(-)-α-bisabolol exerts neuroprotective effects against pentylenetetrazole-induced seizures in rats by targeting inflammation and oxidative stress.

Epilepsy is the most common neurological disorder which is accompanied with behavioral and psychiatric alternations. Current evidences have shown that (-)-α-bisabolol (BSB) possess anti-inflammatory and antioxidative effects in several animal studies. Here, we conducted present study to evaluate its neuroprotective effects against pentylenetetrazole (PTZ)-induced seizures in rats. We used fifty male rats and they were randomly assigned into 5 groups control, BSB100, PTZ, BSB50 + PTZ, BSB100 + PTZ. The animals intraperitoneally received PTZ (45 mg/kg) for ten consecutive days to induce epilepsy model. BSB in doses of 50 and 100 mg/kg was administrated orally one hour before PTZ administration for ten days. The elevated plus maze (EPM) test was carried out to assess anxiety-like behavior. The seizure intensity was evaluated according to modifies Racine's convulsion scale (RCS). Y-maze and passive avoidance were utilized to assess working memory and aversive memory. The expression of pro-inflammatory cytokines and oxidative stress factors were measured using the enzyme-linked immunosorbent assay (ELISA). The neuronal cell loss in the hilar region was assessed using Nissl staining. Results showed that PTZ-treated rats had more seizure intensity, anxiety-like behavior, memory deficits, higher levels of TNF-α, IL-1ß, and oxidative markers. Pre-treatment with BSB 100 significantly inhibited seizure intensity, anxiety-like behavior, and memory deficits; reduced levels of TNF-α, IL-1ß, and MDA oxidative markers. Collectively, outcome of this work shows that BSB at the dose of 100 mg/kg may exert neuroprotective effects by mitigating seizures, oxidative stress, and neuroinflammation, and ameliorates memory and anxiety disorders in the PTZ-induced seizure rats.

Neuroprotective effects of Royal Jelly (RJ) against pentylenetetrazole (PTZ)-induced seizures in rats by targeting inflammation and oxidative stress.

Epilepsy is a chronic neurological condition in which inflammation and oxidative stress play a key role in the pathogenesis. Recently, several studies have suggested that Royal Jelly (RJ) has antioxidant effects. Nevertheless, there is no evidence of its effectiveness against epilepsy. Here, we evaluated its neuroprotective effects at different doses (100 and 200 mg/kg) against pentylenetetrazole (PTZ)-induced seizures. Fifty male Wistar rats were randomly divided into five groups: control, PTZ, RJ100 + PTZ, RJ200 + PTZ and RJ100. In order to establish epilepsy model, 45 mg/kg of PTZ was injected intraperitoneally for 10 consecutive days. Seizure parameters were graded based on Racine's 7-point classification. Elevated-plus maze, Y maze and shuttle box tests were carried out to assess anxiety-like behavior, short-term memory, and passive avoidance memory, respectively. We used ELISA technique to measure the expression of the pro-inflammatory cytokines and oxidative stress factors. Also, neuronal loss in the hippocampal CA3 region was determined using Nissl staining. Our findings showed that PTZ-treated rats had more seizure intensity, anxiety-like behavior, memory dysfunction, higher levels of TNF-α, IL-1ß, and oxidative markers. RJ could allay seizure severity and duration. It also improved memory function as well as anxiety level. In terms of biochemical assessment, RJ gave rise to a significant decrease in the level of IL-1ß, TNF-α and MDA and it restored the activities of GPX and SOD enzymes. Hence, our study shows that RJ contains anti-inflammatory and antioxidative effects which contribute to less neuronal damage in the PTZ-induced epilepsy model.

Neuroprotective Effects of Conditioned Medium of Mesenchymal Stem Cells (MSC-CM) as a Therapy for Ischemic Stroke Recovery: A Systematic Review.

