Semen Strychni Pulveratum and vomicine alleviate neuroinflammation in amyotrophic lateral sclerosis through cGAS-STING-TBK1 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Amyotrophic lateral sclerosis (ALS) is a fetal neuromuscular disorder characterized by the gradual deterioration of motor neurons. Semen Strychni pulveratum (SSP), a processed version of Semen Strychni (SS) powder, is widely used to treat ALS in China. Vomicine is one of the most primary components of SS. However, their pharmacological effects and mechanisms for ALS remain elusive. AIM OF THE STUDY: This study aimed to evaluate the neuroprotective and anti-neuroinflammatory effects of SSP and vomicine, as well as to explore their protective roles in ALS and the underlying mechanisms. MATERIALS AND METHODS: In vivo, 8-week-old hSOD1-WT mice and hSOD1-G93A mice were orally administered different concentrations of SSP (SSP-L = 5.46 mg/ml, SSP-M = 10.92 mg/ml or SSP-H = 16.38 mg/ml) once every other day for 8 weeks. A series of experiments, including body weight measurement, footprint tests, Hematoxylin & Eosin staining, and Nissl staining, were performed to evaluate the preventive effect of SSP. Immunofluorescence staining, western blotting, and RT-qPCR were subsequently performed to evaluate activation of the cGAS-STING-TBK1 pathway in the spinal cord. In vitro, hSOD1G93A NSC-34 cells were treated with vomicine to further explore the pharmacological mechanism of vomicine in the treatment of ALS via the cGAS-STING-TBK1 pathway. RESULTS: SSP improved motor function, body weight loss, gastrocnemius muscle atrophy, and motor neuron loss in the spine and cortex of hSOD1-G93A mice. Furthermore, the cGAS-STING-TBK1 pathway was activated in the spinal cord of hSOD1-G93A mice, with activation predominantly observed in neurons and microglia. However, the levels of cGAS, STING, and pTBK1 proteins and cGAS, IRF3, IL-6, and IL-1ß mRNA were reversed following intervention with SSP. Vomicine not only downregulated the levels of cGAS, TBK1, IL-6 and IFN-ß mRNA, but also the levels of cGAS and STING protein in hSOD1G93A NSC-34 cells. CONCLUSION: This study demonstrated that SSP and vomicine exert neuroprotective and anti-neuroinflammatory effects in the treatment of ALS. SSP and vomicine may reduce neuroinflammation by regulating the cGAS-STING-TBK1 pathway, and could thereby play a role in ALS treatment.

Profiling of antioxidant properties and identification of potential analgesic inhibitory activities of Allophylus villosus and Mycetia sinensis employing in vivo, in vitro, and computational techniques.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional use of plants for medicinal purposes, called phytomedicine, has been known to provide relief from pain. In Bangladesh, the Chakma indigenous community has been using Allophylus villosus and Mycetia sinensis to treat various types of pain and inflammation. AIM OF THE STUDY: The object of this research is to evaluate the effectiveness of these plants in relieving pain and their antioxidant properties using various approaches such as in vitro, in vivo, and computational techniques. Additionally, the investigation will also analyse the phytochemicals present in these plants. MATERIALS AND METHODS: We conducted in vivo analgesic experiment on Swiss albino mice and in-silico inhibitory activities on COX-2 & 15-LOX-2 enzymes. Assessment of DPPH, Anti Radical Activities (ARA), FRAP, H2O2 Free Radical Scavenging, Reducing the power of both plants performed significant % inhibition with tolerable IC50. Qualitative screening of functional groups of phytochemicals was précised by FTIR and GC-MS analysis demonstrated phytochemical investigations. RESULTS: The ethyl acetate (EtOAc) fractioned Mycetia sinensis extract as well as the ethanoic extract and all fractioned extracts of Allophylus villosus have reported a significant percentage (%) of writhing inhibition (p < 0.05) with the concentrated doses 250 mg as well as 500 mg among the Swiss albino mice for writhing observation of analgesic effect. In the silico observation, a molecular-docking investigation has performed according to GC-MS generated 43 phyto-compounds of both plants to screen their binding affinity by targeting COX-2 and 15-LOX-2 enzymes. Consequently, in order to assess and ascertain the effectiveness of the sorted phytocompounds, ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) investigation, DFT (Density-functional theory) by QM (Quantum mechanics), and MDS (Molecular dynamics simulation) were carried out. As the outcome, compounds like 5-(2,4-ditert-butylphenoxy)-5-oxopentanoic acid; 2,4-ditert-butylphenyl 5-hydroxypentanoate; 3,3-diphenyl-5-methyl-3H-pyrazole; 2-O-(6-methylheptan-2-yl) 1-O-octyl benzene-1,2-dicarboxylate and dioctan-3-yl benzene-1,2-dicarboxylate derived from the ethnic plant A. villosus and another ethnic plant M. sinensis extracts enchants magnificent analgesic inhibitions and performed more significant drug like activities with the targeted enzymes. CONCLUSIONS: Phytocompounds from A. villosus & M. sinensis exhibited potential antagonist activity against human 15-lipoxygenase-2 and cyclooxygenase-2 proteins. The effective ester compounds from these plants performed more potential anti-nociceptive activity which could be used as a drug in future.

Cysteine triggered cascade reaction forming coumarin: Visualization of cysteine fluctuation in alcoholic liver disease by a NIR fluorescent probe.

