Oxidative stress-induced neurotoxicity of quantum dots and influencing factors

Abstract

Quantum dots (QDs) have significant potential for treating and diagnosing CNS diseases. Meanwhile, the neurotoxicity of QDs has garnered attention. In this review, we focus on elucidating the mechanisms and consequences of CNS oxidative stress induced by QDs. First, we discussed the pathway of QDs transit into the brain. We then elucidate the relationship between QDs and oxidative stress from in vivo and in vitro studies. Furthermore, the main reasons and adverse outcomes of QDs leading to oxidative stress are discussed. In addition, the primary factors that may affect the neurotoxicity of QDs are analyzed. Finally, we propose potential strategies for mitigating QDs neurotoxicity and outline future perspectives for their development.

Graphical abstract

Plain language summary

Executive summary.

Pathways of quantum dots into the brain

• The blood–brain barrier is one of the main routes for quantum dots (QDs) to enter the CNS, and QDs with a particle size of only a few nanometers can easily cross the blood–brain barrier and enter brain tissue in a short time.

• QDs can also be transported to the brain via the olfactory nerve by inhalation.

QDs induce neurotoxicity through oxidative stress

• Cadmium-based QDs induce changes in oxidative stress levels and neurobehavior in experimental animals or model organisms. In in vitro studies, cadmium-based QDs produced abnormal increases in reactive oxygen species, microglial activation and neuronal damage.

• Noncadmium-based QDs also induce changes in the level of oxidative stress, but this change is twofold, with some QDs exhibiting toxicity and others exhibiting protective effects.

Mechanisms of oxidative damage to the CNS induced by QDs

• QDs entering the CNS cause an abnormal increase in ROS and exceed the body's ability to compensate, resulting in oxidative stress.

• QDs can lead to altered activity of antioxidant enzymes and increased consumption of antioxidants, ultimately causing oxidative damage.

• Accumulation of ROS and dysfunction of the antioxidant system ultimately cause damage to biomolecules, including lipid peroxidation, protein and DNA damage.

Factors influencing the neurotoxicity of QDs

• The chemical composition, particle size, surface functionalization, surface charge and protein corona of QDs affect the neurotoxicity of QDs to varying degrees. Modifying QDs to address these factors will help mitigate the neurotoxicity of QDs.

The development of nanotechnology has led to the widespread application of nanomaterials across several disciplines. Quantum dots (QDs) refer to nanocrystals made up of semiconductor materials from either group III–V or group II–VI elements. These nanocrystals possess a core or shell structure and have a size less than 10 nm in 3D space [1]. QDs exhibit a wide range of excitation wavelengths and a small range of emission wavelengths. By manipulating their size and chemical composition, it is possible to modulate the emission spectrum of QDs, enabling coverage of the whole visible area [2,3]. As an illustration, it can be observed that the particle size of cadmium telluride (CdTe) QDs undergo a rise from 2.5 to 4.0 nm, resulting in a corresponding shift in its emission wavelength from 510 to 660 nm [4]. QDs have demonstrated potential use in the biomedical area, namely for diagnostic and therapeutic purposes. In addition, QDs have been explored for their utility in solar cells, catalytic reactions and display screens (Figure 1) [2,5]. The properties of QDs offer promising prospects in the medical field, particularly in the diagnosis and treatment of CNS diseases. Neurological diseases such as glioblastoma are poorly treated by conventional diagnostic methods, and intraoperative tumor visualization using QDs will significantly improve the degree of glioma resection and patient prognosis [6].

Figure 1.

Luminescent properties and potential applications of quantum dots.

(A) Emission bands of quantum dots (QDs) and fluorescent colors of QDs under ultraviolet light. (B) Cancer incidence and mortality rate globally in 2018. (C) Illustration of receptor-mediated endocytosis of targeting ligand-conjugated graphene QDs loaded with an anticancer drug into a tumor cell and drug release inside the cell. (D) Fluorescent micrographs of three cell lines treated with the carbon quantum dots (CQDs). CQDs confined by double-stranded DNA emit green fluorescence, and CQDs clustered by single-stranded RNA emit red fluorescence. Images were taken under a confocal microscope with an excitation of 488 nm and emission window of 500–560 nm. (E) In vivo fluorescence imaging of nude mice after intravenous injection of CQD-WS solution.

GQD: Graphene quantum dot.

(A) Reproduced with permission from [7]; (B) reproduced with permission from [8]; (C) reproduced with permission from [9]; (D) reproduced with permission from [10]; (E) reproduced from [11] under a CC-BY license.

