Art-based observational training for medical students and surgical residents in two Dutch museums.

Background: Art-based education is gaining interest in the medical field, particularly in specialties with a strong visual focus. Visual arts are increasingly used for the development of observational skills and social competencies. While content and objectives of art-based programs widely differ across medical faculties in the Netherlands, the diverse range of options underscore the interest in and the potential of this educational approach. In this report, we explore the value of art-based observational training for medical students and surgical residents in two prominent Dutch museums in Amsterdam and Rotterdam, respectively. Methods: Our program, conducted at the Rijksmuseum in Amsterdam and Depot Boijmans van Beuningen Museum in Rotterdam engaged medical students (n=24) and surgeons (in training) (n=66) in an interactive workshop focused on art observation led by an experienced art-educator and a clinical professional. Learning objectives were defined and a post-workshop questionnaire was devised to evaluate participants' perceptions, with a specific focus on contribution of the program to professional development. Results: Both residents and surgeons acknowledged that the program had a positive impact on their professional skills. The program learned them to postpone their judgements and contributed to the awareness of their personal bias. Notably, medical students believed in the program's potential contribution to their professional development. Surgeons were more critical in their evaluation, emphasizing the challenge of sustainable improvement of skills within the limited duration of the course. Conclusions: An interactive art-based medical education program was offered to medical students, PhD students, house officers, surgical residents and surgeons in two well known Dutch museums. Participants expressed enthusiasm for the innovative educational approach they experienced at the museums. They learned about the importance of critical observation in their professional work, handling of ambiguity and got the opportunity to practice both observational and communicational skills in a creative manner. The findings indicate that medical students and surgical residents can benefit from art-based observational training, using art as a vehicle to develop their professional competencies.

Fire-induced pine woodland to shrubland transitions in Southern Europe may promote shifts in soil fertility.

Since the mid of the last century, fire recurrence has increased in the Iberian Peninsula and in the overall Mediterranean basin due to changes in land use and climate. The warmer and drier climate projected for this region will further increase the risk of wildfire occurrence and recurrence. Although the impact of wildfires on soil nutrient content in this region has been extensively studied, still few works have assessed this impact on the basis of fire recurrence. This study assesses the changes in soil organic C and nutrient status of mineral soils in two Southern European areas, Várzea (Northern Portugal) and Valencia (Eastern Spain), affected by different levels of fire recurrence and where short fire intervals have promoted a transition from pine woodlands to shrublands. At the short-term (<1year), the amount of soil organic matter was higher in burned than in unburned soils while its quality (represented as labile to total organic matter) was actually lower. In any case, total and labile soil organic matter showed decreasing trends with increasing fire recurrence (one to four fires). At the long-term (>5years), a decline in overall soil fertility with fire recurrence was also observed, with a drop between pine woodlands (one fire) and shrublands (two and three fires), particularly in the soil microsites between shrubs. Our results suggest that the current trend of increasing fire recurrence in Southern Europe may result in losses or alterations of soil organic matter, particularly when fire promotes a transition from pine woodland to shrubland. The results also point to labile organic matter fractions in the intershrub spaces as potential early warning indicators for shifts in soil fertility in response to fire recurrence.

Disparity of basal and therapeutically activated interferon signalling in constraining hepatitis E virus infection.

Hepatitis E virus (HEV) represents one of the foremost causes of acute hepatitis globally. Although there is no proven medication for hepatitis E, pegylated interferon-α (IFN-α) has been used as off-label drug for treating HEV. However, the efficacy and molecular mechanisms of how IFN signalling interacts with HEV remain undefined. As IFN-α has been approved for treating chronic hepatitis C (HCV) for decades and the role of interferon signalling has been well studied in HCV infection, this study aimed to comprehensively investigate virus-host interactions in HEV infection with focusing on the IFN signalling, in comparison with HCV infection. A comprehensive screen of human cytokines and chemokines revealed that IFN-α was the sole humoral factor inhibiting HEV replication. IFN-α treatment exerted a rapid and potent antiviral activity against HCV, whereas it had moderate and delayed anti-HEV effects in vitro and in patients. Surprisingly, blocking the basal IFN pathway by inhibiting JAK1 to phosphorylate STAT1 has resulted in drastic facilitation of HEV, but not HCV infection. Gene silencing of the key components of JAK-STAT cascade of the IFN signalling, including JAK1, STAT1 and interferon regulatory factor 9 (IRF9), stimulated HEV infection. In conclusion, compared to HCV, HEV is less sensitive to IFN treatment. In contrast, the basal IFN cascade could effectively restrict HEV infection. This bears significant implications in management of HEV patients and future therapeutic development.