Alcoholic liver disease (ALD) is a chronic toxic liver injury caused by long-term heavy drinking. Due to the increasing incidence, ALD is becoming one of important medical tasks. Many studies have shown that the main mechanism of liver damage caused by large amounts of alcohol may be related to antioxidant stress. As an important antioxidant, cysteine (Cys) is involved in maintaining the normal redox balance and detoxifying metabolic function of the liver, which may be closely related to the pathogenesis of ALD. Therefore, it is necessary to develop a simple non-invasive method for rapid monitoring of Cys in liver. Thus, a near-infrared (NIR) fluorescent probe DCI-Ac-Cys which undergoes Cys triggered cascade reaction to form coumarin fluorophore is developed. Using the DCI-Ac-Cys, decreased Cys was observed in the liver of ALD mice. Importantly, different levels of Cys were monitored in the livers of ALD mice taking silybin and curcumin with the antioxidant effects, indicating the excellent therapeutic effect on ALD. This study provides the important references for the accurate diagnosis of ALD and the pharmacodynamic evaluation of silybin and curcumin in the treatment of ALD, and support new ideas for the pathogenesis of ALD.

20S-Ginsenoside Rh2, the major bioactive saponin in Panax notoginseng flowers, ameliorates cough by inhibition of NaV1.7 and TRPV1 channel currents and downregulation of TRPV1 expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax notoginseng flowers, which are the buds of the traditional Chinese medicinal herb Sanqi, are widely used in China for their cough-ameliorating properties, with demonstrated therapeutic effects in the treatment of both acute and chronic coughs. However, both the antitussive mechanism and active compound basis of P. notoginseng flowers remain poorly understood. AIM OF THE STUDY: We investigated the antitussive effects of P. notoginseng flowers, identified the bioactive constituents responsible for alleviating cough symptoms, and elucidated the underlying pharmacological mechanisms. MATERIALS AND METHODS: We analyzed the major chemical constituents of aqueous extracts of P. notoginseng flowers using liquid chromatography-mass spectrometry and quantitatively analyzed the key component, 20S-ginsenoside Rh2, using high-performance liquid chromatography. Using a cough reflex model in healthy mice and an ovalbumin-induced, highly sensitive guinea pig cough model, we verified the suppressive effects of P. notoginseng flowers and their saponin constituents on coughing. Furthermore, we explored the mechanisms of action of the key ion channels, NaV1.7 and TRPV1, using whole-cell patch-clamp techniques and molecular docking. Finally, the therapeutic mechanisms of P. notoginseng flowers on pathological cough were revealed using hematoxylin and eosin staining, immunohistochemistry, and western blotting. RESULTS: The active components of P. notoginseng flowers were primarily protopanaxadiol-type saponins, among which 20S-ginsenoside Rh2 had the highest content (51.46 mg/g). In the mouse model, P. notoginseng flowers exhibited antitussive effects comparable to those of pentoxyverine citrate. Although its main saponin component, 20S-ginsenoside Rh2, showed slightly weaker effects, it still demonstrated concentration-dependent inhibition of channel activity. The whole-cell patch-clamp technique and virtual molecular docking showed that Rh2 might exert its effects by directly binding to the NaV1.7 and TRPV1 channels. In the guinea pig model, P. notoginseng flowers and their saponin components not only reduced cough frequency and prolonged the latency period before cough onset, but also significantly inhibited tracheal and pulmonary inflammation and the overexpression of TRPV1. CONCLUSIONS: 20S-Ginsenoside Rh2, the major bioactive saponin in P. notoginseng flowers, exhibits potent antitussive effects. The potential mechanism of action of 20S-Ginsenoside Rh2 in the treatment of cough may involve inhibiting NaV1.7 and TRPV1 channel currents through direct binding to core protein active sites and downregulating TRPV1 expression.

Si-Ni-San alleviates intestinal and liver damage in ulcerative colitis mice by regulating cholesterol metabolism.

ETHNOPHARMACOLOGICAL RELEVANCE: Si-Ni-San (SNS), a traditional Chinese medicinal formula derived from Treatise on Febrile Diseases, is considered effective in the treatment of inflammatory bowel diseases based upon thousands of years of clinical practice. However, the bioactive ingredients and underlying mechanisms are still unclear and need further investigation. AIM OF THE STUDY: This study aimed to evaluate the effect, explore the bioactive ingredients and the underlying mechanisms of SNS in ameliorating ulcerative colitis (UC) and associated liver injury in dextran sodium sulphate (DSS)-induced mouse colitis models. MATERIALS AND METHODS: The effect of SNS (1.5, 3, 6 g/kg) on 3% DSS-induced acute murine colitis was evaluated by disease activity index (DAI), colon length, inflammatory cytokines, hematoxylin-eosin (H&E) staining, tight junction proteins expression, ALT, AST, and oxidative stress indicators. HPLC-ESI-IT/TOF MS was used to analyze the chemical components of SNS and the main xenobiotics in the colon of UC mice after oral administration of SNS. Network pharmacological study was then conducted based on the main xenobiotics. Flow cytometry and immunohistochemistry techniques were used to demonstrate the inhibitory effect of SNS on Th17 cells differentiation and the amelioration of Th17/Treg cell imbalance. LC-MS/MS, Real-time quantitative polymerase chain reaction (RT-qPCR), and western blotting techniques were performed to investigate the oxysterol-Liver X receptor (LXRs) signaling activity in colon. Targeted bile acids metabolomics was conducted to reveal the change of the two major pathways of bile acid synthesis in the liver, and the expression of key metabolic enzymes of bile acids synthesis was characterized by RT-qPCR and western blotting techniques. RESULTS: SNS (1.5, 3, 6 g/kg) decreased the DAI scores, protected intestinal mucosa barrier, suppressed the production of pro-inflammatory cytokines, improved hepatic and splenic enlargement and alleviated liver injury in a dose-dependent manner. A total of 22 components were identified in the colon of SNS (6 g/kg) treated colitis mice, and the top 10 components ranked by relative content were regarded as the potential effective chemical components of SNS, and used to conduct network pharmacology research. The efficacy of SNS was mediated by a reduction of Th17 cell differentiation, restoration of Th17/Treg cell homeostasis in the colon and spleen, and the experimental results were consistent with our hypothesis and the biological mechanism predicted by network pharmacology. Mechanistically, SNS regulated the concentration of 25-OHC and 27-OHC by up-regulated CH25H, CYP27A1 protein expression in colon, thus affected the expression and activity of LXR, ultimately impacted Th17 differentiation and Th17/Treg balance. It was also found that SNS repressed the increase of hepatic cholesterol and reversed the shift of BA synthesis to the acidic pathway in UC mice, which decreased the proportion of non-12-OH BAs in total bile acids (TBAs) and further ameliorated colitis and concomitant liver injury. CONCLUSIONS: This study set the stage for considering SNS as a multi-organ benefited anti-colitis prescription based on the significant effect of ameliorating intestinal and liver damage, and revealed that derivatives of cholesterol, namely oxysterols and bile acids, were closely involved in the mechanism of SNS anti-colitis effect.

