| Background and purposeThis study investigates the differences between ADSCs from rats with Type2diabetes and normal rats by observe the differences of the growth curve and of these two groups, and using ELISA tests to observe the amount of adipokines (leptin adiponectin and tumor necrosis factor-alpha) secreted by ADSCs of both groups, to verify if there were any differences between ADSCs from diabetic rats and normal rats.Methods1. Forty6-week-old male Wistar rats were randomly divided into experimental group and control group, with20rats in each group.The rats in experimental group were intraperitoneally injected with STZ (streptozotocin) buffer solution and given high glucose and high fat diet for4weeks to establish the model of type2diabetes rats.The control group only received routine feeding.2. The inguinal fat pad and epididymis fat pad of rats from both the experimental group and the control group are cut off. ADSCs were extracted from those tissues and cultured by the same method in the same environment. These cells were tested through the osteogenic, chondrogenic and adipogenic differentiation and cell phenotype identification to determine it was ADSCs.When they passaged to the P3generation, adipose-derived stem cells cell growth curve are assay in both groups.3. P3generation adipose-derived stem cells of both groups are cultured in adipogenic differentiation medium for14days to different into adipocytes. Each Medium sample was extracted from Petri dish and detected by ELISA(enzyme linked immunosorbent assay) tests of adipokines leptin, adiponectin and TNF-a(tumor necrosis factor alpha), and Analysis the various difference of adipokines expression between the groups. And the collected data are analysed by the One Way ANOVA method.Results1. ADSCs can be obtained by the method of digesting rat fat from inguinal fat pad and epididymal fat pad with type I collagenase, and cultured in DMEM(High Glucose) medium,and maintained a well function and high level of proliferation ability.By phenotypic analysis and staining after osteogenic, chondrogenic, adipogenic differentiation to verify its versatility, proves that this method can obtain adipose-derived stem cells with strong activity and versatility.2. The growth curve of ADSCs from both the experimental group and the control group are inverted S-shaped, and both have a exponential phase at day2to day8,and both have the fast generation rate in day4.The experimental group of fat stem cells reach plateau time (at day10) later than cells from the control group (at day8)3. Adipocytes differentiated by ADSCs from experimental group have a significantly lower adipogenic rate (less than50%) compared with the control group(more than90%) in the microscope. The leptin level of experimental group (637.25±102.03pg/ml) and control group (122.41±81.27pg/ml) has a significant different (P<0.05). Adiponectin level between the experimental group (42.3±10.5ng/ml) and control group (31.7±8.1ng/ml), also has statistically difference (P<0.05),but tumor necrosis factor a has not statistically difference between experimental group (1763.7±249.4.0pg/ml) and control group (1820.4±316.1pg/ml)(P>0.05) Conclusions1. ADSCs from type2diabetic rats have no significant difference in growth rate comparing to the normal rats’ ADSCs, but a delayed peak has been found in the growth curve of type2diabetic rats’ADSCs.2. Adipocytes derived from type2diabetic rats’ ADSCs secret more leptin and adiponectin than the normal rats’ADSCs, but the levels of tumor necrosis factor-alpha secretion have no significant difference. |
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