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Biomedical subjects

C J Migeon

Publications and source records attributed to C J Migeon.

At least 55 records · Page 3Linked to original sources
PubMed1989

Two distinct areas of unequal crossingover within the steroid 21-hydroxylase genes produce absence of CYP21B.

We mapped crossover sites in chimeric, recombinant CYP21 genes from six patients with salt-losing congenital adrenal hyperplasia (CAH). Nucleotide sequences unique to the CYP21A pseudogene or to the active CYP21B gene were mapped using gene-specific restriction sites and oligonucleotide hybridizations. Each chimeric CYP21 gene in the CYP21-deletion linked haplotypes contained sequences near the 5' end that were characteristic of CYP21A and only a single transition from sequences of CYP21A to those of CYP21B at the 3' end. The transitions all occurred within either of two discrete regions (+470 to +999 and +1375 to +1993). All eight chimeric CYP21 genes coupled with HLA-Bw47 in five unrelated patients had the CYP21A-CYP21B sequence transition within the same gene region (+1375 to +1993). One of the three other "CYP21B deletion" haplotypes (HLA-B7) had a sequence transition within this same region, while in the other two haplotypes (HLA-B61 and HLA-B18) the transition occurred between base pairs +470 and +999. By contrast, both CYP21 genes in a haplotype containing a gene conversion of CYP21B to CYP21A contained apparent transitions between sequences of CYP21A and CYP21B. We conclude that a single, unequal crossingover between the CYP21A and the CYP21B genes yields deletion of the active CYP21 gene and salt-losing CAH and that these crossingovers do not occur randomly within the CYP21 genes of our patients.

Adrenal Hyperplasia, Congenital↗
PubMed1989

Sequence of the intron/exon junctions of the coding region of the human androgen receptor gene and identification of a point mutation in a family with complete androgen insensitivity.

Androgens act through a receptor protein (AR) to mediate sex differentiation and development of the male phenotype. We have isolated the eight exons in the amino acid coding region of the AR gene from a human X chromosome library. Nucleotide sequences of the AR gene intron/exon boundaries were determined for use in designing synthetic oligonucleotide primers to bracket coding exons for amplification by the polymerase chain reaction. Genomic DNA was amplified from 46,XY phenotypic female siblings with complete androgen insensitivity syndrome. AR binding affinity for dihydrotestosterone in the affected siblings was lower than in normal males, but the binding capacity was normal. Sequence analysis of amplified exons demonstrated within the AR steroid-binding domain (exon G) a single guanine to adenine mutation, resulting in replacement of valine with methionine at amino acid residue 866. As expected, the carrier mother had both normal and mutant AR genes. Thus, a single point mutation in the steroid-binding domain of the AR gene correlated with the expression of an AR protein ineffective in stimulating male sexual development.

Alleles↗
PubMed1989

Serum 3 alpha-androstanediol glucuronide measurements in sexually mature women with congenital adrenal hyperplasia during therapy.

Serum 3 alpha-androstanediol glucuronide (3 alpha-diol G) measurements may indicate the extent of androgen metabolism and action in target tissues. To test this supposition we measured serum 3 alpha-diol G concentrations in 23 women with congenital adrenal hyperplasia due to 21-hydroxylase deficiency, including 13 with the salt-losing and 10 with the simple virilizing form, while they were receiving glucocorticoid and, in some cases, mineralocorticoid therapy. Their mean age was 28.3 yr (range, 17.9-38.7 yr). Twenty-four-hour urinary 17-ketosteroid excretion, plasma androstenedione and testosterone levels, and serum 3 alpha-diol G levels were measured during the follicular phase. The values were within or below the normal range in 87%, 78%, 70%, and 91% of the patients, respectively. By contrast, plasma 17-hydroxyprogesterone levels were normal in only 12% of the patients. Serum 3 alpha-diol G levels correlated well with 24-h urinary 17-ketosteroid excretion (r = 0.75) and plasma 17-hydroxyprogesterone (r = 0.77), androstenedione (r = 0.84), and testosterone (r = 0.93) levels. The serum 3 alpha-diol G levels were not significantly different in the women with the salt-losing form and those with the simple virilizing form. However, they were significantly lower (P less than 0.05) in the women with normal menses compared to those with abnormal menses. This finding underscores the validity of serum 3 alpha-diol G measurements as indicators of androgen production and metabolism in women. The excellent correlation between the serum 3 alpha-diol G levels and standard measures of control indicates that the former measurement may be a useful adjunct in the management of women with congenital adrenal hyperplasia.

