The direct detection of small-molecule biomarkers in whole blood remains a significant challenge due to severe interferences from red blood cells (RBCs) and hemoglobin. To address this, we present the first paper-based chemiluminescence analytical device (PCAD) that integrates a core-shell mesoporous silica (CSMS) separation membrane with micellar encapsulation technology. The CSMS membrane acts as a physical barrier to exclude RBCs and simultaneously adsorbs proteins via its mesoporous structure, eliminating background noise without requiring complex pretreatment. Furthermore, we utilize aggregation-induced emission (AIE)-active 1,1,2,3,4,5-hexaphenylsilole (HPS) dyes encapsulated in bis[3,4,6-trichloro-2-(pentyloxycarbonyl)phenyl] oxalate (CPPO)-loaded micelles to overcome the solubility and aggregation limitations of traditional peroxyoxalate systems. Crucially, the micellar environment stabilizes the reactive intermediates, enabling the generation of long-lasting, high-intensity chemiluminescence signals ("glow-type" emission). This feature significantly enhances the signal-to-noise ratio (S/N), ensuring compatibility with low-cost, portable detection systems. As a proof of concept, the PCAD successfully achieves simultaneous determination of three critical metabolic indicators─glucose, uric acid, and xanthine─with low detection limits and excellent reproducibility. The entire process requires only a drop of whole blood without any prior separation steps and delivers results within 45 min using a portable reader. This work establishes a universal, user-friendly platform for on-site diagnosis of metabolic diseases, bridging the gap between laboratory precision and point-of-care convenience.
The Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathway is involved in the pathogenesis of a variety of inflammatory and autoimmune skin diseases, such as atopic dermatitis (AD), psoriasis, alopecia areata (AA), vitiligo, etc. Many cytokines are involved in the occurrence and development of these diseases through the JAK-STAT-mediated intracellular signaling. Small molecule JAK inhibitors (JAKi) have demonstrated substantial efficacy in the treatment of many diseases, especially in inflammatory skin diseases. For example, abrocitinib and upadacitinib have been approved for the treatment of moderate-to-severe AD, and baricitinib has been approved for the treatment of severe AA. Meanwhile, clinical and preclinical studies of various JAKis for other skin diseases are ongoing. However, the traditional classification of inflammatory skin diseases according to the T helper (Th) immune axis fails to distinguish between cytokine signals that are mechanistically dependent on the JAK-STAT pathway and those that are not, thus limiting its utility for guiding targeted therapy. Accordingly, we propose a functional refinement framework that integrates JAK-dependent cytokine modules into the established Th immune axis system. This article reviews the pathogenesis of "JAK-STAT pathway-associated skin diseases" and the available efficacy evidence of JAKi therapy, aiming to improve the mechanistic understanding and rational clinical application of JAKi and provide valuable references for clinical diagnosis and treatment.
Unconventional T cells, including gamma delta (γδ) T cells, mucosal-associated invariant T (MAIT) cells, and CD1d-restricted natural killer T (NKT) cells, comprise a unique component of the immune system. These cells play critical roles in host defense, immune regulation, and disease pathogenesis. In particular, invariant NKT (iNKT) cells have emerged as key immunological orchestrators that bridge innate and adaptive immunity by rapidly recognizing lipid antigens presented by CD1d molecules. This chapter provides a comprehensive overview of iNKT cell biology, including their development, phenotypic and functional heterogeneity, and activation mechanisms. It discusses the role of iNKT cells in pathological conditions in detail. Their ability to directly kill tumor cells, as well as their potential to orchestrate the immune response, opens up new possibilities for anti-cancer therapy. Their contribution to immune response regulation and tolerance highlights their critical role in infectious diseases and transplantation. The chapter summarizes current clinical approaches aimed at harnessing iNKT cells, including in vivo activation strategies, adoptive transfer of ex vivo-expanded cells, and developing chimeric antigen receptor (CAR)-engineered iNKT cells. These emerging therapies have advantages over conventional CAR-T approaches, such as reduced toxicity and the potential for allogeneic use. Finally, the chapter discusses the role of type II NKT cells and their cross-regulatory interactions with iNKT cells. Overall, CD1d-restricted NKT cells are a promising target for next-generation immunotherapies. However, further mechanistic and clinical studies are needed to realize their full therapeutic potential.
Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease characterized by autoantibody production and multi-organ involvement due to dysregulated immune responses. Ongoing research aims to understand the underlying mechanisms of this immune dysregulation, with particular interest in immune checkpoint molecules such as VISTA (encoded by the VSIR gene), which exerts context-dependent immunomodulatory effects. We analyzed a large-scale single-cell RNA-seq (scRNA-seq) dataset (GSE174188) comprising 261 samples (162 SLE cases, 99 controls) to investigate VSIR expression patterns across peripheral blood mononuclear cell (PBMC) subsets in SLE versus controls, using bioinformatics tools. Additionally, fresh PBMCs were isolated from SLE patients and healthy controls, and VSIR mRNA levels were quantified by RT-qPCR in treatment-naïve cases and those receiving Prednisolone, Hydroxychloroquine, or supplementary medications. Associations between VSIR expression and clinical/demographic parameters, including disease activity index, were also evaluated. scRNA-seq analysis revealed significantly upregulated VSIR expression in monocytes from SLE patients compared to healthy controls. In contrast, pseudobulk differential expression analysis of the entire PBMC population, corroborated by RT-qPCR on fresh samples, demonstrated downregulation of VSIR in SLE cases overall (log₂ fold change = -1.23). Notably, VSIR expression differed across treatment groups, with the lowest levels observed in treatment-naïve patients and higher levels in those receiving first-line therapies (prednisolone and hydroxychloroquine). Furthermore, VSIR expression exhibited a significant negative correlation with SLE disease activity index. This study demonstrates decreased VSIR expression in the whole PBMC population in SLE patients, suggesting a potential role in disease pathogenesis processes possibly through altered immune checkpoint regulation. Moreover, VSIR expression was associated with treatment status, with higher expression observed in patients receiving standard first-line therapies.
Cartilage oligomeric matrix protein (COMP) influences extracellular matrix remodeling. We investigated its clinical, prognostic, and immunomodulatory significance in colorectal cancer (CRC). COMP was quantified via ELISA in 107 paired CRC and normal tissues. Expression was correlated with clinicopathological features, mutational profiles, microsatellite instability (MSI), tumor-infiltrating lymphocytes (TILs), immune checkpoints, and multiplex cytokine networks. For transcriptomic validation, the FieldEffectCrc dataset was used for Gene Set Enrichment Analysis (GSEA), and The Cancer Genome Atlas (TCGA) CRC cohort for survival analysis. COMP was significantly upregulated in CRC tissues (p < 0.001) and correlated with advanced T, N, and overall pathological stages (all p < 0.05, tau = 0.18, 0.21, and 0.23, respectively). High COMP expression was linked to restricted immune infiltration (reduced stromal TILs, p < 0.05, tau = -0.23), elevated levels in microsatellite stable (MSS) compared to MSI tumors (p < 0.01), and correlated positively with immune exhaustion markers (T-cell immunoglobulin and mucin-domain containing-3 (TIM-3), galectin-9 (GAL9), sialic acid-binding Ig-like lectin 9 (SIGLEC9)). Transcriptomic data linked high COMP to worse disease-specific and progression-free survival, and enrichment in pro-tumorigenic pathways (epithelial-to-mesenchymal transition, angiogenesis, IL-6 signaling). COMP upregulation defines an immunosuppressive microenvironment in CRC, particularly in MSS tumors. It represents an important prognostic biomarker and potential therapeutic target for overcoming immunotherapy resistance.
