Cellular localization and potential ligands of a novel scavenger receptor class B/CD36 protein homolog (Pt-SRB2) identified in the marine crab, Portunustrituberculatus.

The scavenger receptor class B family proteins (SRB) are multiligand membrane receptor proteins. Herein, a novel SRB homolog (Pt-SRB2) was identified in Portunus trituberculatus. The open reading frame of Pt-SRB2 was predicted to encode 520 amino acid residues comprising a typical CD36 domain. Phylogenetic analysis showed that Pt-SRB2 distinctly clustered with the SRB homologs of most crustaceans and Drosophila but was separate from all vertebrate CD36/SRB. Semi-quantitative and Real-time quantitative PCR revealed that the abundance of Pt-SRB2 transcripts was the highest in hepatopancreas than in other tested tissues. Overexpressed Pt-SRB2 was distributed primarily in the cell membrane and cytoplasm of HEK293T or Drosophila Schneider 2 cells. In crab hemocytes, Pt-SRB2 was distributed primarily in the cell membrane by immunofluorescence staining. In addition, the immunofluorescence staining showed that green fluorescence signals were mainly located in the inner lumen membrane of the hepatopancreatic tubules. Moreover, solid-phase enzyme-linked immunosorbent assay revealed that rPt-SRB2-L exhibited relative high affinity with lipopolysaccharides, and relative moderate binding affinity with lipoteichoic acid or peptidoglycan. Of note, rPt-SRB2-L showed high binding affinity with eicosapentaenoic acid among a series of long-chain polyunsaturated fatty acids. Taken together, this study provided valuable data for understanding the functions of the crab CD36/SRB.

Disruption of a kasB homolog gene (kasB) causes attenuation of cell invasion and virulence of Nocardia seriolae.

Nocardia seriolae is the primary aetiological agent of nocardiosis in fish, which causes mass mortality in freshwater and marine fish. ß-ketoacyl-ACP synthase (KAS) is one of the essential enzymes in the synthesis of mycolic acids (MASs) in Mycobacterium spp. and has been chosen as the target for therapeutic intervention in mycobacterial diseases. In the present study, a kasB homologue gene (kasB) was identified in the genome of N. seriolae, and the gene-deficient mutant (ΔkasB) was generated based on a clinical isolate, XSYC-Ns. Compared to the wild-type (WT) strain, the ΔkasB showed a measurably growth defect in vitro but retained the acid-fastness in acid-fast staining. Observation of the cell ultrastructure showed some alterations in the cell wall of the ΔkasB strain. Compared to its original strain, the cell wall lipid layer seemed sparser, and a wider electron-transparent zone was observed in the cell wall of ΔkasB strain. Moreover, the ΔkasB strain showed impaired ability of cell invasion as well as intracellular survival in the cell line originating from the head-kidney of the large yellow croaker (LYC-hK), compared to its original strain. In addition, the deficiency of ΔkasB significantly attenuated the virulence of N. seriolae in largemouth bass. The present study suggested that the ΔkasB gene might be involved in the synthesis of extracellular cell-wall lipids in N. seriolae and play a crucial role in its pathogenicity.

The galU gene is required for in vivo survival of pseudomonas plecoglossicida in large yellow croaker (Larimichthys crocea).

Pseudomonas plecoglossicida is an important pathogenic bacterium in aquaculture that causes visceral granulomas in large yellow croaker (Larimichthys crocea). Uridine diphosphate glucose phosphorylase encoded by galU plays a key role in biosynthesis of the bacterial envelope, particularly lipopolysaccharide and the capsule. In this study, we inactivated the galU gene in the P. plecoglossicida isolate XSDHY-P. The galU mutant strain showed impaired growth in the early exponential stage and lacked the O polysaccharide side chain in lipopolysaccharide, but almost no defect in biofilm formation was detected. The galU mutant strain also exhibited significantly more sensitivity to the bactericidal action of normal fish serum mediated by the complement system compared to the wild-type strain. In a cell model originating from the head kidney of large yellow croaker, the galU mutant strain showed lower capacities of adhesion, invasion, and intracellular survival compared to the wild-type strain. In addition, the deficiency of the galU mutant drastically decreased bacterial loads in tissues and attenuated P. plecoglossicida virulence in fish. These results suggest that the galU gene of P. plecoglossicida is required for in vivo survival in large yellow croaker.

