Abstract
Obesity is associated with significantly higher mortality rates, and excess adipose tissue is involved in respective pathologies. Here we established a human adipose tissue slice cultures (HATSC) model ex vivo. HATSC match the in vivo cell composition of human adipose tissue with, among others, mature adipocytes, mesenchymal stem cells as well as stroma tissue and immune cells. This is a new method, optimized for live imaging, to study adipose tissue and cell-based mechanisms of obesity in particular. HATSC survival was tested by means of conventional and immunofluorescence histological techniques, functional analyses and live imaging. Surgery-derived tissue was cut with a tissue chopper in 500 μm sections and transferred onto membranes building an air-liquid interface. HATSC were cultured in six-well plates filled with Dulbecco’s Modified Eagle’s Medium (DMEM), insulin, transferrin, and selenium, both with and without serum. After 0, 1, 7 and 14 days in vitro, slices were fixated and analyzed by morphology and Perilipin A for tissue viability. Immunofluorescent staining against IBA1, CD68 and Ki67 was performed to determine macrophage survival and proliferation. These experiments showed preservation of adipose tissue as well as survival and proliferation of monocytes and stroma tissue for at least 14 days in vitro even in the absence of serum. The physiological capabilities of adipocytes were functionally tested by insulin stimulation and measurement of Phospho-Akt on day 7 and 14 in vitro. Viability was further confirmed by live imaging using Calcein-AM (viable cells) and propidium iodide (apoptosis/necrosis). In conclusion, HATSC have been successfully established by preserving the monovacuolar form of adipocytes and surrounding macrophages and connective tissue. This model allows further analysis of mature human adipose tissue biology ex vivo.
Highlights
The prevalence of overweight and obesity, commonly measured by a high body mass index (BMI), is notably increasing worldwide [1]
It can thereby be concluded that human adipose tissue slice cultures (HATSC) provides a platform to investigate human adipose tissue in a controlled ex vivo setting with little ethical constraints
Jo et al did not find an alteration in cell volume during apoptosis, using mathematical models [59]
Summary
The prevalence of overweight and obesity, commonly measured by a high body mass index (BMI), is notably increasing worldwide [1]. Viability of human adipose tissue slice culture. The GSI Helmholtzzentrum fur Schwerionenforschung GmbH (Darmstadt, Germany) provided support in the form of salaries for FR. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section
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