Investigating the effect of corneal herpes simplex virus infection on toll like receptor expression in human peripheral blood mononuclear cells

Herpes Simples Keratitis (HSK) is the commonest cause of infectious blindness in the developed world. It is caused by HSV-l which is a large double stranded DNA virus which can invade the cornea and after treatment remains latent in the trigeminal ganglia. Toll like receptors are key components of the innate immune system and are highly expressed in the corneal epithelium. HSV-l induces upregulation of several Toll Like Receptors (TLRs) and triggers the release of anti-viral cytokines. In certain cases HSV-l has evolved to avoid these innate anti-viral responses and can cause lifelong recurrent infection. This recurrent keratitis causes lesions which are immunoinflammatory in nature, can recur throughout life and cause progressive corneal scarring, vascularisation, thinning and may require a corneal transplant which does not have a good long-term outcome. Understanding the mechanisms that cause this disease may lead to improved or novel therapies that will help the long-term outcome of HSK patients. This study aims to examine the immune responses of HSK patient peripheral blood mononuclear cells (PBMC) before and after treatment compared to healthy donors. Our work has shown that peripheral blood mononuclear cells (PBMCs) show differences in their ability to respond to various TLR ligands both in relation to the cytokines they produce and upregulation of cell surface markers. In particular our results show that IL-1(3, a key proinflammatory cytokine is elevated in serum of active patients compared to inactive patients, thus demonstrating that peripheral immune responses are activated in response to HSV infection of the cornea. In addition our work has shown that although active patients expressed activation markers on T cells and B cells but differences were inconclusive based on patient numbers (active/inactive n=5, control n=4). Further data indicated that the TLR 3 pathway is compromised in active patient PBMCs, potentially having implications for HSV-l viral clearance and viral replication and HSK persists in the cornea

Platelet signalling networks: pathways perturbation demonstrates differential sensitivity of ADP secretion and fibinogen binding.

Platelet signalling responses to single agonists have been identified previously. However a model of the total platelet signalling network is still lacking. In order to gain insights into this network, we explored the effects of a range of platelet-function inhibitors in two independent assays of platelet function, namely fibrinogen binding and ADP secretion. In this study, we targeted the intracellular signalling molecules Syk and PI3K, the prostaglandin synthesis enzyme COX, surface receptors for TxA2 and ADP (P2Y1 and P2Y12) and the integrin cell adhesion molecule, aIIbb3. We demonstrate that the platelet responses of fibrinogen binding and secretion can be differentially affected by the individual inhibitors permitting the generation of a model delineating novel regulatory links in the platelet signal network. Importantly, the model illustrates the interconnections among portions that are traditionally studied as separate modules, promoting a more integrated view of the platelet

The formation of microthrombi in parenchymal microvessels after traumatic brain injury is independent of coagulation factor XI

Microthrombus formation and bleeding worsen the outcome after traumatic brain injury (TBI). The aim of the current study was to characterize these processes in the brain parenchyma after experimental TBI and to determine the involvement of coagulation factor XI (FXI). C57BL/6 mice (n = 101) and FXI-deficient mice (n = 15) were subjected to controlled cortical impact (CCI). Wild-type mice received an inhibitory antibody against FXI (14E11) or control immunoglobulin G 24 h before or 30 or 120 min after CCI. Cerebral microcirculation was visualized in vivo by 2-photon microscopy 2-3 h post-trauma and histopathological outcome was assessed after 24 h. TBI induced hemorrhage and microthrombus formation in the brain parenchyma (p < 0.001). Inhibition of FXI activation or FXI deficiency did not reduce cerebral thrombogenesis, lesion volume, or hemispheric swelling. However, it also did not increase intracranial hemorrhage. Formation of microthrombosis in the brain parenchyma after TBI is independent of the intrinsic coagulation cascade since it was not reduced by inhibition of FXI. However, since targeting FXI has well-established antithrombotic effects in humans and experimental animals, inhibition of FXI could represent a reasonable strategy for the prevention of deep venous thrombosis in immobilized patients with TBI

Systemic IL-1β production as a consequence of corneal HSV-1 infection-contribution to the development of herpes simplex keratitis

This study sought to identify potential therapeutic targets in herpes simplex keratitis (HSK) patients with active and inactive infection by investigating peripheral cytokine production. Peripheral blood mononuclear cells (PBMCs) and serum were prepared from healthy controls and HSK patients during active infection or following treatment (inactive infection). Serum antibody titres were determined by ELISA. Protein expression levels were analysed by Western blot. Cytokine levels were determined by multiplex ELISA. Active corneal herpes simplex virus type 1 (HSV-1) infection resulted in significantly elevated peripheral levels of IL-1β in HSK patients compared to healthy controls, and remained significantly increased following treatment. Elevated production of IL-1β in inactive patients was associated with significantly increased levels of IRF3 and STAT1, key proteins involved in promoting anti-viral immune responses. Our data suggest that inflammation persists beyond the period that it is clinically evident and that enhanced peripheral production of IL-1β may have implications for HSV-1 viral clearance in active and inactive HSK patients

Low expression of pro-apoptotic proteins Bax, Bak and Smac indicates prolonged progression-free survival in chemotherapy-treated metastatic melanoma

Despite the introduction of novel targeted therapies, chemotherapy still remains the primary treatment for metastatic melanoma in poorly funded healthcare environments or in case of disease relapse, with no reliable molecular markers for progression-free survival (PFS) available. As chemotherapy primarily eliminates cancer cells by apoptosis, we here evaluated if the expression of key apoptosis regulators (Bax, Bak, Bcl-2, Bcl-xL, Smac, Procaspase-9, Apaf-1, Procaspase-3 and XIAP) allows prognosticating PFS in stage III/IV melanoma patients. Following antibody validation, marker expression was determined by automated and manual scoring of immunohistochemically stained tissue microarrays (TMAs) constructed from treatment-naive metastatic melanoma biopsies. Interestingly and counter-intuitively, low expression of the pro-apoptotic proteins Bax, Bak and Smac indicated better prognosis (log-rank p  12 months) on a case-by-case basis (area under the receiver operating characteristic curve (ROC AUC) = 0.79). Taken together, our results therefore suggest that Bax, Bak and Smac jointly define a signature with potential clinical utility in chemotherapy-treated metastatic melanoma.status: publishe

Low expression of pro-apoptotic proteins Bax, Bak and Smac indicates prolonged progression-free survival in chemotherapy-treated metastatic melanoma.

Despite the introduction of novel targeted therapies, chemotherapy still remains the primary treatment for metastatic melanoma in poorly funded healthcare environments or in case of disease relapse, with no reliable molecular markers for progression-free survival (PFS) available. As chemotherapy primarily eliminates cancer cells by apoptosis, we here evaluated if the expression of key apoptosis regulators (Bax, Bak, Bcl-2, Bcl-xL, Smac, Procaspase-9, Apaf-1, Procaspase-3 and XIAP) allows prognosticating PFS in stage III/IV melanoma patients. Following antibody validation, marker expression was determined by automated and manual scoring of immunohistochemically stained tissue microarrays (TMAs) constructed from treatment-naive metastatic melanoma biopsies. Interestingly and counter-intuitively, low expression of the pro-apoptotic proteins Bax, Bak and Smac indicated better prognosis (log-rank p 12 months) on a case-by-case basis (area under the receiver operating characteristic curve (ROC AUC) = 0.79). Taken together, our results therefore suggest that Bax, Bak and Smac jointly define a signature with potential clinical utility in chemotherapy-treated metastatic melanoma