Long and Short-term Metformin Consumption as a Potential Therapy to Prevent Complications of COVID-19.

Purpose: The aim of the study is to evaluate the effect of metformin in complication improvement of hospitalized patients with COVID-19. Methods: This was a randomized clinical trial that involved 189 patients with confirmed COVID-19 infection. Patients in the intervention group received metformin-500 mg twice daily. Patients who received metformin before admission were excluded from the control group. Patients who were discharged before taking at least 2000 mg of metformin were excluded from the study. Primary outcomes were vital signs, need for ICU admission, need for intubation, and mortality. Results: Data showed that patients with diabetes with previous metformin in their regimen had lower percentages of ICU admission and death in comparison with patients without diabetes (11.3% vs. 26.1% (P=0.014) and 4.9% vs. 23.9% (P≤0.001), respectively). Admission time characteristics were the same for both groups except for diabetes and hyperlipidemia, which were significantly different between the two groups. Observations of naproxen consumption on endpoints, duration of hospitalization, and the levels of spO2 did not show any significant differences between the intervention and the control group. The adjusted OR for intubation in the intervention group versus the control group was 0.21 [95% CI, 0.04-0.99 (P=0.047)]. Conclusion: In this trial, metformin consumption had no effect on mortality and ICU admission rates in non-diabetic patients. However, metformin improved COVID-19 complications in diabetic patients who had been receiving metformin prior to COVID-19 infection, and it significantly lowered the intubation rates.

Time-dependent influence of infliximab on hemodynamic responses and cardiac injuries of isoproterenol-induced myocardial infarction in rats.

Immune-induced inflammation plays an important role both in aggravating and healing of post myocardial infarction (MI) injuries. Potent anti-inflammatory and local immunomodulatory activity of infliximab has been suggested to have modulating effects on immune responses after MI. The aim of the present study was to evaluate the efficacy of infliximab on hemodynamic responses and myocardial injuries following isoproterenol-induced myocardial infarction. Male Wistar rats, weighting 260 ± 20 g were assigned into ten groups (n = 6) of saline (normal saline), infliximab (7 mg/kg), isoproterenol (100 mg/kg for two consecutive days), and isoproterenol plus infliximab (30 min after the second injection of isoproterenol). The heart tissues and serums were analyzed 24, 48, 72, and 96 h post-MI and hemodynamic parameters, histopathological changes, malondialdehyde (MDA), Total antioxidant capacity (TAC), lactate dehydrogenase (LDH), and lactate levels were assessed in the respective groups. Infliximab partially improved hemodynamic depression in the first days after MI, but the heart became more suppressed later. A similar result also obtained at the MDA tissue levels but not serum levels. Anti-inflammatory effects of Infliximab may improve cardiac function and prevent heart tissue injury early after MI; however, it can worsen the condition later by inhibiting compensatory reactions such as cardiac remodeling and tissue repair.

Oxidized cholesterol exacerbates toll-like receptor 4 expression and activity in the hearts of rats with myocardial infarction.

Introduction: The present study examined the effects of high cholesterol and high oxidized-cholesterol diets on the myocardial expression of TLR4 and pro-inflammatory cytokine in rats. Methods: Male Wistar rats were allocated into 6 groups and fed with a normal diet, cholesterol, and oxidized-cholesterol rich diets with or without isoproterenol-induced myocardial infarction. TLR4 and MyD 88 expression and levels tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) were measured in the heart and serum. Results: Oxidized cholesterol-fed animals had higher serum levels of oxidized low-density lipoprotein (LDL) (263 ± 13 ng/dL) than the cholesterol-fed animals (98 ± 8 ng/dL; P < 0.001). A high level of oxidized-LDL caused fibrotic cell formation and enhanced neutrophil infiltration in the absence of MI. Both cholesterol and oxidized-cholesterol upregulated TLR4 mRNA expression and increased TNF-α and IL-6 production in the hearts of rats with MI. In rats fed with oxidized-cholesterol the serum and myocardial levels of TNF-α (653 ± 42 pg/mL, 1375 ± 121 pg/100 mg, respectively) were higher than MI group (358±24 pg/mL, P < 0.001 and 885 ± 56 pg/100 mg, P < 0.01). A significant correlation was seen between TLR4 expression and infarct size. Conclusion: These findings suggest that cardiac TLR4 is preferentially upregulated by oxidized cholesterol in rats. Oxidized cholesterol may have a critical role in cardiac toxicity in the absence of pathological conditions.

