Improved diagnostic criteria for digital flexor tendon sheath pathology using contrast tenography.

BACKGROUND: Pathology of the digital flexor tendon sheath is a significant cause of lameness in the horse. Imaging is important to identify lesions and inform on prognosis prior to tenoscopic surgery. OBJECTIVES: To use a large population to evaluate 1) the sensitivity and specificity of digital flexor tendon sheath (DFTS) contrast radiographs in diagnosing manica flexoria (MF) tears, deep digital flexor tendon (DDFT) tears and constriction of the palmar/plantar annular ligament (PAL) using novel criteria; 2) predisposition to pathology in signalment and limb affected. STUDY DESIGN: Multicentre retrospective cohort study. METHODS: The medical records of 206 horses with lameness localised to the DFTS, contrast radiographs and subsequent tenoscopic surgery were reviewed. Breed and limb predispositions were evaluated for pathology of the DDFT, MF and PAL constriction. Contrast radiographs of the DFTS were reviewed by four masked operators and for each pathology the sensitivity, specificity and interobserver variability were calculated. RESULTS: Contrast tenography was a sensitive test for MF tears (92% confidence interval [CI] 88.4-94.4%; specificity 56%, CI 51.1-61.1%) and specific for diagnosing DDFT tears (73%, CI 68.6-76.8%; sensitivity 54%, CI 47.8-60.2%) but had a lower sensitivity (71%, CI 65.1-75.9% ) and specificity (45%, CI 39.1-52.0%) for PAL constriction. It had good to substantial interobserver agreement for MF and DDFT tears (Krippendorff's alpha 0.68 and 0.46 respectively). Ponies (57%) and cobs (58%) were significantly more likely to be affected with MF tears (other breeds 20-39%, P = 0.003) and Thoroughbreds (50%), warmbloods (45%) and draught breeds (48%) were more likely to have DDFT tears (other breeds 22-34%, P = 0.01). MF tears and PAL constriction were overrepresented in the hindlimbs compared to DDFT tears in forelimbs. MAIN LIMITATIONS: No standardisation of contrast radiographs was possible. The subjectivity of diagnosis of PAL constriction may also have led to bias. Radiographs were read as JPEGS reducing ability to manipulate images. CONCLUSIONS: Contrast radiography of the DFTS is accurate in the pre-operative diagnosis of DFTS pathologies. Different pathologies are overrepresented in certain breeds and limbs.

Glyco-engineered anti-EGFR mAb elicits ADCC by NK cells from colorectal cancer patients irrespective of chemotherapy.

BACKGROUND: The epidermal growth factor receptor (EGFR) is overexpressed in colorectal cancer (CRC), and is correlated with poor prognosis, making it an attractive target for monoclonal antibody (mAb) therapy. A component of the therapeutic efficacy of IgG1 mAbs is their stimulation of antibody-dependent cellular cytotoxicity (ADCC) by natural killer (NK) cells bearing the CD16 receptor. As NK cells are functionally impaired in cancer patients and may be further compromised upon chemotherapy, it is crucial to assess whether immunotherapeutic strategies aimed at further enhancing ADCC are viable. METHODS: CRC patients before, during and after chemotherapy were immunophenotyped by flow cytometry for major white blood cell populations. ADCC-independent NK cell functionality was assessed in cytotoxicity assays against K562 cells. ADCC-dependent killing of EGFR(+) A431 cancer cells by NK cells was measured with a degranulation assay where ADCC was induced by GA201, an anti-EGFR mAb glyco-engineered to enhance ADCC. RESULTS: Here, we confirm the observation that NK cells in cancer patients are dysfunctional. However, GA201 was able to induce robust NK cell-dependent cytotoxicity in CRC patient NK cells, effectively overcoming their impairment. CONCLUSIONS: These findings support the evaluation of the therapeutic potential of GA201 in combination with chemotherapy in CRC patients.

A novel combination of promoter and enhancers increases transgene expression in vascular smooth muscle cells in vitro and coronary arteries in vivo after adenovirus-mediated gene transfer.

