Effects of aging and reproductive history on bone parameters of common marmoset (Callithrix jacchus) mandibles.

To understand effects of aging and reproductive history in the bones of common marmosets (Callithrix jacchus), mandibles from 79 males and 66 females were analyzed. Dry bone specimen was prepared from dissected mandible, and analyzed using a dual-energy x-ray absorptiometry (DXA) measurement system in terms of bone weight (BnW), bone area (AREA), bone mineral content (BMC), bone mineral density (BMD, ratio of BMC to AREA) and bone mineral ratio (BMR, ratio of BMC to BnW). The mandible bones became porous and thicker with age. The age-related changes in BnW, AREA and BMC showed inflection points at around 1.5-2 Y and 13-15 Y. The period before 1.5-2 Y corresponds to the growth phase, the period between the inflection points is the aging phase, followed by senescence after the second inflection point. BMD increased until 1.5-2 Y and gradually decreased thereafter in males, with a more dramatic decrease in females, probably because of pregnancy and lactation. BMR was stable after reaching its peak by 1 Y, unlike the other parameters we analyzed. BMD of parous female tended to be lower than that of nulliparous female aged 2-5 Y. This study identified some of the particular effects of aging and reproductive history on characteristics of mandible bones in common marmoset.

Effects of aging and delivery order on the breeding capacity of F344/N rats.

Reproductive performance affects the efficiency of the production of animals. Therefore, knowing the reproductive properties of each species or strain of animal is important for proper management of breeding stocks. To elucidate the reproductive properties of female F344/N rats, frequently used for longevity and gerontology research, we monitored the breeding duration, number of deliveries, litter size and weaning rate of their pups. The first delivery was observed at 2.9 ± 0.1 (mean ± standard deviation) months of age (M) and the last delivery was at 15.1 ± 1.8 M. The number of deliveries was 10.4 ± 2.8 (range of 7-16) within the life span of the mother (24.7 ± 5.6 M). The litter sizes at the third (10.0 ± 3.0) or fourth (10.3 ± 2.7) deliveries were higher than for other deliveries. The breeding outcome declined after the fifth delivery yielding reduced litter size or weaning rate, number of delivered mother. These results suggests that though female F344/N rats are able to deliver more than 10 times, the reproductive performance lowered after fifth delivery.

Strategies for Highly Efficient Rabbit Sperm Cryopreservation.

The rabbit is a valuable animal for both the economy and biomedical sciences. Sperm cryopreservation is one of the most efficient ways to preserve rabbit strains because it is easy to collect ejaculate repeatedly from a single male and inseminate artificially into multiple females. During the cooling, freezing and thawing process of sperms, the plasma membrane, cytoplasm and genome structures could be damaged by osmotic stress, cold shock, intracellular ice crystal formation, and excessive production of reactive oxygen species. In this review, we will discuss the progress made during the past years regarding efforts to minimize the cell damage in rabbit sperms, including freezing extender, cryoprotectants, supplements, and procedures.

Small-volume vitrification and rapid warming yield high survivals of one-cell rat embryos in cryotubes†.

To cryopreserve cells, it is essential to avoid intracellular ice formation during cooling and warming. One way to achieve this is to convert the water inside the cells into a non-crystalline glass. It is currently believed that to accomplish this vitrification, the cells must be suspended in a very high concentration (20-40%) of a glass-inducing solute, and subsequently cooled very rapidly. Herein, we report that this belief is erroneous with respect to the vitrification of one-cell rat embryos. In the present study, one-cell rat embryos were vitrified with 5 µL of EFS10 (a mixture of 10% ethylene glycol (EG), 27% Ficoll, and 0.45 M sucrose) in cryotubes at a moderate cooling rate, and warmed at various rates. Survival was assessed according to the ability of the cells to develop into blastocysts and to develop to term. When embryos were vitrified at a 2613 °C/min cooling rate and thawed by adding 1 mL of sucrose solution (0.3 M, 50 °C) at a warming rate of 18 467 °C/min, 58.1 ± 3.5% of the EFS10-vitrified embryos developed into blastocysts, and 50.0 ± 4.7% developed to term. These rates were similar to those of non-treated intact embryos. Using a conventional cryotube, we achieved developmental capabilities in one-cell rat embryos by rapid warming that were comparable to those of intact embryos, even using low concentrations (10%) of cell-permeating cryoprotectant and at low cooling rates.

