Synthetic Tetraploid of Oncidium crispum Lodd. (Orchidaceae).

In ornamental plants, artificial polyploidization has enabled the creation of new cultivars. Due to their high commercial value in the international flower market and their ornamental characteristics, such as the shape, size, color, and durability of their flower, orchids have received great attention in studies of artificial polyploidization. Here we described the protocol used for polyploid induction in Oncidium crispum, an epiphyte species native of southeastern Brazil, of great ornamental interest and widely sold in flower shops. The species stands out for having inflorescence with large flowers, brown in color with yellow spots. In addition, O. crispum has great potential for use in genetic improvement programs since the species is widely used in interspecific crosses. Closed capsules containing mature O. crispum seeds were subjected to running sterilized water for 10 min and then to a 1.5% sodium hypochlorite solution for 10 min. Small portions of seeds were introduced into 50 mL of water-soluble fertilizer with macro- and micronutrients (B>M) plus 0.7% agar. Explants originating from seeds previously in vitro germinated were submitted to 0.05% and 0.1% of colchicine for 4 days and 8 days. Flow cytometry and chromosome counts confirmed that the protocol successfully produced synthetic polyploid plants.

The alleviatory effects of koumine on MSU-induced gouty arthritis via the TLR4/NF-κB/NLRP3 pathway.

The aim of this study was to validate the preventive effects of koumine (KM), a monoterpene indole alkaloid, on gouty arthritis (GA) and to explore its possible mechanisms. C57BL/6 mice were intraperitoneally administered KM (0.8, 2.4 or 7.2 mg/kg), colchicine (3.0 mg/kg) or sterile saline. One hour later, a monosodium urate (MSU) suspension was injected into the right hind paws of the mice to establish an acute gout model. Inflammation symptoms were evaluated at 0, 3, 6, 12 and 24 h, and the mechanical withdrawal threshold was evaluated at 0, 6 and 24 h. After 24 h, the mice were euthanized, and the joint tissue, kidney and blood were collected for subsequent experiments. Histological examination and antioxidant enzyme, kidney index and serum uric acid (UA) measurements were taken. The expression levels of the signalling pathway components were determined. KM effectively alleviated the symptoms of redness, swelling and pain; counteracted inflammatory cell infiltration; and increased antioxidant enzyme levels, reduced kidney index and serum UA levels through regulating UA excretion in MSU-induced mice. The expression of toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB)/nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) signalling pathway proteins and mRNA were reduced in the KM group. These results suggest that KM may be effective in alleviating GA through the TLR4/NF-κB/NLRP3 pathway.

In vitro induction of tetraploids and their phenotypic and transcriptome analysis in Glehnia littoralis.

BACKGROUND: Glehnia littoralis is a medicinal and edible plant species having commercial value and has several hundred years of cultivation history. Polyploid breeding is one of the most important and fastest ways to generate novel varieties. To obtain tetraploids of G. littoralis in vitro, colchicine treatment was given to the seeds and then were screened based on morphology, flow cytometry, and root tip pressing assays. Furthermore, transcriptome analysis was performed to identity the differentially expressed genes associated with phenotypic changes in tetraploid G. littoralis. RESULTS: The results showed that 0.05% (w/v) colchicine treatment for 48 h was effective in inducing tetraploids in G. littoralis. The tetraploid G. littoralis (2n = 4x = 44) was superior in leaf area, leaf thickness, petiole diameter, SPAD value (Chl SPAD), stomatal size, epidermal tissues thickness, palisade tissues thickness, and spongy tissues thickness to the diploid ones, while the stomatal density of tetraploids was significantly lower. Transcriptome sequencing revealed, a total of 1336 differentially expressed genes (DEGs) between tetraploids and diploids. Chromosome doubling may lead to DNA content change and gene dosage effect, which directly affects changes in quantitative traits, with changes such as increased chlorophyll content, larger stomata and thicker tissue of leaves. Several up-regulated DEGs were found related to growth and development in tetraploid G. littoralis such as CKI, PPDK, hisD and MDP1. KEGG pathway enrichment analyses showed that most of DEGs were enriched in metabolic pathways. CONCLUSIONS: This is the first report of the successful induction of tetraploids in G. littoralis. The information presented in this study facilitate breeding programs and molecular breeding of G. littoralis varieties.

