Characterization of large deletions in the DHCR7 gene.

Pathogenic variants in the DHCR7 gene cause Smith-Lemli-Opitz syndrome (SLOS), a defect of cholesterol biosynthesis resulting in an autosomal recessive congenital metabolic malformation disorder. In approximately 4% of patients, the second mutation remains unidentified. In this study, 12 SLOS patients diagnosed clinically and/or by elevated 7-dehydrocholesterol (7-DHC) have been investigated by customized multiplex ligation-dependent probe amplification (MLPA) analysis, because only one DHCR7 sequence variant has been detected. Two unrelated patients of this cohort carry different large deletions in the DHCR7 gene. One patient showed a deletion of exons 3-6. The second patient has a deletion of exons 1 and 2 (non-coding) and lacks the major part of the promoter. These two patients show typical clinical and biochemical phenotypes of SLOS. Second disease-causing mutations are p.(Arg352Trp) and p.(Thr93Met), respectively. Deletion breakpoints were characterized successfully in both cases. Such large deletions are rare in the DHCR7 gene but will resolve some of the patients in whom a second mutation has not been detected.

Craniofrontonasal syndrome in a male due to chromosomal mosaicism involving EFNB1: further insights into a genetic paradox.

Craniofrontonasal syndrome (CFNS) is an X-linked disorder caused by inactivating mutations in the gene for ephrin-B1 (EFNB1). Paradoxically it shows a more severe phenotype in females than in males. As a result of X inactivation cell populations with and without EFNB1 expression are found in EFNB1+/- females. This is thought to initiate a process termed cellular interference which may be responsible for the phenotype in females. We present a boy with severe clinical features of CFNS. In ∼42% of his blood cells we found a supernumerary ring X chromosome containing EFNB1 but lacking XIST. Mosaicism for cell populations with different levels of EFNB1 expression can explain the severe phenotype of this patient. In vitro experiments in Xenopus tissue showed that cells overexpress ephrinB1 cluster and sort out from wild-type cells. Our report provides further evidence that cellular interference contributes to the paradoxical inheritance pattern of CFNS.

Improved GMP-compliant multi-dose production and quality control of 6-[18F]fluoro-L-DOPA.

BACKGROUND: 6-[18F]Fluoro-L-3,4-dihydroxyphenylalanine (FDOPA) is a frequently used radiopharmaceutical for detecting neuroendocrine and brain tumors and for the differential diagnosis of Parkinson's disease. To meet the demand for FDOPA, a high-yield GMP-compliant production method is required. Therefore, this study aimed to improve the FDOPA production and quality control procedures to enable distribution of the radiopharmaceutical over distances.FDOPA was prepared by electrophilic fluorination of the trimethylstannyl precursor with [18F]F2, produced from [18O]2 via the double-shoot approach, leading to FDOPA with higher specific activity as compared to FDOPA which was synthesized, using [18F]F2 produced from 20Ne, leading to FDOPA with a lower specific activity. The quality control of the product was performed using a validated UPLC system and compared with quality control with a conventional HPLC system. Impurities were identified using UPLC-MS. RESULTS: The [18O]2 double-shoot radionuclide production method yielded significantly more [18F]F2 with less carrier F2 than the conventional method starting from 20Ne. After adjustment of radiolabeling parameters substantially higher amounts of FDOPA with higher specific activity could be obtained. Quality control by UPLC was much faster and detected more side-products than HPLC. UPLC-MS showed that the most important side-product was FDOPA-quinone, rather than 6-hydroxydopa as suggested by the European Pharmacopoeia. CONCLUSION: The production and quality control of FDOPA were significantly improved by introducing the [18O]2 double-shoot radionuclide production method, and product analysis by UPLC, respectively. As a result, FDOPA is now routinely available for clinical practice and for distribution over distances.

Obstructive sleep apnoea in craniofacial microsomia: analysis of 755 patients.

A retrospective cohort study was set up to analyse the prevalence and treatment of obstructive sleep apnoea (OSA) in relation to the severity of the deformity in patients with craniofacial microsomia (CFM). This study included a population of 755 patients with CFM from three craniofacial centres. Medical charts were reviewed for severity of the deformity, types of breathing difficulty, age at which breathing difficulty first presented, treatment for OSA, and treatment outcome. In total, 133 patients (17.6%) were diagnosed with OSA. Patients with Pruzansky IIB/III classification or bilateral craniofacial microsomia were significantly more often diagnosed with OSA than unilaterally affected patients of Pruzansky I/IIA classification. The initial treatment of OSA consisted of adenotonsillectomy, tracheotomy, or non-invasive positive pressure ventilation. Thirty-seven patients received more than one treatment (range 1-3). In this study, the prevalence of OSA in patients with CFM was higher than the prevalence in the healthy population described in the literature. Although several treatment modalities are available for the treatment of OSA in patients with CFM, treatment should be individualized and based on clinical symptoms, the severity of the deformity, and comorbidities.

