Single-cell RNA sequencing of equine mesenchymal stromal cells from primary donor-matched tissue sources reveals functional heterogeneity in immune modulation and cell motility.

BACKGROUND: The efficacy of mesenchymal stromal cell (MSC) therapy is thought to depend on the intrinsic heterogeneity of MSC cultures isolated from different tissue sources as well as individual MSCs isolated from the same tissue source, neither of which is well understood. To study this, we used MSC cultures isolated from horses. The horse is recognized as a physiologically relevant large animal model appropriate for translational MSC studies. Moreover, due to its large size the horse allows for the simultaneous collection of adequate samples from multiple tissues of the same animal, and thus, for the unique collection of donor matched MSC cultures from different sources. The latter is much more challenging in mice and humans due to body size and ethical constraints, respectively. METHODS: In the present study, we performed single-cell RNA sequencing (scRNA-seq) on primary equine MSCs that were collected from three donor-matched tissue sources; adipose tissue (AT), bone marrow (BM), and peripheral blood (PB). Based on transcriptional differences detected with scRNA-seq, we performed functional experiments to examine motility and immune regulatory function in distinct MSC populations. RESULTS: We observed both inter- and intra-source heterogeneity across the three sources of equine MSCs. Functional experiments demonstrated that transcriptional differences correspond with phenotypic variance in cellular motility and immune regulatory function. Specifically, we found that (i) differential expression of junctional adhesion molecule 2 (JAM2) between MSC cultures from the three donor-matched tissue sources translated into altered cell motility of BM-derived MSCs when RNA interference was used to knock down this gene, and (ii) differences in C-X-C motif chemokine ligand 6 (CXCL6) expression in clonal MSC lines derived from the same tissue source correlated with the chemoattractive capacity of PB-derived MSCs. CONCLUSIONS: Ultimately, these findings will enhance our understanding of MSC heterogeneity and will lead to improvements in the therapeutic potential of MSCs, accelerating the transition from bench to bedside.

Caffeine consumption disrupts hippocampal long-term potentiation in freely behaving rats.

Caffeine, one of the most commonly consumed psychoactive substances in the world, has long been known to alter neurological functions, such as alertness, attention, and memory. Despite caffeine's popularity, systematic investigations of its effects on synaptic plasticity in the brain are still lacking. Here we used a freely behaving rodent model of long-term potentiation (LTP), a frequently studied form of synaptic plasticity, to assess the effects of caffeine consumption on hippocampal plasticity. LTP, which is a persistent increase in the strength of synaptic connections between neurons, is a cellular mechanism widely considered to underlie the processes of learning and memory. A group of 10-week-old Sprague-Dawley rats were administered caffeine (1 g/L) in their drinking water 3 weeks prior to collection of electrophysiological data. Another group of age-matched animals received tap water and served as controls. Stimulating and recording electrodes were chronically implanted in the perforant pathway (PP) and dentate gyrus (DG) region of the hippocampus, respectively, to permit stable electrophysiological recordings of synaptic transmission at this synapse. Population spike amplitude (PSA) measures of LTP induction and duration were acquired in vivo while animals were freely behaving using a well-established electrophysiological recording protocol. Results indicate caffeine-treated rats (n = 9) had a significantly (P < 0.05) reduced level of LTP induction compared with controls (n = 10). More studies are needed to identify the exact mechanism through which caffeine alters LTP induction in this freely behaving model of synaptic plasticity.