Targeting of Notch, IL-1, and leptin has therapeutic potential in xenograft colorectal cancer.

Background/aim: Colorectal cancer (CRC) is a fatal malignancy type and its occurence still needs to be explored mechanistically. Notch, IL-1, and leptin crosstalk is reported to play a role in the proliferation, migration, and expression of proangiogenic molecules. In this study, we aimed to investigate the effect of inhibition of Notch, IL-1, and leptin on CRC. Materials and methods: To generate colorectal cancer tumor xenografts, 1 × 107 cells from exponentially growing cultures of HCT-15 cells were injected subcutaneously, at the axillary region of the left and right rear flanks of forty NOD.CB17-Prkdcscid/J (NOD/SCID) female mice. The mice were then monitored for the development of tumors and were randomly divided into five groups when tumor sizes reached a volume of approximately 150 mm3. Mice were used to determine the effectiveness of the gamma-secretase inhibitor (DAPT, Notch inhibitor), the interleukin-1 receptor antagonist (Anakinra) and the leptin receptor antagonist (Allo aca) against tumor growth. The mice were euthanized by CO2 inhalation immediately after the treatments finished, and all efforts were made to minimize suffering. Tumors were excissed for RT-PCR and histological analysis. Results: There is an intact Notch, IL-1, and leptin signaling axis, and in vivo antagonism of Notch, IL-1, and leptin affects mRNA and protein expression of inflammatory and angiogenic molecules. Conclusion: Present data suggest that targeting Notch, IL-1, and leptin may be possesses therapeutic potential in CRC.

The Effect of Antioxidant Astaxanthin on Intestinal Ischemia Reperfusion Damage in Rats.

BACKGROUND: Mesenteric ischemia is a frequently encountered disease in surgical clinics, difficult to diagnose, and very mortal if not treated. Our study investigated the effects of astaxanthin, which is known to have potent antioxidant properties and is also known to have anti-inflammatory effects on ischemia-reperfusion (I/R) injury. METHODS: A total of 32 healthy Wistar albino female rats were used in our study. Subjects were randomized and equally divided into 4 groups; control (laparotomy group only), I/R (transient mesenteric ischemia group only), astaxanthin 1 mg/kg and 10 mg/kg doses. The transient ischemia time was 60 minutes and the reperfusion time was 120 minutes. Tissue samples were taken from intracardiac blood and terminal ileum after reperfusion. Superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA) from blood samples, interleukin-1 (IL-1), IL-6, tumor necrosis factor-α (TNFα), Caspase-3, P53 tests from terminal ileum were studied. Tissue samples were also taken for histopathological evaluation. RESULTS: At the end of the study, both doses of astaxanthin were found to significantly reduce MDA level, CAT, and SOD enzymatic activity, whereas higher doses of astaxanthin significantly reduced MDA level, CAT, and SOD enzyme activities. In addition, cytokines such as TNFα, IL-1 and IL-6 were found to be reduced at both doses of astaxanthin, but only significantly inhibited at higher doses. We observed that inhibition of apoptosis reduced caspase-3 activity and P53 and deoxyribonucleic acid (DNA) fragmentation. CONCLUSION: Astaxanthin, a potent antioxidant, and anti-inflammatory, significantly reduces ischemia and reperfusion injury, especially when used at a dose of 10 mg/kg. These data need to be confirmed by larger animal series and clinical studies.

Magnetic Targeting of 5-Fluorouracil-Loaded Liposome-Nanogels for In Vivo Breast Cancer Therapy and the Cytotoxic Effects on Liver and Kidney.

In our previous paper, we demonstrated the ex vivo studies of non-toxic liposome-nanogel systems by which the long-term drug release could be provided from hybrid systems for the 5-fluorouracil (5-FU) drug molecule. The aim of this study was the in vivo magnetic targeting of 5-FU-loaded Fe3O4 nanoparticles including DPPC liposome-based PEGylated nanogels (5-FU loaded Fe3O4LPN) to breast cancer tissue and the investigation of the treatment and cytotoxic effects of that hybrid system to the liver and kidney in CD-1 mice using an external magnetic field. The effectiveness of the control, 5-FU group, Fe3O4LPN, and 5-FU-loaded Fe3O4LPN systems was evaluated using histopathology in terms of p53, ESR, PRG and C-erB-2, and qRT-PCR in terms of TYMS, ESR-1, RPG, and EGRF. Also, the cytotoxicity was analyzed by histopathological evaluation of kidney and liver tissues. Caspase-3 and caspase-9 evaluations were performed by qRT-PCR. The creatinine and ALT levels were also evaluated by comparing the blood samples of all groups. A total of 300-nm TEM-sized Fe3O4LNP hybrid system was successfully prepared. That system significantly decreased the TYMS and ESR1 levels after treatment process and increased the levels of p53 expression. The levels of caspase-3 mRNA did not change during the treatment, but the level of caspase-9 mRNA level was significantly decreased. The magnetically targeted liposome-based nanogel hybrid system is promising an effective therapy for the breast tumor with less liver and kidney damage. This Fe3O4LNP hybrid system could be useful for the similar small molecules.

