materials and methods

1
Materials and methods To monitor the migration of the cells, PDMS (poly-dimethylsiloxane) devices with 100 µL capacity wells and 1 mm long, 10 µm wide and high channels were used. The main setup of this experiment has the cells on the left side with the attractant or chemo attractant on the right side of the channels. The devices were cleaned and sterilized by soaking in 70% ethanol. The devices were seeded with cells and monitored. Standard tissue culture plates served as the substratum for cell adhesion and standard tissue culture protocols were followed. Pictures of the wells and channels were taken with a Nikon Eclipse TI microscope at 100X magnification. The data was processed using AxioVision Rel. 4.8. Acknowledgments We thank Dr. Kambiz Alavi and Mohammadreza Jahangir Moghadam for heading the REU Program. We also thank NSF and UTA for funding and hosting the REU program. NSF grant # EEC-1156801, REU Site: Research Experiences for undergraduates in Sensors and Applications at University of Texas at Arlington Results Cell proliferation and migration were tested on two different cancer cell lines: PC3 (prostate cancer) and MDAMB 231 (breast cancer cells). Cytotoxicity and cell migration were assessed. Cytotoxicity study The cytotoxicity study tested the reaction of the cells with three nanoparticles, gold, iron oxide, and nickel respectively. Summary and conclusions In this study we observed the interaction between cells and nanoparticles. In particular, we investigated the effect of iron oxide particles on cell migration with different attractants. From the cytotoxicity study, the gross observations were that gold and iron oxide affected the cell growth less than nickel nanoparticles which were more cytotoxic. Iron oxide was chosen for migration studies because of its cell compatibility and its magnetic properties. The study suggests that there is migration of PC3 with nanoparticles over a long time. However, in the first 72-hour period, there seemed to be a decrease in cell proliferation and migration in both control and treated cells. Thus, the addition of nanoparticles could inhibit migration over a short time but increase migration over longer periods of time. On the other hand, PC3 and MDA-MB-231 cells lines treated with nanoparticles in the presence of a magnet and EGF on the attractant side, minor contamination was observed. The results suggest that the nanoparticles could have been contaminated since there was little or no contamination in the controls. Thus, proper sterilization of the nanoparticles is key for a successful experiment and the avoidance of contamination. Future work with nanoparticles include continuing the magnetic field migration experiments. Another experiment combining the EGF and magnetic attractants would be to test the potency of EGF while holding the cells in place with a magnet. Trouble shooting of the contamination by the way of testing for bacteria on agar plates and further testing of handling nanoparticles in the presence of cells. Michael Lau , Ian Mackinnon, Akash Sharma, Mahrukh Yasin, Cristian Hernandez, Smitha Rao, Victor Lin, J.-C. Chiao Electrical Engineering Summer REU, The University of Texas at Arlington, Arlington, Texas 76019 Literature cited “Demonstration of Cancer Cell Migration Using a Novel Microfluidic Device,” S. Rao, V. Lin, U. Tata, G. Raj, J.-T. Hsieh, K. Nguyen, and J.-C. Chiao, Journal of Nanotechnology for Engineering and Medicine, Vol. 1, No. 2, May 2010. “Study of Lung-Metastasized Prostate Cancer Cell Line Chemotaxsis to Epidermal Growth Factor with a Biomems Device,” U. Tata, S. M.N Rao, A. Sharma, K. Pabba, K. Pokhrel, B. Adhikari, V. K. Lin and J.-C. Chiao, Advances in Natural Sciences: Nanoscience and Nanotechnology, Vol. 3 No. 3, Sept. 2012. Figure 1: 100X magnification of the left and right wells with the channels connecting them. Further Information Nanoparticle Interaction and Migration in Cancer Cells Day 5 Figure 2: Panels A and B show PC3 cells with gold nanoparticles and after 5 days the cells appeared confluent. Panels C and D depict same conditions using iron oxide particles. Panels E and F with nickel showed necrosis of the cells. Migration studies The migration studies monitored the migration of the cells with Iron oxide nanoparticles. Iron oxide was chosen because of its compatibility of the particle as well as the magnetic characteristics. Each cell line was tested under two different attractants: EGF (epithelial growth factor) and magnets. Day 1 The migration studies revealed the following observations: PC3 • With EGF- the migration was steady for the control and the nanoparticles • With magnet- the nanoparticles showed signs of contamination on day 1 while the control showed signs of contamination on day 3. MDA-MB-231 • With EGF and magnet - the control showed signs of contamination after day 5. The nanoparticle side showed contamination after day 1. Figure 5: The average number of cells in each channel is depicted in the plot. It indicates that there was a gradual increase in migration over each 24-hour period. The decrease in the number of cells in PC3 on day 3 could be attributed to cells that had already moved out of the channels into the right side well. Figure 4: The PC3 migration is present in the EGF migration study for the control and nanoparticle sides. Figure 3: Panels A and B show similar confluency except with MDA-MB 231. Panels C and D display growth despite the clumping from the cells. Panels E and F show high cytotoxicity with nickel. B F B D F Figure 6: Comparing the healthy cells of PC3 of panel A to the contaminated and dead cells of B. The MDAMB231 cells shown in panels C and D share the same results as the PC3. Normal migration is observed in the control of the MDA-MB 231 EGF migration experiment E while dead cells and debris is observed in the nanoparticle side of the MDA-MB 231 EGF experiment in panel F. 1 2 3 4 5 0 2 4 6 8 10 12 14 16 18 Cell Count of PC3 with EGF Control PC3 Nanoparticle PC3 Time Elasped in Days Number of Cells Day 1 Day 5 Day 1 Day 5 A B C D E F Day 1 Day 5 Day 1 Day 5 Day 1 Day 5 Day 1 Day 5 A B C D E F A B C D E F Contact Michael Lau ([email protected]) or Smitha Rao ([email protected]). Abstract Current cancer treatments usually damage the surrounding normal cells as well. New studies have used nanoparticles as a method to battle cancer. In this study, gold, iron oxide and nickel nanoparticles were studied for their cytotoxic effect on cells. Prostate cancer (PC3) and breast cancer (MDA-MB-231) cell lines were used. From the cytotoxicity study, Iron oxide was chosen for its compatibility with the cells and for its magnetic properties. Migration studies with attractants examined the impact of iron oxide nanoparticles in cells under chemotaxis. Initial results suggest increase in cell migration after 72-hours of incubation. Introduction Nanoparticles have been used in previous studies in aiding cancer treatments. This study observed the affect that gold, iron oxide, and nickel nanoparticles have on cancer cells. The cytotoxicity study confirmed the possible use of iron oxide nanoparticles in migration studies. Thus, the migration studies observed chemotaxis of cancer cell using EGF(epithelial growth factor) as an attractant when the cells were treated with nanoparticles. Due to the magnetic properties of iron oxide the possibility of using magnetic fields as an attractant was also tested.

