-
[show abstract]
[hide abstract]
ABSTRACT: Background. Angiotensin II (Ang II) is involved in the direct inhibition of renin release from juxtaglomerular (JG) cells in the kidney
as the negative feedback loop of the renin-angiotensin system. Ang II also modulates renin release via the sympathetic nervous
system, since the renal sympathetic nerves stimulate renin release, and the interaction of the sympathetic nervous system
with Ang II has been demonstrated to occur at multiple levels.
Methods. Experiments were performed in conscious unrestrained rabbits. Ang II (1.0 ng/kg per min) was infused intravenously for 60
min in renal-denervated (Dx; n = 6) and sham-denervated (Sh; n = 6) rabbits, and plasma renin activity (PRA) was determined. Then the intrarenal administration of the Ang II type-1 receptor
(AT1R) antagonist, losartan, or type-2 receptor (AT2R) antagonist, PD123319 was carried out, during infusion of Ang II or
saline in both Dx and Sh.
Results. PRA was decreased in both Sh (5.9 ± 0.6 to 2.8 ± 0.7 ng/ml per h; n = 6, P < 0.01) and Dx (5.7 ± 0.4 to 3.8 ± 0.6 ng/ml per h; n = 6, P < 0.01) during Ang II infusion. The degree of decrease was significantly less in Dx than in Sh (P < 0.05), indicating that the inhibition of renin release by Ang II is associated with renal nerves. The intrarenal administration
of losartan in Sh significantly decreased PRA produced by Ang II (5.6 ± 0.3 to 4.8 ± 0.3 ng/ml per h; n = 6, P < 0.05) vs. saline vehicle (5.7 ± 0.3 to 2.8 ± 0.2 ng/ml per h; n = 6, P < 0.01) (P < 0.05). Blockade with losartan in Dx significantly increased PRA (5.8 ± 0.4 to 6.6 ± 0.4 ng/ml per h; n = 6, P < 0.05) during the infusion of Ang II. Renin response to the Ang II infusion was not influenced by the intrarenal administration
of PD123319 alone, in either Sh or Dx.
Conclusion. Ang II appears to facilitate sympathetic neurotransmission through the postjunctional AT1R leading to renin release.
Clinical and Experimental Nephrology 04/1999; 3(2):89-95. · 1.37 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: 1. The effect of cicletanine, a novel antihypertensive agent with natriuretic activity, on blood pressure and progression of renal failure in 5–6 nephrectomized spontaneously hypertensive rats with salt loading was examined.2. All nephrectomized rats were randomly assigned to one of four groups and their diet was changed from a normal- to a high-salt (5.5% NaCl) diet for the next 10 weeks. Either 10 or 50 mg/kg per day cicletanine (low- and high-dose cicletanine, respectively) or 10 mg/kg per day trichlormethiazide were administered to rats during this period once a day. During the experimental period, urine volume, urinary excretion of sodium, protein, prostaglandin (PG) E2 and 6-keto-PGF1α and systolic blood pressure (SBP) were measured every 2 weeks.3. Systolic blood pressure was significantly reduced by the administration of trichlormethiazide and the higher dose of cicletanine, but not by the lower dose of cicletanine.4. In contrast with changes to SBP, levels of serum creatin- ine in rats treated with both doses of cicletanine were significantly lower than in controls (0.57±0.12, 0.78±0.12 and 1.68±0.26 mg/dL for high- and low-dose cicletanine and control, respectively).5. Urinary excretion of both PGE2 and 6-keto-PGF1α were significantly increased in groups treated with high and low doses of cicletanine compared with control. In rats treated with trichlormethiazide, PGE2 and 6-keto-PGF1α levels were significantly decreased compared with control.6. In contrast with changes in SBP, marked glomerular sclerosis with hyalinosis found in the control group was not ameliorated by trichlormethiazide treatment. These changes were not observed in rats treated with low- and high-dose cicletanine, particularly those treated with the higher dose of cicletanine.7. These data suggest that administration of cicletanine has a beneficial protective effect regarding the progression of renal failure, regardless of the level of blood pressure, through a direct and/or indirect action on the glomerulus.
