Search Results (389)

Podoplanin (PDPN) overexpression is associated with poor clinical outcomes in various tumors. PDPN is involved in malignant tumor progression by promoting invasiveness and metastasis. Therefore, PDPN is considered a promising target of monoclonal antibody (mAb)-based therapy. Because PDPN also plays an essential role in normal cells such as kidney podocytes, cancer specificity is required to reduce adverse effects on normal cells. We developed a cancer-specific mAb (CasMab) against PDPN, PMab-117 (rat IgM, kappa), by immunizing rats with PDPN-overexpressed glioblastoma cells. The recombinant mouse IgG_2a-type PMab-117 (PMab-117-mG_2a) reacted with the PDPN-positive tumor PC-10 and LN319 cells but not with PDPN-knockout LN319 cells in flow cytometry. PMab-117-mG_2a did not react with normal kidney podocytes and normal epithelial cells from the lung bronchus, mammary gland, and corneal. In contrast, one of the non-CasMabs against PDPN, NZ-1, showed high reactivity to PDPN in both tumor and normal cells. Moreover, PMab-117-mG_2a exerted antibody-dependent cellular cytotoxicity in the presence of effector splenocytes. In the human tumor xenograft models, PMab-117-mG_2a exhibited potent antitumor effects. These results indicated that PMab-117-mG_2a could be applied to antibody-based therapy against PDPN-expressing human tumors while reducing the adverse effects. Full article

Nanomedicine holds great potential for revolutionizing medical treatment. Ongoing research and advancements in nanotechnology are continuously expanding the possibilities, promising significant advancements in healthcare. To fully harness the potential of nanotechnology in medical applications, it is crucial to conduct safety evaluations for the nanomedicines that offer effective benefits in the preclinical stage. Our recent efficacy studies indicated that rapamycin perfluorocarbon (PFC) nanoparticles showed promise in mitigating cisplatin-induced acute kidney injury (AKI). As cisplatin is routinely administered to ovarian cancer patients as their first-line chemotherapy, in this study, we focused on evaluating the safety of rapamycin PFC nanoparticles in mice bearing ovarian tumor xenografts. Specifically, this study evaluated the effects of repeat-dose rapamycin PFC nanoparticle treatment on vital organs, the immune system, and tumor growth and assessed pharmacokinetics and biodistribution. Our results indicated that rapamycin PFC nanoparticle treatment did not cause any detectable adverse effects on cardiac, renal, or hepatic functions or on splenocyte populations, but it reduced the splenocyte secretion of IL-10, TNFα, and IL12p70 upon IgM stimulation. The pharmacokinetics and biodistribution results revealed a significant enhancement in the delivery of rapamycin to tumors by rapamycin PFC nanoparticles, which, in turn, led to a significant reduction in ovarian tumor growth. Therefore, rapamycin PFC nanoparticles have the potential to be clinically beneficial in cisplatin-treated ovarian cancer patients. Full article

(1) Background: The adjuvant properties of flagellin from various bacterial species have been extensively studied; however, a systematic comparison of the immunoadjuvant effects of flagellins from different bacterial species is lacking. This study aims to analyze the amino acid sequences and structural features of flagellins from Escherichia coli (FliC_E.C), Salmonella enterica serotype Typhimurium (FliC_S.T), and Pseudomonas aeruginosa (FliC_P.A), and to evaluate their adjuvant activities in terms of Toll-like receptor 5 (TLR5) activation, antibody production, and cytokine responses in a murine model. (2) Methods: Bioinformatics analysis was conducted to compare the amino acid sequences and structural domains (D0, D1, D2, and D3) of flagellins from the three bacterial species. PyMol atomic models were used to confirm structural differences. Toll-like receptor 5 (TLR5) activation assays were performed to measure IL-8 and TNF-α production in vitro. The IgG antibody titers against the model antigen FaeG and cytokine responses, including IL-4 and TNF-α secretion were evaluated in a murine model. (3) Results: Bioinformatics analysis revealed that the D0 and D1 domains are highly conserved, whereas the D2 and D3 domains exhibit significant variability across the three species. Structural analysis via PyMol confirmed these differences, particularly in the D2 and D3 domains. TLR5 activation assays showed that FliC_S.T and FliC_P.A induced higher levels of IL-8 and TNF-α production compared to FliC_E.C, indicating species-specific variations in TLR5 activation. In the murine model, FliC_S.T as an adjuvant produced higher antibody titers against FaeG and increased IL-4 secretion in splenocytes compared to FliC_E.C and FliC_P.A. FliC_P.A induced higher TNF-α expression than FliC_S.T and FliC_E.C, suggesting FliC_S.T and FliC_P.A are more effective at inducing T-cell responses. (4) Conclusions: This study highlights the potential of FliC_S.T and FliC_P.A as potent vaccine adjuvants. The results provide insights into the structure–function relationships of these flagellins and support their application in enhancing immune responses against diverse pathogens. Full article

