Biomedical subjects

H Hisha

Publications and source records attributed to H Hisha.

At least 19 recordsLinked to original sources

PubMed2002

Optimal protocol for total body irradiation for allogeneic bone marrow transplantation in mice.

We have previously demonstrated, using chimeric resistant MRL/lpr mice, that a fractionated total body irradiation (FTBI) (5 Gy x 2 with a 4 h interval on the day before allogeneic bone marrow transplantation (BMT)) is the best conditioning regimen for the treatment of autoimmune diseases in radiosensitive MRL/lpr mice. In the present study, using various standard strains of mice (not radiosensitive mice), we explore the best protocol for irradiation (doses and intervals) as the conditioning regimen for allogeneic BMT. Recipient mice were exposed to various irradiation regimens: a single total body irradiation (TBI) of 9.5 or 12 Gy and FTBI of (5+5) Gy to (7+7) Gy with a 1 to 24 h interval. The method generally utilized for humans ((2+2) Gy with a 4 h interval for 3 days (total 12 Gy)) was also used. One day after the last irradiation, donor BMCs from BALB/c, C3H, or C57BL/6 (B6) mice were transplanted into C3H or B6 mice. The irradiation protocol of (2+2) Gy for 3 days was found to be insufficient to enable the complete removal of recipient immunocompetent cells, since donor-reactive T cells were observed in the recipient spleens and many recipient-type NK and CD4(+) cells were also detected in the recipient hematolymphoid tissues. In all the combinations, the highest survival rate was achieved in the recipients irradiated with (6+6) or (6.5+6.5) Gy with a 4 h interval. In the surviving mice, the hematolymphoid tissues had been fully reconstituted with donor cells.

Animals↗

PubMed2001

Intra-bone marrow injection of allogeneic bone marrow cells: a powerful new strategy for treatment of intractable autoimmune diseases in MRL/lpr mice.

Intractable autoimmune diseases in chimeric resistant MRL/lpr mice were treated by a new bone marrow transplantation (BMT) method consisting of fractionated irradiation, 5.5 Gy x 2, followed by intra-bone marrow (IBM) injection of whole bone marrow cells (BMCs) from allogeneic normal C57BL/6 (B6) mice (5.5 Gy x 2 + IBM). In MRL/lpr mice treated with this method, the number of donor-derived cells in the bone marrow, spleen, and liver rapidly increased (almost 100% donor-derived cells by 14 days after the treatment), and the number of donor-derived hemopoietic progenitor cells concomitantly increased. Furthermore, donor-derived stromal cells were clearly detected in the cultured bone pieces from MRL/lpr mice treated with 5.5 Gy x 2 + IBM. All the recipients thus treated survived more than 1 year (> 60 weeks after birth) and remained free from autoimmune diseases. Autoantibodies decreased to almost normal levels, and abnormal T cells (Thy1.2(+)/B220(+)/CD4(-)/CD8(-)) disappeared. Hematolymphoid cells were reconstituted with donor-derived cells, and newly developed T cells were tolerant to both donor (B6)-type and host (MRL/lpr)-type major histocompatibility complex determinants. Successful cooperation was achieved among T cells, B cells, and antigen-presenting cells when evaluated by in vitro antisheep red blood cell responses. These findings clearly indicate that this new strategy (IBM-BMT) creates the appropriate hemopoietic environment for the early recovery of hemopoiesis and donor cell engraftment, resulting in the complete amelioration of intractable autoimmune diseases in chimeric resistant MRL/lpr mice without recourse to immunosuppressants. This strategy would therefore be suitable for human therapy.

Animals↗

PubMed2001

Major histocompatibility complex restriction between hematopoietic stem cells and stromal cells in vitro.

