Influence of breast milk on secretion of Ghrelin and Peptide YY in rats / 中华实用儿科临床杂志

Objective To study the effect of breast milk on secretion of Ghrelin and Peptide YY (PYY)during different periods in SD rats.Methods Sprague-Dawley (SD) rats were randomly divided into breast milk-fed group(BG) and formula-artificial fed group(FG) with 20 in each group.After 21 days both of the 2 groups were fed by same forage.Ten of each group were experimentized in day 21 equivalently weaning period,and rest rats were experimentized at the 50th day equivalently childhood.Real-time PCR was used to determine the mRNA of PYY,Ghrelin from gastric,colon.Immunohistochemistry was used to determine Ghrelin,PYY protein expression in the gastrointestinal tract tissues.Results There were no difference between body weight and gastrointestinal mucosal development of 2 rat groups in day 21 and day 50(P ＞0.05),mRNA and protein expression of Ghrelin and PYY in breast milk-fed group were higher than formula-artificial fed group in both day 21 and day 50 (all P ＜ 0.05).Conclusions The Ghrelin and PYY levels of breast milk-fed rats is higher than formula-artificial fed ones,and this phenomenon continues to their childhood.Breast milk protects offspring from obesity by influencing the secretion of brain-gut peptide and has long-term consequences on the regulation of food intake and energy balance from neonatal period to their later life.

Enhance efficiency of adenovirus on AAV transfering human RPE cells / 中华实验眼科杂志

Background Adeno-associated virus-based vector is one of most efficient vehicles.It presents with a long-term and efficient transfer and expression of therapeutic genes with minimal toxicity.But its delayed-and low-efficient transgene expression limits the application of AAV vector.To explore an improving method of AAV infecting RPE cells is the hot spot. Objeetive Present study was to investigate whether adeno-associated virus (AAV)combined with low dose non-replieable adenovirus(Ad-null)can enhance its infection efficieney on RPE ceils in vitro. Methods Human RPE cells were isolated from the donate eyeballs under the approval of the Ethic Committee of this hospital.The cells were cultured in DMEM containing 10%fetal bovine serum.AAV particles with enhanced green fluorescence protein(EGFP)were added into the medium alone or in combination with different amount of adenovirus for 30 days.The cells were detected under the fluorescence microscope,and the protein expression levels of report gene EGFP in RPE cells were analyzed with Western blotting assay. Results Melanin granules could be found in cultured RPE cells.EGFP was expressed in RPE cells at 2 days after AAV-EGFP infection and peaked at 12 days and remained for about 3-week duration,showing the green influorescence under the influorescence mwroscope.After the cells were infected by AAV2-EGFP with 0.01 to 1000 MOI Ad-null respectively,the number of cells with green influorescence was obviously increased with the enhanced infiuorescence intensity.The enhance of the infection efficiency began in the 0.1 MOI Ad-null group and peaked in 10 MOI Ad-null group.Dead cells were exhibited in the 100 or more MOI Ad-nulor group.Western blotting assay demonstrated that the protein expression level of EGFP in RPE cells enhanced significantly in 1 and 10 MOI Ad-null groups compared with only AAV infection group. Conclusion These finding suggested that the infection efficiency of AAV can be improved significantly when it is used with low dose Ad-null in vitro.This offers a basis for further study of gene therapy.

Low dosage 5-fluorouracil increases the transfection efficiency of Ad/VEGF-A in mouse lung carcinoma cell line LA795 and inhibits tumor growth / 癌症

