Real-world clinical effectiveness and safety of vedolizumab and anti-tumor necrosis factor alpha treatment in ulcerative colitis and Crohn's disease patients: a German retrospective chart review.

BACKGROUND: Real-world comparisons of biologic treatment outcomes for ulcerative colitis (UC) or Crohn's disease (CD) patients are limited. We sought to evaluate the real-world effectiveness of vedolizumab (VDZ) and anti-tumor necrosis factor alpha (anti-TNFα) in UC and CD patients in Germany. METHODS: A retrospective chart review (15 sites) investigated UC and CD patients who were biologic-treatment naïve (biologic-naïve) or had received no more than one prior anti-TNFα before initiating treatment with VDZ or anti-TNFα between 15 July 2014 and 20 October 2015. Kaplan-Meier analyses assessed time to first chart-documented clinical remission (CR) and symptom resolution (UC: rectal bleeding [RB], stool frequency [SF]; CD: abdominal pain [AP], liquid stools [LS]) and outcome duration. RESULTS: A total of 133 UC (76 VDZ; 57 anti-TNFα) and 174 CD (69 VDZ; 105 anti-TNFα) patients were included. By Week 26, estimated cumulative rates of patients achieving CR or symptom resolution with VDZ vs anti-TNFα treatment were for UC: CR, 53.7% vs 31.7%; RB, 66.8% vs 55.8%; and SF, 59.8% vs 50.7%, respectively; and for CD: CR, 14.4% vs 32.8%; AP, 62.5% vs 56.0%; and LS, 29.9% vs 50.3%, respectively. Outcomes were sustained similarly between treatments, except RB (VDZ vs anti-TNFα: median 38.1 vs 15.1 weeks, P = 0.03). Treatment-related adverse events occurred in 5.3% vs 7.0% (UC) and 8.7% vs 19.0% (CD) of VDZ vs anti-TNFα patients, respectively. CONCLUSIONS: Although there were differences in CR, symptom resolution, and safety profiles, real-world data support both VDZ and anti-TNFα as effective treatment options in UC and CD.

Perceived distress, personality characteristics, coping strategies and psychosocial impairments in a national German multicenter cohort of patients with Crohn's disease and ulcerative colitis.

BACKGROUND AND AIMS: This study examined differences in personality, psychological distress, and stress coping in inflammatory bowel disease (IBD) depending on type of disease and disease activity. We compared patients suffering from Crohn's disease (CD) and ulcerative colitis (UC) with controls. While the literature is replete with distinctive features of the pathogenesis of IBD, the specific differences in psychological impairments are not well studied. METHODS: In this German national multicenter study, participants were recruited from 32 centers. Two hundred ninety-seven questionnaires were included, delivering vast information on disease status and psychological well-being based on validated instruments with a total of 285 variables. RESULTS: CD patients were more affected by psychological impairments than patients suffering from UC or controls. Importantly, patients with active CD scored higher in neuroticism (p < 0.01), psychological distress (p < 0.001) and maladaptive stress coping (escape, p = 0.03; rumination, p < 0.03), but less need for social support (p = 0.001) than controls. In contrast, patients suffering from active UC showed psychological distress (p < 0.04) and maladaptive coping (avoidance, p < 0.03; escape, p = 0.01). Patients in remission seemed to be less affected. In particular, patients with UC in remission were not inflicted by psychological impairments. The group of CD patients in remission however, showed insecurity (p < 0.01) and paranoid ideation (p = 0.04). CONCLUSIONS: We identified specific aspects of psychological impairment in IBD depending on disease and disease activity. Our results underscore the need for psychological support and treatment particularly in active CD.

The metabolic capacity of lipid droplet localized acyl-CoA synthetase 3 is not sufficient to support local triglyceride synthesis independent of the endoplasmic reticulum in A431 cells.