We have previously found that a significant number of hematopoietic progenitors accumulate in engrafted bones with the same major histocompatibility complex (MHC) as the transplanted bone marrow cells. In the present study, to further clarify the MHC restriction between hematopoietic stem cells (HSC) and microenvironment, we carried out cobblestone colony formation assays by culturing HSCs with MHC-matched or -mismatched stromal cell monolayers. The formation of cobblestone colonies under MHC-mismatched stromal cells significantly decreased in comparison with MHC-matched stromal cells. However, the decrease in cobblestone colony formation under MHC-mismatched stromal cells was not significant when using MHC class I-deficient HSC or stromal cells. Taken together with the results using B10 congenic strains, it is suggested that the MHC preference is restricted by MHC class Ia molecules. Treatment with monoclonal antibodies (mAbs) against MHC class Ia molecules of stromal cell phenotypes significantly enhanced the cobblestone colony formation, whereas treatment with mAbs against HSC phenotypes significantly inhibited it. The expression of cytokines to promote hematopoiesis was enhanced by the mAbs against stromal cell phenotypes. The enhancement of cytokine expression was also observed when stromal cells and HSCs were MHC-matched. These results suggest that signaling via the MHC molecules augments stromal cell activity and elicits the MHC restriction.

Animals↗

PubMed2001

Successful allogeneic bone marrow transplantation (BMT) by injection of bone marrow cells via portal vein: stromal cells as BMT-facilitating cells.

We examined the importance of the coadministration of bone marrow (BM) stromal cells with BM cells via the portal vein. A significant increase in the number of day-14 colony-forming unit-spleen (CFU-S) was observed in the recipient mice injected with hemopoietic stem cells (HSCs) along with donor BM stromal cells obtained after three to four weeks of culture. Histological examination revealed that hematopoietic colonies composed of both donor hemopoietic cells and stromal cells coexist in the liver of these mice. However, when donor HSCs plus BM stromal cells were administered i.v., neither the stimulatory effects on CFU-S formation nor the hemopoietic colonies in the recipient liver were observed. These findings suggest that the interaction of HSCs with stromal cells in the liver is the first crucial step for successful engraftment of allogeneic HSCs. It is likely that donor stromal cells and HSCs trapped in the liver migrate into the recipient BM and spleen, where they form CFU-BM and CFU-S, respectively.

Animals↗

PubMed2001

Crucial role of donor-derived stromal cells in successful treatment for intractable autoimmune diseases in mrl/lpr mice by bmt via portal vein.

We have recently established a new bone marrow transplantation (BMT) method for the treatment of intractable autoimmune diseases in MRL/lpr mice; the method consists of fractionated irradiation (5.5 Gy x 2), followed by BMT of whole bone marrow cells (BMCs) from allogeneic C57BL/6 mice via the portal vein (abbreviated as 5.5 Gy x 2 + PV). In the present study, we investigate the mechanisms underlying the early engraftment of donor-derived cells in MRL/lpr mice by this method. In the mice treated with this method, the number of donor-derived cells possessing the mature lineage (Lin) markers rapidly increased in the BM, spleen, and liver; almost 100% were donor-derived cells by 14 days after the treatment. The number of donor-derived hemopoietic progenitor cells (defined as c-kit(+)/Lin(-) cells) increased in the BMCs, hepatic mononuclear cells, and especially spleen cells by 14 days after the treatment. Simultaneously, hemopoietic foci adjoining donor-derived stromal cells were observed in the liver when injected via the PV, but not via the peripheral vein (i.v.). When adherent cell-depleted BMCs were injected via the PV, recipients showed a marked reduction in the survival rate. However, when mice were transplanted with adherent cell-depleted BMCs with cultured stromal cells, all the recipients survived. These findings suggest that not only donor hematopoietic stem cells (HSCs) but also donor stromal cells administered via the PV were trapped in the liver, resulting in the early engraftment of donor HSCs in cooperation with donor-derived stromal cells. This new strategy to facilitate the early recovery of hemopoiesis would therefore be of great advantage in human application.

Animals↗

PubMed2000

Differentiation from thymic B cell progenitors to mature B cells in vitro.

