HLA-G*0105N null allele encodes functional HLA-G isoforms.

Expression of the nonclassical HLA class I antigen, HLA-G, is associated with immune tolerance in view of its role in maintaining the fetus in utero, allowing tumor escape, and favoring graft acceptance. Expressed on invasive trophoblast cells, HLA-G molecules bind inhibitory receptors on maternal T lymphocytes and NK cells, thereby blocking their cytolytic activities and protecting the fetus from maternal immune system attack. The HLA-G gene consists of 15 alleles, including a null allele, HLA-G*0105N. HLA-G*0105N presents a single base deletion, preventing translation of both membrane-bound (HLA-G1) and full-length soluble isoforms (HLA-G5) as well as of the spliced HLA-G4 isoform. The identification of healthy subjects homozygous for this HLA-G null allele suggests that the HLA-G*0105N allele may generate other HLA-G isoforms, such as membrane-bound HLA-G2 and -G3 and the soluble HLA-G6 and -G7 proteins, which may substitute for HLA-G1 and -G5, thus assuming the immune tolerogeneic function of HLA-G. To investigate this point, we cloned genomic HLA-G*0105N DNA and transfected it into an HLA-class I-positive human cell line. The results obtained indicated that HLA-G proteins were indeed present in HLA-G*0105N-transfected cells and were able to protect against NK cell lysis. These findings emphasize the role of the other HLA-G isoforms as immune tolerogeneic molecules that may also contribute to maternal tolerance of the semiallogenic fetus as well as tumor escape and other types of allogeneic tissue acceptance

Expression of HLA-G in human cornea, an immune-privileged tissue.

Human leukocyte antigen (HLA)-G retains the capacity to modulate immune responses, favoring the establishment of tolerance in solid-tissue allotransplants. To better understand the mechanisms that promote corneal allograft survival, we investigated whether HLA-G was an immunoregulatory factor involved in corneal immunology. We therefore sought HLA-G expression in corneal tissues. Corneal transplantation consists in replacing the center of a diseased cornea with normal corneal tissue. Two corneal parts are not used in such surgery: diseased central corneal tissue and peripheral normal cornea. For this study, we used healthy corneas obtained from deceased donors and diseased corneas obtained from patients with pseudophakic bullous keratopathy or keratoconus who had undergone corneal transplantation. Immunohistochemical analysis carried out on the cryopreserved corneas showed a positive immunohistochemical staining with anti-HLA-G, anti-HLA-A, -B, and -C, and anti-HLA class I monoclonal antibodies. Staining was obtained for keratocytes, epithelial cells, and endothelial cells from both healthy and pathologic human corneas, revealing the presence of HLA class I proteins, including HLA-G. HLA-G transcripts were detected in normal cornea by reverse transcriptase-polymerase chain reaction with a classical pattern of alternative splicing. The detection of HLA-G protein in adult corneas leads to the conclusion that this protein may contribute to the maintenance of the privileged immune status of cornea

Expression of HLA-G in inflammatory bowel disease provides a potential way to distinguish between ulcerative colitis and Crohn's disease.

In addition to being involved in nutrient uptake, the epithelial mucosa constitute the first line of defense against microbial pathogens. A direct consequence of this physiological function is a very complex network of immunological interactions that lead to a strong control of the mucosal immune balance. The dysfunction of immunological tolerance is likely to be a cause of inflammatory bowel disease (IBD), ulcerative colitis (UC) and Crohn's disease (CD). HLA-G is a non-classical major histocompatibility complex (HLA) class I molecule, which is highly expressed by human cytotrophoblast cells. These cells play a role in immune tolerance by protecting trophoblasts from being killed by uterine NK cells. Because of the deregulation of immune system activity in IBD, as well as the immunoregulatory role of HLA-G, we have analyzed the expression of HLA-G in intestinal biopsies of patients with UC and CD. Our study shows that the differential expression of HLA-G provides a potential way to distinguish between UC and CD. Although the reason for this differential expression is unclear, it might involve a different mechanism of immune regulation. In addition, we demonstrate that in the lamina propria of the colon of patients with UC, IL-10 is strongly expressed. In conclusion, the presence of HLA-G on the surface of intestinal epithelial cell in patients with UC lends support to the notion that this molecule may serve as a regulator of mucosal immune responses to antigens of undefined origin. Thus, this different pattern of HLA-G expression may help to differentiate between the immunopathogenesis of CD and UC

Spatial and temporal mapping of c-kit and its ligand, stem cell factor expression during human embryonic haemopoiesis.

Receptor tyrosine kinases (RTKs) mediate cellular responses to the extracellular signals involved in the regulation of cell differentiation and proliferation. Ligand binding initiates a cascade of events, such as receptor dimerization and tyrosine phosphorylation. The c-kit gene encodes an RTK for stem cell factor (SCF), (c-kit ligand, KL), both of which play a critical role in the differentiation and growth of haemopoietic stem cells (HSCs). We investigated the expression of the c-kit and SCF genes and the presence of the corresponding proteins in haemopoietic tissues during human embryogenesis. We have examined c-kit and SCF transcripts levels in human embryonic yolk sac, the AGM region, and liver at different stages of gestation (days 25 to 63), using RT-PCR amplification combined with PhosphorImager quantitative analysis and RNase Protection Assay (RPA). Weak levels of SCF gene expression were observed in the AGM region (days 25 to 34) and high levels were found in the early-stage liver (day 34). The expression of c-kit transcript was observed in all studied tissues, but at various levels. The restricted presence of SCF protein following mRNA expression was demonstrated in embryonic liver CD38+ haemopoietic cells by immunocytochemistry. These observations suggest that the biological function of the c-kit receptor plays an important role in the early stages of human haemopoiesis, and that c-kit/SCF signalling is particularly involved in early human definitive haemopoiesis

C-fos and c-myc modulation, mitogenic effect and Ia expression in the P388D1 murine macrophage line treated by immunomodifiers.

The aim of the present study was to investigate whether the early modulation of the c-fos and c-myc oncogenes could give some orientation to the impact of immunomodulators on the monocyte-macrophage lineage. In order to work in a homogeneous system we used the P388D1 mouse macrophage cell-line which is considered as an almost mature macrophage. When P388D1 cells were stimulated by LPS, interferon-gamma or the association of both compounds, no direct correlation could be found between the modulation of DNA synthesis and the early expression of the c-fos and c-myc oncogenes. The positive regulation of Ia antigen expression seemed to correlate with the absence of induction of c-fos oncogene

Classical and non-classical HLA molecules and p16(INK4a) expression in precursors lesions and invasive cervical cancer

Objectives: Viruses and turnout cells may regulate the expression of HLA molecules on the cell surface to escape immune system surveillance. Absence of classical HLA class I molecules may impair the action of specific cytotoxic cells, whereas non-classical HLA class I molecules may regulate innate and adaptive immune cells. We assess here the possible associations between classical/non-classical class I HLA and p16(INK4a) molecule expression in cervical biopsies of women infected with HPV, stratified according to grade of the lesion and HPV type. Study design: Cervical biopsies (N = 74) presenting cervical intraepithelial neoplasia grade 1 (CIN1) (n = 31), CIN2-3 (n = 19), and invasive cancer (n = 14) were evaluated alongside 10 normal cervical specimens. Results: HLA-A/B/C/G staining was observed in the early stages of HPV infection. A significant association was detected between HLA-A/B/C staining and HPV16/18 infection (OR = 0.12, 95%CI: 0.0163-0.7899; p = 0.04). HLA-E expression increased with the progression of the lesion (chi(2)-test for trend = 4.01; p = 0.05), and a significant association was found between HLA-E staining and HPV16/18 infection (OR = 11.25, 95%CI: 2.324-54.465; p = 0.003). Irrespective of the grade of the lesion, HLA-A/B/C staining and p16(INK4a) presented a good concordance (Kappa: 0.67). Conclusions: HLA-E overexpression seemed to be associated with invasive cancer and HPV16/18 infection. (C) 2008 Elsevier Ireland Ltd. All rights reserved.FAPESP[03/01019-5]FAPESP[01/02908-2