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OBJECTIVE: To investigate the hepatotoxicity induced by subchronic crotonaldehyde exposure in male rats and analyze its possible mechanism. METHODS: The specific pathogen-free male Wistar rats were randomly divided into control group, low-, medium-and high-dose crotonaldehyde groups, with 10 rats in each group. The crotonaldehyde solution at doses of 0.0, 2.5, 4.5, and 8.5 mg/kg body weight were given by intragastric administration, once a day, 5 days per week, and continuous for 90 days. The body weight of the rats were weighed during the exposure period. After the exposure, the liver organ coefficients and histopathological changes of the rats were observed. The activities of alanine aminotransferase(ALT) and aspartate aminotransferase(AST) and the level of total bilirubin(TBIL) in the serum of rats were determined by colorimetry. The levels of malondialdehyde(MDA) and reduced glutathione(GSH), and the activities of glutathione peroxide(GSH-Px) and superoxide dismutase(SOD) were determined by colorimetry. The levels of interleukin(IL)-1β, IL-6, tumor necrosis factor alpha(TNF-α) and interferon-γ(IFN-γ) was detected by enzyme-linked immunosorbent assay. RESULTS: At the end of exposure, the increment of body mass of rats in the low-, medium-and high-dose crotonaldehyde groups was lower than that in the control group(P<0.05). The organ coefficients of rats in the middle-and high-dose groups were lower than that in the control group(P<0.05). The liver tissues of the three doses crotonaldehyde groups showed varying degrees of inflammatory cell infiltration. The activities of ALT, AST and the level of TBIL in the serum of rats increased with the increase of the crotonaldehyde exposure dose(P<0.01). With the increase of the crotonaldehyde dose, the level of MDA in rat liver tissue increased(P<0.01), and the level of GSH and the activities of GSH-Px and SOD decreased(P<0.01). The levels of IL-1β, IL-6, TNF-α and IFN-γ in rat liver tissues increased(P<0.05). CONCLUSION: Crotonaldehyde exposure can cause liver tissue damage in rats. Its mechanism of action may be related to the changes of oxidative balance and upregulation of the expression of inflammatory factors in liver tissue. These changes have the dose-effect relationship.
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OBJECTIVE: To observe the effect and mechanism of crotonaldehyde sub-chronic exposure-induced pyroptosis in pulmonary inflammatory reaction in male rats. METHODS: Specific pathogen-free male Wistar rats were randomly divided into control group and low-, medium-and high-dose groups, with 10 rats in each group. Rats were treated with crotonaldehyde at concentrations of 0.0, 2.5, 4.5, and 8.5 mg/kg body weight by intra-gastric administration, once per day for 120 consecutive days. Rats′ body mass was recorded during exposure. After exposure, the rats were sacrificed, and the lung tissues were isolated. The relative expression of reactive oxygen species(ROS) in lung tissues was detected by fluorescent probes at the end of crotonaldehyde exposure. The fluorescent staining of the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3(NLRP3) and Caspase-1 in lung tissues was observed by immunofluorescence microscope. The protein expression of NLRP3, apoptosis-associated spot-like protein(ASC), Caspase-1 precursor(pro-Caspase-1), Caspase-1, interleukin(IL)-1β and IL-18 in lung tissues was determined by Western blotting. RESULTS: At the end of exposure, the body weight gain of rats decreased with the increasing doses of crotonaldehyde(P<0.01). Among them, the body weight gain in the medium-and high-dose groups was lower than that of the control group(P<0.05). After exposure, the lung tissue of each group showed severe inflammatory change with the increasing doses of crotonaldehyde. Immunofluorescence staining showed that the expression of NLRP3 and Caspase-1 in the lung tissues of rats increased in a dose-dependent manner. The relative expression of ROS and the protein of NLRP3, ASC, pro-Caspase-1, Caspase-1, IL-1β and IL-18 in lung tissue of each group increased with the dose of crotonaldehyde(P<0.01). The above indexes of lung tissue in the medium-and high-dose groups were higher than that in the control group(P<0.05). CONCLUSION: Sub-chronic exposure to crotonaldehyde may cause pyroptosis by up-regulating ROS expression in the lung tissues of rats. The up-regulation of Caspase-1 classic dependent pathway leads to pyroptosis, thereby causing inflammatory responses in the lungs.
