BACKGROUND:Diabetic nephropathy (DN) is a clinical condition characterized by progressive decline in kidney function, with persistent albuminuria. DN pathophysiology involves chronic hyperglycaemia, oxidative stress, glomerulosclerosis, interstitial fibrosis and apoptosis. Semaglutide (S) and acarbose (A), antidiabetic drugs with different mechanisms, were explored in this study in a combination regimen to treat experimentally-streptozotocin (STZ)-induced DN in a rat model. METHODS:Fifty-six male rats were divided into 7-groups. Except for the non-diabetic negative control rats (group-I), all other groups had DN induced by an I·P injection of STZ. In group-II & III, STZ-diabetic rats did not receive any medication. Diabetic rats in group-IV (DN/S) and group-V (DN/A), administered semaglutide (30 nmol/kg/week/S.C), and acarbose (50 mg/kg/day/orally), respectively, for 4-weeks. Group-VI [DN/S/A(HD)], administrated both drugs. In group-VII [DN/S/A(LD)], diabetic rats administrated semaglutide (20 nmol/kg/week/S·C) and acarbose (30 mg/kg/day/orally) for 4-weeks. Serum glucose, creatinine/KIM-1 levels and albuminuria were measured. Renal; Nrf-2/HO-1 immunoreactivities, NF-κB/TNF-α/IL-10 levels, TGF-β1/Smad 2/3, and cl-caspase-3/Bax/Bcl-2 expressions were assessed. RESULTS:Compared to solo treatment of either drug, semaglutide/acarbose combination (in high and low doses) regenerated the histopathological features and restored kidney functions through stimulation of Nrf-2/HO-1, attenuation of inflammatory signals via modulation of NF-κB/TNF-α/IL-10 levels. Fibrosis and apoptosis were mitigated by suppression of the TGF-β1/Smad and overall reduction in Bax/Bcl-2 ratio, respectively. CONCLUSION:The combination of semaglutide/acarbose is more effective than the solo treatment with either drug, even upon using lower doses in the combination, suggesting a synergistic effect. The study unveiled a potential therapeutic regimen for DN, that was not evaluated before.

Metformin is a traditional antidiabetic drug for type 2 diabetes mellitus. However, it showed antitumor activity in many types of tumors, and it also has an influence on tumor metastasis in several types of tumors. It is transported through organic cationic transporters (OCTs), OCT1, OCT2, and OCT3, into the cells or into tumor microenvironment (TME). The complex interaction of metformin and its transporters on immune infiltration in TME of different types of tumors of The Cancer Genomic Atlas (TCGA) is not yet studied. The objective of this study is to identify the most suitable therapeutic target of tumors and immune infiltrates for metformin and its transporters in the TME. TIMER2.0, a bioinformatic tool, and other computational analysis were used to investigate this complex interaction; moreover, the identification of metformin target protein in TME is also investigated. The results revealed that the most suitable therapeutic target for metformin and OCTs among 32 types of TCGA data tumor types is Breast Invasive carcinoma (BRCA), and the most relevant immune infiltrate among 14 types of immune infiltrates that yields better prognosis and better therapeutical effect in TME is Macrophage M1. Furthermore, metformin showed a cytotoxic effect and an inhibitory effect on Urokinase Plasminogen Activator (uPA) gene expression in a concentration dependent fashion in MDA-MB-231 breast cancer cell line. This may suggest that metformin is a promising antitumor drug, stimulant for natural antitumor immune infiltrates, and inhibitor for metastasis in breast cancer.

