Inhibition of tyrosine kinase signaling by tyrphostin AG126 downregulates the IL-21/IL-21R and JAK/STAT pathway in the BTBR mouse model of autism.

Autism spectrum disorder (ASD) comprises a broad range of neurodevelopmental disorders that are associated with deficits in social interaction and communication. The tyrosine kinase inhibitor tyrphostin AG126 represents a promising therapeutic agent for several neuroinflammatory disorders. There are currently no treatments available that can improve ASD and we previously showed that AG126 treatment exerts beneficial effects on BTBR T+ Itpr3tf/J (BTBR) mice, a model for autism that shows the core features of ASD; however, the immunological mechanisms and molecular targets associated with this effect were previously unclear. This study was undertaken to delineate the neuroprotective effect of AG126 on BTBR mice. Here, using this mouse model, we investigated the effects of AG126 administration on IL-21R, IL-21, IL-22, TNF-α, NOS2, STAT3, IL-27, and Foxp3 production by CD8+ T cells in the spleen by flow cytometry. We further explored the mRNA and protein expression of IL-21, IL-22, IL-1ß, TNF-α, NOS2, JAK1, STAT3, IL-27, and Foxp3 in brain tissue by RT-PCR, and western blotting. We found that BTBR mice treated with AG126 exhibited significant decreases in IL-21R-, IL-21-, IL-22-, TNF-α-, NOS2-, STAT3-producing, and increases in IL-27- and Foxp3-producing, CD8+ T cells. Our results further demonstrated that AG126 treatment effectively decreased IL-21, IL-22, IL-1ß, TNF-α, NOS2, JAK1, and STAT3, and increased IL-27 and Foxp3 mRNA and protein expression in brain tissues. Our findings suggest that AG126 elicits a neuroprotective response through downregulation of the IL-21/IL-21R and JAK/STAT pathway in BTBR mice, which could represent a promising novel therapeutic target for ASD treatment.

Dysregulation of the expression of HLA-DR, costimulatory molecule, and chemokine receptors on immune cells in children with autism.

Autism spectrum disorder (ASD) is a heterogeneous disorder diagnosed based on the severity of abnormalities in social skills. Several studies have acknowledged the presence of abnormal immune functions among individuals diagnosed with ASD. HLA-DR (human leukocyte antigen-antigen D related) has been shown to play a significant role in several inflammatory and neurological disorders; however, the role of HLA-DR signaling in ASD has not yet been fully clarified. In this study, we investigated the role of HLA-DR signaling in children with ASD. Flow cytometric analysis, using peripheral blood mononuclear cells (PBMCs), revealed the numbers of CD4+, CD8+, CD28+, CXCR4+, and CCR7+ expressing HLA-DR cells in typically developing (TD) controls and children with ASD. We also determined the numbers of IFN-γ+, IL-21+, and Foxp3+ expressing HLA-DR cells in TD controls and in children with ASD using PBMCs. We observed mRNA and protein expression levels of HLA-DR by RT-PCR and western blotting analysis. Our results revealed that children with ASD had significantly increased numbers of HLA-DR+CD4+, HLA-DR+CD8+, CD28+HLA-DR+, HLA-DR+CXCR4+, HLA-DR+CCR7+ cells compared with TD controls. We found that children with ASD showed increased HLA-DR+IFN-γ+ and HLA-DR+IL-21+ and decreased HLA-DR+Foxp3+ expression levels compared with TD controls. Furthermore, children with ASD showed higher HLA-DR mRNA and protein expression levels compared with TD controls. These results indicated that HLA-DR could play an essential role in the immune abnormalities associated with ASD.

Therapeutic potential of carfilzomib, an irreversible proteasome inhibitor, against acetaminophen-induced hepatotoxicity in mice.

