Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties. However, the interplay between endosomal phosphoinositides metabolism and innate immunity is yet to be fully understood. Here, our findings highlight the evolutionary continuity of RAB-10/Rab10's involvement in regulating innate immunity. Upon infection of Caenorhabditis elegans with Pseudomonas aeruginosa, an increase in RAB-10 activity was observed in the intestine. Conversely, when RAB-10 was absent, the intestinal diacylglycerols (DAGs) decreased, and the animal's response to the pathogen was impaired. Further research revealed that UNC-16/JIP3 acts as an RAB-10 effector, facilitating the recruitment of phospholipase EGL-8 to endosomes. This leads to a decrease in endosomal phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and an elevation of DAGs, as well as the activation of the PMK-1/p38 MAPK innate immune pathway. It is noteworthy that the dimerization of UNC-16 is a prerequisite for its interaction with RAB-10(GTP) and the recruitment of EGL-8. Moreover, we ascertained that the rise in RAB-10 activity, due to infection, was attributed to the augmented expression of LET-413/Erbin, and the nuclear receptor NHR-25/NR5A1/2 was determined to be indispensable for this increase. Hence, this study illuminates the significance of endosomal PI(4,5)P2 catabolism in boosting innate immunity and outlines an NHR-25-mediated mechanism for pathogen detection in intestinal epithelia.
Animals ; Caenorhabditis elegans/genetics* ; Endosomes/immunology* ; Caenorhabditis elegans Proteins/immunology* ; Phosphatidylinositol 4,5-Diphosphate/immunology* ; Immunity, Innate ; Pseudomonas aeruginosa/immunology* ; rab GTP-Binding Proteins/genetics* ; Diglycerides/metabolism*

OBJECTIVETo investigate the effect of Qiguiyin Decoction, QGYD) on multidrug-resistant Pseudomonas aeruginosa infection in Sprague-Dawley (SD) rats.

METHODSA pseudomonal infection model in SD rats was established by injecting multidrug-resistant P. aeruginosa intraperitoneally. Infected rats were randomized into four groups treated with Pure water, QGYD, ceftazidime, or combined QGYD and ceftazidime. Blood samples were obtained from the abdominal aorta. Serum was then collected and analyzed by peptide array for immune responsiveness to multidrug-resistant beta-lactamase proteins, including Verona integronen-coded metallo-beta-lactamase 1 (VIM-1), Sao Paulo metallo-beta-lactamase 1 (SPM-1), and Temoniera (TEMs). Blood levels of interleukin-1β (IL-1β), interleukin-4 (IL-4), and interferon-γ (IFN-γ) were assessed by enzyme-linked immunosorbent assay.

RESULTSQGYD enhanced antibody reactivity against VIM-1 [epitopes 7-11 and 36-40] and TEM-1 [epitopes 26-27, 52-55, and 66-70]. QGYD treatment restored the compromised antibody reactivity against VIM-1 [epitopes 53-54 and 56-58] and SPM-1 [epitopes 16-19 and 82-85] following pseudomonal infection. Serum levels of IL-1β and Th1/Th2 in the rats were significantly elevated following pseudomonal infection (P<0.05 orP<0.01). In contrast, QGYD and combination QGYD and ceftazidime treatment restored the elevated serum IL-1β and Th1/Th2 levels to normal (P>0.05).

CONCLUSIONSQGYD improves the immune response to pseudomonal infection in rats by stimulating the production of protective antibodies against drug-resistant proteins VIM-1, SPM-1, and TEM-1. In addition, it protects the immune system and maintains immune responsiveness by restoring IL-1β and Th1/Th2 levels.

Animals ; Antibodies, Bacterial ; blood ; Drug Resistance, Multiple, Bacterial ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Interleukin-1beta ; blood ; Male ; Pseudomonas Infections ; drug therapy ; Pseudomonas aeruginosa ; Rats ; Rats, Sprague-Dawley ; Th1 Cells ; immunology ; Th2 Cells ; immunology ; beta-Lactamases ; immunology

OBJECTIVETo explore the immuno-effect of plasmacytoid dendritic cells (pDC) on bacteria infection induced spontaneous remission (SR) of leukemia.

METHODSBoth pDC and myeloid dendritic cells (mDC) were isolated and purified from leukemic patient with SR and healthy donor by combination of immunomagnetic beads and flow cytometry. pDC were cultured in RPMI1640 medium and stimulated with different bacteria. The T cells proliferation was detected by MTT, and cytokine production by ELISA kits.

RESULTSThe human bacterial pathogen Staphylococcus aureus and Pseudomonas aeruginosa stimulation for 48 h resulted in the maturation of pDC with production of high quantity of IFN-α at (15.34 ± 2.91) ng/ml and (10.38 ± 1.41) ng/ml, respectively, comparing with that of negative group at (1.36 ± 0.13) ng/ml (P<0.01). Activated pDC could promote the differentiation of naive CD4⁺ T cells to Th1 cells with secretion of IFN-γ at (2.16 ± 0.37) ng/ml and (2.73 ± 1.11) ng/ml, respectively, comparing with that of positive control at (2.55 ± 0.23) ng/ml (P > 0.05). Activated pDC showed higher T cell stimulatory capacities [proliferation index (PI) was 4.36 and 4.05, respectively] than that of non-activated pDC (PI was 1.23 and 0.13, respectively) (P < 0.01).

CONCLUSIONStaphylococcus aureus and Pseudomonas aeruginosa activated pDC may play a key role in SR of leukemia following severe infections.

CD4-Positive T-Lymphocytes ; Dendritic Cells ; immunology ; Flow Cytometry ; Humans ; Interferon-alpha ; Leukemia ; diagnosis ; immunology ; Lymphocyte Activation ; Pseudomonas aeruginosa ; immunology ; Remission, Spontaneous ; Staphylococcus aureus ; immunology

The p38 mitogen-activated protein kinase (MAPK) plays an evolutionarily conserved role in the cellular response to microbial infection and environmental stress. Activation of p38 is mediated through phosphorylation by upstream MAPKK, which in turn is activated by MAPKKK. In the Caenorhabditis elegans, the p38 MAPK (also called PMK-1) signaling pathway has been shown to be required in its resistance to bacterial infection. However, how different upstream MAP2Ks and MAP3Ks specifically contribute to the activation of PMK-1 in response to bacterial infection still is not clearly understood. By using double-stranded RNA-mediated interference (RNAi) and genetic mutants of C. elegans, we demonstrate that C. elegans MOM-4, a mammalian TAK1 homolog, is required for the resistance of C. elegans to a P. aeruginosa infection. We have also found that the MKK-4 of C. elegans is required for P. aeruginosa resistance, but not through the regulation of DLK-1. In summary, our results indicate that different upstream MAPKKKs or MAPKKs regulate the activation of PMK-1 in response to P. Aeruginosa.
Animals ; Caenorhabditis elegans ; enzymology ; genetics ; immunology ; microbiology ; Caenorhabditis elegans Proteins ; genetics ; metabolism ; Disease Resistance ; Enzyme Activation ; MAP Kinase Kinase 1 ; metabolism ; MAP Kinase Signaling System ; Membrane Proteins ; deficiency ; genetics ; metabolism ; Mutation ; Pseudomonas Infections ; enzymology ; Pseudomonas aeruginosa ; physiology ; RNA Interference ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo investigate the inhibitory effects of Pseudomonas aeruginosa vaccine (PA-MSHA) on the proliferation of human nasopharyngeal cancer cells and explore the possible mechanism.

METHODSMTT assay was used to determine the cell growth of human nasopharyngeal cancer cell line 5-8F and 6-10B in vitro treated with the vaccine. The cell cycle distribution of the cells was detected by flow cytometry, and the expression levels of apoptosis and cycle-related proteins were evaluated by Western blotting.

RESULTSPA-MSHA treatment significantly suppressed the proliferation of 5-8F and 6-10B cells in a time- and concentration-dependent manner compared with the control group (P<0.05). The cells with PA-MSHA treatment exhibited a decreased percentage of cells entering S phase and a corresponding increase in G(1) phase cells in FACS analysis. The expression of cyclin D(1), CDK4, and CDK6 was significantly up-regulated, Bax protein up-regulated, and the anti-apoptosis protein Bcl-2 down-regulated in PA-MSHA-treated cells.

CONCLUSIONPA-MSHA can suppress the proliferation of nasopharyngeal carcinoma cell in vitro by affecting the cell cycle and promoting cell apoptosis.