Integrin αEß7 is involved in the intestinal barrier injury of sepsis.

BACKGROUND: IL-9-producing CD4(+) T (Th9) cell was related to acute intestinal barrier injury in sepsis. Integrin αEß7 was an important lymphocyte homing receptor on the surface of intestinal Th9 cells. However, the roles of αEß7 in the intestinal injury caused by Th9 cells were not clear in sepsis. METHODS: To investigate the roles of αEß7 in the intestinal injury caused by Th9 cells in sepsis model, the Th9 cells percentages, αEß7, E-cadherin, IL-9, and D-lactate levels in both serum and intestinal tissue were measured. The intestinal histopathology, epithelium apoptosis, and mucosal permeability measurement were also performed. The survival rate of septic rats was recorded daily for 14 days. RESULTS: Rats were assigned to four cohorts: control cohort, sepsis cohort, sepsis+αEß7i (αEß7 inhibition) cohort, and sepsis+αEß7e (αEß7 overexpression) cohort. The Th9 cells percentages, αEß7, IL-9, and D-lactate levels of the sepsis cohort were significantly higher than those of the control cohort. The levels of these variables were also elevated progressively in the sepsis+αEß7i cohort, sepsis cohort, and sepsis+αEß7e cohort. The E-cadherin levels were decreased progressively in the control cohort, sepsis+αEß7i cohort, sepsis cohort, and sepsis+αEß7e cohort. Moreover, αEß7 overexpression could decrease the 14-day survival rate. The findings of histopathology staining, apoptosis detection, and intestinal permeability test also confirmed that the barrier injury was deteriorated or relieved by elevating or decreasing the αEß7 expression levels, respectively. CONCLUSIONS: Integrin αEß7 was closely associated with the intestinal barrier injury caused by Th9 lymphocytes in sepsis.

Identification and function analysis of an immune deficiency homolog in swimming crab, Portunus trituberculatus.

The immune deficiency (IMD) pathway is involved in both antiviral and antibacterial immune responses in Drosophila. IMD protein is the key adaptor to link the extracellular signal and the intracellular reaction to initiate the signal transduction in IMD pathway. In present study, the cDNA of the IMD (Pt-IMD) was identified from a marine crab, Portunus trituberculatus. The Pt-IMD is predicted to encode 170 amino acids with a death domain. Real-Time quantitative PCR analysis showed that Pt-IMD was constitutively expressed in hemocytes, intestine, gill, heart, muscle and hepatopancreas in normal crab. Moreover, the transcript of Pt-IMD in large-granule hemocytes is approximately 6-fold higher than semi-granular cells and agranular cells. Intracellular localization showed Pt-IMD was distributed mainly in the cytoplasm when it was over-expressed in Drosophila Schneider 2 (S2) cell. Functionally, over-expression of Pt-IMD could activate the promoters of Drosophila antimicrobial peptide genes (AMPs) in S2 cell. Furthermore, Pt-IMD expression was also knock-down by RNAi to determine the function of Pt-IMD on regulation of the expression of different antimicrobial peptides (AMPs) in crab. In the primary cultured hemocytes challenged with or without Vibrio alginolyticus, after Pt-IMD was knocked-down by specific long double strand RNA, the expression of anti-lipopolysaccharide factor1 (ALF1), ALF3, crustin1, crustin3, arasin2, hyastatin1and hyastatin3 have been significantly inhibited in normal cell or bacterial infected cell, while the expression of lysozyme was normal in non-infected cells and was significantly induced in bacterial infected cells, which compared to the non-specific dsRNA treated cells.

The Distribution and Source of MRDOs Infection: A Retrospective Study in 8 ICUs, 2013-2019.