Recombinant adenoviruses are employed widely for vascular gene transfer. Vascular smooth muscle cells (SMCs) are a relatively poor target for transgene expression after adenovirus-mediated gene delivery, however, even when expression is regulated by powerful, constitutive viral promoters. The major immediate-early murine cytomegalovirus enhancer/promoter (MIEmCMV) elicits substantially greater transgene expression than the human cytomegalovirus promoter (MIEhCMV) in all cell types in which they have been compared. The Woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) increases transgene expression in numerous cell lines, and fragments of the smooth muscle myosin heavy chain (SMMHC) promoter increase expression within SMC from heterologous promoters. We therefore, compared the expression of beta-galactosidase after adenovirus-mediated gene transfer of lacZ under the transcriptional regulation of a variety of combinations of the promoters and enhancers described, in vitro and in porcine coronary arteries. We demonstrate that inclusion of WPRE and a fragment of the rabbit SMMHC promoter along with MIEmCMV increases beta-galactosidase expression 90-fold in SMC in vitro and approximately 40-fold in coronary arteries, compared with vectors in which expression is regulated by MIEhCMV alone. Expression cassette modification represents a simple method of improving adenovirus-mediated vascular gene transfer efficiency and has important implications for the development of efficient cardiovascular gene therapy strategies.

[Clinic admission?--but not without my child!]. / Klinik?--Aber nicht ohne mein Kind!

Since the end of the 1990s the Westfälische Klinik Lengerich has offered an inpatient psychiatric treatment of an psychiatric ill parent--combined with a pedagogic attendance of the child taken along. The following article describes our experience in that matter, from a ward concentrated on psychotherapy. In selected examples supposed patterns of interaction--following the specific disease--are represented from both the psychotherapeutic und the pedagogic point of view. In practice our plan of treatment and attendance has proved successful for the wellbeing of the patients and their children. Remains to be hoped that--despite difficulties in organisation which will have to be overcome--similar offers can be made in regional psychiatric attendance.

Efficient FLPe recombinase enables scalable production of helper-dependent adenoviral vectors with negligible helper-virus contamination.

Helper-dependent (HD), high-capacity adenoviruses are one of the most efficient and safe gene therapy vectors, capable of mediating long-term expression. Currently, the most widely used system for HD vector production avoids significant contamination with helper virus by using producer cells stably expressing a nuclear-targeted Cre recombinase and an engineered first-generation helper virus with parallel loxP sites flanking its packaging signal. The system requires a final, density-based separation of HD and residual helper viruses by ultracentrifugation to reduce contaminating helper virus to low levels. This separation step hinders large-scale production of clinical-grade HD virus. By using a very efficient recombinase, in vitro-evolved FLPe (ref. 14), to excise the helper virus packaging signal in the producer cells, we have developed a scalable HD vector production method. FLP has previously been shown to mediate maximum levels of excision close to 100% compared to 80% for Cre (ref. 15). Utilizing a common HD plasmid backbone, the FLPe-based system reproducibly yielded HD virus with the same low levels of helper virus contamination before any density-based separation by ultracentrifugation. This should allow large-scale production of HD vectors using column chromatography-based virus purification.

Gene transfer into rat brain using adenoviral vectors.

Recombinant adenovirus vectors are attractive vehicles to deliver genes into the brain for the purposes of neurobiological research and for gene therapy of neurological diseases. This unit provides a comprehensive set of protocols for adenovirus vector-mediated gene transfer to the brain, including introduction of the vector into the brain by stereotaxic injection and preparation and processing of brain tissue for the evaluation of gene transfer. The potential side-effects of administering adenovirus vectors to the brain are discussed in detail. The unit also provides protocols for evaluating these side-effects (e.g., demyelination, inflammation, vector-mediated cytotoxicity, etc.). Finally, critical parameters for obtaining optimal gene transfer with minimum side-effects are presented.

Strong promoters are the key to highly efficient, noninflammatory and noncytotoxic adenoviral-mediated transgene delivery into the brain in vivo.

Using the major immediate early murine cytomegalovirus (MIEmCMV) promoter to drive expression of beta-galactosidase, we have demonstrated that, following adenoviral-mediated transduction of brain cells in vivo, a single viral infectious unit is capable of producing detectable levels of transgene expression and that gene transfer into the brain is close to 100% efficient. By reducing 100-fold the amount of virus needed to detect large numbers of transduced brain cells, we were able to completely eliminate the cellular inflammation and viral cytotoxicity associated with the delivery of adenoviral vectors into the brain compared to saline-injected controls. These results demonstrate that a strong promoter is necessary to allow the use of low concentrations of adenoviral vectors for gene transfer into the brain, thereby eliminating deleterious side effects and increasing the potential efficacy of gene therapy.