Inflammatory bowel disease (IBD) burden is rising globally, yet only subsets of patients benefit from available therapies, underscoring the need for more precise molecular and cellular stratification. In the PREDICT study, we enrolled treatment-naïve pediatric patients with IBD, alongside disorders of gut-brain interaction (DGBI) controls and healthy donors, and profiled their intestinal and blood-derived T cells using single-cell RNA sequencing (scRNA-seq). Across 107 participants, we identify a unique population of cytotoxic CD4⁺ T cells (CD4 CTL) enriched in the inflamed gut of patients with Crohn's disease (CD) and ulcerative colitis. CD4 CTLs are clonally expanded and express cytotoxic effector molecules and IFNG , consistent with antigen-driven activation. Cell-cell interaction analyses implicate macrophage-derived IL-27 as the top candidate for CD4 CTL differentiation, and IL-27 blockade in a mouse model limits CD4 CTL formation. Notably, elevated CD4 CTL frequencies in gut and peripheral blood at diagnosis are associated with subsequent poor outcome of anti-TNF therapy in pediatric CD. Findings in our identification cohort are validated in an independent cohort and through reanalysis of published datasets. Importantly, we designed a simple flow cytometry panel to isolate blood CD4 + CXCR6 + CD27 - T cells, which displayed a CD4 CTL transcriptional phenotype. Together, our results link CD4 CTLs to anti-TNF nonresponse and support their potential as an early, blood-accessible biomarker for treatment stratification in pediatric CD.
Programmed cell death protein 1 (PD-1) has long been considered a central molecule in CD8⁺ T cell exhaustion and immunosuppression. However, recent studies have revealed that PD-1⁺CD8⁺ T cells are not a homogeneous population of terminally dysfunctional cells, but rather constitute key immune cells with significant heterogeneity and functional plasticity within tissue immune microenvironments. PD-1 signaling operates throughout multiple stages of CD8⁺ T cell biology, including thymic development, peripheral activation, chronic antigen stimulation, and tissue residency. By finely regulating T cell receptors (TCRs) signal strength, metabolic state, and transcriptional programs, it deeply participates in cell fate decisions while limiting immunopathology. In chronic infections and tumors, persistent antigen stimulation drives PD-1⁺CD8⁺ T cells to form an exhaustion lineage with a defined differentiation hierarchy, encompassing stem-like precursor cells, effector-like transitional cells, and terminally exhausted cells. PD-1 is not only a characteristic marker of this lineage but also a critical regulatory node through which immune checkpoint blockade therapy exerts its therapeutic effects. Furthermore, in contexts such as tissue-resident memory T cells (TRM), GZMK⁺CD8⁺ T cells, and other disease-associated microenvironments, sustained PD-1 expression often represents an adaptive functional regulatory state rather than mere functional inhibition. This review explores the multidimensional regulatory roles of PD-1 in CD8⁺ T cells, with a focus on elucidating the diverse functions and clinical significance of PD-1⁺CD8⁺ T cells in cancer, chronic infections, and autoimmune diseases.
γδT cells are a subset of innate lymphocytes that play an essential role in anti-infection immunity. However, the functional role of CD8αβ+ γδT cells, a distinct subset of γδT cells, remains poorly characterized during malaria infection. Female C57BL/6 mice were intraperitoneally injected with 1 × 106 Plasmodium yoelii-infected red blood cells (iRBCs). At 12 days post-infection (12 dpi, the peak of parasitemia), spleens were harvested and lymphocytes were isolated. Fluorescence-activated cell sorting (FACS) was performed to determine the frequency, phenotypic features and functional properties of CD8αβ+ γδT cells. Meanwhile, CD45+ lymphocytes were sorted for single-cell RNA sequencing (scRNA-seq) to compare differentially expressed genes between CD8αβ+ γδT and CD8αα+ γδT cell subsets. FACS analysis revealed a significant increase in CD8αβ+ γδT cells following P. yoelii infection. These cells exhibited elevated expression of activation-related molecules, enhanced effector functions, preferential polarization towards an IFN-γ-producing (γδT1) phenotype, and reduced expression of exhaustion markers. scRNA-seq further demonstrated that CD8αβ+ γδT cells upregulated genes involved in DNA replication and repair. Our findings indicate that CD8αβ+ γδT cells upregulate molecules associated with activation and function during P. yoelii infection, suggesting that they may play a more prominent role in host defence against malaria.