ACSL3 is the only long chain fatty acyl-CoA synthetase consistently found on growing and mature lipid droplets (LDs), suggesting that this specific localization has biological relevance. Current models for LD growth propose that triglycerides are synthesized by enzymes at the LD surface, with activated fatty acids provided by LD localized ACSL3, thus allowing growth independent of the ER. Here, we tested this hypothesis by quantifying ACSL3 on LDs from human A431 cells. RNAi of ACSL3 reduced the oleoyl-CoA synthetase activity by 83%, suggesting that ACSL3 is by far the dominant enzyme of A431 cells. Molar quantification revealed that there are 1.4 million ACSL3 molecules within a single cell. Metabolic labeling indicated that each ACSL3 molecule contributed a net gain of 3.1 oleoyl-CoA/s. 3D reconstruction of confocal images demonstrated that 530 individual lipid droplets were present in an average oleate fed A431 cell. A representative single lipid droplet with a diameter of 0.66 µm contained 680 ACSL3 molecules on the surface. Subcellular fractionation showed that at least 68% of ACSL3 remain at the ER even during extensive fatty acid supplementation. High resolution single molecule microscopy confirmed the abundance of cytoplasmic ACSL3 outside of LDs. Model calculations for triglyceride synthesis using only LD localized ACSL3 gave significant slower growth of LDs as observed experimentally. In conclusion, although ACSL3 is an abundant enzyme on A431 LDs, the metabolic capacity is not sufficient to account for LD growth solely by the local synthesis of triglycerides.

Differentially localized acyl-CoA synthetase 4 isoenzymes mediate the metabolic channeling of fatty acids towards phosphatidylinositol.

The acyl-CoA synthetase 4 (ACSL4) has been implicated in carcinogenesis and neuronal development. Acyl-CoA synthetases are essential enzymes of lipid metabolism, and ACSL4 is distinguished by its preference for arachidonic acid. Two human ACSL4 isoforms arising from differential splicing were analyzed by ectopic expression in COS cells. We found that the ACSL4_v1 variant localized to the inner side of the plasma membrane including microvilli, and was also present in the cytosol. ACSL4_v2 contains an additional N-terminal hydrophobic region; this isoform was located at the endoplasmic reticulum and on lipid droplets. A third isoform was designed de novo by appending a mitochondrial targeting signal. All three ACSL4 variants showed the same specific enzyme activity. Overexpression of the isoenzymes increased cellular uptake of arachidonate to the same degree, indicating that the metabolic trapping of fatty acids is independent of the subcellular localization. Remarkably, phospholipid metabolism was changed by ACSL4 expression. Labeling with arachidonate showed that the amount of newly synthesized phosphatidylinositol was increased by all three ACSL4 isoenzymes but not by ACSL1. This was dependent on the expression level and the localization of the ACSL4 isoform. We conclude that in our model system exogenous fatty acids are channeled preferentially towards phosphatidylinositol by ACSL4 overexpression. The differential localization of the endogenous isoenzymes may provide compartment specific precursors of this anionic phospholipid important for many signaling processes.

Protein mediated fatty acid uptake: synergy between CD36/FAT-facilitated transport and acyl-CoA synthetase-driven metabolism.

The mechanism of cellular fatty acid uptake is highly relevant for basic and clinical research. Previous work has demonstrated that fatty acid uptake is facilitated by cell surface membrane proteins as well as by intracellularly localized enzymes. Here, the exogenous expression of the CD36/FAT glycoprotein and the acyl-CoA synthetases FATP4 and ACSL1 in MDCK cells was quantified by comparison to recombinant proteins, and related to the corresponding increases of fatty acid uptake. At the molecular level, CD36/FAT was 30-fold more efficient than either FATP4 or ACSL1 in enhancing fatty acid uptake. Remarkably, co-expression of CD36/FAT with FATP4 or ACSL1 led to a higher increase of fatty acid uptake than expected from the combined individual contributions, whereas co-expression of FATP4 and ACSL1 did not. Immunofluorescence microscopy confirmed the plasma membrane localization of CD36/FAT and the intracellular localization of FATP4 to the endoplasmic reticulum, and of ACSL1 to mitochondria. Concluding, we suggest that fatty acid uptake in our model system is organized by two spatially distinct but synergistic mechanisms: the cell surface protein CD36/FAT directly facilitates fatty acid transport across the plasma membrane, whereas the intracellular acyl-CoA synthetases FATP4 and ACSL1 enhance fatty acid uptake indirectly by metabolic trapping.

CYP3A activity in severe liver cirrhosis correlates with Child-Pugh and model for end-stage liver disease (MELD) scores.