Whole-body insulin resistance and energy expenditure indices, serum lipids, and skeletal muscle metabolome in a state of lipoprotein lipase overexpression.

INTRODUCTION: Overexpression of lipoprotein lipase (LPL) protects against high-fat-diet (HFD)-induced obesity and insulin resistance in transgenic rabbits; however, the molecular mechanisms remain unclear. Skeletal muscle is a major organ responsible for insulin-stimulated glucose uptake and energy expenditure. OBJECTIVES: The main purpose of the current study was to examine the effects of the overexpression of LPL on the skeletal muscle metabolomic profiles to test our hypothesis that the mitochondrial oxidative metabolism would be activated in the skeletal muscle of LPL transgenic rabbits and that the higher mitochondrial oxidative metabolism activity would confer better phenotypic metabolic outcomes. METHODS: Under a HFD, insulin resistance index was measured using the intravenous glucose tolerance test, and total energy expenditure (TEE) was measured by doubly-labeled water in control and LPL transgenic rabbits (n = 12, each group). Serum lipids, such as triglycerides and free fatty acid, were also measured. The skeletal muscle metabolite profile was analyzed using capillary electrophoresis time-of flight mass spectrometry in the two groups (n = 9, each group). A metabolite set enrichment analysis (MSEA) with muscle metabolites and a false discovery rate q < 0.2 was performed to identify significantly different metabolic pathways between the 2 groups. RESULTS: The triglycerides and free fatty acid levels and insulin resistance index were lower, whereas the TEE was higher in the LPL transgenic rabbits than in the control rabbits. Among 165 metabolites detected, the levels of 37 muscle metabolites were significantly different between the 2 groups after false discovery rate correction (q < 0.2). The MSEA revealed that the TCA cycle and proteinogenic amino acid metabolism pathways were significantly different between the 2 groups (P < 0.05). In the MSEA, all four selected metabolites for the TCA cycle (2-oxoglutaric acid, citric acid, malic acid, fumaric acid), as well as eight selected metabolites for proteinogenic amino acid metabolism (asparagine, proline, methionine, phenylalanine, histidine, arginine, leucine, isoleucine) were consistently increased in the transgenic rabbits compared with control rabbits, suggesting that these two metabolic pathways were activated in the transgenic rabbits. Some of the selected metabolites, such as citric acid and methionine, were significantly associated with serum lipids and insulin resistance (P < 0.05). CONCLUSION: The current results suggest that the overexpression of LPL may lead to increased activities of TCA cycle and proteinogenic amino acid metabolism pathways in the skeletal muscle, and these enhancements may play an important role in the biological mechanisms underlying the anti-obesity/anti-diabetes features of LPL overexpression.

Endothelial Lipase Exerts its Anti-Atherogenic Effect through Increased Catabolism of ß-VLDLs.

AIM: Endothelial lipase (EL) plays an important role in lipoprotein metabolism. Our recent study showed that increased hepatic expression of EL attenuates diet-induced hypercholesterolemia, thus subsequently reducing atherosclerosis in transgenic (Tg) rabbits. However, it is yet to be determined whether increased EL activity itself per se is anti-atherogenic or whether the anti-atherogenic effect of EL is exclusively dependent on its lipid-lowering effect. METHODS: To determine the mechanisms underlying EL-mediated anti-atherogenic effect, we fed Tg and non-Tg rabbits diets containing different amounts of cholesterol to make their plasma cholesterol levels similarly high. Sixteen weeks later, we examined their lipoprotein profiles and compared their susceptibility to atherosclerosis. RESULTS: With Tg and non-Tg rabbits having hypercholesterolemia, the plasma lipids and lipoprotein profiles were observed to be similar, while pathological examinations revealed that lesion areas of both aortic and coronary atherosclerosis of Tg rabbits were not significantly different from non-Tg rabbits. Moreover, Tg rabbits exhibited faster clearance of DiI-labeled ß-VLDLs than non-Tg rabbits. CONCLUSION: The results of our study suggest that the enhancement of ß-VLDL catabolism is the major mechanism for atheroprotective effects of EL in Tg rabbits.

Candidate genes involved in the defunctionalization and refunctionalization of the urinary bladder induced by bladder anuria and reperfusion.

