BEE VENOM FOR BREAST CANCER
What Is Breast Cancer?
It is a type of cancer, which originates in the breast due to the uncontrolled growth of harmful cells. In the beginning, there is a small tumor present that can be detected physically, in the form of a lump or can be diagnosed through an x-ray.
But not all of the breast lumps are cancer-prone as most of them are benign. A non-cancerous tumor is just an abnormal growth that does not manifest outside of the breast and is certainly not life-threatening at all. It is highly recommended to detect and get checked any sort of changes or lumps observed in the breast by a professional health care expert, to determine if it's malignant or benign and what are the risks associated with it.
How Does It Begin And What Are Its Types?
Breast cancer can begin in any part of the breast. Possible areas of its origin are:
Breast cancer can also be detected through mammogram screening. An early diagnosis can help initiate proper therapy before the cancer cells can play havoc.
How Is Breast Cancer Treated?Treatment for breast cancer helps in destroying the cancer responsible cells located within nearby tissues and the breast. Different treatment options are:
Surgery
It is perhaps the most common treatment, known as a mastectomy, through which the whole breast is removed. In another process, tissues and tumors are removed and it is known as lumpectomy.
Chemotherapy Makes use of different drugs to eliminate cancer cells and the powerful medicines prescribed can lead to fatigue, hot flashes, early menopause, hair loss and nausea for a long period.
Hormone therapy
In this type of treatment different drugs are used to control hormones that are responsible for aiding the growth of cancer responsible cells. Ovaries are also treated, through medication or surgery, to stop them from producing hormones. Side effects of these medicines can lead to vaginal dryness and hot flashes.
Targeted therapy Under this type of treatment, the natural immune system is strengthened to an extent that it can itself destroy the cancer cells. The medicines used, specifically target the cancer cells present in the breast.
A subject suffering from breast cancer can be given a combination of different treatments at the same time as hormone therapy, chemotherapy, radiation, and even surgery. A comprehensive regime is advised to get rid of the cancer cells completely.
It is important to mention here that all of the treatments, stated above, have severe side effects that can affect other organs of the body too. That is why medical professionals are looking for safe treatment options that do not disturb other functions of the body. In this case, bee venom is one of the most suitable options for treating cancer cells without damaging healthy cells.
Anti-Cancer Effect of Bee Venom on Human MDA-MB-231 Breast Cancer Cells Using Raman Spectroscopy
Abstract
In this clinical study, the apoptotic effect of BV (bee venom) on the human breast cancer cells was examined through principal component analysis (PCA) and Raman spectroscopy. Any sort of biochemical alterations in the cancer cells was evaluated after bee venom treatment. It was noticed that there were different results obtained for different treatment durations and concentrations.
A significant decrease in the Raman vibrations of the proteins and DNA was noticed in the cancer cells that were treated with bee venom having a dose of 3.0µg/mL for 48hours when compared with the control cells.
This finding highly suggests that degradation and denaturation of DNA fragmentation and proteins take place. Results of Raman spectroscopy were by the western blot assay and TUNEL. Thus, Raman spectroscopy along with PCA leads to a label-free and noninvasive tool for assessing cellular changes through anti-cancer properties of bee venom.
Introduction
To come up with effective and safe cancer treatment, it is important to understand the interaction of different anti-cancer mediators with affected cells. It is also well understood that anti-cancer therapy leads to apoptosis of target cancer cells. General Apoptosis is defined through cellular morphological changes like mitochondrial dysfunction, caspase activation, DNA cleavage, membrane blebbing and shrinking.
Western blot and MTT measure the protein synthesis and enzymatic activity, as the endpoints are linked with cell viability. These assays are labor-intensive, time-consuming, destructive, complicated and invasive. They also require a large amount of material to yield just a small quantity of end product. Furthermore, the molecular mechanism of these interactions cannot be evaluated through these assays, as fluorescent labels used during measurement can lead to changes in the biological conditions. Thus, a label-free and non-invasive analytical technique is required for real-time monitoring of the live cells.
The technique used in this clinical study – Raman spectroscopy – is a fast and noninvasive technique that does not even require any sampling before conducting analysis. It also provides detailed quantitative information about the chemical composition, molecular structure and interaction within cells through high selectivity and sensitivity. Raman spectroscopy has been used in different clinical studies to determine: the effects of Actinomycin D on the lung cancer line and to measure molecular changes (time-dependent) in the associated cells during apoptosis. But in this clinical study, Raman spectroscopy is being used for the first time, to determine the anti-cancer effect of bee venom on the breast cancer cells.
