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Renal fibrosis is not reduced by blocking transforming growth factor-β signaling in matrix-producing interstitial cells.

Neelisetty S, Alford C, Reynolds K, Woodbury L, Nlandu-Khodo S, Yang H, Fogo AB, Hao CM, Harris RC, Zent R, Gewin L - Kidney Int. (2015)

Bottom Line: To address this, we deleted the TGF-β type II receptor in matrix-producing interstitial cells using two different inducible Cre models: COL1A2-Cre with a mesenchymal enhancer element and tenascin-Cre that targets medullary interstitial cells, and either the mouse unilateral ureteral obstruction or the aristolochic acid renal injury model.Renal interstitial cells lacking the TGF-β receptor had significantly impaired collagen I production, but, unexpectedly, overall tissue fibrosis was unchanged in the conditional knockouts after renal injury.Thus, abrogating TGF-β signaling in matrix-producing interstitial cells is not sufficient to reduce fibrosis after renal injury.

View Article: PubMed Central - PubMed

Affiliation: Division of Nephrology, Department of Medicine, Vanderbilt Medical Center, Nashville, Tennessee, USA.

ABSTRACT
Transforming growth factor-β (TGF-β) strongly promotes renal tubulointerstitial fibrosis, but the cellular target that mediates its profibrotic actions has not been clearly identified. While in vitro data suggest that TGF-β-induced matrix production is mediated by renal fibroblasts, the role of these cells in TGF-β-dependent tubulointerstitial fibrosis following renal injury is not well defined. To address this, we deleted the TGF-β type II receptor in matrix-producing interstitial cells using two different inducible Cre models: COL1A2-Cre with a mesenchymal enhancer element and tenascin-Cre that targets medullary interstitial cells, and either the mouse unilateral ureteral obstruction or the aristolochic acid renal injury model. Renal interstitial cells lacking the TGF-β receptor had significantly impaired collagen I production, but, unexpectedly, overall tissue fibrosis was unchanged in the conditional knockouts after renal injury. Thus, abrogating TGF-β signaling in matrix-producing interstitial cells is not sufficient to reduce fibrosis after renal injury.

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Expression and localization of COL1A2-Cre/ERT+ (abbreviated COL-Cre) and TenascinC-Cre/ERT+ (abbreviated TNC-Cre) cells. The mT/mG mouse was crossed with COL-Cre and TNC-Cre-containing mice, and Cre activity converts ubiquitous membrane-bound red fluorescence to green (A, B). Distribution of Cre activity for COL-Cre and TNC-Cre is shown in frozen sections of uninjured kidneys (A) as well as in obstructed kidneys (B). Kidneys obstructed for 3 days were converted to single cell suspensions and GFP+ cells were measured. The gating strategy, exclusion of dead cells and doublets, and forward and side scatter characteristics of GFP+ cells are shown in Supplemental Figure 1. An obstructed kidney from mT/mG-containing mouse without Cre is used as negative control (C). The percentage of GFP+ cells as a percentage of viable, single cells is listed for COL-Cre (D) and TNC-Cre (E).
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Figure 1: Expression and localization of COL1A2-Cre/ERT+ (abbreviated COL-Cre) and TenascinC-Cre/ERT+ (abbreviated TNC-Cre) cells. The mT/mG mouse was crossed with COL-Cre and TNC-Cre-containing mice, and Cre activity converts ubiquitous membrane-bound red fluorescence to green (A, B). Distribution of Cre activity for COL-Cre and TNC-Cre is shown in frozen sections of uninjured kidneys (A) as well as in obstructed kidneys (B). Kidneys obstructed for 3 days were converted to single cell suspensions and GFP+ cells were measured. The gating strategy, exclusion of dead cells and doublets, and forward and side scatter characteristics of GFP+ cells are shown in Supplemental Figure 1. An obstructed kidney from mT/mG-containing mouse without Cre is used as negative control (C). The percentage of GFP+ cells as a percentage of viable, single cells is listed for COL-Cre (D) and TNC-Cre (E).

Mentions: We verified that COL-Cre and TNC-Cre activity was present in MPIC by crossing these mice with the mT/mG reporter which has ubiquitous membrane-bound red fluorescence that is converted to green (GFP) by Cre activity.(27) As recombination efficiency is a concern with an inducible system, we used a high dose tamoxifen regimen (see Methods) previously proven to induce efficient recombination in the adult mouse.(28) In the uninjured COL-Cre and TNC-Cre mice, Cre activity localized to peritubular cells primarily in the medullary interstitium (Figure 1A). These GFP+ cells were few in number, consistent with the paucity of collagen I-producing interstitial cells in the healthy kidney. COL-Cre mice had glomerular GFP staining as previously described,(21) but no Cre activity was detectable in the cortex of TNC-Cre mice (Figure 1A).


