Mutations of carnitine palmitoyltransferase II (CPT II) in Japanese patients with CPT II deficiency.

Carnitine palmitoyltransferase II (CPT II) deficiency is an inherited disorder involving beta-oxidation of long-chain fatty acids. CPT II deficiency is a wide-spectrum disorder that includes a lethal neonatal form, an infantile form, and an adult-onset form. However, the ethnic characteristics and the relationship between genotype and clinical manifestation are not well understood. We investigated three non-consanguineous Japanese patients with CPT II deficiency and examined cell lines from 4 unrelated patients and 50 healthy donors. The CPT 2 gene was typed by direct DNA sequencing of polymerase chain reaction-amplified gene products. Case 1 (infantile form) was heterozygous for a phenylalanine to tyrosine substitution at position 383 (p.F383Y) and a novel valine to leucine substitution at 605 (p.V605L). Cases 2, 4, and 5 (infantile form) and case 3 (adult-onset form) were heterozygous for a single mutation at F383Y. Case 6 (adult-onset form) was compound heterozygous at the CPT 2 locus, with deletion of cytosine and thymine at residue 408, resulting in a stop signal at 420 (p.Y408fsX420), and an arginine to cysteine substitution at position 631 (p.R631C). Case 7 (adult-onset form) was homozygous for the p.F383Y mutation. In conclusion, we identified p.F383Y mutations in six of seven patients with CPT II deficiency and two novel variants of the coding gene: p.Y408fsX420 and p.V605L. These mutations differ from those in Caucasian patients, who commonly harbor p.S113L, p.P50H, and p.Q413fsX449 mutations; therefore, our data and those of other Japanese groups suggest that the p.F383Y mutation is significant in Japanese patients with CPT II deficiency.

Ghrelin prevents development of diabetes at adult age in streptozotocin-treated newborn rats.

AIMS/HYPOTHESIS: Ghrelin, a stomach-derived hormone, functions in multiple biological processes, including glucose metabolism and cellular differentiation and proliferation. In this study, we examined whether early treatment with ghrelin can regenerate beta cells of the pancreas in an animal model of diabetes mellitus, the n0-STZ model, in which neonatal rats are injected with streptozotocin (STZ) at birth. METHODS: Following administration of ghrelin to n0-STZ rats from postnatal days 2 to 8, we examined beta cell mass, mRNA expression levels of insulin and of pancreatic and duodenal homeobox 1 (Pdx1) gene, and pancreatic morphology on days 21 and 70. In addition, we investigated the effects of ghrelin on beta cell replication. RESULTS: By day 21, ghrelin treatment increased pancreatic expression of insulin and Pdx1 mRNA in n0-STZ rats. The number of replicating cells was also significantly increased in the ghrelin-treated n0-STZ model. At day 70, n0-STZ rats exhibited hyperglycaemia, despite slight increases in plasma insulin levels. Ghrelin treatment resulted in the improvement of plasma glucose levels, which were associated with normal plasma insulin levels. Pancreatic insulin mRNA and protein levels were significantly increased in ghrelin-treated n0-STZ model animals. CONCLUSIONS/INTERPRETATION: These findings suggest that ghrelin promotes regeneration of beta cells in STZ-treated newborn rats. Thus, early administration of ghrelin may help prevent the development of diabetes in disease-prone subjects after beta cell destruction.

Association of Graves' disease with Evans' syndrome in a patient with IgA nephropathy.

A 20-year-old woman with IgA nephropathy was admitted to Jikei University Hospital for the treatment of rapid deterioration of renal function after receiving 131I-therapy against hyperthyroidism on October 23,1999, and hemodialysis was started. On admission, she was diagnosed as having Evans' syndrome in addition to known Graves' disease. Renal biopsy revealed end-stage renal damage, then, hemodialysis was maintained. Treatment for Evans' syndrome was also started and her general condition gradually improved. The present case implied that "Graves' disease" and "Evans' syndrome" could represent some of the manifestations of an underlying immunological disorder in the patient.

Coordinate regulation of endothelin and adrenomedullin secretion by oxidative stress in endothelial cells.

