Search Results (855)

Background/Objectives: Signaling mediators of PPARγ influence pathways involved in adipogenesis, lipid storage, inflammation, energy-related processes, and glucose utilization. Recent research indicates that PPARγ coregulators, recruited or released during ligand binding, govern specific gene pathways. It was recently discovered that Gα_q, a heterotrimeric G protein subunit, also signals to PPARγ and may significantly affect adipogenesis and glucose sensitivity. Methods: To explore Gα_q’s role in adipocytes, we generated CRISPR-mediated Gα_q (Gnaq) knockout (Gnaq KO) and scramble control cells from 3T3-L1 preadipocytes. Results: The absence of Gα_q resulted in increased lipid accumulation and elevated serine 273 (but not serine 112) phosphorylation of PPARγ. Gα_q deficiency also decreased mitochondrial abundance and respiration in response to PPARγ ligands such as rosiglitazone, pioglitazone, and troglitazone. RNA sequencing comparing differentiated Gnaq KO and control adipocytes identified over 800 differentially expressed genes, including those associated with enhanced lipid metabolism and reduced inflammation. Corresponding PamGene kinome profiling showed increased serine/threonine kinase activity and decreased phosphotyrosine kinase signaling in Gnaq KO adipocytes. Conclusions: These findings support Gα_q as a regulator of adipocyte function, linking kinase signaling pathways to PPARγ-mediated transcription. This research provides mechanistic insights into targeting Gα_q as a potential treatment for individuals with obesity and metabolic disorders. Full article

Leukocyte recruitment from blood into tissues involves sequential adhesive steps, including rolling and integrin-dependent arrest. VLA-4 can support firm adhesion and, in some settings, rolling interactions, whereas CD44–hyaluronan interactions have also been implicated in leukocyte rolling. Here, we used adhesion assays and parallel-plate flow chamber experiments to analyze CD44–hyaluronan-dependent monocyte interactions on ECV304 monolayers and to compare them with α4-integrin-sensitive adhesion on endothelial monolayers. WEHI 78/24 monocytoid cells interacted with ECV304 monolayers in a CD44- and hyaluronan-dependent manner, whereas adhesion to HMEC-1 and bEnd.3 monolayers was sensitive to α4-integrin blockade. Blocking CD44, adding soluble hyaluronan, or treating ECV304 monolayers with hyaluronidase reduced adhesion and rolling. Mixed primary human monocyte preparations also showed CD44-dependent adhesion and rolling on ECV304 monolayers. ECV304 cells are interpreted here not as endothelial cells, but as T24-derived, hyaluronidase-sensitive cellular monolayers useful for functional analysis of CD44–hyaluronan-dependent interactions. These findings support a substrate-dependent functional hierarchy in which CD44–hyaluronan-dependent monocyte rolling becomes detectable when α4-integrin-dependent adhesion is not dominant, while emphasizing the cell-model-based nature of the assay. Full article

Alzheimer’s disease (AD) is a multifactorial neurodegenerative disorder characterized by amyloid-β (Aβ) and the accumulation of tau in the brain, which triggers robust innate immune responses. Growing evidence indicates that neuroinflammation contributes to AD progression by overactivating microglia through the release of cytokines and chemokines. In general, chemokines can disrupt neuronal communication and promote blood–brain barrier permeability. Peripheral immune cells are mobilized into the brain by a gradient of chemokines. These processes link peripheral immune responses with substantial T-cell infiltration into the CNS parenchyma, leptomeninges and cerebrospinal fluid of both AD mice and AD patients. This finding underscores the relevance of the adaptive immune system, particularly T and B cells, in AD neuropathology. T-cell infiltration into the brain can influence amyloid clearance through chemokine signalling. However, chemokines play a critical role in AD by either promoting or suppressing disease progression. The infiltration of peripheral T and B cells into the brain parenchyma can exacerbate neuronal loss, yet it may also exert neuroprotective effects. Despite the presence of CD4⁺ and CD8⁺ T cells in postmortem brains of AD patients, debate continues about their role in AD brains, in terms of whether they are protective or detrimental. Understanding the complex role of chemokines in controlling innate and adaptive immune responses by modulating neuron–glia interactions (involving astrocytes and microglia) may provide novel therapeutic approaches for AD. Targeting chemokine signalling or treating with drugs that can prevent the recruitment of immune cells may be promising strategies for treating AD neuropathology. Therapies that prevent the overactivation of T cells in the brain could lead to protective strategies against AD. In fact, regulatory T cells (Tregs) could delay the onset of cognitive symptoms, because they suppress inflammation and slow the accumulation of Aβ plaques and p-Tau in the brain. Complementary strategies, such as photobiomodulation, nanoparticle, and T-cell-based approaches, could mitigate AD progression in patients. Full article

