Sunitinib displays pulmonary fibrosis in experimental rats: Role of IL-17A dependent pathway.

Sunitinib (SUN) is an FDA approved first line drug for management of metastatic renal cancers and advanced cancerous states of gastrointestinal tract, however, side effects including fibrosis has been reported. Secukinumab (Secu) is an immunoglobulin G1 monoclonal antibody that exhibits anti-inflammatory activity by inhibiting several cellular signaling molecules. This study aimed to examine pulmonary protective potential of Secu in SUN-induced pulmonary fibrosis mediated through inhibition of inflammation via targeting IL-17A associated signaling pathway and using pirfenidone (PFD), an antifibrotic drug approved in 2014 for treatment of pulmonary fibrosis with IL-17A as one of its targets, as a reference drug. Wistar rats (160-200 g) were divided randomly into 4 groups (n = 6); Group 1 served as normal control; Group 2 served as disease control where it was exposed to SUN (25 mg/kg; 3 times weekly orally for 28 days); Group 3 was administered SUN and Secu (3 mg/kg subcutaneous at 0,14 and 28 days) and Group 4 was administered SUN and PFD (100 mg/kg/day orally for 28 days). Pro-inflammatory cytokines IL-1ß, IL-6 and TNF-α were measured in addition to components of IL-17A signaling pathway (TGF-ß, collagen, hydroxyproline). Results revealed that IL-17A-associated signaling pathway was activated in fibrotic lung tissue induced by SUN. Relative to normal control, SUN administration significantly elevated lung organ coefficient, IL-1ß, IL-6, TNF-α, IL-17A, TGF-ß, hydroxyproline and collagen expression. Secu or PFD treatment restored the altered levels to nearly normal values. Our study indicates that IL-17A participates in the development and progression of pulmonary fibrosis in a TGF-ß dependent manner. Hence, components of IL-17A signaling pathway represent potential therapeutic targets for protection and treatment of fibro-proliferative lung disease.

The efficacy of bone marrow-derived mesenchymal stem cells and/or erythropoietin in ameliorating kidney damage in gamma irradiated rats: Role of non-hematopoietic erythropoietin anti-apoptotic signaling.

Radiation-induced multiple organ injury, including γ-radiation nephropathy, is the most common. Even with dose fractionation strategy, residual late side effects are inevitable. Bone marrow-derived mesenchymal stem cells (BM-MSCs) transplantation and erythropoietin (EPO) have shown to be effective in treating chronic kidney disease and associated anemia. This study aimed to evaluate the effect of BM-MSCs and/or EPO in fractionated γ-irradiation induced kidney damage in rats. Adult male Wistar rats were randomized into 2 groups; normal and 8 Gy (fractionated dose of 2 Gy for 4 days) γ-irradiated rats. Animal from both groups were subdivided to receive the following treatments: BM-MSCs (1 × 106 cells/rat, i.v - once), EPO (100 IU/kg, i.p - every other day for 30 days) or their combined treatment (BM-MSCs and EPO). γ-Irradiated rats showed a noticeable elevation in serum urea and creatinine, kidney malondialdehyde (MDA) and caspase 3 activity. They also revealed significant drop in kidney glutathione (GSH) and Bcl2 protein contents. Conspicuously, they revealed down-regulation of renal EPO signaling (EPO, EPOR, pJAK2, pPI3K and pAkt). Conversely, groups treated with BM-MSCs and/or EPO revealed significant modulation in most tested parameters and appeared to be effective in minimizing the hazard effects of radiation. In conclusion, BM-MSCs and/or EPO exhibited therapeutic potentials against nephrotoxicity induced by fractionated dose of γ-irradiation. An effect mediated by antioxidant and non-hematopoietic EPO downstream anti-apoptotic signaling (PI3K/Akt) pathway. EPO potentiate the repair capabilities of BM-MSCs making this combined treatment a promising therapeutic strategy to overcome radiotherapy-induced kidney damage.

