Streptomyces lividans TK24, a widely used host for heterologous protein secretion biotechnology. Green and monomeric red fluorescent proteins were fused behind Sec (SPSec) or Tat (SPTat) signal peptides to direct them through the respective export pathway. Significant secretion of fluorescent eGFP and mRFP was observed exclusively through the Tat and Sec pathways, respectively. Plasmid over-expression was compared to a chromosomally integrated Sec-mRFPsp gene to allow monitoring secretion under high and low level synthesis in various media. Fluorimetric detection of SPSec-mRFP recorded folded states, while immuno-staining detected even non-folded topological intermediates. Secretion of SPSec-mRFP is unexpectedly complex, is regulated independently of cell growth phase and is influenced by the growth regime. At low level synthesis, highly efficient secretion occurs until it is turned off and secretory preforms accumulate. At high level synthesis, the secretory pathway overflows and proteins are driven to folding and subsequent degradation. High-level synthesis of heterologous secretory proteins, whether secretion competent or not, has a drastic effect on the endogenous secretome, depending on their secretion efficiency. These findings lay the foundations of dissecting how protein targeting and secretion are regulated by the interplay between the metabolome, secretion factors and stress responses in the S. lividans model.Fluorescent proteins are a major cell biology tool to analyze protein sub-cellular topology. Here we have applied this technology to study protein secretion in the Gram-positive bacterium  Streptomyces lividans, a useful Gram-positive secretion model, can secrete several heterologous polypeptides of bacterial and eukaryotic origin  of Streptomyces venezuelae CBS762.70  has also been used as a reporter  or monomeric RFP (mRFP) proteins behind Tat or Sec signal peptides. To monitor non-folded, non-fluorescent states intracellularly, we used specific antibodies. We show that eGFP and mRFP can be synthesized at high levels from pIJ486 plasmid derivatives and are excellent reporters for the Tat and Sec system, respectively, and as in E. coli (S. lividans Sec system (up to 300 mg/L). mRFP secretion under low expression regimes was studied with the Sec-mRFPsp gene chromosomally integrated in single copies.To dissect the targeting and secretion regulatory mechanism in  E. coli , each maS. lividans model.These tools allowed us to dissect the Sec pathway, both intra- and extracellularly. Sec secretion is controlled in complex ways in various media and it is not a constitutive process. Media optimal for secretion allow secretion without visible cytoplasmic intermediates. In contrast, media that lead to poor secretion, lead to (the) accumulation of secreted proteins in the cytoplasm and may cause a stress response. These data indicate the existence of complex traffic regulation that decides if a molecule will be secreted or not, while secretion overall does not cease. Therefore, it appears that this process is regulated post-transcriptionally in a way that is specific to certain molecules and not to others. Our study lays the foundations for future dissection of these sophisticated regulatory mechanisms in the S. lividans, the genes for the reporter proteins mRFP and eGFP were cloned in a high copy plasmid pIJ486 behind the Tat-dependent signal peptide XlnC and the Sec-dependent signal peptide Vsi  to form pIJ486-Tat-eGFPsp, pIJ486-Tat-mRFPsp, pIJ486-SecV-eGFPsp, and pIJ486-SecV-mRFPsp, respectively. The secretion of these proteins was tested in various media.To develop fluorescent protein reporters for secretion in enhanced green fluorescent protein (eGFP) as a possible reporter protein for S. lividans TK24 secreted in an active form, and in detectable quantities in the culture medium.We first tested the enhanced GFP (eGFP) gene  or the Sec-routed Vsi (SecV-eGFPsp) proteins  gene  behind ts Figure . The resecretion  and veriecretion .Tat-eGFP after 24 h of growth  and peaked after 48 h .To quantify the amount of eGFP corresponding to a certain fluorescence readout, a calibration curve was made by measuring the fluorescence intensity of known amounts of purified recombinant eGFP , while those of TK24ΔTat-eGFPtatC/pIJ486-sp yielded no detectable immuno-stained signal. A very low level of fluorescent signal was present at the later time points of both cultures , but given the delay in appearance it might be due to lysis of the mycelium , when compared to that through the Tat pathway.Collectively, these data showed that eGFP is secreted efficiently by the Tat system, in significant amounts and that its quantification is rapid and simple. These properties make it a good reporter protein for Tat-dependent protein secretion in  E. coli . In cont and cloned it behind the vsi promoter in pIJ486 . Both fusion-encoding genes were ligated into vector pIJ486 and the resulting constructs were transformed into TK24 and TK24ΔtatC cells. Next, all strains were grown in 24-well plates, with continuous shaking and fluorescence measurement in NB medium  . This also demonstrated that there is no detectable cross-talk between the Tat and Sec pathways for this fusion protein. These observations also corroborate previous findings that secretion yields from the TK24 Sec pathway are superior to those from Tat, for most proteins (SP) Figure . The obsproteins .SecV-mRFP secretion could clearly be detected in the culture supernatant of cells growing in NB medium . We then generated calibration curves of both fluorescence measurements and protein mass as for eGFP  grew as well as wild-type TK24 in 250 mL flasks  and which cross-reacted with the mRFP antibody . Similar results were obtained when mRFP was integrated behind the native promoter and signal peptide of the endogenous lsi gene (S. lividans subtilisin inhibitor) SecL-mRFPsp  and TK24/pIJ486-SecV-mRFPsp (hereafter: high SPSec-mRFP levels synthesized).Cell growth in the presence or absence of SecV-mRFP along the secretion pathway and to identify possible cytoplasmic intermediates, we determined the total amount of SPSecV-mRFP synthesized, and determined its distribution in folded cytoplasmic SecV-mRFP synthesized in the “low level” expressing strain is twice that in the “high level” plasmid-bearing strain. Within 24 h of growth >80% of the low, but only ∼50% of the high level, SPSecV-mRFP synthesized in MM is being secreted. In MM medium, mRFP secretion levels off at 24 h, while exponential cell growth continues SecV-mRFP synthesized in MM, secretion of more than 50% is not observed, indicating that the export capacity of the cell is saturated or other mechanisms prevent it.In the high level SP(c)SecV-mRFP synthesized either at low or high level overall in NB medium is secreted to ∼80%, gradually over 48 h and then levels off.the SPSecV-mRFP synthesized (folded secreted+folded cytoplasmic+non-folded cytoplasmic)  Figure .SecV-mRFP is directed for secretion, while at high levels, the secretion system is saturated and almost half the protein is directed for cytoplasmic folding. In the highest performing NB medium, most of the high level SPSecV-mRFP is directed to secretion, but significant amounts are still detected in the cytoplasm. As these would be detected by immuno-staining and do not yield equivalent amounts of fluorescence, most of these polypeptides would be non-folded. This indicated that it is not the synthesized amounts per se that lead to suboptimal secretion but rather the medium context in which this happens. This implied a particular cellular response to some of the nutrients in NB and that these would facilitate targeting and/or secretion. Such nutrients, may become limiting in MM.We concluded that at low levels of synthesis in MM, most SPS. lividans and to identify what are the non-folded intracelullar states we analyzed the intracellular species of SPSecV-mRFP that were detected by α-mRFP antibodies  with a presumed chaperone-like activity protects these molecules through formation of stoichiometric complexes . A species with an apparent mass slightly lower than that of the mature mRFP also accumulated . Also, some new bands in the secretome of the low level SPSecV-mRFP expressing strain appeared . Nevertheless, the secretion profiles were overall very similar. In contrast, the secretome of the high level SPSecV-mRFP-producing strain, in the earlier time points  were very similar with only minor changes (asterisks). In contrast, the secretomes from high-level SPSecV-mRFP synthesizing cells (lanes 7–9), contained not only a strong band of secreted mRFP, but also several other unknown proteins that are secreted at high levels (empty arrowheads). Some of them are likely to be the same polypeptides as those seen in the corresponding MM secretomes  to ∼30-fold (NB) lower than those of SPSecV-mRFP  showed characteristic secretome changes compared to those of TK24 with an empty plasmid (lanes 1–4). The secretome of SPSecV-eGFP-synthesizing cells showed reduced overall amounts of total proteins compared with TK24 harboring an empty plasmid  and became more prominent at late growth points (compare lanes 11 and 12 to lanes 3 and 4). These data suggest that although synthesis of a poorly secreted polypeptide does not affect cellular growth  of SPSec-mRFP were funneled to the Tat pathway  (Streptoverticillium cinnamoneum (118 mg/L)  (S. coelicolor A3(2) (350 mg/L) (S. coelicolor A3(2) (1070 mg/L) (S. lividans. Endo-xylanase from Aspergillus nidulans (19 