A New Enzyme Immunoassay for the Quantitative Determination of Classical Autotaxins (ATXα, ATXβ, and ATXγ) and Novel Autotaxins (ATXδ and ATXε)

Abstract

BackgroundAutotaxin (ATX) is a secreted enzyme that converts lysophosphatidylcholine to lysophosphatidic acid, a potent bioactive lipid mediator, through its lysophospholipase D activity. Although five alternative splicing isoforms of ATX have been identified as ATXα, ATXβ, ATXγ, ATXδ, and ATXε and the expression patterns of each isoform differ among several tissues, the clinical significance of each isoform remains to be elucidated.MethodsAnti-ATXβ and anti-ATXδ monoclonal antibodies were produced by immunization with recombinant human ATXβ and ATXδ expressed using a baculovirus system, respectively. We then developed enzyme immunoassays to measure the serum concentrations of “classical ATX” (ATXα, ATXβ, and ATXγ) and “novel ATX” (ATXδ and ATXε) antigens and evaluated the usefulness of these assays using human serum samples.ResultsThe with-run and between-run precision, interference, detection limit, and linearity studies for the present assay were well validated. In healthy subjects, the serum concentrations of classical ATX and novel ATX were significantly (P < 0.01) higher in women than in men, while the ratios of classical ATX or novel ATX to total ATX were not different between women and men. The concentrations of both classical ATX and novel ATX in normal pregnant subjects and patients with chronic liver diseases or follicular lymphoma were significantly higher than those in healthy subjects, while the ratio of both ATX isoforms to total ATX did not vary among these groups.ConclusionsWe have developed a new enzyme immunoassay to determine the concentrations of classical ATX and novel ATX in human serum. These assays may be helpful for elucidating the distinct functional roles of each ATX isoform, which are largely unknown at present.

Association between serum autotaxin or phosphatidylserine‐specific phospholipase A1 levels and melanoma

“…First, we compared the serum total ATX and ATX isoforms between healthy subjects and the subjects with melanoma in male or female subjects separately, because the total ATX and ATX isoform levels are higher in female subjects than in male subjects . Although we observed no difference between healthy and melanoma subjects when we analyzed the results in all of the subjects (Fig.…”

“…Classical ATX and novel ATX are widely expressed in human tissues, with relatively high expression levels of classical ATX seen in the brain, placenta, ovary and small intestine, and high levels of novel ATX seen in the small intestine and spleen . Although the predominant isoforms in melanoma cells remain to be elucidated, the results from the present study suggest that a deviation in ATX isoforms may not exist in melanoma, as previously described for patients with follicular lymphoma or liver fibrosis as well as pregnant patients …”

“…The ATX and PS‐PLA 1 antigen levels in the ascites were determined using a two‐site immunoenzymetric assay with the TOSOH AIA system (TOSOH, Tokyo, Japan) . Regarding ATX, because five alternative splicing isoforms of ATX have been identified as ATXα, ATXβ, ATXγ, ATXδ and ATXε, we also measured the classical ATX (ATXα, ATXβ and ATXγ) and novel ATX (ATXδ and ATXε) levels using enzyme immunoassays that we recently developed …”

Analysis of glycero-lysophospholipids in gastric cancerous ascites

“…Regarding ATX, because five alternative splicing isoforms of ATX have been identified as ATXα, ATXβ, ATXγ, ATXδ, and ATXε, we also measured the classical ATX (ATXα, ATXβ, and ATXγ) and novel ATX (ATXδ and ATXε) levels using enzyme immunoassays that we recently developed (25). …”

The critical role and potential target of the autotaxin/lysophosphatidate axis in pancreatic cancer

“…10 To date, five different alternatively spliced isoforms of ATX have been identified, including α, β, γ, δ, and ε, and all of them are catalytically active. 11 Full-length ATX is synthesized as a pre-proenzyme and is secreted by the classical secretory pathway. 12 Although no obvious differences in the core mechanism of involvement in catalysis or physiological activities between different ATX variants have been reported, the differences in gene organization, tissue distribution, and biochemical characterization have been documented.…”