Kaempferol Induces Cell Death in A2780 Ovarian Cancer Cells and Increases Their Sensitivity to Cisplatin by Activation of Cytotoxic Endoplasmic Reticulum-Mediated Autophagy and Inhibition of Protein Kinase B.

This study investigated whether kaempferol could inhibit ovarian cancer (OC) by activation of endoplasmic reticulum (ER) stress and autophagy, and tested its effect on the sensitivity of OC cells to cisplatin (cis-diamminedichloroplatinum, DPP). To study the effect of kaempferol on activation of ER stress and autophagy and find out whether its mechanism of action involves calcium (Ca2+), A2780 OC cells were cultured in DMEM/F12 for 24 h with or without kaempferol (40 µmol/l) in the presence or absence of autophagy or ER stress inhibitors or a calcium chelator. To study the effect of kaempferol on the sensitivity of OC cells to DPP and the potential involvement of modulation of protein kinase B (Akt) expression, A2780 OC were incubated with kaempferol and increasing concentrations of DPP (0-20 µmol/l) and then with kaempferol at its predetermined IC50 (6.8 µmol/l). Compared to control cells, kaempferol increased cell apoptosis (158 %) and decreased viability (53.17 %) and proliferation (49.17 %) of A2780 OC cells. Concomitantly, it increased the protein levels of GRP78, PERK, ATF6, IRE-1, LC3II, beclin 1, and caspase 4, thus suggesting activation of cytotoxic autophagy. This was mediated by increasing intracellular Ca+2 levels. In addition, kaempferol increased the sensitivity of A2780 cells to DPP (IC50 from 6.867 ± 0.99 to 3.73 ± 0.59 µmol/l) by decreasing the protein levels of p-Akt (0.31 ± 0.09 vs 0.12 ± 0.005). In conclusion, the findings of this study encourage the use of kaempferol alone or in combination with DPP to inhibit tumorigenesis of ovarian cells.

Immunomodulatory and anti-cancer potential of cloves (Syzygium aromaticum) bud extract and its phytogenic silver nanoparticles.

Clove plant (Syzygium aromaticum) is one of the Myrtaceae family. It's a common flavor in food and the traditional medicine. The study's objective was to ascertain whether the clove bud aqueous extract (CAE) and CAE + nanosilver have any biological effects on immune cells and HT-29 colon cancer cell line. Nanosilver was produced through green synthesis approach using CAE. Produced nanosilver was characterized via electron microscope (scanning, SEM) and ultraviolet-visible spectroscopy. CAE and CAE + nanosilver were examined for their active biomolecules using FTIR analysis, p53 contents using real-time PCR, apoptosis and cell cycle arrest power on HT-29 cancer cell line via flow cytometerty and immunomodulatory potential utilizing MTT assay. Results cleared that a spherical nanosilver with a diameter range of 53 nm was formed by CAE. There were several active biomolecules in CAE and CAE + nanosilver. CAE and CAE + nanosilver increased the p53 protein expression and apoptotic cell number in HT-29 colon cancer cells. CAE and CAE + nanosilver could arrest HT-29 cells at the phase G2/M. CAE and CAE + nanosilver stimulated quiescent and PHA-pre-treated splenic cells at higher concentrations, and CAE suppressed quiescent splenic cell when diluted. In conclusion, the safe edible Syzygium aromaticum plant can be utilized to make anti-tumor agent, essentially for colon tumor. As Syzygium aromaticum plant could stimulate immune cells, it can be used as immune-stimulatory agent that can help fight tumor and tumor development.

The protective effect of melatonin supplementation against taxol-induced testicular cytotoxicity in adult rats.

