Hemopoietic precursor cell defects in nonanemic but stem cell-deficient W44/W44 mice

Hematopoietic stem cell deficiencies cause a severe macrocytic anemia in W/Wv mice. W44/W44 mice, on the other hand, are not anemic, but, since they accept marrow implants without prior total body irradiation, they have inherited a stem cell lesion. In an attempt to identify the aberrant stem cell(s), we have determined the concentration in W44/W44 marrow of hematopoietic precursors known to be deficient in W/Wv marrow. The in vitro erythroid burst-forming units (BFU-E), the in vivo spleen colony-forming units (CFU-S), and the cells that repopulate the erythroid compartment of stem cell-deficient mice were examined. The progenitors of 7-day bursts are dramatically reduced in W/Wv marrow but are present in normal concentrations in W44/W44 marrow. W44/W44 marrow CFU-S, unlike W/Wv, generate visible spleen colonies 10 days after injection into lethally irradiated recipients. The colonies are, however, smaller and at least 2 times less numerous than those produced from equivalent numbers of +/+ marrow. An additional defect was the inability of W44/W44 stem cells to compete with genetically marked +/+ cells during erythroid repopulation. An estimate of the number of W44/W44 stem cells needed to compete with +/+ cells was provided by enriching W44/W44 progenitors fivefold. Twice as many enriched W44/W44 marrow cells as unfractionated +/+ cells were required to replace competitor cells. This suggests that there are up to 10 times fewer stem cells somewhere in the W44/W44 erythrogenerative pathway. The data support the conclusion that an erythroid progenitor less mature than the BFU-E is one of the cells most severely affected by expression of the mutant gene.     

Fibroblast-dependent growth of mouse mast cells in vitro: duplication of mast cell depletion in mutant mice of W/Wv genotype

In spite of the apparent depletion of mast cells in tissues of mutant mice of W/Wv genotype, cells with many features of mast cells do develop when bone marrow cells of W/Wv mice are cultured in the presence of pokeweed mitogen-stimulated spleen cell-conditioned medium (PWM-SCM). In order to resolve this discrepancy and facilitate the analysis of the W mutation, we attempted to establish an in vitro system in which the in vivo defect of W/Wv mice can be reproduced. Cultured mast cells (CMC) were developed from bone marrow cells of either W/Wv or congenic +/+ mice, and then co-cultured with NIH/3T3 mouse fibroblasts in media supplemented only with fetal calf serum (i.e., in the absence of PWM-SCM). Under this condition, CMC from +/+ mice continued to divide and were maintained for more than 4 weeks. The supportive effect of NIH/3T3 cells required close-range interactions with CMC and was not due to synthesis of the known mast cell growth factors, interleukins 3 and 4. By contrast, CMC from W/Wv mice were not maintained, and the number of mast cells remaining after 4 weeks of co-culture was only 1% of the normal +/+ counterparts. Thus, the humoral factor-independent and cell contact-dependent system presented here revealed the intrinsic defects in growth and differentiation of CMC derived from W/Wv mice and might be useful for biochemical and molecular analysis of the gene product(s) encoded at the W locus.     

Proliferation and differentiation in culture of mast cell progenitors derived from mast cell-deficient mice of genotype W/Wv

Mice of genotype W/Wv have less than 1% of normal mast cells in the skin, stomach, and cecum. In order to further clarify the mechanism of this deficiency, we studied committed mast cell progenitors and multipotent progenitors, which are capable of mast cell differentiation in clonal culture. The relative concentration of mast cell progenitors in the bone marrow, spleen, and peripheral blood of W/Wv mice was similar to that of +/+ mice. However, the cellularity of the marrows of W/Wv mice was 54% of that of their normal littermates. Identification of mast cells was established by metachromatic staining with toluidine blue, transmission electron microscopy, and demonstration of membrane receptors for immunoglobulin E. The time course of colony formation and the morphology of W/Wv mast cell colonies in culture was identical to that of normal littermates. The percentages of mast cells in individual multi-lineage colonies were extremely variable. The histamine content of mast cells derived from W/Wv mice was similar to that of mast cells from +/+ mice. These studies demonstrated the normal capacity for differentiation and proliferation in culture of mast cell progenitors from W/Wv mice.     

