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Paul A Kelly

Publications and source records attributed to Paul A Kelly.

At least 19 recordsLinked to original sources
PubMed2006

A new mechanism for prolactin processing into 16K PRL by secreted cathepsin D.

Cathepsins are lysosomal enzymes that were shown to release the antiangiogenic fragments 16K prolactin (PRL), endostatin, and angiostatin by processing precursors at acidic pH in vitro. However, the physiological relevance of these findings is questionable because the neutral pH of physiological fluids is not compatible with the acidic conditions required for the proteolytic activity of these enzymes. Here we show that cathepsin D secreted from various tissues is able to process PRL into 16K PRL outside the cell. To specifically target extracellular proteolysis, we used tissues from PRL receptor-deficient mice, which are unable to internalize PRL. As assessed by the use of specific inhibitors of proton extruders, we show that the proteolytic activity of cathepsin D requires local acid secretion driven by Na(+)/H(+) exchangers and H(+)/ATPase. Although it is usually assumed that cathepsin-mediated generation of antiangiogenic peptides occurs in the moderately acidic pericellular milieu found in malignant tumors, we propose a new mechanism explaining the extracellular activity of this acidic protease under physiological pH. Our data support the concept that secreted lysosomal enzymes could be involved in the maintenance of angiogenesis dormancy via the generation of active antiangiogenic peptides in nonpathological contexts.

Animals↗
PubMed2006

Growth hormone promotes skeletal muscle cell fusion independent of insulin-like growth factor 1 up-regulation.

Growth hormone (GH) participates in the postnatal regulation of skeletal muscle growth, although the mechanism of action is unclear. Here we show that the mass of skeletal muscles lacking GH receptors is reduced because of a decrease in myofiber size with normal myofiber number. GH signaling controls the size of the differentiated myotubes in a cell-autonomous manner while having no effect on size, proliferation, and differentiation of the myoblast precursor cells. The GH hypertrophic action leads to an increased myonuclear number, indicating that GH facilitates fusion of myoblasts with nascent myotubes. NFATc2, a transcription factor regulating this phase of fusion, is required for GH action because GH is unable to induce hypertrophy of NFATc2-/- myotubes. Finally, we provide three lines of evidence suggesting that GH facilitates cell fusion independent of insulin-like growth factor 1 (IGF-1) up-regulation. First, GH does not regulate IGF-1 expression in myotubes; second, GH action is not mediated by a secreted factor in conditioned medium; third, GH and IGF-1 hypertrophic effects are additive and rely on different signaling pathways. Taken together, these data unravel a specific function of GH in the control of cell fusion, an essential process for muscle growth.

Animals↗
PubMed2006

Drug Insight: prolactin-receptor antagonists, a novel approach to treatment of unresolved systemic and local hyperprolactinemia?

Prolactin is a polypeptide hormone whose major biological actions are related to normal lactation and reproduction. Abnormally high prolactin levels, referred to as hyperprolactinemia, can result in various reproductive disorders. Currently, therapeutic management of hyperprolactinemia relies on dopamine agonists, since dopamine is the primary physiological suppressor of pituitary prolactin production. Epidemiologic studies have shown that prolactin levels in the high-normal range, as well as medications that interfere with dopamine action (e.g. certain antipsychotic drugs), might correlate with increased breast cancer risk. In addition to circulating prolactin, it is now well established that prolactin is also produced locally within various tissues, including breast and prostate. Increasing evidence, mainly from animal studies at present, suggests that excess locally produced prolactin may promote the growth of breast and prostate tumors via an autocrine or paracrine mechanism. These findings have renewed the interest in finding alternative strategies to suppress prolactin actions when dopamine agonists are ineffective. Our studies of the relationship between prolactin structure and function have resulted in the development of pure prolactin-receptor antagonists. These molecules prevent endogenous prolactin from exerting its actions via a competitive mechanism for receptor binding. In this review, we discuss the possible future therapeutic utility of this novel class of compounds.

Animals↗
PubMed2006

Reduced recruitment and survival of primordial and growing follicles in GH receptor-deficient mice.

