Differences and significance of macrophage subtypes and regulatory T lymphocytes in the expression of interstitial fibrosis in diabetic nephropathy at different stages.

BACKGROUND: Diabetic nephropathy (DN) is a microvascular complication of diabetes and a major cause of chronic renal failure in adults. DN is characterized by the infiltration of inflammatory cells in the renal tubular interstitium, resulting in severe interstitial fibrosis, ultimately leading to end-stage renal disease. However, the exact timing at which different phenotypes of inflammatory cells contribute to interstitial fibrosis in DN remains unclear. METHODS: We collected 45 renal tissues diagnosed with DN through renal biopsy between January 2014 and April 2019 from the Pathology Department of Putuo Hospital, affiliated with the Shanghai University of Traditional Chinese Medicine and the Pathology Department of Shanghai Medical College Fudan University. Immunohistochemistry staining was performed on paraffin-embedded sections to analyze the expression patterns of M1 and M2 macrophages and regulatory T lymphocytes. The severity of renal function impairment was categorized into three groups: CKD stage 1 (16 cases), CKD stage 2 (13 cases), and CKD stages 3–5 (16 cases). The degree of tubulointerstitial fibrosis was divided into three groups: mild fibrosis (16 cases), moderate fibrosis (18 cases), and severe fibrosis (11 cases). RESULTS: Our findings revealed significant differences in the levels of inflammatory cell infiltration CD68, CD163, CD3, and CD4, as well as in the M1/M2 ratio among the different groups of tubulointerstitial fibrosis. While the severity of interstitial fibrosis did not impact urinary protein excretion, it demonstrated statistically significant differences in renal function decline. When the samples were categorized based on the degree of renal dysfunction, there were significant differences in the levels of CD68, CD163, CD3, CD4, and Foxp3 among the three groups. The infiltration levels of macrophages and lymphocytes increased significantly with renal function decline, with M2-type macrophages being the predominant subtype. The infiltration level of inflammatory cells was associated with renal function decline but not with the ratio of proteinuria to interstitial fibrosis. Macrophage and lymphocyte infiltration levels were consistent, while regulatory T lymphocytes showed a closer association with M1-type macrophages. The rate of interstitial fibrosis was correlated with renal function indicators and macrophage infiltration, with a positive correlation observed with M2-type macrophages. DISCUSSION: Our study demonstrated an association between regulatory T lymphocytes and M1-type macrophages, which may contribute to early lesions of DN. However, their impact was limited. In the late stage of DN, M2-type macrophages significantly increased, inhibiting the immune response but also producing and releasing numerous pro-fibrotic factors, ultimately exacerbating fibrosis and worsening renal function. The phenotypes of macrophages determined the development and prognosis of DN. Therefore, regulating macrophage polarization and subtype transformation represented a promising target for DN treatment.