Perspectives in familial hypercholesterolemia: with particular reference to associated ischemic heart disease

Seventy heterozygous patients with familial hypercholesterolemia (FH), aged 3 to 91 years (37 men and 33 women), from 38 different families were studied to ascertain their clinical profiles including the pattern of distribution of total cholesterol (TC), Achilles tendon thickness (ATT), and a sex difference in age at the onset of ischemic heart disease (IHD). Eighteen family members who died of IHD were included in this study. The TC level was 332.2 +/- 95.0 mg/dl (mean +/- SD), and the ATT was 12.0 +/- 2.7 mm. IHD was observed in 20 men and 8 women, with an incidence in men 2.5 times higher than that in women. The mean age at the onset of IHD in men was in the sixth decade, one decade younger than in women. IHD was even observed in patients with TC levels of nearly 230 mg/dl. One of the 18 family members was considered a homozygote of the disease by autopsy findings. Twelve of the remaining members died suddenly, and another five had fatal myocardial infarction. We conclude that, (1) IHD is frequently associated with FH, even though TC levels are 230-300 mg/dl, (2) complications of IHD peak earlier in men (sixth decade) than in women (seventh decade) and (3) relatively large numbers of family members died suddenly, especially men.     

Effects of hypercholesterolemia on myocardial ischemia-reperfusion injury in LDL receptor-deficient mice

Hypercholesterolemia is a primary risk factor for atherosclerosis, coronary artery disease, and myocardial infarction. We subjected low density lipoprotein receptor-deficient (LDLr -/-) and control (wild-type) mice to 30 minutes of myocardial ischemia and 120 minutes of reperfusion. Myocardial infarction per area at risk (AAR) was noted under baseline conditions to be significantly (P<0.05) smaller in the LDLr -/- mice compared with wild-type mice (24.7+/-3. 2% and 38.8+/-4.3% of AAR, respectively). Subsequently, mice were fed a high-cholesterol diet (HCD) for 2 or 12 weeks, which resulted in significant increases in serum cholesterol levels in both LDLr -/- and wild-type groups. After 2 weeks of the HCD, the LDLr -/- mice demonstrated a significant elevation (P<0.01) in myocardial necrosis per AAR (50.2+/-5.36% of AAR) compared with the normal-diet LDLr -/- group, whereas the short-term HCD-fed wild-type mice demonstrated no significant difference from baseline. In contrast, wild-type mice fed the HCD for 12 weeks revealed a significant (P<0. 05) decrease in necrosis per AAR, which was 22.5+/-3.2% of the AAR in comparison with that in the normal-diet wild-type mice (38.8+/-4. 3% of AAR). LDLr -/- mice on the same long-term HCD showed a similar significantly (P<0.05) decreased infarct size, which was 13.2+/-4.0% of the AAR. In additional experiments, we determined that myocardial tissue total glutathione (GSH) levels were reduced after 2 weeks of the HCD and were significantly increased after 12 weeks of the HCD in the LDLr -/- mouse heart. These data suggest that short-term cholesterol feeding renders the myocardium of LDLr -/- mice more susceptible to ischemia-reperfusion injury, whereas more long-term hypercholesterolemia confers cardioprotection in the LDLr -/- mouse heart.     

Effects of short-term, diet-induced hypercholesterolemia on systemic hemodynamics, myocardial blood flow, and infarct size in awake dogs with acute myocardial infarction

BACKGROUND. Short-term cholesterol feeding has been shown to affect vasomotor tone and increase infarct size in anesthetized rabbits. The purpose of the study was to determine whether acute hypercholesterolemia reduced collateral flow to ischemic myocardium and increased infarct size in the awake dog. METHODS AND RESULTS. Acute myocardial infarction was produced in awake dogs by a 4-hour left anterior descending coronary artery occlusion followed by 6-hour reperfusion after either a cholesterol-supplemented diet (n = 14) or a control diet of dog chow (n = 15) for 10 days. Infarct size was determined using nitroblue tetrazolium staining. In two subgroups, a 15-minute transient occlusion of the left anterior descending coronary artery was produced before the diet treatments and was compared with occlusion after diet treatments, so that the effects of hypercholesterolemia of collateral flow could be determined by paired comparisons. Cholesterol feeding increased plasma cholesterol to 288 +/- 52 mg/dl, which was twofold to threefold that in the control group (127 +/- 35 mg/dl), but had no effects on baseline systemic hemodynamics and myocardial blood flow. Coronary artery occlusion produced similar increases in heart rate, mean aortic pressure, left atrial pressure, and plasma norepinephrine in both groups of animals. However, cholesterol feeding reduced collateral flow to ischemic myocardium and increased infarct size, compared with the control group. The infarct size correlated with ischemic myocardial blood flow in both groups, but the slopes of regression lines relating the two variables did not differ between the two groups. CONCLUSIONS. Short-term, diet-induced hypercholesterolemia increased infarct size in awake dogs. This change results, at least in part, from a decrease in collateral blood flow to ischemic myocardium during coronary artery occlusion.     