Exploring the mechanism of transformation in Acacia nilotica (Linn.) triggered by colchicine seed treatment.

BACKGROUND: Acacia nilotica Linn. is a widely distributed tree known for its applications in post-harvest and medicinal horticulture. However, its seed-based growth is relatively slow. Seed is a vital component for the propagation of A. nilotica due to its cost-effectiveness, genetic diversity, and ease of handling. Colchicine, commonly used for polyploidy induction in plants, may act as a pollutant at elevated levels. Its optimal concentration for Acacia nilotica's improved growth and development has not yet been determined, and the precise mechanism underlying this phenomenon has not been established. Therefore, this study investigated the impact of optimized colchicine (0.07%) seed treatment on A. nilotica's morphological, anatomical, physiological, fluorescent, and biochemical attributes under controlled conditions, comparing it with a control. RESULTS: Colchicine seed treatment significantly improved various plant attributes compared to control. This included increased shoot length (84.6%), root length (53.5%), shoot fresh weight (59.1%), root fresh weight (42.8%), shoot dry weight (51.5%), root dry weight (40%), fresh biomass (23.6%), stomatal size (35.9%), stomatal density (41.7%), stomatal index (51.2%), leaf thickness (11 times), leaf angle (2.4 times), photosynthetic rate (40%), water use efficiency (2.2 times), substomatal CO2 (36.6%), quantum yield of photosystem II (13.1%), proton flux (3.1 times), proton conductivity (2.3 times), linear electron flow (46.7%), enzymatic activities of catalase (25%), superoxide dismutase (33%), peroxidase (13.5%), and ascorbate peroxidase (28%), 2,2-diphenyl-1-picrylhydrazyl-radical scavenging activities(23%), total antioxidant capacity (59%), total phenolic (23%), and flavonoid content (37%) with less number of days to 80% germination (57.1%), transpiration rate (53.9%), stomatal conductance (67.1%), non-photochemical quenching (82.8%), non-regulatory energy dissipation (24.3%), and H2O2 (25%) and O-2 levels (30%). CONCLUSION: These findings elucidate the intricate mechanism behind the morphological, anatomical, physiological, fluorescent, and biochemical transformative effects of colchicine seed treatment on Acacia nilotica Linn. and offer valuable insights for quick production of A. nilotica's plants with modification and enhancement from seeds through an eco-friendly approach.

Micro tensile tester measurement of biomechanical properties and adhesion force of microtubule-polymerization-inhibited cancer cells.

Both mechanical and adhesion properties of cancer cells are complex and reciprocally related to migration, invasion, and metastasis with large cell deformation. Therefore, we evaluated these properties for human cervical cancer cells (HeLa) simultaneously using our previously developed micro tensile tester system. For efficient evaluation, we developed image analysis software to modify the system. The software can analyze the tensile force in real time. The modified system can evaluate the tensile stiffness of cells to which a large deformation is applied, also evaluate the adhesion strength of cancer cells that adhered to a culture substrate and were cultured for several days with their adhesion maturation. We used the modified system to simultaneously evaluate the stiffness of the cancer cells to which a large deformation was applied and their adhesion strength. The obtained results revealed that the middle phase of tensile stiffness and adhesion force of the microtubule-depolymerized group treated with colchicine (an anti-cancer drug) (stiffness, 13.4 ± 7.5 nN/%; adhesion force, 460.6 ± 258.2 nN) were over two times larger than those of the control group (stiffness, 5.0 ± 3.5 nN/%; adhesion force, 168.2 ± 98.0 nN). Additionally, the same trend was confirmed with the detailed evaluation of cell surface stiffness using an atomic force microscope. Confocal fluorescence microscope observations showed that the stress fibers (SFs) of colchicine-treated cells were aligned in the same direction, and focal adhesions (FAs) of the cells developed around both ends of the SFs and aligned parallel to the developed direction of the SFs. There was a possibility that the microtubule depolymerization by the colchicine treatment induced the development of SFs and FAs and subsequently caused an increment of cell stiffness and adhesion force. From the above results, we concluded the modified system would be applicable to cancer detection and anti-cancer drug efficacy tests.

Low-Dose Colchicine Ameliorates Doxorubicin Cardiotoxicity Via Promoting Autolysosome Degradation.

