Article
pubs.acs.org/Langmuir
ATP-Responsive Controlled Release System Using AptamerFunctionalized Mesoporous Silica Nanoparticles
Xiaoxiao He,† Yingxiang Zhao,† Dinggeng He, Kemin Wang,* Fengzhou Xu, and Jinlu Tang
State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, College of Chemistry and Chemical Engineering,
Hunan University, Key Laboratory for Bio-Nanotechnology and Molecular Engineering of Hunan Province, Changsha 410082,
People’s Republic of China
S Supporting Information
*
ABSTRACT: Adenosine-5′-triphosphate (ATP) is a multifunctional
nucleotide, which plays a vital role in many biological processes, including
muscle contraction, cells functioning, synthesis and degradation of
important cellular compounds, and membrane transport. Thus, the
development of ATP-responsive controlled release system for bioorganism
application is very significative. Here, an original and facile ATP-responsive
controlled release system consisting of mesoporous silica nanoparticles
(MSN) functionalized with an aptamer as cap has been designed. In this system, the ATP aptamer was first hybridized with arm
single-stranded DNA1 (arm ssDNA1) and arm single-stranded DNA2 (arm ssDNA2) to form the sandwich-type DNA structure
and then grafted onto the MSN surface through click chemistry approach, resulting in blockage of pores and inhibition of guest
molecules release. In the presence of ATP, the ATP aptamer combined with ATP and got away from the pore, leaving the arm
ssDNA1 and ssDNA2 on the surface of MSN. The guest molecules can be released because single-stranded DNA is flexible. The
release of the guest molecules from this system then can be triggered by the addition of ATP. As a proof-of-principle, Ru(bipy)32+
was selected as the guest molecules, and the ATP-responsive loading and release of Ru(bipy)32+ have been investigated. The
results demonstrate that the system had excellent loading efficiency (215.0 μmol g−1 SiO2) and the dye release percentage can
reach 83.2% after treatment with 20 mM ATP for 7 h. Moreover, the ATP-responsive behavior shows high selectivity with ATP
analogues. However, the leakage of Ru(bipy)32+ molecule is neglectable if ATP was not added, indicating an excellent capping
efficiency. Interestingly, this system can respond not only to the commercial ATP but also to the ATP extracted from living cells.
By the way, this system is also relatively stable in mouse serum solution at 37 °C. This proof of concept might promote the
application of ATP-responsive devices and can also provide an idea to design various target-responsive systems using other
aptamers as cap.
■
INTRODUCTION
Mesoporous silica nanoparticles (MSN) have been used as a
promising carrier for the design of an “on-command” delivery
system because of their distinctive characteristics, such as large
load capacity, biocompatibility, high thermal stability, homogeneous porosity, inertness, tunable pore sizes (2−10 nm), and
easy functionalization of the external and internal surfaces.1 To
date, many MSN-based controlled-release systems have been
constructed by using polymers,2 nanoparticles,3 small organic
molecules,4 supramolecular assemblies,5 and biomolecules6 as
capping agents. Triggered by the physical or chemical stimuli
such as pH,7 redox,8 temperature,9 competitive binding,10
antigen,11 photoirradiation,12 and enzymes,13 the zero release of
guest molecules from the MSN system can be achieved. Despite
these burgeoning achievements, the development of MSN
controlled-release systems responding to biogenic stimuli such
as intracellular pH and ions, small biomolecules, and some
metabolic products is very popular for practical application in
biomedical fields.
