Liver disease with altered bile acid transport in Niemann-Pick C mice on a high-fat, 1% cholesterol diet
Robert P. Erickson,1
Achyut Bhattacharyya,2
Robert J. Hunter,1
Randall A. Heidenreich,1 and
Nathan J. Cherrington3
Departments of 1Pediatrics, 2Pathology, and 3Pharmacology/Toxicology, University of Arizona, School of Health Sciences, Tucson, Arizona
Submitted 23 December 2004
; accepted in final form 23 March 2005
 |
ABSTRACT
|
---|
Cholestatic hepatitis is frequently found in Niemann-Pick C (NPC) disease. We studied the influence of diet and the low density lipoprotein receptor (LDLR, Ldlr in mice, known to be the source of most of the stored cholesterol) on liver disease in the mouse model of NPC. Npc1/ mice of both sexes, with or without the Ldlr knockout, were fed a 18% fat, 1% cholesterol ("high-fat") diet and were evaluated by chemical and histological methods. Bile acid transporters [multidrug resistance protein (Mrps) 15; Ntcp, Bsep, and OatP1, 2, and 4] were quantitated by real-time RT-PCR. Many mice died prematurely (within 6 wk) with hepatomegaly. Histopathology showed an increase in macrophage and hepatocyte lipids independent of Ldlr genotype. Non-zone-dependent diffuse fibrosis was found in the surviving mice. Serum alanine aminotransferase was elevated in Npc1/ mice on the regular diet and frequently became markedly elevated with the high-fat diet. Serum cholesterol was increased in the controls but not the Npc1/ mice on the high-fat diet; it was massively increased in the Ldlr/ mice. Esterified cholesterol was greatly increased by the high-fat diet, independent of Ldlr genotype.
-Glutamyltransferase was also elevated in Npc1/ mice, more so on the high-fat diet. Mrps 15 were elevated in Npc1/ liver and became more elevated with the high-fat diet; Ntcp, Bsep, and OatP2 were elevated in Npc1/ liver and were suppressed by the high-fat diet. In conclusion, Npc1/ mice on a high-fat diet provide an animal model of NPC cholestatic hepatitis and indicate a role for altered bile acid transport in its pathogenesis.
Niemann-Pick C; low density lipoprotein receptor knockout; neonatal cholestasis; bile acid transporters; non-zone-dependent fibrosis
NIEMANN-PICK C DISEASE (NPC) is a neurodegenerative disorder of childhood associated with a disruption of intracellular cholesterol trafficking (25). Although a neurological presentation is most frequently diagnosed, some cases may be missed because of death from cholestatic liver disease before development of neurological disease (15). In fact, one study concluded that NPC is the second most common genetic cause of liver disease in infancy in the United Kingdom (17). A recent survey found that NPC explained 27% of idiopathic neonatal cholestasis and 8% of all infants evaluated for cholestasis (32). However, unlike neonatal hepatic steatosis (46, 10), this liver disease has not been the subject of intensive investigation. We have taken advantage of a mouse model of NPC1 disease, due to a natural mutation in the Npc1 gene, to further elucidate the pathological and biochemical abnormalities of the liver in this disorder. We have found that a high-fat diet hastens the death of about two-thirds of the Npc1/ mice, with some influence of sex, and induces a number of features of cholestatic hepatitis. Marked changes in bile acid transporters were noted. Because cholesterol is taken up in liver by the low density lipoprotein receptor (Ldlr), we have also taken advantage of the Ldlr knockout to look for genetic interactions between this major component of cholesterol metabolism and liver disease in Npc1/ mice.
 |
MATERIALS AND METHODS
|
---|
Animals.
Npc1NIH mutant mice from the BALB/cJ background were maintained by brother-sister mating of heterozygous animals. Animals were maintained at the University of Arizona Animal Care Facility according to the criteria outlined in the Guide for the Care and Use of Laboratory Animals (National Institutes of Health Assurance No. A-324801); they were given mouse chow containing 6% fat (or 10% for breeding mothers) and water ad libitum. At weaning (
21 days of age), tail tips were removed from mice, and DNA was prepared. PCRs to identify genotypes at the Npc1NIH locus were performed using the primer pairs as described (16). The PCR conditions used 10 mmol/l Tris, pH 8.3, 50 mmol/l KCl, 2.5 mmol/l Mg2+, 200 µmol/l dNTPs, 1.25 units Taq polymerase, and 1 µmol of each primer. DNA (2040 ng) was added at 25°C, and cycles of 30 s at 95°C, 30 s at 61°C, 1 min at 72°C for 35 s, and 10 min at 72°C were used. The products were separated on 1.2% NuSieve agarose gels.
