Total body zinc depletion and its relationship to the development of hyperprolactinemia in chronic renal insufficiency.

Modulation of free plasma zinc levels has been implicated in the increase in plasma prolactin levels seen in patients with chronic renal insufficiency (CRI). The relative importance of this mechanism in comparison to others, however, has not been elucidated. Zinc equilibrium between plasma and red blood cells is partly dependent upon red blood cell carbonic anhydrase (CA). In the present paper, we have investigated the interrelationships among total plasma zinc, leukocyte zinc, prolactin, and erythrocyte CA in patients with CRI. Uremic patients were shown to have significantly increased levels of plasma prolactin and erythrocyte CA activity when compared to normal controls. Moreover, red blood cell CA total concentration and isoenzyme-I and-II levels, as well as plasma zinc were found to be significantly decreased in uremic patients in comparison to normal controls. In patients with CRI, a negative correlation was demonstrated between erythrocyte CA catalytic activity and plasma zinc, as well as between plasma zinc and plasma prolactin levels. Moreover, leukocyte zinc content, which is a reliable indicator of total body zinc stores, was found to be significantly decreased in uremic patients when compared to normal controls. A strong negative correlation between leukocyte zinc content and plasma prolactin levels was documented in CRI patients. Our results suggest that alterations in erythrocyte CA levels, enzymatic activity or isoenzyme profile are most probably mechanistically and etiologically unrelated to the high plasma prolactin levels in CRI patients. Contrariwise, depletion of total body zinc stores, rather than redistribution of this trace metal among extracellular compartments, may represent one of the major contributing mechanisms leading to uremic hyperprolactinemia.

Hyperadrenergic state following acute withdrawal from clonidine used at supratherapeutic doses.

Abrupt cessation of clonidine treatment precipitates a physiological withdrawal syndrome, thought to be due to a hyperactive state of central autonomic and cognitive adrenergic neuronal systems dependent on presynaptic alpha 2-adrenoceptors and/or imidazoline receptors. We hereby describe a 36-year-old male with history of end-stage renal disease, hypertension and medication non-compliance, who presented with severe hypertension and remarkable agitation. His daily clonidine intake was estimated to be 10 mg. The patient had abruptly discontinued his clonidine five days prior to admission. The following indices of adrenergic activity were measured in plasma (normal control values in parentheses): noradrenaline (NA) 8.59 nmol/l (1.32-4.56 nmol/l), adrenaline (Adr) 1.86 nmol/l (0.83-4.20) nmol/l), total 3-methoxy-4-hydroxyphenylglycol (MHPG) 152.8 nmol/l (45.1-111.5 nmol/l), and free MHPG 33.0 nmol/l (12.2-31.4 nmol/l). Plasma clonidine level was 3.53 ng/ml (15.9 nmol/l) with the usual therapeutic level being < 2.0 ng/ml (8.9 nmol/l). Initially, the patient received sedatives and was started on clonidine for the first 24 hours only, after which time period prazosin was started, with good response of his blood pressure and reversal of his mental status changes. At that point, the plasma values of indices of adrenergic activity had decreased compared with their corresponding initial values by the following percentages: NA 60.6%, Adr 22.6%, total MHPG 42.2% and free MHPG 11.5%. Plasma clonidine level had decreased now by 43.6% to an absolute value of 1.99 ng/ml (8.85 nmol/l). We emphasize that physicians should be aware of clonidine's abuse potential and caution should be taken, as well as the appropriate route chosen, when prescribing clonidine in patients who show features of poor compliance to medications and especially in patients with psychoses, suicide potential or personality disorders.

Characterization and purification of the chorionic gonadotropin-like protein binding site in Candida albicans.

Recent studies from our laboratory have shown that human chorionic gonadotropin (hCG), human luteinizing hormone (hLH), and CG-like proteins from Xanthomonas maltophilia (xCG) and Candida albicans (CaCGLP) induce Candida albicans transition from blastospores to hyphal forms. Xanthomonas maltophilia CG-like protein (xCG), hCG, and hLH also bind to Candida albicans blastospores with a high-affinity nM Kd, indicating that these substances can control Candida albicans pathogenicity. The work reported herein describes the purification of the binding site for these CG-like proteins from Candida albicans. The purification developed involved alcohol extraction followed by affinity chromatography. The product obtained was a protein of 64-69 kDa, as analyzed by sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE), Western blot and Sephadex G-100 column chromatography. This binding site reacted in a Western blot with both 125I-hCG and 125I-CaCGLP. Purified CaCGLP was able to displace specifically 125I-hCG bound to Candida albicans blastospores. Scatchard plot analysis showed that the Kd of this reaction was of high affinity in the nM range, and also indicated the presence of one single binding site. These results lead us to conclude: 1) Candida albicans possesses a binding site which is able to bind hCG, hLH and CG-like proteins from Xanthomonas maltophilia and Candida albicans; 2) This binding site is a hydrophobic protein of approximately 64 kDa and; 3) We postulate that CaCGLP is the natural ligand for this binding site and this system is used to control Candida albicans transition and, therefore, pathogenicity.

The presence of a human chorionic gonadotropin-like protein and its binding site in Saccharomyces cerevisiae.

