Extracorporeal membrane oxygenation (ECMO) can deliver effective respiratory and circulatory maintenance to organ donors, improve organ function, and shorten warm ischemic time before harvesting. However, ECMO-supported brain-dead donors (DBDs) still have a high risk of acute kidney injury related to decreased renal oxygen delivery and inflammatory damage, which may cause early graft failure.Kidney transplantation from an ECMO-supported DBD.We found an extremely abnormal “very dark blue” appearance of the graft kidneys from an ECMO-supported DBD during kidney procurement.Rather than discarding the graft kidneys, we performed an on-table biopsy. Pretransplant biopsy results revealed minimal interstitial fibrosis in the section of these graft kidneys.Two candidates received graft kidneys, and the two grafts remained functional until the 8-month follow-up.Currently, there is no standard method for evaluating graft kidney function of ECMO-supported DBDs. Regardless of the donors’ preoperative serum creatinine (SCr) level, estimated glomerular filtration rate (eGFR), or gross appearance of the graft kidney, we believe that it is more reliable to include pretransplant biopsy as a criterion in clinical practice to safely accept kidneys from ECMO-supported DBDs. Compared with kidney transplant using non-ECMO organs, kidney transplant using ECMO-supported brain-dead donors (DBDs) does not appear to have a higher rate of graft dysfunction.,4 and may be a cause of early graft failure. Currently, there is no standard evaluation criteria to assess such graft kidneys. Herein, we report our experience that pretransplant biopsy could be more important than gross appearance, serum creatinine (SCr) level, or estimated glomerular filtration rate (eGFR) to safely accept kidneys from ECMO-supported DBDs.However, kidneys from ECMO-supported DBDs are still at a high risk of acute kidney injury (AKI) related to decreased renal oxygen delivery to inflammatory damage22.1The donor, a 39-year-old male with a history of schizophrenia, fell from a fifth floor window and was taken to Tri-Service General Hospital, Taipei, Taiwan. Computed tomography scan revealed multiple rib fractures, pelvic fracture with active bleeding, and dissection of the descending aorta with intramural hematoma. Consequently, the patient underwent bilateral anterior thoracotomies and transcatheter embolization of the pelvic vessels for acute resuscitation.After 14 days, the patient underwent thoracic endovascular aortic repair of the descending aortic aneurysm. During the procedure, the patient experienced oxygen desaturation and cardiac arrest, and advanced cardiac life support was performed immediately. After return of spontaneous circulation, ECMO was set up due to persistent bradycardia and poor tissue perfusion, possibly caused by pulmonary embolism.Hypoxic-ischemic encephalopathy was not improved after 1 week; therefore, the diagnosis of brain death was made by 2 senior doctors, and the family decided to enter the critical hospice pathway and donate his organs.2.2Organ donation was made at 21 days after the patient's trauma (7 days after the ECMO was placed). The patient's initial SCr on admission was 0.9 mg/dL and rose to 4.7 mg/dL at organ harvesting. During kidney procurement, we discovered an extremely abnormal appearance of the kidneys, which were “very dark blue” in color  and underwent hemodialysis and the other was a 58-year-old female with immunoglobulin A nephropathy leading to ESRD who underwent peritoneal dialysis Table . Both re2.3With the use of immunosuppressive therapy, the SCr of these 2 patients decreased from 8.5 and 17.6 mg/dL to 1.1 and 1.8 mg/dL, respectively, at the 1-month postoperative follow-up, and these 2 grafts remained functional at the 8-month follow-up (Table Written informed consent to publish the case report was provided by the patients, and the consent procedure was approved by the Ethics Committee of Tri-Service General Hospital.3 High incidence of AKI in ECMO-supported patients has been reported in several studies.,6The shortage of kidneys from cadaveric donors is a global problem. As a bridge to brain death declaration, the concept of using ECMO to expand the donor pool has become widely accepted. Despite the advantage of maintaining organ perfusion, using ECMO also carries the risk of decreased renal oxygen delivery and AKI. have described the global kidney score based on pretransplant donor biopsy, while other systems have used SCr, eGFR, or macroscopic anatomy in combination with clinico-histopathological parameters to streamline and determine the allocation of kidneys that otherwise would have been discarded.–12There are various protocols for estimating the quality of a graft kidney. For conventional deceased or living donor kidneys, Remuzzi et al physicians may get confused whether the graft kidneys should be allocated based on pre- or post-ECMO condition.To our knowledge, there is no standard method for evaluating kidneys from ECMO-supported DBDs. Because ECMO activates an acute inflammation-like response and leads to acute tubular necrosis within 48 hours, In general, the macroscopic appearance indirectly reflects graft health, especially when acute tubular necrosis or infarction occurs. However, in this case, histopathological examinations have proven to be more reliable and precise with respect to graft function and survival. This case report suggests that preoperative evaluation and intraoperative frozen section consultation play a more important role in ECMO-supported donation than previously thought.In a recent study, SCr or eGFR level of ECMO-supported DBDs did not seem relevant to kidney graft survival.4Using ECMO maintains organ perfusion, and ECMO-supported kidney donors expand the donor pool, but the risk for AKI related to decreased renal oxygen delivery and inflammatory damage might be a cause of early graft failure.In summary, rather than SCr, eGFR, or macroscopic anatomy, we believe that it is more beneficial to include pretransplant biopsy as a criterion in clinical practice to safely accept kidneys from ECMO-supported DBDs. Moreover, long-term follow-up is required to confirm the benefit of this practice.Conceptualization: Sheng-Tang Wu.Data curation: Chiao-Ching Li, Meng-Han Chou, Yu-Chun Lin.Supervision: Sheng-Tang Wu.Writing – original draft: Pei-Jhang Chiang.Writing – review & editing: Shou-Hung Tang.


To investigate a stem‐adjusted preparation of the femur with a kinked reamer and to determine whether this approach results in higher primary stability of a kinked stem than straight reaming of the intramedullary canal.vs kinked reaming [KR] group). The specimens were analyzed using CT to determine the anchoring length and examined with a mechanical testing system to establish their axial stiffness, torsional stiffness, and migration distance after 10 000 gait cycles.Ten cementless stems with a kinked design were implanted in synthetic femurs after preparation of the femoral canal with 2 different reamer designs . Only for the SR group, a correlation was found between the completely conical anchorage length and absolute stem migration . Regarding the torsional stiffness, no differences were observed between the study groups . In the KR group, significantly higher axial stiffness values were measured than in the SR group .The stem migration distances did not differ significantly between the groups (SR group 0.51 ± 0.16 mm The implantation of a kinked stem after kinked conical intramedullary preparation of the proximal femur showed equivalent results regarding anchoring length, stem migration, and torsional stiffness to those for straight conical reaming. The specimens with kinked reaming showed significantly higher axial stiffness values. Nonetheless, if the prosthetic stem does migrate, it is much less likely that secondary fixation of the implant will be necessary.Currently, implantation of cementless stems is the gold standard for revision and fracture arthroplasty in the proximal femur with the aims of long diaphyseal anchoring and long‐term stability of the prosthesisRegarding the geometry of the prosthesis, implants have been developed with different biomechanical considerations for stable stem anchoring. One type of implant has curved or kinked prosthetic stems because these compensate for the need for an additional osteotomy in the femoral medullary cavity, which may be necessary during the implantation of a straight stem. Curved or kinked stems should follow the anatomical pattern of the femur better than straight stems so that more extensive anchorage is possibleFor straight stems, the femoral bone canal is drilled by a reamer, matching the design of the stem. The implantation of a curved stem occurs after the femur is prepared by drilling with a straight or flexible reamer. The latter is determined by the given femoral geometry. Thereafter, accurate matching between the femur and an anatomic curved stem cannot be achieved. A compromise between the non‐anatomic straight stem and the anatomic curved stem is the kinked stem. This stem is also not anatomically shaped, but it follows the antecurvation or varus form of the femur better than a straight stem. In contrast to a curved stem, a kinked stem allows the customized preparation of the femur with a kinked reamer. The outcomes of femoral revision using cementless fluted modular stems have previously been published by several research groups, and these studies have demonstrated good results at short‐term and long‐term follow‐up evaluationsTherefore, the objective of this study was to evaluate a stem‐adjusted preparation of the femur with a kinked reamer and to determine whether this approach results in greater primary stability of a kinked stem in contrast to the straight reaming of the intramedullary canal.For evaluation in this study, we selected a cementless stem . The standard measurements included a caput–collum–diaphyseal (CCD) angle of 135°, a midshaft outer diameter of 27 mm, and an inner canal diameter of 13 mm. Prior to implantation of the stem into the synthetic bones, femora for the two groups were prepared by sawing the proximal (subtrochanteric) portion. For accurate intramedullary interfaces between the bone and the fluting part of the stem, the resection line was based on the nominal length and diameter of the stem. The inner canals of the femora were reamed to match the stem diameter, which was performed by following the operation instructions that had been provided with the equipment. In the first group, the intramedullary canal preparation was performed with a straight conical reamer  to evaluate the metal–bone interface Fig. A–E. The The experiments were conducted with a servohydraulic testing device . For the load cell, the axial force capacity was ±10 kN, the torque capacity was ±100 Nm, and the accuracy was ±0.5%. The test reports were all grouped automatically by the FastTrack console software used by the multi‐axis fatigue system . We ensured that all hardware devices were working and that the same devices were used for all specimens.To precisely analyze the axial stiffness of the specimen, biomechanical conditions were simulated. We arranged the axes of the femora in a 10° movement towards the median line due to the force transmission in the proximal femur, simulating the single‐legged phase of walking. Depending on the specimen, the condyle region was fixed in a negative imprint in a vice. Then, the stem end was inserted into a suitable femur head. By fixating a compatible inlay on a metal plate of the testing device, the forces were transmitted to the specimen. Vertical loads with a linear ramp‐up/ramp‐down waveform consisting of 10 000 cycles and a load of 2.4 kN were applied to the femur head see Fig. A. For thTo enable testing of the torsional stiffness, the long axes of the femora were aligned vertically. This test was performed in the frontal and sagittal planes using a leveling gage, and the positioning was maintained in relation to that of the loading machine. The end of the stem was attached to a metal plate with a screw. Torsional loads were then applied using a linear ramp‐rotation .For the statistical analysis, a two‐sided Mann–Whitney Each sample was scanned and inspected, and the femur–stem interface areas were then calculated. No significant differences were observed among the contact patterns of the implants against the inner cortical bone Table .P > 0.05). No differences were found between the two groups for local resolution of implant impaction. Despite different intramedullary preparations, the SR group demonstrated conical and multiple‐point anchorage, and the KR group exhibited