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Biomedical subjects

M Marcinkiewicz

Publications and source records attributed to M Marcinkiewicz.

At least 19 recordsLinked to original sources
PubMed2004

Changes of atrial natriuretic peptide in rat supraoptic neurones during pregnancy.

To better understand the role of atrial natriuretic peptide (ANP) in the central regulation of hydro-mineral homeostasis, we analysed its expression in rat hypothalamic neurones during gestation and postpartum. These physiological events are characterized by opposing body fluid regulations. Quantitative in situ hybridization analysis showed that starting from mid-pregnancy, ANP mRNA declined in neurones of the preoptic area, periventricular area, lateral hypothalamus and endorhinal nucleus, and remained low at postpartum. By contrast, magnocellular cells in the supraoptic nucleus (SON) showed four- and 10-fold more ANP mRNA in sections from preterm and postpartum rats, respectively, compared to nonpregnant controls (P < 0.001). Oxytocin mRNA paralleled ANP mRNA expression in the SON, whereas vasopressin mRNA rose in early pregnancy and declined thereafter. High hypothalamic ANP concentration at day 21 of gestation versus nonpregnant rats (3.1 +/- 0.5 versus 1.8 +/- 0.4 ng/mg protein, P < 0.05) suggested that ANP transcript accumulation in the SON is associated with increased utilization of the peptide. The elevation of hypothalamic ANP (two-fold) and ANP receptors by treatment of ovariectomized rats with 17beta-oestradiol (25 micro g/rat, 10 days) was abolished by coadministration of progesterone. Thus, we concluded that elevated oestradiol at term stimulates ANP synthesis and paracrine ANP activation in the hypothalamus. Overall, we provide experimental, anatomical and molecular evidence for ANP regulation in hypothalamic neurones at preterm and after 17beta-oestradiol stimulation. Our study supports the concept that ANP expressed in the SON acts as a peptidergic neurotransmitter involved in water and salt regulation during pregnancy and postpartum.

Adaptation, Physiological↗
PubMed2001

Regional and subunit-specific downregulation of acid-sensing ion channels in the pilocarpine model of epilepsy.

Acid-sensing ion channels (ASICs) constitute a recently discovered family of excitatory cation channels, structurally related to the superfamily of degenerin/epithelial sodium channels. ASIC1b and ASIC3 are highly expressed in primary sensory neurons and are thought to play a role in pain transmission related to acidosis. ASIC1a, ASIC2a, and ASIC2b are also distributed in the central nervous system where their function remains unclear. We investigated here the regulation of their expression during status epilepticus (SE), a condition in which neuronal overexcitation leads to acidosis. In animals treated with pilocarpine (380 mg/kg) to induce SE, we observed a marked decrease of ASIC2b mRNA levels in all hippocampal areas and of ASIC1a mRNA levels in the CA1-2 fields. These changes were also observed after protective treatment from neuronal cell death with diazepam (10 mg/kg) and pentobarbital (30 mg/kg). These findings suggest a key role of channels containing ASIC1a and ASIC2b subunits in both normal and pathological activity of hippocampus.

Acid Sensing Ion Channels↗
PubMed2001

Brain-derived neurotrophic factor superinduction parallels anti-epileptic--neuroprotective treatment in the pilocarpine epilepsy model.

Antiepileptic drugs provide neuroprotection in several animal models of brain damage, including those induced by status epilepticus (SE). The mechanisms involved in this action are unknown, but neurotrophic factors such as brain-derived neurotrophic factor (BDNF) may play a role. In this study we investigated the changes in BDNF levels in rats in which SE had been induced by pilocarpine injection (400 mg/kg i.p.) and continued for several hours (unprotected group). In other animals (protected groups), SE was suppressed after 30 min by intraperitoneal injection of either diazepam (10 mg/kg) + pentobarbital (30 mg/kg) or paraldehyde (0.3 mg/kg). In diazepam + pentobarbital-treated rats the hippocampal damage caused by SE was significantly lower (p < 0.05) than in unprotected animals. In addition, 2 and 24 h after pilocarpine injection, the levels of BDNF mRNA were moderately increased in the unprotected group, but 'superinduced' in protected animals, especially in the neocortex and hippocampus. A time-dependent increase in BDNF immunoreactivity was also found by western blot analysis in rats treated with diazepam + pentobarbital. In contrast, a decrease of BDNF immunoreactivity occurred in the unprotected group. In conclusion, these results show that neuroprotection induced by anti-epileptic drugs in pilocarpine-treated rats is accompanied by strong potentiation of BDNF synthesis in brain regions involved in SE.

