The adsorption mechanism was found to be associated with π-π interaction, hydrogen bonding, and pore filling. Application of the innovative dynamic approach based on fast-field-cycling NMR relaxometry indicates that variations in the retention of water-soluble dyes could be explained by distinct water dynamics in the porous structures of WC and CC. The obtained results suggest that studied biochars will be more effective in adsorbing of cationic than anionic dyes from contaminated effluents.Biochars from wood chips (WC) and corn cobs (CC) were prepared by slow pyrolysis and used for sorption separation of erythrosine B (EB) and thioflavin T (TT) in batch experiments. Biochar-based adsorbents were extensively characterized using FTIR, XRD, SEM-EDX, and XPS techniques. The kinetics studies revealed that adsorption on external surfaces was the rate-limiting step for the removal of TT on both WC and CC biochar, while intraparticle diffusion was the rate-limiting step for the adsorption of EB. Maximal experimental adsorption capacities  In recent decades, there has been an enormous raise in the production and application of synthetic dyes. The global dye and pigment market size was estimated at USD 32.9 billion in 2020, and from 2021 to 2028 is projected to rise at a compound annual growth rate of 5.1% as expected by Grand View Research . In addiNon-specific treatment processes for dye-containing effluents are required because a wide range of different dyes are used in the dyeing of textiles over a short period of time, and the effluents are therefore highly variable in composition ,7. DespiThioflavin T  is a member of the cationic methine dye class, the most prominent group of methine dyes used extensively in the textile industry for the dyeing of polyacrylonitrile fibres . Erythro−1 vs. 97 MJ kg−1), lower global warming potential impact (−0.9 kg CO2eq kg−1 vs. 6.6 kg CO2eq kg−1), and lower environmental impacts than activated carbon. The authors further revealed that the adsorption capacity of biochar for heavy metals can compete with or exceed that of activated carbons. However, in the case of sorption separation of both anionic and cationic synthetic dyes, the adsorption capacities of biochars are extremely variable and vary from tens  is a Freundlich constant expressing adsorption capacity; maxQ (mg g−1) is the Langmuir constant, which gives the maximum adsorption capacity of the biochar with monolayer surface coverage, and b (L mg−1) is constant related to the affinity of biochar binding sites. lich (6)  and Langlich (6)  isothermR2 values for the Freundlich isotherm suggest that the adsorption of TT and EB on WC can be ascribed to Freundlich surface adsorption mechanisms . Both values of the exponent 1/n were less than 1, causing the isotherm curve to decrease at higher concentrations of EB and TT because the accessible sorption sites on the biochar outer and inner surfaces were occupied and fewer EB and TT molecules were adsorbed from the solution (maxQ) at 22 °C calculated from the Langmuir model for TT were 162 mg g−1 (WC) and 48 mg g−1 (CC), respectively. In the case of EB sorption, it must be emphasized that the calculated maxQ values (25.2 mg g−1 for WC and 7.5 mg g−1 for CC) were significantly overestimated compared to the experimentally determined maximum capacities maxexpQ (12.9 mg g−1 for WC and 1.6 mg g−1 for CC). Reflecting values of affinity constant b, both WC and CC biochars exhibited considerably higher affinity for TT (0.134 and 0.056 L mg−1) compared to EB (0.004 and 0.002 L mg−1).solution . The maxWe must note that there are only limited data for EB and TT adsorption by biochar-based adsorbents. However, comparing to other reported adsorbents of inorganic and organic origin , the ads3 g−1, and most of the volume of the CC was due to mesopores (0.019 cm3 g−1). Conversely, WC biochar is characterized by a uniform representation of micro- (0.021 cm3 g−1) and mesopores (0.021 cm3 g−1). Comparison of the original adsorption isotherms . As indsotherms  and the sotherms A,B showeWe also tried to apply an innovative dynamic approach based on FFC NMR relaxometry to understand the nature of the adsorbate–biochar interface interactions . The sorThe presence of crystallite phases such as sylvite, quartz, and calcite  and 44.6 ± 2.4 mg g−1 (CC), and EB 12.7 ± 0.9 (WC) and 1.5 ± 0.4 mg g−1 (CC), respectively indicating a higher affinity of biochars for TT. The pHpzc values of the WC and CC biochars obtained by the pH drift method were 8.3 and 8.5, respectively, and adsorption of both TT and EB was only slightly affected by the solution pH. The use of an innovative dynamic approach based on fast-cyclic NMR relaxometry suggests that substantial differences in the retention of water-soluble dyes can be explained by different water dynamics in the porous structures of WC and CC. Moreover, considering biochar characteristics, the π-π interaction, hydrogen bonding, pore filling, and interactions with mineral components were proposed to contribute to TT and EB removal by both adsorbents. In summary, the obtained results suggest that studied biochars will be more effective in separating cationic than anionic dyes from contaminated effluents.The adsorption properties of TT and EB on the biochars prepared by slow pyrolysis of wood chips (WC) and corn cobs (CC) were successfully investigated and compared. The kinetics studies revealed that adsorption on external surfaces was the rate-limiting step for the removal of TT on both WC and CC biochar, while intraparticle diffusion was the rate-limiting step for the adsorption of EB. Maximal adsorption capacities


Blueberries are concentrated with anthocyanins possessing antioxidant properties. As these properties counter fatigue, blueberry supplementation may improve performance and recovery, particularly in hypoxia, where oxidative stress is elevated.2 = 15.5 %) on separate days after supplementation with four days of blueberries (BLU) or four days of placebo (PLA). Heart rate (HR), oxygen saturation (SaO2) and ratings of perceived exertion (RPE) were monitored during the TT. Blood lactate and fraction of exhaled nitric oxide (FENO) were assessed pre-TT, post-TT, and during recovery.This study examined the effects of blueberry supplementation on running performance, physiological responses, and recovery in normobaric hypoxia. Eleven experienced runners completed a 30-minute time-trial (TT) in normobaric hypoxia . Pre-TT and post-TT FENO did not differ between conditions. Blood lactate recovery following the TT was similar between conditions.No significant differences were observed in the distance run during the TT, HR, SaOFour days of blueberry supplementation did not alter running performance or cardiovascular and perceptual responses in normobaric hypoxia. Supplementation lowered the blood lactate response to running, however, the significance of this finding is uncertain given the absence of an ergogenic effect. The elevated metabolic requirements of exercise accelerate the production of reactive oxygen and nitrogen species (RONS) . In smalConsidering the involvement of oxidative stress in exercise-induced fatigue and that higher levels of oxidative stress occur during exercise performed in hypoxia, strengthening an individual’s antioxidant defenses, thereby preventing an excessive accumulation of RONS, may have an ergogenic effect on performance. Nutritionally, supplementation with polyphenol-rich fruits is one way to enhance an individual’s antioxidant defenses , 8. PolyCurrently, the ergogenic effect of acute supplementation with polyphenol-rich fruits on endurance performance is unclear due to mixed findings . MoreoveiO2=15.5 %) and a graded running test to volitional exhaustion in normoxia to determine maximal oxygen consumption (VO2max). The last two visits served as experimental sessions. During each experimental session, runners completed a 30-minute TT in normobaric hypoxia. Heart rate (HR), rating of perceived exertion (RPE), and oxygen saturation (SaO2) were monitored throughout the TT. Blood lactate was assessed prior to, as well as 5, 15 and 30 min following the TT. Additionally, saliva and fraction of exhaled nitric oxide (FENO) were sampled pre- and post-TT. In the four days before each experimental session, participants supplemented with 4 days of blueberry powder (BLU) or 4 days of a placebo powder (PLA). For each participant, sessions were conducted at the same time of day (± 30 min).This study used a randomized, double-blind, placebo-controlled, crossover design with a 14-day washout period between experimental sessions. Participants visited the laboratory on three occasions. In the familiarization visit, participants performed the 30-minute treadmill time trial (TT) in normobaric hypoxia (F2max = 49.9 ± 9.3 mL·kg− 1·min− 1). All participants were non-smokers; free from