Identification of a unique TGF-β–dependent molecular and functional signature in microglia

In the paper

“This lineage is mainly regulated by CSFR14 and its ligand, IL-347.”

What the sources say

• Partly supported

  The passage labeled Abstract reports that microglial development is strongly dependent on CSF-1R and identifies IL-34 as a CSF-1R ligand. It says IL-34 expression is consistent with an important role in microglial homeostasis, but does not establish that IL-34 mainly regulates the lineage. (LLM, conf=0.85)

  cites [4] Ginhoux, F. et al. Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science 330, 841–845 (2010). sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section (‘IL-34 Contributes to the Maintenance of Microglia in the Adult’) reports that microglia are absent in Csf1r-deficient mice and partially reduced in adult IL-34-deficient mice. This supports a role for CSF1R and IL-34, but the paper limits IL-34’s demonstrated role to region-specific maintenance of adult microglia, not regulation of the microglial lineage generally. (LLM, conf=0.85)

  cites [7] Greter, M. et al. Stroma-derived interleukin-34 controls the development and maintenance of langerhans cells and the maintenance of microglia. Immunity 37, 1050–1060 (2012). sourceExperimental — accuracy not yet measured

In the paper

“we found that these animals did not express TGF-β1 in the brain or spinal cord and we found no TGF-β1-producing T-cells in the CNS of those animals33.”

What the sources say

• Supported

  The Results section, “Generation of adult CNS-TGF-β1 deficient mice,” identifies the transgenic mice lacking both endogenous Tgf-β1 alleles and reports that they did not express TGF-β1 in the brain or spinal cord. The same passage reports no TGF-β1-producing T-cells in their CNS. (LLM, conf=0.95)

  cites [33] Koeglsperger, T. et al. Impaired glutamate recycling and GluN2B-mediated neuronal calcium overload in mice lacking TGF-beta1 in the CNS. Glia 61, 985–1002 (2013). source

In the paper

“adult microglia derive from primitive macrophages4 in a Myb-independent manner5 via PU.1 and IRF8 dependent pathways6.”

What the sources say

• Supported

  Citation 4 is attached to the claim that adult microglia derive from primitive macrophages, not to the separate Myb, PU.1, or IRF8 claims. The paper’s Abstract states that primitive myeloid precursors give rise to microglia in the adult CNS. (LLM, conf=0.93)

  cites [4] Ginhoux, F. et al. Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science 330, 841–845 (2010). source
• ?Not checked — no full text of the sourcecites [5] Schulz, C. et al. A lineage of myeloid cells independent of Myb and hematopoietic stem cells. Science 336, 86–90 (2012). source
• ?Not checked — no full text of the sourcecites [6] Kierdorf, K. et al. Microglia emerge from erythromyeloid precursors via Pu.1- and Irf8-dependent pathways. Nat. Neurosci. 16, 273–280 (2013). source

In the paper

“Our findings are consistent with reports that macrophage progenitors develop from the primitive ectoderm of the yolk sac4 and give rise to tissue macrophages that do not have a monocytic progenitor16,17.”

What the sources say

• Partly supported

  The paper’s Abstract reports that adult microglia derive from primitive myeloid progenitors and that postnatal hematopoietic progenitors do not significantly contribute to their maintenance. This supports the claim for microglia, but not for tissue macrophages generally; the paper also does not identify the yolk sac’s primitive ectoderm as their origin. (LLM, conf=0.88)

  cites [4] Ginhoux, F. et al. Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science 330, 841–845 (2010). sourceExperimental — accuracy not yet measured
• Supported

  The citation to this paper is attached to the claim that tissue macrophages can arise without a monocytic progenitor, not to the preceding claim about primitive ectoderm. The Discussion reports that monocytes are not the progenitors of several tissue-resident macrophage populations in the steady state, and the Introduction describes yolk-sac-derived macrophages maintained independently of monocytes. The paper is not being cited here for the ectoderm claim. (LLM, conf=0.91)

  cites [16] Hashimoto, D. et al. Tissue-resident macrophages self-maintain locally throughout adult life with minimal contribution from circulating monocytes. Immunity 38, 792–804 (2013). source
• Partly supported