Apoptosis ; drug effects ; Cancer Vaccines ; immunology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Nasopharyngeal Neoplasms ; pathology ; Pseudomonas aeruginosa ; immunology

Bacteria survive in nature by forming biofilms on surfaces and probably most, if not all, bacteria (and fungi) are capable of forming biofilms. A biofilm is a structured consortium of bacteria embedded in a self-produced polymer matrix consisting of polysaccharide, protein and extracellular DNA. Bacterial biofilms are resistant to antibiotics, disinfectant chemicals and to phagocytosis and other components of the innate and adaptive inflammatory defense system of the body. It is known, for example, that persistence of staphylococcal infections related to foreign bodies is due to biofilm formation. Likewise, chronic Pseudomonas aeruginosa lung infections in cystic fibrosis patients are caused by biofilm growing mucoid strains. Gradients of nutrients and oxygen exist from the top to the bottom of biofilms and the bacterial cells located in nutrient poor areas have decreased metabolic activity and increased doubling times. These more or less dormant cells are therefore responsible for some of the tolerance to antibiotics. Biofilm growth is associated with an increased level of mutations. Bacteria in biofilms communicate by means of molecules, which activates certain genes responsible for production of virulence factors and, to some extent, biofilm structure. This phenomenon is called quorum sensing and depends upon the concentration of the quorum sensing molecules in a certain niche, which depends on the number of the bacteria. Biofilms can be prevented by antibiotic prophylaxis or early aggressive antibiotic therapy and they can be treated by chronic suppressive antibiotic therapy. Promising strategies may include the use of compounds which can dissolve the biofilm matrix and quorum sensing inhibitors, which increases biofilm susceptibility to antibiotics and phagocytosis.
Animals ; Antibiotic Prophylaxis ; Biofilms ; drug effects ; growth & development ; Chronic Disease ; Cystic Fibrosis ; microbiology ; Drug Resistance, Microbial ; physiology ; Foreign Bodies ; microbiology ; Humans ; Microbial Consortia ; drug effects ; genetics ; immunology ; Phagocytosis ; Pseudomonas Infections ; microbiology ; Pseudomonas aeruginosa ; drug effects ; genetics ; physiology ; Quorum Sensing ; drug effects ; genetics

OBJECTIVETo evaluate the free radical generation and antioxidant enzymes status in murine peritoneal macrophage during in vitro amikacin resistant Pseudomonas aeruginosa (ARPA) treatment with different time interval.

METHODSPeritoneal macrophages were treated with 1×10(8) CFU/mL ARPA cell suspension in vitro for different time interval (1, 2, 3, 6, 12, and 24 h) and super oxide anion generation, NO generation, reduced glutathione level and antioxidant enzymes status were analyzed.

RESULTSSuper oxide anion generation and NO generation got peak at 12 h, indicating maximal free radical generation through activation of NADPH oxidase in murine peritoneal macrophages during ARPA transfection. Reduced glutathione level and antioxidant enzymes status were decreased significantly (P<0.05) with increasing time of ARPA transfection. All the changes in peritoneal macrophages after 12 h in vitro ARPA transfection had significant difference (P<0.05).

CONCLUSIONSFrom this study, it may be summarized that in vitro ARPA infection not only generates excess free radical but also affects the antioxidant system and glutathione cycle in murine peritoneal macrophage.

Amikacin ; pharmacology ; Animals ; Anti-Bacterial Agents ; pharmacology ; Antioxidants ; analysis ; Cells, Cultured ; Drug Resistance, Bacterial ; Free Radicals ; analysis ; Glutathione ; analysis ; Macrophages, Peritoneal ; immunology ; microbiology ; physiology ; Male ; Mice ; Oxidative Stress ; Pseudomonas aeruginosa ; growth & development ; immunology ; Time Factors

BACKGROUNDThe activation of triggering receptor expressed on myeloid cells-1 (TREM-1) in the presence of microbial components amplifies the inflammatory response. The aim of the present study was to investigate the effect of the modulation of the TREM-1 pathway during empyema in rats.

METHODSAdult male Wistar rats were subjected to empyema induced by intrapleural injection of Pseudomonas aeruginosa and Staphylococcus aureus. The animals were treated with LP17 (a synthetic TREM-1 inhibitor), a control peptide, or a vehicle (normal saline). Differential cell count, flow cytometry and histological examination were performed to evaluate local inflammatory alterations. Concentrations of tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6 in both pleural effusion and serum were measured by enzyme-linked immunosorbent assay.

