Mechanical stimulation of nitric oxide synthesizing mechanisms in erythrocytes.

ÜLKER P., Sati L., Celik-Ozenci C., Meiselman H. J., Baskurt O. K.

Biorheology, cilt.46, sa.2, ss.121-32, 2009 (SCI-Expanded) identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 46 Sayı: 2
  • Basım Tarihi: 2009
  • Doi Numarası: 10.3233/bir-2009-0532
  • Dergi Adı: Biorheology
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Sayfa Sayıları: ss.121-32
  • Anahtar Kelimeler: Shear stress, red blood cell NOS, nitric oxide, L-NAME, NOS activation, RED-BLOOD-CELLS, ENDOTHELIAL-CELLS, VASCULAR-TONE, SHEAR-STRESS, SYNTHASE, PHOSPHORYLATION, MODULATION, CALCIUM, OXYHEMOGLOBIN, OXYGENATION
  • Akdeniz Üniversitesi Adresli: Evet

Özet

It has been previously demonstrated that red blood cells (RBC) possess functional nitric oxide ( NO) synthesizing mechanisms. RBC are also equipped with variety of intracellular control mechanisms, and respond to mechanical forces and to various biological stimuli by increased release of ATP. Nitric oxide has also been demonstrated to be released from RBC under certain circumstances, and it has been hypothesized that NO synthase ( NOS), which is located in both the RBC membrane and cytoplasm, might be activated by mechanical factors. The present study aimed at investigating NOS activation and NO export induced by mechanical stress applied to RBC in suspension. Heparinized venous blood samples were obtained from healthy, adult volunteers and their hematocrit adjusted to 0.4 l/l. The RBC suspensions were equilibrated at room temperature (22 +/- 2 degrees C) with either room air or made hypoxic (36 mmHg, similar to 70% saturation) using moisturized 100% nitrogen. The samples were then continuously pumped through a glass tube (diameter = 0.06 cm; length = 33 cm) for 30 min using a dual syringe pump to maintain a wall shear stress of 0.5-2 Pa with NO concentrations in the RBC suspensions measured electrochemically. NO concentration significantly increased under the influence of 2 Pa in hypoxic RBC suspensions: 105.0 +/- 14.2 nM to 127.1 +/- 12.0 nM as the peak value at 20 min of perfusion. No increase was observed at lower levels of shear stress. Plasma nitrite/nitrate concentrations were measured in samples obtained at five minute intervals. Application of fluid shear stress to hypoxic RBC suspensions resulted in a significant, time-dependent increase of plasma nitrite/nitrate levels, reaching to 14.7 +/- 1.5 mu M from a control value of 11.2 +/- 1.3 mu M. The presence of the non-specific NOS inhibitor L-NAME (10(-3) M) prevented this increment. Additionally, both eNOS and serine 1177 phosphorylated eNOS immuno-fluorescence staining in RBC cytoplasm were shown to increase in response to applied shear stress. Our results support the hypothesis that RBC NO synthase is activated and that export of NO from RBC is enhanced by mechanical stress.

Mechanical stimulation of nitric oxide synthesizing mechanisms in erythrocytes

By:Ulker, P (Ulker, P.)[ 1 ] ; Sati, L (Sati, L.)[ 2 ] ; Celik-Ozenci, C (Celik-Ozenci, C.)[ 2 ] ; Meiselman, HJ (Meiselman, H. J.)[ 3 ] ; Baskurt, OK (Baskurt, O. K.)[ 1 ]

 

BIORHEOLOGY

 

Volume: 46

Issue: 2

Pages: 121-132

DOI: 10.3233/BIR-2009-0532

Published: 2009

View Journal Information

BIORHEOLOGY

JCR® Category Quartile in Category
BIOPHYSICS Q4
ENGINEERING, BIOMEDICAL Q3
HEMATOLOGY Q4

Data from the 2012 edition of Journal Citation Reports®

Publisher IOS PRESS, NIEUWE HEMWEG 6B, 1013 BG AMSTERDAM, NETHERLANDS

ISSN: 0006-355X

Research Domain Biophysics
Engineering
Hematology

Abstract

It has been previously demonstrated that red blood cells (RBC) possess functional nitric oxide ( NO) synthesizing mechanisms. RBC are also equipped with variety of intracellular control mechanisms, and respond to mechanical forces and to various biological stimuli by increased release of ATP. Nitric oxide has also been demonstrated to be released from RBC under certain circumstances, and it has been hypothesized that NO synthase ( NOS), which is located in both the RBC membrane and cytoplasm, might be activated by mechanical factors. The present study aimed at investigating NOS activation and NO export induced by mechanical stress applied to RBC in suspension. Heparinized venous blood samples were obtained from healthy, adult volunteers and their hematocrit adjusted to 0.4 l/l. The RBC suspensions were equilibrated at room temperature (22 +/- 2 degrees C) with either room air or made hypoxic (36 mmHg, similar to 70% saturation) using moisturized 100% nitrogen. The samples were then continuously pumped through a glass tube (diameter = 0.06 cm; length = 33 cm) for 30 min using a dual syringe pump to maintain a wall shear stress of 0.5-2 Pa with NO concentrations in the RBC suspensions measured electrochemically. NO concentration significantly increased under the influence of 2 Pa in hypoxic RBC suspensions: 105.0 +/- 14.2 nM to 127.1 +/- 12.0 nM as the peak value at 20 min of perfusion. No increase was observed at lower levels of shear stress. Plasma nitrite/nitrate concentrations were measured in samples obtained at five minute intervals. Application of fluid shear stress to hypoxic RBC suspensions resulted in a significant, time-dependent increase of plasma nitrite/nitrate levels, reaching to 14.7 +/- 1.5 mu M from a control value of 11.2 +/- 1.3 mu M. The presence of the non-specific NOS inhibitor L-NAME (10(-3) M) prevented this increment. Additionally, both eNOS and serine 1177 phosphorylated eNOS immuno-fluorescence staining in RBC cytoplasm were shown to increase in response to applied shear stress. Our results support the hypothesis that RBC NO synthase is activated and that export of NO from RBC is enhanced by mechanical stress.