Quorum sensing bacteria in microplastics epiphytic biofilms and their biological characteristics which potentially impact marine ecosystem.

Microplastics (MPs) have been shown to be a new type of pollutant in the oceans, with complex biofilms attached to their surfaces. Bacteria with quorum sensing (QS) systems are important participants in biofilms. Such bacteria can secrete and detect signal molecules. When a signal molecule reaches its threshold level, bacteria with QS systems can perform several biological functions, such as biofilm formation and antibiotic metabolite production. However, the ecological effects of QS bacteria in biofilm as MPs distribute globally with ocean currents are not to be elucidate yet. In this study, polypropylene and polyvinyl chloride were selected for on-site enrichment to acquire microplastics with biofilms. Eight culturable QS bacteria in the resulting biofilm were isolated by using biosensor assays, and their biodiversity was analyzed. The profiles of the N-acyl-homoserine lactones (AHLs) produced by these bacteria were analyzed by using thin-layer chromatography (TLC)-bioautography and gas chromatography and mass spectrometry (GC-MS). Biofilm-forming properties and several biological characteristics, such as bacteriostasis, algal inhibition, and dimethylsulfoniopropionate (DMSP) degradation, were explored along with QS quenching. Results showed that QS bacteria were mainly affiliated with class Alphaproteobacteria, particularly Rhodobacteraceae, followed by class Gammaproteobacteria. TLC-bioautography and GC-MS analyses revealed that seven AHLs, namely, C6-HSL, C8-HSL, 3-oxo-C6-HSL, 3-oxo-C8-HSL, 3-oxo-C10-HSL, and two unidentified AHLs were produced. The QS system equipped bacteria with strong biofilm-forming capacity and may contribute to the keystone roles of Rhodobacteraceae. In addition, QS bacteria may exacerbate the adverse environmental effects of MPs, such as inducing the misfeeding of planktons on MPs. This study elucidated the diversity of QS bacteria in MP-associated biofilms and provided a new perspective of the effect of key membrane-forming bacteria on the marine ecological environment.

Lpar1-mediated Effects in Endothelial Progenitor Cells Are Crucial for Lung Repair in Acute Respiratory Distress Syndrome/Acute Lung Injury.

Acute respiratory distress syndrome/acute lung injury (ARDS/ALI) involves acute respiratory failure characterized by vascular endothelial and lung alveolar epithelial injury. Endothelial progenitor cells (EPCs) can mediate vasculogenesis. However, the limitations of EPCs, such as low survival and differentiation, are believed to inhibit the effectiveness of autologous cell therapies. This study demonstrated that lysophosphatidic acid (LPA), a bioactive small molecule without immunogenicity, is involved in the survival and antiapoptotic effects in human umbilical cord mesenchymal stem cells. This study aimed to explore whether LPA improves the survival of EPCs, enhancing the cellular therapeutic efficacy in ARDS, and these results will expand the application of LPA in stem cells and regenerative medicine. LPA promoted the colony formation, proliferation, and migration of EPCs and upregulated the expression of vascular endothelial-derived growth factor (VEGF) in EPCs. LPA pretreatment of transplanted EPCs improved the therapeutic effect by increasing EPC numbers in the rat lungs. LPA enhanced EPC proliferation and migration through Lpar1 coupled to Gi/o and Gq/11, respectively. Activation of extracellular signal-related kinase 1/2, or ERK1/2, was related to LPA-induced EPC proliferation but not migration. LPA/Lpar1-mediated Gi/o protein was also shown to be involved in promoting VEGF expression and inhibiting IL-1α expression in EPCs. Low LPA concentrations are present after lung injury; thus, the restoration of LPA may promote endothelial cell homeostasis and lung repair in ARDS. Inhalation of LPA significantly promoted the homing of endogenous EPCs to the lung and reduced lung injury in both rats with LPS-induced ALI and Streptococcus pneumoniae-infected mice. Taken together, these data indicated that LPA/Lpar1-mediated effects in EPCs are involved in maintaining endothelial cell homeostasis and lung tissue repair under physiological conditions.

MicroRNAs combined with the TLR4/TDAG8 mRNAs and proinflammatory cytokines are biomarkers for the rapid diagnosis of sepsis.