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OBJECTIVE: To observe the conditions of sub-chronic crotonaldehyde exposure-induced pulmonary inflammation,oxidative stress and apoptosis in male rats,and to explore the related mechanisms. METHODS: The specific pathogen free Wistar male rats were randomly divided into control group,low-,medium-and high-dose crotonaldehyde groups,with 10 rats in each group. Rats were treated with crotonaldehyde at the concentrations of 0. 0,2. 5,4. 5 and 8. 5 mg/kg body weight by intra-gastric administration,once per day for 120 consecutive days. After the end of treatment,rats were sacrificed,the lungs were weighed and histopathological examination was performed. The levels of malondialdehyde( MDA),superoxide dismutase( SOD) and glutathione peroxidase( GSH-Px) in the serum of rats were determined by colorimetry. The relative expression of reactive oxygen species in lung tissues was detected by fluorescence probe. The apoptosis rate was detected by Tunel staining. The relative expression of B-cell lymphoma( Bcl)-2,Bcl-2 associated X protein( Bax) and cysteine aspartic acid protease-3( Caspase-3) proteins in lung tissue was detected by western blotting.RESULTS: The body weight of the rats in the high-dose group began to decrease after 30 days of exposure( P < 0. 05),and the body weight in the low-and medium-dose groups began to decrease at 90 days exposure( P < 0. 05),when compared with that of the control group at the same time. The body weight of the high-dose group was lower than that of the low-and medium-dose groups began to decrease at 90 days exposure( P < 0. 05). After exposure,the lung tissue of the three doses groups showed different degrees of inflammatory change in a dose-effect relationship. The level of serum MDA in rats increased with the treatment of crotonaldehyde( P < 0. 01). The activities of serum SOD and GSH-Px decreased with the treatment of crotonaldehyde( P < 0. 01). The relative expression of ROS and apoptosis rate in rat lung tissue increased with the treatment of crotonaldehyde( P < 0. 01). The relative expression of Bcl-2 protein and the ratio of Bcl-2/Bax in the lung tissue of rats decreased with the treatment of crotonaldehyde( P < 0. 01). The relative protein expression of Bax and Caspase-3 increased with the treatment of crotonaldehyde( P < 0. 01). CONCLUSION: Crotonaldehyde sub-chronic exposure can cause apoptosis in lung tissue by altering the oxidative balance,leading to inflammatory pathological changes in the lung.
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Objective@#To investigate the effect of subchronic exposure to crotonaldehyde on reproductive damage and oxidative stress in male rate.@*Methods@#Forty male SPF Wistar rats were randomly divided into four group: a control group and 3 exposure groups, 10 per group. The rats in each group were continuously administrated with crotonaldehyde (normal saline) for 1 time/d. For 128 d, the doses were 0.0, 2.5, 4.5, 8.5 mg/kg. After the end of the exposure, the body weight, the weight of the testis and epididymis was measured, and calculating organ coefficient. The left spermatozoon tail was used to determine sperm motility, number and testicular tissue marker enzyme activity : LDH, SDH, ACP, γ-GT; blood biochemical related index concentration: FSH, LH, T; oxidative stress-related indicator concentrations: MDA, SOD, GSH-Px and CAT.@*Results@#Compared with the control group, the weight gain, testicular and epididymis weight, and organ coefficient of the rats in the 4.5 and 8.5 mg/kg groups were decreased, the difference was statistically significant (P<0.05) . In the exposed group, the testicular tissue volume was reduced, the color was dark, and the number of germ cells in some seminiferous tubules was reduced. Compared with the control group, the sperm count and sperm motility of the 4.5 and 8.5 mg/kg groups were significantly lower (P<0.05) ; compared with the control group, 4.5 and 8.5 mg/kg. The activities of serum ACP, LDH, SDH and γ-GT in the exposed group were significantly lower (P<0.05) . Compared with the control group, the serum levels of T in the 8.5 mg/kg group were decreased. The levels of LH and FSH in the 4.5 and 8.5 mg/kg exposure groups were significantly lower (P<0.05) . Compared with the control group, the rats in the 4.5 mg/kg and 8.5 mg/kg exposure groups were compared. The activity of MDA in serum increased, SOD, GSH-Px and CAT activity decreased, the difference was statistically significant (P<0.05) .@*Conclusion@#Crotonaldehyde may cause subchronic reproductive damage and oxidative damage in rats by altering the hormone of the reproductive system, the expression of antioxidant enzymes, and destroying the oxidative balance of the rat.
