Palmatine as a potent immunomodulator: Enhancing resistance to Micropterus salmoides rhabdovirus in largemouth bass through innate immune activation and viral suppression.

Micropterus salmoides rhabdovirus (MSRV) poses a significant threat to aquaculture, causing substantial economic losses. In this study, we evaluated the antiviral efficacy and immunomodulatory potential of palmatine, a plant-derived monomer, against MSRV infection in largemouth bass. Our results demonstrated that palmatine significantly inhibited MSRV replication, with a reduction in viral nucleoprotein expression by 85 % at a safe concentration. Additionally, palmatine pre-treatment of EPC cells enhanced their antiviral capacity, with a maximum inhibition rate of 82 % following 24 h pre-incubation. Palmatine also effectively reduced MSRV-induced cytopathic effects, protecting cellular integrity and maintaining mitochondrial membrane potential. In vivo studies revealed that palmatine immersion at 80 mg/L was non-toxic and significantly suppressed MSRV replication in largemouth bass, increasing survival rates by 53 % over 15 d. Furthermore, palmatine pre-treatment enhanced the fish's resistance to MSRV, with a 78 % inhibition rate of viral replication and a 46 % increase in survival rate. Mechanistically, palmatine activated key immune genes, including IRF3, IRF7, and IFN, indicating its role in boosting innate immune responses. The compound also reduced horizontal transmission of MSRV in a cohabitation model, decreasing viral spread by up to 78 % over nine days. These findings highlight palmatine's potential as a small-molecule immunomodulator in aquaculture, offering a sustainable approach to disease management and enhancing fish health and welfare. Integrating palmatine into fish diets as an immunostimulant could provide a continuous, proactive defense against viral outbreaks, promoting more resilient and sustainable aquaculture practices.

Palmatine protects against atherosclerosis by gut microbiota and phenylalanine metabolism.

Accumulating evidence illuminated that gut microbiota directly modulates the development of atherosclerosis (AS) through interactions with metaflammation. The natural bioactive isoquinoline alkaloid palmatine (PAL), which is extracted from one of the herbs (Coptis chinensis) of the anti-AS formular, is of particular interest due to its pharmacological properties. ApoE-/- mice were administered PAL or vehicle; plaque areas, and stability were assessed by histopathological and immunohistochemistry analysis, serum glycolysis and lipid levels, and inflammation levels were also evaluated. 16S rRNA sequencing and metabolomics analysis were employed to evaluate microbial composition and serum metabolites. Microbial culture experiments were designed to reveal the target microbiota and associated metabolites. Cell culture and transcriptome were performed to elucidate the function of microbial metabolites on THP-1. PAL reduced the area of plaque and necrotic core, improving inflammatory infiltration within plaques, improving glycolipid metabolism, and reducing the levels of serum inflammatory cytokines in a dose-dependent manner. PAL treatment reshaped the composition of the gut microbiota, especially, reducing the relative abundance of Desulfovibrio piger (D. piger) in a dose-dependent manner and serum level of hippuric acid (HA). D. piger was able to convert phenylalanine into 3-phenylpropionic acid (precursor of HA). Finally, we verified HA accelerated the progression of AS and increased the secretions of inflammatory cytokines in vivo and in vitro. In conclusion, PAL exhibited anti-AS effects by regulating the gut microbiota-phenylalanine metabolism axis.

Palmatine reverse aristolochic acid-induced heart failure through activating EGFR pathway via upregulating IKBKB.

