Enzyme Defect Research Articles

Spotlight on Givosiran as a Treatment Option for Adults with Acute Hepatic Porphyria: Design, Development, and Place in Therapy.

Small interfering ribonucleic acids [siRNAs] are short ribonucleic acid (RNA) fragments cleaved from double-stranded RNA molecules that target and bind to specific sequences on messenger RNA (mRNA), leading to their destruction. Therefore, the siRNA down-regulates the formation of selected mRNAs and their protein products. Givosiran is one such siRNA that uses this mechanism to treat acute hepatic porphyrias. Acute hepatic porphyrias are a group of rare, inherited metabolic disorders, characterized by acute potentially life-threatening attacks as well as chronic symptoms with a negative impact on quality of life. It has four types, each associated with distinct enzyme defects in the heme biosynthesis pathway in the liver. By targeting the expression of hepatic 5-aminolevulinic acid [ALA] synthase-1 [ALAS1], givosiran can down-regulate levels of toxic metabolites, leading to biochemical and clinical improvement. Givosiran selectively targets hepatocytes due to its linkage to N-acetylgalactosamine (GalNac) leading to its selective uptake via asialoglycoprotein receptors (ASGPR). We provide an up-to-date literature review regarding givosiran in the context of a clinical overview of the porphyrias, an overview of siRNAs for therapy of human disorders, the design and development of givosiran, key clinical trial results of givosiran for prevention of acute porphyric attacks, emerging concerns regarding chronic use of givosiran, and the overall management of acute hepatic porphyrias. These insights are important not only for the management of acute hepatic porphyrias but also for the emerging field of siRNAs and their role in novel therapies for various diseases.

Glucose 6 phosphate dehydrogenase deficiency in unexplained neonatal hyperbilirubinemia – A study in neonatal care unit of a tertiary care hospital.

Background: Glucose 6 phosphate dehydrogenase (G6PD)deficiency causes impaired production of reduced glutathione and in turn exposes red blood cells to damage by oxidative metabolites with resultant hemolysis. Objective: To study the spectrum of neonatal hyperbilirubinemia , to investigate the relevance of G6PD deficiency in unexplained causes of neonatal hyperbilirubinemia and to look for outcome in cases of deficiency of the enzyme. Material & methods: Cross sectional observational study done on 100 neonates. The inclusion criteria was babies born between 37 and 42 completed weeks of gestation with clinically evident jaundice and those of age upto seven days requiring admission in neonatal care unit .Their age, gender, religion, socioeconomic and residential status noted. History elicited from mother following informed consent and babies were examined clinically.Investigations included complete hemogram, total bilirubin with conjugated and unconjugated assay, red blood cell G6PD assay, thyroid profile and coomb’s test Result: The mean age (mean±s.d.) of patients was 4.5100± 1.4460 days. The study population of 100 babies included 69 males and 31 females with a male to female ratio of 2.23:1. G6PD testing showed deficiency in fifteen patients, whereas eighty five showed normal values of G6PD. All patients in the study population showed normal values of T3, T4, TSH and negative direct coomb’s test. G6PD assay showed a mean value of 9.7291± 2.5480. The mean total bilirubin (mean±s.d.) of patients with deficient G6PD was 20.4533 ± 2.2853 mg/dl. G6PD deficient group showed higher serum bilirubin assay compared to non- deficient ones. Conclusion: G6PD deficiency is a common enzyme defect causing severe indirect hyperbilirubinemiain neonates which can result in kernicterus with neurological damage. Early neonatal screening programmes should be implementedin areas where the deficiency is prevalent. Bangladesh Journal of Medical Science Vol. 21 No. 03 July’22 Page: 669-674

Misdiagnosis in mucopolysaccharidoses.