cardiovascular, respiratory, and oral/gum disease; and free from musculoskeletal injury. None of the participants had been exposed to altitude  within the 3 months prior to the start of the study. Eleven recreational runners  volunteered for this study . Treadmill gradient was increased by 2 % every 2 min for the first 6 min of the test and then by 1 % every 1 min until volitional exhaustion. Oxygen consumption was measured continuously throughout the test using a gas analysis system . VOPrior to the start of each experimental session, participants rinsed their mouth with distilled water and were fitted with a HR sensor and chest strap . Participants were then moved into the altitude chamber  where they rested in a seated position for 10 min. During this time each participant’s food/activity log was reviewed to ensure compliance with study requirements.ENO and blood lactate were administered. Blood lactate was determined using a finger prick and handheld lactate analyzer . The Lactate Pro analyzer, when compared to laboratory-based analyzers, has been shown to be both reliable (Coefficient of variation [CV] ranging from 2.7 to 5.7 %) and valid  [ENO was assessed using a handheld electrochemical analyzer  following previously established guidelines[2, by fingertip pulse oximeter , were recorded throughout the TT while RPE  was recorded every 5 min.Afterwards, participants provided saliva samples and pre-TT measures of F - -1.7) –18. FENOuidelines. Once thuidelines. In expeuidelines. HR and ENO was assessed at 4 min post-TT, and blood lactate measurements were repeated at 5, 15, and 30 min post-TT. To assess lactate recovery, all post-TT values were normalized to the 5-minute post-TT value[Upon completion of the 30-minute TT, saliva was sampled, F-TT value.Saliva was collected to measure cortisol, uric acid (UA), interleukin-6 (IL-6), and C-reactive protein (CRP). Salivary biomarkers were measured due to the ease and non-invasive nature of sample collection. The biomarkers analyzed in the present study have been shown to be valid and reliable indicators of the physiological stress to running, 23. At In the four days leading up to each experimental session, runners ingested either freeze-dried blueberry powder (BLU) . Ingestion of the packets was distributed evenly throughout the day with the final packet ingested 2 h prior to the start of the session. For ingestion, the powder was mixed with about 250 ml of water. The antioxidant and anthocyanin content of the blueberry powder used in the present study Table  was asseDuring each 4-day supplementation period participants were asked to limit the intake of other foods/drinks high in polyphenols as well as refrain from taking other supplements and anti-inflammatory medications. Additionally, participants avoided caffeine and alcohol intake 8 and 24 h (respectively) prior to the start of each session. Participants also abstained from moderate to vigorous intensity physical activity within 24 h of each session. To facilitate this, participants completed a 4-day food and physical activity diary before the first experimental session and were instructed to replicate this diary before the second experimental session. 2, RPE, pre-TT salivary biomarkers, and pre- to post-TT change in salivary biomarkers were analyzed using a paired t-test. Differences in FENO and blood lactate were analyzed using a repeated measures ANOVA with 2 supplement conditions by 2 time points (1 pre- and 1 post-TT measure). Differences in blood lactate recovery were analyzed using a repeated measures ANOVA with 2 supplement conditions by 3 time points . Post hoc analysis was performed using t-tests with a Bonferroni correction. Statistical significance was set at p < 0.05. To estimate the magnitude of the effects from supplementation, Cohen’s d effect sizes (ES) were calculated with the magnitude of effects considered small (0.20–0.49), moderate (0.50–0.79) and large (> 0.80). Additionally, Pearson correlation coefficients were used to explore relationships between the study variables. Data are presented as mean ± SD. All statistical analyses were conducted using SPSS v 20.0 .Normality of the data was evaluated using the Shapiro–Wilk test. Mauchly test of sphericity was performed to assess the homogeneity of data. Where violations were present, Greenhouse–Geisser adjustments were made. Between-supplement differences in 30-min running distance, average HR, maximum HR, average SaO2, and RPE responses during the TT were not different between the two supplement conditions (Table No difference in the distance achieved during the 30-minute TT in normobaric hypoxia was observed between supplement conditions Table . Heart rENO levels were not different between the two supplement conditions (Table p < 0.001) with the increase in PLA being significantly greater than BLU (p = 0.036) (Table p = 0.02). A Cohen’s d showed a moderate effect size (d = 0.76) for the attenuated post-TT increase in blood lactate following BLU. Additionally, 8 of the 11 runners experienced less of an increase in blood lactate in response to BLU  (Table Pre- and post-TT Fns Table . In bothU p = 0.06 (Table U p = 0.06 (Table d = 0.68) for lower pre-exercise cortisol levels in response to BLU. The pre- to post-TT increases in cortisol and IL-6 were significant  with no differences between the two supplement conditions. For UA and CRP, pre- to post-TT changes were not significant.No significant pre-TT differences were observed in any of the salivary biomarkers between PLA and BLU Table . HoweverENO and the PLA vs. BLU difference in the pre-TT to post-TT increase in blood lactate concentration .A moderate negative relationship was observed between the PLA vs. BLU difference in pre-TT FThe present study examined the effect of acute blueberry supplementation on running performance and physiological responses in normobaric hypoxia. Four days of supplementation had no influence on 30-minute TT performance. However, although most of the physiological variables remained unaffected following supplementation, the post-TT increase in blood lactate was attenuated following blueberry supplementation.− 1 or ~ 15 % lower in BLU than PLA (Table iO2 = ~ 17 %) after supplementation with pomegranate extract[iO2 = ~ 15 %) were not altered after 7 days of supplementation with black currant extract[Following the 30-minute TT, blood lactate was 1.7 mmol⋅le extract. In contt extract. The cont extract, particit extract. Furthert extract. With thENO was used as an indirect marker of NO availability. To this end, significant increases in FENO have been observed In response to acute beetroot juice supplementation [ENO were elevated levels of plasma nitrite and nitrate (precursors of NO production); thus suggestive of elevated NO availability in the blood[ENO as nitrate-containing supplements, such as concentrated beetroot juice. However, four days of blueberry supplementation had no effect on FENO as evidenced by the similar pre-TT values. Consequently, it is uncertain if blueberry supplementation had any impact on NO availability, raising questions about the contribution of altered blood flow to the lower blood lactate response. Despite this, there was a moderate negative relationship  between the BLU vs. PLA difference in pre-TT FENO and the BLU vs. PLA difference in the pre-TT to post-TT increase in blood lactate. This suggests that those runners who experienced increases in FENO in response to blueberry supplementation (in comparison to PLA) tended to experience less of an increase in blood lactate following the TT. Thus, as with other forms of polyphenol-rich supplementation, this relationship indicates the physiological response to blueberry supplementation seems to be individualized (i.e. responders and non-responders)[Accounting for the improvements in endothelial function following blueberry supplementation is an increase in NO bioavailability . In the entation , 25. Accthe blood. Considethe blood, albeit sponders). Two facsponders), 30. In iO2=15.4 %)[iO2 = ~ 15 %)[iO2 = ~ 15 %)[2 to sustain the elevated metabolic requirements of the exercising muscle[2 delivery.The absence of any performance advantage following blueberry supplementation is in agreement with recent findings. In a simulated altitude comparable to that of the present study, 10-km treadmill TT performance in trained runners was not significantly altered following a single dose of concentrated beetroot juice F2=15.4 %). Similar= ~ 15 %). When co= ~ 15 %). Further= ~ 15 %). The com=15.4 %. To conclude, four days of blueberry supplementation attenuated the increase in blood lactate to a 30-minute running time trial in normobaric hypoxia. Despite this, there was no improvement in running performance. Consequently, the use of acute blueberry supplementation as an ergogenic aid for endurance performance in hypoxia is not supported.


