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11 pages, 1020 KiB  
Communication
XBB.1.5 COVID-19 mRNA Vaccines Induce Inadequate Mucosal Immunity in Patients with Inflammatory Bowel Disease
by Simon Woelfel, Joel Dütschler, Daniel Junker, Marius König, Georg Leinenkugel, Claudia Krieger, Samuel Truniger, Annett Franke, Seraina Koller, Katline Metzger-Peter, Nicola Frei, STAR SIGN Study Investigators, Werner C. Albrich, Matthias Friedrich, Jan Hendrik Niess, Nicole Schneiderhan-Marra, Alex Dulovic, Wolfgang Korte, Justus J. Bürgi and Stephan Brand
Vaccines 2025, 13(7), 759; https://doi.org/10.3390/vaccines13070759 - 16 Jul 2025
Viewed by 352
Abstract
Background: Mucosal immunity plays a pivotal role in preventing infections with SARS-CoV-2. While COVID-19 mRNA vaccines induce robust systemic immune responses in patients with inflammatory bowel disease (IBD), little is known about their efficacy in the mucosal immune compartment. In this sub-investigation of [...] Read more.
Background: Mucosal immunity plays a pivotal role in preventing infections with SARS-CoV-2. While COVID-19 mRNA vaccines induce robust systemic immune responses in patients with inflammatory bowel disease (IBD), little is known about their efficacy in the mucosal immune compartment. In this sub-investigation of the ongoing STAR-SIGN study, we present the first analysis of mucosal immunity elicited by XBB.1.5 mRNA vaccines in immunocompromised patients with IBD. Methods: IgG and IgA antibodies targeting the receptor-binding domain of the SARS-CoV-2 JN.1 variant were quantified longitudinally in the saliva of IBD patients using the multiplex immunoassay MultiCoV-Ab. Antibody levels were quantified before and 2–4 weeks after vaccination with XBB.1.5 mRNA vaccines. All patients previously received three doses with original COVID-19 vaccines. Results: Mucosal IgG antibodies were readily induced by XBB.1.5 mRNA vaccines (p = 0.0013 comparing pre- and post-vaccination levels). However, mucosal IgA levels were comparable before and after vaccination (p = 0.8233). Consequently, mucosal IgG and IgA antibody levels correlated only moderately before and after immunization (pre-vaccination: r = 0.5294; p = 0.0239; post-vaccination: r = 0.4863; p = 0.0407). Contrary to a previous report in healthy individuals, vaccination did not induce serum IgA in patients with IBD (p = 0.5841 comparing pre- and post-vaccination levels). These data suggest that COVID-19 mRNA vaccines fail to elicit mucosal IgA in patients with IBD. Conclusions: Since mucosal IgA plays a pivotal role in infection control, the lack of IgA induction indicates that patients lack sufficient protection against SARS-CoV-2 infections which warrants the development of mucosal COVID-19 vaccines. Full article
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16 pages, 2024 KiB  
Article
Recovering Immunogenic Orthohantavirus puumalaense N Protein from Pellets of Recombinant Escherichia coli
by Natalya Andreeva, Ekaterina Martynova, Polina Elboeva, Milana Mansurova, Ilnur Salafutdinov, Aleksandr Aimaletdinov, Rafil Khairullin, Diksha Sharma, Manoj Baranwal, Sara Chandy, Dilbar Dalimova, Alisher Abdullaev, Mirakbar Yakubov, Albert Rizvanov, Svetlana Khaiboullina, Yuriy Davidyuk and Emmanuel Kabwe
Vaccines 2025, 13(7), 744; https://doi.org/10.3390/vaccines13070744 - 10 Jul 2025
Viewed by 431
Abstract
(1) Background: Hemorrhagic fever with renal syndrome (HFRS) remains a prevalent zoonosis in Eurasia. Orthohantavirus puumalaense (PUUV), carried by bank voles (Myodes glareolus), is the principal zoonotic pathogen of HFRS in this region. Despite ongoing efforts to develop effective drugs and [...] Read more.
