Paricalcitol attenuates indoxyl sulfate-induced apoptosis through the inhibition of MAPK, Akt, and NF-kB activation in HK-2 cells

• Published in: The Korean journal of internal medicine Vol. 34; no. 1; pp. 146 - 155
• Main Authors: Park, Jung Sun, Choi, Hoon In, Bae, Eun Hui, Ma, Seong Kwon, Kim, Soo Wan
• Format: Journal Article
• Language: English
• Published: Korea (South) The Korean Association of Internal Medicine 01.01.2019; 대한내과학회
• Subjects: apoptosis; Apoptosis - drug effects; Cell Line; Cell Survival - drug effects; Epithelial Cells - drug effects; Epithelial Cells - pathology; Ergocalciferols - pharmacology; Humans; indican; Indican - antagonists & inhibitors; Indican - toxicity; Kidney Tubules, Proximal - drug effects; Kidney Tubules, Proximal - metabolism; Kidney Tubules, Proximal - pathology; MAP Kinase Signaling System - drug effects; Original; paricalcitol; Phosphorylation; Protective Agents - pharmacology; Proto-Oncogene Proteins c-akt - antagonists & inhibitors; proximal tubular epithelial cell; Reactive Oxygen Species - metabolism; signal transduction; Signal Transduction - drug effects; Transcription Factor RelA - antagonists & inhibitors; 내과학; Proximal tubular epithelial cell; Indican; Signal transduction; Paricalcitol; Apoptosis
• ISSN: 1226-3303; 2005-6648
• DOI: 10.3904/kjim.2016.298

Indoxyl sulfate (IS) is a uremic toxin and an important causative factor in the progression of chronic kidney disease. Recently, paricalcitol (19-nor-1,25-dihydroxyvitamin D2) was shown to exhibit protective effects in kidney injury. Here, we investigated the effects of paricalcitol treatment on IS-induced renal tubular injury. The fluorescent dye 2',7'-dichlorofluorescein diacetate was used to measure intracellular reactive oxygen species (ROS) following IS administration in human renal proximal tubular epithelial (HK-2) cells. The effects of IS on cell viability were determined using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays and levels of apoptosis-related proteins (Bcl-2-associated protein X [Bax] and B-cell lymphoma 2 [Bcl-2]), nuclear factor-κB (NF- κB) p65, and phosphorylation of mitogen-activated protein kinase (MAPK) and protein kinase B (Akt) were determined by semiquantitative immunoblotting. The promoter activity of NF-κB was measured by luciferase assays and apoptosis was determined by f low cytometry of cells stained with f luorescein isothiocyanate-conjugated Annexin V protein. IS treatment increased ROS production, decreased cell viability and induced apoptosis in HK-2 cells. IS treatment increased the expression of apoptosis-related protein Bax, decreased Bcl-2 expression, and activated phosphorylation of MAPK, NF-κB p65, and Akt. In contrast, paricalcitol treatment decreased Bax expression, increased Bcl-2 expression, and inhibited phosphorylation of MAPK, NF-κB p65, and Akt in HK-2 cells. NF-κB promoter activity was increased following IS, administration and was counteracted by pretreatment with paricalcitol. Additionally, flow cytometry analysis revealed that IS-induced apoptosis was attenuated by paricalcitol treatment, which resulted in decreased numbers of fluorescein isothiocyanate-conjugated Annexin V positive cells. Treatment with paricalcitol inhibited IS-induced apoptosis by regulating MAPK, NF-κB, and Akt signaling pathway in HK-2 cells.