With this in mind, some pH-responsive MSN controlledrelease systems based on the lower pH of endosomes and
lysosomes in cells have been reported. For example, Shi et al.14
© 2012 American Chemical Society
constructed a kind of nano-MDDS drugs@micelles@MSNs,
which responded to pH quite well in both vitro condition and
vivo condition. Lee et al.15 selected inorganic calcium
phosphate (CaP) as pore blocker by enzyme-mediated
mineralization on the Si-MP surfaces, which was capable of
releasing guest drugs from the CaP-blocked pore under pH
control. Besides the endosomes and lysosomes pH, the
biogenic biomolecules in cells such as glucose,16 collagen,17
thiol-containing molecules,18 and so on can also be used as
stimuli to design MSN controlled-release systems. As one of the
important biogenic biomolecules, ATP is a multifunctional
nucleotide that is the ubiquitous energy currency for all living
organisms through breaking the phosphoanhydride bond. It is
used for many biological processes and can be used as an
indicator for living microbe existence in clinical microbiology,
food quality control, and environmental analyses.19 Therefore,
exploring gate molecules that can specifically recognize ATP
Received: July 9, 2012
Revised: August 11, 2012
Published: August 14, 2012
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transform infrared (FTIR) spectra were obtained from a TENSOR 27
spectrometer, Bruker Instruments Inc., Germany. UV−vis spectra were
collected by DU-800. N2 adsorption−desorption isotherms were
obtained at −196 °C on a Micromeritics ASAP 2010 sorptometer by
static adsorption procedures. Samples were degassed at 100 °C and
10−3 Torr for a minimum of 12 h prior to analysis. Brunauer−
Emmett−Teller (BET) surface areas were calculated from the linear
part of the BET plot according to IUPAC recommendations. Pore size
distribution was estimated from the adsorption branch of the isotherm
by the Barrett−Joyner−Halenda (BJH) method. Small-angle powder
X-ray diffraction patterns (XRD) of the MSN materials were obtained
in a Scintag XDS-2000 powder diffractometer, using Cu Kα irradiation
(λ = 0.154 nm). All fluorescence spectra were recorded on a Hitachi F7000 spectrophotometer in Tris-HCl buffer and physiological saline
solution.
Preparation of Mesoporous Silica Nanoparticle (MSN). 0.50 g
of CTAB was dissolved in 240 mL of pure water. Next, sodium
hydroxide (1.75 mL, 2 M) was added, followed by adjusting the
solution temperature to 80 °C. TEOS (2.50 mL) was added dropwise
into the mixture solution under vigorous stirring. The mixture was
continually stirred for 2 h to give rise to white precipitates. The solid
product was separated by centrifugation, and washed with deionized
water and ethanol several times. Subsequently, the purified nanoparticles were dried in a high vacuum container at 60 °C overnight.
Preparation of Chlorine-Modified MSN (MSN-Cl). 0.70 g of
prepared MSN was refluxed for 20 h in 60 mL of anhydrous toluene
with 0.70 mL of 3-chloropropyltrimethoxysilane (ClTMS) to yield the
chlorine-modified MSN (MSN-Cl). To remove the surfactant
template (CTAB), 0.50 g of MSN-Cl was stirred at 79 °C in a
mixture containing 0.50 mL of HCl (37.2%) and 50 mL of ethanol.
The resulting material was centrifuged and extensively washed with
nanopure water and ethanol. The surfactant-free particles were then
placed under high vacuum with heating at 60 °C to remove the
remaining solvent from the mesopores.
Preparation of MSN-N3. 100 mg of the obtained MSN-Cl sample
was dispersed in 20 mL of anhydrous DMF, followed by transferring it
into a 100 mL Schlenk flask equipped with a Soxhlet extractor under
nitrogen atmosphere, which was filled with a dried molecular sieve
with a 4 Å pore size. The drying process was carried out for 3 h at 90
°C, and the resulting suspension was saturated with 100 mg of NaN3.
After being stirred for 5 h at 80 °C, the obtained particles were washed
three times with 50 mL of water to yield sample MSN-N3.
