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Mice lacking thyroid hormone receptor Beta show enhanced apoptosis and delayed liver commitment for proliferation after partial hepatectomy.

López-Fontal R, Zeini M, Través PG, Gómez-Ferrería M, Aranda A, Sáez GT, Cerdá C, Martín-Sanz P, Hortelano S, Boscá L - PLoS ONE (2010)

Bottom Line: Pharmacologically induced hypothyroidism yielded similar results.Loss of TR activity was associated with enhanced nitrosative stress in the liver remnant, due to an increase in the activity of the nitric oxide synthase (NOS) 2 and 3, caused by a transient decrease in the concentration of asymmetric dimethylarginine (ADMA), a potent NOS inhibitor.Taken together, these results indicate that TRbeta contributes significantly to the rapid initial round of hepatocyte proliferation following PH, and improves the survival of the regenerating liver at later times.

View Article: PubMed Central - PubMed

Affiliation: Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain.

ABSTRACT

Background: The role of thyroid hormones and their receptors (TR) during liver regeneration after partial hepatectomy (PH) was studied using genetic and pharmacologic approaches. Roles in liver regeneration have been suggested for T3, but there is no clear evidence distinguishing the contribution of increased amounts of T3 from the modulation by unoccupied TRs.

Methodology/principal findings: Mice lacking TRalpha1/TRbeta or TRbeta alone fully regenerated liver mass after PH, but showed delayed commitment to the initial round of hepatocyte proliferation and transient but intense apoptosis at 48h post-PH, affecting approximately 30% of the remaining hepatocytes. Pharmacologically induced hypothyroidism yielded similar results. Loss of TR activity was associated with enhanced nitrosative stress in the liver remnant, due to an increase in the activity of the nitric oxide synthase (NOS) 2 and 3, caused by a transient decrease in the concentration of asymmetric dimethylarginine (ADMA), a potent NOS inhibitor. This decrease in the ADMA levels was due to the presence of a higher activity of dimethylarginineaminohydrolase-1 (DDAH-1) in the regenerating liver of animals lacking TRalpha1/TRbeta or TRbeta. DDAH-1 expression and activity was paralleled by the activity of FXR, a transcription factor involved in liver regeneration and up-regulated in the absence of TR.

Conclusions/significance: We report that TRs are not required for liver regeneration; however, hypothyroid mice and TRbeta- or TRalpha1/TRbeta-deficient mice exhibit a delay in the restoration of liver mass, suggesting a specific role for TRbeta in liver regeneration. Altered regenerative responses are related with a delay in the expression of cyclins D1 and E, and the occurrence of liver apoptosis in the absence of activated TRbeta that can be prevented by administration of NOS inhibitors. Taken together, these results indicate that TRbeta contributes significantly to the rapid initial round of hepatocyte proliferation following PH, and improves the survival of the regenerating liver at later times.

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Hydrodynamic transfection of TRβ restores liver regeneration index and impairs caspase activation in regenerating liver of TR KO mice.Before PH, animals (WT, TRβ single KO and TRα1/TRβ double KO mice, referred as ‘KO’) were transfected hydrodynamically with GFP and TRβ expression vectors, or the TRβ-void vector pCX. The expression of GFP and the levels of TRβ (A), the liver mass regeneration index (B) and the levels of procaspase 3 and caspases 3 and 9 (C) were determined 48 h after PH. (D,E) Activities of caspase 9 and caspase 3 using specific peptide substrates were determined in liver extracts obtained 48 h post-PH. Results show the mean ± SD of 5 animals per condition (B,D,E) or a representative section or Western blot out of three (A,C). *P<0.01 vs. the WT condition; #P<0.01 vs. animals of the same genotype transfected with control TRβ-void vector (pCX).
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pone-0008710-g004: Hydrodynamic transfection of TRβ restores liver regeneration index and impairs caspase activation in regenerating liver of TR KO mice.Before PH, animals (WT, TRβ single KO and TRα1/TRβ double KO mice, referred as ‘KO’) were transfected hydrodynamically with GFP and TRβ expression vectors, or the TRβ-void vector pCX. The expression of GFP and the levels of TRβ (A), the liver mass regeneration index (B) and the levels of procaspase 3 and caspases 3 and 9 (C) were determined 48 h after PH. (D,E) Activities of caspase 9 and caspase 3 using specific peptide substrates were determined in liver extracts obtained 48 h post-PH. Results show the mean ± SD of 5 animals per condition (B,D,E) or a representative section or Western blot out of three (A,C). *P<0.01 vs. the WT condition; #P<0.01 vs. animals of the same genotype transfected with control TRβ-void vector (pCX).

Mentions: Mice were injected with a bolus of plasmids encoding TRβ and GFP to allow hydrodynamic transfection of liver in vivo [19]. An average 40–60% hepatocytes stained positive for GFP 24 h post transfection (Fig. 4A). The protein levels of TRβ after hydrodynamic transfection are shown in Fig. 4A. Transfection of TRα1/TRβ double KO or TRβ KO mice with TRβ resulted in a significant increase in liver mass recovery at 48 h after PH (Fig. 4B), suggesting a specific and non-redundant role for TRβ in liver regeneration. Interestingly, overexpression of TRβ in WT animals did not modify the regenerative response (WT condition in Fig. 4B). Ectopic expression of TRβ in the KO mice also markedly decreased the expression and activity of caspase 9 and activity of caspase 3 at 48 h PH, as determined in liver extracts (Fig. 4C,D,E).


