Rhinocerebral mucormycosis in IDDM. Sequential magnetic resonance imaging of long-term survival with intensive therapy.

OBJECTIVE: To describe the clinical course and the utility of computerized tomography (CT) and magnetic resonance imaging (MRI) in the successful management of an often fatal fungal infection in a 12-year-old patient with insulin-dependent diabetes mellitus (IDDM). CASE: The patient was admitted to The University of Mississippi Medical Center (UMC) for the purpose of diabetic ketoacidosis (DKA) management and subsequent intensive therapy for mucormycosis according to nationally accepted standards of care. Strict diabetic control was instituted with frequent monitoring of blood glucose levels and interval assessment of HbA1c. Sequential MRI studies were obtained according to approved patient standards; the clinical and MRI course of the infection was charted. RESULTS: The patient's DKA resolved within 12 h on intravenous fluid repletion and insulin therapy. His sinusitis/rhinitis noted on admission did not respond to intravenous antibiotic therapy and progressed with obvious left orbital involvement and left cranial nerve palsies by 72 h of hospitalization. CT and MRI were invaluable aids to the early diagnosis and design of appropriate surgical and antifungal management of this patient, who survived with minimal left cranial nerve palsies. CONCLUSIONS: Our patient is among the youngest of IDDM patients reported to have survived rhinocerebral mucormycosis. His survival is attributed to early recognition of possible mucormycosis with diagnostic support of CT and MRI, surgical debridement and antifungal therapy, and intensive blood glucose control. Sequential MRI is invaluable to the design of therapy for this type of patient and shows the nearly 3-year recovery from mucormycosis.

Some limitations of using equilibrium dialysis to study human serum albumin-testosterone interaction.

The apparent association constant of testosterone binding to pure human serum albumin was found to be significantly less at 5 g/dl (1.79 x 10(4) M-1) than at 1 g/dl (2.29 x 10(4) M-1). This demonstrates that the common assumption that binding site characteristics are constant with serum protein dilution is not strictly valid. Determination of the percent free testosterone at normal serum protein concentrations by equilibrium dialysis may also be in error by 20% or more due to fluid shifts within the test system.

The rapid ovarian secretory response to pituitary stimulation by the gonadotropin-releasing hormone agonist nafarelin in sexual precocity.

We evaluated a GnRH agonist (GnRHa) as a potential single stimulus to both pituitary and ovarian secretion in 13 girls with true precocious puberty. We compared the GnRH agonist [6-D-(2-naphthyl)alanine]GnRH acetate (nafarelin, Syntex) administered as a single sc injection of 0.2 microgram/kg to GnRH infused iv in a dose of 2 micrograms/kg X h for 3 h and assessed the response of plasma steroid intermediates in estradiol (E2) biosynthesis. Although serum LH and FSH levels increased to similar peaks 3 h after commencing GnRH and nafarelin testing, they rose faster (P less than 0.01 at 1 h) and remained elevated longer (P less than 0.05 at 24 h) after nafarelin administration. At the third hour of testing with either agent, LH and FSH rose 8.8- and 3.4-fold, respectively (P less than 0.001 vs. baseline), whereas the rise in E2 was inconsistent and averaged only one third (P less than 0.02). However, plasma E2 increased later after nafarelin, but not after GnRH, rising from a baseline level of 30 +/- 6 (+/- SEM) to 115 +/- 13 pg/ml at 24 h (P less than 0.001). The least E2 response to nafarelin at this time was 150%. This rise is probably an underestimate of the maximum E2 rise, since a 6-fold response to nafarelin was found at 12 h in patients sampled then. Measurement of steroid intermediates from progesterone and 17 alpha-hydroxypregnenolone to E2 indicated that the response to nafarelin was typical of normal ovarian follicular secretion. That is, plasma levels of the intermediates in E2 biosynthesis rose less than 2-fold, and only the elevations in androstenedione, from 58 +/- 10 to 78 +/- 16 ng/dl (P less than 0.05), and estrone, from 14 +/- 3 to 38 +/- 7 pg/ml (P less than 0.02), at 24 h were significant. The greater effectiveness of nafarelin than GnRH in stimulating E2 secretion appears to be related to the more prolonged gonadotropin response. The magnitude, consistency, specificity, and rapidity of the gonadotropin and E2 responses to nafarelin indicate that this is a promising agent for rapidly testing pituitary and ovarian function simultaneously.

