A novel CYP17A1 deletion causes a functional knockout of the steroid enzyme 17-hydroxylase and 17,20-lyase in a Turkish family and illustrates the precise role of the CYP17A1 gene

A novel homozygous long-range deletion of the CYP17A1 gene abolished protein expression and caused the severest form of 17-hydroxylase deficiency in one kindred of a Turkish family. The affected subjects presented with 46,XY sex reversal and 46,XX lack of pubertal development as well as severe hypertension.

Favourable long-term outcome after immediate treatment of neonatal hyperammonemia due to N-acetylglutamate synthase deficiency

INTRODUCTION: N-Acetylglutamate synthase (NAGS) deficiency is a rare urea cycle disorder, which may present in the neonatal period with severe hyperammonemia and marked neurological impairment. CASE REPORT: We report on a Turkish family with a patient who died due to hyperammonemia in the neonatal period. Reduced activity of NAGS and carbamyl phosphate synthetase were found at autopsy. A second child who developed hyperammonemia on the second day of life was immediately treated with arginine hydrochloride, sodium benzoate and protein restriction. After NAGS deficiency was suspected by enzyme analysis, sodium benzoate was replaced by N-carbamylglutamate (NCG). A third child who developed slight hyperammonemia on the third day of life was treated with NCG before enzyme analysis confirmed reduced NAGS activity. Neither of the patients developed hyperammonemia in the following years. After the human NAGS gene was identified, mutation analysis revealed that the older sibling on NCG therapy was homozygous for a 971G>A (W324X) mutation. The parents and the younger sibling were heterozygous. Therapy was continued in the older sibling until now without any adverse effects and favourable neurodevelopment outcome. In the younger sibling, therapy was stopped without any deterioration of urea cycle function. CONCLUSION: NAGS deficiency can be successfully treated with NCG and arginine hydrochloride with favourable outcome. Molecular diagnostic rather than enzyme analysis should be used in patients with suspected NAGS deficiency.

Carcinoma ex pleomorphic adenoma in a minor salivary gland: report of a case

BACKGROUND: Carcinoma ex pleomorphic adenoma is exceedingly rare in minor salivary glands of the oral cavity. We present a case of carcinoma ex pleomorphic adenoma (CEPA) of the buccal mucosa in a 47-year-old Turkish patient. The buccal mass was of a size of 1.5 cm located in the left cheek. Pleomorphic adenoma was the tentative diagnosis. METHODS: The tumor was removed under local anesthesia. Histopathologic evaluation revealed a preexisting pleomorphic adenoma associated with adenoid tumor component with tubulo-cystic and papillary or pseudopapillary structures; CEPA was diagnosed. Capsular integrity was incomplete with infiltration by islands of metaplastic/dysplastic epithelium. RESULTS: Secondary surgery of the site was performed. No tumor tissue could be detected in the resection specimen. The patient is free of recurrence since 9 months.

Expression and function of 5-hydroxytryptamine 4 receptors in smooth muscle preparations from the duodenum, ileum, and pelvic flexure of horses without gastrointestinal tract disease

OBJECTIVE: To evaluate the expression of the 5-hydroxytryptamine 4 (5-HT4) receptor subtype and investigate the modulating function of those receptors on contractility in intestinal tissues obtained from horses without gastrointestinal tract disease. SAMPLE POPULATION: Smooth muscle preparations from the duodenum, ileum, and pelvic flexure collected immediately after slaughter of 24 horses with no history or signs of gastrointestinal tract disease. PROCEDURES: In isometric organ baths, the contractile activities of smooth muscle preparations in response to 5-hydroxytryptamine and electric field stimulation were assessed; the effect of tegaserod alone or in combination with 5-hydroxytryptamine on contractility of intestinal specimens was also investigated. Presence and distribution of 5-HT4 receptors in intestinal tissues and localization on interstitial cells of Cajal were examined by use of an immunofluorescence technique. RESULTS: Widespread 5-HT4 receptor immunoreactivity was observed in all intestinal smooth muscle layers; 5-HT4 receptors were absent from the myenteric plexus and interstitial cells of Cajal. In electrical field-stimulated tissue preparations of duodenum and pelvic flexure, tegaserod increased the amplitude of smooth muscle contractions in a concentration-dependent manner. Preincubation with tegaserod significantly decreased the basal tone of the 5-HT-evoked contractility in small intestine specimens, compared with the effect of 5-HT alone, thereby confirming that tegaserod was acting as a partial agonist. CONCLUSIONS AND CLINICAL RELEVANCE: In horses, 5-HT4 receptors on smooth muscle cells appear to be involved in the contractile response of the intestinal tract to 5-hydroxytryptamine. Results suggest that tegaserod may be useful for treatment of reduced gastrointestinal tract motility in horses.

