Electrocardiographic characterization of stress‐induced myocardial infarction in atherosclerotic mice

Aim: We have previously shown that mental and hypoxic stress can trigger the development of myocardial infarction (MI) in atherosclerotic apoE^?/? × LDLR^?/? mice. The purpose of the present study was to characterize the interval between stress and MI and determine whether electrophysiological changes precede the precipitation of an infarct by assessing telemetry recordings of the electrocardiogram. Methods: Isoflurane anaesthetized apoE^?/? × LDLR^?/? (n = 16) and C57BL/6J (n = 8) mice were exposed to systemic hypoxia by reducing the inhaled oxygen concentration to 10% for 10 min. Mental stress was induced in eight conscious apoE^?/? × LDLR^?/? and eight C57BL/6J mice by blowing air into the cage. Physiological parameters were recorded every 30 min for 2–6 days by implanted transmitters. Results: During stress all mice developed transient ischaemic STU‐area changes, which returned to normal at the end of stress. During the recovery phase (6 days) 50% (4/8) of the mentally stressed apoE^?/? × LDLR^?/? mice developed increased STU‐area variability (P < 0.05) followed by dramatic STU‐area elevations and spontaneous death at approximately 12–24 h. In hypoxia‐exposed apoE^?/? × LDLR^?/? mice 56% (9/16) developed MI as determined by elevated serum levels of the infarction marker troponin T which correlated with increased variability in the STU‐area (P < 0.05). Conclusion: This is the first mouse model showing that increased STU‐area variability is indicative of MI development in atherosclerotic mice following ischaemic stress. Furthermore, our findings suggest a two‐phase pathway for the infarction development: an initial phase comprising a transient ischaemic response which triggers a delayed second phase of ischaemia and MI.     

载脂蛋白E基因敲除小鼠动脉粥样硬化早期血管外膜成纤维细胞增殖活性增强

目的:探讨动脉外膜成纤维细胞增殖与早期动脉粥样硬化病灶形成的关系。方法:选择6周龄载脂蛋白E基因敲除[apoE(-/-)]小鼠和野生型C57BL/6小鼠,高脂喂养2、4和10周后,在各个时点处死动物前24 h经腹腔注射5-溴-2-脱氧尿嘧啶(BrdU),后选取升主动脉制备连续切片,通过HE染色观察组织形态学的变化,用免疫组化方法观察不同时点血管外膜及内膜BrdU的表达变化。体外培养高脂喂养2周的apoE(-/-)小鼠和C57BL/6小鼠动脉外膜成纤维细胞,通过BrdU掺入法测定细胞增殖活性,流式细胞术测定细胞周期。结果:体内实验发现apoE(-/-)小鼠高脂喂养2周后,在无可见内膜病灶形成之前,首先在主动脉外膜发现BrdU标记的阳性细胞,之后才在损伤内膜观察到BrdU标记细胞。而C57BL/6小鼠在任何时点都未检测到BrdU标记的细胞。体外实验观察到apoE(-/-)小鼠血管外膜成纤维细胞BrdU标记的细胞数显著多于C57BL/6小鼠(P0.01),apoE(-/-)小鼠血管外膜成纤维细胞S期及G2/M期所占百分比明显高于对照组(P0.05)。结论:血管外膜成纤维细胞增殖可能参与早期动脉粥样硬化病灶形成。     