The early diagnosis and treatment of sepsis are of particular importance to patient survival. To obtain novel biomarkers that serve as prompt indicators of sepsis, the current study screened the differentially expressed microRNAs (DEMs) that were associated with sepsis susceptibility. The correlation between the elucidated DEMs and the inflammatory response was also examined. The present study included 40 patients with sepsis and 40 healthy controls. RNA­sequencing technology and bioinformatics analysis were applied to screen the DEMs between the two cohorts. The expression of these DEMs was subsequently verified by performing reverse transcription­quantitative PCR (RT­qPCR). In addition, IL­6, IL­21, C­X­C motif chemokine ligand­8 (CXCL8) and monocyte chemoattractant protein­1 (MCP­1) levels, along with T­cell death­associated gene 8 (TDAG8) and toll­like receptor 4 (TLR4) mRNA expression levels were assessed. The association between microRNA (miRNA/miR)­3663­3p and the secretion of various proinflammatory cytokines or TDAG8 and TLR4 mRNA expressions were subsequently evaluated by linear correlation analysis. The results revealed 305 DEMs (P<0.05; fold change >2) between patients with sepsis and healthy controls. Among these, the top 18 up­ and downregulated miRNAs were selected for RT­qPCR verification. In addition, the serum content of IL­6, IL­21, CXCL8 and MCP­1, and the expression of TDAG8 and TLR4 mRNAs were significantly increased in patients with sepsis compared with healthy controls. Moreover, in patients with sepsis, a positive correlation was identified between miR­3663­3p and the secretion of inflammatory cytokines or TDAG8 and TLR4 mRNA expression. A positive correlation was also elucidated between TDAG8 and TLR4 mRNA expression and proinflammatory cytokine/chemokine secretion. Receiver operating characteristic curve analysis of miR­3663­3p expression, IL­6, IL­21, CXCL8 and MCP­1 secretion and TDAG8 and TLR4 mRNA expression demonstrated that miRNA analysis may be invaluable for the diagnosis of sepsis. Collectively, the results determined that miR­3663­3p may be a potentially powerful diagnostic and predictive biomarker of sepsis and that the combined and simultaneous detection of several biomarkers, including proteins, miRNAs and mRNA may be a reliable approach for the fast diagnosis and early identification of sepsis.

Adipose­derived mesenchymal stem cell­derived HCAR1 regulates immune response in the attenuation of sepsis.

Sepsis serves as a leading cause of admission to and death of patients in the intensive care unit (ICU) and is described as a systemic inflammatory response syndrome caused by abnormal host response to infection. Adipose­derived mesenchymal stem cells (ADSCs) have exhibited reliable and promising clinical application potential in multiple disorders. However, the function and the mechanism of ADSCs in sepsis remain elusive. In the present study, the crucial inhibitory effect of ADSC­derived hydroxy­carboxylic acid receptor 1 (HCAR1) on sepsis was identified. Reverse transcription quantitative­PCR determined that the mRNA expression of HCAR1 was reduced while the mRNA expression of Toll­like receptor 4 (TLR4), major histocompatibility complex class II (MHC II), NOD­like receptor family pyrin domain containing 3 (NLRP3), and the levels of interleukin­1ß (IL­1ß), tumor necrosis factor­α (TNF­α), interleukin­10 (IL­10), and interleukin­18 (IL­18) were enhanced in the peripheral blood of patients with sepsis. The expression of HCAR1 was negatively correlated with TLR4 (r=­0.666), MHC II (r=­0.587), and NLRP3 (r=­0.621) expression and the expression of TLR4 was positively correlated with NLRP3 (r=0.641), IL­1ß (r=0.666), TNF­α (r=0.606), and IL­18 (r=0.624) levels in the samples. Receiver operating characteristic (ROC) curve analysis revealed that the area under the ROC curve (AUC) of HCAR1, TLR4, MHC II and NLRP3 mRNA expression was 0.830, 0.853, 0.735 and 0.945, respectively, in which NLRP3 exhibited the highest diagnostic value, and the AUC values of IL­1ß, IL­18, TNF­α, and IL­10 were 0.751, 0.841, 0.924 and 0.729, respectively, in which TNF­α exhibited the highest diagnostic value. A sepsis rat model was established by injecting lipopolysaccharide (LPS) and the rats were randomly divided into 5 groups, including a normal control group (NC group; n=6), a sepsis model group (LPS group; n=6), an ADSC transplantation group (L + M group; n=6), a combined HCAR1 receptor agonist group [L + HCAR1 inducer (Gi) + M group; n=6], and a combined HCAR1 receptor inhibitor group [L + HCAR1 blocker (Gk) + M group; n=6]. Hematoxylin and eosin staining determined that ADSCs attenuated the lung injury of septic rats and ADSC­derived HCAR1 enhanced the effect of ADSCs. The expression of HCAR1, TLR4, MHC II, NLRP3, IL­1ß, IL­18 and TNF­α levels were suppressed by ADSCs and the effect was further induced by ADSC­derived HCAR1. However, ADSC­derived HCAR1 induced the levels of anti­inflammatory factor IL­10. The negative correlation of HCAR1 expression with TLR4, MHC II, and NLRP3 expression in the peripheral blood and lung tissues of the rats was then identified. It is thus concluded that ADSC­derived HCAR1 regulates immune response in the attenuation of sepsis. ADSC­derived HCAR1 may be a promising therapeutic strategy for sepsis.

