T98G Cell Death Induced by Photothermal Treatment with Hollow

Subscriber access provided by MIDWESTERN UNIVERSITY

Biological and Medical Applications of Materials and Interfaces

T98G Cell Death Induced by Photothermal Treatment with Hollow Gold Nanoshell-Coupled Silica Microrods Prepared from Escherichia Coli Soomin Han, Yoo-Jin Park, Eun-Jung Park, and Younghun Kim ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.8b21199 • Publication Date (Web): 14 Feb 2019 Downloaded from http://pubs.acs.org on February 15, 2019

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ACS Applied Materials & Interfaces

T98G Cell Death Induced by Photothermal Treatment with Hollow Gold Nanoshell-Coupled Silica Microrods Prepared from Escherichia Coli Soomin Han,† Yoo-Jin Park,‡ Eun-Jung Park‡ and Younghun Kim†* †Department

of Chemical Engineering, Kwangwoon University, 20 Kwangwoon-ro, Nowon-gu, Seoul 01897, Korea

‡Graduate

School of East-West Medical Science, Kyung Hee University, Yongin 17104, Korea

Keywords: Photothermal treatment; Escherichia coli; hollow gold nanoshell; near infrared irradiation; T98G cell; tumor cell

ACS Paragon Plus Environment

1

ACS Applied Materials & Interfaces 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 2 of 24

ABSTRACT

As an alternative to traditional cancer treatment, photothermal therapy is a promising method with advantages such as non-invasiveness and high efficiency. Herein, we synthesized armored golden Escherichia coli (AGE) microrods as photothermal agents to evaluate the viability of cancer cell. The hollow gold nanoshell (HAuNS) was synthesized for photothermal effects under near infrared (NIR) region using unicellular E. coli as framework. Coupling HAuNS onto the surface of E. coli@SiO2 enhanced temperature elevation and resulted in high conversion efficiency. The synthesized AGE microrods had excellent photothermal stability under NIR laser irradiation in the five times recycling experiment. The temperature elevation of AGE microrod solution reached 43.7 C, which induced hyperthermia-mediated killing of tumor cells. The results of cytotoxicity test revealed the AGE microrod-induced T98G cell death mediated via apoptosis.


2

Page 3 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Introduction

Metallic nanoparticles (NPs) are gaining momentum for applications in the field of medicine such as drug delivery,1-3 image-guided therapy,4,5 lipolysis,6,7 and cancer treatment.8-12 Photothermal therapy, a method of tumor ablation using the heat induced by light, has received tremendous attention in the field of cancer treatment. This treatment method may effectively ablate the targeted tumor cells and reduce the damage caused to normal cells.13-15 For applications in photothermal therapy, NPs must efficiently convert light into heat. Typical organic16 and inorganic NPs17 with near-infrared (NIR) absorption (700-1,000 nm) have been used to penetrate soft tissues.18 Organic NPs exhibit excellent biocompatibility and low price but may be required in large quantities to facilitate effective photothermal therapy. Inorganic NPs, especially metallic NPs, reveal five-time higher cross-section absorption than organic NPs.19 Metal NPs are needed in a very small amount. Among metal NPs, gold (Au) NPs are the most popular owing to several favorable properties. These particles exhibit excellent biocompatibility than other metal NPs20,21 and their absorbance could be readily changed from visible to NIR region via adjustment of their shape and size.22 The general mechanism for heat generation of Au NPs by NIR irradiation is as follows: laser irradiation leads to localized surface plasmon resonance (LSPR) phenomenon of Au NPs, which induces a strong electromagnetic field. This phenomenon causes enhancement in visible light absorption and excited electrons, which are rapidly equilibrated via electron-electron scattering to create “hot electron” distribution. Induced hot electrons lose their energies by interaction with phonons of the metal lattice and elevate the lattice temperature. Cooling occurs in the final step via heat transfer to the solvent and heat diffusion.23,24 Here, we prepared armored golden Escherichia coli (AGE) microrods and examined their photothermal properties in T98G cells. The hollow gold nanoshell (HAuNS) with absorbance in the NIR region was synthesized and coupled onto the surface of E. coli@SiO2, which was used as a framework. The photothermal effect and biocompatibility were enhanced after coupling of HAuNS to the surface of


3


Page 4 of 24

E. [email protected] Temperature elevation of AGE microrod solution reached 43.7C, which induced hyperthermia in tumor cells. AGE microrod showed excellent stability in various solvents and induced T98G cell death via apoptosis. Therefore, AGE microrods may serve as efficient agents for photothermal therapy.


