Rehmannioside A inhibits the activity of CYP3A4, 2C9 and 2D6 in vitro.

To assess the effect of Rehmannioside A on CYP450s activity and to estimate its inhibitory properties.The effect of Rehmannioside A on the activity of major CYP450s in human liver microsomes (HLMs) was assessed with the corresponding substrates and marker reactions, and compared with a blank control and the respective inhibitors. Suppression of CYP3A4, 2C9 and 2D6 was assessed by the dose-dependent assay and fitted with non-competitive or competitive inhibition models. The inhibition of CYP3A4 was determined in a time-dependent manner.Rehmannioside A suppressed the activity of CYP3A4, 2C9, and 2D6 with IC50 values of 10.08, 12.62, and 16.43 µM, respectively. Suppression of CYP3A4 was fitted to a non-competitive model with Ki value of 5.08 µM, whereas CYP2C9 and 2D6 were fitted to a competitive model with Ki values of 6.25 and 8.14 µM. Additionally, the inhibitory effect on CYP3A4 was time-dependent with KI value of 8.47 µM-1 and a Kinact of 0.048 min-1.In vitro suppression of CYP3A, 2C9 and 2D6 by Rehmannioside A indicated that Rehmannioside A or its source herbs may interact with drugs metabolised by these CYP450s, which could guide the clinical application.

Rea regulates microglial polarization and attenuates neuronal apoptosis via inhibition of the NF-κB and MAPK signalings for spinal cord injury repair.

Inflammation and neuronal apoptosis aggravate the secondary damage after spinal cord injury (SCI). Rehmannioside A (Rea) is a bioactive herbal extract isolated from Rehmanniae radix with low toxicity and neuroprotection effects. Rea treatment inhibited the release of pro-inflammatory mediators from microglial cells, and promoted M2 polarization in vitro, which in turn protected the co-cultured neurons from apoptosis via suppression of the NF-κB and MAPK signalling pathways. Furthermore, daily intraperitoneal injections of 80 mg/kg Rea into a rat model of SCI significantly improved the behavioural and histological indices, promoted M2 microglial polarization, alleviated neuronal apoptosis, and increased motor function recovery. Therefore, Rea is a promising therapeutic option for SCI and should be clinically explored.

Mechanisms of Rehmannioside A Against Systemic Lupus Erythematosus Based on Network Pharmacology, Molecular Docking and Molecular Dynamics Simulation.

The effect of rehmannioside A (ReA) on systemic lupus erythematosus (SLE) is not clear and needs further study. In this study, SLE-related targets were obtained from the DisGeNet and GeneCards databases, while ReA-related targets were obtained from the SwissTarget and SuperPred databases. A protein-protein interaction network of intersected targets was constructed using the STRING platform. After selecting the intersected targets, GO and KEGG enrichment analyses were performed via the R package "clusterProfiler". The relationships between ReA and various core targets were assessed via molecular docking, and molecular dynamics simulation was conducted for optimal core protein-compound complexes obtained by molecular docking. The top five targets in the ranking of degree value were HSP90AA1, HIF1A, PIK3CA, MTOR, and TLR4. Significant biological processes mainly included response to oxidative stress and response to reactive oxygen species. The potential pathways of ReA in the treatment of SLE mainly focused on the PI3K-Akt signaling pathway, neutrophil extracellular trap formation, and Apoptosis. Molecular docking showed that ReA had the highest binding affinity for mTOR, suggesting that mTOR is a key target of ReA against SLE. Molecular dynamics simulations revealed good binding abilities between ReA and mTOR. In conclusion, ReA exerts its effects on SLE through multiple targets and pathways, with mTOR being a key target of ReA against SLE.

Rehmannioside A Inhibits TRAF6/MAPK Pathway and Improves Psoriasis by Interfering with the Interaction of HaCaT Cells with IL-17A.