more conical surface anchorage.Implants with a straight conical reaming preparation corresponded well with the conical anchorage system, and the cortical contact circumference was more than half of the total anchorage length. In contrast, the group with kinked intramedullary femoral preparation exhibited conical surface contacts along nearly two‐thirds of the anchoring track . For the SR group, a correlation was found between the completely conical anchorage length and absolute migration .Torsional loading does not cause any implant migration, but this is not the case for axial loading. Therefore, for all of the implants under investigation, the torsional stiffness remained unchanged from the first to the last gait cycle, and no differences were observed between the study groups et al. found that the torsional stiffness was increased 2.2 times for cylindrical versus flexible reamers in cases with the same type of cementless press‐fit stemMany years have passed since cementless revision arthroplasty with distal fixation was first shown to be a successful method. This technique involves the principles of press‐fit and multiple‐point impactionFor evaluation of this issue, a kinked stem was chosen in this study. This implant is a compromise between the non‐anatomic straight stem and the anatomic curved stem. The kinked stem follows the antecurvation of the femur and allows customized preparation of the femur with a kinked reamer. This conical reamer prototype with a kinked geometry of 3° of angulation is intended to better align with the curvature of the femur than that of a straight conical reamer. In the investigated osseous defect model, the different intramedullary preparations of the proximal femur did not differ in localization or length of fixation of the kinked stem. Conversely, a correlation between the maximal stem migration after straight reaming and the circumferential conical anchoring length was found. The bigger the area of conical stem anchorage, the better was the primary stability of the stem. Femoral preparation with a kinked reamer showed no correlation with the circumferential conical anchoring length, but equivalent results were found for the maximal migration, with less deviation than the straight reamer. It could be assumed that the kinked stem is supported flatly by the 3° angulation after kinked reaming, resulting in a higher primary stability. No differences were found with regard to torsional stiffness or the correlation with the amount of metal–bone interface between the two groups. The torsional stiffness is generated by the star profile of the tested implants. It can be assumed that a short circumferential anchoring length is sufficient to achieve maximal torsional stiffness. In addition, the results showed that the axial stiffness of both groups was completely independent of the conical anchorage length. It is notable that the maximal axial stiffness of the tested stems was significantly increased, with less deviation in the group with kinked reaming than that with straight reaming.in vivo conditions. For revision arthroplasty , the qualities of a synthetic femur are quite different from those of a real human femur, especially when different reamer designs are compared. In conclusion, the implantation of a kinked stem after kinked conical intramedullary preparation of the proximal femur showed equivalent results regarding anchoring length, stem migration, and torsional stiffness to straight conical reaming preparation. In terms of axial stiffness, the specimens with kinked reaming showed significantly higher values. Prospectively, we will advocate the use of a kinked reamer system for the implantation of kinked cementless stems for revision hip arthroplasty.Our study has several limitations. The biomechanical stability of the press‐fit stem and the comparison associated with it in each group depended on equal reaming of the synthetic femur. Reaming is associated with the intramedullary stem length and with the impaction pressure of the prosthesis on radial interference. The main principle of fixation also involves the cortical force distribution and yields an individual press‐fit result. In our opinion, the most crucial limiting factor was adapting a piece of synthetic bone to simulate actual


Lipids and starch are important feedstocks for bioenergy production. Genetic studies on the biosyntheses of lipids and starch in green microalgae have drawn significant attention recently. In these studies, quantifications of lipids and starch are required to clarify the causal effects. While lipids are assayed with similar procedures worldwide, starch in green microalgae has been measured using various methods with deficiencies in accuracy or high cost.http://dx.doi.org/10.17504/protocols.io.2mhgc36.A simple, accurate and low cost procedure for routine quantification of starch in green microalgae was developed. This procedure consists of quick-freezing of the cells, solvent extraction of the pigments, 134 °C autoclaving and glucoamylase double digestions of starch, followed by a glucose assay using the dinitrosalicylic acid reagent. This procedure was optimized to quantify starch in small volumes of green microalgal culture. The accuracy of starch quantification using this procedure was 102.3 ± 2.5% , as indicated by using cornstarch as internal controls. The working protocol is available at This quantification approach overcomes the current problems in the starch quantification of green microalgae such as inaccuracy and high cost. This approach would provide an opportunity to compare the effects of genetic, physiological or cultivation manipulations on the productivity of starch in green microalgae elucidated in different labs, which is essential in the enhancement of lipid productivity studies in microalgae. Biodiesel is superior to bioethanol in terms of production considerations, and their raw materials are lipids and starch, respectively Chisti . Althougo-dianisidine to produce color for the spectrophotometric measurement. The three enzyme reactions compromise the simplicity and accuracy of this approach in addition to the cost considerations. The most costly methods involve the use of commercial assay kits, which are unlikely to be adopted for routine assays.It has been speculated that starch biosynthesis must be suppressed in order to enhance lipid productivity in green microalgae  and the chemical dinitrosalicylic acid were adopted Miller . This prhttp://dx.doi.org/10.17504/protocols.io.2mhgc36.The working protocol is available at Chlamydomonas reinhardtii UTEX 90, a wildtype strain, was purchased from the Culture Collection of Algae at the University of Texas at Austin, USA. The cells were propagated under continuous 150 μmol photon/m2/s white light at 25 °C in a modified Bold 3 N medium which contains 1.1 mM NaNO3, 0.05 mM K2HPO4, 0.16 mM KH2PO4, 0.17 mM CaCl2, 0.3 mM MgSO4, 0.43 mM NaCl and minerals including 6.56 µM FeCl3, 0.25 µM ZnSO4, 2.42 µM MnSO4, 5.69 nM CoSO4, 6.1 nM Na2MoO4, 1 nM Na2SeO3, 6.3 nM NiCl2, described in Table 2 of Berges et al.  were purchased from Sigma-Aldrich, USA. Glucoamylase  was purchased from Tokyo Chemical Industry, Japan . This enzyme completely hydrolyzes soluble starch, amylose, and amylopectin (see in BRENDA database). Potassium sodium tartrate  was purchased from PanReac AppliChem, Spain.Corn starch (S5296), glucose (G5146), dinitrosalicylic acid (D0550), NaOH (S8045), and NaH540) of each mix was measured. A standard curve was built based on the glucose quantity in each mix against its OD540 . A solution of 10 mM was prepared and stored at 4 °C. Serially twofold diluted glucose solutions, 0.5 mL each, were mixed with 2 mL DNS reagent  separately and then heated in boiling water for 5 min. After cooling in tap water, the optical density at 540 nm  method described in the previous “682 around 1.0) were harvested using swing bucket centrifugation  at room temperature. The supernatant was discarded and the cells were transferred to a 2-mL screw cap microtube rapidly. After a brief high-speed centrifugation, the supernatant in the microtube was removed using a pipette and the cells were quickly frozen at − 15 °C in a mix of ice and crude sea salt. One microliter of cold methanol/tetrahydrofuran (v/v = 1/3) was added to the frozen cells. The cells were soaked in the solvent for 30 min in the ice-salt mix and agitated occasionally to extract the pigments. After centrifuging at 16,000g at 4 °C for 10 min, the solvent was discarded and the pellet was dried at 65 °C for 1 h. The dried pellet was repeatedly washed out using the sodium phosphate buffer  and the final volume was adjusted to 5 mL. A small amount of corn starch, weighed using the high accuracy analytical balance aforementioned and listed in Table Ten microliter of the day 5 microalgal culture , which together comprised 1.16% raw weight of corn starch. The amounts of other components such as lipids and minerals were very little as shown in our results and in the USDA database. They were thus ignored in the net weight calculation. Therefore, the net weight of starch (glucose polymer) was 85.3% of the raw weight of the corn starch used in this study.As in the grains of crops, corn starch contains certain levels of water, protein and cellulosic fiber that affect the measurement of the net weight of starch (glucose polymer). Corn starch samples, stored in the lab refrigerator, weighing more than 0.3 g were dried at 105 °C for 2 h in a ceramic crucible. The weight difference of each sample before and after the drying was measured using a high accuracy analytical balance. The water content in the starch was determined to be 13.5 ± 1.3% . The contents of protein and fiber in the corn starch were measured by the U.S. Department of Agriculture  was employed to extract the pigments, and 30 min of extraction gave satisfactory results with the cells of the day 5 culture. After autoclaving at 134 °C and glucoamylase double digestions, the algal cells with or without the additional corn starch  were smashed and centrifuged to collect clear supernatant for the glucose assay. As shown in Table This simple and accurate procedure that was built based on the corn starch internal controls for microalgal starch quantification is also low-cost, due to the use of the thermo-tolerant glucoamylase. This enzyme is able to hydrolyze α-1,6-glucosidic bonds in starch in addition to α-1,4-glucosidic bonds . The cost of the enzyme per assay (4 units/assay) in this study was less than USD 0.5 cent, a dramatic difference compared to USD 14.2 using the Sigma-Aldrich starch assay kit and Euro 2.6 using the Megazyme kit purchased in Taiwan. The mass of the internal controls was measured using a high accuracy analytical balance, which gave the readings very close to their true values. This provides the possibility of gauging the accuracy of the results obtained using this quantification method.340. The two assays do include starch standards, but not internal controls, to calibrate the results from measuring unknown samples. However, the interferences from water contents and other non-glucose components in the starch standards were ignored in the protocols of the two kits. Besides, the three enzyme reactions are also a concern. Generally, more enzyme reactions give more bias in the quantifications.In the study of Rose et al. , the “acThis quantification procedure has advanced starch assay one step further by using the internal controls compared to other methods. In addition, this method has a high sensitivity. The sensitivity of the DNS method described in this procedure is 28 μg of glucose per reaction at the low end of the standard curve Fig. . One micg at room temperature for 10 min) to precipitate the cell debris after the double digestions of the microalgal samples was required to obtain high accuracy of starch measurement.Found in this study, the action of quick-freezing of the cells at − 15 °C right after harvest had positive influence on the accuracy of starch quantification. This might be due to the starch degradation by the endogenous microalgal enzymes at room temperature when the cells were agitated. The measurement accuracy of the internal controls being a little over 100% on average in Table Although it takes 2 days to quantify microalgal starch using this procedure, the work is not intensive since most time in the 2 days is used for autoclaving (about 4 h from the beginning to the end) and enzyme digestion (one night plus 7 h in the second day). This procedure will be a great tool for the studies of biochemical and genetic engineering that involve in starch biosynthesis or degradation in green microalgae.


