[Analysis on drug resistance-associated mutations of multi-drug resistant Mycobacterium tuberculosis based on whole-genome sequencing in China].

Objective: To analyze the resistance mutational profiles of multi-drug resistant Mycobacterium tuberculosis in China and the correlation between major mutation types and genotypes based on the whole-genome sequencing data. Methods: Search and download of the genome-wide sequencing data of M. tuberculosis published in China by August 2019 on NCBI database were conducted. Mutation frequency of drug resistance-related gene loci based on whole-genome sequencing was used to predict the molecular susceptibility of strains, and the correlation between mutation types and genotypes was analyzed. Results: According to the results of molecular resistance and susceptibility profiles, 1 024 MDR strains were identified from 2 019 M. tuberculosis strains. The major mutation types of resistance-related genes to common drugs were katG S315T (73.2%, isoniazid), rpoB S450L (63.1%, rifampicin), rpsL K43R (70.0%, streptomycin), embB M306V (37.4%, ethambutol), pncA_promoter T (-11)C (7.9%, pyrazinamide), gyrA A90V (32.3%, fluoroquinolones), rrs A1401G (67.7%, second-line injection drugs), fabG1_promoter C (-15) T (87.0%, Ethionamide), folC I43T (30.4%, P-aminosalicylic acid). Among them, the frequencies of katG S315T, embB M306V, rpsL K43R, gyrA A90V in lineage 2 were significantly higher than those in lineage 4, and folC I43T was only found in lineage 2. The proportion of katG S315T was significantly higher in the ancient Beijing genotype compared to the modern genotype, in contrast, the proportion of rpsL K43R was significantly higher in modern Beijing genotype, the differences were significant (all P<0.05). Conclusions: The results showed the main mutation types of resistance-related genes of MDR strains to many commonly used anti-tuberculosis drugs in China based on whole-genome sequencing, providing a basis for the development of sensitive and specific rapid molecular detection methods. At the same time, it was also found that the major mutation types of MDR-related genes were related to the genotype of the strains.

[Piperine inhibits the transformation of endothelial cells into fibroblasts].

Objective: To investigate the role of piperine on the transformation of endothelial cells into fibroblasts. Methods: Cultured human umbilical vein endothelial cells (HUVECs, 4-6 passage) were used for the main experiments. The transformation models of endothelial cells into fibroblasts were induced by transforming growth factor ß (TGF-ß) stimulation. HUVECs were divided into 6 groups: control group, TGF-ß group and 4 groups treated with various concentrations of piperine (1, 5, 10, 20 µmol/L). CKK-8 was used to detect cell proliferation. The CD31/α-smooth muscle actin (α-SMA) expression level was detected by fluorescent staining. The vascular endothelial cadherin (VE-cadherin)/vimentin expression was detected by immunofluorescence staining. RT-PCR was used detect the mRNA expressions of transformation markers. Western blot was used to detect the protein expression of snail and twist. Results: TGF-ß increased HUVECs proliferation (P<0.05), which could be significantly inhibited by 10 and 20 µmol/L of piperine, but not by 1 and 5 µmol/L of piperine. Immunofluorescence results demonstrated that TGF-ß increased HUVECs transformation to fibroblasts as shown by downregulated expression of endothelial markers CD31, VE-cadherin, and upregulated expression of α-SMA and vimentin, again, these effects could be attenuated by 10 and 20 µmol/L piperine. The expression levels of collagen type Ⅰ and type Ⅲ were significantly higher in TGF-ß group than in control group (P<0.05), significantly lower in TGF-ß+10 µmol/L piperine group and TGF-ß+20 µmol/L piperine group than in TGF-ß group (P<0.05).In addition, RT-PCR results showed that TGF-ß increased mRNA expression of transformation markers (snail1, snail2, twist1, twist2), while 10 and 20 µmol/L of piperine could significantly downregulated the mRNA expressions of these markers. The protein expression levels of snail and twist were significantly higher in TGF-ß group than in control group (both P<0.05), which was significantly lower in TGF-ß+20 µmol/L piperine group than in TGF-ß group (both P<0.05). Conclusions: Piperine can inhibit the transformation of endothelial cells into fibroblasts. This effect might be viewed as one of the potential mechanisms of reduced myocardial fibrosis post piperine treatment.

Effects of normal lymphatic fluid on rats with sepsis complicated with lung injury.