Clinical application of immune checkpoints in targeted immunotherapy of prostate cancer.

Immunotherapy is considered as an effective method for cancer treatment owing to the induction of specific and long-lasting anti-cancer effects. Immunotherapeutic strategies have shown significant success in human malignancies, particularly in prostate cancer (PCa), a major global health issue regarding its high metastatic rates. In fact, the first cancer vaccine approved by FDA was Provenge, which has been successfully used for treatment of PCa. Despite the remarkable success of cancer immunotherapy in PCa, many of the developed immunotherapy methods show poor therapeutic outcomes. Immunosuppression in tumor microenvironment (TME) induced by non-functional T cells (CD4+ and CD8+), tolerogenic dendritic cells (DCs), and regulatory T cells, has been reported to be the main obstacle to the effectiveness of anti-tumor immune responses induced by an immunotherapy method. The present review particularly focuses on the latest findings of the immune checkpoints (ICPs), including CTLA-4, PD-1, PD-L1, LAG-3, OX40, B7-H3, 4-1BB, VISTA, TIM-3, and ICOS; these checkpoints are able to have immune modulatory effects on the TME of PCa. This paper further discusses different approaches in ICPs targeting therapy and summarizes the latest advances in the clinical application of ICP-targeted therapy as monotherapy or in combination with other cancer therapy modalities in PCa.

STAT3 inhibitory stattic enhances immunogenic cell death induced by chemotherapy in cancer cells.

BACKGROUND: Induction of immunogenic cell death (ICD) is considered a promising strategy for cancer immunotherapy. Stattic is an inhibitor of STAT3, which is found constitutively active in many cancers and plays a major role in cancer progression. OBJECTIVES: In the present study, we proposed to evaluate whether stattic can enhance the effects of chemotherapy in the induction of ICD in cancer cells harboring hyperactive STAT3. METHODS: The growth inhibitory effects of stattic and chemo agents including doxorubicin (DOX) and oxaliplatin (OXP) were evaluated using MTT assay in B16F10 and CT26 cell lines. Flow cytometry was applied to study cell apoptosis and calreticulin (CRT) surface exposure. The levels of high mobility group box 1 (HGMB1), heat shock protein70 (HSP70) and interleukin-12 (IL-12) were measured using ELISA. RESULTS: Treatment of B16F10 and CT26 cells with stattic in combination with DOX resulted in synergistic antitumor effects with combination index being 0.82 and 0.87, respectively. Interestingly, we found a higher level of ICD markers including CRT expression as well as HMGB1 and HSP70 secretion in the cells received combination therapy of stattic and DOX as compared with monotherapies. Moreover, exposure of dendritic cells (DCs) to conditioned media (CM) from cancer cells treated with stattic and/or DOX resulted in secretion of IL-12, which is an indicator of DCs maturation and induction of Th1 response. OXP and stattic monotherapy induced ICD in CT26 cells and stimulated IL-12 secretion by DCs; however, we did not observe a significant increase in the level of ICD in CT26 cells and IL-12 secretion by DCs when CT26 cells were treated with stattic and OXP combination as compared with monotherapy groups. CONCLUSION: These findings indicate that STAT3 inhibitory stattic can increase ICD induced by DOX. Graphical abstract.

Copper sulfate pentahydrate reduced epithelial cytotoxicity induced by lipopolysaccharide from enterogenic bacteria.

