4. Conclusion

100 μg/mL. It was found that the length of NWs affected the internalization, with NWs longer

Semiconductor quantum dots (QDs) are light-emitting particles that have broad excitation spectra, long fluorescence lifetimes compared to traditional fluorescent probes and are more resistant to photobleaching [140, 141]. Also, they can easily be conjugated to proteins [140], which makes them excellent choices for bioimaging [142–146] and other biomedical applications [141, 147, 148]. Tsoi et al. summarized the toxicity of QDs by two mechanisms: degradation with the release of free cadmium (Cd) and generation of ROS [149]. Each design of QD is a unique combination and has its own physicochemical properties that may influence its biological activity and toxicity. As a result, tremendous research efforts have been devoted to produce high quality QDs by optimizing synthetic procedures, as well as functionalizing their

An early study by Derfus et al. demonstrated that CdSe-core QDs oxidized and degraded, releasing Cd ions which induced cell death [151]. When CdSe QDs were exposed to a UV-light for 1, 2, 4 and 8 h and then incubated with hepatocytes, it showed a 6, 42, 83 and 97% decrease in the cells' viability, respectively [143]. Cd is a known carcinogen with potential damage to the renal, skeletal, pulmonary and reproductive systems [152]. Interestingly, Chen et al. showed that the cell viability of HEK293 cells treated with 37.5 nM of 5 nm CdTe QDs was not significantly altered, compared to the control (i.e., untreated cells) after 3 days of incubation [153]. However, high concentrations (300–600 nM) of QDs completely inhibited cell growth from the very beginning. The cytotoxicity of QDs has also been linked to the generation of ROS, which in turn damages cellular proteins, lipids and DNA [149]. The p53 gene was also

Tracking the QDs internalization pathways could help explain their toxicological properties. To this end, microscopy studies showed that QDs localize within cellular endosomes and lysosomes, exposing them to an acidic or oxidative microenvironment [149]. It was determined that the hypochlorous acid present in phagocytic cells oxidized polymer-encapsulated CdS and ZnS-capped CdSe QDs, releasing cadmium, zinc, sulfur and selenium into the cytoplasm. Some studies have suggested that the QDs toxicity might derive from multiple factors including the environment and the QDs physicochemical characteristics (such as size, shape and surface chemistry). A surface coating with a ZnS shell [149] or BSA corona [150] reduced the QDs toxicity. In addition, polymeric coatings (i.e., phospholipid-PEG) and inorganic coatings (e.g., Si) can prevent the release of Cd into the biological media [142]. In a different approach, Soenen et al. studied cell viability using Cd-free QDs (ZnSe/ZnS and InP/ZnS QDs) at concentrations ranging from 0 to 100 nM [154]. Cytotoxic effects were observed starting from 60 nM for ZnSe to 80 nM for InP QDs. Further, no increase in cytotoxicity was reported up to 7 days

shown to be inhibited by CdTe QDs, leading to apoptosis and cell death [150].

after the initial cell labeling compared to normal QDs due to the absence of Cd.

than 5 μm being more difficult to be internalized, due to geometrical restrictions.

surface in order to enhance biocompatibility [142, 150].

3.5. Quantum dots

248 Cytotoxicity

Recent studies on the in-vitro cytotoxicity of carbon structure and semiconductors in biomedical applications were reviewed, taking into account nanoparticles and nanowires/nanorods. A summary of the results of representative studies is provided in Table 1.

Comparisons between the cytotoxicities of those different nanomaterials are generally difficult to make due to the vast range of methods, concentrations, dimensions, cell lines etc. For instance, the concentrations reported in the different studies were typically evaluated using either ICP or Cryogenic TEM. However, the concentration or dose of the nanomaterial plays a significant role in the cytotoxic response as well as the biomedical applications. Similarly, the reported toxicology of the nanomaterials depends on their interaction with the assay. For example, carbon nanostructures interact with the MTT-formazan crystals but not with XTT or INT reagents.

While the concentrations and exposure times are critical factors, the toxicity of these nanostructures is also material-dependent. These relations can be seen in Figure 10, which presents the average values reported for the cell viabilities (ignoring differences in concentrations, incubation times etc.), when exposed to the nanomaterials in the studies covered in Table 1. ZnO NPs showed the highest toxicity, while the lowest has been reported for silicon.

In addition, the particle size plays a major role in the cytotoxic properties of the nanostructure, whereby both the cellular uptake efficiency and pathway are affected, with smaller particles being internalized faster than the larger ones.

