Zeliha Buyukbingol

May nitric oxide molecule have a role in the pathogenesis of human cataract

Recent studies have shown that nitric oxide molecule may have a role in the development of cataract. In this study, we measured the levels of a nitric oxide metabolite (nitrite) in the cataractous and normal human lenses. A modified Griess assay was used to determine the nitrite levels in the lenses as a measure of nitric oxide, based on the spectrophotometric method. Nitrite was detected in 26 (44.1%) cataractous lenses and was found below the detection limit in 33 (55.9%) cataractous lenses. Mean nitrite levels in cataractous lenses (2.77+/-5.26nmol/100mg) was found higher than the normal lenses (0.77+/-0.79nmol/100mg) but this increase was not statistically significant. Comparison of nitrite levels among lenses with various types of cataracts revealed higher levels in lenses with posterior subcapsular cataracts. Hypertensive patients had also significantly higher nitrite levels in their cataractous lenses. The increased levels in the cataractous lenses display a possible role of nitric oxide in the pathogenesis of cataract in human eyes.

Methylsulfonylmethane modulates apoptosis of LPS/IFN-γ-activated RAW 264.7 macrophage-like cells by targeting p53, Bax, Bcl-2, cytochrome c and PARP proteins

Abstract Methylsulfonylmethane (MSM) is a non-toxic, natural organosulfur compound, which is known to possess antioxidant and anti-inflammatory activities. In recent years, MSM has been widely used as a dietary supplement for its beneficial effects against various diseases, especially arthritis. Despite being a popular supplement product, the mechanism of action of MSM is not well known. This study was designed to investigate the effects of MSM on cytotoxic signals induced by lipopolysaccharide (LPS) and interferon-gamma (IFN-γ) in RAW 264.7 macrophage-like cells. The results showed that MSM reversed apoptosis of RAW 264.7 macrophage-like cells at non-cytotoxic concentrations probably through the modulation of apoptotic proteins. After pre-treatment of cells with non-toxic doses of MSM; caspase-3 activation, p53 accumulation, cytochrome c release and Bax/Bcl-2 ratio were significantly decreased and full length poly ADP-ribose polymerase (PARP) was significantly increased. In addition, the loss of mitochondrial membrane potential was decreased with MSM pretreatment in activated macrophages. Since excess nitric oxide production causes apoptosis of macrophages, anti-apoptotic effects of MSM are thought to be mediated by its inhibitor effects on inducible nitric oxide synthase (iNOS) protein and nitric oxide levels. More interestingly, higher doses of MSM exhibited biphasic effects, inhibited cell viability, induced apoptosis of macrophages, increased caspase-3 activity and PARP cleavage. Thus, our results reveal the molecular mechanism of of MSM indicating that MSM supplementation may be beneficial for complications related to nitric oxide-dependent apoptosis in inflammatory conditions. However, the optimum concentration of MSM must be chosen carefully to elicit the desired effect.

Effects of supplementation with a combination of cobalt and ascorbic acid on antioxidant enzymes and lipid peroxidation levels in streptozocin-diabetic rat liver.

The effect of cobalt(II) chloride (CoCl2) and CoCl2 with ascorbic acid (AA) on components of the antioxidant defense system and lipid oxidative damage were studied in controls and streptozotocin-induced diabetic rat livers. Three days after injection, rats received either 0.5 mM CoCl2 or 0.5 mM CoCl2 with a combination of 1 g/L AA in drinking water up to 6 wk. The elevated blood glucose levels in diabetic rats were about 12% restored by oral administration of CoCl2 (0.05 mM) and were significant reduced (46%) following AA addition (1 g/L) to CoCl2. Cobalt therapy effectively decreased the increased activities of catalase (CAT), superoxide dismutase (SOD), and thiobarbituric acid reactant substances (TBARS) but could not restore the increased glutathione peroxidase (GSH-Px) in the liver of diabetic rats. Our findings suggest that cobalt therapy may prove effective in improving the impaired antioxidant status during the early state of diabetes, and ascorbic acid supplementation at this dose potentiates the effectiveness of cobalt action.

