OBJECTIVE: To investigate the role and mechanism of the cyclic guanosine monophosphate-adenosine monophosphate synthase/stimulator of interferon gene (cGAS/STING) pathway in oleic acid-induced acute lung injury (ALI) in mice. METHODS: Male wild-type C57BL/6J mice were randomly divided into five groups (each n = 10): normal control group, ALI model group, and 5, 50, 500 μg/kg inhibitor pretreatment groups. The ALI model was established by tail vein injection of oleic acid (7 mL/kg), while the normal control group received no intervention. The inhibitor pretreatment groups were intraperitoneally injected with the corresponding doses of cGAS inhibitor RU.521 respectively 1 hour before modeling. At 24 hours post-modeling, blood was collected, and mice were sacrificed. Lung tissue pathological changes were observed under light microscopy after hematoxylin-eosin (HE) staining, and pathological scores were assessed. Western blotting was used to detect the protein expressions of cGAS, STING, phosphorylated TANK-binding kinase 1 (p-TBK1), phosphorylated interferon regulatory factor 3 (p-IRF3), and phosphorylated nuclear factor-κB p65 (p-NF-κB p65) in lung tissue. Immunohistochemistry was performed to observe STING and p-NF-κB positive expressions in lung tissue. Serum interferon-β (IFN-β) levels were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Compared with the normal control group, the ALI model group exhibited significant focal alveolar thickening, intra-alveolar hemorrhage, pulmonary capillary congestion, and neutrophil infiltration in the pulmonary interstitium and alveoli, along with markedly increased pathological scores (10.33±0.58 vs. 1.33±0.58, P < 0.05). Protein expressions of cGAS, STING, p-TBK1, p-IRF3, and p-NF-κB p65 in lung tissue significantly increased [cGAS protein (cGAS/β-actin): 1.24±0.02 vs. 0.56±0.02, STING protein (STING/β-actin): 1.27±0.01 vs. 0.55±0.01, p-TBK1 protin (p-TBK1/β-actin): 1.34±0.03 vs. 0.22±0.01, p-IRF3 protein (p-IRF3/β-actin): 1.23±0.02 vs. 0.36±0.01, p-NF-κB p65 protein (p-NF-κB p65/β-actin): 1.30±0.02 vs. 0.53±0.02, all P < 0.05], positive expressions of STING and p-NF-κB in lung tissue were significantly elevated [STING (A value): 0.51±0.03 vs. 0.30±0.07, p-NF-κB (A value): 0.57±0.05 vs. 0.31±0.03, both P < 0.05], and serum IFN-β levels were also significantly higher (ng/L: 256.02±3.84 vs. 64.15±1.17, P < 0.05). The cGAS inhibitor pretreatment groups showed restored alveolar structural integrity, reduced inflammatory cell infiltration, and decreased hemorrhage area, along with dose-dependent lower pathological scores as well as the protein expressions of cGAS, STING, p-TBK1, p-IRF3 and p-NF-κB p65 in lung tissue, with significant differences between the 500 μg/kg inhibitor group and ALI model group [pathological score: 2.67±0.58 vs. 10.33±0.58, cGAS protein (cGAS/β-actin): 0.56±0.03 vs. 1.24±0.02, STING protein (STING/β-actin): 0.67±0.03 vs. 1.27±0.01, p-TBK1 protein (p-TBK1/β-actin): 0.28±0.01 vs. 1.34±0.03, p-IRF3 protein (p-IRF3/β-actin): 0.32±0.01 vs. 1.23±0.02, p-NF-κB p65 protein (p-NF-κB p65/β-actin): 0.63±0.01 vs. 1.30±0.02, all P < 0.05]. Compared with the ALI model group, positive expressions of STING and p-NF-κB in lung tissue were significantly reduced in the 500 μg/kg inhibitor group [STING (A value): 0.40±0.01 vs. 0.51±0.03, p-NF-κB (A value): 0.43±0.02 vs. 0.57±0.05, both P < 0.05], and serum IFN-β levels were also markedly reduced (ng/L: 150.03±6.19 vs. 256.02±3.84, P < 0.05). CONCLUSIONS The cGAS/STING pathway is activated in oleic acid-induced ALI, leading to exacerbated inflammatory responses and increased lung damage. RU.521 can inhibit cGAS, thereby down-regulating the expression of pathway proteins and cytokines, and providing protection to lung tissue.
Animals ; Acute Lung Injury/chemically induced* ; Male ; Nucleotidyltransferases/metabolism* ; Mice ; Signal Transduction ; Mice, Inbred C57BL ; Membrane Proteins/metabolism* ; Oleic Acid/adverse effects* ; Transcription Factor RelA/metabolism* ; Lung/pathology* ; Interferon Regulatory Factor-3/metabolism* ; Disease Models, Animal

