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Biomedical subjects

C L Stewart

Publications and source records attributed to C L Stewart.

17 recordsLinked to original sources
PubMed1992

Blastocyst implantation depends on maternal expression of leukaemia inhibitory factor.

A critical point during mammalian pregnancy is the implantation of the blastocyst when the embryo attaches to the wall of the uterus. The autonomously developing preimplantation embryo then becomes dependent on the maternal environment for its continued development. Little is known about the regulation of implantation, except that a complex interaction between peptide and steroid hormones synchronizes the preparation of the uterus for implantation with the development of the embryo. Whether the implantation event is under maternal or embryonic control is also unclear (reviewed in refs 1, 2). We have previously shown that a cytokine, leukaemia inhibitory factor (LIF), is expressed in the uterine endometrial glands specifically on the fourth day of pregnancy. This burst of expression is under maternal control and always precedes implantation of the blastocyst. Here we report that transient expression of LIF in mice is essential for implantation. Females lacking a functional LIF gene are fertile, but their blastocysts fail to implant and do not develop. The blastocysts, however, are viable and, when transferred to wild-type pseudopregnant recipients, they can implant and develop to term.

Animals↗
PubMed1991

Uterine expression of leukemia inhibitory factor coincides with the onset of blastocyst implantation.

We have analyzed the expression of the cytokine leukemia inhibitory factor (LIF) during embryogenesis and in tissues of neonatal and adult mice. The site of the most abundant LIF expression is the uterine endometrial glands, specifically on day 4 of pregnancy. Analysis of LIF expression in pseudopregnant mice and in females undergoing delayed implantation showed that it is under maternal control and that its expression coincides with blastocyst formation and always precedes implantation. These results suggest that a principal function of LIF in vivo may be to regulate the growth and to initiate implantation of blastocysts.

Animals↗
PubMed1991

FVB/N: an inbred mouse strain preferable for transgenic analyses.

FVB/N mice offer a system suitable for most transgenic experiments and subsequent genetic analyses. The inbred FVB/N strain is characterized by vigorous reproductive performance and consistently large litters. Moreover, fertilized FVB/N eggs contain large and prominent pronuclei, which facilitate microinjection of DNA. The phenotype of large pronuclei in the zygote is a dominant trait associated with the FVB/N oocyte but not the FVB/N sperm. In experiments to generate transgenic mice, the same DNA constructs were injected into three different types of zygotes: FVB/N, C57BL/6J, and (C57BL/6J x SJL/J)F1. FVB/N zygotes survived well after injection, and transgenic animals were obtained with efficiencies similar to the F1 zygotes and much better than the C57BL/6J zygotes. Genetic markers of the FVB/N strain have been analyzed for 44 loci that cover 15 chromosomes and were compared with those of commonly used inbred strains. In addition to the albino FVB/N strain, pigmented congenic strains of FVB/N are being constructed. These features make the FVB/N strain advantageous to use for research with transgenic mice.

Alleles↗
PubMed1991

Embryonal carcinoma cells transfected with ZP3 genes differentially glycosylate similar polypeptides and secrete active mouse sperm receptor.

Mouse and hamster sperm receptors, called mZP3 (approximately 83,000 Mr) and hZP3 (approximately 56,000 Mr), respectively, are glycoproteins located in the ovulated egg zona pellucida. Certain of the glycoprotein O-linked oligosaccharides are essential for sperm receptor activity. Here, we transfected mouse embryonal carcinoma (EC) cells with mZP3 and hZP3 genes placed under control of a constitutive promoter. Transfected cells synthesized and secreted large amounts of the glycoproteins, called EC-mZP3 and EC-hZP3. Although the primary structures of mZP3 and hZP3 polypeptides (44,000 Mr) are very similar to one another, EC-mZP3 (approximately 83,000 Mr) and EC-hZP3 (approximately 49,000 Mr) were glycosylated to very different extents, such that they resembled their egg counterparts. Like egg mZP3, EC-mZP3 inhibited binding of sperm to ovulated eggs and induced sperm to acrosome-react in vitro. In addition, large numbers of sperm bound to aggregates of mZP3-transfected EC cells in vitro. On the other hand, unlike egg hZP3, EC-hZP3 did not exhibit either sperm receptor or acrosome reaction-inducing activity, and sperm failed to bind to aggregates of hZP3-transfected EC cells. Thus, transfected EC cells not only express sperm receptor genes, but also discriminate between very similar polypeptides with respect to glycosylation and, in the case of mZP3, add specific oligosaccharides essential for biological activity. In addition, the results demonstrate that EC cells can serve as a source for large amounts of functional mouse sperm receptor.

