Expression of mic2/CD99 protein and their correlation with Eber-1/LMP-1 in Hodgkin and Reed-Sternberg cells of classical Hodgkin's lymphoma

AIM: To study the expression of mic2/CD99 protein and their correlation with Eber-1/LMP-1 in Hodgkin and Reed-Sternberg cells of classical Hodgkin's lymphoma. METHODS: Immunohistochemical staining, in situ hybrization and tissue microarry technique were used to detect the expressions of mic2/CD99 and Eber-1/LMP-1 of H/RS cells in 43 cases of cHL and 16 cases of NHL. RESULTS: The positive rate of CD99 protein expression in 43 cases of cHL was 2.3％ (1/43), mic2 was 55.8％ (24/43), LMP1 was 58.1％ (25/43) and Eber-1 was 53.5％ (23/43). The expressions of CD99 and mic2 in the NHL group were higher than those in cHL group (P<0.05). The expressions of LMP1 and Eber-1 in NHL group were lower than those in cHL group (P<0.05). The expression of mic2 was higher than CD99 in cHL group (P<0.05). There was no difference between the expression of Eber-1 and LMP1 in cHL group statistically (P>0.05). There was a negative correlation between the expression of CD99 protein and LMP1 in H/RS cells (P<0.05). No correlation was found between the expression of any other markers in H/RS cells and patients' sex (P>0.05). There was a significant correlation between the high expression of LMP1 and a low expression of CD99 in the young patients (P<0.05), but no correlation was found between the expression of mic2 and Eber-1 in young patients (P>0.05). CONCLUSION: There is a negative correlation between the expression of LMP1 and CD99 in Hodgkin and Reed-Sternberg cells of classical Hodgkin's lymphoma.     

Effects of adenovirus-mediated shRNA targeting PTEN on proliferation and apoptosis of activated hepatic stellate cells in vitro

AIM:To investigate the down-regulation of phosphatase and tensin homolog deleted on chromosome 10(PTEN) gene by adenovirus-mediated short hairpin RNA(shRNA) on proliferation and apoptosis of activated hepatic stellate cells(HSCs) in vitro and the related signaling transduction pathways. METHODS:The activated HSCs were cultured in vitro and transfected with recombinant adenovirus expressing shRNA targeting PTEN. The proliferation of HSCs was measured by MTT assay and the apoptosis was assessed by TUNEL and flow cytometry. Western blotting was used to detect the protein levels of PTEN, Bax, Bcl-2, Akt, p-Akt, ERK1/2 and p-ERK1/2 in HSCs, and real-time fluorescent quantitative PCR was applied to detect the mRNA expression of Akt and ERK1. RESULTS:The recombinant adenovirus expressing shRNA targeting PTEN was successfully transfected into activated HSCs in vitro, and significantly promoted the proliferation of HSCs in a time-dependent manner within a certain extent. The apoptotic rate of HSCs was significantly decreased 72 h after transfection(P<0.05). Meanwhile, reduced expression of Bax and elevated expression of Bcl-2 were induced 72 h after transfection(P<0.05). Furthermore, the expression of p-Akt and p-ERK1/2 were increased significantly(P<0.05), while no significant difference in the expression of Akt and ERK1 at mRNA and protein levels was observed(P>0.05). CONCLUSION:Down-regulation of PTEN by adenovirus-mediated shRNA dramatically promotes the proliferation of activated HSCs, and inhibits the apoptosis through Bcl-2/Bax pathway. In addition, the phosphorylation of Akt and ERK1/2 is increased, indicating that PI3K/Akt and ERK1/2 signal transduction pathways may play an important role in the regulation of proliferation and apoptosis of HSCs.     

Expression and function of PTEN in fibroblast-like synoviocytes of rheumatoid arthritis