BACKGROUND: To analyze the distribution and source of MDROs infection in the ICUs and to provide a basis for formulating more effective prevention and control programs for MDROs. METHODS: A retrospective investigation was conducted on MDROs infection in 8 ICUs of a large tertiary hospital from July 2013 to June 2019. A total of 2629 strains of MDROs isolated from 1701 inpatients were selected for analysis. The MDROs of the 8 ICUs were divided into two types of four categories according to source: out-of-hospital (out-of-hospital transfer and community acquisition) and in-hospital (in-hospital transfer and department acquisition) infections. RESULTS: CRAB (41.84%) and CRE (35.07%) accounted for the majority of the infecting MDROs. The detection rates of MRSA, CRAB, CRPA and CRE were 61.24%, 83.75%, 43.01% and 30.15%, respectively. The top three infection sites of MDROs were the lower respiratory tract (81.10%), blood (6.70%) and abdominal cavity (5.80%). The out-of-hospital and in-hospital infection rates of MDROs were 50.51% and 49.49%, respectively; the out-of-hospital infection rates for MRSA, CRAB, CRPA and CRE were 43.56%, 55.91, 64.44% and 44.58%, respectively. The proportions of MRSA, CRAB, CRPA and CRE infections contracted in the department were 40.98%, 36.27%, 25.56% and 46.62%, respectively. There was a statistically significant difference between comprehensive ICU and specialized ICU wards as sources for CRAB infections (P < 0.001). CONCLUSION: The main source of MDROs in the ICU is not the hospital itself entirely. It is particularly important to strengthen the identification of MDRO sources and implement more effective and accurate infection prevention and control measures.

Effects of permissive hypocaloric vs standard enteral feeding on gastrointestinal function and outcomes in sepsis.

BACKGROUND: Intestinal mucosal barrier injury and gastrointestinal dysfunction are important causes of sepsis. However, few studies have investigated the effects of enteral underfeeding on gastrointestinal function in sepsis. Moreover, no consensus on goal enteral caloric intake has been reached in sepsis. AIM: To investigate the effects of different goal caloric requirements of enteral nutrition on the gastrointestinal function and outcomes in the acute phase of sepsis. METHODS: Patients were randomly assigned to receive 30% (defined as group A), 60% (group B), or 100% (group C) of goal caloric requirements of enteral nutrition in this prospective pilot clinical trial. The acute gastrointestinal injury (AGI) grades, incidence of feeding intolerance (FI), daily caloric intake, nutritional and inflammatory markers, and biomarkers of mucosal barrier function were collected during the first 7 d of enteral feeding. The clinical severity and outcome variables were also recorded. RESULTS: A total of 54 septic patients were enrolled. The days to goal calorie of group C (2.55 ± 0.82) were significantly longer than those of group A (3.50 ± 1.51; P = 0.046) or B (4.85 ± 1.68; P < 0.001). The FI incidence of group C (16.5%) was higher than that of group A (5.0%) or B (8.7%) (P = 0.009). No difference in the incidence of FI symptoms was found between groups A and B. The serum levels of barrier function biomarkers of group B were significantly lower than those of group A (P < 0.05) on the 7th day of feeding. The prealbumin and IL-6 levels of group A were lower than those of group B (P < 0.05) on the 7th day of feeding. No significant differences in the clinical outcome variables or 28-d mortality were found among the three groups. CONCLUSION: Early moderate enteral underfeeding (60% of goal requirements) could improve the intestinal barrier function and nutritional and inflammatory status without increasing the incidence of FI symptoms in sepsis. However, further large-scale prospective clinical trials and animal studies are required to test our findings. Moreover, the effects of different protein intake on gastrointestinal function and outcomes should also be investigated in future work.

Interleukin-9 promotes intestinal barrier injury of sepsis: a translational research.

BACKGROUND: Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to infection. Intestinal mucosal barrier injury is one of the important manifestations of sepsis. Interleukin-9 (IL-9) and IL-9-producing CD4(+) T cells were emerging pro-inflammatory mediators with development of intestinal injury. However, it is unclear whether IL-9 is related to the intestinal barrier injury of sepsis. METHODS: To investigate the roles of IL-9-producing CD4(+) T cells and IL-9 in the process of barrier injury in sepsis, serum IL-9-producing CD4(+) T cell percentages, IL-9, and D-lactate levels were measured in septic patients and controls. The markers of barrier function in serum and intestinal tissue were also collected in septic rats. Moreover, the barrier injury degree and survival rate of septic rats were also investigated after increasing or interfering with IL-9 expression. RESULTS: The serum IL-9-producing CD4(+) T cell percentages, IL-9, and D-lactate levels were significantly higher in septic patients or rats than those in controls. IL-9-producing CD4(+) T cells and IL-9 levels were positively correlated with D-lactate levels and had a high predictive value of 28-day mortality in septic patients. The non-survivors had significantly higher serum T cell percentages, IL-9, and D-lactate levels compared with survivors. In septic rats, IL-9 increased the expression levels of D-lactate, whereas that decreased the expression levels of zonula occludens 1. Moreover, the barrier injury was aggravated or alleviated by increasing or interfering with IL-9 expression, respectively. Survival rate analysis also showed that IL-9 decreased the 14-day survival rate of septic rats. CONCLUSION: IL-9 is closely related to intestinal mucosal barrier injury and mortality in sepsis. IL-9 blockade has the potential to improve the barrier injury in sepsis. TRIAL REGISTRATION: The study was registered at ClinicalTrials.gov (ID: NCT03791866, Date: December 2018).

