- Email: [email protected]
Poly-L-lysine modified ITO surface for enhanced cell growth
Available online at www.sciencedirect.com
ScienceDirect Materials Today: Proceedings 5 (2018) 11083–11088
www.materialstoday.com/proceedings
NanoThailand2016
Poly-L-lysine modified ITO surface for enhanced cell growth Suthiwan Udomrata, Supeecha Kumkateb, Theeraporn Puntheeranurakb,*, Tanakorn Osotchana a
Materials Science and Engineering program, and Capability Unit for Nanoscience and Nanotechnology, Faculty of Science, Mahidol University, Bangkok, THAILAND, 10400 b Department of Biology, Faculty of Science, Mahidol University, Bangkok, THAILAND, 10400
Abstract Material surface properties are considered as critical factor for the study of in Vitro cell growth and activities. It provides advantages not only for study of cellular activity but also for biomedical technology such as tissue engineering. Neuron is a typical cell type widely used to examine cell communications, alignments and differentiations. In the present study, a simple pattern of indium tin oxide (ITO) for neuron culture is introduced. Circular inter-digitated design of ITO platform was fabricated by using simple photolithography techniques. In order to enhance cell attachment, the fabricated electrode surface was modified by poly-L-lysine, a positive charge molecule that allows binding site for cell adhesion. SH-SY5Y cell, a human neuroblastoma cell line, was cultured on the modified surface. Results show that cells preferably adhere on the ITO surface while some of them align itself according to the ITO pattern. In addition, the possibility of fabricated nanostructure platform can indicate the potential control of cell precisely guidance and pattern on surface.
© 2017 Elsevier Ltd. All rights reserved. Selection and/or Peer-review under responsibility of The 5th Thailand International Nanotechnology Conference (NanoThailand2016). Keywords: SH-SY5Y; ITO Interdigitated; PLL-coated surface; Cellular platform; Cell adhesion.
___________ * Corresponding author.
E-mail address: [email protected]
2214-7853 © 2017 Elsevier Ltd. All rights reserved. Selection and/or Peer-review under responsibility of The 5th Thailand International Nanotechnology Conference (NanoThailand2016).
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
1. Introduction In recently decade, numerous designs of cellular platform have been intensively develop ranging from a simple glass platform to sophisticate gold-incorporated platform in order to observe and study cellular activities as well as cellular response to specific stimulators. The outstanding properties of the cellular platform for study of cellular activities is that it allows cellular investigation in a non-invasive manner while allow the researchers investigate those cell response for a long period of times. Development of the cellular platform provides many advantages for use, e.g. it grants the new tools for drug screening effect and allows new technique to incorporating cell into implant (1). Cell growth and proliferation on the certain design surface is very crucial. Several research groups develop and fabricate the new platform types from various materials such as biocompatible polymer (2), metal and metal oxide (3), and biocompatible ceramics (4). Among those metal oxide semiconductors, indium-tin-oxide (ITO) is very attractive material since it is non-toxic to cell (5). ITO was introduced as a new electrode platform for cell culture and shows numerous potential uses such as observing cell exocytosis (5) and inducing surface for cell differentiation (6). In addition, the transparent property of ITO makes it capable to observe cell under light microscope. SH-SY5Y, a delicate human neuroblastoma cell line, was intensively studied not only for cell-to-cell communication aspect but control of cell activities also in attractive points. This excitable cell can be induced to differentiate into neuron by several methods, for example, electrical field (6), growth factor, and chemical agent like vitamin A (7). Since this cell is comparable to the neuron cell in morphology, physiology and molecular functions, it is widely used as a model study in neurological disease, therapeutic effect on nervous system, healing and regeneration of the nervous system, and so on. In this work, we develop a simple indium-tin-oxide (ITO) base-platform for cell culture by using photolithography technique. SH-SY5Y cell line was culture on the fabricated platform and observes cell growth. Effect of surface properties as well as surface modification for cell growth is discussed. 2. 2. Material and Method 2.1 Cell culture SH-SY5Y cell line was receipted from Cell Line Service (CLS), Germany. Cells were continued culture until the passage number 65. Culture was supplied with high glucose DMEM–Dulbecco's Modified Eagle Medium (DMEM), 2.5% foetal bovine serum (FBS), 1mM sodium pyruvate, 1% of penicillin and streptomycin (all cell culture reagents were perched from Biochrome, Germany). Culture was controlled at 37 °C and 5% CO2 atmosphere. Cells were subcultured every 7 days (70-80% confluent). In the experiments, cells were cultured for 5 days before seeding. 2.2 Fabrication of Indium-tin-oxide platform for cell culture Commercial ITO coated on glass substrate was cut into small slides. The slides then were cleaned thoroughly before patterning with photoresist dry film by the means of photolithography process and etched by a simple wet etching process with hydrochloric acid. Patterned ITO electrodes were observed by both optical and scanning electron microscope (SEM). In order to create a vial for cell culture, the patterned ITO slide was connected with glass culture tube and polydimethylsiloxane (PDMS) was used to seal and connected the two parts. 2.3 Surface modification and cell culture in ITO vial ITO vial for cell cultures were cleaned and steriled by dry heat at 180 °C for 2 hours. Poly-L-lysine (PLL) at concentration of 0.1% was used to modify the surface to enhance cell adhesion and growth by deposition on the surface for 15 minutes and wash thoroughly before use. Cells were seeded at density of 200 cell/mm2. After seeded cell into the ITO vial, the cell culture medium was changed every day due to a small volume of the medium that
Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
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may not sufficiently provide nutrient for the cells. Cells growth on the surface was observed under optical microscope. 3. 3. Results and Discussions 3.1 Patterned geometry and ITO surface Circular interdigitated was designed to pattern ITO surface for cell culture. Circular design was formerly introduced by Hsiung (8) to reduce sharp corner and increase sensing area. The interdigitated pattern shows finger size at the center is 1,000 μm, the interdigitated fingers size is about 500 μm, and interspacing gap size is 500 μm. The fabricated ITO surface for cell culture is showed in Fig. 1. SEM images revel that there are some defects of the interdigitated figures, for example, the interdigitated finger is not had sharp edge and some areas were over etching. It is well known that the etching process can affect on the finished surface pattern (9) thus it is difficult to control the exactly size of the fingers. However, by adjusting step and time of etching, the overall of the ITO pattern can be controlled.
Fig. 1. ITO platform geometry indicating a) two couples of circular interdigitated design, b) SEM image showing interdigitated finger size and interspacing gap while arrowheads indicating the pattern defect due to fast etching rate, and c) ITO vial for cell culture consisting of ITO interdigiated surface and glass tube. The vial was placed in sterile petri dish to protect the culture from contamination during transfer.
3.2 Effect of surface treatment on cell growth and proliferation Several extracellular matrix molecules were used to modify the surface providing the specific surface properties for cell growth such as cell adhesion, migration, and proliferation (10, 11). Results clearly show that PLL modified ITO surface increase cell adhesion and allow cell proliferation comparing with bare ITO surface. Unsurprisingly,
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
cells grow in the vial of un-treated surface clump and growth in multilayer form (figure 2A and 2B) especially on the glass region, while on the bare ITO region, cell can spread and growth well. It was noticed that cells selective growth and align following the ITO area on both of bare and PLL modifiedsurface whereas on the glass region especially bare ITO platform, cells grow and clump into small cluster. In generally SH-SY5Y cell is semi-adherent cell which mean cell need to adhere on the surface for cell growth and proliferation but cell adhesion ability to surface is not strong. In this case, cells may try to adhere on the glass surface but just only few of them can spread on the surface. Hence most of cells that cannot find some area to occupy but growth on the other cell, which can observe when, cell over growth beyond confluence. According to the proliferation curve, it was noticed that cell grown on bare ITO surface have delay growth compare to the cell that grown on PLL modified-ITO surface. It is well known that cell proliferation can be influent by several factors especially contact inhibition (12, 13). Consider cell in the cluster that cell density is very high thus cell proliferation might be affect by contact inhibition and dominantly observe on proliferation of cell grown on bare ITO surface as illustrated in figure 3. The ITO on glass substrate, which possesses negatively charge of hydroxyl group (OH) these negatively charge provides anchor site for positively charge PLL (14, 15), thus the PLL molecules should evenly distribute on the entire surface. The effect of surface treatment is clearly showed in figure 2, cell growth on the PLL modified surface show better adhesion and proliferation on ITO region than glass surface of both the treat and non-treated surface. The reasons hindrance this phenomenon may involve in the glass changing its surface charge after acid etching especially removing of photoresist dry film by soak in sodium hydroxide (NaOH) Since PLL possesses positively charge that attractive for negatively charge of both glass and ITO surface while allowing the positive anchor site for cell adhesion. Typically cell membrane consists of negative charge protein thus cell may prefer to adhere and spread on the positive charge modified surface rather than on the neutral or negatively charge surface. Consideration of cells selectively grows on the ITO surface, as mention before, the negatively charge of glass surface maybe neutralized by Na+, the ITO surface at that time was covered with the photoreist dry film thus the surface charge of ITO was less altered. So the charge on surface was preserved and allows adsorption of PLL. Besides that ITO surface itself is a composition of metal that provide some of positively charge allowing cell adhesion on the surface. Although cells can adhere and spread on the bare ITO region but the amount of cell adhere over time is less than the cell grow on the treated surface which implied that the anchor site for cell adhesion is not enough. Moreover, adsorption of PLL on the negatively charge surface maybe in monolayer. PLL and other protein molecules are classified into nanoscale regime molecules thus, in this experiment, we also create a nanoscale roughness surface for cell adhesion. However, the effect of this roughness and its properties should have more intensively study in the future.
Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
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Fig. 2. Cell grow on ITO platform showing cell adhere on a, b) bare ITO surface and c,d) on PLL-modified ITO surface after seeding for 1 day and 3 days, repectively.
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
Fig. 3 Amount of cell grown on bare and PLL modified-ITO platform. Discrepancy amount of cell in both conditions is noticed at day 3 after seeding and consistent with fig. 2. 4. Conclusions We successfully demonstrate that biological soft tissue can be grown and incorporated in the metal oxide basesurface by using simple method fabrication. This experiment reveals the role of indium–tin-oxide surface treatment on cell growth and proliferation. Utilizing of surface modification changes surface properties, we can selective and control of cell growth and alignment on the surface by changing surface charge or coating with poly-L-lysine. PLL not only plays the important role for cell adhesion and growth but its monolayer adsorption may also provide nanosurface roughness, which should be intensively study in the future. References [1] AFM. Johnstone, GW. Gross, DG. Weiss, OHU. Schroeder, A. Gramowski, TJ Shafer. NeuroToxicol. 31 (2010) 331-50. [2] F. Pires, Q. Ferreira, CAV. Rodrigues, J. Morgado, FC. Ferreira, Biochimica Biophysica Acta (BBA) - General Subjects. (2015) 1158-68. [3] S. Ahadian, J. Ramon-Azcon, S. Ostrovidov, G. Camci-Unal, Hosseini V, Kaji H. Lab on a Chip. 12 (2012) 3491-503. [4] C Prakash , HK Kansal, BS Pabla, S. J Puri. Mater. Engineer. Perform. 24 (2015) 3622-33. [5] H. Zhao, L. Li, H-J Fan, F. Wang, L-M Jiang, P-G He. Molec. Cell. Biochem. (2012) 309-13. [6] H. Zhao, A. Steiger, M. Nohner, H. Ye. PLOS ONE. 10 (2015). [7] M. Encinas, M. Iglesias, Y. Liu, Wang H, A. Muhaisen, V. J. Ceña. Neurochem. 75 (2000) 991-1003. [8] L-C Hsiung, C-H Yang, C-L Chiu, C-L Chen, Y. Wang, H. Lee. Biosensors and Bioelectronics. 24 (2008) 869-75. [9] CJ. Huang, YK. Su, SL. Wu. Mater. Chem. Phys. 84 (2004) 146-50. [10] OG. Alamdari, E. Seyedjafari, M. Soleimani, N. Ghaemi. J Avicenna. Medic. Biotechnol. 5 (2013) 234-40. [11] S Joo, Yeon J. Kim, E. Lee, N. Hong, W. Sun, Y. Nam. Scientif. Rep. 5 (2015). [12] M. Abercrombie. In Vitro. 6 (1970) 128-42. [13] A. Swat, I. Dolado, JM. Rojas, AR. Nebreda. Molec. Cellul. Biol. 29 (2009) 3332-43. [14] J. Robertus, WR. Browne, BL. Feringa. Chemic. Soc. Rev. 39 (2010) 354-78. [15] R. Wu, Y. Li, Q. Wang, J. Yu, F. Jiang, F. Wang. RSC Advances. 2 (2012) 9887-93.
