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Stem cells derived from neonatal mouse kidney generate functional proximal tubule-like cells and integrate into developing nephrons in vitro.

Ranghini E, Fuente Mora C, Mora CF, Edgar D, Kenny SE, Murray P, Wilm B - PLoS ONE (2013)

Bottom Line: Finally, we compared the ability of KSCs to integrate into developing kidneys ex vivo with that of metanephric mesenchyme cells.We found that KSCs integrated into nascent nephrons to a similar extent as metanephric mesenchyme cells while both were excluded from ureteric bud branches.Our analysis of the behavior of the two cell types shows that some, but not all KSC characteristics are similar to those of the MM.

View Article: PubMed Central - PubMed

Affiliation: Institute of Translational Medicine, Faculty of Health and Life Sciences, The University of Liverpool, Liverpool, United Kingdom.

ABSTRACT
We have recently shown that kidney-derived stem cells (KSCs) isolated from the mouse newborn kidney differentiate into a range of kidney-specific cell types. However, the functionality and integration capacity of these mouse KSCs remain unknown. Therefore, the main objectives of this study were (1) to determine if proximal tubule-like cells, generated in vitro from KSCs, displayed absorptive function typical of proximal tubule cells in vivo, and (2) to establish whether the ability of KSCs to integrate into developing nephrons was comparable with that of metanephric mesenchyme (MM), a transient population of progenitor cells that gives rise to the nephrons during kidney organogenesis. We found that proximal tubule-like cells generated in vitro from mouse KSCs displayed megalin-dependent absorptive function. Subsequently, we used a chimeric kidney rudiment culture system to show that the KSCs could generate proximal tubule cells and podocytes that were appropriately located within the developing nephrons. Finally, we compared the ability of KSCs to integrate into developing kidneys ex vivo with that of metanephric mesenchyme cells. We found that KSCs integrated into nascent nephrons to a similar extent as metanephric mesenchyme cells while both were excluded from ureteric bud branches. Our analysis of the behavior of the two cell types shows that some, but not all KSC characteristics are similar to those of the MM.

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The extent of KSC integration into developing nephrons within chimeric rudiments is similar to that of MM cells.(A) Chimeras generated with QD+ (white)-labelled (i) freshly isolated MM, (ii) MM that had been cultured for 4 days, or (iii) KSCs, were cultured for 3 days and immunostained for Wt1 (green) and laminin-111 (red), or for Pax2 (green). Arrowheads point to QD+ cells integrated within the forming nephrons. Scale bars are 30 µm, 15 µm and 15 µm (upper row), and 36 µm, 31 µm and 10 µm (lower row). (B) The percentage of KSCs that integrated into developing nephrons was significantly lower than that of freshly isolated MM cells, but not statistically different to that of MM cells cultured for 4 days (Student’s t-test; P>0.05). Results are expressed as mean ± SE. n = 3 in each group; for each organoid, 7 random developing nephrons were selected for analysis.
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pone-0062953-g006: The extent of KSC integration into developing nephrons within chimeric rudiments is similar to that of MM cells.(A) Chimeras generated with QD+ (white)-labelled (i) freshly isolated MM, (ii) MM that had been cultured for 4 days, or (iii) KSCs, were cultured for 3 days and immunostained for Wt1 (green) and laminin-111 (red), or for Pax2 (green). Arrowheads point to QD+ cells integrated within the forming nephrons. Scale bars are 30 µm, 15 µm and 15 µm (upper row), and 36 µm, 31 µm and 10 µm (lower row). (B) The percentage of KSCs that integrated into developing nephrons was significantly lower than that of freshly isolated MM cells, but not statistically different to that of MM cells cultured for 4 days (Student’s t-test; P>0.05). Results are expressed as mean ± SE. n = 3 in each group; for each organoid, 7 random developing nephrons were selected for analysis.

Mentions: We next compared the ability of KSCs and in vitro cultured MM cells to integrate into developing nephrons using the kidney rudiment assay. For this purpose, both cell types were labelled with QDs and individually recombined with disaggregated E11.5 kidney rudiments. Following a 3-day culture period, the chimeric organoids were immunostained for Pax2, Wt1 and laminin to visualise developing renal structures. Chimeric organoids containing freshly isolated MM served as positive controls. The integration pattern of QD+ cells was similar in all three types of chimeras, as labelled cells were observed within Wt1+ aggregates and Pax2+ nephron tubules, but rarely detected within the UB (Figure 6A). To compare the extent of integration, the proportion of QD+ cells within developing nephrons was determined for the three types of chimeric organoid, as we have previously undertaken [40]. The percentage of QD+ freshly isolated MM cells, in vitro cultured MM cells and KSCs that integrated into developing nephrons was approximately 13%, 12% and 9%, respectively. Statistical analyses indicated that the extent of cultured MM cell integration was not significantly different than that observed with freshly isolated MM cells or KSCs, but the extent of KSC integration was significantly less than that of freshly isolated MM cells (Figure 6B). These data show that although the KSCs and MM cells are derived from different life stages (i.e., neonatal compared with embryonic), and display very different growth kinetics in culture, their integration behaviour within chimeric kidney rudiments ex vivo is strikingly similar.


