The role of T lymphcytes, especially the Th1 and Th2 lymphocyte phenotypes, in cell mediated immune responses to xenografts has been widely studied [61
]. Th1 lymphocytes produce cytokines such as interleukin (IL)-2, interferon (IFN)-γ, and tumor necrosis factor (TNF)-β leading to macrophage activation, stimulation of complement fixing Ab isotypes (IgG2a
in mice) and differentiation of CD8+
cells to a cytotoxic phenotype [63
]. Activation of this pathway is associated with both allogeneic and xenogeneic transplant rejection [61
]. Th2 lymphocytes produce IL-4, IL-5, IL-6, and IL-10, cytokines that do not activate macrophages and that lead to production of non-complement fixing Ab isotypes (IgG1
in mice). Activation of the Th2 pathway is associated with transplant acceptance [66
The only ECM scaffold material for which the Th1/Th2 response has been characterized is SIS-ECM [69
]. In one study, SIS-ECM was implanted subcutaneously into mice and the host response was compared to the response elicited by either xenogeneic or syngeneic muscle tissue. Histologically, the xenogeneic muscle tissue showed the presence of polymorphonuclear leukocytes (PMNs) within 1 day, with a transition to a mixed population of neutrophils, T lymphocytes, and multinucleate giant cells by day 10. After 28 days, the xenogeneic muscle showed evidence of necrosis, granuloma formation, and encapsulation; all of which are indicative of graft rejection. The syngeneic muscle tissue showed an acute inflammatory response, most of which disappeared by day 10. After 28 days, the residual mild chronic inflammatory response had resolved and the graft site showed organized tissue morphology, consistent with graft acceptance. The host response to the SIS-ECM was similar to that for the syngeneic muscle tissue. At Day 1, PMNs were present at the interface of the host tissue with the SIS-ECM device. After 10 days, the cellular infiltration was reduced and consisted primarily of mononuclear cells. By 28 days, the mononuclear cell infiltrate had diminished and the remodeling response was nearly complete. The mice did not develop an acquired adverse immune response to the SIS-ECM, and analysis of tissue cytokines showed that SIS-ECM strongly increased the expression of IL-4 (Th2), while the expression of IFN-γ (Th1) was 100 fold less than the response elicited by the xenogeneic muscle group. The SIS-ECM implanted mice developed an Ab response that was restricted to the IgG1
isotype, which is most consistent with the Th2 pathway. Thus, although there was a vigorous immune response to the SIS-ECM after implantation, the response was dominated by the Th2 pathway mediators.
To confirm that the immune response to SIS-ECM was due to Th2 restriction rather than lack of sufficient antigen stimulation, Ab responses were measured in mice that received two sequential implants of SIS-ECM 28 days apart [69
]. These animals showed a significant secondary antibody response, but the response was still exclusively of the IgG1
isotype. There was no evidence of Th1 cytokines at the secondary graft site. No deposits of IgG1
Ab were found in the SIS-ECM graft. This double implantation study was also conducted with ECM derived from a different tissue source, specifically the porcine urinary bladder submucosa (UBS). The results were very similar confirming that the source of ECM did not alter the restricted Th2 immune response.
The SIS-ECM has been implanted in T cell KO mice and B cell KO mice [69
]. In the T cell KO mice, no IL-4 expression was found, showing that T cells are the source of the IL-4 mRNA observed in SIS implanted in WT mice. Anti-SIS Ab were absent in both T cell KO mice and B cell KO mice; however, in both cases, the SIS-ECM scaffold was completely remodeled within 28 days. These results confirm that T and B cells do indeed respond to SIS-ECM, but are not required for SIS-ECM acceptance and a constructive remodeling response.
Two mouse models were used to examine the effects of SIS-ECM implantation upon systemic immunity [70
]. Mice implanted with SIS-ECM expressed levels of influenza specific Ab of the IgG1
subtypes after vaccination with a T-dependent subunit vaccine. The vaccine response was comparable to that of mice not implanted with SIS-ECM. Furthermore, challenge of immunized and SIS-implanted mice showed the same survival rate as mice that did not receive the SIS implants. A second model of immune function examined the response to a deliberate bacterial infection following SIS implantation. The mice were immunized with a T-independent polysaccharide vaccine, produced Ab to S. pneumoniae,
and survived a lethal dose of the bacteria with or without SIS-ECM implantation. In several other studies, ECM scaffolds with deliberate bacterial exposure have been shown to resist infection even without previous immunization [14
]. The Th2 response elicited by SIS-ECM does not adversely affect the host’s ability to mount a protective systemic immune response to T-dependent or T-independent vaccines, and to overcome viral or bacterial infections.
Cell mediated immune responses were analyzed using delayed type hypersensitivity and cytotoxic T cell reactions [70
]. In a mouse model of contact dermatitis, topical application of dinitrofluorobenzene led to similar levels of cellular infiltration in both SIS-ECM implanted mice and in mice not implanted with SIS-ECM. Similarly, SIS-ECM implantation did not increase or decrease the ability of mice to reject xenogeneic skin grafts. Thus, SIS-ECM implantation does not impair cell mediated immune responses to antigens.
Since similar studies have not been conducted for other forms of ECM scaffold materials, it is not possible to determine whether they would elicit the same type of host response. Considering the diversity of tissue sources and processing methods from which ECM scaffolds are produced, it seems likely that the host response to biologic scaffold prepared from different sources will vary to a large degree following implantation.