BMC Complementary Medicine and Therapies

Chemical profiling and anticancer activity of Alnus incana dichloromethane fraction on HeLa cells via cell cycle arrest and apoptosis

Abstract Background Cervical cancer remains a global health challenge with persistently high incidence and mortality rates despite advancements in conventional treatments. The therapeutic potential of natural products has gained attention, particularly for their selective cytotoxicity and ability to modulate cancer pathways. Alnus incana (L.) Moench, a species-rich in bioactive compounds, shows potential as an anticancer agent; however, the cytotoxic effects of its leaves dichloromethane (DCM) extract remain underexplored. This study investigates the DCM fraction’s cytotoxicity on various cancer cell lines, with a primary focus on HeLa cells. Methods The cytotoxic effects of the A. incana DCM fraction were evaluated in a dose-dependent manner using the MTT assay on several cancer cell lines, with particular emphasis on HeLa cells. Flow cytometry was used to assess cell cycle arrest and apoptosis, while RT-qPCR quantified changes in the expression of apoptotic markers (Bax, Bcl-2, and p53). Chemical composition analysis was conducted using gas chromatography-mass spectrometry/flame ionization detection (GC-MS/FID) to identify the major bioactive compounds within the fraction. Results The DCM fraction exhibited dose-dependent cytotoxicity in HeLa cells, with an IC50 value of 135.6 µg/mL and a selectivity index (SI) of 2.72 relative to normal cells. Flow cytometry analysis revealed G0/G1 cell cycle arrest, significantly hindering progression through the S and G2/M phases. Moreover, there was a significant increase in both early and late apoptotic cell populations, correlating with the upregulation of pro-apoptotic genes (Bax and p53) and the downregulation of the anti-apoptotic gene Bcl-2. The chemical analysis identified 22 compounds in the unsaponifiable fraction, chiefly terpenoids such as phytol (65.74%). The saponifiable fraction presented a balanced composition of saturated (48.69%) and unsaturated (51.29%) fatty acids, with palmitic acid, linolenic acid, and linoleic acid as the predominant compounds. Conclusion While the DCM fraction’s relatively high IC50 value may limit its utility as a standalone treatment, its ability to induce cell cycle arrest and apoptosis demonstrates its promise as a co-therapeutic agent with conventional anticancer drugs. Further research is essential to elucidate its precise mechanisms of action and to evaluate its efficacy in combination therapies, potentially advancing its role in cervical cancer treatment.

Resveratrol activates MAPK/ERK pathway to regulate oestrogen metabolism in type I endometrial cancer

Abstract Objective Endometrial cancer (EC) is an oestrogen-dependent tumour, the occurrence of which is closely related to an imbalance of oestrogen homeostasis. Our previous studies explored the effects of Resveratrol(Res) on oestrogen metabolism. However, systematic research on the exact mechanism of action of Res is still lacking. Based on network pharmacology, molecular docking and animal experiments, the effects and molecular mechanisms of Res on endometrial cancer were investigated. Methods The target of Res was obtained from the high-throughput experiment and reference-guided database of TCM (HERB) and the Encyclopedia of Traditional Chinese Medicine (ETCM) databases, and the target of endometrial cancer was obtained by using the Genecards database. Venny map was used to obtain the intersection target of Res in the treatment of endometrial cancer, and the protein interaction network of the intersection target was constructed by importing the data into the STRING database. Then, the drug–disease–target interaction network was constructed based on Cytoscape 3.9.1 software. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed for intersection targets using the OmicShare cloud platform. Res and core targets were analysed by molecular docking. EC model mice induced by MNNG were randomly divided into the control group, Res group, MNNG group, MNNG + Res group, and MNNG + Res + MAPK/ERKi group. The protein levels of ERK and p-ERK in the mouse uterus were detected by Western blot. The levels of E1, E2, E3, 16-epiE3, 17-epiE3, 2-MeOE1, 4-MeOE1, 2-MeOE2, 4-MeOE2, 3-MeOE1, 2-OHE1, 4-OHE1, 2-OHE2, 4-OHE2, and 16α-OHE1 in the serum and endometrial tissue of mice were measured by LC‒MS/MS. Results A total of 174 intersection targets of Res anti-endometrial cancer were obtained. The signalling pathways analysed by KEGG enrichment included the AGE-RAGE signalling pathway in diabetic complications, the PI3K-Akt signalling pathway and the MAPK signalling pathway. The top 10 core targets were MAPK3, JUN, TP53, CASP3, TNF, IL1B, AKT1, FOS, VEGFA and INS. Molecular docking showed that in addition to TNF, other targets had good affinity for Res, and the binding activity with MAPK3 was stable. Western blot results showed that Res increased the phosphorylation level of ERK and that MAPK/ERKi decreased ERK activation. In the LC-MS/MS analysis, the levels of 2-MeOE1, 2-MeOE2 and 4-MeOE1 in serum and uterine tissue showed a significantly decreasing trend in the MNNG group, while that of 4-OHE2 was increased (P < 0.05). The concentrations of 4-MeOE1 in serum and 2-MeOE1 and 2-MeOE2 in the endometrial tissue of mice were significantly increased after Res treatment, and those of 4-OHE2 in the serum and uterus of mice were significantly decreased (P < 0.05). Meanwhile, in the MAPK/ERKi intervention group, the effect of Res on the reversal of oestrogen homeostasis imbalance was obviously weakened. Conclusion Res has multiple targets and multiple approaches in the treatment of endometrial cancer. In this study, it was found that Res regulates oestrogen metabolism by activating the MAPK/ERK pathway. This finding provides a new perspective for subsequent research on the treatment of endometrial cancer.

