Aditi Sinha

High-glucose levels and elastin degradation products accelerate osteogenesis in vascular smooth muscle cells

Diabetes mellitus (DM) is a chronic disease in which the body either does not use or produce the glucose metabolising hormone insulin efficiently. Calcification of elastin in the arteries of diabetics is a major predictor of cardiovascular diseases. It has been previously shown that elastin degradation products work synergistically with transforming growth factor-beta 1 (TGF-β1) to induce osteogenesis in vascular smooth muscle cells. In this study, we tested the hypothesis that high concentration of glucose coupled with elastin degradation products and TGF-β1 (a cytokine commonly associated with diabetes) will cause a greater degree of osteogenesis compared to normal vascular cells. Thus, the goal of this study was to analyse the effects of high concentration of glucose, elastin peptides and TGF-β1 on bone-specific markers like alkaline phosphatase (ALP), osteocalcin (OCN) and runt-related transcription factor 2 (RUNX2). We demonstrated using relative gene expression and specific protein assays that elastin degradation products in the presence of high glucose cause the increase in expression of the specific elastin–laminin receptor-1 (ELR-1) and activin receptor-like kinase-5 (ALK-5) present on the surface of the vascular cells, in turn leading to overexpression of typical osteogenic markers like ALP, OCN and RUNX2. Conversely, blocking of ELR-1 and ALK-5 strongly suppressed the expression of the osteogenic proteins. In conclusion, our results indicate that glucose plays an important role in amplifying the osteogenesis induced by elastin peptides and TGF-β1, possibly by activating the ELR-1 and ALK-5 signalling pathways.

Efficacy of Reversal of Aortic Calcification by Chelating Agents

Elastin-specific medial vascular calcification, termed “Monckeberg’s sclerosis,” has been recognized as a major risk factor for various cardiovascular events. We hypothesize that chelating agents, such as disodium ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), and sodium thiosulfate (STS) might reverse elastin calcification by directly removing calcium from calcified tissues into soluble calcium complexes. We assessed the chelating ability of EDTA, DTPA, and STS on removal of calcium from hydroxyapatite (HA) powder, calcified porcine aortic elastin, and calcified human aorta in vitro. We show that both EDTA and DTPA could effectively remove calcium from HA and calcified tissues, while STS was not effective. The tissue architecture was not altered during chelation. In the animal model of aortic elastin-specific calcification, we further show that local periadventitial delivery of EDTA loaded in to poly(lactic-co-glycolic acid) nanoparticles regressed elastin-specific calcification in the aorta. Collectively, the data indicate that elastin-specific medial vascular calcification could be reversed by chelating agents.

Nanoparticle targeting to diseased vasculature for imaging and therapy

UNLABELLED Significant challenges remain in targeting drugs to diseased vasculature; most important being rapid blood flow with high shear, limited availability of stable targets, and heterogeneity and recycling of cellular markers. We developed nanoparticles (NPs) to target degraded elastic lamina, a consistent pathological feature in vascular diseases. In-vitro organ and cell culture experiments demonstrated that these NPs were not taken up by cells, but instead retained within the extracellular space; NP binding was proportional to the extent of elastic lamina damage. With three well-established rodent models of vascular diseases such as aortic aneurysm (calcium chloride mediated aortic injury in rats), atherosclerosis (fat-fed apoE-/- mice), and vascular calcification (warfarin + vitamin K injections in rats), we show precise NPs spatial targeting to degraded vascular elastic lamina while sparing healthy vasculature when NPs were delivered systemically. Nanoparticle targeting degraded elastic lamina is attractive to deliver therapeutic or imaging agents to the diseased vasculature. FROM THE CLINICAL EDITOR This novel work focuses on nanoparticle targeting of degraded elastic lamina in a variety of diseases, including atherosclerosis, vascular calcification, and aneurysm formation, and demonstrates the feasibility to deliver therapeutic or imaging agents to the diseased vasculature.

Prevention of abdominal aortic aneurysm progression by targeted inhibition of matrix metalloproteinase activity with batimastat-loaded nanoparticles.

RATIONALE Matrix metalloproteinases (MMPs)-mediated extracellular matrix destruction is the major cause of development and progression of abdominal aortic aneurysms. Systemic treatments of MMP inhibitors have shown effectiveness in animal models, but it did not translate to clinical success either because of low doses used or systemic side effects of MMP inhibitors. We propose a targeted nanoparticle (NP)-based delivery of MMP inhibitor at low doses to the abdominal aortic aneurysms site. Such therapy will be an attractive option for preventing expansion of aneurysms in patients without systemic side effects. OBJECTIVE Our previous study showed that poly(d,l-lactide) NPs conjugated with an antielastin antibody could be targeted to the site of an aneurysm in a rat model of abdominal aortic aneurysms. In the study reported here, we tested whether such targeted NPs could deliver the MMP inhibitor batimastat (BB-94) to the site of an aneurysm and prevent aneurysmal growth. METHODS AND RESULTS Poly(d,l-lactide) NPs were loaded with BB-94 and conjugated with an elastin antibody. Intravenous injections of elastin antibody-conjugated BB-94-loaded NPs targeted the site of aneurysms and delivered BB-94 in a calcium chloride injury-induced abdominal aortic aneurysms in rats. Such targeted delivery inhibited MMP activity, elastin degradation, calcification, and aneurysmal development in the aorta (269% expansion in control versus 40% elastin antibody-conjugated BB-94-loaded NPs) at a low dose of BB-94. The systemic administration of BB-94 alone at the same dose was ineffective in producing MMP inhibition. CONCLUSIONS Targeted delivery of MMP inhibitors using NPs may be an attractive strategy to inhibit aneurysmal progression.

Elasto-regenerative properties of polyphenols

Abdominal aortic aneurysms (AAA) are progressive dilatations of infra-renal aorta causing structural weakening rendering the aorta prone to rupture. AAA can be potentially stabilized by inhibiting inflammatory enzymes such as matrix metalloproteinases (MMP); however, active regression of AAA is not possible without new elastic fiber regeneration. Here we report the elastogenic benefit of direct delivery of polyphenols such as pentagalloyl glucose (PGG), epigallocatechin gallate (EGCG), and catechin, to smooth muscle cells obtained either from healthy or from aneurysmal rat aorta. Addition of 10 μg/ml PGG and ECGC induce elastin synthesis, organization, and crosslinking while catechin does not. Our results indicate that polyphenols bind to monomeric tropoelastin and enhance coacervation, aid in crosslinking of elastin by increasing lysyl oxidase (LOX) synthesis, and by blocking MMP-2 activity. Thus, polyphenol treatments leads to increased mature elastin fibers synthesis without increasing the production of intracellular tropoelastin.

Periostin as an early marker for elastin mediated vascular smooth muscle cell calcification

Abstract Rationale: Medial vascular calcification (MAC) has been shown to be an active, cell-mediated process. Although several factors have been shown to promote vascular calcification