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Biomedical subjects

B A Croy

Publications and source records attributed to B A Croy.

At least 55 records · Page 3Linked to original sources
PubMed1996

Offspring of xenogeneically-reconstituted scid/scid mice are capable of a primary xenogeneic immune response to DNP-KLH.

Human peripheral blood leukocyte (PBL) reconstitution of severe combined immunodeficient (SCID) mice has provided a small animal model system (hu-PBL-SCID) useful for the study of the human immune system and disease pathogenesis. Transfer of xenogeneic PBL from donors other than humans has also been successful; however, the controversy remains regarding the capability of xenogeneically engrafted lymphocytes to mount a primary immune response. Human cells have been identified in offspring from hu-PBL-SCID but were not evaluated for a primary immune response. In the present study, offspring of bovine PBL-reconstituted SCID mice (F1-PBL-SCID-bo) were assessed for specific immune function. Sera from all of the F1-PBL-SCID-bo contained relatively low levels of bovine IgG 5 weeks after birth but bovine Ig became undetectable by 14 or 18 weeks. Eight F1-PBL-SCID-bo (23 or 27 weeks of age) were immunized with a single dose of 100 mu g dinitrophenyl-keyhole limpet hemocyanin (DNP-KLH). Individual cells secreting bovine antibody were enumerated using the ELISA-plaque assay. One week after immunization, bovine cells secreting bovine immunoglobulin (IgG) specific for DNP-KLH were identified in the spleens from three of the F1-PBL-SCID-bo at a frequency of one antibody-secreting cell per 9 x 10(3) to 1 x 10(6) spleen cells. Thus, xenogeneic lymphocytes, passed from the mother to her offspring, retain the capacity for a primary immune response to DNP-KLH.

Adoptive Transfer↗
PubMed1996

Granulated lymphocytes of pregnancy.

In pregnant mammals, the antigenically distinctive conceptus implants and grows in a uterine environment governed by the maternal immune system. The uterus per se is not immunologically privileged and large numbers of lymphocytes accumulate at implantation sites. In mice and humans, many of these lymphocytes have been identified as uterine natural killer (uNK) cells and exhibit a characteristic granulated morphology. In this review we focus on uNK cells and discuss their origin, differentiation and possible roles in the maintenance of healthy pregnancies. In species with less invasive placentation (ruminants, pigs), lymphocytes with similar granular morphology also appear during gestation and their identity and possible functions are examined.

Animals↗
PubMed1996

A study on the relationship between parity and differentiation of granulated metrial gland cells.

Granulated metrial gland (GMG) cells are a feature consistently associated with rodent pregnancy. Little information is available, however, on the life history of GMG cells in pregnancies other than first pregnancy. To determine if the frequency of GMG cells in the mesometrial triangle of mice is altered by parity, a study of histological sections from pregnant uteri of several females of genotype C.B-17 scid/scid.bg/bg being retired from a breeding program was undertaken. Pregnancies in this study ranged from first to 13th; day 12.5 of gestation was used for all analyses. The frequency of GMG cells in a constant, measured area of the metrial gland on day 12.5 of pregnancy was not significantly altered between first and second pregnancies. A small but statistically significant decline in GMG cell frequency was observed in two females who had given birth to 10 or more litters. However, neither the size of individual day 12.5 GMG cells, nor their granularity changed with parity. The surface area of the placenta in sections was not altered by the decrease in GMG cells. These data suggest that the differentiation of GMG cells from their non-granulated progenitors is a tightly regulated process, that GMG cell progenitors may be fully mature in first pregnancy, and that depletion of GMG cell progenitors is not induced by advanced parity or age.

Animals↗
PubMed1996

Maternal cells are widely distributed in murine fetuses in utero.