[Seasonal variation of microflora in Sinonovacula constricta and its aquacultural pond based on high-throughput sequencing]. / åŸºäºŽé«˜é€šé‡æµ‹åºçš„ç¼¢è›åŠå ¶å »æ®–æ± å¡˜èŒç¾¤ç»“æž„çš„å­£èŠ‚å˜åŒ–.

In order to explore the seasonal changes of the bacterial community structure and the interaction of environmental factors in Sinonovacula constricta ponds, we used high throughput sequencing technology to examine the bacteria of water, sediment, and viscera. The results showed that microflora structure of water samples in winter was significantly different from that in spring, summer and autumn, while there was no significant difference in bacterial community structure of sediment and viscera in different seasons. There was no significant difference of Shannon diversity index in water across different seasons. The Shannon diversity index of sediment and viscera was the lowest in summer and the highest in winter. At the phylum level, Cyanobacteria, Proteobacteria and Tenericutes were the most predominant bacteria in water, sediment, and viscera, respectively. At the genus level, NS3a_marine_group was predominant in winter water, and Synechococcus in the other three seasons. By contrast, dominant bacteria in sediments were norank_f_Anaerolineacea and Nitrospira, and Mycoplasma and Arcobacter were the most abundant bacterial genera in viscera. Synechococcus had a positive correlation with water temperature, COD, PO4--P, NH4+-N, pH, and transparency. The norank_f_Anaerolineacea was positively correlated with water temperature, COD, and TP. Mycoplasma was positively correlated with water temperature, PO4--P, NH4+-N, pH, and transparency. Our results suggest that there were significant differences in the composition and diversity of microflora of S. constricta and ponds in different seasons. Bacteria in water was obviously affected by various environmental factors, especially water temperature and the concentrations of nitrogen and phosphorus.

Characterization, subcellular localization and function analysis of myeloid differentiation factor 88 (Pt-MyD88) in swimming crab, Portunus trituberculatus.

Myeloid differentiation factor 88 (MyD88) is a universal and essential adaptor protein required for the Toll-like receptors (TLRs) pathway activation in invertebrates as well as in vertebrates. Herein, we characterized a MyD88 (Pt-MyD88) cDNA sequence in the swimming crab (Portunus trituberculatus). The Pt-MyD88 ORF is predicted to encode 469 peptides with an N-terminal death domain and a typical C-terminal TIR domain. Real-Time quantitative PCR analysis showed that the Pt-MyD88 transcriptions were constitutively expressed in hemocytes, gill, intestine, heart and muscle in normal crab. The expressions of Pt-MyD88 would be down-regulated by V. alginolyticus or LPS challenge, and be up-regulated by WSSV infection in hemocytes. Intracellular localization showed Pt-MyD88 was distributed mainly in the cytoplasm when it was over-expressed in human cell HEK293T or in Drosophila Schneider 2 (S2). Functionally, over-expression of Pt-MyD88 could either activate the NF-κB in HEK293T cells or activate the promoters of Drosophila antimicrobial peptide genes (AMPs) in S2 cell. In primary cultured hemocytes of swimming crab, after Pt-MyD88 was knocked-down by specific long double strand RNA, the expression of anti-lipopolysaccharide factor1 (ALF1), hyastatin3, crustin1 and crustin3 have been significantly inhibited, while the expression of other AMPs is normal compared to non-specific dsRNA treated cells.

ß-actin gene expression is variable among individuals and not suitable for normalizing mRNA levels in Portunus trituberculatus.