RESULTSAlthough differential counts of each type of leukocytes in pleural effusion were not affected by LP17, a marked reduction in neutrophil numbers was seen in LP17 treated rats due to the reduction of both pleural effusion volume and total cell numbers. LP17 administration impaired concentration elevation in tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6 in both pleural effusion and serum. It was found that survival rate in LP17 treated rats was much higher than that in control rats.

CONCLUSIONThe modulation of the TREM-1 pathway by the use of LP17 appears to be beneficial during empyema in rats in attenuating pleural and systemic inflammatory responses.

Animals ; Empyema ; drug therapy ; immunology ; Male ; Peptides ; pharmacology ; therapeutic use ; Pseudomonas aeruginosa ; immunology ; Rats ; Rats, Wistar ; Receptors, Immunologic ; antagonists & inhibitors ; metabolism ; Signal Transduction ; drug effects ; Staphylococcus aureus ; immunology ; Triggering Receptor Expressed on Myeloid Cells-1

OBJECTIVETo evaluate the efficacy and safety of Pseudomonas aeruginosa MSHA (PA-MSHA) vaccine combined with TAC scheme in the treatment of breast carcinoma.

METHODSA non-blinded, randomized, controlled trial was conducted between January 2008 and June 2008 among 60 patients with breast carcinoma. The patients were randomized into control group (30 cases) with neoadjuvant chemotherapy and PA-MSHA group (30 cases) with PA-MSHA treatment in addition to neoadjuvant chemotherapy. The therapeutic effect, adverse effects, surgical approaches, postoperative complications and cost-effectiveness in both groups were analyzed before and after the treatment.

RESULTSThe response rate in PA-MSHA group were significantly higher than that in the control group at 2, 3, and 4 weeks after neoadjuvant chemotherapy (P<0.05). The Karnofsky scores underwent no significant changes in PA-MSHA group after the chemotherapy, but significantly reduced in the control group (P<0.05). The incidence and severity of the toxic reactions and the rates of subcutaneous fluid, skin flap necrosis and infection in PA-MSHA group were significantly lower than those in the control group. The rate of operation following two neoadjuvant chemotherapy sessions was significantly higher in PA-MSHA group than in the control group. The cost of neoadjuvant chemotherapy for a 1% increment of the response rate was also significantly lower in PA-MSHA than in the control group.

CONCLUSIONPA-MSHA vaccine combined with TAC scheme can significantly enhance the therapeutic effect of breast carcinoma, lowers the rate of postoperative complications, and improve the efficacy of chemotherapy.

Adolescent ; Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Breast Neoplasms ; therapy ; Cancer Vaccines ; therapeutic use ; Cyclophosphamide ; therapeutic use ; Doxorubicin ; therapeutic use ; Female ; Humans ; Middle Aged ; Neoadjuvant Therapy ; methods ; Pseudomonas aeruginosa ; immunology ; Taxoids ; therapeutic use ; Young Adult

OBJECTIVE: To express the recombinant SPLUNC1 protein in HNE1 cells and to study its function of bactericidal and binding to lipopolysaccharide (LPS). METHODS: Full length of SPLUNC1 gene was cloned into pCMV-tag4A vector and stably transfected into HNE1 cell lines, the supernatant of cell cultures was collected. After being treated with the supernatant, the Pseudomonas aeruginosa was seeded to LB soft agar plate, and the bacteria clones were counted and analyzed. For in vitro LPS binding assay, LPS was coated to 96-well plates. We incubated in the plate with SPLUNC1 protein, and detected the binded SPLUNC1 protein by ELISA. Incubating the FITC-LPS with the SPLUNC1 stably transfected or control cells, the intracellular intensity of fluorescence was observed under the fluorescence microscope. RESULTS: SPLUNC1 inhibited the bacteria clone formation obviously. Although the binding efficiency of LPS and SPLUNC1 in vitro was very low, more FITC-LPS entered into the SPLUNC1 stably transfected cells. CONCLUSION SPLUNC1 can inhibit the growth of Pseudomonas aeruginosa and bind LPS, and play an important defensive role in innate immunity of the upper airway.
Cell Line, Tumor ; Glycoproteins ; isolation & purification ; pharmacology ; Humans ; Membrane Proteins ; chemistry ; Nasopharyngeal Neoplasms ; genetics ; pathology ; Phosphoproteins ; isolation & purification ; pharmacology ; Pseudomonas aeruginosa ; drug effects ; Respiratory Mucosa ; chemistry ; immunology ; Respiratory System ; chemistry ; immunology ; Transfection