Objective:To screen plasma exosomal protein molecular markers in patients with spinal cord injury (SCI) by applying Label-Free quantification and bioinformatics analysis.Methods:Fifty plasma specimens from the First Affiliated Hospital of Nanjing Medical University (from January 2021 to June 2022) were collected from SCI patients and healthy people, respectively. Plasma exosomes were isolated using ultracentrifugation and identified by transmission electron microscopy, nanoparticle tracking analysis and western blot. Plasma exosomal differentially expressed proteins (DEPs) were analyzed using Label-Free quantitative proteomics, and DEPs were characterized, annotated, and enriched based on Gene Ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) databases. The screened DEPs were validated by western blot and enzyme linked immunosorbent assay (ELISA) using plasma exosomal specimens.Results:According to the spinal cord injury classification of the American Spinal Injury Association, 14 cases were grade A, 19 cases were grade B, 12 cases were grade C, and 5 cases were grade D. Plasma exosomes of SCI patients and control groups showed typical cup-like morphology, with diameters mainly ranging from 30-200 nm. A total of 493 exosomal proteins were identified by Label-Free quantification, and 126 proteins were screened for differential expression, of which 38 were up-regulated and 88 were down-regulated. GO annotation revealed that DEPs were mainly involved in functions such as protein activation cascade, complement activation and immune response. KEGG pathway analysis revealed that DEPs were involved in biological pathways such as complement and coagulation cascade reactions, proteasome and neurodegenerative disease pathways. Two candidate proteins, APOB and S100A9, were initially screened based on quantitative results from proteomics and bioinformatics analyses. Western blot results showed that the relative expression of S100A9 protein in plasma exosomes of 30 SCI patients (1.62±0.19) was elevated compared with that of 30 control groups (0.86±0.24), and the difference was statistically significant ( t=8.55, P<0.001), while the relative expression of APOB protein (1.06±0.13 and 1.02±0.23) were not statistically significant ( t=0.46, P=0.653). The results of ELISA analysis showed that the expression of S100A9 in plasma exosomes of patients with different degrees of SCI (grade A 197.7±11.7 pg/ml, grade B 151.7±15.2 pg/ml, grade C 136.3±14.7 pg/ml) had statistical significance ( F=69.94, P<0.001), the higher the severity of SCI, the higher the expression of S100A9 in plasma exosomes (A vs. B, q=13.11, P<0.001; A vs. C, q=15.66, P<0.001; B vs. C, q=4.19, P=0.005). Conclusion:S100A9 is a potentially valid plasma exosomal molecular marker for assessing the severity of SCI.

Objective: To discuss the result of treating tibia deformity with fixator assisted nailing technique (FAN). Methods: A total of 5 patients with 7 limbs of tibial deformity-were treated with FAN technique. Etiology: 2 patients with bilateral tibial deformity suffered from Ricket's disease, 2 patients were malunion after tibial fracture, 1 patient was congenital pseudoarthrosis of tibia. This is a retrospective study. The unilateral external fixator was mounted on the medial side of tibia, and thenthe minimal invasive osteotomy was performed. After the deformity was corrected, the intramedullary nail was inserted to fix the tibia. None of the patients need bone autograft. The pre-operation and post-operation medial proximal tibia angle (MPTA), mechanical axis deviation (MAD) and range of motion (ROM) were measured and analyzed. Results: All the 5 patients were followed-up for 12-60 months (average 32 months). The osteotomy site united in 3-5 months (average 4.5 months) post-operatively. According to Paley’s imaging scores, 4 patients were excellent and 1 patient was good. According to Paley's functional result scores, 5 patients were excellent; according to Paley’s bone results evaluation, 4 patients were excellent and 1 patient was good. We achieved desired post-operative MAD (from medial 15 mm-lateral 10 mm) in 6 limbs. The MPTA in 7 limbs was corrected to normal (84°-90°). The ROM was not significantly changed before and after operation. The average ROM of knee before operation was 125°(120°-135°), and average postoperative ROM was 120°(115°-130°), No deep infection or neurovascular injury occurred. All patients were satisfied with the method and results of the operation. Conclusion FAN technique combines the advantage of external fixation and intramedullary nail, and it is a good method to treat tibial deformity, the patient should be carefully evaluated and selected for applying this technique.

