Delivery of enteric neural progenitors with 5-HT4 agonist-loaded nanoparticles and thermosensitive hydrogel enhances cell proliferation and differentiation following transplantation in vivo

Abstract

Cell therapy offers an innovative approach for treating enteric neuropathies. Postnatal gut-derived enteric neural stem/progenitor cells (ENSCs) represent a potential autologous source, but have a limited capacity for proliferation and neuronal differentiation. Since serotonin (5-HT) promotes enteric neuronal growth during embryonic development, we hypothesized that serotonin receptor agonism would augment growth of neurons from transplanted ENSCs. Postnatal ENSCs were isolated from 2-4 week-old mouse colon and cultured with 5-HT4 receptor agonist (RS67506)-loaded liposomal nanoparticles. ENSCs were co-cultured with mouse colon explants in the presence of RS67506-loaded (n=3) or empty nanoparticles (n=3). ENSCs were also transplanted into mouse rectum in vivo with RS67506-loaded (n=8) or blank nanoparticles (n=4) confined in a thermosensitive hydrogel, Pluronic F-127. Neuronal density and proliferation were analyzed immunohistochemically. Cultured ENSCs gave rise to significantly more neurons in the presence of RS67506-loaded nanoparticles. Similarly, colon explants had significantly increased neuronal density when RS67506-loaded nanoparticles were present. Finally, following in vivo cell delivery, co-transplantation of ENSCs with 5-HT4 receptor agonist-loaded nanoparticles led to significantly increased neuronal density and proliferation. We conclude that optimization of postnatal ENSCs can support their use in cell-based therapies for neurointestinal diseases.

Optimizing neurogenic potential of enteric neurospheres for treatment of neurointestinal diseases

“…With approval from the Institutional Animal Care and Use Committee, neurospheres were generated from male and female 3-week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) according to previously published protocols (7, 12). The muscularis propria, which contains the myenteric plexus, was isolated from SI (duodenum through ileum; Fig.…”

“…Tissue preparation and immunohistochemistry were performed as previously described (7). Cells and tissues were fixed in 4% paraformaldehyde.…”

“…Many techniques have been employed to improve the neurogenic potential of enteric neurospheres. We have previously shown that co-transplantation of neurospheres with a serotonin receptor agonist can enhance neuronal differentiation and proliferation (7). Modified dissection techniques to isolate the enteric nerve plexuses from donor tissue have been described in order to generate a purer population (8).…”

Serotonergic Mechanisms Regulating the GI Tract: Experimental Evidence and Therapeutic Relevance

“…5-HT 4 signaling directly impacts postnatal gut-derived enteric neural stem/progenitor cell (ENSCs) proliferation as well. Neuronal proliferation and neuronal differentiation increase significantly in postnatal ENSCs and colon explants cultured with 5-HT 4 receptor agonist (RS67506)-loaded liposomal nanoparticles (Hotta et al 2016). Results are mimicked in vivo as co-transplantation of ENSCs with 5-HT4 receptor agonist-loaded nanoparticles leads to significantly increased neuronal density and proliferation (Hotta et al 2016).…”

Recent advances in regenerative medicine to treat enteric neuropathies: use of human cells

“…Formation of NLBs from dissociated gut cells is now a common method to isolate and study enteric neural progenitors . However, a recent study showed that not all NLBs of cells that form in cultures of dissociated gut cells contain neural stem cells and not all NLBs give rise to neural derivatives under differentiation conditions .…”