Bao-Di Gou

Realgar-induced differentiation is associated with MAPK pathways in HL-60 cells

The clinical efficacy and safety of realgar (arsenic sulfide, As4S4) in the treatment of acute promyelocytic leukemia in China have given rise to an upsurge in research on the underlying mechanism. We prepared realgar nanoparticles (RNPs) to examine their effect on the differentiation of HL‐60 cells. Treatment with RNPs at 6 μM for 72 h induced cell differentiation that was assessed by morphological change, NBT reductive ability, and elevation of CD11b expression at both mRNA and protein levels. The RNP‐induced differentiation was synergized, enhanced and suppressed by the inhibition of p38 MAPK, JNK and ERK pathways, respectively. Our findings demonstrate that MAPK signaling pathways are closely related to the RNP‐induced differentiation in HL‐60 cells.

Lanthanum suppresses osteoblastic differentiation via pertussis toxin-sensitive G protein signaling in rat vascular smooth muscle cells.

A major cellular event in vascular calcification is the phenotypic transformation of vascular smooth muscle cells (VSMCs) into osteoblast‐like cells. After demonstrating that lanthanum chloride (LaCl3) suppresses hydrogen peroxide‐enhanced calcification in rat calcifying vascular cells (CVCs), here we report its effect on the osteoblastic differentiation of rat VSMCs, a process leading to the formation of CVCs. Cells were isolated from aortic media of male SD rats, and passages between three and eight were cultured in Dulbeccols Modified Eagles Medium (DMEM) containing 10% fetal bovine serum (FBS) and 10 mM β‐glycerophosphate (β‐GP) in the presence or absence of LaCl3. Exposure of cells to LaCl3 suppressed the β‐GP‐induced elevations in calcium deposition, alkaline phosphatase (ALP) activity, and Cbfa1/Runx2 expression, as well as the concomitant loss of SM α‐actin. Furthermore, LaCl3 activated the phosphorylation of extracellular signal‐regulated kinase (ERK) and c‐Jun N‐terminal kinase (JNK), and the blockage of either pathway with a specific inhibitor abolished the effects of LaCl3. In addition, pretreatment of the cells with pertussis toxin (PTx), an inhibitor of G protein‐mediated signaling pathway, repealed all the changes induced by LaCl3. These findings demonstrate that LaCl3 suppresses the β‐GP‐induced osteoblastic differentiation and calcification in rat VSMCs, and its effect is mediated by the activation of both ERK and JNK MAPK pathways via PTx‐sensitive G proteins. J. Cell. Biochem. 108: 1184–1191, 2009.

Gadolinium-promoted precipitation of calcium phosphate is associated with profibrotic activation of RAW 264.7 macrophages.

Gadolinium-based contrast agents are now being linked to nephrogenic systemic fibrosis (NSF). The exact mechanism by which gadolinium species act in the pathogenesis of NSF is not fully understood. In this study, we evaluated the effect of gadolinium chloride (GdCl(3)) on the precipitation of calcium phosphate, and examined the role of the gadolinium-containing precipitates in the profibrotic activation of macrophages. In a free-drift system, the induction time was markedly reduced with increasing concentration of GdCl(3), accompanied by alterations of morphology and composition of the precipitates. In complete cell culture medium, the addition of GdCl(3) resulted in formation of particles around 200-300 nm. In an in vitro cellular model with RAW 264.7 macrophages, GdCl(3) increased the production of TGF-beta1 and IL-6 via the activation of PKC and ERK signaling pathway. Our findings demonstrate that GdCl(3) promotes calcium phosphate precipitation and induces profibrotic activation of macrophages.

