Exploring the Interaction of Cobalt Oxide Nanoparticles with Albumin, Leukemia Cancer Cells and Pathogenic Bacteria by Multispectroscopic, Docking, Cellular and Antibacterial Approaches

Abstract

Aim: The interaction of NPs with biological systems may reveal useful details about their pharmacodynamic, anticancer and antibacterial effects.

Methods: Herein, the interaction of as-synthesized Co₃O₄ NPs with HSA was explored by different kinds of fluorescence and CD spectroscopic methods, as well as molecular docking studies. Also, the anticancer effect of Co₃O₄ NPs against leukemia K562 cells was investigated by MTT, LDH, caspase, real-time PCR, ROS, cell cycle, and apoptosis assays. Afterwards, the antibacterial effects of Co₃O₄ NPs against three pathogenic bacteria were disclosed by antibacterial assays.

Results: Different characterization methods such as TEM, DLS, zeta potential and XRD studies proved that fabricated Co₃O₄ NPs by sol-gel method have a diameter of around 50 nm, hydrodynamic radius of 177 nm with a charge distribution of -33.04 mV and a well-defined crystalline phase. Intrinsic, extrinsic, and synchronous fluorescence as well as CD studies, respectively, showed that the HSA undergoes some fluorescence quenching, minor conformational changes, microenvironmental changes as well as no structural changes in the secondary structure, after interaction with Co₃O₄ NPs. Molecular docking results also verified that the spherical clusters with a dimension of 1.5 nm exhibit the most binding energy with HSA molecules. Anticancer assays demonstrated that Co₃O₄ NPs can selectively lead to the reduction of K562 cell viability through the cell membrane damage, activation of caspase-9, -8 and -3, elevation of Bax/Bcl-2 mRNA ratio, ROS production, cell cycle arrest, and apoptosis. Finally, antibacterial assays disclosed that Co₃O₄ NPs can stimulate a promising antibacterial effect against pathogenic bacteria.

Conclusion: In general, these observations can provide useful information for the early stages of nanomaterial applications in therapeutic platforms.

Keywords: antibacterial; anticancer; cobalt oxide; docking; nanoparticle; spectroscopy; synthesis.

Figure 1

(A) TEM images, (B) DLS histogram, (C) XRD pattern, and (D) XRD data of synthesized Co₃O₄ NPs by sol-gel method.

Figure 2

(A) Intrinsic fluorescence, (B) ANS fluorescence, (C) synchronous fluorescence study at Δλ = 60 nm, and (D) synchronous fluorescence study at Δλ = 20 nm of HSA after interaction with different concentrations [0 µg/mL (blue), 1 µg/mL (red), 10 µg/mL (green), 20 µg/mL (purple), 50 µg/mL (brown)] of Co₃O₄ NPs. The dotted circle shows the red or blue shift in the λ_max.

Figure 3

HEX 6.3 outcomes of the interaction of HSA with spherical Co₃O₄ nanoclusters, (A) r=0.5 nm, (B) r=1 nm, (C) r=1.5 nm, (D) r =2 nm.

Figure 4

The resulting docking pose of HSA after interaction with spherical Co₃O₄ nanoclusters, (A) r=0.5 nm, (B) r=1 nm, (C) r=1.5 nm, (D) r =2 nm.

Figure 5

(A) CD spectra, (B) quantified CD data of HSA after interaction with different concentrations [0 µg/mL (blue), 1 µg/mL (red), 10 µg/mL (green), 20 µg/mL (purple), 50 µg/mL (brown)] of Co₃O₄ NPs.

Figure 6

(A) MTT assay of lymphocytes and K562 cells incubated with different concentrations of Co₃O₄ NPs, (B) LDH assay of K562 cells incubated with different concentrations of Co₃O₄ NPs, (C) caspase assay of K562 cells incubated with IC50 concentrations of Co₃O₄ NPs, (D) qPCR assay of K562 cells incubated with IC50 concentrations of Co₃O₄ NPs. *P<0.5 and **P<0.01 relative to negative untreated cells. ^#P<0.5 relative to the activity of caspase-8.

Figure 7

ROS assay of (A) control K562 cells, (B) treated K562 cells incubated with IC50 concentrations of Co₃O₄ NPs, (C) statistical analysis histogram. ***P<0.01 relative to negative untreated cells.

Figure 8

Cell cycle assay of (A) control K562 cells, (B) treated K562 cells incubated with IC50 concentrations of Co₃O₄ NPs, (C) statistical analysis histogram. *P<0.5 and **P<0.01 relative to negative untreated cells.

Figure 9

Annexin-PI assay of (A) control K562 cells, (B) treated K562 cells incubated with IC50 concentrations of Co₃O₄ NPs, (C) statistical analysis histogram. **P<0.1 and ***P<0.001 relative to negative untreated cells.

Figure 10

Visible zone formed by different concentrations (200–3.12µg/mL) of Co₃O₄ NPs against S. aureus, E. coli and P. aeruginosa.