Evolving possible link between PI3K and NO pathways in neuroprotective mechanism of ischemic postconditioning in mice.

The present study was conducted to pharmacologically investigate the influence of NO signaling pathway in PI3K mediated neuroprotective mechanism of ischemic postconditioning (iPoCo). Bilateral carotid artery occlusion (BCAO) of 12 min followed by reperfusion for 24 h was employed to produce ischemia/reperfusion-induced cerebral injury in male Swiss mice. Memory was assessed using Morris water maze test. Degree of motor incoordination was evaluated using inclined beam walk test, rotarod test, and lateral push test. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Brain acetylcholinesterase activity, thiobarbituric acid reactive species (TBARS), nitrite/nitrate and reduced glutathione (GSH) levels were also estimated. BCAO followed by reperfusion produced significant rise in cerebral infarct size, acetylcholinesterase activity, and TBARS levels along with fall in nitrite/nitrate and GSH levels. A significant impairment of memory and motor coordination was also noted. iPoCo consisting of three episodes of 10 s carotid artery occlusion and reperfusion significantly attenuated infarct size, memory impairment, motor incoordination, and altered biochemicals. iPoCo-induced neuroprotective effects were significantly abolished by wortmannin (a selective PI3K inhibitor). However, administration of L-Arginine (a NO precursor) in the presence of wortmannin restored the protective effect of ischemic postconditioning. It may be concluded that neuroprotective mechanism of iPoCo involves PI3K-mediated pathway with NO signaling as an important downstream step.

Neuroprotective mechanism of ischemic postconditioning in mice: a possible relationship between protein kinase C and nitric oxide pathways.

BACKGROUND: The present study was conducted to pharmacologically investigate the role of protein kinase C (PKC) pathway in neuroprotective mechanism of ischemic postconditioning (iPoCo) and determine the influence of nitric oxide (NO) signaling in PKC-mediated effects. MATERIALS AND METHODS: Bilateral carotid artery occlusion of 12 min followed by reperfusion for 24 h was used to produce ischemia and reperfusion (I/R)-induced cerebral injury in male Swiss mice. Memory was assessed using Morris water maze test. Degree of motor incoordination was evaluated using inclined beam-walk test, rota-rod test, and lateral push test. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Brain acetylcholinesterase activity, thiobarbituric acid reactive species, nitrite/nitrate, and reduced glutathione levels were also estimated. RESULTS: Bilateral carotid artery occlusion followed by reperfusion produced significant rise in cerebral infarct size, acetylcholinesterase activity, and thiobarbituric acid reactive species levels along with the fall in nitrite/nitrate and glutathione levels. A significant impairment of memory and motor coordination was also noted. iPoCo consisting of three episodes of 10 s carotid artery occlusion and reperfusion significantly attenuated infarct size, memory impairment, motor incoordination, and altered biochemicals. iPoCo-induced neuroprotective effects were significantly abolished by chelerythrine (a nonselective PKC inhibitor). L-Arginine, an NO precursor significantly attenuated I/R-induced injury and mimicked the neuroprotective effect of postconditioning. Furthermore, this protective effect of L-arginine on I/R injury and iPoCo was abolished when it was coadministered with chelerythrine. CONCLUSIONS: It may be concluded that neuroprotective mechanism of iPoCo involves PKC mediated pathway with NO signaling as an essential step.

Neuroprotective effect of tadalafil, a PDE-5 inhibitor, and its modulation by L-NAME in mouse model of ischemia-reperfusion injury.

BACKGROUND: The present study investigates the neuroprotective effect of tadalafil, a selective phosphodiesterase-5 inhibitor, in a mouse model of ischemia-reperfusion injury. MATERIALS AND METHODS: Bilateral carotid artery occlusion for 12 min followed by reperfusion for 24 h was employed to produce ischemia-reperfusion-induced cerebral injury in male Swiss mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was assessed using Morris water maze test. Degree of motor incoordination was evaluated using inclined beam walk test, rota-rod test, and lateral push test. Brain nitrite/nitrate, brain acetylcholinesterase activity, brain thiobarbituric acid reactive species, and glutathione levels were also estimated. RESULTS: Bilateral carotid artery occlusion, followed by reperfusion, produced a significant rise in cerebral infarct size, brain nitrite/nitrate levels, acetylcholinesterase activity, and thiobarbituric acid reactive species level along with a fall in glutathione. A significant impairment of memory and motor coordination was also noted. Pretreatment of tadalafil significantly attenuated the above effects of ischemia-reperfusion injury. Tadalafil-induced neuroprotective effects were significantly attenuated by administration of L-NAME, a nonselective nitric oxide synthase inhibitor. CONCLUSIONS: Results indicate that tadalafil exerts neuroprotective effects, probably through nitric oxide-dependent pathways. Therefore, phosphodiesterase-5 can be explored as an important target to contain ischemia-reperfusion injury.

