Polymorphisms in the glucocerebrosidase gene and pseudogene urge caution in clinical analysis of Gaucher disease allele c.1448T>C (L444P)

BACKGROUND: Gaucher disease is a potentially severe lysosomal storage disorder caused by mutations in the human glucocerebrosidase gene (GBA). We have developed a multiplexed genetic assay for eight diseases prevalent in the Ashkenazi population: Tay-Sachs, Gaucher type I, Niemann-Pick types A and B, mucolipidosis type IV, familial dysautonomia, Canavan, Bloom syndrome, and Fanconi anemia type C. This assay includes an allelic determination for GBA allele c.1448T>C (L444P). The goal of this study was to clinically evaluate this assay. METHODS: Biotinylated, multiplex PCR products were directly hybridized to capture probes immobilized on fluorescently addressed microspheres. After incubation with streptavidin-conjugated fluorophore, the reactions were analyzed by Luminex IS100. Clinical evaluations were conducted using de-identified patient DNA samples. RESULTS: We evaluated a multiplexed suspension array assay that includes wild-type and mutant genetic determinations for Gaucher disease allele c.1448T>C. Two percent of samples reported to be wild-type by conventional methods were observed to be c.1448T>C heterozygous using our assay. Sequence analysis suggested that this phenomenon was due to co-amplification of the functional gene and a paralogous pseudogene (ΨGBA) due to a polymorphism in the primer-binding site of the latter. Primers for the amplification of this allele were then repositioned to span an upstream deletion in the pseudogene, yielding a much longer amplicon. Although it is widely reported that long amplicons negatively impact amplification or detection efficiency in recently adopted multiplex techniques, this assay design functioned properly and resolved the occurrence of false heterozygosity. CONCLUSION: Although previously available sequence information suggested GBA gene/pseudogene discrimination capabilities with a short amplified product, we identified common single-nucleotide polymorphisms in the pseudogene that required amplification of a larger region for effective discrimination

Lived religion: Rethinking human nature in a neoliberal age

This article considers the relationship between philosophy of religion and an approach to the study of religion, which prioritises the experience of lived religion. Considering how individuals and communities live out their faith challenges some of the assumptions of analytic philosophers of religion regarding the position the philosopher should adopt when approaching the investigation of religion. If philosophy is understood principally as a means for analysing belief, it will have little space for an engagement with what it feels like to live out one’s faith

21-hydroxylase deficiency: From molecular genetics to clinical presentation

Congenital adrenal hyperplasia due to deficiency of the enzyme 21-hydroxylase (21-OH), a cytochrome P450 enzyme located in the endoplasmic reticulum and which catalyzes the conversion of 17-hydroxyprogesterone to 11-deoxycortisol and progestene to deoxycorticosterone, is distinguished in its classical and non-classical form and is also one of the most common autosomal recessive inherited diseases in humans. The classical form appears in a rate between 1:5000 and 1:15,000 among the live neonates of North America and Europe, while the non-classical form occurs in approximately 0.2% of the general white population. This rate is especially high between the Ahskenazi Jews and a part (ie Italians, Hispanics) of the Mediterranean populations. Three alleles are associated with the 21-OH locus and can be combined in several ways in individuals who are either unaffected, heterozygote carriers, or affected with classical or non-classical disease. Variable signs and symptoms of hyperandrogenism, such as hirsutism, acne, virilization of the external genitalia and/or the body, short stature, menstrual irregularities, are common to both types of the disorder. Among the genes responsible for the synthesis of the enzyme 21-OH and the antigens of HLA system, exist both a proven genetic linkage and a proven genetic linkage disequilibrium. HLA-B-w47, HLAB(5) and HILA-B-35 are the most common haplotypes usually met in the classical form, while the haplotype HLA-B14DR1, is the most recurrent in the non-classical form of the disease. The significant advances in molecular biology and gene analysis over the past two decades have led to the development of novel sensitive methods of DNA analysis and study, such as polymerase chain reaction and southern blot analysis. Thus, it has been revealed that the synthesis of enzyme 21-OH is controlled by two genes, the active CYP21B gene and the CYP21A pseudogene. All three forms of the disease have a known sequence of gene changes owing to mutations in isolated proteins or whole series of genes due to translocations or deletions of genetic material

Congenital adrenal hyperplasia because of 21-Hydroxylase deficiency a genetic disorder of interest to obstetricians and gynecologists

