Nitrogen Sources Reprogram Carbon and Nitrogen Metabolism to Promote Andrographolide Biosynthesis in Andrographis paniculata (Burm.f.) Nees Seedlings.

Carbon (C) and nitrogen (N) metabolisms participate in N source-regulated secondary metabolism in medicinal plants, but the specific mechanisms involved remain to be investigated. By using nitrate (NN), ammonium (AN), urea (UN), and glycine (GN), respectively, as sole N sources, we found that N sources remarkably affected the contents of diterpenoid lactone components along with C and N metabolisms reprograming in Andrographis paniculata, as compared to NN, the other three N sources raised the levels of 14-deoxyandrographolide, andrographolide, dehydroandrographolide (except UN), and neoandrographolide (except AN) with a prominent accumulation of farnesyl pyrophosphate (FPP). These N sources also raised the photosynthetic rate and the levels of fructose and/or sucrose but reduced the activities of phosphofructokinase (PFK), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoenolpyruvate carboxylase (PEPC) and pyruvate dehydrogenase (PDH). Conversely, phosphoenolpyruvate carboxykinase (PEPCK) and malate enzyme (ME) activities were upregulated. Simultaneously, citrate, cis-aconitate and isocitrate levels declined, and N assimilation was inhibited. These results indicated that AN, UN and GN reduced the metabolic flow of carbohydrates from glycolysis into the TCA cycle and downstream N assimilation. Furthermore, they enhanced arginine and GABA metabolism, which increased C replenishment of the TCA cycle, and increased ethylene and salicylic acid (SA) levels. Thus, we proposed that the N sources reprogrammed C and N metabolism, attenuating the competition of N assimilation for C, and promoting the synthesis and accumulation of andrographolide through plant hormone signaling. To obtain a higher production of andrographolide in A. paniculata, AN fertilizer is recommended in its N management.

Andrographolide Improves ApoE4-Mediated Blood-Brain Barrier Injury by Alleviating Inflammation.

The pathological and physiological studies of Alzheimer's disease (AD) have been in-depth, and apolipoprotein E4 (ApoE4) has been proven to be highly correlated with AD, and clinical and experimental data show that ApoE4 can cause blood-brain barrier (BBB) injury, and the change of BBB permeability is an important factor affecting the development of AD. Andrographolide (Andro), as the active component of the natural plant Andrographis paniculata, has been proven to have anti-inflammatory and antioxidant effects, which have potential neuroprotective effects. To verify the protective effect of Andro on BBB in a short term, our research group used atorvastatin (Atorva)-mediated zebrafish brain injury model and the ApoE4-mediated cell co-culture model of BBB injury to explore the protective effects and mechanisms of Andro on BBB injury. Studies have shown that Andro can inhibit the activation of CypA/NF-κB/MMP-9 signaling pathway and has achieved the effect of antagonizing the inhibition of ApoE4 on intercellular tight junction proteins (occludin, claudin-5, and ZO-1). At the same time, Andro can inhibit the secretion of cell adhesion molecules (VCAM-1 and ICAM-1) in cells, thereby delaying the occurrence and progression of neuroinflammation and playing a protective role in BBB. In conclusion, Andro is a potent natural product which can protect the blood-brain barrier.

Andrographolide Ameliorates Inflammatory Changes Induced by D-Lactate in Bovine Fibroblast-like Synoviocytes.

During acute ruminal acidosis, the manifestation of aseptic polysynovitis and lameness in cattle has been observed. Evidence suggests that joint inflammation can be attributed to the metabolic alterations induced by D-lactate in fibroblast-like synoviocytes (FLSs). We aimed to investigate whether andrographolide could mitigate the inflammation and metabolic alterations induced by D-lactate in bovine fibroblast-like synoviocytes (bFLSs). To assess this, bFLSs were cultured in the presence or absence of andrographolide. We evaluated its potential interference with the expression of proinflammatory cytokines, COX-2, HIF-1α, and LDHA using RT-qPCR. Furthermore, we investigated its potential interference with PI3K/Akt signaling and IκBα degradation through immunoblotting and flow cytometry, respectively. Our observations revealed that andrographolide reduced the elevation of IL-6, IL-8, COX-2, HIF-1α, and LDHA induced by D-lactate. Additionally, andrographolide demonstrated interference with the PI3K/Akt and NF-κB pathways in bFLSs. In conclusion, our findings suggest that andrographolide can potentially reverse the inflammatory effects and metabolic changes induced by D-lactate in bFLSs, showing promise as a therapeutic intervention for managing these conditions associated with lameness.