Pharmacologic evidence for role of endothelial nitric oxide synthase in neuroprotective mechanism of ischemic postconditioning in mice.

BACKGROUND: The present study was conducted to pharmacologically investigate the isoform-specific role of nitric oxide pathway in neuroprotective mechanism of ischemic postconditioning (iPoCo). MATERIALS AND METHODS: Bilateral carotid artery occlusion of 12 min followed by reperfusion for 24 h was used to produce ischemia- and reperfusion-induced cerebral injury in male Swiss mice. Memory was assessed using Morris water maze test. Degree of motor in-coordination was evaluated using inclined beam-walk test, rotarod test, and lateral push test. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Brain acetylcholinestrase activity, thiobarbituric acid-reactive species, nitrite/nitrate, and reduced glutathione levels were also estimated. Western blotting was performed to determine endothelial nitric oxide synthase (eNOS) expression. RESULTS: Bilateral carotid artery occlusion followed by reperfusion produced significant rise in cerebral infarct size, acetylcholinesterase activity, and thiobarbituric acid-reactive species levels along with fall in nitrite/nitrate, and glutathione and eNOS expression levels. A significant impairment of memory and motor coordination was also noted. iPoCo consisting of three episodes of 10-s carotid artery occlusion and reperfusion significantly attenuated infarct size, memory impairment, motor in-coordination, altered biochemicals, and protein expression levels. iPoCo-induced neuroprotective effects were significantly abolished by L-NAME (a nonselective nitric oxide synthase inhibitor) and L-NIO (a selective eNOS inhibitor). However, aminoguanidine (a selective inducible nitric oxide synthase inhibitor) and 7-nitroindazole (a selective neuronal nitric oxide synthase inhibitor) did not modulate beneficial effects of iPoCo. CONCLUSIONS: It may be concluded that nitric oxide pathway probably plays a vital role with specific involvement of eNOS in neuroprotective mechanism of iPoCo.

Tadalafil enhances the neuroprotective effects of ischemic postconditioning in mice, probably in a nitric oxide associated manner.

This study investigates the modulatory effect of tadalafil, a selective phosphodiesterase (PDE-5) inhibitor, on the neuroprotective effects of ischemic postconditioning (iPoCo) in mice. Bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 h was employed to produce ischemia and reperfusion induced cerebral injury. Cerebral infarct size was measured using TTC staining. Memory was assessed using the Morris water maze test. Degree of motor incoordination was evaluated using inclined beam-walking, rota-rod, and lateral push tests. Brain nitrite/nitrate, acetylcholinesterase activity, TBARS, and glutathione levels were also estimated. BCAO followed by reperfusion produced a significant increase in cerebral infarct size, brain nitrite/nitrate and TBARS levels, and acetylcholinesterase activity along with a reduction in glutathione. Marked impairment of memory and motor coordination was also noted. iPoCo consisting of 3 episodes of 10 s carotid artery occlusion and reperfusion instituted immediately after BCAO significantly decreased infarct size, memory impairment, motor incoordination, and altered biochemistry. Pretreatment with tadalafil mimicked the neuroprotective effects of iPoCo. The tadalafil-induced neuroprotective effects were significantly attenuated by l-NAME, a nonselective NOS inhibitor. We concluded that tadalafil mimics the neuroprotective effects of iPoCo, probably through a nitric oxide dependent pathway, and PDE-5 could be a target of interest with respect to the neuroprotective mechanism of iPoCo.

Secondary prevention by structured semi-interactive stroke prevention package in INDIA (SPRINT INDIA): Findings from the process evaluation of a randomized controlled trial.