AIM:To explore the effects of CD38 on lysosome reformation and cholesterol efflux in macro-phages.METHODS:Bone marrow-derived macrophages from low-density lipoprotein(LDL)receptor knockout(LDLr-/-)mice were cultured as cell model.Live cell imaging system was applied to evaluate the effect of nicotinic acid adenine di-nucleotide phosphate(NAADP)on lysosome number.ELISA was conducted to measure NAADP level in macrophages.After the cells were treated with nicotinic acid(NA),RT-qPCR was conducted to detect CD38 mRNA expression,and Western blot was conducted to observe CD38 protein expression and phosphorylated transcription factor EB(TFEB)level.Laser scanning confocal microscopy was applied to evaluate the influence of CD38/NAADP signaling on lysosome number and cholesterol egression.RESULTS:NAADP remarkably increased lysosome number(P<0.05),and this effect was significantly inhibited by NAADP antagonist NED-19,Ca2+ chelator BAPTA,and calcineurin inhibitor CsA(P<0.05).CD38 markedly enhanced NAADP synthesis in macrophages(P<0.05).NAADP synthetic substrate NA prominently ele-vated the expression of CD38 mRNA and protein(P<0.05).NA significantly decreased the phosphorylated TFEB level;this effect was also attenuated by NED-19,BAPTA and CsA(P<0.05).Disrupting CD38/NAADP signaling pathway markedly inhibited NA-induced enhancement of lysosome number,lysosomal free cholesterol and cytosol cholesterol ester efflux in macrophages(P<0.05).NA-induced enhancement of lysosome number,lysosomal free cholesterol and cytosol cholesterol ester efflux abolished in LDLr/CD38 DKO macrophages(P<0.05),whereas these effects induced by NA were recovered after CD38 gene rescue.CONCLUSION:CD38 triggers lysosome reformation via TFEB and consequently pro-motes the efflux of lysosomal free cholesterol and cytosol cholesterol ester.

Objective:To explore the risk factors of intensive care unit-acquired weakness (ICU-AW) and the characteristics of Medical Research Council (MRC) score and electromyogram.Methods:A case control study was conducted. Patients with mechanical ventilation ≥ 7 days and MRC score admitted to department of respiratory and critical care medicine of China-Japan Friendship Hospital from September 2018 to January 2020 were enrolled, and they were divided into ICU-AW group (MRC score < 48) and non-ICU-AW group (MRC score ≥ 48) according to MRC score. The general situation, past medical history, related risk factors, MRC score, respiratory support mode, laboratory examination results, electromyogram examination results, ICU-AW related treatment, outcome and length of ICU stay were collected, and the differences between the two groups were compared. The risk factors of ICU-AW were analyzed by binary multivariate Logistic regression, and the characteristics of MRC score and electromyogram were analyzed.Results:A total of 60 patients were enrolled in the analysis, including 17 patients in ICU-AW group and 43 patients in non-ICU-AW group. Univariate analysis showed that there were significant differences in acute physiology and chronic health evaluation Ⅱ (APACHEⅡ) score, sequential organ failure assessment (SOFA) score, brain natriuretic peptide (BNP), blood urea nitrogen (BUN) on the first day of ICU admission and the ratio of invasive mechanical ventilation between ICU-AW group and non-ICU-AW group [APACHEⅡ score: 21 (18, 25) vs. 18 (15, 22), SOFA score: 7 (5, 12) vs. 5 (3, 8), BNP (ng/L): 364.3 (210.1, 551.2) vs. 160.1 (66.8, 357.8), BUN (mmol/L): 9.9 (6.2, 17.0) vs. 6.0 (4.8, 9.8), invasive mechanical ventilation ratio: 88.2% vs. 46.5%, all P < 0.05]. Binary multivariate Logistic regression analysis showed no independent risk factor for ICU-AW. The average MRC score of 17 ICU-AW patients was 33±11. The limb weakness was symmetrical, and the proximal limb weakness was the main manifestation. Electromyography examination showed that the results of nerve conduction examination in ICU-AW patients mainly revealed that the amplitude of compound muscle action potential (CMAP) and sensory nerve action potentials (SNAP) were decreased, and the conduction velocity was slowed down; needle electromyography showed increased area of motor unit potential (MUP), prolonged time limit and a large number of spontaneous potentials. Prognosis evaluation showed that compared with non-ICU-AW group, patients in ICU-AW group underwent more tracheotomy (70.6% vs. 11.6%), longer length of ICU stay (days: 57±52 vs. 16±8), and more rehabilitation treatment (58.8% vs. 14.0%), and the differences were statistically significant (all P < 0.01). Conclusions:The occurrence of ICU-AW may be related to high APACHEⅡ score and SOFA score, high levels of BNP and BUN on the first day of ICU admission and the proportion of invasive mechanical ventilation, but the above factors are not independent risk factors for ICU-AW. The MRC score of ICU-AW patients was characterized by symmetrical limb weakness, mainly proximal limb weakness; in electromyography examination, the nerve conduction examination results mainly showed that CMAP and SNAP amplitude were decreased, and conduction velocity was slowed down; needle electromyography examination showed increased MUP area, prolonged duration and a large number of spontaneous potentials.

