Regional heritability mapping method helps explain missing heritability of blood lipid traits in isolated populations.

Single single-nucleotide polymorphism (SNP) genome-wide association studies (SSGWAS) may fail to identify loci with modest effects on a trait. The recently developed regional heritability mapping (RHM) method can potentially identify such loci. In this study, RHM was compared with the SSGWAS for blood lipid traits (high-density lipoprotein (HDL), low-density lipoprotein (LDL), plasma concentrations of total cholesterol (TC) and triglycerides (TG)). Data comprised 2246 adults from isolated populations genotyped using ∼300 000 SNP arrays. The results were compared with large meta-analyses of these traits for validation. Using RHM, two significant regions affecting HDL on chromosomes 15 and 16 and one affecting LDL on chromosome 19 were identified. These regions covered the most significant SNPs associated with HDL and LDL from the meta-analysis. The chromosome 19 region was identified in our data despite the fact that the most significant SNP in the meta-analysis (or any SNP tagging it) was not genotyped in our SNP array. The SSGWAS identified one SNP associated with HDL on chromosome 16 (the top meta-analysis SNP) and one on chromosome 10 (not reported by RHM or in the meta-analysis and hence possibly a false positive association). The results further confirm that RHM can have better power than SSGWAS in detecting causal regions including regions containing crucial ungenotyped variants. This study suggests that RHM can be a useful tool to explain some of the 'missing heritability' of complex trait variation.

A K(ATP) channel gene effect on sleep duration: from genome-wide association studies to function in Drosophila.

Humans sleep approximately a third of their lifetime. The observation that individuals with either long or short sleep duration show associations with metabolic syndrome and psychiatric disorders suggests that the length of sleep is adaptive. Although sleep duration can be influenced by photoperiod (season) and phase of entrainment (chronotype), human familial sleep disorders indicate that there is a strong genetic modulation of sleep. Therefore, we conducted high-density genome-wide association studies for sleep duration in seven European populations (N=4251). We identified an intronic variant (rs11046205; P=3.99 × 10(-8)) in the ABCC9 gene that explains ≈5% of the variation in sleep duration. An influence of season and chronotype on sleep duration was solely observed in the replication sample (N=5949). Meta-analysis of the associations found in a subgroup of the replication sample, chosen for season of entry and chronotype, together with the discovery results showed genome-wide significance. RNA interference knockdown experiments of the conserved ABCC9 homologue in Drosophila neurons renders flies sleepless during the first 3 h of the night. ABCC9 encodes an ATP-sensitive potassium channel subunit (SUR2), serving as a sensor of intracellular energy metabolism.

Global health research priorities: mobilizing the developing world.

The paper focuses on two questions: (i) how to set research priorities in a transparent, systematic, fair and legitimate way?; and (ii) how to mobilize low and middle-income countries to take more ownership in defining their own research policies, rather than merely being passive recipients of international aid for research and development? I propose that the recently developed Child Health and Nutrition Research initiative (CHNRI) methodology is becoming widely accepted as a feasible answer to both those questions. In this paper, I review its numerous applications to date and show how it evolved into a practical and systematic tool that can assist priority setting in health research investments in diverse contexts. The CHNRI methodology also addresses support for different instruments of health research to achieve better balance between fundamental research, translation research and implementation research. The wide application of CHNRI methodology is expected to maximise the potential of health research to reduce disease burden and gradually reduce inequities that exist between support for research on the health problems of the rich and the poor. I believe that this tool will find application within many low and middle-income countries and assist them to pull together their own experts and actively define their priorities for research and development in the coming years.

Comparison of the growth hormone, IGF-1 and insulin in cerebrospinal fluid and serum between patients with motor neuron disease and healthy controls.

