Relationship between the initial serum ammonia level and prognosis in critically ill patients with non-hepatic disease: a retrospective cohort study based on eICU Collaborative Research Database / 西安交通大学学报(医学版)

【Objective】 To investigate the relationship between the initial serum ammonia level and the risk of ICU and hospital mortalities in critically ill patients without hepatic disease. 【Methods】 A retrospective cohort study was conducted among patients admitted to the eICU Collaborative Research Database (eICU-CRD) for a single admission who had serum ammonia test records within 48 hours of the first ICU admission and had no hepatic disease. The age, sex, ethnicity, Acute Physiologic and Chronic Health Evaluation Ⅳ score (APACHE Ⅳ score), treatment methods, complications, and outcomes were extracted. Univariable and multivariable Logistic regression were used to analyze the relationship between serum ammonia level and the risk of mortality. Interactions were used to analyze whether the relationship between serum ammonia level and the risk of mortality differed in subgroups of APACHE Ⅳ scores, age, sex, and ethnicity; subgroup analyses were made. 【Results】 A total of 1 674 patients were included. The multivariable Logistic regression showed that for every 10 μg/dL increase in ammonia, the risk of ICU death increased by 6.9% (OR=1.069, 95% CI: 1.036-1.104), and the risk of hospital death increased by 4.6% (OR=1.046, 95% CI: 1.017-1.076). The risk of ICU death was 1.7 times greater in patients with initial ammonia level of 49-82 μg/dL than in those with <49 μg/dL (OR=1.700, 95% CI: 1.165-2.482), the risk of ICU death was 2.862 times greater in patients with a level of ≥82 μg/dL compared to those with <49 μg/dL (OR=2.862, 95% CI: 1.792-4.570), and the risk of hospital death was 1.844 times higher in the ≥82 μg/dL group than in the <49 μg/dL group (OR=1.844, 95% CI: 1.213-2.804). There were no significant differences between initial ammonia level and the risk of mortalities in different subgroups of APACHEⅣ scores, age, sex, or ethnicity. 【Conclusion】 In critically ill patients without hepatic disease, elevated initial serum ammonia level after ICU admission is associated with a high risk of ICU and hospital mortality.

Role of Nrf2/GPX4 mediated ferroptosis in intestinal injury in sepsis / 中华危重病急救医学

