Características del Comportamiento y niveles correspondientes

Over the past years, BMSCs have been the subject of extensive research and several studies showed that these cells are not lineage restricted and have the ability to cross boundaries and form components of other tissues, like the heart, liver or brain (14-17). Three different stem cell populations within the bone marrow have been identified that have the potential to contribute to tissue repair processes: the HSC population, MSC population and MAPC population. Several studies show the involvement of HSCs in renal regeneration in rodents

Mdr1a/1b(-/-) _{mice Bcrp}(-/-) _mice

Basal characteristics _{Increased diuresis (~200%) Increased diuresis (~300%)}

Normal kidney weight Increased kidney weight Normal gonadal weight Increased gonadal weight Slightly increased blood pressure Normal blood pressure

Renal function Decreased GFR Increased GFR

Proximal tubular dysfunction Normal proximal tubular function Increased excretion of low-molecular weight plasma

proteins Increased excretion of plasma proteins

Basal serum analysis Normal glucose concentration Increased glucose concentration Normal insulin concentration Increased insulin concentration

Hormone regulation Normal testosterone levels Increased testosterone levels (1000%) Normal LH levels Increased LH levels

(18-20), although their contribution is limited ((21), this thesis, chapter 7). Interestingly, we found that the contribution of BMSCs in renal repair increased to 20-30% when bone marrow was isolated from P-gp- or BCRP-deficient mice. MSCs may also directly participate in renal regeneration through differentiation towards proximal tubular cells, but also exhibit a supportive function for other BMSCs migrating towards the injured tissue (22,23). Finally, MAPC are candidate cells in renal repair. Although to date transplantation studies with MAPC in renal repair models have not been performed, a role in renal regeneration is likely because of their capacity to differentiate towards renal epithelial cells in vitro.

Not much is known about the capacity of BMSCs in renal repair in humans. Most studies were performed with rodents and translation to the human situation is difficult. Not only because of species differences, but also because of variations in models applied. For example, most studies in rodents used transplantations performed before the induction of ischemia, while in human studies, transplantation of cells is carried out after the insult. The contribution of bone marrow derived donor cells to repaired tubules was less then 1% (24). HSCs did not give rise to new tubules, but were found to be involved in vasculogenesis (25). This was consistent with findings observed in an animal model of glomerulonephritis, in which bone marrow-derived endothelial and mesangial cells integrate in glomerular structures, and, eventually, contribute to microvascular repair (26). In another study, the migration towards a donor kidney of human mesenchymal progenitor cells was shown, suggesting that donor bone marrow cells might play a role in renal tissue remodeling (27). Hence, the amount of BMSCs engrafting the kidney seems to be small, indicating that other mechanisms of repair are more important. Our findings with transporter-deficient full bone marrow transplantations point to a shift towards a more prominent role for BM-derived cells in remodeling. It is unclear if SP cells or other BM cells are responsible for this beneficial effect. Migration into the kidney of bone marrow cells lacking P-gp or BCRP was faster, but also recovery of renal function was better compared to WT mice. Probably, this phenomenon is connected to the resistance against renal ischemia observed in both knockout mice. An increased resistance to apoptosis in deficient bone marrow cells may have also contributed to the beneficial effect.

It seems likely that the increased population of macrophages and granulocytes in the donor bone marrow of the knockout mice is partly responsible for the increased regenerative

Chapter 9

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process seen in our transplantation study. As a proof of principle, BCRP-deficient mice were transplanted with WT bone marrow. These mice showed a decreased renal function after AKI compared to BCRP-deficient mice without BM transplantation, although renal function was still better as compared to WT bone marrow transplantations in WT mice. This suggests that the contribution of bone marrow-derived cells in renal regeneration is high enough to influence renal function after AKI.

The increased percentage of macrophages and granulocytes within the bone marrow of P-gp- and BCRP-deficient mice might facilitate angiogenesis in the injured kidney, thus supporting renal repair. However, macrophages are also able to influence renal regeneration directly (28,29), although controversial findings were reported. A negative contribution of macrophages to renal regeneration (30), as well as a positive influence of macrophage infiltration in renal regeneration has been reported (28,29). Whether the increased population of macrophages in the bone marrow of BCRP- and P-gp-deficient mice is responsible for the increased regeneration in our studies remains to be determined. Macrophage phenotype might be switched or altered towards a phenotype more involved in tissue regeneration. Mice deficient in P-gp and Multidrug Resistance Protein 1 (MRP1; mrp1/mdr1a/1b-double knockout mice) expression showed a decreased inflammatory response towards cigarette smoking (31). This was likely caused by decreased systemic levels of cytokines and chemokines, but also by the altered dendritic cell migration. Subsequently, this might lead to a decreased inflammatory reaction observed in these animals which could inhibit tissue repair. However, the decreased inflammatory reaction may also have a positive effect in a way that repair mechanisms are enhanced. The composition of cytokines released by these cells could be altered, resulting in a shift between vasculogenesis and inflammation. In line with the results of our study, an increased number of macrophages would indirectly stimulate regeneration by tissue remodeling and vasculogenesis. We observed that the amount of early endothelial progenitor cells in the bone marrow of both knockout mice was increased. The altered balance between inflammation and angiogenesis, controlled by cytokines and chemokines, may result in an increased vascularisation in which also endothelial progenitor cells are involved. Eventually, it is possible that these mechanisms lead to the stimulation of renal regeneration, as we observed in both knockout mice, and even more pronounced in our transplantation experiments.