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RESEARCH |
Department of Histology and Embryology and 1 Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil
Correspondence should be addressed to E Bevilacqua; Email: bevilacq{at}usp.br
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Abstract |
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and complement component C3. Placental trophoblast cells from the second half of gestation show basal phagocytosis that can be dramatically up-regulated by these promoters when microorganisms are inoculated into pregnant animals or introduced into culture systems. Stimulated trophoblast cells phagocytosed organisms more rapidly and in greater numbers than non-stimulated trophoblast exposed to the same numbers of organisms. Taken together, our results indicate that trophoblast cells do not lose their ability to phagocytose during the placentation process, which may imply that trophoblast cells participate in embryonic and fetal innate immune defense through elimination of microorganisms present at the maternal–fetal interface. |
Introduction |
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Over the course of gestation, phagocytic activity decreases significantly, whereas other functional features such as endocrine activity become more prominent (Soares & Glasser 1987, Soares et al. 1993 , 1996, Kanai-Azuma et al. 1994). From a nutritional viewpoint, this decrease is consistent with the differentiation of the placental labyrinth zone, where definitive molecular exchanges are established.
Under in vitro conditions, erythrophagocytosis by rodent trophoblast giant cells has been observed in the presence of lipopolysaccharides (either alone or combined with interferon- (IFN-)), phorbol myristate acetate and all-transretinal (Albieri & Bevilacqua 1996, Gagioti et al. 1996, Albieri et al. 2001). Once phagocytic, trophoblast cells produce and release reactive oxygen and nitrogen species such as superoxide anion, hydrogen peroxide and nitric oxide, suggesting that trophoblast phagocytosis plays a role in microbial defense mechanisms (Gagioti et al. 1995, 1996, 2000). Under in vitro conditions, trophoblast cells are able to phagocytose microorganisms (Drake & Rodger 1987), reinforcing the hypothesis of a possible trophoblast defense mechanism that contributes to removal of infectious agents from the fetal–maternal interface (Simmons & Russel 1962, Pavia 1983, Guilbert et al. 1993, Kanai-Azuma et al. 1994).
In addition to phagocytosis, there are many characteristics that trophoblast giant cells have in common with macrophages, including the ability to invade tissues and the potential for producing or responding to specific regulatory proteins. Such proteins include granulocyte macrophage-colony stimulating factor (GM-CSF) (Athanassakis et al. 2000), colony stimulating factor-1 (CSF-1) (Athanassakis-Vassiliadis et al. 1993, Athanassakis et al. 2000, Guleria & Pollard, 2000, Sasmono et al. 2003), interleukin (IL)-1 (Paulesu et al. 1991, Yamaguchi et al. 1996), IL-6 (Yamaguchi et al. 1996), tumor necrosis factor 
and ß (Kohchi et al. 1991, Whiteside et al. 2003), and transforming growth factor and ß (Sharma, 1998, Schaffer et al. 2003). These lineages share the markers CD4 and CD14 (Athanassakis et al. 1990, Clark et al. 2003), produce specific esterases, and possess receptors for IgG (FcR) (Guilbert et al. 1993), C3b complement system particle (Albieri et al. 1999) and for IFN- (Athanassakis et al. 2000, Albieri et al. 2001). These shared properties have led to the hypothesis that trophoblast cells, although different in lineage from macrophages, are part of a microbial defense system (Guilbert et al. 1993, Guleria & Pollard 2000, Sasmono et al. 2003). Few studies have investigated the degree to which trophoblast cells may contribute to fetal defense. We hypothesized that trophoblast cells, after placental maturation, maintain phagocytic activity associated with functions other than nutrition. In this report, we experimentally demonstrate the ability of mid-gestation trophoblast cells to phagocytose yeast cells Saccharomyces cerevisiae and bacteria Escherichia coli in the presence of promoters of phagocytosis in vivo and in vitro.
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Materials and Methods |
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In vivo phagocytic assays involving S. cerevisiae
Since mediators of immune and inflammatory responses could be of crucial importance in signaling and stimulating trophoblast cells, we tested the ability of the trophoblast to phagocytose yeast cells in vivo. The number of pregnant animals in each of the four experimental groups varied from 1 to 3 time point. In one group (n = 21), macrophages were previously activated by thioglycolate (Cohn, 1978, Cohen et al. 1981, Call et al. 2001, Salins et al. 2001). In this group, the animals received 300 µl 4% sodium thioglycolate in sterile saline, i.p., on day 12.5 of gestation and after 15 h 300 µl of a supersaturated solution of S. cerevisiae, administered i.v. Using the same routes of administration in the other three groups (n = 35) the pregnant females received, respectively, 300 µl of the following solutions: saline solution and then yeast (n = 3 mice/time point), thioglycolate followed by saline (n = 1 mouse/time point) and saline followed by saline (control group, n = 1 mouse/time point).
