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Objective:To investigate the therapeutic effect of conjoint fascia sheath suspension and levator palpebrae muscle shortening on congenital severe blepharoptosis.Methods:From June 2014 to December 2018, 30 cases (40 eyes) of congenital severe ptosis were treated in Ningbo First Hospital and Shaoxing Women and Children＇s Hospital. All patients were corrected by conjoint fascia sheath suspension and levator palpebrae muscle shortening. Six months after operation, the distance between the middle point of upper eyelid margin and corneal reflex point was measured to evaluate the correction effect of blepharoptosis; the improvement of upper eyelid appearance was evaluated by 5-point Likert scale (LS), and the incidence of complications was counted.Results:Thirty patients (40 eyes) were followed up for 6-12 months. Correction effect showed that preoperative marginal reflex distance (MRD) was (0.10±0.05) and postoperative MRD was (3.80±0.55); the difference was statistically significant ( t=0.95, P<0.05); 38 eyes (95%) were corrected and 2 eyes (5%) were undercorrected; appearance of upper eyelid showed that preoperative LS was (0.50±0.05) and postoperative LS (3.80±0.55); the difference was statistically significant ( t=0.98, P<0.05). Only one case was complicated with keratitis caused by improper early nursing after incomplete closure exposure, and the patient was cured by sealing the eyes combined with drug conservative treatment. Conclusions:Combined conjoint fascial sheath suspension and levator palpebrae shortening in the treatment of severe blepharoptosis has significant effect, with the advantages of simple operation, small surgical trauma, low recurrence rate, low incidence of complications, high predictability of surgical results and high satisfactory rate of patients.
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Objective:To observe the effect of dictyophora polysaccharide (DIP) on PINK1/Parkin pathway mediated mitophagy induced by sodium arsenite (NaAsO 2) in human hepatocytes (L-02 cells). Methods:The L-02 cells in logarithmic growth phase and in good condition were divided into control group, NaAsO 2 group (10 μmol/L), DIP group (80 μg/ml), DIP + NaAsO 2 group (80 μg/ml DIP + 10 μmol/L NaAsO 2) , N-acetylcysteine (NAC) group (5 mmol/L), and NAC + NaAsO 2 group (5 mmol/L NAC + 10 μmol/L NaAsO 2). Western blotting was used to detect the expression levels of mitophagy related proteins p62, microtubule-associated protein 1 light chain 3 (LC3)Ⅱ/LC3Ⅰ, PINK1, and Parkin. The mitochondrial stucture and autophagosomes were observed by transmission electron microscope, the fluorescent probe method was used to detect the expression level of intracellular reactive oxygen species (ROS). Results:Compared with the control group, the protein expressions of p62, LC3 Ⅱ/LC3 Ⅰ, PINK1, and Parkin in NaAsO 2 group were higher ( P < 0.05); compared with the NaAsO 2 group, the protein expressions of p62, LC3 Ⅱ/LC3 Ⅰ, PINK1 and Parkin were lower in DIP, DIP + NaAsO 2, NAC, and NAC + NaAsO 2 groups ( P <0.05). According to the transmission electron microscope, compared with the control group, the mitochondria of L-02 cells in NaAsO 2 group were significantly damaged and the number of autophagosomes increased. Compared with NaAsO 2 group, the degree of mitochondrial swelling, vacuolar degeneration and the number of autophagosomes decreased in DIP + NaAsO 2 group. Compared with the control group (33 110.00 ± 2 191.28), the intracellular ROS level in NaAsO 2 group was higher (48 000.00 ± 2 395.31, P < 0.05); the level of intracellular ROS in DIP + NaAsO 2 group (38 670.00 ± 2 620.56) was significantly lower than that in NaAsO 2 group( P < 0.05), and there was no significant change compared with the control group ( P > 0.05). Conclusions:NaAsO 2 can induce PINK1/Parkin mediated mitophagy in L-02 cells. DIP can alleviate NaAsO 2 induced mitophagy. DIP may affect PINK1/Parkin mediated mitophagy induced by NaAsO 2 through the regulation of ROS.