Objective: As a common chronic inflammatory skin disease, psoriasis seriously affects the physical health and psychological well-being of patients. Various clinical treatments for psoriasis have their own drawbacks, so it is important to find effective and safe drugs. Rehmannioside A (ReA) has anti-inflammatory properties and is the main active ingredient in Fuzhengzhiyanghefuzhiyang decoction (FZHFZY), an herbal compound for the treatment of psoriasis. But no studies have been conducted to determine whether ReA alone can treat psoriasis. Therefore, this study was designed to investigate the effect of ReA in the treatment of psoriasis and its potential mechanism of action. Methods: HaCaT cells were treated with ReA and IL-17A alone for 24 h and 48 h, and the most effective concentrations of ReA and interleukin (IL)-17A were found at 25 µg/mL and 100 ng/mL, respectively. A psoriasis cell model was constructed by stimulating HaCaT cells with IL-17A, followed by intervention with ReA. Cell viability and cell cycle distribution were measured by MTT assay and flow cytometry. The expression levels of keratin family members and chemokines were detected by real-time quantitative PCR (RT-qPCR), the levels of pro-inflammatory cytokines by enzyme-linked immunosorbent assay (ELISA), and key proteins of TRAF6/MAPK signaling pathway by Western blot. Results: ReA weaken cell viability, down-regulate the expression of keratin family members (KRT6 and KRT17), restore cell cycle distribution to normal distribution, inhibit the release of pro-inflammatory cytokines (IL-6, IL-8 and IL-1ß) and lower the expression of chemokines (S100A7, S100A9 and CXCL2) by interfering with the interaction between HaCaT cells and IL-17A. Thus, it exerts an anti-psoriatic effect by reducing the inflammatory response and inhibiting abnormal proliferation of HaCaT cells. Mechanistically, ReA inhibited the TRAF6/MAPK signaling pathway activated by IL-17A stimulation in HaCaT cells. Conclusion: ReA has in vitro anti-psoriatic effects and may be a new therapeutic agent for psoriasis.

Potential benefits of Rehmanniae Radix after ancient rice-steaming process in promotion of antioxidant activity in rats' health.

Rice steam processed product of Rehmanniae Radix (RSRR), one of the processed products of Rehmanniae Radix (RR), is popular as an herbal medicine and food. However, the health-promoting effects and mechanisms of RSRR are still unclear. In this study, 10-week-old Sprague-Dawley female rats were treated with different processed products of RR. No organ coefficient differences were observed between RSRR and the control group, indicating that RSRR did not cause damage to the rats. Compared with other RR products, superoxide dismutase, glutathione, and catalase levels were significantly higher and malondialdehyde levels were significantly lower in the RSRR group, indicating that RSRR exerted a better antioxidant effect. Gene expression analysis showed that hemoglobin genes (Hba-a1, Hba-a2, Hbb-bs, Hbb, Hbq1b, Hbb-b1, and LOC103694857) may be potential biomarkers to evaluate the antioxidant effect of RSRR. Antioxidation-related signaling pathways in GO annotation, including cellular oxidant detoxification, hydrogen peroxide metabolic process, hemoglobin complex, and oxygen binding signaling pathways were significantly enriched, indicating these pathways may represent the antioxidant mechanism of RSRR. To explore the main active compounds primarily responsible for the antioxidant activity of RSRR, UPLC-Q-TOF-MS was used and six components (catalpol, rehmannioside A, rehmannioside D, melittoside, ajugol, and verbascoside) were identified in rat serum. Catalpol and rehmannioside A were predicted to be the major active components by network pharmacology. These results suggested that RSRR exhibits antioxidant activity and has health-promoting properties. This study provides a scientific basis for the antioxidant mechanism and clinical use of RSRR.

Antidepressant-like effects of Rehmannioside A on rats induced by chronic unpredictable mild stress through inhibition of endoplasmic reticulum stress and apoptosis of hippocampus.

Depression is one of the most prevalent psychiatric mood diseases worldwide, whose therapy is in urgent need of development. Although the neuroprotective effects of Rehmannioside A (Rea) have been demonstrated, its anti-depressive effect remains unclear. Here, a depression model was induced with chronic unpredictable mild stress (CUMS) in rats. The behavioral trails, including sucrose preference test, forced swim test and open field test were used to determine the success of the CUMS-induced model. The effect of Rea on the neuronal protection, apoptosis and endoplasmic reticulum stress (ERS) was evaluated by HE, NISSL, IF and TUNEL staining, and western blot assays. Mechanically, the MAPK signaling pathway-related proteins expressions were examined by western blot. The results showed that CUMS stimulation evoked a prominent reduction of rat body weight, sucrose preference, and numbers of crossing, rearing and grooming with the enhanced immobility times. Besides, CUMS exposure induced the nuclear shrinkage and damage, as well as the decreased ISSL+ numbers. Moreover, CUMS stimulation increased the relative protein expressions of Bax and Cleaved caspase-3 and the percent of TUNEL positive cells, and decreased the relative protein expressions of Bcl-2. Furthermore, CUMS exposure also increased the relative protein expression of GRP-78, XBP-1, ATF-6, ATF-4 and CHOP. However, the CUMS-induced changes of all these indicators were reversed with Rea introduction in a dose-dependent fashion. Mechanically, Rea supply observably antagonized the CUMS-induced the relative protein levels of p-p38/p-38, p-ERK1/2/ERK1/2 and p-JNK/JNK. Therefore, Rea attenuated depression through suppressing ERS and apoptosis in hippocampus of CUMS-induced rats involved in MAPK signaling.