apoE/LDLR双基因突变小鼠肝脏脂代谢相关基因的表达研究

目的探讨载脂蛋白E和低密度脂蛋白受体双基因缺失（apoE^-/-/LDLR^-/-）小鼠肝脏脂代谢相关基因表达特征与动脉粥样硬化早期病变的关系及发生机制。方法应用RT-PCR技术检测apoE^-/-／LDLR^-/-与野生型小鼠肝脏脂代谢相关基因表达差异,并进行血生化指标检测及主动脉形态学观察。结果所检测的11个脂代谢相关基因中,apoE^-/-／LDLR^-/-小鼠与野生型小鼠相比,载脂蛋白B100和脂肪酸转运体（FAT／CD36）的mRNA水平从14天龄至3月龄均升高。载脂蛋白AⅣ、载脂蛋白AⅤmRNA水平分别于14天龄和3月龄时明显降低。载脂蛋白AⅠ、载脂蛋白F、过氧化物增殖物激活型受体α、肝X受体α、血管生成素样蛋白3、酰基辅酶A氧化酶1及肉碱棕榈酰转移酶ImRNA表达水平无明显差异。血清总胆固醇、总甘油三酯和低密度脂蛋白胆固醇水平分别高于同龄野生型小鼠约7、2和30倍。随年龄增长apoE^-/-／LDLR^-/-小鼠主动脉内膜出现典型的动脉粥样硬化早期病变。结论上述脂代谢相关基因表达变化表明它们在动脉粥样硬化发生发展过程中起重要作用。     

Spontaneous plaque rupture and secondary thrombosis in apolipoprotein E-deficient and LDL receptor-deficient mice

Apolipoprotein E-deficient (apoE(-/-)) and LDL receptor-deficient (LDLR(-/-)) mice develop extensive atherosclerosis, but the occurrence of spontaneous plaque rupture and secondary thrombosis in these models has not been established. The goal of this study was to provide histological evidence of acute complications of atherosclerotic lesions in these mice and to assess their prevalence. Complications of atherosclerosis were initially studied in aortas of control mice which died during previous intervention studies. Coronary arteries and the aortic origin were then systematically assessed in serial sections through the heart of apoE(-/-) and LDLR(-/-) mice. Aortic plaque rupture and/or thrombi were seen in 3 of 82 untreated mice from past intervention studies. Screening of heart sections of 33 older apoE(-/-) mice (age 9-20 months) showed extensive atherosclerosis in one or more coronary arteries of 18 animals. In three coronary arteries, the presence of blood-filled channels within advanced atherosclerotic lesions suggested previous plaque disruption/thrombotic events followed by recanalization. In the aortic origin of the same mice, four deep plaque ruptures (or erosions reaching necrotic core areas) and a large thrombus originating from the core of a disrupted atherosclerotic lesion were observed. Although plaque ruptures/deep erosions were far less frequent than in human populations, these observations demonstrate that spontaneous plaque rupture and secondary thrombosis do occur in apoE(-/-) and LDLR(-/-) mice. These mice may therefore be suitable for studying factors contributing to thrombotic complications of atherosclerosis. However, the frequent absence of a clearly defined single fibrous cap in murine coronary lesions limits their usefulness as a model of fibrous cap rupture.     

Dietary antioxidants decrease serum soluble adhesion molecule (sVCAM-1, sICAM-1) but not chemokine (JE/MCP-1, KC) concentrations, and reduce atherosclerosis in C57BL but not apoE*3 Leiden mice fed an atherogenic diet

Dietary antioxidants are reported to suppress cellular expression of chemokines and adhesion molecules that recruit monocytes to the artery wall during atherosclerosis. In the present study we measured the effect of feeding apoE*3 Leiden mice or their non-transgenic (C57BL) littermates with atherogenic diets either deficient in, or supplemented with, dietary antioxidants (vitamin E, vitamin C and beta-carotene) for 12 weeks, on serum levels of CC (JE/MCP-1) and CXC (KC) chemokines and soluble adhesion molecules (sVCAM-1, sICAM-1) and atherosclerotic lesion size. ApoE*3 Leiden mice developed gross hypercholesterolaemia, and markedly accelerated (10-20 fold; P < 0.0001) atherogenesis, compared with non-transgenic animals. Antioxidant consumption reduced lesion area in non-transgenic, but not apoE*3 Leiden, mice. Serum sVCAM-1 and sICAM-1 levels were significantly (P<0.0001) increased (sVCAM-1 up to 3.9 fold; sICAM-1 up to 2.4 fold) by 4-8 weeks in all groups, and then declined. The initial increase in the concentration of adhesion molecules was reduced by 38%-61% (P < 0.05) by antioxidant consumption, particularly in non-transgenic mice. By contrast, serum chemokine levels tended to increase more rapidly from baseline in apoE*3 Leiden mice, compared with non-transgenic animals, but were unaffected by dietary antioxidants. We conclude that dietary antioxidants reduce circulating soluble adhesion molecules and atherosclerosis in C57BL mice.     

