Mutation Status and Immunohistochemical Correlation of EGFR Mutations in Gastrointestinal Stromal Tumors.

Being one of the leading causes of cancer deaths worldwide and their resistance to conventional treatment methods, made gastrointestinal stromal tumors (GISTs) one of the hot topics in medical research areas in the past decade. To investigate molecular alterations underlying the tumor is of great importance to be able to develop new, targeted treatment options. In this study, GIST samples obtained from 40 Turkish patients were analyzed for actionable epidermal growth factor receptor (EGFR) mutations that are related to treatment regimes in non small cell lung cancer (NSCLC) to understand whether EGFR expression is altered in GISTs. Established alterations in EGFR can make the use of tyrosine kinase inhibitors possible, which are currently used in cancer therapy, especially in NSCLC. Our results indicated that EGFR mutations are rare in GISTs. Further research is needed to sequence whole coding regions of the gene to investigate new actionable mutations in EGFR in an increased sample size.

Investigation of KCNQ1 polymorphisms as biomarkers for cardiovascular diseases in the Turkish Cypriots for establishing preventative medical measures.

Potassium channels are important in transmitting electrical signals through potassium ions transport. These potassium channels are made from signals encoded by KCNQ1 gene. KCNQ1 polymorphisms were associated with many diseases, including many metabolic and cardiovascular diseases and therefore they can be employed as biomarkers. In this study we aimed to investigate KCNQ1 polymorphisms in the Turkish Cypriot population to reveal the allele frequencies specific for this population and use these polymorphisms as biomarkers to develop preventative medical measures. The genotypes of KCNQ1 polymorphisms (rs231361, rs231359, rs151290, rs2283228, rs2237895, rs2237896) were investigated for the first time in Turkish Cypriot population. The correlation between genotypes of these polymorphisms and plasma lipid levels in this population was also explored. The results of this study showed that there was significant differences of the allele frequencies of between rs2283228 allele of C and rs2237896 (P > 0.05) in Turkish Cypriot population. There was no association between the genotypes of the six polymorphisms and the lipid metabolism. This study is the first genetic epidemiology study that investigated the allelic frequencies of KCNQ1 polymorphisms associated with metabolic syndromes as well as cardiovascular diseases. This study proves to be crucial since the etiologic determinants and molecular pathology of cardiovascular diseases have not yet clearly understood. This study showed that genome wide association studies should be designed for preventative medicine in the Turkish Cypriot population.

Bone mineral density measurements in otosclerosis using dual-energy X-ray absorptiometry.

OBJECTIVES: This study aims to evaluate patients with otosclerosis with respect to bone mineral density (BMD) at different regions of interest (ROI), using dual-energy X-ray absorptiometry (DXA). DESIGN: Cross-sectional controlled study. SETTING: Tertiary referral hospital. PARTICIPANTS: The patients with a definite diagnosis of otosclerosis confirmed intra-operatively were defined as the study group (n = 30). The control group consisted of volunteer, healthy subjects with normal hearing (n = 43). MAIN OUTCOME MEASURES: Following an audiometric evaluation, a venous blood sample was obtained and a single BMD measurement using DXA was applied to each participant. RESULTS: The mean BMD, T and Z scores were higher in the otosclerosis group than in the control group in all the regions considered, but not significantly; only the L2-L3 lateral BMD and its T and Z scores were significant (P = 0.036, P = 0.029 and P = 0.036, respectively). CONCLUSION: This study shows that the BMD does not decrease in the presence of otosclerosis despite its genetic and metabolic relevance with osteoporosis. Concerning the L2-L3 lateral BMD measurements, the BMD increased in otosclerosis.

Evaluation of intraabdominal adhesion generating potentials of ankaferd and calcium alginate used as hemostatic agents.