It has been reported that the therapeutic potential of stem cells is mainly mediated by their paracrine factors. In order to identify the effects of conditioned medium of mesenchymal stem cells (MSC-CM) against stroke, a systematic review was conducted. We searched PubMed, Scopus, and ISI Web of Science databases for all available articles relevant to the effects of MSC-CM against the middle cerebral artery occlusion (MCAO) model of ischemic stroke until August 2022. The quality of the included studies was evaluated using The STAIR scale. During the systematic search, a total of 356 published articles were found. A total of 15 datasets were included following screening for eligibility. The type of cerebral ischemia was the MCAO model and CM was obtained from MSCs. The results showed that the therapeutic time window can be considered a crucial factor when researchers use MSC-CM for stroke therapy. In addition, MSC-CM therapy contributes to functional recovery and reduces infarct volume after stroke by targeting different cellular signaling pathways. Our findings showed that MSC-CM therapy has the ability to improve functional recovery and attenuate brain infarct volume after ischemic stroke in preclinical studies. We hope our study accelerates needed progress towards clinical trials.

Apelin-13 attenuates cerebral ischemia/reperfusion injury through regulating inflammation and targeting the JAK2/STAT3 signaling pathway.

BACKGROUND: The precise mechanisms whereby apelin-13 acts against ischemic stroke have remained in the dark. Hence, this study aims to examine the effects of apelin-13 on hypothalamic-pituitary-adrenal (HPA) axis over activation, Jak2-STAT3 signaling pathway, and inflammation following ischemic stroke. METHODS: Middle cerebral artery occlusion (MCAO) was used to induce the cerebral ischemic/reperfusion injury (I/RI). Thirty-five male Wistar rats (250-300 g, 8 weeks old) were randomly divided into sham, MCAO, and intravenous (IV) apelin-13 treated groups which received 10, 20, and 40 µg/kg 5 min before reperfusion (n = 7). Neurological status (modified Longa scoring scale), infarct volume, serum levels of malondialdehyde (MDA), total antioxidant capacity (TAC), interleukin 6 (IL-6), corticosterone, and the expressions of the Jak2/STAT3 were assessed. RESULTS: Our results confirm that IV administration of all three doses of apelin-13 significantly improved neurological defects and reduced infarct volume following cerebral I/RI. Furthermore, we observed that acute stroke caused a rise in the expression of the Jak2/STAT3, IL-6, corticosterone, and MDA content, while apelin-13 could reduce the expression of the Jak2/STAT3 and the serum indices in a dose-dependent manner. The 40 µg/kg dose of apelin-13 was also more effective in reducing the infarct volume and improving TAC. CONCLUSION: Our findings suggest that apelin-13 has protective effects against cerebral I/RI-related inflammation and also could attenuate the HPA axis over activation.

Eotaxin-1 (CCL11) in neuroinflammatory disorders and possible role in COVID-19 neurologic complications.

The related neurologic complications of SARS-CoV-2 infection in COVID-19 patients and survivors comprise symptoms including depression, anxiety, muscle pain, dizziness, headaches, fatigue, and anosmia/hyposmia that may continue for months. Recent studies have been demonstrated that chemokines have brain-specific attraction and effects such as chemotaxis, cell adhesion, modulation of neuroendocrine functions, and neuroinflammation. CCL11 is a member of the eotaxin family that is chemotactic agents for eosinophils and participate in innate immunity. Eotaxins may exert physiological and pathological functions in the central nerve system, and CCL11 may induce neuronal cytotoxicity effects by inducing the production of reactive oxygen species (ROS) in microglia cells. Plasma levels of CCL11 elevated in neuroinflammation and neurodegenerative disorders. COVID-19 patients display elevations in CCL11 levels. As CCL11 plays roles in physiosomatic and neuroinflammation, analyzing the level of this chemokine in COVID-19 patients during hospitalization and to predicting post-COVID-19-related neurologic complications may be worthwhile. Moreover, using chemokine modulators may be helpful in lessening the neurologic complications in such patients.

Necroptosis and RhoA/ROCK pathways: molecular targets of Nesfatin-1 in cardioprotection against myocardial ischemia/reperfusion injury in a rat model.

