BACKGROUND: Immunomodulatory mechanisms of Sertoli cells (SCs) during phylogeny have not been described previously. This study attempted to reveal mechanisms of SC immune modulation in an evolutionary distant host. METHODS: The interaction of the SC cell line derived from Xenopus tropicalis (XtSC) with murine immune cells was studied in vivo and in vitro. The changes in the cytokine production, the intracellular and surface molecules expression on murine immune cells were evaluated after co-culturing with XtSCs. Migration of XtSCs in mouse recipients after intravenous application and subsequent changes in spleen and the testicular immune environment were determined by flow cytometry. RESULTS: The in vitro co-culture model was established, allowing the study of XtSCs interaction with murine immune cells. Intracellular staining of interleukin (IL-)10 revealed a significant increase in its expression in macrophages and B cells co-cultured with XtSCs, compared to both unstimulated cells and xenogeneic control. On the contrary, a significant decrease in Th lymphocytes expressing interferon-gamma was observed. The expression of both PD-1 ligands (PD-L1 and PD-L2) was upregulated on the macrophage surfaces after co-culture with XtSCs, but not with the controls. XtSCs migrated specifically to testes when administered intravenously and modulated systemic and local testicular microenvironment; this was detected by the expression of molecules associated with suppressive phenotype by CD45+ cells in both spleen and testes. CONCLUSION: We have demonstrated for the first time that SCs can migrate and modulate immune response in a phylogenetically distant host. It was further observed that SCs induce expression of molecules associated with immunosuppression, such as IL-10 and PD-1 ligands.

• MeSH
• antigeny CD274 MeSH
• antigeny CD279 * MeSH
• imunita MeSH
• interleukin-10 * MeSH
• ligandy MeSH
• modely nemocí na zvířatech MeSH
• myši MeSH
• Sertoliho buňky MeSH
• transplantace heterologní MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

It is becoming increasingly evident that selecting an optimal source of mesenchymal stromal cells (MSCs) is crucial for the successful outcome of MSC-based therapies. During the search for cells with potent regenerative properties, Sertoli cells (SCs) have been proven to modulate immune response in both in vitro and in vivo models. Based on morphological properties and expression of surface markers, it has been suggested that SCs could be a kind of MSCs, however, this hypothesis has not been fully confirmed. Therefore, we compared several parameters of MSCs and SCs, with the aim to evaluate the therapeutic potential of SCs in regenerative medicine. We showed that SCs successfully underwent osteogenic, chondrogenic and adipogenic differentiation and determined the expression profile of canonical MSC markers on the SC surface. Besides, SCs rescued T helper (Th) cells from undergoing apoptosis, promoted the anti-inflammatory phenotype of these cells, but did not regulate Th cell proliferation. MSCs impaired the Th17-mediated response; on the other hand, SCs suppressed the inflammatory polarisation in general. SCs induced M2 macrophage polarisation more effectively than MSCs. For the first time, we demonstrated here the ability of SCs to transfer mitochondria to immune cells. Our results indicate that SCs are a type of MSCs and modulate the reactivity of the immune system. Therefore, we suggest that SCs are promising candidates for application in regenerative medicine due to their anti-inflammatory and protective effects, especially in the therapies for diseases associated with testicular tissue inflammation.

• MeSH
• antiflogistika MeSH
• imunita MeSH
• lidé MeSH
• mezenchymální kmenové buňky * MeSH
• mitochondrie MeSH
• Sertoliho buňky * MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Nonspecific binding of conjugated antibodies represents a critical step which could significantly influence the results of immunostaining or flow cytometry. In this respect, various staining procedures and distinct cell types can alter the results obtained with different fluorochromes. In this study, we analysed nonspecific binding of R-phycoerythrin (R-PE)-conjugated antibodies to mouse mitogen-stimulated B and T lymphocytes. The cells were fixed, permeabilized and stained using isotype control antibodies conjugated with different fluorochromes and assessed by flow cytometry. R-PE-conjugated antibodies bound to LPS-stimulated B cells, in contrast to Con A-stimulated T cells, independently of their specificity. The percentage of R-PE positive B cells varied, according to the used antibodies or the fixation/permeabilization kit. Nevertheless, up to 30% of R-PE+ B cells after staining with R-PE-conjugated isotype control antibodies was detected. Furthermore, LPS-stimulated B cells bound nonspecifically, in a dose-dependent manner, unconjugated R-PE molecules. Con A-stimulated T cells slightly bound R-PE only in high concentrations. Similarly, the antibodies conjugated with other fluorochromes showed less than 1% of nonspecific binding independently of the manufacturer of antibodies or fixation/permeabilization kits. The data demonstrated that LPS-stimulated B cells, in contrast to Con A-stimulated T cells, bind R-PE nonspecifically following formaldehyde or paraformaldehyde fixation. Therefore, the results based on the use of R-PE-conjugated antibodies should be taken with a precaution.