Elucidation of the key therapeutic targets and potential mechanisms of Andrographolide multi-targets against osteoarthritis via network pharmacological analysis and experimental validation.

OBJECTIVE: Our purpose is to unveil Andrographolide's potential multi-target and multi-mechanism therapeutic effects in treating OA via systematic network pharmacological analysis and cell experimental validation. MATERIALS AND METHODS: Initially, we gathered data from Andrographolide and OA-related databases to obtain information on Andrographolide's biological properties and the targets linked with OA. We developed a bioinformatic network about Andrographolide and OA, whereby we analyzed the network to identify potential therapeutic targets and mechanisms of action of Andrographolide. Subsequently, we used molecular docking to analyze the binding sites of Andrographolide to the target proteins. At the same time, SDF-1 was used to construct an OA cell model to verify the therapeutic effect of Andrographolide on OA and its effect on target proteins. RESULTS: Our experimental results show that Andrographolide has excellent pharmaceutical properties, by Lipinski's rules for drugs, suggesting that this compound can be considered to have a high therapeutic potential in drug development. 233 targets were preliminarily investigated, the mechanisms through which Andrographolide targets OA primarily involve the TNF signaling pathway, PI3K-AKT signaling pathway, IL-17 signaling pathway, and TLR signaling pathway. These mechanisms target OA by influencing immune and inflammatory responses in the joints, regulating apoptosis to prevent chondrocyte death. Finally, TNF-α, STAT3, TP53, IL-6, JUN, IL-1ß, HIF-1α, TGF-ß1, and AKT1 were identified as 9 key targets of Andrographolide anti-OA. In addition, our molecular docking analyzes with cell experimental validation further confirm the network pharmacology results. According to our molecular docking results, Andrographolide can bind to all the hub target proteins and has a good binding ability (binding energy < -5 kcal/mol), with the strongest binding affinity to AKT1 of -9.2 kcal/ mol. The results of cell experiments showed that Andrographolide treatment significantly increased the cell viability and the expression of COL2A1 and ACAN proteins. Moreover, 30 µM Andrographolide significantly reversed SDF-1-induced increases in the protein expression of TNF-α, STAT3, TP53, IL-6, JUN, IL-1ß, HIF-1α, and TGF-ß1, and decreases in the protein expression of AKT1. CONCLUSION: This study provides a comprehensive understanding of the potential therapeutic targets and mechanisms of action of Andrographolide in OA treatment. Our findings suggest that Andrographolide is a promising candidate for drug development in the management of OA.

PCSK5 downregulation promotes the inhibitory effect of andrographolide on glioblastoma through regulating STAT3.

Proprotein convertase subtilisin/kexin type 5 (PCSK5) is a member of the proprotein convertase (PC) family, which processes immature proteins into functional proteins and plays an important role in the process of cell migration and transformation. Andrographolide is a non-peptide compound with PC inhibition and antitumor activity. Our research aimed to investigate the functional role of PCSK5 downregulation combined with Andro on GBM progression. Results from the cancer genome atlas (TCGA) and clinical samples revealed a significant upregulation of PCSK5 in GBM tissues than in non-tumor brain tissues. Higher expression of PCSK5 was correlated with advanced GBM stages and worse patient prognosis. PCSK5 knockdown attenuated the epithelial-mesenchymal transition (EMT)-like properties of GBM cells induced by IL-6. PCSK5 knockdown in combination with Andro treatment significantly inhibited the proliferation and invasion of GBM cells in vitro, as well as tumor growth in vivo. Mechanistically, PCSK5 downregulation reduced the expression of p-STAT3 and Matrix metalloproteinases (MMPs), which could be rescued by the p-STAT3 agonist. STAT3 silencing downregulated the expression of MMPs without affecting PCSK5. Furthermore, Andro in combination with PCSK5 silencing significantly inhibited STAT3/MMPs axis. These observations provided evidence that PCSK5 functioned as a potential tumor promoter by regulating p-STAT3/MMPs and the combination of Andro with PCSK5 silencing might be a good strategy to prevent GBM progression.