Group B Streptococcus (GBS) is an important cause of invasive infection in neonates and infants. Cerebrospinal fluid (CSF) findings and culture may not show evidence of infection early in GBS meningitis. Next-generation sequencing (NGS) has the potential to detect microbial genetic material in patients with infectious diseases. We report two cases of infantile sepsis of GBS meningitis with negative results for CSF culture tests, but positive results for NGS analysis.Enterococcus faecalis was cultured from urine. He was discharged after a total of 1 week of antibiotic treatment. Six days after discharge, he was re-hospitalized for sepsis. Blood and CSF cultures were negative, and E. faecalis was again cultured from urine. He received a total of 3 weeks of ampicillin treatment for enterococcal-induced nephritis and did not relapse thereafter. NGS pathogen searches were retrospectively performed on both blood and CSF collected at the first and second admission. GBS was detected in the CSF collected at the first admission, but no significant pathogen was detected in the other samples. Inadequate treatment for GBS meningitis at the first admission may have caused the recurrence of the disease.Patient 1 was a 22-day-old male infant diagnosed with sepsis and meningitis. His CSF findings showed pleocytosis, decreased glucose, and increased protein levels. However, CSF and blood culture results at admission were negative. He received a total of 3 weeks of treatment with ampicillin and cefotaxime, and showed clinical improvement. GBS was detected through NGS analysis of CSF collected at admission. Patient 2 was a 51-day-old male infant with sepsis. CSF findings on admission were normal, and blood and CSF cultures were also negative. Intravenous ampicillin and cefotaxime treatment were initiated. Treatment was de-escalated to ampicillin alone because Infantile sepsis may present bacterial meningitis that is not diagnosed by either culture testing or CSF findings. NGS analysis for CSF may be useful for confirming the diagnosis of bacterial meningitis. Streptococcus agalactiae; GBS) is a leading cause of sepsis and bacterial meningitis in infants under 3 months of age. The estimated incidence of GBS infection in infants is approximately 0.5 per 1000 live births [Sepsis is one of the leading causes of death among children, and the percentage of all global deaths related to sepsis is highest in children under 1 year of age . Group Be births . GBS infe births  thus, acNext-generation sequencing (NGS) technology makes it possible to analyze large amounts of nucleic acid sequence data contained in samples in a single assay. Therefore, untargeted metagenomic NGS of clinical samples has been applied for the comprehensive diagnosis of infections, including viruses, bacteria, fungi, and parasites . We prevWe describe two pediatric cases of late-onset GBS meningitis with sepsis. They were born by vaginal delivery after uneventful pregnancies. Prenatal GBS screening through a vaginal swab was negative in both cases. Sepsis is defined as a systemic inflammatory response syndrome with an infectious disease. Systemic inflammatory response syndrome refers to four parameters: body temperature, tachycardia, hyperventilation, and white blood cell count. Specifically, it is diagnosed when body temperature is > 38.5 °C or < 36.0 °C, or leukocytosis or leukocytopenia is required, and a total of two or more parameters exist . Tachyca3/μL and a C-reactive protein (CRP) level of 12.0 mg/dL. A lumbar puncture was performed, and examination of the CSF showed pleocytosis , a decreased glucose level of 25 mg/dL, and an increased protein level of 326 mg/dL. He also underwent a sepsis evaluation, including blood and CSF cultures. Leukocytes and bacteria were not seen in Gram staining. He was empirically administered ampicillin (280 mg/kg/day) and cefotaxime (280 mg/kg/day) for bacterial meningitis according to the ESCMID guidelines [A 22-day-old male infant was seen in the emergency room of TOYOTA Memorial Hospital with a 7-h fever history to 38.6 °C and vomiting. He had a bulging anterior fontanelle and peripheral cyanosis. Initial blood examination showed a white blood cell count of 13.1 × 10idelines , and int3/μL) and a CRP elevation of 2.13 mg/dL. He also had peripheral cyanosis and was diagnosed with severe sepsis. He underwent a sepsis evaluation, including blood, CSF, and urine cultures. There was no leukocytosis in either the CSF or urine. He was initially administered intravenous ampicillin (300 mg/kg/day) and cefotaxime (300 mg/kg/day). On day 2 of admission, his vital signs normalized. Blood cultures and CSF cultures were negative, while Enterococcus faecalis (103 CFU/mL) was detected in urine cultures on day 4 of admission. He received ampicillin (200 mg/kg/day) for 1 week before discharge for enterococcal urinary tract infection. He was discharged after confirmation of a continued normal temperature, no abnormalities on physical examination, and improved blood test results. Six days after discharge, he returned to the hospital with a fever of 39.0 °C, tachycardia, and peripheral circulatory insufficiency. He underwent a sepsis evaluation with lumbar puncture and was empirically administered ampicillin (300 mg/kg/day) and cefotaxime (300 mg/kg/day) for sepsis. The CSF findings were within normal limits. No bacteria were cultured from the blood or CSF. A poorly enhanced area in the kidney was found using contrast-enhanced computed tomography (CT) on the day after admission. In addition, 103 CFU/mL E. faecalis was detected in urine culture, and ampicillin (200 mg/kg/day) was administered for enterococcal bacterial nephritis for 3 weeks. Voiding cystourethrogram revealed Grade 4 reflux findings on the right side strongly indicating bacterial nephritis. There was no recurrence of the disease. NGS analysis was performed retrospectively on serum and CSF collected at the first and second admission to detect pathogens. GBS was detected in the CSF collected at the first admission, but no significant pathogen could be detected in the other samples . DNA sequencing libraries were prepared using a Nextera XT library Prep Kit  according to the manufacturer’s instructions with slight modification . An AgilSequence data were processed using the metagenomic analysis pipeline PATHDET version 1.0  to detecPCR using the GBS-specific primers Sag59 (5′-TTTCACCAGCTGTATTAGAAGTA-3′) and Sag190 (5′-GTTCCCTGAACATTATCTTTGAT-3′) was performed for both clinical samples according to methods described previously .Infantile sepsis is occasionally accompanied by bacterial meningitis; however, meningitis is not easy to diagnose in some patients. Bacterial meningitis in the early stage of the disease onset of late-onset GBS infection tends to result in normal CSF findings and is negative for CSF culture testing due to the small number of bacteria . Since bNGS analysis of CSF is advantageous for diagnosing GBS meningitis in pediatric patients with negative CSF cultures. According to a study by Wilson et al. , NGS ideIn Case 2, the NGS result for GBS was positive in CSF; however, the result was negative in blood. A previous report stated that blood cultures are positive in at least half of patients with bacterial meningitis , 22. OthIn conclusion, we report two cases of GBS meningitis that were revealed retrospectively by NGS analysis. We also showed that GBS meningitis was lurking in pediatric sepsis negative for CSF culture testing with normal CSF findings. NGS may be useful as a diagnostic tool in the specific clinical settings presented in the present report.


Full dynamics of biofluid biomarkers have been unknown in patients with Parkinson’s disease (PD). Using data from 396 PD patients and 182 controls in the Parkinson's Progression Markers Initiative (PPMI) database, we estimated long-term temporal trajectories of CSF α-synuclein (α-syn), amyloid-β (Aβ), total tau (t-tau), phosphorylated tau (p-tau) and serum neurofilament light chain (NfL) by integrating function between the baseline levels and annual changes. At baseline, PD patients showed lower CSF α-syn, Aβ, t-tau and p-tau levels than those of the controls. In all PD patients, CSF α-syn and Aβ decreased in a negative exponential pattern before the onset of motor symptoms, whereas CSF t-tau and p-tau, and serum NfL increased. Patients with cognitive impairment exhibited faster decline of Aβ and α-syn and faster rise of t-tau, p-tau and NfL, when compared to those without. Similarly, low Aβ group showed earlier decline of α-syn, faster rise of t-tau, p-tau and NfL, and faster decline of cognitive performances, when compared to high Aβ group. Our results suggest that longitudinal changes in biomarkers can be influenced by cognitive impairment and Aβ burden at baseline. PD patients with Aβ pathology may be associated with early appearance of α-synuclein pathology, rapid progression of axonal degeneration and neurodegeneration, and consequently greater cognitive decline. Pathologically, greater amount of cortical and limbic Lewy bodies (LB), amyloid-β (Aβ) plaque and neurofibrillary tangles are related with the development of dementia in PD5. CSF biomarkers for each pathological protein may reflect the pathological burden in brain, and thereby, long-term changes in CSF biomarkers may be related with both pathological and clinical progression in PD.About 80% of Parkinson’s disease (PD) patients become demented in their clinical course with variable time intervals between the onset of motor symptoms and deterioration of cognition7, while others reported a decrease in CSF α-syn9. Likewise, contradictory results of the longitudinal changes in CSF Aβ1-42 were reported11. These studies observed the changes occurring in relatively short time period (from 18 months to 4 years) at different time points of disease course, and therefore, were insufficient for providing full dynamics of CSF biomarkers.However, several studies observed inconsistent results of longitudinal changes in CSF biomarkers. Some exhibited an increase in CSF α-synuclein (α-syn) along with the disease progression and an association of α-syn with further deterioration of motor and cognitive deficits1-42, total tau (t-tau), phosphorylated tau (p-tau) and serum neurofilament light chain (NfL) by integrating the function between baseline level and annual change rates13. In addition, we investigated effects of cognitive impairment and low CSF Aβ1-42 level on the temporal trajectories of CSF biomarkers.In the present study, using the longitudinal data for