Lianne S.M. Boesten

Mdm2, but not Mdm4, protects terminally differentiated smooth muscle cells from p53-mediated caspase-3-independent cell death

p53 is a potent inhibitor of cell growth and an inducer of apoptosis. During embryonic development, Mdm2 and Mdm4 inhibit the growth suppressive activities of p53. However, whether tight surveillance of p53 activity is required in quiescent cells is unknown. To test this, conditional inactivation of mdm2 and mdm4 was carried out in smooth muscle cells (SMCs). Upon SMC-specific inactivation of mdm2, and not of mdm4, mice rapidly became ill and died. Necropsy showed small intestinal dilation, and histological analyses indicated a severe reduction in the number of intestinal SMCs. Increased p53 levels and activity were detected in the remaining SMCs, and the phenotype was completely rescued on a p53-null background. Interestingly, intestinal SMCs are caspase-3-negative and therefore did not undergo caspase-3-dependent apoptotic cell death. Together, Mdm2, but not Mdm4, prevents accumulation of active p53 in quiescent SMCs and thereby the induction of p53-mediated caspase-3-independent cell death.

Macrophage Low-Density Lipoprotein Receptor-Related Protein Deficiency Enhances Atherosclerosis in ApoE/LDLR Double Knockout Mice

Objective—In vitro studies implicate that the low-density lipoprotein receptor (LDLR)-related protein (LRP) in macrophages has a pro-atherogenic potential. In the present study, we investigated the in vivo role of macrophage specific LRP in atherogenesis independent of its role in the uptake of lipoproteins. Methods and Results—We generated macrophage-specific LRP-deficient mice on an apoE/LDLR double-deficient background. Macrophage LRP deletion did not affect plasma cholesterol and triglyceride levels, lipoprotein distribution, and blood monocyte counts. Nevertheless, macrophage LRP deficiency resulted in a 1.8-fold increase in total atherosclerotic lesion area in the aortic root of 18-week-old mice. Moreover, LRP deficiency also resulted in a relatively higher number of advanced lesions. Whereas macrophage and smooth muscle cell content did not differ between LRP-deficient mice and control littermates, a 1.7-fold increase in collagen content and 2.3-fold decrease in relative number of CD3+ T cells were observed in lesions from macrophage specific LRP-deficient mice. Conclusions—Our data demonstrate that independent of its role in lipoprotein uptake, absence of LRP in macrophages resulted in more advanced atherosclerosis and in lesions that contained more collagen and less CD3+ T cells. In contrast to previous in vitro studies, we conclude that macrophage LRP has an atheroprotective potential and may modulate the extracellular matrix in the atherosclerotic lesions.

Preoperative predictors of postsurgical adhesion formation and the Prevention of Adhesions with Plasminogen Activator (PAPA-study): results of a clinical pilot study

OBJECTIVE To identify predictors of postsurgical adhesion formation in peritoneal fluid and plasma, and assess efficacy and safety of reteplase (recombinant plasminogen activator [r-PA]). DESIGN Prospective randomized study. SETTING University Medical Center. PATIENT(S) Twenty-six abdominal myomectomy patients with early second-look laparoscopy (ESL). INTERVENTION(S) Randomization to IP treatment with 1 mg reteplase in 300 mL Ringers lactate or 300 mL Ringers lactate only. Scoring of adhesions and collecting peritoneal fluid during both surgical procedures and collecting plasma samples at ten time points. MAIN OUTCOME MEASURE(S) Incidence, severity, and extent of adhesions at ESL. Concentrations of C-reactive protein (CRP), tissue-type plasminogen activator (tPA), plasminogen activator inhibitor 1 (PAI-1), and fibrin degradation products (FbDPs). RESULT(S) Significant correlation between the extent of uterine adhesion formation and preoperative plasma levels of CRP (r(s) = 0.558), PAI-1 (r(s) = 0.413), and the change in tPA concentration in peritoneal fluid from initial surgery to ESL (Delta+PA: r(s) = -0.636). No significant differences in adhesion scores between treatment and control groups. CONCLUSION(S) Our finding that preoperative plasma CRP and PAI-1-levels are significantly correlated with extent of adhesion formation points to a role of chronic inflammation in the disease process. Results are highly indicative for the paradigm that adhesions are caused by an insufficiency in peritoneal fibrinolytic capacity. For successful adhesion prevention therapy relatively high amounts of r-PA are required.

