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Biomedical subjects

C J Jones

Publications and source records attributed to C J Jones.

At least 37 records · Page 2Linked to original sources
PubMed2001

Early implantation events in the baboon (Papio anubis) with special reference to the establishment of anchoring villi.

The development of the baboon anchoring villus has been studied from day 14 to day 48 of gestation, using light and electron microscopy. At day 14, cords of trophoblast could be seen streaming into the endometrium, invading maternal vessels and forming blood-filled lacunae; by 20 days gestation some of these had differentiated into distinctive anchoring villi, with an outer covering of syncytiotrophoblast and inner cytotrophoblast cells which differed from those of floating villi in that a subpopulation detached from the syncytium to form an interconnecting network of cells within the centre of the villus. Subsequent migration of cytotrophoblast into the endometrium formed the cytotrophoblastic shell while fibrillin-like extracellular matrix biosynthesis within the body of the villus provided a firm mechanical support. At the trophoblast-decidual interface, a zone of necrosis and phagocytosis initially developed, which became less extensive with time, so that by 40 days a stable interface was evident with only residual pockets of necrosis. During this period, there was differentiation of decidual cells which by 28 days developed characteristic pedunculated cell processes, and later became surrounded by a basal lamina. The factors that control detachment of cytotrophoblast from the syncytium and the biosynthesis of the specialized, fibrillar extracellular matrix, features that are not apparent in other placental villi, require further investigation, possibly by carefully controlled in vitro experimentation.

Animals↗
PubMed2001

Ultrastructure of epithelial plaque formation and stromal cell transformation by post-ovulatory chorionic gonadotrophin treatment in the baboon (Papio anubis).

BACKGROUND: To understand factors controlling endometrial responses to pregnancy, we have established a model using the baboon and examined the effects of infused human chorionic gonadotrophin (HCG) on the preparation of the luminal epithelium and stromal cell differentiation for the establishment of pregnancy. METHODS: The ultrastructure of endometrium from normal day 10 post-ovulation animals, cycling females treated with either HCG or FSH (control), and a day 15 pregnant animal has been compared. RESULTS: In the control endometrium, the luminal epithelium was smooth and regular, with underlying spindle shaped stromal cells. In pregnancy, the luminal epithelium underwent a plaque reaction, while stromal cells enlarged and developed filament-rich cell processes. Infusion of HCG produced changes similar to those seen in pregnancy, with generalized plaque formation and stromal decidualization, while in the animal treated with FSH there was no response. CONCLUSIONS: This study indicates that infusion of HCG into the uterus can duplicate many of the responses of the endometrium to pregnancy, although in this case the plaque reaction involved the whole of the luminal epithelium, rather than only the implantation site as in pregnancy.

Actins↗
PubMed2001

Muc1 and glycan expression in the oviduct and endometrium of a New World monkey, Cebus apella.

Cebus apella is a New World monkey that has a menstrual cycle of 18-23 days with implantation at approximately luteal Day 5. The aim of this study was to characterize by lectin- and antibody-labeling the distribution of Muc1 and associated glycans on the endometrial and oviductal epithelium during the luteal phase of the cycle. Endometrial histology showed a thin endometrium, with glands extending deeply into the myometrium. No obvious evidence of secretory differentiation in cells of either the superficial or the basal segments of glands could be obtained using a panel of antibodies and lectins that marked epithelial glycoprotein, and glycosylation changes observed in some other primate endometrial cycles were not observed in this study. Antibodies to human MUC1 were shown to cross-react with C. apella, and Muc1 was localized to the apical epithelial surfaces of both the endometrial and the tubal epithelium, with stronger expression in the latter. Again, no cyclic changes were noted. Antibodies specific to the isoform Muc1/Sec showed strong staining at the apical tubal epithelium, but no reactivity was detectable in the luminal epithelium of the uterus. This observation suggests differences between the two glycocalyces and could help to explain why C. apella embryos do not implant in this location.

Animals↗
PubMed2001

Human telomerase reverse transcriptase-immortalized MRC-5 and HCA2 human fibroblasts are fully permissive for human cytomegalovirus.