  The Results ('Dual origins of resident macrophages'; page unavailable) find no ongoing steady-state monocyte contribution to several tissue macrophage populations, and the Discussion says these populations can persist without monocyte input in healthy adult mice. That supports monocyte-independent maintenance, but not the broader claim that these macrophages never have a monocytic progenitor. The yolk-sac ectoderm clause is attached to a different citation in the citing sentence. (LLM, conf=0.86)

  cites [17] Yona, S. et al. Fate mapping reveals origins and dynamics of monocytes and tissue macrophages under homeostasis. Immunity 38, 79–91 (2013). sourceExperimental — accuracy not yet measured

In the paper

“Linear discrimination scores were used to assign probabilities to each MS2 spectrum for being assigned correctly and these probabilities were further used to filter the dataset with an MS2 spectra assignment FDR of smaller than a 1% at the protein level58.”

What the sources say

• Partly supported

  The Results section describes using linear discriminant analysis to filter peptide identifications, supporting the method attributed to the paper. But the Experimental Procedures report a 1% peptide FDR and a 1.25% final protein FDR; the phosphopeptide dataset had a 1.7% protein FDR. Neither supports an FDR below 1% at the protein level, and the cited passages do not specify assigning a probability to each MS2 spectrum. (LLM, conf=0.90)

  cites [58] Huttlin, E.L. et al. A tissue-specific atlas of mouse protein phosphorylation and expression. Cell 143, 1174–1189 (2010). sourceExperimental — accuracy not yet measured

In the paper

“Recent work demonstrated that all tissue resident macrophages are different to an extent yet cluster together when compared to monocytes and DCs and that spleen red pulp macrophages were closely associated with microglia14.”

What the sources say

• Partly supported

  The Results sections on tissue macrophage diversity and co-expressed gene modules report differences among the four resident macrophage populations and a shared gene signature that distinguishes them from DCs and blood monocytes. They do not establish that all tissue-resident macrophages cluster together, or that splenic red pulp macrophages are especially closely associated with microglia. The paper instead says DCs clustered more closely than macrophages. Page numbers are unavailable in our copy. (LLM, conf=0.86)

  cites [14] Gautier, E.L. et al. Gene-expression profiles and transcriptional regulatory pathways that underlie the identity and diversity of mouse tissue macrophages. Nat. Immunol. 13, 1118–1128 (2012). sourceExperimental — accuracy not yet measured

In the paper

“Consistent with this, deficits in microglia function have been reported to contribute to synaptic abnormalities which may be present in neuro-developmental disorders41.”

What the sources say

• Supported

  The Abstract says that deficits in microglia function may contribute to synaptic abnormalities seen in some neurodevelopmental disorders. The paper’s conclusion also reports delayed synaptic pruning and excess immature synapses in mice with reduced microglia numbers, supporting the proposed link without claiming it was demonstrated in people. (LLM, conf=0.94)

  cites [41] Paolicelli, R.C. et al. Synaptic pruning by microglia is necessary for normal brain development. Science 333, 1456–1458 (2011). source

In the paper

“We previously reported that CD39 is expressed on adult microglia and can distinguish recruited Ly6C+ monocytes from resident microglia12.”

What the sources say

• Supported

  In the section on reciprocal CD39 and Ly6C expression, the paper reports CD39 expression in microglia from naive adult mouse brains and says that CD39 and Ly6C distinguish resident microglia from peripheral inflammatory monocytes. The Discussion likewise states that CD39 distinguished resident microglia from infiltrating monocytes. (LLM, conf=0.94)

  cites [12] Butovsky, O. et al. Modulating inflammatory monocytes with a unique microRNA gene signature ameliorates murine ALS. J. Clin. Invest. 122, 3063–3087 (2012). source

In the paper

“Tgfb1−/− animals die at 3 weeks of age of a severe multifocal autoinflammatory disorder30 and it has been reported that the loss of TGF-β1 in these mice leads to increased neuronal cell death and microgliosis in the brain47.”

What the sources say

• Supported

  The passage labeled Abstract reports that unmanipulated neonatal Tgfb1−/− mice have increased numbers of apoptotic neurons and widespread microgliosis. The neuronal-degeneration passage also reports significantly more TUNEL-positive cells in knockout brains than in wild-type littermates. No page number is supplied in our copy. (LLM, conf=0.94)

  cites [47] Brionne, T.C., Tesseur, I., Masliah, E. & Wyss-Coray, T. Loss of TGF-beta 1 leads to increased neuronal cell death and microgliosis in mouse brain. Neuron 40, 1133–1145 (2003). source

In the paper

“Our results are consistent with previous studies on the role of microglial in the development and function of the nervous system41,48–50.”