Clinical and Experimental Pharmacology and Physiology 02/1999; 26(3):236 - 241. · 1.85 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Interleukin (IL)-1 and IL-4 synergistically stimulate NF-IL6 activity and IL-6 production in human mesangial cells.Background While interleukin (IL)-4 inhibits pro-inflammatory cytokine expression by human monocytes, we have observed that it potentiates IL-6 production by IL-1-activated human mesangial cells (MC). To study the mechanism of this cell-type specific interaction between IL-1 and IL-4 in MC, we examined the effect of both cytokines on the activities of nuclear factor B (NF-B) and nuclear factor IL-6 (NF-IL6), transcription factors that are essential for IL-6 gene expression.
Kidney International 06/1998; 54(1):71-79. · 6.61 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Acute methotrexate toxicity resulting from methotrexate-induced renal fáilue is a medical emergency requiring extracorporeal
removal of methotrexate. The optimum method of methotrexate removal has not yet been established. We report a case of osteosarcoma
with lung metastasis that was treated with high-dose methotrexate as adjuvant chemotherapy. Although no problems occurred
after the first 5 courses of methotrexate, methotrexate-induced renal failure and methotrexate toxicity appeared after the
sixth course. The patient was treated, either with hemoperfusion plus high-flux hemodialysis, or hemoperfusion alone, and
pre- and post-treatment serum methotrexate concentrations were monitored. The reduction in methotrexate by hemoperfusion alone
for 2 hours was 54%, compared to a mean reduction of 59% by hemoperfusion combined with high-flux hemodialysis, for 3 hours.
Rebound increases in methotrexate levels were small (less than 1 μmol/L) with either method. The combination of hemoperfusion
and hemodialysis resulted in good control of volume status, as well as improvement in serum chemistry values.
Clinical and Experimental Nephrology 02/1998; 2(1):75-79. · 1.37 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: OBJECTIVE
Leptin, the obese gene product, is secreted exclusively by adipocytes and is thought to act as a lipostatic signal that regulates body weight homeostasis. We previously reported that thyroid hormone is one of the up-regulating factors of leptin in vitro. T3, at physiological concentrations, stimulates leptin mRNA expression and leptin secretion by 3T3-L1 adipocytes. The aim of this study was to explore the role of thyroid hormone in the regulation of leptin in humans.DESIGN AND PATIENTSA total of 59 non-obese women aged 38.4 ± 1.8 years (mean ± SEM) were studied: 19 patients with hyperthyroidism, 17 patients with hypothyroidism, and 23 normal control subjects. The correlation between serum leptin concentrations and body mass index (BMI) was analyzed, and serum leptin levels were compared among the three groups.MEASUREMENTSSerum leptin concentrations were measured by radioimmunoassay.RESULTSSerum leptin concentrations after logarithmic transformation were correlated significantly (P < 0.05) with BMI in the hyperthyroid (r = 0.46), the hypothyroid (r = 0.84), and normal (r = 0.63) groups. Even though age, body weight, and BMI were similar in all groups, serum leptin levels in the hypothyroid patients (5.30 ± 1.12 μg/l) were significantly (P < 0.05) lower than in the hyperthyroid and normal groups (6.87 ± 0.66 and 6.58 ± 0.68 μg/l, respectively).CONCLUSIONS
These results indicate that thyroid hormone may play an important role in the appropriate secretion of leptin in humans.
