Browsing by Subject "serum"
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Item Characterization of serum-induced CYP1A1 expression and activity in mouse embryo fibroblasts(2005-07-07) Carmen-Veronica Naira Obianwu; Cornelis Elferink Ph.D.; Xiaodong Cheng Ph.D.; Jonathan B. Ward Ph.D.The AhR is a ligand-activated transcription factor that mediates the toxic effects of environmental contaminants that include TCDD. Using a TCDD dose-response treatment in MEFs, we observed a super induction of CYP1A1 with newborn calf serum (NCS) in the presence (10nM/15nM) of TCDD. In addition to NCS, fetal bovine serum (FBS) also has the capability to yield a CYP1A1 super induction. These results suggest that components within the sera affect the activity of the AhR and consequent CYP1A1 expression. To pursue this idea, characterization of the serum factors were investigated. The findings indicated that serum factor(s) in both sera are heat sensitive at 50◦C, withstand removal from charcoal stripping sera and are ¡Ý 10,000 kDa in size. Using RT-PCR, we found that NCS factors only, could super induce CYP1A1 at the gene level. Moreover, MEFs are the only cells observed in this study that are susceptible to CYP1A1 super induction.Item Development and Analytical Validation of an Enzyme-linked Immunosorbent Assay (ELISA) for the Measurement of Feline Alpha1-proteinase Inhibitor (fa1-PI) in Serum and Feces and the Evaluation of Fecal fa1-PI Concentrations in Cats with Idiopathic Inflammatory Bowel Disease or Gastrointestinal Neoplasia(2012-10-19) Burke, KathrinAlpha1-proteinase inhibitor (alpha1-PI) has been shown to be a useful marker of gastrointestinal protein loss in some species. The objectives of this study were, first, to develop and analytically validate an ELISA for the measurement of alpha1-PI in feces and serum from cats, and, second, to evaluate fecal alpha1-PI concentrations in healthy cats and cats with chronic gastrointestinal disease. The lower detection limits of the ELISA were 0.02 g/L for serum and 0.04 microgram/gram for feces. The observed-to-expected (O/E) ratios for serial dilutions of serum and fecal samples ranged from 100.0 to 129.7% (mean +/- SD: 112.2 +/- 9.9%) and 103.5 to 141.6% (115.6 +/- 12.8%), respectively. The O/E ratios for samples spiked with seven known concentrations of alpha1-PI ranged from 82.3 to 107.8% (94.7 +/- 7.6%) for serum and 78.5 to 148.7% (96.8 +/- 18.2%) for feces. The coefficients of variation for intra-assay and inter-assay variability were <7.9% and <12.1% for serum, and 5.3%, 11.8%, and 14.2% and 7.7%, 10.2%, and 20.4% for feces, respectively. Reference intervals were 0.6 to 1.4 g/L for serum and up to 1.6 microgram/g for feces. We conclude that this ELISA is sufficiently linear, accurate, precise, and reproducible. For the clinical evaluation, twenty cats with clinical signs of chronic gastrointestinal disease and 20 healthy control cats were enrolled. The diseased cats were grouped into two groups: mild to moderate idiopathic inflammatory bowel disease (IBD) (Group A; n=8) and severe IBD or neoplastic disease (Group B; n=12), based on histopathology results of endoscopic biopsies. Fecal alpha1-PI concentrations and serum concentrations of total protein, albumin, globulin, cobalamin, folate, pancreatic lipase immunoreactivity, and trypsin-like immunoreactivity were determined. Nineteen of the 20 diseased cats had increased fecal alpha1-PI concentrations, ranging from 1.9 to 233.6 microgram/g (normal range: <= 1.6 microgram/g). Fecal alpha1-PI concentrations were statistically significantly different between healthy cats and cats of Group A (median: 3.9 microgram/g, range: 1.3 to 9.2 microgram/g, P<0.001) or cats of Group B (median: 20.6 microgram/g, 4.3 to 233.6 microgram/g; P<0.001), and also between cats of Groups A and B (P<0.01). Hypoalbuminemia, hypoproteinemia, and hypocobalaminemia were detected in 88%, 83%, and 56% of the diseased cats, respectively. Our study suggests that increased fecal alpha1-PI concentrations in association with hypoalbuminemia may be a common finding in cats with IBD or GI neoplasia. Furthermore, alpha1-PI concentrations appear to be higher in cats with severe IBD or confirmed GI neoplasia when compared to cats with mild to moderate IBD.