Yi Fu Zhou

Transendocardial delivery of autologous Bone marrow enhances collateral perfusion and regional function in pigs with chronic experimental myocardial ischemia

OBJECTIVES We tested the hypothesis that intramyocardial injection of autologous bone marrow (ABM) promotes collateral development in ischemic porcine myocardium. We also defined, in vitro, whether bone marrow (BM) cells secrete vascular endothelial growth factor (VEGF) and macrophage chemoattractant protein-1 (MCP-1). BACKGROUND The natural processes leading to collateral development are extremely complex, requiring multiple growth factors interacting in concert and in sequence. Because optimal angiogenesis may, therefore, require multiple angiogenic factors, we thought that injection of BM, which contains cells that secrete numerous angiogenic factors, might provide optimal therapeutic angiogenesis. METHODS Bone marrow was cultured four weeks in vitro. Conditioned medium was assayed for VEGF and MCP-1 and was added to cultured pig aortic endothelial cells (PAEC) to assess proliferation. Four weeks after left circumflex ameroid implantation, freshly aspirated ABM (n = 7) or heparinized saline (n = 7) was injected transendocardially into the ischemic zone (0.2 ml/injection at 12 sites). Echocardiography to assess myocardial thickening and microspheres to assess perfusion were performed at rest and during stress. RESULTS Vascular endothelial growth factor and MCP-1 concentrations increased in a time-related manner. The conditioned medium enhanced, in a dose-related manner, PAEC proliferation. Collateral flow (ischemic/normal zone X 100) improved in ABM-treated pigs (ABM: 98 +/- 14 vs. 83 +/- 12 at rest, p = 0.001; 89 +/- 18 vs. 78 +/- 12 during adenosine, p = 0.025; controls: 92 +/- 10 vs. 89 +/- 9 at rest, p = 0.49; 78 +/- 11 vs. 77 +/- 5 during adenosine, p = 0.75). Similarly, contractility increased in ABM-treated pigs (ABM: 83 +/- 21 vs. 60 +/- 32 at rest, p = 0.04; 91 +/- 44 vs. 36 +/- 43 during pacing, p = 0.056; controls: 69 +/- 48 vs. 64 +/- 46 at rest, p = 0.74; 65 +/- 56 vs. 37 +/- 56 during pacing, p = 0.23). CONCLUSIONS Bone marrow cells secrete angiogenic factors that induce endothelial cell proliferation and, when injected transendocardially, augment collateral perfusion and myocardial function in ischemic myocardium.

Association between Prior Cytomegalovirus Infection and the Risk of Restenosis after Coronary Atherectomy

BACKGROUND Restenosis occurs commonly after coronary angioplasty and atherectomy, but the causes of restenosis are poorly understood. Recently, it has been found that cytomegalovirus (CMV) DNA is present in restenotic lesions from atherectomy specimens. This and other evidence suggest that CMV may have a role in the process of restenosis. METHODS We prospectively studied 75 consecutive patients undergoing directional coronary atherectomy for symptomatic coronary artery disease. Before atherectomy was performed, we measured blood levels of anti-CMV IgG antibodies to determine whether previous exposure to CMV increased the risk of restenosis, as determined by coronary angiography performed six months after atherectomy. RESULTS After atherectomy, the mean (+/- SD) minimal luminal diameter of the target vessel was greater in the 49 patients who were seropositive for CMV than in the 26 patients who were seronegative (3.18 +/- 0.51 mm vs. 2.89 +/- 0.45 mm, P=0.01). After six months, however, the seropositive patients had a greater reduction in the luminal diameter (1.24 +/- 0.83 mm vs. 0.68 +/- 0.69 mm, P = 0.003), resulting in a significantly higher rate o restenosis in the seropositive patients (43 percent vs. 8 percent, P = 0.002). In a multivariable logistic-regression model, CMV seropositivity and the CMV titer were independently predictive of restenosis (odds ratios, 12.9 and 8.1, respectively). There was no evidence of acute infection, since the titer of anti-CMV IgG antibodies did not increase over time and tests for anti-CMV IgM antibodies were negative in all patients. CONCLUSIONS Prior infection with CMV is strong independent risk factor for restenosis after coronary atherectomy. If confirmed, these findings may help identify patients at risk for restenosis.

Infection and Atherosclerosis Potential Roles of Pathogen Burden and Molecular Mimicry

Infection has been implicated as a cause of atherosclerosis since the first half of the 19th century. Over the years, sporadic publications have appeared in the literature reflecting a persistent but relatively low level of research activity in this area. In the last decade, however, publications relating to this topic have increased markedly. And very recently, new epidemiological and mechanistic data relating infection to several different diseases, including atherosclerosis, have appeared, stimulating the emergence of important paradigm shifts in how we think about the causes of chronic disease. The following article reviews some of these newer concepts as they relate to a possible role of infection in atherosclerosis.

