SARS-CoV-2 NSP12 utilizes various host splicing factors for replication and splicing regulation.

The RNA-dependent RNA polymerase (RdRp) is a crucial element in the replication and transcription of RNA viruses. Although the RdRps of lethal human coronaviruses severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), SARS-CoV, and Middle East respiratory syndrome coronavirus (MERS-CoV) have been extensively studied, the molecular mechanism of the catalytic subunit NSP12, which is involved in pathogenesis, remains unclear. In this study, the biochemical and cell biological results demonstrate the interactions between SARS-CoV-2 NSP12 and seven host proteins, including three splicing factors (SLU7, PPIL3, and AKAP8). The entry efficacy of SARS-CoV-2 considerably decreased when SLU7 or PPIL3 was knocked out, indicating that abnormal splicing of the host genome was responsible for this occurrence. Furthermore, the polymerase activity and stability of SARS-CoV-2 RdRp were affected by the three splicing factors to varying degrees. In addition, NSP12 and its homologues from SARS-CoV and MERS-CoV suppressed the alternative splicing of cellular genes, which were influenced by the three splicing factors. Overall, our research illustrates that SARS-CoV-2 NSP12 can engage with various splicing factors, thereby impacting virus entry, replication, and gene splicing. This not only improves our understanding of how viruses cause diseases but also lays the foundation for the development of antiviral therapies.

Structure and Anti-HIV Activity of Betulinic Acid Analogues.

Firstly discovered in 1980s, human immunodeficiency virus (HIV) continues to affect more and more people. However, there is no effective drug available for the therapy of HIV infection. Betulinic acid existing in various medicinal herbs and fruits exhibits multiple biological effects, especially its outstanding anti-HIV activity, which has drawn the attentions of many pharmacists. Among the derivatives of betulinic acid, some compounds exhibited inhibitory activities at the nanomolar concentration, and have entered phase II clinical trials. This paper summarizes the current investigations on the anti-HIV activity of betulinic acid analogues, and provides valuable data for subsequent researches.

Diastereoselective Synthesis of CF3-Containing Vicinal Diamines.

The highly diastereoselective synthesis of CF3-containing vicinal diamines by a convenient two-step procedure without the need to isolate the intermediate products is described.

Synthesis and antioxidant activity of curcumin analogs.

Numerous biological activities including antioxidant, antitumor, anti-inflammation, and antivirus of the natural product curcumin were reported. However, the clinical application of it was significantly limited by its instability, poor solubility, less body absorbing, and low bioavailability. This review focuses on the structure modification and antioxidant activity evaluation of curcumin. To study the structure-activity relationship (SAR), five series of curcumin analogs were synthesized and their antioxidant activity were evaluated in vitro. The results showed that electron-donating groups, especially the phenolic hydroxyl group are an essential component to improve the antioxidant activity.

Relationship between serum irisin levels and urinary albumin excretion in patients with type 2 diabetes.

AIM: Irisin is first discovered as a potential mediator of obesity related energy homeostasis. Recent studies indicate that irisin is associated with endothelial dysfunction and atherosclerosis in patients with type 2 diabetes. Our objective was to examine the relationship between irisin and urinary albumin excretion in patients with type 2 diabetes. METHODS: 100 newly diagnosed patients with type 2 diabetes and 100 healthy subjects were selected. Serum irisin levels were measured by ELISA, and urine albumin was measured by radioimmunoassay. High resolution ultrasound was used to measure brachial artery diameter at rest, after reactive hyperemia (flow-mediated arterial dilation, FMD) and after sublingual glyceryltrinitrate. RESULTS: Patients with type 2 diabetes presented decreased irisin levels when compared to controls (14.12±3.93 versus 28.98±2.56ng/ml, P=0.015).Serum irisin levels in the microalbuminuric and macroalbuminuria subgroup were 9.89±1.56ng/ml and 5.67±1.89ng/ml, respectively, which were significantly lower than those in the normoalbuminuria (15.97±3.12ng/ml). In comparison to microalbuminuric subgroup, macroalbuminuria subgroup had lower levels of irisin. By dividing the distribution of serum irisin levels into quartiles, FMD was increased gradually with the increase of serum irisin levels (P<0.001). Multiple stepwise linear regression analysis showed that FMD (ß=0.75, P=0.002), 2-hBG (ß=-0.25, P=0.038) and UAE (ß=-0.87, P=0.008) were significantly associated with irisin. Pearson's correlation analyses showed a negative correlation between irisin and logUAE (r=-0.57) and between FMD and logUAE (r=-0.47), and positive correlations between irisin and FMD (r=0.51). CONCLUSIONS: Decreased plasma levels of irisin seem to be associated with UAE and FMD in patients with type 2 diabetes.

