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C-myc-induced apoptosis in polycystic kidney disease is Bcl-2 and p53 independent.

Trudel M, Lanoix J, Barisoni L, Blouin MJ, Desforges M, L'Italien C, D'Agati V - J. Exp. Med. (1997)

Bottom Line: No renal abnormalities were detected in 13 transgenic lines established, indicating that the PKD phenotype is dependent on functions specific to c-myc.All SBM offspring, irrespective of their p53 genotype, developed PKD with increased renal epithelial apoptotic index.We conclude that the pathogenesis of PKD is c-myc specific and involves a critical imbalance between the opposing processes of cell proliferation and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Institut de Recherches Cliniques de Montréal, Faculté de Médecine de l'Université de Montréal, Montréal, Québec, Canada H2W 1R7.

ABSTRACT
The SBM mouse is a unique transgenic model of polycystic kidney disease (PKD) induced by the dysregulated expression of c-myc in renal tissue. In situ hybridization analysis demonstrated intense signal for the c-myc transgene overlying tubular cystic epithelium in SBM mice. Renal proliferation index in SBM kidneys was 10-fold increased over nontransgenic controls correlating with the presence of epithelial hyperplasia. The specificity of c-myc for the proliferative potential of epithelial cells was demonstrated by substitution of c-myc with the proto-oncogene c-fos or the transforming growth factor (TGF)-alpha within the same construct. No renal abnormalities were detected in 13 transgenic lines established, indicating that the PKD phenotype is dependent on functions specific to c-myc. We also investigated another well characterized function of c-myc, the regulation of apoptosis through pathways involving p53 and members of the bcl-2 family, which induce and inhibit apoptosis, respectively. The SBM kidney tissues, which overexpress c-myc, displayed a markedly elevated (10-100-fold) apoptotic index. However, no significant difference in bcl-2, bax, or p53 expression was observed in SBM kidney compared with controls. Direct proof that the heightened renal cellular apoptosis in PKD is not occurring through p53 was obtained by successive matings between SBM and p53(-/-) mice. All SBM offspring, irrespective of their p53 genotype, developed PKD with increased renal epithelial apoptotic index. In addition, overexpression of both bcl-2 and c-myc in double transgenic mice (SBB+/SBM+) also produced a similar PKD phenotype with a high apoptotic rate, showing that c-myc can bypass bcl-2 in vivo. Thus, the in vivo c-myc apoptotic pathway in SBM mice occurs through a p53- and bcl-2-independent mechanism. We conclude that the pathogenesis of PKD is c-myc specific and involves a critical imbalance between the opposing processes of cell proliferation and apoptosis.

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Detection of SBF mRNA by in situ hybridization in transgenic kidney tissues. (a) In situ hybridization analysis of c-fos mRNA in  transgenic SBF adult kidneys detected intense signal over the epithelial  cells lining the normal-appearing tubules. (Hematoxylin-eosin counterstain, ×200). (b) In situ hybridization for c-fos mRNA in non-transgenic  controls of the same genetic background (C57BL/6 × CBA)F1 is negative, with low background levels. (c) In SBM kidneys, in situ hybridization for c-fos is negative. The renal cysts are visible even with dark-field  optics.
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Figure 4: Detection of SBF mRNA by in situ hybridization in transgenic kidney tissues. (a) In situ hybridization analysis of c-fos mRNA in transgenic SBF adult kidneys detected intense signal over the epithelial cells lining the normal-appearing tubules. (Hematoxylin-eosin counterstain, ×200). (b) In situ hybridization for c-fos mRNA in non-transgenic controls of the same genetic background (C57BL/6 × CBA)F1 is negative, with low background levels. (c) In SBM kidneys, in situ hybridization for c-fos is negative. The renal cysts are visible even with dark-field optics.

Mentions: Fig. 3 a shows RT-PCR analysis of the SBF transgene in the various organs (including kidney, liver, spleen, heart, lung, and brain). The highest levels of SBF transgene expression were obtained in kidney, where levels were demonstrated to be within the linear range (Fig. 3 b). Some expression was also occasionally detectable in spleen and lung, with little or undetectable expression in other organs (Fig. 3 a). This particular organ distribution of transgene expression closely resembles that described previously for SBM mice (35), and it is likely that the transgene is preferentially targeted to the kidney due to common regulatory elements with SBM. Furthermore, to determine more precisely the localization of the c-fos transgene in SBF kidneys, we have carried out in situ hybridization. Adult transgenic SBF kidneys demonstrated high expression of the transgene in tubular epithelial cells throughout the kidney, with particularly intense signal over the medullary collecting tubules (Fig. 4 a). This cellular localization is very similar to that observed for the c-myc transgene. No expression was detected in renal tissue of non transgenic controls (C57BL/6 × CBA)F1 (Fig. 4 b). Furthermore, c-fos expression was also investigated in the SBM mouse kidneys (Fig. 4 c); the absence of signal indicates that c-fos is not induced in the SBM c-myc–dependent cystogenic pathway.