AIM: The present study aimed to evaluate intra-abdominal adhesion generating potential of Ankaferd Blood Stopper (ABS), which was used as postoperative hemostatic agent in the rats that underwent surgery, in comparison with Ca-alginate. MATERIAL AND METHOD: Totally, 30 rats were randomized into 4 groups. In the control group, 1x1 cm peritoneum was removed from the right lower quadrant after cecal abrasion. In the other two study groups, the same procedure was performed after Ankaferd Blood Stopper and Ca-alginate application respectively. RESULTS were evaluated both histopathologically and by adhesion scoring methods. All results underwent statistical analysis. RESULTS: Comparing overall results, no statistically significant difference was found between the sham, control, ABS and Ca-alginate groups (p = 0.099). Paired group comparisons revealed no statistically significant difference between the sham group and the control, ABS, and Ca-alginate groups (p = 0.222, p = 0.222, and p = 0.833 respectively). It was observed that there was no statistically significant difference between the control and ABS groups (p = 0.505), but there was a statistically significant difference between the control and Ca-alginate groups with Bonferroni correction (p = 0.028). Histopathological examination revealed no statistical difference between the groups. CONCLUSION: In conclusion, intra-abdominal adhesion generating potentials of Ca-alginate and ABS were experimentally evaluated and macroscopic and microscopic comparisons revealed no significant difference between sham, control, Ca-alginate, and ABS groups (Fig. 8, Ref. 36). Text in PDF www.elis.sk. agent.

Diffusion tensor magnetic resonance imaging in patients with multiple sclerosis and its relationship with disability.

Multiple sclerosis is a chronic inflammatory demyelinating disease of the central nervous system. Diffusion tensor magnetic resonance imaging (DTI) can yield important information on the in vivo pathological processes affecting water diffusion. The aim of this study was to quantitatively define water diffusion in normal-appearing white matter (NAWM) distant from the plaque, in the plaque, and around the plaque, and to investigate the correlation of these changes with clinical disability. Conventional MRI and DTI scans were conducted in 30 patients with MS and 15 healthy individuals. Fractional anisotropy maps and visible diffusion coefficients were created and integrated with T2-weighted images. Regions of interest (ROIs) were placed on the plaques on the same side, white matter around the plaques and NAWM on the opposite side. Only the white matter of healthy individuals in the control group, and FA and ADC values were obtained for comparison. The highest FA and lowest ADC were detected in the control group at the periventricular region, cerebellar peduncle and at all ROIs irrespective of location. There was a significant difference in comparison to the control group at all ROIs in patients with MS (p < 0.001 for all comparisons). No significant correlation between diffusion parameters and expanded disability state scale (EDSS) scores was found in patients with MS. DTI may provide more accurate information on the damage due to the illness, compared to T2A sequences, but this damage may not be correlated with the clinical disability measured by EDSS score.

Comparison of double inversion recovery and conventional magnetic resonance brain imaging in patients with multiple sclerosis and relations with disease disability.

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system, predominantly affecting the white matter, but also the grey matter. Aim of this study was to detect MS lesions with double inversion recovery (DIR), fluid-attenuated inversion recovery (FLAIR) and T2-weighted magnetic resonance (MR) techniques and determine the sensitivity of these techniques, and the correlation between the number of lesions and expanded disability state scale (EDSS) scores. Thirty-four patients with MS (20 females and 14 males) were included in this study. DIR and conventional MR (T2-A, FLAIR) sequences were obtained. Lesions were counted and classified as belonging to one of seven anatomical regions: cortical, juxtacortical, deep grey matter, deep white matter, mixed white matter-grey matter, periventricular white matter and infratentorial. The correlation between lesion number and EDSS scores was investigated. DIR images showed more intracortical and mixed white matter-grey matter lesions in comparison with both FLAIR and T2 sequences (p=0, p=0 respectively). There was a significant difference between mean lesion numbers at the juxtacortical region, obtained with DIR and T2-weighted images (p = 0.002). The total number of lesions obtained with all methods was similar. DIR brain imaging had the highest sensitivity in the detection of cortical and mixed white matter - grey matter lesions, compared with FLAIR and T2 sequences. In addition, the lesions obtained with DIR images were more easily visualized.

Determinants of the intention of preconception care use: lessons from a multi-ethnic urban population in the Netherlands.

