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SUMMARY: Cadmium is a highly toxic metal and affects the respiratory mucosa. The aim of the study is to show the inflammation and degenerative effect of cadmium on the olfactory mucosa. In this study, eight-week-old Wistar rats with an average weight of 170-190 g were divided into two groups (control and experiment) with 20 animals in each group and used in the experiments. The rats in the experimental group were given 2 mg/kg/day powdered cadmium chloride dissolved in water intraperitoneally every day for two weeks. At the end of the experiment, the nasal cavity was completely removed with anesthesia. Concha nasalis superior was separated, fixed with zinc-Formalin solution and decalcified with 5 % EDTA (Ethylene-diaminetetraacetic acid). After routine histopathological procedure, APAF-1 antibody was used for expression of Hematoxylin-Eosin (HE) and immunohistochemistry. Histopathological examination revealed interruptions in the basement membrane structure due to cadmium and degenerative changes in stem cells, degeneration in sensory cells and pycnosis in nuclei, dilatation in blood vessels and increased inflammation in connective tissue. APAF-1 expression was found to increase in epithelial cells and olfactory glands (Bowman gland) cells. It has been thought that cadmium toxicity increases cell degeneration and inflammation in the olfactory mucosa and may significantly affect cell death and olfactory metabolism by inducing the pro-apoptotic process.
El cadmio es un metal altamente tóxico que afecta la mucosa respiratoria. El objetivo fue mostrar el efecto inflamatorio y degenerativo del cadmio sobre la mucosa olfativa. En este estudio, ratas Wistar de ocho semanas de edad con un peso promedio de 170-190 g se dividieron en dos grupos (control y experimental) con 20 animales en cada grupo. Las ratas del grupo experimental recibieron 2 mg/kg/día de cloruro de cadmio en polvo disuelto en agua por vía intraperitoneal todos los días durante dos semanas. En los animales se exirpó la cavidad nasal bajo anestesia. Se separó la concha nasal superior, se fijó con solución de zinc-Formalina y se descalcificó con EDTA (ácido etilendiaminotetraacético) al 5 %. Después del procedimiento histopatológico de rutina, Hematoxilina- Eosina (HE) e inmunohistoquímica, se utilizó el anticuerpo APAF-1. El examen histopatológico reveló interrupciones en la estructura de la membrana basal debido al cadmio y cambios degenerativos en las células madre, degeneración en las células sensoriales y picnosis en los núcleos, dilatación de los vasos sanguíneos y aumento de la inflamación en el tejido conjuntivo. Se encontró que la expresión de APAF-1 aumenta en las células epiteliales y en las células de las glándulas olfatorias (glándulas de Bowman). Se ha pensado que la toxicidad del cadmio aumenta la degeneración celular y la inflamación en la mucosa olfativa y puede afectar significativamente la muerte celular y el metabolismo olfativo al inducir el proceso proapoptótico.
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Animals , Rats , Olfactory Mucosa/drug effects , Olfactory Mucosa/pathology , Cadmium Chloride/toxicity , Administration, Intranasal , Immunohistochemistry , Rats, Wistar , Apoptotic Protease-Activating Factor 1ABSTRACT
Objective To investigate the effect of calycosin on cerebral ischemia/reperfusion injury and its mechanism. Methods Forty SPF male SD rats were randomly divided into sham group, model group, calycosin group (20 mg/kg), nimodipine group (0.7 mg/kg, positive control group). The occlusion model of middle cerebral artery in rats was established by modified thread occlusion method, and the environment of cerebral ischemia-reperfusion injury was simulated in vivo. Zea longa score was used to detect the neurological deficit of rats after ischemia-reperfusion injury, 2, 3, 5-triphenyltetranitrogen (TTC) was used to detect the volume of cerebral infarction, HE staining was used to detect the pathomorphological changes of nerve cells, Nissl staining was used to observe the changes of nissl bodies, TUNEL staining was used to detect the apoptosis of nerve cells, Western blotting was used to detect the expression of cytochrome C (Cyt C), apoptotic protease activating factor-1 (Apaf-1), Caspase-9 and Caspase-3. Results Compared with the sham group, the neurological deficit symptoms in the model group were significant (P<0.05), the volume of cerebral infarction increased significantly (P<0.05). Under the microscope, it was found that the nerve cells showed contraction of cell body, hyperchromatic and pyknosis of nucleus and poor growth state, the expression of nissl body reduced significantly (P < 0.05), the apoptotic nerve increased significantly (P< 0.05), the expression of Cyt C, Apaf-1, Caspase-9 and Caspase-3 increased significantly (P<0.05). Compared with the model group, the neurological deficit symptoms of calycosin group and nimodipine group reduced significantly (P<0.05), the volume of cerebral infarction reduced significantly (P<0.05). Under the microscope, the damage of nerve cells reduced significantly, the expression of nissl body increased significantly (P<0.05), the apoptotic nerve reduced significantly (P<0.05), the expression of Cyt C, Apaf-1, Caspase-9 and Caspase-3 decreased significantly (P<0.05). Conclusion Calycosin can significantly inhibit the apoptosis of nerve cells and reduce the cerebral ischemia-reperfusion injury. Its mechanism of action is related to the effective regulation of Cyt C/Apaf-1 apoptosis signaling pathway by calycosin.
