Search Results (54)

Background: MSCs possess strong immunoregulatory properties and play a central role in maintaining immune homeostasis by limiting inflammatory responses. Their function is highly plastic and influenced by environmental cues, including viral signals. How SARS-CoV-2-derived antigens affect MSC immunoregulation remains incompletely understood. This study aimed to investigate the impact of SARS-CoV-2 peptides on MSC-mediated immune modulation of T-cells. Methods: MSCs were stimulated directly with SARS-CoV-2 spike protein S peptides or cocultured with SARS-CoV-2 peptide-activated T-cells. TLR4 surface expression and receptor downstream signaling were assessed to evaluate pathway activation. MSC immunoregulatory function was analyzed by measuring suppression of TNF-α and IFN-γ expression and induction of CD4⁺FOXP3⁺ regulatory T-cells. TLR4 inhibition and lipopolysaccharide (LPS) stimulation were used to examine pathway specificity and interaction. Results: SARS-CoV-2 peptides activated TLR4-associated signaling in MSCs, increasing TLR4 expression and NF-κB phosphorylation. Peptide-treated MSCs showed impaired suppression of pro-inflammatory cytokines and reduced induction of regulatory T-cells. TLR4 inhibition prevented these effects. LPS induced similar effects, while combining LPS and peptide stimulation partially restored physiological T-cell cytokine suppression. Conclusions: SARS-CoV-2 peptides modulate MSC immunoregulatory function on T-cells via TLR4-dependent mechanisms. Full article

The survival and activation of both effector and regulatory CD4⁺T cells are promoted by cytokines in a complex series of interactions. Alloantigen-specific Regulatory T cells (Treg) constitutively express IL-2 receptor (CD25) and Foxp3. This discovery arose as the cells that transfer the alloantigen-specific transplant tolerance die in culture with specific alloantigens, unless the cultures are supplemented with cytokines from activated lymphocytes. One such cytokine was IL-2, but other cytokines are essential. We describe how the activation of Treg by antigens depends on cytokines produced by antigen-activated effector T cells. These cytokines also drive in parallel the activation of Treg. The Treg are induced to express similar transcription factors and chemokine receptors and have a similar cytokine responsiveness to the activated T effector cells. The activation of Treg by antigens is a two-step process: the first requires cytokines produced by effector T cells early in their activation, and the second step is driven by cytokines produced later by effector T cells during activation. Cytokines from Type 1 responses promote the induction of Th1-like Treg. Likewise, cytokines produced in Type 2, Type 3, and Tfh responses induce different pathways of Treg activation. Understanding the pathways for the activation and expansion of potent antigen-specific Treg will help produce Treg to control allograft rejection or autoimmunity. Currently, the complexity of the numerous potential pathways of activation of Treg remains incompletely understood. The dogma that IL-2 is the only driver of Treg activation may have hindered the development of highly potent antigen-specific Treg for therapy. Full article

The neonatal heart possesses a unique capacity for reparative healing after myocardial injury, unlike the adult heart. While immune cells, particularly T cells, regulate post-infarction inflammation, their role in age-dependent cardiac repair remains unclear. This study aimed to characterize the temporal activation of T cell subsets and their contribution to immune homeostasis and myocardial repair. Myocardial infarction was induced in mice of different ages, and T cell subsets (CD4+ T cells, CD8+ T cells, and CD4+Foxp3+ T [T-reg] cells) were analyzed using flow cytometry and RNA sequencing. Neonatal hearts exhibited CD4+ T cells, CD8+ T cells, and T-reg cells that gradually increased until seven days post-injury. Transcriptome analysis identified Rcn3 as a neonatal-specific, injury-responsive gene in T-reg cells, with minimal induction in adult and aged hearts, promoting a reparative microenvironment and exerting anti-fibrotic effects via the PI3K/Akt pathway. Under endoplasmic reticulum stress, Rcn3 activated unfolded protein response genes, and Rcn3-conditioned media reduced fibrosis-associated gene expression in adult cardiac fibroblasts. In a conditional knockout mouse model (Lck-cre; Rcn3^fl/fl), Rcn3 deletion in T cells led to impaired cardiac function recovery and increased fibrosis post-injury. These findings suggest that neonatal T-reg cells play a crucial role in cardiac repair, with Rcn3 as a potential therapeutic target for enhancing immune-mediated cardiac repair and limiting pathological remodeling in the adult heart. Full article

