methylcellulose and Anemia--Sideroblastic

We examined the morphological and functional characteristics of erythroblasts derived from marrow erythroid progenitor cells grown in a methylcellulose microculture, which were taken from a female child with rare atypical sideroblastic anaemia (SA) partially responsive to pyridoxine. Colony formation was within the normal range in three successive cultures (median values: 82.25 CFU-E and 16.4 BFU-E derived colonies/6.6 X 10(4) cells) compared to growth by normal cells (65-315 CFU-E and 9-40 BFU-E). We evaluated in vitro differentiation by biochemical microassay of a cytosol enzyme involved in the haem pathway: uroporphyrinogen I synthase (UROS). The UROS values in the erythroid colonies from SA marrow were at the lowere end of the normal range (median values: 6.7 +/- 0.3 and 14.4 +/- 3.8 pmol uroporphyrinogen/h in CFU-E and BFU-E-derived colonies respectively versus 17.4 +/- 7.3 and 25 +/- 7.2 pmol/h in CFU-E and BFU-E colonies from normal subjects. Ultrastructural examination of the SA erythroblasts from non-cultured bone marrow or derived from cultured BFU-E revealed the characteristic deposition of iron in mitochondria around the nucleus of most cells (ringed sideroblasts). However, the majority of cultured cells had marked dyserythropoietic features, with a large number of bilobulated or trilobulated erythroblasts, multiple cytoplasmic vacuoles, numerous abnormalities of the nucleus, and excessive membrane material beneath the plasma membrane, all features difficult to observe in non-cultured marrows.

Topics: Anemia, Sideroblastic; Bone Marrow; Cells, Cultured; Culture Media; Erythroblasts; Female; Hematopoietic Stem Cells; Humans; Infant; Methylcellulose

Bone marrow cells from three patients with idiopathic acquired sideroblastic anemia and one with X-linked sideroblastic anemia were simultaneously cultured in plasma clot and methylcellulose cultures in order to evaluate their erythroid colony (CFUE) cloning potential in these two systems. In contrast to normals, sideroblastic anemia bone marrow cells demonstrated a marked ability to form erythropoietin (Epo)-independent CFUE in methylcellulose culture, and were inhibited in their ability to generate CFUE in plasma clot culture even in the presence of Epo. Addition of citrated plasma to methylcellulose cultures inhibited Epo-independent CFUE growth, but not Epo-dependent growth, by both a normal and patients' bone marrow cells. These results demonstrate that bone marrow cells from some patients with sideroblastic anemia can undergo Epo-independent and Epo-dependent CFUE growth in vitro, and that there is a marked difference in CFUE growth depending on the type of clonal culture method used. It is suggested that the culture microenvironment, plasma, and sensitivity to Epo may be contributing factors which allow specific clones of sideroblastic anemia CFUE-forming cells to proliferate in vitro.

Topics: Anemia, Sideroblastic; Blood; Blood Coagulation; Child; Child, Preschool; Culture Media; Erythropoietin; Female; Humans; Male; Methylcellulose; Middle Aged; Stem Cells