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Donald M Mock – 1st expert on this subject based on the ideXlab platform
in hepg2 cells coexisting carnitine Deficiency masks important indicators of marginal Biotin DeficiencyJournal of Nutrition, 2015Co-Authors: Anna Bogusiewicz, Gunnar Boysen, Donald M MockAbstract:
Background: A large number of birth defects are related to nutrient deficiencies; concern that Biotin Deficiency is teratogenic in humans is reasonable. Surprisingly, studies indicate that increased urinary 3-hydroxyisovalerylcarnitine (3HIAc), a previously validated marker of Biotin Deficiency, is not a valid biomarker in pregnancy.
Objective: In this study we hypothesized that coexisting carnitine Deficiency can prevent the increase in 3HIAc due to Biotin Deficiency.
Methods: We used a 2-factor nutrient depletion design to induce isolated and combined Biotin and carnitine Deficiency in HepG2 cells and then repleted cells with carnitine. To elucidate the metabolic pathogenesis, we quantitated intracellular and extracellular free carnitine, acylcarnitines, and acylcarnitine ratios using liquid chromatography–tandem mass spectrometry.
Results: Relative to Biotin-sufficient, carnitine-sufficient cells, intracellular acetylcarnitine increased by 90%, propionylcarnitine more than doubled, and 3HIAc increased by >10-fold in Biotin-deficient, carnitine-sufficient (BDCS) cells, consistent with a defensive mechanism in which Biotin-deficient cells transesterify the acyl-coenzyme A (acyl-CoA) substrates of the Biotin-dependent carboxylases to the related acylcarnitines. Likewise, in BDCS cells, the ratio of acetylcarnitine to malonylcarnitine and the ratio of propionylcarnitine to methylmalonylcarnitine both more than tripled, and the ratio of 3HIAc to 3-methylglutarylcarnitine (MGc) increased by >10-fold. In Biotin-deficient, carnitine-deficient (BDCD) cells, the 3 substrate-derived acylcarnitines changed little, but the substrate:product ratios were masked to a lesser extent. Moreover, carnitine repletion unmasked Biotin Deficiency in BDCD cells as shown by increases in acetylcarnitine, propionylcarnitine, and 3HIAc (each increased by >50-fold). Likewise, ratios of acetylcarnitine:malonylcarnitine, propionylcarnitine:methylmalonylcarnitine, and 3HIAc:MGc all increased by >8-fold.
Conclusions: Our findings provide strong evidence that coexisting carnitine Deficiency masks some indicators of Biotin Deficiency and support the potential importance of the ratios of acylcarnitines arising from the acyl-CoA substrates and products for Biotin-dependent carboxylases in detecting the Biotin Deficiency that is masked by coexisting carnitine Deficiency.
marginal Biotin Deficiency can be induced experimentally in humans using a cost effective outpatient designJournal of Nutrition, 2012Co-Authors: Shawna L Stratton, Anna Bogusiewicz, Gunnar Boysen, Cindy L Henrich, Thomas D Horvath, Nell I Matthews, Jeffery H Moran, Amanda M Dawson, Suzanne N Owen, Donald M MockAbstract:
To date, marginal, asymptomatic Biotin Deficiency has been successfully induced experimentally by the use of labor-intensive inpatient designs requiring rigorous dietary control. We sought to determine if marginal Biotin Deficiency could be induced in humans in a less expensive outpatient design incorporating a self-selected, mixed general diet. We sought to examine the efficacy of three outpatient study designs: two based on oral avidin dosing and one based on a diet high in undenatured egg white for a period of 28 d. In study design 1, participants (n = 4; 3 women) received avidin in capsules with a Biotin binding capacity of 7 times the estimated dietary Biotin intake of a typical self-selected diet. In study design 2, participants (n = 2; 2 women) received double the amount of avidin capsules (14 times the estimated dietary Biotin intake). In study design 3, participants (n = 5; 3 women) consumed egg-white beverages containing avidin with a Biotin binding capacity of 7 times the estimated dietary Biotin intake. Established indices of Biotin status [lymphocyte propionyl-CoA carboxylase activity; urinary excretion of 3-hydroxyisovaleric acid, 3-hydroxyisovaleryl carnitine (3HIA-carnitine), and Biotin; and plasma concentration of 3HIA-carnitine] indicated that study designs 1 and 2 were not effective in inducing marginal Biotin Deficiency, but study design 3 was as effective as previous inpatient study designs that induced Deficiency by egg-white beverage. Marginal Biotin Deficiency can be induced experimentally by using a cost-effective outpatient design by avidin delivery in egg-white beverages. This design should be useful to the broader nutritional research community.