In Europe and North America, the prevalence of pulmonary nontuberculous mycobacteria (NTM) is increasing. Most pulmonary NTM infections are caused by the Mycobacterium avium complex (MAC). Sadly, the treatment of pulmonary MAC is suboptimal with failure rates ranging from 37% to 58%. Therefore, there is a need to develop new therapeutics. Developing new immunotherapies and studying their interaction with standard or new drugs requires reliable assays. Four different assays including CFSE-based flow cytometry, in vitro protection assays, IFN-γ ELISPOT, and murine infection models were optimized using a reference strain of MAC (ATCC 700898) to help with the development of immunotherapies for MAC. Expansion of proliferating and IFN-γ producing human T cells is optimal after 7 days of stimulation with MAC at a multiplicity of infection (MOI) of 0.1, achieving a stimulation index of 26.5 ± 11.6 (mean ± SE). The in vitro protection assay for MAC works best by co-culturing T cells expanded for 7 days with MAC (MOI 1)-infected autologous macrophages. Aerosol MAC infection of mice allows measurement of the effects of the BCG vaccine and clarithromycin. IFN-γ ELISPOT assays with live MAC (MOI 3) stimulation of splenocytes from mice immunized with BCG help identify differences between unimmunized mice and mice immunized with BCG. In conclusion, multiple assays are available for use to identify MAC-specific effector T cells, which will help in the development of new therapeutics or vaccines against pulmonary MAC. Full article

The anti-inflammatory/immunomodulatory effects of Teucrium montanum L. (TM), a plant distributed in the Mediterranean region, have been insufficiently examined. The effects of the TM ethanol extract were tested in a rat collagen-induced arthritis (CIA) model of rheumatoid arthritis. LC-MS was used for the phytochemical analysis of the TM extract. Dark Agouti rats were immunized with bovine type II collagen (CII) in incomplete Freund’s adjuvant for CIA, and treated with 100 or 200 mg/kg of TM extract daily via oral administration. Clinical and histopathological evaluations and a flow cytometric analysis of the phenotypic and functional characteristics of splenocytes and draining lymph node cells were performed. The cytokines in the paw tissue culture supernatants and anti-CII antibodies in serum were determined by ELISA. The TM extract, with the dominant components verbascoside and luteolin 7-O-rutinoside, reduced the arthritic score and ankle joint inflammation in CIA rats, promoted the antioxidant profile in serum, and lowered pro-inflammatory TNF-α, IL-6 and IL-1β production. It suppressed the activation status of CD11b+ cells by lowering CD86, MHCII and TLR-4 expression, and promoted the Th17/T regulatory cell (Tregs) balance towards Tregs. A lower frequency of B cells was accompanied by a lower level of anti-CII antibodies in treated rats. These findings imply the favorable effect of TM extract on the clinical presentation of CIA, suggesting its anti-inflammatory/immunomodulatory action and potential therapeutic effect. Full article

Recent research has focused on immunotherapy with no side effects as an innovative medical treatment for cancer. However, typical drugs for immunotherapy are very expensive. Here, we propose the use of immunoceramics that activate immune cells by contact with their surface. Previous studies demonstrated that polymers, including the phenylboronic acid group, could activate lymphocytes. This activation may be due to the interaction between the sugar chains in cells and the OH group in B(OH)₃ formed via the dissociation of the BO₂ group. We have clarified that boron-containing apatite (BAp) activated lymphocytes in vitro. In this study, we fabricated the ceramic surfaces using the CaO-P₂O₅-SiO₂-B₂O₃ system (CPSB ceramics) containing BAp as a main crystalline phase. The results of the in vitro evaluation indicated that killer T cells in splenocytes cocultured with the CPSB ceramics were more numerous than in splenocytes cocultured on a control surface. The results of the in vivo evaluation indicated that the CPSB ceramics significantly inhibited tumor growth when CD8-positive T cells were cultured on individual ceramics and subsequently injected into tumor-bearing mice. The present CPSB ceramics are expected to be a valuable biomaterial for immunotherapy. Full article