• MeSH
• B-lymfocyty imunologie   MeSH
• fykoerythrin imunologie   metabolismus   MeSH
• mitogeny imunologie   MeSH
• monoklonální protilátky imunologie   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• T-lymfocyty imunologie   MeSH
• vazebná místa imunologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The clinical manifestation of leishmaniases depends on parasite species, host genetic background, and immune response. Manifestations of human leishmaniases are highly variable, ranging from self-healing skin lesions to fatal visceral disease. The scope of standard model hosts is insufficient to mimic well the wide disease spectrum, which compels the introduction of new model animals for leishmaniasis research. In this article, we study the susceptibility of three Asian rodent species (Cricetulus griseus, Lagurus lagurus, and Phodopus sungorus) to Leishmania major and L. donovani. The external manifestation of the disease, distribution, as well as load of parasites and infectiousness to natural sand fly vectors, were compared with standard models, BALB/c mice and Mesocricetus auratus. No significant differences were found in disease outcomes in animals inoculated with sand fly- or culture-derived parasites. All Asian rodent species were highly susceptible to L. major. Phodopus sungorus showed the non-healing phenotype with the progressive growth of ulcerative lesions and massive parasite loads. Lagurus lagurus and C. griseus represented the healing phenotype, the latter with high infectiousness to vectors, mimicking best the character of natural reservoir hosts. Both, L. lagurus and C. griseus were also highly susceptible to L. donovani, having wider parasite distribution and higher parasite loads and infectiousness than standard model animals.

• Publikační typ
• časopisecké články MeSH

The knowledge of mechanisms of regulation of IL-10 production by B cells remains still very limited. We show here that highly purified mouse B cells stimulated with LPS produce significant levels of IL-10, but Bregs in our model do not express detectable level of either Foxp3 or GATA-3. Nevertheless, IL-10 production by B cells is regulated by cytokines. In activated B cells, IL-10 production was significantly enhanced by IFN-γ and decreased in the presence of IL-4 or TGF-β. These findings are in sharp contrast with the observations in T cells, where IL-10 production correlates with GATA-3 or FoxP3 expression, and the cytokines regulate IL-10 production in a reverse manner than in activated B cells. These results thus show that the production of IL-10 by Bregs is regulated by cytokines independently of the expression of GATA-3 and FoxP3, which is clearly different from GATA-3-dependent IL-10 production by activated Th2 cells and FoxP3 expression in IL-10-producing Tregs.

• MeSH
• aktivace lymfocytů imunologie   MeSH
• faktor 1 indukovatelný hypoxií - podjednotka alfa imunologie   MeSH
• forkhead transkripční faktory metabolismus   MeSH
• interferon gama imunologie   MeSH
• interleukin-10 biosyntéza   MeSH
• interleukin-4 imunologie   MeSH
• kultivované buňky MeSH
• lipopolysacharidy imunologie   MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• regulační B-lymfocyty imunologie   MeSH
• regulační T-lymfocyty imunologie   MeSH
• Th2 buňky imunologie   MeSH
• transformující růstový faktor beta imunologie   MeSH
• transkripční faktor GATA3 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Retinal degenerative disorders are characterized by a local upregulation of inflammatory factors, infiltration with cells of the immune system, a vascular dysfunction and by the damage of retinal cells. There is still a lack of treatment protocols for these diseases. Mesenchymal stem cell (MSC)-based therapy using immunoregulatory, regenerative and differentiating properties of MSCs offers a promising treatment option. In this study, we analyzed the immunomodulatory properties of mouse bone marrow-derived MSCs after their intravitreal delivery to the inflammatory environment in the eye, caused by the application of pro-inflammatory cytokines IL-1β, TNF-α and IFN-γ. The intravitreal administration of these cytokines induces an increased expression of pro-inflammatory molecules such as IL-1α, IL-6, inducible nitric oxide synthase, TNF-α and vascular endothelial growth factor in the retina. However, a significant decrease in the expression of genes for all these pro-inflammatory molecules was observed after the intravitreal injection of MSCs. We further showed that an increased infiltration of the retina with immune cells, mainly with macrophages, which was observed after pro-inflammatory cytokine application, was significantly reduced after the intravitreal application of MSCs. The similar immunosuppressive effects of MSCs were also demonstrated in vitro in cultures of cytokine-stimulated retinal explants and MSCs. Overall, the results show that intravitreal application of MSCs inhibits the early retinal inflammation caused by pro-inflammatory cytokines, and propose MSCs as a promising candidate for stem cell-based therapy of retinal degenerative diseases.