(1) Background: Hemorrhagic fever with renal syndrome (HFRS) remains a prevalent zoonosis in Eurasia. Orthohantavirus puumalaense (PUUV), carried by bank voles (Myodes glareolus), is the principal zoonotic pathogen of HFRS in this region. Despite ongoing efforts to develop effective drugs and vaccines against PUUV, this challenge remains. (2) Aim: In this study, we aimed to express a large quantity of the PUUV recombinant N (rN) protein using E. coli. We also sought to develop a protocol for extracting the rN protein from pellets, solubilizing, and refolding it to restore its native form. This protocol is crucial for producing a large quantity of rN protein to develop vaccines and diagnostic tools for HFRS. (3) Methods; PUUV S segment open reading frame (ORF) coding for N protein was synthesized and cloned into the plasmid vector pET-28 (A+). The ORF was transformed, expressed and induced in BL21(DE3) pLysS E. coli strain. Subsequently, rN protein was purified using immobilized metal affinity and ion chromatography. Immune reactivity of rN protein was tested by employing in house and commercial VektoHanta-IgG kit ELISA methods (both in vitro and in vivo). (4) Results: The best conditions for scaling up the expression of the PUUV rN protein were an incubation temperature of 20 °C during a 20 h incubation period, followed by induction with 0.5 mM IPTG. The most significant protein yield was achieved when the pellets were incubated in denaturing buffer with 8M urea. The highest yield of refolded proteins was attained using non-denaturing buffer (50 mM Tris-HCl) supplemented with arginine. A final 50 μL of PUUV rN protein solution with a concentration of 7 mg/mL was recovered from 1 L of culture. The rN protein elicited an antibody response in vivo and reacted with serum taken from patients with HFRS by ELISA in vitro. (5) Conclusion: Therefore, the orthohantavirus N protein’s ability to elicit immune response in vivo suggests that it can be used to develop vaccines against PUUV after conducting in vitro and in vivo studies to ascertain neutralising antibodies. Full article
(This article belongs to the Special Issue Protein- and Subunit-Based Vaccines)
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20 pages, 1226 KiB  
Article
Transcriptomic Landscape of Paclitaxel-Induced Multidrug Resistance in 3D Cultures of Colon Cancer Cell Line DLD1
by Sandra Dragicevic, Jelena Dinic, Milena Ugrin, Marija Vidovic, Tamara Babic and Aleksandra Nikolic
Int. J. Mol. Sci. 2025, 26(14), 6580; https://doi.org/10.3390/ijms26146580 - 9 Jul 2025
Viewed by 241
Abstract
Multidrug resistance (MDR) significantly contributes to colon cancer recurrence, making it essential to understand its molecular basis for improved therapies. This study aimed to identify genes and pathways involved in resistance to standard chemotherapeutics by comparing transcriptome profiles of sensitive and paclitaxel-induced MDR [...] Read more.
Multidrug resistance (MDR) significantly contributes to colon cancer recurrence, making it essential to understand its molecular basis for improved therapies. This study aimed to identify genes and pathways involved in resistance to standard chemotherapeutics by comparing transcriptome profiles of sensitive and paclitaxel-induced MDR colonospheres. Cell viability and growth were assessed following treatment with 5-fluorouracil, oxaliplatin, irinotecan, bevacizumab, and cetuximab. Drug concentrations in culture media posttreatment were measured using high-performance liquid chromatography (HPLC). RNA sequencing (RNA-seq) of untreated sensitive and resistant colonospheres identified differentially expressed genes linked to baseline resistance. Our results confirmed cross-resistance in the resistant model, showing highest oxaliplatin tolerance may involve mechanisms beyond efflux. Transcriptome analysis highlighted upregulation of PIGR and activation of the ribosomal signaling pathway as potential resistance mediators. Notably, AKR1B10, a gene linked to chemotherapeutic detoxification, was overexpressed, whereas genes related to adhesion and membrane transport were downregulated. The overexpression of ribosomal protein genes suggests ribosome biogenesis plays a key role in acquired resistance. These findings suggest that targeting ribosome biogenesis and specific deregulated genes such as PIGR and AKR1B10 may offer promising strategies to overcome MDR in colon cancer. Full article
(This article belongs to the Special Issue Biological Hallmarks and Therapeutic Strategies in Cancer)
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11 pages, 662 KiB  
Article
Antibody Responses Following Primary Immunization with the Recombinant Herpes Zoster Vaccine (Shingrix®) in VZV Seronegative Immunocompromised Adults
by Andrea Wessely, Ines Zwazl, Melita Poturica, Lukas Weseslindtner, Michael Kundi, Ursula Wiedermann and Angelika Wagner
Vaccines 2025, 13(7), 737; https://doi.org/10.3390/vaccines13070737 - 8 Jul 2025
Viewed by 377
Abstract
Background: Immunocompromised patients are at risk of severe varicella zoster virus (VZV) infection and reactivation. In VZV seronegative immunocompromised persons, live-attenuated VZV vaccination is contraindicated, thus the recombinant herpes zoster vaccine (rHZV) remains a safe alternative, although an off-label application. Yet, data on [...] Read more.