Perilla frutescens leaf extracts alleviate acute lung injury in mice by inhibiting KAT2A.

ETHNOPHARMACOLOGICAL RELEVANCE: Acute lung injury (ALI) can lead to respiratory failure and even death. KAT2A is a key target to suppress the development of inflammation. A herb, perilla frutescens, is an effective treatment for pulmonary inflammatory diseases with anti-inflammatory effects; however, its mechanism of action remains unclear. AIM OF THE STUDY: The purpose of this study was to investigate the therapeutic effect and underlying mechanism of perilla frutescens leaf extracts (PLE), in the treatment of ALI by focusing on its ability to treat inflammation. MATERIALS AND METHODS: In vivo and in vitro models of ALI induced by LPS. Respiratory function, histopathological changes of lung, and BEAS-2B cells damage were assessed upon PLE. This effect is also tested under conditions of KAT2A over expression and KAT2A silencing. RESULTS: PLE significantly attenuated LPS-induced histopathological changes in the lungs, improved respiratory function, and increased survival rate from LPS stimuation background in mice. PLE remarkably suppressed the phosphorylation of STAT3, AKT, ERK (1/2) and the release of cytokines (IL-6, TNF-α, and IL-1ß) induced by LPS via inhibiting the expression of KAT2A. CONCLUSIONS: PLE has a dose-dependent anti-inflammatory effect by inhibiting KAT2A expression to suppress LPS-induced ALI n mice. Our study expands the clinical indications of the traditional medicine PLE and provide a theoretical basis for clinical use of acute lung injury.

Engeletin attenuates the inflammatory response via inhibiting TLR4-NFκB signaling pathway in Crohn's disease-like colitis.

ETHNOPHARMACOLOGICAL RELEVANCE: Smilax glabra rhizome has a long history been used for clinical purposes in traditional Chinese medicinal for treating various inflammatory conditions. Engeletin1 (ENG) is one of the most abundant bioactive compounds found in Smilax glabra rhizome, with anti-inflammatory, antioxidant, and ulcer-preventing activities. AIM OF THE STUDY: The purpose of this study was to investigate the ability of ENG to alleviate inflammatory symptoms and improve epithelial barrier integrity utilize a 2,4,6-trinitrobenzene sulfonic acid2 (TNBS)-induced murine model in Crohn's disease3 (CD)-like colitis, and to characterize the underlying anti-inflammatory mechanisms of action. MATERIALS AND METHODS: A colitis model was established in BALB/c mice and treated with ENG for 7 days. RAW264.7 macrophages were pre-treated with ENG and lipopolysaccharide4 (LPS) stimulation. The mice's weight and colon length were assessed. qPCR and Western blotting were used to analyze gene expression and TLR4-NFκB pathway. Flow cytometry was used to analyze the polarization states of the macrophages. RESULTS: Treatment with ENG was sufficient to significantly alleviate symptoms of inflammation and colonic epithelial barrier integrity in treated mice. Significant inhibition of TNF-α, IL-1ß, and IL-6 expression was observed following ENG treatment in vivo and in vitro. ENG was also determined to be capable of inhibiting the expression of iNOS and CD86, inhibited M1 macrophage polarization in vitro, as well as the TLR4-NFκB signaling pathway. Molecular docking showed a highly stable binding between ENG and TLR4. CONCLUSION: ENG has been proven to alleviate inflammation and ameliorate the damage of epithelial barrier in CD-like colitis. ENG also suppressed the M1 macrophages polarization and the inhibited inflammatory cytokines. TLR4-NFκB signaling pathway, especially TLR4, may be the target of ENG. These data offer a new insight into the therapeutic mechanisms of ENG.

