Abnormal Microscopic Findings in the Placenta Correlate With the Severity of Fetal Heart Failure.

BACKGROUND: This study investigated the association between placental pathology and fetal heart failure.Methods and Results: Singletons with a congenital heart defect (CHD) and/or arrhythmia (n=168) and gestational age-matched controls (n=52) were included in the study. The associations between macro- and microscopic abnormal findings of the placenta and the severity of fetal heart failure were evaluated using the cardiovascular profile (CVP) score. Nine features were microscopically identified and assessed in sections of the placenta: premature villi, edematous villi, fibrotic villi, chorioamnionitis, chorangiosis, fibrin deposition, subchorionic hematoma, infarcted villi, and nucleated red blood cells in villous vessels. Among singletons with CHD and/or arrhythmia, the final CVP score was ≥8 in 140 cases, 6 or 7 in 15 cases, and ≤5 in 13 cases. Microscopic analysis showed that the frequency and severity of premature and edematous villi and increased nucleated red blood cells in villous vessels were greater in cases of fetal heart failure. These microscopic findings were more common and severe in cases with a final CVP score ≤5 than in gestational age-matched controls. The prevalence of abnormal macroscopic findings of the placenta and umbilical cord was similar regardless of the severity of fetal heart failure. CONCLUSIONS: Premature and edematous villi and increased nucleated red blood cells in villous vessels were correlated with the severity of fetal heart failure in cases of CHD and/or arrhythmia.

Utility of perinatal natriuretic peptide for predicting neonatal heart failure.

BACKGROUND: We evaluated the significance of perinatal plasma natriuretic peptide (NP) levels in neonates with congenital heart defects (CHDs) or arrhythmias and determined whether measurement of perinatal plasma NP levels and echocardiographic assessment in utero could predict heart failure after birth. METHODS: The study was conducted between 2012 and 2016 to evaluate the correlation of perinatal atrial NP (ANP) and brain NP (BNP) levels at birth with the modified Ross score after birth and the cardiovascular profile (CVP) score before birth. RESULTS: A total of 122 singletons with CHDs or arrhythmias and 27 controls were analyzed. Neonatal blood sampling was performed at a median of 0.7 h (range, 0.1-1.5) after birth. The neonatal plasma ANP and BNP levels shortly after birth were significantly higher than those in the umbilical artery (UA) plasma. The ANP and BNP levels in UA and neonatal blood were correlated with the modified Ross score. The neonatal plasma ANP and BNP levels and the modified Ross scores were inversely correlated with the CVP score in neonates with CHDs or arrhythmias. The area under the receiver operating characteristic curve of UA ANP levels for predicting neonatal heart failure was highest among those for the CVP score, perinatal plasma ANP and BNP levels, and their combinations. CONCLUSIONS: The plasma ANP and BNP levels increased markedly shortly after birth. Assessment of the UA plasma ANP level at birth and the CVP score in utero may be utilized to predict neonatal heart failure.

Deficiency of Cardiac Natriuretic Peptide Signaling Promotes Peripartum Cardiomyopathy-Like Remodeling in the Mouse Heart.

