Magnetic silica spheres with large nanopores for nucleic acid adsorption and cellular uptake

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Magnetic silica spheres with large nanopores for
nucleic acid adsorptionand cellular uptake

• Jian Liu, Bo Wang, Sandy Budi Hartono
  
• Biomaterials
• University of Queensland, Australia

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contents

• Introduction
• Experimental Section
• Results and Discussion
• Conclusions

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Introduction

• Mesoporous materials
• Large speci?c surface area
• Large pore volume
• Uniform pore size distribution
• Mesoporous silica nanoparticles (MSNs)
• Biocompatibility
• Low toxicity

• Catalysis
• Imaging
• Drug delivery

• Biological application

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Introduction

• Other properties that MSNs required for
• biological application
• Large pore sizes
• Appropriate magnetic properties
• Appropriate functional surface

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Introduction

• 1?Large pore size
• Large internal surface
• Large mesoporous volume
• Cytochrom C 2.63.23.3
  nm
• a-L-arabinofuranosidase 3.99.714.4 nm

Suitable pore sizes for immobilisation of
these proteins can vary from10 to 50 nm
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Introduction

• 2?Magnetic properties
• Bioseparation
• Cell sorting
• Diagnostic analysis
• Simultaneous imaging and drug delivery

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Introduction

• Preparation methods
• Large pore size mesoporous materials
• TemplatesPluronic P123
• Swelling agent1,3,5-trimethyl benzene (TMB)
  or alkanes
• ConditionStrong acidic
• Magnetic mesoporous materials
• TemplatesBrij56 micelles
• ConditionBasic

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Introduction

• 3?Functional surface
• To delivery nucleic acids, the silica
  surface with positive charge is needed to
  electrostatically bind DNA and RNA molecules
• Methods
• Functionalisation with amine-derivative
• group such as APTES
• Conjugations with cationic polymers such as PEI

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Introduction

• Develop synthesis methods to prepare MSNLP
• Establish a surface functionalisation method to
  enable adsorption and delivery of nucleic acids

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Experimental Section

• Synthesis of monodisperse superparamagnetic Fe3O4
  
• nanocrystals

Static conditions at 250? in a Te?on-lined
autoclave for 612 h
The concentration of the magnetic nanocrystals is
10 or 30 mg/ mL and suspended in hexane
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Experimental Section
Synthesis of magnetic silica nanospheres with
large nanopores
PLL
30-glycidox-ypropyltrimethoxysilane (GOPS)
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Experimental Section

• DNA adsorption
• CpG DNA 1826 (5 to 3, TCCATGACGTTCCTGACGTT)
• Measuring A260 absorbance at 260 nm
• Transfection of cells
• CyTM3-labeled miRNA
• Rat kidney epithelial cells (NRK-52E)

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Results and Discussion

• Synthesis of magnetic silica nanospheres with
  large
• nanopores

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Results and Discussion
Fig. 1. SEM (a, c), TEM (b, d-f), and HRTEM (g,
h) images of MSNLP synthesised with different
amount of hexane MSNLP-0-350 (a, b),
MSNLP-10-350 (c, d), MSNLP-10-700 (e),
MSNLP-10-1400 (f-h).
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Results and Discussion
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Results and Discussion

• Brij56 polyoxyethylene 10 cetyl ether, C16H33EO10

N0I0 route
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Results and Discussion

• Magnetic properties of magnetic silica
  nanospheres with
• large nanopores

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Results and Discussion
Fig. (A) Field-dependent magnetisation at 300 K
of MSNLP with different amounts of magnetite a)
MSNLP-10-350, b) MSNLP-10-700, c) MSNLP-10-1400,
and d) MSNLP-30-1400 and (B) the separation
process of MSNLP-30-1400 nanospheres from
solution by magnet (right picture) and their
re-dispersion by as slight shake (left picture).
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Results and Discussion

• Composition of PLL functionalised MSNLP

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Results and Discussion

• Adsorption of DNA on PLL functionalised
• magnetic silica nanospheres with largenanopores

MSNLP-0-350-PLL qm22.5µg/mg MSNLP-10-350-PLL qm
15µg/mg MSNLP-10-1400-PLL qm10µg/mg
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Results and Discussion
Fig. Left panel a-d, cells transfected with
?uorescent oligonucleotide only middle panel
e-h, cells transfected with nanoparticles alone
and right panel i-l, cells transfected with
nanoparticles loaded with ?uorescent
oligonucleotide. From top to bottom cy5 channel
- images of ?uorescence of CyTM3 labeled miRNA
(red), F-actin - images of F-actin stained by
FITC-Phalloidin (green), DAPI - images of nuclei
stained with DAPI (blue), and merge - the merged
picture.
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Conclusions

• Magnetic silica nanospheres with large
  nanopores(13-20 nm) were synthesised for the ?rst
  time
• The saturation magnetisation values can be
  conveniently controlled by changing the amount of
  Fe3O4 magnetic nanocrystals encapsulated
• After functionalisation with PLL, high adsorption
  capacity ranging from 10 to22.5 µg/mg for CpG DNA
  and ef?cient cellular delivery capability for
  miRNA were achieved
• The materials synthesised in this study could ?nd
  broad applications

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Thank You