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Topotecan, a topoisomerase I inhibitor, is an anticancer drug widely used in the therapy of lung, ovarian, colorectal, and breast adenocarcinoma. Due to the primary dose-limiting toxicity of topotecan, which is myelosuppressive, it is necessary to identify other chemotherapeutic agents that can work synergistically with topotecan to increase its efficacy and limit its toxicity. Many studies have shown synergism upon the combination of topotecan with other chemotherapeutic agents such as gemcitabine. Other studies have demonstrated that pre-exposing cells to naturally occurring compounds such as thymoquinone, followed by gemcitabine or oxaliplatin, resulted in higher growth inhibition compared to treatment with gemcitabine or oxaliplatin alone. Our aim was to elucidate the underlying mechanism of action of topotecan in the survival and apoptotic pathways in human colon cancer cell lines in comparison to thymoquinone, to study the proapoptotic and antiproliferative effects of thymoquinone on the effectiveness of the chemotherapeutic agent topotecan, and to investigate the potential synergistic effect of thymoquinone with topotecan. Cells were incubated with different topotecan and thymoquinone concentrations for 24 and 48 hours in order to determine the IC50 for each drug. Combined therapy was then tested with ± 2 values for the IC50 of each drug. The reduction in proliferation was significantly dose- and time-dependent. After determining the best combination (40 µM thymoquinone and 0.6 µM topotecan), cell proteins were extracted after treatment, and the expression levels of B-cell lymphoma 2 and of its associated X protein, proteins p53 and p21, and caspase-9, caspase-3, and caspase-8 were studied by Western blot. In addition, cell cycle analysis and annexin/propidium iodide staining were performed. Both drugs induced apoptosis through a p53-independent mechanism, whereas the expression of p21 was only seen in thymoquinone treatment. Cell cycle arrest in the S phase was detected with each compound separately, while combined treatment only increased the production of fragmented DNA. Both compounds induced apoptosis through the extrinsic pathway after 24 hours; however, after 48 hours, the intrinsic pathway was activated by topotecan treatment only. In conclusion, thymoquinone increased the effectiveness of the chemotherapeutic reagent topotecan by inhibiting proliferation and lowering toxicity through p53- and Bax/Bcl2-independent mechanisms., (Georg Thieme Verlag KG Stuttgart · New York.)

In this review, we summarized published reports that investigated the role of Nigella sativa (NS) and its active constituent, thymoquinone (TQ) in oral health and disease management. The literature studies were preliminary and scanty, but the results revealed that black seed plants have a potential therapeutic effect for oral and dental diseases. Such results are encouraging for the incorporation of these plants in dental therapeutics and hygiene products. However, further detailed preclinical and clinical studies at the cellular and molecular levels are required to investigate the mechanisms of action of NS and its constituents, particularly TQ.

Background: Traumatic brain injury is a leading cause of morbidity and mortality worldwide. We evaluated the neuroprotective effects of thymoquinone (TQ) in a rat model of traumatic brain injury by using biochemical and histopathologic methods for the first time., Materials and Methods: Twenty-four rats were divided into sham (n = 8), trauma (n = 8), and TQ-treated (n = 8) groups. A moderate degree of head trauma was induced with the use of Feeney's falling weight technique, and TQ (5 mg/kg/day) was administered to the TQ-treated group for 7 days. All animals were killed after cardiac perfusion. Brain tissues were extracted immediately after perfusion without damaging the tissues. Biochemical procedures were performed with the serum, and a histopathologic evaluation was performed on the brain tissues. Biochemical experiments included malondialdehyde (MDA), reduced and oxidized coenzyme Q10 analysis, DNA isolation and hydroylazation, and glutathione peroxidase, and superoxide dismutase analyses., Results: Neuron density in contralateral hippocampal regions (CA1, CA2-3, and CA4) 7 days after the trauma decreased significantly in the trauma and TQ-treated groups, compared with that in the control group. Neuron densities in contralateral hippocampal regions (CA1, CA2-3, and CA4) were greater in the TQ-treated group than in the trauma group. TQ did not increase superoxide dismutase or glutathione peroxidase antioxidant levels. However, TQ decreased the MDA levels., Conclusions: These results indicate that TQ has a healing effect on neural cells after head injury and this effect is mediated by decreasing MDA levels in the nuclei and mitochondrial membrane of neurons., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Embelia ribes Burm of Myrsinaceae family has been widely used as an herb in the traditional medicine of India. Embelin is an active component extracted from the fruits of Embelia ribes. It has a wide spectrum of biological activities and is not toxic at low dose. This review focuses on the physical-chemical properties and bioactivities of Embelin, as well as its effects on chronic diseases such as tumors, autoimmune inflammatory diseases, parasitic infections, microbial infections, diabetes, obesity, and cardio-cerebral vascular diseases. The underlying mechanisms of the effects are also discussed. As a multiple-targeted therapeutic agent, Embelin has the potential to be used widely for the treatment of a variety of chronic diseases, including malignant tumors.

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., (© 2016 The Author(s) Published by S. Karger AG, Basel.)

Context: Thymoquinone (TQ), an active component of Nigella sativa L. (Ranunculaceae), possesses anti-inflammatory and anti-oxidative properties. Polycystic ovary syndrome exhibits chronic inflammatory behavior, thus might involve nuclear factor kappa B (NF-κB) signaling and related molecular factors., Objective: The objective of the present study is to investigate and validate the effect of TQ in polycystic ovary (PCO) rat., Materials and Methods: To validate the effect of TQ (1 µM/ml), NF-κB activation, COX2 (cyclooxygenase-2) expression and reactive oxygen species (ROS) induction were studied in the KK1 cell line. To evaluate the effect of TQ (2 mg/200 µl olive oil/rat; sc) with an in vivo system, ovulation rate, levels of key ovulation mediators, and ovarian gelatinases activity were compared in superovulated, PCO, and RU486 + TQ-treated Wistar rats., Results: In vitro studies showed that NF-κB nuclear translocation, COX2, and ROS expression were repressed via TQ supplementation in RU486-treated KK1 cells. Pretreatment of TQ in the PCO rat model induced significant restoration of normal physio-molecular behavior of ovary, such as reduced cysts formation, increased ovulation rate, and normalization of key ovarian factors [like TNF-α-stimulated gene/protein 6, hyaluronan, hyaluronan-binding protein 1, COX2, matrix metalloproteinases (membrane type 1-matrix metalloproteinase, MMP9 and MMP2)], tissue inhibitor of metalloproteinases (TIMP-1 and TIMP-2), and gelatinases (like MMP9 and -2) activity during follicular maturation., Discussion and Conclusion: Overall, most of the above molecular changes are regulated via NF-κB pathway, thus TQ, due to its modulatory effect on the NF-κB signaling, could elevate normal ovarian phenotype and physiological function in the PCO model, indicating its remarkable potential as a remedy for rat PCO.

We aimed to investigate the protective role of thymoquinone (TQ) by targeting its antiapoptotic and antioxidant properties against kidney damage induced by arsenic in rats. We have used the 24 male Sprague-Dawley rats. Rats were divided into three groups. Physiological serum in 10 mL/kg dose as intragastric was given to the control group. Sodium arsenite (10 mg/kg, intragastric by gavage for fifteen days) was given to the arsenic group. Sodium arsenite (10 mg/kg, intragastric by gavage for fifteen days) and TQ (10 mg/kg, intragastric by gavage for 15 days) was given to the arsenic + TQ group. After 15 days, the animals' kidneys were taken theirs, then we have performed histological and apoptotic assessment. Superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) enzyme activities and malondialdehyde (MDA) levels have examined as the oxidative stress parameters. We have determined the levels of arsenic. Increased renal injury and apoptotic cells have been detected in the arsenic group. Degenerative changes in the arsenic + TQ group were diminished. Although the MDA levels were augmented in the arsenic group, SOD, CAT and GSH-Px enzyme activities were lessened than the other groups. Our findings suggest that TQ may impede the oxidative stress, the cells have been damaged and also the generation of apoptotic cells arisen from arsenic. TQ plays a protective role against arsenic-induced toxicity in kidney and may potentially be used as a remedial agent.

