Two patterns of cytokine production by placental macrophages.

INTRODUCTION: Macrophages participate in the regulation immune and morphogenetic events in the placenta. However, these roles remain unclear for placental macrophages (Hofbauer cells). The aims of this study were to characterize the consecutive steps of cytokine production (intracellular synthesis and secretion) in placental macrophages in early and late gestation and to compare the secretory profiles of placental macrophages and villous tissue. METHODS: Macrophages and villous tissue were isolated from placentas obtained from normal pregnancies at either 9-12 or 38-40 weeks of gestation. Intracellular cytokines were determined by flow cytometry after staining with monoclonal antibodies. Secreted cytokines were quantified by cytometric bead array and ELISA. RESULTS: Two patterns of cytokine production were revealed in placental macrophages. Cytokines in the first group (IL-1, IL-6, IL-8, IL-10, TNFα) demonstrated low basal production and were stimulated by bacterial endotoxin. Cytokines in the second group (IL-11, IL-17A, IL-17F, TGF-ß, VEGF) were characterized by constitutive production and did not respond to stimulation. Gestational age-dependent changes were observed: basal secretion of TNFα and IL-8 increased whereas IL-11 and IL-17 secretion decreased in third-trimester macrophages compared with the first-trimester cells. Comparison of cytokine production at the cellular and tissue levels suggested the contribution of the placental macrophages both in intraplacental and extraplacental cytokine production. DISCUSSION: It would be safe to assume that the two patterns of cytokine production, revealed in our study, correspond to two regulatory roles of placental macrophages: "immune" and "morphogenetic". The inflammatory phenotype of macrophages is attenuated in early gestation and increases with the progression of pregnancy. The cytokines of the first group supposedly contribute to both local and extraplacental levels, whereas the cytokine effects of the second group are more likely confined to the placental tissue.

Coordinated expression of TNFα- and VEGF-mediated signaling components by placental macrophages in early and late pregnancy.

INTRODUCTION: Mononuclear phagocytes are thought to significantly contribute to cytokine regulation at the maternal-foetal interface, but the role of placental macrophages has been poorly investigated. TNFα and VEGF were demonstrated to have regulatory effects on basic structures of the placenta, particularly the trophoblast and blood vessels. The aims of this study were to determine the expression of TNFα, VEGF and related receptors in placental macrophages, and how does the participation of placental macrophages alter with gestational age in TNFα- and VEGF-mediated signaling. METHODS: Macrophages were isolated from placental villous tissue from normal pregnancies at either 9-12 or 38-40 weeks gestation. Cell surface receptors (TNFR1, TNFR2, VEGFR1, and VEGFR2) and intracellular TNFα and VEGF were quantified by flow cytometry after antibody staining. Basal and stimulated secretion of both cytokines and soluble TNF receptors was quantified by cytometric bead arrays. Secreted VEGFR1 was measured by ELISA. RESULTS: The expression of TNFR1 and VEGFR1 was remarkably variable and did not change from first to third trimester. There was minimal basal TNFα production in the placental macrophages, but nearly all cells in the population produced VEGF. TNFα and VEGF secretion increased with gestational age accompanied by decreased secretion of the antagonists sTNFR1 and sVEGFR. Macrophages isolated from early term placentas were less effective in responding to bacterial endotoxin. Lipopolysaccharide induced increases in the secretion of TNFα, TNFR1, TNFR2, and VEGFR1 but did not affect the production of VEGF. In late pregnancy, a significant correlation was observed between TNFR1 and VEGFR1. DISCUSSION: The progression of pregnancy is accompanied by the concerted increase in TNFα and VEGF secretion and decrease in the production of their soluble receptors, but the expression of cell surface receptors does not depend on gestational age. The observed patterns of basal and stimulated expression of TNFα and VEGF may reflect the dual immune and morphogenetic roles of placental macrophages in gestation. Compatible patterns of TNFR1 and VEGFR1 expression suggest common regulatory pathways for these receptors.

Comparative phenotypic characterization of human cord blood monocytes and placental macrophages at term.

The expression of surface molecules in cord blood monocytes and placental macrophages was studied using flow cytometry. When compared with monocytes, macrophages presented a decrease in HLA-DR and LAP/TGF-ß1 levels and increased expression of alternative activation markers, especially CD206. No difference in the production of the apoptotic factors TRAIL and TWEAK was observed, whereas the levels of cytokine receptors in monocytes were significantly higher than in macrophages. Most remarkable was the difference in the expression of IL-17 and TNFα receptors. A strong correlation between VEGF and TNFα receptors was revealed in both cell populations. The results obtained in this study provide antigenic phenotypes for two related cell populations and outline the feasible functional alterations during tissue macrophage differentiation.