One of the leading causes of cancer-related deaths worldwide is colorectal cancer (CRC). Extracellular ATP (e-ATP) and purinergic receptors (P2R) play a central role in CRC proliferation and progression. Human antigen R (HuR) is becoming more and more understood to be essential for the expression of genes linked to cancer. The current study demonstrates that ATP can mediate CRC (Caco-2 cells) progression via induction of HuR nucleocytoplasmic shuttling and subsequent expression of cancer-related genes, a consequence mostly mediated via the P2R receptor. It was also noted that suppression of HuR activity by using dihydrotanshinone I (DHTS) prevents cancer-related gene expression and subsequent CRC (Caco-2 cells) progression induced by ATP. The expression of cyclin A2/cyclin-dependent kinase 2 (CDK2), Bcl-2, ProT-α, hypoxia‐inducible factor1-α (HIF1-α), vascular endothelial growth factor A (VEGF-A), transforming growth factor-β (TGF-β) and matrix metallopeptidase 9 (MMP-9) induced by ATP were highly reduced in the presence of either PPADS (non-selective P2R antagonist) or DHTS. In addition, e-ATP-induced Caco-2 cell proliferation as well as cell survival were highly reduced in the presence of either PPADS or DHTS or selective CDK-2 inhibitor (Roscovitine) or selective Bcl-2 inhibitor (ABT-263). Furthermore, it was found that MMP-9 is critical for Caco-2 cells migration induced by e-ATP as demonstrated by a clear reduction in cells migration in the presence of a selective MMP-9 inhibitor (Marimastat). Collectively, these data demonstrate that ATP through P2R activation can induce HuR nucleocytoplasmic shuttling that could be translated into an increase in cancer-related genes expression and subsequent, cell proliferation and progression.

Renal fibrosis is the common endpoint of nearly all chronic and progressive nephropathies. Cell death and sterile inflammation are the main characteristics of renal fibrosis, which can lead to end-stage renal failure. The inflammatory reaction triggered by tissue damage is strongly related to necroptosis, a type of caspase-independent, regulated cell death. Using an animal model of unilateral ureteral obstruction (UUO), the anti-fibrotic effects of sorafenib (SOF), a multi-kinase inhibitor, and edaravone (EDV), a potent antioxidant and free radical scavenger, were examined in rats with obstructive nephropathy. Experimentally, animals were divided randomly into five groups: sham; UUO; UUO + SOF (5 mg/kg/day, P.O.); UUO + EDV (20 mg/kg/day, P.O.); and UUO + SOF + EDV groups. The kidney function biomarkers, oxidant/antioxidant status, renal mRNA expressions of TNF-α, collagen-1α, protein expressions of RIPK-1, RIPK-3, MLKL, caspase-8, HYP, MPO, and TNF-α were all significantly modulated by UUO. Administration of either SOF or EDV significantly attenuated cellular and molecular changes induced by UUO. Also, histopathological changes were improved. Moreover, SOF in combination with EDV, significantly improved UUO-induced renal fibrosis compared with each drug alone. Collectively, administration of either SOF or EDV or both of them significantly attenuated the rats with obstructive nephropathy, possibly by blocking the RIPK-3/MLKL necroptotic pathway and suppressing renal oxidative stress and inflammation.

Extracellular ATP is a potent signaling molecule that regulates different biological activities including proliferation, differentiation as well as cell death.Several studies have implicated extracellular ATP as a mitogenic agent in various cells in vitro.Herein, we aimed to investigate the involvement of ATP in mitogenic processes in vivo via stimulation of P2X7Rs in rat kidney.Results from the current study showed that administration of ATP caused rapid activation of ADAM-17 with subsequent phosphorylation of EGFR and ERK-1/2.Cyclin B1 expression in rat kidney was also increased following ATP administration.Interestingly, a marked decrease in renal ADAM-17 levels as well as EGFR and ERK-1/2 phosphorylation was noticed when the animals were pretreated with the purinergic P2X7R antagonist, A 438079 before ATP administration.Moreover, ATP administration significantly increased serum levels of NO, an effect that was significantly ameliorated by A 438079 pretreatment.Collectively, current findings demonstrate the ability of ATP to activate the ADAM-17-mediated EGFR/ERK-1/2 phosphorylation and to enhance the expression of the cell proliferation gene cyclin B1, an effect that is largely mediated via P2X7Rs and involves NO modulation.Thus, the safety of ATP supplementation as an ergogenic aid for muscle building and performance enhancement should be reassessed.

This study was designed to investigate the potential of extracellular adenosine 5′-triphosphate (ATP) via the P2 X 7 receptor to activate the renal fibrotic processes in rats. The present study demonstrates that administration of ATP rapidly activated transforming growth factor-β (TGF-β) to induce phosphorylation of Smad-2/3. Renal connective tissue growth factor (CTGF) and tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA and protein expressions were also increased following ATP administration. A decrease in TGF-β amount in serum as well as renal Smad-2/3 phosphorylation was noticed in animals pre-treated with the specific antagonist of P2 X 7 receptor, A 438,079. In addition, a significant reduction in mRNA and protein expression of CTGF and TIMP-1were also observed in the kidneys of those animals. Collectively, the current findings demonstrate that ATP has the ability to augment TGF-β-mediated Smad-2/3 phosphorylation and enhance the expression of the pro-fibrotic genes, CTGF and TIMP-1, an effect that is largely mediated via P2 X 7 receptor.

