Molecular genetic analysis of patients carrying steroid 21-hydroxylase deficiency in the Mexican population: identification of possible new mutations and high prevalence of apparent germ-line mutations.

Steroid 21-hydroxylase deficiency is the underlying cause in over 90% of patients with congenital adrenal hyperplasia, an inherited metabolic disorder of adrenal steroidogenesis. We have characterized 94 mutant alleles from 47 unrelated Mexican patients and the corresponding mutant alleles in their parents by amplification of the functional CYP21 gene by PCR, followed by direct sequence analysis. The study included patients diagnosed with the three clinical forms of the disease. Our results revealed: (1) the presence of relatively few mutations or combinations of mutations associated with particular phenotypes; (2) the presence of putative new mutations; (3) the finding of identical genotypes in patients displaying discordant phenotypes; (4) the identification of patients lacking all previous reported mutations; and (5) an apparent high frequency of germ-line mutations. The absence of previously reported mutations in about 22% of the disease alleles, the finding of putative new mutations in some of the patients lacking previously known mutations, and the apparent high prevalence of germ-line mutations make evident the differences in the genetic background leading to this disorder between the Caucasian and the Mexican populations.

Analysis of the glucokinase gene in Mexican families displaying early-onset non-insulin-dependent diabetes mellitus including MODY families.

Non-insulin-dependent diabetes mellitus (NIDDM) is the most common form of diabetes, affecting 5% of the general population. Genetic factors play an important role in the development of the disease. While in other populations NIDDM is usually diagnosed after the fifth decade of life, in Mexico a large proportion of patients develop the disease at an early age (between the third and the fourth decade). In Caucasian population, mutations in the glucokinase gene, the TCF1, and TCF14 genes, have been identified in a subgroup of early-onset NIDDM patients denominated MODY (maturity-onset diabetes of the young), which show an autosomal dominant pattern of inheritance. As a first step in the molecular characterization of Mexican families displaying early-onset NIDDM we searched for mutations in the glucokinase gene through SSCP analysis and/or direct sequencing in 26 individuals from 22 independent families, where at least four can be classified as MODY. No mutations were detected in the exons or the intron-exon boundaries of the gene in any of the screened individuals. The phenotype and clinical profile of some of the studied patients is compatible with that of patients carrying mutations in the TCF1 or TCF14 genes, while others may carry mutations in different loci. Through computer simulation analysis we identified at least four informative families which will be used for further linkage studies.

Fasting and postprandial serum insulin in Mexican adolescents with menstrual disorders.

PURPOSE: To investigate insulin serum concentrations in basal and stimulated conditions in a group of Mexican adolescents presenting menstrual disturbances. METHODS: A total of 77 post-menarchial adolescents were studied: 65 with a chronological age of 15 +/- 1.7 years (mean +/- SD) had persistent anovulation and represented the study group; 12 were normal ovulatory adolescents (15 +/- 1.2 years) and served as controls. Body mass index (BMI), waist to hip ratio (W/H), presence and severity of acne, hirsutism, acanthosis nigricans (AN) and follicular hyperkeratosis were recorded. Transabdominal pelvic ultrasound was performed and LH, FSH, estradiol, prolactin, testosterone, androstenedione and sex hormone binding globulin (SHBG) concentrations were measured in plasma by specific immunoassays. Glucose and insulin levels were determined in venous blood following an overnight fasting and two hours after a standardized breakfast. RESULTS: Anovulatory patients were divided in three groups depending on the presence of AN and overweight (BMI > 25). The insulin concentration in the study patients were remarkably higher than the values reported in the medical literature. Statistically significant differences were also found in fasting and postprandial insulin concentrations among the three anovulatory groups. Insulin values correlated with the severity of AN, W/H ratio, BMI and SHBG serum levels. CONCLUSIONS: Our study indicates that moderate to severe hyperinsulinemia is present in a high proportion of our adolescent anovulatory population. Whether hyperinsulinemia represents a transitory peripubertal event or is a predictive marker of chronic anovulation and metabolic derangement in adult life needs further investigation.

Low frequency of deletion alleles in patients with steroid 21-hydroxylase deficiency in a Mexican population.

Steroid 21-hydroxylase deficiency is caused by mutations in the CYP21 gene. Approximately 95% of mutant alleles are generated by recombination events between the active gene CYP21 and its highly homologous pseudogene, CYP21P. Deletion alleles are generated by unequal crossing over, while point mutations are the result of gene conversion events. Deletions account for 20-25% of the 21-hydroxylase deficiency alleles in most populations studied. We have looked for deletions among 53 unrelated Mexican patients with steroid 21-hydroxylase deficiency and found that deletions represent less than 1% of the disease alleles. These findings suggest that nearly all mutant alleles in our patient population contain point mutations and that the low representation of deletion alleles among clinically diagnosed patients may be due to missing detection of salt wasters, mainly males, who may die during the neonatal period.

