N-acetylcysteine alleviates oxidative stress and apoptosis and prevents skeletal muscle atrophy in type 1 diabetes mellitus through the NRF2/HO-1 pathway.

AIMS: Type 1 diabetes mellitus (T1DM) has been linked to the occurrence of skeletal muscle atrophy. Insulin monotherapy may lead to excessive blood glucose fluctuations. N-acetylcysteine (NAC), a clinically employed antioxidant, possesses cytoprotective, anti-inflammatory, and antioxidant properties. The objective of our study was to evaluate the viability of NAC as a supplementary treatment for T1DM, specifically regarding its therapeutic and preventative impacts on skeletal muscle. MAIN METHODS: Here, we used beagles as T1DM model for 120d to explore the mechanism of NRF2/HO-1-mediated skeletal muscle oxidative stress and apoptosis and the therapeutic effects of NAC. Oxidative stress and apoptosis related factors were analyzed by immunohistochemistry, immunofluorescence, western blotting, and RT-qPCR assay. KEY FINDINGS: The findings indicated that the co-administration of NAC and insulin led to a reduction in creatine kinase levels, preventing weight loss and skeletal muscle atrophy. Improvement in the reduction of muscle fiber cross-sectional area. The expression of Atrogin-1, MuRF-1 and MyoD1 was downregulated, while Myh2 and MyoG were upregulated. In addition, CAT and GSH-Px levels were increased, MDA levels were decreased, and redox was maintained at a steady state. The decreased of key factors in the NRF2/HO-1 pathway, including NRF2, HO-1, NQO1, and SOD1, while KEAP1 increased. In addition, the apoptosis key factors Caspase-3, Bax, and Bak1 were found to be downregulated, while Bcl-2, Bcl-2/Bax, and CytC were upregulated. SIGNIFICANCE: Our findings demonstrated that NAC and insulin mitigate oxidative stress and apoptosis in T1DM skeletal muscle and prevent skeletal muscle atrophy by activating the NRF2/HO-1 pathway.

ChromInst: A single cell sequencing technique to accomplish pre-implantation comprehensive chromosomal screening overnight.

Next Generation Sequencing (NGS) is a powerful tool getting into the field of clinical examination. Its preliminary application in pre-implantation comprehensive chromosomal screening (PCCS) of assisted reproduction (test-tube baby) has shown encouraging outcomes that improves the success rate of in vitro fertilization. However, the conventional NGS library construction is time consuming. In addition with the whole genome amplification (WGA) procedure in prior, makes the single cell NGS assay hardly be accomplished within an adequately short turnover time in supporting fresh embryo implantation. In this work, we established a concise single cell sequencing protocol, ChromInst, in which the single cell WGA and NGS library construction were integrated into a two-step PCR procedure of ~ 2.5hours reaction time. We then validated the feasibility of ChromInst for overnight PCCS assay by examining 14 voluntary donated embryo biopsy samples in a single sequencing run of Miseq with merely 13M reads production. The good compatibility of ChromInst with the restriction of Illumina sequencing technique along with the good library yield uniformity resulted superior data usage efficiency and reads distribution evenness that ensures precisely distinguish of 6 normal embryos from 8 abnormal one with variable chromosomal aneuploidy. The superior succinctness and effectiveness of this protocol permits its utilization in other time limited single cell NGS applications.

Slow-pull and different conventional suction techniques in endoscopic ultrasound-guided fine-needle aspiration of pancreatic solid lesions using 22-gauge needles.

AIM: To evaluate the cytological diagnostic capacity and sample quality of the slow-pull technique and compare them with different suction techniques. METHODS: From July 2010 to December 2015, 102 patients with pancreatic solid lesions who underwent endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) with 22-gauge needles were retrospectively evaluated. EUS-FNA diagnosis was based on a cytological examination, and final diagnosis was based on a comprehensive standard of cytological diagnosis, surgical pathology and clinical or imaging follow-up. Cytological specimens were characterized for cellularity and blood contamination. The cytological diagnostic capacity and sample quality of the slow-pull technique and suction techniques with 5-mL/10-mL/20-mL syringes were analyzed. RESULTS: Of all of the EUS-FNA procedures, the slow-pull technique and suction techniques with 5-mL/10-mL/20-mL syringes were used in 31, 19, 34 and 18 procedures, respectively. There were significant differences between these four suction techniques in terms of cytological diagnostic accuracy (90.3% vs 63.2% vs 58.8% vs 55.6%, P = 0.019), sensitivity (88.2% vs 41.7% vs 40.0% vs 36.4%, P = 0.009) and blood contamination (score ≥ 2 for 29.0% vs 52.6% vs 70.6% vs 72.2%, P = 0.003). The accuracy and sensitivity of the slow-pull technique were significantly higher than those of the suction techniques using 5-mL (P = 0.03, P = 0.014), 10-mL (P = 0.005; P = 0.006) and 20-mL syringes (P = 0.01, P = 0.01). Blood contamination was significantly lower in the slow-pull technique than in the suction techniques with 10-mL (P = 0.001) and 20-mL syringes (P = 0.007). CONCLUSION: The slow-pull technique may increase the cytological diagnostic accuracy and sensitivity with slight blood contamination during EUS-FNA when using 22-gauge needles for solid pancreatic masses.