Single-cell transcriptomics reveals distinct inflammation-induced microglia signatures

Sousa, Carole; Golebiewska, Anna; Poovathingal, Suresh K; Kaoma, Tony; Pires-Afonso, Yolanda; Martina, Silvia; Coowar, Djalil; Azuaje, Francisco; Skupin, Alexander; Balling, Rudi; Biber, Knut; Niclou, Simone P; Michelucci, Alessandro

3. Fig. 2 : a. The authors should show the gene expression of more inflammatory genes, especially upon LPS treatment in vitro. Here, the focus lies too much on homeostatic genes. b. Figs. 2B, 2C and 2D: Isolated monocytes and macrophages should be included as a control. c. Fig. 2E : The relevance of the putative regulation by TGFβ for the following parts of the manuscript is unclear. d. In vitro experiments could also be used to validate findings from the sequencing experiments (see above) 4. Fig. S2 : The inclusion of these data, serving as a putative control of TGFβ as a regulator of the homeostatic function of these cells is relevant information. Yet, the text explains only a fragment of the shown treatments. Why are the treatments included in the study? 5. Fig. 3 : a. How many mice were used for the isolation of microglia and respective subsequent single cell RNASeq? The authors need to mention it in the legend b. The labelling of the heatmap is too small. Showing it without the gene symbols is sufficient. c. The volcano plot is not necessary. d. Descriptions of the -in part very general -results of the gene set enrichment are confusing. 6 . Fig. 4 : a. The authors should label the graphs with a heading to facilitate the understanding of the data shown (e.g. Fig. 4C : "upregulated genes"). b. The whole paragraph in the text can be shortened. profiles between acute inflammatory microglia and Alzheimer's disease-associated microglia on a single cell level. The article presents a useful resource for identifying microglia-specific genes in inflammation. However, many of the described results, such as the downregulation of microglia-specific homeostatic genes during inflammation, have been described previously. In order to increase the significance of this work, the authors should extend their analysis and add additional experiments.