The over usage of multiple antibiotics contributes to the emergence of a whole range of antibiotic-resistant strains of bacteria causing enterogenic infections in poultry science. Therefore, finding an appropriate alternative natural substance carrying an antibacterial capacity would be immensely beneficial. It has been previously discovered that the different types of cupric salts, especially copper sulfate pentahydrate (CuSO4·5H2O), to carry a potent bactericidal capacity. We investigated the neutralizing effect of CuSO4·5H2O (6.25µg/ml) on the reactive oxygen species generation, and expression of MyD88, an essential adaptor protein of Toll-like receptor, and NF-κB in three intestinal epithelial cell lines exposed to 50ng/ml lipopolysaccharide. In order to find the optimal cupric sulfate concentration without enteritis-inducing toxicity, broiler chickens were initially fed with water containing 0.4, 0.5, and 1mg/l during a period of 4days. After determination of appropriate dosage, two broiler chickens and turkey flocks with enteritis were fed with cupric compound for 4days. We found that cupric sulfate can lessen the cytotoxic effect of lipopolysaccharide by reducing the reactive oxygen species content (p<0.05). Additionally, the expression of MyD88 and NF-κB was remarkably down-regulated in the presence of lipopolysaccharide and cupric sulfate. The copper sulfate in doses lower than 0.4mg/ml expressed no cytotoxic effect on the liver, kidney, and the intestinal tract while a concentration of 0.5 and 1mg/ml contributed to a moderate to severe tissue injuries. Pearson Chi-Square analysis revealed the copper cation significantly diminished the rate of mortality during 4-day feeding of broiler chicken and turkey with enteritis (p=0.000). Thus, the results briefed above all confirm the potent anti-bactericidal feature of cupric sulfate during the course of enteritis.

Endothelial cells' biophysical, biochemical, and chromosomal aberrancies in high-glucose condition within the diabetic range.

To date, many studies have been conducted to find out the underlying mechanisms of hyperglycemia-induced complications in diabetes mellitus, attributed to the cellular pathologies of different cells-especially endothelial cells. However, there are still many ambiguities and unresolved issues to be clarified. Here, we investigated the alteration in biophysical and biochemical properties in human umbilical vein endothelial cells exposed to a high-glucose concentration (30mM), comparable to glucose content in type 2 diabetes mellitus, over a course of 120 hours. In addition to a reduction in the rate of cell viability and induction of oxidative stress orchestrated by the high-glucose condition, the dynamic of the fatty acid profile-including polyunsaturated, monounsaturated, and saturated fatty acids-was also altered in favor of saturated fatty acids. Genetic imbalances were also detected at chromosomal level in the cells exposed to the abnormal concentration of glucose after 120 hours. Moreover, the number of tip cells (CD31+ /CD34+ ) and in vitro tubulogenesis capability negatively diminished in comparison to parallel control groups. We found that diabetic hyperglycemia was associated with a decrease in the cell-cell tight junction and upregulation in vascular endothelial cadherin and zonula occludens (ZO)-1 molecules after 72 and 120 hours of exposure to the abnormal glucose concentration, which resulted in a profound reduction in transendothelial electrical resistance. The surface plasmon resonance analysis of the human umbilical vein endothelial cells immobilized on gold-coated sensor chips confirmed the loosening of the cell to cell intercellular junction as well as stable attachment of each cell to the basal surface. Our findings highlighted the disturbing effects of a diabetic hyperglycemia on either biochemical or biophysical properties of endothelial cells.

Methylprednisolone acetate-loaded hydroxyapatite nanoparticles as a potential drug delivery system for treatment of rheumatoid arthritis: In vitro and in vivo evaluations.

