Correlation between mRNA levels and functional role of α1-adrenoceptor subtypes in arteries: evidence of α1L as a functional isoform of the α1A-adrenoceptor

2005 ◽  
Vol 289 (5) ◽  
pp. H1923-H1932 ◽  
Author(s):  
Daniel Martí ◽  
Raquel Miquel ◽  
Khalid Ziani ◽  
Regina Gisbert ◽  
M. Dolores Ivorra ◽  
...  
Keyword(s):  
Mesenteric Artery ◽  
Mesenteric Arteries ◽  
Main Role ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Response Curves ◽  
Inhibitory Potency ◽  
Adrenoceptor Antagonist ◽  
Relevant Role

The mRNA levels for the three α1-adrenoceptor subtypes, α1A, α1B, and α1D, were quantified by real-time RT-PCR in arteries from Wistar rats. The α1D-adrenoceptor was prominent in both aorta (79.0%) and mesenteric artery (68.7%), α1A predominated in tail (61.7%) and small mesenteric artery (73.3%), and both α1A- and α1D-subtypes were expressed at similar levels in iliac artery. The mRNA levels of the α1B-subtype were a minority in all vessels (1.7–11.1%). Concentration-response curves of contraction in response to phenylephrine or relaxation in response to α1-adrenoceptor antagonists on maximal sustained contraction induced by phenylephrine were constructed from control vessels and vessels pretreated with 100 μmol/l chloroethylclonidine (CEC) for 30 min. The significant decrease in the phenylephrine potency observed after CEC treatment together with the inhibitory potency displayed by 8-{2-[4-(2-methoxyphenyl)-1-piperazinyl]-8-azaspiro ( 4 , 5 ) decane-7-dionedihydrochloride} (BMY-7378, an α1D-adrenoceptor antagonist) confirm the relevant role of α1D-adrenoceptors in aorta and iliac and proximal mesenteric arteries. The potency of 5-methylurapidil (an α1A-adrenoceptor antagonist) and the changes in the potency of both BMY-7378 and 5-methylurapidil after CEC treatment provided evidence of a mixed population of α1A- and α1D-adrenoceptors in iliac and distal mesenteric arteries. The low potency of prazosin (pIC50 < 9) as well as the high 5-methylurapidil potency in tail and small mesenteric arteries suggest the main role of α1A/α1L-adrenoceptors with minor participation of the α1D-subtype. The mRNA levels and CEC treatment corroborated this pattern and confirmed that the α1L-adrenoceptor could be a functional isoform of the α1A-subtype.

scholarly journals Quantitative assessment of CYP11B1 and CYP11B2 expression in aldosterone-producing adenomas

2002 ◽  
pp. 795-802 ◽  
Author(s):  
F Fallo ◽  
V Pezzi ◽  
L Barzon ◽  
P Mulatero ◽  
F Veglio ◽  
...  
Keyword(s):  
Real Time ◽  
Secretory Activity ◽  
Zona Glomerulosa ◽  
Zona Fasciculata ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Pathophysiological Role ◽  
Aldosterone Production

BACKGROUND: The presence and pathophysiological role of CYP11B1 (11beta-hydroxylase) gene in the zona glomerulosa of human adrenal cortex is still controversial. METHODS: In order to specifically quantify CYP11B1, CYP11B2 (aldosterone synthase) and CYP17(17alpha-hydroxylase) mRNA levels, we developed a real-time RT-PCR assay and examined the expression in a series of adrenal tIssues, including six normal adrenals from patients adrenalectomized for renal cancer and twelve aldosterone-producing adenomas (APA) from patients with primary aldosteronism. RESULTS: CYP11B1 mRNA levels were clearly detected in normal adrenals, which comprised both zona glomerulosa and fasciculata/reticularis cells, but were also measured at a lower range (P<0.05) in APA. The levels of CYP11B2 mRNA were lower (P<0.005) in normal adrenals than in APA. CYP17 mRNAlevels were similar in normal adrenals and in APA. In patients with APA, CYP11B2 and CYP11B1 mRNA levels were not correlated either with basal aldosterone or with the change from basal aldosterone in response to posture or to dexamethasone. No correlation between CYP11B1 mRNA or CYP11B2 mRNA and the percentage of zona fasciculata-like cells was observed in APA. CONCLUSIONS: Real-time RT-PCR can be reliably used to quantify CYP11B1 and CYP11B2 mRNA levels in adrenal tIssues. Expression of CYP11B1 in hyperfunctioning zona glomerulosa suggests an additional formation of corticosterone via 11beta-hydroxylase, providing further substrate for aldosterone biosynthesis. CYP11B1 and CYP11B2 mRNA levels in APA are not related to the in vivo secretory activity of glomerulosa cells, where post-transcriptional factors might ultimately regulate aldosterone production.

