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Cytochrome c is a type of heme proteins that are widely distributed in living organisms. It consists of heme and apocytochrome c, and has potential applications in bioelectronics, biomedicine and pollutant degradation. However, heterologous overexpression of cytochrome c is still challenging. To date, expression of the cytochrome c from uncultured anaerobic methanotrophic archaea has not been reported, and nothing is known about the function of this cytochrome c. A his tagged cytochrome c was successfully expressed in E. coli by introducing a thrombin at the N-terminus of CytC4 and co-expressing CcmABCDEFGH, which is responsible for the maturation of cytochrome c. Shewanella oneidensis, which naturally has enzymes for cytochrome c maturation, was then used as a host to further increase the expression of CytC4. Indeed, a significantly higher expression of CytC4 was achieved in S. oneidensis when compared with in E. coli. The successful heterologous overexpression of CytC4 will facilitate the exploitation of its physiological functions and biotechnological applications.
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Anaerobiosis , Archaea/metabolism , Cytochromes c/metabolism , Escherichia coli/metabolism , Heme/metabolismABSTRACT
Objective:To investigate the effect of lithium chloride (LiCl) on the gap junctional intercellular communication (GJIC) in human Tenon capsule fibroblasts (HTFs) and its underlying mechanism.Methods:The Tenon capsule tissue of a patient who underwent strabismus surgery in Dezhou People's Hospital in April 2019 was collected and cut into tissue blocks of dimensions 1 mm×1 mm×1 mm.Primary culture and subculture were carried out, and the 4th-generation HTFs were taken for experiment.HTFs were divided into the control group and LiCl treatment group and were cultured with cell medium without or with 80 mmol/L LiCl for another 48 hours according to grouping.The cell scratch and dye labeling technique were used to label the coupling index and evaluate the GJIC function.The expression and localization of Cx43 in HTFs were detected by immunofluorescence staining.The expression levels of Cx43 mRNA and protein were detected by real-time fluorescence quantitative PCR and Western blot, respectively.The study protocol was approved by an Ethics Committee of Dezhou People's Hospital (No.2019-023). Written informed consent was obtained from the subject.Results:The cultured spindle-shaped HTFs grew adhering to the wall showing radial monolayer or vortexlike, and the cytoplasm was vimentin positive.Results of dye tracer experiment of cell scratch showed that the cell coupling index of LiCl treatment group was 9.04±0.53, which was significantly higher than 4.94±0.39 of the control group ( t=-18.79, P<0.01). Immunofluorescence staining showed that the Cx43 fluorescence was dotted in the cell membrane between adjacent cells in the control group, and Cx43 staining was obviously enhanced in the LiCl treatment group.The results of real-time fluorescence quantitative PCR showed that with relative expression level of Cx43 mRNA in the control group set to 1, the relative expression level of Cx43 in the LiCl treatment group was significantly increased to 1.97±0.23, showing a statistical significance between them ( t=-14.426, P<0.01). Western blot showed that the relative expression level of Cx43 protein was 0.871±0.057 in the LiCl treatment group, which was significantly higher than 0.446±0.028 in the control group ( t=-11.682, P<0.01). Conclusions:LiCl can enhance the GJIC function between HTFs by upregulating the expression levels of Cx43 mRNA and protein, suggesting that the enhanced GJIC function by LiCl may be one of the mechanisms of its inhibition on HTFs proliferation.
