Human Osteosarcoma Cell Line Research Articles

The Contribution of the Novel CLTC-VMP1 Fusion Gene to Autophagy Regulation and Energy Metabolism in Cisplatin-Resistant Osteosarcoma.

Osteosarcoma (OS) is a highly malignant tumor, and chemotherapy resistance is a major challenge in the treatment of this disease. This study aims to explore the role of the CLTC-VMP1 gene fusion in the mechanism of chemotherapy resistance in OS and investigate its molecular mechanisms in mediating energy metabolism reprogramming by regulating autophagy and apoptosis balance. Using single-cell transcriptome analysis, the heterogeneity of OS cells and their correlation with resistance to platinum drugs were revealed. Cisplatin-resistant cell lines were established in human OS cell lines for subsequent experiments. Based on transcriptomic analysis, the importance of VMP1 in chemotherapy resistance was confirmed. Lentiviral vectors overexpressing or interfering with VMP1 were used, and it was observed that inhibiting VMP1 could reverse cisplatin resistance, promote cell apoptosis, and inhibit autophagy, as well as mitochondrial respiration and glycolysis. Furthermore, the presence of CLTC-VMP1 gene fusion was validated, and its ability to regulate autophagy and apoptosis balance, promote mitochondrial respiration, and glycolysis was demonstrated. Mouse model experiments further confirmed the promoting effect of CLTC-VMP1 on tumor growth and chemotherapy resistance. In summary, the CLTC-VMP1 gene fusion mediates energy metabolism reprogramming by regulating autophagy and apoptosis balance, which promotes chemotherapy resistance in OS.

Multifunctional zinc oxide loaded stearic acid surfaces on biodegradable magnesium WE43 alloy with hydrophobic, self-cleaning and biocompatible attributes

Biodegradable Mg alloys are one of the most promising materials for implant applications, for which the widespread use is hampered by rapid degradation in physiological environment. The present study explores the development of a novel zinc-oxide (ZnO) loaded stearic acid (SA) coating on medical grade WE43 alloy using a facile, and environment-friendly technique for promising biomaterial surface applications. Morphology and chemical analyses reveal the development of a layered coating surface with uniformly dispersed ZnO nanoparticles in SA matrix. The developed coating exhibits hydrophobic performance (contact angle of 135°) along with the retainment of hydrophobicity under varying chemical (acidic pH of 1 and basic pH of 13) and mechanical (post scratch testing and peel-off studies) conditions. The observed surface water repellency facilitates self-cleaning performance pointing towards its efficacy against potential biofilm formation. The developed coating demonstrates superior resistance against surface degradation during immersion studies in phosphate buffer saline solution (pH = 7.4) and better cell viability with MG-63 (human osteosarcoma) cell line. The approach presented here sets a platform for the development of efficient coatings on biodegradable WE43 alloys surfaces for potential biomaterial technologies.

403 (PB391): Response of canine and human osteosarcoma cell lines to targeted agents compared to clinically relevant exposure

403 (PB391): Response of canine and human osteosarcoma cell lines to targeted agents compared to clinically relevant exposure

Migration and retention of human osteosarcoma cells in bioceramic graft with open channel architecture designed for bone tissue engineering

The microstructure of a porous bioceramic bone graft, especially the pore architecture, plays a crucial role in the performance of the graft. Conventional bioceramic grafts typically feature a random, closed-pore structure, limiting biological activity to the periphery of the graft. This can lead to delay in full integration with the host site. Bioceramic forms with open through pores can perform better because their inner regions are accessible for natural bone remodeling. This study explores the influence of open through pores in a bioceramic graft on the migration and retention of the local cellsin vitro, which will correlate to the rate of healingin vivo.Hydroxyapatite ceramic forms with aligned channels were fabricated using slip casting technique, employing sacrificial fibers. The sorption characteristics across the graft were evaluated using human osteosarcoma cell line. Seven-day cultures showed viable cells within the channels, confirmed by live/dead assay, scanning electron microscope analysis, and cytoskeletal staining, indicating successful cell colonization. The channel architecture effectively enhances cell migration and retention throughout its entire structure, suggesting potential applications in bone tissue engineering based on the results obtained.