Ru(bipy)32+ Loading and Capping. 1.5 mL of 100 μM of arm
ssDNA1, arm ssDNA2, and ATP-aptamer was added into the
centrifuge tube, with heating at 95 °C for 5 min. The DNA mixture
solution was then cooled slowly for hybridization to obtain the ATP
aptamer-containing sandwich-type DNA structure. Separately, 10 mg
of MSN-N3 was soaked in a solution of Ru(bipy)32+ (2 mM) in TrisHCl buffer for 24 h, followed by the addition of 150 nmol of
prehybridized ATP aptamer-containing sandwich-type DNA. Thereafter, 1 μL of CuBr solution (0.1 M in DMSO/tBuOH 3:1) and 2 μL
of the tris-(benzyltriazolylmethylamine) ligand solution (0.1 M in
DMSO/tBuOH 3:1) were added into the mixture and stirred at 4 °C
for 48 h to form the ATP aptamer-containing sandwich-type DNA
capped MSN. The obtained nanoparticles were then centrifuged and
washed thoroughly with Tris-HCl buffer to remove the adsorb guest
molecules and then dried in a vacuum freeze drier to yield the ATP
aptamer-containing sandwich-type DNA-capped MSN (aptamer−
MSN) with encapsulation of Ru(bipy)32+. The control DNAcontaining sandwich-type DNA-capped MSN (denoted as con
DNA−MSN) was obtained by the same procedures above using
control DNA instead of ATP aptamer. All of the washing solutions
were collected, and the loading amount of Ru(bipy)32+ was calculated
from the different absorbances between the initial and left dyes. In
addition, the stability of the aptamer−MSN system in mouse serum at
37 °C has been investigated by estimation of the precipitation of the
aptamer−MSN and the dye leakage from the pores.
Extraction of ATP from the Ramos Cells. Ramos cells were used
to demonstrate the application of this ATP-responsive controlled
release system upon the ATP obtained from the living cells. Ramos
and design the ATP-responsive MSN-based system is very
encouraging.
It is well-known that a variety of fluorescent,20 electrochemical,21 and colorimetric aptasensors22 have been developed
for ATP detection based on the aptamer−ATP interaction.23
Meanwhile, taking advantages of the unique characteristic and
chemical structure of ATP aptamer, the ATP-responsive MSNbased systems have also been reported and demonstrated
improved performance in controlled release. For example, Yang
et al.24 employed aptamer-modified Au nanoparticle to close
the pores, which can be opened in the presence of ATP
through the competitive binding. They have demonstrated for
the first time that the aptamer−target interaction could be used
as a stimuli-responsive mechanism in controlled-release
systems. However, the Au nanoparticles must be involved in
this system. Subsequently, Ö zalp et al.25 selected ATP aptamer,
which extended 8 bases so that it can hybridize with the first 8
bases in its 5′-end to close the pore. As we known, the
extending or decreasing of aptamer sequence might affect its
affinity and selectivity. For overcoming the above limitations,
we design a facile and effective ATP-responsive controlledrelease system using ATP aptamer-functionalized MSN. In the
system, ATP aptamer was hybridized with arm ssDNA1 and
arm ssDNA2 to form sandwich-type DNA structure and
subsequently grafted on the pore outlets of MSN to block the
nanochannels of MSN. In the absence of ATP, the pores of
MSN were blocked, and the leakage of guest molecules was
inhibited. In the presence of ATP, a competitive reaction took
place due to higher affinity and tighter binding of ATP aptamer
with ATP than that of ATP aptamer with arm ssDNA, resulting
in the opening of pores and the release of guest molecules. To
demonstrate the feasibility of this principle, Ru(bipy)32+ was
selected as a model guest molecule to investigate the controlled
release behavior of the aptamer-functionalized MSN system.
■
EXPERIMENTAL SECTION
Chemicals and Materials. N-Cetyltrimethylammonium bromide
(CTAB, ≥99%), CuBr (99.9%), tris(hydroxymethyl)aminomethane
(Tris), and 3-chloropropyltrimethoxysilane (ClTMS, 97+%) were
purchased from Alfa Aesar. Sodium azide (NaN3, 99%) and mouse
serum were obtained from Dingguo reagent Co. (Beijing, China).
