Hasan Alacam

Neuroprotective Effect of Humic Acid on Focal Cerebral Ischemia Injury: an Experimental Study in Rats

Stroke is still a major cause of death and permanent neurological disability. As humic acids are well-known antioxidant molecules, the purpose of this study was to investigate the potential neuroprotective effects of humic acid in a focal cerebral ischemia model. Twenty-four rats were divided equally into three groups. A middle cerebral artery occlusion model was performed in this study where control (group II) and humic acid (group III) were administered intraperitoneally following an ischemic experimental procedure. Group I was evaluated as sham. Malondialdehyde (MDA), superoxide dismutase (SOD), and nuclear respiratory factor-1 (NRF-1) levels were analyzed biochemically on the right side of the ischemic cerebral hemisphere, while ischemic histopathological studies were completed on the left side to investigate the antioxidant status. Biochemical results showed that SOD and NRF-1 levels were significantly increased in the humic acid group (III) compared with the control group (II) while MDA levels were significantly decreased. On histopathological examination, cerebral edema, vacuolization, degeneration, and destruction of neural elements were decreased in the humic acid group (III) compared with the control group (II). Cerebral ischemia was attenuated by humic acid administration. These observations indicate that humic acid may have potential as a therapeutic agent in cerebral ischemia by preventing oxidative stress.

Genistein Exerts Neuroprotective Effect on Focal Cerebral Ischemia Injury in Rats

Brain ischemia and treatment are one of the important topics in neurological science. Free oxygen radicals and inflammation formed after ischemia are accepted as the most important causes of damage. Currently, there are studies on many chemopreventive agents to prevent cerebral ischemia damage. Our aim is to research the preventive effect of the active ingredient in genistein, previously unstudied, on oxidative damage in cerebral ischemia. Rats were randomly divided into three groups: control group (no medication or surgical procedure), ischemia group, and artery ischemia + genistein group, sacrificed at 24 h after ischemia. The harvested brain tissue from the right hemisphere was investigated histopathologically and for tissue biochemistry. Superoxide dismutase and nuclear respiratory factor 1 values decreased after ischemia and they increased after genistein treatment, while increased malondialdehyde levels after ischemia reduced after treatment. Apoptosis-related cysteine peptidase caspase-3 and caspase-9 values increased after ischemia, but reduced after treatment. Our study revealed that genistein treatment in cerebral ischemia reduced oxidative stress and neuronal degeneration. We believe that genistein treatment may be an alternative treatment method.

Cord blood and maternal serum neopterin concentrations in patients with pre-eclampsia.

Abstract Background: Pre-eclampsia is a syndrome characterized by hypertension and proteinuria. The aim of this study was to investigate neopterin concentrations in cord blood and maternal serum in patients with pre-eclampsia and a control group. Methods: Cord blood and maternal serum neopterin were measured in 21 patients with pre-eclampsia and in 27 control subjects. Neopterin concentrations were measured by high performance liquid chromatography. Results: Cord blood neopterin concentrations were significantly increased in patients with pre-eclampsia compared to controls (54.3±16.8 vs. 43.4±8.5 nmol/L, p=0.011, respectively). Maternal serum neopterin (257.3±36.8 vs. 150.9±33.8 nmol/L, p<0.001) was also higher in patients with pre-eclampsia. Conclusions: Cord blood and maternal serum neopterin concentrations are higher in patients with pre-eclampsia. Maternal serum neopterin concentrations used may be used as a marker for the early diagnosis of pre-eclampsia. Clin Chem Lab Med 2010;48:1127–31.

The anticancer effects of desferrioxamine on human breast adenocarcinoma and hepatocellular carcinoma cells.

N-myc downstream-regulated gene 1 (NDRG1) is defined as metastasis suppressor and can be downregulated in many types of cancers, and reported to be an indication of tumor progression in hepatocellular carcinomas. Several in-vivo and in-vitro studies have demonstrated that iron chelators such as Desferrioxamine (DFO) and 1-10 Phenanthroline (PHEN) are effective antitumor agents. It is suggested that these chelators deliver their antitumor activity by acting on the NDRG1 gene expression. It remains unclear why NDRG1 gene expression affects the tumors differently, or becomes affected differently. We consider that this different effect might be caused by variants. Based on this information, we developed specific primers and probes for NDRG1 mRNA variants using bioinformatics analysis, and investigated how DFO and PHEN affected the dynamics of NDRG1 variant on the cell lines of Human Breast Adenocarcinoma (MCF-7) and Hepatocellular Carcinoma (HepG2) that demonstrate opposite action for the relationship NDRG1-metastasis. We administrated various doses of DFO and PHEN into the cells to monitor cell vitality and proliferation with Real time Cell Analyzer. We analyzed the gene expression levels of study groups with Quantitative RT-PCR as well as relative gene expression. Variants of NDRG1 mRNA were transcriptionally regulated after HepG2 and MCF-7 cells were treated by iron chelators, resulting in domination of NDRG1 mRNA Variant 1 (V1) in the HepG2 calls and domination of NDRG1 mRNA Variant 2 (V2) in the MCF-7 cells. Anti-proliferative and cytotoxic effects were observed in the MCF-7 cells whereas an increased proliferation was present in the HepG2 cells.