Increased apoptosis in the heart of genetic hypertension, associated with increased fibroblasts

OBJECTIVE: The present studies were undertaken to identify apoptosis in cardiomyocytes of genetic hypertension and to study the relationship among apoptosis, aging and blood pressure, and the effect of angiotensin-converting enzyme (ACE) inhibitors on apoptosis. METHODS: Apoptosis in the hearts of spontaneously hypertensive rats (SHR) was identified by electron microscopy (EM) and DNA laddering, and quantified from age 3 weeks to 64 weeks in comparison with normotensive rats (WKY). Fibroblasts and protein products of Bcl-2 and Bax were measured by quantitative immunohistochemistry. SHR were treated with ramipril, an ACE inhibitor. RESULTS: The results showed that: (1) ultrastructural characteristics of apoptosis were observed in cardiomyocytes of SHR, with shrinkage of the cell and condensation of the cytoplasm and chromatin. A DNA ladder was shown; (2) a significant increase in apoptosis in SHR began as early as age 4 weeks and reached a plateau at 16 weeks and maintained at high levels up to 64 weeks. Blood pressure (BP) in SHR started to increase significantly at age 5 weeks; (3) fibroblasts were significantly increased in the heart of SHR; (4) the ratio of Bcl-2/Bax was significantly reduced in SHR; and (6) ramipril effectively reduced apoptosis and fibroblasts, and increased the ratio of Bcl-2/Bax. CONCLUSION: Apoptosis occurs in the cardiomyocytes of genetic hypertension although fibroblasts are increased, and a significant, age-dependent increase in apoptosis is observed. The increase in apoptosis occurs before the difference in blood pressure is detectable. The ACE inhibitor ramipril may be useful for prevention of apoptosis in the heart.     

Manipulations of core temperatures in ischemia-reperfusion lung injury in rabbits

The present study was designed to determine the effect of various core temperatures on acute lung injury induced by ischemia-reperfusion (I/R) in our isolated rabbit lung model. Typical acute lung injury was successfully induced by 30 min of ischemia followed by 90 min of reperfusion observation. The I/R elicited a significant increase in pulmonary arterial pressure, microvascular permeability (measured by using the capillary filtration coefficient, Kfc), Delta Kfc ratio, lung weight gain and the protein concentration of the bronchoalveolar lavage fluid. Mild hypothermia significantly attenuated acute lung injury induced by I/R, all parameters having decreased significantly (p<0.05); conversely, mild hyperthermia did not further exacerbate acute lung injury. These experimental data suggest that mild hypothermia significantly ameliorated acute lung injury induced by ischemia-reperfusion in rabbits.     

Increased bone resorption and impaired bone microarchitecture in short-term and extended high-fat diet-induced obesity