Overdose of acetaminophen (APAP) is often associated with hepatotoxicity. Carfilzomib (CFZ) shows multiple pharmacological activities including anti-inflammatory potential. Therefore, this study was undertaken to evaluate the possible therapeutic effects of CFZ against APAP-induced hepatotoxicity. Hepatotoxicity was induced by administration of APAP (350 mg/kg, intraperitoneal). Mice were given CFZ (0.125, 0.25, or 0.5 mg/kg, intraperitoneal) 1.5 h after APAP administration. Animals were sacrificed on 6 h and blood and liver tissue samples were collected for analysis. In CFZ-post-treated group, there was significant and dose-dependent decrease in serum alanine aminotransferase levels. The level of tumor necrosis factor-α (TNF-α), reactive oxygen species, and NO decreased, whereas glutathione increased significantly by CFZ post-treatment. Upregulated mRNA expression of COX-II and iNOS were significantly downregulated by CFZ post-treatment. CFZ may exert its hepatoprotective action by alleviating inflammatory, oxidative, and nitrosative stress via inhibition of TNF-α, COX-II, and iNOS.

Alleviation of Aflatoxin B1-Induced Genomic Damage by Proanthocyanidins via Modulation of DNA Repair.

In order to study the mechanisms underlying the alleviation of aflatoxin B1-induced genomic damage by proanthocyanidins (PAs), we examined the modulation of oxidative DNA damage induced by aflatoxin B1 in PAs-pretreated animals. The effects of PAs on changes in the expression of DNA damage and repair genes induced by aflatoxin B1 were also evaluated in rat marrow cells. Administration of PAs before aflatoxin B1 significantly mitigated aflatoxin B1-induced oxidative DNA damage in a dose-dependent manner. Aflatoxin B1 treatment induced significant alterations in the expression of specific DNA repair genes, and the pre-treatment of rats with PAs ameliorated the altered expression of these genes. Conclusively, PAs protect against aflatoxin B1-induced oxidative DNA damage in rats. These protective effects are attributed to the antioxidant effects of PA and enhanced DNA repair through modulation of DNA repair gene expression. Therefore, PAs are a promising chemoprotective agent for averting genotoxic risks associated with aflatoxin B1 exposure.

Airway oxidative stress causes vascular and hepatic inflammation via upregulation of IL-17A in a murine model of allergic asthma.

Oxidants are generated in asthmatic airways due to infiltration of inflammatory leukocytes and resident cells in the lung. Reactive oxygen species (ROS) such as hydrogen peroxide and superoxide radical may leak into systemic circulation when generated in uncontrolled manner and may impact vasculature. Our previous studies have shown an association between airway inflammation and systemic inflammation; however so far none has investigated the impact of airway oxidative inflammation on hepatic oxidative stress and Th1/Th2/Th17 cytokine markers in liver/vasculature in a murine model of asthma. Therefore, this study investigated the contribution of oxidative stress encountered in asthmatic airways in modulation of systemic/hepatic Th1/Th2/Th17 cytokines balance and hepatic oxidative stress. Mice were sensitized intraperitoneally with cockroach extract (CE) in the presence of aluminum hydroxide followed by several intranasal (i.n.) challenges with CE. Mice were then assessed for systemic/hepatic inflammation through assessment of Th1/Th2/Th17 cytokines and oxidative stress (iNOS, protein nitrotyrosine, lipid peroxides and myeloperoxidase activity). Challenge with CE led to increased Th2/Th17 cytokines in blood/liver and hepatic oxidative stress. However, only Th17 related pro-inflammatory markers were upregulated by hydrogen peroxide (H2O2) inhalation in vasculature and liver, whereas antioxidant treatment, N-acetyl cysteine (NAC) downregulated them. Hepatic oxidative stress was also upregulated by H2O2 inhalation, whereas NAC attenuated it. Therefore, our study shows that airway oxidative inflammation may contribute to systemic inflammation through upregulation of Th17 immune responses in blood/liver and hepatic oxidative stress. This might predispose these patients to increased risk for the development of cardiovascular disorders.

SKP2/P27Kip1 pathway is associated with Advanced Ovarian Cancer in Saudi Patients.

