Supplementary Material for: Danhong Injection Protects Against Hypertension-Induced Renal Injury Via Down-Regulation of Myoglobin Expression in Spontaneously Hypertensive Rats

<b><i>Background/Aims:</i></b> High blood pressure is a major risk factor for chronic kidney disease. Currently, single-target anti-hypertensive drugs are not designed for high blood pressure-related organ damages. Danhong injection (DHI), made from the aqueous extracts of <i>Radix Salviae miltiorrhizae</i> and <i>Flos Carthamus tinctorius</i>, has various pharmacological effects, including BP lowering in SHR, mediated by the reduction of vascular remodeling and the up-regulation of Kallikrein-kinin system published recently by our team, yet if it renders renal protection remains unknown. The current study demonstrated a protective role of DHI in renal injury caused by hypertension and identified its molecular targets in the kidney of spontaneously hypertensive rats (SHR). <b><i>Methods:</i></b> Adult SHR and age/gender-matched normotensive Wistar-Kyoto (WKY) rats were treated with DHI, Losartan, or saline for 4 weeks. Serum levels of Creatinine (CRE), Micro-albumin (mAlb), Beta2-microglobulin (β2-MG), and Uric acid (UA) were detected using ELISA kits. Renal pathology was examined by hematoxylin and Eosin (H&amp;E) stains. Microarray analysis was performed on kidney tissues, and gene expression changes were validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot analyses. <b><i>Results:</i></b> Renal histopathological scores showed that SHR exhibited serious kidney injury compared to normotensive WKY rats. The intervention with DHI potently suppressed the renal injury biomarker (KIM-1) and kidney lesions compared to the untreated hypertensive subjects. Microarray analysis revealed that among the 124 genes that were differentially expressed by DHI treatment in SHR kidney, down-regulation of renal myoglobin (Mb) gene was the most prominent and was subsequently confirmed by qRT-PCR and Western blot analysis. <b><i>Conclusion:</i></b> Hypertension-induced renal injury in SHR may be alleviated by DHI in part by local suppression of Kidney injury molecule-1 and down-regulation of Myoglobin. However, if this effect is independent of the known anti-hypertensive action of DHI in blood vessel remains to be determined.