GATA3 abnormalities in six patients with HDR syndrome.

GATA3 mutations cause HDR (hypoparathyroidism, sensorineural deafness, and renal dysplasia) syndrome and, consistent with the presence of the second DiGeorge syndrome locus (DGS2) proximal to GATA3, distal 10p deletions often leads to HDR and DiGeorge syndromes. Here, we report on six Japanese patients with GATA3 abnormalities. Cases 1-5 had a normal karyotype, and case 6 had a 46,XX,del(10)(p15) karyotype. Cases 1-6 had two or three of the HDR triad features. Case 6 had no DiGeorge syndrome phenotype except for hypoparathyroidism common to HDR and DiGeorge syndromes. Mutation analysis showed heterozygous GATA3 mutations in cases 1-5, i.e., c.404-405insC (p.P135fsX303) in case 1, c.700T>C & c.708-709insC (p.F234L & p.S237fsX303) on the same allele in case 2, c.737-738insG (p.G246fsX303) in case 3, c.824G>T (p.W275L) in case 4, and IVS5+1G>C (splice error) in case 5. Deletion analysis of chromosome 10p revealed loss of GATA3 and preservation of D10S547 in case 6. The results are consistent with the previous finding that GATA3 mutations are usually identified in patients with two or three of the HDR triad features, and provide supportive data for the mapping of DGS2 in the region proximal to D10S547.

Expression of stanniocalcin-1 in gastrointestinal tracts of neonatal and mature rats.

Stanniocalcin-1 (STC-1) produced by ovaries endocrinologically targets to mammary glands and is secreted into milk during lactation. The decline of mother's serum level by STC-1 antiserum administration reduced the milk fat content and the pups' body fat content. Nevertheless, the pups' fecal fat content was increased, suggesting that milk-derived STC-1 could influence intestinal fat absorption. We investigated the STC-1 expression in rat gastrointestinal tissues using immunocytochemistry and in situ hybridization. STC-1 was widely expressed in the chief cells of gastric pits and the cells of intestinal glands. Goblet cells in the small intestine contained STC-1 protein in their mucus. The distribution shows that this peptide is secreted exocrinologically into the gastrointestinal lumen. Quantitative RT-PCR analysis revealed that the expression ratio was higher in the periods of heavy nutritional demand, such as growing and lactation. The endogenous STC-1, similar to milk-derived STC-1, may be involved in digestion and/or absorption in gastrointestinal organs.

Maternal exposure of low dose of TCDD modulates the expression of estrogen receptor subunits of male gonads in offspring.

We have analyzed the effects of low-dose transplacental and lactational exposure of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on gene expression relating to the dioxin and sexual hormone cascade, and demonstrated the effects on testicular growth and sexual maturation in male offspring rats. TCDD (10 ng/kg) was administered to dams on Days 7 and 14 of gestation, and on Days 0, 7 and 14 after delivery. Gene expression of cytochrome P450 family 1 subfamily A polypeptide 1 (CYP1A1) in the liver of 17-day-old rats was significantly increased compared with controls. Furthermore, expression of estrogen receptors (ER)alpha and ERbeta was significantly increased at 17 and 42 days old, respectively in the testis of TCDD-administered rats compared with controls. Although testicular weight and the seminiferous tubule diameter were increased in 17-day-old rats, there was no difference in the number of germ cells between TCDD-treated and control animals. The expressions of androgen receptor and inhibin subunit genes were not significantly changed. These findings suggest that low-dose exposure of TCDD leads to unusual development of the testis by perturbation of steroid hormone homeostasis.

The role of fibroblast growth factor 23 for hypophosphatemia and abnormal regulation of vitamin D metabolism in patients with McCune-Albright syndrome.

McCune-Albright syndrome (MAS) is sometimes complicated by hypophosphatemia and abnormally low levels of 1,25(OH)(2)D in the presence of hypophosphatemia. Recently, fibroblast growth factor 23 (FGF-23) was reported as a phosphaturic and a causal factor of abnormal vitamin D metabolism. This abnormal phosphate and vitamin D metabolism is well known to be found in oncogenic and X-linked hypophosphatemia. We furthermore reported increased circulating plasma FGF-23 levels in patients with oncogenic and X-linked hypophosphatemia. To determine whether FGF-23 may be involved in the pathogenesis of MAS, we measured plasma FGF-23 levels in six MAS patients. As a control for hypophosphatemia, we also investigated the plasma FGF-23 levels in two patients with hereditary hypophosphatemic rickets with hypercalciuria (HHRH). We also investigated the correlation of plasma FGF-23 levels with serum phosphate and 1,25(OH)(2)D levels after short-term pamidronate therapy in three MAS patients. Plasma FGF-23 levels were significantly increased in patients with MAS compared to normal controls, whereas they were not increased in HHRH patients. Serum phosphate levels of the MAS patients were significantly lower than those observed in normal controls. Plasma FGF-23 levels showed significant negative correlation with serum phosphate concentrations. In three MAS patients, pamidronate therapy decreased plasma FGF-23 levels, which showed significant negative correlation with serum 1,25(OH)(2)D concentrations. These data suggested that FGF-23 is a possible causal factor for hypophosphatemia and abnormal vitamin D metabolism in MAS.

Association of the CTLA-4 gene 49 A/G polymorphism with type 1 diabetes and autoimmune thyroid disease in Japanese children.