• MeSH
• antivirové látky farmakologie   MeSH
• cytokiny metabolismus   MeSH
• imunomodulace účinky léků   imunologie   MeSH
• interferon gama farmakologie   MeSH
• interleukin-1beta farmakologie   MeSH
• mediátory zánětu farmakologie   MeSH
• mezenchymální kmenové buňky cytologie   metabolismus   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• oxid dusnatý metabolismus   MeSH
• retina cytologie   účinky léků   imunologie   metabolismus   MeSH
• TNF-alfa farmakologie   MeSH
• zánět imunologie   metabolismus   patologie   prevence a kontrola   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The successful application of mesenchymal stem cells (MSCs) remains a major challenge in stem cell therapy. Currently, several in vitro studies have indicated potentially beneficial interactions of MSCs with immunosuppressive drugs. These interactions can be even more complex in vivo, and it is in this setting that we investigate the effect of MSCs in combination with Cyclosporine A (CsA) on transplantation reaction and allogeneic cell survival. Using an in vivo mouse model, we found that CsA significantly promoted the survival of MSCs in various organs and tissues of the recipients. In addition, compared to treatment with CsA or MSCs alone, the survival of transplanted allogeneic cells was significantly improved after the combined application of MSCs with CsA. We further observed that the combinatory treatment suppressed immune response to the alloantigen challenge and modulated the immune balance by harnessing proinflammatory CD4+T-bet+ and CD4+RORγt+ cell subsets. These changes were accompanied by a significant decrease in IL-17 production along with an elevated level of IL-10. Co-cultivation of purified naive CD4+ cells with peritoneal macrophages isolated from mice treated with MSCs and CsA revealed that MSC-educated macrophages play an important role in the immunomodulatory effect observed on distinct T-cell subpopulations. Taken together, our findings suggest that CsA promotes MSC survival in vivo and that the therapeutic efficacy of the combination of MSCs with CsA is superior to each monotherapy. This combinatory treatment thus represents a promising approach to reducing immunosuppressant dosage while maintaining or even improving the outcome of therapy.

• MeSH
• alografty účinky léků   imunologie   MeSH
• cyklosporin farmakologie   terapeutické užití   MeSH
• cytokiny metabolismus   MeSH
• imunosupresiva farmakologie   terapeutické užití   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• preklinické hodnocení léčiv MeSH
• přežívání štěpu účinky léků   imunologie   MeSH
• T-lymfocyty účinky léků   metabolismus   MeSH
• transplantace mezenchymálních kmenových buněk * MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Pathogenesis of amyotrophic lateral sclerosis (ALS) involves several mechanisms resulting in a shift from a neuroprotective to a neurotoxic immune reaction. A promising tool for ALS treatment is represented by mesenchymal stem cells (MSCs), which possess both regenerative potential and immunomodulatory properties. In this study, we aimed to compare the immunomodulatory properties of MSCs isolated from the bone marrow of patients suffering from ALS and healthy donors. Moreover, the influence of proinflammatory cytokines on the immunoregulatory functions of MSCs was also evaluated. We found that MSCs from ALS patients and healthy donors comparably affected mitogen-stimulated peripheral blood mononuclear cells and reduced the percentage of T helper (Th)1, Th17 and CD8+CD25+ lymphocytes. These MSCs also equally increased the percentage of Th2 and CD4+FOXP3+ T lymphocytes. On the other hand, MSCs from ALS patients decreased more strongly the production of tumour necrosis factor-α than MSCs from healthy donors, but this difference was abrogated in the case of MSCs stimulated with cytokines. Significant differences between cytokine-treated MSCs from ALS patients and healthy donors were detected in the effects on the percentage of CD8+CD25+ and CD4+FOXP3+ T lymphocytes. In general, treatment of MSCs with cytokines results in a potentiation of their effects, but in the case of MSCs from ALS patients, it causes stagnation or even restriction of some of their immunomodulatory properties. We conclude that MSCs from ALS patients exert comparable immunomodulatory effects to MSCs from healthy donors, but respond differently to stimulation with proinflammatory cytokines. Graphical Abstract Treatment of mesenchymal stem cells (MSCs) with cytokines results in a potentiation of their effects, but in the case of MSCs from amyotrophic lateral sclerosis (ALS) patients, it causes stagnation (an equal reduction of the percentage of CD8+CD25+ T lymphocytes) or even restriction (no increase of proportion of CD4+FOXP3+ T lymphocytes) of some of their immunomodulatory properties. It means that MSCs from ALS patients exert comparable immunomodulatory effects to MSCs from healthy donors, but respond differently to stimulation with proinflammatory cytokines.