Neurotrophic effects of the growth hormone (GH), insulin-like growth factor-1 (IGF-1) and insulin on the central nervous system have become more apparent in the past decade. In this study, we measured serum and cerebrospinal fluid (CSF) concentrations of GH, IGF-1 and insulin in 35 patients with motor neuron disease (MND) [24 patients with definite amyotrophic lateral sclerosis (ALS) and 11 patients with progressive bulbar palsy] and in 40 healthy controls. Levels of serum concentrations of GH and IGF-1 did not significantly differ between the MND patient group and the healthy controls, while the level of insulin was significantly decreased (P = 0.0033) in the MND patient group. However, levels of all three examined parameters in CSF were significantly lower in the MND group than in the healthy controls with the statistical significance for IGF-1 and insulin of P < 0.001. This finding has not been reported previously, and further investigations into its association with ALS should establish whether it can be used as an early marker of the disease, or whether it merely represents a consequence of ALS development.

Application of high-dimensional feature selection: evaluation for genomic prediction in man.

In this study, we investigated the effect of five feature selection approaches on the performance of a mixed model (G-BLUP) and a Bayesian (Bayes C) prediction method. We predicted height, high density lipoprotein cholesterol (HDL) and body mass index (BMI) within 2,186 Croatian and into 810 UK individuals using genome-wide SNP data. Using all SNP information Bayes C and G-BLUP had similar predictive performance across all traits within the Croatian data, and for the highly polygenic traits height and BMI when predicting into the UK data. Bayes C outperformed G-BLUP in the prediction of HDL, which is influenced by loci of moderate size, in the UK data. Supervised feature selection of a SNP subset in the G-BLUP framework provided a flexible, generalisable and computationally efficient alternative to Bayes C; but careful evaluation of predictive performance is required when supervised feature selection has been used.

The evidence of mtDNA haplogroup F in a European population and its ethnohistoric implications.

Mitochondrial DNA polymorphism was analysed in a sample of 108 Croatians from the Adriatic Island isolate of Hvar. Besides typically European varieties of human maternal lineages, haplogroup F was found in a considerable frequency (8.3%). This haplogroup is most frequent in southeast Asia but has not been reported before in Europe. The genealogical analysis of haplogroup F cases from Hvar suggested founder effect. Subsequent field work was undertaken to sample and analyse 336 persons from three neighbouring islands (Brac, Korcula and Krk) and 379 more persons from all Croatian mainland counties and to determine if haplogroup F is present in the general population. Only one more case was found in one of the mainland cities, with no known ancestors from Hvar Island. The first published phylogenetic analysis of haplogroup F worldwide is presented, applying the median network method, suggesting several scenarios how this maternal lineage may have been added to the Croatian mtDNA pool.

Genetic epidemiological studies of eastern Adriatic Island isolates, Croatia: objective and strategies.

In this paper, the authors discuss why isolate island populations represent a particularly helpful model for genetic epidemiological studies. A thorough previous anthropological research carried out in Eastern Adriatic island isolates, Croatia, in terms of ethnohistory, geography and current demography is reviewed. The major results of the studies of population genetic structure of those populations, including model-bound and model-free approaches, the analyses of serogenetic polymorphisms and most recent studies using HLA class II, VNTR and STR DNA polymorphisms, are briefly presented. The organization of health care on the islands is analyzed and some relevant details of specific medical problems and some autochtonous diseases in these island populations is noted. The authors present in outline four illustrative examples of research opportunities which are afforded by the unique circumstances found in these isolate communities. These relate to hereditary dwarfism on Krk island, Mal de Meleda on Mljet island, extreme inbreeding on Susak island and population genetics of cancer on the islands of Brac, Hvar, Korcula, Vis and Lastovo. Finally, the authors develop objectives and strategies for a long-term genetic epidemiological research of these populations and suggest that such a programme of investigation would further our understanding of the causes of (rare) diseases which are uniquely important to these communities but also of common diseases which are important contributors to the burden of disease both in these islands and throughout the world.

Holistic approach to analysis of medical data: vulvar cancer.

This paper continues the series of studies introducing holistic approach to analysis of clinical data. Namely, besides the information regarding his/her disease, each hospitalized cancer patient also provides the variety of data regarding his/her psychological, cultural, social, economical, genetic, constitutional and medical background. The aim of this study was to introduce a holistic approach to analysis of medical data, in this case clinical data regarding cancer of the vulva. Such approach requires the collection of data regarding different aspects of the cancer patients, and after the satisfactory sample size is obtained (which should be at least five times greater than the number of examined patient characteristics), the performance of factor analysis. In this study, the authors have processed the data regarding 25 characteristics of all 755 vulvar cancer patients treated between 1938 and 1990 at the Department for Gynecological Oncology of the University Hospital for Gynecology and Obstetrics, Zagreb, Croatia. In factor analysis, the principal components were rotated after the initial extraction (the authors recommended the use of oblimin rotation) in order to obtain better ground for interpretation of the obtained results. The next step in this approach was the stepwise exclusion of characteristics with smallest commonality according to Kaiser-Meyer-Olkin criteria, and retaining the characteristics and components with the most significant impact on the explained system variance. When the number of principal components and initial analyzed characteristics was reduced to 3-4 and 7-10, respectively, the ultimate interpretations and conclusions were made. This approach outlined some clusters of correlations between medical data which are difficult to identify using other statistical procedures, primarily the impacts of various socioeconomic and hereditary-constitutional variables on overall survival.

Holistic approach to analysis of medical data: cancer of the corpus uteri.

Besides the information regarding his/her disease, each hospitalized cancer patient also provides the variety of data regarding his/her psychological, cultural, social, economical, genetic, constitutional and medical background. The aim of this study was to introduce a holistic approach to analysis of medical data, in this case clinical data regarding cancer of the corpus uteri. Such approach requires the collection of data regarding different aspects of the cancer patient, and after the satisfactory sample size is obtained (which should be at least five times greater than the number of examined patient characteristics), the performance of factor analysis. In this study, the authors have processed the data regarding 25 characteristics of 928 corpus uteri cancer patients treated between 1980 and 1990 at the Department for Gynecological Oncology of the University Hospital for Gynecology and Obstetrics, Zagreb, Croatia. In factor analysis, the principal components were rotated after the initial extraction (the authors recommended the use of oblimin rotation) in order to obtain better ground for interpretation of the obtained results. The next step in this approach was the stepwise exclusion of characteristics with smallest communalities according to Kaiser-Meyer-Olkin criteria, and retaining the characteristics and components with the most significant impact on the explained system variance. When the number of principal components and initial analyzed characteristics was reduced to 3-4 and 7-10, respectively, the ultimate interpretations and conclusions were made. This approach outlined some clusters of correlations between medical data which are difficult to identify using other statistical procedures, primarily the impacts of various socioeconomic and hereditary-constitutional variables on overall survival.

MtDNA haplogroups in the populations of Croatian Adriatic Islands.

The number of previous anthropological studies pointed to very complex ethnohistorical processes that shaped the current genetic structure of Croatian island isolates. The scope of this study was limited to the general insight into their founding populations and the overall level of genetic diversity based on the study mtDNA variation. A total of 444 randomly chosen adult individuals from 32 rural communities of the islands of Krk, Brac, Hvar and Korcula were sampled. MtDNA HVS-I region together with RFLP sites diagnostic for main Eurasian and African mtDNA haplogroups were analysed in order to determine the haplogroup structure. The most frequent haplogroups were "H" (27.8-60.2%), "U" (10.2-24.1%), "J" (6.1-9.0%) and "T" (5.1-13.9%), which is similar to the other European and Near Eastern populations. The genetic drift could have been important aspect in history, as there were examples of excess frequencies of certain haplogroups (11.3% of "I" and 7.5% of "W" in Krk, 10.5% of "HV" in Brac, 13.9% of "J" in Hvar and 60.2% of "H" in Korcula). As the settlements on the islands were formed trough several immigratory episodes of genetically distant populations, this analysis (performed at the level of entire islands) showed greater genetic diversity (0.940-0.972) than expected at the level of particular settlements.

Surname as 'cancer risk' in extreme isolates: example from the island of Lastovo, Croatia.

