SelSA-1, a novel HDAC inhibitor demonstrates enhanced chemotherapeutic potential by redox modulation.

Colorectal cancer (CRC) is a multistep disorder resulting from genetic and epigenetic genome changes. It is the third most common malignancy in developed nations accounting for roughly 600,000 deaths annually. Persistent gut inflammation, as observed in inflammatory bowel disease (IBD), is a key risk factor for CRC development. From an epigenetic viewpoint, the pharmacological inhibition of HDACs using HDAC inhibitors such as SAHA has emerged as a suitable anticancer strategy in the recent past. However, the clinical success of these strategies is limited and has risk factors associated with their uses. Thus, considering the critical involvement of epigenetic regulation of key molecular mechanisms in carcinogenesis as well as HDAC inhibitory and anti-tumorigenic properties of Selenium (Se), we aimed to explore the potentially safer and enhanced chemotherapeutic potential of a Se derivative of SAHA namely SelSA-1, in an experimental model of colitis-associated experimental cancer (CAC) model and mechanism involved therein. The in vitro study indicated improved efficiency, specificity, and better safety margin in terms of lower IC50 value of SelSA-1 than SAHA in both NIH3T3 (9.44 and 10.87 µM) and HCT 115 (5.70 and 7.49 µM) cell lines as well on primary colonocytes (5.61 and 6.30 µM) respectively. In an in vivo experimental model, SelSA-1 efficiently demonstrated amelioration of the multiple plaque lesions (MPLs), tumor burden/incidence, and modulation of various histological and morphological parameters. Further, redox-mediated alterations in apoptotic mediators suggested induction of cancer cell apoptosis by SelSA-1. These findings indicate the enhanced chemotherapeutic and pro-resolution effects of SelSA-1 in part mediated through redox modulation of multiple epigenetic and apoptotic pathways.

The relevance of arsenic speciation analysis in health & medicine.

Long term exposure to arsenic through consumption of contaminated groundwater has been a global issue since the last five decades; while from an alternate standpoint, arsenic compounds have emerged as unparallel chemotherapeutic drugs. This review highlights the contribution from arsenic speciation studies that have played a pivotal role in the progression of our understanding of the biological behaviour of arsenic in humans. We also discuss the limitations of the speciation studies and their association with the interpretation of arsenic metabolism. Chromatographic separation followed by spectroscopic detection as well as the utilization of biotinylated pull-down assays, protein microarray and radiolabelled arsenic have been instrumental in identifying hundreds of metabolic arsenic conjugates, while, computational modelling has predicted thousands of them. However, these species exhibit a variegated pattern, which supports more than one hypothesis for the metabolic pathway of arsenic. Thus, the arsenic species are yet to be integrated into a coherent mechanistic pathway depicting its chemicobiological fate. Novel biorelevant arsenic species have been identified due to significant evolution in experimental methodologies. However, these methods are specific for the identification of only a group of arsenicals sharing similar physiochemical properties; and may not be applicable to other constituents of the vast spectrum of arsenic species. Consequently, the identity of arsenic binding partners in vivo and the sequence of events in arsenic metabolism are still elusive. This resonates the need for additional focus on the extraction and characterization of both low and high molecular weight arsenicals in a combinative manner.

Selenium-Based Novel Epigenetic Regulators Offer Effective Chemotherapeutic Alternative with Wider Safety Margins in Experimental Colorectal Cancer.

Colorectal cancer (CRC) is a major cause of morbidity and mortality worldwide. Despite the critical involvement of epigenetic modifications in CRC, the studies on the chemotherapeutic efficacy of various epigenetic regulators remain limited. Considering the key roles of histone deacetylases (HDACs) in the regulation of diverse cellular processes, several HDAC inhibitors are implied as effective therapeutic strategies. In this context, suberoylanilide hydroxamic acid (SAHA), a 2nd-generation HDAC inhibitor, showed limited efficacy in solid tumors. Also, side effects associated with SAHA limit its clinical application. Based on the redox-modulatory and HDAC inhbitiory activities of essential trace element selenium (Se), the anti-carcinogenic potential of Se substituted SAHA, namely, SelSA-1 (25 mg kg-1), was screened for it enhanced anti-tumorigenic role and wider safety profiles in DMH-induced CRC in Balb/c mice. A multipronged approach such as in silico, biochemical, and pharmacokinetics (PK) has been used to screen, characterize, and evaluate these novel compounds in comparison to existing HDAC inhibitor SAHA. This is the first in vivo study indicating the chemotherapeutic potential of Se-based novel epigenetic regulators such as SelSA-1 in any in vivo experimental model of carcinogenesis. Pharmcological and toxicity data indicated better safety margins, bioavailability, tolerance, and elimination rate of SelSA-1 compared to classical HDAC inhibitor SAHA. Further, histological and morphological evidence demonstrated enhanced chemotherapeutic potential of SelSA-1 even at lower pharmacological doses than SAHA. This is the first in vivo study suggesting Se-based novel epigenetic regulators as potential chemotherapeutic alternatives with wider safety margins and enhanced anticancer activities.

Selenium Ameliorates Ibuprofen Induced Testicular Toxicity by Redox Regulation: Running Head: Se protects against NSAID induced testicular toxicity.

Despite the Cox inhibitory anti-inflammatory and antipyretic effects of most widely used non-steroidal anti-inflammatory drugs (NSAIDs), such as Ibuprofen, their chronic use is associated with a plethora of patho-physiological insults. One such toxic effect on testicular tissues is not well studied and the underlying molecular mechanisms remain unexplored. Thus, the current study is designed to evaluate the antioxidant properties of essential trace element selenium (Se) to ameliorative Ibuprofen associated testicular toxic effects. Adult male Wistar rats were divided into 3 groups and fed on diets containing different concentrations of sodium selenite, viz. 0.01 mg/kg (Se- deficient), 0.2 mg/kg (Se-adequate), or 0.5 mg/kg (Se- supplemented) for 8 weeks. After diet feeding schedule, each group was divided into two subgroups i.e., with or without the treatment of Ibuprofen (120 mg/kg Bw). The protective effect of Se was evaluated by measuring testicular Se and selenoproteins status, spermatogenic markers, histopathology and testicular redox status. Ibuprofen diminished seminal volume, sperm count, sperm motility, which correlated well increased testicular reactive oxygen species. Se deficiency exacerbated these detrimental effects of ibuprofen by increasing oxidative stress. Alternatively, Se supplementation through antioxidant enzymes mediated protective effects. Se as essential antioxidant selenoproteins ameliorates Ibuprofen induced male reproductive toxicity.

Redox modulatory protective effects of ω-3 fatty acids rich fish oil against experimental colitis.

Ulcerative colitis (UC), a form of inflammatory bowel disease (IBD), is an immune-modulated disorder characterized by chronic and recurring inflammatory episodes. Oxidative stress and COX pathway of prostaglandin (PG) biosynthesis are indispensable to pathogenesis of UC. Any imbalance between PGs can compromise the mucosal homeostasis, leading to mucosal damage and chronic inflammation. However, blocking these PGs using classical Cox inhibitors such as non-steroidal anti-inflammatory drugs (NSAIDs) can instead aggravate signs of IBD. Therefore, realizing the need for safer and well tolerable alterative treatment approaches, currently, we evaluated the efficacy of n-3 fatty acids rich fish oil (FO) in the resolution of UC. Using a dextran sodium sulfate (DSS) model of experimental colitis, we have demonstrated that supplementation of FO containing 180 mg EPA and 120 mg DHA for 1 month relieved the signs (diarrhea, bloody stools, weight loss) of colitis-associated inflammation. To understand the biophysical changes associated with FO mediated inflammatory regulation, impedance measurement and Fourier transform infrared spectroscopy (FTIR) were done. These changes were also correlated with oxidative stress through markers such as GST, glutathione peroxidase (GPx), LPO, catalase, protein carbonyl content, GR, etc. in colonic mucosa. The modulation of COX mediated pathways in UC-associated inflammation was observed by protein expressions of various pro-inflammatory cytokines such as TNF-α and enzymes of PG synthesis such as COX-2, PGES, TXAS, and anti-inflammatory PGDS. Refuting the earlier reports that suggested the contradictory effects of FO, in the current study, we evidently demonstrated that the protective effects of FO are mediated through molecular mechanisms involving the redox-regulation of metabolism of key lipid metabolites.