the biomarkers in the Parkinson's Progression Markers Initiative (PPMI) study, we estimated long-term temporal trajectories of CSF α-syn, Aβ1-42 , α-syn , t-tau  and p-tau levels  than those of the controls, while the serum NfL levels did not exhibit a significant difference. In addition, PD patients showed worse cognitive performance in MoCA and HVLT delayed recall . Lower CSF α-syn and t-tau levels showed a trend of association with higher MDS-UPDRS III total scores at baseline, however, generalized linear models did not reach statistical significance .Demographic characteristics, clinical measurement and biomarker levels at baseline are summarized in Table 1-42, 274 PD patients were classified as high Aβ1-42 group and remaining 122 patients as low Aβ1-42. PD patients with low Aβ1-42 showed higher frequency of APOE ε4 allele and lower baseline CSF α-syn and p-tau than those with high CSF Aβ1-42. Serum NfL and cognitive performances did not show significant differences between the two groups.311 PD patients showed MoCA total scores greater than 25 at baseline and were classified as PDNC. Although the PDCI group was older age and showed worse parkinsonian motor deficits at baseline than the PDNC, there was no clear difference in baseline biomarker levels between the two groups. According to the baseline CSF Aβ1-42, whereas serum NfL were increased. CSF t-tau and p-tau levels remained almost constant from the baseline to 3rd year of follow-up , Aβ1-42 , t-tau  and p-tau  than control subjects, whereas serum NfL did not exhibit a significant difference during study period.During the follow-up periods, entire PD patients showed an overall trend of decrease in CSF α-syn and Aβ1-42 showed lower estimated annual changes in PD group than control subjects. Estimated annual changes were positive under 839 pg/mL of baseline baseline Aβ1-42 and 1014.1 pg/mL in controls, and then converted into negative at higher levels  and smaller than those in controls  was lower than that of PD patients (2.0 pg/mL/year).  and α-syn  and faster increase of t-tau  and p-tau , disease duration, and interaction between groups and disease duration and subjects as random effect showed significant effect of interaction between disease duration and groups in progressions of CSF Aβ1-42 and, while the PDCU group did not. During the 30 years of symptomatic period, PDCI group showed much greater change in CSF Aβ1-42 (− 48%) than PDCU group , whereas estimated t-tau levels in PDCI group showed a consistent rise (increase of 123.0% during 30 years after the onset). Estimated CSF t-tau levels of PDCI group at motor onset was lower than those of PDCU and control groups, however, became higher 5 and 8 years after motor onset, respectively , t-tau  and p-tau levels , whereas serum NfL did not exhibit significant difference  than high Aβ1-42 group (0.1 years after the onset). As the trajectories of CSF α-syn approached to plateau, difference in CSF α-syn levels between two groups gradually decreased. However, low CSF Aβ1-42 group exhibited still lower estimated CSF α-syn level than that of the high CSF Aβ1-42 group  and 5 years (p-tau) after motor onset. Compared to trajectories in control group, CSF t-tau in PD patients with low CSF Aβ1-42 became higher about 19 years after motor onset, whereas p-tau levels showed similar increase during estimated 30 years. For serum NfL, PD patients with high and low CSF Aβ1-42 exhibited rapid increase of biomarker level compared to controls. In high CSF Aβ1-42 group, estimated serum NfL level increased up to 30.7 pg/mL (Z-score = 1.84) for 30 years, while low CSF Aβ1-42 group showed steady increase in serum NfL level up to 71.1 pg/mL (Z-score = 5.91) at 30 years after the onset  than that in high CSF Aβ1-42 group  than high CSF Aβ1-42 group  at 30 years after the onset  study7, whereas a contrary result was found in a study analyzing full dataset of the same cohort8. In PD without dementia, an increase in CSF α-syn level was observed only in the patients with disease duration longer than 5 years11. Moreover, contrary to our results, a previous analysis including 173 subjects PPMI database reported a significant increase in CSF Aβ1-42 during one year from baseline10. In our temporal trajectory models obtained from all PD patients in PPMI database, overall changes in CSF α-syn, Aβ1-42, t-tau and p-tau levels were only less than Z-score 1.0 during 30 years from the onset and more prominent in the patients with cognitive impairment and low CSF Aβ1-42. We suspect that the inconsistency between the previous studies was attributable to the relatively small long-term changes in CSF biomarkers and a heterogeneity of PD patients included. Indeed, a previous study including full dataset of PPMI demonstrated progressive decline of CSF Aβ1-42 during 3 years from baseline15. Therefore, a consistent and meaningful change can be expected by much longer observational period in a large number of PD patients with relatively homogeneous clinical features vulnerable to changes in CSF biomarkers.Previous studies investigating the longitudinal changes in CSF biomarkers in PD patients have reported inconsistent results18. Nuclear imaging studies showed that decrease in CSF Aβ1-42 and increase in CSF t-tau and p-tau were correlated with increased amyloid and tau burden in brain20. Therefore, our results suggest that PD patients with cognitive impairment at early stage can be a phenotype resulting from an early appearance of LB load and rapid accumulation of amyloid and tau pathologies. LB and amyloid plaque pathologies in cortical and limbic regions are highly associated with the occurrence of dementia in the clinical course of PD21, and the patients with PD dementia were more likely to show lower CSF Aβ1-42 and higher CSF t-tau and p-tau levels, when compared to those with normal cognition24. Similar to our results, PD patients with low CSF Aβ1-42 at early stage showed worse cognitive performance26 and faster decline of cognitive performance in memory, executive and visuospatial function than those with high CSF Aβ1-4228.Although the precise mechanism remains unclear, it was postulated that reduced CSF α-syn in PD may reflect altered dynamic of α-syn, possibly due to sequestration in intracellular LB pathology or enhanced extracellular clearance in α-synucleinopathy30. Axonal degeneration preceding neuronal death and damaged white matter integrity in cellular model and patients with Alzheimer's disease are, for instance, the evidences for an involvement of white matter in neurodegenerative diseases33. Thus, CSF NfL level can be a biomarker for neuroaxonal damage in various kinds of neurodegenerative diseases34. A recent study showed a significant correlation between CSF and serum levels of NfL in PD patients, suggesting that serum NfL also can be a potential biomarker for neurodegeneration replacing CSF NfL35. Although some studies did not find an increased CSF NfL in PD36, the other studies observed an increased CSF or plasma NfL level in PD patients compared to the healthy elderly37. In a large biomarker study including over 3,000 subjects with dementia, patients with late onset AD or dementia with Lewy body (DLB) showed higher CSF NfL than PD subjects30. Therefore, we may expect that additional amyloid pathology in PD enhances an increase of NfL in both CSF and serum. It is interesting to note that estimated changes in serum NfL was much greater than CSF biomarkers in our study. Moreover, PD patients suspicious for having amyloid pathology exhibited great amount  of increase in serum NfL level over the 30 years. This suggests that the NfL level may be a potential biomarker for neurodegeneration induced by amyloid pathology6 or monitoring the disease progression in PD37.NfL is a subunit of neurofilament protein, which is a major component of neuronal cytoskeleton and abundant in myelinated axons with large caliber1-42, t-tau and p-tau may be intercorrelated in PD patients. Postmortem studies showed an association between cortical LB burden, amyloid plaque and tau grade39. Greater postmortem cortical LB burden was expected by the lower antemortem CSF Aβ1-42 level40. In transgenic mice model expressing cortical amyloid, early appearance of widespread α-syn pathology can be induced by an injection of preformed α-syn fibril into the hippocampus41. Similarly, α-syn pathology promoted fibrillization and phosphorylation of tau proteins in cellular and transgenic animal models43. Several studies demonstrated a correlation between CSF α-syn, Aβ1-42 and p-tau levels45. Therefore, amyloid load in central nervous system of PD patients may cause not only neurodegenerative process related with Alzheimer’s disease, but accelerated propagation of α-syn pathology. As stated above, our estimated trajectory of serum NfL suggested a possible impact of amyloid burden on neuroaxonal integrity, and disruption of white matter integrity are related to deterioration of cognitive performance in working memory, attention and executive function, visuospatial skills, and psychomotor speed46. Our estimated trajectories are consistent with previous studies showing predictive value of CSF Aβ1-42 for global and domain specific cognitive decline15.It has been suggested that the dynamics of CSF α-syn, Aβ1-42 or cognition at the time point of baseline assessment in PPMI database. Because the patients had various disease duration at baseline, our study can be limited by the absence of synchronization of CSF Aβ1-42 and cognition levels in dichotomizing groups. In addition, we extrapolated biomarker levels and cognitive outcomes about 30 years of disease duration using longitudinal measurement of 5–7 years. Because the PPMI database only included the PD patients in their early stage of disease course, our study is limited by missing biomarker data for the far advanced PD patients and the changes in biomarkers might be underestimated. Therefore, our temporal trajectory models require to be further revised by including advanced PD patients.In this study, we stratified PD group either by CSF Aβ1-42 may be more vulnerable to LB pathology, neuroaxonal damage, and cognitive impairment in their clinical course. Our study results raised a possibility for the potential role of future amyloid-lowering therapy for delaying the pathological progression and cognitive decline in PD.In summary, our temporal trajectory models suggest that longitudinal changes in biomarkers can vary with amyloid burden or cognitive impairment in PD patients. Early occurrence of cognitive impairment can be