Inhibition of osteosarcoma metastasis in vivo by targeted downregulation of MMP1 and MMP9

Osteosarcoma (OS) mortality stems from lung metastases. Matrix metalloproteinases (MMPs) facilitate metastatic dissemination by degrading extracellular matrix components. Herein we studied the impact of targeted MMP downregulation on OS metastasis. Differential gene expression analysis of human OS cell lines revealed high MMP9 expression in the majority of OS cell lines. Furthermore, 143B, a metastatic OS cell line, exhibited increased MMP1 and MMP9 mRNA levels. Gene set enrichment analysis on metastatic and non-metastatic OS patient specimens indicated epithelial-mesenchymal transition as the most enriched gene set, with MMP9 displaying strong association to genes in this network. Using the same dataset, Kaplan-Meier analysis revealed a correlation between MMP1 expression and dismal patient survival. Hence, we undertook targeted suppression of MMP1 and MMP9 gene expression in OS cell lines. The ability of OS cells to migrate and form colonies was markedly reduced upon MMP1 and MMP9 downregulation, whereas their cell proliferation capacity remained intact. MMP9 downregulation decreased tumor growth and lung metastases area in an orthotopic mouse OS model. Consistently, human OS lung metastasis specimens displayed marked MMP9 protein expression. Our findings highlight the role of MMP1 and MMP9 in OS metastasis, warranting further exploration of simultaneous inhibition of MMPs for future OS therapeutics.

Expression of Matrix Metalloproteinase-9, P53 and B-cl2 Genes in Human Osteosarcoma Cell Line Treated with Ginger Extract

Expression of Matrix Metalloproteinase-9, P53 and B-cl2 Genes in Human Osteosarcoma Cell Line Treated with Ginger Extract

Development of injectable microgel-based scaffolds via enzymatic cross-linking of hyaluronic acid-tyramine/gelatin-tyramine for potential bone tissue engineering

Currently, the healing of large bone defects relies on invasive surgeries and the transplantation of autologous bone. As a less invasive treatment option, the provision of microenvironments that promote the regeneration of defective bones holds great promise. Here, we developed hyaluronic acid (HA)/gelatin (Ge) microgel-based scaffolds to guide bone regeneration. To enable the formation of microgels by enzymatic cross-linking in the presence of horseradish peroxidase (HRP) and hydrogen peroxide (H2O2), we modified the polymers with tyramine (TA). Spectrophotometry and proton nuclear magnetic resonance (1H NMR) spectroscopy analysis confirmed successful tyramine substitution on polymer backbones. To enable the formation of microgels by a water-in-oil emulsion approach, the HRP and H2O2 concentrations were tuned to achieve the gelation in a few seconds. By varying the stirring speed from 600 to 1000 rpm, spherical microgels were produced with an average size of 116 ± 8.7 and 68 ± 4.7 μm, respectively. The results showed that microgels were injectable through needles and showed good biocompatibility with the cultured human osteosarcoma cell line (MG-63). HA/Ge-TA microgels served as a promising substrate for MG-63 cells since they improved the alkaline phosphatase activity and level of calcium deposition. In summary, the developed HA/Ge-TA microgels are promising injectable microgel-based scaffolds in bone tissue engineering.

Polyacrylonitrile‒Chitosan IPN composite scaffolds that closely mimic the human trabecular bone structure for tissue engineering

