One of the most common complications associated with intestinal transplantation (ITx) is graft rejection due to its high donor lymphoid cell burden. It is well known that Peyer’s Patches (PP) are necessary to develop the early response to acute cell rejection (ACR). High doses of immunosuppressants are needed in order to prevent the allogeneic response but have a negative impact in the long term. Regulatory T cells (T regs) are major players in the induction of graft tolerance. On the other hand, goblet cells (GC) actively secrete mucus to the intestinal lumen, having a key role in mucosal protection, therefore, their depletion potentially increases microbial translocation risk.

Allogeneic heterotopic ITx procedures were performed in rats, with Sprague Dawley strain as donors and Wistar GFP+ as recipients. The recipients were divided into two groups: one receiving no immunosuppressant treatment (n-IS group), and the other receiving tacrolimus at a dosage of 0.6 mg/kg/day administered subcutaneously for 7 days as immunosuppressive therapy (TAC group), and the animals were kept alive until day 28. Graft samples were collected at 0, 4, 7, and 10 postoperative days (POD) for the n-IS group, and at 0, 4, 7, 14, 21, and 28 POD for the TAC group. Histopathological diagnosis of ACR was conducted using the Wu score. Flow cytometry assays were employed to determine the frequency of CD3+, CD4+, CD8+ and Tregs, as well as their GFP expression levels within the grafts. Alcian Blue staining was used to count GC in graft samples.

Histopathological analysis revealed that the TAC group developed moderate to severe ACR between 21 and 28 POD (p<0.0001, Kruskal-Wallis test), whereas in the n-IS group it occurred between 7 and 10 POD. Furthermore, a significant correlation was observed between rejection severity and GC loss (p<0.0001, linear regression test). An increase of T regs frequency was observed both in n-IS and TAC group at 4 POD, and this trend was maintained in the TAC group until at least 14 POD and then declined to baseline. The analysis of recipient lymphocyte showed an early turnover o CD4+ and CD8+ populations in PP compartment, with 25 to 50% lymphocytes being of recipient origin since 4 POD. Moreover, in the lamina propria (LP) compartment, the TAC group exhibited delayed recipient CD4+ turnover kinetics until 21 POD, while recipient CD8+ turnover kinetics was delayed until 14 POD. These results are concordant with the progression of the ACR process.

Our results suggest that Tacrolimus at low doses can delay turnover kinetics of T-cell compartment in the lamina propria without affecting turnover in Peyer's Patch compartment. This dynamic allows expansion of T regs and possibly providing transient protection to the integrity of the graft. Pro-homeostatic cells, such as T regs and Goblet cells, decrease during the progression of acute cellular rejection.