AIM:To evaluate the influence of scutellarin on the expression of vascular endothelial growth factor ( VEGF) in high glucose-treated human retinal pigment epithelial cell line ARPE-19 and to observe the effects of scutellarin on the protein expression of VEGF, p-ERK and VEGFR2 in the retinas of type II diabetic rats.METHODS: Cultured ARPE-19 cells were divided into normal control group, scutellarin group, high glucose group and high glucose+scutellarin group.The protein levels of VEGF, p-ERK and VEGFR2 were measured by Western blot.The VEGF release in ARPE-19 cells was detected by ELISA.Normal rats were randomly divided into normal control group and scutellarin group.Diabetic rat model was established by feeding with high-fat diet and injecting with streptozocin, and randomly divided into diabetes group and diabetes treated with scutellarin group.After 16 weeks, the eyes were removed.The morphological changes of the retinas were observed under light microscope with HE staining, and histopathological score was recorded.The expres-sion of VEGF in the retinas was observed by the method of immunohistochemistry.RESULTS:Compared with normal con-trol group, the protein levels of VEGF, p-ERK and VEGFR2 in the ARPE-19 cells decreased in scutellarin group, but in-creased in high glucose group.The histopathological score of the retinas showed significant difference among diabetes group, diabetes treated with scutellarin group and normal control group, and no significant difference between normal con-trol group and scutellarin group was observed.The expression of VEGF increased in diabetic group and was significantly higher than that in scutellarin treatment group (P<0.05).CONCLUSION:Scutellarin inhibits the increased protein le-vels of VEGF, p-ERK and VEGFR2 in ARPE-19 cells, and decreases the expression of VEGF in the retinas of diabetic rats.The suppression of the diabetic retinopathy development by scutellarin may be partly involved in the ERK/MAPK pathway.

BACKGROUND:Hyperbranched cationic amylopectin is a kind of nonviral gene vectors with low toxicity and good transfection efficiency. However, searching for more efficient methods to delivery it into the body and making the genes expressed are being explored.
OBJECTIVE:To study the expression of DMAPA-Amp wrapped green fluorescent protein (GFP) transferred by modified carotid injection into cerebral ischemic area.
METHODS:Male Sprague-Dawley rats subjected to middle cerebral artery infarction were randomly divided into two groups after 24 hours:experimental group (injected with GFP entrapped DMAPA-Amp via the internal carotid artery) and control group (injected with GFP entrapped DMAPA-Amp via the tail vein). These rats were put to death and their brain tissue was removed after 7 days. The expression of GFP was detected by quantitative PCR and western blot assay, and immunofluorescence staining was performed to detect the expression of GFP located near cerebrovascular endothelial cel s by frozen section.
RESULTS AND CONCLUSION:Compared with the control group, the expression of GFP was much higher in the experimental group detected by quantitative PCR and western blot (P< 0.05). Additional y, the expression of GFP located near cerebrovascular endothelial cel s by frozen section was also higher than that in the control group. Modified carotid injection could significantly promote the expression of hyperbranched cationic amylopectin derivates and GFP in the brain tissue of Sprague-Dawley rats undergoing middle cerebral artery infarction compared with tail vein injection, which indicates DMAPA-Amp and modified carotid injection may cast new lights on the therapy for angiogenesis of ischemic stroke.

Objective To investigate the expression of FTO in gastric cancer tissues and the functional significance of FTO in MGC-803 cell line.Methods The FTO mRNA was detected by RT-qPCR in 54 cases of gastric cancer samples and their paired adjacent normal control tissues.The effect of FTO overexpression in MGC-803 on cell proliferation,cell migration and invasion were detected by CCK-8,wound heal and ranswell assays,respectively.Results The expression of FTO mRNA was significantly lower than that of adjacent tissues(P＜O.05).Furthermore,overexpression of FTO in MGC-803 cells inhibited cell proliferation,cell migration and invasion.Conclusions FTO is low expressed in gastric cancer tissues and inhibits gastric cancer cell line MGC-803 proliferation,migration and invasion.FTO is closely associated with the development of gastric cancer.