Congenital adrenal hyperplasia (CAH) due to deficiency of the enzyme 21 -hydroxylase (21 -OH), is distinguished in its classical and nonclassical form and is one of the most common autosomal recessive inherited diseases in humans. The classical form appears between 1:5000 and 1:15000 among the live neonates of North America and Europe, whereas the nonclassical form occurs in approximately 0.2% of the general white populations. Three alleles are associated with the 21-OH locus and can be combined in various ways to individuals who are either unaffected, heterozygote carriers, or affected with the classical or nonclassical disease. Variable signs and symptoms of hyperandrogenism are common to both types of the disorder. In women with CAH, hyperandrogenism may be present, extending from virilization of external genitalia and salt-wasting in classical (C)-CAH cases, to menstrual irregularity, obesity, short stature, infertility or subfertility and skin disorders such as hirsutism, in nonclassical (NC)-CAH cases. These clinical characteristics of NC-CAH cases do not differ unmarkedly from those shown in patients with polycystic ovary syndrome, idiopathic hirsutism, or hyperinsulinemia. The significant advances in molecular biology and gene analysis over the past 2 decades have led to the development of novel sensitive methods of DNA analysis and study, including polymerase chain reaction and Southern blot analysis. Thus it has been revealed that the 21 -OH gene (CYP21A2) and its nonfunctional pseudogene (CYP21A1P) are located on chromosome 6 (6p21.3), sharing a high homology of about 98%. Inactivating mutations occur as complete gene deletions, large gene conversions and pseudogene-derived mutations. Copyright © 2009 by Lippincott Williams & Wilkins

A Thermodynamics Model for the Assessment and Optimisation of Onboard Natural Gas Reforming and Carbon Capture

Abstract The paper examines pre-combustion carbon capture technology (PreCCS) for liquefied natural gas (LNG) propelled shipping from thermodynamics and energy efficiency perspectives. Various types of LNG reformers and CCS units are considered. The steam methane reformer (SMR) was found to be 20% more energy efficient than autothermal (ATR) and methane pyrolysis (MPR) reactors. Pressure swing adsorption (PSA) had a lower energy requirement than membrane separation (MEM), cryogenic separation (CS), and amine absorption (AA) in pre-combustion carbon capture, with PSA needing 0.18 kWh/kg CO2. An integrated system combining SMR and PSA was proposed using waste heat recovery (WHR) from the engine, assuming similar efficiency for LNG and H2 operation, and cooling and liquefying of the CO2 by the LNG. The SMR-PSA system without WHR had an overall efficiency of 33.4% (defined as work at the propeller divided by the total LNG energy consumption). This was improved to 41.7% with WHR and gave a 65% CO2 emission reduction. For a higher CO2 reduction, CCS from the SMR heater could additionally be employed, giving a maximum CO2 removal rate of 86.2% with 39% overall energy efficiency. By comparison, an amine-based post-engine CCS system without reforming could reach similar CO2 removal rates but with 36.6% overall efficiency. The advantages and disadvantages and technology readiness level of PreCCS for onboard operation are discussed. This study offers evidence that pre-combustion CCS can be a serious contender for maritime propulsion decarbonization. Graphical Abstract National Research Foundation (NRF), Prime Minister’s Office, Singapore, under its Campus for Research Excellence and Technological Enterprise (CREATE) programm

The impact of mild hypercholesterolemia on glycemic and hormonal profiles, menstrual characteristics and the ovarian morphology of women with polycystic ovarian syndrome

The severity of polycystic ovarian syndrome (PCOS) has been clearly associated with insulin resistance, obesity and metabolic syndrome. The purpose of the present cross-sectional study is to investigate whether mild hypercholesterolemia alters the biochemical and clinical profile of PCOS patients. Our study is based on a prospectively collected population of women of reproductive age who were diagnosed with PCOS according to the definition of the Rotterdam European Society of Human Reproduction and Embryology/American Society for Reproductive Medicine (ASRM/ESHRE) criteria. For the correlation analysis we used the non-parametric Spearman's rank correlation coefficient. Partial correlation was also performed to control for potential confounders observed in the univariate analysis. Overall, 235 patients were included. Their mean age ranged between 14 and 45 years old and the body mass index (BMI) between 17 and 54. Women with mild hypercholesterolemia had a higher BMI and their fasting insulin was increased as well as indices of insulin resistance [Homeostatic model assessment (HOMA), quantitative insulin sensitivity check index (QUICKI), Matsuda index] compared to women with PCOS with normal cholesterol levels. Correlation statistics suggested that the effect of serum lipids on the hormonal profile of patients was weak. Both low-density lipoprotein (LDL) and high-density lipoprotein (HDL) exerted a significant mild negative correlation to glucose and insulin. However, after controlling the results for BMI and age (the two variables that were found significantly different in the univariate analysis) we observed that this effect was non-significant. Mild hypercholesterolemia does not affect the hormonal profile of patients with PCOS; hence, to date, there is no evidence to suggest its treatment for the correction of menstrual and hormonal abnormalities in PCOS women. © 2018 Walter de Gruyter GmbH, Berlin/Boston

Subclinical hypothyroidism does not influence the metabolic and hormonal profile of women with PCOS

