Effect of senescence marker protein 30 on the proliferation and apoptosis of human lens epithelial cells SRA01/04

AIM： To study the effect of senescence marker protein 30（SMP30） on the proliferation and apoptosis of human lens epithelial cell（HLEC） SRA01/04.METHODS： SMP30 overexpression（OE） and knock down（KD） type cell lines were cultivated by using two groups regucalcin（RGN; SMP30） lentiviral vectors（LVRGN, LV-RGN-RNAi） and the respective negative control virus infect SRA01/04 cells. Western blot and real-time quantitative polymerase chain reaction（q-PCR） analysis were used to determine RGN overexpression and knock down efficiency. We use cell counting kit-8（CCK8） assay to measure cell viability and 5-bromodeoxyuridine（Brd U） assay to test cell proliferation. Cell cycle was measured by PI FACS assay and cell apoptosis was tested by Annexin V-APC assay through flow cytometry. We use Western blot to measure the content of caspase-3 in SRA01/04.RESULTS： We used PCR and Western blot techniques to determine the successful transfection of SMP30 OE and KD SRA01/04 cell lines. By CCK8, Brdu and PI FACS cell cycle assay, it was found that the SMP30 OE group promoted cell proliferation（P〈0.05） compared with the control group, and the KD group inhibited cell proliferation（P〈0.05）. The results of Annexin V-APC signal staining detection indicated that compared with respective control group, the cell apoptosis rate was higher in KD group（P〈0.05） but lower in OE group（P〈0.01）. The expression of caspase-3 was down-regulated in OE group through Western blot assay and up-regulated in KD group compared with respective control group. CONCLUSION： Proliferation of SRA01/04 was promoted by SMP30 OE and apoptosis was suppressed. Increasing the expression of SMP30 may protect HLEC SRA01/04 against apoptosis in cataract.

Construction of a plasmid for human brain-derived neurotrophic factor and its effect on retinal pigment epithelial cell viability

Several studies have investigated the protective functions of brain-derived neurotrophic factor（BDNF） in retinitis pigmentosa. However, a BDNF-based therapy for retinitis pigmentosa is not yet available. To develop an efficient treatment for fundus disease, an eukaryotic expression plasmid was generated and used to transfect human 293 T cells to assess the expression and bioactivity of BDNF on acute retinal pigment epithelial-19（ARPE-19） cells, a human retinal epithelial cell line. After 96 hours of co-culture in a Transwell chamber, ARPE-19 cells exposed to BDNF secreted by 293 T cells were more viable than ARPE-19 cells not exposed to secreted BDNF. Western blot assay showed that Bax levels were downregulated and that Bcl-2 levels were upregulated in human ARPE-19 cells exposed to BDNF. Furthermore, 293 T cells transfected with the BDNF gene steadily secreted the protein. The powerful anti-apoptotic function of this BDNF may be useful for the treatment of retinitis pigmentosa and other retinal degenerative diseases.

Heat shock protein 70 expression in relation to apoptosis in primary bladder transitional cell carcinoma

Bladder carcinoma is the most common tumor in the urinary system. In 1996, a sampleinvestigation showed that bladder carcinoma, in which more than 90% was mainly primary bladder transitional cell carcinoma （BTCC）, was one of the ten highest mortality malignant tumors in China. Bladder carcinoma represented 2% of all malignant tumors and has the fifth most common malignancy in men in Europe and North America.

Effect of Early Intervention with Extract of Huannao Yicong Decoction(还脑益聪方) on the Pathologic Picture of Hippocampus and Neurocyte Apoptosis in APP Transgenic Mice Model of Dementia

Objective：To observe the effect of early intervention using extract of Huannao Yicong Decoction （还脑益聪方,HYD） on the pathological picture of hippocampus,neurocyte apoptosis,and associated regulatory genes inβ-amyloid precursor protein（APP） transgenic mice model of dementia.Methods：Sixty APP695^（V7171） transgenic mice were randomly divided into four groups of 15.The model group was treated with distilled water, the positive control group was treated with donepezil（0.65 mg/kg）,and the two HYD groups were treated with high dose（2.8 g/kg） and low dose（1.4 g/kg） HYD,respectively.All testing drugs were administered through gastrogavage by dissolving in equal volume of distilled water,once a day for six successive months.In addition, a normal control group with 15 healthy C57BL/6J mice of the same age and genetic background was set up with distilled water treatment.The pathologic picture of brain tissue was observed by microscopy with HE stain;the amount of apoptosis cells in the hippocampal CA1 area was detected by TUNEL;and expressions of associated genes,Bcl-2,and Bax were determined by immunohistochemical method.Results：Pathologic pictures of hippocampus showed that in the model group,cells messily arranged,neurons markedly decreased, and the surrounding tissue of some cells was loosened with edema,necrosis,and widened gap with glia cells proliferation.Compared with those in the normal group,the amount of apoptosis cells in the CA1 area was increased,Bcl-2 expression decreased,and Bax expression increased significantly,with markedly reduced Bcl-2/Bax ratio in the model group.Compared to the model group,the pathological changes were significantly milder in the HYD-treated groups,showing rather regularly arranged cells,significantly increased neurons, only few denatured necrotic cells with milder edema,less proliferation of glia cells,and obviously reduced cell apoptosis in CA1 area（P0.05 or P0.01）.Besides,Bcl-2 expression was up-regulated and Bax expression down-regulated,and Bcl-2/Bax ratio significantly increased in the two HYD groups（P0.05 or P0.01）. Conclusion：Early intervention with HYD could improve the abnormal pathologic picture of hippocampus and regulate the expressions of associated genes to suppress cell apoptosis,which might be its mechanism of action in alleviating cognitive functional disorder.