BACKGROUND: The maternal circulatory system and hormone balance both change dynamically during pregnancy, delivery, and the postpartum period. Although atrial natriuretic peptides and brain natriuretic peptides produced in the heart control circulatory homeostasis through their common receptor, NPR1, the physiologic and pathophysiologic roles of endogenous atrial natriuretic peptide/brain natriuretic peptide in the perinatal period are not fully understood. METHODS: To clarify the physiologic and pathophysiologic roles of the endogenous atrial natriuretic peptide/brain natriuretic peptide-NPR1 system during the perinatal period, the phenotype of female wild-type and conventional or tissue-specific Npr1-knockout mice during the perinatal period was examined, especially focusing on maternal heart weight, blood pressure, and cardiac function. RESULTS: In wild-type mice, lactation but not pregnancy induced reversible cardiac hypertrophy accompanied by increases in fetal cardiac gene mRNAs and ERK1/2 (extracellular signaling-regulated kinase) phosphorylation. Npr1-knockout mice exhibited significantly higher plasma aldosterone level than did wild-type mice, severe cardiac hypertrophy accompanied by fibrosis, and left ventricular dysfunction in the lactation period. Npr1-knockout mice showed a high mortality rate over consecutive pregnancy-lactation cycles. In the hearts of Npr1-knockout mice during or after the lactation period, an increase in interleukin-6 mRNA expression, phosphorylation of signal transducer and activator of transcription 3, and activation of the calcineurin-nuclear factor of the activated T cells pathway were observed. Pharmacologic inhibition of the mineralocorticoid receptor or neuron-specific deletion of the mineralocorticoid receptor gene significantly ameliorated cardiac hypertrophy in lactating Npr1-knockout mice. Anti-interleukin-6 receptor antibody administration tended to reduce cardiac hypertrophy in lactating Npr1-knockout mice. CONCLUSIONS: These results suggest that the characteristics of lactation-induced cardiac hypertrophy in wild-type mice are different from exercise-induced cardiac hypertrophy, and that the endogenous atrial natriuretic peptide/brain natriuretic peptide-NPR1 system plays an important role in protecting the maternal heart from interleukin-6-induced inflammation and remodeling in the lactation period, a condition mimicking peripartum cardiomyopathy.

Comparison of physiological functions between neuromedin U-related peptide and neuromedin S-related peptide in the rat central nervous system.

Two novel peptides, neuromedin U precursor-related peptide (NURP) and neuromedin S precursor-related peptide (NSRP), are produced from neuromedin U (NMU) and neuromedin S (NMS) precursors, respectively, as these precursors have multiple consensus sequences for proteolytic processing. Our group has shown previously that one of these two novel peptides, NURP, stimulates body temperature and locomotor activity, but not food intake. However, the physiological function of the other peptide, NSRP, has remained unclear. Therefore, the aim of this study was to characterize differences in the regions of the rat brain targeted by the NMU/NMS peptide family, including NURP and NSRP, and their physiological functions. First, we explored the regions of c-Fos expression after intracerebroventricular (i.c.v.) injection of NURP and NSRP and found that these were fewer than after i.c.v. injection of NMU and NMS in the hypothalamus, possibly because NURP and NSRP cannot activate NMU/NMS receptors. In the ventral subiculum, which is one region of the hippocampus, c-Fos expression was evident only after i.c.v. injection of NURP. We also examined the effects of NSRP on food intake, body temperature and locomotor activity. Like NURP, NSRP increased both body temperature and locomotor activity, but not food intake, indicating that NSRP is also a functional peptide. However, these effects of NSRP were distinctly weaker than those of NURP. These findings suggest differences in the affinity of NURP and/or NSRP for specific receptors, or in their respective biological activities.

Characterization of putative tachykinin peptides in Caenorhabditis elegans.

Tachykinin-like peptides, such as substance P, neurokinin A, and neurokinin B, are among the earliest discovered and best-studied neuropeptide families, and research on them has contributed greatly to our understanding of the endocrine control of many physiological processes. However, there are still many orphan tachykinin receptor homologs for which cognate ligands have not yet been identified, especially in small invertebrates, such as the nematode Caenorhabditis elegans (C. elegans). We here show that the C. elegans nlp-58 gene encodes putative ligands for the orphan G protein-coupled receptor (GPCR) TKR-1, which is a worm ortholog of tachykinin receptors. We first determine, through an unbiased biochemical screen, that a peptide derived from the NLP-58 preprotein stimulates TKR-1. Three mature peptides that are predicted to be generated from NLP-58 show potent agonist activity against TKR-1. We designate these peptides as C. elegans tachykinin (CeTK)-1, -2, and -3. The CeTK peptides contain the C-terminal sequence GLR-amide, which is shared by tachykinin-like peptides in other invertebrate species. nlp-58 exhibits a strongly restricted expression pattern in several neurons, implying that CeTKs behave as neuropeptides. The discovery of CeTKs provides important information to aid our understanding of tachykinin-like peptides and their functional interaction with GPCRs.