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Nanoparticle Interaction and Migration in Cancer Cells. Michael Lau , Ian Mackinnon, Akash Sharma, Mahrukh Yasin , Cristian Hernandez, Smitha Rao, Victor Lin, J.-C. Chiao Electrical Engineering Summer REU, The University of Texas at Arlington, Arlington, Texas 76019. Abstract - PowerPoint PPT Presentation

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Page 1: Materials  and methods

Materials and methodsTo monitor the migration of the cells, PDMS

(poly-dimethylsiloxane) devices with 100 µL capacity wells and 1 mm long, 10 µm wide and high channels were used. The main setup of this experiment has the cells on the left side with the attractant or chemo attractant on the right side of the channels. The devices were cleaned and sterilized by soaking in 70% ethanol. The devices were seeded with cells and monitored. Standard tissue culture plates served as the substratum for cell adhesion and standard tissue culture protocols were followed. Pictures of the wells and channels were taken with a Nikon Eclipse TI microscope at 100X magnification. The data was processed using AxioVision Rel. 4.8.

AcknowledgmentsWe thank Dr. Kambiz Alavi and Mohammadreza Jahangir Moghadam for heading the REU Program. We also thank NSF and UTA for funding and hosting the REU program. NSF grant # EEC-1156801,  REU Site: Research Experiences for undergraduates in Sensors and Applications at University of Texas at Arlington

ResultsCell proliferation and migration were tested on two

different cancer cell lines: PC3 (prostate cancer) and MDAMB 231 (breast cancer cells). Cytotoxicity and cell migration were assessed.

Cytotoxicity studyThe cytotoxicity study tested the reaction of the cells with

three nanoparticles, gold, iron oxide, and nickel respectively.

Summary and conclusionsIn this study we observed the interaction between cells and nanoparticles. In particular, we investigated the effect of iron oxide particles on cell migration with different attractants. From the cytotoxicity study, the gross observations were that gold and iron oxide affected the cell growth less than nickel nanoparticles which were more cytotoxic.

Iron oxide was chosen for migration studies because of its cell compatibility and its magnetic properties. The study suggests that there is migration of PC3 with nanoparticles over a long time. However, in the first 72-hour period, there seemed to be a decrease in cell proliferation and migration in both control and treated cells. Thus, the addition of nanoparticles could inhibit migration over a short time but increase migration over longer periods of time.

On the other hand, PC3 and MDA-MB-231 cells lines treated with nanoparticles in the presence of a magnet and EGF on the attractant side, minor contamination was observed. The results suggest that the nanoparticles could have been contaminated since there was little or no contamination in the controls. Thus, proper sterilization of the nanoparticles is key for a successful experiment and the avoidance of contamination.

