Discussion
Tau pathology is reported to be a biomarker for cognitive impairment
involving cerebral ischemia and neuroinflammation in the early and late
stages of AD (Ballatore et al., 2007; Freude et al., 2005; Iqbal et al.,
2005; Kim et al., 2009; Mandelkow et al., 1996). Children less than five
years old infected with CM exhibited axonal injury caused by increased
plasma tau levels resulting in cognitive impairment and severe mortality
(Datta et al., 2021; Medana et al., 2007). A recent study by Oscar B et
al. states that dysregulation of tau protein contributes to cerebral
vasculopathy and neuronal cell injury impairing cognitive outcomes
during CM (Akide Ndunge et al., 2023). According to the Minxian Dai et
al., aberrant phosphorylation of tau increased the disease severity by
inducing conformational changes in neuronal proteins in experimental
models of CM (Dai et al., 2012b). C57BL/6 mice infected withPlasmodium berghei ANKA is widely known to reiterate the symptoms
of human cerebral malaria. Despite several transgenic models for tau,
ECM pathology globally activates tau without transgenic tau approach.
Tau hyperphosphorylation is propelled by several kinases including cdk5,
glycogen synthase kinase 3 (GSK3β) and mitogen-activated protein kinase
(MAPK) (Hatch et al., 2017). Previously, one of our recent studies
showed a dysregulation of cdk5 signaling, imbalance between kinases
(Ca2+/calmodulin-dependent protein kinase II alpha,
CaMKIIα, protein kinase A, cdk5) and phosphatases (protein phosphatase-1
gamma subunit 1, PP-1γ1) in ECM. Based on our previous studies and
several reports on vasculopathy based tau hyperphosphorylation, we
selected the animal model for CM as an effective model for this study.
Sequestration of infected RBCs cause microvasculature damage inducing
chronic hypoxia mediated elevation of VEGF (Vascular Endothelial Growth
Factor) which results in expression of calcium-dependent proteases,
calpain (Rénia et al., 2012). Chronic hypoxic conditions in the brain
are detected in the form of elevated Hypoxia-inducible factor (HIF)-1α
levels in terminally ill CM animals and human cerebral malaria (Hempel
et al., 2014). Low oxygen levels trigger activation of calpain and
several kinases such as mitogen-activated protein kinases (MAPKs) and
extracellular signal regulated kinases (ERK1/2) activating brain-derived
neurotrophic factor (BDNF) (Gao et al., 2013; Raz et al., 2019; Terraneo
and Samaja, 2017). Finally, all the above conditions lead to the
imbalance of kinase and phosphatases resulting in tau phosphorylation.
We assume that altered vasculature, reduced blood flow and chronic
hypoxia upregulate cdk5, one of the key factors in phosphorylating tau
at several sites such as Ser202, Ser396/404, Thr181and Thr231 (Noble et
al., 2003) in CM. Tau phosphorylation at Ser396 is considered as the
earliest event in Alzheimer’s disease in which cdk5 has been implicated
as potential candidate kinase (Mondragón-Rodríguez et al., 2014; Noble
et al., 2003). Similarly, cdk5/p25 signaling is detected early in
neurodegenerative diseases like Parkinson’s (Ao et al., 2022),
amylotrophic lateral sclerosis (ALS) (Bk et al., 2019), brain injuries
like subarachnoid haemorrhage (Ding et al., 2022), anxiety, depression
(Takahashi et al., 2022), learning disabilities involving cognitive
impairment (Kamiki et al., 2018) and cancers like glioblastoma (GBM)
(Peyressatre et al., 2020). Several research studies have shown that
inhibition of hyperactivated cdk5 based signaling improved hippocampal
neurogenesis and restored cognitive functions in radiation induced
cognitive dysfunction (Zhang et al., 2021). Cdk5 inhibition alleviated
diabetic neurotoxicity related cognitive deficits in diabetes mellitus
animal model (Liu et al., 2019). Drugs such as rolipram (Wachtel, 1983),
cilostazol (Schaler and Myeku, 2018), sildenafil (Sanders, 2020), CM-414
(Cuadrado-Tejedor et al., 2017) reduce tau hyperphosphorylation and
increased dendritic spine density in hippocampal neurons improving
cognition in animal models. Interestingly, recent reports state that
D-pinitol, a naturally occurring inositol alleviate hyperactivation of
p25/cdk5 activity by regulating cyclin‐dependent kinase 5 regulatory
subunit 1 (CDK5R1) and GSK-3 beta (tau phosphorylation regulating gene)
leading to tau dephosphorylation (Medina-Vera et al., 2022). According
to Jeremy Koppel et al., increased extracellular dopamine levels
hyperphosphorylate tau and blocking dopamine D2 receptor
results in robust decrease in tau phosphorylation (Koppel et al., 2019).
Dopaminergic receptors can modulate dopamine and cyclic adenosine
monophosphate regulated phospho protein (DARPP-32), which acts as
molecular switch to activate or inhibit kinases (cAMP-dependent protein
kinase) by activating protein phosphatase-1. We assume that there is a
current necessity to identify the earliest signaling mechanism that
trigger tau hyperphosphorylation and drugs that restore the balance
between kinases and phosphatases in tauopathies. Our study does not
determine phosphatase levels but further research has to be performed
whether polycyclitols restore balance between kinase and phosphatase
levels in ECM.