Circadian rhythm disorder of thrombosis and thrombolysis-related gene expression in apolipoprotein E knock-out mice

Plaque rupture and subsequent embolism as well as thrombosis are major causes of acute myocardial infarction and stroke secondary to atherosclerosis. Pai-1, t-PA, TF and ET-1 are thrombosis- and thrombolysis-related factors which play important roles in thrombosis formation and plaque rupture. Since acute myocardial infarction and stroke are more likely to occur between 6 a.m. and 12 p.m. than at another time of the day, we studied the relationship between circadian rhythm and Pai-1, t-PA, TF and ET-1 in normal and atherosclerotic mice. Atherosclerosis was developed in apoE-/- mice fed a normal diet or a high cholesterol diet. The expression of Pai-1, t-PA, TF and ET-1 in the hearts of control C57BL/6J mice and atherosclerotic mice was measured by real-time RT-PCR at different Zeitgeber times (ZT) including ZT0, ZT4, ZT8, ZT10, ZT12, ZT14, ZT16 and ZT20. The expression of Pai-1, t-PA, TF and ET-1 peaked between ZT14 and ZT16 and bottomed at ZT10 in C57BL/6J mice. Their expression in apoE-/- mice fed a normal diet lost circadian rhythm. Their expression in apoE-/- mice fed a high cholesterol diet peaked at ZT4, indicating a reverse circadian rhythm. Our result indicates that circadian changes in the expression of Pai-1, t-PA, TF and ET-1 may be involved in the onset of myocardial infarction and stroke.     

Identification of a 76-year-old patient with compound heterozygous Familial Hypercholesterolemia by haplotype analysis of the LDL receptor gene

Summary In a large family with clinical characteristics of heterozygous familial hypercholesterolemia (FH) seven restriction fragment length polymorphisms (RFLP) were used to determine lowdensity-lipoprotein receptor (LDLR) gene haplotypes. Following the inheritance of the LDL receptor genes characterized by their seven RFLP haplotypes, two different alleles were found to cosegregate with elevated cholesterol levels within this family. In a 76-year-old man both alleles identified as defective were present, thus classifying this individual as heterozygous compound for FH. In five heterozygous family members one allele was associated with 38% higher cholesterol levels when compared to the other mutant allele in two heterozygous family members. Cosegregation of hypercholesterolemia with the apolipoprotein B (apoB) gene and apolipoprotein E (apoE) gene was excluded by genotyping all individuals for the apoB XbaI RFLP and apoE polymorphisms. These findings are consistent with variable phenotypic expression of the mutant LDLR gene alleles.Abbreviations apo B apolipoprotein B - apo E apolipoprotein E - CAD coronary artery disease - FH familial hypercholesterolemia - LDLR low-density-lipoprotein receptor - MI myocardial infarction - RFLP restriction fragment length polymorphisms     

Experimental Yersinia enterocolitica infection in euthymic and T-cell-deficient athymic nude C57BL/6 mice: comparison of time course, histomorphology, and immune response.

