Encapsulation of Artemisia scoparia extract in chitosan-myristate nanogel with enhanced cytotoxicity and apoptosis against hepatocellular carcinoma cell line (Huh-7)
Introduction
Hepatocellular carcinoma, also known as hepatic cancer, is one of the common cancers in men that leads to many deaths each year (Ramai et al., 2019). In 2018, a total of 841,000 cases of hepatocellular carcinoma with 263,000 deaths were reported worldwide (Le et al., 2019). Currently, surgical operation, chemo-therapeutic and ionizing radiation are the common strategies to treat hepatocellular carcinoma which have low efficacy and high toxicity on normal cells (Cai et al., 2019). In recent years, the natural products such as herbal extracts have attracted the attention of many researchers as alternative anti-cancer agents (Zhai et al., 2019). The herbal extracts are composed of various phytochemicals, many of which are considered as bioactive agents (Huynh et al., 2017). Moreover, herbal extracts contain secondary metabolites with anti-microbial, anti-cancer, anti-inflammatory and anti-oxidant effects (Seca and Pinto, 2018). Generally, there are more than 20,000 species of Artemisia worldwide. Some of the Artemisia species are used for the therapeutic applications, such as sedative factors, cough relief, anti-bacterial and anti-tumor agents, as well as for the malaria therapy (Moulavi et al., 2019). Among 20,000 Artemisia species, 34 species have been found in Iran. A. scoparia belongs to the Asteraceae family which has a high content of secondary metabolites such as flavonoids, terpenoids, and tannins (Naghavi et al., 2014).
The formulation of herbal extracts have gained huge attention to prevent evaporation, degradation of the active components and controlled release of the extract (Iannone et al., 2017). One of these formulation systems is the use of nanoscale delivery systems, which have shown increased biological activities of the extracts due to passive cell absorption strategy (Ong et al., 2017). Among nanomaterials for encapsulation, nanogels are highly suggested due to their high capacity of loading, high stability and release features (Kavetsou et al., 2019). From a structural point of view, the nanogels are three-dimensional compounds which are composed of interconnected polymer chains via self-assembly phenomenon (Abdel-Rashid et al., 2019). Nanogels can be synthesized from biodegradable synthetic materials. Among them some natural polymers such as chitosan (CS) have been used as suitable carrier for several therapeutic agents due to its low cost (Palaniraj et al., 2019). The most important characteristics of CS as a carrier system are biodegradability, biocompatibility, availability, safety, low immunogenicity and cationic charge (Chellappan et al., 2019). It has been shown in different studies that CS-based nanogels are used in tissue engineering, drug delivery and as a carrier for many macromolecules such as plant-derived molecules (Perez-Alvarez et al., 2019). Glucosamine units of CS have provided the hydroxyl and amino groups as the reactive sites for the cross-linking reactions (Ashrafi et al., 2019). The cross-linkers can modify the properties of CS hydrogels and cross-linked hydrogels can absorb a huge amount of different kinds of biological fluids or water (Li et al., 2019). Myristic acid (MA) is a non-toxic alternative cross-linker (C14H28O2) which is favorable to prepare CS nanogel (Nazem et al., 2016). In this study, the CS- myristate nanogel (CS-MA) was designed and synthesized as carrier for A. scoparia extract and its potential anti-cancer activity was evaluated against hepato-carcinoma cell line (Huh-7). CS- MA nanogel is composed of CS as backbone and MA as hydrophobic moiety which are self-assembled in aqueous environments. In this study, the extract of Artemisia scoparia, an Iranian indigenous plant species, was used for the first time for encapsulation studies.
Section snippets
Materials
Chitosan (CS) was purchased from Sigma (USA). The 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and myristic acid (MA) were obtained from Fluka (USA). Acetic acid, Tween 80, glycerol, ethanol and MTT dye were purchased from Merck (Germany). Artemisia scoparia (Herbarium NO. 1319) aerial parts dry powder and Hepato-carcinoma cell line (Huh-7, IBRC C10082) were purchased from Iranian Biological Resource Center. RPMI-1640 medium, fetal bovine serum (FBS) and trypsin-EDTA were purchased from
Characterization of CS-MA nanogel
The CS-MA nanogel was prepared by self-aggregation process through the modification of CS molecules by the hydrophobic MA. As a result, the free amine groups of CS were linked to the carboxylic groups of MA by EDC (Fig. 1).
The structural characterization obtained by FTIR spectroscopy for MA, CS, CS-MA, CS-MA-encapsulated extract and free extract are given in Fig. 2A–E, respectively. In Fig. 2A, the peaks observed at 3412 and 3469 cm−1 belonged to OH stretch, confirming the presence of alcoholic
Conclusion
In the current study, the CS-MA was synthesized using self-assembly process and it was characterized by FTIR, SEM and TEM. The mean diameter of the CS-MA nanogel was below 100 nm with uniform size distribution and spherical structure. The CS-MA nanogel system was then used as an encapsulating material for Artemisia extract in order to increase the cytotoxicity of the extract. Based on our findings, it is concluded that the CS-MA-encapsulated extract induced the apoptosis in Huh-7 cell line. To
CRediT authorship contribution statement
Sara Mosafer Haghighi: Data curation, Formal analysis, Methodology. Farzaneh Tafvizi: Methodology, Project administration, Data curation, Supervision, Writing - review & editing. Amir Mirzaie: Formal analysis, Methodology, Writing - original draft.
