Human Herpes simplex 1 virus infection of endometrial decidual tissue-derived MSC alters HLA-G expression and immunosuppressive functions
Introduction
Mesenchymal stromal/stem cells (MSC), known for their multi-lineage differentiation potential, retain also pleiotropic immunosuppressive functions via a variety of mediators. This inhibitory capacity has raised much interest prompting studies investigating MSC role as an immunosuppressant, in particular in steroid-refractory GvHD after allogeneic hematopoietic stem cell transplantation and in autoimmune diseases [1].
Considering a variety of MSC sources, our previous results demonstrated that one of the players of this immunomodulation can be ascribed to the Human Leukocyte Antigen (HLA)-G [2]. HLA-G is a non-classical HLA class I molecule characterized by four membrane-bound (HLA-G1, -G2, -G3 and -G4) and three secreted soluble isoforms (HLA-G5, -G6, -G7) [3]. HLA-G retains tolerogenic functions, inducing apoptosis of activated CD8+ T cells [4], acting on T regulatory cells [5], modulating the activity of natural killer cells [6] and of dendritic cells [7] and blocking allo-cytotoxic T lymphocyte response [8]. These immune-regulatory properties are mediated by the interaction of HLA-G molecules with specific inhibitory receptors: ILT2 (LILRB1/CD85j), ILT4 (LILRB2/CD85d), CD8 and KIR2DL4 (CD158d) expressed in immune cells [9]. In humans, HLA-G is expressed during physiological conditions in placental trophoblast cells at maternal–fetal interface during pregnancy, in the thymus [5], cornea, nail matrix [6], pancreas [8], monocytes [9], erythroid [10], and endothelial precursors. However, HLA-G expression can be also detected in different immune cell populations, such as T-cells [11], [12], antigen-presenting cells [12], [13], and in immune-regulatory cell populations, such as mesenchymal stem cells [14], [15]. In these contexts, HLA-G expression is fundamental for the maintenance of an immune-privileged site, reducing the activation of the immune responses.
Combining the evidence of MSC as a source of HLA-G [2] and the physiological role of this molecule in female genital tract [16], we here wanted to verify whether human MSC obtained from endometrial decidual tissue (EDT) found within menstrual blood, could express HLA-G with the goal to characterize a more amenable MSC source for cell and gene therapies.
Endometrial tissues, as a source of EDT-MSC, are characterized by a microbiota that plays a crucial role in maintaining health. Disequilibrium of the microbiota has been associated with increased risk of pelvic infections, that could cause alterations in cell functions and molecular expression [17]. Thus, we took advantage of this research project, to additionally address whether viruses, and in particular Human Herpesviruses, as involved in severe opportunistic infections in the patients following allogeneic stem cell transplant [18], [19], was able to infect EDT-MSC and down-modulate HLA-G expression. We used Human Herpes simplex virus 1 (HSV-1) as a model of EDT-MSC infection.
Section snippets
EDT-MSC isolation and culture
EDT-MSC were isolated from menstrual blood as previously reported [20]. Briefly, menstrual blood was collected from healthy female volunteer donors (N = 5) during the first few days of the cycle. Written consent was obtained from each donor and the Local Ethical Committee approved cells donation for research purposes. Each donor has been endowed with a menstrual cup (Yuuki menstrual cup, BETA s.r.o., Czech Republic) to collect blood, which was transferred in Alpha MEM (Gibco) with 1%
EDT-MSC retain basic MSC features MSC
were obtained from menstrual blood samples (n = 5) after approved informed consent and were characterized by microscopic observation and flow cytometry analyses. As seen in Fig. 1A.
EDT-MSC appeared as small spindle-shaped elements whose FACS analyses revealed the positivity for CD90, CD73, CD105 and negative for CD45 and CD34 (Fig. 1B). In vitro expanded EDT-MSC resulted also strongly positive for CD29, CD59, HLA-ABC and CD10 (Fig. 1C) and negative for CD80, HLA-DR (II class), CD184, CD106 and
Discussion
In our present work, we describe that also EDT-MSC express HLA-G molecules and exhibit an immunosuppressive function dependent on culture passages as previously demonstrated for bone-marrow and dental pulp derived MSC [2]. EDT-MSC are permissive to HSV-1 infection and sustain a productive infection, as previously reported for human bone-marrow-derived, adipose-derived MSC and fetal membrane derived-MSC [28], [30].
Interesting, we observed that HSV-1 infection critically impairs the
Author’s contribution
DB carried out HLA-G analysis and lymphoproliferative analysis; FR and GG carried out EDT-MSC purification; AR carried out HSV-1 infections; DC carried out the flow cytometry; RC participated in the ELISA assay; RR and MD performed the statistical analysis, design of the study. All authors read and approved the final manuscript.
Acknowledgements
We thank Iva Pivanti for technical support and Linda M. Sartor for language revision of the manuscript.
Conflict of interest
All authors declare no financial competing interests. All authors declare no non-financial competing interests.
Sources of funding
This work was supported by Italian Health Ministry - Ricerca Finalizzata GR-2011-02346947.
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