miR663 Prevents Epo Inhibition Caused by TNF-Alpha in Normoxia and Hypoxia.

OBJECTIVE: In chronic inflammatory diseases, proinflammatory cytokines such as TNF-α are present in high amounts in the circulation and are associated with anemia in most cases. Experimental studies have shown that TNF-α inhibits the synthesis of erythropoietin (Epo), the main stimulant of hematopoiesis. Our aim was to figure out which microRNAs are involved in the Epo repression by TNF-α. METHODS: First, we determined the dose of TNF-α in HepG2 cells that has no cytotoxic effect by using MTT assays and that inhibits Epo synthesis by qRT-PCR and ELISA. Then, we performed the microRNA array study with TNF-α (20 ng/ml)-treated cells, and the array results were confirmed by qRT-PCR. We transfected the miR663 group with the mimic-miR663 (30 pmol) for 24 hrs; other groups were treated with a transfection reagent followed by treatment of TNF-α for 24 hrs; miR663 groups were treated with TNF-α for 24 hrs; and the control group was incubated with normal medium. We analyzed Epo mRNA levels by qRT-PCR. If mimic-miR663 prevents the Epo repression by TNF-α, more Epo-dependent UT-7 cells would survive. Therefore, we cocultured HepG2 cells with UT-7 cells. The percentage of apoptotic UT-7 cells was determined by TUNEL assays. RESULTS: According to our array study, TNF-α significantly decreases miR663 expression. After transfection of miR663 mimics into HepG2 cells, TNF-alpha was unable to decrease Epo mRNA amounts. Furthermore, mimic-miR663 transfection resulted in a lower apoptosis rate of UT-7 cells in coculture experiments. CONCLUSIONS: miR663 is involved in Epo mRNA production and that is able to prevent or reverse the inhibitory effect of TNF-α. In our coculture study, transfecting HepG2 cells with miR663 mimics decreased the apoptosis of UT-7 cells.

The role of anakinra in the modulation of intestinal cell apoptosis and inflammatory response during ischemia/reperfusion

Background/aim: Even though interleukin-1 receptor antagonist, IL-1Ra, is used in certain inflammatory diseases, its effect on ischemia-reperfusion injury is a current research topic. We aimed to investigate the protective effects of anakinra, an IL-1Ra, on the I/R induced intestinal injury. Materials and methods: The rat model of intestinal ischemia-reperfusion was induced. Rats were randomized into 4 groups: (group 1) control group, (group 2) I/R group, (group 3 and 4) treatment groups (50 mg/kg and 100 mg/kg, respectively). Gene expressions of caspase-3, TNF-α, IL-1α, IL-6, and apoptotic cells in tissue samples were evaluated by PCR and TUNEL methods, respectively. Plasma levels of superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA) were studied by the ELISA method and tissue samples were examined histopathologically as well. Results: Anakinra inhibited the expression of IL-1α, IL-6, and TNF-α and decreased the SOD, CAT, and MDA caused by ischemia- reperfusion injury in both treatment groups. Caspase-3 expression and TUNEL-positive cell number in treatment groups were also less. Histopathologically, anakinra better preserved the villous structure of the small intestine at a dose of 100 mg/kg than 50 mg/kg. Conclusion: Anakinra decreased the intestinal damage caused by ischemia-reperfusion and a dose of 100 mg/kg was found to be histopathologically more effective.

Role of Notch, IL-1 and leptin expression in colorectal cancer.