ScienceDirect Materials Today: Proceedings 5 (2018) 11083–11088
www.materialstoday.com/proceedings
NanoThailand2016
Poly-L-lysine modified ITO surface for enhanced cell growth Suthiwan Udomrata, Supeecha Kumkateb, Theeraporn Puntheeranurakb,*, Tanakorn Osotchana a
Materials Science and Engineering program, and Capability Unit for Nanoscience and Nanotechnology, Faculty of Science, Mahidol University, Bangkok, THAILAND, 10400 b Department of Biology, Faculty of Science, Mahidol University, Bangkok, THAILAND, 10400
Abstract Material surface properties are considered as critical factor for the study of in Vitro cell growth and activities. It provides advantages not only for study of cellular activity but also for biomedical technology such as tissue engineering. Neuron is a typical cell type widely used to examine cell communications, alignments and differentiations. In the present study, a simple pattern of indium tin oxide (ITO) for neuron culture is introduced. Circular inter-digitated design of ITO platform was fabricated by using simple photolithography techniques. In order to enhance cell attachment, the fabricated electrode surface was modified by poly-L-lysine, a positive charge molecule that allows binding site for cell adhesion. SH-SY5Y cell, a human neuroblastoma cell line, was cultured on the modified surface. Results show that cells preferably adhere on the ITO surface while some of them align itself according to the ITO pattern. In addition, the possibility of fabricated nanostructure platform can indicate the potential control of cell precisely guidance and pattern on surface.
© 2017 Elsevier Ltd. All rights reserved. Selection and/or Peer-review under responsibility of The 5th Thailand International Nanotechnology Conference (NanoThailand2016). Keywords: SH-SY5Y; ITO Interdigitated; PLL-coated surface; Cellular platform; Cell adhesion.
___________ * Corresponding author.
E-mail address: [email protected]
2214-7853 © 2017 Elsevier Ltd. All rights reserved. Selection and/or Peer-review under responsibility of The 5th Thailand International Nanotechnology Conference (NanoThailand2016).
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
1. Introduction In recently decade, numerous designs of cellular platform have been intensively develop ranging from a simple glass platform to sophisticate gold-incorporated platform in order to observe and study cellular activities as well as cellular response to specific stimulators. The outstanding properties of the cellular platform for study of cellular activities is that it allows cellular investigation in a non-invasive manner while allow the researchers investigate those cell response for a long period of times. Development of the cellular platform provides many advantages for use, e.g. it grants the new tools for drug screening effect and allows new technique to incorporating cell into implant (1). Cell growth and proliferation on the certain design surface is very crucial. Several research groups develop and fabricate the new platform types from various materials such as biocompatible polymer (2), metal and metal oxide (3), and biocompatible ceramics (4). Among those metal oxide semiconductors, indium-tin-oxide (ITO) is very attractive material since it is non-toxic to cell (5). ITO was introduced as a new electrode platform for cell culture and shows numerous potential uses such as observing cell exocytosis (5) and inducing surface for cell differentiation (6). In addition, the transparent property of ITO makes it capable to observe cell under light microscope. SH-SY5Y, a delicate human neuroblastoma cell line, was intensively studied not only for cell-to-cell communication aspect but control of cell activities also in attractive points. This excitable cell can be induced to differentiate into neuron by several methods, for example, electrical field (6), growth factor, and chemical agent like vitamin A (7). Since this cell is comparable to the neuron cell in morphology, physiology and molecular functions, it is widely used as a model study in neurological disease, therapeutic effect on nervous system, healing and regeneration of the nervous system, and so on. In this work, we develop a simple indium-tin-oxide (ITO) base-platform for cell culture by using photolithography technique. SH-SY5Y cell line was culture on the fabricated platform and observes cell growth. Effect of surface properties as well as surface modification for cell growth is discussed. 2. 2. Material and Method 2.1 Cell culture SH-SY5Y cell line was receipted from Cell Line Service (CLS), Germany. Cells were continued culture until the passage number 65. Culture was supplied with high glucose DMEM–Dulbecco's Modified Eagle Medium (DMEM), 2.5% foetal bovine serum (FBS), 1mM sodium pyruvate, 1% of penicillin and streptomycin (all cell culture reagents were perched from Biochrome, Germany). Culture was controlled at 37 °C and 5% CO2 atmosphere. Cells were subcultured every 7 days (70-80% confluent). In the experiments, cells were cultured for 5 days before seeding. 2.2 Fabrication of Indium-tin-oxide platform for cell culture Commercial ITO coated on glass substrate was cut into small slides. The slides then were cleaned thoroughly before patterning with photoresist dry film by the means of photolithography process and etched by a simple wet etching process with hydrochloric acid. Patterned ITO electrodes were observed by both optical and scanning electron microscope (SEM). In order to create a vial for cell culture, the patterned ITO slide was connected with glass culture tube and polydimethylsiloxane (PDMS) was used to seal and connected the two parts. 2.3 Surface modification and cell culture in ITO vial ITO vial for cell cultures were cleaned and steriled by dry heat at 180 °C for 2 hours. Poly-L-lysine (PLL) at concentration of 0.1% was used to modify the surface to enhance cell adhesion and growth by deposition on the surface for 15 minutes and wash thoroughly before use. Cells were seeded at density of 200 cell/mm2. After seeded cell into the ITO vial, the cell culture medium was changed every day due to a small volume of the medium that
Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
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may not sufficiently provide nutrient for the cells. Cells growth on the surface was observed under optical microscope. 3. 3. Results and Discussions 3.1 Patterned geometry and ITO surface Circular interdigitated was designed to pattern ITO surface for cell culture. Circular design was formerly introduced by Hsiung (8) to reduce sharp corner and increase sensing area. The interdigitated pattern shows finger size at the center is 1,000 μm, the interdigitated fingers size is about 500 μm, and interspacing gap size is 500 μm. The fabricated ITO surface for cell culture is showed in Fig. 1. SEM images revel that there are some defects of the interdigitated figures, for example, the interdigitated finger is not had sharp edge and some areas were over etching. It is well known that the etching process can affect on the finished surface pattern (9) thus it is difficult to control the exactly size of the fingers. However, by adjusting step and time of etching, the overall of the ITO pattern can be controlled.
Fig. 1. ITO platform geometry indicating a) two couples of circular interdigitated design, b) SEM image showing interdigitated finger size and interspacing gap while arrowheads indicating the pattern defect due to fast etching rate, and c) ITO vial for cell culture consisting of ITO interdigiated surface and glass tube. The vial was placed in sterile petri dish to protect the culture from contamination during transfer.
3.2 Effect of surface treatment on cell growth and proliferation Several extracellular matrix molecules were used to modify the surface providing the specific surface properties for cell growth such as cell adhesion, migration, and proliferation (10, 11). Results clearly show that PLL modified ITO surface increase cell adhesion and allow cell proliferation comparing with bare ITO surface. Unsurprisingly,
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
cells grow in the vial of un-treated surface clump and growth in multilayer form (figure 2A and 2B) especially on the glass region, while on the bare ITO region, cell can spread and growth well. It was noticed that cells selective growth and align following the ITO area on both of bare and PLL modifiedsurface whereas on the glass region especially bare ITO platform, cells grow and clump into small cluster. In generally SH-SY5Y cell is semi-adherent cell which mean cell need to adhere on the surface for cell growth and proliferation but cell adhesion ability to surface is not strong. In this case, cells may try to adhere on the glass surface but just only few of them can spread on the surface. Hence most of cells that cannot find some area to occupy but growth on the other cell, which can observe when, cell over growth beyond confluence. According to the proliferation curve, it was noticed that cell grown on bare ITO surface have delay growth compare to the cell that grown on PLL modified-ITO surface. It is well known that cell proliferation can be influent by several factors especially contact inhibition (12, 13). Consider cell in the cluster that cell density is very high thus cell proliferation might be affect by contact inhibition and dominantly observe on proliferation of cell grown on bare ITO surface as illustrated in figure 3. The ITO on glass substrate, which possesses negatively charge of hydroxyl group (OH) these negatively charge provides anchor site for positively charge PLL (14, 15), thus the PLL molecules should evenly distribute on the entire surface. The effect of surface treatment is clearly showed in figure 2, cell growth on the PLL modified surface show better adhesion and proliferation on ITO region than glass surface of both the treat and non-treated surface. The reasons hindrance this phenomenon may involve in the glass changing its surface charge after acid etching especially removing of photoresist dry film by soak in sodium hydroxide (NaOH) Since PLL possesses positively charge that attractive for negatively charge of both glass and ITO surface while allowing the positive anchor site for cell adhesion. Typically cell membrane consists of negative charge protein thus cell may prefer to adhere and spread on the positive charge modified surface rather than on the neutral or negatively charge surface. Consideration of cells selectively grows on the ITO surface, as mention before, the negatively charge of glass surface maybe neutralized by Na+, the ITO surface at that time was covered with the photoreist dry film thus the surface charge of ITO was less altered. So the charge on surface was preserved and allows adsorption of PLL. Besides that ITO surface itself is a composition of metal that provide some of positively charge allowing cell adhesion on the surface. Although cells can adhere and spread on the bare ITO region but the amount of cell adhere over time is less than the cell grow on the treated surface which implied that the anchor site for cell adhesion is not enough. Moreover, adsorption of PLL on the negatively charge surface maybe in monolayer. PLL and other protein molecules are classified into nanoscale regime molecules thus, in this experiment, we also create a nanoscale roughness surface for cell adhesion. However, the effect of this roughness and its properties should have more intensively study in the future.
