Tibetan medicine Ruyi Zhenbao Pill ameliorates neuropathic pain by inhibiting the CXCL10-CXCR3 pathway in spinal cord of spinal nerve ligation model.

ETHNOPHARMACOLOGICAL RELEVANCE: Ruyi Zhenbao Pill (RYZBP) is a traditional Tibetan medicine that has been used for over 300 years in China to treat neurological diseases, specifically neuropathic pain (NP). However, its characteristics and mechanism of action in treating NP remains unclear. AIM OF THE STUDY: Based on animal experiments and transcriptomics to evaluate the characteristics and mechanism of RYZBP in treating NP. METHODS: Mice were divided into six groups using random assignment: sham-operation group, spinal nerve ligation (SNL) group, RYZBP low (0.65 g kg-1), medium (1.30 g kg-1), high (2.60 g kg-1) doses groups, and positive drug pregabalin (PGB, 0.05 g kg-1) group. Mice received intragastrical administered for 14 consecutive days. SNL and intrathecal injection models were employed. The analgesic effects were assessed using the Von Frey test, Acetone test, and Hot Plate test. L5 spinal dorsal horns were collected for transcriptomics on day 15. The potential signaling pathways and Hub genes of RYZBP to ameliorate NP were obtained through transcriptomics and network pharmacology. Molecular docking was utilized to evaluate the binding ability of candidate active ingredients with the Hub genes. Finally, western blot (WB) and immunofluorescence (IF) were used to validate the predicted targets. RESULTS: RYZBP demonstrated a dose-dependent alleviation of mechanical allodynia, cold and heat stimulus-induced pain in SNL mice. Transcriptomics analysis identified 24 differentially expressed genes, and pathway enrichment analysis revealed that the CXCL10-CXCR3 signal axis may be the primary biological pathway through which RYZBP relieve NP. Molecular docking test indicated that the active ingredient in RYZBP exhibit a strong affinity for the target protein CXCL10. WB and IF tests showed that RYZBP can significantly inhibit CXCL10 and CXCR3 and its downstream molecules expression in the spinal dorsal horn of SNL mice. Additionally, intrathecal injection of rmCXCL10 worsened pain hypersensitivity, while RYZBP was able to suppress the pain hypersensitivity response induced by rmCXCL10 and reduce the expression levels of CXCL10 and CXCR3 and its downstream molecules. CONCLUSION: RYZBP had a significant analgesic effect on NP model, and this effect may be related to inhibiting the CXCL10-CXCR3 pathway in the spinal dorsal horn.

[Mechanism of Mongolian drug Naru-3 in initiation of neuroinflammation of neuropathic pain from MMP9/IL-1ß signaling pathway].

Neuropathic pain(NP) has similar phenotypes but different sequential neuroinflammatory mechanisms in the pathological process. It is of great significance to inhibit the initiation of neuroinflammation, which has become a new direction of NP treatment and drug development in recent years. Mongolian drug Naru-3 is clinically effective in the treatment of trigeminal neuralgia, sciatica, and other NPs in a short time, but its pharmacodynamic characteristics and mechanism of analgesia are still unclear. In this study, a spinal nerve ligation(SNL) model simulating clinical peripheral nerve injury was established and the efficacy and mechanism of Naru-3 in the treatment of NPs was discussed by means of behavioral detection, side effect evaluation, network analysis, and experimental verification. Pharmacodynamic results showed that Naru-3 increased the basic pain sensitivity threshold(mechanical hyperalgesia and thermal radiation hyperalgesia) in the initiation of SNL in animals and relieved spontaneous pain, however, there was no significant effect on the basic pain sensitivity threshold and motor coordination function of normal animals under physiological and pathological conditions. Meanwhile, the results of primary screening of target tissues showed that Naru-3 inhibited the second phase of injury-induced nociceptive response of formalin test in mice and reduced the expression of inflammatory factors in the spinal cord. Network analysis discovered that Naru-3 had synergy in the treatment of NP, and its mechanism was associated with core targets such as matrix metalloproteinase-9(MMP9) and interleukin-1ß(IL-1ß). The experiment further took the dorsal root ganglion(DRG) and the stage of patho-logical spinal cord as the research objects, focusing on the core targets of inducing microglial neuroinflammation. By means of Western blot, immunofluorescence, agonists, antagonists, behavior, etc., the mechanism of Naru-3 in exerting NP analgesia may be related to the negative regulation of the MMP9/IL-1ß signaling pathway-mediated microglia p38/IL-1ß inflammatory loop in the activation phase. The relevant research enriches the biological connotation of Naru-3 in the treatment of NP and provides references for clinical rational drug use.

[Baimai Ointment relieves chronic pain induced by chronic compression of dorsal root ganglion in rats by regulating neuroactive ligand-receptor interaction and HIF-1 signaling pathway].