Aristolochic acid (AA) is renowned for engendering nephrotoxicity and teratogenicity. Previous literature has reported that AA treatment resulted in heart failure (HF) via inflammatory pathways. Yet, its implications in HF remain comparatively uncharted territory, particularly with respect to underlying mechanisms. In our study, the zebrafish model was employed to delineate the cardiotoxicity of AA exposure and the restorative capacity of a phytogenic alkaloid palmatine (PAL). PAL restored morphology and blood supply in AA-damaged hearts by o-dianisidine staining, fluorescence imaging, and Hematoxylin and Eosin staining. Furthermore, PAL attenuated the detrimental effects of AA on ATPase activity, implying myocardial energy metabolism recovery. PAL decreased the co-localization of neutrophils with cardiomyocytes, implying an attenuation of the inflammatory response induced by AA. A combination of network pharmacological analysis and qPCR validation shed light on the therapeutic mechanism of PAL against AA-induced heart failure via upregulation of the epidermal growth factor receptor (EGFR) signaling pathway. Subsequent evaluations using a transcriptological testing, inhibitor model, and molecular docking assay corroborated PAL as an IKBKB enzyme activator. The study underscores the possible exploitation of the EGFR pathway as a potential therapeutic target for PAL against AA-induced HF, thus furthering the continued investigation of the toxicology and advancement of protective pharmaceuticals for AA.

Palmatine ameliorates N-methyl-N'-nitrosoguanidine-induced chronic atrophic gastritis through the STAT1/CXCL10 axis.

Chronic atrophic gastritis (CAG) is a prevalent preneoplastic condition of the stomach. Palmatine (PAL), an isoquinoline alkaloid isolated from Rhizoma Coptidis (RC), has significant anti-inflammatory properties and is often used to treat gastrointestinal disorders. However, the mechanism of PAL on CAG remains unclear. In this study, N-methyl-N'-nitrosoguanidine (MNNG) was used to induce CAG inflammatory disease models in vivo and in vitro. The efficacy of five alkaloids in RC and the dose-dependent effects of the most effective PAL in CAG mice were evaluated in two animal experiments. RNA-seq and western blot revealed that PAL significantly improved IL-17, TNF, and NF-kappa B inflammation-related signaling pathways. Further hub gene prediction and experimental validation revealed that PAL modulated the STAT1/CXCL10 axis, thereby exerting attenuation of CAG through the regulation of IL-17, TNF-α, and p-p65 expression. In conclusion, PAL was proposed to mitigate MNNG-induced CAG, potentially through the inhibition of oxidative stress and inflammatory responses via the STAT1/CXCL10 axis. This approach is an effective complement to the use of PAL in the treatment of CAG.

Isolation and Quantification of Palmatine from Fibraurea tinctoria Lour: In Vitro Antioxidant Activity and In Silico Antidiabetic Activity Evaluation.

Purpose: This study aimed to isolate and characterize palmatine from Fibraurea tinctoria Lour stems, quantify its content, and determine its antioxidant and antidiabetic activities. Patients and Methods: Palmatine was isolated from the methanol extract of Fibraurea tinctoria Lour stems by silica gel column chromatography. Structural elucidation of the isolated compounds was performed using spectral data analysis and comparison with the literature. High-Performance Liquid Chromatography (HPLC) was used to quantitatively determine palmatine in the crude methanol extract and fractions. The DPPH and non-enzymatic SOD mimic methods were used to assess the antioxidant activity of the methanol extract, fractions, and isolated compounds. The antidiabetic activity was evaluated in silico by the molecular docking method of alpha-glucosidase and DPP-IV enzymes. Palmatine was used as a test ligand and was compared with berberine and its native ligand or standard compounds. Results: The isolated compound was identified as palmatine. Quantification of palmatine compound by HPLC showed that palmatine was found in the extract and all fractions. In the in vitro antioxidant activity test using the DPPH method, fraction 4 showed the highest activity, with an IC50 value of 91 ppm. In contrast, using the non-enzymatic SOD mimic method, the methanol extract, fraction 5, and isolated compound (palmatine) exhibited very strong antioxidant activity, with IC50 values of 18, 20, and 28 ppm, respectively. The in silico antidiabetic activity of palmatine is thought to have the potential to inhibit these two enzymes. Conclusion: These results showed that Fibraurea tinctoria Lour stems have potential as an antioxidant and antidiabetic agent. Further research on phytochemical and pharmacological is required to validate the use of this plant species for the treatment of various diseases, especially diabetes mellitus.

Palmatine Ameliorates Motor Deficits and Dopaminergic Neuron Loss by Regulating NLRP3 Inflammasome through Mitophagy in Parkinson's Disease Model Mice.

NLRP3 inflammasomes-mediated proinflammatory response and mitochondrial dysfunction play a critical role in the etiology and pathogenesis of Parkinson's disease. Negative regulation of NLRP3 inflammasome activation through mitophagy may be an important strategy to control NLRP3 inflammasome-mediated proinflammatory responses. Palmatine (PAL), an isoquinoline alkaloid found in various of plants, has potent pharmacological effects such as anti-inflammatory and anti-oxidation. However, the specific role of PAL in the pathology of Parkinson's disease remains unclear. In this study, we found that treatment with PAL improved motor deficits and reduced the loss of dopaminergic neurons in MPTP mice. Further results showed that PAL promoted mitophagy and inhibited the proinflammatory response mediated by NLRP3 inflammasomes. In addition, chloroquine (CQ, mitophagy inhibitor) attenuated the ameliorative effects of PAL on the motor deficits and dopaminergic neuron damage, as well as the inhibitory effect of PAL on NLRP3 inflammasome. Collectively, these results provide strong evidence that PAL ameliorates motor deficits and dopaminergic neuron death in Parkinson's disease, and the mechanism may be related to its inhibition of NLRP3 inflammasome activation via promoting mitophagy.

Piperine Enhances Antimalarial Activity of Methyl Gallate and Palmatine Combination.

PURPOSE: Artemisinin combination therapies, the first-line antimalarials in Nigeria, have reportedly suffered multiple failures in malaria treatment, hence the search for novel combination of other compounds. Methyl gallate and palmatine have been reported to exhibit antiplasmodial activities but the antimalarial activity of their combination has not been evaluated. Therefore, the evaluation of the combination of methyl gallate and palmatine for antimalarial activity in vitro and in vivo in the presence of piperine was carried out. MATERIALS AND METHODS: The inhibitory potential of methyl gallate and palmatine combination on ß-hematin (hemozoin) formation was studied in vitro. Also, the antimalarial activity of methyl gallate and palmatine combination with/without a bioenhancer (piperine) was evaluated in Plasmodium berghei NK65-infected mice. RESULTS: Methyl gallate and palmatine in the ratio 3:2 acted synergistically in vitro and had the highest inhibitory effect (IC50 = 0.73 µg/mL) on ß-hematin (hemozoin) formation. The 3:2 combination of methyl gallate and palmatine exhibited no antimalarial activity in vivo in the absence of piperine but caused reduction in parasitemia that exceeded 40% in the presence of piperine at the dose of 25 mg/kg body weight on days 6 and 8 post-inoculation in mice. CONCLUSION: The 3:2 combination of methyl gallate and palmatine in the presence of piperine exhibited antimalarial activity in vivo, possibly by synergistic inhibition of hemozoin formation which may cause accumulation of haem within the food vacuole of Plasmodium spp. and its death.

Palmatine inhibits expression fat mass and obesity associated protein (FTO) and exhibits a curative effect in dextran sulfate sodium (DSS)-induced experimental colitis.

BACKGROUND: Ulcerative colitis (UC) is an inflammatory disease whose pathogenesis and mechanisms have not been fully described. The m6A methylation modification is a general mRNA modification in mammalian cells and is closely associated with the onset and progression of inflammatory bowel disease (IBD). Palmatine (PAL) is a biologically active alkaloid with anti-inflammatory and protective effects in animal models of colitis. Accordingly, we examined the role of PAL on colitis by regulating N6-methyladenosine (m6A) methylation. METHODS: A rat experimental colitis model was established by 5 % dextran sulfate sodium (DSS) in drinking water for seven days, then PAL treatment was administered for seven days. The colonic tissue pathology was assessed using hematoxylin-eosin (HE) and disease activity index (DAI). In in vitro studies, a human, spontaneously immortalized non-cancerous colon mucosal epithelial cell line (NCM460) was exposed to 2 % DSS and treated with PAL and cell viability was assayed using Cell Counting Kit-8 (CCK-8). The levels of tumor necrosis factor α (TNF-α), interleukin (IL)-1ß, IL-6, and IL-8 were detected by enzyme-linked immunosorbent assay (ELISA) kits. The level of Zonula occludens-1 (ZO-1) was dectected by immunofluorescence. Transepithelial electrical resistance (TEER) of cells was also assessed. The methyltransferase-like 3 (METTL3), METTL14, AlkB homologate 5 (ALKBH5), and fat mass and obesity-associated protein (FTO) expression levels were assessed by western blotting. The localized expression of m6A was measured by immunofluorescence. RESULTS: PAL significantly prevented bodyweight loss and shortening of the colon in experimental colitis rats, as well as decreasing the DAI and histological damage scores. Furthermore, PAL inhibited the levels of inflammatory factors (TNF-α, IL-6, IL-8, and IL-1ß) in both DSS treated rats and NCM460 cells. In addition, PAL enhanced the expression level of ZO-1, and increased the transepithelial electrical resistance to repaire intestinal barrier dysfunction. Colitis occurred due to decreased m6A levels, and the increased FTO expression led to a colitis phenotype. PAL markedly enhanced the METTL3 and METTL14 expression levels while decreasing ALKBH5 and FTO expression levels. CONCLUSIONS: The findings demonstrated that PAL improved DSS-induced experimental colitis. This effect was associated with inhibiting FTO expression and regulating m6A methylation.

