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Updated: Jun 28, 2026
Isolation, Characterization, and Purification of Macrophages from Tissues Affected by Obesity-related Inflammation
Published on: April 3, 2017
Macrophage-secreted factors promote a profibrotic phenotype in human preadipocytes
Mayoura Keophiphath1, Vincent Achard, Corneliu Henegar
1Institut National de la Santé et de la Recherche Médicale (INSERM) Unité 872, Centre de recherche des Cordeliers, 15 rue de l'école de Médecine, 75005 Paris, France.
Abstract:
White adipose tissue (WAT) in obese humans is characterized by macrophage accumulation the effects of which on WAT biology are not fully understood. We previously demonstrated that macrophage-secreted factors impair preadipocyte differentiation and induce inflammation, and we described the excessive fibrotic deposition in WAT from obese individuals. Microarray analysis revealed significant overexpression of extracellular matrix (ECM) genes in inflammatory preadipocytes. We show here an organized deposition of fibronectin, collagen I, and tenascin-C and clustering of the ECM receptor alpha5 integrin, characterizing inflammatory preadipocytes. Anti-alpha5 integrin-neutralizing antibody decreased proliferation of these cells, underlining the importance of the fibronectin/integrin partnership. Fibronectin-cultured preadipocytes exhibited increased proliferation and expression of both nuclear factor-kappaB and cyclin D1. Small interfering RNA deletion of nuclear factor-kappaB and cyclin D1 showed that these factors link preadipocyte proliferation with inflammation and ECM remodeling. Macrophage-secreted molecules increased preadipocyte migration through an increase in active/phosphorylated focal adhesion kinase. Gene expression and neutralizing antibody experiments suggest that inhibin beta A, a TGF-beta family member, is a major fibrotic factor. Interactions between preadipocytes and macrophages were favored in a three-dimensional collagen I matrix mimicking the fibrotic context of WAT. Cell-rich regions were immunostained for preadipocytes, proliferation, and macrophages in the vicinity of fibrotic WAT from obese individuals. In conclusion, an inflammatory environment leads to profound modifications of the human preadipocyte phenotype, producing fibrotic components with increased migration and proliferation. This phenomenon might play a role in facilitating the constitution of quiescent preadipocyte pools and eventually in the maintenance and aggravation of increased fat mass in obesity.
Insights
Obesity-associated inflammation in white adipose tissue (WAT) alters preadipocytes, increasing fibrotic factors, proliferation, and migration. This contributes to fat mass expansion in obesity.
Area of Science:
- Adipose tissue biology
- Cellular and molecular mechanisms of obesity
- Extracellular matrix remodeling
Background:
- Obesity involves macrophage accumulation in white adipose tissue (WAT), but its impact on WAT biology is unclear.
- Previous work showed macrophage factors impair preadipocyte differentiation and increase WAT fibrosis.
- Microarray analysis revealed elevated extracellular matrix (ECM) gene expression in inflammatory preadipocytes.
Purpose of the Study:
- To investigate the phenotypic changes in human preadipocytes within an inflammatory WAT environment.
- To identify key molecular players in ECM production, proliferation, and migration of inflammatory preadipocytes.
- To explore the role of fibronectin-integrin interactions and macrophage-secreted factors in preadipocyte behavior.
Main Methods:
- Analysis of ECM gene expression in inflammatory preadipocytes.
- Immunostaining for fibronectin, collagen I, tenascin-C, and alpha5 integrin.
- Use of anti-alpha5 integrin-neutralizing antibody to assess proliferation.
- Small interfering RNA (siRNA) to deplete nuclear factor-kappaB (NF-kB) and cyclin D1.
- Assessment of preadipocyte migration and focal adhesion kinase (FAK) activity.
- Gene expression analysis and neutralizing antibody experiments for inhibin beta A.
- Three-dimensional (3D) cell culture in collagen I matrix.
- Immunohistochemistry on WAT samples from obese individuals.
Main Results:
- Inflammatory preadipocytes exhibit organized deposition of fibronectin, collagen I, tenascin-C, and alpha5 integrin.
- Anti-alpha5 integrin antibody reduced preadipocyte proliferation, highlighting the fibronectin/integrin pathway.
- Fibronectin exposure increased preadipocyte proliferation and expression of NF-kB and cyclin D1.
- NF-kB and cyclin D1 depletion linked preadipocyte proliferation to inflammation and ECM remodeling.
- Macrophage factors enhanced preadipocyte migration via increased FAK phosphorylation.
- Inhibin beta A was identified as a major fibrotic factor.
- Interactions between preadipocytes and macrophages were enhanced in a 3D collagen I matrix.
- Obese WAT showed increased preadipocytes, proliferation, and macrophages near fibrotic areas.
Conclusions:
- Inflammatory environments profoundly alter human preadipocyte phenotype, promoting fibrotic component production, migration, and proliferation.
- The fibronectin/integrin pathway and factors like inhibin beta A are crucial in this process.
- These cellular changes may facilitate the formation of quiescent preadipocyte pools, contributing to increased fat mass in obesity.