Objective To explore the role and mechanism of inducible nitric oxide synthase inhibitor 1 400W in alleviating ischemia-reperfusion injury of human intrahepatic bile duct epithelial cells.Methods Human intrahepatic bile duct epithelial cells (HIBEC) in logarithmic phase were inoculated into culture plate at an appropriate density.The samples were randomly divided into control group (group C),ischemiareperfusion group (group I/R) and ischemia-reperfusion + 1 400W group (group I/R + 1 400W).Group C was cultured routinely;cells in I/R and I/R + 1 400W groups were placed in a three-gas incubator for 12h for simulating ischemia and then normal culture for 6h for simulating reperfusion.The I/R + 1 400W group had a final concentration of 100 μmol/L of 1 400W before ischemia and hypoxia.After reperfusion,cells and culture medium were collected,CCK 8 was used for detecting cell vitality,microplate method for detecting the content of lactate dehydrogenase (LDH) in culture medium,AnnexinV-FITC/PI double stain for detecting apoptosis level,Western blot for analyzing the expressions of endoplasmic reticulum stress (ERS)related protein cysteinyl aspartic acid protease 12 (caspase-12),glucose regulatory protein 78 (GRP78) C/EBP homologous protein (CHOP) and inducible nitric oxide synthase (iNOS).Results As compared with group C,cell viability significantly decreased in I/R and I/R+ 1 400W groups (53.8％ ± 2.3％ vs.100％,66.5 ％ ± 2.8 ％ vs.100 ％) (P＜0.05) while LDH increased markedly in cell culture medium (287.4 ±9.0U/L vs 120.2 ± 8.7U/L,212.0 ± 8.3U/L vs 120.2 ± 8.7U/L) (P＜0.05).Apoptosis accelerated markedly (41.5％±2.3％ vs5.2％±0.5％,32.7％± 1.8％ vs 5.2％±0.5％) (P＜0.05) and the expressions of caspase-12,GRP78,CHOP and iNOS spiked (P＜0.05);as compared with I/R group,cell viability of I/R+ 1 400W group rose while LDH,apoptosis level,caspase-12,GRP78 and CHOP declined in cell culture medium (P＜0.05).Conclusions 1 400W may alleviate ischemia-reperfusion injury of human intrahepatic bile duct epithelial cells and its mechanism may be correlated with a suppression of endoplasrnic reticulum stress.

PURPOSE: An older female predominance has been reported among chronic cough patients in Western countries, which is considered to be associated with a higher cough sensitivity in females. However, the characteristics of Chinese chronic cough patients remain unclear. This study aimed to explore the age and sex distribution as well as their relationship with cough reflex sensitivity to capsaicin in Chinese chronic cough patients. METHODS: We analyzed the demographic features of 1,882 consecutive chronic cough patients who attended our cough clinic in Guangzhou, China. Cough sensitivity to capsaicin, which was defined as the lowest concentration of capsaicin causing 5 coughs or more (C5), was measured in 539 of the 1,882 patients and 68 healthy volunteers. RESULTS: The mean age of the patients was 43.0 ± 13.7 years and patients aged <50 years accounted for more than two-thirds of the study population. Around 87% of the patients were never-smokers. The proportion of females (51.5%) was almost equal to that of males (48.5%). The pattern of the age and sex distribution was consistently reflected within most common causes of chronic cough, while a female predominance was shown in patients with cough-variant asthma and patients aged ≥50 years. Female patients had higher cough sensitivity to capsaicin than male patients (log C5: 1.58 ± 0.84 vs. 2.04 ± 0.84 μmol/L, P = 0.001), and patients aged ≥50 years had higher cough sensitivity to capsaicin than patients aged <50 years. CONCLUSIONS: In China, patients with chronic cough have a roughly equal sex distribution and a middle-aged predominance, irrespective of a higher cough sensitivity to capsaicin in females and older patients. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02591550
Age Distribution ; Asian Continental Ancestry Group ; Asthma ; Capsaicin ; China ; Cough ; Female ; Healthy Volunteers ; Humans ; Male ; Reflex ; Sex Distribution

Objective To investigate the effects of curcumin on the growth and apoptosis of human hepatoma HCC-LM3 cells and explore the molecular mechanisms.Methods The human hepatoma HCC-LM3 cells were treated with different concentrations of curcumin and the cell growth inhibition rate was detected by CCK-8.The effect of curcumin on human hepatoma HCC-LM3 cells was observed.Refer to the relevant literature,the human hepatoma HCC-LM3 cells were treated with the concentration of 2.5,5.0,10.0,15.0,20.0,40.0,60.0 μmol/L of curcumin for 48 hours,taking the 0 μmol/L curcumin as control group,and the cell growth inhibition rate was detected by CCK-8.According to the results of CCK-8,selecting the concentration of 0 μmol/L as control group and the concentration of 10.0,20.0,40.0 μmol/L as experimental groups,which has significant difference on growth inhibition rates.Cell cloning assay was used to detect cell cloning ability,Flow cytometry was used to detect apoptosis,and Western blotting to detect the protein expression levels of Mcl-1,Bax,Bcl-2 and Bcl-xL.The measurement data were expressed in ((x) ± s),and the single factor analysis of variance was used for comparison between groups.Results CCK-8 assay showed that with treated by the concentration of 2.5,5.0,10.0,15.0,20.0,40.0,60.0 μmol/L,the growth inhibition rates were(6.71 ± 3.45)％,(12.33 ± 5.02)％,(20.07 ± 5.60)％,(57.80 ±7.34)％,(78.37 ±6.53)％,(91.73 ±6.14)％ and (96.18 ±3.45)％,suggesting that curcumin could inhibit the growth of human hepatoma HCC-LM3 cells in a dose-dependent manner.Cell clone formation experiment showed that curcumin could inhibit the clone of the human hepatoma HCC-LM3 cells,and the clone of the cells was inhibited significantly when the concentration of the curcumin was over 20.0μmol/L.The result of Annexin V-FITC/PI double staining analysis showed that the apoptotic rates of experimental groups and control groups were (5.20 ± 1.44) ％,(9.90 ± 3.31) ％,(55.67 ± 5.29) ％,(79.63 ±4.71)％,with all the apoptotic rates of experimental group over the control groups (P ＜0.05),suggesting curcumin could induce the apoptosis of human hepatoma HCC-LM3 cells.The Westen blotting showed that curcumin increased the expression of Bax protein while decreasing expression of Mcl-1 protein significantly in concentration-dependent manner (P ＜ 0.05),but have no effect on the expression of Bcl-2 and Bcl-xL proteins.Conclusion Curcumin could inhibit the proliferation and clone of human hepatoma HCC-LM3 cells,and induce apoptosis in a dose dependent manner.

Objectives To investigate the relationship between the degeneration of lumbar multifidus mus-cles and facet joint osteoarthritis(FJOA)by MRI. Methods This retrospective study included 248 patients with low back and leg pain,aged from 55 to 70.MRI was used to measure TCSA,FCSA,fCSA and FI% of the L4~5and L5~S1segments on the both sides of their multifidus muscle in the cross section in T2weighted image.All the cases were divided into three grades:grade 0~1,grade 2 and grade 3 according to their FJOA grades. The relationship between the FJOA grades at the same level and TCSA,FCSA and FI% of both sides was analyzed retrospectively. Results There were significant differences of the FI% of the multifidus muscle between both sides of the two seg-ments(P < 0.05);The TCSA,FCSA and fCSA of the left and right multifidus muscle all showed an increasing trend from L4~5to L5~S1segment and the difference was statistically significant(P<0.05);the FI% of two sides of multifidus muscle was negatively correlated with the right FJOA grade at L5~S1segment(P<0.05).There was a re-lationship between females and right FJOA grade at L5~S1segment(all P < 0.05). Whereas,no significant differ-ence was found in other parameters(all P>0.05).Conclusions There is no relationship between the cross-sec-tional area of the multifidus muscle and the FJOA grade. The higher FI% degree of multifidus muscle in the L5~S1 segment is accompanied with a lower grade of FJOA.The degeneration of multifidus muscle occurs earlier than the degeneration of articular process joints,so the earlier exercise of lumbar muscle is recommended.

Objective: To investigate the efficacy of sequential treatment with first-line administration of second-generation tyrosine kinase inhibitors (TKI) and first-generation TKI (imatinib) in patients with Ph⁺ acute lymphoblastic leukemia (Ph⁺ ALL) followed by allogeneic hematopoietic stem cell transplantation (allo-HSCT). Methods: Retrospective analysis of clinical features and prognosis of 76 newly diagnosed Ph ⁺ALL patients from June 2011 to December 2015 treated by allo-HSCT combined with first-line administration of second-generation or first-generation TKI was performed and the efficacy compared. Results: Of 76 Ph⁺ ALL patients, first-generation TKI was administered in 57 cases, second-generation TKI in 19 cases, including 10 cases of nilotinib and 9 cases of dasatinib. There was no significant difference in age, WBC counts, additional chromosomal abnormalities, time form diagnosis to transplantation, transplantation type, conditioning regimen or TKI initiation time between the two groups. Complete remission (CR) rates at the fourth week of induction therapy in first-generation TKI group and second-generation TKI group was 93.0% and 94.7% (P=1.000), respectively. Major molecular response (MMR, BCR-ABL/ABL reduce 3 log) rates meanwhile were 46.0% and 40.0% (χ²=0.169, P=0.681). Relapse rates before transplantation were 14.0% and 10.5% (P=1.000). MMR rates before transplantation were 54.4% and 68.2% (χ²=1.152, P=0.283). The 2-year overall survival (OS) rates of first-generation and second-generation TKI group were 62.0% and 94.7% (χ²=5.765, P=0.016), 2-year event-free survival (EFS) rates were 46.3% and 84.2% (χ²=5.644, P=0.018), respectively. Univariate analysis showed that second-generation TKI could improve OS (HR=0.126, 95%CI 0.017-0.939, P=0.043). Multiple factors analysis showed that second-generation TKI (HR=0.267, 95%CI 0.081-0.873, P=0.029) and MMR before transplantation (HR=0.496, 95%CI 0.254-0.968, P=0.040) were good independent prognostic factors of EFS. Conclusions There was significant difference in the efficacy of second-generation TKI and first-generation TKI for Ph⁺ ALL patients treated by allo-HSCT. First-line administration of second-generation TKI showed better efficacy than that of first-generation TKI for Ph⁺ ALL patients.