AIM: investigate the expression of PTEN gene in fibroblast-like synoviocytes (FLS) of rheumatoid arthritis (RA).METHODS: FLS were isolated from synovial tissue obtained from the patients with RA, osteoarthritis (OA) or joint trauma. The mRNA expression of PTEN was detected by RT-qPCR. The protein levels of PTEN, p-Akt (Thr308) and total Akt were determined by Western blotting. The phosphorylation status of Akt was analyzed by the protein ratio of p-Akt (Thr308)/total Akt.RESULTS: The mRNA expression of PTEN was significantly lower in RA-FLS than that in OA-FLS and joint trauma-FLS (P < 0.01), while no statistically significant difference was observed between that in OA-FLS and joint trauma-FLS (P < 0.05). Similarly, the protein expression of PTEN in the RA-FLS was much lower than that in the OA-FLS and joint trauma-FLS (P < 0.05), and there was no difference between the latter 2 groups. Moreover, the phosphorylation level of Akt (Thr308) in the RA-FLS was significantly higher than that in the other 2 control groups (P < 0.01), and that in OA-FLS was much lower than that in the joint trauma-FLS (P < 0.01). Finally, Pearson correlation analysis between the phosphorylation level of Akt (Thr308) and PTEN protein expression in the RA-FLSs showed a significant negative correlation (r=-0.994 5, P < 0.01).CONCLUSION: The mRNA and protein expression of PTEN are both decreased in the RA-FLS, which may contribute to the increased phosphorylation level of Akt (Thr308).     

Expression of drug resistance related proteins in osteosarcoma and soft tissue sarcoma and its correlations with chemotherapy resistance

AIM: To explore the expression of four kinds of drug resistance related protein: P-glycoprotein (P-gp), glutathione-S-transferases-π (GST-π), lung resistance protein (LRP), multidrug resistance related protein (MRP) in osteosarcoma cell lines Saos2 and U2OS, and in osteosarcoma and soft tissue sarcoma tissues from 34 patients and their correlations with chemotherapy resistance.METHODS: The expression of protein was detected by flow cytometry (FCM).Chemotherapy resistance was analyzed by MTT assay.RESULTS: Expression of drug resistance related protein was lower in Saos2 cell line than that in U2OS cells.Chemotherapy sensitivity on adriamycin (ADM), cisplatinum (DDP), fluorouracilum (5-Fu), mitomycin (MMC), dacarbazine (DTIC), vincristine (VCR) was higher in Saos2 than those in U2OS cells.After the two cell lines were treated respectively with 1/5 50% inhibitory concentration (IC50) doses of ADM and DDP, the expression increasing range of GST-π was 33%-43%.The nonsensitive rate on ADM, DDP, 5-Fu, MMC, DTIC, VCR, methotrexate (MTX) in 34 patients were 41.18％, 17.65％, 47.06％, 50.00％, 76.47％, 61.76％ and 52.94％, respectively.The expression of P-gp, GST-π, LRP, MRP were 1.54, 2.58, 1.91 and 1.86, respectively.The correlation analysis showed that the expression of P-gp had positive correlation with resistance on ADM (r=0.485, P<0.01), the expression of GST-π had correlations with resistance on ADM, DDP, 5-Fu and MMC.The r value was 0.402, 0.458, 0.364 and 0.500,respectively.The corresponding value of P was <0.05, <0.01, <0.05 and <0.01,respectively.The observed expression of resistance related proteins was not significant difference between different gender, different age, different pathological type and different tumor size in osteosarcoma and soft tissue sarcoma patients (P>0.05).The expression of GST-π was increased dramatically in patients receiving chemotherapy preoperatively.The preoperative level of GST-π was higher in recurrent patients by follow-up survey than that in patients without recurrence (P<0.05).CONCLUSIONS: Individual difference was obvious in chemotherapy sensitivity and expression of resistance related proteins in different patients.Chemotherapy induced the upregulation of GST-π expression.Primary high expression of GST-π was the main mechanism of resistance in osteosarcoma and soft tissue sarcoma patients, and related with prognosis.     

Effects of IL-6 and AG490 on growth of diffuse large B-cell lymphoma and Burkitt lymphoma transplanted into nude mice

