Special proliferative sites are not needed for seeding and proliferation of transfused bone marrow cells in normal syngeneic mice.

Abstract

The widely held view that transfused bone marrow cells will not proliferate in normal mice, not exposed to irradiation or other forms of bone marrow ablation, was reinvestigated. Forty million bone marrow cells from male donors were given to female recipients on each of 5 consecutive days, 5 to 10 times the number customarily used in the past. When the recipients were examined 2-13 weeks after the last transfusion, donor cells were found to average 16-25% of total marrow cells. Similar percentages ofdonor cells were found when variants ofthe enzyme phosphoglycerate kinase determined electrophoretically were used for identification of donor and recipient cells. Evidence is presented that the proportion of donor cells is compatible with a linear dependence on the number of cells transfused over the range tested-i.e., million bone marrow cells injected intravenously. Special proliferative sites thus do not appear to be required.There is a long-standing presumption that intravenously transfused marrow cells proliferate only to a minimal extent, even in congeneic mice (1-3). However, Saxe et aL (4) recently reported the replacement of 10% of marrow cells by donor cells after transfusion of 100 million marrow cells in 5 daily aliquots of 20 million given intravenously, though not after intraperitoneal injection.We have confirmed the substantial proliferation of donor marrow cells after multiple large transfusions. The results appear of particular interest because they make unnecessary the assumption of special proliferative sites for stem cells [colonyforming units, spleen (CFU-S)] thought to be filled in normal marrow and thus precluding seeding of transfused CFU-S (1). MATERIALS AND METHODSMost of the experiments were with CBA mice. To trace the origin ofproliferating bone marrow cells after transfusion, bone marrows of female recipients of cells from male donors were analyzed cytologically for the presence of the Y chromosome as described (3, 5). Suspensions of bone marrow were prepared in phosphate-buffered saline and counted in a Coulter Counter, and the desired number of cells was given intravenously in 0.5 ml of phosphate-buffered saline.Further experiments were performed using donors and recipients carrying A and B alleles of the X chromosome locus PGK-1 of phosphoglycerate kinase (EC 2.7.2.3); the products of these alleles differ in electrophoretic mobility (6). The PGK-1A variant, on a predominantly C3H genetic background, was kindly donated by John West (Oxford University) and was backcrossed onto the CBA/Ca (PGK-1B) genetic background for nine generations. After electrophoretic separation of the two variant enzymes, densitometry provided the percentages of donor and host cells in the recipients' bone marrow or other organs. Full details of the method, modified from that of Buecher et aL (7), will be published elsewhere. Briefly, cell suspensions were prepared in RPMI-1640 medium, centrifuged at 150 X g, and exposed to a 10-sec distilled water shock to lyse the erythrocytes (8). After one washing ...

In vivo trafficking, cell cycle activity, and engraftment potential of phenotypically defined primitive hematopoietic cells after transplantation into irradiated or nonirradiated recipients

“…Nevertheless, our finding that graft cells home more efficiently to nonirradiated BM may partially explain the recent successes of BM transplantation in this scenario. [19][20][21][22][23][24] Better understanding of homing mechanisms may open these areas to manipulation and the possible design of protocols aimed at enhancing trafficking of transplanted HSCs to BM, which may be especially beneficial to those patients undergoing minimally ablated BM transplantation.…”

“…Furthermore, the notion that marrow spaces or "niches" need to be created for successful engraftment of donor HPCs has been recently challenged by studies demonstrating successful engraftment in nonconditioned or minimally conditioned murine and human recipients. [19][20][21][22][23][24] These studies warrant further investigation of the fate and homing patterns of transplanted HPCs in nonmyeloablated or minimally myeloablated recipients.…”

Murine marrow cellularity and the concept of stem cell competition: geographic and quantitative determinants in stem cell biology

“…Total murine cellularity GA Colvin et al addressed the question as to whether niches were a limiting factor for engraftment. 4,[13][14][15][16][17] As will be demonstrated below, considering accurate total murine cellularity numbers, which equate with total stem cell numbers, and calculating theoretical engraftment when 40 million male BALB/c marrow cells are infused into nontreated female BALB/c hosts, and comparing these theoretical engraftment figures with those actually observed in 72 independent determinations, we arrive at virtually identical values. This would indicate that essentially all marrow engraftable stem cells engraft in marrow and that the final percent chimerism is determined solely by stem cell competition.…”

“…In previous studies we, as well as others, have shown high levels of engraftment into nontreated hosts and into minimally treated hosts. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] The key to high levels of engraftment in these models has generally been infusion of high numbers of donor cells to compete with resident or residual host marrow cells. Data from these studies have suggested that marrow space might not need to be 'opened' by cytotoxic host treatment for stem cell engraftment to occur.…”

The marrow stem cell: the continuum

“…Although interpretations of results may have differed, Micklem et al 4 reported similar data in the 1960s and Brecher et al, 5 subsequently, using higher cell levels, showed that significant engraftment could be obtained. We modeled our studies on those of Brecher et al 5 We subsequently showed that therapy that would selectively reduce host stem cells without impacting significantly on marrow cellularity would markedly increase donor chimerism and allow for a reduction in the number of donor marrow cells necessary for engraftment. Exposure of mice to 100 cGy proved to be an ideal nonmyeloablative/stem cell ablative regimen.…”