OBJECTIVE@#To investigate whether ferroptosis exists in sepsis induced intestinal injury, and to verify the association between ferroptosis in sepsis induced intestinal injury and intestinal inflammation and barrier function by stimulating and inhibiting the nuclear factor E2-related factor 2/glutathione peroxidase 4 (Nrf2/GPX4) pathway.@*METHODS@#Forty-eight SPF grade male Sprague-Darvley (SD) rats with a body weight of 220-250 g were divided into sham operation group (Sham group), sepsis group (CLP group), sepsis+iron chelating agent deferoxamine (DFO) group (CLP+DFO group) and sepsis+ferroptosis inducer Erastin group (CLP+Erastin group) using a random number table method, with 12 rats in each group. The sepsis model was established by cecal ligation and puncture (CLP). The Sham group was only performed with abdominal opening and closing operations. After modeling, the CLP+DFO group received subcutaneous injection of 20 mg/kg of DFO, the CLP+Erastin group was intraperitoneally injected with 20 mg/kg of Erastin. Each group received subcutaneous injection of 50 mg/kg physiological saline for fluid resuscitation after surgery, and the survival status of the rats was observed 24 hours after surgery. At 24 hours after model establishment, 6 rats in each group were selected. First, live small intestine tissue was taken for observation of mitochondrial morphology in smooth muscle cells under transmission electron microscopy and determination of reactive oxygen species (ROS). Then, blood was collected from the abdominal aorta and euthanized. The remaining 6 rats were sacrificed after completing blood collection from the abdominal aorta, and then small intestine tissue was taken. Western blotting was used to detect the expression of intestinal injury markers such as Claudin-1 and ferroptosis related proteins GPX4 and Nrf2. Observe the pathological changes of small intestine tissue using hematoxylin-eosin (HE) staining and complete Chiu score; Detection of tumor necrosis factor-α (TNF-α), interleukins (IL-1β, IL-6) levels in serum using enzyme-linked immunosorbent assay (ELISA). The levels of serum iron ions (Fe3+), malondialdehyde (MDA), and D-lactate dehydrogenase (D-LDH) were measured.@*RESULTS@#(1) Compared with the Sham group, the 24-hour survival rate of rats in the CLP group and CLP+Erastin group significantly decreased (66.7%, 50.0% vs. 100%, both P < 0.05), while there was no significant difference in the CLP+DFO group (83.3% vs. 100%, P = 0.25). (2) Western blotting results showed that compared with the Sham group, the expressions of GPX4 and Claudin-1 in the small intestine tissue of the CLP group, CLP+DFO group, and CLP+Erastin group decreased significantly, while the expression of Nrf2 increased significantly (GPX4/β-actin: 0.56±0.02, 1.03±0.01, 0.32±0.01 vs. 1.57±0.01, Claudin-1/β-actin: 0.60±0.04, 0.96±0.07, 0.41±0.01 vs. 1.40±0.01, Nrf2/β-actin: 0.88±0.02, 0.72±0.01, 1.14±0.01 vs. 0.43±0.02, all P < 0.05). Compared with the CLP group, the expressions of GPX4 and Claudin-1 were significantly increased in the CLP+DFO group, while the expression of Nrf2 was significantly reduced. In the CLP+Erastin group, the expressions of GPX4 and Claudin-1 further decreased, while the expression of Nrf2 further increased (all P < 0.05). (3) Under the light microscope, compared with the Sham group, the CLP group, CLP+DFO group, and CLP+Erastin group showed structural disorder in the small intestinal mucosa and submucosal tissue, significant infiltration of inflammatory cells, and destruction of glandular and villous structures. The Chui score was significantly higher (3.25±0.46, 2.00±0.82, 4.50±0.55 vs. 1.25±0.45, all P < 0.05). (4) Under transmission electron microscopy, compared with the Sham group, the mitochondria in the other three groups of small intestinal smooth muscle cells showed varying degrees of volume reduction, increased membrane density, and reduced or disappeared cristae. The CLP+Erastin group showed the most significant changes, while the CLP+DFO group showed only slight changes in mitochondrial morphology. (5) Compared to the Sham group, the CLP group, CLP+DFO group, and CLP+Erastin group had serum levels of TNF-α, IL-1β, IL-6, MDA, D-LDH, and ROS in small intestine tissue were significantly increased, while the serum Fe3+ content was significantly reduced [TNF-α (ng/L): 21.49±1.41, 17.24±1.00, 28.66±2.72 vs. 14.17±1.24; IL-1β (ng/L): 108.40±3.09, 43.19±8.75, 145.70±11.00 vs. 24.50±5.55; IL-6 (ng/L): 112.50±9.76, 45.90±6.52, 151.80±9.38 vs. 12.89±6.11; MDA (μmol/L): 5.61±0.49, 3.89±0.28, 8.56±1.17 vs. 1.86±0.41; D-LDH (kU/L): 39.39±3.22, 25.38±2.34, 53.29±10.53 vs. 10.79±0.52; ROS (fluorescence intensity): 90 712±6 436, 73 278±4 775, 110 913±9 287 vs. 54 318±2 226; Fe3+ (μmol/L): 22.19±1.34, 34.05±1.94, 12.99±1.08 vs. 51.74±11.07; all P < 0.05]. Compared with CLP group, the levels of TNF-α, IL-1β, IL-6, MDA, D-LDH and ROS in CLP+Erastin group were further increased, and the content of Fe3+ was further decreased, the CLP+DFO group was the opposite (all P < 0.05).@*CONCLUSIONS@#Ferroptosis exists in the intestinal injury of septic rats, and stimulating or inhibiting ferroptosis through the Nrf2/GPX4 pathway can effectively intervene in the inflammatory state and intestinal mechanical barrier of the body.

Comparison of three kinds of nutritional screening tools in clinical application of general surgery patients application / 中国现代普通外科进展

Objective:To compare the applicability and consistency of NRS2002,SGA and MNA in general surgery hospitalized patients and analyze the effect in clinical outcomes of NRS resuits with respect to each tool.Method:The 150 patients hospitalized in our hospital were chosen as object of study,to screen and evaluate nutritional risk of patients by NRS 2002,SGA and MNA,respectively,on the second hospital day,then to compare the consistency of NRS results with respect to the four tools on clinical outcomes.Results:The applicabilities of hospital NRS2002,SGA and MNA were alternatively 97.25,97.25％ and 98.35％.The evaluation of patients' NRS corresponding to different four tools was consistent.The effects of screening results of MNA,NRS2002 and SGA on clinical outcomes were most closely related.Conclusion:The three nutritional evaluation tools can be applied to the screening of malnutrition in General Surgery.The effects of screening results of NRS2002 and SGA on clinical outcomes were most closely related.