Nesfatin-1 as a new energy-regulating peptide has been known to display a pivotal role in modulation of cardiovascular functions and protection against ischemia/reperfusion injury. However, the detailed knowledge about molecular mechanisms underlying this protection has not been completely investigated yet. This study was designed to clarify the molecular mechanisms by which nesfatin-1 exert cardioprotection effects against myocardial ischemia-reperfusion (MI/R). Left anterior descending coronary artery (LAD) was ligated for 30 min to create a MI/R model in rats. MI/R rats were treated with three concentrations of nesfatin-1 (10, 15 and 20 µg/kg) then expression of necroptosis and necrosis mediators were measured by western blotting assay. Fibrosis, morphological damages, cardiac function, myocardial injury indictors and oxidative stress factors were evaluated as well. Induction of MI/R model resulted in cardiac dysfunction, oxidative stress, increased activity of RIPK1-RIPK3-MLKL axis and RhoA/ROCK pathway, extension of fibrosis and heart tissue damage. Highest tested concentration of nesfatin-1 markedly improved cardiac function. Moreover, it reduced oxidative stress, collagen deposition, and morphological damages, through inhibiting the expression of necroptosis mediators and also, necrosis including RIPK1, RIPK3, MLKL, ROCK1, and ROCK2 proteins. The lowest and middle tested concentrations of nesfatin-1 failed to exert protective effects against MI/R. These findings have shown that nesfatin-1 can exert cardioprotection against MI/R in a dose dependent manner by suppressing necroptosis via modulation of RIPK1-RIPK3-MLKL axis and RhoA/ROCK/RIP3 signaling pathway.

TLR4 /NF-ĸB and JAK2/STAT3 signaling pathways: Cellular signaling pathways targeted by cell-conditioned medium therapy in protection against ischemic stroke.

Human amniotic membrane-derived mesenchymal stem cell-conditioned medium (hAMSC-CM) has been known to improve neuronal survival following ischemic stroke. The present study was designed to examine whether protective effects of hAMSC-CM against stroke can be linked to reducing neuroinflamation by targeting TLR4 /NF-ĸB and Jak2/Stat3 signaling pathways. Immunohistochemistry of hippocampus and western blot assay were performed to evaluate the expression of TLR4 /NF-ĸB and Jak2/Stat3, respectively. Real-time PCR assay was applied to investigate the mRNA levels of Jak2/Stat3. Hematoxylin and eosin (H&E) staining was used to investigate tissue damage and morphological changes in the CA1 region of hippocampus. Increased brain edema was seen in middle cerebral artery occlusion (MCAO) rats compared to sham. Post-treatment with hAMSC-CM markedly reduced brain edema in comparison with MCAO group (P < 0.05). Compared to sham, significantly increased levels of TLR4 /NF-ĸB and Jak2/Stat3 were seen in MCAO rats. Intravenous injection of hAMSC-CM after reperfusion markedly reduced levels of TLR4 /NF-ĸB and Jak2/Stat3 in hippocampus region (P < 0.05). Tissue damage and neuronal cell increased in the CA1 region of hippocampus that reversed by post-treatment by hAMSC-CM. Interestingly, our finding showed that hAMSC-CM can be considered as good candidate to reduce injury following ischemic stroke by decreasing activity of TLR4 /NF-ĸB and Jak2/Stat3 signaling pathways.

FoxO1 and Wnt/ß-catenin signaling pathway: Molecular targets of human amniotic mesenchymal stem cells-derived conditioned medium (hAMSC-CM) in protection against cerebral ischemia/reperfusion injury.

Ischemia-reperfusion (I/R) injury has weakened the effects of available treatment options for ischemic stroke. Although conditioned medium obtained from human amniotic mesenchymal stem cells (hAMSC-CM) has been reported to exert protective effect against stroke, detailed knowledge about its possible molecular mechanisms is not still completely available. The present study was designed to investigate whether hAMSC-CM can modulate FoxO1 and Wnt/ß-catenin signaling pathway after ischemic stroke to create neuroprotective effects. Middle cerebral artery occlusion (MCAO) model with male Wistar rats was used to evaluate the effects of hAMSC-CM on activities of FoxO1, Wnt/ß-catenin signaling pathway, and endogenous antioxidant system and apoptotic cell death. The results demonstrated that induction of MCAO significantly reduced activities of FoxO1, Wnt/ß-catenin signaling pathway, and endogenous antioxidant system and enhanced apoptotic cell death (P < 0.05). In addition, treatment by hAMSC-CM immediately after cerebral reperfusion resulted in significantly reduced infarct size and increased activities of FoxO1, Wnt/ß-catenin signaling pathway, and restoring endogenous antioxidant system and suppressing apoptotic cell death (P < 0.05). Likewise, increased activity of Wnt/ß-catenin signaling pathway resulted in suppressing the neuroinflammation by inhibiting the expression of TNF-α and increasing the expression of IL-10. These findings demonstrate that hAMSC-CM can be considered as an excellent candidate in the treatment of acute ischemic stroke in clinical routine.