The objective of this study was to improve the therapeutic efficacy of methylprednisolone acetate (MPA) in the treatment of rheumatoid arthritis (RA) by incorporating the drug into the hydroxyapatite (HAp) nanoparticles. The nanoparticles were synthesized using a chemical precipitation technique and their size and morphology were evaluated by dynamic light scattering and scanning electron microscopy (SEM). The solid-state behavior of the nanoparticles was also characterized by operating X-ray powder diffraction (XRPD), differential scanning calorimetry (DSC) and Fourier-transform infrared spectroscopy (FTIR). The Brunauer-Emmett-Teller and Barrett-Joyner-Halenda N2 adsorption/desorption analyses were also performed to determine the surface area, Vm (the volume of the N2 adsorbed on the one gram of the HAp when the monolayer is complete) and the pore size of the samples. Furthermore, the therapeutic efficacy of the prepared nanoformulation on the adjuvant induced arthritic rats was assessed. HAp mesoporous nanoparticles with a particle size of 70.45nm, pore size of 2.71nm and drug loading of 44.53% were obtained. The specific surface area of HAp as well as the Vm values were decreased after the drug loading process. The nanoformulation revealed the slower drug release profile compared to the pure drug. The MTT assay indicated that the MPA-loaded nanoparticles had a lower cytotoxic effect on NIH-3T3 and CAOV-4 cell lines compared to the pure drug. Interestingly, the in vivo study confirmed that the drug-loaded nanoparticles could considerably decrease the paw volume and normalize the hematological abnormalities in the arthritic rats.

A-769662, a direct AMPK activator, attenuates lipopolysaccharide-induced acute heart and lung inflammation in rats.

Activation of AMP­activated protein kinase (AMPK) has been indicated to produce an anti­inflammatory effect through the suppression of toll­like receptor (TLR) activity. In the present study, the investigation was designed to identify the effect of A­769662, a direct activator of AMPK on lipopolysaccharide (LPS)­induced acute lung and heart inflammation in rats. To induce inflammation, an intraperitoneal injection of LPS (0.5 mg/kg) was administered to Wistar rats. The inflammatory parameters and AMPK phosphorylation were then measured 9 h later. For the treatment group, A­769662 (10 mg/kg) was administrated intraperitoneally immediately prior to LPS injection. The results demonstrated that A­769662 attenuated the LPS­induced acute inflammation in the heart and lung tissue, as indicated by the significant reduction in myeloperoxidase activity (P<0.001) and inhibition of tissue damage. This was associated with a significant reduction in tumor necrosis factor­α serum levels (P<0.01) and peripheral neutrophils (P<0.001). Furthermore, A­769662 enhanced AMPK phosphorylation and downregulated the expression of MyD88, a TLR adaptor protein, in the heart tissue. Despite the anti­inflammatory effect of A­769662 on LPS­induced inflammation in the lung tissue, the drug produced no effect on the MyD88 expression levels or AMPK phosphorylation in the tissue. The results of the present study suggested that the administration of A­769662 results in an anti­inflammatory effect in the LPS­induced model of inflammation in rats. The anti­inflammatory activity was demonstrated in the heart and lung tissues and the effect on the cardiac tissue was indicated to be a result of AMPK activation, involving the suppression of TLRs.

Physicochemical characterization and in vivo evaluation of triamcinolone acetonide-loaded hydroxyapatite nanocomposites for treatment of rheumatoid arthritis.

The current study was aimed to investigate the anti-inflammatory effect of triamcinolone acetonide-loaded hydroxyapatite (TA-loaded HAp) nanocomposites in the arthritic rat model. The HAp nanocomposites were synthesized through a chemical precipitation method and the drug was subsequently incorporated into the nanocomposites using an impregnation method. The physicochemical properties as well as cytotoxicity of the prepared nanoformulation were examined as well. To evaluate the therapeutic efficacy of the prepared nanoformulation, the various parameters such as paw volume, haematological parameters and histological studies were assessed in the arthritic rats. The nanocomposites with the particle size of 70.45 nm, pore size of 2.71 nm and drug loading of 41.94% were obtained in this study. The specific surface area (aBET) as well as the volume of nitrogen adsorbed on one gram of HAp to complete the monolayer adsorption (Vm) were decreased after the drug loading process. The prepared nanoformulation revealed the slower drug release profile compared to the pure drug. Furthermore, the obtained data from MTT assay showed that the TA-loaded nanocomposites had a lower cytotoxic effect on NIH-3T3 and CAOV-4 cell lines as compared to the pure drug. Furthermore, TA-loaded HAp nanocomposites demonstrated favorable effects on the paw volume as well as the haematological and histopathological abnormalities in the adjuvant-induced arthritic rats. Therefore, TA-loaded HAp nanocomposites are potentially suggested for treatment of rheumatoid arthritis after further required evaluations.