Immunodeficiency is a pathological state characterized by impaired functional integrity of the immune system, which contributes to the development of various diseases. Natural bioactive peptides are a promising option for improving immune function. This study examined the therapeutic effects and underlying mechanisms of Haematococcus pluvialis peptides (HPP) against CTX-induced immunodeficiency in mice. The results demonstrated that HPP increased bodyweight, immune organ indices, and blood cell count - white blood cells (WBC), red blood cells (RBC), platelets (PLT), hemoglobin (HGB), lymphocytes (Lym), and granulocytes (Gran), as well as serum cytokine levels (interferon-gamma (IFN-γ), interleukin-2 (IL-2), and immunoglobulin A (IgA) - in immunodeficient mice. Haematococcus pluvialis peptides improved the villus length and crypt depth of the small intestine and increased intestinal levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and secretory immunoglobulin A (SIgA). Colonic levels of tight junction proteins - zonula occludens-1 (ZO-1) and occludin - were up-regulated. Fecal microbiota analysis suggested that HPP promoted the enrichment of beneficial bacterial genera (Ligilactobacillus, norank_f_Muribaculaceae, Alistipes) and suppressed pathogenic bacteria (Escherichia-Shigella and Klebsiella). Gut microbial metabolites analysis showed that HPP altered various fecal metabolites involved in lipids and lipid-like molecules, organoheterocyclic compounds, phenylpropanoids and polyketides, as well as organic acids and their derivatives. Fecal microbiota transplantation (FMT) experiments also validated the decisive role of gut microbiota in the immunomodulatory function of HPP. These results offer novel insights into the protective efficacy and underlying mechanisms of HPP for alleviating immunodeficiency, establishing a robust theoretical basis for its application as a promising immunomodulatory agent. © 2026 Society of Chemical Industry.
Circulating monocyte trans-endothelial migration is critical for vascular diseases. The monocyte surface proteins, i.e., integrins and membrane receptors are central in this process. Previous studies demonstrate that ablation of the mitochondrial arginase-2 (ARG2) reduces monocyte/macrophage infiltration in cardiovascular disease. We further investigate whether ARG2 regulates integrin and surface receptor levels in monocytes and facilitates trans-endothelial migration, contributing to atherogenesis. For this purpose, human THP1 cells deficient in ARG2 gene (THP1ARG2-/-) were generated by CRISPR-U™-mediated genome engineering. Atherosclerotic Apoe-/-Arg2+/+ mice and Apoe-/-Arg2-/- mouse models are used. Proteomic profiling, cellular/molecular biology methods, and confocal microscopy were utilized for gene or protein expression analysis. As compared to the control THP1WT cells, the THP1ARG2-/- cells reveal decreased adhesion and trans-endothelial migratory activities towards chemoattractants in a transwell co-culture system, accompanied by lower levels of αL and α4 integrins, CD99 and PECAM1 (CD31) (surface molecules for trans-endothelial activity) and CCR2 (the chemoattractant receptor). The monocyte adhesion to endothelial cells is reduced by blocking LFA-1 (αLβ2) or VLA-4 (α4β1) integrin. Pro-inflammatory polarization of the THP1 cells with LPS does not affect the integrin and surface receptor levels, however, enhances release of several pro-inflammatory cytokines, which is reduced in THP1ARG2-/- cells due to reduced TLR4-ERK-NF-κB signaling. Conditioned medium from the LPS-primed THP1WT reveals higher capacity to enhance endothelial VCAM-1 and ICAM-1 levels than the THP1ARG2-/- cells, that is partly mediated by IL-1β. Moreover, ARG2-dependent TGF-β signaling was found to selectively regulate αL expression in monocytes. Finally, as compared to Apoe-/-Arg2+/+ mice, the ApoE-/-Arg2-/- mice show significantly decreased macrophage integrin, chemoattractant receptor levels, and atherosclerosis. Our study identifies ARG2 as a key regulator of monocyte-mediated vascular inflammation and atherogenesis by controlling integrin, chemoattractant receptor expression, and pro-inflammatory cytokine release through TLR4-ERK-NFκB signaling, highlighting ARG2 as a potential therapeutic target for vascular and chronic inflammatory diseases.
