gms | German Medical Science

Objectives: The immune system is an important initiator of fracture healing. Cytokines secreted by immune cells as well as chemokines secreted by residual cells largely impact the tissue surrounding the fracture gap. Impaired wound/fracture healing frequently observed in diabetics might come from an altered inflammatory response after fracture. Thus, this study aimed at investigating: (i) the effects of monocytic cells cultured under diabetic conditions on osteoblast migration and differentiation, and (ii) the effect of diabetic conditions on the migration of THP-1 cells towards chemokines.

Methods: For collection of RNA and conditioned medium THP-1 cells were cultivated under normoglycemic (+ mannitol) and hyperglycemic (+ glucose) conditions ± insulin for 2 days. Expression/secretion of chemo- and cytokines in THP-1 cells was determined by RT-PCR and ELISA. Migration (under agar spot assay) and differentiation (AP activity, matrix mineralization) of primary human osteoblasts (hOBs) was analyzed in presence of the immune cell conditioned medium. Migration of THP-1 cells to different chemokines was further analyzed under normo- or hyperglycemic medium by Boyden Chamber Assay.

Results and conclusion: THP-1 cells exposed to diabetic conditions show an altered cytokine expression (especially IL-8, TNF-α, and CXCL9) mainly in response to insulin. While IL-8 response was stronger under normoglycemic conditions, TNF-α response was stronger under hyperglycemic conditions (2-fold vs. 3-fold increase). IL-1β expression is 3-fold increased in presence of insulin, resulting in a 10-fold increased secretion under hyperglycemic conditions. Secretion of TNF-α by THP-1 cells is doubled in presence of insulin. Similarly, CXCL9 and CCL2 are more secreted when treating the THP-1 cells with insulin. This altered cytokine expression/secretion is expected to have a direct effect on the migration and differentiation of osteogenic cells. Migration of osteogenic cells was increased by conditioned medium from THP-1 cells but significantly reduced when the THP-1 cells have been cultured under hyperglycemic conditions before. Similarly, THP-1 conditioned medium itself had a positive effect on the differentiation (higher AP activity and matrix mineralization) of hOBs, but this was reduced when the THP-1 cells have been cultured under hyperglycemic conditions before. Additionally, THP-1 cells themselves show reduced migration to chemokines in hyperglycemic conditions. Summarizing our results, the release of chemokines and cytokines from THP-1 cells is altered under diabetic conditions, which in turn has a negative effect on the migration and differentiation of osteogenic cells. This altered cytokine response of monocytic cells due to diabetic conditions opens up new possibilities for treatment of diabetic healing disorders.