OBJECTIVE: To clarify the role of the T-lymphocyte-associated-4 (CTLA-4) polymorphism in the susceptibility to child-onset type 1 diabetes with regard to its clinical characteristics and complications with autoimmune thyroid disease (AITD) in the Japanese population. RESEARCH DESIGN AND METHODS: The CTLA-4 49 A/G polymorphism was detected by the PCR-restriction fragment-length polymorphism (RFLP) method in 97 type 1 diabetic subjects and 20 patients with Graves' disease, a cohort which included 4 patients who also had type 1 diabetes. RESULTS: The genotypes and allele frequencies of this polymorphism did not differ between the type 1 diabetic subjects and the control subjects. The G allele frequency was 63.9% in the type 1 diabetic subjects. The G allele frequency in the subgroup of patients with a high titer of autoantibodies to the GAD antibody (Ab) was 72.9% (P = 0.0499 vs. control subjects); in the subgroup of patients without HLA DRB1*0405, it was 72.6% (P = 0.0271 vs. control subjects); and in the subgroup of patients with a residual beta-cell function, it was 78.6% (P = 0.0391 vs. control subjects). The G allele frequency in the patients with Graves' disease was also significantly higher at 78.1% (P = 0.0405 vs. control subjects). Furthermore, the frequency in our diabetic subjects complicated with Graves' disease was even higher (87.5%). CONCLUSIONS: We have demonstrated that a distinct association exists between the G allele of CTLA-4 and high values of GAD Ab, residual beta-cell function, and the absence of HLA-DRB1*0405.

Functional polymorphisms in the mineralocorticoid receptor and amirolide-sensitive sodium channel genes in a patient with sporadic pseudohypoaldosteronism.

Pseudohypoaldosteronism (PHA) is characterized by urinary salt-wasting in infancy resulting from a congenital resistance to aldosterone involving the genes for the mineralocorticoid receptor (MR) and the amiloride-sensitive sodium channel (ENaC). We identified, in a Japanese patient with sporadic PHA, three homozygous substitutions in the MR gene: G215-->C215, A754-->G754 (Ile180-->Val180), C938-->T938 (Ala241-->Val241), which had previously been reported to occur in healthy populations. Luciferase activities induced by MR with either G215-->C215, Ile180-->Val180, or Ala241-->Val241 substitution were significantly lower than those for wild-type MR with aldosterone at concentrations ranging from 10(-11) to 10(-9) M, 10(-8) M, or 10(-11) to 10(-6) M, respectively. A homozygous A-->G substitution of the donor splice site of alphaENaC intron 4 was found in the patient. The corresponding cDNA exhibited a normal structure, suggesting that this substitution does not alter the splice. The results suggest that each of three MR polymorphisms identified in our patient is functionally and structurally heterogeneous. We hypothesize that two or more "functional" polymorphisms, any of which exhibits only slight effects on MR or ENaC function and is alone incapable causing PHA, may in the right allelic combination induce the negative salt-conservation characteristic of PHA.

Role of the conserved DRY motif on G protein activation of rat angiotensin II receptor type 1A.

To delineate the functional importance of the highly conserved triplet amino acid sequence, Asp-Arg-Tyr (DRY) among G protein-coupled receptors in the second intracellular loop, these residues of rat angiotensin II (Ang II) receptor type 1A (AT(1A)) were changed by alanine or glycine by site-directed mutagenesis. These mutant receptors were stably expressed in CHO-K1 cells, and the binding of Ang II, GTP effect, InsP(3) production, and the acidification of the medium in response to Ang II were determined. The effects of GTPgammaS on Ang II binding in the mutant receptors D125A and D125G were markedly reduced. InsP(3) production of the mutant D125A, D125G, R126A, and R126G was markedly reduced. Extracellular acidification of D125A was not distinguishable from untransfected CHO-K1 cells. Mutant Y127A was able to produce InsP(3) and acidify medium comparable with wild type AT(1A). These results indicate as follows; Asp(125) is essential for intracellular signal transduction involving G protein coupling, Arg(126) is essential for coupling of G(q) protein but not other G proteins, and Tyr(127) is not important for G protein coupling.

Effects of growth hormone on testicular dysfunction induced by cyclophosphamide (CP) in GH-deficient rats.

Growth hormone (GH) is known to accelerate spermatogenesis and maintain gonadal function. In this study, we evaluated the effect of GH on recovery from testicular damage induced by cyclophosphamide (CP). Eleven- to fourteen-week-old GH-deficient Lewis rats (dw/dw) were divided into 4 groups (n = 10 each), with one group serving as controls. In the CP group, CP was intravenously administered in daily doses of 50 mg/kg for 2 days, followed by daily doses of 10 mg/kg for the next 3 days. In the GH group, rat GH was subcutaneously administered at a daily dose of 0.3 mg/kg until the rats were sacrificed. In the CP/GH group, GH and CP administration were started simultaneously. In the CP/preGH group, GH administration was started 14 days before CP administration. Five rats from each group were sacrificed at days 14 and 28 after administration of CP. Spermatogenesis was then evaluated morphometrically by counting numbers of cells at several stages of the spermatogenic cycle. On day 14, there were no significant differences in the numbers of the spermatocytes between CP and CP/GH group. On day 28, the numbers of spermatocytes and motility of spermatozoa in CP/GH group were greater than those of CP group were. In the CP/preGH group, these effects of GH administration were not observed. These results suggested that administration of GH improved testicular function damaged by CP under GH-deficient condition, when GH and CP administration are started simultaneously.