Wenshen Xiaozheng Tang alleviates fibrosis in endometriosis by regulating differentiation and paracrine signaling of endometrium-derived mesenchymal stem cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Wenshen Xiaozheng Tang (WXT), a traditional Chinese medicine (TCM) decoction, is effective for treating endometriosis. However, the effect of WXT on endometrium-derived mesenchymal stem cells (eMSCs) which play a key role in the fibrogenesis of endometriosis requires further elucidation. AIMS OF THE STUDY: The aim of this study was to clarify the potential mechanism of WXT in improving fibrosis in endometriosis by investigating the regulation of WXT on differentiation and paracrine of eMSCs. MATERIALS AND METHODS: The nude mice with endometriosis were randomly divided into model group, WXT group and mifepristone group. After 21 days of treatment, the lesion volume was calculated. Fibrosis in the lesions was evaluated by Masson staining and expression of fibrotic proteins. The differentiation of eMSCs in vivo was explored using a fate-tracking experiment. To further clarify the regulation of WXT on eMSCs, primary eMSCs from the ectopic lesions of endometriosis patients were isolated and characterized. The effect of WXT on the proliferation and differentiation of ectopic eMSCs was examined. To evaluate the role of WXT on the paracrine activity of ectopic eMSCs, the conditioned medium (CM) from ectopic eMSCs pretreated with WXT was collected and applied to treat ectopic endometrial stromal cells (ESCs), after which the expression of fibrotic proteins in ectopic ESCs was assessed. In addition, transcriptome sequencing was used to investigate the regulatory mechanism of WXT on ectopic eMSCs, and western blot and ELISA were employed to determine the key mediator. RESULTS: WXT impeded the growth of ectopic lesions in nude mice with endometriosis and reduced collagen deposition and the expression of fibrotic proteins fibronectin, collagen I, α-SMA and CTGF in the endometriotic lesions. The fate-tracking experiment showed that WXT prevented human eMSCs from differentiating into myofibroblasts in the nude mice. We successfully isolated eMSCs from the lesions of patients with endometriosis and demonstrated that WXT suppressed proliferation and myofibroblast differentiation of ectopic eMSCs. Moreover, the expression of α-SMA, collagen I, fibronectin and CTGF in ectopic ESCs was significantly down-regulated by the CM of ectopic MSCs pretreated with WXT. Combining the results of RNA sequencing, western blot and ELISA, we found that WXT not only reduced thrombospondin 4 expression in ectopic eMSCs, but also decreased thrombospondin 4 secretion from ectopic eMSCs. Thrombospondin 4 concentration-dependently upregulated the expression of collagen I, fibronectin, α-SMA and CTGF in ectopic ESCs, indicating that thrombospondin 4 was a key mediator of WXT in inhibiting the fibrotic process in endometriosis. CONCLUSION: WXT improved fibrosis in endometriosis by regulating differentiation and paracrine signaling of eMSCs. Thrombospondin 4, whose release from ectopic eMSCs is inhibited by WXT, may be a potential target for the treatment of endometriosis.

Sea buckthorn extract mitigates chronic obstructive pulmonary disease by suppression of ferroptosis via scavenging ROS and blocking p53/MAPK pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Sea buckthorn (Hippophae rhamnoides), a traditional Tibetan medicinal herb, exhibits protective effects against cardiovascular and respiratory diseases. Although Sea buckthorn extract (SBE) has been confirmed to alleviate airway inflammation in mice, its therapeutic effect and underlying mechanism on chronic obstructive pulmonary disease (COPD) requires further clarification. AIM OF THE STUDY: To elucidate the alleviative effect and molecular mechanism of SBE on lipopolysaccharides (LPS)/porcine pancreatic elastase (PPE)-induced COPD by blocking ferroptosis. METHODS: The anti-ferroptotic effects of SBE were evaluated in human BEAS-2B bronchial epithelial cells using CCK8, RT-qPCR, western blotting, and transmission electron microscopy. Transwell was employed to detect chemotaxis of neutrophils. COPD model was induced by intranasally administration of LPS/PPE in mice and measured by alterations of histopathology, inflammation, and ferroptosis. RNA-sequencing, western blotting, antioxidant examination, flow cytometry, DARTS, CETSA, and molecular docking were then used to investigate its anti-ferroptotic mechanisms. RESULTS: In vitro, SBE not only suppressed erastin- or RSL3-induced ferroptosis by suppressing lipid peroxides (LPOs) production and glutathione (GSH) depletion, but also suppressed ferroptosis-induced chemotactic migration of neutrophils via reducing mRNA expression of chemokines. In vivo, SBE ameliorated LPS/PPE-induced COPD phenotypes, and inhibited the generation of LPOs, cytokines, and chemokines. RNA-sequencing showed that p53 pathway and mitogen-activated protein kinases (MAPK) pathway were implicated in SBE-mediated anti-ferroptotic action. SBE repressed erastin- or LPS/PPE-induced overactivation of p53 and MAPK pathway, thereby decreasing expression of diamine acetyltransferase 1 (SAT1) and arachidonate 15-lipoxygenase (ALOX15), and increasing expression of glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11). Mechanistically, erastin-induced elevation of reactive oxygen species (ROS) was reduced by SBE through directly scavenging free radicals, thereby contributing to its inhibition of p53 and MAPK pathways. CETSA, DARTS, and molecular docking further showed that ROS-generating enzyme nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) may be the target of SBE. Overexpression of NOX4 partially impaired the anti-ferroptotic activity of SBE. CONCLUSION: Our results demonstrated that SBE mitigated COPD by suppressing p53 and MAPK pro-ferroptosis pathways via directly scavenging ROS and blocking NOX4. These findings also supported the clinical application of Sea buckthorn in COPD therapy.

Assessment of acute, subacute genetic toxicities and immunomodulatory activity of palm (Trachycarpus fortunei) buds.

ETHNOPHARMACOLOGY RELEVANCE: Palm buds are a natural green resource of the forest, which are not only rich in nutrients but contain a large number of phenolic acids and flavonoids, among other components. It has a variety of biological activities such as antioxidant and uterine smooth muscle stimulation. AIM OF THE STUDY: To evaluate the safety of palm buds for use as a nutraceutical product and food by evaluating the toxicity, subacute toxicity and genotoxicity of the young palm buds. Also studied for its immune-enhancing activity. MATERIALS AND METHODS: Acute toxicity tests were performed in mice using the maximum tolerance method, and the manifestations of toxicity and deaths were recorded after administration of 10,000 mg/mL for 14 consecutive d (days) of observations. To assess subacute toxicity, mice were treated with palm buds (750, 1500, or 3000 mg/mL) daily for 28 days. The teratogenicity of palm buds was assessed by the Ames test, the mouse bone marrow cell micronucleus test, and the mouse spermatozoa malformation test. In addition, we evaluated the immune-enhancing ability of palm buds by the mouse carbon profile test, delayed-type metamorphosis reaction, and serum hemolysin assay. RESULTS: In the acute toxicity study, the Median Lethal Dose (LD50) was greater than 10,000 mg/kg bw in both male and female rats. There were also no deaths or serious toxicities in the subacute study. The no-observed-adverse-effect level (NOAEL) was 3000 mg/kg bw. However, the mice's food intake decreased after one week. The medium and high dose groups had a reducing effect on body weight in mice of both sexes. In addition, the changes in organ coefficients of the liver, kidney and stomach in male mice were significantly higher in the high-dose group (3.23 ± 0.35, 0.75 ± 0.05, 0.57 ± 0.05 g) than in the control group (2.94 ± 0.18, 0.58 ± 0.05, 0.50 ± 0.02 g). Hematological analyses showed that all the indices of the rats in each palm sprout dose group were within the normal range. The results of blood biochemical indicators showed that there was a significant reduction in TP in the blood of male mice in the high-dose group (44.6 ± 7.8 g/L) compared to the control group (58.3 ± 15.1 g/L). In histopathological analysis, none of the significant histopathological changes were observed. The results of the immunological experiment in mice showed that the liver coefficient and thymus coefficient of the high-dose group (8400 mg/kg) were significantly lower than the control group. There was no remarkable difference in auricle swelling between each dose palm bud group (1400, 2800, or 8400 mg/kg) and the control group. The anti-volume number of the high-dose group was significantly increased. CONCLUSION: Palm buds have non-toxic effects in vivo and have little effect on non-specific and cellular immunity in the test mice within the dose range of this experiment. The immunoenhancement in mice is mainly achieved through humoral immunity. In conclusion, our results suggest that palm buds are safe for use as healthcare products and food.

