Generation of T lymphocytes from the epithelium and stroma of squamous pre-neoplastic lesions of the uterine cervix

In this study, we have developed a simple and efficient technique for the i solation of viable lymphocytes from the epithelium and stroma of small pre- neoplastic squamous intraepithelial lesions (SIL) of the uterine cervix. Fo llowing the separation of the epithelium from the stroma using dispase II, both biopsy fragments were used to generate T lymphocytes. The stroma-deriv ed lymphocytes were obtained by collecting and culturing the cells migratin g out of the biopsy in the presence of IL2 (50 U/ml). An average of 0.7 x 1 0(6) and 1.4 x 10(6) lymphocytes could be obtained after 20 and 30 days of culture, respectively. For the expansion of lymphocytes derived from the pr e-neoplastic epithelium (SIL) it was necessary to use a combination of irra diated peripheral blood mononuclear cells (PBMC) as a feeder layer with PHA (0.1%), in addition to IL2 (50 U/ml). Interestingly, these lymphocytes cou ld be obtained using either allogeneic or syngeneic PBMCs. With this protoc ol, we were able to generate up to 100 x 10(6) lymphocytes from the epithel ium, the majority of which were T lymphocytes. (C) 1999 Elsevier Science B. V. All rights reserved.