BACKGROUND: Ovarian cancer is the most common gynecological malignancy and constitutes the fifth leading cause of female cancer death. Some biological parameters have prognostic roles in patients with advanced ovarian cancer and their expression may contribute to tumor progression. The aim of this study was to investigate the potential prognostic value of SKP2, genes P27Kip1, K-ras, c-Myc, COX2 and HER2 genes expression in ovarian cancer. MATERIALS AND METHODS: This study was performed on two hundred formalin fixed paraffin embedded ovarian cancer and normal adjacent tissues (NAT). Gene expression levels were assessed using real time PCR and Western blotting. RESULTS: Elevated expression levels of SKP2, K-ras, c-Myc, HER2 and COX2 genes were observed in 61.5% (123/200), 92.5% (185/200), 74% (148/200), 96 % (192/200), 90% (180/200) and 78.5% (157/200) of cancer tissues, respectively. High expression of SKP2 and down-regulation of P27 was associated with advanced stages of cancer. CONCLUSIONS: The association between high expression of c-Myc and SKP2 with low expression of P27 suggested that the Skp2-P27 pathway may play an important role in ovarian carcinogenesis. Reduced expression of P27 is associated with advanced stage of cancer and can be used as a biological marker in clinical routine assessment and management of women with advanced ovarian cancer.

Imiquimod-induced psoriasis-like skin inflammation is suppressed by BET bromodomain inhibitor in mice through RORC/IL-17A pathway modulation.

Psoriasis is one of the most common skin disorders characterized by erythematous plaques that result from hyperproliferative keratinocytes and infiltration of inflammatory leukocytes into dermis and epidermis. Recent studies suggest that IL-23/IL-17A/IL-22 cytokine axis plays an important role in the pathogenesis of psoriasis. The small molecule bromodomain and extraterminal domain (BET) inhibitors, that disrupt interaction of BET proteins with acetylated histones have recently demonstrated efficacy in various models of inflammation through suppression of several pathways, one of them being synthesis of IL-17A/IL-22 which primarily depends on transcription factor, retinoic acid receptor-related orphan receptor C (RORC). However, the efficacy and mechanistic aspect of a BET inhibitor in mouse model of skin inflammation has not been explored previously. Therefore, this study investigated the role of BET inhibitor, JQ-1 in mouse model of psoriasis-like inflammation. Mice were topically applied imiquimod (IMQ) to develop psoriasis-like inflammation on the shaved back and ear followed by assessment of skin inflammation (myeloperoxidase activity, ear thickness, and histopathology), RORC and its signature cytokines (IL-17A/IL-22). JQ-1 suppressed IMQ-induced skin inflammation as reflected by a decrease in ear thickness/myeloperoxidase activity, and RORC/IL-17A/IL-22 expression. Additionally, a RORα/γ agonist SR1078 was utilized to investigate the role of RORC in BET-mediated skin inflammation. SR1078 reversed the protective effect of JQ-1 on skin inflammation at both histological and molecular levels in the IMQ model. The current study suggests that BET bromodomains are involved in psoriasis-like inflammation through induction of RORC/IL-17A pathway. Therefore, inhibition of BET bromodomains may provide a new therapy against skin inflammation.

Carvedilol attenuates inflammatory biomarkers and oxidative stress in a rat model of ulcerative colitis.

Preclinical Research This study evaluated the effects of the carvedilol, a nonselective ß-adrenoceptor anatgonist with α1-adrenoceptor antagonist activity, in a rat model of experimentally induced ulcerative colitis (UC). UC was produced using acetic acid (AA) in animals previously treated with carvedilol (30 mg/kg po, qd) for seven days. Mucus content, lipid peroxidation (LPO) products, sulfhydryl groups, antioxidant enzyme activities, proinflammatory cytokines, prostaglandin E2 and nitric oxide levels were measured in colonic tissues and histopathological changes were assessed. LPO and proinflammatory biomarkers were markedly increased, while mucus content, sulfhydryl groups and enzymatic activities were inhibited in animals administered AA. Pretreatment with carvedilol attenuated LPO elevation, mucus content and sulfhydryl group inhibitions. Antioxidant enzymatic activity and proinflammatory biomarker levels were also restored in carvedilol-pretreated animals. Colonic protection associated with carvedilol pretreatment was further confirmed by histopathological assessment and found to be similar to the standard therapy of mesalazine (100 mg/kg po qd), suggesting that the effects of carvedilol action may be attributable to its anti-inflammatory and antioxidant properties.