Renal fibrosis is not reduced by blocking transforming growth factor-β signaling in matrix-producing interstitial cells.

Neelisetty S, Alford C, Reynolds K, Woodbury L, Nlandu-Khodo S, Yang H, Fogo AB, Hao CM, Harris RC, Zent R, Gewin L - Kidney Int. (2015)

Expression and localization of COL1A2-Cre/ERT+ (abbreviated COL-Cre) and TenascinC-Cre/ERT+ (abbreviated TNC-Cre) cells. The mT/mG mouse was crossed with COL-Cre and TNC-Cre-containing mice, and Cre activity converts ubiquitous membrane-bound red fluorescence to green (A, B). Distribution of Cre activity for COL-Cre and TNC-Cre is shown in frozen sections of uninjured kidneys (A) as well as in obstructed kidneys (B). Kidneys obstructed for 3 days were converted to single cell suspensions and GFP+ cells were measured. The gating strategy, exclusion of dead cells and doublets, and forward and side scatter characteristics of GFP+ cells are shown in Supplemental Figure 1. An obstructed kidney from mT/mG-containing mouse without Cre is used as negative control (C). The percentage of GFP+ cells as a percentage of viable, single cells is listed for COL-Cre (D) and TNC-Cre (E).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4556568&req=5

Figure 1: Expression and localization of COL1A2-Cre/ERT+ (abbreviated COL-Cre) and TenascinC-Cre/ERT+ (abbreviated TNC-Cre) cells. The mT/mG mouse was crossed with COL-Cre and TNC-Cre-containing mice, and Cre activity converts ubiquitous membrane-bound red fluorescence to green (A, B). Distribution of Cre activity for COL-Cre and TNC-Cre is shown in frozen sections of uninjured kidneys (A) as well as in obstructed kidneys (B). Kidneys obstructed for 3 days were converted to single cell suspensions and GFP+ cells were measured. The gating strategy, exclusion of dead cells and doublets, and forward and side scatter characteristics of GFP+ cells are shown in Supplemental Figure 1. An obstructed kidney from mT/mG-containing mouse without Cre is used as negative control (C). The percentage of GFP+ cells as a percentage of viable, single cells is listed for COL-Cre (D) and TNC-Cre (E).
Mentions: We verified that COL-Cre and TNC-Cre activity was present in MPIC by crossing these mice with the mT/mG reporter which has ubiquitous membrane-bound red fluorescence that is converted to green (GFP) by Cre activity.(27) As recombination efficiency is a concern with an inducible system, we used a high dose tamoxifen regimen (see Methods) previously proven to induce efficient recombination in the adult mouse.(28) In the uninjured COL-Cre and TNC-Cre mice, Cre activity localized to peritubular cells primarily in the medullary interstitium (Figure 1A). These GFP+ cells were few in number, consistent with the paucity of collagen I-producing interstitial cells in the healthy kidney. COL-Cre mice had glomerular GFP staining as previously described,(21) but no Cre activity was detectable in the cortex of TNC-Cre mice (Figure 1A).

Bottom Line: To address this, we deleted the TGF-β type II receptor in matrix-producing interstitial cells using two different inducible Cre models: COL1A2-Cre with a mesenchymal enhancer element and tenascin-Cre that targets medullary interstitial cells, and either the mouse unilateral ureteral obstruction or the aristolochic acid renal injury model.Renal interstitial cells lacking the TGF-β receptor had significantly impaired collagen I production, but, unexpectedly, overall tissue fibrosis was unchanged in the conditional knockouts after renal injury.Thus, abrogating TGF-β signaling in matrix-producing interstitial cells is not sufficient to reduce fibrosis after renal injury.

View Article: PubMed Central - PubMed

Affiliation: Division of Nephrology, Department of Medicine, Vanderbilt Medical Center, Nashville, Tennessee, USA.

ABSTRACT
Transforming growth factor-β (TGF-β) strongly promotes renal tubulointerstitial fibrosis, but the cellular target that mediates its profibrotic actions has not been clearly identified. While in vitro data suggest that TGF-β-induced matrix production is mediated by renal fibroblasts, the role of these cells in TGF-β-dependent tubulointerstitial fibrosis following renal injury is not well defined. To address this, we deleted the TGF-β type II receptor in matrix-producing interstitial cells using two different inducible Cre models: COL1A2-Cre with a mesenchymal enhancer element and tenascin-Cre that targets medullary interstitial cells, and either the mouse unilateral ureteral obstruction or the aristolochic acid renal injury model. Renal interstitial cells lacking the TGF-β receptor had significantly impaired collagen I production, but, unexpectedly, overall tissue fibrosis was unchanged in the conditional knockouts after renal injury. Thus, abrogating TGF-β signaling in matrix-producing interstitial cells is not sufficient to reduce fibrosis after renal injury.

Show MeSH
Related in: MedlinePlus