Yuka Fukuda

Endothelin is a potent mitogen for rat vascular smooth muscle cells

The effect of endothelin (ET), a novel endothelium-derived vasoconstrictive peptide, on DNA synthesis was studied in cultured rat vascular smooth muscle cells (VSMC). ET stimulated incorporation of [3H]thymidine into DNA of the quiescent VSMC in a dose-dependent manner; the approximate half-maximal and maximal stimulation for DNA synthesis was induced with 2 x 10(-10) M and 10(-9) M, respectively. The stimulatory effect by ET on DNA synthesis was completely inhibited by the calcium channel blocker nifedipine. ET combined with epidermal growth factor and transforming growth factor-alpha, but not with platelet-derived growth factor, had synergistic effects. These data indicate that ET is a potent mitogen as well as a constrictor for VSMC, suggesting its potential role in the development of vascular disease.

Endothelin is a potent secretagogue for atrial natriuretic peptide in cultured rat atrial myocytes.

Using cultured neonatal rat atrial cardiocytes, we have studied the effect of synthetic porcine endothelin (pET), a novel potent vasoconstrictor isolated from endothelial cells, on the release of immunoreactive (IR) rat atrial natriuretic peptide (rANP). pET stimulated IR-rANP secretion in a dose-dependent manner (10(-10)-10(-7) M) with an approximate half-maximally stimulatory dose of 2 x 10(-10) M. The pET-induced IR-rANP secretion was attenuated by Ca2+-channel blocker nicardipine, but no further stimulation was induced when combined with a Ca2+-channel agonist BAY-K 8644. pET in combination with tetradecanoyl-phorbol-acetate resulted in a synergistic effect on IR-rANP secretion. These data suggest that ET may play as an endogenous secretagogue for rANP by modulating Ca2+ influx through the voltage-dependent Ca2+-channels in atrial cardiocytes.

Specific receptor for endothelin in cultured rat cardiocytes

Specific binding sites for the endothelium-derived vasoconstrictor endothelin (ET) and its effect on cytosolic free Ca2+ concentrations [( Ca2+]i) were studied in a primary culture of cardiocytes from neonatal rats. Binding studies using 125I-labeled-porcine ET as a radioligand revealed the presence of a single class of high-affinity binding sites for ET in cardiocytes with an apparent Kd of 6-9 x 10(-10) M and a Bmax of 50,000-80,000 sites/cell. Neither various vasoconstrictors nor Ca2+-channel blockers affected the binding. Pretreatment with ET substantially reduced the total number of ET receptors without changing their affinity. ET dose-dependently increased [Ca2+]i in fura-2-loaded cardiocytes. These data indicate that cardiocytes have specific ET receptors that are controlled by a down-regulation mechanism, and that ET induces a receptor-mediated increase in [Ca2+]i in cardiocytes.

Endothelin stimulates accumulations for cellular atrial natriuretic peptide and its messenger RNA in rat cardiocytes

The effect of endothelin-1 (ET-1) on secretion and synthesis of rat atrial natriuretic peptide (rANP) as well as its mRNA levels was studied in primary cultures of neonatal rat atrial cardiocytes. ET-1 dose-dependently (10(-10)-10(-7) M) increased media and cellular rANP-like immunoreactivity as well as its cytoplasmic mRNA levels in rat cardiocytes during 24 hrs incubation. These results suggest that ET-1 directly stimulates expression of the rANP gene in cardiocytes, thereby leading to enhanced synthesis and secretion of rANP.

Regional distribution of immunoreactive endothelin in rats

By use of a specific radioimmunoassay for endothelin (ET), the regional distribution of ET-like immunoreactivity (LI) was studied in rats. The antiserum used cross-reacted equally with synthetic porcine and rat ET. Significant amounts of ET-LI are detectable not only in aorta, but also in kidney, lung, heart, liver and central nervous system. Gel chromatography of the tissue extracts revealed size heterogeneity of ET-LI; one major component eluting close to, but slightly larger than standard rat ET and the other minor component with a larger molecular weight. These data indicate that ET-LI is widely distributed throughout the various rat tissues, suggesting its possible involvement in a variety of organ functions.

Effects of protein kinase inhibitors on growth factor-stimulated DNA synthesis in cultured rat vascular smooth muscle cells

The effects of H-7 and ML-9, inhibitors of protein kinase C and myosin light-chain kinase, respectively, on DNA synthesis stimulated by platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) were studied in cultured rat vascular smooth muscle cells (VSMC). H-7 and ML-9 significantly inhibited PDGF-stimulated DNA synthesis in lower concentrations, while both compounds were only effective in inhibiting EGF-induced DNA synthesis in higher concentrations. These data suggest that protein kinase C and myosin light-chain kinase activated by PDGF play a more important role in cell proliferation of VSMC than EGF.

Changes in vasopressin, atrial natriuretic factor, and water homeostasis in the early stage of bronchopulmonary dysplasia.