GH influences female fertility. The goal of the present study was to obtain more insight into the effect of loss of GH signalling, as observed in humans suffering from Laron syndrome, on ovarian function. Therefore, serial paraffin sections of ovaries of untreated and IGF-I-treated female GH receptor knock-out (GHR/GHBP-KO) mice were examined to determine the follicular reserve and the percentage of follicular atresia in each ovary. Our observations demonstrate that the amount of primordial follicles was significantly elevated in GHR/GHBP-KO mice, while the numbers of primary, preantral and antral follicles were lower compared with wild-type values. The reduced number of healthy growing follicles in GHR/GHBP-KO mice was accompanied by a significant increase in the percentage of atretic follicles. IGF-I treatment of GHR/GHBP-KO mice for 14 days resulted in a reduced number of primordial follicles, an increased number of healthy antral follicles, and a decreased percentage of atretic follicles. The results of the present study suggest that GH may play a role, either directly or indirectly, via for instance IGF-I, in the recruitment of primordial follicles into the growing pool. Furthermore, GH seems to protect antral follicles, directly or indirectly from undergoing atresia.

Animals↗
PubMed2006

Local over-expression of prolactin in differentiating mouse mammary gland induces functional defects and benign lesions, but no carcinoma.

Experimental, clinical, and epidemiological data support the growth-promoting role of endocrine prolactin (PRL) in mammary tumors. PRL is also produced by the breast, where it is now recognized to act as a growth/survival factor via autocrine/paracrine mechanisms. Recent transgenic (Tg) mouse models have revealed the pro-oncogenic effect of PRL over-expression in virgin mammary glands. To address the question whether PRL tumorigenicity was maintained on differentiated mammary glands, we generated mammary-specific Tg mice expressing human (h)PRL under the control of the milk whey acidic protein promoter, which directs autocrine hPRL over-expression in late gestation throughout lactation. Minimal levels of transgene expression were detected in the mammary glands of virgin animals, which at best induced partial ductal branching and lobulo-alveolar structures in older nulliparous females. As expected, expression of mammary hPRL dramatically increased at the end of first pregnancy, and from this point it never returned to baseline, although it peaked at each gestation/lactation cycle. Over-expression of hPRL that starts when the gland is already well into the differentiation process led to various morphological mammary alterations, including abnormally differentiated epithelium, atropy of the myoepithelial layer, dilated ducts, cysts, and lymphocytic infiltrates. These phenotypes tended to worsen with successive pregnancies, also reflecting cumulative damage of failure of involution. Although some older, multiparous females developed benign tumors (papillomas and metaplasias), none of the animals studied developed mammary carcinomas. In addition, we noticed that half of the Tg females exhibited lactation defects, leading to significantly increased pup mortality. This phenotype was due neither to failure of milk production nor to modification of its protein content, but rather it was correlated to lipid enrichment of the milk, which, in combination with profoundly altered morphology of the gland, led to impaired milk extrusion through the nipple. In summary, these data show that over-expression of autocrine hPRL in a differentiating mammary gland induces dramatic functional and morphological defects, but not carcinoma. This deserves further investigations on the emerging concept that autocrine PRL may have different effects on pathological development of the mammary gland depending on the differentiation state of the latter.

Animals↗
PubMed2005

Medullary thyroid carcinoma arises in the absence of prolactin signaling.

Prolactin, a pituitary hormone, exerts pleiotropic effects in various cells. These effects are mediated by a membrane receptor highly expressed in many tissues. To analyze prolactin effects on the thyroid gland, we first identified prolactin receptor (PRLR) mRNAs by in situ hybridization. To further evaluate the physiologic relevance of PRLR actions in the thyroid in vivo, we used PRLR knockout mice. Whereas the histologic structure of thyroid of PRLR-null mice was not disturbed, we show that T4 levels are lower in null animals (13.63 +/- 2.98 versus 10.78 +/- 2.25 pmol/L in null mice), confirming that prolactin participates in the control of thyroid metabolism. To further investigate thyroid effects in mice, we measured body temperature and thyroid-stimulating hormone in young and adult male and/or female PRLR-null mice and their normal siblings. Surprisingly, in null animals, we saw medullary thyroid carcinoma (MTC) arising from parafollicular C cells producing calcitonin. The incidence of these carcinomas attained 41% in PRLR-null mice, whereas this malignant tumor occurs sporadically or as a component of the familial cancer syndrome in humans. This finding suggests that PRLR-null mice could represent a valuable animal model for MTC, which could be compared with existing MTC models. These observations suggest a possible link between the appearance of this carcinoma and the absence of prolactin signaling.

Animals↗
PubMed2005

Prolactin and growth hormone regulate adiponectin secretion and receptor expression in adipose tissue.

