Perindopril sensitizes hepatocellular carcinoma to chemotherapy: A possible role of leptin / Wnt/ ß-catenin axis with subsequent inhibition of liver cancer stem cells.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death. The major challenge in managing HCC is the resistance to chemotherapy. Leptin hormone is associated with different oncogenic pathways implicated in drug resistance. Angiotensin II was found to decrease the production and secretion of leptin. Objective: This study investigated the potential role of an ACEI perindopril as a chemosensitizer agent to sorafenib. Method: HCC was induced in mice using a single dose of diethylnitrosamine DENA (200 mg/kg) followed by phenobarbital 0.05% in drinking water for 16 weeks. Mice were then treated with perindopril (1 mg/kg/day), Sorafenib (30 mg/kg/day), or both of them for another four weeks. Leptin, VEGF, MMP-9, Cyclin D1, EpCAM, and ß-catenin were measured using immunoassay while Wnt and ALDH1 were assayed using western blotting assay. Results: Perindopril whether alone or in combination with sorafenib decrease liver enzymes and preserve the liver architecture. Our study revealed that perindopril significantly increased the antineoplastic, antiangiogenic as well as anti-metastatic effects of sorafenib. This effect was correlated with the downregulation of the leptin / Wnt / ß-catenin pathway and overexpression of ALDH1 while downregulation of EpCAM. Conclusion: This study presents perindopril as a potential chemosensitizer agent that works through decreased expression of the leptin / Wnt / ß-catenin pathway.

Ocular films versus film-forming liquid systems for enhanced ocular drug delivery.

The short residence time, corneal barrier functions, and other effective eye protective mechanisms limited the ocular availability after topical application. Ocular inserts are being developed as polymer films for insertion into the conjunctival sac with the goal of increasing ocular availability. Unfortunately, these devices are not convenient for patients and are associated with many problems. The use of in situ gel/film-forming systems may provide promising alternative with comparable efficacy but this requires verification. Therefore, the current study compared ocular inserts with in situ film-forming liquids containing the same polymer components for ocular delivery of pilocarpine nitrate. Solvent casting technique was employed to prepare the inserts using and polyvinyl alcohol (PVA) as film-forming polymer blended with sodium alginate, as bioadhesive polymer. The effect of addition of either carboxymethycellulose, carbopol, polyvinylpyrrolidone, or methylcellulose was investigated. Solid-state characterization of the inserts indicated compatibility of the drug with film component. All inserts were of acceptable bioadhesive parameters and folding endurance that depended on the film composition. In vitro release studies reflected matrix diffusion kinetics for the film and liquid formulations. This confirms the in situ gelation of liquids. The calculated in vivo miotic pharmacokinetics parameters, using albino rabbits, reflected a better rank for the film but the difference was not statistically different from the in situ gel/film-forming systems. Ocular safety, as reflected by tear volume test, indicated acceptable safety of both liquid and inserts to the eye. The study suggested comparable efficacy of film-forming liquids to that of ocular films. Graphical abstract.

The potential beneficial effects of sildenafil and diosmin in experimentally-induced gastric ulcer in rats.

OBJECTIVES: research in the treatment of gastric ulcer has involved the investigation of protective drugs. These drugs may be used as adjacent therapy with the traditional pharmacologic treatment of peptic ulcer. The present study is designed to investigate the gastro protective effects of diosmin (DIO), sildenafil (SILD) and their combinations with ranitidine (RANT) against indomethacin (INDO)-induced gastric ulcer in rats. Additionally, the potential mechanisms of their effect are addressed. METHODS: DIO (100 mg/kg) and SILD (10 mg/kg) were administered by oral route for seven days prior to ulcer induction. Moreover, other rats were treated with RANT (50 mg/kg) not only to compare efficiency of the medications but also, to help clarify potential mechanisms of their effect. Following, after 24 h of fasting, INDO (100 mg/kg) was administered for induction of gastric ulcer. Furthermore, rats in each group were sacrificed 4 h later. Biochemical analysis of DIO, SILD, RANT and their combinations pre-treated host tissues demonstrated reduction in tumor necrosis factor (TNF)-α and malondialdehyde (MDA) contents and concomitant increase in gastric pH, nitric oxide (NO) and reduced glutathione (GSH) contents. RESULT: It is observed, that SILD and DIO pre-treatment showed non-significant effect on gastric juice PH. However, their combinations with RANT is superior to using RANT alone. In addition, the results revealed, that combinations of (RANT and SILD) and (RANT and DIO) showed the highest increase in gastric tissue NO levels. But, these two combinations achieved the lowest MDA levels relative to the control (INDO) group. Despite, all groups displayed non-significant effect on reduced GSH content, (RANT and SILD) group increased GSH concentration by 39.75% relative to INDO group. In addition, DIO, RANT and (RANT and DIO) pre-treatment have anti-apoptotic activity on gastric mucosa. On the other hand, SILD did not affect caspase-3 immunostaining. These results are confirmed by histopathological findings. CONCLUSION: The work outcomes provide a new gastro protective agents in clinical gastropathy. So, this study not only provides an efficient way for peptic ulcer protection, but also it may be considered for future studies in ulcer healing and gastric cancer.