However, the neurotoxicity of QDs is a key issue that needs to be addressed before they can be used clinically. QDs can enter the brain by several routes, including blood–brain barrier (BBB) and trans-olfactory nerve transport [12,13]. QDs may have a range of adverse consequences following their entry into the CNS. The main neurotoxic effects induced by QDs are oxidative stress and inflammation [14,15]. Oxidative stress is a phenomenon caused by an imbalance between the production and elimination of reactive oxygen species (ROS) in organisms [16]. The body itself has a mechanism for scavenging excess ROS, which is the antioxidant system. However, certain exogenous factors, such as QDs, not only promote the generation of ROS, but also inhibit or even destroy the antioxidant system and aggravate oxidative stress [17,18]. Oxidative stress and overproduction of ROS are considered to be the major explanations for neurodegenerative diseases [19].

QDs can cause oxidative stress in the CNS through different pathways. Microglia can be activated into M1 (proinflammatory) or M2 (anti-inflammatory) types after ingestion of QDs [20]. M1 microglia release a large number of inflammatory factors and ROS under the action of QDs and attack the surrounding nerve cells. In addition to exogenous ROS, QDs also lead to the production of endogenous ROS upon accumulation in neurons [21]. This mainly involves damage to mitochondria by QDs, and when it cannot be repaired in time by the cell's repair mechanisms, ROS within the mitochondria is released into the matrix and attacks intracellular biomolecules such as lipids, proteins and nucleic acids. On in vivo studies, high doses of QDs exhibit behavioral changes in addition to causing abnormalities in physiological and biochemical indices [22]. Cadmium-based QDs differed from noncadmium-based QDs in their toxic effects. The release of cadmium ions significantly aggravated oxidative stress and led to more severe damage [23]. The interaction between QDs and living organisms is so complex that it is more difficult to elucidate their neurotoxicity mechanisms. Therefore, a review of the neurotoxicity of QDs, especially the mechanism of oxidative stress, is necessary.

Therefore, although the excellent properties of QDs can play an important role in the diagnostic and therapeutic process of diseases, the potential toxicity has become a major factor hindering their clinical application. In particular, oxidative stress, as the main toxic effect caused by QDs in the CNS, needs to be reviewed to provide reference for further research. In general, QDs are able to transit into the brain and induce neurotoxicity through the BBB, or olfactory nerves. Among them, the release of cadmium ions from cadmium-based QDs exacerbates oxidative stress and oxidative damage. Noncadmium-based QDs have relatively low toxicity and, in certain circumstances, perform antioxidant enzyme-like activity. QD-induced oxidative stress mainly involves a pathological increase in ROS and an imbalance in the antioxidant system. Ultimately, it causes lipid, protein and DNA damage at the molecular level, manifests as necrosis or apoptosis at the cellular level, and causes neurobehavioral changes at the whole animal level.

Pathways of QDs into the brain

Transport across the BBB

Unlike other systems, the nervous system has a unique defense mechanism against foreign chemicals, namely the BBB. The BBB was first discovered in 1885. It is a fundamental barrier structure that separates the brain from the general circulation, protects it from exogenous substances and maintains brain homeostasis [24,25]. It comprises different cell types, such as brain capillary endothelial cells, astrocytes, pericytes and nerve cells, which are tightly connected, further preventing the penetration of foreign substances. Thus, the BBB is a highly selective semipermeable membrane that allows only small, low-molecular-weight, nonpolar compounds to enter the brain [26].

The BBB strictly controls the exchange of substances between the blood and the brain. While protecting the brain from harmful substances, it also prevents most therapeutic drugs from entering the brain. Subsequently, researchers found that nanomaterials can enter the brain directly through the gap between the BBB due to their small particle size. Many studies have confirmed that QDs can cross the BBB in various ways, such as direct penetration, internalization by endothelial cells and receptor-mediated transport (Figure 2A). The latest research shows that multiemission fluorescent carbon QDs can cross the BBB and perform real-time brain imaging, which was initially confirmed by researchers using Kunming mice and zebrafish [27]. In vitro studies showed that QDs were easily internalized by brain microvascular endothelial cells after coupling with fungal metalloproteinases and did not destroy the integrity of the BBB [28]. The constructed QDs nanoprobes can cross the BBB, specifically bind to EGFRvIII, which is primarily distributed on the surface of glioma cells, and produce strong fluorescence, which is helpful to visualize the edge of glioma [29]. In addition, it can also be used as a pH-triggered targeted fluorescent probe for the detection of cerebral ischemic diseases [30]. In vivo experiments showed that CdTe/zinc sulfide (ZnS) QDs could be distributed in various tissues and organs after tail vein injection, including brain tissue [31]. At 3 h after exposure, the cadmium content in brain tissue reached its maximum value, and at 12 h after exposure, the cadmium content in brain tissue decreased significantly, indicating that QDs may be excreted through the kidney (Figure 2B).

Figure 2.

The pathway of quantum dots through the blood–brain barrier into the brain and their distribution and excretion in mice.

