Transport Of Phosphatidylcholine Research Articles

Molecular basis of progressive familial intrahepatic cholestasis 3. A proteomics study.

Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a severe rare liver disease that affects between 1/50,000 and 1/100,000 children. In physiological conditions, bile is produced by the liver and stored in the gallbladder, and then it flows to the small intestine to play its role in fat digestion. To prevent tissue damage, bile acids (BAs) are kept in phospholipid micelles. Mutations in phosphatidyl choline transporter ABCB4 (MDR3) lead to intrahepatic accumulation of free BAs that result in liver damage. PFIC3 onset usually occurs at early ages, progresses rapidly, and the prognosis is poor. Currently, besides the palliative use of ursodeoxycholate, the only available treatment for this disease is liver transplantation, which is really challenging for short-aged patients. To gain insight into the pathogenesis of PFIC3 we have performed an integrated proteomics and phosphoproteomics study in human liver samples to then validate the emerging functional hypotheses in a PFIC3 murine model. We identified 6246 protein groups, 324 proteins among them showing differential expression between control and PFIC3. The phosphoproteomic analysis allowed the identification of 5090 phosphopeptides, from which 215 corresponding to 157 protein groups, were differentially phosphorylated in PFIC3, including MDR3. Regulation of essential cellular processes and structures, such as inflammation, metabolic reprogramming, cytoskeleton and extracellular matrix remodeling, and cell proliferation, were identified as the main drivers of the disease. Our results provide a strong molecular background that significantly contributes to a better understanding of PFIC3 and provides new concepts that might prove useful in the clinical management of patients.

Quantifying Functional Impairment of ABCA3 Variants Associated with Interstitial Lung Disease.

ATP-binding cassette subfamily A member 3 (ABCA3) is a lipid transporter within alveolar type II cells. Patients with bi-allelic variants in ABCA3 may suffer from a variable severity of interstitial lung disease. We characterized and quantified ABCA3 variants' overall lipid transport function by assessing the in vitro impairment of its intracellular trafficking and pumping activity. We expressed the results relative to the wild type, integrated the quantitative readouts from eight different assays and used newly generated data combined with previous results to correlate the variants' function and clinical phenotype. We differentiated normal (within 1 normalized standard deviation (nSD) of the wild-type mean), impaired (within 1 to 3 nSD) and defective (beyond 3 nSD) variants. The transport of phosphatidylcholine from the recycling pathway into ABCA3+ vesicles proved sensitive to the variants' dysfunction. The sum of the quantitated trafficking and pumping predicted a clinical outcome. More than an approximately 50% loss of function was associated with considerable morbidity and mortality. The in vitro quantification of ABCA3 function enables detailed variant characterization, substantially improves the phenotype prediction of genetic variants and possibly supports future treatment decisions.

The neglected biliary mucus and its phosphatidylcholine content: a putative player in pathogenesis of primary cholangitis-a narrative review article.

Primary sclerosing cholangitis (PSC) is a rare progressive cholangitis resulting in cirrhosis and cholangiocellular carcinoma. The pathogenesis is unclear and an effective medical therapy is not available. It is highly associated to ulcerative colitis for which recently a disturbance of the tight junction (TJ) barrier has been claimed as etiologic feature. Genetic mouse models with intestinal TJ disruption showed a defective transport of phosphatidylcholine (PC) to intestinal mucus. Consequently, an ulcerative colitis phenotype developed. In the present study we evaluate whether there is also a paracellular transport of PC through TJ to the apical side of cholangiocytes. As in ulcerative colitis, a TJ defect could lead to deficient PC in biliary mucus. It would impair the protective barrier against aggressive bile acids in bile. Indeed with polarized biliary tumor cells a vectorial transport of PC from basal to luminal side was demonstrated using a transwell culture system. PC was not taken up by the cells but moved paracellularly via TJ to the apical side driven by luminal HCO3− generated by the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) and the anion exchange protein 2 (AE2). If such a TJ-mediated PC translocation to the apical surface of cholangiocytes could be disrupted in a genetic mouse model, a PSC phenotype would be expected. With such an experimental model functional operative therapies can be evaluated. We propose that disruption of TJ mediated paracellular transport of PC to the apical side of cholangiocytes could lead to biliary mucus PC depletion. This may be a pathogenetic factor for development of PSC.