In vitro expression of vascular endothelial growth factor and its receptors by placental macrophages.

The expression of VEGF and membrane-bound and soluble forms of the VEGF-R1 receptor in cultured placental macrophages (trimesters I and III of pregnancy) was studied by flow cytometry, cytometric bead array, and ELISA. Nearly all population of placental macrophages (98%) was capable of producing VEGF during the early and late gestational periods. However, the expression of cellular VEGF-R1 varied from 3.4 to 92%. VEGF secretion was relatively low in the first and third trimesters (0.5 and 1.1 pg/10(5) cells, respectively). Cultured placental macrophages produced soluble receptor sVEGF-R1 in the first and third trimesters (86.4 and 36.4 pg/10(5) cells, respectively). Stimulation with LPS was followed by a 4-fold increase in sVEGF-R1 secretion. Our results indicate that placental macrophages are involved in the autocrine and paracrine regulation in chorionic villi. The data suggest that these cells have a physiological and pathogenetic role in gestation.

Spontaneous and LPS-induced secretion of cytokines by villous chorion tissue.

Secretion of IL-1alpha, IL-1beta, IL-6, IL-10, and TNF-alpha cytokines by villous chorion cultures (7-14 weeks) during normal pregnancy and in spontaneous abortions was studied. Secretion of IL-1alpha and IL-6 increased 4.5 and 7.3 times in miscarriages, while secretion of IL-1beta, IL-10, and TNF-alpha decreased. LPS stimulated the production of IL-1alpha and IL-6 in samples obtained during surgical abortion. LPS stimulated IL-1alpha and TNF-alpha secretion in miscarriages, while the level of IL-6 production decreased significantly. It is hypothesized that increased production of IL-1alpha and IL-6 and attenuation of the antiinflammatory effect of IL-10 play an important role in the pathogenesis of miscarriages at early stages of gestation. The results suggest that cytokine regulation of the fetus rejection is different at early and late stages of gestation.

Local production of interleukins and growth factors in external genital endometriosis.

Secretion of some IL and growth factors (VEGF, IGF-I, TGFbeta) by endometrial tissues and endometrioid heterotopies was studied in vitro in patients with external genital endometriosis of different severity. The production of IL-1beta, IL-2, IL-6, and VEGF in the endometrium increased in severe external genital endometriosis, while the secretion of TGFbeta decreased; hyperproduction of IL-2, IL-6, VEGF and decreased production of TGFbeta were detected in endometrioid foci. Presumably, local cytokine imbalance and increased proliferative activity of endometrial cells are involved in the mechanisms of formation and functioning of endometrioid foci.

Study of cytokine profile and angiogenic potential of peritoneal fluid in patients with external genital endometriosis.

The concentrations of proinflammatory cytokines (IL-1beta, IL-6), vascular endothelium growth factor, tumor growth factor-beta, and insulin-like growth factor-1 were measured in the peritoneal fluid of patients with external genital endometriosis and healthy women by enzyme immunoassay. The effect of peritoneal fluid from patients with external genital endometriosis on proliferative activity of EA.Hy926 human endothelial cells was evaluated by the method based on the analysis of cell cycle by flow cytometry. The concentrations of IL-1beta, IL-6, and insulin-like growth factor-1 were increased in patients with endometriosis in comparison with healthy women. The peritoneal fluid from patients with endometriosis (but not from healthy women) significantly increased mitotic activity of endothelial cells and exhibited high angiogenic potential, which can promote implantation and growth of endometrial transplants. Presumably, insulin-like growth factor-1 stimulates this process.

[On the influence of silicate feeding on metabolism of 137Cs in rabbit]. / O vliianii kormovogo ispol'zovaniia silikatov na metabolizm 137Cs v organizme krolikov.

The features of 137Cs metabolism in the rabbit's organism when feeding on silicates were investigated. By results of two experiments on rabbits radiosorption by of zeolite, saponit, humolit, vermiculite and palygorskit in the gastrointestinal tract was evaluated. It was found that vermiculite and palygorskit had the strongest influence on processes of accumulation and decorporation 137Cs, when a dose of additives was 5% of the weight of forage.

Secretion of proinflammatory cytokines by villous chorion tissue in spontaneous abortion.

Secretion of proinflammatory cytokines IL-1 beta, TNF-alpha, IL-6, IFN-alpha, and IFN-gamma by the chorion tissue (8-12 weeks) was studied in normal gestation and spontaneous abortion. The production of IL-1 beta, TNF-alpha, and IFN-alpha virtually did not change in spontaneous abortion, while IFN-gamma was not secreted in all experimental groups. The production of IL-6 increased more than 2-fold in patients with spontaneous abortion during the first trimester. These data confirm the involvement of this cytokine in the reproductive processes and in the pathogenesis of miscarriage.

