Mesenchymal stem cells belong to multipotential stem cells, which is easy to isolate and culture. Many researchers have been exploring various ways to induce bone marrow mesenchymal stem cells into muscle cells. MSCs were induced into muscle cells by using biochemical and biomechanics approaches. These muscle cells could be used for such clinical applications, as treatment for ischemic cardiomyopathy and post-traumatic repair for muscular tissue. In this article we reviewed the research progress on induction of bone marrow mesenchymal stem cells into muscle cells and isolation, purification, and identification of MSC.

BACKGROUND:E-cadhedn plays an important role in development of liver tissue in the embryonic stage.Therefore,it is importance for investigating the feasibility of dynamic expression of E-cadherin in embryonic stern cell differentiation to in vitro development of liver tissue.OBJECTIVE:To observe the dynamic expression of E-cadherin in embryonic stem cell differentiation and the effect on cell adhesion.DESIGN,TIME AND SETTING:An in vitro cytological observation was performed at Surgical Laboratory of the First Affiliated Hospital of Sun Yat-sen University from December 2007 to December 2008.MATERIALS:Embryonic stem cells of BALB/c mice were obtained from Professor Huang (Department of Ophthalmology,Sun Yat-sen University).Twenty 13-day-old pregnant BALB/c mice of clean grade were provided by the Experimental Animal Centar of Sun Yat-sen University.METHODS:Following trypsinization,embryonic stem cells were suspend-incubated in DMEM culture medium containing fetal bovine serum,2-mercaptoethanol,HEPES,penicillin,and streptomycin.Embryoid body was formed 5 days after normal development and incubated in the culture plate at day 6.Liver tissue which was obtained from 13-day-old pregnant BALB/c mice was prepared for fetal liver cells which were frozen-sectioned as the controls.MAIN OUTCOME MEASURES:E-cadherin expression was detected using reverse transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemistry st varying time points of 1,5,9,13,and 17 days in stages of initial differentiation of embryonic stem cells,formation of embryoid body,and formation of differentiated clusters.Additionally,the effect of E-cadherin expression on cell adhesion was also detected.RESULTS:RT-PCR showed that E-cadherin mRNA expression was not observed at day 1 but peaked at day 5;gradually,the expression was decreased until the expression was stopped at day 17.E-cadherin mRNA expression was strong in fetal liver cells in the control group.Immunocytochemistry showed a similar outcome.Morphologically,embryonic stem cells developed from unicells into compact three-embryonic layer embryoid body and into incompact cell population.CONCLUSION:E-cadherin expression correlates with differentiated cell adhesion;additionally,the lost expression in an in vitro environment may be an important cause for unable regularization of differentiated cells.