Autoimmune type 1 diabetes (T1D) results from the failure of the physiologic regulatory mechanisms that are designed to maintain immune tolerance to pancreatic beta cells. Consequently, the design of strategies to restore tolerance to beta cell antigens is an attractive objective of translational research. We have designed ultrasmall nanoparticles (NPs) loaded with a proinsulin (PI) fusion protein and an agonist for the aryl hydrocarbon receptor (AhR), a transcription factor promoting tolerance induction by different immune cells. We report that a 4 week-treatment with these NPs in non-obese diabetic (NOD) mice starting at disease onset induces temporary and sometimes durable disease remission. Mechanistically, short-term NP treatment induces a rapid depletion of islet infiltrates with a dramatic reduction in the number of CD8⁺ T cells and dendritic cells. This is accompanied by the emergence of B lymphocytes producing IL-10. In the rare mice that undergo durable disease remission, the disappearance of islet infiltrates is associated with the emergence of Foxp3⁺ CD4⁺ regulatory T cells, IFN-γ-producing memory T cells in the spleen, and draining lymph nodes (LNs). We conclude that treatment with these NPs could be of interest in the treatment of recent-onset autoimmune diabetes, but is unlikely to be sufficient for the induction of long-term remission as a stand-alone therapy. Full article

Despite advances in immunotherapy, the treatment of breast cancer still remains a major global problem. In a previous study, we showed that co-blockade of Interleukin-33/ST2 and Programmed death-1/Programmed death-ligand (PD-1/PD-L) signaling pathways strongly slows progression by enhancing the antitumor capacity of natural killer (NK) cells. The main aim of this study is to elucidate the exact effect of co-blockade on the T lymphocyte and macrophage effector cells. 4T1 cells were used to induct breast cancer in female BALB/C and BALB/C ST2^−/− mice. The mice, both BALB/C and BALB/C ST2^−/−, were treated with anti-PD-1 antibody on certain days. After the mice were sacrificed, T cells and macrophages were analyzed using flow cytometry; dual co-blockade increased significantly the percentage of M1 macrophages in the tumor microenvironment, followed by an increase in expression of CD86⁺ and TNFα⁺. T cell accumulation was significantly higher in the spleen and within the tumor microenvironment, with elevation in activation markers such as Interleukin-17, CD69, NKG2D, and FasL and a decrease in Interleukin-10 and FoxP3 expression. Co-blockade of the PD-1/PD-L axes and IL-33/ST2 axes shows promising results in reestablishing an effective immune response and offers a new perspective on improving immune response to breast carcinoma. Full article

Mature dendritic cells (DCs) are known to activate effector immune responses, whereas steady state immature DCs can induce tolerance. Several studies have targeted immature murine quiescent DCs in vivo with antigen, including donor alloantigens, for the induction of tolerance. Receptors expressed by specific DC subsets have been also targeted with antibodies linked with antigens to induce tolerance; for instance, in vivo targeting of the DCIR2⁺ DC subset with donor alloantigen resulted in long-term survival of heart and skin transplants. DCs also express sialic acid immunoglobulin-like lectin (Siglec) receptors, and these have been successfully targeted with myelin oligiodendrocyte glycoprotein (MOG) antigen to induce tolerance in experimental autoimmune encephalomyelitis (EAE). We investigated, in a mismatched model of skin transplant (B6K^d into B6 recipient mice), whether targeting a sialylated alloantigen K^d (Sia-K^d) to Siglecs on recipient DCs promoted transplant survival. The injection of α2,3 Sia-K^d into B6 recipient mice prior to B6K^d skin transplantation, by binding to Batf3 dependent DCs, resulted in prolonged skin graft survival and an increase in CD4⁺CD62L⁺Foxp3⁺ Tregs. Targeting Siglecs on DC subsets in vivo represents a novel way of improving transplant survival. Full article