Background: Immunocompromised patients are at risk of severe varicella zoster virus (VZV) infection and reactivation. In VZV seronegative immunocompromised persons, live-attenuated VZV vaccination is contraindicated, thus the recombinant herpes zoster vaccine (rHZV) remains a safe alternative, although an off-label application. Yet, data on the induction of a VZV-specific immune response in immunocompromised individuals with VZV-specific IgG below the assay’s cut-off are only available for patients after solid-organ transplantation (SOT). Methods: We retrospectively analyzed the induction of VZV-specific IgG antibody levels after vaccination with rHZV in immunocompromised patients who previously tested anti-VZV-IgG negative between March 2018 and January 2024. Results: Of 952 vaccinees screened that received 2 or 3 doses rHZV, depending on the underlying disease, 33 patients (median age 53.0; 51.5% female) with either hematopoietic stem cell transplantation (82%) or high-grade immunosuppressive treatment (18%) fulfilled the inclusion criteria. Upon rHZV vaccination, 88% (29/33) individuals mounted a significant antibody response exceeding the assay’s cut-off level for seropositivity (p < 0.0001). We detected higher geometric mean antibody concentrations after three compared to two doses. However, 12% remained below the assay’s cut-off level and were therefore considered non-responsive. Conclusions: The rHZV is immunogenic in VZV-seronegative immunocompromised individuals and therefore presents a valid option to induce seroconversion. However, antibody testing in high-risk groups should be considered to identify humoral non- and low responders. Full article
(This article belongs to the Special Issue Varicella and Zoster Vaccination)
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9 pages, 639 KiB  
Brief Report
Recombinant IgG1 Fc-μTP-L309C Ameliorates Endogenous Rheumatoid Arthritis in the K/BxN Mouse Model by Decreasing Th1 and Th17 Cells in the Spleen, Lymph Nodes and Joint and Increasing T Regulatory Cells and IL-10 in the Joint
by Bonnie J. B. Lewis, Selena Cen, Ruqayyah J. Almizraq, Beth Binnington, Rolf Spirig, Fabian Käsermann and Donald R. Branch
J. Clin. Med. 2025, 14(13), 4509; https://doi.org/10.3390/jcm14134509 - 25 Jun 2025
Viewed by 361
Abstract
Background/Objectives: Recombinant Fc proteins have been produced that have a protective effect in mouse models of arthritis, such as the K/BxN rheumatoid arthritis model. We have previously shown that a recombinant human IgG1 Fc with a point mutation at position 309, replacing a [...] Read more.
Background/Objectives: Recombinant Fc proteins have been produced that have a protective effect in mouse models of arthritis, such as the K/BxN rheumatoid arthritis model. We have previously shown that a recombinant human IgG1 Fc with a point mutation at position 309, replacing a leucine with a cysteine, fused to the human IgM tailpiece to form a human IgG1 Fc hexamer, rFc-µTP-L309C, effectively prevents neutrophil infiltration into the joints and ameliorates arthritis in the K/BxN serum transfer model and in the endogenous chronic arthritis K/BxN model. We have now investigated the effect of rFc-µTP-L309C on T-cells in the K/BxN chronic arthritis mouse model. Methods: PBMCs were isolated from the spleen, lymph nodes and joint synovial fluid from K/BxN mice having severe chronic arthritis that had been treated with 200 mg/kg rFc-µTP-L309C or human serum albumin (HSA). Flow cytometry was used to isolate the activated CD4+CD44+ T-cells and T-regulatory cells (Tregs). Intracellular staining was used to identify Th1 and Th17 T-cell subsets, and CD4+CD25+FoxP3+ Tregs. ELISA was used to measure levels of IL-10 and TGF-β in synovial fluid. Results: We find that amelioration of the arthritis occurs after treatment with rFc-µTP-L309C and results in a decrease in Th1 cells’ production of IFNγ and Th17 cells’ production of IL-17. Amelioration also results in decreased production of GM-CSF. Moreover, amelioration results in increased Tregs and IL-10 production in the synovial fluid. Conclusions: rFc-µTP-L309C reduces the inflammatory T-cells and increases the regulatory anti-inflammatory T-cells in the chronic arthritis K/BxN mouse model. This effect explains, in part, the ability of rFc-µTP-L309C to ameliorate the arthritis and reduce damage on the articular cartilage of K/BxN mice. Full article
(This article belongs to the Special Issue Updates on Rheumatoid Arthritis: From Diagnosis to Treatment)
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21 pages, 1238 KiB  
Article
An In-House ELISA for Anti-Porcine Circovirus Type 2d (PCV2d) IgG: Analytical Validation and Serological Correlation
by Gyeong-Seo Park, Byoung Joo Seo, Woo Ju Kwon, Yeong Lee Seok, Hyo Jeong Lee, Sung Ho Lee, Minju Kim, MinChul Lee, Chanhee Chae and Chonghan Kim
Vaccines 2025, 13(6), 657; https://doi.org/10.3390/vaccines13060657 - 19 Jun 2025
Viewed by 568
Abstract
Background/Objectives: Porcine circovirus type 2d (PCV2d) is the predominant genotype associated with porcine circovirus-associated disease (PCVAD), leading to significant economic losses. In South Korea, current vaccine lot-release testing relies on a T/C-ratio-based guinea pig assay, which lacks scientific justification and methodological robustness. [...] Read more.