Insights into the interaction and inhibitory action of palmatine on lysozyme fibrillogenesis: Spectroscopic and computational studies.

Interaction under amyloidogenic condition between naturally occurring protoberberine alkaloid palmatine and hen egg white lysozyme was executed by adopting spectrofluorometric and theoretical molecular docking and dynamic simulation analysis. In spetrofluorometric method, different types of experiments were performed to explore the overall mode and mechanism of interaction. Intrinsic fluorescence quenching of lysozyme (Trp residues) by palmatine showed effective binding interaction and also yielded different binding parameters like binding constant, quenching constant and number of binding sites. Synchronous fluorescence quenching and 3D fluorescence map revealed that palmatine was able to change the microenvironment of the interacting site. Fluorescence life time measurements strongly suggested that this interaction was basically static in nature. Molecular docking result matched with fluorimetric experimental data. Efficient drug like interaction of palmatine with lysozyme at low pH and high salt concentration prompted us to analyze its antifibrillation potential. Different assays and microscopic techniques were employed for detailed analysis of lysozyme amyloidosis.Thioflavin T(ThT) assay, Congo Red (CR) assay, 8-anilino-1-naphthalenesulfonic acid (ANS) assay, Nile Red (NR) assay, anisotropy and intrinsic fluorescence measurements confirmed that palmatine successfully retarded and reduced lysozyme fibrillation. Dynamic light scattering (DLS) and atomic force microscopy (AFM) further reiterated the excellent antiamyloidogenic potency of palmatine.

Discovery of palmatine derivatives as potent neuroprotective agents.

Alzheimer's disease is a neurodegenerative disorder characterized by the presence of neurodegenerative lesions and cognitive impairment. In this study, a series of novel palmatine derivatives were designed and synthesized through the introduction of a heteroatom using carbodiimide-mediated condensation. The synthesized compounds were then screened for toxicity and potency, leading to the identification of compound 2q, which exhibited low toxicity and high potency. Our findings demonstrated that compound 2q displayed significant neuroprotective activity in vitro, emerging as a promising candidate for Alzheimer's disease treatment.

Palmatine Attenuated Lipopolysaccharide-Induced Acute Lung Injury by Inhibiting M1 Phenotype Macrophage Polarization via NAMPT/TLR2/CCR1 Signaling.