Deletion of P399_E401 in NADPH cytochrome P450 oxidoreductase results in partial mixed oxidase deficiency

P450 oxidoreductase (POR) is the electron donor for all microsomal P450s including steroidogenic enzymes CYP17A1, CYP19A1 and CYP21A2. We found a novel POR mutation P399_E401del in two unrelated Turkish patients with 46,XX disorder of sexual development. Recombinant POR proteins were produced in yeast and tested for their ability to support steroid metabolizing P450 activities. In comparison to wild-type POR, the P399_E401del protein was found to decrease catalytic efficiency of 21-hydroxylation of progesterone by 68%, 17α-hydroxylation of progesterone by 76%, 17,20-lyase action on 17OH-pregnenolone by 69%, aromatization of androstenedione by 85% and cytochrome c reduction activity by 80%. Protein structure analysis of the three amino acid deletion P399_E401 revealed reduced stability and flexibility of the mutant. In conclusion, P399_E401del is a novel mutation in POR that provides valuable genotype-phenotype and structure-function correlation for mutations in a different region of POR compared to previous studies. Characterization of P399_E401del provides further insight into specificity of different P450s for interaction with POR as well as nature of metabolic disruptions caused by more pronounced effect on specific P450s like CYP17A1 and aromatase.

Fatal myocardial infarction at 4.5 years in a case of homozygous familial hypercholesterolaemia

Management of homozygous familial hypercholesterolaemia is notoriously difficult. For these patients, LDL apheresis is considered the treatment of choice. Treatment initiation is advocated generally from the age of seven years onwards (Thompson et al., Atherosclerosis 198:247-255, 2008). Here, we present the case of a young girl from a large inbred family of Turkish descent with homozygous familial hypercholesterolaemia and fatal outcome at the early age of 4(1/2) years.In conclusion, this case suggests that management of homozygous familial hypercholesterolaemia may require earlier and more aggressive treatment, including LDL apheresis before the age of seven years.

An in vitro study on spontaneous myometrial contractility in the mare during estrus and diestrus

Uterine smooth muscle specimens were collected from euthanatized mares in estrus and diestrus. Longitudinal and circular specimens were mounted in organ baths and the signals transcribed to a Grass polygraph. After equilibration time and 2 g preload, their physiologic isometric contractility was recorded for a continuous 2.0 h. Area under the curve, frequency and time occupied by contractions were studied. Differences between cycle phases, between muscle layers, and over the recorded time periods were statistically evaluated using linear mixed-effect models. In the mare, physiologic contractility of the uterus decreased significantly over time for all variables evaluated (time as covariate on a continuous scale). For area under the curve, there was a significant effect of muscle layer (longitudinal > circular). For frequency, higher values were recorded in estrus for circular smooth muscle layer, whereas higher values were seen in longitudinal smooth muscle layers during diestrus. In longitudinal layer and in diestrus, more time was occupied by contractions than in circular layer, and in estrus. This study is describing physiologic myometrial motility in the organ bath depending on cycle phase.

Membrane microdialysis: Evaluation of a new method to assess splanchnic tissue metabolism

OBJECTIVE: Measuring peritoneal lactate concentrations could be useful for detecting splanchnic hypoperfusion. The aims of this study were to evaluate the properties of a new membrane-based microdialyzer in vitro and to assess the ability of the dialyzer to detect a clinically relevant decrease in splanchnic blood flow in vivo. DESIGN: A membrane-based microdialyzer was first validated in vitro. The same device was tested afterward in a randomized, controlled animal experiment. SETTING: University experimental research laboratory. SUBJECTS: Twenty-four Landrace pigs of both genders. INTERVENTIONS: In vitro: Membrane microdialyzers were kept in warmed sodium lactate baths with lactate concentrations between 2 and 8 mmol/L for 10-120 mins, and microdialysis lactate concentrations were measured repeatedly (210 measurements). In vivo: An extracorporeal shunt with blood reservoir and roller pump was inserted between the proximal and distal abdominal aorta, and a microdialyzer was inserted intraperitoneally. In 12 animals, total splanchnic blood flow (measured by transit time ultrasound) was reduced by a median 43% (range, 13% to 72%) by activating the shunt; 12 animals served as controls. MEASUREMENTS AND MAIN RESULTS: In vitro: The fractional lactate recovery was 0.59 (0.32-0.83) after 60 mins and 0.82 (0.71-0.87) after 90 mins, with no further increase thereafter. At 60 and 90 mins, the fractional recovery was independent of the lactate concentration. In vivo: Abdominal blood flow reduction resulted in an increase in peritoneal microdialysis lactate concentration from 1.7 (0.3-3.8) mmol/L to 2.8 (1.3-6.2) mmol/L (p = .006). At the same time, mesenteric venous-arterial lactate gradient increased from 0.1 (-0.2-0.8) mmol/L to 0.3 (-0.3 -1.8) mmol/L (p = .032), and mesenteric venous-arterial Pco2 gradients increased from 12 (8-19) torr to 21 (11-54) torr (p = .005). CONCLUSIONS: Peritoneal membrane microdialysis provides a method for the assessment of splanchnic ischemia, with potential for clinical application.

