ObjectiveTo detect the expression of motilin in gastric cancer tissues and to explore the relationship between motilin protein expression and clinicopathologic characteristics of gastric cancer. MethodsThe immunohistochemical staining was used to detect the expression of motilin protein in gastric cancer, paracancerous tissues, and normal gastric mucosa tissues. The relationship between motilin protein expression and clinicopathologic characteristics of gastric cancer was analyzed. ResultsThe expression of motilin protein in gastric cancer tissues (1 206.43±631.67) was significantly higher than that in normal gastric mucosa tissues and paracancerous tissues, respectively (Plt;0.01). The difference of motilin protein expression between normal gastric mucosa tissues and paracancerous tissues was not significant (Pgt;0.05). The expression of motilin protein in gastric cancer was correlated with the site of tumor, differentiation degree, and lymph node metastasis (Plt;0.05). ConclusionMotilin may participate in the carcinogenesis of gastric cancer, and correlated with the invasion and metastasis of gastric cancer.
OBJECTIVE: To observe the protein expression of phosphorylated form of P38 mitogen-activated protein kinase(P38MAPK) and c-Jun in hypertrophic scar skin and to explore their influences on the formation and maturation of hypertrophic scar. METHODS: The expression intensity and distribution of phosphorylated form of P38MAPK and c-Jun were examined with immunohistochemistry and pathological methods in 16 cases of hypertrophic scar skin and 8 cases of normal skin. RESULTS: In normal skin, the positive signals of phosphorylated form of P38MAPK mostly distributed in basal lamina cells of epidermis, while c-Jun was mainly located in epidermal cells and endothelial cells. The positive cellular rates of two proteins were 21.3% +/- 3.6% and 33.4% +/- 3.5% respectively. In proliferative hypertrophic scar skin, the particles of phosphorylated P38MAPK and c-Jun were mainly located in epidermal cells and some fibroblasts. The positive cellular rates of two proteins were significantly elevated to 69.5% +/- 3.3% and 59.6% +/- 4.3% respectively (P lt; 0.01). In mature hypertrophic scar, the expression of these proteins decreased but was still higher than that of normal skin. CONCLUSION: The formation and maturation of hypertrophic scar might be associated with the alteration of phosphorylated P38MAPK and c-Jun protein expression in hypertrophic scar.
Objective To explore the expression of the vascular cell adhesion molecule 1 (VCAM1) in the acellular dermal matrix grafting in pigs. Methods Experimental models were established with 15 Inbred Strain mini pigs, 6 full-thichness skin defect wounds, 6 cm × 6 cm in size, were produced on both-side backs of the each pig, and then the pigs were randomly divided into 3 groups. In Group A (n=5, control) the thin auto-skintransplantation alone was made; in Group B (n=5), the grafting was performed in the acellular allo-dermal matrix combined with the thin auto-skins; in Group C (n=5), the grafting was performed in the acellular xeno-dermal matrixcombined with the thin auto-skins. The areas of the wounds were measured and the survival condition of the grafted skins was observed at 3, 9, 21 and 30 days after the grafting. The histological samples were harvested from the grafting area at 3, 6, 9, 12, 21 and 30 days after the procedure. The flow cytometry was employed to analyze the changes in the VCAM1 level in the sample at different time points after the grafting. Results In the 3 groups, the transplanted skin base was easily separated at 3 days after transplantation; the areas of the wound healing accounted for 94%±12%，92%±9%, and 91%±11%, respectively, at 21 days; good wound healing was achieved at 30 days. At 9 and 12 days after transplantation, there was an evidentlyincreased level of the VCAM-1 expression in the tissue samples in the composite skin grafting groups. Compared with the control group, the difference was significant (Plt;0.05); however, the VCAM-1 expression at 3 days was not statistically different between the composite skin grafting groups and the control group after transplantation. In contrast, the level of the VCAM-1 expression was significantly higher at 6 days in the control group than in the composite skin grafting groups (Plt;0.05). The levels of the VCAM-1 expression were significantly lower at 30 days than at 3 days after transplantation in all the 3 groups (plt;0.01). Conclusion The highest level of the VCAM-1 expression can be delayed in the composite skin grafting when compared with that in the thin auto-skins alone, which implies that the VCAM-1 expression may be correlated with angiogenesis and composite skin survival. The VCAM-1 expression is not different between the acellular allo-dermal matrix composite skin grafting groups and the acellular xeno-dermal matrix group.
OBJECTIVE: To explore the expressive characteristics of epidermal growth factor (EGF) and its receptor (EGFR) in tissues of fetal and adult intestines. METHODS: The expression intensity and distribution of EGF and EGFR were detected with pathological and immunohistochemical methods in 6 specimens of adult (16-54 years) intestines and 18 specimens of fetal intestines with different gestational ages (13-31 weeks). RESULTS: Positive protein particles of EGF and EGFR could be detected in tissues of fetal and adult intestines. The protein expressions of EGF and EGFR were elevated progressively with the gestational age. EGF was mainly located in the cytoplasm and extracellular matrix of intestinal villus cells, endothelial cells and tunica serosa epithelial cells, while EGFR chiefly distributed in the cellular membrane of these cells. CONCLUSION: The endogenous EGF and EGFR might be involved in the intestinal development at embryonic stage, in the structural and functional maintenance at adult stage, and in the wound healing after injury.