To elucidate the significance of oxidative stress in the modulation of endothelial functions, we examined the effects of H(2)O(2) on the expression of two endothelium-derived vasoactive peptides, endothelin (ET) and adrenomedullin (Am), and their interaction. H(2)O(2) dose dependently suppressed ET secretion and ET-1 mRNA expression in bovine carotid endothelial cells (ECs). Menadion sodium bisulfate, a redox cycling drug, also decreased ET secretion in a dose-dependent manner. Catalase, a H(2)O(2) reductase, and dl-alpha-tocopherol (vitamin E) significantly inhibited H(2)O(2)-induced suppression of ET secretion. Downregulation of ET-1 mRNA under oxidative stress was regulated at the transcriptional level. In contrast, H(2)O(2) increased Am secretion (and its mRNA expression) accompanied by the augmentation of cAMP production. Am, as well as 8-bromo-cAMP and forskolin decreased ET secretion in a dose-dependent fashion. Furthermore, an anti-Am monoclonal antibody that we developed abolished H(2)O(2)-induced suppression of ET secretion at 6-24 h after the addition of H(2)O(2). H(2)O(2) increased the intracellular Ca(2+) concentration ([Ca(2+)](i)). Moreover, treatment with ionomycin, a Ca(2+) ionophore, and thapsigargin, an inhibitor of endoplasmic reticulum ATPase, decreased ET secretion dose dependently for 3 h. These results suggest that the production of ET was decreased via activation of the Am-cAMP pathway and by the elevation of [Ca(2+)](i) under oxidative stress. These findings elucidate the coordinate expression of two local vascular hormones, ET and Am, under oxidative stress, which may protect against vascular diseases.

Lingual thyroid presenting as acquired hypothyroidism in the adulthood.

There are numerous reported cases of lingual thyroid with an obvious prevalence in pediatric age. Such ectopic thyroid glands are probably quantitatively deficient and thyroid function may be low or at a low normal level. Apparently, most cases of ectopic thyroid tissue develop congenital hypothyroidism, the so-called cretinism. In this report, we describe a very rare adult male case of lingual thyroid who developed hypothyroidism in adulthood; the anomaly remained undiscovered, being without local common symptoms, and permitted a normal life.

Polymer blend implant for ocular delivery of fluorometholone.

Ocular implants containing fluorometholone (FLM) were prepared using blends of poly (DL-lactic acid) (PLA) and polyvinyl pyrrolidone (PVP). The effect of the fraction of PVP content on the release of FLM from the implant was investigated in vitro. The drug was released from the device by approximately following first order kinetics within the period of 40 d. The release rate gradually increased with an increase in the PVP content. The in vivo study after implantation in the anterior chamber of rabbit eyes indicated that the PLA-PVP implant showed a good correlation between the in vitro and in vivo release of FLM. The present polymer blend implant demonstrated a constant level of FLM in the aqueous humor for one month.

Unexplained increase in serum corticosteroid-binding globulin levels in a patient with chronic thyroiditis, pituitary adenoma and empty sella.

Unexplained high serum corticosteroid-binding globulin (CBG) concentrations (mean +/- SD, 74.1 +/- 12.1 micrograms/dl; normal women, 32.5 +/- 5.6 micrograms/dl) were found in an unmarried women who was not pregnant or taking exogenous estrogens. She was also found to suffer from subclinical chronic thyroiditis, pituitary adenoma and empty sella. The increased serum CBG concentrations in this patient were not due to any of the factors known to increase CBG. Consistently high basal serum GH levels and unusual GH responses to GH-releasing factor (GRF) and L-dopa were also noted.

Stimulation by corticotropin-releasing factor of atrial natriuretic peptide and brain natriuretic peptide secretions from cultured neonatal rat cardiomyocytes.

The new functional role of corticotropin-releasing factor (CRF) in the regulation of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) release was investigated using cultured neonatal rat cardiomyocytes. Treatment with CRF (10(-10)-10(-6) M) resulted in dose- and time-dependent increase in ANP and BNP secretion, up to 2.5-fold and 1.8-fold above control values, respectively. The effect was significant at 6 hr and persisted for at least 36 hr. The effect of CRF (10(-7) M) was partially blocked by alpha-helical CRF(9-41) (10(-7) M), a specific CRF receptor antagonist. The effect of CRF (10(-7) M) was not only blunted by cAMP-dependent protein kinase A (PKA) inhibitor, H-89 (10(-5) M), but also by protein kinase C inhibitors, H-7 (50 microM) and Calphostin C (10(-6) M). H-7 (50 microM) and Calphostin C (10(-6) M) alone lowered basal ANP and BNP levels. Furthermore, CRF (10(-7) M) stimulates protein synthesis up to 1.2-fold. These results indicate that CRF stimulates ANP and BNP secretions through the CRF receptor and, at least in part, via PKA activation during cardiac hypertrophy.