Methylsulfonylmethane Induces p53 Independent Apoptosis in HCT-116 Colon Cancer Cells

Methylsulfonylmethane (MSM) is an organic sulfur-containing compound which has been used as a dietary supplement for osteoarthritis. MSM has been shown to reduce oxidative stress and inflammation, as well as exhibit apoptotic or anti-apoptotic effects depending on the cell type or activating stimuli. However, there are still a lot of unknowns about the mechanisms of actions of MSM. In this study, MSM was tested on colon cancer cells. 3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay and flow cytometric analysis revealed that MSM inhibited cell viability and increased apoptotic markers in both HCT-116 p53 +/+ and HCT-116 p53 −/− colon cancer cells. Increased poly (ADP-ribose) polymerase (PARP) fragmentation and caspase-3 activity by MSM also supported these findings. MSM also modulated the expression of various apoptosis-related genes and proteins. Moreover, MSM was found to increase c-Jun N-terminal kinases (JNK) phosphorylation in both cell lines, dose-dependently. In conclusion, our results show for the first time that MSM induces apoptosis in HCT-116 colon cancer cells regardless of their p53 status. Since p53 is defective in >50% of tumors, the ability of MSM to induce apoptosis independently of p53 may offer an advantage in anti-tumor therapy. Moreover, the remarkable effect of MSM on Bim, an apoptotic protein, also suggests its potential use as a novel chemotherapeutic agent for Bim-targeted anti-cancer therapies.

Synthesis and effects of some novel tetrahydronaphthalene derivatives on proliferation and nitric oxide production in lipopolysaccharide activated Raw 264.7 macrophages.

In this study, novel N-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydronaphthalene-2-yl)-carboxamide (6-15) and 5,5,8,8-tetramethyl-5,6,7,8-tetrahydronaphthalene-2-carboxamide (16-32) derivatives were synthesized and their in vitro effects at 5 μM and 50 μM concentrations on proliferation and nitric oxide (NO*) production in lipopolysaccharide (LPS) activated RAW 264.7 macrophage cells were determined. Compounds 12, 17, 24 and 26 were found to decrease nitrite levels in a dose-dependent manner in LPS-activated cells. At the tested concentrations, these compounds did not exhibit cytotoxic effects. Interestingly, compound 27 which contains nitroxide free radical was the most active compound in this series showing 59.2% nitrite inhibition in LPS-activated macrophage cells.

Apoptotic Effects of Some Tetrahydronaphthalene Derivatives on K562 Human Chronic Myelogenous Leukemia Cell Line

BACKGROUND Retinoids which are vitamin A (Retinol) derivatives have been suggested to mediate the inhibition of cancer cell growth and apoptosis. It has been reported that all trans retinoic acid (ATRA) exhibited suppressive effects on different types of leukemia including chronic myelogenous leukemia. OBJECTIVE In the present study, we aim to find out the effects of 6 synthetic N-(3,5,5,8,8-pentamethyl-5,6,7,8- tetrahydronaphthalene-2-yl)-carboxamide derivatives (compound 6-12) on cell viability and apoptotic pathways in K562 human chronic myelogenous leukemia cell line. METHODS Cell viability and apoptosis were examined by spectrophotometric thiazolyl blue tetrazolium bromide (MTT) and caspase-3 assay, western blot, RT-PCR and flow cytometry. RESULTS Our results indicated that compound 6 (5-(1,2-Dithiolan-3-yl)-N-(3,5,5,8,8-pentamethyl-5,6,7,8- tetrahydronaphthalen-2-yl)pentanamide), 8 (4-(3,4-Dimethoxyphenyl)-N-(3,5,5,8,8-pentamethyl-5,6,7,8- tetrahydronaphthalen-2-yl)butanamide) and 11 (E-3-(4-Hydroxy-3-methoxyphenyl)-N-(3,5,5,8,8-pentamethyl- 5,6,7,8-tetrahydronaphthalen-2-yl)acrylamide) exhibited apoptotic effects in K562 human chronic myelogenous leukemia cell line and induced caspase 3, PARP cleavage, Bax/Bcl-2 ratio, Bad and Bim gene expressions. CONCLUSION Some retinoid derivatives tested in this study induced apoptosis of K562 cells which suggest that these compounds may serve as potential agents in the treatment of chronic myelogenous leukemia.