A femoral crossover polytetrafluoroethylene graft was performed in a patient immediately, after a failed iliac endograft limb recanalization, performed 2 years after endovascular aortic aneurysm repair. Although graft patency was confirmed clinically by palpation of groin pulses as well as noninvasively by segmental pressure measurements and Doppler examination, in the immediate postoperative period, duplex ultrasound failed to show any blood flow inside the graft lumen until the fourth postoperative day. Subcutaneous air and air tapped within the wall structure of the graft are possible explanation for the failure to show flow. A 7-mm femoral crossover polytetrafluoroethylene (PTFE) graft was performed after a failed endovascular attempt to recanalize a thrombosed endograft limb  in a paSince the acoustic impedance and elastic stiffness of the expanded PTFE (ePTFE) graft wall are higher than in human native arteries, synthetic grafts greatly attenuate acoustical signal strength.The phenomenon of reflective air within the interstices of an ePTFE graft, early on, merits attention of vascular experts, especially now that more and more femoral crossover bypasses are performed along with EVAR in patents with concomitant unilateral iliac artery chronic occlusionOne decade long example of this phenomenon is, of course, the Viatorr tips endograft, which appears “occluded” early on, until the graft “wets out.” The Viatorr stent graft uses an ePTFE lining that is biocompatible, microporous, nonthrombogenic, and relatively impermeable to blood and tissue and provides a substrate for endothelial lining. The blood-contacting inner layer is made of ePTFE and has a microstructure and mechanical properties that are similar to those of the conventional vascular graft from the same manufacturer .These observations may be helpful for the vascular surgeon's routine practice.


Delayed graft function (DGF) is a frequent complication after deceased donorkidney transplantation with an impact on the prognosis of the transplant.Despite this, long-term impact of DGF on graft function after deceased donorkidney transplantation has not been properly evaluated.The main objective of this study was to evaluate risk factors for DGF and theimpact of its occurrence and length on graft survival and function.A retrospective cohort study was performed in 517 kidney transplantrecipients who received a deceased donor organ between January 2008 andDecember 2013.The incidence of DGF was 69.3% and it was independently associated withdonor's final serum creatinine and age, cold ischemia time, use of antibodyinduction therapy and recipient's diabetes mellitus. The occurrence of DGFwas also associated with a higher incidence of Banff ≥ 1A grade acuterejection (P = 0.017), lower graft function up to six years aftertransplantation and lower death-censored graft survival at 1 and 5 years (P< 0.05). DGF period longer than 14 days was associated with higherincidence of death-censored graft loss (P = 0.038) and poorer graft function(P < 0.001). No differences were found in patient survival.The occurrence of DGF has a long-lasting detrimental impact on graft functionand survival and this impact is even more pronounced when DGF lasts longerthan two weeks. Delayed graft function (DGF) is a frequent complication after deceased donor kidneytransplantation with an impact on the prognosis of the transplant.The consequences of the high DGF incidence are striking in terms of costs, morbidity,and perhaps mortality.Even though the impact of this condition has been extensively reported, theconsequences of DGF length on graft survival and function are less certain. It isconceivable that prolonged duration of DGF may be associated with inferior graftoutcomes. The present study was undertaken to evaluate the risk factors for DGF andthe impact of its duration in long-term kidney graft survival and function.We performed a retrospective cohort study that included 517 kidney transplantrecipients who received a deceased donor organ between January 2008 and December2013 at our institution. Twenty-eight patients were withdrawn from study analysissince the diagnostic criteria for DGF could not be evaluated due to primarynon-function, early deaths, and early graft losses. DGF was defined by the need fordialysis in the first week after transplantation; all patients were followed for atleast one year and up to six years. Risk factors for DGF were evaluated along withrecipient demographic data such as age, gender, race, time on renal replacementtherapy, primary kidney disease, HLA mismatches, presence of donor-specificantibodies, panel-reactive antibodies, and immunosuppressive regimen including theuse of antibody induction therapy. Donor-related variables included demographicdata, final serum creatinine, cause of death, history of hypertension, and beingclassified as expanded criteria donors (ECD).Duration of DGF was measured in days and the last DGF day was considered the one inwhich the last dialysis treatment was undertaken. The outcomes evaluated were: (a)incidence of DGF; (b) incidence of acute rejection evaluated throughout thefollow-up period; (c) estimation of graft glomerular filtration rate (eGFR) by theMDRD equation according to the presence of DGF and its duration; (d) Kaplan-Meieranalysis of patient and graft survival according to the presence or absence of DGFand DGF length.Statistical analyzes included normality evaluations of the data performed byShapiro-Wilk and Kolmogorov-Smirnov tests. Turkey's test was used for the analysisof DGF into quartiles according to its duration. The variables were subjected tounivariate analysis and those that reached a P level ≤ 0.20 were included ina multivariable analysis by Poisson regression in order to independently evaluaterisks factors.All data analyses were performed with the IBM SPSS Statistics program version 20 anda P value < 0.05 was required for statistical significance.The study was approved by the institution's ethics and research committee.Demographic data is shown in Risk factors were classified as donor-related, recipient-related, andgraft-related. They were analyzed by univariate and multivariate methods . Donor'sThe most frequent recipient causes of chronic renal failure were hypertensivenephropathy (24.6%), diabetic nephropathy (21.1%), adult polycystic kidneydisease (12.8%), and chronic glomerulonephritis (7.7%). Donor's causes of deathwere predominantly cerebrovascular disease (56.0%) and trauma (34.6%), whichwere not risk factors for DGF (data not shown).DGF occurred in 339 patients, reaching an incidence of 69.3%. The incidence waselevated over the six yearly cohorts ranging from 59.8% in the lowest and 74.4%in the highest incidence year. Two hundred and eighty patients (57.3%) wouldhave been considered as having DGF if the condition was defined by the need ofmore than one dialysis session in the first post-transplant week.The impact of DGF on graft function is shown in Overall, prior to any exclusion, patients and uncensored grafts survivals at oneand five years after transplantation were 95.6% and 86.5%, and 84.0% and 69.1%,respectively. At one year after transplantation, patient survival in the DGF andnon-DGF groups were 96.5% and 96.0%, respectively and remained essentiallyidentical throughout the follow-up (log rank P = 0.601). However, astatistically significant difference was found in death-censored graft survival(P = 0.038). At one year after transplantation, graft survival was 94.0% (DGFgroup) and 96.6% (non-DGF group), and at five-year, survival were 84.6% and95.0%, respectively (P = 0.038). Sixty-five graft losses occurred in thefollow-up. The main causes of graft loss were vascular thrombosis in 26 cases(40%), rejection in 14 cases (21.5%), and chronic allograft failure in 13 cases(20%). DGF duration did not impact on patient survival up to 6 years aftertransplantation; however, it exerted a significant impact on graft survival. Thegroup of patients with DGF longer than two weeks presented a significant lowergraft survival .In the first year after transplantation, the incidence of biopsy confirmed (Banff≥ 1A) that acute rejection was 24.5% in the DGF group and 14.7% in thegroup of patients without DGF (P = 0.017). Acute rejection superimposed on DGFled to significant lower eGFR and death-censored graft survival. As shown inAt six years after transplantation, death-censored graft survivals were: 95.5% inthe group of recipients without either DGF or acute rejection, 93.1% in thegroup without DGF and with rejection, 89.4% in the group with DGF and withoutrejection, and 73.9% in the group with both DGF and acute rejection. The groupwithout DGF or rejection presented significantly higher survival than the groupof patients with both conditions (P < 0.001). Also, the survival of the groupwithout DGF and with acute rejection was higher than the group with bothconditions (P < 0.001)., Delayed graft function is a frequent complication after deceased donor renaltransplantation and presents as graft acute renal failure resulting many times inpost-transplantation oliguria, need for dialysis, increased allograft immunogenicitywith higher risk of acute rejection, and may lead to decreased graft survival.5Donor-related factors may influence the occurrence of DGF, notably quality of donorintensive care during organ retrieval., 7, The incidence of DGF after deceased donor kidney transplantation presents a widevariation. The current reported average incidence in US is around 30 and 22% forkidney grafts from extended and standard-criteria donors, respectively.3In a recent North-American registry study, outcomes of 29.598 mate kidney transplantsfrom the same donor were evaluated, where only one transplant underwent DGF. Theauthors found that the risk of graft loss associated with DGF in the first yearafter transplantation was 5.35 times higher and remained between 16 and 30% higherafter the first year., 13, In the present work, the incidence of DGF was significantly higher than the meanincidence in the US, and similar with previous Brazilian reports.3Renal grafts from expanded criteria donors have also been associated with a higherDGF incidence compared to grafts from standard criteria donors , 12, 13,4,In the present work, we examined, in a setting of high DGF incidence, the riskfactors and prognostic significance to verify if poorer DGF-related outcomes wouldsustain. As previously reported by Brazilian studies, an elevated incidence of DGFwas found.4Tedesco-Silva and collaborators showed that machine perfusion of kidneys graftssignificantly decreased the incidence of DGF from 61 to 45% compared with staticpreservation in a recent multicenter Brazilian study.,17, Interestingly, the impact of DGF length on graft outcomes has not been frequentlyreported.5Graft recovery of function is another important DGF related concern. Lee andcollaborators evaluated the impact of DFG recovery on graft function and found thatthose kidneys with complete recovery have similar survival to the ones that did nothave DGF. Contrariwise, those with uncomplete recovery presented lower survival andlower GFR at five years after transplantation. In a meta-analysis study, Yarlagadda and collaborators showed that patients with DGFpresent lower graft function compared with those without DGF at 3.5 years aftertransplantation.,24,30,32,DGF has been related to a higher incidence of acute rejection in kidney transplantrecipients.7In our study, patient survival was not influenced by the occurrence or duration ofDGF up to six years after transplantation. The relationship between DGF andmortality has been evaluated in many studies. Yarlagadda and collaborators performeda systematic review and meta-analysis, including twelve studies, evaluating theimpact of DGF on mortality of kidney transplant recipients and found no associationbetween DGF and patient survival up to five years after transplantation.The present study has limitations including being single center and its retrospectivedesign that prevented a better assessment of a number of aspects, but in particular,the quality of donor care. However, we believe that the data brought out significantfindings regarding the impact of DGF length in graft survival and function.In conclusion, in a setting of high DGF incidence, prolonged DGF is associated withinferior graft survival and function. Acute rejection is more frequent in patientswith DGF and its occurrence further aggravates survival and function. The reasonsfor the high DGF incidence could not be identified in the present study. Wespeculate that donor maintenance-related factors might be involved in such findingand that better donor management protocols, care, and organ retrieval might helpimprove this outcome.