AIM: To observe the effects of interleukin-6 (IL-6) and AG490 on diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma (BL) transplanted into nude mice, and to explore the effects of STAT3 activation on growth of these kinds of lymphoma in nude mice and its related mechanisms. METHODS: The nude mouse models with DLBCL and BL were established by transplantation with OCI-LY8 cells and Raji cells, respectively, and were divided into 3 groups:control group, IL-6 group and AG490 group. The body weight of mice and tumor size were measured. Western blot and immunohistochemical staining were used to detect the protein levels of p-STAT3, survivin and vascular endothelial growth factor (VEGF), and real-time PCR was used to detect the mRNA expression of survivin and VEGF. RESULTS: The tumorigenic rate of 2 kinds of tumor cell lines in nude mice was 83.3% (25/30) totally. The tumorigenicity of OCI-LY8 cells (66.7%, 10/15) was significantly lower than that of Raji cells (100%, 15/15) (P<0.05). The tumor size and body weight on days 9 and 10 in IL-6 group increased as compared with the control group, and the total difference value of tumor size between day 1 and day 10 in IL-6 group was obviously larger than that in control group (P<0.05). The positive protein of p-STAT3 was found in the nucleus, while the positive expression of survivin and VEGF was found in the cytoplasm. As compared with control group, the expression of survivin and VEGF was significantly increased (P<0.05), while the protein level of p-STAT3 was not significantly increased in IL-6 group of DLBCL. The protein levels of p-STAT3 and VEGF were significantly decreased (P<0.05), while the expression of survivin did not significantly decreased in AG490 group of DLBCL. The p-STAT3 and VEGF levels significantly increased (P<0.05) in IL-6 group of BL, while the levels of 3 kinds of proteins significantly deceased (P<0.05) in AG490 group of BL, as compared with control group. No statistical difference of mRNA expression of survivin and VEGF among IL-6, AG490 and control groups was observed. CONCLUSION: IL-6 and AG490 affect the growth of DLBCL and BL through activation of STAT3 pathway. The activated STAT3 participates in pathogenesis and progress of DLBCL and BL by up-regulating the expression of survivin and VEGF.     

Effect of over-expression of transcription factor CDX2 on proliferation and cell cycle of human gastric cancer cell line SGC-7901

AIM: To study the effect and the molecular mechanism of CDX2 over-expression on the proliferation, growth and cell cycle of human gastric cancer cell line SGC-7901. METHODS: The SGC-7901 cells in LV-CDX2-GFP group were transfected with the recombinant lentivirus vector LV-CDX2-GFP, the cells in LV-GFP group were transfected with the negative control lentiviral vector for the negative control, and the cells in blank control group were without any treatment. The cell proliferation was detected by CCK-8 assay. The cell cycle distribution was analyzed by flow cytometry. The expression of CDX2, Bax, Bcl-2, cyclin D1 and survivin was determined by semi-quantitative RT-PCR and Wes-tern blotting. RESULTS: Compared with LV-GFP group and blank control group, the proliferation activity of the SGC-7901 cells was significantly lower (P<0.05), the G0/G1 phase proportion increased (P<0.05), the mRNA and protein levels of Bcl-2, cyclin D1 and survivin were reduced (P<0.05), and the mRNA and protein levels of Bax were up-regulated (P<0.05) in LV-CDX2-GFP group. No statistically significant difference of the above indexes was observed (P>0.05) between LV-GFP group and blank control group. CONCLUSION: Over-expression of CDX2 mediated by lentivirus inhibits the proliferation and growth of human gastric cancer SGC-7901 cells and arrestes the cell cycle at G0/G1 phase, which may be related to down-regulation of Bcl-2, cyclin D1 and survivin and up-regulation of Bax.     

Berberine enhances mitomycin C-induced cell cycle arrest and apoptosis of T24 bladder cancer cells

AIM: To observe the effects of the combination of berberin (Ber) and mitomycin C (MMC) on the cell cycle arrest and apoptosis of T24 bladder cancer cells and the underlying mechanisms. METHODS: The T24 cells were exposed to MMC in the presence or absence of difference concentrations of Ber. The viability of the T24 cells was determined by CCK-8 assay. The cell cycle distribution was detected by flow cytometry. The apoptosis was analyzed by flow cytometry with Annexin V-FITC/PI staining, and the protein expression levels of cyclin D1, survivin, CDK2, CDK4, p21 and p27 were determined by Western blot. RESULTS: CCK-8 experiments showed that Ber enhanced the inhibitory effect of MMC on the viability of T24 cells. The results of flow cytometry showed that Ber also enhanced the blockade effect of MMC on T24 cells in G₀/G₁ phase (P<0.05). Compared with the MMC group, Ber increased the expression of p21 and p27 up-regulated by MMC, and decreased the expression of cynlin D1, CDK2 and CDK4 (P<0.05). Meanwhile, Ber promoted MMC to inhibit the expression of survivin (P<0.05). Ber increased the apoptosis of T24 cells induced by MMC (P<0.05). CONCLUSION: Ber significantly enhances the inhibitory effect of MMC on the viability of T24 cells. The mechanism may be related to up-regulation of p21 and p27, thereby inhibiting the expression of cyclin D1, CDK-2 and CDK-4. At the same time, Ber inhibits the protein expression of survivin, which eventually leads to cell arrest in G₀/G₁ phase and promotes apoptosis.     