<p><b>BACKGROUND AND OBJECTIVE</b>Adenovirus vectors were widely used in gene therapy for tumors. We used adenovirus vector to transfer small interfering RNA (siRNA) against vascular epithelium growth factor A (VEGF-A) molecules to mouse lung adenoma LA795 cells and used low dose of chemotherapeutic drugs to further elevate the infection efficiency of adenovirus vector in and therapeutic effect of RNAi on tumor cells.</p><p><b>METHODS</b>LA795 cells were infected by Ad/EGFP and treated with different dosages of gemcitabin, epirubicin, cisplatin, or 5-fluorouracil (5-FU). Cells were observed under fluorescence microscope continuously using green fluorescent protein (GFP) as the reporter gene. The percentage of GFP-positive cells and fluorescent intensity were tested by flow cytometry to determine optimum concentrations of drugs. Ad/siVEGF-A containing VEGF-A siRNA was constructed. Real-time PCR and ELISA were applied to measure the expression level of VEGF-A after LA795 cells were infected by Ad/siVEGF-A and treated with 5-FU. The combination of Ad/siVEGF-A and 5-FU was also applied in treating subcutaneous tumor in mice.</p><p><b>RESULTS</b>Low dose of 5-FU elevated the Ad/EGFP infection in LA795 cells significantly, and also enhanced the effect of Ad/siVEGF-A in down-regulating VEGF-A mRNA and protein levels in tumor cells. When used in tumor in vivo, the combination strategy repressed tumor growth effectively.</p><p><b>CONCLUSION</b>Low dose of 5-FU can enhance the capability of adenovirus infecting tumor cells and promote the efficiency of gene therapy by adenovirus.</p>

Construction and application of conditionally replicative adenovirus for selective cytotoxicity in CEA positive colorectal cancer cells / 上海第二医科大学学报

Objective To construct a new conditionally replicative adenovirus (CRAds) targeting carcinoembryonic antigen (CEA) positive colorectal cancer cells. Methods The DNA fragment of the CEA gene promoter was amplified through PCR and cloned into the vector carrying fusion reporter gene EGFP-Luc to construct expression plasmid pCEA-EGFPLuc. The constructed plasmid pCEA-EGFPLuc was transfected into CEA positive and negative cells by liposome. The activity of CEA gene promoter was evaluated by detecting the expression of EGFP and luciferase activity. The conditionally replicative adenovirus Ad.CEA-E1A/CMV-TK carrying suicide gene HSVtk was constructed, in which the E1A gene was controlled under CEA promoter. CEA positive(Lovo and SW620)and negative tumor cells(HeLa) were infected with Ad.CEA-E1A/ CMV-TK. The selective cytotoxicity of Ad.CEA-E1A/CMV-TK and the synergistic effect of the virus with GCV in CEA positive tumor cells were evaluated by the expression of E1A, cytopathic effect and cell survival rate. Results CEA promoter possesses a good specificity as well as high activity. The expression of E1A only presented in CEA positive tumor cells. After infection with Ad. CEA-E1A/CMV-TK, the cell survival rates of Lovo and SW620 were (36.72±2.49)% and (39.82±4.76)%, respectively, significantly lower than that of Hela[(87.44±2.76)%1 (P<0.01). When combined with GCV, Ad.CEA-E1A/CMV-TK had better oncolytic effect on Lovo and SW620 cells, with cell survival rates of (17.26±3.65) % and (23.93±5.40) %, respectively, significantly lower than those without GCV[(36.72±2.49) % and (39.82±4.76) %, respectively] (P<0.01). Conclusion Ad. CEA-E1A/CMV-TK under the control of CEA promoter has selective cytotoxic effect on CEA positive colorectal cancer cells, and the effect can be enhanced when combined with GCV.

Changes of Peptide with Tyrosine and Ghrelin Levels in Preterm Infants and Their Relationships with Body Weight / 实用儿科临床杂志

Objective To study the role of peptide with tyrosine(PYY) and ghrelin in infants by comparing the difference and correlation of PYY,ghrelin levels and body weight in preterm infants and full-term infants.Methods Radio-immunity was used to determine serum PYY and ghrelin levels in 20 preterm infants in their first,3rd and 7th days,and also in 20 full-term infants at the 7th day.Body weight were recorded in the both groups.Then the levels of 2 hormone were analyzed the correlations with the body weight.Results Serum PYY and ghrelin levels increased remarkably in preterm infants as compared with those in full-term infants[full-term infants:PYY was(601.9?206.2) ng?L-1;ghrelin was(1 064.5?208.6) ng?L-1;preterm infants:PYY was(812.4?153.8) ng?L-1;ghrelin was(1 485.4?409.2) ng?L-1,Pa