Targeting MDR-1 gene expression, BAX/BCL2, caspase-3, and Ki-67 by nanoencapsulated imatinib and hesperidin to enhance anticancer activity and ameliorate cardiotoxicity.

There is a great demand to introduce new approaches into cancer treatment field due to incidence of increased breast cancer all over the world. The current study was designed to evaluate the role of imatinib mesylate (IM) and/or hesperidin (HES) nanoparticles alone or in combination in enhancing the anticancer activity and to investigate the ability of nanoencapsulation to reduce cardiotoxicity of IM in solid Ehrlich carcinoma (SEC)-bearing mice. IM and HES were loaded into PLGA (poly(lactic-co-glycolic acid) polymer. SEC was induced in female albino mice as a model for experimentally induced breast cancer. Mice were randomly divided into eight groups (n = 10). On day 28 from tumor inoculation, mice were sacrificed and blood samples were collected in heparinized tubes for hematological studies, biochemical determination of lactate dehydrogenase (LDH), and glutamic oxaloacetic transaminase (SGOT) levels. In addition, tumor and cardiac tissues were utilized for histopathological examination as well as determination of MDR-1 gene expression. Immunohistochemical staining of BAX and BCL-2 was done. Nano IM- and/or Nano HES-treated groups showed a significant reduction in tumor volume, weight, hematological, cardiac markers, and tumor MDR-1 gene downregulation compared to free conventional treated groups. In conclusion, the use of HES as an adjuvant therapy with IM could improve its cytotoxic effects and limit its cardiac toxicity. Furthermore, nanoencapsulation of IM and/or HES with PLGA polymer showed a remarkable anticancer activity.

Enhanced anticancer activity of combined treatment of imatinib and dipyridamole in solid Ehrlich carcinoma-bearing mice.

The current study was designed to evaluate potential enhancement of the anticancer activity of imatinib mesylate (IM) with dipyridamole (DIP) and to investigate the underlying mechanisms of the combined therapy (IM/DIP) to reduce hepatotoxicity of IM in solid Ehrlich carcinoma (SEC)-bearing mice. SEC was induced in female albino mice as a model for experimentally induced breast cancer. Mice were randomly divided into seven groups (n = 10): SEC vehicle, IM50 (50 mg/kg), IM100 (100 mg/kg), DIP (35 mg/kg), a combination of IM50/DIP and IM100/DIP. On day 28th, mice were sacrificed and blood samples were collected for hematological studies. Biochemical determination of liver markers was evaluated. Glutamic oxaloacetic transaminase (SGOT), glutamic pyruvic transaminase (SGPT) and alkaline phosphatase (ALP) levels were assessed. In addition, MDR-1 gene expression and immunohistochemical staining of BAX and BCL-2 was done. Also, in vitro experiment for determination of IC50 of different treatments and combination index (CI) were assessed in both MCF-7 and HCT-116 cell lines. IM- and/or DIP-treated groups showed a significant reduction in tumor volume, weight, and serum levels of SGOT, SGPT, and AIP compared to vehicle group. In addition, reduction of VEGF, Ki67, and adenosine contents was also reported by treated groups. Also, IM/DIP combination showed lower IC50 than monotherapy. Combination index is less than 1 for IM/DIP combination in both cell lines. DIP as an adjuvant therapy potentiated the cytotoxic effect of IM, ameliorated its hepatic toxicity, and showed synergistic effect with IM in vitro cell lines. Furthermore, the resistance against IM therapy may be overcome by the use of DIP independent on mdr-1 gene expression.

Daclatasvir and Sofosbuvir Mitigate Hepatic Fibrosis Through Downregulation of TNF-α / NF-κB Signaling Pathway.