Mucopolysaccharidosis (MPS) is a group of 13 hereditary metabolic diseases identified in humans (or 14 diseases if considering one MPS type described to date only in mice) in which an enzymatic defect results in the accumulation of glycosaminoglycans (GAG) in the lysosomes of cells. First of all, as a result of GAG storage, the proper functioning of the lysosome is disturbed; then, the cells, and finally, tissue, organs, and the whole organism malfunctions are observed. Due to the rarity, heterogeneity, and multi-systemic and progressive nature of MPS, they present a major diagnostic challenge. Due to the wide variation in symptoms and their similarity to other diseases, MPS is often misdiagnosed, usually as neurological diseases (like autism spectrum disorders, psychomotor hyperactivity, and intellectual disability) or rheumatology and orthopedic disorders (like juvenile idiopathic arthritis, Perthes disease, rickets, and muscular dystrophy). In this review article, we present the problems associated with the possibility of misdiagnosing MPS, discuss what diseases they can be confused with, and suggest ways to reduce these problems in the future.

Beta‐catenin inhibition as a novel therapeutic strategy for porphyria

Background– The porphyrias are metabolic disorders caused by enzymatic defects in heme biosynthesis, leading to excessive accumulation of porphyrins and their precursors in the liver or bone marrow, the most active tissues in heme biosynthesis. The specific patterns of accumulation of the heme precursors aminolevulinic acid (ALA) and other intermediate compounds define clinical features of these diseases such as acute neurovisceral attacks, skin lesions, and mental changes. These debilitating diseases remain incurable, and there is an unmet need to develop effective therapies to treat them. Because the liver is either a source or sink for porphyrins, we investigated the role of Wnt/beta‐catenin pathway in this disease. Wnt/beta‐catenin signaling is an evolutionarily conserved pathway that plays an important role in liver health and pathophysiology by regulating genes important in metabolism and proliferation.Methods– 3,5‐Diethoxycarbonyl‐1,4‐dihydrocollidine (DDC) induces hepatic porphyria in mice and is utilized to identify signaling pathways involved in injury progression. The metabolism of DDC creates a potent inhibitor of ferrochelatase (FC), the terminal enzyme in the heme pathway, which causes the buildup of porphyrin precursors. These precursors can cause cellular abnormalities such as oxidative stress, mitochondrial dysfunction, protein aggregation and inhibition of crucial functions such as autophagy. We studied pharmacological inhibition of Wnt/beta‐catenin signaling by utilizing a beta‐catenin DsiRNA formulated into a lipid nanoparticle in mice fed DDC diet.Results– Our data demonstrates that mice with pharmacologically inhibited beta‐catenin have decreased liver injury due to significantly fewer porphyrin deposits. This is due to suppression of ALA‐S and ALA‐D enzymes which catalyze the first and second steps of heme biosynthesis respectively. Significantly, we identified a strong TCF4 binding site in the intron region of the Alad gene, implicating ALA‐D as a direct Wnt/beta‐catenin target. Our data, therefore, establishes a novel role of the Wnt/beta‐catenin pathway in regulating heme biosynthesis by inhibiting early steps in heme biosynthesis pathway and reducing toxic porphyrin accumulation. Furthermore, our data reveal that mice lacking beta‐catenin have increased induction of autophagy over baseline that contributes to the protection from injury by clearing accumulated toxic porphyrins.Conclusions– These observations collectively offer a novel opportunity to remedy porphyria by targeting the Wnt/beta‐catenin signaling pathway. Our next studies will further validate ALA‐D as the mechanistic target of beta‐catenin in the heme biosynthesis pathway. Because prior research has established crosstalk between autophagy and the Wnt signaling pathway, we will also investigate the role of beta‐catenin in regulating autophagy during porphyria.

