High glucose induces macrophage inflammatory protein-3 alpha in renal proximal tubule cells via a transforming growth factor-beta 1 dependent mechanism.

Abstract

BACKGROUND:

Hyperglycaemia is a causative factor in the pathogenesis of diabetic nephropathy, known to induce chemokines in the kidney. Macrophage inflammatory protein-3 alpha (MIP-3 alpha) is a CC chemokine that has been reported to attract memory T lymphocytes. Our previous microarray study showed significant increased level of MIP-3 alpha in high glucose-induced transcriptional profile in renal proximal tubule cells. Transforming growth factor-beta1 (TGF-beta1) is a key regulator in inflammation and fibrosis in diabetes mellitus setting.

METHODS:

This study aimed to determine the role of TGF beta 1 in high glucose-induced MIP-3 alpha expression. An in vitro model of human proximal tubular cells (HK-2 cells) and an in vivo model of the transgenic (mRen-2)27 diabetic rat, well characterized as a model of human diabetic nephropathy, were used. Small interfering RNA technology was used to silence TGF-beta1 gene in HK-2 cells and subsequent experiments were performed to measure mRNA and protein levels of MIP-3 alpha using real time reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Immunohistochemistry was used to measure the protein level of MIP-3 alpha and CD3 a marker of T lymphocytes in the in vivo model.

RESULTS:

MIP-3 alpha mRNA and protein expression was increased in HK-2 cells by high glucose and TGF-beta1. MIP-3 alpha was up-regulated in the dilated tubules of diabetic rats compared with non-diabetic control animals and CD3 was found to be present around the dilated tubules expressing MIP-3 alpha. This up-regulation was attenuated in the presence of an angiotensin-converting enzyme (ACE) inhibitor. MIP-3 alpha expression significantly decreased in cells in which the TGF-beta1 gene was silenced using small interfering RNA. Furthermore, exposure to high glucose did not induce MIP-3 alpha expression in TGF-beta1 gene silenced cells compared with wild-type cells.

CONCLUSIONS:

In summary, we have uniquely demonstrated that high glucose increases MIP-3 alpha through a TGF beta 1 dependent pathway, suggesting the centrality of TGF-beta1 in both the inflammatory and previously demonstrated fibrotic responses in diabetic nephropathy.

• Authors: Weier Qi, Xinming Chen, Yuan Zhang, John Holian, Ellein Mreich, Richard E Gilbert, Darren J Kelly, Carol A Pollock
• Journal: Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association (Nephrol Dial Transplant) Vol. 22 Issue 11 Pg. 3147-53 (Nov 2007) ISSN: 0931-0509 [Print] England
• PMID: 17664181 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Chemokine CCL20
• DNA Primers
• RNA, Messenger
• Transforming Growth Factor beta1
• Glucose

Topics

• Animals
• Cell Line
• Chemokine CCL20 (biosynthesis, genetics)
• DNA Primers
• Diabetes Mellitus, Experimental (physiopathology)
• Diabetic Nephropathies (physiopathology)
• Female
• Glucose (pharmacology)
• Humans
• Immunohistochemistry
• Kidney Tubules, Proximal (drug effects, physiology)
• RNA, Messenger (genetics)
• Rats
• Reverse Transcriptase Polymerase Chain Reaction
• Transforming Growth Factor beta1 (physiology)