Poecilobdella manillensis Lesson is a well-recognized medicinal leech in traditional Chinese medicine and Guangxi Zhuang ethnic medicine. It has long been used to activate blood circulation and remove blood stasis for the treatment of ischemic stroke. Modern pharmacological research has verified its potent anticoagulant and anti-inflammatory activities. Current studies mainly focus on its polypeptide components that exert antithrombotic effects to improve cerebral ischemia, while the neuroprotective potential and related mechanisms of its small-molecule constituents remain largely unclear. This study aimed to investigate the therapeutic effects of the ethyl acetate extract (EA) of P. manillensis on cerebral ischemia-reperfusion injury and to clarify its underlying molecular mechanism. The chemical constituents of EA were identified by UPLC-Q-TOF-MS/MS. Network pharmacology and molecular docking were used to predict and verify core targets and pathways. Neuroprotective and anti-inflammatory effects of EA were evaluated in a rat MCAO/R model, OGD/R-injured SH-SY5Y cells, and LPS-stimulated BV2 cells, using histological staining, Western blot, immunohistochemistry, and RT-qPCR. Seven small-molecule components were identified in EA, and 314 overlapping targets related to ischemic stroke were screened. Network analysis showed that TLR4 was the core target, and the main enriched pathways included NF-κB, Toll-like receptor, apoptosis and TNF signaling pathways. Consistent with the predicted results, EA significantly reduced cerebral infarct volume and improved neurological deficits in MCAO/R rats, and inhibited neuronal apoptosis and microglial inflammation in vivo. In vitro, EA notably improved the survival of OGD/R-injured neurons and suppressed LPS-induced inflammatory responses in BV2 cells. Meanwhile, EA markedly downregulated the expression of TLR4/NF-κB and NLRP3 inflammasome-related molecules. The present study demonstrated that EA protects against cerebral ischemia-reperfusion injury by inhibiting neuronal apoptosis and TLR4/NF-κB-mediated neuroinflammation. These findings provide a scientific basis for the traditional clinical application of P. manillensis and suggest that EA could serve as a potential therapeutic candidate for ischemic stroke.
Sandhoff disease (SD) is a fatal lysosomal storage disorder caused by β-N-acetylhexosaminidase deficiency, resulting in GM2 ganglioside accumulation, severe neurodegeneration, and chronic neuroinflammation. While enzyme-restoring therapies, such as AAV gene transfer, effectively target the primary enzymatic deficit, neuroinflammation persists and contributes to disease progression, motivating the development of anti-inflammatory adjuncts alongside disease-modifying interventions. Extracellular vesicles released by mesenchymal stromal cells (MSC-EVs) are acellular lipid nanoparticles that contain immunomodulatory molecules and can cross physical barriers, without immunogenicity issues. These features make MSC-EVs promising candidates for anti-inflammatory therapeutics targeting neuropathology. The anti-inflammatory potency of MSC-EVs is increased by priming parent MSCs with pro-inflammatory cytokines, resulting in IFEVs, as referred to in this study. Here, we evaluated the in vitro anti-inflammatory effects of IFEVs in a feline model of SD. IFEVs reduced SD neuroinflammation, lowering IL-6, TNF-α, and IL-1β protein levels in SD neuronal-mixed glia, with concordant transcriptional downregulation of NF-κB/p65 and NLRP3 components and upregulation of arginase 1 mRNA after 48 h of treatment. In SD peripheral blood mononuclear cells, IFEVs significantly increased the proportion of regulatory T cells and the Treg/T-effector cell ratio without inducing cytotoxicity. Together, these findings demonstrate that IFEVs reduced neuroinflammation and altered blood-circulating T-cell populations in an in vitro SD model, supporting their further preclinical development as a complementary immunomodulatory therapy for SD.
MicroRNAs (miRNAs) are single stranded, short, non-coding, and highly conserved RNA molecules with around 22 nucleotides. They regulate gene expression at the co- and post-transcriptional levels. The accumulated research reports showed dysregulated miRNA expression in various female reproductive disorders most notably polycystic ovary syndrome (PCOS). PCOS is an endocrine disorder that affects women of reproductive age. The clinical manifestations of this disease are dominated by hyperandrogenism, polycystic ovaries, and chronic anovulation. Recently, evidence has confirmed abnormal expression of miRNAs in theca cells, granulosa cells, follicular fluid, blood, and serum of women with PCOS. Although some of these reports are controversial, the variability could be attributed to different analysis methods and small sample sizes. Overall, these findings point toward the role of miRNAs in the occurrence and progression of PCOS. However, the exact mechanism of its etiology largely remains unknown. This review takes us beyond the traditional localized dysregulation to explore the regulatory role of the gut microbiota-miRNA axis, exosomal miRNAs and the complexity of competitive endogenous RNA (ceRNA) networks where long non coding RNAs (lncRNAs) and circular RNAs (circRNAs) act as sponges to modulate miRNA activity. miRNAs can be used as pharmacogenomic biomarkers to predict the therapeutic responses to clinical interventions through the integration of machine learning algorithms for enhanced diagnostic accuracy. While research investigations are predominantly based on in vitro and clinical samples, we emphasize the need for more in vivo research to decode the precise etiology of PCOS. This review provides comprehensive update on the functional and clinical potential of miRNAs as non-invasive biomarkers and therapeutic targets for the management of PCOS.