Background/Objectives: Inherited variants in IKZF1 and CDKN2A/2B are among the most consistently reported germline susceptibility markers for childhood acute lymphoblastic leukemia (ALL). Nonetheless, effect sizes differ across ancestry groups, age ranges, and immunophenotypic subtypes, making well-characterized population-specific replication studies valuable for refining ancestry-specific evidence. This study examined two sentinel variants with historical relevance, IKZF1 rs4132601 and CDKN2A rs3731217, within a pediatric Greek cohort. Methods: A case–control study with retrospective case ascertainment and control recruitment through routine pediatric visits was conducted, comprising 50 children and adolescents with ALL and 91 healthy controls from Crete, Greece. Constitutional DNA was isolated from peripheral blood samples collected during remission in cases, while controls provided peripheral blood for targeted germline genotyping. Genotyping was performed using PCR-restriction fragment length polymorphism analysis. We evaluated Hardy–Weinberg equilibrium and genotype and allele distributions and analyzed the data using logistic regression models. Results: Control minor allele frequencies were broadly compatible with public European reference data. Neither IKZF1 rs4132601 nor CDKN2A rs3731217 showed a significant association with susceptibility to childhood ALL under either genotype-based or additive models. Results remained consistent after excluding T-cell ALL cases. Exploratory genotype-phenotype analyses did not reveal robust associations with clinical or molecular features. Conclusions: In this first Greek pediatric assessment of IKZF1 rs4132601 and CDKN2A rs3731217, no significant association with ALL susceptibility was observed. Within the constraints of limited statistical power, these negative findings provide population-specific evidence, refine regional allele-frequency and effect-size estimates, highlight the limitations of relying on single historical sentinel single-nucleotide polymorphisms (SNPs), and support future ancestry-informed and polygenic approaches in southeastern European cohorts. Full article

Streptococcus pneumoniae is a leading cause of community-acquired pneumonia, yet the immune mechanisms required for protection against invasive pulmonary infection remain inadequately understood. Using a murine model of homologous protection against invasive pneumococcal pneumonia, we explored the relative contributions of humoral and cellular immunity using adoptive serum transfer, immune cell depletion, and lung transcriptional profiling. Our findings indicated that passive transfer of immune serum provided robust protection, while neutrophil depletion significantly compromised bacterial control, highlighting that both antibodies and neutrophils are key mediators of protection. In contrast, depletion of CD4⁺ T cells or NK cells did not compromise survival. Although IL-17A has been widely implicated in host defense against pneumococcal infection, IL-17A-deficient mice remained protected, albeit with delayed clearance and reduced early antibody responses. We associate this delay with compensatory upregulation of IL-17F and increased expression of Th1-associated genes in the lungs. Together, these findings indicate that IL-17A is not essential for protection and support a model in which coordinated Th1- and Th17-related cytokine responses collectively promote neutrophil recruitment and effective antibody-mediated defense. These results highlight functional redundancy within the IL-17 cytokine axis and suggest that integrated cytokine networks, rather than individual mediators, underpin protective immunity to pneumococcal pneumonia, with implications for next-generation vaccine design. Full article