Effect of A-769662, a direct AMPK activator, on Tlr-4 expression and activity in mice heart tissue.

OBJECTIVES: TLR-4 activates a number of inflammatory signaling pathways. Also, AMPK could be involved in anti-inflammatory signaling. The aim of this study was to identify whether stimulation of AMPK could inhibit LPS-induced Tlr-4 gene expression in mice hearts. MATERIALS AND METHODS: Heart AMPK activity and/or Tlr-4 expression was stimulated in different mice groups, using respectively IP injection of A-769662 (10 mg/kg) and LPS (2 mg/kg) or a combination of both agents. Moreover, compound-C (20 mg/kg), as an AMPK antagonist, was intraperitoneally co-administrated with both A-769662 and LPS in another group to investigate the role of AMPK activity on Tlr-4 regulation. After 8 hr, in addition to peripheral neutrophil cell count, myocardial p-AMPK, p-ACC as well as MyD88 protein contents and Tlr-4 expression was assessed by Western blotting and real-time qRT-PCR, respectively. TNF-α and IL-6 expression levels were also determined by ELISA. RESULTS: LPS induced heart Tlr-4 expression (P<0.001) associating with an increase in the myocardial MyD88 protein content (P<0.001), elevation of heart TNF-α (P<0.01) and IL-6 (P<0.05) concentrations, and rise in the peripheral neutrophil cell count (P<0.001). Administration of A-769662 decreased LPS-induced Tlr-4 expression (P<0.01) and alleviated peripheral neutrophil cell count (P<0.01). The inhibitory effect of A-769662 on LPS-induced Tlr-4 expression was reversed by antagonizing AMPK with compound-C (P<0.001) which reduced p-AMPK (P<0.05) and p-ACC (P<0.01) myocardial protein contents in the LPS+A-769662 group. CONCLUSION: This study demonstrated that activation of AMPK, by A-769662 agent, could inhibit Tlr-4 expression and activity, suggesting a link between AMPK and Tlr-4 in heart tissue.

Lipopolysaccharide Induced Activation of Toll Like Receptor 4 in Isolated Rat Heart Suggests a Local Immune Response in Myocardium.

BACKGROUND: Myocardial dysfunction is one of the major complications in patients with sepsis where there is a relationship between the blood level of cytokines and the onset of myocardial depression. In many cases of sepsis, the presence of Lipopolysaccharide (LPS) has been established. LPS Binding Protein (LBP) bound endotoxin is recognized by CD14/toll-like receptor-4 (TLR4) complexes in innate immune cells which stimulates TNF-α release. OBJECTIVES: To investigate whether isolated rat heart is capable of producing TNF-α locally through TLR4 activation by LPS. METHODS: Using langendorff method, LPS in 120 mL of the modified Krebs-Henseleit buffer solution (KHBS) at final concentration of 1 µg/mL was perfused at recycling mode. The effect of LPS on cardiac function was evaluated. To assess the TLR4 activity and TNF-α release, western blotting, real time-PCR, and ELISA were used. RESULTS: Compared with control, coronary perfusion pressure (CPP) as well as left ventricular developed pressure (LVDP), maximum and minimum rates of the left ventricular developed pressure (dP/dtmax; dP/dtmin; p<0.001) were depressed to a maximum level after 180 minutes recycling with LPS. This myocardial depression was associated with a significant increase in TLR4 expression (p<0.01), MyD88 activity (p<0.01) and TNF-α (p<0.05) concentration in the heart tissue. CONCLUSION: The results of this study show that heart is capable of producing TNF-α through TLR4 and MyD88 activation independent of classic immune system and suggest a local immune response.

Tacrolimus ameliorates functional disturbances and oxidative stress in isoproterenol-induced myocardial infarction.