Promising anti-cervical cancer efficacy and phytochemical analysis of Vitex doniana Sweet (Verbenaceae) leaf extracts: an in vitro investigation with molecular mechanisms of action

The exploration of natural sources for novel anticancer agents has garnered significant attention in recent years, driven by the need for effective and safe anticancer medications, considering the escalating global burden of cervical cancer. Vitex doniana Sweet (Verbenaceae), a plant with diverse traditional medicinal uses, especially in Africa, has shown promise in this regard due to its rich phytochemical composition. This study investigated the cytotoxic and antiproliferative effects of V. doniana extracts on normal mammalian (Vero-CCL-81) and cervical cancer (HeLa) cells, including cancer-associated gene expression profiles, according to standard procedures. Phytochemical analysis was performed using gas chromatography-mass spectrometry (GC-MS). The findings revealed significant (P < 0.0001) concentration-dependent increases in cytotoxicity and inhibition of cancer cell proliferation, by the aqueous, methanolic, ethyl acetate, and dichloromethane leaf extracts of V. doniana. Notably, differential cytotoxicity profiles were observed among different extracts. The median cytotoxic concentrations (CC These findings provide valuable insights into the potential and molecular mechanisms underlying the anti-cervical cancer effects of V. doniana extracts, their attributable phytocompounds, and highlight their putative value as sources of lead compounds for the development of novel anticancer drugs.

Synergistic overcoming of cisplatin resistance in ovarian cancer by combined Astragalus Membranaceus and cisplatin treatment: network pharmacology and experimental validation

Ovarian cancer (OC) is a leading cause of mortality among gynecologic malignancies. Cisplatin (DDP) is a first-line chemotherapy agent, but resistance to DDP often develops, compromising its efficacy. Astragalus membranaceus (AS), a traditional Chinese medicine, has shown promise in enhancing chemotherapy sensitivity due to its anti-inflammatory and immunomodulatory properties. This study investigates the potential of AS to overcome DDP resistance in OC. We integrated multiple independent DDP-resistant OC datasets identified 337 DDP resistance-associated targets. Network pharmacology identified 20 active compounds in AS, with 22 potential targets related to DDP resistance. GO and KEGG analyses revealed enrichment in pathways involving inflammation and cell adhesion. Survival analysis indicated nine genes significantly associated with OC prognosis and immune infiltration. Molecular docking showed strong binding affinities between AS compounds and these targets. In vitro, assays demonstrated that AS combined with DDP significantly inhibited cell proliferation and migration while inducing apoptosis in DDP-resistant OC cells. Western blot analysis confirmed significant changes in critical proteins (IL1B, IL1A, SERPINE1, ITGA2, and AXL) with combined treatment. AS combined with DDP significantly enhances the inhibition of cell proliferation and migration while promoting apoptosis in DDP-resistant OC cells. These findings suggest that AS could be a valuable adjunct to DDP in overcoming chemoresistance in OC, potentially improving patient outcomes.

The ability of clostridium novyi-NT spores to induce apoptosis via the mitochondrial pathway in mice with HPV-positive cervical cancer tumors derived from the TC-1 cell line

A precise observation is that the cervix's solid tumors possess hypoxic regions where the oxygen concentration drops below 1.5%. Hypoxia negatively impacts the host's immune system and significantly diminishes the effectiveness of several treatments, including radiotherapy and chemotherapy. Utilizing oncolytic spores of Clostridium novyi-NT to target the hypoxic regions of solid tumors has emerged as a noteworthy treatment strategy. The transplantation procedure involved injecting TC-1 cells, capable of expressing HPV-16 E6/7 oncoproteins, into the subcutaneous layer of 6-8-week-old female C57/BL6 mice. The TC-1 cell line, was subcutaneously transplanted into 6-8-week-old female C57/BL6 mice. The tumor-bearing mice were randomly divided into 4 groups, and after selecting the control group, they were treated with different methods. Group 1- control without treatment (0.1 ml sterile PBS intratumor) Group 2- received cisplatin intraperitoneally (10 mg/kg) Group 3- received 10 Our results clearly showed that the injection of Clostridium novyi-NT spores (either intratumorally or intravenously) causes the regression of mouse cervical tumors. Spore germination induces internal apoptosis in cancer cells by inducing ROS production and increasing total cell ceramide, releasing cytochrome c and damaging mitochondria. Additionally, the results provided clear evidence of a significant decrease in the expression of HIF-1 alpha and VEGF proteins among the tumor groups that received spores, when compared to both the cisplatin-treated group and the control group. The study's outcomes demonstrated that the introduction of Clostridium novyi-NT spores triggered apoptosis in cervical cancer cells (derived from the TC-1 cell line) via the mitochondrial pathway, subsequently resulting in tumor regression in a mouse model.

Overcoming cisplatin resistance by targeting the MTDH-PTEN interaction in ovarian cancer with sera derived from rats exposed to Guizhi Fuling wan extract

Abstract Background The well-known traditional Chinese herbal formula Guizhi Fuling Wan (GFW) was recently reported to improve the curative effects of chemotherapy for ovarian cancer with few clinical side effects. The present study aimed to investigate the reversal mechanism of sera derived from rats exposed to Guizhi Fuling Wan extract (GFWE) in cisplatin-resistant human ovarian cancer SKOV3/DDP cells; the proteins examined included phosphatase and tensin homolog (PTEN) and metadherin (MTDH), and the possible protein interaction between PTEN and MTDH was explored. Methods GFWE was administered to healthy Wistar rats, and the sera were collected after five days. The PubMed and CNKI databases were searched for literature on the bioactive blood components in the sera. The systemsDock website was used to predict potential PTEN/MTDH interactions with the compounds. RT-qPCR, western blotting, and immunofluorescence analyses were used to analyze the mRNA and protein levels of MTDH and PTEN. Laser confocal microscopy and coimmunoprecipitation (co-IP) were used to analyze the colocalization and interaction between MTDH and PTEN. Results Sixteen bioactive compounds were identified in GFWE sera after searching the PubMed and CNKI databases. The systemsDock website predicted the potential PTEN/MTDH interactions with the compounds. RT-qPCR, western blotting, and immunofluorescence analyses showed decreased MTDH expression and increased PTEN expression in the sera. Laser confocal microscopy images and coimmunoprecipitation (co-IP) analyses demonstrated that a colocalization and interaction occurred between MTDH and PTEN, and the addition of the sera changed the interaction status. Conclusions GFWE restored sensitivity to cisplatin by inhibiting MTDH expression, inducing PTEN expression, and improving the interaction between MTDH and PTEN in SKOV3/DDP cells, and these proteins and their interaction may serve as potential targets for cancer treatment. The sera may represent a new source of anticancer compounds that could help to manage chemoresistance more efficiently and safely.

