Development and validation of a Brief Diet Quality Assessment Tool in the French-speaking adults from Quebec.

BACKGROUND: The objective of this study was to develop and validate a short, self-administered questionnaire to assess diet quality in clinical settings, using the Alternative Healthy Eating Index (AHEI) as reference. METHODS: A total of 1040 men and women (aged 44.6 ± 14.4 y) completed a validated web-based food frequency questionnaire (webFFQ) and had their height and weight measured (development sample). Participants were categorized arbitrarily according to diet quality (high: AHEI score ≥ 65/110, low: AHEI score < 65/110) based on dietary intake data from the webFFQ. The Brief Diet Quality Assessment Tool was developed using a classification and regression tree (CART) approach and individual answers to the webFFQ among participants considered to have a plausible energy intake (ratio of reported energy intake to basal metabolic rate ≥ 1.2 and < 2.4; n = 1040). A second sample of 3344 older adults (aged 66.5 ± 6.4 y) was used to test the external validity of the Brief Diet Quality Assessment Tool (external validation sample). RESULTS: The decision tree included sequences of 3 to 6 binary questions, yielding 21 different pathways classifying diet quality as being high or low. In the development sample, the area under the receiver operating characteristic (ROC) curve of the predictive model was 0.92, with sensitivity, specificity and agreement values of 89.5, 83.9 and 87.2%. Compared with individuals having a low-quality diet according to the Brief Diet Quality Assessment Tool (mean AHEI 56.7 ± 11.4), individuals classified as having a high-quality diet (mean AHEI 71.3 ± 11.0) were significantly older, and had lower BMI, percent body fat and waist circumference, and had lower blood pressure, triglycerides, cholesterol/HDL ratio and fasting insulin as well as higher HDL-cholesterol concentrations (all P < 0.05). Similar results were observed in the external validation sample, although overall performance of the Brief Diet Quality Assessment Tool was slightly lower than in the development sample, with an area under the ROC curve of 0.79 and sensitivity, specificity and agreement values of 73.0, 69.0 and 71.3%, respectively. CONCLUSION: The CART approach yielded a simple and rapid Brief Diet Quality Assessment Tool that identifies individuals at risk of having a low-quality diet. Further studies are needed to test the performance of this tool in primary care settings.

Assessment of the American Heart Association's "Life's simple 7" score in French-speaking adults from Québec.

BACKGROUND AND AIMS: The "Life's Simple 7" (LS7) metrics were developed by the American Heart Association (AHA) to assess and promote cardiovascular health in the American population. The purpose of this study was to assess the overall cardiovascular health of French-speaking adults from the Province of Quebec using the LS7 score. METHODS AND RESULTS: A total of 777 age and sex-representative participants of five different administrative regions in the Province of Quebec (387 men and 390 women; mean age ± SEM: 41.9 ± 0.1 years) were included in these analyses. Metrics of the LS7 score (smoking, physical activity, diet, body mass index, blood pressure, fasting total cholesterol and blood glucose) were analysed to generate a final score ranging from 0 to 7. Only 0.5% of participants met all criteria for ideal cardiovascular health. The diet metric showed the lowest prevalence of "ideal" scores (4.8%) whereas not smoking was the metric with the highest prevalence (88.1%). Women had a higher LS7 score than men, while age and education level (negative and positive association, respectively; p < 0.0001) were also associated with the LS7 score. CONCLUSION: Consistent with studies conducted among other populations, very few French-speaking adults from the Province of Quebec achieve an ideal cardiovascular health. These data indicate that further public health efforts aimed at promoting the LS7 metrics, focusing primarily on diet, are urgently needed. Specific groups, including older adults and those with lower levels of education, should be targeted when developing cardiovascular health promotion interventions.

[Characteristics associated with depression and post-traumatic stress disorder among childhood sexual abuse women]. / Caractéristiques associées à la dépression et aux symptômes de stress post-traumatique chez les femmes victimes d'agression sexuelle durant l'enfance.