Background: Cytochrome P450 3A4 (CYP3A4) is a key membrane-anchored drug-metabolizing enzyme. Its expression and purification in heterologous systems are severely hindered by low yield and detergent-induced structural inactivation. Although extracellular vesicles (EVs) provide an ideal natural lipid bilayer environment to stabilize membrane proteins, targeted loading remains challenging. The ESCRT and ALIX-binding region (EABR) of CEP55 can efficiently recruit core components of the endosomal sorting complex (ESCRT) to mediate membrane fission. Objectives: This study used the EABR motif to drive the targeted vesicular secretion of CYP3A4, thereby establishing a novel membrane protein engineering platform. Methods and Results: EABR was fused with fluorescent protein, confirming its specific mediation of vesicular secretion. Recombinant plasmids of EABR/CYP3A4 and its reverse mutant (R-EABR) were transfected into HEK293T cells. Western blot and midazolam-based metabolic assays showed that forward EABR significantly enhanced CYP3A4 expression and EV secretion, while R-EABR lost exocytosis function. EVs isolated by ultracentrifugation verified EABR’s role in recruiting ESCRT and improving CYP3A4 activity. Conclusions: Forward CEP55-EABR specifically and efficiently drives vesicular encapsulation of CYP3A4, enhancing its expression and secretion. This ESCRT-mediated strategy avoids destructive purification, provides a stable lipid-rich bioreactor for CYP3A4, and has great translational potential in high-throughput in vitro drug metabolism and screening platforms. Full article

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BrCa), owing to its lack of targetable receptors and resistance to chemical and molecularly targeted therapeutic approaches. While chemotherapy and surgical resection remain the standard of care, these interventions have significant side effects and varying patient outcomes. Thermally ablative focused ultrasound (T-FUS)—a non-invasive and non-ionizing therapy that utilizes targeted acoustic energy to debulk tumors—has displayed immunomodulatory effects in BrCa. However, T-FUS as a monotherapy has had limited clinical efficacy in TNBC due to the presence of anti-inflammatory immunosuppressive myeloid cells (isMCs). We hypothesized that the elimination of isMCs or initiating tumoricidal activity from them would lead to augmented activity of T-FUS. Thus, we interrogated the ability of myeloablative chemotherapies and antibodies; myeloid recruiting chemokine receptor blockade; and TLR agonists to remodel the tumor myeloid populations. Consistent with our previous studies, we found that while myeloablative chemotherapies decreased circulating isMCs, they had little impact on intratumoral isMCs. In contrast, antibodies targeting Ly6C and Ly6G ablated intratumoral isMCs and systemic isMCs, yet their effect was transient and was accompanied by a surprising depletion of T cells. While targeting CCR2, the dominant chemokine receptor for intratumoral isMC diminished a large subset of immunosuppressive cells within the TME; it also depleted T cells and dendritic cells. Contrary to previous studies, TLR stimulation failed to repolarize myeloid cells into a pro-inflammatory, tumoricidal phenotype but did lead to their depletion from the tumor microenvironment (TME) and mobilization of conventional dendritic cells to the draining lymph nodes. We therefore hypothesized that combining isMC depletion and TLR-driven immune activation would enhance FUS efficacy; however, this combinatorial regimen did not enhance overall survival or control tumor volume after T-FUS treatment. Thus, the BrCa TME is highly resistant to approaches intended to remodel the myeloid cell component which fail to synergize with T-FUS-mediated tumor ablation. Full article

