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Obesity, lipodystrophy, diabetes, and hypertension collectively constitute the main features of Metabolic Syndrome (MetS), together with insulin resistance (IR), which is considered as a defining element. MetS generally leads to the development of cardiovascular disease (CVD), which is a determinant cause of mortality and morbidity in humans and animals. Therefore, it is essential to implement and put in place adequate management strategies for the treatment of this disease. Catestatin is a bioactive peptide with 21 amino acids, which is derived through cleaving of the prohormone chromogranin A (CHGA/CgA) that is co-released with catecholamines from secretory vesicles and, which is responsible for hepatic/plasma lipids and insulin levels regulation, improves insulin sensitivity, reduces hypertension and attenuates obesity in murine models. In humans, there were few published studies, which showed that low levels of catestatin are significant risk factors for hypertension in adult patients. These accumulating evidence documents clearly that catestatin peptide (CST) is linked to inflammatory and metabolic syndrome diseases and can be a novel regulator of insulin and lipid levels, blood pressure, and cardiac function. The goal of this review is to provide an overview of the CST effects in metabolic syndrome given its role in metabolic regulation and thus, provide new insights into the use of CST as a diagnostic marker and therapeutic target.

Diabetes insipidus (DI) is a syndrome characterized by the persistent excretion of abnormally large volumes of dilute urine. It can be caused by any of four fundamentally different abnormalities: deficient production of the antidiuretic hormone, arginine vasopressin (AVP) by magnocellular neurons that form the posterior pituitary (hypothalamic DI); impaired renal effects of AVP (nephrogenic DI); reduced AVP secretion due to excessive water intake (primary polydipsia); or degradation of AVP by placental vasopressinase (gestational DI). Each type of DI can be caused or potentiated by other disorders. Hypothalamic and nephrogenic DI can also be caused by mutation of the gene that encodes the AVP prohormone, the AVP-2 receptors in the kidney, or the aquaporin-2 water channels that mediate antidiuresis. Familial hypothalamic DI is usually transmitted in an autosomal dominant mode, but autosomal recessive or X-linked recessive forms also exist. Familial nephrogenic DI is usually transmitted in an X-linked recessive mode but can also be autosomal recessive or dominant. Hence the mode of inheritance does not always indicate the type of DI. Indirect methods of differential diagnosis are also unreliable and the pituitary MRI signal is diminished in both types of familial DI. Thus the determination of plasma AVP and/or the response to desmopressin therapy plus gene sequencing provides the best basis for effective management and family counseling.

Vitamin D differs from other vitamins in that it is not a true vitamin but a prohormone obtained by endogenous synthesis from sun exposure on 7-dehydrocholesterol in the skin and from certain foods and dietary supplements. Its beneficial effects and physiological functions on musculoskeletal health are well understood. Mounting reports have recognized an association of vitamin D deficiency with a myriad of acute and chronic illnesses including autoimmune disorders, infectious diseases, cardiovascular diseases, common cancers, and neurological disorders, suggesting an important role of vitamin D in nonmusculoskeletal functions. Here, the role of vitamin D in skeletal health and present data on vitamin D in other nonmusculoskeletal diseases are reviewed.

Some natural compounds decrease serum levels of thyroid autoantibodies, but results are inconsistent and thyroid function has been evaluated infrequently; moreover, the effects of Aloe on thyroid autoimmunity and function have been examined in very few studies. This study stems from the observation of one co-author, who has Hashimoto’s thyroiditis (HT)-related subclinical hypothyroidism (SCH). Upon checking her biochemical thyroid panel when taking daily Aloe barbardensis Miller juice (ABMJ) for thyroid-unrelated reasons, she noticed a decrease in serum thyroperoxidase autoantibodies (TPOAb) and thyrotropin (TSH) and an increase in serum free thyroxine (FT4). Based on this observation, we enrolled 30 consecutive HT women with levothyroxine-untreated SCH and high TPOAb levels. All of them took ABMJ (50 ml daily) for nine months and were tested for serum TSH, FT4, free triiodothyronine (FT3) and TPOAb. Measurements were performed at baseline and at months 3 and 9. TSH, FT4 and TPOAb improved significantly already at month 3 and further (−61%, +23% and −56%) at month 9. However, FT3 decreased significantly at month 3 (−16%) with no further decrease at month 9, so that the FT4:FT3 ratio increased significantly (+33% and + 49%). At baseline, 100% of women had TSH > 4.0 mU/L and TPOAb > 400 U/ml, but frequencies fell to 0% and 37%, respectively, at month 9. In contrast, a control group (namely, 15 untreated SCH women of comparable age and baseline levels of TSH, FT4, FT3 and TPOAb) had no significant changes in any index. We conclude that the daily intake of 100 ml ABMJ for 9 months in women with HT-related SCH decreases the burden of thyroid autoimmune inflammation. In addition, ABMJ rescues thyrocyte function, with decreased need for conversion of the prohormone T4 into the more active T3 through ABMJ-induced inhibition of T4 deiodination. Keywords: Aloe vera, Subclinical hypothyroidism, Thyroid autoimmunity, Thyroid function