Objective To investigate the effects of structured triglyceride (STG) and physical mixed medium chain/long chain triglycerides (MCT/LCT) on hepatic and renal function and lipometabolism of patients with acute necrotizing pancreatitis (ANP).Methods The clinical data of 30 patients with ANP who were admitted to the PLA General Hospital between January 2012 and June 2014 were prospectively analyzed.A double-blind,randomized,controlled study was performed in 30 patients who were allocated into the experimental group (15 patients received STG) and the control group (15 patients received physical mixed MCT/LCT).All the patients received isometrical nitrogen and isocaloric parenteral nutrition more than 5 days.The levels of alanine transaminase (ALT),aspartate transaminase (AST),glutamyl-transpeptidase (GGT),alkaline phosphatase (ALP),creatinine (Cr),blood urea nitrogen (BUN),triglyceride (TG) and total cholesterol (TC) were assayed before nutritional support treatment and at day 1,3 and 5 after nutritional support therapy.The measurement data with normal distribution was presented as (x) ± s.The skew distribution data were described as M (range).The comparison between groups were evaluated with an independent sample t test or one-way ANOVA.The count data were analyzed using the chi-square test.Results A total of 30 patients were screened for eligibility.The levels of ALT,AST,GGT,ALP,Cr,BUN,TG and TC were changed within a certain range at day 1,3 and 5 after nutritional support treatment.The levels of ALT,AST,GGT,ALP,Cr,BUN and TC before treatment and at day 5after treatment were changed from 29.0 U/L,25.4 U/L,83.2 U/L,(193 ± 115) U/L,(124 ± 97) μmol/L,(8±6)mmol/L and (2.4±1.1)mmol/L to 29.4 U/L,33.0 U/L,77.7 U/L,(172±74)U/L,(117 ±103)μmol/L,(8 ± 5) mmol/L and (2.3 ± 1.0) mmol/L in the experimental group,and from 23.8 U/L,22.9 U/L,96.2 U/L,(148 ± 108) U/L,(82 ± 57) μmol/L,(9 ± 7) mmol/L and (2.5 ± 0.7) mmol/L to 21.3 U/L,24.5 U/L,127.4 U/L,(179 ± 126) U/L,(80 ± 54) μmol/L,(10 ± 6) mmol/L and (2.4 ±0.8) mmol/L in the control group,respectively.There were no significant differences in the changing trends of the levels of ALT,AST,GGT,ALP,Cr,BUN and TC between the 2 groups (F =0.647,1.186,0.282,0.553,0.862,0.182,0.369,P＞0.05).The level of TG in the experimental group from pre-treatment to day 5 after treatment was changed from (1.5 ± 0.6) mmol/L to (1.5 ± 0.7) mmol/L,with increasing trend from pre-treatment to day 1 after treatment and reaching the normal level at day 3 and 5 after treatment.The level of TG in the control group from pre-treatment to day 5 after treatment was changed from (1.5 ± 0.6) mmol/L to (2.4 ± 0.6) mmol/L,with increasing trend from pre-treatment to day 1,3 and 5 after treatments.There were significant differences in the changing trends of TG before and after nutritional support therapy between the 2 groups (F =7.940,P ＜ 0.05).Conclusion STG and physical mixed MCT/LCT don't influence the hepatic and renal function of patients with ANP undergoing parenteral nutritional support therapy,while STG has a better effect of lipometabolism compared with physical mixed MCT/LCT.Registry This study was registered with the UMIN Clinical Trial Registry with the registry number of UMIN000016958

BACKGROUND: Effects of embryonic stem cell-derived hepatocyte-like cell transplantation on oncogenicity of differentiated hepatocyte-like cells and biochemical metabolism of liver should be further studied.OBJECTIVE: To evaluate the therapeutic efficacy of embryonic stem cell-derived hepatocyte-like cell transplantation on the acute liver failure.DESIGN: Randomized controlled study.SETTING: the First Affiliated Hospital of Sun Yat-sen University.MATERIALS: This study was performed at the Central Laboratory, the First Affiliated Hospital of Sun Yat-sen University from January 2005 to February 2006. D3-ES cells extracted from the mice which underwent transfection of green fluorescent protein were graciously presented by professor Huang, Ophthalmology Center of Sun Yat-sen University. Forty 6-week-old D3-129 mice of clean grade and irrespective of gender were provided by Experimental Animal Center of Sun Yat-sen University [certification: SCXK (yue) 2004-0011]. The experimentzl animals were disposed according to ethical criteria.Transforming growth factor, basic fibroblast growth factor, and hepatocyte growth factor were provided by Gibco BRL Company, USA.METHODS: Transforming growth factor, basic fibroblast growth factor, and hepatocyte growth factor were combined to differentiate D3-ES cells into hepatic cells. Cell suspension was poured into liver capsule of 20 mice with 2.0×10(6)cells per mouse. Another 20 mice that determined as the controls were injected with saline. Twenty-four hours later, intraperitoneal injection of 5 μL/20 g carbon tetrachloride was used to induce acute liver failure and to observe quality of life and mean survival time. Twenty-four hours after acute liver failure, vena cava posterior blood was drawn to detect total bilirubin,glutamate-pyruvate transaminase, albumin, blood glucose, pro-time prothrombin time, and other hepatic functional parameters. By scarification, hepatic samples were obtained to evaluate oncogenesis condition, and then HE staining and immunohistochemistry were adopted to detect growth of transplanted cells and albumin expression.MAIN OUTCOME MEASURES: Quality of life, average survival time, hepatic functional parameters, growth of transplanted cells, and oncogenesis condition.RESULTS: Quality of life and average survival time: After the onset of acute liver failure, mice in the control group had incoordination and other symptoms of central nervous system. In addition, 14 mice in the control group and 8 in the transplantation group had abdominal dropsy. Average survival time in the control group was significantly shorter than that in the transplantation group (23, 62 hours, P<0.05). Hepatic functional parameters: Levels of total bilirubin and glutamate-pyruvate transaminase in the control and transplantation groups were higher than those before modeling; levels of albumin and blood glucose were lower than those before modeling; pro-time prothrombin time was significantly longer than that before modeling(P<0.01). Furthermore, levels of total bilirubin and glutamate-pyruvate transaminase in the transplantation group were lower than those in the control group; blood glucose in the transplantation group was higher than that in the control group, and pro-time prothrombin time in the transplantation group was significantly shorter than that in the control group (P<0.05). Growth of transplanted cells and oncogenesis condition: Pathological section demonstrated that structure of liver tissue was not changed remarkably, and tumor was not formed. Moreover, transplanted cells and hepatocyte-like cell were well arranged and combined to express albumin.CONCLUSION: Embryonic stem cell-derived hepatocyte-like cell transplantation can improve quality of life, prolong survival time of model mice with acute liver failure; additionally, transplanted cells may well support biochemical metabolism of liver tissue.

