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BACKGROUND: Studies have shown that adipose-derived stem cells have pluripotent differentiation potential, but only 30%-40% of cells can differentiate into mature adipocytes with low adipogenic differentiation potential. Therefore, how to improve the adipogenic differentiation ability of adipose-derived stem cells is a key problem to be solved in the process of soft tissue regeneration. OBJECTIVE: To observe the relationship between the surface marker CD54 of rabbit adipose-derived stem cells and their adipogenic capacity, and to explore the adipogenic differentiation of CD54+/CD54- adipose-derived stem cells underthe same induction. METHODS: We successfully isolated and cultured the adipose-derived stem cells from inguinal subcutaneous fat pads (3 ml) of New Zealand white rabbits, aged 8-12 weeks, which were induced into multi-differentiation and used to detectsurface markers. We sorted the passage 3 adipose-derived stem cells by immunomagnetic beads and divided into two categories including CD54+ and CD54- adipose-derived stem cells. After 14 days of adipogenic induction, the cells in the two groups were subjected to oil red O staining and were compared by detecting the density of mature adipocytes and lipid droplet contenT.RESULTS AND CONCLUSION: The cultured adipose-derived stem cells possessed the characteristics of mesenchymal stem cells that could differentiate into mature adipocytes, osteoblasts and chondrocytes, with CD29, CD44, CD49d, CD54, CD73, CD90 and CD105 positive expression while CD31, CD34 and CD45 negative expression. Fourteen days after adipogenic induction, the density of mature adipocytes and the intracellular lipid droplet content in the CD54+ group were significantly higher than those in the CD54- group (P < 0.05). We also found that the mRNA expressions of PPARγ,ADD1, C/EBPα related to adipogenic differentiation in the CD54+ group were significantly higher than those in the CD54- group (P < 0.05). Taken together, CD54+ adipose-derived stem cells have excellent adipogenic differentiation capacity.
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ObjectiveTo investigate the effect of bone marrow-derived mesenchymal stem cells (MSCs) on gene and protein expression of toll-like receptor, TLR2/4 of the rat alveolus macrophages so as to discuss the effect of MSCs transplantation on the early inflammatory mediators and the regulatory mechanism of the inflammatory cascade.MethodsThe alveolus macrophages were randomly divided into the PBS control group (n = 24), MSCs control group (n = 24), lipopolysaccharide (LPS) control group (n=24) and LPS + MSCs treatment group (n =24).All groups were taken time-points of 1, 3,6, 12 hours.Positive cell rate of MSCs with surface mark and the expression level of TLR2/4 protein on macrophage were detected by flow cytometry.The mRNA level of TLR2/4 expression on macrophage was detected by RT-PCR and the concentrations of TNF-o in cell culture supernatants by ELISA at the different time points.ResultsCompared with the PBS control group and MSCs control group, TLR2/4 mRNA and protein levels were significantly up-regulated, markedly increased at one hour and peaked at 12 hours (P ＜0.01) and the TNF-α concentrations in cell culture supernatants were significantly up-regulated and peaked at 6 hours in the LPS control group (P ＜ 0.01).Compared with the LPS control group, mRNA and protein level of TLR2/4 and TNF-o concentrations in the cell culture supernatants of the LPS + MSCs group were markedly down-regulated (P ＜ 0.01).ConclusionsWith LPS stimulation, TLR2/4 mRNA and protein expressions as well as TNF-αt concentration increase, when the peak of the TNF-αt concentration appears slightly earlier, indicating that there may be a positive feedback loop between TLR2/4 expression and TNF-ot.MSCs can significantly reduce TLR2/4 mRNA and protein expressions as well as TNF-α concentration after LPS stimulation, indicating that MSCs can break up the positive feedback loop.
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Objective To investigate the effect of bone mesenchymal stem cells on Toll-like receptors (TLR) 2/4 expression in the lungs of rats with acute hemorrhagic necrotizing pancreatitis (AHNP). Methods Seventy SD male rats were randomly divided into sham-operation group (n=10), AHNP group(n=30) and MSCs-treated group(n=30). Masc rate of BMSCs with surface mark were measured by flow cytometer. TLR2/4mRNA expression in the the lung were measured by RT-PCR, and The ratio of Wet/dry and lung histological changs were observed. Results TLR2/4 mRNA could be detected in the lungs with low values in sham-operation group, markedly increased in 3 h, and peaked in 12 h in AHNP group (P＜0.05). Lung injuries were aggravated and the levels of TNF-α in the lung were increased (P＜0. 05) . Treatment with MSCs could effectively inhibit TLR2/4 mRNA expression and relieve lung injuries. The levels of TNF-α in the lung were decreased (P＜0.05). Conclusions The expression of TLR2/4 mRNA is increased in the lungs in AHNP and the lung injuries are aggravated. MSCs could markedly inhibit TLR2/4 mRNA expression in the lungs in AHNP, which would lead to relief of lung injury.
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Objective To investigate the relationship between EphA7 protein and carcinogenesis and development of pancreatic cancer by detecting the expression of EphA7 protein in pancreatic cancer tissues. Methods The expression of EphA7 in 10 cases of normal pancreatic tissue, 51 cases of pancreatic cancer and its adjacent tissues were detected with immunohistechemieal methods and the relationship between EphA7 and pathologic features of pancreatic cancer were analyzed. Results The rate of expression of EphA7 protein in normal pancreatic tissue was 10% (1/10), 47.1% (24/51) in adjacent pancreatic cancer tissues, 94.1% (48/51) in pancreatic cancer tissues. There were significant difference among the three groups (P <0.05). There was no significant correlation between the expression of EphA7 protein and the age, sex, tumor location, tumor size in patients with pancreatic cancer (P > 0.05). However there was significant correlation between the expression of EphA7 protein and the degree of differentiation, clinical staging, lymph node metastasis, or distant metastasis (P < 0.05). Conclusions The abnormally high expression of EphA7 may be relevant with the occurrence and development of the pancreatic cancer.
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AIM: Previous studies paid more attention to the effect of fibroblast immigration in normal surrounding tissue or blood on wound healing. This study explored the morphological character of fibroblasts after heat injury to support the new operation of autogenous epidermis overlapping denatured dermis. METHODS: The experiment was performed at the State Key Laboratory of Trauma, Burn and Combined Injury in the Third Military Medical University of Chinese PLA from April 2006 to April 2007. The dermal fibroblast from the infant prepuce disposed after surgery was cultured in vitro, and divided into 2 groups. In normal group, the dermal fibroblasts were put in water bath at 37 ℃ for 30 seconds; the cells in experimental group were stimulated at different temperatures (50, 51, 52, and 53 ℃) for different time (30, 60, 90, and 180 seconds). Cell survival was detected by MTT; morphology of injured cells were observed under inverted microscope and transmission electron microscope at 3 hours, 1, 2, 3, 4, 5, and 7 days; cell cycle was detected by flow cytometry. RESULTS: At different temperatures and different time, there were significant differences in the survival rate of fibroblast between the experimental groups and normal group (P