Intravenous injection of apelin-13 improves sensory-motor balance deficits caused by cerebral ischemic reperfusion injury in male wistar rats via restoration of nitric oxide.

It has been reported that apelin-13 possesses neuroprotective effects against cerebral ischemia/reperfusion injury (IRI). Disabilities in sense, movement and balance are the major stroke complications which, result in a high rate of mortality. Here, effects of intravenous (IV) injection of apelin-13 on the severity of neural death, infarct volume, neurological defects and its association with nitric oxide (NO) were investigated. A rat model of cerebral IRI was created by middle cerebral artery occlusion (MCAO) for 60 min and restoration of blood flow for 23 h. Animals were randomly assigned into six groups: sham, ischemia (MCAO), vehicle (MCAO + PBS) and three treatment groups (MCAO + apelin-13 in 10, 20, 40 µg/kg doses, IV). All injections were carried out via tail vein injection 5 min before reperfusion. Neural loss and infarct volume were evaluated by Nissl and 2,3,5-triphenyltetrazolium chloride (TTC) staining, respectively. Neurological defects were scored by standard modified criteria. Serum NO was measured by colorimetric method. Apelin-13 in doses of 20 and 40 µg/kg significantly reduced neural death, infarct volume and disturbance of sensory-motor balance compared to control and vehicle groups (p < 0.05). Serum NO levels reduced in MCAO groups compared to sham. Apelin-13 restored serum NO levels at 20 µg/kg dose (p < 0.05). Our data showed beneficial effect of IV injection of apelin-13 on sensory-motor balance defects by reducing neural death and restoration of serum NO levels. The present study shows the validity of apelin-13 in treatment of ischemic stroke in different administration methods.

Donepezil attenuates injury following ischaemic stroke by stimulation of neurogenesis, angiogenesis, and inhibition of inflammation and apoptosis.

Donepezil has proven to be an effective drug to reduce neuronal death and subsequently injury in neurodegenerative diseases. The current study evaluated the neuroprotective effects of donepezil in a rat model of ischaemic stroke and explored possible mechanisms which by this drug may reduce cell death. Temporary middle cerebral artery occlusion (tMCAO) was exerted for 45 min to induce ischaemic stroke. The animals were assigned into five groups: sham, control, and three groups treated with different doses of donepezil. Donepezil was intraperitoneally (IP) injected 4 h after reperfusion for 10 consecutive days. Infarct size was determined using TTC staining. The expression of proteins was evaluated using immunohistochemistry assays. Compared with the control group, infarct size was significantly reduced in tMCAO rats treated with different doses of donepezil. Moreover, our results showed significant decreased expression levels of apoptotic markers and pro-inflammatory mediators after treatment with different doses of donepezil for 10 days (P < 0.05). Likewise, significant increase of brain-derived neurotrophic factor (BDNF) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) proteins were found in tMCAO rats treated with donepezil compared with the control group (P < 0.05). Collectively, our findings show the validity of donepezil as a new therapeutic agent for attenuation of injury following ischaemic stroke through attenuation of inflammation and improvement of mitochondrial function, neurogenesis, and angiogenesis.

Human amniotic membrane mesenchymal stem cells-conditioned medium attenuates myocardial ischemia-reperfusion injury in rats by targeting oxidative stress.