U/mL)  α and interleukin (IL) secreted in  pathway  and Sec- pathway , xyloglu pathway , endoglu73 mg/L) , phospho18 mg/L)  and two ctively) , while l50 mg/L)  and chit70 mg/L) . The val19 U/mL)  and alph19 U/mL) , have alsecreted .SecV-mRFP in CM medium, revealed a previously unknown 2-step secretion pattern  in Luria Bertani medium, supplemented with ampicillin (50 μg/ml) if applicable. Streptomyces cultures are grown at 27–30°C. Protoplast formation and subsequent transformation of S. lividans TK24 as well as E. coli-Streptomyces conjugations were carried out as described , Minimal Medium (MM) , Minimal Medium with 5 g Bacto casamino acids/L (MMC5), Bennet medium (Ben) , Tryptic soy broth (TSB) , Nutrient Broth (NB) without NaCl , and CM medium (4)2SO4, 10.6 g K2HPO4, 5.3 g NaH2PO4, 1 g MgSO4.7H2O, 1 mL minor elements solution ]. For solid medium, MRYE , was excised from the plasmid pIJ486-spSecV-mRFP using XbaI and HindIII and ligated into the respective sites of pTOS  and the mycelia were harvested by centrifugation  and washed twice with sterilized water. After homogenizing the mycelium in 50 mL of sterilized water, the strains were inoculated into 250-mL Erlenmeyer flasks containing 100 mL of media that were described above. In order to have the same amount of mycelia per each inoculum this equation was used: the volume of inoculum = / OD600 (e medium  suppleme)/ OD600 . The flag for 15 min (SIGMA 3–16 KL refrigerated centrifuge). The bacterial pellets were harvested, resuspended in sterilized water and filtered under vacuum using a 0.2 μm pore size filter . The filter was once more dried (overnight 12–24 h in an oven at 60°C) and weighted for DCW determination.To quantify the dry cell weight (DCW), 10 mL of culture was centrifuged at 3800 × S. lividans preculture and incubated at 29°C while shaking . During growth, fluorescence was measured every 15 min.Use of a micro-bioreactor has been described . Briefly® M200 microplate reader (Tecan) . To compare strain performance, mRFP and eGFP fluorescence intensities, obtained at the transition to stationary phases were identified and evaluated. Error propagation was applied to calculate DCW-specific mRFP and eGFP production.Fluorescence measurements were carried out in an InfinitemRFP and eGFP as His-tagged versions in pIMBB316 and pIMBB643, respectively. Plasmids were transformed in E. coli strain BL21 followed by purification of His-eGFP and His-mRFP by metal affinity chromatography (not shown). Calibration curves of both fluorescence measurements and protein mass were generated using titrations of protein amounts . Precipitation of the proteins in the supernatant was, where applicable, carried out with trichloroacetic acid (TCA) . Proteins were separated by SDS–PAGE and as a standard the Precision Plus ProteinTM Standard (All Blue) from Bio-Rad was used. Proteins were visualized by Coomassie Brilliant Blue (CBB) or by Western blotting and immuno-detection with specific antibodies in combination with a suitable secondary alkaline phosphatase-conjugated antibody (Sigma). eGFP and mRFP antibodies were obtained from Immunosource or generated against lab-purified proteins at Davids Biotechnologie, Germany. Quantification of expression signals on the blot was carried using ImageJ software.Extracellular protein fractions of cultures of TM 300 or a LAS4000 system and analyzed using the ImageQuant TL software to compare the intensity of the protein bands to those of the BSA calibration bands or other control proteins. SDS–PAGE and western blotting is described in the Chemicals were from Sigma. Bacto Soytone from DIFCO Laboratories. DNA enzymes were from New England Biolabs and oligonucleotides from Eurogentec. Images were scanned in an ImageQuantMH and KV performed the experiments, cell growth, and analyzed the data. KV, BB, LM, RN, JaK, and AL constructed the clones. JoK and MO performed the cell fitness analyses. JA analyzed the data. AE and SK conceived, managed and supervised the project, and wrote the paper with contribution from MH. All authors read and approved the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


FcμR is a high-affinity receptor for the Fc portion of human IgM. It participates in B cell activation, cell survival and proliferation, but the full range of its functions remains to be elucidated. The receptor has an extracellular immunoglobulin (Ig)-like domain homologous to those in Fcα/μR and pIgR, but unlike these two other IgM receptors which also bind IgA, FcμR exhibits a binding specificity for only IgM-Fc. Previous studies have suggested that the IgM/FcμR interaction mainly involves the Cμ4 domains with possible contributions from either Cμ3 or Cμ2. To define the binding site more precisely, we generated three recombinant IgM-Fc proteins with specific mutations in the Cμ3 and Cμ4 domains, as well as a construct lacking the Cμ2 domains, and analyzed their interaction with the extracellular Ig-like domain of FcμR using surface plasmon resonance analysis. There is a binding site for FcμR in each IgM heavy chain. Neither the absence of the Cμ2 domains nor the quadruple mutant D340S/Q341G/D342S/T343S (in Cμ3 adjacent to Cμ2) affected FcμR binding, whereas double mutant K361D/D416R (in Cμ3 at the Cμ4 interface) substantially decreased binding, and a single mutation Q510R (in Cμ4) completely abolished FcμR binding. We conclude that glutamine at position 510 in Cμ4 is critical for IgM binding to FcμR. This will facilitate discrimination between the distinct effects of FcμR interactions with soluble IgM and with the IgM BCR.  •FcμR binds with similar affinity to pentameric IgM, IgM-Fc homodimer and IgM-Fc lacking the Cμ2 domains.•Each IgM heavy chain can bind one receptor molecule.•A single mutation in the Cμ4 domain of IgM abrogates receptor binding. Identification of the binding sites for these receptors on immunoglobulin Fc regions is therefore critical for understanding the molecular pathways through which they act.The binding of immunoglobulins to their receptors Until the year 2000, the only known human IgM-Fc receptor was polymeric immunoglobulin receptor (pIgR), which binds both IgM and IgA and is expressed on basolateral surfaces of mucus epithelium and ducts of secretory glands . A seconFcμR is a 390-amino acid (aa) polypeptide consisting of a 17-aa signal peptide and 107-aa Ig-like domain, followed by a further 127-aa extracellular region, a 21-aa transmembrane portion that has a charged histidine residue and a 118-aa cytoplasmic tail. The receptor has no N-linked glycosylation site [The present study focuses on the FcμR binding properties of IgM-Fc. We generated the recombinant extracellular Ig-like domain of human FcμR (sIgFcμR), IgM-Fc with and without the Cμ2 domains, and IgM-Fc with site-directed mutations, for binding analysis by surface plasmon resonance (SPR), to identify the structural determinants of IgM-Fc responsible for FcμR binding.Previous work has shown that the Cμ3 and Cμ4 domains of polymeric IgM are involved not only in binding FcμR , but als22.1600 between 0.6 and 0.8, isopropyl β-D-1-thiogalactopyranoside (IPTG) was added to a final concentration of 1 mM to induce protein expression. The cells were grown overnight at 37 °C and pelleted by centrifugation at 4000g for 20 min at 4 °C. The supernatant was discarded, and the cell pellets were stored at −80 °C.The cloned receptor in Zero Blunt TOPO was kindly provided by Prof. H. Kubagawa. The coding sequence for the extracellular Ig-like domain was cloned into the plasmid expression vector pET24+ and expressed in BL21 (DE3) competent cells at 37 °C under the control of the T7 promoter. The oligonucleotides used were 5′-TGAGATCCGGCTGCTAACAAAG-3′ and 3′-TAAAACAAATTGAAATTCTTCCTCTATATGTA-5′. Cells were cultured in 1 L of ampicillin-supplemented (50 μg/mL) LB broth and grown at 37 °C with orbital shaking at 225 rpm. At an OD2.2g for 15 min at 4 °C. The pellets were washed 5 times in 30 mL of 1 M NaCl, 50 mM Tris-HCL, pH 7.5 and 2% Triton-X. After the final centrifugation, the inclusion bodies were dissolved with solubilization buffer , for 1 h at 37 °C. Insoluble contaminants were pelleted by centrifuging at 14000 rpm for 15 min at 4 °C, and the supernatant containing the protein collected. The sIgFcμR protein was slowly diluted 50-fold into ice-cold refolding buffer , with rapid stirring at 4 °C and left to stand for 72 h. Before purification, NaCl and imidazole were added to the refold solution to a final concentration of 0.5 M and 10 mM, respectively and the pH adjusted to between 7 and 8 using 1 M acetic acid. The solution was filtered with 0.45 μM membrane filter (Millipore) before purification by nickel column chromatography. The purity of the refolded sIgFcμR was analyzed by 18% SDS-PAGE.The cell pellets were thawed on ice and re-suspended in 50 mL lysis buffer ). The cells were then passed through a pre-chilled cell disrupter twice, under a pressure of 1000 psi. The disrupted cells were centrifuged at 160002.3Purified serum IgM was purchased from BBI solutions. The coding sequence for His-tagged IgM-Fc (Fcμ2-4) and Fcμ3-4, lacking the tail piece residues and cloned into pcDNA5/FRT, were generously provided by Dr. Katy Doré. The plasmid containing the Fcμ2-4 sequence was the starting construct for all Fcμ2-4 mutagenesis . Mutations were generated using the Q5 site directed mutagenesis kit (New England Biolabs) and verified by sequencing on both DNA strands. The oligonucleotides used were, for M1 K361D mutation: 5′-CTTCCTCACCGATTCCACCAAGTTG-3′ and 3′-ATGCTGGCAAAGGATGGG-5′; for M1 D416R mutation: 5′-CATCAGCGAGCGAGACTGGAATTCCGGGGAGAGGTTCAC-3′ and 3′- CTGGCCTCACCCACGGCG-5′; for M2 Q510R mutation: 5′-GCCTGAGCCCCGAGCCCCAGGCC-3′ and 3′- ATTGGGGCGCTGGTCACATAC-5′; for M3 mutations D340S, Q341G, D342S and T343S: 5′-TCATCAGCCATCCGGGTCTTCGCC-3′ and 3’-TCCAGAGGGGACACACATGGAGGAC-5′. All primers for site directed mutagenesis were designed using NEBasechanger™ (New England Biolabs).2.45 cells/well and cultured overnight in a 5% CO2 incubator at 37 °C using the transfection medium . The appropriate DNA was co-transfected with expression vector pOG44 (100 ng) using Fugene tranfection reagent (Promega) following the protocol supplied by the manufacturer. After 48 h, the cells were split into 24 well plates in transfection medium containing hygromycin B at a final concentration of 100 μg/mL. Once the cells were confluent, they were trypsinised and seeded into 1 L of Freestyle™ serum-free medium containing 5% penicillin-streptomycin and hygromycin B at a final concentration of 100 μg/mL. The cells were grown in spinners for 3–4 weeks and then harvested. Supernatants were centrifuged at 4000 g for 30 min and then filtered through a 0.45 μM filter (Millipore). 