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Genetic polymorphism in the H-2:Qa:Tla region of chromosome 17 is associated with constitutive variation of bodily odor phenotypes which permit individual olfactory recognition among mice. To determine whether known genes in the H-2:Qa:Tla complex are concerned in the constitution of odor phenotypes, mice were tested for their ability to sense a difference between the B6/By (H-2b) and congeneic B6.C-H-2bm1 strains, which differ genetically by mutation of the H-2K gene. As in previous studies of the sensory discrimination of H-2:Qa:Tla phenotypes, mice were trained by reward in a Y maze to distinguish the odors of urine samples, and the successful distinctions of B6/By from B6.C-H-2bm1 were confirmed by transfer of training, without reward, to coded samples of urine from genetically equivalent urine donor mice which the trained mice had not previously encountered. Cosegregation of odor phenotype with H-2b and H-2bm1 was demonstrated by transfer of training to typed H-2b and H-2bm1 homozygous segregants of F2 generations of appropriate crosses. Although it is not excluded that the differences in odor phenotype which distinguish H-2b and H-2bm1 mice are directly related to the structure of the H-2b and H-2bm1 products, it is equally possible that H-2-related odor phenotypes arise from effects of H-2 genetic variation on metabolic pathways either directly, or indirectly through developmental polymorphism.

Murine fetal thymus lobes isolated from both normal and scid/scid mice can be colonized by donor cells from either human bone marrow or human umbilical cord blood in vitro. Subsequent organ culture results in a transient production of a few CD4+ CD8+ (double-positive) cells and then the accumulation of CD4+ or CD8+ (single-positive) T cells. A significant number of immature T-cell intermediates (e.g., CD8low, CD3-/low cells) were present in early organ cultures, suggesting that these were progenitors of the mature CD3+/high single-positive T cells that dominated late cultures. Depletion of mature T cells from the donor-cell populations did not affect their ability to colonize thymus lobes. However, colonization depended on the presence of CD7+ progenitor T cells. Limiting dilution experiments using mature T-cell populations (human peripheral blood leukocytes, human bone marrow cells, and human umbilical cord blood cells) suggested that thymic organ culture supports the growth of progenitor T cells but does not support the growth of mature human T cells. Each of these donor populations produced single-positive populations with different CD4/CD8 ratios, suggesting that precursor cells from different sources differ qualitatively in their capacity to differentiate into T cells.

Successful implementation of bone marrow transplantation for hematopoietic reconstitution is limited by the lack of suitably HLA-matched donors and by the occurrence of graft-versus-host disease that frequently accompanies this procedure. Recent clinical reports have implied that the use of umbilical cord blood as a source of transplantable stem cells may solve these problems. To date, definitive experiments have not been performed to assess the immunological potential of T cells found in umbilical cord blood, which could mediate graft-versus-host disease. In the present study we have observed that umbilical cord blood contains T lymphocytes that appear to be phenotypically immature. In addition, umbilical cord blood lymphocytes appeared to be functionally immature as shown by minimal responses to stimulation with interleukin 2, phytohemagglutinin, or alloantigens. Thus, umbilical cord blood may be more suitable for allogeneic transplantation than bone marrow in that these cord blood cells may not be as capable of mediating graft-versus-host disease.

A DNA-dependent in vitro-coupled transcription-translation system has been prepared from lysates of isolated chloroplasts. These lysates are comparable to those of Escherichia coli in transcriptional and translational fidelity and efficiency in response to a given template DNA. When Nicotiana tabacum chloroplast DNA is used as template with chloroplast lysates (N. tabacum or spinach) or E. coli lysates, NaDodSO4 gel analysis reveals similar polypeptide patterns that are distinct from the patterns obtained with E. coli DNA. Genes in recombinant plasmids containing chloroplast DNA are also expressed in these in vitro systems. DNA . RNA hybridization experiments show that transcripts are synthesized from most of the chloroplast genome. Newly synthesized large subunit of ribulosebisphosphate carboxylase/oxygenate and a transcript of the large subunit gene (rbcL) are observed in chloroplast lysates using as template chloroplast DNA or cloned fragments of tobacco chloroplast DNA that contain the large subunit gene. Results suggest that differential expression of chloroplast genes occurs in vitro. By using cloned chloroplast DNA templates in this homologous system, it is possible to identify and map structural genes for chloroplast proteins.