(A) Components of the blood–brain barrier and modes of drug transport across the membrane. (B) The distribution of the CdTe/ZnS quantum dots in mice (n = 3), quantum dots detected in brain tissue and excreted over time.

AA: Amino acid; BCEC: Brain capillary endothelial cell.

(A) ‘International Journal of Nanomedicine 2019 14 5895-5909’ Originally published by and used with permission from Dove Medical Press Ltd [24]; (B) reproduced with permission from [20].

Translocation via the olfactory nerve

QDs are transported through the BBB after entering the bloodstream and can also enter the brain from the olfactory nerve through the respiratory tract. After inhalation or instillation of nanoparticles (NPs), they enter the respiratory tract; some attach to the olfactory epithelium, are internalized by cells and are transported through axons into the olfactory bulb to the brain tissue. In 2006, it was found that olfactory neuronal pathways could efficiently transport inhaled Mn₂O NPs (30 nm) to the CNS [32]. After continuous inhalation of aerosolized Ag-SiO₂ NPs at a concentration of 1 mg/ml for 6 h in adult Sprague–Dawley rats, NPs elicited transient and differential microglial activation in the olfactory bulb without significant microglial recruitment or oxidative stress, suggesting that NPs may enter the CNS via an intraneural pathway [33]. It is worth mentioning that the surface charge of NPs also affects the penetration of NPs into the brain through the olfactory nerve. Localization of fluorescently labeled NPs in different brain regions was detected after intranasal administration; the positive charge of NPs slowed the arrival of NPs to brain tissue, and the trigeminal pathway was also involved [34]. Other NPs, including QDs and soluble metals, have also been shown to be selectively transferred to the brain via the olfactory bulb [35,36]. Adult C57BL/6 mice inhaled a cadmium selenide (CdSe)/ZnS QDs aerosol for 1 h; the presence of QDs was detected in the respiratory tract and olfactory bulb, and the activation of microglia in the olfactory bulb was detected 3 h later [36]. It is the first study to demonstrate that QDs can be taken up by the olfactory nerve after inhalation and transported rapidly from the nose to the brain by axonal transport.

In addition to olfactory nerve transport, QDs may be transported by sensory nerve endings emitted by the trigeminal nerve [37,38]. However, unfortunately, there are few reports about the trans-sensory nerve terminal transport of QDs. More attention has been paid to the BBB as QDs directly cross the BBB and enter the CNS in a short period of time, and it is also one of the main pathways for the clinical application of QDs in the future.

QDs induce neurotoxicity through oxidative stress

Undoubtedly, most QDs will cause oxidative stress as an early toxic effect after entering the CNS (Table 1). Therefore, the study of the oxidative stress mechanism of QDs is conducive to clarifying the toxic effect mechanism of QDs but can also take oxidative stress as a breakthrough point to study how to prevent the neurotoxicity of QDs. Notably, there are differences in the neurotoxic effects produced by different types of QDs. Cadmium-based QDs have relatively higher fluorescence performance and toxicity than noncadmium-based QDs, and the heavy metal cadmium may cause further damage to the nervous system. Noncadmium-based QDs, such as graphene oxide QDs (GOQDs) may exhibit different effects (toxic and protective) under different conditions.

Table 1.

Summary of neurotoxicity and oxidative stress of quantum dots.

Quantum dots	Size (nm)	Dose	Exposure time	Result	Ref.
CdS	4–12	0.01–100 μg/ml	24 h	The total antioxidant capacity of neurons increased following exposure to the lowest concentrations of CdS, and total oxidant status was decreased following exposure to lower concentrations of CdS	[39]
CdSe/ZnS	6	2 mg/l	72 h	Causes severe neurological and developmental problems	[40]
GQDs	3.4	0–500 μg/ml	24 h	Calcium dysregulation, DNA damage, and cell cycle arrest are key factors leading to the induction of neurotoxicity in GQDs	[41]
CQDs	2.45	5 mg/kg	0–10 h	Functionalized carbon quantum dots selectively accumulate in human tumors in mice to promote targeted therapy	[42]
GQDs GOQDs	2–4	150–2400 g/l 60–960 g/l	7 days	GQDs affect synaptic plasticity by downregulating mRNA levels of NMDA and AMPA receptor family members as well as total glutamine levels in zebrafish larvae	[43]
InP/ZnS	4.5	200, 800 nM	48, 72, 96 hpf	Quantum dots cause developmental toxicity and lead to increased levels of SOD and MDA	[44]
e-CDs	4.8	0–300 μg/ml 0–300 mg/kg	24 h, 30 days	In vivo and in vitro studies show that QDs inhibit oxidative damage through enzyme-like activity	[45]
BPQDs	NM	2.5, 5, 10 mg/l	24, 48, 72, 96 hpf	Induces developmental toxicity, oxidative stress, DNA damage, apoptosis and motor behavioral deficits in early zebrafish development	[46]
Se QDs	5	20 μg/ml 1 mg/kg	24 h, 48 h, 4 weeks	In vitro, Se QDs has neuroprotective effects such as maintaining the intracellular ROS level and improving mitochondrial dysfunctions; in vivo, Se QDs improves the learning and memory ability of AD mice	[47]
MoS2 TPP-MoS2	30 50	10 mg/ml	15 days	TPP-MoS2 QDs stimulated microglial polarization from the inflammatory M1 phenotype to the anti-inflammatory M2 phenotype and demonstrated CAT and SOD enzyme-like activity	[48]
CdTe	3.47	0–10 mg/kg bw	2 h, 24 h, 3 days, 1 weeks	At high doses, exposure to CdTe-QDs resulted in mild dehydration, lethargy, ruffled fur, hunched posture and body weight loss	[49]
InP/ZnS	3–7	2.5, 25 mg/kg bw	1, 3, 7, 14, 28 days	No obvious histopathological abnormalities were observed in the brain	[50]

