The synergistic effect of geldanamycin and trıchostatin A on human bladder cells

AMAÇ: Isı şok protein 90 (IŞP), ATP-bağımlı moleküler bir şaperon olup kanserin temel özellikleri olarak tanımlanan çok sayıdaki onkogenik sinyal proteinlerinin stabilitesi ve fonksiyonu için gereklidir. Bu nedenle, IŞP90 kanserin önlenmesi ve tedavisi için moleküler bir terapötik hedef olarak görülür. Geldanamisin (GA) ilk doğal IŞP90 inhibitörüdür. IŞP90 inhibisyonu, kanser hücrelerinde apoptosisin indüklenmesine neden olur ve kemoterapi direncinde bir azalmaya eşlik edebilir. Bu çalışmada amacımız, Geldanamisin (GA) ve Trikostatin A (TSA)’ın tek başlarına ve/veya kombinasyonlarının kullanımının, insan mesane kanser hücre hattı T24’de transkripsiyonel ve protein düzeyinde apoptotik yolak üzerindeki sinerjik etkilerini araştırmaktır. GEREÇ VE YÖNTEM: Geldanamisin (0–30 µM)'nin hücre canlılığı üzerine olan etkisi WST1 aracılığıyla belirlenmiştir. Belirtilen ilaçların tek başlarına ve birlikte kullanımlarının CASP3 geninin transkripsiyonel ifade düzeylerindeki farklılıklar kantitatif eş zamanlı PCR yöntemiyle belirlenmiştir. Kaspaz3, Bax ve Bcl2 genlerinin translasyonel ifade düzeyleri ise western blot yöntemi ile gerçekleştirilmiştir. BULGULAR: Antiapoptotik gen Bcl2 ifadelenmesi, GA ve GA+TSA kombinasyonu kullanımı sonrasında önemli derecede azalmakta iken, kaspaz-3 ve Bax ifadelenmesi kontrole göre translasyonel düzeyde artmıştır. Aynı zamanda, Cas3 mRNA düzeyi de kontrole göre artmıştır(p<0.05). SONUÇ: GA+TSA kombinasyonu hücre proliferasyonunu azaltmakta ve apoptoza indüklediği görülmüştür. Bu nedenle, IŞP90 inhibitörlerinin mesane kanseri tedavisinde yeni bir yaklaşım sunabileceğini düşünmekteyiz.OBJECTIVE: Heat shock protein 90 (HSP90) is an ATPdependent molecular chaperone required for the stability and function of numerous oncogenic signaling proteins that determine the hallmarks of cancer. Therefore, HSP90 is known as a molecular therapeutic target for the prevention and treatment of cancer. Geldanamycin (GA) was the first identified natural Hsp90 inhibitor. Inhibition of Hsp90 results in the induction of apoptosis in cancer cells, and may be accompanied by a reduction in chemotherapy resistance. This study is aimed to investigate, on transcriptional and protein levels, the synergic effects of unaccompanied and/or combined use of Geldanamycin (GA) and Trichostatin A (TSA) on the apoptotic pathway of human bladder cancer cell line T24. MATERIAL AND METHODS: The effect of geldanamisin (0–30 µM) on cell viability were determined by WST-1. The variation in the expression levels of CASP3 genes was transcriptionally determined by quantitative real-time PCR. The translational expressional levels of caspase-3, Bax and Bcl2 genes were performed by western blot method. RESULTS: The use of GA alone and GA+TSA combinati- on significantly down-regulated the antiapoptotic gene Bcl2 while Caspase-3 and Bax expression were increased at translational levels. Cas3 mRNA level was also increa- sed with respect to control (p<0.05). CONCLUSIONS: The use of geldanamycin+TSA combination reduced cell proliferation and induced apoptosis. Therefore, it can be suggested that HSP90 inhibitors may offer a new approach to consider in the treatment of bladder cancer

The determination of chemotherapeutic effects of oxaliplatin and cisplatin in the prostate cancer cell lines administered gemcitabine