The Baimai Ointment with the effect of relaxing sinew and activating collaterals demonstrates a definite effect on Baimai disease with pain, spasm, stiffness and other symptoms, while the pharmacodynamic characteristics and mechanism of this agent remain unclear. In this study, a rat model of chronic compression of L4 dorsal root ganglion(CCD) was established by lumbar disc herniation, and the efficacy and mechanism of Baimai Ointment in the treatment of CCD were preliminarily explored by behavioral tests, side effect evaluation, network analysis, antagonist and molecular biology verification. The pharmacodynamic experiment indicated that Baimai Ointment significantly improved the pain thresholds(mechanical pain, thermal pain, and cold pain) and gait behavior of CCD model rats without causing tolerance or obvious toxic and side effects. Baimai Ointment inhibited the second-phase nociceptive response of mice in the formalin test, increased the hot plate threshold of normal mice, and down-regulated the expression of inflammatory cytokines in the spinal cord. Network analysis showed that Baimai Ointment had synergistic effect in the treatment of CCD and was related to descending inhibition/facilitation system and neuroinflammation. Furthermore, behavioral tests, Western blot, and immunofluorescence assay revealed that the pain-relieving effect of Baimai Ointment on CCD may be related to the regulation of the interaction between neuroactive ligand and receptors(neuroligands) such as CHRNA7, ADRA2A, and ADRB2, and the down-regulation of the expression of NOS2/pERK/PI3K, the core regulatory element of HIF-1 signaling pathway in spinal microglia. The findings preliminarily reveal the mechanism of relaxing sinew and activating collaterals of Baimai Ointment in the treatment of Baimai disease, providing a reference for the rational drug use and further research of this agent.

Investigation of the mechanism of the anti-cancer effects of Astragalus propinquus Schischkin and Pinellia pedatisecta Schott (A&P) on melanoma via network pharmacology and experimental verification.

Melanoma is a commonly malignant cutaneous tumor in China. Astragalus propinquus Schischkin and Pinellia pedatisecta Schott (A&P) have been clinically used as adjunctive drugs in the treatment of malignant melanoma. However, the effect and mechanism of A&P on melanoma have yet to be explored. The current investigation seeks to characterize the active components of A&P and their potential roles in treating malignant melanoma using network pharmacology and in vitro and in vivo experiments. We first used the traditional Chinese medicine systems pharmacology (TCMSP) database and high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) to identify a total of 13 effective compounds within A&P. 70 common genes were obtained by matching 487 potential genes of A&P with 464 melanoma-related genes, and then we built up protein-protein interaction (PPI) network of these 70 genes, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. The results revealed that A&P might influence the pathobiology of melanoma through the PI3K/Akt pathway. Molecular docking also confirmed that higher content of ingredients in A&P, including hederagenin, quercetin, beta-sitosterol and stigmasterol, had a strong binding activity (affinity < -5 kcal/mol) with the core targets AKT1, MAPK3 and ESR1. Furthermore, we confirmed A&P could inhibit melanoma cells proliferation and induce cells apoptosis through suppressing the PI3K/Akt signaling pathway by in vitro and in vivo xenograft model experiments. These findings indicate that A&P may function as a useful therapy for melanoma through the PI3K/Akt pathway.

Protective effect of apigenin on ischemia/reperfusion injury of the isolated rat heart.

Apigenin (Api), a mainly bioactive component of Apium graveolens L. var. dulce DC. (a traditional Chinese medicinal herb), possesses a wide range of biological activities, including antioxidant effects. It also has been shown to associate with lower prevalence of cardiovascular diseases, but its mechanisms of action remain unclear. The aim of the present study is to investigate the role of Api in isolated rat heart model of ischemia/reperfusion (I/R). Langendorff-perfused isolated rat hearts were used in our study. Api was added to the perfusate before ischemia and during reperfusion in the isolated pulsed rat heart exposed to 30-min ischemia followed by 50-min reperfusion. The treatment with Api conferred a cardioprotective effect, and the treated hearts demonstrated an improved ischemic cardiac functional recovery, a decreased myocardial infarct size, a reduced activities of creatine kinase isoenzyme and lactate dehydrogenase in the coronary flow, a reduced number of apoptotic cardiomyocytes, a reduced activity of caspase-3, up-regulation of the anti-apoptotic protein Bcl-2 and down-regulation of the pro-apoptotic protein Bax. In addition, Api inhibited the phosphorylation of p38 MAPKS during I/R. In conclusion, these observations provide preliminary evidence that Api can protect cardiomyocytes from I-/R-induced injury, at least partially, through the inhibition of p38 MAPKS signaling pathway.

[Application of chromatograph-digitized fingerprint spectrum in identification of indigenous medicinal materials Radix et Rhizoma Rhei from Gansu province].

OBJECTIVE: To establish HPLC-Digitized fingerprint spectrum (HPLC-DFPS) of Radix et Rhizoma Rhei from Gansu province. METHODS: HPLC method was applied. RESULTS AND CONCLUSION: A group of characteristic peaks suggested that HPLC-DFPS can be used for identification of Radix et Rhizoma Rhei from Gansu province.

[Studies on fingerprint spectrum of Ephedra sinica from Gansu province].

OBJECTIVE: To set up the fingerprint spectrum of Ephedra sinica from Gansu province and its extract for controlling the quality. METHODS: HPLC with UV detector was used to analyze the patterns of Ephedra sinica from Gansu province. RESULTS: The fingerprint spectrum of Ephedra sinica from Gansu province was set up. Furthermore, the fingerprint spectrum showed an excellent correlation, and all the parameters of the main co-peaks met the requirements for State Drug Administration's technical regulations on the herb. CONCLUSION: The studies are useful for the quality control of Ephedra sinica from Gansu province.