An intronless gene encoding an additional human serotonin (5-HT) 5-HT1-like receptor subtype was isolated from a human genomic library with probes obtained from degenerate PCR primers used to amplify 5-HT-receptor-specific sequences. The highest degree of homology was found with the 5-HT1E subtype (70%) and the 5-HT1D alpha (63%) and 5-HT1D beta (60%) receptors. RNA for this gene was detected in the human brain but was not detected in kidney, liver, spleen, heart, pancreas, and testes. High-affinity (Kd = 9.2 nM) 3H-labeled 5-HT binding was detected. Competition studies revealed the following rank order of potencies for serotonergic ligands: 5-HT > sumatriptan >> 5-carboxyamidotryptamine > 8-hydroxy-2(di-1-propylamino)tetralin > spiperone. 5-HT produced a dose-dependent inhibition of forskolin-stimulated cAMP accumulation (EC50 = 7.9 nM) in transfected cells. These properties distinguish this receptor from any previously characterized and establish a fifth 5-HT1-like receptor subtype (5-HT1F) coupled to the inhibition of adenylate cyclase.

The purpose of this study was to evaluate human umbilical cord blood as an alternative to bone marrow in the provision of transplantable stem/progenitor cells for hematopoietic reconstitution. Although no direct quantitative assay for human hematopoietic repopulating cells is at present available, the granulocyte-macrophage progenitor cell (CFU-GM) assay has been used with success as a valid indicator of engrafting capability. We examined greater than 100 collections of human umbilical cord blood for their content of nucleated cells and granulocyte-macrophage, erythroid (BFU-E), and multipotential (CFU-GEMM) progenitor cells, in many cases both before and after cryopreservation. First it was determined that granulocyte-macrophage, erythroid, and multipotential progenitor cells remained functionally viable in cord blood untreated except for addition of anticoagulant for at least 3 days at 4 degrees C or 25 degrees C (room temperature), though not at 37 degrees C, implying that these cells could be satisfactorily studied and used or cryopreserved for therapy after transport of cord blood by overnight air freight carriage from a remote obstetrical service. Granulocyte-macrophage progenitor cells from cord blood so received responded normally to stimulation by purified recombinant preparations of granulocyte-macrophage, granulocyte, and macrophage colony-stimulating factors and interleukin 3. The salient finding, based on analysis of 101 cord blood collections, is that the numbers of progenitor cells present in the low-density (less than 1.077 gm/ml) fraction after Ficoll/Hypaque separation typically fell within the range that has been reported for successful engraftment by bone marrow cells. Another observation of practical importance is that procedures to remove erythrocytes or granulocytes prior to freezing, and washing of thawed cells before plating, entailed large losses of progenitor cells, the yield of unwashed progenitor cells from unfractionated cord blood being many times greater. The provisional inference is that human umbilical cord blood from a single individual is typically a sufficient source of cells for autologous (syngeneic) and for major histocompatibility complex-matched allogeneic hematopoietic reconstitution.

Genetically determined body odors that distinguish one mouse from another are termed odortypes. The best known odortypes, highly expressed in urine, are those specified by H-2, the major histocompatibility complex of the mouse, but other odortypes originate from unidentified loci in the rest of the genome, including both sex chromosomes. The definition of H-2 odortypes and evidence that their perception affects reproductive behavior have so far depended on studies with inbred mouse strains whose genetic differences are confined to the H-2 complex of genes. To simulate feral conditions more closely, a freely segregating population was bred from crosses involving four unrelated inbred strains contributing four different H-2 haplotypes. After H-2 typing, this outbred population was divided into four groups of freely segregating mice, comprising the four distinct H-2 genotypes represented, to serve as conventional donors of urine for evaluation in the standard Y-maze system used in the training and testing of mice for H-2 odortype discrimination. With respect to utility in training mice for H-2 odortype discrimination, and to degrees of concordance attained in the Y-maze by trained mice, these urinary H-2 odortype sources from outbred mice were no less effective than urines customarily obtained for those purposes from nonsegregating inbred donors. We conclude that discrimination of H-2 odortypes is not appreciably affected or impaired by the usual concurrent segregation within the genome as a whole.