Effects of Leea indica leaf extracts and its phytoconstituents on natural killer cell-mediated cytotoxicity in human ovarian cancer

Abstract Background The rich biodiversity of medicinal plants and their importance as sources of novel therapeutics and lead compounds warrant further research. Despite advances in debulking surgery and chemotherapy, the risks of recurrence of ovarian cancer and resistance to therapy are significant and the clinical outcomes of ovarian cancer remain poor or even incurable. Objective This study aims to investigate the effects of leaf extracts from a medicinal plant Leea indica and its selected phytoconstituents on human ovarian cancer cells and in combination with oxaliplatin and natural killer (NK) cells. Methods Fresh, healthy leaves of L. indica were harvested and extracted in 70% methanol by maceration. The crude extract was partitioned with n-hexane, dichloromethane and ethyl acetate. Selected extracts and compounds were analyzed for their effects on cell viability of human ovarian cancer cells, NK cell cytotoxicity, and stress ligands expression for NK cell receptors. They were also evaluated for their effects on TNF-α and IL-1β production by enzyme-linked immunosorbent assay in lipopolysaccharide-stimulated human U937 macrophages. Results Leaf extracts of L. indica increased the susceptibility of human ovarian tumor cells to NK cell-mediated cytotoxicity. Treatment of cancer cells with methyl gallate but not gallic acid upregulated the expression of stress ligands. Tumor cells pretreated with combination of methyl gallate and low concentration of oxaliplatin displayed increased levels of stress ligands expression and concomitantly enhanced susceptibility to NK cell-mediated cytolysis. Further, NK cells completely abrogated the growth of methyl gallate-pretreated ovarian cancer cells. The leaf extracts suppressed TNF-α and IL-1β production in human U937 macrophages. Methyl gallate was more potent than gallic acid in down-regulating these cytokine levels. Conclusions We demonstrated for the first time that leaf extracts of L. indica and its phytoconstituent methyl gallate enhanced the susceptibility of ovarian tumor cells to NK cell cytolysis. These results suggest that the combined effect of methyl gallate, oxaliplatin and NK cells in ovarian cancer cells warrants further investigation, for example for refractory ovarian cancer. Our work is a step towards better scientific understanding of the traditional anticancer use of L. indica.

Cordyceps militaris induces apoptosis in ovarian cancer cells through TNF-α/TNFR1-mediated inhibition of NF-κB phosphorylation

Abstract Background Cordyceps militaris (L.) Fr. (C. militaris) exhibits pharmacological activities, including antitumor properties, through the regulation of the nuclear factor kappa B (NF-κB) signaling. Tumor Necrosis Factor (TNF) and TNF-α modulates cell survival and apoptosis through NF- κB signaling. However, the mechanism underlying its mode of action on the NF-κB pathway is unclear. Methods Here, we analyzed the effect of C. militaris extract (CME) on the proliferation of ovarian cancer cells by confirming viability, morphological changes, migration assay. Additionally, CME induced apoptosis was determined by apoptosis assay and apoptotic body formation under TEM. The mechanisms of CME were determined through microarray, immunoblotting and immunocytochemistry. Results CME reduced the viability of cells in a dose-dependent manner and induced morphological changes. We confirmed the decrease in the migration activity of SKOV-3 cells after treatment with CME and the consequent induction of apoptosis. Immunoblotting results showed that the CME-mediated upregulation of tumor necrosis factor receptor 1 (TNFR1) expression induced apoptosis of SKOV-3 cells via the serial activation of caspases. Moreover, CME negatively modulated NF-κB activation via TNFR expression, suggestive of the activation of the extrinsic apoptotic pathway. The binding of TNF-α to TNFR results in the disassociation of IκB from NF-κB and the subsequent translocation of the active NF-κB to the nucleus. CME clearly suppressed NF-κB translocation induced by interleukin (IL-1β) from the cytosol into the nucleus. The decrease in the expression levels of B cell lymphoma (Bcl)-xL and Bcl-2 led to a marked increase in cell apoptosis. Conclusion These results suggest that C. militaris inhibited ovarian cancer cell proliferation, survival, and migration, possibly through the coordination between TNF-α/TNFR1 signaling and NF-κB activation. Taken together, our findings provide a new insight into a novel treatment strategy for ovarian cancer using C. militaris.

Ginger inhibits the invasion of ovarian cancer cells SKOV3 through CLDN7, CLDN11 and CD274 m6A methylation modifications

Abstract Background Ginger is a common aromatic vegetable with a wide range of functional ingredients and considerable medicinal and nutritional properties. Numerous studies have shown that ginger and its active ingredients have suppressive effects on manifold tumours, including ovarian cancer (OC). However, the molecular mechanism by which ginger inhibits OC is not clear. The aim of this study was to investigate the function and mechanism of ginger in OC. Methods The estimation of n6-methyladenosine (m6A) levels was performed using the m6A RNA Methylation Quantification Kit, and RT-qPCR was used to determine the expression of m6A-related genes and proteins. The m6A methylationome was detected by MeRIP-seq, following analysis of the data. Differential methylation of genes was assessed utilizing RT-qPCR and Western Blotting. The effect of ginger on SKOV3 invasion in ovarian cancer cells was investigated using the wound healing assay and transwell assays. Results Ginger significantly reduced the m6A level of OC cells SKOV3. The 3’UTR region is the major site of modification for m6A methylation, and its key molecular activities include Cell Adhesion Molecules, according to meRIP-seq results. Moreover, it was observed that Ginger aids significantly in downregulating the CLDN7, CLDN11 mRNA, and protein expression. The results of wound healing assay and transwell assay showed that ginger significantly inhibited the invasion of OC cells SKOV3. Conclusions Ginger inhibits ovarian cancer cells’ SKOV3 invasion by regulating m6A methylation through CLDN7, CLDN11, and CD274.