A worldwide lack of donor corneas demands the bioengineered corneas be developed as an alternative. The primary objective of the current study was to evaluate the efficacy of acellular porcine corneal stroma (APCS) transplantation in various types of infectious keratitis and identify risk factors that may increase APCS graft failure.In this prospective interventional study, 39 patients with progressive infectious keratitis underwent therapeutic lamellar keratoplasty using APCS and were followed up for 12 months. Data collected for analysis included preoperative characteristics, visual acuity, graft survival and complications. Graft survival was evaluated by the Kaplan–Meier method and compared with the log-rank test.The percentage of eyes that had a visual acuity of 20/40 or better increased from 10.3% preoperatively to 51.2% at 12 months postoperatively. Twelve patients (30.8%) experienced graft failure within the follow-up period. The primary reasons given for graft failure was noninfectious graft melting (n = 5), and the other causes included recurrence of primary infection (n = 4) and extensive graft neovascularization (n = 3). No graft rejection was observed during the follow-up period. A higher relative risk (RR) of graft failure was associated with herpetic keratitis  and graft size larger than 8 mm .APCS transplantation is an alternative treatment option for eyes with medically unresponsive infectious keratitis. Despite the efficacy of therapeutic lamellar keratoplasty with APCS, to achieve a good prognosis, restriction of surgical indications, careful selection of patients and postoperative management must be emphasized.Trial registration Prospective Study of Deep Anterior Lamellar Keratoplasty Using Acellular Porcine Cornea, NCT03105466. Registered 31 August 2016, ClinicalTrails.gov Infective keratitis is considered a major global cause of visual impairment –3. In seAcanthamoeba. In addition, another purpose of the present study was to identify some of the factors that may increase the therapeutic failure rate of APCS transplantation so that future patients with a high risk of graft failure may be excluded and avoidance measures may be taken.APCS is processed from fresh porcine corneas by removing nuclear and cellular materials . The appAcanthamoeba or viral etiology were consecutively enrolled in this prospective cohort study. Infectious keratitis was defined as medically uncontrolled if the disease had significantly progressed after a 2-week of maximum antimicrobial medication or if the cornea was at risk of perforation. All participants completed 12 months of postoperative follow-up. The diagnosis of nonviral infectious keratitis was achieved by smear, microbial culture, confocal microscopy and pathologic examination. The diagnosis of HSK was based on multiple episodes of recurrence and clinical manifestations . The exclusion criteria for APCS transplantation included an infiltration size larger than 9 mm and signs associated with infection that affected the corneal endothelium and anterior chamber. Hypopyon was not an absolute contraindication for ALK as long as it was due to severe intraocular sterile inflammation. Noninfectious hypopyon was distinguished from infectious hypopyon based on clinical appearance. Compared to infectious hypopyon, a noninfectious hypopyon is generally relatively more likely to shift with a change in head position. However, there is substantial overlap between these two entities. Moreover, if the lesions were noted to not extended to the level of Descemet’s membrane (DM), the hypopyon was presumed to be noninfectious. Nonetheless, the early intraocular spread of infection could not be fully excluded. It is even especially difficult to distinguish noninfectious hypopyon from infectious hypopyon in cases with a deep lesion. In light of these considerations, we prudently performed lamellar keratoplasty in cases with hypopyon.Thirty-nine patients (39 eyes) undergoing APCS transplantation in Zhongshan Ophthalmic Center, Sun Yat-Sen University to treat medically uncontrolled infectious keratitis with a bacterial, fungal, 60 radiation.The APCS graft used in the current study was prepared by AiNear Corneal Engineering Corporation  as previously reported . After iAPCS and human donor corneas were processed for routine paraffin embedding. Briefly, 5-µm-thick sections were cut and stained with hematoxylin and eosin (H&E). Tissues were also cut into pieces and processed for scanning electron microscopy  examination as described previously . SEM imaDeep anterior lamellar keratoplasty (DALK) surgery was performed in a minimally modified manner as previously described . BrieflyAcanthamoeba keratitis, topical steroids were withheld for at least 2 weeks, whereas for bacterial and viral infections, topical steroid treatment was initiated immediately after surgery. Topical steroids were generally prescribed four times daily for 6 months, gradually tapered down to a maintenance dose of once daily over a 3-month period, and then continued for another 3 months. Topical tacrolimus was used right after the surgery as an additional immunosuppressive medication and for its anti-inflammatory effect. Sutures were removed approximately 3 months after surgery.To prevent recurrent infection, we postoperatively administrated multiple topical and systemic antimicrobial medications. For nonviral keratitis, systemic antimicrobial treatments  were administered within 1 to 2 weeks postoperatively, while topical antimicrobial medications were usually maintained for at least 1 month after surgery and then tapered over the ensuing weeks if no infection recurred. For HSK, the postoperative antiviral treatment regimen comprised oral acyclovir 400 mg twice a day for 12 months and topical antiviral medications (ganciclovir and interferon) for at least 3 months. In cases of fungal and Clinical measurements included visual acuity, graft survival and postoperative complications. The size of the corneal lesion was reported as the equivalent circular diameter (ECD) and was calculated as previously described . Cornealhttps://www.R-project.org/). Comparisons of continuous data among groups were performed by applying Student’s t-test. Graft survival was evaluated by the Kaplan–Meier method, and comparisons were performed using the log-rank test. Due to the presence of nonproportional hazards between the nonviral and herpetic keratitis groups, the weighted log-rank test was used to compare the survival curves. A weighted log-rank test was carried out using the comp function in the R package “survMisc”, and Fleming–Harrington weights of P = 0, and q = 0.5 were chosen to emphasize differences later in time.Potential risk factors, including indications for grafting, the lesion size, the graft size and preoperative hypopyon, were assessed for associations with the incidence of graft failure. The statistical analyses were performed using SPSS 23.0  or R 3.6.0 (Thirty-nine eligible patients (39 eyes) were implanted with APCS to treat infectious keratitis. As shown in H&E histological stains, these replacements consisted of highly organized collagen fibers, yet no remaining cell (nuclei) were detected. SME revealed that the APCS reserved the structure of a native human corneal stroma but was less porous than the human cornea Fig. .Fig. 1ArAcanthamoeba keratitis . One case was diagnosed as fungal keratitis with concomitant bacterial infection. The size of the corneal lesions ranged from 2.78 to 7.34 mm . Descemetocele formation was observed in two patients with HSK (5.1%). Sterile hypopyon due to intraocular inflammation rather than active intraocular infections was noted in 4 cases (10.3%). Two of these cases were diagnosed with fungal keratitis, one case with Acanthamoeba keratitis and once case with fungal keratitis with concomitant bacterial infection. The trephination sizes of the recipients ranged from 7.25 to 8.50 mm .Demographic features of the recipient are presented in Table Corneal transparency was restored following APCS transplantation in cases without graft failure Fig. . Before Twelve patients presented graft failure within 1 year after keratoplasty Fig. . Of the The preoperative factors that influenced graft outcome are shown in Fig. Acanthamoeba. This study is also the first to disclose the major risk factors for APCS graft failure, and this information may help in future patient selection and the postoperative management of APCS transplantation.Corneal allotransplantation is a well-established technique for treating corneal blindness. However, there is a worldwide shortage of human donor corneas, and this has led to a demand for the development of biocompatible materials as an alternative . EnormouCorneal transplantation performed for active infectious keratitis is associated with a high risk of failure . The graGraft rejection is always the primary concern in xenotransplantation. While APCS implantation is by its nature a xenotransplantation, no evidence of graft rejection was observed in any of our patients at the last follow-up. Our findings are consistent with those presented in previous reports , 21. FreIn our study, five patients developed noninfectious graft melting, which is the most common cause of graft failure. Graft melting is a devastating complication of corneal transplantation. It is a destructive process involving collagen dissolution driven by the excessive release of tissue collagenase . One groAcanthamoeba keratitis varied from 73.9 to 85.7%; in contrast, only 42.9% (3/7) of our patients with HSK achieved therapeutic success. The extremely low survival rate of APCS grafts in active herpetic keratitis observed in this study is consistent with previous reports. For example, Zhang et al. performed APCS