Impact of whole dairy matrix on musculoskeletal health and aging-current knowledge and research gaps.

Dairy products are included in dietary guidelines worldwide, as milk, yoghurt, and cheese are good sources of calcium and protein, vital nutrients for bones and muscle mass maintenance. Bone growth and mineralization occur during infancy and childhood, peak bone mass being attained after early adulthood. A low peak bone mass has consequences later in life, including increased risk of osteoporosis and fractures. Currently, more than 200 million people worldwide suffer from osteoporosis, with approximately 9 million fractures yearly. This poses a tremendous economic burden on health care. Between 5% and 10% of the elderly suffer from sarcopenia, the loss of muscle mass and strength, further increasing the risk of fractures due to falls. Evidence from interventional and observational studies support that fermented dairy products in particular exert beneficial effects on bone growth and mineralization, attenuation of bone loss, and reduce fracture risk. The effect cannot be explained by single nutrients in dairy, which suggests that a combined or matrix effect may be responsible similar to the matrix effects of foods on cardiometabolic health. Recently, several plant-based beverages and products have become available and marketed as substitutes for dairy products, even though their nutrient content differs substantially from dairy. Some of these products have been fortified, in efforts to mimic the nutritional profile of milk, but it is unknown whether the additives have the same bioavailability and beneficial effect as dairy. We conclude that the dairy matrix exerts an effect on bone and muscle health that is more than the sum of its nutrients, and we suggest that whole foods, not only single nutrients, need to be assessed in future observational and intervention studies of health outcomes. Furthermore, the importance of the matrix effect on health outcomes argues in favor of making future dietary guidelines food based.

Correction to: Impact of whole dairy matrix on musculoskeletal health and aging-current knowledge and research gaps.

The article Impact of whole dairy matrix on musculoskeletal health and aging-current knowledge and research gaps written by N.R.W. Geiker, C. Mølgaard, S. Iuliano, R. Rizzoli,Y. Manios, L.J.C. van Loon, J.-M. Lecerf, G. Moschonis, J.-Y. Reginster, I. Givens, A. Astrup.

How French subjects describe well-being from food and eating habits? Development, item reduction and scoring definition of the Well-Being related to Food Questionnaire (Well-BFQ©).

Providing well-being and maintaining good health are main objectives subjects seek from diet. This manuscript describes the development and preliminary validation of an instrument assessing well-being associated with food and eating habits in a general healthy population. Qualitative data from 12 groups of discussion (102 subjects) conducted with healthy subjects were used to develop the core of the Well-being related to Food Questionnaire (Well-BFQ). Twelve other groups of discussion with subjects with joint (n = 34), digestive (n = 32) or repetitive infection complaints (n = 30) were performed to develop items specific to these complaints. Five main themes emerged from the discussions and formed the modular backbone of the questionnaire: "Grocery shopping", "Cooking", "Dining places", "Commensality", "Eating and drinking". Each module has a common structure: items about subject's food behavior and items about immediate and short-term benefits. An additional theme - "Eating habits and health" - assesses subjects' beliefs about expected benefits of food and eating habits on health, disease prevention and protection, and quality of ageing. A preliminary validation was conducted with 444 subjects with balanced diet; non-balanced diet; and standard diet. The structure of the questionnaire was further determined using principal component analyses exploratory factor analyses, with confirmation of the sub-sections food behaviors, immediate benefits (pleasure, security, relaxation), direct short-term benefits (digestion and satiety, energy and psychology), and deferred long-term benefits (eating habits and health). Thirty-three subscales and 14 single items were further defined. Confirmatory analyses confirmed the structure, with overall moderate to excellent convergent and divergent validity and internal consistency reliability. The Well-BFQ is a unique, modular tool that comprehensively assesses the full picture of well-being related to food and eating habits in the general population.

Management of postmenopausal women: Collège National des Gynécologues et Obstétriciens Français (CNGOF) and Groupe d'Etude sur la Ménopause et le Vieillissement (GEMVi) Clinical Practice Guidelines.

