Biomedical subjects

P A Linnemeyer

Publications and source records attributed to P A Linnemeyer.

14 recordsLinked to original sources

PubMed1999

Mouse Ly-49D recognizes H-2Dd and activates natural killer cell cytotoxicity.

Although activation of natural killer (NK) cytotoxicity is generally inhibited by target major histocompatibility complex (MHC) class I expression, subtle features of NK allorecognition suggest that NK cells possess receptors that are activated by target MHC I. The mouse Ly-49D receptor has been shown to activate NK cytotoxicity, although recognition of MHC class I has not been demonstrated previously. To define Ly-49D-ligand interactions, we transfected the mouse Ly-49D receptor into the rat NK line, RNK-16 (RNK.mLy-49D). As expected, anti- Ly-49D monoclonal antibody 12A8 specifically stimulated redirected lysis of the Fc receptor- bearing rat target YB2/0 by RNK.mLy-49D transfectants. RNK.mLy-49D effectors were tested against YB2/0 targets transfected with the mouse MHC I alleles H-2Dd, Db, Kk, or Kb. RNK.mLy-49D cells lysed YB2/0.Dd targets more efficiently than untransfected YB2/0 or YB2/0 transfected with Db, Kk, or Kb. This augmented lysis of H-2Dd targets was specifically inhibited by F(ab')2 anti-Ly-49D (12A8) and F(ab')2 anti-H-2Dd (34-5-8S). RNK.mLy-49D effectors were also able to specifically lyse Concanavalin A blasts isolated from H-2(d) mice (BALB/c, B10.D2, and DBA/2) but not from H-2(b) or H-2(k) mice. These experiments show that the activating receptor Ly-49D specifically interacts with the MHC I antigen, H-2Dd, demonstrating the existence of alloactivating receptors on murine NK cells.

Amino Acid Sequence↗

PubMed1995

Interleukin-2.

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AIDS-Related Opportunistic Infections↗

PubMed1994

Stage-specific expression of activation antigens on NK cells at uterine implantation sites in mice.

The differentiation of the cellular components of the uterine decidua, in particular the life history of NK cells, is poorly understood. With the use of two mAbs that recognize stage-specific activation Ags on NK cells, we investigated the development of NK cells known as granulated metrial gland (GMG) cells. Immunohistochemical analysis demonstrated that mAb 3C2, but not mAb 4H12, recognized numerous cells throughout the uterine decidua basalis during early gestation. Isolated (panned) 3C2+ decidual cells from day 7 of pregnancy co-expressed the NK1.1 Ag, displayed NK cytolytic activity, and proliferated in IL-2-containing media. A small percentage of those cells expressed the GMG-associated Ag 4H12. Immunohistochemical analysis of serial sections at midgestation demonstrated that most of the 3C2+ NK cells co-expressed 4H12 Ag. During the later part of pregnancy, however, 3C2 expression in the decidua was down-regulated, and the cells expressed high levels of 4H12 Ag. When 3C2+ NK cells were isolated from cell suspensions of decidua from 7-day pregnant mice, and cultured in IL-2-containing medium, the cells developed the large and granular morphology characteristics of GMG cells, and acquired 4H12 Ag. These results demonstrate that 4H12+ GMG cells differentiate from 3C2+, NK1.1+, cytolytic precursors that reside in the decidua during early gestation.

Animals↗

PubMed1994

Induction of osteopontin mRNA expression during activation of murine NK cells.

Osteopontin (OPN) is an extracellular phosphorylated glycoprotein expressed in bone, kidney, nervous tissue, bone marrow, and granulated metrial gland (GMG) cells in murine decidua. We recently demonstrated that GMG cells are differentiated natural killer (NK) lineage cells that share phenotypic, functional, and morphologic characteristics with adherent interleukin-2 (IL-2)-activated NK cells. We now show that conditions that induce resting splenic NK cells to develop into adherent, activated cells induce the expression of opn mRNA. Nonstimulated NK cells did not express opn mRNA detectable by Northern analysis. However, expression was evident by day 1-2 of culture of NK cells with IL-2, increased to high levels by day 4, and was maintained at high levels thereafter. Thus, expression of mRNA for OPN, a secreted protein associated with cell adhesion, embryonic development, tissue remodeling, and immune regulation, is up-regulated during the activation of NK cells.

Animals↗

PubMed1993

Prostaglandin E2-induced changes in the phenotype, morphology, and lytic activity of IL-2-activated natural killer cells.