Leflunomide-induced liver injury has been an important problem since its approval. Although, severe cases of leflunomide-induced liver injury leading to hospitalization are rare, the risk is higher with concurrent liver disease or use of other hepatotoxic drugs. The current study was conducted to investigate the potential protective effects of carvedilol and crocin alone and in combination against leflunomide-induced hepatic injury and to clarify the possible mechanism(s) through which carvedilol and crocin may elicit their effects. Fifty male albino mice were allocated into five groups: normal control group, leflunomide group, carvedilol group, crocin group, and combination group. These groups were given vehicle, leflunomide, leflunomide plus carvedilol, leflunomide plus crocin, and leflunomide plus combination of carvedilol and crocin, respectively. The study was conducted for 8 weeks, and different parameters were assessed. The results demonstrated that leflunomide significantly increased the serum levels of AST, ALT, ALP, hepatic MDA, nitrite, mTOR gene, PI3K gene, TGF-β, and the pathological changes alongside with the significant decrease of serum albumin, total protein, hepatic catalase, and GSH. While the coadministration of carvedilol, crocin and their combination with leflunomide significantly decreased the serum levels of AST, ALT, ALP, hepatic MDA, mTOR gene, PI3K gene, TGF-β, and the pathological changes alongside with the significant elevation of serum albumin, total protein, hepatic catalase, and GSH. This study is suggesting several solutions for Leflunomide-induced hepatotoxicity demonstrated by the protective effect of the antihypertensive drug carvedilol, the natural product crocin, and their combination which was demonstrated to be superior to each drug alone.

Aim: The present work was designed to examine of the potential anti-inflammatory effect of rosuvastatin (ROSV) and/or Lefleunamide (LFLU) against Complete Freunds Adjuvant (CFA)-induced arthritis in rats. Methods: The mRNA level of perxisome proliferator-activated receptor-alpha (PPAR-a) was determined using Real-time PCR. The levels of NF-B, iNOS, IL-6, TNF-a and SOD activity were measured using ELISA. The swollen paws were measured using caliper. The GSH level was measured using colorimetric assay. The level of malondialdehyde (MDA) was determined using thiobarbituric acid reactive substances assay kit. Results: ROSV induced the expression of PPAR-a that suppresses NF-kB as demonstrated by a strong reduction in NF-kB level in animals treated with ROSV. Also, ROSV administration reduced the levels of the inflammatory mediators IL-6 and TNF-a. In addition, iNOS and MDA content as well as expression of MMP-9 and MMP-2 induced by CFA is abrogated in animals treated with ROSV. Also GSH content and SOD activity were highly increased in ROSV-treated animals. Furthermore, the size of right paw induced by CFA was reduced in ROSV-treated rats. Moreover, the histopathological alterations induced by CFA were highly improved in animals treated with ROSV. Similar results were also found in animals treated with LFLU. Importantly, similar effects were obtained in rats treated with both ROSV and LFLU in half doses. Conclusion: This study demonstrates that ROSV as well as LFLU has the ability to inhibit rheumatoid arthritis in experimental model induced by CFA. Importantly, concomitant administration of ROSV and LFLU in half doses synergize against rheumatoid arthritis.