HMGB1 Upregulates RAGE to Trigger the Expression of Inflammatory Factors in the Lung Tissue in a Hypoxic Pulmonary Hypertension Rat Model.

Hypoxic pulmonary hypertension (HPH), a form of pulmonary hypertension (PH) caused by hypoxia, could cause serious complications and has a high mortality rate, and no clinically effective treatments are currently available. In this study, we established an HPH preclinical model in rats by simulating clinicopathological indicators of the disease. Our results showed that high mobility group box-1 protein (HMGB1) aggravated the clinical symptoms of HPH. We aimed at establishing protocols and ideas for the clinical treatment of HPH by identifying downstream HMGB1 binding receptors. Our investigation showed that continuous hypoxia could cause significant lung injury in rats. ELISA and western blotting experiments revealed that HPH induces inflammation in the lung tissue and increases the expression of a hypoxia-inducible factor. Testing of lung tissue proteins in vivo and in human pulmonary artery endothelial cells in vitro revealed that the HMGB1-mediated increase in the receptor for advanced glycation end products (RAGE) expression promoted inflammation. In summary, we successfully established an HPH rat model providing a new model for preclinical HPH research and elucidated the role of HMGB1 in HPH. Furthermore, we describe that HMGB1 induced inflammation in the HPH model via RAGE, causing severe lung dysfunction. This study could potentially provide novel ideas and methods for the clinical treatment of HPH.

Diversity and functions of quorum sensing bacteria in the root environment of the Suaeda glauca and Phragmites australis coastal wetlands.

The quorum sensing (QS) system plays a significant role in the bacteria-bacteria or plant-bacteria relationships through signal molecules. However, little is known about the distribution and functional diversity of QS bacteria in the root environment of Suaeda glauca and Phragmites australis in coastal wetlands. We explored the bacterial community by amplicon sequencing and isolated 1050 strains from the rhizosphere soil and root tissues of S. glauca and P. australis in northern China to investigate the bacterial community and AHL producers. AHL activity was found in 76 isolates, and 22 distinct strains were confirmed by 16S rRNA gene sequencing. A substantial number of AHL producers clustered in rhizobiales and sphingomonadale, which derived from the root tissues. AHL producers in the rhizosphere soil mostly belonged to rhodobacterales. The different taxa of AHL producers in the rhizosphere soil and root tissues resulted in a variation of AHL profiles that C6-HSL dominated the AHL profiles in root bacteria compared to the C8-HSL in rhizobacteria, implying different ecological roles for AHL producers in the rhizosphere soil and root tissues. Many AHL producers may form biofilms, and some can degrade DMSP and oil, demonstrating that QS bacteria in the root environment have a wide ecological roles. In our study, for one of the first times here, we explore the distribution and functional variety of AHL producers in the root environment of S. glauca-P. australis. This study expands current knowledge of the relationship between QS bacteria and coastal plants (S. glauca and P. australis), and vital roles of QS bacterial in maintaining the health of coastal wetlands.

Effect of Src tyrosine kinase on a rat model of asthma.

