Synthesis of a novel series of (Z)-3,5-disubstituted thiazolidine-2,4-diones as promising anti-breast cancer agents.

A novel series of (Z)-3,5-disubstituted thiazolidine-2,4-diones 4-16 has been designed and synthesized. Preliminary screening of these compounds for their anti-breast cancer activity revealed that compounds 5, 7, and 9 possess the highest anti-cancer activities. The anti-tumor effects of compounds 5, 7, and 9 were evaluated against human breast cancer cell lines (MCF-7 and MDA-MB-231) and human breast cancer cells. They were also evaluated against normal non-cancerous breast cells, isolated from the same patients, to conclude about their use in a potential targeted therapy. Using MTT uptake method, these three compounds 5, 7, and 9 blunt the proliferation of these cancer cells in a dose-dependent manner with an IC50 of 1.27, 1.50 and 1.31 µM respectively. Interestingly, using flow cytometry analysis these three compounds significantly mediated apoptosis of human breast cancer cells without affecting the survival of normal non-cancerous breast cells that were isolated from the same patients. Mechanistically, these compounds blunt the proliferation of MCF-7 breast cancer cells by robustly decreasing the phosphorylation of AKT, mTOR and the expression of VEGF and HIF-1α. Most importantly, compounds 5, 7, and 9 without affecting the phosphorylation and expression of these crucial cellular factors in normal non-cancerous breast cells that were isolated from the same patients. Additionally, using Western blot analysis the three compounds significantly (P < 0.05) decreased the expression of the anti-apoptotic Bcl-2 members (Bcl-2, Bcl-XL and Mcl-1) and increased the expression of the pro-apoptotic Bcl-2 members (Bak, Bax and Bim) in MCF-7, MDA-MB-231 and human breast cancer cells making these breast cancer cells susceptible for apoptosis induction. Taken together, these data provide great evidences for the inhibitory activity of these compounds against breast cancer cells without affecting the normal breast cells.

Enhanced susceptibility to apoptosis and growth arrest of human breast carcinoma cells treated with silica nanoparticles loaded with monohydroxy flavone compounds.

The treatment of drug-resistant cancer is a clinical challenge, hence screening for novel anticancer drugs is critically important. In this study, we investigated the anti-tumor potential of three plant-derived flavone compounds: 3-hydroxy flavone (3-HF), 6-hydroxy flavone (6-HF), and 7-hydroxy flavone (7-HF), either alone or combined with silica nanoparticles (3-HF + NP, 6-HF + NP, and 7-HF + NP), on the human breast carcinoma cell lines MDA-MB-231 and MCF-7, as well as on non-tumorigenic normal breast epithelial cells (MCF-10). The IC50 values of these flavone compounds loaded with NP (flavones + NP) in these cell lines were determined to be 1.5 µg/mL without affecting the viability of normal MCF-10 cells. Additionally, using annexin V - propidium iodide double-staining followed by flow cytometry analysis, we found that the combination of flavones with NP significantly induced apoptosis in MCF-7 and MDA-MB-231 cancer cells. Furthermore, flavones + NP increased the expression of cytochrome c and caspase-9, mediating the growth arrest of these cancer cells. Most importantly, the combination of flavones with NP significantly abolished the expression of ATF-3, which is responsible for the proliferation and invasion of bone-metastatic breast cancer cells. Our data revealed the potential therapeutic effects of these flavones in fighting breast cancer cells, and provide the first insights concerning the underlying molecular mechanisms.

The Therapeutic Mechanisms of Propolis Against CCl4 -Mediated Liver Injury by Mediating Apoptosis of Activated Hepatic Stellate Cells and Improving the Hepatic Architecture through PI3K/AKT/mTOR, TGF-ß/Smad2, Bcl2/BAX/P53 and iNOS Signaling Pathways.