Expression analysis of Akirin-2, NFκB-p65 and β-catenin proteins in imatinib resistance of chronic myeloid leukemia

ABSTRACT Objective: Chronic myleoid leukemia (CML) is a myeloproliferative disorder characterized with the constitutive activation of Bcr-Abl tyrosine kinase which is a target for imatinib, the first line treatment option for CML. Constitutive activation of NFκB and β-catenin signaling promotes cellular proliferation and survival and resistance to Imatinib therapy in CML. Akirin-2 is a nuclear protein which is required for NFκB dependent gene expression as a cofactor and has been linked to Wnt/beta-catenin pathway. The purpose of this study is to examine Akirin-2, NFκB and β-catenin in imatinib resistance of CML and to test if any direct physical protein–protein interaction exists between NFkB and both β-catenin and Akirin-2. Methods: RT–PCR and western blot were performed to determine Akirin-2, NFκB-p65 and β-catenin gene and protein expressions, Co-immunoprecipitation and chromatin immunoprecipitation analysis were carried out to detect the direct physical interactions and binding of NFκB-p65 and β-catenin proteins to MDR1 promoter region, respectively. Results: β-catenin and NFκB-p65 proteins bound to DNA promoter regions of MDR1 in imatinib-sensitive and resistant CML cells, whereas any direct protein–protein interaction could not be found between NFκB-p65 and Akirin-2 or β-catenin proteins. Nuclear β-catenin and NFκB-p65 levels increased in imatinib resistance. Moreover, increased Akirin-2 protein accumulation in the nucleus was shown for the first time in imatinib resistant CML cells. Discussion: We show for the first time that Akirin-2 can be a novel biomarker in imatinib resistance. Targeting Akirin-2, NFκB and β-catenin genes may provide an opportunity to overcome imatinib resistance in CML.

Effects of glucosamine on LPS/IFN-γ induced RAW 264.7 macrophage apoptosis

Amac: Apoptoz, doku homeostazisinin duzenlenmesi gibi normal fizyolojide ve ceșitli hastaliklarin patofizyolojisinde onemli rol oynayan genetik olarak programlanmiș bir hucre olum mekanizmasidir. Ceșitli hucre tiplerinde mikrobiyal ve viral patojenlere karși bir savunma mekanizmasi olarak ortaya cikan iNOS enzim aktivasyonu ve nitrik oksit sentezi, ateroskleroz, romatoid artrit, diyabet, septik șok, multipl sklerozis gibi inflamatuvar ve immun patolojilerde onemli rol oynamakta, kontrolsuz yuksek duzeydeki nitrik oksit uretimi hucre olumune yol acmaktadir. Yakin zamanda, osteoartrit gibi inflamatuvar bozukluklar icin yararli oldugu iddia edilen glukozamin gunumuzde klinik olarak kullanilmaktadir. Glukozaminin, bașlica immun sistem hucreleri olan makrofajlarin apoptozu uzerine olan etki mekanizmasi hakkinda sinirli bilgi bulunmaktadir. Bu calișmada, farkli konsantrasyonlardaki glukozaminin, LPS/IFNγ ile aktive edilmiș RAW 264.7 makrofajlar uzerindeki etkileri belirlenmiștir. Materyal ve metod: RAW 264.7 makrofaj hucre serisi, LPS/IFNγ ile uyarim oncesinde glukozamin varliginda ve yoklugunda kultur edildi. Nitrit duzeyleri, hucre canliligi, kaspaz-3 aktivitesi, mitokondri membran potansiyeli tayini ve Annexin V-PI hucre boyamasi ile akim sitometrik analizler gercekleștirilmiștir. Sonuclar ve tartișma: Glukozaminin, nitrit duzeylerini inhibe ettigi, mitokondri membran potansiyelini arttirdigi, kaspaz-3 enzim aktivitesini azalttigi ve anlamli olarak antiapoptotik etkiler sergiledigi (p