Studies of the interaction between human protein S and human C4b-binding protein using deletion variants of recombinant human protein S.

Human protein S interacts noncovalently with human C4b-binding protein (C4BP). We have studied this interaction using deletion variants of recombinant human protein S. Two deletion variants were constructed by restriction enzyme digestion and in vitro site-specific mutagenesis of the human protein S cDNA. The variants were stably expressed in C127 cells. Recombinant proteins were purified using Fast Flow Q anion-exchange chromatography. The activated protein C (APC) cofactor activity, C4BP binding properties and reactivity to different monoclonal antibodies against human protein S were examined. The first variant (E variant), which has a deletion of the third epidermal growth factor (EGF)-like domain (deletion of exon VII, corresponding to amino acid residues ASP-160 to Asp-202) expresses normal APC cofactor activity in a plasma system. This activity was inhibited by the addition of purified C4BP. The second variant (L variant), which has a deletion of the C-terminal loop of the sex hormone binding globulin (SHBG)-like domain (deletion of exon XV, corresponding to amino acid residues Asp-583 to Ser-635) also expresses normal APC cofactor activity in plasma. This activity could only be partially inhibited by the addition of purified C4BP. Binding of the recombinant proteins to C4BP was studied in a system using purified proteins. The E variant binds to C4BP with the same affinity similar as recombinant wild type protein S (apparent Kd approximately 10(-10) M). The L variant, however, shows a markedly reduced affinity for binding to C4BP (apparent Kd approximately 10(-7) M).(ABSTRACT TRUNCATED AT 250 WORDS)

Synthesis and evaluation of radiolabeled antagonists for imaging of beta-adrenoceptors in the brain with PET.

Five potent, lipophilic beta-adrenoceptor antagonists (carvedilol, pindolol, toliprolol and fluorinated analogs of bupranolol and penbutolol) were labeled with either carbon-11 or fluorine-18 and evaluated for cerebral beta-adrenoceptor imaging in experimental animals. The standard radioligand for autoradiography of beta-adrenoceptors, [125I]-iodocyanopindolol, was also included in this survey. All compounds showed either very low uptake in rat brain or a regional distribution that was not related to beta-adrenoceptors, whereas some ligands did display specific binding in heart and lungs. Apparently, the criteria of a high affinity and a moderately high lipophilicity were insufficient to predict the suitability of beta-adrenergic antagonists for visualization of beta-adrenoceptors in the central nervous system.

A randomized comparison of transcervical Foley catheter to intravaginal misoprostol for preinduction cervical ripening.

OBJECTIVE: To compare the efficacy of intravaginal misoprostol tablets with transcervical Foley catheter for preinduction cervical ripening. METHODS: Pregnant women who presented for induction of labor with unfavorable cervices (Bishop score less than 6) were assigned randomly to intravaginal misoprostol (50 microg tablet every 4 hours for a maximum of six doses) or 30-mL Foley catheter placed transcervically with maintenance of traction. RESULTS: Among 111 women, 53 were allocated to misoprostol and 58 to Foley bulb. Contractile abnormalities were more frequent in the misoprostol group (20.4%) than the Foley group (0%) (P <.001). No statistically significant differences were noted between groups in change in Bishop score, preinduction cervical ripening times, and total induction times. There were no statistically significant differences in mode of delivery or adverse neonatal outcomes. Uterine rupture occurred in one woman with two previous cesarean deliveries in the misoprostol group. CONCLUSION: Intravaginal misoprostol and transcervical Foley catheter are equivalent for cervical ripening. Uterine contractile abnormalities and meconium passage are more common with misoprostol.

Impact of a pharmacist on drug costs in a coronary care unit.