Peptide hormones represent a major class of hormones that are made from amino acids by specialized endocrine glands. The maturation of bioactive hormones take place in the rough endoplasmic reticulum and Golgi apparatus, where preprohormones are proteolytically cleaved into prohormones, and subsequently into mature peptide hormones. Once the bioactive hormones are released into the circulation, they interact with receptors located on the plasma membrane of target cells, and initiate intracellular signaling pathways to regulate physiological processes including energy metabolism, growth, stress, and reproduction. However, excessive amount of circulating peptide hormones often associates with the presence of tumors. Section 2 discusses 10 peptide hormones as tumor markers and their clinical application in aiding the diagnosis of tumors as well as monitoring the disease process.

Thyroxine (T4) is an iodinated amino acid derivative primarily synthesized in the thyroid epithelial cells. T4 is a major TH but less potent than 3,3′,5-triiodothyronine (T3). T4 has been thought to be a prohormone of the active form, T3. The fact that the binding affinity of nuclear thyroid hormone receptors (TRs) for T3 is one order of magnitude higher than that for T4 indicates that T3 is the natural ligand of TRs. Recent studies have demonstrated physiological actions of T4, which are non-genomic actions. In contrast with genomic actions that are mediated by TRs and appear within several hours or more, by with gene transcriptional activation or suppression, non-genomic actions are initiated at the membrane receptor on integrin αvβ3, other sites on the membrane, or in cytoplasm, and usually appear within minutes. T4 acts on the inward flux of Na+, actin polymerization, and Ca2+-ATP activity, in specific cells or tissues.

Gastrointestinal hormones are peptides released from endocrine cells and neurons in the digestive tract. More than 30 hormone genes are currently known to be expressed in the gastrointestinal tract, which makes the gut the largest hormone-producing organ in the body. Modern biology makes it feasible to conceive the hormones under five headings. (1) The structural homology groups a majority of the hormones into nine families, each of which is assumed to originate from one ancestral gene. (2) The individual hormone gene often has multiple phenotypes due to alternative splicing, tandem organization, or differentiated maturation of the prohormone. By a combination of these mechanisms, more than 100 different hormonally active peptides are released from the gut. (3) Gut hormone genes are also widely expressed outside the gut, some only in extraintestinal endocrine cells and neurons but others also in other cell types. (4) The extraintestinal cells may express different bioactive fragments of the same prohormone due to cell-specific processing pathways. (5) Finally, endocrine cells, neurons, and, for instance, spermatozoa display differential release of gut peptides, so the same peptide may act as a blood-borne hormone, a neurotransmitter, a long-acting growth factor, or a fertility factor.

The pyrokinin/pheromone biosynthesis-activating neuropeptide (PK/PBAN) family is one of the most studied neuropeptide (Np) families in insects and is one of the most important in insect physiology regulation. Physiological functions regulated by the PK/PBAN peptides include development, mating, muscle contraction, and tanning. Members of the family are found in many Lepidopteran and other insect species. In this chapter, we briefly present a historical perspective of the discovery of the PK/PBAN peptides, provide details on the structure of the PK/PBAN genes and the processing of their prohormone to bioactive peptides, describe the distribution of the mRNA and peptides in the insect nervous system, and summarize the current knowledge on the PK/PBAN receptors, their signaling mechanisms, and their biological activity. Employment of the PK/PBAN family of peptides as a basis for designing a novel generation of insect-control agents based on Np antagonists is also briefly discussed.