[The relationship between association of microalbuminuria and retinal vessel diameter in population with essential hypertension].

OBJECTIVE: To investigate the association of urinary albumin-to-creatinine ratio (UACR) and the diameter of retinal vessel in population with essential hypertension in Fujian coastal area. METHODS: Central retinal artery and vein equivalents (CRAE and CRVE) were measured from the avoiding mydriatic digitized photographs and semi-automatic fundus analysis software, as well as albumin and urine creatinine. RESULTS: There were significant differences in CRAE levels among the normal control group, normoalbuminuria with essential hypertension group and microalbuminuria with essential hypertension group [(135.68 ± 10.10) µm, (129.79 ± 10.48) µm, (125.29 ± 11.17) µm, all P values < 0.01]. The CRAE levels were significantly negative correlated with UACR (r = -0.29, P < 0.01). Linear regression analysis showed CRAE was associated with UACR in the patients with hypertension(ß = -5.0, P < 0.01). Logistic regression analysis showed, systolic blood pressure (ß = 1.08, P = 0.02) was risk factor for CRAE abnormality. The CRAE abnormality was increased in turn in the normal control group, normoalbuminuria with the essential hypertension group and microalbuminuria with essential hypertension group (P < 0.01). CONCLUSION: The reduction of central retinal artery diameter are associated with the hypertensive renal damage. UACR and CRAE could be used to evaluate the microvascular lesions and be used as an indicator to assess the target organs damage in essential hypertension patients.

Insulin receptor substrate 1 regulates the cellular differentiation and the matrix metallopeptidase expression of preosteoblastic cells.

Insulin receptor substrate 1 (IRS1) is an essential molecule for the intracellular signaling of IGF1 and insulin, which are potent anabolic regulators of bone metabolism. Osteoblastic IRS1 is essential for maintaining bone turnover; however, the mechanism underlying this regulation remains unclear. To clarify the role of IRS1 in bone metabolism, we employed RNA interference to inhibit IRS1 gene expression and observed the effects of silencing this gene on the proliferation and differentiation of and the expression of matrix metallopeptidase (MMP) and tumor necrosis factor receptor superfamily, member 11b (TNFRSF11B) in MC3T3-E1 cells. Our results showed that IRS1 short hairpin RNAs can effectively suppress the expression of IRS1, and inhibit the phosphorylation of AKT in IRS1 pathway; reduce the expression of MMP2, MMP3, MMP13, and MMP14, decrease the expression of TNFRSF11B and RANKL (also known as tumor necrosis factor (ligand) superfamily, member 11), however increase the RANKL/TNFRSF11B ratio; decrease cell survival, proliferation, and mineralization, and impair the differentiation of MC3T3-E1 cells. The downregulation of IRS1 had no effect on the expression of MMP1. Our findings suggest that IRS1 not only promotes bone formation and mineralization but also might play roles in bone resorption partly via the regulation of MMPs and RANKL/TNFRSF11B ratio, thus regulates the bone turnover.

