单细胞和大量转录组学鉴定结直肠癌中肿瘤特异性CD36+癌症相关成纤维细胞亚群。

IF 20.1 1区 医学 Q1 ONCOLOGY
Jin Wang, Ming-Jia Xi, Qing Lu, Bi-Han Xia, Yu-Zhi Liu, Jin-Lin Yang
{"title":"单细胞和大量转录组学鉴定结直肠癌中肿瘤特异性CD36+癌症相关成纤维细胞亚群。","authors":"Jin Wang,&nbsp;Ming-Jia Xi,&nbsp;Qing Lu,&nbsp;Bi-Han Xia,&nbsp;Yu-Zhi Liu,&nbsp;Jin-Lin Yang","doi":"10.1002/cac2.12506","DOIUrl":null,"url":null,"abstract":"<p>Dear Editor,</p><p>Cancer-associated fibroblasts (CAFs) are highly versatile and plastic cells in the tumor microenvironment. They have been identified as actively involved in cancer progression and metastasis through their various roles in remodeling the extracellular matrix, suppressing the immune response and reprogramming tumor metabolism [<span>1</span>]. However, many challenges exist in revealing the functional phenotypes and mechanisms of CAFs in different cancers due to limited understanding of CAF heterogeneity [<span>2</span>]. Recent advances in single-cell transcriptome technology have enabled the identification of distinct CAF subpopulations by using unique gene signatures in multiple tumor types [<span>2</span>]. In this study, we successfully identified a tumor-specific CD36<sup>+</sup> CAF subpopulation in colorectal cancer (CRC), which was found to be correlated with the number of tumor-infiltrated immune cells.</p><p>Primary CAFs and normal fibroblasts (NFs) were isolated from 5 fresh CRC tissues and paired normal colon tissues. Isolation methods and descriptions of other assays are shown in the Supplementary Methods. Immunofluorescence and Western blotting assays showed that the mesenchymal marker Vimentin was expressed in both NFs and CAFs, while alpha smooth muscle actin (αSMA) and fibroblast-specific protein 1 (FSP1) were overexpressed in CAFs (Supplementary Figure S1A-D). To further investigate gene expression profiles in these fibroblasts, primary NFs and CAFs were subjected to RNA sequencing. The results showed that CD36 was significantly upregulated in CAFs (Supplementary Figure S1E), which was further validated by immunofluorescence and Western blotting assays (Supplementary Figure S1F-G). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of differentially expressed genes between NFs and CAFs indicated significant enrichment in the mitogen-activated protein kinase (MAPK) signaling pathway, phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway and receptor-ligand activity (Supplementary Figure S1H-I), suggesting that CAFs may play a critical role in intracellular and extracellular signal transduction.</p><p>Previous studies have demonstrated that CAFs achieve high heterogeneity and plasticity across different cancer types [<span>1, 2</span>]. To further reveal the characteristics of CAFs in CRC, we performed single-cell RNA-sequencing (scRNA-seq) using isolated NFs and CAFs and detected 21,248 NFs and 18,097 CAFs (Figure 1A). A total of 7 clusters were identified in these fibroblasts by using sample integration analysis based on distinct gene expression signatures. Interestingly, we found that Clusters 3, 5, and 7 were mainly distributed in the CAF group, accounting for 34.5%, 7.7%, and 2.3% of total CAFs, respectively (Figure 1B-E). Cluster gene signature analysis showed that CD36 was expressed in CAF-specific subgroups, such as Clusters 3 and 5; inhibin subunit beta A (INHBA) was mainly expressed in Clusters 3, 5 and 7; whereas fibroblast growth factor 7 (FGF7) and alcohol dehydrogenase 1B (ADH1B) were mainly expressed in Clusters 1 and 2, which made up the majority of NFs (Figure 1F). To further visualize the marker genes of CAFs, we presented single gene expression data by uniform manifold approximation and projection (UMAP) and found that classical CAF markers, such as fibronectin 1 (FN1), fibroblast activation protein (FAP) and actin alpha 2 (ACTA2), cannot differentiate NFs and CAFs well, whereas a gene expression panel composed of CD36, INHBA, forkhead box S1 (FOXS1), keratin 18 (KRT18) and latent transforming growth factor β-binding protein 1 (LTBP1) was specifically expressed in CAFs (Figure 1G), suggesting the possibility that these genes could serve as new CAF markers in CRC.