Deciphering the spatial landscape and plasticity of immunosuppressive fibroblasts in breast cancer.

Although heterogeneity of FAP+ Cancer-Associated Fibroblasts (CAF) has been described in breast cancer, their plasticity and spatial distribution remain poorly understood. Here, we analyze trajectory inference, deconvolute spatial transcriptomics at single-cell level and perform functional assays to generate a high-resolution integrated map of breast cancer (BC), with a focus on inflammatory and myofibroblastic (iCAF/myCAF) FAP+ CAF clusters. We identify 10 spatially-organized FAP+ CAF-related cellular niches, called EcoCellTypes, which are differentially localized within tumors. Consistent with their spatial organization, cancer cells drive the transition of detoxification-associated iCAF (Detox-iCAF) towards immunosuppressive extracellular matrix (ECM)-producing myCAF (ECM-myCAF) via a DPP4- and YAP-dependent mechanism. In turn, ECM-myCAF polarize TREM2+ macrophages, regulatory NK and T cells to induce immunosuppressive EcoCellTypes, while Detox-iCAF are associated with FOLR2+ macrophages in an immuno-protective EcoCellType. FAP+ CAF subpopulations accumulate differently according to the invasive BC status and predict invasive recurrence of ductal carcinoma in situ (DCIS), which could help in identifying low-risk DCIS patients eligible for therapeutic de-escalation.

A multiomics analysis-assisted deep learning model identifies a macrophage-oriented module as a potential therapeutic target in colorectal cancer.

Colorectal cancer (CRC) is a common malignancy involving multiple cellular components. The CRC tumor microenvironment (TME) has been characterized well at single-cell resolution. However, a spatial interaction map of the CRC TME is still elusive. Here, we integrate multiomics analyses and establish a spatial interaction map to improve the prognosis, prediction, and therapeutic development for CRC. We construct a CRC immune module (CCIM) that comprises FOLR2+ macrophages, exhausted CD8+ T cells, tolerant CD8+ T cells, exhausted CD4+ T cells, and regulatory T cells. Multiplex immunohistochemistry is performed to depict the CCIM. Based on this, we utilize advanced deep learning technology to establish a spatial interaction map and predict chemotherapy response. CCIM-Net is constructed, which demonstrates good predictive performance for chemotherapy response in both the training and testing cohorts. Lastly, targeting FOLR2+ macrophage therapeutics is used to disrupt the immunosuppressive CCIM and enhance the chemotherapy response in vivo.

CD163-Mediated Small-Vessel Injury in Alzheimer's Disease: An Exploration from Neuroimaging to Transcriptomics.

Patients with Alzheimer's disease (AD) often present with imaging features indicative of small-vessel injury, among which, white-matter hyperintensities (WMHs) are the most prevalent. However, the underlying mechanism of the association between AD and small-vessel injury is still obscure. The aim of this study is to investigate the mechanism of small-vessel injury in AD. Differential gene expression analyses were conducted to identify the genes related to WMHs separately in mild cognitive impairment (MCI) and cognitively normal (CN) subjects from the ADNI database. The WMH-related genes identified in patients with MCI were considered to be associated with small-vessel injury in early AD. Functional enrichment analyses and a protein-protein interaction (PPI) network were performed to explore the pathway and hub genes related to the mechanism of small-vessel injury in MCI. Subsequently, the Boruta algorithm and support vector machine recursive feature elimination (SVM-RFE) algorithm were performed to identify feature-selection genes. Finally, the mechanism of small-vessel injury was analyzed in MCI from the immunological perspectives; the relationship of feature-selection genes with various immune cells and neuroimaging indices were also explored. Furthermore, 5×FAD mice were used to demonstrate the genes related to small-vessel injury. The results of the logistic regression analyses suggested that WMHs significantly contributed to MCI, the early stage of AD. A total of 276 genes were determined as WMH-related genes in patients with MCI, while 203 WMH-related genes were obtained in CN patients. Among them, only 15 genes overlapped and were thus identified as the crosstalk genes. By employing the Boruta and SVM-RFE algorithms, CD163, ALDH3B1, MIR22HG, DTX2, FOLR2, ALDH2, and ZNF23 were recognized as the feature-selection genes linked to small-vessel injury in MCI. After considering the results from the PPI network, CD163 was finally determined as the critical WMH-related gene in MCI. The expression of CD163 was correlated with fractional anisotropy (FA) values in regions that are vulnerable to small-vessel injury in AD. The immunostaining and RT-qPCR results from the verifying experiments demonstrated that the indicators of small-vessel injury presented in the cortical tissue of 5×FAD mice and related to the upregulation of CD163 expression. CD163 may be the most pivotal candidates related to small-vessel injury in early AD.

