Evaluation of the Content of Aquaporin-5 and Epithelial Sodium Channel in the Lungs of Rats during the Development of Toxic Pulmonary Edema Caused by Intoxication with Acylating Pulmonotoxicants.

We studied the content of aquaporin-5 (AQP5) and epithelial sodium channel (ENaC) in rat lungs during the development of toxic pulmonary edema (TPE) caused by intoxication with phosgene and perfluoroisobutylene (1.5 LC50). The lung body weight index (LBI) was calculated and histological examination of the lung tissues was performed. Localization and expression of AQP5 and ENaC were determined by immunohistochemistry. Intoxication led to a significant (p<0.05) increase in LBI and histological changes typical of TPE 1 and 3 h after the exposure. In 1 and 3 h after phosgene intoxication, the AQP5 and ENaC content significantly (p<0.05) increased in comparison with the control. Similar changes in the AQP5 and ENaC content were observed 1 and 3 h after exposure to perfluoroisobutylene. It was hypothesized that AQP5 plays an important role in the formation of TPE caused by intoxication with acylating pulmonotoxicants. An increase in the content of ENaC can be considered as a compensatory reaction of the body aimed at clearance of the alveolar fluid.

Ultrastructural Changes in the Air-Blood Barrier of Rats in Acute Intoxication with Furoplast Pyrolysis Products.

Rats were exposed to fluoroplast-4 pyrolysis products (sample weight 2.6 g, pyrolysis temperature 440-750°C, pyrolysis duration 4 min) containing perfluoroisobutylene over 15 min. Lung tissue samples for histological and electron microscopic examination were isolated in 3 and 30 min after intoxication and processed routinely. Histological examination revealed no structural changes in the lungs. In ultrathin sections of rat lungs, some changes in the structure of type I pneumocytes were detected in 3 min after the exposure: detachment of cytoplasmic processes and the appearance of transcytosis pores. These changes attested to impaired cell-cell interactions and their adhesion to the basement membrane, where structural disorganization and edema of the collagen matrix were observed. In 30 min following exposure, the signs of damage to type I pneumocytes became more pronounced. The increase in the equivalents of transcellular and paracellular permeability in the alveolar lining profile was observed. No changes in the pulmonary capillary endotheliocytes were detected, which suggest that type I pneumocytes are the primary target of the toxic effect of perfluoroisobutylene. The vulnerability of a particular cell population, in view of specific metabolism of these cells, can be the key to deciphering of the mechanisms of the toxic effect of pyrolysis products of fluorinated polymer materials.

Mechanisms of Pulmonary Toxicity of Perfluoro-n-Alkane Pyrolysis Products with Consideration of the Structural Features of the Blood-Air Barriers.

Perfluoroisobutylene a is pulmonotoxic chemical generated during pyrolysis of perfluoro-nalkanes (polytetrafluoroethylene). The mechanisms of acute pulmonary toxicity induced by perfluoroisobutylene have not been studied yet. The analysis of tissues of brown frogs showed that the products of polytetrafluoroethylene pyrolysis induce typical inflammatory response in the lungs (fluid accumulation, erythrocyte stasis, desquamation of the epithelium, and capillary plethora in lung septa) and oropharyngeal cavity (degeneration of ciliated epithelium, hyperemia of underlying vessels with plasmatic imbibition of the connective tissue, and margination of segmented leukocytes and monocytes). The absence of surfactant is a specific feature of the blood-air barrier of the oropharyngeal cavity in frogs compared to the lungs. It can be hypothesized that toxic effects of perfluoroisobutylene are determined by its influence on epithelial (pneumocytes and cells of nonkeratinized stratified ciliated epithelium) and endothelial cells. Even though the effects of the agent on surfactant cannot be excluded, they do not determine the probability of development of inflammatory response.

Lethal concentration of perfluoroisobutylene induces acute lung injury in mice mediated via cytokine storm, oxidative stress and apoptosis.