The induction of ROS after dissolving the nanostructures in the lysosomes was shown to be the primary underlying cause of the toxicity in several cases, leading to cell death through the apoptotic pathway, due to ROS generation and mitochondrial damage. The acidic condition inside the lysosome increases the digestion of the particles, enhancing the release of ions that affect the viability of the cells. This is a particularly relevant issue in case of CdSe-core QDs, which release Cd ions upon oxidation, leading to fast cell death.

Adding a coating to the nanostructure typically affected both the toxicity and the surface charge of the nanostructure, where cationic surfaces are more toxic than anionic. For instance, the toxicity of QDs was reduced by adding a BSA corona, and the release of Cd was prevented by the addition of polymeric and inorganic coatings. The type of the coatings was shown to affect the cell viability differently.

The cytotoxicity of the nanomaterial depends also on the nanostructure's shape. In this regard, several advantages have been reported for NWs over NPs. For instance, they enhance the drugloading capacity due to their large surface area. An interesting observation from Figure 10 is that NWs/NRs are, on average, less cytotoxic than NPs, with titanium dioxide being the only exception. However, one study has shown that the large surface area of Si NWs has a more toxic effect at lower concentrations compared to NPs. This was attributed to the increased interaction of the nanomaterial with the cells due to the large surface area.


Nanostructure

Surface coating

Nanostructure

Average size Cell line

 Cell viability

Viability test Reference

Propidium

[110]

iodide

staining

MTT assay MTT assay

 [132]

 [124]

concentration

10 mM

13 nm

Human T

40%

lymphocytes

diameter

type

ZnO NPs SiO2 NPs

Si NPs Si NPs Si NPs Si NPs Si NPs Si NWs Si NWs Si NW arrays Si NW arrays Si NW arrays

 Coated with Cu NPs

—

5 μm long,

HeLa cells

50%

MTT assay

 [156] 251

20–100 nm

diameter

 Coated with AgNPs

—

5 μm long,

HeLa cells

 80%

MTT assay

 [156]

http://dx.doi.org/10.5772/intechopen.78616

20–100 nm

diameter

—

 —

5 μm long,

HeLa cells

 98%

MTT assay

 [155]

Review of In Vitro Toxicity of Nanoparticles and Nanorods—Part 2

20–100 nm

diameter

—

1 μg/ml

500 nm long,

breast cancer

90%

MTT assay

 [154]

> cells line (MCF-

100 nm diameter

7/ADR)

—

—

—

—

160 μg/ml

160 μg/ml

160 μg/ml <190 μg/ml

 7 nm

 20 nm

 50 nm

 2 μm long,

55 nm diameter

 HepG2 cells

HeLa and Hep-2

75%

cells

 HepG2 cells

 HepG2 cells

98% 72% 49%

MTT assay MTT assay MTT assay MTT assay

 [152]

 [156]

 [153]

 [136]

 Coated with positively

0–100 mg/ml

 3.9 nm

 Rat alveolar

The EC50 values = 0.38 μg/ml

 MTT assay

 [129]

macrophage

NR8383 cells

charged (NH2)

 Coated with negatively

3 mg/ml

1.6 nm

 Rat alveolar

No cytotoxicity

macrophage

NR8383 cells

charged (COOH)

—

50 μg/mL

 10–20 nm

 monocytes

71%

> (THP-1) cells

—


Nanostructure

Surface coating

Nanostructure

Average size Cell line

 Cell viability

Viability test Reference

250 Cytotoxicity

Tryptan

[35]

blue/

microscopy,

MPP, GSH

Crystal

[44]

violet/optical

density

concentration

0.125 mM/1 h

 N/A

 rat hepatocytes

 80% for pristine, C60(OH)12 and 60%

for C60(OH)24.

type

C60 C60

C60-alanine,

NO2-proline,

polycarboxylic

 derivative

 sodium salt of a

 -NO2, -PVP, -

0.001–0.2 mg/mL

N/A

 HEp-2 cells

 No cytotoxicity

salt of a with 20% viability at 0.01 mg/mL

 except for the sodium

polycarboxylic

 derivative

for C60NO2-proline

0.016–0.2 mg/mL for all others/48 h

 0.03–1 mg/mL/24 h N/A

 HepG2, NHDF,

Maximum inhibition at 1 mg/ ml of

MTT, LDH

[45]

assays

Full–PEG2000

(62%).