scholarly journals Role of NO in arterial vascular function of intertidal fish (Girella laevifrons) and marine fish (Isacia conceptionis)

2016 ◽  
Vol 76 (2) ◽  
pp. 500-505
Author(s):  
F. A. Moraga ◽  
N. Urriola-Urriola
Keyword(s):  
Nitric Oxide ◽  
Sodium Nitroprusside ◽  
Marine Fish ◽  
Vascular Function ◽  
Isometric Tension ◽  
Mesenteric Arteries ◽  
Response Curves ◽  
Intertidal Fish ◽  
Branchial Arteries

Abstract Previous studies performed in intertidal fish (Girella laevifrons),as well as marine fish (Isacia conceptionis), showed that acetylcholine (ACh) produced contractions mediated by cyclooxygenases that were dependent on the area and potency of contraction in several arterial vessels. Given that the role of nitric oxide is poorly understood in fish, the objective of our study was to evaluate the role of nitric oxide in branchial afferent (ABA), branchial efferent (ABE), dorsal (DA) and mesenteric (MA) arterial vessels from both Girella laevifrons and Isacia conceptionis. We studied afferent and efferent branchial, dorsal and mesenteric arteries that were dissected from 6 juvenile specimens. Isometric tension studies were done using dose response curves (DRC) for Ach (10–13 to 10–3 M) and blockade with L-NAME (10–5 M), and DRC for sodium nitroprusside (SNP, a donor of NO). L-NAME produced an attenuation of the contractile response in the dorsal, afferent and efferent branchial arteries and a potentiation of the contraction in the MA. SNP caused 70% dilation in the mesenteric artery and 40% in the dorsal artery. Our results suggest that Ach promotes precarious dilatation in MA mediated by NO; data that is supported by the use of sodium nitroprusside. In contrast, in the vessels DA, ABA and EBA our results support that the pathway Ach-NO-relaxation is absent in both species.

Role of T-type calcium channels in myogenic tone of skeletal muscle resistance arteries

2002 ◽  
Vol 283 (6) ◽  
pp. H2239-H2243 ◽  
Author(s):  
Ed VanBavel ◽  
Oana Sorop ◽  
Ditte Andreasen ◽  
Martin Pfaffendorf ◽  
Boye L. Jensen
Keyword(s):  
Calcium Channels ◽  
Southern Blotting ◽  
Myogenic Tone ◽  
Cremaster Muscle ◽  
Resistance Arteries ◽  
Rt Pcr ◽  
Response Curves ◽  
Potent Effect ◽  
Channel Expression

T-type calcium channels may be involved in the maintenance of myogenic tone. We tested their role in isolated rat cremaster arterioles obtained after CO2anesthesia and decapitation. Total RNA was analyzed by RT-PCR and Southern blotting for calcium channel expression. We observed expression of voltage-operated calcium (CaV) channels CaV3.1 (T-type), CaV3.2 (T-type), and CaV1.2 (L-type) in cremaster arterioles ( n= 3 rats). Amplification products were observed only in the presence of reverse transcriptase and cDNA. Concentration-response curves of the relatively specific L-type blocker verapamil and the relatively specific T-type blockers mibefradil and nickel were made on cannulated vessels with either myogenic tone (75 mmHg) or a similar level of constriction induced by 30 mM K+ at 35 mmHg. Mibefradil and nickel were, respectively, 162-fold and 300-fold more potent in inhibiting myogenic tone compared with K+-induced constriction [log(IC50, M): mibefradil, basal −7.3 ± 0.2 ( n = 9) and K+ −5.1 ± 0.1 ( n = 5); nickel, basal −4.1 ± 0.2 ( n = 5) and K+ −1.6 ± 0.5 ( n = 5); means ± SE]. Verapamil had a 17-fold more potent effect [log(IC50, M): basal −6.6 ± 0.1 ( n = 5); K+ −5.4 ± 0.3 ( n = 4); all log(IC50) P < 0.05, basal vs. K+]. These data suggest that T-type calcium channels are expressed and involved in maintenance of myogenic tone in rat cremaster muscle arterioles.