The impact of clinical pharmacy services on direct drug costs in a coronary care unit (CCU) was studied. An observational, nonrandomized study was conducted on all patients admitted to the CCU to evaluate the impact of clinical pharmacy services on direct drug costs. Clinical pharmacy services were introduced into the CCU in July 1998. Patient characteristics, mean drug costs per admission, mean drug category costs per admission, and total hospital costs per admission were determined for October 1997 to June 1998 (nonintervention period), July 1998 to March 1999 (intervention period 1), and April 1999 to December 1999 (intervention period 2). The Clini-Trend program was used to estimate the total reduction in drug costs associated with documented pharmacist interventions from January to December 1999. Mean patient age, sex, admitting diagnosis-related group, Medicare case-mix index, ventilator days, length of stay, and number of deaths did not differ significantly among the three study periods. Mean +/- S.D. drug costs per admission for the nonintervention period were $374.05 +/- $75.51. With the introduction of clinical pharmacy services, mean +/- S.D. drug costs per admission were $381.94 +/- $66.16 (p > 0.1 for intervention period 1 compared with the nonintervention period) and $233.74 +/- $84.16 (p = 0.002 for intervention period 2 compared with the nonintervention period). From January to December 1999, 4151 pharmacist interventions were documented. The estimated reduction in drug costs associated with the interventions totaled $372,384. A pharmacist's clinical services in the CCU allowed for significant estimated reductions in total drug costs.

Bat-eared fox behavioural ecology and the incidence of rabies in the Serengeti National Park.

This paper provides a brief introduction into some aspects of bat-eared fox biology and social organization that is important to understanding rabies transmission and disease management in susceptible wildlife species (Macdonald 1980; 1993). A detailed description of the effects of two rabies outbreaks on a population of known individuals in the Serengeti National Park is given, Inter- and intrasexual differences in adult mortality rates are reported and discussed.

An improved method for the preparation of [11C]verapamil.

This paper describes an improved preparation of [11C]verapamil by reaction of [11C]methyl triflate with desmethylverapamil. The optimal reaction temperature, amount of precursor and reaction time were assessed. With this method [11C]verapamil can be prepared with a reproducible radiochemical yield of 66 +/- 4% (EOB, based on [11C]methyltriflate). Total synthesis time was 60 min. Radiochemical purity was >99% and specific activities varied between 5 and 30TBq/mmol.

The need for protein containing quality control materials for blood pH and electrolyte analyzers.

In clinical chemistry quality control refers to monitoring of precision and accuracy of the performance of analytical methods. Calibration solutions and (matrixed) control solutions are used in transferring accuracy between definitive method, reference method and field methods. For this purpose aqueous (protein-free), protein-containing and serum-based types of quality control materials having different matrices are available. Here are presented differences in behaviour between aqueous (protein-free) and protein-containing materials. Potentiometry with an electrochemical cell is an often used field method to determine pH and activity of electrolytes with an Ion-Selective Electrode (ISE). Basically, measured e.m.f.'s of calibrators and sample are translated into the activity of the ionic species of the sample by means of the Nernst equation. Besides the standard e.m.f. (E(zero)) of the electrochemical system, the measured e.m.f. includes the ISE-membrane e.m.f. (EISE) and depends on electrolyte type and its concentration and Eij depends on the composition and geometry of the salt bridge. Both EISE and Eij depend on the sample matrix. Protein-containing samples cause a negative bias on the e.m.f. at the liquid junction and a positive bias at the ISE. When EISE and Eij are compared to the mV span of the reference interval, the effects are large for Ca2+, Na+ and Cl-. Exactly the same effects exist for H+, K+, Li+ and Mg2+ but the inaccuracy is less critical for these ions. It may be concluded that only protein containing controls can detect an error that occurs only with measurements on plasma specimens. In practice this means that calibration is checked with protein-free solutions, but that measurements in plasma are best checked with protein-containing solutions.

Evaluation of protein containing quality control materials for blood gas analysis.

Protein containing quality control (QC) material in ampoules for blood gas, pH and electrolyte analysers has been manufactured using buffered protein (Bovine Serum Albumine, BSA) solution with sodium bicarbonate and chloride salts. For comparison a similar QC material but without protein was manufactured. Results obtained with ampouled QC material depend on pre-analytical effects, on matrix effects and on the stability of the material. Pre-analytical variation occurs with closed and/or opened ampoules. The shaking rate of the ampoule must be high (vortexing) and the duration of shaking long enough (15 seconds) to give good reproducibility. Temperature coefficients of protein containing controls are equal to those of protein free controls when incubated at different temperatures. Vigorously shaking of the ampoule gives a protein foam layer resulting in stable values for pH, pCO2 and pO2 during maximally 6 minutes after opening of the ampoule. Concerning matrix effects the CO2 buffer capacity of protein containing QC material is slightly higher compared to that of protein free QC materials as determined by tonometry with CO2/air gas mixtures and measuring pH and plotting log pCO2 vs. pH. The O2 buffer capacity measured as the bias on a properly functioning blood gas analyser is smaller than the bias of protein free QC material. The protein containing quality control is stable for at least 12 months when stored refrigerated at 2-6 degrees C and 28 days at room temperature.