Effects of ERK1/2 on pressure overload-induced cardiomyopathy and heart failure in mice / 中国病理生理杂志

AIM: To explore the changes in extracellular regulated protein kinase (ERK1/2) in the hypertrophic myocardium induced by pressure overload at the different time courses and to determine the molecular mechanism in the myocardium from hypertrophy to heart failure. METHODS: C57/BL mice, aged 12 week old, were subjected to sham-operation (SH) or transversing aortic constriction (TAC) to establish left ventricular hypertrophy. Echocardiographic assessments, hemodynamic determination, organ weight measurement, morphological and histological examination were performed at 1, 4, 8, 12 and 16 weeks after surgery. Meanwhile mRNA levels of atrial natriuretic peptide (ANP), α-myosin heavy chain (α-MHC), bcl-2 and bax were measured by RT-PCR, and ERK1/2 levels were detected by Western blotting. The animals in SH group were performed the same tests then sacrificed at 16 weeks. RESULTS: (1) Compared to SH group, LVESd, LVEDd, Awsth, Awdth, Pwsth and Pwdth progressively increased after TAC. Meanwhile, ejection fraction (EF%) significantly decreased at 16th week (P<0.05). LVSP, dp/dt_(max) and dp/dt_(min) in TAC group were progressively increased after 4 weeks. From 8-12 weeks these parameters maintained stable and then sharply decreased at 16th week (all P<0.05). However, LVEDP was statistically increased at 8th week. These echocardiographic and hemodynamic changes indicated a development of LVH and eventually progressing towards to heart failure. (2) Histologically, cardiac collagen measured by percentage of Sirius red positive stained area and apoptosis index showed progressive increases from 4 to 16 weeks. (3) Compared to SH group, mRNA levels of ANP was time-dependently increased while α-MHC and Bcl-2 were time-dependently decreased. The ratio of Bcl-2 /Bax was decreased. Phosphorylation of ERK1/2 was increased at 4th week, then decreased with age of TAC (all P<0.05). CONCLUSION: Pressure-overload induced by TAC results in a development of LVH from early concentric hypertrophy to late eccentric hypertrophy, and eventually toward cardiac dysfunction or heart failure. Those changes are associated with increase in cell size and cardiac fibrosis. ERK1/2 signaling pathway may involve in the regulation of myocardial cell apoptosis in hypertrophic and failure heart.

Effects of aging on ventricular remodeling and cardiac rupture after acute myocardial infarction in mice / 中华老年医学杂志

Objective To explore the effects of aging on ventricular remodeling and cardiac rupture after acute myocardial infarction in mice. Methods　Male C57BL/6 mice of 3 months and 12 months old were randomly divided into sham operation group and myocardial infarction(MI)group.Following acute myocardial infarction(AMI)modeling induced by open-chest surgery,the events of cardiac rupture were monitored and the echocardiography and hemodynamics were performed on the 7th day after surgery.Zymography,immunohistochemical method and pathological staining were used to measure the activity of matrix metalloproteinases(MMPs),the content of collagen and the degree of inflammatory cell infiltration on the 3rd and 7th days after surgery,respectively. Results　The incidence of cardiac rupture was higher in elderly group than that in young group(38.0% vs.16.0%,X2=6.139,P＜0.05).Compared with young group,significant infarct expansion,left ventricular (LV)remodeling and hemodynamic deterioration were showed in elderly group on the 7th day after surgery(t=5.754,P＜0.05).The degree of inflammatory cell infiltration and the expression of MMP-9 were significantly increased in elderly group on the 3rd day following AMI modeling(P＜0.05),and the collagen content and the expression of type Ⅲ collagen were significantly increased (P＜0.05)compared with young group. Conclusions　Aging is a risk factor for post-infarct cardiac rupture in the mice model.The mechanisms which are responsible for this age-related difference of cardiac rupture are related to increasing degree of inflammatory cell infiltration, overexpression of MMP-9 and type Ⅲ collagen and aggravated early LV remodeling.

Cloning, Expression, Purification of a Fusion Protein of SARS-CoV Nucleocapsid and Spike Protein Fragments and Its Immunological Characteristics / 中国生物化学与分子生物学报

The spike (S) and nucleocapsid (N) proteins, which are responsible for viral binding to cell surface receptors and the formation of ribonucleoprotein complexes during virion assembling, are major structure proteins of severe acute respiratory syndrome coronavirus (SARS-CoV). The expression of recombinant protein may give more accurate result for detecting SARS-CoV infection. A novel fusion protein,comprising of two fragments of N and S proteins from SARS-CoV, was prepared. Our computer-assisted analysis suggested that the immunodominant domains were located in the amino acid residues 1-227 of N protein and 450-650 of S protein, further the fusion of the two fragments did not change the immunochemical characteristics. The complementary DNA(cDNA) encoding N1-227 fused with S450-650 was obtained by sequence overlapping extension (SOE), and named NLS. It was cloned into pET-28a ( + ), an expression vector for His-tag fusion protein. This new constructed fusion protein was prokaryotic expressed in E. coli,and purified by metal chelate affinity chromatography with the purity over 95 %. The purified fusion protein was identified by anti-His monoclonal antibody and convalescence SARS patients serum. The NLS protein based ELISA showed that NLS maintained appropriate antigenicity and specificity to react with the sera of convalescent SARS patients. The functional NLS protein were successfully expressed and purified. And the fusion protein based ELISA can be used for detection of antibodies (Abs) against the S and N proteins of SARS-CoV. It may provided a novel diagnostic tool and have the potential application in developing of anti-SARS vaccine.