AIMS: Impaired liver function often necessitates drug dose adjustment to avoid excessive drug accumulation and adverse events, but a marker for the extent of the required adjustment is lacking. The aim of this study was to investigate whether Child-Pugh (CP) and model for end-stage liver disease (MELD) scores correlate with drug clearance. METHODS: Midazolam was used as a CYP3A probe and its pharmacokinetics were analyzed in 24 patients with mild to severe liver cirrhosis (n = 4, 10 and 10 with CP class A, B and C, respectively) and six patients without liver disease. RESULTS: Both scores correlated well with unbound midazolam clearance (CLu ), unbound midazolam fraction and half-life (all P < 0.01), whereas the unbound steady-state volume of distribution was not significantly changed. In patients with severe liver cirrhosis unbound midazolam clearance was only 14% of controls (CP C: CLu = 843 ± 346 l h(-1), MELD ≥ 15: CLu = 805 ± 474 l h(-1), controls: CLu = 5815 ± 2649 l h(-1), P < 0.01). CONCLUSION: The correlation with unbound midazolam clearance suggests that either score predicts the metabolic capacity of CYP3A, the most relevant drug metabolizing enzyme subfamily in humans.

Flotillin-2 expression in the human gut: from a cell model to human tissue in health and inflammatory bowel diseases.

BACKGROUND AND AIMS: The etiopathogenesis of inflammatory bowel diseases (IBD) remains largely unexplained. Flotillins (flotillin-1 and flotillin-2) are ubiquitous proteins which have been linked to inflammation and regeneration. We hypothesized that alterations in the expression of flotillin-2 in enterocytes may be related to the pathogenesis of IBD as a classical example of an inflammatory disorder of mostly unknown origin. METHODS: Cell and tissue localization of flotillin-2 (and -1) were investigated by immunofluorescent staining in 1. polarized and unpolarized CaCo-2w cells as a model of human enterocytes (native and after TNFα stimulation) and 2. intestinal biopsies from controls, patients with ulcerative colitis (UC) and patients with Crohn's disease (CD). For quantification of flotillin-2, we analyzed its expression in ileal and colonic biopsies from controls, UC patients and CD patients using real-time RT-PCR, Western blot and indirect immunofluorescence. RESULTS: In polarized CaCo-2w cells and human enterocytes in biopsies, flotillins were localized at the basolateral membrane and on subapical vesicles, but not in the apical membrane. Flotillin-2 expression did not differ between UC patients, CD patients and controls. However, it was significantly higher in colonic biopsies compared to ileal biopsies in all groups. CONCLUSIONS: By virtue of its abundant expression in enterocytes, flotillin-2 must have an essential function in intestinal physiology, especially in the colon. Yet our data could not link flotillin-2 to the pathogenesis of IBD.

Real-World Effectiveness of Vedolizumab vs Anti-TNF in Biologic-naïve Crohn's Disease Patients: A 2-year Propensity-score-adjusted Analysis from the VEDOIBD-Study.

BACKGROUND: The aim of this observational, real-world evidence, modified intention-to-treat (mITT) study based on prospectively collected data from the VEDOIBD registry was to compare the effectiveness of vedolizumab (VEDO) vs antitumor necrosis factor (anti-TNF) in biologic-naïve Crohn's disease (CD) patients. METHODS: Between 2017 and 2020, 557 CD patients starting therapy with VEDO or anti-TNF were consecutively enrolled in 45 IBD centers across Germany. Per study protocol, the analysis excluded biologic-experienced patients and those with a missing Harvey-Bradshaw Index score, resulting in a final sample of 327 biologic-naïve CD patients. Clinical remission was measured using the Harvey-Bradshaw Index at the end of induction therapy and after 1 and 2 years. Switching to a different therapy was considered an outcome failure. Propensity score adjustment with inverse probability of treatment weighting was used to correct for confounding. RESULTS: The effectiveness of both VEDO (n = 86) and anti-TNF (n = 241) was remarkably high for induction treatment, but VEDO performed significantly less well than anti-TNF (clinical remission: 56.3% vs 73.9%, P < .05). In contrast, clinical remission after 2 years was significantly better for VEDO compared with anti-TNF (74.2% vs 44.7%; P < .05; odds ratio, 0.45; 95% CI, 0.22-0.94). Remarkably, only 17% of patients switched from VEDO to another biologic vs 44% who received anti-TNF. CONCLUSIONS: The results of this prospective, 2-year, real-world evidence study suggest that the choice of VEDO led to higher remission rates after 2 years compared with anti-TNF. This could support the role of VEDO as a first-line biologic therapy in CD.