In Situ Separation and Analysis of Lipids by Paper Spray Ionization Mass Spectrometry

Paper spray ionization (PSI) is an extractive ambient ionization technique for mass spectrometry (MS), whereby a triangular paper tip serves as the sampling base and the electrospray tip. During PSI, analytes are extracted and transported to the edge of the paper tip by the applied spraying solvent. Analytes can be purified from a sample matrix and separated from each other by this transportation process. In this study, we investigated and utilized the analyte transportation process of PSI for the in situ separation and analysis of lipid mixtures. We found that differential transport of phosphatidylcholine (PC) and triacylglycerol (TAG), the two most abundant lipid classes in animals, occurred during PSI. We also found that the order in which these lipids moved strongly depended on how the spraying solvent was applied to the paper base. The more polar PC moved faster than the less polar TAG during PSI, when a polar solvent was slowly fed into a paper tip, whereas TAG was transported faster than PC when excess solvent was applied to the tip at once. In addition, we achieved a complete separation and detection of PC and TAG by slowly supplying a nonpolar solvent to a PSI tip.

Synthetic human ABCB4 mRNA therapy rescues severe liver disease phenotype in a BALB/c.Abcb4-/- mouse model of PFIC3

Background & Aims:Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a rare lethal autosomal recessive liver disorder caused by loss-of-function variations of the ABCB4 gene, encoding a phosphatidylcholine transporter (ABCB4/MDR3). Currently, no effective treatment exists for PFIC3 outside of liver transplantation.Methods:We have produced and screened chemically and genetically modified mRNA variants encoding human ABCB4 (hABCB4 mRNA) encapsulated in lipid nanoparticles (LNPs). We examined their pharmacological effects in a cell-based model and in a new in vivo mouse model resembling human PFIC3 as a result of homozygous disruption of the Abcb4 gene in fibrosis-susceptible BALB/c.Abcb4−/− mice.Results:We show that treatment with liver-targeted hABCB4 mRNA resulted in de novo expression of functional hABCB4 protein and restored phospholipid transport in cultured cells and in PFIC3 mouse livers. Importantly, repeated injections of the hABCB4 mRNA effectively rescued the severe disease phenotype in young Abcb4−/− mice, with rapid and dramatic normalisation of all clinically relevant parameters such as inflammation, ductular reaction, and liver fibrosis. Synthetic mRNA therapy also promoted favourable hepatocyte-driven liver regeneration to restore normal homeostasis, including liver weight, body weight, liver enzymes, and portal vein blood pressure.Conclusions:Our data provide strong preclinical proof-of-concept for hABCB4 mRNA therapy as a potential treatment option for patients with PFIC3.

Transport mechanism of P4 ATPase phosphatidylcholine flippases.

The P4 ATPases use ATP hydrolysis to transport large lipid substrates across lipid bilayers. The structures of the endosome- and Golgi-localized phosphatidylserine flippases-such as the yeast Drs2 and human ATP8A1-have recently been reported. However, a substrate-binding site on the cytosolic side has not been found, and the transport mechanisms of P4 ATPases with other substrates are unknown. Here, we report structures of the S. cerevisiae Dnf1-Lem3 and Dnf2-Lem3 complexes. We captured substrate phosphatidylcholine molecules on both the exoplasmic and cytosolic sides and found that they have similar structures. Unexpectedly, Lem3 contributes to substrate binding. The conformational transitions of these phosphatidylcholine transporters match those of the phosphatidylserine transporters, suggesting a conserved mechanism among P4 ATPases. Dnf1/Dnf2 have a unique P domain helix-turn-helix insertion that is important for function. Therefore, P4 ATPases may have retained an overall transport mechanism while evolving distinct features for different lipid substrates.