Combined evaluation of plasma B-type natriuretic peptide and urinary liver-type fatty acid-binding protein/creatinine ratio is related to worsening renal function in patients undergoing elective percutaneous coronary intervention.

BACKGROUND: There are few reports on the significance for the combined evaluation of blood humoral factors and urinary biomarkers in terms of worsening renal function (WRF) after coronary angiography (CAG)/percutaneous coronary arterial intervention (PCI). METHOD AND RESULTS: Urinary liver type-fatty acid-binding protein (L-FABP), neutrophil gelatinase associated lipocalin (NGAL), and adrenomedullin (AM) were measured less than 24 h before and 3 h, 6 h, 1 day, and 2 days after CAG/PCI. WRF was defined as a > 20% decrease in the estimated GFR. WRF occurred in seven of 100 patients and the increase in L-FABP/creatinine (Cr) at 1 day after CAG/PCI was significantly higher in the WRF group than in the non-WRF group. Plasma B-type natriuretic peptide (BNP) before CAG/PCI and L-FABP/Cr at 1 day after CAG/PCI were independent predictors for WRF. The areas under the receiver-operating characteristic curves were as follows: 0.760 for BNP before CAG/PCI, 0.731 for L-FABP/Cr at 1 day after CAG/PCI, and 0.892 for BNP and L-FABP/Cr. Urinary AM levels after PCI/CAG were negatively correlated only to serum potassium levels. Gene expressions of AM and AM-receptor were detectable in renal tubule epithelial cells. AM increased intracellular second messenger levels in a dose-dependent manner. CONCLUSIONS: Our results suggest that combined evaluation of plasma BNP and urinary L-FABP/Cr is useful as a predictor of renal dysfunction in CAG/PCI patients.

Neuromedin U suppresses prolactin secretion via dopamine neurons of the arcuate nucleus.

Neuromedin U (NMU) has a precursor that contains one additional peptide consisting of 33 or 36 amino acid residues. Recently, we identified this second peptide from rat brain and designated it neuromedin U precursor-related peptide (NURP), showing it to stimulate prolactin release from the pituitary when injected via the intracerebroventricular (icv) route. Here, we examined whether NMU, like NURP, also stimulates prolactin release. Unlike NURP, icv injection of NMU significantly decreased the secretion of prolactin from the pituitary. This suppression of prolactin release by NMU was observed in hyper-prolactin states such as lactation, stress, pseudopregnancy, domperidone (dopamine antagonist) administration, and icv injection of NURP. Immunohistochemical analysis revealed that icv injection of NMU induced cFos expression in dopaminergic neurons of the arcuate nucleus, but not the substantia nigra. Mice with double knockout of NMU and neuromedin S (NMS), the latter also binding to NMU receptors, showed a significant increase of the plasma prolactin level after domperidone treatment relative to wild-type mice. These results suggest that NMU and NURP may play important reciprocal roles in physiological prolactin secretion.

Transnasal Delivery of the Peptide Agonist Specific to Neuromedin-U Receptor 2 to the Brain for the Treatment of Obesity.

Obesity and metabolic syndrome are threats to the health of large population worldwide as they are associated with high mortality, mainly linked to cardiovascular diseases. Recently, CPN-116 (CPN), which is an agonist peptide specific to neuromedin-U receptor 2 (NMUR2) that is expressed predominantly in the brain, has been developed as a new therapeutic candidate for the treatment of obesity and metabolic syndrome. However, treatment with CPN poses a challenge due to the limited delivery of CPN to the brain. Recent studies have clarified that the direct anatomical connection of the nasal cavity with brain allows delivery of several drugs to the brain. In this study, we confirm the nasal cavity as a promising CPN delivery route to the brain for the treatment of obesity and metabolic syndrome. According to the pharmacokinetic study, the clearance of CPN from the blood was very rapid with a half-life of 3 min. In vitro study on its stability in the serum and cerebrospinal fluid (CSF) indicates that CPN was more stable in the CSF than in the blood. The concentration of CPN in the brain was higher after nasal administration, despite its lower concentrations in the plasma than that after intravenous administration. The study on its pharmacological potency suggests the effective suppression of increased body weight in mice in a dose-dependent manner due to the direct activation of NMUR2 by CPN. This results from the higher concentration of corticosterone in blood after nasal administration of CPN as compared to nasal application of saline. In conclusion, the above findings indicate that the nasal cavity is a promising CPN delivery route to the brain to treat obesity and metabolic syndrome.

