Daily salivary cortisol profile: Insights from the Croatian Late Adolescence Stress Study (CLASS).

INTRODUCTION: The aim of the study was to examine basal hypothalamic-pituitary-adrenal (HPA) axis activity and to determine associations of various covariates (gender, sleep-wake rhythm, demographic, academic, life style and health-related characteristics) with altered daily salivary cortisol profiles in late adolescence. MATERIALS AND METHODS: The total analytic sample consisted of 903 Croatian secondary school students aged 18 - 21 years (median 19 years). Salivary cortisol was sampled at home at three time points over the course of one week and its concentrations were measured by using the enzyme immunoassay. RESULTS: In comparison to males, female students had a higher cortisol awakening response (CAR) (median 4.69, IQR 10.46 and median 3.03, IQR 8.94, respectively; P < 0.001), a steeper ("healthier") diurnal cortisol slope (DCS) (median 0.51, IQR 0.55 and median 0.44, IQR 0.51, respectively; P = 0.001), and a greater area under curve with respect to ground (AUCG) (median 206.79, IQR 111.78 and median 191.46, IQR 104.18, respectively; P < 0.001). Those students who woke-up earlier and were awake longer, had a higher CAR (P < 0.001), a flatter ("less healthy") DCS (P < 0.001), and a greater AUCG (P < 0.001), than students who woke-up later and were awake shorter. Less consistent but still significant predictors of salivary cortisol indexes were age, school behaviour, friendship, diet healthiness and drug abuse. CONCLUSION: Gender and sleep-wake up rhythm were major determinants of the altered daily salivary cortisol profiles in late adolescence. The predictive power of other covariates, although less clear, has a potential for identifying vulnerable subgroups such as male drug users and females without a best friend.

Prevalence and genotyping of the human papillomavirus in the cervical specimens among women of Southern Croatia (Dalmatia County).

AIM: The aim of this study was to evaluate the prevalence of HPV types in the female population with either normal or abnormal Pap smears in Southern Croatia (Dalmatia County) and to evaluate the distribution of HPV types and their association with different cytological findings. METHODS: During one year period, we routinely screened 1,160 women for HR HPV at the Educational Public Health Institute of Split and Dalmatia County, by using Hybrid Capture 2 HPV DNA test. Only HR HPV positive specimens determined by HC2 were further genotyped by polymerase chain reaction (PCR) based assay at the Department of Pathology and Forensic Medicine, Split. RESULTS: Out of the total number of tested women, 406 (35%) of them were positive to HR HPV. The following frequency was observed: HPV 16 in 10.8%, HPV 18 in 7.8%, HPV 31 in 2.3%, HPV 33 in 1.1%, HPV 52 in 0.9%, HPV 45 in 0.6%, and HPV 59 in 0.4% specimens. CONCLUSION: The data obtained from our study indicate that HPV 16 is the most common HPV type found in the cervical specimens among women of Southern Croatia, as in most studies worldwide, followed by HPV 18 and HPV 31. The positivity rates in our study slightly differed for HPV types 18 and 31 from those found in the other studies conducted in Northern Croatia. These suggesting geographic and environmental factors are related to the prevalence of HPV types. We consider it very important to gather information on regional variations in the HPV type spectrum.

P53, MAPK, topoisomerase II alpha and Ki67 immunohistochemical expression and KRAS/BRAF mutation in ovarian serous carcinomas.

