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1.  GPIHBP1 and the intravascular processing of triglyceride-rich lipoproteins 
Journal of internal medicine  2012;272(6):528-540.
Lipoprotein lipase (LPL) is produced by parenchymal cells, mainly adipocytes and myocytes, but its role in hydrolyzing triglycerides in plasma lipoproteins occurs at the capillary lumen. For decades, the mechanism by which LPL reached its site of action in capillaries was unclear, but this mystery was recently solved. GPIHBP1, a GPI-anchored protein of capillary endothelial cells, picks up LPL from the interstitial spaces and shuttles it across endothelial cells to the capillary lumen. When GPIHBP1 is absent, LPL is mislocalized to the interstitial spaces, leading to severe hypertriglyceridemia. Some cases of hypertriglyceridemia in humans are caused by GPIHBP1 mutations that interfere with GPIHBP1's ability to bind LPL, and some are caused by LPL mutations that impair LPL's ability to bind to GPIHBP1. This review will cover recent progress in understanding GPIHBP1's role in health and disease and will discuss some remaining mysteries surrounding the processing of triglyceride-rich lipoproteins.
doi:10.1111/joim.12003
PMCID: PMC3940157  PMID: 23020258
hypertriglyceridemia; chylomicronemia; GPIHBP1; lipoprotein lipase; endothelial cells; lymphocyte antigen 6 proteins
2.  Chylomicronemia mutations yield new insights into interactions between lipoprotein lipase and GPIHBP1 
Human Molecular Genetics  2012;21(13):2961-2972.
Lipoprotein lipase (LPL) is a 448-amino-acid head-to-tail dimeric enzyme that hydrolyzes triglycerides within capillaries. LPL is secreted by parenchymal cells into the interstitial spaces; it then binds to GPIHBP1 (glycosylphosphatidylinositol-anchored high density lipoprotein-binding protein 1) on the basolateral face of endothelial cells and is transported to the capillary lumen. A pair of amino acid substitutions, C418Y and E421K, abolish LPL binding to GPIHBP1, suggesting that the C-terminal portion of LPL is important for GPIHBP1 binding. However, a role for LPL's N terminus has not been excluded, and published evidence has suggested that only full-length homodimers are capable of binding GPIHBP1. Here, we show that LPL's C-terminal domain is sufficient for GPIHBP1 binding. We found, serendipitously, that two LPL missense mutations, G409R and E410V, render LPL susceptible to cleavage at residue 297 (a known furin cleavage site). The C terminus of these mutants (residues 298–448), bound to GPIHBP1 avidly, independent of the N-terminal fragment. We also generated an LPL construct with an in-frame deletion of the N-terminal catalytic domain (residues 50–289); this mutant was secreted but also was cleaved at residue 297. Once again, the C-terminal domain (residues 298–448) bound GPIHBP1 avidly. The binding of the C-terminal fragment to GPIHBP1 was eliminated by C418Y or E421K mutations. After exposure to denaturing conditions, the C-terminal fragment of LPL refolds and binds GPIHBP1 avidly. Thus, the binding of LPL to GPIHBP1 requires only the C-terminal portion of LPL and does not depend on full-length LPL homodimers.
doi:10.1093/hmg/dds127
PMCID: PMC3373243  PMID: 22493000
3.  Transgenic expression and genetic variation of LMF1 affect LPL activity in mice and humans 
Objective
Lipoprotein lipase (LPL) is a principal enzyme in lipoprotein metabolism, tissue lipid utilization and energy metabolism. LPL is synthesized by parenchymal cells in adipose, heart and muscle tissues followed by secretion to extracellular sites, where lipolyic function is exerted. The catalytic activity of LPL is attained during post-translational maturation, which involves glycosylation, folding and subunit assembly within the endoplasmic reticulum (ER). A lipase-chaperone, lipase maturation factor 1 (Lmf1), has recently emerged as a critical factor in this process. Previous studies demonstrated that loss-of-function mutations of Lmf1 result in diminished lipase activity and severe hypertriglyceridemia in mice and human subjects. The objective of this study is to investigate whether, beyond its role as a required factor in lipase maturation, variation in Lmf1 expression is sufficient to modulate LPL activity in vivo.
