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Background and Aims: The incidence of breast cancer is increasing day by day, especially in women. The search for new drugs against breast cancer is the focus of attention in research. Breast cancer and prostate cancer have remarkable biological similarities. Therefore, the 4-(4-chlorophenyl)-3-(1-(2-fluoro-[1,1'-biphenyl]-4-yl)ethyl)-5-((4-fluorobenzyl)thio)-4H1,2,4-triazole (SGK597) compound that is suppressing cell proliferation in prostate cancer, was studied in MCF-7 breast cancer and MCF-10A mammary epithelial cell lines. Methods: The WST-8 method was used to determine cell viability and cytotoxicity of SGK597 in MCF-7 and MCF10-A cell lines. The JC-1 test was applied to determine changes in mitochondrial membrane potential. The protein expression levels of Bax, Bcl-2, and c-PARP associated with apoptosis were determined using Western blot analysis. Results: After 24 and 48 hours of incubation of SGK597, the IC50 values were 28.74 μM and 17.28 μM for MCF-7; 65.9 μM and 50.5 μM for MCF-10A, respectively. Mitochondrial membrane potential showed a tendency toward depolarization in MCF-7 cells as a result of increasing concentration of SGK597, while the same tendency was not seen for MCF-10A. As a result of western blot experiments, no increase in the Bax/Bcl-2 ratio and c-PARP expression level was observed, indicating no apoptosis. Conclusion: It was observed that the compound SGK597 suppressed MCF-7 cell proliferation. These results indicate that SGK597 may be a candidate compound for use as an anticancer agent.

Ustundag, Fumet Duygu/0000-0001-5290-8924 WOS: 000550941000001 PubMed ID: 32693643 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a neurotoxin that damages dopaminergic neurons. Zebrafish has been shown to be a suitable model organism to investigate the molecular pathways in the pathogenesis of Parkinson's disease and also for potential therapeutic agent research. Boron has been shown to play an important role in the neural activity of the brain. Boronic acids are used in combinatorial approaches in drug design and discovery. The effect of 3-pyridinylboronic acid which is an important sub-class of heterocyclic boronic acids has not been evaluated in case of MPTP exposure in zebrafish embryos. Accordingly, this study was designed to investigate the effects of 3-pyridinylboronic acid on MPTP exposed zebrafish embryos focusing on the molecular pathways related to neurodegeneration and apoptosis by RT-PCR. Zebrafish embryos were exposed to MPTP (800 mu M); MPTP + Low Dose 3-Pyridinylboronic acid (50 mu M) (MPTP + LB) and MPTP + High Dose 3-Pyridinylboronic acid (100 mu M) (MPTP + HB) in well plates for 72 hours post fertilization. Results of our study showed that MPTP induced a P53 dependent and Bax mediated apoptosis in zebrafish embryos and 3-pyridinylboronic acid restored the locomotor activity and gene expressions related to mitochondrial dysfunction and oxidative stress due to the deleterious effects of MPTP, in a dose-dependent manner.

© 2022, The Author(s), under exclusive licence to Springer Nature B.V.Background: Preoperative chemoradiotherapy has long been accepted as a method to improve survival and lifetime quality of rectal cancer patients. However, physiologic effects of these therapies largely depend on the resistance of cells to the radiation, type of chemotherapeutic agents and individual responses. As one of the signaling cascades involved in chemo- or radiation- resistance, the present study focused on several proteins involved in pTEN/Akt/mTOR pathway to explore their prognostic significance. Materials and methods: Samples from advanced stage rectal cancer patients were analyzed to detect expression levels of pTEN/Akt/mTOR pathway related proteins pTEN, mLST8, REDD1, BNIP3, SAG and NOXA, together with p53, by RT-qPCR. Kaplan–Meier analysis was used to assess expression-survival relation and correlations among all proteins and clinicopathological features were statistically analyzed. Results.: Except p53, none of the proteins showed prognostic significance. High p53 expression presented clear impact on overall survival and disease free survival. It was also significantly related to pathologic complete response. p53 showed high correlation to local recurrence as well. On the other hand, strong correlation was observed with PTEN expression and tumor response, but not with survival. High associations were also observed between mLST8/REDD1, PTEN and NOXA, confirming their role in the same cascade. Conclusion: The contentious role of p53 as a prognostic biomarker in colorectal cancer was further affirmed, while PTEN and REDD1 could be suggested as potential candidates. Additionally, NOXA emerges as a conjunctive element for different signaling pathways.

