SFEBES2018 Poster Presentations Obesity & metabolism (24 abstracts)
Background: The liver stores 10 μg cobalamin per gram protein and able to hold 50% (11.5 mg) of bodys B12. Making up the bulk of the liver, hepatocytes express receptors regulating cellular uptake of B12 in the liver. However, relationship between circulating and intracellular B12 levels as well as regulation of hepatic uptake of B12 is unexplored.
Objective: To assess the regulation of cellular uptake and storage of B12 in hepatocytes in varying concentrations of circulating B12.
Methods: HepG2 cell line was cultured using B12 deficient EMEM medium and seeded in 500 nM, 400 nM, 200 nM, 100 nM, 50 nM, 20 nM, 10 nM, 1 nM, 100 pM and 25 pM B12-media. B12 concentration within hepatocytes and corresponding conditioned media was determined by electrochemiluminescent immunoassay using a Roche Cobas immunoassay analyzer (Roche Diagnostics UK). Gene and protein expression of transcobalamin II (TCN2) and transcobalamin receptor (TCblR)(CD320) were assessed by RT-PCR and western blots.
Results: Low B12 (25 pM and 100 pM) in condition media resulted in 210280% increase in intracellular levels of B12 in hepatocytes. 1000 pM (1 nM) circulatory B12 resulted in 42.7% storage in hepatocytes. Higher circulating B12 such as 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 400 nM and 500 nM decreased uptake of B12 to 4.7%, 4.3%, 3.1%, 1.6%, 2.3%, 1.3% and 1.8% respectively. We observed increased gene and protein expression of transcobalamin II (TCN2) and receptors (TCblR) (CD320) in hepatocytes under low B12 than higher concentrations.
Conclusion: Our study highlights low circulatory B12 (0100pM) triggers higher intracellular levels of 23 fold and vice versa, supported by increased gene and protein expression of receptors/transporters. Implies active transport of B12 predominate at lower concentrations, possibly due to high expression of receptors/transporters. Hence suggesting the optimal physiological B12 concentration is required rather than overloading with supplements. This provides us impetus to further study whether the tissue-specific effect of low B12 dysregulates hepatic metabolism.
19 Nov 2018 - 21 Nov 2018