BACKGROUND: A significant proportion of childhood sexual abuse victims suffer from psychological sequelae in adulthood. Factors that provide a better understanding for the reasons why some victims develop these sequelae remain under-explored. In this context, the main objective is to examine the specific contribution of the contextual characteristics of childhood sexual abuse, multitype childhood maltreatment and adolescent suicide attempts on the development of depression and post-traumatic stress disorder in adulthood among sexually abused women as children. A secondary objective aimed to establish the prevalence of various forms of childhood maltreatment, adult onset post-traumatic stress disorder and depression among those women. METHODS: The sample included 479 women victims of childhood sexual abuse who participated in two separate surveys taken by women in the province of Quebec. RESULTS: More than half of these women reported at least one other form of childhood maltreatment, 30% of them presented post-traumatic disorder and 40% suffered from depression in adulthood. Regression analysis indicates that post-traumatic stress disorder was associated with early onset childhood sexual abuse and intergenerational continuity of sexual victimization, as well as childhood physical maltreatment and negligence. Depression was associated with childhood psychological maltreatment and negligence, a non-supportive response following child sexual abuse related disclosure and suicide attempt in adolescence. CONCLUSION: These results confirm the need to consider the cumulative effects of various childhood adversity factors in the psychosocial assessment of sexually abused women in early life, thus helping to better understand and treat their psychological sequelae.

Comparison of a Mediterranean to a low-fat diet intervention in adults with type 1 diabetes and metabolic syndrome: A 6-month randomized trial.

BACKGROUND AND AIMS: The metabolic syndrome (MS) is an emerging complication in patients with type 1 diabetes (T1D), with no preventive or therapeutic treatment reported yet. We wanted to compare the impact of two 6-month nutritional interventions, based on a Mediterranean (MED) or a low-fat diet, on waist circumference, anthropometric and metabolic outcomes in patients with both T1D and the MS. METHODS AND RESULTS: Participants were randomized into 2 intervention groups: 1) MED-diet or 2) low-fat diet. The 6-month study included 9 teaching sessions with a registered dietitian. Anthropometric (primary outcome: waist circumference), metabolic and nutritional assessments were performed at inclusion, 3 and 6-month. We used mixed effects models to assess the effects of both interventions. 28 participants were included (50.9 ± 10.3 years old) with a mean BMI of 30.7 ± 3.3 kg/m2 and a waist circumference of 105.5 ± 8.9 cm at inclusion. A trend towards a greater reduction of dietary fat intakes in the low-fat diet group was observed (P-interaction = 0.09). Waist circumference was reduced at 6-month in both groups (-3.5 cm low-fat; -1.5 cm MED-diet) with no significant difference between groups (P-interaction = 0.43). Body mass index also significantly decreased in both groups (-0.7 kg/m2 low-fat; -1.1 kg/m2 MED-diet; P-interaction = 0.56). No significant differences between groups were observed for other metabolic parameters. CONCLUSIONS: This study suggests that a 6-month non-restrictive dietary intervention in patients with T1D and MS could contribute to weight management, without significant differences between interventions for anthropometric and metabolic parameters. Further studies should investigate the long-term benefits of these diets. CLINICAL TRIAL REGISTRY: NCT02821585 (https://clinicaltrials.gov/).

Efficacy and safety of antihyperglycaemic drug regimens added to metformin and sulphonylurea therapy in Type 2 diabetes: a network meta-analysis.

