Inhibition of Overactive TGF-β-Induced Fibrotic Scar Formation Repairs Mouse Spinal Cord Injury
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Spinal cord injury (SCI) often causes disability in humans and other mammals. Here we report that fibrotic scar formation at injury sites prevents recovery after SCI and that the inhibition of fibrotic scar formation significantly improved recovery in adult mice. Neonatal mice were able to fully recover from SCI because they do not experience fibrotic scar formation. Active transforming growth factor-β 1 (TGF-β1) was significantly elevated at SCI sites to recruit mesenchymal stromal/stem cells (MSCs) and induce fibroblast differentiation. Eliminating macrophage lineage cells in LysM-cre::iDTRflox/flox mice significantly decreased TGF-β activity, suggesting macrophages as primary source of TGF-β1. Moreover, TGF-β activity was significantly decreased after selective deletion of TGF-β1 in macrophages in LysM-cre::Tgfb1flox/flox mice. Knocking out of TGF-β type 2 receptor in pericytes of Glast-creERT2::Tgfbr2flox/flox mice also decreased fibrotic scar formation. Interestingly, TGF-β–neutralizing antibody 1D11 treatment induced a greater inhibition of scar formation than any of the knockout mice. Single-cell RNA sequencing revealed that TGF-β downstream signaling was significantly enriched in MSCs, fibroblasts, pericytes, and endothelial cells after SCI, and the lineage trajectory indicates that pericytes differentiate to fibroblasts via MSCs. Most importantly, neonatal mice did not have active TGF-β at the injury site and they recovered completely after SCI. Thus, fibrotic scar may be the primary obstacle that prevents recovery after SCI.
脊髓损伤（Spinal cord injury, SCI）常导致人类及其他哺乳动物出现残疾。本研究证实，损伤部位的纤维化瘢痕形成会阻碍脊髓损伤后的修复，而抑制纤维化瘢痕形成可显著改善成年小鼠的运动功能恢复。新生小鼠能够完全从脊髓损伤中康复，因其不会形成纤维化瘢痕。脊髓损伤部位的活性转化生长因子-β1（transforming growth factor-β1, TGF-β1）水平显著升高，可募集间充质基质/干细胞（mesenchymal stromal/stem cells, MSCs）并诱导成纤维细胞分化。在LysM-cre::iDTRflox/flox小鼠中清除巨噬细胞谱系细胞后，TGF-β活性显著降低，提示巨噬细胞是TGF-β1的主要来源。此外，在LysM-cre::Tgfb1flox/flox小鼠中选择性敲除巨噬细胞内的TGF-β1后，TGF-β活性也显著下降。在Glast-creERT2::Tgfbr2flox/flox小鼠的周细胞中敲除转化生长因子βⅡ型受体（TGF-β type 2 receptor），同样可减少纤维化瘢痕的形成。值得注意的是，使用TGF-β中和抗体1D11进行干预，其对瘢痕形成的抑制效果优于所有基因敲除小鼠模型。单细胞RNA测序结果显示，脊髓损伤后，TGF-β下游信号通路在MSCs、成纤维细胞、周细胞及内皮细胞中显著富集；谱系轨迹分析表明，周细胞可通过MSCs向成纤维细胞分化。最为关键的是，新生小鼠损伤部位不存在活性TGF-β，因此可完全恢复脊髓损伤后的功能。综上，纤维化瘢痕可能是阻碍脊髓损伤后修复的主要障碍。