MRC-5 cells are a well-characterized human diploid fibroblast cell line approved for vaccine production and favoured for the routine propagation of human cytomegalovirus (HCMV). Ectopic expression of telomerase in fibroblasts is capable of overcoming replicative senescence induced by telomere shortening. Following delivery of the hTERT gene to MRC-5 cells using a retrovirus vector three clones were generated that (i) expressed functional telomerase activity, (ii) exhibited telomere extension and (iii) were sustained for >100 population doublings. Immortalized MRC-5-hTERT and also HCA2-hTERT human fibroblasts were both fully permissive for HCMV as determined by plaque assay, studies of virus growth kinetics and measurement of virus yields. Furthermore, telomerase-immortalized HCA2 cells proved capable of supporting the stable maintenance of an EBV-based episomal vector with efficient transgene expression when driven by the HCMV immediate early promoter. An indicator cell line suitable for the efficient detection of HCMV infection was also generated using an episome containing a reporter gene (lacZ) under the control of the HCMV beta-2.7 early promoter. Telomerase immortalization of human fibroblasts will thus facilitate the growth and detection of HCMV and also the generation of helper cell lines for the propagation of HCMV deletion mutants. Immortalization of fibroblasts by telomerase does not affect cell morphology or growth characteristics. The MRC-5-hTERT clones may therefore be suitable for additional applications in virology, cell biology, vaccine production and biotechnology.

Cells, Cultured↗
PubMed2001

The functional regeneration of syncytiotrophoblast in cultured explants of term placenta.

We have investigated the functional characteristics of term human placental villous explants kept in long-term (7-11 days) culture. Fragments of placental villous tissue (approximately 5-10 mg wet wt) were cultured in supplemented CMRL-1066 culture medium for up to 11 days. After the first day of culture, the syncytiotrophoblast appeared vacuolated and eventually degenerated. However, a new syncytiotrophoblast developed by day 4, being indistinguishable from that of a fresh placenta by 11 days. Release of human chorionic gonadotrophin increased and activity of lactate dehydrogenase in culture medium decreased with culture time. Transport variables were measured over the first 7 days of culture. Basal (86)Rb efflux was reduced with time in culture and was inhibited by Ba2+, suggesting the efflux was mediated by K+ channels. At all stages of culture, (86)Rb efflux was stimulated by ATP, hyposmotic medium, and ANG II. A complex pattern of efflux changes with culture time and type of stimulator was observed, suggesting that several compartments of the tissue contributed to stimulated efflux. This culture system provides opportunities for studies of chronic regulation of placental function.

Cell Survival↗
PubMed2001

A novel cell culture model of chondrocyte differentiation during mammalian endochondral ossification.

Endochondral ossification (EO) occurs in the growth plate where chondrocytes pass through discrete stages of proliferation, maturation, hypertrophy, and calcification. We have developed and characterized a novel bovine cell culture model of EO that mirrors these events and will facilitate in vitro studies on factors controlling chondrocyte differentiation. Chondrocytes derived from the epiphyses of long bones of fetal calves were treated with 5-azacytidine (aza-C) for 48 h. Cultures were maintained subsequently without aza-C and harvested at selected time points for analyses of growth and differentiation status. A chondrocytic phenotype associated with an extensive extracellular matrix rich in proteoglycans and collagen types II and VI was observed in aza-C-treated and -untreated cultures. aza-C-treated cultures were characterized by studying the expression of several markers of chondrocyte differentiation. Parathyroid hormone-related protein (PTHrP) and its receptor, both markers of maturation, were expressed at days 5-9. Type X collagen, which is restricted to the stage of hypertrophy, was expressed from day 11 onward. Hypertrophy was confirmed by a 14-fold increase in cell size by day 15 and an increased synthesis of alkaline phosphatase during the hypertrophic period (days 14-28). The addition of PTHrP to aza-C-treated cultures at day 14 led to the down-regulation of type X collagen by 6-fold, showing type X collagen expression is under the control of PTHrP as in vivo. These findings show that aza-C can induce fetal bovine epiphyseal chondrocytes to differentiate in culture in a manner consistent with that which occurs during the EO process in vivo.

Animals↗
PubMed2001

Flow-mediated dilatation following wrist and upper arm occlusion in humans: the contribution of nitric oxide.

