Biomedical subjects

Katherine Macé

Publications and source records attributed to Katherine Macé.

11 recordsLinked to original sources

PubMed2006

Long-term cultured human myotubes decrease contractile gene expression and regulate apoptosis-related genes.

The present study examined time-dependent changes in the gene expression profile of long-term cultured human myotubes. Microarray transcriptional analysis was performed in a primary culture of differentiated myotubes from one subject over seven weeks. This analysis showed a main gradual fall in genes of the contractile apparatus, and a broad upregulation of genes involved in cell development and growth, followed by stress response and signal transduction. Glucose metabolism was also monitored, but no significant alterations in glucose uptake, oxidation or glycogen storage were observed. Mitochondrial membrane potential, or the amount of membrane lipid peroxides, remained similarly unchanged, nor was lactate dehydrogenase leakage observed. Time-dependent changes in eight genes were validated by real-time RT-PCR in primary cultured myotubes from four subjects, of similar age and isolated after equivalent replication cycles in vitro and differentiated over seven weeks. Insulin-like growth factor-binding protein 2 (IGFBP2), a modulator of the IGF signal, was upregulated. The antiapoptotic gene heat-shock 70-kd protein 2 (HSPA2) was induced, whereas the proapoptotic tumor necrosis factor receptor superfamily, member 25 (WSL-1) was suppressed. A decline in the muscle-specific gene M-cadherin and contraction genes, such as slow-twitch troponin I (TNNI1) and myosin heavy chain 2 (MYH2), myosin light chain 1 (MYL1) and myosin-binding protein H (MYBPH), which are expressed in adult fast-twitch muscle, was shown. In summary, these data demonstrate extensive downregulation of contractile genes and modulation of apoptosis-related genes, in favour of cell survival, during maintenance of cultured human myotubes.

Adolescent↗

PubMed2006

Effect of sucrose and saturated-fat diets on mRNA levels of genes limiting muscle fatty acid and glucose supply in rats.

In this study, we examined whether the increased availability of lipids in blood resulting from two types of diet manipulation regulated metabolic gene expression in the skeletal muscle of rats. Feeding for 4 wk on an isocaloric-sucrose or a hypercaloric-fat diet increased plasma TAG in the fed condition by increments of 70 and 40%, respectively, and increased fasting insulinemia (approximately 3-fold) compared with a starch diet. The fat diet impaired glucose tolerance and caused obesity, whereas sucrose-fed rats maintained their normal weight. We analyzed the expression of genes that regulate the exogenous FA supply (LPL, FAT/CD36, FATP1), synthesis (ACC1), glucose (GLUT4, GLUT1, HK2, GFAT1, glycogen phosphorylase) or glycerol (glycerol kinase) provision, or substrate choice for oxidation (PDK4) in gastrocnemius and soleus muscles at the end of the glucose tolerance test. LPL, FAT/CD36, FATP1, PDK4, and GLUT4 mRNA as well as glycogen phosphorylase and glycerol kinase activity levels in both muscles were unchanged by the diets. Increased mRNA levels of GLUT1 (1.6- and 2.6-fold, respectively) and GFAT1 (about 1.7-fold) in gastrocnemius, and of ACC1 (about 1.5-fold) in soleus, were found in both the sucrose and fat groups. In the fat group, HK2 mRNA was also higher (1.8-fold) in the gastrocnemius. Both sucrose and saturated-fat diets prompted hyperinsulinemia and hyperlipemia in rats. These metabolic disturbances did not alter the expression of LPL, FAT/CD36, FATP1, PDK4, and GLUT4 genes or glycogen phosphorylase and glycerol kinase activity levels in either analyzed muscle. Instead, they were linked to the coordinated upregulation in gastrocnemius of genes that govern glucose uptake and the hexosamine pathway, namely, GLUT1 and GFAT1, which might contribute to insulin resistance.

Animals↗

PubMed2006

Protein quality and quantity in cow's milk-based formula for healthy term infants: past, present and future.