Anticuerpos antitiroideos en hipotiroidismo congénito / Antithyroid antibodies in congenital hypothyroidism

El papel que los anticuerpos antitiroideos maternos pueden desempeñar en la etiopatogenia del hipotiroidismo congénito por atireosis es discutible. En este estudio se midieron los niveles séricos de antivuerpos antitiroideos en 12 pacientes hipotiroideos y en sus madres, con la finalidad de detectar la posible asociación entre enfermedad tiroidea autoinmune materna y displasia tiroidea fetal. En dos pacientes hubo anticuerpos antitiroglobulina, lo que apoya la teoría de la existencia de un proceso inmunológico contra la tiroides fetal en las primeras ocho semanas de festación, pero no descarta la existencia y/o coexistencia de otros factores etiopatogénicos

[Biosynthetic growth hormone extracted from mammalian cells used in the management of growth hormone deficient children]. / Hormona de crecimiento biosintética, extraída de células de mamífero en el manejo de niños con deficiencia de hormona de crecimiento.

Biosynthetic growth hormone extracted from mammalian cells was used in eight children with growth hormone deficiency, in search for a clinical safety study. Each child received 0.6 U/kg/week, subcutaneously, during a period of six months, and was evaluated monthly for clinical, anthropometric and laboratory modifications induced by the drug in a physiological or pathological way. The growth velocity increased from less that 4 cm/year to 11.2 +/- 1.08 cm/year, the bone age maintained according to the chronological age, and the anthropometric evaluation showed an harmonious and physiological growth. There was not any undesirable nor secondary changes except transient hypothyroidism in two patients, and unmasked hypophyseal hypothyroidism in another two. We concluded that biosynthetic growth hormone extracted from mammalian cells is a safety drug, and that patients with growth hormone deficiency may be treated with it in order to improve their final height expectancy.

[The use of a biosynthetic human growth hormone in children with a growth hormone deficiency]. / Uso de hormona de crecimiento humana biosintética en niños con deficiencia de hormona de crecimiento.

Ten growth hormone deficient patients (GHDP) (six girls & four boys) with chronological age range six-15 years old, received recombinant human growth hormone (GH) (0.6 UI/kg/week) in three subcutaneous injections per week over 14 months. A complete physical examination was performed monthly; thyroid hormones concentrations every two months, as well as, bone age every four months were determined. Growth velocity was higher in the first six months' treatment (12.73 +/- 1.65 cm/yr vs 10.6 +/- 1.67 cm/yr; previous growth velocity 3.76 +/- 2.4 cm/yr). The variables with better prognosis were: lower previous growth velocity, lower chronological age (< 12 ys), lower bone age (< 8 ys), higher BMI than ideal; and higher acid alkaline phosphate during treatment. Two of ours patients developed biochemical hypothyroidism during the treatment period, and they needed hormonal substitutive therapy. With GH treatment a harmonious growth is obtain with bone age progress according chronological age. The height of GHDP treated with GH do not approaches the height range of their parents, probably due the time of treatment. In the follow-up of all patients treated with GH is important to realize periodical laboratories screenings looking for another hypothalamic and/or hypophyseal deficiencies.

Maternal-fetal carnitine relationship and neonatal ketosis in the rat.

The concentration of ketone bodies in plasma and of carnitine in various maternal, fetal, and neonatal tissues was examined during the developmental period in rats. Plasma ketone levels were low in the fetus, increased 10-fold during the first 24 h postpartom, and thereafter gradually declined such that normal values were found at the end of the suckling period. An almost identical profile was observed for liver carnitine concentrations in the baby rats. The converse was true for heart tissue, the carnitine content of which was low at birth and steadily increased to adult levels with the time of suckling. The primary source of carnitine in neonatal tissues, at least during the first 2 to 3 days postpartum, was shown to be the mother rat whose liver and milk carnitine content was very high at this time and fell as nursing continued. Experiments in which the fate of [14C]butyrobetaine, the immediate precursor of carnitine, was followed after injection into nursing mother rats indicated movement of carnitine from maternal liver leads to maternal plasma leads to milk leads to neonatal tissues. The above findings support the view expressed earlier that one prerequisite for the development of a high ketogenic profile in liver may be an elevation in the tissue carnitine concentration. Additional factors, however, are clearly involved as evidenced by the observation that in the fed state perfused livers from nursing mother rats synthesized ketone bodies from oleic acid at low rates compared with those seen after a 24 h fast, despite the fact that tissue carnitine levels were equally elevated in both groups. This paradox is likely related to the fact that in the fed state such livers also contained large quantities of glycogen, depletion of which through fasting was accompanied by marked acceleration of ketogenesis from oleate. The data indicate, therefore, that maximal ketogenic capacity of the liver requires for its induction an increase in carnitine coupled with a decrease in glycogen content of the tissue.

Role of carnitine in hepatic ketogenesis.

The enhancement of long-chain fatty acid oxidation and ketogenesis in the perfused rat liver, whether induced acutely by treatment of fed animals with anti-insulin serum or glucagon, or over the longer term by starvation or the induction of alloxan diabetes, was found to ba accompanied by a proportional elevation in the tissue carnitine content. Moreover, when added to the medium perfusing livers from fed rats, carnitine stimulated ketogenesis from oleic acid. The findings suggest that the increased fatty acid flux through the carnitine acyltransferase (carnitine palmitoyl-transferase; palmitoyl-CoA:L-carnitine O-palmitoyltransferase; EC 2.3.1.21) reaction brought about by glucagon excess, with or without insulin deficiency, is mediated, at least in part, by elevation in the liver carnitine concentration.