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Objective@#To observe the lung injury of male rats induced by sub-chronic exposure to crotonaldehyde, and to explore the possible mechanism of injury.@*Methods@#Forty SPF male Wistar rats were randomly divided into control group and 3 groups in each group, and each group received 0.0, 2.5, 4.5, 8.5 mg/kg body weight crotonaldehyde solution for continuous intragastric administration. 120 d, once a day. After the end of the exposure, the body weight of the rats was measured, and the lung tissues were quickly separated after cervical dislocation. The organ coefficients were calculated and histopathological examination was performed to determine malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione. Peroxidase (GSH-Px) content; ELISA was used to measure interleukin (IL) -6, IL-1β, and tumor necrosis factor (TNF) -α in lung tissues.@*Results@#Compared with the control group, the weight gain of the rats in the 4.5 and 8.5 mg/kg exposure groups was small, and the lung weight and organ coefficient of the exposed group decreased, the difference was statistically significant (P<0.05). In the exposed group, the lung tissue structure was disordered, the alveolar wall was thickened, and inflammatory cell infiltration was observed. Compared with the control group, the MDA activity in the serum of the rats in the 4.5 mg/kg and 8.5 mg/kg groups increased, and the SOD and GSH-Px activities decreased, the difference was statistically significant (P<0.05). TNF-α levels in the lung tissues of rats exposed to 4.5 mg/kg and 8.5 mg/kg, and levels of (IL) -6 and IL-1β in the lungs of rats in the 2.5, 4.5, and 8.5 mg/kg groups. Significantly increased, the difference was statistically significant (P<0.05) .@*Conclusion@#Crotonaldehyde may induce inflammatory and oxidative stress damage in rats by up-regulating the expression of inflammatory factors in lung tissue and changing the oxidative balance.
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Objective@#To observe the effect of long term crotonaldehyde exposure on heart damage in male rats, and to explore the possible mechanism of toxic action.@*Methods@#24 specific pathogen free healthy male wistar rats were randomly divided into 4 groups with 6 rats in each group. Rats were treated with with 8.5, 4.5, 2.5 and 0.0 mg/kg body weight crotonaldehyde by gavage, once a day for consecutive 150 days. After the last treatment, they were anesthetized and collected blood samples by cardiac puncture. The heart was rapidly separated after cervical dislocation. The cardiac organ coefficient was calculated and the histopathology changes in heart were observed by HE staining. At the same time, the activities of creatine kinase (CK) , lactate dehydrogenase-L (LDH-L) in serum were determined by automatic biochemical analyzer. Moreover, the levels of cardiac troponin (cTnT) , Angiotensin Ⅱ (Ang Ⅱ) , Brain natriuretic peptide (BNP) , Aldosterone (ALD) and interleukin (IL) -6, 8, 1β, interferon (IFN) -γ and tumor necrosis factor (TNF) -α in heart were determined by enzyme linked immunosorbent assay.@*Results@#At the 90d, 120 d, and 150 d exposure, compared with the control group, the body weight gain in 4.5 and 8.5 mg/kg groups were decreased. Moreover, the heart weight in 4.5 and 8.5 mg/kg groups, and heart coefficient in 8.5 mg/kg group were decreased (P<0.05) . With the increasing dosage of crotonaldehyde, the degree of pathological changes in the heart of exposed rats were aggravated. The major pathological changes of heart in 4.5 and 8.5 mg/kg groups could be summarized as lymphocyte infiltration, abnormal cardiac muscle fiber arrangements, necrosis and fibrous connective tissue hyperplasia. Compared with the control group, the serum CK activity in 4.5 mg/kg group, CK and LDH-L activitivies in 8.5 mg/kg group were increased (P<0.05) ; Compared with the control group, the levels of ALD and ANGII in the heart of 4.5 and 8.5 mg/kg groups were increased, BNP level were decreased, and cTNT level in 8.5 mg/kg group were increased (P<0.05) . Compared with the control group, the levels of IL-1β、IL-6、IL-8 in 4.5 mg/kg group and IL-1β、IL-6、IL-8、TNF-α、IFN-γ in 8.5 mg/kg group were increased (P<0.05) .@*Conclusion@#Crotonaldehyde could up-regulate cardiac inflammatory cytokines and alter the balance ofangiotensin-aldosterone-brain natriuretic peptide causing heart damage.