The aim of the present investigation was to study the toxic influences of taxol (TXL) on the testes of rats and the protective impact of melatonin (MLT) against such effects. Rats were classified into control, sham, TXL, MLT, and MLT+TXL-treated groups. Histological and ultrastructural changes were observed in testicular tissues of TXL-intoxicated rats including thickening of tunica albuginea and degenerative alterations in spermatogenic, Sertoli, and Leydig cells. A significant increase (P≤0.05) was found in the thickness of tunica albuginea and numbers of tubules without sperm, apoptotic germinal epithelia, and apoptotic Leydig cells, whereas the diameter of tubules and height of germinal epithelia displayed a significant decrease (P≤0.05) compared with the control, sham, and MLT-treated groups. Immunohistochemically, a marked decrease (P≤0.05) in Bcl-2 immunoreactivity and significant elevation (P≤0.05) in P53 and caspase-3 immunoreactivities were recorded. Co-treatment of MLT and TXL modulated such histological, histomorphometrical, and ultrastructural changes induced by TXL. Also, MLT had a protective effect against testicular apoptosis induced by TXL, as shown by the elevated expression of Bcl-2 and decreased expression of P53 and caspase-3. In conclusion, the current investigation proved that MLT had a protective role against TXL-induced testicular cytotoxicity, which may be a result of inhibition of testicular apoptosis.

Melatonin attenuates thioacetamide-induced liver fibrosis in male rats through modulation of interleukin-6, interleukin-4, apoptosis and urokinase plasminogen activator receptor-associated protein/Endo180.

Liver fibrosis is a chronic progressive disease, its resolution still unclear, and the current study explored the role of melatonin in modulation of interleukin-6 (IL-6), interleukin-4 (IL-4), transforming growth factor beta1 (TGF-ß1) and urokinase plasminogen activator receptor-associated protein/Endo180 (uPARAP/Endo180) pathway in thioacetamide (TAA)-induced hepatotoxicity. Thirty two adult Sprague-Dawley rats were divided into four groups: vehicle control group, TAA-induced liver fibrosis group that was left untreated, melatonin administration before and along with TAA and melatonin along with TAA group. TTA-induced massive liver necrosis, fibrosis around portal tract and increases serum levels of liver enzymes and total bilirubin when compared with control vehicle group. While both melatonin pretreatment and treatment retained liver parenchyma and liver enzymes quite similar to control group and reduced TAA-induced liver injury. Notably, melatonin pretreatment and treatment increased collagen degradation in TAA liver injury by19, 31.7-fold respectively evidence by collagen percentage area. Melatonin also decreased the amount of thiobarbituric acid reactive compounds and retained the reduced glutathione and superoxide dismutase to basal level quite similar to control group. Additionally, melatonin significantly (P value ≤0.05) decreased the levels of TGF-ß1, epidermal growth factor (EGF), hydroxyproline, tissues IL-6, caspase-3, and receptor interacting serine/threonine kinase1 (RIPK1), fibrillin-1, and - smooth muscle actin in the liver tissues while significantly (P value ≤0.05) increasing the levels of IL-4 and uPARAP/Endo180. Due to its anti-inflammatory, anti-apoptotic, and antioxidant capabilities as well as its ability to decrease hepatic stellate cell activation and fibrogenesis, these data imply that melatonin has a powerful anti-fibrotic effect.

Kaempferol protects against streptozotocin-induced diabetic cardiomyopathy in rats by a hypoglycemic effect and upregulating SIRT1.