Integrated bioinformatics and network pharmacology to explore the therapeutic target and molecular mechanisms of Bailing capsule on polycystic ovary syndrome

Abstract Background Polycystic ovary syndrome (PCOS) is a complex endocrine and metabolic disorder that is common in women of reproductive age. The clinical features of PCOS include hyperandrogenemia and polycystic ovarian changes. Bailing capsule (BL), a proprietary Chinese medicine that contains fermented Cordyceps sinensis powder, has been applied to treat PCOS. However, the specific active ingredients of BL and its mechanisms of action are yet to be elucidated. Methods Initially, the effectiveness of BL on PCOS model mice was evaluated. Subsequently, the active ingredients of BL were searched in the TCMSP and TCM Systems Pharmacology databases, and their targets were predicted using Swiss Target Prediction and SEA databases. Furthermore, the GEO gene database was used to screen for differentially expressed genes (DEGs) related to PCOS. Data from Gene Card, OMIM, DDT, and Drugbank databases were then combined to establish a PCOS disease gene library. Cross targets were imported into the STRING database to construct a protein–protein interaction network. In addition, GO and KEGG pathway enrichment analyses were performed using Metascape and DAVID databases and visualized using Cytoscape software and R 4.2.3. The core targets were docked with SYBYL-X software, and their expressions in PCOS mice were further verified using qPCR. Results The core active ingredients of BL were identified to be linoleyl acetate, cholesteryl palmitate, arachidonic acid, among others. Microarray data sets from four groups containing disease and normal samples were obtained from the GEO database. A total of 491 DEGs and 106 drug–disease cross genes were selected. Estrous cycle and ovarian lesions were found to be improved in PCOS model mice following BL treatment. While the levels of testosterone, progesterone, and prolactin decreased, that of estradiol increased. qPCR findings indicated that the expressions of JAK2, PPARG, PI3K, and AKT1 were upregulated, whereas those of ESR1 and IRS1 were downregulated in PCOS model mice. After the administration of BL, the expressions of associated genes were regulated. This study demonstrated that BL exerted anti-PCOS effects via PIK3CA, ESR1, AKT, PPARG, and IRS1 targets affecting PI3K-Akt signaling pathways. Discussion This research clarified the multicomponent, multitarget, and multichannel action of BL and provided a theoretical reference for further investigations on its pharmacological basis and molecular mechanisms against PCOS.

Exploring the mechanism and experimental verification of puerarin in the treatment of endometrial carcinoma based on network pharmacology and bioinformatics analysis

AbstractEndometrial carcinoma is one of the two cancers with rising mortality and morbidity in recent years. In the light of many controversies about its treatment, it is urgent to construct a new prognostic model and to find out new therapeutic directions. As a small drug molecule widely used in clinical treatment and experimental research in China, puerarin has recently been proven to have obvious anti-cancer effects in multiple cancer cells. In this study, bioinformatics analysis and experimental validation were used to explore the potential mechanism of puerarin for endometrial carcinoma and construct a prognostic model. A total of 22 drug-related differential genes were found by constructing a database of drug targets and disease genes. The protein–protein interaction network was constructed for GO and KEGG enrichment analysis to initially explore the potential mechanism of its therapeutic effects. To construct the prognostic model, validation was performed by risk regression analysis and LASSO analysis. Finally, two prognostic genes—PIM1 and BIRC5 were determined to establish high and low risk groups. Kaplan–Meier analysis displayed a higher survival rate in the low-risk group than in the high-risk group. ROC curves indicated the stable and good effect in prediction (one-year AUC is 0.626; two-year AUC is 0.620; three-year AUC is 0.623). The interrelationship between immunity and its disease was explored by immune infiltration analysis. Finally, the potential effect of puerarin on endometrial carcinoma cells was further verified by experiments.

Postbiotic metabolites derived from lactobacillus fermentum as potent antiproliferative bioresources on HeLa cells with promising biocompatibility

Chemotherapy administrations for cervical malignancy possess a variety of unfavorable influences on the human body. Scientists are interested in microbial-derived biomolecules or postbiotics as an alternative therapeutic strategy in malignant patients. This research investigated the mechanisms related to the function of two potential postbiotic Lactobacillus isolates, Lactobacillus fermentum CH and L. fermentum KH, isolated from indigenous Iranian dairy products. The Lactobacillus isolates were recognized through 16S rDNA sequence analysis followed by characterization using morphological and biochemical assays. The bioactivity of postbiotics on the cervical cancer model was also assessed through a cytotoxic study and apoptosis analysis. In addition, the anticancer activity was evaluated by qPCR, followed by a confirmation of the flow cytometry. The results of the bioactivity assay revealed that these postbiotics had suitable anticancer influences on the cervical cancer model (HeLa cells) by increasing BAX, caspase8, and caspase9, followed by a decrease in BCl-2, iKB (Inhibitor of nuclear factor kappa-B), and RelA gene expressions. Thus, the findings of this study signify that the postbiotic derivate from Lactobacillus strains isolated from indigenous Iranian dairy products could be regarded as a topical treatment with a promising curative index due to their effectiveness on cervical malignancy cells.