Tumor-specific activation of folate receptor beta enables reprogramming of immune cells in the tumor microenvironment.

Folate receptors can perform folate transport, cell adhesion, and/or transcription factor functions. The beta isoform of the folate receptor (FRß) has attracted considerable attention as a biomarker for immunosuppressive macrophages and myeloid-derived suppressor cells, however, its role in immunosuppression remains uncharacterized. We demonstrate here that FRß cannot bind folate on healthy tissue macrophages, but does bind folate after macrophage incubation in anti-inflammatory cytokines or cancer cell-conditioned media. We further show that FRß becomes functionally active following macrophage infiltration into solid tumors, and we exploit this tumor-induced activation to target a toll-like receptor 7 agonist specifically to immunosuppressive myeloid cells in solid tumors without altering myeloid cells in healthy tissues. We then use single-cell RNA-seq to characterize the changes in gene expression induced by the targeted repolarization of tumor-associated macrophages and finally show that their repolarization not only changes their own phenotype, but also induces a proinflammatory shift in all other immune cells of the same tumor mass, leading to potent suppression of tumor growth. Because this selective reprogramming of tumor myeloid cells is accompanied by no systemic toxicity, we propose that it should constitute a safe method to reprogram the tumor microenvironment.

WNT-dependent interaction between inflammatory fibroblasts and FOLR2+ macrophages promotes fibrosis in chronic kidney disease.

Chronic kidney disease (CKD) is a public health problem driven by myofibroblast accumulation, leading to interstitial fibrosis. Heterogeneity is a recently recognized characteristic in kidney fibroblasts in CKD, but the role of different populations is still unclear. Here, we characterize a proinflammatory fibroblast population (named CXCL-iFibro), which corresponds to an early state of myofibroblast differentiation in CKD. We demonstrate that CXCL-iFibro co-localize with macrophages in the kidney and participate in their attraction, accumulation, and switch into FOLR2+ macrophages from early CKD stages on. In vitro, macrophages promote the switch of CXCL-iFibro into ECM-secreting myofibroblasts through a WNT/ß-catenin-dependent pathway, thereby suggesting a reciprocal crosstalk between these populations of fibroblasts and macrophages. Finally, the detection of CXCL-iFibro at early stages of CKD is predictive of poor patient prognosis, which shows that the CXCL-iFibro population is an early player in CKD progression and demonstrates the clinical relevance of our findings.

microRNA-622 upregulates cell cycle process by targeting FOLR2 to promote CRC proliferation.

BACKGROUND: Epigenetic alterations contribute greatly to the development and progression of colorectal cancer, and effect of aberrant miR-622 expression is still controversial. This study aimed to discover miR-622 regulation in CRC proliferation. METHODS: miR-622 expression and prognosis were analyzed in clinical CRC samples from Nanfang Hospital. miR-622 regulation on cell cycle and tumor proliferation was discovered, and FOLR2 was screened as functional target of miR-622 using bioinformatics analysis, which was validated via dual luciferase assay and gain-of-function and loss-of-function experiments both in vitro and in vivo. RESULTS: miR-622 overexpression in CRC indicated unfavorable prognosis and it regulated cell cycle to promote tumor growth both in vitro and in vivo. FOLR2 is a specific, functional target of miR-622, which negatively correlates with signature genes in cell cycle process to promote CRC proliferation. CONCLUSIONS: miR-622 upregulates cell cycle process by targeting FOLR2 to promote CRC proliferation, proposing a novel mechanism and treatment target in CRC epigenetic regulation of miR-622.

Presence of onco-fetal neighborhoods in hepatocellular carcinoma is associated with relapse and response to immunotherapy.