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Objective@#To observe the effect of crotonaldehyde long-term exposure on kidney injury in male rats, and to explore the specific mechanism of toxic action.@*Methods@#32 specific pathogen free healthy adult male wistar rats were randomly divided into 4 groups with 8 rats in each group: high-, moderate-, low-dose groups and a control group. Rats were treated with 8.5, 4.5, 2.5 and 0.0 mg/kg body weight crotonaldehyde by gavage, once a day for consecutive 128 days. After the last treatment, they were sacrificed and separated bilateral kidney. Kidney organ coefficients were calculated and the histopathology changes in kidney were observed by HE staining. The activities of malondialdehyde (MDA) , superoxide dismutase (SOD) , glutathion peroxidase (GSH-Px) and the levels of malaondialdehyde uric acid (UA) , urea nitrogen (BUN) , creatinine (CR) in serum were determined in the same time. Moreover, the levels of interleukin (IL) -6, 8, interferon (IFN) -γ and tumor necrosis factor (TNF) -α in kidney were determined by enzyme linked immunosorbent assay.@*Results@#Bilateral kidneys in the 8.5 mg/kg group were reduced in size and dark in color. Under the microscope, the major pathology changes of kidneys could be summarized as summarized as protein cast renal tubule, inflammatory cells and lymphocytes infiltration among kidney cortex. Compared with the control group, the weight gain of rats in 8.5 mg/kg group were smaller, and the weight and organ coefficient of kidney in each groups were significant decreased (P<0.05) . With the increasing dosage of crotonaldehyde, the serum BUN and UA levels in 8.5, 4.5 mg/kg groups, CR level in 8.5 mg/kg group were significant increased (P<0.05) .Compared with the control group, the serum MDA level in 8.5 mg/kg group was significant increased (P<0.05) . However, serum SOD activity in 8.5 mg/kg group and GSH-PX activity in 8.5, 4.5 mg/kg groups was significant decreased (P<0.05) . With the increasing dosage of crotonaldehyde, there are upward trend in the kidney IL-6, IL-8, TNF-α, IFN-γ, β-2 microglobulin levels. Compared with the control group, IL-6, TNF-α, IFN-γ, β-2microglobulin levels in 8.5, 4.5 mg/kg groups and IL-8 level in 8.5 mg/kg group in kidney were significant increased (P<0.05) .@*Conclusion@#Crotonaldehyde could develop the inflammatory factors levels and change the oxidation balance condition in the kidney of male rats, causing the inflammatory and oxidative injures of renal tissues.
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Objective To study the effect of an essential α,β-unsaturated aldehyde from cigarette smoke crotonal-dehyde on myocardial contractile function and intracellular Ca2+function in mice. Methods Hearts of from male C57BL/6 mice were digested by Langendorff to islate the cardiomyocytes. The cardiomyocytes of mice were then in-cubated with crotonaldehyde(1,10,25 and 50 μmol/L) for 6 h,and the control group was treated without croton-aldehyde,then they were evaluated including peak shortening(PS),maximal velocity of shortening/relengthening (±dL/dt),time-to-PS(TPS),time-to-90% relengthening(TR90),fura-2 fluorescence intensity(FFI),intracel-lular Ca2+decay and SERCA 45Ca2+uptake and the expression of Na+-Ca2+exchange were evaluated by Western blot analysis. Results Compared with the control group, the higher concentrations of the crotonaldehyde(25 and 50 μmol/L) groups significantly diminished the PS, ±dL/dt,ΔFFI,SERCA activity and Ca2+decay (P<0.05), as well as prolonged the TR90(P<0.05);however the crotonaldehyde with different concentrations had no effect on the expression of Na+-Ca2+exchange in cardiomyocytes. Conclusions Crotonaldehyde may inhibit cardiomyocyte contraction by suppressing SERCA activity and compromising intracellular Ca2+handling.
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OBJECTIVE: To observe the effect of crotonaldehyde exposure on lung injury in male rats,and to explore the mechanism of toxic action. METHODS: Specific pathogen free healthy adult male Wistar rats were randomly divided into 4groups with 10 rats in each group: a control group and low-,medium- and high-dose groups. Rats were treated with 0. 00,2. 11,4. 22,8. 44 mg / kg body weigh crotonaldehyde by intra-gastric administration,once per day for 25 consecutive days.After the last treatment,rats were secrificed and the lung was isolated. Lung organ coefficients were calculated and the pathologic changes in lung tissues were observed. The levels of interferon( IFN)-γ,interleukin( IL)-1 β,IL-4,IL-6 and tumor necrosis factor( TNF)-α in lung tissue were determined by enzyme-linked immunosorbent assay. RESULTS: The lung tissue in both the medium- and high-dose group showed early inflammatory pathological changes with alveolar structure damage,interval widened and inflammatory cell infiltration,congestion,bronchiolar epithelial hyperplasia,visible red blood cells and inflammatory cells infiltration. The changes in the high-dose group were more severe than that in the medium-dose group. The lung organ coefficient and the levels of IFN-γ,IL-1 β,IL-4,IL-6,TNF-α in lung tissue were increased in dose-effect relations( P < 0. 01). CONCLUSION: Crotonaldehyde can increase the level of inflammatory factors in the lung tissues of rats,causing inflammatory injury of lung tissue.