Williams syndrome associated with chronic renal failure and various endocrinological abnormalities.

A 31-year-old man who had been under regular hemodialysis for 6 months was diagnosed as Williams syndrome (WS) by fluorescence in situ hybridization (FISH) chromosomal analysis. The association of WS and chronic renal failure (CRF) is only rarely encountered. Endocrinological examinations revealed hypergonadotropic hypogonadism. Prolonged and exaggerated responses of adrenocorticotropin (ACTH) to insulin-induced hypoglycemia and corticotropin releasing hormone (CRH) were also noted. While most of the endocrinological abnormalities observed in this patient could be attributed to altered endocrine circumstances in CRF, some findings stand in contrast. Furthermore, the testicular biopsy specimen showed severe hypospermatogenesis. Endocrine disorders observed in this patient may be at least in part, responsible for various clinical features underlying WS.

47 XXY/46 XY mosaic Klinefelter's syndrome presenting with multiple endocrine abnormalities.

We report here a rare case of 47 XXY/46 XY mosaic Klinefelter's syndrome associated with multiple endocrine disorders. A 35-year-old male admitted for the evaluation of renal dysfunction and recurrent bone fractures was diagnosed as having Klinefelter's syndrome by endocrinological examinations and sex chromosome analysis. He has suffered from diabetes mellitus for more than ten years. The serum FSH and LH levels were high together with low free testosterone and estradiol levels. There was a discrepancy between basal serum GH and somatomedin-C levels. On admission, thyroid function revealed thyrotoxicosis with low radioactive iodine uptake and negative thyroid autoantibodies. During hospitalization, serum FT3 and FT4 levels were gradually decreased and serum TSH levels became elevated, leading to the diagnosis of subacute thyroiditis. Serum ACTH levels showed high basal levels with delayed, exaggerated responses to insulin-induced hypoglycemia. Rapid ACTH test (1-24ACTH 0.25 mg) showed low cortisol responses and many of the adrenocortical steroids in plasma and urine were low or low normal. Furthermore, bone mineral density (BMD) by DEXA showed marked osteoporosis. Possible mechanisms underlying these varied endocrine disorders remain to be elucidated.

A rare case of idiopathic hypoparathyroidism with varied neurological manifestations.

A 47-year-old man was admitted for evaluation of unsteady gait, postural instability, and dysarthria. On admission, neurological examinations revealed cerebellar ataxia, extrapyramidal signs including parkinsonism and positive Trousseau's sign. Laboratory findings revealed severe hypocalcemia and hyperphosphatemia, and serum intact parathyroid hormone was not detectable. Brain computed tomography revealed severe calcification of basal ganglia and dentate nuclei. He was diagnosed as idiopathic hypoparathyroidism; treatment with 1 alpha (OH) vitamin D3 brought marked improvement of neurological manifestations. We report a rare case of idiopathic hypoparathyroidism presenting with extrapyramidal and cerebellar dysfunction with a review of literature.

Possible thyroidal involvement in a case of Fabry disease.

Endocrinological evaluations of a 48-year-old man with Fabry disease revealed low levels of serum thyroid hormones and high levels of serum thyrotropin (TSH), indicating that the patient had primary hypothyroidism. Also, an exaggerated growth hormone (GH) response to hypoglycemic stimuli was observed. Thin layer chromatography of the lipid extract of the thyroid gland obtained by biopsy demonstrated marked accumulation of ceramide trihexoside (CTH) and ceramide dihexoside (CDH). These findings strongly suggest that thyroid hypofunction and hypothalamic dysfunction could be involved in Fabry disease.

Corticotropin-releasing factor (CRF) stimulates 45Ca2+ uptake in the mouse corticotroph cell line AtT-20.