Declaration of Competing Interest
The authors declare that they have no conflict of interest.
Acknowledgements
The authors would like to acknowledge the laboratory of Islamic Azad University.
References (42)
- et al.
Mentha piperita essential oils loaded in a chitosan nanogel with inhibitory effect on biofilm formation against S. mutans on the dental surface
Carbohydr. Polym.
(2019) - et al.
Quercetin conjugated poly(beta-amino esters) nanogels for the treatment of cellular oxidative stress
Acta Biomater.
(2015) - et al.
Anti-tumor activity of wogonin, an extract from Scutellaria baicalensis, through regulating different signaling pathways
Chin. J. Nat. Med.
(2017) - et al.
Characterization and in vitro anticancer properties of chitosan-microencapsulated flavan-3-ols-rich grape seed extracts
Int. J. Biol. Macromol.
(2017) - et al.
Preparation, characterization and in vitro release study of carvacrol-loaded chitosan nanoparticles
Colloids Surf. B Biointerfaces
(2011) - et al.
Loss of Peter Pan protein is associated with cell cycle defects and apoptotic events
Biochimica et Biophysica Acta (BBA) -Molecular Cell Research
(2019) - et al.
Formulations of biodegradable Nanogel carriers with 5’-triphosphates of nucleoside analogs that display a reduced cytotoxicity and enhanced drug activity
J. Control. Release
(2007) - et al.
Biodegradable polymeric nanoparticles based drug delivery systems
Colloids Surf. B Biointerfaces
(2010) - et al.
Self-assembled glycol chitosan nanogels containing palmityl-acylated exendin-4 peptide as a long-acting anti-diabetic inhalation system
J. Control. Release
(2012) - et al.
Carboxymethyl chitosan-based nanogels via acid-labile ortho ester linkages mediated enhanced drug delivery
Int. J. Biol. Macromol.
(2019)
Formulation of essential oil-loaded chitosan-alginate nanocapsules
J. Food Drug Anal.
Chlorogenic acid- loaded calcium phosphate chitosan nanogel as biofilm degradative materials
Int. J. Biochem. Cell Biol.
Chitosan nanogels as nanocarriers of polyoxometalates for breast cancer therapies
Carbohydr. Polym.
Soft hydrogel based on modified chitosan containing P. granatum peel extract and its nano-forms: multiparticulate study on chronic wounds treatment
Int. J. Biol. Macromol.
Nanogels: an overview of properties, biomedical applications and obstacles to clinical translation
J. Control. Release
Polyplex Nanogel formulations for drug delivery of cytotoxic nucleoside analogs
J. Control. Release
Molecular targets of β-elemene, a herbal extract used in traditional Chinese medicine and its potential role in cancer therapy
A review Biomedicine & Pharmacotherapy
Encapsulation of Cuminum cyminum essential oils in chitosan-caffeic acid nanogel with enhanced antimicrobial activity against Aspergillus flavus
Ind. Crops Prod.
Nanogel loaded with surfactant based nanovesicles for enhanced ocular delivery of acetazolamide
Int. J. Nanomedicine
Effect of encapsulated Artemisia aucheri. L magnetic nanogel extract on shoulder block in rat
Drug Res. (Stuttg.)
Effect of polyunsaturated fatty acids on proliferation and survival of prostate cancer cells
PLoS One
Cited by (19)
Chitosan inhibits Penicillium expansum possibly by binding to DNA and triggering apoptosis
2024, International Journal of Biological MacromoleculesBiological effect of natural chitosan nanoparticles with transformed roots extract of Leonotis nepetifolia (L.) R.Br. in an in vitro model
2023, Industrial Crops and ProductsEnhanced anticancer effect of Artemisia turcomanica extract in niosomal formation on breast cancer cells: In-vitro study
2023, Nano-Structures and Nano-ObjectsChitosan nanogel for drug delivery and regenerative medicine
2023, Polysaccharide Hydrogels for Drug Delivery and Regenerative MedicineCharacterization and optimization of co-delivery Farnesol-Gingerol Niosomal formulation to enhance anticancer activities against breast cancer cells
2022, Journal of Drug Delivery Science and TechnologyCitation Excerpt :BCL2 is an anti-apoptotic gene most commonly present at the outer and internal mitochondrial membranes. Its weaker expression leads to cell death via the secretion of cytochrome C into the cytoplasm from the mitochondrial matrix that can activate caspase genes [88,89]. Cyclin E is crucial for controlling the phosphorylation of several proteins with key functions in cell growth.
Nanomaterials
2022, Phytochemical Nanodelivery Systems as Potential Biopharmaceuticals