Flippases play specific but distinct roles in the development, pathogenicity, and secondary metabolism of Fusarium graminearum.

The membrane trafficking system is important for compartmentalization of the biosynthesis pathway and secretion of deoxynivalenol (DON) mycotoxin (a virulence factor) in Fusarium graminearum. Flippases are transmembrane lipid transporters and mediate a number of essential physiological steps of membrane trafficking, including vesicle budding, charging, and protein diffusion within the membrane. However, the roles of flippases in secondary metabolism remain unknown in filamentous fungi. Herein, we identified five flippases (FgDnfA, FgDnfB, FgDnfC1, FgDnfC2, and FgDnfD) in F. graminearum and established their specific and redundant functions in the development and pathogenicity of this phytopathogenic fungus. Our results demonstrate that FgDnfA is critical for normal vegetative growth while the other flippases are dispensable. FgDnfA and FgDnfD were found crucial for the fungal pathogenesis, and a remarkable reduction in DON production was observed in ΔFgDNFA and ΔFgDNFD. Deletion of the FgDNFB gene increased DON production to about 30 times that produced by the wild type. Further analysis showed that FgDnfA and FgDnfD have positive roles in the regulation of trichothecene (TRI) genes (TRI1, TRI4, TRI5, TRI6, TRI12, and TRI101) expression and toxisome reorganization, while FgDnfB acts as a negative regulator of DON synthesis. In addition, FgDnfB and FgDnfD have redundant functions in the regulation of phosphatidylcholine transport, and double deletion of FgDNFB and FgDNFD showed serious defects in fungal development, DON synthesis, and virulence. Collectively, our findings reveal the distinct and specific functions of flippase family members in F. graminearum and principally demonstrate that FgDnfA, FgDnfD, and FgDnfB have specific spatiotemporal roles during toxisome biogenesis.

Hydrophilic bile acids prevent liver damage caused by lack of biliary phospholipid in Mdr2−/− mice

Bile acid imbalance causes progressive familial intrahepatic cholestasis type 2 (PFIC2) or type 3 (PFIC3), severe liver diseases associated with genetic defects in the biliary bile acid transporter bile salt export pump (BSEP; ABCB11) or phosphatidylcholine transporter multidrug resistance protein 3 (MDR3; ABCB4), respectively. Mdr2-/- mice (a PFIC3 model) develop progressive cholangitis, ductular proliferation, periportal fibrosis, and hepatocellular carcinoma (HCC) because the nonmicelle-bound bile acids in the bile of these mice are toxic. We asked whether the highly hydrophilic bile acids generated by Bsep-/- mice could protect Mdr2-/- mice from progressive liver damage. We generated double-KO (DKO: Bsep-/- and Mdr2-/- ) mice. Their bile acid composition resembles that of Bsep-/- mice, with increased hydrophilic muricholic acids, tetrahydroxylated bile acids (THBAs), and reduced hydrophobic cholic acid. These mice lack the liver pathology of their Mdr2-/- littermates. The livers of DKO mice have gene expression profiles very similar to Bsep-/- mice, with 4,410 of 6,134 gene expression changes associated with the Mdr2-/- mutation being suppressed. Feeding with THBAs partially alleviates liver damage in the Mdr2-/- mice. Hydrophilic changes to biliary bile acid composition, including introduction of THBA, can prevent the progressive liver pathology associated with the Mdr2-/- (PFIC3) mutation.

Sustained activation of detoxification pathways promotes liver carcinogenesis in response to chronic bile acid-mediated damage.