Transcriptional targeting to anterior pituitary lactotrophic cells using recombinant adenovirus vectors in vitro and in vivo in normal and estrogen/sulpiride-induced hyperplastic anterior pituitaries.

The use of pituitary cell type-specific promoters is a powerful molecular tool to achieve pituitary cell type-specific transcriptional targeting of transgenes encoded by viral vectors. It has recently been proposed that transcriptional targeting of therapeutic genes could be harnessed as a gene therapy strategy for the treatment of pituitary disease. We describe the successful use of the human PRL promoter (hPrl) encoded within recombinant adenovirus vectors to target transgene expression of Herpes Simplex Virus Type 1-Thymidine Kinase (HSV1-TK) or beta-galactosidase to lactotrophic cells in vitro and in vivo. Functionally, the restriction of expression of HSV1-TK to lactotrophic tumor cells, using the hPrl promoter, resulted in the cell type-specific induction of apoptosis in the lactotrophic GH3 tumor cell line, in the presence of ganciclovir (GCV). In the corticotrophic AtT20 cell line, we detected neither HSV1-TK expression, nor apoptosis in the presence of GCV. The hPrl promoter encoded within a recombinant adenoviral vector also restricted transgene expression to lactotrophic cells in primary anterior pituitary (AP) cultures, and importantly, within the anterior pituitary gland in vivo. When the HSV1-TK driven by hPrl promoter was used in an in vivo model ofestrogen/sulpiride lactotroph induced hyperplasia within the AP in situ, the treatment was not effective in either reducing the weight of the gland, the number of lactotrophic cells within the transduced area in vivo, or the circulating PRL levels. This is in contrast to the human cytomegalovirus promoter (hCMV) driving expression of HSV1-TK in the same experimental paradigm, which was effective in reducing pituitary weight and circulating PRL levels. Our results have important implications in the design of gene therapy strategies for pituitary tumors. We demonstrate that both the choice of the in vivo animal model, i.e. adenoma in the AP gland in situ, and the particular gene therapy strategy chosen, i.e. use of strong ubiquitous promoters vs. weaker but cell type-specific promoters, determine the experimental therapeutic outcome.

Postprandial response to a fat tolerance test in young adults with a paternal history of premature coronary heart disease - the EARS II study (European Atherosclerosis Research Study).

BACKGROUND: The European Atherosclerosis Research Study (EARS) I had shown that fasting plasma concentrations of apolipoprotein B (apo B) and triglycerides were the most discriminant variables between offspring with a paternal history of coronary heart disease (CHD) and controls. The EARS II study was undertaken to investigate whether a paternal history of CHD was associated with differences in postprandial lipemia. DESIGN: Male subjects with a paternal history of CHD (cases, n = 407) and age-matched male controls (n = 415) were recruited from 14 European universities. All subjects had an oral fat tolerance test. RESULTS: In the sample as a whole, the postprandial triglyceride responses did not significantly differ between the two groups. However, in the upper tertile of fasting triglycerides, cases displayed a higher area under the curve (5.71 vs. 4.49 mmol.h L-1, P < 0.001), a higher peak (1.76 vs. 1.43 mmol L-1, P < 0.001) and a more delayed time to peak (3.15 vs. 2.91 h, P < 0.05) than controls. In the upper tertile, fasting apo B levels (P < 0.05) and triglyceride area under the curve (P = 0.002) significantly discriminated cases from controls in a multivariate analysis. Cases had also higher Lp C-III:B levels at 4 h than controls (11.2 vs. 9.9 mg dL-1, P < 0.01) and this difference remained significant after adjustment for apo B and triglyceride levels. CONCLUSIONS: These results indicate that in subjects with a moderate elevation of fasting triglycerides, an impaired postprandial response to a fat load constitutes an early biological expression of a paternal history of premature CHD.

The apolipoprotein epsilon4 allele determines prognosis and the effect on prognosis of simvastatin in survivors of myocardial infarction : a substudy of the Scandinavian simvastatin survival study.