AIM: To assess the efficacy and safety of third-line adjuvant antihyperglycaemic agents in people with Type 2 diabetes mellitus failing metformin and sulphonylurea combination therapy. METHODS: We searched MEDLINE, CENTRAL, clinicaltrials.gov and regulatory websites, and conducted a manual search of references in the identified studies. Randomized trials evaluating antihyperglycaemic agents in adults with Type 2 diabetes experiencing poor glycaemic control despite optimized metformin and sulphonylurea therapy (≥ 1500 mg metformin or maximum tolerated dose; ≥ 50% of maximum sulphonylurea dose for ≥ 3 weeks) were included. Data extraction included: study characteristics; change in HbA1c concentration; weight; systolic blood pressure; and relative risk of hypoglycaemia, urinary tract infections; and genital tract infections. A network meta-analysis was performed. RESULTS: A total of 20 trials evaluating 13 antihyperglycaemic agents were included. Compared with placebo/control, all antihyperglycaemic agents reduced HbA1c levels, albeit by differing magnitudes [range 7 mmol/mol (0.6%) for acarbose to 13 mmol/mol (1.20%) for liraglutide]. Sodium glucose cotransporter-2 inhibitors reduced weight (1.43-2.07 kg) whereas thiazolidinediones, glargine and sitagliptin caused weight gain (1.48-3.62 kg) compared with placebo/control. Sodium glucose cotransporter-2 inhibitors, rosiglitazone and liraglutide decreased systolic blood pressure compared with placebo/control, pioglitazone, glargine and sitagliptin (2.41-8.88 mm Hg). Glargine, thiazolidinediones, liraglutide, sitagliptin and canagliflozin increased hypoglycaemia risk compared with placebo/control (relative risk 1.92-7.47), while glargine and rosiglitazone increased hypoglycaemia compared with most antihyperglycaemic agents (relative risk 2.81-7.47). No antihyperglycaemic agent increased the risk of urinary tract infection, but canagliflozin increased the risk of genital tract infection by 3.9-fold compared with placebo/control. CONCLUSIONS: When added to metformin and a sulphonylurea, antihyperglycaemic agents had varying effects on efficacy and safety endpoints. These conclusions should be considered when clinicians choose between possible adjunctive agents.

Gender differences in dietary intakes: what is the contribution of motivational variables?

BACKGROUND: Differences between men and women with respect to dietary intakes and eating behaviours have been reported and could be explained by gender differences in motivational variables associated with the regulation of food intake. The main objectives of the present study were to identify gender differences in dietary intakes, eating behaviours and motivational variables and to determine how motivational variables were associated with dietary intakes and eating behaviours in men and women. METHODS: Sixty-four men and 59 premenopausal women were included in the present study and presented cardiovascular risk factors. The Regulation of Eating Behaviours scale was completed to assess motivational variables. A validated food frequency questionnaire was administered to evaluate dietary intakes and subjects completed the Three-Factor Eating questionnaire to assess eating behaviours. RESULTS: Men had higher energy intake, energy density and percentage of energy from lipids and lower percentage of energy from carbohydrates than women (P ≤ 0.04). Men also had a lower emotional susceptibility to disinhibition than women (P = 0.0001). Women reported a higher score for eating-related self-determined motivation [i.e., eating-related self-determination index (SDI)] than men (P = 0.002). The most notable gender difference in the pattern of associations was that eating-related SDI was negatively associated with energy density (r = -0.30; P = 0.02), only in women. CONCLUSIONS: Women had a better dietary profile and higher eating-related SDI than men. However, gender differences in dietary variables might be explained by a potential gender-specific pattern of association of eating-related SDI with dietary intakes and eating behaviours.

Bright squeezed-vacuum source with 1.1 spatial mode.

Bright squeezed vacuum, a macroscopic nonclassical state of light, can be obtained at the output of a strongly pumped nonseeded traveling-wave optical parametric amplifier (OPA). By constructing the OPA of two consecutive crystals separated by a large distance, we make the squeezed vacuum spatially single-mode without a significant decrease in the brightness or squeezing.

The TGF-ß-Smad3 pathway inhibits CD28-dependent cell growth and proliferation of CD4 T cells.

Transforming growth factor-ß (TGF-ß) maintains self-tolerance through a constitutive inhibitory effect on T-cell reactivity. In most physiological situations, the tolerogenic effects of TGF-ß depend on the canonical signaling molecule Smad3. To characterize how TGF-ß/Smad3 signaling contributes to maintenance of T-cell tolerance, we characterized the transcriptional landscape downstream of TGF-ß/Smad3 signaling in resting or activated CD4 T cells. We report that in the presence of TGF-ß, Smad3 modulates the expression of >400 transcripts. Notably, we identified 40 transcripts whose expression showed Smad3 dependence in both resting and activated cells. This 'signature' confirmed the non-redundant role of Smad3 in TGF-ß biology and identified both known and putative immunoregulatory genes. Moreover, we provide genomic and functional evidence that the TGF-ß/Smad3 pathway regulates T-cell activation and metabolism. In particular, we show that TGF-ß/Smad3 signaling dampens the effect of CD28 stimulation on T-cell growth and proliferation. The impact of TGF-ß/Smad3 signals on T-cell activation was similar to that of the mTOR inhibitor Rapamycin. Considering the importance of co-stimulation on the outcome of T-cell activation, we propose that TGF-ß-Smad3 signaling may maintain T-cell tolerance by suppressing co-stimulation-dependent mobilization of anabolic pathways.

