The normal function of poly (ADP-ribose) polymerase-1 (PARP-1) is the routine repair of DNA damage by adding poly (ADP ribose) polymers in response to a variety of cellular stresses. Recently, it has become widely appreciated that PARP-1 also participates in diverse physiological and pathological functions from cell survival to several forms of cell death and has been implicated in gene transcription, immune responses, inflammation, learning, memory, synaptic functions, angiogenesis and aging. In the CNS, PARP inhibition attenuates injury in pathologies like cerebral ischemia, trauma and excitotoxicity demonstrating a central role of PARP-1 in these pathologies. PARP-1 is also a preferred substrate for several 'suicidal' proteases and the proteolytic action of suicidal proteases (caspases, calpains, cathepsins, granzymes and matrix metalloproteinases (MMPs)) on PARP-1 produces several specific proteolytic cleavage fragments with different molecular weights. These PARP-1 signature fragments are recognized biomarkers for specific patterns of protease activity in unique cell death programs. This review focuses on specific suicidal proteases active towards PARP-1 to generate signature PARP-1 fragments that can identify key proteases and particular forms of cell death involved in pathophysiology. The roles played by some of the PARP-1 fragments and their associated binding partners in the control of different forms of cell death are also discussed.

Inflammatory bowel diseases (IBD) are a complex group of diseases involving alterations in mucosal immunity and gastrointestinal physiology during both initiation and progressive phases of the disease. At the core of these alterations are endothelial cells, whose continual adjustments in structure and function coordinate vascular supply, immune cell emigration, and regulation of the tissue environment. Expansion of the endothelium in IBD (angiogenesis), mediated by inflammatory growth factors, cytokines and chemokines, is a hallmark of active gut disease and is closely related to disease severity. The endothelium in newly formed or inflamed vessels differs from that in normal vessels in the production of and response to inflammatory cytokines, growth factors, and adhesion molecules, altering coagulant capacity, barrier function and blood cell recruitment in injury. This review examines the roles of the endothelium in the initiation and propagation of IBD pathology and distinctive features of the intestinal endothelium contributing to these conditions.

The mobilization and recruitment of blood and lymphatic vasculatures are widely described in inflammatory bowel diseases (IBDs). Although angiogenesis contributes to intense gut inflammation, it remains unclear whether and when lymphangiogenesis amplifies or protects in IBD. The prolonged maintenance of lymphatic (over blood vessels) in inflammation indicates that lymphatic-blood vessel interactions may regulate IBD pathogenesis and restitution. Although lymphatic expansion helps to restore fluid balance and clear cytokines and immune cells, lymphatic failure results in accumulation of these factors and exacerbates IBD. Lymphatic obstruction and remodeling may impair lymphatic pumping, leading to repeated rounds of lymphangiogenesis. Early descriptions of Crohn's disease and ulcerative colitis describe colon lymphatic congestion, remodeling, expansion, and many other features that are recapitulated in experimental IBD and also by intestinal lymphatic obstruction, supporting lymphangitis as a cause and consequence of IBD. Growth factors, cytokines, gut flora, Toll receptors, and leukocytes all regulate inflammation and gut lymphatic remodeling in IBD. This review summarizes the importance of lymphatics and lymphangiogenesis in IBD etiology that may be useful in diagnosis and therapy of gut inflammation.

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Background: Inflammatory cytokines dysregulate microvascular function, yet how cytokines affect lymphatic endothelial cells (LEC) are unclear. Methods and Results: We examined effects of TNF-a, IL-1b, and IFN-g on LEC proliferation, endothelial cell adhesion molecule (ECAM) expression, capillary formation, and barrier changes in murine (SV-LEC) and human LECs (HMEC-1a). Results: All cytokines induced ICAM-1, VCAM-1, MAdCAM-1, and E-selectin in SV-LECs; TNF-a, IL-1b and IFN-g induced ECAMs (but not MAdCAM-1) in HMEC-1a. IL-1b increased, while IFN-g and TNF-a reduced SV-LEC proliferation. While TNF-a induced, IFN-g decreased, and IL-1b did not show any effect on HMEC-1a proliferation. TNF-a, IL-1b, and IFN-g each reduced capillary formation in SV-LEC and in HMEC-1a. TNF-a and IL-1b reduced barrier in SV-LEC and HMEC-1a; IFN-g did not affect SV-LEC barrier, but enhanced HMEC-1a barrier. Inflammatory cytokines alter LEC growth, activation and barrier function in vitro and may disturb lymphatic clearance increasing tissue edema in vivo. Conclusion: Therapies that maintain or restore lymphatic function (including cytokines blockade), may represent important strategies for limiting inflammation.

Background Myocarditis is an inflammatory disease of the cardiac muscle and is mainly caused by viral infections. Viral myocarditis has been proposed to be divided into 3 phases: the acute viral phase, the subacute immune phase, and the chronic cardiac remodeling phase. Although individualized therapy should be applied depending on the phase, no clinical or experimental studies have found biomarkers that distinguish between the 3 phases. Theiler’s murine encephalomyelitis virus belongs to the genus Cardiovirus and can cause myocarditis in susceptible mouse strains. Methods and Results Using this novel model for viral myocarditis induced with Theiler’s murine encephalomyelitis virus, we conducted multivariate analysis including echocardiography, serum troponin and viral RNA titration, and microarray to identify the biomarker candidates that can discriminate the 3 phases. Using C3H mice infected with Theiler’s murine encephalomyelitis virus on 4, 7, and 60 days post infection, we conducted bioinformatics analyses, including principal component analysis and k-means clustering of microarray data, because our traditional cardiac and serum assays, including 2-way comparison of microarray data, did not lead to the identification of a single biomarker. Principal component analysis separated heart samples clearly between the groups of 4, 7, and 60 days post infection. Representative genes contributing to the separation were as follows: 4 and 7 days post infection, innate immunity–related genes, such as Irf7 and Cxcl9; 7 and 60 days post infection, acquired immunity–related genes, such as Cd3g and H2-Aa; and cardiac remodeling–related genes, such as Mmp12 and Gpnmb. Conclusions Sets of molecules, not single molecules, identified by unsupervised principal component analysis, were found to be useful as phase-specific biomarkers.

Calpains, cathepsins and caspases play crucial role in mediating cell death. In the present study we observed a cascade of events involving the three proteases during middle cerebral artery occlusion (MCAo) in Wistar rats. The rats were MCA occluded and reperfused at various time points. We observed a maximal increase in the levels of calpains during 1h and 12 h after reperfusion than permanently occluded rats. Further, these levels were reduced by 1st and 3rd day of reperfusion. Similarly the cathepsin-b levels were significantly increased during 1h and 12 h, of reperfusion, followed by activation of caspase-3 which reached maximal levels by 1st and 3rd day of reperfusion. The sequential activation of calpains, cathepsin-b and cleaved caspase-3 is evident by the Western blot analysis which was further confirmed by the cleavage of substrates like PSD-95 and spectrin. The differences in the regional distribution and elevation of these proteases at different reperfusion time periods indicates that differential mode of cell death occur in the brain during cerebral ischemia in rat model.

BackgroundMultiple sclerosis (MS) is associated with ectopic lymphoid follicle formation. Podoplanin+ (lymphatic marker) T helper17 (Th17) cells and B cell aggregates have been implicated in the formation of tertiary lymphoid organs (TLOs) in MS and experimental autoimmune encephalitis (EAE). Since podoplanin expressed by Th17 cells in MS brains is also expressed by lymphatic endothelium, we investigated whether the pathophysiology of MS involves inductions of lymphatic proteins in the inflamed neurovasculature.MethodsWe assessed the protein levels of lymphatic vessel endothelial hyaluronan receptor and podoplanin, which are specific to the lymphatic system and prospero-homeobox protein-1, angiopoietin-2, vascular endothelial growth factor-D, vascular endothelial growth factor receptor-3, which are expressed by both lymphatic endothelium and neurons. Levels of these proteins were measured in postmortem brains and sera from MS patients, in the myelin proteolipid protein (PLP)-induced EAE and Theiler’s murine encephalomyelitis virus (TMEV) induced demyelinating disease (TMEV-IDD) mouse models and in cell culture models of inflamed neurovasculature.Results and conclusionsIntense staining for LYVE-1 was found in neurons of a subset of MS patients using immunohistochemical approaches. The lymphatic protein, podoplanin, was highly expressed in perivascular inflammatory lesions indicating signaling cross-talks between inflamed brain vasculature and lymphatic proteins in MS. The profiles of these proteins in MS patient sera discriminated between relapsing remitting MS from secondary progressive MS and normal patients. The in vivo findings were confirmed in the in vitro cell culture models of neuroinflammation.