To explore the therapeutic effect of micro-traumatic surgery of loose seton and cutting seton by rubber bands in the treatment of high anorectal fistulae.Application of cutting seton (truss rubber bands) implemented the high part of fistulae and loose seton (ligation rubber band but non-fastened) for the low part of fistulae.133/136 patients undergoing micro-traumatic surgery were cured by one operation,2 cases had pseudo-healing and there was 1 recurrent case.And the curative rate was 97.8％.The microtraumatic surgery of loose seton and cutting seton by rubber bands in the treatment of high anorectal fistula has such multiple advantages as small incision,minor trauma,lesser pain,faster healing and a shorter course of treatment.And it may preserve the proper anal function and the integrity of anal skin.And its clinical efficacy is satisfactory.

Aim To measure the methylation rate of bone marrow mesenchymal stem cells (BMMSCs) using ultra-performance liquid chromatography-tandem mass spectrometry, and determine the methylation rate in the process of senescence.Methods The DNA extracted from BMMSCs would be digested into individual deoxynucleosides using enzymatic hydrolysis.To quantify the global genomic DNA methylation rate,we developed a method using ultra-performance liquid chromatography-tandem mass spectrometry with multiple reaction monitoring to simultaneously measure the levels of 5-methyl deoxycytidine and deoxyguanosine in digested genomic DNA.Results The DNA methylation rate could be analyzed within two minutes after hydrolyzing 1μg DNA for an hour.The detection limit of DNA methylation rate was 5.00×10-4.The coefficient of variance of the intra-day and inter-day precision was within 7.12×10-2 and 0.119, respectively.With the subculture of BMMSCs, the methylation rate gradually decreased.The methylation rate of stem cells was the lowest in 4~6 generation, and gradually increased with the subculture.Conclusions Using this method, the preliminary data on DNA methylation of BMMSCs in the process of senescence are obtained.The method is simple and convenient for sample preprocessing, and the method is fast and accurate with good sensitivity and repeatability.

[ ABSTRACT] AIM:To investigate the role of encapsulated protein transfected into human bone marrow mesen-chymal stem cells ( hBMSCs) by polyethyleneimine ( PEI) , and to optimize the best mole ratio of PEI-proteins.METH-ODS:6 groups of DNase I-PEI complexes were constructed and the best mole ratio was explored by laser scattering analy-sis.The appearance of complexes was presented under transmission electron microscope.Meanwhile, 4 groups of construc-ted GFP-PEI complexes were utilized to transfect into the hBMSCs, which were isolated and expand in vitro.The fluores-cence intensity of transfected cells was observed under confocal microscope.In addition, the cytotoxicity of the complexes on the cell proliferation was detected by MTT assay.The activity of the intracellular proteins was testified by aβ-galactosi-dase staining experiment.RESULTS:When the mole ratio of PEI and protein was adjusted to 4∶1, the complex transfec-tion efficiency was the best, and β-galactosidase color test turned blue.CONCLUSION:PEI has the character of encap-sulating various proteins to nano-complexes.The proteins transfected into bone marrow mesenchymal stem cells are con-firmed to have functional activity.As a protein carrier, PEI is of high efficiency and low toxicity, thus providing a new way for stem cell reprogramming.

Aim To explore the anti-apoptotic function of cardiac progenitor cells(CPCs)-derived exosome in vitro.Method CPCs were isolated from mouse heart using Magnetic Cell Sorting(MACS)system.Flow Cy-tometry(FC)determine the purity of stem cell surface antigen-1 positive(Sca-1 +)CPCs.Exosome was puri-fied from conditional medium,and confirmed by West-ern blot using CD63 as a marker,Nanoparticle Traffic-king Analysis(NTA)was used to detect the diameters and concentration of exosome.Then the cells were di-vided into control groups and CPC-exosome pre-protec-tion groups.H2 O2 was added into H9c2 cells to induce oxidative stress.Western blot was adopted to determine the expression of cleaved caspase-3.Results ① Im-munofluorescence showed that CPCs isolated by MACS were positively expressing Sca-1 protein;FC analysis showed that typical purity of Sca-1 +CPCs from the first
preparations was more than 95%.② WB demonstrated that CD63 of exosome isolated from CCMwas positively expressed,and NTA results showed that the diameters of exosome were (82.33 ±3.06)nm(n =3).Micro-scope detected PKH-26 labeled exosome appeared in the cytoplasma of H9c2 cells.③ Western blot showed the CPC-exosome pre-protection groups significantly down-regulated the levels of cleaved caspase-3 com-pared to the control groups(P <0.05).Conclusion CPC can secrete exosome which carries many important cargos,which can effectively gather in H9c2 cells. CPC-exosome can protect H9c2 cells from the oxidative stress induced by H2 O2 .Our results highlight a new perspective strategy for cardiac disease.

