Sung-Yu Hong

The snake venom disintegrin salmosin induces apoptosis by disassembly of focal adhesions in bovine capillary endothelial cells.

Salmosin, a disintegrin purified from a Korean snake (Agkistrodon halys brevicaudus) venom, interacts with integrin alpha(v)beta(3) and inhibits the proliferation of bovine capillary endothelial (BCE) cells induced by basic fibroblast growth factor (bFGF). We investigated salmosins mechanism of inhibition of BCE cell proliferation by examining changes in the cytoskeleton and activation of integrin-mediated signaling molecules. Salmosin disassembled cortical actins at focal adhesions and induced cells to be rounded and detached, but it did not alter microtubule structures in the early stage of cells being rounded. Immunolocalization of paxillin also demonstrated that focal adhesions were disassembled by salmosin. In salmosin-treated BCE cells, focal adhesion kinase (FAK) was dephosphorylated and expression of paxillin and p130(CAS) was decreased, but PI3 kinase, ILK, and beta-catenin were not expressed in decreased amounts or modified, suggesting that salmosin inactivated FAK-dependent integrin signaling pathways. While BCE cells proliferated normally on plates coated with salmosin, cells treated with salmosin eventually underwent apoptosis. These observations strongly suggest that salmosin disorganizes focal contacts to detach cells by competing with the extracellular matrix (ECM) for direct binding to integrin alpha(v)beta(3) on the cell surface, eventually leading to apoptosis.

Soluble Receptor for Advanced Glycation End Products Alleviates Nephritis in (NZB/NZW)F1 Mice

OBJECTIVE To investigate the efficacy of different doses of the soluble form of the receptor for advanced glycation end products (sRAGE) (conjugated to the Fc portion of immunoglobulin) in the treatment of nephritis in lupus-prone mice, in comparison with the efficacy of combination therapy with mycophenolate mofetil plus prednisolone. METHODS Twenty-eight female (NZB/NZW)F1 mice were divided into 5 groups (untreated, sRAGE [dose groups of 0.5, 1, or 2 μg], or mycophenolate mofetil plus prednisolone). Proteinuria and histologic damage were evaluated. Immune complex deposition and the nuclear translocation of NF-κB in the kidney tissue were assessed by immunofluorescence staining. Serum concentrations of anti-double-stranded DNA (anti-dsDNA) and IgG subclasses were also measured. The population of T cells was evaluated using a fluorescence-activated cell sorter, and expression of intracellular adhesion molecule 1 and vascular cell adhesion molecule 1 in the kidney tissue was assessed by immunohistochemical staining. RESULTS In comparison with untreated mice, mice treated with 1 or 2 μg sRAGE showed significantly reduced proteinuria and attenuated histologic renal damage, with efficacy comparable to that of combination therapy. Treatment with 2 μg sRAGE significantly reduced immune complex deposition and decreased the serum concentrations of anti-dsDNA, IgG2a, IgG2b, and IgG3. In addition, sRAGE interrupted the nuclear translocation of NF-κB in the kidney, resulting in reduction in the expression of downstream genes of NF-κB in vivo and in vitro. Furthermore, sRAGE effectively modified T cell populations. CONCLUSION Treatment with sRAGE significantly improved nephritis in lupus-prone mice, with efficacy comparable to that of standard induction treatment for lupus nephritis. These data suggest that sRAGE has antiinflammatory effects on the pathophysiology of lupus nephritis and could serve as a potent new therapy for this disease.