AIMS: We develop a novel rabbit urinary diversion model of bladder defunctionalization due to bladder anuria followed by refunctionalization due to urine reperfusion to investigate the molecular biological background. To validate the results, we used reverse transcription-polymerase chain reaction (RT-PCR) to analyze human specimens from defunctionalized bladders in patients receiving dialysis before kidney transplantation. METHODS: Female rabbits were divided into three groups: control, defunctionalized, and refunctionalized. The bilateral ureters were anastomosed to vagina in the defunctionalized and refunctionalized groups at 0 weeks. In the refunctionalized group, the unilateral ureter was reanastomosed to the bladder at 8 weeks. RESULTS: The capacity and compliance of the rabbit bladder in the refunctionalized group were significantly lower than those in the control group at 8 weeks and higher than those in the defunctionalized group at 14 weeks. The significant downregulation of IGFBP2, UPK1B, and CST6 in the defunctionalized group compared with that in the control groups, and the significant downregulation of AGTR2 in the refunctionalized group compared with that in the defunctionalized group in the rabbit bladder-muscle DNA microarray were validated by RT-PCR. Human bladder muscle indicated significant downregulation of UPK1B and CST6 and significant downregulation of IGFBP2 in the defunctionalized group, which is consistent with both rabbit bladder-muscle DNA microarray and rabbit bladder RT-PCR results. CONCLUSIONS: The present study using novel model of bladder defunctionalization followed by refunctionalization indicated the consistent downregulation of UPK1B and CST6 in muscle and the consistent downregulation of IGFBP2 in mucosa in process of bladder defunctionalization, which was validated by human specimens.

Macrophage-derived MMP-9 enhances the progression of atherosclerotic lesions and vascular calcification in transgenic rabbits.

Matrix metalloproteinase-9 (MMP-9), or gelatinase B, has been hypothesized to be involved in the progression of atherosclerosis. In the arterial wall, accumulated macrophages secrete considerable amounts of MMP-9 but its pathophysiological functions in atherosclerosis have not been fully elucidated. To examine the hypothesis that macrophage-derived MMP-9 may affect atherosclerosis, we created MMP-9 transgenic (Tg) rabbits to overexpress the rabbit MMP-9 gene under the control of the scavenger receptor A enhancer/promoter and examined their susceptibility to cholesterol diet-induced atherosclerosis. Tg rabbits along with non-Tg rabbits were fed a cholesterol diet for 16 and 28 weeks, and their aortic and coronary atherosclerosis was compared. Gross aortic lesion areas were significantly increased in female Tg rabbits at 28 weeks; however, pathological examination revealed that all the lesions of Tg rabbits fed a cholesterol diet for either 16 or 28 weeks were characterized by increased monocyte/macrophage accumulation and prominent lipid core formation compared with those of non-Tg rabbits. Macrophages isolated from Tg rabbits exhibited higher infiltrative activity towards a chemoattractant, MCP-1 in vitro and augmented capability of hydrolysing extracellular matrix in granulomatous tissue. Surprisingly, the lesions of Tg rabbits showed more advanced lesions with remarkable calcification in both aortas and coronary arteries. In conclusion, macrophage-derived MMP-9 facilitates the infiltration of monocyte/macrophages into the lesions thereby enhancing the progression of atherosclerosis. Increased accumulation of lesional macrophages may promote vascular calcification.

Characteristic features of newly established specific pathogen-free albino large rabbit (JW-AKT): Comparison with Japanese White and New Zealand White rabbits.

The present study was conducted to reveal characteristic features of albino large rabbit (JW-AKT) which we formerly established a specific pathogen-free (SPF) colony. Body weights of JW-AKT rabbit at 52 weeks old was 5.7 ± 0.4 kg in males and 6.4 ± 0.4 kg in females. Weight of body, heart, lung and kidney in JW-AKT rabbit was significantly higher than in Japanese white and New Zealand white rabbits in both sexes. Though the body weight (BW) was rather lower in males, body length and brain weights tended to be higher in males than in females. Since body fat was significantly higher in females, what affects difference in BW is body fat, rather than the physical constitution of female JW-AKT rabbit. No critical sex difference was found in hematological parameters in JW-AKT rabbit. The results indicated that JW-AKT were about 1.5 times larger than the general laboratory rabbits with common properties in hematology. Thus, JW-AKT rabbit could be used as a novel SPF experimental animal model with some advantages in surgical experiments or collection of large amount of biological specimen.

Age-dependent changes of the mandible bone throughout the lifespan in female F344/N rat.

Age-dependent changes of the mandible bone in female F344/N rats, aged 22-1196 days, were analyzed using physiological bone properties and morphology. Bone weight, bone area, bone mineral components, and bone mineral density were assessed using dual-energy X-ray absorptiometry. The bone weight, bone area, bone mineral components, and bone mineral density increased rapidly until approximately 150 days of age, increased gradually thereafter, and then stabilized or decreased after 910 days of age. The ratio of bone mineral components to bone weight (bone mineral ratio) increased rapidly until approximately 43 days of age and stabilized thereafter. Size of the mandible, which was measured at 13 points on mandible surface, increased with age, and the rate of change showed a similar pattern to the other parameters. From a principal component analysis on morphometric measurements, principal component 1 (size factor) increased proportionally with age, whereas principal component 2 (shape factor) decreased until approximately 88 days of age and then increased after 365 days of age. As a result, the scatter plots for principal component 1 and principal component 2 were V-shaped, which indicates that the mandible developed in size, with deformation at younger ages, and recovered its original shape later in life. Our results revealed the occurrence of inflection points at approximately 43, 88, 150, 365, and 910 days of age. Some of these ages corresponded to transition points revealed by the age-dependent changes of the occlusal mandibular condyle and tooth wear in the same rat.

Vitrification of one-cell mouse embryos in cryotubes.

Preventing intracellular ice formation is essential to cryopreserve cells. Prevention can be achieved by converting cell water into a non-crystalline glass, that is, to vitrify. The prevailing belief is that to achieve vitrification, cells must be suspended in a solution containing a high concentration of glass-inducing solutes and cooled rapidly. In this study, we vitrified 1-cell mouse embryos and examined the effect of the cooling rate, the warming rate, and the concentration of cryoprotectant on cell survival. Embryos were vitrified in cryotubes. The vitrification solutions used were EFS20, EFS30, and EFS40, which contained ethylene glycol (20, 30 and 40% v/v, respectively), Ficoll (24%, 21%, and 18% w/v, respectively) and sucrose (0.4 0.35, and 0.3 M, respectively). A 5-µl EFS solution suspended with 1-cell embryos was placed in a cryotube. After 2 min in an EFS solution at 23 °C, embryos were vitrified by direct immersion into liquid nitrogen. The sample was warmed at 34 °C/min, 4,600 °C/min and 6,600 °C/min. With EFS40, the survival was low regardless of the warming rate. With EFS30 and EFS20, survival was also low when the warming rate was low, but increased with higher warming rates, likely due to prevention of intracellular ice formation. When 1-cell embryos were vitrified with EFS20 and warmed rapidly, almost all of the embryos developed to blastocysts in vitro. Moreover, when vitrified 1-cell embryos were transferred to recipients at the 2-cell stage, 43% of them developed to term. In conclusion, we developed a vitrification method for 1-cell mouse embryos by rapid warming using cryotubes.

Increased Hepatic Expression of Endothelial Lipase Inhibits Cholesterol Diet-Induced Hypercholesterolemia and Atherosclerosis in Transgenic Rabbits.

OBJECTIVE: Endothelial lipase (EL) is a key determinant in plasma high-density lipoprotein-cholesterol. However, functional roles of EL on the development of atherosclerosis have not been clarified. We investigated whether hepatic expression of EL affects plasma lipoprotein metabolism and cholesterol diet-induced atherosclerosis. APPROACH AND RESULTS: We generated transgenic (Tg) rabbits expressing the human EL gene in the liver and then examined the effects of EL expression on plasma lipids and lipoproteins and compared the susceptibility of Tg rabbits with cholesterol diet-induced atherosclerosis with non-Tg littermates. On a chow diet, hepatic expression of human EL in Tg rabbits led to remarkable reductions in plasma levels of total cholesterol, phospholipids, and high-density lipoprotein-cholesterol compared with non-Tg controls. On a cholesterol-rich diet for 16 weeks, Tg rabbits exhibited significantly lower hypercholesterolemia and less atherosclerosis than non-Tg littermates. In Tg rabbits, gross lesion area of aortic atherosclerosis was reduced by 52%, and the lesions were characterized by fewer macrophages and smooth muscle cells compared with non-Tg littermates. CONCLUSIONS: Increased hepatic expression of EL attenuates cholesterol diet-induced hypercholesterolemia and protects against atherosclerosis.

Motility and fertility of rabbit sperm cryopreserved using soybean lecithin as an alternative to egg yolk.