Discovery of Pesticide Candidates from Natural Plant Products: Semisynthesis and Characterization of Andrographolide-Based Esters and Study of Their Pesticidal Properties and Toxicology.

To explore the use of nonfood plant-derived secondary metabolites for plant protection, a series of ester derivatives for controlling the major migratory agricultural pests were obtained by structural modification of andrographolide, a labdane diterpenoid isolated from Andrographis paniculata. Compound Id showed good insecticidal activity against the fall armyworm Spodoptera frugiperda Smith. Compounds IIa (LC50: 0.382 mg/mL) and IIIc (LC50: 0.563 mg/mL), the acaricidal activities of which were, respectively, 13.1 and 8.9 times that of andrographolide (LC50: 4.996 mg/mL), exhibited strong acaricidal and control effects against Tetranychus cinnabarinus Boisduval. Against Aphis citricola Van der Goot, compounds IIIc and IVb displayed 3.9- and 3.7-fold pronounced aphicidal activity of andrographolide. Effects of compound Id on three protective enzymes (superoxide dismutase, peroxidase, and catalase) of S. frugiperda were also observed. The obvious differences of epidermal cuticle structures of mites treated with compound IIa were determined by scanning electron microscopy. Structure-activity relationships indicated that 14-ester derivatives of andrographolide showed potential insecticidal/acaricidal activities and can be further utilized as lead compounds.

Andrographolide attenuates sepsis-induced acute kidney injury by inhibiting ferroptosis through the Nrf2/FSP1 pathway.

Sepsis is a systemic inflammatory response syndrome caused by infection, which causes renal dysfunction known as sepsis-associated acute kidney injury (S-AKI). Ferroptosis is a form of lipid peroxidation dependent on iron and reactive oxygen species that differs from other forms of programmed cell death at the morphological and biochemical levels. Andrographolide (AG), a natural diterpenoid lactone compound extracted from Andrographis paniculata, has been shown to have therapeutic effects in kidney disease. In this study, we investigated the novel mechanism by which AG attenuates septic acute kidney injury by inhibiting ferroptosis in renal tubular epithelial cells (HK-2) through the Nrf2/FSP1 pathway. Cecum ligation and puncture (CLP)-induced septic rats and lipopolysaccharide (LPS)-induced HK-2 cells were used for in vivo and in vitro experiments. Firstly, in septic rats and HK-2 cells, AG effectively decreased the levels of kidney injury indicators, including blood creatinine, urea nitrogen, and markers of kidney injury such as neutrophil gelatinase-associated lipid transport protein and kidney injury molecule-1 (KIM-1). In addition, AG prevented ferroptotosis, by avoiding the accumulation of iron and lipid peroxidation, and an increase in SLC7A11 and GPX4 in AG-treated HK-2 cells. Furthermore, AG attenuated mitochondrial damage, including mitochondrial swelling, outer membrane rupture, and a reduction in mitochondrial cristae in LPS-treated HK-2 cells. Ferrostatin-1 (Fer-1), a ferroptosis inhibitor, significantly inhibited LPS-induced ferroptosis in HK-2 cells. Importantly, our results confirm that Nrf2/FSP1 is an important pathway for ferroptosis resistance. Nrf2 siRNA hindered the effect of AG in attenuating acute kidney injury and inhibiting ferroptosis. These findings demonstrate that Nrf2/FSP1-mediated HK-2 ferroptosis is associated with AG, alleviates septic acute kidney injury, and indicates a novel avenue for therapeutic interventions in the treatment of acute kidney injury in sepsis.

Anti-metastatic effects of AGS-30 on breast cancer through the inhibition of M2-like macrophage polarization.