Inability of genetically mast cell-deficient W/Wv mice to acquire resistance against larval Haemaphysalis longicornis ticks

Genetically mast cell-deficient (WB X C57BL/6)F1-W/Wv mice were used to investigate the role of mast cells for the acquisition of resistance against larval Haemaphysalis longicornis ticks. Resistance against ticks was evaluated by reduction in both number and weight of engorged ticks. Although (WB X C57BL/6)F1-+/+ mice with a normal number of mast cells acquired resistance after repeated infestation of ticks, the congenic W/Wv mice did not acquire it. Bone marrow transplantation from the +/+ mice were grafted onto the back of the W/Wv mice, resistance against the ticks was detectable in the grafted skin. In contrast, resistance was not detectable in the skin of the W/Wv mice which had been grafted onto the back of the syngenic W/Wv mice. Thus, we consider that the failure of the W/Wv mice to manifest resistance is attributable to the mast cell depletion.     

Maintenance of granulopoiesis in long-term bone marrow cultures from W/Wv mice and effects of lipopolysaccharide on granulopoiesis in culture

An attempt was made to establish long-term cultures of marrow cells from genetically anaemic W/Wv mice. Two batches of horse sera were used. One batch of horse serum (HS-lot A) supported long-term maintenance (up to 20 weeks) of granulopoiesis in vitro. The number of suspension cells in W/Wv marrow culture was maintained at the same level as that in the control +/+ culture, but the number of granulocyte-macrophage progenitor cells (GM-CFC) and the ratio of immature to mature granulocytes were at a lower level than those in +/+ culture. These data suggest that haemopoietic progenitors in W/Wv cultures maintain a higher level of differentiation, and hence an increased self-renewal than those in +/+ cultures. Another batch of horse serum (HS-lot B) was less effective in the maintenance of the cultures, and the cultures deteriorated within 10 weeks. Addition of bacterial lipopolysaccharide (LPS) induced increased granulopoiesis in +/+ cultures, whereas such treatment resulted in the depletion of suspension cells in W/Wv cultures. The results suggest that haemopoietic cells of W/Wv mouse cannot cope with the strong stimulus for differentiation that occurs after the administration of LPS, although the cells can continue a moderately increased self-renewal and differentiation, as indicated by the results in the culture with HS-lot A.     

Characterization of spleen colonies derived from mice with mutations at the W locus.

Mice with mutations at the W locus have a hemopoietic stem cell defect characterized by an apparent deficiency of spleen colony forming cells (CFU-S). In the present report, we provide evidence that mutant cells form colonies and we compare the characteristics of the colonies derived from mutant and normal cells. To perform the colony-derivation studies, marrow cells were transferred into lethally irradiated congenic hosts that differed from the donors in the ubiquitous genetic marker, glucose phosphate isomerase (GPI-1). Donor GPI-1 comprised over 50% of the marker in the host spleen and marrow by 12 days post injection, regardless of whether the donor was mutant or normal. To characterize the colonies, serially sectioned host spleens were examined microscopically. Colonies are present by 8 days post-transplantation regardless of donor genotype, but mutant colonies are distinctly different from normal colonies. The proportion of blast and granulocyte colonies is always greater in W/Wv than in +/+ recipients. Unlike the W/Wv donors, the +/+ donors generate primarily erythrocyte colonies at 8, 10, and 14 days and mixed colonies at 12 days post-injection. Colonies from the mutant mice are generally smaller but visible colonies do appear by 12 days. The results are consistent with the notion that the anemia in W/Wv mice is caused by the early restriction of differentiating cells to a non-erythrocyte lineage accompanied by the delayed amplification of mutant hemopoietic cells. Whether this means erythrocyte-committed cells are absent or are present but unable to respond to the appropriate cytokines is not possible to determine from the current experiments.     