Hepato-protective effect of rutin via IL-6/STAT3 pathway in CCl4-induced hepatotoxicity in rats.

BACKGROUND: Carbon tetrachloride (CCl4) induces hepatotoxicity in animal models, including the increased blood flow and cytokine accumulation that are characteristic of tissue inflammation. The present study investigates the hepato-protective effect of rutin on CCl4-induced hepatotoxicity in rats. RESULTS: Forty male Wistar rats were divided into four groups. Group I (control group) received 1 mL/kg of dimethyl sulfoxide intragastrically and 3 mL/kg olive oil intraperitoneally twice a week for 4 weeks. Group II received 70 mg/kg rutin intragastrically. Groups III and IV received CCl4 (3 mL/kg, 30 % in olive oil) intraperitoneally twice a week for 4 weeks. Group IV received 70 mg/kg rutin intragastrically after 48 h of CCl4 treatment. Liver enzyme levels were determined in all studied groups. Expression of the following genes were monitored with real-time PCR: interleukin-6 (IL-6), dual-specificity protein kinase 5 (MEK5), Fas-associated death domain protein (FADD), epidermal growth factor (EGF), signal transducer and activator of transcription 3 (STAT3), Janus kinase (JAK), B-cell lymphoma 2 (Bcl2) and B-cell lymphoma-extra-large (Bcl-XL). The CCl4 groups showed significant increases in biochemical markers of hepatotoxicity and up-regulation of expression levels of IL-6, Bcl-XL, MEK5, FADD, EGF, STAT3 and JAK compared with the control group. However, CCl4 administration resulted in significant down-regulation of Bcl2 expression compared with the control group. Interestingly, rutin supplementation completely reversed the biochemical markers of hepatotoxicity and the gene expression alterations induced by CCl4. CONCLUSION: CCl4 administration causes alteration in expression of IL-6/STAT3 pathway genes, resulting in hepatotoxicity. Rutin protects against CCl4-induced hepatotoxicity by reversing these expression changes.

Hepato-protective effect of rutin via IL-6/STAT3 pathway in CCl4-induced hepatotoxicity in rats

BACKGROUND: Carbon tetrachloride (CCl4) induces hepatotoxicity in animal models, including the increased blood flow and cytokine accumulation that are characteristic of tissue inflammation. The present study investigates the hepato-protective effect of rutin on CCl4-induced hepatotoxicity in rats. RESULTS: Forty male Wistar rats were divided into four groups. Group I (control group) received 1 mL/kg of dimethyl sulfoxide intragastrically and 3 mL/kg olive oil intraperitoneally twice a week for 4 weeks. Group II received 70 mg/ kg rutin intragastrically. Groups III and IV received CCl4 (3 mL/kg, 30 % in olive oil) intraperitoneally twice a week for 4 weeks. Group IV received 70 mg/kg rutin intragastrically after 48 h of CCl4 treatment. Liver enzyme levels were determined in all studied groups. Expression of the following genes were monitored with real-time PCR: interleukin-6 (IL-6), dual-specificity protein kinase 5 (MEK5), Fas-associated death domain protein (FADD), epidermal growth factor (EGF), signal transducer and activator of transcription 3 (STAT3), Janus kinase (JAK), B-cell lymphoma 2 (Bcl2) and B-cell lymphoma-extra-large (Bcl-XL). The CCl4 groups showed significant increases in biochemical markers of hepatotoxicity and up-regulation of expression levels of IL-6, Bcl-XL, MEK5, FADD, EGF, STAT3 and JAK compared with the control group. However, CCl4 administration resulted in significant down-regulation of Bcl2 expression compared with the control group. Interestingly, rutin supplementation completely reversed the biochemical markers of hepatotoxicity and the gene expression alterations induced by CCl4. CONCLUSION: CCl4 administration causes alteration in expression of IL-6/STAT3 pathway genes, resulting in hepatotoxicity. Rutin protects against CCl4-induced hepatotoxicity by reversing these expression changes.

