Expression and role of Oct3/4, Nanog and Sox2 in regeneration of rat tracheal epithelium.

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.

Expression and role of Notch signalling in the regeneration of rat tracheal epithelium.

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 gamma-secretase inhibitor or Jag-1 peptide after 5-FU treatment and another group were pre-treated with the gamma-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 gamma-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.

[The expression of C-FOS, C-JUN and phosphotyrosine gene products in lung cancer].

An immunohistochemical study on C-FOS, C-JUN and phosphotyrosine (P-Tyr) cancer gene products was performed. The results showed that C-FOS had the lowest frequency of expression and P-Tyr had the highest. The positive reactions of the three cancer gene products were observed in the nucleus, nuclear membrane and cytoplasm. The expression of C-FOS in normal bronchial and alveolar tissue was 3.8% and 1.6% respectively. But in lung cancer it was 60%. The simultaneous positive expression of C-FOS and C-JUN was 56% (54 cases). Negative C-FOS and positive C-JUN was 32%. Positive C-FOS and negative C-JUN was less than 4% (4 cases). Although C-FOS and C-JUN formed a hetero-dimer by zipper structure, the C-FOS had the ability of single expression.

[Observations on the relation of gastric carcinoma to Epstein-Barr virus].

Epstein-Barr virus (EBV) has been detected in the tumor cells of some cases of gastric carcinoma. The monoclonal expansion of EBV contained in tumor cells and increased serum antibody titers suggest that gastric carcinoma may be associated with EBV infection. It has been known for many years that EBV is associated with nasopharyngeal carcinoma which has a high incidence rate in China. The relation of EBV infection to gastric carcinoma is not clear. 110 cases of gastric carcinoma of Chinese patients were studied by use of EBV encoded small RNA (EBER-1) in situ hybridization. EBV was detected in 3 of the 49 cases (6.1%) from Shenyang in north China and 1 of the 61 cases (1.6%) from Changsha in south China. The EBV positive rate in gastric carcinoma of Chinese is not as high as it is in Americans and Japanese.

[Studies on the chemical constituents of Lonicera macranthoides Hand. -Mazz].

Three bisdesmosidic triterpenoid saponins I-III, have been isolated from the flowers of Lonicera macranthoides Hand. -Mazz. By spectral (IR, MS, 1H-NMR and 13C-NMR) and chemical methods, I was proved to be identical with the known dipsacoside B, II and III were new compounds named macranthoidin A and macranthoidin B. Their structures were identified as follows: The prosapogenin of II, macranthoside A(IIb), was elucidated as 3-O-beta-D-glucopyranosyl-(1-3)-alpha-L-rhamnopyranosyl-(1-2)-alpha-L- arabinopyranosyl 3 beta, 23-dihydroxyl-olean-12-en-28-olic acid, II was established as 3-O-beta-D-glucopyranosyl-(1-3)-alpha-L-rhamnopyranosyl-(1-2)-alpha-L- arabinopyranosyl 3 beta,23-dihydroxyl-olean-12-en-28-O-beta-D-glucopyranosyl-(1-6)-beta-D- glucopyranoside. The prosapogenin of III, macranthoside B(IIIb), was elucidated as 3-O-beta-D-glucopyranosyl-(1-4)-beta-D-glucopyranosyl-(1-3)-alpha-L- rhamnopyranosyl-(1-2)-alpha-L-arabinopyranosyl 3 beta,23-dihydroxyl-olean-12-en-28-olic acid, III was established as 3-O-beta-D-glucopyranosyl-(1-4)-beta-D-glucopyranosyl-(1-3)-alpha-L- rhamnopyranosyl-(1-2)-alpha-L-arabinopyranosyl 3 beta,23-dihydroxyl-olean-12-en-28-O-beta-D-glucopyranosyl-(1-6)-beta-D- glucopyranoside. IIb and IIIb have not been reported in the literature.

[Synthesis and antibacterial action of 1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(4-aroylthiocarbamoyl-1-piperaziny l)-3-quinoline carboxylic acids].

Thirteen new 1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(4-aroyl-thiocarbamoyl-1-piperazinyl)-3- quinoline carboxylic acids were prepared. Their structures were characterized by elemental analysis, IR, HNMR and MS spectra. Preliminary pharmacological tests indicated that some of compounds Ia-m possess strong inhibiting activity against Escherichia coli, Bacillus subtilis and Proteus at concentration of 100 micrograms/ml.