Design and synthesis of peptidic partial agonists of human neuromedin U receptor 1 with enhanced serum stability.

Neuromedin U (NMU) activates two receptors (NMUR1 and NMUR2) and is a promising candidate for development of drugs to combat obesity. Previously, we obtained hexapeptides as selective full NMUR agonists. Development of a partial agonist which mildly activates receptors is an effective strategy which lead to an understanding of the functions of NMU receptors. In 2014, we reported hexapeptide 3 (CPN-124) as an NMUR1-selective partial agonist but its selectivity and serum stability were unsatisfactory. Herein, we report the development of a hexapeptide-type partial agonist (8, CPN-223) based on a peptide (3) but with higher NMUR1-selectivity and enhanced serum stability. A structure-activity relationship study of synthetic pentapeptide derivatives suggested that a hexapeptide is a minimum structure consistent with both good NMUR1-selective agonistic activity and serum stability.

A chemically stable peptide agonist to neuromedin U receptor type 2.

Neuromedin U (NMU) is a peptide with appetite suppressive activity and other physiological activities via activation of the NMU receptors NMUR1 and NMUR2. In 2014, we reported the first NMUR2 selective agonist, 3-cyclohexylpropionyl-Leu-Leu-Dap-Pro-Arg-Asn-NH2 (CPN-116). However, we found that CPN-116 in phosphate buffer is unstable because of Nα-to-Nß acyl migration at the Dap residue. In this study, the chemical stability of CPN-116 was evaluated under various conditions, and it was found to be relatively stable in buffers such as HEPES and MES. We also performed a structure-activity relationship study to obtain an NMUR2-selective agonist with improved chemical stability. Consequently, CPN-219 bearing a Dab residue in place of Dap emerged as a next-generation hexapeptidic NMUR2 agonist.

Identification and signaling characterization of four urotensin II receptor subtypes in the western clawed frog, Xenopus tropicalis.

Urotensin II (UII) is involved, via the UII receptor (UTR), in many physiological and pathological processes, including vasoconstriction, locomotion, osmoregulation, immune response, and metabolic syndrome. In silico studies have revealed the presence of four or five distinct UTR (UTR1-UTR5) gene sequences in nonmammalian vertebrates. However, the functionality of these receptor subtypes and their associations to signaling pathways are unclear. In this study, full-length cDNAs encoding four distinct UTR subtypes (UTR1, UTR3, UTR4, and UTR5) were isolated from the western clawed frog (Xenopus tropicalis). In functional analyses, homologous Xenopus UII stimulation of cells expressing UTR1 or UTR5 induced intracellular calcoum mobilization and phosphorylation of extracellular signal-regulated kinase 1/2. Cells expressing UTR3 or UTR4 did not show this response. Furthermore, UII induced the phosphorylation of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) through the UII-UTR1/5 system. However, intracellular cAMP accumulation was not observed, suggesting that UII-induced CREB phosphorylation is caused by a signaling pathway different from that involving Gs protein. In contrast, the administration of UII to cells increased the phosphorylation of guanine nucleotide exchange factor-H1 (GEF-H1) and myosin light chain 2 (MLC2) in all UTR subtypes. These results define four distinct UTR functional subtypes and are consistent with the molecular evolution of UTR subtypes in vertebrates. Further understanding of signaling properties associated with UTR subtypes may help in clarifying the functional roles associated with UII-UTR interactions in nonmammalian vertebrates.

