Molecular Landscape of the Mouse Adrenal Gland and Adjacent Adipose by Spatial Transcriptomics
Curation statements for this article:-
Curated by eLife
eLife Assessment
This useful study provides a spatial transcriptomic analysis of the mouse adrenal gland that could have implications for future research and applications. The authors present solid results that allow the dissection of the cell signalling pathways and cellular composition of different zones of the adrenal glands in the mouse model; they propose new zone-specific gene markers and specific intra- and inter-zonal signaling pathways based on receptor-ligand expression patterns. Their web tool is user-friendly and will be helpful for adrenal scientists; however, the validation of crucial results of the large dataset is necessary. There are also several contradictory results/interpretations, and the opportunity to dissect the sexually dimorphic gene expression pattern and mouse-human interspecies differences is a missed opportunity.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
Abstract
The adrenal glands play a vital role in maintaining homeostasis and managing stress through the production and secretion of steroid hormones. Despite extensive research, the molecular mechanisms underlying adrenal zonation and cellular differentiation remain poorly understood. By employing spatial transcriptomics, this study has mapped the adult CD1 IGS mouse adrenal gland, thereby identifying unique genetic markers of zonal differentiation and dynamic cellular interactions. Five cellular clusters, corresponding to the cortex and medulla compartments, were identified, along with two adipose tissue clusters (brown and white). These findings confirm the centripetal differentiation model, highlighting the gradual transition of cell populations from the capsule through cortical zones. Through ligand-receptor interaction analysis, a complex regulatory network governing inter- and intra-zone communication was identified, thereby emphasising the adrenal gland’s central role in integrating endocrine and neuroendocrine signals, particularly in response to stress. This comprehensive spatial transcriptomic map of the adult mouse adrenal gland provides original insights into adrenal biology and constitutes a valuable resource for future research.
Article activity feed
-
eLife Assessment
This useful study provides a spatial transcriptomic analysis of the mouse adrenal gland that could have implications for future research and applications. The authors present solid results that allow the dissection of the cell signalling pathways and cellular composition of different zones of the adrenal glands in the mouse model; they propose new zone-specific gene markers and specific intra- and inter-zonal signaling pathways based on receptor-ligand expression patterns. Their web tool is user-friendly and will be helpful for adrenal scientists; however, the validation of crucial results of the large dataset is necessary. There are also several contradictory results/interpretations, and the opportunity to dissect the sexually dimorphic gene expression pattern and mouse-human interspecies differences is a missed …
eLife Assessment
This useful study provides a spatial transcriptomic analysis of the mouse adrenal gland that could have implications for future research and applications. The authors present solid results that allow the dissection of the cell signalling pathways and cellular composition of different zones of the adrenal glands in the mouse model; they propose new zone-specific gene markers and specific intra- and inter-zonal signaling pathways based on receptor-ligand expression patterns. Their web tool is user-friendly and will be helpful for adrenal scientists; however, the validation of crucial results of the large dataset is necessary. There are also several contradictory results/interpretations, and the opportunity to dissect the sexually dimorphic gene expression pattern and mouse-human interspecies differences is a missed opportunity.
-
Reviewer #1 (Public review):
Summary:
This study employs spatial transcriptomics to explore the molecular architecture of the adult mouse adrenal gland and the adjacent adipose tissue. The research aimed to identify zonation-specific genetic markers, elucidate cellular differentiation patterns, and investigate inter- and intra-zone communication within the adrenal gland. The findings support the centripetal differentiation model, highlighting the transition of cell populations across different cortical zones. The study also integrates ligand-receptor interaction analysis to uncover the adrenal gland's role in endocrine and neuroendocrine signaling, particularly in stress response. This high-resolution spatial transcriptomic map provides novel insights into adrenal gland biology and is a resource for further investigations.
Strengths:
The…
Reviewer #1 (Public review):
Summary:
This study employs spatial transcriptomics to explore the molecular architecture of the adult mouse adrenal gland and the adjacent adipose tissue. The research aimed to identify zonation-specific genetic markers, elucidate cellular differentiation patterns, and investigate inter- and intra-zone communication within the adrenal gland. The findings support the centripetal differentiation model, highlighting the transition of cell populations across different cortical zones. The study also integrates ligand-receptor interaction analysis to uncover the adrenal gland's role in endocrine and neuroendocrine signaling, particularly in stress response. This high-resolution spatial transcriptomic map provides novel insights into adrenal gland biology and is a resource for further investigations.
Strengths:
The study, using the latest technologies and methods such as Visium CytAssist technology, UMAP & Seurat analysis, Gene Ontology (GO) & KEGG pathway enrichment analysis, Monocle3, and CellChat analysis, performed three-dimensional analysis, which has been challenging to achieve using the two-dimensional transcriptomics that have been commonly used up until now.