Methods and Results
To assess the effects of Lmf1 overexpression in adipose and muscle tissues, we generated aP2-Lmf1 and Mck-Lmf1 transgenic mice. Characterization of relevant tissues revealed increased LPL activity in both mouse strains. In the omental and subcutaneous adipose depots, Lmf1 overexpression was associated with increased LPL specific activity without changes in LPL mass. In contrast, increased LPL activity was due to elevated LPL protein level in heart and gonadal adipose tissue. To extend these studies to humans, we detected association between LMF1 gene variants and post-heparin LPL activity in a dyslipidemic cohort.
Conclusions
Our results suggest that variation in Lmf1 expression is a post-translational determinant of LPL activity.
doi:10.1161/ATVBAHA.112.245696
PMCID: PMC3331946  PMID: 22345169
4.  Localization of lipoprotein lipase and GPIHBP1 in mouse pancreas: effects of diet and leptin deficiency 
BMC Physiology  2012;12:14.
Background
Lipoprotein lipase (LPL) hydrolyzes triglycerides in plasma lipoproteins and enables uptake of lipolysis products for energy production or storage in tissues. Our aim was to study the localization of LPL and its endothelial anchoring protein glycosylphosphatidylinositol-anchored high density lipoprotein-binding protein 1 (GPIHBP1) in mouse pancreas, and effects of diet and leptin deficiency on their expression patterns. For this, immunofluorescence microscopy was used on pancreatic tissue from C57BL/6 mouse embryos (E18), adult mice on normal or high-fat diet, and adult ob/ob-mice treated or not with leptin. The distribution of LPL and GPIHBP1 was compared to insulin, glucagon and CD31. Heparin injections were used to discriminate between intracellular and extracellular LPL.
Results
In the exocrine pancreas LPL was found in capillaries, and was mostly co-localized with GPIHBP1. LPL was releasable by heparin, indicating localization on cell surfaces. Within the islets, most of the LPL was associated with beta cells and could not be released by heparin, indicating that the enzyme remained mostly within cells. Staining for LPL was found also in the glucagon-producing alpha cells, both in embryos (E18) and in adult mice. Only small amounts of LPL were found together with GPIHBP1 within the capillaries of islets. Neither a high fat diet nor fasting/re-feeding markedly altered the distribution pattern of LPL or GPIHBP1 in mouse pancreas. Islets from ob/ob mice appeared completely deficient of LPL in the beta cells, while LPL-staining was normal in alpha cells and in the exocrine pancreas. Leptin treatment of ob/ob mice for 12 days reversed this pattern, so that most of the islets expressed LPL in beta cells.
Conclusions
We conclude that both LPL and GPIHBP1 are present in mouse pancreas, and that LPL expression in beta cells is dependent on leptin.
doi:10.1186/1472-6793-12-14
PMCID: PMC3537605  PMID: 23186339
Lipoprotein lipase; Diabetes mellitus; Islet cells; Exocrine pancreas; Endothelium; Ob/ob mice; High fat diet; Heparin; qPCR; Immunofluorescence
5.  Cholesterol Intake Modulates Plasma Triglyceride Levels in GPIHBP1-deficient Mice 
Objective
Adult GPIHBP1-deficient mice (Gpihbp1−/−) have severe hypertriglyceridemia; however, the plasma triglyceride levels are only mildly elevated during the suckling phase when lipoprotein lipase (Lpl) is expressed at high levels in the liver. Lpl expression in the liver can be induced in adult mice with dietary cholesterol. We therefore hypothesized that plasma triglyceride levels in adult Gpihbp1−/− mice would be sensitive to cholesterol intake.
Methods and Results
After 4–8 weeks on a western diet containing 0.15% cholesterol, plasma triglyceride levels in Gpihbp1−/− mice were 10,000–12,000 mg/dl. When 0.005% ezetimibe was added to the diet to block cholesterol absorption, Lpl expression in the liver was reduced significantly, and the plasma triglyceride levels were significantly higher (>15,000 mg/dl). We also assessed plasma triglyceride levels in Gpihbp1−/− mice fed western diets containing either high (1.3%) or low (0.05%) amounts of cholesterol. The high-cholesterol diet significantly increased Lpl expression in the liver and lowered plasma triglyceride levels.
Conclusions
Treatment of Gpihbp1−/− mice with ezetimibe lowers Lpl expression in the liver and increases plasma triglyceride levels. A high-cholesterol diet had the opposite effects. Thus, cholesterol intake modulates plasma triglyceride levels in Gpihbp1−/− mice.
doi:10.1161/ATVBAHA.110.214403
PMCID: PMC2959134  PMID: 20814015
lipoprotein lipase; chylomicronemia; hypertriglyceridemia; GPIHBP1
6.  Deletion of GPIHBP1 causing severe chylomicronemia 
Lipoprotein lipase (LPL) is a hydrolase that cleaves circulating triglycerides to release fatty acids to the surrounding tissues. The enzyme is synthesized in parenchymal cells and is transported to its site of action on the capillary endothelium by glycophosphatidylinositol (GPI)-anchored high-density lipoprotein-binding protein 1 (GPIHBP1). Inactivating mutations in LPL; in its cofactor, apolipoprotein (Apo) C2; or in GPIHBP1 cause severe hypertriglyceridemia. Here we describe an individual with complete deficiency of GPIHBP1. The proband was an Asian Indian boy who had severe chylomicronemia at 2 months of age. Array-based copy-number analysis of his genomic DNA revealed homozygosity for a 17.5-kb deletion that included GPIHBP1. A 44-year-old aunt with a history of hypertriglyceridemia and pancreatitis was also homozygous for the deletion. A bolus of intravenously administered heparin caused a rapid increase in circulating LPL and decreased plasma triglyceride levels in control individuals but not in two GPIHBP1-deficient patients. Thus, short-term treatment with heparin failed to attenuate the hypertriglyceridemia in patients with GPIHBP1 deficiency. The increasing resolution of copy number microarrays and their widespread adoption for routine cytogenetic analysis is likely to reveal a greater role for submicroscopic deletions in Mendelian conditions. We describe the first neonate with complete GPIHBP1 deficiency due to homozygosity for a deletion of GPIHBP1.
Electronic supplementary material
The online version of this article (doi:10.1007/s10545-011-9406-5) contains supplementary material, which is available to authorized users.
doi:10.1007/s10545-011-9406-5
PMCID: PMC3319888  PMID: 22008945
7.  GPIHBP1 Is Responsible for the Entry of Lipoprotein Lipase into Capillaries 
Cell metabolism  2010;12(1):42-52.
SUMMARY
The lipolytic processing of triglyceride-rich lipoproteins by lipoprotein lipase (LPL) is the central event in plasma lipid metabolism, providing lipids for storage in adipose tissue and fuel for vital organs such as the heart. LPL is synthesized and secreted by myocytes and adipocytes but then finds its way into the lumen of capillaries, where it hydrolyzes lipoprotein triglycerides. The mechanism by which LPL reaches the lumen of capillaries represents one of the most persistent mysteries of plasma lipid metabolism. Here, we show that GPIHBP1 is responsible for the transport of LPL into capillaries. In Gpihbp1-deficient mice, LPL is mislocalized to the interstitial spaces surrounding myocytes and adipocytes. Also, we show that GPIHBP1 is located at the basolateral surface of capillary endothelial cells and actively transports LPL across endothelial cells. Our experiments define the function of GPIHBP1 in triglyceride metabolism and provide a mechanism for the transport of LPL into capillaries.
doi:10.1016/j.cmet.2010.04.016
PMCID: PMC2913606  PMID: 20620994
9.  GPIHBP1—an endothelial cell molecule important for the lipolytic processing of chylomicrons 
Current opinion in lipidology  2007;18(4):389-396.
Purpose of review
This review summarizes recent data indicating that glycosylphosphatidylinositol-anchored high density lipoprotein–binding protein 1 (GPIHBP1) plays a key role in the lipolytic processing of chylomicrons.
