Pre-proenkephalin 1 (Penk1) is a pro-neuropeptide that belongs to the typical opioid peptide’s family, having analgesic properties. We previously found Penk1 to be the most downregulated gene in a whole gene profiling analysis performed in osteoblasts subjected to microgravity as a model of mechanical unloading. In this work, Penk1 downregulation was confirmed in the bones of two in vivo models of mechanical unloading: tail-suspended and botulinum toxin A (botox)-injected mice. Consistently, in the sera from healthy volunteers subjected to bed rest, we observed an inverse correlation between PENK1 and bed rest duration. These results prompted us to investigate a role for this factor in bone. Penk1 was highly expressed in mouse bone, but its global deletion failed to impact bone metabolism in vivo. Indeed, Penk1 knock out (Penk1^−/−) mice did not show an overt bone phenotype compared to the WT littermates. Conversely, in vitro Penk1 gene expression progressively increased during osteoblast differentiation and its transient silencing in mature osteoblasts by siRNAs upregulated the transcription of the Sost1 gene encoding sclerostin, and decreased Wnt3a and Col1a1 mRNAs, suggesting an altered osteoblast activity due to an impairment of the Wnt pathway. In line with this, osteoblasts treated with the Penk1 encoded peptide, Met-enkephalin, showed an increase of Osx and Col1a1 mRNAs and enhanced nodule mineralization. Interestingly, primary osteoblasts isolated from Penk1^−/− mice showed lower metabolic activity, ALP activity, and nodule mineralization, as well as a lower number of CFU-F compared to osteoblasts isolated from WT mice, suggesting that, unlike the transient inhibition, the chronic Penk1 deletion affects both osteoblast differentiation and activity. Taken together, these results highlight a role for Penk1 in the regulation of the response of the bone to mechanical unloading, potentially acting on osteoblast differentiation and activity in a cell-autonomous manner.

前脑啡肽原 1 (Penk1) 是一种前神经肽，属于典型的阿片肽家族，具有镇痛特性。我们之前发现，在以微重力作为机械卸载模型的成骨细胞中进行的全基因谱分析中， Penk1是下调幅度最大的基因。在这项工作中，Penk1下调在两种体内机械卸载模型的骨骼中得到证实：悬尾小鼠和注射肉毒杆菌毒素 A (botox) 的小鼠。一致地，在卧床休息的健康志愿者的血清中，我们观察到 PENK1 与卧床时间之间呈负相关。这些结果促使我们研究该因子在骨骼中的作用。Penk1在小鼠骨骼中高表达，但其整体缺失未能影响体内骨代谢。事实上，与 WT 同窝小鼠相比， Penk1敲除 ( Penk1 ^−/− ) 小鼠并未表现出明显的骨表型。相反，体外Penk1基因表达在成骨细胞分化过程中逐渐增加，并且siRNA在成熟成骨细胞中的短暂沉默上调了编码硬化素的Sost1基因的转录，并减少了Wnt3a和Col1a1 mRNA，这表明由于Wnt受损而导致成骨细胞活性改变途径.与此一致的是，用Penk1编码肽 Met-脑啡肽处理的成骨细胞显示Osx和Col1a1 mRNA增加，结节矿化增强。有趣的是，与从 WT 小鼠中分离的成骨细胞相比，从Penk1 ^−/−小鼠中分离的原代成骨细胞表现出较低的代谢活性、ALP 活性和结节矿化，以及较低的 CFU-F 数量，这表明与瞬时抑制不同，慢性Penk1缺失会影响成骨细胞的分化和活性。总而言之，这些结果强调了 Penk1 在调节骨骼对机械卸载的反应中的作用，可能以细胞自主的方式作用于成骨细胞的分化和活性。