BACKGROUND: Carriers of the epsilon4 allele of the apolipoprotein E gene are at a higher risk of coronary heart disease than individuals with other genotypes. We examined whether the risk of death or a major coronary event in survivors of myocardial infarction depended on apolipoprotein E genotype and whether the benefits of treatment with simvastatin differed between genotypes. METHODS AND RESULTS: Cox proportional hazards models were used to analyze 5.5 years of follow-up data from 966 Danish and Finnish myocardial infarction survivors enrolled in the Scandinavian Simvastatin Survival Study. A total of 16% of the 166 epsilon4 carriers in the placebo group died compared with 9% of the 312 patients without the allele, which corresponds to a mortality risk ratio of 1.8 (95% confidence interval, 1.1 to 3.1). The risk ratio was unaffected by considerations of sex, age, concurrent angina, diabetes, smoking, and serum lipids in multivariate analyses. Simvastatin treatment reduced the mortality risk to 0.33 (95% confidence interval, 0.16 to 0.69) in epsilon4 carriers and to 0.66 (95% confidence interval, 0. 35 to 1.24) in other patients (P=0.23 for treatment by genotype interaction). Apolipoprotein E genotype did not predict the risk of a major coronary event. Baseline serum levels of lipoprotein(a) also predicted mortality risk and could be combined with epsilon4-carrier status to define 3 groups of patients with different prognoses and benefits from treatment. CONCLUSIONS: Myocardial infarction survivors with the epsilon4 allele have a nearly 2-fold increased risk of dying compared with other patients, and the excess mortality can be abolished by treatment with simvastatin.

A comparison of lifestyle, genetic, bioclinical and biochemical variables of offspring with and without family histories of premature coronary heart disease: the experience of the European Atherosclerosis Research Studies.

BACKGROUND: Lifestyle, genetic, bioclinical and biochemical factors of European university students aged 18-26 years, with and without documented paternal histories of premature coronary heart disease, have been compared in the European Atherosclerosis Research Studies (EARS) I and II. OBJECTIVE: To highlight consistencies and inconsistencies between findings in the two studies. METHODS: All measurements were made according to strict protocols, by trained technicians using validated methods. The results for men in EARS I are compared with those from EARS II which was confined to men. RESULTS: In both studies we found no differences between cases and controls in lifestyle factors and bioclinical factors except that controls were taller. We found inconsistent differences between obesity indices and antecedents of arterial hypertension. In both studies we found consistent differences between cases and controls in levels of total cholesterol and apolipoprotein B, both levels being higher in cases. The lack of any difference between levels of high-density lipoprotein cholesterol and apolipoprotein A1 was also found consistently. Inconsistent differences were found for levels of triglycerides and apolipoprotein E. For most of the candidate genes that were studied, no differences between cases and controls were found, but different polymorphisms were associated with levels of lipids, apoproteins and fibrinogen independently of case-control status. Some of these associations were potentiated by lifestyle factors. The interaction between genetic and environmental factors is further illustrated with results from the association of apolipoprotein E polymorphism with the level of apolipoprotein B and a variety of other determinants of apolipoprotein B level. CONCLUSIONS: In the EARS studies a documented family history of premature coronary heart disease was mainly expressed in terms of biochemical factors that are determined both by nature and by nurture.

The intrinsic factor-vitamin B12 receptor, cubilin, is a high-affinity apolipoprotein A-I receptor facilitating endocytosis of high-density lipoprotein.

Cubilin is the intestinal receptor for the endocytosis of intrinsic factor-vitamin B12. However, several lines of evidence, including a high expression in kidney and yolk sac, indicate it may have additional functions. We isolated apolipoprotein A-I (apoA-I), the main protein of high-density lipoprotein (HDL), using cubilin affinity chromatography. Surface plasmon resonance analysis demonstrated a high-affinity binding of apoA-I and HDL to cubilin, and cubilin-expressing yolk sac cells showed efficient 125I-HDL endocytosis that could be inhibited by IgG antibodies against apoA-I and cubilin. The physiological relevance of the cubilin-apoA-I interaction was further emphasized by urinary apoA-I loss in some known cases of functional cubilin deficiency. Therefore, cubilin is a receptor in epithelial apoA-I/HDL metabolism.

Two common mutations (D9N, N291S) in lipoprotein lipase: a cumulative analysis of their influence on plasma lipids and lipoproteins in men and women.