Real-world effectiveness of vedolizumab compared to anti-TNF agents in biologic-naïve patients with ulcerative colitis: A two-year propensity-score-adjusted analysis from the prospective, observational VEDOIBD -study.

BACKGROUND: This observational real-world evidence (RWE) study is based on prospectively collected data from the VEDOIBD registry study. AIM: To compare the effectiveness of vedolizumab and anti-TNF agents in biologic-naïve patients with ulcerative colitis (UC) at the end of induction and during maintenance treatment. METHODS: Between 2017 and 2020, we enrolled 512 patients with UC starting therapy with vedolizumab or an anti-TNF agent in 45 IBD centres across Germany. We excluded biologic-experienced patients and those with missing partial Mayo (pMayo) outcomes; this resulted in a final sample of 314 (182 on vedolizumab and 132 on an anti-TNF agent). The primary outcome was clinical remission measured using pMayo score; any switch to a different biologic agent was considered an outcome failure (modified ITT analysis). We used propensity score adjustment with inverse probability of treatment weighting to correct for confounding. RESULTS: During induction therapy, clinical remission was relatively low and similar in vedolizumab- and anti-TNF-treated patients (23% vs. 30.4%, p = 0.204). However, clinical remission rates after two years were significantly higher for vedolizumab-treated patients than those treated with an anti-TNF agent (43.2% vs. 25.8%, p < 0.011). Among patients treated with vedolzumab, 29% switched to other biologics, versus 54% who had received an anti-TNF agent. CONCLUSION: After two years of treatment, vedolizumab resulted in higher remission rates than anti-TNF agents.

The N-terminal region of acyl-CoA synthetase 3 is essential for both the localization on lipid droplets and the function in fatty acid uptake.

Cytosolic lipid droplets (LDs) are storage organelles for neutral lipids derived from endogenous metabolism. Acyl-CoA synthetase family proteins are essential enzymes in this biosynthetic pathway, contributing activated fatty acids. Fluorescence microscopy showed that ACSL3 is localized to the endoplasmic reticulum (ER) and LDs, with the distribution dependent on the cell type and the supply of fatty acids. The N-terminus of ACSL3 was necessary and sufficient for targeting reporter proteins correctly, as demonstrated by subcellular fractionation and confocal microscopy. The N-terminal region of ACSL3 was also found to be functionally required for the enzyme activity. Selective permeabilization and in silico analysis suggest that ACSL3 assumes a hairpin membrane topology, with the N-terminal hydrophobic amino acids forming an amphipathic helix restricted to the cytosolic leaflet of the ER membrane. ACSL3 was effectively translocated from the ER to nascent LDs when neutral lipid synthesis was stimulated by the external addition of fatty acids. Cellular fatty acid uptake was increased by overexpression and reduced by RNA interference of ACSL3. In conclusion, the structural organization of ACSL3 allows the fast and efficient movement from the ER to emerging LDs. ACSL3 not only esterifies fatty acids with CoA but is also involved in the cellular uptake of fatty acids, presumably indirectly by metabolic trapping. The unique localization of the acyl-CoA synthetase ACSL3 on LDs suggests a function in the local synthesis of lipids.

Mucosal protection by phosphatidylcholine.

The colonic mucus serves a first barrier towards invasion of commensal bacteria in stools to prevent inflammation. One essential component of intestinal mucus is phosphatidylcholine (PC) which represents more than 90% of the phospholipids in mucus indicative for a selective transport of PC into this compartment. It is arranged in lamellar structures as surfactant-like particles which provide a hydrophobic surface on top of the hydrated mucus gel to prevent the invasion of bacteria from intestinal lumen. In ulcerative colitis (UC), the mucus PC content is reduced by 70%, irrespective of the state of inflammation. Thus, it could represent an intrinsic primary pathogenetic condition predisposing to bacterial invasion and the precipitation of inflammation. Since PC was shown to be mainly secreted by the ileal mucosa from where it is assumed to move distally to the colon, the PC content along the colonic wall towards the rectum gradually thins, with the least PC content in the rectum. This explains the start of the clinical manifestation of UC in the rectum and the expansion from there to the upper parts of the colon. In three clinical trials, when missing mucus PC in UC was supplemented by an oral, delayed release PC preparation, the inflammation improved and even resolved after a 3-month treatment course. The data indicate the essential role of the mucus PC content for protection against inflammation in colon.