The housekeeping gene encoding ß-actin appears to be the most widely-used internal reference for gene expression studies in experimental animals or their cell lines. However, the effectiveness of ß-actin to normalize mRNA levels expression in many crustacean species is still object of debate. To date, it is still unclear if ß-actin is suitable to be utilized as the internal reference in qualitative real-time gene expression study in crab species. To address this concern, we evaluated 5 candidate reference genes encoding ß-actin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), cyclophilin A, elongation factor 1-α (EF1-α), and 18 S ribosomal RNA (18 S rRNA) in the swimming crabs (Portunus trituberculatus) models. Our data showed that the ß-actin gene expression varied significantly across individual swimming crab individuals in gills or hemocytes and the expression of 18 S rRNA, EF1-α, cyclophilin or GAPDH gene were relatively stable compared to that of ß-actin. Moreover, the expression stability of the reference genes among different tissues in normal crabs or after WSSV challenge was also tested by geNorm and NormFinder software. Among tissues, 18 S rRNA was most stably expressed in different tissues, followed by cyclophilin A and EF1-α, compared to ß-actin and GAPDH. Upon to viral simulation, GAPDH was found to be the most stable internal control gene in gills and cyclophilin A was ranked as the most stable gene in hemocytes.

[Effects of Organic Pollutants on the Bacterioplankton Community in Hangzhou Bay].

Hangzhou Bay suffers from intensive anthropogenic disturbances and a huge amount of terrestrial inputs, and thus has become one of the most seriously contaminated coastal zones in China. There is evidence that microbes play a dominant role in pollutant biodegradation and serve as biomarkers for pollution levels. However, it remains unclear how the bacterioplankton communities respond to organic contaminants. To fill this knowledge gap, we collected surface water samples (0.5 m below the surface layer) from 13 sites across Hangzhou Bay and 8 control sites across its adjacent offshore areas. Using Illumina sequencing based on analysis of the bacterial 16S rRNA gene, we explored the effects of increasing organic pollution levels on the bacterioplankton community compositions (BCCs). The results revealed that the organic pollution level (A) in Hangzhou Bay (13.2±1.6) was significantly (P<0.001) higher than in the control zone (5.4±3.0). The distribution and diversity of bacterioplankton communities were significantly distinct between the two zones. The dominant bacterioplankton lineages in Hangzhou Bay were γ-Proteobacteria (24.4%±5.5%), α-Proteobacteria (16.5%±7.7%), and Planctomycetes (13.9%±8.6%), whereas those in the adjacent zones were Cyanobacteria (20.1%±7.5%), Bacteroidetes (18.4%±1.5%), Actinobacteria (17.5%±4.2%), γ-Proteobacteria (16.6%±1.2%), and α-Proteobacteria (14.3%±1.7%). Multivariate regression tree (MRT) analysis showed that the bacterioplankton community diversity was primarily affected by suspended particulates (SP), nitrite, oil, and organic pollutants, which respectively explained 22.0%, 6.5%, 6.0%, and 5.5% of the variance in diversity. Redundancy analysis (RDA) illustrated that the bacterioplankton community distribution was controlled by organic pollutants, COD, Chla, TN, nitrate, and salinity, which cumulatively governed 71.0% of the variation in BCCs. Organic pollutants alone controlled 6.5% variance, which was higher than any other single factor. Additionally, 35 sensitive species were identified via the indicator value method and their relative abundances were significantly associated (P<0.05 in each case) with the organic pollution level, thereby indicating their potential for evaluating coastal pollution. Collectively, our work demonstrates that BCCs are sensitive to coastal pollution and provides biomarkers for elevated pollution levels.

Identification of a novel class B scavenger receptor homologue in Portunus trituberculatus: Molecular cloning and microbial ligand binding.

Class B scavenger receptors (SRBs), which are present in mammals and insects, have been implicated in a wide range of functions. Herein, a novel SRB homologue, PtSRB, was cloned from the swimming crab, Portunus trituberculatus. PtSRB has 538 amino acid residues, and it consists of two transmembrane regions, a large extracellular loop, and two intracellular tails. A phylogenetic analysis showed that PtSRB distinctly clustered with Marsupenaeus japonicas SRB-1 and most Drosophila SRB homologues, including Croquemort, Peste, NinaD, and Santa Maria, but was separate from the Drosophila sensory neuron membrane protein, MjSRB-2, and all vertebrate SRBs. Real-time quantitative PCR analyses showed that the PtSRB gene was constitutively expressed in all tissues tested. When PtSRB was overexpressed in human embryonic kidney 293T cells, it was distributed in the membrane and cytoplasm. Moreover, in vitro assays showed that rPtSRB bound microbial lipopolysaccharide with low affinity, and lipoteichoic acid and peptidoglycan with high affinity. PtSRB transcripts were down-regulated after challenge with Vibrio alginolyticus or white spot syndrome virus, but not after a Candida lusitaniae challenge. This study provides valuable data for understanding the role of SRBs in the host defense against microbial pathogens, which will facilitate future studies of host-pathogen interactions in crabs.