The role of the thymic microenvironment in the development of murine thymic B cells has yet to be fully clarified. We therefore investigate the microenvironment that supports the development of mature thymic B cells (sIg+/B220+/CD43-B cells) from thymic B cell progenitors with immunophenotypes of sIg-/B220med/CD43+ cells. As we have previously reported, thymic B cells generated from these progenitors in the thymus are CD5+ B cells. We next study the in vitro condition that supports the differentiation of thymic B cell progenitors. Stromal cells (from the bone marrow or thymus), thymus-derived cell lines with the character of thymic nurse cells (TNCs) or thymic epithelial cells (TECs), or the bone marrow-derived cell line (MS-5) are tested for their ability to support B-lymphopoiesis from thymic B cell progenitors. Interestingly, thymic stromal cells (but neither stromal cells from the bone marrow nor stromal cell lines) support the differentiation of thymic B cell progenitors into thymic B cells in the presence of IL-7. Cortical epithelia (but not medullary epithelia, thymic macrophages or dendritic cells) are found to contribute to thymic B cell differentiation. Surface phenotype and Ig rearrangement analyses reveal that mature B cells generated in this condition are primarily CD5+ B cells, indicating that the thymic microenvironment (particularly cortical epithelia) determines the differentiation of thymic B cells.

Animals↗

PubMed2000

Evidence for migration of donor bone marrow stromal cells into recipient thymus after bone marrow transplantation plus bone grafts: A role of stromal cells in positive selection.

Intrathymic T-cell differentiation is characterized by two selection events: positive and negative selection. It has been shown that thymic epithelial cells in the cortex are involved in the positive selection, while macrophages and dendritic cells, derived from hemopoietic stem cells, are involved in the negative selection. Here we investigate whether donor-derived bone marrow stromal cells can migrate into the thymus and participate there in positive selection after bone marrow transplantation plus bone grafts (to recruit bone marrow stromal cells). Allogeneic bone marrow transplantation with or without bone grafts was carried out in the [C57BL/6-->C3H] combination. Fluorescence-activated cell sorter analyses of recipient thymic adherent cells showed that donor-type bone marrow stromal cells exist in the thymus of mice that received bone marrow plus bone grafts but not in the mice that received bone marrow cells alone. Histological examination using confocal microscopy also confirmed the existence of donor-type stromal cells in the thymus of mice that received bone marrow cells plus bones. Both T-cell proliferation and plaque-forming cell assays indicated that the T cells of such mice show donor-type major histocompatibility complex-restriction. These findings strongly suggest that stromal cells can migrate from the bone marrow to the thymus, where they participate in the positive selection of thymocytes.

Animals↗

PubMed2000

A new method for bone marrow cell harvesting.

To minimize contamination of bone marrow cells (BMCs) with T cells from the peripheral blood, a new "perfusion method" for collecting BMCs is proposed using cynomolgus monkeys. Two BM puncture needles are inserted into a long bone such as the humerus, femur, or tibia. One needle is connected to an extension tube and the end of the tube is inserted into a culture flask to collect the BM fluid. The other needle is connected to a syringe containing 30 ml of phosphate-buffered saline. The solution is pushed gently from the syringe into the medullary cavity, and the medium containing the BM fluid is collected into the culture flask. There is significantly less contamination with peripheral blood, determined from the frequencies of CD4(+) and CD8(+) T cells, when using this method (<6%) than when using the conventional method (>20%) consisting of multiple BM aspirations from the iliac crest. Furthermore, the number and progenitor activities of the cells harvested using this "perfusion method" are greater than those harvested using the conventional aspiration method. This perfusion method was carried out 42 times using 15 cynomolgus monkeys, and no complications such as pulmonary infarction or paralysis were observed. These findings suggest that the "perfusion method" is safe and simple and would be of great advantage in obtaining pure BMCs, resulting in a less frequent occurrence of acute graft-versus-host-disease in allogeneic BM transplantation.

Animals↗

PubMed1999

c-kit<low Pluripotent hemopoietic stem cells form CFU-S on day 16.