</p><p>To confirm the scRNA-seq results, we conducted multiplex immunohistochemistry (mIHC) staining and found that CD36 was overexpressed in both tumor cells and tumor stroma, especially in αSMA<sup>+</sup> fibroblasts (Figure 1H-I). CD36 staining using a CRC tissue microarray showed similar results (Supplementary Figure S2A-B). Survival analysis showed that higher expression of CD36 in tumor cells and stromal cells indicated poorer prognosis in CRC patients (Supplementary Figure S2C). In addition, we also performed survival analyses using gene expression data from The Cancer Genome Atlas-Colon Adenocarcinoma (TCGA-COAD) cohort and found that a panel of CAF marker genes [INHBA, FOXS1, periostin (POSTN), thrombospondin 2 (THBS2)] also indicated a poor prognosis in CRC patients (Supplementary Figure S2D-E).</p><p>Overall, we identified a CD36<sup>+</sup> CAF subpopulation in CRC and suggested the potential of CD36 as a specific CAF marker. Moreover, the survival analysis indicated that CAF marker genes (CD36, INHBA, FOXS1, POSTN, THBS2) may be promising prognostic indicators for CRC patients. Similarly, some previous studies have also indicated that CD36<sup>+</sup> CAFs and INHBA<sup>+</sup> CAFs may be predictors of a poor prognosis for hepatocellular carcinoma (HCC) [<span>3</span>] and gastric cancer patients [<span>4</span>], respectively. Mechanically, CD36<sup>+</sup> CAFs promote the formation of an immunosuppressive microenvironment in HCC [<span>3</span>]. To further investigate whether CD36 and INHBA are involved in immune microenvironment modulation in CRC, we analyzed gene expression data from the TCGA-COAD cohort and found that the expression of CD36 and INHBA was positively correlated with the expression of immunosuppressive factors and protumorigenic immune cell markers, such as C-X-C motif chemokine ligand 12 (CXCL12), tumor necrosis factor ligand superfamily member 4 (TNFSF4), T-cell immunoglobulin domain and mucin domain-3 (TIM-3) and CD163 (Supplementary Figure S3A-B). In agreement, estimation of the immune cell infiltration using the CIBERSORT method showed a significant positive correlation between M2 macrophage infiltration and CD36 expression as well as a significant negative correlation between CD8<sup>+</sup> T cell infiltration and INHBA expression (Supplementary Figure S3C-H). Based on these findings, we performed mIHC staining using CRC tissues and found that the infiltration of CD8<sup>+</sup> T cells in CD36<sup>+</sup> CAF-rich regions was significantly reduced (Supplementary Figure S4A-B), while the infiltration of CD68<sup>+</sup> CD163<sup>+</sup> macrophages in CD36<sup>+</sup> CAF-rich regions was increased (Supplementary Figure S4C-D).</p><p>CD36, a scavenger receptor expressed in tumor cells [<span>5</span>], regulatory T cells [<span>6</span>], CD8<sup>+</sup> T cells [<span>7</span>], and cancer-associated macrophages [<span>8</span>], promotes tumor metastasis and induces an immunosuppressive phenotype in different cancer types [<span>5-8</span>]. Tumor cells with elevated CD36 expression exhibit a unique ability to initiate metastasis, and the presence of CD36<sup>+</sup> metastasis-initiating cells associates with a poor prognosis of numerous cancers [<span>9</span>]. However, a previous study demonstrated that CD36 repressed colorectal tumorigenesis by inhibiting glycolysis and that its expression was gradually reduced from adenomas to carcinomas [<span>10</span>]. In our study, we identified that CD36 was overexpressed both in CRC cells and tumor stroma by using multiple methods, and we revealed that high expression of CD36 in CRC indicated poor patient outcomes. In addition, we found that the presence of CD36<sup>+</sup> CAFs was associated with decreased CD8<sup>+</sup> T cell infiltration and increased CD68<sup>+</sup> CD163<sup>+</sup> macrophage infiltration in CRC, suggesting that CD36<sup>+</sup> CAFs were associated with the suppressive immune phenotype. However, whether and how CD36<sup>+</sup> CAFs affect these immune cells remains to be further investigated.</p><p>Overall, CD36<sup>+</sup> CAFs are a poor prognostic factor in CRC and are associated with immune cell infiltration. Our work highlights the importance of identifying tumor-specific CAF subpopulations in understanding the heterogeneous tumor microenvironment.