0.1% sodium azide was added and the supernatants were stored at 4 °C. Proteins were first purified on an AKTA Prime system (Amersham) using nickel column chromatography and then on the Gilson HPLC using a Superdex 200, 10/300 column .All IgM proteins were stably transfected into human cells. HEK293 cells were seeded in 96-well plates to 2 × 102.5MALLS studies were performed in-line with SEC on mutant IgM protein samples to assess mono-dispersity and the molecular mass of the protein samples. The peaks corresponding to the homodimeric Fcμ2-4 mutants from SEC were run on the Superdex 200 Increase 5/150 column  using an in-line miniDAWN multi-angle light scattering detector and an Optilab DSP Interferometric Refractometer (Wyatt Technology). The data were analyzed using the ASTRA 4.9 software (Wyatt Technology).2.6Far-UV (190-280) CD spectra were acquired on a Chirascan Plus spectropolarimeter (Applied Photophysics Ltd) flushed continuously with pure evaporated nitrogen throughout the experiments. Measurements were recorded in a 0.5 mm strain-free rectangular cell using a 2 nm spectral bandwidth, 1 nm step-size and 1 s measurement time-per-point. The concentrations of the protein samples used ranged between 0.1 mg/mL - 0.4 mg/mL and buffer used was 10 mM HEPES and 150 mM NaCl at pH 7.4. All spectra were acquired at 25 °C and buffer baseline corrected before analysis.2.7D values represent the fitting of the 1:1 binding model to the experimental data.All SPR assays were performed on a Biacore T200 instrument  at 25 °C in running buffer . Serum IgM, IgM-Fc, Fcμ3-4 and IgM-Fc (Fcμ2-4) mutants were immobilized on Biacore CM5 chips in 10 mM sodium acetate, pH 4.5, using standard amine coupling chemistry. The analyte was sIgFcμR in concentrations ranging from 0.25 μM to 12 μM. In each experiment, the binding curves were corrected by subtracting the signal obtained from the control flow cell. Non-specific binding was <30%. Steady state binding curves were analyzed using Biacore T200 evaluation software. The errors in the KTo investigate the stoichiometry of sIgFcμR binding to IgM or its homodimeric fragments, the following equation was used to calculate the activity of the immobilized IgM or fragments :max of sIgFcμR bound)/ (Mol. Wt. of sIgFcμR × RU of IgM or homodimeric fragment immobilized).% Activity = 100 × (Mol. Wt. of IgM or homodimeric fragment × R3c (molecular weight 12.4 kDa). The far-UV CD spectrum showed a characteristic β-sheet signature of a folded polypeptide with a broad minimum near 218 nm  was expressed as described in Material and Methods. After affinity purification, SEC was conducted to ensure the purity of the protein A. Fractir 218 nm B. SPR anectively . This shmax  to the number of RU immobilized on the sensor chip, taking into account the relative molecular weights , indicated that each IgM heavy chain has a binding site for sIgFcμR. The number of RU immobilized for IgM, IgM-Fc and Fcμ3-4 was 1959, 876 and 1343 respectively, and the sIgFcμR Rmax values were 229, 253 and 472. If the stoichiometry of binding is assumed to be one sIgFcμR to one IgM-Fc homodimer, then the apparent binding activities are 165%, 171% and 139% respectively ; since no immobilized protein can have an activity of >100% (and it is commonly much less), this indicates that each heavy chain must be able to bind one sIgFcμR molecule (i.e. 2:1 sIgFcμR:IgM-Fc homodimer).SPR has limitaitons as a method to determine the absolute stoichiometry between, in this case, an antibody or fragment and its receptor, due to inevitable loss of activity caused by chemical coupling of the antibody or fragment to the matrix. However, since the initial baseline signal in RU is proportional to the mass of antibody or fragment immobilizsed, and the additional signal in RU is proportional to the mass of receptor bound, it is possible to calculate the theoretical maximum expected signal assuimg a particular stoichiometry of binding. For each of the proteins, IgM, IgM-Fc and Fcμ3-4, the ratio of RSite-directed mutagenesis was applied to study IgM-Fc binding to sIgFcμR using SPR. While there is currently no crystal structure for human IgM-Fc or any of its domains, assumptions about their structural features can be made based on the individual domain structures of mouse IgM-Fc  and the The IgM-Fc mutants were secreted from human embryonic kidney (HEK) cells. After purification by affinity chromatography followed by SEC (data not shown), the proteins were characterized by SEC-MALLS and far-UV CD. Similarly, non-mutated IgM-Fc (wild-type) was prepared to act as a positive control in the experiments. For all four preparations, SEC-MALLS analysis revealed single peaks with a weight-averaged molar mass between 80 and 84 kDa, within experimental error of one another and of IgM-Fc homodimer, proving that the samples were not aggregated .Fig. 4ChThe far-UV CD profiles of all the three IgM-Fc mutants compared well with that of wild-type, suggesting that the native states of the mutants were not perturbed by the respective mutations and that all were properly folded .Fig. 5ChD values for wild-type and the quadruple mutant are shown in SPR analysis was used to assess the binding of sIgFcμR to immobilized wild-type, single- (Q510R), double- (K361D/D416R) and quadruple- (D340S/Q341G/D342S/T343S) mutant IgM-Fc. The sensorgrams for the four IgM-Fc species and the resulting KD = 1.45 ± 0.03 μM) almost indistinguishable within experimental error from the wild-type (KD = 1.83 ± 0.09 μM). This shows that residues D340, Q341, D342 and T343, which correspond to key elements of the IgE-Fc/FcεRIα and homologous IgG-Fc/FcγR interactions, play no role in the binding of IgM-Fc to FcμR. The double mutation of residues 361 and 416 in Cμ3 (M1) substantially reduced sIgFcμR binding; the location of K361 and D416 at the Cμ3/Cμ4 interface corresponds to that of the CD23 and FcαR binding sites in IgE and IgA respectively. . Further investigation with single mutations would be required to determine the contribution of each. Finally, mutation of residue 510 in Cμ4 (M2) showed no binding of sIgFcμR at all concentrations tested, indicating that Q510 plays a critical role in the IgM-Fc/FcμR interaction.The mutation of residues 340–343 in Cμ3 generated an IgM-Fc mutant (M3) with sIgFcμR binding parameters  within the Cμ3 and Cμ4 domains since an Fcμeraction ,17. A laeraction . In ordeD ~ 1–2.5 μΜ was determined by SPR. This is ~ 100 fold weaker than cell surface FcμR binding to pentameric serum IgM, which is reported as KD ~ 10 nM [i.e. an avidity effect).The binding of purified sIgFcμR to whole serum IgM (largely pentameric), IgM-Fc (Fcμ2-4), and the Fc sub-fragment Fcμ3-4, was investigated by SPR . It is c ~ 10 nM ; the difIn order to determine the FcμR binding site on IgM, we generated three IgM-Fc mutants, M1, M2 and M3  and studLloyd et al. proposed a model of the interaction between IgM-Fc and FcμR, based upon molecular dynamics simulations, that involved several residues from Cμ4, but not Q510, together with a contribution from Cμ2 . Their eIn cryoEM structures and models of pentameric and hexameric IgM , the Q51e.g. FDCs and B cells in germinal centres or epithelial cells and B cells) and could lead to competition for IgM. However, the Cμ4 domain has also been identified as the binding site for the erythrocyte membrane protein pfEMP1 expressed by cells infected with Plasmodium falciparum [The functional significance of this clustering of IgM receptor binding sites in Cμ4 is unclear, although some different cell types which express these receptors do co-localise (lciparum , and intlciparum .5There have been contradictory findings in relation to the function of FcμR . The ideR.A.C. and B.J.S. designed the research. R.A.C. and R.A.N. performed and analyzed the experiments. All authors contributed to writing the paper.Rosemary Nyamboya: Formal Analysis, Investigation, Visualization, Writing-Original Draft, Writing-Review and Editing.Brian Sutton: Conceptualization, Funding, Project Administration, Writing-Review and Editing.Rosaleen Calvert: Conceptualization, Formal Analysis, Investigation, Project Administration, Supervision, Validation, Visualization, Writing-Review and Editing.The authors declare that they have no conflicts of interest with the contents of this article.