Animals↗
PubMed1991

Development to term of mouse androgenetic aggregation chimeras.

Diploid androgenetic eggs contain two sperm-derived genomes, and only rarely develop to the early somite stage. Also, previous studies have indicated that androgenetic eggs cannot be rescued in aggregation chimeras beyond embryonic stages. Paradoxically, in blastocyst injection chimeras made with androgenetic embryonic stem (ES) cells of the 129/Sv strain, we previously obtained considerable improvement in developmental potential. Although considerable death occurred in utero, overtly normal chimeric fetuses and occasional postnatal chimeras that developed skeletal abnormalities were observed. Consequently, we have re-evaluated the developmental potential of androgenetic aggregation chimeras utilizing androgenetic eggs of the 129/Sv strain, and of the BALB/c and CD-1 strains for comparison. Regardless of strain, androgenetic aggregation chimeras were generally more inviable than previously observed with androgenetic ES cell chimeras, and often the embryoproper was abnormal even when an androgenetic contribution was detected only in the extra-embryonic membranes. This is at least a partial explanation of the greater viability of androgenetic ES cell chimeras, as ES cells do not colonize significantly certain extra-embryonic tissues. Nevertheless, in the 129/Sv strain, occasional development of chimeras to term was obtained, and one chimera that survived postnatally developed identical skeletal abnormalities to those observed previously in androgenetic ES cell chimeras. This result demonstrates that at least one example of paternal imprinting is faithfully conserved in androgenetic ES cells. Also, the postnatal chimerism shows that androgenetic eggs can give rise to terminally differentiated cell types, and are therefore pluripotent. In contrast, only possibly one BALB/c and no CD-1 androgenetic aggregation chimeras developed to term. Therefore, the developmental potential of androgenetic aggregation chimeras is to some extent dependent on mouse strain.

Animals↗
PubMed1991

Lack of graft rejection in a renal transplant recipient with AIDS.

A renal transplant recipient developed evidence of human immunodeficiency virus (HIV) infection and severe opportunistic infection 44 months after transplantation. A strikingly reduced dosage of pharmacologic immunosuppression was required to maintain renal graft function. This may be the result of impaired helper T-cell function associated with the acquired immunodeficiency syndrome (AIDS).

Acquired Immunodeficiency Syndrome↗
PubMed1990

Androgenetic mouse embryonic stem cells are pluripotent and cause skeletal defects in chimeras: implications for genetic imprinting.

The inviability of diploid androgenetic and parthenogenetic embryos suggests imprinting of paternal and maternal genes during germ cell development, and differential expression of loci depending on parental inheritance appears to be involved. To facilitate identification of imprinted genes, we have derived diploid androgenetic embryonic stem (ES) cell lines. In contrast to normal ES cells, they form tumors composed almost entirely of striated muscle when injected subcutaneously into adult mice. They also form chimeras following blastocyst injection, although many chimeras die at early postnatal stages. Surviving chimeras develop skeletal abnormalities, particularly in the rib cartilage. These results demonstrate that androgenetic ES cells are pluripotent and point to stage- and cell-specific expression of developmentally important imprinted genes.

Animals↗
PubMed1989

The histology of distraction osteogenesis using different external fixators.

The local biology of distraction osteogenesis using Ilizarov and Wagner external fixators was compared in unilateral tibial lengthenings in two groups of eight dogs. Serial roentgenograms with histological correlation revealed some basic similarities as well as distinct differences between the two groups. All dogs fully bridged the experimental gap by intramembranous ossification. Subtle histological changes in the alignment of collagen bundles and subsequent ossification reflected the mechanical configuration of each fixator. The multiplanar, full-pin Ilizarov system delivered concentric load to the site of osteogenesis, inducing perfectly parallel columns of new bone. The uniplanar, one-half pin Wagner system induced angulation of the collagen and subsequent bone columns on the basis of eccentric load to the osteogenic interface. Despite a significant difference in axial rigidity, the Ilizarov and Wagner external fixators induced osteogenesis of equal volume.