Objective: Thymoquinone (TQ) is an antioxidant and anti-apoptotic substance found in the Nigella sativa plant. Alpha-tocopherol (α-TP) is a potent antioxidant. We aimed to determine whether or not TQ and TP have a protective effect against lower limb ischemia-reperfusion (IR) injury of muscle and the sciatic nerve., Materials and Methods: A single dose of TQ 25 mg/kg was given intraperitoneally to the TQ group, a single dose of α-TP 200 mg/kg was given intraperitoneally to the α-TP group. IR was performed for 45 minutes after the drugs' applications., Results: While serum levels of malondialdehyde (MDA) and interleukin-6 (IL-6) of the IR group were significantly higher than those of the TQ plus α-TP, TQ and α-TP groups (p<0.001, p<0.001, p=0.008, respectively) and IL-6 (all p<0.001), the reduced glutathione (GSH) level of the IR group was lower than that of the other three groups. While neuronal nitric oxide synthase activity of nerve tissues of the IR group was significantly lower than that of the TQ plus α-TP group, the muscle tissue caspase-3 activity was higher than that of the TQ plus α-TP group., Conclusions: Administration of TQ plus α-TP may strongly protect muscle and nerve tissues against IR injury via their synergistic effects.

Aims: The present investigation was designed to evaluate the effect of thymoquinone in a septic animal model and to explore the role of nitric oxide (NO) in the process., Main Methods: To achieve this, mice (n=12 per group) were treated in parallel with thymoquinone (0.75mg/kg/day) and/or NG-nitro-l-arginine methyl ester (L-NAME; 400μg/g/day) prior to sepsis induction with live Escherichia coli., Key Findings: Thymoquinone significantly improved renal and hepatic functions alone and in combination with L-NAME. This was associated with less NO production and lower oxidative stress in treated animals. Tumor necrosis factor-α concentration with thymoquinone and L-NAME were 36.27±3.41pg/ml and 56.55±5.85pg/ml, respectively, as opposed to 141.11±6.46pg/ml in septic controls. Similarly, Interleukin-1α, 2, 6 and 10 levels decreased significantly upon treatment with thymoquinone and L-NAME as compared with untreated septic animals. NF-κB and NF-κB-DNA binding activity in nuclear proteins were also significantly down-regulated. Vascular responsiveness studies in isolated mouse aortae demonstrated a reduced relaxation to acetylcholine exposure in septic mice treated with thymoquinone., Significance: These findings suggest that thymoquinone prevents sequels of the multiple organ failure syndrome of sepsis by modulating the production of NO and its inflammatory sequela, and adjusting vascular responsiveness., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Objective: Lynch syndrome is caused by germline mutations in DNA mismatch repair genes leading to microsatellite instability (MSI) and colorectal cancer. Mesalazine, commonly used for the treatment of UC, reduces MSI in vitro. Here, we tested natural compounds for such activity and applied mesalazine and thymoquinone in a Msh2(loxP/loxP) Villin-Cre mouse model for Lynch syndrome., Design: Flow cytometry was used for quantitation of mutation rates at a CA13 microsatellite in human colon cancer (HCT116) cells that had been stably transfected with pIREShyg2-enhanced green fluorescent protein/CA13, a reporter for frameshift mutations. Mice were treated for 43 weeks with mesalazine, thymoquinone or control chow. Intestines were analysed for tumour incidence, tumour multiplicity and size. MSI testing was performed from microdissected normal intestinal or tumour tissue, compared with mouse tails and quantified by the number of mutations per marker (NMPM)., Results: Besides mesalazine, thymoquinone significantly improved replication fidelity at 1.25 and 2.5 µM in HCT116 cells. In Msh2(loxP/loxP) Villin-Cre mice, tumour incidence was reduced by mesalazine from 94% to 69% (p=0.04) and to 56% (p=0.003) by thymoquinone. The mean number of tumours was reduced from 3.1 to 1.4 by mesalazine (p=0.004) and to 1.1 by thymoquinone (p<0.001). Interestingly, MSI was reduced in normal intestinal tissue from 1.5 to 1.2 NMPM (p=0.006) and to 1.1 NMPM (p=0.01) by mesalazine and thymoquinone, respectively. Thymoquinone, but not mesalazine, reduced MSI in tumours., Conclusions: Mesalazine and thymoquinone reduce tumour incidence and multiplicity in Msh2(loxP/loxP) Villin-Cre mice by reduction of MSI independent of a functional mismatch repair system. Both substances are candidate compounds for chemoprevention in Lynch syndrome mutation carriers., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

Background: Ovarian cancer is the most lethal gynecologic malignancy, with limited treatment options for chemoresistant disease. An important link between inflammation and peritoneal spread of ovarian cancer is NF-κB signaling. Thymoquinone (TQ) exerts multiple anti-tumorigenic cellular effects, including NF-κB inhibition. We aimed to investigate the therapeutic potential of TQ in an established murine syngeneic model of ovarian cancer., Methods: ID8-NGL mouse ovarian cancer cells stably expressing an NF-κB reporter transgene were injected intra-peritoneally into C57BL/6 mice, and mice were treated with TQ or vehicle for 10 or 30 days. TQ was combined with the macrophage depleting drug, liposomal clodronate, in selected experiments. Effects on peritoneal tumor burden were measured by volume of ascites, number of peritoneal implants and mesenteric tumor mass. NF-κB reporter activity and markers of proliferation and apoptosis were measured in tumors and in confirmatory in vitro experiments. Protein or mRNA expression of M1 (anti-tumor) and M2 (pro-tumor) macrophage markers, and soluble cytokine profiles, were examined from harvested ascites fluid, peritoneal lavages and/or tumor sections. 2-tailed Mann-Whitney tests were used for measuring differences between groups in in vivo experiments., Results: Consistent with its effects in vitro, TQ reduced proliferation and increased apoptosis in ID8-NGL tumors after 10 and 30 day treatment. Prolonged TQ treatment did not significantly alter tumor number or mass compared to vehicle, but rather exerted an overall deleterious effect by stimulating ascites formation. Increased ascites was accompanied by elevated NF-κB activity in tumors and macrophages, increased pro-tumor M2 macrophages and expression of pro-tumorigenic soluble factors such as VEGF in ascites fluid, and increased tumor infiltration of M2 macrophages. In contrast, a 10 day exposure to TQ produced no ascites, and reduced tumor NF-κB activity, M2 macrophages and soluble VEGF levels. Peritoneal macrophage depletion by clodronate significantly reduced tumor burden. However, TQ-stimulated ascites was further enhanced by co-treatment with clodronate, with macrophages present overwhelmingly of the M2 phenotype., Conclusions: Our findings show that pro-tumorigenic microenvironmental effects limited the efficacy of TQ in a syngeneic mouse model of ovarian cancer, and provide caution regarding its potential use in clinical trials in ovarian cancer patients.

Neurodegenerative disorders are generally characterized by abnormal aggregation and deposition of specific proteins. Amyloid beta (Aβ)-associated neurodegenerative disorder is characterized by an oxidative damage that, in turn, leads to some behavioral changes before the establishment of dementia such as depression and anxiety. In the current study, we investigated the effect of heat shock protein 90 inhibitor geldanamycin (GA) administration 24 h before Aβ injection. In our experiment, 7 days after Aβ injection, elevated plus maze and forced swimming test were conducted to assess anxiety and depression-like behaviors. Levels of autophagy markers and malondialdehyde (MDA) and also activity of catalase in the hippocampus of rats were evaluated. Our behavioral analyses demonstrated that GA pretreatment can significantly decrease anxiety- and depression-like behaviors in Aβ-injected rats. Also, levels of autophagy markers including Atg12, Atg7, and LC3-II increased, while MDA level decreased and the activity of catalase increased in rats pretreated with GA compared to Aβ-injected rats. Thus, we assumed that GA, at least in part, ameliorated Aβ-mediated anxiety and depression by inducing autophagy and improving antioxidant defense system.

The aim of this study was to investigate the protective effects of thymoquinone treatment on cholestatic rats with liver injury. Thirty-two Sprague-Dawley rats were divided randomly into four groups: normal control, bile duct ligation model control, low-dose thymoquinone (25 mg/kg), and high-dose thymoquinone (50 mg/kg). Thymoquinone gavage was administered continuously 3 days before bile duct ligation, and saline, at the same volume, was administered to the control group. The rats were sacrificed after 2 weeks of treatment, and the liver tissues were obtained and frozen. The contents of hydroxyproline (HP), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the homogenate of the liver tissues were determined to evaluate the changes in hepatic tissue pathology by fibrosis scoring. The HP and MDA levels were significantly lower and the SOD and GPx levels were significantly higher in the thymoquinone-treatment group than the corresponding levels in the model control group, and the differences were statistically significant (P < 0.05) and dose-dependent. The hepatic necrosis areas and hepatic fibrosis scores of the thymoquinone-treatment groups were significantly lower than those of the model group (P < 0.05). Thymoquinone increased the antioxidative capacity of liver and reduced the oxidative stress damage to the liver. Thymoquinone can be used as a liver protectant in patients with cholestasis.