Radiation and haematopoiesis in Harwell steel mice

Haematological information on steel (Sl) mice is limited largely to Sl/Sld mice of Bar Harbor stock (WC.B6 F1). Therefore, two Harwell alleles, SlgbH and Slcon, were investigated. In the steady state both heterozygotes were modestly anaemic, homozygous Slcon and compound Slcon/SlgbH more so. On perturbation by X-irradiation Slcon/SlgbH showed a decrease in median lethal dose (MLD)--6.5 Gy, Slcon/+ and Slcon/Slcon slightly less change (7.5 Gy) compared with +/+, 8 Gy. In recovery from sublethal doses single heterozygotes, double heterozygotes with Wv, and compounds showed no delay in restoration of the count of red blood corpuscles (RBC) such as that seen in typical W mice (e.g. Wv/+, W/Wv). Effects on Slcon/Slcon and Slcon/SlgbH differ from those reported for Sl/Sld in that they show normal growth of spleen colonies when used as lethally irradiated recipients of bone marrow, they support growth of implanted bone marrow to form radiation chimaeras. When Harwell steel mice are donors of bone marrow to lethally irradiated +/+ mice the chimaeras ultimately are not anaemic; when lethally irradiated Harwell steel mice are recipients of +/+ marrow they remain macrocytically anaemic. One deduces that, for normal development and production of normal RBC in the steady state, the erythron requires intrinsic factors determined by wild type alleles at the W locus and extrinsic factors determined by wild type alleles at the Sl locus. Mutant alleles at either locus may determine macrocytosis. Two mutant alleles at either locus are still more deleterious, often lethal. Whereas mutant W alleles may also influence the pluripotent haematopoietic stem cell (HSC) leading to reduced MLD on X-irradiation, a similarly reduced MLD for Sl mutants may represent an increased need for and consumption of products of the haematopoietic stem cells rather than truly increased radiosensitivity, since the Do for spleen colony-forming units is the same for Slcon/SlgbH as +/+ mice.     

Hybrid resistance to parental mast cell precursors in the skin of (WB X C57BL/6)F1-W/Wv mice

When bone marrow cells of (WB X C57BL/6)F1-+/+ (WBB6F1-+/+) and WB-+/+ (WB) mice were directly injected into the skin of genetically mast cell-deficient WBB6F1-W/Wv mice, mast cell clusters appeared at the injection sites. However, the number of WB bone marrow cells necessary for appearance of mast cell clusters was significantly larger than when bone marrow cells of WBB6F1-+/+ mice were used. When WB bone marrow cells were mixed either with WB thymus cells or with silica particles, the proportion of injection sites at which mast cell clusters appeared increased to the level that was observed after the injection of the same number of WBB6F1-+/+ bone marrow cells. When suckling WBB6F1-W/Wv mice of less than or equal to 18 days of age were used as recipients, bone marrow cells of WBB6F1-+/+ and WB mice produced mast cell clusters with a comparable efficiency. Both syngeneic thymus cells and silica particles are known to abrogate the hybrid resistance that is observed in the spleen against parental hematopoietic stem cells. The hybrid resistance in the spleen is not detectable in suckling mice, either. Thus, the poor growth of mast cell precursors in the skin and the poor growth of hematopoietic stem cells in the spleen seem to be regulated by the same mechanism.     