What the sources say

• Supported

  The paper’s conclusion (section “Synaptic Pruning by Microglia Is Necessary for Normal Brain Development”) reports that microglia eliminate synapses during development and that disrupted pruning in mice is associated with immature brain circuitry. It therefore supports the cited background point that microglia have a role in nervous-system development and function. (LLM, conf=0.91)

  cites [41] Paolicelli, R.C. et al. Synaptic pruning by microglia is necessary for normal brain development. Science 333, 1456–1458 (2011). source
• ?Not checked — no full text of the sourcecites [48] Wake, H., Moorhouse, A.J., Jinno, S., Kohsaka, S. & Nabekura, J. Resting microglia directly monitor the functional state of synapses in vivo and determine the fate of ischemic terminals. J. Neurosci. 29, 3974–3980 (2009). source
• Partly supported

  The Results section, “Glia-mediated inflammation contributes to the death of TH+ neurons” (page unavailable), shows that activated microglia can produce factors toxic to neurons. The Discussion, “Nurr1 exerts anti-inflammatory and neuroprotective effects in glia” (page unavailable), likewise links microglial activity to neuronal survival. But the paper does not establish a role for microglia in nervous-system development; its reference to development concerns Nurr1’s role in dopaminergic neurons. (LLM, conf=0.82)

  cites [49] Saijo, K. et al. A Nurr1/CoREST pathway in microglia and astrocytes protects dopaminergic neurons from inflammation-induced death. Cell 137, 47–59 (2009). sourceExperimental — accuracy not yet measured
• Supported

  In “Studying Neuronal-Microglial Interactions during Brain Development” (p. 924), the paper reports that microglia continuously engulf apoptotic neurons during normal brain development. “Microglia in the Zebrafish” also describes their capacity to remove dying neurons and phagocytose bacteria, supporting the broad reference to microglial roles in the developing nervous system. (LLM, conf=0.88)

  cites [50] Peri, F. & Nusslein-Volhard, C. Live imaging of neuronal degradation by microglia reveals a role for v0-ATPase a1 in phagosomal fusion in vivo. Cell 133, 916–927 (2008). source

In the paper

“The blood bran barrier begins to form at E13.5 and may isolate the developing brain from the contribution of fetal liver hematopoiesis40.”

What the sources say

• Supported

  The Figure 1 caption in “The yolk sac hypothesis of microglial origin” states the claim directly: the blood–brain barrier starts to form from E13.5 and may isolate the developing brain from fetal liver hematopoiesis. The discussion in “Evidence for the persistence of the embryonic wave of microglia” also describes restricted access as a possibility, not a certainty. (LLM, conf=0.95)

  cites [40] Ginhoux, F., Lim, S., Hoeffel, G., Low, D. & Huber, T. Origin and differentiation of microglia. Front. Cell. Neurosci. 7, 45 (2013). source

In the paper

“Subsequent reports showed that microglia arise early during development from progenitors in the embryonic yolk sac4,38,39.”

What the sources say

• Supported

  The paper reports that, in mice, the first wave of microglia is specified before the end of embryonic day 8 and that Runx1+ progenitors migrate from the yolk sac into the brain between E8.5 and E9.5 (passages labeled Abstract). This supports the claim of an early embryonic yolk-sac origin. (LLM, conf=0.94)

  cites [4] Ginhoux, F. et al. Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science 330, 841–845 (2010). source
• Partly supported

  The section “Neutral Red Labels Macrophages in Living Embryos” describes young macrophages migrating from the yolk sac, and “A Phenotypic Transition to ‘Early Microglia’” identifies brain and retinal macrophages as early microglia. These findings concern zebrafish; the cited passages do not establish that mouse microglia arise from embryonic yolk-sac progenitors. Page numbers are unavailable in our copy. (LLM, conf=0.84)

  cites [38] Herbomel, P., Thisse, B. & Thisse, C. Zebrafish early macrophages colonize cephalic mesenchyme and developing brain, retina, and epidermis through a M-CSF receptor–dependent invasive process. Dev. Biol. 238, 274–288 (2001). sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [39] Chan, W.Y., Kohsaka, S. & Rezaie, P. The origin and cell lineage of microglia: new concepts. Brain Res. Rev. 53, 344–354 (2007). source