Special diet.
The high-fat diet was no. 112280 from Dyets (Bethlehem, PA). It contains 18% butter fat and 2% corn oil with 1% additional cholesterol. Over 50 Npc1/ mice of various Ldlr genotypes were placed on this diet at weaning (21 days). Npc1+/+ and Npc1+/ sibling mice (83% of the weaned mice, since Npc1/ are found at a significantly lower fraction than 25%) were used as controls.
LDL receptor knockout mice.
Ldlr/ mice on the C57BL6/J background were the knockouts developed by Ishibashi and colleagues (14) and were obtained from Jackson Laboratory (Bar Harbor, ME). They were interbred with the Npc1NIH heterozygotes (homozygous recessives are sterile) on the BALB/cJ background to produce double heterozygotes (Ldlr+/, Npc1+/; F1, C57BL/6J, BALB/cJ). These mice were interbred to produce the nine possible genotypes of Npc1 and Ldlr (Npc1+/+, Ldlr+/+; Npcl+/+, Ldlr+/; Npc1+/+, Ldlr/; Npc1+/, Ldlr+/+; Npc1+/,Ldlr+/; Npc1+/, Ld1r/; Npc1/, Ld1r+/+; Npc1/, Ld1r+/; and Npc1/, Ld1r/), which were on an F2, C57BL6/J, BALB/cJ background. The last three genotypes were experimental, whereas the third and fifth genotypes were the major controls. Ldrl typing was performed after the method of Gaw and colleagues (11); however, we could not use both PCR primers successfully in the same reaction. Thus the typing for neo (for the presence of the knockout allele) was performed as described, whereas the Ldlr typing used the describe primers and ionic conditions but slightly varying cycle conditions: 3 min at 96°C, then 30 s at 96°C, 30 s at 62°C, 1.5 min at 72°C x 30, followed by 10 min at 72°C and product characterization as described above.
Histology.
Thick section (3 mm) were fixed in 10% buffered formalin and processed for routine tissue processing, including paraffin embedding. Tissue sections (4 µm) were subsequently examined by hematoxylin and eosin stain. Selected sections were examined with Periodic acid-Schiff (PAS; to delineate glycogenated hepatocytes) and trichrome (to document extent of fibrosis) stains. Additional immunoperoxidase stains were undertaken to delineate hepatocytes [cytokeratin-(818)] and to delineate Kupffer cells (CD68). For immunohistochemical staining, 4-µm sections were placed on a charged slide that already contained necessary positive controls. Slides were dried in a 70° oven for 30 min. Bar-coded sections were processed as follows: 10 min in xylene to deparaffinize and then through two washes of 100% ethanol, two washes of 95% ethanol, one wash of 80% ethanol, and then in tap water. The slides were then transferred to plastic racks and processed in the Ventana Medical Systems apparatus for automated immunohistochemistry. Reagents were loaded in carousels at recommended dilutions. Diaminobenzidine was the chromogen used. Slides were counterstained with hematoxylin and covered with a cover slip.
Cholesterol determination.
Cholesterol concentrations were determined using the cholesterol oxidase method of Heider and Boyett (13). Briefly, liver homogenates were extracted with hexane-isopropanol (3:2) for 1 h, followed by centrifugation to pellet the protein. The organic solution was removed, dried, and spotted on a TLC plate (Redi-Plate, Silica Gel G; Fisher Scientific). To separate neutral lipids, hexane-ethyl ether-glacial acetic acid (1,000:30:1.1) was used as the mobile phase. After being stained with iodine vapor, unesterified cholesterol and esterified cholesterol were scraped from the plate, extracted with hexane, and dried in a glass tube. To each tube, 0.4 ml assay reagent [0.1 ml cholesterol oxidase (8 U/ml), 0.1 ml peroxidase (3,000 U/ml), 1.0 ml p-hydroxyphenylacetic acid (0.15 mg/ml), and 8.8 ml of 0.05 mol/l sodium phosphate buffer, pH 7.0] was added and incubated for 20 min at room temperature. The reaction was terminated by adding 0.8 ml of 0.5 mol/l NaOH. A Perkin-Elmer model LS-40 fluorescence spectrometer (excitation at 325 nm and emission at 415 nm) was used for measuring fluorescence. Unknown cholesterol concentrations were determined from known cholesterol standards using linear regression and normalized to the amount of protein determined using the BCA Protein Assay Kit (Pierce, Rockford, IL).