Successful bone tissue engineering involves managing several important parameters, such as the design of intercommunicating porous structures, pore sizes, and the material and mechanical suitability of the material. In our work, we focused on the preparation of a synthetic scaffold that morphologically mimics the structure of human trabecular bone. The scaffolds were fabricated through the thermal modification (TM) of polyacrylonitrile. The scaffold strength was supported by a crosslinked chitosan supporting network. The prepared scaffold has imprinted pores of the appropriate size to facilitate the ingrowth and proliferation of human osteoblasts throughout the entire pore volume created by a primary porogen, sodium chloride. The resulting material has a dual porous morphological structure, in which adjustable larger pores support cellular ingrowth into the scaffold, whereas smaller pores, created using succinonitrile (SCN) as a secondary porogen, increase the diffusion of oxygen and nutrients to developing cells. The mechanical properties of the scaffold were promoted by the use of a secondary interpenetrating network (IPN) based on chitosan. The incorporation of secondary IPNs led to a significant improvement in the mechanical characteristics of the scaffold. Two crosslinking agents were used: the widely utilized glutaraldehyde (GA) and its green, nontoxic alternative, genipin (GEN).The current study introduces a synthetic scaffold that effectively mimics the structure of human trabecular bone, providing a conducive environment for osteoblast proliferation and ingrowth. We incorporated a dual porous morphology and a secondary IPN based on chitosan and found that the scaffold exhibited improved mechanical properties and nutrient diffusion capabilities. This research highlights the potential of these scaffolds for major advancements in bone tissue engineering, demonstrating their noncytotoxic nature and ability to support the adhesion and proliferation of the human osteosarcoma cell line SAOS-2. The described scaffold preparation technique may offer distinct advantages over current high-end methods such as 3D printing. The primary benefit lies in its very simple instrumentation, which enables the creation of a complex trabecular bone morphology with both macro- and microporosity in the bulk, which is still challenging for common 3D printers. Additionally, IPN with chitosan can be postmodified, substantially expanding the potential applications of these scaffolds.

Ezetimibe Inhibits Cell Viability and Invasion by Suppressing PI3K/AKT Signaling Pathway in Human Osteosarcoma Cells

Introduction: Osteosarcoma is one of the most prevalent malignant bone tumors with a poor overall prognosis and mainly happens in children and adolescents. Current therapy strategies still possess a lot of limitations, and new and efficient strategies are required. Ezetimibe was previously reported to have its anti-tumor effect in various tumors, but the investigation of Ezetimibe on osteosarcoma is still limited. Aims: This study explores whether ezetimibe could exert an anti-tumor effect on human osteosarcoma cell lines, U2OS, and Saos-2. Methods: The effect of ezetimibe on the proliferation of osteosarcoma was explored by CCK-8 and colony formation assay. The role of ezetimibe on osteosarcoma cell migration and invasion was explored by wound healing assay and transwell assay. The role of ezetimibe on osteosarcoma cell apoptosis was explored by PI/Annexin V analysis. In addition, a western blot was performed to verify the phenotype. Results: The flow cytometry assay indicated that ezetimibe could promote osteosarcoma apoptosis. Western blot assay further demonstrated the effect of ezetimibe on proliferation, migration, invasion and apoptosis- related proteins. Finally, the deep anti-tumor effect of ezetimibe contributed to suppressing the PI3K/AKT signaling pathway. Conclusion: Present data indicated that ezetimibe has an antitumor effect on osteosarcoma and could be considered a future osteosarcoma treatment.

Influence of Metamizole on Antitumour Activity of Risedronate Sodium in In Vitro Studies on Canine (D-17) and Human (U-2 OS) Osteosarcoma Cell Lines.

The availability of metamizole varies greatly around the world. There are countries such as the USA, UK, or Australia where the use of metamizole is completely forbidden, and there are also countries where this drug is available only on prescription (e.g., Greece, Italy, Spain, etc.) and those in which it is sold OTC-over the counter (e.g., most Asian and South American countries). Metamizole, as a drug with a strong analgesic effect, is used as an alternative to other non-steroidal anti-inflammatory drugs, alone or in combination with opioid drugs. Risedronate sodium is a third-generation bisphosphonate commonly used in orthopaedic and metabolic diseases of the musculoskeletal system, including hypercalcemia, postmenopausal osteoporosis, Paget's disease, etc. The aim of this study was to check whether there were any pharmacological interactions between metamizole and risedronate sodium in in vitro studies. Cell viability was assessed using the MTT method, the number of apoptotic cells was assessed using the labelling TUNEL method, and the cell cycle assessment was performed with a flow cytometer and propidium iodide. This was a pilot study, which is why only two cancer cell lines were tested: D-17 of canine osteosarcoma and U-2 OS of human osteosarcoma. Exposure of the canine osteosarcoma cell line to a combination of risedronate sodium (100 µg/mL) and metamizole (50, 5, and 0.5 µg/mL) resulted in the complete abolition of the cytoprotective activity of metamizole. In the human osteosarcoma cell line, the cytotoxic effect of risedronate sodium was entirely eliminated in the presence of 50 µg/mL of metamizole. The cytoprotective and anti-apoptotic effect of metamizole in combination with risedronate sodium in the tested human and canine osteosarcoma cell lines indicates an urgent need for further in vivo studies to confirm or disprove the potential dose-dependent undesirable effect of such a therapy.

