Role of P38 MAPK on MMP Activity in Photothrombotic Stroke Mice as Measured using an Ultrafast MMP Activatable Probe

Abstract

Matrix metalloproteinases (MMPs) exert a dual effect in ischemic stroke and thus represent an ideal target for detection and therapy. However, to date, all clinical trials of MMP inhibitors have failed, and alternative drug candidates and therapeutic targets are urgently required. Nonetheless, further investigations are limited by the lack of non-invasive imaging techniques. Here, we report a novel, fast and ultrasensitive MMP activatable optical imaging probe for the dynamic visualization of MMP activity in photothrombotic stroke mice. This probe provides a significant signal enhancement in as little as 15 min, with the highest signal intensity occurring at 1 h post-injection, and shows high sensitivity in measuring MMP activity alterations, which makes it specifically suitable for the real-time visualization of MMP activity and drug discovery in preclinical research. Moreover, using this probe, we successfully demonstrate that the regulation of the p38 mitogen-activated protein kinase (MAPK) signal pathway is capable of modulating MMP activity after stroke, revealing a novel regulatory mechanism of postischemic brain damage and overcoming the limitations of traditional therapeutic strategies associated with MMP inhibitors by using a non-invasive molecular imaging method.

Continuous administration of the p38α inhibitor neflamapimod during the subacute phase after transient ischemia-induced stroke in the rat promotes dose-dependent functional recovery accompanied by increase in brain BDNF protein level

“…This neflamapimod treatment study is clearly different from all previously reported studies of other small molecule p38 MAPK inhibitors in various experimental stroke models [25,[28][29][30][31][32] both in terms of the delay in treatment initiation to 48 hours post reperfusion and the six-week prolonged dosing duration and overall treatment study focus. All previously reported studies of p38 MAPK inhibitors that most potently inhibit the p38 isoform, including SB203580 [25], SB239063 [28,29], RWJ67657 [31], or VCP979 [32], involved compound administration before or soon after the experimental stroke during the acute phase, with treatment initiations at different time points, ranging from as early as 1 hour pre stroke to as late as 24 hours post stroke. Additionally, all prior p38 inhibitor studies had a focus on demonstrating neuroprotective effects that resulted in significant reduction of infarct volume (assessed as early as 6 hours post stroke) and subsequent improvements of neurological defects assessed at 24 hours [28,29], 2 days [25], 7 days [32] or 14 days post stroke [31].…”

“…Activation of p38 MAPK, including p38, signaling after experimental ischemic stroke in rodents has been demonstrated in neurons, astrocytes and microglia [21][22][23][24][25], and p38 has been established as a driver of neuroinflammation-mediated cell death in the acute phase of ischemic stroke [26,27]. Therefore, several inhibitors of p38 MAPK that exhibit different potency and kinase selectivity, all of them most potently blocking p38 versus other p38 isoforms (p38, p38, p38) have been administered during the acute phase with treatment initiation within the first 24 hours after stroke in experimental models of cerebral ischemia, and all of them have provided robust neuroprotection [25,[28][29][30][31][32].…”

“…Time course analyses of phospho-p38 (i.e. the activated from of p38) expression in experimental rat stroke models reported increased phospo-p38, including phospho-p38, immediately after cerebral ischemia within neurons and other cell types, and, despite some fluctuations observed in some of the studies, also demonstrated that phospho-p38 remained elevated and associated with neurons, as well as other cell types, for >4 days, and in one study up to 14 days, implying that p38-activation during the subacute phase could be partly responsible for poor recovery from stroke [22,24,25,31]. A one week study of a novel p38 inhibitor, the tetra-substituted thiophene VCP979, evaluated as a treatment of phothothrombotic ischemic stroke in streptozocin-induced Type 2 diabetes mellitus (T2DM) mice demonstrated beneficial efficacy resulting in neuroprotection as well as axonal/white matter PAGE 6 remodeling within the motor cortex [32].…”

“…All previously reported studies of p38 MAPK inhibitors that most potently inhibit the p38 isoform, including SB203580 [25], SB239063 [28,29], RWJ67657 [31], or VCP979 [32], involved compound administration before or soon after the experimental stroke during the acute phase, with treatment initiations at different time points, ranging from as early as 1 hour pre stroke to as late as 24 hours post stroke. Additionally, all prior p38 inhibitor studies had a focus on demonstrating neuroprotective effects that resulted in significant reduction of infarct volume (assessed as early as 6 hours post stroke) and subsequent improvements of neurological defects assessed at 24 hours [28,29], 2 days [25], 7 days [32] or 14 days post stroke [31]. The two most recent p38 inhibitor studies [31,32] were also the two longest ones prior to the present study.…”

“…Additionally, all prior p38 inhibitor studies had a focus on demonstrating neuroprotective effects that resulted in significant reduction of infarct volume (assessed as early as 6 hours post stroke) and subsequent improvements of neurological defects assessed at 24 hours [28,29], 2 days [25], 7 days [32] or 14 days post stroke [31]. The two most recent p38 inhibitor studies [31,32] were also the two longest ones prior to the present study. Interestingly, they did demonstrate the validity of mNSS [51] as a readout for functional recovery attributable to neuroprotection via p38 inhibition, that was measurable during the subacute phase after photothrombotic ischemic stroke in mice at one and two weeks, respectively [31,32].…”