Fluorescent copper nanoclusters as a nano-dye for DNA methyltransferase activity analysis and inhibitor screening.

Fluorescent copper nanoslusters (CuNCs) as a new class of fluorophores have attracted more and more attention due to their ease of synthesis, excellent optical properties, and low cost. In this study, a novel label-free fluorescent method was developed for the detection of DNA methyltransferases based on template length-dependent of dsDNA-CuNCs. In the absence of DNA adenine methylation methyltransferase (Dam MTase), the dsDNA containing the methylation-responsive sequence could effectively template the formation of fluorescent CuNCs with bright fluorescence. When the dsDNA substrate is methylated by Dam MTase, the methylation-sensitive restriction endonuclease Dpn I cleaves the methylated dsDNA and produces shorter dsDNA product, which fails to template fluorescent CuNCs. So, the Dam MTase activity could be identified by the changes of CuNCs' fluorescence. Based on this method, a linear range of 0.5-10 U/mL was achieved with high sensitivity and selectivity. Moreover, we also demonstrate the proposed method can be applied to evaluation and screening of inhibitors for Dam MTase.

Gene expression profiles of rat MMECs with different glucose levels and fgl2 gene silencing.

BACKGROUND: Cardiac microvascular endothelial cells (MMECs) is one of the key factors in the process of diabetic cardiomyopathy, a common chronic complication of diabetes. Fibrinogen-like protein 2 (FGL2) is linked to apoptosis, angiogenesis, and inflammatory response, all of which also occur in diabetes. Thus, we investigate the role of FGL 2 and other genes in the pathology of diabetic cardiomyopathy. METHODS: In the present study, we used high-throughput microarray to profile gene expression in rat myocardial MMECs with or without silencing the fgl2 gene and in different glucose environments. We use volcanic maps to isolate genes with significantly different expression levels between conditions, using the standard statistical criteria of fold changes ≥1.5 and P-values ≤0.05. From this list, we identified genes with the most signicant changes in RNA levels and confirmed their protein-level changes with Western blot. Furthermore, bioinformatic analysis predicts possible pathophysiology and clinical relevance of these proteins in diabetic cardiomyopathy. RESULTS: We identified 17 upregulated and 15 downregulated genes caused by silencing fgl2 gene. Most of them are involved in metabolism, ion transport, cell membrane surface recognition signal modification, inflammatory response, and immune response. Using Western blot, we were able to confirm protein-level expression changes of three genes. Specifically, in both normal and high glucose conditions, silencing fgl2 significantly decreased the expression levels of CCL3 and PLAGL1 while increasing the expression level of CTSC. Significantly, bioinformatic analyses show that CCL3 is related to type 1 diabetes, PLAGL1 to cardiomyocytes, and CTSC to albuminuria in type 2 diabetes. CONCLUSIONS: Our study provides clues for further studies on the mechanism of diabetic cardiomyopathy as well as function of FGL2 in this process, potentially offering new therapeutic strategies for treating diabetic cardiomyopathy.

A novel endovenous laser ablation strategy of treatment of greater saphenous vein varicosities with difficult wire placement.

AIM: To investigate the efficacy and early postoperative morbidity of a novel endovenous laser ablation (IEVLA) strategy of treatment of the great saphenous vein (GSV) with difficulty of wire placement. METHODS: Sixty patients with serious GSV incompetence in 73 limbs were randomized into two treatment groups: Group 1 underwent traditional endovenous laser ablation (TEVLA) surgery and group 2 received IEVLA. Local pain, ecchymosis, induration, paraesthesia in treated regions, thrombotic diseases, vein diameter, treated vein length, delivered energy, operation duration, success rate in placement of the laser fiber and venous clinical severity (VCS) scores were recorded for both group. Follow-up were conducted on the 2nd day, 7th day, and 1st, 2nd, 3rd and 6th month postoperatively. RESULTS: In group 1, induration was present in 18 cases, ecchymosis in 19, paraesthesia in 9, pulmonary embolism (PE) in 1 case, and deep vein thrombus (DVT) in 3. While in group 2, induration present in 29, ecchymosis in 23, paraesthesia in 17 with and no patients were complicated with PE or DVT. Although no difference in improvement of VCS score existed between the two groups at each follow-up time point, group 2 had significantly shorter operation time and higher success rate (P < 0.05). CONCLUSION: IEVLA is a more effective and safe technique for treatment of serious GSV varicosities with difficulty of wire placement.