• MeSH
• amyotrofická laterální skleróza imunologie   MeSH
• buňky kostní dřeně imunologie   MeSH
• cytokiny metabolismus   MeSH
• imunologické faktory farmakologie   MeSH
• imunomodulace MeSH
• leukocyty mononukleární účinky léků   imunologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mezenchymální kmenové buňky imunologie   MeSH
• mitogeny farmakologie   MeSH
• T-lymfocyty pomocné-indukující účinky léků   imunologie   MeSH
• TNF-alfa biosyntéza   MeSH
• zdraví dobrovolníci pro lékařské studie MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Regulatory T cells (Tregs) play a critical role in the maintenance of a pregnancy. While the kinetics of the number of peripheral blood Tregs has been satisfactorily described in mouse models, analysis of these cell populations in human pregnancy is complicated by high variability in the quantity of Tregs and inconsistencies in the markers used for detecting different types of Treg. In the light of this, we set out to investigate the kinetics of various types of Treg, including CD45RA, GARP and PD-1(+) Tregs, in the peripheral blood of pregnant women in the first, second and third trimester, and at the time of delivery. Tregs, defined as a CD4(+)CD25(++)CD127(dim)Foxp3(+) population of leucocytes, were detected using flow cytometry. Natural thymus-derived Tregs and induced Tregs in the peripheral blood were distinguished by the expression or absence of a Helios marker, respectively. Our results showed that during normal pregnancy the sizes of various Treg subpopulations varied across women and also in an individual woman did not remain constant but varied significantly, most notable being the decrease observed at the time of delivery. Helios(-) cells were significantly less frequent in the peripheral blood of healthy pregnant women than Helios(+) cells, and the majority of Tregs were Helios(+)PD-1(+) Tregs.

• MeSH
• buněčná diferenciace MeSH
• forkhead transkripční faktory metabolismus   MeSH
• imunofenotypizace MeSH
• kultivované buňky MeSH
• lidé MeSH
• počet lymfocytů MeSH
• průtoková cytometrie MeSH
• regulační T-lymfocyty imunologie   MeSH
• T-lymfocyty - podskupiny imunologie   MeSH
• těhotenství imunologie   MeSH
• thymus imunologie   MeSH
• transformující růstový faktor beta krev   MeSH
• transkripční faktor Ikaros metabolismus   MeSH
• zdraví dobrovolníci pro lékařské studie MeSH
• Check Tag
• lidé MeSH
• těhotenství imunologie   MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Immunosuppressive drugs are used to suppress graft rejection after transplantation and for the treatment of various diseases. The main limitations of their use in clinical settings are severe side effects, therefore alternative approaches are desirable. In this respect, mesenchymal stem cells (MSCs) possess a regenerative and immunomodulatory capacity that has generated considerable interest for their use in cell-based therapy. Currently, MSCs are tested in many clinical trials, including the treatment of diseases which require simultaneous immunosuppressive treatment. Since the molecular targets of immunosuppressive drugs are also present in MSCs, we investigated whether immunosuppressive drugs interact with the activity of MSCs. Human MSCs isolated from the bone marrow (BM) or adipose tissue (AT) were cultured in the presence of clinical doses of five widely used immunosuppressive drugs (cyclosporine A, mycophenolate mofetil, rapamycin, prednisone and dexamethasone), and the influence of these drugs on several factors related to the immunosuppressive properties of MSCs, including the expression of immunomodulatory enzymes, various growth factors, cytokines, chemokines, adhesion molecules and proapoptotic ligands, was assessed. Glucocorticoids, especially dexamethasone, showed the most prominent effects on both types of MSCs and suppressed the expression of the majority of the factors that were tested. A significant increase of hepatocyte growth factor production in AT-MSCs and of indoleamine 2,3-dioxygenase expression in both types of MSCs were the only exceptions. In conclusion, clinically relevant doses of inhibitors of calcineurin, mTOR and IMPDH and glucocorticoids interfere with MSC functions, but do not restrain their immunosuppressive properties. These findings should be taken into account before preparing immunosuppressive strategies combining the use of immunosuppressive drugs and MSCs.

• MeSH
• imunosupresiva farmakologie   MeSH
• interferon gama agonisté   biosyntéza   MeSH
• kultivované buňky MeSH
• leukocyty mononukleární účinky léků   metabolismus   MeSH
• lidé MeSH
• mezenchymální kmenové buňky účinky léků   metabolismus   MeSH
• proliferace buněk účinky léků   fyziologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH