Deep Proteome Profiling Reveals Signatures of Age and Sex Differences in Paw Skin and Sciatic Nerve of Naïve Mice

The age and sex of studied animals profoundly impact experimental outcomes in animal-based preclinical biomedical research. However, most preclinical studies in mice use a wide-spanning age range from 4 to 14 weeks and do not assess study parameters in male and female mice in parallel. This raises concerns regarding reproducibility and neglects potentially relevant age and sex differences. Furthermore, the molecular setup of tissues in dependence of age and sex is unknown in naïve mice. Here, we employed an optimized quantitative proteomics workflow in order to deeply profile mouse paw skin and sciatic nerve (SCN) – two tissues, which are crucially implicated in nociception and pain as well as diverse diseases induced by inflammation, trauma, and demyelination. Remarkably, we uncovered significant differences when comparing (i) male and female mice, and, in parallel, (ii) adolescent mice (4 weeks) with adult mice (14 weeks). Age was identified as a major discriminator of analyzed samples irrespective of tissue type. Moreover, our analysis enabled us to decipher protein subsets and networks that exhibit differential abundance in dependence on the age and/or sex of mice. Notably, among these were proteins and signaling pathways with known relevance for (patho)physiology, such as homeostasis and epidermal signaling in skin and, in SCN, multiple myelin proteins and regulators of neuronal development. In addition, extensive comparisons with available databases revealed that we quantified approx. 50% of gene products that were implicated in distinct skin diseases and pain, many of which exhibited significant abundance changes in dependence on age and/or sex. Taken together, our study emphasizes the need for accurate age matching and uncovers hitherto unknown sex and age differences at the level of proteins and protein networks. Overall, we provide a unique systems biology proteome resource, which facilitates mechanistic insights into somatosensory and skin biology in dependence on age and sex - a prerequisite for successful preclinical studies in mouse disease models.