Recent studies reported that many different biochemical forms of B-type-related peptides circulate in human blood. In particular, a significant amount of the prohormone peptide (i.e., proBNP108) can be detected in plasma of patients with heart failure. These data indicate that the posttranslational maturation processing of the B-type natriuretic peptide (BNP) precursor may not be efficient in heart failure. The aim of this chapter is to describe the biochemical pathways of proBNP108 maturation and to discuss the pathophysiological relevance of alteration of the posttranslational maturation mechanisms in heart failure. An impaired cardiac endocrine function was proposed to explain the altered electrolyte and fluid homeostasis occurring in chronic heart failure. Recent studies demonstrated that a great part of BNPs assayed by immunoassay methods in healthy subjects and in patients with cardiovascular disease is devoid of biological activity. These findings suggest that an alteration in posttranslational maturation of BNP precursor may promote the resistance to biological action of BNP in patients with heart failure at a prereceptor level. These studies also open a new and more complex scenario regarding the circulating BNPs. The active hormone (i.e., BNP1-32) may be produced even in vivo from the circulating precursor proBNP108 by plasma enzyme degradation, such as the soluble form of corin, possibly able to process the circulating intact precursor of natriuretic hormones. As a future perspective, the simultaneous measurement of the proBNP1-108 and the active peptide BNP1-32 with more specific methods could allow a more accurate estimation of both production/secretion of B-type-related peptides from cardiomyocytes and the true activity of the cardiac endocrine function.

Knowledge of the mechanisms of prohormone processing has been growing steadily over the last decade. Studies with knockout mice have defined the individual steps of the pathways and have emphasized the physiological importance of the process. Although the phenotypes caused by deficiency of the processing enzymes have traditionally been ascribed to lack of product, they may also be caused by an oversupply of the precursor, since evidence is now accumulating that the precursors are biologically active through different receptors from the mature hormones. The processing pathways leading to the gastrointestinal hormones gastrin-releasing peptide, gastrin, and Sonic Hedgehog are presented as examples. The crystal structures of many of the processing enzymes have now been determined, their catalytic mechanisms defined, and selective inhibitors identified. Further improvements in inhibitor potency and specificity may lead to novel therapies for diseases as diverse as obesity and cancer.

Background This study compared NT-proBNP levels and the association with cardiovascular markers between Africans and Caucasians from South Africa. Methods This cross-sectional study involved 201 Africans and 255 Caucasians from the North West province, South Africa. Serum NT-proBNP concentrations, blood pressure, pulse wave velocity and arterial compliance were measured. Results NT-proBNP levels were significantly higher (P < 0.001) in Africans than Caucasians, also after adjusting for gender, body mass index (BMI) and pulse wave velocity (P = 0.008). This significant difference became borderline significant after adjusting for systolic blood pressure (SBP) (P = 0.060), and non-significant after adjusting for arterial compliance (P = 0.35). In single regression, a significant positive correlation of NT-proBNP with SBP (r = 0.26; P < 0.001) and pulse pressure (PP) (r = 0.28; P < 0.001) were shown for Africans only. After multiple adjustments, the associations of NT-proBNP with SBP and PP remained significant in Africans (SBP: β = 0.187, P < 0.01; PP: β = 0.234, P < 0.001), with no significant associations in Caucasians. Conclusions NT-proBNP levels were higher in Africans than Caucasians, independently of BMI and gender. This difference was partly driven by higher SBP and lower arterial compliance in Africans. NT-proBNP was persistently associated with SBP and PP in Africans, but not in Caucasians. These associations may suggest early vascular changes contributing to cardiac alterations in Africans. http://dx.doi.org/10.1016/j.hlc.2011.10.009

Publisher Summary The mechanisms and outcome of cerebral ischemic injury are influenced strongly by biological sex and the availability of female and male sex steroids to the brain. This chapter evaluates the data suggesting that biological sex determines clinical and experimental stroke outcomes, reviews special considerations for women with stroke, summarizes the controversy behind estrogen's antiischemic and neuroprotective properties, andintroduces the novel idea that androgens contribute to ischemic sensitivity in the male brain. The epidemiology of ischemic stroke is sexually dimorphic in that ischemic events occur with greater frequency in men than in women regardless of country of origin and ethnic culture. The potential for sex-specific responses to treatments or the possibility that endogenous or exogenous sex steroids may interact with treatments is observed. The pleiotropic estrogens are linked to the early protection from stroke in women. The mechanism attenuating the beneficial effects of estrogen is loss of arterial estrogen receptors. Loss of the estrogen receptors occurs with aging and is more prominent in atherosclerotic vessels. Male sex is an acknowledged risk factor for human stroke. Testosterone acts as a type of prohormone with two distinct conversion pathways. In all androgen-sensitive tissues, the steroid is converted intracellularly to either the more potent androgen dihydrotestosterone or to E2. It is hypothesized that androgens enhance ischemic damage and may play a role in male stroke.