Src tyrosine kinase is a protein encoded by the Src gene. The present study aimed to determine the role of Src protein kinase in asthma using small interfering RNA (siRNA) technology. Several Src siRNAs were designed and the most effective siRNA pair was selected. A rat model of asthma was established using ovalbumin, and the rats were treated with Src siRNA, empty vector or phosphate-buffered saline (PBS). A non-asthmatic control group was also established. The rats were clinically observed and Src mRNA and protein levels were measured by reverse transcription-quantitative PCR and western blot analysis, respectively. Pathological observation of the lung tissue, counting of white blood cells (WBCs) and eosinophils (EOSs) and analysis of the concentrations of IL-5, IL-33 and IFN-γ in the bronchoalveolar lavage fluid were performed. The expression levels of Src mRNA in the control, PBS, empty vector and siRNA groups were 110±30.7x103, 253±55.4x103, 254±41.3x103 and 180±50.9x103, respectively. Histochemical analysis of the lung tissue of rats in the siRNA group exhibited a relatively complete lung structure and little damage to the alveolar cavity. Src protein expression and IL-5, IL-33 levels, WBC and EOS levels were positively correlated with Src mRNA expression, while the IFN-γ concentration was negatively correlated with Src mRNA expression. These results indicate that Src knockdown inhibits the release of tracheal inflammatory factors and significantly alleviates asthma in rats. In conclusion, the present study utilized a gene transfer technique to interfere with the expression of Src in rats, which decreased the levels of IL-5, IL-33, WBCs and EOSs and increased the level of IFN-γ; these changes effectively ameliorated the condition of the trachea in asthmatic rats.

Effect of surgical starting time and season on prognosis in octogenarians with colorectal cancer: a retrospective study.

Aim: To investigate the effect of surgical starting time and season on the prognosis of octogenarians with colorectal cancer. Patients & methods: A total of 291 patients aged 80 years or above who received elective colectomy for colorectal cancer between January 2007 and December 2018 in the National Cancer Center in China were included. Results: No significant time- or season-dependent difference in overall survival for all clinical stages was found in the study. Comparing perioperative outcomes, the morning group had a longer operative time than the afternoon group (p = 0.03), but no significant difference was found based on the season of colectomy. Conclusion: These findings provide insights into clinical outcomes for colorectal cancer patients aged more than 80 years.

Recurrent studies have demonstrated that in heart surgery, different surgical starting times can affect the patients' outcomes, mainly due to the 24-h cyclic variations in heart function. This variability also exists in bowel function. The surgical outcomes of elderly patients aged over 80 years are more susceptible to external factors due to their frailty, so we wanted to compare the differences in prognosis of elderly patients who underwent surgery at different times and seasons.

Screening of Biomarkers Involved in Idiopathic Pulmonary Fibrosis and Regulation of Upstream miRNAs.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is the most common type of fatal interstitial lung disease and IPF patients usually have a poor prognosis. Biomarkers that can predict the occurrence, process and prognosis of IPF will be useful for its diagnosis and treatment. This study aimed to identify the potential biomarkers of IPF and analyze the regulation of upstream miRNAs. METHODS: The miRNA and gene expression profiles were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) and miRNAs (DEMs) between IPF and normal groups were identified. After co-expression gene pair analysis, functional enrichment analysis was performed. Then, the potential biomarkers of IPF were screened and validated. Finally, the upstream regulatory miRNA of biomarkers was predicted. RESULTS: A total of 343 DEGs and 21 DEMs were identified between IPF and normal samples. CLDN18, COL6A3, MYRF, PRSS8, RRAS, and SBNO1 were identified as potential IPF biomarkers. In addition, 17 miRNA-target relationship pairs were obtained. The up-regulation of hsa-miR-657, hsa-miR-671-5p, hsa-miR-198, and hsa-miR-940 could regulate the down-regulation of MYRF and the up-regulation of hsa-miR-198 and hsa-miR-373-3p could regulate the down-regulation of RRAS and CLDN18, respectively. Our data indicated that PRSS8, hsa-miR-614, and hsa-miR-503-5p might be involved in the migration and invasion of IPF related cells. CONCLUSIONS: CLDN18, COL6A3, MYRF, PRSS8, RRAS, and SBNO1 might be potential IPF biomarkers. However, the specific role of these genes and miRNA in IPF needs further experimental research.

Long-term effect of anesthesia choice on patients with hepatocellular carcinoma undergoing open liver resection.