Dual PPARα/γ Agonist Tesaglitazar Reduces Atherosclerosis in Insulin-Resistant and Hypercholesterolemic ApoE*3Leiden Mice

Objective—We investigated whether the dual PPAR&agr;/&ggr; agonist tesaglitazar has anti-atherogenic effects in ApoE*3Leiden mice with reduced insulin sensitivity. Methods and Results—ApoE*3Leiden transgenic mice were fed a high-fat (HF) insulin-resistance–inducing diet. One group received a high-cholesterol (HC) supplement (1% wt/wt; HC group). A second group received the same HC supplement along with tesaglitazar (T) 0.5 &mgr;mol/kg diet (T group). A third (control) group received a low-cholesterol (LC) supplement (0.1% wt/wt; LC group). Tesaglitazar decreased plasma cholesterol by 20% compared with the HC group; cholesterol levels were similar in the T and LC groups. Compared with the HC group, tesaglitazar caused a 92% reduction in atherosclerosis, whereas a 56% reduction was seen in the cholesterol-matched LC group. Furthermore, tesaglitazar treatment significantly reduced lesion number beyond that expected from cholesterol lowering and induced a shift to less severe lesions. Concomitantly, tesaglitazar reduced macrophage-rich and collagen areas. In addition, tesaglitazar reduced inflammatory markers, including plasma SAA levels, the number of adhering monocytes, and nuclear factor &kgr;B-activity in the vessel wall. Conclusions—Tesaglitazar has anti-atherosclerotic effects in the mouse model that go beyond plasma cholesterol lowering, possibly caused by a combination of altered lipoprotein profiles and anti-inflammatory vascular effects.

Macrophage p53 controls macrophage death in atherosclerotic lesions of apolipoprotein E deficient mice

The cellular composition of atherosclerotic lesions is determined by many factors including cell infiltration, proliferation and cell death. Tumor suppressor gene p53 has been shown to regulate both cell proliferation and cell death in many cell types. In the present study, we investigated the role of macrophage p53 in the pathogenesis of early and advanced atherosclerosis. Using the Cre-loxP system we found that absence of macrophage p53 (p53(del)) strongly reduces apoptosis of macrophages both in early and advanced atherosclerotic lesions (-59% and -37%, respectively). Consequently, in advanced atherosclerosis, reduced apoptosis upon absence of macrophage p53, coincided with increased acellular necrotic core formation (+96%), increased macrophage content (+24%), and reduced cholesterol cleft accumulation (-41%). Proliferation was not affected by the absence of macrophage p53 in both early and advanced atherosclerosis. However, these significant changes in lesional cell death did not affect total lesion area in both early and advanced atherosclerosis, neither in the aortic root nor in the aortic arch and thoracic aorta in ApoE-deficient mice. Our data demonstrate that macrophage p53 is an important regulator of macrophage apoptosis, thereby preventing necrotic death of lesional macrophages. The regulation of this cell death balance directly affects lesion composition.

Macrophage retinoblastoma deficiency leads to enhanced atherosclerosis development in ApoE-deficient mice

The cellular composition of an atherosclerotic lesion is determined by cell infiltration, proliferation, and apoptosis. The tumor suppressor gene retinoblastoma (Rb) has been shown to regulate both cell proliferation and cell death in many cell types. To study the role of macrophage Rb in the development of atherosclerosis, we used apoE‐deficient mice with a macrophage‐restricted deletion of Rb (Rbdel mice) and control littermates (Rbfl mice). After 12 wk feeding a cholesterol‐rich diet, the Rbdel mice showed a 51% increase in atherosclerotic lesion area with a 39% increase in the relative number of advanced lesions. Atherosclerotic lesions showed a 13% decrease in relative macrophage area and a 46% increase in relative smooth muscle cell area, reflecting the more advanced state of the lesions. The increase in atherosclerosis was independent of in vitro macrophage modified lipoprotein uptake or cytokine production. Whereas macrophage‐restricted Rb deletion did not affect lesional macrophage apoptosis, a clear 2.6‐fold increase in lesional macrophage proliferation was observed. These studies demonstrate that macrophage Rb is a suppressing factor in the progression of atherosclerosis by reducing macrophage proliferation.—Boesten, L. S. M., Zadelaar, A. S. M., van Nieuwkoop, A., Hu, L., Jonkers, J., van de Water, B., Gijbels, M. J. J. van der Made, I., de Winther, M. P. J., Havekes, L. M., van Vlijmen, B. J. M. Macrophage retinoblastoma deficiency leads to enhanced atherosclerosis development in ApoE‐deficient mice. FASEB J. 20, E18–E26 (2006)