Passage of maternal cells into conceptuses in utero is recognized but poorly defined in species with hemochorial placentation. Despite the potential importance for such a phenomenon in vertical disease transmission, only limited data address the frequency of material to fetal cell trafficking or the developmental stage of its initiation. A murine model system, involving transfer of LacZ-, scid/scid, or wild type (+/+) blastocysts to pseudo-pregnant, LacZ+ transgenic ROSA26 females provided both flow cytometric and in situ information. In 100% of the late-gestation pregnancies studied, nucleated LacZ+ maternal cells crossed to conceptuses. In 90% of scid/scid fetuses, nucleated maternal cells were present in at least one lymphoid organ and often in more than one organ. Thymus was the most frequent site for maternal cell detection while the highest proportions of maternal cells were found in liver. Maternal cells were also visualized in fetal lung, heart, and bone marrow. Maternal cell trafficking into scid/scid fetuses commenced about midgestation, coincident with maturation of a placental circulation. In late-gestation +/+ fetuses, maternal cells were found extensively throughout bone marrow but not in other organs. The presence of maternal cells within primary lymphoid organs of fetuses may influence the repertoire of the developing fetal immune system and may be an underappreciated mechanism for vertical disease transmission.

Animals↗
PubMed1996

Maintenance of decidual cell reaction by androgens in the mouse.

In the mouse, estrogen and progesterone are required to prime the uterus for decidual cell reaction (DCR) in response to an intraluminal stimulus and, once DCR is induced, progesterone is required to maintain DCR. However, some evidence indicates that certain nonprogestational steroid hormones may also be involved in regulating DCR. The present study determined whether androgen plays any role in DCR. Adult CD1 mice were ovariectomized and treated with a regimen of estradiol and progesterone to prime the uterus for DCR and to maintain DCR. Sesame oil was injected into the uterine lumen to induce DCR on Day 5 of the treatment. DCR was determined by deciduomal weight-the difference between the wet weights of oil-injected and noninjected uterine horns. Testosterone, given at 1 mg/day during Days 3-5, could not replace progesterone in priming the uterus for DCR. However, the same dose of testosterone given during Days 6-8 maintained DCR. Alkaline phosphatase activity, a bio-marker for DCR, was present in the deciduoma maintained by either progesterone or testosterone, although the distribution of this enzyme activity was more intense in the antimesometrial pole in progesterone-maintained deciduoma. A nonaromatizable androgen, 5 alpha-dihydrotestosterone (DHT), was also effective in maintaining DCR, and this action of DHT was blocked by an androgen receptor antagonist, hydroxyflutamide. The relative potency of DHT in maintaining DCR was similar to that of progesterone. However, the regression of the deciduoma appeared to be advanced in DHT-treated mice. Ovariectomy on Day 6 of pregnancy resulted in resorption of the conceptus and regression of the decidua within 48 h. Treatment with DHT at the time of ovariectomy could not prevent fetal resorption, but it delayed the regression of decidua, as indicated, in part, by the presence of granulated metrial gland cells. In summary, androgen cannot prime the uterus for DCR, but it can maintain DCR once it is induced. The physiological significance of this finding remains to be determined.

Alkaline Phosphatase↗
PubMed1996

Pregnancy-associated uterine granulated metrial gland cells in mutant and transgenic mice.

PROBLEM: Granulated metrial gland (GMG) cells are pregnancy-specific uterine lymphocytes found in rodents. The lineage relationships of GMG cells are incompletely defined, although genetic and immunophenotyping studies suggest membership in the natural killer (NK) cell lineage. Pregnancy-specific functions have been postulated for GMG cells, but no successful depletion of these cells has been achieved that would permit assessment of any critical roles that might influence gestational outcome. METHOD: Routine histological methods for light microscopy were used to assess implantation sites from wild-type mice and mice of the following genotypes: tgE26, mi/mi, and p53 knockout. RESULTS: GMG cells are transient, histamine-negative cells found in the pregnant uteri of most mice. Pregnancies in the NK and T-cell-deficient tgE26 were characterized by infrequent GMG cells, very small placentae, and an elevated rate of fetal and perinatal mortality. In term placentae of mi/mi with dystocia. GMG cells were found in a new location along the plane of placental separation, and they appeared degranulated. A normal life-history was observed for GMG cells in p53 knockout mice. CONCLUSION: Pregnancies in mutant and transgenic mice indicate that GMG cells are natural killer cells and have critical roles in placental development and pregnancy success. The disappearance of GMG cells at term is independent of p53 gene expression.

Animals↗
PubMed1996

Accounting for the peripartum loss of granulated metrial gland cells, a natural killer cell population, from the pregnant mouse uterus.

