[Effect of different mechanical ventilation modes on patient-ventilator synchrony and diaphragm function in rabbit model of acute respiratory distress syndrome].

Objective: To observe the effect of different modes of mechanical ventilation on patient-ventilator synchrony and diaphragm function in rabbits with acute respiratory distress syndrome(ARDS). Methods: Eighteen New Zealand rabbit models of ARDS were induced by intratracheal infusion hydrochloric acid until the oxygenation index (PaO(2)/FiO(2)) was less than 200 mmHg, and then divided into three groups with random number: assisted-controlled mechanical ventilation (A/C) group, pressure support ventilation (PSV) group and neurally adjusted ventilatory assist (NAVA) group. All of them were ventilated for four hours with the targeted tidal volume (V(T)) (6 ml/kg) and the positive end-expiratory pressure (PEEP) titrated with the maximum oxygenation method. Gas exchange, pulmonary mechanics and patient-ventilator synchrony were determined during 4 h of ventilation and the concentrations of malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione (GSH) in diaphragm were measured after 4 h of ventilation. The q test was used for the multiple comparison of the sample mean. Results: There were no significant differences in PaO(2)/FiO(2) between three groups during ventilation 1-4 h (F=1.029, P>0.05). The V(T) in NAVA group was obviously lower than that in PSV group and the respiratory rate (RR) and the electrical activity of diaphragm(EAdi) were higher than those in A/C group(all P<0.05).The trigger delay and off cycle delay the in NAVA group were markedly lower than those in A/C and PSV group during ventilation 1-4 h(F=14.312, 9.342, both P<0.05). Asynchrony index in NAVA group (3.1%±1.0%) was obviously lower than those in A/C group (22.3%±5.2%) and PSV group(8.4%±2.3%) (F=7.192, P<0.05). In NAVA group, peak EAdi (EAdi(peak)) and peak airway pressure (Ppeak) were markedly correlated (r=0.97±0.16, P<0.05), while Ppeak delivery in A/C and PSV group was not correlated to EAdi(peak) (r=0.38±0.13,0.46±0.15, both P>0.05).Compared with A/C group, the concentration of MDA in the diaphragm in NAVA group was obviously lower(P<0.05). SOD and GSH level inthe diaphragm in NAVA group were both obviously higher than those in A/C group (both P<0.05). Conclusions: It is helpful to avoid eccentric contraction of diaphragm, lessen oxidative stress and alleviate ventilator-related diaphragm dysfunction by keeping spontaneous breathing as far as possible and subject-ventilator synchrony when ventilation in ARDS with NAVA.

Biological characteristics of osteoclast exosomes and their role in the osteogenic differentiation of somatic cells prior to osteogenesis.

This study aimed to investigate the biological characteristics of osteoclast exocrine bodies and their role in the differentiation of somatic cells, so as to find out the key factors involved in osteoclast exosomatic growth and osteogenesis. RANKL (Receptor Activator for Nuclear Factor-κ B Ligand) induced factor was used to induce the osteoclast differentiation of Raw 264.7 cells, and TRAP (Tartrate resistant acid phosphatase) staining was employed to identify induced cells. Ultra-filtration centrifugation was used to separate OC-exosomes from osteoclast supernatant, while Western blot was employed to detect the expression characteristics of exosomal proteins CD9 and CD63. PKH67 labeled exosomes were observed to target kusao cells, which were divided into 3 groups, i.e., the complete medium group (group A), the osteoblast induced group (group B), and the osteogenesis induced liquid + OC-exosomes group (group C). The medium was changed on the next day and after 14-day culture. Using Western blot, alizarin red staining and Von Kossa silver staining, the role of OC-exosomes in the differentiation of kusao cells was clarified. Results showed that TRAP staining showed osteoclasts as irregular and TRAP positive giant cells with a red multicore and a large volume. Microcapsule membrane structures with a uniform size were detected in osteoclast supernatant, and the expression of CD9 and CD63 proteins was confirmed by Western blot. In addition, the Western blot results showed that the expression of RUNX2 (Runt-related transcription factor 2) protein in group B was 1.254 times of that in group A and 2.636 times of that in group C. Furthermore, alizarin red staining showed that the ratios of calcium salt deposition area to the total area in group A, group B and group C were 0.208%, 3.469%, and 20.724%, respectively. Von Kossa silver staining showed that the ratios of calcium salt deposition area to the total area in group A, group B and group C were 0.064%, 2.636%, and 20.872%, respectively. To sum up, OC-exosomes can promote the osteogenic differentiation of osteoblast cells (kusao cells).