Multiplex Mass Spectrometry for Enzymatic Activities Screening of Lysosomal Storage Diseases

Lysosomal storage diseases (LSDs) are a group of rare and inherited disorders of lysosome function and they are classified into more than 50 types (e.g., mucopolysaccharidoses, neuronal ceroid lipofuscinoses, Fabry’s Disease; Pompe Disease, etc.). The majority of LSDs result from defective lysosomal enzyme activity leading to accumulation of endogenous macromolecules in the lysosome causing severe disease symptoms and finally death. For some LSDs, treatment is available in the form of enzyme replacement therapy with good results if the therapy is started as soon as possible, making the diagnosis of crucial importance. State of art now in LSDs diagnostic employs in first stage, enzymatic determinations by fluorimetry or by mass spectrometry (in few cases).In this report, we describe a highly specific and sensitive diagnostics on dry blood spots (DBSs) for (i) molecular determinations of LSDs, particularly mucopolysaccharidoses (MPSs) and neuronal ceroid lipofuscinoses (NCLs), by simultaneous fluorimetric and mass spectrometric analysis using new types of substrate derivatives based on different alkyl‐umbelliferone; (ii) clinical diagnostics of LSDs by multiplex‐MS‐MRM analysis using new type of substrate derivatives and (iii) clinical validation of the new developed methods using DBS samples for patients with established LSDs. We have developed a high‐throughput single and multiplexing assay for rare diseases (e.g. MPS 1, MPS 2, MPS 6, NCL 2, NCL 10, Fabry Disease, etc.) in DBSs. A common substrate for the fluorimetric measurement of a lysosomal enzyme activity is a molecule with 2 different parts which are bound covalently: a group recognized by the target enzyme (e.g. sugar, lipid) and 4‐umbelliferone and the product resulted after the enzymatic reaction is always 4‐methyl umbelliferone. In our tests, the enzymatic activity levels in DBSs were determinate by fluorimetry or multiple reactions monitoring mass spectrometry in the presence of an internal standard showing a good statistical correlation in singles assays. Moreover, we developed duplex and triplex assays for the diagnosis of different LSDs from the muccopolysacharidoses family using modified substrates based on different coumarin derivates obtained through Pechman condensation. The new umbelliferone derivatives were synthesize by substituting groups in position 4 and 7 of the alpha‐ coumarin molecule with others e.g. 4‐ ethyl‐ umbelliferone, 4‐propyl‐umbelliferone, etc.In conclusion, by quantification of umbelliferone derivatives as products of enzymatic reactions either by fluorimetry or mass spectrometry, the newly proposed substrates can be easily use for clinical diagnosis of several LSDs.

Understanding Inborn Errors of Metabolism through Metabolomics.

Inborn errors of metabolism (IEMs) are rare diseases caused by a defect in a single enzyme, co-factor, or transport protein. For most IEMs, no effective treatment is available and the exact disease mechanism is unknown. The application of metabolomics and, more specifically, tracer metabolomics in IEM research can help to elucidate these disease mechanisms and hence direct novel therapeutic interventions. In this review, we will describe the different approaches to metabolomics in IEM research. We will discuss the strengths and weaknesses of the different sample types that can be used (biofluids, tissues or cells from model organisms; modified cell lines; and patient fibroblasts) and when each of them is appropriate to use.

High Frequency of Glucose-6-Phosphate Dehydrogenase Deficiency in Patients Diagnosed with Celiac Disease.

Celiac disease (CD) is characterized by a proinflammatory state associated with the production of reactive oxygen species, i.e., a condition of oxidative stress. In this study, we tested the hypothesis that the inherited deficiency of glucose-6-phosphate dehydrogenase (G6PD), by causing impaired antioxidant defense, may increase the risk of CD. Methods: A retrospective monocentric case-control study was performed using the clinical records of 8338 outpatients (64.6% women) scheduled for upper endoscopy between 2002 and 2021 in Northern Sardinia. Overall, 627 were found to have CD (7.5%), and 1027 resulted to be G6PD-deficiency carriers (12.3%). Since randomization was impractical, the potential covariates imbalance between cases and controls was minimized using a 1:2 propensity-score-matched (PSM) analysis. Results: Overall, G6PD deficiency was associated with increased risk of CD (odds ratio (OR) 1.50; 95% confidence interval (CI) 1.19–1.90). The PSM procedure identified 1027 G6PD-deficient and 2054 normal patients. Logistic regression including the propensity score detected for G6PD deficiency an OR of 1.48 (95%CI 1.13–1.95; p = 0.004). Conclusions: Our findings show that the enzyme defect was significantly and positively associated with CD, in line with the pro-oxidant impact of the enzyme defect observed in animal models and humans.

