Chemerin isoform analysis in human biofluids using an LC/MRM-MS-based targeted proteomics approach with stable isotope-labeled standard.

Targeted proteomics has advantages over earlier conventional technologies for protein detection. We developed and validated an LC/MRM-MS-based targeted proteomic method combined with immunoaffinity precipitation for the enrichment and detection of low abundance chemerin isoforms in human biofluids. After tryptic digestion, each chemerin isoform was characterized by isoform-specific peptides, and the absolute quantification was achieved by using stable isotope-labeled peptides as internal standards. In serum, follicular fluid and synovial fluid, a total of 6 chemerin isoforms were identified and quantified, among which a novel natural isoform 153Q was discovered for the first time. The relative content of the six chemerin isoforms in human serum was 157S ≫ 156F ≫ 158K > 154F ≥ 155A > 153Q in the ratio of 25:17:5:2.5:2.2:1, respectively. The absolute contents were in the range of 88-3.5 ng/mL. This distribution remained consistent among the 3 biofluids analyzed. Total chemerin were found to be increased in both polycystic ovary syndrome (serum and follicular fluid) and rheumatoid arthritis (serum) patients. However, chemerin isoform analysis revealed that only 156F & 157S were increased in the former, while 155A, 156F & 157S were increased in the latter. This demonstrates the potential of this method in detailed characterization of changes in chemerin isoforms that may be of clinical relevance.

Alterations of Sphingolipid Metabolism in Different Types of Polycystic Ovary Syndrome.

The roles of sphingolipids in polycystic ovary syndrome (PCOS) are still unknown. This study aimed to investigate the sphingolipid characteristics for different types of PCOS using liquid chromatography-mass spectrometry (LC-MS). A total of 107 women with PCOS and 37 healthy women as normal controls were studied. PCOS patients were further classified into non-obesity with insulin resistance (IR) (NOIR), obesity with IR (OIR), and non-obesity and non-IR (NIR) subgroups. A total of 87 serum sphingolipids, including 9 sphingosines, 3 sphinganines, 1 sphingosine-1-phosphate (S1P), 19 ceramides (Cers), 1 ceramide-1-phosphate, 44 sphingomyelins (SMs), 4 hexosylceramides, and 6 lactosylceramides (LacCers) were analyzed using an improved sphingolipidomic approach based on LC-MS. Notable elevations in the levels of S1P, Cer, and SM were observed in PCOS patients when compared with healthy women, and SM species with long saturated acyl chains showed potential as novel biomarkers of PCOS. In addition, the level of LacCer was only elevated in NIR, and there was almost no change in NOIR and OIR. This study is the first to report the comprehensive sphingolipidomic profiling of different subgroups of PCOS with or without IR or obesity and suggests that serum sphingolipids might be useful as diagnostic biomarkers for different types of PCOS.