The unique gene expression patterns were demonstrated for each adrenal zone. Spatial transcriptomics confirmed unique gene expression patterns for each adrenal zone (ZG, ZF, ZX, medulla). The centripetal differentiation model shows the migration of the progenitor cells from the adrenal capsule towards the inner cortex. Key genetic markers were identified in each adrenal zone and adjacent adipose tissues. In addition, CellChat analysis identified major signaling pathways, including Wnt signaling, Hedgehog signaling, IGF2-IGF2R interactions, and Neuropeptide Y (NPY) signaling in the medulla. All these results offer a valuable dataset for future adrenal biology research, with potential applications in disease modeling and therapeutic target identification.
The results, high-resolution mapping of adrenal gland zonation, validation of the centripetal differentiation model, perspective on cell-cell communication, and potential translational impact on human adrenal gland function and disorders, are quite noble.
Weaknesses:
The reviewer requests that the following issues be addressed in the text:
(1) The study focuses only on adult male mice, which limits insights into developmental and sex-specific differences. What do the authors predict about the gender and age difference?
(2) Despite advanced methodologies, single-cell heterogeneity may not be fully captured, as Visium technology has limited spatial resolution.
(3) While the study suggests that ZX might have a role in androgen synthesis, further functional validation is required.
(4) The study is primarily descriptive, lacking in-depth mechanistic experiments to validate cell-cell communication interactions. It is quite interesting to suggest cell-cell communication, but the authors are still required to provide some evidence to support it.
(5) The data supports the conclusions, particularly in validating the centripetal differentiation model using Monocle3 trajectory analysis. However, functional validation experiments (e.g., gene knockout studies) would strengthen the findings, especially regarding ZX function and ligand-receptor interactions.
-
Reviewer #2 (Public review):
This study by M. Blatkiewicz et al. seeks to define the spatial gene expression pattern of the adult male mouse adrenal gland using current spatial transcriptomic techniques. They propose new zone-specific gene markers and specific intra- and inter-zonal signaling pathways based on receptor-ligand expression patterns. Their web tool is user-friendly and will be helpful for adrenal scientists. The manuscript is easy to follow, but validation of crucial results of the large dataset is missing. There are also several contradictory results/interpretations, and the opportunity to dissect the sexually dimorphic gene expression pattern and mouse-human interspecies differences is a missed opportunity.
(1) The authors used 10-week-old CD1 male mouse adrenal glands to assess the spatial transcriptomics of the adrenal …
Reviewer #2 (Public review):
This study by M. Blatkiewicz et al. seeks to define the spatial gene expression pattern of the adult male mouse adrenal gland using current spatial transcriptomic techniques. They propose new zone-specific gene markers and specific intra- and inter-zonal signaling pathways based on receptor-ligand expression patterns. Their web tool is user-friendly and will be helpful for adrenal scientists. The manuscript is easy to follow, but validation of crucial results of the large dataset is missing. There are also several contradictory results/interpretations, and the opportunity to dissect the sexually dimorphic gene expression pattern and mouse-human interspecies differences is a missed opportunity.
(1) The authors used 10-week-old CD1 male mouse adrenal glands to assess the spatial transcriptomics of the adrenal gland. As they also mentioned, male mice typically lose their zone-X after puberty (around 6-8 weeks of age). However, their analysis in 10-week-old mice suggests that zone-X covers most of the adrenal cortex. As shown in Figure 3A, the dots between the zona glomerulosa and the medulla are mostly positive for zone-X, which would suggest that the zona fasciculata represents a relative minority of the overall adult adrenal cortex. Is this correct? Is the presence of zone-X in sexually mature adult male mice unique to the CD1 strain? Providing histology data in support of this conclusion, using zone-specific markers combined with RNA in situ hybridization or immunofluorescence techniques in the CD1 male adrenal gland, would help to interpret these data further. Given the relatively low resolution of their gene expression profiles, it is possible there is overlap between the zona fasciculata and the zone-X.
(2) The pseudotime trajectory analysis confirms prior reports in the literature showing zonal transdifferentiation but does not provide novel insight. It would be nice to know what gene expression patterns correlate (positively or negatively) based on an unbiased analysis.
(3) The authors suggest that they identified new zonal markers, but it would be nice to see confirmation of some of these markers (e.g., Frmpd4, Oca2, Sphkap for the ZG or Cited1, Nat8f5 for the ZF, etc. ) with in situ or immunofluorescence combined with known markers such as Dab2, Cyp11b2, or Cyp11b1.