Adiponectin is a hormone secreted from adipose tissue, and serum levels are decreased with obesity and insulin resistance. Because prolactin (PRL) and growth hormone (GH) can affect insulin sensitivity, we investigated the effects of these hormones on the regulation of adiponectin in human adipose tissue in vitro and in rodents in vivo. Adiponectin secretion was significantly suppressed by PRL and GH in in vitro cultured human adipose tissue. Furthermore, PRL increased adiponectin receptor 1 (AdipoR1) mRNA expression and GH decreased AdipoR2 expression in the cultured human adipose tissue. In transgenic mice expressing GH, and female mice expressing PRL, serum levels of adiponectin were decreased. In contrast, GH receptor deficient mice had elevated adiponectin levels, while PRL receptor deficient mice were unaffected. In conclusion, we demonstrate gene expression of AdipoR1 and AdipoR2 in human adipose tissue for the first time, and show that these are differentially regulated by PRL and GH. Both PRL and GH reduced adiponectin secretion in human adipose tissue in vitro and in mice in vivo. Decreased serum adiponectin levels have been associated with insulin resistance, and our data in human tissue and in transgenic mice suggest a role for adiponectin in PRL and GH induced insulin resistance.

Adult↗
PubMed2005

DYRK1A enhances the mitogen-activated protein kinase cascade in PC12 cells by forming a complex with Ras, B-Raf, and MEK1.

Dual-specificity tyrosine-phosphorylated and regulated kinase 1A (Dyrk1A) is the human homologue of the Drosophila mnb (minibrain) gene. In Drosophila, mnb is involved in postembryonic neurogenesis. In human, DYRK1A maps within the Down syndrome critical region of chromosome 21 and is overexpressed in Down syndrome embryonic brain. Despite its potential involvement in the neurobiological alterations observed in Down syndrome patients, the biological functions of the serine/threonine kinase DYRK1A have not been identified yet. Here, we report that DYRK1A overexpression potentiates nerve growth factor (NGF)-mediated PC12 neuronal differentiation by up-regulating the Ras/MAP kinase signaling pathway independently of its kinase activity. Furthermore, we show that DYRK1A prolongs the kinetics of ERK activation by interacting with Ras, B-Raf, and MEK1 to facilitate the formation of a Ras/B-Raf/MEK1 multiprotein complex. These data indicate that DYRK1A may play a critical role in Ras-dependent transducing signals that are required for promoting or maintaining neuronal differentiation and suggest that overexpression of DYRK1A may contribute to the neurological abnormalities observed in Down syndrome patients.

Animals↗
PubMed2005

Zero-length cross-linking reveals that tight interactions between the extracellular and transmembrane domains of the luteinizing hormone receptor persist during receptor activation.

Several molecular models of glycoprotein hormone receptor activation have been proposed. It has been suggested that ligand binding to the ectodomain (ECD) leads to major changes in intramolecular interactions between the ECD and the transmembrane domain. We studied these intramolecular modifications by generating a recombinant LH/CG receptor (LHR) bearing an intramolecular cleavage site. We did this by inserting a furin site at position 316 in the hinge region of the ECD (LHR_Fur316). Affinity for human chorionic gonadotropin (hCG) and cAMP production upon hCG stimulation was identical to those of wild-type LHR. Western blot analysis showed that the LHR_Fur316 receptor was cleaved into two subunits linked by disulfide bridges. Chemical shedding of the ECD from the transmembrane domain did not increase basal adenylate cyclase activity, indicating that the first 294 residues did not act as an inverse agonist. The truncated LHR_316 was still activated by hCG but with an EC50 higher than that for the wild-type receptor. Zero length cross-linking was used to study intramolecular interactions between the two domains of LHR_Fur316. Cross-linking efficiency was similar for the basal and activated states, which indicated that the two domains interacted closely in the basal state, and this tight interaction persisted during activation. Our data suggest that activation of the LHR results from subtle modifications of intramolecular interactions between the two domains and low-affinity binding of hCG to the extracellular loops or residues preceding the first transmembrane segment.

Aspartic Acid↗
PubMed2005

Development and potential clinical uses of human prolactin receptor antagonists.

There is a large body of literature showing that prolactin (PRL) exerts growth-promoting activities in breast cancer, and possibly in prostate cancer and prostate hyperplasia. In addition, increasing evidence argues for the involvement of locally produced (autocrine) PRL, perhaps even more than pituitary-secreted (endocrine) PRL, in tumor growth. Because dopamine analogs are unable to inhibit PRL production in extrapituitary sites, alternative strategies need investigation. To that end, several PRL receptor antagonists have been developed by introducing various mutations into its natural ligands. For all but one of these analogs, the mechanism of action involves a competition with endogenous PRL for receptor binding. Such compounds are thus candidates to counteract the undesired actions of PRL, not only in tumors, but also in dopamine-resistant prolactinomas. In this review, we describe the different versions of antagonists that have been developed, with emphasis on the controversies regarding their characterization, and the limits for their potential development as a drug. The most recently developed antagonist, Delta1-9-G129R-hPRL, is the only one that is totally devoid of residual agonistic activity, meaning it acts as pure antagonist. We discuss to what extent this new molecule could be considered as a lead compound for inhibiting the actions of human PRL in the above-mentioned diseases. We also speculate on the multiple questions that could be addressed with respect to the therapeutic use of PRL receptor antagonists in patients.