Heat shock proteins (HSPs) are induced by cellular stress and function as molecular chaperones that regulate protein folding. Diabetes impairs the function/expression of many HSPs, including HSP70 and HSP90, key regulators of pathological mechanisms involved in diabetes complications. Therefore, we investigated whether pharmacological HSP90 inhibition ameliorates diabetes-associated renal damage and atheroprogression in a mouse model of combined hyperglycemia and hyperlipidemia (streptozotocin-induced diabetic apolipoprotein E-deficient mouse). Treatment of diabetic mice with 17-dimethylaminoethylamino-17-demethoxygeldanamycin (DMAG, 2 and 4 mg/kg, 10 weeks) improved renal function, as evidenced by dose-dependent decreases in albuminuria, renal lesions (mesangial expansion, leukocyte infiltration, and fibrosis), and expression of proinflammatory and profibrotic genes. Furthermore, DMAG significantly reduced atherosclerotic lesions and induced a more stable plaque phenotype, characterized by lower content of lipids, leukocytes, and inflammatory markers, and increased collagen and smooth muscle cell content. Mechanistically, the renoprotective and antiatherosclerotic effects of DMAG are mediated by the induction of protective HSP70 along with inactivation of nuclear factor-κB (NF-κB) and signal transducers and activators of transcription (STAT) and target gene expression, both in diabetic mice and in cultured cells under hyperglycemic and proinflammatory conditions. In conclusion, HSP90 inhibition by DMAG restrains the progression of renal and vascular damage in experimental diabetes, with potential implications for the prevention of diabetes complications., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Background: The 90-kDa heat-shock protein (heat-shock protein 90) is an abundant cytosolic chaperone, and inhibition of heat-shock protein 90 by 17-allylamino-17-demethoxygeldanamycin (17-AAG) compromises transforming growth factor (TGF)-β-mediated transcriptional responses by enhancing TGF-β receptor I and II degradation, thus preventing Smad2/3 activation. In this study, the authors evaluated whether heat-shock protein 90 regulates TGF-β signaling in the pathogenesis and treatment of keloids., Methods: Keloid fibroblasts were treated with 17-AAG (10 μM), and mRNA levels of collagen types I and III were determined by real-time reverse- transcriptase polymerase chain reaction. Also, secreted TGF-β1 was assessed by enzyme-linked immunosorbent assay. The effect of 17-AAG on protein levels of Smad2/3 complex was determined by Western blot analysis. In addition, in 17-AAG-treated keloid spheroids, the collagen deposition and expression of major extracellular matrix proteins were investigated by means of Masson trichrome staining and immunohistochemistry., Results: The authors found that heat-shock protein 90 is overexpressed in human keloid tissue compared with adjacent normal tissue, and 17-AAG decreased mRNA levels of type I collagen, secreted TGF-ß1, and Smad2/3 complex protein expression in keloid fibroblasts. Masson trichrome staining revealed that collagen deposition was decreased in 17-AAG-treated keloid spheroids, and immunohistochemical analysis showed that expression of collagen types I and III, elastin, and fibronectin was markedly decreased in 17-AAG-treated keloid spheroids., Conclusion: These results suggest that the antifibrotic action of heat-shock protein 90 inhibitors such as 17-AAG may have therapeutic effects on keloids.

Background: To determine the effect of thymoquinone on adhesion formation in a rat caecotomy/suture model., Material and Methods: Thirty wistar rats were randomized into three groups: The control group received saline and the thymoquinone group received 10 mg/kg thymoquinone after cecal caecotomy/suture model. In the sham group the abdominal wall was closed without any abrasion to the cecum. On day 15, adhesions were classified, and histopathological samples were taken., Results: There were no incisional hernias or wound dehiscences. In comparing adhesion scores, a significant difference was found between the thymoquinone and the control groups (p < 0.05). The grade of inflammation for the thymoquinone and the sham groups were significantly lower than that of the control group (p < 0.01 and p < 0.001, respectively). Hydroxyproline levels were significantly lower in the sham and thymoquinone groups compared to the control group (p < 0.05)., Conclusions: Based on the results of this study in a rat peritoneal adhesion model, intraperitoneal administered thymoquinone has a strong anti-adhesive effect., (© Acta Chirurgica Belgica.)

Objective: To determine the effect of thymoquinone and methotrexate on blood urea and serum creatinine in arthritic rats., Study Design: Experimental, comparative study., Place and Duration of Study: Postgraduate Medical Institute, Lahore, from March to August 2013., Methodology: Thirty two female Sprague-Dawley rats were randomized into four equal groups (n=8); group A(healthy control), group B (positive control), group C (Thymoquinone treated) and group D (Methotrexate treated). Arthritis developed within two weeks after a single pristane injection. Total leukocyte count, blood urea and serum creatinine were taken at day 0, 15 and 30. While clinical score of inflammation was taken at day 0 and then on every alternate day., Results: Development of arthritis and renal involvement was accompanied by significant raise in total leukocyte count, clinical score of inflammation, blood urea and serum creatinine as compared to healthy control rats (group A) till day 15 (p < 0.001). From day 15 to day 30 both thymoquinone (group C) and methotrexate (group D) significantly lowered the total leukocyte count, clinical score of inflammation and improved blood urea and serum creatinine as compared to arthritic rats (group B) (p < 0.001). Methotrexate was found a bit more effective than thymoquinone., Conclusion: Evaluation of results supported the beneficial effects of thymoquinone in renal injury produced by rheumatoid arthritis.

Prostate cancer is one of the most common cancer types worldwide. In 2014, there were an estimated 233,000 new cases and 29,480 mortalities in the United States. Androgen deprivation therapy, also called androgen suppression therapy, targets androgen signaling and remains the standard treatment for patients with advanced prostate cancer; however, responses to treatment are not durable and most patients advance to castrate-resistant prostate cancer. Therefore, novel therapeutic strategies to treat prostate cancer are urgently required. Heat shock protein 90 (Hsp90) is a chaperone protein that has been shown to regulate the progression of tumor cells. Numerous Hsp90 inhibitors show anti-tumor activity and several of them have entered clinical trials. Geldanamycin (GA) was identified as the first Hsp90 inhibitor, but shows hepatotoxicity at its effective concentrations, limiting its clinical use. In previous studies by our group, the GA derivative 17-ABAG was designed and synthesized. The present study showed that 17-ABAG inhibits the proliferation and induces apoptosis of LNCaP, an androgen-dependent prostate cancer cell line, in vitro through a classic apoptotic pathway. 17-ABAG also downregulated the Hsp90 client protein and inhibited androgen receptor nuclear localization in LNCaP cells. In addition, 17-ABAG suppressed the growth of LNCaP xenograft tumors without any obvious side-effects. The present study demonstrated that 17-ABAG is a promising anti-tumor agent and warrants further validation in prospective studies.

Objectives: Heat Shock Protein 90 (HSP90) is a molecular chaperone that stabilizes many oncogenic proteins. HSP90 inhibitors may sensitize tumors to cytotoxic agents by causing client protein degradation. Gemcitabine, which has modest activity in pancreas cancer, activates Chk1, a client protein of HSP90. This phase II trial was designed to determine whether 17AAG could enhance the clinical activity of gemcitabine through degradation of Chk1 in patients with stage IV pancreatic cancer., Methods: A multicenter, prospective study combining gemcitabine and 17AAG enrolled patients with stage IV pancreatic adenocarcinoma, adequate liver and kidney function, ECOG performance status 0-2, and no prior chemotherapy for metastatic disease. The primary goal was to achieve a 60 % overall survival at 6 months. Sixty-six patients were planned for accrual, with an interim analysis after 25 patients enrolled., Results: After a futility analysis to achieve the endpoint, accrual was halted with 21 patients enrolled. No complete or partial responses were seen. Forty percent of patients were alive at 6 months. Median overall survival was 5.4 months. Tolerability was moderate, with 65 % of patients having ≥ grade 3 adverse events (AE), and 15 % having grade 4 events., Conclusions: The lack of clinical activity suggests that targeting Chk1 by inhibiting HSP90 is not important in pancreatic cancer sensitivity to gemcitabine alone. Further studies of HSP90 targeted agents with gemcitabine alone are not warranted.