Prednisolone does not affect direct-acting antivirals against hepatitis C, but inhibits interferon-alpha production by plasmacytoid dendritic cells.

BACKGROUND: Chronic hepatitis C virus (HCV) infection compromises long-term outcomes of liver transplantation. Although glucocorticosteroid-based immunosuppression is commonly used, discussion is ongoing on the effect of prednisolone (Pred) on HCV recurrence and response to antiviral therapy post transplantation. Recently, new drugs (direct-acting antivirals) have been approved for the treatment of HCV, however, it remains unknown whether their antiviral activity is affected by Pred. The aim of this study was to investigate the effects of Pred on the antiviral activity of asunaprevir (Asu), daclatasvir (Dac), ribavirin (RBV), and interferon-alpha (IFN-α), and on plasmacytoid dendritic cells (PDCs), the main IFN-α-producing immune cells. METHODS: The effects of Pred and antiviral compounds were tested in both a subgenomic and infectious HCV replication model. Furthermore, effects were tested on human PDCs stimulated with a Toll-like receptor-7 ligand. RESULT: Pred did not directly affect HCV replication and did not inhibit the antiviral action of Asu, Dac, RBV, or IFN-α. Stimulated PDCs potently suppressed HCV replication. This suppression was reversed by treating PDCs with Pred. Pred significantly decreased IFN-α production by PDCs without affecting cell viability. When Asu and Dac were combined with PDCs, a significant cooperative antiviral effect was observed. CONCLUSION: This study shows that Pred acts on the antiviral function of PDCs. Pred does not affect the antiviral action of Asu, Dac, RBV, or IFN-α. This implies that there is no contraindication to combine antiviral therapies with Pred in the post-transplantation management of HCV recurrence.

Canine hepacivirus and idiopathic hepatitis in dogs from a Dutch cohort.

Liver diseases are highly prevalent in the general dog population, though the etiology is often unknown. Recently a homolog of human hepatitis C virus was discovered in dogs with respiratory infections. Although this canine hepacivirus (CHV) was detectable in some liver samples, a clear link with liver disease has not been established. A recent study by Bexfield et al. showed that in a large cohort of dogs from the UK with idiopathic hepatitis, no evidence can be found for exposure to, or carrier state of CHV both in liver and in serum. The authors however state that 'the absence of CHV infection on dogs from the UK might not represent the global ecology of the virus'. We investigated CHV-infection in 267 liver biopsies from 120 dogs idiopathic hepatitis and 135 control animals, in a population from the Netherlands. Using a highly sensitive PCR assay for CHV-NS3, no CHV was detected in all 267 liver samples. Our data show that the lack of association between canine hepacivirus and chronic liver disease in dogs is not limited to the UK, but is also found in an independent cohort from the European continent.

Sensitive detection of hepatocellular injury in chronic hepatitis C patients with circulating hepatocyte-derived microRNA-122.

