Diabetic nephropathy (DN) is a frequent and severe complication of diabetes that is structurally characterized by glomerular basement membrane thickening, extracellular matrix accumulation, and destabilization of podocyte foot processes. MicroRNAs (miRNAs) are dysregulated in DN, but identification of the specific miRs involved remains incomplete. Here, we confirm that the peripheral blood from patients with diabetes and the kidneys of animals with type 1 or 2 diabetes have low levels of miR-23b compared with those of their nondiabetic counterparts. Furthermore, exposure to high glucose downregulated miR-23b in cultured kidney cells. In contrast, renal expression of Ras GTPase-activating protein SH3 domain-binding protein 2 (G3BP2), a putative miR-23b target, increased in DN. In vitro, overexpression of miR-23b decreased, and inhibition of miR-23b increased, G3BP2 expression levels. Bioinformatics analysis also revealed p53 binding sites in the miR-23b promoter; in vitro inhibition of p53 or the upstream p38 mitogen-activated protein kinase (p38MAPK) upregulated miR-23b expression in high-glucose conditions. In turn, inhibition of G3BP2 or overexpression of miR-23b downregulated p53 and p38MAPK expression in high-glucose conditions. In vivo, overexpression of miR23b or inhibition of p53 in db/db mice reversed hyperalbuminuria and kidney fibrosis, whereas miR-23b antagomir treatment promoted renal fibrosis and increased albuminuria in wild-type mice. These data suggest that hyperglycemia regulates pathogenic processes in DN through an miR-23b/G3BP2 feedback circuit involving p38MAPK and p53. In conclusion, these results reveal a role for miR-23b in DN and indicate a novel potential therapeutic target.

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Diabetic nephropathy is characterized by persistent albuminuria, progressive decline in GFR, and secondary hypertension. MicroRNAs are dysregulated in diabetic nephropathy, but identification of the specific microRNAs involved remains incomplete. Here, we show that the peripheral blood from patients with diabetes and the kidneys of animals with type 1 or 2 diabetes have low levels of microRNA-25 (miR-25) compared with those of their nondiabetic counterparts. Furthermore, treatment with high glucose decreased the expression of miR-25 in cultured kidney cells. In / mice, systemic administration of an miR-25 agomir repressed glomerular fibrosis and reduced high BP. Notably, knockdown of miR-25 in normal mice by systemic administration of an miR-25 antagomir resulted in increased proteinuria, extracellular matrix accumulation, podocyte foot process effacement, and hypertension with renin-angiotensin system activation. However, excessive miR-25 did not cause kidney dysfunction in wild-type mice. RNA sequencing showed the alteration of miR-25 target genes in antagomir-treated mice, including the Ras-related gene CDC42. , cotransfection with the miR-25 antagomir repressed luciferase activity from a reporter construct containing the CDC42 3' untranslated region. In conclusion, these results reveal a role for miR-25 in diabetic nephropathy and indicate a potential novel therapeutic target for this disease.

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Abstract. For assessment of biosimilarity, the US Food and Drug Administration (FDA) recommends a stepwise approach for obtaining the totality-of-the-evidence for demonstrating biosimilarity between a proposed biosimilar product and an innovative (reference) biological product. The stepwise approach starts with analytical studies for functional and structural characterization at various stages of manufacturing process of the proposed biosimilar product. Analytical similarity assessment involves identification of critical quality attributes (CQAs) that are relevant to clinical outcomes. FDA proposes first classifying the identified CQAs into three tiers according to their criticality or risking ranking relevant to clinical outcomes and then performing equivalence test (for CQAs in Tier 1), quality range approach (for CQAs in Tier 2), and raw data or graphical presentation (for CQAs in Tier 3) for obtaining totality-of-the-evidence for demonstrating biosimilarity between the proposed biosimilar product with the reference product. In practice, some debatable issues are evitably raised due to this complicated process of analytical similarity assessment. In this article, these debatable are described and discussed.KEY WORDS: equivalence test; fixed SD approach; quality range approach; stepwise approach; tiered approach; totality-of-the-evidence.