Flow-mediated dilatation (FMD) of the brachial artery assessed by high-resolution ultrasound is widely used to measure endothelial function. However, the technique is not standardized, with different groups using occlusion of either the wrist or the upper arm to induce increased blood flow. The validity of the test as a marker of endothelial function rests on the assumption that the dilatation observed is endothelium-dependent and mediated by nitric oxide (NO). We sought to compare the NO component of brachial artery dilatation observed following wrist or upper arm occlusion. Dilatation was assessed before and during intra-arterial infusion of the NO synthase inhibitor N(G)-monomethyl-L-arginine (L-NMMA) following occlusion of (i) the wrist (distal to ultrasound probe) and (ii) the upper arm (proximal to ultrasound probe) for 5 min in ten healthy males. Dilatation was significantly greater after upper arm occlusion (upper arm, 11.62+/-3.17%; wrist, 7.25+/-2.49%; P=0.003). During L-NMMA infusion, dilatation after wrist occlusion was abolished (from 7.25+/-2.49% to 0.16+/-2.24%; P<0.001), whereas dilatation after upper arm occlusion was only partially attenuated (from 11.62+/-3.17% to 7.51+/-2.34%; P=0.006). The peak flow stimulus was similar after wrist and upper arm occlusion. We conclude that dilatation following upper arm occlusion is greater than that observed after wrist occlusion, despite a similar peak flow stimulus. L-NMMA infusion revealed that FMD following wrist occlusion is mediated exclusively by NO, while dilatation following upper arm occlusion comprises a substantial component not mediated by NO, most probably related to tissue ischaemia around the brachial artery. FMD following wrist occlusion may be a more valid marker of endothelial function than dilatation following upper arm occlusion.

Adult↗
PubMed2000

Olf-1, a neuron-specific transcription factor, can activate the herpes simplex virus type 1-infected cell protein 0 promoter.

Herpes simplex virus type 1 (HSV-1) establishes a life-long latent infection in sensory neurons of infected individuals. Infected cell protein 0 (ICP0) is important for productive infection and reactivation from latency. Thus, activation of ICP0 expression in neurons is likely to be important for reactivation from latency. In a mouse neuroblastoma cell line, ICP0 promoter activity is high compared with other strong viral promoters. In contrast, promoter activity is low in non-neuronal cells. DNase I footprinting assays indicated that three distinct motifs in the ICP0 promoter are bound by nuclear factors. One of these motifs contains a binding site for a novel helix-loop-helix olfactory neuron-specific transcription factor (Olf-1). Gel shift assays and supershift assays using an Olf-1-specific antibody demonstrated that mouse neuroblastoma cells express Olf-1, which is bound to the Olf-1-like site in the ICP0 promoter. Deletion of the putative Olf-1 motif reduced ICP0 promoter activity more than 5-fold in mouse neuroblastoma cells and prevented trans-activation by an Olf-1 expression vector. We hypothesize that the Olf-1-binding site activates ICP0 promoter activity in neurons during reactivation from latency.

Animals↗
PubMed2000

Posttranslational modification and plasma membrane localization of the Drosophila melanogaster presenilin.

Mutations in two genes, presenilin 1 (PS1) and presenilin 2, are linked to early onset cases of familial Alzheimer's disease. The presenilins are thought to contribute to the pathogenesis of Alzheimer's disease by directly or indirectly affecting the proteolytic processing of the amyloid precursor protein. They have also been implicated in the proteolytic processing of Notch. In PS1-deficient mammalian cells, the proteolytic release of the Notch intracellular domain is reduced. Likewise, loss-of-function mutations in Drosophila presenilin (Psn) prevent the production of the intracellular Notch signaling fragment and lead to phenotypes resembling Notch mutants. Here we characterize the Drosophila Psn protein and demonstrate that it undergoes a proteolytic cleavage. We describe Psn expression at different developmental stages of the fly and show Psn localization near both apical and basal plasma membranes. Furthermore, we demonstrate that portions of the Psn protein span the plasma membrane in S2 cells.

Alternative Splicing↗
PubMed2000

A vhs-like activity of bovine herpesvirus-1.