Biosynthesized Silver Nanoparticles from Cyperus conglomeratus Root Extract Inhibit Osteogenic Differentiation of Immortalized Mesenchymal Stromal Cells.

Silver nanoparticles (AgNPs) are a focus of huge interest in biological research, including stem cell research. AgNPs synthesized using Cyperus conglomeratus root extract have been previously reported but their effects on mesenchymal stromal cells have yet to be investigated. The aim of this study is to investigate the effects of C. conglomeratus-derived AgNPs on adipogenesis and osteogenesis of mesenchymal stromal cells. AgNPs were synthesized using C. conglomeratus root extract, and the phytochemicals involved in AgNPs synthesis were analyzed using gas chromatography-mass spectrometry (GCMS). The cytotoxicity of the AgNPs was tested on telomerase-transformed immortalized human bone marrow-derived MSCs-hTERT (iMSC3) and human osteosarcoma cell line (MG-63) using MTT and apoptosis assays. The uptake of AgNPs by both cells was confirmed using inductively coupled plasma-optical emission spectrometry (ICP-OES). Furthermore, the effect of AgNPs on iMSC3 adipogenesis and osteogenesis was analyzed using stain quantification and reverse transcription- quantitative polymerase chain reaction (RT-qPCR). The phytochemicals predominately identified in both the AgNPs and C. conglomeratus root extract were carbohydrates. The AgNP concentrations tested using MTT and apoptosis assays (0.5-64 µg/ml and 1,4 and 32 µg/ml, respectively) showed no significant cytotoxicity on iMSC3 and MG-63. The AgNPs were internalized in a concentration-dependent manner in both cell types. Additionally, the AgNPs exhibited a significant negative effect on osteogenesis but not on adipogenesis. C. conglomeratus-derived AgNPs had an impact on the differentiation capacity of iMSC3. Our results indicated that C. conglomeratus AgNPs and the associated phytochemicals could exhibit potential medical applications.

MiR-488-3p Represses Malignant Behaviors and Facilitates Autophagy of Osteosarcoma Cells by Targeting Neurensin-2.

Osteosarcoma (OS) is a primary bone sarcoma that primarily affects children and adolescents and poses significant challenges in terms of treatment. microRNAs (miRNAs) have been implicated in OS cell growth and regulation. This study sought to investigate the role of hsa-miR-488-3p in autophagy and apoptosis of OS cells. The expression of miR-488-3p was examined in normal human osteoblasts and OS cell lines (U2OS, Saos2, and OS 99-1) using RT-qPCR. U2OS cells were transfected with miR-488- 3p-mimic, and cell viability, apoptosis, migration, and invasion were assessed using CCK-8, flow cytometry, and Transwell assays, respectively. Western blotting and immunofluorescence were employed to measure apoptosis- and autophagy-related protein levels, as well as the autophagosome marker LC3. The binding sites between miR-488-3p and neurensin-2 (NRSN2) were predicted using online bioinformatics tools and confirmed by a dual-luciferase assay. Functional rescue experiments were conducted by co-transfecting miR-488-3p-mimic and pcDNA3.1-NRSN2 into U2OS cells to validate the effects of the miR-488-3p/NRSN2 axis on OS cell behaviors. Additionally, 3-MA, an autophagy inhibitor, was used to investigate the relationship between miR- 488-3p/NRSN2 and cell apoptosis and autophagy. miR-488-3p was found to be downregulated in OS cell lines, and its over-expression inhibited the viability, migration, and invasion while promoting apoptosis of U2OS cells. NRSN2 was identified as a direct target of miR-488-3p. Over-expression of NRSN2 partially counteracted the inhibitory effects of miR-488-3p on malignant behaviors of U2OS cells. Furthermore, miR- 488-3p induced autophagy in U2OS cells through NRSN2-mediated mechanisms. The autophagy inhibitor 3-MA partially reversed the effects of the miR-488-3p/NRSN2 axis in U2OS cells. Our findings demonstrate that miR-488-3p suppresses malignant behaviors and promotes autophagy in OS cells by targeting NRSN2. This study provides insights into the role of miR-488-3p in OS pathogenesis and suggests its potential as a therapeutic target for OS treatment.

