Novel genes and pathways modulated by syndecan-1: implications for the proliferation and cell-cycle regulation of malignant mesothelioma cells

Our plain-English summary of Novel genes and pathways modulated by syndecan-1: implications for the proliferation and cell-cycle regulation of malignant mesothelioma cells, by T. Szatmári, F. Mundt, G. Heidari-Hamedani, et al (PloS One, 2012).

The study

• Authors: T. Szatmári, F. Mundt, G. Heidari-Hamedani, F. Zong, E. Ferolla, A. Alexeyenko, A. Hjerpe, K. Dobra
• Journal: PloS One
• Published: 2012;7(10):e48091
• Cited by: 37 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Malignant pleural mesothelioma is a highly malignant tumor, originating from mesothelial cells of the serous cavities. In mesothelioma the expression of syndecan-1 correlates to epithelioid morphology and inhibition of growth and migration. Our previous data suggest a complex role of syndecan-1 in mesothelioma cell proliferation although the exact underlying molecular mechanisms are not completely elucidated. The aim of this study is therefore to disclose critical genes and pathways affected by syndecan-1 in mesothelioma; in order to better understand its importance for tumor cell growth and proliferation. We modulated the expression of syndecan-1 in a human mesothelioma cell line via both overexpression and silencing, and followed the transcriptomic responses with microarray analysis. To project the transcriptome analysis on the full-dimensional picture of cellular regulation, we applied pathway analysis using Ingenuity Pathway Analysis (IPA) and a novel method of network enrichment analysis (NEA) which elucidated signaling relations between differentially expressed genes and pathways acting via various molecular mechanisms. Syndecan-1 overexpression had profound effects on genes involved in regulation of cell growth, cell cycle progression, adhesion, migration and extracellular matrix organization. In particular, expression of several growth factors, interleukins, and enzymes of importance for heparan sulfate sulfation pattern, extracellular matrix proteins and proteoglycans were significantly altered. Syndecan-1 silencing had less powerful effect on the transcriptome compared to overexpression, which can be explained by the already low initial syndecan-1 level of these cells. Nevertheless, 14 genes showed response to both up- and downregulation of syndecan-1. The "cytokine - cytokine-receptor interaction", the TGF-β, EGF, VEGF and ERK/MAPK pathways were enriched in both experimental settings. Most strikingly, nearly all analyzed pathways related to cell cycle were enriched after syndecan-1 silencing and depleted after syndecan-1 overexpression. Syndecan-1 regulates proliferation in a highly complex way, although the exact contribution of the altered pathways necessitates further functional studies.

In plain English

This laboratory study used one human malignant mesothelioma cell line, called STAV-AB. These cells naturally make little syndecan-1, a protein found on the cell surface. The authors raised syndecan-1 levels in some cells and lowered ('silenced') it in others. They then measured changes in the activity of thousands of genes and analysed which biological pathways were affected. The study did not examine asbestos exposure. Raising syndecan-1 changed far more genes than lowering it: with a 2-fold cut-off, 1,124 genes versus 21. Fourteen genes changed in both experiments. In silenced cells compared with control cells, growth fell, doubling time rose from 21.6 to 27.9 hours, and more cells sat in the G0/G1 phase of the cell cycle. Apoptosis (a form of cell death) did not change significantly. In network analysis, cell-cycle pathways were enriched after silencing, while many of the same pathways were depleted after overexpression. The authors concluded that syndecan-1 helps maintain a delicate balance regulating cell proliferation. They noted that they used only one cell line and that further functional studies are needed.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: laboratory experiment on a cultured cell line: syndecan-1 was overexpressed or silenced, gene activity was measured by microarray, and results were analysed with pathway and network methods (GSEA, Ingenuity Pathway Analysis, network enrichment analysis), plus growth, cell-cycle and apoptosis assays
• Who was studied: one human malignant mesothelioma cell line (STAV-AB), with epithelioid morphology and low natural syndecan-1, grown in the laboratory at Karolinska Institutet, Stockholm; no patients were studied
• Exposure: No asbestos or fiber exposure was studied. The experimental change was in syndecan-1 level: stable overexpression using a plasmid carrying the full-length human syndecan-1 gene (control: empty vector), and silencing with three syndecan-1-specific siRNA constructs (control: scrambled siRNA)
• Main results:
  • With a 2-fold change cut-off, expression of 1,124 genes was significantly altered in syndecan-1 overexpressing cells, but only 21 genes in syndecan-1 silenced cells
  • With a 1.5-fold cut-off, 2,389 genes were differentially expressed in overexpressing cells and 103 in silenced cells
  • Fourteen genes were altered by both overexpression and silencing
  • Silencing reduced syndecan-1 mRNA by 90 to 95% and syndecan-1 protein by 43%, measured 24 hours after transfection and compared with scrambled control
  • Cell proliferation significantly decreased in silenced cells. The article describes the effect as 25% of the control at 24 hours and 40% of the control at 48 hours
  • Doubling time rose from 21.6 hours in scrambled control cells to 27.9 hours in syndecan-1 silenced cells
  • In silenced cells, the proportion of cells in G2/M was significantly reduced after 24 hours, and more cells were in G0/G1, compared with scrambled control
  • No significant difference in apoptosis between silenced cells and scrambled control
  • In overexpressing cells, 51 of 783 proliferation-related genes on the chip were downregulated and 74 were upregulated
  • In overexpressing cells, 99 cell-cycle regulation genes were differentially expressed, most of them downregulated
  • In overexpressing cells, IL33, IL6 and IL8 were increased more than 10-fold
  • In network enrichment analysis of more than 1,600 pathways, 939 were significantly altered in overexpressing cells and 234 in silenced cells
  • Cell-cycle pathways were enriched when syndecan-1 was silenced, and many of the same pathways were depleted when it was overexpressed
  • cdc42-related pathways were enriched whether syndecan-1 was raised or lowered, although the cdc42 gene itself was not differentially regulated
  • Ingenuity Pathway Analysis found five pathways significantly altered by both overexpression and silencing: IL10, IL6, HGF, ERK5 and ERK/MAPK signaling
  • At the protein level, 7 of 11 proteins tested showed changes in the same direction as the microarray results
• The authors' conclusion: The authors concluded that syndecan-1 is important in maintaining a delicate balance that regulates cell proliferation in these mesothelioma cells. They found that the gene changes, both increases and decreases, ultimately led to reduced proliferation
• Limitations the authors state:
  • Only one cell line was used, so whether these pathways apply generally needs further studies
  • The contribution of each altered pathway needs further functional studies
  • Values from the confirmatory RT-PCR tests differed in some cases from the microarray values, although the direction of change agreed

Topics

Medical terms in this study, explained in our Lexicon: Malignant Mesothelioma, Pleural Mesothelioma, Mesothelial Cells, Serous Cavities, Mesothelioma, Epithelioid, Interleukins, Cytokine.

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