The chaperone heat shock protein 90 (Hsp90), a cell stress-inducible molecule that regulates activity of many client proteins responsible for cellular growth, differentiation and apoptosis, has been proposed as an important therapeutic target in patients with malignancies. More recently, its active participation in (auto)immune processes has been recognized as evidenced by amelioration of inflammatory disease pathways through pharmacological inhibition of Hsp90 in rodent models of autoimmune encephalomyelitis, rheumatoid arthritis and systemic lupus erythematosus. Based on own current research results, this viewpoint essay provides important insights that Hsp90 is also involved as a notable pathophysiological factor in autoimmune blistering dermatoses including epidermolysis bullosa acquisita, bullous pemphigoid and possibly dermatitis herpetiformis. The observed in vitro, ex vivo and in vivo efficacy of anti-Hsp90 treatment in experimental models of autoimmune bullous diseases and its underlying multimodal anti-inflammatory mechanisms of interference with key contributors to autoimmune-mediated blister formation supports the introduction of selective non-toxic Hsp90 inhibitors into the clinical setting for the treatment of patients with these disorders., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Current evidence shows that the majority of the damage induced during sepsis is pursuant to induction and overproduction of endogenous cytokines. Embelin has been reported to suppress cytokine expressions in inflammatory disorders. The present study was designed to investigate the effects of embelin on cecal and ligation and puncture (CLP)-induced rat sepsis. Single-dose administration of embelin 1 h after surgery significantly improved survival of rats with CLP-induced sepsis. In addition, embelin treatment reduced the serum levels of pro-inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 and decreased organ inflammation and injuries. Moreover, embelin suppressed the activation of p65 subunit of nuclear factor-kappa B (NF-κB) and signal transducers and activators of transcription 3 (STAT3). Collectively, these results indicated that embelin ameliorates sepsis in rats through suppressing STAT3 and NF-κB pathways.

Traumatic brain injury (TBI) is caused by both primary and secondary injury mechanisms, all of which cause neuronal cell death and functional deficits. Both apoptosis and autophagy participated in neuronal cell death and functional loss induced following TBI. Preclinical findings implicate that 17-allylamino-demethoxygeldanamycin (17-AAG), an anticancer drug in clinical, present neuroprotection actions in multiple neurological disorders, but whether 17-AAG is capable of modulating neuronal autophagy has never been addressed. The present study was designed to determine the hypothesis that17-AAG treatment could confer neuroprotection in a rat model of TBI. We also used an autophagy inhibitor 3-methyladenine (3-MA) as well as an autophagy inducer rapamycin (RAPA) to test its underlining mechanisms. Our results showed that post-TBI administration of 17-AAG could attenuate brain edema, decrease neuronal death, as well as improve the recovery of motor function. Afterwards, in our model, 17-AAG treatment protected against TBI-induced apoptosis activation as well as enhanced neuronal autophagy. The present study provides novel clues in understanding the mechanisms of which 17-AAG exerts its neuroprotective activity on neurological disorders., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Context: Papillary thyroid cancer (PTC) is the second most common cancer in females in Saudi Arabia. However, the pathogenesis of PTC is still not fully elucidated., Objective: To identify potential genes that play important role in progression of PTC, we studied the role of X-linked inhibitor of apoptosis protein (XIAP) as a potential prognostic marker and therapeutic target in a large cohort of PTC samples and cell lines., Design: A DNA microarray chip was used to screen for gene copy number. XIAP expression was assessed by immunohistochemistry in a tissue microarray format on a cohort of 1022 clinical samples. In vitro and in vivo studies were performed using Embelin and/or LY294002 on PTC cell lines., Results: XIAP was found to be amplified in 14 of 29 and overexpressed in 48.8% of PTC cases. XIAP overexpression was significantly associated with old age, extrathyroidal extension, tumor size, nodal involvement, tall-cell variant, advanced stage disease, and significantly poor disease-free survival (P = .0341). XIAP was also significantly associated with phosphorylated AKT (P < .0001), Bcl-Xl (P < .0001), and Ki67 (P = .0006) proteins. Embelin treatment caused growth inhibition and apoptosis in PTC cell lines and induced tumor regression in PTC xenograft in nude mice. Finally, the combination of suboptimal doses of Embelin and LY294002 induced a synergistic apoptotic response in PTC cells., Conclusion: XIAP dysregulation in PTC confers an aggressive phenotype with poor outcome. In vitro and in vivo studies using an XIAP inhibitor suggest that this subgroup of PTC with overexpression of XIAP can be therapeutically targeted, either alone or in combination, to induce efficient apoptosis in these cancers.

Context: Leishmaniasis is a complex disease with a broad spectrum of clinical presentations., Objective: We evaluated the anti-leishmanial effects of Nigella sativa L. (Ranunculaceae) against Leishmania tropica and Leishmania infantum with an in vitro model., Materials and Methods: Antileishmanial effects of essential oil and methanolic extract of N. sativa (0-200 µg/mL) and thymoquinone (0-25 µg/mL) on promastigotes of both species and their cytotoxicity activities against murine macrophages were evaluated using the MTT assay at 24, 48, and 72 h. Moreover, their leishmanicidal effects against amastigotes were investigated in a macrophage model, for 48 and 72 h., Results: The findings showed that essential oil (L. tropica IC50 9.3 μg/mL and L. infantum IC50 11.7 μg/mL) and methanolic extract (L. tropica IC50 14.8 μg/mL and L. infantum IC50 15.7 μg/mL) of N. sativa, particularly thymoquinone (L. tropica IC50 1.16 μg/mL and L. infantum IC50 1.47 μg/mL), had potent antileishmanial activity on promastigotes of both species after 72 h. In addition, essential oil (L. tropica IC50 21.4 μg/mL and L. infantum IC50 26.3 μg/mL), methanolic extract (L. tropica IC50 30.8 μg/mL and L. infantum IC50 34.6 μg/mL), and thymoquinone (L. tropica IC50 2.1 μg/mL and L. infantum IC50 2.6 μg/mL) mediated a significant decrease in the growth rate of amastigote forms of both species. Thymoquinone (CC50 38.8 μg/mL) exhibited higher cytotoxic effects against murine macrophages than the other extracts., Conclusion: N. sativa, especially its active principle, thymoquinone, showed a potent leishmanicidal activity against L. tropica and L.infantum with an in vitro model.

Ethnopharmacological Relevance: Embelia ribes Burm. f. (Fam. Myrsinaceae) locally known as Vidanga have been used for treating tumors, ascites, bronchitis, jaundice, diseases of the heart and brain in traditional Indian medicine. However, no scientific studies providing new insights in its pharmacological properties with respect to acute respiratory distress syndrome have been investigated., Aim: The present investigation aimed to elucidate the effectiveness of Embelin isolated from Embelia ribes seeds on attenuation of LPS-induced acute respiratory distress syndrome in murine models., Methods: Embelin (5, 10 and 20 mg/kg/day, i.p.) and Roflumilast (1 mg/kg/day, p.o.) were administered for four days and prior to LPS in rats (i.t.). Four hour after LPS challenge animals were anesthesized and bronchoalveolar lavage was done with ice-cold phosphate buffer. Assessment of BAL fluid was done for albumin, total protein, total cell and neutrophil count, TNF-α levels, nitrosoative stress. Superior lobe of right lung was used for histopathologic evaluation. Inferior lobe of right lung was used to obtain lung edema. Left lung was used for myeloperoxidase estimation. Arterial blood was collected immediately and analyzed for pH, pO2 and pCO2 were estimated., Results: Pretreatment with embelin (5, 10 and 20 mg/kg, i.p.) decreased lung edema, mononucleated cellular infiltration, nitrate/nitrite, total protein, albumin concentrations, TNF-α in the bronchoalveolar lavage fluid and myeloperoxidase activity in lung homogenate. Embelin markedly prevented pO2 down-regulation and pCO2 augmentation. Additionally, it attenuated lung histopathological changes in acute respiratory distress syndrome model., Conclusion: The study demonstrates the effectiveness of Embelia ribes Burm. f. (Fam. Myrsinaceae) seeds in acute respiratory distress syndrome possibly related to its anti-inflammatory and protective effect against LPS induced airway inflammation by reducing nitrosative stress, reducing physiological parameters of blood gas change, TNF-α and mononucleated cellular infiltration indicating it as a potential therapeutic agent for acute respiratory distress syndrome., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Unlabelled: Human noroviruses (HuNoV) are a significant cause of acute gastroenteritis in the developed world, and yet our understanding of the molecular pathways involved in norovirus replication and pathogenesis has been limited by the inability to efficiently culture these viruses in the laboratory. Using the murine norovirus (MNV) model, we have recently identified a network of host factors that interact with the 5' and 3' extremities of the norovirus RNA genome. In addition to a number of well-known cellular RNA binding proteins, the molecular chaperone Hsp90 was identified as a component of the ribonucleoprotein complex. Here, we show that the inhibition of Hsp90 activity negatively impacts norovirus replication in cell culture. Small-molecule-mediated inhibition of Hsp90 activity using 17-DMAG (17-dimethylaminoethylamino-17-demethoxygeldanamycin) revealed that Hsp90 plays a pleiotropic role in the norovirus life cycle but that the stability of the viral capsid protein is integrally linked to Hsp90 activity. Furthermore, we demonstrate that both the MNV-1 and the HuNoV capsid proteins require Hsp90 activity for their stability and that targeting Hsp90 in vivo can significantly reduce virus replication. In summary, we demonstrate that targeting cellular proteostasis can inhibit norovirus replication, identifying a potential novel therapeutic target for the treatment of norovirus infections., Importance: HuNoV are a major cause of acute gastroenteritis around the world. RNA viruses, including noroviruses, rely heavily on host cell proteins and pathways for all aspects of their life cycle. Here, we identify one such protein, the molecular chaperone Hsp90, as an important factor required during the norovirus life cycle. We demonstrate that both murine and human noroviruses require the activity of Hsp90 for the stability of their capsid proteins. Furthermore, we demonstrate that targeting Hsp90 activity in vivo using small molecule inhibitors also reduces infectious virus production. Given the considerable interest in the development of Hsp90 inhibitors for use in cancer therapeutics, we identify here a new target that could be explored for the development of antiviral strategies to control norovirus outbreaks and treat chronic norovirus infection in immunosuppressed patients., (Copyright © 2015, Vashist et al.)