In the paper

“These animals have defects in extracellular glutamate homeostasis and synaptic plasticity33”

What the sources say

• Supported

  The Introduction reports reduced hippocampal glutamate uptake and aberrant synaptic plasticity in CNS-TGFβ1-deficient mice. The Results section also reports impaired synaptic plasticity and identifies a role for TGF-β1 in extracellular glutamate homeostasis. Page numbers are not provided in our copy. (LLM, conf=0.93)

  cites [33] Koeglsperger, T. et al. Impaired glutamate recycling and GluN2B-mediated neuronal calcium overload in mice lacking TGF-beta1 in the CNS. Glia 61, 985–1002 (2013). source

In the paper

“it has been reported that endogenous TGF-β1 is essential for normal murine Langerhans cell development35 and TGF-β1 acts directly on Langerhans cells through an autocrine/paracrine loop36.”

What the sources say

• Partly supported

  Citation 35 attaches to the first clause, not the separate claim about a direct autocrine/paracrine loop. The Abstract concludes that endogenous TGF-β1 is essential for normal murine Langerhans cell development *or epidermal localization*. The Discussion says the nature of that requirement has not been determined, so this source does not establish a developmental requirement specifically. (LLM, conf=0.86)

  cites [35] Borkowski, T.A., Letterio, J.J., Farr, A.G. & Udey, M.C. A role for endogenous transforming growth factor beta 1 in Langerhans cell biology: the skin of transforming growth factor beta 1 null mice is devoid of epidermal Langerhans cells. J. Exp. Med. 184, 2417–2422 (1996). sourceExperimental — accuracy not yet measured
• Supported

  Citation 36 is attached to the claim that TGF-β1 acts directly on Langerhans cells through an autocrine/paracrine loop. The Abstract states this conclusion, and the Results and Discussion section reports that loss of TGF-β1 signaling in Langerhans cells demonstrates a direct effect. (LLM, conf=0.94)

  cites [36] Kaplan, D.H. et al. Autocrine/paracrine TGFbeta1 is required for the development of epidermal Langerhans cells. J. Exp. Med. 204, 2545–2552 (2007). source

In the paper

“We also found that organ-resident macrophages in the gut and liver are “microglia-like” as they express Gas6 and Mertk and these genes may have a role in maintaining tissue homeostasis which is consistent with a recent transcriptional profiling of murine tissue macrophages14.”

What the sources say

• Partly supported

  The Results section on distinct molecular signatures identifies MerTK as an apoptotic-cell phagocytosis receptor expressed across the macrophage populations compared. The Results section on macrophage identification also finds MerTK on liver macrophages. Our copy does not establish a tissue-homeostasis role for MerTK or Gas6, and the Gas6 search found no relevant passage. (LLM, conf=0.81)

  cites [14] Gautier, E.L. et al. Gene-expression profiles and transcriptional regulatory pathways that underlie the identity and diversity of mouse tissue macrophages. Nat. Immunol. 13, 1118–1128 (2012). sourceExperimental — accuracy not yet measured

In the paper

“TGF-β and their receptors are expressed at low levels by both neurons and glial cells42,43, and their expression is upregulated in response to a wide range of neuronal insults and during aging44,45.”

What the sources say

• ?Not checked — no full text of the sourcecites [42] Flanders, K.C. et al. Localization and actions of transforming growth factor-beta s in the embryonic nervous system. Development 113, 183–191 (1991). source
• Partly supported

