Effects of nano propolis on the proliferation and wound healing: in vitro study on A375 melanoma cancer cell line.
Effects of nano propolis on the proliferation and wound healing: in vitro study on A375 melanoma cancer cell line.
Nassim Valivand,1Shima Bidabad,2Shiva Ansari,3Hajie Lotfi,4,*Nematollah Gheibi,5
1. 1.Department of medical biotechnology, School of paramedical sciences, Qazvin University of Medical Sciences, Qazvin, Iran 2. Student Research Committee, Qazvin University of Medical Sciences, Qazvin, Iran 2. 1.Department of medical biotechnology, School of paramedical sciences, Qazvin University of Medical Sciences, Qazvin, Iran 2. Student Research Committee, Qazvin University of Medical Sciences, Qazvin, Iran 3. 1.Department of medical biotechnology, School of paramedical sciences, Qazvin University of Medical Sciences, Qazvin, Iran 2. Student Research Committee, Qazvin University of Medical Sciences, Qazvin, Iran 4. 1.Student Research Committee, Qazvin University of Medical Sciences, Qazvin, Iran 3 Cellular and Molecular Research Center, Research Institute for prevention of Non-Communicable Disease, Qazvin University of Medical Sciences, Qazvin, Iran 5. Department of Biochemistry and Genetics, Cellular and Molecular Research Center, Research Institute for Non-communicable Diseases, Qazvin University of Medical Sciences, Qazvin, Iran
Introduction: Skin cancer is the most common type of cancer. It is a disease in which malignant (cancerous) cells are formed in melanocytes.
Melanoma is the most aggressive skin tumor with malignancy, metastasis, and high mortality. Malignant melanoma comprises 2% of all cancers.
The success rate of current chemotherapy is relatively low because of multidrug resistance and its side effects. Therefore, there is a need to discover new, safe, and effective compounds and low-cost against melanoma.
Propolis, as an herbal compound (an emerging strategy for preventing or treating melanoma), can be used for cancer treatment due to flavonoids in the composition.
Bees produce Propolis by mixing the enzymes in their saliva with different parts of the plant. Propolis has anti-bacterial, anti-fungal, anti-parasitic, antioxidant, anti-inflammatory, and antitumor properties, as well as a booster for the immune system. An essential issue for using Propolis is overcoming to limitations of bulk Propolis and protein structural changes. Hence, converting Propolis into nanoparticles could become more reactive following the reduction of their surface area to volume.
Methods: 1. propolis extraction:
To extraction of Iranian Propolis (using 200gr Propolis with 1700ml ethanol 70%), the mixture was kept in the dark container for 14 days with moderate shaking. The resulting extract was filtered, and after rotary evaporating, it was freeze-dried. Propolis chemical contents were evaluated by X-ray crystallography (XRD).
2. nano propolis preparation:
Propolis dried (powder was dissolved in 80% alcohol) sonicated with Tween 80 and 20 and distilled water. Then characterization of nanoparticles was determined by DLS and Zeta potential.
3. Cell viability: first A375 cells (5*103 ) were seeded on a 96-well plate with high glucose DMEM containing 10% FBS (fetal bovine serum) and 1% Pen Strep for 24h. Then, the cells were treated with different concentrations of Propolis for 24 and 48h. Negative and positive control contained medium and 15% DMSO, respectively. MTT solution was added to each well and incubated for 24h. Next, DMSO was added to dissolve the blue formazan crystals. Absorbance was read at 570 and 630 nm by a microplate reader.
4. wound healing:
After seeding the cells in the 12-well plate and receiving 80% confluency, the cells were treated with IC50 concentration and a control sample. Then, a scratch was created in each well using a blue tip, and the wells were imaged for 48h.
Results: 1.Nano propolis characterize:
The DLS showed particle sizes from 1 to 40 nm, and the zeta potential determined the surface charge to be -110, which prevents aggregation.
2.MTT assay:
Nanoparticle-propolis had different effects in two time periods of 24 and 48 hours with concentrations of 10, 20, 40, 50, 60, 70, 80, 90, and 100(µl/ml). The IC50 in these two times was shown as 86(µl/ml) and 60(µl/ml), respectively. Compared to the control group, cell viability decreased significantly in 48 hours. Therefore, the effective concentration and the optimal time to continue the study were 60(µ l/ml) and 48 h, respectively.
3.Wound healing:
The wound width in the control group changed from 0.5 to 0.1(µm) from 0 to 48h compared with the IC50-treated group (with no changes). The correlation between time and wound healing was negative, with a coefficient of -0.84.
Conclusion: The study of nano propolis on the A375 cancer cell line showed that it could have cytotoxic effects in a dose-dependent manner, which was proven using the MTT test. in addition, the wound healing test also showed that nano propolis could prevent cell migration. As a result, nano propolis can be a suitable candidate as a therapeutic nutritional supplement along with chemotherapy drugs. Of course, extensive clinical studies are needed to confirm the positive effects of this natural product.