One of the most common causes of cancer mortality worldwide is hepatocellular carcinoma (HCC). Extracellular signal-regulated kinase (ERK1/2) pathway has been shown to play an important role in the development and progression of HCC. Here, we demonstrate that the immunosuppressive agent cyclosporin A (CsA) has the ability to increase the cellular growth in HCC (HepG2 cells) via activation of ERK1/2 signaling cascade. It was found that ERK1/2 phosphorylation induced by CsA was highly reduced in the presence of the reactive oxygen species (ROS) scavenger polyethylene glycol-superoxide dismutase (PEG-SOD). Furthermore, it was observed that inhibition of metalloproteinase activity using TAPI-2 prevents ERK1/2 activation by CsA. Moreover, a disintegrin and metalloproteinase domain 17 (ADAM-17) activity was found to be critical for ERK phosphorylation by CsA. In addition, CsA-induced ERK phosphorylation was highly reduced in the presence of either neutralizing anti-heparin-binding-epidermal growth factor (HB-EGF) antibody or UO126 (MEK inhibitor). By using the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor AG1478, it was found that EGFR is critical for ERK phosphorylation induced by CsA. Furthermore, CsA-induced cell proliferation was strongly reduced in the presence of either PEG-SOD or TAPI-2 or neutralizing anti-ADAM17 antibody or neutralizing anti-HB-EGF antibody or AG1478 or UO126. Collectively, these data demonstrate that CsA has the ability to activate ERK1/2 signaling cascade that could be translated into an increase in HepG2 cell proliferation. Furthermore, these data support the role of ROS, ADAM-17, and EGFR in ERK1/2 signaling activation and subsequent cell proliferation induced by CsA in HepG2 cells.

The calcineurin inhibitor, cyclosporin A (CsA) is one of the most common immunosuppressive agents used in organ transplantation. However, its clinical use is often limited by several unwanted effects including nephrotoxicity and hepatotoxicity. By using immunohistochemical and ELISA techniques, it was found that CsA administration causes a rapid activation of a disintegrin and metalloproteases-17 (ADAM-17), epidermal growth factor receptor (EGFR) and subsequent ERK1/2 phosphorylation in the liver and kidney of albino mice. Furthermore, this study presents mechanistic relevance of this signaling cascade involving reactive oxygen species (ROS)-mediated ADAM-17/EGFR/ERK1/2 activation as indicated by a clear reduction in ADAM-17 and EGFR activities as well as ERK1/2 phosphorylation when the animals pretreated with Polyethylene glycol-superoxide dismutase (PEG-SOD) before CsA administration. Collectively, our findings demonstrate that CsA has the ability to activate ADAM-17-mediated EGFR/ERK1/2 phosphorylation in the liver and kidney of albino mice in ROS-dependent manner. Finally, these data may support the concept of using antioxidant therapy as a valuable approach for the prevention of CsA-induced nephrotoxicity and hepatotoxicity.

Overproduction of reactive oxygen species (ROS) and a consequent imbalance between oxidants and endogenously produced antioxidants have been reported to play an important role in liver disease. Recently, the wheat germ oil (WGO) has attracted much attention due to its excellent nutritional value. WGO has a high content of vitamin E, the most powerful natural antioxidant. It has been reported that WGO has the ability to protect against hepatotoxicity induced by a variety of oxygen radical generators. WGO has been shown to inhibit lipid peroxidation induced by ROS in liver tissue. Furthermore, it has been reported that WGO has the ability to improve not only the serum levels of liver enzymes but also the histopathological changes induced by ROS in liver tissue. Thus, a daily dose of WGO is the perfect way to enrich diet with a high concentration of the most powerful natural antioxidant vitamin E that may protect the liver through its unique cytoprotective properties.

Cyclosporin A (CsA) is the most common immunosuppressive drug used in organ transplantation. However, the clinical use of CsA is often limited by several side effects including hepatotoxicity. In the present study, it was found that administration of CsA causes a rapid activation of TGF-β/Smad signaling cascade and subsequent expression of the profibrotic genes connective tissue growth factor (CTGF) and tissue inhibitors of matrix metallproteinases-1 (TIMP-1) in rat liver. In addition, Smad phosphorylation and subsequent CTGF and TIMP-1 expression were markedly reduced in the presence of neutralizing monoclonal TGFβ1–3 antibody. Furthermore, CsA administration significantly increased the serum levels of the liver enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) as well as lipid peroxidation in hepatic tissues. Moreover, significant reduction in the hepatic content of reduced glutathione (GSH), superoxide dismutase (SOD), and catalase (CAT) was observed in CsA-alone-treated animals. Histopathological changes were also observed in CsA-alone-treated rats. Pretreatment of animals with Vitamin E (Vit E) before CsA administration significantly reduced TGF-β level as well as Smad phosphorylation and subsequent CTGF and TIMP-1 expression. Furthermore, administration of PEG-SOD clearly attenuated TGF-β/Smad signaling induced by CsA. Moreover, concomitant administration of Vit E along with CsA significantly ameliorated the histopathological changes and improved liver function as well as the antioxidant capacity. Finally, this study shows that the immunosuppressive efficiency of CsA was not altered in the presence of Vit E. These data may support the concept of using antioxidant therapy as a valuable approach for the prevention of CsA-induced tissue fibrosis.