We assessed the effect of two common mutations in the lipoprotein lipase gene (LPL), D9N and N291S, which have been shown to modulate plasma lipids in a wide spectrum of patients. A total of 1114 men and 1 144 women from the Framingham Offspring Study (FOS) were analyzed for these two LPL variants. Subsequently, the association with fasting plasma lipids and risk of coronary artery disease (CHD) was determined. We extended our study by calculating weighed means of lipids and lipoproteins in carriers and non-carriers for these LPL mutations in patients with genetic dyslipidemias, CHD patients and healthy controls. In the FOS sample, the D9N and N291S alleles were associated with lower high-density lipoprotein-cholesterol (HDL-C) (delta = - 0.07 mmol/ 1, p = 0.03) and a trend towards increased triglycerides (delta = 0.25 mmol/ 1, p = 0.07). In women, a trend towards the high triglyceride, low HDL-C phenotype was evident (delta = - 0.02 mmol/1 for HDL-C and delta = 0.14 mmol/l for triglycerides, respectively). Cumulative analysis of other studies of male carriers of the D9N and N291S revealed higher levels of triglycerides (D291N; 2.60(1.85) mmol/l vs. 1.62(1.18) mmol/l: p < 0.0001) (D9N; 1.94 (1.19) mmol/l vs. 1.74(1.17) mmol/l: p < 0.001) and lower HDL-C (N291S; 1.04(0.32) mmol/l vs. 1.15(0.28) mmol/l: p < 0.0001) (D9N; 1.08(0.24) mmol/l vs. 1.16(0.28) mmol/l: p < 0.0001). In females, results differed with higher TG levels (N291S; 1.70(0.99) mmol/l vs. 1.10(0.63) mmol/l: p < 0.001) (D9N; 1.08(0.76) mmol/l vs. 0.96(0.51) mmol/l: p < 0.01) and lower HDL-C levels (N291S; 1.27(0.33) mmol/l vs. 1.51(0.32) mmol/l: p < 0.0001); however, the HDL-C levels for D9N carriers were similar to non-carriers (D9N; 1.52(0.29) mmol/l vs. 1.53(0.35) mmol/l: p = 0.83). Our data provide evidence that common variants of the LPL gene are significant modulators of lipid and lipoprotein levels in both men and women.

'Complete' spinal cord injury does not block perceptual responses to genital self-stimulation in women.

BACKGROUND: A priori hypothesis: vaginal and/or cervical self-stimulation will not produce perceptual responses in women with "complete" spinal cord injury (SCI) at or above the highest level of entry of the hypogastric nerves (T10-12) but will produce perceptual responses if SCI is below T-10. DESIGN: Women with complete SCI were assigned to a group with "upper" (T-10 and/or above) (n = 6) or "lower" (below T-10) (n = 10) SCI; uninjured women (n = 5) constituted a control group. Perceptual response to vaginal and/or cervical self-stimulation was quantified as magnitude of analgesia to calibrated finger compressive force. SETTING: Rutgers, The State University of New Jersey, Human Physiology Laboratory, College of Nursing, Newark. PARTICIPANTS: Consecutive samples of first 16 of 34 women with SCI who responded to nationwide advertisements, met inclusion criteria, and volunteered; control group was the first 5 respondents. INTERVENTION: Vaginal or cervical (cervix uteri) self-stimulation applied for 12 minutes, interspersed with non-stimulation periods, while measuring analgesia. MAIN OUTCOME MEASURE: Quantify analgesia magnitude to vaginal or cervical self-stimulation. RESULTS: Significant analgesia was produced in the uninjured group and the group with lower SCI, supporting the hypothesis. Unexpectedly, significant analgesia was also produced in the group with upper SCI. Women in the group with upper SCI also experienced menstrual discomfort, awareness of vaginal and/or cervical stimulation per se, and orgasms. CONCLUSIONS: (1) Genitospinal visceral afferent pathways function in the women in the group with upper SCI, although unrecognized by the American Spinal Injury Association criteria, and/or (2) there exists a functional genital afferent pathway that bypasses the spinal cord and projects directly to the brain, which we propose to be via the vagus nerves.

Lipoprotein lipase variants D9N and N291S are associated with increased plasma triglyceride and lower high-density lipoprotein cholesterol concentrations: studies in the fasting and postprandial states: the European Atherosclerosis Research Studies.

