Effect Of Signal Transduction Pathway Research Articles

The effect of NF-κB on NLRP3 of lung tissue in rats with acute lung injury induced by phosgene

Objective To observe the effect of signal transduction pathway of nuclear factor-kappa B (NF-κB) on Nod-like receptor family, pyrin domain-containing 3 (NLRP3) inflammasome and pyroptosis in rats with acute lung injury induced (ALI) by phosgene. Methods The rats were randomly(random number) divided into 3 groups: air exposure control group, phosgene exposure group and PDTC group with phosgene exposure after 100 mg/kg pyrrolidine dithiocarbamate (PDTC) administration. The specimens of serum, bronchoalveolar lavage fluid (BALF) and lung tissue were collected 6 h after exposure. Morphological changes were observed by HE staining. The expression of NLRP3 in the lung of three groups was detected by immunohistochemistry. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expressions of NF-κB p65,NLRP3, ASC and caspase-1 mRNA in the lung tissue. NF-κB p65, NLRP3, ASC and caspase-1 protein levels in the lung tissue were quantified by Western blot. The concentrations of IL-1β, IL-18 and IL-33 in the serum and BALF were measured by enzyme-linked immunosorbent assay (ELISA). Pyroptosis was observed by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL). Results The model of phosgene-induced ALI was successfully established in rats. Morphological changes with inflammatory cell infiltration were observed in the lung tissues of phosgene group, in which NLRP3 positive cells also could be observed by immunohistochemical staining. The mRNA expressions and protein levels of NF-κB p65, NLRP3 and caspase-1 in lung tissues were significantly increased (P<0.05) in phosgene group, compared with air control group. The mRNA expressions and protein levels of NF-κB p65, NLRP3 and caspase-1 in lung tissues were significantly decreased (P<0.05) in PDTC group, compared with phosgene group. The IL-1β, IL-18 and IL-33 protein levels in serum and BALF were significantly increased (P<0.05) in phosgene group, compared with air control group. The IL-1β, IL-18 and IL-33 protein levels in serum and BALF were significantly decreased (P<0.05) in PDTC group, compared with phosgene group. TUNEL results showed that pyroptosis in the lung tissue obviously increased in phosgene group, while decreased in PDTC group. Conclusions NLRP3 inflammasome and lung cell pyroptosis were induced through NF-κB signal transduction pathway in rats with acute lung injury caused by phosgene inhalation. Blockade of NF-κB can alleviate acute lung injury by down-regulating the expression of NLRP3 inflammasome to inhibit pyroptosis. Key words: Nuclear factor-kappa B (NF-κB); Phosgene; acute lung injury; Pyrin domain-containing 3 (NLRP3) innammasome; Inhalation; pyroptosis; Inflammation; Pyrrolidinedithiocarbamic acid (PDTC)

The effect of signal transduction pathway of triggering receptor-1 expressed on myeloid cells in acute lung injury induced by paraquat in rats

To investigate the transduction pathway of triggering receptor-1 expressed on myeloid cells (TREM-1) in acute lung injure induced by paraquat in rats through the activating or blocking TREM-1, to observe the effect of signal transduction pathway in the acute lung injure induced-paraquat. 80 SD rats were randomly divided into normal saline control group (n=20) , PQ poisoning group (n=20) , antibody group (n=20) , and LP17 group (n=20). poisoning group, antibody group and LP17 group were given saline diluting PQ 80 mg/kg of disposable lavage after 2 h, a single set of intraperitoneal injection of anti-TREM-1 mAb (250 g/kg) , tail intravenous LP17 group synthetic peptide (3.5 mg/kg) , poisoning group was given equal normal saline intraperitoneal injection, control group given normal saline 1 mg/kg after 2 h after lavage, given the amount of intraperitoneal injection of saline solution. The expression of NF-κB in lung tissue was determined by immunohistochemistry.The levels of TNF-a, IL-10, TREM-1, and soluble TREM-1 (sTREM-1) in lung tissue and serum were measured by ELISA. Pathology changes of lung were observed under light microscope, and lung score of pathology was compared. Administration of anti-TREM-1 mAb after PQ poisoning modeling significantly increased the NF-κB expression in lung tissue at 48 h, resulting in a large number of pro-inflammatory cytokines releasing in the lung tissue and serum and lung pathology injury score increasing.Administration of LP17 after modeling significantly down-·regulated the expressions of NF-κB and proinflammatory cytokines, while led to a slight increase of anti-inflammatory cytokines and a decline of lung pathology injury score. TREM-1 may involve in inflammatory response by promoting the generation of inflammatory factors via NF-κB pathway, thus lead to lung pathological changes.

