TPO (Thyroid Peroxidase) is known to be one of the major genes involved in congenital hypothyroid patients with thyroid dyshormonogenesis. The present study aims to validate high-resolution melting (HRM) curve analysis as a substitute method for Sanger sequencing, focusing on the frequently observed non-synonymous mutations c.1117G>T, c.1193G>C, and c.2173A>C in the TPO gene in patients from Bangladesh. We enrolled 36 confirmed cases of congenital hypothyroid patients with dyshormonogenesis to establish the HRM method. Blood specimens were collected, and DNA was extracted followed by PCR and Sanger sequencing. Among the 36 specimens, 20 were pre-sequenced, and variants were characterized through Sanger sequencing. Following pre-sequencing, the 20 pre-sequenced specimens underwent real-time PCR-HRM curve analysis to determine the proper HRM condition for separating the three variations from the wild-type state into heterozygous and homozygous states. Furthermore, 16 unknown specimens were subjected to HRM analysis to validate the method. This method demonstrated a sensitivity and specificity of 100 percent in accurately discerning wild-type alleles from both homozygous and heterozygous states of c.1117G>T (23/36; 63.8%), c.1193G>C (30/36; 83.3%), and c.2173A>C (23/36; 63.8%) variants frequently encountered among 36 Bangladeshi patients. The HRM data was found to be similar to the sequencing result, thus confirming the validity of the HRM approach for TPO gene variant detection. In conclusion, HRM-based molecular technique targeting variants c.1117G>T, c.1193G>C, and c.2173A>C could be used as a high throughput, rapid, reliable, and cost-effective screening approach for the detection of all common mutations in TPO gene in Bangladeshi patients with dyshormonogenesis.
Congenital Hypothyroidism , Humans , Bangladesh , Congenital Hypothyroidism/diagnosis , Congenital Hypothyroidism/genetics , Mutation , DNA , Real-Time Polymerase Chain Reaction
Objectives: This study determined the baseline hypothalamic-pituitary-adrenal axis hormonal levels and their associated factors in noncritically ill hospitalized patients with coronavirus disease 2019 (COVID-19). Methodology: This cross-sectional study was carried out in 91 noncritical RT-PCR-confirmed COVID-19 patients (aged 18 to 65 years) recruited consecutively from the COVID unit of two tertiary care hospitals over a period of six months. After the screening, relevant history and physical examinations were done, and blood was drawn between 07:00 am to 09:00 am in a fasting state to measure serum cortisol and plasma adrenocorticotropic hormone (ACTH) by chemiluminescent microparticle immunoassay. Results: Of 91 patients, 54, 26, and 11 had mild, moderate, and severe COVID-19, respectively. Median values of serum cortisol (p = 0.057) and plasma ACTH (p = 0.910) were statistically similar among the severity groups. Considering a cortisol cut-off of 276 nmol/L (<10 µg/dL), the highest percent of adrenal insufficiency was present in severe (27.3%), followed by mild (25.9%) and least in the moderate (3.8%) COVID-19 cases. Using the cortisol/ACTH ratio >15, only 6.6% had enough reserve. Conclusions: The adrenocortical response was compromised in a significant percentage of noncritically ill hospitalized patients with COVID-19, with the highest percentage of adrenal insufficiency present in severely infected cases. The HPA axis parameters of serum cortisol, plasma ACTH and cortisol/ACTH were similar across the severity of noncritical patients with COVID-19.