Liver enzymes.
Serum alanine aminotransferase (ALT) and
-glutamyltransferase (GGT) were performed by standard techniques in the University Animal Care Diagnostic Laboratory of the Arizona Health Sciences Center.
Branched DNA signal amplification assay.
Total RNA was isolated using RNAzol B reagent (Tel-Test, Friendswood, TX) as per the manufacturer's protocol. Each RNA pellet was resuspended in 0.2 ml of 10 mM Tris·HCl buffer, pH 8.0. The concentration of total RNA in each sample was quantified spectrophotometrically at 260 nm. RNA integrity and quality were analyzed by gel electrophoresis with ethidium bromide staining. The quality of RNA samples was judged by the integrity and relative ratio of 28S and 18S rRNA bands.
Reagents required for RNA analysis (i.e., lysis buffer, amplifier/label probe buffer, and substrate solution) were supplied in the QuantiGene Discover kit (Genospectra, Fremont, CA). Specific oligonucleotide probe sets were diluted in lysis buffer. Total RNA (1 µg/µl; 10 µl) was added to each well of a 96-well plate containing capture hybridization buffer [0.05 M HEPES sodium salt, 0.05 M HEPES free acid, 0.037 M lithium lauryl sulfate, 0.5% (vol/vol) Micr-O-protect, 8 mM EDTA, and 0.3% (wt/vol) nucleic acid blocking agent] and 50 µl diluted probe set (50, 100, and 200 fmol/µl capture, blocker, and label probes, respectively). Total RNA was allowed to hybridize to each probe set containing all probes for a given transcript (blocker probes, capture probes, and label probes) overnight at 53°C. Subsequent hybridization and posthybridization wash steps were carried out according to the manufacturer's direction, and luminescence was measured with the Quantiplex 320 branched DNA Luminometer (Bayer Diagnostics) interfaced with Quantiplex Data Management Software version 5.02 (Bayer Diagnostics) for analysis of luminescence from 96-well plates. The probes used for Oatp2, Oatp4, and Bsep are provided in Table 1. Probes for multidrug resistance protein (Mrp) 3 are described by Cherrington et al. (7); those for Mrp 1, 2, 4, and 5, Ntcp, and Oatp1 are from Aleksunes et al. (1).
 |
RESULTS
|
---|
General.
As previously noted, heterozygosity or homozygosity for the Ldlr knockout has little effect on the neurological course of the disease when mice are on a regular diet (6% fat; see Refs. 9 and 46). We placed Npc1/ mice of varying Ldlr genotypes on a high-fat diet (18% total fat, 1% cholesterol) to study the effects of high serum cholesterol on the disorder. The high-fat, 1% cholesterol diet caused a dramatic disease in Npc1/ mice independent of Ldlr knockout genotype [
die (or are moribund and killed to obtain tissues) before 43 days]. The survivors live about as long as Npc1/ mice on a normal diet (mostly
80 days but a few as early as 57 days: on a regular diet Npc1/ live 74.2 ± 7.4 days). The early death does not correlate with the Ldlr status of the mice but is influenced by sex, with more males dying early (
2 early vs. late, male vs. female, P
0.02).
We surmised that this severe early lethality might be because of liver disease, since the livers were greatly enlarged postmortem. Npc1/ mice have mildly enlarged livers on a regular diet (6% of body wt compared with 4% for normal mice). There is a sex difference for weight on the regular diet but not the high-fat diet (males are heavier) and no sex difference in liver weight as a percentage of body weight. Thus this is the variable compared. It increased dramatically to 14% in the early decedents and up to 22% in the longer-surviving Npc1/ mice. The high-fat, 1% cholesterol diet only increased the weight of normal mouse livers to 8% of body weight, independent of Ldlr status. Although not significant, there was a trend for Npc11, Ldlr/ mice to have less hepatomegaly.