Resistance to systemic therapy remains the defining challenge in the management of gastric cancer (GC) and esophageal adenocarcinoma (EAC). While genomic drivers of resistance are well characterized, traditional bulk profiling has failed to capture the physical rules governing tumor survival within the complex tissue ecosystem. Emerging data from 2024–2025, leveraging high-resolution spatial transcriptomics and multi-omics, have recontextualized resistance as a phenomenon of “spatial privilege” rather than solely an intrinsic cellular fate. This review summarizes recent evidence to define “architectural refuges”: distinct spatial niches that physically shield malignant clones from cytotoxic and targeted agents. We delineate three critical resistance domains common to upper gastrointestinal adenocarcinomas: (1) The “Excluded” Niche, where specific cancer-associated fibroblast (CAF) subpopulations (iCAFs vs. myCAFs) and stiffened extracellular matrix create hypovascular zones that limit drug delivery; (2) the “Immune-Tolerant” Niche, characterized by the spatial exclusion of CD8+ T cells and the recruitment of suppressive myeloid populations via the MIF/CD74 and USP14 axes; and (3) the “Metabolic” Niche, where mitochondrial heterogeneity and lipid metabolic symbiosis establish nutrient-deprived niches that select for stem-like, dormant states. By mapping these conserved spatial determinants from primary GEJ tumors to peritoneal and distant metastases, we argue that overcoming resistance requires an advancement: moving beyond targeting individual mutations to dismantling the multicellular architecture that sustains malignancy. Full article

Unexplained infertility affects up to 30% of couples and has been associated with heat shock proteins (HSP) and endometrial stress. HSPs and their co-chaperones are part of a complex network of proteins responsible for maintaining protein homeostasis and cell survival. This exploratory hypothesis-generating study investigated the possible relationship between HSPs and unexplained infertility. Twenty-five women were recruited from an IVF clinic. Eleven were confirmed for unexplained infertility (UI), while fourteen were age- and body mass index (BMI)-matched couples with confirmed male factor infertility (MFI), acting as controls. Blood samples were obtained at day 21 of the luteal phase, and plasma measurement of 19 HSPs and co-chaperones undertaken using the slow off-rate modified aptamer (SomaScan) platform. Welch’s t-test and a permutation test were used to compare group means, and Pearson’s correlations to examine relationships with HSPs. Of the 19 proteins measured, plasma HSP70 was decreased (permutation p = 0.002) in cases with unexplained infertility, while HSC70 and STIP1 were increased (permutation p = 0.017 and p = 0.001, respectively) when compared to MFI control. HSP70 was negatively correlated to both HSC70 and STIP 1 in UI (r = −0.77, permutation p = 0.017; −0.80, permutation p = 0.003, respectively), but not in MFI, whilst HSC70 and STIP1 were positively correlated in both UI and MFI (r = 0.93, permutation p = 0.001; r = 0.65, permutation p = 0.035, respectively). The HSP70-HSC70-STIP1 axis showed HSC70-STIP1 coupling with an inverse relationship with inducible HSP70, findings that may suggest dysregulation of constitutive and stress-inducible chaperone systems in UI. Full article

Solid tumors typically expand in a “cold” immunosuppressive tumor microenvironment (TME) and resist killing by CAR T cells or conventional therapy. Herein, we show that intratumoral injection of C3a and C5a receptor 1 (C3ar/C5ar1) pharmaceutical antagonists in an in situ forming implant (ISFI) can robustly suppress such tumors. Antagonizing autocrine C3ar/C5ar1 signaling in eight human and murine cancers of diverse lineages was universally anti-mitotic and pro-apoptotic in vitro, and growth-repressive in vivo. In contrast to i.p. administration of C3ar/C5ar1 antagonists to tumor-bearing mice, injecting the antagonists intratumorally in slow release poly (lactic-co-glycolic acid) (PLGA) polymer caused near-complete tumor elimination. The focused blockade of C3ar/C5ar1 GPCR signaling in an intratumoral ISFI opposed solid cancers by jointly repressing cancer cell viability/growth, tumor-associated angiogenesis, and myeloid-derived suppressor cell (MDSC) recruitment. Thus, the sustained blockade of C3ar/C5ar1 signaling in an intratumoral ISFI uninterruptedly disrupts three processes essential for solid cancer growth while avoiding adverse effects on other cell types. Our findings may apply to multiple cancer types in which discrete tumor masses can be targeted. Full article