To elucidate the role of T lymphocytes in primary infection with Yersinia enterocolitica, we investigated the elimination rate of this pathogen, the histomorphology of tissue lesions, and the immune responses of athymic T-cell-deficient C57BL/6 nude mice and their euthymic littermates after parenteral infection with Y. enterocolitica of serotype O:8. While a low inoculum of 3 x 10(2) Y. enterocolitica cells (about 0.01 times the median lethal dose for normal C57BL/6 mice) was cleared by normal C57BL/6 mice within 7 to 10 days, athymic nude C57BL/6 mice developed progressive infections after this inoculum, leading to death on days 20 to 25 postinfection (p.i.). While normal C57BL/6 mice experienced short-term transient infections, nude mice exhibited a biphasic, progressive infectious process. Thus, in the early phase (days 1 to 7 p.i.), a rapid influx of CD11b/18-positive cells (Mac-1 antigen) and natural killer cells was evident in the spleens and livers of the nude mice. The late phase (from day 8 p.i. onward) was characterized by a rapid progression of the infection and a further influx of CD11b/18-positive cells into the liver accompanied by an increase in bacterial counts and development of tissue lesions particularly in the liver and spleen. In normal mice, granuloma-like lesions composed of CD11b/18-, CD4-, and CD8-positive cells could be observed. However, granulomata were not found in nude mice. Yersinia-specific immunoglobulin G antibodies appeared on day 15 p.i. in the sera of normal mice, while nude mice failed to develop significant antibody titers. Adoptive transfer of Yersinia-specific T cells into athymic nude mice mediated resistance to Y. enterocolitica infection and restored both the ability of granuloma formation and the production of specific antibodies. In summary, the data presented herein strongly suggest that T lymphocytes play an essential role in the defense of C57BL/6 mice against Y. enterocolitica.     

Induction of macrophage scavenger receptor MARCO in nonalcoholic steatohepatitis indicates possible involvement of endotoxin in its pathogenic process

Nonalcoholic steatohepatitis (NASH) is one of the life-threatening hepatic diseases; however, its pathogenesis is still unknown. To evaluate the causative role of hyperlipidaemia and high-fat diet, we compared C57BL/6 mice with inherited hyperlipidaemic model mice (LDLR(-/-)mice and ApoE(-/-) mice) fed a normal or a high-fat diet. LDLR(-/-) and ApoE(-/-) mice fed the normal diet showed significantly higher serum cholesterol level than that of C57BL/6 mice fed the high-fat diet. These mice, however, have shown neither significant elevation of serum alanine transaminase (ALT) level nor histopathologic features of steatohepatitis. High-fat diet groups of all three strains showed histopathological characteristics of steatohepatitis with elevated serum ALT levels and high expression of macrophage scavenger receptor MARCO mRNA in the liver. Semiquantitative endotoxin analysis showed an elevated serum endotoxin level in the portal vein but not in the vena cava in ApoE(-/-) mice fed the high-fat diet. These results indicate that long-term feeding of a high-fat diet induces NASH, whereas hyperlipidaemia alone is not enough to induce NASH. Liver-restricted induction of MARCO in mice with high-fat diet and portal endotoxaemia in ApoE(-/-) mice fed the high-fat diet suggest the possible involvement of endotoxin in the pathogenesis of NASH.     

LPS binding protein is important in the airway response to inhaled endotoxin

BACKGROUND: Inhaled endotoxin is a risk factor for asthma exacerbation, and endotoxin inhalation by itself recapitulates many of the classical features of asthma in mice, including reversible airflow obstruction and inflammation, airways hyperresponsiveness, and airway remodeling. OBJECTIVE: Our objective was to determine the importance of LPS binding protein (LBP) in the response to inhaled LPS. METHODS: We challenged LBP-deficient mice (C57BL/6(LBP-/-)) and C57BL/6 mice with inhaled endotoxin for 4 hours, 5 days, or 4 weeks, followed by 3 days of recovery. RESULTS: LBP in the lung was significantly increased in LPS-exposed C57BL/6 mice from all 3 groups. Only LPS-exposed C57BL/6 mice had significantly enhanced airway responsiveness to inhaled methacholine. Total lavage cells in LPS-exposed C57BL/6(LBP-/-) mice were significantly reduced compared with those seen in LPS-exposed C57BL/6 mice; however, the percentage of PMNs was similarly increased in both the C57BL/6 and C57BL/6(LBP-/-) mice. TNF-alpha, IL-1 beta, and IL-6 protein concentrations in whole-lung lavage fluid from C57BL/6(LBP-/-) mice were also significantly reduced when compared with those seen in C57BL/6 mice. In C57BL/6(LBP-/-) mice submucosal cell proliferation was significantly reduced in the 1-week group when compared with that seen in similarly exposed C57BL/6 mice. The 4-week exposed C57BL/6 mice had significantly thickened airway submucosa and significantly increased lavaged TGF-beta(1) protein compared with that seen in C57BL/6(LBP-/-) mice. CONCLUSIONS: These findings indicate that LBP is one of the critical molecules regulating the acute and chronic airway response to inhaled LPS.     