Oncogenic Raf-1 regulates epithelial to mesenchymal transition via distinct signal transduction pathways in an immortalized mouse hepatic cell line.

The epithelial to mesenchymal transition (EMT) is considered to be an important event during malignant tumor progression and metastasis. Although Raf/MEK/ERK signaling causes EMT, the mechanisms, including the signaling pathways, are as yet unclear. In the present study we have examined the effects of signal transduction pathways on oncogenic Raf-1-induced EMT, using an immortalized mouse hepatic cell line. Oncogenic Raf-1-induced EMT is characterized by down-regulation of adherens and tight junctions and the reorganization of actin. An active Raf-1 gene was introduced into a mouse hepatic cell line which was then treated with the MAP kinase inhibitor PD98059, the p38 MAP kinase inhibitor SB203580, the PI3 kinase inhibitor LY294002 or the c-Src tyrosine kinase inhibitor PP2. The expression and localization of the adherens and tight junction proteins E-cadherin, occludin, ZO-1, claudin-1 and claudin-2 were determined by western blotting, RT-PCR and immunocytochemistry. The barrier function of tight junctions was assessed by measurements of transepithelial electric resistance (TER) and permeability in terms of fluxes of [(14)C]mannitol and [(14)C]inulin. In Raf-1-transfected cells expression of occludin and claudin-2 was markedly down-regulated at the protein and mRNA levels and the TER value was decreased, while the permeability was increased. The distribution of ZO-1, pancadherin and F-actin was changed from linear to zipper-like structures at cell borders. In Raf-1-transfected cells treated with PD98059 and SB203580, but not LY294002, expression and localization of claudin-2, but not occludin, recovered, together with barrier function, measured as the TER value. The distributions of ZO-1, pancadherin and F-actin also recovered on treatment with PD98059 and SB203580, but not LY294002. Expression and localization of occludin recovered slightly on treatment with PP2. Thus, oncogenic Raf-1 regulates EMT via distinct MAP kinase, p38 MAP kinase and c-Src tyrosine kinase signal pathways in the mouse hepatic cell line.

Testing effects of signal transduction pathways on cadherin junctional complex assembly using quantitative image analysis

Cadherin adhesion molecules function in numerous cell biological processes that influence embryo development, normal cell physiology, and pathophysiology of many disease processes. Cadherins nucleate the assembly of the adherens junction, a cell-to-cell adhesion plaque that is prominent in simple epithelial cells and found in many cell types. Numerous cell biological approaches have been used to study this interesting class of molecules. Here, we outline methodology used in our studies of junctional complexes to examine effects of signaling molecules on assembly mechanisms. This is a quantitative method that allows the investigator to test the combined effect of two different signaling processes to determine whether these two signals act in concert within the same pathway. We discuss how this method could be generalized to other studies to examine consequences of various experimental manipulations on the assembly of cellular structures.

Alterations of signal transduction pathways involved in 2,3,7,8-tetrachlorodibenzo- p-dioxin-induced malignant transformation of human cells in culture

Effects of signal transduction pathways in TCDD-induced neoplastic transformation of human cells were assessed with respect to PLC-coupled signaling pathways, adenylyl cyclase-mediated responses and PKC isozyme expressions. A lower stimulation of the intracellular free calcium levels with exposure to extracellular ATP or histamine was observed in the transformed cells, as compared to the parental cells. While the steady-state level of IP3 was higher in the transformed cells, the magnitude of stimulation of I P3 generation by ATP or histamine was significantly lower in the transformed cells than the parental cells. These results indicate that a downregulation of PLC-coupled signaling pathways may be involved in the TCDD-induced transformation of human cells. While the steady-state levels of cAMP accumulation were similar between the two cell lines, treatment of PGE 2, a potent differentiation inducer, stimulated a higher accumulation of CAMP in the parental cells but isoproterenol, a typical β-adrenergic agonist, did not induce a significant difference between the two cell lines. These results suggest that desensitization of cAMP-mediated responses to extracellular signals including differentiation signals may be associated with a possible mechanism of the carcinogenesis. Elevated expression of PKC-α, -γ, -ζ, ε, -λ, and -ι were observed ion TCDD-transformed cells, indicating a possible association of altered expression of PKC isozymes with TCDD-induced transformation of human cells. The present study demonstrates that alterations of signal transduction pathways are involved in the TCDD-induced transformation of human cells and provides, a valuable basis to investigate effects of signaling pathway as a possible mechanism of TCDD-induced carcinogenesis in human cells.