Ürogenital kanserler içerisinde erkeklerde görülme sıklığı açısından ilk sırada yer alan prostat kanseri, kanser ilişkili ölümler arasında ikinci sırada yer almaktadır. Mevcut araştırmalar, spesifik hasta popülasyonlarında ilave avantajlar sağlamak için platin-temelli kemoterapötikler ile hedeflenmiş tedavi kombinasyonlarına odaklanmıştır. Cerrahi veya radrasyonu takiben platin bazlı kombinasyon kemoterapisi kullanımı en fazla yararı sağlamaktadır. Bu çalışmanın amacı, platin temelli kemoterapötikler sisplatin ve oksaliplatin ile Gemsitabinin tek başına ve kombine dozlarının insan prostat kanserli hücre hatları olan DU-145 ve PC3 üzerindeki hücre proliferasyonu ve apoptotik yolaklar üzerine etkilerini belirlenmektedir. Prostat kanserli hücre hatlarında bu ilaçların tek başlarına veya kombine dozlarının, hücrelerin canlılığı üzerine olan etkileri WST-1 yöntemiyle ve CASP3, CASP8 ve CASP9 genlerinin mRNA düzeylerinde ifadelenme değişiklikleri RT-PCR ile belirlendi. Sonuç olarak, içsel ve dışsal yolakta görev alan CASP8 ve CASP9 genlerinin mRNA düzeyleri değerlendirildiğinde sisplatinin içsel yolaktan (CASP9) ve oxaliplatinin ise dışsal yolaktan (CASP8) apoptozu indüklediği tespit edildi. Bu göstermektedir ki; platin temelli kemoterapötik olan oxaliplatin ve sisplatin hedef yolakta farklı proteinler üzerinden etki etmektedir. Oxaliplatinin tek başına veya gemsitabin ile kombinasyonlarının apoptotik yolakta sisplatinden daha etkin olduğunu belirlendi.While prostate cancer takes first place in terms of prevalence among genitourinary cancers in men, it is second among cancer related deaths. Current researches are focused on combining targeted therapy with platinum-based chemotherapy for achieve additional avantages for spesific patient populations. Use of platin-based combination chemotherapy followed by surgical resection and/or radiation ensures the most benefit terapy. The aim of this study was to compare the effectiveness of platinum-based chemotherapeutics Cisplatin and Oxaliplatin used single or combination doses with Gemsitabine on apoptosis and cell proliferation of DU145 and PC3 human prostate cancer cell lines. The effects of drugs alone or combinations on cell viability were determined by WST-1 method. The mRNA expression levels of Caspase-3, Caspase-8, Caspase-9 genes were analyzed by real time polymerase chain reaction (RT-PCR). As a result, when evaluated the mRNA levels of Caspase-8 and Caspase-9 genes that play role intrinsic and extrinsic pathway, it was faund that Cisplatin induced apoptosis by intrinsic pathway, and Cisplatin induced apoptosis by extrinsic pathway. It is determined that, only Oxaliplatin or along with Gemsitabin combinations are more effective on apoptotic pathway than Sisplatin

Protective effect of taurine against acrylamide-induced oxidative stress in rats

Bu çalışmada, sıçanlarda akrilamid ile oluşturulan oksidatif strese karşı antioksidan ajan olan taurinin koruyucu etkisinin araştırılması amaçlandı. Toplam 35 Wistar erkek sıçan standart rodent yemiyle beslendi ve 5 eşit gruba ayrıldı. I. Grup: kontrol grubu, II. Grup: sadece 15 mg/kg akrilamid 60 gün boyunca gastrik gavaj yolu ile verildi. III., IV. ve V. gruplara sırasıyla akrilamid (15 mg/kg) ve taurin 50, 100 ve 200 mg/kg/gün olacak şekilde ağızdan gastrik gavaj ile verildi. Akrilamid uygulaması sıçanların kan ve dokularında malondialdehid seviyelerinde önemli bir artışa ve glutatyon seviyelerinde azalmaya neden oldu. Buna ilave olarak, akrilamid uygulaması ile süperoksid dismutaz ve katalaz aktivitelerinin sıçan eritrosit ve dokularında azaldığı belirlendi. Taurin uygulamasının akrilamid ile indüklenen oksidatif stres ve lipid peroksidasyonu azaltarak vücudun antioksidan savunma sistemini iyileştirdiği belirlendi. Bunun yanı sıra, taurin uygulaması ile akrilamid verilen sıçanların karaciğer ve beyin dokularındaki IFN-γ, IL-1β, TNF-α ve NFĸB mRNA ekspresyon düzeylerinin azaldığı gözlendi. Ayrıca, taurin dokularda akrilamid ile indüklenen oksidatif stres sonucu oluşan histopatolojik değişikliklere karşı koruyucu etki gösterdi. Sonuçta doza bağlı olarak taurinin sıçanlarda antioksidan savunma mekanizmasını artırarak akrilamidle indüklenen oksidatif strese karşı koruyucu etki gösterdiği belirlendi.The aim of the present study was to evaluate the protective effect of taurine on acrylamide-induced oxidative stress in rats. Totally, 35 Wistar albino male rats were fed standard rodent diet and divided into 5 equal groups. I. Group: control group, II. Group: only 15 mg/kg acrylamide treated by gastric gavage for 60 days. III., IV., and V. group received 15 mg/kg acrylamide with taurine at dose of 50, 100 and 200 mg/kg/day by gastric gavage. Treatment with acrylamide significantly increased malondialdehyde levels and decreased glutathione levels in blood and tissues of rats. Additionally, acrylamide treatment decreased superoxide dismutase and catalase activities in erythrocyte and tissues of rats. However, treatment of taurine inhibited acrylamide-induced oxidative stress, lipid peroxidation, and enhanced the antioxidant enzymes activities. Besides, mRNA expression levels of IFN-γ, IL-1β, TNF-α, and NFĸB in liver and brain of acrylamide exposed rats were decreased with taurine treatment. Moreover, taurine exhibited protective action against the acrylamide-induced histopathological changes in tissues. In conclusion, it was determined that taurine in a dose-dependent manner has a protective effect against acrylamide induced oxidative stress by increasing antioxidant defense mechanism in rats