Future work with nanoparticles include continuing the magnetic field migration experiments. Another experiment combining the EGF and magnetic attractants would be to test the potency of EGF while holding the cells in place with a magnet. Trouble shooting of the contamination by the way of testing for bacteria on agar plates and further testing of handling nanoparticles in the presence of cells.

Michael Lau, Ian Mackinnon, Akash Sharma, Mahrukh Yasin, Cristian Hernandez, Smitha Rao, Victor Lin, J.-C. Chiao

Electrical Engineering Summer REU, The University of Texas at Arlington, Arlington, Texas 76019

Literature cited“Demonstration of Cancer Cell Migration Using a Novel Microfluidic Device,” S. Rao, V. Lin, U. Tata, G. Raj, J.-T. Hsieh, K. Nguyen, and J.-C. Chiao, Journal of Nanotechnology for Engineering and Medicine, Vol. 1, No. 2, May 2010.

“Study of Lung-Metastasized Prostate Cancer Cell Line Chemotaxsis to Epidermal

Growth Factor with a Biomems Device,” U. Tata, S. M.N Rao, A. Sharma, K. Pabba, K. Pokhrel, B. Adhikari, V. K. Lin and J.-C. Chiao, Advances in Natural Sciences: Nanoscience and Nanotechnology, Vol. 3 No. 3, Sept. 2012.

Figure 1: 100X magnification of the left and right wells with the channels connecting them.

Further Information

Nanoparticle Interaction and Migration in Cancer Cells

Day 5

Figure 2: Panels A and B show PC3 cells with gold nanoparticles and after 5 days the cells appeared confluent. Panels C and D depict same conditions using iron oxide particles. Panels E and F with nickel showed necrosis of the cells.

Migration studiesThe migration studies monitored the migration of the cells

with Iron oxide nanoparticles. Iron oxide was chosen because of its compatibility of the particle as well as the magnetic characteristics. Each cell line was tested under two different attractants: EGF (epithelial growth factor) and magnets.

Day 1

The migration studies revealed the following observations:• PC3

• With EGF- the migration was steady for the control and the nanoparticles

• With magnet- the nanoparticles showed signs of contamination on day 1 while the control showed signs of contamination on day 3.

• MDA-MB-231• With EGF and magnet - the control showed signs of

contamination after day 5. The nanoparticle side showed contamination after day 1.

Figure 5: The average number of cells in each channel is depicted in the plot. It indicates that there was a gradual increase in migration over each 24-hour period. The decrease in the number of cells in PC3 on day 3 could be attributed to cells that had already moved out of the channels into the right side well.

Figure 4: The PC3 migration is present in the EGF migration study for the control and nanoparticle sides.

Figure 3: Panels A and B show similar confluency except with MDA-MB 231. Panels C and D display growth despite the clumping from the cells. Panels E and F show high cytotoxicity with nickel.

B F

B D F

Figure 6: Comparing the healthy cells of PC3 of panel A to the contaminated and dead cells of B. The MDAMB231 cells shown in panels C and D share the same results as the PC3. Normal migration is observed in the control of the MDA-MB 231 EGF migration experiment E while dead cells and debris is observed in the nanoparticle side of the MDA-MB 231 EGF experiment in panel F.

1 2 3 4 50

2

4

6

8

10

12

14

16

18

Cell Count of PC3 with EGF

Control PC3

Nanoparticle PC3

Time Elasped in Days

Num

ber o

f Cel

ls

Day 1

Day 5

Day 1

Day 5

A

B

C

D

E

F

Day 1

Day 5

Day 1

Day 5

Day 1

Day 5

Day 1

Day 5

A

B

C

D

E

F

A

B

C

D

E

F

Contact Michael Lau ([email protected]) or Smitha Rao ([email protected]).

AbstractCurrent cancer treatments usually damage the

surrounding normal cells as well. New studies have used nanoparticles as a method to battle cancer. In this study, gold, iron oxide and nickel nanoparticles were studied for their cytotoxic effect on cells. Prostate cancer (PC3) and breast cancer (MDA-MB-231) cell lines were used. From the cytotoxicity study, Iron oxide was chosen for its compatibility with the cells and for its magnetic properties. Migration studies with attractants examined the impact of iron oxide nanoparticles in cells under chemotaxis. Initial results suggest increase in cell migration after 72-hours of incubation.

IntroductionNanoparticles have been used in previous

studies in aiding cancer treatments. This study observed the affect that gold, iron oxide, and nickel nanoparticles have on cancer cells. The cytotoxicity study confirmed the possible use of iron oxide nanoparticles in migration studies. Thus, the migration studies observed chemotaxis of cancer cell using EGF(epithelial growth factor) as an attractant when the cells were treated with nanoparticles. Due to the magnetic properties of iron oxide the possibility of using magnetic fields as an attractant was also tested.