Using Ly5 congenic mice, we characterized the early differentiation step of pluripotent hemopoietic stem cells. Lineage- (Lin-)/CD71- cells in the bone marrow cells were separated into major histocompatibility complex (MHC) class I(high)/c-kit(low) and MHC class I(high)/c-kit<low populations from C57BL/6 Ly5.1 male mice. These two populations (1,000 cells) were transplanted into lethally irradiated (5.5 Gy x 2) C57BL/6 Ly5.2 female mice. Colony-forming unit in spleen (CFU-S) assays were carried out on days 10, 12, 14, 16, and 20. In the mice that received c-kit(low) cells, CFU-S were first detected on day 12, and the CFU-S counts gradually increased. In contrast, no visible colony was detected until day 14 in the mice that received c-kit<low cells; CFU-S were first observed on day 16. Donor-derived (Ly5.1+) cells, such as B cells, T cells, and myeloid cells, were detected by fluorescence-activated cell sorter analyses, and donor-derived erythroid cells were detected by polymerase chain reaction analyses using Y-chromosome-specific primers. Donor-derived cells in the recipients of c-kit(low) cells were detected in the spleen, bone marrow, and peripheral blood on day 12 after transplantation, while they were detected on day 16 in the mice that received c-kit<low cells. Therefore, c-kit<low cells have the capacity not only to form CFU-S on day 16 but also to reconstitute the recipients with donor-derived hematolymphoid cells 16 days after transplantation.

Animals↗

PubMed1999

HGF activates signal transduction from EPO receptor on human cord blood CD34+/CD45+ cells.

Hepatocyte growth factor (HGF) is a multifunctional cytokine with early hematopoiesis-stimulatory activity. Here, we focus on its erythropoiesis-stimulatory effect on highly purified human hematopoietic progenitor cells (CD34+/CD45+ cells) derived from the cord blood. In immunoblot analyses, c-met protein (a receptor of HGF) was detected in the CD34+/CD45+ cells, although the expression levels were different among samples. The c-met expression was facilitated by incubation of the cells with stem cell factor (SCF) or interleukin 3 (IL-3), even if the expression level had been low. IL-6, G-CSF, or erythropoietin (EPO) did not show such a stimulatory effect on the c-met expression of the cells. When HGF was added to the CD34+/CD45+ cells in the presence of SCF, the numbers of CD36+/CD11b- cells (very early erythroid lineage cells) and BFU-E increased. EPO-dependent tyrosine phosphorylation of Stat 5 also increased, but the EPO receptor (EPO-R) expression remained unchanged in the CD34+/CD45+ cells treated with SCF + HGF. Our present study suggests that stimulation of the HGF/c-met signal is concomitant with induction of c-met protein by SCF. The subsequent enhancement of signal transduction via the activation of Stat 5 from the EPO-R plays a crucial role in the commitment of hematopoietic stem cells into erythroid lineage cells.

Antigens, CD34↗

PubMed1999

Age-dependent abnormalities of hematopoietic stem cells in (NZW x BXSB)F1 mice.

The (NZW x BXSB)F1 (W/BF1) mouse is known as an autoimmune-prone strain which develops lupus nephritis, thrombocytopenia due to platelet-specific autoantibodies, leukocytosis, and myocardial infarction. In this experiment, we investigated the age-dependent abnormalities of the hematopoietic stem cells (HSCs) and hematopoiesis in this mouse. White blood cell counts (especially Mac-1- or Gr-1-positive cells) in the peripheral blood of 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or normal mice. To investigate whether the abnormal hematopoiesis can be attributed to the HSCs of W/BF1 mice, colony-forming unit in spleen (CFU-S) and colony-forming unit in culture (CFU-C) assays were performed. Day 12 CFU-S counts of 12-week-old W/BF1 mice significantly increased in comparison with those of four-week-old W/BF1 mice or normal mice. In the CFU-C assay, CFU-GEMM and CFU-GM counts in 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or control mice. The bone marrow cells (BMCs) from 12-week-old W/BF1 mice showed a high level of G-CSF and a low level of GM-CSF in mRNA expression. To examine the effect of HSCs from 12-week-old W/BF1 mice on the onset of autoimmune diseases and the abnormal hematopoiesis, T- and B-cell-depleted BMCs of four-week-old or 12-week-old W/BF1 mice were transplanted to C3H mice. Recipient C3H mice that had received the BMCs from 12-week-old W/BF1 mice showed an earlier onset of autoimmune diseases and a shorter survival rate than those that had received the BMCs from four-week-old W/BF1 mice. These data suggest that the HSCs from 12-week-old W/BF1 mice showing the symptoms of autoimmune diseases have the capacity to induce autoimmune diseases earlier than the HSCs from four-week-old W/BF1 mice.