Animals↗
PubMed2001

Selective expression of the proprotein convertases furin, pc5, and pc7 in proliferating vascular smooth muscle cells of the rat aorta in vitro.

The aim of this study was to investigate whether transformation of quiescent vascular smooth muscle cells (VSMCs) into proliferating secretory cells is accompanied by an expression of processing enzymes that activate de novo-synthesized growth factors. Three enzymes belonging to the family of the kexin/subtilisin-like mammalian proprotein convertases (PCs), furin, PC5, and PC7, were found to be upregulated after balloon denudation in vivo. To determine their importance in these cell processes, we investigated their gene regulation using a short-term organ culture system. After incubation of rat aorta for 4 and 24 hr in serum-free medium, we demonstrated a significant induction of VSMC proliferation. The affected subset of VSMCs, positive for alpha-smooth muscle actin, also expressed proliferating cell nuclear antigen (PCNA). Our results revealed a parallel upregulation of furin, PC5, and PC7 in PCNA-immunolabeled cells. As a substrate model for comparison with PCs we used nerve growth factor (NGF). NGF is known to be activated by PCs. As shown by Northern blotting analysis, NGF mRNA concentration was significantly increased in cultured explants. NGF was released into the culture medium. In conclusion, both PCs and NGF are coordinately modulated on induction of VSMC proliferation.

Animals↗
PubMed2000

Molecular cloning and biochemical characterization of a new mouse testis soluble-zinc-metallopeptidase of the neprilysin family.

Because of their roles in controlling the activity of several bio-active peptides, members of the neprilysin family of zinc metallopeptidases have been identified as putative targets for the design of therapeutic agents. Presently, six members have been reported, these are: neprilysin, endothelin-converting enzyme (ECE)-1 and ECE-2, the Kell blood group protein, PHEX (product of the phosphate-regulating gene with homologies to endopeptidase on the X chromosome) and X-converting enzyme (XCE). In order to identify new members of this important family of peptidases, we designed a reverse transcriptase-PCR strategy based on conserved amino acid sequences of neprilysin, ECE-1 and PHEX. We now report the cloning from mouse testis of a novel neprilysin-like peptidase that we called NL1. NL1 is a glycoprotein that, among the members of the family, shows the strongest sequence identity with neprilysin. However, in contrast with neprilysin and other members of the family which are type II integral membrane proteins, NL1 was secreted when expressed in cultured mammalian cells, likely due to cleavage by a subtilisin-like convertase at a furin-like site located 22 amino acid residues in the C-terminus of the transmembrane domain. The recombinant enzyme exhibited neprilysin-like peptidase activity and was efficiently inhibited by phosphoramidon and thiorphan, two inhibitors of neprilysin. Northern blot analysis and in situ hybridization showed that NL1 mRNA was found predominantly in testis, specifically in round and elongated spermatids. This distribution of NL1 mRNA suggests that it could be involved in sperm formation or other processes related to fertility.

Amino Acid Sequence↗
PubMed2000

Localization of retinal dehydrogenase type 1 in the stomach and intestine.

Retinal dehydrogenase type 1 (RALDH1) is involved in the biosynthesis of retinoic acid (RA), a modulator of gene expression and cell differentiation. RALDH1 mRNA transcripts are present in the stomach and small intestine, and their expression is regulated by vitamin A status. In situ hybridization demonstrated RALDH1 mRNA expression in epithelial cells of the stomach, small intestine, and large intestine. Strong hybridization was also seen in the lamina propria of small intestinal mucosa and the smooth muscle layer of the small and large intestines. Immunocytochemical localization revealed RALDH1 staining in parietal cells of the stomach and prismatic cells of the small and large intestines. The presence of RALDH1 protein was also detectable within supportive glial cells around neuronal fibers throughout the muscular layers of the stomach as well as the small and large intestines. These data suggest an important role for RALDH1 in generating RA needed for the differentiation of specific epithelial cells in the stomach and intestines.

Aldehyde Oxidoreductases↗
PubMed2000

Study of salivary response to continuous infusion of cerulein and secretin in healthy subjects.