Parkinson\"s disease (PD) is one of the most common forms of neurodegenerative diseases and research on potential therapeutic agents for PD continues. Rotenone is a neurotoxin that can pass the blood-brain barrier and is used to generate PD models in experimental animals. Boron is a microelement necessary for neural activity in the brain. Antioxidant, non-cytotoxic, antigenotoxic, anti-carcinogenic effects of boric acid, the salt compound of boron has been reported before. Boronic acids have been approved for treatment by FDA and are included in drug discovery studies and pyridine boronic acids are a subclass of heterocyclic boronic acids used in drug design and discovery as substituted pyridines based on crystal engineering principles. The aim of our study was to determine the effect of 3-pyridinylboronic acid in rotenone-exposed zebrafish embryos, focusing on oxidant-antioxidant parameters and gene expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2) target genes gclm, gclc, hmox1a, nqo1, and PD related genes, brain-derived neurotrophic factor, dj1, and tnfa alpha. Zebrafish embryos were exposed to Rotenone (10 mu g/l); Low Dose 3-Pyridinylboronic acid (100 mu M); High Dose 3-Pyridinylboronic acid (200 mu M); Rotenone + Low Dose-3-Pyridinylboronic acid (10 mu g/l + 100 mu M); Rotenone + High Dose-3-Pyridinylboronic acid (10 mu g/l + 200 mu M) in well plates for 96 h post-fertilization (hpf). Our study showed for the first time that 3-pyridinylboronic acid, as a novel sub-class of the heterocyclic boronic acid compound, improved locomotor activities, ameliorated oxidant-antioxidant status by decreasing LPO and NO levels, and normalized the expressions of bdnf, dj1, tnf alpha and Nrf2 target genes hmox1a and nqo1 in rotenone exposed zebrafish embryos. On the other hand, it caused the deterioration of the oxidant-antioxidant balance in the control group through increased lipid peroxidation, nitric oxide levels, and decreased antioxidant enzymes. We believe that these results should be interpreted in the context of the dose-toxicity and benefit-harm relationship of the effects of 3-pyridinylboronic.

BACKGROUND/AIMS Humic acid (HA) formed by the decomposition of organic matter is a high-molecular-weight polymer. Humic-derived substances are potential drugs for human health, and their role in the prevention and treatment of diseases, like diabetes, antiviral, UV-B protective effect, cancer, and cardiovascular disease, is emphasized in various studies. It has antiulcerogenic, antitumor. Yang et al. demonstrated that HA exerts an antiproliferative effect and growth inhibition on HL-60 cells through the induction of apoptosis. A study showed that the antiulcer effect of Shilajit (HAs extracted from Shilajit) samples may be due to antimicrobial, anti-inflammatory, antianxiety, antioxidant, healing, and regenerative effects. The present study aimed to investigate the effects of HA on cultures of the chronic myeloid leukemia cell line in vitro. MATERIAL and METHODS We determined the apoptotic effect of HA in K562 cells by using various molecular methods. For cell viability, HA was tested in vitro against K562 cell lines using the MTT colorimetric process. Bax, bcl-2, casp-3, and casp-9 expressions were quantified by Western blot. Apoptosis analysis has been made with the Annexin-V/PI method. RESULTS The IC50 values of HA were determined 100 mg mL(-1) for the K562 cell line. HA has been shown to inhibit cell proliferation. As the bcl-2/bax ratio decreased and increasing the levels of caspase-3 and caspase-9, the cells underwent apoptosis. CONCLUSION As a result, HA showed the anticancer effect on leukemia cells. In the future, HA can be used as a chemopreventive agent in leukemia treatments.