The present work was conducted to research the potential mechanism of palmatine (PAL) on lipopolysaccharide (LPS)-caused acute lung injury (ALI). Network pharmacology and bioinformatic analyses were carried out. Mice were intragastrically treated with PAL and intratracheally stimulated with LPS. LPS-induced RAW264.7 cells were employed for the in vitro model. The MPO activity, W/D ratio, neutrophils, total cell number in BALF, and histopathological alteration were examined. The levels of TNF-α, IL-1ß, IL-6, IL-18, IL-4, and IL-10 in serum, BALF, and supernatant were examined by ELISA. The mRNA expressions of iNOS, CD68, Arg1, Ym1, and CD206 and protein expressions of NAMPT, TLR2, CCR1, and NLRP3 inflammasome were detected by PCR, WB, and immunofluorescence. The NAMPT inhibitor FK866, TLR2 inhibitor C29, CCR1 inhibitor BX471, NAMPT-overexpression (OE) plasmid, and TLR2-OE plasmid were used for mechanism research. As a result, PAL relieved the symptoms of ALI. PAL inhibited M1 phenotype indices and promoted M2 phenotype indices in both LPS-induced mice and RAW264.7 cells. PAL also inhibited the expressions of NAMPT, TLR2, CCR1, and NLRP3 inflammasome. The treatments with FK866, NAMPT-OE plasmid, C29, TLR2-OE plasmid, and BX471 proved that PAL exerted its effect via NAMPT/TLR2/CCR1. Molecular docking suggested that PAL might combine with NAMPT. In conclusion, PAL ameliorated LPS-induced ALI by inhibiting M1 phenotype macrophage polarization via NAMPT/TLR2/CCR1 signaling.

Palmatine Alleviates Particulate Matter-Induced Acute Lung Injury by Inhibiting Pyroptosis via Activating the Nrf2-Related Pathway.

Particulate matter (PM) induces and enhances oxidative stress and inflammation, leading to a variety of respiratory diseases, including acute lung injury. Exploring new treatments for PM-induced lung injury has long been of interest to researchers. Palmatine (PAL) is a natural extract derived from plants that has been reported in many studies to alleviate inflammatory diseases. Our study was designed to explore whether PAL can alleviate acute lung injury caused by PM. The acute lung injury model was established by instilling PM (4 mg/kg) into the airway of mice, and PAL (50 mg/kg and 100 m/kg) was administrated orally as the treatment groups. The effect and mechanism of PAL treatment were examined by immunofluorescence, immunohistochemistry, Western Blotting, ELISA, and other experiments. The results showed that oral administration of PAL (50 mg/kg and 100 m/kg) could significantly alleviate lung inflammation and acute lung injury caused by PM. In terms of mechanism, we found that PAL (50 mg/kg) exerts anti-inflammatory and anti-damage effects mainly by enhancing the activation of the Nrf2-related antioxidant pathway and inhibiting the activation of the NLRP3-related pyroptosis pathway in mice. These mechanisms have also been verified in our cell experiments. Further cell experiments showed that PAL may reduce intracellular reactive oxygen species (ROS) by activating Nrf2-related pathways, thereby inhibiting the activation of NLRP3-related pyroptosis pathway induced by PM in Beas-2B cell. Our study suggests that PAL can be a new option for PM-induced acute lung injury.

Protective Effects of Coptis chinensis Rhizome Extract and Its Constituents (Berberine, Coptisine, and Palmatine) against α-Synuclein Neurotoxicity in Dopaminergic SH-SY5Y Cells.

Parkinson's disease (PD) is a common neurodegenerative disease with progressive loss of dopaminergic neurons in substantia nigra and the presence of α-synuclein-immunoreactive inclusions. Gaucher's disease is caused by homozygous mutations in ß-glucocerebrosidase gene (GBA). GBA mutation carriers have an increased risk of PD. Coptis chinensis (C. chinensis) rhizome extract is a major herb widely used to treat human diseases. This study examined the association of GBA L444P mutation with Taiwanese PD in 1016 cases and 539 controls. In addition, the protective effects of C. chinensis rhizome extract and its active constituents (berberine, coptisine, and palmatine) against PD were assayed using GBA reporter cells, LC3 reporter cells, and cells expressing mutated (A53T) α-synuclein. Case-control study revealed that GBA L444P carriers had a 3.93-fold increased risk of PD (95% confidence interval (CI): 1.37-11.24, p = 0.006) compared to normal controls. Both C. chinensis rhizome extract and its constituents exhibited chemical chaperone activity to reduce α-synuclein aggregation. Promoter reporter and endogenous GBA protein analyses revealed that C. chinensis rhizome extract and its constituents upregulated GBA expression in 293 cells. In addition, C. chinensis rhizome extract and its constituents induced autophagy in DsRed-LC3-expressing 293 cells. In SH-SY5Y cells expressing A53T α-synuclein, C. chinensis rhizome extract and its constituents reduced α-synuclein aggregation and associated neurotoxicity by upregulating GBA expression and activating autophagy. The results of reducing α-synuclein aggregation, enhancing GBA expression and autophagy, and protecting against α-synuclein neurotoxicity open up the therapeutic potentials of C. chinensis rhizome extract and constituents for PD.