BACKGROUND/AIMS: Propolis is one of the most promising natural products, exhibiting not only therapeutic but also prophylactic actions. Propolis has several biological and pharmacological properties, including hepatoprotective activities. The present study aimed to investigate the underlying molecular mechanisms of propolis against CCl4-mediated liver fibrosis. METHODS: Three groups of male BALB/c mice (n=15/ group) were used: group 1 comprised control mice; groups 2 and 3 were injected with CCl4 for the induction of liver fibrosis. Group 3 was then orally supplemented with propolis (100 mg/kg body weight) for four weeks. Different techniques were used to monitor the antifibrotic effects of propolis, including histopathological investigations using H&E, Masson's trichrome and Sirius red staining; Western blotting; flow cytometry; and ELISA. RESULTS: We found that the induction of liver fibrosis by CCl4 was associated with a significant increase in hepatic collagen and α-smooth muscle actin (α-SMA) expression. Moreover, CCl4-treated mice also exhibited histopathological alterations in the liver architecture. Additionally, the liver of CCl4-treated mice exhibited a marked increase in proinflammatory signals, such as increased expression of HSP70 and increased levels of proinflammatory cytokines and ROS. Mechanistically, the liver of CCl4-treated mice exhibited a significant increase in the phosphorylation of AKT and mTOR; upregulation of the expression of BAX and cytochrome C; downregulation of the expression of Bcl2; a significant elevation in the levels of TGF-ß followed by increased phosphorylation of SMAD2; and a marked increase in the expression of P53 and iNOS. Interestingly, oral supplementation of CCl4-treated mice with propolis significantly abolished hepatic collagen deposition, abrogated inflammatory signals and oxidative stress, restored CCl4-mediated alterations in the signaling cascades, and hence repaired the hepatic architecture nearly to the normal architecture observed in the control mice. CONCLUSION: Our findings revealed the therapeutic potential and the underlying mechanisms of propolis against liver fibrosis.

Camel Whey Protein Disrupts the Cross-Talk Between PI3K and BCL-2 Signals and Mediates Apoptosis in Primary Acute Myeloid Leukemia Cells.

In the present study, we investigated the impact of camel whey protein (CWP) on the survival of primary acute myeloid leukemia (AML) cells that were isolated from 20 patients diagnosed with AML. We found that CWP induced apoptosis in the primary AML cells without affecting the normal PBMCs that were isolated from healthy individuals, as determined by PI/annexin V double staining followed by flow-cytometry analysis. Furthermore, we demonstrated that these primary AML cells exhibited aberrant phosphorylation of AKT, mTOR and STAT3. Treatment of AML cells with CWP mediated significant reduction in the phosphorylation of AKT, mTOR and STAT3. Additionally, we demonstrated that blockade of PI3K/AKT signaling pathway by wortmannin (WM) impaired the expression of Bcl-2 and BclXL in the primary AML cells, suggesting an essential cross-talk between PI3K and Bcl-2 that maintains the survival of AML cells. In this context, treatment of AML cells with CWP disrupted the PI3K/Bcl-2 cross-talk; significantly downregulated the expression of anti-apoptotic Bcl-2 family members Bcl-2 and BclXL; markedly upregulated the expression of the pro-apoptotic Bcl-2 family members Bak and Bax; and subsequently sensitized tumor cells to growth arrest. Our data revealed the therapeutic potential of CWP and the underlying mechanisms against leukemia.

Camel whey protein protects lymphocytes from apoptosis via the PI3K-AKT, NF-κB, ATF-3, and HSP-70 signaling pathways in heat-stressed male mice.

Heat stress (HS) is an environmental factor that depresses the immune systems that mediate dysfunctional immune cells. Camel whey protein (CWP) can scavenge free radicals and enhance immunity. This study investigated the impact of dietary supplementation with CWP on immune dysfunction induced by exposure to HS. Male mice (n = 45) were distributed among 3 groups: control group; HS group; and HS mice that were orally administered CWP (HS + CWP group). The HS group exhibited elevated levels of reactive oxygen species (ROS) and pro-inflammatory cytokines (interleukin (IL)-1ß, IL-6, tumor necrosis factor-α) as well as a significant reduction in the IL-2 and IL-4 levels. Exposure to HS resulted in impaired phosphorylation of AKT and IκB-α (nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha); increased expression of activating transcription factor 3 (ATF-3) and 70 kDa heat shock proteins (HSP70); and aberrant distribution of CD3+ T cells and CD20+ B cells in the thymus and spleen. Interestingly, HS mice treated with CWP presented significantly restored levels of reactive oxygen species and pro-inflammatory cytokines near the levels observed in the control mice. Furthermore, supplementation of HS mice with CWP enhanced the phosphorylation of AKT and IκB-α; attenuated the expression of ATF-3, HSP70, and HSP90; and improved T and B cell distributions in the thymus and spleen. Our findings reveal a potential immunomodulatory effect of CWP in attenuating immune dysfunction induced by exposure to thermal stress.

