AIM: To study the effects of cotransplantation of donor-derived bone marrow mesenchymal stem cells on graft versus host disease in a rat allogeneic bone marrow transplantation model. METHODS: Fisher 344 rat bone marrow MSCs were isolated and cultured to the fifth passage (P5) in vitro . The recipient Wistar rats were conditioned with lethal total body irradiation and transplanted with F344 rat bone marrow cells and spleen cells in the presence or absence of (P5) MSCs. The onset time of graft versus host disease (GVHD), incidence of GVHD and survival time were monitored. RESULTS: Cotransplantation of MSCs deferred the onset time of GVHD[(19.1?1.7) d vs (15.6?1.5) d, P

AIM:To investigate the characteristics of Ets-1 and VEGF expression and distribution in the experimental diabetic rat retina.METHODS:Diabetes was induced by intraperitoneal injection of streptozotocin (STZ).At 4 weeks after STZ-injection,animals were sacrificed.Total proteins were isolated from retinas of experimental and control eyes and were assessed by Western blot analysis.Frozen cross sections of eyeballs with 14um thickness were used to perform double immunoffuorescence staining with anti-Ets-1 and anti-VEGF antibodies.RESULTS:Both Ets-1 and VEGF expression were up-regulaled in the diabetic retina,the distribution of Ets-1 and VEGF was identical to each other,and the two proteins were almostlocalized in all retinal layers.CONCLUSION:Ets-1 might contribute to the pathologic progress of the diabetic retina induced by VEGF.

AIM: To determine the involvement of Ets-1 in the pathological progress of the experimental diabetic retina. METHODS: Diabetes was induced by intraperitoneal injection of STZ. Total RNA and Total proteins were isolated from retinas of experimental and control eyes at 4 weeks after STZ-injection and were assessed by Northern blot analysis and Western blot analysis, respectively.RESULTS: Expression of both Ets-1 mRNA and Ets-1 protein was significantly increased in the experimental diabetic rat's retina after STZ-injection compared with the control group (P＜0.001).CONCLUSION: Our results indicated that Ets-1 was involved in the pathological progress of experimental diabetic retina.Further studies should be conducted to focus on the relationship between Ets-1 and VEGF in the diabetic retina.

Objective To investigate the role of oxidative stress in the epithelial-to-mesenchymal transdifferentiation (EMT) of peritoneal mesothelial cells in rat model of peritoneal fibrosis and the effect of probucol on peritoneal fibrosis. Methods The rat model of peritoneal fibrosis was induced by 4.25% high glucose peritoneal dialysis fluid (PDF). The rats were randomly divided into 4 groups:the control group, the saline group, the peritoneal fibrosis group, and the probucol group. A 4 hour peritoneal equilibration test (PET) was performed 4 weeks later. The peritoneal function and net ultrafiltration (UF) volume were determined. The level of malondialdehyde (MDA) and glutathione peroxidase (GSH-Px) in peritoneal tissue were examined. The histology of peritoneal membrane was evaluated by light microscopy. E-cadherin and α-smooth muscle actin (α-SMA) protein expression was evaluated by immunohistochemical method and Western blot.Results The mesothelial cells were detached from peritoneal membrane in peritoneal firbosis rats. Comparing with the control rats, the thickness of visceral peritoneum, the level of MDA, and the-SMA protein expression were increased while the net ultrafiltration volume, the level of GSH-Px and E-cadherin protein expression were decreased in peritoneal firbosis rats. All these changes were reversed in the rats treated with probucol.Conclusion Oxidative stress plays an important role in transdifferentiation of peritoneal mesothelial cell in the peritoneal fibrosis rats. Probucol can improve structure and function of peritoneum, and partially reverse the EMT by reducing the oxidative stress.

Objective To investigate the effects of connective tissue growth factor (CTGF) siRNA delivered by pRetro-Super (PRS) retrovirus vector on extracellular matrix and VEGF expression in human peritoneal mesothelial cells (HPMC). Methods Four pairs of oligonucleotides including 64 bp DNA were designed and synthesized in vitro according to siRNA target sequence and PRS retrovirus desire.PRS-CTGF-siRNA1-4 recombinant retrovirus vectors were constructed.The recombinant retrovirus vectors containing CTGF-siRNA were transferred into PT67 packaging cell lines with lipefectamine 2000,then infected HPMC.mRNA expression was determined by semi-quantitative RT-PCR and protein expression was determined by Western blot.Results Both mRNA and protein expressions of CTGF,FN,Col I,laminin (LN) and VEGF were significantly increased in HPMC with 5 μg/L TGF-β1 stimulation (P＜0.01,respectively).CTGF,FN,Col I,LN mRNA and protein and VEGF mRNA expression stimulated by TGF-β1 were significantly decreased in HPMC infected with PRS-CTGF-siRNA1～4 retrovirus vectors (P＜0.01,respectively).The inhibitory rates on CTGF were 69.3%,22.2%,27.4% and 38.8%,respectively (P＜0.01).At the same time,there was also a significant reduction of VEGF protein expression in HPMC infected with PRS-CTGF-siRNA1 vector (P＜0.01).There was no significant difference in HPMC infected with PRS void vector. Conclusion CTGF siRNA delivered by PRS retrovirus vector can effectively inhibit the enhancement of extracellular matrix and VEGF expression stimulated by TGF-β1 in HPMC.