AGS-30, a new andrographolide derivative, showed significant anticancer and anti-angiogenic characteristics. However, its role in controlling macrophage polarization and tumor immune response is unknown. Thus, the main goals of this study are to investigate how AGS-30 regulates macrophage polarization and how it suppresses breast cancer metastasis. AGS-30 inhibited IL-4 and IL-13-induced RAW 264.7 and THP-1 macrophages into M2-like phenotype. However, AGS-30 did not affect the LPS and IFN-γ-induced polarization of M1-like macrophages. AGS-30 reduced the mRNA expressions of CD206, Arg-1, Fizz-1, Ym-1, VEGF, IL-10, MMP2, and MMP9 in M2-like macrophages in a concentration-dependent manner. In contrast, andrographolide treatment at 5 µM did not affect M1-like and M2-like macrophage polarization. The conditioned medium from M2-like macrophages increased 4T1 breast cancer cell migration and invasion, whereas AGS-30 inhibited these effects. In the 4T1 breast tumor xenograft mice, the tumor volume and weight were reduced without affecting body weight after receiving AGS-30. AGS-30 treatment also reduced lung and liver metastasis, with reduced STAT6, CD31, VEGF, and Ki67 protein expressions. Moreover, the tumors had considerably fewer M2-like macrophages and Arg-1 expression, but the proportion of M1-like macrophages and iNOS expression increased after AGS-30 treatment. Same results were found in the tail vein metastasis model. In conclusion, this study shows that AGS-30 inhibits breast cancer growth and metastasis, probably through inhibiting M2-like macrophage polarization. Our findings suggest that AGS-30 may be a potential immunotherapeutic alternative for metastatic breast cancer.

Andrographolide sensitizes KRAS-mutant colorectal cancer cells to cetuximab by inhibiting the EGFR/AKT and PDGFRß/AKT signaling pathways.

BACKGROUND: Cetuximab, an inhibitor targeting EGFR, is widely applied in clinical management of colorectal cancer (CRC). Nevertheless, drug resistance induced by KRAS-mutations limits cetuximab's anti-cancer effectiveness. Furthermore, the persistent activation of EGFR-independent AKT is another significant factor in cetuximab resistance. Nevertheless, the mechanism that EGFR-independent AKT drives cetuximab resistance remains unclear. Thus, highlighting the need to optimize therapies to overcome cetuximab resistance and also to explore the underlying mechanism. PURPOSE: This work aimed to investigate whether and how andrographolide enhance the therapeutic efficacy of cetuximab in KRAS-mutant CRC cells by modulating AKT. METHODS: The viabilities of CRC cell lines were analyzed by CCK-8. The intracellular proteins phosphorylation levels were investigated by Human Phospho-kinase Antibody Array analysis. Knockdown and transfection of PDGFRß were used to evaluate the role of andrographolide on PDGFRß. The western blotting was used to investigate Wnt/ß-catenin pathways, PI3K/AKT, and EMT in KRAS-mutant CRC cells. The animal models including subcutaneous tumor and lung metastasis were performed to assess tumor response to therapy in vivo. RESULTS: Andrographolide was demonstrated to decrease the expression of PI3K and AKT through targeting PDGFRß and EGFR, and it enhanced cetuximab effect on KRAS-mutant CRC cells by this mechanism. Meanwhile, andrographolide helped cetuximab to inhibit Wnt/ß-catenin, CRC cell migration and reduced Vimentin expression, while increasing that of E-cadherin. Lastly, co-treatment with cetuximab and andrographolide reduced the growth of KRAS-mutant tumors and pulmonary metastases in vivo. CONCLUSIONS: Our findings suggest that andrographolide can overcome the KRAS-mutant CRC cells' resistance to cetuximab through inhibiting the EGFR/PI3K/AKT and PDGFRß /AKT signaling pathways. This research provided a possible theory that andrographolide sensitizes KRAS-mutant tumor to EGFR TKI.

Andrographolide acts with dexamethasone to inhibit the growth of acute lymphoblastic leukemia CEM­C1 cells via the regulation of the autophagy­dependent PI3K/AKT/mTOR signaling pathway.