Recent findings
Lipoprotein lipase (LpL) hydrolyzes triglycerides in chylomicrons at the luminal surface of the capillaries in heart, adipose tissue, and skeletal muscle. However, the endothelial cell molecule that facilitates the lipolytic processing of chylomicrons has never been clearly defined. Mice lacking GPIHBP1 manifest chylomicronemia, with plasma triglyceride levels as high as 5,000 mg/dl. In wild-type mice, GPIHBP1 is expressed on the luminal surface of capillaries in heart, adipose tissue, and skeletal muscle. Cells transfected with GPIHBP1 bind both chylomicrons and LpL avidly.
Summary
The chylomicronemia in Gpihbp1-deficient mice, the fact that GPIHBP1 is located within the lumen of capillaries, and the fact that GPIHBP1 binds LpL and chylomicrons suggest that GPIHBP1 is a key platform for the lipolytic processing of triglyceride-rich lipoproteins.
doi:10.1097/MOL.0b013e3281527914
PMCID: PMC2888298  PMID: 17620854
Chylomicronemia; lipoprotein lipase; hypertriglyceridemia; GPI-anchored proteins
10.  Chylomicronemia with Low Postheparin Lipoprotein Lipase Levels in the Setting of GPIHBP1 Defects 
Background
Recent studies in mice have established that an endothelial cell protein, GPIHBP1, is essential for the lipolytic processing of triglyceride-rich lipoproteins.
Methods and Results
We report the discovery of a homozygous missense mutation in GPIHBP1 in a young boy with severe chylomicronemia. The mutation, p.C65Y, replaces a conserved cysteine in the GPIHBP1’s Ly6 domain with a tyrosine and is predicted to perturb protein structure by interfering with the formation of a disulfide bond. Studies with transfected CHO cells showed that GPIHBP1-C65Y reaches the cell surface but has lost the ability to bind LPL. When the GPIHBP1-C65Y homozygote was given an intravenous bolus of heparin, only trace amounts of LPL entered the plasma. We also observed very low levels of LPL in the postheparin plasma of a chylomicronemic subject who was homozygous for a different GPIHBP1 mutation (p.Q115P). When the GPIHBP1-Q115P homozygote was given a 6-h infusion of heparin, significant amounts of LPL appeared in the plasma, resulting in a fall in the plasma triglyceride levels from 1780 mg/dl to 120 mg/dl.
Conclusions
We identified a novel GPIHBP1 missense mutation (p.C65Y) associated with defective LPL binding in a young boy with severe chylomicronemia. We also show that homozygosity for the C65Y or Q115P mutations is associated with low levels of LPL in the postheparin plasma, demonstrating that GPIHBP1 is important for plasma triglyceride metabolism in humans.
doi:10.1161/CIRCGENETICS.109.908905
PMCID: PMC2858258  PMID: 20124439
lipoprotein lipase; GPIHBP1; triglycerides
11.  Chylomicronemia With a Mutant GPIHBP1 (Q115P) That Cannot Bind Lipoprotein Lipase 
Objective
GPIHBP1 is an endothelial cell protein that binds lipoprotein lipase (LPL) and chylomicrons. Because GPIHBP1 deficiency causes chylomicronemia in mice, we sought to determine whether some cases of chylomicronemia in humans could be attributable to defective GPIHBP1 proteins.
Methods and Results
Patients with severe hypertriglyceridemia (n=60, with plasma triglycerides above the 95th percentile for age and gender) were screened for mutations in GPIHBP1. A homozygous GPIHBP1 mutation (c.344A>C) that changed a highly conserved glutamine at residue 115 to a proline (p.Q115P) was identified in a 33-year-old male with lifelong chylomicronemia. The patient had failure-to-thrive as a child but had no history of pancreatitis. He had no mutations in LPL, APOA5, or APOC2. The Q115P substitution did not affect the ability of GPIHBP1 to reach the cell surface. However, unlike wild-type GPIHBP1, GPIHBP1-Q115P lacked the ability to bind LPL or chylomicrons (d <1.006 g/mL lipoproteins from Gpihbp1−/− mice). Mouse GPIHBP1 with the corresponding mutation (Q114P) also could not bind LPL.