Based on network pharmacology and molecular docking technology to explore the pharmacodynamic components and mechanism of Gynostemmae Pentaphylli Herba reversing Cervical intraepithelial neoplasia

To investigate the pharmacodynamic components of Gynostemmae Pentaphylli Herba reversing Cervical intraepithelial neoplasia (CIN) were investigated by network pharmacology, and the mechanism of action was analyzed by molecular docking technology. The effective components and targets of Gynostemmae Pentaphylli Herba and the disease target of CIN were searched in TCMSP, Pubchem, Swiss Target Prediction, GenCards, WebGestalt, and STRING. Based on the above data and the Cytoscape software, we mapped the protein-protein interaction (PPI) co-expression network. The mechanism of Gynostemmae Pentaphylli Herba CIN treatment was identified from the enrichment analysis perspective. We performed molecular docking on the AutoDock. Finally, we carried out cell experiments for verification. Eighty-five targets matching the active ingredients of Gynostemmae Pentaphylli Herba, and 2512 CIN-related action targets were obtained. The nodal degree values of five components and the target of Gynostemmae Pentaphylli Herba in the top 5 were IL6, IL1β, TNF, TP53, and PTGS2. There were 52 intersection targets of the effective active ingredient of Gynostemmae Pentaphylli Herba and CIN. The PPI network map suggested that the main active ingredient MOL000098 (Quercetin) had the most targets (40), followed by MOL000351 (Rhamnazin) (17). Gene Ontology (GO) analysis yielded the regulation of transcription from RNA polymerase II promoter, cytoplasm, extracellular space and enzyme binding, zinc ion binding, etc. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway identified 114 signaling pathways. These pathways were mainly enriched in positive regulation of transcription from RNA polymerase II promoter, positive regulation of gene expression, cytoplasm, extracellular space, enzyme binding, zinc ion binding, Pathways in cancer, Fluid shear stress and atherosclerosis, etc. The binding energies of IL6, IL1β, TNF, TP53, and PTGS2 with Quercetin, Rhamnazin, Gypenoside XXVII_qt and Gypenoside XXVIII_qt were all less than -5 kcal·mol Gynostemmae Pentaphylli Herba may reverse CIN through targeted action of Quercetin, Rhamnazin, and gypenosides, as well as various pathways. Our study preliminarily explored the pharmacodynamic components and mechanism of the reversal of CIN by Gynostemmae Pentaphylli Herba and verified the possible mechanism of action through cell and molecular biology experiments to understand the pharmacological mechanism of Gynostemmae Pentaphylli Herba in CIN and provide data reference for new drugs and clinical trials research, which will be of great significance for clinical treatment decisions.

Ethanolic extract from Sophora moorcroftiana inhibit cell proliferation and alter the mechanical properties of human cervical cancer

Abstract Background Cervical cancer is one of the most common gynecological malignancies. Previous studies have shown that the ethanol extract of Sophora moorcroftiana seeds (EESMS) possesses an antiproliferative effect on several tumors in vitro. Therefore, in this study, we assessed the impact of EESMS on human cervical carcinoma (HeLa) cell proliferation. Methods The proliferation and apoptotic effects of HeLa cells treated with EESMS were evaluated using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay, dual acridine orange/ethidium bromide double staining, flow cytometry, and western blotting. Single-cell level atomic force microscopy (AFM) was conducted to detect the mechanical properties of HeLa cells, and proteomics and bioinformatics methods were used to elucidate the molecular mechanisms of EESMS. Results EESMS treatment inhibited HeLa cell proliferation by blocking the G0/G1 phase, increasing the expression of Caspase-3 and affecting its mechanical properties, and the EESMS indicated no significant inhibitory effect on mouse fibroblasts L929 cell line. In total, 218 differentially expressed proteins were identified using two-dimensional electrophoresis, and eight differentially expressed proteins were successfully identified using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry. The differentially expressed proteins were involved in various cellular and biological processes. Conclusion This study provides a perspective on how cells change through biomechanics and a further theoretical foundation for the future application of Sophora moorcroftiana as a novel low-toxicity chemotherapy medication for treating human cervical cancer.

Antiproliferative activities of some selected Nigerian medicinal plants against breast, liver, and cervical cancer cells

Abstract Background Phytochemicals have become a growing source of alternative medicine in developing countries due to the poor prognosis, high cost of conventional pharmaceuticals, and undesirable effects associated with mainstream cancer treatment. Objective This study was aimed at investigating the anticancer effect of some selected Nigerian medicinal plants used in cancer treatment. These include ethanol extracts of Dialium guineense root (DGR), Dialium guineense leaves (DGL), Jateorhiza macrantha leaves (JML), Musanga cecropioides leaves (MCL), Musanga cecropioides stembark (MCSB), Piptadeniastrum africanum stembark (PASB), Piptadeniastrum africanum root (PAR), Pupalia lappacea flower tops (PLF), Raphiostylis beninensis root (RBR), Raphiostylis beninensis leaves (RBL), Ritchiea capparoides leaves (RCL), Ritchiea capparoides stembark (RCSB), and Triplochiton scleroxylon stembark (TSB). Methods The cytotoxic activity of the extracts was examined using a brine shrimp lethality assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay against three cancer cell lines, including MCF-7, HUH-7, and HeLa. The selectivity of all extracts towards cancer cells was investigated using normal lung fibroblasts (MRC-5). Cell migration and colony-forming assays of active extracts against MCF-7 cells were also performed. Additionally, the total polyphenolic contents of the active extracts were estimated using standard methods. Results The extract of PASB had the highest cytotoxicity (LC50 = 1.58 μg/mL) on the brine shrimps compared to vincristine sulphate (LC50 = 2.24 μg/mL). In the cell viability assay, all the extracts produced significant (p &lt; 0.05) growth inhibitory effects against all cell lines tested in a dose-dependent manner. All extracts were selective to cancer cells at varying degrees. Worth mentioning are the extracts of MCL, DGR, RBR, and PASB, which exhibited 14-, 7-, 6- and 2-fold selectivity toward MCF-7 cancer cells relative to normal lung fibroblast (MRC-5), respectively. These four extracts also significantly inhibited cell migration and colony formation in MCF-7-treated cells in dose-dependent manners. Considerable amounts of phenolics, flavonoids, and proanthocyanidins were detected in all extracts evaluated. Conclusion These findings advocate the continued development of MCL, DGR, RBR, and PASB as potential chemotherapeutic agents.