During pregnancy, many large granular NK lineage cells that are recognized by mAb 4H12 accumulate at uterine implantation sites. Similar large, granular, 4H12+ NK cells accumulate during the 2nd wk of culture of IL-2-activated NK cells. Inasmuch as uterine NK cell lytic activity is suppressed during pregnancy, apparently due to the effects of PGE2, we have analyzed the effect of PGE2 on the phenotype, morphology, and lytic activity of IL-2-activated NK cells as a model for NK cell differentiation at uterine implantation sites. When cultures of IL-2-activated NK cells were supplemented with 1 microM PGE2, the cells increased in size and granularity, accompanied by an increase in the number of cells expressing the 4H12 Ag. This effect was specific to PGE2 because PGF2 alpha at the same concentration had no effect. In addition, changes in morphology and up-regulation of 4H12 Ag expression could be augmented by 100 mM dibutyryl cAMP, but not 100 mM dibutyryl cGMP, suggesting that the PGE2 effects are mediated by changes in cytoplasmic cAMP levels. When indomethacin was added to cultures of IL-2-activated NK cells at the beginning of the culture period, 4H12 expression was markedly reduced. Indomethacin also increased the cytolytic capacity of NK cells and reduced the number of cells that developed the large granular morphology characteristic of 4H12+ cells. When activated NK cells were sorted into 4H12+ and 4H12- populations and tested for their ability to kill YAC-1 cells, we found that 4H12+ NK cells have lower lytic activity as compared to the 4H12- subset. Furthermore, when activated NK cells were labeled with [3H]TdR, combined with 4H12 labeling, we found that the 4H12+ subset was not proliferative. These results are likely to have direct relevance to NK cell differentiation at uterine implantation sites where NK cell activity is suppressed by PGE2, 4H12 expression is up-regulated and NK cells differentiate into large, granular and relatively nonlytic cells termed granulated metrial gland cells.

Animals↗

PubMed1992

Stimulation of growth and ion uptake in bean leaves by red and blue light.

Red and blue light both stimulate growth and ion accumulation in bean (Phaseolus vulgaris L.) leaves, and previous studies showed that the growth response is mediated by phytochrome and a blue-light receptor. Results of this study confirm that there is an additional photosynthetic contribution from the growing cells that supports ion uptake and growth. Disc expansion in the light was enhanced by exogenous K(+) and Rb(+), but was not specific for anions. Light increased K(+) accumulation and the rate of (86)Rb(+) uptake by discs, over darkness, with no effect of light quality. The photosynthetic inhibitor, 3-(3,4-dichlorophenyl)-1,1-dimethylurea, inhibited light-driven (86)Rb(+) uptake by 75%. Light quality caused differences in short-term kinetics of growth and acidification of the leaf surface. At comparable fluence rates (50 mumol m(-2) s(-1)), continuous exposure to blue light increased the growth rate 3-fold after a 2-min lag, whereas red light caused a smaller growth response after a lag of 12 min. In contrast, the acidification of the leaf surface normally associated with growth was stimulated 3-fold by red light but only slightly (1.3-fold) by blue light. This result shows that, in addition to acidification caused by red light, a second mechanism specifically stimulated by blue light is normally functioning in light-driven leaf growth.

Journal Article↗

PubMed1991

Murine granulated metrial gland cells at uterine implantation sites are natural killer lineage cells.

Granulated metrial gland (GMG) cells, a population of morphologically distinct, bone marrow-derived cells in murine decidua that react with mAb 4H12, are shown in this report to express NK-specific Ag and to become cytolytic to the NK cell target YAC-1 when cultured in the lymphokine IL-2. When 1-mm3 explants of 8-day decidual tissue were cultured with IL-2, large numbers of 4H12+ GMG cells migrated out of the tissue. Migration was dependent on the amount of IL-2 used. This explant technique was used to isolate a pure population of GMG cells. The migratory activated GMG cells were phenotypically 4H12+, NK1.1+, LGL-1+/-, CD3-, and MAC-1-. Furthermore, the IL-2-activated GMG cells killed YAC-1 but not P815 cells in a 4-h 51Cr-release cytotoxicity assay. 4H12+ GMG cells from collagenase-digested decidual tissue also were analyzed for the presence of NK lineage Ag by flow cytometry and shown to coexpress the NK-associated Ag NK1.1 and ASGM1 but not the T cell Ag CD3 or macrophage Ag MAC-1 or F4/80. GMG cells isolated by collagenase digestion did not express LGL-1, an Ag associated with lytic NK cells. Our results demonstrate that GMG cells express Ag and functions characteristic of NK cells, and thus GMG cells can be assigned to the NK lineage. The possible relevance of NK cells at implantation sites is discussed.