The DNA methylation drift of the atherosclerotic aorta increases with lesion progression

Background: Atherosclerosis severity-independent alterations in DNA methylation, a reversible and highly regulated DNA modification, have been detected in aortic atheromas, thus supporting the hypothesis that epigenetic mechanisms participate in the pathogenesis of atherosclerosis. One yet unaddressed issue is whether the progression of atherosclerosis is associated with an increase in DNA methylation drift in the vascular tissue. The purpose of the study was to identify CpG methylation profiles that vary with the progression of aterosclerosis in the human aorta. Methods: We interrogated a set of donor-matched atherosclerotic and normal aortic samples ranging from histological grade III to VII, with a high-density (>450,000 CpG sites) DNA methylation microarray. Results: We detected a correlation between histological grade and intra-pair differential methylation for 1,985 autosomal CpGs, the vast majority of which drifted towards hypermethylation with lesion progression. The identified CpG loci map to genes that are regulated by known critical transcription factors involved in aterosclerosis and participate in inflammatory and immune responses. Functional relevance was corroborated by crossing the DNA methylation profiles with expression data obtained in the same human aorta sample set, by a transcriptome-wide analysis of murine atherosclerotic aortas and from available public databases. Conclusions: Our work identifies for the first time atherosclerosis progression-specific DNA methylation profiles in the vascular tissue. These findings provide potential novel markers of lesion severity and targets to counteract the progression of the atherom

DNMT1 expression level in urogenital cancers

DNA metilasyonunun en önemli epigenetik mekanizmalardan biri olduğu düşünülmekte ve DNA metiltransferazlar (DNMT) tarafından katalizlenmektedir. Artmış DNMT1 ürogenital sistem tümörlerinde sıklıkla görülmektedir ancak bu artışın nedeni tam olarak bilinmemektedir. Çalışmamızda ürogenital kanserlerinde transkripsiyonel ve protein düzeyde WNT/ß-katenin sinyal yolağının DNMT1 aşırı ifadelenmesi üzerine etkisi ve DNMT1 stabilitesinden sorumlu olan UHRF1 ve HAUSP enzimlerin değişen ifade düzeylerinin belirlenmesi amaçlanmıştır. SB216763'in sitotoksitesi, mesane, renal hücre karsinoma ve prostat kanser hücrelerinde zamana ve doza bağımlı olarak WST1 aracılığıyla, CCND1 (ß-katenin) ve WIF-1 hedef genlerin ifadelenme değişimleri real-time PCR and ß-katenin, DNMT1, pGSK3ß(Ser9), HAUSP ve UHRF1 protein düzeyleri western blot kullanılarak belirlendi. Bulgularımıza göre, SB216763 düşük dozlarda T24, Caki-2, PC3 hücrelerinde hücre proliferasyonunu arttırırken yüksek dozlarda ise hücre proliferasyonunun inhibisyonuna neden olmaktadır. Her iki durumda da CCND1 mRNA(p<0,001) ve protein düzeyinin arttığı belirlendi. Aynı şartlar altında ß-katenin düzeyinde artışına paralel olarak DNMT1 protein düzeyinde artma belirlendi. Ayrıca HAUSP ve UHRF1 ise kanser özgü olarak protein düzeylerinde artma veya azalma gözlemlendi. Bizim sonuçlarımız, Wnt/ß-katenin sinyal yolağının moleküler etkilerinin DNMT1 üzerine etkilerinin tam olarak belirlenmesine kanserlerinin tedavisi için yeni moleküler hedeflerin tanımlanmasına katkı sağlayabilecektir.DNA methylation is considered as the most important epigenetic mechanism and is catalyzed by DNA methyltransferases (DNMT). DNMT1 abundance has been seen frequently in urogenital system tumors but the reasons for these increased are not well understood. We aimed to determinate effect of Wnt/ß-catenin signal pathway on overexpression of DNMT1 and aberrant expression of UHRF1 and HAUSP responsible for stability of DNMT1 at transcriptional and protein levels in urogenital cancer. Cytotoxic effect of SB216763 in bladder cancer, renal cell carcinoma and prostate cancer cells was detected time and dose dependent manner with WST1, expression aberration of target genes CCND1(ß-catenin) and WIF-1 using real-time PCR and protein levels of ß-catenin, DNMT1, pGSK3ß(Ser9), HAUSP and UHRF1 using western blot. Our results indicated that SB216763 caused an increased cell proliferation at low dose in the meantime high dose caused a decreased cell proliferation in cells. In both cases, SB216273 was increased CCND1 mRNA, pGSK3ß(Ser9) and ß-catenin, DNMT1 at protein level. Also, HAUSP and UHRF1 up/down-regulated at the same doses (p<0,01) depending on the types of cancer. Also, we showed that ß-catenin and UHRF1 were decreased after reexpression of WIF-1 following treatment with DAC. Our findings may offer a new approach determination of molecular effect of Wnt/ß-catenin signal pathway on DNMT1 would allow us to identify new molecular targets for treatment of the cancer