Interferons (IFNs) play fundamental roles in cancer immunity. We have previously shown that conditional ablation of Notch pathway genes in a mouse model of glioma results in impaired IFNγ signaling and immunosuppressive tumors. However, it remained unclear whether the interaction between the Notch and IFN signaling pathways could be leveraged to counteract immune evasion in glioma. Here, we investigated whether expression of the intrinsically active Notch intracellular domain (NICD) could enhance IFN responses in glioma cells. Using a doxycycline (Dox)-inducible system, we overexpressed (OE) NICD in U-251MG human glioma cells. NICD-OE dramatically potentiated STAT1 phosphorylation in response to stimulation with either IFNγ or IFNα. Moreover, NICD-OE induced the expression of the transcription factor IRF1, a regulator of IFN signaling responses. Notably, NICD-OE in U-251MG human glioma cells boosted the IFNγ-dependent transcription of the CXCL9 and CXCL10 genes, which encode cytokines that regulate T cell function. Accordingly, NICD-OE in vivo promoted cytotoxic T lymphocyte recruitment to the tumor and reduced tumor cell proliferation in a murine glioma model. Hence, we have identified a signaling network that could be exploited to enhance anti-tumor immunity in glioma subtypes. Full article

Hepatocellular carcinoma (HCC) is a leading cause of cancer mortality worldwide, increasingly driven by metabolic dysfunction-associated steatotic liver disease alongside viral and alcohol-related cirrhosis. The tolerogenic immune environment of the liver enables tumor immune escape, with regulatory T cells (Tregs) playing a central role. This review synthesizes human-focused evidence (tissues, blood, clinical cohorts, and single-cell/spatial studies) through September 2025 to define how Tregs are recruited, maintained, and functionally deployed in HCC. Across datasets, intratumoral effector-like Tregs (eTregs) expressing ICOS, CTLA-4, CCR8, and CD39/CD73 accumulate within tumors and co-localize with exhausted cytotoxic PD-1^hi CD8⁺ T cells and suppressive myeloid cells. Recruitment is driven mainly by CCL20–CCR6 and CCL22/CCL17–CCR4 signaling, while CCR8 marks highly suppressive tumor-resident Tregs. Their persistence is supported by TGF-β, IL-10, IL-35, adenosine signaling, IL-2 sequestration, and metabolic adaptation. Spatial biomarkers, including ICOS⁺/CCR8⁺ eTreg density and CD8:Treg ratios, associate with prognosis and emerging immunotherapy responses. Etiology further shapes immune architecture: HBV-related HCC often forms Treg-exhausted T-cell niches around viral antigens, whereas MASLD/MASH promotes stromal and metabolic barriers that may reduce PD-(L)1 efficacy. Current treatments (PD-(L)1 blockade with anti-VEGF or CTLA-4, and some TKIs) intersect with Treg biology, while emerging strategies targeting CCR8, CCR4, ICOS, or the adenosine pathway aim to selectively disrupt intratumoral eTreg networks. This review underscores that an etiology-aware, spatial-biomarker framework may guide the integration of selective Treg targeting with PD-(L)1-based therapies in HCC. Full article

Radiotherapy (RT) is a cornerstone of cancer treatment, traditionally recognized for its direct cytotoxic effects via DNA damage. However, emerging evidence highlights RT as a profound modulator of the tumor microenvironment (TME), acting as a “double-edged sword” that greatly influences the success of immune checkpoint inhibitors (ICIs). On the one hand, RT acts like an in situ vaccine, causing immunogenic cell death and activating the cGAS-STING pathway, which leads to dendritic cell maturation, T-cell infiltration, and reactive PD-L1 expression. This effect can turn “cold” tumors into “hot” ones, making them more responsive to immune checkpoint blockade. On the other hand, RT can lead to resistance to ICIs by promoting an immunosuppressive environment, recruiting regulatory T cells, M2 macrophages, and myeloid-derived suppressor cells. This review analyzes the mechanisms behind this immunological duality and assesses how parameters such as dose, fractionation, and particle type (e.g., carbon ion versus photon therapy) can be optimized to enhance immune activation. Lastly, we discuss future strategies that focus on innate immunity and tumor metabolism, showing how targeting nutrient depletion and ferroptosis can break down immunosuppressive barriers and position RT as an essential component of precision immuno-oncology. Full article