</p><p>This work was supported by grants from the National Natural Science Foundation of China (No. 82173253), the Sichuan Province Science and Technology Support Program (No. 2022YFH0003 and No. 2023NSFSC1900), and the Postdoctoral Research Foundation of China (No. 2022M712260).</p><p>The authors disclose no conflicts.</p><p>Not applicable.</p><p>This study was approved by the Ethics Committee of West China Hospital (No. 2021-179). All human samples were obtained with informed content.</p><p>JW and JLY designed the outline; JW, MJX, and BHX performed the experiments; JW, QL, and YZL analyzed the data. JW and JLY wrote the manuscript; JW, YZL and JLY supervised and edited the manuscript. 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They have been identified as actively involved in cancer progression and metastasis through their various roles in remodeling the extracellular matrix, suppressing the immune response and reprogramming tumor metabolism [<span>1</span>]. However, many challenges exist in revealing the functional phenotypes and mechanisms of CAFs in different cancers due to limited understanding of CAF heterogeneity [<span>2</span>]. Recent advances in single-cell transcriptome technology have enabled the identification of distinct CAF subpopulations by using unique gene signatures in multiple tumor types [<span>2</span>]. In this study, we successfully identified a tumor-specific CD36<sup>+</sup> CAF subpopulation in colorectal cancer (CRC), which was found to be correlated with the number of tumor-infiltrated immune cells.</p><p>Primary CAFs and normal fibroblasts (NFs) were isolated from 5 fresh CRC tissues and paired normal colon tissues. Isolation methods and descriptions of other assays are shown in the Supplementary Methods. Immunofluorescence and Western blotting assays showed that the mesenchymal marker Vimentin was expressed in both NFs and CAFs, while alpha smooth muscle actin (αSMA) and fibroblast-specific protein 1 (FSP1) were overexpressed in CAFs (Supplementary Figure S1A-D). To further investigate gene expression profiles in these fibroblasts, primary NFs and CAFs were subjected to RNA sequencing. The results showed that CD36 was significantly upregulated in CAFs (Supplementary Figure S1E), which was further validated by immunofluorescence and Western blotting assays (Supplementary Figure S1F-G). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of differentially expressed genes between NFs and CAFs indicated significant enrichment in the mitogen-activated protein kinase (MAPK) signaling pathway, phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway and receptor-ligand activity (Supplementary Figure S1H-I), suggesting that CAFs may play a critical role in intracellular and extracellular signal transduction.</p><p>Previous studies have demonstrated that CAFs achieve high heterogeneity and plasticity across different cancer types [<span>1, 2</span>]. To further reveal the characteristics of CAFs in CRC, we performed single-cell RNA-sequencing (scRNA-seq) using isolated NFs and CAFs and detected 21,248 NFs and 18,097 CAFs (Figure 1A). A total of 7 clusters were identified in these fibroblasts by using sample integration analysis based on distinct gene expression signatures. Interestingly, we found that Clusters 3, 5, and 7 were mainly distributed in the CAF group, accounting for 34.5%, 7.7%, and 2.3% of total CAFs, respectively (Figure 1B-E). Cluster gene signature analysis showed that CD36 was expressed in CAF-specific subgroups, such as Clusters 3 and 5; inhibin subunit beta A (INHBA) was mainly expressed in Clusters 3, 5 and 7; whereas fibroblast growth factor 7 (FGF7) and alcohol dehydrogenase 1B (ADH1B) were mainly expressed in Clusters 1 and 2, which made up the majority of NFs (Figure 1F). To further visualize the marker genes of CAFs, we presented single gene expression data by uniform manifold approximation and projection (UMAP) and found that classical CAF markers, such as fibronectin 1 (FN1), fibroblast activation protein (FAP) and actin alpha 2 (ACTA2), cannot differentiate NFs and CAFs well, whereas a gene expression panel composed of CD36, INHBA, forkhead box S1 (FOXS1), keratin 18 (KRT18) and latent transforming growth factor β-binding protein 1 (LTBP1) was specifically expressed in CAFs (Figure 1G), suggesting the possibility that these genes could serve as new CAF markers in CRC.