urinary excretion of 3 hydroxyisovaleryl carnitine is an early and sensitive indicator of marginal Biotin Deficiency in humansJournal of Nutrition, 2011Co-Authors: Shawna L Stratton, Anna Bogusiewicz, Cindy L Henrich, Thomas D Horvath, Nell I Matthews, Horace J Spencer, Jeffery H Moran, Donald M MockAbstract:
Mounting evidence indicates that marginal Biotin Deficiency is not rare, contrary to previous assumptions. Accordingly, robust indicators of Biotin status would be useful. In a study of 10 healthy adults, we recently provided evidence that abnormally increased plasma concentration of 3-hydroxyisovaleryl carnitine (3HIA-carnitine) is a sensitive indicator of marginal Biotin Deficiency. We sought to determine whether urinary excretion of 3HIA-carnitine (expressed as the ratio to urinary creatinine) significantly increases in marginal Biotin Deficiency. Marginal, asymptomatic Biotin Deficiency was induced experimentally in the same 10 healthy adults (8 women) by feeding undenatured egg white with meals for 28 d. Biotin status was repleted by a mixed general diet plus Biotin supplementation. Urinary excretion of 3HIA-carnitine was determined by liquid chromatography-tandem MS on d 0, 14, and 28 (depletion) and on d 35 and 50 (repletion). Mean urinary 3HIA-carnitine concentration increased with depletion (P < 0.0001; d 0 vs. 28) and decreased with repletion (P = 0.0002; d 28 vs. 50). Urinary 3HIA-carnitine excretion was greater than the upper limit of normal in 9 of 10 participants by d 14 and decreased to within normal limits by d 50 in all participants. This study provides evidence that urinary excretion of 3HIA-carnitine is an early and sensitive indicator of marginal Biotin Deficiency. The ease of collection of untimed urine samples and application of a new analytical method with simplified sample preparation suggest that urinary 3HIA-carnitine is likely to be a useful indicator for large population studies.
Nell I Mock – 2nd expert on this subject based on the ideXlab platform
Biotin Deficiency reduces expression of SLC19A3, a potential Biotin transporter, in leukocytes from human bloodExperimental biology, 2020Co-Authors: Tatyana I Vlasova, Nell I Mock, Shawna L Stratton, Amanda M Wells, Donald M. MocksAbstract:
In evaluating potential indicators of Biotin status, we quantitated the expression of Biotin-related genes in leukocytes from human blood of normal subjects before and after inducing marginal Biotin Deficiency. Biotin Deficiency was induced experimentally by feeding an egg-white diet for 28 d. Gene expression was quantitated for the following Biotin-related proteins: methylcrotonyl-CoA carboxylase chains A (MCCA) and B (MCCB); propionyl-CoA carboxylase chains A (PCCA) and B (PCCB); pyruvate carboxylase (PC); acetyl-CoA carboxylase isoforms A (ACCA) and B (ACCB); holocarboxylase synthetase (HCS); Biotinidase; and 2 potential Biotin transporters: sodium-dependent multivitamin transporter (SMVT) and solute carrier family 19 member 3 (SLC19A3). For 7 subjects who successfully completed the study, the abundance of the specific mRNAs was determined by quantitative real-time RT-PCR at d 0 and 28. At d 28, SLC1 9A3 expression had decreased to 33% of d 0 (P < 0.02 by two-tailed, paired t test). Expression of MCCA, PCCA, PC, ACCA, ACCB, HCS, Biotinidase, and SMVT decreased to ∼80% of d 0 (P < 0.05). Expression of the MCCB and PCCB chains that do not carry the Biotin-binding motif did not change significantly; we speculate that expression of the Biotin-binding chains of Biotin-dependent carboxylases is more responsive to Biotin status changes. These data provide evidence that expression of SLC19A3 is a relatively sensitive indicator of marginal Biotin Deficiency.