[Efficacy and safety of procalcitonin guidance in reducing the duration of antibiotic treatment of sepsis patients].

Objective: To assesse the efficacy and safety of procalcitonin-guided antibiotic treatment of sepsis patients in intensive care units (ICU). Methods: A prospective, randomised, controlled trial was gone in ICU of Northern Jiangsu People's Hospital.Between January 2013 and December 2015.One hundred and fifty-six patients assessed for eligibility were randomly assigned to the procalcitonin-guided group (PCT group, 79) or to regular antibiotic group (RAT group, 77). Patients who received antibiotics for presumed infection according to principle of antimicrobial usage.In the procalcitonin-guided group, a non-binding advice to discontinue antibiotics was provided if procalcitonin concentration had decreased by 90% or more of its peak value or to 0.25 µg/L or lower.In the regular antibiotic group, patients were treated according to principle of antimicrobial usage.The general status of the patient, antimicrobial drug use time, length of ICU stay, hospital stay time, number of cases of recurrence in 28 days and number of cases of death in 28 days were compared between the two groups. Results: There were no statistical significance in age, gender, blood culture positive rate, and chronic underlying diseases (P>0.05). While APACHE Ⅱ score of PCT group was (22.7±4.7) points, which was higher than that of RAT group (19.9±4.2) (P<0.05). Log Rank test results showed that the time of antimicrobial drug usage was significantly reduced in PCT group than RAT group [days: 8.3±0.3, 95% confidence interval (95%CI 7.9-9.1) vs 10.1±0.4, 95% confidence interval (95%CI 9.2-11.3), Log Rank value 31.85, P=0.000]. There was no significant difference in length of hospital stay, ICU stay time, number of cases of recurrence in 28 days and number of death in 28 days between two groups (P>0.05). Conclusion: Procalcitonin guidance stimulates reduction of duration of treatment and daily defined doses in critically ill patients with a presumed bacterial infection.This reduction does not affect the length of hospital stay, ICU stay time, number of cases of recurrence in 28 days and number of death in 28 days.

[The role and mechanism of mesenchymal stem cell in modulating human pulmonary microvascule endothelial cell permeability via paracrine hepatocyte growth factor].

Objective: To explore the role and mechanism of mesenchymal stem cell (MSC) in modulating human pulmonary microvascular endothelial cell (HPMECs) permeability via hepatocyte growth factor (HGF). Methods: The study introduced a co-cultured model between HPMECs and human mesenchymal stem cell conditioned media (MSC-CM) collected from MSCs after 24 h hypoxia culture, and meanwhile HGF was neutralized in MSC-CM by anti-HGF antibody respectively, followed by lipopolysaccharide (LPS) stimulation. Finally, the following measurements were performed: the permeability of HPMECs, the protein expression of vascular endothelial cadherin (VE-cadherin), Occludin in HPMECs by Western blot, HPMECs apoptosis by Annexin V-FITC/PI and HPMECs proliferation by 3-(4, 5)-dimethylthiahiazo (-z-y1)-3, 5-di- phenytetrazoliumromide(MTT). Results: Compared to LPS group (4.15±0.88), MSC-CM reduced endothelial paracellular permeability injured by LPS(1.56±0.36, P<0.01), however, the MSC-CM effect was significantly blocked by anti-HGF antibody(3.11±0.74, P<0.05). Furthermore MSC-CM significantly increased the expression of VE-cadherin(0.71±0.05 vs. 0.38±0.19, P<0.05)and Occludin protein(0.96±0.05 vs. 0.51±0.02, P<0.05) in HPMECs, which was significantly blocked by anti-HGF antibody (P<0.05). MSC-CM significantly reduced the number of early apoptotic cells (6.82±1.80 vs. 17.09±1.89, P<0.05). However, the effect of MSC-CM was significantly blocked by neutralizing HGF (12.07±0.98, P<0.01). The cell viability results by MTT assay confirmed that MSC-CM(6.82±1.80, P<0.05)restored cell viability to a greater extent than LPS stimulation only(0.47±0.09), and meanwhile the MSC-CM effect was significantly inhibited by neutralizing HGF from MSC-CM with anti-HGF antibody (0.69±0.29, P<0.05). Conclusion: HGF secreted by MSCs reduces endothelial cell paracelluar permeability induced by LPS, and the possible mechanisms include remodelling of endothelial intercellular adherence junction, promoting endothelial cell proliferation and restraining endothelial cell apoptosis.

