Search

This study explored the effect of commercial ripened Pu-erh tea (Y562, China Tea (Yunnan) Industry Co. Ltd.) on improving the health of populations with abnormal glucose and lipid metabolism. Qualified volunteers were recruited for nutritional epidemiological and clinical intervention studies. The volunteers were asked to drink the tea for three months. Their body composition (body mass, body mass index (BMI), body fat ratio and visceral fat), body circumferences (waist circumference, hip circumference and neck circumference), glucose metabolism-related indicators (fasting blood glucose, insulin, C-peptide, glycosylated hemoglobin), lipid metabolism-related indicators (total cholesterol, triglycerides, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, apolipoprotein A1 and B), blood uric acid levels, liver fat contents, and gut microbiota were tested and compared before and after drinking the tea to explore its health benefits. The results showed that after drinking Y562 without any change in diet or exercise for three months, the body mass, BMI, body circumferences, blood lipids (high-density lipoprotein cholesterol, apolipoprotein A1 and B), and liver fat contents of the volunteers were significantly improved (P 0.05). Moreover, the gut microbiota was improved, showing increased uniformity. The relative abundance of Lactobacillus, Clostridium, Akkermansia muciniphila, Clostridium screenens and Streptococcus thermophilus was increased, while that of Streptococcus, Lactococcus, Bacteroides, Bifidobacterium and Paraacteroides distasonis was decreased. This study suggests that ripened Pu-erh tea Y562 has the potential to help reduce the body mass, improve the body shape, reduce blood glucose and lipids, and alleviate non-alcoholic fatty liver disease (NAFLD). The underlying mechanism may be related to the regulation of the intestinal microecology and the bile acid pathways by Pu-erh tea.

Objective To establish a stable-expressing human cell cycle regulatory protein 50A (CDC50A) SKOV3 cell strain expressing vector containing CDC50A by molecular cloning. Methods CDC50A was introduced into the expression vector pLVX-IRES-GFP by molecular cloning method. The pLVX-CDC50A-GFP expression plasmid was packaged by lentivirus and stably transfected into SKOV3. SKOV3 cell strain with high-expressing CDC50A was established. The expression of Flag protein was analyzed by flow cytometry, Western blot and immunofluorescence. Results The construction of pLVX-CDC50A-GFP expressing vector was confirmed by gel electrophoresis and endonuclease digestion. The efficiency of infection was observed qualitatively under fluorescence microscope and detected quantitatively by flow cytometry as 86.5%. Flag-tag protein was detected by Western blot to present only in the positive group. Conclusions The SKOV3 cell strain was established by infecting with lentivirus of pLVX-CDC50A-GFP, laying a foundation for the mechanism study of CDC50A in ovarian cancer.

Objective To construct the expression vectors carrying human Type Ⅳ-P-type phospholipids transferases 11A or 11B (ATP11A or ATP11B), and to detect the up-regulation effect on ATP11A or ATP11B. Methods The primers were designed according to ATP11A or ATP11B sequence and cloned into EZ-TTM Cloning Vector. The primers were identified by enzyme digestion and verified by sequencing. ATP11A and ATP11B sequences were digested and directed to expression vectors pLVX-IRES-mCherry and pLVX-IRES-RFP, respectively. Then the recombinant expression plasmids with immunofluorescence protein were transfected into HEK-293T using liposomes. Finally, the expression of the recombinant cell strains was verified by sequencing, fluorescence microscopy, flow cytometry and Western blot. Results The construction of ATP11A and ATP11B cloning vectors was confirmed by gel electrophoresis and was analyzed by endonuclease. The efficiency of transfection was observed qualitatively under microscope, detected quantitatively by flow cytometry as 38.9%,41.9% respectively, and the tagged proteins were detected by Western blot to present only in the positive groups. Conclusions The expression vectors of human ATP11A and ATP11B are successfully constructed and their expression levels could be effec-tively upregulated,which lay a foundation for future mechanism studies involved to ATP11A or ATP11B.

Objective To validate the stemness bio-function of side population (SP) cells from epithelial ovarian cancer cell lines and to detect the differentially expressed cellular membrane proteins in SP cells. Methods SP cells of human ovarian cancer cell lines SKOV3 and A2780 which secreted bisbenzimide Hoechst33342 and located as side population were sorted by flow cytometry. Bio-functions of stemness of SP cells were validated. Through stable isotope labeling by amino acids in cell culture (SILAC), differentially expressed membrane proteins in SP cells were detected. Results SP cells of SKOV3 and A2780 were sorted successfully. The ratio of SP cells was about 0.5% and increased significantly through repeated sorting. SP cells differentiated into non-SP (NSP) cells. Stemness of SP cells was validated in vitro, including cell clone forming, resistance to cisplatin and paclitaxel in NOD/SCID mice. SILAC technique was used to detect differentially expressed membrane proteins in SP cells. Results showed that 1 561 and 1 989 differentially expressed proteins were detected respectively in SKOV3 and A2780 SP cells. Among those significantly up-regulated proteins in SKOV3 SP cells, 12 proteins were also up-regulated in A2780 SP cells. On the contrary, 13 proteins were down-regulated both in SKOV3 and A2780 SP cells. Conclusions SP cells of human epithelial ovarian cancer cells show stemness. And differently expressed membrane proteins in SP cells were detected which would be the base of further screening surface biomarker of ovarian cancer stem-like cells.