Background: Euglena gracilis (E. gracilis), a species of unicellular algae, can accumulate large amounts of β-1,3-glucan paramylon, a polysaccharide, in its cytoplasm and has recently attracted interest as a bioproduct due to its various health benefits. In this study, the immune-enhancing effect of E. gracilis powder (EP) was investigated in vitro and in vivo. Methods: In vitro, the production of NO and cytokines and the mechanism of the signaling pathway of β-1,3-glucan were identified in RAW264.7 cells. In vivo, cyclophosphamide-induced (CP-induced) immunosuppressed C57BL/6 female mice were orally administered with three different concentrations (100, 300, and 600 mg/kg) of EP daily. After 14 days, the organs and whole blood were collected from each animal for further study. Results: The weight loss of CP-treated mice was reversed by treatment with EP to levels comparable to those of control mice. In addition, the frequencies of NK1.1⁺, CD3⁺, CD4⁺, CD8⁺, and B220⁺ in immune cells isolated from the spleen were increased by EP treatment compared with water or RG. The secretion of TNF-α, IFN-γ, and IL-12 from splenocytes was also increased by EP treatment, as was the level of IgM in the serum of the mice. Finally, EP treatment specifically upregulated the expression of dectin-1 in the liver of CP-treated mice. Conclusions: E. gracilis could be a good candidate for a natural immune stimulator in the innate and adaptive response by secreting TNF-α, IFN-γ, and IL-12 through stimulating dectin-1 expression on the surface of immune cells. Full article

Allergic rhinitis (AR) is a common chronic disease that significantly impacts the quality of life. Lidocaine is known to have anti-inflammatory and immunomodulatory effects. This study evaluated the effect of lidocaine analogs in a Dermatophagoides pteronyssinus (DP)-induced AR mouse model. An AR model was developed using BALB/c mice via intraperitoneal sensitization with DP and intranasal challenge with DP. One hour before stimulation with DP, lidocaine analogs, EI137 and EI341 (at a dose of 0.5 or 5 ug/g), were administered intranasally. Nasal symptoms and serum total IgE, interleukin (IL)-4, IL-10, interferon (IFN)-γ, and tumor necrosis factor (TNF)-α levels were evaluated. Reverse-transcription polymerase chain reaction was used to determine IL-4, IL-10, and IFN-γ, as well as the expression of their mRNA transcription factors in the sinonasal mucosa. Histologic changes were evaluated using hematoxylin and eosin and periodic acid–Schiff staining. The DP-induced AR mouse model had increased serum levels of total IgE and cytokines. EI137 and EI341 significantly suppressed the levels of total IgE, IL-4, and TNF-α. Intranasal instillation of EI137 and EI341 significantly inhibited IL-4, IL-10, and IFN-γ mRNA expression, as well as inflammatory cells and mucus-producing goblet cells. Lidocaine analogs also suppressed DP-stimulated IL-4, IFN-γ, and IFN-γ production by splenocytes. Intranasal instillation of EI137 and EI341 exhibited anti-allergic and anti-inflammatory effects, influenced by Th1 and Th2 inflammatory cytokines. These lidocaine analogs suppressed DP-induced sinonasal mucosal inflammation, inflammatory cell infiltration, and mucus hypersecretion. Full article