Clinical Endocrinology 02/1998; 48(3):299 - 302. · 3.17 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: A full-length cDNA for the rat kidney mitochondrial cytochrome P450 mixed function oxidase, 25-hydroxyvitamin D3-1α-hydroxylase (P4501α), was cloned from a vitamin D-deficient rat kidney cDNA library and subcloned into the mammalian expression
vector pcDNA 3.1(+). When P4501α cDNA was transfected into COS-7 transformed monkey kidney cells, they expressed 25-hydroxyvitamin
D3-1α-hydroxylase activity. The sequence analysis showed that P4501α was of 2,469 bp long and contained an ORF encoding 501
amino acids. The deduced amino acid sequence showed a 53% similarity and 44% identity to the vitamin D3-25-hydroxylase (CYP27), whereas it has 42.6% similarity and 34% identity with the 25-hydroxyvitamin D3-24-hydroxylase (CYP24). Thus, it composes a new subfamily of the CYP27 family. Further, it is more closely related to the
CYP27 than to the CYP24. The expression of P4501α mRNA was greatly increased in the kidney of vitamin D-deficient rats. In
rats with the enhanced renal production of 1α,25-dihydroxyvitamin D3 (rats fed a low Ca diet), P4501α mRNA was greatly increased in the renal proximal convoluted tubules.
Proceedings of the National Academy of Sciences 11/1997; 94(24):12920-12925. · 9.68 Impact Factor
-
Toru Nabika,
Tamiyo Ito,
Kazuhiro Kitada,
Tadao Serikawa,
Tomoji Mashimo,
Florent Soubrier,
Cécile Julier,
Yoichi Ohno, Takao Saruta,
Hisao Tanase,
Junichi Masuda,
Yukio Yamori,
Yasuo Nara
Mammalian Genome 04/1997; 8(3):215-217. · 2.89 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Summary1. We investigated the effects of antihypertensive treatment (8 weeks) with four different agents (trichlormethiazide, atenolol, nicardipine and enalapril) on baroreceptor function in 28 week old spontaneously hypertensive rats (SHR) to measure aortic depressor nerve (ADN) activity.2. Threshold pressure (P th) of ADN activity was elevated and the gain sensitivity of the pressure–-activity curve, as determined by the maximum gain (Gmax) of a logistic function curve, was depressed in untreated SHR compared to those in untreated Wistar-Kyoto (WKY) rats.3. Treatment with the four agents similarly reduced blood pressure in SHR. Each of the four agents induced a decrease in Pth and an increase in Gmax to a similar extent in SHR.4. These findings suggest that antihypertensive therapy in chronic hypertension augments baroreceptor function through the lowering of blood pressure but not through specific pharmacological actions.
Clinical and Experimental Pharmacology and Physiology 10/1995; 22(s1):S67 - S69. · 1.85 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The effects of low-dose endothelin on systemic haemodynamics and vasoactive hormones were examined in conscious dogs. In addition, we examined the effects of endothelin on pressor responses to noradrenaline and angiotensin II and the baroreflex regulation of heart rate in conscious dogs. Continuous infusion of 40 fmol/kg per min endothelin for 40min induced a mild but significant reduction in mean arterial pressure from 89.1 +/- 1.7 to 82.7 +/- 2.0mmHg (P<0.05), associated with decreases in total peripheral resistance 20 min later. A 400 fmol/kg per min dose of endothelin, on the other hand, induced a gradual elevation of mean arterial pressure from 89.2 +/- 2.3 to 96.8 +/- 2.0 mmHg (P<0.05), associated with increases in total peripheral resistance over 30 min. The 40fmol/kg per min dose of endothelin infusion induced a significant reduction in plasma arginine vasopressin (AVP; P<0.05) and elevations of plasma atrial natriuretic peptide (ANP; P<0.05), plasma prostaglandin E2 (PGE2; P<0.05) and plasma 6-keto-prostaglandin F1[alpha] (6-keto-PGF1[alpha]; P<0.05). The 400 fmol/kg per min dose produced elevations of AVP, ANP, PGE2 and 6-keto-PGF1[alpha] (P<0.05). Pressor responses to noradrenaline and angiotensin II were significantly attenuated during continuous infusion of 40 fmol/kg per min endothelin, whereas 400 fmol/kg per min endothelin did not induce any significant changes compared with the control. Furthermore, baroreflex sensitivity was attenuated with 40 fmol/kg per min endothelin but did not show any significant changes at 400 fmol/kg per min. These data demonstrate that low-dose endothelin has effects on haemodynamics and hormonal responses which are the opposite of those of the pharmacological dose employed, suggesting that the earlier hypothesized physiological role of endothelin has now been challenged.