[Ru(bipy)3]Cl2 (bipy = 2,2′-bipyridine) dye (Ru(bipy)32+) was
purchased from Sigma-Aldrich. Sodium hydroxide (NaOH), ammonium hydroxide solution (25%), N,N-dimethylformamide (DMF), and
tetraethylorthosilicate (TEOS, 28%) were purchased from Xilong
reagent Co. (Guangdong, China). Adenosine 5′-triphosphate (ATP),
guanosine 5′-triphosphate (GTP), cytosine 5′-triphosphate (CTP),
and uridine 5′-triphosphate (UTP) were purchased from ShangHai
BoYa Biotechnology Co. Ltd. All buffers were prepared with ultrapure
Milli-Q water (resistance >18.2 MΩ cm−1). The oligonucleotides were
synthesized by Sangon Biotechnology Inc. (Shanghai, China). The
sequences are as follows: ATP-aptamer, 5′-CAC CTG GGG GAG
TAT TGC GGA GGA AGG TT-3′; arm single-stranded DNA1 (arm
ssDNA1), 5′-alkyne-TTC CTC CGC A-3′; arm single-stranded DNA2
(arm ssDNA2), 5′-alkyne-ATA CTC CC-3′; nonaptamer structure
DNA (control DNA), 5′-TTT TTT TGG GAG TAT TGC GGA
GGA ATT TT-3′. The Tris-HCl buffer contains 10 mM Tris and 100
mM NaCl, pH 7.4. Ramos cells (B cell line, human Burkitt’s
lymphoma) were purchased from the Cancer Institute & Hospital
(Chinese Academy of Medical Sciences).
Characterization. High-resolution transmission electron microscopy (HRTEM) image was obtained from a JEOL 3010 microscope
with an accelerating voltage of 100 kV. Scanning electron microscopy
(SEM) image was obtained from a JSM-6700F microscope. The
hydrodynamic diameter and size distribution of MSN were measured
by dynamic light scattering (DLS), Malvern Inc., England. Fourier
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cells were cultured in RPMI 1640 medium supplemented with 15%
fetal bovine serum (FBS) and 100 IU/mL penicillin−streptomycin
solution and incubated at 37 °C in a humidified incubator containing 5
wt %/vol CO 2 . The cell density was determined using a
hemocytometer prior to any experiments. Well cultured Ramos cells
were centrifuged at 3000 rpm for 5 min to remove the growth medium
and washed with physiological saline solution three times, and then
resuspended in 1 mL of physiological saline solution. The suspension
was experimentally subjected to three freeze−thaw cycles (30 min
each at −20 and +40 °C) to break the cell membrane so that the ATP
can skip into the physiological saline solution.
Ru(bipy)32+ Releasing. One milligram of aptamer−MSN was
dispersed in 1 mL of Tris-HCl buffer containing 20 mM commercial
ATP at 25 °C. Subsequently, 0.20 mL of supernatant was taken
periodically from the suspension at 25 °C followed by centrifugation
(15 000 rpm, 10 min). The release of Ru(bipy)32+ from the pore voids
to the buffer solution was determined by fluorescence emission
spectroscopy (ex at 454 nm, em at 598 nm). In addition, the response
of the aptamer−MSN to the ATP obtained from living cells has also
been investigated following the same procedures.
of as-synthesized MSN was investigated by scanning electron
microscopy (SEM), transmission electron microscopy (TEM),
dynamic light scattering (DLS), X-ray diffraction patterns
(XRD), and N2 adsorption−desorption isotherms. As shown in
Figure 1, the SEM image (Figure 1a) illustrated that the as-
Figure 1. SEM (a) and TEM (b) images of as-synthesized MSN.
■
RESULTS AND DISCUSSION
For the design of the stated controlled release system, two
components were chosen, a solid supporter and the ATPresponsive molecule-gated switch. In this work, MCM-41-type
MSN was selected as a suitable inorganic material because of its
unique features. For the gating mechanism, our attention was
focused on the ATP aptamer. The working principle was
illustrated in Scheme 1. ATP aptamer, arm ssDNA1, and arm
synthesized MSN had narrow size distribution. The TEM
image (Figure 1b) showed a typical hexagonally arranged
porosity and well disparity of MSN containing parallel pores
with two openings. From the TEM image, it was demonstrated
that the diameter of MSN was about 80 nm. Here, we also
measured the particle size in aqueous suspension by DLS. It
was shown that the diameter of MSN was 208 nm with a
polydispersity index (PDI) of 0.105 (Figure S1). The reason for
this was that the DLS values of nanoparticles in aqueous
solution are always larger than solid-state diameters in a
monolayer in air by TEM due to the swell in aqueous solution.