Acetylmelodorinol isolated from Sphaerocoryne affinis seeds inhibits cell proliferation and activates apoptosis on HeLa cells

Abstract Background Cervical cancer is a major global health concern with a high prevalence in low- and middle-income countries. Natural products, particularly plant-derived compounds, have shown immense potential for developing anticancer drugs. In this study, we aimed to investigate the anticancer properties of the pericarp and seeds of Sphaerocoryne affinis fruit on human cervical carcinoma cells (HeLa) and isolate the bioactive compound from the active fraction. Methods We prepared solvent fractions from the ethanol extracts of the pericarp and the seed portion by partitioning and assessing their cytotoxicity on HeLa cells. Subsequently, we collected acetylmelodorinol (AM), an anticancer compound, from the ethyl acetate fraction of seeds and determined its structure using nuclear magnetic resonance. We employed cytotoxicity assay, western blotting, Annexin V apoptosis assay, measurement of intracellular reactive oxygen species (ROS) levels, 4′,6-diamidino-2-phenylindole (DAPI) staining, and a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, to evaluate the anticancer properties of AM on HeLa. Results The solvent fractions from the seed displayed considerably higher cytotoxic activity against HeLa cells than those of the pericarp. We isolated and identified acetylmelodorinol as an anticancer compound from the ethyl acetate fraction from S. affinis seed extract. Treatment with acetylmelodorinol inhibited HeLa cell proliferation with an IC50 value of 2.62 ± 0.57 µg/mL. Furthermore, this study demonstrated that acetylmelodorinol treatment disrupted cell cycle progression by reducing the expression of cyclin E, CDK1/2, and AKT/mTOR pathways, increasing the intracellular ROS levels, reducing BCL-2/BCL-XL expression, causing DNA fragmentation and nuclear shrinkage, and triggering apoptosis through caspase 3 and 9 activation in a dose-and time-dependent manner. Conclusion In contrast to previous reports, this study focuses on the inhibitory effects of AM on the AKT/mTOR pathway, leading to a reduction in cell proliferation in cervical cancer cells. Our findings highlight the promising potential of acetylmelodorinol as an effective treatment for cervical cancer. Additionally, this study establishes a foundation for investigating the molecular mechanisms underlying AM’s properties, fostering further exploration into plant-based cancer therapies.

Heracleum persicum Desf. ex Fisch., C.A.Mey. &amp; Avé-Lall. fruit essential oil: content, antimicrobial activity and cytotoxicity against ovarian cancer cell line

Abstract Background One of the most important aromatic species of the Apiaceae family is Heracleum persicum Desf. ex Fisch., C.A.Mey. &amp; Avé-Lall, which used as a spice and condiment in food. It is widely used in traditional Iranian medicine due to its anti-toxic properties. The aim of this study was to evaluate the essential oil of this plant in terms of content and ingredients, cytotoxicity and antimicrobial activity. Methods The fruit of H. persicum was collected in June 2019 from Maragheh region of Kashan, Iran. The essential oil was extracted by water distillation using Clevenger for 3 h. Identification analysis of H. persicum fruit essential oil (HPFEO) components was performed using gas chromatography-mass spectrometry (GC–MS). Evaluation of the effect of the HPFEO on the growth and proliferation of ovarian cancer cell line (OVCAR-3) was performed using MTT colorimetric method. Its antimicrobial activity was evaluated by agar diffusion method, minimum inhibitory concentration (MIC) and minimum bactericidal/fungal lethality concentration (MBC/MFC). Results The results of analysis of the HPFEO by GC–MS showed that there were 35 compounds with 99.54% relative content. Hexyl butyrate (35.24%), octyl 2-methylbutyrate (11.65%), octyl isobutyrate (9.23%), and octyl acetate (8.42%) were the predominant compounds. Examination of cell survival showed that the viability of cells decreased depending on the concentration of the HPFEO in 24 h and had value of IC50 ~ 12.08 μg/ml against OVCAR-3 cell line. The strongest inhibitory activity of the HPFEO was against Gram-negative bacteria Pseudomonas aeruginosa and Salmonella paratyphi-A serotype (MIC &lt; 62.50 μg/mL). Also, the strong inhibitory and lethal activity of this essential oil against Candida albicans (MIC and MBC 250 μg/mL was one degree weaker compared to nystatin (MIC = 125 μg/mL). Conclusions Thus, HPFEO, in addition to its traditional use, may have a strong and potential potential for the production of anti-proliferative and antimicrobial drugs.

Potent anti-cancer activity of Sphaerocoryne affinis fruit against cervical cancer HeLa cells via inhibition of cell proliferation and induction of apoptosis

Abstract Background Cervical cancer remains a significant global health issue, highlighting the need for effective therapeutic strategies. Given that Sphaerocoryne affinis (SA) has shown potential anti-cancer activity in several cancer types, herein, we investigate the effects of SA fruit (SAF) on human cervical cancer HeLa cells and their underlying mechanisms of action. Methods SAF extract cytotoxicity was assessed in various cancer cell lines. The effects of the hexane fraction (SAF-Hex) on HeLa cell viability, cell cycle protein expression, apoptosis, and DNA damage were evaluated using cytotoxicity assays, Western blotting, quantitative PCR, 4′,6-diamidino-2-phenylindole (DAPI) staining, and a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Results SAF-Hex selectively inhibited HeLa cell viability with an IC50 of 4.20 ± 0.36 µg/mL and a selectivity index of 5.11 ± 0.58. The time-dependent cytotoxicity assay showed decreased cell survival after 48 h of treatment, accompanied by morphological changes and apoptotic bodies in HeLa cells. SAF-Hex also suppressed HeLa cell cycle proteins (Cyclin E, CDK2, and CDK1), reduced PCNA transcription, and diminished AKT and mTOR activation, thus inhibiting cell proliferation. The increased γH2AX expression, DNA fragmentation, and caspases-3 and -9 activation indicated SAF-Hex-induced DNA damage and apoptosis. However, the BAX/BCL-2 ratio remained unchanged, and BAX and BCL2 expression was attenuated. Conclusion SAF-Hex effectively inhibits HeLa cell proliferation and induces DNA damage in that cervical cancer cell line activating apoptosis through the intrinsic pathway. Interestingly, the BAX/BCL-2 ratio remained unchanged while BAX and BCL2 transcription was attenuated. Hence, further research is required to explore this unexpected finding and facilitate the development of novel therapies targeting cervical cancer HeLa cells.