4

Page 5 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Results and Discussion Preparation and characterization of AGE microrods The synthesis scheme of AGE microrods is shown in Figure 1. To prepare AGE microrods, we first synthesized HAuNS via galvanic replacement reaction between AgNPs and HAuCl4. This reaction facilitated the formation of hollow structures of HAuNS through dissolution of AgCl, resulting in change in the absorbance peak from visible to NIR region.26 TEM images revealed the hollow structure and rough surface of HAuNS (Figure 2a). The surface roughness of HAuNS may be helpful to enhance the SPR band by tip plasmon mode.27 The particle size of HAuNS measured by TEM is ca. 115.9 ± 22.7 nm, and the sizes in the axial/longitudinal direction for AGE particles was 4156.4/951.5 nm.

Figure 1. Scheme of AGE microrod preparation using E. coli template and photothermal treatment of tumor cells using NIR laser irradiation.


5


Page 6 of 24

Figure 2. TEM images. (a) HAuNS, (b) E. coli@SiO2, and AGE microrod (c) before irradiation and (d) after irradiation with five cycles of on/off NIR laser treatment.

For the enhancement of photothermal effects, E. coli@SiO2 were synthesized. E. coli is a rod shaped bacterium that is easily cultivated, and was used as a framework for photothermal agents. Silica coating on the surface of E. coli induced cell death and maintained the solid and free-standing vehicle. E. coli@SiO2 were functionalized with APTES to allow HAuNS coupling. Polyvinylpyrrolidone (PVP) was simultaneously coated onto the surface of HAuNS to enhance stability, and the PVP-coated HAuNS were coupled onto APTES-modified E. coli@SiO2 surfaces via electrostatic attraction. The coupling reaction induced a strong electromagnetic field between adjacent HAuNS28 and changed the absorbance peak of AGE microrods with a red shift, thereby enhancing photothermal effects.29 As shown in Figure 3, coupling of HAuNS onto E. coli@SiO2 resulted in a slight shift in LSPR peak from 819 to 839 nm. In TEM images, HAuNS was found uniformly decorated on the surface of silica microrods (Figure 2c).


6

Page 7 of 24

1.0 0.9

E. coli@SiO2 HAuNS

0.8

AGE microrod

0.7

Absorbance

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


0.6 0.5 0.4 0.3 0.2 0.1 0.0 300

400

500

600

700

800

900

1000

1100

Wavelength (nm) Figure 3. UV-vis absorption spectra of E. coli@SiO2, HAuNS, and AGE microrods.

Photothermal performance AGE microrods had stronger absorbance in NIR region than HAuNS, indicative of the enhancement in photothermal effects. To evaluate photothermal effects, 808 nm NIR laser at a power density of 2 W/cm2 was irradiated for 15 min on the different concentrations of aqueous solution containing AGE microrods (25, 50, 75, and 100 μg/mL). Both pure water and HAuNS (concentration: 50 μg/mL) solution were evaluated under same irradiation conditions for comparison. As the temperature of aqueous solutions reached a steady state (15 min), the laser was turned-off to allow cooling of the solution to room temperature (15 min). Different concentrations (25, 50, 75, and 100 μg/ml) of AGE microrod solutions showed temperature elevations of 39.4C, 43.7C, 46.8C, and 50.9C, respectively (Figure 4a). However, temperature of pure water and HAuNS solutions at 50-μg/mL concentration reached 34.1C and 39.2C, respectively (Figure 4b). E. coli framework was positively influenced by the enhancement in temperature elevation via coupling effect. In addition, the solution containing AGE microrods reached temperatures


7


Page 8 of 24

that induced hyperthermia in tumor cells at concentrations between 50 and 75 μg/mL. As cell death induced by hyperthermia is mediated at temperatures between 40C and 47C, the elevation of temperature above 50C is thought to cause DNA damage and denaturation.30,31

Figure 4. Temperature changes. (a) AGE microrod solutions at different concentrations (25, 50, 75, and 100 μg/mL) and (b) pure water, HAuNS, and AGE microrod solutions at a concentration of 50 μg/mL under NIR irradiation for 15 min (808 nm, 2 W/cm2).