Mice lacking thyroid hormone receptor Beta show enhanced apoptosis and delayed liver commitment for proliferation after partial hepatectomy.

López-Fontal R, Zeini M, Través PG, Gómez-Ferrería M, Aranda A, Sáez GT, Cerdá C, Martín-Sanz P, Hortelano S, Boscá L - PLoS ONE (2010)

Hydrodynamic transfection of TRβ restores liver regeneration index and impairs caspase activation in regenerating liver of TR KO mice.Before PH, animals (WT, TRβ single KO and TRα1/TRβ double KO mice, referred as ‘KO’) were transfected hydrodynamically with GFP and TRβ expression vectors, or the TRβ-void vector pCX. The expression of GFP and the levels of TRβ (A), the liver mass regeneration index (B) and the levels of procaspase 3 and caspases 3 and 9 (C) were determined 48 h after PH. (D,E) Activities of caspase 9 and caspase 3 using specific peptide substrates were determined in liver extracts obtained 48 h post-PH. Results show the mean ± SD of 5 animals per condition (B,D,E) or a representative section or Western blot out of three (A,C). *P<0.01 vs. the WT condition; #P<0.01 vs. animals of the same genotype transfected with control TRβ-void vector (pCX).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2806828&req=5

pone-0008710-g004: Hydrodynamic transfection of TRβ restores liver regeneration index and impairs caspase activation in regenerating liver of TR KO mice.Before PH, animals (WT, TRβ single KO and TRα1/TRβ double KO mice, referred as ‘KO’) were transfected hydrodynamically with GFP and TRβ expression vectors, or the TRβ-void vector pCX. The expression of GFP and the levels of TRβ (A), the liver mass regeneration index (B) and the levels of procaspase 3 and caspases 3 and 9 (C) were determined 48 h after PH. (D,E) Activities of caspase 9 and caspase 3 using specific peptide substrates were determined in liver extracts obtained 48 h post-PH. Results show the mean ± SD of 5 animals per condition (B,D,E) or a representative section or Western blot out of three (A,C). *P<0.01 vs. the WT condition; #P<0.01 vs. animals of the same genotype transfected with control TRβ-void vector (pCX).
Mentions: Mice were injected with a bolus of plasmids encoding TRβ and GFP to allow hydrodynamic transfection of liver in vivo [19]. An average 40–60% hepatocytes stained positive for GFP 24 h post transfection (Fig. 4A). The protein levels of TRβ after hydrodynamic transfection are shown in Fig. 4A. Transfection of TRα1/TRβ double KO or TRβ KO mice with TRβ resulted in a significant increase in liver mass recovery at 48 h after PH (Fig. 4B), suggesting a specific and non-redundant role for TRβ in liver regeneration. Interestingly, overexpression of TRβ in WT animals did not modify the regenerative response (WT condition in Fig. 4B). Ectopic expression of TRβ in the KO mice also markedly decreased the expression and activity of caspase 9 and activity of caspase 3 at 48 h PH, as determined in liver extracts (Fig. 4C,D,E).

Bottom Line: Pharmacologically induced hypothyroidism yielded similar results.Loss of TR activity was associated with enhanced nitrosative stress in the liver remnant, due to an increase in the activity of the nitric oxide synthase (NOS) 2 and 3, caused by a transient decrease in the concentration of asymmetric dimethylarginine (ADMA), a potent NOS inhibitor.Taken together, these results indicate that TRbeta contributes significantly to the rapid initial round of hepatocyte proliferation following PH, and improves the survival of the regenerating liver at later times.

View Article: PubMed Central - PubMed

Affiliation: Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain.

ABSTRACT

Background: The role of thyroid hormones and their receptors (TR) during liver regeneration after partial hepatectomy (PH) was studied using genetic and pharmacologic approaches. Roles in liver regeneration have been suggested for T3, but there is no clear evidence distinguishing the contribution of increased amounts of T3 from the modulation by unoccupied TRs.

Methodology/principal findings: Mice lacking TRalpha1/TRbeta or TRbeta alone fully regenerated liver mass after PH, but showed delayed commitment to the initial round of hepatocyte proliferation and transient but intense apoptosis at 48h post-PH, affecting approximately 30% of the remaining hepatocytes. Pharmacologically induced hypothyroidism yielded similar results. Loss of TR activity was associated with enhanced nitrosative stress in the liver remnant, due to an increase in the activity of the nitric oxide synthase (NOS) 2 and 3, caused by a transient decrease in the concentration of asymmetric dimethylarginine (ADMA), a potent NOS inhibitor. This decrease in the ADMA levels was due to the presence of a higher activity of dimethylarginineaminohydrolase-1 (DDAH-1) in the regenerating liver of animals lacking TRalpha1/TRbeta or TRbeta. DDAH-1 expression and activity was paralleled by the activity of FXR, a transcription factor involved in liver regeneration and up-regulated in the absence of TR.

Conclusions/significance: We report that TRs are not required for liver regeneration; however, hypothyroid mice and TRbeta- or TRalpha1/TRbeta-deficient mice exhibit a delay in the restoration of liver mass, suggesting a specific role for TRbeta in liver regeneration. Altered regenerative responses are related with a delay in the expression of cyclins D1 and E, and the occurrence of liver apoptosis in the absence of activated TRbeta that can be prevented by administration of NOS inhibitors. Taken together, these results indicate that TRbeta contributes significantly to the rapid initial round of hepatocyte proliferation following PH, and improves the survival of the regenerating liver at later times.

Show MeSH
Related in: MedlinePlus