Influences of hucMSC-exosomes on VEGF and BMP-2 expression in SNFH rats.

OBJECTIVE: To investigate the influences of human umbilical cord mesenchymal stem cell-derived exosomes (hucMSC-exosome) on steroid-induced necrosis of the femoral head (SNFH) and the expressions of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) in rats. PATIENTS AND METHODS: A total of 20 male Sprague-Dawley rats were randomly divided into SNFH group and SNFH + hucMSC-exosome group using a random number table. Prednisolone acetate (24.5 mg/kg) was injected twice a week to establish the rat model of SNFH, and hucMSC-exosome in a certain dose was additionally injected into the marrow cavity in SNFH + hucMSC-exosome group. After 3 weeks, the influences of hucMSC-exosome on the pathological changes and apoptosis of the femoral head in SNFH rats were detected via hematoxylin-eosin (H&E) staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. In addition, the expressions of cluster of differentiation 31 (CD31), VEGF, and BMP-2 in bone tissues in both groups were detected via immunohistochemical staining, and the messenger ribonucleic acid (mRNA) and protein expression levels of VEGF and BMP-2 in necrotic bone tissues in both groups were detected via Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and Western blotting. RESULTS: The results of H&E staining revealed that the fibrous callus formation was good, the new trabecular structure was more obvious, the number of vacuum cleft declined, and there were fewer enlarged adipocytes in SNFH + hucMSC-exosome group compared with SNFH group. The results of TUNEL staining showed that the number of apoptotic cells in femoral head tissues was smaller in SNFH + hucMSC-exosome group (p<0.05). According to the results of immunohistochemistry, hucMSC-exosome could increase the expression of vascular endothelial marker CD31 in SNFH rats (p<0.05). Besides, the results of RT-PCR, immunostaining and Western blotting manifested that both the mRNA and protein levels of BMP-2 and VEGF in femoral head tissues were significantly increased in SNFH + hucMSC-exosome group (p<0.05). CONCLUSIONS: HucMSC-exosome can improve SNFH in rats, whose mechanism may be related to the up-regulation of VEGF and BMP-2 by exosomes.

LncRNA HOTAIR participates in the development and progression of adrenocortical carcinoma via regulating cell cycle.

OBJECTIVE: To explore the role of HOTAIR in the pathogenesis of adrenocortical carcinoma (ACC) and its underlying mechanism. PATIENTS AND METHODS: Differentially expressed lncRNA (HOTAIR) in ACC was screened out from the GEO database. The survival analysis and ROC curve were performed according to HOTAIR expressions in ACC patients. The correlation between HOTAIR expression and clinical information of ACC patients was analyzed by chi-square test. The univariate and multivariate COX regression analysis was carried out to analyze the relationship between HOTAIR expression, disease-free survival (DFS) and overall survival (OS) of ACC patients. We then detected HOTAIR expression in 77 ACC tissues and 30 normal tissues by qRT-PCR (quantitative Real-time polymerase chain reaction). ACC cell lines were further screened out for the following in vitro experiments. After altering HOTAIR expression in ACC cells by plasmid transfection, proliferation and cell cycle were detected by Cell Counting Kit-8 (CCK-8) and colony formation assay, respectively. Finally, Western blot was utilized to detect expressions of cell cycle-related genes in ACC cells. RESULTS: HOTAIR was overexpressed in ACC tissues than that of normal tissues. HOTAIR expression was remarkably increased in ACC with T3 and T4 stage than that of T1 and T2 stage. Moreover, HOTAIR expression was remarkably increased in ACC with stage III and IV than that of stage I and II. HOTAIR was an independent prognostic factor for DFS and OS of ACC patients. For in vitro experiments, inhibited proliferation and arrested cell cycle were observed in H295R cells transfected with si-HOTAIR. Opposite results were obtained after SW-13 cells were transfected with HOTAIR overexpression plasmid. Furthermore, expressions of cell cycle-related genes, including Cyclin D1, p-Rb and p-GSK3ß were remarkably decreased after HOTAIR knockdown. CONCLUSIONS: We demonstrated for the first time that HOTAIR is overexpressed in ACC and is a prognostic risk factor in ACC patients. HOTAIR participates in the development and progression of ACC via shortening cell cycle and promoting proliferation of ACC cells.