Palliative in-patient cancer treatment in an anthroposophic hospital: I. Treatment patterns and compliance with anthroposophic medicine

BACKGROUND: Complementary and alternative medicine (CAM) and most of all anthroposophic medicine (AM) are important features of cancer treatment in Switzerland. While the number of epidemiological investigations into the use of such therapies is increasing, there is a distinct lack of reports regarding the combination of conventional and CAM methods. PATIENTS AND METHODS: 144 in-patients with advanced epithelial cancers were enrolled in a prospective quality-of-life (QoL) study at the Lukas Klinik (LK), Arlesheim, Switzerland. Tumor-related treatment was assessed 4 months prior to admission, during hospitalization and 4 months after baseline. OBJECTIVE: We aimed at giving a detailed account of conventional, AM and CAM treatment patterns in palliative care, before, during and after hospitalization, with emphasis on compliance with AM after discharge. RESULTS: Certain conventional treatments featured less during hospitalization than before but were resumed after discharge (chemotherapy, radiotherapy, sleeping pills, psychoactive drugs). Hormone therapy, corticosteroids, analgesics WHO III and antidepressants remained constant. AM treatment consisted of Iscador? (mistletoe), other plant- or mineral-derived medication, baths, massage, eurythmy, art therapy, counseling and lactovegetarian diet. Compliance after discharge was highest with Iscador (90%) and lowest with art therapy (14%). Many patients remained in the care of AM physicians. Other CAM and psychological methods were initially used by 39.9% of patients. After 4 months, the use had decreased with few exceptions. CONCLUSION: During holistic palliative treatment in an anthroposophic hospital, certain conventional treatments featured less whereas others remained constant. After discharge, chemotherapy returned to previous levels, AM compliance remained high, the use of other CAM therapies low.

Keratin 8 sequence variants in patients with pancreatitis and pancreatic cancer

Keratin 8 (KRT8) is one of the major intermediate filament proteins expressed in single-layered epithelia of the gastrointestinal tract. Transgenic mice over-expressing human KRT8 display pancreatic mononuclear infiltration, interstitial fibrosis and dysplasia of acinar cells resulting in exocrine pancreatic insufficiency. These experimental data are in accordance with a recent report describing an association between KRT8 variations and chronic pancreatitis. This prompted us to investigate KRT8 polymorphisms in patients with pancreatic disorders. The KRT8 Y54H and G62C polymorphisms were assessed in a cohort of patients with acute and chronic pancreatitis of various aetiologies or pancreatic cancer originating from Austria (n=16), the Czech Republic (n=90), Germany (n=1698), Great Britain (n=36), India (n=60), Italy (n=143), the Netherlands (n=128), Romania (n=3), Spain (n=133), and Switzerland (n=129). We also studied 4,234 control subjects from these countries and 1,492 control subjects originating from Benin, Cameroon, Ethiopia, Ecuador, and Turkey. Polymorphisms were analysed by melting curve analysis with fluorescence resonance energy transfer probes. The frequency of G62C did not differ between patients with acute or chronic pancreatitis, pancreatic adenocarcinoma and control individuals. The frequency of G62C varied in European populations from 0.4 to 3.8%, showing a northwest to southeast decline. The Y54H alteration was not detected in any of the 2,436 patients. Only 3/4,580 (0.07%) European, Turkish and Indian control subjects were heterozygous for Y54H in contrast to 34/951 (3.6%) control subjects of African descent. Our data suggest that the KRT8 alterations, Y54H and G62C, do not predispose patients to the development of pancreatitis or pancreatic cancer.

Intraindividual comparison of human radial and internal thoracic arteries in vitro and the effect of preoperative calcium blocker therapy

The patency rate of radial artery (RA) conduits is considerably lower than that of internal thoracic artery (ITA) grafts and the evidence suggests that this is due to a clinically suspected higher incidence of vasospasm. The aim of this study was, therefore, to compare intraindividually the pharmacological reactivity of RA with that of ITA. Both RA and ITA were taken from the same patients and investigated in parallel. Changes in tone were monitored isometrically on ring preparations from both arteries in organ baths with modified Krebs-Henseleit solution containing 1.25 mm calcium chloride at 1 g passive preload. In intraindividual comparisons maximal receptor-mediated contractile responses to noradrenaline and endothelin-1 and endothelium-dependent acetylcholine-induced relaxant responses revealed no differences between both arteries. By contrast, depolarization-induced contractions to potassium chloride (KCl) appeared to be significantly higher in RA than in ITA. Further analysis, however, revealed that this difference was due to preoperative calcium entry blocker (Ca(2+)eB) therapy. Compared with control tissues, maximal responses to KCl were significantly attenuated in the ITA but unchanged in RA when arteries were obtained from patients with preoperative Ca(2+)eB therapy. The present results suggested that functional responses to pharmacological stimuli of both RA and ITA were quite similar. Preoperative Ca(2+)eB therapy, however, attenuated markedly responses to KCl of the ITA leaving those of RA unchanged. These results, demonstrating a lower sensitivity to Ca(2+)eB of RA, therefore suggested that in addition to Ca(2+)eB other classes of drug may be more effective at reducing the propensity of RA conduits to vasospasm.