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J B Turpen

Publications and source records attributed to J B Turpen.

At least 19 recordsLinked to original sources
PubMed2001

Cloning and developmental expression of Xenopus Stat1.

Proteins of the STAT family (Signal Transducers and Activators of Transcription) are latent cytoplasmic factors which, upon phosphorylation, are translocated to the nucleus where they participate in gene activation. In this report, we describe the cloning and developmental expression of a Xenopus homolog of Stat1. XStat1 is highly conserved, exhibiting greater than 90% identity in the critical DNA binding, SH2, SH3 and trans-activation domains with both human and murine Stat1. Using RT-PCR, we show that XStat1 is present as a maternal message during early development of Xenopus. The maternal message is translated during cleavage and its product is phosphorylated on tyrosine, a prerequisite for functional activation. During cleavage and gastrula stages, XStat1 is widely expressed throughout the developing embryo. During neurulation and early tailbud stages, XStat1 is expressed in both dorsal axial and ventral tissues. By late tailbud, dorsal XStat1 expression domains are associated with the developing pharyngeal arches and pronephros. These regions of the embryo correspond to the future location of the thymus, sites of dorsal hematopoietic activity, and one location where melanocytes differentiate.

Amino Acid Sequence↗
PubMed1997

Bipotential primitive-definitive hematopoietic progenitors in the vertebrate embryo.

Two regions of the vertebrate embryo, the blood islands and the dorsal lateral plate (DLP), participate in early hematopoietic development. In Xenopus, primitive erythrocytes are derived solely from the ventral blood islands (VBI), while definitive hematopoietic cells such as lymphocytes are derived from both VBI and DLP. We have utilized a transplantation technique to demonstrate in vivo that all hematopoietic cells (embryonic, fetal, or adult) originate from ventral mesoderm. Reciprocal grafts between VBI and DLP demonstrated that both regions are bipotential with respect to primitive and definitive hematopoiesis. Commitment of the VBI to primitive erythropoiesis and restriction of the DLP to definitive hematopoiesis occurs during neurula stages. Thus, hematopoietic development involves the induction of the blood program on the ventral axis of the embryo followed by environmentally regulated specification to the primitive or definitive lineages.

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PubMed1995

Intraembryonic origin of hepatic hematopoiesis in Xenopus laevis.

The liver is a major site of hematopoietic stem cell differentiation during vertebrate development. Hepatic hematopoiesis is dependent on colonization of the organ by extrinsically derived stem cells which, in mammals, are thought to originate only in the yolk sac. However, in birds and amphibians two distinct embryonic stem cell sources have been identified. The yolk sac or extraembryonic compartment is associated with the developing vitelline veins, and the para-aortic or intraembryonic compartment is associated with the dorsal aortae and postcardinal veins. The homologous compartments in the Xenopus embryo are the ventral blood island (extraembryonic) and dorsal lateral plate (intraembryonic) mesoderms, which contribute to primitive larval erythrocyte and definitive late larval and adult erythroid populations, respectively. The role of these embryonic stem cell sources in hepatic hematopoiesis has not been determined. We have examined the development of hepatic hematopoiesis in Xenopus 2N/3N stem cell chimeras using two-color FACS analysis. DNA content was determined using Hoechst 33342, and subpopulations of hematopoietic cells were identified with specific mAbs. Here we show that hepatic erythrocytes, leukocytes, and B lymphocytes in the liver of Xenopus larvae were derived from stem cells that originated from the intraembryonic mesoderm.

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PubMed1992

Differential stem cell contributions to thymocyte succession during development of Xenopus laevis.

The contribution of two embryonic stem cell compartments to the developing thymus in the amphibian Xenopus was examined throughout the larval, postmetamorphic, and adult periods. Hematopoietic chimeras were produced by transplanting either the ventral blood islands (VBI) or the dorsal stem cell compartment (DSC) from diploid donors onto triploid hosts. The DNA content of isolated nuclei harvested from the thymus and circulating E populations was analyzed using propidium iodide staining and flow cytometry. The DNA content of mitotic figures derived from PHA reactive splenocytes was analyzed using the Feulgen reaction and microdensitometry. These data suggested that both the VBI and DSC contribute to the thymocyte populations from the earliest developmental stages examined. Moreover, the contribution of both stem cell compartments was cyclic. However, the periods of these cycles were different. Both VBI- and DSC-derived cells entered the thymus 4 days postfertilization. VBI-derived thymocytes were at a minimum at 28 days postfertilization, reached a maximum at 35 days postfertilization and a second minimum at 42 days postfertilization. However, DSC-derived cells reached a maximum at 28 days, a minimum at 35 days, and a second maximum at 42 days. The PHA-reactive splenocyte population followed a similar temporal pattern. In contrast, the VBI-derived E population was at a maximum during early development and steadily declined throughout the larval period. DSC-derived E were undetectable during early development but steadily increased throughout the larval period. Both VBI- and DSC-derived hematopoietic cells persisted after metamorphosis and contributed to all populations examined in adult frogs. Because of temporal differences in the VBI and DSC contributions to the developing thymus, these data suggest heterogeneity within the thymocyte population associated with the embryonic origin of the colonizing stem cells.