Objective To investigate the relevant risk factors for fungal infection following operation of the gastrointestinal neo- plasm and offer supporting data for the prevention of fungal infection.Methods Medical records from 116 patients who under- went the operation of gastrointestinal neoplasm in the special group of this hospital from January 2006 to June 2006 were retro- spectively reviewed on the relevant risk factors by univariate and multivariate Logistic regression analysis.Results Of the 116 patients reviewed, 18 had fungal infection.Forty-six samples were positive for fungal pathogen.The most frequently isolated fungal strain was Candida albicans (15/20) and the most common infection site was gastrointestinal tract (14/18).Fungal in- fection after the operation of gastrointestinal neoplasm was significantly relevant with the duration of antibiotic use, duration of post-operative fasting, low serum albumin, high blood glucose and complication of bacterial infection.The duration of antibiotic use was a significantly independent risk factor.Conclusions Reasonable antibiotic use, nutritional support, early enteral nutri- tion and control of blood glucose should be taken into account after the operation of gastrointestinal neoplasm in order to prevent fungal infections.

OBJECTIVETo analyze the effects of three surgical operations in the treatment of Pilon fracture of Rüedi-Allgower type III, and put forward the best therapeutic method.

METHODSThe clinical data of 33 patients with Pilon fracture who received surgical operations (plaster immobilization group, 10 cases; distal tibia anatomical plate group, 11 cases; external fixation with limited internal fixation group, 12 cases) from October 2009 to January 2012 were analyzed. There were 5 males and 5 females, ranging in age from 24 to 61 years in the plaster immobilization group. There were 7 males and 4 females, ranging in age from 21 to 64 years in the distal tibia anatomical plate group. There were 7 males and 5 females, ranging in age from 23 to 67 years in the external fixation with limited internal fixation group. The Ankle X-ray of Pilon fracture after operation, ankle score, early and late complications were collected. Bourne system was used to evaluate ankle joint function.

RESULTSAfter 8 months to 3 years follow-up, it was found that three kinds of treatment had significant differences in the outcomes and complications (P < 0.05): the external fixation with limited internal fixation group got the best results. The number of anatomic reduction cases in the external fixation with limited internal fixation group (7 cases) and the distal tibia anatomical plate group (8 cases) was more than the plaster immobilization group (2 cases). According to the ankle score, 8 patients got an excellent result, 3 good and 1 poor in the limited internal fixation group ,which was better than those of distal tibia anatomical plate group (5 excellent, 4 good and 2 poor) and the plaster immobilization group (3 excellent, 4 good and 3 poor). The number of early and late complications in the external fixation with limited internal fixation group was more than those in the plaster immobilization group and the distal tibia anatomical plate group (P< 0.05).

CONCLUSIONTreatment of external fixation with limited internal fixation in the treatment of Pilon fracture of Rüedi-Allgower type III is effective and safe.

Adult ; Aged ; Case-Control Studies ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Retrospective Studies ; Tibial Fractures ; diagnostic imaging ; surgery ; Tomography, X-Ray Computed ; Treatment Outcome ; Young Adult

Objective To determine the effect of smad anchor for receptor activation (SARA) on renal tubular epithelial to mesenchymal transtion (EMT) induced by high glucose and to investigate the associated mechanism.Methods HK-2 cells were exposed to high glucose (30 mmol/L).HK-2 cells were transfected with the plasmids of wild-type SARA [SARA (WT)] or SARA mutant [SARA with SBD deletion,called SARA (dSBD)] and then was stimulated by high glucose.The gene expression was assayed by real-time PCR and the protein expression was detected by Western blotting.Results During the process of high glucose-induced EMT of HK-2 cells,the gene and protein expression of SARA were down-regulated.The expression of TGF-β1 and Smad3 increased after stimulation of high glucose in HK-2.However,the Smad2 mRNA expression increased while its protein expression was down-regulated in a time-dependent manner.Smad2 and Smad3 were activated by high glucose stimulation and Smad3 kept activation for longer time than Smad2.Compared with high glucose group,over-expression of SARA by transfection of SARA (WT) up-regulated the expression of zona occludens(ZO)1 and down-regulated the expression of vimentin (P＜0.05).However,SARA (dSBD) had no such effects on above expressions.The Smad2 protein expression increased along with the over-expression of SARA.Meanwhile,over-expression of SARA prolonged the activation time of Smad2 and shortened the activation time of Smad3.Conclusions TGF-β1 signaling is activated and SARA expression is down-regulated during the process of high glucose-induced EMT in HK-2 cells.Over-expression of SARA can inhibit the EMT via increase of Smad2 protein expression and longer activation time of Smad2.