8

Page 9 of 24

The NPs were dispersed in phosphate-buffered saline (PBS) and Dulbecco’s modified Eagle’s medium (DMEM) at 50-μg/mL concentration and the temperature elevation in AGE microrods was evaluated. The temperature change was monitored under NIR laser irradiation for 15 min (2 W/cm2). As shown in Figure 5, the temperature of AGE microrods in PBS and DMEM increased to 43.1C and 42.4C, respectively. The temperature elevation profile in different solutions was similar to that in DI water.

46 PBS DMEM water

44 42

o

Temperature ( C)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


40 38 36 34 32 30 0

1

2

3

4

5

6

7

8

9

10 11 12 13 14 15 16

Time (min)

Figure 5. Temperature changes in AGE microrods in different types of solvents (pure water, PBS, and DMEM) at a concentration of 50 μg/mL under NIR irradiation for 15 min (808 nm, 2 W/cm2).

Calculation of photothermal conversion efficiency of AGE microrods The photothermal conversion efficiency of AGE microrods was determined by recording the temperature change in AGE microrod solution as a function of time under 808 nm laser irradiation (2 W/cm2) until the temperature reached a steady state (15 min). Photothermal conversion efficiency (η) was calculated with the following equations.32,33 The total energy balance equation of the system was derived


9


Page 10 of 24

from Equation (1):

∑𝑚 𝐶

𝑑𝑇 = 𝑄𝐴𝐺𝐸 + 𝑄𝑑𝑖𝑠 ― 𝑄𝑠𝑢𝑟𝑟…(1) 𝑑𝑡

𝑖 𝑝,𝑖

𝑖

Where, 𝑚𝑖 and 𝐶𝑝,𝑖 are mass and heat capacity of AGE microrods, quartz cuvette, and water in the system, respectively; 𝑇 and t are the system temperature and time, respectively; 𝑄𝐴𝐺𝐸 is the energy changed by AGE microrods from the radiant energy; 𝑄𝑑𝑖𝑠 is the heat dissipation from the light absorbed by the quartz cuvette; and 𝑄𝑠𝑢𝑟𝑟 presents the energy loss from the surface by air. 𝑄𝐴𝐺𝐸 is induced by electron-phonon relaxation of AGE microrod surfaces under 808 nm laser irradiation. 𝑄𝐴𝐺𝐸 = I(1 ― 10 ―𝐴808)η…(2) Where, η is the photothermal conversion efficiency, I is the 808 nm laser power (2 W/cm2), and A808 is the absorbance of the AGE microrod solutions at 808 nm. The heat loss term 𝑄𝑠𝑢𝑟𝑟 is represented by following equation (3): 𝑄𝑠𝑢𝑟𝑟 = ℎ𝐴(𝑇 ― 𝑇𝑠𝑢𝑟𝑟)…(3) Where, A is surface area of the container, h is the heat transfer coefficient, and 𝑇 and 𝑇𝑠𝑢𝑟𝑟 are the system temperature and ambient temperature of the surrounding, respectively. Laser-induced energy 𝑄𝑑𝑖𝑠 was 0.00108 W, as measured using a quartz cuvette cell containing pure water. Steady state is the time when the system temperature reached the maximum value. During steady state, input energy is equal to output energy. Thus, Equation (1) is described as below: 𝑑𝑇 = 𝑄𝐴𝐺𝐸 + 𝑄𝑑𝑖𝑠 ― 𝑄𝑠𝑢𝑟𝑟 = 0…(4) 𝑑𝑡

∑𝑚 𝐶

𝑖 𝑝,𝑖

𝑖

Combining the equation (2), (3), and (4), photothermal conversion efficiency may be determined using Equation (5): η=

ℎ𝐴(𝑇𝑚𝑎𝑥 ― 𝑇𝑠𝑢𝑟𝑟) ― 𝑄𝑑𝑖𝑠 𝐼(1 ― 10 ―𝐴808)

…(5)

To calculate photothermal conversion efficiency, ℎ𝐴 value needs to be calculated. To determine ℎ𝐴, dimensionless driving force temperature θ and time constant 𝜏𝑠 are defined by Equation (6) and (7):