Purification and identification of native forms of goldfish neuromedin U from brain and gut.

Neuromedin U (NMU) plays important roles in energy homeostasis in rodents and birds. Previously, our group has isolated four cDNAs encoding precursor proteins of NMU from the goldfish brain and gut, and it was assumed that these transcripts are produced by alternative splicing. We have also demonstrated that intracerebroventricular (ICV) injection of putative goldfish NMU inhibits food intake. However, as native goldfish NMU has not yet been identified, we attempted to purify it from goldfish brain and gut extracts. To assess NMU activity in fractions at each purification step, we measured changes in the intracellular concentrations of Ca2+ using HEK293 cells expressing goldfish NMU-R1 or -R2. We isolated a 25-amino-acid peptide (NMU-25) from the brain and gut and found that its primary structure is similar to that of mammalian NMU. Another 21-amino-acid peptide (NMU-21) was purified from the brain, but not from the gut. Furthermore, a 9-amino-acid peptide (NMU-9) identical to the C-terminus of NMU-21 and -25 was also isolated from the brain and gut. Treatment with synthetic NMU-9, -21 and -25 dose-dependently increased the intracellular Ca2+ concentration in mammalian cells expressing goldfish NMU-R1 and -R2. We also examined the effect of ICV-administered synthetic goldfish NMUs on goldfish food intake. NMU-25 inhibited food intake to the same degree as NMU-21. However, the inhibitory effect of NMU-9 was slightly weaker than those of NMU-21 and -25. These results indicate that several molecular forms of NMU exist in the goldfish brain and gut, and that all of them play physiological roles via NMU-R1 and NMU-R2.

Maternal biomarkers for fetal heart failure in fetuses with congenital heart defects or arrhythmias.

BACKGROUND: Diagnosis of fetal heart failure depends primarily on fetal ultrasonography assessment. Our recent study demonstrated that plasma natriuretic peptide levels in umbilical cord blood were correlated with the severity of heart failure in fetuses with congenital heart defects or arrhythmias. However, percutaneous umbilical blood sampling is an invasive procedure, and therefore, less or noninvasive biomarkers reflecting fetal heart failure are required. OBJECTIVE: The aim of this study was to investigate the possibility of whether maternal serum biomarkers can diagnose fetal heart failure in fetuses with congenital heart defects or arrhythmias. STUDY DESIGN: This exploratory cross-sectional study was conducted at a tertiary pediatric cardiac center. A total of 50 singletons with fetal congenital heart defects or arrhythmias and 50 controls who were registered in the National Cerebral and Cardiovascular Center Biobank from 2013 to 2016 were included. Maternal serum samples obtained during the third trimester were analyzed for 2 hormones and 36 cytokines using the Bio-Plex Pro Human Cancer Biomarker panels 1 and 2. We comprehensively analyzed the association between maternal serum biomarkers and ultrasonography findings or fetal arrhythmia status. Fetal heart failure was defined as a cardiovascular profile score ≤7. RESULTS: Of 37 fetuses with congenital heart defects, heart failure was found in 1 case of tricuspid valve dysplasia with moderate tricuspid regurgitation. Of 13 fetuses with arrhythmias, 5 had heart failure at 28-33 weeks of gestation. Maternal serum cytokine and hormone concentrations were compared between patients with and without fetal heart failure at 28-33 weeks of gestation (n = 6 and n = 61, respectively). Sixty-one fetuses without heart failure consisted of 10 with congenital heart defect, 6 with arrhythmia, and 45 controls. Maternal serum concentrations of tumor necrosis factor-α, interleukin-6, soluble Fas ligand, transforming growth factor-α, and vascular endothelial growth factor-D were significantly higher when fetuses had heart failure than when they did not (P < .05), whereas maternal serum concentrations of heparin-binding epidermal growth factor-like growth factor were significantly lower when fetuses had heart failure than when they did not (P < .05). Multivariate analysis showed that maternal serum concentrations of tumor necrosis factor-α, vascular endothelial growth factor-D, and heparin-binding epidermal growth factor-like growth factor were independently associated with fetal heart failure. The cutoff values were as follows: tumor necrosis factor-α, 68 pg/mL (sensitivity of 50.0%, specificity of 93.4%, positive likelihood ratio of 7.6, negative likelihood ratio of 0.5); vascular endothelial growth factor-D, 1156 pg/mL (sensitivity of 50.0%, specificity of 93.4%, positive likelihood ratio of 7.6, negative likelihood ratio of 0.5); and heparin-binding epidermal growth factor-like growth factor, 90 pg/mL (sensitivity of 83.3%, specificity of 83.6%, positive likelihood ratio of 5.1, negative likelihood ratio of 0.2). The combination of these 3 cytokines showed sensitivity of 100%, specificity of 80.3%, positive likelihood ratio of 5.1, and negative likelihood ratio of 0. In the absence of fetal heart failure, concentrations of all maternal serum cytokines and hormones were similar in cases of fetal congenital heart defects and controls, while maternal serum soluble CD40 ligand concentrations were increased only in fetal arrhythmias. CONCLUSION: Maternal serum concentrations of tumor necrosis factor-α, vascular endothelial growth factor-D, and heparin-binding epidermal growth factor-like growth factor were associated with fetal heart failure.

