PO.TB10.13 · 肿瘤生物学
基质细胞SPARC驱动胰腺导管腺癌中的肿瘤-神经-基质串扰及神经周围浸润
Stromal SPARC drives tumor-nerve-stroma crosstalk and perineural invasion in pancreatic ductal adenocarcinoma
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：神经周围浸润（PNI）在胰腺导管腺癌（PDAC）中频繁发生，并导致不良预后。然而，支撑肿瘤嗜神经性的基质-神经机制仍知之甚少。我们假设基质成纤维细胞分泌的可溶性因子与神经及癌细胞协同作用，驱动PNI相关的肿瘤侵袭性。
方法：建立垂直Transwell三培养系统，上室为人PDAC细胞（MIA PaCa-2），下室为新生小鼠背根神经节（DRG）与人胰腺星状细胞（PSC）。培养24 h后定量PDAC细胞迁移。对各条件（对照、DRG、PSC、DRG + PSC）的条件培养基采用基于强度的定量方法进行LC-MS/MS分析；候选蛋白通过与迁移的相关性（Pearson r ≥ 0.95）及在DRG + PSC中≥ 2倍富集进行筛选。利用TCGA-PDAC和GTEx数据集评估临床意义。通过CIBERSORTx反卷积和单细胞RNA测序分析推断细胞起源。在81例切除的PDAC中，通过免疫组化和半自动化QuPath评分评估基质与上皮SPARC蛋白表达及其与PNI的关联。
结果：与对照或任一单细胞条件相比，DRG + PSC三培养显著增强PDAC迁移（均p < 0.001）。蛋白质组学分析鉴定出富含半胱氨酸的酸性分泌蛋白（SPARC）为DRG + PSC条件下最富集的蛋白之一，其丰度与迁移活性密切相关。在转录组数据集中，SPARC在PDAC中相对于正常胰腺显著上调，且高SPARC表达与总生存期缩短相关。CIBERSORTx和单细胞分析将SPARC表达主要定位于成纤维细胞/癌症相关成纤维细胞（CAF）群体。在临床标本中，SPARC蛋白集中于促结缔组织增生的基质中，显著高于肿瘤上皮（p < 0.0001）。在我们的机构队列中，PNI阳性肿瘤的基质SPARC水平高于PNI阴性肿瘤（p < 0.0001），在TCGA-PDAC中结果一致（p = 0.032）。
结论：神经-基质调控的微环境加速PDAC细胞迁移，CAF来源的SPARC是肿瘤-神经串扰及PNI的关键驱动因素。通过细胞外基质重塑和促进神经浸润，SPARC成为PDAC中面向基质、抗PNI策略的有前景的生物标志物和治疗靶点。
查看英文原文 English abstract
Background: Perineural invasion (PNI) is frequent in pancreatic ductal adenocarcinoma (PDAC) and contributes to poor outcomes. However, the stromal-neural mechanisms that underlie tumor neurotropism remain poorly understood. We hypothesized that soluble factors secreted by stromal fibroblasts, in cooperation with nerves and cancer cells, drive PNI-related tumor aggressiveness.
Methods: A vertical transwell tri-culture system was established using human PDAC cells (MIA PaCa-2) in the upper chamber and neonatal mouse dorsal root ganglia (DRG) with human pancreatic stellate cells (PSCs) in the lower chamber. PDAC cell migration was quantified after 24 h. Conditioned media from each condition (control, DRG, PSC, DRG + PSC) were analyzed by LC-MS/MS with intensity-based quantification; candidate proteins were selected by correlation with migration (Pearson r ≥ 0.95) and ≥ 2-fold enrichment in DRG + PSC. Clinical significance was assessed using TCGA-PDAC and GTEx datasets. Cell-of-origin was inferred by CIBERSORTx deconvolution and single-cell RNA-seq analysis. Stromal versus epithelial SPARC protein expression and its association with PNI were evaluated by immunohistochemistry and semi-automated QuPath scoring in 81 resected PDACs.
Results: Tri-culture with DRG + PSC significantly enhanced PDAC migration compared with control or either single-cell condition (all p < 0.001). Proteomic profiling identified Secreted Protein Acidic and Cysteine-Rich (SPARC) as one of the most enriched proteins in DRG + PSC condition, with abundance strongly correlating with migratory activity. In transcriptomic datasets, SPARC was markedly upregulated in PDAC relative to normal pancreas, and high SPARC expression was associated with shorter overall survival. CIBERSORTx and single-cell analyses localized SPARC expression primarily to fibroblast/cancer-associated fibroblast (CAF) populations. In clinical specimens, SPARC protein was concentrated in the desmoplastic stroma and significantly higher than in tumor epithelium (p < 0.0001). Stromal SPARC levels were elevated in PNI-positive versus PNI-negative tumors in our institutional cohort (p < 0.0001), with consistent results in TCGA-PDAC (p = 0.032).
Conclusions: A nerve-stroma-conditioned microenvironment accelerates PDAC cell migration, with CAF-derived SPARC serving as a pivotal driver of tumor-nerve crosstalk and PNI. Through extracellular matrix remodeling and promotion of neural infiltration, SPARC emerges as a promising biomarker and therapeutic target for stroma-directed, anti-PNI strategies in PDAC.
利益披露 Disclosure
S. Furuhashi, None..
R. Muraki, None..
Y. Morita, None..
D. Nishiwaki, None..
A. Matsumoto, None..
Y. Shimizu, None..
T. Murakami, None..
M. Takeda, None..
H. Kikuchi, None..
Y. Hiramatsu, None..
H. Takeuchi, None.