The development of infant formula with optimized protein quality and quantity has been, and still is, the subject of intense investigation. A better understanding of the protein composition of breast milk and infant needs in association with technological breakthroughs in cow's milk fractionation, has led to the development of infant formulas with a protein content that is closer to that of human milk. Today, infant formulas with a protein/energy ratio of 1.8 g/100 kcal are commercially available. These formulas have been shown to be safe and nutritionally adequate for term infants. However, the short-term and potentially long-term metabolic benefits of formulas with reduced protein content have still to be elucidated and are currently under investigation. In addition to providing amino acids as building blocks for growth, milk is the source of numerous bioactive factors/hormones which are involved in multiple physiological processes. Continuous efforts are being made to identify new bioactive compounds in human milk. However, a better understanding of their biological functions in suckling infants as well as a comparison with their bovine counterparts are needed. Technological processes, which preserve some bioactive factors in cow's milk already exist. These processes could be applied to infant formulas.

Animals↗

PubMed2005

Liver X receptor preferentially activates de novo lipogenesis in human preadipocytes.

The liver X receptor (LXR) was demonstrated to play a key role in cholesterol metabolism in liver, intestine and macrophage. However, its function on the regulation of preadipocyte differentiation remains unclear since contradictory results were reported. The objective of the present study was to unravel the functionality of LXR in human preadipocytes. We show that the LXR agonist T0901317 strongly stimulated the expression of SREBP-1c and the lipogenic enzymes ACC-1, FAS and SCD-1 in both the human preadipose cell line Chub-S7 as well as human primary stromal vascular fraction (SVF) cells. The effects on gene expression were associated with the stimulation of de novo lipogenesis in both cell models, resulting in the induction of lipid accumulation. In contrast with a PPARgamma agonist (BRL49653), T0901317 enhanced only slightly the expression of PPARgamma dependent genes (PPARgamma, aP2 and adiponectin) in Chub-S7 cells and failed to change their expression in human SVF cells. These results show that LXR stimulated preferentially triglyceride accumulation in human preadipocytes via the induction of de novo lipogenesis, rather than activating the differentiation process through PPARgamma activation.

Adipocytes↗

PubMed2005

Aspergillus oryzae produces compounds inhibiting cholesterol biosynthesis downstream of dihydrolanosterol.

The formation of cholesterol synthesis inhibiting molecules by five different strains of the koji mold Aspergillus oryzae was studied. After growing these strains on a complex liquid medium we found in crude organic phase extracts and specific fractions there from compounds inhibiting cholesterol synthesis in human hepatic T9A4 cells in vitro at enzyme sites downstream of dihydrolanosterol. This was evidenced by using different radioactively labeled precursors, namely acetate, mevalonate, 24,25-dihydro-[24,25-(3)H2]-lanosterol or [3-(3)H]-lathosterol.

Aspergillus oryzae↗

PubMed2004

beta3-adrenoceptor agonist prevents alterations of muscle diacylglycerol and adipose tissue phospholipids induced by a cafeteria diet.

BACKGROUND: Insulin resistance induced by a high fat diet has been associated with alterations in lipid content and composition in skeletal muscle and adipose tissue. Administration of beta3-adrenoceptor (beta3-AR) agonists was recently reported to prevent insulin resistance induced by a high fat diet, such as the cafeteria diet. The objective of the present study was to determine whether a selective beta3-AR agonist (ZD7114) could prevent alterations of the lipid profile of skeletal muscle and adipose tissue lipids induced by a cafeteria diet. METHODS: Male Sprague-Dawley rats fed a cafeteria diet were treated orally with either the beta3-AR agonist ZD7114 (1 mg/kg per day) or the vehicle for 60 days. Rats fed a chow diet were used as a reference group. In addition to the determination of body weight and insulin plasma level, lipid content and fatty acid composition in gastronemius and in epididymal adipose tissue were measured by gas-liquid chromatography, at the end of the study. RESULTS: In addition to higher body weights and plasma insulin concentrations, rats fed a cafeteria diet had greater triacylglycerol (TAG) and diacylglycerol (DAG) accumulation in skeletal muscle, contrary to animals fed a chow diet. As expected, ZD7114 treatment prevented the excessive weight gain and hyperinsulinemia induced by the cafeteria diet. Furthermore, in ZD7114 treated rats, intramyocellular DAG levels were lower and the proportion of polyunsaturated fatty acids, particularly arachidonic acid, in adipose tissue phospholipids was higher than in animals fed a cafeteria diet. CONCLUSIONS: These results show that activation of the beta3-AR was able to prevent lipid alterations in muscle and adipose tissue associated with insulin resistance induced by the cafeteria diet. These changes in intramyocellular DAG levels and adipose tissue PL composition may contribute to the improved insulin sensitivity associated with beta3-AR activation.