Mast cell-derived TNF contributes to airway hyperreactivity, inflammation, and TH2 cytokine production in an asthma model in mice

BACKGROUND: Mast cells, IgE, and TNF, which have been implicated in human atopic asthma, contribute significantly to the allergic airway inflammation induced by ovalbumin (OVA) challenge in mice sensitized with OVA without alum. However, it is not clear to what extent mast cells represent a significant source of TNF in this mouse model. OBJECTIVE: We investigated the importance of mast cell-derived TNF in a mast cell-dependent model of OVA-induced airway hyperreactivity (AHR) and allergic airway inflammation. METHODS: Features of this model of airway inflammation were analyzed in C57BL/6J-wild-type mice, mast cell-deficient C57BL/6J-Kit(W-sh)(/W-sh) mice, and C57BL/6J Kit(W-sh/W-sh) mice that had been systemically engrafted with bone marrow-derived cultured mast cells from C57BL/6J-wild-type or C57BL/6J-TNF(-/-) mice. RESULTS: Ovalbumin-induced AHR and airway inflammation were significantly reduced in mast cell-deficient Kit(W-sh/W-sh) mice versus wild-type mice. By contrast, Kit(W-sh/W-sh) mice that had been engrafted with wild-type but not with TNF(-/-) bone marrow-derived cultured mast cells exhibited responses very similar to those observed in wild-type mice. Mast cells and mast cell-derived TNF were not required for induction of OVA-specific memory T cells in the sensitization phase, but significantly enhanced lymphocyte recruitment and T(H)2 cytokine production in the challenge phase. CONCLUSION: Mast cell-derived TNF contributes significantly to the pathogenesis of mast cell-dependent and IgE-dependent, OVA-induced allergic inflammation and AHR in mice, perhaps in part by enhancing lymphocyte recruitment and T(H)2 cytokine production. CLINICAL IMPLICATIONS: Our findings in mice support the hypothesis that mast cell-derived TNF can promote allergic inflammation and AHR in asthma.     

Induction of persistent autoimmune haemolytic anaemia in mice by combined use of rat erythrocyte preimmunization and chronic GVHR

Female F1 mice (BDF1: C57BL/6 x DBA/2, BCF1: C57BL/6 x C3H/HeN, BBF1: C57BL/6 x BALB/c) were preimmunized with rat erythrocytes and then received splenic T cells from parental female C57BL/6 mice injected with hydrocortisone acetate before killing. In BDF1 and BCF1 mice, direct Coombs' test (DCT) positivity persisted for more than 4 months in almost all mice. They also showed haematological sign of anaemia, and the survival of infused 51Cr-labelled parental C57BL/6 erythrocytes was reduced. In BBF1 mice, DCT positivity returned to negative within 2 months. Autoantibodies were eluted from DCT-positive erythrocytes and their specificities were examined. They were capable of binding equally well to erythrocytes from all strains of mice tested, including the donor mice. They also cross-reacted with rat erythrocytes but not with sheep, rabbit or human erythrocytes. Isotypes of the autoantibodies eluted were also examined. Interestingly, isotypes of the autoantibodies obtained from individual mice were confined to one or other subclass of IgG isotype: IgG1, IgG2a or IgG2b.     