Mitochondrial transcription factor A (TFAM) is a key regulator of mitochondrial DNA transcription and replication. T cell-specific TFAM-deficient mice are immunocompromised, often succumbing to viral infection, and their T cells are unresponsive to T-cell antigen receptor (TCR) stimulation, suggesting that TFAM-mediated mitochondrial activity regulates T-cell activity, such as effector function and memory formation. In contrast to the attenuation of immune response, TFAM deficiency may also induce inflammatory responses, raising the possibility that TFAM deficiency results in inflammation-mediated autoimmune responses. Thus, besides regulating T-cell function, TFAM plays important roles in autoimmune responses. However, its role in autoimmune diseases remains uncertain. We aimed to investigate the role of TFAM in autoimmune diseases using T cell-specific TFAM-deficient mice. We detected anti-double-strand (ds) DNA antibody, and interferon alpha (IFNα) and IFNγ in the serum of TFAMfl/fl CD4Cre mice after 30 weeks of age. Mononuclear cell infiltration was observed in the kidneys. TFAM-deficient T cells exhibited leakage of mitochondrial DNA into the cytoplasm. Cytoplasmic mitochondrial DNA was associated with activation of TANK-binding kinase 1 (TBK1) and IFN regulatory factor 3 (IRF3)-the downstream molecules of the nucleic acid sensor cyclic guanosine monophosphate-adenosine monophosphate synthase/stimulator of IFN genes (cGAS/STING) machinery. mRNA expression of type I IFN genes was elevated in T cells from TFAMfl/fl CD4Cre mice. The suppressive function of Foxp3+ regulatory T (Treg) cells, which play a major role in establishment of peripheral tolerance, was reduced in the absence of TFAM. Our findings suggest that T cells in TFAM-deficient conditions may contribute to immune instability and autoimmune responses by inducing type I IFN-mediated inflammatory responses.
Selenium (Se) is an essential micronutrient that participates in redox regulation and cellular homeostasis. Selenium-containing ionic liquids (ILsSe) have emerged as hybrid compounds combining the properties of organoselenium molecules with the tunable features of ionic liquids. In this study, we evaluated the cytotoxicity, interaction with mercury (Hg2+), pharmacokinetic profile, and antiproliferative activity of three ILsSe (C1, C2, and C3). Human peripheral blood mononuclear cells (PBMCs) were used to assess cytotoxicity, showing that ILsSe were generally well tolerated at low micromolar concentrations, although their effects were dependent on exposure time and dose. The interaction of ILsSe with Hg2+ was investigated using a thiol oxidase assay, with diphenyl diselenide (DPDS) as a reference compound. The results indicate that ILsSe can interact with Hg2+, likely through the formation of Se-Hg species. In parallel, in silico absorption, distribution, metabolism, excretion, and toxicity (ADMET) analysis suggested that these compounds have physicochemical properties compatible with central nervous system exposure, including predicted blood-brain barrier permeability. In U87-MG glioblastoma cells, compound C3 exhibited concentration and time-dependent antiproliferative effects at low micromolar concentrations. These effects were associated with alterations in redox homeostasis, modulation of cell cycle progression, and induction of late-stage cell death. In general, ILsSe are biologically active compounds whose effects depend on dose and exposure time, warranting further investigation to clarify their toxicological profile and mechanisms of action. This study highlights the importance and biological potential of selenium-containing compounds.