Protective effects of camel whey protein against scrotal heat-mediated damage and infertility in the mouse testis through YAP/Nrf2 and PPAR-gamma signaling pathways.

Elevation of scrotal temperature is one of the most important causes of impaired spermatogenesis and male infertility, but the exact mechanism remains controversial. The present study investigated the impact of camel whey protein (CWP) on the mechanisms of heat stress (HS)-mediated testicular damage in male mice. Exposure to HS was associated with significant increase in the testicular tissues' oxidative stress. Mechanistically, exposure to HS resulted in upregulation of P53 and Nrf2 expressions; downregulation of Bcl2 and PPAR-γ expressions; and induction of testicular Leydig cell hyperplasia. Because Leydig cells produce testosterone up on stimulation with Luteinizing hormone (LH), HS mice also exhibited significant reduction in the serum testosterone levels followed by significant reduction in the percentages of progressively motile sperm and higher percentages of immotile sperm, when compared with those of control mice. Interestingly, treatment of HS mice with CWP significantly restored the levels of ROS and the activities of antioxidant enzymes in the testicular tissues nearly to those observed in control mice. Furthermore, CWP supplemented HS mice exhibited complete restoration of Bcl2, P53, Nrf2, and PPAR-γ expressions; testicular Leydig cell distribution; significant higher levels of testosterone levels; and hence higher percentages of progressively motile sperm and lower percentages of immotile sperm as compared to HS mice. Our findings reveal the protective effects of CWP against testis injury and infertility induced by exposure to HS by rescuing functional Leydig cells. Additionally, the present study has shed light on the molecular mechanisms underlying improved testicular damage following CWP treatment.

Curcumin analogue 1,5-bis(4-hydroxy-3-((4-methylpiperazin-1-yl)methyl)phenyl)penta-1,4-dien-3-one mediates growth arrest and apoptosis by targeting the PI3K/AKT/mTOR and PKC-theta signaling pathways in human breast carcinoma cells.

Recent developments in the literature have demonstrated that curcumin exhibit antioxidant properties supporting its anti-inflammatory, chemopreventive and antitumoral activities against aggressive and recurrent cancers. Despite the valuable findings of curcumin against different cancer cells, the clinical use of curcumin in cancer treatment is limited due to its extremely low aqueous solubility and instability, which lead to poor in vivo bioavailability and limited therapeutic effects. We therefore focused in the present study to evaluate the anti-tumor potential of curcumin analogues on the human breast carcinoma cell lines MDA-MB-231 and MCF-7, as well as their effects on non-tumorigenic normal breast epithelial cells (MCF-10). The IC50 values of curcumin analogue J1 in these cancer cell lines were determined to be 5 ng/ml and 10 ng/ml, in MDA-MB-231 and MCF-7 cells respectively. Interestingly, at these concentrations, the J1 did not affect the viability of non-tumorigenic normal breast epithelial cells MCF-10. Furthermore, we found that J1 strongly induced growth arrest of these cancer cells by modulating the mitochondrial membrane potentials without significant effect on normal MCF-10 cells using JC-1 staining and flow cytometry analysis. Using annexin-V/PI double staining assay followed by flow cytometry analysis, we found that J1 robustly enhanced the induction of apoptosis by increasing the activity of caspases in MDA-MB-231 and MCF-7 cancer cells. In addition, treatment of breast cancer cells with J1 revealed that, in contrast to the expression of cyclin B1, this curcumin analogue vigorously decreased the expression of cyclin A, CDK2 and cyclin E and subsequently sensitized tumor cells to cell cycle arrest. Most importantly, the phosphorylation of AKT, mTOR and PKC-theta in J1-treated cancer cells was markedly decreased and hence affecting the survival of these cancer cells. Most interestingly, J1-treated cancer cells exhibited a significant inhibition in the activation of RhoA followed by reduction in actin polymerization and cytoskeletal rearrangement in response to CXCL12. Our data reveal the therapeutic potential of the curcumin analogue J1 and the underlying mechanisms to fight breast cancer cells.