Objective:To explore the protective effect and related mechanism of rebamipide on non-steroid anti-inflammatory drug (NSAID) related small intestinal mucosal injury.Methods:A total of 21 C57BL/6 mice were selected and by random number table method, they were divided into negative control group (0.9% NaCl gavage for four days), indomethacin modeling group (20 mg/kg indomethacin gavage for four days) and rebamipide intervention group (20 mg/kg indomethacin gavage for four hours and then 320 mg·kg -1·d -1 rebamipide gavage for four days), seven mice in each group. After modeling, the injury of mice intestinal mucosa of indomethacin modeling group and rebamipide intervention group was evaluated by gross observation as well as pathological analysis. The serum levels of interleukin (IL)-6, IL-10, trefoil factor 3 (TFF3), prostaglandin E2 (PGE2) and epidermal growth factor (EGF) in mice were detected by enzyme-linked immunosorbent assay (ELISA). The expression of IL-6, IL-10, TFF3, cyclooxygenase 2( COX2) and EGF at mRNA level of mice small intestinal tissues were examined by real-time quantitative polymerase chain reaction (qRT-PCR). And the relative expression of TFF3, COX2 and EGF at protein level of mice small intestinal tissues were determined by Western blotting. Levene test and independent sample t test were used for statistical analysis. Results:The scores of gross observation and histopathology of mice small intestinal mucosa injury of rebamipide intervention group were both lower than those of indomethacin modeling group (2.80±0.45 vs. 4.60±1.14, 1.67±0.52 vs. 3.00±0.71), and the differences were statistically significant ( t=2.667 and 3.618, P=0.029 and 0.006). The mouse serum level of IL-6 and the expression of IL-6 at mRNA level in intestinal tissues of indomethacin modeling group were both higher than those of the negative control group, however the serum level of IL-10 was lower than that of the negative control group ((48.83±5.40) ng/L vs. (40.96±5.92) ng/L, 5.23±2.36 vs. 1.12±0.56, (168.50±10.57) ng/L vs. (186.30±7.77) ng/L), and the differences were statistically significant ( t=2.307, 3.372 and 3.366; P=0.047, 0.007 and 0.012). The expression of IL-6 at mRNA level in mice small intestinal tissues of rebamipide intervention group was lower than that of indomethacin modeling group (1.74±0.82 vs. 5.23±2.36), however, the expression of IL-10 at mRNA level was higher than that of indomethacin modeling group (6.44±3.46 vs. 1.22±0.83), and the differences were statistically significant ( t=3.409 and 3.025, P=0.008 and 0.014). The serum levels of TFF3, PGE2 and EGF, the expression of TFF3 at mRNA level of small intestinal tissues, the relative expression of COX2 and EGF at protein level of small intestinal tissues of indomethacin modeling group were all lower than those of the negative control group ((131.20±16.37) ng/L vs. (150.30±9.66) ng/L, (32.68±6.88) ng/L vs. (41.51±3.20) ng/L, (112.70±17.17) ng/L vs. (138.20±10.10) ng/L, 0.43±0.22 vs. 1.20±0.50, 0.33±0.25 vs. 1.30±0.43, 0.28±0.19 vs. 1.15±0.10), and the differences were statistically significant ( t=2.290, 2.645, 2.867, 3.097, 3.405 and 7.106; P=0.048, 0.021, 0.025, 0.017, 0.027 and 0.002). The mice serum levels of PGE2 and EGF, expression of TFF3, COX2 and EGF at mRNA level of small intestinal tissues, as well as the expression of TFF3 and EGF at protein level of small intestinal tissues of rebamipide intervention group were all higher than those of indomethacin modeling group ((43.55±5.28) ng/L vs. (32.68±6.88) ng/L, (153.30±15.66) ng/L vs. (112.70±17.17) ng/L, 2.48±1.70 vs. 0.43±0.22, 2.95±1.56 vs. 0.88±0.45, 3.97±2.54 vs. 0.98±0.76, 1.47±0.26 vs. 0.72±0.35, 1.08±0.36 vs. 0.28±0.19), and the differences were statistically significant ( t= 2.711, 3.658, 2.656, 2.856, 2.524, 3.013 and 3.435; P=0.024, 0.008, 0.026, 0.019, 0.033, 0.039 and 0.026). Conclusions:Rebamipide alleviates small intestinal mucosal injury induced by indomethacin by inhibiting the expression of inflammatory factors and promoting the expression of intestinal mucosal protective factors suggesting that rebamipide plays a protective role in NSAID related small intestinal injury by maintaining the chemical barrier of the intestinal mucosal.