Liver fibrosis results from chronic inflammation that precipitates excessive accumulation of extracellular matrix. Oxidative stress is involved in its pathogenesis. This study aimed to elucidate the potential antifibrotic effect of the NADPH oxidase (NOX) inhibitor, apocynin against concanavalin A (ConA)-induced immunological model of liver fibrosis, and to investigate the ability of the antioxidant, alpha-lipoic acid (α-LA) to potentiate this effect. Rats were treated with apocynin and/or α-LA for six weeks. Hepatotoxicity indices, oxidative stress, insulin, NOXs, inflammatory and liver fibrosis markers were assessed. Treatment of animals with apocynin and α-LA significantly ameliorated the changes in liver functions and histopathological architecture induced by ConA. Liver fibrosis induced by ConA was evident where alpha-smooth muscle actin and transforming growth factor- beta1 were elevated, which was further confirmed by Masson's trichrome stain and increased hydroxyproline. Co-treatment with apocynin and α-LA significantly reduced their expression. Besides, apocynin and α-LA significantly ameliorated oxidative stress injury evoked by ConA, as evidenced by enhancing reduced glutathione content, antioxidant enzymes activities and decreasing lipid peroxides. ConA induced a significant elevation in serum insulin level and inflammatory markers; tumor necrosis factor-alpha, interleukin-6 and nuclear factor kappa b. Furthermore, the mRNA tissue expression of NOXs 1 and 4 was found to be elevated in the ConA group. All these elevations were significantly reduced by apocynin and α-LA co-treatment. These findings indicate that using apocynin and α-LA in combination possess marked antifibrotic effects, and that NOX enzymes are partially involved in the pathogenesis of ConA-induced liver fibrosis.

Viral hepatitis-induced oxidative stress accompanied by increased levels of transforming growth factor-β (TGF-β) and hepatic fibrosis are hallmarks of hepatitis C virus infection. The present study was designed to investigate the potential protective effect of propolis against liver injury induced by concanavalin A (Con A), a T-cell-dependent model that causes an immune-mediated hepatitis in a similar pattern to the one induced by viral infections. In the present study, rats were randomly divided into four groups. The first group (control) was administered the vehicle of Con A (i.v.) for 24 h. The second group received Con A (12 mg/kg body weight i.v.) for 24 h. The third group received propolis (300 mg/kg by oral gavage) 5 days before and concurrently with Con A for 24 h. The last group received propolis alone. Following a single injection of Con A, histopathological changes as well as significant reduction in albumin level were observed. In addition, serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin were significantly increased. These increases correlated with an increase in lipid peroxidation and downregulation of reduced glutathione (GSH) as well as superoxide dismutase (SOD) and catalase activities in liver tissue. Furthermore, these changes were associated with an increase in serum levels of the inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) as well as the profibrotic cytokine TGF-β. Moreover, TGF- β activation was accompanied with an increase in Smad phosphorylation. Interestingly, concomitant administration of propolis along with Con A significantly attenuated all these negative effects and improved liver function indicating that propolis has the ability to protect rats from Con A-induced hepatitis.

Aim: This study was designed to evaluate the possible modulatory effect of vitamine E on tacrolimus (FK506)-induced renal injury in rats.Methods: Twenty-four male Wistar rats (6 animals in each group) were used in this study. The first group (Control) received normal saline intraperitoneal (i.p.) daily for 21 days. The second group received FK506 (1 mg/kg/day i.p.) daily for 21 days. The third group was administered Vitamin E (250 mg/kg/day by oral gavage) 5 days before and concurrently during FK506 administration daily for 21 days. The fourth group received Vitamin E alone (as previously described in the third group).Results: Administration of FK506 significantly increased blood urea nitrogen and serum creatinine levels. In addition, FK506 has also reduced the renal content of reduced glutathione, as well as the enzymatic activities of superoxide dismutase and catalase. Furthermore, these effects were associated with an increase in lipid peroxidation, inducible NO-synthase (iNOS), and NF-kappa B expression. Moreover, histopathological examinations showed severe damage of the renal tissues in animals treated with FK506. Interestingly, it was found that concomitant administration of vitamin E along with FK506 ameliorated all these parameters and improved renal function. Furthermore, the immunosuppressive effect of FK506 was not affected by vitamin E.Conclusion: The findings of the present study suggest that concomitant use of vitamin E might be useful in reducing nephrotoxicity induced by FK506.

