Tamas Ordog

The intestinal epithelium is exposed to diverse combinations of microbiota-derived compounds; however, the mechanisms by which the host integrates these signals remain poorly defined. Studying two highly abundant microbial metabolites, we identified the purine metabolite hypoxanthine as an effector metabolite that directly drives signaling and the short-chain fatty acid butyrate as a regulatory metabolite that conditions host responsiveness. Specifically, hypoxanthine activates the adenosine A1 receptor–TRPC4 axis in enterochromaffin (EC) cells, triggering calcium influx and serotonin release resulting in accelerated gastrointestinal transit locally and increased platelet activation systemically. In contrast, butyrate epigenetically upregulates specific G protein–coupled receptors and ion channels to enhance response to hypoxanthine and the neurotransmitters norepinephrine and dopamine. These findings define a cooperative signaling framework and highlight the role of EC cells as a distinct epithelial signaling hub that senses and integrates microbial metabolite signals to drive physiological responses. Our findings provide a mechanistic foundation for therapeutic strategies that leverage combinatorial microbial signaling.

Intestinal epithelial cells (IEC) are the primary cell type in direct contact with stimuli from the luminal microbiota, playing a critical role in host-microbe interactions. However, how IECs communicate with underlying immune cells to maintain homeostasis remains poorly understood. In addition, the mechanisms by which IECs sense microbiota-derived stimuli to initiate this crosstalk are not yet fully understood. Here, we demonstrate that oral administration of the gut microbiota metabolite butyrate induces sustained IL-10 production in CD4⁺ T cells, conferring long-lasting protection against intestinal inflammation even after treatment withdrawal. This persistent immunoregulatory effect is also observed in germ-free mice, indicating the establishment of a butyrate-conditioned intestinal environment. By metabolomic profiling, we identify N1-acetylspermidine as a metabolite contributing to the IL-10-inducing activity of butyrate-treated IEC-conditioned medium. Mechanistically, butyrate induces sustained transcriptional and epigenetic activation of the acetylpolyamine biosynthetic enzyme Sat1 in IECs, which catalyzes the acetylation of spermidine to generate N1-acetylspermidine, and this activation persists after butyrate withdrawal. Together, our findings reveal a mechanism by which butyrate sensing establishes persistent crosstalk between IECs and T cells, thereby maintaining intestinal immune tolerance through epigenetic regulation and reprogramming of epithelial metabolism.

Violin plot; overall abundance

Protein abundance by compartment

Figure 4 from Spatially Resolved, Multiregion Proteomics for Prediction of Immunotherapy Outcome in Deficient Mismatch Repair Metastatic Colorectal Cancer

Purpose:Digital proteomic profiling was performed to identify spatial context in relationship to patient response and survival after anti–PD-1 therapy in metastatic colorectal cancer.Experimental Design:Primary colorectal cancers with deficient mismatch repair from patients treated with anti–PD-1 antibodies were analyzed (N = 30) using digital spatial profiling (GeoMx nCounter). At the invasive margin, 71 proteins were profiled in 10 regions of interest/slide that were segmented into 3 compartments labeled with pan-cytokeratin (epithelia), CD45 (stromal cells), and SYTO13 (nuclei). In an independent cohort (n = 13), digital spatial profiling data and single-cell transcriptomic data were analyzed. Differential protein abundance, after Benjamini–Hochberg correction, was examined by clinical response and progression-free survival (PFS) using multivariable Cox regression.Results:Protein abundance varied significantly between epithelial and stromal compartments. Nonresponders to anti–PD-1 showed higher fibronectin and smooth muscle actin abundance in the epithelial compartment that was associated with significantly shorter PFS (adjusted HR: 6.49 and 4.52, respectively; P < 0.05). In CD45+ stroma, increased expression of proteins related to T cells (CD3 and CD4), NK cells (CD56), antigen presentation (CD40), immune activation (CD27, ICOS), and apoptosis (GZMA) were found in responders (vs nonresponders) to anti–PD-1; each marker was significantly associated with longer patient PFS (0.02 < adjusted HR < 0.17; P < 0.05). In a separate cohort, consistent results by compartment were found for fibronectin and CD56. Gene expression data revealed that fibronectin and smooth muscle actin were primarily derived from cancer-associated fibroblasts.Conclusions:Spatially resolved protein profiles within microenvironments of deficient mismatch repair colorectal cancers can influence patient response and survival after anti–PD-1, highlighting their potential clinical significance.

Spatial multiomic technologies combined with single-cell profiling offer unprecedented opportunities to elucidate the molecular and cellular architecture of complex tissues. Combining spatial barcoding of nuclei within individual cryosections prepared from tissue blocks embedded in optimal cutting temperature (OCT) freezing medium with single-cell multiomic assays into a simple and efficient workflow can facilitate the annotation and spatially resolved molecular analysis of cells in tissue types. This study reports a spatial cleavage under targets & tagmentation (CUT&Tag) multiome approach that simultaneously profiles histone H3K27ac modification and gene expression in a single frozen brain section. Following spatial barcoding, single nuclei are isolated and subjected to CUT&Tag, combined capture of tagmented DNAs, RNAs, and spatial barcodes, library preparation, and sequencing. The workflow generates spatially annotated epigenomic and transcriptomic profiles from the same nuclei, enabling assignment of the multiomic information to anatomical coordinates. Integrating epigenomic information with spatial transcriptomic data increases the fidelity of cell-type identification and reveals spatially organized regulatory programs and cell-cell interactions within intact tissues.

Immune dysfunction has been increasingly implicated in bipolar disorder, but the underlying mechanisms remain unclear. To address this, we profile 833 genome-wide chromatin immunoprecipitation sequencing datasets spanning five histone marks in peripheral blood immune cells from 88 Type I bipolar disorder patients and 92 controls, integrating them with whole-genome sequencing and clinical data. We identify disease-associated cis -regulatory elements and genetically influenced regulatory elements, revealing immune signatures and pathways involving calcium signaling and endoplasmic reticulum transport. By integrating genetic risk variants, differential and genetically influenced regulatory elements, and regulatory element—gene links, we prioritize 39 driver genes, 28 of which are exclusively supported by blood evidence. We further stratify patients into five epigenomic subtypes with distinct clinical features and genetic risk profiles and identify compounds that reverse disease-associated epigenomic signatures. Here, we combine immune epigenomics with genetics and clinical traits to identify driver genes, patient subtypes, and therapeutic candidates, highlighting immune contributions to type I bipolar disorder pathogenesis.