OBJECTIVE: The effects of intravenous infusion of cerulein and secretin on the secretion and biochemical composition of whole saliva in humans were studied. STUDY DESIGN: A total of 33 subjects were divided into 3 groups, which underwent intravenous infusion with saline solution (NaCl 0.15 mol/L(-1)); saline solution, cerulein (1.25 x 10(-3) microg kg(-1) min(-1)), and cerulein supplemented with secretin (2.5 x 10(-2) clinical units kg(-1) min(-1)); and saline solution, secretin (2.5 x 10(-2) clinical units kg(-1) min(-1)), and secretin supplemented with cerulein (1.25 x 10(-3) microg kg(-1) min(-1)). RESULTS: Cerulein reduced salivary flow rate, bicarbonate concentration and output, and protein output and increased amylase activity. The inhibitory effect of cerulein on salivary flow rate and bicarbonate concentration and output prevailed when an infusion of cerulein was supplemented with secretin. Cerulein and secretin acting together increased protein concentration. Secretin alone decreased salivary flow rate and bicarbonate concentration, whereas secretin supplemented with cerulein not only decreased salivary flow rate, bicarbonate concentration, and bicarbonate output but also increased protein concentration. CONCLUSION: The effect of secretin and cerulein on salivary secretion and its composition is quite different from that observed in the pancreas.

Adult↗
PubMed2000

Coordinated expression of beta-amyloid precursor protein and the putative beta-secretase BACE and alpha-secretase ADAM10 in mouse and human brain.

To define the enzymes involved in the etiology of Alzheimer's disease, we compared in mouse and human brain the mRNA levels and cellular localization of the ubiquitous beta-amyloid precursor protein (beta-APP) with those of the putative alpha-secretases ADAM10 and ADAM17 and the beta-secretases BACE and BACE2. In situ hybridization performed in mice during prenatal and postnatal development and in adulthood revealed the coexpression of beta-APP, BACE, and ADAM10. The patterns of BACE2 and ADAM17 only partially overlapped with that of beta-APP. beta-APP, BACE, and ADAM10 mRNAs have also been detected by northern blot in human brain cortex of normal subjects and in Alzheimer's disease subjects. In situ hybridization performed using combined biotin- and radiolabeled riboprobes provided evidence for the coexpression of beta-APP with BACE and ADAM10 in human cortical neurons. Our data provide cytochemical evidence supporting the role of ADAM10 and BACE as authentic alpha- and beta-secretases.

ADAM Proteins↗
PubMed2000

The potential role of the esophageal pre-epithelial barrier components in the maintenance of integrity of the esophageal mucosa in patients with endoscopically negative gastroesophageal reflux disease.

OBJECTIVE: Patients with gastroesophageal reflux disease (GERD) accompanied by erosive reflux esophagitis (RE) exhibit an impairment within the esophageal pre-epithelial barrier protective components that may facilitate the development and/or progression of the mucosal injury. Little is known, however, whether such impairment is a general phenomenon affecting all patients with GERD or whether this is a characteristic feature only of patients with erosive RE. We therefore studied the rate of secretion of esophageal inorganic and organic protective factors in patients with endoscopically negative [E (-)] GERD and compared these results with the corresponding values in asymptomatic volunteers (CTRL). METHODS: The study was conducted on 33 white asymptomatic volunteers and 10 white patients with a long history of GERD confirmed by 24-h pH monitoring and a grossly negative upper endoscopy. Esophageal secretion was collected during mucosal exposure to NaCl, HCl, HC/pepsin and NaCl using the esophageal perfusion catheter. In collected samples all investigated parameters were measured. RESULTS: The pH of esophageal secretion and its content of bicarbonate, EGF, and PGE2 in patients with E (-) GERD and asymptomatic volunteers were similar. Unexpectedly, the rate of esophageal glycoconjugate (predominantly mucin) secretion was significantly higher in patients with E (-) GERD than in controls during perfusion with HCl (p < 0.05). Furthermore, secretion of protein in patients with E (-) GERD was significantly higher than in the control group during the mucosal exposure to HCl/Pepsin (p < 0.05). The nonbicarbonate buffer secretion during perfusion with HCl and HCl/Pepsin as well as the rate of esophageal TGFalpha output during infusion of final saline in patients with E (-) GERD were significantly lower than in CTRL group (p < 0.05). CONCLUSIONS: Our data indicate that patients with E (-) GERD have an esophageal secretory potential, in terms of glycoconjugate and protein, higher than that in asymptomatic controls. This phenomenon in patients with E (-) GERD may, by enhancing the quantity of the esophageal pre-epithelial barrier, help to prevent the development of erosive esophagitis. A significantly lower esophageal secretory response in patients with E (-) GERD in terms of nonbicarbonate buffers and TGFalpha may facilitate the development of GERD symptoms and histological changes of GERD, respectively.