Hepatic angiomyolipoma: identification of an efferent vessel to be hepatic vein by contrast-enhanced harmonic ultrasound.

We report two patients with rare hepatic angiomyolipoma and demonstrate the special tumour haemodynamics with contrast-enhanced harmonic ultrasound. This reliably identified the efferent vessel of the hepatic angiomyolipoma to be the hepatic vein in both cases, which corresponded well with that seen on conventional angiography and CT angiography. This haemodynamic finding may be an important characteristic of hepatic angiomyolipoma, and facilitate the differential diagnosis from other benign and malignant hepatic tumours.

Co-localization of tissue factor protein and its mRNA in human blood monocytes.

Triggering of both the extrinsic and intrinsic coagulation pathways is mediated by the cell surface receptor Tissue Factor (TF). The ability to observe the cell surface TF protein and its mRNA at the single cell level would facilitate our understanding of the cellular biology of TF in health and diseased states. Employing the methods of immuno-gold silver staining and in situ hybridization using non-isotopically labelled oligoprobes, tissue factor antigen and mRNA were detected simultaneously in endotoxin stimulated human peripheral blood monocytes.

Production of tissue factor by monocyte progenitor cells.

Tissue factor (TF) is known to be produced by monocytes from human peripheral blood. However the production of this factor by haematopoietic progenitor cells is not yet known. We thus studied human monocyte progenitor cells isolated from bone marrow of normal and diseased individuals. These cells were non-adherent, monocytic and able to phagocytose particles ranging from 0.3-1 microns. Unactivated partial thromboplastin time clotting assay demonstrated procoagulant activity consistent with TF function, which was blocked by a neutralizing anti-TF monoclonal antibody, G12. The production of TF messenger RNA was demonstrated on dot blot and northern blot analysis utilizing an oligonucleotide probe.

Prognostic value of dobutamine stress test in patients with septic shock.

AIM: To evaluate the prognostic effect of dobutamine stress test in patients with septic shock. METHODS: Patients with septic shock received intravenous infusion of dobutamine at 10 microg . kg-1 . min-1 for 1 h. Hemodynamics and oxygen metabolism was observed. A patient who was able to increase oxygen consumption index (VO2) by >15 % was designated as a responder to the test. RESULTS: In 47 patients with septic shock, twenty one responders and twenty six nonresponders were identified, and mortality was 33.3 % and 76.9 % respectively. After the dobutamine infusion, the responders showed increases in cardiac index (18.1 %), oxygen delivery index (12.7 %), VO2 (38.6 %), and oxygen extraction ratio (18.0 %), and reductions in systemic vascular resistance index (18.5 %). The nonresponders demonstrated increases in cardiac index (11.5 %), but no change in oxygen delivery, VO2, and oxygen extraction ratio. CONCLUSION: Dobutamine stress test might be a good predictor of outcome in patients with septic shock.

Transforming growth factor-beta 1 in rheumatoid synovial membrane and cartilage/pannus junction.

Transforming growth factor (TGF)-beta has been shown to promote tissue repair and have immunosuppressive actions, and has been proposed to have a role in rheumatoid arthritis (RA). Using immunohistochemical techniques with rabbit F(ab')2 antibodies raised against recombinant human TGF-beta 1, we have detected TGF-beta 1 in the synovial tissue and cartilage/pannus junction (CPJ) from 18/18 patients with RA. TGF-beta 1 was found predominantly in the thickened synovial lining layer in RA, but also detected in a perivascular pattern in the synovial interstitium as well as in occasional cells in the lymphoid aggregates. At the CPJ it was found both in cells at the distinct junction as well as in the transitional region of the diffuse fibroblastic zone. The cells staining for TGF-beta 1 were identified by double immunofluorescence staining as being from the monocyte/macrophage series as well as the type B synovial lining cells. TGF-beta 1 was also detected in the synovial membrane sections from 4/4 patients with systemic lupus erythematosus/mixed connective tissue disease and 5/8 patients with osteoarthritis, in a similar distribution to that seen in RA, and in the lining layer of 1/7 normal synovial membranes. These results add to histological evidence confirming that TGF-beta 1 is present in RA synovial cells and those from other arthritides. The distributions of TGF-beta 1 in RA synovial membrane reflects its known actions, as it can be detected at the CPJ, where it could induce repair, and close to activated cells upon which it may exert an immunosuppressive action.