Stem cells derived from neonatal mouse kidney generate functional proximal tubule-like cells and integrate into developing nephrons in vitro.

Ranghini E, Fuente Mora C, Mora CF, Edgar D, Kenny SE, Murray P, Wilm B - PLoS ONE (2013)

The extent of KSC integration into developing nephrons within chimeric rudiments is similar to that of MM cells.(A) Chimeras generated with QD+ (white)-labelled (i) freshly isolated MM, (ii) MM that had been cultured for 4 days, or (iii) KSCs, were cultured for 3 days and immunostained for Wt1 (green) and laminin-111 (red), or for Pax2 (green). Arrowheads point to QD+ cells integrated within the forming nephrons. Scale bars are 30 µm, 15 µm and 15 µm (upper row), and 36 µm, 31 µm and 10 µm (lower row). (B) The percentage of KSCs that integrated into developing nephrons was significantly lower than that of freshly isolated MM cells, but not statistically different to that of MM cells cultured for 4 days (Student’s t-test; P>0.05). Results are expressed as mean ± SE. n = 3 in each group; for each organoid, 7 random developing nephrons were selected for analysis.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3646983&req=5

pone-0062953-g006: The extent of KSC integration into developing nephrons within chimeric rudiments is similar to that of MM cells.(A) Chimeras generated with QD+ (white)-labelled (i) freshly isolated MM, (ii) MM that had been cultured for 4 days, or (iii) KSCs, were cultured for 3 days and immunostained for Wt1 (green) and laminin-111 (red), or for Pax2 (green). Arrowheads point to QD+ cells integrated within the forming nephrons. Scale bars are 30 µm, 15 µm and 15 µm (upper row), and 36 µm, 31 µm and 10 µm (lower row). (B) The percentage of KSCs that integrated into developing nephrons was significantly lower than that of freshly isolated MM cells, but not statistically different to that of MM cells cultured for 4 days (Student’s t-test; P>0.05). Results are expressed as mean ± SE. n = 3 in each group; for each organoid, 7 random developing nephrons were selected for analysis.
Mentions: We next compared the ability of KSCs and in vitro cultured MM cells to integrate into developing nephrons using the kidney rudiment assay. For this purpose, both cell types were labelled with QDs and individually recombined with disaggregated E11.5 kidney rudiments. Following a 3-day culture period, the chimeric organoids were immunostained for Pax2, Wt1 and laminin to visualise developing renal structures. Chimeric organoids containing freshly isolated MM served as positive controls. The integration pattern of QD+ cells was similar in all three types of chimeras, as labelled cells were observed within Wt1+ aggregates and Pax2+ nephron tubules, but rarely detected within the UB (Figure 6A). To compare the extent of integration, the proportion of QD+ cells within developing nephrons was determined for the three types of chimeric organoid, as we have previously undertaken [40]. The percentage of QD+ freshly isolated MM cells, in vitro cultured MM cells and KSCs that integrated into developing nephrons was approximately 13%, 12% and 9%, respectively. Statistical analyses indicated that the extent of cultured MM cell integration was not significantly different than that observed with freshly isolated MM cells or KSCs, but the extent of KSC integration was significantly less than that of freshly isolated MM cells (Figure 6B). These data show that although the KSCs and MM cells are derived from different life stages (i.e., neonatal compared with embryonic), and display very different growth kinetics in culture, their integration behaviour within chimeric kidney rudiments ex vivo is strikingly similar.

Bottom Line: Finally, we compared the ability of KSCs to integrate into developing kidneys ex vivo with that of metanephric mesenchyme cells.We found that KSCs integrated into nascent nephrons to a similar extent as metanephric mesenchyme cells while both were excluded from ureteric bud branches.Our analysis of the behavior of the two cell types shows that some, but not all KSC characteristics are similar to those of the MM.

View Article: PubMed Central - PubMed

Affiliation: Institute of Translational Medicine, Faculty of Health and Life Sciences, The University of Liverpool, Liverpool, United Kingdom.

ABSTRACT
We have recently shown that kidney-derived stem cells (KSCs) isolated from the mouse newborn kidney differentiate into a range of kidney-specific cell types. However, the functionality and integration capacity of these mouse KSCs remain unknown. Therefore, the main objectives of this study were (1) to determine if proximal tubule-like cells, generated in vitro from KSCs, displayed absorptive function typical of proximal tubule cells in vivo, and (2) to establish whether the ability of KSCs to integrate into developing nephrons was comparable with that of metanephric mesenchyme (MM), a transient population of progenitor cells that gives rise to the nephrons during kidney organogenesis. We found that proximal tubule-like cells generated in vitro from mouse KSCs displayed megalin-dependent absorptive function. Subsequently, we used a chimeric kidney rudiment culture system to show that the KSCs could generate proximal tubule cells and podocytes that were appropriately located within the developing nephrons. Finally, we compared the ability of KSCs to integrate into developing kidneys ex vivo with that of metanephric mesenchyme cells. We found that KSCs integrated into nascent nephrons to a similar extent as metanephric mesenchyme cells while both were excluded from ureteric bud branches. Our analysis of the behavior of the two cell types shows that some, but not all KSC characteristics are similar to those of the MM.

Show MeSH
Related in: MedlinePlus