  The Introduction states that TGF-β1 is upregulated under neuropathological conditions, and the Results show that cultured astrocytes express TGF-β receptors. These passages do not establish low-level expression of TGF-β and its receptors in both neurons and glial cells, or upregulation of both TGF-β and its receptors during aging. (LLM, conf=0.85)

  cites [43] Hamby, M.E., Hewett, J.A. & Hewett, S.J. Smad3-dependent signaling underlies the TGF-beta1–mediated enhancement in astrocytic iNOS expression. Glia 58, 1282–1291 (2010). sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [44] Henrich-Noack, P., Prehn, J.H. & Krieglstein, J. TGF-beta 1 protects hippocampal neurons against degeneration caused by transient global ischemia. Dose-response relationship and potential neuroprotective mechanisms. Stroke 27, 1609–1614, discussion 1615 (1996). source
• ?Not checked — no full text of the sourcecites [45] Wiessner, C. et al. Expression of transforming growth factor-beta 1 and interleukin-1 beta mRNA in rat brain following transient forebrain ischemia. Acta Neuropathol. 86, 439–446 (1993). source

In the paper

“we found increased microglial expression of C1q, Csfr1 and Cd34 genes, which are known to be expressed on tissue resident macrophages14.”

What the sources say

• ?Could not verify

  The Results identify macrophage-associated genes, but the passages retrieved for C1q, Cd34, Csf1r and the printed spelling Csfr1 do not identify the claimed three-gene set. The Methods describe sorting peritoneal macrophages as CD115hi, without linking that marker to the gene named in the claim. Our copy therefore does not establish the attributed expression claim. (LLM, conf=0.72)

  cites [14] Gautier, E.L. et al. Gene-expression profiles and transcriptional regulatory pathways that underlie the identity and diversity of mouse tissue macrophages. Nat. Immunol. 13, 1118–1128 (2012). sourceExperimental — accuracy not yet measured

In the paper

“We have previously reported that these mice have defects related to glutamate recycling and synaptic plasticity33.”

What the sources say

• Supported

  The Discussion reports reduced glutamate recycling in CNS TGF-β1-deficient mice alongside impaired long-term potentiation and facilitated long-term depression. The Results also link decreased glutamate uptake in these mice to altered synaptic plasticity. The supplied passages do not provide page numbers. (LLM, conf=0.94)

  cites [33] Koeglsperger, T. et al. Impaired glutamate recycling and GluN2B-mediated neuronal calcium overload in mice lacking TGF-beta1 in the CNS. Glia 61, 985–1002 (2013). source

In the paper

“Quantitative analysis showed significant reduction of CD39+CD11b+ cells and a reciprocal increase in Ly6C+CD11b+ cells at 20, 90 and 160 days in CNS-TGFβ1−/− mice (Fig. 7h).”

In the paper

“We found 8 microRNAs that were highly expressed in microglia.”

In the paper

“This microglia signature was not observed in microglial lines or in monocytes recruited to the CNS and was also observed in human microglia.”

In the paper

“FACS analysis of human cells showed that anti-P2ry12 stained a subpopulation of CD14+ myeloid cells isolated from fresh post-mortem brain but did not stain peripheral blood CD14+ monocytes (Fig. 4c).”

In the paper

“Apoe was highly expressed in newborn and primary microglia and had minimal expression in adult microglia (Fig. 5c).”

In the paper

“Apoe expression was downregulated in microglia during development and expression of Apoe reciprocally correlated with the induction of microglial genes (Supplementary Fig. 9c).”

In the paper

“We found that IL2TGF-β1-Tg-TGF-β1−/− mice were protected from lethal inflammation, survived into adulthood and were indistinguishable from IL2TGF-β1-Tg-TGF-β1+/− littermates at the age of 4–10 weeks with no signs of inflammation or organ dysfunction.”

In the paper

“We found that there was significant reduction of F4/80+CD11b+ cells as compared to wild type mice and that the level of CD11b expression was reduced in CNS-TGFβ1−/− mice (Fig. 7f).”

In the paper

“FACS analysis shows that anti-FCRLS and anti-P2ry12 antibodies stain adult microglia isolated by conventional methods22 from naïve murine brain but did not stain CD11b-gated myeloid cells from murine spleen, bone marrow and peripheral blood (Fig. 4a).”

What the sources say

• ?Not checked — no full text of the sourcecites [22] Cardona, A.E., Huang, D., Sasse, M.E. & Ransohoff, R.M. Isolation of murine microglial cells for RNA analysis or flow cytometry. Nat. Protoc. 1, 1947–1951 (2006). source

In the paper

“We found that the top microglial functions in naïve adult microglia were related to nervous system development and were not observed in monocytes (Supplementary Fig. 2).”