Animals↗
PubMed2005

Atrophy of S6K1(-/-) skeletal muscle cells reveals distinct mTOR effectors for cell cycle and size control.

The mammalian target of rapamycin (mTOR) and Akt proteins regulate various steps of muscle development and growth, but the physiological relevance and the downstream effectors are under investigation. Here we show that S6 kinase 1 (S6K1), a protein kinase activated by nutrients and insulin-like growth factors (IGFs), is essential for the control of muscle cytoplasmic volume by Akt and mTOR. Deletion of S6K1 does not affect myoblast cell proliferation but reduces myoblast size to the same extent as that observed with mTOR inhibition by rapamycin. In the differentiated state, S6K1(-/-) myotubes have a normal number of nuclei but are smaller, and their hypertrophic response to IGF1, nutrients and membrane-targeted Akt is blunted. These growth defects reveal that mTOR requires distinct effectors for the control of muscle cell cycle and size, potentially opening new avenues of therapeutic intervention against neoplasia or muscle atrophy.

Animals↗
PubMed2005

Diet-induced obesity impairs mammary development and lactogenesis in murine mammary gland.

We have developed a mouse model of diet-induced obesity that shows numerous abnormalities relating to mammary gland function. Animals ate approximately 40% more calories when offered a high-fat diet and gained weight at three times the rate of controls. They exhibited reduced conception rates, increased peripartum pup mortality, and impaired lactogenesis. The impairment of lactogenesis involved lipid accumulation in the secretory epithelial cells indicative of an absence of copius milk secretion. Expression of mRNAs for beta-casein, whey acid protein, and alpha-lactalbumin were all decreased immediately postpartum but recovered as lactation was established over 2-3 days. Expression of acetyl-CoA carboxylase (ACC)-alpha mRNA was also decreased at parturition as was the total enzyme activity, although there was a compensatory increase in the proportion in the active state. By day 10 of lactation, the proportion of ACC in the active state was also decreased in obese animals, indicative of suppression of de novo fatty acid synthesis resulting from the supply of preformed fatty acids in the diet. Although obese animals consumed more calories in the nonpregnant and early pregnant states, they showed a marked depression in fat intake around day 9 of pregnancy before food intake recovered in later pregnancy. Food intake increased dramatically in both lean and obese animals during lactation although total calories consumed were identical in both groups. Thus, despite access to high-energy diets, the obese animals mobilized even more adipose tissue during lactation than their lean counterparts. Obese animals also exhibited marked abnormalities in alveolar development of the mammary gland, which may partially explain the delay in differentiation evident during lactogenesis.

Acetyl-CoA Carboxylase↗
PubMed2005

Benign breast diseases.

Benign breast diseases have always been neglected in comparison to cancer, despite the fact that there are many more patients with such diseases than patients presenting to a breast clinic for cancer. Like normal breast tissues, benign breast diseases are under a complex system of controls by both systemic hormonal and local factors. In this review, we attempt to present an overview of the latest knowledge concerning the epidemiology, classification, clinical presentation, management, and physiopathology of these disorders.

Adolescent↗
PubMed2004

Roles of the lactogens and somatogens in perinatal and postnatal metabolism and growth: studies of a novel mouse model combining lactogen resistance and growth hormone deficiency.

To delineate the roles of the lactogens and GH in the control of perinatal and postnatal growth, fat deposition, insulin production, and insulin action, we generated a novel mouse model that combines resistance to all lactogenic hormones with a severe deficiency of pituitary GH. The model was created by breeding PRL receptor (PRLR)-deficient (knockout) males with GH-deficient (little) females. In contrast to mice with isolated GH or PRLR deficiencies, double-mutant (lactogen-resistant and GH-deficient) mice on d 7 of life had growth failure and hypoglycemia. These findings suggest that lactogens and GH act in concert to facilitate weight gain and glucose homeostasis during the perinatal period. Plasma insulin and IGF-I and IGF-II concentrations were decreased in both GH-deficient and double-mutant neonates but were normal in PRLR-deficient mice. Body weights of the double mutants were reduced markedly during the first 3-4 months of age, and adults had striking reductions in femur length, plasma IGF-I and IGF binding protein-3 concentrations, and femoral bone mineral density. By age 6-12 months, however, the double-mutant mice developed obesity, hyperleptinemia, fasting hyperglycemia, relative hypoinsulinemia, insulin resistance, and glucose intolerance; males were affected to a greater degree than females. The combination of perinatal growth failure and late-onset obesity and insulin resistance suggests that the lactogen-resistant/GH-deficient mouse may serve as a model for the development of the metabolic syndrome.