An assessment of mast-cell deficient mice (W/Wv) as a model system to study the role of histamine in implantation and deciduoma formation

The ovaries from mast cell-normal (+/+) and mast cell-deficient (W/Wv) mice were examined with light and electron microscopy. In addition the effect of ovariectomy and subsequent steroid treatment on total uterine histamine content, total mast cell numbers and surface and glandular epithelial cell heights was measured. The ovaries of +/+ mice were normal, displaying various stages of follicular growth and atresia and numerous corpora lutea; the ovaries of W/Wv mice lacked follicles and corpora lutea but contained numerous hyperplastic interstitial cells which contained numerous lipid droplets, vesiculated mitochondria and abundant endoplasmic reticulum suggestive of steroid synthesis. Steroid treatment of ovariectomized +/+ and W/Wv mice caused a significant increase in uterine wet weight and endometrial surface and glandular epithelial cell heights. In +/+ mice, steroid treatment caused a concomitant increase in total mast cells per uterine horn while mast cells were totally absent in W/Wv mice. The increase in uterine histamine in +/+ mice is consistent with the increase in mast cell numbers. Measurable amounts of uterine histamine, which increases slightly after steroid treatment, were demonstrated in W/Wv mice. Since the uteri of +/+ and W/Wv mice respond to steroids in a similar manner with the sole exception being histamine content and mast cell numbers, our results demonstrate the potential of using these animals to investigate the role(s) of uterine mast cells and non-mast cell uterine histamine in the process of implantation and the formation of a decidual cell response.     

Effect of alpha-fluoromethylhistidine on the histamine content of the brain of W/Wv mice devoid of mast cells: turnover of brain histamine

In the brains of W/Wv mutant mice that have no mast cells, the histidine decarboxylase (HDC) level is as high as in the brain of congenic normal mice (+/+), but the histamine content is 53% of that of +/+ mice. The effects of alpha-fluoromethylhistidine (alpha-FMH) on the HDC activity and histamine content of the brain of W/Wv and +/+ mice were examined. In both strains, 30 min after i.p. injection of alpha-FMH the HDC activity of the brain had decreased to 10% of that in untreated mice. The histamine content decreased more gradually, and after 6 h about half of the control level remained in +/+ mice, whereas histamine had disappeared almost completely in W/Wv mice. It is concluded that the portion of the histamine content that was depleted by HDC inhibitor in a short time is derived from non-mast cells, probably neural cells. The half-life of histamine in the brain of W/Wv mice was estimated from the time-dependent decrease in the histamine content of the brain after administration of alpha-FMH: 48 min in the forebrain, 103 min in the midbrain, and 66 min in the hindbrain.     

Regulatory mechanisms of mast cell differentiation

Mast cells are a progeny of the multipotential hematopoietic stem cell. Most of progenies of the stem cell complete their differentiation within the bone marrow, but precursors of mast cells leave the bone marrow, migrate in blood, and invade into tissues. After the invasion, precursors proliferate and differentiate into mast cells. An appreciable proportion of mast cells retain proliferative potential after differentiation, and even after degranulation, some mast cells can proliferate and recover the original morphology. Proliferation of mast cells are regulated by both T cell-derived factors (i.e., IL-3 and IL-4) and fibroblast-derived factor(s). Mice of either W/Wv or Sl/Sld genotype lack mast cells, but mast cells do develop when bone marrow cells of W/Wv or Sl/Sld mice were cultured in the presence of T cell-derived factors. Mast cells derived from W/Wv mice cannot respond fibroblast-derived factor(s) and fibroblasts derived from Sl/Sld mice cannot support mast cells of normal mouse origin. Phenotypes of mast cells are determined by the environment in which the mast cells differentiated. However, when mast cells are transplanted into a new environment which is different from the original one, the mast cells acquire the phenotype which are dependent on the second environment.     

Implantation, deciduoma formation and live births in mast cell-deficient mice (W/Wv)

The intraluminal injection of oil produced deciduoma formation in ovariectomized, mast cell-normal (+/+) and mast cell-deficient (W/Wv) mice that were treated with exogenous steroids. Oil injection and trauma (e.g. sutures) also produced a deciduoma in ovariectomized +/+ and W/Wv mice that had received a single control (+/+) ovary transplanted under the kidney capsule. After transfers of donor blastocysts, implantation and live births were obtained in +/+ and W/Wv mice containing a single ovary transplant. Our results demonstrate that uterine mast cells are not required for the production of a decidual cell response, implantation, gestation or the birth of live offspring in mice.     