BACKGROUND: Variations at the DNA level with moderate effects on biochemical variables may be important for the occurrence of disease at the population level, if they are common. Two mutations in the LPL gene, N9 and S291, are associated with variation in fasting plasma concentrations of HDL cholesterol (HDL-C) and triglycerides (TG). We investigated whether these mutants were more frequent in offspring of cases with premature coronary disease and analyzed the effects on fasting plasma lipids and postprandial TG. METHODS AND RESULTS: Students with and without paternal history of myocardial infarction (cases and control subjects [controls]) were studied in the European Atherosclerosis Research Studies I and II (EARS-I and -II). Allelic frequencies for the N9 and S291 mutations did not differ between cases and control subjects. The N9 mutation was identified in 4.2% of all subjects in EARS-I, and carriers had higher fasting TG levels (P<.001) than noncarriers. In an oral fat tolerance test, there were no differences in postprandial TG between carriers and noncarriers of the N9 allele. The S291 mutation was identified in 3.1% of all subjects in EARS-I, and carriers had lower fasting HDL-C levels (P<.005) than noncarriers. There was a significant interaction between S291 genotype and body mass index on fasting TG levels (P<.01). In the cases, carriers of the S291 allele had higher TG levels 6 hours postprandially (P<.04) than did noncarriers. CONCLUSIONS: The two LPL mutations are common and may predispose to elevated TG and decreased HDL-C concentrations, even in young subjects. In the case of the S291 mutation, this effect appears to be mediated via delayed postprandial TG clearance. Moreover, even moderate obesity potentiates the TG-raising and HDL-lowering effects associated with the S291 allele.

Phenotypic variation in patients heterozygous for familial defective apolipoprotein B (FDB) in three European countries.

A glutamine-for-arginine substitution at amino acid position 3500 of apolipoprotein B (apo B) causes synthesis of LDL with reduced binding affinity to the LDL receptor (LDLR). The associated clinical syndrome has been named familial defective apolipoprotein B- 100 (FDB). In 205 FDB patients from Germany (n = 73). The Netherlands (n = 87), and Denmark (n = 45), we tried to assess determinants of variation in lipid concentrations. Besides age, sex, and geographic origin, variation in the LDLR gene was the most powerful determinant of variation in total cholesterol and LDL cholesterol levels. Polymorphic variation in the LDLR gene (SfaNI, exon 2; Nco I, exon 18) was associated with total cholesterol (TC) and LDL cholesterol (LDL-C) variation in women (SfaNI: P = .04 and .03 for TC and LDL-C, respectively; Nco I; P = .003 and .006, respectively), whereas the Ava II (exon 13) and the Pvu II (intron 15) polymorphisms were not. Combined information from all three LDLR exon polymorphisms showed that subjects with at least one S + A + N + allele had 13% to 20% higher TC than non-S + A + N + subjects (P = .02 [TC, men]; P = .01 [LDL-C, men]; P = .005 [TC, women]; and P = .004 [LDL-C, women]) and, together with age and geographic origin, accounted for 20% (women) and 19% (men) of the variation in LDL-C. The expected association of the apo E genotypes (e3e2, e3e3, and e3e4) with cholesterol concentrations was seen in S + A + N + but not in non-S + A + N + subjects and in P-P- but not in P + P + or P + P- subjects. With regard to clinical expression, FDB patients had lower TC and LDL-C levels and a lower prevalence of cardiovascular disease than 101 Danish patients with familial hypercholesterolemia.

Isoform-specific binding of human apolipoprotein E to the non-amyloid beta component of Alzheimer's disease amyloid.

The non-A beta component (NAC) of Alzheimer's disease amyloid is a newly discovered 35 amino acid peptide found to be closely linked to the beta-amyloid fibrils in senile plaques. Apolipoprotein E (apoE) is another prominent constituent of senile plaques. In vitro studies have shown that apoE binds beta-amyloid (A beta) with high avidity, but it is unknown to what extent apoE interacts with NAC. We examined the interactions between apoE and NAC and found that apoE bound synthetic NAC, forming a complex that resisted reducing agents and separation on SDS-PAGE. The complex could be formed using apoE from either purified human very low density lipoprotein (VLDL) particles, unfractionated human cerebrospinal fluid (CSF), or recombinant protein. The binding was established within 15 min upon mixing, and the interaction between NAC and apoE was dose-dependent and specific as revealed by competition experiments. The NAC-apoE complex was affected by non-physiological pH, but not by reducing agents such as DTT or beta-mercaptoethanol. ApoE exists in different isoforms of which the apoE3 genotype is the most frequent. Notably, the apoE4 genotype has been linked to late-onset Alzheimer's disease. This study presents evidence that apoE3 as well as apoE4 bind NAC, but the binding to apoE4 is about twice as strong as to apoE3. The isoform-specific binding of NAC to apoE may thus play an important role in amyloidogenesis and in the sequestering of apoE in senile plaques during the progress of Alzheimer's disease.