Objective To study the growth inhibition,apoptosis induction effects of cinobufagin(CB)on human osteosarcoma(OS) cell line U2OS,MG63 and SaO2 in vitro and the underlying mechanism of action of cinobufagin in OS cells.Methods Cell viability was assessed by MTT assay.Cell-cycle status,apoptosis-inducing effects were evaluated by flow cytometry,fluorescent staining and DNA fragmentation assays.Inhibitors of apoptosis proteins (IAPs) and Bcl-2 family proteins including Bax,cleaved-PARP,xIAP,cIAP-1,survivin and p65 were tested by Western blot.Results MTT assay showed that CB could inhibited the growth of U2OS,MG63 and SaO2 cells in a dose-and time-dependent manner.The 48 h IC50 of CB on U2OS,MG63 and SaO2 cells were (104.83± 16.96) nmol/L,(47.07±7.5) nmol/L,and (136.72±10.08) nmol/L respectively.The induction of G2/M cell-cycle arrest was seen in the cells treated with CB.After cells were cultured for 12 h in the presence of 100 nmol/L CB,the percentages of cells in the G0/G1 phase were decreased,while G2/M phase were increased in U2OS,MG63 and SaOS2 cells,respectively.The results showed CB inhibited the proliferation of osteosarcoma cells through blocking the cell cycle in G2/M phase.Induction of apoptosis was confirmed by Hoechst 33258 and Annexin V/PI staining.After treating with 100 nmol/L CB for 48 h,the extents of apoptosis were 33.6％±6.4％,36.4％±7.8％ and 29.3％±5.1％,respectively.These results indicate that the anti-tumor activity of cinobufagin in osteosarcoma cells was due to a G2/M cell cycle arrest and apoptosis inducing effect.Western blot showed that CB could induce the apoptosis related family proteins Bax,cleaved-PARP up-regulation,xIAP,cIAP-1,survivin and p65 downregulation in OS cells.Conclusion CB can inhibit the cell viability and induce G2/M cell cycle arrest and apoptosis in U2OS,MG63 and SaO2 cells.The apoptosis-inducing effect of CB is confirmed by the regulation of apoptosis related proteins IAPs and Bcl-2 in vitro.

Background: Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy, with a high incidence and poor prognosis. In the past several decades, hundreds of proteins have been reported to be associated with the prog-nosis of ESCC, but none has been widely accepted to guide clinical care. This study aimed to identify proteins with great potential for predicting prognosis of ESCC. Methods: We conducted a systematic review on immunohistochemical (IHC) prognostic markers of ESCC accord-ing to the 2009 Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) Guidelines. Literature related to IHC prognostic markers of ESCC were searched from PubMed, Embase, Web of Science, and Cochrane Library until January 30th, 2017. The risk of bias of these original studies was evaluated using the Quality in Prognosis Studies (QUIPS) tool. Results: We identified 11 emerging IHC markers with reproducible results, including eight markers [epidermal growth factor receptor (EGFR), Cyclin D1, vascular endothelial growth factor (VEGF), Survivin, Podoplanin, Fascin, phosphorylated mammalian target of rapamycin (p-mTOR), and pyruvate kinase M2 (PKM2)] indicating unfavorable prognosis and 3 markers (P27, P16, and E-cadherin) indicating favorable prognosis of ESCC. Conclusion: Strong evidence supports that these 11 emerging IHC markers or their combinations may be useful in predicting prognosis and aiding personalized therapy decision-making for ESCC patients.