Researches on fruit juice production from stanley variety black plums grown in İnegöl

Marmara Bölgesi için elverişli çeşitler içinde yer alan Stanley çeşidi erik,sofralık ve kurutmalık olarak değerlendirilmektedir. Ancak özellikle son yıllarda İnegöl yöresinde, Stanley çeşidi erik üretiminde büyük bir artma meydana gelerek, yöre halkı ürünü değerlendirmede bir dar boğaza girmiş ve çözüm için resmi kurum ve kuruluşlara başvuruda bulunmuşlardı Bu gerçekten hareketle üreticilere, yöre halkının kurdukları kooperatife ve sanayi kuruluşlarına Stanley eriğe ait bir değerlendirme şekli sunmak üzere çeşitli çalışmalar yapılmış ve bu çalışma ile de bir reçete geliştirilmiştir. Bu çalışmada, fiziksel ve kimyasal özellikleri saptanan Stanley çeşidi erik materyal olarak kullanılarak, diğer erik değerlendirme çeşitlerine alternatif olabilecek, berrak tipte meyve suyu üretim olanakları araştırılmıştır. Reçetelerin düzenlenmesinde preslenip, durultulan naturel meyve suyunun bir bölümü direkt şişelenerek, kalan meyve suyunda asit ve şeker ilavesiyle 3 farklı tipte ayarlama yapılmıştır. Şişelenen 4 tip erik suyu 3 ay depolandıktan sonra; fiziksel, kimyasal ve duyusal özellikleri saptanmak üzere değerlendirmeye alınmışlardır. Yapılan analizler sonucunda; 1) Materyal olarak kullanılan Stanley çeşidi eriğin, albenili renkleri ve lezzetli tatları nedeniyle, berrak tipte meyve suyu yapımına uygun olduğu, 2) Duyusal değerlendirme sonucunda, 14 ve 15 briksli olarak üretilen erik sularının daha çok beğenildikleri anlaşılmıştır.Stanley variety of plum is among the convenient varieties of Marmara Region. This plum can be thought of one which can be evaluated for dining and drying. However, especially in recent years, in İnegöl region. Stanley variety of plum had a big increase in its production and people in that region have had great difficulties to make increase in value of their products. In order to find a solution to their difficulties, they applied to the government agencies. By taking these facts into the consideration, some studies have been done for presenting an evaluation related to Stanley plum to the producers, to the cooperative established by the people of that region,and to the industrial organizations. With this study, a prescription reflecting the evaluation has been developed.In this study the physical and Chemical properties of Stanley variety of plum are determined and by using this plum as material, possiblilities of producing clear type of fruit juice being an alternative to other plum evaluation variations are searched. In the design of prescription, a part of presse! and clarified natural fruit juice is bottled directly, to the rest of it, with the addition of acid and sugar, three different types of adjustment are done

The effect of heat shock protein 90 inhibitors on histone 4 lysine 20 methylation in bladder cancer