a clinical feature suspecting more rapid growth of LB and AD type pathologies. PD patients with low CSF Aβhttp://ppmi-info.org), anonymized and de-identified results as of Feb 2020 were downloaded. Briefly, the PPMI study is an observational cohort study aimed at identifying biomarkers of PD progression. The eligibility criteria include PD patients older than 30 years of age, diagnosed with PD within the last 2 years with a Hoehn & Yahr (H&Y) stage not greater than II, untreated, and exhibiting striatal dopaminergic dysfunction on 123I-N-3-fluoropropyl-2β-carbomethoxy-3β-4-iodophenyl tropane (123I-FP-CIT) single photon emission computed tomography (SPECT). The detailed study protocol can be found at www.ppmi-info.org.Study design is summarized in Fig. 1-42, p-tau and serum NfL at least twice. Age-at-onset (AAO) was defined as the time interval between date of birth and self-declared motor onset, and disease duration as the time interval between the self-declared motor onset and sampling date. The severity of parkinsonian motor deficits was assessed by the Movement Disorder Society sponsored Unified Parkinson Disease Rating Scale (MDS-UPDRS) part III and H&Y stage. To estimate the effect of baseline CSF Aβ1-42 on longitudinal cognitive performance, we included total scores of Montreal Cognitive Assessment (MoCA), Letter-Number Sequencing (LNS) test, and delayed recall scores of Hopkins Verbal Learning Test (HVLT). In addition, genotyping of Apolipoprotein E (APOE) was included in the analyses. We classified PD patients into cognitively unimpaired  and cognitively impaired  groups47.We included PD and control subjects who had undergone measurement of CSF α-syn, AβThe PPMI study is registered at ClinicalTrials.gov (NCT01141023). Each PPMI site received approval from an ethics committee on human experimentation prior to study initiation. All research was performed in accordance with the relevant guidelines and regulations. The institutional review board of Pusan National University approved secondary analyses of publically shared data. The informed written consent has been waived off by the institutional review board of Pusan National University.http://ppmi-info.org) and elsewhere48. The coded frozen aliquots of CSF were transferred from the PPMI Biorepository Core laboratories to the University of Pennsylvania and to Covance for analyses. CSF Aβ1-42, t-tau and p-tau were measured using the electrochemiluminescence (ECL) immunoassays on a fully automated cobas e601 analyzer . The concentration of α-syn in CSF samples were analyzed using commercially available sandwich type ELISA kits , as previously described26. Serum NfL level was measured by the 2-step digital immunoassay using Single Molecule Array (Simoa) technology . In the present study, CSF α-syn samples with low hemoglobin (< 200 ng/mL)9 were included in the analyses. As proposed by a recent study analyzing PPMI database15, we used cut-off 683.45 pg/mL for deciding high and low Aβ1-42 groups. In addition, we excluded the PD patients with baseline biomarker levels more than 3 interquartile ranges below first quartile or above the third quartile.CSF was collected by standardized lumbar puncture procedures. Shipment and storage were performed as described in the PPMI biological manual  as predictor and biomarker or scores of cognitive function tests as responder variable. In case of control group, time interval from baseline (year) was used as predictor term. In addition, CSF biomarker levels and scores of cognitive function tests at the onset of motor symptoms were calculated using the linear regression model. Then, we acquired a curve for annual changes in each biomarker as a function of baseline values by using restricted cubic spline with 4 knots. The knots were placed at 5-, 35-, 65- and 95-percentile values of baseline levels. When sample size was under 100, we applied 3 knots at 5-, 50-, 95-percentile values. Estimated levels as a function of time in year were acquired by using the modified Euler’s method for solving the first order differential equation. Finally, based on the calculated biomarker levels at motor onset, we plotted trajectories of estimated CSF biomarker levels between the time points at 5 years before and 30 years after the onset of motor symptoms. To compare with control subjects, we also calculated Z-score using the mean and SD obtained from baseline values of control groups. In addition, we estimated temporal trajectories of biomarkers and scores of cognitive function tests in control group using same method. Calculated trajectories of control group were anchored to time axis (age) using mean values of biomarkers and cognitive performances and median age at baseline visit of PPMI study. For comparison between PD and control groups, we converted age into disease duration using following formula: disease duration = age – median of age at motor onset in PD patients.As described in the previous literaturesp < 0.05. Estimation of temporal trajectories and statistical analyses were conducted by custom script written in R software  with Hmisc and rms packages.Age, disease duration, education period and MDS-UPDRS III total scores were compared using independent t-test. Chi-square test was employed to compare categorical variables including sex ratio, APOE e4 allele frequency, and H&Y stage. Comparison of longitudinal biomarker levels and cognitive outcomes between groups were testing by linear mixed effect models using group, disease duration, and interaction between group and disease duration as fixed effects and subjects as random effect. Biomarker levels and cognitive outcomes at baseline were tested using generalized linear model covariated with age and sex. Statistical significance was defined as Supplementary Information.


Objectives: Zirconia (Y-TZP) ceramics are considered as posterior fixed partial denture (FPD) materials; however, their applications are limited due to chipping. The use of monolithic lithium disilicate (LiDi) glass ceramics in posterior FPDs can be advantageous. This in vitro study aims to compare the loads until failure of posterior Y-TZP-FPDs and LiDi-FPDs before and after aging. Ceramic materials have attracted increasing attention in dentistry and have emerged as an effective alternative for the fabrication of dental restorations in several applications . AccordiSilicate ceramics are suitable for single-tooth restorations in cases with high esthetic demands owing to their excellent optical properties; however, these ceramics do not have the high fracture strengths of zirconium dioxide ceramics, which are about 1200 MPa .2O:SiO2 ratio determines the crystalline structure of these materials, thereby indicating which LiDi or lithium aluminosilicate is formed from the raw materials [Therefore, the use of silicate ceramics is limited to veneers, onlays, and single crowns in the anterior region . Lithiumaterials . In LiDiaterials ,9,10. Hoaterials .For the frameworks of fixed partial dentures (FPDs), zirconia (Y-TZP) is considered as the preferred material, owing to its superior flexural strength ,12,13,14LiDi glass ceramics do not need to be veneered to imitate the naturalness of human teeth . Single-2SiO3 phase of LiDi-ceramics appears to dissolve more slowly than the glass matrix. This results in in the development of a rough surface [For FPDs comprising pressed LiDi, three-unit restorations up to the second premolar as a distal abutment are recommended . Fabrica surface . Additio surface . However surface , LiDi pr surface  and FPDs surface , do not  surface , or are  surface . TherefoEthics Statement:This article does not contain any studies with human participants or animals performed by any of the authors.Informed Consent:For this type of study, formal consent is not required. The authors have no conflict of interest and no funding to declare.Herein, for replacing the first premolars, 16 three-unit LiDi-FPDs (IPS e.max computer-aided design (CAD)) and 32 three-unit Y-TZP-frameworks  were fabricated ((computer-aided design/computer-aided manufacturing (CAD/CAM)). Eight FPDs per design group were artificially aged before all FPDs were loaded until failure. The data were analyzed using one- and two-way non-parametric analysis of variance and Tukey’s post hoc tests (α = 0.05).Herein, 48 stainless steel test models were fabricated with identical tooth abutments , representing a canine and second premolar A,B. The n = 16 FPDs per group): monolithic LiDi, conventionally veneered Y-TZP, and pressed-on Y-TZP. All of the FPDs were fabricated using a master FPD constructed of wax as a template.The following FPDs were fabricated and fitted to the test models forming three FPD-type groups (2. The dimensions of the connector between the canine and first premolar were 5.81 mm high and 3 mm wide (19.75 mm2). The surface area of the posterior connector was 21.81 mm2 (6.38 mm high and 4.37 mm wide). Further processing was performed in accordance with the manufacturers’ specifications. The monolithic restorations were milled  and crystallized at 850 °C  were fabricated using CAD/CAM technology A. The wa2. The frameworks were milled  and sintered for 2 h at 1450 °C.Zirconia frameworks were fabricated using CAD/CAM technology . The master FPD was scanned and virtually reduced by 1 mm to obtain the minimum thickness of the veneering ceramics . In accordance with the manufacturers’ instructions, the frameworks retained a minimum thickness of 0.7 mm and the connector dimensions were 19.75 and 21.81 mmHerein, 16 Y-TZP frameworks were conventionally veneered and glazed using a low-fusing nano-fluorapatite glass ceramic  A. The veAdditional 16 Y-TZP frameworks were veneered using the press-on technique B. The frAll the FPDs were cemented with glass ionomer cement  in a standardized manner  using a universal testing machine . Before cementing, the intaglio surface was airborne-particle abraded with 115-µm alumina at 0.15 MPa.Half of the FPDs in each design group were exposed to 1.2 million biaxial chewing cycles . To achieve this, the pontic was vertically loaded with constantly increasing force . A piece of tin foil (1-mm thick) was placed between the specimen and plunger (diameter: 4 mm) of the universal testing machine to avoid stress peaks. Failure was defined as a 10% drop in force during static loading , which coincided with the complete fracture of the examined restorations. Specimens showing complete fracture after only artificial aging were considered to exhibit a failure load of 50 N. If fracture gaps or the chipping of