As chronic hepatitis C patients with progressive disease can present themselves with normal ALT levels, more sensitive biomarkers are needed. MicroRNAs are newly discovered small noncoding RNAs that are stable and detectable in the circulation. We aimed to investigate the association between hepatocyte-derived microRNAs in serum and liver injury in patients with chronic hepatitis C. The hepatocyte-derived miR-122 and miR-192 were analysed in sera of 102 chronic HCV-infected patients and 24 healthy controls. Serum levels of miR-122 and miR-192 correlated strongly with ALT (R = 0.67 and R = 0.65, respectively, P < 0.001 for both). Median levels of miR-122 and miR-192 in HCV-infected patients were 23 times and 8 times higher as in healthy controls (P < 0.001 for both). Even within the HCV-infected patients with a normal ALT (n = 38), the levels of miR-122 and miR-192 were 12 times and 4 times higher compared with healthy controls (P < 0.001 for both). Multivariate logistic regression analyses showed that only miR-122 was a significant predictor of the presence of chronic HCV infection (P = 0.026). Importantly, miR-122 was also superior in discriminating chronic HCV-infected patients with a normal ALT from healthy controls compared with the ALT level (AUC = 0.97 vs AUC = 0.78, P = 0.007). In conclusion, our study confirmed that liver injury is associated with high levels of hepatocyte-derived microRNAs in circulation and demonstrated that in particular miR-122 is a sensitive marker to distinguish chronic hepatitis C patients from healthy controls. More sensitive blood markers would benefit especially those patients with minor levels of hepatocellular injury, who are not identified by current screening with ALT testing.

Divergent composition but similar function of soil food webs of individual plants: plant species and community effects.

Soils are extremely rich in biodiversity, and soil organisms play pivotal roles in supporting terrestrial life, but the role that individual plants and plant communities play in influencing the diversity and functioning of soil food webs remains highly debated. Plants, as primary producers and providers of resources to the soil food web, are of vital importance for the composition, structure, and functioning of soil communities. However, whether natural soil food webs that are completely open to immigration and emigration differ underneath individual plants remains unknown. In a biodiversity restoration experiment we first compared the soil nematode communities of 228 individual plants belonging to eight herbaceous species. We included grass, leguminous, and non-leguminous species. Each individual plant grew intermingled with other species, but all plant species had a different nematode community. Moreover, nematode communities were more similar when plant individuals were growing in the same as compared to different plant communities, and these effects were most apparent for the groups of bacterivorous, carnivorous, and omnivorous nematodes. Subsequently, we analyzed the composition, structure, and functioning of the complete soil food webs of 58 individual plants, belonging to two of the plant species, Lotus corniculatus (Fabaceae) and Plantago lanceolata (Plantaginaceae). We isolated and identified more than 150 taxa/groups of soil organisms. The soil community composition and structure of the entire food webs were influenced both by the species identity of the plant individual and the surrounding plant community. Unexpectedly, plant identity had the strongest effects on decomposing soil organisms, widely believed to be generalist feeders. In contrast, quantitative food web modeling showed that the composition of the plant community influenced nitrogen mineralization under individual plants, but that plant species identity did not affect nitrogen or carbon mineralization or food web stability. Hence, the composition and structure of entire soil food webs vary at the scale of individual plants and are strongly influenced by the species identity of the plant. However, the ecosystem functions these food webs provide are determined by the identity of the entire plant community.

Empirical and theoretical challenges in aboveground-belowground ecology.

A growing body of evidence shows that aboveground and belowground communities and processes are intrinsically linked, and that feedbacks between these subsystems have important implications for community structure and ecosystem functioning. Almost all studies on this topic have been carried out from an empirical perspective and in specific ecological settings or contexts. Belowground interactions operate at different spatial and temporal scales. Due to the relatively low mobility and high survival of organisms in the soil, plants have longer lasting legacy effects belowground than aboveground. Our current challenge is to understand how aboveground-belowground biotic interactions operate across spatial and temporal scales, and how they depend on, as well as influence, the abiotic environment. Because empirical capacities are too limited to explore all possible combinations of interactions and environmental settings, we explore where and how they can be supported by theoretical approaches to develop testable predictions and to generalise empirical results. We review four key areas where a combined aboveground-belowground approach offers perspectives for enhancing ecological understanding, namely succession, agro-ecosystems, biological invasions and global change impacts on ecosystems. In plant succession, differences in scales between aboveground and belowground biota, as well as between species interactions and ecosystem processes, have important implications for the rate and direction of community change. Aboveground as well as belowground interactions either enhance or reduce rates of plant species replacement. Moreover, the outcomes of the interactions depend on abiotic conditions and plant life history characteristics, which may vary with successional position. We exemplify where translation of the current conceptual succession models into more predictive models can help targeting empirical studies and generalising their results. Then, we discuss how understanding succession may help to enhance managing arable crops, grasslands and invasive plants, as well as provide insights into the effects of global change on community re-organisation and ecosystem processes.