Dachengqi decoction dispensing granule ameliorates LPS-induced acute lung injury by inhibiting PANoptosis in vivo and in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Acute lung injury (ALI) is a serious health-threatening syndrome of intense inflammatory response in the lungs, with progression leading to acute respiratory distress syndrome (ARDS). Dachengqi decoction dispensing granule (DDG) has a pulmonary protective role, but its potential modulatory mechanism to alleviate ALI needs further excavation. AIM OF THE STUDY: This study aims to investigate the effect and potential mechanism of DDG on lipopolysaccharide (LPS)-induced ALI models in vivo and in vitro. MATERIALS AND METHODS: LPS-treated Balb/c mice and BEAS-2B cells were used to construct in vivo and in vitro ALI models, respectively. Hematoxylin-eosin (HE), Wet weight/Dry weight (W/D) calculation of lung tissue, and total protein and Lactic dehydrogenase (LDH) assays in BALF were performed to assess the extent of lung tissue injury and pulmonary edema. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-1ß), and interleukin-18 (IL-18) in BALF, serum, and cell supernatant. The qRT-PCR was used to detect inflammatory factors, Z-DNA binding protein 1 (ZBP1), and receptor-interacting protein kinase 1 (RIPK1) expression in lung tissues and BEAS-2B cells. Double immunofluorescence staining and co-immunoprecipitation were used to detect the relative expression and co-localization of ZBP1 and RIPK1. The effects of LPS and DDG on BEAS-2B cell activity were detected by Cell Counting Kit-8 (CCK-8). Western blot (WB) was performed to analyze the expression of PANoptosis-related proteins in lung tissues and BEAS-2B cells. RESULTS: In vivo, DDG pretreatment could dose-dependently improve the pathological changes of lung tissue in ALI mice, and reduce the W/D ratio of lung, total protein concentration, and LDH content in BALF. In vitro, DDG reversed the inhibitory effect of LPS on BEAS-2B cell viability. Meanwhile, DDG significantly reduced the levels of inflammatory factors in vitro and in vivo. In addition, DDG could inhibit the expression levels of PANoptosis-related proteins, especially the upstream key regulatory molecules ZBP1 and RIPK1. CONCLUSION: DDG could inhibit excessive inflammation and PANoptosis to alleviate LPS-induced ALI, thus possessing good anti-inflammatory and lung-protective effects. This study establishes a theoretical basis for the further development of DDG and provides a new prospect for ALI treatment by targeting PANoptosis.

Ameliorative potential of ethanol extract of Calyptrochilum emarginatum leaves on scopolamine-induced amnesia in male swiss mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Calyptrochilum emarginatum (Afzel. Ex Sw.) Schltr. (Orchidaceae) is a traditional medicinal plant known for its antimicrobial properties and efficacy in managing convulsive fever and menstrual disorders and addressing conditions such as malaria, tuberculosis, and cough. AIM OF THE STUDY: The study aims to examine the memory-enhancing and neuroprotective properties of ethanol extract of Calyptrochilum emarginatum leaves (EECEL) in scopolamine-induced amnesia mice model. MATERIALS AND METHODS: Forty-two male mice were divided into six groups (n = 7). Group 1 served as control, administered distilled water (10 mL/kg, p. o), group 2 received scopolamine only (3 mg/kg, i. p.), groups 3 to 6 received pretreatments of EECEL (50, 100, and 200 mg/kg, p. o.) and donepezil (1 mg/kg, p. o.) 30 min before scopolamine (3 mg/kg), for seven days. Following treatments, behavioral (learning and memory) assessments were carried out, while biochemical (acetylcholinesterase activity, oxidative stress markers, inflammatory cytokines markers) and histological evaluations were done after euthanasia. RESULTS: Scopolamine significantly impaired spatial, long term and recognition memory. Nevertheless, administration of EECEL (50, 100, and 200 mg/kg orally) enhanced memory function in mice, as observed in the Y maze [F (5, 30) = 20.23, p < 0.0001], Morris water maze [F (10, 90) = 3.105, p = 0.0019; [F (5, 30) = 21.13, p < 0.0001]], and novel object recognition tasks [F (5, 30) = 37.22, p < 0.0001)]. Scopolamine-treated mice exhibited significant dysfunction in the cholinergic system, as evidenced by elevated AChE activity [0.099 ± 0.005 vs. 0.063 ± 0.004 mol/min/g] with an elevation in oxidative stress. On the other hand, administration of EECEL counteracted these consequences by reducing AChE activity, mitigating oxidative damage, reducing pro-inflammatory cytokines, and preventing degeneration of neurons. CONCLUSION: The results demonstrated that EECEL effectively mitigates scopolamine-induced memory impairment via an oxido-inflammatory mechanism and modulation of the central cholinergic system.