© 2022New thiosemicarbazides (3, 5-6), 1,2,4-triazoles (14-15) and thioethers (22-68) from derived (S)-Naproxen were synthesized in this study. The structure of these compounds were elucidated by spectral (FT-IR, 1H NMR, 13C NMR) methods, besides elemental analysis and TLC. The molecular binding of the compounds on MetAP-2 was performed. Anticancer effects of the synthesized compounds were studied by using MTT assay method on MCF-7 (includes oestrogene and progesterone receptors) and MDA-MB-231 (lacks estrogen and progesterone receptors) adenocarcinoma cell lines at 0, 10, 25, 50, 75 and 100 μM concentrations for 24 h. The IC50 values of novel (S)-Naproxen derivatives were determined between from 5 to 100 μM on MCF-7 breast cancer cell line and MDA-MB-231 cell lines. The apoptotic activity of selected compounds 22 and 42 were first analyzed by Annexin V staining using Tali Image-Based Cytometer. Mitochondrial membrane potential changes determined in fluorescence plate reader following JC-1 stain for compounds 22 and 42 in MCF-7 and MDA-MB-231 cells. The effect of these compounds on the cell viability 4T1 mouse mammary tumor cell line was tested at 1 to 5 times of calculated IC50 value (IC50x1, IC50x2, IC50x3, IC50x4, and IC50x5). Next in order to determine the toxicity of the combination of compound 51 and Docetaxel, WST-1 cell viability and proliferation assay was performed with 4T1.

Understanding cell responses to the topography they are interacting with has a key role in designing surfaces due to the distinctiveness in the responses of different cell types. Thus far, a variety of surface textures have been fabricated, and the cellular responses of diversified cell lines to the surface textures have been assessed together with surface chemistry. However, the results reported in the literature are contradictory, and also not in-depth for inferring the relevance between cells, surface chemistry, and surface topography. Starting from this point of view, we focused on fabricating surfaces having extracellular matrix-like surface patterns and investigated the influence of patterning on human ovarian cancer cells. In this study, hemispherical protrusion-shaped, nanotextured surfaces were prepared via colloidal lithography and polymer casting methods using monolayer templates prepared from 280 nm, 210 nm, and 99 nm polystyrene particles and polydimethylsiloxane moulds. Then, the surface textures were transferred to biocompatible polycaprolactone films. After the characterisation of the surfaces via atomic force microscopy, X-ray photoelectron spectroscopy, and contact angle measurements, the cellular response to topography was evaluated by cell attachment, viability, and apoptosis studies. The results were compared with non-textured surfaces and control plate wells. The results showed that human ovarian cancer cell attachment increased with nanotexturing, which suggests that nanotexturing may be a promising approach for cancer cell modulation, and may have the potential to introduce new strategies for cancer treatment.

Clinical and experimental studies show that epilepsy affects cardiac function; however, the underlying molecular mechanism has not been fully elucidated. Inwardly-rectifying potassium (Kir) channels transport K+ ions into excitable cells such as neurons and cardiomyocytes; they control the cell excitability by acting towards the repolarization phase of cardiac action potential. Kir channel expression has been previously shown to vary in epileptic brains, at the same time seizures are known to affect the autonomic nervous system. Kir channel expression in cardiac tissue is a possible mechanism for the explanation of cardiac pathology in epilepsy. We investigate the expression of Kir channels in epileptic cardiac tissue by using pentylenetetrazole (PTZ)-kindling model in rats. Our molecular analyses showed significant decrease in cardiac Kir channel mRNA and protein expression of PTZ-kindled rats. Interestingly, both Kir2.x, which directs IK1 flux in ventricular tissue and Kir3.x, which is responsible for IKACh in the atria, were observed to decrease significantly. Kir channel expression also differs between females and males. This is the first study to our knowledge in epileptic cardiac tissue showing the expression of Kir channels. Our results show that Kir channels may play a role in cardiac pathology associated with epilepsy.

