Endocannabinoids And Energy Homeostasis

National Institute On Alcohol Abuse And Alcoholism

Investigators

Linked publications & trials

Abstract

In previous studies we demonstrated that both the endocannabinoid/CB1 receptor (CB1R) system and inducible nitric oxide synthase (iNOS) are upregulated in various fibrotic tissues and contribute to the initiation and progression of fibrosis. This has led us to design and synthesize hybrid CB1R/iNOS inhibitors that can simultaneously engage these 2 targets resulting in an increase in their anti-fibrotic efficacy compared to engaging only one of these targets. We were able to prove this concept in different preclinical models of liver, lung and skin fibrosis, using the hybrid CB1R/iNOS inhibitor MRI-1867 (JCI Insight 2016; 2017; Clin Transl Med 2021; Front Endocrinol 2021). In a hypothesis article, we make the case for the therapeutic potential of a hybrid CB1/iNOS inhibitor in mitigating the fibroproliferative changes of long-Covid in the lung and some other tissues. The case is based on the documented role of pulmonary CB1R in inflammatory and fibroproliferative changes in the lung, as reviewed in Br J Pharmacology 179:2121, 2022. In other projects we are analyzing the role of peripheral CB1 receptors in hepatocytes vs. adipocytes in obesity-related, non-alcoholic fatty liver disease (NAFLD). We earlier reported that the steatosis of mice with high-fat diet-induced obesity (DIO) is nearly completely reversed by treatment with a peripherally restricted CB1 antagonist even though mutant mice that express CB1 receptors in hepatocytes only on a global CB1 knockout background accumulate very little hepatic fat when fed the same high-fat diet (JCI 2010). This suggested that diet-induced steatosis may be promoted by extrahepatic peripheral CB1 receptors, such as those in adipocytes, a known source of fatty acids released and taken up by the liver for the synthesis of hepatic triglycerides. Indeed, a recent publication indicated that mice with conditional knockout of CB1R in adipocytes are resistant to DIO and the associated steatosis (JCI 127:4148, 2017). Based on this we decided to test whether adipocyte CB1 are not only necessary but also sufficient for the development of diet-induced steatosis. For this, we are generating a rescue model, i.e. mutant mice with transgenic expression of CB1 in adipocytes only on a global CB1R-KO background. To achieve this, tThe gRNA to mouse ROSA26 gene, the donor vector containing in reverse orientation the "Adipoq promoter-Kozak-Mouse Cnr1 CDS-rBG pA" cassette, and Cas9 mRNA were co-injected into fertilized mouse eggs to generate targeted knockin offspring. F0 founder animals were identified by PCR followed by sequence analysis, which were bred to wildtype mice to test germline transmission and F1 animal generation. Heterozygote breeding pairs were then interbred to yield homozygote transgenes, which were then bred with global CB1R-KO mice to generate the required genotype. This breeding program is ongoing. As mentioned above, hepatic CB1R have minimal role in the development of diet-induced hepatic steatosis, which led some to question their functional relevance. However, this view ignores the possible roe of hepatic CB1R in the reversal of steatosis by CB1 blockade, which has not yet been tested (JCI Another related project is aimed to explore the role of hepatic CB1R