Natural killer (NK) cells become a prominent cell population in the rodent uterus during pregnancy. The mature, heavily granulated form of these cells is rare in virgin or postpartum uteri. Death, migration, or dedifferentiation could account for the disappearance of these cells from late gestation uteri. We asked whether uterine NK cells, also known as granulated metrial gland (GMG) cells, die in situ and if expression of Fas antigen is essential for their death. Late in gestation, fragmentation of nuclear DNA was detected histologically by OH-end labeling, as were ultrastructural changes suggesting cell death. NK cells developed in and were lost from the uteri of pregnant Fas antigen-deficient lpr/lpr mice. Postpartum samples of retained placentas contained some residual NK cells that had moved from regions of uterine musculature toward the uterine lumen and were being expelled with the placenta. Thus, both cell death and placental separation remove NK cells from the peripartum uterus.

Animals↗
PubMed1995

Effects of triiodothyronine supplements on splenic natural killer cells in malnourished weanling mice.

The main purpose of this investigation was to determine whether exogenous triiodothyronine (T3) administered according to a protocol known to prevent depression in acquired immunity in weanling murine protein-energy malnutrition (PEM) would, likewise, influence the splenic natural killer (NK) cell in this disease. Weanling mice of disparate inbred strains, C57BL/6J and CBA/J, were subjected to wasting PEM produced by means of two low-protein diets (0.5% crude protein) identical in every respect except that one diet contained supplemental T3 (0.2 micrograms/g diet). NK cell lytic activity toward YAC-1 targets was assessed in vitro using suspensions containing 0.5 x 10(6) mononuclear spleen cells. Lytic activity in this assay was low in mice fed the unsupplemented low-protein diet, but was not depressed in malnourished animals given exogenous T3. Surface marker analysis using the NK cell-specific antigen, NK 1.1 (PK 136), revealed no effect of the low-protein diet or of exogenous T3 on the proportion of splenic mononuclear cells exhibiting NK 1.1+ phenotype. Previous investigations have shown that acquired immune competence in PEM can be manipulated, by means of endocrine hormonal intervention, independently of continued wasting disease. The present results extend this fundamental new concept to include an innate immune function, namely NK cell lytic activity. In this system of experimental PEM, exogenous T3 prevented depression in NK cell lytic activity expressed on a per cell basis. The malnourished weanling rodent is a particularly powerful experimental system with which to investigate the mechanisms whereby thyroid hormones influence NK cells.

Animals↗
PubMed1994

Immunohistochemical analysis of beta 1-integrin receptors displayed by murine uterine natural killer cells over the course of successful pregnancy.

Granulated metrial gland (GMG) cells are a natural killer (NK) cell-like population present in large numbers in the pregnant rodent uterus. By day 8 of gestation GMG cells are large and granulated and localized to the mesometrial side of each implantation site. GMG cells appear to be highly migratory both in vivo and in vitro; however, little is known regarding their functions. Using indirect fluorescence immunohistochemistry, murine uteri and implantation sites were studied on successive days of gestation to characterize the extracellular matrix receptors of the VLA-integrin family displayed by GMG cells. On days 3 and 6 of gestation, double immunostaining using the monoclonal antibody LGL-1 was employed to recognize GMG cells because their morphology early in pregnancy resembles that of other lymphocytes. Between days 8-15 of gestation, GMG cells can be recognized by their unique morphology. The day 3 and day 6 LGL-1+ cells were positive for all antigens examined; that is, beta 1 plus alpha 1, alpha 3, alpha 4, alpha 5 and alpha 6. From days 8-15 of gestation, GMG cells were beta 1+, alpha 4+, alpha 5+ but alpha 1-, alpha 3-, alpha 6-. Thus, between days 6-8 of gestation, major changes occur in the uterine NK/GMG cell population which include the loss of the surface molecules VLA alpha 1, alpha 3 and alpha 6 or the rapid expansion of NK cells not expressing these proteins. It is postulated that major changes in the functions of uterine NK cells are likely to be associated with these alterations in cell surface phenotype and that functional studies of uterine NK cells should focus upon this relatively early gestational time point.