Plasma adiponectin concentration is strongly associated with VLDL-TG catabolism in postmenopausal women.

BACKGROUND AND AIMS: To investigate associations between plasma adiponectin concentration and very-low density lipoprotein-triglyceride (VLDL-TG) secretion and catabolism in postmenopausal women. METHODS AND RESULTS: This cross-sectional study included 30 postmenopausal women. Plasma adiponectin concentration was measured by ELISA. Insulin sensitivity was assessed by a 2-h euglycemic-hyperinsulinemic clamp. Fasting plasma glucose (FPG) and 2-hour plasma glucose (2hPG) were measured during an oral glucose tolerance test. The calculation of VLDL-TG fractional catabolic rate (FCR) and VLDL-TG total secretion rate (TSR) were based on the monoexponential decrease of TG-[²H5] glycerol values obtained following the administration of a ²H5-glycerol bolus. Plasma adiponectin concentration was negatively associated with VLDL-TG TSR (r=-0.50; p=0.005) and positively associated with VLDL-TG FCR (r=0.54; p<0.002). This latter association remained significant after further adjustments for insulin sensitivity, visceral adipose tissue, HDL-C, FPG and 2hPG concentrations. In a multivariate model including adiponectin, insulin sensitivity and 2hPG, plasma adiponectin level was the strongest correlate of VLDL-TG FCR. CONCLUSIONS: Elevated plasma adiponectin concentration is associated with a favourable VLDL-TG metabolism.

Heterogeneity among Ly-49C natural killer (NK) cells: characterization of highly related receptors with differing functions and expression patterns.

Ly-49C is a member of the polymorphic family of murine NK cell inhibitory receptors. The 5E6 antibody that defines a subset of NK cells responsible for the rejection of parental H-2d bone marrow by F1 mice has been shown previously to react with Ly-49C. Here, the 5E6 antibody was found to detect two Ly-49C-related molecules in B6 mice. Two cDNA clones were isolated from B6 NK cells, one identical to previously reported Ly-49CB6 and the other a novel cDNA. The deduced amino acid sequence of the latter differs from that of Ly-49CBALB at only 4 residues, whereas the previously reported Ly-49CB6 differs at 22 residues. Flow cytometric analyses of COS cells transfected with the two cDNAs showed that the 5E6 antibody binds to both Ly-49 molecules, while another anti-Ly-49C antibody, 4LO3311, binds to the newly described Ly-49C but not the previously reported Ly-49CB6. Two-color flow cytometric analysis detected 5E6+4LO3311- as well as 5E6+4LO3311+ subsets of NK cells from B6, but not BALB/c, mice. The level of Ly-49C expression on B6 NK cells detected by the 4LO3311 antibody was substantially lower than that on BALB/c NK cells. Binding specificity of the novel Ly-49CB6 was indistinguishable from that of Ly-49CBALB, whereas no binding was detectable with previously reported Ly-49CB6. These results demonstrate that the newly described Ly-49CB6, not the previously reported Ly-49CB6, is the probable B6 allelic form of Ly-49C. The previously reported Ly-49CB6 must be encoded by a separate gene and should be renamed Ly-49I. The implication of these results with respect to the role of Ly-49C in hybrid resistance is discussed.

Couple dissatisfaction and eating profile: a mediation effect of coping style.

