Multiple Immune and Genetic Mechanisms Contribute to Cmv5s-Driven Susceptibility and Tissue Damage during Acute Murine Cytomegalovirus Infection.

The MHC class I molecule H-2Dk conveys resistance to acute murine CMV infection in both C57L (H-2Dk transgenic) and MA/My mice. M.H2k/b mice are on an MA/My background aside from a C57L-derived region spanning the MHC (Cmv5s), which diminishes this resistance and causes significant spleen histopathology. To hone in on the effector elements within the Cmv5s interval, we generated several Cmv5-recombinant congenic mouse strains and screened them in vivo, allowing us to narrow the phenotype-associated interval >6-fold and segment the genetic mechanism to at least two independent loci within the MHC region. In addition, we sought to further characterize the Cmv5s-associated phenotypes in their temporal appearance and potential direct relationship to viral load. To this end, we found that Cmv5s histopathology and NK cell activation could not be fully mirrored in the MA/My mice with increased viral dose, and that marginal zone destruction was the first apparent Cmv5s phenotype, being reliably quantified as early as 2 d postinfection in the M.H2k/b mice, prior to divergence in viral load, weight loss, or NK cell phenotype. Finally, we further dissect NK cell involvement, finding no intrinsic differences in NK cell function, despite increased upregulation of activation markers and checkpoint receptors. In conclusion, these data dissect the genetic and immunologic underpinnings of Cmv5 and reveal a model in which polymorphism within the MHC region of the genome leads to the development of tissue damage and corrupts protective NK cell immunity during acute viral infection.

Obesity-Induced Coronary Microvascular Disease Is Prevented by iNOS Deletion and Reversed by iNOS Inhibition.

Coronary microvascular disease (CMD) caused by obesity and diabetes is major contributor to heart failure with preserved ejection fraction; however, the mechanisms underlying CMD are not well understood. Using cardiac magnetic resonance applied to mice fed a high-fat, high-sucrose diet as a model of CMD, we elucidated the role of inducible nitric oxide synthase (iNOS) and 1400W, an iNOS antagonist, in CMD. Global iNOS deletion prevented CMD along with the associated oxidative stress and diastolic and subclinical systolic dysfunction. The 1400W treatment reversed established CMD and oxidative stress and preserved systolic/diastolic function in mice fed a high-fat, high-sucrose diet. Thus, iNOS may represent a therapeutic target for CMD.

Loss of Endogenously Cycling Adult Cardiomyocytes Worsens Myocardial Function.

RATIONALE: Endogenously cycling adult cardiomyocytes increase after myocardial infarction (MI) but remain scarce and are generally thought not to contribute to myocardial function. However, this broadly held assumption has not been tested, mainly because of the lack of transgenic reporters that restrict Cre expression to adult cardiomyocytes that reenter the cell cycle. OBJECTIVE: We created and validated a new transgenic mouse, αMHC (alpha myosin heavy chain)-MerDreMer-Ki67p-RoxedCre (denoted αDKRC [cardiomyocyte-specific αMHC-MerDreMer-Ki67p-RoxedCre]) that restricts Cre expression to cycling adult cardiomyocytes and uniquely integrates spatial and temporal adult cardiomyocyte cycling events based on the DNA specificities of orthologous Dre and Cre recombinases. We then created αDKRC::DTA mice that expressed an inducible diphtheria toxin in adult cycling cardiomyocytes and examined the effects of ablating these endogenously cycling cardiomyocytes on myocardial function after ischemic-reperfusion (I/R) MI. METHODS AND RESULTS: A tandem αDKRC transgene was designed, validated in cultured cells, and used to make transgenic mice. The αDKRC transgene integrated between MYH6 and MYH7 and did not disrupt expression of the surrounding genes. Compared with controls, αDKRC::RLTG (Rox-Lox-tdTomato-eGFP) mice treated with Tamoxifen expressed tdTomato+ in cardiomyocytes with rare Bromodeoxyuridine+, eGFP+ cardiomyocytes, consistent with reentry of the cell cycle. We then pretreated αDKRC::RLTG mice with Tamoxifen to activate the reporter before sham or reperfusion (I/R) MI surgeries. Compared with Sham surgery, the I/R MI group had increased single and paired eGFP+ (enhanced green fluorescent protein)+ cardiomyocytes predominantly in the border zones (5.8±0.5 versus 3.3±0.3 cardiomyocytes per 10-micron section, N=8-9 mice per group, n=16-24 sections per mouse), indicative of cycled cardiomyocytes. The single to paired eGFP+ cardiomyocyte ratio was ≈9 to 1 (5.2±0.4 single versus 0.6±0.2 paired cardiomyocytes) in the I/R MI group after MI, suggesting that cycling cardiomyocytes were more likely to undergo polyploidy than replication. The ablation of endogenously cycling adult cardiomyocytes in αDKRC::DTA (diphtheria) mice caused progressive worsening left ventricular chamber size and function after I/R MI, compared with controls. CONCLUSIONS: Although scarce, endogenously cycling adult cardiomyocytes contribute to myocardial function after injury, suggesting that these cells may be physiologically relevant.