BACKGROUND: The inflammatory responses play a major role in the pathogenesis of acute myocardial infarction (MI). Early inhibition of inflammation may improve post MI cardiac function. The aim of this study was to investigate the effects of tacrolimus on cardiac function, hemodynamic parameters as well as histopathologic and electrocardiographic changes in isoproterenol-induced myocardial infarction. METHODS: Male Wistar rats were randomly divided into six groups of control, isoproterenol alone, tacrolimus alone, and isoproterenol plus tacrolimus (0.5, 1 and 2 mg/kg). Isoproterenol (100 mg/kg) was injected subcutaneously for two consecutive days to induce myocardial infarction, and simultaneously tacrolimus was administered orally twice a day for three days. RESULTS AND CONCLUSIONS: Administration of isoproterenol resulted in myocardial edema and necrosis as well as a marked reduction in the left ventricular systolic pressure (LVSP), left ventricular contractility (LVdP/dtmax) and relaxation (LVdP/dtmin) along with a severe elevation in left ventricular end-diastolic pressure (LVEDP). Isoproterenol also elevated the ST-segment and suppressed the R-amplitude and R-R interval on ECG. It was found that all doses of tacrolimus could amend the ECG pattern and ameliorated the isoproterenol induced disturbances in cardiac function. Acute and short term treatment with tacrolimus at dose of 2 mg/kg significantly (P < 0.001) improved LVdP/dtmax from 2712 ± 82 in myocardial infarcted rats to 4592 ± 149 mmHg/sec. Similarly, tacrolimus lowered LVEDP from 17.6 ± 0.68 in MI group to the value of 5.6 ± 0.22 mmHg (P < 0.001). Furthermore, tacrolimus was found to reduce malondialdehyde concentration in serum and myocardium by 50-70% (P < 0.001).

Pioglitazone prevents morphine antinociception tolerance and withdrawal symptoms in rats.

Long-term exposure to opiates induces tolerance to the analgesic effect and dependence. The purpose of the present study is to investigate the effects of pioglitazone, a peroxisome proliferator-activated receptors gamma (PPAR-γ) agonist, on the morphine-induced tolerance and dependence. Groups of rats received morphine in combination with a vehicle or pioglitazone (5, 10, 20, and 40 mg/kg) daily. Thirty minutes before pioglitazone (40 mg/kg), GW-9662, a selective PPAR-γ antagonist, (2 mg/kg) was administrated in order to evaluate the possible role of the PPAR-γ. Nociception was assessed by a tail flick apparatus, and the percentage of the maximal possible effect was calculated as well. For 9 days, rats received additive doses of morphine to induce dependence. Naloxone was administrated 2 h after the morphine last dose, and withdrawal symptoms were recorded for 45 min. Morphine administration to rats over a duration of 17 days resulted in the development of tolerance, whereas pioglitazone (40 mg/kg) delayed the day of the established tolerance for 15 days. Administration of pioglitazone also prevented morphine-induced 50 % effective dose (ED50) shift to the right in the dose-response curve and increased the global analgesic effect of morphine. In addition, pioglitazone decreased the total withdrawal score significantly, whereas GW-9662 significantly reversed the pioglitazone effects on the morphine tolerance and dependence. The prevention of the morphine-induced glia activation and the proinflammatory responses were the possible mechanisms for pioglitazone effect on delaying the morphine tolerance and attenuating the dependence.

Chronic treatment with metformin suppresses toll-like receptor 4 signaling and attenuates left ventricular dysfunction following myocardial infarction.

Acute treatment with metformin has a protective effect in myocardial infarction by suppression of inflammatory responses due to activation of AMP-activated protein kinase (AMPK). In the present study, the effect of chronic pre-treatment with metformin on cardiac dysfunction and toll-like receptor 4 (TLR4) activities following myocardial infarction and their relation with AMPK were assessed. Male Wistar rats were randomly assigned to one of 5 groups (n=6): normal control and groups were injected isoproterenol after chronic pre-treatment with 0, 25, 50, or 100mg/kg of metformin twice daily for 14 days. Isoproterenol (100mg/kg) was injected subcutaneously on the 13th and 14th days to induce acute myocardial infarction. Isoproterenol alone decreased left ventricular systolic pressure and myocardial contractility indexed as LVdp/dtmax and LVdp/dtmin. The left ventricular dysfunction was significantly lower in the groups treated with 25 and 50mg/kg of metformin. Metfromin markedly lowered isoproterenol-induced elevation in the levels of TLR4 mRNA, myeloid differentiation protein 88 (MyD88), tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6) in the heart tissues. Similar changes were also seen in the serum levels of TNF-α and IL-6. However, the lower doses of 25 and 50mg/kg were more effective than 100mg/kg. Phosphorylated AMPKα (p-AMPK) in the myocardium was significantly elevated by 25mg/kg of metformin, slightly by 50mg/kg, but not by 100mg/kg. Chronic pre-treatment with metformin reduces post-myocardial infarction cardiac dysfunction and suppresses inflammatory responses, possibly through inhibition of TLR4 activities. This mechanism can be considered as a target to protect infarcted myocardium.