The aim of this study was to analyze whether there are surnames which appear more frequently among the ancestors of cancer cases in a small isolate, in comparison to the ancestral surnames of the healthy controls, using the classic case-control design. The chosen setting was the island of Lastovo, Croatia, located more than 100 kilometers from the nearest coastal region. The period of study was 1970-1995, during which a total of 76 cancer cases were recorded in a population of approximately 800. The comparison of surname frequencies was performed in current and in five ancestral generations. The leading hypothesis was that, if inbreeding and common ancestry contributed to the development of the disease, then those phenomena should be reflected in increasing frequency of some surnames among ancestors, identifying the 'hidden' consanguinity, or 'following' cancer-promoting genes on the Y-chromosome. The results imply that there are surnames representing a classic "risk" for cancer, but also those "protecting" from its development, which all underscores the importance of founder effect and genetic predisposition to the disease in a small, reproductively isolated population. All of the results become more evident and increasingly significant when analyzed in more distant ancestral generations.

Inbreeding and nephrolithiasis in Croatian island isolates.

The aim of this study was to investigate a recessive genetic component in susceptibility to nephrolithiasis (NL) by comparing its prevalence in highly inbred, moderately inbred and non-inbred villages of three Croatian islands: Brac, Hvar and Korcula. The average inbreeding coefficient of each village population (F) was estimated in a random sample of 20-30% adults from 14 villages using Wright's path method (based on genealogical information), isonymy data and average deviation from Hardy-Weinberg expectations for MN, Ss and Kk serogenetic polymorphisms. The six villages with the greatest genealogical F value (0.025-0.049; current total population N=1,401), the four with intermediate value (0.012-0.015; N=998) and the four with the smallest value (0.002-0.008; N=1,500) were chosen for this study. Medical records of entire populations were reviewed and the diagnosis of NL was established according to unified criteria, based on the information from the specialists in general hospitals and on the agreement in diagnosis between the investigators and local general practitioners. The variance in environmental and socio-cultural factors between villages was shown to be minimal. Prevalence of NL in each village was standardised by sex and age to the total population of all 14 villages. The standardized prevalence of NL was 1.5% in the group of villages with low F, 2.3% in the group with moderate F (p<0.10), and 5.4% in the group with high F (p<0.001). The correlation factor between F values by villages and the standardized prevalence of NL was 0.45 (p<0.05). The study showed that a large number of predominantly recessive genetic factors might mediate the susceptibility to formation of renal stones in humans.

Inbreeding and learning disability in Croatian island isolates.

The aim of this study was to investigate the prevalence of learning disability (LD) in isolate populations with different inbreeding coefficients (F). Prevalence of LD and F were determined in 10 villages from five Croatian islands: Brac, Hvar, Korcula, Lastovo and Susak. For the purpose of this study, LD was defined as the inability to attend the public school system. As the elementary schools (grade 1-8) in the place of the study are both public and compulsory, the assessment of child's inability to attend the school is performed at the age of six. This is required by all children in the country based on standard set of tests of cognitive performance defined by the Ministry of Education and Culture of the Republic of Croatia. The average inbreeding coefficients in each village population (F) were estimated in a random sample of 20-30% adults in each of the 10 villages based on 4 ancestral generations and using Wright's path method. Prevalence of LD ranged from 0.43% to 2.47%, and the inbreeding coefficients ranged from 0.8% to 4.9%. The Pearson's correlation coefficient between F and LD prevalence was 0.80 (p < 0.01). Although the relative risk per 5% inbreeding appeared very high (about 10), the absolute risk only increased from 0.18% to 1.77%. The genetic effect of inbreeding (GEI) was approximately 0.69% and the population-attributable fraction 76.6%. A review of the literature and the results of this study lead to a conclusion that a very large number of predominantly recessive genetic factors might mediate the genetic susceptibility to various forms of LD in these populations.

Cancer incidence in eastern Adriatic isolates, Croatia: examples from the islands of Krk, Cres, Losinj, Rab and Pag.

