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Stromal cell-derived NGF controls sympathetic innervation in subcutaneous fat

  • Xia Meng
    Affiliations
    Institute for Immunology and Department of Basic Medical Sciences, School of Medicine, Tsinghua University, and Tsinghua-Peking Center for Life Sciences, Beijing, 100084, China
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  • Jian Chen
    Correspondence
    Correspondence:
    Affiliations
    Chinese Institute for Brain Research, Beijing, 102206, China
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  • Wenwen Zeng
    Correspondence
    Correspondence:
    Affiliations
    Institute for Immunology and Department of Basic Medical Sciences, School of Medicine, Tsinghua University, and Tsinghua-Peking Center for Life Sciences, Beijing, 100084, China

    Beijing Key Laboratory for Immunological Research on Chronic Diseases, Beijing, 100084, China
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Open AccessPublished:August 10, 2022DOI:https://doi.org/10.1016/j.jlr.2022.100264

      Graphical abstract

      The densely distributed sympathetic nerves in the mouse subcutaneous inguinal white adipose tissues (iWAT) promote the thermogenic capacity to counteract the cold environment. The sympathetic innervation within iWAT is established during early development and persists throughout adulthood (
      • Chi J.
      • Lin Z.
      • Barr W.
      • Crane A.
      • Zhu X.G.
      • Cohen P.
      Early postnatal interactions between beige adipocytes and sympathetic neurites regulate innervation of subcutaneous fat.
      ). Previous studies have indicated that the sympathetic nerve outgrowth could be modulated by neurotrophic factors, resulting in altered parenchymal sympathetic density at the adult stage (
      • Meng X.
      • Qian X.
      • Ding X.
      • Wang W.
      • Yin X.
      • Zhuang G.
      • Zeng W.
      Eosinophils regulate intra-adipose axonal plasticity.
      ,
      • Xie H.
      • Heier C.
      • Meng X.
      • Bakiri L.
      • Pototschnig I.
      • Tang Z.
      • Schauer S.
      • Baumgartner V.J.
      • Grabner G.F.
      • Schabbauer G.
      • Wolinski H.
      • Robertson G.R.
      • Hoefler G.
      • Zeng W.
      • Wagner E.F.
      • Schweiger M.
      • Zechner R.
      An immune-sympathetic neuron communication axis guides adipose tissue browning in cancer-associated cachexia.
      ). Intriguingly, expression profiles examining the iWAT-resident cells reveal abundant expression of nerve growth factor (Ngf) in the stromal cells, in contrast to other cell types including adipocytes (
      • Meng X.
      • Qian X.
      • Ding X.
      • Wang W.
      • Yin X.
      • Zhuang G.
      • Zeng W.
      Eosinophils regulate intra-adipose axonal plasticity.
      ). To determine whether stromal cell-derived NGF may play a significant role in sympathetic innervation, we bred Ngffl/fl mice generated in the previous study (
      • Meng X.
      • Qian X.
      • Ding X.
      • Wang W.
      • Yin X.
      • Zhuang G.
      • Zeng W.
      Eosinophils regulate intra-adipose axonal plasticity.
      ) with Prrx1Cre mice to achieve Ngf deletion in stromal cells. Transcript analysis verified that Ngf expression reduced drastically in iWAT, in agreement with a predominant contribution of NGF from stromal cells. Next, we performed whole-mount immunostaining and volume-fluorescence imaging using antibodies against tyrosine hydroxylase (TH) and protein gene product 9.5 (PGP9.5) to visualize sympathetic nerves and total peripheral nerves, respectively. Remarkably, at the whole tissue level, sympathetic nerves immunolabeled by TH were largely abrogated in the iWAT parenchyma of Prrx1Cre; Ngffl/fl mice compared to the control Ngffl/fl mice (Movie S1 for Ngffl/fl, and S2 for Prrx1Cre; Ngffl/fl). Consistent with the predominant distribution of sympathetic nerve type in the iWAT, total nerves immunolabeled by PGP9.5 also showed defective branching in Prrx1Cre; Ngffl/fl mice, with only a few by-passing nerve bundles visible (Movie S5 for Ngffl/fl, and S6 for Prrx1Cre; Ngffl/fl). Moreover, the fine nerve fibers imaged at high magnification within inguinal regions proximal to the lymph node presented a similar result (Movie S3 and S4 for TH, Movie S7 and S8 for PGP9.5). Taken together, these data suggest that stromal cell-derived NGF plays a dominant role in the establishment of intra-adipose sympathetic innervations.
      EQUIPMENT: Ultramicroscope II with a 1.1x 0.1 NA objective lens (Miltenyi Biotec, 1x zoom) or 4x 0.35 NA objective lens (Miltenyi Biotec, 2x zoom).
      REAGENTS AND ANIMALS: The whole-mount immunostaining and volume-fluorescence imaging were carried out as previously described (
      • Meng X.
      • Qian X.
      • Ding X.
      • Wang W.
      • Yin X.
      • Zhuang G.
      • Zeng W.
      Eosinophils regulate intra-adipose axonal plasticity.
      ) with the following antibodies: rabbit anti-TH (Millipore catalog no. AB152, RRID: AB_390204), rabbit anti-PGP9.5 (Proteintech catalog no. 14730–1-AP, RRID: AB_2210497), and Alexa Fluor 647-goat anti-rabbit (Thermo Fisher Scientific). Prrx1Cre mice were obtained from Jackson Laboratory (JAX 005584, RRID: IMSR_JAX:005584).
      SOFTWARE: Imaris (Oxford Instruments).

      Supplementary data

      References

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        Early postnatal interactions between beige adipocytes and sympathetic neurites regulate innervation of subcutaneous fat.
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        Eosinophils regulate intra-adipose axonal plasticity.
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        An immune-sympathetic neuron communication axis guides adipose tissue browning in cancer-associated cachexia.
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