Cytochrome P450 2J2 (CYP2J2) is a known arachidonic acid (AA) epoxygenase that mediates the formation of four bioactive regioisomers of cis-epoxyeicosatrienoic acids (EETs). Although its expression in the liver is low, CYP2J2 is mainly observed in extrahepatic tissues, including the small intestine, pancreas, lung, and heart. Changes in CYP2J2 levels or activity by xenobiotics, disease states, or polymorphisms are proposed to lead to various organ dysfunctions. Several studies have investigated the regulation of CYP2J2 and EET formation in various cell lines and have demonstrated that such regulation is tissue-dependent. In addition, studies linking CYP2J2 polymorphisms to the risk of developing cardiovascular disease (CVD) yielded contradictory results. This review will focus on the mechanisms of regulation of CYP2J2 by inducers, inhibitors, and oxidative stress modeling certain disease states in various cell lines and tissues. The implication of CYP2J2 expression, polymorphisms, activity and, as a result, EET levels in the pathophysiology of diabetes and CVD will also be discussed.
Epoxyeicosatrienoic acids (EETs) are metabolites of arachidonic acid (AA) oxidation that have important cardioprotective and signaling -6 polyunsaturated fatty acid (PUFA) that is prone to autoxidation. Although hydroperoxides and isoprostanes are major autoxidation products of AA, EETs are also formed from the largely overlooked peroxyl radical addition mechanism. While autoxidation yields both cis- and trans-EETs, cytochrome P450 (CYP) epoxygenases have been shown to exclusively catalyze the formation of all regioisomer cis-EETs, on each of the double bonds. In plasma and red blood cell (RBC) membranes, cis- and trans-EETs have been observed, and both have multiple physiological functions. We developed a sensitive ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) assay that separates cis- and trans- isomers of EETs and applied it to determine the relative distribution of cis- vs. trans-EETs in reaction mixtures of AA subjected to free radical oxidation in benzene and liposomes in vitro. We also determined the in vivo distribution of EETs in several tissues, including human and mouse heart, and RBC membranes. We then measured EET levels in heart and RBC of young mice compared to old. Formation of EETs in free radical reactions of AA in benzene and in liposomes exhibited time- and AA concentration-dependent increase and trans-EET levels were higher than cis-EETs under both conditions. In contrast, cis-EET levels were overall higher in biological samples. In general, trans-EETs increased with mouse age more than cis-EETs. We propose a mechanism for the non-enzymatic formation of cis- and trans-EETs involving addition of the peroxyl radical to one of AA's double bonds followed by bond rotation and intramolecular homolytic substitution (SHi). Enzymatic formation of cis-EETs by cytochrome P450 most likely occurs via a one-step concerted mechanism that does not allow bond rotation. The ability to accurately measure circulating EETs resulting from autoxidation or enzymatic reactions in plasma and RBC membranes will allow for future studies investigating how these important signaling lipids correlate with heart disease outcomes.
The twist energy parameter (E T ) that governs the supercoiling free energy, and the linking difference (Δl) are measured for p30 δ DNA in solutions containing 0 to 40 w/v% ethylene glycol (EG). A plot of E T vs. −ln a w , where a w is the water activity, displays the full (reverse) sigmoidal profile of a discrete structural transition. A general theory for the effect of added osmolyte on a cooperative structural transition between two duplex states, 1□ 2, is formulated in terms of parameters applicable to individual base-pairs subunits. The resulting fraction of base-pairs in the 2-state ( ), is incorporated into expressions for the effective torsion and bending elastic constants, the effective twist energy parameter ( ), and the change in intrinsic twist (δl 0 ). Fitting the expression for to the measured E T -values yields reasonably unambiguous estimates of E T 1 and E T 2 , the midpoint value (ln a w ) 1/2 , and midpoint slope (∂E T /∂ln a w ) 1/2 , but does not yield unambiguous estimates of the equilibrium constant ( K 0 ), the difference in DNA-water preferential interaction coefficient (ΔΓ), or the inverse cooperativity parameter, J. Fitting a non-cooperative model (assumed J=1.0) to the data yields, K 0 = 0.067, and ΔΓ = − 30.0 per base-pair (bp). Essentially equivalent fits are provided by models with a wide range of correlated J, ΔΓ, and K 0 values. Other results favor ΔΓ in the range − 1.0 to 0, which then requires K 0 ≥ 0.914, and a cooperativity parameter, 1/J ≥ 30.0 bp. The measured δl 0 and circular dichroism (CD) at 272 nm are found to be compatible with curves predicted using the same -values that best-fit the E T -data. At least 7 to 10 % of the base-pairs are inferred to exist in the 2-state in 0.1 M NaCl in the complete absence of added osmolyte. Compared with the 1-state, the 2-state has a ~2.0-to 2.1-fold greater torsion elastic constant, a ~0.70-fold smaller bending elastic constant, a ~0.91-fold smaller E T -value, a ~0.2 % lower intrinsic twist, a somewhat lower CD near both 272 and 245 nm, and less water and/or more EG in its neighborhood. However, the relative change in preferential interaction coefficient associated with the transition is likely rather slight.
CYP2J2, a member of the Cytochrome P450 family of enzymes, is the most abundant epoxygenase in the heart and has multifunctional properties including bioactivation of arachidonic acid to epoxyeicosatrienoic acids, which, in turn, have been implicated in mediating several cardiovascular conditions. Using a proteomic approach, we found that CYP2J2 expression is lower in cardiac tissue from patients with cardiomyopathy compared to controls. In order to better elucidate the complex role played by CYP2J2 in cardiac cells, we performed targeted silencing of CYP2J2 expression in human adult ventricular cardiomyocytes and interrogated whole genome transcriptional responses. We found that knockdown of CYP2J2 elicits widespread alterations in gene expression of ventricular cardiomyocytes and leads to the activation of a diverse repertoire of programs, including those involved in ion channel signaling, development, extracellular matrix, and metabolism. Several members of the differentially up-regulated ion channel module have well-known pathogenetic roles in cardiac dysrhythmias. By leveraging causal network and upstream regulator analysis, we identified several candidate drivers of the observed transcriptional response to CYP2J2 silencing; these master regulators have been implicated in aberrant cardiac remodeling, heart failure, and myocyte injury and repair. Collectively, our study demonstrates that CYP2J2 plays a central and multifaceted role in cardiomyocyte homeostasis and provides a framework for identifying critical regulators and pathways influenced by this gene in cardiovascular health and disease.
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