OBJECTIVETo construct the plasmid expression vector pSIH1-H1-copGFP for RNA interference against vascular endothelial growth factor C (VEGF-C) and to evaluate its effect on the expression of VEGF-C mRNA in gastric cancer cells after transfection.

METHODSThree siRNAs of genome sequence of VEGF-C gene were retrieved from GenBank and one negative chain was used as control. Four siRNAs were cloned into plasmid pSIH1-H1-copGFP,which were then transfected into gastric cancer cells (SGC7901). The expression of VEGF-C mRNA was analyzed by RT-PCR.

RESULTSThe recombinant plasmid of pSIH1-H1-copGFP specific for VEGF-C was confirmed by gene sequencing analysis. The target sequence obtained was completely consistent with the design. Transfection efficiency of the three siRNAs ranged from 60% to 70%. After transfection, the expression of VEGF-C mRNA in SGC7901 cells was significantly inhibited. Inhibition rates of VEGF-C mRNA expression were 35.4%, 33.8% and 81.5% in the three siRNA plasmid vectors, respectively.

CONCLUSIONThe siRNA expression plasmid vector against VEGF-C mRNA is successfully constructed, and RNAi may be a useful technique to inhibit the lymphangiogenesis of gastric cancer.

Cell Line, Tumor ; Genetic Vectors ; Humans ; Plasmids ; RNA Interference ; RNA, Small Interfering ; Stomach Neoplasms ; genetics ; Transfection ; Vascular Endothelial Growth Factor C ; genetics

Objective:To investigate the difference of vitamin A and E levels in children with different respiratory diseases at different ages.Methods:A total of 671 children in Hunan Children's Hospital from July 2017 to October 2019 were selected as the disease group, including 197 cases of pneumonia, 152 cases of recurrent respiratory tract infection, 91 cases of asthma, 88 cases of cough variant asthma and 143 cases of Mycoplasma pneumoniae pneumonia; At the same time, 245 healthy children were selected as the normal group. The serum vitamin A and vitamin E levels of the two groups were detected by high performance liquid chromatography (HPLC).Results:⑴ The vitamin A level [(0.31±0.09)mg/L] of the disease group was lower than the normal group [(0.35±0.25)mg/L], and the vitamin E level [(8.92±2.57)mg/L] was lower than the normal group [(9.62±2.79)mg/L], with statistically significant difference ( P<0.05); ⑵ The level of vitamin A in the disease group at the age of >1-3 years [(0.32±0.09)mg/L] was lower than that in the normal group of the same age group [(0.35±0.08)mg/L]; the level of vitamin A in the disease group at the age of >3-6 years old [(0.30±0.08)mg/L] was lower than that of the same age group [(0.32±0.07)mg/L], with statistically significant difference ( P<0.05); ⑶ The vitamin E level of the disease group at >1-3 years old [(9.23±2.56)mg/L], >3-6 [(8.02±1.86)mg/L] and >6-14 years old [(8.02±1.82)mg/L] were lower than that of the same age normal group [(9.76±2.81)mg/L, (9.67±2.87)mg/L, (9.19±2.58)mg/L], with statistically significant difference ( P<0.05); ⑷ There were significant differences in vitamin A levels among different age in disease group ( P<0.05). Among them, the children with high risk of subclinical deficiency accounted for the largest proportion (45.78%) in the 6-month-1-year-old group, and the proportion of children with normal vitamin A levels in other age groups was the largest; ⑸ There are significant differences in vitamin E levels in different age groups in the disease group ( P<0.05), the levels in the normal range accounts for the largest proportion of all ages; ⑹ The levels of vitamin A and vitamin E in mycoplasma pneumoniae infection group were increased compared with in recurrent respiratory infection group , asthma group, and cough variant asthma group, and the difference was statistically significant ( P<0.05). Compared with the pneumonia group, the level of vitamin E increased in the recurrent respiratory infection group, and the difference was statistically significant ( P<0.05); The vitamin E levels in the cough variant asthma group were reduced compared with the repeated respiratory infection group, asthma group and pneumonia group ( P<0.05). Conclusions:The Vitamin A and E levels of children suffering from respiratory diseases are lower than those of normal children. The Vitamin A and E levels of different respiratory diseases and different age groups are different. Vitamin A and E supplementation may be significantly targeted according to different ages and different respiratory diseases in clinical practice.

AIM: To investigate the effects of antitumor drug paclitaxel(PTX)on the proliferation, apoptosis, cell cycle, cell morphology, and related protein expression of Müller cells, and to evaluate its potential toxicity to the retina.METHODS:Müller cells were cultured in vitro and divided into two groups: control group(normal medium)and PTX group. Retinal Müller cells were treated with different concentrations of PTX(0.005, 0.05, 0.5 and 5mg/L)for varying durations(12, 24, 36, 48 and 72h). The CCK8 method was used to assess the effects of different concentrations of PTX and treatment duration on the proliferation Müller cells. Flow cytometry was employed to investigate the impact of different concentrations of PTX on Müller cells apoptosis and cell cycle arrest. Immunofluorescence was used to observe morphological changes in Müller cells. The effects of PTX on the expression of apoptosis-related proteins and aquaporins were analyzed by Western blot and qRT-PCR.RESULTS: PTX exhibits the ability to inhibit the proliferation of Müller cells when cultured in vitro. The efficacy of this inhibition was found to be dependent on both the concentration of the drug and the duration of the stimulation. Higher concentrations of the drug and longer stimulation times resulted in a weaker ability of the cells to proliferate. Additionally, PTX also induces apoptosis in Müller cells, with increased drug concentrations and longer stimulation times leading to higher apoptosis rates. Flow cytometry analysis demonstrates that PTX arrests Müller cells in the G2-M phase of the cell cycle. Moreover, there is a distinct change in cell morphology, with a shift from the typical appearance characterized by clear and slender fibrous structures to a rounder morphology, accompanied by a significant decrease in cell numbers. Further, our findings reveal that there is a transient increase in the expression of cytoinflammatory factors following drug treatment compared to the control group. However, discontinuation of drug stimulation can alleviate this heightened expression. In treated cells, the expression of the CA XIV protein is upregulated compared to the control group, while the expression of vascular endothelial growth factor(VEGF)is downregulated(P<0.05). Additionally, the levels of inflammatory factors in the PTX group are significantly higher than those in the control group(P<0.05), suggesting that PTX has the potential to disrupt the retinal barrier function.CONCLUSION: PTX affects the proliferation and apoptosis of Müller cells, with the effects dependent on stimulation duration and drug concentration. In addition, PTX blocks the Müller cell cycle at the G2-M phase and alters cell morphology, leading to a transient upregulation of inflammatory factors and affecting the integrity of the retinal barrier. These findings indicate the potential toxicity of the antitumor drug PTX to the retina.

Aim To research the effect of PDG on bone metabolism in young rats. Methods The experimental rats were randomly divided into contro group, PDG-25 group and PDG-50 group. PDG-25 group and PDG-50 group were given PDG at the dose of 25 mg·kg