In the paper

“The P2ry12 signal was not observed on GFAP+ astrocytes or NeuN+ neurons (Fig. 4f) or peripheral tissue resident macrophages in splenic red and white pulp, lung, kidney, liver and skin (Supplementary Fig. 8a).”

In the paper

“we identified genes (e.g., Fcrls, P2ry12, Tmem119, Olfml3, Hexb, Tgfbr1, Gpr34 and Sall1), which appear to be expressed in microglia as we did not detect them in immune cells, Ly6C monocytes or organ specific macrophages.”

In the paper

“Mass spectrometry identified a total of 1991 proteins (Supplementary Fig. 1a,b and Source data Fig. 1).”

In the paper

“recruited GFP+ monocytes do not express P2ry12 and resident GFP− microglia are positive for P2ry12 in both naïve and EAE mice.”

In the paper

“we also found that miR-15b, miR-148a and miR-223 were highly expressed in human monocytes with miR-223 the most selectively expressed miRNA.”

In the paper

“IL2TGF-β1-Tg-TGF-β1+/− mice do not show any clinical or pathological abnormalities and breed well.”

In the paper

“A set of identified 399 microglial genes and 611 monocyte genes (see Fig. 1a and Supplementary Table 1) were imported into MetaCore to build an analysis of functional ontologies including GeneGo process, GeneGo disease process, canonical pathway maps, and networks.”

In the paper

“CNS-TGFβ1−/− mice were normal until they reached 100–120 days of age when they developed motor abnormalities (Supplementary Movie 1) as measured by decreased body weight and reduced rotorod performance (Fig. 7j).”

In the paper

“However, at E14.5 we found a significant reduction in CD39+CD11b+ cells (from 82.7% to 14.7%) associated with reduced surface expression of CD11b.”

In the paper

“In summary, we have identified a TGFβ-1 dependent microglial signature and structures on the surface of microglia that provide the ability to distinguish microglia from infiltrating myeloid cells in the CNS.”

In the paper

“Commonly studied microglia cell lines and primary microglia do not express the adult microglial molecular signature and do not express the signature after culture with TGF-β1.”

In the paper

“In addition, the MG400 chip contains 40 inflammation-related genes which were significantly affected in EAE, APP/PS1 and SOD1 mice (see Supplementary Table 6).”

In the paper

“mouse anti-GFAP (1:300; BD Pharmingen, clone 2E1, validated in ref. 33)”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites ref. 33

In the paper

“The tissue shown in Fig. 4d was stained with a rabbit anti-P2ry12 pAb (1:200, validated in ref. 21).”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites ref. 21

In the paper

“Tgfb1−/− animals die at 3 weeks of age of a severe multifocal autoinflammatory disorder30 and it has been reported that the loss of TGF-β1 in these mice leads to increased neuronal cell death and microgliosis in the brain47.”

What the sources say

• ?Not checked — no full text of the sourcecites [30] Shull, M.M. et al. Targeted disruption of the mouse transforming growth factor-beta 1 gene results in multifocal inflammatory disease. Nature 359, 693–699 (1992). source

In the paper

“Data collection and analysis were performed blind to the conditions of the experiments.”

In the paper

“A MS2 spectra assignment false discovery rate (FDR) of less than 1% was achieved by applying the target-decoy database search strategy60.”

What the sources say

• ?Not checked — no full text of the sourcecites [60] Elias, J.E. & Gygi, S.P. Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry. Nat. Methods 4, 207–214 (2007). source

In the paper

“others have reported that in vitro, TGF-β1 signaling induces a quiescent microglial phenotype25 and promotes resting microglial properties26”

What the sources say

• ?Not checked — no full text of the sourcecites [25] Abutbul, S. et al. TGF-beta signaling through SMAD2/3 induces the quiescent microglial phenotype within the CNS environment. Glia 60, 1160–1171 (2012). source
• ?Not checked — no full text of the sourcecites [26] Schilling, T., Nitsch, R., Heinemann, U., Haas, D. & Eder, C. Astrocyte-released cytokines induce ramification and outward K+ channel expression in microglia via distinct signaling pathways. Eur. J. Neurosci. 14, 463–473 (2001). source

In the paper

“Thus, primitive macrophages exit the yolk sac blood islands at the onset of circulation and colonize the neuroepithelium from E9.5 to give rise to microglia.”