BPQD: Black phosphorus quantum dot; bw: Body weight; CAT: Catalase; CdS: Cadmium sulfide; CdSe: Cadmium selenide; CdTe: Cadmium telluride; CQD: Carbon quantum dot; e-CD: Enzyme-like activity of carbon dot; GQD: Graphene quantum dot; GOQD: Graphene oxide quantum dot; hpf: Hours post fertilization; MDA: Malondialdehyde; NM: Not mentioned; Se: Selenium; SOD: Superoxide dismutase; ZnS: Zinc sulfide.

Cadmium-based QDs

Cadmium-based QDs mainly include CdTe, CdSe and cadmium sulfide (CdS). On this basis, there are different modified QDs. The significant difference between cadmium-based QDs and other QDs is their heavy metal core, which gives them excellent fluorescence performance but also brings some safety risks. The cytotoxicity of CdTe QDs and CdCl₂ at the same dose was compared, and the results showed that CdTe QDs had a more substantial inhibitory effect on cells, indicating that cadmium-based QDs may be more toxic due to the nano-effect [51,52]. In 2009, it was pointed out that the oxidative damage of CdS QDs was mainly caused by ROS at low concentrations, by both ROS and Cd²⁺ at medium concentrations, and by Cd²⁺ at high doses [23]. Interestingly, the fluorescence intensity of CdTe QDs decreased while the Cd²⁺ concentration increased as the pH value decreased [53]. This implies that CdTe QDs may accelerate degradation in the acidic environment of intracellular lysosomes. In addition, Cd²⁺ release is time dependent, and its intracellular concentration will reach a dynamic equilibrium over time. When there is enough Cd²⁺ chelator, the dynamic equilibrium will be destroyed and CdTe QDs will be degraded entirely [54].

The in vivo neurotoxicity study of cadmium-based QDs found that exposure to QDs could trigger behavioral changes, which may be caused by oxidative stress. After MPA-CdTe QDs were injected into the hippocampus of rats, the learning efficiency and spatial memory were impaired using the open field test and Y-maze test, which may be due to the pathological changes and ultrastructural damage of hippocampal neurons and synapses [22]. Further studies showed that CdTe QDs caused cell death and apoptosis in primary cultured rat hippocampal neurons in a dose- and time-dependent manner, and this process was accompanied by an increase in intracellular ROS and calcium levels [55]. Buffet et al. compared CdS QDs at the same concentration (10 μg/l) with the toxicity of soluble cadmium in clams, and the results indicated that CdS QDs caused behavioral damage in clams accompanied by significant increases in catalase (CAT) and glutathione-S-transferase [56]. These results suggested that QDs induced oxidative stress, and the unique nano-effect caused additional damage.

Exposure of zebrafish eggs to CdSe/ZnS QDs for 5 days resulted in elevated chromium content in larvae, impaired locomotor behavior and an increased abundance of mRNA expression associated with oxidative stress [57]. The findings suggest that exposure to QDs may induce alterations in the oxidative stress levels of zebrafish larvae, subsequently impacting their nervous system and resulting in changes in motor behavior. Both 3-mercaptopropionic acid (MPA)-CdTe QDs and MPA-CdS-CdTe QDs induced a dose-dependent delayed hatching effect in zebrafish and subsequently induced transporter proteins such as mrp 1 and mrp 2 in zebrafish embryos, in addition to elevated oxidative stress, which may also be regulated by signals such as oxidative stress [58]. CdSe QDs also induced neurotoxicity in zebrafish, resulting in increased expression of antioxidant enzymes, excessive ROS and activation of apoptosis through the caspase-3-mediated signaling pathway [59]. On the one hand, cadmium-based QDs themselves can promote the generation of ROS and change the level of oxidative stress, and on the other hand, Cd²⁺ released from QDs may also aggravate this phenomenon [60].