Three novel Toll genes (PtToll1-3) identified from a marine crab, Portunus trituberculatus: Different tissue expression and response to pathogens.

The Toll signaling pathway is one of the most important regulators of the immune response in both vertebrates and invertebrates. Herein, three novel Toll (PtToll1-3) cDNA sequences were cloned from the swimming crab, Portunus trituberculatus. PtToll1 has 1003 amino acid residues and consists of an extracellular domain containing 15 leucine-rich repeats (LRRs) and a cytoplasmic Toll/interleukin-1 receptor (TIR) domain of 139 residues. PtToll2 encodes 1196 peptides, with an extracellular domain containing 28 LRRs and a cytoplasmic TIR domain. PtToll3 is 1229 residues long and contains 26 LRRs and a cytoplasmic TIR domain. Based on sequence and phylogenetic analyses, PtToll1 distinctly clustered with almost all crustacean Tolls, except Litopenaeus vannamei Toll3. However, PtToll2 and PtToll3 were separated from most reported crustacean Tolls, which mostly clustered with Drosophila melanogaster (Dm) Toll8, L. vannamei Toll3, and DmToll6. Reverse transcription PCR and real-time quantitative PCR analyses showed that PtToll1 and PtToll3 were constitutively expressed in all tissues tested, but PtToll2 mRNA was only highly enriched in gills. Upon challenges with Vibrio alginolyticus, Candida lusitaniae, or white spot syndrome virus (WSSV), the three Tolls exhibited different responses: the PtToll1 transcript was up-regulated in response to C. lusitaniae or V. alginolyticus challenge, but did not respond to WSSV challenge; both PtToll2 and PtToll3 mRNAs were down-regulated 12 h after C. lusitaniae or V. alginolyticus infection. However, WSSV elicited the expression of PtToll2 at 6 h post-infection, but suppressed transcription of PtToll3 at 24 h post-infection. The study provides valuable data for understanding the role of Toll pathways in the host defense against microbial pathogens, which will facilitate future studies on host-pathogen interactions in crabs.

First description and expression analysis of tumor necrosis factor receptor-associated factor 6 (TRAF6) from the swimming crab, Portunus trituberculatus.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a cytoplasmic adapter protein that mediates signals induced by the tumor necrosis factor receptor (TNFR) superfamily and the interleukin-1 receptor (IL-1R). In the present study, the full-length cDNA of TRAF6 (Pt-TRAF6) was identified in a marine crab, Portunus trituberculatus. Pt-TRAF6 ORF is predicted to encode a 599-amino acid protein, including a RING type zinc finger, two TRAF-type zinc fingers, and a meprin and TRAF homology (MATH) domain. The overall amino acid sequence identity between Pt-TRAF6 and other TRAF6s ranged from 50.9 to 51.3% for shrimp and from 16.1 to 19.4% for insects. The Pt-TRAF6 gene contains six exons and five introns, which is different from the organization of the insect TRAF6 gene. Pt-TRAF6 transcripts were broadly expressed in all tissues tested, and their expression was higher in hemocytes, gills, the intestine, and heart than in muscle. Interestingly, the level of Pt-TRAF6 transcript differed between male and female crabs. After Vibrio alginolyticus or lipopolysaccharide (LPS) challenge, the Pt-TRAF6 transcript was down-regulated in hemocytes and up-regulated in gills. Moreover, Pt-TRAF6 expression was altered sooner in the LPS challenge group than in the V. alginolyticus challenge group. These results indicate that Pt-TRAF6 may respond to Gram-negative bacterial infections.

Efficacy of linezolid on gram-positive bacterial infection in elderly patients and the risk factors associated with thrombocytopenia.