To quantify yeast particles, blood smears were prepared from tail blood. Test and control mice were topically anesthetized (5% xylocaine, AstraZeneca Pharmaceutical Ind., São Paulo, SP, Brazil) and, under minimal stress, the tail skin was perforated with a 36-gauge needle. A few drops of blood were immediately collected and, after 15 min, spread onto three histological slides. Blood collection was repeated at 1 h and some animals were immediately killed by cervical dislocation. Blood collection and subsequent death were then carried out at 2, 3, 6, 9 and 24 h after yeast administration. Placentas and spleens were collected for morphological studies and for tissue smears. For the smears, three fragments of placenta and spleen from each animal were macerated on a histological slide and washed in PBS to remove excess tissue. The remaining material was then air-dried, fixed in methanol and stained with Giemsa or hematoxylineosin. At the time of each assay, blood was collected from all living mice and their blood films were analyzed. For the morphological studies, some placentas and minced spleen fragments from each euthanized mouse were processed histologically.
Light and electron microscopic analyses
For light microscopic analysis, the placenta and spleen were cut into small fragments of 2–3 mm in thickness and 5–8 mm in length and fixed for 2 h in 4% paraformaldehyde (Sigma) in 0.1 M PBS, pH 7.4, containing 0.1 M sucrose. After fixation, the specimens were washed in PBS, dehydrated in ethanol and embedded in plastic resin (Historesin, Leica Microsystems Nusslock, Heidelberg, Germany) or paraplast (Merck Chemical Co., São Paulo, Brazil). Sections of 2 µm in thickness were stained with 1% Toluidine blue in distilled water, or Giemsa. For ultra-structural studies, placentas and spleens were cut into 2-mm3 fragments and fixed in 2% glutaraldehyde in 0.1 M PBS, pH 7.4, containing 0.1 M sucrose, for 12 h, post-fixed in 1% osmium tetroxide for 1 h, dehydrated in ethanol and embedded in Spurr’s resin (Sigma). The placentas were first cut into four quadrants and the fragments were obtained from the most angular region, in such a way that decidua, junctional zone and the labyrinth were always represented in 2 to 3 small fragments. The placental fragments obtained from each placental region were then carefully identified in separated flasks. For ultrastructural analyses, samples from each segment were randomly selected. Thin sections were stained with 1% uranyl acetate and 0.5% lead citrate, and examined in a JEOL CX-II-100 electron microscope.
In vitro phagocytic assay involving S. cerevisiae and E. coli
Placental cells, obtained as described below, were cultured for 48 h, then used for phagocytic assays employing S. cerevisiae and E. coli as targets for phagocytosis. The effects of IFN- (Sigma) and complement component C3 (as normal mouse plasma prepared in our laboratory) on this phagocytic activity were also evaluated. Phagocytosis was quantified and qualitatively analyzed through morphological examination.
Placental cell primary cultures
Fragments from each day-13.5 placenta were cut into very small pieces in a laminar flow hood (Vecco Ind., Campinas, Brazil). The fragments were treated with 15 ml trypsin (Instituto Adolfo Lutz, São Paulo, Brazil) in sterile tubes for 20 min at room temperature followed by 20 min incubation at 37 °C. The trypsin was then inactivated in Dulbecco’s modified Eagle’s medium (D-MEM, Sigma) containing 20% fetal calf serum (FCS, Cultilab S.A., Campinas, Brazil) and the material was centrifuged at 1000 r.p.m. for 10 min. The supernatant was resuspended in 1 ml culture medium supplemented with FCS and introduced into a 60 mm culture plate containing six removable 18mm diameter glass cover slips. The cultures were maintained under standard culture conditions for 48 h to allow attachment and spreading of the placental cells. The characterization of the trophoblast cells within the placental cell population was performed using morphological patterns associated with cytokeratin A-B immunolocalization.