Importance: The PIK3CA mutation is one of the most common mutations in head and neck squamous cell carcinoma (HNSCC). Through this research we attempt to elicit the role of oncogene dependence and effects of targeted therapy on this PIK3CA mutation., Objectives: (1) To determine the role of oncogene dependence on PIK3CA-one of the more common and targetable oncogenes in HNSCC, and (2) to evaluate the consequence of this oncogene on the effectiveness of newly developed targeted therapies., Design, Setting, and Participants: This was a cell culture-based, in vitro study performed at an academic research laboratory assessing the viability of PIK3CA-mutated head and neck cell lines when treated with targeted therapy., Exposures: PIK3CA-mutated head and neck cell lines were treated with 17-AAG, GDC-0941, trametinib, and BEZ-235., Main Outcomes and Measures: Assessment of cell viability of HNSCC cell lines characterized for PIK3CA mutations or SCC25 cells engineered to express the PIK3CA hotspot mutations E545K or H1047R., Results: Surprisingly, in engineered cell lines, the hotspot E545K and H1047R mutations conferred increased, rather than reduced, IC50 assay measurements when treated with the respective HSP90, PI3K, and MEK inhibitors, 17-AAG, GDC-0941, and trametinib, compared with the SCC25 control cell lines. When treated with BEZ-235, H1047R-expressing cell lines showed increased sensitivity to inhibition compared with control, whereas those expressing E545K showed slightly increased sensitivity of unclear significance., Conclusions and Relevance: (1) The PIK3CA mutations within our engineered cell model did not lead to enhanced oncogene-dependent cell death when treated with direct inhibition of the PI3K enzyme yet did show increased sensitivity compared with control with dual PI3K/mTOR inhibition. (2) Oncogene addiction to PIK3CA hotspot mutations, if it occurs, is likely to evolve in vivo in the context of additional molecular changes that remain to be identified. Additional study is required to develop new model systems and approaches to determine the role of targeted therapy in the treatment of PI3K-overactive HNSCC tumors.

Objectives: This study was designed to evaluate the effects of chronic cyclosporine A (CsA) treatment and acute renal ischaemia/reperfusion (I/R) on the kidney and liver in thymoquinone (TQ)-treated rats., Methods: In the CsA study, adult male rats were divided into control, CsA (25 mg/kg per day), TQ (10 mg/kg per day) and CsA + TQ groups, and rat treatment was for 28 days. In the I/R study, adult male rats were divided into sham-operated, I/R (renal ischaemia for 60 min followed by 60 min reperfusion) and TQ + I/R (TQ 10 mg/kg, 24 h and 1 h before ischaemia) groups., Key Findings: CsA treatment and renal I/R caused kidney and liver dysfunction as evaluated by histopathological changes and biochemical parameters. TQ treatment reduced elevated serum indices back to control levels and ameliorated CsA-induced kidney and liver histopathological changes. In renal and hepatic tissues, CsA and renal I/R induced significant increases in malondialdehyde levels with significant decreases in reduced glutathione levels and superoxide dismutase activities. Such changes in oxidative stress markers were counteracted by TQ treatment., Conclusions: Kidney and liver injury due to CsA or renal I/R can be significantly reduced by TQ, which resets the oxidant/antioxidant balance of the affected organs through scavenging free radicals and antilipoperoxidative effects., (© 2015 Royal Pharmaceutical Society.)

The protective effect of thymoquinone was investigated against cadmium-induced testicular toxicity in rats. Testicular toxicity was induced by a single intraperitoneal (i.p.) injection of cadmium chloride (2 mg kg(-1) ). Thymoquinone treatment (10 mg kg(-1)  day(-1) , i.p.) was applied for five consecutive days, starting 3 days before cadmium administration. Thymoquinone significantly attenuated the cadmium-induced decreases in serum testosterone, and testicular reduced glutathione and superoxide dismutase activity and significantly decreased the elevations of testicular malondialdehyde, nitric oxide and cadmium ion levels resulted from cadmium chloride administration. Also, thymoquinone ameliorated the cadmium-induced testicular tissue injury observed by histopathological examination. In addition, thymoquinone significantly decreased the cadmium-induced expression of inducible nitric oxide synthase, tumour necrosis factor-α, cyclooxygenase-2, nuclear factor-κB and caspase-3 in testicular tissue. It was concluded that thymoquinone, through its antioxidant and anti-inflammatory activities, may represent a potential candidate to protect the testes against the detrimental effect of cadmium exposure., (© 2014 Blackwell Verlag GmbH.)

Herbal medicine has attracted great attention in the recent years and is increasingly used as alternatives to chemical drugs. Several lines of evidence support the positive impact of medicinal plants in the prevention and cure of a wide range of diseases. Thymoquinone (TQ) is the most abundant constituent of the volatile oil of Nigella sativa seeds and most properties of N sativa are mainly attributed to TQ. A number of pharmacological actions of TQ have been investigated including anti-oxidant, anti-inflammatory, immunomodulatory, anti-histaminic, anti-microbial and anti-tumor effects. It has also gastroprotective, hepatoprotective, nephroprotective and neuroprotective activities. In addition, positive effects of TQ in cardiovascular disorders, diabetes, reproductive disorders and respiratory ailments, as well as in the treatment of bone complications as well as fibrosis have been shown. In addition, a large body of data shows that TQ has very low adverse effects and no serious toxicity. More recently, a great deal of attention has been given to this dietary phytochemical with an increasing interest to investigate it in pre-clinical and clinical researches for assessing its health benefits. Here we report on and analyze numerous properties of the active ingredient of N. sativa seeds, TQ, in the context of its therapeutic potentials for a wide range of illnesses. We also summarize the drug's possible mechanisms of action. The evidence reported sugests that TQ should be developed as a novel drug in clinical trials., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Objective: To determine the anti-inflammatory effects of thymoquinone on body weight, clinical score of inflammation, total leukocyte count and differential leukocyte count in arthritic rats and compare it with that of methotrexate., Methods: The study was conducted at the Post-Graduate Medical Institute, Lahore, from March to September 2013, and comprised female Sprague-Dawley rats randomised into four equal groups; group A (healthy control), group B (positive control), group C (thymoquinone treated) and group D (methotrexate treated). Arthritis developed in Group B, C and D within two weeks after a single intra-dermal injection of pristane. Body-weight measured on electronic balance in grams and clinical score of inflammation scored on macroscopic scoring system were monitored on every alternate day while total leukocyte count and differential leukocyte count were taken at day 0, 16 and 30. After day 15, groups A and B were given 0.5ml of distilled water by intra-peritoneal injection daily for 15 consecutive days; group C was given thymoquinone 2mg/kg by intra-peritoneal injection daily for 15 consecutive days, and group D received methotrexate 0.5mg/kg by intra-peritoneal injection, daily for 15 consecutive days. SPSS 20 was used for statistical analysis., Results: The 32 rats in the study were randomised into four groups of 8(25%) each. In group A the body-weight continued to increase and reached a mean of 144.13±10.8% of the baseline at day 30. In group B the weight reduced to 93.13±4.19% at day 16 and to 88.3±6.97% at day 30. In groups C and D the weight reduced to 87.25±7.69% and 88.5±7.07% respectively at day 16; then the animals in the two groups regained their weight which increased to 108.63±10.89% and 103.38±6.25% respectively at day 30. The score was zero in group A throughout the study period. The score of group B, which was zero at day 0, reached a mean of 16±0 at day 16. In groups C and D, the mean score increased till day 16 and reached 16±1 and 16±0 respectively, and then reduced to 5±2 and 4±1 at day 30 respectively., Conclusions: Evaluation of data supported the anti-inflammatory activities of thymoquinone, so it may be investigated as an effective anti-inflammatory drug in rheumatoid arthritis.