Effects of Glimepiride on metabolic parameters and cardiovascular risk factors in patients with newly diagnosed type 2 diabetes mellitus.

BACKGROUND: To investigate the effects of Glimepiride on blood glucose in patients with newly diagnosed type 2 diabetes mellitus (T2DM) in connection with plasma lipoproteins and plasminogen activity. METHODS: A total of 565 T2DM patients were received Glimepiride (n=333) or Glibenclamide (n=232) for 12 weeks. We observed the level of blood glucose (BG), glycated hemoglobin (HbA1C), the insulin resistance (IR) state, plasma lipoproteins, tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type I (PAI-1) before and after a 12 weeks of treatment. RESULTS: After 12 weeks with Glimepiride treatment, significant reductions were observed in fasting blood glucose (FBG) and 2-h postprandial BG(PBG), HbA1C (from 8.60+/-3.10 to 7.10+/-1.60%) and HOMA-IR (from 4.11+/-0.85 to 2.42+/-0.91%). The level of total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) were significantly decreased, whereas that of high-density lipoprotein (HDL) was increased markedly with statistical significance. In addition, there was an obvious improvement in t-PA activity (from 0.225+/-0.11 to 0.457+/-0.177IU/ml); whereas the PAI-1 activity was decreased significantly (from 0.898+/-0.168 to 0.533+/-0.215AU/ml). No significant changes were observed in plasma lipoprotein profiles and plasminogen activity in Glibenclamide receiving group. CONCLUSIONS: Glimepiride can rapidly and stably improve glycemic control and lipoprotein metabolism, significantly alleviate insulin resistance and enhance fibrinolytic activity.

[Proliferation, differentiation, and gene expression profile of osteoblast of patient with spondyloepiphyseal dysplasia tarda with progressive arthropathy].

OBJECTIVE: To explore the proliferation, differentiation, and gene expression profile of the osteoblasts (OBs) of patient with spondyloepiphyseal dysplasia tarda with progressive arthropathy (SEDT-PA). METHODS: OBs from the head of femur of a SEDT-PA patient resected during operation were cultured. ELISA was used to examine the bone-specific alkaline phosphatase (BAP), osteocalcin (OC), and osteoprotegrin (OPC) in the lysate of OBs. The proliferation and survival of the OBs were evaluated with MTT method and (3)H-TDR incorporation. Flow cytometry was used to observe the cell cycle. Northern blotting was used to evaluate the mRNA expression of WISP3, a member of the CCN family, mRNA in the OBs. Gene differential expression microarray was used to determine the cDNA expression. Osteoblasts from the head of femur of a patient about the same age with fracture of femur neck was used as control. RESULTS: The cultured OBs from the SEDT-PA patient showed a higher survival capacity (0.86 +/- 0.04 vs 0.71 +/- 0.10) and more (3)H-TDR incorporation (1363 +/- 350 vs 867 +/- 128). The expressions of OC and OPG were down-regulated to the 1/4.3 and 1/4.69 of the control respectively. There was no significant difference in the expression of BAP. WISP3 was very lowly expressed in the OBs of the SEDT-PA patient. In comparison with the normal control, there were up-regulation of 22 genes, including those coding chemotactic factors and inflammatory factors, and down-regulation of 16 genes in the OBs of the SEDT-PA patient with a synthetic effect of more rapid proliferation of OBs and reduction of synthesis of extracellular matrix, resulting in osteopenia. CONCLUSION: Not significantly different between SEDT-PA patient and normal person, the WISP3 expression may not be a direct factor of SEDT-PA.

[Effects of estradiol and progesterone on the expression of insulin receptor substrate in human osteoblasts].

