Histamine-releasing factor/translationally controlled tumor protein (HRF/TCTP)-induced histamine release is enhanced with SHIP-1 knockdown in cultured human mast cell and basophil models.
Previously, we demonstrated a negative correlation between histamine release to histamine-releasing factor/translationally controlled tumor protein (HRF/TCTP) and protein levels of SHIP-1 in human basophils. The present study was conducted to investigate whether suppressing SHIP-1 using small interfering (si)RNA technology would alter the releasability of culture-derived mast cells and basophils, as determined by HRF/TCTP histamine release. Frozen CD34+ cells were obtained from the Fred Hutchinson Cancer Research Center (Seattle, WA, USA). Cells were grown in StemPro-34 medium containing cytokines: mast cells with IL-6 and stem cell factor (100 ng/ml each) for 6-8 weeks and basophils with IL-3 (6.7 ng/ml) for 2-3 weeks. siRNA transfections were performed during Week 6 for mast cells and Week 2 for basophils with siRNA for SHIP-1 or a negative control siRNA. Changes in SHIP-1 expression were determined by Western blot. The functional knockdown was measured by HRF/TCTP-induced histamine release. siRNA knockdown of SHIP-1 in mast cells ranged from 31% to 82%, mean 65 +/- 12%, compared with control (n=4). Histamine release to HRF/TCTP was increased only slightly in two experiments. SHIP-1 knockdown in basophils ranged from 34% to 69%, mean 51.8 +/- 7% (n=4). Histamine release to HRF/TCTP in these basophils was dependent on the amount of SHIP knockdown. Mast cells and basophils derived from CD34+ precursor cells represent suitable models for transfection studies. Reducing SHIP-1 protein in cultured mast cells and in cultured basophils increases releasability of the cells.
Langdon
Jacqueline M
JM
The John Hopkins Asthma and Allergy Center, Baltimore, MD 21224, USA.
Schroeder
John T
JT
Vonakis
Becky M
BM
Bieneman
Anja P
AP
Chichester
Kristin
K
Macdonald
Susan M
SM
D001491:Basophils / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D014408:Biomarkers, Tumor / Q000494:pharmacology*; D002478:Cells, Cultured; D005434:Flow Cytometry; D006636:Histamine Release / Q000187:drug effects*; D006801:Humans; D007073:Immunoglobulin E / Q000494:pharmacology; D000072181:Inositol Polyphosphate 5-Phosphatases; D008407:Mast Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D037342:Pertussis Toxin / Q000494:pharmacology*; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000172:deficiency* / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D000090324:Tumor Protein, Translationally-Controlled 1
D014408:Biomarkers, Tumor; D034741:RNA, Small Interfering; C000715647:TPT1 protein, human; D000090324:Tumor Protein, Translationally-Controlled 1; D007073:Immunoglobulin E; D037342:Pertussis Toxin; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; C000606430:INPP5D protein, human; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases