Tissue factor deficiency causes cardiac fibrosis and left ventricular dysfunction.

Exposure of blood to tissue factor (TF) activates the extrinsic (TF:FVIIa) and intrinsic (FVIIIa:FIXa) pathways of coagulation. In this study, we found that mice expressing low levels of human TF ( approximately 1% of wild-type levels) in an mTF(-/-) background had significantly shorter lifespans than wild-type mice, in part, because of spontaneous fatal hemorrhages. All low-TF mice exhibited a selective heart defect that consisted of hemosiderin deposition and fibrosis. Direct intracardiac measurement demonstrated a 30% reduction (P < 0.001) in left ventricular function in 8-month-old low-TF mice compared with age-matched wild-type mice. Mice expressing low levels of murine FVII ( approximately 1% of wild-type levels) exhibited a similar pattern of hemosiderin deposition and fibrosis in their hearts. In contrast, FIX(-/-) mice, a model of hemophilia B, had normal hearts. Cardiac fibrosis in low-TF and low-FVII mice appears to be caused by hemorrhage from cardiac vessels due to impaired hemostasis. We propose that TF expression by cardiac myocytes provides a secondary hemostatic barrier to protect the heart from hemorrhage.

Targeted gene disruption of matrix metalloproteinase-9 (gelatinase B) suppresses development of experimental abdominal aortic aneurysms.

Abdominal aortic aneurysms represent a life-threatening condition characterized by chronic inflammation, destructive remodeling of the extracellular matrix, and increased local expression of matrix metalloproteinases (MMPs). Both 92-kD gelatinase (MMP-9) and macrophage elastase (MMP-12) have been implicated in this disease, but it is not known if either is necessary in aneurysmal degeneration. We show here that transient elastase perfusion of the mouse aorta results in delayed aneurysm development that is temporally associated with transmural mononuclear inflammation, increased local production of several elastolytic MMPs, and progressive destruction of the elastic lamellae. Elastase-induced aneurysmal degeneration was suppressed by treatment with a nonselective MMP inhibitor (doxycycline) and by targeted gene disruption of MMP-9, but not by isolated deficiency of MMP-12. Bone marrow transplantation from wild-type mice prevented the aneurysm-resistant phenotype in MMP-9-deficient animals, and wild-type mice acquired aneurysm resistance after transplantation from MMP-9-deficient donors. These results demonstrate that inflammatory cell expression of MMP-9 plays a critical role in an experimental model of aortic aneurysm disease, suggesting that therapeutic strategies targeting MMP-9 may limit the growth of small abdominal aortic aneurysms.

Inflammatory cell-derived NO modulates cardiac allograft contractile and electrophysiological function.

We previously demonstrated that inhibition of inducible nitric oxide (NO) synthase (iNOS) ameliorated acute cardiac allograft rejection. This study used a rat cardiac transplant model to characterize contractile and electrophysiological dysfunction during early acute rejection, further examine the role of NO and iNOS in this process, and determine which cells expressed iNOS during early rejection. During early acute rejection, before significant myocyte necrosis, allograft papillary muscles had reduced tension development and rates of tension development and decline during beta-adrenergic, adenylate cyclase, and calcium stimulation compared with isograft and normals [e.g., tension of 36 (allograft) vs. 73 (isograft) mN/mm2 during calcium stimulation, P < 0.001]. Allografts had resting membrane potential depolarization and reduced action potential amplitude and upstroke velocity. iNOS mRNA was expressed in infiltrating inflammatory cells but not in allograft myocytes, endothelial cells, or isografts. Corticosteroids attenuated allograft contractile and electrophysiological dysfunction and inhibited iNOS enzyme activity. Direct iNOS inhibition with aminoguanidine inhibited NO production and prevented allograft contractile and electrophysiological dysfunction (e.g., tension of 64 mN/mm2 during calcium stimulation, P < 0.001). We conclude that 1) early allograft rejection caused contractile and electrophysiological dysfunction that was largely mediated by iNOS expression in infiltrating inflammatory cells, 2) corticosteroid-mediated amelioration of allograft contractile and electrophysiological dysfunction may reflect inhibition of iNOS, and 3) iNOS inhibition may offer an alternative in management of immune-mediated myocardial dysfunction.

Ventricular myocytes isolated from rejecting cardiac allografts exhibit a reduced beta-adrenergic contractile response.

