| Objective:Tumor-draining lymph nodes(TDLN) are an excellent source of Tumor-reaetive T lymphoeytes and the adoptive transfer of these cells Is capable of mediating the regression of tumors established. Mature DC is add into TDLN to investigate TDLN antitumor activity.Method:1 The monocytes collected from patient's peripheral blood were isolated by density gradient centrifugation using lymphocyte isolating solution under axenic condition. The morphological characteristics of DCs from untreated group and TNF-αgroup were observed under inverted microscope and scanning electron microscope.2 Expressions of CD1a, CD83, CD80 and CD86 on DCs of untreated group, TNF-αgroup were detected by flow cytometry.3 Expressions of CD3, CD83, CD8 and CD4 on TDLNs from patien by flow cytometry.4 Proliferation of T cells stimulated by DC, and effect on cell growth of TE1 treated with different proportion (DC :TDLN, 1:12.5, 1:25, 1:50, 1:100) and different treatment times (24h, 48h and 72h) were determined by MTT method.5 The level of IL-12 and IFN-γand TNF-a in the culture supernatant of DCs from DC lone and DCTDLN were detected by ELISA.6 Expression of Bcl-2,Bax,survivin mRNA in TE1 cells after treated with DCTDLN(1:1.25,1:;25,1:50,1:100) for 24h were detected by semi-quantitative RT-PCR.Result:1 After 5 days cultured with GM-CSF and IL-4 in vitro, human CBMC developed into immature DCs with typical morphological characteristics, the cells were small and a little dendrites on the cellular surface, and they were grown in clustering. The CBMC that were stimulated by TNF-аcytokines for 2 days developed into mature DCs with typical morphological characteristics, the cells enlarged and were irregular in shape, many thin and long dendrites on the cellular surface.2 Observed by scanning electron microscope, CBMC stimulated by TNF-аfor 2 days developed into mature DCs with typical morphological characteristics, abundance of cytoplasm with more mitochondrion and rough endoplasmic reticulum, the caryon heterochromatin were under the karyolemma, and fewer lysosome and vacuolus.3 Expressions of CD1a, CD83, CD80 and CD86 on mature DCs surface were up-regulated significantly by TNF-a, compared to the control group untreated with stimulating factor (P<0.05).4 The result of phenotype analysis showed that the expressions of CD3 on TDLNs reached 79.91±4.97% and CD83 on TDLNs reached 21.36±4.33%。5 MTT analysis showed that, DCTDLN (1:12.5, 1:25, 1:50, 1:100) can significantly inhibit the growth of TE1 cells in vitro (p<0.05). The growth inhibition is in dose-dependent and time-dependent manner.The greatest inhibit rate is 1:50 observed in the treatment group of TE1 cells by DCTDLN for 24 hours;6 MTT analysis showed that, DCTDLN (1:12.5, 1:25, 1:50, 1:100) can't significantly inhibit the growth of TE1 cells in vitro.7 DCs can enhance obviously production of IL-12, IFN-γand TNF-a expression cells in TDLN (P<0.05).8 Expression of Bcl-2, Bax, survivin mRNA on TE1 cells were up regulated after treated with different proportion of DC:TDLN (1:12.5, 1:25, 1:50, 1:100) for 24h, there was statistically significant differences compared to the control group (P<0.05).Conclusion: 1 1 It is an effective way to cultivate large amount of active DC in peripheral blood from esophageal carcinoma patients by using GM-CSF ,IL-4 and TNF-a together,which will lay a foundation for further clinical trial.2 TNF-a is important for developed into mature DCs. Expressions of CD1a, CD83, CD80 and CD86 on mature DCs surface were up-regulated significantly by TNF-a.3 IL-2 is important for CBNC developed into T cell.4 DCTDLN cell in killing TE1 capacity is greater than killing A375 capacity, TDLN may have been sensitized in vivo, to a certain degree of specificity of recognition,that is to add DC to improve TDLN pecific anti ability.5 DCTDLN (1:12.5, 1:25, 1:50, 1:100) can significantly inhibit the growth of TE1 cells in vitro (p<0.05). The growth inhibition is in dose-dependent6 Proliferation of T cells stimulated by DC, and effect on cell growth of TE1 treated with different times (24h, 48h and 72h) can significantly inhibit the growth of TE1 cells in vitro (p<0.05). The growth inhibition is in time-dependent manner.7 These studies demonstrated that specific antitumor reaetivity of TDLN cells could be enhanced significant after activated by autologo.
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