Acute lymphoblastic leukemia (ALL) is one of the most common malignant tumor types of the circulatory system. Dexamethasone (DEX) acts on the glucocorticoid (GC) receptor (GR) and is a first-line chemotherapy drug for ALL. However, long-term or high-dose applications of the drug can not only cause adverse reactions, such as osteoporosis and high blood pressure, but can also cause downregulation of GR and lead to drug resistance. In the present study, reverse transcription-quantitative PCR, western blotting and LysoTracker Red staining were used to observe the effects of DEX and andrographolide (AND; a botanical with antitumorigenic properties) combined treatment. It was found that AND enhanced the sensitivity of CEM-C1 cells, a GC-resistant cell line, to DEX, and synergistically upregulated GR both at the transcriptional and post-transcriptional level with DEX. The combination of AND with DEX synergistically alkalized lysosomal lumen and downregulated the expression of autophagy-related genes Beclin1 and microtubule-associated 1 protein light chain 3 (LC3), thereby inhibiting autophagy. Knocking down LC3 expression enhanced GR expression, suggesting that GR was regulated by autophagy. Furthermore, compared with the monotherapy group (AND or DEX in isolation), AND interacted with DEX to activate the autophagy-dependent PI3K/AKT/mTOR signaling pathway by enhancing the phosphorylation of PI3K, AKT and mTOR, thereby decreasing GR degradation and increasing the sensitivity of cells to GCs. In conclusion, the present study demonstrated that AND exhibited a synergistic anti-ALL effect with DEX via upregulation of GR, which was orchestrated by the autophagy-related PI3K/AKT/mTOR signaling pathway. The results of the present study therefore provided novel research avenues and strategies for the treatment of ALL.

Targeting autophagy to counteract neuroinflammation: A novel antidepressant strategy.

Depression is a common disease that affects physical and mental health and imposes a considerable burden on afflicted individuals and their families worldwide. Depression is associated with a high rate of disability and suicide. It causes a severe decline in productivity and quality of life. Unfortunately, the pathophysiological mechanisms underlying depression have not been fully elucidated, and the risk of its treatment is still presented. Studies have shown that the expression of autophagic markers in the brain and peripheral inflammatory mediators are dysregulated in depression. Autophagy-related genes regulate the level of autophagy and change the inflammatory response in depression. Depression is related to several aspects of immunity. The regulation of the immune system and inflammation by autophagy may lead to the development or deterioration of mental disorders. This review highlights the role of autophagy and neuroinflammation in the pathophysiology of depression, sumaries the autophagy-targeting small moleculars, and discusses a novel therapeutic strategy based on anti-inflammatory mechanisms that target autophagy to treat the disease.

Andrographolide reverts multidrug resistance in KBChR 8-5 cells through AKT signaling pathway.

Multidrug resistance (MDR) is a major obstacle in cancer chemotherapy. P-glycoprotein (P-gp) one of the ATP-binding cassette (ABC) transporters plays an important role in MDR. In this study, we examined the sensitizing property of andrographolide (Andro) to reverse MDR in the drug-resistant KBChR 8-5 cells. Andro exhibited increased cytotoxicity in a concentration-dependent manner in the P-gp overexpressing KBChR 8-5 cells. Furthermore, Andro showed synergistic interactions with PTX and DOX in this drug-resistant cells. Andro co-administration enhanced PTX- and DOX-induced cytotoxicity and reduced cell proliferation in the MDR cancer cells. Moreover, reactive oxygen species (ROS) were elevated with a decrease in the mitochondrial membrane potential (MMP) during Andro and chemotherapeutic drugs combination treatment in the drug-resistant cells. Furthermore, Andro and PTX-induced cell cycle arrest was observed in the drug-resistant cell. We also noticed that the expression of ABCB1 and AKT were downregulated during Andro (4 µM) treatment. Furthermore, Andro treatment enhanced the expression of caspase 3 and caspase 9 in the combinational groups that support the enhanced apoptotic cell death in drug-resistant cancer cells. Therefore, the results reveal that Andro plays a role in the reversal of P-gp-mediated MDR in KBChR 8-5 cells which might be due to regulating ABCB1/AKT signaling pathway.

Development and Optimization of Andrographis paniculata Extract-Loaded Self-Microemulsifying Drug Delivery System Using Experimental Design Model.