Marrubium vulgare L. methanolic extract inhibits inflammatory response and prevents cardiomyocyte fibrosis in isoproterenol-induced acute myocardial infarction in rats.

INTRODUCTION: Nowadays, finding new therapeutic compounds from natural products for treatment and prevention of a variety of diseases including cardiovascular disorders is getting a great deal of attention. This approach would result in finding new drugs which are more effective and have fewer side effects than the conventional medicines. The present study was designed to investigate the anti-inflammatory effect of the methanolic extract of Marrubiumvulgare, a popular traditional medicinal herb, on isoproterenol-induced myocardial infarction (MI) in rat model. METHODS: Male Wistar rats were assigned to 6 groups of control, sham, isoproterenol, and treatment with 10, 20, and 40 mg/kg/12h of the extract given orally concurrent with MI induction. A subcutaneous injection of isoproterenol (100 mg/kg/day) for two consecutive days was used to induce MI. Then, histopathological changes and inflammatory markers were evaluated. RESULTS: Isoproterenol injection increased inflammatory response, as shown by a significant increase in peripheral neutrophil count, myocardial myeloperoxidase (MPO) activity and serum levels of creatinine kinase-MB (CK-MB) and TNF-α (p<0.001). In the groups treated with 10, 20 and 40 mg/kg of M.vulgare extract serum CK-MB was subsided by 55.4%, 52.2% and 69%, respectively. Also treatment with the extract (40 mg/kg) significantly reduced (p<0.001) MPO activity in MI group. The levels of TNF-α was also considerably declined in the serums of MI group (p<0.001). In addition, peripheral neutrophil count, was significantly lowered by all doses of the extract (p<0.001). Interstitial fibrosis significantly was attenuated in treated groups compared with control MI group. CONCLUSION: The results of study demonstrate that the M. vulgare extract has strong protective effects against isoproterenol-induced myocardial infarction and it seems possible that this protection is due to its anti-inflammatory effects.

Selenium effect on oxidative stress factors in septic rats.

PURPOSE: Severe oxidative stress is an important event that occurs in patients with sepsis. The body has extensive and multiple defense mechanisms against the reactive oxygen species (ROS) produced during inflammation and sepsis. One of these mechanisms includes a group of enzymes that utilize selenium as their cofactor. The purpose of this study is investigating of Selenium effect on oxidative stress factors in animal model of sepsis. METHODS: Sepsis was induced by caecal ligation and puncture (CLP) method. 30 Male Wistar rats were divided into following groups: sham group; CLP group; 100 µg/kg Selenium- treated CLP group. 12 hours after inducing sepsis animals were killed and lungs were removed. One of the lungs was frozen in liquid nitrogen and kept at -70°C for enzymatic activity analysis and the other was kept in formalin 10% until tissue section preparation performed for histopathological studies. RESULTS: The Myeloperoxidase (MPO) activity was decreased in Selenium- treated CLP group. Inflammation score of lung tissue was lowered in Selenium- treated CLP group, but it wasn't statically significant. Level of glutathione peroxidase (GPx) was higher in CLP and Selenium- treated CLP groups. CONCLUSION: It seems that Selenium has protective effect on lung inflammation during acute lung injury. Also it may improve some stress oxidative profile during CLP model of sepsis.

Cardioprotective effect of methanolic extract of Marrubium vulgare L. on isoproterenol-induced acute myocardial infarction in rats.