Insights into ultrasonic treatment on the mechanism of proteolysis and taste improvement of defective dry-cured ham

To investigate the effects of ultrasonic treatment on proteolysis and taste development of defective dry-cured ham, sensory attributes, enzyme activities, protein degradation and free amino acids were evaluated after different ultrasonic treatments. The ultrasonic treatment of 1000 W & 50 °C significantly increased the intensities of overall taste, umami, sweetness and richness, and decreased bitterness values compared with other groups. The residual activities of DPP I and cathepsin B + L in 1000 W & 50 °C maintained 48.71% and 24.94% of control group, respectively; the intense degradation of structural proteins was observed by label-free proteomics, accordingly. The contents of total free amino acids from 4522.64 mg/100 g muscles in control group increased to 5838.75 mg/100 g muscles in 1000 W & 50 °C; the largest increase of sweet and umami amino acids observed in 1000 W & 50 °C was responsible for the improvement of taste quality of defective dry-cured ham.

3,4,5-Trihydroxypiperidine Based Multivalent Glucocerebrosidase (GCase) Enhancers.

The synthesis of five new multivalent derivatives of a trihydroxypiperidine iminosugar was accomplished through copper catalyzed alkyne‐azide cycloaddition (CuAAC) reaction of an azido ending piperidine and several propargylated scaffolds. The resulting multivalent architectures were assayed as inhibitors of lysosomal GCase, the defective enzyme in Gaucher disease. The multivalent compounds resulted in much more potent inhibitors than a parent monovalent reference compound, thus showing a good multivalent effect. Biological investigation of these compounds as pharmacological chaperones revealed that the trivalent derivative (12) gives a 2‐fold recovery of the GCase activity on Gaucher patient fibroblasts bearing the L444P/L444P mutations responsible for neuropathies. Additionally, a thermal denaturation experiment showed its ability to impart stability to the recombinant enzyme used in therapy.

Corticosteroid suppresses urea-cycle-related gene expressions in ornithine transcarbamylase deficiency

BackgroundOrnithine transcarbamylase deficiency (OTCD) is most common among urea cycle disorders (UCDs), defined by defects in enzymes associated with ureagenesis. Corticosteroid administration to UCD patients, including OTCD patients, is suggested to be avoided, as it may induce life-threatening hyperammonemia. The mechanism has been considered nitrogen overload due to the catabolic effect of corticosteroids; however, the pathophysiological process is unclear.MethodsTo elucidate the mechanism of hyperammonemia induced by corticosteroid administration in OTCD patients, we analyzed a mouse model by administering corticosteroids to OTCspf−ash mice deficient in the OTC gene. Dexamethasone (DEX; 20 mg/kg) was administered to the OTCspf−ash and wild-type (WT) mice at 0 and 24 h, and the serum ammonia concentrations, the levels of the hepatic metabolites, and the gene expressions related with ammonia metabolism in the livers and muscles were analyzed.ResultsThe ammonia levels in Otcspf−ash mice that were administered DEX tended to increase at 24 h and increased significantly at 48 h. The metabolomic analysis showed that the levels of citrulline, arginine, and ornithine did not differ significantly between Otcspf−ash mice that were administered DEX and normal saline; however, the level of aspartate was increased drastically in Otcspf−ash mice owing to DEX administration (P < 0.01). Among the enzymes associated with the urea cycle, mRNA expressions of carbamoyl-phosphate synthase 1, ornithine transcarbamylase, arginosuccinate synthase 1, and arginosuccinate lyase in the livers were significantly downregulated by DEX administration in both the Otcspf−ash and WT mice (P < 0.01). Among the enzymes associated with catabolism, mRNA expression of Muscle RING-finger protein-1 in the muscles was significantly upregulated in the muscles of WT mice by DEX administration (P < 0.05).ConclusionsWe elucidated that corticosteroid administration induced hyperammonemia in Otcspf−ash mice by not only muscle catabolism but also suppressing urea-cycle-related gene expressions. Since the urea cycle intermediate amino acids, such as arginine, might not be effective because of the suppressed expression of urea-cycle-related genes by corticosteroid administration, we should consider an early intervention by renal replacement therapy in cases of UCD patients induced by corticosteroids to avoid brain injuries or fatal outcomes.