Journal Article↗

PubMed2004

Fiber type- and fatty acid composition-dependent effects of high-fat diets on rat muscle triacylglyceride and fatty acid transporter protein-1 content.

Intramuscular triacylglyceride (TAG) is considered an independent marker of insulin resistance in humans. Here, we examined the effect of high-fat diets, based on distinct fatty acid compositions (saturated, monounsaturated or n-6 polyunsaturated), on TAG levels and fatty acid transporter protein (FATP-1) expression in 2 rat muscles that differ in their fiber type, soleus, and gastrocnemius; the relationship to whole body glucose intolerance was also studied. Compared with carbohydrate-fed rats, the groups subjected to any one of the high-fat diets consistently exhibited enhanced body weight gain and adiposity, elevated plasma free fatty acids and TAG in the fed condition, hyperinsulinemia, and glucose intolerance. TAG content was consistently higher in soleus than in gastrocnemius, but was only significantly elevated by the n-6 polyunsaturated-based diet. FATP-1 levels in soleus were double those in gastrocnemius muscle in carbohydrate-fed animals. High-fat diets caused an elevation in FATP-1 protein content in soleus, but a reduction in gastrocnemius. In conclusion, the hyperinsulinemic hyperlipidemic condition upregulates FATP-1 expression in soleus and downregulates that of gastrocnemius. Hypercaloric saturated, monounsaturated, or n-6 polyunsaturated lipid diets cause equivalent whole body insulin resistance in rats, but only an n-6 polyunsaturated acid-based diet triggers intramuscular TAG accumulation.

Adipose Tissue↗

PubMed2003

Immortalization of human preadipocytes.

Primary human preadipocytes in culture are characterized by a low proliferative capacity associated with a rapid decline of differentiation ability during subculturing; thereby limiting their use as cellular model. Cellular immortalization constitutes an interesting approach for establishing cell lines presenting an unlimited life span and a maintained differentiation capacity. Different procedures for developing immortalized human preadipocytes are discussed in this review. Transformation of human preadipocytes with the simian virus 40 large T-antigen (SV40 T-Ag) permitted the development of immortalized cells; however these cells could not maintain their capacity to differentiate into adipocytes. This limitation may be explained by the ability of SV40 T-Ag to inhibit transcriptional factors involved in the differentiation of preadipocyte. Reconstitution of the telomerase activity by stable expression of the hTERT (human telomerase catalytic subunit) gene was able to partially extend the lifespan of primary preadipocytes but not to promote cellular immortalization. However, a combined expression of hTERT and the E7 oncoprotein of human papillomavirus type 16, generated human preadipocytes with both an unlimited life span and a preserved adipogenic potential. This approach appears to be an effective method for establishing human preadipose cell lines for studying adipocyte differentiation and metabolism.

Adipocytes↗

PubMed2003

3-methylindole-induced toxicity to human bronchial epithelial cell lines.

Transfected BEAS-2B cells that express different cytochrome P450 enzymes were used to assess whether human bronchial epithelial cell lines are target cells for 3-methylindole (3MI)-induced damage. Four different transfected BEAS-2B lines overexpressing P450s 2A6, 3A4, 2F1, and 2E1 (B-CMV2A6, B-CMV3A4, B-CMV2F1, and B-CMV2E1), respectively, were compared. The B-CMV2F1 and B-CMV3A4 cells were the most susceptible to 3MI-mediated cytotoxicity, measured by leakage of lactate dehydrogenase into the medium after a 48-h incubation. The toxicity was ameliorated by pretreatment with 1-aminobenzotriazole (ABT). Depletion of glutathione with diethylmaleate decreased the onset and increased the extent of cell death with 3MI. Thus, 3MI is cytotoxic to immortalized bronchial epithelial cells overexpressing 2F1 without concomitant depletion of GSH, but depletion of GSH modestly enhances the cytotoxicity of 3MI to human lung cells. Additional studies clearly demonstrated that a low concentration of 3MI (10 micro M) induced apoptosis in BEAS-2B cells that was measured by DNA fragmentation, and apoptosis was inhibited by the presence of ABT. The B-CMV2F1 cells overexpressing 2F1 demonstrated increased apoptosis (measured by Annexin-V binding) at 24 h with 100 micro M 3MI. Therefore, CYP2F1 in human bronchial epithelial lung cells may bioactivate 3MI to 3-methyleneindolenine, which induces programmed cell death at relatively low concentrations. Human lung cells may be susceptible to this prototypical pneumotoxicant.