Infliximab ameliorates tumor necrosis factor-alpha exacerbated renal insulin resistance induced in rats by regulating insulin signaling pathway.

Infliximab (IFX), a monoclonal antibody for tumor necrosis factor-alpha (TNF-α), is known to restore blood glucose homeostasis. However, its effects on improving renal insulin resistance (IR) are not yet studied. So we investigate the impact of infliximab on renal insulin signaling pathway in IR rat model regarding to metformin (MET). The induced IR was confirmed by a high oral glucose tolerance test, an elevation of lipid profile and the homeostatic model assessment of insulin resistance 2 (HOMA-IR 2) values. Subsequently, IR rats were concurrently treated with either MET (100 mg/kg/day) or IFX (one dose 5 mg/kg) besides IR and normal control (NC) groups. Four weeks later, IR control rats displayed hyperglycemia, hyperinsulinemia and elevation in HOMA-IR 2, renal function markers and renal tissue TNF-α, interleukins-1ß and 6 (Il-1ß, IL-6) and suppressor of cytokines signaling 3 (SOCS3) contents as well as glomerulosclerosis when compared to NC group. Additionally, the phosphorylation of renal insulin receptor substrate 1 (IRS1), phosphatidylinositol 3-kinase (PI3K) and protein kinase B (Akt) were markedly impaired. Treatment with either MET or IFX significantly improved IR and kidney functions. The effects of the drugs were achieved by the downregulation of renal inflammatory cytokines and SOCS3 levels and the amelioration of the renal IRS1/PI3K/Akt pathway. In conclusion, MET and IFX ameliorated the TNF-α worsening effect on IR in rat renal tissues by regulating insulin signaling. Interestingly, infliximab was superior to metformin in regulating insulin signaling pathway. Therefore, infliximab could be used as an adjuvant therapy in improving renal IR.

Inhibition of tumor necrosis factor-α enhanced the antifibrotic effect of empagliflozin in an animal model with renal insulin resistance.

Insulin resistance (IR) has emerged as one of the main risk factors for renal fibrosis (RF) that represents a common stage in almost all chronic kidney disease. The present study aims to investigate the inhibitory effect of empagliflozin (EMPA "a sodium-glucose co-transporter 2 inhibitor") and infliximab [IFX "a tumor necrosis factor-α (TNF-α) antibody"] on RF in rats with induced IR. IR was induced by adding 10% fructose in drinking water for 20 weeks. Thereafter, fructose-induced IR rats were concurrently treated with EMPA (30 mg/kg), IFX (1 dose 5 mg/kg), or EMPA + IFX for 4 weeks, in addition to IR control group (received 10% fructose in water) and normal control (NC) group. Rats with IR displayed hyperglycemia, deterioration in kidney functions, glomerulosclerosis, and collagen fiber deposition in renal tissues as compared to NC. This was associated with downregulation of the renal sirtuin 1 (Sirt 1) expression along with higher renal tissue TNF-α and transforming growth factor-ß1 (TGF-ß1) levels. Both EMPA and IFX significantly modulated the aforementioned fibrotic cytokines, upregulated the renal Sirt 1 expression, and attenuated RF compared to IR control group. Of note, IFX effect was superior to that of EMPA. However, the combination of EMPA and IFX alleviated RF to a greater extent surpassing the monotherapy. This may be attributed to the further upregulation of renal Sirt 1 in addition to the downregulation of fibrotic cytokines. These findings suggest that the combination of EMPA and IFX offers additional benefits and may represent a promising therapeutic option for RF.

Modulation of brain insulin signaling in Alzheimer's disease: New insight on the protective role of green coffee bean extract.