INTRODUCTION: Secondary Prevention by Structured Semi-Interactive Stroke Prevention Package in INDIA Trial delivered secondary stroke awareness intervention to sub-acute stroke patients in form of workbook, videos and SMS across 31 centres in 12 languages. Trial was stopped for futility due to fewer vascular outcomes than anticipated. Trial results indicated that trial intervention, did not lead to reduction in vascular events. We carried out process evaluation, to evaluate trial implementation and participant's perspectives, to comprehend the trial's futile outcomes. MATERIALS AND METHODS: Using mixed methods approach, qualitative interviews and quantitative data from case report forms, workbooks and questionnaires were analysed to measure intervention fidelity and contamination. Using purposive sampling, 115 interviews of patient-caregiver dyads and health professionals at 11 centres and 2 focus group discussions were held. RESULTS AND DISCUSSION: Iterative thematic analysis of qualitative data was done with RE-AIM and realist models. There was good fidelity to intervention and adherence to protocol; however, there was dilution of inclusion criteria by randomly enrolling uneducated and caregiver-dependent patients. Centre coordinators provided counselling to both arms, not specified by protocol, causing bias. Coordinators found it difficult to keep patients motivated to view intervention which was corroborated by fidelity questionnaire showing decreased viewing of intervention for a year. Cardiovascular protection improved in routine care by virtue of participating in trial. No contamination of intervention was reported. CONCLUSION: The intervention was acceptable by patients and caregivers, which could be made a community-based programme. Reasons identified for decreased viewing were repetitive content and non-availability of personal cellular device.

Discovery of potent inhibitors for Mpro enzyme of SARS-COV2 by multi-stage in-silico screening of Alkannin/shikonin.

Novel coronavirus disease, a serious challenge for the healthcare system, has diverted all the researchers toward the exploration of potential targets, compounds or vaccines for the management of this disease. Mpro enzyme was found to be crucial for replication of this virus which makes this enzyme an attractive drug target for SARS-CoV-2. Diverse pharmacological profile of Alkannin/shikonin (A/S) derivatives build up curiosity to study their antiviral profile. Therefore, current study utilises various computational tools to screen and evaluate all the discovered A/S derivatives to inhibit the Mpro enzyme for its anti-viral activity. Results revealed that the A/S has a very good tendency to inhibit the catalytic activity of the enzyme. Moreover, (5 R,6R)-5,8-dihydroxy-6-methoxy-3,4,5,6-tetrahydro-2H-benzo[a]anthracene-1, 7, 12-trione, an A/S derivative was found to possess drug-likeliness properties and a good ADME profile. Moreover, its complex with Mpro enzyme was found stable for 50 ns which makes it a very promising ligand to treat COVID-19.

Phytoconstituents from ten natural herbs as potent inhibitors of main protease enzyme of SARS-COV-2: In silico study.

Background: Lack of treatment of novel Coronavirus disease led to the search of specific antivirals that are capable to inhibit the replication of the virus. The plant kingdom has demonstrated to be an important source of new molecules with antiviral potential. Purpose: The present study aims to utilize various computational tools to identify the most eligible drug candidate that have capabilities to halt the replication of SARS-COV-2 virus by inhibiting Main protease (Mpro) enzyme. Methods: We have selected plants whose extracts have inhibitory potential against previously discovered coronaviruses. Their phytoconstituents were surveyed and a library of 100 molecules was prepared. Then, computational tools such as molecular docking, ADMET and molecular dynamic simulations were utilized to screen the compounds and evaluate them against Mpro enzyme. Results: All the phytoconstituents showed good binding affinities towards Mpro enzyme. Among them laurolitsine possesses the highest binding affinity i.e. -294.1533 kcal/mol. On ADMET analysis of best three ligands were simulated for 1.2 ns, then the stable ligand among them was further simulated for 20 ns. Results revealed that no conformational changes were observed in the laurolitsine w.r.t. protein residues and low RMSD value suggested that the Laurolitsine-protein complex was stable for 20 ns. Conclusion: Laurolitsine, an active constituent of roots of Lindera aggregata, was found to be having good ADMET profile and have capabilities to halt the activity of the enzyme. Therefore, this makes laurolitsine a good drug candidate for the treatment of COVID-19.

Investigation of the role of non-selective calcium channel blocker (flunarizine) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice.

