Amyotrophic lateral sclerosis (ALS) is a motor neuron disease characterized by progressive distal axonopathy that precedes actual motor neuron death. Triggers for neuromuscular junction degeneration remain to be determined, but the axon repulsion factor semaphorin 3A (Sema3A), which is derived from terminal Schwann cells, is a plausible candidate. This study examines the hypothesis that Sema3A signaling through its motor neuron neuropilin-1 (NRP1) receptor triggers distal axonopathy and muscle denervation in the SOD1G93A mouse model of ALS. Neuropilin-1 was found to be expressed in axonal terminals at the mouse neuromuscular junction in vivo and in NSC-34 motor neuron–like cells in vitro. In differentiated NSC-34 cells, an anti-NRP1A antibody that selectively blocks Sema3A binding to NRP1 prevented Sema3A-induced growth cone collapse. Furthermore, intraperitoneal injections of anti-NRP1A antibody administered twice weekly from age 40 days significantly delayed and even temporarily reversed motor functional decline while prolonging the life span of SOD1G93A mice. Histologic evaluation at 90 and 125 days revealed that anti-NRP1A antibody reduced neuromuscular junction denervation and attenuated pathologic alterations in ventral roots at late-stage disease. These data suggest that peripheral NRP1A signaling is involved in the pathobiology of this ALS model and that antagonizing Sema3A/NRP1 binding or downstream signals could have implications for the treatment of ALS.

Serpins maintain haemostasis through regulation of serine proteinases in the thrombotic and thrombolytic pathways. Viruses encode serpins that can alter thrombotic and thrombolytic responses producing, in some cases, disseminated intravascular coagulation (DIC). However, it has not been precisely defined how viral serpins induce these profound responses. The rabbit myxoma viral serpin, Serp-1 inhibits urokinase- and tissue-type plasminogen activators (uPA and tPA), plasmin and factor Xa in vitro and exhibits remarkable anti-inflammatory activity in various animal models. The effects of Serp-1 on activation of human platelets, endothelial cells, monocytes and T cells that mediate thrombosis and innate immune responses were therefore examined. We found that Serp-1 attenuated platelet and mononuclear cell adhesion to fibronectin and collagen. Serp-1 similarly inhibited monocyte migration into the peritoneum. Serp-1 inhibition of monocyte migration was lost in uPA receptor (uPAR) deficient mice. Serp-1 bound to the plasma membrane surface and altered uPA activation of endothelial cells (p=0.001), thrombin activation of platelets (p=0.021) and phorbol ester activation of endothelial (p=0.047), monocyte (p=0.011) and Jurkat T cells (p=0.012) as measured by intracellular calcium. Modulation of cellular activation was confirmed by membrane fluidity analysis. Microarray analysis of Serp-1 treated endothelial cells revealed alterations in Inositol 1,4,5-triphosphate receptor type II (ITPR2) a calcium-regulating gene. This study demonstrates the unique capacity of a viral serpin, Serp-1 to modify adhesion, activation, gene expression and calcium homeostasis in a wide range of cells that regulate coagulation and inflammation. Endothelial cells potentially represent a pivotal regulatory point for Serp-1 anti-inflammatory activity.

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Brain microvascular alterations are thought to contribute to the development of stroke and dementia. Structural changes in capillaries of elderly patients correlate positively with advanced age and dementia. The objective of this study is to use laser-induced fluorescence spectroscopy to compare structural (collagen content) and functional (apoptosis) parameters in brain tissues and isolated vessels of AD patients to age-matched controls. Our results show significantly higher fluorescent labeling for apoptosis in AD vessels compared to controls. Also, there is significantly higher autofluorescence (reflecting levels of collagen and other proteins that autofluoresce) in AD brain and vessels compared to controls. Western blot analysis of collagen subtypes shows elevated type I and type III and reduced type IV levels in AD vessels. These data demonstrate that changes in the amount and type of collagen occur in AD brain and suggest that cerebral vessel injury is part of AD pathology.