Abdominal aortic aneurysm (AAA) is a common degenerative vascular disease whose pathogenesis is associated with activation of multiple signaling pathways including Jun NH2-terminal kinases (JNK) and NF-κB. It is now well recognized that these pathways are chaperoned by the heat shock protein 90 (Hsp90), suggesting that inhibition of Hsp90 may be a novel strategy for inhibiting AAAs. The aim of this study is to investigate whether inhibition of Hsp90 by 17-DMAG (17-dimethyl-aminothylamino-17-demethoxy-geldanamycin) attenuates ANG II-induced AAA formation in mice, and, if so, to elucidate the mechanisms involved. Apolipoprotein E-null mice were infused with ANG II to induce AAA formation and simultaneously treated by intraperitoneal injection with either vehicle or 17-DMAG for 4 wk. ANG II infusion induced AAA formation in 80% of mice, which was accompanied by increased matrix metalloproteinase (MMP) activity, enhanced tissue inflammation, oxidative stress, and neovascularization. Importantly, these effects were inhibited by 17-DMAG treatment. Mechanistically, we showed that 17-DMAG prevented the formation and progression of AAA through its inhibitory effects on diverse biological pathways including 1) by blocking ANG II-induced phosphorylation of ERK1/2 and JNK that are critically involved in the regulation of MMPs in vascular smooth muscle cells, 2) by inhibiting IκB kinase expression and expression of MCP-1, and 3) by attenuating ANG II-stimulated angiogenic processes critical to AAA formation. Our results demonstrate that inhibition of Hsp90 by 17-DMAG effectively attenuates ANG II-induced AAA formation by simultaneously inhibiting vascular inflammation, extracellular matrix degradation, and angiogenesis, which are critical in the formation and progression of AAAs., (Copyright © 2015 the American Physiological Society.)

Embelin has been used to treat fever and inflammatory diseases for thousands of years. Although reports indicate that embelin has antiinflammatory effects, its effects on myocardial injury following cardiac arrest (CA) have not been previously explored. In this study, we aim to investigate the protective effects of embelin on myocardial ischemia-reperfusion injury (IRI) following CA in a rabbit model. Pro-inflammatory (TNF-α, IL-1β, and IL-6) cytokines, cardiac troponin I (cTnI), necrosis ratio, apoptotic index (AI), hemodynamics, nuclear factor-kappa B (NF-κB) p65, and histological damage have been measured or evaluated. Embelin reverts TNF-α, IL-1β, and IL-6 to basal levels and reduces the serum level of cTnI, the necrosis ratio, the AI, and the expression of NF-κB p65. Meanwhile, it improves the hemodynamics and myocardial function. Moreover, embelin-treated groups also showed improved myocardial morphology. Our results indicate that embelin may protect the heart against myocardial IRI following CA via its antiinflammatory abilities.

The effect of 17-allylamino-17-demethoxygeldanamycin (17-AAG), an hsp90 inhibitor, alone or in combination with paclitaxel on survival of anaplastic thyroid carcinoma (ATC) was evaluated. In 8505C and CAL62 cells, after treatment of 17-AAG, cell viability decreased, and the percentage of dead cells increased. 17-AAG did not cause cleavage of caspase-3 protein, and change expression of IAPs. Pretreatment of z-VAD-fmk did not alter cell viability and the percentage of dead cells. In 17-AAG-treated cells, knockdown of p53 rescued growth inhibition, while cycloheximide attenuated cell death. When cells were treated with both 17-AAG and paclitaxel, all of the combination index values were higher than 1, indicating antagonism between 17-AAG and paclitaxel. In 17-AAG- and paclitaxel-treated cells, compared with paclitaxel alone-treated cells, the protein levels of hsp90, hsp70, and hsc70 increased. In conclusion, our results suggest that 17-AAG induces non-apoptotic cell death requiring de novo protein synthesis in ATC cells. Moreover, these results demonstrate that 17-AAG antagonizes paclitaxel with concomitant alterations in hsp90 client proteins in ATC cells.

Background: The gemcitabine-insensitivity remains the main challenge for pancreatic cancer treatment. Thymoquinone, the predominant bioactive ingredient of Nigella sativa, has been shown to possess promising anti-cancer and chemo-sensitizing effects on pancreatic cancer, however, its meticulous mechanism is still indistinct., Aim: The objective of the present study was to investigate the potency of thymoquinone in combination with gemcitabine in inducing apoptosis and preventing the development of gemcitabine-insensitivity in pancreatic cancer cells., Methods: The anti-tumor effects of thymoquinone and gemcitabine were analyzed via evaluation of alterations of cell viability, tumor weight, apoptosis-related proteins, caspase-3, -9 activities and NF-κB DNA binding activity in pancreatic cancer cells in vitro and PANC-1 cells orthotopic xenograft in vivo., Results: Thymoquinone pretreatment following gemcitabine treatment synergistically caused an increase in pancreatic cancer cells apoptosis and tumor growth inhibition both in vitro and in vivo. The novel combinational regimen also contributes to alterations of multiple molecular signaling targets, such as the suppression of Notch1, NICD accompanying with up-regulation of PTEN, the inactivation of Akt/mTOR/S6 signaling pathways, and the suppression of phosphorylation and nuclear translocation of p65 induced by TNF-α. Thymoquinone pretreatment and gemcitabine also induced down-regulation of anti-apoptotic Bcl-2, Bcl-xL, XIAP and up-regulation and activation of pro-apoptotic molecules including Caspase-3, Caspase-9, Bax and increased release of cytochrome c., Conclusions: This novel modality of thymoquinone pretreatment can enhance the anti-cancer activity of gemcitabine and may be a promising option in the treatment of pancreatic cancer.

Hsp90 inhibitors have become well-studied antitumor agents for their selective property against tumors versus normal cells. The combined treatment of Hsp90 inhibitor and conventional photon radiation also showed more effective tumor growth delay than radiation alone. However, little is known regarding the combined treatment of Hsp90 inhibitor and heavy-ion irradiation. In this study, SQ5 human lung tumor cells were used in vitro for clonogenic cell survival and in vivo for tumor growth delay measurement using a mouse xenograft model after 17-allylamino-17-demethoxygeldanamycin (17AAG) pretreatment and carbon ion irradiation. Repair of DNA double strand breaks (DSBs) was also assessed along with expressions of DSB repair-related proteins. Cell cycle analysis after the combined treatment was also performed. The combined treatment of 17AAG and carbon ions revealed a promising treatment option in both in vitro and in vivo studies. One likely cause of this effectiveness was shown to be the inhibition of homologous recombination repair by 17AAG. The more intensified G2 cell cycle delay was also associated with the combined treatment when compared with carbon ion treatment alone. Our findings indicate that the combination of Hsp90 inhibition and heavy-ion irradiation provides a new effective therapeutic alternative for treatment of solid tumors., (© 2015 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Epilepsy is a common neurological disorder that leads to neuronal excitability and provoke various forms of cellular reorganization in the brain. In this study, we investigate the anti-convulsant and neuroprotective effects of thymoquinone (TQ) and vitamin C against pentylenetetrazole (PTZ)-induced generalized seizures. Epileptic seizures were induced in adult rats using systemic intraperitoneal injections of PTZ (50 mg/kg) for 7 days. Animals pretreated with either TQ or vitamin C or in combination attenuated PTZ-induced seizures and mortality in rats as well neurodegeneration in the cells. Compared to PTZ, TQ and vitamin C significantly prolonged the onset of seizures (p > 0.05) as well decrease the high-grade seizures. Analysis of electroencephalogram (EEG) recordings revealed that TQ or vitamin C supplementation significantly reduced polyspike and epileptiform discharges. Epileptic seizures caused a decline in expression of gamma-aminobutyric acid B1 receptor (GABAB1R) (p > 0.05), unchanged expression of protein kinase A (PKA), decreased calcium/calmodulin-dependent protein kinase II (CaMKII) (p > 0.05) and inhibit the phosphorylation of cAMP response element-binding protein (CREB) (p > 0.05) in cortex and hippocampus, respectively, compared with control. Changes in expression of GABAB1R, CaMKII and CREB by PTZ were reversed by TQ and vitamin C supplementation. Moreover, PTZ significantly increased Bax, decreased Bcl-2 expression and finally the activation of caspase-3. TQ and vitamin C pretreatment reversed all these deleterious effects induced by PTZ. TQ and vitamin C showed anticonvulsant effects via activation of GABAB1R/CaMKII/CREB pathway and suggest a potential therapeutic role in epilepsy.