Adrenal Insufficiency , COVID-19 , Humans , Hypothalamo-Hypophyseal System/physiology , Hydrocortisone , SARS-CoV-2 , Cross-Sectional Studies , Pituitary-Adrenal System/physiology , Adrenocorticotropic Hormone , Adrenal Insufficiency/diagnosis , 17-Hydroxycorticosteroids
BACKGROUND: In individuals with coronavirus disease 2019 (COVID-19), male subjects have consistently been linked to poor severity and prognosis. Data on sex hormones in non-critical COVID-19-infected patients are scarce. The aim of this study was to assess the status of total testosterone (TT) and dehydroepiandrosterone sulfate (DHEAS) among noncritical patients with COVID-19 according to sex and their associations with clinical and biochemical features. MATERIALS AND METHODS: This cross-sectional observational study was done in the COVID-19 unit of a University hospital during the period of September 2021 to February 2022 among 91 adults (18-65 years) with reverse transcriptase- polymerase chain reaction confirmed noncritical COVID-19 patients. Blood was drawn by venipuncture before receiving steroids between 07:00 to 09:00 a.m. in a fasting state to measure serum TT and DHEAS by chemiluminescent microparticle immunoassay. Diagnosis and classification of COVID-19 were done according to World Health Organization's interim guidance. Age- and sex-specific laboratory reference values were used to classify the TT and DHEAS status of the patients. RESULTS: Only three males (8.1%) had low TT and the rest had normal TT. On the other hand, 15 (27.8%) of the females had high TT with normal levels in the rest. Similarly, 11 (29.7%) males had low DHEAS. Females had low, normal, and high DHEAS in four (7.4%), 48 (88.9%), and two (3.7%) cases respectively. Males with moderate severity of COVID-19 had significantly lower DHEAS (post hoc P=0.038) than the mild group. Both TT (P=0.008) and DHEAS (P=0.023) significantly correlated with neutrophils/lymphocytes ratio and only DHEAS with platelets/lymphocytes ratio (P=0.044) in males. In females, TT significantly correlated with serum sodium (P=0.034). CONCLUSION: In noncritical COVID-19 patients, substantial gender variations in TT and DHEAS were detected and correlated with severity markers in males.
OBJECTIVE: Polycystic ovary (PCO), a diagnostic component of polycystic ovary syndrome (PCOS), requires either an ovarian volume (OV) criterion or a follicle number per ovary (FNPO) criterion. This study investigated the association of OV and FNPO criteria with various manifestations of PCOS. METHODS: This cross-sectional study was conducted at a university hospital among 100 patients newly diagnosed with PCOS (according to the revised Rotterdam criteria). Fasting blood samples were collected to measure glucose, total testosterone (TT), luteinizing hormone (LH), follicle-stimulating hormone (FSH), lipid, insulin, and hemoglobin A1c levels. An oral glucose tolerance test was performed. Transabdominal or transvaginal ultrasound of the ovaries was done, depending on patients' marital status. All investigations were conducted in the follicular phase of the menstrual cycle. OV >10 mL and/or FNPO ≥12 indicated PCO. A homeostasis model assessment of insulin resistance (IR) value ≥2.6 indicated IR, and metabolic syndrome (MS) was defined according to the international harmonization criteria. RESULTS: Seventy-six participants fulfilled the OV criterion, 70 fulfilled the FNPO criterion, and 89 overall had PCO. Both maximum OV and mean OV had a significant correlation with TT levels (
Turner syndrome (TS) is the most common cause of short stature and delayed puberty in females. Approximately half of the patients have the classic form with a genotype of 45,XO, one-fourth of patients have different mosaic forms, and the remaining one-fourth have structural abnormalities on the X chromosome. Among the structural abnormalities, the most common is isochromosome Xq. Females with structural variants of TS can present with delayed menarche, amenorrhea, and infertility rather than classic manifestations of TS. This study describes two rare variants of TS. One was a structural abnormality on the X chromosome, 46X,iso(Xq), and the other involves a mosaic variety of TS, including isochromosome X in the form of 45,XO/46X,iso(Xq). Both patients presented with short stature and secondary amenorrhea without classic manifestations of TS. In TS with or without mosaicism, the frequency of isochromosomes is reported to be about 15% to 18%. Owing to the absence of classical manifestations of TS, diagnosis may be delayed or missed. Therefore, females of short stature with secondary amenorrhea should be evaluated for rare variants of TS by chromosomal analysis.