Confirmation of hepatitis in the Npc1/ mice was found from enzyme studies and histology. Serum ALTs were mildly elevated in Npc1/ mice on a regular diet but mostly became strongly elevated when the mice were placed on the high-fat diet (Table 2). This increase did not occur in the normal mice placed on the high-fat, 1% cholesterol diet. These differences were not significant because of the high scatter in the values. Serum GGT was already high in Npc1/ mice on a regular diet and became markedly higher on the high-fat diet (Table 2). The high-fat diet elevated serum cholesterols markedly in control Npc1+/ but not in Npc1/ mice (Table 3). Npc1/, Ldlr/ mice had lower cholesterols than did Npc1+/ mice.
Histology.
Standard hematoxylin and eosin staining shows massive accumulation of presumed lipid droplets in the livers of Npc1/ mice on a regular diet independent of LDLR genotype (Fig. 1A). The fatty nature of these was confirmed with Oil red-O staining (data not shown). This was enhanced on the high-fat diet (Fig. 1B). Negative PAS staining confirmed that the vacuoles were not glycogen (Fig. 2). When CD68 and cytokeratin 18 immunohistochemistry were used to distinguish between macrophage storage and hepatocyte storage, lipid accumulation is noted in both cell types (Fig. 3) but is more marked in hepatocytes. Trichrome staining does not show much fibrosis in the younger mice but mild diffuse fibrosis in older mice, which is not zone dependent, as would be found in nonalcoholic hepatic steatosis (Fig. 4). Such fibrosis was only seen in Npc1/ and when on the high-fat diet.

View larger version (136K):
[in this window]
[in a new window]
|
Fig. 1. Lipid accumulation in Nieman-Pick C (NPC)-deficient (Npc1/) livers of mice on a high-fat, 1% cholesterol diet. Hematoxylin- and eosin-stained liver sections at x400 magnification showing pale cells with presumed lipid accumulation and cell degeneration with inflammation (arrow) in Ldlr/ mice. A: regular diet; B: high-fat diet.
|
|

View larger version (156K):
[in this window]
[in a new window]
|
Fig. 2. Lack of glycogen in Npc1/ livers of mice on a high-fat, 1% cholesterol diet. Periodic acid-Schiff (PAS)-stained liver tissue at x400 magnification supporting lipid accumulation, and not glycogen, in hepatocytes on high-fat diet in Ldlr/ mouse.
|
|

View larger version (135K):
[in this window]
[in a new window]
|
Fig. 3. Lipid accumulation in hepatocytes of Npc1/ mice on a high-fat, 1% cholesterol diet. Indirect immunoperoxidase stains for Kupffer cells (CD68; A, arrow) and for hepatocytes (cytokeratin 8/18; B, arrow). The lipid accumulation is primarily in hepatocytes.
|
|

View larger version (151K):
[in this window]
[in a new window]
|
Fig. 4. Liver fibrosis in Npc1/ mice on a high-fat, 1% cholesterol diet. Trichrome-stained section at x400 magnification showing pericentral and minimal sinusoidal fibrosis (arrow) on a high-fat diet, Ldlr/ mouse.
|
|
Liver cholesterol.
Unesterified cholesterol is stored massively in the livers of Npc1/ mice independent of Ldlr genotype (Table 4). On the regular diet, esterified cholesterol is decreased compared with normals in Npc1/ livers, and this is relatively unchanged by Ldlr genotype. On high-fat diets, there is more variable storage of unesterified cholesterol, which is still greatly increased compared with normals on a regular diet. However, esterified cholesterol is now quite markedly elevated. This great alteration in the pattern of storage of the esterified cholesterol does not correlate with the serum level of cholesterol (Table 3).
Changes in bile acid transporters.
As seen in Fig. 5A, mRNAs for Mrps 1, 2, 3, and 5 were elevated in Npc1/ livers on a regular diet; they became more markedly elevated on the high-fat diet, and Mrp 4 became elevated. In contrast, mRNAs for Ntcp, Bsep, and OatP2 were initially elevated in Npc1/ liver. The levels of mRNA for Ntcp, Bsep, and OatP1 and -2 were suppressed by the high-fat diet in the Npc1/ while only OatP1 mRNA levels were suppressed in normals.