BACKGROUND: The only clinically approved drug that reduces doxorubicin cardiotoxicity is dexrazoxane, but its application is limited due to the risk of secondary malignancies. So, exploring alternative effective molecules to attenuate its cardiotoxicity is crucial. Colchicine is a safe and well-tolerated drug that helps reduce the production of reactive oxygen species. High doses of colchicine have been reported to block the fusion of autophagosomes and lysosomes in cancer cells. However, the impact of colchicine on the autophagy activity within cardiomyocytes remains inadequately elucidated. Recent studies have highlighted the beneficial effects of colchicine on patients with pericarditis, postprocedural atrial fibrillation, and coronary artery disease. It remains ambiguous how colchicine regulates autophagic flux in doxorubicin-induced heart failure. METHODS AND RESULTS: Doxorubicin was administered to establish models of heart failure both in vivo and in vitro. Prior studies have reported that doxorubicin impeded the breakdown of autophagic vacuoles, resulting in damaged mitochondria and the accumulation of reactive oxygen species. Following the administration of a low dose of colchicine (0.1 mg/kg, daily), significant improvements were observed in heart function (left ventricular ejection fraction: doxorubicin group versus treatment group=43.75%±3.614% versus 57.07%±2.968%, P=0.0373). In terms of mechanism, a low dose of colchicine facilitated the degradation of autolysosomes, thereby mitigating doxorubicin-induced cardiotoxicity. CONCLUSIONS: Our research has shown that a low dose of colchicine is pivotal in restoring the autophagy activity, thereby attenuating the cardiotoxicity induced by doxorubicin. Consequently, colchicine emerges as a promising therapeutic candidate to improve doxorubicin cardiotoxicity.

Modification of the phenyl ring B of phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates by pyridinyl moiety leads to novel antimitotics targeting the colchicine-binding site.

A series of 8 novel pyridinyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates (PYRIB-SOs) were designed, prepared and evaluated for their mechanism of action. PYRIB-SOs were found to have antiproliferative activity in the nanomolar to submicromolar range on several breast cancer cell lines. Moreover, subsequent biofunctional assays indicated that the most potent PYRIB-SOs 1-3 act as antimitotics binding to the colchicine-binding site (C-BS) of α, ß-tubulin and that they arrest the cell cycle progression in the G2/M phase. Microtubule immunofluorescence and tubulin polymerisation assay confirm that they disrupt the cytoskeleton through inhibition of tubulin polymerisation as observed with microtubule-destabilising agents. They also show good overall theoretical physicochemical, pharmacokinetic and druglike properties. Overall, these results show that PYRIB-SOs is a new family of promising antimitotics to be further studied in vivo for biopharmaceutical and pharmacodynamic evaluations.

Anticancer potential of thiocolchicoside and lauric acid loaded chitosan nanogel against oral cancer cell lines: a comprehensive study.

The present study explored the anticancer activity of a Chitosan-based nanogel incorporating thiocolchicoside and lauric acid (CTL) against oral cancer cell lines (KB-1). Cell viability, AO/EtBr dual staining and Cell cycle analysis were done to evaluate the impact of CTL nanogel on oral cancer cells. Real-time PCR was performed to analyze proapoptotic and antiapoptotic gene expression in CTL-treated KB-1 cells. Further, molecular docking analysis was conducted to explore the interaction of our key ingredient, thiocolchicoside and its binding affinities. The CTL nanogel demonstrated potent anticancer activity by inhibiting oral cancer cell proliferation and inducing cell cycle arrest in cancer cells. Gene expression analysis indicated alterations in Bax and Bcl-2 genes; CTL nanogel treatment increased Bax mRNA expression and inhibited the Bcl-2 mRNA expression, which showed potential mechanisms of the CTL nanogel's anticancer action. It was found that thiocolchicoside can stabilize the protein's function or restore it as a tumour suppressor. The CTL nanogel exhibited excellent cytotoxicity and potent anticancer effects, making it a potential candidate for non-toxic chemotherapy in cancer nanomedicine. Furthermore, the nanogel's ability to modulate proapoptotic gene expression highlights its potential for targeted cancer therapy. This research contributes to the growing interest in Chitosan-based nanogels and their potential applications in cancer treatment.

A polarity-responsive lysosomes-nucleus translocation probe for the dual-emissive visualization of cell apoptosis.