Corticotropin-releasing factor (CRF) stimulates adrenocorticotropin (ACTH) release via the adenylate cyclase/cAMP-dependent protein kinase system. Because calcium is necessary for receptor-mediated release of ACTH, we have examined the effect of CRF on 45Ca2+ uptake in a corticotroph cell line model, AtT-20. Treatment of AtT-20 cells with CRF (10(-9)-10(-6) M) resulted in dose- and time-dependent increases in 45Ca2+ uptake, up to 2.2-fold above control values. The effect was statistically significant at 1 min and persisted for at least 10 min. Treatment with forskolin (1-30 microM), 8-Br-cAMP (0.5 mM), cholera toxin (CT, 100 ng/ml) and K+ (20 mM) also increased cell-associated 45Ca2+. The effect of K+ was completely blocked by nifedipine (100 microM), whereas the effects of CRF (10(-8) M) were only partially inhibited by this calcium channel antagonist. These data suggested a role of voltage-dependent calcium channels in 45Ca2+ uptake. Short term pretreatment (1-2 h) of AtT-20 cells with CRF (10(-8) M) significantly desensitized both CRF-stimulated cAMP accumulation and ACTH release, but did not attenuate CRF-stimulated 45Ca2+ uptake. Pretreatment with CRF (10(-8) M) for 4 h did not alter CT- or forskolin-stimulated cAMP accumulation and ACTH release. This suggests that the molecular mechanisms of desensitization are proximal to adenylate cyclase. Conversely, long term pretreatment (24 h) of AtT-20 cells with CRF (10(-8) M) induced significant desensitization of CRF-stimulated 45Ca2+ uptake. These results indicate that CRF stimulates calcium uptake in AtT-20 cells via cAMP-dependent and cAMP-independent mechanisms, and that the cellular mechanisms involved in desensitization of cAMP accumulation and ACTH release and those involved in desensitization of calcium uptake are qualitatively different.

IgA myeloma complicated by fractures of the bones and hyperviscosity syndrome--report of an autopsied case.

A case of IgA (kappa type) myeloma complicated by fractures of the bones and hyperviscosity syndrome is presented. A 56-year-old woman who had been followed as an outpatient with diabetes mellitus for about 16 years, developed multiple myeloma. She also showed symptoms and signs of hyperviscosity syndrome; hemorrhagic diathesis, blurred vision and episodes of transient ischemic attacks of the brain, and fractures of the bones by small powers of trauma. At autopsy, almost all bones showed infiltration of multiple myeloma of IgA-kappa type and severe osteoporosis accompanied with proliferation of osteoclasts. The association of IgA myeloma with hyperviscosity syndrome and/or bone destruction was discussed. The "serum hyperviscosity syndrome" has been described clinically as the triad of bleeding, visual signs and symptoms, and neurologic manifestations. And this syndrome has been associated frequently with macroglobulinemia of Waldenström, occasionally with immunoglobulin (Ig) G myeloma, rarely with IgA myeloma, and with other dysproteinemia. Myeloma is also characterized by extensive bone destruction and is accompanied by susceptibility to fracture, although Waldenström's macroglobulinemia, acute leukemia or chronic leukemia are rarely associated with bone resorption. The present report describes a patient with IgA myeloma complicated by hyperviscosity syndrome and multiple bone fractures.

A method for predicting steady-state rate of skin penetration in vivo.

A simple in vivo method was proposed for predicting the steady-state rate of penetration of drugs across the stratum corneum. Both the diffusion coefficient and the partition coefficient in the stratum corneum can be determined by the amounts of drug in the stratum corneum at two time intervals under transient conditions after transdermal drug application. The amount of drug entering the stratum corneum is determined by 20 strippings with an adhesive tape. The steady-state rate of penetration was then calculated for the thickness of the stratum corneum and the concentration of the donor solution. The steady-state rates of penetration of ascorbic acid and estradiol across hairless mouse skin were evaluated from this in vivo approach and compared with those obtained from in vitro penetration experiment using excised hairless mouse skin. The data confirmed that the proposed in vivo method can predict the steady-state rate of penetration of these drugs across the stratum corneum in normal skin.

Stimulation of growth hormone secretion by central administration of atrial natriuretic polypeptide in the rat.

Intracerebroventricular (icv) injection of alpha-human atrial natriuretic polypeptide (alpha hANP) or alpha-rat ANP (0.6 and 3 nmol/rat) elicited an increase in plasma GH levels both in conscious freely moving rats and in urethane-anesthetized rats when given at the trough of spontaneous GH secretion. Antiserum specific for rat GRF did not affect the plasma GH increase induced by icv injection of alpha hANP. Intracerebroventricular injection of alpha ANP (3 nmol/rat) failed to stimulate GH secretion in conscious rats pretreated with cysteamine (30 mg/100 g BW, sc), a depletor of somatostatin (SRIF), and in conscious rats during constant iv infusion of SRIF (55 ng/ml). GH release induced by iv injection of synthetic rat GRF (200 ng/100 g BW) was exaggerated by alpha hANP (3 nmol/rat, icv) in conscious rats. These results suggest that central ANP stimulates pituitary GH secretion possibly by inhibiting SRIF release from the hypothalamus in the rat.