transplantation to treat 13 cases of herpes simplex virus (HSV)-related corneal ulcers or scarring [Several preoperative factors that exerted a profound impact on graft survival were identified in this study. First, we noted that APCS transplantation had a much worse prognosis in clinically active herpetic eyes than in eyes with nonviral microbial keratitis. In our series, the 1-year graft survival rate for cases of bacterial, fungal and scarring  and obseHowever, under some conditions, such as medically uncontrolled ulceration, ulceration at risk of perforation, and insufficient fresh donor corneas to meet demands, APCS transplantation may be applied. There are some management considerations that could improve surgical prognosis. HSK is typically characterized by recurrent episodes of infections, because HSV can be latent in the trigeminal ganglion . After kIn our study, another preoperative factor associated with the incidence of graft failure was graft size. We found that APCS grafts performed with a size ≥ 8.0 mm had a higher risk than smaller grafts of developing recurrent infection (22.2% vs 6.7%), graft melting (33.3% vs 6.7%) or extensive graft neovascularization (22.2% vs 3.3%). Williams et al. and Volker-Dieben et al. conducted a large retrospective analysis to evaluate the effect of graft size on graft outcomes. Both of these authors showed that graft failure rates were higher for grafts exceeding of 8.0 mm . In therMoreover, we found that compared to human corneal transplantation, in APCS transplantation, the graft size seemed to exert a more significant influence on graft failure. We suggest that this difference could be due to two reasons. First, as we discussed above, the use of a large graft might increase the chance that the graft will interact with immune cells derived from the corneal limbus. Even worse, APCS grafts are more prone than human donor corneas to interact with collagenase released by immune cells than donor human cornea . Second,A relatively short follow up period is a limitation of this study. Because serious complications, such as recurrent infectious and graft rejection, are more likely to occur within 1 year after corneal transplantation, our patients were followed up for at least 12 months. A future study will be designed to verify the long-term success of APCS transplantation for infectious keratitis.Based on these findings, although the number of patients was limited and the follow-up duration was short, this study provides some insights that will be valuable for the future application of APCS, including surgical indications and postoperative management.Additional file 1: Figure S1. Slit-lamp biomicroscopy performed at 2 (a) and 3 (b) years after implantation with acellular porcine corneal stroma (APCS). The implant was well integrated into the recipient cornea. The patient had a visual acuity of 20/25 in the operated eye.


There is a need for efficient and “off-the-shelf” grafts in urethral reconstructive surgery. Currently available surgical techniques require harvesting of grafts from autologous sites, with increased risk of surgical complications and added patient discomfort. Therefore, a cost-effective and cell-free graft with adequate regenerative potential has a great chance to be translated into clinical practice. Tubular cell-free collagen grafts were prepared by varying the collagen density and fiber distribution, thereby creating a polarized low fiber density collagen graft (LD-graft). A uniform, high fiber density collagen graft (HD-graft) was engineered as a control. These two grafts were implanted to bridge a 2 cm long iatrogenic urethral defect in a rabbit model. Histology revealed that rabbits implanted with the LD-graft had a better smooth muscle regeneration compared to the HD-graft. The overall functional outcome assessed by contrast voiding cystourethrography showed patency of the urethra in 90% for the LD-graft and in 66.6% for the HD-graft. Functional regeneration of the rabbit implanted with the LD-graft could further be demonstrated by successful mating, resulting in healthy offspring. In conclusion, cell-free low-density polarized collagen grafts show better urethral regeneration than high-density collagen grafts. This increases the risk of complications, the possible lack of tissue availability and patient discomfort. Certain oral pathologies are contraindications for buccal mucosal graft harvesting2. Both cell-free and cell-seeded urethral grafts have been investigated. To date, cell-based grafts have shown better functional results in preclinical large animal models and inconclusive results in subsequent clinical trials3. In 2015 and 2017, the clinical outcome of a commercial cell-seeded tissue engineered buccal mucosa product, was published5. This product requires a buccal biopsy to be taken, followed by ex-vivo cell culturing in a GMP facility and a 3-week manufacturing time. The cost-effectiveness and efficacy of these grafts have been debated7. Therefore, it is unlikely that this approach will become standard clinical practice for the patient with limited financial resources or insurance coverage. To overcome these problems we have targeted the development of a cell-free urethral graft that is cost-effective and adequate also to be used in third world countries.There is a clinical need for efficient and cost-effective grafts in urethral reconstructive surgery. Current surgical techniques require harvesting from autologous sites eg. buccal mucosa and penile skin tissueIn vitro the mechanical niche is an important parameter to be considered for controlling cell fate. For example, mesenchymal stem cells are known to differentiate into different cell lineages, depending on the substrate stiffness8. Previous studies have shown that fluid content or collagen fiber density relates to the stiffness and influenced material properties of the collagen gels10. The more extracted fluid from the collagen gels, the stiffer the collagen material gets. In this manuscript we address the question if collagen fiber density of a collagen graft influences the functional regeneration of the rabbit urethra? Two different cell-free bovine tubular collagen grafts were manufactured, one with polarized and low collagen fiber density distribution (LD-graft) and one control with uniform and high collagen fiber density distribution (HD-graft). Polarization was achieved by specific compression and the collagen density increased towards the luminal side of the tube. These grafts were implanted in rabbits and were examined post-implantation by histology and immunohistochemistry, and the functional outcome was assessed by voiding cystourethrography and mating.10. To fabricate the LD-graft, the gelated collagen was manually compressed by rolling it on filter paper, followed by an air-drying step . 8.5 mL of collagen solution were added to 0.8 mL of 10 × MEM and neutralized with 1 M NaOH (approx. 1.85 mL). The neutralized solution was set in a mold previously used for manufacturing tubular constructs of 3 mm inner diametertep Fig. . To fabrtep Fig. . MonitorThe samples were fixed with 1% tannic acid and 1.25% glutaraldehyde, then washed with 0.1 M cacodylate, and dehydrated in increasing ethanol concentrations prior to critical point drying. Thereafter coated with gold/palladium and imaged at a voltage of 10 kV using a scanning electron microscope .10.Burst pressure, ultimate tensile strength (UTS) and Young’s modulus of grafts (N = 4 in each group) were measured with an electronic manometer (Extech instruments HD750) or an Instron tensile machine  as described previously10. Rabbits from the long-term study group were involved in the in-house breeding program of the Animal Experimental Unit of the Faculty of Medicine, University Malaya.Following approval by the Animal Ethics Committees of the Canton of Vaud (Authorization number: VD-2740), Switzerland and of the Faculty of Medicine of the University of Malaya (Authorization number: 2013-07-19/SUR/R/TCR), Kuala Lumpur the experiments were performed on New Zealand white male rabbits  in Lausanne and Kuala Lumpur. All experiments were done in compliance with national directives for the care and use of laboratory animals. The surgical steps and post-operative monitoring were done as previously describedAnimals were submitted under general anesthesia for macroscopic evaluation and voiding cysto-urethrography (Visipaque 270 mg/mL). All images were collected with a Philips BV Pulsera. The diameter of the urethra was measured utilizing a scale. Knowing that the graft was sutured at 0.5 cm proximal to the base of the glans and it measured 2 cm in length, the position of the graft could be determined on the radioscopic images. It was then possible to estimate the potential presence of stenosis at the anastomotic sites. Stenosis was defined as a persistent 50% reduction of the diameter of the urethra at the same location.10.Entire penises were harvested and fixed in 4% PFA. Specimens were embedded in paraffin, and 8 µm thick sections were prepared. Antibodies used: Mouse anti-α-smooth muscle actin , Goat anti-uroplakin-2 , Mouse anti-caldesmon , Rabbit anti-CD31 , Donkey anti-mouse Alexa 647 , and Donkey anti-goat Alexa 546 . Images were taken with a Leica DM5500 microscope  and with a LSM 700 microscope . Alpha smooth muscle actin (α-SMA) expression was quantified with Fiji imaging program (ImageJ) as previously describedStatistical analyses were performed using Prism v5.0a (GraphPad), using the test mentioned in the figure legends. A p-value of less than 0.05 was considered significant. All error bars in diagrams represent the standard deviation (SD).The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.By applying a fast compression step, resulting in a less than 50% liquid loss followed by a slow air-drying step, we could control the liquid content of our produced grafts and therefore also the collagen grafts final density Fig. . This prThe LD-grafts were associated with a fistula formation rate of 15.4% (2 out of 13 rabbits) compared to one of 33% (3 out of 9 rabbits) in the HD-graft Fig. . No uretLD-grafts were implanted in 5 rabbits for 9 months and 2 rabbits for 11 months. No complications were noted in these rabbits Fig. . At 9 anin vivo, we had to develop two fabrication methods to produce both a polarized low- density and a uniform high-density collagen graft. Our rabbit results show that the low-density graft had an improved smooth muscle regeneration compared to the implanted high-density graft.In order to answer the question if collagen fiber density of a graft influences the urethral regeneration 10. The more extracted fluid from the collagen gels, the stiffer and more mechanical strong the collagen material gets. We experienced the same for our two grafts, where the surgeons reported that the HD-graft was more mechanically strong compared to the LD-graft. The HD-graft was designed to serve as a control, to represent a dense and stiff matrix (approximately 0.1 MPa), although it was not in the similar high range as the well studied SIS grafts 11, it was still two times stiffer than the LD-graft. Importantly, the SIS is reported to have been tried with patients for urethral reconstruction but has failed and is