OBJECTIVES: Ischemic heart diseases (IHD) are one of the major causes of death worldwide. Studies have shown that mesenchymal stem cells can secrete and release conditioned medium (CM) which has biological activities and can repair tissue injury. This study aimed to investigate the effects of human amniotic membrane mesenchymal stem cells (hAMCs)-CM on myocardial ischemia/reperfusion (I/R) injury in rats by targeting oxidative stress. MATERIALS AND METHODS: Male Wistar rats (40 rats, weighing 200-250 g) were randomly divided into four groups: Sham, myocardial infarction (MI), MI + culture media, and MI + conditioned medium. MI was induced by ligation of the left anterior descending coronary artery for 30 min. After 15 min of reperfusion, intramyocardial injections of hAMCs-CM or culture media (150 µl) were performed. At the end of the experiment, serum levels of cardiac troponin-I (cTn-I), myocardial levels of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GPx), as well as cardiac histological changes were evaluated. RESULTS: HAMCs-CM significantly decreased cTn-I and MDA levels and increased SOD and GPx activities (P<0.05). In addition, hAMCs-CM improved cardiac histological changes and decreased myocardial injury percentage (P<0.05). CONCLUSION: This study showed that hAMCs-CM has cardioprotective effects in the I/R injury condition. Reduction of oxidative stress by hAMCs-CM plays a significant role in this context. Based on the results of this study, it can be concluded that hAMCs-CM can be offered as a therapeutic candidate for I/R injury in the future, but more research is needed.

Comparison of the effects of intramyocardial and intravenous injections of human mesenchymal stem cells on cardiac regeneration after heart failure.

OBJECTIVES: Existing studies have demonstrated that intravenous and intramyocardial-administrated mesenchymal stem cells (MSCs) lead to tissue repair after cardiac disorders. We compared the efficiency of both administration methods. MATERIALS AND METHODS: A rat model of isoproterenol-induced heart failure (ISO-HF) was established to compare the effects of intravenous and intramyocardial-administrated MSCs on cardiac fibrosis and function. The animals were randomly assigned into six groups: i) control or normal, ii) ISO-HF (HF) iii) ISO-HF rats treated with intramyocardial administration of culture medium (HF+IM/CM), iv) ISO-HF rats treated with intravenous administration of culture medium ( HF+IV/CM), v) ISO-HF rats treated with intravenous administration of MSCs (HF+IV/MSCs), vi) ISO-HF rats treated with intramyocardial administration of MSCs ( HF+IM/MSCs). Cultured MSCs and culture medium were administrated at 4 weeks after final injection of ISO. Heart function, identification of MSCs, osteogenic differentiation, adipogenic differentiation, cardiac fibrosis and tissue damage were evaluated by echocardiography, flow-cytometery, von Kossa, oil red O, Masson's trichrome and H & E staining, respectively. RESULTS: Both intravenous and intramyocardial MSCs therapy significantly improved heart function and reduced cardiac fibrosis and tissue damage (P<0.05), whereas the cultured medium had no beneficial effects. CONCLUSION: In sum, our results confirm the validity of both administration methods in recovery of HF, but more future research is required.

Conditioned medium obtained from human amniotic mesenchymal stem cells attenuates focal cerebral ischemia/reperfusion injury in rats by targeting mTOR pathway.

Conditioned medium obtained from human amniotic mesenchymal stem cells (hAMSC-CM) was recently shown to have many antioxidant, antiapoptotic and proangiogenic growth factors. The present study was performed to investigate whether protective effects of hAMSC-CM against focal cerebral ischemia/ reperfusion (I/R) injury is associated with modulation of the mammalian target of rapamycin (mTOR) pathway. A rat model of middle cerebral artery occlusion (MCAO) was created and the animals were divided into three groups including sham, MCAO and MCAO + hAMSC-CM. Drug was administrated immediately after cerebral reperfusion (i.v). The expressions of mTOR, p-mTOR and LC3 were measured using Western blotting and real time-PCR, respectively. Apoptosis and neuronal loss were determined using TUNEL and Nissl staining, respectively. Infarct volume and the blood-brain barrier (BBB) damage were evaluated using 2,3,5-triphenyltetrazolium chloride (TTC) staining and Evans Blue (EB) uptake, respectively. Compared with sham, significant infarct volume, apoptotic cell death, and neuronal loss were found in MCAO rats that reversed by hAMSC-CM (P < 0.05). Likewise, MCAO rats exhibited increased mRNA level of light-chain 3 (LC3) and the LC3II/LC3I ratio as well as decreased expression level of p-mTOR that reversed by hAMSC-CM (P < 0.05). There were no significant differences in the expression of total mTOR among the experimental groups. In summary, our results demonstrate that hAMSC-CM gives rise to neuroprotection following ischemic stroke by restoring mTOR activity and inhibiting autophagy.