Conclusions
A homozygous missense mutation in GPIHBP1 (Q115P) was identified in a patient with chylomicronemia. The mutation eliminated the ability of GPIHBP1 to bind LPL and chylomicrons, strongly suggesting that it caused the patient’s chylomicronemia.
doi:10.1161/ATVBAHA.109.186577
PMCID: PMC2811263  PMID: 19304573
lipoprotein; lipase; human; chylomicronemia; hypertriglyceridemia; GPIHBP1
12.  GPIHBP1 and lipolysis: an update 
Current opinion in lipidology  2009;20(3):211-216.
Purpose of review
This review will provide an update on the structure of GPIHBP1, a 28-kDa glycosylphosphatidylinositol-anchored glycoprotein, and its role in the lipolytic processing of triglyceride-rich lipoproteins.
Recent findings
Gpihbp1 knockout mice on a chow diet have milky plasma and plasma triglyceride levels of more than 3000 mg/dl. GPIHBP1 is located on the luminal surface of endothelial cells in tissues where lipolysis occurs: heart, skeletal muscle, and adipose tissue. The pattern of lipoprotein lipase (LPL) release into the plasma after an intravenous injection of heparin is abnormal in Gpihbp1-deficient mice, suggesting that GPIHBP1 plays a direct role in binding LPL within the tissues of mice. Transfection of CHO cells with a GPIHBP1 expression vector confers on cells the ability to bind both LPL and chylomicrons. Two regions of GPIHBP1 are required for the binding of LPL – an amino-terminal acidic domain and the cysteine-rich Ly6 domain. GPIHBP1 expression in mice changes with fasting and refeeding and is regulated in part by peroxisome proliferator-activated receptor-γ.
Summary
GPIHBP1, an endothelial cell-surface glycoprotein, binds LPL and is required for the lipolytic processing of triglyceride-rich lipoproteins.
doi:10.1097/MOL.0b013e32832ac026
PMCID: PMC2810420  PMID: 19369870
chylomicrons; endothelial; lipoprotein lipase; PPARγ
13.  Normal binding of lipoprotein lipase, chylomicrons, and apo-AV to GPIHBP1 containing a G56R amino acid substitution 
Biochimica et biophysica acta  2007;1771(12):1464-1468.
Summary
GPIHBP1 is an endothelial cell protein that serves as a platform for lipoprotein lipase–mediated processing of triglyceride-rich lipoproteins within the capillaries of heart, adipose tissue, and skeletal muscle. The absence of GPIHBP1 causes severe chylomicronemia. A hallmark of GPIHBP1 is the ability to bind lipoprotein lipase, chylomicrons, and apolipoprotein (apo-) AV. A homozygous G56R mutation in GPIHBP1 was recently identified in two brothers with chylomicronemia, and the authors of that study suggested that the G56R substitution was responsible for the hyperlipidemia. In this study, we created a human GPIHBP1 expression vector, introduced the G56R mutation, and tested the ability of the mutant GPIHBP1 to reach the cell surface and bind lipoprotein lipase, chylomicrons, and apo-AV. Our studies revealed that the G56R substitution did not affect the ability of GPIHBP1 to reach the cell surface, nor did the amino acid substitution have any discernible effect on the binding of lipoprotein lipase, chylomicrons, or apo-AV.
doi:10.1016/j.bbalip.2007.10.005
PMCID: PMC2266775  PMID: 17997385
chylomicronemia; GPIHBP1; hypertriglyceridemia; apolipoprotein AV; lipoprotein lipase
14.  Glycosylphosphatidylinositol-anchored high density lipoprotein–binding protein 1 plays a critical role in the lipolytic processing of chylomicrons 
Cell metabolism  2007;5(4):279-291.
