Influence of Fasting Status and Sample Preparation on Metabolic Biomarker Measurements in Postmenopausal Women

<div><p>Background</p><p>Epidemiologic data linking metabolic markers-such as insulin, insulin-like growth factors (IGFs)-and adipose tissue-derived factors with cancer are inconsistent. Between-study differences in blood collection protocols, in particular participant’s fasting status, may influence measurements.</p><p>Methods</p><p>We investigated the impact of fasting status and blood sample processing time on components of the insulin/IGF axis and in adipokines in a controlled feeding study of 45 healthy postmenopausal-women aged 50–75 years. Fasting blood samples were drawn (T0), after which subjects ate a standardized breakfast; subsequent blood draws were made at 1 hour (T1), 3 hours (T3), and 6 hours (T6) after breakfast. Serum samples were assayed for insulin, C-peptide, total- and free-IGF-I, IGF-binding protein [BP]-1 and -3, total and high molecular weight (HMW)-adiponectin, retinol binding protein-4, plasminogen activator inhibitor (PAI)-1, and resistin.</p><p>Results</p><p>Insulin and C-peptide levels followed similar postprandial trajectories; intra-class correlation coefficients [ICC] for insulin = 0.75, (95%CI:0.64–0.97) and C-peptide (ICC = 0.66, 95%CI:0.54–0.77) were similarly correlated in fasting (Spearman correlation, <i>r</i> = 0.78, 95%CI:0.64–0.88) and postprandial states (T1, <i>r</i> = 0.77 (95%CI: 0.62–0.87); T3,<i>r</i> = 0.78 (95%CI: 0.63–0.87); T6,<i>r</i> = 0.77 (95%CI: 0.61–0.87)). Free-IGF-I and IGFBP-1 levels were also affected by fasting status, whereas total-IGF-I and IGFBP-3 levels remained unchanged. Levels of adipokines were largely insensitive to fasting status and blood sample processing delays.</p><p>Conclusion</p><p>Several components of the insulin/IGF axis were significantly impacted by fasting state and in particular, C-peptide levels were substantially altered postprandially and in a similar manner to insulin.</p></div

Characteristics of study subjects.

<p>Characteristics of study subjects.</p

Medians of insulin/IGF-1 axis parameters and adipokines after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast, and intraclass correlations between measurements on blood samples collected at all time-points.

<p>Medians of insulin/IGF-1 axis parameters and adipokines after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast, and intraclass correlations between measurements on blood samples collected at all time-points.</p

Medians and median absolute deviation (MAD) of serum (A) adiponectin, (B) HMW adiponectin, (C) resistin, (D) PAI-I, and (E) RBP-4 after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast.

<p>Medians and median absolute deviation (MAD) of serum (A) adiponectin, (B) HMW adiponectin, (C) resistin, (D) PAI-I, and (E) RBP-4 after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast.</p

Medians and median absolute deviation of serum (A) C-peptide, (B) insulin, (C) total IGF-1, (D) free-IGF-1, (E) IGFBP-1, and (F) IGFBP-3 after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast.

<p>Medians and median absolute deviation of serum (A) C-peptide, (B) insulin, (C) total IGF-1, (D) free-IGF-1, (E) IGFBP-1, and (F) IGFBP-3 after an overnight fast (T0), and 1 hour (T1), 3 hours (T3), and 6 hours (T6) after the research breakfast.</p

Spearman’s correlation matrix for the insulin/IGF-1 axis parameters and adipokines measured on blood samples collected 1 hour after the research breakfast (T1).

<p>Spearman’s correlation matrix for the insulin/IGF-1 axis parameters and adipokines measured on blood samples collected 1 hour after the research breakfast (T1).</p

Adjusted^a odds ratios (ORs) and 95% confidence intervals (CIs) for the associations between circulating metabolite concentrations and esophageal adenocarcinoma incidence, in the FINBAR Study: 2002–2004.

<p>Adjusted<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0190325#t002fn001" target="_blank">^a</a> odds ratios (ORs) and 95% confidence intervals (CIs) for the associations between circulating metabolite concentrations and esophageal adenocarcinoma incidence, in the FINBAR Study: 2002–2004.</p

Mean circulating metabolite concentrations among esophageal adenocarcinoma cases and controls, in the FINBAR Study: 2002–2004.

<p>Mean circulating metabolite concentrations among esophageal adenocarcinoma cases and controls, in the FINBAR Study: 2002–2004.</p

Relationship of circulating insulin-like growth factor-I and binding proteins 1-7 with mammographic density among women undergoing image-guided diagnostic breast biopsy.

BACKGROUND: Mammographic density (MD) is a strong breast cancer risk factor that reflects fibroglandular and adipose tissue composition, but its biologic underpinnings are poorly understood. Insulin-like growth factor binding proteins (IGFBPs) are markers that may be associated with MD given their hypothesized role in breast carcinogenesis. IGFBPs sequester IGF-I, limiting its bioavailability. Prior studies have found positive associations between circulating IGF-I and the IGF-I:IGFBP-3 ratio and breast cancer risk. We evaluated the associations of IGF-I, IGFBP-3, and six other IGFBPs with MD. METHODS: Serum IGF measures were quantified in 296 women, ages 40-65, undergoing diagnostic image-guided breast biopsy. Volumetric density measures (MD-V) were assessed in pre-biopsy digital mammograms using single X-ray absorptiometry. Area density measures (MD-A) were estimated by computer-assisted thresholding software. Age, body mass index (BMI), and BMI RESULTS: IGF-I and IGFBP-3 were not strongly associated with MD after BMI adjustment. In multivariable analyses among premenopausal women, IGFBP-2 was positively associated with both percent MD-V (β = 1.49, p value = 0.02) and MD-A (β = 1.55, p value = 0.05). Among postmenopausal women, positive relationships between IGFBP-2 and percent MD-V (β = 2.04, p = 0.003) were observed; the positive associations between IGFBP-2 and percent MD-V were stronger among lean women (BMI  CONCLUSIONS: In this comprehensive study of IGFBPs and MD, we observed a novel positive association between IGFBP-2 and MD, particularly among women with lower BMI. In concert with in vitro studies suggesting a dual role of IGFBP-2 on breast tissue, promoting cell proliferation as well as inhibiting tumorigenesis, our findings suggest that further studies assessing the role of IGFBP-2 in breast tissue composition, in addition to IGF-1 and IGFBP-3, are warranted.</p