Novel detoxifier of spironolactone against triptolide-induced hepatotoxicity through inhibition of RPB1 degradation.

ETHNOPHARMACOLOGICAL RELEVANCE: Triptolide is a major bioactive and toxic ingredient isolated from the traditional Chinese herb Tripterygium wilfordii (T. wilfordii) Hook F. It exhibits potent antitumor, immunosuppressive, and anti-inflammatory biological activities; however, its clinical application is hindered by severe systemic toxicity. Two preparations of T. wilfordii, including T. wilfordii glycoside tablets and T. wilfordii tablets, containing triptolide, are commonly used in clinical practice. However, their adverse side effects, particularly hepatotoxicity, limit their safe use. Therefore, it is crucial to discover potent and specific detoxification medicines for triptolide. AIM OF THE STUDY: This study aimed to investigate the detoxification effects and potential mechanism of action of spironolactone on triptolide-induced hepatotoxicity to provide a potential detoxifying strategy for triptolide, thereby promoting the safe applications of T. wilfordii preparations in clinical settings. MATERIALS AND METHODS: Cell viability was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and crystal violet staining. Nuclear fragmentation was visualized using 4',6-diamidino-2-phenylindole (DAPI) staining, and protein expression was analyzed by Western blotting. The inhibitory effect of spironolactone on triptolide-induced hepatotoxicity was evaluated by examining the effects of spironolactone on serum alanine aminotransferase and aspartate aminotransferase levels, as well as liver pathology in a mouse model of triptolide-induced acute hepatotoxicity. Furthermore, a survival assay was performed to investigate the effects of spironolactone on the survival rate of mice exposed to a lethal dose of triptolide. The effect of spironolactone on triptolide-induced global transcriptional repression was assessed through 5-ethynyl uridine staining. RESULTS: Triptolide treatment decreased the cell viability, increased the nuclear fragmentation and the cleaved caspase-3 levels in both hepatoma cells and hepatocytes. It also increased the alanine aminotransferase and aspartate aminotransferase levels, induced the hepatocyte swelling and necrosis, and led to seven deaths out of 11 mice. The above effects could be mitigated by pretreatment with spironolactone. Additionally, molecular mechanism exploration unveiled that spironolactone inhibited triptolide-induced DNA-directed RNA polymerase II subunit RPB1 degradation, consequently increased the fluorescence intensity of 5-ethynyl uridine staining for nascent RNA. CONCLUSIONS: This study shows that spironolactone exhibits a potent detoxification role against triptolide hepatotoxicity, through inhibition of RPB1 degradation induced by triptolide and, in turn, retardation of global transcriptional inhibition in affected cells. These findings suggest a potential detoxification strategy for triptolide that may contribute to the safe use of T. wilfordii preparations.

Uncovering the pharmacological mechanisms of Patchouli essential oil for treating ulcerative colitis.

ETHNOPHARMACOLOGICAL RELEVANCE: Pogostemonis Herba has long been used in traditional Chinese medicine to treat inflammatory disorders. Patchouli essential oil (PEO) is the primary component of Pogostemonis Herba, and it has been suggested to offer curative potential when applied to treat ulcerative colitis (UC). However, the pharmacological mechanisms of PEO for treating UC remain to be clarified. AIM OF THE STUDY: To elucidate the pharmacological mechanisms of PEO for treating UC. METHODS AND RESULTS: In the present study, transcriptomic and network pharmacology approaches were combined to clarify the mechanisms of PEO for treating UC. Our results reveal that rectal PEO administration in UC model mice significantly alleviated symptoms of UC. In addition, PEO effectively suppressed colonic inflammation and oxidative stress. Mechanistically, PEO can ameliorate UC mice by modulating gut microbiota, inhibiting inflammatory targets (OPTC, PTN, IFIT3, EGFR, and TLR4), and inhibiting the PI3K-AKT pathway. Next, the 11 potential bioactive components that play a role in PEO's anti-UC mechanism were identified, and the therapeutic efficacy of the pogostone (a bioactive component) in UC mice was partially validated. CONCLUSION: This study highlights the mechanisms through which PEO can treat UC, providing a rigorous scientific foundation for future efforts to develop and apply PEO for treating UC.

Diosmetin-7-O-ß-D-glucopyranoside from Pogostemonis Herba alleviated SARS-CoV-2-induced pneumonia by reshaping macrophage polarization and limiting viral replication.

ETHNOPHARMACOLOGICAL RELEVANCE: Viral pneumonia is the leading cause of death after SARS-CoV-2 infection. Despite effective at early stage, long-term treatment with glucocorticoids can lead to a variety of adverse effects and limited benefits. The Chinese traditional herb Pogostemonis Herba is the aerial part of Pogostemon Cablin (Blanco) Benth., which has potent antiviral, antibacterial, anti-inflammatory, and anticancer effects. It was used widely for treating various throat and respiratory diseases, including COVID-19, viral infection, cough, allergic asthma, acute lung injury and lung cancer. AIM OF THE STUDY: To investigate the antiviral and anti-inflammatory effects of chemical compounds from Pogostemonis Herba in SARS-CoV-2-infected hACE2-overexpressing mouse macrophage RAW264.7 cells and hACE2 transgenic mice. MATERIALS AND METHODS: The hACE2-overexpressing RAW264.7 cells were exposed with SARS-CoV-2. The cell viability was detected by CCK8 assay and cell apoptotic rate was by flow cytometric assay. The expressions of macrophage M1 phenotype markers (TNF-α and IL-6) and M2 markers (IL-10 and Arg-1) as well as the viral loads were detected by qPCR. The mice were inoculated intranasally with SARS-CoV-2 omicron variant to induce viral pneumonia. The levels of macrophages, neutrophils, and T cells in the lung tissues of infected mice were analyzed by full spectrum flow cytometry. The expressions of key proteins were detected by Western blot assay. RESULTS: Diosmetin-7-O-ß-D-glucopyranoside (DG) presented the strongest anti-SARS-CoV-2 activity. Intervention with DG at the concentrations of 0.625-2.5 µM not only reduced the viral replication, cell apoptosis, and the productions of inflammatory cytokines (IL-6 and TNF-α) in SARS-CoV-2-infected RAW264.7 cells, but also reversed macrophage polarity from M1 to M2 phenotype. Furthermore, treatment with DG (25-100 mg/kg) alleviated acute lung injury, and reduced macrophage infiltration in SARS-COV-2-infected mice. Mechanistically, DG inhibited SARS-COV-2 gene expression and HK3 translation via targeting YTHDF1, resulting in the inactivation of glycolysis-mediated NF-κB pathway. CONCLUSIONS: DG exerted the potent antiviral and anti-inflammatory activities. It reduced pneumonia in SARS-COV-2-infected mice via inhibiting the viral replication and accelerating M2 macrophage polarization via targeting YTHDF1, indicating its potential for COVID-19 treatment.