Background/Objectives: Porcine circovirus type 2d (PCV2d) is the predominant genotype associated with porcine circovirus-associated disease (PCVAD), leading to significant economic losses. In South Korea, current vaccine lot-release testing relies on a T/C-ratio-based guinea pig assay, which lacks scientific justification and methodological robustness. This study aimed to develop and validate a statistically defined in-house ELISA using rabbit-derived polyclonal antibodies against PCV2d for the standardized evaluation of immunogenicity. Methods: Polyclonal IgG was generated by immunizing a rabbit with inactivated PCV2d, and it was purified through Protein A chromatography. Guinea pigs (n = 18) were immunized with IMMUNIS® DMVac, an inactivated PCV2d vaccine candidate developed by WOOGENE B&G, at different doses. In-house ELISA parameters were optimized (antigen coating, blocking agent, and substrate incubation), and analytical performance was evaluated by ROC, linearity, reproducibility, and specificity. Sera from guinea pigs and pigs were analyzed under validated conditions. Results: The optimal performance was achieved using 105 genomic copies/mL of the antigen coating and a 5% BSA blocking agent. The assay showed strong diagnostic accuracy (AUC = 0.97), reproducibility (CVs < 5%), and linearity (R2 = 0.9890). Specificity tests with PCV2a, PCV2b, and PRRSV showed minimal cross-reactivity (<7%). The cross-species comparison revealed a positive correlation (R2 = 0.1815) and acceptable agreement (bias = −0.21) between guinea pig and porcine sera. The validated cut-off (S/P = 0.4) enabled accurate classification across both species and aligned well with commercial kits. Conclusions: The in-house ELISA offers a robust, reproducible, and scientifically validated platform for immunogenicity verification, supporting its application in Korea’s national lot-release system. Homologous competition assays with PCV2d are planned to further confirm antigen specificity. Full article
(This article belongs to the Special Issue Animal Diseases: Immune Response and Vaccines)
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14 pages, 940 KiB  
Article
The Influence of Genotype and Seasonality on the Sow Colostrum Quality and Immunoglobulin G Content
by Kristina Gvozdanović, Vice Čuljak, Vladimir Margeta, Ivona Djurkin Kušec, Boris Antunović, Dalida Galović and Goran Kušec
Animals 2025, 15(12), 1802; https://doi.org/10.3390/ani15121802 - 18 Jun 2025
Viewed by 372
Abstract
The aim of this study was to investigate the influence of genotype (GT) and seasonality (NP) on the quality parameters of sow colostrum and evaluate the efficiency of the radial immunodiffusion (RID) analysis and the Brix refractometer in determining the IgG concentration. This [...] Read more.
The aim of this study was to investigate the influence of genotype (GT) and seasonality (NP) on the quality parameters of sow colostrum and evaluate the efficiency of the radial immunodiffusion (RID) analysis and the Brix refractometer in determining the IgG concentration. This study was conducted on 240 sows that originated from two genotypes, namely GT1 (TOPIGS, n = 120) and GT2 (Pig Improvement Company, n = 120), during the three farrowing periods: the winter farrowing period (WNP, n = 80), the summer farrowing period (SMP, n = 80) and the spring farrowing period (SSP, n = 80). The significant interaction effect was observed for protein (p < 0.0001), lactose (p < 0.05) and non-fat solids (SNT) (p < 0.001). At the same time, the interaction effect influenced the IgG concentration measured with the Brix refractometer (p < 0.0001) and RID (p < 0.0001). Pearson’s correlation coefficient showed that Brix percentage was positively correlated with RID results (r = 0.52, p < 0.0001), while the Bland–Altman plots indicated a mean bias of −1.93. Partial eta-squared analysis (η2) showed that the genotype explained the largest proportion of variance in fat content (η2 = 0.136) and IgG concentration (η2 = 0.164), while interaction effects were largest for protein (η2 = 0.072). The results of this study show that genotype and seasonality influence sow colostrum quality, which indicates the importance of genotype−seasonality interactions in breeding programs for optimizing the colostrum quality and piglet survival. Full article
(This article belongs to the Special Issue Genetic Research for Improving Livestock Heat Stress Resistance)
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11 pages, 1047 KiB  
Brief Report
Light Chain Isotype and Antibody-Specificity Impact on Virus Neutralization
by Lin Sun, Roman Palt, Georg Schütz, Esther Föderl-Höbenreich, Laura Brod, Antonia Hermle, Anja Lux, Herta Steinkellner and Somanath Kallolimath
Antibodies 2025, 14(2), 50; https://doi.org/10.3390/antib14020050 - 17 Jun 2025
Viewed by 400
Abstract
Therapeutic antibodies with lambda light chains (λ-Abs) are underrepresented compared to kappa light chains (κ-Abs). Here, we evaluated two SARS-CoV-2-specific monoclonal antibodies (mAbs) that exhibit high (P5C3) and low (H4) antigen binding as κ and λ variants. mAbs expressed in glycoengineered Nicotiana benthamiana [...] Read more.
Therapeutic antibodies with lambda light chains (λ-Abs) are underrepresented compared to kappa light chains (κ-Abs). Here, we evaluated two SARS-CoV-2-specific monoclonal antibodies (mAbs) that exhibit high (P5C3) and low (H4) antigen binding as κ and λ variants. mAbs expressed in glycoengineered Nicotiana benthamiana did not show differences in expression levels, glycosylation, and antigen binding, while κ-Abs exhibited slightly increased thermodynamic stability over λ-Abs. SARS-CoV-2 neutralization and IgG-FcγR immune complex studies revealed increased activities of H4 IgG1κ compared to H4 IgG1λ, with no differences observed between P5C3 variants. Our results indicate that constant light chain variability and Ab specificity contribute to Ab features, a fact that should be considered in engineering therapeutics. Full article
(This article belongs to the Section Antibody Discovery and Engineering)
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19 pages, 3312 KiB  
Article
Integrated Plasma and Tumor Proteomics of Nasopharyngeal Carcinoma in a Moroccan Cohort
by Ayman Reffai, Michelle Hori, Ravali Adusumilli, Abel Bermudez, Houssam Haddad, Nezha Tawfiq, Sharon Pitteri, Mohcine Bennani Mechita and Parag Mallick
Int. J. Mol. Sci. 2025, 26(12), 5771; https://doi.org/10.3390/ijms26125771 - 16 Jun 2025
Viewed by 448
Abstract
Nasopharyngeal carcinoma (NPC) is a multifactorial disease mainly affecting the Southeast Asian and North African populations. Critically, there is a dearth of available circulating biomarkers for NPC. Additionally, as of this writing, there have been no prior plasma proteomics studies on NPC in [...] Read more.