Individuals' ways of coping with psychological stress have often been associated with body weight regulation through their impact on eating behaviours. In particular, emotion-oriented and distraction-oriented coping styles have been steadily related to disordered eating. Couple dissatisfaction may be experienced as an important psychological stressor and could therefore affect eating behaviours through the use of inadequate coping strategies. The study proposes 1) to compare women reporting a low vs a high level of couple satisfaction, and 2) to test mediational models including couple satisfaction, coping styles, and eating variables. Analyses were performed among 65 overweight/obese premenopausal women who reported being weight-preoccupied. Women exhibiting couple dissatisfaction (34.8%) showed a higher level of EDE-Q restraint, more intense concerns about eating and shape, a higher level of disinhibition and susceptibility to hunger and endorsed more often a distraction-oriented coping style, independently of their body weight. Furthermore, distraction- oriented coping style seemed to be a valid mediator of the relation between couple dissatisfaction and eating behaviours. Since non-normative eating behaviours, namely disinhibition and susceptibility to hunger, have been particularly linked to a higher body weight status, it is relevant to extend the scope of interest to more distal contributing factors, such as couple dissatisfaction.

Impact of high-fat /low-carbohydrate, high-, low-glycaemic index or low-caloric meals on glucose regulation during aerobic exercise in Type 2 diabetes.

AIMS: A decrement in blood glucose (BG) may be observed in patients with Type 2 diabetes (T2DM) when exercise is performed after a meal, in contrast to fasting. We determined the impact of different pre-exercise meal macronutrient compositions with modulation of the glycaemic index (GI) on glucose regulation during exercise in patients with T2DM. METHODS: Using a randomized, single-blind crossover design, 10 sedentary men performed five exercise sessions, once after an overnight fast, and also after each of four test meals, consisting of a high-fat/low-carbohydrate meal, a high-GI meal, a low-GI meal, and a low-calorie meal. RESULTS: Pre-exercise BG and insulin levels were comparable for all four meals. Exercise decreased BG and insulin levels during all meal conditions (all P < 0.001) compared with the fasting state in which BG levels did not change. The magnitude of BG and insulin decrements was similar after consuming the low-calorie, the high-GI and the high-fat/low-carbohydrate meals, whereas the low-GI meal induced the lowest BG fall. Adrenaline response was higher after consumption of the high-, the low-GI and the low-caloric meals compared with the high-fat/low-carbohydrate meal and with the fasting state (P < 0.05). CONCLUSIONS: This study underlines the beneficial effect of low-GI foods and the differential impact of pre-exercise meal macronutrient composition on BG decrease. This may protect against exercise-induced hypoglycaemia, and reiterates the safety of exercising while fasting in T2DM patients.

Associations between self-reported vegetable and fruit intake assessed with a new web-based 24-h dietary recall and serum carotenoids in free-living adults: a relative validation study.

The aim of the present study was to assess the relative validity of a new web-based 24-h dietary recall (R24W) in terms of vegetable and fruit (VF) intake assessment using serum carotenoid concentrations as reference biomarkers. A total of seventy-four women and seventy-three men (mean age 47·5 (sd 13·3) years; mean BMI 25·5 (sd 4·4) kg/m2) completed the R24W four times to assess their VF intake. Serum carotenoids were obtained from 12-h fasted blood samples and measured by HPLC. Raw and de-attenuated partial Spearman's correlations were performed to determine how usual vegetable and/or fruit intake was associated with serum carotenoids. Relevant confounders were selected using a stepwise regression analysis. Finally, cross-classification was used to determine agreement between intake of VF and serum carotenoids. Intake of total dietary carotenoids was significantly associated (r 0·40; P < 0·01) with total serum carotenoids (without lycopene). Total VF intake was also associated with total serum carotenoid concentrations without lycopene (r 0·44; P < 0·01). HDL-cholesterol, waist circumference and age were identified as confounders in the association between total VF intake and total serum carotenoids (without lycopene). De-attenuated partial correlation adjusted for these confounders increased the associations between dietary carotenoids and total serum carotenoids without lycopene (r 0·49; P < 0·01) and between total VF intake and total serum carotenoids without lycopene (r 0·48; P < 0·01). Almost 80 % of respondents were classified in the same or the adjacent quartile for total VF intake and total serum carotenoids without lycopene, while less than 6 % were classified in the opposite quartile. Overall, these observations support the appropriateness of the R24W to assess the dietary intake of VF.

Effect of a nutritional intervention promoting the Mediterranean food pattern on anthropometric profile in healthy women from the Québec city metropolitan area.

