Biochemical aspects of olfactory mechanisms

Electron microscopic examination of intact olfactory epithelia in different animals yielded the same pattern of morphological organisation of the cellular organelles. Attempts have been made to isolate cilia from the primary olfactory neurons in order to study the molecular mechanisms involved in olfaction. The results indicated that the purity and quantity of the ciliary preparation obtained by current membrane fractionation methods was not suitable for biochemical investigations. Also ciliated columnar respiratory cells have been found in the olfactory mucosa of sheep. This feature presents difficulties for future neuronal membrane isolation attempts. Two basic problems in olfaction, coding and transduction have also been studied in this thesis. Chemical modification methods have been employed to study the coding mechanism at the peripheral level in the frog. With a variety of group specific protein reagents, proteolytic enzymes and photoaffinity odorants, it has been possible to specifically modify olfactory receptors. The non-penetrant sulfhydryl reagent, mersalyl irreversibly inhibited the EOG responses to odorants. This effect was found to be prevented by n-amyl acetate. Application of this odorant in solution before, during and after the treatment of the mucosa with mersalyl resulted not only in protection of the responses to n-amyl acetate, but also to other odorants as well. Responses to some odorants did not show any recovery providing evidence for the occurrence of different receptor sites. To investigate the possible involvement of cAMP in the olfactory transduction mechanism, a number of chemicals were applied to the frog's olfactory mucosa. Phosphodiesterase inhibitors and membrane- permeable cyclic AMP derivatives decreased the EOG responses. One contribution of this thesis to the field of olfaction has been to show that olfaction has biochemical features common to other receptor systems

Regulation of Shootin1 Gene Expression Involves NGF-induced Alternative Splicing during Neuronal Differentiation of PC12 Cells

Shootin1 is a protein involved in neuronal polarization, and has been shown to be a key molecule for the positive/negative feedback loop for axon induction required during neuronal symmetry breaking. To better understand the molecular basis of shootin1 dynamics, we analysed the regulatory pathways and the expressional status of shootin1 gene during NGF-induced neuronal differentiation. We demonstrated that the isoform-1 and isoform-2 of shootin1 is differentially expressed during neuronal differentiation. By blocking individual downstream pathways of NGF signalling, we found that PI3K/Akt pathway plays a major role in the expression of shootin1 isoform-2. Western blot and RT-PCR results showed that the isoform-1 of shootin1 is constitutively expressed, while the isoform-2 is expressed in a manner that is strictly dependent on NGF-stimulation. Isoform-specific RT-PCR results demonstrated that the differential expression of the isoform-1 and isoform-2 of shootin1 is a consequence of alternative splicing of shootin1 pre-mRNA, in response to NGF-signalling. Collectively these findings provide the first information on the molecular mechanisms regulating the expression of shootin1 gene and represent the first example of NGF-induced alternative splicing process that has a regulatory role in neuritogenesis

The Effects of Cisplatin on MTOR, AKT, CCND1 and STAT3 mRNA Expressions on HeLa Cells

WOS: 000505071500002Objective: Cervical cancer is one of the most tumors seen in womans and second leading cause of cancer death in women. Cisplatin (CDDP), a platin based compound, is the most important chemotherapeutic agent and effectively used for the treatment of sarcomas and solid tumors. It binds to the DNA with crosslinks and leads inhibition of replication, causing DNA damage. As a result of this action, apoptotic pathways are induced and cell death occurs. In our study, we aimed to investigate the effect of CDDP on relative mRNA expression of mTOR, AKT, Cyclin D1 (CCND1) ve STAT3 on cervical cancer cell line. Method: In this study, HeLa cells were treated with different concentrations of CDDP at 24 and 48 hours. Cell viability was determined by XTT method. Moreover, after treatment with selected doses of cisplatin, quantitative mRNA expression of mTOR, AKT, CCND1 and STAT-3 genes was analyzed using Real-Time PCR. Results: IC50 concentration of CDDP was found to be about 60 mu M for 24h and 8 mu M for 48h treatment. Moreover, all analyzed genes' expression was shown to diminish only after 24 h treatment. On the other hand, no statistically significant change was found after 48 h cisplatin exposure with respect to quantitative mRNA expression. Conclusion: In summary, different mRNA expression pattern was found after CDDP treatment regarding to exposure time. Our study has been contributed the literature in terms of detecting the effect of conventional chemotherapeutic CDDP on cell survival pathways