Adipose Tissue↗
PubMed2004

Cathepsin D processes human prolactin into multiple 16K-like N-terminal fragments: study of their antiangiogenic properties and physiological relevance.

16K prolactin (PRL) is the name given to the 16-kDa N-terminal fragment obtained by proteolysis of rat PRL by tissue extracts or cell lysates, in which cathepsin D was identified as the candidate protease. Based on its antiangiogenic activity, 16K PRL is potentially a physiological inhibitor of tumor growth. Full-length human PRL (hPRL) was reported to be resistant to cathepsin D, suggesting that antiangiogenic 16K PRL may be physiologically irrelevant in humans. In this study, we show that hPRL can be cleaved by cathepsin D or mammary cell extracts under the same conditions as described earlier for rat PRL, although with lower efficiency. In contrast to the rat hormone, hPRL proteolysis generates three 16K-like fragments, which were identified by N-terminal sequencing and mass spectrometry as corresponding to amino acids 1-132 (15 kDa), 1-147 (16.5 kDa), and 1-150 (17 kDa). Biochemical and mutagenetic studies showed that the species-specific digestion pattern is due to subtle differences in primary and tertiary structures of rat and human hormones. The antiangiogenic activity of N-terminal hPRL fragments was assessed by the inhibition of growth factor-induced thymidine uptake and MAPK activation in bovine umbilical endothelial cells. Finally, an N-terminal hPRL fragment comigrating with the proteolytic 17-kDa fragment was identified in human pituitary adenomas, suggesting that the physiological relevance of antiangiogenic N-terminal hPRL fragments needs to be reevaluated in humans.

Amino Acid Sequence↗
PubMed2003

The N-terminus of human prolactin modulates its biological properties.

The N-terminus is the most divergent region within the prolactin (PRL)/placental lactogen (PL)/growth hormone (GH) family. Since all of these ligands are able to activate the lactogen receptor, it has been usually assumed that the N-terminus plays no major role in biological actions of any family member. In this study, we generated several analogs of human PRL in which the N-terminus was truncated by 9 and iteratively up to the 14 first residues. Truncation did not alter protein folding, and it even decreased the formation of PRL aggregates that appear during the purification of refolded protein. Removal of the entire N-terminal loop (14 residues) decreased the affinity for the receptor by two-three-fold, and reduced the ability of the hormone to activate the human lactogen receptor. In contrast, removal of 13 or less residues improves receptor activation since these analogs are able to produce supra-maximal activities in a transcriptional bioassay, or in proliferation assays exhibit dose-response curves that are less bell-shaped, which reflects enhanced stabilization of receptor dimers. Altogether, these data suggest that the N-terminus of PRL is actually slightly detrimental to bioactivity, but may be required for other properties of the hormone.

Amino Acid Sequence↗
PubMed2003

Development of pure prolactin receptor antagonists.

Prolactin (PRL) promotes tumor growth in various experimental models and leads to prostate hyperplasia and mammary neoplasia in PRL transgenic mice. Increasing experimental evidence argues for the involvement of autocrine PRL in this process. PRL receptor antagonists have been developed to counteract these undesired proliferative actions of PRL. However, all forms of PRL receptor antagonists obtained to date exhibit partial agonism, preventing their therapeutic use as full antagonists. In the present study, we describe the development of new human PRL antagonists devoid of agonistic properties and therefore able to act as pure antagonists. This was demonstrated using several in vitro bioassays, including highly sensitive assays able to detect extremely low levels of receptor activation. These new compounds also act as pure antagonists in vivo, as assessed by analyzing their ability to competitively inhibit PRL-triggered signaling cascades in various target tissues (liver, mammary gland, and prostate). Finally, by using transgenic mice expressing PRL specifically in the prostate, which exhibit constitutively activated signaling cascades paralleling hyperplasia, we show that these new PRL analogs are able to completely revert PRL-activated events. These second generation human PRL antagonists are good candidates to be used as inhibitors of growth-promoting actions of PRL.

Amino Acid Motifs↗