(C) Lippincott-Raven Publishers.
Journal of Hypertension 03/1991; 9(4). · 4.02 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The effects of the calcium entry blocker nifedipine on blood pressure, heart rate, plasma renin activity, aldosterone, noradrenaline and adrenaline were studied in 23 normotensive subjects in the supine and upright positions. Nifedipine, 10 mg administered sublingually, lowered mean blood pressure and increased heart rate, plasma noradrenaline and renin activity without increasing plasma aldosterone in the supine position. The increase in plasma aldosterone in response to upright posture was inhibited by nifedipine, whereas the rise in plasma noradrenaline was augmented. These results suggest that intracellular calcium is important as a regulator of aldosterone secretion as well as of vascular tone in normotensive subjects.
Clinical Endocrinology 04/1986; 24(5):565 - 570. · 3.17 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: To investigate the interaction between endothelin (ET) and the nitric oxide system, we examined the effects of ET-1 and ET-3 on the induction of inducible nitric oxide synthase (iNOS) and guanosine triphosphate cyclohydrolase I (GTP:CHI), the rate-limiting enzyme of de novo synthesis of the cofactor tetrahydrobiopterin (BH4), in rat mesangial cells. ET-1 inhibited the nitrite accumulation induced by a combination of interleukin-1β, tumor necrosis factor-α, and lipo-polysaccharide in a concentration-dependent manner. The inhibitory effect of ET-3 was less potent than that of ET-1. A selective ETA antagonist, BQ-485, and an ETA and ETB antagonist, TAK-044, abolished the inhibitory effects of ET-1, whereas the selective ETB antagonist BQ-788 had no effect on the inhibition produced by ET-1. These observations indicate that ET-1 inhibits cytokine-stimulated nitrite accumulation through the ETA receptor. Western blot analysis showed that the suppression of nitrite accumulation was accompanied by a decrease in iNOS protein. Northern blot analysis showed that ET-1 inhibited the expression of both iNOS and GTP: CHI mRNA. In conclusion, ET-1 inhibits cytokine-stimulated nitric oxide production through the ETA receptor by suppressing the expression of iNOS and GTP: CHI mRNA in rat mesangial cells.
Pharmacology 08/1970; 53(4):241-249. · 1.79 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The role of the renin-angiotensin system in polyuric patients was studied in 5 patients with pituitary diabetes insipidus, 1 patient with nephrogenic diabetes insipidus and 3 patients with psychogenic polydipsia by determining the plasma renin activity and infusing an angiotensin II analog, 1-Sar, 8-Ile angiotensin II. In all patients with diabetes insipidus, plasma renin activity was markedly increased and the blood pressure was reduced in 5 of 6 patients by the administration of the angiotensin II analog. In the other patient, the blood pressure remained unchanged. The plasma renin activity and response of blood pressure to the angiotensin II analog in a patient with nephrogenic diabetes insipidus were not significantly different from there of patients with pituitary diabetes insipidus. On the other hand, in all patients with psychogenic polydipsia, plasma renin activity was normal or low, and the blood pressure increased with the administration of the angiotensin II analog. These results suggest that evaluation of the renin-angiotensin system by determining plasma renin activity and infusing an angiotensin II analog is useful in differentiating between diabetes insipidus and psychogenic polydipsia.