High Serum Advanced Glycation End-Products Predict Coronary Artery Disease Irrespective of Arterial Stiffness in Diabetic Patients

Background and Objectives Advanced glycation end-products (AGEs) contribute to the development of atherosclerosis. We investigated whether serum AGEs are related to the presence or severity of coronary artery disease (CAD), and explored the association between serum AGEs and arterial stiffness according to diabetes status in patients suspected of having CAD. Subjects and Methods The measurement of serum AGEs and brachial-ankle pulse wave velocity (baPWV) were performed in 145 consecutive patients (63±9 years, 58% men) who received a coronary angiogram for evaluation of CAD. Results Forty-four diabetics and 101 non-diabetics were classified into three subgroups based on the number of diseased vessels with obstructive CAD: 0, 1, and 2 or more vessel diseases (VDs). Serum AGEs were significantly higher in diabetics with obstructive CAD than in those without obstructive CAD (2.16±0.29 vs. 1.85±0.29 mU/mL, p=0.010) and were significantly correlated with the number of VDs only in diabetics (r=0.504, p<0.001). Serum AGEs were not significantly correlated with baPWV in diabetics or non-diabetics. In receiver operating characteristics analysis, the cut-off value of serum AGEs as a predictor of obstructive CAD was 1.98 mU/mL, with 64% sensitivity and 63% specificity in diabetics. In multiple regression analysis, serum AGEs independently predicted obstructive CAD and were associated with the number of VDs in diabetics. Conclusion Serum AGEs independently predict obstructive CAD and the severity of coronary atherosclerosis irrespective of arterial stiffness only in diabetics. Evaluation of PWV and serum AGEs together may be more effective to identify the risk of CAD in diabetic individuals.

Suppressive effect and mechanism of saxatilin, a disintegrin from Korean snake (Gloydius saxatilis), in vascular smooth muscle cells

RGD-peptides can inhibit the binding of ligands to certain beta3 integrins, alphaIIbbeta3 and alphavbeta3, both of which are involved in neointimal hyperplasia that contributes to atherosclerosis and restenosis of arterial walls. Saxatilin, a disintegrin from a Korean snake (Gloydius saxatilis), interacts with integrins alphaIIbbeta3 and alphavbeta3. It suppressed the adhesion of human coronary artery smooth muscle cells (HCASMCs) to vitronectin with an IC(50) of 2.5 microM, and growth factor (PDGF-BB or bFGF)-induced proliferation was inhibited at an IC(50) of 25 microM. Saxatilin disassembled the actin cytoskeleton of focal adhesion and induced cell detachment. This disassembly of focal adhesion in saxatilin-treated HCASMCs involved caspase-induced paxillin degradation. Saxatilin temporally phosphorylated FAK and ERKs and affected the cell cycle of HCASMCs by increasing CDK inhibitors (p21 and p27) and reducing cyclins (D1/2 and E). These results may have significant implications for integrin antagonistic therapy used for the treatment of atherosclerosis and restenosis.

Targeted ultrasound imaging of apoptosis with annexin a5 microbubbles in acute Doxorubicin-induced cardiotoxicity.

BACKGROUND The aim of this study was to assess the feasibility of targeted ultrasound imaging on apoptosis with annexin A5 microbubbles (A5MB) in acute doxorubicin-induced cardiotoxicity. METHODS Avidinated and octafluoropropan-filled phospholipid microbubbles were conjugated with biotinylated annexin A5. To confirm the specific binding of A5MB, flow cytometry was performed with hydrogen peroxide induced apoptosis in rat aorta smooth muscle cells incubated with fluorescein-5-isothiocyanate (FITC) labeled annexin A5 and A5MB. Adult male rats were injected intraperitoneally with 5 mg/kg doxorubicin weekly for 3 weeks (n = 5). Control rats were injected with normal saline (n = 5). At 24 hours after the final treatment, triggering imaging was performed 15 min after an intravenous bolus injection of A5MB for washout of freely circulating microbubbles. After echocardiography, the heart was isolated for histological detection of apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. RESULTS In the in vitro tests, fluorescence intensity was low for healthy cells and high for apoptotic cells when incubated with FITC-labeled annexin A5 and A5MB. Rats treated with doxorubicin showed significant contrast opacification of the myocardium on contrast echocardiography using A5MB. However, no opacification was observed in control rats. Apoptosis was confirmed by TUNEL assay in doxorubicin treated rats. CONCLUSION Acute doxorubicin-induced cardiomyopathy based on early apoptosis can be assessed and imaged with targeted ultrasound imaging using A5MB in rats.