10

Page 11 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


θ≡

𝑇 ― 𝑇𝑠𝑢𝑟𝑟

…(6) 𝑇𝑚𝑎𝑥 ― 𝑇𝑠𝑢𝑟𝑟

∑ 𝑚𝑖𝐶𝑝,𝑖 𝑖 𝜏𝑠 = …(7) ℎ𝐴 When the laser is turned off, the value of 𝑄𝐴𝐺𝐸 + 𝑄𝑑𝑖𝑠 is 0. Thus, 𝜏𝑠 may be calculated using Equation (8): t ≡ ― 𝜏𝑠ln 𝜃…(8) As shown in Figure 6a, 𝜏𝑠 was determined to be 242.22 s and ℎ𝐴 value was 0.005478 W/K. In addition, total 𝑚𝐶𝑝 was 13.27 J/K, and the absorbance at 808 nm and (𝑇𝑚𝑎𝑥 ― 𝑇𝑠𝑢𝑟𝑟) were 0.798 and 13.7 K, respectively. The photothermal conversion efficiency was calculated as 44.57% at 50-μg/mL AGE microrods. However, photothermal conversion efficiency of 75-μg/mL AGE microrods and 50-μg/mL HAuNS were calculated to be 39.56% and 28.19%, respectively. The temperature of AGE microrods was higher at 75 μg/mL than at 50 μg/mL. It means high cytotoxicity and low photothermal conversion efficiency. Therefore, 50 μg/mL concentration was used as the efficient dose of AGE microrods to kill tumor cells. The stability of AGE microrods was evaluated in the NIR laser on/off experiment. The laser was turned-on for 15 min and then turned-off for 15 min. This process was repeated for five cycles. During five cycles of laser on/off, no significant decrease in temperature was observed (Figure 6b). After the recycling test, AGE microrods maintained their shape without significant separation of HAuNS from the surface of E. coli@SiO2 (Figure 2d). High photothermal conversion efficiency and excellent photothermal stability indicate that AGE microrods are excellent photothermal agents for heat-mediated tumor treatment.


11


Page 12 of 24

Figure 6. (a) Plot of linear fitting time data versus negative natural logarithm of driving force temperature obtained from the cooling stage and (b) temperature changes of AGE microrod solutions at a concentration of 50 μg/mL during five cycles of on/off NIR laser irradiation (808 nm, 2 W/cm2).


12

Page 13 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Cytotoxicity and photothermal toxicity in vitro NPs with good biocompatibility and photothermal effects have great potential as photothermal agents. To evaluate biocompatibility, cytotoxicity of HAuNS and AGE microrods was tested in T98G glioma cells. The cultured cells were divided into seven groups (control, HAuNS-treated, and AGE microrod-treated groups at different concentrations [12.5, 25, and 50 μg/mL]). T98G cells were exposed to different concentrations of HAuNS and AGE microrods for 24 h. The viability of cells was similar after treatment with HAuNS and AGE microrods at 12.5 and 25 μg/mL concentrations. However, at HAuNS concentration of 50 μg/mL, the cell viability drastically decreased to 52%, while the viability of cells was 81.9% following treatment with AGE microrods (Figure 7a). E. coli framework helped enhance the biocompatibility of these microrods. Photothermal toxicity was measured under NIR laser irradiation for 10 min (808 nm, 2 W/cm2). After irradiation, the viability of cells treated with HAuNS and AGE microrods decreased to 5.23% and 6.01%, respectively. This effect was associated with the photothermal heating. No significant change in T98G cell viability was observed following irradiation with NIR laser. The use of E. coli@SiO2 as a framework facilitated the change in the shape of the particles, which became elongated and had excellent chance to interact with the cell surface owing to the increased surface area as compared with spherical NPs. Thus, AGE microrods induced more drastic cell death effects. There are two different cell death mechanisms: necrosis and apoptosis. Apoptosis is “clean” cell death pathway and is known to reduce external damages. However, necrosis destroys cell functions and causes inflammation, owing to the leakage of the cytoplasmic contents of the cells to the extracellular space. Thus, apoptosis is preferred over necrosis during photothermal treatment. As shown in Figure 8, the feature of T98G cell death was similar to that observed during apoptosis. Therefore, AGE microrods have excellent properties as a potential photothermal agent.