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Objective:To evaluate the accuracy of mini-fluid challenge test in predicting fluid responsiveness in elderly patients undergoing surgery in prone position.Methods:Forty-eight elderly patients, aged ≥ 65 yr, with body mass index of 18-30 kg/m 2, of American Society of Anesthesiologists physical status Ⅱ or Ⅲ, scheduled for elective spinal surgery, were enrolled.Patients were mechanically ventilated using a volume-controlled mode with a tidal volume of 6 ml/kg of ideal body weight during operation.A radial arterial catheter was inserted and connected to FloTrac/Vigileo system to monitor hemodynamic parameters.At 5 min after prone position, volume expansion was started when the hemodynamics was stable: lactated Ringer′s solution 1 ml/kg was intravenously infused over 1 min, 1 min later lactated Ringer′s solution 1 ml/kg was intravenously infused over 1 min, 1 min later lactated Ringer′s solution 1 ml/kg was intravenously infused over 1 min, and 1 min later operation was started.After tracheal intubation and before prone position (T 1), 5 min of prone position (T 2), 1 min after 1st infusion of 1 ml/kg liquid (T 3), and 1 min after 2nd infusion of 1 ml/kg liquid (T 4) and 1 min after infusion of 3 ml/kg fluid (T 5), heart rate, mean arterial pressure, cardiac output, cardiac index, stroke volume, stroke volume index (SVI), stroke volume variability (SVV), pulse pressure variability (PPV), and changes in SVI induced by rapid infusion of 1, 2 and 5 ml/kg crystalloid (ΔSVI 1 ml/kg, ΔSVI 2 ml/kg, ΔSVI 5 ml/kg) were calculated.Positive fluid challenges were defined as an increase in SVI of 10% or more from baseline, and the patients were divided into responder group (R) and non-responder group (NR). Receiver operating characteristic curves predicting fluid responsiveness were generated for ΔSVI 1 ml/kg, ΔSVI 2 ml/kg, SVV and PPV, and areas under the receiver operating characteristic curves (AUC) were calculated. Results:Thirty patients were enrolled in group R and 18 cases in group NR.The AUC of ΔSVI 1 ml/kg in predicting fluid volume responsiveness was 0.87 with a diagnostic threshold of 7%, a sensitivity of 80%, and a specificity of 83%.The AUC of ΔSVI 2 ml/kg in predicting fluid responsiveness was 0.928 with a diagnostic threshold of 8%, a sensitivity of 78%, and a specificity of 89%.The AUC of SVV and PPV in predicting fluid responsiveness was 0.65 and 0.53, respectively. Conclusion:Mini-fluid challenge test guided by ΔSVI can predict fluid responsiveness in elderly patients undergoing surgery in prone position, and rapid infusion of 2 ml/kg crystalloid provides better accuracy than 1 ml/kg.
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Objective To set up and apply a standardized communication system of adolescents and young adults (AYAs) cancer patients,in order to improve AYAs cancer patients' psychological distress and other negative emotions,as well as promote social support and quality of life of patients.Methods A AYAs cancer patients standardized communication system,suitable for China's national conditions,was preliminarily built.Using randomized controlled trials,a total of 171 subjects,selected from 486 cases of AYAs cancer patients,in the Third Xiangya Hospital of Central South University and Hunan Cancer Hospital from August to September in 2016,were intervened with a set of standardized communication system.The Mental Distress Thermometer (DT),Hospital Anxiety and Depression Scale (HADS),Social Support Rating Scale (SSRS) and Concise Health Status Questionnaire (SF-36) were used as evaluation indexes to observe the psychological distress,emotion,social support and quality of life of the three groups of subjects before intervention,immediately after intervention,1 month after intervention and 3 months after intervention.Results At the follow-up of 3 months after intervention,38 cases were lost,14 cases in communication group (final n =43),11 cases in music therapy group (final n =46),and 13 cases in routine group (final n =44)..There was no significant difference in the evaluation indexes between the communication group,music treatment group and the routine group before intervention (P ＞ 0.05).There were statistically significant differences in scores of psychological pain,anxiety and depression,and social support in the 3 groups,before and immediately after intervention,1 month and 3 months after intervention (P ＜ 0.05).The scores of psychological pain,anxiety and depression,social support and quality of life in the communication group 1 month after intervention were statistically significant compared with those in the music group and the routine group (P ＜ 0.05).Conclusions Compared to music therapy and regular care,the standardized communication system has significant effect on improving the psychological distress of AYAs with cancer,and can also improve their social support level and quality of life.The clinical validation of the standardized communication system can provide reference for psychological rehabilitation of cancer survivors.