Rehmannioside A improves cognitive impairment and alleviates ferroptosis via activating PI3K/AKT/Nrf2 and SLC7A11/GPX4 signaling pathway after ischemia.

ETHNOPHARMACOLOGICAL RELEVANCE: Rehmannioside A is derived from Rehmannia glutinosa Libosch, which is widely used as an important ingredient in diverse traditional Chinese medicines to treat diseases caused by "kidney deficiency" such as cerebral arteriosclerosis, aging-related stroke and dementia in China. Recent studies have proved that Rehmannia glutinosa Libosch and Rehmannioside A can improve memory capability and recover nerve damage. AIM OF THE STUDY: To investigate the effect of Rehmannioside A on cognitive impairment after ischemia in rats and SH-SY5Y cells, and further evaluate the anti-oxidative and anti-ferroptosis mechanisms. MATERIALS AND METHODS: Differentially expressed proteins (DEPs) in patients after cerebral ischemic stroke were revealed by a RayBio protein array. Cognitive impairment model was established by middle cerebral artery occlusion and reperfusion (MCAO) 14 days in rats. Rehmannioside A was administered intraperitoneally injection at dose of 80 mg/kg. The SH-SY5Y cells were exposed to H2O2 for 24 h and treated with Rehmannioside A (80 µM) for 24 h. The neuroprotecion of Rehmannioside A were evaluated by infarct volume (TTC), neurological defects (Garcia score) and learning memory (Morris water maze test) in vivo, and cell viability (CCK-8 or LDH) in vitro. Superoxide dismutase (SOD), malondialdehyde (MDA) and myeloperoxidase (MPO) activity of rats, glutathione (GSH), oxidized glutathione (GSSG) and nicotinamide adenine dinucleotide phosphate (NADPH) of cells were detected by biochemical assay. Intracellular reactive oxygen species (ROS) were measured by DCFH-DA assay. Myeloperoxidase (MPO), PI3 kinase (PI3K), p-PI3K, Akt, p-Akt, heme oxygenase-1 (HO-1), nuclear factor-E2-related factor 2 (Nrf2), SLC7A11, glutathione peroxidase 4 (GPX4) of the cerebral cortex in rats or SH-SY5Y cells were examined by western blotting. RESULTS: Compared with model group, the cognitive impairment and neurological deficits of Rehmannioside A group were significantly improved, and the cerebral infarction was reduced in MCAO rats. Moreover, the cell viability obviously increased and the H2O2-induced toxicity was reduced in Rehmannioside A group. Further research indicated that the expression of p-PI3K, p-Akt, nuclear Nrf2, HO-1 and SLC7A11 in Rehmannioside A group was significantly higher than model group. CONCLUSION: Rehmannioside A has neuroprotection effect and improves cognitive impairment after cerebral ischemia by inhibiting ferroptosis and activating PI3K/AKT/Nrf2 and SLC7A11/GPX4 signaling pathway. These findings provide valuable insight into the pathogenesis and therapeutic target of ischemic stroke.

Rehmannioside A attenuates cognitive deficits in rats with vascular dementia (VD) through suppressing oxidative stress, inflammation and apoptosis.

Vascular dementia (VD) is a degenerative cerebrovascular disorder, leading to progressive decline of cognitive abilities and memory. Rehmannioside A (ReA) is isolated from Rehmanniae Radix, which exhibits protective role against various diseases. The present study was performed to calculate the possible neuroprotective effects of ReA on VD. Here, the morris water maze (MWM) test and electrophysiological recordings indicated that ReA reduced cognitive deficits. Additionally, through hematoxylin and eosin (H&E) and Nissl staining, ReA attenuated the histological alterations of hippocampus in rats with VD. ReA group significantly reduced oxidative stress, inflammatory response and apoptosis in the hippocampus of rats with VD, which was linked to the activation of nuclear erythroid related factor-2 (Nrf2), while the inactivation of nuclear factor-κB (NF-κB) and Caspase-3. Further, the anti-oxidative, anti-inflammatory and anti-apoptosis abilities of ReA were confirmed in cells stimulated by hydrogen peroxide. Overall, the results above demonstrated the protective effects of ReA against cognitive deficits and indicated the potential value of ReA in the therapy of VD in future.