Perfluoroisobutylene (PFIB) is a highly toxic gas that targets the lungs. Low-level inhalation of the gas can lead to acute lung injury (ALI), pulmonary edema and even death. No specific anti-PFIB drugs are currently available and the pathogenesis of PFIB-induced ALI is not fully understood. Early direct oxidative injury and a secondary hyper-inflammatory response are recognized as the primary mechanisms of PFIB-induced ALI. In the present study, our data demonstrate for the first time that a cytokine storm is associated with PFIB-induced ALI. Levels of 10 pro-inflammatory cytokines and one anti-inflammatory cytokine were significantly increased in lung tissues of PFIB-exposed mice. PFIB inhalation additionally led to significant oxidative stress in lung tissue. Inflammation-associated CD11b+Ly6G+Ly6Cint neutrophils and CD11b+Ly6G-Ly6Chi monocytes were significantly increased in blood in association with PFIB-induced ALI. Bcl-2/Bax-mediated lung cell apoptosis was significantly increased at 1 h, followed by a sustained decrease after 1 h, which was significant at 4-8 h in PFIB-exposed mice. This suppression of apoptosis is possibly associated with the Akt-signaling pathway.

[The protective effect of N-acetylcysteine on acute lung injury induced by PFIB inhalation].

Objective: To study the protective effect of N-acetylcysteine on acute lung injury induced by PFIB inhalation and its mechanism. Methods: Survival experiment: 48 male ICR (CD-1) mice were randomly divided into 4 groups, i. e., PFIB control group, NAC prevention group, NAC treatment group, and NAC prevention + treatment group, each group contains 12 animals. The mice of PFIB C group were exposed to PFIB without any treatment. The mice of NAC P group were exposed to PFIB 30min after NAC administration. The mice of NAC T group were exposed to PFIB 1h before NAC administration, The mice of NAC P+T group were administrated with NAC twice (30 min before and 1h after PFIB inhalation) . 150 mg/kg NAC was given by each time. The 7 days survival rate of mice after lethal dose PFIB exposure was observed. 18 male Wistar rats were randomly divided into 3 groups i.e., normal control group (N-C) , PFIB control group (PFIB-C) and NAC prevention group (NAC-P) , with each group contains 6 animals in the second experiment. The rats of N-C group received no treatment. The rats of NAC-P group and PFIB-C group were exposed to PFIB 30min after treatment of NAC (420 mg/Kg, i.p.) and saline, respectively. The respiratory functions of animals were tested before and 24 h after PFIB inhalation. The arterial blood gas was analyzed after rats were anesthetized 24 hours post sublethal dose PFIB exposure. Then samples of BALF, plasma and lung tissue were collected. Wet lung/body weight ratio, protein and phospholipid content in BALF, and T-SOD, GSH, GSH-Px in plasma and lung tissue were measured. The expression of Peroxiredoxin 2 was detected by Westernblot assay. Results: NAC prevention can significantly improve the survival of mice exposed to a lethal dose PFIB while NAC treatment is ineffective. Severe lung edema was observed in rats 24 h after PFIB exposure. Compared to N-C group, the wet lung/body weight ratio, protein and phospholipid content in BALF, and respiratory rate of PFIB control group all increased significantly (P<0.01) . The arterial oxygen partial pressure (PaO(2)) reduced significantly (P<0.05) . The GSH-Px activity in lung tissue reduced significantly (P<0.01) while the expression of Peroxiredoxin 2 increased significantly (P<0.01) . NAC prophylaxis significantly reduced the wet lung/body weight ratio, protein and phospholipid content in BALF, respiratory rate of rats exposed to PFIB (P<0.01) . Compared with PFIB-C group, the PaO(2) (P<0.05) and the activity of GSH-Px (P<0.01) and the expression of Peroxiredoxin 2 in lung tissue (P<0.01) were increased significantly. Conclusion: Acute lung injury induced by PFIB inhalation is related to oxidative stress caused by the stimulation to lung. induced and pulmonary subjected to stimulate the generation of exposure, NAC prevention can regulation of the redox system in lung tissue and protect target organ of the treated animals effectively.