Objective: Serious health problems in humans are caused by arsenic (As) exposure, which is wide spread in the environment. Sodium arsenite (SAs), capable of inducing macromolecular damage is evaluated for its damaging effect in the blood vessels, liver and kidneys of Wistar rats. This study was undertaken to investigate the ameliorative effects of thymoquinone on SAs-induced oxidative and inflammatory damages in the serum of male Wistar rats., Materials and Methods: Wistar Albino rats divided into three groups of nine rats each were administered to controls saline (10 mg/kg), SAs (10 mg/kg), and SAs plus thymoquinone (10 mg/kg/day) for two weeks orally. Biochemical tests were analyzed by a otoanalyzer; nitric oxide levels specthrophometrically, and cytokines were measured by ELISA method in the rat serum samples., Results: Inflammatory cytokines and some biochemical variables were found to be increased in the SAs group compared to control group. On the other hand, thymoquinone supressed these laboratory signs, which are thought to be the characteristic signs of SAs toxicity, most probably by its ameliorative effects including anti-inflammatory and antioxidant properties., Conclusions: From the results obtained, thymoquinone mitigates SAs-induced adverse effects in the serum of rats, which suggest that it may attenuate inflammation implicated in endotelial dysfunction.

Objective: To investigate whether thymoquinone has any eliminative effects against inner-ear damage caused by acoustic trauma., Methods: Thirty-two male rats were divided into four groups. Group 1 was only exposed to acoustic trauma. Group 2 was given thymoquinone 24 hours before acoustic trauma and continued to receive it for 10 days after the trauma. Group 3 was only treated with thymoquinone, for 10 days. Group 4, the control group, suffered no trauma and received saline instead of thymoquinone. Groups 1 and 2 were exposed to acoustic trauma using 105 dB SPL white noise for 4 hours., Results: There was a significant decrease in distortion product otoacoustic emission values and an increase in auditory brainstem response thresholds in group 1 on days 1, 5 and 10, compared with baseline measurements. In group 2, a decrease in distortion product otoacoustic emission values and an increase in auditory brainstem response threshold were observed on day 1 after acoustic trauma, but measurements were comparable to baseline values on days 5 and 10. In group 3, thymoquinone had no detrimental effects on hearing. Similarly, the control group showed stable results., Conclusion: Thymoquinone was demonstrated to be a reparative rather than preventive treatment that could be used to relieve acoustic trauma.

Aim: We aimed to investigate efficacy of intraperitoneally administered Thymoquinone (TQ) in acetaminophen (APAP) induced renal toxicity., Material and Method: Forty Wistar Albino rats were randomly divided into 4 groups of ten rats each. Control group was untreated group while rats in TQ group were treated with single dose TQ. In APAP group rats were treated with single dose acetaminophen. In TQ + APAP group TQ and APAP were administered respectively. Rats were sacrificed at 24th hour; urea, creatinine and nitric oxide levels were measured also malondialdehyde activity were assessed in renal tissue specimens. Tissue damage scores were recorded in histopathological assessment., Results: Urea and creatinine levels were found significantly higher in APAP group than control group (p < 0.003). Urea and creatinine levels in APAP + TQ treated group were significantly lower than APAP treated group (p < 0.01). Serum NO activity and tissue MDA levels were higher in APAP group than control group (p ≤ 0.002). In contrast to APAP treated group serum NO activity and tissue MDA levels were found significantly lower in TQ + APAP group (p ≤ 0.03). In APAP group significant histopathological change was found compared with control group (p ≤ 0.001) where there was no significant change between control and TQ treated groups (p > 0.05). In APAP group we found significantly higher tissue damage scores than control group (p ≤ 0.001). In APAP + TQ group lower tissue damage scores were found compared with APAP group (p ≤ 0.001) while higher scores were found compared with control group (p ≤ 0.001)., Conclusion: We can conclude that TQ treatment has therapeutical effect in APAP induced nephrotoxicity in rats., (Copyright © 2014 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.)

Introduction: Previous studies have demonstrated that thymoquinone has protective effects against ischemia reperfusion injury to various organs like lungs, kidneys and liver in different experimental models., Objective: We aimed to determine whether thymoquinone has favorable effects on lung, renal, heart tissues and oxidative stress in abdominal aorta ischemia-reperfusion injury., Methods: Thirty rats were divided into three groups as sham (n=10), control (n=10) and thymoquinone (TQ) treatment group (n=10). Control and TQ-treatment groups underwent abdominal aorta ischemia for 45 minutes followed by a 120-min period of reperfusion. In the TQ-treatment group, thymoquinone was given 5 minutes. before reperfusion at a dose of 20 mg/kg via an intraperitoneal route. Total antioxidant capacity, total oxidative status (TOS), and oxidative stress index (OSI) in blood serum were measured and lung, kidney, and heart tissue histopathology were evaluated with light microscopy., Results: Total oxidative status and oxidative stress index activity in blood samples were statistically higher in the control group compared to the sham and TQ-treatment groups (P<0.001 for TOS and OSI). Control group injury scores were statistically higher compared to sham and TQ-treatment groups (P<0.001 for all comparisons)., Conclusion: Thymoquinone administered intraperitoneally was effective in reducing oxidative stress and histopathologic injury in an acute abdominal aorta ischemia-reperfusion rat model.

Objective: To investigate the effect of Embelin, an inhibitor of X-Linked Inhibitor of Apoptosis Protein (XIAP), on inflammation and bone erosion in a collagen antibody induced arthritis (CAIA) in mice., Methods: Four groups of mice (n = 6 per group) were allocated: CAIA untreated mice, CAIA treated with Prednisolone (10 mg/kg/day), CAIA treated with low dose Embelin (30 mg/kg/day), and CAIA treated with high dose Embelin (50 mg/kg/day). Joint inflammation was evaluated using clinical paw score and histological assessments. Bone erosion was assessed using micro-CT, tartrate resistant acid phosphatase (TRAP) staining, and serum carboxy-terminal collagen crosslinks (CTX-1) ELISA. Immunohistochemistry was used to detect XIAP protein. TUNEL was performed to identify apoptotic cells., Results: Low dose, but not high dose Embelin, suppressed inflammation as reflected by lower paw scores (P < 0.05) and lower histological scores for inflammation. Low dose Embelin reduced serum CTX-1 (P < 0.05) and demonstrated lower histological score and TRAP counting, and slightly higher bone volume as compared to CAIA untreated mice. XIAP expression was not reduced but TUNEL positive cells were more abundant in Embelin treated CAIA mice., Conclusion: Low dose Embelin suppressed inflammation and serum CTX-1 in CAIA mice, indicating a potential use for Embelin to treat pathological bone loss.

Background: Gentamicin (GEN) is considered as a main aminoglycoside antibiotic medicine. The top therapeutic side effect of GEN is nephrotoxicity. The current research was proposed to determine the protective effect of thymoquinone (TQ) on GEN-induced acute renal failure (ARF)., Methods: The rats were divided into 6 groups: sham group, control group, GEN-treated group and the TQ-treated groups. At the end of the research period, the serum blood urea nitrogen (BUN) and creatinine (Cr) were measured. The kidneys were then removed for evaluating malondialdehyde (MDA), reduced glutathione (GSH), interleukins IL-6, IL-10, IL-18 and IL-1β, tumor necrosis factor α (TNF-α), superoxide dismutase (SOD) and glutathione peroxidase (GPx) levels., Results: GEN induced a raise in the levels of serum Cr, BUN and also the levels of MDA, IL-6, IL-18, IL-1β and TNF-α with decease in GSH, SOD, GPx and IL-10 in the kidney (p < 0.05). The data illustrated that the significant elevation in the levels of serum Cr, BUN, MDA, IL-6, IL-18, IL-1β, TNF-α and also the reduction of GSH, SOD, SOD, GPx, IL-10 in the kidney were ameliorated in the TQ-treated groups versus the untreated group, in a dose-dependent manner (p < 0.05)., Conclusion: The present investigation proposes that TQ may be ameliorated ARF through modulation of the oxidative stress and inflammatory responses., (© 2015 S. Karger AG, Basel.)