</p><p>To confirm the scRNA-seq results, we conducted multiplex immunohistochemistry (mIHC) staining and found that CD36 was overexpressed in both tumor cells and tumor stroma, especially in αSMA<sup>+</sup> fibroblasts (Figure 1H-I). CD36 staining using a CRC tissue microarray showed similar results (Supplementary Figure S2A-B). Survival analysis showed that higher expression of CD36 in tumor cells and stromal cells indicated poorer prognosis in CRC patients (Supplementary Figure S2C). In addition, we also performed survival analyses using gene expression data from The Cancer Genome Atlas-Colon Adenocarcinoma (TCGA-COAD) cohort and found that a panel of CAF marker genes [INHBA, FOXS1, periostin (POSTN), thrombospondin 2 (THBS2)] also indicated a poor prognosis in CRC patients (Supplementary Figure S2D-E).</p><p>Overall, we identified a CD36<sup>+</sup> CAF subpopulation in CRC and suggested the potential of CD36 as a specific CAF marker. Moreover, the survival analysis indicated that CAF marker genes (CD36, INHBA, FOXS1, POSTN, THBS2) may be promising prognostic indicators for CRC patients. Similarly, some previous studies have also indicated that CD36<sup>+</sup> CAFs and INHBA<sup>+</sup> CAFs may be predictors of a poor prognosis for hepatocellular carcinoma (HCC) [<span>3</span>] and gastric cancer patients [<span>4</span>], respectively. Mechanically, CD36<sup>+</sup> CAFs promote the formation of an immunosuppressive microenvironment in HCC [<span>3</span>]. To further investigate whether CD36 and INHBA are involved in immune microenvironment modulation in CRC, we analyzed gene expression data from the TCGA-COAD cohort and found that the expression of CD36 and INHBA was positively correlated with the expression of immunosuppressive factors and protumorigenic immune cell markers, such as C-X-C motif chemokine ligand 12 (CXCL12), tumor necrosis factor ligand superfamily member 4 (TNFSF4), T-cell immunoglobulin domain and mucin domain-3 (TIM-3) and CD163 (Supplementary Figure S3A-B). 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Based on these findings, we performed mIHC staining using CRC tissues and found that the infiltration of CD8<sup>+</sup> T cells in CD36<sup>+</sup> CAF-rich regions was significantly reduced (Supplementary Figure S4A-B), while the infiltration of CD68<sup>+</sup> CD163<sup>+</sup> macrophages in CD36<sup>+</sup> CAF-rich regions was increased (Supplementary Figure S4C-D).</p><p>CD36, a scavenger receptor expressed in tumor cells [<span>5</span>], regulatory T cells [<span>6</span>], CD8<sup>+</sup> T cells [<span>7</span>], and cancer-associated macrophages [<span>8</span>], promotes tumor metastasis and induces an immunosuppressive phenotype in different cancer types [<span>5-8</span>]. Tumor cells with elevated CD36 expression exhibit a unique ability to initiate metastasis, and the presence of CD36<sup>+</sup> metastasis-initiating cells associates with a poor prognosis of numerous cancers [<span>9</span>]. However, a previous study demonstrated that CD36 repressed colorectal tumorigenesis by inhibiting glycolysis and that its expression was gradually reduced from adenomas to carcinomas [<span>10</span>]. In our study, we identified that CD36 was overexpressed both in CRC cells and tumor stroma by using multiple methods, and we revealed that high expression of CD36 in CRC indicated poor patient outcomes. In addition, we found that the presence of CD36<sup>+</sup> CAFs was associated with decreased CD8<sup>+</sup> T cell infiltration and increased CD68<sup>+</sup> CD163<sup>+</sup> macrophage infiltration in CRC, suggesting that CD36<sup>+</sup> CAFs were associated with the suppressive immune phenotype. However, whether and how CD36<sup>+</sup> CAFs affect these immune cells remains to be further investigated.</p><p>Overall, CD36<sup>+</sup> CAFs are a poor prognostic factor in CRC and are associated with immune cell infiltration. 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引用次数: 0

