Enrique G. Olivares

Follicular Dendritic Cells Are Related to Bone Marrow Stromal Cell Progenitors and to Myofibroblasts

Follicular dendritic cells (FDC) are involved in the presentation of native Ags to B cells during the secondary immune response. Some authors consider FDC to be hemopoietic cells, whereas others believe them to be mesenchymal cells. The low proportion of FDC in the lymphoid follicle, together with technical difficulties in their isolation, make these cells difficult to study. We show that Fibroblast Medium can be used successfully to isolate and maintain FDC lines. In this culture medium, we obtained 18 FDC lines from human tonsils, which proliferated for as long as 18 wk and showed a stable Ag phenotype as detected by flow cytometry and RT-PCR. FDC lines were CD45-negative and expressed Ags associated to FDC (CD21, CD23, CD35, CD40, CD73, BAFF, ICAM-1, and VCAM-1) and Ags specific for FDC (DRC-1, CNA.42, and HJ2). These cell lines were also able to bind B cells and secrete CXCL13, functional activities characteristic of FDC. Nevertheless, the additional expression of STRO-1, together with CD10, CD13, CD29, CD34, CD63, CD73, CD90, ICAM-1, VCAM-1, HLA-DR, alkaline phosphatase, and α-smooth muscle actin (α-SM actin) indicated that FDC are closely related to bone marrow stromal cell progenitors. The expression of α-SM actin also relates FDC with myofibroblasts. Like myofibroblasts, FDC lines expressed stress fibers containing α-SM actin and were able to contract collagen gels under the effect of TGFβ1 and platelet-derived growth factor. These findings suggest that FDC are a specialized form of myofibroblast and derive from bone marrow stromal cell progenitors.

Immune phenotype and cytotoxic activity of lymphocytes from human term decidua against trophoblast

Flow cytometric data were used to compare the phenotype of term decidual lymphocytes and peripheral blood lymphocytes. Unlike peripheral blood lymphocytes, a significant percentage of CD3+, CD4+, CD8+ and CD16+ term decidual lymphocyte populations expressed the CD69 activation marker. The relative proportions of CD38 in CD3+, CD4+ and CD8+ populations were more than twice as large in term decidual lymphocytes as in peripheral blood lymphocytes. As reported for early decidual lymphocytes, the expression of CD38 and CD69 by term decidual lymphocytes suggests that these cells are also regionally activated. However, term decidual lymphocytes showed no spontaneous cytotoxicity against normal trophoblast or its tumoral counterpart, JEG cells. After stimulation with interleukin-2, these lymphocytes became cytotoxic, as did peripheral blood lymphocytes. The relevance of this latter result to the immune control of the physiological and pathological invasion of the decidua by the trophoblast is discussed.

Antigenic Phenotype of Cultured Human Osteoblast-Like Cells

Background/Aims: Osteoblasts are classically considered to play an important role during bone tissue development, and to be involved in the formation of mineralized bone matrix. Recent reports have suggested that they can also exert some activities directly associated with the immune system (cytokine synthesis and antigen presentation). Moreover, some authors have found antigens on osteoblast-like cells normally expressed by other cells with a common origin in bone marrow.Methods: We isolated and cultured human osteoblast-like lines and studied their antigenic phenotype with flow cytometry using monoclonal antibodies against antigens associated with hematopoietic cells.Results: Cultured cells expressed CD34, but were negative for CD45. B cell antigens CD20 and CD23 and myelomonocytic antigens CD11b, CD13, and CD16 were detected. Expression of CD3, CD14, CD15 and CD68 was negative, whereas CD25 expression was positive. CD56, an antigen expressed on NK cells, was positive. These cells were CD10, CD44, CD54, CD80, CD86 and HLA-DR positive, as previously described. An antigen specific to follicular dendritic cells was also observed on cultured osteoblast-like cells.Conclusions: The antigenic phenotypes of human osteoblast-like cells and FDC are similar. These data suggest that osteoblasts may be functionally related to certain dendritic cells and may play an additional role in bone tissue to that classically assigned.

Constitutive Secretion of Interleukin-6 by Human Decidual Stromal Cells in Culture. Regulatory Effect of Progesterone

PROBLEM: Although several studies have demonstrated that decidual stromal cells (DSC) can secrete cytokines in culture, none of these studies documented the purity of the cultures. Since other cells of the decidua, such as macrophages and epithelial cells, also produce cytokines, it is important to ensure purity of the culture so that cytokine production can be ascribed with confidence to DSC.

Decidual Lymphocytes of Human Spontaneous Abortions Induce Apoptosis but Not Necrosis in JEG-3 Extravillous Trophoblast Cells

Abstract The human decidua contains an unusually high proportion of lymphocytes, mainly NK and T cells, which are potentially cytotoxic to the trophoblast when they are stimulated with certain cytokines. Given the high incidence of spontaneous abortion in humans and other species, our working hypothesis is that decidual lymphocytes are involved in immunological mechanisms that attack the trophoblast and induce abortion when any gestational problem arises. To test this hypothesis, flow cytometry was used to compare decidual lymphocyte populations in first-trimester spontaneous abortions and elective terminations of first-trimester pregnancy. We found significantly higher proportions of decidual lymphocytes that expressed activation markers, and of T cells (mainly T helper cells) in spontaneous abortions than in elective terminations of pregnancy. Decidual lymphocytes from spontaneous abortion, like decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, were however, unable to lyse the JEG-3 extravillous cytotrophoblast cell line in a 51Cr-release assay. Nevertheless, decidual lymphocytes from spontaneous abortion, unlike decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, induced apoptosis in JEG-3 cells as determined by DNA fragment-release assay. Hematoxylin and eosin staining showed a significantly higher proportion of apoptotic JEG-3 cells when these cells were treated with decidual lymphocytes from spontaneous abortion than when JEG-3 cells were cultured with decidual lymphocytes from elective termination of pregnancy. The ultrastructural signs of apoptosis were confirmed by electron microscopy. These data support the hypothesis that activated decidual lymphocytes participate in human spontaneous abortion by inducing apoptosis but not necrosis of the trophoblast.