The present study was designed to investigate the role of flunarizine (a non-selective calcium channel blocker) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice. Bilateral carotid artery occlusion of 12min followed by reperfusion for 24h was given to induce cerebral injury in male Swiss mice. The assessment of learning & memory was performed by Morris water maze test; motor in-coordination was evaluated by rota rod, lateral push and inclined beam walking tests; cerebral infarct size was quantified by triphenyltetrazolium chloride staining. In addition, reduced glutathione (GSH), total calcium and acetylcholinesterase (AChE) activity were also estimated in aged brain tissue. Donepezil treated group served as a positive control in this study. Ischemia reperfusion (I/R) injury produced significant increase in cerebral infarct size. A significant loss of memory along with impairment of motor performance was also noted. Further, I/R injury also produced significant increase in levels of total calcium, AChE activity and decrease in GSH levels. Pretreatment of flunarizine significantly attenuated I/R induced infarct size, behavioral and biochemical changes. Hence, it may be concluded that, a non-selective calcium channel blocker can be useful in I/R associated cognitive dysfunction due to its anti-oxidant, anti-infarct and modulatory actions of neurotransmitters & calcium channels.

Pharmacological evidence for connection of nitric oxide-mediated pathways in neuroprotective mechanism of ischemic postconditioning in mice.

INTRODUCTION: Postconditioning (PoCo) is an adaptive phenomenon whereby brief repetitive cycles of ischemia with intermittent reperfusion instituted immediately after prolonged ischemia at the onset of prolonged reperfusion elicit tissue protection. PoCo is noted to exert a protective effect in various organs like heart, liver, kidney and brain. Various triggers, mediators and end effectors are suggested to contribute to the protective effect of PoCo. However, the neuroprotective mechanism of PoCo is poorly understood. OBJECTIVES: The present study has been designed to investigate the role of nitric oxide pathway in the neuroprotective mechanism of ischemic postconditioning (iPoCo) employing a mouse model of global cerebral ischemia and reperfusion-induced injury. MATERIALS AND METHODS: Bilateral carotid artery occlusion (BCAO) of 12 min followed by reperfusion for 24 h was employed to produce ischemia and reperfusion (I/R)-induced cerebral injury in mice. Cerebral injury was assessed in the terms of cerebral infarct, memory impairment and motor in-coordination. Brain nitrite/nitrate; acetylcholinesterase activity, thiobarbituric acid reactive species (TBARS) and glutathione level were also estimated. RESULTS: BCAO followed by reperfusion produced a significant rise in cerebral infarct size, memory impairment and motor incoordination. Further a rise in acetylcholinesterase activity and TBARS level along with fall in brain nitrite/nitrate and glutathione levels was also noted. iPoCo consisting of three episodes of 10 s carotid artery occlusion and reperfusion (instituted immediately after BCAO) significantly attenuated infarct size, memory impairment, motor incoordination as well as altered biochemicals. iPoCo-induced neuroprotective effects were significantly abolished by pretreatment of L-NAME, a nonselective NOS inhibitor. CONCLUSION: It may be concluded that the nitric oxide pathway probably plays a vital role in the neuroprotective mechanism of iPoCo.

Neuroprotective effect of gadolinium: a stretch-activated calcium channel blocker in mouse model of ischemia-reperfusion injury.

The present study was designed to investigate the potential of gadolinium, a stretch-activated calcium channel blocker in ischemic reperfusion (I/R)-induced brain injury in mice. Bilateral carotid artery occlusion of 12 min followed by reperfusion for 24 h was given to induce cerebral injury in male Swiss mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was assessed using Morris water maze test and motor incoordination was evaluated using rota-rod, lateral push, and inclined beam walking tests. In addition, total calcium, thiobarbituric acid reactive substance (TBARS), reduced glutathione (GSH), and acetylcholinesterase (AChE) activity were also estimated in brain tissue. I/R injury produced a significant increase in cerebral infarct size. A significant loss of memory along with impairment of motor performance was also noted. Furthermore, I/R injury also produced a significant increase in levels of TBARS, total calcium, AChE activity, and a decrease in GSH levels. Pretreatment of gadolinium significantly attenuated I/R-induced infarct size, behavioral and biochemical changes. On the basis of the present findings, we can suggest that opening of stretch-activated calcium channel may play a critical role in ischemic reperfusion-induced brain injury and that gadolinium has neuroprotective potential in I/R-induced injury.