1 INTRODUCTION
Sudden cardiac arrest (CA) is a leading cause of global mortality
[1]. Despite much advances in optimizing the techniques of
cardiopulmonary resuscitation (CPR), the overall prognosis is still
unsatisfactory after out-of-hospital CA with successful resuscitation,
mainly due to the post-CA syndrome [2, 3]. Brain injury represents
an essential hallmark of the pathophysiology of the post-CA syndrome
[4],
profoundly
impairs the neurological function and conduces to lifelong disability
among CA survivors and even death. However, no clinically
effective
pharmacological intervention is available to reduce neurologic
deficiency in patients with CA/CPR at present [5].
Currently, most neuroprotective methods after CA/CPR have focused on
protecting neurons. However, endothelial dysfunction, an inevitable
pathologic process of ischemia/reperfusion during CA plays a governing
role in the progression of post-CA brain injury and evokes further
neuronal damages [6]. The blood-brain barrier (BBB) integrity is
compromised by CA/CPR, leading to intractable cerebral edema. More
significantly, cerebral edema exacerbates clinical outcomes and becomes
a
potent
prognostication after CA [7-9]. Microglia/macrophages are
spectacularly plastic and obtain multiple subtypes, such as
pro-inflammatory and anti-inflammatory status, to fulfill different
activities in health and disease. Altered microglia/macrophages
functional subtypes and consequent neuroinflammation have an intimate
relationship with the integrity of BBB. The unwanted entry of serum
proteins into brain after the collapse of BBB is a pivotal cause of
neuroinflammation [10], facilitating the transition of
microglia/macrophages to a pro-inflammatory status. The pro-inflammatory
phenotype of microglia, in turn, can simultaneously exaggerate BBB
insult [11]. In this way, a vicious circle between BBB breakdown and
pro-inflammatory microglia/macrophages status is created to escalate
post-CA/CPR brain injury continuously. Therefore, only drugs that
modulate multiple targets such as BBB destruction, neuroinflammation and
neuronal injury are likely to achieve clinical translation.
Although the molecular mechanisms responsible for controlling the
restrictive feature of BBB remain
largely
elusive, transient receptor potential M4 (TRPM4), a nonselective
monovalent cation channel activated by elevated intracellular
Ca2+, has been shown to be critical in regulating the
BBB function [12]. Studies conducted by us and others indicated that
pharmacological inhibition of the subunits of sulfonylurea receptor
1-transient receptor potential M4 (SUR1-TRPM4) or gene deletion ofTrpm4 could function in preserving the BBB integrity in animal
models of status epilepticus [13], acute ischemic stroke [14],
spinal cord injury [15], intracerebral hemorrhage [16], and
subarachnoid hemorrhage [17]. Indeed, we have reported that
glibenclamide, a selective inhibitor of SUR1, effectively improved
survival and neurologic outcome in rodent models of CA/CPR [18-20],
but its protective effect on BBB after CA/CPR and whether it works by
blocking the SUR1-TRPM4 channel remain unclear. Furthermore, the
SUR1-TRPM4 channel was shown to be expressed in microglia and
participated in regulating pro-inflammatory gene expression [21],
implying that TRPM4 channel may affect microglia polarization.
Therefore, the therapeutic manipulation of the BBB function and
phenotypic shift of microglia/macrophages by interfering with the TRPM4
channel may serve as a promising opportunity for minimizing brain injury
resulting from CA/CPR.
Flufenamic acid (FFA) is a non-steroidal anti-inflammatory drug that has
been applied for analgesia against pain related to rheumatic disorders.
Since the TRPM4 channel was found to be highly sensitive to FFA and can
be inhibited by low concentrations of FFA, FFA has attracted extensive
attention as a convenient and relatively selective TRPM4 inhibitor to
study the physiological effects of TRPM4 [22, 23]. Emerging studies
suggested that FFA conferred neuroprotection against several
neurological diseases, such
as
spinal cord injury [24, 25], Alzheimer’s disease [26], and
epilepsy [27]. FFA inhibited capillary fragmentation and secondary
hemorrhage via blocking TRPM4 after spinal cord injury [24, 25].
These studies prompted us to logically postulate that FFA could
elicit
its neuroprotective effects through ameliorating BBB breakdown and
improving neurologic outcome by inhibiting TRPM4.
Here, we investigated whether FFA could improve neurologic outcome in a
mouse model of CA/CPR. Moreover, we aimed to explore whether gene
deletion of Trpm4 (Trpm4−/− ) exerts
similar effect to FFA and whether blockage of TRPM4 represents part of
the mechanism accounting for FFA-mediated neuroprotection in CA/CPR.