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Objective:To investigate the effects of sodium arsenite (NaAsO 2) on the expression of sterol regulatory element-binding protein-1c (SREBP-1c), peroxisome proliferator activated receptor α (PPARα) and fatty acid synthase (FAS) in human liver cells (L-02 cells). Methods:L-02 cells were cultured in vitro, and exposed to NaAsO 2 at 0 (control), 2, 4, 8, 16, 32, 64 and 128 μmol/L for 24 h, respectively, and the cell survival rate was determined by CCK-8 method. And a fitting curve was made to calculate the half inhibitory concentration (IC 50), subsequent experiments were carried out with 0, 1/8, 1/4 and 1/2 of IC 50 as arsenic exposure doses. Glycerol phosphate oxidase-catalase (GPO-PAP) method was used to detect the content of triglyceride (TG) in cells; the mRNA expression levels of SREBP-1c, PPARα and FAS were detected by Real-time PCR; and the protein expression levels of SREBP-1c and PPARα were detected by Western blotting. Results:The cell survival rates of 8, 16, 32, 64 and 128 μmol/L NaAsO 2 groups [(92.000 ± 1.414)%, (91.000 ± 0.000)%, (76.500 ± 0.707)%, (53.000 ± 1.412)%, (47.000 ± 1.412)%] were significantly lower than that of the control group [(100.000 ± 0.000)%, P < 0.01]. The IC 50 was 64 μmol/L, and subsequent experiments were conducted with 0 (control), 8, 16 and 32 μmol/L NaAsO 2, respectively. Compared with the control group [(1.000 ± 0.000) mmol/g prot], TG contents of 8, 16 and 32 μmol/L NaAsO 2 groups [(0.691 ± 0.064), (0.474 ± 0.162), (0.184 ± 0.045) mmol/g prot] were significant decreased ( P < 0.01). Compared with the control group, the mRNA expression levels of SREBP-1c, PPARα, FAS, and the protein expression levels of SREBP-1c and PPARα in NaAsO 2 groups were significantly decreased ( P < 0.01 or < 0.05). Correlation analysis showed that NaAsO 2 content was negatively correlated with TG content, SREBP-1c and PPARα protein expression levels ( r =-0.954,- 0.875,-0.965, P < 0.01). Conclusion:NaAsO 2 can reduce the TG content and the expression of lipid metabolism related genes SREBP-1c, PPARα and FAS in L-02 cells, suggesting that arsenic-induced liver injury can cause lipid metabolism disorders.
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Objective:To explore the role of nuclear factor-E2-related factor 2 (Nrf2) signaling pathway in oxidative damage caused by sodium arsenite (NaAsO 2) in human normal liver cells (L-02), and to provide experimental basis for the study of oxidative damage mechanism of liver damage caused by arsenic. Methods:L-02 cells were cultured in vitro and treated with 0 (control), 25, 50, 75, 100, 125, and 150 μmol/L NaAsO 2, respectively, for 24 h. The half-inhibitory concentration (IC 50) was calculated according to the cell survival rate by CCK8, and L-02 cells were treated with 0, 1/8, 1/4 and 1/2 dose of IC 50 of NaAsO 2, respectively, for grouping experiments. Protein expressions of Nrf2, heme oxygenase-1 (HO-1), NADH quinone oxidoreductase 1 (NQO1) and glutathione peroxidase 1 (GPx1) in L-02 cells and L-02 nucleus were detected by Western blotting. Results:The result of CCK8 showed that the survival rates of L-02 cells in 25, 50, 75, 100, 125, 150 μmol/L NaAsO 2 groups were [(69.53 ± 0.06)%, (41.33 ± 0.08)%, (23.65 ± 0.04)%, (26.51 ± 0.04)%, (31.63 ± 0.01)%, (26.24 ± 0.02)%], which were significantly lower than that of the control group[(100 ± 0.00)%]. The differences were statistically significant ( P < 0.05). The IC 50 calculated by cell survival was 40 μmol/L, and the NaAsO 2 doses used in the experiment were 0 (control), 5, 10, and 20 μmol/L. Western blotting results showed that, compared with the control group, the