Chronic inflammation promotes oncogenic transformation and tumor progression. Many inflammatory agents also generate a toxic microenvironment, implying that adaptive mechanisms must be deployed for cells to survive and undergo transformation in such unfavorable contexts. A paradigmatic case is represented by cancers occurring in pediatric patients with genetic defects of hepatocyte phosphatidylcholine transporters and in the corresponding mouse model (Mdr2-/- mice), in which impaired bile salt emulsification leads to chronic hepatocyte damage and inflammation, eventually resulting in oncogenic transformation. By combining genomics and metabolomics, we found that the transition from inflammation to cancer in Mdr2-/- mice was linked to the sustained transcriptional activation of metabolic detoxification systems and transporters by the Constitutive Androstane Receptor (CAR), a hepatocyte-specific nuclear receptor. Activation of CAR-dependent gene expression programs coincided with reduced content of toxic bile acids in cancer nodules relative to inflamed livers. Treatment of Mdr2-/- mice with a CAR inhibitor blocked cancer progression and caused a partial regression of existing tumors. These results indicate that the acquisition of resistance to endo- or xeno-biotic toxicity is critical for cancers that develop in toxic microenvironments.

Comparison of in silico prediction and experimental assessment of ABCB4 variants identified in patients with biliary diseases

Genetic variations of the phosphatidylcholine transporter, ABCB4 cause several biliary diseases. The large number of reported variations makes it difficult to foresee a comprehensive study of each variation. To appreciate the reliability of in silico prediction programs, 1) we confronted them with the assessment in cell models of two ABCB4 variations (E528D and P1161S) identified in patients with low phospholipid-associated cholelithiasis (LPAC); 2) we extended the confrontation to 19 variations that we had previously characterized in cellulo. Four programs (Provean, Polyphen-2, PhD-SNP and MutPred) were used to predict the degree of pathogenicity. The E528D and P1161S variants were studied in transfected HEK293 and HepG2 cells by immunofluorescence, immunoblotting and measurement of phosphatidylcholine secretion. All prediction tools qualified the P1161S variation as deleterious, but provided conflicting results for E528D. In cell models, both mutants were expressed and localized as the wild type but their activity was significantly reduced, by 48% (P1161S) and 33% (E528D). These functional defects best correlated with MutPred predictions. MutPred program also proved the most accurate to predict the pathogenicity of the 19 ABCB4 variants that we previously characterized in cell models, and the most sensitive to predict the pathogenicity of 65 additional mutations of the Human Gene Mutation Database. These results confirm the pathogenicity of E528D and P1161S variations and suggest that even a moderate decrease (by less than 50%) of phosphatidylcholine secretion can cause LPAC syndrome. They highlight the reliability of in silico prediction tools, most notably MutPred, as a first approach to predict the pathogenicity of ABCB4 variants.

TLR4 Deficiency Protects against Hepatic Fibrosis and Diethylnitrosamine-Induced Pre-Carcinogenic Liver Injury in Fibrotic Liver.

BackgroundThe development of hepatocellular carcinoma (HCC) is a common consequence of advanced liver fibrosis but the interactions between fibrogenesis and carcinogenesis are still poorly understood. Recently it has been shown that HCC promotion depends on Toll-like receptor (TLR) 4. Pre-cancerogenous events can be modelled in mice by the administration of a single dose of diethylnitrosamine (DEN), with HCC formation depending amongst others on interleukin (IL) 6 production. Mice lacking the hepatocanalicular phosphatidylcholine transporter ABCB4 develop liver fibrosis spontaneously, resemble patients with sclerosing cholangitis due to mutations of the orthologous human gene, and represent a valid model to study tumour formation in pre-injured cholestatic liver. The aim of this study was to investigate DEN-induced liver injury in TLR4-deficient mice with biliary fibrosis.MethodsABCB4-deficient mice on the FVB/NJ genetic background were crossed to two distinct genetic backgrounds (TLR4-sufficient C3H/HeN and TLR4-deficient C3H/HeJ) for more than 10 generations. The two congenic knockout and the two corresponding wild-type mouse lines were treated with a single dose of DEN for 48 hours. Phenotypic differences were assessed by measuring hepatic collagen contents, inflammatory markers (ALT, CRP, IL6) as well as hepatic apoptosis (TUNEL) and proliferation (Ki67) rates.ResultsHepatic collagen accumulation is significantly reduced in ABCB4-/-:TLR4-/-double-deficient mice. After DEN challenge, apoptosis, proliferation and inflammatory markers are decreased in TLR4-deficient in comparison to TLR4-sufficient mice. When combining ABCB4 and TLR4 deficiency with DEN treatment, hepatic IL6 expression and proliferation rates are lowest in fibrotic livers from the double-deficient line. Consistent with these effects, selective digestive tract decontamination in ABCB4-/- mice also led to reduced tumor size and number after DEN.ConclusionThis study demonstrates that liver injury upon DEN challenge depends on pre-existing fibrosis and genetic background. The generation of ABCB4-/: TLR4-/- double-deficient mice illustrates that TLR4-deficiency protects against hepatic injury in a preclinical mouse model of chronic liver disease.

