COOH-terminal Disruption of Lipoprotein Lipase in Mice Is Lethal in Homozygotes, but Heterozygotes Have Elevated Triglycerides and Impaired Enzyme Activity*

The role of the enzyme lipoprotein lipase (LPL) in atherosclerosis is uncertain. To generate an animal model of LPL deficiency, we targeted the LPL gene in embryonic stem cells with a vector designed to disrupt the COOH terminus of the protein and used these cells to generate LPL-deficient mice. Germ line transmission of the disrupted LPL allele was achieved with two chi meric males, and offspring from each of these animals were phenotypically identical. Pups homozygous (-/- ) for LPL deficiency died within 48 h of birth with ex treme elevations of serum triglycerides (13,327 mg/dl) associated with essentially absent LPL enzyme activity in heart and carcass. Newborn heterozygous (+/-) LPL deficient pups had lower LPL enzyme activity and higher triglycerides (370 versus 121 mg/dl) than wild type (+/+) littermates. Adult heterozygotes had higher triglycerides than wild type mice with ad libitum feed ing (236 mg/dl for +/- versus 88 mg/dl for +/+) and after fasting for 4 h (98 mg/dl for +/- versus 51 for

scure. Elevated triglycerides may cause vascular damage through both direct and indirect mechanisms. Remnants of triglyceride-rich lipoproteins may be directly atherogenic (2), a hypothesis bolstered by the recent demonstration of triglyceride-rich lipoproteins in human atherosclerotic lesions (3). Elevated triglycerides might affect vascular health indirectly by decreasing HDU cholesterol levels, by rendering LDL particles more atherogenic, or by affecting clotting.
Defects in lipoprotein lipase (LPL) can cause hypertriglyceridemia. LPL is increasingly recognized as a multifunctional protein. In addition to hydrolyzing triglycerides in VLDL and chylomicrons, the LPL protein may also function as an apolipoprotein, associating with the surface of various lipoproteins to promote binding to the low density lipoprotein (LDL) receptor-related protein/nj-macroglobulin receptor (4), LDL receptor (5,6), and extracellular proteoglycans (7,8). The LPL protein can be functionally divided into two major domains: an NH 2terminal domain containing the catalytically active site, and a COOH-terminal domain (9). The latter is essential for catalytic activity (10), binds lipoproteins (11), and is probably responsible for the LPL-mediated catabolism of triglyceride-rich lipoproteins by the LDL receptor-related protein/oj-macroglobulin receptor (12).
LPL could also impact atherogenesis independent of effects on circulating triglycerides. Macrophages from human and rabbit atherosclerotic lesions express LPL (13)(14)(15), and higher levels of macrophage LPL expression are found in inbred strains of mice that are susceptible to atherosclerosis (16), suggesting that local expression of LPL could promote uptake of lipoproteins by vascular tissue (17). Thus genetic defects in LPL could have opposing effects on atherosclerotic risk; systemically decreased function could increase the concentration of atherogenic, triglyceride-rich particles, but decreased macrophage expression could decrease foam cell formation.
Human heterozygous LPL deficiency is probably common. It has been postulated to form a subset of familial combined hyperlipidemia (18), a very common genetic disorder associated with vascular disease. However, heterozygous LPL deficiency could have phenotypically different effects depending on genetic and physiological backgrounds. This might explain why some families with heterozygous LPL deficiency manifest familial hypertriglyceridemia (19), while others have lipid abnormalities associated with increased atherosclerotic risk (20). Further complicating study of human heterozygous LPL defi-FIG. 1. Strategy for inactivation of the mouse LPL gene. Schematic diagrams of the targeting vector, native (wild type) mouse gene, a successfully targeted allele, and the predicted restriction fragments resulting from digestion of genomic DNA with HindIII and Tthl11I are shown. S represents SacI, TK represents thymidine kinase (for negative selection in the presence of ganciclovir), and NEG represents the neomycin resistance cassette (for positive selection with G418). Also shown is the location of the LPL genomic probe (spanning the intron 5/exon 6 junction) used for detection of fragments by Southern blotting. ciency is the fact that detection of this disorder is difficult given the large number of mutations in the LPL gene (21).
With a goal of generating an animal model of LPL deficiency within a homogeneous genetic background suitable for studying atherosclerosis, we have inactivated the LPL gene in mice by homologous recombination using a targeting vector designed to disrupt the COOH-terminal domain of the LPL protein.

EXPERIMENTAL PROCEDURES
Vector Construction and Generation of LPL-deficient Mice-An 8.3-kb SacUSau3AI genomic fragment was isolated from a AEMBL3 library constructed using Balb/c DNA (22). Restriction mapping, Southern blotting (with detection by oligonucleotides based on the mouse LPL eDNA sequence in Ref. 23), and sequencing showed that this fragment contained exons 7-10 of the mouse gene with essentially the same structure described by Zechner et al. (24). A 1.8-kb EcoRUHindlII fragment of the plasmid pRJ-l containing the neomycin resistance cassette driven by the PGK promoter was inserted by blunt-end ligation at a HindlII site in exon 8. This manipulation was shown to abolish the HindlII site and interrupt exon 8, resulting in a predicted mouse LPL protein disrupted following the leucine residue at position 380 as numbered in Ref. 23. This modified genomic clone was then inserted at the XhoI site of a Bluescript plasmid (a gift from Fred Fiedorek, Chapel Hill, NC) containing the 3.4-kb thymidine kinase fragment from pHSV-106 to generate the targeting vector.
E14 ES cell electroporation, positive (using G418) and negative (using ganciclovir) selection, and injections into the blastocysts of C57BL/6J embryos were carried out as described (25). Chimeric (on the basis of coat color) males were mated with C57BL/6J females to generate Fl animals, and these Fl animals were crossed with each other to generate the mice characterized in this study.
Animal Genotyping-DNA isolated from tail (for most animals) or liver (from recently expired animals) was subjected to Southern blotting or PCR. For Southern blotting, 5-15 f.Lg of DNA was electrophoresed followed by acid depurination, treatment with NaOH, and transfer using standard techniques. Blots were probed with a random-primed 0.8-kb BamHUPvulI fragment of the mouse LPL gene spanning the 3' end of intron 5 and the 5' end of exon 6. As expected, preliminary blots showed that this probe did not hybridize with the targeting vector. For PCR genotyping of potential heterozygotes, an upstream primer corresponding to mouse exon 8 (5'-TTT ACA CGG AGG TGG ACA TCG GA) and a downstream primer corresponding to a region near the 3' end of the neomycin resistance cassette (5'-TCG CCT TCT ATC GCC TTC TTG AC) were used in reactions containing genomic DNA and 2 mM MgCl 2 subjected to the following cycling parameters: 5 min at 94°C for 1 cycle, and 1 min at 94°C/2 min at 55°C/3 min at 72°C for 30 cycles.
Mouse Housing, Handling, and Diet-Animal rooms were illuminated between 7:00 a.m. and 7:00 p.m. Mice were fed a 50/50 mixture of PicoLab rodent chow 20 and mouse chow 20 (product numbers 5053 and 5058) with a total fat content of -6.75%. Animals were weaned at 21 days of age. For blood collection via the retro-orbital plexus, animals were lightly anesthetized with methoxyflurane. Deep anesthesia was used for those animals subjected to exsanguination via inferior vena cava venipuncture.
Lipid and Lipoprotein Analysis-Triglycerides, cholesterol, and phospholipids were assayed enzymatically using commercially available kits (Wako Pure Chemical Industries, Ltd., Osaka, Japan). For measurements of total lipids, serum was kept at 4°C or on ice and assayed within 2 h for triglycerides and phospholipids to decrease the chances of endogenous hydrolysis (26). Cholesterol determinations were generally done the following day after storing samples at -70 "C. HDL cholesterol was measured after polyethylene glycol precipitation of apo B-containing lipoproteins (27). For separation of lipoproteins by gel filtration, serum samples from two animals with similar total lipids were pooled and separated by FPLC using two Superose 6 columns in series (28). The 50 fractions collected per condition were analyzed the same day for triglycerides and the following day for phospholipids and cholesterol.
Determination of LPL Enzyme Activity, mRNA, and Protein-LPL activity was assayed as the salt-inhibitable ability of triplicate samples to hydrolyze an emulsion containing radiolabeled triolein (29). For postheparin plasma activity, animals were injected intraperitoneally with 200 units of porcine heparin; 30 min later blood was collected from the inferior vena cava. Previous studies have shown this procedure to be suitable in mice (30). For other assays, individual tissues were flash frozen in liquid nitrogen, weighed, and made 4% (w/v) in assay buffer (29). For newborn pups, the heart was removed and processed, the milk-filled stomach was removed and discarded (to prevent dilution of radioactive emulsion by milk lipids), and the remaining carcass (minus heart and stomach) was frozen, weighed, and homogenized in assay buffer. For determination of LPL message, total RNA was prepared from tissues by centrifugation in cesium chloride and mRNA was detected by Northern blotting as described previously (31). For determination of LPL protein, tissue extracts were subjected to SDS-polyacrylamide gel electrophoresis and Western blotted, and LPL protein was detected with chicken anti-bovine milk LPL as described (29). Each gel included one lane loaded with purified bovine LPL as a positive control.
Statistical Information-Differences were assessed using unpaired, two-tailed t tests unless otherwise specified. For the data shown in Fig.  4, ANOVA was specifically not used because groups contained different numbers of animals.

RESULTS
The strategy for inactivating the mouse LPL gene is shown in Fig. 1. The mouse LPL gene has 10 exons, 9 of which are translated. The targeting vector contains~3 kb of mouse DNA upstream and~5 kb downstream of the exon 8 HindIII site where the neomycin resistance cassette (NEO) was inserted. Predicted restriction fragments detected by the LPL probe spanning the intron 5/exon 6 junction are shown at the bottom of the figure for native and targeted genes. With HindIII, the native fragment is 8.4 kb but the targeted fragment is 14.4 kb since insertion of NEO abolishes the natural HindIII site in Additional confirmation of authe ntic targe ti ng is shown in panel C. Tail DNA from animals shown to be +/+ or +/-by Southern blotting was subjected to PCR using an up stream primer just 5' to the NEO in sertion site in exon 8 and a downst rea m pri mer complementary to NEO . The expecte d -600-bp band was amplified from heterozygotes (+/-, lanes 2 and 3) but not wild type a ni mals (+ /+ , lan es 1 an d 4). Thi s as say was subsequently used for rapid genotyping. In oth er studies, confirmation of the homozygous state was also performed by PCR using the primers of panel C in conjun ction with primers (sepa rate d by -127 bp) flanking the NEO insertio n site in exon 8. With the latter primer s, wild type anima ls showed a 127-bp band, homozygotes had a -1.8-kb band (due to the presence of the NEO cassette between th e primer s ), and het er ozygotes had both bands (data not shown).
Homozygote s were born via ble, initi ally a ppea red healthy, but un iforml y died withi n 48 h . The 100% mortali ty for t his geno type was sign ifican tly higher t han dea th rate s for +/and + / + animals ( Table I). Autop sy find ings from four homozygotes (data not shown ) were non specific, although atelectasis a nd congestion in t he lungs and hepatic congestion were see n consis te ntly . There were no pancreatic abno rm ali ti es det ected.
Also shown in Tabl e I are seru m lipid s from mice -12 h after birth. Homozygotes had marked elevations of t riglycerides , phospholipids, and total choleste rol, bu t no detectable HDL cholesterol. Thi s extre me hyperlipidemia was feeding-dependen t . One homozygote was kept from nursing (verified by th e absence of milk in the stoma ch), and in this single animal at the time of sacrifice lipids wer e: triglycerides, 296 mg/dl; cholesterol, 95 mg/dl; phospholipids, 178 mg/dl. Triglycerides (lanes 1, 5, a n d 9 ) and correctly target ed ES cell clon es (lanes 2·4 , 6-8 , a nd 10) was dig ested with HindIII or Tthl11l , Southern blotted , and det ected with the LPL prob e indica ted sche matica lly in Fi g. 1 (lanes 1-4, 9, a nd 10) or a NEO probe (lanes [5][6][7][8], consisting ofa BamHIIPvuII fragm en t complementary to th e ph osphotransfer ase codin g region from t he vecto r pKJ-l. Correctly ta rget ed H indIII-cu t DNA contained th e pr ed icted 14.4-kb mutant band , wh ich hyb rid ized wit h both LPL a nd NEO pr obes. Correctly targeted Tth 11lI-cut DNA containe d the pr edicted 7.7-kb mu t an t ba nd (lane 10). Panel B shows DNA from wild typ e (+/+), hete rozygou s (+/-), a nd homozygous (-/-) mice a fter HindIII or Tth11 11 dig estion. As expected, heterozygotes have bot h m utant a nd native alleles after digestion with bot h enzymes wh ile homozygotes ha ve on ly t he m uta nt a llele (14.4 kb for HindIII , 7.7 kb for Tth 11lI). Panel C sho ws a PCR assay using the same DNA from t he +/mice in pan el B. Assays wer e perform ed as descri bed under "Experi mental Procedures" using a n up st ream primer complementary to exon 8 of the mouse LPL ge ne and a downst ream pri mer compleme ntary to NEO. As expected, t he pred icted -600·bp band was seen in DNA fro m + /mi ce a nd wit h th e targe ti ng vector (la beled Plasmid ) but not in +/+ mice or in the nega tive contro l lane (labeled No DNA ). type animals at 12 h. HDL cholesterol levels were not decreased in +1-animals; in fact, HDL cholesterol was almost significantly higher in +/compared to +/+ pups (p = 0.0730).