Method
Human breast cancer cells were acquired from the American Type Culture Collection, USA. Cells were cultured in the Dulbecco’s upgraded Eagle medium, which was supplemented with antibiotics and a 10% FBS (fetal bovine serum), in a humid atmosphere having 95% air and 5%CO2. PBMLs (human peripheral blood mononuclear lymphocytes) were later isolated from blood acquired from the donors through Ficoll-hypaque gradient centrifugation.
PBMLs obtained were set into 2 different brands: the upper band had PBMC, while the lower band had neutrophils. PBMLs were first harvested and then washed thrice in the PBS. Cells were subjected to bee venom having a composition: 50% melittin, 2.5% apamin, 2% MCD peptide, 2% amine, 1.5 % hyaluronidase, 0.5% histidine, 1% lyso-PLA, 12% PLA and 10% to 17% of norepinephrine, dopamine, acid phosphomonoesterase, invertase, glucosidase, and protease inhibitor – these ingredient were nearly 100% pure and subjected to incubation for some time of 24 hours.
Cells were seeded in 96 well plates and later treated with different doses of bee venom and saline for different periods. Once the treatment was done, cell viability was examined through the Cell Counting Kit-8. The apoptosis detection system was used by the guidelines of the supplier. Treated cells were washed with PBC twice and after that, a 1%parafromaldehyde (ice cold) was also added for 20 minutes. Cells were incubated for an hour at 37 degrees centigrade in a dark environment in 50µL of a buffer consisting of fluorescein-12-dUTP also having terminal deoxynucleotidyl transferase to label the 3'-OH ends of a fragmented DNA.
After this reaction, cells were washed again with a PBS solution and stained with a mixture of 250µg of RNase and 5.0µg/mL of BSA. Surface exposure of the phosphatidylserine by apoptotic cells was assessed through the flow cytometer after adding annexin V-V-FITC and stained at the same time with PI. Obtained samples were examined on the FACSort flow cytometer with the help of Lysis II software.
To carry out Raman spectroscopic measurement a gold-coated substrate was used. A layer of Cr was added so that there could be a proper adhesion between the glass substrate and the gold film. 10 individual cells were taken from each group. Raman spectra were acquired by using the SENTERRA confocal Raman system, equipped with a diode laser source (785nm and 100mW) having a resolution of 3cm. Raman signals were collected through a 100x MPLN N.A 0.9 Olympus that generates a laser spot of 1µm.
Preprocessing of data and Raman spectral acquisition was managed through OPUS software. To extract a pure sample of Raman spectra, an automated algorithm for removing the autofluorescence background was applied to the measured data. Raman measurements were noted down in an accumulation time of 60seconds in a range of 600cm-1 to 1750cm-1. Raman spectra of the selected cells were calculated as an average of the ten samples.
Results
To determine the cytotoxic effects of bee venom on the human normal and breast cancer cells, the viability of cells was determined through the CCK-8 assay. It was found that BV significantly hindered the proliferation of cancer cells. To further differentiate between the susceptibilities of normal cells and cancer cells, the effects of bee venom in the PBMLs was explored. As a result, BV did not present any sort of cytotoxic effect on the PBLMs at a dose of 12.4µg/mL after 72hours of incubation.
While the results gathered through Raman spectroscopy, in combination with PCA, demonstrated accordance with the results acquired through conventional biological assays like western blot assays, TUNEL, and viability. This technique is best for determining the apoptotic effects of bee venom through a label-free and non-invasive quality assessment of the cellular changes.
The results obtained from Raman spectroscopy, combined with PCA showed good agreement with the results obtained using conventional biological assays, such as viability, TUNEL, and western blot assays, as described in the previous section. Thus, this technique can be used to assess the apoptotic effect of BV using a noninvasive, label-free quality valuation of the cellular changes.
Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6238932/
Synergistic Effects Induced By Combined Treatments of Aqueous Extract of Propolis and Venom
Abstract
Breast cancer can be regarded as a heterogeneous disease that leads to a wide number of deaths. TNBC (triple-negative breast cancer) is one of the harshest cancers that results due to the complete failure of chemotherapy. That is why it is crucial to develop an alternative therapy that could control TNBC. In this clinical study, the synergistic effects of bee venom and propolis on the TNBC and MCF-7 cell lines were carefully evaluated.
The above stated, breast cancer cell lines were cultured in MEM and DMEM medium respectively and supplemented with fetal bovine serum 10%, penicillin-streptomycin and glutamine 1% at a temperature of 37 degrees centigrade in a 5% CO2 incubator. 0.01mg/ml of insulin was added to human breast cancer cell media while 1% of NEEA (non-essential amino acid) was added to the MCF-7 media.