Beneficial Effect of Vigconic VI-28 Capsule on Cyclophosphamide-Treated C57BL/6 Mice Carrying Lewis Lung Cancer Tumor / 中国肿瘤生物治疗杂志

Objective: To investigate whether Vigconic VI-28 capsule, a formulated traditional Chinese medicine containing radix Ginseng, cornu cervi pantotrichum and semen cuscutae, can assist tumor chemotherapy in a mouse model. Methods: Female C57BL/6 mice were s. c. injected with viable Lewis lung cancer ( LLC) cells (106 cells/mouse) . The mice were then treated with cyclosposphamide (Cyp, 40 mg/kg bodyweight, once every other day). One group was intra-gastrically given 2% VI-28 (0.5 ml/mouse, every other day) during the course of the chemotherapy. By day 28, the mice were sacrificed and their thymic indices and tumor indices were calculated and compared. Splenocytes were collected for analysis of their immunological status. Histological study was carried to examine the solid tumors. Results: Fourteen days after the injection of LLC cells, solid tumors developed in most of the animals, reaching 1 ~ 1.8 cm diameters by 28 th day Compared with mice of the LCC + Cyp group, thymus glands from the LLC + Cyp + VI-28 group were significantly heavier. Splenocytes of the same group responded better to ConA stimulation in vitro. Histochemical examination of the tumor tissues revealed that tumors of the Cyp + VI-28 group were better differentiated (less aggressive) than that of the Cyp group. Conclusion: Vigconic VI-28 capsule can promote recovery of immune system in mice undergoing chemotherapy and help Cyp to control the growth of tumor cells in vivo.

DNA vaccination encoding variable region gene of the T cell receptor induces regulatory immune response in BALB/C mice / 中国免疫学杂志

Objective:To investigate regulatory immune response induced by DNA vaccination encoding T cell receptor V?5.2 or V?2.1 chain predominantly displayed on ovalbumin (OVA)-specific T cell clone.Methods:BALB/C mice were vaccinated with pcDNA3.1 encoding T cell receptor (TCR) V?5.2 or V?2.1 chain respectively.Using RT-PCR,transcription of the recombined plasmids was analysed. Cell proliferation was measured by 3H-TdR incorporation.CTL response was assayed by JAM test.Immuno-fluorescent assay was used to examine the anti-TCR antibody level and the number V?2 +T cells.Results:RT-PCR analysis showed that the recombined plasmids can be transcripted in vivo and in vitro.DNA vaccine with TCR variable chain effectively induced TCR-specific humoral and cellular immune response,V?2 +T cells was not depleted by V?2.1 TCR-DNA vaccination,but rather was anergy.Conclusion:Regulatory immune response can be induced by DNA vaccination encoding TCR V? or V? region in normal mice.

Molecular mechanisms and biology significance of polysaccharide recognition by the immune system / 中国免疫学杂志

As monosaccharide polymers,polysaccharides are widely distributed in nature,they are found in microorganisms,plants and animal cells and tissues.Glycosylation is of importance to maintain the structural stability and biological activity of protein molecules in eukaryotic cells.Polysaccharides represent the most important antigens recognized by immune system.The innate immune system senses invading microbes via their pathogen-associated molecular patterns,using pattern-recognition receptors on the surface of innate immune cells,many of which are polysaccharides uniquely expressed in certain microorganisms.Polysaccharides can also be processed intracellularly and presented to ??-T cells by MHC molecules.Polysaccharides can activate B lymphocytes,with or without T cell help,and induce production of specific antibodies.Amongst the polysaccharide-specific antibodies in human sera,some are products of adaptive humoral responses against microbial infection and others autoantibodies to glycan epitopes of host cells.Patients with autoimmune disorders tend to have higher titers of antibodies against various glycans.Immune clearance of senescence cells depends on the recognition of altered glycan epitopes on cell surface.Modification of glycan epitopes on tumour cell surface is at least partially responsible for activating antitumor immunity.Specific recognition of polysaccharides by the immune system is of importance to immunological defence,autoimmunity,anti-tumour immunity and immunological homeostasis,it will be the new focus of immunology research in the near future.