Genetic control of natural resistance in mouse macrophages regulating intracellular Legionella pneumophila multiplication in vitro.

It is known that Legionella pneumophila proliferates in peritoneal macrophage cultures derived from A/J mice but not in macrophage cultures derived from many other strains, including C57BL/6 mice. To analyze the genetic control of this trait and the location of the Legionella resistance-susceptibility gene, we prepared segregating progeny of A/J and C57BL/6 mice and determined the levels of susceptibility of individual mice. Peritoneal macrophages were collected by injecting thioglycolate medium, and macrophage monolayers were infected in vitro with L. pneumophila Philadelphia-1. Counting of colonies on buffered charcoal yeast extract agar plates and Gimenez staining of macrophage monolayers were carried out daily. There was a 10-fold increase in bacterial burden 1 day after infection and a 100-fold increase after 2 days in A/J (susceptible) macrophages. The increase in bacterial burden was always less than 10-fold in macrophages from C57BL/6 (resistant) progenitors, A/J x C57BL/6 F1 hybrids, and C57BL/6 x F1 backcross progeny. The ratios of resistant individuals to susceptible individuals were 22:6 for F2 progeny and 20:22 for A/J x F1 backcross progeny. The fact that the organism did not proliferate in macrophages from B10.A mice demonstrated that major histocompatibility antigens did not regulate the macrophage resistance of C57BL/6-derived mice. The sex and coat color genes of mice were not linked to the resistance-susceptibility gene. We suggest that resistance and susceptibility are controlled by a single gene or closely linked genes which are autosomal and that the resistance allele is dominant. The results of a comparison of the strain distribution pattern of this trait with the distribution pattern of 185 allelic markers in A/J x C57BL/6 and C57BL/6 x A/J recombinant inbred strains suggest that this susceptibility-resistance gene is located in the proximal part of chromosome 15.     

Identification of genetic loci controlling bacterial clearance in experimental Salmonella enteritidis infection: an unexpected role of Nramp1 (Slc11a1) in the persistence of infection in mice

The Gram-negative bacteria, Salmonella, cause a broad spectrum of clinical diseases in both animals and humans ranging from asymptomatic carriage to life-threatening sepsis. We have developed a model to study the contribution of genetic factors to the susceptibility of 129sv and C57BL/6J inbred mice to Salmonella enteritidis during the late phase of infection. C57BL/6J mice were able to eliminate completely sublethal inoculums of S. enteritidis from their reticuloendothelial system, whereas 129sv mice could not even after 60 days post inoculation. A genome scan performed on 302 (C57BL/6J x 129sv) F2 progeny identified three dominant loci (designated Ses1 to Ses3) that are associated with disease susceptibility in 129sv mice. Two highly significant linkages were identified on chromosomes 1 (Ses1) and 7 (Ses2) with respective LOD scores of 9.9 (P = 1.4 x 10(-11)) at D1Mcg5 and 4.0 (P = 1.9 x 10(-5)) at D7Mit62. One highly suggestive QTL was located on chromosomes15 (Ses3) with a LOD score 3.4 (P = 1.2 x 10(-4)). The estimated effects of Ses1, Ses2 and Ses3 on the bacterial clearance were greater in females. Using a model of three loci, with interaction between Ses1 and Ses2 and sex as a covariate, the three QTLs explained 32% of the phenotypic variance. The candidacy of Nramp1 as the gene for Ses1 was evaluated using mice carrying a null allele at Nramp1 (129sv-Nramp1(tm1Mcg)). These mice have a significantly lower spleen bacterial load compared to the wild-type 129sv mice, strongly suggesting the involvement of Nramp1 in controlling S. enteritidis clearance during the late phase of infection.     