As expected, LPL enzyme activity was decreased in +/and essentially absent in -/-pups (Fig. 3). For these experiments, the heart was removed and assayed, then the carcass minus the heart (and milk-filled stomach, as described under "Experimental Procedures") was assayed separately.
Heterozygotes developed normally. Over 100 animals were weighed at weekly intervals for the first two months oflife, and although +/animals tended to be slightly heavier than +/+ littermates, this difference was not significant (p = 0.5513).
Heterozygotes had higher triglycerides than wild type animals with fasting and with ad libitum feeding ( Fig. 4 and Table II). With fasting, triglycerides were~2-fold elevated in +/versus +/+ animals regardless of sex. With ad libitum feeding, the genotype-specific differences in triglycerides were amplified with the highest elevations seen in heterozygous LPL-deficient females. There were no significant differences between groups for total cholesterol or phospholipids.
As with neonates, HDL cholesterol levels were not lower in adult +/-animals. Ad libitum fed adults with heterozygous LPL deficiency tended to have higher HDL cholesterols than their +/+ littermates (118 ± 14 for +/versus 100 ::' ::: 9 for +/+, Lipoprotein analysis by gel filtration chromatography showed that the increased triglycerides in heterozygous LPLdeficient animals were due to an increase in VLDUchylomi- DISCUSSION We present evidence consistent with the inactivation of the LPL gene in mice. Targeting of the LPL gene with a vector designed to disrupt the COOH terminus of the LPL protein results in severe hypertriglyceridemia and essentially absent enzyme activity in homozygotes. These animals die within 2 days of birth. Heterozygotes have decreased enzyme activity and mild to moderate hypertriglyceridemia with both feeding and fasting. The homozygous and heterozygous states appear to have different effects on HDL levels. Homozygous LPL deficiency in humans causes a distinctive phenotype, the chylomicronemia syndrome, that is not necessarily lethal (33). Naturally occurring defects in LPL activity have also been described in cats (34), mice lacking mast cells (W/ WV mice; Ref. 35), and mice homozygous for combined lipase deficiency (cld / cld mice; Ref. 36). Our homozygotes resemble cld / cld mice. Triglycerides are similar at 12 h postpartum (~1O,OOO mg/dl for cld/cld, -13,000 mg/dl for LPL -/-), and animals uniformly die within 48 h of birth. cld / cld mice presumably die because of microinfarcts in critical organs caused by dense packing of chylomicrons in capillaries (37), and a similar mechanism probably operates in LPL -/-mice. The crons (Fig. 5, top panels, fractions 1-10). Consistent with measurements of HDL cholesterol by PEG precipitation, HDL cholesterol measured by gel filtration tended to be higher in +/animals (middle panels, fractions 30--38). In males, "shoulders" to the right of the VLDL triglyceride peak (fractions [10][11][12][13][14][15] and to the left of the HDL cholesterol peak (fractions 20--30) were consistently more prominent in heterozygotes.
LPL enzyme activity was decreased in animals carrying one copy of the mutant allele (Fig. 6). Activity in plasma 30 min after the intraperitoneal injection of heparin (panel A) was 43% lower in heterozygotes (146 ::' ::: 25 /-Lmol of FFAImllh for +/versus 258 ::' ::: 19 for +/+, n = 4 animals per genotype, p = 0.0121). This decrease was also reflected in enzyme activity assayed in individual tissues (panel BJ. For heart, kidney, epididymal/parametrial and inguinal adipose tissue, lung, psoas muscle, and brain, LPL activity was consistently decreased in +/versus +/+ animals. These differences were statistically significant only for heart and kidney (both p < 0.05) due to considerable animal-to-animal variation, especially for adipose tissue. Since these experiments were performed using the F2 generation representing a mixture of the the 129/Ola and C57BU6J strains, the genetic heterogeneity of these animals is probably a major source of the observed variation in LPL enzyme activity.