BACKGROUND: Hepatic fibrosis is the major issue in chronic liver diseases such as chronic hepatitis C virus (HCV). The newly approved direct acting antiviral (DAA) agents such as Sofosbuvir (SOF) and daclatasvir (DAC) have been found to be associated with decreased fibrotic markers in HCV patients. AIM: This study tried to explore whether the reported antifibrotic effect of these drugs is antiviral dependent or drug induced. METHOD: Hepatic fibrosis was induced by (0.5ml/kg) CCl4 IP twice a week for six weeks. SOF (20 mg/kg/d) and DAC (30 mg/kg/d) were added in the last four weeks of treatments. Liver functions, fibrotic markers such as Hyaluronic acid and metalloproteinase-9 were detected using immunoassay. The expression of TNF-α/NF-κB signaling pathway as well as Bcl-2 were done using immunoassay. RESULTS: SOF and DAC exerted a potent antifibrotic effect evidenced by their activity against hyaluronic acid HA and metalloproteinase MMP-9 significantly (P≤0.001). This effect was further proved histopathologically where liver tissues from rats treated by drugs showed marked inhibition of collagen precipitation as well as inhibition of HSCs activation. This antifibrotic action was associated with decreased expression of TNF-α /NF-κB signaling pathway and induction of Bcl-2. CONCLUSION: SOF/ DAC antifibrotic effect is independent of its antiviral activity. The molecular events associated with this effect were the downregulation of TNF-α / NF-κB signaling pathway and induction of Bcl-2.

Protective effects of mirtazapine and chrysin on experimentally induced testicular damage in rats.

Mirtazapine is an antidepressant with prominent antioxidant effects. Chrysin, a natural flavone, exhibits multiple pharmacological actions. This study was designed to investigate the potential protective effects of chrysin and mirtazapine against nitrofurazone-induced testicular damage in rats. Possible underlying mechanisms such as oxidative stress, inflammation and apoptosis were also investigated. Testicular damage was induced by oral administration of nitrofurazone (50mg/kg/day) for two weeks. Chrysin (25 and 50mg/kg/day, p.o.) and mirtazapine (15 and 30mg/kg/day, p.o.) were applied for three weeks, starting one week before nitrofurazone administration. Prophylactic treatment with chrysin and mirtazapine attenuated the elevation of serum acid phosphatase enzyme activity and halted the decline of sperm count and sperm viability resulted from nitrofurazone administration. Moreover, both agents ameliorated nitrofurazone-induced lipid peroxidation, glutathione depletion, elevation in tumor necrosis factor-α level and reduction in c-kit level in rat testes. With respect to apoptosis, immunohistochemical analysis revealed that chrysin and mirtazapine reduced the expression of caspase-3 in testicular tissue which was induced by nitrofurazone. Histopathological findings further supported the protective effects of both drugs against nitrofurazone-induced testicular injury. These findings suggest that the cytoprotective effects of chrysin and mirtazapine on rat testes were associated with suppression of oxidative stress and apoptotic tissue damage. Generally, chrysin prophylactic treatment showed a superior testicular protection than mirtazapine at the tested doses.

Oxamate potentiates taxol chemotherapeutic efficacy in experimentally-induced solid ehrlich carcinoma (SEC) in mice.

Several human cancers including the breast display elevated expression of Lactate dehydrogenase-A (LDH-A), the enzyme that converts pyruvate to lactate and oxidizes NADH to NAD+. Indeed, tumor lactate levels correlate with increased metastasis, tumor recurrence, and poor outcome. Lactate also plays roles in promoting tumor inflammation and as a signaling molecule that stimulates tumor angiogenesis. Because of its essential role in cancer metabolism, LDH-A has been considered as a potential target for combination cancer therapy. Therefore, the current study investigated the possible anti-tumor effect of LDH inhibitor (oxamate) in a murine model of breast cancer [Solid Ehrlich Carcinoma (SEC)], alone and in combination with Taxol chemotherapy. The potential underlying mechanisms were also investigated. The results indicated that oxamate induced significant anti-tumor activity against the SEC. Mechanistically, the combination treatment was more efficient than paclitaxel monotherapy in reducing ATP, MDA, TNF-α and Il-17 contents in SEC. Moreover, the apoptotic and anti-angiogenic effects of the combination treatment were triggered more efficiently as compared to paclitaxel monotherapy, Therefore, oxamate may represent a promising agent that enhance the antitumor activity of paclitaxel.

Combination of tadalafil and diltiazem attenuates renal ischemia reperfusion-induced acute renal failure in rats.