13


Page 14 of 24

Figure 7. Viability of T98G cells treated for 24 h with HAuNS or AGE microrod solutions. (a) Viability in the presence of different concentrations (0, 12.5, 25, and 50 μg/mL). (b) Viability after treatment with 50 μg/mL of agents with or without irradiation for 10 min (808 nm, 2 W/cm2).


14

Page 15 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 8. Optical images of T98G cells. (a, b) PBS, (c, d) HAuNS, and (e, f) AGE microrod. (a, c, e) Before and (b, d, f) after NIR laser irradiation for 10 min (808 nm, 2 W/cm2).


15


Page 16 of 24

3. Conclusions AGE microrods were synthesized to facilitate killing of T98G glioma cell mediated by the heat induced by photothermal effects. HAuNS with absorbance in NIR region was synthesized by galvanic replacement reaction. E. coli cells were used as the framework to enhance the photothermal effects via coupling reaction and were covered with SiO2 to improve stability. E. coli@SiO2 were functionalized with amino groups and uniformly coupled with HAuNS via electrostatic interaction. The synthesized AGE microrods had strong absorbance in NIR region. Irradiation of HAuNS and AGE microrod solutions with NIR laser resulted in temperature elevation, which was higher for AGE microrods than HAuNS; thus, AGE microrods showed better photothermal conversion efficiency. Moreover, AGE microrods demonstrated excellent stability under NIR laser irradiation in five times recycling experiment. The results of cytotoxicity assay revealed the better biocompatibility of AGE microrods than HAuNS and photothermal effects that induce T98G cell death via apoptosis. The synthesized AGE microrod may serve as a potential photothermal agent for T98G glioma cell death.


16

Page 17 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


4. Experimental Section Synthesis of HAuNS HAuNS was synthesized using Ag NPs and HAuCl4 in a galvanic replacement reaction. Ag NPs were synthesized with 0.025 M of silver nitrate, and 1 wt% of trisodium citrate (TSC) was added to 80 mL of boiled deionized (DI) water. After 20 min, the color of the solution changed from yellow to green; the solution was cooled down at room temperature. A total of 5 mL of Ag NPs was dispersed in 3 mM HAuCl4, followed by injection of 5 mL of 10 mM ascorbic acid. For enhancement in the dispersion stability, 0.1 g/mL of PVP was added into the solution and stirred at 50C for 6 h.

Synthesis of E. coli@SiO2 E. coli@SiO2 was used as a framework for microrods. E. coli was routinely cultured at 37C for 48 h in Luria-Bertani (LB) medium containing yeast extract (0.5 g), tryptone (1 g), and sodium chloride (1 g) in 100 mL of DI water. Cell culture medium was removed by centrifugation and cells were washed thrice with DI water. The obtained E. coli cells were re-dispersed in 15 mL DI water and 10 mL of the redispersed E. coli solution was added to fresh LB medium. Extra solution was dispersed in 80% ethanol solution for silica coating. E. coli@SiO2 were synthesized by a modified Stober method. The re-dispersed E. coli solution was added to a mixture of 80 v/v% ethanol and 20 v/v% DI water, and 1 mL of ammonium hydroxide (NH4OH) was added to the mixture, followed by dropwise addition of tetraethyl orthosilicate (TEOS) solution. The mixture was stirred for 6 h and washed thrice with ethanol. The washed E. coli@SiO2 particles were re-dispersed in ethanol and dried.

Preparation of AGE microrods To couple HAuNS on E. coli@SiO2, (3-aminopropyl) triethoxysilane (APTES) was slowly added to 0.01 g of E. coli@SiO2 dispersed in 40 mL ethanol. The mixture was stirred at 50C for 6 h to ensure complete functionalization and for the removal of extra solution using centrifugation to collect E.


17


Page 18 of 24

coli@SiO2. The collected E. coli@SiO2 were washed thrice with ethanol and re-dispersed in DI water. The APTES-functionalized E. coli@SiO2 solution was added to PVP-coated HAuNS solution. After stirring for 12 h, the mixture was centrifuged and washed thrice with DI water to obtain AGE microrods.

Characterization and photothermal performance The morphology of the synthesized AGE microrods was investigated with transmission electron microscopy (TEM, JEOL, JEM-2010). The hydrodynamic diameter and zeta potential were determined by dynamic light scattering (DLS, Photal, ELS-Z) and their absorption spectra from UV-visible to NIR were collected (Shimadzu, UV-1800). To investigate photothermal performance, different concentrations of AGE microrods (0, 25, 50, 75, and 100 μg/mL) were dispersed in a quartz cuvette and irradiated for 15 min using NIR laser (808 nm, 2 W/cm2). The temperature of each solution was measured every 1 min by NIR imaging camera (FLIR System, FLIR ONE).

