Ozge Cemiloglu Ulker

Estimation of ochratoxin A in some Turkish populations: An analysis in urine as a simple, sensitive and reliable biomarker

In this study, ochratoxin A (OTA) occurrence and level in human urine samples, collected from four different regions of Turkey was analyzed by NaHCO(3) dilution, immunoaffinity column clean-up and high-performance liquid chromatography with fluorescent detection. For the repeatability of the method, RSD (%) values were found between 3.83 and 8.86, for the accuracy, the recovery values were found between 85.7% and 110.5% and limit of detection and limit of quantification of the method were measured as 0.006 and 0.018 ng mL(-1) respectively. For the analysis, first morning urine samples were collected and the results were adjusted with creatinine levels. From the total collected samples of 233 larger amounts of 83% was contaminated with OTA. Among the calculated to be OTA levels, positive sample rate, average OTA amount and the highest contamination was found in Ankara. (Positive sample rate; 90.1%, average OTA concentration; 14.34 ng g(-1) creatinine and highest OTA value; 75.60 ng g(-1) creatinine). In order to define the exposure profile of OTA in human a questionnaire was conducted among the voluntaries as well. But related to the gender, age, dietary habits, coffee consumption, smoking and alcohol habits of the volunteers, no correlation was found with the OTA exposure.

Serum and BAL cytokine and antioxidant enzyme levels at different stages of pneumoconiosis in coal workers

Coal workers’ pneumoconiosis (CWP) is an occupational pulmonary disease that occurs by chronic inhalation of coal dust. CWP is divided into two stages depending on the extent of the disease, as simple pneumoconiosis (SP) and progressive massive fibrosis (PMF). In the present study, serum and bronchoalveolar lavage (BAL) cytokine (interleukin-1β [IL-1β], IL-6, tumor necrosis factor-α [TNF-α], transforming growth factor-β [TGF-β]) and antioxidant enzymes levels, their relation with the disease severity, and whether they can be considered as biological markers were investigated. Serum and BAL levels of IL-1β, IL-6, and TNF-α were higher in SP and PMF patient groups compared with that in active and retired miner groups. Serum and BAL IL-1β, IL-6, and TNF-α levels were also found to be higher in patients with PMF compared with the SP group. BAL superoxide dismutase (SOD), glutathione peroxidase, and catalase levels and serum SOD level were increased in both patient groups compared with the control group. In addition, mean serum and BAL TGF-β levels were found to be increased in patients with SP compared with PMF group. Based on these results, BAL and serum cytokine and antioxidant enzymes levels were evaluated and discussed as potential biomarkers for different stages of CWP.

Evaluation of auricular lymph node cell lymphocyte proliferation and cytokine production as non-radioactive endpoints during murine contact allergy

The murine local lymph node assay (LLNA) has been developed as a test method to assess allergic contact dermatitis. In spite of the validity of the LLNA, attention was drawn to the two disadvantages: use of radioactivity for in vivo measurement of lymph node cell proliferation ([3H]-thymidine labeling) and the possibility of false positive results caused by non-specific cell activation as a result of inflammatory processes in the skin (irritation). We aimed to investigate the following non-radioactive endpoints of LLNA: 5-bromo-2′-deoxyuridine (BrdU) incorporation ex vivo and in vivo, in vivo and ex vivo cytokine production with or without phytohemagglutinin (PHA) stimulation. Here, 8-12-week-old female BALB/c mice were treated topically with the strong sensitizer 2,4-dinitrochlorobenzene (DNCB) in acetone:olive oil (AOO, 4:1 [v/v]) at levels of 0.025, 0.05, 0.01, or 0.25% (w/v). Ear thickness was also measured to determine the differentiation index (DI) indicating the proportion of non-specific activation due to irritating properties of test compound. At the concentration of 0.05%, stimulation index (SI) value was found to be 3 for DNCB based on in vivo and ex vivo BrdU incorporation. The results of the in vivo and ex vivo non-radioactive LLNA assays were compatible both with each other and with previous radioactive LLNA data. Our results indicate that non-radioactive endpoints may be used as an alternative to the [3H]-thymidine LLNA. The levels of TH1 cytokines (IL-2 and IFNγ) and TH2 cytokines (IL-4 and IL-5) in lymph node cell cultures were significantly (P < 0.01) increased when DNCB was applied at the concentrations of 0.05 and 0.1%, respectively. As the DI was > 1, the applied concentrations of DNCB caused only allergic effect but not any irritant effect. This study reports that the use of these non-radioactive endpoints can assess allergic contact dermatitis caused by chemicals.

Allergenicity evaluation of fragrance mix and its ingredients by using ex vivo local lymph node assay-BrdU endpoints.

The present studies were performed to compare the differences between sensitization potency of fragrance mix and its ingredients (oak moss absolute, isoeugenol, eugenol, cinnamal, hydroxycitronellal, geraniol, cinnamic alcohol, alpha amyl cinnamal), by using ex vivo LLNA-BrdU ELISA. The SI and EC3 values were calculated and potency classification was found for the mixture and for each ingredients. TH1 cytokines (IL-2, IFN-γ) and TH2 cytokines (IL-4, IL-5) releases from lymph node cell culture were also investigated as contact sensitization endpoints. The EC3 values were calculated and the potency of contact sensitization were classified for fragrance mix, oak moss absolute, isoeugenol, eugenol, cinnamal, hydroxycitronellal, geraniol, cinnamic alcohol, alpha amyl cinnamal respectively: 4.4% (moderate), 3.4% (moderate), 0.88% (strong), 16.6% (weak), 1.91% (moderate), 9.77% (moderate), 13.1% (weak), 17.93% (weak), 7.74% (moderate). According to our results it should be concluded that exposure to fragrance mix does not constitute an evidently increased hazard compared to exposure to each of the eight fragrance ingredients separately. Cytokine analyses results indicate that both TH1 and TH2 cytokines are involved in the regulation of murine contact allergy and can be considered as useful endpoints.

Evaluation of non-radioactive endpoints of ex vivo local lymph node assay-BrdU to investigate select contact sensitizers.

The present study sought to verify the utility of the non-radioactive endpoints LLNA BrdU (5-bromo-2′-deoxyuridine) ex vivo incorporation and cytokine release using auricular lymph node cells isolated from BALB/c mice topically treated with a strong (formaldehyde or p-phenylene-diamine [PPD]), moderate sensitizer (cinnamal), or weak sensitizer (eugenol). Stimulation index (SI) and EC3 values were calculated for each agent. Based on the results of ex vivo LLNA-BrdU assays, EC3 values were calculated to be 0.29, 0.09, 1.91, and 16.60% for formaldehyde, PPD, cinnamal, and eugenol, respectively. These results were in good agreement with data from previous standard radioactive LLNA. Cytokine analyses indicated TH1 and TH2 cytokine involvement in the regulation of murine contact allergy and these could be utilized as endpoints in assessments of contact allergy in mice. In conclusion, the current study provided evidence that the non-radioactive endpoint LLNA BrdU ex vivo incorporation could be of use as a viable alternative approach to assess the skin sensitization potential of test compound with respect to improving animal welfare. This is of particular importance in the case of any laboratory where it might be difficult to handle and/or readily employ radioisotopes. Further studies will be required to confirm—across test agents—the reproducibility as well as the limits of utility of this new ex vivo BrdU method.

Correlation of Ochratoxin A Exposure to Urinary Levels of 8-Hydroxydeoxyguanosine and Malondialdehyde in a Turkish Population

Ochratoxin A is one of the most abundant food- contaminating mycotoxins in the world that is immunosuppressive, genotoxic, teratogenic and carcinogenic. Malondialdehyde is a naturally occurring product of lipid peroxidation that is mutagenic and carcinogenic. 8-Hydroxydeoxyguanosine is produced during the interaction of reactive oxygen species and DNA. In this study, Ochratoxin A, malondialdehyde and 8-Hydroxydeoxyguanosine levels of individuals in the study group were measured and results were correlated with each other. Additionally, the correlation of biomarker levels to smoking habit, alcohol and coffee consumption, age and gender of individuals was investigated. As a result of these assessments, a significant correlation was observed between Ochratoxin A exposures and malondialdehyde and 8-Hydroxydeoxyguanosine levels.

Utilization of the ex vivo LLNA: BrdU-ELISA to distinguish the sensitizers from irritants in respect of 3 end points-lymphocyte proliferation, ear swelling, and cytokine profiles.

Dermal exposure to chemicals may result in allergic or irritant contact dermatitis. In this study, we performed ex vivo local lymph node assay: bromodeoxyuridine-enzyme-linked immunosorbent assay (LLNA: BrdU-ELISA) to compare the differences between irritation and sensitization potency of some chemicals in terms of the 3 end points: lymphocyte proliferation, cytokine profiles (interleukin 2 [IL-2], interferon-γ (IFN-γ), IL-4, IL-5, IL-1, and tumor necrosis factor α [TNF-α]), and ear swelling. Different concentrations of the following well-known sensitizers and irritant chemicals were applied to mice: dinitrochlorobenzene, eugenol, isoeugenol, sodium lauryl sulfate (SLS), and croton oil. According to the lymph node results; the auricular lymph node weights and lymph node cell counts increased after application of both sensitizers and irritants in high concentrations. On the other hand, according to lymph node cell proliferation results, there was a 3-fold increase in proliferation of lymph node cells (stimulation index) for sensitizer chemicals and SLS in the applied concentrations; however, there was not a 3-fold increase for croton oil and negative control. The SLS gave a false-positive response. Cytokine analysis demonstrated that 4 cytokines including IL-2, IFN-γ, IL-4, and IL-5 were released in lymph node cell cultures, with a clear dose trend for sensitizers whereas only TNF-α was released in response to irritants. Taken together, our results suggest that the ex vivo LLNA: BrdU-ELISA method can be useful for discriminating irritants and allergens.

Investigation of allergenicity of some cosmetic mixtures by using ex vivo local lymph node assay-BrdU endpoints.

Background: Balsam of Peru and fragrance mix are commonly used in cosmetic products. Allergy to fragrance is the most common cause of cosmetic contact dermatitis. Methods: In the present study, ex vivo local lymph node assay-5-bromo-2′-deoxyuridine (LLNA-BrdU) was used to evaluate the dermal sensitization potential of these cosmetic mixtures. The stimulation index values and estimated concentration (EC3) values were calculated and the potency classification was found for each mixture. At the same time, in order to measure the irritant effect without having to use additional animals, a combination of ex vivo LLNA-BrdU and the irritancy assay was conducted. Results: Th1 [interleukin (IL)-2, interferon-γ] and Th2 cytokine (IL-4, IL-5) releases from lymph node cell culture were investigated as non-radioactive endpoints. According to the results of ex vivo LLNA-BrdU assays, EC3 values were found to be 3.09% (moderate) for balsam of Peru and 4.44% (moderate) for fragrance mix. Cytokine analysis results indicate that both Th1 and Th2 cytokines are involved in the regulation of murine contact allergy and can be considered as useful endpoints. Conclusion: In conclusion, according to our results, fragrance mix and balsam of Peru can be considered as moderate sensitizers; however, in high concentrations, both of them have irritation properties. The cytokines investigated can be considered as the endpoints of the ex vivo LLNA-BrdU assay.

The fruit extract of Berberis crataegina DC: exerts potent antioxidant activity and protects DNA integrity

BackgroundDried fruits of Berberis crataegina (Berberidaceae) have been frequently consumed as food garniture in Turkish cuisine, while its fruit paste has been used to increase stamina and in particular to prevent from cardiovascular dysfunctions in Northeastern Black Sea region of Turkey. This study investigated this folkloric information in order to explain the claimed healing effects as well as to evaluate possible risks.MethodsTotal phenolic, flavonoid and proanthocyanidin contents and antioxidant capacity of the methanolic fruit extract were evaluated through several in vitro assays. The cytotoxic and genotoxic effects of B. crataegina fruit extract were also assessed in both cervical cancer cell line (HeLa) and human peripheral blood lymphocytes.ResultsThe extract showed protective effects against ferric-induced oxidative stress and had a relatively good antioxidant activity. It also ameliorated the H2O2 mediated DNA damage in lymphocytes, suggesting the protective effect against oxidative DNA damage.ConclusionThe methanolic extract of B. crataegina fruits may be a potential antioxidant nutrient and also may exert a protective role against lipid peroxidation as well as oxidative DNA damage.

Investigation of hypersensitivity potential of diacetyl by determining cytokine profiles

Exposure to chemicals in workplace settings leads to significant occupational diseases related to hypersensitivity reactions. In recent years, diacetyl which is used as a food additive is thought to be as an important hazard due to its sensitization potency for worker’s health. Therefore, in this study, we aimed to investigate the sensitization potency of diacetyl for the purpose of its hazard evaluation. Nonradioactive ex vivo local lymph node assay: BrdU-enzyme-linked immunosorbent assay (ELISA) method with short-term and long-term exposure protocols were conducted based on animal welfare principles. As end points, lymphocyte proliferation, cytokine releases, and total serum IgE levels were measured by ELISA method. After short-term dermal exposure to diacetyl, primary Th1 cytokine interleukin-2 (IL-2) and Th2 cytokines IL-4 and IL-13 levels were significantly increased relatively to vehicle control, whereas such significant increases were not observed in long-term exposure. According to our measurements of IgE levels after long-term dermal exposures to chemicals, diacetyl led to significant increase. In conclusion, the findings that showed significant increases in IL-13 and total serum IgE levels induced with diacetyl can be relevant to respiratory sensitizing hazard of this chemical.