Clinical and experimental evidence suggested that anesthesia choice can influence cancer progression and patients' outcomes by modulating tumor microenvironment and tumorigenic pathways. Curative resection is the mainstay of therapy for hepatocellular carcinoma (HCC), which is an intractable disease due to high recurrence and poor prognosis. However, different anesthetics may play different roles in alleviating surgery-induced stress response and inflammatory cytokines release that are considered to be closely associated with proliferation, invasion and metastasis of tumor cells. Propofol, sevoflurane, non-steroidal anti-inflammatory drugs and local anesthetics have shown to exert anti-tumor effect on HCC mainly through regulating microRNAs or signaling pathways, while other inhalational agents, dexmedetomidine and opioids have the potential to promote tumor growth. In terms of anesthetic methods and analgesia strategies, propofol based total intravenous anesthesia and thoracic epidural analgesia could be preferred for HCC patients undergoing open liver resection rather than inhalational anesthesia. Local anesthesia techniques have great potential to attenuate perioperative stress response, hence they may contribute to more favorable outcomes. This review summarized the relations between different anesthesia choices and HCC patients' long-term outcomes as well as their underlying mechanisms. Due to the complexity of molecules interactions and signaling pathways, further studies are warranted to confirm these results so as to optimize anesthesia strategy for HCC patients.

Quorum Sensing Bacteria in the Phycosphere of HAB Microalgae and Their Ecological Functions Related to Cross-Kingdom Interactions.

It has been proven that the relationship between microalgae and bacteria affects the dynamic process of harmful algal blooms (HABs). Microalgae-associated microorganisms widely exist in the phycosphere and play an essential role in algae-bacteria cross-kingdom interactions. Among these processes, quorum sensing (QS), as a communication system of bacteria, is thought to participate in algae-bacteria interactions. However, the species of QS bacteria in the phycosphere and their ecological function are still unknown. In this study, microalgae-associated microorganisms with a QS system were screened by the biosensor method and identified based on 16S rRNA gene analysis. The types and number of acyl-L-homoserine lactone (AHL) signalling molecules produced by QS bacteria were analysed by thin layer chromatography (TLC) bioautography and gas chromatography-mass spectrometer (GC-MS). The film formation, ß-dimethylmercaptopropionic (DMSP) degradation and algae growth effects of QS bacteria were investigated. The results showed that 113 QS bacteria were isolated from 842 microalgae-associated bacteria. Detection of AHL molecules in 10 different species of QS bacteria showed that most of them were N-(3-Oxodecanoyl)-L-homoserine lactone (OC10-HSL), N-Octanoyl-L-homoserine lactone (C8-HSL) and N-(3-Oxooctanoyl)-L-homoserine lactone (OC8-HSL). All 10 QS bacteria had film-forming ability, and they could degrade DMSP (except strain E26). The crude metabolic extracts of the 10 QS bacteria can inhibit or promote microalgae growth to different degrees. Our study is helpful to understand the role of microalgae-associated microorganisms with the QS system in algae-bacteria interactions and community succession of HAB microalgae.

Complete Genome Sequence of a Quorum-Sensing Bacterium, Oceanicola sp. Strain D3, Isolated from a Microplastic Surface in Coastal Water of Qingdao, China.

Oceanicola sp. strain D3 was isolated from the plastisphere of polyvinyl chloride (PVC) in coastal water of Qingdao, China. Here, we present the complete genome sequence of strain D3, which consists of a chromosome of 3,926,685 bp with a G+C content of 64.49% and 4,964 coding DNA sequences. This is the first report of a quorum-sensing (QS) system in an Oceanicola sp. strain.

Marine microplastic-associated bacterial community succession in response to geography, exposure time, and plastic type in China's coastal seawaters.

Microplastics have emerged as new pollutants in oceans. Nevertheless, information of the long-term variations in the composition of plastic-associated microbial communities in coastal waters remains limited. This study applied high-throughput sequencing to investigate the successional stages of microbial communities attached to polypropylene and polyvinyl chloride microplastics exposed for one year in the coastal seawater of China. The composition of plastisphere microbial communities varied remarkably across geographical locations and exposure times. The dominant bacteria in the plastisphere were affiliated with the Alphaproteobacteria class, particularly Rhodobacteraceae, followed by the Gammaproteobacteria class. Scanning electron microscopy analysis revealed that the microplastics showed signs of degradation. Microbial communities showed adaptations to plastisphere including more diverse microbial community and greater "xenobiotics biodegradation and metabolism" in metabolic pathway analysis. The findings elucidate the long-term changes in the community composition of microorganisms that colonize microplastics and expand the understanding of plastisphere microbial communities present in the marine environment.