Till date, there has been no study conducted on the effect of
polycyclitols on cognition and behavior. To our knowledge, our findings
represent the first evidence describing the role of polycyclitol
derivatives on learning and memory functions in ECM. Christopher D
Morrone et al., reported that administration of scyllo -inositol
reduced the levels of amyloid beta and reversed the cognitive decline in
TgF344-AD rats (Morrone et al., 2020). However, failure ofscyllo -inositol (6 ), in phase-II trials resulted in a
setback to this molecule which otherwise exhibited significant
Aβ -42 lowering in patient brains. This failure was attributed to
its toxicity and non-specific binding with the target protein possibly
because of its small size. Considering this failure of scylloinositol (6 ) against Aβ , it was an opportunity to
evaluate our recently accessed polycyclitols against the other important
concerned protein ‘tau’ and map their potential in controlling its
hyperphosphorylation (Rashid et al., 2020). Due to the bulkier size of
polycyclitols (15 , 16-18 ), we expect them to have
increased number of interacting sites which might result in a better
therapeutic potential for neurodegenerative diseases involving
tauopathy. According to a recent reports, tau based therapeutics could
play a potential role in reversing the neuronal cell damage and
cognitive impairment in ECM (Akide Ndunge et al., 2023; Dai et al.,
2012b). ARM alone impairs blood-brain barrier in experimental models of
CM despite improving survivability by parasite clearance (Brejt and
Golightly, 2019; Dai et al., 2012a; Gul et al., 2021). In agreement to
previous research studies, our group showed ARM therapy alone is unable
to restore cognition (Kumar and Babu, 2022). ARM monotherapy also holds
the current problem of anti-malarial resistance in patients with severe
malaria. Therefore, we administered our adjuncts (SR4-01 to 04) along
with ARM instead of ARM alone in our study.
RMCBS scores show that rescue therapy successfully alleviated
neurobehavioral impairment after few days of administration. We strongly
assume that the restoration of RMCBS behavioral parameters is solely due
to the ARM as is the first line therapy for CM clearing parasites in the
blood. According to Clemmer et al., survivability of ARM treated animals
is near to 50 percent (Clemmer et al., 2011), and the same was observed
after rescue ARM adjunctive therapy on day 30. Based on outcomes of
Barnes maze experiment, we assume that SR4-02 (15 ) and SR4-04
(18 ) treated animals restored the spatial reference memory with
minimal error rates. The same experimental groups exhibited a retrieval
of working memory in spontaneous alternation method of T-maze
experiment. SR4-02 group also showed improved learning and novelty in
novel-object recognition test which was similar to the control group.
SR4-01 (16 ) and SR4-03 (17 ) groups showed increased
side preference while performing T-maze experiment. The increase in
error rates in SR4-01 could be due to loss of neuronal density in
hippocampal regions and expression of lower levels of phospho tau, p25
inducing neurotoxic stimuli. SR4-03 group exhibited improved neuronal
arborization pattern but cognitive impairment still persisted. SR4-03
treated brain sections show improved neuronal arborization with lower
restoration of cognition. One of the reasons could be that SR4-01 and
SR4-03 contain a carbonate functionality in common compared to SR4-02
and SR4-04, which makes the former compounds highly reactive and
accelerate ROS generation (Juan et al., 2021)(Sparrow et al., 2003). We
assume that both SR4-01 and SR4-03 drugs with ARM interacts with
hemoglobin, producing toxic oxygen free radicals; the other reason could
be the non-compliance of dosage standardization.
Under pathological conditions, hyperphosphorylated tau localized to
somatodendritic compartments impair neuronal arborization by
dysregulating microtubule regulatory proteins, affecting microtubule
stability and resulting in loss of neuronal density (LaPointe et al.,
2009)(Kimura et al., 2014). Neuronal loss due to tau
hyperphosphorylation is observed in hippocampal, cortical pyramidal
neurons and striatal neurons in Alzheimer’s, Parkinson’s (Kaul et al.,
2011) and dementia with Lewy bodies (Hatch et al., 2017). Previously,
our group showed extensive neurodegeneration associated with
upregulation of NADPH oxidase 2 in CA1 and dentate gyrus regions of
hippocampus (Kumar and Babu, 2022) also known as early vulnerable
regions of neuronal damage in neurodegenerative diseases (Davolio and
Greenamyre, 1995; Planche et al., 2018). Hyperphosphorylation of tau
dissociates microtubules affecting the distal ends followed by proximal
ends of the dendrites of pyramidal neurons in AD (Regalado-Reyes et al.,
2020). As per the findings of Golgi-cox staining, we assume that
decreased lengths of distal and proximal neurons of cortical and
hippocampus is a key indicator for the loss of neuronal plasticity in
CM. p25 and cdk5 hyperactivation followed by tau pathology could
deteriorate dendritic arborization pattern of the pyramidal hippocampal
and cortical neurons in ECM. We assume that hyperphosphorylated form of
tau and accumulation of p25 could be a potential reason for loss of
neuronal density in CM. Restoration of neuronal density after
administration of adjunctives SR4-02, SR4-03 and SR4-04 could be due to
significant reduction of phosphorylated tau, p25 neurotoxicity and total
cdk5 levels after ECM.