Hematopoietic Stem Cell Transplantation Cures Oc/Oc Mice from Osteopetrosis.

Abstract Background: Infantile malignant osteopeterosis is a rare autosomal recessive disorder affecting osteoclast function. Fifty % of the affected children have a mutation in the Tcirg1 gene coding for one subunit of an osteoclast specific proton pump. This pump is used by the osteoclast for acidification to enable bone resorption. Patients with this disease have normal or elevated numbers of non-functional osteoclasts, leading to increase in bone mass. The dense, sclerotic bones cause among other things pancytopenia, hepatosplenomegaly and progressive visual loss. The only curative treatment so far is hematopoietic stem cell transplantation (SCT). Without treatment, this disease results in death of 70% of the patients by the age of 6 years. The oc/oc mouse has a mutation in the gene homologous to Tcirg1 giving rise to similar symptoms as in patients. However, previous attempts to cure these mice with SCT have been unsuccessful (M.F. Seifert and S.C. Marks, Jr, Tissue & Cell 1987). Aim: To determine if early hematopoietic SCT can cure oc/oc mice from osteopetrosis. Material and Methods: One day old litters from oc/+ breeding pairs where given a radiation dose of 400 or 600 cGy. Four hours later the pups where injected ip with 1 or 5x106 lineage depleted (lin−) bone marrow (BM) cells from normal mice. Three weeks after injection, pups were genotyped and engraftment levels (based on CD45 variant expression) were checked 3 and 12 weeks after injection. X-ray examination of mutant and normal mice were performed. Results: Irradiated oc/oc mice transplanted with normal lin− BM survive past the normal lifespan of 26 days and show normalisation of the bone structure. 18 weeks after transplantation oc/oc mice are still alive. Levels of engraftment were as follows: Engraftment in transplanted mice 400 cGy 1x106 cells 400 cGy 5x106 cells 600 cGy 5x106 cells 3 w 12 w 3 w 12 w 3 w 12 w Percentage of engrafted cells in transplanted oc/oc mice and control littermates oc/oc 15,1 38,8 30,0 48,3 21,2 48,3 wt or oc/+ 13,4 10,8 28,1 28,5 19,3 28,5 Even though they increase in weight, the oc/oc mice remain smaller then their littermates. In contrast to untreated animals where teeth eruption is delayed or absent, teeth develop but with abnormal structure and shape making them not usable. If the transplant was delayed until day 8 the mice did not survive longer than untransplanted oc/oc mice. Discussion: Previous attempts to cure these mice with transplantation were not successful with only 2 out of 29 mice surviving past normal lifespan. However, these transplants were performed with non-purified cells and without matching for tissue type. We found 400 cGy and 5 million cells to be optimal for successful transplantation of 1 day old oc/oc mice. Engraftment levels were generally higher in oc/oc mice than in transplanted oc/+ or wt littermates indicating a smaller competing stem cell pool in these mice or a survival advantage of normal transplanted cells compared to stem cells in oc/oc mice. It appears that an engraftment level of 10–15 % in enough to make the mice survive long term and correct the disease phenotype.