A limited repertoire of mutations of the luteinizing hormone (LH) receptor gene in familial and sporadic patients with male LH-independent precocious puberty.

Herein, we report mutation analysis of the LH receptor gene in 17 males with LH-independent precocious puberty, of which 8 were familial and 9 had a negative family history. A total of 7 different mutations (all previously reported) were detected in 12 patients. Among 10 European familial male-limited precocious puberty (FMPP) patients who had a LH receptor gene mutation, none had the Asp578Gly mutation, which is responsible for the vast majority of cases in the U.S. The restricted number of activating mutations of the LH receptor observed in this and other studies of FMPP strongly suggests that an activating phenotype is associated with very specific sites in the receptor protein. Clinical follow-up of the 5 patients who did not have LH receptor mutations shows that such cases most likely do not have true FMPP. LH receptor mutation analysis provides a sensitive tool for distinguishing true FMPP from other causes of early-onset LH-independent puberty in males.

Two tumors detected by thyroid assessment in two children.

OBJECTIVE: To describe the early detection of two tumors in two children by recognition of unusual features in initial thyroid assessments. METHODS: We present the clinical findings and results of laboratory studies in two children. In addition, we describe RET proto-oncogene studies in one of them. RESULTS: A 14.5-year-old boy was referred for assessment because of short stature in conjunction with lack of physical growth and development. His physical examination was remarkable for height at the 50th percentile (height age, 11.5 years), weight at the 50th percentile (weight age, 13 years), and prepubertal male status. Pertinent laboratory findings were a normal thyroid-stimulating hormone (TSH) level but low free thyroxine (FT4) index. These findings prompted an immediate magnetic resonance imaging study of the head. A parasellar tumor was detected and removed; histopathologic examination revealed that it was a craniopharyngioma. The patient requires lifelong multihormonal therapy for his panhypopituitarism and has responded with physical growth. Our second patient, a 7.5-year-old girl, was referred because of a painless left thyroid nodule of 4 months' duration. Her physical examination was remarkable for (1) upper lip swelling, (2) intermittent rash, and (3) a goiter with painless mobile left and right nodules. Normal levels of TSH and FT4, serum calcitonin of 6,192 pg/mL, and a fine-needle biopsy specimen that stained strongly for calcitonin were obtained at her first clinic visit. A total thyroidectomy was performed and confirmed the presence of medullary thyroid carcinoma. Genetic studies showed that she was positive for the RET multiple endocrine neoplasia IIB mutation. After 4 years of follow-up, the patient had serum calcitonin levels that remained low (<2.2 pg/mL). CONCLUSION: Attention to thyroid physical findings and laboratory studies can promptly lead to correct diagnoses and management of some rare and life-threatening tumors in children.

Estradiol inhibition of pituitary luteinizing hormone release is antagonized by serum proteins.

We tested the influence of protein binding upon the rapid 17 beta-estradiol (E2) inhibitory effect on luteinizing hormone (LH) responsiveness to LH releasing hormone (LHRH) in perifused rat anterior pituitaries. LHRH pulses were given before 1 h after changing the perifusion medium to contain various combinations of protein and E2. In the absence of albumin an E2 concentration of about 27 pg/ml inhibited LH secretion in response to LHRH by 50% (judged from the change in secretion ratio, i.e LH response to LHRH pulse no. 2 divided by that to pulse no. 1). Albumin (1 g/dl) and human plasma enhanced the self-priming effect of LHRH (P less than 0.1) and blunted the slope of the dose-response relationship between E2 and LH secretion. These effects suggest a stimulatory effect of proteins on LH secretion which is independent of E2. However, proteins appear to antagonize the E2 effect in part by binding E2. A total E2 dose of 270 pg/ml or more was required to achieve about 50% inhibition of the LH secretion ratio in the presence of albumin. Since 20% of E2 was unbound in the presence of albumin, the 50% effective dose of free E2 was about 54 pg/ml. This closely approximates the comparably effective dose of E2 in the absence of protein. Although a total E2 concentration of 216 pg/ml was required for a 50%-inhibitory E2 effect in the presence of heat-inactivated plasma, this was equivalent to between 28 and 37 pg/ml of free E2 in these experiments. The effect of E2 in a total dose of 216 pg/ml was attenuated more by the use of TEBG-positive plasma than of TEBG-negative (heat-inactivated) plasma (P less than 0.01). Our data indicate that the E2 inhibitory effect upon LH responsiveness to LHRH is attenuated by albumin and TEBG to approximately the extent expected by binding of E2 to these proteins. Our data are certainly contrary to those expected if either albumin or TEBG augmented this E2 action. Our data support the concept that the bioavailable fraction of plasma sex steroids is that which is free from binding to plasma proteins.