Regulation of telomerase activity is thought to participate in the cellular response to ionizing radiation. Epigenetic mechanisms play a role in this regulation, as well as other mechanisms such as transcription, phosphorylation, etc. Here, we investigated chromatin modifications in telomerase promoter upon exposure to ionizing radiation in human mesenchymal stem cells (hMSC) and telomerase-immortalized hMSCs (hMSC-telo1) together with a hMSC-telo1 cell line in which TRF2 expression was partially repressed (siTRF2 hMSC-telo1). Histone methylations and acetylations were compared in all cell lines after exposure to various doses of ionizing radiation (0.1, 1, 2.5 and 15 Gy) using chromatin immunoprecipitation assay. hTERT gene was shown to be quickly regulated through H3, H4 acetylations, as well as with H3K4 and H3K9 methylations, following radiation exposure, although the kinetic of hMSC-telo1 cells were different, indicative of the higher radioresistivity of these cells. To the author's surprise, there was an upregulation of endogenous telomerase activity in the hMSC-telo1 cells, even though the cells had already expressed high levels of ectopic hTERT. Our results show that telomerase regulation is one of the primary actions in response to damage and epigenetic factors play a major role in this regulation. Our results also suggested that partial silencing of TRF2 enhances the radiosensitivity of these cells, and endogenous telomerase is upregulated upon radiation, even under ectopic expression of hTERT in these cells.

An approach for cancer treatment is modulation of tumor microenvironment. Based on the role of extracellular matrix in cell modulation, fabrication of textured materials mimicking extracellular matrix could provide novel opportunities such as determining cancer cell behaviour. With this background, in this work, we have fabricated doxorubicin hydrochloride loaded nanotextured films which promote topographical attachment of cancer cells to film surface, and eliminate cells by release of the anti-cancer drug encapsulated within the films. These films are designed to be placed during surgical removal of the tumor with the intent to prevent ovarian cancer recurrence by capturing cancer cell residuals. With this aim, hemispherical protrusion shaped surface textures were acquired using colloidal lithography technique using 280 nm, 210 nm or 99 nm polystyrene particles. Once moulds were formed, nanotextured films were obtained by casting water-in-oil stable polycaprolactone emulsions encapsulating doxorubicin hydrochloride. Films were then characterized, and evaluated as drug delivery systems. According to results, we found that template morphologies were successfully transferred to films by atomic force microscopy studies. Hydrophilic surfaces were formed with contact angle values around 40 degrees.In-vitrodrug release studies indicated that nanotextured films best fit into the Higuchi model, and similar to 30% of the drug is released from the films within 60 days. Cell culture results indicated increases in the attachment and viability of human ovarian cancer cells to nanotextured surfaces, particularly to the film fabricated using 99 nm particles. Our results demonstrated that delivery of anti-cancer drugs by use of nanotextured materials could be efficient in cancer therapy, and may offer new possibilities for cancer treatment.

The dopamine transporter (DAT) plays a role in the termination of dopaminergic neurotransmission; thereby it is accepted as the primary target of various psychostimulants. N-terminal phosphorylation of DAT has been proposed as a regulator in different DAT functions, such as amphetamine-induced efflux or PKC/PKA-mediated responses. To understand the role of N-terminal conformational changes in dopamine transporter structure and function, the fluorescence resonance energy transfer (FRET) method was applied to various DAT constructs fluorescently labeled at the N-terminus and substrate-induced conformational changes were determined using rhodamine-labeled cocaine analog JHC1-64 in three DAT mutants. The results indicated that the construct with YFP inserted into the N-terminal position-55 displayed effective interaction with the substrate and simultaneous mutation of two serine residues (S7 and S12) to alanine or aspartic acid demonstrated similar phenotypes as their wild-type (WT) counterparts. FRET was detectable for N-terminal p55 YFP WT, Ser/Ala, and Ser/Asp forms, but there was no significant difference among the three mutants, contrary to our expectations based on the previously proposed roles of these serine residues. In addition, three mutants (K264A, D345A, and Y335A) implemented in the position-55 YFP background were also investigated and the importance of Y335 in the translocation cycle and in the process of substrate release was verified.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly used as anti-inflammatory and analgesic agents. This family of drugs suppresses prostaglandin synthesis through inhibition of cyclooxygenase (COX) enzymes. Recent studies showed that anti-carcinogenic effects of these drugs are especially mediated by COX-2 enzyme. Etodolac is a COX-2 inhibitor and though not perfectly selective, it exhibits “preferential selectivity„ for COX-2. In this study, the anti-proliferative and apoptotic effects of etodolac and its hydrazone or triazole derivatives (SGK 206 and SGK 242, respectively), were investigated on prostate cancer cell line PC-3 and human colorectal carcinoma cell line HT-29. Our data showed that SGK 206 and SGK 242 were more effective in the inhibition of proliferation and induction of apoptosis compared to etodolac in both cell lines.

The anticancer activity of Zingiber officinalis (ginger) is an area of active research. However, data is quite limited regarding its action and mechanism, especially in hematologic cancer types. Here, antiproliferative and apoptotic effects of whole extract of the rhizome of Zingiber officinalis (ZOWE), was investigated in K562 cell line derived from a chronic myeloid leukemia (CML) patient. Various concentrations of whole extract (0, 10, 25, 50 and 100 μM) were tested in K562 cultures. Cytotoxicity and apoptosis was assessed with appropriate methods, as well as cellular ROS levels. In this study, we showed that ZOWE inhibited proliferation of K562 cells substantially, when compared to peripheral blood mononuclear cells (PBMCs) isolated from healthy donor. Increased Bax/Bcl-2 ratio, reduced mitochondrial membrane potential and increased PARP cleavage demonstrated that ZOWE inhibited proliferation by induction of apoptosis. These changes were coupled with an increase of reactive oxygen species (ROS) production. Furthermore, ZOWE addition to the culture also reduced expression levels of survival proteins pAkt and survivin, in a concentration dependent manner. Our results clearly mark that ZOWE causes to a reduction in cell viability, an induction of apoptosis and elevation in ROS levels in chronic myeloid leukemia cells and effects are significantly different from healthy peripheral blood mononuclear cells, further supporting its potential therapeutic value.

Tolmetin hydrazide and a novel series of tolmetin hydrazide-hydrazones 4a-l were synthesized in this study. The structures of the new compounds were determined by spectral (FT-IR, (1)H NMR) methods. N'-[(2,6-Dichlorophenyl)methylidene]-2-[1-methyl-5-(4-methylbenzoyl)-1H-pyrrol-2-yl]acetohydrazide (4g) was evaluated in vitro using the MTT colorimetric method against the colon cancer cell lines HCT-116 (ATCC, CCL-247) and HT-29 (ATCC, HTB-38) to determine growth inhibition and cell viability at different doses. Compound 4g exhibited anti-cancer activity with an IC50 value of 76 μM against colon cancer line HT-29 (ATCC, HTB-38) and did not display cytotoxicity toward control NIH3T3 mouse embryonic fibroblast cells compared to tolmetin. In addition, this compound was evaluated for caspase-3, caspase-8, caspase-9, and annexin-V activation in the apoptotic pathway, which plays a key role in the treatment of cancer. We demonstrated that the anti-cancer activity of this compound was due to the activation of caspase-8 and caspase-9 involved in the apoptotic pathway. In addition, in this study, we investigated the catalytical effect of COX on the HT-29 cancer line, the apoptotic mechanism, and the moleculer binding of tolmetin and compound 4g on the COX enzyme active site.

4-Chloro-3-({[(substitutedamino)carbonothioyl]amino}sulfonyl)-N-(2-methyl-2,3-dihydro-1H-indole-1-yl)benzamide (1–20) and 4-chloro-3-({[3-(substituted)-4-oxo-1,3-thiazolidine-2-ylidene]amino}sulfonyl)-N-(2-methyl-2,3-dihydro-1H-indole-1-yl)benzamide derivatives (21–31) were synthesized from 4-chloro-N-(2-methyl-2,3-dihydroindol-1-yl)-3-sulfamoylbenzamide (indapamide). 4-Chloro-3-({[(4-chlorophenyl) amino) carbonothioyl]amino}sulfonyl)-N-(2-methyl-2,3-dihydro-1H-indole-1-yl)benzamide 12 demonstrated the highest proapoptotic activity among all synthesized compounds on melanoma cell lines MDA–MB-435 with 3.7% growth inhibition at the concentration of 10 µM. Compound 12 (SGK 266) was evaluated in vitro using the MTT colorimetric method against melanoma cancer cell line MDA–MB435 growth inhibition for different doses and exhibited anticancer activity with IC50 values of 85–95 µM against melanoma cancer cell line MDA–MB435. In addition, this compound was investigated as inhibitors of four physiologically relevant human carbonic anhydrase isoforms, hCA I, II, IX and XII. The compund inhibited these enzymes with IC50 values ranging between 0.72 and 1.60 µM.

Tolmetin hydrazide and a novel series of tolmetin hydrazide-hydrazones 4a-l were synthesized in this study. The structures of the new compounds were determined by spectral (FT-IR, (1)H NMR) methods. N'-[(2,6-Dichlorophenyl)methylidene]-2-[1-methyl-5-(4-methylbenzoyl)-1H-pyrrol-2-yl]acetohydrazide (4g) was evaluated in vitro using the MTT colorimetric method against the colon cancer cell lines HCT-116 (ATCC, CCL-247) and HT-29 (ATCC, HTB-38) to determine growth inhibition and cell viability at different doses. Compound 4g exhibited anti-cancer activity with an IC50 value of 76 μM against colon cancer line HT-29 (ATCC, HTB-38) and did not display cytotoxicity toward control NIH3T3 mouse embryonic fibroblast cells compared to tolmetin. In addition, this compound was evaluated for caspase-3, caspase-8, caspase-9, and annexin-V activation in the apoptotic pathway, which plays a key role in the treatment of cancer. We demonstrated that the anti-cancer activity of this compound was due to the activation of caspase-8 and caspase-9 involved in the apoptotic pathway. In addition, in this study, we investigated the catalytical effect of COX on the HT-29 cancer line, the apoptotic mechanism, and the moleculer binding of tolmetin and compound 4g on the COX enzyme active site.

BACKGROUND: Telomeres are protective caps consisted of specific tandem repeats (5′-TTAGGG-3′). Shortening of telomeres at each cell division is known as "mitotic clock" of the cells, which renders telomeres as important regulators of lifespan. TRF2 is one of the critical members of shelterin complex, which is a protein complex responsible from the preservation of cap structure, and loss or mutation of TRF2 results in DNA damage, senescence or apoptosis. Since cancer is frequently associated with aberrant cell cycle progression, defective DNA repair or apoptosis pathways, TRF2 could be one likely candidate for cancer therapy. Here we investigated the prognostic role of TRF2 levels in cervical cancer patients. Fold-induction rates were evaluated with respect to median values after real-time PCR analysis. Overall survival, distant disease-free and local recurrence-free survival rates were calculated using Kaplan-Meier long rank test. RESULTS: Both five year overall- and disease-free survival rates were longer in patients with higher TRF2 expression compared to lower expression, but results were not statistically significant (69.2% vs 28.9%, respectively). Mean local recurrence-free survivals (LRF) were very close ( 58.6, CI: 44.3-72.9 vs 54.5, CI: 32.1-76.9 months) for high and low expressions, respectively. Cumulative proportion of LRF at the end of five year period was 76.9% for high and 57.1% for low TRF2 expression (P = 0.75). Statistically significant difference was found between survival ratios and Bcl-xL and p53 gene expressions, but not with TRF2. A respectable correlation between TRF2 expression and apoptosis along with distant metastasis was noted (P = 0.045 and 0.036, respectively). Additionally, high TRF2 expression levels had a positive impact in five year survival rate of stage IIIB-IVA patients (P = 0.04). CONCLUSIONS: Our results support the role of TRF2 in apoptosis and imply a positive relation with distant metastases and survival in advanced stage patients. The remarkable difference in survival periods of patients with different TRF2 expressions suggest that TRF2 may be a candidate factor to estimate survival for cervical cancer, a preliminary observation which should further be verified with a larger cohort.