AIM: The aim of these recommendations is to set forth an individualized approach to the management of early postmenopausal women (i.e., within the first 10 years after natural menopause) covering all aspects of lifestyle and therapeutic management, with or without menopause hormone therapy (MHT). MATERIALS AND METHODS: Literature review and consensus of French expert opinion. Recommendations were graded according to the HAS methodology and levels of evidence derived from the international literature, except when there was no good-quality evidence. SUMMARY RECOMMENDATIONS: The beginning of menopause is an ideal time for each woman to evaluate her health status by assessing her bone, cardiovascular, and cancer-related risk factors that may be amplified by postmenopausal estrogen deficiency and by reviewing her lifestyle habits. Improving lifestyle, including nutrition and physical activity, and avoiding risk factors (notably smoking), should be recommended to all women. MHT remains the most effective treatment for vasomotor symptoms but it could be also recommended as first-line treatment for the prevention of osteoporosis in early postmenopausal women at low to moderate risk for fracture. The risks of MHT differ depending on its type, dose, duration of use, route of administration, timing of initiation, and whether a progestogen is used. There is reasonable evidence that using transdermal estradiol in association with micronized progesterone or dydrogesterone may limit both the venous thromboembolic risk associated with oral estrogens and the risk of breast cancer associated with synthetic progestins. Treatment should be individualized to each woman, by using the best available evidence to maximize benefits and minimize risks, with periodic reevaluation of its benefit-risk balance. For bothersome genitourinary syndrome of menopause (GSM) symptoms, vaginal treatment with lubricants and moisturizers is recommended as first-line treatment together with low-dose vaginal estrogen therapy, depending on the clinical course. No recommendation of an optimal duration of MHT can be made, but it must take into consideration the initial indication for MHT as well as each woman's benefit-risk balance. Management of gynecological side-effects of MHT is also examined. These recommendations are endorsed by the Groupe d'Etude sur la Ménopause et le Vieillissement hormonal (GEMVI) and the Collège National des Gynécologues-Obstétriciens Français (CNGOF).

[Nutritional advices for postmenopausal woman. Postmenopausal women management: CNGOF and GEMVi clinical practice guidelines]. / Conseils nutritionnels pour la femme ménopausée. RPC Les femmes ménopausées du CNGOF et du GEMVi.

Menopause is a key period for health due to physiological changes, particularly of body composition (with decrease of lean mass and increase of fat mass) and of body fat distribution, leading to a higher risk for bone and muscular health and cardiometabolic health. Nutritional advices, associated to physical activity advices, may partially prevent these effects. The energy balance will be moderately negative if there is a weight gain, while the protein intake will be preserved and a regular physical activity will be increased. A Mediterranean style diet will be beneficial on cardiovascular health. Dairy products will be preserved, but restrictive and dietary exclusion will be avoided.

Effects of two marine dietary supplements with high calcium content on calcium metabolism and biochemical marker of bone resorption.

BACKGROUND/OBJECTIVE: Calcium is essential for the bone metabolism but daily calcium requirements are not met in a significant proportion of the population. Fortunately, oral calcium supplementation can help to meet these needs; however, the calcium bioavailability depends on the calcium sources. The calcium absorption and bioavailability of dietary supplements from marine sources are not known. The objectives of this study were to evaluate the effects of two marine dietary supplements with a high calcium content: a fishbone powder (Phoscalim) and a ray cartilage hydrolysate (Glycollagene), in comparison with milk, and a placebo (maltodextrin), on calcium metabolism and a biochemical marker of bone resorption, using the oral calcium tolerance test. SUBJECTS: Twenty male volunteers were randomized to eat 836 mg of calcium from different sources compared to maltodextrin during a Latin square study. Serum calcium concentrations and other parameters of the calcium metabolism, such as serum intact parathyroid hormone (iPTH) and serum C telopeptides (s-CTX), were measured after an acute oral calcium load based on the Pak protocol. RESULTS: An increase in serum-corrected calcium areas under the curve (AUC) occurred with Phoscalim and Glycollagene when compared to milk. Significantly lower iPTH concentrations were observed with Glycollagene than with milk at T0+1 h, T0+3 h, T0+6 h and with Phoscalim than with milk at T0+6 h. A significantly lower s-CTX concentration was observed with Glycollagene than with milk and Phoscalim at T0+6 h. Furthermore, the urinary calcium/creatinine ratio increased significantly more with Glycollagen than with milk in T0 h+3 h and T3 h+6 h. CONCLUSION: These two dietary supplements from marine sources constitute oral calcium sources when compared to milk on calcium absorption and bone resorption markers on short time.