View larger version (25K):
[in this window]
[in a new window]
|
Fig. 5. Hepatic transporter mRNA levels determined by QuantiGene signal amplification. Total hepatic RNA from control and Npc1/ mice on a regular and high-fat diet was analyzed by the branched DNA assay. Data are expressed as relative light units (RLU)/10 µg total RNA ± SE. A: multidrug resistance protein (Mrp) transporters. B: Ntcp, Bsep, and Oatp transporters.
|
|
 |
DISCUSSION
|
---|
The early description of the mutant mice now known to be Npc1/ mice included extensive descriptions of the liver pathology and storage material (18, 20, 2224, 27). The time course of this accumulation was also studied and found to be at its maximum level at 6 wk of age, with the weights of liver and spleen decreasing in parallel with the decrease in body weight thereafter (19). The accumulation of hepatic unesterified cholesterol was found to be increased when the mice were fed a 10% fat, 1% cholesterol diet (22). Mutational analyses of the human NPC1 gene have not disclosed particular mutants associated with liver disease (3). We have studied the interaction of this deficiency, the LDL receptor deficiency, and a high-fat, 1% cholesterol diet.
The rapid death with the high-fat, 1% cholesterol diet in two-thirds of the mice was a very different phenotype than had previously been observed. Npc1/ mice usually achieve near-normal adult weight and then develop neurological symptoms, although these early lethal mice did not nearly achieve such a weight and wasted away with few neurological symptoms. The segregation into two groups was independent of Ldlr genotype (Table 1) but was influenced by sex, with males more severely affected. This is in contrast to the mdr2 knockout mice where females are more severely affected (29) but in accord with cholic acid-fed, bile salt export pump knockout mice where males are more severely affected (30). Because our mice came from an F2 of the Balb/cJ strain (source of Npc1) with the C57BL/6 strain (source of Ldlr receptor), it was possible that a gene was segregating to cause this difference. The ratio of 29 to 15 is not significantly different from a 3 to 1 ratio, which could be explained by a recessive gene protecting the mice.
The early lethality was associated with greatly increased liver weights, up to an average of 14% of total body weight in the early lethal mice and even more in the late survivors. Thus liver size was not the sole cause of early death. The rapid death might involve an interaction of hepatic encephalopathy with the neurodegeneration of Npc1/, but the precise cause is unclear. Although not statistically significant, there was a trend for the Ldlr knockout to protect the mice from this massive increase in liver weight, which is consistent with the role of LDLR in cholesterol uptake in mouse liver, although LDLR is not involved in mice to the degree that it is in human liver (14). This may be the reason that variation in it had relatively little effect, contrary to our expectation, on the liver disease. The high-fat diet was also associated with increases in serum ALT (although these were not statistically significant given large variation in the values) and GGT in the Npc1+/ mice. Serum cholesterol was also increased by the high-fat, 1% cholesterol diet, although this occurred more or less independent of the Npc1 genotype but, of course, was greatly affected by the Ldlr knockout.
The unesterified cholesterol content of the liver tended to be reduced with the high-fat diet (Table 4). In contrast, esterified cholesterol, which is normally very low in Npc1/ mice on a regular diet, became very greatly elevated, again relatively unaffected by Ldlr status (Table 4). These findings of massive cholesterol storage were confirmed by liver histology. Massive fat storage, more so in hepatocytes than macrophages, was documented (Figs. 13). In the older surviving mice, mild diffuse fibrosis, which was zone independent, was found (Fig. 4). Collagen deposition in Npc1/ mouse livers on a regular diet has been shown previously (12). Because recent data suggest that the degree of alteration in cholesterol homeostasis in NPC1 is correlated with a deficiency in the generation of 25- and 27-hydroxycholesterol from LDL cholesterol (9), these hydroxy cholesterols need further study in these mice.
A major function of hepatic transporters is to move a wide range of organic substances across cell membranes, thereby facilitating the excretion of these compounds from blood into bile. During periods of hepatic stress, such as sepsis or cholestasis, the downregulation of several sinusoidal uptake transporters, such as Ntcp, may be one compensatory mechanism to protect the liver against the accumulation of toxic substances. Conversely, the upregulation of sinusoidal excretion transporters such as Mrp 3 may also compensate by exporting these compounds from the liver back into the blood. Thus the putative role of altered regulation of hepatic transporters during stress is to decrease the concentration of potentially toxic molecules in the liver. In the present study, mRNA levels for several uptake transporters were higher in Npc1/ mice than in control mice but significantly reduced by the high-fat diet. Additionally, the expression of several Mrp transporters was elevated in Npc1/ mice and was further increased by the high-fat diet. Whether the transcriptional mechanisms that are responsible for altered regulation of transporters during cholestatic stress are the same mechanisms responsible for the changes observed in Npc1/ mice or with the high-fat diet remains to be determined.