Bovine herpesvirus 1 (BHV-1) is a major pathogen of cattle, causing significant disease including immunosuppression in infected animals. In vitro, the surface expression of major histocompatibility complex (MHC) class I molecules, crucial for an appropriate anti-viral immune response of the host, is down-regulated by BHV-1 infection. Northern blot analyses revealed that the mRNAs for MHC class I and class II molecules were significantly down-regulated in BHV-1 infected cells, starting as early as 2 h after infection. Furthermore, mRNA expression of the two house keeping genes actin and glyceraldehyde-6-phosphate dehydrogenase (GAPDH) was also repressed after infection. This BHV-1 induced effect on cellular metabolism resembled the virion host shutoff (vhs) activity of herpes simplex virus (HSV). Similar to the HSV vhs activity, the putative BHV-1 vhs activity was not abrogated in cells infected in the presence of actinomycin D (ActD) which suggested that no viral gene expression is required for the vhs function and the putative vhs protein is associated with the virion. Sequence comparison indicated a BHV-1 open reading frame having a 60% similarity to the HSV vhs sequence. This putative BHV-1 open reading frame contained the four conserved regions of the alphaherpesvirus vhs protein. Since an HSV vhs-mutant exhibited less virulence and good immunogenicity, we suggest that a BHV-1 vhs- mutant may hold promising potential as a candidate vaccine.

Actins↗
PubMed2000

Dietary supplementation with marine omega-3 fatty acids improve systemic large artery endothelial function in subjects with hypercholesterolemia.

OBJECTIVE: This work was undertaken to determine whether dietary supplementation with marine omega-3 fatty acids improve systemic large artery endothelial function in subjects with hypercholesterolemia. BACKGROUND: Marine omega-3 fatty acids improve vascular function, but the underlying mechanism(s) are unclear. We studied the effects of marine omega-3 fatty acids on large artery endothelial function in subjects with hypercholesterolemia. METHODS: Hypercholesterolemic subjects with no other known cause for endothelial dysfunction were recruited to a prospective, placebo-controlled, randomized, double-blind, parallel-group study. Treatment with omega-3 fatty acids at a dose of 4 g/day (n = 15/group) was compared with placebo, at the beginning (day 0) and end (day 120) of a four-month treatment period. Endothelial function was assessed pre- and posttreatment by noninvasive ultrasonic vessel wall tracking of brachial artery flow-mediated dilation (FMD). RESULTS: Treatment with marine omega-3 fatty acids resulted in a significant improvement in FMD (0.05 +/- 0.12 to 0.12 +/- 0.07 mm, p < 0.05) and a significant reduction in triglycerides (2.07 +/- 1.13 to 1.73 +/- 0.95 mmol/liter, p < 0.05), whereas treatment with placebo resulted in no change in FMD (0.03 +/- 0.10 to 0.04 +/- 0.10 mm) or triglycerides (2.29 +/- 2.09 to 2.05 +/- 1.36 mmol/liter) (both p < 0.05 treated compared with control). Responses to sublingual glyceryl trinitrate were unchanged. CONCLUSIONS: Marine omega-3 fatty acids improve large artery endothelium-dependent dilation in subjects with hypercholesterolemia without affecting endothelium-independent dilation.

Arteriosclerosis↗
PubMed2000

Localization of pN-type IIA procollagen on adult bovine vitreous collagen fibrils.

Type II procollagen is synthesized in long (type IIA) and short (type IIB) forms because of alternative splicing of mRNA; the long form containing an additional cysteine-rich domain in the amino-propeptide. An antiserum (IIA) that recognizes this domain was used for immunolocalization studies on adult bovine vitreous at light and electron microscopic levels and for Western blot analyses. The immunolocalization studies revealed labelling by the IIA antiserum of the vitreous collagen fibrils. This labelling was removed by prior extraction of the fibrils with 6 M guanidine hydrochloride (GuHCl) and the extract was shown to contain pN-type IIA procollagen. Adult vitreous collagen fibrils are coated with pN-type IIA procollagen, a finding with potential implications for vitreous collagen fibril structure and function.

Animals↗
PubMed2000

Growth factor-extracellular matrix synergy in the control of trophoblast invasion.