OBJECTIVES: To investigate the determinants of the intention of preconception care use of women in a multi-ethnic urban population. METHODS: The ASE-model-a health behaviour model-was used as an explanatory framework. A representative sample was taken from the municipal population registers of two districts in Rotterdam, the Netherlands, 2009-2010. 3,225 women (aged 15-60 years) received a questionnaire, which was returned by 631: 133 Dutch, 157 Turkish and Moroccan, and 341 Surinamese and Antillean. Descriptive, univariate and multivariate analyses were performed. RESULTS: The multiple logistic analyses showed that intention to attend preconception care was significantly higher in women with a Turkish and Moroccan background (ß 1.02, P = 0.006), a higher maternal age (ß 0.04, P = 0.008) and a positive attitude (ß 0.50, P < 0.001). Having no relationship (ß -1.16, P = 0.004), multiparity with previous adverse perinatal outcome (ß -1.32, P = 0.001), a high educational level (ß -1.23, P = 0.03), having paid work (ß -0.72, P = 0.01) and experienced barriers level (ß -0.15, P = 0.003) were associated with less intention to use preconception care. CONCLUSIONS: Modifiable determinants as attitude and barriers can be addressed to enhance preconception care attendance.

The levels of nitric oxide and asymmetric dimethylarginine in the rat endometriosis model.

AIM: To investigate the levels of nitric oxide (NO) and asymmetric dimethylarginine (ADMA) in all the rat endometriosis models. MATERIAL & METHODS: Forty-one rats with endometriotic implants were divided into four groups (1 to 4) and administered infliximab, etanercept, letrozole and control, respectively. There were 11 rats in group 5 (normal). The size of implants, plasma ADMA and nitrate/nitrite (NO(x) ) levels and histological score were assessed. RESULTS: In groups 1, 2 and 3, plasma ADMA levels were higher than groups 4 and 5, 296.8 ± 66.2, 285.9 ± 35.7, 200.3 ± 41.0, 125.3 ± 16.7, 111.3 ± 6.5 µmol/L, respectively, while NO(x) levels were lower than groups of control and normal 19.6 ± 3.8, 19.8 ± 4.4, 39.3 ± 6.1, 80.5 ± 5.3, and 91.1 ± 5.0 µmol/L, respectively. CONCLUSIONS: Infliximab, etanercept and letrozole have regressed endometriotic implants, decreased plasma NO(x) levels, and increased plasma ADMA levels.

Effects of citicoline used alone and in combination with mild hypothermia on apoptosis induced by focal cerebral ischemia in rats.

The effects of citicoline used either alone or in combination with hypothermia on the suppression of apoptotic processes after transient focal cerebral ischemia were investigated. Middle cerebral artery occlusion (MCAo) was performed for 2 hours on Sprague-Dawley (SD) rats using intraluminal thread insertion. The treatment groups were as follows: Group 1, sham-operated; Group 2, saline; Group 3, citicoline (400mg/kg intraperitoneal.); Group 4, hypothermia (34+/-1 degrees C); Group 5, citicoline+hypothermia. All rats were reperfused for 24 hours, and after sacrifice and transcardiac perfusion, immunohistochemical studies were performed for markers of apoptosis. In Group 2, the Bcl-2 immunostaining score (mean+/-standard deviation, 0.71+/-0.75) was lower compared to Groups 3, 4 and 5 (2.33+/-0.81; 3.00+/-0.00; 2.20+/-0.83; p<0.05). There was higher expression of caspase-3 proteins in Group 2 (2.28+/-0.95) compared to Group 5 (1.50+/-0.83; p<0.05). Bax proteins were also increased in Group 2 (1.85+/-1.06) compared to Group 5 (0.40+/-0.54) and in Group 4 (2.00+/-0.00) compared to Group 5 (0.40+/-0.54; p<0.05). Significant differences in caspase-9 immunostaining scores were found in Group 2 (2.29+/-0.96) compared to Group 5 (0.20+/-0.44) (p<0.05); Group 3 (1.00+/-0.70) compared to Group 5 (0.20+/-0.44; p<0.05); and Group 4 (3.00+/-0.00; p<0.05) compared to Group 5 (0.40+/-0.54; p<0.05). Thus by suppressing apoptotic processes citicoline with hypothermia is more effective than either used alone in ameliorating cerebral damage after transient focal ischemia.