Chagas disease (ChD) is a neglected parasitic infection for which the first-choice trypanocidal drug benznidazole (BZ) has limited efficacy and marked systemic toxicity. Considering the need and urgency for more effective and safe drugs and the emergence of BZ-resistent parasites, we integrated molecular synthesis, physicochemical modeling, in vitro and in vivo strategies to investigate the antiparasitic, anti-inflammatory and cardioprotective relevance of a new nitroimidazole-based molecule 1-(2-(2-methoxy-5-nitro-4-propylphenoxy)ethyl)-2-methyl-5-nitro-1H-imidazole (MNPI) for ChD treatment. After chemical synthesis and molecular characterization, Trypanosoma cruzi (epimastigotes and trypomastigotes), H9c2 cardiomyocytes, and T. cruzi-infected mice were used to determine MNPI selectivity, hepatotoxicity, antiparasitic, anti-inflammatory and cardioprotective potential. BZ was used as the reference antiparasitic control. MNPI presented physicochemical characteristics compatible with orally bioactive drugs, potent antiparasitic activity, anti-infective efficacy in H9c2 cells, high selectivity indices (13.35 μM and 72.28 μM) against epimastigotes and trypomastigotes, and low cytotoxicity (CC50 351.3 ± 20.0) in vitro, outperforming BZ selectivity (35.24 μM and 149.20 μM) and cytotoxicity (CC50 768.4 ± 86.4). Alone, MNPI reduced blood parasitism, heart inflammation, cytokine (TNF-α, IFN-γ) and anti-T. cruzi antibody (IgG) levels without inducing hepatotoxicity in T. cruzi-infected mice. MNPI combined with BZ improved humoral response and blood parasitism control and prevented mortality in T. cruzi-infected mice. Our findings indicate that MNPI combined the pharmacological effects of its precursor molecules, acting as a promising antiparasitic, anti-inflammatory, and cardioprotective compound in T. cruzi infection. By interacting with BZ to induce superior anti-inflammatory and cardioprotective effects, MNPI may contribute to the development of innovative chemotherapeutic strategies for ChD treatment.
Emergent helminthiases are increasingly impacting global health in both humans and animals, especially given the limited efficacy of existing drugs against these infections. Neuroangiostrongyliasis, an eosinophilic meningitis caused by Angiostrongylus cantonensis, currently lacks effective treatment, highlighting the need for novel anthelmintics. We previously identified cinnamoyl-benzylpiperazine, a simplified analogue of cinnarizine, as an effective anthelmintic agent against first (L1) and third-stage (L3) larvae of A. cantonensis in vitro. In the present work, structural modifications on the active prototype cinnamoyl-benzylpiperazine were performed, prioritizing the improvement of solubility and the provision of balanced physicochemical properties compatible with blood-brain barrier permeability, alongside anthelmintic activity. A set of 31 compounds divided into two series (I-cinnamoyl and II-benzoyl) was synthesized and tested against L1 and L3 larvae, yielding EC50 values ranging from 4.1 to 27.6 μM. SAR analyses revealed that the activity of set I is strongly associated with balanced electronic density in both the cinnamamide and amine regions (described by ionization potential descriptors), whereas modifications in the charge distribution of the molecules (indicated by topological charge descriptors) appear to determine the anthelmintic activity of set II. None of the compounds displayed significant toxicity to HaCaT mammalian cells (up to 200 μM) or Caenorhabditis elegans worms (up to 1000 μM), denoting specific activity against A. cantonensis. The balanced polarity of compound 4a-I (EC50 L1 4.7 μM; L3 10.2 μM) and the localized charge density provided by the methoxy group in compounds 1c-II (EC50 L1 5.5 μM; L3 10.9 μM) and 5c-II (EC50 L1 4.9 μM; L3 11.9 μM) seem important for interacting with the putative target in the helminth. Collectively, the substituents in these molecules provided improved drug-likeness over the previous set of compounds, and represent noteworthy derivatives for further investigation against A. cantonensis in vitro.