The major histocompatibility complex (MHC) is the prime but not exclusive determinant of genetically specific constitutive body odors, termed odor types, represented strongly in urine of the mouse. Perception of MHC-determined odor types influences reproductive behavior in the contexts of mate choice and maintenance of early pregnancy, tending to favor the propagation of one MHC type over another. How MHC genotype determines MHC odor type is unknown. One possible explanation is that differential odorants are generated by populations of commensal microorganisms whose composition is somehow geared to MHC diversity. This hypothesis was tested in the Y-maze system in which mice are trained to distinguish the urinary odors of MHC-congenic mice. First, it was shown that mice could readily be trained to distinguish the urines of germfree MHC-congenic mice. Second, it was shown that mice trained to distinguish the urines of conventionally maintained MHC-congenic mice could as readily distinguish the urines of germfree MHC-congenic mice. These results imply that MHC-determined odor types do not depend on odorants generated by microorganisms.

Odortypes--namely, body odors that distinguish one individual from another on the basis of genetic polymorphism at the major histocompatibility complex and other loci--are a fundamental element in the social life and reproductive behavior of the mouse, including familial imprinting, mate choice, and control of early pregnancy. Odortypes are strongly represented in urine. During mouse pregnancy, an outcrossed mother's urine acquires fetal major histocompatibility complex odortypes of paternal origin, an observation that we took as the focus of a search for odortypes in humans, using a fully automated computer-programmed olfactometer in which trained rats are known to distinguish precisely the odortypes of another species. Five women provided urine samples before and after birth, which in each case appropriately trained rats were found to distinguish in the olfactometer. Whether this olfactory distinction of mothers' urine before and after birth reflects in part the odortype and hence genotype of the fetus, and not just the state of pregnancy per se, was tested in a second study in which each mother's postpartum urine was mixed either with urine from her own infant or with urine of a different, same-aged infant. Responses of trained rats were more positive with respect to the former (congruous) mixtures than to the latter (incongruous) mixtures, implying that, as in the mouse, human fetal odortypes of paternal genomic origin are represented in the odortype of the mother, doubtless by circulatory transfer of the pertinent odorants.

The extended H-2 complex of genes in the mouse includes at least three loci that independently specify distinctive body odors, "odortypes," whose differential recognition influences mating choice and affects the maintenance of early pregnancy. A prime experimental method of identifying H-2 odortypes is the specially designed Y-maze in which mice are trained, by water deprivation and reward, to distinguish odors conducted to the arms of the maze from H-2-dissimilar mice or their urines. It is confirmed that H-2-dissimilar infant mice, unlike adult mice, are not distinguished by trained mice in the Y-maze. However, a previous conclusion that infant mice do not express H-2 odortypes is shown to be incorrect, because the urines of H-2-dissimilar infant mice, even at 1 day of age, were distinguished in the Y-maze. Thus urine, ingested by the mother, clearly could suffice for her to distinguish her own from other H-2-dissimilar pups. Further, urine would seem to be a unique source of H-2 odortypes. If, as we believe, H-2 odortypes represent mostly compound odors composed by H-2 genetic variation in the urinary output of odorous metabolites, as distinct from simple odors that depend on chemical differences of single odorants, then the kidney, which is not responsible for H-2 odortype specificity, may nevertheless impart a unique character to urinary odortypes by virtue of differential excretion/resorption processing of various constituent odorous metabolites. In that case, various organs and tissues, among which the hematopoietic/lymphoid system is known to contribute to H-2 odortype specificity, may exhibit tissue-specific varieties of H-2 odortypes, their products having not yet been subjected to renal processing.