Functional stability of microbial communities in contaminated soils near a zinc smelter (Budel, the Netherlands).

Environmental pollution causes adverse effects on many levels of ecosystem organization; it might affect the use efficiency of available resources which will make the system more sensitive to subsequent stress. Alternatively the development of community tolerance may make the system more resistant to additional stresses. In this study we investigate the functional stability, measured in the terms of resistance and resilience, of microbial populations inhabiting contaminated soils near a zinc smelter. With functional stability we mean that we look at processes rather than at population dynamics. We measure changes in respiration and bacterial growth rate in response to addition of stress (lead, salt) or disturbance (heat). We used soils that differ in the level of pollution with zinc and cadmium originating from an adjacent smelter. Our results showed, with regard to respiration, that the most polluted soils have the lowest stability to salt (stress) and heat (disturbance). This confirms the hypothesis that more stressed systems have less energy to cope with additional stress or disturbance. However, bacterial growth rates were affected in a different way than respiration. There was no difference between the soils in resistance and resilience to addition of lead. In case of salt treatment, the least polluted soils showed highest stability. In contrast, the least polluted soils were the least stable to increased temperature, which supports the hypothesis that more stressed soils are more stable to additional stress/disturbance due to properties they gained when exposed to the first stress (pollution by the smelter). Thus, the responses of microbial processes to stress, their nature and size, depend on the kinds of stress factors, especially whether a subsequent stress is similar to the first stress, in terms of the mechanism with which the organisms deal with the stress.

Ring-reinforced prosthesis for paracolostomy hernia.

BACKGROUND: The reported recurrence rate after paracolostomy hernia repair, with or without mesh, varies between 47 and 100%. We developed a polypropylene mesh prosthesis with a central opening, reinforced with a polypropylene ring, that can be implanted suprafascially through a mucocutaneous incision. METHODS: The results of a consecutive series of 46 patients with paracolostomy hernia, who were implanted with this prosthesis between October 1988 and November 2000 in 12 hospitals, were analyzed. The mean follow-up was 5 (range 1-13) years. RESULTS: The early infection rate was 4.3%. Two patients were lost to follow-up. The late infection rate was 2.3%. Seven patients (15.9%) developed a recurrent parastomal hernia and were all reoperated. The prosthesis was definitively removed in 10 patients. CONCLUSION: Although implantation of this new prosthesis is not without complications, it offers the lowest reported recurrence rate until now.

Progestogenic effects of tibolone on human endometrial cancer cells.

Tibolone, a synthetic steroid acting in a tissue-specific manner and used in hormone replacement therapy, is converted into three active metabolites: a Delta(4) isomer (exerting progestogenic and androgenic effects) and two hydroxy metabolites, 3 alpha-hydroxytibolone (3 alpha-OH-tibolone) and 3beta-OH-tibolone (exerting estrogenic effects). In the present study an endometrial carcinoma cell line (Ishikawa PRAB-36) was used to investigate the progestogenic properties of tibolone and its metabolites. This cell line contains progesterone receptors A and B, but lacks estrogen and androgen receptors. When tibolone was added to the cells, complete conversion into the progestogenic/androgenic Delta(4) isomer was observed within 6 d. Furthermore, when cells were cultured with tibolone or when the Delta(4) isomer or the established progestagen medroxyprogesterone acetate was added to the medium, marked inhibition of growth was observed. Interestingly, 3 beta-OH-tibolone also induces some inhibition of growth. These growth inhibitions were not observed in progesterone receptor-negative parental Ishikawa cells, and progestagen-induced growth inhibition of PRAB-36 cells could readily be reversed using the antiprogestagen Org-31489. Upon measuring the expression of two progesterone-regulated genes (fibronectin and IGF-binding protein-3), tibolone, the Delta(4) isomer and medroxyprogesterone acetate showed similar gene expression regulation. These results indicate that tibolone, the Delta(4) metabolite, and to some extent 3 beta-OH-tibolone exert progestogenic effects. Tibolone and most likely 3 beta-OH-tibolone are converted into the Delta(4) metabolite.

Implications of removing a normal appendix.