Butyrate inhibits the mitochondrial complex Ι to mediate mitochondria-dependent apoptosis of cervical cancer cells

Abstract Background Cervical cancer (CC) is a common gynecological malignancy with high morbidity worldwide. Butyrate, a short-chain fatty acid produced by intestinal flora, has been reported to inhibit cervical carcinogenesis. This study aimed to investigate the pro-apoptotic effects of butyrate on CC and the underlying mechanisms. Methods Human HeLa and Ca Ski cells were used in this study. Cell proliferation, cell migration and invasion were detected by CCK-8 and EdU staining, transwell and wound healing assay, respectively. Cell cycle, mitochondrial membrane potential and apoptosis were evaluated by flow cytometry. Western blot and RT-qPCR were carried out to examine the related genes and proteins to the mitochondrial complex Ι and apoptosis. Metabolite changes were analyzed by energy metabolomics and assay kits. The association between G protein-coupled receptor 41, 43, 109a and CC prognosis was analyzed using data from The Cancer Genome Atlas (TCGA). Results CCK-8 results showed significant inhibition of CC cell proliferation induced by butyrate treatment, which was confirmed by EdU staining and cell cycle detection. Data from the transwell and wound healing assay revealed that CC cell migration was dramatically reduced following butyrate treatment. Additionally, invasiveness was also decreased by butyrate. Western blot analysis showed that cleaved Caspase 3 and cleaved PARP, the enforcers of apoptosis, were increased by butyrate treatment. The results of Annexin V/PI staining and TUNEL also showed an increase in butyrate-induced apoptotic cells. Expression of Cytochrome C (Cytc), Caspase 9, Bax, but not Caspase 12 or 8, were up-regulated under butyrate exposure. Mechanistically, the decrease in mitochondrial NADH and NAD + levels after treatment with butyrate was observed by energy metabolomics and the NAD+/NADH Assay Kit, similar to the effects of the complex Ι inhibitor rotenone. Western blot results also demonstrated that the constituent proteins of mitochondrial complex Ι were reduced by butyrate. Furthermore, mitochondria-dependent apoptosis has been shown to be initiated by inhibition of the complex Ι. Conclusion Collectively, our results revealed that butyrate inhibited the proliferation, migration and invasion of CC cells, and induced mitochondrial-dependent apoptosis by inhibiting mitochondrial complex Ι.

Role of limonin in anticancer effects of Evodia rutaecarpa on ovarian cancer cells

Abstract Background Ovarian cancer therapy generally involves systemic chemotherapy with anticancer drugs; however, chemotherapy with a platinum-based drug has often been shown to cause adverse reactions and drug resistance in ovarian cancer patients. Evodia rutaecarpa (ER) reportedly shows anticancer activity against various types of cancer cells. However, the effects of ER have not yet been fully uncovered in ovarian cancer. Methods In the present study, we investigated the anticancer effects of an ER extract and its components against the ovarian cancer cell lines SKOV-33, A2780, RMUG-S and a cisplatin-resistant SKOV-3 cell line (CisR SKOV-3). Cell viability and colony formation assays along with subcellular fractionation analysis, immunoblotting, and immunofluorescence staining were performed. Results ER treatment led to a significant reduction in the viability of SKOV-3 cells. Moreover, limonin, a compound found in ER, reduced the viability of both serous-type (SKOV-3 and A2780) and mucinous-type (RMUG-S) ovarian cancer cells by inducing apoptosis via activation of the p53 signaling pathway. Furthermore, limonin reversed the drug resistance through activation of apoptosis in CisR SKOV-3. Conclusion Taken together, our findings suggest that limonin contributes to the anti-ovarian cancer effects of ER by inducing apoptosis via activation of the p53 signaling pathway.

Methanolic neem (Azadirachta indica) stem bark extract induces cell cycle arrest, apoptosis and inhibits the migration of cervical cancer cells in vitro

Abstract Background Cervical cancer remains one of the significant causes of mortality in women due to the limitations of current treatment strategies and their associated side effects. Investigation of alternative medicine, including phytomedicine, has shown effective anti-cancer potential with fewer side effects. Azadirachta indica (commonly known as neem) is known for its medicinal properties. The present study investigated the anti-cancer potential of methanolic neem stem bark extract (MNBE) against cervical cancer using HeLa, SiHa, and ME-180 cell lines. Methods Cytotoxic effect of MNBE on cultured cell lines was evaluated by MTT and clonogenic assay. The growth-inhibiting effect of MNBE was further confirmed by performing cell cycle analysis and apoptosis assay using flow cytometry. The anti-migratory effect of MNBE was evaluated by using wound healing and Boyden chamber assay. Real-time PCR was used to determine the mRNA expression, and western blot and flow cytometry was used to determine the protein levels of growth and migration-related genes. Results MNBE significantly suppressed the growth and survival of cervical cancer cells in a dose-dependent manner by inducing cell cycle arrest and apoptosis. In addition, the growth inhibitory effect of MNBE was specific to cervical cancer cells than normal cells. Cell cycle arrest was correlated to transcriptional downregulation of cyclin dependent kinase 1 (CDK1), cyclin A, and cyclin B. Additionally, MNBE treatment resulted in the upregulation of active caspase-3 protein and downregulation of prosurvival genes, Bcl2, and survivin at mRNA level and NFkB-p65 at the protein level. Furthermore, MNBE inhibited the migration of cervical cancer cells accompanied by modulation of migration-related genes, including zona occludens-1 (ZO-1), matrix metalloproteinase 2 (MMP2), focal adhesion kinase (FAK), N-cadherin, snail, and E-cadherin. Conclusion In summary, the present study provides the first evidence of MNBE in restricting cervical cancer cell growth and migration, which warrants further investigation for developing novel anti-cancer drugs.