Objective: To investigate the efficacy of first-line administration of generic dasatinib or first-generation TKI (imatinib) in patients with Philadelphia chromosome positive acute lymphoblastic leukemia (Ph⁺ ALL) treated by hematopoietic stem cell transplantation (HSCT). Methods: Clinical features and prognoses of 63 newly diagnosed Ph⁺ ALL patients from Jan 2014 to June 2017 treated by HSCT combined with first-line administration of generic dasatinib or imatinib were retrospective analyzed. Results: Of 63 Ph⁺ ALL patients, 31 cases were administered generic dasatinib, and the other 32 ones imatinib. Complete remission (CR) rates at the fourth week of induction therapy in generic dasatinib and imatinib groups were 96.8% and 93.8% (P=1.000) , respectively. Meanwhile major molecular response (MMR; BCR-ABL/ABL reduce 3log) rates were 41.9% and 43.8% (χ²=0.021, P=0.884), respectively. Relapse rates before transplantation were 6.5% and 12.5% (P=0.672), respectively. MMR rates before HSCT were 83.9% and 68.8% (χ²=1.985, P=0.159), respectively. The 20-monthes overall survival (OS) rates of generic dasatinib and imatinib groups were 95.5% and 76.5% (χ²=0.990, P=0.320) respectively; 20-monthes event-free survival (EFS) rates were 93.5% and 61.4% (χ²=5.926, P=0.015), respectively. Statistically significant differences of EFS were reached. Multiple factors analysis showed that generic dasatinib (HR=0.201, 95% CI 0.045-0.896, P=0.035) and MMR before transplantation (HR=0.344, 95% CI 0.124-0.956, CI=0.041) could improve EFS. Conclusions First-line administration of generic dasatinib could improve EFS for Ph⁺ALL patients treated by HSCT when compered with imatinib.

Objective To investigate fatigue in children with acute lymphoblastic leukemia (ALL) during chemotherapy,and influencing factors.Methods From Feburary 2014 to August 2016, 216 children with ALL,from 5 to 15 years old,treated in a Grade III,Level A children's hospital in Zhengzhou,were surveyed by Peds QLTM multidimensional fatigue scale (Chinese version) and Questionnaire on discomfort in ALL children during chemotherapy.Results During chemotherapy,total scores of fatigue in the ALL children were (31.71±16.66) ,with fatigue of the whole body (11.67±6.29),of sleeping (10.99±5.57),and of cognition (9.10±6.84);during chemotherapy,incidence of fatigue of the whole body was 90.74%,of sleeping 95.83%, and of cognition 81.94%. Main influencing factors of fatigue of the whole body were weight loss,appetite loss and lack of energy,accounting for 72.8% of fatigue variation;that of sleeping were weight loss and irritability, accounting for 51.2% of fatigue variation;and that of cognition were difficulty in concentration and oral ulcer, accounting for 44.2% of fatigue variation.Conclusions During chemotherapy,fatigue in ALL children is common and is below average level. When managing fatigue symptoms in ALL children,nurses should actively treat the symptoms,so as to reduce the level of fatigue.

Objective To explore the expression profile of Wnt4 in rat kidney during renal development and its effect on renal development. Methods Rats with embryonic age of 18 days (E 18 d) , 20 days (E 20 d) as well as postnatal age of 0 day (P 0 d), 1 day (P 1 d), 3 days (P 3 d), 5 days (P 5 d) and 7 days (P 7 d) were selected. Expression levels of Wnt4 in rat kidney during renal development were quantified by immunohistochemistry and Western blot in all time points. Results Immuno?histochemistry analysis showed that during E 18 d to P 7 d, Wnt4 mainly expressed in proximal tubules, ureteric bud, comma shaped bodies and S shaped bodies of nephrogenic zone;the expression in the distal tubule was weak;the expression in renal corpuscle decreased with time;Western blot analysis showed that the expression of Wnt4 in rat kidney began to decrease from E 18 d and reached bottom at P 1 d then rise again until P 7 d when it dropped again. Conclusion During renal development, Wnt4 proteins were involved in the development of the nephrogenic zone through regulating canonical Wnt/β-catenin signaling pathway, and was involved in extension of proximal tubules by inducing the non canonical Wnt/PCP signaling pathway. Expression of Wnt4 protein in rat kidney was closely related to nephron formation and development of proximal tubules.