sb-290157 and Stroke

The complement system is a key regulator of the innate immune response against diseased tissue that functions across multiple organ systems. Dysregulation of complement contributes to the pathogenesis of a number of neurological diseases including stroke. The C3a anaphylatoxin, via its cognate C3a receptor (C3aR), mediates inflammation by promoting breakdown of the blood-brain barrier and the massive infiltration of leukocytes into ischemic brain in experimental stroke models. Studies utilizing complement deficient mice as well as pharmacologic C3aR antagonists have shown a reduction in tissue injury and mortality in murine stroke models. The development of tissue-specific C3aR knockout mice and more specific C3aR antagonists is warranted to facilitate our understanding of the role of the C3aR in brain ischemia with the ultimate goal of clinical translation of therapies targeting C3aR in stroke patients.

Topics: Animals; Arginine; Benzhydryl Compounds; Blood-Brain Barrier; Complement Activation; Complement C3a; Complement Inactivating Agents; Disease Models, Animal; Humans; Immunity, Innate; Mice; Mice, Knockout; Neuroimmunomodulation; Neuroprotective Agents; Receptors, Complement; Stroke; Translational Research, Biomedical

Topics: Animals; Arginine; Benzhydryl Compounds; Mice; Stroke; Thrombolytic Therapy

Worldwide, millions of individuals suffer an ischemic stroke each year, causing major disability, especially in the elderly, where stroke is the number one cause of disability. However, to date, no effective therapy exists that targets the functional recovery after stroke. After necrosis, neuroinflammation is a common feature of the acute stroke and a major obstacle to tissue restoration. In the lesioned area, the dying neurons release chemotactic signals, such as fractalkine/CX3CL1, which evoke "eat-me" signals that are recognized by microglia expressing complement C3a receptor (C3aR), resulting in phagocytosis of the dying but still viable neurons, known as secondary phagocytosis. Using a mouse model of stroke and two-photon microscopy, we aimed to attenuate poststroke phagocytosis of the dying but still viable neurons by using SB 290157, an antagonist of C3aR. We found that intracortical administration of SB 290157 reduced the number of inflammatory microglial cells expressing ED1 and Iba1 antigens at the lesion site. We could show, in vivo, that two days after a needle-induced cortical lesion there were less microglial cells present around the injury site, displaying less high-order branches and an increase in the lower order ones, suggesting an attenuated phagocytic phenotype in treated animals as compared with controls. We conclude that the C3aR antagonist, SB 290157, may be used in the future to limit the neuronal death by limiting secondary phagocytosis after stroke.

Topics: Animals; Arginine; Benzhydryl Compounds; Disease Models, Animal; Mice; Microglia; Neurons; Phagocytosis; Receptors, Complement; Recovery of Function; Stroke; Trifluoroacetic Acid

The use of intravenous tissue plasminogen activator (tPA) in the treatment of ischemic stroke is limited by its propensity to exacerbate brain edema and hemorrhage. The mechanisms underlying these deleterious effects of tPA remain incompletely understood. The purpose of this study was to delineate a pathway of tPA-mediated complement cascade activation in stroke and to determine whether complement inhibition ameliorates the adverse effects of post-ischemic tPA administration. We found that tPA promotes C3 cleavage both in vitro and in ischemic brain through a plasmin-mediated extrinsic pathway. Using cell culture models, we then showed that the C3a-receptor is strongly expressed on ischemic endothelium and that exogenous C3a dramatically enhances endothelial cell permeability. Next, we assessed the effect of tPA administration on brain edema and hemorrhage in a transient model of focal cerebral ischemia in C57BL/6 mice. We found that intravenous tPA exacerbates brain edema and hemorrhage in stroke, and that these effects are abrogated by a small-molecule antagonist of the C3a receptor. These findings establish for the first time that intravenous tPA dramatically upregulates complement cascade activation in ischemic brain and that pharmacologic complement inhibition protects against the adverse effects of tPA-mediated thrombolysis in stroke.

Topics: Animals; Arginine; Benzhydryl Compounds; Brain; Brain Edema; Cell Death; Cells, Cultured; Complement C3; Endothelial Cells; Fibrinolysin; Hemoglobins; Hemorrhage; Humans; Immunohistochemistry; Infarction, Middle Cerebral Artery; Male; Mice; Mice, Inbred C57BL; Stroke; Tissue Plasminogen Activator

Vascular smooth muscle cells (VSMCs) derived from spontaneously hypertensive rats (SHR) show exaggerated growth with a synthetic phenotype and angiotensin II (Ang II) production associated with increased production of complement (C3). We hypothesized that C3 is involved in the growth of mesangial cells (MCs) from hypertensive rats. We examined the effects of a C3a receptor inhibitor on proliferation, phenotype and Ang II generation in MCs from stroke prone-spontaneously hypertensive rats (SHR)-SP, SHR and Wistar-Kyoto (WKY) rats. Expression of C3 and C3a receptor were evaluated by immunohistochemical staining of the renal cortex. We examined the effects of the C3a inhibitor, SB290157, on proliferation, the expression of phenotype-marker mRNAs and Ang II production in cells from SHR-SP, SHR and WKY rats. Immunostaining of C3 was stronger in SHR and SHRSP glomeruli. MCs from SHR-SP and SHR abundantly express pre-pro C3 mRNA. SB290157 significantly inhibited basal DNA synthesis and proliferation of MCs from SHR-SP and SHR. Expression of osteopontin mRNA in MCs from SHR-SP and SHR was decreased with SB290157 treatment, whereas MC basal expression of α-SMA mRNA was decreased. SB290157 significantly decreased the production of Ang II in MCs from SHR-SP and SHR. Endogenous C3a promotes exaggerated growth with a synthetic phenotype and the production of Ang II in MCs from SHR-SP and SHR. The C3 and C3a receptor system may primarily be involved in the pathogenesis of renal remodeling in hypertensive rats.

Topics: Actins; Angiotensin II; Animals; Arginine; Benzhydryl Compounds; Cell Proliferation; Complement C3a; Gene Expression; Hypertension; Kidney Cortex; Male; Mesangial Cells; Myocytes, Smooth Muscle; Osteopontin; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Receptors, Complement; RNA, Messenger; Stroke