Haploinsufficiency for Stard7 is associated with enhanced allergic responses in lung and skin.

Allergic asthma is a chronic inflammatory disorder that affects ∼20% of the population worldwide. Microarray analyses of nasal epithelial cells from acute asthmatic patients detected a 50% decrease in expression of Stard7, an intracellular phosphatidylcholine transport protein. To determine whether loss of Stard7 expression promotes allergic responses, mice were generated in which one allele of the Stard7 locus was globally disrupted (Stard7 (+/-) mice). OVA sensitization and challenge of Stard7(+/-) mice resulted in a significant increase in pulmonary inflammation, mucous cell metaplasia, airway hyperresponsiveness, and OVA-specific IgE compared with OVA-sensitized/challenged wild-type (WT) mice. This exacerbation was largely Th2-mediated with a significant increase in CD4(+)IL-13(+) T cells and IL-4, IL-5, and IL-13 cytokines. The loss of Stard7 was also associated with increased lung epithelial permeability and activation of proinflammatory dendritic cells in sensitized and/or challenged Stard7 (+/-) mice. Notably, OVA-pulsed dendritic cells from Stard7(+/-) mice were sufficient to confer an exaggerated allergic response in OVA-challenged WT mice, although airway hyperresponsiveness was greater in Stard7(+/-) recipients compared with WT recipients. Enhanced allergic responses in the lung were accompanied by age-dependent development of spontaneous atopic dermatitis. Overall, these data suggest that Stard7 is an important component of a novel protective pathway in tissues exposed to the extracellular environment.

Position 834 in TM6 plays an important role in cholesterol and phosphatidylcholine transport by ABCA1.

ATP-binding cassette protein A1 (ABCA1) plays a key role in eliminating excess cholesterol from peripheral cells by generating nascent high-density lipoprotein (HDL). However, it remains unclear whether both phospholipids and cholesterol are directly loaded onto apolipoprotein A-I (apoA-I) by ABCA1. To identify the amino acid residues of ABCA1 involved in substrate recognition and transport, we applied arginine scan mutagenesis to residues L821-E843 of human ABCA1 and predicted the environment to which each residue is exposed. The relative surface expression of each mutant suggested that residues L821-E843 pass through the plasma membrane as TM6, and the four residues (S826, F830, L834, and V837) of TM6 are exposed to the hydrophilic internal cavity of ABCA1. Furthermore, we showed that L834 is critical for the function of ABCA1.