Effects of ketamine, propofol, and ketofol on proinflammatory cytokines and markers of oxidative stress in a rat model of endotoxemia-induced acute lung injury

Intravenous lipopolysaccharide (LPS) leads to acute lung injury (ALI) in rats. The purpose of this study was to examine the anti-inflammatory and antioxidant efficacy of ketamine, propofol, and ketofol in a rat model of ALI. We induced ALI in rats via intravenous injection of LPS (15 mg kg(-1)). The animals were randomly separated into five groups: control, LPS only, LPS + ketamine (10 mg.kg(-1).h(-1)), LPS + propofol (10 mg.kg(-1).h(-1)), LPS + ketofol (5 mg.kg(-1).h(-1) ketamine + 5 mg.kg(-1).h(-1) propofol). LPS resulted in an increase in the release of pro-inflammatory cytokines, mRNA expression related with inflammation, production of nitric oxide, and lipid peroxidation. Ketamine prevented the increase in markers of oxidative stress and inflammation mediators, both in plasma and lung tissue. Propofol decreased the levels of cytokines in plasma and lung tissue, whereas it had no effect on the IL-1-beta level in lung tissue. Ketamine downregulated mediators of lung tissue inflammation and reduced the level of circulating cytokines and protected lung tissue against lipid peroxidation. Ketofol decreased the level of INF-alpha and IL-1 beta in plasma, as well as expression of cyclooxygenase-2 mRNA and the nitrate/nitrite level in lung tissue. The results of this investigation support the hypothesis that ketamine may be effective in preventing ALI

SISTER-CHROMATID EXCHANGE INDUCING EFFECT OF SMOKELESS TOBACCO USING ON T-LYMPHOCYTE CHROMOSOMES

A kind of a smokeless tobacco (Maras powder) is widely used instead of cigarettes in the South Eastern region of Turkey. In this study we investigated the sister-chromatid exchange (SCE) inducing effect of this powder on the chromosomes of its users compared with smokers and nonsmokers using standard cell culture methods and SCE staining techniques. Average SCE per metaphase and total SCEs increased significantly among both smokeless tobacco users and smokers compared to nonsmokers (p < 0.01). However, the effect is significantly lower in smokeless tobacco users than in smokers (p < 0.05)

Investigation of Ocular Neovascularization-Related Genes and Oxidative Stress in Diabetic Rat Eye Tissues After Resveratrol Treatment

Changes in vascular endothelial growth factor (VEGF), angiotensin-converting enzyme (ACE), matrix metalloproteinase (MMP)-9, and endothelial nitric oxide synthase (eNOS) mRNA expression profiles and oxidative stress in the eye tissue microenviroment may have important roles in ocular neovascularization and permeability in proliferative diabetic retinopathy. The present study investigated the effects of resveratrol (RSV) treatment on the mRNA expression profile of VEGF, ACE, MMP-9, and eNOS, which are associated with vascular neovascularization, and glutathione, protein carbonyl, and nitrite-nitrate levels, which are markers of oxidative stress in eyes of diabetic rats. Twenty-four Wistar albino male rats were divided into four groups. After diabetes induction with streptozotocin (10 mg/kg/day) RSV was administered to the RSV and diabetes mellitus (DM) + RSV groups for 4 weeks. The mRNA levels were measured by quantitative real-time polymerase chain reaction assay, and biochemical estimations were determined with spectrophotometric assays in eye homogenates. The mRNA expression levels of VEGF, ACE, and MMP-9 were increased in the DM group compared with the control group, and RSV treatment decreased their mRNA levels. Expression of eNOS mRNA was increased in the RSV and DM groups and decreased in the DM + RSV group. Nitrite-nitrate levels and protein carbonyl content were increased and glutathione levels were decreased in the DM group compared with controls. Consequently, these data suggest that RSV suppressed the expression of eNOS, which is actively involved in the inflammation and healing process in chronic diabetes. Although oxidative stress was increased in eye tissue from diabetic rats, mRNA levels of VEGF, MMP-9, and ACE genes associated with vascular remodeling did not change in diabetic eyes