Wound healing: time to look for intelligent, 'natural' immunological approaches?

There is now good evidence that cytokines and growth factors are key factors in tissue repair and often exert anti-infective activities. However, engineering such factors for global use, even in the most remote places, is not realistic. Instead, we propose to examine how such factors work and to evaluate the reparative tools generously provided by 'nature.' We used two approaches to address these objectives. The first approach was to reappraise the internal capacity of the factors contributing the most to healing in the body, i.e., blood platelets. The second was to revisit natural agents such as whey proteins, (honey) bee venom and propolis. The platelet approach elucidates the inflammation spectrum from physiology to pathology, whereas milk and honey derivatives accelerate diabetic wound healing. Thus, this review aims at offering a fresh view of how wound healing can be addressed by natural means.

Camel Whey Protein Protects B and T Cells from Apoptosis by Suppressing Activating Transcription Factor-3 (ATF-3)-Mediated Oxidative Stress and Enhancing Phosphorylation of AKT and IκB-α in Type I Diabetic Mice.

BACKGROUND: Diabetes mellitus (DM) is associated with severe immune system complications. Camel whey protein (CWP) decreases free radicals (ROS) and modulates immune functions, but its effect on DM-impaired immune systems has not been studied. We investigated the impact of CWP on the immune system in a Type 1 diabetes mouse model. METHODS: Three experimental groups were used: (1) non-diabetic control; (2) diabetic; and (3) CWP-treated diabetic mice. RESULTS: Induction of diabetes by streptozotocin was associated with reduction of body weight and insulin level, increase in glucose level and pro-inflammatory cytokines (IL-1ß, IL-6, and TNF-α), and reduction in IL-2 and IL-4 levels. Upregulated ATF-3 expression was followed by a marked elevation in ROS levels. Lymphocytes from diabetic mice exhibited increased apoptosis through decreased phosphorylation of AKT and IκB-α, increased infiltration of T cells in the spleen and thymus, and decreased B cell numbers in the spleen. Supplementation with CWP decreased the levels of proinflammatory cytokines, ROS, and ATF-3 expression, and increased the levels of IL-4. Treatment with CWP decreased apoptosis by enhancing the phosphorylation of AKT and IκB-α as well as T-cell and B-cell distribution in the spleen and thymus. CONCLUSIONS: Our findings suggest the beneficial effects of CWP supplementation during diabetes on decreasing and orchestrating the redox status and subsequently rescuing the immune cells from exhaustion.

Bee Venom Accelerates Wound Healing in Diabetic Mice by Suppressing Activating Transcription Factor-3 (ATF-3) and Inducible Nitric Oxide Synthase (iNOS)-Mediated Oxidative Stress and Recruiting Bone Marrow-Derived Endothelial Progenitor Cells.

Multiple mechanisms contribute to impaired diabetic wound healing including impaired neovascularization and deficient endothelial progenitor cell (EPC) recruitment. Bee venom (BV) has been used as an anti-inflammatory agent for the treatment of several diseases. Nevertheless, the effect of BV on the healing of diabetic wounds has not been studied. Therefore, in this study, we investigated the impact of BV on diabetic wound closure in a type I diabetic mouse model. Three experimental groups were used: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice treated with BV. We found that the diabetic mice exhibited delayed wound closure characterized by a significant decrease in collagen production and prolonged elevation of inflammatory cytokines levels in wounded tissue compared to control non-diabetic mice. Additionally, wounded tissue in diabetic mice revealed aberrantly up-regulated expression of ATF-3 and iNOS followed by a marked elevation in free radical levels. Impaired diabetic wound healing was also characterized by a significant elevation in caspase-3, -8, and -9 activity and a marked reduction in the expression of TGF-ß and VEGF, which led to decreased neovascularization and angiogenesis of the injured tissue by impairing EPC mobilization. Interestingly, BV treatment significantly enhanced wound closure in diabetic mice by increasing collagen production and restoring the levels of inflammatory cytokines, free radical, TGF-ß, and VEGF. Most importantly, BV-treated diabetic mice exhibited mobilized long-lived EPCs by inhibiting caspase activity in the wounded tissue. Our findings reveal the molecular mechanisms underlying improved diabetic wound healing and closure following BV treatment. J. Cell. Physiol. 231: 2159-2171, 2016. © 2016 Wiley Periodicals, Inc.