This study investigated if kaempferol could attenuate the oxidative, inflammatory, and fibrotic damage of the left ventricles (LVs) in streptozotocin (STZ)-diabetic rats by modulating silent mating type information regulation 2 homolog 1 (SIRT1) signaling. Adult male rats were divided into 5 groups (n = 12/each) as control, control + kaempferol, STZ-induced diabetes mellitus (STZ-DM), STZ-DM + kaempferol, and STZ-DM + kaempferol + EX-527, a sirtuin 1 (SIRT1) inhibitor. Administration of kaempferol to diabetic rats significantly preserved the systolic and diastolic functions of the LVs that was associated with a significant reduction in ventricular collagen deposition, infiltration of inflammatory cells, and protein expression of Bcl2-associated X protein (Bax), cleaved caspase-3, and cytochrome-C. In both the control and diabetic rats, kaempferol attenuated the loss in body weights, reduced fasting glucose levels, and increased fasting insulin levels and HOMA-ß. Besides, kaempferol lowered the levels of reactive oxygen species (ROS), malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6), downregulated the transforming growth factor-ß1 (TGF-ß1) and reduced the nuclear levels of NF-κB p65. In concomitant, kaempferol increased the LV levels of manganese superoxide dismutase (MnSOD) and glutathione (GSH) and stimulated the total protein levels of Bcl2, the nuclear activity of SIRT1, and nuclear levels of nuclear factor erythroid 2-related factor 2 (Nrf2). These events were associated with increased deacetylase activity and total levels of SIRT1 and a parallel decrease in the acetylation of Nrf2, NF-κB, smad2, and FOXO1. In conclusion: kaempferol attenuate diabetic cardiomyopathy in STZ-treated rats through its hypoglycaemic and insulin-releasing effects, as well as a cardiac independent mechanism that involves activation of SIRT1.

Dehydroepiandrosterone protects against acetaminophen-induced liver damage in rats by upregulation of Bcl-2 and activation of sirt signalling.

The study examined the protective effect of exogenous administration of dehydroepiandrosterone (DHEA) against acetaminophen (APAP) -induced liver damage in rats and tested the underlying mechanism(s). Male rats were divided into 5 groups as control, control + DHEA, APAP, APAP + DHEA, and APAP + DHEA + EX-527 (SIRT1 inhibitor). Treatments were conducted for 10 days and then followed by intragastric administration of a single dose of APAP. DHEA not only reduced serum alanine transaminase (ALT) and aspartate aminotransferase (AST) but also preserved the liver structures. Besides, DHEA reduced hepatic levels of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), Bax, cleaved caspase-3. In the livers of both the control and APAP-treated rats, DHEA suppressed the generation of reactive oxygen species (ROS) and malondialdehyde (MDA), increased levels of glutathione (GSH), MnSOD (SOD2), and Bcl-2 levels, lowered Bax/Bcl-2 ratio, enhanced the activity of nuclear factor erythroid-derived 2-like 2 (Nrf2), and inhibited nuclear factor kappaB (NF-κB) p65. All these effects coincided with a significant increase in the levels and activity of SIRT1 and a reduction in the acetylation of Nrf2, p53, forkhead box class O transcription factor 1 (FOXO1), and NF-κB p65. Except for Bcl-2, treating the rats with EX-527 abolished the beneficial effects of DHEA on all these markers. In conclusion, DHEA prevents APAP-induced liver damage by concomitant upregulation of Bcl-2 and SIRT1-dependent effect.

Heat shock protein expression independently predicts survival outcome in schistosomiasis-associated urinary bladder cancer.

BACKGROUND: Heat shock proteins (HSPs) are synthesized by cells in response to various stress conditions, including carcinogenesis. These molecules have been studied in several malignancies, among which bladder carcinoma. This is the first study attempting to clarify the significance of HSP27 and HSP70 in schistosomiasis-associated bladder carcinoma and their relation to prognosis. METHODS: HSP27 and HSP70 were localized immunohistochemically in tissue sections from 75 chistosomiasis-associated bladder carcinomas. Their expression was correlated with clinical and pathological features and their impact on 5-year disease-free survival was studied with univariate and multivariate analysis. RESULTS: In all, 45 and 51 patients were positive for HSP27 and HSP70 expression, respectively. A significant correlation was found between expression of both HSPs and tumor grade, stage, DNA ploidy and recurrence. In univariate analysis, a statistically significant association of HSP27 and HSP70 expression with 5-year disease-free survival was found. In a multivariate Cox proportional hazards model, both HSP27 and HSP70 maintained a statistically significant impact on survival. CONCLUSIONS: The current results indicate that expression of HSP27 and HSP70 may have prognostic relevance in patients with schistosomiasis-associated bladder cancer. HSPs may be useful markers for patients with this type of bladder carcinoma and may be used for predicting disease progression.