protein expression levels of Nrf2, HO-1 in L-02 and HO-1 in the L-02 cells nucleus in the 5, 10 and 20 μmol/L NaAsO 2 groups were significantly higher ( P < 0.05). Compared with the control group, the expression levels of GPx1 protein in L-02 cells of 10 and 20 μmol/L NaAsO 2 groups were decreased ( P < 0.05). Compared with the control group, the expression levels of Nrf2 protein in L-02 nucleus in 10 and 20 μmol/L NaAsO 2 groups were significantly increased ( P < 0.05); the expression level of NQO1 protein in L-02 nucleus in 5 μmol/L NaAsO 2 group was significantly increased ( P < 0.05). Conclusion:NaAsO 2 has an effect on the expression of Nrf2 signaling pathway related factors in L-02 cells, and the mechanism of oxidative damage caused by NaAsO 2 in L-02 cells may be related to Nrf2 signaling pathway.
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Objective:To explore the mechanism of apoptosis induced by sodium arsenite (NaAsO 2) in human hepatic cells (L-02) through reactive oxygen species (ROS) accumulation and mitochondrial dysfunction, and provide experimental evidence for the mechanism of arsenic poisoning. Methods:L-02 cells were divided into control group, NaAsO 2 group (10 μmol/L NaAsO 2), N-acetylcysteine (NAC) group (5 mmol/L NAC), and NaAsO 2 + NAC group (10 μmol/L NaAsO 2, 5 mmol/L NAC), and were cultured in vitro for 24 h. The intracellular ROS level, mitochondrial membrane potential depolarization ratio and cell apoptosis rate were measured by dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescence probe, JC-1 staining and Annexin V-FITC/PI double staining, respectively; the mRNA and the protein of Caspase 3, cytochrome C (Cyt-C) and cytochrome C oxidaseⅣ (COXⅣ) were detected by real time fluorescence quantitative PCR (qRT-PCR) and Western blotting, respectively. Results:There were statistically significant differences in intracellular ROS levels (3 857 392.33 ± 44 928.39, 4 515 288.00 ± 32 660.64, 3 670 150.67 ± 101 987.69, 4 035 235.67 ± 99 995.30), mitochondrial membrane potential depolarization ratios (2.16 ± 0.54, 7.95 ± 0.52, 2.70 ± 0.29, 1.01 ± 0.23) and total apoptosis rates (1.45 ± 0.03, 4.27 ± 0.17, 1.87 ± 0.12, 2.52 ± 0.35) between groups ( F = 62.62, 159.81, 112.70, P < 0.05). There were statistically significant differences in Caspase 3, Cyt-C, COXⅣ mRNA expression levels ( F = 9.20, 7.33, 14.87, P < 0.05) and in cleaved-Caspase 3, Cyt-C, COXⅣ protein expression levels( F = 31.42, 8.01, 83.30, P < 0.05) between groups. Compared with the control group, the intracellular ROS level, mitochondrial membrane potential depolarization ratio and total apoptosis rate were significantly increased ( P < 0.05); Caspase3, Cyt-C mRNA and protein expression levels were significantly increased ( P < 0.05), and COXⅣ mRNA and cleaved-Caspase 3, Cyt-C protein expression levels were significantly decreased ( P < 0.05) in NaAsO 2 group. Compared with the NaAsO 2 group, the intracellular ROS level, mitochondrial membrane potential depolarization ratio and total apoptosis rate of NaAsO 2 + NAC group were significantly decreased ( P < 0.05); the Caspase3, Cyt-C mRNA and cleaved-Caspase 3, Cyt-C protein expression levels were significantly decreased ( P < 0.05), the COX Ⅳ mRNA and protein expression levels were significantly increased ( P < 0.05). Conclusions:NaAsO 2 stimulates L-02 cells to produce excessive ROS, which induces mitochondrial depolarization and further triggers mitochondrial damage, resulting in increased release of Cyt-C and activation of the mitochondrial apoptosis pathway that Caspase 3 protein induces apoptosis in L-02 cells, which may be one of the main mechanisms of arsenic-induced liver injury.