Immunolocalization of cytokeratin filaments
Cells grown on the cover slips were fixed in 4% paraformaldehyde in 0.1 M PBS, pH 7.2, and washed several times in 0.02 M TBS (tris-hydroxymethyl aminomethane buffered saline, Amersham Biosciences, Uppsala, Sweden), pH 8.2, containing 0.05% saponin (Polyscience Inc., Warrington, PA, USA). Blocking of endogenous alkaline phosphatase was verified by incubation for 8 min in 5% acetic acid, and that of nonspecific binding by incubation for 1 h at 37 °C with TBS-1% bovine serum albumin (BSA, Sigma) containing 0.05% saponin and 0.2% glycine, pH 8.2. Rat IgG anti-mouse cytokeratin A-B (Troma, DSHB, University of Iowa, Iowa City, IA, USA) was used as the primary antibody at a dilution of 1:10 and incubated at 4 °C overnight. The secondary antibody used was either biotinylated anti-rat IgG or Cy3 anti-rat IgG (both from Sigma), at 37 °C for 1 h of incubation. The material receiving the biotinylated antibody was developed using an ABC kit (alkaline phosphatase-avidinbiotin complex, BioStain Super kit, Biomeda Co., Foster City, CA, USA) as recommended by the manufacturer and analyzed by conventional light microscopy. Cytokeratin-positive cells were counted in random areas and expressed as a percentage of the total number per microscopic field for each pregnant animal. Cytokeratin characterization was carried out after culture preparation of at least one coverslip from each placenta.
Phagocytic assays involving S. cerevisiae
The phagocytic assays were performed using S. cerevisiae (zymosan), either conjugated with Texas-red (Molecular Probes, Eugene, OR, USA) or unlabeled (Sigma) at a concentration of 7 x 105 particles/culture dish, for direct observation by fluorescence or light/electron microscopy respectively. In the experimental groups, IFN- (100 U/ml) was added to the culture medium 12 h before zymosan inoculation. To test the effect of the complement component C3 on phagocytosis by the placental trophoblast, yeast cells pre-treated with C3-sufficient mouse plasma (20 µl/ml), either alone or combined with IFN-, were also used as an additional treatment. Control cultures received either only zymosan particles or no treatment. To obtain opsonized-zymosan particles, zymosan was pretreated according to techniques described by Albieri and coworkers (1999). The final product formed C3b-bound zymosan (Lima & Diasda-Silva 1970) and was used in the phagocytic assays at the particle concentration mentioned above. Each experimental treatment was then analyzed 1, 2, 3, 6, 9 and 24 h after introduction of the yeast into the culture system, creating 18 experimental and 12 control groups. For each group, samples were assayed in triplicate.
The ability of placental cells to ingest zymosan particles was measured by counting the total number of cytokeratin-positive cells, as well as the total number of cytokeratin-positive cells containing internalized yeast cells, per microscopic field in a total of 10 random fields/culture dish. The phagocytic index (PI) was calculated as the percentage of trophoblast cells within the trophoblast population, which contained phagosomes (PI = number of trophoblast cells with zymosan-internalized particles x 100/total number of trophoblast cells). Counts were performed using a Nikon Optiphot light microscope or a Nikon eclipse E800 fluorescence microscope at a final magnification of 400 x . Results are given in the text as means±S.D. The mean values obtained for each experiment were compared with the control using ANOVA. Values of P < 0.05 were considered significant. For morphological and quantitative assays, the specimens incubated with Texas red-zymosan particles were fixed in 4% paraformaldehyde, counterstained with 4',6-diamidino-2-phenylindole (DAPI, Sigma) for observation by fluorescence microscopy or counterstained with 1% Toluidine blue for conventional light microscopy and mounted on histological slides with Diazabicyclo [2.2.2] octane (DABCO, Sigma). All specimens that received fluorescent particles (zymosan or E. coli ) were also submitted to concomitant cytokeratin immunodetection as previously described. Some cultures were also prepared for electron microscopy to accompany the stages of the zymosan internalization.
Ultrastructural studies
Cultured cells were gently scraped from the culture dish and inserted into 2% soft agarose (Gibco, USA) in PBS. After gelling, the agar was cut into small fragments and prepared as previously described for embedding in Spurr’s resin. Ultrathin (60-nm) sections were stained with 0.2% uranyl acetate and 0.5% lead citrate before examination in a JEOL CX-II-100 electron microscope.