Inhibition of gene expression of carnitine palmitoyltransferase I and heart fatty acid binding protein in cyclophosphamide and ifosfamide-induced acute cardiotoxic rat models.

This study investigated whether cyclophosphamide (CP) and ifosfamide (IFO) therapy alters the expression of the key genes engaged in long-chain fatty acid (LCFA) oxidation outside rat heart mitochondria, and if so, whether these alterations should be viewed as a mechanism during CP- and IFO-induced cardiotoxicity. Adult male Wistar albino rats were assigned to one of the six treatment groups: Rats in group 1 (control) and group 2 (L-carnitine) were injected intraperitoneal (i.p.) with normal saline and L-carnitine (200 mg/kg/day), respectively, for 10 successive days. Animals in group 3 (CP group) were injected i.p. with normal saline for 5 days before and 5 days after a single dose of CP (200 mg/kg, i.p.). Rats in group 4 (IFO group) received normal saline for 5 successive days followed by IFO (50 mg/kg/day, i.p.) for 5 successive days. Rats in group 5 (CP-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days after a single dose of CP as group 3. Rats in group 6 (IFO-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days concomitant with IFO as group 4. Immediately, after the last dose of the treatment protocol, blood samples were withdrawn and animals were killed for biochemical, histopathological and gene expression studies. Treatment with CP and IFO significantly decreased expression of heart fatty acid binding protein (H-FABP) and carnitine palmitoyltransferase I (CPT I) genes in cardiac tissues. Moreover, CP but not IFO significantly increased acetyl-CoA carboxylase2 mRNA expression. Conversely, IFO but not CP significantly decreased mRNA expression of malonyl-CoA decarboxylase. Both CP and IFO significantly increased serum lactate dehydrogenase, creatine kinase isoenzyme MB and malonyl-CoA content and histopathological lesions in cardiac tissues. Interestingly, carnitine supplementation completely reversed all the biochemical, histopathological and gene expression changes induced by CP and IFO to the control values, except CPT I mRNA, and protein expression remained inhibited by IFO. Data from the current study suggest, for the first time, that (1) CP and IFO therapy is associated with the inhibition of the expression of H-FABP and CPT I genes in cardiac tissues with the consequent inhibition of mitochondrial transport and oxidation of LCFA. (2) The progressive increase in cardiotoxicity enzymatic indices and the decrease in H-FABP and CPT I expression may point to the possible contribution of these genes to CP- and IFO-induced cardiotoxicity.

Protective effect of rutin on the antioxidant genes expression in hypercholestrolemic male Westar rat.

BACKGROUND: High-cholesterol diet (HCD) increases the oxidative stress in different tissues leading to many diseases. Rutin (RT) is a natural flavonoid (vitamin p), which possesses an antioxidant activity with protective potential. The present study aimed to examine the potential effects of rutin on hypercholesterolemia-induced hepatotoxicity in rat. METHODS: Male Wistar rats were divided into four groups: GI) control (Rat chow), GII) Rutin (0.2% in rat chow), GIII) HCD (1% cholesterol and 0.5% cholic acid in rat chow) and GIV) rutin (0.2%) + HCD. RESULTS: Rutin in combination with HCD induced a significant protective effect against the hepatotoxicity by reducing the plasma level of alanine transaminase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), and low-density lipoprotein (LDL). The HCD (GII) showed a decrease in glutathione peroxidase (GPx), glutathione reductase (GR) and increase in glutathione S transferase α (GSTα), sulfiredoxin-1(Srx1), glutamate-cysteine ligase (GCL) and paraoxonase-1(PON-1) genes expression levels. CONCLUSION: Treatment with rutin reversed all the altered genes induced by HCD nearly to the control levels. The present study concluded that the HCD feedings altered the expression levels of some genes involved in the oxidative stress pathway resulting in DNA damage and hepatotoxicity. Rutin have a hepatoprotective effect through the mechanism of enhancing the antioxidant effect via amelioration of oxidative stress genes.

