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Objective To analyze the differential expression of D2 and D3 alternative splicing sites of human kinectin in human hepatocellular carcinoma (HCC) tissues, adjacent non-cancerous tissues and normal liver tissues, and to investigate a possible relationship between alternative splicing sites of kinectin and hepatocarcinogenesis. Methods The cDNA was obtained by RT-PCR in 45 coupled HCC cancerous and adjacent tissues, and 10 normal liver tissues. The difference in the expression of D2 and D3 alternative splicing sites in cDNA was examined by semi-quantitative PCR, and statistical analysis was performed. Results The ratio of D2L (long segment contains of the D2 region)/D2S (short segment that does not contain a D2 zone) in hepatocellular cancerous tissues was 2.709 ± 1.025, the ratio of D2L/D2S in adjacent non-cancerous tissues was 1.564 ± 0.357, and the ratio of D2L/D2S in normal liver tissues was 1.507 ± 0.499. The differences were statistically significant (F=29.698, P 0.05). Conclusion Variant containing D2 is over expressed in cancerous tissues and this alteration may be tumor associated.
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Based on the known IRAK4 inhibitors MK-32 and AU-5,we designed and synthesized 12 pyridine-based target compounds by adopting open-ring and hybrid strategies,and combining molecular docking technology.The bioassays determined by radioisotope labeling demonstrated that the target compounds displayed good inhibitory activity against IRAK4.Among them,the IC50 value of 5 compounds was less than 1 μmol/L,suggesting that these compounds may be candidates for further investigation.
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BACKGROUND:Studies have shown that Schwann cells form a Bunger band in the basement tube and guide the extension of regenerating axons after peripheral nerve injury, but the exact mechanism remains to be explored. OBJECTIVE:To explore the effect of Wal erian degeneration on biological characteristics and secretory function of Schwann cells in rats with sciatic nerve injury. METHODS:A rat model of sciatic nerve injury was established and divided into two groups:sciatic nerve transection group and surgical control group. Schwann cells were isolated and cultured from sciatic nerve segments by one enzyme digestion. The cellmorphology was observed under light microscope and S-100 protein expression was determined by immunofluorescence staining. After subculture, the first generation of Schwann cells were chosen to draw the growth curve by the counting method within 14 days. The cellactivity was detected by MTT assay. The adhesion of Schwann cells was examined by acid phosphatase analysis and the concentration of nerve growth factor was detected by ELISA method. RESULTS AND CONCLUSION:At 14 days after primary culture, a great number of Schwann cells were observed near the edges of nerve segments in the sciatic nerve transection group, but only smal number of Schwann cells scattered around nerve segments in the control group. Schwann cells in both groups showed S-100 positive expression. At 3 days after subculture, Schwann cells reached the logarithm proliferative phase, the cellnumber and proliferation absorbance values in both groups were increased along with time extension. Furthermore, the number of Schwann cells and absorbance value in the sciatic nerve transection group were significantly higher than those of control group (P<0.05). The adhesion ability in the sciatic nerve transection group was also significantly higher than those in the control group (P<0.05). ELISA results showed that, the concentrations of nerve growth factor in the sciatic nerve transection group were significantly higher than those in the control group at 4, 6, 8, 10, 12 and 14 days (P<0.05). After sciatic nerve injury, Wal erian degeneration can induce Schwann cells dedifferentiate into the precursors, significantly influence the biological function of Schwann cells, promote the proliferation of Schwann cells within the short term, secrete large amounts of neurotrophic factors, enhance celladhesion, and provide a suitable microenvironment for regenerated axons. In addition, it creates the necessary microenvironment for peripheral nerve regeneration.
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BACKGROUND: Schwann cells are the seed cells of neural repair, and it is a key to harvest a large number of Schwann cells with high purity and activity. OBJECTIVE: To compare the in vitro culture, purification, and morphology of Schwann cells between neonatal and adult rats, and investigate a simple and feasible culture method to harvest high-purity Schwann cells. METHODS: Totally 30 Sprague-Dawley rats, comprising 20 neonatal (1-3 days after birth, neonatal group) and 10 adult (weighing 150-200 g, adult group) rats, were included. Following double-enzyme digestion and two incubations, Schwann cells were isolated and purified by differential attachment. Cell morphology and attaching speed were determined through the use of inverted microscope. Cells were counted and cell purity was calculated. Cell proliferative ability was detected by MTT microcolorimetry. Curves of cell proliferation in each group were depicted to determine proliferative speed. Schwann cells were identified by S-100 immunochemistry.RESULTS AND CONCLUSION: Compared with fibroblasts, neonatal rat Schwann cells exhibited faster, while adult rat Schwann cells showed slower, attaching speed. Both neonatal and adult groups yielded over 96% cell purity. MTT microcolorimetry results revealed that Schwann cells proliferated actively in neonatal and adult groups. Cell proliferative curves show that neonatal rat Schwann cells proliferated faster than adult rat Schwann cells (P < 0.05). S-100 immunochemistry results showed positive results in both groups. All these findings suggest that double-enzyme digestion and two incubations followed by differential attachment is a satisfactory method to harvest considerable Schwann cells with high purity and activity. Neonatal rat Schwann cells show stronger proliferative, attaching capacities than adult rat Schwann cells.
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BACKGROUND:Schwann cells transplantation can change the local micro-environment and help to repair the injured neural tissue, so getting a large number of highly purified and active Schwann cells is the key of the study. OBJECTIVE: To search for a simple and rapid method to extract and purify the Schwann cells.METHODS: Rats were divided randomly into two groups, namely, in vivo pre-degeneration of sciatic nerve resection group and untreated control group, with 20 rats in each group. Under sterile conditions, the rat sciatic nerves were cut off at post-operative 7 days, Schwann cells were extracted by using mixed enzyme digestion and tissue mass transplantation; through low enzyme digestion and twice inoculation to differential adhesion, Schwann cells were purified. Cell morphology was observed under phase contrast microscope and identified by mmunofluorescence staining; cell purity was calculated; MTT method assay was used to determine the capacity of cell proliferation.RESULTS AND CONCLUSION: At 7 days of the culture, the experimental group showed the typical bipolar or bipolar Schwann cells, with connections between cells; in control group, cell processes were shorter and less associated with the surrounding cells. Following S-100 immunofluorescence staining, cells were positive for green expression.Cells proliferated rapidly in the experimental group and formed a swirling shape at 15 days, there were a relatively small number of fibroblasts, at the purity of 96.1%; in the control group, the cells proliferated slowly, with many fibroblasts at a low purity. MTT assay showed that primary cultured Schwann cell proliferated weakly in both groups; compared with the control group, the proliferation of subcultured Schwann cells in the experimental group was markedly increased (P < 0.05 or 0.01), and reached a peak 3 4 days later. The results confirmed that in vivo denaturing, in vitro hybrid enzyme digestion, tissue mass transplantation combined with low enzyme digestion, separation of double-differential adhesion of Schwann cells is a simple and rapid method to extract and purify Schwann cells.