Cytotoxicity and photothermal toxicity in vitro Human brain glioblastoma cells (T98G) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum and 100 units/mL of penicillin/streptomycin. The cytotoxicity of HAuNS and AGE microrods against T98G cells was evaluated with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, a colorimetric assay used for assessing cell metabolic activity. T98G cells (2  104 cells/well) were overnight cultured in four-well plates and treated with culture medium containing different concentrations of HAuNS and AGE microrods (0, 12.5, 25, and 50 μg/mL). T98G cells were eventually exposed to NIR laser (808 nm, 2 W/cm2) for 10 min and incubated for 24 h at 37C. After removing the supernatant, MTT solution (20 μg/mL, 200 μL/well) was added to each well, and the cells were incubated for another 3 h at 37C. The formazan crystals formed were dissolved in dimethyl sulfoxide and the absorbance at 540 nm


18

Page 19 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


in each well was measured using a microplate spectrophotometer system (VictorTM × 3, PerkinElmer, Waltham, MA, USA). The treated T98G cells were observed under an optical microscope.

AUTHOR INFORMATION Corresponding authors E-mail: [email protected] (Y.K.) ORCID Younghun Kim: 0000-0002-2570-3808 Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes The authors declare no competing financial interest.

Acknowledgements This work was supported by the National Research Foundation of Korea (NRF-2017R1A2B4001829).


19


Page 20 of 24

REFERENCES (1) Alea-Reyes, M. E.; González, A., Calpena, A. C.; Ramos-López, D.; de Lapuente, J.; Pérez-García, L. Gemini Pyridinium Amphiphiles for the Synthesis and Stabilization of Gold Nanoparticles for Drug Delivery. J. Colloid Interface Sci. 2017, 502, 172–183. (2) Oh, J. Y.; Kim, H. S.; Palanikumar, L.; Go, E. M.; Jana, B.; Park, S. A.; Kim, H. Y.; Kim, K.; Seo, J. K.; Kwak, S. K.; Kim, C.; Kang, S.; Ryu, J. H. Cloaking Nanoparticles with Protein Corona Shield for Targeted Drug Delivery. Nat. Commun. 2018, 9, 4548. (3) Than, A.; Liu, C.; Chang, H.; Duong, P. K.; Cheung, C. M. G.; Xu, C.; Wang, X.; Chen, P. SelfImplantable Double-Layered Micro-Drug-Reservoirs for Efficient and Controlled Ocular Drug Delivery. Nat. Commun. 2018, 9, 4433. (4) Liu, L.; Cui, H.; Yan, S.; Jing, X.; Wang, D.; Meng, L. Gold Nanostars Decorated MnO2Nnanosheets for Magnetic Resonance Imaging and Photothermal Erasion of Lung Cancer Cell. Mater. Today Commun. 2018, 16, 97–104. (5) Ke, H.; Yue, X.; Wang, J.; Xing, S.; Zhang, Q.; Dai, Z.; Tian, J; Wang, S.; Jin, Y. Gold Nanoshelled Liquid Perfluorocarbon Nanocapsules for Combined Dual Modal Ultrasound/CT Imaging and Photothermal Therapy of Cancer. Small. 2014, 10, 1220–1227. (6) Lee, J. H.; Jeong, H. S.; Lee, D. H.; Beack, S.; Kim, T.; Lee, G. H.; Park, W. C.; Kim, C.; Kim, K. S.; Hahn, S. K. Targeted Hyaluronate-Hollow Gold Nanosphere Conjugate for Anti-Obesity Photothermal Lipolysis. ACS Biomater. Sci. Eng. 2017, 3, 3646–3653. (7) Sheng, W.; Alhasan, A. H.; DiBernardo, G.; Almutairi, K. M.; Rubin, J. P.; DiBernardo, B. E.; Almutairi, A. Gold Nanoparticle-Assisted Selective Photothermolysis of Adipose Tissue (NanoLipo). Plast. Reconstr. Surg. Glob. Open. 2014, 2, 1–8. (8) Liu, Y.; Xu, J.; Choi, H. H.; Han, C.; Fang, Y.; Li, Y.; Van der Jeught, K.; Xu, H.; Zhang, L.; Frieden, M.; Wang, L.; Eyvani, H.; Sun, Y.; Zhao, G.; Zhang, Y.; Liu, S.; Wan, J.; Huang, C.; Ji, G.; Lu, X.; He, X.; Zhang, X. Targeting 17q23 Amplicon to Overcome the Resistance to Anti-HER2 Therapy in HER2+ Breast Cancer. Nat. Commun. 2018, 9, 4718.