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@#Objective To observe the effect of electroacupuncture on Caspase-9, cytochrome C (Cyt-C) and apoptotic protease activating factor 1 (Apaf-1) protein in rats with neurogenic bladder after complete spinal cord injury. Methods A total of 60 female Sprague-Dawley rats were involved, in which 36 rats were randomly selected to establish the neurogenic bladder model with T10 spinal cord transected. 24 successful models selected were randomly divided into model group (n=12) and electroacupuncture group (n=12). The other 24 rats were also randomly divided into sham group (n=12) and blank group (n=12). The electroacupuncture group was electroacupunctured on Ciliao (BL32), Zhongji (RN3), Sanyinjiao (SP6) and Dazhui (GV14) for seven days. All of them were assessed with urodynamic test, TUNEL method was used to determine the apoptosis rate of the spinal cord, and Western blotting was used to detect the expression of Caspase-9, Cyt-C and Apaf-1 protein. Results Compared with the blank group and the sham group, the maximum bladder capacity and compliance decreased (P<0.01), the urinary bladder pressure and leakage point pressure increased (P<0.05); the apoptosis rate of the spinal cord increased (P<0.001); the expression of Caspase-9, Cyt-C and Apaf-1 protein increased (P<0.05) in the model group and the electroacupuncture group. Compared with the model group, the maximum bladder capacity and compliance increased (P<0.01), the urinary bladder pressure and leakage point pressure decreased (P<0.05); the apoptosis rate of the spinal cord decreased (P<0.05); the expression of Caspase-9, Cyt-C and Apaf-1 protein decreased (P<0.05) in the electroacupuncture group. Conclusion Electroacupuncture on Ciliao, Zhongji, Sanyinjiao and Dazhui could improve the bladder function after spinal cord injury in rats. The mechanisms may be related with the down-regulation of Caspase-9, Cyt-C and Apaf-1 protein to reduce apoptosis.
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Objective To explore the effects of prostaglandin E1 (PGE1) on the expression of Apaf-1 and TLR4 in rats with cerebral ischemia-reperfusion(CIR) injury.Methods 32 healthy adult male Wistar rats were randomly divided into four groups,which were sham operated group (n=8),CIR model group (n=8) and PGE1 pretreated groups (low dose,12 μg · kg-1;high dose,24 μg · kg-1,n =8).Rat model of cerebral ischemia/reperfusion was established by bilateral common carotid artery ligation.The expression of Apaf-1 and TLR4 was detected with immunohistochemical staining method in hippocampus and epencephalon.Results After 20 min of ischemia and reperfusion for 24 h,compared with sham operated group (Apaf-1:hippocampus (0.87±0.78),epencephalon (0.67 ±0.43);TLR4:hippocampus (2.43 ± 1.17),epencephalon (1.97± 1.033)),the number of positive cells of Apaf-1 (hippocampus (11.83± 2.26);epencephalon(5.80±1.30) and TLR4 (hippocampus(16.90±2.86);epencephalon(12.90±2.66)) was increased in CIR model group (P＜0.05).Compared with CIR model group,the positive cell numbers of Apaf-1 (hippocampus:low dose(9.83±2.12),high dose(5.50± 1.17);epencephalon:low dose(4.87± 1.38),high dose(2.73±1.172)) and TLR4 (hippocampus:low dose (11.53± 2.40),high dose (9.13 ± 2.54);epencephalon:low dose (9.07 ± 2.07),high dose (4.47 ± 1.68)) were reduced dose-dependently in PGE 1 pretreatment all group (all P ＜0.05).Conclusion PGE1 can inhibit the expression of Apaf-1 and TLR4 in hippocampus and epencephalon of rat with cerebral ischemia-reperfusion injury.