Objective To study the effects of Interferon(IFN)in chronic hepatitis B patients genotype B and C.Methods 20 cases were genotype B ,23 cases were genotype C,all the patients were treated with 5 million units of IFN-a-lb,im,qod,for 12 months,viral markers,liver function and adverse drug reactions were observed.Results There was no statistically significant difference between B,C genotype in the negative conversion rate of HBV-DNA (60.0% ,39.1%),the negative conversion rate of HBeAg(42.9% ,30.8%),anti-HBe seroconversion rate(35.7% ,23.1 %)and the rate ALT normalization(85.0% ,73.9%)(t = 1.86,0.69,0.68 ,0.79,P ＞ 0.05).Conclusion The results suggested that therapeutic efficacy of IFN-α-lb was not significantly different between B,C genotypes.

Objective To evaluate the long following-up outcome of the medial gastrocnemius muscle transferring reconstruction the patella tendon after the wide resection of aggressive bone tumors in the proximal tibia. Methods With the 69 patients of the osteogenetic sarcoma in the proximal tibia were treated with the wide resection and reconstruction the patella tendon. After the long following up the knee extensor,function and complications were evaluated. Results With the 69 patients, the 45 survival patients were followed up for the average 68.6 (24-128) months. The local recurrence rate was 8.7%(6/69). The strength of knee extending was in the average of grade 4.2(3.6-5.0), the degree of knee flexion was in the average of 95°(75°-135°), the degree of knee extension was in the average of-2°(0°-12°), the knees of five patients cannot fully extension. The MSTS functional score was in the average of 77% (23.1/30). Conclusion During the limb salvage of the proximal tibial aggressive bone tumors, the medial gastrocnemius muscle transferring reconstruction the patella tendon could offer the knee extension strength; improve the soft tissue coverage and functional results.

Objective To characterize and compare the different biological behaviors of two novel human osteosarcoma cell lines, Zos and Zos-M, established respectively from the primary site and the skip metastasis of an osteosarcoma patient. Methods Two novel human osteosarcoma cell lines, Zos and Zos-M,were developed using tissue plant culture method. The vitro examinations included observations of morphology, analysis of karyotype and cell cycle, calculation of doubling time and growth curve, detection of osteoblastic markers and matrigel invasion assay. Subcutaneous, intratibial and intravenous inoculations into nude mice were performed to study the in vivo tumorigenicity and metastatic potentials of both cell lines.MTT were used to detect sensitivity of the cell lines to chemotherapeutic drugs. RT-PCR was performed to assess the expression of and some metastasis-related genes. Results Both cell lines proliferated actively and remained stable for more than 100 passages in vitro without interruption. The morphology and expression of osteoblastic markers of Zos and Zos-M were conformed to the characteristic of osteosarcoma. The karyotype analysis displayed aneuploidy and various structural abnormalities. The population doubling time of Zos and Zos-M were 33.65 h and 31.58 h respectively. Both cell lines were less sensitive to the current chemotherapy protocols compared to U-2OS. Zos and Zos-M were 100% tumorigenic by subcutaneous and othotopic injection. 37.5% of nude mice injected Zos-M and none of nude mice injected Zos developed lung metastasis after intravenous injection. The comparison of the expression patterns of some metastasis-related genes revealed that the decreased expression of cadherin-11 in Zos-M may correlate with a high potential of metastases. Conclusion The two novel established human osteosarcoma cell lines, Zos and Zos-M and related animal models could serve as models for the study of drug resistance and screening of new therapeutics for osteosarcoma. In addition, the study also provide tools for the study of metastasis because the same genetic background and different potential of metastasis of Zos and Zos-M.