Thymoquinone Rescues T Lymphocytes from Gamma Irradiation-Induced Apoptosis and Exhaustion by Modulating Pro-Inflammatory Cytokine Levels and PD-1, Bax, and Bcl-2 Signaling.

BACKGROUND/AIMS: Recent studies have shown that thymoquinone (TQ) exerts protective effects against ionizing radiation-induced cataracts in lens after total cranium irradiation of rats. Nevertheless, there is no published work investigated the effects of TQ on T cell development and biology in animal models exposed to gamma radiation. Therefore, in the present study we focused on determining the effects of TQ on radiation damage in the thymus, radiation-induced T cell imbalance, and on immune dysfunction induced by gamma-rays. METHODS: Three groups of rats were used: a control group, a gamma-irradiated group, and a gamma-irradiated group that was orally supplemented with TQ. Serum lipid profiles, malondialdehyde (MDA) levels, and pro-inflammatory cytokine levels were measured to assess gamma irradiation-induced oxidative stress and inflammatory capacity. T cell apoptosis was evaluated by annexin V/propidium iodide staining followed by flow cytometry analysis. The expression of pro-apoptotic proteins such as Bax and caspase-3, the anti-apoptotic protein Bcl-2, and an exhaustion marker of T cells (PD-1) in CD4+ and CD8+ T cell populations was evaluated using flow cytometry analysis. The T cell architecture of the thymus gland was evaluated by histological analysis. RESULTS: Exposure to gamma radiation increased triglyceride, cholesterol, LDL-C, MDA, TNF-α and IL-6 levels and decreased HDL-C levels. The altered lipid profile and MDA and pro-inflammatory cytokine (TNF-α and IL-6) levels induced by exposure to gamma radiation were significantly restored in TQ-treated gamma-irradiated rats. Rats exposed to gamma radiation exhibited increased exhaustion of T lymphocytes via down-regulation of Bcl-2 expression and upregulation of PD-1, Bax, and caspase-3 expression, which sensitized these cells to apoptosis. Interestingly, treatment of gamma-irradiated rats with TQ decreased T cell exhaustion and apoptosis by modulating the expression of Bcl-2, PD-1, Bax, and caspase-3. CONCLUSIONS: Our results provide evidence for the beneficial effects of TQ as an effective radioprotective candidate that enhances cellular immunity.

Oral supplementation of diabetic mice with propolis restores the proliferation capacity and chemotaxis of B and T lymphocytes towards CCL21 and CXCL12 by modulating the lipid profile, the pro-inflammatory cytokine levels and oxidative stress.

BACKGROUND: Type 1 diabetes mellitus (T1D) is a chronic autoimmune disease caused by the selective destruction of pancreatic ß cells, followed by hyperglycemia, oxidative stress and the subsequent extensive impairment of immune cell functions, a phenomenon responsible for the development of chronic diabetic complications. Propolis, a natural bee product that is extensively used in foods and beverages, significantly benefits human health. Specifically, propolis exerts antioxidant, anti-inflammatory and analgesic effects that may improve diabetic complications. To further elucidate the potential benefits of propolis, the present study investigated the effect of dietary supplementation with propolis on the plasma cytokine profiles, free radical levels, lipid profile and lymphocyte proliferation and chemotaxis in a streptozotocin (STZ)-induced type I diabetic mouse model. METHODS: Thirty male mice were equally distributed into 3 experimental groups: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice supplemented daily with an ethanol-soluble derivative of propolis (100 mg/kg body weight) for 1 month. RESULTS: First, the induction of diabetes in mice was associated with hyperglycemia and significant decreases in the insulin level and the lymphocyte count. In this context, diabetic mice exhibited severe diabetic complications, as demonstrated by a significant decrease in the levels of IL-2, IL-4 and IL-7, prolonged elevation of the levels of pro-inflammatory cytokines (IL-1ß, IL-6 and TNF-α) and reactive oxygen species (ROS) and altered lipid profiles compared with control non-diabetic mice. Moreover, antigen stimulation of B and T lymphocytes markedly reduced the proliferative capacity and chemotaxis of these cells towards CCL21 and CXCL12 in diabetic mice compared with control mice. Interestingly, compared with diabetes induction alone, treatment of diabetic mice with propolis significantly restored the plasma cytokine and ROS levels and the lipid profile to nearly normal levels. Most importantly, compared with untreated diabetic mice, diabetic mice treated with propolis exhibited significantly enhanced lymphocyte proliferation and chemotaxis towards CCL21 and CXCL12. CONCLUSION: Our findings reveal the potential immuno-modulatory effects of propolis, which acts as a natural antioxidant to enhance the function of immune cells during diabetes.