Leukocyte Cell Population Data for Hematology Analyzer-Based Distinction of Clonal-versus-Non-Clonal Lymphocytosis: A Real-World Testing Experience.

Automated blood counts revealing lymphocytosis necessitate smear reviews. Even expert morphological evaluation may however, fail to differentiate a benign-versus-malignant etiology without further testing. Automated analyser-derived quantitative data on leukocyte cell populations remain undertested for distinguishing such etiologies. Instrument manufacturers claim that if successful, they may be used to generate software flags that help under-resourced laboratories better triage hemogram specimens requiring further testing. We tested the diagnostic accuracy of volume-conductivity-scatter (VCS) indices together with complete blood count (CBC) parameters in such scenarios. We compared LH780-derived (Beckman Coulter, FL, USA) CBC + VCS parameters from patients with clonal lymphoproliferations (n = 42, including 30 chronic lymphocytic leukemia cases) versus 83 controls with absolute or relative lymphocytosis (derivation cohort). Diagnostic performances of 11 logistic regression equations derived were subsequently evaluated on two specific validation cohorts (n = 130 and n = 1465). Clonal lymphocytoses showed significantly lower hemoglobin and higher leukocyte counts but similar lymphocyte percentages (LY %) vis-à-vis controls. The most significant, albeit overlapping predictor of clonality was the absolute lymphocyte count, LY# (47.8 ± 48.4 × 109/L vs. 2.9 ± 1.4 × 109/L in clonal vs. benign cases). In eleven logistic regression equations constructed using four combinatorial approaches, only the models with LY# (highest sensitivity/specificity of 99.3%/100%) and the lymphocytic VCS parameters alone (highest sensitivity/specificity of 76.2%/90.2%) performed consistently in both validation cohorts. Lymphocytic VCS parameters were moderately successful in distinguishing benign-versus-malignant lymphocytes. Other approaches of CBC-plus-VCS parameters did not sustain their initial excellent performances in the validation cohorts, highlighting a need for careful appraisal and better standardization of automated cellular analysis technologies.

Epigenetic regulation of inflammatory gene expression in macrophages by selenium.

Acetylation of histone and non-histone proteins by histone acetyltransferases plays a pivotal role in the expression of proinflammatory genes. Given the importance of dietary selenium in mitigating inflammation, we hypothesized that selenium supplementation may regulate inflammatory gene expression at the epigenetic level. The effect of selenium towards histone acetylation was examined in both in vitro and in vivo models of inflammation by chromatin immunoprecipitation assays and immunoblotting. Our results indicated that selenium supplementation, as selenite, decreased acetylation of histone H4 at K12 and K16 in COX-2 and TNFα promoters, and of the p65 subunit of the redox sensitive transcription factor NFκB in primary and immortalized macrophages. On the other hand, selenomethionine had a much weaker effect. Selenite treatment of HIV-1-infected human monocytes also significantly decreased the acetylation of H4 at K12 and K16 on the HIV-1 promoter, supporting the down-regulation of proviral expression by selenium. A similar decrease in histone acetylation was also seen in the colonic extracts of mice treated with dextran sodium sulfate that correlated well with the levels of selenium in the diet. Bone-marrow-derived macrophages from Trsp(fl/fl)Cre(LysM) mice that lack expression of selenoproteins in macrophages confirmed the important role of selenoproteins in the inhibition of histone H4 acetylation. Our studies suggest that the ability of selenoproteins to skew the metabolism of arachidonic acid contributes, in part, to their ability to inhibit histone acetylation. In summary, our studies suggest a new role for selenoproteins in the epigenetic modulation of proinflammatory genes.

Crucial role of macrophage selenoproteins in experimental colitis.

