Acceleration of gene transfection efficiency in neuroblastoma cells through polyethyleneimine/poly(methyl methacrylate) core-shell magnetic nanoparticles

Figures & data

Table 1 Polymerase chain reaction primers used for tryptophan hydroxylase-2 cloning and semiquantitative assay of GAPDH and tryptophan hydroxylase-2 gene expressions

Primer	Sequence	Product (Kb)	Reference
TPH-2-NheI_pGL-CMV	5′-CCT gCT AgC gCC TTC CTC TCA ATC TC-3′	1.5	The present study
TPH-2-XbaI_pGL-CMV	5′-CCC gCT CTA gAT AgT TCC Agg CAT CAA ATC C-3′
GAPDH sense	5′-gAC CAC AgT CCA TgC CAT CAC T-3′	0.4	Divya and Pillai^Citation22
GAPDH antisense	5′-TCC ACC ACC CTg TTg CTg TAg-3′
TPH-2 sense	5′-AAC CAC TAT TgT gAC gCT gAA TCC TCC AgA gAA-3′	0.2	The present study
TPH-2 antisense	5′-ACC CAT AAC CCA TCg CCA CAT CCA CAA AA-3′

Abbreviations: CMV, cytomegalovirus; TPH-2, tryptophan hydroxylase-2.

Figure 1 Transmission electron microscopic image. (A) Mag-PEI and histogram showing size distribution. (B) Mag-PEI nanoparticles with high magnetic loading.

Abbreviation: Mag-PEI, magnetic poly(methyl methacrylate) core/ polyethyleneimine shell.

Table 2 Size and zeta potential of mag-PEI nanoparticle/DNA magnetoplex at N/P ratios of 0.4/1, 0.8/1, 1.6/1, 4.3/1, 8.7/1, and 17.5/1

	N/P	Size (nm)	Zeta potential (mV)	PDI
Mag-PEI NP		123.8 ± 3.1	39.3 ± 1.9	0.26 ± 0.02
Mag-PEI NP/DNA	0.4/1	231.3 ± 24.7	6.3 ± 1.7	0.36 ± 0.03
	0.8/1	370.0 ± 32.4	25 ± 1.4	0.45 ± 0.05
	1.6/1	298.5 ± 65	34.8 ± 2.3	0.75 ± 0.15
	4.3/1	286.8 ± 23.3	22.4 ± 3.2	0.42 ± 0.06
	8.7/1	251.3 ± 5.1	5.6 ± 0.5	0.34 ± 0.06
	17.5/1	215.4 ± 17.5	3.3 ± 0.7	0.36 ± 0.05

Abbreviations: Mag-PEI NP, magnetic poly(methyl methacrylate) core/polyethyleneimine shell nanoparticles; PDI, polydispersity index.

Figure 2 Gel retardation assay.

Notes: One microgram of plasmid DNA was applied to the magnetoplex with mag- PEI nanoparticles at different N/P ratios. Lane 1 is the control DNA without mag-PEI nanoparticles. Lanes 2–7 represent mag-PEI NP/DNA magnetoplexes with N/P ratios of 0.4/1, 0.8/1, 1.6/1, 4.3/1, 8.7/1, and 17.5/1.

Abbreviation: Mag-PEI, magnetic poly(methyl methacrylate) core/polyethyleneimine shell.

Figure 3 Atomic force microscopy images of mag-PEI nanoparticles (A) mag-PEI nanoparticles forming magnetoplexes with DNA at N/P ratios of 0.81/1 (B) and 4.3/1 (C).

Abbreviation: Mag-PEI, magnetic poly(methyl methacrylate) core/polyethyleneimine shell.

Figure 4 Transfection efficiency (A) and cytotoxicity (B) of mag-PEI nanoparticles at 15, 30, 60, 120, and 180 minutes in LAN-5 cells.

Notes: The transfection efficiency and cytotoxicity was compared with positive control Lipofectamine 2000™, PolyMAG, and negative controls (naked DNA, plasmid pGL-3-basic containing CMV promoter/enhancer). *Significant differences between cells transfected with and without a magnetic plate in each transfection reagent (P < 0.05). The gray and white bars show the results of cells incubated with or without magnetic induction, respectively.

Abbreviation: Mag-PEI, magnetic poly(methyl methacrylate) core/polyethyleneimine shell.

Figure 5 Confocal image of LAN-5 cells 24 hours after transfection. Cells incubated with or without a magnetic plate for (A) 60 minutes and (B) 180 minutes were used for investigation of the cellular uptake of mag-PEI nanoparticles.

Note: Green, acridine orange-stained live cells; red, RITC-stained mag-PEI nanoparticles.

Abbreviations: RITC, rhodamine-B-isothiocyanate; Mag-PEI, magnetic poly(methyl methacrylate) core/polyethyleneimine shell.

Figure 6 Semiquantitative reverse-transcriptase polymerase chain reaction result shows expression of the TPH-2 gene in LAN-5 24 hours after transfection by mag-PEI nanoparticles compared with positive control Lipofectamine 2000™ and PolyMAG, and negative control (naked DNA).

Notes: *Significant differences between cells transfected with and without magnetic plate in each transfection reagent (P < 0.05). The gray bars and white bars show the results of cells incubated with and without magnetic induction, respectively.

Abbreviations: Mag-PEI, magnetic poly(methyl methacrylate) core/polyethyleneimine shell; TPH-2, tryptophan hydroxylase-2.

DivyaCSPillaiMRAntitumor action of curcumin in human papillomavirus associated cells involves downregulation of viral oncogenes, prevention of NFkB and AP-1 translocation, and modulation of apoptosisMol Carcinog20064532033216526022

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