Pediatric Graves' disease: therapeutic options and experience with radioiodine at the University of Mississippi Medical Center.

BACKGROUND: Pediatric Graves' disease can be life-threatening, and it adversely alters growth and development. Controversies concerning optimal therapy led us to review our 40 pediatric patients treated for Graves' disease from 1988 to 1996 to assess efficiency, efficacy, and safety of current therapy options. METHODS: Diagnosis of Graves' disease required clinical hyperthyroidism with supportive laboratory studies. Patients were given informed choices of therapy, which divided them into three groups. RESULTS: In group 1, 17 patients received antithyroid medications for 0.3 years to 6.0 years. Three required surgical thyroidectomy. Remissions (with or without thyroxine therapy) were achieved after 2 years to 5 years in 11 (65%). In group 2, 15 patients received antithyroid medications for 0.3 years to 5.0 years before receiving radioactive iodine (131I). One also required surgical thyroidectomy. Remissions were achieved after 1 year to 5 years in 10 (67%). In group 3, eight patients received initial 131I therapy. Remissions were achieved within 1 year in 7 (88%). CONCLUSIONS: Our results agree with and expand upon published reports on Graves' disease. Our data support early use of 131I as efficient, effective, and safe therapy for pediatric Graves' disease.

Plasma free testosterone in the diagnosis of adolescent polycystic ovary syndrome.

We have employed plasma free testosterone concentrations to diagnose polycystic ovary syndrome in adolescence after demonstrating that mature plasma free androgen levels are achieved by midpuberty. Among 43 female volunteers, 10 to 16 years of age, followed up in a mixed longitudinal-cross-sectional study, we found that three (7%) had elevated plasma free testosterone levels in the absence of symptoms or signs of hyperandrogenemia. Of two who were followed up, the abnormality resolved on completion of sexual maturation in one, and in the other polycystic ovary syndrome was subsequently diagnosed. We diagnosed this syndrome in 14 adolescents as the basis of menstrual disorders associated with hirsutism, acne, or obesity. The diagnosis was based on finding elevated free testosterone values, which did not decrease after adrenal suppression by dexamethasone. Serum concentrations of luteinizing hormone were elevated in most but not all. In some of these patients further confirmation of polycystic ovary syndrome was obtained by laparoscopy or ultrasound examination. These studies demonstrate that measurement of plasma free testosterone before and after administration of dexamethasone appears to be the most sensitive single method for detecting polycystic ovary syndrome.

Direct inhibitory effect of estradiol on pituitary luteinizing hormone responsiveness to luteinizing hormone releasing hormone is specific and of rapid onset.