Development of atherosclerosis in Balb/c apolipoprotein E-deficient mice

BACKGROUND: Since its creation in 1992 by gene inactivation via gene targeting, the apolipoprotein E "knockout" mouse has become the most widely used rodent model for the study of atherosclerosis. Commercially available apolipoprotein E(-/-) mice are bred on a C57BL/6J background. The goal of the present study was to investigate the development of atherosclerosis in apolipoprotein E-deficient mice generated on a Balb/c background. METHODS: We compared serum cholesterol concentrations and the development of atherosclerotic lesions in heterozygous Balb/c [apolipoprotein E(+/-)] mice fed regular rodent chow, Balb/c apolipoprotein E-deficient mice fed regular chow, and Balb/c apolipoprotein E-deficient mice fed a high-fat diet for up to 30 weeks. Expression of the chemokine JE (murine homologue of MCP-1), as well as the adhesion molecules E-selectin, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1, in the aortas of knockout mice fed a high-fat diet was measured by enzyme-linked immunosorbent assay. RESULTS: Balb/c apolipoprotein E-deficient mice develop atherosclerotic lesions in a reproducible temporal and morphological pattern. Total serum cholesterol concentrations in Balb/c apolipoprotein E-deficient mice fed regular chow or a high-fat diet, respectively, closely parallel those reported for C57BL/6J apolipoprotein E-deficient mice. The expression of all three adhesion molecules in the aorta follows a similar temporal pattern, peaking in the first 15 weeks, whereas JE concentrations peak around 23 weeks. CONCLUSION: The availability of Balb/c apolipoprotein E-deficient mice will facilitate the study of atherosclerosis in a mouse strain that can concomitantly develop other pathological states that are not readily inducible in mice with the C57BL/6J background.     

Progressive loss of synaptic integrity in human apolipoprotein E4 targeted replacement mice and attenuation by apolipoprotein E2

Inheritance of the APOE4 allele is a well established genetic risk factor linked to the development of late onset Alzheimer's disease. As the major lipid transport protein in the central nervous system, apolipoprotein (apo) E plays an important role in the assembly and maintenance of synaptic connections. Our previous work showed that 7 month old human apoE4 targeted replacement (TR) mice displayed significant synaptic deficits in the principal neurons of the lateral amygdala, a region that is critical for memory formation and also one of the primary regions affected in Alzheimer's disease, compared to apoE3 TR mice. In the current study, we determined how age and varying APOE genotype affect synaptic integrity of amygdala neurons by comparing electrophysiological and morphometric properties in C57BL6, apoE knockout, and human apoE3, E4 and E2/4 TR mice at 1 month and 7 months. The apoE4 TR mice exhibited the lowest level of excitatory synaptic activity and dendritic arbor compared to other cohorts at both ages, and became progressively worse by 7 months. In contrast, the apoE3 TR mice exhibited the highest synaptic activity and dendritic arbor of all cohorts at both ages. C57BL6 mice displayed virtually identical synaptic activity to apoE3 TR mice at 1 month; however this activity decreased by 7 months. ApoE knockout mice exhibited a similar synaptic activity profile with apoE4 TR mice at 7 months. Consistent with previous reports that APOE2 confers protection, the apoE4-dependent deficits in excitatory activity were significantly attenuated in apoE2/4 TR mice at both ages. These findings demonstrate that expression of human apoE4 contributes to functional deficits in the amygdala very early in development and may be responsible for altering neuronal circuitry that eventually leads to cognitive and affective disorders later in life.     