20

Page 21 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(9) Cao, y.; Li, S.; Chen, C.; Wang, D.; Wu, T.; Dong, H.; Zhang, X. Rattle-Type Au@Cu2−xS Hollow Mesoporous Nanocrystals with Enhanced Photothermal Efficiency for Intracellular Oncogenic MicroRNA Detection and Chemo-Photothermal Terapy. Biomaterials. 2018, 158, 23–33. (10) Chen, J.; Glaus, C.; Laforest, R.; Zhang, Q.; Yang, M.; Gidding, M.; Welch, M. J.; Xia, Y. Gold Nanocages as Photothermal Transducers for Cancer Treatment. Small. 2010, 6, 811–817. (11) Kang, S.; Lee, J.; Ryu, S.; Kwon, Y.; Kim, K. H.; Jeong, D. H.; Paik, S. R.; Kim, B. S. Gold Nanoparticle/Graphene Oxide Hybrid Sheets Attached on Mesenchymal Stem Cells for Effective Photothermal Cancer Therapy. Chem. Mater. 2017, 29, 3461–3476. (12) Lerner, M. I.; Mikhaylov, G.; Tsukanov, A. A.; Lozhkomoev, A. S.; Gutmanas, E.; Gotman, I.; Bratovs, A.; Turk, V.; Turk, B.; Psakhye, S. G.; Vasiljeva, O. Crumpled Aluminum Hydroxide Nanostructures as a Microenvironment Dysregulation Agent for Cancer Treatment. Nano Lett. 2018, 18, 5401–5410. (13) Nakamura, T.; Tamura, A.; Murotani, H.; Oishi, M.; Jinji, Y.; Matsuishi, K.; Nagasaki, Y. Large Payloads of Gold Nanoparticles into the Polyamine Network Core of Stimuli-Responsive PEGylated Nanogels for Selective and Noninvasive Cancer Photothermal Therapy. Nanoscale. 2010, 2, 739–746. (14) Zou, L.; Wang, H.; He, B.; Zeng, L.; Tan, T.; Cao, H.; He, X.; Zhang, Z.; Guo, S.; Li, Y. Current Approaches of Photothermal Therapy in Treating Cancer Metastasis with Nanotherapeutics. Theranostics. 2016, 6, 762–772. (15) Jo, H. H.; Youn, H. G.; Lee, S. H., Ban, C. G. Ultra-Effective Photothermal Therapy for Prostate Cancer Cells using Dual Aptamer-Modified Gold Nanostars. J. Mater. Chem. B. 2014, 2, 4862–4867. (16) Zhou, B.; Li, Y.; Niu, G.; Lan, M.; Jia, Q.; Liang, Q. Near-Infrared Organic Dye-Based Nanoagent for the Photothermal Therapy of Cancer. ACS Appl. Mater. Interfaces. 2016, 8, 29899–29905. (17) Tian, Q.; Hu, J.; Zhu, Y.; Zou, R.; Chen, Z.; Yang, S.; Li, R.; Su, Q.; Han, Y.; Liu, X. Sub-10 nm


