We report a the detailed spatiotemporal evaluation of the local and systemic immune responses following allogeneic CDC transplantation for myocardial repair. Allogeneic CDC transplantation without immunosuppression is safe, and produces structural and functional benefits post-MI by stimulating endogenous cardiac regeneration. This indirect mechanism of action, shared by syngeneic cells, explains why benefits persist despite the temporary engraftment of transplanted cells.
CDCs represent an attractive cell type for heart repair and regeneration. CDCs are clonogenic and exhibit multineage potential, thus fulfilling key criteria for heart-derived stem cells15
. Over the past 6 years, we have demonstrated that CDCs can improve cardiac function post-MI in mice5,7,21
. Importantly, several independent labs worldwide have reproduced the published methodology and verified CDCs’ identity and utility26–32
. On the other hand, critiques of the cardiosphere methodology have appeared33,34
, but, as we have pointed out in detailed rebuttals8,15
, these studies did not follow published protocols for CDC isolation and expansion, and the methodological variations likely explain the negative results. With regard to clinical translation, highly positive results from a proof-of-concept clinical study utilizing autologous CDCs—the CADUCEUS (CArdiosphere-Derived aUtologous stem CElls to reverse ventricUlar dySfunction; NCT008933603
) trial— have recently been reported35
The avoidance of immunologic rejection renders autologous therapy attractive, but serious disadvantages dampen enthusiasm. Patient-specific tissue harvesting and cell processing result in a delay to therapy and introduce possible variations in cell potency related to patient age and disease4
. Here, we tested the specific hypothesis that allogeneic CDCs are hypoimmunogenic in vivo
and can survive in the infarcted myocardium for a critical period of time in order to stimulate endogenous reparative and regenerative pathways, resulting in sustained benefit. We find that allogeneic CDC transplantation without immunosuppression induces only a transient mild local immune reaction in a rat MI model. In the clinical setting, development of an immune response after allogeneic CDC delivery to the heart could theoretically lead to: a) immune-related myocardial damage (which based on our findings would be unlikely, since no foci of myocardial damage were detected even after xenogeneic CDC transplantation); and b) allosensitization of the cell recipient, which in turn could: i) complicate repeat dosing with the same batch of cells (a problem that could be easily overcome by administering cells from different donors); and ii) complicate future organ transplantation (if the CDC donor and organ donor share similar HLA haplotypes). We did not detect any circulating anti-donor antibodies after allogeneic CDC transplantation, implying that no significant increase in panel reactive antibodies would occur; the absence of such sensitization at baseline increases the likelihood of allograft survival36
. In addition, the small inoculum associated with CDC therapy (compared for example with the volumes used in blood transfusions) make sensitization of the recipient improbable. Nevertheless, the possibility of recipient allosensitization should be investigated in large animals as a prelude to studies in human subjects.
We have also shown that transient and rather paltry short-term cell survival suffices to produce dramatic lasting benefits. Despite lower cell engraftment, allogeneic CDC transplantation generates structural and functional benefits which are indistinguishable from syngeneic transplantation, and persist 6 months post-MI. The equivalence of allogeneic and syngeneic transplantation is not surprising once we recognize and accept the central paradox: few, briefly-present transplanted cells are sufficient to produce large, durable benefits by amplifying endogenous pathways of repair and regeneration, rather than by directly generating new transplanted tissue. This indirect “amplifier effect”, impressive as it may be, is not yet fully understood2
. Even though we show that allogeneic CDCs stimulate host cardiomyocyte cycling, endogenous stem cell recruitment and angiogenesis in the post-MI setting, it is unclear whether these phenomena can account for the totality of the observed benefit; other mechanisms could involve cytoprotection of the host tissue, or modulation of inflammatory processes resulting in better infarct healing. In addition, it is unclear how much of the benefit is attributable to the identified paracrine factors; IGF1 and HGF have been shown to mobilize resident cardiac stem cells37
, while VEGF is well-known to stimulate angiogenesis38
. Alternatively, other factors39
may also play important roles. Identification of the appropriate “cocktail” of beneficial growth factors and incorporation into a formulation enabling sustained and controlled local release after cardiac delivery is a conceptually attractive approach. However, cell-mediated contact-dependent mechanisms may also contribute to the observed effects.
Regardless of the mechanism, in practice, the current work opens up a new treatment paradigm: CDCs could be grown in large numbers from allogeneic heart tissue in a central facility under strict quality control and banked for future use, enabling safe and effective myocardial repair in a timely, cost-efficient manner. Potential sources of allogeneic heart tissue include hearts explanted from organ donors but not used for transplantation, cadaveric hearts from the recently deceased, and surgical discards. Hearts obtained from organ donors (but not used for transplantation) have the inherent advantage that donors are, by definition, healthy and have been previously HLA-typed and screened for infectious diseases, with the tissue maintained viable and sterile until processed. Hearts from organ donors after cardiac death are particularly attractive, since they are rarely used for transplant, although kidneys, liver, and pancreas are commonly used40
. In 2008 there were 832 organ donors after cardiac death in the US, and no hearts were used for cardiac transplantation41
; these hearts represent one pool from which source tissue can be obtained for allogeneic CDC culture. Cadaveric hearts from healthy, non-infectious donors could be also be used, however, the tissue is not optimally stored and samples would have to be obtained with low post-mortem intervals. Surgical discards are yet another source option; while these specimens are more abundant, donors are apt to have an existing cardiac disorder or other comorbidities (which may or may not hamper cell quality).
In conclusion, we demonstrate that allogeneic CDC transplantation without immunosuppression is safe, promotes cardiac regeneration and improves heart function in a rat MI model, mainly through stimulation of endogenous repair mechanisms. This indirect mechanism of action rationalizes the lasting benefit brought about by ephemeral transplanted cells, in that the new tissue originates from the recipient rather than the donor. This work motivates the testing of allogeneic human CDCs as a potential clinical product for cellular cardiomyoplasty.