Effect of one-year vitamin C- and E-supplementation on cerebrospinal fluid oxidation parameters and clinical course in Alzheimer's disease.

Antioxidant vitamins are being widely discussed as a therapeutic option in Alzheimer's disease (AD). We recently found that supplementation with vitamin C and E over 1 month leads to an increase of their levels in cerebrospinal fluid (CSF) and a reduction of CSF lipid peroxidation. In the present study, we followed-up the biochemical and clinical effect of vitamin C and E supplementation in an open clinical trial over 1 year. Twelve AD patients stably taking a cholinesterase inhibitor were supplemented with vitamin C (1,000 mg/day) and E (400 I.U./day), while 11 patients taking cholinergic medication only served as a control group. Cognition was assessed at baseline, after 6 months and 12 months using the Mini-Mental State Examination; a more detailed testing of cognitive function was performed at baseline and after 12 months. From eight of the vitamin-supplemented patients, CSF was taken at baseline, after 1 month and after 1 year to measure the antioxidant effect of vitamin supplementation on CSF lipids using a recently established in vitro oxidation assay. CSF antioxidant vitamins were significantly increased after 1 month and 1 year of supplementation, while in vitro oxidation of CSF lipids was significantly reduced only after 1 year of the supplementation. The clinical course of AD did not significantly differ between the vitamin and the control group. We conclude that supplementation with vitamins E and C did not have a significant effect on the course of AD over 1 year despite of a limited antioxidant effect that could be observed in CSF.

Plasma triglycerides after oral glucose load specifically associate with metabolic risk markers in healthy type 2 diabetes offspring.

OBJECTIVE: To assess the potential of plasma triglycerides measured after glucose load as biomarker for insulin resistance and cardiovascular risk. METHODS: An oral glucose tolerance test (OGTT, n=91) was performed in healthy type 2 diabetes offspring. Plasma lipids, lipoproteins, glucose and hormones were quantified in fasting and post-challenge samples. RESULTS: During the OGTT total plasma triglycerides decreased in most subjects, however, they increased in some individuals and this increase was strongly associated with metabolic risk factors. Subjects with increasing triglycerides (n=18) were more obese and insulin resistant than those with the most pronounced triglyceride decrease (n=18), as indicated by higher HOMA-IR, BMI and waist circumference. Correlation analysis (n=91) demonstrated that the changes of total plasma and VLDL-associated triglycerides between 0 h and 2 h (Δ-TG, Δ-VLDL-T) were strongly associated with risk factors. Δ-TG, and especially Δ-VLDL-T, correlated better than fasting triglycerides with waist circumference, waist-to-hip ratio and fasting glucose. The correlations remained significant after adjustment for gender, age and HDL cholesterol. CONCLUSION: The observed increase of triglycerides after glucose load in subjects with signs of insulin resistance and obesity suggests that post-glucose triglyceride change is a potential novel biomarker for early detection of metabolic risk. The specific association of post-glucose triglyceride change with abdominal obesity and fasting glucose suggests a link to hepatic steatosis and insulin resistance.

Research integrity and publication ethics.

The basic principle for professional conduct of science in all countries and all disciplines is honesty towards oneself and towards others. Therefore it is utmost important that the scientific community prevents scientific misconduct by fostering research integrity. This commentary reports on the experience of a German 'Ombudsman' and relates it to the international concepts of good scientific practice as well as the questions of publication ethics. Biomedical research seems to be most susceptible for scientific misconduct since internationally we see many of the cases in this field. Here possible explanations for the observed misconduct are discussed as well as ways to prevent it. The intention is to both alert scientists and ultimately to adjust the scientific system in a way which allows the next generation of scientists to develop their careers in true research integrity.

A highly effective, nontoxic T1 MR contrast agent based on ultrasmall PEGylated iron oxide nanoparticles.

In this study we systematically developed a potential MR T(1) contrast agent based on very small PEGylated iron oxide nanoparticles. We adjusted the size of the crystalline core providing suitable relaxometric properties. In addition, a dense and optimized PEG coating provides high stability under physiological conditions together with low cytotoxicity and low nonspecific phagocytosis into macrophage cells as a part of the reticulo endothelial system at biologically relevant concentrations. The as developed contrast agent has the lowest r(2)/r(1) ratio (2.4) at 1.41 T reported so far for PEGylated iron oxide nanoparticles as well as a r(1) relaxivity (7.3 mM(-1) s(-1)) that is two times higher compared to that of Magnevist as a typical T(1) contrast agent based on gadolinium as a clinical standard.