Numerous in vitro studies have found that cadmium-based QDs can cause an abnormal increase in ROS and lead to oxidative stress, which is further confirmed by in vivo studies that the toxic effect of cadmium-based QDs is highly related to oxidative stress. The higher the concentration of Cd in BV2 cells after QDs exposure, the higher the degree of cell damage [51]. After CdTe/ZnS QDs treatment, microglia were activated to the M1 phenotype, and glycolysis was changed from oxidative phosphorylation to aerobic glycolysis through the mTOR signaling pathway [61]. M1 microglial cells are proinflammatory cells, and neurotoxic molecules such as free radicals and inflammatory factors are expressed and released in large quantities in the cells, leading to neuronal damage. In a nonhypoxic environment, QDs increased the glycolysis level of microglia by 86% and inhibited the aerobic respiration level by up to 54% [31]. In addition, cadmium-based QD exposure activated NF-κB, which was involved in the initiation of NLRP3 inflammasome and pro-IL-1β expression. After that, excessive ROS production induced by QDs triggered activation of the NLRP3 inflammasome and resulted in the processing of pro-Il-1β by active caspase-1 to mature IL-1β release and inflammatory cell death, namely pyroptosis [62].

Noncadmium-based QDs

Although noncadmium-based QDs are not as good as cadmium-containing QDS in imaging, they have lower toxicity, so they have also received extensive attention from the industry. By comparing the toxicity of several QDs through acute toxicity tests, detection of growth inhibition, enzyme activity and metabolites, the toxicity of several QDs from small to large was obtained as follows: CuInS₂/ZnS QDs < no doped carbon QDs (CQDs) < N doped CQDs < N, S doped CQDs < CdS QDs < CdTe QDs [63]. Similar to cadmium-based QDs, noncadmium-based QDs generally also cause an abnormal increase in ROS and lead to oxidative stress. Using Caenorhabditis elegans as a model to evaluate the neurotoxicity of Ag₂Se QDs, it was found that such QDs could accumulate in the nematode in a dose- and time-dependent manner, causing an increase in ROS and resulting in a shortened lifespan and impaired neurobehavior of the nematode, significantly when the exposure concentration exceeded 0.1 μM [64]. Pericardial edema and tail bending were observed in Chinese rare carp embryos after exposure to CuInS₂/ZnS QDs. Further studies showed that SOD and malondialdehyde levels were significantly increased, and DNA damage was induced [65]. Graphene QD (GQD) exposure caused a change in the motor behavior of C. elegans, which was manifested as a significant reduction in the movement frequency of body bending, head swing and pharyngeal suction. In addition, the average speed, bending angle frequency and wavelength of crawling movements were significantly reduced after exposure. This suggests that there could be damage to the neurons [66].

Some QDs showed a completely different effect from others, that is, a positive effect. Different from general nanomaterials, biocompatible GOQDs can reduce oxidative stress and inhibit neurotoxicity [67]. GOQDs can play a role similar to CAT after entering the zebrafish brain, thereby improving neurotoxicity. GQDs were also used to mitigate neurotoxicity by eliminating hydroxyl radicals and nitric oxide and inducing cytoprotective autophagy [68]. Carbon dots also showed the opposite effect, showing a greater scavenging capacity for free radicals and can protect cells from oxidative damage [69,70]. In addition, CQDs extended the lifespan of paraquat-impaired C. elegans and herbicide-mediated ablation of dopamine neurons [71]. However, it should be noted that these protective effects appear to be effective only under certain conditions and are still neurotoxic in some cases [72]. In general, the protective mechanism of these QDs is often through the intervention of oxidative stress, inhibiting the generation of free radicals in the cells and thereby reducing oxidative damage. Although the protective effect of these QDs conflicts with the neurotoxic effects of nanomaterials described above, there seem to be some mechanisms that we have not yet explored that are worth further exploration by our researchers.

Mechanisms of oxidative damage to the CNS induced by QDs

The neurotoxicity of QDs is dose dependent, with cellular damage progressively worsening with increasing doses and even causing neuronal damage. At the molecular level, QDs promote the generation of ROS; at the cellular level, QDs cause damage to biomolecules and apoptosis; at the organ level, QDs may eventually cause neurodegenerative diseases (Figure 3). These potential safety hazards severely limit the clinical application of QDs and are a challenge that needs to be urgently addressed.

Figure 3.

Mechanism of quantum dots causing neurotoxicity by inducing oxidative stress.

(A & B) Formation of reactive oxygen species and reactive nitrogen species in the mitochondrial matrix and their adverse consequences. (C) Schematic representation of toxicity induced by quantum dots in live cell. (D) The adverse outcome of oxidative stress in the central nervous system.

MT: Metallothionein; QD: Quantum dot; ROS: Reactive oxygen species.