摘要

同样,之前的一些研究也表明,CD36+ CAFs 和 INHBA+ CAFs 可分别预测肝细胞癌(HCC)[3] 和胃癌患者[4] 的不良预后。从机制上讲,CD36+ CAFs 可促进 HCC 中免疫抑制微环境的形成 [3]。为了进一步研究 CD36 和 INHBA 是否参与了 CRC 的免疫微环境调控,我们分析了 TCGA-COAD 队列中的基因表达数据,发现 CD36 和 INHBA 的表达与免疫抑制因子和原发肿瘤性免疫细胞标志物的表达呈正相关、如 C-X-C motif趋化因子配体 12(CXCL12)、肿瘤坏死因子配体超家族成员 4(TNFSF4)、T 细胞免疫球蛋白结构域和粘蛋白结构域-3(TIM-3)和 CD163 的表达呈正相关(补充图 S3A-B)。同样,使用 CIBERSORT 方法估计免疫细胞浸润显示,M2 巨噬细胞浸润与 CD36 表达呈显著正相关,CD8+ T 细胞浸润与 INHBA 表达呈显著负相关(补充图 S3C-H)。基于这些发现,我们利用 CRC 组织进行了 mIHC 染色,发现 CD36+ CAF 富集区的 CD8+ T 细胞浸润显著减少(补充图 S4A-B),而 CD36+ CAF 富集区的 CD68+ CD163+ 巨噬细胞浸润增加(补充图 S4C-D)。CD36 是一种在肿瘤细胞[5]、调节性 T 细胞[6]、CD8+ T 细胞[7]和癌症相关巨噬细胞[8]中表达的清道夫受体,可促进肿瘤转移并在不同癌症类型中诱导免疫抑制表型[5-8]。CD36 表达增高的肿瘤细胞具有独特的转移启动能力,CD36+ 转移启动细胞的存在与多种癌症的不良预后有关 [9]。然而,之前的一项研究表明,CD36 通过抑制糖酵解抑制结直肠肿瘤的发生,而且从腺瘤到癌变,CD36 的表达逐渐减少 [10]。在我们的研究中,我们通过多种方法发现 CD36 在 CRC 细胞和肿瘤基质中均有过表达,并揭示了 CD36 在 CRC 中的高表达预示着患者的不良预后。此外,我们还发现 CD36+ CAFs 的存在与 CRC 中 CD8+ T 细胞浸润减少和 CD68+ CD163+ 巨噬细胞浸润增加有关,这表明 CD36+ CAFs 与抑制性免疫表型有关。总之,CD36+ CAFs是CRC的不良预后因素,并与免疫细胞浸润有关。我们的工作强调了鉴定肿瘤特异性 CAF 亚群对了解异质性肿瘤微环境的重要性。本研究获得了华西医院伦理委员会的批准(编号:2021-179)。JW和JLY设计了研究大纲;JW、MJX和BHX进行了实验;JW、QL和YZL分析了数据。JW和JLY撰写了手稿;JW、YZL和JLY对手稿进行了指导和编辑。所有作者阅读并批准了最终手稿。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Single-cell and bulk transcriptomics identifies a tumor-specific CD36+ cancer-associated fibroblast subpopulation in colorectal cancer

Single-cell and bulk transcriptomics identifies a tumor-specific CD36+ cancer-associated fibroblast subpopulation in colorectal cancer

Dear Editor,

Cancer-associated fibroblasts (CAFs) are highly versatile and plastic cells in the tumor microenvironment. They have been identified as actively involved in cancer progression and metastasis through their various roles in remodeling the extracellular matrix, suppressing the immune response and reprogramming tumor metabolism [1]. However, many challenges exist in revealing the functional phenotypes and mechanisms of CAFs in different cancers due to limited understanding of CAF heterogeneity [2]. Recent advances in single-cell transcriptome technology have enabled the identification of distinct CAF subpopulations by using unique gene signatures in multiple tumor types [2]. In this study, we successfully identified a tumor-specific CD36+ CAF subpopulation in colorectal cancer (CRC), which was found to be correlated with the number of tumor-infiltrated immune cells.