Onco-fetal reprogramming of the tumor ecosystem induces fetal developmental signatures in the tumor microenvironment, leading to immunosuppressive features. Here, we employed single-cell RNA sequencing, spatial transcriptomics and bulk RNA sequencing to delineate specific cell subsets involved in hepatocellular carcinoma (HCC) relapse and response to immunotherapy. We identified POSTN+ extracellular matrix cancer-associated fibroblasts (EM CAFs) as a prominent onco-fetal interacting hub, promoting tumor progression. Cell-cell communication and spatial transcriptomics analysis revealed crosstalk and co-localization of onco-fetal cells, including POSTN+ CAFs, FOLR2+ macrophages and PLVAP+ endothelial cells. Further analyses suggest an association between onco-fetal reprogramming and epithelial-mesenchymal transition (EMT), tumor cell proliferation and recruitment of Treg cells, ultimately influencing early relapse and response to immunotherapy. In summary, our study identifies POSTN+ CAFs as part of the HCC onco-fetal niche and highlights its potential influence in EMT, relapse and immunotherapy response, paving the way for the use of onco-fetal signatures for therapeutic stratification.

Folate receptor overexpression induces toxicity in a diet-dependent manner in C. elegans.

Folate receptor (FR) alpha (FOLR1) and beta (FOLR2) are membrane-anchored folate transporters that are expressed at low levels in normal tissues, while their expression is strongly increased in several cancers. Intriguingly, although the function of these receptors in, for example, development and cancer has been studied intensively, their role in aging is still unknown. To address this, we utilized Caenorhabditis elegans, in which FOLR-1 is the sole ortholog of folate receptors. We found that the loss of FOLR-1 does not affect reproduction, physical condition, proteostasis or lifespan, indicating that it is not required for folate transport to maintain health. Interestingly, we found that FOLR-1 is detectably expressed only in uterine-vulval cells, and that the histone-binding protein LIN-53 inhibits its expression in other tissues. Furthermore, whereas knockdown of lin-53 is known to shorten lifespan, we found that the loss of FOLR-1 partially rescues this phenotype, suggesting that elevated folr-1 expression is detrimental for health. Indeed, our data demonstrate that overexpression of folr-1 is toxic, and that this phenotype is dependent on diet. Altogether, this work could serve as a basis for further studies to elucidate the organismal effects of abnormal FR expression in diseases such as cancer.

Functional Annotation and Gene Set Analysis of Gastric Cancer Risk Loci in a Korean Population.

PURPOSE: We aimed to identify the associated single nucleotide polymorphisms (SNPs) with gastric cancer (GC) risk by genome-wide association study (GWAS) and to explore the pathway enrichment of implicated genes and gene-sets with expression patterns. MATERIALS AND METHODS: The study population was comprised of 1,253 GC cases and 4,827 controls from National Cancer Center and an urban community of the Korean Genome Epidemiology Study and their genotyping was performed. SNPs were annotated, and mapped to genes to prioritize by three mapping approaches by functional mapping and annotation (FUMA). The gene-based analysis and gene-set analysis were conducted with full GWAS summary data using MAGMA. Gene-set pathway enrichment test with those prioritized genes were performed. RESULTS: In GWAS, rs2303771, a nonsynonymous variant of KLHDC4 gene was top SNP associated significantly with GC (odds ratio, 2.59; p=1.32×10-83). In post-GWAS, 71 genes were prioritized. In gene-based GWAS, seven genes were under significant p < 3.80×10-6 (0.05/13,114); DEFB108B had the lowest p=5.94×10-15, followed by FAM86C1 (p=1.74×10-14), PSCA (p=1.81×10-14), and KLHDC4 (p=5.00×10-10). In gene prioritizing, KLDHC4 was the only gene mapped with all three gene-mapping approaches. In pathway enrichment test with prioritized genes, FOLR2, PSCA, LY6K, LYPD2, and LY6E showed strong enrichment related to cellular component of membrane; a post-translation modification by synthesis of glycosylphosphatidylinositol (GPI)-anchored proteins pathway. CONCLUSION: While 37 SNPs were significantly associated with the risk of GC, genes involved in signaling pathways related to purine metabolism and GPI-anchored protein in cell membrane are pinpointed to be playing important role in GC.