Differential expression of enzymes in thymidylate biosynthesis in zebrafish at different developmental stages: implications for dtymk mutation-caused neurodegenerative disorders

BackgroundDeoxythymidine triphosphate (dTTP) is an essential building block of DNA, and defects in enzymes involved in dTTP synthesis cause neurodegenerative disorders. For instance, mutations in DTYMK, the gene coding for thymidylate kinase (TMPK), cause severe microcephaly in human. However, the mechanism behind this is not well-understood. Here we used the zebrafish model and studied (i) TMPK, an enzyme required for both the de novo and the salvage pathways of dTTP synthesis, and (ii) thymidine kinases (TK) of the salvage pathway in order to understand their role in neuropathology.ResultsOur findings reveal that maternal-stored dNTPs are only sufficient for 6 cell division cycles, and the levels of dNTPs are inversely correlated to cell cycle length during early embryogenesis. TMPK and TK activities are prominent in the cytosol of embryos, larvae and adult fish and brain contains the highest TMPK activity. During early development, TMPK activity increased gradually from 6 hpf and a profound increase was observed at 72 hpf, and TMPK activity reached its maximal level at 96 hpf, and remained at high level until 144 hpf. The expression of dtymk encoded Dtymk protein correlated to its mRNA expression and neuronal development but not to the TMPK activity detected. However, despite the high TMPK activity detected at later stages of development, the Dtymk protein was undetectable. Furthermore, the TMPK enzyme detected at later stages showed similar biochemical properties as the Dtymk enzyme but was not recognized by the Dtymk specific antibody.ConclusionsOur results suggest that active dNTP synthesis in early embryogenesis is vital and that Dtymk is essential for neurodevelopment, which is supported by a recent study of dtymk knockout zebrafish with neurological disorder and lethal outcomes. Furthermore, there is a novel TMPK-like enzyme expressed at later stages of development.

Liver transplantation in glycogen storage disease: a single-center experience

BackgroundGlycogen storage diseases (GSDs) are inherited glycogen metabolic disorders which have various subtypes. GSDs of type I, III, IV, VI, and IX show liver involvement and are considered as hepatic types of GSDs. Thus, liver transplantation (LT) has been proposed as a final therapy for these types of GSD. LT corrects the primary hepatic enzyme defect; however, the long-term outcomes of LT in these patients have not been extensively evaluated so far. There are few reports in the English literature about the outcome of GSD patients after LT. There has been no report from Iran. The present retrospective study aimed to evaluate the long-term outcomes of eight patients with GSD types I, III, and IV who underwent LT in the affiliated hospitals of Shiraz University of Medical Sciences, from March 2013 to June 2021. During this period, there were no patients with GSD VI and IX identified in this center.ResultsThe median time of diagnosis of the GSDs and at transplant was 1 year and 11 years, respectively. All eight transplanted patients were alive at the time of follow-up in this study. None of them required a re-transplant. All of the patients showed normalized liver enzymes after LT with no sign of hypoglycemia.ConclusionsLT is an achievable treatment for end-stage hepatic involvement of GSDs with a cure for metabolic deficiency. Our experience in these eight patients shows a favorable outcome with no mortality and no major complication.