Nephron 08/1970; 32(1):14-17. · 13.26 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Increases in arterial blood pressure cause cumulative changes in tissue structure and function, resulting ultimately in end-organ damage. One of the pathological hallmarks of hypertensive tissue injury is an increase in tissue fibrosis, which leads to reductions in tissue compliance and function. Fibrosis (or sclerosis) occurs as result of marked changes in the amount and composition of the extracellular matrix. This extracellular matrix is a complex mixture of structural proteins and glycoproteins, including collagens, fibronectins, and proteoglycans. Hypertension is known to be associated with increases in the synthesis of extracellular matrix proteins and changes in their degradation. These processes are mediated by several mediators, in particular the renin-angiotensin-aldosterone system. Since these changes play an important role in the formation of vascular sclerosis, cardiac dysfunction, and renal damage, understanding the mechanisms, and finding interventions to prevent or reverse these changes are clinically important. In this review we discuss the alterations in the extracellular matrix during hypertension, as well as the effects of antihypertensive agents in animal models and human patients.
-
[show abstract]
[hide abstract]
ABSTRACT: It is well known that α2-adrenergic agonism inhibits insulin secretion and stimulates glucagon secretion in both animal and human studies. Recently, α2-adrenergic blockers (DG-5128, MK-912, and SL 84.0418) have been studied as antihyperglycemic agents in human subjects. To clarify the action mechanism(s) of these agents, we investigated the effects of α2 agonists and antagonists (10−10 to 10−4 mol/L) and pretreatment by pertussis toxin (PTX) on glucose-stimulated insulin secretion using the hamster insulinoma cell line HiT-T15. The imidazoline-derivative α2-adrenoceptor agonists clonidine and oxymetazoline at concentrations as low as 10−8 mol/L significantly inhibited glucose-stimulated insulin secretion by 63% and 65%, respectively (P < .01 for both). These inhibitory effects were abolished by 20-hour preincubation of these cells with PTX 100 ng/mL. The imidazoline-derivative α2-adrenoceptor antagonist DG-5128 at a concentration of 10−4 mol/L doubled insulin secretion with or without pretreatment by PTX (P < .01 for both). Furthermore, both clonidine and oxymetazoline at a high concentration of 10−4 mol/L stimulated insulin secretion with pretreatment of the cells by PTX (P < .05 for both). These results indicate that glucose-stimulated insulin secretion is inhibited by α2-adrenoceptor agonists through PTX-sensitive G-protein in HIT-T15 cells. It is also suggested that imidazoline compounds at high concentrations directly stimulate insulin secretion.
Metabolism. 46(10):1146-1149.
-
Shin Hisahara,
Takashi Araki,
Fumihiro Sugiyama,
Ken-Ichi Yagami,
Misao Suzuki,
Kuniya Abe,
Ken-Ichi Yamamura,
Jun-ichi Miyazaki,
Takashi Momoi, Takao Saruta,
Claude C.A. Bernard,
Hideyuki Okano,
Masayuki Miura
[show abstract]
[hide abstract]
ABSTRACT: The mechanisms underlying oligodendrocyte (OLG) loss and the precise roles played by OLG death in human demyelinating diseases such as multiple sclerosis (MS), and in the rodent model of MS, experimental autoimmune encephalomyelitis (EAE), remain to be elucidated. To clarify the involvement of OLG death in EAE, we have generated transgenic mice that express the baculovirus anti-apoptotic protein p35 in OLGs through the Cre-loxP system. OLGs from cre/p35 transgenic mice were resistant to tumor necrosis factor–α-, anti-Fas antibody- and interferon–γ-induced cell death. cre/p35 transgenic mice were resistant to EAE induction by immunization with the myelin oligodendrocyte glycoprotein. The numbers of infiltrating T cells and macrophages/microglia in the EAE lesions were significantly reduced, as were the numbers of apoptotic OLGs expressing the activated form of caspase–3. Thus, inhibition of apoptosis in OLGs by p35 expression alleviated the severity of the neurological manifestations observed in autoimmune demyelinating diseases.