BACKGROUND: During the embryonic stage, E-cadherin expression plays a critical role in the formation of hepatic tissue.OBJECTIVE: E-cadherin gene was transfected into mouse embryonic stem cells (ESCs) to observe its effects on adhesive capacity of differentiated cells.DESIGN, TIME AND SETTING: A cytological in vitro observation was performed at the Laboratory of Surgery, First Hospital Affiliated to Sun Yat-sen University between December 2007 and December 2008.MATERIALS: BALB/c mice at gestational 13 days, of clean grade, were provided by Laboratory Animal Center, Sun Yat-sen University. BALB/c mouse ESCs were preserved by professor Huang Bing from the Department of Ophthalmology, Sun Yat-sen University. CMV promoter-containing eukaryotic expression plasmid pEGFP-N1 was gifted by doctor Lu Zhi-yue from Medical College, Sun Yat-sen University.METHODS: Total RNA was extracted from BALB/c mouse fresh hepatic tissue and synthesized into cDNA by reverse transcription (RT). The synthesized cDNA was used as a template to perform a polymerase chain reaction (PCR) that amplifies a targeted fragment. Following double enzyme digestion, pEGFP-E-cadherin plasmids were reconstructed and transfected into mouse ESCs. In vitro differentiation of transfected mouse ESCs was performed.MAIN OUTCOME MEASURES: Detection of E-cadherin expression in the differentiation system using RT-PCR and immunocytochemistry and observation of adhesive capacity of differentiated cells.RESULTS: E-cadherin gene-transfected ESCs could stably express E-cadherin during differentiational 1-17 days, while non-transfected ESCs expressed a decreasing amount of E-cadherin. The adhesive capacity of differentiated cells that stably expressed E-cadherin was markedly enhanced. Compact cell connection and multi-layer growth state remained at 19 days. While non-transfected ESCs gradually changed from embryoid bodies into noncohesive cell populations.CONCLUSION: Differentiating E-cadherin ESCs exhibit markedly enhanced adhesive capacity and maintain multi-layer growth state.

AIM:To observe the myogenic effect of bone marrow-derived mesenchymal stem cell(MSC)transplantation after chemical-induction and stress-induction on denervated skeletal muscle atrophy in vivo. METHODS:The experiment was performed at the Animal Experimental Center and Central Laboratory in Xinhua Hospital affiliated to Shanghai Jiao Tong University from April to October 2007.SD rats were provided by B&K Universal Group Limited.The experimental procedure was consistent with animal ethical standard.The MSCs from male SD rat femurs and tibia were isolated by density gradient centrifugation.The third passage of MSCs after chemical-induction and stress-induction was labeled with DAPI before transplantation.Thirty-six 8-week-old SD rats were randomly divided into control group and experimental group(n=18).The animal model of denervated gastrocnemius muscles were made by cutting the left sciatic nerve and creating nerve defect about 1 cm. MSCs were percutaneously transplanted to medial and lateral gastrocnemius of the experimental group rats,while low carbohydrates DMEM culture solution without MSCs and fetal bovine serum were transplanted to gastrocnemius of the control group.Bilateral gastrocnemius muscles of each rat from the two groups were weighed 4,8 and 12 weeks postoperatively after examining the motor unit potential and fibrillation potential.The gastrocnemius muscles underwent HE staining and image analysis to measure cross-section area of muscle fiber.The amount of protein was detected by BCA method. RESULTS:Muscular atrophy was observed 2 weeks after denervation.The motor unit waveshape of gastrocnemius became single; time limit became long;voltage became low,and the fibrillation potential norientation wave was increased.Differences were observed between two groups 4 weeks and 8 weeks,but no differences were observed after 12 weeks.Cells with fluorescence were observed in transplantation sites of the experimental group,but not in the control group.The wet weight remnant rate,cross-sectional area of fiber remnant rate and muscle amount protein content remnant rate in rates transplanted with MSCs were significantly lower than those in the control group 4 and 8 weeks after surgery(P