Phosphatidylcholine as a constituent in the colonic mucosal barrier--physiological and clinical relevance.

Phosphatidylcholine (PC) is an important constituent of the gastrointestinal tract. PC molecules are not only important in intestinal cell membranes but also receiving increasing attention as protective agents in the gastrointestinal barrier. They are largely responsible for establishing the hydrophobic surface of the colon. Decreased phospholipids in colonic mucus could be linked to the pathogenesis of ulcerative colitis, a chronic inflammatory bowel disease. Clinical studies revealed that therapeutic addition of PC to the colonic mucus of these patients alleviated the inflammatory activity. This positive role is still elusive, however, we hypothesized that luminal PC has two possible functions: first, it is essential for surface hydrophobicity, and second, it is integrated into the plasma membrane of enterocytes and it modulates the signaling state of the mucosa. The membrane structure and lipid composition of cells is a regulatory component of the inflammatory signaling pathways. In this perspective, we will shortly summarize what is known about the localization and protective properties of PC in the colonic mucosa before turning to its evident medical importance. We will discuss how PC contributes to our understanding of the pathogenesis of ulcerative colitis and how reinforcing the luminal phospholipid monolayer can be used as a therapeutic concept in humans.

FATP4 contributes as an enzyme to the basal and insulin-mediated fatty acid uptake of C2C12 muscle cells.

The function of membrane proteins in long-chain fatty acid transport is controversial. The acyl-CoA synthetase fatty acid transport protein-4 (FATP4) has been suggested to facilitate fatty acid uptake indirectly by its enzymatic activity, or directly by transport across the plasma membrane. Here, we investigated the function of FATP4 in basal and insulin mediated fatty acid uptake in C(2)C(12) muscle cells, a model system relevant for fatty acid metabolism. Stable expression of exogenous FATP4 resulted in a twofold higher fatty acyl-CoA synthetase activity, and cellular uptake of oleate was enhanced similarly. Kinetic analysis demonstrated that FATP4 allowed the cells to reach apparent saturation of fatty acid uptake at a twofold higher level compared with control. Short-term treatment with insulin increased fatty acid uptake in line with previous reports. Surprisingly, insulin increased the acyl-CoA synthetase activity of C(2)C(12) cells within minutes. This effect was sensitive to inhibition of insulin signaling by wortmannin. Affinity purified FATP4 prepared from insulin-treated cells showed an enhanced enzyme activity, suggesting it constitutes a novel target of short-term metabolic regulation by insulin. This offers a new mechanistic explanation for the concomitantly observed enhanced fatty acid uptake. FATP4 was colocalized to the endoplasmic reticulum by double immunofluorescence and subcellular fractionation, clearly distinct from the plasma membrane. Importantly, neither differentiation into myotubes nor insulin treatment changed the localization of FATP4. We conclude that FATP4 functions by its intrinsic enzymatic activity. This is in line with the concept that intracellular metabolism plays a significant role in cellular fatty acid uptake.

Morphologic patterns of autoimmune pancreatitis in CT and MRI.

BACKGROUND/AIMS: To retrospectively evaluate the morphologic characteristics of autoimmune pancreatitis (AIP) using MRI and CT. METHODS: 86 dynamic contrast-enhanced CT and MRI scans in 36 AIP patients were evaluated regarding: different enlargement types, abnormalities of the main pancreatic duct (MPD), morphology of the parenchyma and other associated findings. RESULTS: 3 types of enlargement were found: (1) a focal type (28%), (2) a diffuse type (involving the entire pancreas, 11%) and (3) a combined type (56%). The MPD was usually dilated together with focal or diffuse narrowing in 67% (24/36). Unenhanced MRI showed AIP area in 56% (mostly T(1) hypo- and T(2) hyperattenuating), and CT in 10% (hypoattenuating). The arterial phase depicted similar patterns for CT and MRI (hypoattenuating in 58 and 52%, respectively). Venous and late venous phase patterns were usually hyperattenuating in MRI (65 and 74%, late enhancement), while CT mostly showed no signal differences (isoattenuating in 57 and 75%), yielding significant differences between CT and MRI for the venous (p < 0.0001) and the late phase (p = 0.025). Miscellaneous findings were: rim sign (25%), pseudocysts (8%) and infiltration of large vessels (11%). CONCLUSIONS: The 'late-enhancement' sign seems to be a key feature and is best detectable with MRI. MRI may be recommended in the diagnostic workup of AIP patients. and IAP.