A multicenter RCT of noninvasive ventilation in pneumonia-induced early mild acute respiratory distress syndrome.

RATIONALE: Our pilot study suggested that noninvasive ventilation (NIV) reduced the need for intubation compared with conventional administration of oxygen on patients with "early" stage of mild acute respiratory distress syndrome (ARDS, PaO2/FIO2 between 200 and 300). OBJECTIVES: To evaluate whether early NIV can reduce the need for invasive ventilation in patients with pneumonia-induced early mild ARDS. METHODS: Prospective, multicenter, randomized controlled trial (RCT) of NIV compared with conventional administration of oxygen through a Venturi mask. Primary outcome included the numbers of patients who met the intubation criteria. RESULTS: Two hundred subjects were randomized to NIV (n = 102) or control (n = 98) groups from 21 centers. Baseline characteristics were similar in the two groups. In the NIV group, PaO2/FIO2 became significantly higher than in the control group at 2 h after randomization and remained stable for the first 72 h. NIV did not decrease the proportion of patients requiring intubation than in the control group (11/102 vs. 9/98, 10.8% vs. 9.2%, p = 0.706). The ICU mortality was similar in the two groups (7/102 vs. 7/98, 4.9% vs. 3.1%, p = 0.721). Multivariate analysis showed minute ventilation greater than 11 L/min at 48 h was the independent risk factor for NIV failure (OR, 1.176 [95% CI, 1.005-1.379], p = 0.043). CONCLUSIONS: Treatment with NIV did not reduce the need for intubation among patients with pneumonia-induced early mild ARDS, despite the improved PaO2/FIO2 observed with NIV compared with standard oxygen therapy. High minute ventilation may predict NIV failure. TRIAL REGISTRATION: NCT01581229 . Registered 19 April 2012.

[Effects of hypoxia on the expression of HMGB1 and related inflammatory factors in human pulmonary artery smooth muscle and pulmonary artery endothelial cells].

OBJECTIVE: To investigate the effects of hypoxia on the expressions of high mobility group box-1(HMGB1), HMGB1 receptors and inflammatory factors in human pulmonary artery smooth muscle cell( HPASMC) and human pulmonary artery endothelial cells (HPAEC).The effects of HMGB1 on the proliferation and migration activity of the two kinds of cells were detected. METHODS: HPASMC and HPAEC were cultured under hypoxic conditions (Hypoxia at 1% oxygen, Hypoxia group) and normoxic conditions(Control group) . The mRNA levels of HMGB1, Toll-like receptors 2,4,9 (TLR2 , TLR4, TLR9), the receptor of advanced glycation end products(RAGE) , CD24 and IL-6 ,TNF-α,CXCL8 were detected by real-time PCR. Cell proliferation was measured by MTS. Cell migration was analysed by wound healing test. RESULTS: Compared with control group, the expressions of HMGB1 mRNA and RAGE mRNA in both HPASMC and HPAEC were increased significantly (P＜0.05 and 0.01). Meanwhile, the expressions of CD24 mRNA in HPAEC and IL-6 mRNA in HPASMC of hypoxia group were increased significantly (P＜0.05). MTS results showed that HMGB1 inhibited the proliferation of HPAEC at 345 pmol/L significantly (P＜0.01). HMGB1 had no effect on the proliferation of HPASMC. Wound healing test showed that HMGB1 had no significant effect on HPASMC and HPAEC. CONCLUSION: HMGB1 was produced by HPAEC and HPASMC induced by hypoxia. HMGB1 induces endothelial barrier dysfunction by inhibiting HPAEC proliferation. Hypoxia stimulates HPASMC to produce inflammatory cytokines.

Isolation, fine structure and morphology studies of galactomannan from endosperm of Gleditsia japonica var. delavayi.

The water-soluble polysaccharides extracted from endosperm of Gleditsia japonica var. delavayi seeds (EGSP) were identified as galactomannan having the M/G ratio of 2.54-2.66 and a weight average molecular weight (Mw) of 1913 kDa. The molecular structure of EGSP was determined by periodate oxidation, Smith degradation, methylation, FTIR and NMR spectroscopy. The main chain is composed of ß-1,4-d-mannopyranose and the branches composed of single α-1-d-galactopyranose. We had also established a model to speculate the fine structure of galactomannan molecules and given preliminary results. The I2-KI test indicated that there were many branches on the EGSP backbone and no starch in EGSP. The CD spectra and Congo red test showed EGSP was random coil conformations in solution and could form a small quantity of helical conformation under alkaline conditions. The microstructure of morphology was observed by OM, SEM and AFM. The results showed that the fibers composed of multiple microfibers formed a network construction by entangling with each other.