Animals↗
PubMed1994

Preliminary characterization of lymphoid hybridoma cell lines derived from the pregnant mouse uterus.

Seven independent cell lines were derived from the fusion of migratory cells recovered from explant cultures of metrial glands to SP 2/0, a non-Ig secreting B cell myeloma. The migrating cells came from a pool of metrial glands from day 6-8 pregnant random bred CD1 mice and were assumed to be cells early in the differentiation pathway to granulated metrial gland (GMG) cells. The fused cells were cloned twice at the limiting dilution. Hybridization was confirmed by quantitation of cellular DNA using propidium iodide staining and by karyotyping. Electron microscopy revealed that each of the hybrid cell lines was composed of cells which were lymphoid in appearance, but lacked the granules found in mature GMG cells. The surface phenotype of all lines is CD45+, LGL-1-, asialo GM-1-, IgG-, IgM-, CD3- and CD25- (p55 of IL-2 receptor). Although the hybridomas lack those phenotypic markers which were used to show that GMG cells are related to the natural killer (NK) cell lineage (ie LGL-1, asialo GM-1), they do express the pan-leukocyte marker CD45 as well as the lytic protein, perforin, at levels intermediate to those of SP 2/0 cells and GMG cells. In addition, the hybridomas were observed to preferentially bind the NK target cell YAC and to be capable of lytic activity at temperatures below 30 degrees C. Because these hybridomas may represent fusion to an early progenitor cell of the NK/GMG cell lineage, their continued characterization is of merit.

Animals↗
PubMed1994

Granulated metrial gland cells: hypotheses concerning possible functions during murine gestation.

Granulated metrial gland (GMG) cells are morphologically distinctive lymphoid cells found in the murine uterus only during gestation. The life history of GMG cells suggests that they have important roles during mammalian gestation but these roles have been difficult to define. Genetic and immunologic data suggest that GMG cells may be a specialized subset of natural killer (NK) lymphocytes. This has directed research on GMG cell functions towards questions of effector cell-target cell interactions. A broader range of potential functions is discussed and shifts in functional roles played by GMG cells are proposed over the course of gestation.

Animals↗
PubMed1994

Lysis of porcine trophoblast cells by endometrial natural killer-like effector cells in vitro does not require interleukin-2.

Cells with cytotoxic activity against the cell line K562 and expressing perforin have been demonstrated in endometrial cells isolated from pigs early in pregnancy. This study was designed to determine whether porcine trophoblast cells were susceptible to these endometrial effector cells in vitro. Pregnant gilts (n = 8) were slaughtered between Days 17 and 20 of gestation. Immediately after slaughter, both the endometrial effector cells and trophoblast cells were isolated enzymatically from each animal. Enzymatically dispersed endometrial cells were further fractionated by size at unit gravity, whereas trophoblast cells were enriched by discontinuous density centrifugation on Percoll. Cytolytic activity was evaluated against Na2 51CrO4-labeled trophoblast and K562 cells. Comparison was made between freshly prepared and interleukin-2 (IL-2)-stimulated effector cells. The effect of prostaglandin E2 (PGE2) was tested by including it directly in the 51Cr-release assay. The results indicated that porcine trophoblast cells, like K562 cells, could be recognized and directly lysed by endometrial effector cells. Preculture of effector cells with IL-2 was not required for target lysis but enhanced their cytolytic activity against both trophoblast and K562 targets. In contrast, PGE2 exhibited highly suppressive effects on the cytotoxic activity of both freshly isolated and IL-2-stimulated endometrial effector cells. Conjugate assays demonstrated the binding of trophoblast and K562 targets by effector cells of similar morphology. Cold-target inhibition assays suggested that the effectors in porcine endometrial cell preparations that killed trophoblast and K562 cells were the same NK cell-like population.

Animals↗
PubMed1994

Signs and lesions of experimental Sendai virus infection in two genetically distinct strains of SCID/beige mice.