[The behavioral goals of the coronary patient: No longer smoke, eat better, move more and better]. / Les objectifs comportementaux du patient coronarien : ne plus fumer, manger mieux, bouger plus et mieux.

The behavioral goals of the coronary patient require active management by the cardiologist. Every smoker must be clearly informed about the cardiovascular consequences of smoking and the major benefits of smoking cessation. The only advice to "quit smoking" is not enough. Validated "treatments" (cognitive-behavioral therapy, nicotine replacement therapy, varenicline, bupropion) must be used, with a precise strategy and prolonged follow-up. All drugs assistance can be prescribed in coronary patients and nicotine replacement therapy can even be used just after a myocardial infarction. Nutrition plays a significant role in cardiovascular prevention. Counseling today is based on solid evidence, although evidence is harder to obtain than with drugs. It should no longer be advisable only to "suppress cooked fats and starches" because these recommendations are unclear and/or false. Today we need positive food-based benchmarks and complex dietary patterns in which fruits and vegetables, fish, whole grains, pulses, nuts, olive oil and a diet closed to the Mediterranean diet. Dairy products have their place. Sugary foods should be limited especially in case of overweight and metabolic syndrome. Physical activity is part of good nutrition. Indeed, the fight against a very sedentary lifestyle and physical inactivity in coronary and heart failure patients is part of the lifelong treatment of these patients. The cardiologist and the general practitioner must be much more involved in their prescription and education to hope for good compliance.

Repertoire cloning of lupus anti-DNA autoantibodies.

To investigate the autoantibody repertoire associated with SLE, we have created phage display IgG Fab libraries from two clinically active SLE patients and from the healthy identical twin of one of these patients. The libraries from the lupus discordant twins were found to both include unusually large representations of the V(H)5 gene family. By panning with DNA, the SLE libraries each yielded IgG anti-double-stranded (ds) DNA autoantibodies, which are characteristic of lupus disease. These included a V(H)5 autoantibody from the affected twin, that has a targeted cluster of mutations that potentially improves binding affinity. The recovered IgG anti-dsDNA autoantibodies expressed the same idiotypes associated with the in vivo IgG anti-dsDNA response of the respective SLE donor. Heavy-light chain shuffling experiments demonstrated a case in which the in vitro creation of anti-dsDNA binding activity required restrictive pairing of a heavy chain with Vlambda light chains similar to those in circulating anti-dsDNA autoantibodies. By contrast, IgG anti-ds autoantibodies could not be recovered from the library from the healthy twin, or from shuffled libraries with heavy chains from the healthy twin. These repertoire analyses illustrate how inheritance and somatic processes interplay to produce lupus-associated IgG autoantibodies.

[Some notions about possible harmlessness and/or toxicity of the antioxidant micronutriments]. / Quelques notions sur l'innocuité et la toxicité éventuelles des supplémentations en micronutriments antioxydants.

Report # 8 of the Age Related Eye Disease Study (AREDS) showed the interest of a cocktail of antioxidant micronutrients for the uni- or bilateral intermediate forms and the unilateral evolved forms of Age related Macular Degeneration. This use of supranutritional amounts aiming at obtaining a therapeutic effect corresponds to the concept of "neutraceuticals" which can be opposed to the concept of nutritional amounts. Although the AREDS was carried out under strict conditions, the evolution of knowledge in micronutrition since its design has led to some criticism of both the amounts of the micronutrients and the composition of the formulation. For example several authors pointed out that beta-carotene used at 3 fold the daily recommendations could have harmful effects, especially among smokers or former smokers. Other authors pointed out that vitamin E, at amounts corresponding to 40 to 60 fold the amounts recommended could be correlated with a lethal risk. We develop here some notions about safety and/or harmlessness of the antioxidant micronutrients.

Overexpression of the multidrug resistance-associated protein (MRP) gene in vincristine but not doxorubicin-selected multidrug-resistant murine erythroleukemia cells.