Background/Objectives: Inflammatory bowel disease (IBD) is a chronic inflammatory condition encompassing ulcerative colitis (UC) and Crohn’s disease (CD). The role of environmental factors in the pathogenesis of IBD remains elusive. Nevertheless, evidence suggests a pivotal role of viruses, specifically Epstein–Barr virus (EBV), in the progression of IBD through mechanisms such as molecular mimicry and bystander activation. Our previous findings demonstrate EBV DNA’s significant role in exacerbating colitis symptoms and elevating the levels of the pro-autoimmune cytokine interleukin-17A (IL-17A) in an IBD mouse model via toll-like receptor 9 (TLR9). Therefore, we aimed to examine the role of EBV particles in the pathogenesis of IBD, and the potential role of TLR9 inhibition in ameliorating disease outcomes. Methods: Three days post colitis induction, EBV particles were intra-rectally injected into female C57BL/6J mice, followed by the intra-peritoneal administration of TLR9 inhibitor. Thereupon, mice were monitored daily and the disease activity index (DAI), colon lengths, and damage scores, as well as the number of cells, double-positive for IL-17A+ and IFN-γ+, and triple-positive for IL-17A+, IFN-γ+, and FOXP3+, were evaluated. Results: Our findings revealed a significant role of TLR9 inhibition in mitigating colitis features in an EBV-injected IBD mouse model compared to the control group. Conclusions: These results indicate an essential role of TLR9 in initiating immune responses against recurrent EBV reactivation events, which ultimately contributes to inflammation aggravation in IBD patients. Consequently, TLR9 could serve as a potential therapeutic target to alleviate the severe symptoms of IBD in EBV-infected individuals. Full article

Autoimmune diseases such as multiple sclerosis (MS) are characterized by a loss of self-tolerance, driven by diminished regulatory T cell (Treg) function and elevated Th1/Th17 responses. Existing therapies broadly suppress the immune system without correcting this imbalance, often leading to adverse effects. LPX3, a novel small-molecule T cell immunoglobulin and mucin domain-containing 3 and 4 (Tim-3/4) receptor agonist, was developed to restore immune tolerance via Treg induction. In this study, LPX3 was formulated into a liposomal oral delivery system, enabling efficient uptake through the gastrointestinal tract and lymphatic targeting. In vitro and in vivo analyses confirmed LPX3’s ability to expand CD4⁺Foxp3⁺ Tregs in a dose-dependent manner. In a MOG-induced experimental autoimmune encephalomyelitis (EAE) mouse model of MS, both prophylactic and therapeutic oral administration of LPX3 significantly delayed disease onset, reduced symptom severity, and improved survival. Importantly, efficacy was achieved without antigen co-delivery, indicating an antigen-independent mechanism of immune modulation. LPX3 liposomes showed deep lymph node penetration and colocalization with immune cells, supporting its functional delivery to key immunological sites. These findings suggest LPX3 is a promising candidate for treating autoimmune diseases by re-establishing immune regulation through oral, antigen-agnostic tolerance induction. Full article

Introduction: Advanced chronic lymphocytic leukemia (CLL) is accompanied by increased circulating regulatory T cells (Tregs) and increased susceptibility to severe infections, which were also shown to entail a striking induction of FOXP3 expression in Tregs. As homeostasis of the most suppressive CD45RA⁻FOXP3^high activated Treg (aTreg) subset differs, it is critical to analyse homeostatic signalling in Treg subsets. Materials and Methods: In this study, by using conventional and imaging flow cytometry, we monitored STAT5 signalling/phosphorylation (pSTAT5) and investigated Treg subsets in relation to the Binet stage, the total tumor mass score (TTM) and the disease course during a follow-up of 37 patients with CLL. Results: The aTreg percentage was significantly increased among CD4⁺ T cells from patients with advanced disease and significantly correlated with the TTM. A subgroup of patients with higher aTreg percentages among CD4⁺FOXP3⁺ T cells at the start of therapy was characterised by more frequent episodes of severe infections during follow-up. Conclusions: The results suggesting that an aTreg fraction could represent a possible marker of a severe disease course with infectious complications. Augmented homeostatic STAT5 signalling could support aTreg expansion, as higher pSTAT5 levels were significantly correlated with an increased aTreg frequency among CD4⁺FOXP3⁺ T cells during the follow-up of patients on therapy, as well as following SARS-CoV-2 antigen-specific stimulation in vitro. Full article

