The Saliva and Muscle Study (SaMu): Rationale and Protocol for Associations between Salivary Microbiome and Accelerated Muscle Ageing.

Background: The gut microbiome is recognized as a pivotal factor in the pathophysiology of sarcopenia-a condition marked by the accelerated loss of muscle strength, mass and function with ageing. Despite this well-known gut-muscle axis, the potential links between other microbial ecosystems and sarcopenia remain largely unexplored. The oral microbiome has been linked to various age-related health conditions such as rheumatoid arthritis and colorectal cancer. However, its potential association with sarcopenia is unknown. The Saliva and Muscle (SaMu) study seeks to address this knowledge gap.

Methods: The SaMu study comprises three sequential phases. In phase 1, a cross-sectional analysis will be conducted on a cohort of 200 individuals aged 70 years or older to examine the relationship between salivary microbiome and sarcopenia status. Participants will be recruited in the three main places of living: general community, assisted living facilities and nursing homes. The salivary microbiome composition will be evaluated utilizing shotgun metagenomics sequencing, while sarcopenia status will be determined through muscle mass (determined by whole-body bioelectrical impedance analysis and calf circumference), muscle strength (grip strength and the 5-times-sit-to-stand test) and physical performance (usual walking speed). In addition to investigating the microbiome composition, the study aims to elucidate microbiome functions by exploring potential omic associations with sarcopenia. To achieve this, salivary proteomics, metabolomics and quorum sensing peptidomics will be performed. Covariates that will be measured include clinical variables (sociodemographic factors, health status, health-related behaviours, oral health and quality of life) as well as blood variables (immune profiling, hormones, kidney and liver function, electrolytes and haematocrit). In phase 2, an in-depth mechanistic analysis will be performed on an envisaged subcohort of 50 participants. This analysis will explore pathways in muscle tissue using histology, genomics and transcriptomics, focusing on (maximal) 25 healthy older adults and (maximal) 25 with severe sarcopenia. Phase 3 involves a two-year clinical follow-up of the initial participants from the cross-sectional analysis, along with a resampling of blood and saliva. Additionally, secondary outcomes like falls, hospitalization and mortality will be examined.

Discussion: Using a salivary multi-omics approach, SaMu primarily aims to clarify the associations between the oral microbiome and sarcopenia. SaMu is expected to contribute to the discovery of predictive biomarkers of sarcopenia as well as to the identification of potential novel targets to prevent/tackle sarcopenia. This study-protocol is submitted for registration at the ISRCTN registry.