(A & B) Reproduced from [73] under a CC-BY license; (C) reproduced with permission from [5]; (D) reproduced from [74] under a CC-BY license.

ROS accumulation

ROS are the by-products of cellular aerobic respiration, which play an essential role in the normal life cycle of cells as cell signal transduction molecules at low doses [75]. However, ROS is an indispensable link in life activities and one of the leading causes of damage. Under the influence of endogenous or exogenous stimuli, it can induce a rapid elevation in ROS levels and the excessive accumulation of ROS cannot be efficiently eliminated in a timely manner, resulting in the generation of oxidative stress (Figure 3A & B). ROS is not a single substance but a group of substances, including superoxide (O₂·-), hydrogen peroxide (H₂O₂), hydroxyl radical (·OH) and singlet oxygen (¹O₂) [76,77]. Excessive ROS production can cause irreversible damage to biofilms, nucleic acids and proteins and even cause neurodegenerative diseases such as Parkinson's disease and Alzheimer's disease.

ROS is frequently observed as the instigator of a cascade of adverse effects caused by QDs. Numerous studies have consistently demonstrated that introducing a specific dosage of QDs into the CNS typically triggers excessive production of ROS, subsequently leading to a series of detrimental events. A significant increase in ROS levels was detected in model organisms such as C. elegans and in cultured cells such as microglia after QDs exposure [14,51,78]. Intracellular ROS is mainly produced by mitochondria, with a small amount also generated in other organelles. The mitochondrial respiratory chain is an essential source of ROS production. Indo et al. found that more ROS were induced in cells with mitochondrial respiratory chain inhibition and mtDNA damage [79]. ROS production was found to be associated with Akt, p38 and JNK signaling pathways. Resveratrol, H₂S and 43°C hyperthermia reduced ROS production and cell apoptosis after inhibiting the related signaling pathways, respectively [80]. QDs could not only induce the increase of endogenous ROS but also induce the production of exogenous ROS. For example, copper-doped zinc oxide QDs and GO QDs can produce ROS under light, including O₂·-, ·OH and ¹O₂ [81,82]. Different types of ROS may play different roles in the process of QD oxidative damage. However, there are few studies on this aspect, and further exploration is still needed. Although the body's antioxidant system can remove ROS to a certain extent, excessive ROS exceeds the antioxidant capacity of the antioxidant system, leading to the disorder of the antioxidant system.

Dysfunction of the antioxidant system

The body's antioxidant system is critical to the life activities of cells. It can effectively remove excessive free radicals in the body and avoid the damage of free radicals to the cell membrane and biological macromolecules. The exposure to QDs will inhibit the antioxidant system and have adverse effects. The brain antioxidant system consists of antioxidant enzymes (e.g., SOD and CAT) and nonenzymes (e.g., GSH and ascorbic acid). The destruction of the antioxidant system of QDs will lead to oxidative stress. In molecular experiments, MPA-CdTe QDs significantly changed the molecular conformation of SOD and CAT, increasing CAT activity and inhibiting SOD activity. It also induces oxidative stress and apoptosis at the cellular level [83]. Cd QDs were found to bind with antioxidant enzymes through hydrophobic forces, leading to static quenching of the protein's intrinsic fluorescence and the formation of a new complex. At the same time, the skeleton and secondary structure (reduced α-helix) of the two antioxidant enzymes were also affected [84,85]. In addition, it was found that CdTe QDs could also cause GSH depletion and reduce the scavenging ability of the liver and kidney to ROS in a time- and dose-dependent manner, thereby inducing oxidative damage in tissues [86]. N-GQDs can strongly interfere with the redox-sensitive system by selectively inhibiting the activity of endogenous antioxidant enzymes in zebrafish. This may be due to competitive inhibition of the electron transfer process to destroy antioxidant enzyme activity [18]. GOQDs could also cause oxidative stress by affecting the antioxidant system, which was manifested as an increase in the activity of SOD, an increase in the activity of CAT, and a dose-dependent increase in the production of GSH and the degree of lipid peroxidation [87]. It is worth noting that with aging, the antioxidant capacity of the body decreases, and it may be more sensitive to exposure to QDs [88]. Therefore, the exposure of QDs to the elderly or the possible clinical medical application should be kept more cautious. As an oxidative defense mechanism of the body, the antioxidant system is dysregulated under the influence of QDs, and excessive ROS damages the cell membrane and biological macromolecules, eventually causing cell apoptosis (Figure 3C).