Real-time magnetic resonance imaging and quantification of lipoprotein metabolism in vivo using nanocrystals.

Semiconductor quantum dots and superparamagnetic iron oxide nanocrystals have physical properties that are well suited for biomedical imaging. Previously, we have shown that iron oxide nanocrystals embedded within the lipid core of micelles show optimized characteristics for quantitative imaging. Here, we embed quantum dots and superparamagnetic iron oxide nanocrystals in the core of lipoproteins--micelles that transport lipids and other hydrophobic substances in the blood--and show that it is possible to image and quantify the kinetics of lipoprotein metabolism in vivo using fluorescence and dynamic magnetic resonance imaging. The lipoproteins were taken up by liver cells in wild-type mice and displayed defective clearance in knock-out mice lacking a lipoprotein receptor or its ligand, indicating that the nanocrystals did not influence the specificity of the metabolic process. Using this strategy it is possible to study the clearance of lipoproteins in metabolic disorders and to improve the contrast in clinical imaging.

Insulin stimulates hepatic low density lipoprotein receptor-related protein 1 (LRP1) to increase postprandial lipoprotein clearance.

BACKGROUND: While the role of insulin in glucose uptake and its aberration in diabetes are well established, the effect of insulin on lipoprotein clearance in the postprandial phase is not yet fully understood. The dietary lipids are carried in chylomicron remnants (CR) which are taken up into the liver mainly via LDLR-related protein 1 (LRP1). In this study, the effect of insulin on LRP1-mediated hepatic CR uptake was investigated. METHODS: The study was based on determining the subcellular localisation of LRP1 by subcellular fractionation and immunofluorescence microscopy and correlating those findings with the hepatic uptake of fluorescently or radioactively labelled LRP1-specific ligands and CR in hepatoma cells, primary hepatocytes and mouse models. RESULTS AND CONCLUSION: In vitro and in vivo, insulin stimulated the translocation of hepatic LRP1 from intracellular vesicles to the plasma membrane, which correlates with an increased uptake of LRP1-specific ligands. In wild-type mice, a glucose-induced insulin response increased the hepatic uptake of LRP1 ligands while in leptin-deficient obese mice (ob/ob), which are characterised by hepatic insulin resistance, insulin-inducible LRP1 ligand uptake was abolished. Finally, upon hepatic LRP1 knockdown, insulin no longer significantly enhanced CR uptake into the liver. The insulin-induced LRP1-mediated CR uptake, as demonstrated here, suggests that impaired hepatic LRP1 translocation can contribute to the postprandial lipaemia in insulin resistance.

Liver TAG transiently decreases while PL n-3 and n-6 fatty acids are persistently elevated in insulin resistant mice.

Changes in fatty acid metabolism associated with insulin resistance have been described in rats and humans but have not been well characterized in the frequently used mouse model of diet-induced obesity. To analyse the early phase as well as established insulin resistance, C57BL/6 mice were placed for 1 or 16 weeks on a high fat diet (1w-HFD, 16w-HFD). Endocrine and metabolic parameters indicated that 1w-HFD mice showed a moderate but significant induction of insulin resistance while 16w-HFD mice exhibited profound obesity-associated insulin resistance and dyslipidemias. Significant alterations in fatty acid composition were observed in plasma and liver in both groups. Liver phospholipid-associated arachidonate and docosahexaenoate were increased in both 1w-HFD and 16w-HFD mice, possibly due to increased expression of the desaturases Fads1 and Fads2. Unexpectedly, SCD1 activity and gene expression in liver were decreased in the 1w-HFD group accompanied by diminished total hepatic lipid levels, while they were increased in chronically fed mice. Our data indicate that the early phase of HFD-induced insulin resistance is not associated with elevated liver lipid concentration. Furthermore, the early and persistent rise of arachidonate and docosahexaenoate indicates that insulin resistance is not due to insufficient availability (or concentrations) of polyunsaturated fatty acids as postulated previously.

Acute-phase serum amyloid A as a marker of insulin resistance in mice.