miR-221 changes resistance of lung cancer cells to gefitinib by regulating PTEN

AIM: To explore the effect of microRNA-221 (miR-221) on resistance of lung cancer cells to gefitinib, and to investigate its related mechanism. METHODS: RT-qPCR was used to detect the levels of miR-221 expression between gefitinib-sensitive cell line PC9 and gefitinib-resistant cell line PC9/GR. The PC9/GR cells were transfected with miR-221 inhibitor by Lipofectamine 2000. The drug sensitivity of these cells to gefitinib was determined by CCK-8 assay. The protein expression level of phosphatase and tensin homologue deleted on chromosome ten (PTEN) was determined by Western blot. The 3'-UTR of PTEN was cloned into luciferase reporter vector and its luciferase activity was detected to verify whether miR-221 targets PTEN. RESULTS: The expression level of miR-221 in the PC9/GR cells was significantly higher than that in the PC9 cells (P<0.05). The protein expression level of PTEN in the PC9/GR cells was lower than that in the PC9 cells (P<0.05). The IC₅₀ of gefitinib was significantly reduced in the PC9/GR cells after transfection with miR-221 inhibitor (P<0.05). The protein expression level of PTEN in the cells transfected with miR-221 inhibitor was increased as compared with control group and blank group (P<0.05). Inhibition of miR-221 expression enhanced the enzymatic activity of luciferase reporter vector of PTEN. CONCLUSION: miR-221 enhances the resistance of lung cancer cells to gefitinib by down-regulating the protein expression of PTEN.     

Expression of Tpl-2 in colorectal carcinogenesis

AIM: To analyze the relationship between Tpl-2 (tumor progression locus 2)expression and clinicopathological parameters of colorectal carcinoma by investigating the expression of Tpl-2 in adjacent normal mucosa, colorectal adenomas and colorectal carcinoma. METHODS: Tpl-2 expression in normal mucosa, adenoma and carcinoma was examined and compared in a set of tissue microarrays by immunohistochemistry. The potential relationship between Tpl-2 expression and clinicopathological features was analyzed. RESULTS: The expression of Tpl-2 in carcinoma was significantly increased compared to the adenoma and normal mucosa (P<0.01). No significant difference was detected between the adenoma and normal mucosa (P>0.05). Meanwhile, the correlation between Tpl-2 expression and lymph node metastasis (N stage) and TNM stage (P<0.05) was observed. However, the correlation between the Tpl-2 expression and clinicopathological features of colorectal cancer including sex, age, body mass index (BMI), tumor size, histological differentiation, invasive depth (T stage),distant metastasis(M stage) and K-ras mutation (P>0.05) was not found. CONCLUSION: Tpl-2 has a relevance to the development of colorectal cancer as a promotive factor in the colorectal carcinogenesis.     

Prognostic prediction of PTEN and Her-2 expression in breast cancer

AIM:To study the prognositic value of PTEN and Her-2 expression in primary breast cancer. METHODS:81 breast cancer specimens with 15 years follow-up were obtained from 1989 to 2004. Immunohistochemical methods were used to detect the expression of PTEN and Her-2 in 81 paraffin-embedded specimens. The correlation between expression of PTEN and clinipathological factors was discussed with the Chi-square test. The survival rate analysis results were calculated with Kaplan-meier method.Long-rank test and Cox model by SPSS 10.0 software. 〖JP+1〗RESULTS:(1) PTEN expression significantly affects 5-year and 10-year survival rate of breast cancer (P<0.01 and P<0.05), and significantly negative correlation with the Her-2 expression was observed. (2) The patients with negative PTEN combined with positive Her-2 expression had worse prognosis. (3) Tumor sizes, postoperative therapy and PTEN expression had significant relation with the 5-year survival rate (P<0.05), and ER, Her2 and PTEN expression had significant relation with the 10-year survival rate (P<0.05). CONCLUSION:PTEN and Her-2 expression significantly affects 5-year and 10-year survival rate in patients with breast cancer, and may be biomarkers and important prognostic factors in breast cancer.     

芍药切花贮藏后水分与膜脂过氧化的研究

试材为芍药‘莲台’品种 (Paeonialactiflora‘Liantai’) ,采自山东荷泽 ,在花蕾萼片疏松、外层花瓣显现真正花色时采样 ,用水洗去叶片和花蕾上的分泌物 ,采后 40h火车运到实验室。在水中剪截成花枝长 35cm ,留 2～ 3片复叶 ,基部在 15cm深水中浸 2h复水 ,10 0 0倍的百菌清浸泡 1min ,晾干 ,普通白纸包裹后封入聚乙烯塑料袋中 ,于 0～2℃干藏。定期取样测定花瓣各项指标 ,均测 3次重复。定期取 5枝花在蒸馏水中瓶插 (室内散射光 ,温度 2 5℃±3℃ ,相对湿度 40 %～ 6 0 %) ,每天称花枝和 (水 +瓶 )质量 ,计算吸水量和失水量 ,记录瓶插寿命…     

Effects of intermittent high glucose on apoptosis and PTEN expression in islet cells

AIM: To investigate whether the increase in PTEN expression is related to apoptosis, and whether it is regulated by reactive oxygen species(ROS). METHODS: The rat islet cells were divided into constant low glucose group (group L), constant high glucose group (group H), glucose fluctuation group (group F), low glucose after high glucose group (group HL) and low glucose after fluctuation group (group FL). The ROS level, apoptotic rate, intracellular calcium, insulin release and PTEN protein expression were analyzed. RESULTS: Compared with groups H and L, the insulin secretion decreased, and intracellular calcium, ROS level, PTEN protein expression and apoptotic rate increased in group F (P<0.05). Compared with group H, the intracellular calcium, ROS level, PTEN protein expression and apoptotic rate in group HL decreased, but were still higher than those in group L (P<0.05). Compared with group F, the intracellular calcium, ROS level, PTEN protein expression and apoptotic rate in group FL decreased, but were still higher than those in group L (P<0.05). CONCLUSION: Glucose fluctuation can cause the apoptosis of islet cells more easily than constant high glucose. This may be related to the change of intracellular calcium and increase in oxidative stress which promotes PTEN expression. The recovery of glucose level to some extent relieves oxidative stress, decrease PTEN expression and reduce cell damage.     

Effect of mouse dermis-derived mesenchymal stem cells on expression of collagen and TGF-β1 in scleroderma fibroblasts

AIM: To investigate the change of collagen component and the expression of TGF-β₁ in the co-culture of mdMSCs and human fibroblast of scleroderma (hsFb) in vitro,and to probe the therapy possibility of mdMSCs on scleroderma.METHODS: The cultivated mdMSCs were isolated by tissue digested-adherented-subcultured in low-serum medium.The changes of hydroxyproline (Hyp) and TGF-β₁ in co-cultured mdMSCs and human normal fibroblast (hnFb) were determined at day 4 and 8 by samples basic hydrolysis and ELISA respectively.RESULTS: The Hyp production and TGF-β₁ expression of hsFb were significant higher than that of hnFb on day 8 (P<0.05),no difference among the various hsFb/mdMSCs co-cultured Transwell system was found (P>0.05).The TGF-β₁ production in hsFb/mdMSCs 2.5×104 co-culture Transwell system was significant higher than that in hsFb culture alone on day 4 (P<0.05),but there was on difference between them on day 8 (P>0.05).No correlation between the production of Hyp and TGF-β₁ in co-cultured Transwell system (r＝0.221,P>0.05) was observed.However,the production of Hyp and TGF-β₁ showed significant positive correlation under the condition that hnFb or hsFb was cultured alone (P<0.05).CONCLUSION: In vitro,mdMSCs couldn't effectively reduce the production of Hyp and TGF-β₁ by hsFb in Transwell system.The mdMSCs may not effectively treat scleroderma by the effect on hsFb.     

Hypothalamic expression of orexin A and lipid metabolism disorder in rats with alimentary obesity

AIM: To evaluate the effect of orexin A in rat hypothalamus on lipid metabolism disorder in rats with alimentary obesity induced by high-fat diet.METHODS: The rat model of alimentary obesity was induced by high-fat diet. The levels of insulin, triglyceride (TG) and total cholesterol (TC) in the serum were detected by luminescent immunoassay and enzymic method. The mRNA expression of orexin A in rat hypothalamus was determined by real-time PCR.RESULTS: There were statistically significant differences of weight, body fat content, and Lee's index between high-fat diet group and control group after 8-week feeding of high-fat diet. Compared to control animals, the levels of insulin, TG and TC in the rats with alimentary obesity significantly increased by 50％, 94％ and 43％, respectively (P<0.05). The expression of orexin A in rat hypothalamus significantly decreased by 57％, and had significant negative correlation with Lee's index, insulin, TG and TC. Their correlation coefficients were r=-0.798 (P<0.05), r=-0.868 (P<0.05), r=-0.981(P<0.05) and r=-0.815 (P<0.05), respectively. CONCLUSION: Alimentary obesity and lipid metabolism disorder induced by high-fat diet are correlated with down-regulation of orexin A expression in rat hypothalamus.     