Loss of EpCAM expression in breast cancer derived serum exosomes: role of proteolytic cleavage.

OBJECTIVE: Cancer cells in the body release soluble and membranous factors that manipulate the tumor environment to facilitate growth and survival. Recent years have provided evidence that small microvesicles that are termed exosomes may play a pivotal role in this process. Exosomes are membrane vesicles with a size of 40-100 nm that are released by both tumor and normal cells and can be found in various body fluids. Tumor-derived exosomes carry functional proteins, mRNAs, and miRNAs and could serve as novel platform for tumor diagnosis and prognosis. However, marker proteins that allow enrichment of tumor-derived exosomes over normal exosomes are less well defined. METHODS: We used Western blot analysis and antibody coupled magnetic beads to characterize CD24 and EpCAM as markers for exosomes. We investigated ovarian carcinoma ascites, pleural effusions and serum of breast carcinoma patients. As non-tumor derived control we used exosomes from ascites of liver cirrhosis patients. RESULTS: Exosomes could be isolated from all body fluids and contained marker proteins as well as miRNAs. We observed that CD24 and EpCAM were selectively present on ascites exosomes of tumor patients and copurified together on anti-EpCAM or anti-CD24 magnetic beads. In breast cancer patients CD24 was present but EpCAM was absent from serum exosomes. Instead, the intact EpCAM ectodomain was recovered in a soluble form. We provide evidence that EpCAM can be cleaved from exosomes via serum metalloproteinase(s). CONCLUSION: Loss of EpCAM on serum exosomes may hamper enrichment by immune-affinity isolation. We suggest that CD24 could be an additional marker for the enrichment of tumor-derived exosomes from blood.

Reduced hydrophobicity of the colonic mucosal surface in ulcerative colitis as a hint at a physicochemical barrier defect.

PURPOSE: There is increasing evidence that a defect of the gastrointestinal mucosal barrier is important for the development of inflammatory bowel diseases (IBD). The hydrophobicity of the colonic mucosal surface is a measure of its resistance to luminal antigens, e.g. of bacterial origin. Therefore, the purpose of this study was to determine this parameter in patients suffering from IBD. METHODS: Nineteen patients with ulcerative colitis (UC), ten patients with Crohn's disease (CD) and 20 controls were examined. All underwent colonic surgery at the University Hospital Heidelberg. Clinical disease activity was determined. From every subject, colonic tissue specimens were obtained, and hydrophobicity of the mucosal surface was determined with a goniometer by multiple plateau contact angle measurements. Histological evaluation of disease activity was performed in directly adjacent tissue specimens. RESULTS: Hydrophobicity of the colonic mucosal surface, expressed as plateau contact angles, was significantly reduced in patients with UC (mean ± SEM, 47.8° ± 3.4°) compared to those with CD (72.0° ± 5.2°) and controls (72.5° ± 5.6°; over-all P = 0.0004; UC versus controls, P < 0.001; UC versus CD, P < 0.05; CD versus controls, P > 0.05). Between mucosal hydrophobicity and clinical disease activity, as well as mucosal hydrophobicity and histological disease activity, no significant correlation was found. CONCLUSIONS: The results suggest a defective physicochemical barrier as an essential factor in the pathogenesis of UC, but not CD. The fact that no correlation was found between mucosal hydrophobicity and disease activity may indicate that the loss of mucosal hydrophobicity in UC is not exclusively a secondary effect due to inflammation.

Overexpression of CD36 and acyl-CoA synthetases FATP2, FATP4 and ACSL1 increases fatty acid uptake in human hepatoma cells.