We have employed a perifusion technique to explore the time course and specificity of 17 beta-estradiol (E2) effects directly upon luteinizing hormone (LH) release from the isolated rat anterior pituitary under pulsatile luteinizing hormone releasing hormone (LHRH) stimulation. We first characterized the perifusion system and fitted the data to a simple dose-response model. Multiple perifusion studies were then performed with pulses of LHRH at approximately half-maximal response concentration (10(-8) M); LH responses to an initial LHRH pulse (#1) were compared with LH responses to a second LHRH pulse (#2) given at variable times after addition of E2, antiestrogen (LY 117018), and/or 17 alpha-estradiol (17 alpha-E2). Using this approach, we found that the direct inhibitory effect of E2 upon LH responsiveness to LHRH was rapid and specific. The ratio of LH secretion in response to LHRH pulse #2 to that in response to LHRH pulse #1 (LH secretion ratio) decreased steadily during 1 h of exposure to E2. This inhibition was significant (P less than 0.01) by 36 min of E2 exposure. It represented more than the removal of a LHRH self-priming effect because the LH secretion ratios were significantly less than 1.0 [0.80 +/- 0.05 (SEM), P less than 0.01] within 36 min of E2 exposure. The inhibitory effect was not seen when LY 117018 was added with E2, nor when 17 alpha-E2 replaced E2. The specificity of this rapid E2 inhibitory effect upon pituitary LH responsiveness to LHRH strongly suggests that it is receptor mediated. The rapidity of this apparent receptor-mediated estrogen effect suggests that it is a very rapid consequence of nuclear translocation of the E2-receptor complex.

Phosphorylation of histone catalyzed by a bovine brain protein kinase.

The use of polyethyleneimine-cellulose thin layer sheets to follow the phosphorylation of histone and decomposition of ATP catalyzed by an adenosine 3':5'-monophosphate (cyclic AMP)-stimulated protein kinase, protein kinase I, has made possible a more detailed analysis of the time course of these reactions than has been achieved previously be observing only recovered phosphorylated protein. When [gamma-32P] ATP was employed, significant error was introduced by the presence of 32Pi at the solvent front on these sheets, and this limited the accuracy of the available information. However, the analysis of assays performed with [U-14C] ATP was straightforward and appeared to have an accuracy comparable to that of the present standard assay. This appears to be the first use of [U-14C] ATP to assay protein kinases. Our physical characterization of protein kinase I showed it to be a homogeneous protein species by polyacrylamide gel electrophoresis, sodium dodecyl sulfate gel electrophoresis and analytical ultracentrifugation. Kinetic studies with protein kinase I indicated the absence of histone phosphatase and cyclic AMP phosphodiesterase activity. Furthermore, the ATPase activity seen is believed to be intimately associated with the protein kinase action, particularly in view of the observed dependence of the rate of Pi production on the presence of cyclic AMP. The kinetic data for the phosphorylation of histone catalyzed by protein kinase I under full stimulation by cyclic AMP are consistent with a double displacement mechanism.

Ionic inhibition of catalytic phosphorylation of histone by bovine brain protein kinase.

The effects of various ions commonly found in protein kinase assays upon the rate of histone phosphorylation catalyzed by the highly purified bovine brain enzyme, protein kinase I, have been investigated. Sodium, potassium, and magnesium were found to inhibit histone phosphorylation by protein kinase I in a similar manner. The degree of inhibition by any of these cations was demonstrated to be directly proportional to the square root of the ionic strength of the assay medium. The relationship between the ionic strength of the assay medium and the rate of histone phosphorylation catalyzed by protein kinase I was employed to correct the rate of histone phosphorylation at various magnesium acetate concentrations to a standard ionic strength. When this was done an analysis of the previously postulated rate law for histone phosphorylation c atalyzed by protein kinase I gave a binding constant for the magnesium-ATP complex which was in agreement with that expected for this complex on the basis of various binding constants available in the literature. These results demonstrate that it is unnecessary to postulate a specific ion inhibition process for protein kinase I by the ions employed in this study. They also support the reasonable assumption that magnesium ion binds to ATP at or prior to the rate-determining step in histone phosphorylation catalyzed by protein kinase I. The expression developed in this paper for the effect of ionic strength upon protein kinase I activity can now be used to correct activity measurements made under various assay conditions to a standard assay state, allowing facile comparisons of kinetic data. It should be possible to develop similar expressions for other protein kinases and substrates to permit useful interpretation of kinetic data.

Administration of low-dose estrogen rapidly and directly stimulates growth hormone production.