Cisplatin (cis-diammine dichloroplatinum (II), CDDP) is a widely used drug for treatment of various types of cancers. However, CDDP-induced nephrotoxicity remains the main dose-limiting side effect. Retinoids are a group of vitamin A-related compounds that exert their effects through retinoid receptors activation. In this study, we investigated the effect of CDDP treatment on retinoic acid receptor-α (RAR-α) and retinoid X receptor-α (RXR-α) expression. In addition, we investigated the possible modulatory effects of RAR agonist, all-trans retinoic acid (ATRA), on CDDP-induced nephrotoxicity. Rats were treated with saline, DMSO, CDDP, ATRA, or CDDP/ATRA. Twenty-four hours after the last ATRA injection, rats were killed; blood samples were collected; kidneys were dissected; and biochemical, immunohistochemical, and histological examinations were performed. Our results revealed that CDDP treatment significantly increased serum levels of creatinine and urea, with concomitant decrease in serum albumin. Moreover, reduced glutathione (GSH) content as well as superoxide dismutase (SOD) and catalase (CAT) activities were significantly reduced with concurrent increase in kidney malondialdehyde (MDA) content following CDDP treatment. Furthermore, CDDP markedly upregulated tubular RAR-α, RXR-α, fibrin, and inducible nitric oxide synthase (iNOS) protein expression. Although administration of ATRA to control rats did not produce marked alterations in kidney function parameters, administration of ATRA to CDDP-treated rats significantly exacerbated CDDP-induced nephrotoxicity. In addition, CDDP/ATRA co-treatment significantly increased RAR-α, RXR-α, fibrin, and iNOS protein expression compared to CDDP alone. In conclusion, we report, for the first time, the crucial role of retinoid receptors in CDDP-induced nephrotoxicity. Moreover, our findings indicate that co-administration of ATRA with CDDP, although beneficial on the therapeutic effects, their deleterious effects on the kidney may limit their clinical use.

Context: Cyclosporin A (CsA) is one of the most important immunosuppressive agents. However, its clinical use is strongly limited by several side effects including hepatotoxicity which remains a major clinical problem. Involvement of reactive oxygen species (ROS) in CsA-induced hepatotoxicity has been reported.Objective: This study investigates the potential protective role of wheat germ oil (WGO) as an antioxidant against CsA-induced hepatotoxicity.Materials and methods: Twenty-four male Wistar albino rats (six animals in each group) received castor oil, the vehicle of CsA i.p. (control) or either CsA (25 mg/kg/d i.p.), WGO (900 mg/kg/d by oral gavage), or CsA in combination with WGO daily for 21 d.Results: CsA administration significantly increased serum levels of the liver enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST). In addition, an increase in lipid peroxidation, inducible NO-synthase (iNOS), and NF-kappa B expression were observed in hepatic tissues of CsA-alone-treated rats. Furthermore, significant reduction in the hepatic content of reduced glutathione (GSH), superoxide dismutase (SOD), and catalase (CAT) was also observed in CsA-alone-treated animals. Moreover, histopathological changes occurred in CsA-alonetreated rats. Concomitant administration of WGO along with CsA improved all these parameters. Most interestingly, the immunosuppressive effect of CsA was not affected by WGO.Conclusion: The present study suggests that concomitant use of WGO might be useful in reducing liver toxicity induced by CsA via inhibition of ROS, iNOS, and NF-kB expression.