Honey bee venom, used in this clinical study, was gathered through the electro stimulation process while 30% extract of propolis and bee venom was obtained from Bisboaca Simona Elena and the Dostetean Cornelia, respectively. Propolis treatment was done as follows: the aqueous solution was subjected to centrifugation at 5000xg for 10 minutes to get rid of any sort of sediment. Later the supernatant was filtered further to remove any contaminants. Bee venom was suspended in the DNAse/RNAse free water. Bee venom and propolis were further diluted with the help of DNAse/RNAse free water to a serial concentration of 0.1:100 to 30:50 ratios.
The synergistic effect induced by bee venom and propolis was evaluated through a combination therapy based on each treatment done according to IC50 for the two cancer cell lines. Subsequent serial dilution was also conducted similarly. MTT assay was performed to judge anti-cancer activity promoted by the two treatments done. For this reason, 2x104 cells were plated in the 96 well plates having 200µl of culturing media. Before starting the treatment, 200µl of fresh media containing the above described ascending concentrations of bee venom or propolis or their combination were also added to well plates. Incubation was done for 24hours and then the medium was moved away and finally, 100µl of MTT was added to well plates for 1 hour.
Result
Results found indicated that both of the cell lines demonstrated a similar level of sensitivity to propolis at a diluted solution of 0.072mg/ml to 0.09mg/ml. IC50 for BV on the MCF10 cells was at 1mg/ml which was 20 times higher than the 0.05mg/ml in the human breast cancer cells. After combining bee venom with propolis, a synergistic effect was observed at a higher concentration, nearly 5 and 2 times more powerful than the two treatments done alone.
Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4777451/
Conclusion
Both clinical studies, discussed above, clearly indicate that bee venom alone and in combination can help control the activity of human breast cancer cells. The important thing is to determine a perfect dose for treating subjects of breast cancer without any side effects.
Further reading:
Honeybee venom and melittin suppress growth factor receptor activation in HER2-enriched and triple-negative breast cancer (.pdf)
It is a type of cancer, which originates in the breast due to the uncontrolled growth of harmful cells. In the beginning, there is a small tumor present that can be detected physically, in the form of a lump or can be diagnosed through an x-ray.
But not all of the breast lumps are cancer-prone as most of them are benign. A non-cancerous tumor is just an abnormal growth that does not manifest outside of the breast and is certainly not life-threatening at all. It is highly recommended to detect and get checked any sort of changes or lumps observed in the breast by a professional health care expert, to determine if it's malignant or benign and what are the risks associated with it.
How Does It Begin And What Are Its Types?
Breast cancer can begin in any part of the breast. Possible areas of its origin are:
- In ducts that supply milk to the nipple. This type of cancer is known as duct cancer.
- In glands that produce milk. This type of cancer is known as lobular cancer.
- In tissues of the breasts. They are known as lymphomas and sarcomas.
- Less common types of breast cancers like angiosarcoma and phyllodes tumor.
Breast cancer can also be detected through mammogram screening. An early diagnosis can help initiate proper therapy before the cancer cells can play havoc.
How Is Breast Cancer Treated?Treatment for breast cancer helps in destroying the cancer responsible cells located within nearby tissues and the breast. Different treatment options are:
Surgery
It is perhaps the most common treatment, known as a mastectomy, through which the whole breast is removed. In another process, tissues and tumors are removed and it is known as lumpectomy.
Chemotherapy Makes use of different drugs to eliminate cancer cells and the powerful medicines prescribed can lead to fatigue, hot flashes, early menopause, hair loss and nausea for a long period.
Hormone therapy
In this type of treatment different drugs are used to control hormones that are responsible for aiding the growth of cancer responsible cells. Ovaries are also treated, through medication or surgery, to stop them from producing hormones. Side effects of these medicines can lead to vaginal dryness and hot flashes.
Targeted therapy Under this type of treatment, the natural immune system is strengthened to an extent that it can itself destroy the cancer cells. The medicines used, specifically target the cancer cells present in the breast.
A subject suffering from breast cancer can be given a combination of different treatments at the same time as hormone therapy, chemotherapy, radiation, and even surgery. A comprehensive regime is advised to get rid of the cancer cells completely.
It is important to mention here that all of the treatments, stated above, have severe side effects that can affect other organs of the body too. That is why medical professionals are looking for safe treatment options that do not disturb other functions of the body. In this case, bee venom is one of the most suitable options for treating cancer cells without damaging healthy cells.