The protective effect of N-acetylcysteine on acute lung injury induced by PFIB inhalation / 中华劳动卫生职业病杂志

Objective@#To study the protective effect of N-acetylcysteine on acute lung injury induced by PFIB inhalation and its mechanism.@*Methods@#Survival experiment: 48 male ICR (CD-1) mice were randomly divided into 4 groups, i. e., PFIB control group, NAC prevention group, NAC treatment group, and NAC prevention + treatment group, each group contains 12 animals. The mice of PFIB C group were exposed to PFIB without any treatment. The mice of NAC P group were exposed to PFIB 30min after NAC administration. The mice of NAC T group were exposed to PFIB 1h before NAC administration, The mice of NAC P+T group were administrated with NAC twice (30 min before and 1h after PFIB inhalation) . 150 mg/kg NAC was given by each time. The 7 days survival rate of mice after lethal dose PFIB exposure was observed. 18 male Wistar rats were randomly divided into 3 groups i.e., normal control group (N-C) , PFIB control group (PFIB-C) and NAC prevention group (NAC-P) , with each group contains 6 animals in the second experiment. The rats of N-C group received no treatment. The rats of NAC-P group and PFIB-C group were exposed to PFIB 30min after treatment of NAC (420 mg/Kg, i.p.) and saline, respectively. The respiratory functions of animals were tested before and 24 h after PFIB inhalation. The arterial blood gas was analyzed after rats were anesthetized 24 hours post sublethal dose PFIB exposure. Then samples of BALF, plasma and lung tissue were collected. Wet lung/body weight ratio, protein and phospholipid content in BALF, and T-SOD, GSH, GSH-Px in plasma and lung tissue were measured. The expression of Peroxiredoxin 2 was detected by Westernblot assay.@*Results@#NAC prevention can significantly improve the survival of mice exposed to a lethal dose PFIB while NAC treatment is ineffective. Severe lung edema was observed in rats 24 h after PFIB exposure. Compared to N-C group, the wet lung/body weight ratio, protein and phospholipid content in BALF, and respiratory rate of PFIB control group all increased significantly (P<0.01) . The arterial oxygen partial pressure (PaO2) reduced significantly (P<0.05) . The GSH-Px activity in lung tissue reduced significantly (P<0.01) while the expression of Peroxiredoxin 2 increased significantly (P<0.01) . NAC prophylaxis significantly reduced the wet lung/body weight ratio, protein and phospholipid content in BALF, respiratory rate of rats exposed to PFIB (P<0.01) . Compared with PFIB-C group, the PaO2 (P<0.05) and the activity of GSH-Px (P<0.01) and the expression of Peroxiredoxin 2 in lung tissue (P<0.01) were increased significantly.@*Conclusion@#Acute lung injury induced by PFIB inhalation is related to oxidative stress caused by the stimulation to lung. induced and pulmonary subjected to stimulate the generation of exposure, NAC prevention can regulation of the redox system in lung tissue and protect target organ of the treated animals effectively.

Long-term toxicity to respiratory system of rats by low-dose perfluoroisobutylene exposure once and possible beneficial effect of early intervention via Qingkailing injection / 中国药理学与毒理学杂志

OBJECTIVE To clarify the long-term toxicity to the respiratory system in a rat model of acute lung injury (ALI) induced by a single low-dose of perfluoroisobutylene(PFIB) inhalation expo?sure,and observe the possible beneficial effect of early intervention via Qingkailing(QKL) injection. METHODS Totally 224 male Wistar rats were randomly divided into 4 groups:normal control group in which air exposure was followed by a saline 10 mL · kg-1(ip),QKL control group in which QKL 10 mL · kg-1 was ip given after air exposure,PFIB exposure group in which rats were exposed to PFIB 280 mg·m-3 for 5 min only,and QKL treatment group in which QKL 10 mL·kg-1 was given ip at 1 h after PFIB exposure. Lung functions of rats were measured at 24 h,3,6,12,24,36 and 48 weeks after exposure. The arterial blood gas,lung coefficient,protein content in bronchoalveolar lavage fluid(BALF),hydroxy?proline(HYP) content in lung tissue and plasma,and other indicators were detected or analyzed. RESULTS Within 24 h after PFIB exposure,the lung coefficient and protein content in BALF were increased significantly(P<0.01),whereas the PaO2(P<0.01) and SaO2(P<0.05) indices in arterial blood decreased significantly in PFIB group compared with normal control. The inhalation time , exhalation time,tidal volume(TV),expired volume(EV)and relaxed time were reduced significantly (P<0.01). However,all the above indicators returned to normal in 3 weeks,but TV,EV and peak expiratory flow were significantly lower than in normal group at 48 weeks(P<0.05). HYP contents in lung tissues,compared with normal control(P<0.05),were reduced significantly within 24 h after PFIB exposure,increased significantly in 6 weeks(P<0.05),then returned to normal in 12 weeks. HYP contents in plasma increased significantly compared with normal control(P<0.05) within 24 h after PFIB exposure but returned to normal in 3 weeks. The protein contents in BALF of QKL treatment group were significantly lower than those in PFIB group(P<0.01) within 24 h after PFIB exposure. From 24 h to 24 weeks after PFIB exposure,changes of pulmonary functions were similar to those in PFIB group. At 48 weeks,TV and EV in QKL treatment group were more significantly increased than those in PFIB group(P<0.05). CONCLUSION Rats with ALI induced by a single low dose of PFIB exposure undergo compensatory repair except for pulmonary capacity and pulmonary ventilation functions. Early treatment with QKL reduces protein content of BALF and alleviates pulmonary edema,and has some beneficial effect on lung function recovery later.