Animals↗
PubMed1988

Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells.

Embryonic stem (ES) cells, the totipotent outgrowths of blastocysts, can be cultured and manipulated in vitro and then returned to the embryonic environment where they develop normally and can contribute to all cell lineages. Maintenance of the stem-cell phenotype in vitro requires the presence of a feeder layer of fibroblasts or of a soluble factor, differentiation inhibitory activity (DIA) produced by a number of sources; in the absence of DIA the ES cells differentiate into a wide variety of cell types. We recently noted several similarities between partially purified DIA and a haemopoietic regulator, myeloid leukaemia inhibitory factor (LIF), a molecule which induces differentiation in M1 myeloid leukaemic cells and which we have recently purified, cloned and characterized. We demonstrate here that purified, recombinant LIF can substitute for DIA in the maintenance of totipotent ES cell lines that retain the potential to form chimaeric mice.

Animals↗
PubMed1987

Expression of retroviral vectors in transgenic mice obtained by embryo infection.

Pre-implantation embryos were infected with the retroviral vector MMCV-neo, which carries the neomycin resistance (neo) gene and the v-myc gene. Three transgenic substrains (M-TKneo 1-3) were derived which stably transmit a single intact copy of the vector. In all of the substrains, expression of the neo gene from the internal thymidine kinase (TK) promoter was detected, with two of the substrains expressing the gene in all tissues analysed. In the third substrain, the vector had integrated on the X chromosome and neo expression varied between different tissues. A second series of transgenic mice were obtained with the retroviral vector SAX, in which the human adenosine deaminase cDNA (ADA) is under the control of an internal SV40 promoter. Four substrains (M-SAX 1-4) were analysed; however, no expression of the ADA cDNA was detected. In all mice, no expression was found of the genes under the control of the viral 5' long terminal repeats (LTRs). In the M-TKneo substrains the vector was hypomethylated irrespective of its expression whereas in the M-SAX mice the vector was hypermethylated. These results demonstrate for the first time that the TK promoter can apparently express a gene in all tissues of adult mice and that retroviral vectors with internal promoters may provide an alternative to DNA injection for the efficient expression of genes in transgenic mice.

Animals↗
PubMed1985

Expression of foreign genes from retroviral vectors in mouse teratocarcinoma chimaeras.

The formation of chimaeric mice from embryonic stem cells (EC and ES) carrying and exposing foreign genes is described. Two retroviral vectors were used to stably introduce genes, by virus infection and selection into EC and ES cells. In the first series EC cell clones were isolated that contained a single intact copy of the vector, which expressed the neomycin resistance gene (neo) from the 5' long terminal repeat (LTR). Following the formation of chimaeras with one of the clones, expression of the foreign gene was found in all chimaeric tissues examined. In the second series, a vector was used which contained the OK10 v-myc oncogene under the control of the 5' LTR as well as the neo gene expressed from an internal thymidine kinase (TK) promoter. ES cell clones were again isolated and used to form chimaeras. Although expression of the TK neo transcripts was maintained in the chimaeric tissues analysed, no expression of the v-myc transcripts from the 5' LTR was detected. These results show that genes can be introduced into mice, using clones of embryonic stem cells selected in vitro and that the expression of the genes can be maintained probably throughout development and in all tissues. The advantages this approach offers compared with DNA injection into eggs and the use of retroviruses as vectors for introducing genes into mice are discussed.

Animals↗
PubMed1985

Microvascular and histologic effect of circumferential wire on appositional bone growth in immature dogs.