Anti-Cancer Effect of Bee Venom on Human MDA-MB-231 Breast Cancer Cells Using Raman Spectroscopy
Abstract
In this clinical study, the apoptotic effect of BV (bee venom) on the human breast cancer cells was examined through principal component analysis (PCA) and Raman spectroscopy. Any sort of biochemical alterations in the cancer cells was evaluated after bee venom treatment. It was noticed that there were different results obtained for different treatment durations and concentrations.
A significant decrease in the Raman vibrations of the proteins and DNA was noticed in the cancer cells that were treated with bee venom having a dose of 3.0µg/mL for 48hours when compared with the control cells.
This finding highly suggests that degradation and denaturation of DNA fragmentation and proteins take place. Results of Raman spectroscopy were by the western blot assay and TUNEL. Thus, Raman spectroscopy along with PCA leads to a label-free and noninvasive tool for assessing cellular changes through anti-cancer properties of bee venom.
Introduction
To come up with effective and safe cancer treatment, it is important to understand the interaction of different anti-cancer mediators with affected cells. It is also well understood that anti-cancer therapy leads to apoptosis of target cancer cells. General Apoptosis is defined through cellular morphological changes like mitochondrial dysfunction, caspase activation, DNA cleavage, membrane blebbing and shrinking.
Western blot and MTT measure the protein synthesis and enzymatic activity, as the endpoints are linked with cell viability. These assays are labor-intensive, time-consuming, destructive, complicated and invasive. They also require a large amount of material to yield just a small quantity of end product. Furthermore, the molecular mechanism of these interactions cannot be evaluated through these assays, as fluorescent labels used during measurement can lead to changes in the biological conditions. Thus, a label-free and non-invasive analytical technique is required for real-time monitoring of the live cells.
The technique used in this clinical study – Raman spectroscopy – is a fast and noninvasive technique that does not even require any sampling before conducting analysis. It also provides detailed quantitative information about the chemical composition, molecular structure and interaction within cells through high selectivity and sensitivity. Raman spectroscopy has been used in different clinical studies to determine: the effects of Actinomycin D on the lung cancer line and to measure molecular changes (time-dependent) in the associated cells during apoptosis. But in this clinical study, Raman spectroscopy is being used for the first time, to determine the anti-cancer effect of bee venom on the breast cancer cells.
Method
Human breast cancer cells were acquired from the American Type Culture Collection, USA. Cells were cultured in the Dulbecco’s upgraded Eagle medium, which was supplemented with antibiotics and a 10% FBS (fetal bovine serum), in a humid atmosphere having 95% air and 5%CO2. PBMLs (human peripheral blood mononuclear lymphocytes) were later isolated from blood acquired from the donors through Ficoll-hypaque gradient centrifugation.
PBMLs obtained were set into 2 different brands: the upper band had PBMC, while the lower band had neutrophils. PBMLs were first harvested and then washed thrice in the PBS. Cells were subjected to bee venom having a composition: 50% melittin, 2.5% apamin, 2% MCD peptide, 2% amine, 1.5 % hyaluronidase, 0.5% histidine, 1% lyso-PLA, 12% PLA and 10% to 17% of norepinephrine, dopamine, acid phosphomonoesterase, invertase, glucosidase, and protease inhibitor – these ingredient were nearly 100% pure and subjected to incubation for some time of 24 hours.
Cells were seeded in 96 well plates and later treated with different doses of bee venom and saline for different periods. Once the treatment was done, cell viability was examined through the Cell Counting Kit-8. The apoptosis detection system was used by the guidelines of the supplier. Treated cells were washed with PBC twice and after that, a 1%parafromaldehyde (ice cold) was also added for 20 minutes. Cells were incubated for an hour at 37 degrees centigrade in a dark environment in 50µL of a buffer consisting of fluorescein-12-dUTP also having terminal deoxynucleotidyl transferase to label the 3'-OH ends of a fragmented DNA.
After this reaction, cells were washed again with a PBS solution and stained with a mixture of 250µg of RNase and 5.0µg/mL of BSA. Surface exposure of the phosphatidylserine by apoptotic cells was assessed through the flow cytometer after adding annexin V-V-FITC and stained at the same time with PI. Obtained samples were examined on the FACSort flow cytometer with the help of Lysis II software.