Visualization of cell apoptosis is a critical task playing central roles in the fundamental studies in biology, pathology, and biomedicine. Dual-emissive fluorescent probes are desired molecular tools for study on apoptosis, which however were rarely reported. Herein, utilizing the polarity differences between lysosomes and nucleus, a translocation type of fluorescent probe (NA-S) was developed for the dual-color visualization of cell apoptosis. NA-S was designed to be polarity sensitive, bearing alkalescence group, and with DNA affinity. In living cells, NA-S targeted the lysosomes to give blue fluorescence, which translocated into the nucleus during cell apoptosis to give green emission. Thereby, the cell apoptosis could be visualized with NA-S in dual-emissive manner. With the unique probe, the cell apoptosis induced by oxidative stress, UV irradiation, rotenone, colchicine, and paclitaxel have been successfully visualized.

Atrial Fibrosis and Inflammation in Postoperative Atrial Fibrillation: Comparative Effects of Amiodarone, Colchicine, or Exosomes.

BACKGROUND: Extracellular vesicles (EVs) isolated from human heart-derived cells have shown promise in suppressing inflammation and fibroblast proliferation. However, their precise benefits in atrial fibrillation (AF) prevention and the role of their antifibrotic/anti-inflammatory properties remain unclear. OBJECTIVES: The purpose of this study was to conduct a head-to-head comparison of antiarrhythmic strategies to prevent postoperative AF using a rat model of sterile pericarditis. Specifically, we aimed to assess the efficacy of amiodarone (a classic antiarrhythmic drug), colchicine (an anti-inflammatory agent), and EVs derived from human heart-derived cells, which possess anti-inflammatory and antifibrotic properties, on AF induction, inflammation, and fibrosis progression. METHODS: Heart-derived cells were cultured from human atrial appendages under serum-free xenogen-free conditions. Middle-aged Sprague Dawley rats were randomized into different groups, including sham operation, sterile pericarditis with amiodarone treatment, sterile pericarditis with colchicine treatment (2 dose levels), and sterile pericarditis with intra-atrial injection of EVs or vehicle. Invasive electrophysiological testing was performed 3 days after surgery before sacrifice. RESULTS: Sterile pericarditis increased the likelihood of inducing AF. Colchicine and EVs exhibited anti-inflammatory effects, but only EV treatment significantly reduced AF probability, whereas colchicine showed a positive trend without statistical significance. EVs and high-dose colchicine reduced atrial fibrosis by 46% ± 2% and 26% ± 2%, respectively. Amiodarone prevented AF induction but had no effect on inflammation or fibrosis. CONCLUSIONS: In this study, both amiodarone and EVs prevented AF, whereas treatment with colchicine was ineffective. The additional anti-inflammatory and antifibrotic effects of EVs suggest their potential as a comprehensive therapeutic approach for AF prevention, surpassing the effects of amiodarone or colchicine.

Structure-based design and synthesis of BML284 derivatives: A novel class of colchicine-site noncovalent tubulin degradation agents.

Our previous studies have demonstrated that BML284 is a colchicine-site tubulin degradation agent. To improve its antiproliferative properties, 45 derivatives or analogs of BML284 were designed and synthesized based on the cocrystal structure of BML284 and tubulin. Among them, 5i was the most potent derivative, with IC50 values ranging from 0.02 to 0.05 µM against the five tested tumor cell lines. Structure-activity relationship studies verified that the N1 atom of the pyrimidine ring was the key functional group for its tubulin degradation ability. The 5i-tubulin cocrystal complex revealed that the binding pattern of 5i to tubulin is similar to that of BML284. However, replacing the benzodioxole ring with an indole ring strengthened the hydrogen bond formed by the 2-amino group with E198, which improved the antiproliferative activity of 5i. Compound 5i effectively suppressed tumor growth at an intravenous dose of 40 mg/kg (every 2 days) in paclitaxel sensitive A2780S and paclitaxel resistant A2780T ovarian xenograft models, with tumor growth inhibition values of 79.4% and 82.0%, respectively, without apparent side effects, showing its potential to overcome multidrug resistance. This study provided a successful example of crystal structure-guided discovery of 5i as a colchicine-targeted tubulin degradation agent, expanding the scope of targeted protein degradation.

Autotetraploid Induction of Three A-Genome Wild Peanut Species, Arachis cardenasii, A. correntina, and A. diogoi.