A near-infrared ratiometric fluorescent probe for the sensing and imaging of sulfur dioxide derivatives in living systems.

As a reactive sulfur species, sulfur dioxide (SO2) and its derivatives play crucial role in various physiological processes, which can maintain redox homeostasis at normal levels and lead to the occurrence of many diseases at abnormal levels. So, the development of a suitable fluorescent probe is a crucial step in advancing our understanding of the role of SO2 derivatives in living organisms. Herein, we developed a near-infrared fluorescent probe (SP) based on the ICT mechanism to monitor SO2 derivatives in living organisms in a ratiometric manner. The probe SP exhibited excellent selectivity, good sensitivity, fast response rate (within 50 s), and low detection limit (1.79 µM). In addition, the cell experiment results suggested that the SP has been successfully employed for the real-time monitoring of endogenous and exogenous SO2 derivatives with negligible cytotoxicity. Moreover, SP was effective in detecting SO2 derivatives in mice.

Triterpenes from Ganoderma lucidum inhibit hepatocellular carcinoma by regulating enhancer-associated lncRNA in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Ganoderma lucidum (G. lucidum) has been widely used as adjuvant of anti-tumor therapy for variety tumors. The bioactive ingredients of G. lucidum mainly include triterpenes, such as Ganoderic acid A, Ganoderic acid B, Ganoderenic acid A, Ganoderenic acid B, Ganoderenic acid D, and Ganoderic acid X. However, the effects and underlying mechanisms of G. lucidum are often challenging in hepatocellular carcinoma (HCC) treatment. AIM OF THE STUDY: To explore the potential role and mechanism of enhancer-associated lncRNAs (en-lncRNAs) in G. lucidum treated HCC through the in vivo and in vitro experiments. MATERIALS AND METHODS: Hepa1-6-bearing C57 BL/6 mice model were established to evaluate the therapeutic efficacy of G. lucidum treated HCC. Ki67 and TUNEL staining were used to detect the tumor cell proliferation and apoptosis in vivo. The Mouse lncRNA 4*180K array was implemented to identify the differentially expressed (DE) lncRNAs and mRNAs of G. lucidum treated tumor mice. The constructed lncRNA-mRNA co-expression network and bioinformatics analysis were used to selected core en-lncRNAs and its neighboring genes. The UPLC-MS method was used to identify the triterpenes of G. lucidum, and the in vitro experiments were used to verify which triterpene monomers regulated en-lncRNAs in tumor cells. Finally, a stable knockdown/overexpression cell lines were used to confirm the relationship between en-lncRNA and neighboring gene. RESULTS: Ki67 and TUNEL staining demonstrated G. lucidum significantly inhibited tumor growth, suppressed cell proliferation and induced apoptosis in vivo. Transcriptomic analysis revealed the existence of 126 DE lncRNAs high correlated with 454 co-expressed mRNAs in G. lucidum treated tumor mice. Based on lncRNA-mRNA network and qRT-PCR validation, 6 core lncRNAs were selected and considered high correlated with G. lucidum treatment. Bioinformatics analysis revealed FR036820 and FR121302 might act as enhancers, and qRT-PCR results suggested FR121302 might enhance Popdc2 mRNA level in HCC. Furthermore, 6 main triterpene monomers of G. lucidum were identified by UPLC-MS method, and in vitro experiments showed FR121302 and Popdc2 were significantly suppressed by Ganoderenic acid A and Ganoderenic acid B, respectively. The knock/overexpression results demonstrated that FR121302 activating and enhancing Popdc2 expression levels, and Ganoderenic acid A and Ganoderenic acid B dramatically suppressed FR121302 and decreased Popdc2 level in Hepa1-6 cells. CONCLUSIONS: Enhancer-associated lncRNA plays a crucial role as an enhancer during hepatocarcinogenesis, and triterpenes of G. lucidum significantly inhibited tumor cell proliferation and induced apoptosis by regulating en-lncRNAs. Our study demonstrated Ganoderenic acid A and Ganoderenic acid B suppressed en-lncRNA FR121302 may be one of the critical strategies of G. lucidum inhibit hepatocellular carcinoma growth.

Topical frankincense treatment for frostbite based on microcirculation improvements.

