Amit Kumar

Pharmacologic investigations on the role of Sirt-1 in neuroprotective mechanism of postconditioning in mice.

BACKGROUND Cerebral ischemia-reperfusion (I-R) injury is one of the primary causes of ischemic stroke. Ischemic postconditioning (iPoCo) is evolving as an important adaptive technique to contain I-R injury. Some recent studies have shown neuroprotective effects of iPoCo. However, the neuroprotective mechanism of iPoCo is not clear. So, the present study has been undertaken to investigate the possible role of Sirtinol, a selective class III histone deacetylase (HDAC) inhibitor in the neuroprotective mechanism of iPoCo in mice. MATERIAL AND METHODS Bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 h was used to produce I-R-induced cerebral injury in Swiss albino mice. iPoCo involving three episodes of 10-s carotid artery occlusion and reperfusion instituted immediately after BCAO just before prolonged reperfusion of 24 h. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was evaluated using a Morris water maze test. Rotarod test, inclined beam-walking test, and neurologic severity score (NSS) were used to assess motor incoordination. Acetylcholine esterase levels, brain thiobarbituric acid reactive species (TBARS), and glutathione level were also estimated. RESULTS BCAO for 12 min followed by reperfusion for 24 h produced a significant rise in cerebral infarct size and NSS along with impairment of memory and motor coordination and biochemical alteration (↑acetylcholine esterase, ↓glutathione, and ↑TBARS). iPoCo, involving three episodes of 10-s carotid artery occlusion with intermittent reperfusion of 10 s applied just after ischemic insult of 12 min produced a significant decrease in cerebral infarct size and NSS along with the reversal of I-R-induced impairment of memory and motor coordination. iPoCo-induced neuroprotective effects were significantly abolished by pretreatment with selective SIRT 1 (class III HDAC) blocker Sirtinol (10 mg/kg intraperitoneal). CONCLUSIONS It may be concluded that the neuroprotective effect of iPoCo probably involves activation of SIRT 1 (class III HDAC) enzyme.

Silymarin ameliorates memory deficits and neuropathological changes in mouse model of high-fat-diet-induced experimental dementia

A huge body evidences suggest that obesity is the single great risk factor for the development of dementia. Recently, silymarin, a flavonoid, clinically in use as a hepatoprotectant, has been reported to prevent amyloid beta-induced memory impairment by reducing oxidative stress and inflammation in mice brain. However, its potential in high-fat-diet (HFD)-induced dementia has not yet been investigated. Therefore, the present study is designed to explore the role of silymarin in HFD-induced experimental dementia in mice. Morris water maze test was employed to assess learning and memory. Various biochemical estimations including brain acetylcholinerstarse activity (AchE), thiobarbituric acid-reactive species (TBARS) level, reduced glutathione level (GSH), nirate/nitrite, and myeloperoxidase (MPO) activity were measured. Serum cholesterol level was also determined. HFD significantly impaired the cognitive abilities, along with increasing brain AchE, TBARS, MPO, nitrate/nitrite, and serum cholesterol levels. Marked reduction of brain GSH levels was observed. On the contrary, silymarin significantly reversed HFD-induced cognitive deficits and the biochemical changes. The present study indicates strong potential of silymarin in HFD-induced experimental dementia.

Efficacy of Cilostazol a selective phosphodiesterase-3 inhibitor in rat model of Streptozotocin diabetes induced vascular dementia

The present study has been designed to investigate the potential of Cilostazol a phosphodiesterase-3 (PDE-3) inhibitor in diabetes-induced vascular dementia (Vad) employing Wistar rats. A single dose of Streptozotocin (STZ) was used for the induction of diabetes and subsequent Vad in rats. Memory and learning abilities of rats were evaluated with Morris water maze (MWM) test. Serum glucose, body weight, vascular endothelial function, serum nitrite/nitrate levels, brain oxidative stress levels (viz. brain thiobarbituric acid reactive species and reduced glutathione levels), inflammatory markers (viz. brain myeloperoxidase activity and neutrophil infiltration in the brain hippocampal area) and brain acetylcholinesterase activity were also tested. Donepezil was used as positive control. Streptozotocin treated animals showed poor performance on MWM indicating impairment of learning and memory abilities with a significant reduction in body weight, vascular endothelial function, serum nitrite/nitrate levels, along with an increase in serum glucose, brain oxidative stress levels, inflammatory changes and brain acetylcholinesterase activity. Treatment with selective PDE-3 inhibitor, Cilostazol significantly attenuated, diabetes-induced impairment of learning and memory; endothelial dysfunction, and changes in various biochemical parameters. It is concluded that selective PDE-3 inhibitor, Cilostazol may be considered as the potential pharmacological agent for the management of diabetes-induced vascular dementia.