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Objective To study the effect of celecoxib on learning and memory function,cyclooxygenase(COX-2) and the apoptotic protease-activating factor-1(Apaf-1) protein expression after traumatic brain injury in rat.Methods A total of 72 adult male Wistar rats were equally and randomly divided into the normal control group,sham operation group,trauma group and Celecoxib treatment group.Postoperative 72 h-reperfusion was performed for taking brain specimens.The immunohistochemical method and Western blot were used to respectively detect COX-2 and Apaf-1 protein expression change;the Morris water maze test was adopted to detect the learning and memory function on preoperative 5 d and at postoperative 72 h.Results The COX-2 and Apaf-1 protein expression in the trauma group was significantly higher than that in other groups (P＜0.05),and the protein expression in the treatment group and trauma group was decreased,but still higher than that in the sham operation group and normal group(P＜ 0.05);in the Morris water maze test,the prolongation of escape latency time in the trauma group was maximal among 4 groups (P ＜0.05),but the treatment group had a shorter time compared with the trauma group (P＜0.05).Conclusion Craniocerebral trauma can cause different degrees of learning and memory dysfunction,and COX-2 inhibitor celecoxib can downregulate the expression of COX-2 and Apaf-1 protein,inhibit inflammation reaction and cellular apoptosis,and improve the learning and memory dysfunction after traumatic brain injury.
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Objective To investigate the expression and clinical significance of apoptotic protease activating factor 1 (Apaf-1) and bax in prostate cancer (PCa) and benign prostatic hyperplasia (BPH). Methods Immunohistochemistry was used to detect the expression of Apaf-1 and bax in the tissues from 45 PCa patients and 60 BPH patients. Results The positive rates of Apaf-1 and bax in PCa tissues were 22.22%(10/45) and 20.00 % (9/45), respectively, while those in BPH tissues were 48.33 % (29/60) and 46.67 % (28/60). There was a statistically significant difference in the expressions of Apaf-1 and bax between two groups (P0.05), but they were correlated with the pathological grade and clinical stage of PCa (P< 0.05). The expressions of Apaf-1 and bax in PCa tissues were lower than those in BPH tissues. There was a positive correlation between the expression of Apaf-1 and bax (r=0.535, P<0.01). Conclusion Apaf-1 and bax might be correlated with the carcinogenesis and development of PCa.
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Objective To investigate effects of celecoxib on the expression of cyclooxygenase-2 (COX-2), apoptotic protease activation factor-1 (Apaf-1) and function of mobility in rat model of severe craniocerebral trauma. Methods For?ty-eight adult male Wistar rats were randomly divided by random number table into four groups. Normal group was given no manipulation. Sham group was given scalp incision and sutured. The severe closed craniocerebral injury model was estab?lished via Foda method in rats of injury group. Treatment group was given intraperitoneal injection of celecoxib [ 250 mg/(kg·6 h)] on the basis of injury group. The intraperitoneal injection of same volume of normal saline was given in the other three groups. Samples were taken altogether after 72 hours. Changes of COX-2 and Apaf-1 were detected by immunohistochemis?try and Western blot assay. Ten days after the restoration, six rats were taken from each group for assessing neurological im?pairment scale (NSS). Results The expression levels of COX-2 and Apaf-1 were significantly higher in injury group than those of other groups. The expression levels of COX-2 and Apaf-1 were significantly lower in treatment group than those of injury group but the levels were significantly higher than those of sham group and normal group (P < 0.05). NSS scores showed that rats in treatment group improved mobility compared with that of injury group (P<0.05), but there was difference compared with Sham group and control group (P<0.05). Conclusion Celecoxib, with its specific inhibitoty effect on pro?tein COX-2, can effectively reduce inflammatory reactions lower the expression of Apaf-1 and reduce apoptosis of neurons, improving the prognosis of dysfunction of mobility after craniocerebral injury.
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Objective To study the expressions of apoptotic protease activating factor-1(Apaf-1)and astrocyte elevated gene-1(AEG-1)in colonic carcinoma,and to explore their correlations with the clinical path-ological features. Methods The expressions of Apaf-1 and AEG-1 were detected in 63 colonic carcinoma sam-ples and 30 normal colonic mucosa adjacent to tumor nest by immunohistochemical method,and their correla-tions with clinical features of colonic carcinoma were analyzed. Results The positive expressions of Apaf-1 and AEG-1 in colonic carcinoma were 23. 81%(15 / 63)and 68. 25%(43 / 63),respectively. The positive expre-ssions of Apaf-1 and AEG-1 in normal colonic mucosa were 76. 67%(23 / 30)and 26. 67%(8 / 30),respec-tively. The positive expression rate of AEG-1 was significantly higher in colonic carcinoma than that in normal tissue(χ2 = 14. 192,P = 0. 000). However,the expression of Apaf-1 was signi-ficantly lower in colonic carci-noma than that in normal tissue(χ2 = 23. 497,P = 0. 000). The expression of Apaf-1 was negatively correlated to the expression of AEG-1(r = - 0. 339,P = 0. 007). The expressions of AEG-1 and Apaf-1 were associated with differentiation degree(χ2 = 4. 643,P = 0. 031;χ2 = 12. 034,P = 0. 001)and clinical stage(χ2 = 6. 628, P = 0. 010;χ2 = 8. 246,P = 0. 004),but they were not correlated with age(χ2 = 1. 462,P = 0. 227;χ2 =2. 401,P = 0. 121)and tumor size(χ2 = 0. 333,P = 0. 564;χ2 = 0. 590,P = 0. 442). Conclusion The expression of AEG-1 is up-regulated in colonic carcinoma,but the expression of Apaf-1 is down-regulated,with a significant negative correlation. Apaf-1 and AEG-1 may be closely related to the occurrence and development of colon carcinoma. Therefore,combination detection of Apaf-1 and AEG-1 may be more valuable for the prog-nosis evaluation of colonic carcinoma.