Phagocytic assays using E. coli
To follow the progression of phagocytosis of bacterial cells by the placental trophoblast, bacteria expressing green fluorescent protein (GFP) were used (Ouahrani-Bettache et al. 1999). Since GFP absorbs light at a maximum excitation of 395 nm and fluoresces at a maximum emission of 510 nm, it permits the visualization of a single bacterium under a fluorescence microscope. Plasmid pRL1, which contains the GFP gene under the strong inducible promoter (Ptac) was constructed and introduced into E. coli. The transfected cells were grown overnight in LB medium supplemented with ampicillin and chloramphenicol. Addition of isopropylthio-ß galactosidase (IPTG) to the culture was not necessary because the bacteria expressed GFP even in the absence of the inducer, probably due to the natural leakiness of Ptac. Bacteria (1 x 105) were introduced into each of the following trophoblast cell cultures: treated with IFN- for 12 h, treated with C3-sufficient-plasma for 12 h, treated with IFN- and C3-sufficient-plasma for 12 h, and untreated. To obtain C3-sufficient plasma, mouse plasma was collected as described by Albieri and co-workers (1999) and added to the culture medium at a concentration of 10%, without previous heat-inactivation. At 1, 2, 3, 6, 9 and 24 h after the introduction of E. coli into the culture system, the cells were harvested, fixed in 4% paraformaldehyde and submitted to immunofluorescence procedures for cytokeratin A-B localization, mounted on histological slides with DABCO, and observed under a fluorescence microscope. Bacterial phagocytosis was observed either by the green fluorescent light emitted by the cloned GFP under a fluorescence microscope or with a JEOL CX-II-100 electron microscope as described above.
Construction of plasmid pRL1
Plasmid pRL1 was constructed by introducing Ptac into the polylinker of plasmid pBBR1-GFP (Ouahani-Bettache et al. 1999). Plasmid pBBR1-GFP carries a small polylinker followed by the promoterless GFP and chloramphenicol acetyl transferase (CAT) genes in tandem. Introduction of a promoter sequence at the polylinker site in the right orientation enhances the synthesis of both GFP and CAT proteins. A 270-bp DNA fragment containing Ptac was obtained by digesting plasmid pKK223-3 (Amersham) with BamHI (MBI-Fermentas). The Ptac was then ligated to the polylinker of plasmid pBBR1-GFP previously digested with BglII (MBI-Fermentas, Vilnius, Lithuania). The product of the ligation was electroporated into electrocompetent DH10B E. coli, which were plated on LB plates supplemented with 100 µg/ml ampicillin, 20 µg/ml chloramphenicol and 1 mM IPTG. The pRL1-transformed cells that expressed both CAT and GFP proteins were used in the phagocytic assays.
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Results |
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). Animals treated with thioglycolate presented leukocyte-attached particles and internalized yeast cells (Fig. 1b, inset). This occurred 3–6 h earlier in treated animals than in controls (data not shown). Yeast cells were also found in the spleen and in fetal and maternal components of the placenta. In the spleen, free and phagocytosed yeast cells were seen in both the thioglycolate-stimulated and non-stimulated experimental groups throughout the entire period analyzed (data not shown). From 6 h after inoculation, yeast cells seen in placental smears were apparently attached to leukocytes or trophoblast cells, or were still internalized in the cytoplasm of such cells (Fig. 1c, d). Paraplast placental sections (Fig. 1e, f) and ultrastructural analysis (Fig. 1g) also confirmed yeast access to the extra-vascular placental spaces and yeast internalization into the trophoblast cell cytoplasm.
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) and were morphologically distinct from the endodermal cells. Most of the negative cells presented a fibroblast typical morphology (14%) and the remaining cells (less than 1%) probably included decidual cells, endothelial cells, macrophages and uterine NK cells. In all experimental phagocytic assays, the phagocytosis exhibited by the placental cells was estimated in cells morphologically identified as trophoblast cells containing cytokeratin-positive filaments.
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) and giant cells (Fig. 2e, g). In general, non-giant cells displayed features similar to labyrinth zone cells, either organized in clusters, with cellular limits of difficult distinction and resembling a syncytial disposition (Fig. 2d,f), or as individual cells (Fig. 2e, g). However, the number of cells exhibiting phagocytic activity, as well as the number of phagosomes, varied markedly as a function of the experimental treatment, time of exposure to the phagocytic agent and type of particle (Figs 2–6).
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, inset). Addition of IFN- and/or C3-opsonized particles significantly altered the percentage of cells exhibiting phagocytic activity (Fig. 3c–h, Fig. 4). At the morphological level, it was also observed that the numbers of internalized particles per cell were altered (Fig. 3). At 6 and 24 h of treatment, C3-opsonized particles stimulated an approximate 1.5-fold increase in the phagocytic activity of the cultured cells (P < 0.05), whereas IFN- apparently led to earlier alterations (Fig. 4). At 2 h after the IFN- stimuli, there was a 2.7-fold increase (P < 0.05) in the phagocytic rate of the placental cells, followed by a significant increase of 1.3-fold at both 6 and 24 h of the treatment (P < 0.05). The concomitant presence of IFN- and C3 led to synergistic increases in phagocytosis rates during all experimental periods (Fig. 3g–h, Fig. 4), but the most pronounced effect occurred after 2 h of treatment (4.2-, 2.4- and 2.3-fold increases after 2, 6 and 24 h respectively, P < 0.05).