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Cancer is the most cause death of among adolescents and young adults (AYAs).Psychological distress caused by cancer affects AYAs' effective coping abilities of disease,physical symptoms and treatment.This paper mainly introduces the related concepts,screening tools and intervention progress of psychological distress of AYAs cancer patients to deepen the understanding of these among clinical professionals and provide reference for implement effective interventions to patients.
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Abstract The Asian gypsy moth (Lymantria dispar) is a serious pest of forest and shade trees in many Asian and some European countries. However, there have been few studies of L. dispar genetic information and comprehensive genetic analyses of this species are needed in order to understand its genetic and metabolic sensitivities, such as the molting mechanism during larval development. In this study, high-throughput sequencing technology was used to sequence the transcriptome of the Asian subspecies of the gyspy moth, after which a comprehensive analysis of chitin metabolism was undertaken. We generated 37,750,380 high-quality reads and assembled them into contigs. A total of 37,098 unigenes were identified, of which 15,901 were annotated in the NCBI non-redundant protein database and 9,613 were annotated in the Swiss-Prot database. We mapped 4,329 unigenes onto 317 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database. Chitin metabolism unigenes were found in the transcriptome and the data indicated that a variety of enzymes was involved in chitin catabolic and biosynthetic pathways.
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Objective To study the predictive value of high-sensitivity C-reactive protein (hs-CRP) to perineonate mortality in early onset severe pre-eclampsia (EOSPE) pregnant women. Methods Seventy-five pregnant women with EOSPE (EOSPE group) and 75 pregnant women with gestational hypertension (control group) were selected. The serum tumor necrosis factor (TNF)-α, interleukin (IL)-2 and hs-CRP levels were detected. The data of perineonate death in EOSPE group were recorded, then the pregnant women were divided into death subgroup and survival subgroup. Results The hs-CRP, TNF-αand IL-2 levels in EOSPE group were significantly higher than those in control group:(7.2 ± 2.1) mg/L vs. (2.5 ± 1.0) mg/L, (28.9 ± 5.3) ng/L vs. (12.4 ± 3.5) ng/L and (134.3 ± 34.6) ng/L vs. (104.2 ± 28.5) ng/L, and there were statistical differences (P0.05). Receiver operating characteristic curve analysis result showed that the best threshold of hs-CRP for predicting the perineonate mortality caused by EOSPE was 7.98 mg/L, and the area under the curve was 0.779 (95％CI 0.637 to 0.922), with a sensitivity of 0.83 and a specificity of 0.68. Conclusions Inflammatory reaction is involved in the pathophysiological process of EOSPE, and hs-CRP is a good index to predict perineonate death in EOSPE pregnant women.
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Objective To explore the changes of serum lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) and the level of lipids in patients with obstructive sleep apnea hypopnea syndrome (OSAHS).Methods A total of 94 candidates with sleep disorders between January-July 2011 at outpatient department were monitored with polysomnography.According to apnea-hypopnea index (AHI),they were divided into mild-to-moderate OSAHS (5 ≤ AHI ≤ 30) (n =27),severe OSAHS (AHI ＞ 30) (n =37) and normal control groups (AHI ＜ 5) (n =30).After polysomnography,their blood samples were obtained to measure the levels of serum LOX-1,triglycerides (TG),cholesterol (TC),high-density lipoprotein (HDL-C) and low-density lipoprotein (LDL-C).Results The serum level of LOX-1 in severe OSAHS group was significantly higher than that in the mild-to-moderate and control groups (P ＜ 0.01).The serum level of LOX-1 in OSAHS patients was positively correlated with AHI and longest apnea time (LAT)(r =0.645 & 0.501 respectively,both P ＜ 0.01) and was negatively correlated with SaO2 (r =-0.647,P ＜0.01).No significant difference existed in serum lipids in all groups (P ＞ 0.05).Conclusions Intermittent hypoxia caused by OSAHS increases the level of LOX-1 to further promote the formation and development of atherosclerotic in patients with OSAHS.The levels of lipid can not effectively predict the severity of lipid metabolism disorder in patients with OSAHS.