Transforming growth factor- (TGF- and downstream Smad signaling pathways have been found to be the most important pathways involved in tissue fibrosis via induction of the profibrotic genes connective tissue growth factor (CTGF) and tissue inhibitors of matrix metalloproteinase-1 (TIMP-1).Cyclosporin A (CsA) is one of the most efficient immunosuppressive drugs that has been shown to induce a rapid activation of TGF-Smad signaling pathway.The present work demonstrates that CsA similar to TGF- causes a rapid phosphorylation of Smad-2 in A549 lung epithelial cells which is abrogated by the addition of neutralizing TGF--antibody.By using the TGF-receptor I kinase inhibitor, activin receptor-like kinase-5 (ALK-5) a critical involvement of TGF- receptor in CsA-triggered Smad signaling was depicted.Furthermore, TGF-release and Smad-2 phosphorylation induced by CsA were highly reduced in the presence of the reactive oxygen species (ROS) scavenger N-acetyl-cysteine (NAC), indicating that ROS is required for TGF- release and Smad-2 phosphorylation induced by CsA.Moreover, the present work demonstrates that CsA via ROS generation and activation of TGF-/Smad signaling cascade can cause an increase in the expression of the profibrotic genes CTGF and TIMP-1 in A549 lung epithelial cells.Collectively, these data demonstrate that CsA causes a rapid activation of the fibrogenic Smad signaling cascade in A549 lung epithelial cells via generation of ROS and subsequent activation of latent TGFβ that is sufficient to elicit fibrogenic cell responses as indicated by increasing the expression of the fibrogenic genes CTGF and TIMP-1 that play an important role in tissue fibrosis.

Cyclosporin A (CsA) is one of the most important immunosuppressive agents and therefore is widely used in organ transplantation. However, the clinical use of CsA is strongly limited by several side effects including hepatotoxicity which remains a major clinical problem. Transforming growth factor-b (TGF-b) and downstream Smad signaling pathways have been found to play an important role in liver fibrosis via induction of profibrotic genes such as tissue inhibitors of matrix metallproteinases-1 (TIMP-1), and connective tissue growth factor (CTGF). The present work demonstrates that treatment of animals with CsA causes a rapid activation of TGF-b/Smad signalingcascade in rat liver as demonstrated by an increase in plasma TGFb level and Smad-2 phosphorylation. Activation of TGF-b/Smad signalingcascade was accompanied by activation of Smad-dependent expression of TIMP-1and CTGF. However, concomitant administration of neutralizing anti-TGF-b antibody markedly reduced Smad-2 phosphorylation as well as CTGF and TIMP-1 expression induced by CsA. Furthermore, it was found that administration of the antioxidant N-acetyl cysteine (NAC) along with CsA significantly reduced plasma TGF-b level, Smad-2 phosphorylation as well as CTGF and TIMP-1 expression. These data demonstrates for the first time that administration of CsA causes a rapid activation of TGF-b/Smad signaling pathway and subsequent CTGF and TIMP-1 expression in rat liver.

Several animal studies suggest a role of platelet-derived growth factors (PDGFs) particularly A and B in atherosclerosis. Previously, it has been shown that viral infections have the ability to initiate and accelerate atherosclerosis in animal models. Recently, it has been reported that IL-18 has a pro-atherogenic character. Moreover, viral infections have been shown to be associated with induction of IL-18 bioactivity. By using human predendritic KG1 cells, we sought to assess PDGF-AA production under the influence of IL-18 and the byproduct of viral replication, dsRNA-mimetic poly (I:C). Here we demonstrate that poly (I:C) and IL-18 have the ability to induce PDGF-AA expression. In addition, costimulation of KG-1 cells with both IL-18 plus poly (I:C) shows an additive effect on PDGF-AA production. Furthermore, we demonstrate that neither p38 nor SAPK/JNK is required for PDGF-AA production by both PIC and IL-18. However, the expression of PDGF-AA has been found to be associated with increased activation of NF-κB and enhancement of DNA-binding capacity of NF-κB as shown by electrophoretic mobility shift assay (EMSA) and supershift analysis. Collectively, this study demonstrates that the byproduct of viral replication, dsRNA [poly (I:C)], and IL-18 have the ability to induce PDGF-AA in NF-κB-dependent manner. Furthermore, dsRNA act in an additive way with IL-18 to induce PDGF-AA which plays a major role in atherosclerosis. These data might help to understand the pro-atherogenic character of IL-18 and molecular mechanisms of viral infection-induced atherosclerosis.