OBJECTIVE: Infection can be caused by severe burnt, trauma and hypoxia, further causing systemic inflammatory syndrome or sepsis. The sepsis occurs in about 2% of all hospitalizations and ranges from 6% to 30% in intensive care unit (ICU) in developed countries This study aimed to investigate the effects of normal lymph fluid on sepsis complicated with pulmonary injury. MATERIALS AND METHODS: Wistar rats were prepared for sepsis complicated with acute pulmonary model via cecal ligation and puncture, and received normal lymph fluid injection 60 min later. Artery blood-gas index, wet/dry weight (W/D) ratio of lung, Myeloperoxidase (MPO) and superoxide dismutase (SOD) activity of lung tissues were measured, along with protein content and cell count in bronchoalveolar lavage fluid (BALF). Real-time PCR (RT-PCR) and Western blot were employed to measure expression of nuclear factor κB (NF-κB) in lung tissues, whilst enzyme linked immunosorbent assay (ELISA) was used to analyze serum expression of tumor necrosis factor α (TNF-α) and interleukin 2 (IL-2). RESULTS: Model group had significantly depressed PaO2 and pH value, higher W/D ratio, and MPO activity, lower SOD activity, higher protein and cell count of BALF, and up-regulation of TNF-α, IL-2 and NF-κB (p < 0.05 compared to sham group). Infusion of lymph fluid effectively improved blood-gas function, decreased W/D ratio, MPO activity, elevated SOD activity, and lowered TNF-α, IL-2 and NF-κB expression (p < 0.05 compared to model group). CONCLUSIONS: Normal lymph fluid can inhibit NF-κB expression, suppress inflammation, and improve blood-gas exchange in lung tissues. Therefore, the normal lymph fluid could effectively relieve the sepsis complicated with pulmonary injury.

The role of configurational disorder on plastic and dynamic deformation in Cu64Zr36 metallic glasses: A molecular dynamics analysis.

The varying degrees of configurational disorder in metallic glasses are investigated quantitatively by molecular dynamics studies. A parameter, the quasi-nearest atom, is used to characterize the configurational disorder in metallic glasses. Our observations suggest configurational disorder play a role in structural heterogeneity, plasticity and dynamic relaxations in metallic glasses. The broad configurational disorder regions distribution is the indicator of abundant potential deformation units and relaxations. Plastic flow, as well as relaxation, is believed to start at configurational disorder regions. The width of the shear bands and dynamic relaxations can then be regulated by the degree of configurational disorder regions in metallic glasses.

Structural evolution of nanoscale metallic glasses during high-pressure torsion: A molecular dynamics analysis.

Structural evolution in nanoscale Cu50Zr50 metallic glasses during high-pressure torsion is investigated using molecular dynamics simulations. Results show that the strong cooperation of shear transformations can be realized by high-pressure torsion in nanoscale Cu50Zr50 metallic glasses at room temperature. It is further shown that high-pressure torsion could prompt atoms to possess lower five-fold symmetries and higher potential energies, making them more likely to participate in shear transformations. Meanwhile, a higher torsion period leads to a greater degree of forced cooperative flow. And the pronounced forced cooperative flow at room temperature under high-pressure torsion permits the study of the shear transformation, its activation and characteristics, and its relationship to the deformations behaviors. This research not only provides an important platform for probing the atomic-level understanding of the fundamental mechanisms of high-pressure torsion in metallic glasses, but also leads to higher stresses and homogeneous flow near lower temperatures which is impossible previously.

MiRNA-21 reverses high glucose and high insulin induced insulin resistance in 3T3-L1 adipocytes through targeting phosphatase and tensin homologue.

AIMS/HYPOTHESIS: Our previous study showed there was a change of microRNA (miRNA) expression profile, and miR-21 was significantly down regulated in insulin-resistant adipocytes (IR-adipocytes). Phosphatase and tensin homologs deleted on chromosome 10 (PTEN), a negative regulator of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, was identified to be a target gene of miR-21, which suggested miR-21 might be associated with insulin resistance (IR) or diabetes. However, it is not known whether miR-21 play any role in the development of IR in 3T3-L1 adipocytes. METHODS: Normal adipocytes and adipocytes transfected with pre-miR-21(pmiR-21) or negative control (pNeg) were treated with high glucose and high insulin for 24 h, insulin-stimulated glucose uptake was determined by 2-Deoxyglucose transport assay, miR-21 expression level was measured by using quantitative real-time RT-PCR (qRT-PCR). The protein expression levels of PTEN, Akt, phospho-Akt (Ser473), IRß, GSK3ß, phospho-GSK3ß (Ser9) and GLUT4 were detected by western blotting assay. RESULTS: We further confirmed that miR-21 was down regulated in IR-adipocytes by qRT-PCR. Over-expression of miR-21 significantly increased insulin-induced glucose uptake and decreased PTEN protein expression, while it had no significant effect on PTEN mRNA expression in IR-adipocytes. Moreover, over-expressing miR-21 significantly increased insulin-induced phosphorylation of AKT (Ser473), GSK3ß (Ser9) and the translocation of glucose transporter 4 (GLUT4) in IR-adipocytes. CONCLUSIONS: In this study, our data demonstrate that miR-21 reverses high glucose and high insulin induced IR in 3T3-L1 adipocytes, possibly through modulating the PTEN-AKT pathway, and miR-21 may be a new therapeutic target for metabolic diseases such as T2DM and obesity.