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Objective:To evaluate the effect of TBK1 overexpression on hypoxia-reoxygenation (H/R) injury in isolated mouse cardiomyocytes subjected to high glucose and the relationship with mitochondrial autophagy.Methods:Normally cultured log-phase HL-1 mouse cardiomyocytes were inoculated in a 6-well plate at a density of 1×10 6 cells/ml and were divided into 4 groups ( n=10 each) using a random number table method: control group (group C), high glucose group (group HG), high glucose and H/R group (group HG+ H/R), and TBK1 overexpression group (group TBK1). The cells were incubated in culture medium with 1% fetal bovine serum and 1% double antibody for 24 h when the cell density reached 50%.When the cell density reached 80%, pcDNA3.1 (+ ) was used as a vector to achieve TBK1 overexpression.The cells were cultured with high glucose medium (33 mmol/L) for 24 h, exposed to 94% N 2+ 5% CO 2+ 1% O 2 for 24 h in an incubator at 37℃ followed by 12 h reoxygenation in an incubator containing 5% CO 2 at 37°C to establish the model of H/R injury to cardiomyocytes subjected to high glucose.After reoxygenation, CCK-8 assay was used to detect cell viability, the activity of lactic dehydrogenase (LDH) in supernatant was detected using LDH kit, mitochondrial contents were determined using Mito-Tracter green fluorescent probe, and the expression of TBK1 and mitophagy-related proteins PINK1, Parkin, LC3B and P62 was detected by Western blot. Results:Compared with group C, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, mitochondrial contents were decreased, the expression of TBK1, PINK1, Parkin and LC3B was down-regulated, and the expression of P62 was up-regulated in HG group and HG+ H/R group ( P<0.05). Compared with group HG, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, mitochondrial contents were decreased, the expression of TBK1, PINK1, Parkin and LC3B was down-regulated, and the expression of P62 was up-regulated in group HG+ H/R ( P<0.05). Compared with group HG+ H/R, the the cell viability was significantly increased, the activity of LDH in supernatant was decreased, mitochondrial contents were increased, the expression of TBK1, PINK1, Parkin and LC3B was up-regulated, and the expression of P62 was down-regulated in group TBK1 ( P<0.05). Conclusion:The mechanism by which TBK1 overexpression reduces the H/R injury is related to restoring mitophagy in isolated mouse cardiomyocytes subjected to high glucose.
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Objective To explore the effect of particulate matter (PM 2.5) on airway inflammation in asthmatic mice and the intervention effect of Honokiol.Methods Fifty female BALB/c mice were divided into 5 groups according random number table,group A:normal control group;group B:asthma model group;group C:PM 2.5 low dose exposure asthma group;group D:PM 2.5 high dose exposure asthma group:group E:Honokiol group.Asthmatic mouse models were established by ovalbumin(OVA) sensitization and challenge.On day 0 and 7,B-E groups were intraperitoneally with injection 100 mg/L OVA and Al (OH)3 for sensitization;on day 14 to 21,10 g/L OVA solution was given 30 min per day to challenge.During challenge phrase,C-D groups were received different doze intratracheal injection of PM 2.5 respectively,every 7 days,total 4 times.On this basis,the mice in group E received Honokiol intragastfic administration.The mice in group A were carried out by using saline instead of OVA.Mice were sacrificed 24 h after the final inhalation challenge,and for the recovered bronchoalveolarlavage fluid(BALF) of the left lung was used for differential inflammatory cell counts,HE staining and pathological examination were performed on the right lung.The expression of Toll-like receptor 4 (TLR4) and nuclear factor (NF)-κB at mRNA level were detected by real-time flurescence quantitative polymerase chain reaction (qPCR).Flow cytometry analysis was performed to measure the levels of Th17 and Treg cells.Results Compared with group A,mice in group B,group C and group D expressed more serious disorsers of bronchial epithelial cells,alveolar wall congestion and edema,increased mucus secretion in the airway and infiltration of inflammatory cells in lung,and those in group D was more obvious than those in group C and group E,significantly reduced respiratory inflammation compared with group E[(8.56 ± 3.28) × 108/L,0.041 5 ± 0.013 5],the total number of inflammatory cell counts in group C and group D were (20.28 ± 11.16) × 108/L and (27.38 ± 14.64) × 108/L,eosinophils proportion were 0.177 8 ±0.064 9 and 0.229 1 ±0.098 7,there were statistically significant differences(all P ＜ 0.05);compared with group E (1.60 ± 0.28,1.54 ± 0.25),the expression of TLR4 mRNA and NF-κB mRNA in group C and group D (2.56 ± 0.49,3.21 ± 0.61;2.42 ± 0.30,2.83 ± 0.32) were significantly higher,and there were statistically significant differences(all P ＜0.05),group D was more higher than those in group C (all P ＜ 0.05);compared with group E(0.018 3 ± 0.002 3),the expression of Th17 in group C and group D (0.043 9 ±0.008 9 and 0.052 2 ±0.011 8) were significantly higher,and there were statistically significant differences(all P ＜0.05);compared with group E(0.064 5 ±0.003 8),the expression of Treg in group C and group D (0.038 2 ± 0.004 2) and (0.022 7 ± 0.003 3) were significantly lower,and there were statistically significant differences(all P ＜ 0.05);and those of group E were improved remarkably.Conclusion PM 2.5 exposure can aggravate airway inflammation in asthmatic mice,and the damage to airway is more obvious when exposed to high dose of PM 2.5,Honokiol can relieve PM 2.5 exposure of asthmatic airway inflammation through down regulation the expression of TLR4 and NF-κB and Th17 and regulating the balance of Th17 and Treg cells.