Modulatory role of a constitutively active population of α1D-adrenoceptors in conductance arteries

2002 ◽  
Vol 282 (2) ◽  
pp. H475-H481 ◽  
Author(s):  
Khalid Ziani ◽  
Regina Gisbert ◽  
Maria Antonia Noguera ◽  
Maria Dolores Ivorra ◽  
Pilar D'Ocon
Keyword(s):  
Mesenteric Artery ◽  
Functional Role ◽  
Mesenteric Arteries ◽  
The Other ◽  
Active Population ◽  
Abrupt Changes ◽  
Resting Tone ◽  
Constitutively Active

A constitutively active population of α1D-adrenoceptors in iliac and proximal, distal, and small mesenteric rat arteries was studied. The increase in resting tone (IRT) that evidences it was observed only in iliac and proximal mesenteric and was inhibited by prazosin (pIC50 = 9.57), 5-methylurapidil (pIC50 = 7.61), and BMY 7378 (pIC50 = 8.77). Chloroethylchlonidine (100 μmol/l) did not affect IRT, but when added before the other antagonists it blocked their effect. The potency shown by BMY 7378 confirms the α1D-subtype as responsible for IRT. BMY 7378 displayed greater inhibition of adrenergic responses in iliac (pIC50 = 7.57 ± 0.11) and proximal mesenteric arteries (pIC50 = 8.05 ± 0.2) than in distal (pIC50 = 6.94 ± 0.13) or small mesenteric arteries (pIC50 = 6.30 ± 0.14), which confirms the functional role of the α1D-adrenoceptor in iliac and proximal mesenteric arteries. This subtype prevents abrupt changes in iliac and proximal mesenteric artery caliber when the agonist disappears, and this modulatory role is evidenced by the slower decay in the response to norepinephrine after removal.

scholarly journals Interleukin-1 Receptors Are Differentially Expressed in Normal and Psoriatic T Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Attila Bebes ◽  
Ferenc Kovács-Sólyom ◽  
Judit Prihoda ◽  
Róbert Kui ◽  
Lajos Kemény ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Interleukin 1 ◽  
Effector T Cells ◽  
Receptor Expression ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Healthy Control

This study was carried out to examine the possible role of interleukin-1 (IL-1) in the functional insufficiency of regulatory T cells in psoriasis, by comparing the expression of IL-1 receptors on healthy control and psoriatic T cells. Patients with moderate-to-severe chronic plaque psoriasis and healthy volunteers, matched in age and sex, were selected for all experiments. CD4+CD25−effector and CD4+CD25+CD127lowregulatory T cells were separated and used for the experiments. Expression of the mRNA of IL-1 receptors (IL-1R1, IL-1R2, and sIL-1R2) was determined by quantitative real-time RT-PCR. Cell surface IL-1 receptor expression was assessed by flow cytometry. Relative expression of the signal transmitting IL-1 receptor type 1 (IL-1R1) mRNA is higher in resting psoriatic effector and regulatory T cells, and activation induces higher IL-1R1 protein expression in psoriatic T cells than in healthy cells. Psoriatic regulatory and effector T cells express increased mRNA levels of the decoy IL-1 receptors (IL-1R2 and sIL-1R2) upon activation compared to healthy counterparts. Psoriatic T cells release slightly more sIL-1R2 into their surrounding than healthy T cells. In conclusion, changes in the expression of IL-1 receptors in psoriatic regulatory and effector T cells could contribute to the pathogenesis of psoriasis.

scholarly journals Vasorelaxant Effect Induced by the Essential Oil of Ocotea duckei Vattimo Leaves and Its Main Constituent, Trans-caryophyllene, in Rat Mesenteric Artery

2020 ◽  
pp. 31-39
Author(s):  
Tays Amanda Felisberto Gonçalves ◽  
Renildo Moura da Cunha ◽  
Dionatas Ulises de Oliveira Meneguetti ◽  
Marcio Roberto Viana Santos ◽  
José Maria Barbosa- Filho ◽  
...  
Keyword(s):  
Essential Oil ◽  
Mesenteric Artery ◽  
Calcium Release ◽  
Mesenteric Arteries ◽  
Dependent Manner ◽  
Main Constituent ◽  
Response Curves ◽  
Vasorelaxant Effect ◽  
Rat Mesenteric Artery ◽  
Isolated Rat

Aims: To evaluate the vasorelaxant effect induced by the essential oil of the leaves of O. duckei Vattimo (ODEO) and its main constituent, trans-caryophyllene, in rat superior mesenteric arteries. Methodology: Isolated rat superior mesenteric rings were suspended by cotton threads for isometric tension recordings in Tyrode’s solution at 37ºC, gassed with 95% O2 and 5% CO2 and different ODEO concentrations (0.1-300 μg/mL) or trans-caryophyllene (1-1000 μg/mL) were added cumulatively to the organ baths. Results: Vasorelaxant effect induced by the essential oil of Ocotea duckei leaves (ODEO) and its main constituent, trans-caryophyllene (60.54 %), was evaluated in this work. In intact isolated rat superior mesenteric rings ODEO (0.1-300 μg/mL, n=6) induced concentration-dependent relaxation of tonus induced by phenylephrine (10 µM) or K+-depolarizing solution (KCl 80 mM) (IC50=31±5, 5±0.4 µg/mL, respectively, n=6). The relaxations of phenylephrine-induced contractions were not significantly attenuated after removal of the vascular endothelium (IC50=25±5 µg/mL). ODEO antagonized the concentration-response curves to CaCl2 (10-6-3x10-2 M) and Bay K 8644 (10-10-3x10-6 M). Furthermore, in nominally without calcium solution, ODEO significantly inhibited, in a concentration-dependent manner, transient contractions induced by 10 µM phenylephrine or 20 µM caffeine. Trans-caryophyllene induced vasorelaxations, however, this effect was 18.6 times less potent when compared to ODEO-induced vasorelaxations. Conclusion: The relaxant effect induced by ODEO in rat superior mesenteric artery rings is endothelium-independent and seems to be related to both, inhibition of Ca2+ influx through L-type voltage-gated Ca2+-channels sensitive to dihydropyridines and inhibition of the calcium release from intracellular IP3-and caffeine-sensitive stores.

scholarly journals Interaction of Perivascular Adipose Tissue and Sympathetic Nerves in Arteries From Normotensive and Hypertensive Rats

2016 ◽  
pp. S391-S399 ◽  
Author(s):  
J. TÖRÖK ◽  
A. ZEMANČÍKOVÁ ◽  
Z. KOCIANOVÁ
Keyword(s):  
Adipose Tissue ◽  
Electrical Stimulation ◽  
Mesenteric Artery ◽  
Sympathetic Nerves ◽  
Mesenteric Arteries ◽  
Inhibitory Influence ◽  
Hypertensive Rats ◽  
Response Curves ◽  
Perivascular Adipose Tissue ◽  
Endogenous Noradrenaline

The inhibitory action of perivascular adipose tissue (PVAT) in modulation of arterial contraction has been recently recognized and contrasted with the prohypertensive effect of obesity in humans. In this study we demonstrated that PVAT might have opposing effect on sympatho-adrenergic contractions in different rat conduit arteries. In superior mesenteric artery isolated from normotensive Wistar-Kyoto rats (WKY), PVAT exhibited inhibitory influence on the contractions to exogenous noradrenaline as well as to endogenous noradrenaline released from arterial sympathetic nerves during transmural electrical stimulation or after application of tyramine. In contrast, the abdominal aorta with intact PVAT responded with larger contractions to transmural electrical stimulation and tyramine when compared to the aorta after removing PVAT; the responses to noradrenaline were similar in both. This indicates that PVAT may contain additional sources of endogenous noradrenaline which could be responsible for the main difference in the modulatory effect of PVAT on adrenergic contractions between abdominal aortas and superior mesenteric arteries. In spontaneously hypertensive rats (SHR), the anticontractile effect of PVAT in mesenteric arteries was reduced, and the removal of PVAT completely eliminated the difference in the dose-response curves to exogenous noradrenaline between SHR and WKY. These results suggest that in mesenteric artery isolated from SHR, the impaired anticontractile influence of PVAT might significantly contribute to its increased sensitivity to adrenergic stimuli.

scholarly journals Lisinopril Alters Contribution of Nitric Oxide and KCa Channels to Vasodilatation in Small Mesenteric Arteries of Spontaneously Hypertensive Rats

2015 ◽  
pp. 39-49 ◽  
Author(s):  
S. ALBARWANI ◽  
S. AL-SIYABI ◽  
I. AL-HUSSEINI ◽  
A. AL-ISMAIL ◽  
I. AL-LAWATI ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Spontaneously Hypertensive Rats ◽  
Mesenteric Arteries ◽  
Bkca Channel ◽  
Hypertensive Rats ◽  
Response Curves ◽  
Spontaneously Hypertensive ◽  
Kca Channels ◽  
Reserve System

To investigate lisinopril effect on the contribution of nitric oxide (NO) and KCa channels to acetylcholine (ACh)-induced relaxation in isolated mesenteric arteries of spontaneously hypertensive rats (SHRs). Third branch mesenteric arteries isolated from lisinopril treated SHR rats (20 mg/kg/day for ten weeks, SHR-T) or untreated (SHR-UT) or normotensive WKY rats were mounted on tension myograph and ACh concentration-response curves were obtained. Westernblotting of eNOS and KCa channels was performed. ACh-induced relaxations were similar in all groups while L-NMMA and indomethacin caused significant rightward shift only in SHR-T group. Apamin and TRAM-34 (SKCa and IKCa channels blockers, respectively) significantly attenuated ACh-induced maximal relaxation by similar magnitude in vessels from all three groups. In the presence of L-NMMA, indomethacin, apamin and TRAM-34 further attenuated ACh-induced relaxation only in SHR-T. Furthermore, lisinopril treatment increased expression of eNOS, SKCa and BKCa proteins. Lisinopril treatment increased expression of eNOS, SKCa, BKCa channel proteins and increased the contribution of NO to ACh-mediated relaxation. This increased role of NO was apparent only when EDHF component was blocked by inhibiting SKCa and IKCa channels. Such may suggest that in mesenteric arteries, non-EDHF component functions act as a reserve system to provide compensatory vasodilatation if (and when) hyperpolarization that is mediated by SKCa and IKCa channels is reduced.

255 GENE EXPRESSION OF Cox 5a, 5b, AND 6b1 AND THEIR ROLES IN PRE-IMPLANTATION OF MOUSE EMBRYOS

2006 ◽  
Vol 18 (2) ◽  
pp. 235
Author(s):  
S.-E. Lee ◽  
X.-Y. Li ◽  
X.-S. Cui ◽  
N.-H. Kim
Keyword(s):  
Rna Interference ◽  
Mrna Expression ◽  
Real Time ◽  
Oocyte Maturation ◽  
Blastocyst Stage ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Target Mrna ◽  
Protein Levels

Despite clear evidence of regulation of mitochondrial respiration by nuclear encoded genes, cytochrome oxidase (Cox), little information is available on their expression and functional roles during early embryonic development. To examine the role of Cox in oocyte maturation and embryogenesis, we first characterized mRNA and protein levels of nuclear encoded genes, Cox 5a, 5b, and 6b1, in mouse oocytes and during early embryogenesis, using real-time RT-PCR and immunocytochemistry. We then examined the possible role of these genes in oocyte maturation and pre-implantation development using RNA interference analysis. The relative abundances of Cox 5a, 5b, and 6b1 transcripts was measured by real time RT-PCR. After normalization by comparison to histone H2a mRNA levels, the mRNA expression of Cox 5a, 5b, and 6b1 were found to be considerable in mature oocytes and zygotes, but reduced slightly in 2-cell embryos. From the 2-cell to the blastocyst stage, mRNA expression is dependent on the number of blastomeres, as expression increases only gradually with development. Immunocytochemical studies revealed that Cox 5a, 5b, and 6b1 proteins were expressed in all blastomeres of the blastocyst. Injection of Cox 5a, 5b, or 6b1 siRNA into GV stage oocytes decreased expression of the target mRNA specifically, while not affecting the expression of mRNAs for the other subunits in mature oocytes. Similarly, each siRNA injection into zygotes specifically reduced target mRNA expression at the 2-cell, morula and blastocyst stages (P < 0.05). Silencing of mRNA expression by RNA interference (siRNA) did not inhibit oocyte maturation or developmental events up to the morula and blastocyst stages. The expression level of mtDNA9, as well as overall levels of mitochondrial mRNAs, was not different following injection of siRNA for Cox 5a, 5b, or 6b1. However, it is evident that the number of mitochondria in siRNA treated blastocysts was greatly reduced, and they appeared to be morphologically abnormal. Significantly higher apoptosis and lower cell numbers were observed in siRNA treated blastocysts. Real time RT PCR revealed that silencing of Cox 5a, 5b, and 6b1 decreased mRNA and protein levels of E-cadherin. These results suggest that the Cox subunits, Cox 5a, 5b, and 6b1, play an important role in mitochondrial function during pre-implantation development. This work was funded by a grant from the National Research Laboratory Program in Korea.

90 ENDOGENOUS MODIFICATIONS OF AURORA B AND AURORA C KINASE EXPRESSION IN MOUSE OOCYTES AND EARLY EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 146
Author(s):  
C. A. Lima ◽  
V. Huntress ◽  
E. W. Overström
Keyword(s):  
Protein Localization ◽  
Somatic Cells ◽  
Early Embryo ◽  
Vital Role ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Mouse Oocytes ◽  
Reverse Transcription Pcr ◽  
Early Embryos

The Aurora (Aur) proteins are a family of serine/threonine kinases that play fundamental roles in controlling M-phase progression. Previous reports have shown that AurB is vital for proper completion of karyokinesis and cytokinesis in somatic cells. The role of AurC in somatic cells has been found to be much less significant, whereas it appears to play an important role in spermatogenesis. The role of these Aur proteins is not well characterized in mouse oocytes and early embryos. The objective of this study was to assess changes of AurB and AurC mRNA and protein expression in mouse oocytes and early embryos as development progresses through the activation of the zygotic genome. Oocytes and embryos were collected from the oviducts of hormone-stimulated CF-1 mice. After culturing for varying amounts of time, cumulus-denuded samples were either fixed for immunofluorescence microscopy studies or lysed for analysis of mRNA levels through the use of reverse transcription-PCR (RT-PCR). Samples were processed for immunofluorescence using markers of spindle morphology (tubulin) and AurB. Analysis of relative levels of AurB and AurC mRNA were assessed by RT-PCR methods. Marked differences were observed in the localization of AurB when unfertilized oocytes or prezygotic genome activation (ZGA) embryos were compared with post-ZGA samples. There was no evidence of AurB protein localized to the mitotic spindle or resultant midbody in oocyte and early embryo samples. Embryos fixed post-ZGA demonstrated AurB localization, as is conventionally found in somatic cells. The AurB protein was found co-localized with DNA in metaphase stage blastomeres and associated with the midbody in blastomeres near completion of cytokinesis. Relative levels of AurB mRNA were not found to be significantly different when pre- and post-ZGA samples were compared. A significant decrease in relative levels of AurC mRNA was observed in fertilized pre-ZGA samples when compared with unfertilized oocyte counterparts. These observations demonstrate significant differences in the status of AurB and AurC mRNA levels and protein localization in mouse oocytes and early embryos when compared with somatic cells. Given earlier reports showing the vital role of AurC in spermatogenesis, the elevated levels of AurC mRNA observed in prefertilization oocytes may be indicative of a similar role of AurC during oogenesis. Elucidating temporal and localization details of Aur expression is vital to gaining further understanding of cell-cycle regulation in oogenesis and early embryogenesis.

Sign in / Sign up

Export Citation Format

Share Document