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Pre-analytical-related variability influencing serum peptide profiles demonstrated in a mass spectrometry-based search for colorectal and prostate cancer biomarkers

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Karczmarski J., Rubel T., Mikula M., Wolski J., Rutkowski A., Zagorowicz E., Dadlez M., Ostrowski J.

2013

vol. 60

issue 3

417-425

Although the degradome, which comprises proteolytic fragments of blood proteins, presents a potential source of diagnostic biomarkers, studies on cancer peptide biomarkers have provided inconsistent conclusions. In the present study, we reevaluated the usefulness of serum degradome analyses for searching peptide cancer biomarker candidates. Particular attention was paid to pre-analytical factors influencing the variability of determined peptide levels, including clotting time and control group selection. Studies were conducted on 44 and 86 serum samples collected from cancer patients and healthy individuals, respectively, using liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS)-based analyses. We identified 1373 unique peptides, nearly 40% of which originated from five blood proteins: fibrinogen alpha chain, apolipoprotein A-IV (APOA4), complement C3, apolipoprotein A-I, and alpha-1-antitrypsin. A set of 118 and 88 peptides exhibited highly significant differences (adjusted p-value ≤ 0.01 and fold change ≥ 2) in pair-wise comparisons of control vs. prostate cancer and control vs. colorectal cancer, respectively, with 37 peptides displaying a consistent direction of change for these pair-wise comparisons. The levels of 67 peptides differed significantly in serum samples collected from healthy individuals immediately prior to colonoscopy and those who underwent colonoscopic examination at least four weeks earlier. Of them, 49 peptides originated from APOA4. Whereas earlier studies, including ours, have utilized fragments of fibrinopeptide A (FPA) to distinguish cancer from healthy cases, here we show that their absolute abundance is a sensitive indicator of clotting time. These observations may have implications for future serum peptidome studies since these issues have not previously been recognized.

Gene expression alterations induced by low molecular weight heparin during bowel anastomosis healing in rats

81%

Krześniak N., Paziewska A., Rubel T., Skrzypczak M., Mikula M., Dzwonek A., Goryca K., Wyrwicz L. S., Jarosz D., Laubitz D., Woszczyński M., Bielecki K., Ostrowski J.

2011

vol. 58

issue 1

79-87

Colon anastomosis is therapeutically challenging because multiple, usually undetectable factors influence a spectrum of repair mechanisms. We hypothesized that low molecular weight heparins, routinely administered perioperatively, may differentially affect gene expression related to colon healing. Twenty pairs of untreated and enoxaparin-treated rats underwent left-side hemicolectomy with a primary end-to-end anastomosis. Normal colon and anastomotic bowel segments were resected on day 0 and on days 1, 3, 5, and 7 after surgery, respectively. Serial anastomosis transverse cross-sections were evaluated microscopically and by microarray (Rat Genome 230 2.0, Affymetrix). Differentially expressed probe sets were annotated with Gene Ontology. We also examined the influence of enoxaparin on fibroblast proliferation and viability in vitro. Among the 5476 probe sets, we identified differential expression at each healing time point, yielding 79 subcategories. Most indicated genes were involved in wound healing, including multicellular organismal development, locomotory behavior, immune response, cell adhesion, inflammatory response, cell-cell signaling, blood vessel development, and tissue remodeling. Although we found no intensity differences in histological features of healing between enoxaparin-treated and control rats, treatment did induce significant expression changes during early healing. Of these changes, 83 probe sets exhibited at least twofold changes and represented different functional annotations, including inflammatory response, regulation of transcription, regulation of apoptosis, and angiogenesis. Fibroblast culture confirmed an anti-viability effect of enoxaparin. Enoxaparin affects colon wound-related gene expression profiles, but further studies will resolve whether heparin treatment is a risk factor after intestinal surgery, at least in some patients.

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2 Acta Biochimica Polonica

1 2013

1 2011

2 Mikula M.

2 Ostrowski J.

2 Rubel T.

1 Bielecki K.

1 Dadlez M.

1 Dzwonek A.

1 Goryca K.

1 Jarosz D.

1 Karczmarski J.

1 Krześniak N.

1 Laubitz D.

1 Paziewska A.

1 Rutkowski A.

1 Skrzypczak M.

1 Wolski J.

1 Woszczyński M.

1 Wyrwicz L. S.

1 Zagorowicz E.

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Pre-analytical-related variability influencing serum peptide profiles demonstrated in a mass spectrometry-based search for colorectal and prostate cancer biomarkers

Gene expression alterations induced by low molecular weight heparin during bowel anastomosis healing in rats