Berberine Metabolites Stimulate GLP-1 Secretion by Alleviating Oxidative Stress and Mitochondrial Dysfunction.

Berberine (BBR) is a principal component of Rhizoma coptidis known for its therapeutic potential in treating diseases such as type 2 diabetes mellitus (T2DM) and obesity. Despite the trace levels of BBR in plasma, it's believed that its metabolites play a pivotal role in its biological activities. While BBR is recognized to promote GLP-1 production in intestinal L cells, the cytoprotective effects of its metabolites on these cells are yet to be explored. The present study investigates the effects of BBR metabolites on GLP-1 secretion and the underlying mechanisms. Our results revealed that, out of six BBR metabolites, berberrubine (BBB) and palmatine (PMT) significantly increased the production and glucose-stimulated secretion of GLP-1 in GLUTag cells. Notably, both BBB and PMT could facilitate GLP-1 and insulin secretion and enhance glucose tolerance in standard mice. Moreover, a single dose of PMT could markedly increase plasma GLP-1 and improve glucose tolerance in mice with obesity induced by a high-fat diet. In palmitic acid or TNF[Formula: see text]-treated GLUTag cells, BBB and PMT alleviated cell death, oxidative stress, and mitochondrial dysfunction. Furthermore, they could effectively reverse inflammation-induced inhibition of the Akt signaling pathway. In general, these insights suggest that the beneficial effects of orally administered BBR on GLP-1 secretion are largely attributed to the pharmacological activity of BBB and PMT by their above cytoprotective effects on L cells, which provide important ideas for stimulating GLP-1 secretion and the treatment of T2DM.

Evaluation of iso-quinoline alkaloids by RP-HPLC-PDA in different Berberis species collected from Western Himalayan Region.

The genus Berberis has high significance in Indian and other traditional systems of medicine due to the presence of iso-quinoline alkaloids. This study was conducted to record the metabolic variation in eight Berberis spp. collected from the Western Himalayan region. The RP-HPLC-PDA chromatogram separates the markers jatrorrhizine, palmatine and berberine at specific retention time, 11.45, 16.41 and 18.15 min respectively. The method was validated on linearity, precision and recovery indices as per International Conference on Harmonisation (ICH) guidelines. The maximum content of berberine, palmatine and jatrorrhizine was found in Berberis asiatica (1.74% ± 0.032, 1.63% ± 0.028 and 0.264% ± 0.012, respectively) on % dry weight basis. This study will help in phyto-marker-based identification and differentiation of various Berberis species, which can be used as an alternative to the official drug Daruharidra i.e. Berberis aristata.

Palmatine alleviates cardiac fibrosis by inhibiting fibroblast activation through the STAT3 pathway.

Cardiac fibrosis, the hallmark of cardiovascular disease, is characterized by excessive deposition of extracellular matrix in the heart. Emerging evidence indicates that cardiac fibroblasts (CFs) play pivotal roles in driving cardiac fibrosis. However, due to incomplete insights into CFs, there are limited effective approaches to prevent or reverse cardiac fibrosis currently. Palmatine, a protoberberine alkaloid extracted from traditional Chinese botanical remedies, possesses diverse biological effects. This study investigated the potential therapeutic value and mechanism of palmatine against cardiac fibrosis. Adult male C57BL/6 mice were treated with vehicle, isoproterenol (ISO), or ISO plus palmatine for one week. After echocardiography assessment, mice hearts were collected for histopathology, real-time polymerase chain reaction, and Western blot analyses. Primary rat CFs were utilized in vitro. Compared to control, ISO-treated mice exhibited cardiac hypertrophy and structural abnormalities; however, treatment with palmatine ameliorated these effects of ISO. Moreover, palmatine treatment mitigated ISO-induced cardiac fibrosis. Network pharmacology and molecular docking analysis showed that palmatine strongly binds the regulators of cardiac fibrosis including signal transducer and activator of transcription 3 (STAT3) and mammalian target of rapamycin. Furthermore, palmatine reduced the elevated fibrotic factor expressions and overactivated STAT3 induced by ISO, Transformed growth factor ß1 (TGF-ß1), or interleukin-6 both in vivo and in vitro. Additionally, blocking STAT3 suppressed the TGF-ß1-induced CF activation. Collectively, these data demonstrated that palmatine attenuated cardiac fibrosis partly by inhibiting fibroblast activation through the STAT3 pathway. This provides an experimental basis for the clinical treatment of cardiac fibrosis with palmatine.