Cyclooxygenase-2 expression in astrocytes and microglia in human oligodendroglioma and astrocytoma.

Cyclooxygenases (cox) are potent mediators of inflammation and two cox-isoenzymes, cox-1, cox-2, are described to date. Cox-2 is cytokine-inducible in inflammatory cells and enhanced cox-2 expression has been attributed a key role in the development of edema and immunomodulation in pathologically altered brain tissues. In normal cerebral cortex cox-2 is present only in neurons, but not in the glial or vascular endothelial cells. The function of microglia in glioma biology is unclear. Microglia have both neurotrophic and neurotoxic functions and have been shown to release a variety of cytokines. Our preliminary results showed that the expression pattern of cox-2 is predominantly neuronal although glial expression was observed with the correlation of high malignancy. In this study we aimed to assess the phenotypes (astrocyte, microglia) of the cox-2-expressing glial cells in various types of human gliomas and to compare their expression patterns. For this purpose we employed dual immunohistochemistry for cox-2 and GFAP (astrocyte) or LCA-MAC (microglia-macrophage) in archival formalin-fixed, paraffin embedded human tissue diagnosed as oligodendroglioma and/or astrocytoma. The results showed that cox-2 immunoreactivity is up-regulated in the neurons according to the tumor grade. Most of the cox-2 immunoreactive glia were GFAP-positive in anaplastic oligodendrogliomas and at lesser extend in glioblastomas. Cox-2 and LCA co-localization was detected in more glial cells in glioblastomas. It may be speculated that the induction of cox-2 in microglia may contribute to the deleterious effects of prostanoids in cerebral edema formation during the progression of oligodendrogliomas. The detection of cox-2 in astrocytes surrounding the necrotic areas might be important to develop new strategies, such as the usage of cox-2 inhibitors combine with chemotherapy and radiotherapy in the treatment of glioma patients.

Immunohistochemical detection of p53 protein in basal cell skin cancer after microwave-assisted antigen retrieval.

p53 is the most frequently altered tumor-suppressor gene in skin cancer. In normal tissues the p53 protein (wild type) has a very short half-life and it is not detectable immunohistochemically. In contrast, the mutant p53 protein has an extended half-life in tumor cells and can be detected by immunohistochemical methods. p53 is widely used as an indicator of tumor aggression and progression. Fixation methods especially formaldehyde based fixation may mask the immunohistochemical detection of p53 protein but antigen retrieval methods enhance the inmmunohistochemical detection of p53 protein by remodification of protein structure. This study was designed to evaluate the efficacy of different fixatives, of microwaving and microwave pretreatment method to retrieve p53 immunoreactivity in paraffin-embedded non-lesioned (adjacent normal tissue) human skin samples or pathological human skin samples diagnosed as basal cell carcinoma. The samples were fixed at RT and/or in microwave oven either in neutral buffered formalin or alcohol for different time periods. For antigen retrieval, the sections were irradiated in a microwave oven for 5 cycles in 10 mM citrate buffer (pH 6.00). In this study the effects of six different fixation methods on the immunohistochemical staining have been investigated in basal cell tumor specimens. The application of antigen retrieval method was also examined and compared. Optimal results were obtained using samples fixed in alcohol either at room temperature (24 h) or in microwave oven.

Extended pedigree with multiple cases of XX sex reversal in the absence of SRY and of a mutation at the SOX9 locus.

It is well established that testicular differentiation of the human embryonic gonad depends on the action of the Y-chromosomal gene SRY. However, exceptional cases such as SRY-negative cases of 46,XX testicular disorder of sexual development (DSD), and of 46,XX ovotesticular DSD document that testicular tissue can develop in the absence of the SRY gene. These SRY-negative XX sex reversal cases are very rare and usually sporadic, but a few familial cases have been reported. We present a large, consanguineous family with nine affected individuals with phenotypes ranging from 46,XX testicular DSD to 46,XX ovotesticular DSD, with predominance of male characteristics. Absence of SRY in peripheral blood was documented by fluorescence in situ hybridization (FISH) and PCR analysis in all nine affected individuals, and by FISH analysis on gonadal sections with testicular tissue in four affected individuals. By quantitative PCR, a duplication of the SOX9 gene was excluded. In addition, as linkage analysis showed that the nine affected members of the family do not share a common SOX9 haplotype, any mutation at the SOX9 locus could be ruled out. Together, these findings implicate a mutation at a sex-determining locus other than SRY and SOX9 as the cause for the XX sex reversal trait in this family.

A simple and rapid microwave-assisted hematoxylin and eosin staining method using 1,1,1 trichloroethane as a dewaxing and a clearing agent.

The use and practicability of microwave-assisted staining procedures in routine histopathology has been well established for more than 17 years. In the study reported here, we aimed to examine an alternative approach that would shorten the duration of dewaxing and clearing steps of hematoxylin and eosin (H & E) staining of paraffin sections by using a microwave oven. Although xylene is one of the most popular dewaxing and clearing agents, its flammability restricts its use in a microwave oven; thus we preferred 1,1,1 trichloroethane, which is not flammable, as the dewaxing and clearing agent in the present study. In Group I and Group II (control groups), intestine was processed with xylene and 1,1,1 trichloroethane, respectively. The sections were then stained with H & E according to the conventional staining protocol at room temperature and subdivided into two groups according to the duration of dewaxing and clearing in xylene. In Groups III and IV (experimental groups) similar tissues were processed with xylene and 1,1,1 trichloroethane, respectively; however, sections from these groups were divided into four subgroups to study the period required for dewaxing and clearing in 1,1,1 trichloroethane, then stained with H & E in the microwave oven at 360 W for 30 sec. Our conventional H & E staining procedure, which includes dewaxing, staining and clearing of sections, requires approximately 90 min, while our method using 1,1,1 trichloroethane and microwave heating required only 2 min. Our alternative method for H & E staining not only reduced the procedure time significantly, but also yielded staining quality equal or superior to those stained the conventional way. Our results suggest that 1,1,1 trichloroethane can be used effectively and safely as a dewaxing and clearing agent for H & E staining in a microwave oven.

Rapid polymerisation with microwave irradiation for transmission electron microscopy.

Successful results of microwave polymerisation of different epoxy formulations have been reported in the literature. The present study was intended to shorten the time needed for polymerisation of epoxy resin by the use of a microwave technique. A standard double fixation and tissue processing was applied to samples of rat kidney tissue. Tissue samples from the control group were polymerised in a conventional oven at 60 degrees C for 48 h, while tissue from the experimental group was irradiated in a microwave oven, initially at 900 W for 10 min and then at 360 W for another 100 min. During this irradiation, the sealed BEEM capsules were submerged in a water bath, so that the temperature rise was uniform and constant. This resulted in a homogeneous and rapid polymerisation. The cutting properties of the blocks in both groups were similar and no noticeable difference in the quality of the sections was evident when evaluated with TEM. The results showed that the use of a microwave oven reduced the time needed for the polymerisation of Epon blocks without any loss in quality.

Bias and precision of cholesterol analysis by physician's office analyzers.

We studied the bias and precision of serum cholesterol analysis by physician's office analyzers. Total imprecision (CV range, %) for analysis of serum pools was: Abbott Vision 1.5%-1.9%; Ames Seralyzer 3.9%-4.5%; BMD Reflotron 2.3%-3.8%; Chrometrics Cholesterol Test System 2.3%-2.8%; Kodak DT-60 1.6%-2.2%. The Ames Seralyzer exhibited an excessive between-run component of variation. We collected, from 109 volunteers, samples of venous serum, heparin-treated whole blood, heparin-treated plasma, and fingerstick whole blood, and analyzed each type (where possible) with each system; serum was also analyzed in duplicate (by a proposed Reference Method) at the Centers for Disease Control. For assays with serum, the BMD Reflotron and Kodak DT-60 exhibited negative bias. All systems gave lower results for plasma and whole blood than for serum. All systems except the Kodak DT-60 were less precise for analysis of patients' sera than for analysis of serum pools; between-specimen variables may influence the results of these systems.