Adrenal Hyperplasia, Congenital↗
PubMed1988

Studies of up-regulation of androgen receptors in genital skin fibroblasts.

Prolonged exposure of genital skin fibroblasts (GSF) to dihydrotestosterone (DHT) increases androgen receptor binding of steroid, a process termed 'up-regulation'. Because the extent of up-regulation appears to be quite variable, we have investigated the optimal conditions and the molecular mechanisms that control this phenomenon in seven strains of normal neonatal GSF. When GSF were incubated for 1-48 h with 3 nM methyltrienolone (R1881), maximal up-regulation was reached by 20 h and remained constant thereafter. With DHT, rapid steroid metabolism required replenishment of DHT for maximum up-regulation. Up-regulation levels following 20 h incubation with DHT (including steroid replenishment) and R1881 were 2.07-fold (range = 1.1-3.3) and 2.35-fold (range = 1.86-3.33), respectively. The greater variability observed with DHT may be related to variable rates of steroid catabolism among cell strains. Half-maximal up-regulation was attained at 0.29 nM R1881, which approximates the Kd. Maximal up-regulation was reached only with continuous exposure to R1881 for 24 h. It was completely inhibited by actinomycin D (0.5 micrograms/ml) or cycloheximide (10 micrograms/ml). Following up-regulation, removal of R1881 for 24 h resulted in a highly variable decline of androgen receptors among cell strains. Maximal up-regulation could be reinduced by exposure to R1881 again for an additional 24 h. During up-regulation, androgen receptor levels in nuclei and nuclear matrix rose with increments comparable to those obtained in whole cells. We conclude that the extent of up-regulation and its rate of decline differ greatly among normal cell strains. Hence, its study in cells of patients with androgen insensitivity may have limited value.(ABSTRACT TRUNCATED AT 250 WORDS)

Cells, Cultured↗
PubMed1988

Restriction fragment analysis of duplication of the fourth component of complement (C4A).

The two genes encoding the fourth component of complement (C4A and C4B) reside between HLA-B and HLA-DR on human chromosome 6. Two kilobases downstream from each C4 gene lies a 21-hydroxylase gene (CA21HA and CA21HB, respectively). Utilizing the method of Southern blotting and a 5'-end 2.4-kb BamHI/KpnI fragment of the C4 cDNA, we have analyzed TaqI-digested DNA from four pedigrees with one or more extended haplotypes containing a C4A duplication, as demonstrated by protein electrophoresis and segregation analysis. Two C4A protein duplications (C4A*2,A*3,C4B*QO and C4A*3,A*5,C4B*QO) segregated with two large TaqI DNA restriction fragments (7.0 and 6.0). In pedigree Fi, one individual homozygous for HLA-A3,B35,C4,DR1,DQ1,BFF,C2C,-C4A2,3,C4BQO had TaqI 7.0- and 6.0-kb restriction fragments with equal hybridization intensities as measured by two-dimensional densitometry (7.0/6.0 kb = 0.83, SD = 0.12, N = 7). A hybridization probe for the 21-hydroxylase gene also demonstrated equal gene dosage (CA21HA/CA21HB = 1.01). DNA from another individual (Ma I-2) with a different C4A gene duplication (C4A*3,A*5,C4B*QO) also had equal densitometry measurements (7.0/6.0 kb = 1.07). We conclude that two extended haplotypes from unrelated pedigrees have two C4 genes and both C4 genes encode separate C4A alleles. These findings are compatible with a gene conversion event of C4B to C4A.