Isoproterenol injection (100 mg/kg; sc) produced changes in ECG pattern including ST-segment elevation and suppressed R-amplitude. The methanolic extract of M. vulgare at doses of 10, 20, and 40 mg/kg significantly amended the ECG changes. A severe myocardial necrosis and edematous along with a sharp reduction in the arterial blood pressure, left ventricular contractility (LVdP/dt(max or min)), but a marked increase in the left ventricular end-diastolic pressure (LVEDP) were seen in the isoproterenol group. All parameters were significantly improved by the extract treatment. The extract (10 mg/kg) strongly increased LVdP/dt(max). Similarly, treatment with 40 mg/kg of M. vulgare lowered the elevated LVEDP and the heart to body weight ratio. In addition to in vitro antioxidant activity, the extract suppressed markedly the elevation of malondialdehyde levels both in serum and in myocardium. The results demonstrate that M. vulgare protects myocardium against isoproterenol-induced acute myocardial infarction and suggest that the effects could be related to antioxidant activities.

Protective effects of methylsulfonylmethane on hemodynamics and oxidative stress in monocrotaline-induced pulmonary hypertensive rats.

Methylsulfonylmethane (MSM) is naturally occurring organic sulfur that is known as a potent antioxidant/anti-inflammatory compound. The aim of this study was to investigate the effect of MSM on hemodynamics functions and oxidative stress in rats with monocrotaline- (MCT-) induced pulmonary arterial hypertension (PAH). Wistar rats were randomly assigned to 38-days treatment. MSM was administered to rats at 100, 200, and 400 mg/kg/day doses 10 days before a single dose of 60 mg/kg, IP, MCT. Hemodynamics of ventricles were determined by Powerlab AD instrument. Blood samples were obtained to evaluate changes in the antioxidative system including activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and the level of reduced glutathione (GSH) and malondialdehyde (MDA). Improvements in cardiopulmonary hemodynamics were observed in the MSM-treated pulmonary arterial hypertensive rats, with a significant reduction in right ventricular systolic pressure (RSVP) and an increase in the mean arterial pressure (MAP). The values of CAT, SOD, GSH-px activities, and GSH were significantly lower in MCT-induced PAH (P < 0.01), but they were recovered to control levels of MSM-treated groups. Our present results suggest that long-term administration of the MSM attenuates MCT-induced PAH in rats through modulation of oxidative stress and antioxidant defense.

Short-term treatment with metformin suppresses toll like receptors (TLRs) activity in isoproterenol-induced myocardial infarction in rat: are AMPK and TLRs connected?

AMP-activated protein kinase (AMPK) is a key sensor of cellular energy. The activation of AMPK by metformin prevents cardiac remodeling after myocardial infarction (MI). Besides, the innate immune response through TLRs is activated during MI. In the present study, the effects of short-term treatment with metformin on TLRs activity and its relation with AMPK in isoproterenol-induced MI were assessed in rats. To induce MI, a subcutaneous injection of isoproterenol was given to Wistar rats for two consecutive days. Metformin (25, 50, and 100mg/kg) was orally administered to rats twice daily for two days. Interstitial fibrosis was dose-dependently attenuated in the treated groups in comparison to the MI group (score: 1.25 ± 0.28 with 100 mg/kg metformin versus 3.5 ± 0.28; P<0.001). Further, metformin reduced TLR-dependent inflammatory cytokines as indexed by reduced myocardial levels of TNFα (maximum 68%; P<0.001) and IL6 (maximum 84%; P<0.001) as well as by reduced myocardial MPO activity (25%; P<0.01). It was found that the level of phosphorylated AMPK was significantly upregulated by 165% (P<0.001) when treated with 100 mg/kg of metformin, but not with 25 and 50mg/kg. This was associated with a remarkable suppression of TLR4 expression and reduction of protein level of TLRs adapter protein, MyD88 (P<0.01) in the infarcted myocardium. These results suggest that AMPK activation by metformin and the subsequent suppression of TLRs activity could be considered as a target in protecting the infarcted heart, which may indicate a link between AMPK and TLRs.