SPP1+ TAM subpopulations in tumor microenvironment promote intravasation and metastasis of head and neck squamous cell carcinoma.

Macrophages are heterogeneous cells that play multifaceted roles in cancer progression and metastasis. However, the phenotypic diversity of tumor-associated macrophages (TAMs) in head and neck squamous carcinomas (HNSCC) remains poorly characterized. Here, we comprehensively analyzed the HNSCC single-cell transcriptomic dataset (GSE172577) and identified 5 subsets of myeloid-driven cells as TAMs using Seurat. Deciphering the lineage trajectory of TAMs, we revealed that FCN1+ TAMs could give rise to pro-angiogenesis SPP1+CCL18+ and SPP1+FOLR2+ populations through SPP1-CCL18+ and CXCL9+CXCL10+ TAMs. SPP1+CCL18+ and SPP1+FOLR2+ TAMs harbored pro-angiogenic and metastatic transcriptional programs and were correlated with poor survival of HNSCC patients. Our immunostaining examination revealed that infiltration of SPP1+ TAMs is associated with lymph node metastasis and poor prognosis in patients with HNSCC. Cell-cell communication analysis implied that SPP1+ TAM populations may employ SPP1 signaling to activate metastasis-related ECs. In vitro and in vivo studies, we demonstrated that SPP1hi TAMs enhanced tumor intravasation and metastasis in HNSCC in a manner dependent on the secretion of SPP1, CCL18, and CXCL8. Taken together, our study characterized the cellular heterogeneity of TAM populations and identified two SPP1+ TAM populations that play key roles in HNSCC intravasation and metastasis and serve as predictive markers for patients with HNSCC.

Recent Advances in Folates and Autoantibodies against Folate Receptors in Early Pregnancy and Miscarriage.

Though firstly identified in cerebral folate deficiency, autoantibodies against folate receptors (FRAbs) have been implicated in pregnancy complications such as miscarriage; however, the underlying mechanism needs to be further elaborated. FRAbs can be produced via sensitization mediated by folate-binding protein as well as gene mutation, aberrant modulation, or degradation of folate receptors (FRs). FRAbs may interfere with folate internalization and metabolism through blocking or binding with FRs. Interestingly, different types of FRs are expressed on trophoblast cells, decidual epithelium or stroma, and macrophages at the maternal-fetal interface, implying FRAbs may be involved in the critical events necessary for a successful pregnancy. Thus, we propose that FRAbs may disturb pregnancy establishment and maintenance by modulating trophoblastic biofunctions, placental development, decidualization, and decidua homeostasis as well as the functions of FOLR2+ macrophages. In light of these findings, FRAbs may be a critical factor in pathological pregnancy, and deserve careful consideration in therapies involving folic acid supplementation for pregnancy complications.

Mutational analysis of FOLR1 and FOLR2 genes in children with Myelomeningocele.

Myelomeningocele (MMC) is a congenital disease. For a long time, molecular mechanism of MMC, the role of folate receptor and transporter proteins remain unclear. Folate from maternal lumen to developing embryo is carried out with the help of folate transporters (SLC46A1, SLC19A1, FOLH1 and SLC25A32) and folate receptor (FOLR1, FOLR2 and FOLR3). Due to the loss of function of these important genes, complications can facilitate the risk of MMC. This study focused on the mutational analysis of FOLR1 and FOLR2 genes in children suffering from MMC. Myelomeningocele is a rare disorder so twenty blood samples from the children were collected. Primers of selected exons for FOLR1 and FOLR2 genes were designed with the help of PrimerFox software. Extracted DNA was amplified, and PCR based mutational analysis was done to check any type of mutation/SNPs in these genes. Sanger sequencing method was performed to confirm mutation in FOLR1 and FOLR2 genes. The results showed that certain environmental factors (smoking, low socio-economic status of mother bearing MMC fetus) were found to be significantly (P<0.05) associated with MMC but no mutation in the selected exons of FOLR1 and FOLR2 genes was detected. Thus, genetic variations in the folate transporter gene may have no role in the progression of MMC in the studied population.

