Traumatic brain injury (TBI) is a major risk factor for late-onset neurodegenerative disorders, including Alzheimer’s disease (AD). We investigated whether mild TBI (mTBI) at the presymptomatic stage differentially affects the gut microbiota–endocannabinoidome (eCBome) axis. Caco-2 cells were treated as follows: (i) untreated (CTRL), (ii) IL-1β (10 ng/ml for 24 h), (iii) 2-AG 10 μM (24 h), (iv) 2-AG 10 μM + IL-1β (30 min 2-AG IL-1β 24 h). We characterized proteome-wide quantitative changes in this cell model under different treatment conditions using the LFQ proteomics approach. Our findings identify the gut microbiota–endocannabinoidome axis as a key mediator of long-term outcomes after brain trauma.