In this study, we characterize from Saccharomyces cerevisiae: 1) a protein that has immunological similarities to human chorionic gonadotropin (hCG), and 2) a binding site for this hCG-like protein which also binds hCG and human luteinizing hormone (hLH). Saccharomyces cerevisiae chorionic gonadotropin-like protein (ScCGLP) was purified in several steps. This protein when analyzed by SDS-PAGE, under nondenaturing conditions, produced two bands, one at 110-kDa, and another at 57.5-kDa. Under denaturing conditions, only the 57.5-kDa band appeared. This 57.5-kDa band also reacted in a Western blot, using a polyclonal antibody directed against hCG. Purified ScCGLP reacted in the following hCG immunoassays: 1) polyclonal rabbit anti-hCG equilibrium assay, 2) carboxyl-tail hCG equilibrium assay, 3) two equilibrium assays using monoclonal antibodies, and 4) free alpha-chain subunit equilibrium assay using a monoclonal antibody. Characterization of hCG/hLH binding sites in Saccharomyces cerevisiae was performed, and the ability of the ScCGLP to displace I125-hCG was also shown. Human CG and hLH were able to compete for I125-hCG binding to Saccharomyces cerevisiae blastospores (Kds of approximately 10(-8) M), while ScCGLP competed with higher affinity (Kd = 9.41 x 10(-10) M). The hCG-like immunoactivity was also present in Saccharomyces growth media, as well as in all brands of commercial beer studied.

Estradiol stimulates cortisol production by adrenal cells in estrogen-dependent primary adrenocortical nodular dysplasia.

Adrenal glands from a patient with ACTH-independent Cushing's syndrome, whose symptoms worsened during pregnancy and oral contraceptive use, were cultured in different concentrations of estradiol. Estradiol stimulated cortisol secretion in a dose-response manner in the absence of ACTH. Since immunoglobulins G from this patient did not stimulate corticosterone production in a mouse adrenal bioassay, an adrenal-stimulating immunoglobulin is unlikely to be the cause of adrenal hyperfunction in this case. This is the first description of estradiol stimulation of cortisol production by cultured adrenal cells in ACTH-independent Cushing's syndrome.

Characterization of a human chorionic gonadotropin-like protein from Candida albicans.

Studies from our laboratory have demonstrated that human CG (hCG), human LH, (hLH), and an hCG-like protein extracted from Xanthomonas maltophilia were able to induce Candida albicans transition from the blastospore to the germ tube stage. In the present study, we describe the characterization of an hCG-like material extracted from Candida albicans blastospores (CaCGLP), which is potent in inducing transition and presumably represents the endogenous transition-inducing substance. This material was extracted from Candida albicans blastospores with glacial acetic acid and purified by affinity chromatography using a polyclonal rabbit anti-hCG antibody. The product obtained is a 68-kilodalton single band protein, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis. Under reduced conditions a protein smear is seen. Amino acid analysis showed a predominance of glycine (22%), followed by serine (12%), and glutamate (12%). This protein reacted in the following hCG immunoassays: 1) a polyclonal rabbit anti-hCG equilibrium assay, 2) a carboxyl-tail hCG equilibrium assay, 3) two hCG equilibrium assays using monoclonal antibodies (CG no. 4 and CG no. 9), 4) a free alpha-subunit equilibrium assay using a monoclonal antibody, and 5) an ultrasensitive immunoradiometric assay for hCG which does not cross-react with hLH, nor the free beta-subunit of hCG. The CaCGLP showed no reaction in a specific hLH immunoradiometric assay. When CaCGLP was tested in the transition assay, in the presence of 4% rat serum, it was found that this protein was 100 times more potent than hCG in producing Candida albicans transition. We conclude that Candida albicans produces a protein that has certain tertiary structure similarities to hCG and that this material is able to induce germ tube formation. We postulate that CaCGLP has an autocrine/paracrine effect in Candida albicans as a transition factor to control its own pathogenicity.

Partial nucleotide sequence of the Xanthomonas maltophilia chorionic gonadotropin-like receptor.

Xanthomonas maltophilia possesses a unique high-affinity binding site which binds human chorionic gonadotropin (hCG), but not human luteinizing hormone (hLH) or other glycoprotein hormones. We designed primers from the known nucleotide sequence of the human LH/CG receptor, spanning an area extending from transmembrane region 2 to transmembrane region 6. Genomic DNA extracted from Xanthomonas maltophilia was used to obtain a PCR amplified product using the above primers. The primary amplification product was cloned in a pCR11 TA cloning vector, and the partial nucleotide sequence of the gene determined. This determined sequence showed 73% identity with the human, as well as the rat LH/CG receptor. Comparison of the translated protein sequence with the human, rat and porcine receptor protein sequences showed a 52% similarity.

Stimulation of Candida albicans transition by human chorionic gonadotrophin and a bacterial protein.

Candida albicans, a dimorphic fungus, is involved commonly in human infections with the mycelium form more associated with pathogenicity. The influence of various hormones and a bacterial protein on the transition from blastospore to mycelium was assessed. Human luteinizing hormone (hLH), chorionic gonadotrophin (hCG), and an hCG-like material purified from a bacteria, Xanthomonas maltophilia (PCG), were able to increase the rate of transition when compared with the controls. The effect of the two hormones and the bacterial peptide were specific, as human follicle stimulating hormone (hFSH), thyroid stimulating hormone (hTSH), growth hormone (hGH), prolactin (hPrl) and rat and bovine LH (rLH, bLH), and bovine albumin and gamma globulin did not affect the transition. The binding of 125I-hCG or 125I-LH to spheroplasts of Candida albicans were competitively displaced by hCG, hLH, and PCG. Scatchard analysis of binding of all three ligands revealed two binding sites with a high-affinity nM Kd. Thus, hCG, hLH, and PCG induce transition of Candida albicans from a blastospore state to a mycelium form, suggesting that these hormones may modify the pathogenicity of Candida albicans.