The nanosized vesicles secreted from various cell types into the surrounding extracellular space are called extracellular vesicles (EVs). Although mesenchymal stem cell-derived EVs are known to have immunomodulatory effects in asthmatic mice, the role of identified pulmonary genes in the suppression of allergic airway inflammation remains to be elucidated. Moreover, the major genes responsible for immune regulation in allergic airway diseases have not been well documented. This study aims to evaluate the immunomodulatory effects of secretoglobin family 1C member 1 (SCGB1C1) on asthmatic mouse models. C57BL/6 mice were sensitized to ovalbumin (OVA) using intraperitoneal injection and were intranasally challenged with OVA. To evaluate the effect of SCGB1C1 on allergic airway inflammation, 5 μg/50 μL of SCGB1C1 was administrated intranasally before an OVA challenge. We evaluated airway hyperresponsiveness (AHR), total inflammatory cells, eosinophils in the bronchoalveolar lavage fluid (BALF), lung histology, serum immunoglobulin (Ig), the cytokine profiles of BALF and lung-draining lymph nodes (LLN), and the T cell populations in LLNs. The intranasal administration of SCGB1C1 significantly inhibited AHR, the presence of eosinophils in BALF, eosinophilic inflammation, goblet cell hyperplasia in the lung, and serum total and allergen-specific IgE. SCGB1C1 treatment significantly decreased the expression of interleukin (IL)-5 in the BALF and IL-4 in the LLN, but significantly increased the expression of IL-10 and transforming growth factor (TGF)-β in the BALF. Furthermore, SCGB1C1 treatment notably increased the populations of CD4⁺CD25⁺Foxp3⁺ regulatory T cells (Tregs) in asthmatic mice. The intranasal administration of SCGB1C1 provides a significant reduction in allergic airway inflammation and improvement of lung function through the induction of Treg expansion. Therefore, SCGB1C1 may be the major regulator responsible for suppressing allergic airway inflammation. Full article

We have previously performed preclinical studies with the oxidized mannan-conjugated peptide MOG35–55 (OM-MOG35–55) in vivo (EAE mouse model) and in vitro (human peripheral blood) and demonstrated that OM-MOG35–55 suppresses antigen-specific T cell responses associated with autoimmune demyelination. Based on these results, we developed different types of dendritic cells (DCs) from the peripheral blood monocytes of patients with multiple sclerosis (MS) or healthy controls presenting OM-MOG35–55 or MOG-35–55 to autologous T cells to investigate the tolerogenic potential of OM-MOG35–55 for its possible use in MS therapy. To this end, monocytes were differentiated into different DC types in the presence of IL-4+GM-CSF ± dexamethasone (DEXA) ± vitamin D3 (VITD3). At the end of their differentiation, the DCs were loaded with peptides and co-cultured with T cells +IL-2 for 4 antigen presentation cycles. The phenotypes of the DC and T cell populations were analyzed using flow cytometry and the secreted cytokines using flow cytometry or ELISA. On day 8, the monocytes had converted into DCs expressing the typical markers of mature or immature phenotypes. Co-culture of T cells with all DC types for 4 antigen presentation cycles resulted in an increase in memory CD4+ T cells compared to memory CD8+ T cells and a suppressive shift in secreted cytokines, mainly due to increased TGF-β1 levels. The best tolerogenic effect was obtained when patient CD4+ T cells were co-cultured with VITD3-DCs presenting OM-MOG35–55, resulting in the highest levels of CD4+PD-1+ T cells and CD4+CD25+Foxp3+ Τ cells. In conclusion, the tolerance induction protocols presented in this work demonstrate that OM-MOG35–55 could form the basis for the development of personalized therapeutic vaccines or immunomodulatory treatments for MS. Full article

Forkhead box protein 3 (FoxP3) is a key transcription factor responsible for the development, maturation, and function of regulatory T cells (Tregs). The FoxP3 pre-mRNA is subject to alternative splicing, resulting in the translation of multiple splice variants. We have shown that Tregs from patients with amyotrophic lateral sclerosis (ALS) have reduced expression of full-length (FL) FoxP3, while other truncated splice variants are expressed predominantly. A correlation was observed between the reduced number of Tregs in the peripheral blood of ALS patients, reduced total FoxP3 mRNA, and reduced mRNA of its FL splice variant. Induction of FL FoxP3 was achieved using splice-switching oligonucleotides capable of base pairing with FoxP3 pre-mRNA and selectively modulating the inclusion of exons 2 and 7 in the mature mRNA. Selective expression of FL FoxP3 resulted in the induction of CD127^low, CD152, and Helios-positive cells, while the cell markers CD4 and CD25 were not altered. Such Tregs had an increased proliferative activity and a higher frequency of cell divisions per day. The increased suppressive activity of Tregs with the induced FL FoxP3 splice variant was associated with the increased synthesis of the pro-apoptotic granzymes A and B, and perforin, IL-10, and IL-35, which are responsible for contact-independent suppression, and with the increased ability to suppress telomerase in target cells. The upregulation of Treg suppressive and proliferative activity using splice-switching oligonucleotides to induce the predominant expression of the FoxP3 FL variant is a promising approach for regenerative cell therapy in Treg-associated diseases. Full article

