Nöske Katharina, Stark Hans-Jürgen, Nevaril Leonard, Berning Manuel, Langbein Lutz, Goyal Ashish, Diederichs Sven, Boukamp Petra
Department of Genetics of Skin Carcinogenesis, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany.
Department of RNA Biology and Cancer, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany.
Int J Mol Sci. 2016 Jan 28;17(2):167. doi: 10.3390/ijms17020167.
Despite decades of skin research, regulation of proliferation and homeostasis in human epidermis is still insufficiently understood. To address the role of mitoses in tissue regulation, we utilized human long-term skin equivalents and systematically assessed mitoses during early epidermal development and long-term epidermal regeneration. We now demonstrate four different orientations: (1) horizontal, i.e., parallel to the basement membrane (BM) and suggestive of symmetric divisions; (2) oblique with an angle of 45°-70°; or (3) perpendicular, suggestive of asymmetric division. In addition, we demonstrate a fourth substantial fraction of suprabasal mitoses, many of which are committed to differentiation (Keratin K10-positive). As verified also for normal human skin, this spatial mitotic organization is part of the regulatory program of human epidermal tissue homeostasis. As a potential marker for asymmetric division, we investigated for Numb and found that it was evenly spread in almost all undifferentiated keratinocytes, but indeed asymmetrically distributed in some mitoses and particularly frequent under differentiation-repressing low-calcium conditions. Numb deletion (stable knockdown by CRISPR/Cas9), however, did not affect proliferation, neither in a three-day follow up study by life cell imaging nor during a 14-day culture period, suggesting that Numb is not essential for the general control of keratinocyte division.
尽管对皮肤进行了数十年的研究，但对人类表皮中增殖和内稳态的调节仍了解不足。为了探讨有丝分裂在组织调节中的作用，我们利用人类长期皮肤等效物，并在早期表皮发育和长期表皮再生过程中系统地评估有丝分裂。我们现在展示了四种不同的方向：（1）水平方向，即平行于基底膜（BM），提示对称分裂；（2）倾斜角度为45°-70°；或（3）垂直方向，提示不对称分裂。此外，我们还展示了基底上层有丝分裂的第四个重要部分，其中许多细胞致力于分化（角蛋白K10阳性）。正如对正常人类皮肤所验证的那样，这种空间有丝分裂组织是人类表皮组织内稳态调节程序的一部分。作为不对称分裂的潜在标志物，我们研究了Numb，发现它在几乎所有未分化的角质形成细胞中均匀分布，但在某些有丝分裂中确实不对称分布，并且在抑制分化的低钙条件下尤为常见。然而，Numb缺失（通过CRISPR/Cas9稳定敲低）在通过活细胞成像进行的为期三天的随访研究中以及在为期14天的培养期内均未影响增殖，这表明Numb对于角质形成细胞分裂的总体控制并非必不可少。
