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Proline-rich inositol polyphosphate 5-phosphatase (PIPP) is one of the signal-modifying enzymes that play pivotal regulatory roles in PI3K signalling pathway. The aim of this study was to determine the role of PIPP in early development of fertilized mouse eggs, via inhibition of Akt activity and subsequent downstream signalling events. The mRNA transcript levels of endogenous PIPP and Akt1, Akt2, Akt3 were detected in G, S, G and M phases of fertilized mouse eggs by RT-PCR. Levels of exogenous PIPP, phosphorylated Akt at Ser473, dephosphorylated cdc2 at Tyr15 and levels of CCNB1, were detected respectively by immunoblotting. Changes in Akt localization were observed by fluoroimmunoassay; meanwhile, changes in activity of Akt and its downstream MPF were detected. Percentages of cells undergoing division were determined by counting, using a dissecting microscope. PIPP and Akt1 transcripts were detectable in G, S, G and M phases of fertilized mouse eggs, but Akt2 and Akt3 were not. We also observed that overexpression of PIPP in fertilized eggs decreased expression of phosphorylated Akt at Ser473 and altered membrane localization of phosphorylated Akt at Ser473 specifically. Furthermore, overexpression of PIPP resulted in decreases in mitosis-phase promoting factor activity, level of dephosphorylated cdc2 at Tyr15 and cleavage rate of fertilized mouse eggs. Our data suggest, for the first time, that PIPP may affect development of fertilized mouse eggs by inhibition of level of phosphorylated Akt at Ser473 and subsequent inhibition of downstream signal cascades. [ABSTRACT FROM AUTHOR]

Objectives: To explore the role of Oct3/4, Nanog and Sox2 in regeneration of rat tracheal epithelium. Materials and methods: An ex vivo model of rat tracheal epithelial regeneration using 5-fluorouracil (5-FU) was developed, to induce injury. Expression levels of Oct3/4, Nanog and Sox2 were examined using Western blot analysis, RT-PCR or microscopically observed immunofluorescence, and cell morphological changes were observed using HE staining, during the recovery process. Results: Oct3/4, Nanog and Sox2 were not detectable in normal tracheal epithelium. After treatment with 5-FU, the normally proliferating tracheal epithelium desquamated and only a few cells in G0 phase of the cell cycle were left on the basement membrane and Oct3/4, Nanog and Sox2 could be observed at this time. Thereafter, the number of Oct3/4-, Nanog- and Sox2-positive cells increased gradually. When the cells differentiated into ciliate cells, mucous cells or basal cells, and restored pseudostratified mucociliary epithelium, the number of Oct3/4-, Nanog- and Sox2-positive cells decreased and gradually disappeared. Conclusions: G0 phase cells with resistance to 5-FU damage expressed Oct3/4, Nanog and Sox2. This indicated that these cells were undifferentiated, but had the ability to terminally differentiate into downstream-type cells. They possessed stem cell properties. The results are consistent with Oct3/4, Nanog and Sox2-expressing cells being considered as tracheal stem cells. [ABSTRACT FROM AUTHOR]

Objectives: This study is to explore the role of Notch signalling during the regeneration of rat tracheal epithelium after injury induced by 5-fluorouracil (5-FU). Materials and methods: We developed an ex vivo model of rat tracheal epithelial regeneration using 5-FU to induce injury. Expression levels of members of the Notch signalling pathway, ABCG2, CK19, and proliferating cell nuclear antigen (PCNA) were examined by reverse transcription–polymerase chain reaction, Western blot, and immunofluorescence. One group of tracheas were cultured in the medium with a γ-secretase inhibitor or Jag-1 peptide after 5-FU treatment and another group were pre-treated with the γ-secretase inhibitor or Jag-1 peptide before 5-FU treatment. The expression changes of ABCG2, CK19, and PCNA were examined by Western blot or immunofluorescence and the morphologic changes were observed by haematoxylin and eosin stain during the recovery process. Results: Expression levels of Notch3, Jagged1, and Hey1 were increased in rat tracheal epithelial cells after treatment with 5-FU. During injury recovery, disruption of Notch signalling by treatment with the γ-secretase inhibitor reduced expression of ABCG2 and PCNA, but promoted expression of CK19, while persistent activation of Notch signalling promoted expression of ABCG2 and PCNA, but reduced expression of CK19. Under both conditions, recovery from injury was reduced. However, blocking Notch signalling prior to 5-FU treatment led to the complete blockage of recovery, while activating Notch signalling before 5-FU treatment promoted recovery. Conclusions: During tracheal epithelial regeneration, Notch signalling maintains an undifferentiated state and promotes proliferation among a population of tracheal epithelial cells. [ABSTRACT FROM AUTHOR]

Objective: The aim of this study was to evaluate the individual and combined diagnostic utility of carcinoembryonic antigen (CEA), cytokeratin 19 fragments (CK19) and HBME-1 in pleural effusions of patients with lung cancer. Study design: CEA, CK19 and HBME-1 were detected by immunocytochemistry in pleural effusions from patients with lung cancer (86 cases) and without lung cancer (40 cases). Results: CEA and CK19 expression were significantly higher in the carcinoma cell group and in three subgrouped as adenocarcinoma (AC), squamous cell carcinoma (SCC) and small cell lung cancer than in the mesothelial cell group, whereas HBME-1 expression was lower in the former group ( P < 0.01). In the subgrouped tumours, CEA expression was higher in AC than in SCC ( P < 0.05), whereas HBME-1 expression was higher in SCC than in AC ( P < 0.01). Used alone, CK19 had the highest sensitivity (95.3%) and accuracy (93.7%), whereas CEA had the highest specificity (97.5%). When combinations of antibodies were evaluated together and membrane staining with HBME-1 taken as a negative outcome, CK19 and HBME-1 gave a high diagnostic performance: sensitivity of 100.0% and accuracy of 95.2% respectively. Conclusion: A panel of CEA, CK19 and HBME-1 monoclonal antibodies proved to be suitable for distinguishing carcinoma cells from reactive mesothelial cells in pleural effusions. [ABSTRACT FROM AUTHOR]