The nature of immunoglobulin M (IgM) nephropathy has been controversial for a long time, but it is now considered an independent disease like immunoglobulin A nephropathy. IgM nephropathy has been known to have various clinical manifestations ranging from asymptomatic hematuria and/or proteinuria to nephrotic syndrome. Recently, one case of IgM nephropathy manifesting as crescentic glomerulonephritis (GN) was reported in a child.We experienced a case of IgM nephropathy that manifested clinically as nephritic and nephrotic syndrome with pathologically confirmed crescentic GN in a 30-year-old woman. We administered a calcineurin inhibitor and corticosteroids to treat the ongoing nephrotic syndrome after remission of crescentic GN. As a result, her proteinuria was significantly reduced and edema improved.We described a case of IgM nephropathy in an adult patient who initially developed crescentic GN with nephritic and nephrotic syndrome. This case report could contribute to a deeper understanding of IgM nephropathy. Based on two seminal reports in the 1970s , immunogHere, we describe the case of a 30-year-old woman diagnosed with IgM nephropathy that manifested as acute nephritic and nephrotic syndrome with pathologically confirmed crescentic GN.3/μL; total protein level, 5.1 g/dL; serum albumin level, 2.01 g/dL; serum creatinine level, 1.14 mg/dL (corresponding to an estimated glomerular filtration rate [eGFR] of 64.9 mL/min/1.73 m2); and total cholesterol level, 395 mg/dL. Urinalysis revealed proteinuria (2+) and hematuria . The spot urine protein-to-creatinine ratio (UPCR) was 7.32 g/g, and the spot urine albumin-to-creatinine ratio was 5.24 g/g, with nonselective glomerular proteinuria detected by urine electrophoresis. The results of other serological tests were positive for the antinuclear antibody (1:160), and negative for a speckled pattern and anti-neutrophil cytoplasmic antibody (ANCA); additionally, her serum complements were within their reference ranges. All viral serological markers were negative. Kidney biopsy was performed. Twenty of 30 glomeruli showed cellular or fibrocellular crescents with mesangial proliferation. There was focal mild to moderate acute and chronic inflammatory cell infiltration with mild interstitial fibrosis and tubular atrophy, mainly along the crescentic glomeruli. There was occasional fibrinoid necrosis in the crescentic glomeruli, but there was no vasculitis in the interstitium. Some glomeruli showing mesangial proliferation or segmental sclerotic change without fibrinoid necrosis were observed. The immunofluorescent study revealed diffuse global immunofluorescent activity for IgM (1+), immunoglobulin G (IgG) (trace), and complement 3 (C3) (trace) in the mesangium. The corresponding electron-dense deposit was confirmed by the electron microscopic (EM) examination . Her serum creatinine level continuously increased to 1.66 mg/dL  after admission and then decreased to 1.09 mg/dL  when she was discharged on the eighth day of admission. Three months later, her serum creatinine level was stabilized at 1.11 mg/dL , and microscopic hematuria completely disappeared. However, she steadily complained of generalized edema, and considerable nephrotic-range proteinuria persisted at a spot urine UPCR of 7.73 g/g. Given the tendency of resistance to cyclophosphamide treatments, rituximab was administered intravenously (375 mg/m2 per week) for 4 weeks alternatively. After the first dose of rituximab, her UPCR decreased to 0.27 g/g. However, after 4 weeks of rituximab therapy, the UPCR increased again to 5.59 g/g.As a treatment regimen for crescentic GN, three pulses of intravenous methylprednisolone (750 mg/day) were administered, followed by daily oral prednisone and intravenous cyclophosphamide (500 mg/m2) and spot urine UPCR was 1.49 g/g. Owing to the risk of nephrotoxicity, we replaced cyclosporine with tacrolimus. At the recent follow-up, renal function and proteinuria were stabilized with a serum creatinine level of 1.23 mg/dl  and UPCR of 0.88 g/g, without symptoms of edema , IgG (trace), and C3 (trace) were observed, with electron-dense deposits in the mesangium, like in the first biopsy Fig. . A diagnThis is the first case report of an adult patient who was diagnosed with IgM nephropathy manifesting as crescentic GN and nephrotic syndrome. The pathological findings of IgM nephropathy have been diversely reported to date, from almost normal histological findings such as MCD to findings such as FSGS. Connor et al.  recentlyRegarding the clinical manifestation of IgM nephropathy, subnephrotic proteinuria and nephrotic syndrome were commonly observed in the studies involving adults, but no patient presenting with crescentic GN has been reported –9. AlthoOur patient was treated initially with pulse methylprednisolone followed by daily oral prednisone and intravenous cyclophosphamide as empirical therapy for idiopathic crescentic GN. After treatment, renal function was stabilized, but nephrotic-range proteinuria continued. On the basis of reports on the therapeutic effect of rituximab in IgM nephropathy , 17, we In addition to the previous case report involving a girl in Pakistan, this case report provides evidence of IgM nephropathy manifesting as crescentic GN, particularly in an adult. This case report will help expand physicians’ knowledge on the clinical and pathological spectrum of IgM nephropathy. For a better understanding of IgM nephropathy, more evidence is needed from studies with a larger number of patients and longitudinal research designs.


Given the benefits of physical activity for health and survival, clinicians are seeking opportunities for cancer patients to become more active independent of rehabilitation programs that are small, time-limited, and location specific. This proof-of-concept study evaluated a community-based physical activity program (Curves™) for increasing physical activity among women diagnosed and treated for breast cancer.n = 14) received the community physical activity memberships (Curves™), guidelines, and a pedometer. This group was compared to women (n = 16) who received physical activity guidelines and a pedometer on changes in physical activity. In study 2, women (n = 66) completed self-report questionnaires after Curves™ memberships expired to evaluate the program. Study 3 was a qualitative study exploring the benefits and barriers of the physical activity program among women (n = 6) who attended Curves™ regularly.Women were recruited from a breast cancer clinic through physician chart review. In study 1, women (Provision of memberships to a community-based physical activity program did not improve physical activity levels beyond educational and information resources. However, there are a number of advantages to community-based physical activity programs, and the women offer a number of suggestions for improvements for community physical activity opportunities aimed at breast cancer survivors.Women-only community-based physical activity programs may be a viable option to help introduce women to get active after treatment.Trial registration ISRCTN, ISRCTN14747810. Registered on 18 October 2017—Retrospectively registered, 10.1186/ISRCTN14747810 Breast cancer is the most common cancer diagnosis among Canadian women . With 5-The benefits of physical activity (PA) for breast cancer survivors (BCS) are well-documented , 4. Evidcancer PA programs and there is a need to identify and evaluate women-specific PA opportunities offered in the community.Community-based PA opportunities may be ideal for BCS , 15. ComCurves™ is a women-only circuit training facility with numerous locations across North America. This program may be particularly attractive to BCS as it is women-only and offers PA coaching support that is not in the context of cancer care . This PAThe purpose of this current proof-of-concept research was to evaluate Curves™ as a PA program for BCS. This purpose is addressed with three interrelated studies focused on a sample of BCS who were eligible for Curves™ memberships from a local breast cancer clinic in a large urban city in Canada. Specifically, 203 women accepted complimentary memberships to the PA program over a 1-year period and served as the sampling frame for the three studies in the current program of research. All participants across the three studies were mutually exclusive . Generally, participants were eligible for the studies if they (a) had been diagnosed with breast cancer; (b) were ≥ 18 years of age; and (c) could read and understand English or French. The flow of participants among all three studies is depicted in Fig. Using a proof-of-concept paradigm , the purExercise Guidelines for People with Cancer were not yet published [All participants were screened by the head surgical oncologist using chart review and informed of the study in person at a regularly scheduled medical appointment. Participants were randomly assigned to the community PA program (Curves™) or a lifestyle PA program. Participation in the Curves™ group involved accepting a complimentary membership in addition to general PA Guidelines , a canceublished . Women wParticipants in the lifestyle PA group were given all lifestyle intervention materials without a complimentary membership. These women were asked to follow the recommendations outlined in the educational and guideline materials, which included striving towards 150 min of moderate-to-vigorous PA per week . All pard = 0.37 and 0.38 between standard of care and either pedometer or combined print material and pedometer intervention groups. Using G*Power repeated measured analysis of variance F-test family with alpha = 0.05 and power of 0.08, 2 groups with 2 assessments, and a correlation among repeated measures estimated at 0.5, a total of N = 58 BCS were required for recruitment.Sample size was calculated based on the findings from Vallance et al.  who repoSocio-demographic variables included age; ethnicity; highest education achieved, and marital status. Cancer-related variables included stage of cancer diagnosis, time since diagnosis and treatment, and types of cancer treatment received . Finally, body mass index (BMI) was calculated from self-reported weight and height.Women self-reported PA using Short Questionnaire to Assess Health-Enhancing PA (SQUASH)  which isWomen were also given a pedometer  to wear each day from time awakening until bedtime except for during water activities and showering/bathing. A logbook was also provided to keep track of daily steps. Average number of steps per week was computed. For pedometer assessment to align with main analyses, average steps for week 1 (baseline) and week 12 were used in analyses.d effect sizes were also calculated and reported.Descriptive statistics (means and standard deviations or frequencies) were calculated. To address the main purpose, a repeated measures univariate analysis of variance (RM ANOVA) model was estimated to assess differences in the PA program and lifestyle groups on PA over time. Where appropriate, Cohen’s N = 30; n = 14 Curves™ and n = 16 lifestyle PA) can be found in Table p < 0.05) different on any measured descriptive variable. Means and standard deviations for PA levels by groups over the 12 weeks are presented in Table F = 0.80, p = 0.38; Group F = 0.91, p = 0.35, or interaction, F = 0.10, p = 0.92.A total of 38 BCS were recruited and consented to the study, although 8 women dropped out of the study before baseline assessment. A summary of demographic characteristics of the analytical sample of participants  = 0.67, p = 0.24, group effect, F = 0.31, p = 0.59, or time × group interaction, F = 0.93, p = 0.35.For pedometer data, seven women from the Curves™ group and 12 women from lifestyle group complied with wearing the pedometer . These 19 women provided 1593 days of pedometer data and median days worn per week was seven. Means and standard deviations for the average step count over the week of assessment for the pedometer data for those who wore the device are reported in Table Overall, there were no differences in the groups based on PA participation. In this proof-of-concept study, the lack of differences may be attributed to a small sample size, additional sociocultural factors that were not measured in this study that may have influenced the groups unequally, or ineffective memberships to Curves. Understanding participation trends and experiences in the community-based program is important to better explain these findings.Given the findings of the first study, the purpose of study 2 was to explore attendance at the Curves™ PA program, identify potential personal and cancer-specific factors associated with attendance, and describe the barriers and facilitators to participating in the program.One year following the provision of complimentary program memberships, 173 BCS who were not involved in study 1 were invited to participate in study 2. Sixty-six BCS (response rate of 38.2%) provided informed consent and completed a mailed questionnaire 1 year following the provision of their Curves™ membership. Questionnaires were completed and returned by mail.Sociodemographic and cancer history variables were assessed as described in study one.Frequency of program attendance was assessed to examine use of memberships . Additionally, participants were asked to describe their program participation at . Curves™ PA engagement was dichotomized as either no or little participation (= 0) compared to regular participation (= 1) to further explore predictors of behavior.extent of met needs  rated as 1 = met none of my needs to 4 = met all of my needs, satisfaction with Curves™ service  with responses of 1 = very dissatisfied to 4 = very satisfied, and likelihood of choosing Curves™ in the future  rated as 1 = definitely not to 4 = definitely yes. Specifics of Curves™ location  were also examined.A questionnaire was developed to evaluate participants’ experiences at the Curves™ program. Questions were modeled from common