Nasopharyngeal carcinoma (NPC) is a multifactorial disease mainly affecting the Southeast Asian and North African populations. Critically, there is a dearth of available circulating biomarkers for NPC. Additionally, as of this writing, there have been no prior plasma proteomics studies on NPC in the Moroccan population. Accordingly, there has been no integrated analysis of tumor and plasma for NPC in the Moroccan sub-population. Label-free proteomics analysis was conducted on 25 samples of Moroccan origin (10 NPC samples and 15 healthy control samples). Each sample was depleted of albumin, fractionated into eight fractions, and then analyzed using Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS). A total of 291 proteins and 2702 unique peptides were identified across all samples. In total, 16 proteins were differentially expressed (DEPs) between NPC cases and healthy individuals. Of these, three showed prognostic significance, while four demonstrated diagnostic potential. A pathway analysis showed significantly enriched terms related to the immune response and chronic inflammation, revealing acute-phase proteins as differentially expressed. The investigation of patients with early and advanced stages of NPC revealed two DEPs, while four additional DEPs were identified across the three defined clusters of NPC. Across all comparisons, DEPs, such as H2A, IGHG2, SERPINA3, SAA1, CRP, PIGR, and APOA2, have shown potential as biomarkers for NPC, with several being identified for the first time. We additionally compared the plasma proteomic profile of NPC with the tumor proteomic profile, highlighting that deeper proteomics analysis of plasma may be required to quantify additional putative biomarkers that may be shed from the tumor into the blood. Our research presents the first plasma proteomic profile of NPC in Morocco and North Africa, identifying proteins that might ultimately have diagnostic and prognostic potential. Full article
(This article belongs to the Special Issue Genomics and Proteomics of Cancer)
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43 pages, 4992 KiB  
Article
Restorative Effects of Synbiotics on Colonic Ultrastructure and Oxidative Stress in Dogs with Chronic Enteropathy
by Dipak Kumar Sahoo, Tracey Stewart, Emily M. Lindgreen, Bhakti Patel, Ashish Patel, Jigneshkumar N. Trivedi, Valerie Parker, Adam J. Rudinsky, Jenessa A. Winston, Agnes Bourgois-Mochel, Jonathan P. Mochel, Karin Allenspach, Romy M. Heilmann and Albert E. Jergens
Antioxidants 2025, 14(6), 727; https://doi.org/10.3390/antiox14060727 - 13 Jun 2025
Viewed by 2369
Abstract
Synbiotics can be used to reduce intestinal inflammation and mitigate dysbiosis in dogs with chronic inflammatory enteropathy (CIE). Prior research has not assessed the colonic mucosal ultrastructure of dogs with active CIE treated with synbiotics, nor has it determined a possible association between [...] Read more.
Synbiotics can be used to reduce intestinal inflammation and mitigate dysbiosis in dogs with chronic inflammatory enteropathy (CIE). Prior research has not assessed the colonic mucosal ultrastructure of dogs with active CIE treated with synbiotics, nor has it determined a possible association between morphologic injury and signaling pathways. Twenty client-owned dogs diagnosed with CIE were randomized to receive either a hydrolyzed diet (placebo; PL) or a hydrolyzed diet supplemented with synbiotic-IgY (SYN) for 6 weeks. Endoscopic biopsies of the colon were obtained for histopathologic, ultrastructural, and molecular analyses and were compared before and after treatment. Using transmission electron microscopy (TEM), an analysis of the ultrastructural alterations in microvilli length (MVL), mitochondria (MITO), and rough endoplasmic reticulum (ER) was compared between treatment groups. To explore potential signaling pathways that might modulate MITO and ER stress, a transcriptomic analysis was also performed. The degree of mucosal ultrastructural pathology differed among individual dogs before and after treatment. Morphologic alterations in enterocytes, MVL, MITO, and ER were detected without significant differences between PL and SYN dogs prior to treatment. Notable changes in ultrastructural alterations were identified post-treatment, with SYN-treated dogs exhibiting significant improvement in MVL, MITO, and ER injury scores compared to PL-treated dogs. Transcriptomic profiling showed many pathways and key genes to be associated with MITO and ER injury. Multiple signaling pathways and their associated genes with protective effects, including fibroblast growth factor 2 (FGF2), fibroblast growth factor 7 (FGF7), fibroblast growth factor 10 (FGF10), synaptic Ras GTPase activating protein 1 (SynGAP1), RAS guanyl releasing protein 2 (RASGRP2), RAS guanyl releasing protein 3 (RASGRP3), thrombospondin 1 (THBS1), colony stimulating factor 1 (CSF1), colony stimulating factor 3 (CSF3), interleukin 21 receptor (IL21R), collagen type VI alpha 6 chain (COL6A6), ectodysplasin A receptor (EDAR), forkhead box P3 (FoxP3), follistatin (FST), gremlin 1 (GREM1), myocyte enhancer factor 2B (MEF2B), neuregulin 1 (NRG1), collagen type I alpha 1 chain (COL1A1), hepatocyte growth factor (HGF), 5-hydroxytryptamine receptor 7 (HTR7), and platelet derived growth factor receptor beta (PDGFR-β), were upregulated with SYN treatment. Differential gene expression was associated with improved MITO and ER ultrastructural integrity and a reduction in oxidative stress. Conversely, other genes, such as protein kinase cAMP-activated catalytic subunit beta (PRKACB), phospholipase A2 group XIIB (PLA2G12B), calmodulin 1 (CALM1), calmodulin 2 (CALM2), and interleukin-18 (IL18), which have harmful effects, were downregulated following SYN treatment. In dogs treated with PL, genes including PRKACB and CALM2 were upregulated, while other genes, such as FGF2, FGF10, SynGAP1, RASGRP2, RASGRP3, and IL21R, were downregulated. Dogs with CIE have colonic ultrastructural pathology at diagnosis, which improves following synbiotic treatment. Ultrastructural improvement is associated with an upregulation of protective genes and a downregulation of harmful genes that mediate their effects through multiple signaling pathways. Full article
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16 pages, 339 KiB  
Article
Humoral and T-Cell-Mediated Immunity Against Phlebotomus perniciosus Salivary Proteins in Dogs from a Leishmaniosis-Endemic Area
by Núria Balsells-Aguilar, Maria Magdalena Alcover, Marta Baxarias, Alejandra Álvarez-Fernández, Lourdes Alarcón, Petra Sumova, Petr Volf and Laia Solano-Gallego
Pathogens 2025, 14(6), 576; https://doi.org/10.3390/pathogens14060576 - 10 Jun 2025
Viewed by 484
Abstract
Compounds in sand fly saliva elicit specific immune responses that may play a role in the establishment of canine Leishmania infection. Although canine antibodies to anti-sand fly saliva antigens have been extensively studied, little is known about cellular immune responses against Phlebotomus perniciosus [...] Read more.
Compounds in sand fly saliva elicit specific immune responses that may play a role in the establishment of canine Leishmania infection. Although canine antibodies to anti-sand fly saliva antigens have been extensively studied, little is known about cellular immune responses against Phlebotomus perniciosus salivary proteins. This study aimed to explore humoral and T-cell-mediated immunity against P. perniciosus salivary proteins in dogs (n = 85) from Mallorca (Spain), a leishmaniosis-endemic area, and find correlations with demographic (age, sex, and breed) and parasite-specific immunological parameters. Anti-sand fly saliva IgG was examined using a P. perniciosus whole salivary gland homogenate (SGH) ELISA and recombinant salivary protein rSP03B ELISA. Interferon gamma (IFN-γ) release whole blood assays with L. infantum soluble antigen (LSA), SGH, and rSP03B were also performed. Positive correlations were found between IgG levels in the SGH and rSP03B tests and between concentrations of SGH IFN-γ and rSP03B IFN-γ. While concentrations of SGH IFN-γ and rSP03B IFN-γ were low and produced only by a minority of dogs (less than 20%), high levels and frequencies of LSA IFN-γ as well as anti-saliva IgG for SGH and rSP03B were detected in a majority of dogs (61% and 75%, respectively). LSA IFN-γ levels were positively correlated with age and Leishmania-specific antibodies. In conclusion, dogs from a leishmaniosis-endemic area presented high humoral immunity against P. perniciosus salivary proteins, but their cellular immunity to these proteins was low and less frequent. Full article
(This article belongs to the Special Issue Pets, Wildlife and Parasites—2nd Edition)
18 pages, 5566 KiB  
Article
Multi-Strain Probiotics Alleviate Food Allergy-Induced Neurobehavioral Abnormalities by Regulating Gut Microbiota and Metabolites
by Shouxun Hu, Luanluan Li, Chunyan Zhou, Yue Zhang and Xiaodan Yu
Nutrients 2025, 17(12), 1955; https://doi.org/10.3390/nu17121955 - 8 Jun 2025
Viewed by 832
Abstract
Background and aim: Neurobehavioral changes associated with food allergies have been reported, but the therapeutic effects of probiotics have not been fully explored. Our study aimed to investigate the impact of multi-strain probiotics on neurobehavioral outcomes and to elucidate the underlying mechanism via [...] Read more.