At the periphery of the human placenta, trophoblast attaches to the uterine wall. The tissue interface contains many anchoring sites, with cytotrophoblast columns that form bridges between the overlying extraembryonic (villous) mesenchyme and the maternal decidual stroma beneath. From the periphery of these columns, large numbers of trophoblast cells detach, migrate through the decidua and eventually colonize and transform maternal arteries. In this way the placenta increases and gives priority to the maternal blood supply to the conceptus. We have shown that when early villous tissue is explanted on a collagen gel in serum-free medium, anchoring-site morphogenesis occurs. Thus, in the presence of placental mesenchyme but in the absence of maternal cells, contact with a permissive extracellular matrix (ECM) is necessary and sufficient for cytotrophoblast column development. Proliferation of trophoblast occurs, followed by differentiation into a columnar cell phenotype in which cells remain attached to one another and to the ECM. At this stage, interaction between fibronectin and integrin alpha5beta1 at the cell surface stabilizes the column and the cells remain as a contiguous multilayered sheet. However, the addition of serum-free conditioned medium from first-trimester placental fibroblasts stimulates cytotrophoblast to detach from the distal column and migrate in streams across the ECM. The removal of insulin-like growth factor I (IGF-I) from the fibroblast medium decreases streaming activity, whereas the addition of exogenous IGF-I (10 ng/ml) to serum-free medium produces a streaming phenotype. In contrast, transforming growth factor beta1 (10 ng/ml) maintains the cells in a tight sheet. These results suggest the possibility of a paracrine interaction between villous mesenchyme and cytotrophoblast in anchoring sites to stimulate the infiltration of the maternal ECM by trophoblast. Such a mechanism would be self-limiting because the signal diminishes with distance from the placenta.

Animals↗
PubMed2000

Mosaic characteristics of human endometrial epithelium in vitro: analysis of secretory markers and cell surface ultrastructure.

Specific terminal carbohydrate structures and mucin-associated glycans increase in expression within the human endometrial epithelium during the secretory phase of the menstrual cycle but exhibit wide intercellular variation. We postulated that variation in glycosylation between cells would produce differences in the glycocalyx and result in complex mixtures of cells bearing different combinations of glycans. MUC-1 mucin, keratan sulphate and fucosylated lactosaminoglycans were examined in epithelial gland fragment cultures with antibodies (HMFG1, 5D4) and a lectin (Dolichos biflorus agglutinin). The glycocalyx was examined by transmission and high resolution scanning electron microscopy. The data were related to patterns of expression seen in vivo. The MUC-1 mucin was expressed relatively uniformly in culture, but heterogeneity was evident in mucin sialylation within the epithelial cell population. Double labelling of gland explant cultures for combinations of fucosylated lactosaminoglycans, keratan sulphate and MUC-1 demonstrated cells expressing all combinations of these markers. Ultrastructural examination confirmed remarkable intercellular variation in the glycocalyx. Though the human endometrial epithelium is relatively morphologically homogeneous, these observations reveal complex variations of cell surface glycosylation between neighbouring cells and suggest that secretory function might vary in a similar fashion.

Adult↗
PubMed2000

Evidence for a telomere-independent "clock" limiting RAS oncogene-driven proliferation of human thyroid epithelial cells.

An initiating role for RAS oncogene mutation in several epithelial cancers is supported by its high incidence in early-stage tumors and its ability to induce proliferation in the corresponding normal cells in vitro. Using retroviral transduction of thyroid epithelial cells as a model we ask here: (i) how mutant RAS can induce long-term proliferation in an epithelial cell in contrast to the premature senescence observed in fibroblasts; and (ii) what is the "clock" which eventually triggers spontaneous growth arrest even in epithelial clones generated by mutant RAS. The early response to RAS activation in thyroid epithelial cells showed two features not seen in fibroblasts: (i) a marked decrease in expression of the cyclin-dependent kinase inhibitor (CDKI) p27(kip1) and (ii) the absence of any induction of p21(waf1). When proliferation eventually ceased (after up to 20 population doublings) this occurred despite undiminished expression of mutant RAS and was tightly correlated with a return to the initial high level of p27(kip1) expression, together with the de novo appearance of p16(ink4a). Importantly, neither the CDKI changes nor the proliferative life span of RAS-induced epithelial clones was altered by induction of telomerase activity through forced expression of the catalytic subunit, hTERT, at levels sufficient to immortalize human fibroblasts. These data provide a basis for cell-type differences in sensitivity to RAS-induced proliferation which may explain the corresponding tumor-type specificity of RAS mutation. They also show for the first time in a primary human cell model that a telomere-independent mechanism can limit not only physiological but also oncogene-driven proliferation, pointing therefore to a tumour suppressor mechanism additional, or alternative, to the telomere clock.

Catalytic Domain↗