Topical application of propolis enhances cutaneous wound healing by promoting TGF-beta/Smad-mediated collagen production in a streptozotocin-induced type I diabetic mouse model.

BACKGROUND/AIMS: Impaired wound healing is considered to be one of the most serious complications associated with diabetes as it significantly increases the susceptibility of patients to infection. Propolis is a natural bee product used extensively in foods and beverages that has significant benefits to human health. In particular, propolis has antioxidant, anti-inflammatory and analgesic effects that could be useful for improving wound healing. In this study, we investigated the effects of topical application of propolis on the healing and closure of diabetic wounds in a streptozotocin (STZ)-induced type I diabetic mouse model. METHODS: Sixty male mice were distributed equally into 3 experimental groups: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice treated daily with a topical application of propolis. RESULTS: We found that diabetic mice exhibited delayed wound closure characterized by a significant decrease in the levels of TGF-ß1 and a prolonged elevation of the levels of inflammatory cytokines (IL-1ß, IL-6 and TNF-α) and MMP9 in wound tissues compared with control non-diabetic mice. Moreover, the wound tissues of diabetic mice showed a marked reduction in the phosphorylation of Smad2 and Smad3 as well as a marked reduction in collagen production. Interestingly, compared with untreated diabetic mice, topical application of propolis significantly enhanced the closure of diabetic wounds and decreased the levels of IL-1ß, IL-6, TNF-α and MMP9 to near normal levels. Most importantly, compared with untreated diabetic mice, the treatment of diabetic mice with propolis significantly enhanced the production of collagen via the TGF-ß1/Smad2,3 signaling axis in wounded tissues. CONCLUSION: Our findings reveal the molecular mechanisms underlying the improved healing and closure of diabetic wounds following topical propolis application.

Increased levels of type 1 interferon in a type 1 diabetic mouse model induce the elimination of B cells from the periphery by apoptosis and increase their retention in the spleen.

BACKGROUND: The autoimmune disease type 1 diabetes mellitus (T1D) is associated with a defect in the immune response, which increases susceptibility to infection. We recently demonstrated that prolonged elevated levels of type 1 interferon (IFN) induce lymphocyte exhaustion during T1D. AIMS: In the present study, we further investigated the effect of blocking the type I IFN receptor signaling pathway on diabetic dyslipidemia, in which an abnormal lipid profile leads to the exhaustion of B cells and alteration of their distribution and functions. METHODS: T1D was induced in a mouse model by an intraperitoneal injection of a single dose (60 mg/kg) of streptozotocin (STZ). Three groups of mice were examined: a non-diabetic control group, a diabetic group and a diabetic group treated with an anti-IFN (alpha, beta and omega) receptor 1 (IFNAR1) blocking antibody to block type I IFN signaling. RESULTS: We observed that induction of T1D was accompanied by a marked destruction of ß cells and a reduction in the insulin levels in the diabetic group. Diabetic mice exhibited many changes, including alterations in their lipid profiles, expansion of splenic B cells, increased caspase-3, -8 and -9 activity, and apoptosis in peripheral B cells. Blocking type 1 IFN signaling in diabetic mice significantly returned the insulin and lipid profiles to normal levels, subsequently restored the B cell distribution, and rescued the peripheral B cells from apoptosis. CONCLUSION: Our data suggest the potential role of type I IFN in mediating diabetic dyslipidemia and an exhausted state of B cells during T1D.

Cytotoxic cardenolides from the latex of Calotropis procera.