Detection of interleukin-6 and interleukin-1 production in human thyroid epithelial cells by non-radioactive in situ hybridization and immunohistochemical methods.

Human endocrine thyroid epithelial cells have been described to produce cytokines in vitro. In order to determine whether they do so in vivo during thyroiditis, parallel studies on mRNA expression with a non-radioactive in situ hybridization technique and immunohistochemical detection for the protein were performed on frozen sections of thyroid samples from autoimmune thyroiditis (Graves' disease and Hashimoto's thyroiditis), non-toxic goitre and normal thyroid tissue. cDNA probes were sulphonated and their hybridization with mRNA was detected with a sulphonyl-specific monoclonal antibody. This signal was amplified and visualized with the alkaline phosphatase-anti-alkaline phosphatase (APAAP) system. The protein products were detected with immuno-purified rabbit F(ab')2 antibody fragments recognizing recombinant human cytokines, visualized by the immunoperoxidase technique. Each sample was studied at the two levels. Both interleukin-6 mRNA and protein were found in the endocrine cells. There was no obvious difference between autoimmune thyroiditis and non-toxic goitre. However, normal thyroid epithelial cells produced less interleukin-6. Interleukin-1 alpha mRNA and its protein were found in epithelial cells from Hashimoto's thyroiditis samples, but not in the others, except one Graves' disease sample, in which only mRNA was detected. Interleukin-1 beta was not detected in these cells, its mRNA was only found in one of the Graves' disease samples. These cytokines were also detected in some infiltrating cells.

Expression of intercellular adhesion molecule-1 and lymphocyte function-associated antigen-3 on human thyroid epithelial cells in Graves' and Hashimoto's diseases.

Human endocrine thyroid epithelial cells (TEC) from autoimmune thyroiditis which express HLA Class II antigens have been shown to present autoantigens to T cells for a TEC-specific immune response. Since the initiation of a specific immune response also involves antigen-receptor independent interactions between accessory molecules, such as lymphocyte function-associated antigen-1 (LFA-1) with intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-3 (LFA-3) with CD2, it was of interest to determine whether TEC can express the adhesion molecules (ICAM-1 and LFA-3) which augment the efficiency of antigen presentation. Cultured TEC were studied for their expression of ICAM-1 and LFA-3 by immunofluorescence. Those derived from Graves' disease expressed these molecules after stimulation with recombinant human interferon-gamma (IFN gamma) or with recombinant human tumour necrosis factor-alpha (TNF alpha). However, using the same stimuli, TEC from non-toxic goitre were induced to express ICAM-1, but not LFA-3. To establish whether ICAM-1 and LFA-3 on TEC were expressed in vivo during the disease process, antibodies against these molecules were incubated with frozen sections of autoimmune thyroiditis, including Graves' and Hashimoto's diseases, and non-toxic goitre. Both ICAM-1 and LFA-3 were highly expressed in the autoimmune diseases, but not in non-toxic goitre. These findings establish that TEC are able to express adhesion molecules and suggest the possible involvement of these adhesion molecules in the TEC-specific immune response in autoimmune thyroiditis.

Detection of in vivo production of tumour necrosis factor-alpha by human thyroid epithelial cells.

We have established previously that human thyroid epithelial cells (TEC) from patients with autoimmune thyroiditis are able to synthesize cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6). This paper examines TEC in sections from autoimmune thyroiditis for the in vivo production of tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) using the combined techniques of in situ hybridization and immunohistochemistry. Thyroid tissue from patients with Graves' disease, Hashimoto's disease and non-toxic goitre was examined and both mRNA and the protein of TNF-alpha were detected in TEC on frozen sections. Representative figures of only Graves' samples are illustrated in this paper. In contrast, using the same methods, IFN-gamma was detected only in the infiltrating cells and not in TEC of thyroid tissue from the patients.