A recent study of plasma cholesterol and biliary lipid secretion in Npc1/ mice used a 2-wk supplementation of the diet with 2% cholesterol (2). This shorter treatment did not result in the dramatic increases in liver weight and frequent early mortality that we found with the higher total fat and cholesterol diet given continuously. However, the 2-wk treatment with a high-cholesterol diet did result in increases in plasma cholesterol in the Npc1/ mice, a trend that we observed but which was not statistically significant in our hands. Amigo et al. (2) found large increases in cholesterol output and lower increases in bile salts and phospholipid output in bile on the 2% cholesterol diet. These authors also found that the ATP-binding cassette proteins were differentially expressed in Npc1/ vs. control mice after consumption of the 2% cholesterol diet for 2 wk. ABCA1 expression increased by 100% in Npc1/ mice but did not change in controls on the diet, whereas ABCG5 and -8 were upregulated by 5080% in the controls but did not increase in the Npc1/ mice (1). ABCG5 and -8 are important in biliary cholesterol secretion (33) and are targets, as is ABCA1, of the nuclear oxysterol receptor LXR (26). Thus Amigo et al.'s (2) findings indicate that the LXR pathway is differentially, or not at all, activated by cholesterol in Npc1/ mice. Thus deficiencies in intracellular cholesterol metabolism presumably inhibit signaling via LXR for these ABC proteins, while LXR, or other signaling pathways such as the farnesoid X receptor (21, 28), may mediate other pathological changes in gene expression.
In summary, we have found that feeding Npc1/ mice a high-fat, 1% cholesterol diet resulted in early lethality for a majority of the mice. This lethality was associated with massive storage of esterified and unesterified cholesterol and elevations in liver enzymes. A number of bile acid transporters were elevated in Npc1/ livers on a regular diet but were differentially regulated on the high-fat diet. The Npc1//Ld1r/ mice had lower serum cholesterol than Npc1/ mice on both diets. It appears that the cholestatic hepatitis, which is frequently found in human NPC disease, was exacerbated by the high-fat diet in this mouse model of NPC1.
 |
GRANTS
|
---|
This work was supported by National Institutes of Health Grants EB-00343-03 and ES-011646.
 |
ACKNOWLEDGMENTS
|
---|
We thank Donelle Myers for secretarial support and Dr. William S. Garver for help with cholesterol determinations.
Current address for R. J. Hunter: Dept. of Surgery, Univ. of Arizona School of Health Sciences, Tucson, AZ 85724.
 |
FOOTNOTES
|
---|
Address for reprint requests and other correspondence: Robert P. Erickson at the Dept. of Pediatrics/Rm. 4341B, 1501 N. Campbell Ave., P. O. Box 245073, Tucson, Arizona 85724-5073 (e-mail: erickson{at}peds.arizona.edu)
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
 |
REFERENCES
|
---|
- Aleksunes LM, Slitt AL, Cherrington NJ, Thibodeau MS, Klaassen CD, and Manautou JE. Differential expression of mouse hepatic transporter genes in response to acetaminophen and carbon tetrachloride. Tox Sci 83: 4452, 2005.[ISI]
- Amigo L, Mendoza H, Castro J, Quinones V, Miquel JF, and Zanlungo S. Relevance of Niemann-Pick type C1 protein expression in controlling plasma cholesterol and biliary lipid secretion in mice. Hepatology 36: 819828, 2002.[ISI][Medline]
