Synthetic Ultrafine Carbon Black Toxicity in Cell Free System and Animals
Date Issued
2005
Date
2005
Author(s)
Chen, Li-Chen
DOI
zh-TW
Abstract
Nanoparticles are increasingly used in science, technology and medicine, and they are produced for specific purposes which cannot be met by large particles and bulk material. However, recent evidences reveal that nanoparticles are likely to be highly reactive with biological systems. Currently environmental, healthy and safety regulation regarding to nanoparticles have not be well-established.
Recent studies indicated that artificial nanoparticals-induced health effects may be associated with the generation of reactive oxygen species (ROS). However, the exact relationship remains unclear. In order to investigate the possible mechanism, we used a cell-free system study to verify the relationship between particle size, total surface area and ROS generation. Further, we investigated the effects of nanoparticle exposure on oxidative stress and pulmonary inflammation on spontaneously hypertensive rats (SHR) and Sprague Dawley (SD) rats.
In cell-free system, ultrafine carbon black (ufCB) with average diameter of 15, 51 and 95nm were suspended in phosphate buffered saline (PBS) in 200, 400 and 800 μg/ml for 1hr. DCF (2’,7’-dichlorofluorescin) assay was used to determine the ROS generation of ufCB. In animal study, SHR and SD rats were intratracheally instillation of 15, 51 and 95nm ufCB in 500 and 1000μg, separately. Animals administrated PBS were treated as control group. Animals were sacrificed 24hr after treatment. Bronchoalveolar lavage fluid (BALF) was collected for pulmonary inflammation analysis. Serum 8-OHdG (8-hydroxy-2'-deoxyguanosine) and peripheral blood DNA single-strand breaks (DNA SSB) were determined to evaluate the effects of oxidative stress.
Our results revealed that the generation of ROS increased with the instilled mass concentration in each particle size. At the same mass concentration, smaller particle size of CB produced greater ROS. Interestingly, the generation of ROS was highly correlated with total surface area of particles (R2=0.83). In animal study, SHR treated with 1000µg of 15nm ufCB had significantly increased the proportion of neutrophils in BALF as compared to SHR with larger particle treatment. No significant increased pulmonary inflammation was observed in SHR treated with 51 and 95 nm ufCB at same dose. SHR with 15nm ufCB treatment demonstrated increased serum 8-OHdG, although it did not reach statistical difference. In addition, SHR treated with 15nm ufCB showed significant increased DNA SSB in tail moment and ％ of tail length as compare to the controls (P<0.05). However, the DNA SSB was reduced with the increasing ufCB treatment with 51 and 95 nm in SHR. In SD rat model, we found the proportion of neutrophils in BALF was significantly increased in SD rats treated with 15nm ufCB as compared to the controls (P<0.05). However, there was no significant elevation of serum 8-OHdG and peripheral blood DNA SSB after ufCB treatment. In SHR, interestingly, we found correlation between oxidative stress and pulmonary inflammation, BALF total cells correlated with serum 8-OHdG (R2=0.23; P<0.05) and the proportion of neutrophils in BALF correlated with peripheral blood DNA SSB (tail moment R2=0.36, % of tail length R2=0.27, % of tail intensity R2=0.25; P<0.05). We also found that serum 8-OHdG statistically correlated with peripheral blood DNA SSB (tail moment R2=0.6, % of tail length R2=0.44, % of tail intensity R2=0.43; P<0.05). In SD rats, serum 8-OHdG correlated with BALF LDH (lactate dehydrogenase) (R2=0.21; P<0.05). However, there was no significant association found between 8-OHdG, peripheral blood DNA SSB and other pulmonary inflammation indicators. Serum 8-OHdG and BALF total cells at baseline or their trends for either exposure dose or total surface area in SHR was higher than those in SD rats. After adjusting for control, the trend of BALF total cells in SHR was higher than those in SD rats. However, the trend of DNA damage indicators, which was adjusted for control, in SHR was higher than those in SD rats only for the exposure to 15nm nanoparticles.
In summary, we found nanoparticles generated ROS in cell free system and the generation of ROS was associated with total surface area of particle. In vivo study found that exposure to nanoparticles caused ROS generation and pulmonary inflammation. In SHR susceptible animal study we found that exposure to ufCB would increase burden on lung inflammation and oxidative damage which were related to total surface area of particles. Our study also indicates that susceptible animals maybe subject to increased risk of lung inflammation and oxidative DNA damage. This may have important policy implication. However, more studies are needed to clarify the above findings.
Subjects
奈米微粒
反應性氧化物種
發炎反應
nanoparticle
reactive oxygen species
inflammation
Type
thesis
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