A comprehensive assessment of palmatine as anticonvulsant agent - In vivo and in silico studies.

Previously, we demonstrated that palmatine (PALM) - an isoquinoline alkaloid from Berberis sibrica radix, exerted antiseizure activity in the pentylenetetrazole (PTZ)-induced seizure assay in larval zebrafish. The aim of the present study was to more precisely characterize PALM as a potential anticonvulsant drug candidate. A range of zebrafish and mouse seizure/epilepsy models were applied in the investigation. Immunostaining analysis was conducted to assess the changes in mouse brains, while in silico molecular modelling was performed to determine potential targets for PALM. Accordingly, PALM had anticonvulsant effect in ethyl 2-ketopent-4-enoate (EKP)-induced seizure assay in zebrafish larvae as well as in the 6 Hz-induced psychomotor seizure threshold and timed infusion PTZ tests in mice. The protective effect in the EKP-induced seizure assay was confirmed in the local field potential recordings. PALM did not affect seizures in the gabra1a knockout line of zebrafish larvae. In the scn1Lab-/- zebrafish line, pretreatment with PALM potentiated seizure-like behaviour of larvae. Repetitive treatment with PALM, however, did not reduce development of PTZ-induced seizure activity nor prevent the loss of parvalbumin-interneurons in the hippocampus of the PTZ kindled mice. In silico molecular modelling revealed that the noted anticonvulsant effect of PALM in EKP-induced seizure assay might result from its interactions with glutamic acid decarboxylase and/or via AMPA receptor non-competitive antagonism. Our study has demonstrated the anticonvulsant activity of PALM in some experimental models of seizures, including a model of pharmacoresistant seizures induced by EKP. These results indicate that PALM might be a suitable new drug candidate but the precise mechanism of its anticonvulsant activity has to be determined.

Antiviral effect of palmatine against infectious bronchitis virus through regulation of NF-κB/IRF7/JAK-STAT signalling pathway and apoptosis.

1. Infectious bronchitis virus (IBV), a gamma-coronavirus, can infect chickens of all ages and leads to an acute contact respiratory infection. This study evaluated the anti-viral activity of palmatine, a natural non-flavonoid alkaloid, against IBV in chicken embryo kidney (CEK) cells.2. The half toxic concentration (CC50) of palmatine was 672.92 µM, the half inhibitory concentration (IC50) of palmatine against IBV was 7.76 µM and the selection index (SI) was 86.74.3. Mode of action assay showed that palmatine was able to directly inactivate IBV and inhibited the adsorption, penetration and intracellular replication of IBV.4. Palmatine significantly upregulated TRAF6, TAB1 and IKK-ß compared with the IBV-infected group, leading to the increased expressions of pro-inflammatory cytokines IL-1ß and TNF-α in the downstream NF-κB signalling pathway.5. Palmatine significantly up-regulated the levels of MDA5, MAVS, IRF7, IFN-α and IFN-ß in the IRF7 pathway, inducing type I interferon production. It up-regulated the expression of 2'5'-oligoadenylate synthase (OAS) in the JAK-STAT pathway.6. IBV infection induced cell apoptosis and palmatine-treatment delayed the process of apoptosis by regulation of the expression of apoptosis-related genes (BAX, BCL-2, CASPASE-3 and CASPASE-8).7. Palmatine could exert anti-IBV activity through regulation of NF-κB/IRF7/JAK-STAT signalling pathways and apoptosis, providing a theoretical basis for the utilisation of palmatine to treat IBV infection.