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Fig. 2. Cell grow on ITO platform showing cell adhere on a, b) bare ITO surface and c,d) on PLL-modified ITO surface after seeding for 1 day and 3 days, repectively.
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Suthiwan Udomrat et.al./ Materials Today: Proceedings 5 (2018) 11083–11088
Fig. 3 Amount of cell grown on bare and PLL modified-ITO platform. Discrepancy amount of cell in both conditions is noticed at day 3 after seeding and consistent with fig. 2. 4. Conclusions We successfully demonstrate that biological soft tissue can be grown and incorporated in the metal oxide basesurface by using simple method fabrication. This experiment reveals the role of indium–tin-oxide surface treatment on cell growth and proliferation. Utilizing of surface modification changes surface properties, we can selective and control of cell growth and alignment on the surface by changing surface charge or coating with poly-L-lysine. PLL not only plays the important role for cell adhesion and growth but its monolayer adsorption may also provide nanosurface roughness, which should be intensively study in the future. References [1] AFM. Johnstone, GW. Gross, DG. Weiss, OHU. Schroeder, A. Gramowski, TJ Shafer. NeuroToxicol. 31 (2010) 331-50. [2] F. Pires, Q. Ferreira, CAV. Rodrigues, J. Morgado, FC. Ferreira, Biochimica Biophysica Acta (BBA) - General Subjects. (2015) 1158-68. [3] S. Ahadian, J. Ramon-Azcon, S. Ostrovidov, G. Camci-Unal, Hosseini V, Kaji H. Lab on a Chip. 12 (2012) 3491-503. [4] C Prakash , HK Kansal, BS Pabla, S. J Puri. Mater. Engineer. Perform. 24 (2015) 3622-33. [5] H. Zhao, L. Li, H-J Fan, F. Wang, L-M Jiang, P-G He. Molec. Cell. Biochem. (2012) 309-13. [6] H. Zhao, A. Steiger, M. Nohner, H. Ye. PLOS ONE. 10 (2015). [7] M. Encinas, M. Iglesias, Y. Liu, Wang H, A. Muhaisen, V. J. Ceña. Neurochem. 75 (2000) 991-1003. [8] L-C Hsiung, C-H Yang, C-L Chiu, C-L Chen, Y. Wang, H. Lee. Biosensors and Bioelectronics. 24 (2008) 869-75. [9] CJ. Huang, YK. Su, SL. Wu. Mater. Chem. Phys. 84 (2004) 146-50. [10] OG. Alamdari, E. Seyedjafari, M. Soleimani, N. Ghaemi. J Avicenna. Medic. Biotechnol. 5 (2013) 234-40. [11] S Joo, Yeon J. Kim, E. Lee, N. Hong, W. Sun, Y. Nam. Scientif. Rep. 5 (2015). [12] M. Abercrombie. In Vitro. 6 (1970) 128-42. [13] A. Swat, I. Dolado, JM. Rojas, AR. Nebreda. Molec. Cellul. Biol. 29 (2009) 3332-43. [14] J. Robertus, WR. Browne, BL. Feringa. Chemic. Soc. Rev. 39 (2010) 354-78. [15] R. Wu, Y. Li, Q. Wang, J. Yu, F. Jiang, F. Wang. RSC Advances. 2 (2012) 9887-93.