Objective To determine the effect of 2 transforming growth factor β1 (TGF-β1) short hairpin RNA (shRNA) expression plasmids (pcDU6-A1-A2 and pcDU6-B1-B2) on proliferation, TGF-β1, connective tissue growth factor (CTGF), and fibronectin (FN) expression induced by human serum albumin (HAS) in HK2 cells. Methods A vector plasmid containing the TGF-β1 shRNA was generated. An HK2 cell line was used in the study. The 2 TGF-β1 shRNA expression plasmids were transfected into cultured HK2 cells by lipofectamine 2000. Cellular proliferation was assessed by tetrazolium salt colorimetry. The semi-quantitative reverse transcriptive PCR was performed to detect the expression of TGF-β1,CTGF, and FN mRNA. Levels of TGF-β1 and FN protein were measured with a sandwich enzyme-linked immunosorbent assay. Results After treating with 5 g/L HAS for 24 hours in HK2 cells, cellular proliferating capacity increased significantly (P＜0.05). The expression levels of TGF-β1, CTGF, and FN mRNA were upregulated in HK2 cells stimulated by 5 g/L HAS, and levels of TGF-β1 and FN protein in the culture supernatant increased (P＜0.05). The introduction of pcDU6-A1-A2 and pcDU6-B1-B2 resulted in significant reduction of cellular proliferation activity, and the expression levels of TGF-β1, CTGF, and FN mRNA were downregulated (P＜0.05). Levels of TGF-β1 and FN protein in the culture supernatant decreased (P＜0.05) after 12 or 24 hours of TGF-β1 shRNA transfection into HK2 cells There was no significant difference in the expression levels of TGF-β1, CTGF, and FN mRNA between the 2 pcDU6 vector plasmid mediated TGF-β1 shRNA groups (P＞0.05). Conclusion pcDU6 vector plasmid mediated TGF-β1 shRNAs could obviously inhibit the expression levels of TGF-β1, CTGF, FN and cellular proliferation stimulated by HAS in HK2 cells, which may be related to the mediation of TGF-β1.

OBJECTIVETo establish the model of bone mesenchymal stem cell-derived smooth muscle cells (BMSC-SMCs) and investigate the role of BMSC-SMCs in the development and progression of artherosclerosis.

METHODSBMSCs were isolated from the femoral bone of SD rats by adherent tissue culture method, and vascular smooth muscle cells (VSMCs) were obtained from the thoracic aorta. The differentiation of BMSCs into BMSC-SMCs was induced in the conditioned medium. The specific markers of BMSCs and BMSC-SMCs were identified by immunofluorescence (IF) staining. After treatment with 80 mg/L oxidative low-density lipoprotein (ox-LDL) for 72 h, the growth characteristics of BMSC-SMCs and VSMCs were observed. Flow cytometry was applied to analyze the cell cycle of BMSC-SMCs and VSMCs.

RESULTSBMCS-SMCs transformed into foam cells after treatment with ox-LDL, which was more obvious in comparison with VSMCs. The growth curve of BMSC-SMCs and VSMCs presented with an S-shape pattern with the cell doubling time of 20 and 32 h, which was reduced to 15 and 28 h after treatment with 80 mg/L ox-LDL, respectively. Flow cytometry showed that exposure to 80 mg/L ox-LDL significantly increased G(0)/G(1) and decreased S and G(2)/M phase cells in both BMSC-SMCs (P<0.01, n=3) and VSMCs (P<0.05, n=3) in comparison with the control cells.

CONCLUSIONBMSC-SMC might be involved in the formation of fatty core and accelerate the development of atherosclerosis.

Animals ; Atherosclerosis ; etiology ; Bone Marrow Cells ; cytology ; Cell Differentiation ; physiology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Female ; Foam Cells ; cytology ; Lipoproteins, LDL ; pharmacology ; Male ; Mesenchymal Stromal Cells ; cytology ; Muscle, Smooth, Vascular ; cytology ; Rats ; Rats, Sprague-Dawley