OBJECTIVE: To evaluate the impact of a nutritional intervention promoting a Mediterranean food pattern on anthropometric profile in healthy women. DESIGN: Nutritional intervention study. SETTING: Laval University, Canada. SUBJECTS: Seventy-seven healthy women started the study and four did not complete the study. METHODS: A 12-week nutritional intervention in free-living conditions consisted of two group courses on nutrition and seven individual sessions with a dietitian. A follow-up visit was performed 12 weeks after the end of the intervention (week 24). A Mediterranean dietary score (MedScore), based on the 11 components of the Mediterranean pyramid, was established to evaluate the adherence to the Mediterranean food pattern. RESULTS: Small but significant decreases in body weight and waist circumference were observed after 12 weeks of intervention (0.5 kg and 1.2 cm, respectively (P<0.01)). Increase in partial MedScore for legumes, nuts and seeds (increase in consumption) as well as increase in partial MedScore for sweets (decrease in consumption) were significantly associated with changes in waist circumference (r=-0.36, P=0.001; r=0.24, P=0.05, respectively). No association was observed between changes in anthropometric profile and changes in the consumption of olive oil. CONCLUSION: Changes in dietary food pattern, more specifically an increase in the consumption of legumes, nuts and seeds, and a decrease in the consumption of sweets, were associated with some beneficial changes in anthropometric profile.

A novel approach for the identification of efficient combination therapies in primary human acute myeloid leukemia specimens.

Appropriate culture methods for the interrogation of primary leukemic samples were hitherto lacking and current assays for compound screening are not adapted for large-scale investigation of synergistic combinations. In this study, we report a novel approach that efficiently distills synthetic lethal interactions between small molecules active on primary human acute myeloid leukemia (AML) specimens. In single-dose experiments and under culture conditions preserving leukemia stem cell activity, our strategy considerably reduces the number of tests needed for the identification of promising compound combinations. Initially conducted with a selected library of 5000 small molecules and 20 primary AML specimens, it reveals 5 broad classes of sensitized therapeutic target pathways along with their synergistic patient-specific fingerprints. This novel method opens new avenues for the development of AML personalized therapeutics and may be generalized to other tumor types, for which in vitro cancer stem cell cultures have been developed.

Transcriptome analysis of G protein-coupled receptors in distinct genetic subgroups of acute myeloid leukemia: identification of potential disease-specific targets.

Acute myeloid leukemia (AML) is associated with poor clinical outcome and the development of more effective therapies is urgently needed. G protein-coupled receptors (GPCRs) represent attractive therapeutic targets, accounting for approximately 30% of all targets of marketed drugs. Using next-generation sequencing, we studied the expression of 772 GPCRs in 148 genetically diverse AML specimens, normal blood and bone marrow cell populations as well as cord blood-derived CD34-positive cells. Among these receptors, 30 are overexpressed and 19 are downregulated in AML samples compared with normal CD34-positive cells. Upregulated GPCRs are enriched in chemokine (CCR1, CXCR4, CCR2, CX3CR1, CCR7 and CCRL2), adhesion (CD97, EMR1, EMR2 and GPR114) and purine (including P2RY2 and P2RY13) receptor subfamilies. The downregulated receptors include adhesion GPCRs, such as LPHN1, GPR125, GPR56, CELSR3 and GPR126, protease-activated receptors (F2R and F2RL1) and the Frizzled family receptors SMO and FZD6. Interestingly, specific deregulation was observed in genetically distinct subgroups of AML, thereby identifying different potential therapeutic targets in these frequent AML subgroups.

Proteogenomic-based discovery of minor histocompatibility antigens with suitable features for immunotherapy of hematologic cancers.

Pre-clinical studies have shown that injection of allogeneic T cells primed against a single minor histocompatibility antigen (MiHA) could cure hematologic cancers (HC) without causing any toxicity to the host. However, translation of this approach in humans has been hampered by the paucity of molecularly defined human MiHAs. Using a novel proteogenomic approach, we have analyzed cells from 13 volunteers and discovered a vast repertoire of MiHAs presented by the most common HLA haplotype in European Americans: HLA-A*02:01;B*44:03. Notably, out of >6000 MiHAs, we have identified a set of 39 MiHAs that share optimal features for immunotherapy of HCs. These 'optimal MiHAs' are coded by common alleles of genes that are preferentially expressed in hematopoietic cells. Bioinformatic modeling based on MiHA allelic frequencies showed that the 39 optimal MiHAs would enable MiHA-targeted immunotherapy of practically all HLA-A*02:01;B*44:03 patients. Further extension of this strategy to a few additional HLA haplotypes would allow treatment of almost all patients.