Pharmacological investigations on possible role of Src kinases in neuroprotective mechanism of ischemic postconditioning in mice

The present study has been undertaken to investigate the possible role of Src Kinases in a neuroprotective mechanism of ischemic postconditioning in mice. Bilateral carotid artery occlusion for 12 min followed by reperfusion for 24 h produced a significant increase in cerebral infarct size and neurological severity score along with impairment of memory and motor coordination. Ischemic postconditioning involving three episodes of 10 s carotid artery occlusion with intermittent reperfusion of 10 s proceeding ischemic insult of 12 min, produced a significant decrease in cerebral infarct size and neurological severity score along with reversal of ischemia-reperfusion induced impairment of memory and motor coordination. Ischemic postconditioning induced neuroprotective effects were significantly attenuated by pre-treatment of selective Src Kinase inhibitors SU-6656 (4 mg/kg i.p.) and PP1 (0.2 mg/kg i.p.). It may be concluded that the neuroprotective effect of ischemic postconditioning probably involves activation of Src Kinase pathway.

Possible role of pannexin 1/P2x7 purinoceptor in neuroprotective mechanism of ischemic postconditioning in mice

BACKGROUND Previous studies have suggested a significant role of pannexin 1 (Panx1)/P2X7 receptor complex in cardioprotective mechanism of ischemic preconditioning and postconditioning (IPC). The present study has been undertaken to investigate whether Panx1/P2X7 purinoceptors are also involved in the neuroprotective mechanism of IPC in mice. MATERIALS AND METHODS Bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 h was used to produce ischemia-reperfusion-induced cerebral injury in Swiss albino mice. For IPC immediately after BCAO of 12 min, three cycles of 10-s ischemia and reperfusion each were given and then prolonged reperfusion of 24 h was used. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was evaluated using a Morris water maze test. Rotarod test, inclined beam walking test, and neurologic severity score (NSS) were used to assess motor dysfunction. Acetylcholine esterase levels, brain thiobarbituric acid reactive species, and glutathione level were also estimated. RESULTS BCAO followed by reperfusion produced a significant increase in cerebral infarct size, NSS along with impairment of memory and motor dysfunction. It also increased brain acetylcholine esterase, thiobarbituric acid reactive species levels, and decreased the glutathione level. IPC produced a significant decrease in the cerebral infarct size and NSS along with reversal of ischemia-reperfusion-induced impairment of memory, motor dysfunction, and altered biochemical levels in the brain. IPC-induced neuroprotective effects were significantly abolished by pretreatment of mefloquine (15.0 mg/kg orally; 30.0 mg/kg orally), blocker of Panx1/P2X7 purinoceptor. CONCLUSIONS Therefore, activation of Panx1/P2X7 purinoceptors appears to play a significant role in the neuroprotective mechanism of IPC.

Inhibitor of Phosphodiestearse-4 improves memory deficits, oxidative stress, neuroinflammation and neuropathological alterations in mouse models of dementia of Alzheimer’s Type

The study investigates the potential of Rolipram a phosphodiesterase-4 inhibitor in cognitive deficits induced by streptozotocin (STZ, 3mg/kg intracerebroventricularly) and natural ageing in mice. Morris water maze (MWM) test was employed to evaluate learning and memory of the animals. Extent of oxidative stress was measured by estimating the levels of brain glutathione (GSH) and thiobarbituric acid reactive species (TBARS). Brain acetylcholinestrase (AChE) activity was also estimated. The brain activity of myeloperoxidase (MPO) was measured as a marker of inflammation. STZ and ageing results in marked decline in MWM performance of the animals, reflecting impairment of learning and memory. STZ treated mice and aged mice exhibited a marked accentuation of AChE activity, TBARS and MPO activity along with fall in GSH level. Further the stained micrographs of STZ treated mice and aged mice indicate pathological changes, severe neutrophilic infiltration and amyloid deposition. Rolipram treatment significantly attenuated STZ induced and age related memory deficits, biochemical and histopathological alterations. The findings demonstrate the potential of Rolipram in memory dysfunctions which may probably be attributed to its anti-cholinesterase, anti-amyloid, anti-oxidative and anti-inflammatory effects. The study concludes that PDE-4 can be explored as a potential therapeutic target in dementia.

Calcineurin inhibitors improve memory loss and neuropathological changes in mouse model of dementia.