The low-angle reflection typical of a hexagonal array, which
could be indexed as (100) Bragg peaks, further confirmed the
structure of MSN (Figure 2a). Moreover, the N2 adsorption−
desorption isotherms (Figure 2b) of the material presented an
Scheme 1. Schematic Illustration of Aptamer-Based ATPResponsive MSN System
ssDNA2 were first hybridized to each other to form ATP
aptamer-containing sandwich-type DNA structure. The immobilization of the sandwich-type DNA structure on the
surface of MSN would then result in the blockage of pores and
the package of guest molecules. However, the addition of ATP
as the target molecule induced a competitive displacement
reaction to the sandwich-type DNA structure. ATP aptamer
combined with ATP molecule and departed from the sandwichtype DNA structure. The guest molecules could be released
because the left flexible arm ssDNA on the surface of MSN
could not block the pores. To clearly show the loading and
release processes, Ru(bipy)32+ dye was chosen as a model guest
molecule.
Following this procedure, we first synthesized MSN
according to the previously reported procedure.18 The structure
Figure 2. (a) XRD and (b) BET nitrogen sorption isotherms of MSN.
Inset: BJH pore volume and pore size distribution plots of MSN.
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adsorption step at an intermediate P/P0 value (0.2−0.4) typical
of MCM-41-type structure. The application of BET model
resulted in a value that the total surface area is 963.5 m2 g−1.
The BJH curve showed a narrow pore size distribution of MSN
with an average pore diameter of 2.8 nm and pore volume of
1.0 cm3 g−1 (Figure 2b, inset). The characteristics of the assynthesized MSN were all summarized in Table S1.
The as-synthesized MSN was then modified with a chlorine
group and further treated with sodium azide in DMF to
obtained MSN-N3. Ru(bipy)32+ was then chosen as guest
molecule to load into the pores of MSN-N3 by soaking the
particles into 2 mM of Ru(bipy)32+ solution. Subsequently, the
preobtained ATP aptamer-containing sandwich-type DNA
structure was immobilized on the MSN-N3 surface to block
the pores and reserve the Ru(bipy)32+, giving rise to the
aptamer−MSN samples. The loading amount of Ru(bipy)32+
was determined to be 215.0 μmol g−1 SiO2 calculated using the
standard curve of Ru(bipy)32+ (Figure S2 in the Supporting
Information). The successful conversion of the MSN surface
was confirmed by FTIR. As shown in Figure 3, the absorption
Figure 4. Time course of Ru(bipy)32+ release profiles: (a) aptamer−
MSN without ATP, (b) con DNA−MSN without ATP, (c) aptamer−
MSN with 20 mM ATP, and (d) con DNA−MSN with 20 mM ATP.
pores would be opened. As for the con DNA−MSN, in the
presence of ATP, the release of Ru(bipy)32+ from the pores was
much lower than that from the aptamer−MSN system (Figure
4d). Of course, 21.2% release was still observed when the con
DNA−MSN was treated with 20 mM ATP for 7 h. The most
probable reason was that the control DNA sequence has the
same 18 bases as that of the ATP−aptamer. Therefore, the ATP
might also nonspecifically bind with the control DNA and
result in some release. In addition, the amount of released
Ru(bipy)32+ from the aptamer−MSN was dependent on the
added amount of ATP molecules (Figure 5). In a lower ATP
Figure 3. FTIR spectra of materials before and after modification: (a)
MSN, (b) MSN-N3, and (c) aptamer−MSN.
band around 2110 cm−1 was assigned to the azide stretch. MSN
(Figure 3a) had no adsorption around 2110 cm−1, while MSNN3 (Figure 3b) showed obvious absorption band at that
wavenumber, indicating that the −N3 group was successfully
grafted onto the surface of MSN. As compared to MSN-N3, the
adsorption band of aptamer−MSN (Figure 3c) around 2110
cm−1 was distinctively declining, confirming the reaction
between alkynes and −N3 group. It suggested that the ATP
aptamer-containing sandwich-type DNA structure was successfully tethered to MSN-N3. The quantity of sandwich-type DNA
anchored on the surface of MSN was determined by UV−vis
spectroscopy to be approximately 1.6 μmol g−1 SiO2 (Figure S3
in the Supporting Information).