The patterns of adhesion and phagocytosis of E. coli and zymosan were similar, although a faster phagocytosis rate was observed for bacteria (Fig. 5). Internalized particles could be seen as soon as 1 h after introducing the bacteria to the culture. After 3 h, these particles became quite prominent in the trophoblast giant and non-giant cells (Fig. 5 a,b). At 6 h after bacterial inoculation, the trophoblast cytoplasm showed either scarce particles or diffusely fragmented fluorescent debris (Fig. 5 c,d). In addition to the cell morphology, the presence of cytokeratin filaments was used to verify the trophoblastic nature of the phagocytic cells (Fig. 5e–h). At the ultrastructural level, these temporal differences could also be seen (Fig. 6). At 6 h after bacteria inoculation, the cytoplasm of the trophoblast cells exhibited a large number of phagosomic vacuoles containing heterogeneous, unrecognizable material (Fig. 6 c,d), whereas recognizable particles still in the process of internalization were frequently seen in the samples receiving zymosan particles (Fig. 6 a,b).
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Discussion |
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Particles of S. cerevisiae (zymosan) were used for the in vivo experimental design because they are easily recognizable in light and electron microscopic analysis and are not capable of invading cells, thus assuring that the internalized particles result from phagocytosis. During the first 24 h following inoculation, yeast cells were found in the blood, spleens and placentas of the pregnant host animals. In those same animals, leukocytes, macrophages and trophoblast cells showed conspicuous signs of phagocytosis, with or without thioglycolate activation of macrophages. At the placental level, the onset of yeast cell phagocytosis by leukocytes and trophoblast cells began as early as 3 h after inoculation. The yeast cell phagocytosis was faster than that described for pathogenic bacteria such as Coxiella burnetti, Chlamydia psittaci, Listeria monocytogenes and Brucella abortus (Redline & Lu 1988, Baumgartner & Bachman 1992, Tobias et al. 1993, Buendia et al. 1998). It is possible that these microorganisms have defense mechanisms for avoiding immediate recognition by the immunological defense system of the host organism. In the case of S. cerevisiae in particular, this is an unexpected finding.
Treatment with thioglycolate did not significantly alter the pattern of yeast phagocytosis by either leukocytes and/or placental cells, but did appear to interfere with the kinetics of the process. Phagocytosis was triggered earlier in the thioglycolate-treated animals, suggesting that macrophages may, either directly or indirectly, orchestrate the phagocytic process in other cell types, including placental cells. This may be particularly significant in the case of trophoblast cells because it is indicative of a maternal–fetal dialogue through regulatory molecules of the maternal defense system. Direct stimulation of the trophoblast by thioglycolate might also be involved in this activation. However, in our in vitro studies (data not shown), we found no evidence of such a mechanism.
Using primary placental cell cultures, we also identified and estimated the proportion of trophoblast cells involved in phagocytosis under different situations. Cytokeratin A-B (CK) immunolocalization (Brület et al. 1980) and morphological analysis were employed in order to verify the lineage of the phagocytic cells. Although phagocytosis was immediately observed when yeast cells or E. coli were added to the system, E. coli particles were apparently internalized more rapidly than were zymosan particles. Phagocytosis of E. coli peaked 3 h after the bacteria were introduced into the culture system, whereas maximum yeast phagocytosis was achieved at approximately 6 h after culture inoculation. The reasons for these differences are unclear, but may involve recognition, attachment and mechanisms for internalization and degradation – all dependent upon the nature of the target organisms. In this model system, trophoblast giant cells (giant, CK+ individual cells) and trophoblast cells that are morphologically compatible with those present in the labyrinth zone (small, clustered CK+ cells) exhibit phagocytosis. In normal gestation, labyrinth cells are usually involved in metabolic exchange rather than in phagocytic activity. However, Athanassakis et al.(1987) reported some phagocytic activity in the labyrinth, suggesting that these cells may be endowed with this ability as an additional nutritional mechanism.