Since its discovery in 1999, the stomach-derived hormone ghrelin has been studied intensively. Proghrelin is 94 amino acids long in mammals and this undergoes proteolytic processing to produce ghrelin [residues 1-28 of proghrelin(1-94)] and the C-terminal peptide C-ghrelin, which likely contains the entire 66 amino acids of the prohormone C-terminus. The accumulating data identifies ghrelin as having important roles in growth hormone (GH) release, appetite, metabolism, energy balance, cardiovascular function, reproduction, and bone growth. The most striking feature of ghrelin is that it can be acylated at its third amino acid residue (usually Ser), usually in the form of n-octanoyl group (C8:0). Approximately 10-20% of circulating ghrelin is acylated and this feature confers its GH releasing ability, mediated by the GH secretagogue receptor (GHSR). In contrast, the remaining 80-90% of circulating ghrelin is desacylated. Desacyl ghrelin was initially thought to be inactive, but recent in vivo and in vitro evidences have identified biological actions for this peptide, independent of GHSR. Whether C-ghrelin has bioactivity remains to be determined, but it is known that plasma concentrations of this peptide respond to endocrine and metabolic manipulations in the same fashion as ghrelin itself. A third putative proghrelin peptide, termed "obestatin" has been mooted, but confirmatory biochemical and functional evidences supporting the existence of this peptide have not been forthcoming, suggesting it to be a biochemical miscalculation. This chapter will address biochemical aspects of proghrelin peptides and point to potential avenues for future work.

The natriuretic peptide system includes atrial (ANP) and brain, or B-type (BNP), natriuretic peptide. Brain natriuretic peptide is a 32 amino acid cardiac natriuretic peptide, which was originally isolated from porcine brain tissue. The human BNP gene is located in chromosome 1 and encodes the 108 amino acid prohormone pro-BNP, which circulates with the 32 amino acid biologically active BNP and the N terminal 76 amino acid component of the prohormone (NT-proBNP). These three components can be measured by immunoassay. BNP is released by cardiac myocytes of the ventricles in response to myocardial wall stress. This is brought on by increased transmural wall tension and elevations of end diastolic pressure, ventricular volume expansion and pressure overload. The secretion of this important peptide serves to regulate sodium and water balance by the kidneys and causes vasodilatation of arteries, which benefits the failing heart. Elevated BNP levels are detected in the blood of patients with heart failure, but BNP levels are normal in patients with shortness of breath caused by pulmonary disease. The rapid measurement of levels of BNP in the blood has proved useful in the diagnosis of heart failure. More importantly, in emergency rooms, the BNP test rapidly clarifies whether severe shortness of breath is caused by heart failure or by pulmonary disease.

We have established that decentralization (cutting sympathetic nerve trunk) of the superior cervical ganglia bilaterally reduces the magnitude of allergic inflammation in the airways of rats. The magnitude of anaphylactic and endotoxic hypotension, and of gastrointestinal inflammation was also reduced. This anti-inflammatory activity was dependent upon intact submandibular glands. Reconstitution of sialadenectomized (removal of submandibular glands) rats with soluble extracts of the submandibular glands identified two polypeptides with anti-inflammatory activities. The sequences of these polypeptides were found within a prohormone, submandibular gland rat 1 (SMR1). The C-terminal peptide TDIFEGG, has been studied extensively. Using sequential amino acid substitutions and systematic removal of C terminal or N terminal amino acids, we established that the tripeptide FEG is biologically active. Modification of FEG to the D-isomeric feG, enhances its activity in some assay systems. We postulated that feG would inhibit airways inflammation, and tested this using a model ofallergic asthma, namely the Brown Norway rat sensitized to ovalbumin (OA). Sensitized rats were challenged 14 to 21 days later with aerosolized OA. This challenge markedly increased numbers of inflammatory cells recovered from the airways after 24 hour (29×106, n=23) compared to saline controls (1×106, n=4). The infiltrating cells included macrophages (10×106), neutrophils (9×106) and eosinophils (9×106). Intravenous (0.25 mg/kg) or oral feG (1 mg/kg) given 30 min prior to OA significantly inhibited influx of inflammatory cells by 50 to 70%. feG reduced inflammatory cell infiltration when given 30 min before to 3 to 6 hour post allergen exposure. Oral feG reduced the numbers of macrophages, neutrophils and eosinophils. One of the mechanisms underlying the effects of feG may be its ability to inhibit PAF-induced expression of CD11b on purified human neutrophils. It is possible that feG may be useful in the treatment of allergic asthma, given either as an oral prophylactic, or as a post exposure therapeutic. The cervical sympathetic nerve trunk-submandibular gland axis of neuroendocrine regulation of inflammation may be dysfunctional in inflammatory diseases and provide opportunities for new therapeutic intervention. This axis may be sensitive to modulation by central and peripheral neural mechanisms that influence its function/dysfunction.