BACKGROUND: The diagnosis of acute appendicitis remains difficult and therefore 15-30% of the removed appendices appear to be normal. The aim of this study was to investigate morbidity, mortality and costs of removing a normal appendix in patients with suspected appendicitis. PATIENTS AND METHODS: Retrospective study of patients who underwent a negative appendectomy for suspected appendicitis in the period 1991-1999 with a median follow-up of 4.4 years. Patients who underwent an elective appendectomy or appendectomy for other reasons were excluded. RESULTS: In 285 patients (70% women, 30% men) a normal appendix was removed. In 192 (67%) patients a muscle-splitting incision was performed, in 6 (2%) a median laparotomy, and in 51 (18%) the normal appendix was removed by laparoscopy. In 36 patients (13%) a diagnostic laparoscopy was converted to a muscle-splitting incision. Complications occurred in 16 (6%) patients, in 5 (2%) a re-operation was needed. The mean hospital stay was 4.4 (SE 2.8) days, in case of complications 7.4 (SE 4.2) days. The mean extra hospital costs of a negative appendectomy were EUR 2712. CONCLUSION: The removal of a normal appendix has considerable complications and costs. In an attempt to prevent these costs, extra diagnostic tools should be considered. Expensive diagnostic tools such as diagnostic laparoscopy should be used selectively in order not to further increase costs.

Implications of removing a normal appendix.

BACKGROUND: The diagnosis of acute appendicitis remains difficult, and therefore 15-30% of the removed appendices appear to be normal. The aim of this study is to investigate the morbidity, mortality and costs of removing a normal appendix in patients with suspected appendicitis. PATIENTS AND METHODS: A retrospective study was performed on patients who underwent a negative appendectomy for suspected appendicitis in the period 1991-1999 with a median follow-up of 4.4 years. Patients who underwent an elective appendectomy or appendectomy for other reasons were excluded. RESULTS: In 285 patients (70% women, 30% men) a normal appendix was removed. In 192 (67%) patients a muscle-splitting incision was performed, in 6 (2%) a median laparotomy, and in 51 (18%) the normal appendix was removed by laparoscopy. In 36 patients (13%) a diagnostic laparoscopy was converted to a muscle-splitting incision. Complications occurred in 16 (6%) patients, in 5 (2%) a reoperation was needed. The mean hospital stay was 4.4 (SE 2.8) days, in case of complication 7.4 (SE 4.2) days. The mean extra hospital costs of a negative appendectomy were EUR 2,712. CONCLUSION: The removal of a normal appendix has considerable complications and costs. In an attempt to prevent these costs, extra diagnostic tools should be considered. Expensive diagnostic tools as diagnostic laparoscopy should be used selectively in order to not further exceed costs.

Phenotypic variation in a family with partial androgen insensitivity syndrome explained by differences in 5alpha dihydrotestosterone availability.

Mutations in the androgen receptor (AR) gene result in a wide range of phenotypes of the androgen insensitivity syndrome (AIS). Inter- and intrafamilial differences in the phenotypic expression of identical AR mutations are known, suggesting modifying factors in establishing the phenotype. Two 46,XY siblings with partial AIS sharing the same AR gene mutation, R846H, but showing very different phenotypes are studied. Their parents are first cousins. One sibling with grade 5 AIS was raised as a girl; the other sibling with grade 3 AIS was raised as a boy. In both siblings serum levels of hormones were measured; a sex hormone-binding globulin (SHBG) suppression test was completed; and mutation analysis of the AR gene, Scatchard, and SDS-PAGE analysis of the AR protein was performed. Furthermore, 5alpha-reductase 2 expression and activity in genital skin fibroblasts were investigated, and the 5alpha-reductase 2 gene was sequenced. The decrease in SHBG serum levels in a SHBG suppression test did not suggest differences in androgen sensitivity as the cause of the phenotypic variation. Also, androgen binding characteristics of the AR, AR expression levels, and the phosphorylation pattern of the AR on hormone binding were identical in both siblings. However, 5alpha-reductase 2 activity was normal in genital skin fibroblasts from the phenotypic male patient but undetectable in genital skin fibroblasts from the phenotypic female patient. The lack of 5alpha-reductase 2 activity was due to absent or reduced expression of 5alpha-reductase 2 in genital skin fibroblasts from the phenotypic female patient. Exon and flanking intron sequences of the 5alpha-reductase 2 gene showed no mutations in either sibling. Additional intragenic polymorphic marker analysis gave no evidence for different inherited alleles for the 5alpha-reductase 2 gene in the two siblings. Therefore, the absent or reduced expression of 5alpha-reductase 2 is likely to be additional to the AIS. Distinct phenotypic variation in this family was caused by 5alpha-reductase 2 deficiency, additional to AIS. This 5alpha-reductase deficiency is due to absence of expression of the 5alpha-reductase iso-enzyme 2 as shown by molecular studies. The distinct phenotypic variation in AIS here is explained by differences in the availability of 5alpha-dihydrotestosterone during embryonic sex differentiation.