A-genome Arachis species (AA; 2n = 2x = 20) are commonly used as secondary germplasm sources in cultivated peanut breeding, Arachis hypogaea L. (AABB; 2n = 4x = 40), for the introgression of various biotic and abiotic stress resistance genes. Genome doubling is critical to overcoming the hybridization barrier of infertility that arises from ploidy-level differences between wild germplasm and cultivated peanuts. To develop improved genome doubling methods, four trials of various concentrations of the mitotic inhibitor treatments colchicine, oryzalin, and trifluralin were tested on the seedlings and seeds of three A-genome species, A. cardenasii, A. correntina, and A. diogoi. A total of 494 seeds/seedlings were treated in the present four trials, with trials 1 to 3 including different concentrations of the three chemical treatments on seedlings, and trial 4 focusing on the treatment period of 5 mM colchicine solution treatment of seeds. A small number of tetraploids were produced from the colchicine and oryzalin gel treatments of seedlings, but all these tetraploid seedlings reverted to diploid or mixoploid states within six months of treatment. In contrast, the 6-h colchicine solution treatment of seeds showed the highest tetraploid conversion rate (6-13% of total treated seeds or 25-40% of surviving seedlings), and the tetraploid plants were repeatedly tested as stable tetraploids. In addition, visibly and statistically larger leaves and flowers were produced by the tetraploid versions of these three species compared to their diploid versions. As a result, stable tetraploid plants of each A-genome species were produced, and a 5 mM colchicine seed treatment is recommended for A-genome and related wild Arachis species genome doubling.

Structure-based approaches for the design of 6-aryl-1-(3,4,5-trimethoxyphenyl)-1H-benzo[d][1,2,3]triazoles as tubulin polymerization inhibitors.

The colchicine binding site on tubulin has been widely acknowledged as an attractive target for anticancer drug exploitation. Here, we reported the structural optimization of the lead compound 4, which was proved in our previous work as a colchicine binding site inhibitor (CBSI). Based on docking researches for the active binding conformation of compound 4, a series of novel 6-aryl-1-(3,4,5-trimethoxyphenyl)-1H-benzo[d][1,2,3]triazole derivatives (9a-9x) were developed by replacing a CH group in the 1H-benzo[d]imidazole skeleton of compound 4 with a nitrogen atom as a hydrogen bond acceptor. Among them, compound 9a showed the strongest antiproliferative activity with IC50 values ranging from 14 to 45 nM against three human cancer cell lines (MCF-7, SGC-7901 and A549), lower than that of compound 4. Mechanistic studies indicated that compound 9a could inhibit tubulin polymerization, destroy the microtubule skeleton, block the cell cycle in G2/M phase, induce cancer cell apoptosis, prevent cancer cell migration and colony formation. Moreover, compound 9a significantly inhibited tumor growth in vivo without observable toxicity in the mice 4T1 xenograft tumor model. In conclusion, this report shows a successful case of the structure-based design approach of a potent tubulin polymerization inhibitor for cancer treatment.

Investigating the Promising Anticancer Activity of Cetuximab and Fenbendazole Combination as Dual CBS and VEGFR-2 Inhibitors and Endowed with Apoptotic Potential.

In this work, the cytotoxicity of monoclonal antibody (Cetuximab, Ce) and Fenbendazole (Fen), as well as their combination therapy were tested with the MTT assay. On the other side, Ce, Fen, and a combination between them were subjected to a colchicine-tubulin binding test, which was conducted and compared to Colchicine as a reference standard. Besides, Ce, Fen, and the combination of them were tested against the VEGFR-2 target receptor, compared to Sorafenib as the standard medication. Moreover, the qRT-PCR technique was used to investigate the levels of apoptotic genes (p53 and Bax) and anti-apoptotic gene (Bcl-2) as well. Also, the effect of Ce, Fen, and the combination of them on the level of ROS was studied. Furthermore, the cell cycle analysis and Annexin V apoptosis assay were carried out for Ce, Fen, and a combination of them. In addition, the molecular docking studies were used to describe the molecular levels of interactions for both (Fen and colchicine) or (Fen and sorafenib) within the binding pockets of the colchicine binding site (CBS) and vascular endothelial growth factor-2 receptor (VEGFR-2), respectively.

Network Pharmacology Analysis of the Therapeutic Potential of Colchicine in Acute Lung Injury.