Cytomegalovirus infection increases development of atherosclerosis in Apolipoprotein-E knockout mice

Cytomegalovirus (CMV) infection has been associated with coronary artery disease, but it is unknown whether the virus can causally contribute to atherogenesis. To determine whether the virus has this capacity, we infected an atherosclerotic-prone mouse strain (C57BL/6J apoE-/-) with murine CMV. At 14 days of age, 30 mice received CMV (30000 pfu) ip and 30 received virus free media. At 13 and 16 weeks atherosclerotic lesion size was measured from aortic sinus cross-sections. Infection did not alter plasma levels of cholesterol, triglycerides, and high density lipoprotein (HDL); however, 4 weeks after infection IFNgamma levels were elevated (infection vs control: 156+/-49 vs 50+/-22 pg/ml, P=0.04). No differences in lesion size were present at 13 weeks post infection. However, by 16 weeks mean aortic sinus lesion area (mm(2)x10(3)+/-SEM; N=75) in the CMV-infected mice was significantly greater than in uninfected mice (74+/-6 vs 57+/-6; P=0.04). CMV caused the greatest increase (34%) in lesion size in females (103+/-9 vs 77+/-10; P=0.05; N=35). These results provide additional evidence implicating CMV as a causal agent of atherosclerosis, at least in an animal model.

Human cytomegalovirus increases modified low density lipoprotein uptake and scavenger receptor mRNA expression in vascular smooth muscle cells.

Evidence suggests a possible role for human cytomegalovirus (HCMV) in the development of arteriosclerosis. One of the earliest events in plaque formation is the accumulation of lipid-laden foam cells, derived from macrophages and smooth muscle cells (SMCs). The lipid accumulation that occurs depends upon the uptake of oxidized LDL (Ox-LDL), a process in which the scavenger receptor (SR) has been postulated to play an important role. We therefore examined the effects of HCMV on this process. We demonstrate that HCMV infection of human SMCs increases modified LDL uptake and stimulates class A SR gene (SR-A) mRNA expression. In addition, infection of rat SMCs with HCMV, which causes immediate early gene expression (IE72/IE84), but no early or late HCMV gene products and no cytopathic effects, also increases SMC uptake of Ox-LDL and acetylated LDL, with either effect blocked by an excess of either cold Ox-LDL or acetylated-LDL, and by fucoidin, an SR competitor. Cotransfection of an IE72, but not an IE84, expression plasmid and a plasmid containing a Class A SR promoter/reporter gene construct enhances SR promoter activity. Since increased Ox-LDL uptake is believed to play an important role in arteriosclerosis, these results provide a link between HCMV infection and arteriosclerotic plaque formation.

Akt Controls Vascular Smooth Muscle Cell Proliferation In Vitro and In Vivo by Delaying G1/S Exit

Abstract— Constitutive activation of serine/threonine kinase Akt causes uncontrolled cell-cycle progression in different cell types and in malignancy. To investigate how Akt activation modulates cell-cycle progression in vascular smooth muscle cells (SMCs) in vitro and in the intact animal, we inhibited Akt-dependent signaling by adenovirus-mediated transfection of a dominant-negative Akt mutant (AA-Akt). We observed reduced proliferation rate (P <0.01), DNA synthesis (P <0.01), and a significant arrest in G1/S exit (P <0.01) both in vitro in response to serum stimulation and in vivo after vascular injury. In vivo transfection of the balloon-injured vessel with AA-Akt reduced SMC proliferation, resulting in decreased neointima compared with control virus (P <0.01). These effects were at least in part modulated, both in vitro and in vivo, by increased p21Cip1 expression, as demonstrated by lack of effect of AA-Akt on cell proliferation in p21−/− mouse SMCs. In conclusion, this study demonstrates that Akt-dependent signaling enhances cell-cycle progression of nontransformed SMCs in vitro and in response to vascular injury in the intact animal. These results suggest a role for Akt signaling in modulating the response of normal tissues to stress and the response of the arterial wall to acute and possibly repetitive injuries that ultimately contribute to restenosis and atherosclerosis.

