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Novel functional changes during podocyte differentiation: increase of oxidative resistance and H-ferritin expression.

Bányai E, Balogh E, Fagyas M, Arosio P, Hendrik Z, Király G, Nagy G, Tánczos B, Pócsi I, Balla G, Balla J, Bánfalvi G, Jeney V - Oxid Med Cell Longev (2014)

Bottom Line: We observed that differentiated podocytes were highly resistant to oxidants such as H2O2 and heme when applied separately or in combination, whereas undifferentiated cells were prone to such challenges.Elevated oxidative resistance of differentiated podocytes was associated with increased activities of antioxidant enzymes and H-ferritin expression.Immunohistochemical analysis of normal human kidney specimens revealed that podocytes highly express H-ferritin in vivo as well.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Debrecen, Debrecen 4032, Hungary.

ABSTRACT
Podocytes are highly specialized, arborized epithelial cells covering the outer surface of the glomerular tuft in the kidney. Terminally differentiated podocytes are unable to go through cell division and hereby they are lacking a key property for regeneration after a toxic injury. Podocytes are long-lived cells but, to date, little is known about the mechanisms that support their stress resistance. Our aim was to investigate whether the well-known morphological changes during podocyte differentiation are accompanied by changes in oxidative resistance in a manner that could support their long-term survival. We used a conditionally immortalized human podocyte cell line to study the morphological and functional changes during differentiation. We followed the differentiation process for 14 days by time-lapse microscopy. During this period nondifferentiated podocytes gradually transformed into large, nonproliferating, frequently multinucleated cells, with enlarged nuclei and opened chromatin structure. We observed that differentiated podocytes were highly resistant to oxidants such as H2O2 and heme when applied separately or in combination, whereas undifferentiated cells were prone to such challenges. Elevated oxidative resistance of differentiated podocytes was associated with increased activities of antioxidant enzymes and H-ferritin expression. Immunohistochemical analysis of normal human kidney specimens revealed that podocytes highly express H-ferritin in vivo as well.

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Podocyte growth under nonpermissive (37°C) condition. At 50–60% confluence, nondifferentiated cells were placed from 33°C to 37°C to induce differentiation which was followed by video microscopy for 14 days. Cell numbers were determined by using time-lapse phase contrast images that were acquired one frame every 10 minutes. Graph is a representative of 3 experiments with similar results.
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fig1: Podocyte growth under nonpermissive (37°C) condition. At 50–60% confluence, nondifferentiated cells were placed from 33°C to 37°C to induce differentiation which was followed by video microscopy for 14 days. Cell numbers were determined by using time-lapse phase contrast images that were acquired one frame every 10 minutes. Graph is a representative of 3 experiments with similar results.

Mentions: We placed nondifferentiated human podocytes expressing the thermosensitive A58 T antigen at 50–60% confluency from 33°C to 37°C to induce differentiation. We followed changes in cell number and morphology by video microscopy for 14 days. By analysing the growth curve we observed that the initial exponential growth phase lasted for 9 days (Figure 1). After cell number reached its peak between days 9 and 10, we saw a slight decrease in cell number followed by stabilization of the culture (days 10–14) when we could hardly detect any mitotic event. When the temperature was shifted to 37°C the morphology of nondifferentiated podocytes gradually transformed into that of differentiated cells (Figures 1 and 2). At permissive temperature (33°C) nondifferentiated podocytes showed typical epithelial cobblestone morphology. In contrast, differentiated podocytes are characterized by enlarged cell bodies with an irregular shape (Figure 2(a), broken outlines) and the formation of processes involving shorter and rounded (Figure 2(a), white arrows) as well as longer and spindle-like projections (Figure 2(a), black arrows). Formations of spindle-like projections were often preceded by partial retraction of cytoplasmatic protrusions. Many differentiated cells developed cellular hypertrophy, some of the cells died (Figure 2(a), red asterisks), and others started to arborize. We detected numerous binucleated but nonproliferating cells. After day 9 we registered a reduction of cell number, but the motility of cells remained high even after differentiation, and the culture proved capability of migration until all bare fields of the culturing vessel were completely covered.