Circumferential wires were placed around both midshaft femora of six immature dogs. On one side the wires were placed under the periosteum and on the other side the wires were placed over the periosteum. All wires were tightened to an equivalent tension. A diffuse growth of periosteal new bone occurred in those femora in which wires were placed under the periosteum but not in those over the periosteum. Corresponding microangiographs at 3 weeks in those femora in which wires were placed under the periosteum revealed dramatically increased medullary and periosteal vascularity coupled with histologic active trabecular bone formation. In those femora in which the wires were placed over the periosteum, even though the placement of the wires should supposedly have been the most detrimental, vascularity was not restricted. There were perfused vessels within the cortex directly under the wires. At 8 weeks the wires in both preparations were becoming encased in the growing cortical bone. Cerclage did not devitalize immature bone nor did it restrict adjacent appositional bone growth.

Angiography↗
PubMed1985

Clustered homeo boxes are differentially expressed during murine development.

A murine homeo box (m6-12) known to be expressed during differentiation of embryonal carcinoma cells lies within a cluster of homeo boxes located in 30 kilobase pairs of genomic DNA. We have established the organization of the boxes within this complex, as well as the nucleotide sequence of a second box, m5-4. Similar to the m6-12 box, expression of novel m5-4 transcripts is induced upon differentiation of embryonal carcinoma cells. Transcripts of genes containing m6-12 were found in embryonic tissue during almost all stages of prenatal development studied, whereas expression of m5-4 was detected only in 12 day embryonic tissue. Finally, we have described the differential expression of these homeo-box-containing regions in various adult tissues.

Amino Acid Sequence↗
PubMed1985

Vascularization of cancellous chip bone grafts.

Fresh autogenous, frozen allogeneic, and commercially prepared xenogeneic cancellous chip bone grafts were placed into plug holes in the tibia and into surgically simulated nonunion defects in the ulna of dogs. The microvascular and correlated histologic reactions were studied at given times up to 12 weeks after the graft implantation. Marked and characteristic differences were noted among the 3 graft materials. Fresh autogenous chips were extensively vascularized by 1 week, and subsequent microscopic healing was rapid. Allogeneic chips were incorporated into all defects, but vascularization and histologic healing rates were slower than those with the fresh autogenous graft material. The xenogeneic graft was poorly incorporated. A foreign body reaction was seen histologically, and corresponding vascularization of the graft was slow and incomplete.

Angiography↗
PubMed1985

Transitional nephrology.

During growth from fetal life to early childhood, renal function and parameters used to assess renal function undergo developmental changes. This article provides a brief review of these maturational changes and how they affect the evaluation and management of young patients with diseases affecting the genitourinary system. Included is a discussion of renal blood flow, glomerular filtration, sodium and water balance, clinical assessment of renal function, and age-related effects of nephrotoxins.

Adult↗
PubMed1979

Experimental reaming of the proximal femur and acrylic cement implantation: vascular and histologic effects.

The technique of microangiography and correlated histology was used, in experiments on the proximal femora of mature mongrel dogs, to study the effects of simple medullary reaming and of reaming followed by insertion of acrylic cement. The hip joints were not disturbed. A segment of Steinmann pin, representing the stem of a hip prosthesis, was inserted into the cement as it was polymerizing. The most significant results, after reaming alone, were devascularization and apparent necrosis of large areas of the cortex of the subtrochanteric femoral diaphysis. Full recovery was observed in 6 months. However, when acrylic cement was introduced following the reaming, extensive necrosis of the inner layers of the diaphyseal cortex was still present after a year, when the observations were terminated. The presence of necrosis of bone was judged by the lack of visible osteocytes in the lacunae. After the devascularization caused by filling the diaphyseal medulla with acrylic cement, the apparently necrotic cortex was revitalized without osteoclasia. That is in contrast to the results in another study in which osteoclasia and osteoneogenesis were required to restore cortex with empty cell lacunae. This raises the question of dormant osteonecrosis in contrast to total osteonecrosis. These studies show, in thin histologic preparations, acrylic bone cement in situ in the tissues. A fibrous membrane was found at the cement-bone interface in most areas. It was thicker in the diaphysis than in the metaphysis. The physiologic effects of acrylic cement implantation are considered to be likely factors in the loosening which is observed clinically when the femoral components of hip prostheses are under protracted stress.

Acrylic Resins↗