Objective: Alzheimer's disease (AD), a neurodegenerative disorder, involves brain insulin signaling cascades and insulin resistance (IR). Because of limited treatment options, new treatment strategies are mandatory. Green coffee bean extract (GCBE) was reported to attenuate IR and improve brain energy metabolism. We aimed to investigate the possible use of GCBE as a prophylactic strategy to delay the onset of AD or combined with pioglitazone (PIO) as a strategy to retard the progression of AD.Methods: Rats received 10% fructose in drinking water for 18 weeks to induce AD. GCBE-prophylactic group received GCBE for 22 weeks started 4 weeks prior to fructose administration. The PIO group treated with PIO for 6 weeks started on week 12 of fructose administration. The GCBE+PIO group received GCBE for 22 weeks started 4 weeks prior to fructose administration and treated with PIO for the last 6 weeks of fructose administration.Results: Pretreatment with GCBE, either alone or combined with PIO, alleviated IR-induced AD changes. GCBE improved cognition, decreased serine phosphorylation of insulin receptor substrate, increased phosphoinositol-3 kinase (PI3K) activity and protein kinase B (Akt) gene expression, decreased glycogen synthase kinase-3ß (GS3Kß) gene expression and Tau hyperphosphorylation.Discussion: GCBE exerted neuroprotective effects against IR-induced AD mediated by alleviating IR and modulating brain insulin signaling cascade.

Smooth muscle-specific deletion of MnSOD exacerbates diabetes-induced bladder dysfunction in mice.

Bladder dysfunction in diabetes progresses gradually over time. However, the mechanisms of the development are not clear. We tested the hypothesis that oxidative stress plays a key role in the development of diabetic bladder dysfunction using an inducible smooth muscle (SM)-specific superoxide dismutase 2 (Sod2) gene knockout (SM-Sod2 KO) mouse model. Eight-week-old male Sod2lox/lox, SM-CreERT2(ki)Cre/+ mice and wild-type mice were assigned to diabetic or control groups. 4-Hydroxytamoxifen was injected into Sod2lox/lox, SM-CreERT2(ki)Cre/+ mice to activate CreERT2-mediated deletion of Sod2. Diabetes was induced by injection of streptozotocin, whereas control mice were injected with vehicle. Nine weeks later, bladder function was evaluated, and bladders were harvested for immunoblot analysis. Wild-type diabetic mice presented compensated bladder function along with increased nitrotyrosine and MnSOD in detrusor muscle. Induction of diabetes in SM-Sod2 KO mice caused deteriorated bladder function and even greater increases in nitrotyrosine compared with wild-type diabetic mice. Expression levels of apoptosis regulator Bax and cleaved caspase-3 were increased, but apoptosis regulator Bcl-2 expression was decreased in detrusor muscle of both diabetic groups, with more pronounced effects in SM-Sod2 KO diabetic mice. Our findings demonstrate that exaggerated oxidative stress can accelerate the development of bladder dysfunction in diabetic mice and the enhanced activation of apoptotic pathways in the bladder may be involved in the process.

Nicotine and vascular endothelial dysfunction in female ovariectomized rats: role of estrogen replacement therapy.

OBJECTIVES: The protective effects of estrogen replacement therapy (ERT) against oxidative injury and endothelial dysfunction in the aortic tissues induced with nicotine in ovariectomized (OVX) rats were investigated. METHODS: Female rats were divided into a sham-operated group (n = 8) and four groups in which OVX rats received either vehicle (0.1 ml sesame oil, i.m., n = 8), or nicotine (0.1 mg/kg, s.c., n = 8), or estradiol benzoate (0.1 mg/kg, i.m., n = 8), or both nicotine and estradiol benzoate (n = 8) starting at week 5 after the surgery and continuing for the following 6 weeks. KEY FINDINGS: ERT was effective in preventing the rise in plasma lipid profile, atherogenic index and the level of induced endothelin-1 (ET-1) in nicotine-treated OVX rats. It also reduced aortic malondialdehyde, hydroxyproline levels, calcium content and caspase-3 expression induced in nicotine-treated OVX rats. ERT increased serum estradiol, high-density lipoprotein cholesterol and nitric oxide levels in nicotine-treated OVX rats. Furthermore, ERT was effective in restoring reduced glutathione and cyclic guanosine monophosphate contents and endothelial nitric oxide synthase expression in aortic tissues of nicotine-treated OVX rats. CONCLUSIONS: Short-term ERT could be a promising therapeutic strategy to minimize nicotine-induced oxidative stress and vascular endothelial dysfunction in menopausal women subjected to environmental smoke.