Single-cell profiling reveals the trajectory of FOLR2-expressing tumor-associated macrophages to regulatory T cells in the progression of lung adenocarcinoma.

An immunosuppressive microenvironment enriched with regulatory CD4+ T lymphocytes (Tregs) facilitates the progression of lung adenocarcinoma (LUAD). This study aims to investigate the cellular mechanism underlying the formation of the immunosuppressive microenvironment in LUAD. LUAD samples (n = 12) and normal lung samples (n = 3) were obtained from patients with different pathological stages of LUAD. Single-cell RNA sequencing was performed to classify cellular components and analyze the transcriptomes, including transcription factors/targets and chemokine ligands/receptors, followed by bioinformatics study such as pseudotime analysis. Myeloid cells and T cells were the most abundant cell types in tumors and normal lung tissues, while tumor-associated macrophage-folate receptor 2 (TAM-FOLR2) and CD4+ nuclear receptor subfamily 4 group A member 3 (NR4A3) exhibited sharp increases in invasive adenocarcinoma (IA). The enrichment of TAM-FOLR2 in IA might result from alveolar resident macrophage-resistin (ARM-RETN) transformation and recruitment of dendritic cells (DCs) and other TAMs, as evidenced by temporal trajectories and differential expression profiles of chemokine ligands/receptors versus those in the early stages of tumors. High expression of CCL17/19/22 was observed in IA as well as in DCs, along with the strong interaction of TAM-FOLR2 with DCs. The results of pseudotime analysis suggested that CD4+NR4A3 might potentially convert to CD4+FOXP3, further supported by the high expression of NR4A3 target genes in CD4+FOXP3 cells. This study provides a single-cell transcriptome atlas from preinvasive to invasive LUAD and reveals a potential ARM-RETN/TAM-FOLR2/DCs/CD4+NR4A3/CD4+FOXP3 trajectory in shaping the immune suppressive microenvironment along the pathogenesis of LUAD.

Roux-en-Y gastric bypass affects the expression of genes related to the intestinal folate metabolism pathway in obese women.

OBJECTIVES: Roux-en-Y gastric bypass (RYGB) promotes sustained weight loss, and the resulting new gastrointestinal anatomy can contribute to nutritional depletions. Folate deficiency is one of the most frequently observed nutritional deficiencies after RYGB. The aim of this study was to assess whether RYGB affects the expression of genes related to the intestinal folate metabolism pathway as an additional molecular mechanism contributing to its postoperative deficiency. METHODS: Biopsies from the duodenum, jejunum, and ileum of 20 obese women were collected before and 3 mo after RYGB. The expression of genes involved in intestinal folate metabolism was assessed by microarray and reverse transcriptase polymerase chain reaction (RT-qPCR). Folate intake (7-d food record) and plasma levels (electrochemiluminescence) also were measured. RESULTS: Compared with the preoperative phase, transcriptomic alterations were observed in all intestinal segments studied after RYBG, mainly marked by decreased expression of genes encoding folate transporters/receptors and increased expression of genes involved in folate biosynthesis (P < 0.05). Reduced folate intake and plasma folate levels were also observed simultaneously (P < 0.05). Plasma folate concentrations correlated inversely with intestinal FOLR2 and SHMT2 genes (P < 0.001). CONCLUSION: The present findings suggested that impaired expression of genes related to intestinal folate metabolism may contribute to the early systemic deficiency after RYGB and highlight a potential transcriptomic reprogramming of the intestine in response to RYGB to compensate for folate depletion induced by this surgical technique.

scRNA-seq reveals aging-related immune cell types and regulators in vaginal wall from elderly women with pelvic organ prolapse.