Combined Targeted and Untargeted Profiling of HeLa Cells Deficient in Purine De Novo Synthesis.

Three genetically determined enzyme defects of purine de novo synthesis (PDNS) have been identified so far in humans: adenylosuccinate lyase (ADSL) deficiency, 5-amino-4-imidazole carboxamide-ribosiduria (AICA-ribosiduria), and deficiency in bifunctional enzyme phosphoribosylaminoimidazole carboxylase and phosphoribosylaminoimidazolesuccinocarboxamide synthase (PAICS). Clinical signs of these defects are mainly neurological, such as seizures, psychomotor retardation, epilepsy, autistic features, etc. This work aims to describe the metabolic changes of CRISPR-Cas9 genome-edited HeLa cells deficient in the individual steps of PDNS to better understand known and potential defects of the pathway in humans. High-performance liquid chromatography coupled with mass spectrometry was used for both targeted and untargeted metabolomic analyses. The statistically significant features from the untargeted study were identified by fragmentation analysis. Data from the targeted analysis were processed in Cytoscape software to visualize the most affected metabolic pathways. Statistical significance of PDNS intermediates preceding deficient enzymes was the highest (p-values 10 × 10−7–10 × 10−15) in comparison with the metabolites from other pathways (p-values of up to 10 × 10−7). Disturbed PDNS resulted in an altered pool of adenine and guanine nucleotides. However, the adenylate energy charge was not different from controls. Different profiles of acylcarnitines observed among deficient cell lines might be associated with a specific enzyme deficiency rather than global changes related to the PDNS pathway. Changes detected in one-carbon metabolism might reduce the methylation activity of the deficient cells, thus affecting the modification state of DNA, RNA, and proteins.

Abstract 64: Clinical and hormonal profile of congenital adrenal hyperplasia from a tertiary centre in India

Background: Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders due to defect in enzymes involved in adrenal steroidogenesis with varied clinical presentations.Aims and Objectives: To understand the clinical and hormonal profile of individuals with CAH.Results: Twenty nine patients were included in this cross sectional study ( males (n) =15 (51.7%)) and females (n=14 (48.3%)) (adults=11) (born out of consanguineous marriage = 22 and 3 pairs of siblings). Thirteen patients had salt wasting (SW) phenotype, 14 had simple virilizing (SV) and 2 had non classical CAH (NCCAH). The presentation of SW phenotype was salt wasting crisis in males and ambiguous genitalia in females. The presentation of SV phenotype was clitoromegaly in females; precocious puberty and premature adrenarche in males. All the patients with NCCAH were females with virilization and primary amenorrhea. Majority of patients are on hydrocortisone (82.7%), the rest on prednisolone (13.7%) and dexamethasone (3.4%). Adult females with SV and NCCAH phenotypes had higher 17OHP levels at follow up. Mean adult height of patients with NCCAH was better than SV patients (154.5 cms vs 145.2 cms)Conclusions: The presentation of CAH may be varied but a screening at birth will help us to detect it early.

Advances in Thymidine Kinase 2 Deficiency: Clinical Aspects, Translational Progress, and Emerging Therapies.

Defects in the replication, maintenance, and repair of mitochondrial DNA (mtDNA) constitute a growing and genetically heterogeneous group of mitochondrial disorders. Multiple genes participate in these processes, including thymidine kinase 2 (TK2) encoding the mitochondrial matrix protein TK2, a critical component of the mitochondrial nucleotide salvage pathway. TK2 deficiency (TK2d) causes mtDNA depletion, multiple deletions, or both, which manifest predominantly as mitochondrial myopathy. A wide clinical spectrum phenotype includes a severe, rapidly progressive, early onset form (median survival: < 2 years); a less severe childhood-onset form; and a late-onset form with a variably slower rate of progression. Clinical presentation typically includes progressive weakness of limb, neck, facial, oropharyngeal, and respiratory muscle, whereas limb myopathy with ptosis, ophthalmoparesis, and respiratory involvement is more common in the late-onset form. Deoxynucleoside monophosphates and deoxynucleosides that can bypass the TK2 enzyme defect have been assessed in a mouse model, as well as under open-label compassionate use (expanded access) in TK2d patients, indicating clinical efficacy with a favorable side-effect profile. This treatment is currently undergoing testing in clinical trials intended to support approval in the US and European Union (EU). In the early expanded access program, growth differentiation factor 15 (GDF-15) appears to be a useful biomarker that correlates with therapeutic response. With the advent of a specific treatment and given the high morbidity and mortality associated with TK2d, clinicians need to know how to recognize and diagnose this disorder. Here, we summarize translational research about this rare condition emphasizing clinical aspects.

