Aim
The purpose of present study is to compare the protective effects of raw and salt-processed Achyranthes on lipopolysaccharide (LPS) - induced acute kidney injury in mice and chemically characterize their extracts. Method: The monomer components of raw and salt-processed Achyranthes extracts were analyzed using high performance liquid chromatography (HPLC). The aggregation and distribution of 2-Deoxy-D-glucose (2-DG) near infrared fluorescence probe in mice was examined with a small animal imaging systems. The pathological and morphological changes of kidneys were observed by H&E staining, and the serum urea nitrogen (BUN) and serum creatinine (Scr) levels were used to evaluate the renal function. The levels of cytokines in serum were detected by cytometric bead array. Flow cytometry assay was performed to assess the apoptosis and reactive oxygen species (ROS) in the kidney cells, and cell surface marker expression including CD45+, F4/80+, and Ly-6G+. The estrogenic activities of the raw and salt-processed Achyranthes were observed by uterine weight gain test in sexually immature mice. Western blot was used to detect the protein expression levels in the kidney.
Background
Traditional Chinese medicine suggests that Radix Achyranthis Bidentatae nourishes and protects the kidneys, the effect of which is enhanced following a salt treatment. Raw and salt-processed Achyranthes bidentata are produced via different processing techniques from the same crude Achyranthes root. The anti-inflammatory and immunomodulatory properties of this plant have been verified earlier. However, there is a scarcity of experimental evidence for the renal-protective effects.
Conclusions
The renal-protective effect of raw and salt-processed Achyranthes was exhibited through antiapoptotic and antioxidant mechanisms via an estrogen-like pathway, along with a modulation of the inflammatory response by regulating immune cells. Ginsenoside Ro and Chikusetsu saponin IVa were found to be the key factors to enhance the protective effect of salt-processed Achyranthes.
Results
Chemical analysis showed that the salt-processed Achyranthes contained more ginsenoside Ro and chikusetsusaponin Ⅳa than the raw Achyranthes, but there was no difference in the contents of β-ecdysterone, 25R-inokosterone, and 25S-inokosterone.in vivo near-infrared fluorescence imaging showed a significant reduced inflammation in the AKI mice. Histological studies showed that the raw and salt-processed Achyranthes markedly decreased the inflammatory infiltration, swelling and vacuolar degeneration in renal tissues and the Scr and BUN. Importantly, the raw and salt-processed Achyranthes extracts demonstrated different degrees of inhibition on the LPS-induced AKI, with salt-processed Achyranthes showing better inhibition. Results of flow cytometry showed a significant inhibition of IFN-γ, TNF-α, and IL-2, and promoted IL-10, along with reduced macrophages (CD45 + F4/80+), neutrophils (CD45+ Ly-6G+) and phagocytes. Furthermore, the extracts reduced the accumulation of ROS and apoptosis in the kidney, and also regulated the expression of apoptosis marker proteins TLR4, Bcl-2, Bax, cleaved caspase 3 and cleaved caspase 9 levels. Notably, they increased ERα, ERβ, and GPR30 in the renal tissues of AKI mice and LPS non-treated mice. In the subsequent experiments, it was found that the raw and salt-processed Achyranthes extracts increased the uterine coefficient in sexually immature mice, improved the LPS-induced decrease in NRK52e cell viability, and reduced the apoptosis, which could be antagonized by ICI182, 780 (estrogen receptor-unspecific antagonist, Faslodex). Conclusions: The renal-protective effect of raw and salt-processed Achyranthes was exhibited through antiapoptotic and antioxidant mechanisms via an estrogen-like pathway, along with a modulation of the inflammatory response by regulating immune cells. Ginsenoside Ro and Chikusetsu saponin IVa were found to be the key factors to enhance the protective effect of salt-processed Achyranthes.