Previous experiments using multicellular preparations from rejecting transplanted animal hearts have indicated that a decrease in the contractile response to beta-adrenergic stimulation accompanies acute rejection. The precise mechanism of this decreased beta-adrenergic response, which may limit the inotropic reserve in human transplant patients during rejection, is currently unknown. In order to determine whether the decreased beta-adrenergic response is an intrinsic property of the cardiac myocytes from rejecting hearts, we examined the effects of beta-adrenergic stimulation (isoproterenol, 10(-8) and 10(-6) M) on the cell shortening of single myocytes isolated from native rat hearts (untransplanted Lewis strain), non-rejecting isografts (Lewis to Lewis heterotopic transplant), and rejecting allografts (Lewis to ACI transplant). The myocytes from isografts demonstrated a significantly increased contractile response to isoproterenol compared to native myocytes (presumably due to denervation supersensitivity), whereas the myocytes from allografts demonstrated a greatly decreased response to isoproterenol compared to both native and isograft myocytes. These results demonstrate that the decreased beta-adrenergic contractile response is an intrinsic property of the rejecting cardiac myocyte.

Major histocompatibility complex-specific prolongation of murine skin and cardiac allograft survival after in vivo depletion of V beta+ T cells.

The preferential usage of certain T cell receptor (TCR) V beta genes has been well established in several major histocompatibility complex (MHC)-restricted immune responses. However, V beta usage among allogeneic responses remains unclear. Because recent findings of ours and others indicate that V beta 8 predominates in certain Ld-restricted, peptide-specific responses, we examined the V beta 8 usage in allogeneic responses to Ld. To selectively recognize the Ld molecule, cells from BALB/c-H-2dm2 (dm2), the Ld-loss mutant mouse, were stimulated in vitro or in vivo with wild-type BALB/c cells. We report here that after the intraperitoneal administration of the anti-V beta 8 monoclonal antibody (mAb) F23.1, peripheral V beta 8 T cells were depleted from dm2 mice. This in vivo depletion abrogated the ability of dm2 splenocytes to mount a primary response to Ld molecules. This abrogation was specific, since the response of V beta 8-depleted dm2 cells to Kb/Db antigens was the same as that of control nondepleted dm2 cells. Furthermore, in vivo depletion of V beta 8 cells was found to cause a dramatic prolongation of Ld-disparate skin grafts (mean survival time [MST] 22.1 +/- 2.1 vs. 10.3 +/- 1.1 d for saline-treated controls, or 10.9 +/- 1.7 d for controls treated with mAb KJ23 to V beta 17). By contrast, V beta 8 depletion had no effect on recipients grafted with haplotype-mismatched skin or single Dk-locus-disparate skin. These findings demonstrate that V beta 8+ T cells predominate in allogeneic response to Ld but not other alloantigens. The effect of V beta 8 depletion was found to be even more dramatic on recipients grafted with Ld-disparate vascularized heart transplants (MST > 100 vs. 8.6 +/- 0.5 d for controls). In total, these findings establish the efficacy of using mAb to the V beta gene family to specifically and significantly enhance the survival of allografts. The implications of detecting V beta 8 usage in both alloreactive or MHC-restricted TCR responses to the same class I molecule are discussed.

Influence of activation origin, lead number, and lead configuration on the noninvasive electrophysiologic detection of cardiac allograft rejection.

An improved method of cardiac allograft surveillance, less invasive than endomyocardial biopsy (EMB), is required as the scope of cardiac transplantation increases. The correlation between changes in telemetrically recorded intramyocardial electrograms and the degree of rejection was examined. The electrophysiological parameter assessed was the unipolar peak-to-peak amplitude (UPPA), defined as the magnitude of the first, fast negative deflection of the QRS complex in unipolar electrograms. A canine model of intrathoracic heterotopic cardiac transplantation was employed in which each graft was instrumented with four unipolar intramyocardial electrodes (two left ventricle [LV] and two right ventricle [RV]) connected to two telemetric pacemakers. Immunosuppression was begun at operation and continued for a variable period of time following the acquisition of control data (postoperative days 6-10). Graft status was monitored via biweekly EMB. Intramyocardial electrograms were recorded from each lead during sinus rhythm and 36 atrial and ventricular pacing protocols on a daily basis. The daily UPPA was calculated for each rhythm sequence and expressed as percent of control UPPA. EMBs were qualitatively graded and assigned a quantitative rejection score (0 = none; 3 = severe). Criteria for rejection were rejection score (RS) greater than or equal to 0.66 or daily UPPA less than 85% control. Eight animals were included for study and, in all animals, rejection was documented. A total of 30 EMBs were obtained; in 11 EMBs rejection was present by histological criteria. Analysis of grouped UPPA data from all four leads revealed a sensitivity and specificity of 91% and 84%, respectively, in the detection of rejection. Analysis of grouped sinus UPPA data from the two LV leads revealed a sensitivity of 94% and 91%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)