Life threatening conditions characterized by renal ischemia/reperfusion (RIR) such as kidney transplantation, partial nephrectomy, renal artery angioplasty, cardiopulmonary bypass and aortic bypass surgery, continue to be among the most frequent causes of acute renal failure. The current study investigated the possible protective effects of tadalafil alone and in combination with diltiazem in experimentally-induced renal ischemia/reperfusion injury in rats. Possible underlying mechanisms were also investigated such as oxidative stress and inflammation. Rats were divided into sham-operated and I/R-operated groups. Anesthetized rats (urethane 1.3g/kg) were subjected to bilateral ischemia for 30min by occlusion of renal pedicles, then reperfused for 6h. Rats in the vehicle I/R group showed a significant (p˂0.05) increase in kidney malondialdehyde (MDA) content; myeloperoxidase (MPO) activity; TNF-α and IL-1ß contents. In addition significant (p˂0.05) increase in intercellular adhesion molecule-1(ICAM-1) content, BUN and creatinine levels, along with significant decrease in kidney superoxide dismutase (SOD) activity. In addition, marked diffuse histopathological damage and severe cytoplasmic staining of caspase-3 were detected. Pretreatment with combination of tadalafil (5mg/kg bdwt) and diltiazem (5mg/kg bdwt) resulted in reversal of the increased biochemical parameters investigated. Also, histopathological examination revealed partial return to normal cellular architecture. In conclusion, pretreatment with tadalafil and diltiazem combination protected against RIR injury.

PPARγ-dependent anti-tumor and immunomodulatory actions of pioglitazone.

Peroxisome proliferator-activated receptor-γ (PPARγ) has been reported to play important roles in carcinogenesis. The current study was carried out to assess the possible anti-tumor effects of pioglitazone (PIO), a PPARγ agonist, in a mouse mammary carcinoma model, i.e. a solid Ehrlich carcinoma (SEC). Effects of PIO on tumor-induced immune dysfunction, and the possibility that PIO may modulate the anti-tumor and immunomodulatory effects of doxorubicin (DOX) were also studied. The effects in tumor-bearing hosts of several doses of PIO (100 mg/kg, per os), with and without the added presence of DOX (2 mg/kg, IP), was investigated in vivo; end-points evaluated included assessment of tumor volume, splenic lymphocyte profiles/functionality, tumor necrosis factor (TNF)-α content, as well as apoptosis and expression of nuclear factor-κB (NF-κB) among the tumor cells. The data indicate that PIO induced significant anti-tumor activity against the SEC. PIO treatments also significantly mitigated both tumor- and doxorubicin-induced declines in immune parameters assessed here. Moreover, PIO led to decreased NF-κB nuclear expression, and, in doing so, appeared to chemo-sensitize these tumor cells to DOX-induced apoptosis. All pioglitazone-studied effects were antagonized by GW9662, a selective PPARγ antagonist.

Histamine protects against the acute phase of experimentally-induced hepatic ischemia/re-perfusion.

Histamine, involved in many inflammatory reactions and immune responses, is reported to suppress--via H4R stimulation--injury concomitant with the late phase of warm hepatic ischemia/re-perfusion (I/R). The current study investigated the possible effects of histamine on the acute phase of hepatic I/R injury, and the possible underlying mechanisms like oxidative stress and release of inflammatory cytokines (e.g., tumor necrosis factor (TNF)-α nd interleukin [IL]-12). Rats were divided into naïve, sham-operated, and I/R groups. The I/R group was divided into sub-groups and pre-treated with histaminergic ligands before induction of ischemia. Anesthetized rats were subjected to warm ischemia for 30 min by occlusion of the portal vein and hepatic artery, then re-perfused for 90 min. Rats in the control I/R group showed significant increases in hepatic malondialdehyde (MDA), TNFα, and IL-12 contents, and in plasma alanine transaminase (ALT) and aspartate transaminase (AST) levels, along with significant decreases in hepatic reduced glutathione (GSH) content and marked diffuse histopathologic damage. Pre-treatment with histamine resulted in significant mitigation of each of these end-points. The protective effect of histamine was not antagonized by pre-treatment with mepyramine (H1R antagonist) or ranitidine (H2R antagonist) and completely reversed by pre-treatment with thioperamide (H3R and H4R antagonist). In addition, the histamine protective effect was mimicked by pre-treatment of rats with clozapine (H4R agonist). These observations strongly suggested that histamine has a protective effect against hepatic I/R-mediated tissue injury during the acute phase, and this effect was mediated through an H4R stimulation that led to a decrease in IL-12 and TNFα production--outcomes that consequently decreased localized oxidative stress and afforded hepatic protection in general.

Effect of genetic polymorphisms on the development of secondary failure to sulfonylurea in egyptian patients with type 2 diabetes.