ABSTRACT: Arginine vasopressin (AVP), atrial natriuretic factor, and water balance were examined in the infants with or without hronchopulmonary dysplasia (BPD) during the first 4 wk of life. Fourteen premature infants, nine in the early stage of BPD secondary to respiratory distress syndrome (BPD infants) and five healthy low birthwt infants (LBW infants), were the subjects of this study. The water and sodium balance, renal function, and plasma AVP and atrial natriuretic factor concentrations were determined during the first 4 wk of life. Plasma AVP and atrial natriuretic factor levels of BPD infants at the 4th wk of life were higher than those of LBW infants at the corresponding age. Urine osmolality was higher and free water clearance was lower in BPD infants at the 4th wk of life when compared with each parameter in LBW infants, respectively. PaCo2 of BPD infants at the 4th wk of life was more elevated than that of LBW infants. These results suggest that elevated plasma AVP level may be related with pulmonary abnormalities and that atrial natriuretic factor may hence compensate the water retention resulted from the functionally activated AVP in the early stage of BPD.

Reliability of Urinary N-Acetyl-β-D-Glucosaminidase as an Indicator of Renal Tubular Damage in Neonates

Urinary N-acetyl-beta-D-glucosaminidase (NAG) was determined in infants during the first year of life to clarify its evolutional change and the reliability of this enzyme as a marker of renal tubular damage in the newborn period. During the first 2 weeks of life, there was no difference in the urinary levels between preterm (gestational age of 33-36 weeks) and full-term infants. The urinary NAG index (NAG is expressed as the ratio of the enzyme activity to milligrams of creatinine) during the first month of life was significantly elevated when compared with that of any other succeeding period and gradually decreased during the first year of life. The index in newborn infants on tobramycin was significantly elevated and maintained a higher value during the 5 days after withdrawal of the medication. Infants with asphyxia had a higher urinary NAG index during the first week of life. These data suggest that the urinary NAG index is a useful indicator of renal tubular damage during the newborn period.

Effects of aldosterone and atrial natriuretic peptide on water and electrolyte homeostasis of sick neonates

ABSTRACT: To clarify fluid homeostasis in neonates, 27 sick neonates (51 had respiratory distress syndrome, 12 had hyperbilirubinemia, and nine were low birth wt infants) were studied on 13 to 15 days of age. The infants were stabilized and required neither ventilation nor intravenous fluids at the time of study. All infants were breast fed, and their sodium intake ranged from 1.7 to 2.9 (2.2 ± 0.3) mEq/kg/d. Gestational age ranged from 30 to 39 (34 ± 3.4) wk and birth wt ranged from 1080 to 3280 (2,070 ± 672) g. Creatinine clearance (Ccr), fractional excretion of sodium (FENa), fluid and electrolyte balance, plasma aldosterone and human atrial natriuretic peptide (ANP) concentrations were determined and analyzed according to conceptional age (CA) to clarify their maturational changes. None of the infants had a negative sodium balance or hyponatremia. Ccr remained almost constant until the 36th wk of CA, after which an abrupt increase occurred. FENa and plasma aldosterone level of infants less than the 32nd wk of CA were high (0.75 ± 0.33%, 2868 ± 1153 pg/ml, respectively) when compared with those in infants of the 33 to 34th wk of CA (0.47 ± 0.12%, 1,663, ± 488 pg/ml, respectively). There was a negative correlation between FENa and plasma aldosterone level in all cases (p<0.01). Plasma ANP level of all infants were higher than those of healthy adults (40 ± 20 pg/ml). A good correlation was observed between Ccr and plasma ANP level in infants less than 36th wk of CA. These results show that the high plasma aldosterone concentrations possibly act to enhance the reabsorptive capacity of the distal nephron for sodium during development under conditions of low sodium intake and suggest that ANP may be released to control the extracellular space in premature infants with a limited Ccr.

Maturational Changes and Origin of Urinary Human Epidermal Growth Factor in the Neonatal Period

In order to clarify the characteristics of urinary human epidermal growth factor (hEGF) excretion in the newborn period, we examined hEGF in the 41 newborn infants; 12 healthy preterm infants (group A), 10 healthy full-term infants (group B), 12 full-term infants with neonatal asphyxia (group C), 7 full-term infants treated with tobramycin (group D) during the first week of life. Renal function tests, i.e. creatinine clearance (Ccr), and N-acetyl-beta-D-glucosaminidase (NAG) as a parameter of renal tubular damage, were examined in 29 full-term infants. Urinary hEGF excretion showed a linear increase with gestational age. During the first week of life, urinary hEGF excretion in group B increased with age. However, urinary hEGF excretion in group A remained at the constant level through the study period. Ccr in group C was significantly decreased through the study period when compared with that in group B. NAG indices in group C during the first week of life and those in group D on days 5-7 of life were more elevated than those in group B. Urinary hEGF excretion in group C on days 4-7 of life and that in group D on day 7 of life were significantly decreased when compared with that in group B. These results suggest that urinary hEGF excretion is related to the maturation and that the source of urinary hEGF may be renal tubular cells.