An increasing number of studies have shown that angiogenesis has an important role in the progression of cancer. The growth of a new network of blood vessels is crucial for tumor growth and metastasis, which is promoted by several proangiogenic factors. Leptin, an essential adipokine that is secreted from fat tissue, is one of these pro-angiogenic factors. It has been shown that the inhibition of leptin-induced angiogenesis resulted in decreased levels of vascular endothelial growth factor (VEGF)/VEGFR2, hypoxia inducible factor (HIF) 1α, NF-κB, IL-1 and Notch and reduced the tumor growth in breast cancer. Leptin induces angiogenesis in breast cancer either by upregulating VEGFR2 in endothelial cells or by increasing VEGF/VEGFR2 expression through the Notch, IL-1 and leptin crosstalk outcome (NILCO) pathway. NILCO is a novel mechanism that interacts with proinflammatory and proangiogenic signals, which are critical for cell proliferation and angiogenesis in cancer. Several studies have shown that components of NILCO may affect human cancer incidence and progression. However, to the best of our knowledge, the interactions between Notch, IL-1 and leptin in human colorectal cancer have not been yet studied at the molecular level. The aim of the present study was to investigate the expression levels of genes related to the NILCO pathway in human colorectal cancer specimens. The current results demonstrated that leptin, leptin receptor (ObR) b, Notch-1, Notch-4, IL-1α, IL-1ß, IL-1R, IL-6, JAK-2, STAT-1, STAT-3, VEGFA, VEGFR1, VEGFR2, TNF-α and NF-κB mRNA expression levels in the cancer tissue were increased compared with the normal tissue. No significant changes in the mRNA expression levels of Jagged-1, HIF-1α and TNF receptor 1 were observed. Western blotting revealed that the protein expression levels of IκB were increased in the cancer tissue compared with normal tissue, whereas HIF-1α and phosphorylated STAT-1 levels were decreased. IL-6 and VEGFA plasma concentrations were statistically raised and the leptin plasma concentration was also raised, although significantly, patients with cancer compared with control individuals. Together, the present findings indicated that Notch, IL-1 and leptin may serve a crucial role in the development of colorectal cancer.

Erythropoietin Protects the Kidney by Regulating the Effect of TNF-α in L-NAME-Induced Hypertensive Rats.

BACKGROUND/AIMS: Hypertension is the leading cause of death worldwide. Chronic high blood pressure induces inflammation. Tumor necrosis factor (TNF)-α plays a major role in inflammation and also depresses the synthesis of erythropoietin, which exerts protective effects on tissue; however, the mechanism is still unclear. We investigated the protective effect of erythropoietin against tissue damage caused by hypertension in the kidney and whether this effect was suppressed by TNF-α. METHODS: First, we detected the optimum chronic dose for darbepoetin-α (Depo), which is a long-acting erythropoietin analog for rats. We separated 60 female adult rats into 6 groups: control, Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME), L-NAME+Depo, L-NAME+Remicade (an anti-TNF-α antibody), L-NAME+Depo+Remicade, Depo, and control. After 1 month of treatment, we measured cardiovascular parameters, took blood samples, sacrificed the rats, and removed kidneys for analyses. RESULTS: The apoptotic index and the plasma and kidney mRNA levels of TNF-α increased in the L-NAME group and decreased in all other treatment groups. Macrophage accumulation increased in the L-NAME and L-NAME+Remicade groups, while it decreased in the Depo group. The mRNA abundance of TNF receptor 1 (TNFR1) decreased slightly in the Depo group and TNFR2 increased significantly in the same group. CONCLUSION: Erythropoietin protects kidney tissue against hypertension by preventing the apoptotic effects of TNF-α by blocking macrophage accumulation, decreasing TNF-α levels, and switching the TNF-α receptors from the apoptotic receptor TNFR1 to the proliferative receptor TNFR2.

Comparison of a ceramidase inhibitor (ceranib-2) with C2 ceramide and cisplatin on cytotoxicity and apoptosis of glioma cells.

Inhibiting ceramidase activity in cancer cells has been identified as a promising target for cancer therapy in recent studies. uhTs, we examined the possible role of ceranib-2, a novel ceramidase inhibitor, on growth and apoptotic mechanisms of the human normal glia cell line (HNA), human glioma cell lines (T-98G and U-87MG), and a rat glioma cell line (C6). We also compared the results with the effects of C2 ceramide and cisplatin. We determined the in vitro survival rate with MTT assay, apoptosis with flow cytometry, gene expressions with qRT-PCR, and statistical significance by one-way analysis of variance together with Tukey's test. Calculated from MTT outcomes, the inhibitory ranking was as follows: T-98G > U-87MG > C6 > HNA. Ceranib-2 had the most growth-suppressive activity on human T-98G cells with an IC50 of 7 µM for 24 h and 0.9 µM for 48 h. Only the 25 µM dose of ceranib-2 induced apoptosis of human T-98G and U-87MG cells after 24 h of treatment; however, it increased apoptosis of C6 cells dose- and time-dependently. Ceranib-2 increased the cytochrome c gene expression level during 24 h in T-98G cells. Ceranib-2 had cytotoxic and apoptotic effects on glioma cells but the cytotoxic effect was weaker on normal glia cells. This cytotoxicity was stronger than that of C2 ceramide and cisplatin.

Parenchymal Pressure Inconsistency in Different Brain Areas After Kaolin Injection into the Subarachnoid Space of Neonatal Rats.

AIM: To describe the relationship between the parenchymal pressure changes and the development of hydrocephalus in kaolininjected neonatal rats according to cerebral regions and time intervals of developing hydrocephalus. MATERIAL AND METHODS: Neonatal rats aged 2 to 3 days were examined in 5 groups as kaolin frontal "K-F", kaolin parietal "KP", saline frontal "SF-F", saline parietal "SF-P" and control "C", based on the injected material and injection sites. All injections were performed into the cortical subarachnoid space of the right frontal and right parietal regions. The fifth group was injection free. On the 3 < sup > rd < /sup > , 7 < sup > th < /sup > , 15 < sup > th < /sup > , 30 < sup > th < /sup > and 60 < sup > th < /sup > days after injection, parenchymal pressures (PP) of 5-7 rats from each group were measured from different regions. RESULTS: We compared the control group with saline-injected and kaolin-injected groups and found statistically significant parenchymal pressure differences based on regional measurements. In the kaolin groups, the mean PP values were obviously higher than the saline-injected group. Within each kaolin-injected group, the pressure values were variable and inconsistent regarding the parenchymal regions. CONCLUSION: Hydrocephalus cannot be totally explained with existent "bulk-flow" or "hydrodynamic" theories. Although our experimental design was planned to develop hydrocephalus according to the bulk flow theory, our results were more compatible with the hydrodynamic theory. The present comments on the occurrence and pathogenesis of hydrocephalus are still open to debate and may require further comprehensive studies.

Vasoactive intestinal peptide, a promising agent for myopia?

AIM: To investigate the role of vasoactive intestinal peptide (VIP) in form-deprivation myopia (FDM). METHODS: FDM was created in three groups of eight chicks by placing a translucent diffuser on their right eyes. Intravitreal injections of saline and VIP were applied once a day into the occluded eyes of groups 2 and 3, respectively. Retinoscopy and axial length (AL) measurements were performed on the first and 8th days of diffuser wear. The retina mRNA levels of the VIP receptors and the ZENK protein in right eyes of the three groups and left eyes of the first group on day 8 were determined using real time polymerase chain reaction (PCR). RESULTS: The median final refraction (D) in right eyes were -13.75 (-16.00, -12.00), -11.50 (-12.50, -7.50), and -1.50 (-4.75, -0.75) in groups 1, 2, and 3, respectively (P<0.001). The median AL (mm) in right eyes were 10.65 (10.00, 11.10), 9.90 (9.70, 10.00), and 9.20 (9.15, 9.25) in groups 1, 2, and 3, respectively (P<0.001). The median delta-delta cycle threshold (CT) values for the VIP2 receptors were 1.07 (0.82, 1.43), 1.22 (0.98, 1.65), 0.29 (0.22, 0.45) in right eyes of groups 1, 2, and 3, and 1.18 (0.90, 1.37) in left eyes of group 1, respectively (P=0.001). The median delta-delta CT values for the ZENK protein were 1.07 (0.63, 5.03), 3.55 (2.20, 5.55), undetectable in right eyes of groups 1, 2, and 3 and 1.89 (0.21, 4.73) in left eyes of group 1, respectively (P=0.001). CONCLUSION: VIP has potential inhibitory effects in the development of FDM.

Apium graveolens Extract Inhibits Cell Proliferation and Expression of Vascular Endothelial Growth Factor and Induces Apoptosis in the Human Prostatic Carcinoma Cell Line LNCaP.

Apium graveolens has been shown to inhibit the growth of a variety of cancer tissues. In this study, we investigated the anticancer effect of A. graveolens on the human prostatic carcinoma cell line LNCaP. LNCaP cells were treated with increasing concentrations of an ethanolic extract of A. graveolens ranging from 1000 to 3000 µg/mL, and viability was determined after 24 and 48 h using the XTT cell proliferation assay. The levels of cleaved poly (ADP-ribose) polymerase (PARP), one of the best biomarkers of apoptosis, were analyzed. Finally, quantitative gene expression analysis of vascular endothelial growth factor (VEGF), a critical mediator of angiogenesis, was performed using real-time reverse transcription-polymerase chain reaction. A. graveolens extract inhibited cell viability in both a time- and dose-dependent manner. Data from cleaved PARP assays suggested that A. graveolens caused induction of apoptosis in these cells. Treatment of cells with A. graveolens also resulted in downregulation of VEGF expression. This study showed that the antiproliferative effect exerted by an ethanolic extract of A. graveolens is triggered by induction of apoptosis. We also demonstrated that VEGF expression was downregulated by treatment with A. graveolens extract.

Protective effect of chitosan treatment against acetaminophen-induced hepatotoxicity.

Acetaminophen (APAP) is the most commonly reported toxic ingestion in the world. Severe liver injury resulting from overdose or chronic use of APAP remains a significant clinical problem. In recent years, the mechanisms underlying liver injury caused by APAP have become much better understood. We have studied the protective effect of chitosan supplementation against APAP-induced hepatotoxicity with respect to changes in the levels of total and lipid-bound sialic acid in the serum and in the liver tissue and changes in the activity of diagnostic marker enzymes, lipid peroxidation, and ceruloplasmin oxidase enzyme in normal and experimental groups of rats. During the experimental period, chitosan (200 mg/kg body weight per day) was administered to APAP + chitosan-treated rats by oral gavage. Results showed that treatment with APAP induced a significant increase in the serum alanine aminotransferase and alkaline phosphatase activities, in total and lipid-bound sialic acids levels, and in the liver lipid peroxide content. The administration of chitosan significantly prevented APAP-induced alterations in the levels of diagnostic marker enzymes, total sialic acid, lipid-bound sialic acid, and malondialdehyde in the experimental groups of rats. Furthermore, chitosan administration increased the activity of ceruloplasmin oxidase. In conclusion, our results suggest that chitosan has a protective effect on APAP-induced hepatic injury in rats. The study sheds light on the therapeutic potential of chitosan in an APAP-induced hepatotoxicity model.

Protective effects of the nuclear factor kappa B inhibitor pyrrolidine dithiocarbamate in bladder ischemia-reperfusion injury in rats.

The aim of the present study was to evaluate the protective effects of the NF-кB inhibition with pyrrolidine-dithiocarbamate (PDTC) in ischemia-reperfusion (I/R) injury in the rat bladder. Twenty-four Sprague-Dawley male rats were divided into three groups. Group I; (n = 8) control, group II; (n = 8) I/R group; group III (n = 8) I/R and PDTC treatment. Superoxide dismutase (SOD), catalase (CAT), and gluatathione-S-transferase (GST) enzymes was studied in bladder tissue. Lipid peroxidation (as TBARS) levels in tissue homogenate were measured with thiobarbituric acid reaction. All the slides were stained with NF-кB, p53 and HSP60 immunohistochemistry for detection genome destruction and tissue stress, respectively. Our results show that the mean TBARS levels were significantly higher in group II (p < 0.05). The TBARS levels were significantly decreased in group III compared with the group II (p < 0.05). CAT, SOD and GST activities were decreased in group II, but these enzymes levels were significantly increased in group III according to the group II (p < 0.05). Under microscopic evaluation NF-кB expression increased significantly in group II compared to the group I (p < 0.05) and then decreased in group III (p < 0.05). HSP60 and p53 expression in group II was increased significantly compared with group I. Under microscopic evaluation we detected that HSP60 and p53 expression was increased significantly in group II compared with group I. In group III PDTC administration was decreased the HSP60 and p53 expression, this difference was statistically significant (p < 0.05). The results of the present study have demonstrated that NF-кB inhibition with PDTC protects and provides beneficial effects on ischemia/reperfusion stress related bladder tissue destruction.

Effect of αlipoic acid and silymarin on bladder outlet obstruction.

The aim of the present study was to determine whether the treatment of obstructed rat bladders with αlipoic acid (ALA) and silymarin reverses the biochemical and physiological responses to bladder outlet obstruction (BOO). A total of 32 adult Sprague Dawley rats were divided into four groups (n=8 per group): sham (placebo surgery) animals with no treatment (group 1); control animals with surgically induced BOO (group 2); obstructed rats treated with ALA (group 3); and obstructed rats treated with silymarin (group 4). Histological evaluation, bladder weights, collagen structure, TdT-mediated biotin nick end-labeling (TUNEL), inducible nitric oxide sentase (iNOS) mRNA levels, malondialdehyde (MDA) levels and tumor necrosis factor (TNF) levels were investigated. The ALA-treated group had similar bladder weights, collagen levels and TUNEL positivity and decreased iNOS levels compared with the control group, while the silymarin group exhibited further differences. Serum MDA and TNF-α levels were both decreased in the ALA and silymarin groups. ALA treatment reduced the increased oxidative stress and bladder inflammation caused by BOO and may contribute to the protection of bladder function.

Effects of rotenone on inducible nitric oxide synthase and cyclooxygenase-2 mRNA levels detected by real-time PCR in a rat bladder ischemia/reperfusion model.

We aimed to determine whether rotenone treatment prevents induced ischemia/reperfusion (I/R) damage in rat bladders by detecting inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) levels by real-time PCR (RT-PCR). A total of 18 Sprague-Dawley albino rats were used in this experiment. The experimental groups each consisted of 6 rats and were treated as follows: group I, control; group II, I/R; group III, rotenone + I/R. In the control group, the rat bladders were removed by lower abdominal incision without any procedure. In the I/R group, 1 h prior to the ischemia 1 cc physiological serum was administered and the abdominal aortas were clamped for 1 h to achieve bladder ischemia. Following the ischemia, reperfusion was induced for 1 h and the bladders were removed. In the rotenone + I/R group, the rats were treated with 25 mg/kg rotenone intraperitoneally. The iNOS and COX-2 mRNA levels in each group were detected using RT-PCR. In the I/R group, the COX-2 levels in the bladder tissue were higher compared with the control group (P<0.05). The COX-2 levels in the rotenone-treated group were statistically lower compared with the I/R group (P<0.01). Vascularization and edema were markedly increased in the I/R group. Following rotenone treatment these were abrogated inversely to inflammation. Although iNOS levels were slightly higher in the I/R group compared with the control group, iNOS levels did not decrease and no significant difference was observed between the groups with regard to rotenone treatment (P>0.05). We suggest that rotenone may be used clinically to treat I/R damage due to its diminishing effect on COX-2 levels.

Effects of quercetin on apoptosis, NF-κB and NOS gene expression in renal ischemia/reperfusion injury.

The aim of this study was to investigate the effects of quercetin on nitric oxide synthase (NOS), nuclear factor-κB (NF-κB) and apoptosis in renal ischemia/reperfusion (I/R) injury in rats. A total of 42 Sprague-Dawley rats were divided into three groups. The control, I/R and I/R+quercetin (I/R+Q) groups were treated with quercetin (50 mg/kg intraperitoneal) 1 h prior to the induction of ischemia. Tissue malondialdehyde (MDA) and glutathione (GSH) levels were determined by high-performance liquid chromatography (HPLC). p53, endothelial NOS (eNOS) and NF-κB expression were assessed immunohistochemically, and apoptosis assesment was performed using terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) assay. The mRNA levels of inducible NOS (iNOS) in renal tissue were determined by real-time polymerase chain reaction (RT-PCR). MDA levels were significantly decreased in the quercetin group compared to the I/R group. However, GSH levels were significantly increased with quercetin treatment in the I/R group. Histological results, the number of apoptotic and p53-positive cells, NF-κB and eNOS expression levels were significantly decreased in the quercetin treatment group compared to the I/R group. iNOS gene expression increased in the I/R group, but no significant difference was found between the I/R and quercetin treatment groups. Therefore, quercetin not only has antioxidant and anti-apoptotic activities, but also has an inhibitory effect on eNOS and NF-κB for renal tissue protection during I/R injury in rats. Therefore, quercetin may be a promising renoprotective therapeutic agent.

Lack of association between plasminogen activator inhibitor type-1 (PAI-1) gene 4G/5G polymorphism and osteoarthritis.

