Comparative transcriptomics reveal highly conserved regional programs between porcine and human colonic enteric nervous system

The enteric nerve system (ENS) coordinates diverse gastrointestinal functions. The porcine gastrointestinal tract is increasingly regarded as a useful translational model. However, porcine ENS molecular profiling and its similarity to that of human are lacking. Bulk RNA sequencing (RNA-seq) coupled with laser-capture microdissection and single-cell RNA-seq on myenteric ganglia (MG) addressed that discrepant transcriptional programs explained the functional characteristics between inner submucosal and/or MG in porcine proximal or/and distal colon (p-pC, p-dC). Comparative bulk transcriptomics of MG in corresponding colonic regions of porcine and human revealed highly conserved programs existing in p-pC and p-dC, which interpreted >90% of their transcriptomic responses to vagal nerve stimulation (VNS), suggesting that p-pC and p-dC could serve as predictors in translational studies. At bulk and single-cell resolution, we highlighted specific conserved programs, executing inflammatory modulation in porcine with VNS. This study offers rich foundations to understand human colonic functions and neuromodulation using porcine model. Overall design: The colonic specimens with full thickness from three castrated male and three intact female naïve Yucatan minipigs and three castrated male Yucatan minipigs undergoing vagal nerve stimulation (VNS) were removed from the proximal colon (p-pC) and distal colon (p-dC). The full thickness of colonic specimens were dissected from healthy margin of the post-operative ascending and descending colon (h-aC, h-dC, 4 of each) from 12 patients (4 males and 8 females) with colonic adenocarcinoma. A range of 25-40 ganglia/subject were harvested from inner submucosal ganglia (ISG) and myenteric ganglia (MG) respectively of p-pC and p-dC and from MG of h-aC and h-dC using LMD-6000 Laser Micro-dissection System. Total 46 samples were sequenced on an Illumina HiSeq 3000 sequencer. In addition, the muscularis externa containing myenteric ganglia from each p-pC and p-dC of 4 naïve porcine (2 of each sex) were processed for cell suspension preparation, which were loaded on the 10x Genomics Chromium platform and sequenced on an Illumina NextSeq 500 Sequencer.