Although obesity traditionally has been considered a condition of low risk for osteoporosis, this classic view has recently been questioned. The aim of this study was to assess bone microarchitecture and turnover in a mouse model of high-fat diet-induced obesity. Seven-week-old male C57BL/6J mice (n = 18) were randomized into 3 diet groups. One third (n = 6) received a low-fat diet for 24 weeks, one third was kept on an extended high-fat diet (eHF), and the remaining was switched from low-fat to high-fat chow 3 weeks before sacrifice (sHF). Serum levels of insulin, leptin, adiponectin, osteocalcin, and cross-linked telopeptides of type I collagen (CTX) were measured. In addition, bone microarchitecture was analyzed by micro-computed tomography; and lumbar spine bone density was assessed by dual-energy x-ray absorptiometry. The CTX, body weight, insulin, and leptin were significantly elevated in obese animals (sHF: +48%, +24%, +265%, and +102%; eHF: +43%, +52%, +761%, and +292%). The CTX, body weight, insulin, and leptin showed a negative correlation with bone density and bone volume. Interestingly, short-term high-fat chow caused similar bone loss as extended high-fat feeding. Bone volume was decreased by 12% in sHF and 19% in eHF. Bone mineral density was 25% (sHF) and 27% (eHF) lower when compared with control mice on low-fat diet. As assessed by the structure model index, bone microarchitecture changed from plate- to rod-like appearance upon high-fat challenge. Trabecular and cortical thickness remained unaffected. Short-term and extended high-fat diet-induced obesity caused significant bone loss in male C57BL/6J mice mainly because of resorptive changes in trabecular architecture.     

Age-associated ultrastructural changes in the aortic intima of rats with diet-induced hypercholesterolemia

The ultrastructure of the aortic intima and serum lipid levels in Fischer 344 rats were examined at the ages of 12, 18 and 24 months after feeding the animals an atherogenic diet (2% cholesterol, 0.25% sodium cholate, 5% beef fat) for 6 months. Structural changes in the intima were noticeable only at the age of 24 months. In control rats, the endothelial cells were irregular in shape and each had a well-developed Golgi complex and a few lipid droplets. Simultaneously, reticular, basal lamina-like material and electron-dense granules of extracellular liposomes accumulated in the subendothelium. In fat-fed rats, these structural changes were more conspicuous in association with hypercholesterolemia, and numerous monocytes with lipid droplets were attached to the endothelium, occasionally invading into the subendothelium. Slight foam cell lesions were evident in the intima. The finding that older rats were more susceptible to the atherogenic diet suggests that atherogenesis in the rat is promoted by intrinsic age-associated changes in the aortic intima.     

Y-27632对家兔心肌缺血／再灌注损伤的有益作用

目的研究Rho激酶抑制剂Y-27632对家兔心肌缺血／再灌注损伤（IRI）是否有心脏保护作用。方法健康家兔18只，体重2．0～2．5kg，随机分为3组：IRI组、Y-27632（Y）组和假手术（C）组。用结扎左冠状动脉前降支60min后剪断结扎线恢复血流30min的方法建立在体心肌IRI模型，Y组于再灌注前5min，经耳缘静脉注射Y-276320．35mg/kg。检测左心室收缩功能，血浆肌酸激酶（CK）、乳酸脱氢酶（LDH）及心肌缺血、梗死范围。结果IRI组缺血／再灌注导致左心室舒缩功能受损，左心室收缩峰压（LVSP）、左心室内压最大上升速率（＋dp／dtmax）下降，左心室舒张末期压力（LVEDP）升高，CK、LDH的释放增加，心肌梗死范围为（5．3±0．7）％；Y组在再灌注早期LVSP、±dp／dtmax下降的幅度和LVEDP升高的幅度、CK和LDH的释放，均明显低于IRI组，组间比较P均〈0．05。心肌缺血、梗死范围也明显较IRI组小（缺血：40．5％ vs 51．3％，P〈0．05；梗死：1．7％ vs 5．3％，P〈0．01）。结论Y-27632对家兔心肌IRI有部分心脏保护作用。     

Increased bone turnover in patients with hypercholesterolemia

Osteoporosis has been linked with arteriosclerotic vascular diseases, suggesting that hypercholesterolemia or dyslipidemia may be a common pathogenetic factor underlying these diseases. However, little is known about the relationship between osteoporosis and hypercholesterolemia. The purpose of this study was, therefore, to investigate the effects of hypercholesterolemia upon bone metabolism, by measuring bone turnover markers in hypercholesterolemic patients. This study included 281 Japanese patients with hypercholesterolemia, and 267 control subjects. Serum bone-specific alkaline phosphatase (BAP) of the patients was significantly higher than that of the controls in women. Serum N-terminal telopeptide of type I collagen (NTx) of the patients was significantly higher than that of the controls in both men and women. In addition, both BAP and NTx in men showed a significantly negative correlation with high density lipoprotein cholesterol (HDL-C). On the other hand, in women, both BAP and NTx showed a significantly positive correlation with total cholesterol and low density lipoprotein cholesterol (LDL-C). These results indicate increased bone turnover in hypercholesterolemic or dyslipidemic patients regardless of gender, and suggest the importance of treating hypercholesterolemia or dyslipidemia in order to prevent not only arteriosclerotic complications but also osteoporotic bone loss and subsequent fractures.     