Early-life nutrition significantly impacts vaccination efficacy in infants, whose immune response to vaccines is weaker compared to adults. This study investigated vaccination efficacy in female C57Bl/6JOlaHsd mice (6 weeks old) fed diets with 0.7% galacto-oligosaccharides (GOS)/long-chain fructo-oligosaccharides (lcFOS) (9:1), 0.3% human milk oligosaccharides (HMOS), or a combination (GFH) for 14 days prior to and during vaccination. Delayed-type hypersensitivity (DTH) was measured by assessing ear swelling following an intradermal challenge. Influvac-specific IgG1 and IgG2a levels were assessed using ELISAs, while splenic T and B lymphocytes were analyzed for frequency and activation via flow cytometry. Additionally, cytokine production was evaluated using murine splenocytes co-cultured with influenza-loaded dendritic cells. Mice on the GFH diet showed a significantly enhanced DTH response (p < 0.05), increased serological IgG1 levels, and a significant rise in memory B lymphocytes (CD27+ B220+ CD19+). GFH-fed mice also exhibited more activated splenic Th1 cells (CD69+ CXCR3+ CD4+) and higher IFN-γ production after ex vivo restimulation (p < 0.05). These findings suggest that GOS/lcFOS and HMOS, particularly in combination, enhance vaccine responses by improving memory B cells, IgG production, and Th1 cell activation, supporting the potential use of these prebiotics in infant formula for better early-life immune development. Full article

Senecavirus A (SVA) is a picornavirus that is endemic in swine, causing a vesicular disease clinically indistinguishable from other vesicular diseases, like foot-and-mouth disease. The widespread viral circulation, constant evolution, and economic losses caused to the swine industry emphasize the need for measures to control the agent. In this study, we evaluated the immunogenicity of a whole-virus-inactivated vaccine using a representative contemporary Brazilian SVA strain in Balb/ByJ mice. The animals were vaccinated with two doses by an intramuscular route. The humoral response induced by the vaccination was evaluated by an in-house ELISA assay for IgG detection. The cellular response was assessed by flow cytometry after in vitro SVA stimulation in splenocyte cultures from vaccinated and non-vaccinated groups. Protection against SVA was assessed in the experimental groups following an oral challenge with the homologous virus. The vaccination induced high levels of IgG antibodies and the proliferation of CD45R/B220⁺sIgM⁺, CD3e⁺CD69⁺, and CD3e⁺CD4⁺CD44⁺CD62L⁻ cells. These results indicate the immunogenicity and safety of the vaccine formulation in a murine model and the induction of humoral and cellular response against SVA. Full article

Effects of pulsed electromagnetic fields (PEMF) on immunological factors in a 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD) model were investigated. Hairless mice were randomly assigned to control, acetone and olive oil solution-treated (AOO), PEMF 15 Hz, PEMF 75 Hz, and sham groups (n = 5 each). AOO solution was used to dissolve DNCB. Both PEMF and sham groups were exposed to similar DNCB doses, causing similar AD symptoms. After AD induction for five weeks, only the PEMF groups were exposed to PEMF stimulations (15 Hz, 75 Hz, and 15 mT) inside the solenoid coil, for two weeks. In both groups, splenomegaly was observed, as AD was induced by hyperimmune reactions caused by DNCB sensitization. However, splenomegaly did not occur in the PEMF-exposed groups, and spleen weight decreased similarly to that of the control. Hence, the total splenocytes in the PEMF group were similar to those in the control group, whereas the sham group showed three times the number of splenocytes compared with the PEMF group. The serum immunoglobulin E levels did not significantly change in the PEMF group; however, they increased more than fourfold in the sham group. These results demonstrate that PEMF stimulation ameliorated the abnormal symptoms caused by hyperimmune reactions. Full article

Virus-like nanoparticles (VNP) are regarded as efficient vaccination platforms and have proven to be useful for the non-anaphylactogenic delivery of allergen-specific immunotherapy in preclinical models previously. Herein, we sought to determine the mode of VNP uptake by antigen presenting cells (APC). Accordingly, we screened a collection of substances known to inhibit different uptake pathways by APC. The human leukemia monocytic cell line THP-1 and the murine dendritic cell line DC 2.4 were examined for the uptake of fluorescently labelled VNP in the presence or absence of inhibitors. The inhibitory effect of candidate substances that blocked VNP uptake in APC lines was subsequently evaluated in studies with primary APC present in splenocyte and lung cell homogenates in vitro and upon intratracheal application of VNP in vivo. The uptake of allergen-specific VNP in vitro and in vivo was mainly observed by macrophages and CD103⁺ dendritic cells and was sensitive to inhibitors that block macropinocytosis, such as hyperosmolarity induced by sucrose or the polyphenol compound Rottlerin at low micromolar concentrations but not by other inhibitors. Also, T-cell proliferation induced by allergen-specific VNP was significantly reduced by both substances. In contrast, substances that stimulate macropinocytosis, such as Heparin and phorbol myristate acetate (PMA), increased VNP-uptake and may, thus, help modulate allergen-specific T-cell responses. We have identified macropinocytosis as the principal uptake mechanism of APC for allergen-specific VNP in vitro and in vivo, paving the way for further improvement of VNP-based therapies, especially those that can be used for tolerance induction in allergy, in the future. Full article