Three new cardenolides (3, 9 and 10), along with eight known ones, were isolated from the latex of Calotropis procera. The structural determination was accomplished by the 1D- and 2D-NMR spectra as well as HRESIMS analysis. The growth inhibitory activity of the latex and its sub-fractions as well as isolated compounds was evaluated against human A549 and Hela cell lines. The results exhibited that latex had strong growth inhibitory activity with IC50s of (3.37 µM, A-549) and (6.45 µM, Hela). Among the four extracts (hexane, chloroform, ethyl acetate and aqueous), chloroform extract displayed the highest potential cytotoxic activity, with IC50s of (0.985 µM, A-549) and (1.471 µM, Hela). All the isolated compounds displayed various degrees of cytotoxic activity and the highest activity was observed by calactin (1) with IC50s values of (0.036 µM, A-549) and (0.083 µM, Hela). None of these isolated compounds exhibited good antimicrobial activity evaluated by determination of their MICs using the broth microdilution method against various infectious pathogens. The structure-activity relationships for cytotoxic activity were also discussed.

Effect of a high fat, high sucrose diet on the promotion of non-alcoholic fatty liver disease in male rats: the ameliorative role of three natural compounds.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is a multifactorial disease with a complex pathophysiology. The clinical features of NAFLD include obesity, insulin resistance (IR) and dyslipidemia. Consumption of a diet high in saturated fats and sucrose is an important factor in the increasing occurrence of these metabolic disorders, primarily NAFLD and IR. We sought to assess the role of a high-fat, high-sucrose (HFS) diet in the promotion of NAFLD and to evaluate the effects of quercetin (Q), berberine (BB) and o-coumaric acid (CA) on modulation of these disorders. METHODS: Fifty male rats were divided into 2 main groups as follows: group 1 comprised 10 rats fed a standard diet (SD), and group 2 comprised 40 rats fed an HFS diet for 6 weeks and then subdivided equally into 4 groups; one of these groups served as the HFS diet and each of the other three groups received daily supplementation with either Q, CA or BB for 6 weeks. RESULTS: In the present study, several metabolic disorders were induced in our laboratory animal model, as evidenced by histological and biochemical changes. These alterations included serum and hepatic dyslipidemia (i.e., increased triglyceride, total cholesterol and low-density lipoprotein levels and decreased high-density lipoprotein levels), alterations in metabolic enzyme activities (lipase, glycerol-3-phosphate dehydrogenase, and glucose-6-phosphate dehydrogenase), histological changes in the liver (micro- and macrovesicular steatosis) and the downregulation of peroxisome proliferator-activated receptor γ (PPARγ) in adipose tissue and the liver. Daily oral supplementation with Q, CA or BB for 6 weeks after NAFLD induction had a hypolipidemic action and modulated metabolic markers. CONCLUSION: We showed that an HFS diet is able to promote NAFLD, and our results suggest that CA and BB are promising complementary supplements that can ameliorate the metabolic disorders associated with an HFS diet; however, Q requires further investigation.

Increased susceptibility to apoptosis and growth arrest of human breast cancer cells treated by a snake venom-loaded silica nanoparticles.

BACKGROUND: The development of effective treatments against metastatic cancers, including breast cancer, is among the most important challenges in current experimental and clinical cancer research. We recently demonstrated that Walterinnesia aegyptia venom (WEV), either alone or in combination with silica nanoparticles (WEV+NP), resulted in the growth arrest and apoptosis of different cancer cell lines. AIMS: In the present study, we evaluated the impact of WEV alone and WEV+NP on human breast cancer cells isolated from cancer biopsies. METHODS: The potential effects of WEV alone and WEV+NP on the proliferation, induction of apoptosis and generation of free radicals in breast cancer cells isolated from 80 patients clinically diagnosed with breast cancer were evaluated by flow cytometry and ELISA. RESULTS: WEV alone and WEV+NP inhibited the proliferation, altered the cell cycle and enhanced the induction of apoptosis of the breast cancer cells by increasing the activities of caspase-3, caspase-8 and caspase-9. In addition, the combination of WEV and NP robustly sensitized the breast cancer cells to growth arrest and apoptosis by increasing the generation of free radicals, including reactive oxygen species (ROS), hydroperoxide and nitric oxide. The combination of WEV with NP significantly enhanced the anti-tumor effect of WEV in breast cancer cells. CONCLUSION: Our data indicate the therapeutic potential of the nanoparticle-sustained delivery of snake venom for the treatment of breast cancer.