Continuous guest molecules release experiments had been
done to test the gate property of the designed system. The
ATP-triggered release of Ru(bipy)32+ was monitored by
fluorescence emission spectroscopy at 598 nm. To both
Ru(bipy)32+ loaded aptamer−MSN and con DNA−MSN
(Figure 4a and b), a negligible Ru(bipy)32+ release from the
pores was observed when ATP was not added, indicating a
good retention efficiency of Ru(bipy)32+ in the pores of the
MSN by virtue of capping with sandwich-type DNA structure.
In contrast, the Ru(bipy)32+ release from the aptamer−MSN
system reached 83.2% of the total load after the introduction of
20 mM ATP for 7 h (Figure 4c). The result demonstrated that
the aptamer-containing sandwich-type DNA structure could be
dissociated through a competitive binding with ATP, and the
Figure 5. Controlled release of Ru(bipy)32+ from aptamer−MSN
system triggered by ATP as a function of concentration, measured
after 48 h.
concentration region, the fluorescence intensity of the released
Ru(bipy)32+ increased dramatically with the increase of added
ATP amount. While the rising rate of fluorescence intensity
slowed when the ATP concentration was higher than 5 mM, it
was mainly a constant when the ATP concentration was higher
than 20 mM. The maximum release was observed at 20 mM of
ATP, suggesting that the ATP aptamer combined with ATP
thoroughly at this ATP concentration.
In the design of controlled release systems, the selectivity is
very important. Further control experiments were implemented
to investigate the selectivity of the aptamer−MSN system.
Figure 6 illustrated that ATP analogues, such as GTP, CTP,
and UTP, induced a small quantity of guest molecules to
release, while ATP induced a dramatic release of loaded guest
molecules. This result obviously indicated that the aptamer−
MSN system had sufficient selectivity to ATP and was able to
discriminate ATP from its analogues.
In addition, the stability of the particle is very important in
cargo release application. To explore the application potential
of the aptamer−MSN system, we investigated its stability in
mouse serum at 37 °C by estimation of the precipitation of the
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It is well-known ATP is a multifunctional nucleoside
triphosphate and widely used in cells as energy metabolism.
To demonstrate whether the developed aptamer−MSN
controlled release system could be responsive to the ATP
obtained from living cells, we investigated the release effect of
the aptamer−MSN upon the ATP in the Ramos cells extracts.
As shown in Figure 8, the fluorescence intensities in the
Figure 6. Selectivity of the aptamer−MSN system. The concentration
of ATP, CTP, UTP, and GTP is 20 mM, respectively. The results were
measured after incubation for 7 h.
particles and the dye release. As can be seen in Figure 7, the
Ru(bipy)32+ released percentage was only about 4.1% when the
Figure 8. Dye release effect of the aptamer-based drug delivery system
using ATP extracted from living cells as stimuli: (a) aptamer−MSN in
physiological saline solution, (b) aptamer−MSN in broken cell
suspension (7.1 × 107 cells mL−1), (c) con DNA−MSN in
physiological saline solution, and (d) con DNA−MSN in broken
cell suspension (7.1 × 107 cells mL−1). All were treated for 7 h.
supernatant of aptamer−MSN and con DNA−MSN were both
relatively low when they were treated with the pure
physiological saline solution at 25 °C for 7 h, indicating few
leakages of guest. Differently, the fluorescence intensity in the
supernatant of aptamer−MSN was greatly increased after it was
incubated in the physiological saline solution containing
extracted ATP from Ramos cells for 7 h, suggesting that the
aptamer−MSN system could also respond to ATP, which was
extracted from the living cells. What is more, the fluorescence
intensity in the supernatant of con DNA−MSN was much
lower than that of aptamer−MSN, indicating that the aptamer−
MSN system also kept its good selectivity in cell lysates.
