Oral Delivery of R-spondin1-Loaded Small Extracellular Vesicles Activates WNT Signalling Pathway to Accelerate Intestinal Injury Repair and Reverse Ageing.

RSPO1 protein can be loaded onto the surface of small extracellular vesicles (sEV) via heparan sulfate proteoglycans (HSPGs).

• RSPO1 binding to sEV was mediated through heparan sulfate proteoglycans present on the vesicle surface.
• The resulting construct, termed evRSPO1, retained active RSPO1 protein on the sEV surface.
• Heparinase treatment or HSPG-deficient conditions disrupted RSPO1 loading, confirming the HSPG-dependent mechanism.

sEV-delivered RSPO1 (evRSPO1) effectively induces WNT/β-catenin signalling activity in vitro.

• evRSPO1 enhanced WNT/β-catenin signalling-inducing activity compared to free RSPO1 or unloaded sEV.
• evRSPO1 promoted intestinal stem cell (ISC) proliferation in cell-based assays.
• evRSPO1 supported intestinal organoid growth in vitro, demonstrating functional WNT pathway activation.

Oral administration of evRSPO1 activates the WNT/β-catenin signalling pathway specifically in the cryptic stem cell niche in vivo.

• Following oral delivery, evRSPO1 reached the intestinal crypts and activated WNT/β-catenin signalling in the stem cell niche.
• Activation was demonstrated in a radiation-induced intestinal injury mouse model.
• The sEV vehicle enabled evRSPO1 to overcome biological barriers associated with oral delivery of protein therapeutics.

Oral evRSPO1 accelerated tissue repair and regeneration in a radiation-induced intestinal injury model.

• Radiation-induced intestinal injury was used as the in vivo injury model.
• evRSPO1 treatment accelerated intestinal tissue repair and regeneration compared to controls.
• The therapeutic effect was attributed to ISC proliferation driven by WNT/β-catenin pathway activation in the cryptic niche.

evRSPO1 treatment induced ISC proliferation and reversed intestinal senescence phenotypes in aged mice.

• Aged mice treated with evRSPO1 showed increased intestinal stem cell proliferation.
• evRSPO1 reversed the intestinal senescence phenotype observed in aged mice.
• These results suggest evRSPO1 has potential for intestinal rejuvenation in the context of ageing.

The study establishes evRSPO1 as a potential first-in-class orally deliverable therapeutic for intestinal regeneration.

• Effective oral delivery of functional WNT signalling agonists to the gut was described as a previously challenging problem.
• The sEV platform overcame biological barriers to enable oral delivery of active RSPO1 protein.
• The authors describe evRSPO1 as a 'potential first-in-class, orally deliverable therapeutic' for ISC activation, intestinal tissue repair, and rejuvenation.