Since the first approval of CTLA-4 blockade for melanoma, immune checkpoint inhibitors (ICIs) have expanded into a major class of cancer therapy, with more than 100 FDA-approved oncological indications across metastatic and earlier-stage disease settings, including use as monotherapy and in combination regimens. Preclinical research has largely focused on myocarditis and atherosclerosis, but a wider set of phenotypes, such as non-inflammatory left ventricular dysfunction (NILVD), arrhythmias, and vasculitis, can be observed, and they are rarely connected within a single mechanistic model. We aim to build a systems-oriented, mechanistic framework of the most widely studied biological processes; it will link the main checkpoint pathways to relevant cardiac and vascular cell types, molecular pathways, immune synapses, and candidate biomarkers. We searched PubMed, Scopus, and Web of Science using combinations of terms for immune checkpoint inhibition and cardiovascular-immune-related adverse events that provide mechanistic insight into cardiac-immune-related adverse reactions (irAEs). An AI-assisted semantic clustering approach was used only to organize the included literature. The integrated framework identifies PD-1/PD-L1 as the dominant mechanistic hub linking T-cell activation, endothelial recruitment, myocardial injury, and vascular inflammation. Across phenotypes, a shared immune core involving checkpoint pathways, cytokine signaling, and leukocyte trafficking coexists with phenotype-restricted mediators that may bias injury toward myocarditis, vascular inflammation, conduction-system disease, or NILVD. KEGG analyses support the enrichment of T-cell receptor signaling, Th17 differentiation, JAK-STAT signaling, cytokine–cytokine receptor interaction, and lipid and atherosclerosis pathways. Candidate biomarkers emerging from the reviewed literature include troponin, IL-6, CXCL9/CXCL10/CXCL13, S100A family proteins, ROCK2, HLA-linked susceptibility signals, and T-cell receptor clonality markers. The AI-assisted clustering broadly recapitulated the expert-defined thematic structure while identifying finer semantic neighborhoods within the literature. This framework provides a support map for further hypotheses about toxicity patterns with current and next-generation checkpoint strategies on the cardiac system, while AI-assisted clustering provides a complementary method for organizing the literature rather than an independent source of biological inference. Full article

Background: Cytokine-induced JAK–STAT signaling becomes dysregulated in chronic human diseases, including cancer and autoimmunity, and contributes to immune cell dysfunction. A cytokine-independent approach to activating STAT proteins could “hardwire” pro-survival and effector programs in immune cells to sustain function within diseased tissues. Engineered variants of the herpesvirus saimiri tyrosine kinase interacting protein (TIP) can recruit the SRC family kinase (SFK) LCK to drive STAT phosphorylation and activation. Here, we evaluated the interactome of a TIP-derived, cytokine-independent STAT5 activator and determined whether it could induce STAT5 activation in immune cell lines and primary human CD8+ T cells. Methods: A STAT5 activator (aSTAT5) was characterized by proteomics using affinity purification mass spectrometry (AP-MS) to define its interactome and STAT5 binding specificity. STAT5 phosphorylation was assessed in hematopoietic cell lines and primary human CD8+ T cells. Results: Proteomic analysis confirmed preferential association of aSTAT5 with STAT5 relative to other proteins. In cell-based assays, aSTAT5 induced robust STAT5 phosphorylation in LCK-expressing NK-92 and Jurkat T cells, whereas phosphorylation was not observed in Raji B cells or RAW 264.7 macrophages despite expression of closely related SFKs and STAT5. Cytokine-independent STAT5 phosphorylation supported the viability of NK-92 cells and primary human CD8+ T cells during cytokine withdrawal and preserved the cytotoxic function of CAR T cells. Conclusions: We defined the interactome of a cytokine-independent STAT5 activator and demonstrated its capacity to maintain survival and function in human CD8+ T cells and NK-92 cells. These findings underscore the translational potential of engineered, cytokine-independent STAT5 activation for immune cell therapies. Full article