后适应在兔心肌缺血再灌注损伤中的实验研究

目的：观察缺血后适应对兔心肌缺血再灌注损伤（IRI）的保护作用。方法将30只新西兰大白兔随机分为缺血-再灌注组（IR组）,缺血后适应组,后适应加5-羟基癸酸盐（5-HD）组,延迟后适应组及假手术组5组,每组6只。采用硫代巴比妥酸比色法检测5组大白兔心肌组织丙二醛含量,依文思兰加氯化四氮唑兰双重染色法及称重法测定心肌梗死范围大小。结果丙二醛含量缺血后适应组显著低于IR组和延迟后适应组（均P＜0.01）,与后适应加5-HD组相近,但显著高于假手术组（P＜0.05）。心肌梗死面积在IR组、后适应加5-HD组、延迟后适应组之差异无统计学意义（P＞0.05）,但大于缺血后适应组,差异均有显著统计学意义（均P＜0.01）。结论缺血后适应在兔心肌IRI模型中有明显的心肌保护作用;特异性线粒体ATP敏感钾通道抑制剂5-HD可完全取消后适应的心肌保护作用;延迟5min后实行后适应操作其保护作用完全消失。     

Increased susceptibility to diet-induced obesity in histamine-deficient mice

BACKGROUND AND AIM: The neurotransmitter histamine is involved in the regulation of appetite and in the development of age-related obesity in mice. Furthermore, histamine is a mediator of the anorexigenic action of leptin. The aim of the present study was to investigate a possible role of histamine in the development of high-fat diet (HFD)-induced obesity. METHODS: Histamine-deficient histidine decarboxylase knock-out (HDC-KO) mice and C57BL/6J wild-type (WT) mice were given either a standard diet (STD) or HFD for 8 weeks. Body weight, 24-hour caloric intake, epididymal adipose tissue size, plasma leptin concentration and quantitative expression of leptin receptor (Ob-R) mRNA were measured. RESULTS: Both HDC-KO and WT mice fed an HFD for 8 weeks increased their body weight significantly more than STD-fed mice. A significant difference in body weight gain between HDC-KO mice fed an HFD or an STD was seen after 2 weeks, whereas a significant difference in body weight gain was first observed after 5 weeks in WT mice. After 8 weeks 24-hour caloric intake was significantly lower in HFD- than in STD-fed WT mice. In HDC-KO mice no difference in caloric intake was observed between HFD- and STD-fed mice. After 8 weeks epididymal adipose tissue size and plasma leptin concentration had increased significantly in HFD-fed WT and HDC-KO mice compared to their STD-fed controls. Epididymal adipose tissue size was higher in HDC-KO than WT mice, both in STD- and HFD-fed mice. A significant decrease in Ob-R mRNA in HFD-fed HDC-KO mice compared to STD-fed HDC-KO mice was observed, while no such difference was observed in WT mice. CONCLUSION: Based on our results, we conclude that histamine plays a role in the development of HFD-induced obesity.     

衰老大鼠心肌对缺血再灌注损伤的敏感性研究

目的观察衰老大鼠心肌对缺血再灌注损伤的敏感性,并探讨其可能的机制。方法成年和老年雄性Wistar大鼠各12只,分为4组：成年假手术组、成年缺血再灌注组、老年假手术组和老年缺血再灌注组,每组6只,进行缺血30 min再灌注3 h。脱氧核糖核苷酸末端转移酶介导的缺口末端标记(TUNEL)法检测心肌细胞凋亡,比色法检测心肌组织半胱胺酸蛋白酶蛋白-3 (caspase-3)活性,化学发光法测定心肌组织总一氧化氮(NOx)含量,ELISA法检测心肌过氧亚硝基(ONOO~-)含量。结果老年组缺血再灌注引起心肌细胞凋亡的程度明显高于成年组,凋亡指数分别为(14．6±1.7)%、(19．0±2．1)%,差异有统计学意义(P＜0．05)；caspase-3活性：成年组为(340±32)μmol/mg,老年组为(436±35)μmol/mg,差异有统计学意义(P＜0．05)；心肌组织中NOx含量：成年缺血再灌注组、老年缺血再灌注组分别为成年假手术组的(2．1±0．2)、(4．4±0．5)倍,差异有统计学意义(P＜0．05)；ONOO~-含量：成年缺血再灌注组和老年缺血再灌注组分别为(4．68±0．15)nmol/g、(7．25±0．18)nmol/g,差异有统计学意义(P＜0．05)。结论老年大鼠对心肌缺血再灌注损伤的敏感性增加,可能的原因是老年鼠心脏中NO的毒性衍生物ONOO~-含量增加,从而导致功能蛋白的硝基化。     

多胺代谢与大鼠心肌缺血再灌注损伤关系的实验研究

目的 观察缺血再灌注不同时期大鼠心肌多胺代谢变化规律,探讨多胺代谢与心肌缺血再灌注损伤的关系.方法 采用结扎冠状动脉方法复制大鼠在体心肌缺血再灌注损伤模型,应用逆转录聚合酶链反应(RT-PCR)、Western免疫印迹(Western blot)方法分别测定正常、缺血再灌注2 h、6 h、12 h和24 h时心肌鸟氨酸脱羧酶(ODC)和精胺/精脒乙酰转移酶(SSAT)mRNA的转录和蛋白表达水平,并用高效液相色谱仪测定多胺含量变化.结果 心肌缺血再灌注后ODC和SSAT mRNA的转录和蛋白表达均上调,至再灌注24 h时,与假手术组比,ODC mRNA和SSAT mRNA转录分别增加了3.1倍和3.8倍(P＜0.01),ODC和SSAT的蛋白表达分别增加了3.1倍和2.9倍(P＜0.01).精胺、精脒和多胺总代谢池含量减少,至再灌注24 h时,分别比假手术组少了33.6%、35.3%和32.9%,而腐胺多了58.9%(P＜0.01).结论 心肌缺血再灌注损伤可导致多胺代谢失衡,二者密切相关.     

Haplotype of the angiotensinogen gene is associated with coronary heart disease in familial hypercholesterolemia

OBJECTIVE: Familial hypercholesterolemia is characterized by high plasma low-density lipoprotein cholesterol levels and premature coronary heart disease. Despite the monogenetic origin of familial hypercholesterolemia, the incidence of coronary heart disease varies considerably among patients, which is only partly explained by classical risk factors. Hypertension is an important risk factor for coronary heart disease that is associated with angiotensinogen levels. Therefore, we analyzed the angiotensinogen gene as a modifier gene for coronary heart disease risk in patients with familial hypercholesterolemia. METHODS: In a cohort of 1785 familial hypercholesterolemia patients, we reconstructed five frequent haplotypes of the angiotensinogen gene, based on four polymorphisms. The five haplotypes cover approximately 98% of the genetic diversity accounted for by these four polymorphisms. The associations between the haplotypes and coronary heart disease were analyzed with the haplo.stats program, adjusted for age, sex and smoking. RESULTS: Patients homozygous for the C allele of the 4072 T>C polymorphism had a 34% increased coronary heart disease risk (P = 0.017) compared to patients homozygous for the T allele. Haplotype H3, consisting of the minor allele of the 4072T>C polymorphism and the major alleles of the other polymorphisms, had a frequency of 15% and was associated with a 45% increased coronary heart disease risk (P = 0.006) compared to the wild-type haplotype H1. CONCLUSIONS: We conclude that genetic variation in the angiotensinogen gene contributes to coronary heart disease risk in patients with familial hypercholesterolemia.     