Acute-phase serum amyloid A (A-SAA) was shown recently to correlate with obesity and insulin resistance in humans. However, the mechanisms linking obesity-associated inflammation and elevated plasma A-SAA to insulin resistance are poorly understood. Using high-fat diet- (HFD-) fed mice, we found that plasma A-SAA was increased early upon HFD feeding and was tightly associated with systemic insulin resistance. Plasma A-SAA elevation was due to induction of Saa1 and Saa2 expression in liver but not in adipose tissue. In adipose tissue Saa3 was the predominant isoform and the earliest inflammatory marker induced, suggesting it is important for initiation of adipose tissue inflammation. To assess the potential impact of A-SAA on adipose tissue insulin resistance, we treated 3T3-L1 adipocytes with recombinant A-SAA. Intriguingly, physiological levels of A-SAA caused alterations in gene expression closely resembling those observed in HFD-fed mice. Proinflammatory genes (Ccl2, Saa3) were induced while genes critical for insulin sensitivity (Irs1, Adipoq, Glut4) were down-regulated. Our data identify HFD-fed mice as a suitable model to study A-SAA as a biomarker and a novel possible mediator of insulin resistance.

Recycling of apolipoprotein E is not associated with cholesterol efflux in neuronal cells.

After receptor-mediated endocytosis of apolipoprotein E (apoE)-containing lipoproteins in hepatocytes, the isoform apoE3 is efficiently recycled in a process which is associated with cholesterol efflux. Recycling and cholesterol efflux are greatly reduced when apoE4 is the only isoform present. ApoE is the main apolipoprotein in cerebrospinal fluid, and it plays a pivotal role in maintaining cholesterol homeostasis in the brain. The isoform apoE4 is associated with an increased risk of Alzheimer's disease and it has been postulated that high intracellular cholesterol levels promote the amyloidogenic processing of amyloid precursor protein. Therefore we investigated the cellular processing of different apoE isoforms as well as the associated cholesterol efflux in the murine neuronal cell line HT-22. Uptake of apoE3-containing lipoproteins resulted in the expected recycling while, as seen in non-neuronal cells, recycling of apoE4 was significantly reduced. However, despite these differences in apoE recycling, there was no difference in rates of cholesterol efflux. Therefore we conclude that in this neuronal cell model the reduced recycling of apoE4 does not affect cellular cholesterol metabolism.

Characterization of lipid metabolism in insulin-sensitive adipocytes differentiated from immortalized human mesenchymal stem cells.

There is a great demand for cell models to study human adipocyte function. Here we describe the adipogenic differentiation of a telomerase-immortalized human mesenchymal stem cell line (hMSC-Tert) that maintains numerous features of terminally differentiated adipocytes even after prolonged withdrawal of the peroxisome proliferator activated receptor gamma (PPARgamma) agonist rosiglitazone. Differentiated hMSC-Tert developed the characteristic monolocular phenotype of mature adipocytes. The expression of adipocyte specific markers was highly increased during differentiation. Most importantly, the presence of the PPARgamma agonist rosiglitazone was not required for the stable expression of lipoprotein lipase, adipocyte fatty acid binding protein and perilipin on mRNA and protein levels. Adiponectin expression was post-transcriptionally down-regulated in the absence of rosiglitazone. Insulin sensitivity as measured by insulin-induced phosphorylation of Akt and S6 ribosomal protein was also independent of rosiglitazone. In addition to commonly used adipogenic markers, we investigated further PPARgamma-stimulated proteins with a role in lipid metabolism. We observed an increase of lipoprotein receptor (VLDLR, LRP1) and apolipoprotein E expression during differentiation. Despite this increased expression, the receptor-mediated endocytosis of lipoproteins was decreased in differentiated adipocytes, suggesting that these proteins may have an additional function in adipose tissue beyond lipoprotein uptake.

The effect of loss of excess weight on the metabolic risk factors after bariatric surgery in morbidly and super-obese patients.

BACKGROUND: Changes in metabolic risk factors such as dyslipidemia and hyperinsulinemia as well as levels of sex hormones and leptin were studied in morbidly obese (MO) and super-obese (SO) patients during excess weight loss (EWL), separately in males and females. METHODS: In this prospective clinical intervention study, 431 patients were included (361 females and 70 males). There were 217 patients with MO (BMI 40-49.9 kg/m2) and 214 patients with SO (BMI > or =50 kg/m2). All patients underwent restrictive bariatric operations. Metabolic parameters (lipids, insulin, leptin, hepatic transaminases, uric acid, and sex hormones) were measured before obesity surgery and at defined postoperative points of EWL (25%, 50%, 75% and 100%). RESULTS: Successful weight reduction of 25% EWL was achieved by 94% of patients at 2 months. With this moderate EWL, most of the patients already improved their risk profile considerably, including normalization of insulin levels. Additional EWL led to a further amelioration of risk profile in all patients, including normalization of triglyceride levels. Male MO and SO patients had a worse metabolic situation preoperatively and a greater benefit after weight loss. Even though SO patients did not lose as much excess weight as MO patients, they did profit comparably. CONCLUSION: Bariatric surgery is a valuable tool not only to reduce excess weight in severely obese patients but also to improve the metabolic risk profile within a short time-frame. This benefit is most pronounced in high-risk males.

