Qianli Yu

Role of T Lymphocytes in Hypertension-Induced Cardiac Extracellular Matrix Remodeling

Cardiac remodeling in response to pressure overload involves reorganization of the myocytes and extracellular matrix (ECM). Neurohormonal pathways have been described as effector pathways in left ventricular ECM reorganization in response to pressure overload; we now are assessing the role of the T lymphocyte in this process. Mice with defined differences in T-lymphocyte function (C57BL/6 SCID, C57BL/6 WT, and BALB/c) were treated with 50 mg/L of NG-nitro-l-arginine methyl ester in their drinking water for 30 days. The immune function of C57BL/6 WT mice was T-helper type 1 (TH1), BALB/c was TH2, and C57BL/6 SCID was null. The arterial blood pressure increased by 30% in all of the strains of mice. However, ventricular stiffness significantly decreased in the C57 SCID, significantly increased in the BALB/c, and did not change in the C57 WT. The characterization of matrix metalloproteinase induction and activation on day 30 was associated with T-lymphocyte function. The total cardiac fibrillar collagen, percentage of fibrillar collagen cross-linking, and the activity of the cross-linking enzyme lysyl oxidase-like–3 (LOXL-3) significantly decreased in the C57 SCID, significantly increased in the BALB/c, and did not change in the C57 WT. This study revealed that the LOXL-3 pathway, namely, gene expression, enzymatic activities, and LOXL-3–mediated collagen cross-linking, was associated with ventricular stiffness and incongruence with lymphocyte function. These data support the concept that the T lymphocytes may play a fundamental regulatory role in cardiac ECM composition through modulation of collagen synthesis, degradation, and cross-linking.

Heart disease, methamphetamine and AIDS.

Methamphetamine (MA) not only affects the nervous system but also has cardiac toxicity and immunosuppressive properties. This manuscript will provide support that there is a relationship between MA use and heart disease as well as immune dysfunction. The cardiovascular manifestations of acute MA use include tachycardia, atrioventricular arrhythmias, myocardial ischemia, myocardial ischemia and hypertension, resulting in cardiac lesions. Chronic use of MA causes cardiomyopathy including cellular infiltration, myocardial hypertrophy, myocardium rupture and fibrosis. The increased catecholamine levels are responsible for the cardiac lesions induced by MA. The additional problem with MA use is its potential to disrupt the immune system function leading to suppression of mitogen-stimulated lymphocyte, a reduction in circulating lymphocyte numbers and alternation T-lymphocyte cytokine secretion as well as B cell proinflammatory cytokine secretion. Concomitant MA use and Human Immunodeficiency Virus (HIV) infection not only enhances immunosuppression associated with HIV but also increases the heart disease occurrence with a coincidentally complication of AIDS or AIDS medications.

Truncation of Titin’s Elastic PEVK Region Leads to Cardiomyopathy With Diastolic Dysfunction

Rationale: The giant protein titin plays key roles in myofilament assembly and determines the passive mechanical properties of the sarcomere. The cardiac titin molecule has 2 mayor elastic elements, the N2B and the PEVK region. Both have been suggested to determine the elastic properties of the heart with loss of function data only available for the N2B region. Objective: The purpose of this study was to investigate the contribution of titin’s proline–glutamate–valine–lysine (PEVK) region to biomechanics and growth of the heart. Methods and Results: We removed a portion of the PEVK segment (exons 219 to 225; 282 aa) that corresponds to the PEVK element of N2B titin, the main cardiac titin isoform. Adult homozygous PEVK knockout (KO) mice developed diastolic dysfunction, as determined by pressure-volume loops, echocardiography, isolated heart experiments, and muscle mechanics. Immunoelectron microscopy revealed increased strain of the N2B element, a spring region retained in the PEVK-KO. Interestingly, the PEVK-KO mice had hypertrophied hearts with an induction of the hypertrophy and fetal gene response that includes upregulation of FHL proteins. This contrasts the cardiac atrophy phenotype with decreased FHL2 levels that result from the deletion of the N2B element. Conclusions: Titin’s PEVK region contributes to the elastic properties of the cardiac ventricle. Our findings are consistent with a model in which strain of the N2B spring element and expression of FHL proteins trigger cardiac hypertrophy. These novel findings provide a molecular basis for the future differential therapy of isolated diastolic dysfunction versus more complex cardiomyopathies.

Chronic methamphetamine exposure alters immune function in normal and retrovirus-infected mice.

Methamphetamine (MA) abuse represents a growing problem in the USA with an increase of sudden death. To evaluate the immune function alterations due to chronic methamphetamine use, we examined C57BL/C mice with LP-BM5 retrovirus infection plus methamphetamine exposure. Mice were randomly assigned to the following groups: placebo, placebo retrovirus-infected, uninfected MA treated and retrovirus-infected MA treated. Placebo, MA-treated groups were intraperitoneally injected with saline, MA, respectively, with a gradually increasing dose from 15 to 40 mg/kg for 12 weeks (5 days/week). Con A- and LPS-induced mitogenesis of splenocytes, cytokine production by splenocytes culture and lipid peroxides in the liver were measured. Heart tissue histopathology was analyzed in all the groups with murine cytomegalovirus (CMV) superinfection. Our data showed that MA treatment significantly decreased production of IL-2 and interferon gamma (IFN-gamma) in uninfected mice but did not further suppress the reduced Th1 cytokines in retrovirus-infected mice. There were no significant effects on cytokines IL-4 and IL-6. However, tumor necrosis factor (TNF-alpha) was significantly increased in both uninfected and infected mice due to MA treatment. Lipid peroxides in liver were significantly increased both in uninfected and retrovirus-infected mice due to MA exposure. Vitamin E levels in liver were significantly decreased in uninfected mice due to MA treatment. CMV superinfection greatly increased the cardiac lesions in retrovirus-infected mice while no significant histopathology changes were detected due to MA treatment. Our data suggest that MA has immunomodulation activity, suppressing Th1 cytokine production and enhancing some Th2 cytokine secretion, as well as increasing lipid peroxides in uninfected mice. The interaction between LP-BM5 and MA remains unclear.

IL-18 induction of osteopontin mediates cardiac fibrosis and diastolic dysfunction in mice

Osteopontin (OPN), a key component of the extracellular matrix, is associated with the fibrotic process during tissue remodeling. OPN and the cytokine interleukin (IL)-18 have been shown to be overexpressed in an array of human cardiac pathologies. In the present study, we determined the role of IL-18 in the regulation of cardiac OPN expression and the subsequent interstitial fibrosis and diastolic dysfunction. We demonstrated parallel increases in IL-18, OPN expression, and interstitial fibrosis in murine models of left ventricular pressure and volume overload. Exogenous recombinant (r)IL-18 administered for 2 wk increased cardiac OPN expression, interstitial fibrosis, and diastolic dysfunction. Stimulation of the T helper (Th)1 lymphocyte phenotype with a selective toll-like receptor (TLR)9 agonist induced cardiac IL-18 and OPN expression, which was associated with increased cardiac fibrillar collagen concentrations and interstitial fibrosis resulting in diastolic dysfunction. rIL-18 induced OPN expression and protein levels in primary of cardiac fibroblast cultures. Conditioned media from TLR9-stimulated T lymphocyte cultures induced IL-18 and OPN expression in cardiac fibroblasts, while blockade of the IL-18 receptor with a neutralizing antibody abolished the increase in OPN expression. Furthermore, a mutation in the transcriptional factor interferon regulatory factor (IRF)1 or IRF1 small interfering RNA (siRNA) resulted in the decreased expression of IL-18 and OPN in cardiac fibroblasts. With pressure overload, IRF1-mutant mice showed downregulation of IL-18 and OPN expression in cardiac tissue, reduced cardiac fibrotic development, and increased left ventricular function compared with wild type. These results provide direct evidence that the induction of IL-18 regulates OPN-mediated cardiac fibrosis and diastolic dysfunction.

