High glucose alters tendon homeostasis through downregulation of the AMPK/Egr1 pathway

Yu-Fu Wu,Jingyu Sun,Hsing-Kuo Wang,Jui-Sheng Sun,Yuan-Hung Chao,Hong-Wei Chang

doi:10.1038/srep44199

Abstract

Diabetes mellitus (DM) is associated with higher risk of tendinopathy, which reduces tolerance to exercise and functional activities and affects lifestyle and glycemic control. Expression of tendon-related genes and matrix metabolism in tenocytes are essential for maintaining physiological functions of tendon. However, the molecular mechanisms involved in diabetic tendinopathy remain unclear. We hypothesized that high glucose (HG) alters the characteristics of tenocyte. Using in vitro 2-week culture of tenocytes, we found that expression of tendon-related genes, including Egr1, Mkx, TGF-β1, Col1a2, and Bgn, was significantly decreased in HG culture and that higher glucose consumption occurred. Down-regulation of Egr1 by siRNA decreased Scx, Mkx, TGF-β1, Col1a1, Col1a2, and Bgn expression. Blocking AMPK activation with Compound C reduced the expression of Egr1, Scx, TGF-β1, Col1a1, Col1a2, and Bgn in the low glucose condition. In addition, histological examination of tendons from diabetic mice displayed larger interfibrillar space and uneven glycoprotein deposition. Thus, we concluded that high glucose alters tendon homeostasis through downregulation of the AMPK/Egr1 pathway and the expression of downstream tendon-related genes in tenocytes. The findings render a molecular basis of the mechanism of diabetic tendinopathy and may help develop preventive and therapeutic strategies for the pathology.

Highlights

Diabetes mellitus (DM) is associated with higher risk of tendinopathy, which reduces tolerance to exercise and functional activities and affects lifestyle and glycemic control
After surgical repair in the rotator cuff, diabetic patients with poorer glycemic control were found to have a higher retear rate[24]. These findings suggest that long-sustained high concentration of blood glucose is associated with diabetic tendinopathy
We introduced an in vitro diabetic-mimicking culture condition to investigate the effect of high glucose on tenocyte traits

Summary

Results

No significant effect on cell growth and apoptosis. Throughout the two-week culture, similar viability levels of tenocytes were observed in low- and high-glucose medium (LG and HG) with or without insulin at each time point (Fig. 1A). To examine whether higher glucose consumption resulted in even lower tendon-related gene expression, we tested the expression of three crucial transcription factors, including Scx, Mkx, and Egrl. At day 7, expression of Egr[1], TGF-β1, and Bgn were suppressed by insulin under the HG condition, with levels ~67%, ~68%, and ~84% as high as the non-insulin HG group, respectively (Fig. 4A and C). After inhibiting Egr[1] to a level of 0.28 ± 0.04-times as high as that of the control group, expression levels of all the target genes, including TGF-β1, Bgn, Col1a1, and Col1a2, were significantly suppressed (Fig. 5B–D). We hypothesized that increased glucose consumption in tenocytes under HG conditions resulted in inactivation of AMPK signaling, which in turn affected the expression of tendon-related genes. Through quantitative image analysis by ImageJ software, we confirmed that the average interfibrillar area was significantly larger in diabetic tendons than in healthy tendons (Fig. 7B)

Discussion

Author Contributions

Additional Information