Apoptosis↗

PubMed2002

Comparative aflatoxin B(1) activation and cytotoxicity in human bronchial cells expressing cytochromes P450 1A2 and 3A4.

Some epidemiological evidence suggests a link between the inhalation of aflatoxin B(1) (AFB(1))-contaminated grain dusts and increased lung cancer risk. However, the mechanisms of AFB(1) activation and action in human lung are not well understood. We compared AFB(1) action in SV40 immortalized human bronchial epithelial cells (BEAS-2B) with two transfected cell lines that stably express human cytochromes P450 (CYPs) 1A2 (B-CMV1A2) and 3A4 (B3A4), the principal CYPs thought to activate this mycotoxin in human liver. All three cell types retained catalytically active glutathione S-transferase, the key phase II enzyme that detoxifies metabolically activated AFB(1). B-CMV1A2 and B3A4 cells expressed methoxyresorufin-O-demethylase (MROD) and nifedipine oxidase activities, respectively, and were 3000- and 70-fold more susceptible, respectively, to the cytotoxic effects of AFB(1) than the control cell line (BEAS-2B). When cultured with a range of low, environmentally relevant AFB(1) concentrations (0.02-1.5 microM), control cells formed barely detectable AFB(1)-DNA adducts, whereas B-CMV1A2 cells formed significantly more adducts than B3A4 cells. In B-CMV1A2 cells, formation of AFB(1)-DNA adducts was inhibited by the CYP 1A2 inhibitor 7,8-benzoflavone, whereas formation of AFB(1)-DNA adducts in B3A4 cells was inhibited by the CYP 3A4 inhibitor 17alpha-ethynylestradiol. Competitive reverse transcription-PCR analysis showed that only the CYP-transfected cell lines expressed CYP mRNA. When adjusted for CYP mRNA expression, B-CMV1A2 cells were more efficient in the formation of cytotoxic and DNA-alkylating species at low AFB(1) concentrations, whereas B3A4 cells were more efficient at high concentrations. Our results affirm the hypothesis that, as in human liver microsomes, CYP 1A2 in human lung cells appears to have a more important role than CYP 3A4 in the bioactivation of low AFB(1) concentrations associated with many human exposures. Therefore, it is possible that under conditions in which appropriate CYPs are expressed in lung, inhalation of AFB(1) may result in increased risk of lung cancer in exposed persons.

Aflatoxin B1↗

PubMed2002

Cyclosporin A-induced free radical generation is not mediated by cytochrome P-450.

1. Reactive oxygen species (ROS) have been proposed to play a role in the side effects of the immunosuppressive drug cyclosporin A (CsA). 2. The aim of this study was to investigate whether cytochrome P-450 (CYP) dependent metabolism of CsA could be responsible for ROS generation since it has been suggested that CsA may influence the CYP system to produce ROS. 3. We show that CsA (1 -- 10 microM) generated antioxidant-inhibitable ROS in rat aortic smooth muscle cells (RASMC) using the fluorescent probe 2,7-dichlorofluorescin diacetate. 4. Using cytochrome c as substrate, we show that CsA (10 microM) did not inhibit NADPH cytochrome P-450 reductase in microsomes prepared from rat liver, kidney or RASMC. 5. CsA (10 microM) did not uncouple the electron flow from NADPH via NADPH cytochrome P-450 reductase to the CYP enzymes because CsA did not inhibit the metabolism of substrates selective for several CYP enzymes that do not metabolize CsA in rat liver microsomes. 6. CsA (10 microM) did not generate more radicals in CYP 3A4 expressing immortalized human liver epithelial cells (T5-3A4 cells) than in control cells that do not express CYP 3A4. 7. Neither diphenylene iodonium nor the CYP 3A inhibitor ketoconazole were able to block ROS formation in rat aortic smooth muscle or T5-3A4 cells. 8. These results demonstrate that CYP enzymes do not contribute to CsA-induced ROS formation and that CsA neither inhibits NADPH cytochrome P-450 reductase nor the electron transfer to the CYP enzymes.

Animals↗