Introduction: In the pathology of pelvic organ prolapse (POP), little is known about the contributing role of pelvic microenvironment. Also, the age-related differences in pelvic microenvironment of POP patients is always ignored. In the present study, we investigated the age-related differences in pelvic microenvironment between Young POP patients and Old POP patients, and the novel cell types and critical regulators which contributes to the age-related differences. Methods: Single-cell transcriptomic analyses were used to detect the changes in cell composition and gene expression from the pelvic microenvironment of control group (<60 years), Young POP group (<60 years) and Old POP group (>60 years). Then, immunohistochemistry and immunofluorescence were used to verify the novel cell types and critical regulators in the pelvic microenvironment. Furthermore, histopathological alteration and mechanical property alteration in POP with different ages were revealed by vaginal tissue histology and biomechanical testing. Results: The up-regulated biological process in Old women with POP is mainly related to chronic inflammation, while the up-regulated biological process in Young women with POP is mainly related to extracellular matrix metabolism. Meantime, CSF3+ endothelial cells and FOLR2+ macrophages were found to play a central role in inducing pelvic chronic inflammation. Furthermore, the collagen fiber and mechanical property of POP patients decreased with aging. Conclusions: Taken together, this work provides a valuable resource for deciphering the aging-related immune cell types and the critical regulators in pelvic microenvironment. With better understanding of normal and abnormal events in this pelvic microenvironment, we provided rationales of personalized medicine for POP patients with different ages.

Single-cell RNA sequencing reveals unique monocyte-derived interstitial macrophage subsets during lipopolysaccharide-induced acute lung inflammation.

Interstitial macrophages (IMs) reside in the lung tissue surrounding key structures including airways, vessels, and alveoli. Recent work has described IM heterogeneity during homeostasis, however, there are limited data on IMs during inflammation. We sought to characterize IM origin, subsets, and transcriptomic profiles during homeostasis and lipopolysaccharide (LPS) induced acute lung inflammation. During homeostasis, we used three complementary methods, spectral flow cytometry, single-cell RNA-sequencing, and gene regulatory network enrichment, to demonstrate that IMs can be divided into two core subsets distinguished by surface and transcriptional expression of folate receptor ß (Folr2/FRß). These subsets inhabited distinct niches within the lung interstitium. Within FRß+ IMs we identified a subpopulation marked by coexpression of LYVE1. During acute LPS-induced inflammation, lung IM numbers expand. Lineage tracing revealed IM expansion was due to recruitment of monocyte-derived IMs. At the peak of inflammation, recruited IMs were comprised two unique subsets defined by expression of genes associated with interferon signaling and glycolytic pathways. As recruited IMs matured, they adopted the overall transcriptional state of FRß- resident IMs but retained expression in several origin-specific genes, such as IL-1ß. FRß+ IMs were of near-pure resident origin. Taken together our data show that during LPS-induced inflammation, there are distinct populations of IMs that likely have unique functions. FRΒ+ IMs comprise a stable, resident population, whereas FRß- ΙΜs represent a mixed population of resident and recruited IMs.

Elevated expression of receptors for EGF, PDGF, transferrin and folate within murine and human lupus nephritis kidneys.

OBJECTIVE: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease where the body's immune system targets cells and tissue in numerous organs, including the kidneys. Lupus nephritis (LN) is a highly heterogeneous disease, and diagnosis is difficult because clinical manifestations vary widely among patients. Comprehensive proteomic studies reported recently in LN have identified several urinary proteins which are also cell-surface receptors. If indeed these receptor proteins are also hyper-expressed within the kidneys, ligands to these receptors may be useful for drug targeting. METHODS: scRNA sequence data analysis and immunohistochemistry were performed on LN kidneys for expression of four implicated receptors, EGFR, FOL2R2, PDGF-RB, and TFRC. RESULTS: In reported scRNA sequencing studies from 21 LN patients and 3 healthy control renal biopsies or renal-infiltrating immune cells from 24 LN biopsies, EGFR, FOLR2, PDGF-Rb, and TFRC were all hyper expressed within LN kidneys in comparison to healthy kidneys, either within resident renal cells or infiltrating leukocytes. Immunohistochemistry staining of murine lupus renal biopsies from lupus mice revealed EGFR, FOLR2, TFRC and PDGF-RB were elevated in LN kidneys. Immunohistochemistry staining of human Class II, Class III, and Class IV kidney tissue sections revealed EGFR, TFRC, and PDGF-RB were significantly elevated in proliferative LN kidneys. CONCLUSION: These findings underscore the potential of EGFR, TFRC, FOLR2, and PDGF-RB as promising receptors for potential drug-targeting in LN.