theories of behavior change including Theory of Planned Behavior  and ExpeWomen were asked to list three things they liked about the program and three things they disliked about the PA program in an open-ended question which were then coded based on common barriers for PA , 42 inclt-test and Chi square analyses. Open-ended responses were coded by two independent researchers and categorized into barriers and facilitators.Descriptive statistics were used to characterise the sample based on sociodemographic information, cancer history and self-report PA. In the main analysis, descriptive statistics, specifically means, standard deviations and frequencies (%) were used to characterize the women’s participation (attendance) and evaluation of the Curves™ PA program. Differences in personal or cancer-related factors among women who regularly participated in the Curves™ program compared to those women who did not participate were tested using n = 5), injury precluding exercise (n = 1), and moved or the Curves™ gym in neighbourhood closed (n = 4). Reasons for drop-out, in which women could report as many as they wanted, included: poor perception of the program characteristics including loud music, preference for other activities, no health benefits attained, and distance to a PA program location.Participant characteristics are summarized in Table Women who attended the program were generally satisfied with their experience, reported that the program met their needs, and reported that they would choose Curves™ again if they were going to exercise at a fitness facility. However, none of the women had purchased a Curves™ membership at the time of completing the questionnaire. There were no differences between groups of women who attended Curves™ compared to those who did not on any personal or cancer-related factors.Based on a qualitative assessment of open-ended responses to the list of facilitators and barriers pertaining to the program, there were 110 reported favourable characteristics  and 83 barriers  of the program. See Table There were a number of barriers and facilitators to program participation that may be used to inform future program development for community PA opportunities among women with breast cancer. Offering variety in programs and flexibility in participation, such as the “drop in” and short circuit nature of the program, appear to be critical to favourable perceptions. This finding is in line with recent empirical evidence on the importance of perceptions of variety in PA , 44. TheStudy 3 was a qualitative study among a unique sample of women who attended Curves™ and who were purposefully selected to participate in individual interviews aimed at understanding the experience of participating in the Curves™ PA program.Women were purposefully sampled if they had attended Curves™ at least once a month for the duration of the membership and that they had been diagnosed with breast cancer within 5 years prior to study Fig. . Women wAudio files were transcribed verbatim and transcripts were analyzed using inductive thematic analysis . TranscrN = 6) were an average of 55 years old, identified as Caucasian (100%), mostly (67%) university educated and married (50%). They were all breast cancer survivors (100%) who had completed treatment an average of 3.7 (SD = 1.1) prior to the study commencing. Treatment included lumpectomy (67%), single or double mastectomy (33%), reconstructive surgery (33%), chemotherapy (50%), radiotherapy (83%) and hormonal therapy (67%). Participants reported that they attended Curves™ on a regular basis ranging from once per week to multiple times per week. Most of the women (n = 5) attended the PA program for almost a full year; however, none renewed their program membership. Two main themes and six sub-themes were identified generally around the motivation and barriers of the PA program  workout atmosphere, (ii) goal achievement, and (iii) social influences.Some women (33%) felt that the atmosphere created by the program was inspiring and made them feel like moving. The circuit was described as motivational based on program duration, intensity being personal and challenging yet manageable, ease of getting on machines, variety of machines, and music.Women mentioned weight management as a program benefit. Most (83%) also discussed the program helped them meet goals related to feeling better, finding an enjoyable activity and helped them re-gain a sense of normality.A majority (67%) of women expressed social outcomes related to attending the program. Women attending the program felt that the staff, few other BCS, and other women attending the program were supportive. Specifically, BCS felt that trainers hired at the location she attended were helpful and inspirational. They appreciated that the staff focused on them as women and not as BCS.The negative or challenges to PA that were experienced or perceived at Curves™ were coded into the main theme of barriers, and there were three sub-themes: (i) location and scheduling; (ii) circuit design, and (iii) lack of cancer-specific support.Lack of locations near participants’ homes was an issue. Similarly, circuit schedules and seasonal schedules were perceived as inconvenient by participants.Two women mentioned the circuit itself did not meet their exercise intensity needs. This may have been a reason for why some women did not renew their memberships. Furthermore, half of the women mentioned a lack of program variety. While these were program challenges, it was also apparent that the program was a platform that led to other PA opportunities.Improvements centred around specific types of social support. For instance, having specific, one-on-one support from staff members would be helpful for fostering attendance. Similarly, one participant felt that she lacked informational support from the program. Based on BCS’ perceptions and experiences, it was clear that further tailored social support was valuable for their interest and enjoyment.BCS who attended Curves™ experienced a number of benefits and reported a number of factors that influenced their decision to discontinue. These factors were centered on specific program logistics that could be altered for improved sustainability. Specifically, community PA programs should be accessible and flexible in hours of operation, should hire knowledgeable staff specific to breast cancer  and PA, and should offer a variety of PA plans in both number of exercises as well as intensity. Based on the strengths of the program, the duration of the activity was highly valued . The women-only focus and motivated staff were also important. These factors overall are not surprising given the plethora of research identifying lack of perceived social support, lack of facilities or facilities that are close by and lack of motivation as barriers to engaging in PA for both BCS and other cancer survivors , 41, 46.The overall purpose of this research was to evaluate Curves™ as a community-based PA program for BCS. This was accomplished through three studies with independent samples, which independently aimed to (a) compare two ecologically valid programs for changes in PA levels in BCS (study 1); (b) evaluate attendance, attitudes and beliefs about characteristics of the Curves™ program (study 2); and (c) explore experiences of BCS who attended Curves™ through interviews (study 3). Based on results of study 1, Curves™ did not significantly increase PA levels in BCS who received free, 1-year memberships. Findings from studies two and three suggest that BCS who attended the program were not interested in the program for long. While a few women discussed ease of the program, there was no adaptation over time that limited interest once women improved. Nonetheless, it may be that similar community-based PA programs may be a promising opportunity for initiating PA among BCS who can then gain confidence and skill to advance to other physical activities. The women-only focus, time commitment of 30 min, simple circuit, and encouraging environment are all positive features that could be adopted in survivorship PA programs that may also be transition opportunities for BCS to gain competence, social support, and a sense of personal agency and control. In this way, sustainability of the PA program may not be a primary goal—the program can be effective at fostering a sense of normalcy among BCS and building a foundation for PA in a community setting outside of survivorship programming.While current findings confirm there are many barriers to PA which developed from the Curves™ environment, these barriers are consistent with situational contexts of community programs more generally, including distance to closest location, hours of operation and staff . FurtherIn contrast to previous findings, women did not discuss other common environmental or situational barriers such as lack of time, inclement weather and absence of equipment  or diseaThis work is not without its limitations. Specifically, the sample may not have been representative of BCS from this large urban centre in that they were predominately Caucasian, affluent and well-educated. The sample size for study 1 was small, and many of the women had already received memberships thus precluding random assignment to intervention groups. There are also a number of additional sociocultural factors and conditions reflecting intersections of race, identity, value systems, health perceptions, and socioeconomic position that were not assessed in study 1 that may have precluded finding differences in the intervention groups. The response rate for study 2 was low, although consistent with self-report survey research . AdditioBased on strengths and weaknesses of the PA program circuit, survivorship programs can benefit from integrating women-only, supportive, small class size, short time commitment, self-paced circuit training PA features while also highlighting trained staff in oncology and fitness, adding variety and challenge, and regular adaptations based on training gains. Future interventions targeting BCS should account for these features. It is important to note that the PA program and other similar community-based programs may be a gateway opportunity to help women initiate PA in a safe and comfortable environment. Understanding the importance of community PA programs is essential for translating research into practice  and help


Text-based programs have been shown to effectively address a wide variety of health issues. Although little research examines short message service (SMS) text messaging program characteristics that predict participant retention and attrition, features of SMS text message programs, such as program duration and intensity, message content, and the participants’ context, may have an impact. The impact of stop messages—messages with instructions for how to drop out of an SMS text message program—may be particularly important to investigate.The aim of this study was to describe attrition from Text2BHealthy, a text-based nutrition and physical activity promotion program for parents of low-income elementary school children, and to determine the impact of message content and number of stop messages received on attrition.Using data from 972 parents enrolled in Text2BHealthy, we created Kaplan-Meier curves to estimate differences in program duration for different SMS text message types, including nutrition, physical activity, stop, and other messages. Covariates, including rurality and number of stop messages received, were included.P<.001). Furthermore, each additional stop message received increased the probability of attrition . The degree of rurality also had a significant effect on the probability of attrition, with metropolitan county participants more likely to drop out of the program than rural county participants. The interaction between SMS text message type and total number of stop messages received had a significant effect on attrition, with the effect of the number of stop messages received dependent on the SMS text message type.Retention rates by school ranged from 74% (60/81) to 95.0% (132/139), with an average retention rate of 85.7% (833/972) across all schools. Program duration ranged from 7 to 282 days, with a median program duration of 233 days and an average program duration of 211.7 days. Among those who dropped out, program duration ranged from 7 to 247 days, with a median program duration of 102.5 days. Receiving a stop message increased the probability of attrition compared with receiving messages about nutrition, physical activity, or other topics (hazard ratio=51.5, 95% CI 32.46-81.7; This study demonstrates the potential of SMS text message programs to retain participants over time. Furthermore, this study suggests that the probability of attrition increases substantially when participants receive messages with instructions for dropping out of the program. Program planners should carefully consider the impact of stop messages and other program content and characteristics on program retention. Additional research is needed to identify participant, programmatic, and contextual predictors of program duration and to explicate the relationship between program duration and program efficacy. Using text messages, also known as short message service (SMS), to deliver or supplement health interventions has increased in popularity in recent years. SMS text messages are an appealing mode of program delivery largely because they can be easily and inexpensively used to reach a broad audience . In the d=0.329 , indicating a small effect across studies [Text-based programs have been successfully used to address a wide variety of health issues such as weight loss , smokingAlthough many studies have shown that retention is generally high in SMS text message programs ,10-12, rUnderstanding when and why participants tend to drop out of SMS text message programs is imperative for effective program planning. There is some evidence that when participants