Fluorescent ZnS Quantum Dots-Phosphoethanolamine Nanoconjugates for Bioimaging Live Cells in Cancer Research

Abstract

Many human diseases, including metabolic, immune, and central nervous system disorders, as well as several types of cancers, are the consequence of an important alteration in lipid-related metabolic biomolecules. Although recognized that one of the most important metabolic hallmarks of cancer cells is deregulation of lipid metabolism, the multiple complex signaling pathways are poorly understood yet. Thus, in this research, novel nanoconjugates made of ZnS quantum dots (QDs) were directly synthesized in aqueous media using phosphoethanolamine (PEA) as the capping ligand, which is an important biomolecule naturally present in cells for de novo biosynthesis of fatty acids and phospholipids involved in the cell structure (e.g., membrane), differentiation, and cancer growth. These QD-PEA bio-nanoconjugates were characterized by spectroscopical and morphological techniques. The results demonstrated that fluorescent ZnS nanocrystalline QDs were produced with uniform spherical morphology and estimated sizes of 3.3 ± 0.6 nm. These nanoconjugates indicated core-shell colloidal nanostructures (ZnS QD-PEA) with the hydrodynamic diameter (H _D) of 26.0 ± 3.5 nm and ζ-potential centered at -30.0 ± 4.5 mV. The cell viability response using mitochondrial activity assay in vitroconfirmed no cytotoxicity at several concentrations of PEA (biomolecule) and the ZnS-PEA nanoconjugates. Moreover, these nanoconjugates effectively behaved as fluorescent nanomarkers for tracking the endocytic pathways of cancer cells using confocal laser scanning microscopy bioimaging. Hence, these results proved that biofunctionalized ZnS-PEA nanoprobes offer prospective tools for cellular bioimaging with encouraging forecast for future applications as active fluorescent biomarker conjugates in metabolic-related cancer research.

Figure 1

ZnS–PEA QDs characterization: (A) UV–vis spectroscopy; (B) PL spectroscopy; (C) TEM image; (D) HRTEM image with the typical QD size (diameter) and nanocrystal lattice fringes (ED pattern); (E) histogram of nanoparticle size distribution; and (F) schematic representation of QDs with indication of ZP and hydrodynamic radius.

Figure 2

Cell viability responses by MTT assay after 48 h of incubation of (A) SAOS and MCF7 cell lines with PEA at different concentrations and (B) at concentration of 100 μM with different cell types.

Figure 3

Cell viability responses by MTT assay of incubation of ZnS–PEA with SAOS and MCF7 cell lines for (A) 48 h, 5 days, and 7 days and (B) after incubation with different cell lines for 48 h (at 350 μM equivalent of PEA biomolecule; 10 nM of ZnS–PEA colloidal nanoconjugates).

Figure 4

Confocal microscopy imaging of the cellular uptake of the ZnS–PEA nanoconjugates by MCF7 cells vs incubation time (15, 30, and 60 min): (A) PL image and (B) PL + bright field image (scale bar = 10 μm).

Figure 5

Confocal microscopy imaging of the cellular uptake of the ZnS–PEA nanoconjugates by SAOS cells vs incubation time (15, 30, and 60 min): (A) PL image and (B) PL + bright field image (scale bar = 10 μm).

Figure 6

Confocal microscopy imaging of the cellular uptake of the ZnS–PEA nanoconjugates by HEK 293T cells vs incubation time (15, 30, and 60 min): (A) PL image and (B) PL + bright field image (scale bar = 10 μm).

Figure 7

(A) Fluorescence image (inset: bright field and PL images merged; white arrows: nucleus) and intensity fluorescence profiles along (B) A–A line and (C) B–B line for MCF7 cell line after 60 min of incubation with ZnS–PEA nanoconjugates.

Figure 8

CLSM images showing intracellular uptake and distribution of ZnS–PEA nanoconjugates by (A) SAOS and (B) HEK 293T cells after 60 min of incubation. (a) Images of DNA staining with TO-PRO-3 in red color; (b) images of ZnS–PEA green emission; and (c) merged images of both biomarkers.

Figure 9

Cell uptake profile based on MFI as function of incubation time for (a) MCF7, (b) SAOS, and (c) HEK 293T cell lines.

Figure 10

Schematic representation of cellular uptake and endocytosis pathway of ZnS–PEA conjugates (not to scale).

Figure 11

Fluorescence images showing (A) red emission of LysoTracker, (B) green emission of ZnS–PEA, (C) overlapped CLSM images of red and green emissions, and (D) bright field image + overlapped PL emissions for SAOS cells after 60 min of incubation with ZnS–PEA QD.