Age of Onset↗

PubMed1999

Experimental autoimmune thyroiditis induced by thyroglobulin-pulsed dendritic cells.

Dendritic cells (DCs), which are the most effective professional antigen-presenting cells (APCs), initiate and regulate immune responses. In this report, we examine the role of DCs in the induction of autoimmune thyroiditis. Experimental autoimmune thyroiditis (EAT) induced by immunization with thyroglobulin (Tg) plus adjuvant is considered to be an animal model of autoimmune thyroiditis, and is categorized as a T cell-mediated autoimmune disease. To examine the contribution of DCs to EAT, naive DCs were purified from high responder B10BR mice and pulsed with various concentrations of porcine Tg (pTg). These pTg-pulsed DCs were transferred without adjuvant to syngenic BIOBR mice to induce EAT. Mice that had received pTg-pulsed DCs showed thyroiditis, and the degree of thyroiditis induced was positively correlated to the amounts of pTg used for the incubation (pulsing) of DCs. The severity of thyroiditis was also correlated to the amounts of anti-pTg IgG2a antibodies and IFN-gamma in the recipient sera, but not to IL-4 or IL-10, indicating that Th1 cells are mainly activated by pTg-pulsed DCs and attributable to the pathogenesis of EAT.

Animals↗

PubMed1998

Stimulatory effects of hepatocyte growth factor on hemopoiesis of SCF/c-kit system-deficient mice.

In this study, we report that W/W mutant mice, which have severe macrocytic anemia caused by a deficit of extracellular domain in c-kit molecules and therefore die perinatally, have hemopoietic stem cells (HSCs) and mature hematolymphoid cells in the bone marrow (BM), thymus, and spleen, although there are significant decreases in cell counts. Moreover, the mitogen-induced proliferative response, mixed lymphocyte reaction, and anti-SRBC plaque formation of spleen cells in W/W mice are similar to those in age-matched +/? littermates and normal mice, suggesting that the SCF/c-kit system is necessary for cell proliferation but not essential for HSCs to differentiate. We next examine the stimulatory effects of hepatocyte growth factor (HGF) on hemopoiesis in W/W mice. HGF has a stimulatory effect on the colony formation (CFU-C) of W/W BM cells when cultured using either a methylcellulose assay (containing cytokines) or a long-term culture (LTC) assay. A similar stimulatory effect of HGF is observed in the other W or SI locus-mutant mice (W/Wv and SI/SId mice), which show less severe anemia than W/W. The numbers of nonadherent cells and cobblestone colonies significantly increase in the LTCs using their BM cells. In addition, in vivo administration of HGF shows a transient increase in the CFU-C counts in BM cells and peripheral blood cells. RBC, WBC, and platelet counts also increased. These results suggest that the SCF/c-kit system is not essential to hemopoiesis but that a compensatory system such as the HGF/c-met system functions in the SCF/c-kit system-deficient mice.

Anemia, Macrocytic↗

PubMed1997

Isolation and identification of hematopoietic stem cell-stimulating substances from Kampo (Japanese herbal) medicine, Juzen-taiho-to.