Publisher Summary Studies using antisense inhibition have been criticized because of the possible nonspecific cellular effects complicating interpretation of the results. In light of these concerns, several controls have been performed to ensure that the effects of antisense expression on the processing of pro-cholecystokinin (CCK) are specific. The two most important controls performed were to measure the effects of antisense expression on target messenger ribonucleic acid (mRNA) and target protein. Another control that was performed was to measure the effect of polycystin 2 (PC2) message and protein level in antisense PC1-transfected cells and vice versa. Although PC1 and PC2 are homologous genes within the same family of endoproteases, the effect of antisense is specific for each. These observations suggest that the effect of antisense on the RNA level is complex but specific and may occur at multiple levels to produce inhibition of the protein product. Knowledge of the actual enzymes responsible for prohormone cleavages in specific tissues lags behind progress in other areas of the peptide hormone field owing to the technical difficulty of identification, isolation, characterization, and inhibition of these enzymes. As important as the PC enzymes appear to be, it is possible that proteases other than the PCs are involved in prohormone processing.

Publisher Summary The functional unit of the thyroid is the follicle, a cystlike structure lined by a single-layer epithelium, the follicular epithelium, enclosing a central cavity: the colloid. The latter one represents a special extracellular compartment that serves as a reservoir for storage of iodinated thyroglobulin—that is, a prohormone from which thyroid hormones are released on appropriate stimulus. Thyroglobulin is synthesized within the endoplasmic reticulum of the follicular cells (or thyrocytes) and is secreted into the colloid. Iodide reaches this compartment separately where it is eventually coupled to thyroglobulin. This chapter reviews the evidence for the existence of an iodide channel in the apical membrane of the thyrocyte.

The renin–angiotensin system (RAS) is an extracellular hormonal system implicated in acute, homeostatic control of peripheral vascular resistance and electrolyte homeostasis. In this tightly regulated system, physiological regulators of blood pressure and fluid balance induce the production of the potent vasoactive angiotensin peptides by sequential proteolysis of the angiotensinogen (AGT) prohormone. AGT is the only known precursor of the angiotensin peptides, whose circulating concentrations influence the tonic activity of the RAS. AGT abundance is regulated at the transcriptional level through hormonal and cell-type specific regulators. In this review, we will discuss the identified mechanisms controlling AGT expression separately for the rodent and human genes. The most intensively investigated gene (rodent AGT) is regulated constitutively by multiple positive- and negative-acting cis factors that function in a cell-type dependent fashion. Inducible rodent AGT expression is mediated through a multihormone-inducible enhancer that integrates signals from steroid and cytokine hormones into AGT transcription. We review recent advances in understanding the mechanism of the nuclear factor-κB (NF-κB) family in mediating cytokine-induced AGT expression and our recent discoveries on the existence of differentially inducible pools of cytoplasmic NF-κB. Constitutive control of the human AGT gene will be discussed; there is surprisingly little information on the cis - and trans -acting regulators controlling inducible expression of human AGT. Finally, we will explore some of the recent developments in gene linkage studies where human AGT alleles have been associated with hypertensive phenotypes through a mechanism that may involve enhanced transcription. These studies have provided a molecular explanation for a subset of heritable hypertensive disorders in humans.

A number of peptides have been identified in the central nervous system of the freshwater snail, Lymnaea stagnalis, that function as hormones and neurotransmitters/neuromodulators. These peptides are typically proteolytically processed from larger prohormones mostly at sites composed of single or multiple basic amino acid residues. Previously we demonstrated a diversity of putative prohormone convertases that may be involved in prohormone processing in the Lymnaea brain. In the present report, we have characterized a cDNA clone encoding a putative endoprotease of 837 amino acids. The primary structure of the endoprotease (Lfur2) was comparable to that of human furin and contained a putative catalytic domain, a Cys-rich domain, and a transmembrane region. The catalytic domain of Lfur2 demonstrated about 70% residue identity when compared with human furin, PACE4 and Drosophila Dfur1 and dKLIP-1. The Lfur2 gene was expressed in the central nervous system as well as various peripheral tissues of Lymnaea.