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Objective To investigate the effectiveness of decitabine demethylation in treatment of acute leukemia.Methods Methylation specific PCR (MSP) was used to detected the methylation status of Apaf-1 gene promoter.10 cases entering the group.MSP was used to detected the 10 cases methylation status of Apaf-1 promoter between pre-and post-treatment of dicitabine.RT-PCR method used was to detect the differential expression levels of Apaf-1 mRNA in acute leukemia bone marrow mononuclear cell between preand post-treatment of decitabine.Results In post-treatment of decitabine,6 cases Apaf-1 gene promoter was demethylated.The loss expression of Apaf-1 mRNA re-expressed in 4 cases.6 cases Apaf-1 mRNA still express deletion.6 cases patients have Apaf-1 mRNA exprssion deletion,However,4 cases Apaf-1 gene was demethylated,2 cases methylated in post-treatment,maybe related to allele deletion or allelic varriants.Conclusion Post treatment of decitabine.Apaf-1 gene promotor was demethylated and repress the expression of Apaf-1 mRNA,play a key role in apoptosis maybe a new method for treatment of acute leukemia.
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Objective To investigate the effect of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on mitochondrial pathway of apoptosis in rats with neonatal necrotizing enterocolitis (NEC).Methods Sprague-Dawley neonatal rats were randomly divided into three groups with ten in each.NEC group rats were formula fed,and hypoxia exposed by 100％ N2 for 90 s and cold stress at 4 ℃ for 10 min twice a day for three days.Additionally,rats in HB-EGF group received HB-EGF 800μg/kg by gavage four times a day for three days.Rats in control group were given breast milk feeding for three days without any interventions.Seventy-two hours after born,all neonatal rats were sacrificed after fasting for 12 h,from which the terminal ileum was removed.HE-staining was done for histologic evaluation.Mitochondrial ultrastructure was observed under electron microscopy.Cytochrome C was detected by immunohistochemical analysis and apoptosis inducing factor (AIF) and apoptotic protease activating factor-1 (APAF-1) were measured by Western blot.Analysis of variance and q-test were used to compare the difference among groups.Results (1) The incidence of NEC in HB-EGF group was lower than that in NEC group (2/10 vs 9/10,x2 =7.27,P＜0.01).(2) In NEC group,mitochondria in epithelial cells and muscle cells of intestine were significantly swelling,appearing many electron-lucent zones in matrix.Ultrastructure of mitochondria were severely damaged.In HB-EGF group,mitochondria were less swelling and showed milder damage than those in NEC group.(3) The expression of cytochrome C in ileal tissue in NEC group was higher than that in control group (0.030±0.018 vs 0.002±0.001,q=6.15,P＜0.01).The expression of cytoehrome C in ileal tissue in HB-EGF group was lower than that in NEC group (0.014±0.018 vs 0.030±0.018,q=3.53,P＜0.05).The expression of APAF-1 and AIF in NEC group was higher than those in control group (1.364±0.299 vs 0.215±0.033,q=15.31,P＜0.05;0.181±0.050 vs0.127±0.045,q=3.71,P＜0.05).Compared to NEC group,the expression of APAF-1 was lower (0.455±0.123 vs 1.364±0.299,q=4.04,P＜0.05) and the expression of AIF was higher (0.289±0.045 vs 0.181±0.050,q=7.32,P＜0.05) in HB-EGF group.Conclusions HB-EGF could reduce the incidence of NEC in neonatal rats by inhibiting the mitochondrial pathway related apoptosis through down regulation of APAF-1.
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Objective:To explore the relationship between apoptotic protease activating factor 1(APAF1) gene and oral squamous cell carcinoma (OSCC). Methods:The mRNA expression of APAF1 gene were detected with semiquantitative RT-PCR method in 18 cases of normal oral mucous membrane and 32 cases of OSCC tissues. Results:The expression of APAF1 gene was significantly decreased in OSCC tissues comparing with the normal oral mucous membrane(P