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PubMed1991

Expression of a leukocyte-specific antigen during ontogeny in Xenopus laevis.

The monoclonal antibody CL21 recognizes a determinant present on the surface of leukocytes, but not on erythrocytes or nonhemopoietic tissue. The CL21 antigen was first expressed at 48 hr of development at 20 degrees C (stage 28) on embryonic cells cultured from lateral plate mesoderm. Based on immunofluorescence staining and flow cytometric analysis, the distribution of fluorescence intensity of larval thymocytes and splenocytes was unimodal. Distributions of dull and bright cells were detected in both adult thymocytes and splenocytes. These different subpopulations appeared during the late perimetamorphic period. Adult splenocytes were metabolically activated when cultured in the presence of mAb CL21 bound to a substrate but not in the presence of mAb CL21 in suspension. Immunoprecipitation and SDS-PAGE under nonreducing conditions revealed that a single 180-kD molecule was expressed on thymocytes. Analysis of splenocytes demonstrated the presence of two molecules having similar molecular mass that resolved to a single band under reducing conditions.

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PubMed1989

Location of hemopoietic stem cells influences frequency of lymphoid engraftment in Xenopus embryos.

The first hemopoietic stem cells to differentiate in Xenopus embryos arise from ventral blood island (VBI) mesoderm. Progeny of these stem cells contribute to larval E, macrophage, thymocyte, and B lymphocyte populations. When small pieces of mesoderm are transplanted to a central location within the VBI, the contribution of this mesoderm is predominantly to erythropoiesis and engraftment of lymphoid populations is minimal. The present experiments examined the influence of position within the VBI on the contribution of single stem cells to lymphoid populations. Pieces of diploid VBI mesoderm, containing an average of one hemopoietic stem cell, were transplanted to either a central or a peripheral location within the defined boundaries of the VBI of triploid, stage matched embryos. The number of animals with donor-derived cells in lymphoid populations was markedly increased when stem cells were grafted to a peripheral position. In three cases, stem cells contributed to lymphoid populations at the exclusion of erythroid populations. These data were consistent with the notion of either a lymphoid stem cell or restricted B and T lymphocyte precursors. These data also suggested that during embryogenesis, stochastic differentiation of hemopoietic stem cells was influenced by regional differences in the VBI microenvironment.

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PubMed1989

Precursor immigration and thymocyte succession during larval development and metamorphosis in Xenopus.

The developing thymus in Xenopus was examined at four different levels: 1) precursor immigration of cytogenetically distinct embryonic stem cells; 2) waves of colonization during tadpole life and metamorphosis; 3) inter-thymic exchange of cells between separate lobes; and 4) development of cortical and medullary thymocytes. Based on the flow cytometric analysis of cytogenetically distinct thymocytes, there were at least two periods of stem cell immigration into the thymus, one during early larval life and the second before or during metamorphosis. Within the thymus, cohorts of cells derived from the first wave of immigration expanded at different times. The initial expansion occurred before 35 days of development. Cells involved in the second period of expansion were also derived from the initial immigrants, expanded after 35 days, and resulted in a turnover of thymocytes during the larval period. Precursor cells entering the thymus during metamorphosis expanded and resulted in an additional replacement of thymocytes. Cortical and medullary thymocytes were isolated from animals that received embryonic stem cell grafts. No differences in the presence or absence, or in the percentages, of donor thymocytes in these different fractions were observed. When limiting numbers of stem cells were transplanted, several cases of asymmetrical thymic lobe colonization were observed. These data suggested that an inter-thymic exchange of cells did not occur during larval life.

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PubMed1989

Experimental analysis of ventral blood island hematopoiesis in Xenopus embryonic chimeras.