Publisher Summary Prohormone structure governs proteolytic processing and sorting in the Golgi complex. In the trans-Golgi network (TGN), neuropeptides are segregated from other secretory proteins and are packaged into dense-core vesicles (DCVs). These vesicles are then transported to the nerve terminal and their contents are released by exocytosis when the appropriate signal is received by the neuron. Processing begins with the removal of the signal sequence as the nascent peptide enters the endoplasmic reticulum . Hence, elucidating the molecular and cellular mechanisms governing these processing events is the key to fully understanding the regulation of neuropeptide biosynthesis. This chapter discusses (1) the structural features of the prohormone important for processing, (2) the processing enzymes, and (3) how the processing sequence of the egg-laying hormone (ELH) prohormone influences the sorting of peptides derived from this molecule. Using the ELH prohormone to study the relationship between processing and sorting has opened up new avenues of thought. In the past, it was learned that cellular mechanisms can control the relative levels and spatial localization of different neuropeptides derived from a common precursor. However, recent results indicate that perturbation of a processing site can cause processing in a different locale in the cell and result in altered pathways of sorting. Defining the hierarchy of processing enzymes and their localization will be crucial to future work in this area and to understand how the design of prohormones complements the repertoire of processing enzymes.

Publisher Summary The members of the vasopressin/oxytocin hormone superfamily are widely distributed throughout the animal kingdom, and all hormones belonging to this superfamily are structurally related nonapeptides, having five invariant amino acid residues, including two cysteines, that form a disulphide bridge and an amidated carboxyterminal glycine. This chapter discusses the presence of vasopressin- and oxytocin-related peptides in invertebrates as well as their functions and explores the structural characteristics of the Lymnaea conopressin precursor in an evolutionary perspective. Only four different vasopressin- and oxytocin-related peptides have been structurally identified in invertebrate species, including Lysconopressin from the gastropod molluscs Conus geographus and Lymnaea stagnalis , Arg-conopressin from the gastropod mollusc Conus striatus , the vasopressin-like diuretic hormone from the insect Locusta migratoria , and cephalotocin from the cephalopod mollusc Octopus vufgaris . The results presented in the chapter show for the first time that in invertebrates a vasopressin-related prohormone is present, which has the same organization as its vertebrate counterparts.

Publisher Summary This chapter describes the studies conducted for atrial natriuretic factor (ANF) receptor assays. Cardiac atria are believed to play an important role in the regulation of extracellular fluid volume and electrolyte homeostasis. Distension of the atrial walls or an increase in intra-atrial pressure induces water and salt excretion. The ANF is synthesized in atrial cardiocytes and other tissues as a 126 amino acid prohormone. Its biological activity resides on the C-terminal part of the molecule, the 28 amino acid C-terminal peptide Ser99–Tyr126, which is released into the circulation. All the actions of ANF can be explained by its binding to specific receptors identified on plasma membranes of cells in a great variety of target tissues. Modern biochemical techniques and molecular biology have been used to identify, characterize, and clone ANF receptors with respect to molecular weight, composition, and regulation. Antireceptor antibodies could be a powerful tool in physiological studies to block receptors. Using molecular biology reagents, such as recombinant DNA probes, the number of genes that encode ANF receptors could be determined.

Publisher Summary A biosynthetic precursor, proparathyroid hormone (proPTH), to parathyroid hormone (PTH) has been identified recently in bovine human, and, tentatively, in chick and rat parathyroid tissue. The prohormone from the bovine contains an additional hexapeptide sequence at the amino terminus of PTH. Four of the six additional amino acids are positively charged (1 arginine and 3 lysine residues), providing a ready means for the physical separation of the prohormone from PTH by gel electrophoresis or ion-exchange chromatography. The initial identification of the precursor was accomplished by electrophoretic analysis of the radioactive tissue proteins synthesized during short incubations of the tissue in vitro with radioactive amino acids (pulse labeling). This chapter describes the techniques that have been used to identify the prohormone in bovine parathyroid glands. Similar techniques have been used for the identification of biosynthetic precursors for other polypeptide hormones, for example, insulin and glueagon and for identification of precursors from human, chick, and rat parathyroids. The procedures described here include methods (1) for location of parathyroid glands in the bovine, (2) for demonstration of the precursor to product relationship between proPTH and PTH by in vitro pulse labeling of the parathyroid proteins, and (3) for analyses of the chemical properties of PTH and proPTH by use of selective labeling with radioactive amino acids followed by limited cleavage and eleetrophoretie separation of the labeled peptides.