Aim: The present study was designed to investigate the potential of Cyclosporine (CsA) and Tacrolimus, the inhibitors of calcineurin (CaN) in cognitive deficits of mice. Methods: Streptozotocin [STZ, 3 mg/kg, injected intracerebroventricular (i.c.v.)] was used to induce memory deficits in NIH mice, while aged mice separately taken served as a natural model of dementia. Morris water maze (MWM) test was employed to evaluate learning and memory of the animals. A battery of biochemical and histopathological studies was also performed. Extent of oxidative stress was measured by estimating the levels of brain glutathione (GSH) and thiobarbituric acid reactive species (TBARS). Brain acetylcholinestrase (AChE) activity was estimated to assess cholinergic activity. The brain level of myeloperoxidase (MPO) was measured as a marker of inflammation. Results: STZ i.c.v. and aging results in marked decline in MWM performance of the animals, reflecting impairment of learning and memory. STZ i.c.v. treated mice and aged mice exhibited a marked accentuation of AChE activity, TBARS and MPO levels along with a fall in GSH level. Further the stained micrographs of STZ treated mice and aged mice indicate pathological changes, severe neutrophilic infiltration and amyloid deposition. Cyclosporine and Tacrolimus treatment significantly attenuated STZ induced and age related memory deficits, biochemical and histopathological alterations. Conclusion: The findings demonstrate the potential of CaN inhibitors Cyclosporine and Tacrolimus in memory dysfunctions which may probably be attributed to anti‐cholinesterase, anti‐amyloid, anti‐oxidative and anti‐inflammatory effects. It is concluded that CaN can be explored as a potential therapeutic target in dementia.

Pharmacology of Src family kinases and therapeutic implications of their modulators

Src family kinases (SFKs), the largest family of nonreceptor tyrosine kinases, include 10 members. Src was the first gene product discovered to have intrinsic protein tyrosine kinase activity. Src is widely expressed in many cell types and can have different locations within a cell; the subcellular location of Src can affect its function. Src can associate with cellular membranes, such as the plasma membrane, the perinuclear membrane, and the endosomal membrane. SFKs actions on mammalian cells are pleiotropic and include effect on cell morphology, adhesion, migration, invasion, proliferation, differentiation, and survival. SFKs at one end have been documented to play some important physiological functions; on the other end, they have been described in the pathophysiology of some disorders. In this review article, an exhaustive attempt has been made to unearth pharmacology of SFKs and therapeutic implications of SFKs modulators.

Amelioration of ischemia-reperfusion induced functional and biochemical deficit in mice by Ocimum kilimandscharicum leaf extract

The genus Ocimum (family Lamiaceae) has been revered for its diverse biological activities. Various species have been used traditionally to treat CNS disorders and are proven to have neuroprotective effect that is often attributed to their significant antioxidant activity. Ocimum kilimandscharicum (Karpoora Thulasi), a prominent member of this genus is reported to have marked antioxidant activity but its neuroprotective potential has not been explored. Thus, present study was designed to evaluate the cerebroprotective effect of O. kilimandscharicum leaf extract (OKLE) in mice against ischemia reperfusion (I-R) induced brain injury. Bilateral common carotid artery occlusion (BCCAO) for 15min followed by 24h reperfusion was used to induce brain damage in Swiss Albino mice. Animals were treated with OKLE (200 and 400mg/kg, po) once daily for 7days after I-R. Morris water maze and elevated plus maze tests were used to assess long and short term memory while neurological severity score was used to determine motor coordination. Histopathological evaluation (TTC staining) along with brain biochemical parameters (TBARS, reduced GSH and SOD activity) were determined to outline neuroprotective mechanism of OKLE. I-R resulted in marked cognitive impairments, motor incoordination in mice, significant brain damage and increased oxidative stress. Treatment with OKLE produced functional recovery in mice which is manifested by improved memory and motor coordination; reduced cerebral infarct size and brain oxidative stress (TBARS levels) and elevated endogenous antioxidants (reduced GSH and SOD activity). In addition, OKLE showed DPPH radical scavenging and reducing power in-vitro. These results show that O. kilimandscharicum mitigated the neurodegenerative changed induced by I-R in mice probably due to its antioxidant activity.

Evidence for the role of histaminergic pathways in neuroprotective mechanism of ischemic postconditioning in mice

The present study has been designed to investigate the possible role of histaminergic pathway in neuroprotective mechanism of ischemic postconditioning (iPoCo). Bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 h was employed to produce I/R‐induced cerebral injury in National Institutes of Health mice mice. iPoCo involving three episodes of carotid artery occlusion and reperfusion of 10 sec each was instituted immediately after BCAO just before prolonged reperfusion. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was evaluated using Morris water maze test. Rotarod test, inclined beam‐walking test, and neurological severity score (NSS) were performed to assess motor incoordination and sensorimotor abilities. Brain acetylcholine esterase (AChE) activity, brain myeloperoxidase (MPO) activity, brain thiobarbituric acid‐reactive species (TBARS), and glutathione level (GSH) were also estimated. BCAO produced a significant rise in cerebral infarct size and NSS along with impairment of memory and motor coordination and biochemical alteration (↑AChE, ↑MPO ↓GSH, and ↑TBARS). iPoCo attenuated the deleterious effect of BCAO on infarct size, memory, NSS, motor coordination, and biochemical markers. Pretreatment of carnosine (a histamine [HA] precursor) potentiated the neuroprotective effects of iPoCo, whereas pretreatment of ketotifen (HA H1 receptor blocker and mast cell stabilizer) abolished the protective effects of iPoCo as well as that of carnosine on iPoCo. It may be concluded that neuroprotective effect of iPoCo probably involves activation of histaminergic pathways.