Demographic and auxological features of the children at admission and vital signs pre intervention.

<p>*data are expressed as median (interquartile range)</p><p>Demographic and auxological features of the children at admission and vital signs pre intervention.</p

Flow diagram of the progress through the phases of the randomised trial of two groups (animal-assisted therapy and standard care after surgery).

<p>Flow diagram of the progress through the phases of the randomised trial of two groups (animal-assisted therapy and standard care after surgery).</p

Comparison of the model-derived profiles for heart rate (HR), diastolic blood pressure (DBP), oxygen saturation (SpO₂), cerebral prefrontal oxygenation (HbO₂) in animal-assisted therapy (AAT) and standard (STAND) groups.

<p>All models are adjusted for time of anesthesia</p

Vital signs measured in treated and standard groups.

<p>Differences and 95%CI are derived from repeated measurements models (including main effects for time and groups and their interaction) adjusted for anesthesia time.</p

Characterization and localization of rat EGFP-MSC.

<p>1: Characteristic spindle-shaped morphology of bone marrow derived MSC obtained from one rat. Magnification X4. 2: Immunophenotype of one representative culture-expanded rat EGFP-MSC. Rat MSC were negative for CD11b and CD45 and positive for CD90, CD49e and CD29. 3: Osteogenic and adipogenic differentiation capacity of rat MSC. Differentiation into osteoblasts was demonstrated by the histological detection of Alkaline phosphatase activity (a) and calcium depositions positive for Alizarin Red (b). Differentiation into adipocytes was revealed by the formation of lipid droplets stained with Oil Red O (c). Magnification X20. 4: Representative renal sections of rats treated with MSC sacrified one day after ureteral ligation. Arrows indicate EGFP positive MSC. Immunohistochemistry for EGFP showing glomerular (a) tubular and interstitial (b) localization of MSC, magnification X200. Self fluorescence showed EGFP-MSC presence (c), magnification X400.</p

Renal fibrosis.

<p>A: Upper: Representative renal sections of IHC for FSP1 in all groups at day 7 (X200). Bottom: Representative renal sections of Masson’s trichrome staining in all groups at day 21 (X200). B: <i>Renal tissue TGF</i>β <i>levels</i>. Columns represent tissue TGFβ levels after 7 days from ureteral obstruction in all groups. Data are means ± SD. *p<0.001 vs sham, ^p<0.05 vs Vehicle D7. C: <i>FSP1 positive cells</i>. Columns represent FSP1 positive cells/HPF (means ± SD) at day 21. *p<0.001 vs all groups, °p<0.005 vs vehicle D21. D: <i>Interstitial fibrosis score</i> Columns represents the score of interstitial fibrosis (means ± SD) at day 21. *p<0.05 vs sham, °p<0.001 vs vehicle D21.</p

Tubular damage.

<p>A: <i>Tubular diameters</i>: Hematoxylin-eosin staining of representative renal sections of all groups of rats at day 7. Magnification, X200. B: <i>Tubular diameters</i> columns represent tubule diameters/HPF at day 7, results expressed as means ± SD. *p<0.0001 vs sham, ^p<0.05 vs MSC D7, ACEi D7 and ACEi+MSC D7. C: <i>Broken tubules</i>. Columns represent percentage of broken tubules/HPF at day 21. Results were expressed as means ± SD. *p<0.0001 vs sham, ^p<0.01 vs MSC D21 and Vehicle D21.</p

RENmRNA expression in co-coltures of MSC and HK2 or Mϕ.

<p>A: RENmRNA expression in HK2 cultured in low glucose (LG) medium, high glucose (HG) medium, in co-coltures with MSC (1:2, 1:20, 1:200) for 4 h. Columns indicate RENmRNA expression normalized to the βactin expression and converted in fold change. °p<0.05 vs HK2 LG; HK2 HG + MSC 1:2 4h. ^p< 0.01 vs HK2 HG + MSC 1:20 4h, HK2 HG + MSC 1:200 4h. *p<0.01 vs HK2 LG. B: RENmRNA expression in circulating monocytes (MC), macrophages (Mϕ), MSC+Mϕ co-coltures (1:2, 1:20, 1:200) for 4 days. Columns indicate RENmRNA expression normalized to βactin expression and converted in fold change. ^p<0.01 vs MC, *p<0.005 vs Mϕ. C: RENmRNA expression in HK2 cultured in HG medium in presence and absence of IL10 (10 ng/ml) for 1 h. *p<0.001 vs HK2 HG. D: RENmRNA expression in HK2 cultured in HG medium, in MSC/HK2 HG co-coltures (1:2) in presence and absence of anti IL10 blocking antibody at concentration 100 ng/ml for 4 h. Columns indicate RENmRNA expression normalized to the βactin expression and converted in fold change. *p<0.0001 vs HK2 HG, HK2 HG+MSC+anti IL10 Ab. E: RENmRNA expression in Mϕ cultured in presence and absence of IL10 (10 ng/ml) for 4 h. Columns indicate REN mRNA expression normalized to βactin expression and converted in fold change. *p<0.05 vs Mϕ. F: RENmRNA expression in Mϕ, MSC/Mϕ co-coltures (1:2) in presence and absence of anti IL10 blocking antibody at concentration 100 ng/ml for 24 h. Columns indicate RENmRNA expression normalized to βactin expression and converted in fold change. *p<0.05 vs Mϕ and Mϕ+MSC+IL10 Ab.</p

HuRmRNA, RENmRNA expression and renal Renin levels.

<p>A: HuRmRNA expression in Sham, Vehicle and MSC groups one day after ureteral ligation determined by real time PCR. Columns indicate HuRmRNA expression normalized to the βactin expression and converted in fold change. *p<0.001 vs Sham and MSC D1. B: RENmRNA expression in Sham, Vehicle, and MSC groups one day after ureteral ligation determined by real time PCR. Columns indicate RENmRNA expression normalized to the βactin expression and converted in fold change. *p<0.001 vs Sham, MSC D1. C: Renal Renin levels in Sham, Vehicle and MSC groups one day after ureteral ligation. Columns represent Renin levels. Data are means ± SD. °p<0.005 vs MSC D1, *p<0.0001 vs sham.</p

REN mRNA, renal Renin, serum AII and aldosterone levels.

<p>D7 indicates the day after ureteral ligation. A: RENmRNA expression in all groups of rats determined by real time PCR. Columns indicate RENmRNA expression normalized to the βactin expression and converted in fold change. °p<0.001 vs sham, *p<0.05 vs vehicle D7, ACEi D7 and ACEi+MSC D7, #p<0.05 vs ACEi D7. B: Renin protein levels in renal tissue in all groups of rats. Columns represent Renin protein levels. Data are means ± SD. °p<0.005 vs sham, MSC D7 and ACEi+MSC D7, *p<0.05 vs vehicle D7 and ACEiD7. C: AII serum levels in all groups of rats. Columns represent AII levels. Data are means ± SD. °p<0.005 vs all groups. D: Aldosterone serum levels in all groups of rats. Columns represent aldosterone serum levels. Data are means ± SD. °p<0.005 vs all groups.</p