Publisher Summary This chapter discusses the combined antibody-high-performance liquid chromatography approach to assess prohormone processing. This approach creates special problems for the protein biochemist and physiologist because the amino acid sequences of the various peptides may not be known with certainty (the peptide has not been sequenced, only deduced from a complementary DNA (cDNA)), the regulatory steps involved in the biosynthesis, processing, and secretion of the peptides are unclear, and their functional roles are often unexplored. Various tissues may metabolize the same precursor peptides by different routes, and these products may have different functional consequences depending on the locus of secretion and the physiological/pathological status of the organism. These circumstances emphasize the importance of adopting a coordinated and multifaceted approach in detailing a peptide's fate from biosynthesis to function. Perhaps, through additional biochemical and physiological manipulations, we will be able to dissect more clearly the contribution of different gonadal factors to the biosynthesis, processing, transport, secretion, and/or degradation of these various processed products from luteinizing hormone releasing hormone (LHRH) neurons.

Publisher Summary This chapter presents the scheme by which peptide hormones are synthesized. The mRNA is transcribed in the nucleus of the cell. There is evidence that some mRNA's may be posttranscriptionally modified, that is, that pro-mRNA's may be made that in turn give rise to smaller mRNA's. The mRNA's are exported from the nucleus to the cytoplasm. There they are translated by ribosomes. The translation product is the preprohormone, the N-terminus of which is the signal or leader sequence. This amino acid sequence is typically quite hydrophobic and it passes into and through the endoplasmic membrane. It is then removed from the nascent peptide chain by a “signalase,” yielding the prohormone. Inside the endoplasmic space, the prohormone is processed into biologically active smaller molecules. This processing involves proteolysis and, in some cases, addition of functional groups. A number of strategies have been employed for characterizing preprohormones and prohormones. The earliest approach involved pulse labeling prohormones with radiolabeled amino acids and then isolating them.

Human β-lipotropin (β h -LPH) consists of 89 amino acids with glutamic acid as the NH 2 - and COOH-terminal residue and its primary structure has been proposed. In order to securely establish the primary structure of β h -LPH, the hormone has been synthesized by the new segment-coupling method in aqueous solution. The synthetic product was shown to be identical to native β h -LPH by paper electrophoresis, isoelectric focusing, high performance liquid chromatography, lipolytic activity in isolated rabbit fat cells and radio-immunoassay. Since β-lipotropin is the prohormone for β-endorphin, some recent studies on β-endorphin are discussed.

Peptides related to β-endorphin (residues 1–31)were extracted from rat pituitary and brain, resolved by chromatography and determined by RIA. In the pituitary, the biologically potent form of β-endorphin was the major immunoreactive peptide in the anterior region while the inactive acetylated forms of β-endorphin and C′-fragment (residues 1–27 and 1–26) were the major peptides of the pars intermedia. In the brain, the biologically potent form of β-endorphin was present in the hypothalamus as the major immunoreactive peptide; in the amygdala and mid brain it was accompanied by des histidine C′-fragment (β-endorphin 1–26). In the hippocampus, colliculae and brain stem, N-acetylated forms of the β-endorphin related peptides, which are inert as opiates, predominated. The results demonstrate the existence in these tissues of two distinct mechanisms for processing of the C-terminal region of the ACTH-endorphin prohormone.