Conditioned medium obtained from mesenchymal stem cells attenuates focal cerebral ischemia reperfusion injury through activation of ERK1/ERK2-BDNF signaling pathway.

Previous studies have shown that conditioned medium (CM) obtained from mesenchymal stem cells (MSCs) might exert neuroprotective effects against focal cerebral ischemia reperfusion (I/R) injury. This study was conducted to investigate if CM obtained from MSCs gives rise to neuroprotection by targeting neurogenesis. To induce focal cerebral ischemia in rats, middle cerebral artery (MCA) was occluded for 1 h and the amniotic mesenchymal stem cells-conditioned medium (AMSC-CM) at the dose of 0.5 µl was administered 30 min after reperfusion by stereotactic intracerebral infusion. The animals were randomly divided into three groups: sham operated animals received all procedures except occlusion of MCA (sham, n = 12), I/R group only received occlusion of MCA (MCAO, n = 17), treatment group received MCAO + 0.5 µl of AMSC-CM (MCAO + AMSC-CM, n = 17). The expression of Phospho-ERK1/ERK2, BDNF, VEGF and NGF were determined using immunohistochemical assay. Neuronal loss and DNA fragmentation were evaluated by Nissl and TUNEL assay, respectively. Our results demonstrated that the expression of Phospho-ERK1/ERK2 and BDNF, VEGF and NGF significantly decreased in MCAO rats and was reversed by AMSC-CM. Likewise, AMSC-CM markedly reduced neuronal loss and DNA fragmentation at 24 h after reperfusion. In sum, our study showed that AMSC-CM administration at the onset of reperfusion led to neuroprotection by activating neuronal ERK1/ERK2-BDNF signaling pathway, neurogenesis, angiogenesis as well as suppression of apoptosis.

Conditioned medium obtained from human amniotic membrane-derived mesenchymal stem cell attenuates heart failure injury in rats.

OBJECTIVES: Heart failure (HF) is one of the leading causes of death worldwide. Due to beneficial effects of stem cells, paracrine secretion of them has recently been used by researchers. The purpose of this study was to investigate the effects of intravenous injection (IV) of conditioned medium (CM) of human amniotic membrane-derived mesenchymal stem cell (MSC-CM) on HF. MATERIALS AND METHODS: Male Wistar rats (n=35, 180 g) were randomly divided into five groups: sham, HF, HF+MSC-CM, HF+culture medium and HF+phosphate-buffered saline (PBS). To induce HF, isoproterenol (170 mg/kg/d) was injected subcutaneously for 4 consecutive days. After 28 days, induction of HF was evaluated by echocardiography. A day after echocardiography, 50 µg culture medium/5 ml PBS in HF+culture medium group, 50 µg MSC-CM/5 ml PBS in HF+MSC-CM group and 5 ml PBS in HF+PBS group were injected two times for 4 successive days. The echocardiography was performed 4 weeks after the last injection of isoproterenol. To evaluate the fibrosis, morphology, and cardiac function, Trichrome Masson's staining, Hematoxylin and Eosin staining and echocardiography were performed, respectively. RESULTS: CM significantly increased fractional shortening and ejection fraction, and also significantly decreased apoptotic nuclear condensation. Moreover, significant decreased level of fibrosis and increased level of angiogenesis was observed in the treatment group (P<0.05). CONCLUSION: Our results indicated that IV injection of CM has therapeutic effects on HF by reducing fibrosis and preventing the progression of failure due to its paracrine effects.