Adult↗
PubMed2000

Cellular localization of neprilysin in mouse bone tissue and putative role in hydrolysis of osteogenic peptides.

The regulation of osteoblast and osteoclast metabolism is mediated by both hormones and local bone peptide factors. Peptides and hormones are under control of membrane peptidases such as Neprilysin (NEP). NEP is a widely distributed cell-surface zinc-metallopeptidase that is involved in the regulation of several important physiological processes by controlling the half-life of bioactive peptides. Although NEP is known to be present in skeletal tissues, neither its cellular localization nor its function have been established. To address this question, we examined NEP distribution in bones of postnatal mouse. In situ hybridization (ISH) and immunohistochemistry showed that NEP messenger RNA (mRNA) and protein are associated with bone-forming cells including presumptive osteoblast precursors, preosteoblasts, osteoblasts, and osteocytes. NEP levels in newborn and adult mice bones also were compared by immunoblotting. Higher amounts of NEP immunoreactivity were observed in newborn as compared with adult bones, suggesting a relationship between NEP expression and bone growth. To further explore this hypothesis, we monitored in vitro NEP proteolytic activity using a series of synthetic osteogenic peptides such as parathyroid hormone-related peptide 1-43 (PTHrP1-34), osteostatin (PTHrP107-139), osteogenic growth peptide (OGP), calcitonin, alpha-calcitonin gene-related peptide (alpha-CGRP), and PTH1-34. Except for PTH1-34, all peptides were found to be NEP substrates.

Aging↗
PubMed2000

Developmental expression and tissue distribution of Phex protein: effect of the Hyp mutation and relationship to bone markers.

Mutations in PHEX, a phosphate-regulating gene with homology to endopeptidases on the X chromosome, are responsible for X-linked hypophosphatemia (XLH). The murine Hyp homologue has the phenotypic features of XLH and harbors a large deletion in the 3' region of the Phex gene. We characterized the developmental expression and tissue distribution of Phex protein, using a monoclonal antibody against human PHEX, examined the effect of the Hyp mutation on Phex expression, and compared neprilysin (NEP), osteocalcin, and parathyroid hormone/parathyroid hormone-related protein (PTH/PTHrP) receptor gene expression in bone of normal and Hyp mice. Phex encodes a 100- to 105-kDa glycoprotein, which is present in bones and teeth of normal mice but not Hyp animals. These results were confirmed by in situ hybridization (ISH) and ribonuclease protection assay. Phex protein expression in femur and calvaria decreases with age, suggesting a correlation between Phex expression and bone formation. Immunohistochemical studies detected Phex protein in osteoblasts, osteocytes, and odontoblasts, but not in osteoblast precursors. In contrast to Phex, the abundance of NEP messenger RNA (mRNA) and protein is not significantly altered in Hyp bone. Similarly, osteocalcin and PTH/PTHrP receptor gene expression are not compromised in bone of Hyp mice. Our results are consistent with the hypothesis that loss of Phex function affects the mineralizing activity of osteoblasts rather than their differentiation.

Animals↗
PubMed1999

The subtilisin/kexin family of precursor convertases. Emphasis on PC1, PC2/7B2, POMC and the novel enzyme SKI-1.

Proopiomelanocortin (POMC) is a precursor to various, bioactive peptides including ACTH, beta LPH, alpha MSH, and beta endorphin (beta END). Processing of POMC at dibasic residues is tissue-specific and is performed by either PC1 alone (resulting in ACTH and beta LPH, anterior pituitary corticotrophes) or by a combination of PC1 and PC2 (yielding alpha MSH and beta END, pituitary neurointermediate lobe and hypothalamus). The PC2-specific binding protein 7B2 is intimately involved in the zymogen activation of proPC2 into PC2. Structure-function studies of these enzymes demonstrated the presence of N- and C-terminal domains, as well as specific amino acids within the catalytic segment that influence the degree of activity of each enzyme and the interaction of PC2 with 7B2. The tissue distribution, plasticity of expression, and the multiple precursors that are differentially cleaved by PC1 and/or PC2, predict a wide array of combinatorial activities of these convertases within the endocrine and neuroendocrine system. The phenotypic consequences of the absence of genetic expression of either PC1 or PC2 are now explored using knockout mice and in human patients suffering from obesity and diabetes.

Animals↗
PubMed1999

Mammalian subtilisin/kexin isozyme SKI-1: A widely expressed proprotein convertase with a unique cleavage specificity and cellular localization.