Publisher Summary Several years of search for a putative factor in brain, which activates opiate receptors, has culminated in the isolation of two pentapeptides, Met-enkephalin [Ia] and Leu-enkephalin [Ib] from guinea pig brain. Both have been synthesized and showed typical opioid activity in vitro and analgesic effects in vivo . This chapter reviews Peptide hormones that are of other than hypothalamic or pituitary origin. The structures of seven species-different calcitonins have been determined. Conventional syntheses of porcine, human and salmon calcitonin provided large supplies of homogeneous peptides for physiological, pharmacological, and clinical investigations. Salmon calcitonin is far more potent in man than human calcitonin that may be attributable to increased metabolic stability. Secretion of calcitonin from porcine thyroid glands is effected by activation of C-cell adenyl cyclase by either ionized Ca or by hormones (glucagon, gastrin, CCK, epinephrine). Calcitonin effects are age-dependent and are very low in normal adult humans. 6-Lipotropin is thus another example of a second order prohormone, that is, a well-defined peptide hormone serving as a precursor for a smaller peptide with entirely different hormonal activities. The characterization of the opioid peptides may suggest new approaches to the development of nonaddictive analgesics.

Angiotensin II (AII) receptors in the central nervous system mediate three pressor responses–enhanced sympathetic outflow, release of ADH and dipsogenesis. We defined a topographical distribution of the prohormone of angiotensin, angiotensinogen, in the CNS and find a positive correlation with published norepinephrine (NE) concentrations. Direct, simultaneous measurements of angiotensinogen and NE in various brain sections support this correlation. We interpret these results as indicating that AII may control NE metabolism.

Growth hormone (GH) should be prepared from frozen pituitary glands. Organic solvents denature the GH molecule, and make it less readily extractable, aggregats are formed, and the activity is reduced. Aggregated human (h) GH produce hGH antibodies, and may give therapy resistance. The hGH molecule is a single chain polypeptide with 191 amino acids with two intrachain disulfide bridges. Removal of the fragment 135–146 by limited plasmin digestion increased the biologic activity to a great extent. The native pituitary GH may be a prohormone, and the sequence 95–134 has been considered to be an eventual somatotropic core. The stimulation of longitudinal growth is probably mediated by production of new growth factors, whereas synthesis of tissue proteins is a direct anabolic effect. Additionally GH has a mammotropic and lymphotropic effect, and influences carbohydrate and lipid metabolism. The secretion of GH is regulated by hypophysiotropic hormones synthesized in hypothalamic nuclei and transported to the pituitary gland through portal capillaries. The release of GH inhibitory hormone and an eventual GH releasing hormone is controlled by aminergic neurons and glucoreceptors.

Publisher Summary This chapter discusses the chemistry of growth hormone and lactogenic hormone. The various human and animal growth hormones and lactogens differ to a greater or lesser extent from one another in amino acid sequence but share common biological effects and functions. Hence, comparisons of restricted or extended sequences of these molecules are frequently made to assess the degree of homology, with the ultimate aim of identifying the structural features important for biological activity. Human growth hormone preparations subjected to limited plasmin digestion retain full activity and in some assays have actually shown increased growth-promoting activity. Moreover, it is possible that the growth hormone molecule as stored in the pituitary and released into the circulation may be a precursor or prohormone that is converted peripherally into a modified form or a smaller fragment. Comparisons of amino acid sequence have shed some light on the evolution of the growth hormones and lactogens. Thus, the similarity among the sequences of growth hormones, prolactins, and placental lactogen make it almost certain that all three of these molecules have evolved from a common primordial precursor. The growth hormones and lactogenic hormones are single-chain polypeptides with 2 or 3 intrachain disulfide bridges and about 200 residues. They lack carbohydrate, in contrast to the gonadotropins and thyrotropins that have multiple sites of carbohydrate attachment in a two-chain structure.

Publisher Summary This chapter examines cholesterol metabolism in terms of the molecular events that are responsible for constructing cyclopentanoperhydrophenanthrene derivatives from small molecular precursors. There is good reason for centering a discussion of steroid biogenesis around cholesterol, not only because this substance has received the major share of attention but also because cholesterol synthesis appears to be an integral part of the processes leading to the physiologically active steroids, namely, the bile acids, and the adrenocortical and sex hormones. It seems further than the syntheses of cholesterol, of ergosterol, and of the sterols in higher plants follow the same general plan and branch from each other only a t a relatively late stage. It is largely a matter of definition whether a discussion of the present subject properly belongs to “Vitamins and Hormones.” As a major, and probably obligatory, precursor of progesterone and, by way of progesterone also of the androgens, the estrogens, and the corticoids, cholesterol qualifies at least as a prohormone. Secondly, since cholesterol is known to be obligatory for the growth of representatives of at least two animal phyla, protozoa and insects, it is technically correct to regard cholesterol as a vitamin. It seems clear that the traditional classification of cell constituents on the basis of nutritional or functional parameters is far too narrow, at least for the case of cholesterol.