Pathophysiological analysis of primary biliary cirrhosis focusing on choline/phospholipid metabolism

Injury to biliary epithelial cells caused by disorders in bile composition may be the initial step in the pathogenesis of primary biliary cirrhosis (PBC). We therefore examined choline/phospholipid metabolism in livers of patients with PBC. Hepatic levels of mRNA encoded by choline metabolism-related genes in early stage PBC patients were quantified by real-time RT-PCR. Serum cholesterol and triglyceride concentrations in each lipoprotein compartment and serum/tissue choline levels were also measured. OCT1 expression was quantified by genotype (rs683369 and rs622342). Serum choline concentrations were significantly higher in PBC patients than in normal individuals, with the concentrations in the former lowered by treatment with fibrates. Hepatic choline levels were markedly lower in PBC patients than in controls. The levels of expression of genes associated with choline uptake (OCT1 and CTL1), phosphatidylcholine synthesis (PEMT and BHMT), and phosphatidylcholine transport (MDR3) were significantly upregulated in PBC compared with control livers. Serum cholesterol concentrations and the cholesterol/triglyceride ratio in serum very low density lipoprotein were markedly higher in PBC patients than in controls. In PBC liver, OCT1 protein levels were lower in patients with minor (CG/GG at rs683369 and/or CC at rs622342) than major (CC at rs683369 and AA at rs622342) genotypes of the OCT1 gene. During early stage PBC, hepatocellular choline uptake and PC synthesis become dysregulated. OCT1 genotypes may influence the pathogenesis of PBC.

Expression of MDR1 and MDR3 gene products in paclitaxel-, doxorubicin- and vincristine-resistant cell lines

Multiple drug resistance is one of the main reasons for low chemotherapeutic efficiency in cancer patients. The proteins that are most frequently implicated to play a role in this mechanism are transmembrane proteins that are members of the ABC family. The most important ABC protein is MDR1 (ABCB1), which is expressed in over fifty percent of drug-resistant cancers. The phosphatidylcholine transporter, MDR3 (ABCB4), exhibits high homology with MDR1. An increasing body of evidence suggests that MDR3 plays a role in drug resistance. In the present study, we used doxorubicin-, paclitaxel- and vincristine-resistant cancer cell lines. A chemosensitivity assay MTT test was performed to assess drug resistance. Quantitative real-time polymerase chain reaction analyses were performed to determine the mRNA expression levels of the MDR1 and MDR3 genes. We observed dose-dependent responses to doxorubicin, paclitaxel and vincristine in the investigated cell lines. In all of the drug-resistant cell lines that we studied, we observed increased MDR1 and MDR3 transcript levels. In a doxorubicin-resistant variant of the LoVo cell line (LoVoDx), MDR3 was expressed at higher levels than MDR1. We also observed high correlations between MDR3 expression and resistance to doxorubicin and paclitaxel. Our results suggest that MDR3 plays an active and important role in drug resistance in the investigated cell lines.

Neutral effects of common variation in the hepatocanalicular phosphatidylcholine transporter ABCB4 and the bile salt export pump ABCB11 on genetic cholangiocarcinoma risk

Neutral effects of common variation in the hepatocanalicular phosphatidylcholine transporter ABCB4 and the bile salt export pump ABCB11 on genetic cholangiocarcinoma risk

The hepatic phosphatidylcholine transporter ABCB4 as modulator of glucose homeostasis

The hepatic phosphatidylcholine (PC) transporter ATP-binding cassette (ABC) B4 flops PC from hepatocytes into bile, and its dysfunction causes chronic cholestasis and fibrosis. Because a nuclear receptor-dependent PC pathway has been determined to exert antidiabetic effects, we now analyzed the role of ABCB4 in glucose metabolism. We bred congenic Abcb4-knockout (Abcb4(-/-)) mice on the fibrosis-susceptible BALB/cJ background. Knockout mice and wild-type controls were phenotyped by measuring plasma glucose concentrations, intraperitoneal glucose tolerance, hepatic RNA expression profiles, and liver histology. In addition, 4 procholestatic ABCB4 gene variants were correlated with blood glucose levels in 682 individuals from 2 independent European cohorts. Systemic glucose levels differ significantly between Abcb4(-/-) mice and wild-type controls, and knockout mice display improved glucose tolerance with significantly lower area under the curve values on intraperitoneal glucose challenge. Of note, hepatic expression of the antidiabetic nuclear receptor 5A2 (LRH-1) is induced consistently in Abcb4(-/-) mice, and its specific rare PC ligands are detected in liver by mass spectrometry imaging. In humans, serum glucose levels are associated significantly with the common ABCB4 variant c.711A>T. In summary, ABCB4 might play a critical role in glucose homeostasis in mice and humans. We speculate that the effects could be mediated via LRH-1-dependent PC pathways.