The Utility of Genomic Testing for Hyperphenylalaninemia.

Hyperphenylalaninemia (HPA), the most common amino acid metabolism disorder, is caused by defects in enzymes involved in phenylalanine metabolism, with the consequent accumulation of phenylalanine and its secondary metabolites in body fluids and tissues. Clinical manifestations of HPA include mental retardation, and its early diagnosis with timely treatment can improve the prognosis of affected patients. Due to the genetic complexity and heterogeneity of HPA, high-throughput molecular technologies, such as next-generation sequencing (NGS), are becoming indispensable tools to fully characterize the etiology, helping clinicians to promptly identify the exact patients’ genotype and determine the appropriate treatment. In this review, after a brief overview of the key enzymes involved in phenylalanine metabolism, we represent the wide spectrum of genes and their variants associated with HPA and discuss the utility of genomic testing for improved diagnosis and clinical management of HPA.

COMPARISON OF SINGLE DOSE AND MULTI-DOSE hCG STIMULATION TESTS

Objective: The human chorionic gonadotropin (hCG) stimulation test is widely used to assess testicular steroidogenesis. This test evaluates Leydig cell function and helps to evaluate testosterone synthesis in testicular enzyme defects, hypogonadism, 46,XY sexual development disorders (DSD), and micropenis. This study will compare testosterone responses to a single dose of 5000 IU/m2 and three consecutive days of 1500 IU/m2 hCG regimens. Materials and Methods: The study evaluated 18 patients who were admitted with micropenis, hypospadias, or undescended testis (Group 1) and 18 patients with a molecular diagnosis of androgen receptor insensitivity (56%) or 5α reductase deficiency (44%) (Group 2). The median (interquartile range; IQR) age of the patients were 1.4 (0.8-4.2) years for Group 1 and 0.7 (0.3-0.8) years for Group 2. Baseline testosterone levels were checked and after a single dose of hCG 5000 IU/m2 injection into the first group, testosterone levels were observed on day one and day four. In the second group, after the injection of hCG 1500 IU/m2 for three days, testosterone levels were checked on the fourth day. Results: There was no significant difference in height, weight, BMI SDS, and baseline testosterone levels between groups. After hCG, stimulation testosterone responses did not vary among the groups. Testosterone levels were increased 40-fold and 40.1- fold on day four compared to the baseline in Group 1 and Group 2. In Group 1, testosterone response was significantly higher on day four compared to day one (p=0.001). Conclusions: The hCG tests performed with a single dose or multidoses showed that there was no difference between testosterone responses and these two tests were applicable the 5000 IU/m2 hCG test requires fewer injections and is easy to administer, this test may be preferred to an evaluation of the Leydig cell function. For an adequate testosterone response, the researchers suggest measuring testosterone levels on the fourth day after the single dose of 5000 IU/m2 .