Extracorporeal membrane oxygenation (ECMO) is a viable ultimate support therapy for patients with severe cardiorespiratory failure. Antimicrobial agents are commonly prescribed in critically ill patients with ECMO. However, an increasing number of studies have shown that ECMO circuit is associated with significant pharmacokinetic (PK) alterations, including the increased volume of distribution and reduced the clearance. In addition, the critical illness pathophysiology can also influence PK of antimicrobial, such as systemic inflammation, excessive fluid resuscitation, hypo-albuminemia, worsening hepatic or renal function. These PK alterations may increase the possibility of therapeutic failure or toxicity. Therefore, this study reviews published studies of the effects of ECMO on PK of antimicrobial agents in adults and makes preliminary recommendations on possible dosing regimen.

Rv2742 is a novel gene identified from Mycobacterium tuberculosis H37Rv by the proteogenomics strategy. The aim of this study was to establish a system of soluble expression and purification of the missing protein Rv2742 in M. tuberculosis H37Rv, to provide reference for further research on the biological function of Rv2742. The soluble protein was not successfully induced by prokaryotic expression vectors pGEX-4T-2-Rv2742, pET-32a-Rv2742, pET-28a-Rv2742 and pMAL-c2X-Rv2742. After the codon of novel gene Rv2742 was optimized according to E. coli codon usage frequency, only the recombinant strain containing plasmid pMAL-c2X-Rv2742 could produce soluble products of Rv2742 encoding gene. In addition, the expression effects of the desired fusion protein were also analyzed under different conditions including hosts, culture temperatures and IPTG concentrations. The optimum expression conditions were as follows: Rosetta (DE3) host, 16 °C culture temperature and 0.5 mmol/L IPTG. After being purified by affinity chromatography with amylose resin, the fusion protein sequence was confirmed by LC-MS/MS. These results indicated that the novel gene Rv2742 product could be successfully induced and expressed in a soluble form by the expression system pMAL-c2X with MBP tag. Our findings provide reference for studies on potential interaction and immunogenicity.
Chromatography, Liquid ; Cloning, Molecular ; Escherichia coli ; Mycobacterium tuberculosis ; genetics ; Recombinant Fusion Proteins ; Tandem Mass Spectrometry

Objective To investigate the problems during using, standard and quality management of the POCT glucose meters in hospital, to analyze the solutions, and to provide reference data for improving the test level of POCT in hospital.Methods The amount, brand and application of portable glucose meters in the hospital were obtained by 3 rounds of surveillance from May to July in 2013.All of those glucose meters were taken part in external quality assessment of Clinical Laboratory Center of National Health and Family Planning Commission.The test results of those glucose meters were compared with that of automatic biochemical analyzer, the comparison results were then analyzed.Results The POCT glucose meters possessed 5 brands in our hospital, and the amount and type of glucose meters in some clinical departments were often changed.When 4 brands which were detected as quality control samples by ministry of health, the accuracy of detection results of 3 concentration of brand Ⅲ were substandard, the CV% of two levels were 11.9%and 10.1%respectively, the remaining 3 brands were in line with the requirements.The qualified percentages of 3 times of comparison were 85.0%, 92.0%and 97.4%.Conclusions The hospital should select the brand of portable glucose meters reasonably, correct use of glucose meters, and it is very necessary to build indoor and interstitial quality evaluation system, at the same time, suggesting the hospital to establish the POCT quality management team, to carry out the instrument comparison periodically, so to guarantee the accurate, reliable results of POCT in hospital.

Objective To evaluate the effect of single operation video intubationscope assisted by disposable mouthpiece in orotracheal intubation. Methods 100 patients undergoing general anesthesia were randomly divided into two groups with 50 cases in each: mouthpiece group (group M): single operation video intubationscope assisted by disposable mouthpiece in orotracheal intubation was performed; control group (group C): Video intubationscope oral intubation assisted by helper were applied. The BP, MAP, HR and SpO2 in the two groups were recorded during intubation. The success rate of intubation, duration of glottis exposure, duration of intubation and complications were recorded. Results Oral-tracheal intubation with video intubationscope were successfully completed for all 100 pa-tients, SpO2 during intubation in two groups was maintained above 95.0%, there was no significant hemodynamic changes in two groups. There were no significant difference in the one-time success rate of intubation, duration of glottis exposure and duration of intubation between group M and group C [92.0%vs 88.0%, (13.0 ±7.0) vs (14.0 ±8.0), (20.0 ± 10.0) vs (21.0 ± 11.0), > 0.05]. No significant complications were reported. Conclusion Compared with video intubationscope oral intubation assisted by helper, single operation video intubationscope assisted by dis-posable mouthpiece in orotracheal intubation also is feasible and effective without needing assistant, it is a simple and convenient technology worthy of application.