Background and aim: Neurobehavioral changes associated with food allergies have been reported, but the therapeutic effects of probiotics have not been fully explored. Our study aimed to investigate the impact of multi-strain probiotics on neurobehavioral outcomes and to elucidate the underlying mechanism via the microbiota-gut-brain axis. Methods: C57BL/6J Male mice were randomly divided into the following three groups: (1) control group; (2) OVA-sensitized group; (3) OVA-sensitized group treated with multi-strain probiotics (OVA + P). Anaphylactic reactions and behavioral abnormalities were assessed by histological, immunological, and behavioral analyses. To further elucidate the underlying mechanisms, the prefrontal cortex was collected for microglial morphological analysis, while serum and fecal samples were obtained for untargeted metabolomic profiling and 16S rDNA-based gut microbiota analysis, respectively. Results: Multi-strain probiotics significantly alleviated anaphylactic reactions in OVA-sensitized mice, as evidenced by reduced serum IgE levels, decreased Th2 cytokines, and reduced epithelial damage. Meanwhile, neurobehavioral symptoms were alleviated, including anxiety-like and depression-like behaviors, repetitive behaviors, social avoidance, and impaired attention. Mechanistically, probiotics administration suppressed production of inflammatory cytokines (TNF-α, IL-1β and IL-6) and inhibited activation of M1 microglia in the prefrontal cortex, which might contribute to neuron recovery. Furthermore, multi-omics analysis revealed that amino acid metabolism restoration in OVA + P mice, particularly carboxylic acids and derivatives, which was remarkably correlated with alterations in gut microbiota and behaviors related to FA. Conclusions: Gut microbiota and its amino acid metabolites mediate the therapeutic effects of multi-strain probiotics on FA-induced behavioral abnormalities. These effects occur alongside the suppression of neuroinflammation and microglial activation in the prefrontal cortex. Our findings highlight the neuroimmune regulatory role of the gut-microbiota-brain axis and support the potential use of probiotics as an intervention for FA-induced brain dysfunctions. Full article
(This article belongs to the Section Prebiotics and Probiotics)
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11 pages, 1940 KiB  
Case Report
Pituitary Apoplexy in a Child with Short Stature and Possible Recent SARS-CoV-2 Infection
by Carmen Gabriela Barbu, Luminita Nicoleta Cima, Marian Andrei, Simona Vasilache, Mihaela Țarnă, Ileana Olguta Rizea, Carmen Sorina Martin, Anca Elena Sîrbu and Simona Fica
Diagnostics 2025, 15(12), 1453; https://doi.org/10.3390/diagnostics15121453 - 7 Jun 2025
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Abstract
Background and Clinical Significance: Pituitary apoplexy is an extremely rare condition in children and adolescents with a rapid onset due to acute hemorrhage, infarction, or both in the pituitary gland. Most frequently, pituitary apoplexy is an asymptomatic or subclinical entity. Few cases of [...] Read more.
Background and Clinical Significance: Pituitary apoplexy is an extremely rare condition in children and adolescents with a rapid onset due to acute hemorrhage, infarction, or both in the pituitary gland. Most frequently, pituitary apoplexy is an asymptomatic or subclinical entity. Few cases of pituitary apoplexy with concurrent SARS-CoV-2 infection or COVID-19 vaccination have been reported. Case Presentation: We present the case of a 13-year-8-month-old boy who presented in our pediatric endocrinology department for the evaluation of short stature. He was previously diagnosed with secondary hypothyroidism and was treated with levothyroxine. At admission, clinical examination revealed a height of 141 cm (−2.68 SD/−2.4 SD corrected for mid-parental height), normal weight (60th centile), Tanner-stage G2P1, and delayed bone age. Basal IGF1 was normal, but the tests performed to assess the GH reserve confirmed the GH deficiency (peak GH value 3.11 ng/mL after clonidine/0.95 ng/mL after insulin). The brain MRI revealed a subacute pituitary hemorrhage. Thrombophilia and coagulopathies were excluded by further testing. Anti-SARS-CoV-2 (anti-S-protein IgG) antibodies (>200 BAU/mL) were compatible with COVID-19 infection, indicating a possible association between these two entities. At 3-month follow-up, physical examination showed a 3 cm height gain and advancing pubertal development (G4P2). Newer MRI found changes consistent with resolving hemorrhage. The patient was provided immediately with recombinant human GH and aromatase inhibitor therapy to maximize GH treatment response. During follow-up, the rGH dose was adjusted based on IGF1 values, and after 3 years and 10 months, rGH treatment was stopped, reaching a height of 172.3 cm (−0.51 SD) and surpassing the initial prediction of 164.5 cm. Conclusions: Pituitary apoplexy, an even rarer complication in the pediatric population, may be associated with SARS-CoV-2 infection. Further studies are necessary to better understand the intertwining of those conditions. Full article
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13 pages, 538 KiB  
Article
Gut Microbiota Differences in Infants with Cow-Milk-Induced Allergic Proctocolitis: A Comparative Cross-Sectional Study
by Zeliha Haytoglu, Dilek Ozcan and Derya Ufuk Altintas
Children 2025, 12(6), 734; https://doi.org/10.3390/children12060734 - 5 Jun 2025
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Abstract
Background: Cow-milk-induced allergic proctocolitis (CMIAP) is a non-IgE-mediated food hypersensitivity that often resolves spontaneously but may predispose infants to IgE-mediated allergies and eosinophilic gastrointestinal disorders. Understanding its pathophysiology is crucial for microbiota-based interventions. Methods: We enrolled 32 exclusively breastfed infants—16 with [...] Read more.
