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Objective·To explore the possible pathogenic gene of total anomalous pulmonary venous connection (TAPVC) by whole exon sequencing and verify its function.Methods·One hundred TAPVC children (case group) and one hundred and twenty healthy children (control group) in Xinhua Hospital and Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine from 2014 to 2019 were included. The blood samples from the two groups of children were collected, and whole blood genomic DNA was extracted for exon sequencing to screen out the potential pathogenic genes of TAPVC. Harmful mutation sites of pathogenic genes were screened through Mutation Taster, SIFT and PolyPhen-2 websites, and then conducted by Sanger sequencing. The wild-type (wild-type group) and mutant (mutant group) plasmids of PDX1 were transfected into HUVEC cells. Quantitative real-time PCR (qPCR) and Western blotting were used to detect the effects of mutations on mRNA and protein levels of PDX1, respectively. The STRING database was used to analyze the interaction between proteins, and qPCR was used to determine the expressions of downstream genes regulated by PDX1.Results·Pathogenic PDX1 was found in TAPVC children, and Sanger sequencing revealed two novel variants in the gene: c.C237A (P33T) and c. C237G (P33A). Compared with the wild-type group, there was no significant difference in PDX1 mRNA levels in the two mutant groups, but there was a significant increase in relative protein expression of the CA group and CG group, which was 2.9 and 3.4 times higher than the wild-type group, respectively (P=0.000, P=0.001). Protein interaction analysis demonstrated that PDX1 was associated with SOX17. qPCR results showed that overexpression of PDX1 could downregulate the expression of SOX17 in HUVEC.Conclusion·The two novel PDX1 missense mutations can affect the process of PDX1 post-transcriptional translation, indicating that PDX1 may participate in the occurrence and development of TAPVC by regulating SOX17.

This article summarizes the clinical features and follow-up findings of insulin autoimmune syndrome in two patients with type 2 diabetes after the use of lipoic acid,along with literature review,with an attempt to increase the awareness of lipoic acid-induced insulin autoimmune syndrome among clinicians and improve its diagnosis and treatment.

Objective: We screened and identified a strain capable of enantioelectively hydrolyzing methyl (R,S)-N-(2,6-dimethylphenyl) alaninate (MAP), a key intermediate for the synthesis of metalaxyl, followed by cloning and expressing the esterase in E. coli., Methods: We used MAP as the sole carbon source in the medium inoculated with an active sludge specimen to enrich the target microorganism. The strain with the highest hydrolysis activity and enatioelectivity was identified by 16S rRNA sequence analysis, morphological observation and physiological and biochemical properties. From the gene library of the strain, the DNA sequence fragment containing the target gene was found. By DNA sequence analysis and PCR amplification, the esterase gene was obtained. It was ligated with plasmids pET28a (+), then transformed into E. coli BL21Gold (DE3) plysS., Results: We isolated a gram-negative bacterial strain capable of enantioelective hydrolyzing MAP. It was identified as Achromobacter denitrificans. From its gene library, the esterase gene named EHest was found. The recombinant EHest-pET28a(+)-BL21Gold (DE3) plysS was constructed. The recombinant expressed esterase (EHesterase) capable of catalyzing enatioelective hydrolysis of methyl (R,S)-N-(2,6-dimethylphenyl) alaninate. Its size was 27 kDa. The expression activity was 27.1 times as high as that in the original strain. Hydrolysis of MAP (5% M/V) by EHesterase for 1 h at 37℃, the substrate conversion was 29.5% and ee p of the product acid (major in R configuration) was 85.1%. The optimum pH was 9.0 and temperature 50℃., Conclusions: A new isolate Achromobacter denitrificans 1104 capable of enantioelective hydrolyzing MAP was found and identified.

The bovine jugular veins were divided into two groups and treated with 4% EX-313 and 0.5% glutaraldehyde respectively, and then they were examined with naked-eye, microscope and scanning electron microscope. Biomechnics test and dorsal implantation in rats were performed. The aquired data were processed and subjected to t-test. The EX-313 fixed material was more pliable than the glutaraldehyde treated material, and the former had higher anticalcification than the latter. In conclusion, the hydrophilia crosslinking agent EX-313 is superior to glutaraldehyde in treating biomaterials, and the bovine jugular vein tanned with EX-313 should be a promising material for repairing in cardiovascular surgery.