(4) The authors mention a gradual transition between the zones. It would be interesting to know whether transition zones exist between the zona glomerulosa and the zona fasciculata or the zona fasciculata and the zone-X.
(5) The authors note using Visium cyst assist, but they do not discuss the advantages of this system compared to other systems. Explanation of the approximate resolution of their analysis (e.g., how many cells were pooled in the wells) would help readers to interpret their data. It would also be nice to compare it to other spatial transcriptomic analyses of human adrenals, given the differences between the zonation of human and mouse adrenals.
(6) Interestingly, CellChat analysis suggests possible communication between the medulla and the zona fasciculata and zona glomerulosa. How do the authors explain the transfer of these molecules from the medulla to the outer zones given centripetal blood flow in the adrenal? Also, how does the fact that Igf2 expression has been shown to be expressed in the capsule (PMID: 22266195) affect the interpretation of their data?
(7) The study misses the opportunity to dissect sexually dimorphic gene expression patterns in the mouse adrenal. For example, the authors could have focused on the role of stem cells between male and female mouse adrenals, which have been reported to differ (PMID: 31104943). In addition, the authors could have focused on the sexually dimorphic zone-X and its regulation by sex hormone signaling.
(8) The capsule is classified as a connective tissue, which may be misleading given its important role as a signaling center in the adrenal. Genes enriched in typical connective tissues do not include many of the genes that seem to define the adrenal capsule. Also, some of the capsule markers appear to be found in the zona glomerulosa. Is this a result of not being able to fully resolve the small layer of zG cells and the even smaller layer of capsular cells? Guided reclustering of the cells based on known markers and separation of capsule and connective tissue might help to present their data on adrenal zonation more clearly.
-
Reviewer #3 (Public review):
Summary:
In summary, the scientists used Visium spatial transcriptomics technology to create a thorough spatial transcriptomic atlas of the adult male mouse adrenal gland and the adipose tissues that surround it. Their primary goals were to map the cell communication network, determine the differentiation direction of various cell types, and find marker genes for various adrenal zones.
Strengths:
(1) Undoubtedly, one of the biggest strengths of the manuscript is a spatial transcriptomic o mouse adrenal gland tissue, which, to my knowledge, has not been done before.
(2) Comprehensive Zonal Characterization: Seven distinct clusters were identified, corresponding to known anatomical and functional regions (ZG, ZF, ZX, medulla, connective tissue, brown and white adipose tissue), each with robust marker gene sets.
Reviewer #3 (Public review):
Summary:
In summary, the scientists used Visium spatial transcriptomics technology to create a thorough spatial transcriptomic atlas of the adult male mouse adrenal gland and the adipose tissues that surround it. Their primary goals were to map the cell communication network, determine the differentiation direction of various cell types, and find marker genes for various adrenal zones.
Strengths:
(1) Undoubtedly, one of the biggest strengths of the manuscript is a spatial transcriptomic o mouse adrenal gland tissue, which, to my knowledge, has not been done before.
(2) Comprehensive Zonal Characterization: Seven distinct clusters were identified, corresponding to known anatomical and functional regions (ZG, ZF, ZX, medulla, connective tissue, brown and white adipose tissue), each with robust marker gene sets.
(3) The authors manage to integrate advanced bioinformatical tools such as CellChatDB, Monocle3, and CARD to study the relationship between cell types and differentiation of the tissue.
(4) The authors manage to identify novel marker genes for some adrenal zones.
Weaknesses:
(1) The study focused only on one adult male CD1 IGS mouse, which is a limiting factor for other strains, ages, or females, especially given the sexual dimorphism of the ZX. Although the authors claim that four slices of the adrenal gland have been processed on Visium and sequenced, for "clarity," they show only one, which might bias the results.
(2) Lack of detailed QC analysis of the Visium slide.
(3) The study misses the functional validation of the novel marker genes - this needs to be addressed.
(4) What worries me a lot is the fact that, actually, there might be more than one cell present within a Visium spot, so the only way to define zones is by anatomical observation rather than cellular composition.
(5) In cell chat analysis, the authors show the strength of the interactions, but miss out on the number of interactions.
Conclusions:
The authors' stated goals were mostly accomplished:
By mapping the mouse adrenal gland's molecular landscape, they were able to clearly establish unique molecular signatures for every anatomical zone.
Pseudotime study of the cell progression from the capsule through ZG, ZF, and ZX demonstrates that the data strongly support the centripetal differentiation concept. Conclusions on the functional importance of newly discovered marker genes are conjectural and need additional experimental support.
Nevertheless, several findings are still tentative and will need more experimental support, especially when it comes to the significance of ZX persistence and the functional involvement of recently discovered marker genes.
-