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Objective To investigate the protective effect of Honokiol on the airway inflammation induced by particulate matter 2.5(PM2.5)in the asthmatic mice and its mechanism.Methods Fifty male specific pathogen free (SPF)Balb/c mice were randomly divided into 5 groups.Group A:normal control group;group B:asthmatic model group;group C:PM2.5 exposure asthmatic group;group D:TAK -242 group;group E:Honokiol group. Asthmatic mouse models were established by ovalbumin(OVA)sensitization and challenge.On days 0 and 7,the mice in B-E groups were injected intraperitoneally with injection 100 mg/L OVA and aluminum hydroxide for sensitization;on days 14 to 21,10 g/L OVA solution was given 30 min per day to challenge.During challenge phrase,the mice in C -E groups received intratracheal injection of PM2.5,every other day,4 times totally.On this basis,the mice in group D re-ceived TAK-242 intraperitoneal injection,and the mice in group E received honokiol intragastric administration.Group A was given saline instead of OVA.Animals were sacrificed 24 h after the final inhalation challenge,and the bronchoal-veolar lavage fluid(BALF)of the left lung was used for differential inflammatory cell counts.The expressions of Toll-like receptors 4(TLR4)and nuclear factor(NF)-κB at mRNA level were detected by real-time quantitative PCR. Flow cytometry analysis was performed to measure the levels of Th17 and Treg cells.Results Compared with group A,mice in group B and group C expressed more serious disorders of bronchial epithelial cells,alveolar wall congestion and edema,increased mucus secretion in the airway and infiltration of inflammatory cells in the lung,and those in group C were more obvious than those of group B and group E significantly reduced respiratory inflammation;compared with group A[(4.15 ± 1.35)×108/L,0.012 0 ± 0.002 3],the total number of inflammatory cell counts[(16.79 ± 5.62)×108/L and(24.58 ± 13.46)×108/L],eosinophils proportions(0.113 8 ± 0.022 3 and 0.197 8 ± 0.084 9)in group B and group C,were significantly higher,and the differences were statistically significant(all P<0.05);The total number of inflammatory cell counts and eosinophils proportion in group E(8.56 ± 3.28)×108/L and 0.041 5 ± 0.013 5)were significantly lower than those in group C,and the differences were statistically significant(all P <0.05);The expressions of TLR4 mRNA and NF-κB mRNA in group B and C(1.85 ± 0.56,1.82 ± 0.28 and 2.97 ± 0.41,2.83 ± 0.32)were significantly higher,and the differences were statistically significant(all P <0.05);The expressions of TLR4 mRNA and NF-κB mRNA in group E(1.60 ± 0.28,1.54 ± 0.25)was significantly lower than those in group C,and the differences were statistically significant(all P<0.05);the expressions of Th17 in group B and C[(2.89 ± 0.61)% and(4.96 ± 0.27)%]were significantly higher than those of group A[(1.03 ± 0.35)%] (all P<0.05);The expression of Th17 in group E[(1.83 ± 0.23)%]was significantly lower than that of group C,and the differences were statistically significant(P<0.05);the expressions of Treg in group B and C[(4.96 ± 0.35)%and(2.27 ± 0.41)%]were significantly lower than those of group A[(7.37 ± 0.56)%],and the differences were sta-tistically significant(all P<0.05);The expression of Treg in group E was significantly increased[(6.45 ± 0.38)%] compared with that in group C,and the difference were statistically significant(P<0.05);and those of group D and E were improved remarkably.Conclusions Honokiol can relieve PM2.5 exposure of asthmatic airway inflammation through down-regulating the expression of TLR4 and NF-κB and Th17 and regulating the balance of Th17 and Treg cells.
