Peptidoglycan-Mediated Bone Marrow Autonomic Neuropathy Impairs Hematopoietic Stem/Progenitor Cells via a NOD1-Dependent Pathway in db/db Mice

Figure 6

Chemical sympathectomy blocked the protective effects of ML130 on HSPC populations. (a) Representative IHC images of bone marrow sections stained with tyrosine hydroxylase (left panel) and NF200 (right panel). (b, c) db/db and db/m mice were treated with PGN, PGN+ML130, or vehicle after the chemical sympathectomy induced by 6-OHDA. 6-OHDA caused severe bone marrow denervation, as determined by the reduced numbers of (b) try-OH⁺ and (c) NF200⁺ neurons in the mouse femur ( per group). (d) Representative plots of flow cytometry analysis showing HSPC populations (LSK, LT-HSC, and ST-HSC). (e–g) The (f) LT-HSC percentage out of total live cells was dramatically reduced in sympathectomized (6-OHDA) diabetic animals regardless of ML130 treatment determined by multicolor flow cytometry studies. The (e) total LSK and (g) ST-HSC populations were not affected by 6-OHDA ( per group). (h) The CFU assay showed increased CFU-G/M/GM of bone marrow cells cotreated with 6-OHDA and ML130 compared to those treated with ML130 alone ( per group). (i) LSK cells from each group were cultured and tested for colonies in the LTC-IC assay. 6-OHDA blocked the beneficial effect of ML130 on PGN-induced reduction in the total number of colonies (per group). (j) For the in vivo BrdU incorporation study, the bar graph in the (j) shows the percentages of BrdU⁺ CD34^-LSK cells were decreased after he chemical sympathectomy induced by 6-OHDA irrespective of diabetic state by FACS analysis ( per group). Data represent . PGN: peptidoglycan; V: vehicle; LSK: Lin^-/CD127^-/Sca-1⁺c-Kit⁺; LT-HSC: long-term repopulating hematopoietic stem cell; ST-HSC: short-term repopulating hematopoietic stem cell. , ; ^&compared to db/db+PGN; ^§compared to db/db+V; ^#compared to db/db+PGN+ML130; ^※compared to db/m+PGN+ML130; ^¶compared to db/db+PGN+ML130.

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