drop out, they do so early in the program; Coa and Patrick found that among those who dropped out of a diet and physical activity SMS text message program, 65%  did so within the first 2 weeks . As SMS stop messages. Stop messages provide participants with instructions for withdrawing from the program. Due to a concern of the program funding agency about participants without SMS text messaging in their data plans incurring costs to receive program messages, stop messages were a required feature of the program. It is unknown whether and how much such messages increase attrition. In addition, the Text2BHealthy program was tailored to particular elementary schools located throughout the state of Maryland. These schools varied widely in terms of the degree of rurality of the area. Participants in rural areas might be less likely to drop out of a text-based health promotion program as rural areas tend to have fewer health services and programs available [The purpose of this study was to describe attrition from Text2BHealthy, an SMS text message-based healthy eating and physical activity promotion program. Due to the lack of existing research about attrition from SMS text message programs, certain features of Text2BHealthy thought to be most likely to influence attrition were selected for examination. For example, in the program, several types of message content were delivered, including messages about nutrition, physical activity, and vailable .In this study, we sought to discern (1) how long participants remained in the program before dropping out; (2) whether particular types of messages, particularly stop messages, increased the likelihood that participants will drop out of the program; (3) whether the number of stop messages received increased the likelihood that participants will drop out of the program; (4) whether school rurality was associated with attrition; and (5) whether the impact of the number of stop messages received differs by message type.stop to the program short code or responding to any message with a message that included stop. As required by the program funding agency, SMS text messages informing participants about how to leave the program were sent approximately every 6 weeks.Text2BHealthy is a Maryland Supplemental Nutrition Assistance Program-Education (SNAP-Ed) nutrition and physical activity promotion program delivered by SMS text messages to parents of elementary school children . ParentsParticipants included in this study were 972 parents of children attending a selection of low-income elementary schools with youth SNAP-Ed programs in Maryland and participating in the Text2BHealthy program. Parents were recruited through school events and program promotional items sent home at the beginning of the school year. They were enrolled by either providing their phone number to program staff or enrolling themselves through a keyword texted to a short code. Data for this study came from the September 2012 to June 2013 program year, which includes enrolled parents from 10 elementary schools in 5 Maryland counties and Baltimore City. Parents could enroll in and drop out of Text2BHealthy at any point during the program year.The outcome of interest for this study was attrition from the Text2BHealthy program . Attrition data were recorded by the Web platform that was used to send messages to participants. When participants sent an SMS text message to the program phone number indicating that they wanted to be removed from the program, the Web platform would automatically and instantly remove them from the list of participants to receive SMS text messages and record the date and time that they left the program. Program enrollment date and dropout date were used to calculate program duration  in days.Survival analyses were conducted to examine differences in participant attrition from the program. The primary predictor for survival time, in days, was message type. Message type was created by coding the last SMS text message received (either the last message before dropout or the last message sent during the program year) into 5 categories: nutrition; physical activity; stop messages describing how to drop out of the program; stop messages combined with content about nutrition or physical activity; and a variety of other content that included evaluation questions, goal-setting, and healthy event notifications and other messages not explicitly addressing nutrition and physical activity actions see .Several covariates were included in the models, including rurality and the number of stop messages received during the program year. Rurality was determined for each school using the United States Department of Agriculture’s 2013 Rural-Urban Continuum Codes  for the county where the school was located. These codes range from 1 to 9, with 1 to 3 indicating metropolitan areas and 4 to 9 indicating nonmetropolitan areas .Frequencies were used to determine retention rates. Mean and median program durations were calculated to determine how long participants remained in the program before dropping out. Kaplan-Meier curves were created to estimate differences in participant attrition for different message types. Additional models that included total number of stop messages received, RUCC, and interactions between message type and total number of stop messages received were run. Using the survival package, Cox proportional hazards models were fit to estimate hazard ratios. A graphical inspection of the residuals was done to test the proportional hazards assumption. Pairwise comparisons were conducted to determine the effect of the number of stop messages by message type. All analyses were run in R Statistical Computing Package .There were 972 participants during the 2012 to 2013 program year. Among the 10 participating schools, the average free and reduced meals (FARM) rate, indicating the percentage of students in the school receiving free or reduced price meals, was 80.9%. According to the 2013 RUCC , 7 out o23=916.6; P<.001). Among those who did drop out of the program, participants were more likely to drop out after having received a stop message, followed by a stop message paired with nutrition or physical activity content and then by physical activity content alone. Participants were least likely to drop out after receiving a nutrition message. On the basis of these findings, Cox proportional hazards models were fit to estimate the effect of several covariates on program attrition.Kaplan-Meier survival curves indicate differences in program attrition by SMS text message type see . A log rP<.001). The addition of covariates to the model attenuates this relationship; however, the magnitude of the effect of stop messages remains substantial in comparison with nutrition messages. In model 2, which examines SMS text message type and the total number of stop messages received, the hazard ratio decreases substantially to 10.36 . In addition, model 2 shows that the more stop messages a participant receives, the greater the probability of attrition. In model 3, rurality of the program site impacts attrition, as the addition of the RUCC increases the hazard ratio to 11.60 .Model 1 examines the predictive effect of SMS text message type on program attrition see . As seenP<.001), whereas living in a nonmetro county with a population between 2500 and 19,999 people (RUCC=6) is associated with lower, but statistically insignificant probability of dropping out of the program , respectively, compared with living in a metro county with populations greater than 1 million (RUCC=1).Living in a metro county with populations under 250,000 (RUCC=3) is associated with higher probability of dropping out of the program . The interaction between stop messages and the total number of stop messages received, however, has the largest effect on likelihood of dropping out of the program , suggesting a possible moderating effect of the total number of stop messages on the association between receiving stop messages and attrition rate in the program.In model 4, the final model in this analysis, interaction terms between the total number of stop messages and SMS text message types are included. The effect of the interaction term was explored through pairwise comparisons by message type (see Pairwise Comparisons by Message Type section). The addition of all the covariates and the interaction terms to the model leads to a 99% decrease in the effect of stop message only from model 1, and the association is no longer statistically significant  were retained through the end of the school year, indicating possible exposure to key messages over a lengthy period. This is consistent with other literature, including 2 meta-analyses that found a mean retention rate of 86% 72 were r across aWe also found that the median program duration for participants who dropped out of the program was high (102.5 days), meaning that many participants were exposed to a substantial amount of program content despite ultimately unenrolling from the program. This observation is inconsistent with research conducted by Coa and Patrick , showingIn examining the context of program participants, we found that parents living in rural counties were less likely to drop out than parents living in more metropolitan counties. This finding may be explained by the relative scarcity of health services and programs in rural areas. Participants in rural areas may be less likely to drop out of a text-based health promotion program, as rural areas tend to have fewer health services and programs available . It is lWe found that the effect of the number of stop messages received during the program period differs by SMS text message type. In particular, receiving 6 stop messages results in greater probabilities of attrition for all SMS text message types than receiving 3 stop messages, suggesting a possible dose-response relationship between number of stop messages received and the likelihood of dropping out of the program. In addition, the interaction between stop messages and the total number of stop messages received yields the largest effect on attrition compared with the interaction between other message types and the number of stop messages received. Although we were unable to find any other research examining the effect of stop message receipt on attrition, these findings might echo dose-response observations in another study showing that number of messages received was associated with positive behavior changes related to weight management . These fProgram planners needing to send instructions on how to drop out of a program should consider limiting the number of times this information is provided, as it is possible that a greater frequency of such information has negative implications for program retention. In addition, a better understanding of the ways in which participants’ characteristics impact probability of dropping out after receiving a stop message could also improve retention. In particular, certain participants might be more responsive to stop messages, such as those who join many text-based programs and those with limited facility with SMS text messaging who might drop out accidentally or misunderstand the intention of the stop message. Furthermore, future research may identify the best ways of informing participants about how to remove themselves from the program and illuminate which groups of participants may be expected to already know how to remove themselves from any SMS text message program.This study has a number of important limitations. First, we were unable to control for individual demographic characteristics that might explain differences in attrition. Second, although the data analysis accounts for different frequencies with which particular messages were sent, the relatively small number of messages combining nutrition content with a stop message and physical activity with a stop message signifies a small number of possible dropout events to observe, resulting in limited or inadequate statistical power. We, therefore, combined these 2 SMS text message types with stand-alone stop messages into 1 group for analysis, which may obscure differences that might have been detected with more observations. It is also possible that certain messages within a message category impacted attrition differentially, but because of somewhat varied content across schools, we were unable to examine attrition probabilities for each unique message. Third, although we are able to link attrition events to the most recently sent message, this does not necessarily indicate that a participant chose to drop out of the program because of the content of this particular message. In addition, program data indicate only that messages were sent to a functioning cellular number, not whether participants read the messages they receive or whether the messages were impactful. Therefore, participant retention is not itself an indication of either participant engagement or program efficacy.This study of attrition in the Text2BHealthy program demonstrates the potential of SMS text message programs to retain participants over a long program duration. In examining the patterns of attrition, we have provided evidence that the probability of attrition increases when participants receive SMS text messages with instructions about withdrawing from the program. Program planners should carefully consider how and how often to provide such information to minimize its effect on retention and determine other possible message content and characteristics that may undermine retention. Despite substantial progress in understanding best practices in SMS text message program design and implementation, more research is needed to determine participant, programmatic, and contextual predictors of program duration and attrition to mitigate their impact in SMS text message programs. Furthermore, the relationship between program duration and attrition and targeted behavioral outcomes also necessitates examination.