Immunospecificity of albumin detected in 1.6 million-year-old fossils from Venta Micena in Orce, Granada, Spain

The Orce skull fragment from southern Spain, dated at 1.6 Myr, has been a subject of heated controversy since it was first discovered in 1982. If it is hominid, as its discoverers contend, it is by far the oldest fossil hominid yet found in western Europe and implies that human populations settled this region much earlier than was previously realized. Numerous stone artifacts found at the Orce sites provide evidence that hominids were indeed present there in the Lower Pleistocene. Some paleontologists maintain that the 8 cm diameter occipital fragment is from a horse, not a hominid. Two independent investigations of the residual proteins in the skull were undertaken, one at the University of Granada in Spain, the other at the University of California, San Francisco. Two immunological methods of comparable sensitivity were employed for detection and species attribution of protein extracted from fossil bone: the Granada team used an enzyme-linked-immunosorbent assay (ELISA), and the UCSF team used a radioimmunoassay (RIA). Both teams obtained reactions characteristic of human albumin in the Orce skull and horse albumin in some of the horse fossils. These results support the lithic evidence that hominids were living in Andalusia 1.6 million years ago.

Antigen phenotype of cultured decidual stromal cells of human term decidua

We previously reported that decidual stromal cells (DSC) from early human decidua express antigens associated with hematopoietic cells and develop different immune functions. Here we study the antigenic phenotype of DSC from term decidua and compare it with the phenotype reported for DSC from early decidua. Decidual stromal cells were isolated from human term deciduas and maintained in culture until highly purified DSC cultures were obtained. Most term DSC, like most early DSC, expressed CD10. Term DSC expressed antigens specific for follicular dendritic cells (FDC), such as DRC-1 (CD21L) and HJ2, together with CD21, CD23 and CD80, which are detected on FDC as well. Also like early DSC, term DSC were negative for CD3, CD14, CD15 and CD45. Although early DSC were reported to be HLA-DR-positive and CD86-positive, these antigens were not expressed by term DSC. These discrepant results suggest that two types of cells, or cells at different stages of differentiation (decidualization) were selected during culture of decidual cells from different periods of gestation. This possibility was further supported by the finding that term DSC expressed desmin and prolactin, two markers of decidualization, whereas these molecules have not previously been detected in early DSC.

Phagocytosis by fresh and cultured human decidual stromal cells: opposite effects of interleukin-1α and progesterone

Flow cytometry and transmission electron microscopy have been employed to show that a proportion of fresh and cultured human decidual stromal cells phagocytose latex particles. Phagocytosis of Escherichia coli by cultured decidual stromal cells was, however, very low. Stimulation of cultured decidual stromal cells with interleukin-1 alpha enhanced phagocytosis of both latex particles and E. coli. In contrast, when decidual stromal cells were cultured with progesterone under decidualizing conditions, phagocytic activity was reduced. These results suggest the existence of an immune-endocrine circuit involving decidual stromal cells.

Human Decidual Stromal Cells Protect Lymphocytes from Apoptosis

Human decidual stromal cells (DSC) have been shown to be involved in different immune functions that may be relevant for the relationship between the mother and fetus and hence for successful pregnancy. The expression of death ligands by fetal trophoblast and maternal decidual cells has been proposed as a mechanism for the establishment of materno-fetal immunotolerance. This study intended to elucidate the interrelations between DSC and lymphocytes. We analyzed the expression and function of death receptors and ligands in DSC maintained in culture. These DSC lines expressed CD95 and TNF-related apoptosis-inducing ligand receptor-2 (TRAIL-R2), although they were resistant to death receptor-mediated apoptosis. Regarding the expression of CD95L and TRAIL, it was variable among DSC lines although none of them induced apoptosis in death ligand-sensitive Jurkat T cells. Interestingly, most of the DSC lines, as well as fresh DSC, reduced apoptosis in Jurkat cells induced by anti-CD95 antibody and recombinant TRAIL. The protective effect of DSC was observed when they were co-cultured with Jurkat cells in Transwell plates, indicating that DSC may produce soluble factors of importance for lymphocyte survival. Moreover, the viability of peripheral blood lymphocytes and decidual lymphocytes was improved when co-cultured with DSC. Our results suggest that DSC, far from inducing apoptosis, may be relevant in the regulation of lymphocyte survival at the materno-fetal interface.

Expression of adhesion molecules by endothelial cells of early human decidua.

The expression of adhesion molecules by endothelial cells (EC) of early human decidua was studied with monoclonal antibodies and the immunoperoxidase technique. Although E-selectin, INCAM-110 and VCAM-1 were poorly detected on decidual EC, ICAM-1, P-selectin and DR antigens were highly expressed by these cells, some of which showed high endothelial venule-like morphology. Our results suggest that decidual EC are activated, and are probably involved in the active recruitment of leucocytes.