Assessment of G-CSF and GM-CSF mRNA expression in peripheral blood mononuclear cells from patients with severe congenital neutropenia and in human myeloid leukemic cell lines.
Patients with severe congenital neutropenia (SCN), also called Kostmann syndrome, are unable to generate sufficient peripheral blood granulocytes owing to an arrest of myeloid differentiation at the level of promyelocytes. Similarly, myeloid leukemic cells show a maturation arrest at different stages of myeloid maturation coupled with uncontrolled proliferation. Among other cells, defective production of or defective response to granulocyte/macrophage colony-stimulating factor (GM-CSF) or granulocyte CSF (G-CSF) might be involved in the pathophysiology of these disorders of hematopoiesis. Reverse transcription of messenger RNA and subsequent specific amplification by the polymerase chain reaction (RT-PCR) served as a sensitive technique to detect G-CSF and GM-CSF gene expression. We have tested two alternative assays for the specific quantitation of transcript levels for G-CSF. Applying one assay we could demonstrate that: 1) peripheral blood monocytes from 5 patients with SCN are able to express G-CSF and GM-CSF messenger RNA, suggesting that defective production of these factors is not responsible for the neutropenia in this condition; 2) messenger RNA levels from 5 SCN patients were on average higher than the levels determined for three healthy volunteers; 3) 7 of 9 of the examined myeloid cell lines express GM-CSF and all of them G-CSF mRNA. These results show that quantitative PCR techniques can be used as simple tools to elucidate aspects of the pathophysiology of hematologic disorders concerning the production of CSFs.
Bernhardt
T M
TM
Department of Pediatric Hematology and Oncology, Medizinische Hochschule Hannover, Germany.
Burchardt
E R
ER
Welte
K
K
D000328:Adult; D001483:Base Sequence; D002648:Child; D002675:Child, Preschool; D005260:Female; D005634:Fructose-Bisphosphate Aldolase / Q000235:genetics; D015870:Gene Expression; D016179:Granulocyte Colony-Stimulating Factor / Q000235:genetics*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D006801:Humans; D007223:Infant; D007951:Leukemia, Myeloid / Q000097:blood*; D007963:Leukocytes, Mononuclear / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D009503:Neutropenia / Q000097:blood / Q000151:congenital*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000097:blood*; D012334:RNA, Neoplasm / Q000097:blood; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D012334:RNA, Neoplasm; D016179:Granulocyte Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D005634:Fructose-Bisphosphate Aldolase