Genetically mast cell-deficient W/Wv mice as a tool for studies of differentiation and function of mast cells

Genetically mast cell-deficient W/Wv mice are useful for the analysis of mast cell biology, especially as recipients of bone marrow cells and skin pieces. Inasmuch as suspension and clonal cultures of mast cells have been developed, we combined these in vivo and in vitro systems. Suspension-cultured mast cells had morphological and biochemical characteristics similar to those of mucosal mast cells (MMC). However, i.p. injection of such cultured mast cells gave rise to development of cells with characteristics similar to those of connective tissue mast cells (CTMC). When peritoneal cells of normal +/+ mice were cultured in methylcellulose, pure mast cell colonies appeared. Cells from individual mast cell colonies were divided and injected into the skin and stomach wall of W/Wv mice; CTMC developed in the skin and MMC in the stomach mucosa. This indicates the presence of a common precursor for CTMC and MMC. Morphology of such bipotent mast cell precursors was studied by using micromanipulation. About 4% of morphologically identifiable peritoneal mast cells may function as the bipotent precursors. Although W/Wv mice showed a defect in resistance against ixodid ticks, injection of suspension-cultured mast cells normalized the defect. The four examples mentioned above indicate that combinations of in vivo and in vitro systems increase the usefulness of W/Wv mice.     

Normalization of anti-tick response of mast cell-deficient W/Wv mice by intracutaneous injection of cultured mast cells

Genetically mast cell-deficient (WB X C57BL/6)F1-W/Wv mice show a defect in manifestation of resistance against larval Haemaphysalis longicornis ticks. In order to obtain direct evidence for anti-tick roles of mast cells, we examined whether intracutaneous injection of cultured mast cells normalized the defect of W/Wv mice. Bone marrow cells of (WB X C57BL/6)F1-+/+ mice were cultured in the presence of pokeweed mitogen-stimulated spleen cell-conditioned medium. More than 95% of the cultured cells were identified as immature mast cells 4 wk after the initiation of the culture; the cells were harvested and directly injected into the skin of W/Wv mice. Mast cells appeared at the injection sites, where the resistance against the ticks was observed. Thus, mast cells developing at the injection sites seem to play an essential role for manifestation of resistance.     

The importance of mast cells for the neutrophil influx in immune complex-induced peritonitis in mice

The role of mast cells in polymorphonuclear leukocyte (PMN) influx in Ag-antibody complex-induced peritonitis was evaluated in mast cell-deficient WBB6F1-W/Wv (W/Wv) mice and their normal littermates, WBB6F1-+/+ (+/+). Peritoneal cell influx was evaluated after i.p. injection of preformed immune complexes. The first significant elevation in the PMN count over PBS-treated controls in +/+ mice was observed 2 h after stimulation. During the period of maximum leukocyte concentrations (6 to 10 h), the increase in total cell count was 5-fold and in PMN 25-fold. In W/Wv mice the PMN influx started 2 h later than in the +/+ mice, and the maximum response (8 to 10 h) was only 50% of that in controls. Reconstitution of mast cells in W/Wv mice for 2 wk or more restored the PMN response to immune complexes. Mast cell release due to AG-antibody complexes was evaluated by measuring fluorescence intensity after berberine sulfate staining for heparin in mast cells from unstimulated as well as stimulated +/+ mice. There was a significant decrease in fluorescence intensity as early as 15 min after stimulation. By 30 min the fluorescence intensity had declined by 65%. This indicates extensive mast cell release that started before PMN mobilization. These experiments demonstrate that mast cells make a significant contribution to immune complex-induced inflammation.     