Differences in gene expression patterns, revealed by RNA-seq analysis, between various Sanfilippo and Morquio disease subtypes

Mucopolysaccharidoses (MPS) are genetic disorders that affect up to 1 in 25,000 births. They are caused by dysfunctions of lysosomal hydrolases that degrade glycosaminoglycans (GAGs) which accumulate in cells, damaging their proper functioning. There are 7 types of MPS, distinguished by the kind of accumulated GAG and the defective enzyme, which differ significantly in the course of the disease. Despite the storage of the same GAGs, two of them (MPS III and IV) are divided into subtypes. While the course of MPS IV A and B is similar, the variability between MPS III A, B, C and D is high. This suggests that there are additional aspects that could influence the course of the disease. Therefore, the aim of this study was to determine differences of patterns of gene expression between all MPS III and IV subtypes. Transcriptomic studies, carried out with dermal fibroblasts from patients with all MPS III and IV subtypes, showed a significant variation in the gene expression pattern between individual MPS III subtypes, in contrast to MPS IV. Detailed analysis of transcripts with altered expression levels between MPS III subtypes indicated that these transcripts are mainly involved in maintaining the proper structure of connective tissue (COL4A1, COL4A2, COMP) and the structure of ribosomes (RPL10, RPL23, RPLP2). The results presented in this study indicate a significant role of genetic factors in the diversified course of MPS III subtypes.

Inherited Disorders of Thyroid Hormone Metabolism Defect Caused by the Dysregulation of Selenoprotein Expression.

Consistent activation and functioning of thyroid hormones are essential to the human body as a whole, especially in controlling the metabolic rate of all organs and systems. Impaired sensitivity to thyroid hormones describes any process that interferes with the effectiveness of thyroid hormones. The genetic origin of inherited thyroid hormone defects and the investigation of genetic defects upon the processing of thyroid hormones are of utmost importance. Impaired sensitivity to thyroid hormone can be categorized into three conditions: thyroid hormone cell membrane transport defect (THCMTD), thyroid hormone metabolism defect (THMD), and thyroid hormone action defect (THAD). THMD is caused by defects in the synthesis and processing of deiodinases that convert the prohormone thyroxine (T4) to the active hormone triiodothyronine (T3). Deiodinase, a selenoprotein, requires unique translation machinery that is collectively composed of the selenocysteine (Sec) insertion sequence (SECIS) elements, Sec-insertion sequence-binding protein 2 (SECISBP2), Sec-specific eukaryotic elongation factor (EEFSEC), and Sec-specific tRNA (TRU-TCA1-1), which leads to the recognition of the UGA codon as a Sec codon for translation into the growing polypeptide. In addition, THMD could be expanded to the defects of enzymes that are involved in thyroid hormone conjugation, such as glucuronidation and sulphation. Paucity of inherited disorders in this category leaves them beyond the scope of this review. This review attempts to specifically explore the genomic causes and effects that result in a significant deficiency of T3 hormones due to inadequate function of deiodinases. Moreover, along with SECISBP2, TRU-TCA1-1, and deiodinase type-1 (DIO1) mutations, this review describes the variants in DIO2 single nucleotide polymorphism (SNP) and thyroid stimulating hormone receptor (TSHR) that result in the reduced activity of DIO2 and subsequent abnormal conversion of T3 from T4. Finally, this review provides additional insight into the general functionality of selenium supplementation and T3/T4 combination treatment in patients with hypothyroidism, suggesting the steps that need to be taken in the future.

High anion gap metabolic acidosis as a result of 5-oxoproline

High anion gap metabolic acidosis is a common acid–base disorder encountered in hospitalized patients and more so in the intensive care unit. The most common causes are accumulation of lactate, ketones, and urea and ingestion of certain toxins. In the absence of these, lesser-known etiologies such as accumulation of D-lactate or pyroglutamic acid (PGA) should be suspected. PGA (5-oxoproline) is an endogenous organic acid that can accumulate in serum to cause anionic gap metabolic acidosis. This can occur with inherited defects in enzymes that participate in the y-glutamyl cycle or can be acquired in association with therapeutic paracetamol use, malnutrition, sepsis, antibiotics, and renal impairment. Our case illustrates that prompt recognition of this entity and discontinuation of offending agents such as paracetamol result in rapid recovery, thus reducing the hospital stay.