Inflammatory bowel disease (IBD) is characterized by chronic intestinal inflammation and its treatment varies widely; however, when inflammation is high, a complete nutrient containing pre-digested elemental diet (ED) is used to preserve the intestinal tract. In this study, we investigated the mechanisms underlying the effectiveness of EDs for IBD using mice. C57BL/6 mice were orally treated with the ED (5 mL/day) and its ingredient L-tryptophan (Trp) (1–100 mg/kg), respectively. Flow cytometry analysis revealed that treatment with the ED and Trp (10 and 100 mg/kg) significantly increased the percentage of splenic CD4+-/CD25+-/Foxp3+ regulatory T cells (Tregs). In the 2% DSS-induced colitis-mouse model, Trp administration (100 mg/kg) led to a significant decrease in TNF-α and increase in IL-10 in the serum as well as a significant decrease in the inflammation score. Furthermore, the aryl hydrocarbon receptor (AhR) agonistic activity, which is a key function of Treg induction, of Trp and 15 Trp metabolites was characterized using a highly sensitive DR-EcoScreen cell assay. Five Trp metabolites, including L-kynurenine, acted as AhR agonists, while Trp did not. Taken together, these results suggest that the ED treatment has a Trp-dependent immunoregulatory effect, and several Trp metabolites that activate the AhR might contribute to induction of remission in patients with IBD. Full article

Atopic dermatitis (AD) is a recurrent allergic disease characterized by symptoms such as itching, redness, swelling, dryness, scaling skin, inflammation, and tissue damage. The underlying pathogenesis of AD remains unclear. Steroid drugs are commonly used in the clinical treatment of AD; however, their long-term use may lead to associated complications. Numerous studies have indicated that probiotics could modulate the immune system, enhance immune function, or suppress excessive immune responses. In this study, Lactobacillus paracasei subsp. paracasei NTU 101 (NTU 101) was orally administered for a duration of 4 weeks, followed by the induction of AD using ovalbumin (OVA) in a mouse model. The skin condition of the stimulated site was observed during the induction period. Subsequently, the serum immunoglobulin E (IgE) content, splenocyte T cell typing, and skin histological interpretation were examined to evaluate the efficacy of NTU 101 in alleviating AD symptoms in allergen-exposed animals. The findings indicated that administering NTU 101 beforehand effectively alleviated skin symptoms in animals with AD. It reduced the infiltration of inflammatory cells in skin tissue sections, and compared to the OVA group, there was a significant reduction in the thickening of the epidermal cell layer (decreased from 89.0 ± 20.2 µM to 48.6 ± 16.0 µM) and dermis layer (decreased from 310.3 ± 69.0 µM to 209.7 ± 55.5 µM). Moreover, the proportion of regulatory T (Treg) cells and T helper 2 (Th2) cells in splenocytes significantly increased, while the proportions of T helper 1 (Th1) and T helper 17 (Th17) cells did not differ. It is speculated that the potential mechanism by which NTU 101 prevents AD involves increasing the expression of Forkhead box protein P3 (FOXP3) and promoting Treg cell maturation, thereby alleviating allergic reaction symptoms associated with AD. Full article

The reduced burden of helminth parasites in industrialized countries is probably one of the reasons for the increased prevalence of autoimmune disorders such as multiple sclerosis (MS). The current study aimed to evaluate the potential preventive effects of hydatid cyst fluid (HCF) on the disease severity in an EAE mouse model of MS. EAE-induced mice were treated with HCF before and after EAE induction. An RT-PCR-based evaluation of IFN-γ, IL-1β, TNF, T-bet, IL-4, GATA3, IL-17, RoRγ, TGF-β, and FOXP3 expression levels in splenocytes and an ELISA-based analysis of IFN-γ and IL-4 levels in cell culture supernatant of splenocytes were performed. Histopathological examinations of mice during the study were also conducted. The expression levels of T-bet, IL-4, GATA3, TGF-β, and FOXP3 in EAE + HCF mice were significantly higher compared to EAE + PBS mice. In the EAE + HCF group, the expression levels of IFN-γ, IL-1β, and TNF were significantly lower than in the EAE + PBS group. The histopathological results showed significantly reduced inflammation and demyelination in EAE + HCF mice compared to EAE + PBS mice. Our study provides proof-of-concept in the EAE mouse model of MS that helminth-derived products such as HCF have a potential prophylactic effect on MS development and present a novel potential therapeutic strategy. Full article