Effects of MF-tricyclic, a selective cyclooxygenase-2 inhibitor, on atherosclerosis progression and susceptibility to cytomegalovirus replication in apolipoprotein-E knockout mice ☆

OBJECTIVES We examined whether selective cyclooxygenase-2 (COX-2) inhibition in apolipoprotein-E (apoE) deficient mice reduces cytomegalovirus (CMV) replication, and determined whether COX-2 anti-inflammatory activity leads to decreased atherosclerosis. BACKGROUND Evidence suggests that CMV infection contributes to atherosclerosis and that this occurs in part through inflammatory mechanisms. Cyclooxygenase-2 inhibitors are potent anti-inflammatory agents. They also inhibit CMV replication in vitro. METHODS The apoE deficient mice were either treated or not treated with a selective COX-2 inhibitor, and either infected or not infected with CMV. Viral deoxyribonucleic acid load in salivary glands was determined by quantitative polymerase chain reaction. Atherosclerotic lesion analysis was performed by standard methods. RESULTS In vivo COX-2 inhibition, unexpectedly increased viral load: in the CMV-infected animals viral load was 2.58 +/- 1.0 in the nontreated group, 4.74 +/- 1.38 in the group treated with 12 mg/kg/day MF-tricyclic, and 6.51 +/- 1.64 in the group treated with 24 mg/kg/day MF-tricyclic (p trend = 0.050). This increased viral load was paralleled by increased anti-CMV antibody titers. Most surprisingly, COX-2 inhibition significantly increased early atherosclerotic lesion area, independent of viral infection. CONCLUSIONS Our study demonstrates that selective inhibition of COX-2 in vivo increases viral load. The finding that inhibition of COX-2 increases atherosclerosis development in apoE deficient mice suggests, unexpectedly, that this enzyme exerts antiatherosclerosis activity, at least in this model.

Lack of association of restenosis following coronary angioplasty with elevated C-reactive protein levels or seropositivity to chlamydia pneumoniae

Seventy-five consecutive patients undergoing directional coronary atherectomy were evaluated by measuring anti-Chlamydia immunoglobulin G and anticytomegalovirus immunoglobulin G antibodies, and serum levels of C-reactive proteins (before atherectomy). The results showed that although both Chlamydia infection and elevated C-reactive protein levels are associated with coronary artery disease and coronary artery disease events, neither of these appears to play a role in the development of restenosis.

Chronic non-vascular cytomegalovirus infection: effects on the neointimal response to experimental vascular injury

OBJECTIVE Epidemiologic and mechanistic evidence implicates a role for cytomegalovirus (CMV) in atherogenesis. Recently, we demonstrated that CMV has the capacity to causally contribute to atherogenesis; acute infection of rats with rat CMV (RCMV) 1 day after carotid artery injury increased neointimal accumulation. Importantly, in the injured vessel infectious virus could not be detected and viral genome was present only transiently, suggesting that additional mechanisms play a role in the virus-induced exacerbation of the vascular injury response other than the changes caused by direct infection of vessel wall cells. The present investigation was designed to determine whether chronic persistent RCMV infection, more relevant to the clinical situation, also exacerbates the response to injury and, if so, whether similar mechanisms are operative. METHODS Sixty 3-week-old male Spraque-Dawley rats received an i.p. injection of either 10(6) TCID50 RCMV (Priscott strain) or normal saline. The left carotid artery was balloon-injured 3 months after infection. Rats were killed 6 weeks later. This model produces persistent infection, as demonstrated by presence of infectious virus in the salivary glands at time of sacrifice. RESULTS The neointima to media (N/M) ratio of the injured vessel was 41% greater in the RCMV-infected than in control rats (1.40 +/- 0.48 vs. 0.99 +/- 0.45; P = 0.003). The aorta never contained infectious RCMV, and exhibited RCMV DNA, detected by PCR, only transiently. The persistent infection of non-vascular tissues was associated with increased serum levels of IL-2, IL-4 and IFN-gamma. CONCLUSIONS CMV infection of young rats causes persistent infection of non-vascular tissues and increased cytokine levels. The neointimal response to subsequent vascular injury is increased, despite absence of virus from the vessel wall. These findings, as in acute infection following vascular injury, suggest that inflammatory and immune responses to chronic persistent CMV infection contribute to an exaggerated response to vascular injury.

Host Response Differences to Infection May Affect Disease susceptibility in atherosclerotic patients

Publisher Summary Epidemiological studies for infections in human populations have shown the associations between atherosclerotic disease and positive serology. Despite not all authors reported positive associations, such studies have made essential contributions to the totality of evidence. Basic researches have demonstrated pathogen antigens or nucleic acids in arterial tissues taken from patients undergoing vascular operation or autopsy. These data supported the epidemiological findings, and provided direct observation of what actually happens in human. Although a clear mechanism for the infection in the development atherosclerosis has not been established, numerous investigations that have attempted to link atherosclerosis in humans with infection with various pathogens, such as CMV, Chlamydia pneumoniae and Helicobacter pylori, have led to the suggestion that infection-related cardiovascular disease is an inflammation and immune reaction-mediated process. The susceptibility to the atherogenic effects of infectious agents depend, at least in part, on the capacity of the host to pathogen induced inflammatory activity or immunr responses. Therefore, it is likely that a complete understanding of the potential role of infection in atherosclerosis will be achieved only after the more complex interrelations between a pathogen and the hosts response to the pathogen are elucidated.