Palmatine ameliorated lipopolysaccharide-induced sepsis-associated encephalopathy mice by regulating the microbiota-gut-brain axis.

BACKGROUND: Sepsis-associated encephalopathy (SAE), a common neurological complication from sepsis, is widespread among patients in intensive care unit and is linked to substantial morbidity and mortality rates, thus posing a substantial menace to human health. Due to the intricate nature of SAE's pathogenesis, there remains a dearth of efficacious therapeutic protocols, encompassing pharmaceutical agents and treatment modalities, up until the present time. Palmatine exhibits distinctive benefits in the regulation of inflammation for the improvement of sepsis. Nevertheless, the precise functions of palmatine in treating SAE and its underlying mechanism have yet to be elucidated. PURPOSE: This study aimed to evaluate efficiency of palmatine in SAE mice and its underlying mechanisms. STUDY DESIGN AND METHODS: Behavioral experiments, percent survival rate analysis, histological analysis, immunofluorescence staining, ELISA analysis, were performed to evaluate the efficiency of palmatine in SAE mice. Quantibody® mouse inflammation array glass chip was performed to observe the effects of palmatine on inflammation storm in SAE mice. Real-time quantitative and western blotting analyzes were employed to examine the expression of relevant targets in the Notch1/nuclear factor-kappa B (NF-κB) pathway. Finally, brain tissues metabolomics-based analyzes were performed to detect the differentially expressed metabolites and metabolic pathways. The fecal samples were subjected to microbial 16S rRNA analysis and untargeted metabolomics analysis in order to identify the specific flora and metabolites associated with SAE, thereby further investigating the mechanism of palmatine in SAE mice. RESULTS: Our results showed that palmatine significantly improved nerve function, reduced cell apoptosis in brain tissue, and decreased inflammatory cytokine levels in SAE induced-LPS mice. Meanwhile, our results demonstrate the potential of palmatine in modulating key components of the Notch1/NF-κB pathway, enhancing the expression of tight junction proteins, improving intestinal permeability, promoting the growth of beneficial bacteria (such as Lachnospiraceae_NK4A136_group), inhibiting the proliferation of harmful bacteria (such as Escherichia-Shigella), and mitigating metabolic disorders. Ultimately, these observed effects contribute to the therapeutic efficacy of palmatine in treating SAE. CONCLUSION: The findings of our study have provided confirmation regarding the efficacy of palmatine in the treatment of SAE, thereby establishing a solid foundation for further exploration into SAE therapy and the advancement and investigation of palmatine.

Determination of four different components in Danggui Liuhuang decoction by HPLC-MS/MS / 药学实践与服务

Objective To establish the method of simultaneous determination of four main components of Danggui Liuhuang Decoction, including phellodendrine, palmatine, calycosin, and ferulic acid and provide reference for the quality control of Danggui Liuhuang Decoction. Methods Based on the HPLC-MS/MS analysis method, the positive ion data acquisition mode were adopted for the mass spectrometry detection and the four main components were quantified with multiple reaction monitoring mode (MRM) by ESI source. The chromatographic column was Agilent Extend-C18 (5 μm, 4.6 mm×250 mm), and gradient elution was performed with methanol and 0.5% formic acid in water. Results The linear range of phellodendrine was from 2-200 nmol/ml, and the linear range of palmatine, calycosin and ferulic acid was from 20-2 000 nmol/ml. The contents of the four components in the seven batches of Danggui Liuhuang Decoction were relatively stable, among which ferulic acid was mainly found in Phellodendrine and Coptidis; Phellodendrine was only detected in cortex phellodendri; the content of calycosin in Scutellaria baicalensis and Astragalus was higher; palmatine was detected in both Phellodendron and Astragalus. Conclusion The method had high sensitivity, good specificity and sample stability, which could meet the requirements of quantitative analysis of Traditional Chinese Medicine compounds, and could provide reference for further pharmacokinetics study on the content changes of traditional Chinese medicine compounds in biological samples.