Studies On The Pulmonary Toxicity Of Indium-Tin Oxide In Rats

Objectives The main purpose was to study the pulmonary toxicological effects and pathogenesis of indium-tin oxide（ITO）.The experiment would be useful for filling the blank for ITO toxicity research and provide a scientific basis for formulating indium occupational exposure limits and treating of indium poisoning.Methods Healthy male Wistar rats were randomly divided into five groups,namely,the control group,ITO₃ mg/kg group,ITO 6 mg/kg group,In₂O₃ 5.4 mg/kg group and SnO₂ 0.6 mg/kg group（each group n=8）.After ether anesthesia,the rats were intratracheally administered solution,twice a week,for eight weeks.Control rats were given physiological sodium chloride solution only.In the exposure period,we weighed the weight of all rats weekly and observed the condition of drinking,food,activities and so on.The rats were euthanized serially up to eight weeks after the final instillation and followed by collecting samples of lung to weigh.We collected the blood by abdominal aorta,centrifuged and measured the SP-A,SP-D,KL-6 and GM-CSF levels in serum by enzyme-linked immunoassay.After microwave digestion,inductively coupled plasma mass spectrometry was used to determine the indium content in blood and lung tissues.We determined the levels of T-SOD,MDA,T-AOC and LDH in lung tissue homogenate.The lung tissue of HE staining was used to observe the pathological changes.The ultrastructure of lung tissues was observed by transmission electron microscopy.The expressions of SP-A,SP-D,KL-6 and GM-CSF in lung were evaluated by the immunohistochemical method and the protein expressions of Bax and Caspase-3 were detected by the western blot method.Results 1 The basic situation of rats in all groups were normal and no obvious clinical symptoms were observed during the experiment period.Body weights of all groups always gained during the instillation period.Relative lung weights among all the exposed groups were significantly increased compared to that in the control group（P<0.05）.2 Histopathological changes: the rats’ lung tissues structure was clear and alveolar structure was complete in the control group;the alveolar structure in ITO₃mg group was relatively intact,but scattered eosinophilic granular materials and deposited single ITO particles could be seen in the alveolar space;the alveolar space of treated rats in ITO 6 mg group and In₂O₃ group was filled with eosinophilic particles and gathered a large amount of brown granules,but changes of ITO 6 mg group were more obvious than those in In₂O₃ group;SnO₂ group rats’ lung structure was complete,accompanied by a small amount of inflammatory cells infiltration.3 Transmission electron microscopy（TEM）examinations: the lungs of control rats and SnO₂ group rats showed clear structure of villi,relatively bigger nucleus,well developed endoplasmic reticulum,rich mitochondria crest and more concentrically arranged lamellar bodies in type II alveolar epithelial cells;type II epithelial cells of rats exposed to ITO and In₂O₃ of could be seen sparse microvillus,slightly irregular karyotype,a little of expansion endoplasmic reticulum,cavitation matrix,swelling mitochondria,vacuolation of lamellar corpuscles,the vacuolated phenomenon of laminated annular structures in ITO 6mg group was more serious than that in ITO₃mg group and In₂O₃ group.4 The blood indium concentrations in ITO 6mg group and In₂O₃ group were obviously increased,but the blood indium concentration in In₂O₃ group and SnO₂ group was lower than that of in ITO 6mg group;the contents of indium in lungs in the two ITO groups and In₂O₃ group gradually increased（P<0.05）.5 The level of total superoxide dismutase（T-SOD）in two ITO groups were higher than that in the control group,but the level of T-SOD in In₂O₃ group and SnO₂ group was lower than that in ITO 6mg group.The level of malondialdehyde（MDA）activity and lactate dehydrogenase（LDH）activity in lungs among both ITO groups are significantly increased（P<0.05）.6 The serum GM-CSF level among all treated groups was significantly increased;the serum SP-A contents in ITO₃mg group,In₂O₃ group and SnO₂ group were distinctly reduced,but the concentration of serum SP-A in ITO₃mg group was obviously lower than that in ITO 6mg group;the contents of serum SP-D in two ITO groups and In₂O₃ group were significantly lower that in control group,but the content of serum SP-D in SnO₂ group was higher than that in ITO 6mg group（P<0.05）.7 Immunohistochemical results: the expressions of SP-A in ITO 6mg group and In₂O₃ group were increased,but the expressions of SP-A in ITO₃mg group,In₂O₃ group and SnO₂ group were much lower than that in ITO 6mg group;the expressions of SP-D in ITO 6mg group and In₂O₃ group were increased,but the expressions of SP-D in ITO₃mg group and SnO₂ group were lower than those in ITO 6mg group;the expression of KL-6 in ITO 6mg group was increased,but the expressions of KL-6 in ITO₃mg group,In₂O₃ group and SnO₂ group was much lower than those in ITO 6mg group;the expressions of GM-CSF in ITO 6mg group,In₂O₃ group and SnO₂ group were increased,but the expressions of GM-CSF in ITO₃mg group,In₂O₃ group and SnO₂ group was much lower than those in ITO 6mg group（P<0.05）.8 Weatern blot results: the expression level of Bax and Caspase-3 protein in the lungs increased significantly in two ITO groups and In₂O₃ group,but the expression level of Bax and Caspase-3 protein in the lungs in In₂O₃ group and SnO₂ group was much lower than that in ITO 6mg group（P<0.05）.Conclusions 1 The exposure of ITO lead to the deposition of ITO particles in rats’ lung tissues,may make indium concentration in blood and lung increase significantly.2 The exposure of ITO may induce oxidative and affect the activity of T-SOD and MDA.3 The exposure of ITO and In₂O₃ lead to the deposition of ITO and In₂O₃ particles in rats’ lung tissues,the pulmonary surfactant was accumulated in alveoli for the reason that the remove of pulmonary surfactant was blocked,the expressions of SP-A,SP-D,KL-6 and GM-CSF in lungs were increased,which may be relation with the formation of pulmonary alveolar proteinosis.4 It may suggest that the pulmonary toxicity of ITO was more serious than that of single chemical composition（indium oxide and tin oxide）under the same dose.