To explore the expression of Wnt-1 during the process of inducing neural stem cells (NSCs) into neurons by using all-trans-retinoic acid (ATRA) in vitro and the effect of Wnt-1 on NSCs differentiation. Methods NSCs isolated from cerebral cortex of SD rat embryo (12-16 days’ gestation) were cultured. The concentration of cells at passage 3 were adjusted to 1 × 106 cells /mL and treated with ATRA at 0.5, 1.0, 5.0 and 10.0 μmol/L, respectively. Differentiation ratio of NSCsinto neurons in each group was detected by double-labelling immunofluorescence technique and flow cytometry, and 1.0 μmol/ L was selected as the best concentration for ATRA to promote NSCs differentiation. In experimental group, NSCs at passage 3 were cultured with ATRA at 1.0 μmol/L in vitro, and expression of Wnt-1 was detected by immunocytochemistry staining, realtime flurescent quantitive PCR and Western blot at 3, 5, 7 and 9 days after culture, respectively. The cells at passage 3 receiving no ATRA served as control group. Results Immunocytochemistry staining: in the control group, there was l ittle Wnt-1 protein expression; in the experimental group, peak expression of Wnt-1 and numerous positive cells occurred at 3 days after culture, the positive expression of Wnt-1 was still evident at 5 days after culture, and there was significant difference between two groups in integrated absorbance (IA) value at 3 and 5 days after culture(P lt; 0.05), obvious decrease of positive expression of Wnt-1 was evident, and no significant difference was evident between two groups in IA value at 7 and 9 days (P gt; 0.05). Real-time fluorescence quantitative PCR: the relative expression of Wnt-1 mRNA in the control group was 0.021 7 ± 0.072 1; the relative expression of Wnt-1 mRNA in the experimental group at 3, 5, 7 and 9 days was 0.512 2 ± 0.280 0, 0.216 4 ± 0.887 0, 0.038 5 ± 0.299 4 and 0.035 5 ± 0.309 5, respectively, indicating the value decreased over time, and there were significant difference between two groups at 3 and 5 days (P lt; 0.05), and no significant difference at 7 and 9 days (P gt; 0.05) . Western blot detection: specific and visible staining band was noted; in the control group, Wnt-1 protein expression was 0.005 1 ± 0.558 3; in the experimental group, Wnt-1 protein expression at 3, 5, 7 and 9 days was 0.451 7 ± 0.071 3, 0.311 7 ± 0.080 5, 0.007 3 ± 0.052 7 and 0.004 7 ± 0.931 4, respectively, suggesting the value decreased over time; there were significant differences between two groups at 3 and 5 days (P lt; 0.05), and no significant differences at 7 and 9 days (P gt; 0.05). Conclusion With the induction of ATRA at 1.0 μmol/L, Wnt-1 and NSCs differentiation in early stage are positively correlated. Its possible mechanism may rely on the activation of such signals as classic Wnt-1 signal pathway, indicating Wnt-1 relates to the differentation of NSCs into neurons.
【摘要】 目的 观察不同种培养基中重组人色素上皮衍生因子(rPEDF)融合蛋白的表达。 方法 将前期研究已构建的pET28aPEDF原核表达重组体转化E.coli BL21大肠杆菌表达宿主菌,酶切鉴定阳性菌落后,分别在M9和LB培养基中用异丙基βD硫代半乳糖(IPTG,IsopropylbetaDthiogalactoside)诱导表达,SDSPAGE电泳检测表达的PEDF蛋白, 美国ImagePro Plus 分析系统进行蛋白定量分析。结果 LB和M9培养基中均获得相对分子质量约54×103的rPEDF融合蛋白。但LB培养基获得的是rPEDF融合蛋白的包涵体,目的蛋白占总蛋白含量为21046%,M9培养基获得的是可溶性的rPEDF的融合蛋白,目的蛋白占总蛋白含量的1231%。结论 不同种培养基中均有rPEDF 融合蛋白的表达。【Abstract】 Objective To observe the express of recombinant pigment epithelial derivative facto (rPEDF) in the different medium. Methods The pET28aPEDF was transformed into E.coli BL21. After the colonies were positive identification which were induced by IsopropylbetaDthiogalactoside in medium M9 and LB. The PEDF protein were detected by SDSPAGE and analyzed by American ImagePro Plus system. Results LB and M9 medium obtained the relative molecular mass about 54×103 rPEDF fusion protein. But LB medium obtained the inclusion bodys of rPEDF fusion protein，the purpose protein account for 21.046%；LB medium obtained the soluble rPEDF fusion protein，the purpose protein account for 12.31%. Conclusion The rPEDF protein was expressed in the different medium.