the veneering ceramic could have been visually or acoustically recognized, the determination of this incident was confirmed subjectively as an additional notice (data not shown).p ≤ 0.024), the collected data were analyzed using two- and one-way non-parametric analysis of variance (ANOVA) and Tukey’s post hoc tests [p = 0.257). The level of significance was set to 5% (α = 0.05).Statistical analyses were performed using an IBM SPSS 25.0 . As normal distribution could not be assumed , 1609 ± 427 and 1685 ± 480 N (p = 1.000) for e.max ZirPress-FPDs, and 1541 ± 438 and 1557 ± 643 N (p = 1.000) for e.max Ceram-FPDs. After aging, higher failure loads were observed for e.max ZirPress-FPDs than those for LiDi-FPDs (p = 0.042).The mean failure loads of LiDi-FPDs were 1293 ± 237 and 996 ± 516 N , while both artificial aging (p = 0.397) and the interaction of these terms (p = 0.511) were not statistically significant. In the pairwise tests, the failure loads for FPDs of one type did not differ in a statistically significant manner between test specimens with and without artificial aging (p ≥ 0.712). Considering the unaged FPDs, there were no statistically significant differences in failure loads between the different FPD types (p ≥ 0.677). However, for the artificially aged FPDs, statistically higher failure loads were found for pressed-on Y-TZP-FPDs than those for monolithic LiDi-FPDs (p = 0.042), while other comparisons in the group of the aged FPDs were again not statistically significant (p ≥ 0.262) (The failure loads of the FPDs are provided in ≥ 0.262) .All-ceramic restorations have excellent esthetic characteristics. Y-TZP-ceramics are high-performance ceramics that have been established for the framework of posterior FPDs. However, chipping is a major clinical problem when using veneered FDPs. CAD/CAM technology has made it such that monolithic LiDi-ceramics do not need to be veneered.In this study, three-unit Y-TZP-FPDs and LiDi-FPDs replacing the first premolar with and without artificial aging were compared considering their failure load in an in vitro model. The performance of posterior LiDi-FPDs was comparable to that of the veneered CAD/CAM-fabricated FPDs. The average load-to-failure values of the investigated LiDi-FPDs were 1293 N without aging and 996 N with aging, which is comparable to other results . MoreoveThe load-to-failure values for the pressed-on Y-TZP-FPDs ranged from 1609 (without aging) to 1685 N (with aging). The conventionally veneered Y-TZP-FPDs achieved results ranging from 1541 N (without aging) to 1557 N (with aging). This outcome is confirmed by other studies that have tested these types of FDPs in vitro ,43.Artificial aging is usually part of in vitro studies. The simulated aging scenario of a wearing period of approximately five years was used . The temArtificial aging had no significant influence on the tested FPDs. However, previous studies have determined different results. It is possible that a more extensive aging process would have a stronger influence on the samples. This could lead to significant differences. The load-to-failure values of LiDi-FPDs  were observed to decrease from 1832 to 410 N after undergoing chewing simulations . MoreoveThe test method chosen in this study—the elastic support of tooth abutments during aging and testing—and the use of a tin foil between the plunger and the occlusal ceramic surface during loading probably ensured a continuous increase in stress and minimized additional stress peaks. Compared to a rigid tooth abutment support and failure loading without an occlusal foil, the initiation of cracks can be reduced. In partial accordance with clinical use, this could explain the approximately constant fracture forces that are independent of the aging process. The influence of patient-specific conditions, such as occlusion and oral hygiene, should not be ignored considering real oral cavity conditions ,51.The present study verified that there were no significant differences regarding the load-to-failure between the two tested veneering techniques. According to cohesive fractures of the veneering (cracks and chipping), the over-pressed zirconia FPDs performed better than the conventionally veneered Y-TZP-FPDs . In this study, chipping did not occur for the examined Y-TZP-FPDs. Three LiDi-FPDs displayed cohesive fractures after chewing simulations in different fracture patterns . These findings are in contrast with those of previous in vivo studies ,25,48,52However, despite the fact that artificial aging has been used in this in vitro study, it can hardly imitate the complex structure of the oral cavity and a possible microbiological effect on the framework structure cannot be excluded. In addition, the sample size in this study was reduced to a statistically sensible minimum. Due to the explorative character of the present study, p-values should be interpreted descriptively. It is a weakness of this study that an a priori sample size calculation did not take place. Instead, a convenience sample was chosen for which statistically significant effects had been found in previous studies ,54,55,56In summary, FPDs constructed of LiDi glass ceramics performed well compared to veneered zirconia FPDs. The results of load-to-failure tests confirm the applicability of three-unit LiDi-FPDs for replacing first premolars.Within its limitations, this in vitro study indicates that the failure load of LiDi-FPDs may be sufficiently high for the prosthetic replacement of a first premolar. Conducting clinical trials to confirm this finding appears warranted.


This paper describes the education and case management profile of sports chiropractors with the Federation of International Sports Chiropractors (FICS) postgraduate qualification: International Chiropractic Sport Science Practitioner (ICSSP). The ICSSP is the predominant international sports chiropractic qualification.n = 240) in 2014. A cross-sectional survey, carried out between 22/10/2014 and 22/12/2014,was utilized with a 39-item web-based survey examining practitioner, practice and clinical management characteristics, and was distributed via email to all sports chiropractors who held an ICSSP qualification (n = 154). 36% of chiropractors were aged between 31 and 40 years, just over three quarters were male, and 27% had been in practice for 5–10 years. The majority of respondents were based in North America. All sports chiropractors surveyed reported treating neuromusculoskeletal conditions outside of the spine. 91% utilized a multimodal approach in most of their treatments, prescribing rehabilitative exercises in 76% of consultations. Almost 64% of respondents reported current treatment of professional athletes, and 78% reported current treatment of semi-professional athletes, whilst the vast majority of those surveyed endorsed past treatment of professional (91%) and semi-professional (95%) athletes. All respondents reported referring to a range of conventional and allied health providers.The survey response rate was 64% , and referring to and co-managing with other health practitioners.The online version contains supplementary material available at 10.1186/s12998-021-00405-1. The International Federation of Sports Chiropractic/Fédération Internationale de Chiropratique du Sport (FICS), founded in 1987, is comprised of national chiropractic sports councils worldwide and has affiliations with international organizations within the chiropractic profession and the world of sports. FICS has created a post-professional qualification program  for practitioners working at sporting events that consists of a minimum of an online educational component, a hands-on component, and a field-based experience requirement  but a 2009 survey of the United States general chiropractic profession by Christensen et al.. revealed that more than three-quarters of general chiropractors use passive adjunctive care procedures including ice packs, trigger point therapy, braces, and electrical stimulation . The higIn conjunction to passive modalities, rehabilitative exercises were prescribed by our sports chiropractors in 76%  of visits. Comparatively, previous studies have shown general chiropractors prescribing rehabilitation on a less frequent basis at 31% of visits, suggesting those working with active populations maybe more likely to prescribe exercises , 17. DesChiropractors are often labelled “spine only” due to reasons that range from self-promotion by chiropractors to misrepresentations by non-chiropractors –20. Our This survey also provides information on the self-education of sports chiropractors who are sometimes labelled as not conforming to evidence- based practice , 21. As Sports chiropractors are increasingly being used by elite sporting teams: the current NFL season saw 41 team chiropractors working with all 32 teams, and chiropractic services were offered in the Summer Olympic Games polyclinic in 2008, 2012, 2016 and 2020. As our study reveals, the vast majority of ICSSP chiropractors surveyed have at some stage in their careers treated professional  and semi-professional  athletes, with 20%  currently working with a sports team full time.There are those that hold the view that chiropractors do not integrate well in a main-stream multi-disciplinary team, nor do they interact with other professionals on a regular basis, as was shown in Busse et al.’s survey of orthopaedic surgeons . Whilst A strength of this study is that it attracted a good response rate of 64%, which is likely to be representative of the FICS based sports chiropractors , 27. WhiAdditionally, the survey was written in English and therefore limited to English speaking FICS chiropractors. However, despite the name FICS, the ICSSP/D qualification is only presented in English and therefore those who have completed it would be expected to be proficient in English.This preliminary study described the activities of sports chiropractors. Future research could investigate different geographical, practice and practitioner characteristics, comparing sports chiropractors at different levels of involvement  as well as describing in detail the treatment outcomes obtained by sports chiropractors.This study of sports chiropractors describes a small but well-educated workforce, that treats high-level athletes, manages a wide range of spine and non-spinal neuromusculoskeletal conditions, utilises a multimodal approach (including active and passive strategies), and refers to and co-manages with other health practitioners.Additional file 1: Web survey questionnaire.