The protective effect of melatonin supplementation against taxol-induced testicular cytotoxicity in adult rats

The aim of the present investigation was to study the toxic influences of taxol (TXL) on the testes of rats and the protective impact of melatonin (MLT) against such effects. Rats were classified into control, sham, TXL, MLT, and MLT+TXL-treated groups. Histological and ultrastructural changes were observed in testicular tissues of TXL-intoxicated rats including thickening of tunica albuginea and degenerative alterations in spermatogenic, Sertoli, and Leydig cells. A significant increase (P≤0.05) was found in the thickness of tunica albuginea and numbers of tubules without sperm, apoptotic germinal epithelia, and apoptotic Leydig cells, whereas the diameter of tubules and height of germinal epithelia displayed a significant decrease (P≤0.05) compared with the control, sham, and MLT-treated groups. Immunohistochemically, a marked decrease (P≤0.05) in Bcl-2 immunoreactivity and significant elevation (P≤0.05) in P53 and caspase-3 immunoreactivities were recorded. Co-treatment of MLT and TXL modulated such histological, histomorphometrical, and ultrastructural changes induced by TXL. Also, MLT had a protective effect against testicular apoptosis induced by TXL, as shown by the elevated expression of Bcl-2 and decreased expression of P53 and caspase-3. In conclusion, the current investigation proved that MLT had a protective role against TXL-induced testicular cytotoxicity, which may be a result of inhibition of testicular apoptosis.

Prognostic value of carcinoma-associated antigen MK-1 in urinary bladder carcinoma.

BACKGROUND: The present study has been performed to evaluate the expression of MK-1 in schistosomiasis-associated squamous cell carcinoma of the urinary bladder and to correlate this new marker with the conventional histopathological parameters. PATIENTS AND METHODS: Paraffin sections of 5-microm thickness from 81 cases were prepared for hematoxylin and eosin staining and immunohistochemical analysis of MK-1 expression was carried out. RESULTS: Forty-six cases (56.8%) were positive for MK-1 protein expression. Significant correlations between MK-1 expression and tumor grade (p=0.004), schistosoma (p=0.031), DNA ploidy (p=0.001), and tumor recurrence (p<0.001) were observed. MK-1, sex, tumor grade, stage, schistosoma, DNA ploidy, and recurrence were evaluated in relation to outcome. Univariate and multivariate analysis of survival were performed. The overall 5-year survival was 51.85%. In univariate analysis, MK-1 expression, tumor grade, DNA ploidy, and recurrence had a significant impact on the survival of these patients. In a Cox proportional hazards model, recurrence maintained its significant impact on survival. CONCLUSIONS: These findings suggest that MK-1 is a prognostic marker for recurrence: 34 (87.2%) of 39 recurrent cases were positive for MK-1 expression. However, only recurrence was an independent prognostic factor in patients with schistosomiasis- associated squamous cell carcinoma of the bladder.

Prognostic value of p53 and MDM2 expression in bilharziasis-associated squamous cell carcinoma of the urinary bladder.

The aim of this study was to elucidate the associations between immunostaining for MDM2 and p53, their respective expression in squamous cell carcinoma of the urinary bladder, and the value of these variables for predicting treatment outcome after cystectomy. Inactivation of TP53 might play a role in the development and progression of bladder cancer. Complex formation with the MDM2 product is one mechanism that inactivates the p53 protein. Therefore, the MDM2 and the p53 protein were investigated to study potential interactions in bladder cancer. Fifty archival bladder tissue specimens were immunohistochemically stained using monoclonal antibodies against p53 and MDM2. Staining for p53 was observed in 48% of the specimens and staining for MDM2 in 20%. Univariate analysis demonstrated a significant correlation between p53 accumulation and survival (p = 0.0101), while the correlation between MDM2 and survival was not significant (p = 0.7183). The combined expression of MDM2 and p53 doest not add to the prognostic information provided by p53 alone.