C-myc-induced apoptosis in polycystic kidney disease is Bcl-2 and p53 independent.

Trudel M, Lanoix J, Barisoni L, Blouin MJ, Desforges M, L'Italien C, D'Agati V - J. Exp. Med. (1997)

Detection of SBF mRNA by in situ hybridization in transgenic kidney tissues. (a) In situ hybridization analysis of c-fos mRNA in  transgenic SBF adult kidneys detected intense signal over the epithelial  cells lining the normal-appearing tubules. (Hematoxylin-eosin counterstain, ×200). (b) In situ hybridization for c-fos mRNA in non-transgenic  controls of the same genetic background (C57BL/6 × CBA)F1 is negative, with low background levels. (c) In SBM kidneys, in situ hybridization for c-fos is negative. The renal cysts are visible even with dark-field  optics.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199149&req=5

Figure 4: Detection of SBF mRNA by in situ hybridization in transgenic kidney tissues. (a) In situ hybridization analysis of c-fos mRNA in transgenic SBF adult kidneys detected intense signal over the epithelial cells lining the normal-appearing tubules. (Hematoxylin-eosin counterstain, ×200). (b) In situ hybridization for c-fos mRNA in non-transgenic controls of the same genetic background (C57BL/6 × CBA)F1 is negative, with low background levels. (c) In SBM kidneys, in situ hybridization for c-fos is negative. The renal cysts are visible even with dark-field optics.
Mentions: Fig. 3 a shows RT-PCR analysis of the SBF transgene in the various organs (including kidney, liver, spleen, heart, lung, and brain). The highest levels of SBF transgene expression were obtained in kidney, where levels were demonstrated to be within the linear range (Fig. 3 b). Some expression was also occasionally detectable in spleen and lung, with little or undetectable expression in other organs (Fig. 3 a). This particular organ distribution of transgene expression closely resembles that described previously for SBM mice (35), and it is likely that the transgene is preferentially targeted to the kidney due to common regulatory elements with SBM. Furthermore, to determine more precisely the localization of the c-fos transgene in SBF kidneys, we have carried out in situ hybridization. Adult transgenic SBF kidneys demonstrated high expression of the transgene in tubular epithelial cells throughout the kidney, with particularly intense signal over the medullary collecting tubules (Fig. 4 a). This cellular localization is very similar to that observed for the c-myc transgene. No expression was detected in renal tissue of non transgenic controls (C57BL/6 × CBA)F1 (Fig. 4 b). Furthermore, c-fos expression was also investigated in the SBM mouse kidneys (Fig. 4 c); the absence of signal indicates that c-fos is not induced in the SBM c-myc–dependent cystogenic pathway.

Bottom Line: No renal abnormalities were detected in 13 transgenic lines established, indicating that the PKD phenotype is dependent on functions specific to c-myc.All SBM offspring, irrespective of their p53 genotype, developed PKD with increased renal epithelial apoptotic index.We conclude that the pathogenesis of PKD is c-myc specific and involves a critical imbalance between the opposing processes of cell proliferation and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Institut de Recherches Cliniques de Montréal, Faculté de Médecine de l'Université de Montréal, Montréal, Québec, Canada H2W 1R7.

ABSTRACT
The SBM mouse is a unique transgenic model of polycystic kidney disease (PKD) induced by the dysregulated expression of c-myc in renal tissue. In situ hybridization analysis demonstrated intense signal for the c-myc transgene overlying tubular cystic epithelium in SBM mice. Renal proliferation index in SBM kidneys was 10-fold increased over nontransgenic controls correlating with the presence of epithelial hyperplasia. The specificity of c-myc for the proliferative potential of epithelial cells was demonstrated by substitution of c-myc with the proto-oncogene c-fos or the transforming growth factor (TGF)-alpha within the same construct. No renal abnormalities were detected in 13 transgenic lines established, indicating that the PKD phenotype is dependent on functions specific to c-myc. We also investigated another well characterized function of c-myc, the regulation of apoptosis through pathways involving p53 and members of the bcl-2 family, which induce and inhibit apoptosis, respectively. The SBM kidney tissues, which overexpress c-myc, displayed a markedly elevated (10-100-fold) apoptotic index. However, no significant difference in bcl-2, bax, or p53 expression was observed in SBM kidney compared with controls. Direct proof that the heightened renal cellular apoptosis in PKD is not occurring through p53 was obtained by successive matings between SBM and p53(-/-) mice. All SBM offspring, irrespective of their p53 genotype, developed PKD with increased renal epithelial apoptotic index. In addition, overexpression of both bcl-2 and c-myc in double transgenic mice (SBB+/SBM+) also produced a similar PKD phenotype with a high apoptotic rate, showing that c-myc can bypass bcl-2 in vivo. Thus, the in vivo c-myc apoptotic pathway in SBM mice occurs through a p53- and bcl-2-independent mechanism. We conclude that the pathogenesis of PKD is c-myc specific and involves a critical imbalance between the opposing processes of cell proliferation and apoptosis.

Show MeSH
Related in: MedlinePlus