Effects of PTEN on apoptosis,oxidative damage and immune inflammatory factors in myocardial H9c2 cells with anoxia/reoxygenation

AIM: To investigate the effects of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) on the apoptosis, oxidative damage and immune inflammatory factors in myocardial H9c2 cells with anoxia/reoxygenation (A/R). METHODS: The H9c2 cells were used to establish a model of A/R. The H9c2 cells were transfected with PTEN small interfering RNA (siRNA) and negative control. After A/R, the expression of PTEN at mRNA and protein levels was determined by RT-PCR and Western blot, respectively. The cell viability was measured by MTT assay. The apoptosis was analyzed by flow cytometry. Xanthine oxidase method was used to determine superoxide dismutase (SOD) activity. The content of malondialdehyde (MDA) was detected by thiobarbituric acid method. The lactate dehydrogenase (LDH) activity in the supernatant was evaluated by 4-dinitrophenylhydrazine method. The levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6 in culture supernatant were examined by ELISA. The protein levels of cleaved caspase-3, Bax and FasL in the cells were determined by Western blot. RESULTS: After A/R, the expression of PTEN at mRNA and protein levels was significantly increased in the H9c2 cells (P<0.05). The mRNA and protein levels of PTEN were decreased significantly after transfection with PTEN siRNA (P<0.05). The viability of H9c2 cells was decreased after A/R, while the apoptotic rate was increased. The protein levels of cleaved caspase-3, Bax and FasL were increased in the cells. The MDA level was elevated, the activity of SOD was decreased, and the levels of LDH, TNF-α, IL-1β and IL-6 in the culture supernatant were increased (P<0.05). Down-regulation of PTEN partly antagonized the effects of A/R on the viability, apoptotic rate, MDA content, SOD activity, and the levels of LDH, TNF-α, IL-1β and IL-6 in culture supernatant. CONCLUSION: Down-regulation of PTEN attenuates oxidative damage induced by A/R, reduces apoptosis and secretion levels of TNF-α, IL-1β and IL-6 in the H9c2 cells.     

PTEN,VEGF, COX-2 and angiogenesis in JAK2 V617F positive myeloroliferative neoplasms

AIM: To investigate the expression of PTEN, VEGF and COX-2, and the relationship with angiogenesis in the bone tissues of JAK2 V617F mutation positive myeloroliferative neoplasm(MPN)patients. METHODS: JAK2 V617F positive MPN patients (n=42) were selected in the First Hospital of Baoding including 25 cases of newly diagnosed group and 17 cases of treatment group with IFN-α 2b, and 10 cases of idiopathic thrombocytopenic purpura (ITP) patients served as controls. The ratio of mutant and wild type JAK2 was detected by real-time PCR. The protein levels of p-JAK2, PTEN, VEGF and COX-2, and microvascular density (MVD) marked with CD105 in pathological tissues of bone marrow were detected by immunohistochemistry. RESULTS: The levels of p-JAK2, VEGF, COX-2 and MVD in newly diagnosed group were significantly higher than those in control group. However, the expression level of PTEN was significantly lower than that in control group. The levels of p-JAK2, VEGF, COX-2 and MVD in treatment group were significantly lower than those in newly diagnosed group, while the expression level of PTEN was significantly higher than that in newly diagnosed group. JAK2 V617F mutation burden was positively correlated with VEGF, COX-2 and MVD (P<0.05). PTEN was negatively correlated with VEGF and MVD (P<0.05). The levels of p-JAK2, VEGF, COX-2 and MVD in JAK2 V617F/JAK2 ratio≥0.5 group were significantly higher than those in JAK2 V617F/JAK2 ratio<0.5 group, but PTEN showed the contrary to the above.CONCLUSION: PTEN, VEGF, COX-2 and JAK2 V617F are involved in the pathogenesis of angiogenesis in MPN patients.     

