RNAseq from normal human epidermal keratinocytes treated with C. acnes 6919 conditioned media and propionic acid

Lipid synthesis is necessary for formation of epithelial barriers and homeostasis with external microbes. An analysis of the response of human keratinocytes to several different commensal bacteria on the skin revealed that Cutibacterium acnes induced a large increase in essential lipids including triglycerides, ceramides, cholesterol, and free fatty acids. A similar response occurred in mouse epidermis and in human skin affected with acne. Further analysis showed that this increase in lipids was mediated by short-chain fatty acids produced by Cutibacterium acnes and was dependent on increased expression of several lipid synthesis genes including glycerol3-phosphate-acyltransferase-3. Inhibition or RNA silencing of peroxisome proliferator–activated receptor–α (PPARα), but not PPARβ and PPARγ, blocked this response. The increase in keratinocyte lipid content improved innate barrier functions including antimicrobial activity, paracellular diffusion, and transepidermal water loss. These results reveal that metabolites from a common commensal bacterium have a previously unappreciated influence on the composition of epidermal lipids.

Methods
Normal neonatal human epidermal keratinocytes (NHEKs) were treated with 8mM of propionic acid or with 15% of C. acnes conditioned media or RCM media (control) for 4 days in triplicate. Then, RNA was extracted using the PureLink RNA mini kit. Isolated RNA was submitted to the UCSD IGM Genomics Center for RNA-sequencing performed on a high-output run V4 platform (Illumina, USA) with a single read 100 cycle runs.

脂质合成对于上皮屏障的构建以及维持机体与外界微生物间的稳态平衡不可或缺。针对人类角质形成细胞（keratinocytes）与皮肤内数种不同共生菌的应答反应开展分析后发现，痤疮丙酸杆菌（Cutibacterium acnes）可诱导甘油三酯、神经酰胺、胆固醇及游离脂肪酸等必需脂质的大量合成。该应答效应在小鼠表皮以及痤疮患者的皮肤组织中同样得以重现。进一步研究表明，脂质合成的增加由痤疮丙酸杆菌产生的短链脂肪酸所介导，且依赖于包括甘油3-磷酸酰基转移酶3（glycerol3-phosphate-acyltransferase-3）在内的多种脂质合成基因的表达上调。抑制过氧化物酶体增殖物激活受体α（PPARα）或对其进行RNA沉默，可阻断该应答效应，但对PPARβ与PPARγ无此调控作用。角质形成细胞内脂质含量的提升可增强天然屏障功能，涵盖抗菌活性、细胞旁扩散能力及经皮水分丢失（transepidermal water loss）调控等多个维度。上述研究结果证实，常见共生菌产生的代谢物可对表皮脂质组成产生此前未被认知的调控作用。

方法
将正常新生儿表皮角质形成细胞（NHEKs）分别以8mM丙酸、15%痤疮丙酸杆菌条件培养基或RCM培养基（对照组）进行处理，每组设置3个生物学复孔，培养4天。随后采用PureLink RNA迷你试剂盒提取总RNA。提取获得的RNA被送至加州大学圣地亚哥分校IGM基因组中心，依托美国因美纳（Illumina）公司的高通量测序V4平台（High-Output Run V4）开展单端100循环的RNA测序。