SEMA4D compromises blood–brain barrier, activates microglia, and inhibits remyelination in neurodegenerative disease

AbstractMultiple sclerosis (MS) is a chronic neuroinflammatory disease characterized by immune cell infiltration of CNS, blood–brain barrier (BBB) breakdown, localized myelin destruction, and progressive neuronal degeneration. There exists a significant need to identify novel therapeutic targets and strategies that effectively and safely disrupt and even reverse disease pathophysiology. Signaling cascades initiated by semaphorin 4D (SEMA4D) induce glial activation, neuronal process collapse, inhibit migration and differentiation of oligodendrocyte precursor cells (OPCs), and disrupt endothelial tight junctions forming the BBB. To target SEMA4D, we generated a monoclonal antibody that recognizes mouse, rat, monkey and human SEMA4D with high affinity and blocks interaction between SEMA4D and its cognate receptors. In vitro, anti-SEMA4D reverses the inhibitory effects of recombinant SEMA4D on OPC survival and differentiation. In vivo, anti-SEMA4D significantly attenuates experimental autoimmune encephalomyelitis in multiple rodent models by preserving BBB integrity and axonal myelination and can be shown to promote migration of OPC to the site of lesions and improve myelin status following chemically-induced demyelination. Our study underscores SEMA4D as a key factor in CNS disease and supports the further development of antibody-based inhibition of SEMA4D as a novel therapeutic strategy for MS and other neurologic diseases with evidence of demyelination and/or compromise to the neurovascular unit

Ion currents and proliferation in MCF-7 human breast cancer cells.

Previous work in our laboratory has demonstrated that in order to proliferate MCF-7 human breast cancer cells have to undergo a change of the membrane potential from a very depolarized to a more hyperpolarized. That change occurs at a hypothetical regulatory point located in early G1 phase, D control point. Potassium (K) channel antagonists quinidine, linogliride, glibenclamide, 4-aminopyridine (4-AP) and tetraethylammonium (TEA) inhibited MCF-7 cell proliferation. Among those blockers three (quinidine, linogliride and glibenclamide) also produced a reversible G0/G1 arrest. On the basis that these three drugs are antagonists of ATP-sensitive K (KA{dollar}\\sb{lcub}\\rm ATP{rcub}{dollar}) channels, we concluded that in MCF-7 cells it is the activation of K{dollar}\\sb{lcub}\\rm ATP{rcub}{dollar} channels at the D control point that promotes progression through G1 phase. The goal of our work was to identify in MCF-7 cells ion current(s) whose activation is required for passage through early G1 phase. To achieve this goal we used whole-cell and single-channel configurations of patch-clamp technique. We expected that the ion current-regulator of the G1 progression would: (1) have a reversal (i.e., zero current) potential near the equilibrium potential for K{dollar}\\sp+{dollar} ({dollar}-{dollar}84 mV); (2) be blocked by physiological concentrations of intracellular ATP; (3) be blocked by linogliride, glibenclamide and quinidine, but not by TEA. Our study demonstrated that MCF-7 cells contain ATP-sensitive K{dollar}\\sp+{dollar} current with the pharmacological profile of the regulator of G1 progression. The conductance density of the current was very low in cells arrested in early G1 phase (prior to D control point) with quinidine that further confirmed the involvement into G1 progression. We also identified in MCF-7 cells a small-conductance (8.5 pS) channel whose reversal potential was close to the equilibrium potential for K{dollar}\\sp+.{dollar} Furthermore, the channel was inhibited by quinidine. We concluded that this channel underlies the current-regulator of G1 progression