This study was conducted to investigate whether soy lecithin can be used as an alternative cryoprotectant to establish a procedure that does not require the use of egg yolk to cryopreserve rabbit strains. Semen from Japanese White rabbits was frozen with HEPES extender containing 20% egg yolk (EYH), 0.5% (Lec-0.5), 1.5% (Lec-1.5), 2.5% (Lec-2.5), or 3.5% (Lec-3.5; wt/vol) lecithin (type IV-S, ≥30%), and the motility of thawed sperm was analyzed. The sperm motility in the Lec-1.5 group was significantly higher than that in the Lec-2.5 and 3.5 groups and equivalent to the EYH group. From 17 rounds of artificial insemination with frozen-thawed sperm in the EYH and Lec-1.5 groups, 12 rabbits in both groups were pregnant (70.6%) and delivered offspring. The litter size was 3.3 in the EYH group and 5.1 in the Lec-1.5 group. These results indicate that soy lecithin can be used as a substitute for egg yolk as a cryoprotectant on the basis of motility and fertility of the frozen-thawed rabbit sperm and that 1.5% lecithin (type IV-S, ≥30%) in the semen extender was the optimum concentration for rabbit sperm cryopreservation.

Effects of type III antifreeze protein on sperm and embryo cryopreservation in rabbit.

We investigated the effects of antifreeze protein (AFP) III supplementation on the cryopreservation of rabbit sperm cells and embryos. Ejaculated semen was collected from male Japanese white (JW) rabbits and divided into four AFP-supplemented groups (0.1 µg/ml, 1 µg/ml, 10 µg/ml, 100 µg/ml) and one control group with no AFP-supplementation. The semen samples were treated with egg-yolk HEPES extender containing 6% acetamide before the sperm was cooled from room temperature to 5 °C, then packed into sperm straws. The straws were frozen in steam of liquid nitrogen (LN2) and then preserved in the LN2. The motility of the sperm after thawing in 37 °C water was analyzed. The percentage of rapidly motile sperm in the 1 µg/ml AFP group was significantly higher than in the control group. Morulae were collected from female JW rabbits and divided into three AFP-supplemented groups (100 ng/ml, 500 ng/ml, 1000 ng/ml) and one control group. The morulae, immersed in an embryo-freezing solution (M199-HEPES containing 20% ethylene glycol, 20% dimethylsulfoxide, 10% fetal bovine serum and 0.25 M sucrose), were packed into open pulled embryo straws and vitrified in LN2. The frozen embryos were thawed in the embryo-freezing solution, and the rates of embryo survival and development to blastocyte stage were analyzed after incubation for 72 h. The development rate of the embryos in the 500 ng/ml AFP group was significantly higher than in the control group, but that in the 1000 ng/ml AFP group was significantly lower. In conclusion, the appropriate dose of AFP III increased the number of rapidly motile sperm and embryo survival following freezing and thawing. The results suggest that supplementation with AFP III can increase the efficiency of cryopreservation of rabbit sperm cells and embryos.

Effects of cholesterol-loaded cyclodextrins on the rate and the quality of motility in frozen and thawed rabbit sperm.

The motility of sperm after freezing and thawing is critical for effective cryopreservation. It is known that supplementation with cholesterol-loaded cyclodextrin (CLC) improves cryosurvival of sperm in various animals. To clarify the effects of supplementation with CLC on rabbit sperm motility after freezing and thawing, rabbit sperm motility was analyzed using a computer-assisted sperm analysis system. Sperm motility with CLC supplementation was 29.4 ± 9.6% (mean ± SD), which was significantly higher than that of controls (20.8 ± 7.1%, P<0.05). The curvilinear velocity of sperm with CLC exceeded that of controls, whereas the values for linearity and wobble were significantly lower in sperm with CLC compared with controls. After artificial insemination, 44.3% of recovered ova were fertilized in the CLC-supplemented group, which was higher than the percentage in the control group (36.4%). The results indicate that supplementation with CLC improves the rate and quality of motility in rabbit sperm after freezing and thawing, and would be advantageous for successful cryopreservation.

Populations of follicles in F344/N rats during aging.

Follicular populations were investigated in female F344/N rats to better understand the aging process of the rat ovary. Ovaries dissected at various ages (spanning 1-36 months old) were submitted for histological examination. The total number of primordial, growing (primary and secondary), tertiary, and atretic follicles as well as corpora lutea (CL) were counted in hematoxylin-eosin- and azocarmine-aniline-blue-stained ovarian sections. The number of healthy follicles including primordial, growing and tertiary follicles decreased rapidly between the first and third months and gradually thereafter. CL were found in 3-month-old rats, and their number remained unchanged until 18 months of age, at which point it decreased. The number of atretic follicles started to increase in rats older than 18 months, which corresponded to the cessation of estrous cyclicity. Several healthy follicles and CL were observed even in 36-month-old rats.