Background: Cow-milk-induced allergic proctocolitis (CMIAP) is a non-IgE-mediated food hypersensitivity that often resolves spontaneously but may predispose infants to IgE-mediated allergies and eosinophilic gastrointestinal disorders. Understanding its pathophysiology is crucial for microbiota-based interventions. Methods: We enrolled 32 exclusively breastfed infants—16 with confirmed cases of CMIAP and 16 age-matched healthy controls. The cohorts were sex-balanced (8 F/8 M), term-born (gestational age ± SD: 40 ± 1.2 vs. 39 ± 1.3 weeks), vaginally delivered, and sampled at a mean age of 2.0 ± 0.44 months (range 1.5–3.0) vs. 2.4 ± 0.66 months (range 1.5–3.5). Faecal samples underwent 16S rRNA gene sequencing on the Illumina NovaSeq platform, with diversity and differential abundance analyses. Results: The maternal dairy intake was similar (total dairy: 250 ± 80 vs. 240 ± 75 mL/day; yoghurt: 2.3 ± 1.0 vs. 2.5 ± 1.2 days/week; p = 0.72). Bray–Curtis dissimilarity assessments revealed distinct microbiota in infants with CMIAP. Infants with CMIAP had a lower abundance of Bifidobacterium (log2FC−2.27; q = 0.022; ANCOM-BC), Collinsella (−29.35; padj < 0.0001; DESeq2), and Limosilactobacillus (−8.01; padj = 0.0285; DESeq2; q < 0.0001; ANCOM-BC) compared with controls. In contrast, Hungatella (+24.99; padj < 0.0001; DESeq2), Veillonella (+4.73; padj = 0.0221; DESeq2), Citrobacter (+10.44; padj = 0.0124; DESeq2), and Ruminococcus gnavus (+2.69; q < 0.0001; ANCOM–BC) were more abundant in the CMIAP group. Conclusions: Infants with CMIAP exhibit gut dysbiosis, which is characterised by the depletion of beneficial commensals and the enrichment of potential pathogens, independent of maternal dairy intake. Further studies should establish whether these microbiota alterations are causal or consequential in CMIAP. Full article
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12 pages, 941 KiB  
Article
Prevalence of Various Systemic and Organ-Specific Autoimmune Markers in Addison’s Disease Patients Compared to Healthy Controls
by Aylin Feyzullova, Georgi Kirilov, Atanaska Elenkova, Dobromir Tanev, Krassimir Kalinov, Sabina Zacharieva and Ralitsa Robeva
J. Clin. Med. 2025, 14(11), 3951; https://doi.org/10.3390/jcm14113951 - 3 Jun 2025
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Abstract
Background: Addison’s disease (AD) is a rare disorder that often develops in the context of autoimmune polyglandular syndromes. However, the prevalence of rheumatological autoimmune diseases and corresponding autoimmune markers in AD is poorly investigated. Therefore, the present study aims to explore systemic and [...] Read more.
Background: Addison’s disease (AD) is a rare disorder that often develops in the context of autoimmune polyglandular syndromes. However, the prevalence of rheumatological autoimmune diseases and corresponding autoimmune markers in AD is poorly investigated. Therefore, the present study aims to explore systemic and organ-specific immune markers in a cohort of AD patients from a single tertiary endocrine center. Material and methods: In total, 43 adult AD patients and 31 controls were included in the study. 21-hydroxylase autoantibodies (21OHAb), glutamic acid decarboxylase autoantibodies (GADAbs), zinc transporter-8 autoantibodies (ZnT8Abs), antibodies against nuclear antigens (ANAs), autoantibodies against cyclic citrullinated peptides (CCPAbs), rheumatoid factors (RFs), IgG autoantibodies against cardiolipin (ACLAbs), and autoantibodies against beta-2-Glycoprotein I (β2-GPIAbs) were measured in all participants. Results: An increased prevalence of antibodies against RFs (27.91% vs. 0%, p < 0.001) and ANAs (13.95% vs. 0%, p = 0.037) was found in AD patients compared to controls. Moreover, the titers of 21-hydroxylase and RF antibodies correlated positively (r = +0.269, p = 0.020). The AD patients tended to show an increased prevalence of subthreshold ACL antibody reactivity compared to controls. All patients diagnosed with type 1 diabetes mellitus were GADAb- but not ZnT8Ab-positive. Conclusions: The results show an increased prevalence of ANA and RF positivity in AD patients compared to controls and a significant association between 21-OHAb and RF positivity. ZnT8Ab positivity was not typical for adult AD patients from our ethnic group, while GADAbs were an essential marker for autoimmune diabetes mellitus. Extensive studies in different ethnic groups are needed to establish the clinical significance of various immunological markers for AD comorbidity and the appropriate follow-up protocols for patients with different antibody positivity. Full article
(This article belongs to the Section Endocrinology & Metabolism)
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