The ability of trophoblast cells to phagocytose was also tested by using phagocytic mediators such as IFN- and complement component C3. Results of morphological and quantitative analyses suggest that both treatments enhanced the phagocytic rate, indicating an increase in the number of cells exhibiting microorganisms within phagosomes, as well as in the number of phagosomes per cell. In comparison with IFN-, C3 protein appeared to stimulate a greater number of cells to phagocytose. However, the highest phagocytic levels were obtained using both treatments concomitantly, suggesting that their effects are synergistic.
The IFN--mediated phagocytic response observed here might be a direct consequence of IFN- receptor and ß chains up-regulation in the function of the IFN- treatment. Similar up-regulation has been described in macrophages (Mantovani, 1981 , 1987, Hiraí et al. 1985, Heyworth & Badwey 1990, Lewis & McGee 1992) and in early stages of gestation (Albieri et al. 1999, 2001). In our laboratory, studies investigating the possibility that up-regulation may also occur in placental trophoblast cells are currently underway.
Involvement of the C3 protein in the phagocytic process of trophoblast cells is notable, but not unique. In a receptor-specific mediated process, C3b-opsonized particles have been shown to be more susceptible to recognition, adhesion and phagocytosis (Bianco et al. 1975, Mantovani 1987). In the present study, we were also able clearly to associate the presence of C3-opsonized particles with increased numbers of internalized particles in the trophoblast cell population, implying that these molecules facilitate trophoblast cell phagocytosis in a similar process. Although we did not analyze expression of C3b receptor in placental cells, this receptor has been detected at the trophoblast giant-cell surface in the earlier stages of gestation (Albieri et al. 1999). This and other evidence in the literature (Athanassakis et al. 1987, Drake & Rodger 1987, Guilbert et al. 1993, Albieri et al. 1999 , 2001, Guleria & Pollard 2000, Sasmono et al. 2003) strengthens the argument that trophoblast cells play roles in placental defense mechanisms. In mice, CSF-1 of maternal origin interacts with trophoblast cells, inducing the synthesis of neutrophil chemoattractants and macrophage inflammatory protein-2, both of which are potent mediators of the maternal immune response against bacterial infection (Guleria & Pollard 2000). Moreover, T-lymphocytes act on trophoblast cells, stimulating them to proliferate and to exhibit phagocytic behavior (Athanassakis et al. 1987).
Taken together, these data clearly indicate that placental trophoblast cells are not only able to express phagocytosis in vivo and in vitro but, when properly stimulated via various activation pathways, are also capable of increasing phagocytic activity. Our results also corroborate other findings that suggest a trophoblast defense mechanism (Athanassakis et al. 1987, Drake & Rodger 1987, Guilbert et al. 1993, Albieri et al. 1999, 2001, Guleria & Pollard 2000). We conclude that these cells are capable of selectively removing microorganisms from the maternal–fetal interface and in a culture environment.
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Footnotes |
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References |
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Albieri A & Bevilacqua E 1996 Induction of erythrophagocytic activity in cultured mouse trophoblast cells by phorbol myristate acetate and all-transretinal. Placenta 17 507–512.[CrossRef][Web of Science][Medline]
Albieri A, Kipnis T & Bevilacqua E 1999 Possible role of activated complement component C3 in phagocytic activity exhibited by the mouse trophoblast. American Journal of Reproductive Immunology 41 343–352.
Albieri A, Amarante AM, Gagioti S & Bevilacqua E 2001 In vitro behavior of post implanting and placental trophoblast cells during zymosan challenging and interferon- stimulation. Placenta 22 A11.
Athanassakis I, Bleackley RC, Patetkau V, Guilbert L, Barr PJ & Wegmann TG 1987 The immunostimulatory effect of T cells and T cell lymphokines on murine fetally derived placental cells. Journal of Immunology 138 37–44.[Abstract]
Athanassakis I, Chaouat G & Wegmann TG 1990 The effects of anti-CD4 and CD-8 antibody treatment on placental growth and function in allogeneic and syngeneic murine pregnancy. Cellular Immunology 129 13–21.[CrossRef][Web of Science][Medline]
Athanassakis I, Papamatheakis J, Bouris G & Vassiliadis S 2000 Interferon-gamma induces differentiation of ectoplacental cone cells to phenotypically distinct trophoblasts. Developmental Comparative Immunology 24 633–672.
Athanassakis-Vassiliadis I, Papamatheakis J & Vassiliadis S 1993 Specific CSF-1 binding on murine placental trophoblasts and macrophages serves as a link to placental growth. Journal of Receptor Research 13 739–751.[Web of Science][Medline]
Baumgartner W & Bachman N 1992 Histological and immunocytochemical characterization of Coxiella burnet–associated lesions in the murine uterus and placenta. Infection and Immunity 60 5232–5241.