Objective To investigate the trend of liver function changes in untreated adult patients with Graves’ disease in China, and to analyze the associated factors. Methods Patients with newly diagnosed as well as recurrent Graves’ disease from January 2006 to August 2014 were enrolled. They were over 18 years old and did not receive any treatments, Examination of liver function, thyroid function, and thyroid related antibodies as well as tests regarding virus hepatitis were performed. Results A total of 1 254 patients were enrolled. 33 patients with virus hepatitis were ruled out. Ultimately, 1 221 patients matched the criteria of our trial, with 347 males and 874 females [(39. 3 ± 9. 5) year old]. After inclusion, they were assigned to 2 groups according to their liver function results(605 in normal group and 616 in abnormal group). Compared to normal group, patients in the abnormal group were older [(40. 1 ± 9. 2 vs 38. 5 ± 8. 7) year old, P<0. 05] and with higher proportion of females(81. 8% vs 61. 2% , P<0. 05). Regarding the thyroid function and related antibody tests, some patients yielded results that were extremely high so as to exceed the upper limit of the normal range. These patients were more frequently seen in the group with abnormal liver function. The patients whose thyroid function parameters exceeded the upper limit had higher level of alanine aminotransferase[ALT,(37. 69 ± 7. 51 vs 31. 90 ± 5. 95) U/ L, P<0. 05], aspartate aminotransferase[AST, (31. 97 ± 5. 09 vs 27. 88 ± 3. 82) U/ L, P<0. 05], direct bilirubin[DBiL, (5. 58 ± 0. 77 vs 4. 54 ± 0. 71) μmol/ L, P<0. 05]than the group whose thyroid function on the detected range. In the patients with all results detected, patients in abnormal liver function group had higher level of triiodthyronine[T3 , (5. 42 ± 0. 29 vs 4. 94 ± 0. 33) nmol/ L, P<0. 05], thyroxin[T4 ,(217. 53 ±14. 32 vs 204. 22 ±13. 54) nmol/ L, P<0. 05], free triiodthyronine[FT3 ,(15. 88 ± 2. 86 vs 14. 48 ±4. 83) pmol/ L, P<0. 05], free thyroxin[FT4 ,(48. 91 ±8. 45 vs 42. 95 ±6. 14) pmol/ L, P<0. 05], thyroid peroxidase antibody[ TPOAb, (402. 75 ± 89. 99 vs 210. 70 ± 44. 63) IU/ ml, P < 0. 05] and thyrotrophin receptor antibody[TRAb,(14. 08 ± 5. 24 vs 9. 04 ± 2. 58) IU/ L, P<0. 05]. Multivariate logistic regression analysis revealed that patients’ age(OR=0. 98, 95% CI 0. 97-0. 99), gender(OR=0. 94, 95% CI 0. 91-0. 97), level of FT4 (OR=3. 08, 95% CI 2. 19-4. 32), TPOAb(OR = 0. 98, 95% CI 0. 97-0. 99), and TRAb(OR = 1. 07, 95% CI 1. 01-1. 12) were independent risk factors of their liver dysfunction. Conclusion Graves’ disease may lead to liver dysfunction, which is much more common and severe in elder and female patients, as well as patients who are suffering from hyperthyroidism and raised level of thyroid related antibodies.

ObjectiveTo investigate the feasibility and safety of Proseal-laryngeal mask airway(PLMA) for anesthesia in endoscopic thyroidectomy.MethodsForty ASA class Ⅰ or Ⅱ patients underwent endoscopic thyroidectomy via breast approach were randomly divided into laryngeal mask venting group( Group P,n =20) and endotracheal tube venting group( Group T,n =20).Both groups were used the same anesthesia induction,maintenance and breathing parameter adjustment.The HR and MAP in both groups were recorded respectively before intubation,while intubation,2 min after intubation,before and after head back out center position,before extubation and 2 min after exbutation.The adverse effects of respiratory tract were recorded respectively.ResultsThere were no significant changes in HR and MAP in Group P before and after intubation,as well as before and after extubation( all P ＞ 0.05 ) ;In Group T,the HR and MAP while intubation,2 min after intubation are higher than that before intubation (all P ＞ 0.05).The HR and MAP in Group T,2 min after exbutation are higher than before extubation(all P ＞ 0.05 ).The incidence rate of sore throat and hoarse postoperatively were lower in Group P than in Group T( all P ＞ 0.05 ).ConclusionPLMA could be safely and effectively used in the anesthesia of endoscopic thyroidectomy.