Lipid peroxidation, protein & DNA damage

QDs have been found to have detrimental effects on biofilms and biological macromolecules, resulting in lipid peroxidation, protein degradation and DNA damage. These adverse effects ultimately impact the CNS, contributing to the development of many disorders [73]. Lipid peroxidation is a free radical chain reaction that spreads rapidly and affects many lipid molecules, for example, excess hydroxyl radicals and peroxynitrite cause lipid peroxidation and destruction of cell membranes and lipoproteins [89]. The increase in lipid peroxidation level was closely related to the exposure to QDs [90,91]. After NRK cells were treated with QDs, the contents of ROS and MDA were significantly increased, and the activities of CAT, SOD and GSH-PX were decreased. Subsequent in vivo studies showed that the activation of the Nrf2-Keap1 signaling pathway was inhibited [92]. The Nrf2-Keap1 signaling pathway is crucial for organisms to resist oxidative stress. Keap1 inhibits Nrf2 activity under resting conditions, while Nrf2 is released from Keap1-mediated inhibition during stress and transacts chemical signals to regulate a series of cytoprotective genes [93]. The inhibition of this pathway affects the body's antioxidant defense system and aggravates cell damage.

DNA damage is also one of the adverse consequences of QDs. Both cadmium-based QDs and noncadmium-based QDs induced DNA damage effects [94–96]. However, DNA cannot interact with QDs, so DNA damage is not caused by the adduct of CdTe QDs with DNA but by increased Cd ion concentration and secondary oxidative damage [51,97]. In addition to DNA damage, biofilm damage and depletion of endogenous reduced glutathione were evident in QD-exposed BV2 cells [51]. After QDs could not compensate for the damage to biological macromolecules, they further caused damage to organelles. Mitochondrial dysfunction and oxidative stress due to excessive production of ROS are standard features of neurodegenerative diseases such as Parkinson's disease and Alzheimer's disease (Figure 3D) [98]. Fortunately, the chance of exposure to highly toxic QDs in our daily lives is slight, with the possibility of occupational exposure and short-term exposure in future clinical applications.

Factors influencing the neurotoxicity of QDs

QDs are a type of nanomaterial, and as such, their potential toxicity to the human body is quite different from that of conventional poisons, such as chemical composition, particle size, surface functionalization/coating, surface charge and protein corona. The effects of these factors on toxicity are reflected in various aspects, of which we are concerned mainly with oxidative stress.

The effect of particle size of QDs on Escherichia coli was compared by using E. coli as a research object, and the result was that the smaller the particle size, the greater the inhibitory effect on E. coli [99]. QDs, as a nanomaterial of only a few nanometers, can be distributed more efficiently across various barriers of living organisms to various tissues and organs throughout the body than other nanomaterials or conventional compounds. After exposure to graphite, graphite oxide nanosheets and GQDs, it was found that the first two were mainly distributed in the pharynx and intestine of C. elegans. In contrast, QDs were widely distributed in the body of C. elegans [66]. Differences in gene expression in the rat hippocampus treated with 2.2 and 3.5 nm CdTe QDs were found by whole-transcriptome sequencing, and 3.5 nm may cause more severe damage [100]. In vitro experiments demonstrated that 2.2 nm CdTe QDs induced higher ROS levels and lower cell viability (Figure 4A & B) [101]. QDs with smaller particle sizes produced higher levels of ROS, resulting in more severe effects at the cellular or overall animal level.

Figure 4.

Different properties of quantum dots have different effects on toxicity.

(A & B) Alterations in BV2 cell viability and ROS after exposure to 2.2 and 3.5 nm CdTe QDs. (C–H) Cell viabilities and intracellular ROS levels of P. chrysosporium and E. coli exposed to different concentrations of Cd²⁺, cadmium selenide (CdSe) and CdSe/zinc sulfide QDs for 24 h, respectively.

CdTe QD: Cadmium telluride quantum dot; ROS: Reactive oxygen species.

(A & B) Reproduced with permission from [101]; (C–H) reproduced with permission from [102].

Chemical composition has a significant effect on the toxicity of QDs, e.g., Ag₂Se QDs (2.8 nm) and CdTe QDs (2.2 nm) with similar particle size inhibited BV2 cells differently at a dose of 160 nM, with cell viability of the former at 40% and the latter dropping to less than 20% after 24 h of exposure [103]. It is worth noticing that the influence of the chemical composition of QDs on toxicity is not limited to different elements. However, the ratio of the elements may also be an essential influence. Increased selenium content in CdSe QDs reduces free cadmium ion levels and mitigates oxidative stress by affecting the antioxidant system [104].