- Bauer P, Knoblich R, Bauer C, Finckh U, Hufen A, Kropp J, Braun S, Kustermann-Kuhn B, Schmidt D, Harzer K, and Rolfs A. NPC1: Complete genomic sequence, mutation analysis, and characterization of haplotypes. Hum Mutat 19: 3038, 2002.[CrossRef][ISI][Medline]
- Boison D, Scheurer L, Zumsteg V, Rulicke T, Litynski P, Fowler B, Brandner S, and Mohler H. Neonatal hepatic steatosis by disruption of the adenosine kinase gene. Proc Natl Acad Sci USA 99: 69856990, 2002.[Abstract/Free Full Text]
- Boles RG, Buck EA, Blitzer MG, Platt MS, Cowan TM, Martin SK, Yoon H, Madsen JA, Reyes-Mugica M, and Rinaldo P. Retrospective biochemical screening of fatty acid oxidation disorders in postmortem livers of 418 cases of sudden death in the first year of life. J Pediatr 132: 924933, 1998.[ISI][Medline]
- Brivet M, Boutron A, Slama A, Costa C, Thuillier L, Demaugre F, Rabier D, Saudubray JM, and Bonnefont JP. Defects in activation and transport of fatty acids. J Inherit Metab Dis 22: 428441, 1999.[CrossRef][ISI][Medline]
- Cherrington NJ, Slitt AL, Maher JM, Zhang XX, Zhang J, Huang W, Wan YJ, Moore DD, and Klaassen CD. Induction of multidrug resistance protein 3 (mrp3) in vivo is independent of constitutive androstane receptor. Drug Metab Dispos 31: 13151319, 2003.[Abstract/Free Full Text]
- Erickson RP, Garver WS, Camargo F, Hossain GS, and Heidenreich RA. Pharmacological and genetic modifications of somatic cholesterol do not substantially alter the course of CNS disease in Niemann-Pick C mice. J Inherit Metab Dis 23: 5462, 2000.[CrossRef][ISI][Medline]
- Frolov A, Zielinski SE, Crowley JR, Dudley-Rucker N, Schaffer JE, and Ory DS. NPC1 and NPC2 regulate cellular cholesterol homeostasis through generation of low density lipoprotein cholesterol-derived oxysterols. J Biol Chem 278: 2551725525, 2003.[Abstract/Free Full Text]
- Fromenty B and Pessayre D. Impaired mitochondrial function in microvesicular steatosis. Effects of drugs, ethanol, hormones and cytokines. J Hepatol 26: 4353, 1997.[CrossRef][ISI][Medline]
- Gaw A, Mancini FP, and Ishibashi S. Rapid genotyping of low density lipoprotein receptor knockout mice using a polymerase chain reaction technique. Lab Anim Sci 29: 447449, 1995.
- Guo J, Trouard TP, Galons JP, Erickson R, and Gillies RJ. Magnetization transfer contrast image in Niemann pick type C mouse liver. J Magn Reson Imaging 18: 321327, 2003.[CrossRef][ISI][Medline]
- Heider JG and Boyett RL. The picomole determination of free and total cholesterol in cells in culture. J Lipid Res 19: 514518, 1978.[Abstract]
- Ishibashi S, Brown MS, Goldstein JL, Gerard RD, Hammer RE, and Herz J. Hypercholesterolemia in low density lipoprotein receptor knockout mice and its reversal by adenovirus-mediated gene delivery. J Clin Invest 92: 883893, 1993.[ISI][Medline]
- Kelly DA, Portmann B, Mowat AP, Sherlock S, and Lake BD. Niemann-Pick disease type C: diagnosis and outcome in children, with particular reference to liver disease. J Pediatr 123: 242247, 1993.[ISI][Medline]
- Loftus SK, Morris JA, Carstea ED, Gu JZ, Cummings C, Brown A, Ellison J, Ohno K, Rosenfeld MA, Tagle DA, Pentchev PG, and Pavan WJ. Murine model of Niemann-Pick C disease: mutation in a cholesterol homeostasis gene. Science 277: 232235, 1997.[Abstract/Free Full Text]
- Mieli-Vergani G, Howard ER, and Mowat AP. Liver disease in infancy: a 20-year perspective. Gut Suppl: S123S128, 1991.