Apolipoprotein E inhibits neointimal hyperplasia after arterial injury in mice

The potential cytostatic function of apolipoprotein (apo) E in vivo was explored by measuring neointimal hyperplasia in response to vascular injury in apoE-deficient and apoE-overexpressing transgenic mice. Results showed a significant increase in medial thickness, medial area, and neointimal formation after vascular injury in both apoE knockout and wild-type C57BL/6 mice. Immunochemical analysis with smooth muscle alpha-actin-specific antibodies revealed that the neointima contained proliferating smooth muscle cells. Neointimal area was 3.4-fold greater, and the intima/medial ratio as well as stenotic luminal area was more pronounced in apoE(-/-) mice than those observed in control mice (P < 0.05). The human apoE3 transgenic mice in FVB/N genetic background were then used to verify a direct effect of apoE in protection against neointimal hyperplasia in response to mechanically induced vascular injury. Results showed that neointimal area was reduced threefold to fourfold in mice overexpressing the human apoE3 transgene (P < 0.05). Importantly, suppression of neointimal formation in the apoE transgenic mice also abolished the luminal stenosis observed in their nontransgenic FVB/N counterparts. These results documented a direct role of apoE in modulating vascular response to injury, suggesting that increasing apoE level may be beneficial in protection against restenosis after vascular surgery.     

CCL21/CCR7对小鼠急性心肌梗死后炎症反应及梗死面积的影响

目的:探讨抗CCL21处理对急性心肌梗死(acute myocardial infarction,AMI)后小鼠炎症反应和心肌损伤的影响.方法:结扎冠脉前降支构建C57BL/6小鼠心肌梗死模型,随机分为假手术组(Sham)、对照组(isotype-IgG control)和CCL21单抗干预组(goat anti-mouse CCL21 mAb).qPCR检测AMI后心肌组织CCL21受体CCR7表达,ELISA法检测抗CCL21处理对心肌梗死后小鼠血清炎症趋化因子和肌钙蛋白的影响.HE染色评估心肌梗死后心肌组织炎性细胞浸润.1和7 d后Evans blue/TTC双染评估梗死区和危险区面积.结果:AMI导致小鼠血清CCL21水平和心肌组织CCR7在心肌组织内表达水平显著增高.冠脉结扎后小鼠血清白细胞趋化因子水平显著增高,anti-CCL21干预组小鼠血清CXCL1等趋化因子水平均显著低于对照isotype-IgG组.给予anti-CCL21处理有效逆转了cTnI和心肌酶LDH-1升高,并显著降低心肌梗死区面积.结论:抗CCL21处理可显著抑制心肌梗死后炎症反应并降低小鼠心肌梗死面积.     

Differential expression of cysteine and aspartic proteases during progression of atherosclerosis in apolipoprotein E-deficient mice

Several groups of proteolytic enzymes are able to degrade components of the extracellular matrix. During atherosclerosis, matrix remodeling is believed to influence the migration and proliferation of cells within the plaque. In the present study, gene expression of several proteases and their inhibitors was analyzed during the development of atherosclerosis in apolipoprotein E-deficient (ApoE-/-) mice. Quantitative real-time polymerase chain reaction was used to study gene expression of proteases after 10 and 20 weeks in ApoE-/- and C57BL/6 mice and in atherosclerotic lesions and nonaffected regions of the same ApoE-/- mouse. Some of the differentially expressed proteolytic enzymes were studied by immunohistochemistry. The matrix metalloproteinase (MMP)-9 and its inhibitor TIMP-1 were differentially expressed and the expression increased with time. Urokinase-type plasminogen activator showed no major expression. In contrast, cathepsins B, D, L, and S all showed strong and increased expression in ApoE-/- mice compared to C57BL/6 mice whereas the expression of their inhibitor, cystatin C, did not differ between the two mouse strains. The expression of cathepsins was mainly localized to the lesions and not to nonaffected regions of the aorta of ApoE-/- mice. Furthermore, cathepsin expression was similar to the expression of the macrophage marker macrosialin (CD68) although expression of cathepsins B, D, and L could be demonstrated in healthy C57BL/6 mice and in nonaffected vessel segments of atherosclerotic ApoE-/- mice. Cathepsin S mRNA expression was restricted to lesions of ApoE-/- mice. Furthermore, cathepsin S was the only cathepsin that was expressed in the media and absent in lipid-rich regions. All cathepsins studied showed intimal expression, the degree and localization of which differed between individual cathepsins. In conclusion, increased expression of several cathepsins in atherosclerotic lesions suggests that these proteases may participate in the remodeling of extracellular matrix associated with the atherosclerotic process.