Summary
The triglycerides in chylomicrons are hydrolyzed by lipoprotein lipase (LpL) along the luminal surface of the capillaries. However, the endothelial cell molecule that facilitates chylomicron processing by LpL has not yet been defined. Here, we show that glycosylphosphatidylinositol-anchored high density lipoprotein–binding protein 1 (GPIHBP1) plays a critical role in the lipolytic processing of chylomicrons. Gpihbp1-deficient mice exhibit a striking accumulation of chylomicrons in the plasma, even on a low-fat diet, resulting in milky plasma and plasma triglyceride levels as high as 5,000 mg/dl. Normally, Gpihbp1 is expressed highly in heart and adipose tissue, the same tissues that express high levels of LpL. In these tissues, GPIHBP1 is located on the luminal face of the capillary endothelium. Expression of GPIHBP1 in cultured cells confers the ability to bind both LpL and chylomicrons. These studies strongly suggest that GPIHBP1 is an important platform for the LpL-mediated processing of chylomicrons in capillaries.
doi:10.1016/j.cmet.2007.02.002
PMCID: PMC1913910  PMID: 17403372
15.  Liver heparan sulfate proteoglycans mediate clearance of triglyceride-rich lipoproteins independently of LDL receptor family members 
Journal of Clinical Investigation  2007;117(1):153-164.
We examined the role of hepatic heparan sulfate in triglyceride-rich lipoprotein metabolism by inactivating the biosynthetic gene GlcNAc N-deacetylase/N-sulfotransferase 1 (Ndst1) in hepatocytes using the Cre-loxP system, which resulted in an approximately 50% reduction in sulfation of liver heparan sulfate. Mice were viable and healthy, but they accumulated triglyceride-rich lipoprotein particles containing apoB-100, apoB-48, apoE, and apoCI-IV. Compounding the mutation with LDL receptor deficiency caused enhanced accumulation of both cholesterol- and triglyceride-rich particles compared with mice lacking only LDL receptors, suggesting that heparan sulfate participates in the clearance of cholesterol-rich lipoproteins as well. Mutant mice synthesized VLDL normally but showed reduced plasma clearance of human VLDL and a corresponding reduction in hepatic VLDL uptake. Retinyl ester excursion studies revealed that clearance of intestinally derived lipoproteins also depended on hepatocyte heparan sulfate. These findings show that under normal physiological conditions, hepatic heparan sulfate proteoglycans play a crucial role in the clearance of both intestinally derived and hepatic lipoprotein particles.
doi:10.1172/JCI29154
PMCID: PMC1716206  PMID: 17200715
16.  Effects of the PPARγ agonist pioglitazone on lipoprotein metabolism in patients with type 2 diabetes mellitus 
Journal of Clinical Investigation  2005;115(5):1323-1332.
Elevated plasma levels of VLDL triglycerides (TGs) are characteristic of patients with type 2 diabetes mellitus (T2DM) and are associated with increased production rates (PRs) of VLDL TGs and apoB. Lipoprotein lipase–mediated (LPL-mediated) lipolysis of VLDL TGs may also be reduced in T2DM if the level of LPL is decreased and/or the level of plasma apoC-III, an inhibitor of LPL-mediated lipolysis, is increased. We studied the effects of pioglitazone (Pio), a PPARγ agonist that improves insulin sensitivity, on lipoprotein metabolism in patients with T2DM. Pio treatment reduced TG levels by increasing the fractional clearance rate (FCR) of VLDL TGs from the circulation, without changing direct removal of VLDL particles. This indicated increased lipolysis of VLDL TGs during Pio treatment, a mechanism supported by our finding of increased plasma LPL mass and decreased levels of plasma apoC-III. Lower apoC-III levels were due to reduced apoC-III PRs. We saw no effects of Pio on the PR of either VLDL TG or VLDL apoB. Thus, Pio, a PPARγ agonist, reduced VLDL TG levels by increasing LPL mass and inhibiting apoC-III PR. These 2 changes were associated with an increased FCR of VLDL TGs, almost certainly due to increased LPL-mediated lipolysis.
doi:10.1172/JCI200523219
PMCID: PMC1070635  PMID: 15841215
17.  Lipoprotein lipase (LpL) on the surface of cardiomyocytes increases lipid uptake and produces a cardiomyopathy 
Journal of Clinical Investigation  2003;111(3):419-426.