We tested the concept that estrogen directly stimulates growth hormone (GH) production by determining whether low-dose treatment with ethinyl estradiol increases the GH reserve, as assessed by levodopa administration, without inhibiting somatomedin-C (Sm-C) levels. Twenty-three prepubertal short normal children underwent levodopa tests before and after being treated with ethinyl estradiol. One bedtime dose of ethinyl estradiol (20 to 40 micrograms/sq m, n = 8) resulted in a significant increase in GH levels during levodopa testing, with no significant change in Sm-C levels (0.27 +/- 0.03 vs 0.36 +/- 0.1 units/mL). Two days of a comparable ethinyl estradiol dose (n = 12) raised the mean basal GH level (2.4 +/- 0.4 vs 9 +/- 3 ng/mL) and had a similar effect on peak GH response, without affecting the mean Sm-C level. Eighteen of the 23 patients responded (maximum GH level, greater than or equal to 7 ng/mL) to levodopa before estrogen; all 20 children who received ethinyl estradiol priming in a dose of 20 micrograms/sq m or more also responded. We conclude that low-dose estrogen therapy rapidly stimulates GH production without decreasing Sm-C plasma levels. These results support the concept that the estrogen effect is direct. This action may be important for the stimulation of growth by estrogen. This effect can be conveniently employed to enhance the specificity of the levodopa test for profound GH deficiency.

Growth hormone deficiency impedes the rise in plasma insulin-like growth factor I levels associated with precocious puberty.

We tested the hypothesis that growth hormone (GH) mediates the rise in insulin-like growth factor I (IGF-I) concentrations in children with precocious puberty. We studied three groups of patients. Group 1 included six children with GH deficiency and precocious puberty (precocious GH-deficient); group 2 included 10 GH-sufficient patients with idiopathic true precocious puberty (precocious GH-sufficient); and group 3 included 9 prepubertal children with GH deficiency (prepubertal GH-deficient). Growth rates, pubertal status, and plasma IGF-I concentrations were determined at regular intervals. The precocious children with GH deficiency had a mean (+/- SD) growth rate of 7.2 +/- 2.1 significantly below that of the precocious GH-sufficient patients (10.5 +/- 2.5 cm/yr, p less than 0.05) but above that of the prepubertal GH-deficient children (3.9 +/- 1.4 cm/yr, p less than 0.05). The mean IGF-I concentration in the precocious GH-deficient children was 0.77 +/- 0.39 U/ml, significantly lower than the mean level of 2.2 +/- 0.67 U/ml in the precocious GH-sufficient patients (p less than 0.01). However, precocious GH-deficient patients had significantly higher IGF-I values than the prepubertal GH-deficient children (0.24 +/- 0.10 U/ml, p less than 0.05). IGF-I values did not rise with the onset of precocious puberty in four of the precocious GH-deficient children evaluated before and after the development of precocious puberty. However, three patients who began GH treatment did have a rise in plasma IGF-I concentrations to levels of 1.2, 3.4, and 3.7 U/ml, respectively. These findings are compatible with the concept that sex steroids increase IGF-I levels in precocious puberty primarily by increasing GH production. A small but direct effect of sex steroids on IGF-I production may also exist. The onset of precocious puberty in children with organic GH deficiency may mask the abnormal growth pattern of these children and delay diagnosis; determinations of plasma IGF-I concentrations may be helpful in assessing the GH status of these patients.

The relationship of maternal erythrocyte oxygen transport parameters to intrauterine growth retardation.

