Devang N. Patel

Interleukin-18-induced human coronary artery smooth muscle cell migration is dependent on NF-κB- and AP-1-mediated matrix metalloproteinase-9 expression and is inhibited by atorvastatin

The proliferation and migration of arterial smooth muscle cells (SMCs) are key events in the vascular restenosis that frequently follows angioplasty. Furthermore, SMC migration and neointimal hyperplasia are promoted by degradation of the extracellular matrix by matrix metalloproteinases (MMPs). Because we demonstrated previously that the proinflammatory and proatherogenic cytokine interleukin-18 (IL-18) stimulates SMC proliferation (Chandrasekar, B., Mummidi, S., Valente, A. J., Patel, D. N., Bailey, S. R., Freeman, G. L., Hatano, M., Tokuhisa, T., and Jensen, L. E. (2005) J. Biol. Chem. 280, 26263–26277), we investigated whether IL-18 induces SMC migration in an MMP-dependent manner and whether the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor atorvastatin can inhibit this response. IL-18 treatment increased both mRNA and protein expression of MMP9 in human coronary artery SMCs. Gel shift, enzyme-linked immunosorbent, and chromatin immunoprecipitation assays revealed a strong induction of IL-18-mediated AP-1 (c-Fos, c-Jun, and Fra-1) and NF-κB (p50 and p65) activation and stimulation of MMP9 promoter-dependent reporter gene activity in an AP-1- and NF-κB-dependent manner. Ectopic expression of p65, c-Fos, c-Jun, and Fra-1 induced MMP9 promoter activity. Specific antisense or small interfering RNA reagents for these transcription factors reduced IL-18-mediated MMP9 transcription. Furthermore, IL-18 stimulated SMC migration in an MMP9-dependent manner. Atorvastatin effectively suppressed IL-18-mediated AP-1 and NF-κB activation, MMP9 expression, and SMC migration. Together, our results indicate for the first time that the proatherogenic cytokine IL-18 induces human coronary artery SMC migration in an MMP9-dependent manner. Atorvastatin inhibits IL-18-mediated aortic SMC migration and has therapeutic potential for attenuating the progression of atherosclerosis and restenosis.

Interleukin-17 Stimulates C-reactive Protein Expression in Hepatocytes and Smooth Muscle Cells via p38 MAPK and ERK1/2-dependent NF-κB and C/EBPβ Activation

Elevated systemic levels of the acute phase C-reactive protein (CRP) are predictors of future cardiovascular events. There is evidence that CRP may also play a direct role in atherogenesis. Here we determined whether the proinflammatory interleukin (IL)-17 stimulates CRP expression in hepatocytes (Hep3B cell line and primary hepatocytes) and coronary artery smooth muscle cells (CASMC). Our results demonstrate that IL-17 potently induces CRP expression in Hep3B cells independent of IL-1β and IL-6. IL-17 induced CRP promoter-driven reporter gene activity that could be attenuated by dominant negative IκBα or C/EBPβ knockdown and stimulated both NF-κB and C/EBP DNA binding and reporter gene activities. Targeting NF-κB and C/EBPβ activation by pharmacological inhibitors, small interfering RNA interference and adenoviral transduction of dominant negative expression vectors blocked IL-17-mediated CRP induction. Overexpression of wild type p50, p65, and C/EBPβ stimulated CRP transcription. IL-17 stimulated p38 MAPK and ERK1/2 activation, and SB203580 and PD98059 blunted IL-17-mediated NF-κB and C/EBP activation and CRP transcription. These results, confirmed in primary human hepatocytes and CASMC, demonstrate for the first time that IL-17 is a potent inducer of CRP expression via p38 MAPK and ERK1/2-dependent NF-κB and C/EBPβ activation and suggest that IL-17 may mediate chronic inflammation, atherosclerosis, and thrombosis.

Stability of self-assembled monolayers on titanium and gold.

Methyl- and hydroxyl-terminated phosphonic acid self-assembled monolayers (SAMs) were coated on Ti from aqueous solution. Dodecyl phosphate and dodecyltrichlorosilane SAMs were also coated on Ti using solution-phase deposition. The stability of SAMs on Ti was investigated in Tris-buffered saline (TBS) at 37 degrees C using X-ray photoelectron spectroscopy, contact angle goniometry, and atomic force microscopy. For comparison purposes, a hydroxyl-terminated thiol SAM was coated on Au, and its stability was also investigated under similar conditions. In TBS, a significant proportion of phosphonic acid or phosphate molecules were desorbed from the Ti surface within 1 day, while the trichlorosilane SAM on Ti or thiol SAM on Au was stable for up to 7 days under similar conditions. The stability of hydroxyl-terminated phosphonic acid SAM coated Ti and thiol SAM coated Au was investigated in ambient air and ultraviolet (UV) light. In ambient air, the phosphonic acid SAM on Ti was stable for up to 14 days, while the thiol SAM on Au was not stable for 1 day. Under UV-radiation exposure, the alkyl chains of the phosphonic acid SAM were decomposed, leaving only the phosphonate groups on the Ti surface after 12 h. Under similar conditions, decomposition of alkyl chains of the thiol SAM was observed on the Au surface accompanied by oxidation of thiolates.