[Hypacusis in acquired hypothyroidism (author's transl)]. / Hypakusis bei erworbener Hypothyreose.

We examined 56 patients with latent or clinical hypothyroidism and a control group of 18 patients with acute hypothyroidism. We were especially interested in the localization and extent of hearing abnormalities. 45% of the patients with chronic hypothyroidism showed a hearing loss. 2 of these patients had a pure conductive loss, 7 a combined hearing loss and 16 a sensorineural hearing loss. We demonstrated hair cell damage in 13 of the patients with chronic hypothyroidism. This loss was mild to moderate in over 90% of the cases. Although several patients had basilar-cochlear hearing losses, no characteristic audiograms could be found. 50% of the patients showed a small improvement in hearing after thyroid substitution therapy. The results of our examinations are discussed in detail.

[Hypacusis in acquired hypothyroidism (author's transl)]. / Hypakusis bei erworbener Hypothyreose.

We examined 56 patients with latent or clinically manifest hypothyroidism and 18 patients with acute athyreosis as a control group. We were especially interested in the localization and extent of hearing abnormalities. 45% of the patients with chronic hypothyroidism showed a hearing loss. Of these, two patients had a pure conductive loss, seven a combined hearing loss and 16 a sensori-neural hearing loss. We demonstrated hair cell damage in 13 of the patients with chronic hypothyroidism. The hearing loss was mild to moderate in over 90% of the cases. Although several patients had a basocochlear hearing loss, no characteristic audiogram was found. There was a correlation between the conductive loss and the severity of the hypothyroidism. 50% of the patients showed a significant improvement in hearing after substitution therapy. The results of our examinations are discussed in detail.

[Allergic contact dermatitis due to Asteraceae (Compositae). Cross reaction with Liatris spicata]. / Allergische Kontaktdermatitis auf Asteraceae (Kompositen). Kreuzreaktion mit Liatris spicata.

Case reports of 2 florists suffering from contact dermatitis to Asteraceae, especially to Chrysanthema are described. In addition, in both patients allergic patch-test reactions to another Asteraceae species (Liatris spicata) could be demonstrated. This Asteraceae species is a native flower (Blazing Star) in the USA, but in Germany we find the flower in shops, and sometimes in parks and gardens. Genus Liatris contains about 30 species. Numerous sesquiterpene lactons are isolated from Liatris species which are probably cross reactive with sesquiterpene lactons from Asteraceae native to Europe.

[Remission behavior following low-dose 13-cis-retinoic acid in papulopustular acne]. / Remissionsverhalten nach niedrigdosierter 13-cis-Retinsäuretherapie bei Acne papulo-pustulosa.

64 patients suffering from severe papulopustular acne previously resistant to therapy were treated with 13-cis retinoic acid (isotretinoin) at a dosage of 0.05, 0.1, or 0.2 mg/kg body weight for 20 weeks. After treatment, we performed follow-up examinations for 12 months concerning remission and side effects. After six months, more than half of the patients were free of relapses, even those treated with low doses, although there was a trend in favor of the 0.2 mg/kg body weight dose. According to lesion counts, the therapeutic effect of isotretinoin was even better after discontinuation of the drug. Mucocutaneous side effects disappeared within four to six weeks. In severe papulopustular acne, previously showing inadequate response to therapy, at least 0.2 mg/kg body weight isotretinoin should be given to ensure a good therapeutic result. These findings correspond with those observed in the initial reduction of lesions.

[13-cis-retinoic acid. Low dosage oral use in acne papulopustulosa. Results of a multicenter study]. / 13-cis-Retinsäure. Niedrig dosierte orale Anwendung bei Acne papulopustulosa. Ergebnisse einer multizentrischen Studie.

Successful treatment of acne conglobata with 13-cis-retinoic acid (isotretinoin, Ro 4-3780) has already been reported in this journal [25, 36]. The aim of the present study was to treat severe forms of papulopustular acne, unresponsive to conventional therapy, with low doses of 13-cis-retinoic acid to obtain good results with few side effects. A total of 191 patients from 14 departments of dermatology in the Federal Republic of Germany received 13-cis-retinoic acid under open randomized conditions in parallel dose groups of 0.05, 0.1, and 0.2 mg/kg body weight for 20 weeks in order to establish efficacy and tolerance. All inflammatory and non-inflammatory skin lesions were counted. The intensity of seborrhea was graded by using a scale. Adverse effects as well as laboratory values were registered. After 20 weeks of treatment a 79% (0.05 mg), 80% (0.1 mg), and 84% (0.2 mg) decrease in the number of inflammatory skin lesions was seen. Fourteen patients in the lowest dose group were considered to be dropouts. The decrease in non-inflammatory skin lesions was less marked and amounted to between 49% and 69%. A significant reduction of seborrhea could be observed in all patients. The main side effect was dryness of the skin and mucous membranes, which was, however, of low intensity. The elevation of triglyceride and cholesterol levels reported with higher doses of 13-cis-acid, especially in patients with high-risk factors, were not encountered in this study.