Using reverse transcriptase-PCR and degenerate oligonucleotides derived from the active-site residues of subtilisin/kexin-like serine proteinases, we have identified a highly conserved and phylogenetically ancestral human, rat, and mouse type I membrane-bound proteinase called subtilisin/kexin-isozyme-1 (SKI-1). Computer databank searches reveal that human SKI-1 was cloned previously but with no identified function. In situ hybridization demonstrates that SKI-1 mRNA is present in most tissues and cells. Cleavage specificity studies show that SKI-1 generates a 28-kDa product from the 32-kDa brain-derived neurotrophic factor precursor, cleaving at an RGLT downward arrowSL bond. In the endoplasmic reticulum of either LoVo or HK293 cells, proSKI-1 is processed into two membrane-bound forms of SKI-1 (120 and 106 kDa) differing by the nature of their N-glycosylation. Late along the secretory pathway some of the membrane-bound enzyme is shed into the medium as a 98-kDa form. Immunocytochemical analysis of stably transfected HK293 cells shows that SKI-1 is present in the Golgi apparatus and within small punctate structures reminiscent of endosomes. In vitro studies suggest that SKI-1 is a Ca2+-dependent serine proteinase exhibiting a wide pH optimum for cleavage of pro-brain-derived neurotrophic factor.

Amino Acid Sequence↗
PubMed1999

Nerve growth factor and proprotein convertases furin and PC7 in transected sciatic nerves and in nerve segments cultured in conditioned media: their presence in Schwann cells, macrophages, and smooth muscle cells.

Synthesis of proteins such as nerve growth factor (NGF) is induced after nerve lesion. The NGF precursor (pro-NGF) requires a posttranslational processing by proprotein convertases to become active. In this report, we re-examine the localization of NGF protein and mRNA in injured nerve and show that the candidate pro-NGF convertases furin and PC 7 colocalize with NGF in non-neuronal cells in nerve. By Northern blot analysis, 1.5-kb and 1.3-kb NGF mRNAs were shown to be increased in distal and immediately proximal nerve segments on days 1, 4, and 14 after lesion; by Western blot analysis, NGF proteins of high molecular weight were detected after injury. In vivo, two phases of NGF immunopositivity were observed, in macrophages and perivascular cells shortly after lesion and in endoneurial cells on day 1 and 4. To identify the cells containing NGF, nerve segments were incubated in serum-containing medium with or without conditioning by white blood cells isolated from the circulation. Both hybridization and immunoreactivity signals for NGF were elevated after incubation of nerve segments for 4 hours in conditioned media, so that cells with NGF immunoreactivity could be identified by antibodies to specific cell markers. In these nerve fragments, Schwann cells, perivascular smooth muscle cells, and macrophages contained NGF immunoreactivity. The concentration of furin and PC7 mRNA also increased in lesioned nerves. By immunocytochemical investigation of nerve explants, furin and PC7 were detected in endoneurial cells, macrophages and perivascular cells and were colocalized with NGF. These in vitro and in vivo findings suggest that both furin and PC7 are associated with NGF in several cell types of the sciatic nerve and, hence, may be implicated in intracellular processing of pro-NGF.

Animals↗
PubMed1999

Influence of secretin and caerulein on acid phosphatase activity in human saliva.

Experimental studies have demonstrated a structural and functional correlation between the pancreas and salivary glands. Therefore, one may suppose that caerulein plus secretin, used in functional exploration of the exocrine pancreas, could exert an influence on acid phosphatase activity (AcP) and its isoenzymes (tartrate sensitive-TsAcP, tartrate resistant-TRAP acid phosphatases) in whole saliva in humans. The current study was conducted in 14 asymptomatic volunteers (six woman and eight men, mean age 34, range 27-45). Salivary secretions were collected under basal conditions (intravenous infusion of 0.15 mol l-1 NaCl) and during intravenous infusion of secretin plus caerulein at different doses. Stimulation with secretin (50 mKE kg-1 min-1) and caerulein (2.5 ng kg-1 min-1) resulted in a marked and rapid decrease of salivary flow rate. TRAP activity decreased during the first 20 min of hormonal infusion (1.09 +/- 0.19 mU ml-1 vs. 1.93 +/- 0.18 mU ml-1, 1.16 +/- 0.12 mU ml-1 vs. 1.86 +/- 0.17 mU ml-1, p < 0.05). AcP and TsAcP activity during intravenous infusion of secretin plus caerulein remained unchanged. These results point to a wider than expected participation of the digestive system in hormonal control of biochemical processes taking place inside the oral cavity.

Acid Phosphatase↗