Bevilacqua E & Abrahamsohn PA 1988 Ultrastructure of trophoblast giant cell transformation during the invasive stage of implantation of the mouse embryo. Journal of Morphology 198 341–351.[CrossRef][Web of Science][Medline]
Bevilacqua E & Abrahamsohn PA 1989 Trophoblast invasion during implantation of the mouse embryo. Archivos de Biología y Medicina Experimentales 22 107–108.[Medline]
Bianco C, Griffi FM Jr & Silverstein C 1975 Studies of the macrophage complement receptor: alteration of receptor function upon macrophage activation. Journal of Experimental Medicine 141 1278–1290.
Billington WD 1971 Biology of the trophoblast. In Advances in Reproductive Physiology, pp 27–66. Ed. MNH Bishop. London: Logos Press.
Brület P, Babinet C, Kemler R & Jacob F 1980 Monoclonal antibodies against trophectoderm-specific markers during mouse blastocyst formation. PNAS 77 4113–4117.
Buendia AJ, Sanchez J, Martinez M, Camara P, Navarro JA, Rodolakis A & Salinas J 1998 Kinetics of infection and effects on placental cell populations in a murine model of Chlamydia psittaci-induced abortion. Infection and Immunity 66 2128–2134.
Call DR, Nemzek JK, Ebong SJ, Bolgos GL, Newcomb DE & Remick DG 2001 Ratio of local to systemic chemokine concentrations regulates neutrophil recruitment. American Journal of Pathology 158 715–721.
Clark DA, Yu G, Arck PC & Gorezynski RM 2003 MD-1 is a critical part of the mechanism causing TH-1 cytokine-triggered murine fetal loss syndrome. American Journal of Reproductive Immunology 49 297–307.[CrossRef]
Cohen MS, Ryan JL & Root RK 1981 The oxidative metabolism of thioglycolate-elicited macrophages: the relationship between oxygen, superoxide and hydrogen peroxide and the effect of monolayer formation. Journal of Immunology 127 1007–1011.[Abstract]
Cohn ZA 1978 The activation of mononuclear phagocytes: fact, fancy and future. Journal of Immunology 121 813–816.
Drake BL & Rodger JC 1987 Phagocytic properties of cultured murine trophoblast. Placenta 8 129–134.[Web of Science][Medline]
Enders AC & Schlafke S 1969 Cytological aspects of trophoblast uterine interactions in early implantation. American Journal of Anatomy 125 1–29.
Gagioti S, Colepicolo P & Bevilacqua E 1995 Post implantation mouse embryos have the capability to generate and release reactive oxygen species. Reproduction, Fertility and Development 7 1111–1116.[CrossRef][Medline]
Gagioti S, Colepicolo P & Bevilacqua E 1996 Reactive oxygen species and the phagocytosis process of hemochorial trophoblast. Ciência e Cultura 48 37–42.
Gagioti S, Scavone C & Bevilacqua E 2000 Participation of the mouse implanting trophoblast in nitric oxide production during pregnancy. Biology of Reproduction 62 260–268.
Guilbert L, Robertson SA & Wegmann TG 1993 The trophoblast as an integral component of macrophage–cytokine network. Immunology Cell Biology 71 49–57.[CrossRef]
Guleria I & Pollard JW 2000 The trophoblast is a component of the innate immunosystem during pregnancy. Nature Medicine 6 589–593.[CrossRef][Web of Science][Medline]
Heyworth PG & Badwey JA 1990 Protein phosphorylation associated with the simulation of neutrophils. Modulation of superoxide production by protein kinase C and calcium. Journal of Bioenergetics and Biomembranes 22 1–26.[CrossRef][Web of Science][Medline]
Hiraí K, Ohno Y & Ogawa K 1985 Cytochemistry of hydrogen peroxide production by cultured activated macrophages in phagocytosis. Acta Histochemica et Cytochemica 18 69–75.
Kanai-Azuma M, Kanai Y, Kurohmaru M, Tachi C, Yazaki K & Hayashi Y 1994 Giant cell transformation of trophoblast cells in mice. Endocrine Journal 41 (Suppl) 33–41.
Kohchi C, Mizuno D & Soma G 1991 Expression of TNF-alpha and TNF-beta transcripts in murine embryonal carcinoma cells and trophoblast cell. Nucleic Acids Symposium Series 25 119–120.
Lewis C & McGee JOD 1992 The Macrophage. Oxford: IRL Press.
Lima AO & Diasda-Silva W 1970 Immunologia, Imunopatologia, Alergia - Métodos. Rio de Janeiro: Guanabara Koogan.