There were tissue-specific differences in the magnitude of the decrease in enzyme activity associated with the mutant allele. For example, in multiple assays involving six different side-by-side comparisons of age and weight-matched +/+ versus +/animals, the decrease in heart activity was consistently less than 50% (mean decrease 28.4%) while the decrease in renal activity was consistently greater than 50% (mean decrease 62.4%).
LPL message and protein were decreased in heterozygotes (Fig. 7). Both the 3.6-and 3.4-kb LPL mRNA species detected by others in mouse tissues (23) were decreased in +1-(lane 2) compared to +/+ (lane 1) adipose tissue. Similar differences were seen in comparisons of adipose tissue total RNA from five different pairs of +/+ and +/animals. A decrease in LPL mRNA for heterozygotes was also seen in heart (not shown). Blood was obtained by retro-orbital bleeding from mice at age 7-9 weeks between 10:00 a.m. and noon. Serum was assayed immediately or stored at -70 DC for assay within 24 h. Data from the left side of the figure represent mice fasted for 12 h, and data from the right side represent ad libitum fed mice. +/+ indicates wild type, and +/indicates heterozygotes as determined by PCR genotyping of tail DNA. Data represent means :' :: S.E. for 3-8 animals per group.

TABLE II
Serum triglycerides in adult heterozygous LPL-de{icient and wild type mice Data are presented as mean:':: S.E. in mg/dl. LPL +/-mice suggest that, with the exception of the LPL -/state, LPL activity in mice has a limited role in determining HDL cholesterol levels.
That heterozygous LPL-deficient mice have increased triglycerides seems sensible, but this result was not predictable. In humans, 94% of young heterozygotes have normal triglycerides (19) as do mice heterozygous for the cld mutation (36). The mechanism responsible for hypertriglyceridemia in our mice, especially with fasting, is uncertain. Mutations generally cause gain of function, loss of function, or have dominant negative effects. The latter class of mutation could be relevant to our mice since active LPL is probably a dimer and one could envision an inactive but stable monomer having a prolonged dominant negative effect on triglyceride metabolism. This mecha-cld mutation affects the posttranslational processing of both LPL and hepatic lipase (38), but the similarities between LPL -/-and cld/cld mice suggest that LPL is the lipase most critical for neonatal survival.
HDL cholesterol was absent from the serum of LPL -/mice, an expected finding since the major sources of lipid for HDL generation are thought to be remnant cholesterol and phospholipid from chylomicronIVLDL metabolism. Whether apolipoprotein A-I, the major protein ofHDL, is produced normally in LPL -/-animals is unknown. These mice could be used to determine if nascent HDL-like particles are present in the absence of LPL activity.
In human populations, lower levels of LPL enzyme activity are associated with lower levels ofHDL cholesterol (39). Given this association, one might predict that the lower (but not absent) LPL activity ofLPL +/-mice would also be associated with lower HDL cholesterol. This was not observed. There was no difference in HDL cholesterol between +/and +/+ adult mice, and levels were almost significantly higher in heterozygous neonates (Table I). At least three groups have overexpressed human LPL in transgenic mice (40)(41)(42), and all report decreases in triglyceride-rich lipoproteins without significant effects on HDL cholesterol. These results and our findings in   1 an d 2 ), an d adipose tissue protein was subjected to Western blotting (lan es 3 an d 4). For la nes 1 an d 2, 20 fJ.g of tota l RNA was separated on agarose gels containi ng formaldehyde. Equa l loadin g and equa l RNA integri ty for each genotype was verifie d by analysis of ethidiu m-stained 28 and 18 S ribosomal RNA int en sities before tran sfer . Th e LPL message (indicated by th e 3.6-kb a nd 3.4-kb ma rkers) was detected using a ran dom-primed mouse LPL eDNA. For lanes 3 and 4 , 1 fJ.g of protein was sepa rated by SDS-polyacry lami de gel electrophoresis wit h the -55-kDa LPL prot ein (visible in both lan es at the position indica ted by the arrowhead to the righ t of lane 4 ) detected usin g an iodinated chicken a nt i-bovine LPL a nt ibody. N umb ers to the left of lan e 3 ind icat e the positio ns of protein size sta ndards. Resu lt s shown for LPL mRNA are representati ve of comparisons of 5 different pairs of + /+ and +/ani ma ls . Resul ts for LPL protein a re representative of comparisons of 3 differen t pair s of +/+ an d + / -ani mals . 1 2 a po CI prob ably oper ates through a si mila r mechanism to increase triglycerides (48). Over expression of a po CII , a cofactor for LPL activity, surpris ingly a lso incre ases triglycerides , perhap s by inte rfering with th e ability of VLDL to bind glycosaminoglycan s (49). Each of th ese models was gene ra ted using human t ran sgen es in mice. Hetero zygous LPL-d eficient mice have the theoretical a dva ntage of man ifesting hyp ertriglycer idemia that is not depend ent on th e expression of a pr otein from a nothe r species . Anoth er potential a dvantage of our model is t he lack of a n effect on HDL choles te rol levels. In species with cholesteryl es te r tran sfer pr otein activity, triglyceride concentrations are in versely related to HDL choles te rol levels, making it difficult to decide whethe r ben eficial effects on vascul ar disea se a re du e to decr eased triglycerides or incre ase d HDL-C. LPL +/mice sui tably backcrossed into th e C57BU6J background could be useful for determining how hypertriglycerid emi a a lone affect s at he ros cleros is. In addition, assessing athero sclerotic disea se in LPL +/mice in the setting of diab et es mellitus, ethanol intak e, and hypoth yroidism , or a fter crossing th ese animals with cholesteryl es te r trans fer protein tran sgenic mice, with heter ozygotes for a po E deficiency , or with heterozygote s for LDL receptor deficien cy, could pr ovide ins ight into whether commonly observed human lipid ph enotyp es confer atherosclerotic risk. ism does not a ppea r to be operative in our +/mice. LPL activity (Fig. 6), message, a nd pr otein (Fig. 7) are lower in th e tissu es of heterozygotes, making it lik ely th at our mutation causes loss of functi on t hro ugh a decr ease in LPL protein .
Cons is te nt with this hyp oth esis, COOH-ter minal truncation of human LPL at residue 38 1, essentially the sa me site disrupted in mouse LPL in th e current study, cau ses a loss of catalytic activity as well as a decrease in pro tein mass in transien t tran sfection st udies (43). Thus, t he ph enotype of mice gene rated by COOH-te r mina l disruption of LPL may be simila r to th e ph en otyp e gene ra te d by targeting regions of th e LPL gene 5' to th e exon 8 site chosen in this st udy.
Why th en do LPL + /mice have elevated tri glycerides in the fasted state on a low fat diet? At least four expla na tions are possible. First , the disruption of t he COOH terminus of th e LPL molecul e could some how impa ir th e metabolism oftriglyceride-ri ch lipoprotein s as sugges te d by cell culture st udies (12). We do not find evidence of a protein with altered size in + /ti ssu es to su pport this hypoth esis, bu t Western blots were don e with a single antibody, rai sing th e possibility th at a n a lte re d pr otein is pr oduced bu t not recogn ized by our an tib ody. Second, LP L deficiency could a ffect triglyceride-rich particle composition a nd impair clearanc e. Third, LPL deficiency could reduc e hepa tic reuptak e of apo B-containing particles with a net effect of lipoprotein over production by the liver (44). Our enzyme assay was not sufficiently se ns itive to detect LPL activity in mou se liver, although we have been able to det ect very low levels of LPL pr otein in th is tissue by Western blotting (not shown). Fourth, LPL deficiency is like ly to increase th e concentration of remnan t particles in these a nima ls, a possibility suggeste d by t he "shoulders " of particl es seen to the right of the triglyceride peak a nd to the left of th e cholesterol peak for males in Fig. 5. Remnants are noncomp etiti ve inhibitors ofLPL activity, a nd even low concent rations of t hese particles could subs ta ntially interfere with VLDL hydrolysis (45).
There are oth er mou se model s of elevate d triglycerides. Overexpression of apo CII I in mice results in hypertriglyceridemia (46 ). Thi s condition is corrected by over expression of apo E (47), suppor ti ng th e hypoth esis that excess ap o CII I displaces a po E from triglyceride-ri ch lip opr oteins, thereby hindering clearance by a po E-depend en t mechanism s. Overexpression of