Inflammation is a hallmark of inflammatory bowel disease (IBD) that involves macrophages. Given the inverse link between selenium (Se) status and IBD-induced inflammation, our objective was to demonstrate that selenoproteins in macrophages were essential to suppress proinflammatory mediators, in part, by the modulation of arachidonic acid metabolism. Acute colitis was induced using 4% dextran sodium sulfate in wild-type mice maintained on Se-deficient (<0.01 ppm Se), Se-adequate (0.08 ppm; sodium selenite), and two supraphysiological levels in the form of Se-supplemented (0.4 ppm; sodium selenite) and high Se (1.0 ppm; sodium selenite) diets. Selenocysteinyl transfer RNA knockout mice (Trsp(fl/fl)LysM(Cre)) were used to examine the role of selenoproteins in macrophages on disease progression and severity using histopathological evaluation, expression of proinflammatory and anti-inflammatory genes, and modulation of PG metabolites in urine and plasma. Whereas Se-deficient and Se-adequate mice showed increased colitis and exhibited poor survival, Se supplementation at 0.4 and 1.0 ppm increased survival of mice and decreased colitis-associated inflammation with an upregulation of expression of proinflammatory and anti-inflammatory genes. Metabolomic profiling of urine suggested increased oxidation of PGE2 at supraphysiological levels of Se that also correlated well with Se-dependent upregulation of 15-hydroxy-PG dehydrogenase (15-PGDH) in macrophages. Pharmacological inhibition of 15-PGDH, lack of selenoprotein expression in macrophages, and depletion of infiltrating macrophages indicated that macrophage-specific selenoproteins and upregulation of 15-PGDH expression were key for Se-dependent anti-inflammatory and proresolving effects. Selenoproteins in macrophages protect mice from dextran sodium sulfate-colitis by enhancing 15-PGDH-dependent oxidation of PGE2 to alleviate inflammation, suggesting a therapeutic role for Se in IBD.

Selenium suppresses leukemia through the action of endogenous eicosanoids.

Eradicating cancer stem-like cells (CSC) may be essential to fully eradicate cancer. Metabolic changes in CSC could hold a key to their targeting. Here, we report that the dietary micronutrient selenium can trigger apoptosis of CSC derived from chronic or acute myelogenous leukemias when administered at supraphysiologic but nontoxic doses. In leukemia CSC, selenium treatment activated ATM-p53-dependent apoptosis accompanied by increased intracellular levels of reactive oxygen species. Importantly, the same treatment did not trigger apoptosis in hematopoietic stem cells. Serial transplantation studies with BCR-ABL-expressing CSC revealed that the selenium status in mice was a key determinant of CSC survival. Selenium action relied upon the endogenous production of the cyclooxygenase-derived prostaglandins Δ(12)-PGJ2 and 15d-PGJ2. Accordingly, nonsteroidal anti-inflammatory drugs and NADPH oxidase inhibitors abrogated the ability of selenium to trigger apoptosis in leukemia CSC. Our results reveal how selenium-dependent modulation of arachidonic acid metabolism can be directed to trigger apoptosis of primary human and murine CSC in leukemia.

Evaluation of the stability, bioavailability, and hypersensitivity of the omega-3 derived anti-leukemic prostaglandin: Δ(12)-prostaglandin J3.

Previous studies have demonstrated the ability of an eicosapentaenoic acid (EPA)-derived endogenous cyclopentenone prostaglandin (CyPG) metabolite, Δ(12)-PGJ3, to selectively target leukemic stem cells, but not the normal hematopoietic stems cells, in in vitro and in vivo models of chronic myelogenous leukemia (CML). Here we evaluated the stability, bioavailability, and hypersensitivity of Δ(12)-PGJ3. The stability of Δ(12)-PGJ3 was evaluated under simulated conditions using artificial gastric and intestinal juice. The bioavailability of Δ(12)-PGJ3 in systemic circulation was demonstrated upon intraperitoneal injection into mice by LC-MS/MS. Δ(12)-PGJ3 being a downstream metabolite of PGD3 was tested in vitro using primary mouse bone marrow-derived mast cells (BMMCs) and in vivo mouse models for airway hypersensitivity. ZK118182, a synthetic PG analog with potent PGD2 receptor (DP)-agonist activity and a drug candidate in current clinical trials, was used for toxicological comparison. Δ(12)-PGJ3 was relatively more stable in simulated gastric juice than in simulated intestinal juice that followed first-order kinetics of degradation. Intraperitoneal injection into mice revealed that Δ(12)-PGJ3 was bioavailable and well absorbed into systemic circulation with a Cmax of 263 µg/L at 12 h. Treatment of BMMCs with ZK118182 for 12 h resulted in increased production of histamine, while Δ(12)-PGJ3 did not induce degranulation in BMMCs nor increase histamine. In addition, in vivo testing for hypersensitivity in mice showed that ZK118182 induces higher airways hyperresponsiveness when compared Δ(12)-PGJ3 and/or PBS control. Based on the stability studies, our data indicates that intraperitoneal route of administration of Δ(12)-PGJ3 was favorable than oral administration to achieve effective pharmacological levels in the plasma against leukemia. Δ(12)-PGJ3 failed to increase histamine and IL-4 in BMMCs, which is in agreement with reduced airway hyperresponsiveness in mice. In summary, our studies suggest Δ(12)-PGJ3 to be a promising bioactive metabolite for further evaluation as a potential drug candidate for treating CML.