therefor not recommended to use in urethral reconstruction12. According to literature the native rabbit urethra seems to have a stiffness of approximately 0.2 MPa, which is more in the range of our engineered scaffolds as compared to the SIS urethral scaffolds13.Previous studies have shown that fluid content or collagen fiber density relates to the stiffness and influences material properties of the collagen gels12. However, when these graft are seeded with cells they have shown promising result in pre-clinical models but ultimately only limited success in clinical applications17. In our graft implantation study in rabbits, the LD-graft exhibited more ingrowth of smooth muscle cells with formation of muscle bundles, due to its less dense collagen structure or, whereas the HD-graft demonstrated faster urothelial regeneration due to its denser collagen structure. This confirms similar reports where denser/stiffer grafts have been implanted in animals and showed good urothelial regeneration but has lacked an adequate smooth muscle regeneration18. The accelerated ingrowth of smooth muscle cells into the low-density grafts is likely due to the lower surface area of collagen that needs to be degraded and remodeled by the infiltrating cells. Examining closer the HD-graft, it showed limited smooth muscle cell ingrowth in the center of grafted areas, but a normal muscle bundle structure was observed at the interface of the graft and native tissue (Supplementary Figure We would like to challenge the current opinion in urethral reconstructive surgery that a cell-free graft is ineffective to bridge a urethral defect, and to rephrase this statement to: A cell-free graft, which is too dense or stiff, is ineffective for urethral defect grafting. Published studies have already demonstrated the failure of using stiff cell-free grafts in this clinical indication19. By 6 months, the LD-graft had significantly more expression of α-SMA compared to the HD-grafts. At 11 months, cystourethrography showed urethral patency and normal urine flow and bladder voiding. Successful mating resulting in progeny could also be seen proving clinical relevant functional regeneration of the grafted areas.Improved smooth muscle regeneration is an advantage, as the primary pathology of urethral stricture is due to muscular fibrosis21. Although the grafts were prepared under sterile conditions, the production needs to be transferred into a GMP setting to allow translation of this technology to human application. This translation is allowed if the bioburden is lower than the sterility assurance levels set by governing authorities. To further reduce the bioburden, final sterilization can be attempted, however, it is known that sterilization can change the material properties of the grafts which might compromise the final outcome of the graft22.Our study has limitations. The rabbit model used had an artificially created urethral defect, with healthy urethral tissue on the edges. The graft was implanted to replace a totally excised urethral segment, what is not always mandatory as in clinical practice only partial circumferential replacement is often performedAn off-the-shelf urethral graft without the incorporation of cells and growth factors, showing adequate urothelial and smooth muscle cell ingrowth into the implanted graft, represents an important pre-clinical breakthrough. This was demonstrated by the comparable encouraging results of experimental studies conducted in two research centers in Switzerland and Malaysia opening the way for clinical translation of this cost effective, cell-free grafting for application in urethral reconstruction.Supplementary material


Autologous platelet rich plasma (PRP) has significant benefits facilitating improved graft take on wound beds due to hemostasis, adhesive and healing properties. This study aimed at effective use of PRP in wound beds on graft take irrespective of etiology as compared to conventional methods of mechanical fixation using sutures and staples. Forty cases including 20 in control and 20 in PRP groups admitted to the Department of Plastic Surgery at Vydehi Institute of Medical Sciences and Research Centre, Bangalore were enrolled between October 2015 and September 2017. Freshly prepared autologous PRP was applied on wound beds in the treated group, while conventional mechanical fixation methods like staples and sutures were used in the control group for the fixation of the skin grafts. Most significant result was the instant graft take to the wound bed irrespective of the etiology besides hemostasis and healing properties in the PRP treated group which resulted in considerable reduction of surgeon’s time required for the removal of sutures and staples at the final stages. Also, only 10% with graft edema were noted in the PRP treated patients as compared to 68% in the control group. The inner dressings and skin graft were dry in the PRP group and the post-operative etching, weeping and pain at the graft site reduced. The cosmetic appearance of this scar was better in the PRP group besides post-operative edema and graft loss. The study recommends use of PRP at the recipient site of split thickness skin graft. Platelets secrete stored intercellular mediators and cytokines from the cytoplasmic pool and release their α-granule content after aggregation. Also, cell proliferation, angiogenesis and cell migration are stimulated resulting in tissue regeneration. The anti-inflammatory and angiogenetic effects related to the other properties of platelets are well known.1-3Split thickness skin graft is generally done for soft skin coverage in view of its broad application for use due to ease of harvest. The healing process is undertaken through three stages of anchorage,4 The addition of PRP to SSG recipient sites seems to enhance primary healing and reduce healing time, likely as a result of shearing force reduction and enhancement of the wound environment with growth factors.5-7The application of autologous platelet rich plasma (PRP) to the split thickness skin graft sites is considered and theorized to provide immediate skin graft anchorage, as well as inosculation of the split thickness skin graft (SSG) with nutrient rich blood media. This study was performed to report the time to >90% primary healing of SSGs augmented with application of PRP in a high-risk patient population  parameters in both groups i.e. hematoma, discharge from graft site with significant graft loss, (ii) graft edema, (iii) frequency of dressings, (iv) duration of stay in plastic surgery unit and (v) cost effectiveness. In the PRP group, autologous PRP was topically applied on wound beds for graft anchorage, whereas in the control group, the graft was applied and fixed using conventional methods of suturing of staples. Autologous PRP helps to achieve stable hemostasis as it mimics the final steps of coagulation cascade. It brings about instant adhesion of graft to bed preventing any collection under the graft or undue shear.Randomized control study in 40 patients during 24 months from October 2015 to September 2017 was carried for obtaining the necessary ethical clearances. Also, written informed consent was obtained from all the patients included in the study and they were divided into two distinct groups of 20 each and the inclusion criteria were (i) Patients with acute and chronic traumatic wounds, (ii) Infective wounds, (iii) Post-burn wounds, (iv) Wounds following release of post-burn and post traumatic scar contractures, and (v) Co-morbidities like diabetes and hypertension and those on aspirin analogues. Patients who were positive for HIV, HbsAg and HCV and those with coagulation disorders and malignancy were excluded. A randomized procedure was adopted for the 40 patients and was divided into two equal groups of 20 each.In order to prepare the PRP, aseptic procedures were adopted for drawing the blood preferably from femoral vein alternatively using two 10 ml syringes containing anticoagulant before transferring to 10 ml vacutainers containing 1 ml citrate phosphate, distros-adenine (CPD-A) anticoagulant (freshly obtained from blood bank). PRP was prepared by a double centrifugation process. Whole blood was withdrawn from the patient in 5 ml plain vacutainer tubes. Four milliliter of 3.2% ACD-A anticoagulant was added per vacutainer. The PRP was extracted by a double centrifugation process using a REMI R-303 Table Top Centrifuge. The first ‘hard’ spin was given at 3000 RPM for 20 minutes. After the first spin, the whole blood was separated into 2 layers. The RBCs were collected at the bottom and the plasma buffy coat layer present above was aspirated using a wide bore needle.The transferred plasma was given a second ‘soft’ spin at 1000 RPM for 10 mins. Following this centrifugation, the test tubes showed a collection of platelet pellets with a few RBC at bottom of the tubes with the lower one-third containing platelet rich plasma and the upper two-third containing platelet poor plasma (PPP). The superficial two-third PPP was discarded and the PRP is extracted into a plain test tube. The remaining contents of the tube were discarded 0.05 mL/1 mL calcium chloride was added to the test tube containing the PRP which acts as a clot activator. The PRP was prepared just before the patient was being put on the operating table, since the shelf life of freshly prepared autologous PRP is approximately 2 hours . After the wound beds were prepared, topical application of the harvested PRP was done and the SSGs were placed over the recipient site. The graft appeared to relatively adhere itself to the ulcer bed within a span of 3-5 minutes. This was confirmed by moving the graft with a finger . Once thIn order to compare the effectiveness of PRP with that of the suturing for fixing the SSG, the raw area under the neck and the right forearm were prepared for the placement of SSG harvested from the thigh with which the neck was covered with the SSG and fixed with sutures, while in the lower part of the neck, the SSG was placed after the PRP application for fixation. This procedure helped in the comparison of the effectiveness of PRP in the fixation of the graft with that of mechanical fixation using suturing .Regarding the control group, the wounds were surgically debrided, hemostasis was secured and lavage was done for both the selected groups. In the PRP group, application was done topically on the wounds beds through the cannula from the syringe and instant anchorage of skin graft to wound bed was confirmed by moving the graft on the bed with the finger, which was performed by the assistants who were blind to the study. In the control group, sutures or staplers were used to secure the graft to the wound margins and bed -6. In thAs per the standard practice, the inspection of the graft was carried out during the early post-operative period taking note of the wetness of outer dressing, odor and pain in both the groups. The identified parameters like hematomas and discharge from the graft site, the frequency of discharge from graft site with significant graft loss, graft edema, the frequency of dressings and duration of stay in Plastic Surgery Unit were noted. The number of days of stay in the hospital was specifically noted for