The objectives of this study were to develop an optimized formulation for an Andrographis paniculata extract (AGPE)-loaded self-microemulsifying drug delivery system (SMEDDS) using an experimental design and evaluate the characteristics of the developed SMEDDS. The solubility of andrographolide (AGP) in various solvents was investigated. The pseudo-ternary phase was constructed to provide an optimal range for each component to form microemulsions (MEs). The formulation was optimized using an I-optimal design mixture type, where the physical stability, droplet size, polydispersity index, and zeta potential were examined. Soft capsules of the optimized AGPE-loaded SMEDDS were manufactured. The dissolution and ex vivo membrane permeation were studied. Oleic acid, Tween® 80, and PEG 400 were the best solubilizers for AGP. The promising surfactant to co-surfactant ratio to generate ME was 3:1. The optimized SMEDDS contained 68.998% Tween® 80, with 13.257% oleic acid and 17.745% PEG 400. The assayed content of AGP, uniformity of dosage unit, and stability complied with the expected specifications. The dissolution and membrane permeability of AGPE-loaded SMEDDS was significantly improved from the A. paniculata extract (p < 0.05). All in all, the developed optimized AGPE-loaded SMEDDS was proven to contain optimal composition and AGP content where a stable ME could spontaneously be formed with enhanced delivery efficacy.

An Integrated Network Pharmacology and RNA-seq Approach for Exploring the Protective Effect of Andrographolide in Doxorubicin-Induced Cardiotoxicity.

PURPOSE: Doxorubicin (Dox) is clinically limited due to its dose-dependent cardiotoxicity. Andrographolide (Andro) has been confirmed to exert cardiovascular protective activities. This study aimed to investigate protective effects of Andro in Dox-induced cardiotoxicity (DIC). METHODS: The cardiotoxicity models were induced by Dox in vitro and in vivo. The viability and apoptosis of H9c2 cells and the myocardial function of c57BL/6 mice were accessed with and without Andro pretreatment. Network pharmacology and RNA-seq were employed to explore the mechanism of Andro in DIC. The protein levels of Bax, Bcl2, NLRP3, Caspase-1 p20, and IL-1ß were qualified as well. RESULTS: In vitro, Dox facilitated the downregulation of cell viability and upregulation of cell apoptosis, after Andro pretreatment, the above symptoms were remarkably reversed. In vivo, Andro could alleviate Dox-induced cardiac dysfunction and apoptosis, manifesting elevation of LVPWs, LVPWd, EF% and FS%, suppression of CK, CK-MB, c-Tnl and LDH, and inhibition of TUNEL-positive cells. Using network pharmacology, we collected and visualized 108 co-targets of Andro and DIC, which were associated with apoptosis, PI3K-AKT signaling pathway, and others. RNA-seq identified 276 differentially expressed genes, which were enriched in response to oxidative stress, protein phosphorylation, and others. Both network pharmacology and RNA-seq analysis identified Tap1 and Timp1 as key targets of Andro in DIC. RT-QPCR validation confirmed that the mRNA levels of Tap1 and Timp1 were consistent with the sequenced results. Moreover, the high expression of NLRP3, Caspase-1 p20, and IL-1ß in the Dox group was reduced by Andro. CONCLUSIONS: Andro could attenuate DIC through suppression of Tap1 and Timp1 and inhibition of NLRP3 inflammasome activation, serving as a promising cardioprotective drug.

In vitro and in vivo study of andrographolide nanoparticles for the treatment of Mycoplasma pneumoniae pneumonia.