Green synthesized chitosan-coated iron oxide nanocomposite using Cissus quadrangularis plant extract for antibacterial, antioxidant and anticancer applications

Metal oxide nanoparticles (NPs) have demonstrated their potential to eliminate a wide range of drug-resistant bacterial pathogens and cancer cells in recent years. Iron oxide (Fe3O4) NPs have received significant attention due to their wide range of uses in the field of biomedicine. In specific, biopolymer-encapsulated Fe3O4 NPs were applied in cancer therapy owing to their biocompatibility and biodegradability. The current investigation describes a bio-inspired approach to synthesize chitosan-based Fe3O4 nanocomposite (NC) employing Cissus quadrangularis (CQ) plant extract in a sustainable manner. The physicochemical, morphological, compositional, surface texture and thermal stability properties of the biopolymer-coated nanocomposites were characterised using FT-IR, UV–vis, XRD, XPS, HR-SEM, TEM, TGA, BET and TGA analyses. The antibacterial efficacy was examined in strains of Escherichia coli and Staphylococcus aureus, which verified that the growth of bacterial pathogens was inhibited by a substantial increase in the concentration of composites. The DPPH assay results were utilized to assess the free radical scavenging activity. The in vitro anticancer activity was found to be significant against the human osteosarcoma (MG-63) cell line, with an IC50 value of 54 ± 0.50 μg/mL. The enhanced biocompatibility of NCs was confirmed by cytotoxic testing on human RBCs and normal L929 fibroblast cells.

Characterization of Iron Oxide Nanotubes Obtained by Anodic Oxidation for Biomedical Applications-In Vitro Studies.

To improve the biocompatibility and bioactivity of biodegradable iron-based materials, nanostructured surfaces formed by metal oxides offer a promising strategy for surface functionalization. To explore this potential, iron oxide nanotubes were synthesized on pure iron (Fe) using an anodic oxidation process (50 V-30 min, using an ethylene glycol solution containing 0.3% NH4F and 3% H2O, at a speed of 100 rpm). A nanotube layer composed mainly of α-Fe2O3 with diameters between 60 and 70 nm was obtained. The effect of the Fe-oxide nanotube layer on cell viability and morphology was evaluated by in vitro studies using a human osteosarcoma cell line (SaOs-2 cells). The results showed that the presence of this layer did not harm the viability or morphology of the cells. Furthermore, cells cultured on anodized surfaces showed higher metabolic activity than those on non-anodized surfaces. This research suggests that growing a layer of Fe oxide nanotubes on pure Fe is a promising method for functionalizing and improving the cytocompatibility of iron substrates. This opens up new opportunities for biomedical applications, including the development of cardiovascular stents or osteosynthesis implants.

The exposure to extremely low frequency electromagnetic-fields inhibits the growth and potentiates the sensitivity to chemotherapy of bidimensional and tridimensional human osteosarcoma models

We previously established a thermodynamical model to calculate the specific frequencies of extremely low frequency-electromagnetic field (ELF-EMF) able to arrest the growth of cancer cells. In the present study, for the first time, we investigated the efficacy of this technology on osteosarcoma, and we applied a precise frequency of the electromagnetic field on three human osteosarcoma cell lines, grown as adherent cells and spheroids. We evaluated the antitumour efficacy of irradiation in terms of response to chemotherapeutic treatments, which is usually poor in this type of cancer. Importantly, the results of this novel combinatorial approach revealed that the specific exposure can potentiate the efficacy of several chemotherapeutic drugs, both on bidimensional and tridimensional cancer models. The effectiveness of cisplatinum, methotrexate, ifosfamide and doxorubicin was greatly increased by the concomitant application of the specific ELF-EMF. Moreover, our experiments confirmed that ELF-EMF inhibited the proliferation and modulated the mitochondrial metabolism of all cancer models tested, whereas mesenchymal cells were not affected. The latter finding is extremely valuable, given the importance of preserving the cell reservoir necessary for tissue regeneration after chemotherapy. Altogether, this novel evidence opens new avenues to the clinical applications of ELF-EMF in oncology.

The EphA2 Receptor Regulates Invasiveness and Drug Sensitivity in Canine and Human Osteosarcoma Cells.

Osteosarcoma is an aggressive bone cancer affecting both humans and dogs, often leading to pulmonary metastasis. Despite surgery and chemotherapy being the primary treatment modalities, survival rates remain low in both species, underscoring the urgent need for more efficacious therapeutic options. Accumulating evidence indicates numerous biological and clinical similarities between human and canine osteosarcoma, making it an ideal choice for comparative oncological research that should benefit both species. The EphA2 receptor has been implicated in controlling invasive responses across different human malignancies, and its expression is associated with poor prognosis. In this study, we utilized a comparative approach to match EphA2 functions in human and canine osteosarcoma models. Our objectives were to assess EphA2 levels and its pro-malignant action in osteosarcoma cells of both species. We found that EphA2 is overexpressed in most of both canine and human osteosarcoma cell lines, while its silencing significantly reduced cell viability, migration, and invasion. Moreover, EphA2 silencing enhanced the sensitivity of osteosarcoma cells to cisplatin, a drug commonly used for treating this cancer. Furthermore, inhibition of EphA2 expression led to a significant reduction in tumor development capability of canine osteosarcoma cells. Our data suggest that these EphA2 effects are likely mediated through various signaling mechanisms, including the SRC, AKT, and ERK-MAPK pathways. Collectively, our findings indicate that EphA2 promotes malignant behaviors in both human and canine osteosarcoma and that targeting EphA2, either alone or in combination with chemotherapy, could offer potential benefits to osteosarcoma patients.

Release of Exosomal PD-L1 in Bone and Soft Tissue Sarcomas and Its Relationship to Radiotherapy.

(1) Background: Exosomal PD-L1 has garnered attention owing to its role in instigating systemic immune suppression. The objective of this study is to elucidate whether bone and soft tissue sarcoma cells possess the capacity to secrete functionally active exosomal PD-L1 and whether radiotherapy (RT) induces the exosomal PD-L1 release. (2) Methods: Human osteosarcoma cell line 143B and human fibrosarcoma cell line HT1080 were utilized. Exosomes were isolated from the culture medium and blood via ultracentrifugation. The expression of PD-L1 on both tumor cells and exosomes was evaluated. The inhibitory effect on PBMC was employed to assess the activity of exosomal PD-L1. Post radiotherapy, changes in PD-L1 expression were compared. (3) Results: Exosomal PD-L1 was detected in the culture medium of tumor cells but was absent in the culture medium of PD-L1 knockout cells. Exosomal PD-L1 exhibited an inhibitory effect on PBMC activation. In tumor-bearing mice, human-derived exosomal PD-L1 was detected in the bloodstream. Following radiotherapy, tumor cells upregulated PD-L1, and human-derived exosomal PD-L1 were detected in the bloodstream. (4) Conclusions: Exosomal PD-L1 emanates from bone and soft tissue sarcoma cells and is disseminated into the circulatory system. The levels of PD-L1 in tumor cells and the release of exosomal PD-L1 were augmented after irradiation with RT.

Mild Hyperthermia-Induced Thermogenesis in the Endoplasmic Reticulum Defines Stress Response Mechanisms.

Previous studies reported that a mild, non-protein-denaturing, fever-like temperature increase induced the unfolded protein response (UPR) in mammalian cells. Our dSTORM super-resolution microscopy experiments revealed that the master regulator of the UPR, the IRE1 (inositol-requiring enzyme 1) protein, is clustered as a result of UPR activation in a human osteosarcoma cell line (U2OS) upon mild heat stress. Using ER thermo yellow, a temperature-sensitive fluorescent probe targeted to the endoplasmic reticulum (ER), we detected significant intracellular thermogenesis in mouse embryonic fibroblast (MEF) cells. Temperatures reached at least 8 °C higher than the external environment (40 °C), resulting in exceptionally high ER temperatures similar to those previously described for mitochondria. Mild heat-induced thermogenesis in the ER of MEF cells was likely due to the uncoupling of the Ca2+/ATPase (SERCA) pump. The high ER temperatures initiated a pronounced cytosolic heat-shock response in MEF cells, which was significantly lower in U2OS cells in which both the ER thermogenesis and SERCA pump uncoupling were absent. Our results suggest that depending on intrinsic cellular properties, mild hyperthermia-induced intracellular thermogenesis defines the cellular response mechanism and determines the outcome of hyperthermic stress.

In vitro Molecular Mechanisms of Anticancer Activity of Stevioside in Human Osteosarcoma Cell Lines (Sarcoma Osteogenic)

Abstract Introduction: Osteosarcoma (OS) is a rare and aggressive form of bone cancer that primarily affects the long bones of the body, such as the arms and legs. It is characterized by the uncontrolled growth of malignant cells in the bone tissue, leading to the formation of abnormal and painful bone masses. Steviol glycosides have been widely used as natural noncalorie sweeteners and are the collective name of the sweet substances found naturally in the plant Stevia rebaudiana, which is commonly called Stevia. Our study aimed to analyze the anticancer activity of Stevioside in OS. Materials and Methods: Stevioside was applied to OS cells, and the levels of Bcl xL, Bcl-2, and Bax were then estimated. The results of three separate studies, each carried out in triplicate, were expressed as the mean ± standard errors of the mean (SEM). One-way ANOVA was used for statistical analysis. Results: The findings showed that the effect of Stevioside on sarcoma osteogenic cells with mean ± SEM as 0.74 ± 0.05, 0.69 ± 0.09, 0.46 ± 0.09 for Bcl-xL gene, 0.98 ± 0.06, 0.58 ± 0.07, 0.5 ± 0.07 for Bcl-2 gene, and 1.2 ± 0.08, 1.45 ± 0.11, 1.67 ± 0.12 for Bax gene, respectively, when treated with untreated control cells. Conclusion: The study concludes its action against bone OS cells was significant with apoptotic induction. Stevia has a wide range of health benefits as well as being a plant-based diet it has less of side effects and promoting features even by intaking it daily along with other medicines.

Quantitative Analysis of Isopimpinellin from Ammi majus L. Fruits and Evaluation of Its Biological Effect on Selected Human Tumor Cells.

Ammi majus L. (Apiaceae) is a medicinal plant with a well-documented history in phytotherapy. The aim of the present work was to isolate isopimpinellin (5,8-methoxypsoralen; IsoP) from the fruit of this plant and evaluate its biological activity against selected tumor cell lines. The methanol extract obtained with the use of an accelerated solvent extraction (ASE) method was the most suitable for the quantitative analysis of coumarins in the A. majus fruit matrix. The coumarin content was estimated by RP-HPLC/DAD, and the amount of IsoP was found to be 404.14 mg/100 g dry wt., constituting 24.56% of the total coumarin fraction (1.65 g/100 g). This, along with the presence of xanthotoxin (368.04 mg/100 g, 22.36%) and bergapten (253.05 mg/100 g, 15.38%), confirmed A. majus fruits as an excellent source of these compounds. IsoP was isolated (99.8% purity) by combined liquid chromatography/centrifugal partition chromatography (LC/CPC) and tested for the first time on its antiproliferative activity against human colorectal adenocarcinoma (HT29, SW620), osteosarcoma (Saos-2, HOS), and multiple myeloma (RPMI8226, U266) cell lines. MTT assay results (96 h incubation) demonstrated a dose- and cell line-dependent decrease in cell proliferation/viability, with the strongest effect of IsoP against the Saos-2 cell line (IC50; 42.59 µM), medium effect against U266, HT-29, and RPMI8226 (IC50 = 84.14, 95.53, and 105.0 µM, respectively), and very weak activity against invasive HOS (IC50; 321.6 µM) and SW620 (IC50; 711.30 µM) cells, as well as normal human skin fibroblasts (HSFs), with IC50; 410.7 µM. The mechanistic study on the Saos-2 cell line showed that IsoP was able to reduce DNA synthesis and trigger apoptosis via caspase-3 activation. In general, IsoP was found to have more potency towards cancerous cells (except for HOS and SW620) than against healthy cells. The Selective Index (SI) was determined, underlining the higher selectivity of IsoP towards cancer cells compared to healthy cells (SI = 9.62 against Saos-2). All these results suggest that IsoP might be a promising molecule in the chemo-prevention and treatment of primary osteosarcoma.