OBJECTIVE: This study investigated the possibility that genetic factors, such as polymorphism of K inward rectifier subunit (Kir6.2), E23K, and Arg(972) polymorphism of insulin receptor sub-strate-1 (IRS-1), may predispose patients to sulfonylurea failure. METHODS: A total of 100 unrelated Egyptian patients with type 2 diabetes were recruited. They were divided into two equal groups: group I consisted of patients with secondary failure to sulfonylurea (hemoglobin A(1c) ≥ 8% despite sulfonylurea therapy) while group II consisted of patients whose condition was controlled with oral therapy. RESULTS: Of all the patients, 45% and 14% were carriers of the K allele and Arg(972) variants respectively. The frequency of the K allele was 34% among patients with diabetes that was controlled with oral therapy and 56% among patients with secondary failure to sulfonylurea. The frequency of the Arg(972) IRS-1 variant was 6% among patients with diabetes controlled with oral therapy and 22% among patients with secondary failure. CONCLUSION: The E23K variant of the Kir6.2 gene and Arg(972) IRS-1 variants are associated with increased risk for secondary failure to sulfonylurea.

Insulinotropic and anti-inflammatory effects of rosiglitazone in experimental autoimmune diabetes.

Cytokines and nitric oxide (NO) are involved in the pathogenesis of autoimmune diabetes mellitus (DM). Rosiglitazone is an insulin-sensitizing drug that is a ligand for the nuclear receptor peroxisome proliferator-activated receptor-gamma (PPAR-gamma). The anti-inflammatory and immunomodulating properties of PPAR-gamma have been documented. The aim of this study is to investigate the effectiveness of rosiglitazone in autoimmune DM and to clarify the possible mechanism(s) involved. Autoimmune DM was induced in adult male Balb/c mice by co-administration of cyclosporin A and multiple low doses of streptozotocin. Diabetic mice were treated daily with rosiglitazone (7 mg/kg, p.o.) for 21 days. Blood glucose level (BGL), serum insulin level and pancreatic levels of tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma) and NO were measured. Histopathological examination and immunohistochemical determination of CD4 and CD8 T lymphocytes in the pancreatic islets were performed. In addition, analysis of pancreatic protein expression was carried out. The results showed that rosiglitazone treatment resulted in a significant decrease in the BGL and the pancreatic levels of TNF-alpha, IFN-gamma and NO compared to diabetic mice. The serum insulin level was significantly increased after rosiglitazone treatment compared to diabetic mice. The destroyed pancreatic islets were regenerated and became free from both CD4 and CD8 T cells after treatment. Furthermore, many changes in pancreatic protein expression were observed. These results suggest that rosiglitazone has a beneficial effect in the treatment of autoimmune diabetes, an effect that seemed to be a secondary consequence of its anti-inflammatory and immunomodulating properties and might be reflected at the level of protein expression.

The potential role of cyclooxygenase-2 inhibitors in the treatment of experimentally-induced mammary tumour: does celecoxib enhance the anti-tumour activity of doxorubicin?

The potential anti-tumour activity of non-steroidal anti-inflammatory drugs (NSAIDS) has been previously discussed. This study was undertaken to assess the possible anti-tumour activity of the cyclooxygenase-2 (COX-2) inhibitor; celecoxib in an animal model of mammary carcinoma; the solid Ehrlich carcinoma (SEC). The possibility that celecoxib may modulate the anti-tumour activity of doxorubicin on the SEC was also studied. Some of the possible mechanisms underlying such modulation were investigated. The anti-tumour activity of celecoxib (25 mg kg(-1)), diclofenac (12.5 mg kg(-1)) and doxorubicin (2 mg kg(-1)) either alone or in combination were investigated on SEC in vivo through the assessment of tumour growth delay (TGD) and tumour volume (TV), changes in tumour DNA content and nitric oxide (NO) levels, immunohistochemical staining of the tumour suppressor gene product; p53 histopathological examination and determination of apoptotic index of SEC. In addition, the influence of these drugs on the DNA fragmentation pattern of Ehrlich carcinoma cells (ECC) was studied. It was found that both celecoxib and diclofenac lack the anti-tumour activity on SEC. In addition there was a significant increase in doxorubicin anti-tumour activity when administered in combination with celecoxib. Moreover, it was found that both celecoxib and diclofenac have the potential to inhibit the function of P-glycoprotein (P-gp) in ECC using rhodamine uptake and efflux assays. Therefore, the current study suggested the chemosensitizing potential of celecoxib in the SEC animal model of mammary tumour, which could be explained in part on the basis of inhibition of P-gp function, with possible enhancement of doxorubicin anti-tumour activity.