As an extension of previous research this study investigates the incidence of cancer in five genetic isolate island populations of the Eastern Adriatic, Croatia. Thorough anthropological research over the past three decades has established some of those populations as outstanding examples of genetic isolates. A previous study which found higher cancer incidence in 5 Eastern Adriatic islands than in a control population supported a hypothesis that among the founders of these populations there were genetic variants (especially with recessive inheritance) responsible for genetic susceptibility to certain types of cancer. This study sought to investigate cancer incidence in 5 further island populations. All cancer cases in five island populations (Krk, Cres, Losinj, Rab and Pag) over the 20-year period (1971 to 1990) was extracted from the data of the Croatian Cancer Registry. The mainland populations of Istrian and Primorsko-Goranska County, characterized by similar environmental factors but an outbred genetic structure, represented a control population. After standardization by by sex and age, cancer incidence was higher in the island populations than in the control population in both sexes. The cancer sites primarily responsible for the excess incidence were prostate, stomach and pancreatic cancer in males, and ovarian, breast, stomach, bowel, and brain cancer in females. The reasons for the increased cancer incidence are uncertain and may be due to different environmental exposure between the two populations. However, it is possible that genetic isolation and inbreeding are important factors. Further investigations of cancer in these isolate populations are warranted to explore these findings further.

A large cross-sectional study of health attitudes, knowledge, behaviour and risks in the post-war Croatian population (the First Croatian Health Project).

As the liberation of occupied Croatian territories ended the war in the country in 1995, the Ministry of Health and Croatian Health Insurance Institute have agreed to create the new framework for developing a long-term strategy of public health planning, prevention and intervention. They provided financial resources to develop the First Croatian Health Project, the rest of the support coming from the World Bank loan and the National Institute of Public Health. A large cross-sectional study was designed aiming to assess health attitudes, knowledge, behaviour and risks in the post-war Croatian population. The large field study was carried out by the Institute for Anthropological Research with technical support from the National Institute of Public Health. The field study was completed between 1995-1997. It included about 10,000 adult volunteers from all 21 Croatian counties. The geographic distribution of the sample covered both coastal and continental areas of Croatia and included rural and urban environments. The specific measurements included antropometry (body mass index and blood pressure). From each examinee a blood sample was collected from which the levels of total plasma cholesterol (TC), triglycerides (TG), HDL-cholesterol (High Density Lipoprotein), LDL-cholesterol (Low Density Lipoprotein), lipoprotein Lp(a), and haemostatic risk factor fibrinogen (F) were determined. The detailed data were collected on the general knowledge and attitudes on health issues, followed by specific investigation of smoking history, alcohol consumption, nutrition habits, physical activity, family history of chronic non-communicable diseases and occupational exposures. From the initial database a targeted sample of 5,840 persons of both sexes, aged 18-65, was created corresponding by age, sex and geographic distribution to the general Croatian population. This paper summarises and discusses the main findings of the project within this representative sample of Croatian population.

Breast cancer prognosis. I. Prognostic factors in patients with node-negative (N0) breast cancer.

Predictors of breast cancer survival were investigated among 196 node-negative (N0) breast cancer patients treated at the Department of Surgery of the University Hospital for Tumors in Zagreb between 1969-1988. Selected prognostic factors included patient age, delay in treatment, tumor size, histologic grade of malignancy, estrogen receptor status, progesterone receptor status, tumor site, and type of surgical treatment. Among these predictors, only tumor size (p < 0.001) and pathohistologic grade of malignancy (p < 0.001) caused significant differences in 5-year overall survival rates (T1-92.1%, T2-89.8%, T3-64.1% and T4-45.4%; grade I-95.0%; II-80.6%; III-63.6%). The authors conclude that among eight selected clinical characteristics, only tumor size and pathohistological malignancy grade can serve as helpful predictors in determination of the probability of 5-year overall survival among node-negative breast cancer patients.

Breast cancer prognosis. II. Prognostic factors in patients with node-positive (N1-3) breast cancer.