Young people with visual impairment (YPWVI) face several challenges in their everyday lives. However, little is known about interventions that focus on promoting their participation that contributes to health and well-being and is considered the most relevant outcome in rehabilitation. Objectives: This study investigated the clinical outcomes and acceptability of a new one-year, multiform, group-based rehabilitation program for YPWVI. The aim of the pilot program was to support them becoming more independent and to promote their participation. Rehabilitation consisted of group-meetings in an institutional setting, online group meetings, individually tailored one-on-one guidance, individual online discussions and parents’ group meetings. Fifteen young persons with visual impairment were recruited and 14 completed the intervention, six of whom were blind or had severe visual impairment and eight had mild visual loss. Methodology: The study utilized a mixed methods triangulation design. Clinical outcome measures were goal attainment scaling (GAS) and occupational performance (COPM) completed with qualitative interview data. Focus group interviews with participants and parents were used to evaluate the acceptability of the program. Results: GAS-rated personal goals were widely achieved and the scores of both performance and satisfaction scales of COPM improved. Overall, the rehabilitation program proved to be acceptable. Group-based rehabilitation was deemed very important and it enabled peer support. However, two-day periods of inpatient rehabilitation, proved to be too short, whereas five-day periods were considered to disturb schoolwork. Conclusions: Group-based multi-form rehabilitation for YPWVI can have a positive impact on activity and participation of the participants. The program can support independence and the achievement of rehabilitation goals. The group format was applauded for providing social support and company. The program required some structural modifications. Young people with visual impairment face challenges in many areas of their lives. Disabled young people may often remain children longer than their peers due to, for example, overprotective parents and helpers, a lack of assistive devices or the nature of their disability . The selThese difficulties may hamper visually impaired young people’s transition to adulthood, and lead to potential difficulties in full participation as adults. Participation contributes to health and well-being, and in rehabilitation, participation is considered as the most relevant outcome . The IntRehabilitation is an integral component of health services, ensuring that people can realize their full functional potential in their own living and working environments . In relaIn Finland, the rehabilitation of people with visual impairments is funded either by municipalities or by the Social Insurance Institution (Kela). Kela organizes medical rehabilitation services for people who have severe limitations in activity and severe restrictions in participation and pays them rehabilitation allowances. The total number of young people aged 15 to 24 in Finland in 2016 was 633,704, of whom 761 were visually impaired: 450 male, 311 female . FinlandKela developed and arranged (2011–2012) a new one-year group-based outpatient rehabilitation program for young people with visual impairment to support them in their everyday lives. The general aims of the program were to support young people in becoming more independent and to promote their participation. The program was developed and arranged together with the Finnish Federation of the Visually Impaired (FFVI), an advocacy organization for the blind and the partially sighted, which also provides a special rehabilitation service with social element. The aims of this pilot study were to describe clinical outcomes and the acceptability of the piloted program.A one-year, multiform, group-based rehabilitation program for visually impaired young people was arranged in two regional groups, one in the Helsinki capital area (7 participants) and one in western Finland (8 participants). The rehabilitation consisted of both inpatient group meetings in institutional settings and online, and individually tailored one-on-one guidance in individually selected settings . Group meetings in an institutional setting were arranged in four modules. The first module (five days) dealt with themes of interpersonal skills and life management. At this phase, a great deal of effort was put into building group cohesion among participants. The themes of the second period (two days) were social and psychological well-being, sexuality and self-determination, leisure time, and hobbies. The third module (two days) dealt with education, work and the role of the employer (in Finland disabled people employ their assistants). During the last module (five days) the participants were given practical guidance and tips for everyday life. They also received support in planning their lives and looking forward to the future. Each day included six hours of rehabilitation and three hours of other supporting activities. In addition to professional supervisors, two young persons with visual impairment acted as a part-time support team for the participants. One-on-one guidance in individually selected settings included interviews before and after the program and three to five meetings based on the individual needs and wishes of the young people. The focus was on the performance of practical tasks in conjunction with personal goals. Participants learned, for example, the use of public transport, made college visits and learned to use ATM machines or their assistive devices. Participants were asked to arrange one-on-one meetings with supervisors independently without their parents’ support. In addition, supervisors encouraged participants to travel independently to the meetings. Both real time and asynchronous tele-rehabilitation were used in the online interventions. Real time monthly group meetings (chat rooms) were built around primary themes of the rehabilitation. Some group meetings did not have a predefined theme. Supervisors provided information on an as-needed basis during chat room meetings. Participants completed tasks related to the rehabilitation themes, and in the Moodle learning platform, participants had opportunities to read more information about the themes. Supervisors supported participants with personal e-mails. Tools for this section were financed by the Finnish Ministry of Education and Culture. Parents had one-day group meetings twice, once in the beginning of the rehabilitation and once at the end of it. The parents program included also individual meetings in the beginning and at the end of the rehabilitation. Parents had the opportunity to reflect the strengths of their children and their own parenthood as well as to suggest their wishes for the program. The rehabilitation professionals’ peaceful presence was considered highly important. The parents’ sessions supported the agenda of the children’s rehabilitation. They had group conversations about being a parent to a growing up young person with visual impairment, and about supporting their children becoming independent. They also discussed the different roles of parenthood. Contents of their children’s rehabilitation and plans for the future were presented to the parents. Parents were provided information about the FFVI. Counsellors presented their role and services to the children and youth with visual impairments and their families. At the end of the rehabilitation parents were given advice on how to support their children’s independent living and travelling. Ethical approval for the study  was obtained from the Ethical Review Board of the Social Insurance Institution of Finland in Helsinki. All participants gave their written informed consent for participation in the study.Inclusion criteria were age (16–22 years), visual impairment , motivation for rehabilitation, lack of other severe illnesses , lack of difficult cognitive problems and independence in daily activities . Fourteen young people with visual impairment participated in the program from the beginning to the end. They were 16–22 years of age, nine of them were male and five were female. Six were blind or had severe visual impairment and eight had mild visual impairment. All attended school—either in primary, vocational, or higher education—and took part in the rehabilitation program during their leisure time. One participant discontinued during its course. This study utilized a mixed methods triangulation design . In thisClinical outcome measures were the rehabilitees’ occupational performance and rehabilitation goal attainment. The measurements were carried out within the rehabilitation program by the rehabilitation professionals in the beginning and the end of the program. The Canadian Occupational Performance Measure (COPM) was used to capture self-perceived problems in everyday activities and participation. COPM is a semi-structured interview designed to identify the activities that a person wants, needs or is expected to perform . An occuGoal Attainment Scaling (GAS) was used to set and score the participants’ individual goals. GAS is a method of scoring the extent to which individual goals are achieved in the course of an intervention. Participants have their own outcome measures, but standardized scoring allows for statistical analysis. At the beginning of the intervention, each participant established two to five individual goals for rehabilitation with support from professionals. Each goal was rated on a five-point scale so that the “expected outcome” fell in the middle of the scale at a score of zero. Also the levels for “somewhat less” and “much less”, “somewhat more” and “much more” were defined. At the end of the rehabilitation program, the participants evaluated their goal achievement and scored the achieved performance ,22.n = 14) and their parents (n = 22) were interviewed by the first author of this article at the end of the rehabilitation program. The young persons were interviewed individually and their parents as couples when two parents took part in the study. Using open-ended questions modified according to the circumstances, the interviews focused on the impact of the rehabilitation program on social and independent living skills of the visually impaired participants. Additional discussions were held to describe the challenges young people experience in participation. This topic has been reported in another article about this pilot study . They faced challenges in participation most frequently with regard to domestic life, leisure activities, major life areas, interpersonal interaction and relationships, and mobility. Scores of both performance and satisfaction scales of the COPM improved at the end of the rehabilitation program. The group level mean score in the performance of the rated activities improved from 6.1 to 7.7 and satisfaction in the rated activities improved from 6.5 to 7.9.Each participant identified one to six goals in their rehabilitation program. In all, 39 goals were identified . Nearly all goals were associated with the ICF component Activity and Participation. At the end of the program, the group mean for the GAS T-score was 56.6 (range 41–72), which means that, on average, participants’ performance in their individually chosen goals was above the expected level at the end of the program. All participants progressed in some of their chosen goals. However, outcomes were individual and there were huge differences between participants. For some participants the change was related primarily to increased awareness and information on visual impairment . Overall, the participants had trained and learned many everyday skills during the program. However, many of the skills, such as more independent mobility and doing housework required long term practice and parental support before the participants were able to use the skills independently in their everyday life. Eleven out of fourteen participants could transfer at least one learned skill into everyday life. These skills were for example using public transport, transferring food from kettle to plate, frying an egg and using a bankcard.n = 6), acquiring goods and services (n = 5), and preparing meals (n = 4). These problems were dealt with during the program, and nine participants—most of them were blind––had goals related to domestic life in their rehabilitation program . Most of the goals related to acquiring good and services, such as shopping. Learning to shop is challenging especially for blind persons, as it requires learning routes and finding the courage to ask for help. One participant remarked that: “One achievement during the program has been that I have learned to go independently to the shop and ask for help there”. In addition to the guidance during the program, learning to shop has required parental support and guidance. Two other participants reported having practiced shopping, but not being able to do it independently at the end of the program. The largest proportion of the participants’ problems (24.6%) were related to domestic life such as doing housework  were associated with major life areas, most of them were related to basic economic transactions such as using an ATM machine. Many of the participants, mainly mildly visually impaired, had a goal  related to major life areas, such as school and higher education and basic economic transaction. Six participants progressed in educational matters. They had discussed professional and study choices or improved in their motivation to study. Two participants learned to use a bank-card and were able to transfer the skill into everyday life: “At least I learned to use bank card, I use it regularly.”Nearly one fifth of the participants’ problems  of the participants’ problems were associated with recreation and leisure. This included problems related to meeting friends, participating in sports and playing games. However, only two participants had goals related to these in their rehabilitation program. One of them got active in camp activities offered by FFVI.A large proportion  associated with interpersonal interactions and relationships, all participants said that they were happy to meet other visually impaired young persons during the program. Group based rehabilitation was a good platform to learn interaction skills with peers. One participant was happy about having learned to ask strangers for help during the program. “One of the participants told that: “I do not usually get along in groups, here it has gone surprisingly well”. Another participant said: “It is good to find out that I am not the only one.” And a third one: “In this program it is so different to be with other visually impaired, everything is better.” Some participants formed new friendships with each other during the program, but only few of them reported to keep in touch after the program. Many parents had the same question at the end of the program: how to tackle with loneliness?