Objective To observe the effects of vascular endothelial growth factor antisense oligonucleotide (VEGF-ASODN) on expression of vascular endothelial growth factor (VEGF) and growth in gastric cancer cells. Methods The VEGF-ASODN was synthesized artificially with phosphorothioic acid. After transfecting with VEGF-ASODN in gastric cancer cells SGC-7901, the initial copy number of mRNA was detected by real-time RT-PCR, and the quantity of VEGF protein in both cell and supernatant were detected by ELISA. The levels of expression of survivin protein in cells were measured by Western blot. FCM and MTT method were used to detect cellular apoptosis and the activity of cells, respectively. The effect of transfection on the growth of cells was evaluated by growth curve. Results The copy number of VEGR mRNA, protein levels of VEGF in the cells and in culture fluid all decreased when the concentration of transfected VEGF-ASODN increased, as well as the levels of survivin protein （P＜0.05）. The ratio of apoptosis increased, the activity of cells also decreased as the concentration of transfected VEGF-ASODN increased （P＜0.05）. Conclusion Transfection with VEGF-ASODN in gastric cancer cells SGC-7901 can inhibit the expressions of VEGF and survivin remarkably. It can enhance cellular apoptosis and suppress growth of cells.
Objective To investigate the expression of COX-2 in human cervical cancer and explore their relationship between the COX-2 expression and the clinicopathologic characteristic of cervical cancer. Methods The published studies were searched in the CBMdisc (1979 to 2009), CNKI (1979 to 2009), VIP (1989 to 2009) and WANFANG Database (1982 to 2009), and other relevant journals were also hand searched, to identify all the relevant case-control trials. The quality of the included studies was assessed. The Cochrane Collaboration’s software RevMan 4.2.10 was used to test the heterogeneity, overall effect and publication bias of the combined studies. Results A total of 9 studies were recruited. As for the positive rate of COX-2 expression, significant differences was tested between cervical cancer vs. normal cervical tissues, lymph node metastasi vs. non-lymph node metastasi, clinical stages I-II vs. clinical stages III-IV, cell differentiation G1 vs. cell differentiation G2-G3 and cervical squamous cell carcinoma vs. adenocarcinoma with OR (95%CI) at 28.03 (9.53 to 82.50), 5.16 (3.36 to 7.93), 0.53 (0.33 to 0.84), 3.11 (1.86 to 5.22) and 5.00 (2.68 to 9.35) respectively. Conclusions According to the domestic evidence, higher COX-2 expression might be associated with cervical cancer. However, more high quality case-control studies are expected for further study.
Objective To investigate the expression of enhancer of zeste homolog 2 （EZH2） mRNA in colorectal cancer tissues， and to explore the relationship with clinicopathological features of it. Methods PCR was used to examine the expression of EZH2 mRNA in colorectal cancer tissues， paracancerous tissues， and normal epithelium tissues of 27 patients with colorectal cancer. Meanwhile， the relationship between the expression of EZH2 mRNA and clinicopathological features of colorectal cancer were analyzed. Results Expression level of EZH2 mRNA in colorectal cancer tissues， paracancerous tissues， and normal epithelium tissues were 3.01±1.29， 1.20±0.69， and 0.87±0.37 respectively， and the expression value of colorectal cancer tissues at the top （P＜0.01）， but there were no significant difference between the paracancerous tissues and normal epithelium tissues （P=0.403）. Expression of EZH2 mRNA were related to TNM stages （P=0.002） and degree of differentiation （P=0.005）， but were not related to age （P=0.388）， gender （P=0.963）， diameter of tumor （P=0.579）， histological type （P=0.945）， location of tumor （P=0.611）， and lymph node metastasis （P=0.449）. Conclusions EZH2 mRNA expresses highly in colorectal cancer tissues， so it may play an essential role in the emergence and development of colorectal cancer. EZH2 mRNA can be used as one of the referenced indexes to determine the malignant degree and process of colorectal cancer.
Objective To construct the recombined DNA pcDNA3.1-hBMP-2 and transfect into human marrow stromal stem cells (MSCs) in vitro, and to explore theeffects of transfection on cellular proliferation and expression of vascular endothelial growth factor (VEGF). Methods The expression of human bone morphogenetic protein 2(hBMP-2) in these cells after transfection was determined by in situ hybridization and immunohistochemical analysis and Western blot analysis. The changes of cell proliferation were observed by flow cytometry. The effects of BMP-2 gene transfection on expression of VEGF in the cells were analyzed by in situ hybridization of VEGF cDNA probe. Results Stable expressionof hBMP-2 in pcDNA3.1-hBMP-2 transfected MSCs was confirmed in the levels of mRNA and protein.Cellular proportion in S period increased, which indicated that the synthesis of cell DNA increased. The expression of VEGF in the cells increased obviously. Conclusion With the help of lipofectamine, the pcDNA3.1-hBMP-2 were transfected into human MSCs successfully. hBMP-2 plays an important role in promoting cellular proliferation and vascular generation during bone repair.