-
[show abstract]
[hide abstract]
ABSTRACT: We have examined the effects of sex hormones on calcium-dependent NO production and protein levels of NO synthase in cultured human aortic endothelial cells, which were treated with various doses of 17β-estradiol and testosterone for 8–48 h. Treatment with 17β-estradiol enhanced calcium-dependent NO production, but testosterone had exerted no effect. Western blot using monoclonal anti-human endothelial NO synthase antibody clarified that increased NO production by 17β-estradiol treatment was accompanied by increased NO synthase protein. Our results provide evidence that human endothelial NO synthase can be regulated by estrogens.
FEBS Letters.
-
[show abstract]
[hide abstract]
ABSTRACT: Our purpose was to determine the effect of ovariectomy on intracellular Ca2+ mobilization and platelet aggregation in sodium induced hypertension. At the age of 12 weeks ovariectomy or sham operation was performed in female Dahl-Iwai salt sensitive rats on a 0.3% NaCl diet. Four weeks later we assessed the effects of ovariectomy and an 8% NaCl diet on agonist induced intracellular Ca2+ mobilization in fura-2 loaded platelets and platelet aggregation. Ovariectomy enhanced the increase of systolic blood pressure and heart to body weight ratio on an 8% NaCl diet. However, thrombin evoked intracellular Ca2+ was not correlated with systolic blood pressure (r = −0.338, P = .17), and was lowered by sodium loading and ovariectomy (360 ± 23 to 285 ± 9, 296 ± 10 nmol/L, P < .05). Furthermore, the ionomycin induced intracellular calcium fraction in the absence of external Ca2+ that reflected internal Ca2+ discharge capacity was reduced in ovariectomized rats compared with sham operated rats on an 8% NaCl diet (648 ± 15 v 768 ± 35 nmol/L, P < .05). The internal Ca2+ discharge capacity was inversely correlated with systolic blood pressure (r = −0.506, P = .03). In addition to the decreased internal Ca2+ discharge capacity, intracellular Ca2+-independent platelet aggregation by phorbol 12-myristate 13-acetate, a protein kinase C activator, was significantly enhanced in hypertensive rats. We concluded that ovariectomy enhanced sodium induced hypertension associated with the decreased internal Ca2+ discharge capacity and increased platelet aggregation in Dahl-Iwai salt-sensitive rats.
American Journal of Hypertension.
-
Hirotaka Shibata,
Yayoi Ikeda,
Tokuo Mukai,
Ken-ichirou Morohashi,
Isao Kurihara,
Takashi Ando,
Toshihiko Suzuki,
Sakiko Kobayashi,
Masaru Murai,
Ikuo Saito, Takao Saruta
[show abstract]
[hide abstract]
ABSTRACT: Chicken ovalbumin upstream promoter–transcription factor (COUP–TF), DAX-1, and steroidogenic factor-1 (SF-1) are orphan members of the nuclear hormone receptor superfamily. COUP–TF and DAX-1 have been shown to negatively regulate the transcriptional activity of SF-1, a steroidogenic cell-specific activator of various steroidogenic cytochrome P450 genes. We therefore examined the expression levels and immunolocalization of COUP–TF, DAX-1, and SF-1 in human adrenal gland (NL) and adrenocortical adenomas, and compared the results with CYP17 expression levels and its enzyme activities to study their potential correlation with adrenocortical steroidogenesis. In NL (n = 10), expressions of COUP–TF, DAX-1, and SF-1 were detected in the nuclei of adrenocortical cells, but not in the medulla. In cortisol-producing adenomas causing Cushing syndrome (CS, n = 20), CYP17 expression was upregulated (298 ± 2% vs NL 98 ± 4%), whereas expression levels of both COUP–TFs (COUP–TFI, 52 ± 5% vs NL 98 ± 4%; COUP–TFII, 18 ± 4% vs NL 98 ± 4%) and DAX-1 (42 ± 4% vs NL 100 ± 4%) were reduced. In deoxycorticosterone-producing adenomas (DOC, n = 2), on the other hand, CYP17 expression was extremely reduced (8 and 12% vs NL 98 ± 4%), whereas DAX-1 expression increased markedly (350 and 360% vs NL 100 ± 4%). Expression levels of SF-1 did not differ between NL (100 ± 8%) and CS (106 ± 10%), but its expression appeared to be decreased in DOC (25 and 20%). These results showed CYP17 expression to be upregulated and downregulated in CS and DOC, respectively, in a manner reciprocal to that of its repressors, COUP–TF and/or DAX-1. In summary, the results indicate that co-localization of COUP–TF, DAX-1, and SF-1 in NL was lost in adrenocortical tumors and that these orphan receptors play an important role in the regulation of steroidogenesis in human adrenals.