Size and surface effects on the MRI relaxivity of manganese ferrite nanoparticle contrast agents.

Superparamagnetic MnFe2O4 nanocrystals of different sizes were synthesized in high-boiling ether solvent and transferred into water using three different approaches. First, we applied a ligand exchange in order to form a water soluble polymer shell. Second, the particles were embedded into an amphiphilic polymer shell. Third, the nanoparticles were embedded into large micelles formed by lipids. Although all approaches lead to effective negative contrast enhancement, we observed significant differences concerning the magnitude of this effect. The transverse relaxivity, in particular r2*, is greatly higher for the micellar system compared to the polymer-coated particles using same-sized nanoparticles. We also observed an increase in transverse relaxivities with increasing particle size for the polymer-coated nanocrystals. The results are qualitatively compared with theoretical models describing the dependence of relaxivity on the size of magnetic spheres.

Comparison of metabolic risk factors between severely and very severely obese patients.

OBJECTIVE: A prospective clinical intervention study was performed to estimate the metabolic risk factors in patients with very severe obesity (VSO) vs. severe obesity (SO). RESEARCH METHODS AND PROCEDURES: Two hundred twenty-eight VSO (BMI > or = 50 kg/m(2)) and 221 SO patients (BMI = 40 to 49.9 kg/m(2)) participated in the study (367 women and 82 men). Metabolic measurements included plasma lipids, glucose and insulin, hemoglobin A(1c), leptin, and sex hormones, as well as hepatic steatosis in a subgroup of patients. Subgroups of patients with non-insulin-dependent diabetes and hyperlipidemia (HLP) were examined. RESULTS: The most unexpected result of our study was that VSO men showed significantly better lipid profiles than SO men. Furthermore, 18% of VSO men had no metabolic aberrations, whereas all SO men did. The advantageous metabolic status of VSO men was associated with sex hormone changes that favor gynoid fat distribution. The beneficial metabolic situation with VSO seems to be sex specific for men. DISCUSSION: This study shows that the metabolic situation in VSO is not more severe than in the less obese cohort. These findings distinctly differ from the positive associations that have previously been reported between BMI, lipids, and other metabolic indices among individuals whose BMI is <40 kg/m(2).

Apolipoprotein E recycling: implications for dyslipidemia and atherosclerosis.

After receptor-mediated endocytosis, the intracellular fate of triglyceride-rich lipoproteins (TRLs) is far more complex than the classical degradation pathway of low-density lipoproteins. Once internalized, TRLs disintegrate in peripheral endosomes, followed by a differential sorting of TRL components. Although core lipids and apolipoprotein B are targeted to lysosomes, the majority of TRL-derived apolipoprotein E (apoE) remains in peripheral recycling endosomes. This pool of TRL-derived apoE is then mobilized by high-density lipoproteins (HDLs) or HDL-derived apoA-I to be recycled back to the plasma membrane, followed by apoE resecretion and the subsequent formation of apoE-containing HDL. The HDL-induced recycling of apoE is accompanied by cholesterol efflux and involves the internalization and targeting of HDL-derived apoA-I to endosomes containing both apoE and cholesterol. These findings point to a yet unknown intracellular link between TRL-derived apoE, cellular cholesterol transport, and HDL metabolism. Recent studies provide first evidence that impaired recycling of TRL-derived apoE4, but not apoE3, is associated with intracellular cholesterol accumulation, which might explain some well-documented effects of apoE4 on HDL metabolism. This review summarizes the current understanding of apoE recycling and its potential role in the regulation of plasma apoE levels in the postprandial state.

Investigation into the role of the hormone sensitive lipase -60C>G promoter variant in morbid obesity.