 for J774 (41%) and U937

Caco2, HUVEC,

U931, J774 A1

C60 SWCNTs

SWCNTs

SWCNTs

MWCNTs

MWCNTs

TiO2 NPs

—

600 μg/mL

 5 nm

L929 mouse

<70%

MTT assay

 [107]

diameter

fibroblast cells

 3 and 23% of Fe impurities

 5–60 and 72 h

μg/mL/24, 48

2–50 nm

PC12 cells

 70 and 20% viability at highest

CCK-8

 [73]

concentration

respectively

 and time exposure,

diameter,

50 μm length

 Pristine, COOH

12.5–200 μg/mL/

10–20 nm

human normal

60 and 80% viability at highest

concentration

respectively

 and time exposure,

diameter, 10–

liver cell line L02

> 30 μm length

24, 48 and 72 h

 Gd-NPs as catalysts and PEG 50–100

 Collagen

15 15 days.

μg/mL/4 h to

0.7–1.6 nm

BACs

No cytotoxicity

> diameter, N/

A

μg/mL/12–

N/A

 NIH/3 T3

70% viability at highest

and time exposure

concentration

Tryptan

[55]

blue/

microscopy,

Live/dead

CellTiter-GloV® assay

[68]

fibroblasts

48 h

 Pristine and PEG

0.1–100 μg/mL/

0.7–1.6 nm

PC12 cells

 30 and 50% viability in MTT at highest

concentration,

values for XTT and 10–20% LDH

leakage

respectively.

 Higher

MTT, XTT,

[58]

LDH, DCF, GSH assays WST-1 assay,

[59]

Live/dead

diameter,

0.2–3 μm

length

24 h

PEG of various sizes

 and

Pristine, C60(OH)12,

C60(OH)24,

Review of In Vitro Toxicity of Nanoparticles and Nanorods—Part 2 http://dx.doi.org/10.5772/intechopen.78616 251


Table 1. Summary of in-vitro cytotoxicity studies with different kinds of nanoparticles (NPs) and nanowires (NWs), NWs with aspect ratio <10 are often called nanorods (NR), SWCNT the abbreviation of single-walled carbon nanotube, MWCNT for multiwalled carbon nanotube and QD for quantum dots.

While all these studies contributed to obtain a better picture of the cytotoxicity of nanomaterials and the underlying mechanisms, it is a persisting issue that a consistent measurement and reporting system will be needed for future studies. This will not only enable performing more accurate comparisons of the toxicological characteristics of nanostructures, but also to better

Figure 10. Average cell viability when exposed to nanomaterials reported in Table 1, considering nanoparticles (NP),

, single-walled carbon nanotubes (SWCNT), multiwalled carbon

Review of In Vitro Toxicity of Nanoparticles and Nanorods—Part 2

http://dx.doi.org/10.5772/intechopen.78616

253

)

Research reported in this publication was supported by the King Abdullah University of

<sup>1</sup> and Jürgen Kosel

1 Division of Biological and Environmental Sciences and Engineering, King Abdullah

2 Division of Computer, Electrical and Mathematical Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal, Kingdom of Saudi Arabia

2 \* 1 ,

Jose E. Perez1,2, Nouf Alsharif1,2, Aldo I. Martínez-Banderas1,2, Basmah Othman

University of Science and Technology, Thuwal, Kingdom of Saudi Arabia

evaluate the potential of using them for biomedical applications.

Policy: https://www.intechopen.com/authorship-policy.html.

, Timothy Ravasi

\*Address all correspondence to: jurgen.kosel@kaust.edu.sa

Acknowledgements

Author details

Jasmeen Merzaban

Science and Technology (KAUST).

nanorods (NR), nanowires (NW), Fullerene (C60

nanotubes (MWCNT) and Quantum Dots (QDs).

1

Figure 10. Average cell viability when exposed to nanomaterials reported in Table 1, considering nanoparticles (NP), nanorods (NR), nanowires (NW), Fullerene (C60), single-walled carbon nanotubes (SWCNT), multiwalled carbon nanotubes (MWCNT) and Quantum Dots (QDs).

While all these studies contributed to obtain a better picture of the cytotoxicity of nanomaterials and the underlying mechanisms, it is a persisting issue that a consistent measurement and reporting system will be needed for future studies. This will not only enable performing more accurate comparisons of the toxicological characteristics of nanostructures, but also to better evaluate the potential of using them for biomedical applications.