Multidrug-resistant sublines of the murine erythroleukemia cell line PC4 were sequentially selected in increasing vincristine concentrations (5-160 ng/ml). The low- and intermediate-level resistant cell lines, selected in < or = 40 ng/ml of vincristine, demonstrated resistance to Vinca alkaloids and to an epipodophyllotoxin but little or none to an anthracycline. The expression of murine mdr genes, as analyzed by Northern blotting, revealed a baseline expression of murine mdr2 in parental cells that was unchanged in the drug-resistant cell lines. Overexpression of mdr3 was observed only in the highest-level resistant cell line, PC-V160, whereas mdr1 mRNA was not detected in any of the cell lines. The polymerase chain reaction, using mdr3-specific primers, excluded the possibility that low levels of P-glycoprotein expression contributed to the resistance phenotype in the low and intermediate-level resistant cell lines. Northern blot analysis using a human complementary DNA probe for the multidrug resistance-associated protein (MRP) demonstrated overexpression of murine mrp in each of the vincristine-selected sublines. Genomic amplification of the mrp gene was coincident with mrp overexpression. The expression of mrp was also examined in two series of previously characterized doxorubicin-selected cell lines derived from parental PC4 and C7D murine erythroleukemia cells. In contrast to the vincristine-selected cell lines, overexpression of mrp was not detected. These studies demonstrate that, in murine erythroleukemia cells selected for vincristine resistance, overexpression of murine mrp occurred prior to that for murine mdr. In contrast to human MRP, selection for vincristine, but not doxorubicin resistance, resulted in the overexpression of murine mrp.

Determination of the LDL receptor binding capacity of human lymphocytes by immunocytofluorimetric assay.

The determination of the LDL receptor binding capacity of human blood lymphocytes was assessed by indirect immunocytofluorimetric assay. To produce the maximal synthesis of the LDL receptor, the cholesterol efflux was enhanced by incubation of lymphocytes with HDL3 subfractions. The binding capacity of the LDL receptor was measured by incubation at 4 degrees C either with LDL and rabbit anti-LDL immunoglobulins or with peptide receptor antibody (ARP-Ig) raised against the NH2-terminal sequence of the LDL receptor. Thereafter complexes were incubated with fluorescein-labelled anti-rabbit immunoglobulin (FITC-Ig). Fluorescence flow cytometry was used to quantify the number of fluorescent lymphocytes and results were expressed as the percentage of lymphocytes with a fluorescent intensity above the threshold. Using preimmune rabbit immunoglobulin and then FITC-Ig, only 5-10% of cells were fluorescent. Neither LDL nor ARP-Ig could bind to homozygous familial hypercholesterolemia (FH) lymphocytes. Normal lymphocytes preincubated with HDL3 could bind LDL or ARP-Ig, the number of fluorescent cells being 59 and 39.2% respectively. Subjects with confirmed or suspected heterozygous FH demonstrated cell fluorescence at about half the normal level.

Heavy chain dominance in the binding of DNA by a lupus mouse monoclonal autoantibody.

Antibodies H241 and 2C10 are lupus mouse IgG autoantibodies that bind native DNA. In previous experiments, oligonucleotide antigens affinity-labeled both H and L chains of H241 but only the H chain of antibody 2C10. Primary structures of the V regions of the 2C10 H and L chains and the H241 L chain, determined from cDNA, help to explain the previous affinity-labeling experiments. The 2C10 L chain CDRs had several Asp residues and a net negative charge of five, whereas the 2C10 H chain CDRs had four Arg residues and a net positive charge of five. The L chain CDRs of H241 had a net positive charge of one. [The H241 H chain cDNA sequence was published previously by Gangemi et al. (1993) J. Immun. 151, 4660-4671]. Plasmid vectors were used for bacterial expression of H and L chains of 2C10 alone and in combinations in single chain Fv (scFv) molecules. The H chain alone bound native DNA as well as or better than the H-plus-L chain scFv. The H chain alone also bound Z-DNA. Combination of the 2C10 H chain with the L chain of an anti-Z-DNA antibody maintained the selectivity for Z-DNA, whereas its combination with the 2C10 L chain (in the 2C10 Fab) yielded selective B-DNA binding. The results with 2C10 match other examples in which the H chain is sufficient for DNA binding but selectivity is modulated by the L chain. The H chain binding to autoantigen may reflect selective events in early stages of B cell development.

Effect of phytosterols/stanols on LDL concentration and other surrogate markers of cardiovascular risk.