Adrenal Hyperplasia, Congenital↗
PubMed1988

Deletion of the steroid-binding domain of the human androgen receptor gene in one family with complete androgen insensitivity syndrome: evidence for further genetic heterogeneity in this syndrome.

The cloning of a cDNA for the human androgen receptor gene has resulted in the availability of cDNA probes that span various parts of the gene, including the entire steroid-binding domain and part of the DNA-binding domain, as well as part of the 5' region of the gene. The radiolabeled probes were used to screen for androgen receptor mutations on Southern blots prepared by restriction endonuclease digestion of genomic DNA from human subjects with complete androgen insensitivity syndrome (AIS). In this investigation, we considered only patients presenting complete AIS and with the androgen receptor (-) form as the most probable subjects to show a gene deletion. One subject from each of six unrelated families with the receptor (-) form of complete AIS and 10 normal subjects (6 females and 4 males) were studied. In the 10 normal subjects and in 5 of the 6 patients, identical DNA restriction fragment patterns were observed with EcoRI and BamHI. In one affected individual, a partial deletion of the androgen receptor gene involving the steroid-binding domain was detected. Analysis of other members of this family confirmed the apparent gene deletion. Our data provide direct proof that complete AIS in some families can result from a deletion of the androgen receptor structural gene. However, other families do not demonstrate such a deletion, suggesting that point mutations (or small, undetectable deletions) may also result in the receptor (-) form of complete AIS, adding further to the genetic heterogeneity of this syndrome.

Blotting, Southern↗
PubMed1988

Down-regulation of the glucocorticoid receptor by dexamethasone in cultured human skin fibroblasts: implications for the regulation of aromatase activity.

Human genital skin fibroblasts grown in cell culture contain aromatase activity and glucocorticoid receptors, providing a model for studying the relationship between changes in levels of dexamethasone (DEX) receptor binding and changes in the response of aromatase to DEX. Incubation of cells with media containing DEX produced a time-dependent decline in receptor number, with a nadir at 18-24 h. When DEX was removed from the media, only 40-60% of the lost receptor content was recovered. Binding of DEX to its receptor declined in a dose-dependent fashion after 24-h exposure to this glucocorticoid; the concentration of DEX that produced a half-maximal decline (2.2 nmol/L) was in the same range as the Kd for the receptor-DEX complex (14 nmol/L). Incubation of cells with DEX for 20 h also reduced the level of DEX receptor binding in purified nuclei and nuclear matrix by 76% and 89%, respectively. When subcellular fractions were prepared after incubation of cells with DEX for 1-24 h, whole cell, cytosolic and nuclear DEX receptor binding declined in parallel with time. Incubation of skin fibroblasts with DEX resulted in progressive stimulation of aromatase activity, with a peak at 24 h followed by a return to baseline at 72 h. The initial stimulation of aromatase was mediated by DEX receptors. However, the decline in aromatase activity after prolonged DEX exposure did not appear to be due to a decline in the level of DEX receptor binding. The data supporting this last conclusion included the following. When cells were washed free of DEX after 48 h, DEX receptor binding recovered within 24 h, whereas aromatase activity could not be maximally restimulated until 36 h; when cells were incubated with media containing DEX and cycloheximide for 1-48 h, DEX receptor binding declined to a nadir by 24 h, whereas aromatase activity rose continuously up to 48 h. These findings are consistent with the concept that the aromatase gene in skin fibroblasts is subject to both positive and negative regulation.

Aromatase↗
PubMed1987

Prevalence of polymorphic 21-hydroxylase gene (CA21HB) mutations in salt-losing congenital adrenal hyperplasia.

Using genomic restriction analysis of 14 unrelated patients with salt-losing congenital adrenal hyperplasia, we identified three different CA21HB mutation patterns: no detectable restriction fragment abnormalities (16/28 haplotypes), deletion of the active CA21HB gene (9/28), and apparent conversion of the active CA21HB gene to the pseudogene CA21HA (3/28). CA21HB gene deletion was associated with HLA-Bw47 in 6 haplotypes and with absent C4B expression in 7. A variety of HLA and C4 types was associated with the other mutations. Apparent conversion of CA21HB to CA21HA was identified by the disparity between the intensity ratios for the major TaqI and BglII hybridization fragments.

Adrenal Hyperplasia, Congenital↗
PubMed1987

Fertility rates in female patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Among 80 women with congenital adrenal hyperplasia due to 21-hydroxylase deficiency (40 with the simple virilizing form and 40 with the salt-losing form), 40 reported having an adequate introitus and being heterosexually active. In 15 of 25 patients with the simple virilizing form, 25 pregnancies resulted in 20 normal children, whereas only 1 of 15 women with the salt-losing form became pregnant; this pregnancy was electively terminated. Several factors seem to be responsible for the low fertility rates: noncompliance with therapy was probably high, as suggested by hirsutism and poor endocrine follow-up in 25 percent of patients; whereas 49 patients had regular menstrual periods, 14 had irregular periods, 10 had amenorrhea, 5 had undergone hysterectomy, and 2 had entered menopause; 87 percent of patients with salt loss and 50 percent of those with simple virilization (P less than 0.001) had remained single; the vaginal introitus was reported to be inadequate for intercourse by 35 percent of patients (53 percent of those with salt loss and 18 percent of those with simple virilization; P less than 0.002); and heterosexual activity was reported less frequently among patients with an inadequate introitus. The status of the introitus seemed to have a more important role in the sexual activity reported than did the degree of prenatal exposure to androgen (which was higher among patients with salt loss than among those with simple virilization). However, our data did not rule out an effect of androgen exposure on female fetuses. Our experience indicates that improved surgical correction of the external genitalia and better compliance with therapy will be necessary to improve fertility rates among women with congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Adolescent↗
PubMed1987

Gonadal function in patients with Down syndrome.

Gonadal function was evaluated in 100 home-reared persons with Down syndrome (DS) including 53 boys and men and 47 girls and women. In order to definitively evaluate gonadal function in our subjects, all patients with abnormal thyroid function were excluded from endocrine analysis. Among the men, the frequency of hypospadias and cryptorchidism was similar to that of the general population. In both men and women, the ages for the onset and completion of puberty were also normal. However, among adult men with DS, the mean stretched penile length and the mean testicular volume were significantly below the mean value of normal men. In the 23 men with DS, the mean serum levels of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were significantly elevated above the mean for normal men. By contrast, the mean plasma level of testosterone (T) was normal, suggesting a diagnosis of partial gonadal deficiency. Among the 14 women in the study, 6 had primary gonadal dysfunction. In prepubertal children, serum FSH levels in 3 boys and 5 girls were more than 2 SD above the mean for normal children, while serum levels of LH in 3 boys and 2 girls were abnormally elevated. When gonadal function was evaluated in male infants, serum levels of FSH were above the normal in 6 of 11 subjects. Serum LH was abnormally elevated in 3 of 8 female infants. Our data argue that primary gonadal deficiency is common in DS, that it is progressive from birth to adolescence, and that it is clearly manifest in adult patients.

Adolescent↗
PubMed1987

The influence of 4-hydroxy-4-androstene-3,17-dione on androgen metabolism and action in cultured human foreskin fibroblasts.