Snake (Walterinnesia aegyptia) venom-loaded silica nanoparticles induce apoptosis and growth arrest in human prostate cancer cells.

Prostate cancer (PCa) is the most commonly diagnosed cancer in men. The progression and invasion of PCa are normally mediated by the overexpression of chemokine receptors (CKRs) and the interaction between CKRs and their cognate ligands. We recently demonstrated that venom extracted from Walterinnesia aegyptia (WEV) either alone or in combination with silica nanoparticles (WEV+NP) mediated the growth arrest and apoptosis of breast cancer cells. In the present study, we evaluated the impact of WEV alone and WEV+NP on the migration, invasion, proliferation and apoptosis of prostate cancer cells. We found that WEV alone and WEV+NP decreased the viability of all cell types tested (PCa cells isolated from patient samples, PC3 cells and LNCaP cells) using an MTT assay. The IC(50) values were determined to be 10 and 5 µg/mL for WEV alone and WEV+NP, respectively. WEV+NP decreased the surface expression of the CKRs CXCR3, CXCR4, CXCR5 and CXCR6 to a greater extent than WEV alone and subsequently reduced migration and the invasion response of the cells to the cognate ligands of the CKRs (CXCL10, CXCL12, CXCL13 and CXCL16, respectively). Using a CFSE proliferation assay, we found that WEV+NP strongly inhibited epidermal growth factor-mediated PCa cell proliferation. Furthermore, analysis of the cell cycle indicated that WEV+NP strongly altered the cell cycle of PCa cells and enhanced the induction of apoptosis. Finally, we demonstrated that WEV+NP robustly decreased the expression of anti-apoptotic effectors, such as B cell Lymphoma-2 (Bcl-2), B cell Lymphoma-extra large (Bcl-(XL)) and myeloid cell leukemia sequence-1 (Mcl-1), and increased the expression of pro-apoptotic effectors, such as Bcl-2 homologous antagonist/killer (Bak), Bcl-2-associated X protein (Bax) and Bcl-2-interacting mediator of cell death (Bim). WEV+NP also altered the membrane potential of mitochondria in the PCa cells. Our data reveal the potential of nanoparticle-sustained delivery of snake venom as effective treatments for prostate cancer.

Enhanced anticancer efficacy of snake venom combined with silica nanoparticles in a murine model of human multiple myeloma: molecular targets for cell cycle arrest and apoptosis induction.

Multiple myeloma (MM) is a clonal disease of plasma cells that reside in the bone marrow (BM). MM is an incurable disease; thus, screening for novel anti-myeloma drugs remains critically important. We recently described a silica nanoparticle-based snake venom delivery model that targets cancer cells, but not normal cells. Using this model, we demonstrated a strong enhancement of the antitumor activity of snake venom extracted from Walterinnesia aegyptia (WEV) in two breast carcinoma cell lines when the venom was combined with silica nanoparticles (WEV+NP). In the present study, we aimed to delineate the in vivo therapeutic efficacy of WEV+NP in an MM-bearing experimental nude mouse model. We found that treatment with WEV+NP or WEV alone significantly inhibited tumor growth compared to treatment with NP or vehicle. WEV+NP- and WEV-treated cancer cells exhibited marked elevations in oxidative stress and robust reductions in the levels of interleukin-6 (IL-6) and B cell-activating factor (BAFF). WEV+NP also decreased the surface expression of the chemokine receptors CXCR3, CXCR4 and CXCR6 to a greater extent than WEV alone, and WEV+NP subsequently reduced migration in response to the cognate ligands CXCL10, CXCL12 and CXCL16. Furthermore, we found that WEV+NP strongly inhibited insulin-like growth factor 1 (EGF-1)- and IL-6-mediated MM cell proliferation, altered the cell cycle and enhanced the induction of apoptosis of MM cells. In addition, the results of treatment with WEV+NP or WEV alone revealed that the combination of WEV with NP robustly decreased the expression of cyclin D1, Bcl-2 and the phosphorylation of AKT; increased the expression of cyclin B1; altered the mitochondrial membrane potential; increased the activity of caspase-3, -8 and -9; and sensitized MM cells to growth arrest and apoptosis. Our data reveal the therapeutic potential of the nanoparticle-sustained delivery of snake venom to fight cancer cells.