We have previously found that TJ-48 has the capacity to accelerate recovery from hematopoietic injury induced by radiation and the anti-cancer drug mitomycin C (MMC). The effects are found to be due to its stimulation of spleen colony-forming unit (CFU-S) counts on day 14. In the present study, we attempt to isolate and purify the active components in TJ-48 extracts using a new in vitro hematopoietic stem cell (HSC) assay method. n-Hexane extract from TJ-48 shows a significant stimulatory activity. The extract is further fractionated by silica gel chromatography and HPLC in order to identify its active components. 1H-NMR and GC-EI-MS indicate that the active fraction is composed of free fatty acids (oleic acid and linolenic acid). When 27 kinds of free fatty acids (commercially available) are tested using the HSC proliferating assay, oleic acid, elaidic acid, and linolenic acid are found to have potent activity. The administration of oleic acid to MMC-treated mice enhances CFU-S counts on days 8 and 14 to twice the control group. These findings strongly suggest that fatty acids contained in TJ-48 actively promote the proliferation of HSCs. Although many mechanisms seem to be involved in the stimulation of HSC proliferation, we speculate that at least one of the signals is mediated by stromal cells, rather than any direct interaction with the HSCs.

Animals↗

PubMed1997

Pluripotent hemopoietic stem cells are c-kit<low.

Pluripotent hemopoietic stem cells (P-HSCs) were thought to be c-kit+, but recent reports indicate that they are c-kit(low). In the present report, we provide evidence using Ly5 congenic mice that P-HSCs are c-kit(<low). Lineage-negative (Lin-)/CD71- cells among bone marrow cells (BMCs) from C57BL/6 Ly5.1 mice were separated into major histocompatibility complex class I(high) (class I(high))/c-kit(low) and class I(high)/ c-kit(<low) populations. Each population (500 cells) was transplanted into lethally (9.0 Gy) irradiated C57BL/6 Ly5.2 congenic mice along with Ly5.2 (2 x 10(5)) compromised cells. Donor-derived Ly5.1+ cells were detected 6 months after transplantation in primary recipients reconstituted with either class I(high)/c-kit(low) or class I(high)/c-kit(<low) cells. BMCs (1 x 10(6)) from the primary recipients were further transplanted into secondary recipients (Ly5.2 mice) to assess their long term repopulating activity. Six months after bone marrow transplantation, Ly5.1+ cells in all lineages were detected only in secondary recipients that had been given BMCs from the primary recipients reconstituted with class I(high)/c-kit(<low) cells but not in cells that were class I(high)/c-kit(low). When the BMCs (1 x 10(6)) of these secondary recipients were further transplanted into tertiary recipients, all tertiary recipients that had been given BMCs from the secondary recipients originally reconstituted with Lin-/CD71-/class I(high)/c-kit(low) cells died within 10 days whereas all six tertiary recipients originally reconstituted with Lin-/CD71-/class I(high)/c-kit(<low) cells showed donor (Ly5.1+)-derived cells in their peripheral blood. In the single tertiary recipient that was killed, donor-derived T cells, B cells, macrophages, and granulocytes also were detected in several major hematolymphoid organs. The remaining five mice continue to survive more than 6 months after the tertiary bone marrow transplantation.

Animals↗

PubMed1997

Distinct qualitative differences between normal and abnormal hemopoietic stem cells in vivo and in vitro.

The transplantation of partially purified hemopoietic stem cells (HSCs) plus the engraftment of bone from autoimmune-prone mice ((NZW x BXSB)F1 (W/BF1) mice) induces autoimmune diseases in major histocompatibility complex (MHC)-incompatible normal C3H/HeN mice. In contrast, W/BF1 mice die of infection or anemia within three weeks due to a failure in hemopoietic reconstitution when the mice receive partially purified HSCs plus bones from normal C3H/HeN mice, although they survive more than a year without showing any symptoms of autoimmune diseases when they receive T cell-depleted bone marrow cells (without bone grafts) from normal mice. This finding suggests that abnormal HSCs can proliferate even in MHC-incompatible microenvironments, while normal HSCs cannot. This is confirmed by spleen colony-forming assays (CFU-S) on day 12, using pluripotent HSCs (P-HSCs). The P-HSCs of old (> 4 mo) W/BF1 mice (after the development of autoimmune diseases) form high CFU-S counts on day 12 even in the allogeneic C3H environment, although the P-HSCs of normal mice form high CFU-S counts only in the MHC-compatible environments. In addition, abnormal P-HSCs of autoimmune-prone mice can proliferate in vitro in collaboration with MHC-incompatible stromal cells, although normal HSCs do so in collaboration with MHC-compatible stromal cells, but not MHC-incompatible stromal cells. These findings indicate that abnormal P-HSCs are more "resilient" than normal P-HSCs.