The frequencies and potentialities of hematopoietic stem cells from 20-hr-old Xenopus embryos were examined by transplanting cytogenetically distinct ventral blood island tissue from diploid to triploid embryos. Thirty-five-day-old larvae were examined for the presence of donor-derived cells in their erythrocyte, thymocyte, and B lymphocyte populations by analyzing DNA content using flow cytometry. These experiments demonstrated that B lymphocytes, as well as erythrocytes and thymocytes, were derived from the ventral blood island. Data obtained by transplanting graded sized pieces of ventral blood island suggested that restricted erythroid precursors were present within the region by 20 hr postfertilization. Differentiation of both B- and T-lymphoid precursors from small pieces of ventral blood island was markedly enhanced when this tissue was grafted onto peripheral areas within the blood island region. Analysis of these data using repopulation statistics suggested that circulating larval erythrocytes of ventral blood island origin were derived from six or seven precursors. Each lobe of the thymus was colonized by three precursors, one of which was ventral blood island derived.

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PubMed1986

Analysis of hemopoietic lineage of accessory cells in the developing thymus of Xenopus laevis.

The developmental history of accessory cells in the thymus was studied by grafting hemopoietic stem cells into cytogenetically distinct frog embryos (diploid-2N or triploid-3N) before the establishment of circulation and overt differentiation and colonization of the thymus. The DNA content of cortical thymocytes and circulating erythrocytes was quantified by staining with propidium iodide and measuring the amount of red fluorescence emitted by individual nuclei with the use of flow cytometry. Accessory cells from thymic medulla were separated by incubating for 2 hr on glass slides. For comparison, the developmental history of peritoneal macrophages was examined as representative, myeloid-derived phagocytic cells. DNA content of adherent cells was quantified by staining with the DNA-specific Feulgen reaction and measuring light absorption of individual nuclei by microdensitometry. Thymic accessory cells were subdivided into phagocytic and nonphagocytic phenotypes on the basis of latex bead ingestion. Phagocytic cells in the thymus were usually nonspecific esterase positive and phenotypically resembled peritoneal macrophages. Nonphagocytic cells from the thymus were usually esterase negative and had a dendritic morphology characterized by branched cytoplasmic extensions. Nonphagocytic cells were positive for cytoplasmic RNA based on staining with methyl green-pyronin Y. Phagocytic cells from both the thymus and the peritoneal cavity had no levels of cytoplasmic RNA detectable by this method. Analysis of the embryonic derivation of thymic accessory cells, based on the proportion of cells carrying the cytogenetic marker, demonstrated that thymic lymphocytes and thymic accessory cells were a concordant pair of cells, distinct from myeloid-derived erythrocytes and possibly macrophages. These experiments provide circumstantial evidence suggesting thymocytes and thymic accessory cells could arise from a bipotential precursor that diverges into these separate lineages after colonization of the epithelial thymic rudiment during early development.

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PubMed1985

Dorsal lateral plate mesoderm influences proliferation and differentiation of hemopoietic stem cells derived from ventral lateral plate mesoderm during early development of Xenopus laevis embryos.

The developmental pattern of dorsal and ventral lateral plate mesoderm was examined by explanting these regions and culturing them in serum-free conditions for 10 days. Under these circumstances, ventral lateral plate mesoderm gave rise to hemopoietic cells, and dorsal lateral plate mesoderm gave rise to stromal cells. Comparison of the differentiation pattern observed when dorsal and ventral mesoderm were cultured separately versus the pattern observed when dorsal and ventral mesoderm were cultured together revealed that dorsal mesoderm influenced the differentiation of ventral-derived hemopoietic cells. Ventral mesoderm cultured alone gave rise to erythrocytes and granulocytes. However, in the presence of dorsal mesoderm, ventral mesoderm gave rise to erythrocytes, monocytes, and granulocytes. Moreover, dorsal mesoderm functioned to maintain the proliferation and differentiation potential of progenitor cells as determined using a colony-forming cell assay (CFU-c).

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PubMed1985

Thymic non-lymphoid cells.

In formulating this summary of our simon-pure knowledge of the structure/function relationships in the thymus, we decided that the time may have come to introduce a suitable dose of cynicism to balance the sometimes hopeless optimism of the past. Are the non-lymphoid cells of the thymus necessary for thymic function? Probably, but not to the extent or uniqueness that some authors including ourselves have previously claimed; T cells can probably differentiate in other tissues but may acquire their preference for MHC class II in the thymus. Mouse thymic lymphoid cell traffic and surface phenotype has recently been summarized pictorally by Scollay and Shortman [95]. Briefly stated, within the thymus, cells are hatched, matched and then dispatched. Minimally, the non-lymphoid cells act either as scenically varied obstacles along the way, nurseries for newborn T cells, or as tombstones for life's disenfranchized, effete and autoaggressive thymocytes. Hassall's corpuscles are morphological structures unique to the thymus, which are most useful to medical students for identification of this tissue. Their function remains one of life's great mysteries. Morphologically, they are suitable companions to the more recently described strange multicellular complexes of lymphocytes and epithelial cells which might be functionally important. The thymus of the much studied inbred, environmentally mollycoddled, laboratory mouse has been often and majestically described. It is probably typical for that of man and most mammals. It may, however, be unrepresentative of the thymus of stressed and parasitized wild animals. Diseases of the thymus generally can be categorized as not having enough thymus, having a neoplastic thymus or having a thymus which does not work properly. The bottom line in our knowledge of thymic nonlymphoid cells is that if you are born without them, you get sick and die; unless, of course, you are a nude mouse in Omaha, in which case you just freeze to death.