Cytokines and placental macrophages in regulation of birth activity.

The article reviews present notions on functional activity of cytokines of the fetoplacental complex. Particular emphasis is placed on the role of these molecules in the regulation of gestation processes and in pregnancy incompetence. The mechanism of the involvement of placental macrophages and their products in gestation and delivery is discussed.

Structural alterations of human immortal astrocytes after inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase by lovastatin.

This investigation was designed to study the specific structural alterations of astroglial cells following the inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, which is a key enzyme in cholesterol synthesis. In our previous investigations, it has been demonstrated that lovastatin, which is a specific competitive inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, effectively inhibited cholesterol synthesis in human primary and immortal astrocytes in serum and lipid-free media, and we also showed that the effects on both astrocytes of primary cultures and immortal astrocytes (ASCh-7) were very similar. In the present study we therefore examined effects of lovastatin at a concentration of 100 ng/ml on human immortal astrocytes (ASCh-7) using electron microscopic analysis. We have found that lovastatin significantly affects human immortal astrocytes, resulting in degenerative ultrastructural changes including accumulation of a large number of phagosomes, oedema of the cytoplasm and destruction of gliofilaments. The results obtained suggest that the human immortal astrocyte cell line (ASCh-7) can be not only very useful for screening drugs in pharmacological research but can also provide a useful model for examining fine structural alterations of cells following selective disturbances of metabolism.

An in vitro study of the effects of lovastatin on human fetal brain cells.

We used various cultures of embryonic brain cells and a line of immortal astrocytes as in vitro model systems to study the direct effects of the hypolipidemic drug lovastatin on developing human CNS cells. Our data showed that pharmacological concentrations of the drug significantly affected growth and development of neuronal and astroglial cells in serum- and lipid-free media. Lovastatin at concentrations of 0.01-1000 ng/ml effectively inhibited intracellular cholesterol synthesis in primary and immortal astrocytes as well as in glial-neuronal reaggregated cultures. Primary astrocytes were more sensitive to minimal concentrations of the drug than their immortal counterparts and glial-neuronal aggregates. A concentration of 100 ng/ml of lovastatin significantly increased activity of LDL receptors both in primary and immortal astrocytes by about 100% and 50%, respectively (p < 0.001 and p < 0.01, respectively). Proliferation of immortal astrocytes in serum-free medium was entirely inhibited by 100 ng/ml of lovastatin. By contrast, a concentration of 5 ng/ml of lovastatin had no significant effect on cell proliferation. Long-term exposure of human brain explants to 100 ng/ml of lovastatin resulted in detrimental ultrastructural changes in neuronal and glial cells and led to cell death. Our data suggest that lovastatin is neurotoxic to developing brain cells and we propose that its in vivo adverse effects on the CNS may be attributed, at least in part, to its direct influence on human neurons and astrocytes as observed in vitro.

[The production of "pure" cultures of neurons from the brain of human embryos]. / Poluchenie "chistykh" kul'tur neironov golovnogo mozga émbrionov cheloveka.

A method has been developed for preparing essentially pure primary cultures of neurons from 10-12 week old fetal human brain. This method is based on the 1) capacity of neurons to form multicellular aggregates, 2) cultivation in chemically defined medium, and 3) treatment with cytosine arabinoside as antimitotic agent. This procedure allowed the preparation of highly purified (95 per cent and more) neuronal aggregate cultures. Ultrastructural analysis of these cultures following one and 8 days has revealed their partial differentiation during cultivation.

[An electron microscopic study of neuronal development in organotypic cultures of the anlage of the cerebral cortex from a human embryo]. / Elektronno-mikroskopicheskoe issledovanie razvitiia neironov v organotipicheskikh kul'turakh zakladki kory golovnogo mozga émbriona cheloveka.

Data are provided on cytodifferentiation of the cerebral cortex cultured cells taken from 10-12 week old embryos of man. It is shown that low differentiated neuroblasts well survive in culture for 21 days. Mature granular cells and middle pyramidal neurons are revealed in cultures. The number of morphological criteria may testify to the maturity of neurons: the presence of the Nissl substance, differentiation of dendrites and axons; the presence of various types of synapses. The absence of myelinized fibres testifies to the insufficient maturity of the cultures, that is probably associated with employing the low differentiated nervous tissue for cultivation and with insufficient cultivation period.