Lanthanum chloride bidirectionally influences calcification in bovine vascular smooth muscle cells

Vascular calcification (VC) is frequent prevalence in patients with chronic kidney disease (CKD) and atherosclerosis. Lanthanum carbonate is used as an orally administered phosphate‐binding agent to reduce the gastrointestinal absorption of phosphate and ameliorate VC in advanced CKD. In this study, we used bovine vascular smooth muscle cells as a model VC in vitro and studied the effects of lanthanum chloride on calcium deposition. Exposure of cells to LaCl3 at the concentration of 0.1 µM suppressed the β‐glycerophosphate‐induced alkaline phosphatase activity and calcium deposition. Furthermore, LaCl3 upregulated the β‐glycerophosphate‐suppressed expression of calcium‐sensing receptor. In contrast to the inhibitory effect of LaCl3 on calcium deposition, higher level lanthanum (50 µM) was found to promote immediately precipitation of calcium phosphate in cell culture medium. At this concentration, LaCl3 was found to induce cell apoptosis which involves caspases‐9 and ‐3. These data indicate that the promotory effect of LaCl3 on calcium deposition is likely mediated by induction of apoptosis. Our in vitro findings do suggest that, in the context of raised lanthanum, greater attention should be paid to potential toxic effects associated to the use of lanthanide‐based drugs. J. Cell. Biochem. 113: 1776–1786, 2012.

Association of oxidative stress with realgar-induced differentiation in human leukemia HL-60 cells.

Background: Realgar (arsenic sulfide, As4S4) has been shown to have clinical efficacy in patients with newly diagnosed and relapsed acute promyelocytic leukemia. Mechanistic studies have demonstrated that realgar is able to induce cell differentiation. Methods: The oxidative stress in the realgar-induced differentiation was examined with human leukemia HL-60 cells. Cell differentiation was evaluated by the expression of cell surface antigen CD11b and nitroblue tetrazolium assay. The activities of catalase and superoxide dismutase were measured spectrophotometrically. Flow cytometry was used to assess cell cycle distribution and apoptosis, the cellular level of reactive oxygen species (ROS) and glutathione, as well as mitochondrial transmembrane potential (MTP). Results: The realgar-induced differentiation was enhanced by hydrogen peroxide, and preceded with drastic changes in ROS and catalase, as well as small changes in superoxide dismutase and the reduced form of glutathione. MTP values at 24 h were in linear proportion to the CD11b expression at 48 h when no apoptosis was observed. Conclusion: Oxidative stress and stress-related MTP decrease are associated with realgar-induced differentiation in HL-60 cells.

Preparation of Realgar Nanoparticle Suspension and Its Inhibition Effect on the Proliferation of Human Myelocytic Leukaemia HL-60 Cells

Realgar (As4S4) has recently been proved effective in the treatment of leukaemia in clinical trials. However, the poor solubility of realgar makes it difficult to conduct mechanistic study in cellular level. In this work, we prepared realgar nanoparticle (RNP) suspension and examined its effect on the proliferation of human myelocytic leukaemia HL-60 cells. The average diameter of the particles was 143 nm and was stable against coalescence over a 15-month storage. The suspension inhibited cell proliferation dose-dependently at concentrations from 10 to 60 µM (As), a fact shedding light on the mechanism of realgars clinical effectiveness against hematopoietic malignancies.

Quantitative chemical relations at pseudo-equilibrium in amorphous calcium phosphate formation

Amorphous calcium phosphate (ACP) is a precursor to crystalline hydroxyapatite, and has been found to be present in zebrafish and mouse bone formation. Most of the previous studies on ACP formation are qualititative in nature. Here, we report a quantitative correlation between the solution composition and the amount of ACP. Solutions with the initial Ca/P molar ratios between 0.5 and 1.5 were made either by fixing calcium and varying phosphate concentration (Series 1), or by fixing phosphate and varying calcium concentration (Series 2). From the two reaction series, we derived pseudo-equilibrium equations, and determined the threshold concentrations for ACP formation and its maximum amount, which are present as constants in these equations. Based on these results, we proposed a mechanism for ACP formation and derived the formation constant that depends on both the solution composition and the ACP amount. This work might represent an advance toward understanding the basic aspects of solution chemistry involving clusters and an amorphous phase of calcium phosphate, and the “two series” approach could be applicable to the systematical study of the amorphous phase of other sparingly soluble electrolytes. Moreover, these findings might provide new insight into the relevant physiological phenomena and be helpful for the rational design of functional materials.