UNLABELLED: Objective : Linezolid is active against drug-resistant gram-positive bacteria. However, the efficacy and safety of linezolid in the treatment of the elderly have not been well characterized. The purpose of this study was to evaluate the efficacy of linezolid in the treatment of the elderly with gram-positive bacterial infection and to investigate the risk factors associated with the development of thrombocytopenia in these patients. METHODOLOGY: This was a retrospective analysis of 50 elderly patients who were treated with intravenous linezolid for gram-positive bacterial infection. Clinical data and bacteriological responses were assessed. Risk factors associated with thrombocytopenia in elderly patients were analyzed. RESULTS: The overall clinical cure rate of linezolid was 74%, and the bacteriological eradication rate was 69%. Thrombocytopenia occurred in 24 patients, and thrombocytopenia was associated with both the duration of treatment (P = 0.005) and the baseline platelet count (P = 0.042). Based on a logistic regression analysis, the baseline platelet count <200×10(9)/L (OR = 0.244; 95% CI = 0.068- 0.874; P = 0.030) was identified as the only significant risk factor for linezolid-associated thrombocytopenia in elderly patients. The mean platelet count decreased significantly from the 7(th) day of treatment, and decreased to the lowest value 1-2 days after the end of therapy. Conclusions : Linezolid is effective and safe for the elderly with gram-positive bacterial infections. Adverse effects such as thrombocytopenia are of greater concern. Platelet counts should be monitored in patients who are treated with linezolid and that measures should be taken in advance to avoid hemorrhagic tendencies.

Ruscogenin attenuates monocrotaline-induced pulmonary hypertension in rats.

Inflammation, endothelial dysfunction, and thrombosis contribute to the pathogenesis and development of human pulmonary arterial hypertension (PAH). The aim of this study was to investigate the effects of ruscogenin, a natural anti-inflammatory and anti-thrombotic agent, on the development of monocrotaline (MCT)-induced PAH in rats. Our results revealed that ruscogenin had favorable effects on hemodynamics and pulmonary vascular remodeling, preventing the development of PAH 3 weeks after MCT. In addition, ruscogenin resulted in markedly reduced expression of inflammatory cytokine and leukocyte infiltration via the inhibition of nuclear factor (NF)-κB activity in rat lungs. Ruscogenin also attenuated MCT-induced endothelial cell apoptosis in the remodeled pulmonary arterioles and rescued destruction of endothelial cell membrane proteins such as eNOS, caveolin-1, and CD31. Our findings suggest that ruscogenin might have therapeutic benefits for PAH patients.

Inhibition of COX-2 and activation of peroxisome proliferator-activated receptor gamma synergistically inhibits proliferation and induces apoptosis of human pancreatic carcinoma cells.

Although inhibition of cyclooxygenase-2 (COX-2) or activation of peroxisome proliferators-activated receptor gamma (PPAR-gamma) leads to growth inhibition in malignancies, the synergistic anti-tumor effects of combination of COX-2 inhibitor (NS-398) and PPAR-gamma agonist (rosiglitazone) on the human pancreatic cancer cells remains unknown. Here, we evaluated the effects of NS-398 and/or rosiglitazone on the cell proliferation and apoptosis in a pancreatic cancer cell line, SW1990. NS-398 and rosiglitazone decreased cell proliferation in a dose- and time-dependent manner. Proliferating cell nuclear antigen (PCNA) labeling index significantly decreased in the cells treated with either NS-398 or rosiglitazone. Both NS-398 and rosiglitazone alone induced apoptotic cell death of SW1990. The combination of NS-398 and rosiglitazone exerted synergistic effects on proliferation inhibition, and apoptosis induction in SW1990 cells, with down-regulation of Bcl-2 and up-regulation of Bax expression. Our results indicate that simultaneous targeting of COX-2 and PPAR-gamma inhibits pancreatic cancer development more effectively than targeting each molecule alone.

Blockade of cholecystokinin-2 receptor and cyclooxygenase-2 synergistically induces cell apoptosis, and inhibits the proliferation of human gastric cancer cells in vitro.

Gastrin and cyclooxygenase-2 (COX-2) play important roles in the carcinogenesis and progression of gastric cancer. However, it remains unknown whether the combination of cholecystokinin-2 (CCK-2) receptor antagonist plus COX-2 inhibitor exerts synergistic anti-tumor effects on human gastric cancer. Here, we demonstrated that the combination of AG-041R (a CCK-2 receptor antagonist) plus NS-398 (a selective COX-2 inhibitor) treatment had synergistic effects on proliferation inhibition, apoptosis induction, down-regulation of Bcl-2 and up-regulation of Bax expression in MKN-45 cells. These results indicate that simultaneous targeting of CCK-2 receptor and COX-2 may inhibit gastric cancer development more effectively than targeting either molecule alone.