Bullous pemphigoid (BP) is an autoimmune blistering disease with an increasing incidence in recent years; however, the underlying immune regulatory mechanisms remain largely unclear. As a critical signalling hub linking innate and adaptive immunity, stimulator of interferon genes (STING) has recently been implicated in the pathogenesis of various autoimmune diseases and may regulate tissue inflammation and immune homeostasis through modulation of CD4+ T cell responses. In this study, we found that STING expression was significantly increased in lesional skin tissues from patients with BP compared with healthy controls. Transcriptomic analysis further revealed that differentially expressed genes in peripheral blood CD4+ T cells from BP patients were primarily enriched in the JAK-STAT signalling pathway, T cell activation and differentiation, and type I interferon (IFN-I)-related pathways. Pharmacological inhibition of STING markedly attenuated the aberrant activation of these signalling pathways. Moreover, qRT-PCR analysis confirmed that the mRNA levels of STING1, JAK1, and CXCR5 were significantly elevated in BP patients, whereas treatment with the STING inhibitor C176 suppressed the expression of these molecules. Collectively, our findings suggest that STING may contribute to BP immunopathogenesis by regulating the JAK-STAT signalling axis and promoting abnormal CD4+ T cell activation and differentiation, providing new insights into the molecular mechanisms underlying BP and identifying potential therapeutic targets.
Ataxia-Telangiectasia (A-T) is a multisystem disorder caused by loss of A-T mutated (ATM) protein activity, characterized clinically by immunodeficiency and cerebellar ataxia. ATM is a master regulator of DNA damage responses and loss of ATM function is accompanied by persistent activation of PARP1 leading to depletion of intracellular NAD+ and dysfunction of a series of cellular signalling pathways dependent on NAD+, providing a mechanistic rationale for NAD+ augmentation therapy. We performed a clinical trial of NAD+ augmentation with nicotinamide riboside (NR) over 24 months in A-T patients where we observed improved coordination and eye movements in A-T patients. Here, by using peripheral blood mononuclear cells, we performed longitudinal transcriptome profiling to define molecular signatures of A-T and to assess pathway-level responses to NR supplementation. A-T patients exhibited reproducible transcriptomic alterations involving immune, vascular, and inflammatory pathways. NAD+ augmentation was associated with suppression of interferon response genes and modulation of networks correlated with neurological improvement. These findings establish systemic molecular signatures of A-T and identify potential blood-based biomarkers that reflect disease processes and therapeutic response, supporting the use of NAD+ augmentation as a disease-modifying strategy in A-T by dampening interferon signalling.
Diabetic retinopathy (DR), a leading cause of blindness, is driven by hyperglycemia-induced neurovascular damage. Emerging evidence indicates that PANoptosis, an integrated inflammatory programmed cell death modality encompassing apoptosis, pyroptosis, and necroptosis, participates in the progression of diabetic retinal damage; however, the supporting evidence varies substantially across clinical specimens, diabetic animal models, high-glucose cultured retinal cells, and non-DR inflammatory disease models. This review systematically summarizes the latest advances in PANoptosis-associated mechanisms underlying DR pathogenesis, focusing on PANoptosome signaling networks, non-coding RNA-mediated regulation, and immune-metabolic crosstalk. We outline promising candidate biomarkers including PANoptosis-related gene signatures and inflammatory cell death molecules, and critically evaluate multiple translational therapeutic strategies covering small-molecule inhibitors, gene intervention, and nanomedicine delivery. Importantly, we further address context-dependent dual roles of PANoptosis, potential safety risks of non-selective PANoptosis blockade, challenges in blood-retinal barrier penetration, systemic immune side effects, and pathological heterogeneity among DR subtypes and disease stages. Since PANoptosis also fundamentally contributes to host defense and retinal innate immune homeostasis, DR therapeutic strategies should prioritize biomarker-guided, local ocular delivery, time-dependent stage intervention, and cell-type-specific fine tuning, rather than generalized systemic suppression. By reconciling mechanistic progress with unresolved translational bottlenecks, this review proposes that PANoptosis serves as a dynamic and evolving conceptual framework for interpreting inflammatory neurovascular degeneration in DR, instead of an entirely confirmed clinical therapeutic target. We emphasize the urgent need for DR-specific mechanistic verification, longitudinal biomarker cohort studies, and well-designed multicenter clinical trials to advance safe and precise targeted interventions.