Proliferation and differentiation of spermatogonial stem cells in the w/wv mutant mouse testis

Mutations in the dominant-white spotting (W; c-kit) and stem cell factor (Sl; SCF) genes, which encode the transmembrane tyrosine kinase receptor and its ligand, respectively, affect both the proliferation and differentiation of many types of stem cells. Almost all homozygous W or Sl mutant mice are sterile because of the lack of differentiated germ cells or spermatogonial stem cells. To characterize spermatogenesis in c-kit/SCF mutants and to understand the role of c-kit signal transduction in spermatogonial stem cells, the existence, proliferation, and differentiation of spermatogonia were examined in the W/Wv mutant mouse testis. In the present study, some of the W/Wv mutant testes completely lacked spermatogonia, and many of the remaining testes contained only a few spermatogonia. Examination of the proliferative activity of the W/Wv mutant spermatogonia by transplantation of enhanced green fluorescent protein (eGFP)-labeled W/Wv spermatogonia into the seminiferous tubules of normal SCF (W/Wv) or SCF mutant (Sl/Sld) mice demonstrated that the W/Wv spermatogonia had the ability to settle and proliferate, but not to differentiate, in the recipient seminiferous tubules. Although the germ cells in the adult W/Wv testis were c-kit-receptor protein-negative undifferentiated type A spermatogonia, the juvenile germ cells were able to differentiate into spermatogonia that expressed the c-kit-receptor protein. Furthermore, differentiated germ cells with the c-kit-receptor protein on the cell surface could be induced by GnRH antagonist treatment, even in the adult W/Wv testis. These results indicate that all the spermatogonial stem cell characteristics of settlement, proliferation, and differentiation can be demonstrated without stimulating the c-kit-receptor signal. The c-kit/SCF signal transduction system appears to be necessary for the maintenance and proliferation of differentiated c-kit receptor-positive spermatogonia but not for the initial step of spermatogonial cell differentiation.     

Different radiosensitivities of mast-cell precursors in the bone marrow and skin of mice

Although tissue mast cells are derived from the bone marrow, some descendants of bone marrow-derived precursors retain the ability to proliferate and differentiate into mast cells even after localization in the skin. The purpose of the present study was to determine the D0 values for mast-cell precursors in the bone marrow and those localized in the skin. Bone marrow cells were removed from (WB X C57BL/6)F1-+/+ mice after various doses of irradiation and injected into the skin of the congenic W/Wv mice which were genetically without mast cells. Radiosensitivity of mast-cell precursors in the bone marrow was evaluated by determining the proportion of the injection sites at which mast cells did not appear. For the assay of the radiosensitivity of mast-cell precursors localized in the skin, pieces of skin were removed from beige C57BL/6 (bgJ/bgJ. Chediak-Higashi syndrome) mice after various doses of irradiation and grafted onto the back of the normal C57BL/6 mice. Radiosensitivity of mast-cell precursors in the skin was evaluated by determining the decrease of beige-type mast cells which possessed giant granules. Mast-cell precursors in the bone marrow were much more radiosensitive than those localized in the skin. D0 value was about 100 rad for the former and about 800 rad for the latter.     

Mast cells are important in the development of hypersensitivity pneumonitis. A study with mast-cell-deficient mice

We have previously reported that C57B1/6 mice develop lung lesions similar to human hypersensitivity pneumonitis (HP) by repeated transnasal administration of Thermoactinomyces vulgaris antigen. Since the HP-like lesions were induced via respiratory route and by the causative antigen in human HP (farmer's lung), it seems that this murine model is useful for investigating the cell-to-cell interactions in human HP. To clarify the involvement of mast cells (MC) in the development of HP, T. vulgaris (90 micrograms/day) was transnasally administered to MC-deficient WBB6F1-W/Wv mice (W/Wv) and their littermates (+/+) five times a wk for 3 wk. When the lungs were examined by scoring pathological findings and lung indexes, HP-like lesions were significantly less severe in W/Wv than in +/+, whose lesions were equivalent to those of C57B1/6. Bone-marrow-derived cultured MC from +/+ mice (98% purity) were obtained by in vitro culture mixed with WEHI-3B-derived conditioned medium which contained IL-3. When these MC were adoptively transferred to W/Wv mice (10(7) cells/mouse), the HP-like lesions in W/Wv mice were enhanced to be as severe as those in +/+. Importantly, significant numbers of MC were found in the lungs of MC-transferred W/Wv mice. These results suggest that MC play an important role in the development of the murine experimental HP.     