- Miyawaki S, Mitsuoka S, Sakiyama T, and Kitagawa T. Sphingomyelinosis, a new mutation in the mouse: a model of Niemann-Pick disease in humans. J Hered 73: 257263, 1982.[ISI][Medline]
- Miyawaki S, Mitsuoka S, Sakiyama T, and Kitagawa T. Time course of hepatic lipids accumulation in a strain of mice with an inherited deficiency of sphingomyelinase. J Hered 74: 465468, 1983.[ISI][Medline]
- Morris MD, Bhuvaneswaran C, Shio H, and Fowler S. Lysosome lipid storage disorder in NCTR-BALB/c mice. I. Description of the disease and genetics. Am J Pathol 108: 140149, 1982.[Abstract]
- Parks DJ, Blanchard SG, Bledsoe RK, Chandra G, Consler TG, Kliewer SA, Stimmel JB, Willson TM, Zavacki AM, Moore DD, and Lehmann JM. Bile acids: natural ligands for an orphan nuclear receptor. Science 284: 13651368, 1999.[Abstract/Free Full Text]
- Pentchev PG, Boothe AD, Kruth HS, Weintroub H, Stivers J, and Brady RO. A genetic storage disorder in BALB/C mice with a metabolic block in esterification of exogenous cholesterol. J Biol Chem 259: 57845791, 1984.[Abstract/Free Full Text]
- Pentchev PG, Gal AE, Booth AD, Omodeo-Sale F, Fouks J, Neumeyer BA, Quirk JM, Dawson G, and Brady RO. A lysosomal storage disorder in mice characterized by a dual deficiency of sphingomyelinase and glucocerebrosidase. Biochim Biophys Acta 619: 669679, 1980.[ISI][Medline]
- Pentchev PG, Boothe AD, Kruth HS, Weintroub H, Stivers J, and Brady RO. A genetic storage disorder in BALB/C mice with a metabolic block in esterification of exogenous cholesterol. J Biol Chem 259: 57845791, 1984.[Abstract/Free Full Text]
- Pentchev PG, Vanier MT, Suzuki K, and Patterson MC. Nieman-Pick disease type C. A cellular cholesterol lipidosis. In: The Metabolic and Molecular Basis of Inherited Disease (8th ed.), edited by Scriver CR, Beaudet AL, Sly WS, and Valle D. New York: McGraw-Hill, 1995, p. 55875627.
- Repa JJ, Berge KE, Pomajzl C, Richardson JA, Hobbs H, and Mangelsdorf DJ. Regulation of ATP-binding cassette sterol transporters ABCG5 and ABCG8 by the liver X receptors alpha and beta. J Biol Chem 277: 1879318800, 2002.[Abstract/Free Full Text]
- Shio H, Fowler S, Bhuvaneswaran C, and Morris MD. Lysosome lipid storage disorder in NCTR-BALB/c mice. II. Morphologic and cytochemical studies. Am J Pathol 108: 150159, 1982.[Abstract]
- Sinal CJ, Tohkin M, Miyata M, Ward JM, Lambert G, and Gonzalez FJ. Targeted disruption of the nuclear receptor FXR/BAR impairs bile acid and lipid homeostasis. Cell 102: 731744, 2000.[CrossRef][ISI][Medline]
- van Nieuwerk CMJ, Groen AK, Ottenhoff R, van Wijland M, van den Bergh Weerman and MA, Thtgat GNJ, Offerhaus JJA, and Oude Elferink RPJ. The role of bile salt composition in liver pathology of mdr2 (-/-) mice: differences between males and females. J Hepatol 26: 138145, 1997.[CrossRef][ISI][Medline]
- Wang R, Lam P, Liu L, Forrest D, Yousef IM, Mignault D, Phillips MJ, and Ling V. Severe Cholestasis induced by cholic acid feeding in knockout mice of sister of p-glycoprotein. Hepatology 38: 14891499, 2003.[ISI][Medline]
- Xie C, Turley SD, and Dietschy JM. Centripetal cholesterol flow from the extrahepatic organs through the liver is normal in mice with mutated Niemann-Pick type C protein (NPC1). J Lipid Res 41: 12781289, 2000.[Abstract/Free Full Text]
- Yerushalmi B, Sokol RJ, Narkewicz MR, Smith D, Ashmead JW, and Wenger DA. Niemann-Pick disease type C in neonatal cholestasis at a North American center. J Pediatr Gastroenterol Nutr 35: 4450, 2002.[CrossRef][ISI][Medline]
- Yu L, Hammer RE, Li-Hawkins J, Von Bergmann K, Lutjohann D, Cohen JC, and Hobbs HH. Disruption of Abcg5 and Abcg8 in mice reveals their crucial role in biliary cholesterol secretion. Proc Natl Acad Sci USA 99: 1623716242, 2002.[Abstract/Free Full Text]
Copyright © 2005 by the American Physiological Society.