Effect of Comprehensive Nursing on Pain Relief, Comfort and Burden of Family Care of Infantile Anal Fistula.

Objective: To explore the effect of comprehensive nursing on pain relief, comfort and burden of family care of infantile anal fistula. Methods: This was a randomized, double-blind, controlled clinical trial. A total of 106 cases of children with anal fistula from January 2021 to December 2021 were selected and divided into observation group and control group, there were 53 cases in each group. The control group were underwent with routine nursing intervention. The observation group were underwent with video health education, peer support, music relaxation training, physiotherapy and auricular point pressing on the basis of routine nursing therapy and other measures (comprehensive nursing). The wound healing time, comfort score, complication rate and family care burden of two groups were compared. Results: After intervention, the scores of pain degree in the observation group were significantly lower than those in the control group (4.02 ± 0.85 vs 5.89 ± 2.36, p < 0.05), and the scores of comfort degree in the observation group were higher than those in the control group (70.23 ± 5.98 vs 46.88 ± 5.23, p < 0.05). After intervention, the wound healing time of the observation group was shorter than that of the control group (3.98 ± 0.63 vs 5.77 ± 1.02, p < 0.05), and the crying times of the observation group were less than that of the control group (1.22 ± 0.26 vs 4.02 ± 0.48, p < 0.05). After intervention, the total scores of family members' care load in the observation group were significantly lower than those in the control group (37.26 ± 4.78 vs 55.99 ± 5.02, p < 0.05). Conclusion: Comprehensive nursing can effectively promote wound healing in infantile anal fistula, reduce pain, prompt children's comfort, reduce the number of children crying, and reduce the burden of care for children's families.

Construction of high-quality Caco-2 three-frame cDNA library and its application to yeast two-hybrid for the human astrovirus protein-protein interaction.

Human epithelial colorectal adenocarcinoma (Caco-2) cells are widely used as an in vitro model of the human small intestinal mucosa. Caco-2 cells are host cells of the human astrovirus (HAstV) and other enteroviruses. High quality cDNA libraries are pertinent resources and critical tools for protein-protein interaction research, but are currently unavailable for Caco-2 cells. To construct a three-open reading frame, full length-expression cDNA library from the Caco-2 cell line for application to HAstV protein-protein interaction screening, total RNA was extracted from Caco-2 cells. The switching mechanism at the 5' end of the RNA transcript technique was used for cDNA synthesis. Double-stranded cDNA was digested by Sfi I and ligated to reconstruct a pGADT7-Sfi I three-frame vector. The ligation mixture was transformed into Escherichia coli HST08 premium electro cells by electroporation to construct the primary cDNA library. The library capacity was 1.0×10(6)clones. Gel electrophoresis results indicated that the fragments ranged from 0.5kb to 4.2kb. Randomly picked clones show that the recombination rate was 100%. The three-frame primary cDNA library plasmid mixture (5×10(5)cfu) was also transformed into E. coli HST08 premium electro cells, and all clones were harvested to amplify the cDNA library. To detect the sufficiency of the cDNA library, HAstV capsid protein as bait was screened and tested against the Caco-2 cDNA library by a yeast two-hybrid (Y2H) system. A total of 20 proteins were found to interact with the capsid protein. These results showed that a high-quality three-frame cDNA library from Caco-2 cells was successfully constructed. This library was efficient for the application to the Y2H system, and could be used for future research.

Screening effective short interfering RNA/short hairpin RNA for inhibition of human astrovirus ORF2 gene expression in cultured cells.

In this study, we have evaluated four different 21-nt duplexes of small interfering RNA (siRNA-469, siRNA-852, siRNA-1802 and siRNA-1806) that specifically target the ORF2 gene of human astrovirus (HAstV) in inhibiting HAstV capsid protein expression in transfected BHK-21 cells. Furthermore, fluorescence analysis, real-time quantitative PCR (RT-qPCR) and western blot assays showed that pGPU6/GFP/Neo-shRNA inhibits ORF2 gene expression in Caco2 cells. The results indicate that siRNA/shRNA-469 and siRNA/shRNA-1802 can interfere with capsid protein expression in cell culture, and this provides a powerful tool for the study of HAstV gene functions and the biological properties of the capsid protein.

[Construction and expression of six deletion mutants of human astrovirus C-terminal nsP1a/4 protein].

Human astrovirus (HAstV) is one of the leading causes of actue virual diarrhea in infants. HAstV-induced epithdlial cell apoptosis plays an important role in the pathogenesis of HAstV infection. Our previous study indicated that HAstV non-structural protein nsPla C-terminal protein nsPla/4 was the major apoptosis functional protein and probably contained the main apoptosis domains. In order to screen for astrovirus encoded apoptotic protien, nsPla/4 and six turncated proteins, which possessed nsPla/4 protein different function domain ,were cloned into green fluorescent protein (GFP) vector pEG-FP-N3. After 24-72 h transfection, the fusion protein expression in BHK21 cells, was analysis by fluorescence microscope and Western blot. The results indicated seven fusion proteins were observed successfully in BHK21 cell after transfected for 24 h. Western blot analysis showed that the level of fusion protein expressed in BHK21 cells was increased significantly at 72h compared to 48h in transfected cells. The successful expression of deletion mutants of nsPla/4 protein was an important foundation to gain further insights into the function of apoptosis domains of nsPla/4 protein and it would also provide research platform to further confirm the molecule pathogenic mechanism of human astrovirus.