The effect of daily administration of IL-18 on cardiac structure and function.

Recently, the cytokine Interleukin-18 (IL-18) has been shown to be increased as a result of cardiac surgery. Elevated IL-18 has been associated with neurological dysfunction, systemic inflammatory response syndrome (SIRS), and multiple organ dysfunction syndrome (MODS) post open-heart surgery. The intent of the study contained herein was to determine the effect of IL-18 administration on cardiac function and structure. Eight C57BL/6 female mice were treated daily with 0.5μg/mouse of recombinant IL-18 for 7 days. Long axis echocardiography (ECHO) measurements of the anatomical and hemodynamic function of the heart for all mice were studied 24h after the last dose. The left ventricular wet weights increased from 84 ± 1 to 93 ± 3 mg when comparing the placebo (n = 8) with the IL-18 groups, respectively (p = 0.01). With ECHO analysis, IL-18 significantly increased left ventricular (LV) mass, the left atrium dimensions (LA), and the left ventricular posterior wall thickness (LVPW) over the 8-day time period (p < 0.01). There was a 5-fold increase in interstitial cardiac collagen content and a 30% increase in myocyte size in the IL-18 compared with the control groups (p < 0.01). Administration of IL-18 appears to induce interstitial fibrosis and myocyte hypertrophy, resulting in increased ventricular stiffness. Thus, increased IL-18 during and post open-heart surgical procedures may induce left ventricular diastolic dysfunction and affect post-operative outcomes.

Characterization of High-Salt and High-Fat Diets on Cardiac and Vascular Function in Mice

This study compared two established dietary formulations, high salt and high fat-high carbohydrate, separately or in combination on the induction cardiovascular dysfunction. One-month-old C57BL/6J mice were fed one of the following diets for 3 mo: (1) control diet consisting of a high fat-high simple carbohydrate (HFHSC); (2) 8% NaCl diet (HS); or (3) HFHSC diet supplemented with 1% NaCl (HFHS). After 3 mo, the HFHSC mice demonstrated significantly increased end-diastolic volume and end-systolic volume, specifically increases of 35% and 78%, respectively (p<0.01) and a reduction of ventricular stiffness by 27% (p=0.015). The HS mice exhibited arterial hypertension with an increase of 33% in maximum end-systolic pressure (p=.024) and a decrease of 44% in arterial elastance (p=0.020), corroborated by an increase in the heart weight to body weight ratios (p=0.002) and vascular types I and III collagen (p=0.03 and p=0.0008, respectively). The HFHS group revealed a striking response of 230% to the α1-adrenergic challenge (p=0.00034). These data suggest that the HFHSC diet causes dilaeed cardiomyopathy, whereas the HS diet produces arterial hypertension and the HFHS diet causes a vascular dysfunctional state that was highly responsive to α-adrenergic stimulation.

Effects of chronic methamphetamine exposure on heart function in uninfected and retrovirus-infected mice.

Methamphetamine (MA) increases catecholamine levels, which have detrimental effects on heart function through vasoconstriction, myocardial hypertrophy, and fibrosis. Murine retrovirus infection induces dilated cardiomyopathy (DCM). The present study investigated the cardiovascular effects of chronic MA treatment on uninfected and retrovirus-infected mice. C57BL/6 mice were studied after 12 weeks treatment. The four study groups were (group I) uninfected, MA placebo; (group II) infected, MA placebo; (group III) uninfected, MA treatment; and (group IV) infected and MA treatment. MA injections were given i.p. once a day for 5 days/week with a increasing dose from 15 mg/kg to 40 mg/kg. Left ventricular mechanics were measured in situ a using Millar conductance catheter system for pressure-volume loop analysis. Cardiac pathology was determined with histological analysis. In the uninfected mice, the load independent contractile parameters, pre-load recruitable stroke work (PRSW) and dP/dt(max) vs. Ved, significantly decreased by 32% and 35% in MA treated mice when compared to the saline injected mice. In retrovirus-infected mice, although there were no significant difference in Ees, PRSW, and dP/dt(max) vs. Ved due to MA treatment, they were increased 45%, 15% and 42% respectively when compared to saline treated mice. No further lowered heart function during murine AIDS may be due to the counteraction of the retroviral DCM and the MA induced myocardial fibrosis and hypertrophy (thickening of the ventricular walls). This is supported by increases in the End-diastolic volume (Ved, 38%) and End-systolic volume (Ves, 84%) in the retrovirus-infected saline injected mice, the decreases of 33% and 17% in the uninfected MA-treated mice, but no significant changes in the retrovirus-infected MA treated mice when compared to uninfected saline injected mice. These data suggest that MA induced myocardial cellular changes compensate for retrovirus induced DCM.

T-lymphocytes mediate left ventricular fibrillar collagen cross-linking and diastolic dysfunction in mice

Aberrant concentrations of cardiac extracellular matrix (ECM) fibrillar collagen cross-linking have been proposed to be an underlying cause of cardiac diastolic dysfunction however the role of the adaptive immune system in this process has yet to be investigated. Fibrillar collagen cross-linking is a product of the enzymatic activities of lysyl oxidase (LOX and LOXL-3) released by the cardiac fibroblast and possibly cardiac myocytes. Our hypothesis is that stimulation of the TH1 lymphocytes activates lysyl oxidase mediated ECM cross-linking and thereby alters left ventricular function. Three-month old C57BL/J female mice were treated with selective TH1 lymphocyte inducers - T-cell receptor Vβ peptides (TCR). After 6 weeks, candidate gene expression, tissue enzymatic activity, ECM composition, and left ventricular mechanics were quantified. Lymphocyte gene expression and cytokine assay revealed TH1 immune polarization with TCR administration which was associated with a 2.6-fold and 3.1-fold increase of LOX and LOXL3 gene expression, respectively, and a 55% increase in cardiac LOX enzymatic activity. The ECM cross-linked fibrillar collagen increased by 95% when compared with the control. Concurrently, there was a 33% increased ventricular stiffness, decreased cardiac output, and normal ejection fraction. These data implicate the TH1 lymphocyte in the pathogenesis of diastolic dysfunction which has potential clinical application in the pathogenesis of diastolic heart failure.

Effect of angiotensin II on primary cardiac fibroblast matrix metalloproteinase activities

Left ventricular dysfunction is associated with reperfusion injury occurring during open-heart surgery. There is an increased secretion of angiotensin II (Ang II) and increased matrix metalloproteinases (MMPs) activities associated with open-heart surgery that may affect the cardiac extracellular matrix (ECM). The goal of this study was to determine the effects of Ang II and selective angiotensin II receptor (AT1-R and AT2-R) blockers on the enzymatic activities of MMPs in primary adult murine cardiac fibroblasts (CF). Our hypothesis is that Ang II, with and without a selective receptor blocker, differentially affects CF MMPs activities. The CF were treated with Ang II (10-6 M) and doses of AT1-R and AT2-R blockers (losartan and PD123319, respectively) at doses of 10-7 to 10-5 M for 48 hours. The Ang IIstimulated CF reduced collagenase activities by only 24% (p =0.004); however, the MMP-2 and MMP-9 gelatinase activities were reduced by 42% and 39%, respectively (p =0.022). The losartan dose dependently increased MMP-2 (p =0.02) and MMP-9 (ns). PD123319 at 10-5 M significantly reduced MMP-2 and MMP-9 activities compared with the Ang II group (p =0.014 and p =0.02, respectively). The doses of PD123319 at 10-6 and 10-7 M increased the MMP-2 and MMP-9 enzymatic activities significantly above the Ang II only group. Thus, Ang II and AT1-R and AT2-R differentially affect the collagenase and gelatinase MMPs activities released by cardiac fibroblasts. Perfusion (2007) 22, 51-55.