Protection against doxorubicin cardiomyopathy in rats: role of phosphodiesterase inhibitors type 4.

Selective cardiotoxicity of doxorubicin (DOX) remains a significant and dose-limiting clinical problem. The mechanisms implicated are not yet fully defined but may involve the production of reactive oxygen species or expression of cytokines. Although patients with advanced congestive heart failure express elevated circulating levels of tumour necrosis factor-alpha (TNFalpha), little is known about the prognostic importance and regulation of TNF in the heart in cardiac disease states. Here we tested whether the expression of TNFalpha, along with oxidative stress, is associated with the development of DOX-induced cardiomyopathy (DOX-CM) and whether concurrent treatment with taurine (Taur), an antioxidant, or rolipram (Rolp), a TNFalpha inhibitor, offer a certain protection against DOX cardiotoxic properties. DOX (cumulative dose, 12 mg kg(-1)) was administered to rats in six equal (intraperitoneal) injections over a period of 6 weeks. Cardiomyopathy was evident by myocardial cell damage, which was characterized by a dense indented nucleus with peripheral heterochromatin condensation and distorted mitochondria, as well as significant increase in serum levels of creatine kinase and lactate dehydrogenase. DOX also induced an increment (P<0.001) in serum TNF and plasma nitric oxide levels. The extent of left ventricular (LV) superoxide anion, lipid peroxide measured as malondialdehyde, catalase and calcium content were markedly elevated, whereas superoxide dismutase, total and non-protein-bound thiol were dramatically decreased in DOX-treated rats. Exaggeration of DOX-CM was achieved by intraperitoneal injection of lipopolysaccharide (LPS) (1 mg kg(-1)) 18 h before sampling and evaluated by highly significant increase in heart enzymes (P<0.001), oxidative stress biomarkers and TNFalpha production. Pre- and co-treatment of DOX or DOX-LPS rats with Taur (1% daily supplemented in drinking water, 10 days before and concurrent with DOX) or Rolp (3 mg kg(-1), intraperitoneally, one dose before DOX administration then every 2 weeks throughout the experimental period) ameliorated the deleterious effect of both DOX and LPS on the aforementioned parameters. Meanwhile, it is noteworthy that Rolp exhibited a more preferable effect on serum TNFalpha level. Taur and rolipram also restored the myocardial apoptosis induced by DOX. In conclusion, a cumulative dose of DOX affected free radical and TNFalpha production in the heart of an experimental cardiomyopathy animal model. The current results suggest that down-regulation of these radicals and cytokines could be maintained by using the free radical scavenger Taur or, more favourably, the TNFalpha inhibitor Rolp.

Hyperhomocysteinaemia and cardiovascular risk in female ovariectomized rats: role of folic acid and hormone replacement therapy.