Methodology for estimation of cerebrospinal fluid (CSF) tracer clearance could have wide clinical application in predicting excretion of intrathecal drugs and metabolic solutes from brain metabolism and for diagnostic workup of CSF disturbances.The MRI contrast agent gadobutrol (Gadovist) was used as a CSF tracer and injected into the lumbar CSF. Gadobutrol is contained outside blood vessels of the CNS and is eliminated along extravascular pathways, analogous to many CNS metabolites and intrathecal drugs. Tracer enrichment was verified and assessed in CSF by MRI at the level of the cisterna magna in parallel with obtaining blood samples through 48 hours.n = 29), both enrichment within CSF and blood coincided in time. Blood concentration profiles of gadobutrol through 48 hours varied between patients diagnosed with CSF leakage (n = 4), idiopathic normal pressure hydrocephalus dementia (n = 7), pineal cysts (n = 8), and idiopathic intracranial hypertension (n = 4).In a reference patient cohort  was preceded by far peak tracer enhancement at MRI in extracranial lymphatic structures (at about 24 hours), as shown in previous studies, indicating a major role of the spinal canal in CSF clearance capacity.The work was supported by the Department of Neurosurgery, Oslo University Hospital; the Norwegian Institute for Air Research; and the University of Oslo.  Methodology for estimation of cerebrospinal fluid (CSF) molecular clearance would be beneficial in several ways. Intrathecal administration of drugs is presently emerging as a promising route for treatment of a range of CNS diseases, including neuroinflammatory, neurodegenerative, neurooncologic, and neurovascular diseases –5. AlthoThe ability to predict clearance of endogenous molecules through CSF would also be crucial in assessment of neurodegenerative disease and dementia, and particularly in the preclinical phase of disease. Recent studies on animals and humans point at an instrumental role in extravascular clearance of metabolic waste products from CNS and its dependency on age and sleep , 8 for dHere, we studied the clinical feasibility and utility of estimating CSF tracer clearance from the craniospinal compartment using the gadolinium-based (Gd-based) MRI contrast agent gadobutrol as CSF tracer. Gadobutrol is a hydrophilic, stable compound hypothesized to be excreted from CSF along clearance pathways similar to those of other drugs and endogenous metabolites not being cleared at the BBB. After intrathecal administration of the tracer, we consecutively measured blood concentration of Gd over time in parallel with standardized T1 MRI sequences for assessment of tracer enhancement in CSF. The CSF tracer clearance to blood was assessed in a reference patient cohort (REF) with no identified CSF disturbance, and in individuals with CSF disturbances, including a cohort with the dementia subtype idiopathic normal pressure hydrocephalus (iNPH).This study included 52 individuals . In 29 phttps://doi.org/10.1172/jci.insight.147063DS1). For the REF group, tracer enrichment in CSF as shown by MRI (Following lumbar injection of CSF tracer (0.5 mmol gadobutrol), the tracer enriched CSF within the cisterna magna , but GFR was neither correlated with tracer enrichment in cisterna magna nor correlated with tracer concentrations in blood. The GFR is the most important measure for clearance function of the kidneys   with subsequent improvement of symptoms, tracer enrichment in cisterna magna was higher than in REF subjects after 1 and 2–2.5 hours , whereasWe established a 2-compartment model with first-order elimination from the central compartment; the model described the data adequately . The fint1/2,absorption) of gadobutrol to blood, the time taken for one-half of the administered dose to be absorbed from CSF to blood, was used to assess gadobutrol clearance from CSF to blood. In the reference group, mean t1/2,absorption was 6.3 ± 4.4 hours, and the maximum concentration (Cmax) 1.4 ± 0.6 μM was achieved after 9.8 ± 4.7 hours  may also need to be considered. Half-life of gadobutrol in blood is 1.5 hours .Some limitations should be noted. In this study, the amount of gadobutrol injected intrathecally was standardized to 0.5 mmol, regardless of fundamental patient characteristics such as dimensions of the craniospinal compartment, which may be a determining factor for CSF clearance capacity. Dose adjustments at individual level should be considered in future applications. The most optimal time points for retrieving blood samples could probably also be further sophisticated, both in the setting of suspected hyperaccelerated and delayed CSF clearance.It may be considered a limitation that intrathecal gadobutrol is used off-label. In our experience, intrathecal gadobutrol in a dose of 0.5 mmol is safe , 25. MorThe present blood concentrations of gadobutrol were measured in whole blood that was partly coagulated, which implied an extensive cleanup procedure involving homogenization of the samples. Gadobutrol is highly water soluble and eliminated from plasma through renal excretion, and it would be more convenient to analyze gadobutrol in whole blood samples with added anticoagulants or maybe in the plasma or serum fraction. Only a small amount of sample is needed (<0.5 mL) for gadobutrol analysis in blood. An optimization of the cleanup and analysis procedure may decrease the detection limit of the method even more. Bearing in mind that the amount of gadobutrol measured in the blood samples was at the higher end of optimal measuring range, this implies that the gadobutrol dosage needed for the purpose of CSF clearance assessment alone can be reduced significantly.It also needs to be resolved to which extent assessment of gadobutrol clearance from CSF to blood is valid as a surrogate marker to predict clearance along similar pathways for neurotoxic by-products of metabolism . In this respect, benefits of using contrast agents are several but are primarily their hydrophilic properties, stability in biological tissue, and ability to remain strictly confined to the outside of CNS blood vessels, thereby tracing extravascular CSF excretion pathways.Finally, this study includes selected groups of patients, examined for tentative CSF disturbances of unclear cause, who we presently have in our database with whole blood samples after intrathecal contrast-enhanced MRI. The mechanisms underlying symptoms in these cases are unclear. Moreover, this is most evident for individuals with symptomatic nonhydrocephalic pineal cysts (PCs), in which some may present with pathologic intracranial pressure (ICP) indices  and evenCSF tracer clearance assessment is clinically feasible and may provide a way to predict extravascular clearance of intrathecal drugs and endogenous metabolites from the CNS. The time course of tracer level in blood concurred well with that in CSF, which was estimated here semiquantitatively using MRI. The peak of tracer in blood preceded by far the peak of enhancement in extracranial lymphatic structures as shown in previous studies, indicating that the spinal canal has a major role in CSF clearance. Further studies are required to more precisely determine the clinical utility of the methodology and show how it differentiates different neurological diseases.Individuals who were candidates for intrathecal contrast-enhanced MRI were referred to Department of Neurosurgery, Oslo University Hospital, for tentative CSF disturbances. Patients who showed no evidence of CSF disturbance were denoted REF subjects. In these individuals, no indication for surgery was found. Patients with SIH who had a defined leakage requiring surgery to close the leakage were denoted as SIH. The diagnosis of iNPH was based on clinical, imaging findings, and results of ICP monitoring, as previously described , 62. AddNo patients with a history of hypersensitivity reactions to contrast agents, severe allergy reactions in general, or evidence of renal dysfunction were included. We also did not include pregnant or breastfeeding women or individuals under 18 or more than 80 years of age.The intrathecal injection was performed by an interventional neuroradiologist. Correct position of the spinal needle in the subarachnoid space was verified by CSF backflow from the puncture needle, and injecting of a small amount  of 270 mg L/mL iodixanol  confirmed unrestricted distribution of radiopaque contrast agent in the lumbar subarachnoid space. The MRI contrast agent gadobutrol  was given intrathecal in a volume of 0.5 mL (1.0 mmol/mL) at a speed of a few seconds. The study design with repeated MRI acquisitions and blood samples is presented in A 3 Tesla Philips Ingenia MRI scanner  was used, with equal imaging protocol settings at each acquisition to obtain sagittal 3D T1-weighted volume scans. The imaging parameters included: repetition time = “shortest” , echo time = “shortest” , Flip angle = 8 degrees, field of view = 256 × 256 cm and matrix = 256 × 256 pixels (reconstructed 512 × 512), and slice thickness 1 mm. The MRI exams were categorized as follows: 1, 2–2.5, 4–5, 5–6.5, 22–26, and 46–54 hours. Tracer enrichment in CSF of cisterna magna was assessed by placing region of interest within the CSF . To corrIntravenous blood samples were obtained and kept in a freezer at –80°C. The blood samples were thawed and transferred to a 15 mL polyethylene vial and homogenized with an Ultra-Turrax homogenizer (IKA T18). Blanks were regularly prepared with a similar procedure with 6 mL ultrapure deionized Milli-Q water to control for background contamination from the Ultra-Turrax. Aliquots of the homogenized blood samples (approximately 0.6 g) were then subjected to microwave-assisted digestion with concentrated ultrapure distilled nitric acid mixed with ultrapure deionized Milli-Q water (3 mL water and 5 mL nitric acid). The samples were digested in an UltraCLAVE single reaction chamber microwave oven (Milestone) according to a 60-minute stepwise heating program, where hold time for maximum temperature (250°C) was 15 minutes. After digestion, the samples were allowed to cool down to room temperature into their vessels before being transferred to 50 mL polyethylene vials (VWR) and diluted with deionized ultrapure water. Two certified plant reference material from the National Institute of Standards & Technology (NIST), 1515 apple leaves and 1547 peach leaves containing 3 and 1 μg/g Gd, respectively, were subjected to similar microwave-assisted digestion to assess the recovery of Gd. Mean recoveries of Gd in the NIST apple leaves and NIST peach leaves were 95% ± 4.1% SD and 103% ± 7.7% SD, respectively. The blood samples were analyzed for Gd by inductively coupled plasma mass spectrometry . Indium at 0.1 μg/L was used as internal standard. The samples were quantified using a 5-point standard curve (0.01–10 μg/L), and results were corrected for blank values. Representative detection limits for the analytics, estimated as 3 times the SD of 20 method blanks from 5 analytical runs, was 0.0009 μg/L. To ensure the correct quantification of the metals, 2 reference solutions of known element concentrations (0.1 and 1.0 μg/L) were analyzed in addition to in-house prepared standards. A deviation of 9% from the given concentration in the reference solution was accepted. Blanks were regularly analyzed to control for background contamination. The measured Gd concentrations in the blood samples were recalculated to gadobutrol concentrations.To accurately describe individual tracer (gadobutrol) pharmacokinetics, a population pharmacokinetic model was developed. A nonparametric adaptive grid approach implemented in Pmetrics (version 1.9.7) for R (version 4.0.0) was used . A totalt1/2,absorption (time to 50% of tracer dose absorbed from CSF to blood) was used as a measure of molecular CSF clearance to blood, and determined by dividing the natural logarithm of 2 by the model estimated transfer rate constant from CSF to the central compartment. Area under curve from 0 to infinity (AUC0-¥) was calculated with the trapezoidal approximation from the individual posterior predicted concentrations using the “makeAUC”-function” in Pmetrics.Posterior individual parameter values as well as posterior individually predicted concentrations obtained from the final population pharmacokinetic model were used for all pharmacokinetic calculations. The t test samples (2 tailed). For statistical analysis, we applied SPSS version 26 (IBM Corporation) or Stata/SE 15.0 (StataCorp LLC). A P value of 0.05 was considered significant.Repeated measurements were assessed with linear-mixed models using a subject-specific random intercept with an independent structure of residual errors at 1–24 and at 48 hours  using maximum likelihood estimation. We tested the difference between the groups at the different times after intrathecal CSF tracer with pairwise comparisons. The model-based parameters were compared between groups, using independent The study was approved by the Regional Committee for Medical and Health Research Ethics of Health Region South-East, Oslo, Norway (2015/96); the IRB of Oslo University Hospital (2015/1868); and the National Medicines Agency, Oslo, Norway (15/04932-7). Participants of this prospective and observational study were included after written and oral informed consent were provided from January 2017 to May 2018.PKE and GR conceptualized and designed the project. PKE and AL handled the blood samples. EM and HU performed the blood analysis. HC and MHH created the pharmacokinetic model. AHP, MHH, and PKE performed the statistical analysis. PKE and GR supervised and performed the administration and writing of the original draft. PKE, EM, UH, AL, AHP, BH, and GR wrote, reviewed, and edited the manuscript. All authors approved the final version of the manuscript.