Adenosine promotes bFGF protein and bFGF mRNA expression in human umbilical vein endothelial cells in vitro

AIM: To investigate the influence of adenosine on human umbilical vein endothelial cells (HUVEC) bFGF protein production and bFGF mRNA expression. METHODS: Immunohistochemistry staining was performed to detect bFGF protein. RT-PCR was performed to detect bFGF mRNA expression. RESULTS: Immunohistochemistry study demonstrated that there was only a small amount of bFGF positive cells and the color was weak in control group (without adenosine). In groups treated with 10-4 mol/L and 10-6 mol/L adenosine, bFGF protein was significantly higher than that in control group (P<0.05). In 10-8 mol/L and 10-10 mol/L adenosine groups, there were no significant differences compared with control group (P>0.05). RT-PCR showed that in 10-4 mol/L and 10-6 mol/L adenosine groups, bFGF mRNA expression was higher than that in control group (P<0.05), while the difference between 10-8 mol/L adenosine group and control group was not significant (P>0.05). CONCLUSION: Adenosine may promote HUVEC proliferation and angiogenesis partly through inducing bFGF expression.     

Effect of tumor suppressor phosphatase and tensin homology deleted in chromosome 10 on renal interstitial fibrosis in IgA nephropathy

AIM: To explore the potential correlation between the expression of phosphatase and tensin homology deleted on chromosome 10(PTEN) and RIF in IgA nephropathy. METHODS: Forty-seven patients diagnosed as primary IgA nephropathy by renal histology were involved. 10 specimens from normal renal tissue of renal carcinoma served as control group. Tubulointerstitial lesion (TIL) was classified by using Katafuchi scale, including no TIL (group I), mild TIL (group II), moderate TIL (group III), severe TIL (group IV). The expressions of PTEN, TGF-β1, α-SMA and ColⅢ in renal tissue were detected by immunohistochemistry. PTEN mRNA was detected by in situ hybridization. RESULTS: Renal tissues from renal biopsy showed abundant expressions of PTEN and PTEN mRNA in endochylema of renal tubular epithelial cells, and negligible expression in glomeruli. With the progress of TIF in IgA nephropathy, the expressions of PTEN and PTEN mRNA decreased gradually (P<0.05). In contrast, the expressions of TGF-β1, α-SMA and ColⅢ increased significantly (P<0.05). The expressions of PTEN and PTEN mRNA in renal tissues were positively with eGFR and urine osmotic pressure (P<0.01), negatively correlated with the expressions of TGF-β1, α-SMA, ColⅢ, urinary protein excretion for 24 h, the rate of sclerosis glomeruli and the scores of vascular lesion (P<0.01). CONCLUSION: These results suggest the loss of PTEN expression in RIF of IgA nephropathy. Moreover, TGF-β signaling induces epithelial to mesenchymal transition and extracellular matrix accumulation possibly through a mechanism dependent on the downregulation of PTEN.     

miR-499 attenuates myocardial injury in rats by targeting PTEN

AIM:To investigate the effect of microRNA-499 (miR-499) on myocardial injury in rats and its mechanism. METHODS:The rat model of ischemia-reperfusion (I/R) injury was established and the myocardial cells were primarily cultured. The expression level of miR-499 was detected by RT-qPCR. After hydrogen peroxide stimulation, CCK-8 assay and LDH kit were used to detect the viability and LDH release of the cells transfected with miR-499 mimic and siR-PTEN. The targeting relationship between miR-499 and PTEN was predicted by TargetScan and confirmed by luciferase test. RT-qPCR and Western blot were used to detect the effect of miR-499 mimic and inhibitor on the mRNA and protein expression levels of PTEN. After miR-499 inhibitor and siR-PTEN were co-transfected into the cells, CCK-8 assay and LDH kit were used to detect the viability and LDH release of the myocardial cells induced by hydrogen peroxide. RESULTS:The expression level of miR-499 in the I/R rats was increased rapidly, and then was decreased gradually in a time-dependent manner (P<0.05). miR-499 mimic and siR-PTEN significantly promoted the viability and decreased the LDH release of cardiomyocytes induced by hydrogen peroxide (P<0.05). miR-499 and PTEN had a targeting relationship. The expression of PTEN was significantly down-regulated by miR-499 mimic, and up-regulated by miR-499 inhibitor (P<0.05). Transfection with siR-PTEN reversed the inhibitory effect of miR-499 inhibitor on the cells. CONCLUSION:miR-499 attenuates the myocardial injury in rats by targeting PTEN.