Hyperhomocysteinaemia is an independent risk factor for arteriosclerosis, recurrent thromboembolic complications and osteoporosis. After menopause, a high level of total homocysteine seems to be secondary to the altered hormonal status. Hormone replacement therapy (HRT) limits the development of coronary artery disease through a variety of mechanisms. One such mechanism is through affecting homocysteine metabolism. Folate and vitamin B12 deficiencies are considered to be major risks for hyperhomocysteinaemia. This study, therefore, was undertaken to examine whether lowering homocysteine with HRT or folic acid in ovariectomized rats could attenuate cardiovascular complications. Sixty sexually mature female Wistar rats were ovariectomized. Three weeks later, they were treated with estradiol (15 microg kg(-1), every two weeks, i.m.) or folic acid (90 microg daily, orally), either alone or in a combined form for four weeks. In addition, groups of ovariectomized rats (positive control) and healthy rats (negative control) were given cottonseed oil. Blood samples were then collected for serum and plasma separation. Serum total homocysteine, folate, estradiol, plasma nitric oxide (NO), lipid profile, and susceptibility of non-high-density-lipoprotein cholesterol (non HDLC) content to oxidation were determined. In ovariectomized rats, hyperhomocysteinaemia was established and associated with significant increments of both atherogenic indexes (total cholesterol/HDLC, low-density-lipoprotein cholesterol (LDLC)/HDLC) and susceptibility of their non HDLC to oxidation. However, plasma NO, serum folate, and estradiol levels significantly decreased. HRT and folic acid significantly reduced total homocysteine and susceptibility of non HDLC to oxidation and increased plasma NO content. Moreover, a significant negative correlation was found between total homocysteine versus folate and estradiol (r = -0.5, P < 0.01; r = -0.25, P < 0.05, respectively). Meanwhile, a positive correlation with the susceptibility of lipoprotein to oxidation was observed (r = 0.85, P < 0.001). In conclusion, a low folate level is found to be associated with elevated total homocysteine. Folic acid supplementation, either individually or in a combined form with HRT, has a beneficial effect in low estrogen status subsequent to ovariectomy.

A novel concept to preserve the beneficial effects of hormone replacement therapy in bilaterally female ovariectomized rats: role of lovastatin therapy.

Estrogen replacement therapy (ERT) is claimed to reduce cardiovascular mortality by about 50% in postmenopausal women. This improvement is caused by favorable changes in lipid and lipoproteins metabolism, however, it also increases the incidence of the endometrial hyperplasia. Addition of progestin to ERT, referred to as hormone replacement therapy (HRT), has been shown to successively reduce this risk to the endometrium. Unfortunately, it has an adverse effect on high-density lipoprotein cholesterol (HDLC) concentration, thus compromising the benefits of ERT. Therefore the issue here whether HRT given alone and/or concomitantly with 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (lovastatin) could exert any significant additional favorable effect on the lipid profile in bilaterally ovariectomized female rats. Sixty female Wistar rats were ovariectomized and treated with ERT (0.625 mg kg (-1)estradiol, E (2), IM every 2 weeks), HRT (estradiol plus progesterone, E (2)+ P, 0.625 mg kg (-1)estradiol and 5 mg progesterone kg (-1) respectively, IM every 2 weeks), and lovastatin (20 mg kg (-1)day (-1)orally) plus HRT (L + HRT) for 6 weeks. Blood aliquots were collected for serum and plasma separation. Serum vitamin E and plasma levels of C-reactive protein (CRP), nitric oxide (NO), lipid profile, and the susceptibility of non-HDLC to oxidation were determined. Moreover, thoracic aortas were dissected and directed for measurement of its lipid peroxide and NO contents. Treatment of ovariectomized rats with HRT showed a significant decrease ( P< 0.0001) in HDLC concentration compared to the group treated alone with ERT and increase ( P< 0.0001) in CRP levels compared to ovariectomized rats. HDLC and CRP are two powerful and significant predictors for increased cardiovascular risk in postmenopausal women. Addition of lovastatin as a complementary therapy to HRT revealed a significant 27% increment in HDLC and 48% decrement in CRP concentrations. Moreover, it significantly increased vitamin E, each of plasma and tissue content of NO and decreased atherogenic indexes (TC/HDLC, LDLC/HDLC), aortic lipid peroxide and susceptibility of non-HDLC to oxidation. In conclusion, this current study demonstrated that lovastatin together with continuous combined HRT seems to be more effective in the secondary prevention of coronary heart disease not only due to lipid lowering properties but also related to several other additive effects such as modification of endothelial function and inflammatory responses.