Plant sterols and stanols are well-known to reduce LDL-cholesterol (LDL-C) concentrations. It is generally accepted that supplementation with 2g/day of sterols/stanols leads to a 10% reduction in LDL. However, most of the clinical trials supporting this conclusion were of short-term duration, and the results of longer interventions are scanty. In four studies, interventions lasting>6 months were carried out and the LDL-C-lowering effects were maintained over this longer duration, although some results suggest that a reduced effect may be observed with sterols, while stanols maintain their effect. In any case, the data are too limited to be definitive. In a free-living population as well as in multiparametric interventional studies, however, the LDL-C-lowering effect has been confirmed, although to a lesser extent than in clinical studies. In the absence of data on cardiovascular morbidity and mortality, data for surrogate markers of cardiovascular risk could be considered adequate alternatives. Several studies have been conducted on this basis, but their results failed to demonstrate any favourable effects. The present report summarizes the different results obtained in long-term studies, and in those comparing the effects of sterols and stanols on lipids and other surrogate markers of cardiovascular risk.

Postprandial glycaemic and insulinaemic responses in adults after consumption of dairy desserts and pound cakes containing short-chain fructo-oligosaccharides used to replace sugars.

The present studies aimed to evaluate the glycaemic and insulinaemic responses, in healthy adults, to short-chain fructo-oligosaccharides (scFOS) from sucrose used to replace sugars in foods. Two study populations aged 18-50 years were recruited and they consumed dairy desserts or pound cakes containing either standard sugar content or scFOS to replace 30 % of the sugar content. For each study, the two products were tested once under a double-blind and cross-over design with at least 7 d between the two tests. Glucose and insulin were measured using standard methods in blood samples collected with a venous catheter for 120 min during a kinetic test. For the dairy desserts, replacing 30 % of the sugars with scFOS significantly reduced postprandial glycaemic (AUC0-120 min; P = 0·020) and insulinaemic (AUC0-120 min; P = 0·003) responses. For the pound cakes, the glycaemic response was not altered (AUC0-120 min; P =  0·322) while the insulinaemic response tended to be lower (AUC0-120 min; P = 0·067). This study showed that scFOS can be used to replace sugars with the benefit of lowering the postprandial glycaemic response without increasing the insulinaemic response. The effect might be modulated by other parameters (e.g. fat content) of the food matrices.

Comparison between fat intake assessed by a 3-day food record and phospholipid fatty acid composition of red blood cells: results from the Monitoring of Cardiovascular Disease-Lille Study.

We investigated the relationship between assessment of fatty acid intake by a 3-day food record and by capillary gas chromatography of erythrocyte phospholipid fatty acid. The study was performed in a sample of 244 men aged 45 to 66 years from the general population who were participating in the Monitoring of Cardiovascular Disease (MONICA)-Lille survey. The relationship between each nutrient and food item and erythrocyte phospholipid fatty acid was investigated by a regression model on proportion including each food item and nutrient as a dependent variable and percentage of fatty acid and covariables (nonalcoholic energy intake, age, alcohol intake, and smoking) as independent variables. Polyunsaturated fat and linoleic acid intake were positively correlated with linoleic acid content of erythrocytes (beta = 0.641 and 0.604, respectively, P < .001). Monounsaturated and saturated fat intake were correlated with oleic acid (beta = 0.375 and 0.373, respectively, P < .01). Fish intake correlated positively with docosahexaenoic acid (DHA) (beta = 0.383, P < .001) and negatively with arachidonic acid (beta = -0.509, P < .01). These data confirm, on a group level, a good relationship between assessment of polyunsaturated fat intake by a 3-day record and linoleic acid content of erythrocyte membranes. These data suggest that erythrocyte oleic acid content is a marker of both saturated and monounsaturated fat intake.

The relationship between the phospholipid fatty acid composition of red blood cells, plasma lipids, and apolipoproteins.

This study examined the relationship between the fatty acid composition of red blood cell phospholipids and lipid markers of atherosclerotic risk in an urban male population aged 45 to 66 years. There was a surprisingly significant positive association between the docosahexaenoic acid ([DHA] 22:6n-3) content of erythrocyte phospholipids and the following risk markers: plasma cholesterol (P < .01), low-density lipoprotein (LDL) cholesterol (P < .01), apolipoprotein (apo) B (P < .05), and apo B-containing lipoprotein particles (P < .05) recognized by a monoclonal antibody (LpBL3). On the other hand, phospholipid alpha-linolenate was positively correlated with apo A-I and high-density lipoprotein (HDL) cholesterol levels (P < .05), while arachidonate showed an inverse relationship with plasma cholesterol (P < .05). There was a negative association between palmitoleic acid and apo B (P < .01) and LpBL3 (P < .001); the latter showed a negative association with stearic acid (P < .001). These interesting findings emphasize the beneficial effect on atherosclerotic risk markers of dietary n-6 polyunsaturated and monounsaturated fatty acids, and suggest that long-chain n-3 polyunsaturated fatty acids (DHA) could have an adverse effect on some of the lipid risk markers.