Background: Both visceral adiposity index (VAI) and lipid accumulation product (LAP) can be used to assess insulin resistance (IR) and metabolic syndrome (MetS) which are required for management of even lean polycystic ovary syndrome (PCOS) (body mass index [BMI] <23 kg/m2). Aim: This study was aimed to see the magnitude of associations of VAI and LAP with cardiometabolic risk factors including IR and MetS in lean PCOS. Study Setting and Design: This cross-sectional study was done amongst 62 newly detected lean PCOS patients and 58 age- and BMI-matched healthy controls. Materials and Methods: PCOS was diagnosed according to the Revised 2003 Rotterdam Consensus criteria. Along with relevant clinical data, fasting blood was taken to measure glucose, insulin and lipid profile by glucose oxidase, chemiluminescent microparticle immunoassay and by glycerol phosphate dehydrogenase-peroxidase method, respectively. IR was calculated by homeostasis model of IR (HOMA-IR). VAI and LAP were calculated from BMI, waist circumference, triglyceride and high-density lipoprotein cholesterol by using sex-specific formulae. Statistical Analysis Used: Linear and binary regression analyses and receiver operating characteristics curve (ROC) analyses were done as appropriate. Results: Only LAP had predictive associations with age, systolic and diastolic blood pressure and total and low-density lipoprotein cholesterol. Both VAI and LAP had predictive associations with history of subfertility and MetS. LAP had moderate discriminating index for IR with cut-off of HOMA-IR of 2.3. Both VAI and LAP had excellent discriminating index for MetS in lean PCOS patients. Conclusions: LAP had more associations with cardiometabolic risks than VAI and was a moderate discriminator of IR in lean PCOS.
Although thyroid dyshormonogenesis (TDH) accounts for 10-20% of congenital hypothyroidism (CH), the molecular etiology of TDH is unknown in Bangladesh. Thyroid peroxidase (TPO) is most frequently associated with TDH and the present study investigated the spectrum of TPO mutations in Bangladeshi patients and analyzed the effects of mutations on TPO protein structure through in silico approach. Sequencing-based analysis of TPO gene revealed four mutations in 36 diagnosed patients with TDH including three nonsynonymous mutations, namely, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, and one synonymous mutation p.Pro715Pro. Homology modelling-based analysis of predicted structures of MPO-like domain (TPO142-738) and the full-length TPO protein (TPO1-933) revealed differences between mutant and wild type structures. Molecular docking studies were performed between predicted structures and heme. TPO1-933 predicted structure showed more reliable results in terms of interactions with the heme prosthetic group as the binding energies were -11.5 kcal/mol, -3.2 kcal/mol, -11.5 kcal/mol, and -7.9 kcal/mol for WT, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, respectively, implying that p.Ala373Ser and p.Thr725Pro mutations were more damaging than p.Ser398Thr. However, for the TPO142-738 predicted structures, the binding energies were -11.9 kcal/mol, -10.8 kcal/mol, -2.5 kcal/mol, and -5.3 kcal/mol for the wild type protein, mutant proteins with p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro substitutions, respectively. However, when the interactions between the crucial residues including residues His239, Arg396, Glu399, and His494 of TPO protein and heme were taken into consideration using both TPO1-933 and TPO142-738 predicted structures, it appeared that p.Ala373Ser and p.Thr725Pro could affect the interactions more severely than the p.Ser398Thr. Validation of the molecular docking results was performed by computer simulation in terms of quantum mechanics/molecular mechanics (QM/MM) and molecular dynamics (MD) simulation. In conclusion, the substitutions mutations, namely, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, had been involved in Bangladeshi patients with TDH and molecular docking-based study revealed that these mutations had damaging effect on the TPO protein activity.
Autoantigens/genetics , Congenital Hypothyroidism/genetics , Iodide Peroxidase/genetics , Iron-Binding Proteins/genetics , Mutation/genetics , Structure-Activity Relationship , Adolescent , Amino Acid Substitution/genetics , Autoantigens/chemistry , Bangladesh/epidemiology , Child , Child, Preschool , Computer Simulation , Congenital Hypothyroidism/epidemiology , Congenital Hypothyroidism/pathology , Female , Genotype , Humans , Iodide Peroxidase/chemistry , Iron-Binding Proteins/chemistry , Male , Models, Molecular , Molecular Docking Simulation , Phenotype , Thyroid Gland/metabolism , Thyroid Gland/pathology