Mantovani B 1981 Phagocytosis of immune complex mediated by IgM and C3b by macrophages from mice treated with glycogen. Journal of Immunology 126 127–130.[Abstract]
Mantovani B 1987 Phagocytosis of in vitro-aged erythrocytes: a sharp distinction between activated and normal macrophage. Experimental Cell Research 173 282–288.[CrossRef][Web of Science][Medline]
Mehrotra PK 1988 Ultrastructure of mouse ectoplacental cone cells. Biological Structures and Morphogenesis 1 63–68.[Medline]
Muntener M & Hsu YC 1977 Development of trophoblast and placenta in the mouse. Acta Anatomica 98 241–252.[Web of Science][Medline]
Ouahrani-Bettache S, Porte F, Teyssier J, Liautard JP & Kohler S 1999 pBBR1-GFP: a broad-host-range vector for prokaryotic promoter studies. Biotechniques 26 620–622.[Web of Science][Medline]
Paulesu L, King A, Loke YW, Cintorino M, Bellizzi E & Boraschi D 1991 lmmunohistochemical localization of IL-1 beta in normal human placenta. Lymphokine Cytokine Research 10 443–448.
Pavia CS 1983 Expression of cell-mediated antimicrobial immunity by mouse trophoblast monolayers. Journal of Infection Diseases 146 1006–1010.
Redline RW & Lu CY 1988 Specific defects in the anti-lysterial immune response in discrete regions of the murine uterus and placenta account for susceptibility to infection. Journal of Immunology 140 3947–3955.[Abstract]
Salins S, Newlon C, Winden P, Klein TW & Friedman H 2001 Differential induction of gamma interferon in Legionella pneumophila infected macrophages from Balb/c and A/J mice. Infection and Immunity 69 3605–3610.
Sasmono RT, Oceandy D, Pollard JW, Tong W, Pavli P, Wainwright BJ, Ostrowski MC, Himes SR & Hume DA 2003 A macrophage colony-stimulating factor receptor green fluorescent protein transgene is expressed throughout the mononuclear phagocyte system of the mouse. Blood 101 1155–1163.
Schaffer I, Scheid A, Spielmann P, Breymann C, Zimmermann R, Meudi M, Gassmann M, Marti HH & Wenger RH 2003 Oxygen-regulated expression of TGF-beta 3, a growth factor involved in trophoblast differentiation. Placenta 24 941–950.[CrossRef][Web of Science][Medline]
Sharma RK 1998 Mouse trophoblast cell lines: I - relationship between invasive potential and TGF-beta 1. In Vivo 12 431–440.[Web of Science][Medline]
Simmons RL & Russell PS 1962 The antigenicity of mouse trophoblast. Annals of New York Academy of Sciences 99 717–732.
Soares MJ & Glasser SR 1987 Placental lactogen production and functional differentiation of rat trophoblast cells in vitro. Journal of Reproduction and Fertility 79 335–341.
Soares MJ, Faria TN, Hamilin GP, Lu XJ & Deb S 1993 Trophoblast cell differentiation: expression of the placental prolactin family. In Trophoblast Cells: Pathways for Maternal–Embryonic Communications, pp 45–67. Eds MJ Soares, S Handwerger & F Talamares. New York: Springer-Verlag.
Soares MJ, Chapman BM, Rasmussen CA, Tamei T & Orwing KE 1996 Differentiation of trophoblast endocrine cells. Placenta 17 277–289.[CrossRef][Web of Science][Medline]
Tobias L, Cordes DO & Schuring GG 1993 Placental pathology of the pregnant mouse inoculated with Brucella abortus strain 2308. Veterinary Pathology 30 119–129.[Abstract]
Welsh AO & Enders AC 1987 Trophoblast–decidual cell interaction and establishment of maternal blood circulation in the parietal yolk sac placenta of the rat. Anatomical Record 217 203–219.[CrossRef][Medline]
Whiteside FJ, Boucaut KJ, Teh A, Garcia-Aragon J, Harvey MB & Herington AC 2003 Elevated concentration of TNF-alpha induces trophoblast differentiation in mouse blastocyst outgrowths. Cell and Tissue Research 314 275–280.[CrossRef][Web of Science][Medline]
Yamaguchi M, Sawada K & Miyake A 1996 Lipopolysaccharides selectively inhibit mouse placental lactogen-II secretion through stimulation of interleukin-1 alpha (IL-1 alpha) and IL-6 production. Journal of Endocrinological Investigation 7 415–421.
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