Drug delivery from gold and titanium surfaces using self-assembled monolayers.

Currently available drug-eluting stents (DES) use polymers for coating and releasing drugs. Increasing evidence suggests that inflammatory and hypersensitive reactions are caused by such polymer coatings. This study focused on developing new techniques for delivering drugs directly from metal implant surfaces. Hydroxyl-terminated self-assembled monolayers (SAMs) were coated on Au and Ti surfaces. Therapeutic self-assembled monolayers (TSAMs) were prepared by chemically attaching the model drug, flufenamic acid, to SAM coated metal surfaces. Three different methods of esterification (acid chloride esterification, dry heat esterification, and direct esterification) were explored to attach flufenamic acid to SAMs. TSAMs were characterized using X-ray photoelectron spectroscopy, fluorescence microscopy, atomic force microscopy, and contact angle goniometry. These techniques collectively confirmed the attachment of drug onto SAM coated metal surfaces. In vitro drug release was investigated by immersing TSAM coated metal specimens in tris-buffered saline (TBS) at 37 degrees C for 28 days. TBS was analyzed at 1, 3, 7, 14, 21, and 28 days for the amount of drug eluted using high performance liquid chromatography. Large data scatter was observed for the release profiles of TSAMs prepared by acid chloride esterification. TSAMs prepared by dry heat and direct esterification methods showed an initial burst release of the drug followed by a sustained slow release for up to 2 weeks. Thus, this study suggests the potential for using self-assembled monolayers as an alternate system for delivering drugs from coronary stents and other metal implants.

Drug loading of nanoporous TiO2 films

The loading of therapeutic amounts of drug on a nanoporous TiO(2) surface is described. This novel drug-loading scheme on a biocompatible surface, when employed on medical implants, will benefit patients who require the deployment of drug-eluting implants. Anticoagulants, analgesics and antibiotics can be considered on the associated implants for drug delivery during the time of maximal pain or risk for patients undergoing orthopedic procedures. Therefore, this scheme will maximize the chances of patient recovery.

Interaction of Endothelial Cells with Self-Assembled Monolayers for Potential Use in Drug-Eluting Coronary Stents

Drug-eluting stents (DES) are implanted in patients to treat in-stent restenosis. Commercially available DES use polymers for coating and releasing drugs. Several studies have showed that polymer coatings cause adverse reactions. Delayed endothelialization of polymer-coated DES leads to late stent thrombosis. Recently, the potential for using self-assembled monolayers (self-assembled monolayers (SAMs)-organic constructs composed of (a) chemical groups which attach to metal surfaces, (b) long hydrocarbon chains, and (c) terminal functional groups) as an alternate drug delivery system for coronary stents has been demonstrated. In this study, the interaction of human aortic endothelial cells (HAECs) with SAMs and therapeutic SAMs (therapeutic self-assembled monolayers (TSAMs)-SAMs derivatized with the drug, flufenamic acid) was investigated. HAECs were cultured on plain glass, control, SAMs-, and TSAMs-coated titanium (Ti) and gold (Au) specimens. The viability and proliferation of HAECs were investigated using MTT colorimetric assay. The adhesion of HAECs on SAMs and TSAMs was equivalent to that of control metal surfaces and superior to that of plain glass surfaces. The cells continued to proliferate on both SAMs and TSAMs even though the rate of proliferation was slower than plain glass or control-Ti. The spreading of HAECs on TSAMs with typical polygonal shape indicated that these surfaces are conducive to endothelialization. The expression of surface adhesion protein, platelet endothelial cell adhesion molecule-1, on TSAMs indicated that the endothelial cells preserved their phenotype on these surfaces. Thus, this study demonstrated that SAMs and TSAMs do not elicit an adverse response from endothelial cells in in vitro conditions.

Nanotechnology in cardiovascular medicine.

Nanotechnology is a new field of science and technology that has already had significant impact in the development of novel products in industry. In medicine, application of nanotechnology has the potential to develop new imaging agents, pharmaceutical drugs and medical devices with unique physical and chemical properties. This article reviews the potential for various nanoparticles in cardiovascular imaging and therapeutics, nanoporous structures for sensing and implant based drug delivery, and self‐assembled monolayers for surface modification and implant based drug delivery.