Chemotherapy can promote liver metastasis by enhancing metastatic niche formation in mice.

BACKGROUND: Some chemotherapeutic agents have been reported to promote lung metastasis. However, there have been no reports regarding chemotherapy-induced liver metastasis. We hypothesized that chemotherapy might also enhance liver metastasis. The present study aimed to create a chemotherapy-enhanced liver metastasis mouse model and investigate its mechanism. MATERIALS AND METHODS: Mice were pretreated with cisplatin, vincristine, or saline by intraperitoneal injection. Next, B16F10 mouse melanoma cells and BE(2)-C human neuroblastoma cells were injected into the spleens of C57BL/6 and BALB/c nu/nu mice, respectively, to induce experimental liver metastasis, and the number of liver nodules was determined. We also analyzed the effect of chemotherapy on changes of the liver tissue regarding representative metastasis-promoting factors using real-time quantitative polymerase chain reaction and immunohistochemical and histological analysis. RESULTS: Cisplatin increased the number of nodules by 4.7-fold in the B16F10 liver metastasis model. Vincristine increased the number of nodules by 3.8-fold in the BE(2)-C liver metastasis model. Cisplatin increased mRNA levels of matrix-metalloproteinase (MMP)-2 and periostin, while vincristine increased MMP-9 and S100A8/9 levels in liver tissues. Cisplatin induced fibrosis, whereas vincristine induced neutrophil recruitment in liver tissues according to histological and immunohistochemical analysis. CONCLUSIONS: We concluded that cisplatin or vincristine could enhance liver metastasis of mouse melanoma cells or human neuroblastoma cells, respectively. In addition, the mRNA expression of MMP-2 and periostin, or MMP-9 and S100A8/9 is increased by cisplatin or vincristine pretreatment, possibly resulting in fibrosis or neutrophil recruitment, respectively. These niche factors might be associated with increased liver metastasis.

Amniotic Fluid Natriuretic Peptide Levels in Fetuses With Congenital Heart Defects or Arrhythmias.

BACKGROUND: We have previously demonstrated that umbilical cord plasma natriuretic peptide (NP) levels reflect the severity of heart failure (HF) in fetuses with congenital heart defects (CHD). The aim of this study was to evaluate the significance of amniotic fluid (AF) NP levels in the assessment of HF in fetuses with CHD or arrhythmia. Methods and Results: This was a prospective observational study at a tertiary pediatric cardiac center. A total of 95 singletons with CHD or arrhythmia, and 96 controls from 2012 to 2015 were analyzed. AF concentrations of atrial NP (ANP), B-type NP (BNP) and N-terminal pro-B-type NP (NT-proBNP) at birth were compared with ultrasonographic assessment of fetal HF using the cardiovascular profile (CVP) score. Multivariate analysis showed that a CVP score ≤5 and preterm birth are independently associated with high AF NT-proBNP levels. AF NT-proBNP levels of fetuses with CHD or arrhythmia inversely correlated with CVP score (P for trend <0.01). In contrast, AF concentrations of ANP and BNP were extremely low, and it was difficult to assess the degree of fetal HF based on them. CONCLUSIONS: AF NT-proBNP concentrations increase in stepwise fashion with the severity of HF in fetuses with CHD or arrhythmia; it was the optimal NP for assessing the fetal HF.