both the groups particularly to determine the advantages of using PRP towards the hospital stay, if any, and tabulated. In our study, the patients were followed up for a period of 3 months from the date of discharge to assess any scar hypertrophy in the early post-operative period. In order to undertake statistical analysis of data, SPSS software , in each of the groups were used. The number of patients indicating instant adhesion advantage, graft edema, discharge from graft site, hematoma with significant graft loss and scar hypertrophy were calculated in each group of patients. Hematoma formation, discharge from graft site formation, graft edema, and graft loss between the groups were matched using the Chi square test. Frequency of dressing and duration of stay between the groups was matched using the sample t test. For comparison, computations were done between control and PRP group with regard to day of first graft inspection, frequency of post-operative dressings and stay in plastic surgery unit. Difference in proportion between two groups was tested through Chi-square test. P≤0.05 was considered for statistical significance.In p<0.001) with the average age of 38.4 years in the control group and 32.5 years in the PRP group (p<0.001). Most interesting part was the duration of stay in which the PRP group had an average duration of stay of 15.25 days as compared to 17.3 days by the control group leading to savings both for the patients and for the hospital in terms of expenditure and satisfaction of the patients. The frequency of change of dressing was 2.75 in comparison to 8.15 with the inner dressings being observed to be significant dry in the PRP group (As seen in RP group . p<0.05). The edema was much reduced in the PRP group, as compared to the control group (p<0.05). With regard to the post operative graft loss, the PRP group significantly reduced the number of losses and the post operating morbidity (p<0.05). The p<0.0001). The total expenditure for the PRP group was around 15%, while that of the control group was 85% (The post operative hematoma at the donor site reduced post operative morbidity in the PRP group (orbidity . The Vanorbidity . The pos was 85% .13 and more than 800 proteins and bioactive factors.11 The results clearly showed considerable benefits including hemostasis, speeding up of operating time, frequency of postoperative dressings and the time of the surgery. In our study the preferred anti-coagulant used was CPD-A, because of the fact that the citrate binds calcium to create anti coagulation.13The entire procedure was aimed at application of platelet rich plasma in surgery prior to resurfacing with skin graft for facilitating instant and stable adhesion of the SSG to the wound bed without any mechanical fixation. The PRP normally has a platelet concentration of about 10,00,000 platelets/µl.14-17 In order to activate the platelets, calcium chloride was used in the preparation of the PRP for the activation, degranulation and release of growth factors. It was noted that platelet activation plays a major role in exocytosis, cytoplasmic α degranulation with significant burst of growth factor, platelet, derived growth factor, epidermal growth factor, platelet derived growth factor as well as vascular endothelial growth factor.18-20In order to increase platelet activation and the coagulation system in our studies, the blood was drawn immediately prior to the surgery. The supernatant plasma as well as the buffy coat containing high platelet concentration was used in our study taking into consideration the fact that the platelet concentration is always more than in the white blood cells. It was also noted that the activation of the platelet increases in anti-inflammatory cytokines in the presence of hepatocyte GF.21-23 The most noteworthy part of the study was that in all the patients treated with PRP, there was instant adherence of the skin graft to the wound bed contrary to the controlled group, where this was absent. The major advantage was the adhesive nature of PRP,23 whereas we had to use sutures and staples in the control group. The instant fixation of the skin graft without suturing or stapling resulted in considerable reduction of surgeon’s time required for the removal of sutures and staples at the final stages. In our PRP treated patients, we observed only 10% with graft edema compared to 68% patients in the control group, which had graft edema for more than a week and the most noticeable fact was that the inner dressings as well as the skin graft was observed to be dry in the PRP group. In our study PRP was used in all etiological groups and it was found to yield remarkably good results as it is well known in the various phases of SSG and plasmatic expiation in the first 24-48 hours, second stage of capillary in growth and the last stage of revascularization taking note of the fact that the grafts viability is critically dependent in the first two phases of the graft take.2,22 In the PRP group, it was noted that about 1 % of the patients developed hematoma under the graft, requiring secondary grafting as compared to 15% in the control group. In the PRP group, there was a significant reduction in the number of dressings within the first 15 days as compared to the control group which had to undergo frequent dressings within the initial 15 days. This resulted in cost saving in the dressing and the time of the nursing staff and the surgeon and is of considerable advantage in a plastic surgery unit. Our study showed that in the PRP group, scar hypertrophy was not seen mainly due to quick adhesion, less of graft edema as well as the collection under the graft.23-29The stage of capillary inosculation and early circulation reduced graft edema with application of PRP The hospital stay of the PRP group was 11 days as compared to the control group which was 17 days resulting in significant cost saving. All the parameters which were studied in our set up were found significantly different between the PRP and the control group. It is considered beneficial in all the aspects with regard to the patient and the surgeon using autologous PRP in grafting, while having considerable practical benefits of effective graft take on the wound beds.The authors declare no conflict of interest.


Their chain-length distributions (CLDs) varied with longan cultivar; Chuliang showed a larger proportion of longer amylopectin chains with a degree of polymerization (DP) 30~100. This is attributed to the slightly higher relative crystallinity of Chuliang longan seed starch. Apparent differences were also detected in amylose structure. Caopu showed a higher amylose content than Chuliang and Shixia, resulting in its lower gelatinization temperatures and enthalpy change. All longan seed starch had a typical A-type crystal structure with relative crystallinity ranging 28.6–28.9%. For raw starch, Caopu showed the lowest digestion rate, followed by Chuliang; Shixia showed the highest. This is because Caopu had the highest amylose content. Chuliang had a more intact structure than Shixia, as suggested by its higher crystallinity, although they had similar amylose content. After being fully gelatinized, all starch showed a similar digestion process, indicating that the digestibility of gelatinized starch does not differ with starch source or structure.Starch was isolated from longan seeds of three widely distributed cultivars  in China. Comparisons of the multi-level structure of the starch of longan seeds among various cultivars were made, and the relations between these structural and property characteristics are discussed. The isolated starch, accounting for 44.9–49.5% ( Dimocarpus longan Lour.), belonging to the Sapindaceae family, is a prevalent fruit widely distributed in subtropical areas such as China, Thailand, Malaysia, and Vietnam [Dimocarpus longan cv. Chuliang, Shixia, and Caopu are the predominant ones cultivated in China. Longan seeds are normally considered as agricultural waste in longan fruit processing, resulting in incredible resource waste and environmental pressure on a large scale. In recent years, more and more attention has been directed to longan seeds because they contain several bioactive compounds, including polysaccharides, phenolic compounds, and flavonoids, etc., and exhibit multiple functional properties, such as antioxidant, anti-inflammatory, anticancer, and anti-tyrosinase effects [Longan  linkages as backbones and by α-(1 → 6) linkages at branching points. As a complex macromolecule, starch has a multiple-level structure . In addiIn this study, seeds from three widely distributed longan cultivars  in China were used as raw materials. Starch was isolated from the longan seeds and compared with normal maize starch. The multi-level structure of the starch, including granular, crystalline, and molecular structures, was characterized and these structural features were used to explain differences in their gelatinization and in vitro digestion properties. The results of this study will provide guidance on the application of longan seed starch in food and non-food products, and thus will be beneficial for the development of the global longan industry.w/w) of starch, 6.0–8.1% (w/w) of protein, and 2.0–3.4% (w/w) of fat. The starch content among the longan seeds from the three cultivars was not significantly different, indicating that the genetic background and growth environment of longan does not have a significant influence on the accumulation of starch in longan seeds. In addition, significant differences were observed in the protein and fat content among the three cultivars: Shixia showed the highest protein content while Chuliang had the highest fat content.The content of starch, protein, and fat in longan seeds from three longan cultivars are summarized in The morphology of the starch granules isolated from the longan seeds, as compared with normal maize starch, was characterized using scanning electron microscopy (SEM), with images shown in θ, which is the same as the crystalline type of normal maize starch [The crystalline structure of starch was analyzed using X-ray diffraction (XRD), and the diffraction patterns are displayed in e starch . The relde(logX) in The chain-length distributions (CLDs) of all the starch samples were determined by size exclusion chromatography (SEC) after fully debranching the side chains of starch using isoamylase. The results are expressed as the SEC weight distribution of glucan chains from debranched starch as a function of DP (X), denoted as w1 chains) and longer amylopectin chains that span at least two lamellae (B2 and longer chains), respectively [As can be seen from ectively . Small dectively . The higThe amylose CLDs were different among the three longan cultivars. The highest amylose content was detected in Caopu (30.1%), followed by normal maize starch , peak (Tp), and conclusion (Tc) temperatures and enthalpy change (∆H) of seed starch from three longan cultivars were determined using a differential scanning calorimeter (DSC), and the results are summarized in o, Tp, and Tc) of the seed starch, with Chuliang showing the highest gelatinization temperatures and Caopu exhibiting the lowest. The enthalpy changes of longan seed starch