High performance liquid chromatography with ultraviolet detection for the determination of SYUIQ-5, a novel telomerase inhibitor for cancer therapy: application to an enzyme kinetic study in rat liver microsomes

A sensitive assay for the determination of SYUIQ-5, a novel telomerase inhibitor and anti-tumor drug, in rat liver microsomes was developed by using high-performance liquid chromatography with ultraviolet detection. SYUIQ-5 was incubated in vitro with liver microsomes from rats pre-treated with control vehicle, beta-naphthofIavone, phenobarbital, 20% ethanol or dexamethasone. The analytes were extracted with diethyl ether and separated a C(18) 5-microm analytical column. Elution was conducted with 30 mM dipotassium hydrogen phosphate (pH 8.0)-methanol-triethylamine (30:70:0.05, v/v/v) at a flow-rate of 1.0 ml/min and the detection of UV absorbance was conducted at 278 nm. Intra-day and inter-day precision and accuracy of the method were within 10%. The mean analytical recoveries of SYUIQ-5 ranged from 78.8 to 95.3%. The linearity of the calibration curve was in the range of 1.0-80.0 microM. The lower limit of quantification (LOQ) was 1.0 microM. Kinetic analysis showed that beta-naphthofIavone and dexamethasone significantly induced SYUIQ-5 metabolism, suggesting that cytochrome P450 1A and 3A are the major contributor to SYUIQ-5 metabolism in rat liver microsomes.     

In situ proteolysis for protein crystallization and structure determination

We tested the general applicability of in situ proteolysis to form protein crystals suitable for structure determination by adding a protease (chymotrypsin or trypsin) digestion step to crystallization trials of 55 bacterial and 14 human proteins that had proven recalcitrant to our best efforts at crystallization or structure determination. This is a work in progress; so far we determined structures of 9 bacterial proteins and the human aminoimidazole ribonucleotide synthetase (AIRS) domain.     

MicroRNA-200c promotes osteogenic differentiation of human bone mesenchymal stem cells through activating the AKT/β-Catenin signaling pathway via downregulating Myd88

During the human bone formation, the event of osteogenic differentiation of human bone mesenchymal stem cells (hBMSCs) is vital, and recent evidence has emphasized the important role of microRNAs (miRNAs) in osteogenic differentiation of hBMSCs. This study aims to examine the potential effects of miR-200c in osteogenic differentiation of hBMSCs and understand their underlying mechanisms. HBMSCs were obtained via human bone marrow. During osteogenic induction and differentiation, cells were transfected with different plasmids with the intention of investigating the roles of miR-200c on osteogenic differentiation, calcium salt deposition, alkaline-phosphatase (ALP) activity, mineralized nodule formation, osteocalcin (OCN) content, and proliferation of osteoblasts. Following transfection, dual luciferase reporter gene assay was conducted so as to explore the correlation between miR-200c and Myd88. Moreover, the AKT/β-Catenin signaling pathway was blocked with an AKT/β-Catenin inhibitor, AKTi, to investigate its involvement. The hBMSCs were successfully isolated from human bone marrow. Myd88 was determined as a target gene of miR-200c. Gain and loss-of-function assays confirmed that overexpression of miR-200c, or silencing of Myd88 promoted osteogenic differentiation, increased calcium salt deposition, ALP activity, mineralized nodule formation, and enhanced the proliferation of osteoblasts following osteogenic differentiation of hBMSCs. Meanwhile, the downregulation of miR-200c has been shown to have the opposite effect. Furthermore, these findings showed that the miR-200c overexpression activated the AKT/β-Catenin signaling pathway by targeting Myd88. To sum up, the miR-200c upregulation induces osteogenic differentiation of hBMSCs by activating the AKT/β-Catenin signaling pathway via the inhibition of Myd88, providing a target for treatment of bone repair.     

Platelet-derived growth factor BB mediated regulation of 12-lipoxygenase in porcine aortic smooth muscle cells

Platelet-derived growth factor BB (PDGF) is a potent mitogen and chemoattractant for vascular smooth muscle cells (VSMC). In the present study, we have examined the effects of PDGF on the 12-lipoxygenase (12-LO) pathway of arachidonate metabolism in porcine aortic VSMC (PVSMC). The rationale for this is previous studies showing that LO products have growth and chemotactic effects in VSMC and that another VSMC growth factor, angiotensin II, is a potent positive regulator of 12-LO activity and expression. We observed that PDGF causes a significant increase in the formation of the 12-LO product, 12-hydroxyeicosatetraenoic acid (12-HETE) in PVSMC. In addition, PDGF also markedly increased leukocyte-type 12-LO messenger RNA and protein expression. PDGF-induced PVSMC migration was inhibited significantly by two LO blockers but not by a cyclooxygenase blocker. Furthermore, although the proliferative effects of PDGF on PVSMC were not altered by cell culture under hyperglycemic conditions (25 mM glucose, HG), the chemotactic effects of PDGF as well as those of 10% fetal calf serum were significantly greater in cells cultured in HG as compared to normal glucose conditions (5.5 mM), thus indicating a potential new mechanism for the accelerated cardiovascular disease usually observed in diabetes. These results indicate a novel mechanism for the biological effects of PDGF in leading to cardiovascular disease. © 1996 Wiley-Liss, Inc.     

Parasite richness/sampling effort/host range: the fancy three-piece jigsaw puzzle

In this article, Jean-Fran?ois Guégan and Clive Kennedy propose an alternative explanation for the confounding effects of host geographical range and sampling effort on parasite species richness using pathway analysis procedure. They suggest that much of the species richness revealed by sampling effort is also a reflection of host range. Thus, the total contribution of host range logically incorporates a contribution from sampling effort. The implications of indirect effects of host range on richness estimates have not previously been discussed, and the authors here attempt to redress the balance. The contribution of host range to richness, as derived from control of sampling effort on richness estimates, therefore, is a mathematical expression that does not take into account the cause-and-effect nature of things.     

Receptome profiling identifies KREMEN1 and ASGR1 as alternative functional receptors of SARS-CoV-2

Host cellular receptors play key roles in the determination of virus tropism and pathogenesis.However,little is known about SARS-CoV-2 host receptors with the e...