Heat shock protein 90 (HSP90), an ATP-dependent molecular chaperone required for the stability and function of numerous oncogenic signaling, is one of the hallmarks of cancer. Recent years, the studies showed that HSP90 plays a pivotal role in epigenetic pathways. Epigenetic regulation plays an important role in the etiology of bladder cancer. The aim of the present study was to investigate the effect of HSP90 proteins on DNA methylation and the levels of inactivated histone methylation markers in bladder cancers. The cytotoxic effect of geldanamycin (GA), a HSP90-specific inhibitor, in human bladder cancer cell line, T24, was studied by using WST1 (both time and dose-dependent), qPCR for the expression aberration of target genes DNMT1 and WIF-1 and western blot for the protein levels of DNMT1, Histone H4, Histone 4 lysine monomethylation (H4K20me1), Histone 4 lysine trimethylation (H4K20me3), Akt1, pAkt1 (S473) and Lysine methyltransferase 5C (KMT5C). High-dose GA treatment decreased cell proliferation. After the GA treatment, DNMT1 decreased at both transcriptional and translational levels due to Akt1 and pAkt1 (S473) inhibition. Following the GA-induced decrease in DNMT1, re-expression of WIF-1 gene was found at mRNA. In addition, the GA treatment resulted in doseand time-dependent upregulation/downregulation of histone post-translational modifications (H4K20me1 and H4K20me3) and the KMT5C enzyme responsible for these modifications. There was no significant change in the H4 protein level. These findings may offer a new approach for the determination of the molecular effect of HSP90 on epigenetic regulation and the identification of new molecular targets (HSP90 client proteins) for bladder cancer treatment

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DNA methylation is considered as one of the most important epigenetic mechanisms and it is catalyzed by DNA methyltransferases (DNMTs). DNMT1 abundance has been frequently seen in urogenital system tumors but the reasons for this abundance are not well understood. We aimed to look into the effects of Wnt/β-catenin signaling pathway on overexpression of DNMT1 and aberrant expression of UHRF1 and HAUSP which are responsible for stability of DNMT1 at transcriptional and protein levels in urogenital cancers. In this context, firstly, Wnt/β-catenin signaling pathway was activated by using SB216763 which is a glycogen synthase kinase-3 (GSK3) β inhibitor. Cell proliferation levels in bladder cancer cells, renal cell carcinoma, and prostate cancer cells treated with GSK3β inhibitor (SB216763) were detected by WST-1 reagent. WIF-1 gene methylation profile was determined by methylation-specific PCR (MSP); expression levels of target genes β-catenin and WIF-1 by real-time PCR; and protein levels of β-catenin, DNMT1, pGSK3β(Ser9), HAUSP, and UHRF1 by Western Blot. Our results indicated that treatment with SB216763 caused an increased cell proliferation at low dose. mRNA levels of β-catenin increased after treatment with SB216273 and protein levels of pGSK3β(Ser9), β-catenin, and DNMT1 increased in comparison to control. HAUSP and UHRF1 were either up-regulated or down-regulated at the same doses depending on the type of cancer. Also, we showed that protein levels of DNMT1, β-catenin, HAUSP, and UHRF1 decreased after re-expression of WIF-1 following treatment with DAC. In Caki-2 cells, β-catenin pathway might have accounted for the stability of DNMT1 expression, whereas such relation is not valid for T24 and PC3 cells. Our findings may offer a new approach for determination of molecular effects of Wnt/β-catenin signal pathway on DNMT1. This may allow us to identify new molecular targets for the treatment of urogenital cancers

A Local Study on Antenatal Features of Preterm Births at 26-32 Versus 33-36 Weeks of Pregnancy

Objective: The antenatal features of pregnancies affect the incidence of preterm births. This retrospective study from Trakya University of Northwestern Turkey, describes antenatal factors involved in preterm births at 26-32 weeks of pregnancy and compares with those involved in preterm births at 33-36 weeks. Study Design: The records of preterm births at 26-32 weeks (earlier preterm births, n=419) and at 33-36 weeks (later preterm births, n=158) during the years 2002-2010 were reviewed and the demographic, obstetric and medical features were evaluated retrospectively. The data was expressed as numbers and percentages and analyzed by SPSS 20.0. Results: Iron supplementation [OR 0.27 (0.16-0.45), p=<0.001], short cervix [OR 9.12 (2.09-39.73) p=0.003] and infection [OR 2.6 (1.2-5.6) p=0.014] were important factors in the emergence of earlier preterm births which occurred at the rate of 1.4%. Conclusions: Earlier preterm births at 26-32 weeks of pregnancy, which compose an obstetric issue, are associated with several antenatal risk factors such as nutrition, cervical problems and infections in Northwestern Turkey