BACKGROUND: Hormone sensitive lipase (HSL) plays a central role in free fatty acid homeostasis in adipose tissue and in pancreatic beta-cells, where it contributes to the control of insulin secretion by generating long-chain fatty acids. AIM: We examined the frequency and association of the functional HSL promoter variant, -60C>G, in a German cohort of morbidly obese women (N=239) and men (N=55) and compared the frequency to a cohort of 199 blood donors, recruited from the same region. RESULTS: The rare allele frequency of -60C>G, in the obese individuals was significantly lower 0.031 (95% CI 0.02, 0.04), than that in the blood donors 0.061 (95% CI 0.04, 0.08) p=0.05. The association of the HSL -60C>G with lipid and glucose parameters was examined in the obese women (there were too few men for comparative analysis). In the obese women, those heterozygous for the -60G had significantly higher glucose levels compared to CC women, 142.71 (+/-16.23) mg/dl vs. 110.34 (+/-1.79) mg/dl, respectively (p=0.0001). There was no statistically significant difference in other parameters. CONCLUSION: This study confirms a role for HSL in glucose homeostasis and the reduced frequency of the low expressing -60G promoter variant in obese individuals, together with existing published data, suggests that this allele might be protective against obesity.

Histological changes in the liver of morbidly obese patients: correlation with metabolic parameters.

BACKGROUND: Obesity is a major risk factor for fatty liver disease. The purpose of this study was: 1) to determine the degree of steatosis, inflammation and fibrosis in liver biopsies of morbidly obese patients in relation to their body fat distribution and metabolic status, and 2) to examine the course of liver enzyme changes with surgically-induced weight loss. METHODS: The study population included 179 morbidly obese bariatric surgical patients (82% female, 18% male, mean age 39+/-0.7 (SEM) years, BMI 52+/-0.6 kg/m2, excess body weight 80+/-1.8 kg). All patients tested negative for hepatitis and HIV. Liver biopsies were taken intra-operatively. Hepatic enzyme activities were measured along with lipid parameters, fasting glucose, insulin and leptin. RESULTS: Liver biopsies showed that 47% of morbidly obese females and 85% of males had >30% of hepatocytes filled with fat droplets. Clinically significant hepatic steatosis was associated (P<0.01) with: a) metabolic aberrations, i.e.hyperlipidemia, hyperglycemia, b) male gender, c) abdominal adiposity, and d) elevated hepatic aminotransferase activities. Hepatic inflammation was found in 47% of females and 55% of males, and 'moderate' fibrosis occurred in 12% of males and 6% of females. Postoperatively, the activity of hepatic aminotransferases declined after an initial increase in response to weight loss, with normalization of values occurring at an excess weight loss of 50% (P<0.0001). CONCLUSION: The majority of morbidly obese patients have >30% steatosis of the liver. The incidence of steatosis is higher for males than females, possibly due to their visceral obesity and associated metabolic aberrations.

Knockdown of hepatic ABCA1 by RNA interference decreases plasma HDL cholesterol levels and influences postprandial lipemia in mice.

OBJECTIVE: To investigate the impact of hepatic ABCA1 on systemic lipoprotein metabolism in vivo by an adenovirus-mediated RNA interference approach. METHODS AND RESULTS: Efficiency of plasmid-based small interference RNA (siRNA)-induced knockdown of cotransfected murine ATP binding cassette transporter A1 (mABCA1) in HEK-293 cells was judged by RT-polymerase chain reaction, immunofluorescence, and Western blot analysis. The most effective plasmid was used to generate a recombinant adenovirus as a tool to selectively downregulate ABCA1 expression in mouse liver (C57BL/6). In comparison to controls, Western blot analysis from liver membrane proteins of Ad-anti-ABCA1 infected mice resulted in an approximately 50% reduction of endogenous ABCA1 and a clear upregulation of apolipoprotein E. Fast protein liquid chromatography analysis of plasma revealed that hepatic ABCA1 protein reduction was associated with an approximately 40% decrease of HDL cholesterol and a reduction of HDL-associated apolipoprotein A-I and E. In the fasted state, other lipoprotein classes were not affected. To analyze the influence of ABCA1 downregulation on postprandial lipemia, infected mice were given a gastric load of radiolabeled trioleate in olive oil. In Ad-anti-ABCA1 infected mice, the postprandial increase of chylomicrons and chylomicron-associated apolipoproteins B and E was significantly reduced as compared with controls. CONCLUSIONS: Hepatic ABCA1 contributes to HDL plasma levels and influences postprandial lipemia.

Expression of LRP1 by human osteoblasts: a mechanism for the delivery of lipoproteins and vitamin K1 to bone.

UNLABELLED: Accumulating clinical and experimental data show the importance of dietary lipids and lipophilic vitamins, such as vitamin K1, for bone formation. The molecular mechanism of how they enter the osteoblast is unknown. Here we describe the expression of the multifunctional LRP1 by human osteoblasts in vitro and in vivo. We provide evidence that LRP1 plays an important role in the uptake of postprandial lipoproteins and vitamin K1 by human osteoblasts. INTRODUCTION: Chylomicrons (CM) and their remnants (CR) represent the postprandial plasma carriers of dietary lipids. Dietary vitamin K1 is known to be transported in the circulation as part of CM/CR and is required by osteoblasts as an essential co-factor for the gamma-carboxylation of bone matrix proteins. The molecular mechanisms underlying the delivery of lipophilic substances to bone are not understood. In this study, the expression and function of CM/CR receptors was examined in human osteoblasts. MATERIALS AND METHODS: Four human osteoblast-like cell lines were analyzed: two osteosarcoma lines (MG63, SaOS-2) and two telomerase-immortalized human bone marrow stromal cell lines (hMSC-TERT [4] and [20]) after 1,25(OH)2 vitamin D3 induction of osteoblastic differentiation (hMSC-TERT-OB). Receptor expression was examined by Western blotting and immunohistochemistry of normal human bone sections. Endocytotic receptor function was analyzed by cellular uptake assays using fluorescent and radiolabeled human CR. Vitamin K1-enriched CR (CR-K1) were generated in vivo after oral vitamin administration and vitamin K1 uptake by osteoblasts was measured by HPLC. The effect of CR-K1 uptake on osteocalcin carboxylation was measured by ELISA. RESULTS: Osteoblasts exhibit high levels of protein expression of the CR receptors LRP1 and LDLR. VLDLR is expressed to a lower degree. Immunohistochemistry of normal human bone sections showed strong LRP1 expression by osteoblasts and marrow stromal cells. Uptake of fluorescent CR by osteoblasts resulted in the typical pattern of receptor-mediated endocytosis. CR uptake was stimulated by the exogenous addition of the lipoprotein receptor ligands apolipoprotein E and lipoprotein lipase. Uptake was reduced by the known LRP1 inhibitors RAP, lactoferrin, and suramin, but not by LDL, which exclusively binds to the LDLR. Vitamin K1 uptake by hMSC-TERT-OB after incubation with CR-K1 was also shown to be sensitive to LPL stimulation and the LRP1 specific inhibitor lactoferrin. CR-K1 uptake into osteoblasts stimulated the gamma-carboxylation of osteocalcin. CONCLUSION: Human osteoblasts express receptors of the LDLR family with a capacity for vitamin K1 uptake through CR endocytosis, a novel mechanism for the delivery of dietary lipids and lipophilic vitamins to human bone. The current data suggest that, among the expressed receptors, LRP1 plays a predominant role.

FHR-4A: a new factor H-related protein is encoded by the human FHR-4 gene.

We describe a new member of the human Factor H protein family, termed Factor H-related protein 4A (FHR-4A). The corresponding cDNA sequence was isolated and encodes a secreted protein of 559 amino acids, with a predicted molecular weight of 63.2 kDa. Apparently, this novel cDNA is derived from the human FHR-4 gene. Genetic analysis shows that the human FHR-4 gene is composed of 10 coding exons, and two distinct mRNA transcripts are derived from this gene by alternative splicing. The short FHR-4B form represents a truncated variant and encodes a secreted protein of five domains (previously termed FHR-4). The long transcript encodes the novel FHR-4A protein that is composed of nine complement control protein (CCP) domains. A unique feature of FHR-4A is the tandem arrangement of four CCP domains forming a 'natural dimer' of the short isoform. The FHR-4A protein is identified in human plasma as a 86 kDa protein. The difference between the predicted and observed molecular masses is explained by glycosylation. Comparison of the deduced protein sequence of FHR-4A with peptides from a 86 kDa apolipoprotein described by us earlier suggests that the long form, FHR-4A, represents this apoprotein. In summary, FHR-4A is a new Factor H-related protein with a unique domain composition, that is, an internal duplication of four CCP domains. To our knowledge, FHR-4A provides the first evidence for alternative splicing among Factor H-related genes.