Impact of perfluoroisobutylene exposure on pulmonary microvascular endothelial cells of rats / 中国药理学与毒理学杂志

OBJECTIVE To investigate the change of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),intercellular adhesion molecules(ICAM-1),matrix metalloproteinases 2 (MMP-2) and MMP-9 contents in cultured pulmonary microvascular endothelial cells (PMVECs) in rats after perfluoroisobutylene (PFIB) exposure. METHODS PMVECs were separated and purified using a modified method of implantation of pulmonary tissues. After identification,PMVECs were divided into the normal control group and the PFIB-exposed groups(n=3). The PFIB-exposed groups inhaled PFIB at the concentration of 200 mg · m-3 for 5 min in a flow-past header,while the normal control group were PMVECs in quiescent condition. The supernatants and lysates of PMVECs were harvested at 0.5,1,2,4 and 8 h,respec?tively, after execution. The contents of TNF-α,IL-1β,ICAM-1,MMP-2 and MMP-9 were measured by ELISA,and the activity of MMP-2 and MMP-9 was measured by gelatin zymography. RESULTS① According to the morphologic characteristics of cell growth and the expression of specificity antigens and the bind experiment of phytohemagglutinin,the cells separated and purified by modified method shared the characteristics of PMVECs.②TNF-αwas rapidly expressed by PMVECs at 0.5 h post PFIB stimulation and the maximum value was achieved at 2 h post PFIB stimulation(P<0.05). The newly synthesized TNF-α was slowly released out of the cells. The maximum TNF-α in the supernatant was achieved at 4 h post stimulation.③Within 2 h of stimulation,PMVECs synthesized a large amount of IL-1β and peaks at 2 h. However,IL-1βwas never released to the extracellular milieu.④The amount of ICAM-1 was rapidly synthesized by PMVECs after PFIB stimulation,but at a low level.⑤After stimulation with PFIB,MMP-2 in the supernatant of PMVECs culture was gradually increased,peaked at 2 h and then decreased subsequently. The biological activity of MMP-2 in the supernatant was also enhanced after PFIB stimulation. PFIB did not stimulate synthesis or secretion of MMP-9,indicating that PMVECs were not the main source of MMP-9 during PFIB inhalation-induced acute lung injury. CONCLUSION PFIB stimulates the surviving PMVECs to synthesize a large amount of TNF-α,IL-1β, MMP-2 and conjunctive ICAM-1.

Preventive effect of senecio cannabifolius less.II on perfluoroisobutylene inhalation-induced acute lung injury in rats / 国际药学研究杂志