In the paper

“All qRT-PCRs were performed in duplicate or triplicate, and the data are presented as mean ± standard error of mean (s.e.m).”

In the paper

“Numerous studies have demonstrated that TGF-β1 exerts a protective effect against various neuronal insults46.”

What the sources say

• ?Not checked — no full text of the sourcecites [46] Flanders, K.C., Ren, R.F. & Lippa, C.F. Transforming growth factor-betas in neurodegenerative disease. Prog. Neurobiol. 54, 71–85 (1998). source

In the paper

“No statistical methods were used to predetermine sample sizes, but our sample sizes are similar to those reported in previous publication12.”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites previous publication12

In the paper

“RNA quality and integrity were assessed using Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA) with all samples having high-quality RNA [RNA integrity number (RIN) = 8.7–10].”

In the paper

“It is well known that TAM family receptors/ligands are expressed in macrophages15.”

What the sources say

• ?Not checked — no full text of the sourcecites [15] Rothlin, C.V., Ghosh, S., Zuniga, E.I., Oldstone, M.B. & Lemke, G. TAM receptors are pleiotropic inhibitors of the innate immune response. Cell 131, 1124–1136 (2007). source

In the paper

“which confirms previous observations that PU.1 deficient mice do not develop myeloid cells19,20.”

What the sources say

• ?Not checked — no full text of the sourcecites [19] Scott, E.W. et al. PU.1 functions in a cell-autonomous manner to control the differentiation of multipotential lymphoid-myeloid progenitors. Immunity 6, 437–447 (1997). source
• ?Not checked — no full text of the sourcecites [20] Beers, D.R. et al. Wild-type microglia extend survival in PU.1 knockout mice with familial amyotrophic lateral sclerosis. Proc. Natl. Acad. Sci. USA 103, 16021–16026 (2006). source

In the paper

“Also, data for each experiment were collected and processed randomly and animals were assigned to various experimental groups randomly as well.”

In the paper

“Thus, embryonic and early post-natal microglia acquire the features of adult microglia after post-natal day four, possibly as newly-proliferated microglial cells begin to express differentiated functions.”

In the paper

“No statistical methods were used to predetermine sample sizes, but our sample sizes are similar to those reported in previous publication12.”

In the paper

“rabbit anti-Iba1 (1:500; Wako, cat. no. 019-19741, validated in ref. 12)”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites ref. 12

In the paper

“mouse anti-NeuN (1:500; Millipore, validated in ref. 12)”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites ref. 12

In the paper

“The quality of cDNA and fragmented cDNA was assessed in the Agilent bioanalyzer.”

In the paper

“In summary, lack of TGF-β1 affected microglial development beginning at E14.5 but did not affect microglial progenitors at the E10.5 stage.”

In the paper

“Because we identified genes related to neuronal function both in cultured as well as ex vivo adult microglia, the presence of these genes does not simply represent extracellular absorption as the microglial genes we identified were upregulated by exogenous TGF-β1 in vitro.”

In the paper

“To minimize technical (nonbiological) variability among arrays, densitometry values between arrays were normalized using the Robust Multichip Average function and further transformed to the logarithmic scale (log2).”

In the paper

“On the C57BL6/Sv129 background, 60–80% of (statistically expected) homozygous TGF-β1−/− conceptuses reached parturition as described29.”

What the sources say

• ?Not checked — no full text of the sourcecites [29] Kulkarni, A.B. et al. Transforming growth factor beta 1 null mutation in mice causes excessive inflammatory response and early death. Proc. Natl. Acad. Sci. USA 90, 770–774 (1993). source

In the paper

“Also, data for each experiment were collected and processed randomly and animals were assigned to various experimental groups randomly as well.”

In the paper

“To validate these findings, we performed qPCR analysis of microglial miRNAs (miR-125b-5p, miR-99a, miR-342-3p) and inflammatory Ly6C+ miRNAs (miR-15b, miR-148a, miR-223) and found a similar expression pattern that we observed by Nanostring (Fig. 3c,d).”

In the paper

“Control sections (not treated with primary antibody) were used to distinguish specific from nonspecific staining or autofluorescent components.”