Monocyte tissue factor response is decreased in patients with hyperlipidemia.

Monocytes are potent regulators of blood coagulation through the expression of tissue factor (TF) on stimulation and of tissue factor pathway inhibitor (TFPI), a selective inhibitor of TF pathway. As hyperlipidemia can modify some monocyte functions, we compared the TF and TFPI expression by circulating monocytes and the plasma TFPI levels between 65 healthy normolipemic controls and 38 nontreated hyperlipemic patients. TF and TFPI relationships with plasma lipoproteins are also examined. TF and TFPI expression were evaluated in peripheral mononuclear cells after isolation from blood by density gradient centrifugation and after short culture with or without lipopolysaccharide (LPS). TF and TFPI activity and antigen were measured in mononuclear cell lysates using amidolytic assay and enzyme-linked immunosorbent assay, respectively. TFPI activity and antigen were measured in plasma using the same methods. Plasma factor VII (FVII) activity and antigen were also determined. LPS-stimulated monocyte TF activity and antigen were lower in hyperlipidemic patients than in controls (0.0001

Serum Lp AI lipoprotein particles and other antiatherogenic lipid parameters in normolipidaemic obese subjects.

The android pattern of body fat distribution has been shown to increase the risk of metabolic and coronary heart disease. Protective lipid markers against cardiovascular disease were studied in 98 obese normolipidaemic, non diabetic, non-smoker subjects over 18 years of age according to regional distribution of adipose tissue as estimated by the waist ship ratio (WHR) and overall obesity as estimated by the body mass index (BMI). WHR was inversely correlated with Lp AI (r = 0.46) and HDL-cholesterol (r = 0.37). BMI was not correlated with protective lipid parameters but only with triglycerides. After adjustment, Lp AI was lower in men and in upper body obese women (p < 0.05). Lp AI is a better indicator of body fat distribution than HDL-cholesterol or apo AI, and its variations appear to be indirectly related to gender, menopause, and age, thereby influencing body fat distribution (the main factor accounting for Lp AI variation). Lp AI was inversely correlated with WHR in gluteal-femoral obese women but not in abdominally obese women or men, possibly because of a threshold effect.

Differential changes in genome structure and expression of the mdr gene family in multidrug-resistant murine erythroleukemia cell lines.

The genome structure and expression of mdr genes were examined in multidrug-resistant sublines of two different murine (DBA/2J) Friend erythroleukemia cell lines, PC4 and C7D, derived by stepwise exposure to increasing concentrations of adriamycin beginning with 5 ng/ml. The PC4 cell lines selected in higher drug concentrations (80-1280 ng/ml) demonstrated amplification of all three mdr genes with preferential amplification of mdr3. Overexpression of the mdr2 and mdr3 genes accompanied their genomic amplification; however, expression of mdr1 was not seen despite amplification. In the C7D cell lines selected with higher drug concentrations (40-160 ng/ ml), amplification and overexpression of mdr1 and mdr2 without mdr3 was observed. Increased expression of mdr1 occurred prior to gene amplification. The distribution of mdr-specific genes in micrococcal nuclease-generated chromatin fractions differing in transcriptionally active sequences and proteins was different between the parent and drug-resistant sublines. An enrichment (two- to threefold) of mdr3 genes in the H1-depleted mononucleosome fraction enriched for actively transcribed genes (e.g., globin) was detected by Southern analysis of chromatin fractions in PC4-80 cells (selected in 80 ng/ml of adriamycin and overexpressing mdr3), compared to the parental cells. mdr3 enrichment was also detected using a new PCR-based method, which examined mdr3 genes and repetitive sequences. Of note, the H1-depleted chromatin fraction from PC4-20 showed enrichment of the mdr3 gene, although mdr3 expression was not detected in the cell line. These studies showed a different pattern of gene amplification and overexpression in genetically related erythroleukemia cell lines selected for resistance to the same chemotherapeutic agent. A change in chromatin organization of mdr genes preceded overexpression and amplification of the mdr3 gene.