Quantitative evaluation of Oryeongsan and its action on water regulation in renal inner medullary collecting duct cells

Ethnopharmacological relevance: Oryeongsan (ORS, Wulingsan) has been reported to possess renal protective effects from renal diseases such as diabetes-induced renal damage, and nephrocalcinosis. Aim of the study: This study was conducted to evaluate the quantitative analysis and the inhibitory effect of ORS on hypertonic stress-induced water channel and apoptosis in murine inner medullary collecting duct cell line (mIMCD-3). Materials and methods: Chromatographic and NMR spectroscopic analysis were performed and water balance regulation was determined by Western blot, RT-PCR, and immunofluorescnece. Results: Seven active principles (5-hydroxymethylfurfural, alismoxide, methyl-trans-cinnamate, adenine, guanosine, adenosine, and ferulic acid) in ORS were isolated and the structures were identified mainly by NMR spectroscopic analysis. In addition, contents of these metabolites in ORS were evaluated by HPLC analysis. Pretreatment with ORS significantly attenuated the hypertonic stress (175 mM NaCl)-induced increase in protein levels of AQP2 and apical membrane insertion. ORS also attenuated osmolyte sodium-myo-inositol transporter (SMIT) expression and tonicity-responsive enhancer binding protein (TonEBP) mRNA under hypertonic stress. Those actions of ORS presented the similar effect of PICA inhibitor which AQP2 expression throughout the inhibition of vasopressin-mediated cAMP/PKA signal pathway. On the other hand, pretreatment with ORS attenuated hypertonic stress -induced cell death. Hypertonic stress -induced Bax or caspase-3 expression was decreased by ORS, resulting in anti-apoptotic effect. Conclusions: The present data suggest that the beneficial effect of ORS in water balance and apoptosis against hypertonic stress of renal collecting ducts. (C) 2016 Published by Elsevier Ireland Ltd.