Chlamydia trachomatis (Ct) is the most common cause of bacterial sexually transmitted infections (STIs) worldwide. Ct infections are often asymptomatic in women, leading to severe reproductive tract sequelae. Development of a vaccine against Chlamydia is crucial. The Chlamydia major outer membrane protein (MOMP) is a prime vaccine antigen candidate, and it can elicit both neutralizing antibodies and protective CD4+ T cell responses. We have previously designed chimeric antigens composed of immunogenic variable regions (VDs) and conserved regions (CDs) of MOMP from Chlamydia muridarum (Cm) expressed into a carrier protein (PorB), and we have shown that these were protective in a mouse model of Cm respiratory infection. Here, we generated corresponding constructs based on MOMP from Ct serovar F. Preliminary structure analysis of the three antigens, PorB/VD1-3, PorB/VD1-4 and PorB/VD1-2-4, showed that they retained structure features consistent with those of PorB. The antigens induced robust humoral and cellular responses in mice with different genetic backgrounds. The antibodies were cross-reactive against Ct, but only anti-PorB/VD1-4 and anti-PorB/VD1-2-4 IgG antibodies were neutralizing, likely due to the antigen specificity. The cellular responses included proliferation in vitro and production of IFN-γ by splenocytes following Ct re-stimulation. Our results support further investigation of the PorB/VD antigens as potential protective candidates for a Chlamydia subunit vaccine. Full article

Marek’s disease virus (MDV) can cause severe immunosuppression in chickens. Our previous study showed that infection with very virulent plus (vv+) MDV strains of one-day-old commercial meat-type chickens possessing maternal antibodies against MDV resulted in severe depletion of splenocytes at 28–30 days of age. In the present study, we have investigated the effect of vv+MDV strain 686 on splenic immunophenotypes at 6, 20, and 30 days post-infection (dpi). Both live and dead cells were analyzed, and the data were statistically compared to the uninfected control. The results revealed a decrease in the total live cell population starting on day 20, primarily affecting B cells, CD8β+, and gamma delta (γδ) T cells, while the frequencies of both live and dead CD3+ and CD4+ T cells were increased. The MHC-I expression of CD3+ and CD4+ T cells was higher at 20 and 30 dpi, while the expression of MHC-II on these cells was downregulated at 6 dpi but was upregulated at 30 dpi. Collectively, these results suggest that maternal antibodies seem to delay the negative effects of vv+MDV on the splenic lymphoid populations, albeit being non-protective. Our results emphasize the importance of MD vaccination in vv+MDV endemic areas. Full article

β-Casomorphin-7 (BCM), a breakdown product of milk β-casein, exhibits opioid activity. Opioids are known to affect the immune system, but the effects of BCM on ulcerative colitis (UC) are not clear. We examined the effects of BCM on mucosal immunity using a mouse dextran sulfate sodium-induced colitis model and an in vitro CD8+ T cell activation model. Human UC patients were examined to reveal the relationship between CD10 and mucosal immunity. Combined treatment of the colitis model with thiorphan (TOP) inhibited BCM degradation by suppressing CD10 in the intestinal mucosa, activating mouse mucosal CD8, and suppressing CD4 and Treg. In the CD8+ T cell in vitro activation assay using mouse splenocytes, BCM inhibited the oxidative phosphorylation (OXPHOS) of CD8+ T cells and induced the glycolytic pathway, promoting their activation. Conversely, in a culture system, BCM suppressed OXPHOS and decreased defensin α production in IEC6 mouse intestinal epithelial cells. In the mouse model, BCM reduced defensin α and butyrate levels in the colonic mucosa. During the active phase of human ulcerative colitis, the downward regulation of ileal CD10 expression by CpG methylation of the gene promoter was observed, resulting in increased CD8 activation and decreased defensin α and butyrate levels. BCM is a potential aggravating factor for UC and should be considered in the design of dietary therapy. In addition, decreased CD10 expression may serve as an indicator of UC activity and recurrence, but further clinical studies are needed. Full article