This study was conducted in Turkish osteoarthritis patients to determine the frequency of 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene and to examine the role of this polymorphism in osteoarthritis development. Genomic DNA obtained from 200 persons (140 patients with osteoarthritis and 60 healthy controls) was used in the study. DNA was amplified by polymerase chain reaction using 4G allele- and 5G allele-specific primers. Polymerase chain reaction products were assessed with CCD camera by being exposed to 2% agarose gel electrophoresis. No statistically significant difference between the groups with respect to genotype distribution was found (P > 0.05) in the study. The 4G allele frequency was indicated as 44% and 5G allele was as 56% in patients, whereas this was 45-55% in the control group. This study has established that 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene do not play a role in the development of osteoarthritis in the Turkish population.

Anti-apoptotic and antioxidant effect of leptin on CCl4-induced acute liver injury in rats.

The aim of this study is to investigate the effect of leptin in rats on carbon tetrachloride (CCl(4)) induced acute liver damage using immunohistochemical methods for apoptosis and biochemical parameters. In this experimental study, 18 Spraque-Dawley rats were divided into three groups viz; control, CCl(4) and CCl(4)+leptin treatment. 0.8 ml/kg olive oil was administered intraperitoneally (i.p.) to the control group and 0.8 ml/kg CCl(4) (1:1 dissolved in olive oil) was administered i.p. to the CCl(4) and CCl(4)+leptin treatment groups, respectively. After 6 h of administrating CCl(4), CCl(4)+leptin treatment group was given i.p. leptin (10 µg/kg). Twenty-four hours after administrating CCl(4) all of the groups were euthanized. Biochemical assessments were performed using serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), plasma tumor necrosis factor alpha (TNF-α) levels and tissue malondialdehyde (MDA), and TNF-α levels. Histological assessments were then performed using Hematoxylin&Eosin (H&E) staining in light microscope and apoptosis assessment using Terminal Transferase dUTP Nick End Labeling (TUNEL)-staining. Serum AST, ALT, ALP and plasma TNF-α levels, tissue MDA and TNF-α levels had all increased in CCl(4) group, but were found to be significantly decreased in CCl(4)+leptin treatment group. Moreover, TUNEL-positive cell counts in liver had significantly increased in CCl(4) group, but decreased in CCl(4)+leptin treatment group (P < 0.05). The results of our study the biochemical, histological and TUNEL-staining showed that leptin has treatment effects on liver CCl(4) induced injury. It plays a role as a potent free radical scavenger, a powerful antioxidant and it also has anti-apoptotic effects.

Association of angiotensin converting enzyme (ACE) gene I/D polymorphism and polycystic ovary syndrome (PCOS).

This study was conducted in Turkish patients with polycystic ovary syndrome to determine the frequency of I/D polymorphism genotypes of angiotensin converting enzyme gene, and to examine the role of this polymorphism in polycystic ovary syndrome development. Genomic DNA obtained from 200 persons (100 patients with polycystic ovary syndrome and 100 healthy controls) was used in the study. DNA was multiplied by polymerase chain reaction using I and D allele-specific primers. Polymerase chain reaction products were assessed with a charge coupled device (CCD) camera by being exposed to 2% agarose gel electrophoresis. There was statistically significant difference between the groups with respect to genotype distribution (p<0.001). The D allele frequency was indicated as 68% and I allele was as 32% in the patients, whereas it was 51.5-48.5% respectively in the control group. As a result of our study we may assert that angiotensin converting enzyme gene I/D polymorphism DD genotype should be considered as a genetic marker in polycystic ovary syndrome development in this Turkish study population.

Plasminogen activator inhibitor type-1 gene 4G/5G polymorphism and polycystic ovary syndrome.

AIM: This study was conducted in Turkish patients with polycystic ovary syndrome (PCOS) to determine the frequency of 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene and to examine the role of this polymorphism in PCOS development. MATERIALS AND METHODS: Genomic DNA obtained from 200 persons (100 patients with PCOS and 100 healthy controls) was used in the study. DNA was amplified by polymerase chain reaction using 4G and 5G allele-specific primers. Polymerase chain reaction products were assessed with charged-couple device camera after exposing to 2% agarose gel electrophoresis. RESULTS: No statistically significant difference between the groups with respect to genotype distribution was found (p>0.05) in the study. The 4G and 5G allele frequencies were indicated as 51.5% and 48.5% in patients, respectively, whereas this was 51%-49% in the control group. CONCLUSION: Consequently, it has been established by this study that the 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene do not play a role in the development of PCOS in the Turkish population.