showed a similar trend. Chuliang and Shixia displayed higher enthalpy change values  than Caopu (13.5 J/g).Starch gelatinization corresponds to the dissociation of amylopectin double helices from a semi-crystalline structure to an amorphous conformation. The gelatinization temperatures, including onset  were not correlated with the amylopectin CLD nor the crystallinity of the starch. Therefore, differences in the gelatinization temperatures of the three longan seed starch samples most likely resulted from variations in the amylose content. This is in accordance with previously published results showing that starch gelatinization temperatures and enthalpy change are negatively related to amylose content [Starch gelatinization properties could be influenced by multiple factors including the granule morphology, crystalline structure, amylose content, and molecular structure of the starch, etc. . In the  content ,18. In a content .The isolated longan seed starch samples were enzymatically digested by pancreatin and amyloglucosidase in vitro to stimulate their digestion process in food. Although starch is normally consumed after cooking, it sometimes cannot be fully gelatinized because of factors such as food processing conditions or lack of water in food systems, etc. Therefore, the in vitro starch digestion properties of longan seed starch, as compared with normal maize starch, were evaluated in both raw and freshly gelatinized states in this study. The resulting digestion curves are exhibited in As shown in −3 min−1) might be attributed to its higher amylose content  than Caopu, although its amylose content (29.1%) was similar to Caopu. This is probably because of the porous structure of maize starch granules, while longan seed starch showed a smooth surface as shown in the SEM images displayed in −3 min−1) was similar to normal maize starch, whereas that of Chuliang (7.7 × 10−3 min−1) was significantly lower, although the amylose content of the Chuliang seed starch was similar to Shixia. This could be explained by the higher crystallinity of Chuliang (29.2%) compared with Shixia (28.6%), resulting in a more intact structure which is more difficult for enzymes to digest.The digestion mechanism of native starch has been reported to be quite complex as it is associated with several interconnected factors, including the granule architecture , crystalline structure , and molecular structure  . The sigIn terms of freshly gelatinized starch samples, much quicker digestion processes were detected, as shown in Normal maize starch was purchased from Tiancheng Corn Development Co. Ltd., Siping, China. Chuliang and Shixia longan seeds were collected from local orchards in Maoming Gaozhou  in July 2017, while Caopu longan seeds were from orchards in Chaozhou  in July 2017.Aspergillus niger), isoamylase (from Pseudomonas sp.), total starch (AA/AMG) assay kit, and D-glucose (GOPOD Format) kit were purchased from Megazyme International, Ltd. . Pullulan standards with known peak molecular weights were purchased from Polymer Standards Service (PSS) GmbH . Dimethyl sulfoxide  was purchased from Merck Co. Inc. . Other chemicals were reagent-grade and used as received.Pancreatin from porcine pancreas was purchased from Sigma-Aldrich Pty. Ltd. . Amyloglucosidase  assay kit. The crude protein content was estimated from the nitrogen content of longan seeds using a Kjeltec instrument  with a conversion factor of 5.18 . The cruw/v) overnight, after which they were ground into a slurry using a homogenizer . The milled longan seed slurry was filtered through a 106 µm screen. The residues remaining on the screen were collected and mixed again with sodium bisulfite solution. The mixture was homogenized and filtered again. These steps were repeated until the residue quality no longer decreased. All filtrate fractions were combined and then stirred for 1.5 h with a mixed solution containing NaCl solution (0.1 M) and toluene at a ratio of 9:1. The mixture was left to stand until the starch granules had settled at the bottom. The upper toluene layer and NaCl solution were removed. The NaCl solution/toluene steps were repeated until the toluene layer became clear. The isolated starch samples were washed with water and then with ethanol three times, followed by a drying step in a 40 °C oven.Starch was isolated from longan seeds following the wet-milling method described by Wang et al. . Longan Starch samples were spread onto a circular metal stub covered with double-sided adhesive tape and were coated with platinum by a sputter coater . Images of starch granules were acquired using a scanning electron microscope  under an accelerating voltage of 10 kV and at multiple magnifications .X-ray patterns were performed with an X-ray diffractometer  at 44 kV and 27 mA Cu-Ka radiation. Diffractograms were obtained from 3–45° (2θ) at a scan rate of 5°/min. Starch samples were equilibrated in a saturated relative humidity chamber for 24 h at 25 °C before analysis . The relv/v) solution (DMSO/LiBr solution) before being injected into the SEC system with DMSO/LiBr solution at a flow rate of 0.6 mL/min. Size separation of the starch chains was performed using a pre-, Gram 100, and Gram 1000 columns  at 80 °C. Amylose content of the longan seed starch was calculated from the CLDs of each starch as the ratio of the area under the curve of amylose CLD to that of the whole CLD curve.The CLDs of isolated longan seed starch were characterized using an Agilent 1260 Infinity size exclusion chromatography (SEC) system  with a refractive index detector  following a previously published method . Starch o), peak (Tp) and conclusion (Tc) temperatures were calculated using Pyris software .Starch  was weighed in a high-pressure pan and then distilled water (three times the weight of the starch) was added. The pan was immediately sealed and allowed to equilibrate at room temperature for 1 h. Each crucible was loaded in a DSC  and then scanned from 30 to 110 °C at a heating rate of 10 °C/min . The ent2, 0.49 mM MgCl2, and 0.02% NaN3) at 37 °C. Digesta (0.1 mL) was collected at 0, 30, 60, 90, 120, 180, 240, 300, 360, 420, 480, and 540 min, after which 0.9 mL of absolute ethanol was immediately added to deactivate the enzymes. In terms of cooked starch digestion, the isolated longan seed starch  was first gelatinized in 2 mL of distilled water by cooking in a boiling water bath for 30 min. After cooling down, each freshly gelatinized starch was digested using 0.25 mg of pancreatin and 12.5 μL of amyloglucosidase in a sodium acetate buffer solution following a procedure similar to that for raw starch, except that starch digesta (0.1 mL) was collected at 0, 20, 30, 60, 120, 180, 240, and 300 min.In vitro starch digestion of raw and cooked longan seed starch was carried out using pancreatin and amyloglucosidase following a previously described method with modifications . Raw lont is the percentage of starch digested at time t (min), C∞ is the estimated percentage of starch digested by the end of digestion, and k is the digestion rate coefficient [The digestibility of both raw and cooked starch samples was calculated from the content of glucose released during digestion. The digestogram was then fitted to first-order kinetics using logarithm of the slope (LOS) plot following the equation:fficient ,33.p < 0.05.Results were expressed as the mean and standard deviation of at least duplicate measurements. Analysis of variance (ANOVA) with the general linear model and Tukey’s pairwise comparisons were used to analyze data in Minitab 16 . Significant differences of the mean values were determined at Longan seeds of Chuliang, Shixia, and Caopu cultivars are rich in starch with contents ranging from 44.9–49.5%. The seed starch of the three longan cultivars had similar granule shape and size, with most granules having smooth surfaces and an oval or irregular polygonal shape. The CLDs characterized using SEC showed differences among the seed starch of the three longan cultivars, although the same crystalline type (A-type) was detected with the relative crystallinity varying from 28.9 to 29.2%. Chuliang longan seed starch had the highest proportion of amylopectin chains with DP 30–100, leading to its relatively higher crystallinity than the others. The starch also differed in its amylose structure. The amylose content of longan seed starch varied with longan cultivar, with Caopu exhibiting a significantly higher value (30.1%) than the Shixia (26.6%) and Chuliang (26.7%) cultivars. This contributes to the lower gelatinization temperatures and enthalpy change of Caopu than the others. For raw longan seed starch, the digestion rate differed significantly with cultivar, which is attributed to their distinct molecular and crystalline structures. After full gelatinization, however, the digestion profile of the seed starch of the longan cultivars became similar, suggesting that the digestion process remains the same no matter the starch structure, botanical source, or growth.


Dear editor, in which the authors presented their views about theimpact factor provided by the Journal Citation Report (JCR). However, I believe that thereaders of this journal should be aware that BJCVS is also indexed by another globalcitation database, Scopus. As JCR generates a list of journals indexed by Web ofScience, SCImago generates annually a list of journals indexed by Scopus, which aresorted by their SCImago journal rank (SJR) for each of its subject categories. Hence,according to data from SCImago, BJCVS is categorized as Q3 in three subject categories:“Cardiology and Cardiovascular Medicine”, “Medicine (miscellaneous)”, and “Surgery”.I read with great interest the latest editorial published by the Brazilian Journal ofCardiovascular Surgery (BJCVS)As the journal has been indexed by Web of Science since 2016, I extracted data publishedby BJCVS from Scopus for the same time period to establish other journal’sattributes.First, I searched for the documents published by other journals that cite articlespublished by BJCVS. I sorted the citing journals by the total number of BJCVS articlescited and determined to which quartile each of the journals belonged, according to theirSJR2017. As As the editorial mentions, most of the contributions accepted by the journal are writtenby Brazilian researchers from Brazil, followed at a distance by researchers from Turkey,China, United States of America, and India. However, a closer look at the country ofaffiliation of those researchers that use the information published by BJCVS revealsthat 480 articles were written by Brazilian researchers, 341 were authored byresearchers from United States of America, 191 were written by Chinese researchers, 89were authored by British researchers, and 80 were written by German researchers. Thisinformation is relevant to readers as it confirms the international visibility of thework being published by BJCVS..Perhaps it is time to begin using metrics that are more meaningful to theresearchers.Currently, many members of the research community are considering the use of new metricsto evaluate journals where to submit their manuscripts, following the guidelines set atthe San Francisco Declaration on Research Assessment, also known asDORA