Evaluation of serum endothelial cell specific molecule-1 (endocan) levels as a biomarker in patients with pulmonary thromboembolism.

The aim of this study was to investigate the relationship between pulmonary thromboembolism (PTE) and serum endocan levels. The study included 46 patients with a diagnosis of PTE and control group (25 healthy individuals). Serum endocan levels in all participants’ blood samples were measured. The average age of the individuals was 61.76 ± 16.39 years. There was a significant difference in the serum endocan levels between the patients and those of the control group [321.93 ng/l (111.35–2511.33) and 192.77 ng/l (118.30–309.02), respectively; P < 0.030]. The serum endocan levels in the submassive [469.41 ng/l (258.13–800.54)] and the massive PTE groups [719.18 ng/l (319.84–2511.33)] were statistically higher than those in the control group [192.77 ng/l (118.30–309.02)] (P < 0.001 and P < 0.001, respectively). In addition, there was a statistically significant difference between the serum endocan levels of the nonmassive PTE group [188.57 ng/l (111.35–685.56)] and the submassive PTE group (P < 0.01). The serum endocan levels correlated with the international normalization ratio (INR), right ventricular dilatation (RVD) and SBP (r = 0.418, P = 0.004; r = 0.659, P < 0.001; r = −0.425, P = 0.003, respectively). In conclusion, serum endocan levels can be considered a practicable biomarker to determine the severity of PTEs and follow-up thrombolytic therapy.

Ouabain Targets the Unfolded Protein Response for Selective Killing of HepG2 Cells During Glucose Deprivation

Ouabain is a cardiotonic steroid and specific inhibitor of the Na(+)/K(+)-ATPase. The relationship between ouabain treatment and the unfolded protein response (UPR) in cells is not precisely understood. Therefore, we studied the possible effects of ouabain on proliferation, apoptosis, and the UPR. HepG2 cells were cultured overnight and then treated with various concentrations of ouabain (0.75 to 750 nM) in the absence or presence of 10 mM 2-deoxyglucose (2-DG) for 48 hours. We also used real-time polymerase chain reaction to obtain quantitative measurements of expression levels of Grp78, Grp94, CHOP, MTJ-1, HKII, MDR-1, MRP-1, HO-1, and Par-4. Cell number, viability, and proliferation of HepG2 cells were monitored with a real-time cell analyzer system (xCELLigence). We show that ouabain modulates the UPR transcription program and induces cell death in glucose-deprived tumor cells. Ouabain at all concentrations showed no cytotoxicity whereas all concentrations were very effective under 2-DG stress conditions. Our findings show that disruption of the UPR during glucose deprivation could be an attractive approach for selective cancer cell killing and could provide a chemical basis for developing UPR-targeting drugs against solid tumors. Ouabain use as an adjunct to conventional cancer therapy also warrants vigorous investigation.

The role of asymmetric dimethyl arginine and oxidant/antioxidant system in preeclampsia.

Preeclampsia is a syndrome characterized by hypertension and proteinuria. The aim of this study is to find the relationship between preeclampsia, asymmetric dimethyl arginine (ADMA), and the oxidant/antioxidant system. Twenty-one preeclamptic and 28 normal pregnant women were included in this study. In cord bloods, ADMA and oxidant/antioxidant parameters were measured. Asymmetric dimethyl arginine levels were significantly increased in preeclamptic pregnancies compared to the control group (p = 0.006). The activities of antioxidant enzymes and malondialdehyde levels were increased in the preeclamptic group compared to the control group (p < 0.001, p = 0.022, p < 0.001, p < 0.001, respectively). Development of endothelial dysfunction and oxidative stress may play a role in developing preeclampsia.

Paraoxonase-1 activity determination via paraoxon substrate yields no significant difference in mild hyperhomocysteinemia

Elevated plasma homocystein (Hcy) level has been recognized as an important risk factor for a number of cardiovascular diseases, peripheral arterial occlusive disease and venous thrombosis. A part of Hcy in the organism is turned to homocysteine thiolactone (HcyT) via a ring closure reaction, which gains rate in hyperhomocysteinemia, and in turn undergoes a hydrolytic reaction back to Hcy by paraoxonase enzyme (PON). Since this is a protective reflex action enzyme against hyperhomocysteinemia, we investigated how a mild hyperhomocysteinemic nutritional habit affected serum PON activity in a population-based study. The difference detected via enzymatic activity using the paraoxon substrate was statistically non-significant (p=0.19), suggesting a defective performance to reflect the expected significance. Determination of serum PON activity via substrate paraoxon yielded no significant difference in an acute mild hyperhomocysteinemic diet model in humans.

The evaluation of different treatment protocols for trauma-induced lung injury in rats

BACKGROUND Lung contusion is an important factor that affects mortality and morbidity of lung injury after blunt chest trauma (BCT). The present study aims to evaluate the effectiveness of different treatment regimens on BCT-induced lung injury. METHODS A total of 35 Sprague Dawley rats were divided into five experimental groups (n=7): sham, control; BCT; BCT + MP, BCT group treated with methylprednisolone (MP; 30 mg/kg on first day and 3 mg/kg/d on the following days); BCT + Q, BCT group treated with quercetin (Q; 50 mg/kg/d for seven days); and BCT + MP + Q, BCT group treated with the same doses of MP and Q. Serum Clara Cell Protein-16 (CC-16), thiobarbituric acid reactive substances (TBARS), and superoxide dismutase (SOD) levels were analyzed to determine histopathological changes in the lung tissues. RESULTS Elevated serum CC-16 and TBARS levels and reduced serum SOD levels were found in the BCT group compared to the Sham group. There was a significant change in the serum CC-16 levels in the BCT + MP group compared to the Sham group. Serum TBARS levels were significantly lower in the BCT + MP and BCT + Q group compared to the BCT group. The combined therapy regimen yielded significantly decreased CC-16 levels and increased serum SOD levels compared to the individual treatment groups. Serum TBARS levels did not significantly differ between the BCT + MP + Q group and the other treatment groups. Compared to the BCT + MP + Q group, the BCT + MP group showed significantly lower alveolar edema (AED) and alveolar exudate (AEX) scores, while the BCT + Q group showed significantly lower peribronchial inflammatory cell infiltration (PICI) and AED scores. CONCLUSIONS The combined usage of quercetin and low dose MP treatment after initial high dose MP at the early stage of lung injury after BCT is more effective.

Cytotoxic effect of fluvastatin on MCF-7 cells possibly through a reduction of the mRNA expression levels of SGK1 and CAV1.

Fluvastatin (FLU) prevents the conversion of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) to mevalonic acid by inhibiting HMG-CoA reductase and decreases cholesterol level. Although the effects of FLU treatment on several cancer types through many mechanisms have been identified, its relationship with unfolded protein response and apoptosis has not been clearly understood. In this recent study, we aimed to investigate the cytotoxic effect of Fluvastatin on MCF-7 cells and define the transcriptional regulation of specific genes during the occurrence of this cytotoxic effect. We administered 0.62, 2.5, 5, and 40 μM FLU on MCF-7 cells singly and in combination with 2-deoxyglucose (2-DG), and we monitored cell viability and proliferation for 48 hours using real-time cell analyzer system (xCELLigence). At the same time, we measured the mRNA expression levels of glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein, homologous protein (CHOP), caveolin-1 (CAV1), NDRG1 Variant 1 and Variant 2, HMOX1, SGK1, and prostate apoptosis response-4 (PAR4) genes using quantitative real-time polymerase chain reaction (LightCycler 480 II). We accepted GAPDH gene and control groups as the reference gene and calibrator, respectively. We performed relative gene expression analyses of the study groups using the QIAGEN 2009 Relative Expression Software Tool (REST). FLU revealed an antiproliferative and cytotoxic effect on MCF-7 cells, while causing the transcriptional regulation of many genes. Of these genes, the mRNA expressions of CHOP, heme oxygenase 1 (HMOX1), N-myc downstream-regulated gene 1 (NDRG1) V1, and NDRG1 V2 increased. On the other hand, the mRNA expression levels of SGK1 and CAV1 decreased. The antiproliferative effects of FLU may be related to the decreased expression levels of SGK1 and CAV1.