Effects of HMGB1 on ischemia-reperfusion injury in the rat heart

BACKGROUND: Coronary ischemia-reperfusion (I/R) injury causes cardiomyocyte necrosis in a multi-step process that includes an inflammatory reaction. A recent study has suggested that high-mobility group box 1 (HMGB1) is a late mediator of lethal sepsis and an early mediator of inflammation and necrosis following I/R injury. In the present study a neutralizing monoclonal antibody (mAb) for HMGB1 was used to clarify the role of HMGB1 in cardiac I/R injury. METHODS AND RESULTS: Rats underwent 30 min of left coronary artery occlusion followed by 60 min reperfusion. An intravenous injection of anti-HMGB1 mAb or control IgG was administered just before reperfusion. The infarct size was enlarged in the anti-HMGB1 mAb group in comparison with the control group (p<0.05). The treatment of anti-HMGB1 mAb significantly increased the plasma troponin-T and norepinephrine (NE) content in the heart in comparison with the control (p<0.05). Moreover, the production of dihydroxyphenylglycol was reduced in the anti-HMGB1-treated group (p<0.05). CONCLUSION: This study shows for the first time the effects of treatment with neutralizing anti-HMGB1 mAb on I/R injury in the rat heart. The findings support the novel view that I/R-induced HMGB1 may be an important factor in the modulation of interstitial NE.     

Protease-activated receptor-2 modulates myocardial ischemia-reperfusion injury in the rat heart

Protease-activated receptor-2 (PAR-2) is a member of seven transmembrane domain G protein-coupled receptors activated by proteolytic cleavage whose better known member is the thrombin receptor. The pathophysiological role of PAR-2 remains poorly understood. Because PAR-2 is involved in inflammatory and injury response events, we investigated the role of PAR-2 in experimental myocardial ischemia-reperfusion injury. We show for the first time that PAR-2 activation protects against reperfusion-injury. After PAR-2-activating peptide (2AP) infusion, we found a significant recovery of myocardial function and decrease in oxidation at reflow. Indeed, the glutathione cycle (glutathione and oxidized glutathione) and lipid peroxidation analysis showed a reduced oxidative reperfusion-injury. Moreover, ischemic risk zone and creatine kinase release were decreased after PAR-2AP treatment. These events were coupled to elevation of PAR-2 and tumor necrosis factor alpha (TNFalpha) expression in both nuclear extracts and whole heart homogenates. The recovery of coronary flow was not reverted by L-nitroarginine methylester, indicating a NO-independent pathway for this effect. Genistein, a tyrosine kinase inhibitor, did not revert the PAR-2AP effect. During early reperfusion injury in vivo not only oxygen radicals are produced but also numerous proinflammatory mediators promoting neutrophil and monocyte targeting. In this context, we show that TNFalpha and PAR-2 are involved in signaling in pathophysiological conditions, such as myocardial ischemia-reperfusion. At the same time, because TNFalpha may exert pro-inflammatory actions and PAR-2 may constitute one of the first protective mechanisms that signals a primary inflammatory response, our data support the concept that this network may regulate body responses to tissue injury.     

Intravascular hemolysis increases atherogenicity of diet-induced hypercholesterolemia in rabbits in spite of heme oxygenase-1 gene and protein induction

Free radical mediated oxidation of apoB lipoproteins in the arterial intima appears to contribute to atherogenicity of the entrapped particles. A plausible pathogenic mechanism for oxidation is the one induced by heme leaking from erythrocytes that is then carried into the arterial wall by its high affinity for lipoproteins. In the intima, in the presence of H(2)O(2) secreted by macrophages, heme can be a potent oxidant. To study the role of heme as a promoter of oxidative stress damage in vivo we used a model of intravascular hemolysis (IVH) caused by phenylhydrazine in rabbits with and without diet-induced moderate hypercholesterolemia (MHC). Evaluation of the antioxidant status of plasma indicated that at the end of the treatment period this was compromised by the MHC-IVH. After 10 weeks the animals with combined MHC-IVH showed more of the aorta surface covered by lesions (27%+/-8, mean (SD) than the animals with only MHC (11%+/-7), in spite of having similar plasma levels of VLDL+LDL lipoproteins. The animals with only IVH, as well as the controls, showed minimal lesions (<1%). Heme oxygenase (HO-1) expression in aorta and other tissues was markedly increased in the group with MHC-IVH and it was correlated with the extent of IVH. The data suggest that the oxidative stress associated with IVH potentiates the atherogenicity of moderate hypercholesterolemia and that in spite of a strong induction of HO-1 this is not sufficient to counteract the atherogenicity of the combined condition.