Background: This study employed integrated network pharmacology approach to explore the mechanisms underlying the protective effect of colchicine against acute lung injury (ALI). Methods: We analyzed the expression profiles from 13 patients with sepsis-related ALI and 21 controls to identify differentially expressed genes and key modules. ALI-related genes were curated using databases such as DisGeNET, Therapeutic Target, and Comparative Toxicogenomics Database to curate ALI-related genes. Drug target fishing for colchicine was conducted using the DrugBank, BATMAN-TCM, STITCH, and SwissTargetPrediction. Potential drug-disease interactions were determined by intersecting ALI-associated genes with colchicine target genes. We performed comprehensive pathway and process enrichment analyses on these genes. A protein-protein interaction network was constructed, and topological analysis was executed. Additionally, an ALI mouse model was established to evaluate the effect of colchicine on CXCL12 and CXCR4 levels through western blot analysis. Results: Analysis revealed 23 potential colchicine-ALI interaction genes from the intersection of 253 ALI-associated genes and 389 colchicine targets. Functional enrichment analysis highlighted several inflammation-related pathways, such as cytokine-mediated signaling pathway, CXCR chemokine receptor binding, NF-kappa B signaling pathway, TNF signaling pathway, and IL-17 signaling pathway. The protein-protein interaction network demonstrated complex interactions for CXCL12 and CXCR4 among other candidate genes, with significant topological interaction degrees. In vivo studies showed that colchicine significantly reduced elevated CXCL12 and CXCR4 levels in ALI mice. Conclusion: Our findings suggest that colchicine's therapeutic effect on ALI might derive from its anti-inflammatory properties. Further research is needed to explore the specific mechanisms of colchicine's interaction with sepsis-induced ALI.

Genotoxic effects and mitosis aberrations of chromium (VI) on root cells of Vicia faba and its molecular docking analysis.

Like other heavy metals, Cr (VI) is a powerful carcinogen and mutagen agent. Its toxic effects on plants are well considered. In order to elucidate its adverse effects, the present work aims to study the mitosis aberrations of Cr (VI) on the Vicia faba root-cells and its molecular docking analysis to understand the genotoxicity mechanisms. In-vivo, Vicia faba plants were exposed to 50 and 100 µM Cr (VI) for 48 h. In-silico, molecular docking and molecular dynamics simulation were used to study the interactions between dichromate and tubulin tyrosine ligase T2R-TTL (PDBID: 5XIW) with reference to Colchicine (microtubule inhibitor). According to our results, Cr (VI) affects growth and cell division and also induces many mitosis aberrations such as chromosome sticking, anaphase/telophase bridges, lagging chromosomes and fragmentation during all phases of mitosis. On the one hand, Cr (VI) reduces mitotic index and promotes micronuclei induction. The in-silico results showed that dichromate establishes very strong bonds at the binding site of the tubulin tyrosine ligase T2R-TTL, with a binding affinity of -5.17 Kcal/Mol and an inhibition constant of 163.59 µM. These interactions are similar to those of colchicine with this protein, so dichromate could be a very potent inhibitor of this protein's activity. TTL plays a fundamental role in the tyrosination/detyrosination of tubulin, which is crucial to the regulation of the microtubule cytoskeleton. Its inhibition leads to the appearance of many morphogenic abnormalities such as mitosis aberrations. In conclusion, our data confirm the highest genotoxicity effects of Cr (VI) on Vicia faba root-cells.

Colchicine Alleviates Rosacea by Inhibiting Neutrophil Inflammation Activated by the TLR2 Pathway.