21


Page 22 of 24

Fe3O4@Cu2-xS Core-Shell Nanoparticles for Dual-Modal Imaging and Photothermal Therapy. J. Am. Chem. Soc. 2013, 135, 8571–8577. (18) Yun, S. H.; Kwok, S. J. J. Light in Diagnosis, Therapy and Surgery. Nat. Biomed. Eng. 2017, 1, 0008. (19) Jain, P. K.; Lee, K. S.; El-Sayed, I. H.; El-Sayed, M. A. Calculated Absorption and Scattering Properties of Gold Nanoparticles of Different Size, Shape, and Composition: Applications in Biological Imaging and Biomedicine. J. Phys. Chem. B. 2006, 110, 7238–7248. (20) Connor, E. E.; Mwamuka, J.; Gole, A.; Murphy, C. J.; Wyatt, M. D. Gold Nanoparticles Are Taken Up by Human Cells but Do Not Cause Acute Cytotoxicity. Small. 2005, 1, 325–327. (21) Li, X.; Hu, Z.; Ma, J.; Wang, X.; Zhang, Y.; Wang, W.; Yuan, Z. The Systematic Evaluation of SizeDependent Toxicity and Multi-Time Biodistribution of Gold Nanoparticles. Colloids Surfaces B Biointerfaces. 2018, 167, 260–266. (22) Qiu, J.; Wei, W. D. Surface Plasmon-Mediated Photothermal Chemistry. J. Phys. Chem. C. 2014, 118, 20735–20749. (23) Webb, J. A.; Bardhan, R. Emerging Advances in Nanomedicine with Engineered Gold Nanostructures. Nanoscale. 2014, 6, 2502–2530. (24) Qin Z.; Bischof, J. C. Thermophysical and Biological Responses of Gold Nanoparticle Laser Heating. Chem. Soc. Rev. 2012, 41, 1191–1217. (25) Lee, H.; Han, S.; Kim, Y. Synthesis of Gold-Spikes Decorated Biomimetic Silica Microrod for Photothermal Agents. J. Ind. Eng. Chem. 2018, 58, 33–37. (26) Ma, N. Jiang, Y. W., Zhang, X.; Wu, H.; Myers, J. N.; Liu, P.; Jin, H.; Gu, N.; He, N.; Wu, F. G.; Chen, Z. Enhanced Radiosensitization of Gold Nanospikes via Hyperthermia in Combined Cancer


22

Page 23 of 24 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Radiation and Photothermal Therapy. ACS Appl. Mater. Interfaces. 2016, 8, 28480–28494. (27) Barbosa, S.; Agrawal, A.; Rodríguez-Lorenzo, L.; Pastoriza-Santos, I.; Alvarez-Puebla, R. A.; Kornowski, A.; Weller, H.; Liz-Marzán, L. M. Tuning Size and Sensing Properties in Colloidal Gold Nanostars. Langmuir. 2010, 26, 14943–14950. (28) Serrano-Montes, A. B.; Langer, J.; Henriksen-Lacey, M.; Jimenez De Aberasturi, D.; Solís, D. M.; Taboada, J. M.; Obelleiro, F.; Sentosun, K.; Bals, S.; Bekdemir, A.; Stellacci, F.; Liz-Marzán, L. M. Gold Nanostar-Coated Polystyrene Beads as Multifunctional Nanoprobes for SERS Bioimaging. J. Phys. Chem. C. 2016, 120, 20860–20868. (29) Jain, P. K.; Huang, W.; El-Sayed, M. A. On the Universal Scaling Behavior of the Distance Decay of Plasmon Coupling in Metal Nanoparticle Pairs: A Plasmon Ruler Equation. Nano Lett. 2007, 7, 2080– 2088. (30) Harmon, B. V.; Corder, A. M.; Collins, R. J.; Gobé, G. C.; Allen, J.; Allan, D. J.; Kerr, J. F. R. Cell Death Induced in a Murine Mastocytoma by 42-47°C Heating in Vitro: Evidence that the Form of Death Changes from Apoptosis to Necrosis Above a Critical Heat Load. Int. J. Radiat. Biol. 1990, 58, 845–858. (31) Roti, J. L. Cellular Responses to Hyperthermia (40-46°C): Cell Killing and Molecular Events. Int. J. Hyperth. 2008, 24, 3–15. (32) Roper, D. K.; Ahn, W.; Hoepfner, M. Microscale Heat Transfer Transduced by Surface Plasmon Resonant Gold Nanoparticles. J. Phys. Chem. C 2007, 111, 3636–3641. (33) Li, K. C.; Chu, H. C.; Lin, Y.; Tuan, H. Y.; Hu, Y. C. PEGylated Copper Nanowires as a Novel Photothermal Therapy Agent. ACS Appl. Mater. Interfaces 2016, 8, 12082–12090.


23


Page 24 of 24

TOC


24

T98G Cell Death Induced by Photothermal Treatment with Hollow

Recommend Documents