Role of Fibroblasts in Glioblastoma

Glioblastoma (GBM) is a type of brain cancer that is difficult to treat due to its complex interaction with its surrounding microenvironment. While previous studies have focused on the role of immune and endothelial cells in this microenvironment, the potential role of cancer-associated fibroblasts (CAFs) has been largely overlooked. While some studies have suggested the presence of CAFs in GBM, they have not comprehensively profiled these cells or determined their effects on GBM and its microenvironment.

To address this gap in knowledge, researchers used a method called serial trypsinization to isolate those cancer-associated fibroblasts from GBM tissue samples and analyzed them at a genetic level to confirm their identity. They then studied the effects of these cells on GBM cells and their surrounding environment both in laboratory cultures and in live animal models. Indeed, this research could provide valuable insights into the role of CAFs in GBM and may help inform future treatment strategies.

VAMPIRE analysis, machine learning and scRNA-seq

Researchers used a modified visually aided morpho-phenotyping recognition (VAMPIRE) analysis and a machine-learning logistic regression classifier to identify and quantify cancer-associated fibroblasts (CAFs) in newly diagnosed glioblastoma (GBM) patient samples. By performing serial trypsinization on GBM samples, the researchers were able to identify CAF-like cells resistant to trypsinization, and bulk RNA-Seq analysis showed that these cells exhibited a transcriptomic profile similar to that of breast CAFs.

Single-cell RNA-Seq analysis also revealed that 86.5% of these cells expressed at least 1 of 9 CAF markers, including ACTA2, FAP, PDGFRA, PDGFRB, PDPN, S100A4, TNC, VIM, and COL1A1. Overall, the study supports the hypothesis that these cells are GBM CAFs. Furthermore, the researchers analyzed scRNA-Seq results from 12 patient GBMs and used clustering algorithms to identify 18 robust cell clusters, including CAFs, in each of the patient GBMs.

CAFs promote tumor growth in glioblastoma through activation of HIF-1α pathway and upregulation of cancer progression pathways

Let's take a look at the results of the study. The researchers found that CAFs mostly belonged to the fully differentiated subtype and were spatially correlated with the mesenchymal-like signature, M2 protumoral macrophages and glioblastoma stem cell markers. GBM CAFs activated the HIF-1α signaling pathway and upregulated cancer progression pathways, including EMT and cell proliferation in GSCs, which increased the frequency of GSCs and neurosphere formation.

In addition, the researchers report that CAFs are more abundant in the GBM subventricular zone (SVZ), which is home to neural stem cells thought to give rise to GBM stem cells. CAFs promote tumor growth by supporting self-renewal and differentiation of GBM stem cells. The addition of CAFs to GBM neurospheres caused tumors to grow faster in vivo, whereas this effect does not occur in normal fibroblasts. The results suggest that targeting CAFs may be a promising therapeutic strategy for GBM.

At a glance: discovery of cancer-associated fibroblasts in glioblastoma enhances understanding of aggressive brain cancer

In summary, researchers have discovered that CAFs also exist in glioblastoma (GBM), a highly aggressive brain cancer. While there has been debate among researchers about whether CAFs exist in GBM, this study used serial trypsinization of GBM cells to isolate and identify CAF-like cells. The researchers found that these CAF-like cells existed in multiple subtypes. One of them promoted the growth of glioma stem cells, a rare cell type that contributes to GBM therapeutic resistance.

Additionally, the CAF-produced extra domain A splice variant of fibronectin was found to promote M2 macrophage polarization and shift GBM vasculature to a larger, hypertrophied phenotype. These results suggest that CAFs alter the GBM microenvironment through multiple mechanisms that contribute to the aggressive biology and treatment refractoriness of GBM.

The authors state:

Overall, our findings provide compelling evidence that GBM CAFs promote GBM growth, insight that can be exploited for therapeutic benefit.
Jain S, Rick JW, Joshi RS, et al. Single-cell RNA sequencing and spatial transcriptomics reveal cancer-associated fibroblasts in glioblastoma with protumoral effects. The Journal of Clinical Investigation. 2023 Mar;133(5):e147087. DOI: 10.1172/jci147087 .