Objective To study the changes of trans-diaphragmatic pressure (Ptra) and its correlation with esophageal pressure (Peso) through ARDS piglet model.Methods Five piglets were enrolled in the study.Peso,gastric pressure (Pgas) and intra-thoracic pressure (Pint) was monitored through balloon inserted.The data before ARDS serve as control.ARDS was produced in the piglets through saline lavage.The pressure were observed and the Ptra were calculated.The pressure changes and correlation between Ptra and Peso were analyzed as well.Linear regression with the coefficient of determination and t-test were used as appropriate.Significance was assumed for P ＜ 0.05.Results Peso,Pgas and Pint before ARDS were 7.3 ± 1.9,25.5 ± 2.4,- 1.23 ± 0.21 cmH2O,Ptra was 18.2 ± 1.6 cmH2O.While after ARDS,the data were 4.7 ± 1.4,31.1 ± 3.1 and - 1.79 ± 0.28 cmH2O,and Ptra was 26.4 ± 2.1 cmH2 O,and all these changes were obviously ( P ＜ 0.05 ).The correlation between Pint and Peso,Pint and Ptra (A) and Ptra ( B ) were 0.93 ± 0.025,0.88 ± 0.023 and 0.87 ± 0.37 before ARDS.After ARDS,the correlation changed to be 0.82 ±0.21,0.81 ±0.20 and 0.78 ±0.31.Although a bit decreased,the correlation was still positive (P ＜ 0.01 ).Conclusions There existed good correlations between Peso and Ptra as well as between Pint and Peso before or after ARDS.Ptra was increased obviously after ARDS,which could lead to respiratory muscle fatigue.

BACKGROUND: Previous studies have demonstrated that the Chinese porous tantalum made in China has non-toxicity and good biocompatibility, which can promote osteogenesis. OBJECTIVE: To investigate the effects of transforming growth factor β1 on proliferation, cel cycle and secretion of osteoblasts on porous tantalum/MG63 osteoblast-like cel composites. METHODS: Passage 3 MG63 osteoblast-like cel suspension (1×109/L) was seeded onto the porous tantalum, then the cel composites were inoculated in the medium with 0, 0.5, 5 and 10 μg/L transforming growth factor β1, respectively. The proliferation of osteoblasts was detected by cel counting kit-8 assay at 1-13 days after inoculation; the cel morphology and ultrastructure observed by scanning electron microscope and transmission electron microscopy; and level of col agen type I detected by enzyme-linked immunosorbent assay. RESULTS AND CONCLUSON: 0.5, 5, 10 μg/L transforming growth factor β1 could promote the osteoblast proliferation, and cel proliferation in the 5 μg/L transforming growth factor β1 group was higher than that in the other groups; in the 5 μg/L transforming growth factor β1 group, laminated osteoblasts adhered on the surface and grew into inner of porous tantalum, which extended more pseudopodia toward the scaffold; osteoblasts-secreted matrix could cover the scaffold and numerous rough endoplasmic reticulum, free ribosomes, dense mitochondria, Golgi apparatus as wel as matrix vesicles could be found in the cytoplasm. In addition, the level of col agen type I in the 5 μg/L transforming growth factor β1 group was significantly higher than that in the other groups (P < 0.05). These results indicate that transforming growth factor β1 can promote proliferation, and col agen type I secretion of osteoblasts on porous tantalum/MG63 osteoblast-like cel composites, and the optimum mass concentration of transforming growth factor β1 is 5 μg/L.