The pathogenesis of Sendai virus infection was studied in genetically immunodeficient mice of genotype scid/scid.bg/bg (SCID-beige) using C.B-17 SCID-beige mice, a BALB/c-related strain that expresses the same major histocompatibility complex as the Sendai virus-susceptible DBA/2 (H-2d). Mice were inoculated intranasally with isolate 771076 of Sendai virus, then killed at 2-day intervals beginning on day 4 post-inoculation. Clinical signs were evident beginning at 8 to 10 days post-inoculation, and all animals remaining were killed in extremis by 14 to 17 days post-inoculation. Lesions in inoculated mice were confined to the respiratory tract. In the nasal passages, a nonresolving rhinitis, with epithelial hyperplasia/metaplasia occurred. Cranioventral bronchopneumonitis was characterized by marked hyperplasia and necrosis of epithelial cells lining airways and with leukocytic infiltration. At the alveolar level, there was marked hypertrophy and hyperplasia of type II pneumocytes, mobilization of alveolar macrophages, and obliteration of the normal architecture in severely affected areas. Viral antigen was evident beginning at 4 days post-inoculation and persisted in affected areas throughout the duration of the study. Because immunocompetent C57BL/6 mice are known to be genetically resistant to Sendai virus, the susceptibility of C57BL/6 SCID-beige to Sendai virus was then compared to that of C.B-17 SCID-beige mice. In age-matched animals of the two strains, there was no evidence of natural resistance to Sendai virus infection in the immunodeficient C57BL/6 strain compared to the C.B-17 mice. These studies indicate that the genetic differences in susceptibility of two strains of immunocompetent mice to Sendai virus infection are eliminated by expression of the mutations scid and beige.

Animals↗
PubMed1994

Clones of tumor cells derived from a single primary human lung tumor reveal different patterns of beta 1 integrin expression.

Previously we reported that over 75% of human non-small cell lung cancers overexpress the beta 1 integrin VLA-2 on their surface and show an increase in the mRNA encoding the alpha-2 chain of this integrin. These results suggested the possibility that the overproduction and overexpression of one or more of the beta 1 integrin may be involved in the pathogenesis of human lung tumors by modulating the invasive and/or metastatic potential of the tumor. We report here the generation and characterization of multiple clones of tumor cells derived from the primary culture of cells obtained from biopsy tissue of an aggressive human squamous cell lung tumor. We show that these tumor clones (or clonotypes) exhibit seven different yet stable phenotypes with respect to the expression of five members of the beta 1 integrin family. These results illustrate that a primary human lung tumor consists of multiple subpopulations of cells that while indistinguishable by ultrastructure are heterogeneous with respect to their beta 1 integrins. The availability of these distinct tumor clonotypes derived from a single tumor biopsy have made it possible to test the assumption that the beta 1 integrins play a role in tumor progression. The feasibility of this approach is demonstrated here by the intravenous inoculation of different human tumor clonotypes into severe combined immunodeficient (scid) mice. Our preliminary results with a pair of tumor clonotypes differing in VLA-1 and VLA-2 expression level reveal that the clonotype with high level of VLA-1 and VLA-2 displays a substantial increase in the experimental engraftment and metastasis of the human tumor cells in scid mice.

Animals↗
PubMed1994

Enhancement of pancreatic tumor metastasis in transgenic immunodeficient mice.

Metastatic pancreatic cancer presents a bleak prognosis. Typically, human tumor development has been modelled in animals by generating transgenic mice carrying an oncogene, and metastasis studied by engrafting human tumor cells into immunodeficient mice. We derived mouse lines that spontaneously develop metastatic pancreatic cancer by crossing a transgenic line that develops primary pancreatic adenocarcinomas with lines that are deficient for different lymphocyte components of the immune system. We obtained transgenics carrying the SCID mutation resulting in loss of B and T cell function, those carrying the beige mutation resulting in impaired NK cell and macrophage activity, and those carrying both mutations. Although human graft studies indicated that the SCID mutation permits metastasis of different types of tumor cells, in our mice its effect on metastasis of the pancreatic tumor was minimal. In contrast, the beige mutation resulted in metastasis in almost 90% of the animals. The SCID and beige mutations synergistically resulted in faster growing tumors. Both primary tumors and metastases contained undifferentiated and differentiated cell types. The tissue distribution of metastases was similar to that recorded from human patients with pancreatic cancer, suggesting that mechanisms underlying metastasis in these mice could be similar to those involved in human disease.

Adenocarcinoma↗