BACKGROUND: Understanding the mechanisms of long chain fatty acid (LCFA) uptake in hepatic cells is of high medical importance to treat and to prevent fatty liver disease (FLD). ACSs (Acyl-CoA synthetases) are a family of enzymes that catalyze the esterification of fatty acids (FA) with CoA. Recent studies suggest that ACS enzymes drive the uptake of LCFA indirectly by their enzymatic activity and could promote special metabolic pathways dependent on their localization.The only protein located at the plasma membrane which has consistently been shown to enhance FA uptake is CD36. AIMS: The current study investigated whether ACSs and CD36 could regulate hepatic LCFA uptake. METHODS AND RESULTS: FATP2 and FATP4 were both localized to the ER of HuH7 and HepG2 cells as shown by double immunofluorescence in comparison to marker proteins. ACSL1 was located at mitochondria in both cell lines. Overexpression of FATP2, FATP4 and ACSL1 highly increased ACS activity as well as the uptake of [3H]-oleic acid and fluorescent Bodipy-C12 (B12) fatty acid. Quantitative FACS analysis showed a correlation between ACS expression levels and B12 uptake. FATP2 had the highest effect on B12 uptake of all proteins tested. CD36 was mainly localized at the plasma membrane. Whereas [3H]-oleic acid uptake was increased after overexpression, CD36 had no effect on B12 uptake. CONCLUSION: Uptake of LCFA into hepatoma cells can be regulated by the expression levels of intracellular enzymes. We propose that ACS enzymes drive FA uptake indirectly by esterification. Therefore these molecules are potential targets for treatment of nonalcoholic fatty liver disease (NAFLD) or steatohepatitis (NASH).

Delayed release phosphatidylcholine in chronic-active ulcerative colitis: a randomized, double-blinded, dose finding study.

BACKGROUND: In 2 preceding studies, delayed release phosphatidylcholine (rPC) was found to (a) improve disease activity and (b) withdraw steroids in patients with chronic-active ulcerative colitis. GOAL: Objective of the study was to determine the most effective rPC dose with least adverse events. STUDY: A randomized, dose-controlled, double-blinded study. Four groups of 10 patients each with nonsteroid-treated, chronic-active ulcerative pancolitis with a clinical activity index (CAI) and endoscopic activity index (EAI) >or=7. Patients were treated with oral rPC at doses of 0.5, 1, 3, and 4 g daily over 12 weeks. RESULTS: The CAI changes from baseline to the end of the study were 2.5 (0.5 g), 7.0 (1 g), 5.5 (3 g), and 6.0 (4 g dose arm). Significant improvement of the CAI was registered between the lowest rPC dose of 0.5 g (control group) and all higher doses of 1.0, 3.0, and 4.0-g rPC (Por=50% CAI improvement) were 70% in all of the effective dose groups (1 to 4 g, P=0.003). This was paralleled by the EAI improvement and by the rates of mucosal healing. Median time to clinical response was 5 (IQR 2 to 8) weeks. Bloating was registered in 40% of the patients irrespective of the treatment dose. Three of the 10 patients in the 4 g dose group reported nausea. CONCLUSION: We found a saturable dose response of rPC in the treatment of chronic-active ulcerative colitis with effective doses >or=1 g per day; doses of 3 and 4 g seem to be superior in achieving remission.

Phosphatidylcholine (lecithin) and the mucus layer: Evidence of therapeutic efficacy in ulcerative colitis?

Colonic mucus protects against attacks from bacteria in stool. One component of mucus is phosphatidylcholine (PC) which is thought to be arranged as continuous lamellar layer in the apical mucus and to be responsible for establishing a protective hydrophobic surface. This 'intestinal surfactant' plays a key role in mucosal defense. Thus, a defective PC layer contributes to the development of inflammation. Analysis of rectoscopically acquired mucus aliquots revealed a 70% decrease in PC content in ulcerative colitis (UC) compared to Crohn's disease (CD) and healthy controls - independent of disease activity. Accordingly, we propose that lack of mucus PC is a key pathogenetic factor in UC. In clinical studies a delayed-release oral PC preparation (rPC) was found to substitute the lack of PC in rectal mucus. Indeed, in non-steroid-treated active UC, 53% of rPC patients reached remission [clinical activity index (CAI) ≤3] compared to 10% of placebo patients (p ≤ 0.001). Endoscopic and histologic findings improved concomitantly. A second trial with 60 chronic-active, steroid-dependent UC patients was conducted to test for steroid-sparing effects. Complete steroid withdrawal with a concomitant achievement of remission (CAI ≤3) or clinical response (≥50% CAI improvement) was reached in 15 PC-treated patients (50%) but only in 3 (10%) placebo patients (p = 0.002). In conclusion, intrinsic reduction of PC (lecithin) in colonic mucus may be a key pathogenetic feature of UC. Topical supplement of PC by a delayed-released oral PC preparation is effective in resolving inflammatory activity of UC and may develop to a first-choice therapy for this disease.