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Objective@#To observe the predictive value of serial platelet function testing (PFT) on outcome in patients undergoing complex percutaneous coronary intervention (PCI).@*Methods@#Six hundred and two consecutive patients undergoing complex PCI in Anzhen hospital were enrolled during October 2011 to June 2012.Adenosine diphosphate(ADP)-induced platelet aggregation was measured by light transmission aggregometry on the first, sixth and twelfth month after PCI and the mean value was calculated.The cut-off value of high on-treatment platelet reactivity (HTPR) was defined as 40%.The primary endpoint was major adverse cardiovascular and cerebral event (MACCE). Clinical outcomes were analyzed by the Kaplan-Meier method and differences were compared using the log-rank test.Multivariate analyses by Cox proportion hazards regression were applied to identify variables independently associated with the adverse outcomes.@*Results@#Five hundred and eighty-five patients (HTPR, n=285; non-HTPR, n=280) finished the follow-up ((28.47±7.45) months). A total of 33 cases of MACCE were observed during the follow-up, among which 29 cases(8.42%) were in HTPR group and 9 cases (3.21%) in the non-HTPR group.Kaplan-Meier analysis suggested that HTPR was associated with an increased incidence of MACCE (log-rank test, P=0.01). The Cox multivariate analysis indicated that HTPR was an independent risk factor of MACCE (HR=2.69, 95%CI 1.23-5.85, P=0.01) in patients undergoing complex PCI.Incidence of MACCE was similar between HTRP patients receiving standard dual antiplatelet therapy (DAPT) or prolonged DAPT (>12 months).@*Conclusion@#Serial PFT could predict the long-term prognosis of patients underwent complex PCI.
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Objective To explore the relation between puncturing at superficial venules and effectiveness of acupoint application in summer to treat winter disease. Methods 4872 chronic bronchitis patients who received therapy of acupoint application in summer for treating winter diseases were divided into a superficial-venule group (including 994 patients) and a non-superficial-venuls group (including 3878 patients), according to the conditions that if or not they had superficial venuls. Both groups received the treatment of acupoint application combined with plum-blossom needles quick puncture. On this basis, the superficial-venule group was additional treated by puncturing at superficial venules. Results The cure rate and total effective rate were 73.2% and 98.1% in the superficial-venule group, and 35.9% and 90.8% in the non-superficial-venuls group. There was a significant difference between the two groups in their therapeutic effect (P＜0.01).Conclusion The treatment of acupoint application combined with puncturing at superficial venules are effective in treating chronic bronchitis.
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This paper reports a simple method using the mesentery as a naturai mountingsubstrate for the study of mesenteric free cells under the scanning electron micros-cope.Mouse mesentery was cut into pieces about Ⅰ square centimeter in size andadhered on to neutral filter paper.The specimen were fixed in 5% glutaraldehyde and1% osmium tetroxide for 20 minutes,each step washed lightly with pH 7.4 PBS,dehydrated with ascending series of alcohol,put into amyl acetate,critical pointdried with CO_2,coated with gold and examined in scanning electron microscope.The morphologic features of free cells in the peritoneal lumen,such as polymorpho-nuclear leucocytes,macrophages and lymphocytes could be observed satisfactorily.We studied in particular the relationship between immunoreactive cells and targetcells(fetal liver cell)injected into the peritoneal cavity,Polymorphonuclear leuco-cytes could be seen invading into and eroding the target cells and macrophage cau-sing their swelling and lysis.Cooperative action of immunoreactive cells and anincrease in number of milk spots were also observed.