Primary CAFs and normal fibroblasts (NFs) were isolated from 5 fresh CRC tissues and paired normal colon tissues. Isolation methods and descriptions of other assays are shown in the Supplementary Methods. Immunofluorescence and Western blotting assays showed that the mesenchymal marker Vimentin was expressed in both NFs and CAFs, while alpha smooth muscle actin (αSMA) and fibroblast-specific protein 1 (FSP1) were overexpressed in CAFs (Supplementary Figure S1A-D). To further investigate gene expression profiles in these fibroblasts, primary NFs and CAFs were subjected to RNA sequencing. The results showed that CD36 was significantly upregulated in CAFs (Supplementary Figure S1E), which was further validated by immunofluorescence and Western blotting assays (Supplementary Figure S1F-G). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of differentially expressed genes between NFs and CAFs indicated significant enrichment in the mitogen-activated protein kinase (MAPK) signaling pathway, phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway and receptor-ligand activity (Supplementary Figure S1H-I), suggesting that CAFs may play a critical role in intracellular and extracellular signal transduction.

Previous studies have demonstrated that CAFs achieve high heterogeneity and plasticity across different cancer types [1, 2]. To further reveal the characteristics of CAFs in CRC, we performed single-cell RNA-sequencing (scRNA-seq) using isolated NFs and CAFs and detected 21,248 NFs and 18,097 CAFs (Figure 1A). A total of 7 clusters were identified in these fibroblasts by using sample integration analysis based on distinct gene expression signatures. Interestingly, we found that Clusters 3, 5, and 7 were mainly distributed in the CAF group, accounting for 34.5%, 7.7%, and 2.3% of total CAFs, respectively (Figure 1B-E). Cluster gene signature analysis showed that CD36 was expressed in CAF-specific subgroups, such as Clusters 3 and 5; inhibin subunit beta A (INHBA) was mainly expressed in Clusters 3, 5 and 7; whereas fibroblast growth factor 7 (FGF7) and alcohol dehydrogenase 1B (ADH1B) were mainly expressed in Clusters 1 and 2, which made up the majority of NFs (Figure 1F). To further visualize the marker genes of CAFs, we presented single gene expression data by uniform manifold approximation and projection (UMAP) and found that classical CAF markers, such as fibronectin 1 (FN1), fibroblast activation protein (FAP) and actin alpha 2 (ACTA2), cannot differentiate NFs and CAFs well, whereas a gene expression panel composed of CD36, INHBA, forkhead box S1 (FOXS1), keratin 18 (KRT18) and latent transforming growth factor β-binding protein 1 (LTBP1) was specifically expressed in CAFs (Figure 1G), suggesting the possibility that these genes could serve as new CAF markers in CRC.

To confirm the scRNA-seq results, we conducted multiplex immunohistochemistry (mIHC) staining and found that CD36 was overexpressed in both tumor cells and tumor stroma, especially in αSMA+ fibroblasts (Figure 1H-I). CD36 staining using a CRC tissue microarray showed similar results (Supplementary Figure S2A-B). Survival analysis showed that higher expression of CD36 in tumor cells and stromal cells indicated poorer prognosis in CRC patients (Supplementary Figure S2C). In addition, we also performed survival analyses using gene expression data from The Cancer Genome Atlas-Colon Adenocarcinoma (TCGA-COAD) cohort and found that a panel of CAF marker genes [INHBA, FOXS1, periostin (POSTN), thrombospondin 2 (THBS2)] also indicated a poor prognosis in CRC patients (Supplementary Figure S2D-E).