OBJECTIVE(S): The emergence of antibiotic resistance has led to suboptimal treatment outcomes for Mycoplasma pneumoniae pneumonia (MPP). Exploring naturally occurring drug components that are both effective against MPP and non-toxic may be a promising choice. This study aimed to investigate the therapeutic effect of andrographolide nanoparticles on pneumonia caused by Mycoplasma pneumoniae infection. METHODS: Andrographolide alginate-poloxamer nanoparticles (AND-ALG-POL/NPs) were obtained by wet medium grinding, and the characterization and in vitro release of the prepared andrographolide nanoparticles were examined by high performance liquid chromatography, particle size analyzer, zeta potential meter and transmission electron microscopy. The cytotoxicity and anti-inflammatory effects of AND-ALG-POL/NPs were evaluated in vitro by MP-infected lung epithelial cells BEAS-2B. Symptoms of pneumonia, total cell count, total protein content and inflammatory factor levels in BALF were assessed by MP-induced pneumonia in BALB/c mice treated with AND-ALG-POL/NPs, and histopathological studies were performed on lung tissues from experimental animals. RESULTS: The results showed that the prepared AND-ALG-POL/NPs were homogeneous spherical with a diameter of 180 ± 23 nm, a zeta potential of (-14.4 ± 2.1) mV, an average encapsulation rate of 87.74 ± 0.87 %, and an average drug loading of 13.17 ± 0.54 %. AND-ALG-POL/NPs were capable of slow release in vitro and showed significant inhibitory ability against MP (P < 0.001). However, AND-ALG-POL/NPs were not cytotoxic to normal cells and alleviated MP infection-induced apoptosis and elevated inflammatory factors. In the in vivo experiments, AND-ALG-POL/NPs alleviated the symptoms of pneumonia in MPP mice, reduced the abnormally elevated total cell count, total protein content and inflammatory factor levels in BALF, and alleviated lung tissue edema, inflammatory cell infiltration and apoptosis (P < 0.001). Meanwhile, the therapeutic effects of AND-ALG-POL/NPs on MPP were similar to those of azithromycin (AZM) and higher than those of andrographolide (AND) free monotherapy (P < 0.001). CONCLUSION: In summary, the prepared AND-ALG-POL/NPs can effectively inhibit MPP in vitro and in vivo, and the effect is similar to that of AZM. Therefore, AND- ALG - POL/NPs have the potential to replace AZM as a potential drug for the treatment of MPP.

Sustained release polymer and surfactant based solid dispersion of andrographolide exhibited improved solubility, dissolution, pharmacokinetics, and pharmacological activity.

Andrographolide (AD) is a potent natural product with a wide range of pharmacological activities. However, it has low oral bioavailability due to poor solubility and dissolution rate. Solid dispersion (SD) is a promising technique to improve the solubility and dissolution rate of such molecules. In this study, SD formulation of AD was prepared using Kollidon-SR (KSR) and Poloxamer-407 (P-407) as carriers. SD was prepared using the solvent evaporation method and evaluated for the modulation of solubility of AD. The developed SD formulation was characterized using scanning electron microscopy (SEM), differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD), and Fourier transform infrared spectroscopy (FTIR). Further, dissolution rate, yield, drug content, stability, flowability, and pharmacokinetic profile of SD were evaluated. The compatibility of SD with the Caco-2 cells and its impact on the P-glycoprotein (P-gp) mediated efflux was also investigated. Furthermore, carrageenan-induced paw edema, and adjuvant-induced arthritic model were used to evaluate the efficacy of SD. The results showed that SD3 (AD + KSR + P-407, 1:6:8) exhibited the highest solubility and dissolution rate, and significantly improved pharmacokinetic profile compared to native AD. SD3 was found to be stable during storage and displayed excellent yield, drug content, and flowability. This formulation was found to be compatible with the Caco-2 cells and retarded the efflux of P-gp substrate. SD3 also demonstrated substantially better efficacy than native AD in terms of paw edema inhibition (carrageenan-induced paw edema, Wistar rats), and overall improvement of disease condition (in terms of paw edema, arthritic score, AST, ALT, cytokines, radiological changes, and histopathology) in arthritic Wistar rats. In conclusion, SD3 exhibited improved solubility, dissolution rate, pharmacokinetic profile, and pharmacological activity than native AD.

Molecular docking and dynamics simulations revealed the potential inhibitory activity of honey-iQfood ingredients against GSK-3ß and CDK5 protein targets for brain health.

Honey-iQfood is an herbal supplement made of a mixture of polyherbal extracts and wild honey. The mixture is traditionally claimed to improve various conditions related to brain cells and functions including dementia and Alzheimer's disease. Glycogen synthase kinase-3 beta (GSK-3ß) and cyclin-dependent kinase 5 (CDK5) have been identified as being involved in the pathological hyperphosphorylation of tau proteins, which leads to the formation of neurofibrillary tangles and causes Alzheimer's disease. Therefore, this study was conducted to confirm the traditional claims by detection of active compounds, namely curcumin, gallic acid, catechin, rosmarinic acid, and andrographolide in the raw materials of Honey-iQfood through HPLC analysis, molecular docking, and dynamic simulations. Two potential compounds, andrographolide, and rosmarinic acid, produced the best binding affinities following the molecular docking of the active compounds against the GSK-3ß and CDK5 targets. Andrographolide binds with GSK-3ß at -8.2 kcal/mol, whereas rosmarinic acid binds to CDK5 targets at -8.6 kcal/mol. Molecular dynamics was further carried out to confirm the docking results and clarify their dynamic properties such as RMSD, RMSF, rGyr, SASA, PSA, and binding free energy. CDK5-andrographolide complexes had the best MM-GBSA score (-83.63 kcal/mol) compared to other complexes, indicating the better interaction profile and stability of the complex. These findings warrant further research into andrographolide and rosmarinic acid as efficient inhibitors of tau protein hyperphosphorylation to verify their therapeutic potential in brain-related illnesses.Communicated by Ramaswamy H. Sarma.

