Combination Therapy With VELCADE and Tissue Plasminogen Activator Is Neuroprotective in Aged Rats After Stroke and Targets MicroRNA-146a and the Toll-Like Receptor Signaling Pathway

Abstract

Objective Activation of the toll-like receptor (TLR) signaling pathway exacerbates ischemic brain damage. The present study tested the hypothesis that combination treatment with VELCADE and tissue plasminogen activator (tPA) modulates the TLR signaling pathway on cerebral vasculature, which leads to neuroprotection in aged rats after stroke. Methods and Results Focal cerebral ischemia acutely increased TLR2, TLR4, and interleukin-1 receptor activated kinases 1 (IRAK1) immunoreactivity on fibrin/fibrinogen positive vessels in aged rats. Monotherapy of tPA further amplified these signals. However, VELCADE in combination with tPA blocked stroke- and tPA-potentiated vascular TLR signals, leading to robust reduction of infarct volume compared with respective monotherapies. Quantitative RT-PCR analysis of cerebral endothelial cells isolated by laser capture microdissection revealed that the combination treatment increased miR-l46a levels, which was inversely associated with the reduction of vascular IRAK1 immunoreactivity. In vitro, fibrin upregulated IRAK1 and TLR4 expression and downregulated miR-146a on primary human cerebral endothelial cells. VELCADE elevated miR-146 levels and abolished fibrin-increased IRAK1 proteins. Conclusions Stroke acutely activates the TLR signaling pathway on cerebral vasculature. Upregulation of miR-146a and inactivation of ischemia and tPA-potentiated TLR signaling pathway by VELCADE may play an important role in the neuroprotective effect of the combination therapy of VELCADE and tPA for acute stroke.

Brain Endothelial miR-146a Negatively Modulates T-Cell Adhesion through Repressing Multiple Targets to Inhibit NF-κB Activation

“…15 Unexpectedly, we also observed that siIRAK1 and/or siTRAF6 decreased VCAM1 protein levels without affecting its corresponding mRNA levels. It is possible that the role of these two signaling proteins in the inflammatory activation of brain endothelial cells induced by cytokines is a complex process and that, in addition to their role in NF-κB-mediated transcriptional activation, IRAK1, and TRAF6 may regulate inflammatory molecules such as VCAM1 at the posttranslational level.…”

“…11 The anti-inflammatory role of miR-146a has been demonstrated in several cell types including monocytes, 10 T cells, 12 astrocytes, 13 human umbilical vein endothelial cells (HUVECs), 14 and human BECs. 15 However, its function in BECs remains to be elucidated. Here we show that miR-146a is upregulated in cytokine-activated BECs and decreases leukocyte adhesion by inhibiting NF-κB through repressing not only TRAF6 and IRAK1 but also RhoA and nuclear factor of activated T cells 5 (NFAT5).…”

Non-coding RNAs and neuroprotection after acute CNS injuries

“…1). A combination therapy of VELCADE and tissue plasminogen activator was found to provide neuroprotection in aged rats after stroke by simultaneously upregulating miR-146a and inactivating toll-like receptor signaling pathway (Zhang et al, 2012a) (Fig. 1).…”

Angiogenesis-regulating microRNAs and Ischemic Stroke

“…Several other studies from different groups also documented that direct modulation of miR-23a [183], miR-181 [184], miR-29b [185], and Let7f [186] can provide neuroprotective roles in rodent experimental stroke models. Accordingly, miRs such as miR-331 and miR-885-3p [187], miR-146a [188], and miR-199a-5p [189] can also mediate valproic acid, VELCADE-tissue plasminogen activator combination therapy, Vitamin E-mediated neuroprotection or improvements in neurological deficits, and motor performance in rodent stroke models. Interestingly, stroke is also able to alter miR expression in neural progenitor cells of the subventricular zone (SVZ) and in particular, miR-124a mediates stroke-induced neurogenesis by targeting the JAG-Notch signaling pathway [190].…”