“It would then be easier in high school”.Two participants had communication-related goals. One of them wanted to learn discussion skills, as he considered it important to learn how to initiate discussions. At the end of the program, he thought that he had progressed according to his plan. Another participant wanted to learn to use computer-based communication techniques: Only few participants had mobility-related goals, but supervisors required each one of them to try independent mobility and to use public transport independently when they travelled to the inpatient module of the rehabilitation. Some participants reported that their ability to use public transport had improved during the program, when earlier they had primarily taken a taxi.Along with the program came independent mobility. [Our daughter] has travelled to program meetings independently. She was forced to do these tasks, but I believe that this has been good for her.”Especially parents applauded this: ““I have learned to take more responsibility of my own life”.Overall, the participants reported that the program supported them in looking after themselves and gave them visions about more independent living. Support from and relationships with supervisors and peers, and an encouraging attitude during the program gave a boost to the participants. Most of them became more active and were encouraged to look after themselves. “During the program I learned to fry eggs, and now I have been able to do so at home, and my mother lets me.” Still, many parents do the tasks automatically for their children. Two participants considered their parents overprotective. One of them thought that she did get some help with this during the program: “In a way this program has helped my mother more than me, she has learned to trust me more. She has learned to trust, to not always worry, that she always does so naturally.”Many participants reported the importance of their parents in encouraging and teaching independent living skills. In some families, parents allowed participants to practice their skills: At the beginning of the program, some participants either had improper assistive devices or were not able or willing to use proper devices efficiently. During the program, seven participants were either purchased new assistive devices or were referred to assistive device assessment. They also practiced to use a white cane and computers. Both the program participants and their parents considered the length of the rehabilitation program suitable. One year was a suitable time frame to work toward their goal achievement. Five-day modules made it possible to focus on rehabilitation topics with time and were easy to cope with. Most participants considered the two-day inpatient module to be too short with too much time spent in the program. The participants considered the contents of the rehabilitation modules to be worthwhile.Leisure time during inpatient periods was very important for the participants as it gave them the opportunity to spend time with their peers. Participants considered having slightly older peers as role models supporting them during the inpatient module to be an important component of the program that set a good example. “It has been good to have two groups. The group is small enough to learn to know each other…In a bigger group it is more difficult to get your voice heard. And those practical tasks, such as cooking, are easier in smaller group. Everyone could participate.” However, the two groups were different from each other. One group also spent leisure time together during the inpatient rehabilitation periods, whereas the other divided into smaller subgroups. The participants reported that it was unproblematic and nice to have both blind and mildly visually impaired participants in the same group: “It is only good to have both, the activity is not dependent on if you have sight or not, we all are similar, however.” Also, most parents considered this arrangement good, as it provided an opportunity for mildly visually impaired participants to help the blind participants, which might raise their self-esteem. On the other hand, blind participants were able to show the mildly visually impaired participants that you can manage even with severe challenges. On the other hand, some parents thought that it would be better to have a more homogenous group, because it would allow focusing on disability-centered problems. For example, asking for help is different for participants whose visual impairment is not visible than for those who are blind. Participants and parents highlighted the importance of not making changes to groups during the rehabilitation process. Young people with visual impairment considered group-based rehabilitation to be very important. The group provided the opportunity to get to know other visually impaired young people, learn from them, make friends and get peer support. The participants described the atmosphere in the groups as good and accepting. The size of the group (seven people) was considered good as it enabled both getting to know other people and individual participation. One participant said: “I would have liked encouragement with how to guide the child, to get support in how to give her more space.”Parents considered their program important, but too short. Time was spent largely on providing them with information and the time left over was not sufficient for discussions. Especially parents would have liked to have some spare time together. One parent considered getting much new information in such a short time as too heavy. Also, the time between parents’ meetings was lengthy and some of the parents proposed one extra day for parents in the middle of the program. Parents who were new to the topic of visual impairment considered the opportunity for peer support to be especially important. Some parents would have preferred more support during the program: Both the project participants and their parents considered one-on-one guidance to be useful and supportive. One-on-one guidance was implemented in real-life environments and situations that were important and meaningful for the participants and supported their rehabilitation goals. Performing practical tasks with a connection to personal goals with the support of the supervisors prompted true pleasure and a sense of goal achievement. Many of the young participants considered the support by professional supervisors to be more helpful than the support given by their parents. It is easier to accept support and feedback from an outsider.Online rehabilitation divided opinions, with girls being more positive and boys more critical. Some participants took part in all of the real time meetings and gained a lot from the asynchronous tasks, while some participated in only a few meetings and did not read or do any of the tasks in the learning platform. Most participants described that chat rooms were important places to meet other young people, when face-to-face meetings were impossible to arrange. The findings suggest that online rehabilitation supported direct communication between young people and interaction through the rehabilitation professionals decreased during the rehabilitation program. Furthermore, some participants and parents reported that online rehabilitation developed participants’ information and communication skills.”During one-to-one guidance we can focus on the place where we are and visit different places, in the chat we focus more on ideology.” Parents expressed that they would have preferred more information about the online intervention. Many participants reported that they had major difficulties in coordinating their school schedule with participating in the online chat room. Participation in the online interventions was disrupted also due to technical issues. Some participants experienced the online rehabilitation as unhelpful, for example the contents of the interventions were not relevant to them. One participant reported: No adverse effects were reported other than inpatient rehabilitation modules during school weeks having disturbed studying and students had to make up for all from school. However, even with the need to make up for missed time in school, the participants were not willing to have institutional rehabilitation modules during school holidays. Additionally, parents expressed concerns about their children managing both school and rehabilitation at the same time because school is very time-consuming for them. Results of this study show that a multiform, group-based rehabilitation program for young people with visual impairment is beneficial in promoting activities and participation. Due to, for example, parental overprotection and a lack of activities, it is beneficial to provide them extra support on their path towards independence. Some of the participants in this study will never be fully independent, but will live with interdependence instead, in which a person does some things with assistance and others without . In thesGroup-based rehabilitation proved to be especially important for the young participants. Like the participants in this study, young persons with disabilities consider themselves lonely more often than those without disability ,5. SociaFurthermore, peer support from more experienced young persons with visual impairment is important, as this way young people can better understand their potential and their parents can understand that having expectations for their children is both realistic and a critical part of their child’s development . Peer suInterestingly, participants were divided on the online components of the intervention and criticized them widely. Young people tend to spend their social lives on the Internet—according to a Finnish study, 16% of youths aged 13–16 spend more time with their friends on the Internet than face-to-face and 41% have good friends with Internet contact only . Young pAfter the completion of this program, the use of tele-rehabilitation has increased widely. As of yet, there seems to be only little evidence about tele-rehabilitation among visually impaired people . HoweverThe results of this study show that the program succeeded to focus not on underlying capacities but instead on the different kinds of skills needed in participation. Concrete action and being engaged in something that is meaningful and important for the individual concerned is a way for people to become what they want and have the potential to become . ThroughThe program for parents was too limited in this program. Parents play a crucial role in the socialization process of young people with visual impairment . There aOverall, the program is appropriate for young people with visual impairments. The program still requires some structural modifications. Possibly two or three 3–4-day inpatient rehabilitation modules would produce a similar outcome and disturb studying less. Online intervention components could be limited to informal discussions. Parents need a more intense program that could partly take place online.The strength of the study lies in its mixed methods triangulation design describing how visually impaired young people and their parent report the outcomes and acceptability of the piloted program. Subjective COPM and GAS measures were used in addition to the interviews as the method of data collection. The use of a mixed-method design has both theoretical and practical strengths by involving stakeholders in the research process and by enhancing context sensitivity in line with models such as the ICF, that leads to the potential ecological and external validity of research .Thus theThe group exceeded a mean normalized T-score of 50 on GAS, indicating that, on average, goals were above the expected level. On the other hand, this—as well as the broad range of the T-scores—may show difficulties in the goal-setting process, leading participants to set goals that were either too easy or too difficult to achieve. Originally, the COPM and GAS were intended to be used in a combination in this intervention. The COPM was first used by an external occupational therapist to identify the problems of young persons after which GAS was used by the group supervisors to define the goals for implementation. However, the ICF linking of the COPM problems and GAS goals showed that the setting of GAS goals was not based on the COPM assessment. The main reason for this was lack of cooperation between professionals. On the one hand, the group supervisors had thorough information on the participants from their medical background data and personal interviews that may have had impact on setting of GAS goals. Previous research  has showA multiform, group-based rehabilitation program for young people with visual impairment is beneficial in promoting activities and participation. It is necessary in a rehabilitation program to focus on the goals and areas that the participants themselves consider important. The crucial elements of the program are practical support and focus on action within real life contexts, both individual and group interventions, peer support, and parental support. Difficulties in activities and participation, as well as restricted environmental factors of young people with visual impairment should be recognized in all health care, social care and educational settings to promote full participation and wellbeing of the young.