Organically modified silica nanoparticles: A nonviral vector for in vivo gene delivery and expression in the brain
- Dhruba J. Bharali*,
- Ilona Klejbor*,†,‡,
- Ewa K. Stachowiak*,†,
- Purnendu Dutta*,
- Indrajit Roy*,
- Navjot Kaur*,
- Earl J. Bergey*,
- Paras N. Prasad*,§, and
- Michal K. Stachowiak*,†,§
- *Institute for Lasers, Photonics, and Biophotonics, Department of Chemistry, State University of New York, Buffalo, NY 14260; †Molecular and Structural Neurobiology and Gene Therapy Program, Department of Pathology and Anatomical Sciences, State University of New York, Buffalo, NY 14214; and ‡Department of Anatomy and Neurobiology, Medical University of Gdansk, 80-210 Gdansk, Poland
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Communicated by Tobin J. Marks, Northwestern University, Evanston, IL, June 13,2005 (received for review March 28, 2005)
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Fig. 1.Transmission electron micrograph of ORMOSIL/pEGFP-N2 nanoparticles.
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Fig. 2.ORMOSIL nanoparticle transfection in the SNc. (A) DNA-free ORMOSIL injection showing no substantial immunostaining for EGFP. (B–E) Injection of ORMOSIL-pEGFP-N2 complex into SNc. (B) Multiple cells with typical dopaminergic neuron morphology are immunostained positive for EGFP. (C) No immunostaining is observed without primary anti-EGFP Ab. (D) EGFP immunostaining of neuron-shaped cells (higher magnification). (E) Transfected EGFP (green) is expressed in TH-immunopositive (red) dopaminergic neuron.
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Fig. 3.Expression of EGFP in multiple brain areas after injection of ORMOSIL-pEGFP-N2 into the brain LV. Brain sections were immunostained with EGFP antibodies as described in Materials and Methods.(A and B) Control ORMOSIL nanoparticles. (A) The region surrounding the LV. Str, striatum; Sep, septum; cc, corpus callosum. (B) The hippocampal region adjacent to the ventricle. No cellular staining is detected in either region by using anti-EGFP immunocytochemistry. (C–F) ORMOSIL/pEGFP-N2 particles. Injection resulted in EGFP immunostaining of the neuron-shaped cells in dorsal lateral (d), lateral (l), and medial (m) septal nuclei (C); in the adjacent striatal region of the brain (D); cingulate and motor cortex (E); and pyramidal neurons of the CA3 hippocampal region (F).
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Fig. 4.Transfection of ORMOSIL-pEGFP-N2 complex into the LV cells of the SVZ. Mice were transfected with ORMOSIL/pEGFP-N2 by injection into the brain LV. (A and B) Seven days postmortem EGFP immunostaining is shown at low magnification (A) and at higher magnification (B) of the positive region to visualize transfected cells. (C and D) In vivo imaging of EGFP fluorescence in cells in the LV. Ten days after transfection, mice were subjected to the second stereotaxic surgery, and a miniature fiber-optic Cell-viZio probe was inserted into the anterior dorsal region (C) or the posterior region (D) of the LV >15 μm from the medial ventricular wall. Dynamic sequences were recorded, and selected frames are shown. The complete dynamic sequences are included in Movie 1, which is published as supporting information on the PNAS web site.
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Fig. 5.Modulation of cell proliferation by using ORMOSIL transfection of nonmembrane/nucleus-targeted FGFR1(SP-/NLS). Control ORMOSIL (A, C, and E) or ORMOSIL/pFGFR1(SP-/NLS) (B, D, and F) was injected into the anterior region of the brain LV. Seven days later, the animals were injected with BrdUrd (i.p.) and were perfused 5 h later. Sagittal brain sections were immunostained for FGFR1 or DNA that had incorporated BrdUrd. (A and B) Immunostaining of SVZ with FGFR1 McAb6. (C and D) BrdUrd immunostaining of cell nuclei in SVZ and adjacent tissue. (E and F) BrdUrd immunostaining of cells in the rostral migratory stream close to SVZ.
Footnotes
- Copyright © 2005, The National Academy of Sciences