4-hydroxy-4-androstene-3,17-dione (4-OHA) has been shown to be a potent inhibitor of aromatase activity. It is effective in the control of estrogen-dependent processes in female subjects and may potentially be useful in the treatment of estrogen-dependent processes in men. Human foreskin fibroblasts grown in cell culture provide a model to investigate the effects of 4-OHA on extraglandular aromatase activity as well as the ability of the compound to influence androgen receptor binding and the 5 alpha-reduction of testosterone (T). Initial experiments were carried out to determine the potency of 4-OHA in genital skin fibroblasts by incubating cells with 4-OHA over a range of concentrations. When aromatase activity was determined at a substrate concentration close to the apparent Km of the enzyme, a 44% inhibition of enzyme activity occurred at a mean concentration of 5 nM 4-OHA. Enzyme kinetic studies analyzed by Eadie-Hofstee plots demonstrated competitive inhibition by 4-OHA with a mean apparent Ki of 2.7 nM. When 5 alpha-reductase activity was determined in the presence of 200 nM [3H]T, in the absence or presence of 4-OHA, a 50% inhibition of enzyme activity occurred at an inhibitor concentration of 3 microM. In androgen receptor binding studies, 4-OHA possessed 1% of the affinity of dihydrotestosterone (DHT) for [3H]DHT binding sites. In summary: 4-OHA is a potent and specific inhibitor of aromatase activity in human genital skin fibroblasts, the affinity of the enzyme for 4-OHA being greater than its affinity for the substrate, androstenedione. The influence of 4-OHA on 5 alpha-reductase activity and androgen receptor binding is minimal.

Androgens↗
PubMed1987

Effects of 10-(2-propynyl)-estr-4-ene-3,17-dione (MDL 18,962)--a mechanism-based irreversible inhibitor of aromatase--in cultured human foreskin fibroblasts.

In male subjects, peripheral aromatization of androgens accounts for most of the estrogen production, and skin is an important site of such enzymatic activity. We have studied the effects of a mechanism-based, irreversible aromatase inhibitor, 10-(2-propynyl)-estr-4-ene-3,17-dione (MDL 18,962) on androgen action and metabolism in cultured human foreskin fibroblasts. Cells were incubated simultaneously in the presence of substrate, androstenedione, and inhibitor, MDL 18,962. Aromatase activity was linear with time up to 3 h of incubation at 37 degrees C in the absence and presence of 1.0-10 nM inhibitor. The IC50 for four different cell strains ranged from 4.0 to 8.6 nM MDL 18,962. Kinetic analysis of competitive inhibition by the Eadie-Hofstee method yielded an apparent Ki of 2.75 nM for the inhibitor. Preincubation of cells with MDL 18,962 resulted in irreversible inhibition of aromatase activity which was time- and concentration-dependent. We calculated a Ki of 7.6 nM for MDL 18,962. Preincubation of cells with 25 nM MDL 18,962 suppressed enzyme activity for up to 6 h following removal of the inhibitor, before a return of enzyme activity due to synthesis of new enzyme. MDL 18,962 (0.2-20 microM) did not influence the 5 alpha-reduction of testosterone (200 nM). In addition, binding of dihydrotestosterone (2 nM) to androgen receptors was not affected by MDL 18,962 (25-1000 nM). In summary, MDL 18,962 is a specific, high potency inhibitor of aromatase. By virtue of its high binding affinity to the enzyme active site, it competes very effectively with substrate, resulting in irreversible inactivation of aromatase.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed1987

HCG stimulation in children with cryptorchidism.

Fifty-eight children with cryptorchidism have been given hCG stimulation testing, 31 with bilateral cryptorchidism, 22 with unilateral, and 5 with prior unsuccessful orchiopexy. Hormonal studies were carried out prior to and following stimulation. In bilateral cryptorchidism, bilateral descent was observed in 32 percent of cases and in unilateral, the success rate was 55 percent. From their data, the authors concluded that hCG is not indicated if patients present elevated LH and FSH levels or if the basal T levels are in the pubertal range. In the other subjects, the hCG test will permit the determination of the presence or absence of testosterone production and in some cases it results in testicular descent. Finally, in cases of failure that require surgery, the hCG will stimulate tissue growth enhancing the success of orchiopexy.

Adolescent↗
PubMed1987

Coupling of HLA-A3,Cw6,Bw47,DR7 and a normal CA21HB steroid 21-hydroxylase gene in the Old Order Amish.