Objective To preliminarily investigate the effect and possible mechanisms of Senecio cannabifolius Less. II (FHC- n) on perfluoroisobutylene (PFIB) inhalation-induced acute lung injury. Methods Totally 156 rats were randomly assigned to three groups: the control group， the PFIB group and the FHC- H prevention group， with 32， 62 and 62 rats in each group respectively. The FHC- II prevention group were given FHC- II three times per day at the dosage of 340 mg/kg before PFIB exposure. 1 h after the last time of FHC- h administration，the FHC- i prevention group were exposured to gaseous PFIB (0.2 mg/L) for 10 minutes in a static whole-body exposure inhalation system. The survival rate of the rats were recorded at 1，2，4，8，16，24，48 and 72 h post PFIB exposure; the lung index and total protein content in bronchoalveolar lavage fluid (BALF) were measured at 1 h，2 h，4 h，8 h，16 h and 24 h; IL-1β and IL-8 in sera were assayed by enzyme-linked immunosorbent assay (ELISA) at 1，2，4，8 and 16 h post PFIB exposure and the histopathological examination of the lung tissue was performed at 8 h post PFIB exposure. Results FHC-II significantly reduced the content of the total protein in BALF，lung index and the levels of IL-1β and IL-8 in aera as well， and dramatically alleviated the histopathological changes in the lung tissue. Conclusion FHC- D demonstrates some preventive effect on PFIB inhalation-induced acute lung injury in rats.

Preventive effect of Senecio cannabifolius Less.Ⅱon perfluoroisobutylene inhalation-induced acute lung injury in rats / 国际药学研究杂志

Objective To preliminarily investigate the effect and possible mechanisms of Senecio cannabifolius Less.Ⅱ(FHC-Ⅱ) on perfluoroisobutylene (PFIB) inhalation-induced acute lung injury. Methods Totally 156 rats were randomly assigned to three groups: the control group, the PFIB group and the FHC-Ⅱ prevention group, with 32, 62 and 62 rats in each group respectively. The FHC-Ⅱprevention group were given FHC-Ⅱthree times per day at the dosage of 340 mg/kg before PFIB exposure. 1 h after the last time of FHC-Ⅱ administration, the FHC-Ⅱ prevention group were exposured to gaseous PFIB (0.2 mg/L) for 10 minutes in a static whole-body exposure inhalation system. The survival rate of the rats were recorded at 1, 2, 4, 8, 16, 24, 48 and 72 h post PFIB exposure;the lung index and total protein content in bronchoalveolar lavage fluid (BALF) were measured at 1 h, 2 h, 4 h, 8 h, 16 h and 24 h; IL-1β and IL-8 in sera were assayed by enzyme-linked immunosorbent assay (ELISA) at 1, 2, 4, 8 and 16 h post PFIB exposure and the histopathological examination of the lung tissue was performed at 8 h post PFIB exposure. Results FHC-II significantly reduced the content of the total protein in BALF, lung index and the levels of IL-1β and IL-8 in aera as well, and dramatically alleviated the histopathological changes in the lung tissue. Conclusion FHC-Ⅱ demonstrates some preventive effect on PFIB inhalation-induced acute lung injury in rats.

Role of angiotensinⅡin perfluoroisobutylene-induced acute lung injury in rats / 军事医学

Objective To investigate tentatively the role of angiotensionⅡ( AngⅡ) in perfluoroisobutylene ( PFIB)-in-duced acute lung injury ( ALI) in rats.Methods Twenty-eight male Wistar rats were randomly divided into one control group(0 h) and six PFIB-exposed groups which were executed at 1, 2, 4, 8, 16 and 24 h after PFIB exposure (n=4). The PFIB-exposed groups inhaled PFIB at a concentration of 145 mg/m3 for 8 min in a flow-past header while the control group was exposed to the filtered air in a similar manner .After execution at the corresponding time-point, the samples of the lung, serum and brochoalveolar lavage fluid (BALF) were harvested.The measurement of the lung wet-to-dry weight ratio ( W/D) and total protein content in BALF , and the histopathological examination of the lung were carried out to evalu -ate the degree of lung injury .The over-time changes in the content of AngⅡin the lung homogenates and blood plasma and the activity of angiotensin converting enzyme ( ACE) in the lung tissue were observed .Results The lung W/D and total protein content in BALF were increased significantly at 16 h after PFIB exposure with severe acute lung edema and abun-dant neutrophil exudation to the alveoli , which were alleviated dramatically at 24 h after PFIB exposure .The content of AngⅡin the lung homogenate showed a tendency of increase during the first 8 hours with significant decrease at 16 and 24 h after exposure.However, the content of AngⅡin the plasma and the activity of ACE in the lung experienced of fluctuations , but without significant difference compared to the control group .Conclusion There is no obvious correlation between the extent of lung injury and that of AngⅡin the lung.The pathological significance of AngⅡin PFIB-induced ALI needs to be further clarified.