Subclinical hypothyroidism (SCH) is present in 5%-10% of polycystic ovary syndrome (PCOS) patients. To date, its impact on the metabolic and hormonal profile of those women remains controversial. The purpose of our study is to evaluate the impact of SCH on the glycemic, lipid and hormonal profile of PCOS patients. We conducted a prospective case control study of patients that attended the Department of Gynecological Endocrinology of our hospital. Overall, 280 women with PCOS were enrolled during a time period of 7 years (2009-2015). Twenty-one patients (7.5%) suffered from SCH. The anthropometric characteristics were comparable among women with PCOS and those with SCH + PCOS. The prevalence of acne, hirsutism and anovulation did not differ. Significant differences were observed in the 2-h oral glucose tolerance test (OGTT) (p = 0.003 for glucose and p = 0.046 for insulin). The QUICKI, Matsuda and homeostatic model assessment-insulin resistance (HOMA-IR) indices where, however, similar. No difference in serum lipids was observed. Slightly elevated levels of follicle stimulating hormone (FSH) and testosterone were noted. The remaining hormonal parameters remained similar among groups. Similarly, the ovarian volume and the endometrial thickness did not differ. The impact of SCH on the metabolic and hormonal profile of PCOS patients seems to be negligible. Future studies are needed in the field and their conduct in a multi-institutional basis seems to be required, given the small prevalence of SCH among women with PCOS. © 2017 2017 Walter de Gruyter GmbH, Berlin/Boston

The impact of the metabolic syndrome on bone mass density: A prospective case control study

Obesity and metabolic syndrome (MetS) during the perimenopausal period and in menopause have been linked to altered bone mass density (BMD) in various experimental studies. However, current clinical studies provide conflicting results in this field. The purpose of the present study was to evaluate this association. We conducted a prospective case control study that was based on a consecutive series of menopausal women who attended the Reproductive Endocrinology Outpatient Clinic of our hospital between January 2013 and December 2016. One hundred and forty post-menopausal women were included in the present study. After stratifying the women in two groups according to the presence of MetS we observed that bone turnover markers remained unaffected by the presence of MetS (p > 0.05). On the other hand, both the T- A nd Z-scores of women with MetS were significantly higher compared to healthly postmenopausal women [T-score: 0.4 (-0.7 to 1.3) vs.-1 (-1.62 to-0.1), p < 0.001] [Z-score: 0.55 (-0.3 to 1.7) vs.-0.4 (-1.1 to 0.4), p = 0.003]. According to the findings of our study the presence of MetS during the perimenopausal years seems to have a mild benefit on bone mass density. The pathophysiology that underlies this effect remains unclear as bone turnover markers seem to be unaffected by MetS. © 2018 2018 Walter de Gruyter GmbH, Berlin/Boston

New reliable biochemical marker for screening 21 alpha-hydroxylase deficiency without index person among hirsute women in agreement with HLA-haplotyping

Late onset congenital adrenal hyperplasia due to 21 alpha-hydroxylase deficiency (LO21OH def), as many other diseases, is the cause of hirsutism, menstrual disorders, infertility (PCO-like symptoms). We evaluated the reliability of a new biochemical marker for screening LO-21OH def in 47 women with PCO-like symptoms and 11 men, members of their families, comparing the results of separation using this new marker with those of HLA-haplotyping in 21 members of the patient population. All subjects were stimulated with 0.25 mg synthetic ACTH iv. Serum progesterone (P), 17-hydroxyprogesterone (17-OHP) and cortisol (F) at 0, 15, 30, 45 and 60 min following ACTH administration were determined and the new marker, namely the difference between 60min and 0min of the ratio F/17-OHP [Delta F/17-OHP (60 min -0 min)] was calculated. According to the established biochemical criteria for the detection of LO-21OH def cases, (Gurai 30 min greater than or equal to 12 ng/dl/min and 17-OHP 60 min greater than or equal to 12 ng/ml for severe 21-OH def and Gutai 30 min<6.5 ng/dl/min and 17-OHP 60 min<5 ng/ml for ‘’healthy” individuals regarding 21-OH def) two groups, A and B respectively, were separated from the patient population. In group A (n=8), with LO-21OH def, the new marker showed negative values in all cases, while in group B (n=9), without LO-21OH def, this marker was positive. The remaining subjects, depending on the results of the new marker were separated in 2 subgroups, C-neg (n=28), with negative values, composed, consequently, of members with 21-OH def and C-pos, (n=13), with positive values, composed, consequently, of subjects with absence of LO-21OH def. HLA-typing was in agreement with the results of screening by the new marker, in 20 out of 21 cases, while there was only one false negative result. In conclusion, the proposed biochemical marker Delta F/17-OHP (60 min-0 min) seems to be a reliable parameter for the LO-21OH def detection among young women with PCO-like symptoms as well as males suspected for congenital adrenal hyperplasia