BACKGROUND: We investigated the immunohistochemical expression of p53, MAPK, topoisomerase II alpha (topoII alpha) and Ki67 in ovarian serous carcinomas (OSCs) along with mutational analysis for KRAS and BRAF. METHODS: Eighty one cases of OSCs were reviewed and examined immunohistochemically using antibodies against p53, MAPK, topoII alpha and Ki67. Staining was evaluated as a percentage of immunopositive cells with cut-off levels at 10% for p53 and topoII alpha, and 5% for MAPK. The Ki67 immunoexpression was assessed by means of Olympus Image Analysis System as a percentage of immunopositive cells in 1000 tumor cells. KRAS and BRAF mutational analysis was performed on 73 available microdissected samples. RESULTS: Of 81 cases of OSCs 13.6% were of low-grade and 86.4% were of high-grade morphology. In the high-grade group there was a significantly higher immunoexpression of p53 (P < 0.001) and topoII alpha (P = 0.001), with Ki67 median 56.5 vs. 19 in low-grade group (P < 0.001). The difference in immunoexpression of active MAPK between low- and high-grade group was also significant (P = 0.003). MAPK positive immunostaining was detected in 63.6% of low-grade vs. 17.1% of high-grade OSCs. The frequency of KRAS mutation was significantly higher in low-grade as compared to high-grade group (P = 0.006). None of the samples had BRAF mutation. In addition, we detected positive MAPK immunoexpression in 13/59 samples with wild-type KRAS, suggesting that activation of MAPK pathway is not ultimately related either to KRAS or BRAF mutation. Seven morphologically high-grade samples (11.7%) showed both KRAS mutation and p53 immunopositivity. CONCLUSIONS: Although this study is limited by its humble number of low-grade samples, our data fit the proposed dualistic pathway of ovarian carcinogenesis. Mutational analysis for KRAS and BRAF discloses some possible interactions between different tumorigenic pathways of low- and high-grade carcinomas. Immunohistochemical staining for MAPK was not sufficiently sensitive, nor specific, to precisely predict the KRAS mutation. However, it appears to be quite reliable in ruling out a KRAS mutation if the staining is negative. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9283563368804632.

Ki-67 quantitative evaluation as a marker of cervical intraepithelial neoplasia and human papillomavirus infection.

OBJECTIVE: To assess the value of Ki-67 quantitative analysis in cervical intraepithelial neoplasia (CIN) in relation to CIN grading and human papillomavirus (HPV) group typing. METHODS: Cervical samples selected retrospectively from 106 cases were analyzed immunohistochemically for Ki-67-positive nuclei in 3 epithelial layers and by polymerase chain reaction for HPV typing. RESULTS: The proportion of high-risk HPV positivity was 0% in normal controls and 30% in CIN 1, 57% in CIN 2, and 90% in CIN 3 groups, and there was no low-risk HPV finding in CIN 2 and CIN 3 cases (P < 0.001). High-risk HPV-positive cases exhibited significantly more Ki-67-positive nuclei per 100-mum basal membrane, which were more frequent in the middle and upper third layers of the epithelium compared with low-risk HPV and HPV-negative cases (P < 0.001). The differences among the CIN groups in the total number and in the percentages of Ki-67-positive nuclei in the lower, middle, and upper third layers of the epithelium were significant (P < 0.001). With the cutoff value of more than 33% Ki-67-positive nuclei in the middle and the upper third layers of the epithelium, Ki-67 staining demonstrated 98.4% sensitivity (60/61 cases) and 97.8% specificity (44/45 cases) for the detection of CIN 2/CIN 3 in our study group. CONCLUSIONS: The Ki-67 immunostaining proved to be predictive for high-risk HPV infection, and it can differentiate reactive lesions from cervical dysplasias. Ki-67 quantitative analysis in 3 epithelial layers is a sensitive and specific method of differentiation between CIN 1 and CIN 2/CIN 3 grades and can be a valuable adjunctive method for more accurate CIN grading.

Human papillomavirus DNA typing in the cervical specimens among women of Split and Dalmatian County.

Human papillomavirus (HPV) infection is the main cause of cervical cancer, the second most common cancer in women worldwide. More than 200 types of HPV have been described, and within this range more than 40 types attack epithelium of genital tract. The types that were most commonly related to the development of cervical cancer are called high-risk types (HR HPV). There are very few studies about HPV prevalence in Croatia and there is an absolute lack of data for Split and Dalmatian County. Therefore, during a 6 month period, we routinely screened 570 women for HPV DNA at the Educational Public Health Institute of Split and Dalmatian County. HR HPV was detected in cervical exfoliated cells, by using Hybrid Capture 2 HPV DNA test. Out of the total number of tested women, 200 (35%) of them were positive to HR HPV Polymerase chain reaction (PCR) based assays were employed for HR HPV genotyping in positive specimens. The following frequency was observed: HPV 16 in 10%, HPV 18 in 6.1%, HPV 31 in 2.6%, HPV 33 in 1.9%, HPV 52 in 1.4%, HPV 59 in 0.7%, HPV 45 in 0.4% specimens, while 11.9% of tested specimens currently remained untyped. It is necessary to expand this study to a larger number of women, in order to better evaluate genital HPV types distribution among women in this region.