University of California San Francisco

Department of Anesthesia and Perioperative Care

1.

Wilhelmsen K, Khakpour S, Tran A, Sheehan K, Schumacher M, Xu F, Hellman J. The Endocannabinoid/Endovanilloid N-Arachidonoyl Dopamine (NADA) and Synthetic Cannabinoid WIN55,212-2 Abate the Inflammatory Activation of Human Endothelial Cells. J Biol Chem. 2014 May 9;289(19):13079-100. (IF: 4.651)

During health endothelial cells maintain vascular homeostasis, and mediate critical aspects of the host’s innate immune response to infection. It is widely believed that during sepsis and inflammatory critical illness, endothelial cell dysfunction contributes to systemic inflammation, and to the coagulopathy, vascular leak, and excessive neutrophil activity that promote organ failure and death. The endogenous cannabinoid system, discovered in the 1990’s has been postulated to maintain homeostasis in a variety of systems. Endogenously produced cannabinoids (endocannabinoids) are derived from arachidonic acid, and are ligands for the cannabinoid receptors 1 and 2 (CB1R and CB2R) and for the transient receptor potential channel V1 (TRPV1). Several endocannabinoids have been reported to modulate aspects of leukocyte inflammatory responses, but thus far the role of the endothelial endocannabinoid system during inflammation has not been thoroughly assessed. We report that endothelial cells express CBRs, TRPV1, and endocannabinoid metabolic enzymes, and that the endocannabinoid N-arachidonoyl dopamine (NADA) and synthetic cannabinoid WIN55,212-2, but not anandamide (AEA) nor 2-arachidonoylglycerol (2-AG), reduce endothelial cell secretion of cytokines and expression of adhesion molecules, and decrease neutrophil adherence to endothelial cell monolayers activated by TNFa, endotoxin or bacterial lipopeptides. We observed that the effects of NADA on the inflammatory activation of endothelial cells are attenuated by CBR antagonists and augmented by TRPV1 antagonists. These data suggest that NADA constrains the endothelial cell inflammatory response through opposing actions at CBRs and TRPV1. Our study suggests that the endothelial endocannabinoid system may play an important role in the resolution of endothelial inflammation, and thus could potentially be targeted to ameliorate inflammatory critical illness.

2.

Chen W, Sun Z, Han Z, Jun K, Camus M, Wankhede M, Mao L, Arnold T, Young WL, Su H. De novo cerebrovascular malformation in the adult mouse after endothelial Alk1 deletion and angiogenic stimulation. Stroke. 2014 Mar;45(3):900-2. (IF: 6.018)

Brain arteriovenous malformation (bAVM) is an important risk factor for intracranial hemorrhage, especially in children and young adults, and the factors contributing to its formation have not been identified. In humans, a deficiency of activin receptor-like kinase 1 (Alk1) causes type 2 hereditary hemorrhagic telangiectasia (HHT2). HHT2 patients can have AVMs in multiple organs, including the brain, and the bAVM phenotype in those patients is similar to that of sporadic bAVM. To identify gene mutation, in which cell types will trigger bAVM formation, we used an HHT2 mouse model and deleted Alk1 in different cell types using a conditional gene knockout strategy. We found that deletion of the Alk1 gene in endothelial cells is essential in initiating de novo bAVM formation.

3.

McAuley DF, Curley GF, Hamid UI, Laffey JG, Abbott J, McKenna DH, Fang X, Matthay MA, Lee JW. Clinical grade allogeneic human mesenchymal stem cells restore alveolar fluid clearance in human lungs rejected for transplantation. American journal of physiology. Lung cellular and molecular physiology. 2014;306(9):L809-815. (IF: 4.04)