Novel functional changes during podocyte differentiation: increase of oxidative resistance and H-ferritin expression.

Bányai E, Balogh E, Fagyas M, Arosio P, Hendrik Z, Király G, Nagy G, Tánczos B, Pócsi I, Balla G, Balla J, Bánfalvi G, Jeney V - Oxid Med Cell Longev (2014)

Podocyte growth under nonpermissive (37°C) condition. At 50–60% confluence, nondifferentiated cells were placed from 33°C to 37°C to induce differentiation which was followed by video microscopy for 14 days. Cell numbers were determined by using time-lapse phase contrast images that were acquired one frame every 10 minutes. Graph is a representative of 3 experiments with similar results.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4109136&req=5

fig1: Podocyte growth under nonpermissive (37°C) condition. At 50–60% confluence, nondifferentiated cells were placed from 33°C to 37°C to induce differentiation which was followed by video microscopy for 14 days. Cell numbers were determined by using time-lapse phase contrast images that were acquired one frame every 10 minutes. Graph is a representative of 3 experiments with similar results.
Mentions: We placed nondifferentiated human podocytes expressing the thermosensitive A58 T antigen at 50–60% confluency from 33°C to 37°C to induce differentiation. We followed changes in cell number and morphology by video microscopy for 14 days. By analysing the growth curve we observed that the initial exponential growth phase lasted for 9 days (Figure 1). After cell number reached its peak between days 9 and 10, we saw a slight decrease in cell number followed by stabilization of the culture (days 10–14) when we could hardly detect any mitotic event. When the temperature was shifted to 37°C the morphology of nondifferentiated podocytes gradually transformed into that of differentiated cells (Figures 1 and 2). At permissive temperature (33°C) nondifferentiated podocytes showed typical epithelial cobblestone morphology. In contrast, differentiated podocytes are characterized by enlarged cell bodies with an irregular shape (Figure 2(a), broken outlines) and the formation of processes involving shorter and rounded (Figure 2(a), white arrows) as well as longer and spindle-like projections (Figure 2(a), black arrows). Formations of spindle-like projections were often preceded by partial retraction of cytoplasmatic protrusions. Many differentiated cells developed cellular hypertrophy, some of the cells died (Figure 2(a), red asterisks), and others started to arborize. We detected numerous binucleated but nonproliferating cells. After day 9 we registered a reduction of cell number, but the motility of cells remained high even after differentiation, and the culture proved capability of migration until all bare fields of the culturing vessel were completely covered.

Bottom Line: We observed that differentiated podocytes were highly resistant to oxidants such as H2O2 and heme when applied separately or in combination, whereas undifferentiated cells were prone to such challenges.Elevated oxidative resistance of differentiated podocytes was associated with increased activities of antioxidant enzymes and H-ferritin expression.Immunohistochemical analysis of normal human kidney specimens revealed that podocytes highly express H-ferritin in vivo as well.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Debrecen, Debrecen 4032, Hungary.

ABSTRACT
Podocytes are highly specialized, arborized epithelial cells covering the outer surface of the glomerular tuft in the kidney. Terminally differentiated podocytes are unable to go through cell division and hereby they are lacking a key property for regeneration after a toxic injury. Podocytes are long-lived cells but, to date, little is known about the mechanisms that support their stress resistance. Our aim was to investigate whether the well-known morphological changes during podocyte differentiation are accompanied by changes in oxidative resistance in a manner that could support their long-term survival. We used a conditionally immortalized human podocyte cell line to study the morphological and functional changes during differentiation. We followed the differentiation process for 14 days by time-lapse microscopy. During this period nondifferentiated podocytes gradually transformed into large, nonproliferating, frequently multinucleated cells, with enlarged nuclei and opened chromatin structure. We observed that differentiated podocytes were highly resistant to oxidants such as H2O2 and heme when applied separately or in combination, whereas undifferentiated cells were prone to such challenges. Elevated oxidative resistance of differentiated podocytes was associated with increased activities of antioxidant enzymes and H-ferritin expression. Immunohistochemical analysis of normal human kidney specimens revealed that podocytes highly express H-ferritin in vivo as well.

Show MeSH
Related in: MedlinePlus