Molecular Genetics and Metabolism.
-
[show abstract]
[hide abstract]
ABSTRACT: Receptors with a heptahelical structure initiate signal transduction by interacting with specific Gα proteins. The aim of this study was to analyze the ability of type 1 (AT1) and type 2 (AT2) angiotensin receptors to recognize the receptor coupling regions of Gα proteins using our previously described technique (Ikezu, T., Okamoto, T., Komatsuzaki, K., Matsui, T., Martyn, J.A.J., Nishimoto, I., 1996. Negative transactivation of cAMP response element by familial Alzheimer's mutants of APP. EMBO J. 15, 2468–2475; Komatsuzaki, K., Murayama, Y., Giambarella, U., Ogata, E., Seino, S., Nishimoto, I., 1996. A novel system that reports the G-proteins linked to a given receptor: a study of the type 3 somatostatin receptor. FEBS Lett. 406, 165–170). Chimeric Gαs protein constructs, whose receptor binding regions contained sequences from the four major families of Gα proteins (Gαq, Gαi, Gα12, Gαs), were cotransfected with AT1 or AT2 receptors in COS cells, then stimulated with angiotensin II (Ang II). Changes in cellular cAMP were assayed on cell lysates by enzyme immunoassay. In the case of the Gαq family, cotransfection of AT1 with Gα11/Gαs, Gα14/Gαs, Gα16/Gαs, elicited significant increases in cAMP after agonist stimulation. Confirmatory results were found using an independent [35S]GTPγS binding assay. Further examination using chimeric G proteins for Gα12 proteins and Gαi family proteins provided evidence that the AT1 receptor can recognize sequences from Gα12, Gαi1/i2, Gαz, Gαo, while both receptors interacted with Gαi3. These results provide a Gα protein recognition database for both AT1 and AT2 receptors, which may be important for understanding the full spectrum of cellular responses mediated by the hormone Ang II.
Molecular and Cellular Endocrinology.
-
[show abstract]
[hide abstract]
ABSTRACT: Heterogeneity in Madin-Darby canine kidney (MDCK) epithelial cells has been reported, however, its details have not been well described. In the present study, we show that subclones obtained from a MDCK cell line could be divided into two morphologically and biochemically distinct cell types with different hormonal responsiveness. Clones of the first type, motile clones, which had extended and flattened cytoplasm, were devoid of carbonic anhydrase activity. Clones of the second type, nonmotile clones, formed colonies of cuboidal cells and showed carbonic anhydrase activity. Motile clones synthesized cAMP in response to arginine vasopressin, prostaglanin E1, and isoproterenol but not glucagon. In contrast, nonmotile clones responded to all of these hormones. These findings suggest MDCK cells have multiple cellular origins. The motile clones have characteristics similar to the principal cells of the collecting system, whereas the nonmotile clones may be derived from the thick ascending limb or the intercalated cell. Our studies also demonstrate a significant influence of culture condition on MDCK cellular behavior (carbonic anhydrase activity, Na+/K+-ATPase activity and vasopressin responsiveness). Therefore, physiologic and biochemical experiments with MDCK cells must be interpreted with reservations about cellular heterogeneity as well as differences induced by culture conditions.
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research.