Cytotoxic and immunomodulatory properties of Tinospora cordifolia, Boerhaavia diffusa, Berberis aristata, and Ocimum basilicum extracts against HPV-positive cervical cancer cell line

Cervical cancer remains a significant public health concern, especially for low-and middle-income countries. This study explores the dual potential of medicinal plant extracts as both anticancer agents and immunomodulators, particularly in HPV-positive cervical cancer, while also addressing the limitations of conventional chemotherapy. Extracts from Tinospora cordifolia, Boerhaavia diffusa, Berberis aristata, and Ocimum basilicum were tested on CaSki and HEK 293 cells alongside cisplatin. Cytotoxicity, genotoxicity, cell migration, HPV DNA inhibition, IFNG secretion, and cell cycle modulation were assessed using established biochemical and immunological assays, including qPCR, ELISA, and flow cytometry. Among the extracts, B. aristata demonstrated the strongest anticancer effects on cancer cells while exhibiting minimal impact on normal cells, highlighting its therapeutic potential. It also significantly reduced cell migration and has synergistic affect with cisplatin. B. diffusa emerged as the most effective in inhibiting HPV 16 DNA, suggesting its role in viral suppression. Immunomodulatory analysis revealed that T. cordifolia and O. basilicum significantly enhanced IFNG secretion, indicative of robust immune activation. Each tested extracts induced G0/G1 phase cell cycle arrest, with T. cordifolia showing the most pronounced effect. This study highlights the novel therapeutic promise of integrating plant extracts into HPV-positive cervical cancer management. B. aristata stands out for its cytotoxicity and anti-migration properties, while T. cordifolia offers significant immunomodulatory benefits. These findings pave the way for further research into combining natural products with conventional therapies for safer, more effective cancer treatments.

Urtica dioica leaf infusion enhances cisplatin-induced apoptosis in ovarian cancer cells in vitro

Urtica Dioica (UD) or stinging nettle has been widely used for its therapeutic benefits and biological activities. Recent studies have reported its beneficial effect in treating cancer, most importantly when combined with chemotherapeutic drugs. To our knowledge, no studies investigated the potential effect of UD to enhance the sensitivity of ovarian cancer cells to cisplatin. In this study we aim to investigate whether this combination might possess anti-proliferative, pro-apoptotic, and anti-metastatic properties on one of the most aggressive ovarian cancer cell lines, SKOV-3 cells, in vitro. To elucidate its therapeutic values, cytotoxicity and DNA fragmentation assays were performed along with cell cycle and apoptosis assays using flow cytometry, RT-qPCR, and western blot analysis. Additionally, wound healing and trans-well migration assays were used to study the effect of this combination on the motility of ovarian cancer cells. Results showed that the combination of nettle extract and cisplatin significantly decreased the proliferation of SKOV-3 cells in a dose and time-dependent manner compared to each treatment alone by inducing cellular death as revealed by major apoptotic markers including phosphatidylserine translocation to the outer membrane leaflet, DNA fragmentation, and the upregulation of cleaved PARP protein. Further evaluation verified the activation of extrinsic apoptosis via the caspase-dependent pathway as demonstrated by the upregulated expression levels of caspases 3 and 8. Finally, the combination of nettle tea and cisplatin showed an inhibitory effect on the motility and migratory capacities of SKOV-3 cells. As a result, Urtica Dioica leaf infusion was found effective in enhancing cisplatin-induced apoptosis while inhibiting the tumor progression of one of the most aggressive ovarian cancer cells in vitro.

In vitro analysis of the molecular mechanisms of ursolic acid against ovarian cancer

Ovarian cancer is one of most common gynaecologic malignancy and ranks third in cancer-related deaths among women. Ursolic acid (UA) is a pharmacologically active pentacyclic triterpenoid isolated from a large variety of vegetables, fruits and many traditional medicinal plants. However, the mechanism of action of UA in inhibiting the proliferation of ovarian cancer cells remains unclear. Consequently, this experiment was designed to elucidate the mechanism of action of UA in inhibiting the proliferation of ovarian cancer cells in greater detail.The results indicated that UA was capable of effectively inhibiting the proliferation, migration, and colony formation of ovarian cancer cells.UA was observed to up-regulate Bcl-2-associated X protein(BAX)and cysteinyl aspartate specific proteinase 3 (Caspase3) expression and down-regulating B-cell lymphoma-2(Bcl-2) expression.Meanwhile, UA up-regulated Sequestosome 1(p62)expression and down-regulated coiled-coil, moesin-like BCL2-interacting protein(Becline1), microtubule-associated proteins light chain 3(LC3), Phosphoinositide 3-Kinase(PI3K), andProtein Kinase B( AKT) expression, thus effectively inhibiting autophagy in ovarian cancer cells.Furthermore, UA upregulated pancreatic ER kinase (PKR)-like ER kinase (PERK), eukaryotic translation initiation factor 2 A(eIF2α), and The C/EBP Homologous Protein(CHOP) expression.In addition UA upregulates PERK, eIF2α, and CHOP expression and effectively promotes endoplasmic reticulum stress(ERS).In conclusion, UA can inhibit ovarian cancer cell proliferation, migration, colony formation, and may inhibit tumor cell autophagy by promoting tumor cell ERS, and ultimately promote ovarian cancer cell apoptosis.