Δ12-prostaglandin J3, an omega-3 fatty acid-derived metabolite, selectively ablates leukemia stem cells in mice.

Targeting cancer stem cells is of paramount importance in successfully preventing cancer relapse. Recently, in silico screening of public gene-expression datasets identified cyclooxygenase-derived cyclopentenone prostaglandins (CyPGs) as likely agents to target malignant stem cells. We show here that Δ(12)-PGJ(3), a novel and naturally produced CyPG from the dietary fish-oil ω-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA; 20:5) alleviates the development of leukemia in 2 well-studied murine models of leukemia. IP administration of Δ(12)-PGJ(3) to mice infected with Friend erythroleukemia virus or those expressing the chronic myelogenous leukemia oncoprotein BCR-ABL in the hematopoietic stem cell pool completely restored normal hematologic parameters, splenic histology, and enhanced survival. More importantly, Δ(12)-PGJ(3) selectively targeted leukemia stem cells (LSCs) for apoptosis in the spleen and BM. This treatment completely eradicated LSCs in vivo, as demonstrated by the inability of donor cells from treated mice to cause leukemia in secondary transplantations. Given the potency of ω-3 polyunsaturated fatty acid-derived CyPGs and the well-known refractoriness of LSCs to currently used clinical agents, Δ(12)-PGJ(3) may represent a new chemotherapeutic for leukemia that targets LSCs.

Selenoprotein-dependent up-regulation of hematopoietic prostaglandin D2 synthase in macrophages is mediated through the activation of peroxisome proliferator-activated receptor (PPAR) gamma.

The plasticity of macrophages is evident from their dual role in inflammation and resolution of inflammation that are accompanied by changes in the transcriptome and metabolome. Along these lines, we have previously demonstrated that the micronutrient selenium increases macrophage production of arachidonic acid (AA)-derived anti-inflammatory 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)) and decreases the proinflammatory PGE(2). Here, we hypothesized that selenium modulated the metabolism of AA by a differential regulation of various prostaglandin (PG) synthases favoring the production of PGD(2) metabolites, Δ(12)-PGJ(2) and 15d-PGJ(2). A dose-dependent increase in the expression of hematopoietic-PGD(2) synthase (H-PGDS) by selenium and a corresponding increase in Δ(12)-PGJ(2) and 15d-PGJ(2) in RAW264.7 macrophages and primary bone marrow-derived macrophages was observed. Studies with organic non-bioavailable forms of selenium and the genetic manipulation of cellular selenium incorporation machinery indicated that selenoproteins were necessary for H-PGDS expression and 15d-PGJ(2) production. Treatment of selenium-deficient macrophages with rosiglitazone, a peroxisome proliferator-activated receptor γ ligand, up-regulated H-PGDS. Furthermore, electrophoretic mobility shift assays indicated the presence of an active peroxisome proliferator-activated receptor-response element in murine Hpgds promoter suggesting a positive feedback mechanism of H-PGDS expression. Alternatively, the expression of nuclear factor-κB-dependent thromboxane synthase and microsomal PGE(2) synthase was down-regulated by selenium. Using a Friend virus infection model of murine leukemia, the onset of leukemia was observed only in selenium-deficient and indomethacin-treated selenium-supplemented mice but not in the selenium-supplemented group or those treated with 15d-PGJ(2). These results suggest the importance of selenium in the shunting of AA metabolism toward the production of PGD(2) metabolites, which may have clinical implications.

The regulation of erythropoiesis by selenium in mice.