Design, synthesis, and anti-cancer evaluation of C-14 arylcarbamate derivatives of andrographolide.

In this study, we explored a concise and mild synthetic route to produce novel C-14 arylcarbamate derivatives of andrographolide, a known anti-inflammatory and anticancer natural product. Upon assessing their anti-cancer efficacy against pancreatic ductal adenocarcinoma (PDAC) cells, some derivatives showed stronger cytotoxicity against PANC-1 cells than andrographolide. In addition, we demonstrated one derivative, compound 3m, effectively reduced the expression of oncogenic p53 mutant proteins (p53R273H and p53R248W), proliferation, and migration in PDAC lines, PANC-1 and MIA PaCa-2. Accordingly, the novel derivative holds promise as an anti-cancer agent against pancreatic cancer. In summary, our study broadens the derivative library of andrographolide and develops an arylcarbamate derivative of andrographolide with promising anticancer activity against PDAC.

Andrographolide protect against lipopolysacharides induced vascular endothelium dysfunction by abrogation of oxidative stress and chronic inflammation in Sprague-Dawley rats.

The development of heart disease involves interconnected factors such as oxidative stress, inflammation, and vascular dysfunction. Andrographolide (AG), known for its potent antioxidant and anti-inflammatory properties, has the potential to counteract lipopolysaccharides (LPS)-induced endothelial dysfunction by reducing oxidative stress and inflammation. Our research aimed to investigate the effects of AG on alleviating vascular endothelium dysfunction, oxidative stress, and inflammation in an experimental model induced by LPS. To create chronic vascular endothelium dysfunction, inflammation, and oxidative stress, rats received weekly injections of LPS via their tail vein over a 6-week period. The study evaluated the therapeutic effects of orally administered AG (50 mg/kg/day) on diseased conditions. We conducted aortic histology and measured nitric oxide (NO) thresholds, superoxide dismutase (SOD) activity, constitutive nitric oxide (cNOS) activity, and inducible nitric oxide (iNOS) levels, alongside several inflammatory biomarkers. To evaluate endothelial dysfunction, we assessed endothelium-dependent and endothelium-independent vasorelaxation in aortas through histopathological and various immunoassays examinations. Vascular Endothelial inflammatory activity was consequently enhanced in LPS groups animals when compared to normal control, also endothelial performance were dependently improved by AG therapy. IL-1ß and tumors necrosis factor levels in the aorta decreased in a dose-dependent manner after exogenous AG delivery to LPS-treated rats. However, in current research work aortic SOD activity, NO levels, and cNOS activity increased, whereas aortic malondialdehyde levels and iNOS activity decreased after the AG treatment. These findings suggest that long-term AG therapy could be considered as a potential therapy to avoid vascular endothelial dysfunction and major nonobstructive coronary artery disease.

Evaluation of natural products from virtual screenings as SARS-CoV-2 main protease inhibitors using combinational experiments.

Recently, andrographolide, kaempferol, maslinic acid, rutin, and schaftoside have been identified as potent SARS-CoV-2 main protease (Mpro) inhibitors via molecular docking studies. However, no comprehensive in vitro testing of these compounds against Mpro has been conducted. In this study, we rigorously evaluated the in vitro inhibition of Mpro by these compounds using combinational experiments, including fluorescence resonance energy transfer (FRET), fluorescence polarization (FP), and dimerization-dependent red fluorescent protein (ddRFP) assays. Our data revealed that these compounds are not Mpro inhibitors based on the results from a set of in vitro assays. These results suggest that an efficient combination of a molecular docking approach and an experimental assay is essential for the discovery of Mpro inhibitors in the future.