Dopamine has been shown to influence blood pressure by regulating renal sodium excretion through direct interaction with the dopamine receptors, especially with the Dopamine D1 receptor (DRD1). To better understand the role of polymorphisms in those effects, we investigated the association between two polymorphic sites in the DRD1 promoter region (A-48G, G-94A) and essential hypertension in the Turkish population. The DRD1 variants were genotyped by restriction fragment length polymorphism (RFLP) analysis. A total of 205 unrelated individuals were enrolled in the study. We found that genotype distributions and allele frequencies of the control and hypertensive subjects were very similar and did not show any significant difference with respect to blood pressure (BP) and hypertension. Contribution of the gene variances in BP or hypertension by sex differences and dependence on body mass index (BMI) were also evaluated. Distribution of genotypes and allele frequencies were found to be in line with previous reports. However, increments detected in hypertensive subjects were far from being statistically significant.

Amac: Dopamin tasiyicilar (DAT) dopaminin sinaptik gerialimindan ve boylelikle sinirsel iletinin cabuk sonlanmasindan sorumludur. Bu proteinler aliskanlik yapici kokain, amfetamin gibi maddelerin de etki merkezlerini olusturur. Proteinin N-ucunda bulunan Serin 7 ve 12’nin fosforilasyon bakimindan kritik amino asitler oldugu bilinmektedir. Bu calismada N-ucu ile kokain baglama bolgesi arasindaki uzaklik, yaban tip ve Ser7/12Asp ve Ser7/12Ala mutantlarinda belirlenmistir. Gerec ve Yontem: Sari-floresan protein (YFP) ve mutasyonlar DAT’in N-terminal ucuna 2-asamali PCR ile yerlestirilmistir. Dopamin geri-alim testleri yapilmis ve konfokal mikroskopta verici-agartma yontemi ile Forster rezonans enerji transferi (FRET) mesafeleri belirlenmistir. Bulgular: Tum N-ucundan YFP-etiketlenmis DAT proteinleri duzgun olarak hucre yuzeyine tasinmistir. Km degerleri sirasiyla yaban tip, Ser7/12Ala and Ser7/12/Asp mutantlari icin 0.76, 0.75 ve 1.24 mM olarak saptanmistir. FRET etkinligi yaban-tip DAT icin 0.15 olarak bulunmus ve FRET uzakligi 68.4 A olarak bulunmustur. Diger konstraktlarda FRET etkinlikleri negatiftir ve enerji transferi saptanmamistir. Sonuc: Veriler bize N-ucu bolgesinin kokain baglama bolgesi ile FRET mesafesinde oldugunu ve kritik serin rezidulerinin mutasyonlarinin (Ser7/12Asp) bu mesafeyi arttirdigini gostermektedir

Pinar Mega Tiber,1 Latife Baloglu,2 Sevgi Ozden,3 Zerrin Ozgen,4 Hazan Ozyurt,3 Makbule Eren,3 Oya Orun11Department of Biophysics, Marmara University, School of Medicine, Maltepe, Istanbul, Turkey; 2Laboratory Medicine, Karolinska Institute Biomedical Laboratory Science, Stockholm, Sweden; 3Clinic of Radiation Oncology, Dr Lutfi Kirdar Kartal Education and Research Hospital, Istanbul, Turkey; 4Department of Radiation Oncology, Marmara University, School of Medicine, Kaynarca, Istanbul, TurkeyObjective: To evaluate the expressions of several apoptotic pathway proteins in relation to clinical parameters and survival in patients with cervical carcinoma.Methods: A total of 20 patients with clinically advanced staged carcinoma of cervix (International Federation of Gynecology and Obstetrics [FIGO] stage IIB-IVA) aged from 40 to 75 years were included in this study. The expression profile of anti-apoptotic protein (sensitive to apoptosis gene [SAG]), mitochondrial apoptotic proteins (B-cell lymphoma-extra-large [Bcl-xL] and Bcl-2 homologous antagonist/killer [Bak]), and tumor suppressor proteins (p73 and p53) were examined by real-time polymerase chain reaction experiments along with their relation to clinical parameters and survival analyses during follow-up for 5 to 8 years.Results: No significant difference was found in the expressions of SAG, Bcl-xL, Bak, p73 and p53 proteins with respect to stage and grade of tumor. A significant positive correlation was noted between SAG and Bcl-xL genes (r=0.752, P