title: Mechanical perturbations trigger endothelial nitric oxide synthase activity in human red blood cells.
abstract: Nitric oxide (NO), a vascular signaling molecule, is primarily produced by endothelial NO synthase. Recently, a functional endothelial NO synthase (eNOS) was described in red blood cells (RBC). The RBC-eNOS contributes to the intravascular NO pool and regulates physiological functions. However the regulatory mechanisms and clinical implications of RBC-eNOS are unknown. The present study investigated regulation and functions of RBC-eNOS under mechanical stimulation. This study shows that mechanical stimuli perturb RBC membrane, which triggers a signaling cascade to activate the eNOS. Extracellular NO level, estimated by the 4-Amino-5-Methylamino-2', 7'-Difluorofluorescein Diacetate probe, was significantly increased under mechanical stimuli. Immunostaining and western blot studies confirmed that the mechanical stimuli phosphorylate the serine 1177 moiety of RBC-eNOS, and activates the enzyme. The NO produced by activation of RBC-eNOS in vortexed RBCs promoted important endothelial functions such as migration and vascular sprouting. We also show that mechanical perturbation facilitates nitrosylation of RBC proteins via eNOS activation. The results of the study confirm that mechanical perturbations sensitize RBC-eNOS to produce NO, which ultimately defines physiological boundaries of RBC structure and functions. Therefore, we propose that mild physical perturbations before, after, or during storage can improve viability of RBCs in blood banks.
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Shunmugan
authors.initials: S
authors.lastname: Nagarajan
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Rajendran Kadarkarai
authors.initials: RK
authors.lastname: Raj
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Venkatesan
authors.initials: V
authors.lastname: Saravanakumar
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Uma Maheswari
authors.initials: UM
authors.lastname: Balaguru
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Jyotirmaya
authors.initials: J
authors.lastname: Behera
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Vinoth Kumar
authors.initials: VK
authors.lastname: Rajendran
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Yogarajan
authors.initials: Y
authors.lastname: Shathya
authors.affiliation: Pondicherry university, Pondicherry, India.
authors.forename: B Mohammed Jaffar
authors.initials: BM
authors.lastname: Ali
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.
authors.forename: Venil
authors.initials: V
authors.lastname: Sumantran
authors.affiliation: VascularBiology Laboratory, AU-KBC Research Centre, Chennai, Tamil Nadu, India.|Department of Biotechnology, Anna University, Chennai, Tamil Nadu, India.
authors.forename: Suvro
authors.initials: S
authors.lastname: Chatterjee
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D002465:Cell Movement; D004789:Enzyme Activation; D004912:Erythrocytes / Q000201:enzymology*; D015870:Gene Expression; D006801:Humans; D040542:Mechanotransduction, Cellular; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism*
chemical_list: D009569:Nitric Oxide; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III