Interleukin-18 Suppresses Adiponectin Expression in 3T3-L1 Adipocytes via a Novel Signal Transduction Pathway Involving ERK1/2-dependent NFATc4 Phosphorylation

An inverse correlation between the pro-inflammatory cytokine interleukin-18 and the anti-atherogenic adipokine adiponectin has been reported in the chronic pathological conditions obesity, insulin resistance, coronary artery disease, and metabolic syndrome. We investigated whether this relationship is coincidental or has a causal basis. Here we show that interleukin-18 (IL-18) suppresses adiponectin transcription, mRNA expression, and secretion by 3T3-L1 adipocytes. IL-18 suppresses adiponectin promoter-reporter activity, an effect reversed by deletion or mutation of the NFATc4 core DNA-binding site. IL-18 induces NFATc4 phosphorylation (Ser676), nuclear translocation, and in vivo DNA binding. IL-18 induces ERK1/2 phosphorylation and enzyme activity, and pretreatment with the MEK inhibitor U0126, ERK1/2 inhibitor PD98059, or small interference RNA targeted to ERK1/2 attenuates ERK1/2 activation and NFATc4 phosphorylation. Finally, inhibition of ERK1/2 or NFATc4 knockdown reverses IL-18-mediated adiponectin suppression. In contrast to its inhibitory effects on adiponectin expression, IL-18 potently stimulates PAI-1 secretion. These data demonstrate for the first time that IL-18 selectively suppresses adiponectin expression via ERK1/2-dependent NFATc4 activation and suggest that the inverse relationship observed between IL-18 and adiponectin in various chronic pathological conditions is causally related. Thus, targeting IL-18 expression may enhance adiponectin expression and mitigate disease progression.

Long-term effects of novel biolimus eluting DEVAX AXXESS plus nitinol self-expanding stent in a porcine coronary model

Objective: The purpose of this study was to evaluate the long‐term effects of the DEVAX AXXESS biolimus eluting stent (BES) in a porcine coronary model, compared with those of bare metal stent (BMS) and polymer only stent (POS) controls. Background: Excessive neointimal growth has been identified as a major cause of late failure of percutaneous coronary interventions. The effect of drug eluting from self‐expanding stents for prevention of neointimal hyperplasia has not been studied before. The DEVAX AXXESS is a self‐expanding nickel titanium stent, coated with antiproliferative compound‐biolimus. Methods: Twenty juvenile farm swine, 25–35 kg in weight, 3–6 months in age were used. Each animal received a stent to the left anterior descending artery, left circumflex or right coronary arteries as permitted per anatomy. The chronic vascular response after BES implantation was compared with that after BMS and POS implantation at 28, 90, and 180 days follow‐up. Results: The 28‐day outcome by quantitative coronary angiography (QCA) showed significant increase in minimal luminal diameter (MLD) in the BES (MLD: 2.90 ± 0.97, 2.39 ± 0.90, 1.59 ± 0.91; P = 0.009) compared with BMS and POS, respectively. By histomorphometric analysis, there was also a corresponding significant reduction in neointimal tissue proliferation in the BES (average neointimal area: 2.78 ± 0.07, 5.46 ± 0.66, 8.42 ± 0.85; P = 0.002) compared with that in BMS and POS controls, respectively at 28‐days follow‐up. At 90 and 180 days, the mean neointimal area was not significantly different between the BES and the controls. Conclusions: BES favorably modulates the neointimal tissue formation for 28 days, in the porcine coronary model. Long‐term inhibition of neointimal hyperplasia is not sustained most likely because of the delayed cellular proliferation and inflammation in the vessel wall.

Periodontal disease in association with systemic levels of interleukin-18 and CXC ligand 16 in patients undergoing cardiac catheterization

BACKGROUND Interleukin (IL)-18 is a proinflammatory cytokine that is present in chronically inflamed tissues; IL-18 was positively associated with periodontitis and coronary artery disease (CAD). CXC ligand (CXCL) 16, a recently discovered chemokine, was identified in atherosclerotic lesions; its role in periodontal disease is largely unknown. This research study correlates periodontal parameters with systemic levels of IL-18 and CXCL16. METHODS Fifty-one patients who presented for clinically indicated coronary angiography received full-mouth periodontal examinations. The periodontal status of patients was defined using frequency distributions of probing depth (PD), clinical attachment loss (AL), and bleeding on probing (BOP). Blood samples were collected during cardiac catheterization, and plasma levels of IL-18 and CXCL16 were analyzed. The severity of CAD was determined by the presence and extent of coronary artery stenosis. Correlations between periodontal parameters, levels of inflammatory mediators, and CAD status were analyzed. RESULTS The extent of BOP exhibited a significant positive correlation with IL-18 in the Spearman rank correlation analysis (P = 0.039), which indicated a correlation between periodontal inflammation and systemic IL-18 levels. When multiple regression analysis was performed, the extent of clinical AL > or =3 mm (P = 0.045) and > or =5 mm (P = 0.024) exhibited an association with IL-18, whereas CXCL16 was associated with clinical AL > or =5 mm (P = 0.040) and PD > or =7 mm (P = 0.047). CONCLUSION A significant correlation is identified between periodontitis and systemic levels of IL-18 and CXCL16 in patients undergoing diagnostic coronary angiography.