CCM2 and PAK4 act downstream of atrial natriuretic peptide signaling to promote cell spreading.

Atrial natriuretic peptide (ANP) is a cardiac hormone released by the atrium in response to stretching forces. Via its receptor, guanylyl cyclase-A (GC-A), ANP maintains cardiovascular homeostasis by exerting diuretic, natriuretic, and hypotensive effects mediated, in part, by endothelial cells. Both in vivo and in vitro, ANP enhances endothelial barrier function by reducing RhoA activity and reorganizing the actin cytoskeleton. We established mouse endothelial cells that stably express GC-A and used them to analyze the molecular mechanisms responsible for actin reorganization. Stimulation by ANP resulted in phosphorylation of myosin light chain (MLC) and promotion of cell spreading. p21-activated kinase 4 (PAK4) and cerebral cavernous malformations 2 (CCM2), a scaffold protein involved in a cerebrovascular disease, were required for the phosphorylation of MLC and promotion of cell spreading by ANP. Finally, in addition to the GC domain, the kinase homology domain of GC-A was also required for ANP/GC-A signaling. Our results indicate that CCM2 and PAK4 are important downstream mediators of ANP/GC-A signaling involved in cell spreading, an important initial step in the enhancement of endothelial barrier function.

Atrial natriuretic peptide prevents cancer metastasis through vascular endothelial cells.

Most patients suffering from cancer die of metastatic disease. Surgical removal of solid tumors is performed as an initial attempt to cure patients; however, surgery is often accompanied with trauma, which can promote early recurrence by provoking detachment of tumor cells into the blood stream or inducing systemic inflammation or both. We have previously reported that administration of atrial natriuretic peptide (ANP) during the perioperative period reduces inflammatory response and has a prophylactic effect on postoperative cardiopulmonary complications in lung cancer surgery. Here we demonstrate that cancer recurrence after curative surgery was significantly lower in ANP-treated patients than in control patients (surgery alone). ANP is known to bind specifically to NPR1 [also called guanylyl cyclase-A (GC-A) receptor]. In mouse models, we found that metastasis of GC-A-nonexpressing tumor cells (i.e., B16 mouse melanoma cells) to the lung was increased in vascular endothelium-specific GC-A knockout mice and decreased in vascular endothelium-specific GC-A transgenic mice compared with control mice. We examined the effect of ANP on tumor metastasis in mice treated with lipopolysaccharide, which mimics systemic inflammation induced by surgical stress. ANP inhibited the adhesion of cancer cells to pulmonary arterial and micro-vascular endothelial cells by suppressing the E-selectin expression that is promoted by inflammation. These results suggest that ANP prevents cancer metastasis by inhibiting the adhesion of tumor cells to inflamed endothelial cells.

Neuromedin U precursor-related peptide (NURP) exerts neuromedin U-like sympathetic nerve action in the rat.

