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Repair-related activation of hedgehog signaling in stromal cells promotes intrahepatic hypothyroidism.

Bohinc BN, Michelotti G, Xie G, Pang H, Suzuki A, Guy CD, Piercy D, Kruger L, Swiderska-Syn M, Machado M, Pereira T, Zavacki AM, Abdelmalek M, Diehl AM - Endocrinology (2014)

Bottom Line: Repair-related changes in deiodinases were accompanied by reduced hepatic TH content and TH-regulated gene expression.Moreover, the decreases in the free T3 to rT3 and free T4 to rT3 ratios distinguished advanced from mild fibrosis, even in individuals with similar serum levels of TSH and free T4.In conclusion, the Hedgehog-dependent changes in liver stromal cells drive repair-related changes in hepatic deiodinase expression that promote intrahepatic hypothyroidism, thereby limiting exposure to T3, an important factor for cellular differentiation.

View Article: PubMed Central - PubMed

Affiliation: Divisions of Endocrinology, Diabetes, and Metabolism (B.N.B., D.P.) and Gastroenterology (G.M., G.X., A.S., L.K., M.S.-S., M.M., T.P., M.A., A.M.D.) and Departments of Biostatistics and Bioinformatics (H.P.) and Pathology (C.D.G.), Duke University, Durham, North Carolina 27710; and Division of Endocrinology, Diabetes, and Metabolism (A.M.Z.), Brigham and Women's Hospital, Boston, Massachusetts 02115.

ABSTRACT
Thyroid hormone (TH) is important for tissue repair because it regulates cellular differentiation. Intrahepatic TH activity is controlled by both serum TH levels and hepatic deiodinases. TH substrate (T4) is converted into active hormone (T3) by deiodinase 1 (D1) but into inactive hormone (rT3) by deiodinase 3 (D3). Although the relative expressions of D1 and D3 are known to change during liver injury, the cell types and signaling mechanisms involved are unclear. We evaluated the hypothesis that changes in hepatic deiodinases result from repair-related activation of the Hedgehog pathway in stromal cells. We localized deiodinase expression, assessed changes during injury, and determined how targeted manipulation of Hedgehog signaling in stromal cells impacted hepatic deiodinase expression, TH content, and TH action in rodents. Humans with chronic liver disease were also studied. In healthy liver, hepatocytes strongly expressed D1 and stromal cells weakly expressed D3. During injury, hepatocyte expression of D1 decreased, whereas stromal expression of D3 increased, particularly in myofibroblasts. Conditionally disrupting Hedgehog signaling in myofibroblasts normalized deiodinase expression. Repair-related changes in deiodinases were accompanied by reduced hepatic TH content and TH-regulated gene expression. In patients, this was reflected by increased serum rT3. Moreover, the decreases in the free T3 to rT3 and free T4 to rT3 ratios distinguished advanced from mild fibrosis, even in individuals with similar serum levels of TSH and free T4. In conclusion, the Hedgehog-dependent changes in liver stromal cells drive repair-related changes in hepatic deiodinase expression that promote intrahepatic hypothyroidism, thereby limiting exposure to T3, an important factor for cellular differentiation.

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Redistribution of deiodinase expression in injured livers. Immunohistochemistry for DIO1 (A) and DIO3 (B) on day 14 after BDL are shown (magnification, ×10 and ×40). Costaining for D3 (brown) and desmin (green), a stellate cell marker (C), CD68, a macrophage marker (green) (D), and CD31, a marker of liver sinusoidal endothelial cells (green) (E) are shown (magnification, ×100).
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Figure 3: Redistribution of deiodinase expression in injured livers. Immunohistochemistry for DIO1 (A) and DIO3 (B) on day 14 after BDL are shown (magnification, ×10 and ×40). Costaining for D3 (brown) and desmin (green), a stellate cell marker (C), CD68, a macrophage marker (green) (D), and CD31, a marker of liver sinusoidal endothelial cells (green) (E) are shown (magnification, ×100).

Mentions: To determine whether the observed changes in deiodinase expression reflected changes in residual resident liver cells, as opposed to nonspecific effects caused by liver cell loss and/or infiltrating inflammatory cells, additional immunostaining was done (Figure 3). Surprisingly, the types of liver cells producing deiodinases changed dramatically after liver injury. In injured livers, hepatocytes were no longer the dominant D1-expressing cell. Rather, D1 accumulated in ductular type cells (Figure 3A). D3 protein remained in stromal cells (Figure 3B) and mainly colocalized with markers for HSCs (desmin, Figure 3C), KCs (CD68, Figure 3D), and LSECs (CD31, Figure 3E). Desmin, CD68, and CD31 costains were performed by IHC and revealed that, although stellate cells were the major producers of D3 after injury (comprising >60% of D3 expressing cells), approximately 30% of D3-positive cells were CD68 positive and 10% were CD31 positive. In both instances, CD68- and CD31-producing cells increased in a time-dependent manner after BDL injury, with most cells seen by day 7. Hence, the accumulation of these stromal cells likely contributed to the injury-related shift from D1 to D3 that was demonstrated by analysis of whole liver mRNA and protein (Figure 2, A–D). Thus, liver injury recapitulates the fetal-like state of D3 predominance due not only to a loss of D1 in hepatocytes but also because D3 is dramatically increased in the hepatic stromal compartment.