Animals↗

PubMed1991

Monoclonal antibody 4H12 recognizes subsets of adherent-lymphokine activated killer cells and splenic natural killer cells from pregnant and neonatal mice.

Using a new mAb, 4H12, that recognizes a plasma membrane-associated Ag on granulated metrial gland cells, we identified subsets of murine NK cells in spleen cell-derived adherent lymphokine activated killer cells and in the spleens of neonatal and pregnant mice. In spleen cell adherent-lymphokine-activated killer cultures, 4H12 Ag was detected on a small subset of cells after 7 days culture and expression increased with time to 70% of the cells after 21 days culture. 4H12+ cells were large (up to 70 microns) and granular. The Ag was also detected in the cytoplasmic granules of some, but not all 4H12 surface positive cells. Coexpression studies indicated 4H12+ cells were predominantly positive for the NK1.1 and ASGM1 Ag, negative for the MAC-1 and F4/80 Ag, and +/- for the LGL-1 and CD3 Ag. Subsets of 4H12+/-, LGL-1+/- exhibited morphologic characteristics restricted to specific phenotypic subsets. The 4H12-/LGL-1+ subset was shown to contain the smallest, least granular cells, whereas the 4H12+/LGL-1+/- subsets contained the largest and most granular cells. Although 4H12 expression was negligible in the spleens of normal adult mice, spleen cells of neonatal and pregnant mice contained subsets of NK1.1+ cells that coexpressed 4H12. The 4H12+/NK1.1+ and 4H12-/NK1.1+ subsets displayed differential levels of NK1.1 expression. 4H12+ NK cells were NK1.1 low to high, whereas 4H12- NK cells were NK1.1 high only. The functional significance of subsets of NK cells in IL-2 culture and in the spleens of neonatal and pregnant mice remains to be elucidated.

Animals↗

PubMed1990

A monoclonal antibody, 4H12, recognizes a surface antigen found on granulated metrial gland cells in the murine decidua.

A monoclonal antibody (MAb), designated 4H12, was selected for reactivity to a surface antigen on PYS-2 teratocarcinoma cells. 4H12 was the product of a fusion of lymphoid cells of a non-immunized pregnant C57BL/6 mouse to NS-1 myeloma cells. Initial studies utilizing immunohistochemistry revealed that MAb 4H12 bound to an antigen found on cells in the decidua basalis of 7-, 8- and 10-day pregnant mice. Antigen-positive cells of 11--19-day pregnant mice were also found predominantly in the decidua. A few antigen-positive cells were found in the labyrinth of the placenta and up against Reichert's membrane. Antigen-positive cells were morphologically and spatially distinct, oval to round with large periodic acid Schiff positive granules. Indirect immunofluorescent (IIF) labeling of decidual cultures showed antigen on the surface of cells that were small, oval to round and adherent. The antigen recognized by MAb 4H12 was removed from tissue sections with trypsin and protease and therefore is suggested to be a protein. We conclude that MAb 4H12 recognizes a surface antigen found on cells historically described as granulated metrial gland (GMG) cells. This MAb should greatly facilitate the further analysis of the life history and function of GMG cells during pregnancy.

Animals↗

PubMed1990

Characterization and effect of light on the plasma membrane H(+) -ATPase of bean leaves.

Proton excretion from bean (Phaseolus vulgaris L.) leaf cells is increased by bright white light. To test whether this could be due, at least in part, to an increase in plasma membrane (PM) ATPase activity, PM vesicles were isolated from primary leaves by phase partitioning and used to characterize PM ATPase activity and changes in response to light. ATPase activity was characterized as magnesium ion dependent, vanadate sensitive, and slightly stimulated by potassium chloride. The pH optimum was 6.5, the Km was approximately 0.30 millimolar ATP, and the activity was about 60% latent. PM vesicles were prepared from leaves of plants grown for 11 days in dim red light (growing slowly) or grown for 10 days in dim red light and then transferred to bright white-light for 1 day (growing rapidly). For both light treatments, ATPase specific activity was approximately 600 to 700 nanomoles per milligram protein per minute, and the latency, Km, and sensitivity to potassium chloride were also similar. PM vesicles from plants grown in complete darkness, however, exhibited a twofold greater specific activity. We conclude that the promotion of leaf growth and proton excretion by bright white light is not due to an increase in ATPase specific activity. Light does influence ATPase activity, however; both dim red light and bright white light decreased the ATPase specific activity by nearly 50% as compared with dark-grown leaves.