Human apolipoprotein A-II protects against diet-induced atherosclerosis in transgenic rabbits.

OBJECTIVE: Apolipoprotein (apo) A-II is the second major apo of high-density lipoproteins, yet its pathophysiological roles in the development of atherosclerosis remain unknown. We aimed to examine whether apo A-II plays any role in atherogenesis and, if so, to elucidate the mechanism involved. METHODS AND RESULTS: We compared the susceptibility of human apo A-II transgenic (Tg) rabbits to cholesterol diet-induced atherosclerosis with non-Tg littermate rabbits. Tg rabbits developed significantly less aortic and coronary atherosclerosis than their non-Tg littermates, while total plasma cholesterol levels were similar. Atherosclerotic lesions of Tg rabbits were characterized by reduced macrophages and smooth muscle cells, and apo A-II immunoreactive proteins were frequently detected in the lesions. Tg rabbits exhibited low levels of plasma C-reactive protein and blood leukocytes compared with non-Tg rabbits, and high-density lipoproteins of Tg rabbit plasma exerted stronger cholesterol efflux activity and inhibitory effects on the inflammatory cytokine expression by macrophages in vitro than high-density lipoproteins isolated from non-Tg rabbits. In addition, ß-very-low-density lipoproteins of Tg rabbits were less sensitive to copper-induced oxidation than ß-very-low-density lipoproteins of non-Tg rabbits. CONCLUSIONS: These results suggest that enrichment of apo A-II in high-density lipoprotein particles has atheroprotective effects and apo A-II may become a target for the treatment of atherosclerosis.

Life span of common marmoset (Callithrix jacchus) at CLEA Japan breeding colony.

The life span and survival parameters of the common marmoset (Callithrix jacchus) in a breeding colony at CLEA Japan, Inc. were investigated. The average life span of male marmosets was 148.5 ± 6.1 (mean ± SE) months of age (M), which was significantly longer (P < 0.01) than that of females (111.7 ± 6.0 M). Additionally, the male population reached 25-, 50-, 75- and 90 %-mortality at an older age than the female population. However, the maximum life span in males (259.9 M) was shorter than in females (262.5 M). The survival of females shows a relatively continuous decline; however, the male marmosets show a slight decline in survival during the first 7-9 years and then a dramatic decrease and another slight decline after 14-16 year of age in survival, i.e., a lifespan curve similar to what is observed in colonies of aging rodents and humans. The sex-associated difference in life span was caused by reproductive burden on the females. The present study reported a longer than expected life span of the marmoset, and a long-lived animal can be a powerful model for senescence and longevity sciences.

Endothelial lipase mediates HDL levels in normal and hyperlipidemic rabbits.

AIM: Existing evidence suggests that endothelial lipase (EL) plays an important role in high-densitylipoprotein (HDL) metabolism. Because rabbits are a useful animal model for the study of human lipid metabolism and atherosclerosis, we characterized rabbit EL (rEL) expression and investigated its relationship with plasma HDL levels in normal and hyperlipidemic rabbits. METHODS: We cloned the rEL cDNA and analyzed the EL tissue expression using Northern blotting, real-time RT-PCR, Western blotting, and in situ hybridization. We evaluated the effects of rEL antisense on plasma HDL levels. RESULTS: We found that rEL mRNA was highly expressed in cholesterol synthesis-related organs, including the liver, testis, and adrenal along with its expression in the lung, kidney, bone marrow, and small intestine. Interestingly, Watanabe heritable hyperlipidemic (WHHL) rabbits, a model of human familial hypercholesterolemia, had lower plasma levels of HDLs than normal rabbits. The plasma HDL levels in WHHL rabbits were inversely associated with high levels of plasma rEL proteins and hepatic expression of rEL mRNA. Injection of rEL-specific antisense oligonucleotides into rabbits resulted in the elevation of plasma large HDLs. Furthermore, we demonstrated that rEL mRNA was expressed by both endothelial cells and macrophages in the lesions of aortic atherosclerosis of WHHL rabbits. CONCLUSIONS: rEL is expressed in multiple tissues and may have many physiological and pathophysiological functions, such as in the regulation of cholesterol metabolism and atherosclerosis. Our results suggest that EL is an important regulator of plasma HDL levels in rabbits.