Liver Transplantation for Familial Intra Hepatic Cholestastic Syndrome FICS Type 3

Purpose: To discuss transplantation for patients with FICS. Methods: IRB approval is not required for case reports. We present a case of a 47-year-old male who presented with fulminant hepatic failure and underwent successful liver transplantation. All of the patient's serologies were negative. He had no known exposure to hepatic toxins. Twelve years prior his then 53 year old sister underwent transplantation for a similar indication. She also had no known exposure to hepatotoxins and her serologies were all negative. She succumbed shortly thereafter secondary to overwhelming sepsis. Explant pathology revealed pan lobar necrosis, patchy portal inflammation and mild hepatocytic cholestasis in both siblings. These findings and clinical scenarios are most consistent with MDR3 mutations associated with progressive familial intra hepatic cholestasis. The patient has a remaining sister with no signs of liver disease. Progressive familial intrahepatic cholestastis was first described by Clayton et al. in 1965. It was originally called Byler Disease. There are several distinct phenotypes. It appears that defects affecting the hepatocytes, bile secretions and bile acid metabolism play a major role in pathogenesis. Genetic mutations in hepatocellular transport system genes have been implicated. While type 1 and type 2 FICS usually affect children, FICS type 3 is seen in adults and is caused by mutations in MDR3 gene encoding the biliary phospholipid phosphatidyl choline transporter. Patients with FICS 3 usually exhibit high serum GGTP. Later stage liver histology is consistent with biliary cirrhosis. The liver pathology may result by the toxic effect of the bile acids on bile canaliculi and biliary epithelium. Patients with this syndrome usually present later in life. Treatment usually involves ursodeoxycholic acid. Since our patient (and his sister) presented with fulminant hepatic failure, ursodeoxycholic acid treatment was unsuccessful. We are in the process of performing genetic examinations on the recipient and his remaining sister. If she is MDR3 positive we will likely initiate ursodeoxycholic acid treatment. Conclusion: Familial intrahepatic cholestatic syndrome Type 3 is an uncommon cause of liver failure. Nevertheless, it may have significant ramifications for the health of the entire kindreds. Appropriate screening and intervention are paramount. In the case of liver failure caused by FICS3, liver transplantation appears to be indicated.

Disruption of the Lipid-Transporting LdMT-LdRos3 Complex in Leishmania donovani Affects Membrane Lipid Asymmetry but Not Host Cell Invasion

Maintenance and regulation of the asymmetric lipid distribution across eukaryotic plasma membranes is governed by the concerted action of specific membrane proteins controlling lipid movement across the bilayer. Here, we show that the miltefosine transporter (LdMT), a member of the P4-ATPase subfamily in Leishmania donovani, and the Cdc50-like protein LdRos3 form a stable complex that plays an essential role in maintaining phospholipid asymmetry in the parasite plasma membrane. Loss of either LdMT or LdRos3 abolishes ATP-dependent transport of NBD-labelled phosphatidylethanolamine (PE) and phosphatidylcholine from the outer to the inner plasma membrane leaflet and results in an increased cell surface exposure of endogenous PE. We also find that promastigotes of L. donovani lack any detectable amount of phosphatidylserine (PS) but retain their infectivity in THP-1-derived macrophages. Likewise, infectivity was unchanged for parasites without LdMT-LdRos3 complexes. We conclude that exposure of PS and PE to the exoplasmic leaflet is not crucial for the infectivity of L. donovani promastigotes.

PPARs, RXRs, and Drug‐Metabolizing Enzymes

PPARs, RXRs, and Drug‐Metabolizing Enzymes