Two distinct protein isoforms are encoded by ntk, a csk-related tyrosine protein kinase gene.

Recently, we and others have cloned cDNAs encoding a second member of the Csk family of inhibitory tyrosine protein kinases, which we have termed Ntk. Intriguingly, the mouse ntk cDNA sequences published by two independent groups differed by the presence or absence of a 136 nucleotide-insert near their 5' ends. In this report, we demonstrate that this 136 nucleotide-sequence likely corresponds to a complete exon in the ntk gene (termed exon 2), and that the two types of cDNAs/transcripts are produced by alternative splicing. Using ribonuclease protection assays, it was also established that brain and lymphoid organs, as well as most hemopoietic cells, predominantly expressed ntk transcripts lacking exon 2. In contrast, selected hemopoietic cell lines, such as the immature myeloid cell lines 32D cl3(G) and WEHI-3B, exclusively possessed exon 2-bearing RNAs. Interestingly, exon 2 introduced a novel in-frame upstream AUG in the ntk transcript, which is in the appropriate context for translation initiation. Evidence was obtained that this AUG is utilized in vivo, and that it extends the amino-terminal sequence of Ntk by 40 amino acids. Indeed, while exon 2-deficient ntk RNAs were translated into a 52 kilodalton (kDa) polypeptide (p52ntk), those bearing exon 2 produced a 56 kDa protein (p56ntk). Furthermore, p56ntk, but not p52ntk, was recognized by an antiserum directed against the novel amino-terminal sequence encoded by exon 2. Additional biochemical characterizations showed that p52ntk and p56ntk were localized to the cytoplasm, and that they partially accumulated in the detergent-insoluble cellular fraction. This last finding suggested that the Ntk proteins can associate with the cytoskeleton. Finally, through linkage analysis of two multilocus crosses, the ntk gene was mapped to Chromosome 10 in the mouse. Taken together, these data showed that ntk, a csk-related tyrosine protein kinase gene, encodes two protein isoforms expressed in distinct cell types. Moreover, they raised the possibility that Ntk may be involved in the regulation of Src-like enzymes in detergent-insoluble cellular compartments.

Effects of acute changes of plasma free fatty acids on intramyocellular fat content and insulin resistance in healthy subjects.

The reason for the 3- to 4-h delay between a rise in plasma free fatty acid (FFA) levels and the development of insulin resistance remains unknown. In the current study, we have tested the hypothesis that the delay may be caused by the need for plasma FFAs to first enter muscle cells and to be re-esterified there before causing insulin resistance. To this end, we have determined intramyocellular triglyceride (IMCL-TG) content with proton nuclear magnetic resonance ((1)H-NMR) spectroscopy in healthy volunteers before and 4 h after lowering of plasma FFAs (with euglycemic-hyperinsulinemic clamping) or after increasing plasma FFAs (with lipid plus heparin infusions). Increasing plasma FFAs (from 516 to 1,207 micromol/l or from 464 to 1,857 micromol/l, respectively) was associated with acute increases in IMCL-TG from 100 to 109 +/- 5% (P < 0.05) or to 133 +/- 11% (P < 0.01), respectively, and with a significant increase in insulin resistance (P < 0.05 after 3.5 h). Lowering of plasma FFAs from 560 to 41 micromol/l was associated with a tendency for IMCL-TG to decrease (from 100 to 95 +/- 3%). Changes in plasma FFAs correlated linearly with IMCL-TG (r = 0.74, P < 0.003). The demonstration that acute changes in plasma FFAs were accompanied by corresponding changes in IMCL-TG and with the development of insulin resistance, taken together with previous reports of a close correlation between IMCL-TG and insulin resistance, supported the notion that accumulation of IMCL-TG is a step in the development of FFA-induced insulin resistance.