To carry out Raman spectroscopic measurement a gold-coated substrate was used. A layer of Cr was added so that there could be a proper adhesion between the glass substrate and the gold film. 10 individual cells were taken from each group. Raman spectra were acquired by using the SENTERRA confocal Raman system, equipped with a diode laser source (785nm and 100mW) having a resolution of 3cm. Raman signals were collected through a 100x MPLN N.A 0.9 Olympus that generates a laser spot of 1µm.
Preprocessing of data and Raman spectral acquisition was managed through OPUS software. To extract a pure sample of Raman spectra, an automated algorithm for removing the autofluorescence background was applied to the measured data. Raman measurements were noted down in an accumulation time of 60seconds in a range of 600cm-1 to 1750cm-1. Raman spectra of the selected cells were calculated as an average of the ten samples.
Results
To determine the cytotoxic effects of bee venom on the human normal and breast cancer cells, the viability of cells was determined through the CCK-8 assay. It was found that BV significantly hindered the proliferation of cancer cells. To further differentiate between the susceptibilities of normal cells and cancer cells, the effects of bee venom in the PBMLs was explored. As a result, BV did not present any sort of cytotoxic effect on the PBLMs at a dose of 12.4µg/mL after 72hours of incubation.
While the results gathered through Raman spectroscopy, in combination with PCA, demonstrated accordance with the results acquired through conventional biological assays like western blot assays, TUNEL, and viability. This technique is best for determining the apoptotic effects of bee venom through a label-free and non-invasive quality assessment of the cellular changes.
The results obtained from Raman spectroscopy, combined with PCA showed good agreement with the results obtained using conventional biological assays, such as viability, TUNEL, and western blot assays, as described in the previous section. Thus, this technique can be used to assess the apoptotic effect of BV using a noninvasive, label-free quality valuation of the cellular changes.
Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6238932/
Synergistic Effects Induced By Combined Treatments of Aqueous Extract of Propolis and Venom
Abstract
Breast cancer can be regarded as a heterogeneous disease that leads to a wide number of deaths. TNBC (triple-negative breast cancer) is one of the harshest cancers that results due to the complete failure of chemotherapy. That is why it is crucial to develop an alternative therapy that could control TNBC. In this clinical study, the synergistic effects of bee venom and propolis on the TNBC and MCF-7 cell lines were carefully evaluated.
The above stated, breast cancer cell lines were cultured in MEM and DMEM medium respectively and supplemented with fetal bovine serum 10%, penicillin-streptomycin and glutamine 1% at a temperature of 37 degrees centigrade in a 5% CO2 incubator. 0.01mg/ml of insulin was added to human breast cancer cell media while 1% of NEEA (non-essential amino acid) was added to the MCF-7 media.
Honey bee venom, used in this clinical study, was gathered through the electro stimulation process while 30% extract of propolis and bee venom was obtained from Bisboaca Simona Elena and the Dostetean Cornelia, respectively. Propolis treatment was done as follows: the aqueous solution was subjected to centrifugation at 5000xg for 10 minutes to get rid of any sort of sediment. Later the supernatant was filtered further to remove any contaminants. Bee venom was suspended in the DNAse/RNAse free water. Bee venom and propolis were further diluted with the help of DNAse/RNAse free water to a serial concentration of 0.1:100 to 30:50 ratios.
The synergistic effect induced by bee venom and propolis was evaluated through a combination therapy based on each treatment done according to IC50 for the two cancer cell lines. Subsequent serial dilution was also conducted similarly. MTT assay was performed to judge anti-cancer activity promoted by the two treatments done. For this reason, 2x104 cells were plated in the 96 well plates having 200µl of culturing media. Before starting the treatment, 200µl of fresh media containing the above described ascending concentrations of bee venom or propolis or their combination were also added to well plates. Incubation was done for 24hours and then the medium was moved away and finally, 100µl of MTT was added to well plates for 1 hour.
Result
Results found indicated that both of the cell lines demonstrated a similar level of sensitivity to propolis at a diluted solution of 0.072mg/ml to 0.09mg/ml. IC50 for BV on the MCF10 cells was at 1mg/ml which was 20 times higher than the 0.05mg/ml in the human breast cancer cells. After combining bee venom with propolis, a synergistic effect was observed at a higher concentration, nearly 5 and 2 times more powerful than the two treatments done alone.
Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4777451/
Conclusion
Both clinical studies, discussed above, clearly indicate that bee venom alone and in combination can help control the activity of human breast cancer cells. The important thing is to determine a perfect dose for treating subjects of breast cancer without any side effects.
Further reading:
Honeybee venom and melittin suppress growth factor receptor activation in HER2-enriched and triple-negative breast cancer (.pdf)