It has been suggested that novel peptide that is produced from the neuromedin U (NMU) precursor may exist, as this precursor contains multiple consensus sequences for proteolytic processing. Recently, we identified two mature novel peptides comprising 33 and 36 residues in the rat brain, which were designated neuromedin U precursor-related peptide (NURP) 33 and 36. In the present study, we compared the roles of NURP33 and 36 with that of NMU, as neither activates the NMU receptors. Immunoreactivity for NMU and NURPs was widely present in the central nervous system and showed a similar distribution. Intracerebroventricular (icv) injection of NURP33 in rats increased locomotor activity, energy expenditure, heart rate and back surface temperature (BS-T), similarly to NMU or NURP36. NMU treatment reduced food intake, but NURP33 did not. Pretreatment with the ß3 blocker, SR59230A, and the cyclooxygenase blocker, indomethacin, inhibited the NURP33- or NMU-induced increase of BS-T. In addition, icv injection of NURP33 or NMU increased the expression of mRNA for cyclooxygenase 2 in the hypothalamus and for uncoupling protein 1 in the brown adipose tissue. These results suggest that although NURP33 and 36 do not activate the NMU receptors, they might exert NMU-like sympathetic nerve action in the brain.

Neuromedin U suppresses glucose-stimulated insulin secretion in pancreatic ß cells.

Neuromedin U (NMU), a highly conserved peptide in mammals, is implicated in energy homeostasis and glycemic control, and may also be involved in the regulation of adipoinsular axis function. However, the role of NMU in regulating insulin secretion has not been clearly established. In this study, we investigated the role of NMU in the regulation of insulin secretion both in vitro and in vivo. We found that NMU and NMU receptor (NMUR) 1 were expressed in mouse islets and ß cell-derived MIN6-K8 cells. In mice, NMU suppressed glucose-stimulated insulin secretion (GSIS) both in vitro and in vivo. Additionally, an NMUR1 agonist inhibited GSIS in both MIN6-K8 cells and mice islets. Moreover, NMU attenuated intracellular Ca2+ influx in MIN6-K8 cells, potentially causing a decrease in insulin secretion. siNmu-transfected MIN6-K8 cells showed elevated GSIS. Treatment with anti-NMU IgG increased GSIS in isolated mouse pancreatic islets. These results suggested that NMU can act directly on ß cells through NMUR1 in an autocrine or paracrine fashion to suppress insulin secretion. Collectively, our results highlight the crucial role of NMU in suppressing pancreatic insulin secretion, and may improve our understanding of glucose homeostasis.

Atrial natriuretic peptide protects against bleomycin-induced pulmonary fibrosis via vascular endothelial cells in mice : ANP for pulmonary fibrosis.

BACKGROUND: Pulmonary fibrosis is a life-threatening disease characterized by progressive dyspnea and worsening pulmonary function. Atrial natriuretic peptide (ANP), a heart-derived secretory peptide used clinically in Japan for the treatment of acute heart failure, exerts a wide range of protective effects on various organs, including the heart, blood vessels, kidneys, and lungs. Its therapeutic properties are characterized by anti-inflammatory and anti-fibrotic activities mediated by the guanylyl cyclase-A (GC-A) receptor. We hypothesized that ANP would have anti-fibrotic and anti-inflammatory effects on bleomycin (BLM)-induced pulmonary fibrosis in mice. METHODS: Mice were divided into three groups: normal control, BLM with vehicle, and BLM with ANP. ANP (0.5 µg/kg/min via osmotic-pump, subcutaneously) or vehicle administration was started before BLM administration (1 mg/kg) and continued until the mice were sacrificed. At 7 or 21 days after BLM administration, fibrotic changes and infiltration of inflammatory cells in the lungs were assessed based on histological findings and analysis of bronchoalveolar lavage fluid. In addition, fibrosis and inflammation induced by BLM were evaluated in vascular endothelium-specific GC-A overexpressed mice. Finally, attenuation of transforming growth factor-ß (TGF-ß) signaling by ANP was studied using immortalized mouse endothelial cells stably expressing GC-A receptor. RESULTS: ANP significantly decreased lung fibrotic area and infiltration of inflammatory cells in lungs after BLM administration. Furthermore, similar effects of ANP were observed in vascular endothelium-specific GC-A overexpressed mice. In cultured mouse endothelial cells, ANP reduced phosphorylation of Smad2 after TGF-ß stimulation. CONCLUSIONS: ANP exerts protective effects on BLM-induced pulmonary fibrosis via vascular endothelial cells.