SIÇANLARDA DOCA-TUZ MODELİ İLE OLUSTURULAN HİPERTANSİYONA BAĞLI DAMAR ENDOTEL DİSFONKSİYONU PARAMETRELERİ VE ROCK EKSPRESYONU VE AKTİVASYONUNUN KARSILASTIRILMASI

Bu çalısmanın amacı; DOCA-Tuz hipertansiyon modeli uygulanan sıçanlarda hipertansiyona bağlı damar endotel disfonksiyon parametreleri ile ROCK ekspresyonu ve aktivasyonunun incelenmesidir. Çalısmada 4 hafta kontrol, 4 hafta DOCA-tuz, 12 hafta kontrol ve 12 hafta DOCA-tuz olmak üzere dört farklı deney grubu kullanılmıstır. DOCA gruplarına haftada iki kez 15 mg/kg DOCA enjeksiyonu ek olarak %1 NaCl ve %0.2 KCl içeren içme suyu uygulanmıstır. Bu süre boyunca tüm grupların kan basıncı değerleri haftada bir kez ölçülmüstür. 4 ve 12 hafta sonunda deney gruplarındaki sıçanlardan izole edilen torasik aort halkaları izole organ deneylerinde kullanılmıstır. Fenilefrinle kastırılmıs dokularda 10-5-10-8M asetilkolin konsantrasyon-cevap eğrileri çıkarılmıstır. Ayrıca izole edilen aortların bir kısmında western blot yöntemi ile ROCK I, ROCK II ve pMYPT ekspresyonları incelenmistir. Deneylerin sonucunda DOCA uygulanan gruplardaki kan basıncı değerlerinin 1. haftadan baslayarak anlamlı olarak arttığı bulunmustur. Asetilkolin gevseme yanıtlarından elde edilen EC50 değerleri, 4 ve 12 hafta DOCA-tuz gruplarında kontrole göre anlamlı olarak değismezken, Emaks değerlerinin anlamlı olarak azaldığı tespit edilmistir. ROCK I, ROCK II ve pMYPT ekspresyonları hipertansiyona bağlı olarak değismemistir. ROCK I ekspresyonu hem kontrol hem de DOCA gruplarında yasa bağlı olarak anlamlı olarak artmıstır. ROCK II ekspresyonu ise 12 hafta kontrol grubunda, 4 hafta kontrol grubuna göre anlamlı olarak azalmıstır. Son olarak pMYPT ekspresyonunda, 4 hafta ve 12 hafta DOCA-tuz uygulanmıs gruplar arasındaki fark istatistiksel olarak anlamlı bulunmustur. Bu sonuçlar, DOCA-tuz uygulamasıyla indüklenen hipertansiyona bağlı olarak endotel disfonksiyonu gelistiğini, fakat disfonksiyonun derecesinin hipertansiyonun süresine göre değismediğini ve ROCK ekspresyonu ve aktivasyonunun DOCA-tuz hipertansiyon modeline aracılık etmediğini düsündürmekle birlikte, kas kasılmasında rol oynayan ROCK proteinlerinin ekspresyon ve aktivasyonunun yasa bağlı olarak değisiklik gösterdiğine isaret etmektedir

Investıgatıon Of Hypertensıon Induced Endoplasmıc Retıculum stress In Vessels

Hipertansiyon oldukça karmaşık bir patogeneze sahiptir ve moleküler etiyolojisi henüz tam olarak aydınlatılamamıştır. Katlanmamış/yanlış katlanmış proteinlerin endoplazmik retikulumda birikmesi durumu olan endoplazmik retikulum stresi (ERS) oldukça çeşitli sayıda hücresel strese cevaben oluşabilir. ERS yolağı kardiyovasküler hastalıklar için potansiyel bir hedef olarak tanımlanmıştır. Bu çalışmada ERS inhibisyonunun, damarlarda hipertansiyonda oluşan değişimler üzerindeki etkileri araştırılmıştır. Hipertansiyon unilateral nefrektomi, deoksikortikosteron asetat (DOCA) enjeksiyonu (20 mg/kg, haftada iki kez) ve içme sularına % 1 NaCl ile % 0,2 KCl eklenmesi yoluyla erkek Wistar albino sıçanlarda (8 haftalık) oluşturulmuştur. Hayvanlar 12 hafta boyunca kan basınçları ölçülerek takip edilmiştir. Bir ERS inhibitörü olan tauroursodeoksikolik asit (TUDCA) (150 mg/kg/gün, ip), sıçanlara son dört hafta boyunca uygulanmıştır. Deney sonunda, hayvanlardan izole edilen torasik aortlarda kasılma ve gevşeme cevapları kaydedilmiştir. Total antioksidan kapasite ve bir nitrik oksit (NO) göstergesi olan nitrit seviyeleri spektrofotometrik olarak plazma örneklerinde ölçülmüştür. SERCA2, IP3R, PERK, p- PERK, Bax ve Bcl2 ekspresyon seviyeleri Western blot analizi ile torasik aort örneklerinde incelenmiştir. Bu çalışmada DOCA-tuz hipertansiyon modelinde ERS inhibisyonunun, sistolik kan basıncını azalttığı; endotel işlevlerinde iyileşme sağladığı; kasılma cevaplarını değiştirdiği; plazma NO seviyelerini artırdığı; aort SERCA2 ve IP3R ekspresyonlarını geri çevirdiği; aortta Bcl2 ekspresyonunu artırdığı gösterilmiştir. Elde edilen bulgular, ERS inhibisyonunun hipertansiyondaki iyileştirici etkisinde, NO seviyelerini artırması, SERCA2 ve IP3R ekspresyonlarını düzeltmesi ve antiapoptotik etkileri ile birlikte damar işlevlerini düzeltici etkisinin rol oynayabileceğini düşündürmektedir. Bu sonuçlar, ERS inhibisyonunun hipertansiyon üzerindeki olumlu etkisinin moleküler mekanizmalarına ışık tutmakta ve yeni tedaviler için potansiyel bir hedef öne sürmektedir.Hypertension has complex pathogenesis therefore the molecular etiology remains poorly elucidated. Endoplasmic reticulum stress (ERS) which is a condition of the unfolded/misfolded protein accumulation in endoplasmic reticulum, may occur in response to a wide variety of cellular stress. ERS pathway has been defined as a potential target for cardiovascular disease. In the present study, effects of ERS inhibition on hypertensioninduced alterations in vessel were investigated. Hypertension was induced through unilateral nephrectomy, deoxycorticosterone acetate (DOCA) injection (20 mg/kg, twice a week) and 1 % NaCl with 0,2 % KCI by adding to drinking water in male Wistar albino rats (8 weeks old). Animals were monitored by measuring blood pressures for 12 weeks. An ERS inhibitor tauroursodeoxycolic acid (TUDCA) (150 mg/kg/day, i.p.), was administrated to animals for last four weeks. At the end of the experiment, contraction and relaxation responses of isolated thoracic aortas were recorded. Total antioxidant capacity and nitrite levels, as an indicator of nitric oxide (NO), were measured spectrophotometrically in plasma samples. The expression levels of SERCA2, IP3R, PERK, p-PERK, Bax and Bcl2 were examined by Western blot analysis in thoracic aorta. In the present study, it has been shown that ERS inhibition in DOCA-salt induced hypertension, reduced systolic blood pressure; improved endothelial dysfunction; changed vascular contractions; enhanced plasma NO levels; restored aortic expressions of SERCA2 and IP3R; increased aortic Bcl2 expression. These obtained data suggest that the beneficial effects of ERS inhibition on hypertension may be related to the protection of vessel functions together with increasing NO levels, restoring of SERCA2 and IP3R expressions and antiapoptotic effects. These results provide a new insight into the molecular mechanisms of the beneficial effects of ERS inhibition on hypertension and suggest a potential target for novel therapies

Hypertension-induced cardiac impairment is reversed by the inhibition of endoplasmic reticulum stress

WOS:000488634600001PubMed:31579948Objectives Endoplasmic reticulum stress (ERS) has been shown to play a crucial role in the pathogenesis of hypertension. However, the role and mechanisms of ERS on hypertension-induced cardiac functional and morphological changes remain unclear. In this study, the effect of ERS inhibition with tauroursodeoxycholic acid (TUDCA) on hypertension-induced cardiac remodelling was examined. Methods Hypertension was induced by deoxycorticosterone-acetate (DOCA) and salt administration in uni-nephrectomized rats for 12 weeks. TUDCA was administered for the last four weeks. Rhythmic activity and contractions of the right atrium and left papillary muscle (LPM) were recorded. In the left ventricle, the expression of various proteins was examined and histopathological evaluation was performed. Key findings Hypertension-induced increments in systolic blood pressure and ventricular contractions were reversed by TUDCA. In the hypertensive heart, while expressions of glucose-regulated protein-78 (GRP78), phospho-dsRNA-activated protein kinase-like ER kinase (p-PERK), sarcoplasmic reticulum Ca-ATPase-2 (SERCA2), matrix metalloproteinase-2 (MMP-2) and nuclear NF-kappa B p65 increased; Bcl-2 (B-cell lymphoma-2) expression decreased and the altered levels of all these markers were restored by TUDCA. In the microscopic examination, TUDCA treatment attenuated hypertension-stimulated cardiac inflammation and fibrosis. Conclusions These results suggest that ERS inhibition may ameliorate cardiac contractility through improving ERS-associated calcium mishandling, apoptosis, inflammation and fibrosis, thereby offering therapeutic potential in hypertension-induced cardiac dysfunction.Ankara University Research FoundationAnkara University [16B0230004]This work was supported by Ankara University Research Foundation [Grant number 16B0230004]

The effects of resveratrol and exercise on age and gender-dependent alterations of vascular functions and biomarkers

WOS:000442386400026PubMed:29908346The purpose of this study was to determine the effects of resveratrol and regular aerobic exercise on vascular functions and biomarkers related to vessel responsiveness in an age and gender-dependent manner. The study used young (3 months) and old (12 months) male and female Wistar albino rats. Resveratrol was given in the drinking water (0.05 mg/ml; approximately 7.5 mg/kg) for 6 weeks.In the exercise group, all rats performed treadmill running at 20 m/min on a 0 degrees incline, 40 min/day, 3 times a week, for 6 weeks. Acetylcholine-induced, endothelium-dependent and sodium nitroprusside-mediated, endothelium-independent relaxations of rat thoracic aorta and blood levels of biomarkers were separately changed by resveratrol intake and exercise-training in an age and gender-dependent manner. Antioxidant enzymes and eNOS expressions in vessels were elevated by resveratrol and exercise. Resveratrol and exercise enhanced gene expressions of non-selective PDE1, 2, 3 and cAMP selective PDE4 but not cGMP selective PDE5 in the aorta. In addition, the aortic mRNA expression of inflammation markers were altered by resveratrol and exercise-training. The results of the study demonstrated that vessel responsiveness and biomarkers related to vascular functions were altered by resveratrol consumption and exercise-training in an age and gender-dependent manner.Ankara University Research FoundationAnkara University [12B3330001]This study was supported by a research grant, 12B3330001 from the Ankara University Research Foundation

The effects of LXR agonist GW3965 on vascular reactivity and inflammation in hypertensive rat aorta

WOS:000448413900034PubMed:30366037Aims: Liver X receptors (LXRs) play an important role in the regulation of cholesterol, fatty acid and glucose metabolisms together with inflammatory processes. In the present study, the effects of LXR agonist GW3965 on vascular reactivity and expression of functional proteins in DOCA-Salt induced hypertension were examined. Main methods: Hypertension was induced through unilateral nephrectomy and deoxycorticosterone-acetate (DOCA) injection (20 mg/kg, twice a week) for 6 weeks in male Wistar albino rats (8 weeks old). An LXR agonist GW3965 (10 mg/kg/day, i.p.) was administered to animals for last seven days. Key findings: GW3965 treatment reduced systolic blood pressures in hypertensive rats. Acetylcholine-induced endothelium-dependent and sodium nitroprusside-induced endothelium-independent vasorelaxations were decreased in hypertensive rats but not affected by GW3965. GW3965 treatment enhanced plasma nitrite levels in normotensive rats. KCl and phenylephrine (Phe)-induced vasocontractions were reduced in hypertensive groups and increased with GW3965 treatment. Decreased sarco/endoplasmic reticulum Ca2+- ATPase2 (SERCA2) expression in the hypertensive aorta was not changed by GW3965 treatment. Expression of inositoltrisphosphate receptor1 (IP3R1) was increased by GW3965 in normotensive animals. The nuclear factor kappaB (NF-kappa B) and tumor necrosis factor alpha (TNF-alpha) expressions were increased in hypertensive rats and reduced by GW3965 treatment. Significance: The results of study indicate that the LXR agonist, GW3965, exhibited a beneficial effect on increased blood pressure and improved hypertension-induced impairment in contractile activity of vessel and inflammatory markers in vascular tissue. Therefore, these effects of LXR agonists on vessel should be taken into account in experimental or therapeutic approaches to hypertension.TUBITAK-3001 projectTurkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [114S170]; Ankara University Research FoundationAnkara University [16B0230004]This study was supported by research grants, from TUBITAK-3001 project (114S170) and Ankara University Research Foundation (16B0230004)

Age- and sex-dependent alteration of functions and epigenetic modifications of vessel and endothelium related biomarkers.

Aging is a main risk factor for development of cardiovascular diseases associated with the impairment of endothelial function in both sexes. In the present study, age-related changes in vascular responsiveness, epigenetic modifications of vessel wall, and blood biomarkers related to endothelial functions were examined in an age- and sex-dependent manner. Acetylcholine (ACh)-induced relaxations of the aorta were decreased in 3-, 6-, and 12-month-old rats compared to those in 1-month-old female rats. In males, maximum relaxations related to ACh were higher in 1- and 6-month-old rats than in 3- and 12-month-old rats. Plasma levels of nitric oxide (NO) and asymmetric dimethylarginine (ADMA) decreased with age in female rats, and total antioxidant capacity (TAC) and hydrogen sulfide (H 2S) levels displayed biphasic alterations. In male rats, plasma levels of NO, TAC, and ADMA decreased with age, and H2S levels increased. Aging also caused a sex-dependent alteration in epigenetic modification of vessels. Expressions of H3K27me2, H3K27me3, H3K36me2, and H3K36me3 were much higher in vessels of 12-month-old female rats compared to those in younger age groups. These results indicate that vascular functions, epigenetic modifications of vessels, and plasma levels of endothelium-related biomarkers are affected by age and sex. These findings could be important for the assessment of vascular status over the course of the life span

Inhibition of endoplasmic reticulum stress protected DOCA-salt hypertension-induced vascular dysfunction

WOS:000462806800005PubMed:30458302Hypertension has complex vascular pathogenesis and therefore the molecular etiology remains poorly elucidated. Endoplasmic reticulum stress (ERS), which is a condition of the unfolded/misfolded protein accumulation in the endoplasmic reticulum, has been defined as a potential target for cardiovascular disease. In the present study, the effects of ERS inhibition on hypertension-induced alterations in the vessels were investigated. In male Wistar albino rats, hypertension was induced through unilateral nephrectomy, deoxycorticosteroneacetate (DOCA) injection (20 mg/kg, twice a week) and 1% NaCl with 0.2% KCI added to drinking water for 12 weeks. An ERS inhibitor, tauroursodeoxycolic acid (TUDCA) (150 mg/kg/day, i.p.), was administered for the final four weeks. ERS inhibition in DOCA-salt induced hypertension was observed to have reduced systolic blood pressure, improved endothelial dysfunction, enhanced plasma nitric oxide (NO) level, reduced protein expressions of phosphorylated-double-stranded RNA-activated protein kinase-like endoplasmic reticulum kinase (pPERK), 78 kDa glucose-regulated protein (GRP78), Inositol trisphosphate receptor1 (IP(3)R1) and Epidermal growth factor receptor (EGFR), increased expressions of endoplasmic reticulum Ca2+-ATPase2 (SERCA2) and B cell lymphoma2 (Bcl2) in vessels. These findings suggest that the beneficial effects of ERS inhibition on hypertension may be related to protection of vessel functions through restoration of endoplasmic reticulum calcium homeostasis, and apoptotic and mitotic pathways.Ankara University Research FoundationAnkara University [16B0230004]This study was supported by a research grant (16B0230004) from the Ankara University Research Foundation. The authors are grateful to Prof. Dr. H. Gurdal and B. Dalkilic for assisting with the Western Blot analysis

Diurnal Temporal Blood H2S Variations Correlate with the Circadian Rhythm of Vascular Contraction and Blood Pressure

Background: It is well known that blood pressure has a circadian rhythm in rat. However, the underlying mechanisms that modulate circadian rhythm of blood pressure have not been fully clarified. The aim of this study was to investigate the probable mechanisms that regulate time dependent variation of blood pressure. In present study, the correlations among the following s:alpha-1 adrenoceptor stimulated aortic contractions, thoracic aortic expression of Rho-kinase ll and myosin phosphatase target subunit-1 and blood biomarkers (nitric oxide, hydrogen sulfide {[}H2S] and total antioxidant capacity) that regulate blood pressure at six different times of the day and night were examined. Materials and Methods: Systolic blood pressure was measured every 4 h during a 24 h period in male albino Wistar rats by tail-cuff plethysmography. At each time point, contraction and relaxation responses of isolated thoracic aortas were recorded. The expression of protein from aortas was determined by western blot method. Nitric oxide, total antioxidant capacity and H2S levels were measured spectrophotometrically in plasma samples. One-way analysis of variance and student t-test was used to determine statistical differences. Results: Rat systolic blood pressure displayed a circadian rhythm, which reached the maximum at 05:00 am and minimum at 09;00 am. Diurnal variation of phenylephrine-induced contractions in the isolated thoracic aorta was also observed. Although, the Rho-kinase inhibitor Y-27632 reduced phenylephrine-induced contractions, the circadian pattern of the contractions did not change. Interestingly, Rho-kinase II and myosin phosphatase target subunit -1 protein expression in the thoracic aorta did not show significant changes throughout the day. Further, plasma levels of nitric oxide and total antioxidant capacity did not vary during the day. However, H2S levels in the systemic circulation showed circadian variation, which was the maximum at 01:00 am and minimum at 05;00 am. Conclusions: These results suggest that, in addition to alpha-1 adrenoceptor sensitivity of vessels, the circadian rhythm of plasma H2S could contribute to diurnal blood pressure variations. This highlights a potential novel experimental and therapeutic approach to blood pressure regulation

Age- and sex-dependent alteration of functions and epigenetic modifications of vessel and endothelium related biomarkers

Aging is a main risk factor for development of cardiovascular diseases associated with the impairment of endothelial function in both sexes. In the present study, age-related changes in vascular responsiveness, epigenetic modifications of vessel wall, and blood biomarkers related to endothelial functions were examined in an age- and sex-dependent manner. Acetylcholine (ACh)-induced relaxations of the aorta were decreased in 3-, 6-, and 12-month-old rats compared to those in 1-month-old female rats. In males, maximum relaxations related to ACh were higher in 1- and 6-month-old rats than in 3- and 12-month-old rats. Plasma levels of nitric oxide (NO) and asymmetric dimethylarginine (ADMA) decreased with age in female rats, and total antioxidant capacity (TAG) and hydrogen sulfide (H2S) levels displayed biphasic alterations. In male rats, plasma levels of NO, TAG, and ADMA decreased with age, and H2S levels increased. Aging also caused a sex-dependent alteration in epigenetic modification of vessels. Expressions of H3K27me2, H3K27me3, H3K36me2, and H3K36me3 were much higher in vessels of 12-month-old female rats compared to those in younger age groups. These results indicate that vascular functions, epigenetic modifications of vessels, and plasma levels of endothelium-related biomarkers are affected by age and sex. These findings could be important for the assessment of vascular status over the course of the life span