A normal appendix found during diagnostic laparoscopy should not be removed.

BACKGROUND: Diagnostic laparoscopy has been introduced as a new diagnostic tool for suspected appendicitis. While the normal appendix used to be removed routinely, laparoscopy allows us to leave a normal looking appendix in place. This latter strategy is, however, not generally accepted. The long-term results of not removing a normal looking appendix were evaluated. METHODS: This was a prospective evaluation of 109 diagnostic laparoscopies for suspected appendicitis in which a normal looking appendix was left in place. After a median follow-up of 4.4 years a telephone questionnaire was performed. RESULTS: There were no false-negative laparoscopies. In 65 patients (60 per cent) another diagnosis was obtained (group 1). In 44 patients (40 per cent) no diagnosis was obtained (group 2). After a median interval of 8 months, 15 patients presented to the emergency department for symptoms possibly involving the appendix, during the median follow-up of 4.4 years. This resulted in readmission of nine patients, of whom eight were reoperated. In only one patient (1 per cent) was a histologically proven appendicitis found and the appendix removed. Some 105 patients were eligible for follow-up. Of the 100 patients interviewed (95 per cent), nine patients (9 per cent) (six in group 1 and three in group 2) still had recurrent pain in the right lower abdominal quadrant. There were no differences between patients with or without another diagnosis obtained during preceding laparoscopy. CONCLUSION: It is safe to leave a normal looking appendix in place when a diagnostic laparoscopy for suspected appendicitis is performed, even if another diagnosis cannot be found at laparoscopy.

Selective use of diagnostic laparoscopy in patients with suspected appendicitis.

BACKGROUND: Diagnostic laparoscopy has been introduced as a new diagnostic tool for patients with acute appendicitis. We performed diagnostic laparoscopy when the clinical diagnosis of appendicitis was in doubt. The aims of this study were to evaluate this strategy and to analyze the efficacy of diagnostic laparoscopy in patients with suspected appendicitis. PATIENTS AND METHODS: All patients referred to our hospital with suspected appendicitis during the period 1994-1997 were evaluated prospectively. The clinical diagnosis was determined by the surgeon or resident on call based on the patient's history, physical examination, and leukocyte count. The patients were divided into three groups: group 1: appendicitis not likely. These patients were observed for 24 h or discharged. When they showed signs of appendicitis in 24 h, they were transferred to either group 2 or 3; group 2: doubt concerning diagnosis. These patients underwent diagnostic laparoscopy, and appendectomy was performed if indicated; group 3: In these patients the diagnosis appendicitis was felt to be certain. They were treated by primary appendectomy by an open procedure. In this study, 1,050 patients, 531 women (51%), 389 men (37%), and 130 children (12%) <11 yrs, were evaluated. RESULTS: Altogether, 377 diagnostic laparoscopies were performed, leaving 109 healthy-looking appendices in place. This reduced the negative appendectomy rate from 25% to 14% in all surgically managed patients. The negative appendectomy rate for the women in group 2 was reduced from 49% to 14%, and for the men from 22% to 11%, so it also seemed worthwhile to perform diagnostic laparoscopy in men. Because the appendix sana was left in place in only three children, the benefit from laparoscopy is relatively small for children. In 48% of these patients a second diagnosis was obtained, most of them gynecologic in nature. There were no false-negative laparoscopies and no complications resulting from the laparoscopic procedure. CONCLUSIONS: Diagnostic laparoscopy is a safe procedure that reduced the appendix sana rate without increasing the total number of operations. It is a useful method for obtaining other, mostly gynecologic, diagnoses. To further reduce the appendix sana rate, better criteria for laparoscopic assessment of the appendix are needed.

