Stem Cell Research

Expression of LGR5 RNA (brown dots) in human colorectal cancer tissue, RNA in situ hybridization (ISH) using RNAscope<sup>®</sup> 2.0 HD Reagent Kit-BROWN.

Expression of LGR5 RNA (brown dots) in human colorectal cancer tissue, RNA in situ hybridization (ISH) using RNAscope® 2.0 HD Reagent Kit-BROWN.

Detection of Olfm4 crypt base columnar (CBCs) stem cells with RNA in situ hybridization (ISH) using RNAscope<sup>®</sup> 2.0 HD Reagent Kit-BROWN.

Detection of Olfm4 crypt base columnar (CBCs) stem cells with RNA in situ hybridization (ISH) using RNAscope® 2.0 HD Reagent Kit-BROWN.

The rapid and expansive field of stem cell biology has demonstrated the remarkable capabilities of these cells to self-renew, differentiate and reprogram. Yet the field continues to expand with studies trying to elucidate stem cell populations, characterize stem cell markers and identify the signals secreted from stem cells. RNAscope® in situ hybridization (ISH) technology enables cell-specific localization of RNA transcripts quickly and precisely for markers of stem cell populations.
The RNAscope® assay can be used to

  • Identify, characterize and locate stem cell populations

  • Reveal markers of stem cell maintenance and regeneration

  • Identify long non-coding RNAs in stem cells

  • Detect stem cell markers when no reliable antibodies are available

In a recent publication, Baker et. al. characterize LGR5-stem cells in colorectal adenomas and carcinomas. This study not only sheds light on the origins of serrated adenomas, but also implicates LGR5 mRNA as an alternative marker for discerning between conventional and serrated colonic adenomas. Use of the RNAscope® assay allows for detection of LGR5 expression in human samples that would otherwise be impossible due to the lack of adequate and reliable antibodies for LGR5.

The identification of the precise cells that produce Wnt in bone had been hindered due to the lack of appropriate tools. With the use of RNAscope®, Tan et. al. were able to examine expression of 19 Wnt genes in bone. The authors not only confirmed that the relative levels of Wnt expression are similar to transcriptome profiling studies, but were also able to provide new information on the cellular distribution of each of the 19 Wnt genes examined in bone. Because of the high resolution capabilities of RNAscope® technology, the authors were able to identify the precise anatomical regions where Wnt and Wnt target genes are expressed in bone. These findings contributed to the overall conclusion that Osterix-expressing cells produce their own Wnt that then activate Wnt signaling within these same cells, and in turn regulate their own proliferation and differentiation.

The mechanism for maintaining a stem cell niche is all about boundaries, involving the formation and maintenance of lineage-specific cellular and molecular boundaries. A 2013 publication by Barry et. al. in the January issue of Nature demonstrates how regulation of Wnt signaling by YAP is essential in the formation and maintenance of intestinal stem cells (ISC) and has implications for how YAP regulates the regenerative response. To determine if YAP expression was reducing ISC numbers, the authors used RNAscope® in situ assays to perform in situ hybridization (ISH) for Olfm4, which marks crypt base columnar (CBCs) stem cells.

Previously, Yan et al. used the RNAscope® in situ assays Olfm4 Assay to identify novel biomarkers demarcating two functionally distinct ISC populations in the intestinal crypt. These authors showed that Lgr5+ and Bmi+ ISCs demonstrate significant differences in basal proliferation status, response to canonical Wnt pathway modulation and response to acute radiation injury.

RNAscope® in situ hybridization assays are ideal tools to visualize stem cell-specific gene expression. The exquisite sensitivity and specificity of RNAscope® in situ assays make it a powerful technique for molecular identification and characterization of rare stem cell populations in complex tissues that can easily be viewed with any standard brightfield microscope. With RNAscope® in situ hybridization, you can quickly and easily visualize molecular markers for stem cells such as Lgr5, Bmi1 and Olfm4 at single-molecule resolution in routine clinical FFPE specimens.

Recorded Webinar

Watch recorded webinar to learn more about the ACD study: Visualization of LGR5+ Stem Cells and the immune response in the inflamed mouse colon with RNAscope® Assay presented by Courtney Anderson, Senior Scientist at ACD.

Watch Webinar

Application Review

Cellular localization of RNA Expression in Stem Cells using RNAscope® Technology

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