Contributions
Abstract: PB1900
Type: Publication Only
Background
We previously reported that human interleukin 1-beta (IL-1-b) stimulated bone-marrow stromal myofibroblasts to express a hematopoietic molecule CD34. We also reported that a some fraction of myofibroblasts from bone marrow of acute myelogenous leukemia (AML) patients and from chronic myelogenous leukemia (CML) ones showed characteristics of original AML blasts and CML cells, such as chromosomal translocations and the expression of myeloid markers. However, when myofibroblasts were not separated and cultured totally, that is, bulk cultures mixed with normal and leukemic myofibroblasts, leukemia-specific markers were detected in DNA level, but not expressed in RNA levels for a long term cultures.
Aims
We showed in this report that myofibroblasts positive for leukemic markers was dormant and G0 phase in the mixed normal and leukemic myofibroblasts, and when IL-1-beta was added in the cultures, the fusion transcript was observed.
Methods
Bone marrow samples were collected from informed CML patients, which were separated with density centrifugation method. Cells were cultured in DMEM with 20% FCS to prepare myofibroblasts. The obtained myofibroblasts were cultured for one month, and FISH was analyzed with bcr and abl probes. Cells were further cultured in DMEM/F12 medium supplemented with 20% KSR and with recombinant human IL-1-b for one week. The morphological changes and the expression of specific genes were observed.
Results
When bone marrow-derived stromal myofibroblast obtained from CML patients were cultured in an ordinary DMEM with 10% FCS medium, FISH analysis detected 1-5% BCR-ABL fusion chromosome in the cultures; however, RT-PCR analyses revealed that leukemia-specific Bcr-Abl transcript was not detected. When CML-derived myofibroblasts were cultured in DMEM/F12 medium with IL-1-beta for one week, Bcr-Abl transcript was detected with RT-PCR analysis. And, CD41 molecule and GATA-2 transcription factor were also detected in the cultures.
Conclusion
Serum-free culture with IL-1-beta induced activation of CML-derived myofibroblasts that were dormant in the ordinary culture. In this system hematopoietic progenitor-cells may promote to grow.
Session topic: 7. Chronic myeloid leukemia – Biology & Translational Research
Keyword(s): BCR-ABL, Chronic myeloid leukemia, Stromal cell
Abstract: PB1900
Type: Publication Only
Background
We previously reported that human interleukin 1-beta (IL-1-b) stimulated bone-marrow stromal myofibroblasts to express a hematopoietic molecule CD34. We also reported that a some fraction of myofibroblasts from bone marrow of acute myelogenous leukemia (AML) patients and from chronic myelogenous leukemia (CML) ones showed characteristics of original AML blasts and CML cells, such as chromosomal translocations and the expression of myeloid markers. However, when myofibroblasts were not separated and cultured totally, that is, bulk cultures mixed with normal and leukemic myofibroblasts, leukemia-specific markers were detected in DNA level, but not expressed in RNA levels for a long term cultures.
Aims
We showed in this report that myofibroblasts positive for leukemic markers was dormant and G0 phase in the mixed normal and leukemic myofibroblasts, and when IL-1-beta was added in the cultures, the fusion transcript was observed.
Methods
Bone marrow samples were collected from informed CML patients, which were separated with density centrifugation method. Cells were cultured in DMEM with 20% FCS to prepare myofibroblasts. The obtained myofibroblasts were cultured for one month, and FISH was analyzed with bcr and abl probes. Cells were further cultured in DMEM/F12 medium supplemented with 20% KSR and with recombinant human IL-1-b for one week. The morphological changes and the expression of specific genes were observed.
Results
When bone marrow-derived stromal myofibroblast obtained from CML patients were cultured in an ordinary DMEM with 10% FCS medium, FISH analysis detected 1-5% BCR-ABL fusion chromosome in the cultures; however, RT-PCR analyses revealed that leukemia-specific Bcr-Abl transcript was not detected. When CML-derived myofibroblasts were cultured in DMEM/F12 medium with IL-1-beta for one week, Bcr-Abl transcript was detected with RT-PCR analysis. And, CD41 molecule and GATA-2 transcription factor were also detected in the cultures.
Conclusion
Serum-free culture with IL-1-beta induced activation of CML-derived myofibroblasts that were dormant in the ordinary culture. In this system hematopoietic progenitor-cells may promote to grow.
Session topic: 7. Chronic myeloid leukemia – Biology & Translational Research
Keyword(s): BCR-ABL, Chronic myeloid leukemia, Stromal cell