Rehmanniae Radix in osteoporosis: A review of traditional Chinese medicinal uses, phytochemistry, pharmacokinetics and pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Emerging clinical usage and pharmacological effects have been achieved in using Rehmanniae Radix either singly or in combination with other herbs to treat skeletal diseases in traditional Chinese medicine (TCM) in the recent years. This study is aimed to provide a comprehensive review about the historical TCM interpretation of the action of Rehmanniae Radix in osteoporosis, its usage in clinical trials and osteoporotic models, its main phytochemical constituents, and its pharmacokinetics. MATERIALS AND METHODS: Several databases included PubMed, China Knowledge Resource Integrated Database, China Science and Technology Journal Database, National Science and Technology Library and the Web of Science Database were consulted to locate the publications pertaining to Rehmanniae Radix. The initial inquiry was conducted for the presence of the following terms combinations in the abstracts: Rehmanniae Radix, Dihuang, phytochemistry, pharmacokinetics, osteoporosis, bone, osteoclast and osteoblast. About 330 research papers and reviews were consulted. RESULTS: In TCM, Rehmanniae Radix exerts the anti-osteoporotic effect via regulating the functions of kidney and liver as well as improving blood circulation. 107 clinical trials are identified that used Rehmanniae Radix in combination with other herbs to treat post-menopausal, senile and secondary osteoporosis. Most of the clinical trials are characterized by high efficacy and no obvious adverse effects. However, the efficacies of these clinical trials are limited because of small patient sample size, short treatment duration and poor clinical design. In addition, TCM herbs under the clinical study are not clear because of a lack of standardization and authentication. The pharmacokinetics data demonstrate that the ingredients of Rehmanniae Radix are widely distributed after administration, and that catalpol and ajugol as well as acetoside are supposed to be the active constituents. More than 140 individual compounds have been currently isolated from this plant and reported to show pleiotropic effects on various diseases. Rehmanniae Radix displays bone protecting features in the osteoporosis models via the delicate balance between osteoclastogenesis and osteoblastogenesis through single herb extracts and its isolated compounds. CONCLUSIONS: The successful inclusion of Rehmanniae Radix in clinical trials and preclinical studies for the management of osteoporosis has attracted rising attentions for identifying potential anti-osteoporotic candidates from this plant and clinical existing TCM formulas, which will further speed up anti-osteoporosis drug discovery processes. Properly designed and well controlled prospective studies are still needed to further demonstrate bone protective actions and safe use of this herb and its ingredients.

Jia-Jian-Di-Huang-Yin-Zi decoction reduces apoptosis induced by both mitochondrial and endoplasmic reticulum caspase12 pathways in the mouse model of Parkinson's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: As a classical prescription of traditional Chinese medicine (TCM), Jia-Jian-Di-Huang-Yin-Zi decoction (JJDHYZ) has been used to treat the symptoms of neurological disorders with a long history. AIM OF THE STUDY: To evaluate the effects and possible mechanisms of JJDHYZ on a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced subacute mouse model of Parkinson's disease. MATERIALS AND METHODS: Adult male C57BL/6 mice were randomly divided into five groups: control, MPTP, JJDHYZ low dosage (JJDHYZ-L, 8.5g/kg/day), medium dosage (JJDHYZ-M, 17g/kg/day) and high dosage (JJDHYZ-H, 34g/kg/day). Behavioral tests, immunohistochemistry, immunofluorescence, and high-performance liquid chromatography (HPLC) were conducted to evaluate the neuroprotective effects of JJDHYZ. The mechanism was further explored using TdT-mediated dUTP nick end labeling staining and transmission electron microscopy. The protein expression of Bax, Bcl-2, cytochrome c, full-length caspase9, cleaved caspase9, cleaved caspase3, caspase12 and C/EBP homologous protein was assessed. The toxicity on hepatocytes and renal cells was detected using the enzyme-linked immunosorbent assay kits. RESULTS: JJDHYZ-H restored the behavior performance impaired by MPTP, and reduced the loss of tyrosine hydroxylase. Additionally, it blocked the apoptosis, activated cleaved caspase3 and protected the ultrastructural integrity of mitochondria by regulating the expression of proteins in both mitochondrial and endoplasmic reticulum (ER) caspase12 pathways. CONCLUSIONS: JJDHYZ-H showed behavior recovery and dopamine neuron protection by inhibiting the apoptotic activities associated with mitochondrial and ER caspase12 pathways.