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Objective To investigate the anti-inflammatory and immunosuppressive effects of honokiol in a mouse model of particulate matter ( PM ) 2.5-induced asthma .Methods Female SPF BALB/c mice were randomly divided into five groups:normal saline group (group A), ovalbumin (OVA)-sensitized group ( group B), PM2.5-exposed+OVA-sensitized group ( group C), dexamethasone-treated group (group D) and honokiol-treated group (group E).All mice except those in group A were sensitized and challenged with OVA, and the mice in groups C, D and E were exposed to PM2.5 every two days since the first challenge.Samples of lung sections were stained with hematoxylin and eosin (HE) to observe in-flammatory infiltration.Bronchoalveolar lavage fluid (BALF) and PBMCs were collected from each mouse . Expression of RORγt and Foxp3 at mRNA level was detected by quantitative real-time PCR.Flow cytometry analysis was performed to measure the percentages of Th 17 and Treg cells.ELISA was performed to measure the levels of IFN-γ, IL-10 and IL-17 in the supernatants of cell culture .Results Compared with group B , group C showed an enhanced expression of RORγt at mRNA level, increased IL-17 level and up-regulated percentage of Th17 cells (all P<0.05), but a suppressed expression of Foxp3 at mRNA level, decreased IL-10 level and down-regulated percentage of Th17 cells (all P<0.05).No significant difference in the per-centage of Th1 cells or in the expression of Th 1-related cytokines was observed .The expression of RORγt at mRNA level, IL-17 level and the percentage of Th 17 cells were decreased in PM2.5-exposed mice upon honokiol intervention (all P<0.05), while the expression of Foxp3 at mRNA level, IL-10 level and the per-centage of Treg cells were increased after honokiol intervention (all P<0.05).Honokiol had similar efficacy to dexamethasone in the treatment of asthma .Conclusion Honokiol can alleviate airway inflammation in mice with PM2.5 exposure-induced asthma through regulating the percentages of Th 17 and Treg cells.
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<p><b>OBJECTIVE</b>To compare the difference in the clinical efficacy on type M prostatitis between the combined therapy of acupuncture and isolated-ginger moxibustion and tamsulosin.</p><p><b>METHODS</b>One hundred and ten patients of type III prostatitis were randomized into an acupuncture and moxibustion group and a tamsulosin group, 55 cases in each one. In the acupuncture and moxibustion group, acupuncture and isolated-ginger moxibustion were adopted. Two groups of acupoints were selected, named (1) Guanyuan (CV 4), Qugu (CV 2) and Sanyinjiao (SP 6); (2) Yaoyangguan (GV 3), Pangguangshu (BL 28) and Zhibian (BL 54). The two groups of points were used by acupuncture alternatively and only one group was selected a day. Isolated-ginger moxibustion was applied to Guanyuan (CV 4) and Zhibian (BL 54), once a day, 10 treatments made one session, and totally 3 sessions were required. In the tamsulosin group, tamsulosin was prescribed for oral administration, 0.2 mg, twice a day for 1 month. The National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) score and expressed prostatic secretion (EPS) score were observed in the patients of the two groups.</p><p><b>RESULTS</b>NIH-CPSI and EPS scores after treatment were all reduced apparently as compared with those before treatment in the two groups (all P < 0.05). The improvements in the acupuncture and moxibustion group were more obvious than those in the tamsulosin group (all P < 0.05). In 3 months follow-up, NIH-CPSI score in the acupuncture and moxibustion group was reduced apparently as compared with the tamsulosin group (P < 0.05). The curative rate and total effective, rate were 20.0% (11/55) and 85.5% (47/55) in the acupuncture and moxibustion group, and were 3.6% (2/55) and 61.8% (34/55) in the tamsulosin group respectively (both P < 0.05).</p><p><b>CONCLUSION</b>The combined therapy of acupuncture and isolated-ginger moxibustion achieves the good effect of relieving the symptoms of type III prostatitis and recovery of EPS, better than those treated with tamsulosin. This combined therapy spresents the better long-term efficacy.</p>
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Adult , Humans , Male , Middle Aged , Young Adult , Acupuncture Points , Acupuncture Therapy , Ginger , Chemistry , Moxibustion , Prostatitis , TherapeuticsABSTRACT
Purpose To investigate the clinical pathological features, pathological diagnosis and differential diagnosis of the non-Wilms’ renal tumors in children. Methods The clinical pathological data and immunohistochemical findings of 15 cases of non-Wilms’ renal tumors were retrospectively analyzed with review of the relevant literature. Results In 7/15 (46. 67%) cases of cystic nephroma ( CN) , microscopically the variable size of cysts were lined by a single layer flattened, cuboidal epithelia, separated by fi-broblastic stroma;in 4/15 (26. 67%) cases of rhabdoid tumor of kidney (RTK), histological characteristics were rhabdoid cells and cytoplasmic inclusions;in 2/15 (13. 33%) cases of clear cell sarcoma (CCSK), the classical pattern of histology demonstrated nests, cords of ovoid epithelioid or spindled clear cells separated by fibrovascular septa, which had a marked ’ chicken wire’ pattern of small blood vessels;in 1/15 (6. 67%) case of congenital mesoblastic nephroma (CMN), the tumor was mainly composed of spindle cells arranged in fascicular pattern, in which fetal renal tissue was seen;in 1/15 (6. 67%) case of granularity renal cell carcinoma, micro-scopically, the cancer cells with eosinophilic granular cytoplasm arranged in nests, cords and tubular pattern. Conclusions Although non-Wilms’ renal tumors is a small part of all kidney tumor, it is difficult to diagnose and distinguish from other tumors before opera-tion. The pathological diagnosis is the most reliable method for differential diagnosis of non-Wilms’ renal tumor.