Predictors of breast cancer survival were investigated among 282 node-positive (N1-3) breast cancer patients treated at the Department of Surgery of the University Hospital for Tumors in Zagreb between 1969-1988. Selected prognostic factors included patient age, delay in treatment, tumor size, type of lymph-node affection, pathohistological grade of malignancy, estrogen receptor status, progesterone receptor status, tumor site, and type of surgical treatment. Among these predictors, only tumor size (p < 0.001), type of lymph-node affection (p < 0.001), malignancy grade (p < 0.001), and progesterone receptor status (p < 0.001) revealed a significant impact on a 5-year overall survival rates (T1-100%, T2-64.5%, T3-54.5% and T4-23.7%; N1-60.2%, N2-20.8%, N3-30.3%; grade I-85.5%, II-59.4%, III-30.5%; PgR+ 63.8%, PgR--26.4%). The authors conclude that among nine selected characteristics, only tumor size, type of lymph-node affection, pathohistological malignancy grade, and progesterone receptor status can be helpful predictors in the determination of the probability of 5-year overall survival among node-positive breast cancer patients.

Breast cancer prognosis. III. Prognostic factors in patients with distant metastases (M1) at the time of diagnosis.

Predictors of breast cancer survival were investigated among 66 patients who had distant metastases at the time of diagnosis (M1). All of these patients were treated at the Department of Surgery of the University Hospital for Tumors in Zagreb between 1969-1988. Selected prognostic factors included patient age, delay in treatment, tumor size, type of lymph node affection, response to palliative surgical treatment and administered chemotherapy or hormonal therapy, and site of metastasizing. Among these predictors, several of them revealed a significant impact on a median survival (in months) of these patients: the most important was the response to chemotherapy (p < 0.001), followed by site of metastasizing (p < 0.05) and primary tumor size (p < 0.05). Palliative surgical treatment, apart from improvement of life quality, played no role in determining the survival among breast cancer patients with a distant disease.

Differences between male and female breast cancer. I. Epidemiological features.

The authors' interest was focused on the direct determination of differences between the salient features of breast cancer in men and women in a large number of patients. Various epidemiological features were analyzed in a group of 386 male and 21,491 female breast cancer patients registered in the Cancer Registry of the Republic of Croatia from 1968 until 1988. This was a population-based study, and all male and female breast cancer cases recorded during the mentioned period were included in the study. During this 21-year period, the total incidence rate of breast cancer in men was about 0.83/100,000, and the mortality 0.2,/100.000. In the same interval, in women the rates doubled from 30 to 60/100,000, and 13 to 29/100,000, respectively. The highest incidence rates were recorded in the 80-84 age group in men, and 60-64 age group in women. The mean age at diagnosis was 63.4 years in men and 58.2 years in women. The distribution in terms of TNM-stages in men was: stage I-10.10%; II-40.67%; III-32.40%; IV-16.83%; in women; I-12.19%; II-49.36%; III-27-58%; IV-10.86%. The results showed that breast cancer in men had epidemiological characteristics of a sporadic disease, occurring continuously at a specific mean frequency in the general population and being little affected by environmental impacts. Conversely, in women affected by the disease there was a tendency of continuous rise in the morbidity and mortality, which might have been due to the trigger action of one or more environmental factors.(ABSTRACT TRUNCATED AT 250 WORDS)

Locally recurring primary myxoma of the breast: an evidence of malignant alteration.

Breast is an extremely rare localization of myxoma. To date, only five cases have been reported, including one case of nerve sheath myxoma. In this report, the authors present a 37-year-old female patient who was admitted to the University Hospital for Tumors in Zagreb, Croatia, in 1979, with a 2-month history of gradually enlarging, painless subcutaneous mass in her right breast. On physical examination, a firm, relatively well circumscribed mass measuring 3 x 2 cm was detected in the upper outer quadrant and partly beneath the areola. The mass was freely movable to the overlying skin, causing no retraction. There was no family history of a breast disease or cancer. Patient's past history revealed no major illnesses or trauma. The tumor was removed by simple local excision, but it recurred locally in three episodes between 1981 and 1986. After that, the tumor underwent malignant alteration to myxosarcoma on its fourth (in 1992), and to myxoid liposarcoma on its fifth recurrence (in 1993). No such sequence of events has been reported in the literature so far.