Azides↗

PubMed1989

A monoclonal antibody, MA21, recognizes a surface component that is present on F9 teratocarcinoma cells and that appears vectorially on the trophectoderm of peri-implantation-stage mouse blastocysts.

A monoclonal antibody (MAb) "MA21", derived from lymphoid tissue of a multiparous mouse and selected for binding to mouse teratocarcinoma cell line F9, recognizes a surface antigen that appears on peri-implantation-stage mouse blastocysts. In indirect immunofluorescence assays, MAb MA21 does not bind to 1-cell-through morula-stage embryos, nor to early, 3.5-day post-coitum (p.c.) blastocysts. When 3.5-day p.c. blastocysts are maintained 17 h in vitro and then assayed, MAb MA21 binds to a limited number of trophectoderm cells that are centered at the embryonic pole. As culture time lengthens, the number of antigen-expressing trophectoderm cells increases, forming a cap that spreads from the embryonic pole into the abembryonic region. Embryos maintained 48 h in vitro bind MAb MA21 over as much as 100% of the trophectoderm surface. MAb MA21 does not bind to the inner cell mass. When mouse pregnancy uteri are assayed by the immunoperoxidase method, MAb MA21 binds to extra-embryonic ectoderm and trophectoderm of 5-day p.c. implanted blastocysts, but does not bind to 6-day p.c. blastocysts. MAb MA21 recognizes a component with an estimated mol. wt of 44,000 from NP-40 detergent extracts of F9 cells and peri-implantation-stage mouse blastocysts. The component appears to be firmly associated with the plasma membrane; it is resistant to removal by high salt or moderate concentrations of non-ionic detergent.

Animals↗

PubMedDate not supplied

A trophoblast specific antigen, recognized by monoclonal antibody MA21, locates a unique trophoblast cell population in the murine placenta.

Monoclonal antibody MA21 recognized a 44kDa plasma membrane protein on F9 teratocarcinoma cells, trophectoderm of mouse peri-implantation-stage blastocyst and ectoplacental cone cells of 5 day postcoitum implanted blastocyst (Vernon, Linnemeyer and Hamilton, 1989). We show here that this antigen is expressed by trophoblast cells of the maturing placenta. Immunohistochemical assays of early and mature placental tissue sections, indirect immunofluorescence labelling of placental cultures and blastocyst outgrowths in vitro, and immunoprecipitation of 35S-labelled NP-40 extracts of placental cultures indicate the presence of a plasma membrane-associated antigen with the same characteristics as MA21 antigen of peri-implantation embryos and F9 teratocarcinoma cells. In sections of placentae, antigen-positive cells are always situated in a thin layer between trophoblastic giant cells and maternal tissue. In cultures of postimplantation stage embryos, attached trophoblast cells express MA21 antigen initially, but following transformation to the giant cell state, antigen is no longer expressed. These results indicate the presence of a plasma membrane protein antigen associated with a distinct population of cells believed to be trophoblast. We believe that these cells are the foremost trophoblast cells opposing maternal decidua and that they may give rise to secondary trophoblastic giant cells.

Animals↗

PubMedDate not supplied

Hamster monoclonal antibodies to murine natural killer cells.

To further the study of natural killer (NK) cells we have produced hamster monoclonal antibodies (MAbs) with reactivities to mouse NK cells. MAbs 4A2 and 3C2 were obtained by fusing spleen cells from Syrian hamsters immunized with IL-2-activated NK cells with Fox-NY myeloma cells. 4A2 antigen was expressed by bone marrow (BM)-derived IL-2-responsive NK cell precursors, by mature NK cells of the BM, and by a highly lytic subset of splenic NK cells, in addition to IL-2-activated NK cells. 3C2 antigen was also expressed by BM-derived NK cell precursors, by mature NK cells in the BM, at low levels by splenic NK cells, and at high levels by IL-2-activated NK cells. These MAbs are likely to provide useful reagents for the study of the life history and functional significance of NK cells.

Animals↗

PubMedDate not supplied

Natural killer cells in the nonpregnant murine uterus.

Recent studies on the origin and function of the large granular cells that accumulate at implantation sites in the rodent uterus during pregnancy have shown that these cells are highly differentiated natural killer (NK) cells. These findings raise questions about the presence and regulation of NK cells in the normal, nonpregnant uterus. For example, do NK cells comprise a major or minor population of leukocytes in the uterus? Are these uterine NK cells (uNK) similar in phenotype and function to NK cells in other organs? Is the population of uNK cells maintained by local proliferation and/or by influx via the blood stream from the bone marrow? This brief review will examine our current understanding of those questions based on the experimental literature on rodents and our own recent studies.

Animals↗