Animals↗

PubMed1997

Induction of c-kit molecules on human CD34+/c-kit < low cells: evidence for CD34+/c-kit < low cells as primitive hematopoietic stem cells.

c-kit, a receptor for stem cell factor, has been widely accepted as a distinctive marker for hematopoietic stem cells. However, the level of c-kit expression on pluripotent hematopoietic stem cells is still controversial in mice and humans. We purified CD34+/c-kit < low cells (phenotypically c-kit-negative but only detectable at the message level) from human cord blood and examined their maturational steps in relation to the expression of c-kit molecules. When the CD34+/c-kit < low cells were cultured with cytokines (flt 3 ligand, interleukin 6 and interleukin 7) plus immobilized anti-CD34 monoclonal antibody (to crosslink CD34 molecules), c-kit molecules were clearly induced within 24 h. The c-kit expression gradually increased until day 8. When CD34+/c-kit(low) or CD34+/c-kit+ cells that had been induced from CD34+/c-kit < low cells were resorted and recultured using a methylcellulose culture system, they showed the same colony-forming ability as the freshly isolated CD34+/c-kit(low) or CD34+/c-kit+ cells, respectively. Furthermore, CD34+/c-kit < low cells have a similar hematopoietic potential to CD34+/c-kit(low) cells in assays for long-term culture initiating cell and colony-forming unit culture generated from long-term cultures. These findings suggest that CD34+/c-kit < low cells mature into CD34+/c-kit(low) and CD34+/c-kit+ cells, and acquire the reactivity to various humoral hematopoietic stimuli. Moreover, CD34+/c-kit < low cells showed a low level of rhodamine 123 retention, suggesting that CD34+/c-kit < low cells have multidrug resistance. Therefore, the CD34+/c-kit < low cells without colony-forming unit-granulocyte-erythroid-macrophage-megakaryocyte activity are also a pluripotent hematopoietic stem cell population, and the expression of c-kit on c-kit < low cells is the first maturational step of hematopoiesis.

Antigens, CD34↗

PubMed1997

Intrathymically injected hemopoietic stem cells can differentiate into all lineage cells in the thymus: differences between c-kit+ cells and c-kit < low cells.

To investigate whether hemopoietic stem cells (HSCs) can differentiate into all lineage cells even in the thymus, we injected two types of HSCs (c-kit+ and c-kit < low cells) obtained from C57BL/6 Ly5.1 mice directly into the thymus of 7.5 Gy-irradiated C57BL/6 Ly5.2 mice. When c-kit < low cells (low density/lineage-/CD71-/major histocompatibility complex class I high/Sca-1+/Thy-1low/ c-kit < low) were injected, donor-derived (Ly5.1) cells were detected on day 8 after intrathymic (i.t.) injection, and the number reached a maximum on day 24 after injection. Granulocytes and macrophages were also detected on day 8 after injection. However, B220+ B cells were observed on day 13. Eighteen days after i.t. injection, the injected lobes showed red color due to the synchronous development of erythroid cells. Histological studies revealed the development not only of erythroid lineage cells but also of megakaryocytes in the thymus. In contrast, when c-kit+ cells were injected, a significant number of donor-derived cells were detected on day 5 after i.t. injection (three days earlier than in the case of c-kit < low cell injection). The differentiation into erythroid lineage cells was also observed six days earlier than when c-kit < low HSCs were injected. These findings suggest that c-kit < low HSCs are more primitive than c-kit+ HSCs, although both can differentiate into all lineage cells after i.t. injection.

Animals↗