OBJECTIVE: To investigate the effects of 17beta-estradiol (E(2)) and progesterone (P) on the expression of IRS in human osteoblast cells (HOBs) and further study the mechanism of E(2) and P on postmenopausal osteoporosis (PMOP). METHODS: Osteoblasts from cancellous bones of normal adults were cultured and underwent intervention of E(2) and P respectively. Semi-quantitative RT-PCR was used to measure the expression of mRNAs of IRS-1 and IRS-2. The expression of the proteins of IRS-1 and IRS-2 were detected by Western blotting. The expressions of phosphorylated IRS protein (P-IRS) was examined by immunocoprecipitation/Western blotting. RESULTS: The expression level of IRS-2 mRNA in the HOBs undergoing intervention of E(2) and P were 421% +/- 68% and 327% +/- 54% that of the control group respectively (P < 0.05 and P < 0.01). The expression level of IRS-2 mRNA in the HOBs undergoing intervention of E(2) combined with P was 496% +/- 54% that of the control group (P < 0.01). However, the expression level of IRS-1 was not significantly influenced by E(2) and/or P (all P > 0.05). The expression level of IRS-2 protein and phophorylated IRS-2 protein in the HOBs undergoing intervention of E(2), P, and E(2) combined with P were 487% +/- 65%, 507% +/- 54%, 552% +/- 47% and 483% +/- 52%, 527% +/- 76%, and 717% +/- 86% that of the control group (P < 0.05, P < 0.01, and P < 0.05). The P-IRS-1 protein level of the HOBs undergoing intervention of E(2), P, and E(2) combined with P were 54% +/- 7.6%, 37% +/- 4.2%, and 21.2% +/- 3.0% that of the control group (P < 0.05, P < 0.01, and P < 0.01). CONCLUSION: E(2) and P up-regulate the expression of IRS-2 mRNA in HOBs. Combined intervention of E(2) and P positively creates a synergistic effect on the up-regulation of IRS-2 mRNA. E(2) and P have no effects on the expression of IRS-1 mRNA. E(2) and P increase the expression of IRS-2 protein and P-IRS-2. Combined intervention of E(2) and P presents a positive synergistic effect on the up-regulation of P-IRS-2. However, they show no effect on the expression of IRS-1 protein, and down regulate the phosphorylation of IRS-1 protein.

[Characterization of biologic behaviors and cDNA expression of articular chondrocytes in spondyloepiphyseal dysplasia tarda with progressive arthropathy].

OBJECTIVE: Spondyloepiphyseal dysplasia tarda with progressive arthropathy (SEDT-PA) is an autosomal-recessive hereditary disorder of cartilage homeostasis. The pathogenesis of SEDT-PA is unknown though it has been demonstrated that CCN6 is the SEDT-PA causing gene. The present study characterized the biologic behaviors and cDNA differential expression profile of articular chondrocytes in SEDT-PA. METHODS: The morphologic and functional features of growth, proliferation, differentiation and DNA synthesis of SEDT-PA chondrocytes were determined with cell growth curve, (3)H-TDR incorporation, MTT and flow cytometry. cDNA differential expression profile was carried out with gene microarray containing 8000 genes. Differentially expressed genes were verified by RT-PCR, Western blot and immunohistochemistry. RESULTS: As compared with normal control, the variant chondrocytes were much larger in size with an enhanced ability of proliferation and DNA synthesis and increased ratio of G(2)-M (10.72% vs 0.11%) to S phases (37.0% vs 15.8%). Matrix metalloproteinases (MMPs), which decompose the extra-cellular matrix of the cartilage, accumulated at the endochylema and failed in exocytosis, which lead to the negative feedback of the mRNA transcriptions. The mRNA expression of MMPs was down-regulated. At the same time, the mRNA expression of genes related to cell growth, proliferation and progression of cell cycles were up-regulated, while most of those associated with extracellular matrix, non-collagen proteins and poly-proteoglycans were down-regulated. CONCLUSIONS: The accumulation and failure in exocytosis of the MMPs decompose the extra-cellular matrix of the cartilage. The decreased expression of matrix proteins and polyproteoglycans is involved in the pathogenesis of arthropathy resulted from CCN6 mutation.