Figure 7. Time course of Ru(bipy)32+ release profile in mouse serum
at 37 °C. Inset: (a) mouse serum; (b) aptamer−MSN in mouse serum,
0 h; (c) aptamer−MSN in mouse serum, 3 h; (d) aptamer−MSN in
mouse serum, 7 h; and (e) the centrifugation of aptamer−MSN after
incubating in mouse serum for 7 h.
aptamer−MSN particles were treated with mouse serum at 37
°C for 3 h. When the incubation time was further increased to 7
h, the Ru(bipy)32+ release percentage was increase to 9.2%. By
comparison with the aptamer−MSN that was incubated with
20 mM ATP for 7 h, the dye release of aptamer−MSN in the
mouse serum at 37 °C for 7 h was much lower. These results
indicated that the ATP aptamer containing sandwich-type DNA
can also effectively block the pores of MSN in the mouse
serum. The photo pictures inset in Figure 7 displayed the
precipitation procedure of aptamer−MSN in mouse serum at
37 °C. The color of mouse serum (Figure 7, inset a) was pale
yellow and transparent, while the color of aptamer−MSN
dispersed mixture (Figure 7, inset b) was deeper and
nontransparent, implying that the particles dispersed in
mouse serum well. As displayed in Figure 7, inset c, there
was a small quantity of orange-yellow particles precipitate in the
bottom of the centrifuge tube after it was dispersed in mouse
serum for 3 h due to the action of gravity, but the supernatant
was still relatively opaque. It suggested that the majority of the
particles were still well-dispersed in the serum. Differently, a
large proportion of the orange-yellow particles precipitated 7 h
later, and the supernatant became semitransparent (Figure 7,
inset d). When the mixture was centrifuged, as Figure 7, inset e
displayed, the supernatant of the sample was transparent again,
and the color was almost the same as that of the mouse serum,
confirming there was no obvious leakage of Ru(bipy)32+ and
good blockage behavior of the sandwich-type DNA. The data
above indicated that the aptamer−MSN was stable in
physiological-like conditions.
■
CONCLUSION
We have designed an easy-to-achieve and prevalent aptamerbased ATP-responsive MSN system. In this system, ATP
aptamer was hybridized with two arm ssDNA to form the
sandwich-type DNA structure and then tethered to the surface
of MSN to inhibit the guest molecules from skipping out of the
pores. The Ru(bipy)32+ molecules as model guests had been
encapsulated into pores of MSN. The release profile was
dependent on the leaving of aptamer due to the competitive
binding between ATP and ATP aptamer. The results
demonstrated that the system had a high loading amount of
guests (215.0 μmol g−1 SiO2) and good release behavior in the
presence of ATP. Furthermore, the system responded not only
to the commercial ATP but also to the ATP, which was
extracted from the living cells. This proof of concept might
promote the application of aptamer in constructing other
target-responsive MSN-based drug delivery systems by using
aptamers of other targets such as ions, small biological
molecules, biological macromolecules, bacteria, and cells
instead of ATP−aptamer. Also, it can promote the application
of ATP-responsive systems.
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ASSOCIATED CONTENT
S Supporting Information
*
Size distribution of as-synthesized MSN in aqueous suspension
measured by dynamic light scattering (DLS), standard curve of
Ru(bipy)32+ measured by UV−vis spectrometer and UV−vis
spectra of initial and left Ru(bipy)32+, and a table corresponding
to their characterization. This material is available free of charge
via the Internet at http://pubs.acs.org.
■
AUTHOR INFORMATION
Corresponding Author
*E-mail: [email protected].
Author Contributions
†
These authors contributed equally.
Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS
This work was supported in part by the Project of Natural
Science Foundation of China (21175039, 20905023, and
21190044), Key Technologies Research and Development
Program of China (2011AA02a114), Research Fund for the
Doctoral Program of Higher Education of China
(20110161110016), and the project supported by Hunan
Provincial Natural Science Foundation and Hunan Provincial
Science and Technology Plan of China (10JJ7002,
2011FJ2001).
■
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