Early transvaginal ultrasonography versus early cerclage in women with an unclear history of incompetent cervix.

OBJECTIVE: Our aim was to compare outcomes in women with a questionable history of incompetent cervix, followed up with early transvaginal ultrasonography, with outcomes in women who had early cerclage. STUDY DESIGN: Charts were reviewed and patients identified for incompetent cervix from our obstetric database from 1995 through 1997. We included women who had an unclear history of incompetent cervix as follows: second-trimester loss or termination, > or =3 first-trimester terminations, cone biopsy or loop electrosurgical excision, or exposure to diethylstilbestrol. The primary outcome variable was gestational age at delivery. RESULTS: A total of 106 women were included, 45 in the early cerclage group and 61 in the early transvaginal ultrasonography group. The mean gestational age at delivery was 35.1 weeks for the early cerclage group versus 36.1 weeks for the early transvaginal ultrasonography group. CONCLUSION: In women with an unclear history of incompetent cervix, early cerclage does not appear to offer significant benefit over early transvaginal ultrasonography.

A comparative bioavailability study of carbamazepine tablets and a chewable tablet formulation.

Differences in product formulations have been shown to affect the therapeutic response by altering the relative bioavailability and pharmacokinetics of a drug. The relative bioavailability and pharmacokinetics of carbamazepine tablets (CBZ) and a chewable tablet formulation were evaluated in 10 normal healthy subjects (five men and five women). The study utilized a randomized, crossover design with a 4-week washout period between doses. Blood samples were collected at 0, 1, 2, 3, 4, 6, 8, 10, 14, 24, 30, 36, and 48 h following a 200-mg dose. Plasma samples were assayed by fluorescence polarization immunoassay. Ke, Cmax, Tmax, area under the curve (AUC), and relative bioavailability were estimated using traditional pharmacokinetic methods and compared by paired t test. A statistically significant higher Cmax (3.81 +/- 81 vs. 4.64 +/- .80 mg/L) was observed with the chewable tablet formulation but was not thought to be clinically relevant. No significant differences between formulations for Ke (0.022 +/- 0.007 vs. 0.025 +/- 0.008 h-1 h), Tmax (7.49 +/- 2.69 vs. 6.04 +/- 2.7 h), AUC 48 h (119 +/- 22 vs. 133 +/- 13 mg/h/L), or AUCO--infinity ( 221 +/- 40 vs. 203 +/- 41 mg/h/L) were noted. Absorption was variable for both preparations. The relative bioavailability using the tablet as the standard formulation was (0.92 +/- 0.22). Transient, mild side effects were noted in three subjects with the chewable tablet alone, and one subject experienced side effects with both formulations. It was concluded that CBZ tablets and chewable tablets may be used interchangeably; however, considerable intra- and intersubject variability exists, and the need for patient monitoring is emphasized.

Lyophilized bovine hemoglobin as a possible reference material for the determination of hemoglobin derivatives in human blood.

We investigated the suitability of a lyophilized bovine hemoglobin (LBH) preparation containing various fractions of oxyhemoglobin (O2Hb), carboxyhemoglobin (COHb), and methemoglobin (MetHb) for quality assessment in multicomponent analysis (MCA) of hemoglobin derivatives. It was demonstrated that a stable preparation of these components after reconstitution yields a hemoglobin solution that is spectrophotometrically equivalent with a fresh bovine hemoglobin solution. The preparation was found to be stable for at least 1 year when it is kept at 2-8 degrees C and for 1 h after reconstitution. We determined the fractions of O2Hb, COHb, and MetHb of several LBH preparations, using the complete spectra of 480-650 nm with 2-nm intervals and absorptivities as determined for pure LBH solutions. A field trial involving various types of multiwavelength hemoglobin photometers showed the suitability of LBH as a quality-control material. Computer models of the various common multiwavelength hemoglobin photometers may be useful for establishing more accurate target values of LBH preparations for each type of photometer and for studying the importance of the influence of specific factors such as wavelength selection, absorptivity values, and interfering dyes.