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PubMed1985

Hemopoietic differentiation potential of cultured lateral plate mesoderm explanted from Rana pipiens embryos at successive developmental stages.

Ventral blood island mesoderm and dorsal lateral plate mesoderm were removed from Rana pipiens embryos at successive developmental stages (stages 13-19; 50-118 h) and cultured as individual explants in serum-free medium. After 5-7 days, the cultures were harvested, and differential counts were made of Wright-Giemsa-stained cells. Ventral blood island explants gave rise to cells of the myeloid lineage, suggesting that ventral blood island mesoderm was committed to hemopoiesis at the time of explant. Although erythrocytes were present in the cultures, granulocytes and monocyte/macrophages predominated. This differentiation profile occurred without the addition of any exogenous humoral factors. Monocyte/macrophages and immature precursor cells exhibited recurring inverse fluctuations with respect to one another. In all cases examined, cultures of dorsal lateral plate mesoderm showed marginal hemopoietic cell differentiation, suggesting a requirement for exogenous humoral factors and/or cell-cell interactions. When viewed in the context of previous studies from our laboratory, these results demonstrate that, in the amphibian embryo, there are two sources of hemopoietic stem cells separated both in space and time.

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PubMed1984

Differential contribution of dorsal and ventral lateral plate mesoderm to hemopoiesis during Rana pipiens embryogenesis.

Data obtained from studies on the origin and development of hemopoietic cells in several classes of vertebrate embryos argue for two distinct sources of hemopoietic cells, the intraembryonic dorsal lateral plate and the extraembryonic ventral blood island/yolk sac. In the present study, a stage by stage comparison of the hemopoietic potential of both of these regions was made during development of the frog, Rana pipiens. Either dorsal lateral plate or ventral blood island mesoderm was reciprocally transplanted between cytogenetically labeled triploid and diploid embryos. The ratio of donor-derived cells to host-derived cells (labeling index) was determined from Feulgen-stained DNA measurements of cells harvested from hemopoietic organs of young larvae. Blood island transplants consistently resulted in larvae with positive labeling of the circulating blood. Transplanted dorsal mesoderm supplied mesonephric granulocytes and thymocytes, but not circulating erythrocytes to larvae. However, the contribution of dorsal mesoderm to larval hemopoiesis fluctuated with respect to embryonic stage at transplantation.

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PubMed1983

Peritoneal exudate in larval Rana pipiens contains cells that are embryologically derived from dorsal lateral plate mesoderm.

The embryonic origin of cells functioning in a peritoneal exudate response of larval Rana pipiens was investigated. Normal larva received injections of sodium caseinate on day 0 and cells in the peritoneal cavity were assayed on days 3, 5, 7, 9, 11, 13 and 16. Experimental animals that received cytogenetically labeled transplants of dorsal mesoderm at 67-72 hours of development were injected with sodium caseinate on day 0 and assayed on days 5, 7 and 9. Feulgen-DNA microdensitometric analysis showed that the peritoneal exudate from days 5, 7, 9 contained hemopoietic cells that were derived from the embryonic transplant. Therefore, dorsal mesoderm gave rise to cells capable of functioning in a peritoneal exudate response to injections of sodium caseinate.

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PubMed1983

Dual contribution of embryonic ventral blood island and dorsal lateral plate mesoderm during ontogeny of hemopoietic cells in Xenopus laevis.

The early embryonic development of hemopoietic cells in Xenopus laevis was examined. Either dorsal lateral plate (DLP) or ventral blood island (VBI) mesoderm was reciprocally transplanted between cytogenetically distinct (2N or 3N) stage 14 to 19 (neural fold) embryos. F-DNA content of circulating erythrocytes was assayed at stages 40, 41, 43, 45, and 49. The F-DNA content of cells in the thymus and mesonephros was assayed at stage 49. F-DNA values were used to distinguish between donor or host origin of hemopoietic cells in individual animals. The results demonstrated that DLP mesoderm gave rise to a population of stem cells that colonized the thymus and mesonephros, but not the blood. VBI mesoderm gave rise to a population of stem cells that colonized the blood and thymus, but not the mesonephros. These experiments show that there are two stem cell compartments in the amphibian embryo, separated in both space and time.

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