lymphocyte propionyl coa carboxylase and its activation by Biotin are sensitive indicators of marginal Biotin Deficiency in humansThe American Journal of Clinical Nutrition, 2006Co-Authors: Shawna L Stratton, Nell I Mock, Anna Bogusiewicz, Amanda M Wells, Matthew M Mock, Donald M MockAbstract:
BACKGROUND: Marginal Biotin Deficiency may be a human teratogen. A Biotin status indicator that is not dependent on renal function may be useful in studies of Biotin status during pregnancy. A previous study of experimental Biotin Deficiency suggested that propionyl-coenzyme A carboxylase (PCC) activity in peripheral blood lymphocytes (PBLs) is a sensitive indicator of Biotin status. OBJECTIVE: We examined the utility of measuring PCC activity and the activation of PCC by Biotin in detecting marginal Biotin Deficiency. DESIGN: Marginal Biotin Deficiency was induced in 7 adults (3 women) by egg-white feeding for 28 d. Blood and urine were obtained on days 0, 14, and 28 (depletion phase) and 44 and 65 (repletion phase). PBLs were incubated with (activated) or without (control) Biotin before PCC assay. The activation coefficient of PCC is the ratio of PCC activity in activated PBLs to that in control PBLs. The significance of differences for all measurements was tested by repeated-measures analysis of variance with Fisher’s post hoc test and Bonferroni correction. RESULTS: Changes in the urinary excretion of Biotin and of 3-hydroxyisovaleric acid confirmed that marginal Biotin Deficiency was successfully induced. By day 14, PCC activity had decreased (P < 0.0001) to below the lower limit of normal in all subjects. By day 28, the activation coefficient of PCC had increased significantly (P = 0.003) and was above the upper limit of normal in 6 of 7 subjects. CONCLUSION: PCC activity is the most sensitive indicator of Biotin status tested to date. In future pregnancy studies, the use of lymphocyte PCC activity data should prove valuable in the assessment of Biotin status.
Biotin Deficiency reduces expression of slc19a3 a potential Biotin transporter in leukocytes from human bloodJournal of Nutrition, 2005Co-Authors: Tatyana I Vlasova, Nell I Mock, Shawna L Stratton, Amanda M Wells, Donald M MockAbstract:
Biotin Deficiency is teratogenic in mice (1,2) and may be teratogenic in humans (3). Valid indicators of marginal and moderate Biotin Deficiency would be useful in investigating the role of Biotin Deficiency in birth defects and in other illnesses hypothesized to be Biotin related (4 –7). Emerging evidence indicates that Biotin plays a role in gene expression (8 –12). In addition to acting as a cofactor for Biotin-dependent carboxylases, Biotin stimulates expression of hepatic glucokinase (8) and represses expression of hepatic phosphoenolpyruvate carboxylase (9) in vivo, and expression of the Biotin-related enzymes propionyl-CoA carboxylase chain A (PCCA),4 acetyl-CoA carboxylase isoform A (ACCA), and holocarboxylase synthetase (HCS) in cultured human hepatoblastoma cells and normal fibroblasts (10). However, no such studies have been performed in humans in vivo. In this study, we examined the expression of specific Biotin-related genes as indicators of marginal, asymptomatic Biotin Deficiency and assessed gene response to marginal Biotin Deficiency.
In mammals, Biotin is a coenzyme for 5 Biotin-dependent carboxylases: methylcrotonyl-CoA carboxylase (MCC), propionyl-CoA carboxylase (PCC), pyruvate carboxylase (PC), and the 2 isoforms of ACC (ACCA and ACCB). The active forms of the enzymes (holocarboxylases) contain Biotin covalently bound to lysine residues; the attachment of Biotin to the corresponding apocarboxylase is catalyzed by HCS.
Biotin is transported into eukaryotic cells by Biotin transporters located in cell membranes. Three Biotin transporters have been proposed in human cells: 1) the sodium-dependent multivitamin transporter (SMVT) (13,14); 2) the solute carrier family 19 member 3 (SLC19A3) (15,16); and 3) the monocarboxylate transporter R1 (17). This third transporter was proposed after this study was initiated and was not examined here.
