[The effect of retrovirus-mediated hTRT transfection into cultured oral keratinocytes].

PURPOSE: Human telomerase reverse transcriptase (hTRT) was transfected into cultured oral keratinocytes (OKC) mediated by pBABE-tert recombined retrovirus to investigate the effect on OKC lifespan. METHODS: pBABE-tert recombined retrovirus loaded with hTRT gene was amplified by transfected PT67 cells, and then transfected into cultured OKC in vitro. The positive clones of OKC were separated by puromycin and subcultured. Telomerase activity was analyzed by telomerase PCR-ELISA and PCR-PAGE. RESULTS: The hTRT positive clones of OKC showed telomerase expression, with extending lifespan to 8-9 passages. CONCLUSIONS: The hTRT transfected OKC can prolong doubly lifespan but not be immortalized, which indicates that cellular immortality mechanism is complicated and multi-controled. Telomerase activity is the key for cell immortalization but not the only impact factor.

[Construction and application of the tissue bank and database of oral mucosa precancerous lesions in the Yangtze delta].

PURPOSE: To construct a database and a tissue bank of oral mucosa precancerous lesions and to estimate the application values. METHODS: Patients in the Yangtze delta suffering oral mucosa precancerous lesions were enrolled into this study. The patients' clinical data and samples of oral precancerous mucosa, salivary and blood were collected to create a tissue bank, based on which a database was constructed using Microsoft Access software, Brower/Server structure and ASP language. RESULTS: The tissue bank and database of oral mucosa precancerous lesions were successfully built. The procedure to harvest, store and transport the samples had been standardized. The database showed good interactive interface, convenient for data collection, query and share in the internet. CONCLUSIONS: We constructed the tissue bank and database of oral mucosa precancerous lesions for the first time, which not only help preserve the biological resource of oral mucosa precancerous lesions, but also provide enormous convenience in clinical work, researching and teaching. Supported by Research Fund of Science and Technology Committee of Shanghai Municipality (08ZR1416700).

[The function of osteopontin and other cytokines in the diagnosis of Sjoigren's syndrome].

PURPOSE: To explore the relationship of serum concentrations of osteopontin(OPN), TNF-α and TGF-ß1 in Sjoigren's syndrome patients. METHODS: Forty Sjoigren's syndrome (SS) patients were determined by pathological examination, which included 15 primary SS and 25 secondary SS. Serum level of OPN, TNF-alpha and TGF-beta 1 were examined by enzyme linked immunosorbent assay (ELISA). Multiple correlation analysis was performed using SPSS13.0 software package. RESULTS: Serum OPN, TNF-alpha levels increased in SS compared with healthy subjects (P<0.05).Serum TGF-beta 1 level in secondary SS significantly increased than primary SS (P<0.05). On the contrary, serum TGF-beta1 level had no significant change compared with the controls. OPN was correlated with TGF-beta 1. CONCLUSIONS: OPN and TNF-alpha may be used as adjunctive diagnostic parameter. TGF-beta 1 can be used as an index in differential diagnosis of primary and secondary SS. OPN and TGF-beta 1 have different effects and interactions between them, demonstrating mutuality, diversity and integrity of the cytokine network.

[Study of oral precancerous lesion and oral cancer by using micro PET/CT].

PURPOSE: To study oral precancerous lesion and oral cancer by using micro PET/CT. METHODS: Thirty-nine SD rats were divided into experimental group and control group. 33 of them were raised with a 4-nitroquinoline-1-oxide (4NQO) solution with the concentration of 0.002% during the first 13 weeks, and then changed to normal water. The other 6 rats drank normal water all the time. During 25th to 30th week of the experiment, 2-Deoxy-2-[F-18]fluoro-D-glucose (FDG)-PET/CT was performed for these rats. One day after imaging, pathological examination was performed. SUVmax and T/NT were investigated according to pathological results. SAS6.0 software package was used for statistical analysis. RESULTS: There was no significant difference in SUVmax among the normal group, precancerous group and cancerous group (P>0.05). There was significant difference in T/NT(muscle, brain) between the normal group and the cancerous group (P<0.05). But there was no significant difference between the normal group and the precancerous group (P>0.05); and no significant difference between the precancerous group and the cancerous group (P>0.05). The T/NT (muscle, brain) ratios increased along with the increase of the pathologic grade of the lesions. There was no significant difference in T/NT(thyroid) among the three groups and no correlation between the T/NT(thyroid) ratios and the pathologic grade. CONCLUSIONS: Micro PET/CT, as a non-invasive technology, may contribute to the dynamic studies of the process of carcinogenesis. T/NT(muscle, brain) ratios could show the degree of lesions of rat's tongue during carcinogenesis.

Squamous cell carcinoma development in previously diagnosed oral lichen planus: de novo or transformation?

OBJECTIVES: The objectives of this study were to investigate the incidence of oral squamous cell carcinoma (OSCC) developing in lesions that were previously diagnosed as oral lichen planus (OLP), and to evaluate potential contributing factors that might be associated with an increased risk for the development of OSCC in these patients. STUDY DESIGN: We retrospectively reviewed a relatively large cohort of 518 patients with OLP who received long-term follow-up (range, 6 months-21.5 years). RESULTS: There were 353 females and 165 males. Of these, 5 (0.96%) patients developed OSCC with a mean duration of 70 months. All were females with no history of smoking or alcohol use. Four of them received corticosteroid therapy. Notably, 1 of these patients received systemic corticosteroid therapy 13 months before transformation, and died of metastatic disease 46 months after transformation. CONCLUSIONS: The incidence of OSCC developing in lesions previously diagnosed as OLP is less than 1%, and females were more commonly affected. These cases appear to represent the transformation of OLP into OSCC, however it cannot be entirely ruled out that these cases may represent de novo OSCC.

[Transfecing hTRT into normal human oral keratinocytes by liposome].

PURPOSE: To introduce human telomerase reverse transcriptase (hTRT) into normal human oral keratinocytes(OKC) to expand their life-span. METHODS: The pEGFP-hTRT was transferred into OKCs by lipofecAMINE2000 and transferring(TF). Expression of hTERT in transfectants was examined by fluorescence microscopy, laser confocal microscopy. Transfection efficiency was examined by flow cytometer. Exogenous hTERT gene was detected by RT-PCR. The transfeced cells were selected by G418 for stable expression. RESULTS: OKCs were transferred with pEGFP-hTRT by lipofecAMINE2000 and TF. The transfection efficiency was 15.34%. Exogenous hTERT gene could be detected by RT-PCR. The transfeced cells exhibited transient expression of exogenous hTERT gene. CONCLUSIONS: The recombinant vector pEGFP-hTRT can be transfected into OKC by liposome. TF can improve transfection efficiency. Transient expression of exogenous hTERT is possible in oral keratinocytes.

[Construction of recombinant pEGFP/hTRT plasmid].

OBJECTIVE: To construct a plasmid which has a reporter gene for exploring the role of human telomerase reverse transcriptase(hTRT) in in-vitro cell cultivation. METHODS: hTRT was cut by restricted enzyme from plasmid pGRN145 and inserted to plasmid pEGFP-C1 (enhanced green fluorescent protein). RESULTS: Restricted enzyme analysis and DNA sequencing showed that the sequence of the pEGFP -hTRT transgenic plasmid was correct. CONCLUSION: The recombinant vector pEGFP-hTRT has been successfully constructed, and it can be used as a transgenic plasmid in generating immortalized cell lines.