Alleviating effects of morin against experimentally-induced diabetic osteopenia.

BACKGROUND: Plant flavonoids are emerging as potent therapeutic drugs effective against a wide range of aging diseases particularly bone metabolic disorders. Morin (3,5,7,20,40-pentahydroxyflavone), a member of flavonols, is an important bioactive compound by interacting with nucleic acids, enzymes and protein. The present study was designed to investigate the putative beneficial effect of morin on diabetic osteopenia in rats. METHODS: Streptozotocin (STZ)-induced diabetic model was used by considering 300 mg/dl fasting glucose level as diabetic. Morin (15 and 30 mg/kg) was treated for five consecutive weeks to diabetic rats. Serum levels of glucose, insulin, deoxypyridinoline cross links (DPD), osteocalcin (OC), bone specific alkaline phosphatase (BALP), telopeptides of collagen type I (CTX), interleukin 1 beta (IL-1ß), interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α), thiobarbituric acid reactive substance (TBARS) and reduced glutathione (GSH) were estimated. Femoral bones were taken for micro CT scan to measure trabecular bone mineral density (BMD) and other morphometric parameters. RESULTS: Significant bone loss was documented as the level of bone turnover parameters including DPD, OC, BALP and CTX were increased in serum of diabetic rats. Morin treatment significantly attenuated these elevated levels. Bone micro-CT scan of diabetic rats showed a significant impairment in trabecular bone microarchitecture, density and other morphometric parameters. These impairments were significantly ameliorated by morin administration. Serum levels of glucose, TBARS, IL-1ß, IL-6 and TNF-α were significantly elevated, while the level of insulin and GSH was decreased in diabetic rats. These serum changes in diabetic rats were bring back to normal values after 5 weeks morin treatment. CONCLUSION: These findings revealed the protective effect of morin against diabetic induced osteopenia. We believed that this effect is through its both the anti-inflammatory and antioxidant properties.

Downregulation of oxidative and nitrosative apoptotic signaling by L-carnitine in Ifosfamide-induced Fanconi syndrome rat model.

It is well documented that ifosfamide (IFO) therapy is associated with sever nephropathy in the form of Fanconi syndrome. Although oxidative stress has been reported as a major player in IFO-induced Fanconi syndrome, no mechanism for this effect has been ascertained. Therefore, this study has been initiated to investigate, on gene expression level, the mechanism of IFO-induce nephrotoxicity and those whereby carnitine supplementation attenuates this serious side effect of IFO. To achieve the ultimate goals of this study, adult male rats were assigned to one of four treatment groups, namely, control, L-carnitine, IFO, and IFO plus L-carnitine. Administration of IFO for 5 days significantly increased serum creatinine, blood urea nitrogen (BUN), and total nitrate/nitrite (NOx) production in kidney tissues. In addition, IFO significantly increased mRNA expression of inducible nitric oxide synthase (iNOS), caspase-9, and caspase-3 and significantly decreased expression of glutathione peroxides (GPx), catalase (CAT), and Bcl2 in kidney tissues. Administration of L-carnitine to IFO-treated rats resulted in a complete reversal of the all biochemical and gene expression changes, induced by IFO, to the control values. Data from this study suggest that L-carnitine prevents the development of IFO-induced nephrotoxicity via downregulation of oxidative and nitrosative apoptotic signaling in kidney tissues.