Surface functionalization of QDs is the linking of QDs with suitable ligands so that the QDs can have different efficacy, such as targeting an organ or tissue. OH-GQDs are more potent in inducing histone modification imbalances and lead to a higher latency of impaired neural differentiation than NH₂-GQDs, suggesting a neurodevelopmental toxicity-specific effect of surface functionalization of GQDs [105]. The surface coatings of the QDs had different effects on cytotoxicity. The IC₅₀ values of CdTe QDs with different coatings were different: GSH-CdTe QDs (15.3 nM) < MPA-CdTe QDs (56.2 nM) < NAC-CdTe QDs (89.8 nM) [106]. It is noteworthy that all three QDs were primarily localized in the mitochondria upon cell entry and induced a marked increase in ROS. In addition, the study confirmed that the presence of a surface coating effectively reduced the release of heavy metal cadmium ions, thereby mitigating the toxic effects. The toxicity of QDs is mainly due to the increase in ROS, caused by their unique nanoproperties such as particle size, charge, specific surface area and shape. The release of heavy metal ions is typically a side effect of heavy metal-based QDs. It was shown that the magnitude of cytotoxicity (Figure 4C–H) induced by Cd²⁺, CdSe and CdSe/ZnS QDs was as follows: CdSe QDs > CdSe/ZnS QDs > Cd²⁺ [102]. The toxicity of CdTe QDs synthesized with 2-mercaptoacetic acid and MPA as a coating material was compared, and it was found that these two QDs can impair mitochondrial respiration and induce mitochondrial permeability transition [107]. Notably, the surface coating mitigates the toxicity of QDs and improves the fluorescence performance of QDs. Compared with CdTe QDs, CdTe/ZnS QDs are less toxic and more luminescent, with a quantum yield 1.7-times higher than bare-core QDs [108].

In addition to the above factors affecting the toxicity of QDs, other factors such as surface charge [109,110], protein corona [111] and exposure mode [112] affect toxicity to varying degrees. Moreover, the chirality of QDs could possibly exert a significant influence [113].

Conclusion

QDs have the potential to be transferred across the BBB or through the olfactory nerve, opening up new avenues for the diagnosis and treatment of CNS problems. Nevertheless, the safety risk associated with the potential neurotoxicity of QDs arises from their ability to generate ROS and induce oxidative stress, leading to various deleterious effects. In addition, the presence of cadmium ions aggravated oxidative damage. Through in vivo and in vitro studies, it was found that both cadmium-based QDs and noncadmium-based QDs affect ROS levels and induce oxidative stress. Furthermore, in vivo studies have found that QDs cause neurobehavioral changes, including reduced learning ability, impaired memory and altered motor behavior. In addition to inducing ROS generation, QDs also affect the antioxidant system, primarily in the inhibition or destruction of antioxidant enzymes. When excess ROS cannot be eliminated through the antioxidant system, they will attack biological membranes and various biomolecules, leading to neuronal damage or even apoptosis. It has also been found that some QDs exhibit the protective effect, and that such QDs alleviate oxidative stress by scavenging ROS through the exertion of enzyme-like activity. It involves a more complex mechanism and is one of the possible future directions for QDs.

Future perspective

In general, although QDs exhibit toxic effects through oxidative stress, it was found that the toxic effects of QDs can be controlled. It is specifically manifested in the improvement of the dose, particle size, chemical composition, surface charge, surface modification and functionalization of the QDs to reduce their toxicity. Recent studies have shown that combining QDs with appropriate ligands to target cancer cells results in ROS overproduction, activating a series of apoptotic signaling pathways to promote cancer cell apoptosis, and ultimately inhibiting or even curing cancer [114–118]. The toxic effects of QDs can be inhibited and regulated by the body's own mechanisms. For instance, the cellular autophagy mechanism can limit the toxic damage that QDs cause to some extent. After knocking down PINK1, the mitochondrial autophagy pathway mediated by PINK1/Parkin was inhibited, and the cellular damage was aggravated [119]. Therefore, QDs have immeasurable potential for biomedical applications. The development of QDs with robust fluorescence performance, excellent biocompatibility and reduced toxicity will be the predominant focus in the future. The discovery of QDs has unlocked new avenues in nanoscience, and the continued progress and application of QDs technology will undoubtedly propel societal advancements and bring about significant benefits to humankind.

Funding Statement

This work was supported by the National Natural Science Foundation of China (nos. 82173545, 21876026 and 31671034), the Natural Science Foundation of Jiangsu Province (no. BK20180371), the Postgraduate Research & Practice Innovation Program of Jiangsu Province (no. KYCX22_0300) and Fundamental Research Funds for the Central Universities (no. 2242023k30020). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Author contributions

Conception: Q Fang. Initial manuscript drafting: Q Fang. Review and final manuscript preparation: Q Fang and M Tang.

Financial disclosure

This work was supported by the National Natural Science Foundation of China (nos. 82173545, 21876026 and 31671034), the Natural Science Foundation of Jiangsu Province (no. BK20180371), the Postgraduate Research & Practice Innovation Program of Jiangsu Province (no. KYCX22_0300) and Fundamental Research Funds for the Central Universities (no. 2242023k30020). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Competing interests disclosure

The authors have no competing interests or relevant affiliations with any organization or entity with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Writing disclosure

No writing assistance was utilized in the production of this manuscript.