AIM: To study the effects of fibroblast growth factor 8 (FGF8) on directional differentiation of human dental pulp stem cells (hDPSCs) into odontoblasts and pulp tissue.METHODS: hDPSCs were isolated and cultured, and identified with flow cytometry by detecting cell surface markers of hDPSCs.FGF8 at concentration of 50 μg/L was added into the mineralization fluid to induce the differentiation of the hDPSCs.The mRNA expression of dentin sialophosphoprotein (DSPP), alkaline phosphatase (ALP), bone sialoprotein (BSP) and core-binding factor alpha 1 (Cbfa-1) in differentiated cells was detected by real-time PCR.FGF8 and mouse E11.5 dental epithelium formed restructuring cell group with hDPSCs, and then the restructuring cell group was transplanted under renal capsule membrane in nude mice for tissue culture.DNA in situ hybridization was used to identify the sources of odontoblasts and pulp cells.RESULTS: The surface markers of CD29 and CD90 showed positive in isolated hDPSCs.FGF8 induced hDPSCs to form a distinct mineralization nodule, and the expression of dentin-specific proteins, DSPP, BSP and Cbfa-1, was increased.hDPSCs were induced to differentiate into odontoblasts and pulp cells by E11.5 dental epithelium and FGF8.CONCLUSION: FGF8 can assist dental epithelium to induce directional differetiation of hDPSCs into odontoblasts and pulp cells, and formation of dentin and dental pulp cavity structure.

BACKGROUND:Previous studies have shown that home-made porous tantalum has non-toxicity and good biocompatibility, and can promote osteogenesis. Herein, we explore the mechanisms of tantalum-bone interface osseointegration.OBJECTIVE:To observe the morphological characteristics and expressions of integrin β1 and fibronectin on the interface between porous tantalum and bone tissues after implantation into the right rabbit femur, and to evaluate the biological mechanisms of tantalum-bone interface osseointegration.METHODS: Animal models of bilateral femoral condyle defects were made in Japanese big ear rabbits. Porous tantalum rod and allogeneic bone were respectively implanted into the left (experimental group) and right (control group) femur of rabbits. The animal specimens at the bone defect region were taken and made into paraffin sections and hard tissue sections at postoperative 2, 4, 8 weeks for morphological observation of new bone at the junction between the tantalum rod and host bone under light microscope, for osteogenic observation of the tantalum-bone interface under scanning electron microscope, and for immunohistochemical detection of integrin β1 and fibronectin expression.RESULTS AND CONCLUSION:Porous tantalum was bonded closely with the host bone. The loose and thick fibrous capsule was observed in the early stage and became thinner in the late stage shown by hematoxylin-eosin staining. The new bone was visible on tantalum-bone interface. Hard tissue slicing observation showed that the new bone was seen on the porous tantalum-bone interface, blood capillaries grew into the pores at postoperative 2 weeks and the pores were full of new bone tissues at postoperative 4 and 8 weeks. Under the scanning electron microscope, the osteoblasts appeared on the tantalum surface and in the pores at the early stage, and bone maturation and lamelar bone were seen at the late stage. The immunohistochemical results showed that the expression of integrin β1 in the experimental group was significantly lower than that in the control group at postoperative 2 weeks (P < 0.05), but the expression of fibronectin had no significant difference between the two groups (P > 0.05). In addition, there was a decline trend in the expression of integrin β1 and fibronectin atpostoperative 2, 4, 8 weeks. To conclude, the porous tantalum material is beneficial to enhance adhesion of osteoblasts on the surface and inside the micro-pores. Increased expression of integrin β1 and fibronectin on the tantalum-bone interface at early stage may promote early osteogenesis, while their decreased expression at bone maturing stage can promote osseointegration and bone remodeling.