Biotinidase catalyzes the release of covalently bound Biotin from Biotinyl-peptides generated by the turnover of intracellular proteins and releases Biotin from dietary proteins during digestion (18). Biotinidase is also likely important in catalyzing the covalent binding of Biotin to histones (19).
In this study, we evaluated expression of Biotin-related genes as potential indicators of marginal, asymptomatic Biotin Deficiency. Gene expression was quantitated in leukocytes of 7 healthy humans after 28 d of progressive Biotin Deficiency.
Antonio Velazquezarellano – 3rd expert on this subject based on the ideXlab platform
functional and metabolic implications of Biotin Deficiency for the rat heartMolecular Genetics and Metabolism, 2008Co-Authors: Antonio Velazquezarellano, Maria De La Luz Hernandezesquivel, Rafael Moreno Sanchez, Daniel Ortegacuellar, Nayeli Rodriguezfuentes, Saul Cano, Alfonso Leondelrio, Karla CarvajalAbstract:
The tricarboxylic acid (TCA) cycle is the main ATP provider for the heart. TCA carbons must be replenished by anaplerosis for normal cardiac function. Biotin is cofactor of the anaplerotic enzymes pyruvate and propionyl-CoA carboxylases. Here, we found that in Biotin deficient rats, both carboxylases decreased 90% in adipose tissue, jejunum and spleen, but in heart they conserved about 60% residual activity. We then investigated if under Biotin Deficiency (BtDEF), the heart is able to maintain its function in vivo and in isolated conditions, and during ischemia and reperfusion, where metabolism drastically shifts from oxidative to mainly glycolytic. Neither glucose nor octanoate oxidation were severely affected in BtDEF hearts, as assessed by mechanical performance, oxygen uptake or high-energy metabolite content; however, myocardial hexokinase activity and lactate concentration were reduced in deficient hearts. When challenged by ischemia and reperfusion injury, BtDEF hearts did not suffer more damage than the controls, although they lowered significantly their performance, when changed to ischemic conditions, which may have clinical implications. Post-ischemic increase in ADP/ATP ratio was similar in both groups, but during reperfusion there was higher rhythm perturbation in BtDEF hearts. By being relatively insensitive to Biotin Deficiency, cardiac tissue seems to be able to replenish TCA cycle intermediates and to maintain ATP synthesis.
Biotin Deficiency affects both synthesis and degradation of pyruvate carboxylase in rat primary hepatocyte culturesMolecular Genetics and Metabolism, 2007Co-Authors: Nayeli Rodriguezfuentes, Itzel Lopezrosas, Gabriela Romancisneros, Antonio VelazquezarellanoAbstract:
Abstract Pyruvate carboxylase (PC) is a Biotin-dependent enzyme that plays a crucial role in gluconeogenesis, lipogenesis, Krebs cycle anaplerosis and amino acid catabolism. Biotin Deficiency reduces its mass besides its activity. Enzyme mass is the result of its cellular turnover, i.e., its rates of synthesis and degradation. We have now investigated, by a pulse and chase approach in cultured primary hepatocytes, the effects of Biotin Deficiency on these rates. Wistar rats were fed a Biotin-deficient diet and the controls were fed the same diet supplemented with Biotin; their Biotin status was monitored measuring lymphocytes propionyl-CoA carboxylase activity and urinary 3-hydroxyisovaleric acid. After 6–7 weeks primary hepatocytes were cultured in Biotin-deficient or complete DMEM. PC activity was determined by measuring the incorporation of 14 C-bicarbonate into acid-non-volatile products, and its mass by streptavidin Western blots. Its synthesis rate was estimated from [ 35 S] methionine incorporation into anti-PC antibody immunoprecipitate. Its degradation rate was calculated from the loss of radioactivity from previously labeled hepatocytes, in a medium containing an excess of non-radioactive methionine. PC synthesis rate in Biotin-deficient hepatocytes was approximately 4.5-fold lower than in the controls, and its degradation rate was 5.1-fold higher. Therefore, the decrement of PC mass during Biotin Deficiency results both from a decrease in its synthesis and an increase in its degradation rates. To our knowledge, this is the first instance where a mammalian enzyme cofactor is necessary to sustain both processes.