HLA-Bw47, a rare human histocompatibility antigen, occurs in strong linkage disequilibrium with HLA-A3,Cw6,DR7 and salt-losing congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency and is associated with a contiguous deletion of the active CA21HB gene and the C4B complement gene. We studied the HLA-A3,Cw6,Bw47,DR7 haplotype in 10 subjects of the Old Order Amish of Lancaster County in Pennsylvania and found that this haplotype, which occurs with a similar frequency in this group as in the general caucasoid population, has C4B and Ca21HB genes. These C4B and CA21HB genes are expressed as assessed by C4 typing and iv ACTH testing, respectively. Serological studies indicate that the HLA-D loci of this Amish haplotype are the same as those in patients with HLA-Bw47 and CAH, but different from HLA-D loci coupled to HLA-B13, which some workers have proposed is the progenitor genotype for HLA-Bw47. Our studies demonstrate that 1) HLA-Bw47 is not an invariant marker for salt-losing CAH due to 21-hydroxylase deficiency, and 2) the HLA-Bw47 phenotype coupled to CAH is not derived from the HLA-B13 genotype by a single mutation.

Adrenal Hyperplasia, Congenital↗
PubMed1987

Familial hypersecretion of adrenal androgens transmitted as a dominant, non-HLA linked trait.

Clinical evidence of adrenal androgen hyperfunction (premature pubarche, hirsutism, amenorrhea) occurred in the studied proband, her mother, maternal aunt (twin sisters), and maternal great-grandmother. The basal levels of androgen in the first three were variably elevated. In all the members of this family who were tested, the response of 17-hydroxyprogesterone and progesterone to adrenocorticotropic hormone stimulation was either normal or of the type seen in heterozygotes for congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Of particular importance is the fact that neither the proband nor her mother or maternal aunt had the type of response seen in homozygotes presenting the attenuated form of congenital adrenal hyperplasia. The disorder appears to be a familial condition resulting in excessive levels of adrenal androgens beginning during childhood years, causing hirsutism and amenorrhea and interfering with normal pubertal and adult ovarian function. Glucocorticoid therapy suppresses adrenal androgen levels; in two individuals, conception occurred twice in each during such treatment in otherwise amenorrheic individuals. The pattern of transmission of the disorder appears to be either autosomal or X-linked dominant, and not linked to the homologous leucocytic antibodies (HLA) region of the sixth chromosome.

Adolescent↗
PubMed1986

McCune-Albright syndrome. Long-term follow-up.

This article describes clinical follow-up of 15 patients--13 females and two males--with McCune-Albright syndrome. Osseous fractures occurred only during childhood, while hearing impairment due to temporal bone involvement occurred in four of six adults. Four females with precocious puberty had final heights that were not different from the mean for normal females; they eventually developed regular menses, and two had children. Persistent hyperthyroidism requiring ablative therapy occurred in three subjects, while hypophosphatemia occurred in three subjects. The protean manifestations of this disorder suggest that it results from a basic defect of cellular regulation. We postulate that its varied endocrine abnormalities result from altered regulation of intracellular cyclic adenosine monophosphate effects.

Acromegaly↗
PubMed1986

Restriction maps and restriction fragment length polymorphisms of the human 21-hydroxylase genes.

Restriction maps were constructed for the two human 21-hydroxylase genes (21-OHA and 21-OHB) by using DNA from subjects homozygous for a deletion of each gene. Comparing the patterns of these two genes, a KpnI restriction site occurred in the 21-OHA gene in place of a TaqI site in the 21-OHB gene about 1-kb from the 5' end of the gene, and an extra EcoRI site was located 500 bp 5' to the common EcoRI site. The DNA of fourteen unrelated normal subjects was digested with nine restriction endonucleases (AccI, BamHI, BgIII, EcoRI, HindIII, KpnI, MspI, SacI and TaqI). Restriction fragment length polymorphisms were found with EcoRI, HindIII and AccI that resulted from polymorphic endonuclease sites outside the genes.

Adrenal Hyperplasia, Congenital↗