Drug permeation across the skin: effect of penetrant hydrophilicity.

The hydrophilicity of progesterone, a lipophilic steroid itself, was progressively increased by incorporating one or more hydroxy substituents at different positions on the steroidal skeleton. Effects of these hydrophilic substituents on the permeation of progesterone across the intact skin and stripped skin of the hairless mouse were studied using a hydrodynamically well-calibrated in vitro skin permeation system. The steady-state rate of permeation across the intact skin and stripped skin was found to be approximately proportional to the solubility of drugs in the stratum corneum or in the viable skin, respectively. Furthermore, the solubility of progesterone and its hydroxyl derivatives in the stratum corneum was noted to decrease gradually as the hydrophilicity of the penetrant increased. This finding was similar to that of a previously reported study of drug permeation across the lipophilic silicone membrane. However, the solubility of these progestins in the viable skin was observed to be dependent not only on the penetrant hydrophilicity but also on the position of the OH group on the penetrant molecule. The diffusivity of progesterone and its hydroxyl derivatives across the stratum corneum and viable skin was almost independent of the hydrophilicity of the drugs.

Stimulation by serotonin of immunoreactive peptide histidine isoleucine release from rat hypothalamus in vitro.

The effect of serotonin (5-HT) on the release of peptide histidine isoleucine-like immunoreactivity (PHI-LI) from rat hypothalamus was investigated in vitro with a perifusion system. A high potassium concentration (56 mM) stimulated PHI-LI release in a calcium-dependent manner. PHI-LI release was dose-relatedly stimulated by 5-HT (10(-6), 10(-5) and 10(-4) M). PHI-LI release induced by 5-HT (10(-5) M) was abolished by cyproheptadine (10(-4) M), a 5-HT antagonist. These results suggest that 5-HT has a stimulating effect on PHI release from the hypothalamus.

Involvement of peptide histidine isoleucine (PHI) in prolactin secretion induced by serotonin in rats.

The possible role of hypothalamic peptide histidine isoleucine (PHI) in prolactin (PRL) secretion induced by serotoninergic mechanisms was investigated in male rats using a passive immunization technique. Intracerebroventricular injection of serotonin (5HT, 10 micrograms/rat) raised plasma PRL levels both in urethane-anesthetized rats and in conscious rats pretreated with normal rabbit serum (0.5 ml/rat, iv, 30 min before). Plasma PRL responses to 5HT were blunted in these animals when they were pretreated with rabbit antiserum specific for PHI (0.5 ml/rat, iv, 30 min before) (mean +/- SE peak plasma PRL: anesthetized rats 271.3 +/- 38.3 ng/ml vs 150.0 +/- 12.6 ng/ml, p less than 0.01, conscious rats 54.3 +/- 6.8 ng/ml vs 30.7 +/- 4.1 ng/ml, p less than 0.025). These results suggest that hypothalamic PHI is involved, at least in part, in PRL secretion induced by central serotoninergic stimulation in the rat.

Stimulation by leumorphin of prolactin secretion from the pituitary in rats.

The effect of leumorphin (LM), one of big leu-enkephalins derived from preproenkephalin B, on PRL secretion was studied in the rat in vivo and in vitro. Intracerebroventricular injection of synthetic porcine LM (0.06-6 nmol/rat) caused a dose-related increase in plasma PRL levels in urethane-anesthetized male rats and in conscious freely moving rats. Intravenous injection of LM (3 nmol/100 g BW) also raised plasma PRL levels in these animals. The plasma PRL response to intracerebroventricular LM (0.6 nmol/rat) was blunted by naloxone (125 micrograms/100 g BW, iv). The stimulating effect of LM on PRL release was the most potent among the peptides derived from preproenkephalin B. In in vitro studies, PRL release from superfused anterior pituitary cells was stimulated in a dose-related manner by LM (10(-9)-10(-6) M), and the effect was blunted by naloxone (10(-5) M). These results suggest that LM has a potent stimulating effect on PRL secretion from the pituitary in the rat by acting, at least in part, directly at the pituitary through an opiate receptor.