Differences in circulating concentrations of total, free and bound leptin relate to gender and body composition in adult humans

Abstract

We describe a radioimmunoassay (RIA) for total leptin and a gel filtration procedure for the separation of free and bound leptin in human serum. The RIA, based on a locally prepared antibody, has a minimum detection limit of 0.9 ng/mL, a working range (CV < 10%) of 2.5-50 ng/mL, inter-assay precision of 10.2, 7.2 and 8.9%CV at 7.9, 15.4 and 30.0 ng/mL, respectively, 94% recovery of exogenous leptin (range 81.1-120.6%), exhibited parallelism and demonstrated no significant cross-reactivity or interferences. A difference plot of results from this method and those from a commercially available kit (Linco Research) demonstrated satisfactory agreement up to concentrations of 50 ng/mL total leptin, with no significant bias. A gender-dependent correlation was obtained between body mass index (BMI) and total leptin (r = 0.91, P<0.001, n = 75 for men; r = 0.79, P<0.001, n = 72 for women), with women having higher leptin concentrations than men for any given BMI. Gel filtration studies (inter-assay precision: 4.7%CV, n = 18) demonstrated that a variable fraction (between 10% and 40%) of total leptin in serum was bound with high affinity (Keq = 1.0-1.45 x 10(9) L/mol) to a non-albumin, non-lipid macromolecule. Binding affinities were found to be similar irrespective of gender or fat mass. A significant positive correlation between free or bound leptin concentrations and BMI was obtained for both men and women (r = 0.87-0.94); free and bound leptin concentrations were also significantly higher in women (P<0.01) than in men for any given BMI, and higher in obese (P<0.01) than in lean individuals. We conclude that leptin 'resistance' associated with obesity cannot be accounted for by reduced free leptin concentrations in serum and that the methods described are suitable for the investigation of total, free and bound leptin for both clinical and research purposes.

Lack of association between body mass index and plasma adiponectin levels in healthy adults

“…The regression analysis of male leptin to BMI was based on data from 10 groups (Figure 2a The plasma level of leptin was shown to be significantly higher in women compared with men. 31 This gender difference in plasma leptin level was also significant in our data analysis (mean ± s.d., 7.75 ± 7.13 vs 22.5 ± 12.2 ng ml À1 , M vs F). In contrast, the plasma level of adiponectin was not significantly different between men and women in our data analysis (mean ± s.d., 7.34 ± 2.74 vs 9.31 ± 4.81 mg ml À1 , M vs F).…”

“…Furthermore, studies reporting mean values combining observations from both genders were not used because of the known gender difference in percentage body fat, [28][29][30] fat distribution 29 and plasma leptin level. 31 Overall, the three exclusion criteria, concerns on subject health, young age and a lack of gender focus, led to the majority of the data exclusion.…”

“…27 We limited the analysis to those studies providing gender-specific data because women are known to have higher percentage body fat than men at the same BMI. [28][29][30] To validate the quality of our analyses, plasma leptin level, which was known to increase with obesity and found to be higher in women, 31 was also analyzed here as a positive control.…”

Effects of a beverage rich in (poly)phenols on established and novel risk markers for vascular disease in medically uncomplicated overweight or obese subjects: A four week randomized placebo-controlled trial

“…Insulin was measured using the Mercodiaultrasenstive insulin ELISA. Leptin analysis was performed using an in-house assay; the methodology has been previously published (19). t-PA was measured with an ELISA (Biopool AB, Umea, Sweden).…”

The relationship between the insulin-like growth factor-1 axis, weight loss, an inflammation-based score and survival in patients with inoperable non-small cell lung cancer

“…Leptin was quantified by an in-house radio-immunoassay, which has been described and validated elsewhere 19 .…”