Redox modulation by antioxidants, such as selenium (Se), has emerged as an important regulator of erythropoiesis. Using Se-deficient (0.04 ppm), Se-adequate (0.1 ppm), and Se-supplemented (0.4 ppm) C57/BL6 mice, we show that Se deficiency caused anemia, when compared to the Se-supplemented and Se-adequate groups. Increased denaturation of hemoglobin, methemoglobin, protein carbonyls, lipid peroxidation, Heinz bodies, and osmotic fragility of erythrocytes were observed in Se-deficient mice. Increased oxidative stress upregulated forkhead transcription factor (FoxO3a) and hypoxia-inducible factor-(HIF)1α in the spleen and kidney of Se-deficient murine as well as in the proerythroblast G1E cells cultured in Se-deficient media. A significant increase in the expression of erythropoietin, a downstream target of HIF1α, and expansion of stress erythroid progenitors (burst forming units-erythroid) were seen in the Se-deficient mice. Despite the increase in erythroid progenitors, lowered reticulocytes suggest a defective erythroid differentiation pathway. While Se deficiency led to increased nuclear levels of active FoxO3a, Se-adequate conditions reversed this effect and increased nuclear export by its binding partner, 14-3-3ßζ, that is under the redox control of selenoproteins. In summary, these results provide insight into the importance of adequate Se nutrition in regulating red cell homeostasis by mitigating oxidative stress-dependent modulation of FoxO3a and HIF1α to effect differentiation of erythroid progenitors.

Inhibition of TLR4-induced IκB kinase activity by the RON receptor tyrosine kinase and its ligand, macrophage-stimulating protein.

The RON receptor tyrosine kinase regulates the balance between classical (M1) and alternative (M2) macrophage activation. In primary macrophages, the ligand for Ron, macrophage-stimulating protein (MSP), inhibits the expression of inducible NO synthase, a marker of classically activated macrophages, whereas promoting the expression of arginase I, a marker of alternative activation. Ron(-/-) mice express increased levels of IL-12, a product of classically activated macrophages, after endotoxin administration, resulting in increased serum IFN-γ levels and enhanced susceptibility to septic shock. In this study, we demonstrate that MSP inhibits LPS-induced IL-12p40 expression, and this inhibition is dependent on the docking site tyrosines in Ron. To further define this inhibition, we examined the effect of Ron on signaling pathways downstream of Ron. We found that MSP does not inhibit the MyD88-independent activation of IFN regulatory factor 3 and production of IFN-ß in response to LPS, nor does it inhibit MyD88-dependent TGF-ß-activated kinase phosphorylation or MAPK activation in primary macrophages. However, the induction of IκB kinase activity, IκB degradation, and DNA binding of NF-κB after LPS stimulation is delayed in the presence of MSP. In addition, Ron inhibits serine phosphorylation of p65 and NF-κB transcriptional activity induced by LPS stimulation of TLR4. Finally, MSP inhibits the NF-κB-dependent upregulation of the nuclear IκB family member, IκBζ, a positive regulator of secondary response genes including IL-12p40. LPS also induces expression of Ron and an N-terminally truncated form of Ron, Sf-Ron, in primary macrophages, suggesting that the upregulation of Ron by LPS could provide classical feedback regulation of TLR signaling.

Synthesis and evaluation of the anti-inflammatory properties of selenium-derivatives of celecoxib.

Celecoxib is a selective cyclooxygenase (COX)-2 inhibitor used to treat inflammation, while selenium is known to down-regulate the transcription of COX-2 and other pro-inflammatory genes. To expand the anti-inflammatory property, wherein celecoxib could inhibit pro-inflammatory gene expression at extremely low doses, we incorporated selenium (Se) into two Se-derivatives of celecoxib, namely; selenocoxib-2 and selenocoxib-3. In vitro kinetic assays of the inhibition of purified human COX-2 activity by these compounds indicated that celecoxib and selenocoxib-3 had identical K(I) values of 2.3 and 2.4µM; while selenocoxib-2 had a lower K(I) of 0.72µM. Furthermore, selenocoxib-2 inhibited lipopolysaccharide-induced activation of NF-κB leading to the down-regulation of expression of COX-2, iNOS, and TNFα more effectively than selenocoxib-3 and celecoxib in RAW264.7 macrophages and murine bone marrow-derived macrophages. Studies with rat liver microsomes followed by UPLC-MS-MS analysis indicated the formation of selenenylsulfide conjugates of selenocoxib-2 with N-acetylcysteine. Selenocoxib-2 was found to release minor amounts of Se that was effectively inhibited by the CYP inhibitor, sulphaphenazole. While these studies suggest that selenocoxib-2, but not celecoxib and selenocoxib-3, targets upstream events in the NF-κB signaling axis, the ability to effectively suppress NF-κB activation independent of cellular selenoprotein synthesis opens possibilities for a new generation of COX-2 inhibitors with significant and broader anti-inflammatory potential.