Lysophosphatidic acid enhances human umbilical cord mesenchymal stem cell viability without differentiation via LPA receptor mediating manner.

Human umbilical cord mesenchymal stem cells (hUC-MSCs) are potential stromal cells which are regarded as the most feasible stem cell group in cell therapy. The maintenance of cell survival without differentiation is important in cell transplantation and stem cell therapy. However, negative factors exist in cell transplantation. Lysophosphatidic acid (LPA) is a non-antigenic small molecule phospholipid which induced several fundamental cellular responses, such as cell proliferation, apoptosis and migration. In this study we aimed to explore the effects of LPA on the survival and differentiation of MSCs and its availability in cell therapy. We found that LPA stimulated hUC-MSC proliferation and protected hUC-MSCs from lipopolysaccharide (LPS) induced apoptosis. We also observed that CD29, CD44, CD73, CD90 and CD105 were expressed, whereas CD34 and CD45 were not expressed in hUC-MSCs, and these makers have no change in LPA containing medium, which indicated that LPA accelerated the survival of hUC-MSCs in an undifferentiating status. We also demonstrated that higher expressed LPAR1 involved in LPA stimulated cell survival action. LPA stimulated cell proliferation was associated with LPAR1 mediated Gi/o-proteins/ERK1/2 pathway. On the other hand, LPA protected hUC-MSCs from LPS-induced apoptosis through suppressing caspase-3 activation by LPAR1 coupled with a G protein, but not Gi/o or Gq/11 in hUC-MSC. Collectively, this study demonstrated that LPA increased the proliferation and survival of hUC-MSCs without differentiation through LPAR1 mediated manner. Our findings provide that LPA as a anti-apoptotic agent having potential application prospect in cell transplantation and stem cell therapy.

[Expressions of G2A and OGR1 in peripheral blood cells of patients with hypoxia induced pulmonary hypertension].

OBJECTIVE: To detect the expression changes of proton-sensing receptor G2 accumulation (G2A) and ovarian cancer G protein-coupled receptor 1(OGR1) in human peripheral blood cells in hypoxic pulmonary hypertension patients(HPH). METHODS: Thirty-one patients with HPH were enrolled for HPH group(16 men and 15 women,age:(65.19±5.86) and thirty healthy persons were enrolled for the control group (NC group). The peripheral blood samples were collected and the mRNA expressions of G2A and OGR1 were determined by using real-time fluorescent quantitative PCR. The serum levels of tumor necrosis factor-α (TNF-α) were detected by using enzyme-linked immunosorbent assay (ELISA). RESULTS: PaCO2 was increased significantly in HPH group than that of the NC group (P<0.05). Forced expiratory volume in 1 sencond(FEV1)pro% and FEV1/forced vital capacity(FVC)in HPH group were significant lower than those of the NC group(P<0.05). The expressions of peripheral blood G2A mRNA and TNF-α in HPH group were increased dramatically than those of the NC group(P<0.05). The expressions of OGR1 mRNA in peripheral blood had no difference between HPH group and NC group. The expressions of G2A and TNF-α in HPH group were positively related to pulmonary artery pressure significantly. CONCLUSIONS: The expression of proton-sensing receptor G2A and the level of TNF-α are increased in peripheral blood cells of patients with pulmonary hypertension.The expressions of TNF-α,G2A and pulmonary artery pressure have positive correlation in the HPH group.

The influence of genetic polymorphisms in TLR4 and TIRAP, and their expression levels in peripheral blood, on susceptibility to sepsis.

The present study aimed to investigate whether genetic polymorphisms in the Toll-like receptor (TLR)-4 and Toll/interleukin-1 receptor (TIR)-associated protein (TIRAP) genes, and/or their expression levels, influence the susceptibility of a patient to sepsis. A total of 106 patients with sepsis were divided into two groups on the basis of their acute physiology and chronic health evaluation (APACHE) II scores: i) Sepsis group A (APACHE II <20) and ii) Sepsis group B (APACHE II >20). In addition, 100 healthy volunteers were enrolled into the control group. Polymerase chain reaction-restriction fragment length polymorphism assay was used to detect the following genetic polymorphisms: The Ser180Leu allele of the TIRAP gene and the Asp299Gly and Thr399I1e alleles of the TLR4 gene. Furthermore, the protein expression levels of TLR4 and TIRAP were analyzed using an enzyme-linked immunosorbent assay. Genetic polymorphisms were not detected for the TLR4 and TIRAP genes; however, the protein expression levels of TLR4 and TIRAP differed significantly between the control, sepsis A and sepsis B groups (P<0.01). An APACHE II score of 20 was used as a baseline in order to differentiate sepsis severity. Pearson analysis demonstrated that TLR4 and TIRAP protein expression levels were positively correlated with sepsis severity (r=0.931 and 0.972; P<0.05), and TLR4 protein expression levels were positively correlated with those of TIRAP (r=0.936; P<0.05). The results of the present study suggested that the protein expression levels of, but not genetic polymorphisms in, TLR4 and TIRAP were associated with the severity of sepsis.

[Role of endothelial progenitor cell transplantation in rats with sepsis].

OBJECTIVE: To investigate the role of endothelial progenitor cells (EPCs) transplantation in rats with sepsis induced by endotoxin (lipopolysaccharides, LPS). METHODS: Sixty clean grade Sprague-Dawley (SD) rats with genetic background were divided into three groups according to random number table method: control group, model group, and EPCs transplantation group, with 20 rats in each group. The sepsis model was reproduced by intravenous delivery of LPS 5 mg/kg. Rats in control group were injected with the same amount of normal saline. EPCs were isolated, and cultured and identified were fluorescently labeled with the green fluorescent protein (GFP) adenoviral transfection method. The EPC transplantation group was injected with LPS, then a fluorescently labeled EPCs suspension was injected via the tail vein 1 hour later. The expression of fluorescent markers of EPCs was detected with both small animal in vivo imaging instrument and frozen section. Seven days after transplantation, abdominal aorta blood was collected to determine interleukins (IL-6 and IL-10) in peripheral blood with enzyme linked immunosorbent assay (ELISA), and the lung, liver, and kidney tissues were harvested, the wet/dry ratio of the lung (W/D) was calculated, and hematoxylin and eosin (HE) staining was performed to observe, the change in histopathology. Toll-like receptor 4 (TLR4) mRNA expression in lung, liver, and kidney tissues was determined with real-time reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The positive rate of EPCs cells with double marking of CD133 and CD34 was 99.0% at the 5th generation of subculture by using flow cytometry. After the transplantation of EPCs labeled with the green fluorescent protein, the appearance of fluorescence indicated that EPCs were mainly localized in the chest, and a stronger fluorescence was observed near the blood vessels. EPCs transplantation could significantly reduce the inflammatory cell infiltration and cell damage in lung, liver, and kidney tissue in septic rats. Compared with control group, the expression of IL-6 and IL-10 in the peripheral blood, W/D ratio, and TLR4 mRNA in lung, liver, and kidney were increased significantly in the model group. Compared with model group, the expressions of IL-6 and IL-10 in the peripheral blood were significantly reduced after EPCs transplantation [IL-6 (µg/L): 2.127±0.118 vs. 2.664±0.438, IL-10 (ng/L): 24.5±3.9 vs. 31.5±3.8, both P<0.01]. EPCs transplantation reduced the W/D ratio of lung, liver and kidney tissues (lung: 4.68±0.24 vs. 5.48±0.15, liver: 3.33±0.11 vs. 3.94±0.09, kidney: 4.08±0.20 vs. 4.84±0.21, all P<0.05], and down-regulated the expression of TLR4 mRNA (×10(3), lung: 782±131 vs. 1 136±126, liver: 39.1±14.0 vs. 69.2±8.7, kidney: 52.2±15.2 vs. 83.5±17.1, all P<0.01). CONCLUSIONS: EPCs can enter the lung, liver and kidney tissues of the rat successfully after transplantation of EPCs via vein. EPCs transplantation can down-regulate pro-inflammatory process, help to recover the balance of pro- and anti-inflammatory processes, alleviate the damage to the lung, liver, and kidney tissue significantly.