17Beta-hydroxysteroid dehydrogenase-3 deficiency: diagnosis, phenotypic variability, population genetics, and worldwide distribution of ancient and de novo mutations.

17Beta-hydroxysteroid dehydrogenase-3 (17betaHSD3) deficiency is an autosomal recessive form of male pseudohermaphroditism caused by mutations in the HSD17B3 gene. In a nationwide study on male pseudohermaphroditism among all pediatric endocrinologists and clinical geneticists in The Netherlands, 18 17betaHSD3-deficient index cases were identified, 12 of whom initially had received the tentative diagnosis androgen insensitivity syndrome (AIS). The phenotypes and genotypes of these patients were studied. Endocrine diagnostic methods were evaluated in comparison to mutation analysis of the HSD17B3 gene. RT-PCR studies were performed on testicular ribonucleic acid of patients homozygous for two different splice site mutations. The minimal incidence of 17betaHSD3 deficiency in The Netherlands and the corresponding carrier frequency were calculated. Haplotype analysis of the chromosomal region of the HSD17B3 gene in Europeans, North Americans, Latin Americans, Australians, and Arabs was used to establish whether recurrent identical mutations were ancient or had repeatedly occurred de novo. In genotypically identical cases, phenotypic variation for external sexual development was observed. Gonadotropin-stimulated serum testosterone/androstenedione ratios in 17betaHSD3-deficient patients were discriminative in all cases and did not overlap with ratios in normal controls or with ratios in AIS patients. In all investigated patients both HSD17B3 alleles were mutated. The intronic mutations 325 + 4;A-->T and 655-1;G-->A disrupted normal splicing, but a small amount of wild-type messenger ribonucleic acid was still made in patients homozygous for 655-1;G-->A. The minimal incidence of 17betaHSD3 deficiency in The Netherlands was shown to be 1: 147,000, with a heterozygote frequency of 1:135. At least 4 mutations, 325 + 4;A-->T, N74T, 655-1;G-->A, and R80Q, found worldwide, appeared to be ancient and originating from genetic founders. Their dispersion could be reconstructed through historical analysis. The HSD17B3 gene mutations 326-1;G-->C and P282L were de novo mutations. 17betaHSD3 deficiency can be reliably diagnosed by endocrine evaluation and mutation analysis. Phenotypic variation can occur between families with the same homozygous mutations. The incidence of 17betaHSD3 deficiency is 0.65 times the incidence of AIS, which is thought to be the most frequent known cause of male pseudohermaphroditism without dysgenic gonads. A global inventory of affected cases demonstrated the ancient origin of at least four mutations. The mutational history of this genetic locus offers views into human diversity and disease, provided by national and international collaboration.

Mechanisms of androgen receptor activation and function.

Androgens play a crucial role in several stages of male development and in the maintenance of the male phenotype. Androgens act in their target cells via an interaction with the androgen receptor, resulting in direct regulation of gene expression. The androgen receptor is a phosphoprotein and modulation of the phosphorylation status of the receptor influences ligand-binding and consequently transcription activation of androgen responsive genes. Androgen binding induces a conformational change in the ligand-binding domain, accompanied by additional receptor phosphorylation. Subsequently the liganded androgen receptor interacts with specific androgen response elements in the regulatory regions of androgen target genes, resulting in stimulation of gene expression. Anti-androgens induce a different conformational change of the ligand-binding domain, which does not or only partially result in stimulation of transactivation. Interestingly, different anti-androgens can induce different inactive conformations of the androgen receptor ligand-binding domain. Recent evidence strongly supports a ligand dependent functional interaction between the ligand-binding domain and the NH2-terminal transactivating domain of the androgen receptor. Two regions in the NH2-terminal domain are involved in this interaction, whereas in the ligand-binding domain the AF-2 AD core region is involved.