Protective effects of fulvotomentosides on cadmium-induced hepatotoxicity.

Fulvotomentosides (Ful) is the total saponins of Lonicera fulvotomentosa. In the present study, we examined the effects of Ful on cadmium (CdCl2)-induced acute liver injury in mice. Ful pretreatment (150 mg.kg-1, sc x 3 d) remarkably decreased CdCl2 (3.7 mg Cd.kg-1, iv)-induced liver damage as indicated by serum activities of alanine aminotransferase and sorbitol dehydrogenase. Distribution of Cd to 12 organs and hepatic subcellular fractions was determined 2 h after Cd challenge. Ful pretreatment did not produce a marked shift in the distribution of Cd to various organs, but markedly altered the hepatic subcellular distribution of Cd, with more Cd bound to metallothionein (MT) in the cytosol, less in the nuclear, mitochondrial, and microsomal fractions. Ful pretreatment produced a dose-dependent increase in hepatic MT as determined by the Cd.hemoglobin assay. In conclusion, Ful protected against Cd hepatotoxicity by inducing MT, which binds Cd in the cytosol and lowers the amount of Cd available to other critical organelles and proteins.

Protective effects of fulvotomentosides on acetaminophen-induced hepatotoxicity.

Fulvotomentosides (Ful) is the total saponins of Lonicera fulvotomentosa. In the present study, we examined the effect of Ful on acetaminophen (AA)-induced hepatotoxicity in mice. Ful pretreatment (75-225 mg.kg-1, sc x 3 d) significantly decreased AA (500 mg.kg-1, ip)-induced liver damage as indicated by serum activities of alanine aminotransferase and sorbitol dehydrogenase. Ful pretreatment (225 mg.kg-1, sc x 3 d) decreased hepatic cytochrome P-450, cytochrome b5, and NADPH-cytochrome c reductase by approximately 15-20%. Microsomes from Ful-pretreated mice, incubated in vitro with AA, produced less AA-glutathione. A 28% increase in urinary excretion of AA-glucuronide was observed in Ful (150 mg.kg-1, sc x 3 d) pretreated mice. Ful pretreatment had no influence on liver UDP-glucuronic acid concentration, but increased hepatic glucuronyltransferase activity towards AA. In summary, Ful pretreatment protects against AA-induced hepatotoxicity. One of the mechanisms for this protection appears to be the decreased AA toxic activation via P-450, as well as increased detoxication via glucuronidation of AA.

[Establishment of immunohistochemical technique for polyamine and its application in research of lung cancer].

Antiputrescine, antispermidine and antispermine were used to detect polyamine in normal and cancer tissues of lung by PAP method. Sections of lung cancer, which were known to have high concentration of polyamine, were used as positive control. Results showed highly significant difference in the staining between the experimental and control groups. Immunohistochemical assay further demonstrated that polyamine located mainly in the nucleus, nucleolus and cytoplasm of the cancer cells and was evenly distributed. The staining results of normal lung tissue and epithelium of bronchi were negative or weakly positive. The morphological finding of this experiment is considered important for further study on the correlation of polyamine with the differentiation, proliferation and malignant change of cells.

[Studies on the chemical constituents of Lonicera fulvotomentosa Hsu et S.C. Cheng].

Five saponins (I-V) have been isolated from the flowers of Lonicera fulvotomentosa Hsu et S.C. Cheng (Caprifoliaceae). This paper reports the structural determination of V, a new triterpenoid saponin named fulvotomentoside A, and characterization of I and II. Fulvotomentoside A, C58H94O26, was obtained as white crystalline needles, mp 215-7 degrees C, [a]D27.5-14.9 degrees (c 0.98, MeOH). Its structure was elucidated to be 3-O-beta-D-xylopyranosyl-(1-3)-alpha-L-rhamnopyranosyl-(1-2)- alpha-L-arabanopyranosyl-hederagenin-28-O-beta-glu cop yranosyl-(1-4)-beta-D- glucopyranoside mainly by spectroscopic analysis (IR, MS, 1H and 13CNMR) and chemical degradation. I, C41H66O12, mp 259-262 degrees C, and II, C46H74O16, mp 222-7 degrees C, were identified to be alpha-hederin and sapindoside B, respectively.