ETHNOPHARMACOLOGICAL RELEVANCE: The Chinese traditional medicine frankincense, which can promote blood circulation, is often used to treat skin lesions, including frostbite. AIM OF THE STUDY: To explore the properties of frankincense oil extract (FOE) and its active ingredients and their effect on frostbite wound recovery as an approach to understand the mechanism associated with microcirculation-improvement therapy. MATERIALS AND METHODS: The microcirculation-improving effects of FOE and its active ingredients were evaluated using liquid nitrogen-induced frostbite animal models. The rewarming capacity of FOE on the skin was determined through infrared detection, and frostbite wound healing was evaluated following haematoxylin and eosin (H&E) staining and fibre analysis. Moreover, related factors were examined to determine the anti-apoptotic, anti-inflammatory, and microcirculatory properties of FOE and its active ingredients on affected tissue in the context of frostbite. RESULTS: FOE and its active ingredients rapidly rewarmed wound tissue after frostbite by increasing the temperature. Moreover, these treatments improved wound healing and restored skin structure through collagen and elastin fibre remodelling. In addition, they exerted anti-apoptotic effects by decreasing the number of apoptotic cells, reducing caspase-3 expression, and eliciting anti-inflammatory effects by decreasing COX-2 and ß-catenin expression. They also improved microcirculatory disorders by decreasing HIF-1α expression and increasing CD31 expression. CONCLUSIONS: FOE and its active components can effectively treat frostbite by enhancing microcirculation, inhibiting the infiltration of inflammatory cells, decreasing cell apoptosis, and exerting antinociceptive effects. These findings highlight FOE as a new treatment option for frostbite, providing patients with an effective therapeutic strategy.

In vivo and in vitro study on the regulatory mechanism of XiaoChaiHu decoction on PANoptosis in sepsis-induced cardiomyopathy.

ETHNOPHARMACOLOGICAL RELEVANCE: In accordance with the tenets of traditional Chinese medicine, sepsis is categorized into three distinct syndromes: heat syndrome, blood stasis syndrome, and deficiency syndrome. Xiaochaihu decoction (XCHD) has many functions, including the capacity to protect the liver, cholagogue, antipyretic, anti-inflammatory, and anti-pathogenic microorganisms. XCHD exerts the effect of clearing heat and reconciling Shaoyang. The XCHD contains many efficacious active ingredients, yet the mechanism of sepsis-induced cardiomyopathy (SIC) remains elusive. AIM OF THE STUDY: To investigate the molecular mechanisms underlying the protective effects of XCHD against SIC using an integrated approach combining network pharmacology and molecular biology techniques. MATERIALS AND METHODS: Network pharmacology methods identified the active ingredients, target proteins, and pathways affected by XCHD in the context of SIC. We conducted in vivo experiments using mice with lipopolysaccharide-induced SIC, evaluating cardiac function through echocardiography and histology. XCHD-containing serum was analyzed to determine its principal active components using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The effects of XCHD-containing serum on SIC were further tested in vitro in LPS-treated H9c2 cardiac cells. Protein expression levels were quantified via Western blotting and enzyme-linked immunosorbent assay (ELISA). Additionally, molecular docking was performed between the active components and ZBP1, a potential target protein. Overexpression of ZBP1 in H9c2 cells allowed for a deeper exploration of its role in modulating SIC-associated gene expression. RESULTS: UPLC-MS/MS identified 31 shared XCHD and XCHD-containing serum components. These included organic acids, terpenoids, and flavonoids, which have been identified as the active components of XCHD. Our findings revealed that XCHD alleviated LPS-induced myocardial injury, improved cardiac function, and preserved cardiomyocyte morphology in mice. In vitro studies, we demonstrated that XCHD-containing serum significantly suppressed the expression of inflammatory cytokines (IL-6, IL-1ß, and TNF-α) in LPS-induced H9c2 cells. Mechanistic investigations showed that XCHD downregulated genes associated with PANoptosis, a novel cell death pathway, suggesting its protective role in sepsis-damaged hearts. Conversely, overexpression of ZBP1 abolished the protective effects of XCHD and amplified PANoptosis-related gene expression. CONCLUSIONS: Our study provides the first evidence supporting the protective effects of XCHD against SIC, both in vitro and in vivo. The underlying mechanism involves the inhibition of ZBP1-initiated PANoptosis, offering new insights into treating SIC using XCHD.

Exploring antiviral effect and mechanism of Jinye Baidu granules（JYBD）against influenza A virus through network pharmacology and in vitro and invivo experiments.

ETHNOPHARMACOLOGICAL RELEVANCE: Jinye Baidu granules (JYBD) have been used to treat acute respiratory tract infections and demonstrated clinical efficacy for the treatment of emerging or epidemic respiratory viruses such as SARS-CoV-2 and influenza virus. AIM OF THE STUDY: This study is to investigate the antiviral effect of JYBD against influenza A viruses (IAV) in vitro and in vivo and elucidate its underlying mechanism. MATERIALS AND METHODS: Ultra-high-performance liquid chromatography connected with Orbitrap mass spectrometer (UHPLC-Orbitrap MS) was employed to describe the chemical profile of JYBD. The potential pathways and targets involved in JYBD against IAV infection were predicted by network pharmacology. The efficacy and mechanism of JYBD were validated through both in vivo and in vitro experiments. Moreover, combination therapy with JYBD and the classic anti-influenza drugs was also investigated. RESULTS: A total of 126 compounds were identified by UHPLC-Orbitrap MS, of which 9 compounds were unambiguously confirmed with reference standards. JYBD could significantly inhibit the replication of multiple strains of IAV, especially oseltamivir-resistant strains. The results of qRT-PCR and WB demonstrated that JYBD could inhibit the excessive induction of pro-inflammatory cytokines induced by IAV infection and regulate inflammatory response through inhibiting JAK/STAT, NF-κB and MAPK pathways. Moreover, both JYBD monotherapy or in combination with oseltamivir could alleviate IAV-induced severe lung injury in mice. CONCLUSIONS: JYBD could inhibit IAV replication and mitigate virus-induced excessive inflammatory response. Combinations of JYBD and neuraminidase inhibitors conferred synergistic suppression of IAV both in vitro and in vivo. It might provide a scientific basis for clinical applications of JYBD against influenza virus infected diseases.