Diminished reproductive potential of male mice in response to selenium-induced oxidative stress: involvement of HSP70, HSP70-2, and MSJ-1.

The oxidative stress imposed by nutritional variations in selenium (Se) has plausible role in reproductive toxicology and affects the reproductive potential. Also, the expression of heat shock proteins (HSPs) is a highly regulated event throughout the process of spermatogenesis and is modulated by stressful stimuli. This prompted us to investigate the possibility that Se-induced oxidative stress may affect the fertility status by altering the expressions of the constitutive and inducible HSP70 proteins, having crucial role in spermatogenesis. Different Se status-deficient, adequate, and excess, male Balb/c mice were created by feeding yeast-based Se-deficient diet (group I) and deficient diet supplemented with Se as sodium selenite at 0.2 and 1 ppm Se (group II and III) for a period of 8 weeks. After completion of the diet-feeding schedule, a significant decrease in the Se and glutathione peroxidase (GSH-Px) levels was observed in the Se-deficient group (I), whereas Se-excess group (III) demonstrated an increase. Increased levels of reactive oxygen species, malondialdehyde, and alterations in the redox status in both groups I and III indicated oxidative-stressed conditions. There was an overall reduced fertility status in mice supplemented with Se-deficient and Se-excess diet. The mRNA and protein expression of HSP70 was found to be elevated in these two groups, whereas the expression patterns of HSP70-2 and MSJ-1 demonstrated a reverse trend. In vitro CDC2 kinase assay showed reduced kinase activity in group I and group III. These findings suggest that Se-induced oxidative stress by differentially regulating various HSP70s can affect its downstream factors having crucially important role in differentiation of germ cells and completion of spermatogenesis. Therefore, it can provide an insight into the mechanism(s) by which the oxidative stress-induced reproductive toxicity can lead to increased apoptosis/growth arrest and infertility. This will thus add new dimensions to the molecular mechanism underlying the human male infertility and open new vistas in the development of various chemo-preventive methods.

Selenium variation induced oxidative stress regulates p53 dependent germ cell apoptosis: plausible involvement of HSP70-2.

BACKGROUND: Selenium at altered concentration causes abnormal spermatogenesis and male infertility. However, the exact mechanism behind this is still unexplored. AIMS: It was aimed to investigate if Se induced oxidative stress alters the expressions of testis specific HSP70-2 protein, that is crucial in normal spermatogenesis. The study was extended to delineate the apoptotic process after this change if any. METHODS: To create different Se status-deficient, adequate and excess, male Balb/c mice were fed yeast based Se deficient diet (group I) and this diet supplemented with Se as sodium selenite at 0.2 and 1 ppm Se (group II and III, respectively) for 8 weeks. RESULTS: After the feeding schedule, a dose dependent change in the Se levels and GSH-Px activity was observed in samples of different Se diet fed group animals as reported in earlier studies. Changes in the redox status in both groups I and III indicated oxidative stress conditions. The mRNA and protein expression of HSP70-2 was found to be reduced in group I and III, whereas, the expressions of p53 demonstrated a reverse trend. Increased apoptosis was observed in the group I and III animals as indicated by increased apoptotic index (AI) on the TUNEL stained sections and by DNA fragmentation indicating DNA damage in these groups. CONCLUSION: These findings suggest that Se variations induced oxidative stress leads to germ cell apoptosis and downregulation of HSP70-2. This study suggests that there can be a possible link between these two events and the fate of HSP70-2 in case of oxidative damage can provide an insight into the mechanism(s) by which the nutritional variation induced oxidative stress can affect reproductive potential and thus demonstrates the importance of nutrition at molecular level as well.