Background:China's accelerating development and increasingly important role in global health engagement create a great demand for global health professionals including international consulting experts.This study reported the detailed development and evaluation of an international consulting training for global health workforce.Methods:Based on Kirkpatrick's model,a mixed-methods approach was used to evaluate the effectiveness of the training.Quantitative and qualitative data on participants' reaction,learning,and application of the learned knowledge and skills were collected by a training evaluation survey at the ending of training and a follow-up interview in three months after the training.Results:Thirty-six participants attended the training and 34 of them completed quantitative investigation.The training satisfaction evaluations were positive,for which participants rated the training program highly and over 90％ of them agreed with the usefulness of the training.About knowledge and skills change,participants showed improved consulting knowledge and skills from pre-to post-training (P ＜ 0.001).A total of 23 participants accepted follow-up interview,and most participants applied knowledge and skills learned from the training in their daily work or study.However,only 30.4％ of participants applied their learning in the consulting program.The largest barrier of application was the lack of consulting opportunities.In addition,almost all the participants reported that they would be glad to attend more training courses in the future.Conclusion:The international consulting training program was well-received,and was feasible to improve the consulting service competence of global health professionals.According to participants' feedback,it is essential to develop and expand consulting training in the field of global health.

Objective:To investigate the correlation between vitamin D deficiency and the severity of symptoms in children with vasovagal syncope (VVS).Methods:A prospective study was conducted. One hundred and twenty-two children diagnosed with VVS by head up tilt test in Department of Pediatric Cardiology and 130 healthy children without symptoms who underwent physical examination in the outpatient department of Child Healthcare Department of Second Hospital of Lanzhou University from December 2019 to May 2021 were selected and assigned to VVS group and control group, respectively. According to the diagnostic criteria of vitamin D deficiency, children in the VVS group were assigned to three subgroups: non-vitamin D deficiency, vitamin D deficiency, and severe vitamin D deficiency. All children underwent detailed history taking, physical examination, and level determination of serum 25 (OH) D. Children in the VVS group were scored for orthostatic intolerance (OI) symptoms including 10 symptoms: syncope, dizziness, nausea, palpitation, headache, tremor, chest tightness, blurred vision, profuse perspiration, and attention deficit. The differences in the age, gender, body mass index, blood pressure, and serum 25 (OH) D levels between VVS group and control group, and the differences regarding the age, gender, body mass index, blood pressure, serum 25 (OH) D levels and symptom scores among the three VVS subgroups were compared. Comparisons were performed using independent sample t test, ANOVA analysis, Chi square test and rank sum test. Pearson correlation analysis was used to analyze the correlation between serum 25 (OH) D levels and OI symptom scores in children with VVS. Results:The serum 25 (OH) D levels were significantly lower in the VVS group than those in the control group ((31±11) vs. (46±10) nmol/L, t=10.89, P<0.001). Vitamin D deficiency was more frequent in the VVS group (73.0% (89/122) vs. 24.6% (32/130), χ2=58.91, P<0.001). There were significant differences among the severe vitamin D deficiency subgroup, vitamin D deficiency subgroup, and non-vitamin D deficiency subgroup regarding the serum 25 (OH) D levels ((9.8±0.4) vs. (26.6±6.5) vs. (45.8±5.9) nmol/L, F=142.77, P<0.001) and the OI symptom scores ((14±1) vs. (10±2) vs. (7±2) scores, F=44.97, P<0.001). The scores of syncope, nausea, profuse perspiration, blurred vision and dizziness among the severe vitamin D deficiency subgroup, vitamin D deficiency subgroup, and non-vitamin D deficiency subgroup were statistically significant ( H=9.01, 7.52, 12.11, 7.07 and 9.54, respectively, all P<0.05). Pearson correlation analysis showed that the serum 25 (OH) D levels were negatively correlated with OI symptom scores in children with VVS ( r=-0.769, P<0.001). Conclusions:VVS children have significant vitamin D deficiency. The severity of symptoms increases with decreasing of vitamin D level. Syncope, nausea, and profuse perspiration are more likely to occur in children with severe vitamin D deficiency, and dizziness and blurred vision are more likely to occur in children with vitamin D deficiency.