Lipoprotein lipase is the principal enzyme that hydrolyzes circulating triglycerides and liberates free fatty acids that can be used as energy by cardiac muscle. Although lipoprotein lipase is expressed by and is found on the surface of cardiomyocytes, its transfer to the luminal surface of endothelial cells is thought to be required for lipoprotein lipase actions. To study whether nontransferable lipoprotein lipase has physiological actions, we placed an α-myosin heavy-chain promoter upstream of a human lipoprotein lipase minigene construct with a glycosylphosphatidylinositol anchoring sequence on the carboxyl terminal region. Hearts of transgenic mice expressed the altered lipoprotein lipase, and the protein localized to the surface of cardiomyocytes. Hearts, but not postheparin plasma, of these mice contained human lipoprotein lipase activity. More lipid accumulated in hearts expressing the transgene; the myocytes were enlarged and exhibited abnormal architecture. Hearts of transgenic mice were dilated, and left ventricular systolic function was impaired. Thus, lipoprotein lipase expressed on the surface of cardiomyocytes can increase lipid uptake and produce cardiomyopathy.
doi:10.1172/JCI200316751
PMCID: PMC151861  PMID: 12569168
18.  Transgenic rabbits expressing human lipoprotein lipase 
Cytotechnology  2000;33(1-3):93-99.
To study the functions of lipoprotein lipase (LPL) in lipid and lipoprotein metabolism and the relationship between LPL and atherosclerosis, we generated transgenic rabbits expressing the human LPL gene. A total of 4045 Japanese whiterabbit embryos were microinjected with a 3.8-kb SalI/HindIII fragment containing the chicken β-actin promoter, human LPL cDNA and rabbit β-globin with poly (A) signals, and then transplanted into 116 recipient rabbits. Of the 166 pups born, six pups were transgenic as confirmed by Southern blot analysis. ANorthern blot analysis revealed that human LPL was expressed by a number of tissues including the heart, kidney, adrenal gland and intestine. One transgenic rabbit showed up to 3-foldincreased LPL activity in post-heparin plasma compared to thatin nontransgenic rabbits. Human LPL expression in various tissues of transgenic rabbits was further elucidated by in situ hybridization and immunostaining. Since rabbits are superior to mice as a model of atherosclerosis, this transgenicrabbit model should provide a valuable tool for the study of LPL in lipid metabolism and atherosclerosis.
doi:10.1023/A:1008115429679
PMCID: PMC3466728  PMID: 19002816
atherosclerosis; lipoprotein; lipoprotein lipase; transgenic rabbit
19.  Heparin-binding defective lipoprotein lipase is unstable and causes abnormalities in lipid delivery to tissues 
Journal of Clinical Investigation  2001;107(9):1183-1192.
Lipoprotein lipase (LpL) binding to heparan sulfate proteoglycans (HSPGs) is hypothesized to stabilize the enzyme, localize LpL in specific capillary beds, and route lipoprotein lipids to the underlying tissues. To test these hypotheses in vivo, we created mice expressing a human LpL minigene (hLpLHBM) carrying a mutated heparin-binding site. Three basic amino acids in the carboxyl terminal region of LpL were mutated, yielding an active enzyme with reduced heparin binding. Mice expressing hLpLHBM accumulated inactive human LpL (hLpL) protein in preheparin blood. hLpLHBM rapidly lost activity during a 37°C incubation, confirming a requirement for heparin binding to stabilize LpL. Nevertheless, expression of hLpLHBM prevented the neonatal demise of LpL knockout mice. On the LpL-deficient background hLpLHBM expression led to defective targeting of lipids to tissues. Compared with mice expressing native hLpL in the muscle, hLpLHBM transgenic mice had increased postprandial FFAs, decreased lipid uptake in muscle tissue, and increased lipid uptake in kidneys. Thus, heparin association is required for LpL stability and normal physiologic functions. These experiments confirm in vivo that association with HSPGs can provide a means to maintain proteins in their stable conformations and to anchor them at sites where their activity is required.
PMCID: PMC209279  PMID: 11342582

Results 1-19 (19)