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Objective To investigate the changes in serum level of vascular endothelial growth factor (VEGF), nitric oxide (NO), erythropoietin (EPO) and plasma level of carbon monoxide (CO) in patients with obstructive sleep apnea-hypopnea syndrome (OSAHS) and their clinical significance. Methods Sixty-four patients diagnosed as OSAHS by polysomnography (PSG), were divided into two groups according to apnea-hypopnea index (AHI), 20 in mild group [5/h≤AHI≤20/h, with a mean of (12± 5)/hi and 44 in moderate-severe group[AHI > 20/h, with a mean of (63±23)/h]. Blood samples were obtained from all the patients after PSG for measurement of levels of serum VEGF, NO and EPO and plasma CO. Results Levels of serum VEGF [(101±91) ng/L], NO [(10.3±3.3) μmol/L]and plasma CO [(0.56±0. 35) mg/L]in moderate-severe OSAHS group were significantly higher than those in mild group (all P <0.05). There was no significantly difference in serum level of EPO between the two groups (P> 0.05). Levels of serum VEGF, NO and plasma CO in OSAHS patients positively correlated with AHI and percentage of time with percutaneous arterial blood oxygen saturation lower than 0. 9 (SLT90) of all sleep time (all P < 0. 05), but reversely correlated with the lowest arterial blood oxygen saturation (LSaO2) at night (P>0.05). No correlation between EPO and all the indicators was found (P>0.05). Multiple linear regression analysis showed that LSaO2 was an independent risk factor to affect levels of serum VEGF and plasma CO (R2=0.198, P=0.001, and R2=0.210, P=0.000, respectively) and SLT90 was an independent risk factor to affect serum level of NO (R2= 0. 148, P=0.004) in OSAHS patients. Conclusion Hypoxia at night is a main cause leading to increased level of serum VEGF, NO and plasma CO in OSAHS patients.
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Objective To study the relationship between oxidative stress and endothelial cell apoptosis in patients with obstructive sleep apnea-hypoventilation syndrome (OSAHS). Methods Sixth-seven patients definitely diagnosed by potysomnography (PSG) as OSAHS were divided into three groups according to their apnea-hypoventilation index (AHI), 14 in mild group (5 < AHI≤20), 21 in moderate group (20 < AHI≤40) and 32 in severe group (AHI 40). And, 18 healthy persons (AHI <5) were recruited as controls. Blood samples were obtained form all of them after PSG performance for measuring apoptotic endothelial cells (CD146AnnV+) and serum levels of superoxide dismutase (SOD) and malondialdehyde (MDA). Results Serum level of MDA and CD146AnnV+ in moderate and severe OSAHS group were significantly higher than those in control group (all P < 0.05). Serum level of SOD in moderate and severe OSAHS group was significantly lower than that in control group (P < 0.05). CD146AnnV+ correlated positively with AHI, the longest apnea time (LAT) and oxygen desaturation index (ODI) (r = 0.778, 0.609 and 0.689, respectively, all P < 0.05) and correlated reversely with saturation of arterial blood oxygen at night (SaO2min) (r =-0.635, P < 0.01). CD146AAnnV+ correlated positively with serum level of MDA (r = 0. 698, P < 0.01), and correlated reversely with serum level of SOD (r =-0.705, P < 0.01). Results of linear multivariate regression analysis showed that AHI, serum levels of SOD and MDA were independent risk factors for endothelial cells apoptosis in patients with OSAHS. Conclusions There existed oxidative stress due to intermittent hypoxia in patients with OSAHS, which could be one of the major causes in exacerbating endothelial damage.