Overall, we identified a CD36+ CAF subpopulation in CRC and suggested the potential of CD36 as a specific CAF marker. Moreover, the survival analysis indicated that CAF marker genes (CD36, INHBA, FOXS1, POSTN, THBS2) may be promising prognostic indicators for CRC patients. Similarly, some previous studies have also indicated that CD36+ CAFs and INHBA+ CAFs may be predictors of a poor prognosis for hepatocellular carcinoma (HCC) [3] and gastric cancer patients [4], respectively. Mechanically, CD36+ CAFs promote the formation of an immunosuppressive microenvironment in HCC [3]. To further investigate whether CD36 and INHBA are involved in immune microenvironment modulation in CRC, we analyzed gene expression data from the TCGA-COAD cohort and found that the expression of CD36 and INHBA was positively correlated with the expression of immunosuppressive factors and protumorigenic immune cell markers, such as C-X-C motif chemokine ligand 12 (CXCL12), tumor necrosis factor ligand superfamily member 4 (TNFSF4), T-cell immunoglobulin domain and mucin domain-3 (TIM-3) and CD163 (Supplementary Figure S3A-B). In agreement, estimation of the immune cell infiltration using the CIBERSORT method showed a significant positive correlation between M2 macrophage infiltration and CD36 expression as well as a significant negative correlation between CD8+ T cell infiltration and INHBA expression (Supplementary Figure S3C-H). Based on these findings, we performed mIHC staining using CRC tissues and found that the infiltration of CD8+ T cells in CD36+ CAF-rich regions was significantly reduced (Supplementary Figure S4A-B), while the infiltration of CD68+ CD163+ macrophages in CD36+ CAF-rich regions was increased (Supplementary Figure S4C-D).

CD36, a scavenger receptor expressed in tumor cells [5], regulatory T cells [6], CD8+ T cells [7], and cancer-associated macrophages [8], promotes tumor metastasis and induces an immunosuppressive phenotype in different cancer types [5-8]. Tumor cells with elevated CD36 expression exhibit a unique ability to initiate metastasis, and the presence of CD36+ metastasis-initiating cells associates with a poor prognosis of numerous cancers [9]. However, a previous study demonstrated that CD36 repressed colorectal tumorigenesis by inhibiting glycolysis and that its expression was gradually reduced from adenomas to carcinomas [10]. In our study, we identified that CD36 was overexpressed both in CRC cells and tumor stroma by using multiple methods, and we revealed that high expression of CD36 in CRC indicated poor patient outcomes. In addition, we found that the presence of CD36+ CAFs was associated with decreased CD8+ T cell infiltration and increased CD68+ CD163+ macrophage infiltration in CRC, suggesting that CD36+ CAFs were associated with the suppressive immune phenotype. However, whether and how CD36+ CAFs affect these immune cells remains to be further investigated.

Overall, CD36+ CAFs are a poor prognostic factor in CRC and are associated with immune cell infiltration. Our work highlights the importance of identifying tumor-specific CAF subpopulations in understanding the heterogeneous tumor microenvironment.

This work was supported by grants from the National Natural Science Foundation of China (No. 82173253), the Sichuan Province Science and Technology Support Program (No. 2022YFH0003 and No. 2023NSFSC1900), and the Postdoctoral Research Foundation of China (No. 2022M712260).

The authors disclose no conflicts.

Not applicable.

This study was approved by the Ethics Committee of West China Hospital (No. 2021-179). All human samples were obtained with informed content.

JW and JLY designed the outline; JW, MJX, and BHX performed the experiments; JW, QL, and YZL analyzed the data. JW and JLY wrote the manuscript; JW, YZL and JLY supervised and edited the manuscript. All authors read and approved the final manuscript.

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来源期刊
Cancer Communications
Cancer Communications Biochemistry, Genetics and Molecular Biology-Cancer Research
CiteScore
25.50
自引率
4.30%
发文量
153
审稿时长
4 weeks
期刊介绍: Cancer Communications is an open access, peer-reviewed online journal that encompasses basic, clinical, and translational cancer research. The journal welcomes submissions concerning clinical trials, epidemiology, molecular and cellular biology, and genetics.
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