Rosacea is a chronic facial inflammatory skin disease that occurs with dysfunction of the immune system. Colchicine was reported to have anti-inflammatory properties. However, the impact of colchicine on rosacea remains unclear. In the present study, the phenotype of rosacea lesions was evaluated by the redness score, inflammatory biomarkers were analyzed by reverse transcription PCR (RT‒PCR), and the infiltration of inflammatory cells was assessed by IHC analysis and immunofluorescence in a rosacea-like mouse model. In vitro, RT‒PCR was used to identify the inflammatory factors that Toll-like receptor 2 (TLR2) agonist caused neutrophils to produce, and immunofluorescence and coimmunoprecipitation were used to identify putative signalling pathways. We found that skin erythema and histopathological alterations, as well as elevated proinflammatory factors (IL-1ß, IL-6, TNFα, CXCL2) and CAMP, were significantly ameliorated by colchicine treatment in LL37-induced rosacea-like mice. In addition, colchicine reduced the colocalization of TLR2 and neutrophils and the formation of neutrophil extracellular trap networks (NET) in mouse lesions. In neutrophils, colchicine markedly reduced TLR2 agonist-induced inflammatory biomarker expression, NET formation, and ROS production. Moreover, we found that LL37 could bind to TLR2 upon activation of TLR2 in neutrophils. Importantly, colchicine could repress the combination of TLR2 and LL37 in vivo. Finally, bioinformatics methods further validated the key molecules of neutrophil-related inflammation in rosacea, which is consistent with our experimental findings. Collectively, colchicine ameliorated rosacea-like dermatitis by regulating the neutrophil immune response activated by the TLR2 pathway, indicating that it could be an effective therapeutic option for patients with rosacea.

Design, synthesis, and bioevaluation of 1h-pyrrolo[3,2-c]pyridine derivatives as colchicine-binding site inhibitors with potent anticancer activities.

A new series of 1H-pyrrolo[3,2-c]pyridine derivatives were designed and synthesised as colchicine-binding site inhibitors. Preliminary biological evaluations showed that most of the target compounds displayed moderate to excellent antitumor activities against three cancer cell lines (HeLa, SGC-7901, and MCF-7) in vitro. Among them, 10t exhibited the most potent activities against three cancer cell lines with IC50 values ranging from 0.12 to 0.21 µM. Tubulin polymerisation experiments indicated that 10t potently inhibited tubulin polymerisation at concentrations of 3 µM and 5 µM, and immunostaining assays revealed that 10t remarkably disrupted tubulin microtubule dynamics at a concentration of 0.12 µM. Furthermore, cell cycle studies and cell apoptosis analyses demonstrated that 10t at concentrations of 0.12 µM, 0.24 µM, and 0.36 µM significantly caused G2/M phase cell cycle arrest and apoptosis. The results of molecular modelling studies suggested that 10t interacts with tubulin by forming hydrogen bonds with colchicine sites Thrα179 and Asnß349. In addition, the prediction of physicochemical properties disclosed that 10t conformed well to the Lipinski's rule of five.

Anti-rheumatic colchicine phytochemical exhibits potent antiviral activities against avian and seasonal Influenza A viruses (IAVs) via targeting different stages of IAV replication cycle.

BACKGROUND: The continuous evolution of drug-resistant influenza viruses highlights the necessity for repurposing naturally-derived and safe phytochemicals with anti-influenza activity as novel broad-spectrum anti-influenza medications. METHODS: In this study, nitrogenous alkaloids were tested for their viral inhibitory activity against influenza A/H1N1 and A/H5N1 viruses. The cytotoxicity of tested alkaloids on MDCK showed a high safety range (CC50 > 200 µg/ml), permitting the screening for their anti-influenza potential. RESULTS: Herein, atropine sulphate, pilocarpine hydrochloride and colchicine displayed anti-H5N1 activities with IC50 values of 2.300, 0.210 and 0.111 µg/ml, respectively. Validation of the IC50 values was further depicted by testing the three highly effective alkaloids, based on their potent IC50 values against seasonal influenza A/H1N1 virus, showing comparable IC50 values of 0.204, 0.637 and 0.326 µg/ml, respectively. Further investigation suggests that colchicine could suppress viral infection by primarily interfering with IAV replication and inhibiting viral adsorption, while atropine sulphate and pilocarpine hydrochloride could directly affect the virus in a cell-free virucidal effect. Interestingly, the in silico molecular docking studies suggest the abilities of atropine, pilocarpine, and colchicine to bind correctly inside the active sites of the neuraminidases of both influenza A/H1N1 and A/H5N1 viruses. The three alkaloids exhibited good binding energies as well as excellent binding modes that were similar to the co-crystallized ligands. On the other hand, consistent with in vitro results, only colchicine could bind correctly against the M2-proton channel of influenza A viruses (IAVs). This might explicate the in vitro antiviral activity of colchicine at the replication stage of the virus replication cycle. CONCLUSION: This study highlighted the anti-influenza efficacy of biologically active alkaloids including colchicine. Therefore, these alkaloids should be further characterized in vivo (preclinical and clinical studies) to be developed as anti-IAV agents.