A Method for Automated Histological Characterisation of Human Peripheral and Orbital Adipose Tissue

Abstract

Purpose: : Adipose depots represent targets of endocrine disease. The metabolic syndrome has a direct effect on omental fat where adipocyte hypertrophy and hyperplasia give rise to obesity. By contrast, orbital fat is vulnerable to adipogenesis in states of thyroid dysregulation. Published methods to quantify adipose cellularity involve either subjective counting or enzymatic tissue disruption. In this study, we describe an automated technique to characterise adipocyte morphology in human adipose depots.

Methods: : Paired omental (OM) and subcutaneous (SC) fat was obtained from 8 patients undergoing elective laparotomy (median BMI 26.4kg/m²). Orbital fat (OF) was harvested from 14 patients (7 female) undergoing orbital or lid surgery. Patients with orbital tumours, those on exogenous corticosteroids, or with underlying endocrine disease were excluded from the study. Formalin fixed 3µm sections were stained with haematoxylin and eosin and photographed in triplicate using an Olympus– BH–2–RFC microscope and MicroFire^TM – S99808 camera. Image–Pro^R Plus (v4.0, Media Cybernetics) parameters were optimised to identify an adipocyte outline in section for analysis.

Results: : Three parameters (cell diameter, perimeter, feret) confirmed that OM cells were significantly smaller than the SC adipocytes (p<0.05). BMI correlated significantly with OM diameter (p<0.05) but not SC. In general, OF parameters were similar to those of OM, with the exception of the radius ratio that was larger for the OF depot. Median cell diameters were OM 52.1µm (range 47.1–77.3); SC 78.0µm (range 53.8–112.6); and OF 52.6µm (35.4–64.4). Cell count/area of interest did not differentiate significantly between depots, although area of the cell achieved borderline significance. The OM depot had statistically the least roundness (p<0.05).

Conclusions: : Cell diameter, perimeter and roundness were identified as optimal parameters. SC adipocytes were larger than OM and OF depots, and the OF cells resembled the OM in morphology. Consistent with earlier studies, OM cell diameter increased with BMI, suggesting a significant contribution of cell hypertrophy to increasing fat mass. Our methodology generates detailed descriptive data and represents a powerful and flexible tool for characterising adipocyte morphology

Keywords: imaging/image analysis: non-clinical • comparative anatomy • computational modeling