Background: Thymoquinone is an active ingredient isolated from Nigella sativa (Black Seed). This study aimed to evaluate the in vitro and in vivo anti-leukemic effects of thymoquinone on WEHI-3 cells., Methodology/principal Findings: The cytotoxic effect of thymoquinone was assessed using an MTT assay, while the inhibitory effect of thymoquinone on murine WEHI-3 cell growth was due to the induction of apoptosis, as evidenced by chromatin condensation dye, Hoechst 33342 and acridine orange/propidium iodide fluorescent staining. In addition, Annexin V staining for early apoptosis was performed using flowcytometric analysis. Apoptosis was found to be associated with the cell cycle arrest at the S phase. Expression of Bax, Bcl2 and HSP 70 proteins were observed by western blotting. The effects of thymoquinone on BALB/c mice injected with WEHI-3 cells were indicated by the decrease in the body, spleen and liver weights of the animal, as compared to the control., Conclusion: Thymoquinone promoted natural killer cell activities. This compound showed high toxicity against WEHI-3 cell line which was confirmed by an increase of the early apoptosis, followed by up-regulation of the anti-apoptotic protein, Bcl2, and down-regulation of the apoptotic protein, Bax. On the other hand, high reduction of the spleen and liver weight, and significant histopathology study of spleen and liver confirmed that thymoquinone inhibited WEHI-3 growth in the BALB/c mice. Results from this study highlight the potential of thymoquinone to be developed as an anti-leukemic agent.

This study was conducted to assess the preventive effect of two plant constituents, thymoquinone and curcumin, on gentamicin-induced deleterious effect on liver function, integrity and histological architecture. The gentamicin was intraperitoneally injected to rats at dose level of 100 mg/kg b.w. (every other day) for 21 days. The thymoquinone and curcumin were concurrently and orally administered at dose level of 20 mg/kg b.w. (every other day) to gentamicin-injected rats. The present data indicated that thymoquinone and curcumin significantly prevented the gentamicin-induced elevations of serum AST, ALT and LDH activities as well as tumor necrosis factor alpha (TNF-α) and total bilirubin levels. On the other hand, both agents markedly ameliorated the gentamicin-induced decrease in serum total protein, albumin and albumin/globulin ratio. In addition, the gentamicin-induced liver histological alterations including hydropic degeneration of hepatocytes, fatty changes, inflammatory cell infiltration and congestion of portal vein were successfully amended by thymoquinone and curcumin. The elevated proapoptotic proteins caspase 3 and Bax expression in cytoplasm and nucleus of hepatocytes of gentamicin-injected rats were reduced to normal value as a result of thymoquinone and curcumin administration while the lowered expression of antiapoptotic protein Bcl-2 was increased. Based on the previous findings, it can be concluded that thymoquinone and curcumin successfully prevents the deleterious effects on liver function and histological integrity to more or less the same degree by enhancing anti-oxidant defense system, suppression of oxidative stress and attenuation of inflammation and apoptosis.

Background: Metabolic syndrome is an important risk factor for cardiovascular disease (CVD) occurrence and mortality. CVDs are leading cause of death worldwide. Recently, there has been an increasing interest in the use of herbal medicines with more efficiency and minimal undesirable effects than chemical drugs for a variety of disorders including CVD. Nigella sativa and its active constituent, thymoquinone, have been documented to exhibit antidiabetic, antiobesity, hypotensive and hypolipidemic properties., Aim: In this review, we discussed the most relevant articles to find out the role of N. sativa in different components of metabolic syndrome and CVD risk factors including high blood pressure, obesity, dyslipidemia and high blood glucose., Conclusions: This review suggests a potential role of N. sativa and TQ in the management of metabolic syndrome, however more studies should be conducted to evaluate their effectiveness.

Background: Leishmaniasis remains a worldwide public health problem. The limited therapeutic options, drug toxicity and reports of resistance, reinforce the need for the development of new treatment options. Previously, we showed that 17-(allylamino)-17-demethoxygeldanamycin (17-AAG), a Heat Shock Protein 90 (HSP90)-specific inhibitor, reduces L. (L.) amazonensis infection in vitro. Herein, we expand the current knowledge on the leishmanicidal activity of 17-AAG against cutaneous leishmaniasis, employing an experimental model of infection with L. (V.) braziliensis., Methodology/principal Findings: Exposure of axenic L. (V.) braziliensis promastigotes to 17-AAG resulted in direct dose-dependent parasite killing. These results were extended to L. (V.) braziliensis-infected macrophages, an effect that was dissociated from the production of nitric oxide (NO), superoxide (O(-2)) or inflammatory mediators such as TNF-α, IL-6 and MCP-1. The leishmanicidal effect was then demonstrated in vivo, employing BALB/c mice infected with L. braziliensis. In this model, 17-AAG treatment resulted in smaller skin lesions and parasite counts were also significantly reduced. Lastly, 17-AAG showed a similar effect to amphotericin B regarding the ability to reduce parasite viability., Conclusion/significance: 17-AAG effectively inhibited the growth of L. braziliensis, both in vitro and in vivo. Given the chronicity of L. (V.) braziliensis infection and its association with mucocutaneous leishmaniasis, 17-AAG can be envisaged as a new chemotherapeutic alternative for cutaneous Leishmaniasis.

The bioactive natural products (plant secondary metabolites) are widely known to possess therapeutic value for the prevention and treatment of various chronic diseases including cancer. Thymoquinone (2-methyl-5-isopropyl-1,4-benzoquinone; TQ), a monoterpene present in black cumin seeds, exhibits pleiotropic pharmacological activities including antioxidant, anti-inflammatory, antidiabetic and antitumor effects. TQ inhibits experimental carcinogenesis in a wide range of animal models and has been shown to arrest the growth of various cancer cells in culture as well as xenograft tumors in vivo. The mechanistic basis of anticancer effects of TQ includes the inhibition of carcinogen metabolizing enzyme activity and oxidative damage of cellular macromolecules, attenuation of inflammation, induction of cell cycle arrest and apoptosis in tumor cells, blockade of tumor angiogenesis, and suppression of migration, invasion and metastasis of cancer cells. TQ shows synergistic and/or potentiating anticancer effects when combined with clinically used chemotherapeutic agents. At the molecular level, TQ targets various components of intracellular signaling pathways, particularly a variety of upstream kinases and transcription factors, which are aberrantly activated during the course of tumorigenesis., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Introduction: Extramedullary myeloma that occurs during the clinical course of multiple myeloma is rare but is an independent poor prognostic factor with mortality of 73% and median survival of 12 months despite aggressive therapies including novel agents. The clinicopathological aspects, biology and management of extramedullary myelomas are poorly understood. Our case highlights the pathobiological aspects of this important but rare entity, and the repercussions of modern therapies., Case Presentation: A 60-year-old Caucasian man initially presented with an anterior rib fracture. Subsequent workup revealed stage IIIB immunoglobulin G lambda multiple myeloma. A bone marrow biopsy showed sheets of plasma cells, harboring unfavorable cytogenetics including deletion of 17p and t(4;14). He achieved near complete remission and resolution of karyotypic abnormalities with three cycles of induction doxorubicin, thalidomide, and dexamethasone (clinical trial). This was followed by high-dose melphalan and autologous stem cell transplant. He relapsed 1 year later. His bone marrow at that time showed only a few scattered polyclonal plasma cells. He received three cycles of bortezomib and tanespimycin (clinical trial) and achieved very good partial response. He again relapsed 1 year later with multiple large peripheral soft tissue masses and lymph nodes. Biopsies of the peripheral lesions were consistent with extramedullary myeloma, but repeat bone marrow biopsy continued to show no evidence of intramedullary disease., Conclusions: This is one of the few cases reported that illustrates the differential response of extramedullary compared to intramedullary myeloma to multiple standard combination therapies including novel therapeutics and transplant, resulting in a very short survival. Several mechanisms for intra-to-extra medullary migration and hence the differential treatment response have been hypothesized. Physicians should be aware of this problem during treatment with immunomodulatory drugs and proteasome inhibitors not only in relapsed but also in front-line setting. In such cases, there is a potential role for evolving targeted therapeutics as we continue to better understand the tumor biology.

Background: Spinal cord injury (SCI) is one of the most devastating conditions leading to neurological impairment and disabilities. The aim of the study was to investigate the potential neuroprotective effect of thymoquinone (TQ) histopathologically in an experimental model of traumatic spinal cord injury (SCI)., Methods: Twenty-four male Wistar albino rats were randomly divided into 4 groups: control group; SCI group; SCI-induced and 10 mg/kg/day TQ administered group; SCI-induced and 30 mg/kg/day TQ administered group. TQ was given as intraperitoneal for three days prior to injury and four days following injury. Spinal cord segment between T8 and T10 were taken for histopathologic examination. Hemorrhage, spongiosis and liquefactive necrosis were analyzed semiquantatively for histopathological changes., Results: Administration of TQ at a dose of 10 mg/kg did not cause any significant change on the histological features of neuronal degeneration as compared to the SCI group (p=0.269); however, 30 mg/kg TQ significantly decreased the histological features of spinal cord damage below that of the SCI group (p=0.011)., Conclusion: Data from this study suggest that TQ supplementation attenuates trauma induced spinal cord damage. Thus, TQ needs to be taken into consideration, for it may have a neuroprotective effect in trauma induced spinal cord damage.