Formation of mast cell colonies in methylcellulose by mouse peritoneal cells and differentiation of these cloned cells in both the skin and the gastric mucosa of W/Wv mice: evidence that a common precursor can give rise to both "connective tissue-type" and "mucosal" mast cells

We investigated the issue of mast cell heterogeneity by cloning mast cell colonies from peritoneal cells in methylcellulose, injecting the cloned cells into the skin and stomach of mast cell-deficient (WB X C57BL/6)F1-W/Wv (WBB6F1-W/Wv) mice, and staining the mast cells that developed in these sites with Berberine sulfate, a fluorescent dye that identifies heparin-containing mast cells. When peritoneal cells of nontreated WBB6F1-+/+ mice were plated in methylcellulose containing pokeweed mitogen-stimulated spleen cell conditioned medium, pure mast cell colonies developed. In contrast, the peritoneal cavity of genetically mast cell-deficient WBB6F1-W/Wv mice lacked the progenitor cells that made mast-cell colonies. The clonal nature of the mast cell colonies was determined by using the giant granules of C57BL/6-bgJ/bgJ mice as a marker: even when mixture of peritoneal cells of C57BL/6-bgJ/bgJ mice and C57BL/6-+/+ mice were plated, all of the resulting colonies consisted of either bgJ/bgJ-type mast cells alone or +/+-type mast cells alone. Individual mast c 11 colonies of WBB6F1-+/+ mouse origin were divided into two parts; one part was directly injected into the wall of the glandular stomach of a WBB6F1-W/Wv mouse, and another part was injected into the skin of the same W/Wv mouse. Injections of 14 of 46 such colonies resulted in development of mast cells in both the "connective tissues" (skin or stomach muscle or both) and the stomach mucosa. Mast cells in the connective tissues were stained with Berberine-sulfate, indicating that they contained heparin, whereas mast cells in the stomach mucosa were not. These results suggest that a single precursor cell can give rise to both "connective tissue-type" and "mucosal" mast cells.     

White-spotting mutations affect the regenerative differentiation of testicular germ cells: demonstration by experimental cryptorchidism and its surgical reversal.

The effect of white-spotting (W) mutations on differentiation of testicular germ cells was investigated by using experimental cryptorchidism and its surgical reversal. All mutant mice used in this study (Wv/+, Wsh/+, Wf/+ and Wf/Wf) showed normal fertility and well-ordered spermatogenesis, as in congenic +/+ mice. In the cryptorchid testis, which contains only type A spermatogonia as germ cells, the number and the proliferative activity of type A spermatogonia in mutant mice were comparable to +/+ mice. On the other hand, surgical reversal of the cryptorchid testis in mutants resulted in impaired regenerative differentiation of germ cells. Although complete recovery of spermatogenesis was observed in +/+ mice, testicular weight in Wsh/+, Wf/+ and Wf/Wf mice recovered to approximately 60-70% of intact levels, and some portions of seminiferous epithelium showed incomplete spermatogenesis. In Wv/+ mice, however, ability to recover the weight was completely lost, and only type A spermatogonia existed as germ cells in seminiferous tubules 3 mo after surgical reversal. These results suggest that W mutation affects the differentiation through type A spermatogonia to type B spermatogonia, indicating the functional significance of W (c-kit) in early spermatogenesis.