Translation is a fundamental step in regulation of gene expression and abnormalities in this process may lead to cancer. In eukaryotic cells, translation of mRNA is mainly regulated by many eukaryotic initiation factors ( eIFs) . EIF3 plays an impor-tant role in translational regulation, cell growth and oncogenesis. The largest subunit of eIF3, eIF3a may play a role as a regulator of mRNAs. The relationship between eIF3a and oncogenesis has been found. Moreover, the eIF3a mRNA is ubiquitously ex-pressed in different cancer cells and can modulate the cell cycle. However, some studies indicate that eIF3a could provide protec-tion against evolution into higher malignancy and reduce the re-sistance to chemotherapy . The patients of high eIF3a expression could get a better prognosis . In fact, the role of eIF3a is still un-clear in cancer cells. EIF3a may be involved in the process of tumor pathophysiology, but its regulatory role is undulatory.

Objective:To identify potential risk factors of varus collapse after unstable proximal humerus fracture treated with locking plates.Methods:A retrospective case series study was conducted on data of 146 patients with unstable proximal humerus fracture stabilized by locking plates at Xinhua Hospital, Shanghai Jiaotong University School of Medicine from January 2008 through December 2014. These patients were classified into varus collapse group ( n=39) and non-varus collapse group ( n=107) according to the occurrence of varus collapse. The gender, age, bone mineral density, cause of injury, fracture Neer classification, fracture type (varus or valgus), surgical timing, surgical techniques (medial support, cancellous bone graft, suture augmentation), number of humeral head screws and reduction quality were recorded. Potential risk factors were evaluated using univariate analysis and multivariate Logistic regression. The subjective reliability analysis was performed for Neer classification and medial fracture assessments. Results:Varus collapse group had higher ratio of osteoporosis, varus fracture, lack of medial column support, absence of suture augmentation and varus malreductin compared to non-varus collapse group ( P<0.05). While the two groups had no significant differences in gender, age, fracture classification, allogeneic cancellous bone transplantation and number of humeral head screws ( P>0.05). Moreover, the Logistic regression analysis indicated that osteoporosis, varus fracture, lack of medial column support, absence of suture augmentation and varus malreduction were major independent risk factors for varus collapse in proximal humerus fractures ( P<0.05). Among these risk variables, the lack of medial column support showed the strongest correlation of varus collapse after proximal humerus fractures treated with locking plates ( OR=9.62), and varus malreduction was another remarkable risk factor ( OR=8.39). The reliability of Neer classification and medial fracture assessments between interobservers and intraobservers was good. Conclusion:The risk factors for varus collapse after unstable proximal humerus fracture treated with locking plate are osteoporosis, varus fracture, lack of medial column support, absence of suture augmentation and varus malreduction.

Objective:To investigate the effect of passive motion and immobilization on shoulder function early after arthroscopic repair of rotator cuff tears.Methods:A retrospective case-control study was conducted to analyze the clinical data of 78 patients with rotator cuff tear admitted to Xinhua Hospital, Shanghai Jiaotong University School of Medicine from January 2016 to December 2017. There were 36 males and 42 females, aged 35-78 years [(62.7±3.2)years]. There were 36 patients with medium-sized tears (1-3 cm), 31 with small tears (<1 cm), and 11 with partial articular supraspinatus tendon avulsion (PASTA). All patients underwent arthroscopic rotator cuff repair. Forty-three patients started rehabilitation exercise immediately after operation (motion group). Thirty-five patients were immobilized with shoulder abduction brace for 6 weeks, and started rehabilitation exercise at week 7 (immobilization group). The range of motion, visual analogue pain score (VAS), simplified shoulder joint function test (SST) and Constant shoulder joint score were compared between the two groups before surgery, 6 weeks, 3 months and 12 months after surgery. The healing results were assessed by ultrasound 12 months after surgery. Complications were observed.Results:All the patients were followed up for 12-16 months [(13.7±1.3)months]. There were 7 patients with shoulder joint stiffness in each group (motion group: 16%, immobilization group: 20%) ( P<0.05). There were no significant differences between the two groups in VAS, SST or Constant score at postoperative 6 weeks, 3 months and 12 months ( P>0.05). The forward flexion and external rotation with the arm at the side in immobilization group was (124.9±12.9)° and 25(20, 30)° at postoperative 6 weeks, significantly improved in motion group [(136.6±16.7)°, 30(25, 40)°] ( P<0.05). There were no significant differences between the two groups in forward flexion and external rotation with the arm at the side at postoperative 3 and 12 months ( P>0.05). There were no significant differences between the two groups in internal rotation at postoperative 6 weeks, 3 months, and 12 months ( P>0.05). All rotator cuffs were healed verified by ultrasound at postoperative 12 months. No infection or implant displacement occurred after operation. Conclusions:For arthroscopic repair of medium-sized tears, small tears and PASTA, early postoperative rehabilitation exercises have advantage in improving range of motion only at early stage when compared to immobilization, which disappears with time. Moreover, the two methods have no significant differences in improving postoperative pain and shoulder function.

OBJECTIVETo investigate an method for hepatic differentiation from embryonic stem cells (ES cells) in vitro and the resulting differentiation ratio, in order to develop a procedure for producing a new type of hepatocyte for hepatocyte replacement therapy in the treatment of liver failure.

METHODSES cells from Balb/C mice were cultured and maintained in an undifferentiated state in gelatin-coated dishes using Dulbecco's modified Eagle's medium (DMEM) containing 1000 U/ml leukemia inhibitory factor (LIF). Then, LIF was withdrawn from the DMEM to allow the ES cells to develop into embryonic bodies (EBs). EBs were plated onto tissue culture dishes, and growth factors such as acidicfibroblast growth factor (aFGF) and hepatocyte growth factor (HGF) were added to the medium to promote directional differentiation. The course of development and differentiation was observed dynamically using an inversion microscope. The expression of hepatic proteins, such as alpha-fetoprotein (AFP), albumin (ALB), cytokeratin 8 (CK8), cytokeratin 18 (CK18), in cytoplasm was analyzed by immunocytochemistry (ICC). The concentration of ALB in the medium was determined dynamically by radioimmunoassay (RIA).

RESULTSES cells replicated as clones, without differentiating, in DMEM containing LIF. They developed into EBs in medium without LIF. Our ICC assay showed that differentiating cells did not express hepatic proteins, such as AFP, ALB, CK8, and CK18 until day 7, day 9, day 11, and day 11, respectively (up to 2 days later when growth factors are not present). The concentration of AFP in the medium was first detected on day 8, at a concentration of 3.4 ng/ml, and increased to 22.8 ng/ml by day 15. The concentration of ALB in the medium was 0.2 micro g/ml on day 11, and increased to 2.2 micro g/ml by day 15. ALB-positive cells under ICC manifest morphological structures were consistent with normal mouse hepatocytes. The differentiation ratio of hepatocytes in the ES cell differentiation system was 30% on day 15 (significantly lower without the presence of growth factors).

CONCLUSIONSES cells can differentiate into mature hepatocytes. Growth factors, such as aFGF and HGF, can enhance this differentiation and produce sufficient numbers of functional hepatocytes. This method may be a reliable new way of differentiating ES cells into hepatocytes for use in replacement therapy in the treatment of liver failure.

Animals ; Cell Differentiation ; physiology ; Cells, Cultured ; Embryo, Mammalian ; cytology ; Hepatocytes ; cytology ; Liver ; cytology ; Mice ; Mice, Inbred BALB C ; Stem Cells ; cytology