The relation of fetal growth and maternal oxygen transport as assessed by red blood cell 2,3-diphosphoglycerate, hemoglobin oxygen affinity, hemoglobin, pH, and PCO2 was evaluated in 21 pregnant women. The study was performed in the third trimester and each subject evaluated had sonographic evidence of fetal growth retardation without other obvious abnormalities. Decreased maternal 2,3-diphosphoglycerate/hemoglobin molar ratio and hemoglobin oxygen affinity were related linearly to the birth weight normalized for the expected sea level values of gestational age expressed as a birth weight (gestational age-normalized) Z score. The correlation coefficients and p values were r = 0.71, p less than 0.001 and r = 0.67, p less than 0.001, respectively. The ponderal index-normalized Z score correlated with the 2,3-diphosphoglycerate/hemoglobin molar ratio (r = 0.46, p less than 0.04), but the relation was not as strong as the birth weight-normalized Z score. The crown-heel length/head circumference ratio did not correlate with the 2,3-diphosphoglycerate/hemoglobin molar ratio (r = 0.29, NS). The birth weight (gestational age)-normalized Z score did not correlate with hemoglobin, PCO2, or pH. In the regulation of hemoglobin oxygen affinity, calculations indicated that the 2,3-diphosphoglycerate/hemoglobin molar ratio played a highly significant role (p less than 0.001), pH was minimally significant (p less than 0.025), but PCO2 had little or no significant effects in this study. It appears that fetal growth is related to the maternal red blood cell oxygen transport parameters 2,3-diphosphoglycerate/hemoglobin molar ratio and hemoglobin oxygen affinity. Moreover, the 2,3-diphosphoglycerate/hemoglobin molar ratio is the principal regulator of hemoglobin oxygen affinity.

Preserving adult height potential in girls with idiopathic true precocious puberty.

We designed a prospective study of height potential in girls with idiopathic precocious puberty, comparing the presenting features of girls with and without evidence of reduced adult height potential. The 14 girls with impaired adult height prognoses (group 1) were reexamined after treatment with a gonadotropin releasing hormone agonist, nafarelin. The seven girls with the prognosis of unimpaired height (group 2) were followed without therapy. We found that the group could be distinguished at initial examination by the greater bone age/height age ratio of group 1 (mean +/- SEM: 1.4 +/- 0.06 vs 1.0 +/- 0.05; p less than 0.005) and by the greater difference between predicted height and target height in group 1. The mean predicted height in group 1 was significantly less than the mean target height (150.7 +/- 2.1 vs 165.4 +/- 3.0 cm; p less than 0.005), whereas the mean predicted and target heights in group 2 were similar (165.4 +/- 3.0 vs 164.3 +/- 2.1 cm). Initial estradiol levels were also greater in group 1 than in group 2 (21.6 vs 10.6 pg/ml; p less than 0.05), although this difference was not sustained during follow-up. In group 1, nafarelin therapy suppressed the pituitary-gonadal axis, and although there was a transient reduction in height potential in girls with the youngest bone ages during the first 6 months of therapy, 2 years of treatment slightly improved predicted heights from 150.7 +/- 2.1 to 152.7 +/- 2.0 cm (p less than 0.05). Height predictions also increased without therapy during the 2-year observation period in group 2, from 165.4 +/- 3.0 to 168.7 +/- 4.1 cm (p less than 0.05). Our data indicate that gonadotropin releasing hormone agonist therapy preserves height potential in girls with an initially impaired height prognosis, and that height potential is preserved without therapy in those with a good initial height prognosis.

Cosegregation of missense mutations of the luteinizing hormone receptor gene with familial male-limited precocious puberty.

Familial male-limited precocious puberty is a male-limited autosomal dominant condition. It is characterized by increased testosterone synthesis in the absence of testicular stimulation by luteinizing hormone (LH). We hypothesised that an abnormal configuration of the LH receptor might autonomously activate G protein coupling, and thereby cause the overproduction of testosterone in this condition. To test this hypothesis, we screened for mutations in a part of the LH receptor gene that is important for G protein binding. DNA sequence variation was detected in 2 out of 5 families with male-limited precocious puberty by the single strand conformation polymorphism technique. Direct sequencing demonstrated different single nucleotide substitutions in the sixth transmembrane region of the LH receptor gene. The mutations cosegregated with the disorder in both families (lod score 5.76 without recombination). Both mutations cause an amino acid substitution in the sixth transmembrane domain, close to the C-terminal portion of the third cytoplasmatic loop, a region which is important for the binding of G proteins. We conclude that familial male-limited precocious puberty cosegregates with missense mutations in the LH receptor gene. These findings support the hypothesis that increased activity of the LH receptor is the pathogenetic mechanism that causes the abnormal pubertal development in this condition.