Repair-related activation of hedgehog signaling in stromal cells promotes intrahepatic hypothyroidism.

Bohinc BN, Michelotti G, Xie G, Pang H, Suzuki A, Guy CD, Piercy D, Kruger L, Swiderska-Syn M, Machado M, Pereira T, Zavacki AM, Abdelmalek M, Diehl AM - Endocrinology (2014)

Redistribution of deiodinase expression in injured livers. Immunohistochemistry for DIO1 (A) and DIO3 (B) on day 14 after BDL are shown (magnification, ×10 and ×40). Costaining for D3 (brown) and desmin (green), a stellate cell marker (C), CD68, a macrophage marker (green) (D), and CD31, a marker of liver sinusoidal endothelial cells (green) (E) are shown (magnification, ×100).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256825&req=5

Figure 3: Redistribution of deiodinase expression in injured livers. Immunohistochemistry for DIO1 (A) and DIO3 (B) on day 14 after BDL are shown (magnification, ×10 and ×40). Costaining for D3 (brown) and desmin (green), a stellate cell marker (C), CD68, a macrophage marker (green) (D), and CD31, a marker of liver sinusoidal endothelial cells (green) (E) are shown (magnification, ×100).
Mentions: To determine whether the observed changes in deiodinase expression reflected changes in residual resident liver cells, as opposed to nonspecific effects caused by liver cell loss and/or infiltrating inflammatory cells, additional immunostaining was done (Figure 3). Surprisingly, the types of liver cells producing deiodinases changed dramatically after liver injury. In injured livers, hepatocytes were no longer the dominant D1-expressing cell. Rather, D1 accumulated in ductular type cells (Figure 3A). D3 protein remained in stromal cells (Figure 3B) and mainly colocalized with markers for HSCs (desmin, Figure 3C), KCs (CD68, Figure 3D), and LSECs (CD31, Figure 3E). Desmin, CD68, and CD31 costains were performed by IHC and revealed that, although stellate cells were the major producers of D3 after injury (comprising >60% of D3 expressing cells), approximately 30% of D3-positive cells were CD68 positive and 10% were CD31 positive. In both instances, CD68- and CD31-producing cells increased in a time-dependent manner after BDL injury, with most cells seen by day 7. Hence, the accumulation of these stromal cells likely contributed to the injury-related shift from D1 to D3 that was demonstrated by analysis of whole liver mRNA and protein (Figure 2, A–D). Thus, liver injury recapitulates the fetal-like state of D3 predominance due not only to a loss of D1 in hepatocytes but also because D3 is dramatically increased in the hepatic stromal compartment.

Bottom Line: Repair-related changes in deiodinases were accompanied by reduced hepatic TH content and TH-regulated gene expression.Moreover, the decreases in the free T3 to rT3 and free T4 to rT3 ratios distinguished advanced from mild fibrosis, even in individuals with similar serum levels of TSH and free T4.In conclusion, the Hedgehog-dependent changes in liver stromal cells drive repair-related changes in hepatic deiodinase expression that promote intrahepatic hypothyroidism, thereby limiting exposure to T3, an important factor for cellular differentiation.

View Article: PubMed Central - PubMed

Affiliation: Divisions of Endocrinology, Diabetes, and Metabolism (B.N.B., D.P.) and Gastroenterology (G.M., G.X., A.S., L.K., M.S.-S., M.M., T.P., M.A., A.M.D.) and Departments of Biostatistics and Bioinformatics (H.P.) and Pathology (C.D.G.), Duke University, Durham, North Carolina 27710; and Division of Endocrinology, Diabetes, and Metabolism (A.M.Z.), Brigham and Women's Hospital, Boston, Massachusetts 02115.

ABSTRACT
Thyroid hormone (TH) is important for tissue repair because it regulates cellular differentiation. Intrahepatic TH activity is controlled by both serum TH levels and hepatic deiodinases. TH substrate (T4) is converted into active hormone (T3) by deiodinase 1 (D1) but into inactive hormone (rT3) by deiodinase 3 (D3). Although the relative expressions of D1 and D3 are known to change during liver injury, the cell types and signaling mechanisms involved are unclear. We evaluated the hypothesis that changes in hepatic deiodinases result from repair-related activation of the Hedgehog pathway in stromal cells. We localized deiodinase expression, assessed changes during injury, and determined how targeted manipulation of Hedgehog signaling in stromal cells impacted hepatic deiodinase expression, TH content, and TH action in rodents. Humans with chronic liver disease were also studied. In healthy liver, hepatocytes strongly expressed D1 and stromal cells weakly expressed D3. During injury, hepatocyte expression of D1 decreased, whereas stromal expression of D3 increased, particularly in myofibroblasts. Conditionally disrupting Hedgehog signaling in myofibroblasts normalized deiodinase expression. Repair-related changes in deiodinases were accompanied by reduced hepatic TH content and TH-regulated gene expression. In patients, this was reflected by increased serum rT3. Moreover, the decreases in the free T3 to rT3 and free T4 to rT3 ratios distinguished advanced from mild fibrosis, even in individuals with similar serum levels of TSH and free T4. In conclusion, the Hedgehog-dependent changes in liver stromal cells drive repair-related changes in hepatic deiodinase expression that promote intrahepatic hypothyroidism, thereby limiting exposure to T3, an important factor for cellular differentiation.

Show MeSH
Related in: MedlinePlus