Long-term effect of a single perfluoroisobutylene exposure induced acute lung injury in mice / 中国药理学与毒理学杂志

OBJECTIVE To investigate whether the pulmanary fibrosis formed after a single PFIB exposure.METHODS A total of 70 male mice were exposed to PFIB 130 mg·m-3 for 5 min.Pulmonary edema of 10 mice was evaluated by lung indices at 24 h after PFIB exposure.Pathological changes and collagen deposition were detected by hematoxylin and eosin (HE) and Sirius red stainings in the other mice,changes in collagen content in lungs and plasma by measuring the respective hydroxyproline content at 2,4,6,8,12 and 16 weeks after PFIB exposure.RESULTS Severe pulmonary edema was observed at 24 h after PFIB exposure.At day 14 after PFIB exposure,inflammatory cell infiltration,alveolar septum thickening,interstitial and alveolar edema and protein leakage were noticed.Collagens types Ⅰ and Ⅲ on the wall of vessel and bronchi were severely damaged,but considerable amount of collagen type Ⅲ deposited on the alveolar wall.The content of hydroxyproline considerably decreased in the lungs but increased significantly in the plasma up to six weeks.Hydroxyproline in lungs and plasma began to recover at the end of 8 weeks,and then returned to normal.At 16 weeks,they recovered to normal level.At the end of 4 weeks,the lung lesions and the collagens at the wall of vessel and bronchi began to recover gradually; collagen typeⅢ at the alveolar wall was gradually absorbed,too.At 16 weeks,the lungs almost recovered to normal level.CONCLUSION At earlier phase after PFIB exposure,the excessive collagens destruction in lungs is observed,but no pulmonary fibrosis forms at the later phase.

Variation and clinical application value of fluoride concentrations in the serum and urine in patients with acute organic fluoride poisoning by inhaling / 中华急诊医学杂志

Objective To investigate the changes of serum and urine fluorion organic fluoride poisoning by inhaling, and to probe into the clinical application value of concentrations in different degrees and at different time in patients with acute evaluating the sertm and urine fluorion concentration in acute organic fluoride poisoning by inhaling. Method A study was carried out in 23 patients, who suffered from acute organic fluoride poisoning by inhaling and were admitted Zhejiang Quhua Hospital, from December 2000 to December 2008. According to the occupational acute organic fluoride poisoning diagnostic criteria(GBZ66-2002),23 patients were divided into mild poisoning group,moderate poisoning group and severe poisoning group. Serum and urine fluorion concentration of patients at 1,2,3,4,5 d after poisoning were measured by using Ion-Selective Electrodes. Fluorion concentration of 10 staffs of Fluorine chemical company was also measured at the same period as the control group. The values of serum and urine fluorion concentration were analyzed. Differences in serum and urine fluorion concentration between groups at different time points were compared by repeated measures ANOVA and variability were deemed as statistical significance when P ＜ 0.05. Results Compared with mild poisoning group, there was no statistically significant difference ( P ＞ 0.05) in serum and urine fluorion concentration at the same time point in 1 to 5 days after poisoning in moderate poisoning group, but there was statistically significant differences ( P ＜ 0.05 or P ＜0.01) in severe poisoning group. Compared with moderate poisoning group, there was statistically significant difference ( P ＜ 0.05) in serum and urine fluorion concentration at the same time point in 1 to 5 days after poisoning in severe poisoning group. Serum fluorion concentration in 1 to 5 days after poisoning in each poisoning groupswere statistically higher than those in control group ( P ＜ 0.05), but there was statistically significant elevation ( P＜ 0.05) in urine fluorion concentration only in 1 day in mild poisoning group, in 1 to 3 days in moderate poisoning groups, in 1 to 5 days in severe poisoning group. Conclusions Serum fluorion concentration can be used as the severity index of diagnosis and determine the extent in 5 days after acute organic fluoride poisoning by inhaling,and urine fluorion concentration can also be used as diagnostic indicators of intoxication, but only in earlier stage or severe poisoning.