The need to increase the donor pool for lung transplantation is a major public health issue. Recently, a new technique of ex vivo lung perfusion (EVLP) has been developed to extend the donor pool size. EVLP allows “rehabilitation” of marginal donor lungs initially rejected for transplantation by allowing a short duration of perfusion and oxygenation with ventilation prior to transplantation. EVLP has also become an ideal method to test the effects of pharmacologic and/or gene- or cell-based therapy prior to surgery to improve the success of lung transplantation. Using an ex vivo human lung preparation, the current studies were designed to determine if intravenous clinical-grade human mesenchymal stem cells (MSC) would be effective in restoring AFC in donor lungs that had been deemed unsuitable for transplantation; impaired AFC contributes to primary graft dysfunction, a major cause of morbidity and mortality after lung transplantation in the form of non-cardiogenic pulmonary edema. The human lungs were perfused with 5% albumin in a balanced electrolyte solution and oxygenated with continuous positive airway pressure. Baseline AFC was measured in the control lobe and if AFC was impaired (defined as <10%/h), the lungs received either 5 × 106 MSC added to the perfusate or perfusion only as a control. AFC was measured in a different lung lobe at 4 h. Intravenous administration of human MSC restored AFC in the injured lungs to a normal level, an effect that was dependent on MSC secretion of keratinocyte growth factor. In contrast, perfusion only did not increase AFC. In summary, administration of allogeneic human MSC was effective in restoring the capacity of the alveolar epithelium to remove pulmonary edema fluid at a normal rate, further rehabilitating marginal donor lungs and potentially developing a technique to increase donor pool size.

4.

Lee BH, Chan JT, Kraeva E, Peterson K, Sall JW. Isoflurane exposure in newborn rats induces long-term cognitive dysfunction in males but not females. Neuropharmacology. 2014;83:9-17. (IF: 4.819)

In a recently published human and animal study we noted a possible difference in cognitive outcome between male and female children that were exposed to anesthesia at an early age (Stratmann et al, Neuropsychopharmacology 2014). This finding was further investigated using a rat model of isoflurane exposure on day of life seven. All anesthesia exposed groups showed the expected increase in brain cell death with no difference by sex. One month later using a series of 4 increasingly difficult object recognition/association tasks we found isoflurane exposed males impaired in 3 of the 4 tasks, while females were not impaired in any task. In further testing, all animals showed appropriate social interaction, but isoflurane exposed males were impaired in social recognition. These findings suggest that males may have a greater risk of cognitive impairment after early life anesthesia exposure and that brain cell death which was similar between groups is likely not the primary cause for the deficits we observed. http://www.sciencedirect.com/science/article/pii/S0028390814001063

5.

Zavala K, Lee J, Chong J, Sharma M, Eilers H, Schumacher MA. The anticancer antibiotic mithramycin-A inhibits TRPV1 expression in dorsal root ganglion neurons. Neurosci Lett. 2014 Aug 22;578:211-6. doi: 10.1016/j.neulet.2014.01.021. Epub 2014 Jan 25.

PMID: 24468003

Estimates of the number of individuals suffering from chronic pain in the United States and its physical, emotional and financial toll are staggering. Although tremendous clinical advances have been made, our ability to prevent or reverse the development of chronic pain states remains a largely unresolved clinical challenge. Activation of peripheral nociceptors by products of inflammation has been shown to be dependent on specific sensory transducing elements such as the capsaicin receptor, TRPV1. Transient receptor potential vanilloid type -1 (TRPV1) directs complex roles in signal transduction including the detection of noxious stimuli arising from cellular injury and inflammation. One line of investigation has been related to understanding the consequence of TRPV1 up-regulation in sensory neurons under chronic pathophysiologic conditions such as inflammation or nerve injury. Within this context, we have serendipitously determined that the principal promoter for the capsaicin receptor (TRPV1) contains a GC-box binding motif within promoter P2 that is essential for TRPV1 transcriptional activity. A search for small-molecule inhibitors acting at GC-box motifs revealed the anticancer agent mithramycin-A (plicamycin) an aureolic acid polyketide produced by a species of the soil bacterium Streptomyces. Mithramycin-A has been described as having its anticancer / antiproliferative effect through binding to GC- and / or GT-box DNA binding motifs and either displacing or preventing the binding of transcription factors that utilize GC-box motifs within their promoter site. Based on our model of Sp1 and Sp4 binding to the GC-box domain within the rTRPV1 promoter region, we tested whether the small molecule, mithramcyin-A, would alter endogenous TRPV1 expression in cultured rat primary sensory neurons. Mithramycin-A dose-dependently (10-50nM) decreased endogenous TRPV1 mRNA content and appeared to decrease TRPV1-like protein expression in DRG neurons. We also observed that mithramycin-A directed a decrease in the number of capsaicin-responsive DRG neurons without a significant change in the capsaicin-response magnitudes. Interestingly, mithramycin-A also reduced the mRNA encoding Sp1 and Sp4 in DRG neurons, transcription factors previously found to positively regulate TRPV1 expression in sensory neurons. Taken together Mithramycin-A blocks expression of TRPV1 in cultured DRG neurons and may serve as a model for the development of transcriptional inhibitors in chronic pain therapeutics.

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