Single-cell profiling of healthy human kidney reveals features of sex-based transcriptional programs and tissue-specific immunity.

Knowledge of the transcriptional programs underpinning the functions of human kidney cell populations at homeostasis is limited. We present a single-cell perspective of healthy human kidney from 19 living donors, with equal contribution from males and females, profiling the transcriptome of 27677 cells to map human kidney at high resolution. Sex-based differences in gene expression within proximal tubular cells were observed, specifically, increased anti-oxidant metallothionein genes in females and aerobic metabolism-related genes in males. Functional differences in metabolism were confirmed in proximal tubular cells, with male cells exhibiting higher oxidative phosphorylation and higher levels of energy precursor metabolites. We identified kidney-specific lymphocyte populations with unique transcriptional profiles indicative of kidney-adapted functions. Significant heterogeneity in myeloid cells was observed, with a MRC1+LYVE1+FOLR2+C1QC+ population representing a predominant population in healthy kidney. This study provides a detailed cellular map of healthy human kidney, and explores the complexity of parenchymal and kidney-resident immune cells.

Identification of a six-gene prognostic signature for bladder cancer associated macrophage.

As major components of the tumor microenvironment (TME), tumor-associated macrophages (TAMs) play an exceedingly complicated role in tumor progression and tumorigenesis. However, few studies have reported the specific TAM gene signature in bladder cancer. Herein, this study focused on developing a TAM-related prognostic model in bladder cancer patients based on The Cancer Genome Atlas (TCGA) data. Weighted Gene Co-Expression Network Analysis (WGCNA) was used to identify key genes related to TAM (M2 macrophage). Gene ontology (GO) enrichment and the Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway analysis showed the functional categories of the key genes. Simultaneously, we used the Least Absolute Shrinkage and Selection Operator (LASSO) and univariate and multivariate Cox regressions to establish a TMA-related prognostic model containing six key genes: TBXAS1, GYPC, HPGDS, GAB3, ADORA3, and FOLR2. Subsequently, single-cell sequencing data downloaded from Gene Expression Omnibus (GEO) suggested that the six genes in the prognostic model were expressed in TAM specifically and may be involved in TAM polarization. In summary, our research uncovered six-TAM related genes that may have an effect on risk stratification in bladder cancer patients and could be regarded as potential TAM-related biomarkers.

A Population of TIM4+FOLR2+ Macrophages Localized in Tertiary Lymphoid Structures Correlates to an Active Immune Infiltrate Across Several Cancer Types.

TIM4 has previously been associated with antitumor immunity, yet the pattern of expression and the function of this receptor across human cancer tissues remain poorly explored. Here we combined extensive immunolabeling of human tissues with in silico analysis of pan-cancer transcriptomic data sets to explore the clinical significance of TIM4 expression. Our results unveil that TIM4 is expressed on a fraction of cavity macrophages (CATIM4+MΦ) of carcinoma patients. Moreover, we uncover a high expression of TIM4 on macrophages of the T-cell zone of the carcinoma-associated tertiary lymphoid structures (TLSTIM4+MΦ). In silico analysis of a pan-cancer data set revealed a positive correlation between TIM4 expression and markers of B cells, effector CD8+ T cells, and a 12-chemokine signature defining tertiary lymphoid structure. In addition, TLSTIM4+MΦ were enriched in cancers displaying microsatellite instability and high CD8+ T-cell infiltration, confirming their association with immune-reactive tumors. Both CATIM4+MΦ and TLSTIM4+MΦ express FOLR2, a marker of tissue-resident MΦ. However, CATIM4+MΦ had a higher expression of the immunosuppressive molecules TREM2, IL10, and TGFß as compared with TLSTIM4+MΦ. By analyzing a scRNA sequence data set of tumor-associated myeloid cells, we identified two TIM4+FOLR2+ clusters coherent with CATIM4+MΦ and TLSTIM4+MΦ. We defined specific gene signatures for each subset and found that the CATIM4+ MΦ signature was associated with worse patient survival. In contrast, TLSTIM4+MΦ gene signature positively correlates with a better prognosis. Together, these data illustrate that TIM4 marks two distinct macrophage populations with distinct phenotypes and tissue localization and that may have opposing roles in tumor immunity.