In vitro excess ammonia production in human myeloma cell lines

Abstract

It is well known that cases with multiple myeloma reveal various clinical manifestations such as pancytopenia, hyperproteinemia, renal dysfunction, bone lesions, hypercalcemia and immunodeficiency. Recently, a few more clinical features associated with myeloma, such as salivary type hyperamylasemia and elevated serum C-reactive protein (CRP) concentration, have been reported. The elevation of CRP is thought to be related to interleukin-6 (IL-6) production by myeloma cells, because of identification of IL-6 as an autocrine and/or paracrine growth factor for myeloma cells. More recently, there have been several reports of cases with myeloma associated with hyperammonemia. This hyperammonemia is not considered to be due to liver dysfunction, because in most of these cases tests revealed normal hepatic function, and some cases showed different patterns of serum amino acid distribution than that associated with hepatic failure. However, there have been no apparent observations of ammonia production by myeloma cells. In this study, we used six human myeloma cell lines including KMS-18, which was recently established from a myeloma case associated with hyperammonemia. These lines were treated with MRA (mycoplasma removal agent) to observe ammonia production in vitro. They produced and released significantly higher levels of ammonia into culture medium than non-myeloma hematological cell lines or the HepG2 human hepatic carcinoma cell line. Although attempts to analyze the relative expression levels of the enzymes related to ammonia biosynthesis using the reverse transcriptase-polymerase chain reaction assay failed to detect any differences between these myeloma lines and other cell lines, in vitro excess ammonia production by the myeloma cells was confirmed and the relevance to clinical manifestations is discussed.

Lactate dehydrogenase production and release in a newly established human myeloma cell line

“…For example, production of the bone-resorbing cytokines, such as IL-1␤ and lymphotoxin, by myeloma cells is related to osteoclastic bone destruction and hypercalcemia [41][42][43]. Disturbance of consciousness may be caused by hyperammonemia in patients with ammoniaproducing myeloma [44,45]. Also, amylase production and release by myeloma cells have been reported [46,47].…”

Dependence on glutamine uptake and glutamine addiction characterize myeloma cells: a new attractive target

“…1,2 It is a historical notion that the growth of MM cells was limited by depletion of L-glutamine (Gln) 3 and that MM cells may produce an excess of ammonium (NH 4 1 ). 4,5 Hyperammonemia with or without the related encephalopathy has been reported as a possible rare clinical manifestation in relapsed/refractory MM patients with high mortality rate.…”

“…1,2 It is a historical notion that the growth of MM cells was limited by depletion of L-glutamine (Gln) 3 and that MM cells may produce an excess of ammonium (NH 4 1 ). 4,5 Hyperammonemia with or without the related encephalopathy has been reported as a possible rare clinical manifestation in relapsed/refractory MM patients with high mortality rate. [6][7][8][9][10][11][12] Recently, multivariate analysis based on 1 H-NMR spectroscopy analysis of serum samples has shown that a specific metabolic profile characterized MM patients vs healthy controls, including Gln levels significantly lower in the MM group.…”

“…13 Overall, these data suggest that Gln is highly metabolized in MM cells. To satisfy metabolic requirements of Gln, mammalian cells rely on glutamine synthetase (GS), the enzyme that obtains Gln from glutamate (Glu) and NH 4 1 . 14, 15 Moreover, a variety of carriers, operate Gln influx, such as the Na 1 -dependent transporters SNAT1-2 and ASCT2, and the Na 1 -independent transporter LAT1.…”

“…16 Gln is a substrate of several enzymes, playing an important role in various processes, such as the synthesis of nucleotides, other amino acids, or glucosamine. 17,18 Moreover, through the activity of glutaminases (GLS1 and GLS2), which hydrolyze the amide group obtaining NH 4 1 and Glu, Gln may fuel the intracellular pool of the Krebs cycle intermediate and anaplerotic substrate 2-oxoglutarate (2-OG, a-ketoglutarate). 17,18 Some types of human tumor cells exhibit an high requirement for Gln ("glutamine addiction") 15 and…”