Reduced RhoA Activity Mediates Acute Alcohol Intoxication‐Induced Inhibition of Lymphatic Myogenic Constriction Despite Increased Cytosolic [<scp><scp>Ca</scp></scp><sup>2+</sup>]

Daines

Doggett

et al. 2016

JAHA

Self Cite

BackgroundMicrovascular leakage of plasma proteins is a hallmark of inflammation that leads to tissue dysfunction. There are no current therapeutic strategies to reduce microvascular permeability. The purpose of this study was to identify the role of Rnd3, an atypical Rho family GTPase, in the control of endothelial barrier integrity. The potential therapeutic benefit of Rnd3 protein delivery to ameliorate microvascular leakage was also investigated.Methods and ResultsUsing immunofluorescence microscopy, Rnd3 was observed primarily in cytoplasmic areas around the nuclei of human umbilical vein endothelial cells. Permeability to fluorescein isothiocyanate–albumin and transendothelial electrical resistance of human umbilical vein endothelial cell monolayers served as indices of barrier function, and RhoA, Rac1, and Cdc42 activities were determined using G‐LISA assays. Overexpression of Rnd3 significantly reduced the magnitude of thrombin‐induced barrier dysfunction, and abolished thrombin‐induced Rac1 inactivation. Depleting Rnd3 expression with siRNA significantly extended the time course of thrombin‐induced barrier dysfunction and Rac1 inactivation. Time‐lapse microscopy of human umbilical vein endothelial cells expressing GFP‐actin showed that co‐expression of mCherry‐Rnd3 attenuated thrombin‐induced reductions in local lamellipodia that accompany endothelial barrier dysfunction. Lastly, a novel Rnd3 protein delivery method reduced microvascular leakage in a rat model of hemorrhagic shock and resuscitation, assessed by both intravital microscopic observation of extravasation of fluorescein isothiocyanate–albumin from the mesenteric microcirculation, and direct determination of solute permeability in intact isolated venules.ConclusionsThe data suggest that Rnd3 can shift the balance of RhoA and Rac1 signaling in endothelial cells. In addition, our findings suggest the therapeutic, anti‐inflammatory potential of delivering Rnd3 to promote endothelial barrier recovery during inflammatory challenge.

Section: Methodsmentioning

confidence: 99%

Rnd3 as a Novel Target to Ameliorate Microvascular Leakage

Daines

Doggett

et al. 2016

JAHA

Self Cite

“…6,7,24,25 Briefly, rats were anesthetized with ketamine and xylazine (90 and 9 mg/kg, respectively). A sterile catheter was aseptically placed into the antrum of the stomach and exteriorized at the nape of the neck.…”

Section: Animalsmentioning

confidence: 99%

“…7,23 Briefly, upon entry into the cytoplasm, AM portion of the dye is cleaved, making fura-2 membrane-impermeable. Measurements were collected in isolated lymphatics loaded with fura-2 by illuminating at alternating wavelengths of 340 and 380 nm via a dichroic mirror (400 nm; Chroma Technology Corp. 400DCLP) for durations of 50 ms each (the shortest time we could obtain sufficient signal with our hardware), over a period of 1-2 min.…”

Section: Ratiometric Measurement Of [Ca 2 + ] Imentioning

confidence: 99%

“…[2][3][4][5] In previous studies, we demonstrated that acute alcohol intoxication (AAI) modulates the pump function of mesenteric lymphatics. 6,7 We have chosen to study AAI-induced changes in lymphatic pumping because AAI complicates recovery from injury and infection by impairing cardiovascular function, fluid homeostasis, and host defense. 8 The gut lymphatics are a primary route for nonbacterial, tissue injury factors released from the GI tract.…”

mentioning

confidence: 99%

“…This uncoupling was reflected in the AAI-induced increased magnitude of phasic Ca +2 mobilization and attenuated lymphatic myogenic constriction by impaired RhoA/ROCK pathwaymediated Ca +2 sensitivity. 6,7 To expand understanding of the molecular mechanisms that mediate the AAI-induced increase in Ca 2 + transient magnitude of lymphatic smooth muscle, in the present study, we investigated the potential role of sarcolemmal Ca 2 + entry through L-type Ca 2 + channels and its uptake/release from the sarcoplasmic reticulum (SR) in rat isolated mesenteric lymphatic vessels.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Mechanisms of Acute Alcohol Intoxication-Induced Modulation of Cyclic Mobilization of [Ca²⁺] in Rat Mesenteric Lymphatic Vessels

Souza‐Smith

Kerut

Lymphatic Research and Biology

et al. 2015

Self Cite

Lymphatic Vessel Network Structure and Physiology

Comprehensive Physiology

Yang

Scallan

et al. 2018

Self Cite

267

258

The lymphatic system is comprised of a network of vessels interrelated with lymphoid tissue, which has the holistic function to maintain the local physiologic environment for every cell in all tissues of the body. The lymphatic system maintains extracellular fluid homeostasis favorable for optimal tissue function, removing substances that arise due to metabolism or cell death, and optimizing immunity against bacteria, viruses, parasites, and other antigens. This article provides a comprehensive review of important findings over the past century along with recent advances in the understanding of the anatomy and physiology of lymphatic vessels, including tissue/organ specificity, development, mechanisms of lymph formation and transport, lymphangiogenesis, and the roles of lymphatics in disease.