Hypothermia to 33 °C did not result in a significant reduction in CSF outflow resistance compared with normothermia controls . Conclusions: Hypothermia treatment was protective in terms of ICP rise prevention, infarct volume reduction, and may be implicated in CSF outflow resistance post-stroke. Further investigations are warranted to elucidate the mechanisms of ICP elevation and hypothermia treatment.Background: Elevated intracranial pressure (ICP) occurs 18–24 h after ischaemic stroke and is implicated as a potential cause of early neurological deterioration. Increased resistance to cerebrospinal fluid (CSF) outflow after ischaemic stroke is a proposed mechanism for ICP elevation. Ultra-short duration hypothermia prevents ICP elevation 24 h post-stroke in rats. We aimed to determine whether hypothermia would reduce CSF outflow resistance post-stroke. Methods: Transient middle cerebral artery occlusion was performed, followed by gradual cooling to 33 °C. At 18 h post-stroke, CSF outflow resistance was measured using a steady-state infusion method. Results: Hypothermia to 33 °C prevented ICP elevation 18 h post-stroke (hypothermia ∆ICP = 0.8 ± 3.6 mmHg vs. normothermia ∆ICP = 4.4 ± 2.0 mmHg,  We have previously demonstrated that there is a damaging, transient increase in intracranial pressure (ICP) occurring 18–24 h post-stroke in rats ,4,5,6. EThe Monroe–Kellie doctrine ,12 descrCSF volume can be altered in the craniospinal space by two pathways; increased CSF production and decreased CSF outflow, both of which would result in an overall increase in the volume of CSF within the compartment. CSF volume is very difficult to quantify in vivo but we are able to determine insights into CSF dynamics by measuring CSF production or CSF outflow resistance  in real time. Previous work from our group has demonstrated that there is an increase in CSF outflow resistance in rats subjected to cortical stroke compared with sham rats 18 h post-stroke . A recenWe have previously demonstrated that ultra-short duration hypothermia, using clinically relevant cooling rates, prevents ICP elevation and reduces infarct volume 24 h post-stroke in rats . If the n = 32, Animal Services Unit, University of Newcastle, Callaghan, Australia) weighing 280–350 g were used for this study. Animals were housed under standard conditions in a 12 h light-dark cycle with unlimited access to food and water. All experimental procedures were in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes and were approved by the Animal Care and Ethics Committee of the University of Newcastle (A-2013-343 and A-2020-003).Adult male (11–12 weeks old) outbred Wistar rats (2:O2. Incision sites were cleaned and injected subcutaneously (s.c.) with 2 mg/kg 0.05% Bupivacaine . Body temperature was regulated throughout the surgery with a rectal temperature thermocouple . The femoral artery was cannulated with a catheter (1 and 2 French silicone tubing) for continuous monitoring of arterial blood pressure and heart rate. After stroke surgery and hypothermia treatment, rectal paracetamol  was administered for overnight pain relief. Saline injections  were also given to prevent dehydration and animals were returned to their cages with free access to softened laboratory chow and water.Rats were anesthetized with isoflurane  in 50:50 NDatalogger implantation was performed for continuous monitoring of core body temperature during experimental protocols and recovery. Implantation was performed according to previously described methods . To summCranial surgery was performed according to previously described methods . To summTransient middle cerebral artery occlusion (MCAo) was carried out according to our established protocol ,18. To sWe opted for the gradual cooling hypothermia paradigm as opposed to the traditional rapid cooling techniques that are common in pre-clinical literature. This choice was based on previous findings that show that clinically relevant cooling rates prevent ICP elevation and reduce infarct volume . Using tCSF outflow resistance was calculated using a steady-state infusion method ,20 with 3) = infarct volume ×  [Animals were euthanized at 24 h post-stroke. They were transcardially perfused with saline. Brains were removed and sliced into 2 mm sections on a rat brain matrix. Tryphenyltetrazolium chloride (TTC) staining was performed to confirm the presence of ischaemic stroke by identification of infarcted tissue. These same brains were then fixed in neutral-buffered formalin before processing for haematoxylin and eosin (H&E) staining for infarct volume quantification. Images were scanned using a high-resolution scanner  and infarct volumes were analyzed using the Aperio ImageScope program. Infarct analysis was conducted by an independent investigator blinded to treatment allocation. Infarct volumes were corrected for oedema by applying the formula: corrected infarct volume (mmt-test (unpaired) were used to compare differences between hypothermia treatment and normothermia control groups. Relationships were determined by Pearson r correlation. Data are presented as mean ± standard deviation (SD) unless otherwise stated. Statistical significance was accepted at the level of p < 0.05.Subarachnoid haemorrhage, equipment malfunction and lack of sufficient LDF drop at occlusion were pre-specified exclusion criteria. For outflow resistance measurements, lack of reliable ICP trace and complications during aCSF infusion that prevented ICP readings from at least 4 infusion rates, were also prespecified exclusions. Sample sizes were based on a similar study  which can = 4), abnormal animal physiology (n = 1), death during aCSF infusion (n = 1), poor oxygenation post-surgical intervention (n = 3), subdural bleed during cranial surgery (n = 1), surgical error during MCAo (n = 1), death at reperfusion with no clear cause detected at post-mortem (n = 1), subarachnoid haemorrhage confirmed at post-mortem (n = 2), overnight death with no apparent cause detected at post-mortem (n = 1), and ICP equipment malfunction (n = 2). One animal in the hypothermia group experienced a fever overnight, which may have contributed to an ICP rise in this animal. The animal was included in all analyses, as fever complication was not a pre-specified exclusion criterion. Three animals had infusions stopped before the 30 μL/min infusion rate was reached, because an ICP level of 40 mmHg was reached. These animals were included in all analyses.A total of 15 animals were included in this study: 9 treated with hypothermia to 33 °C and 6 normothermia controls. A total of 17 animals were excluded. Reasons for exclusion were lack of sufficient LDF drop at occlusion , outflow resistance  and infarct volume }.The Shapiro–Wilk normality test indicated that ICP, outflow resistance and infarct volume variables were normally distributed {ΔICP  was not significantly different between normothermia controls and hypothermia treated animals 18 h post-stroke , and if true, would be of physiological relevance. It is likely that ICP amplifies the effects of many hours of change in the balance between CSF production and outflow resistance. Therefore, what may seem to be minor differences in the outflow resistance rates between groups may result in quite significant differences when measured over a longer time scale.Our results suggest that hypothermia treatment may influence CSF outflow post-stroke, with animals treated with hypothermia having lower outflow resistance rates than normothermia controls, albeit not statistically significant. The recent study by Alshuhri et al.  demonstrOur results suggest that the involvement of CSF outflow resistance as a mechanism for ICP elevation cannot be discarded, however it is important to note that it may not be the only mechanism responsible for this pressure rise post-stroke. CSF production is the other compartment of CSF dynamics that influences the volume of CSF in the craniospinal compartment at any given time. An indirect estimation of CSF production rates by others could not detect an increase in CSF production post-stroke . TherefoAnother possible mechanism of interest relates to the glymphatic system. Glymphatic circulation drives CSF into the brain along periarterial spaces and drives interstitial fluid out . This flIt is also possible that hypothermia may prevent post-stroke ICP rise by mechanisms unrelated to CSF dynamics. A recent study by Kalisvaart et al.  suggestsWe have demonstrated that gradual cooling successfully prevents ICP elevation, reduces infarct volume post-stroke, and has a role in reducing CSF outflow resistance. A limitation of this study relates to the animal variability in the CSF outflow analysis. Another limitation of this study is the choice of anaesthetic used. Previous studies have reported that anaesthesia can alter responses in CSF flow  and haveIn conclusion, our results suggest that hypothermia treatment has important effects on CSF volume dynamics. CSF outflow resistance has been implicated as a mechanism for ICP elevation following experimental stroke. Moreover, hypothermia treatment has routinely been shown to prevent this ICP rise, suggesting hypothermic cerebroprotection acts directly on this mechanism. Having a mechanistic understanding of how hypothermia prevents ICP elevation is crucial if the treatment is to be translated to stroke patients. Therefore, these results are the first steps towards elucidating the mechanism involved in intracranial pressure elevation and hypothermia treatment following experimental stroke.