Transforming growth factor ß-1 stimulates profibrotic epithelial signaling to activate pericyte-myofibroblast transition in obstructive kidney fibrosis.

Pericytes have been identified as the major source of precursors of scar-producing myofibroblasts during kidney fibrosis. The underlying mechanisms triggering pericyte-myofibroblast transition are poorly understood. Transforming growth factor ß-1 (TGF-ß1) is well recognized as a pluripotent cytokine that drives organ fibrosis. We investigated the role of TGF-ß1 in inducing profibrotic signaling from epithelial cells to activate pericyte-myofibroblast transition. Increased expression of TGF-ß1 was detected predominantly in injured epithelium after unilateral ureteral obstruction, whereas downstream signaling from the TGF-ß1 receptor increased in both injured epithelium and pericytes. In mice with ureteral obstruction that were treated with the pan anti-TGF-ß antibody (1D11) or TGF-ß receptor type I inhibitor (SB431542), kidney pericyte-myofibroblast transition was blunted. The consequence was marked attenuation of fibrosis. In addition, epithelial cell cycle G2/M arrest and production of profibrotic cytokines were both attenuated. Although TGF-ß1 alone did not trigger pericyte proliferation in vitro, it robustly induced α smooth muscle actin (α-SMA). In cultured kidney epithelial cells, TGF-ß1 stimulated G2/M arrest and production of profibrotic cytokines that had the capacity to stimulate proliferation and transition of pericytes to myofibroblasts. In conclusion, this study identified a novel link between injured epithelium and pericyte-myofibroblast transition through TGF-ß1 during kidney fibrosis.

Role of lung pericytes and resident fibroblasts in the pathogenesis of pulmonary fibrosis.

RATIONALE: The origin of cells that make pathologic fibrillar collagen matrix in lung disease has been controversial. Recent studies suggest mesenchymal cells may contribute directly to fibrosis. OBJECTIVES: To characterize discrete populations of mesenchymal cells in the normal mouse lung and to map their fate after bleomycin-induced lung injury. METHODS: We mapped the fate of Foxd1-expressing embryonic progenitors and their progeny during lung development, adult homeostasis, and after fibrosing injury in Foxd1-Cre; Rs26-tdTomato-R mice. We studied collagen-I(α)1-producing cells in normal and diseased lungs using Coll-GFP(Tg) mice. MEASUREMENTS AND MAIN RESULTS: Foxd1-expressing embryonic progenitors enter lung buds before 13.5 days post-conception, expand, and form an extensive lineage of mesenchymal cells that have characteristics of pericytes. A collagen-I(α)1-expressing mesenchymal population of distinct lineage is also found in adult lung, with features of a resident fibroblast. In contrast to resident fibroblasts, Foxd1 progenitor-derived pericytes are enriched in transcripts for innate immunity, vascular development, WNT signaling pathway, and cell migration. Foxd1 progenitor-derived pericytes expand after bleomycin lung injury, and activate expression of collagen-I(α)1 and the myofibroblast marker αSMA in fibrotic foci. In addition, our studies suggest a distinct lineage of collagen-I(α)1-expressing resident fibroblasts that also expands after lung injury is a second major source of myofibroblasts. CONCLUSIONS: We conclude that the lung contains an extensive population of Foxd1 progenitor-derived pericytes that are an important lung myofibroblast precursor population.

In vitro analysis of transport and metabolism of 4'-thiothymidine in human tumor cells.

INTRODUCTION: The use of thymidine (TdR) and thymidine analogs such as 3'-fluoro-3'-deoxythymidine (FLT) as positron emission tomography (PET)-based proliferation markers can provide information on tumor response to treatment. Studies on another TdR analog, 4'-thiothymidine (4DST), suggest that it might be a better PET-based proliferation tracer than either TdR or FLT. 4DST is resistant to the catabolism that complicates analysis of TdR in PET studies, but unlike FLT, 4DST is incorporated into DNA. METHODS: To further evaluate 4DST, the kinetics of 4DST transport and metabolism were determined and compared to FLT and TdR. Transport and metabolism of FLT, TdR and 4DST were examined in the human adenocarcinoma cell line A549 under exponential-growth conditions. Single cell suspensions were incubated in buffer supplemented with radiolabeled tracer in the presence or absence of nitrobenzylmercaptopurine ribonucleoside (NBMPR), an inhibitor of equilibrative nucleoside transporters (ENT). Kinetics of tracer uptake was determined in whole cells and tracer metabolism measured by high performance liquid chromatography of cell lysates. RESULTS: TdR and 4DST were qualitatively similar in terms of ENT-dependent transport, shapes of uptake curves, and relative levels of DNA incorporation. FLT did not incorporate into DNA, showed a significant temperature effect for uptake, and its transport had a significant NBMPR-resistant component. Overall 4DST metabolism was significantly slower than either TdR or FLT. CONCLUSIONS: 4DST provides a good alternative for TdR in PET and has advantages over FLT in proliferation measurement. However, slow 4DST metabolism and the short half-life of the (11)C label might limit widespread use in PET.

MicroRNA-21 promotes fibrosis of the kidney by silencing metabolic pathways.

Scarring of the kidney is a major public health concern, directly promoting loss of kidney function. To understand the role of microRNA (miRNA) in the progression of kidney scarring in response to injury, we investigated changes in miRNA expression in two kidney fibrosis models and identified 24 commonly up-regulated miRNAs. Among them, miR-21 was highly elevated in both animal models and in human transplanted kidneys with nephropathy. Deletion of miR-21 in mice resulted in no overt abnormality. However, miR-21(-/-) mice suffered far less interstitial fibrosis in response to kidney injury, a phenotype duplicated in wild-type mice treated with anti-miR-21 oligonucleotides. Global derepression of miR-21 target mRNAs was readily detectable in miR-21(-/-) kidneys after injury. Analysis of gene expression profiles up-regulated in the absence of miR-21 identified groups of genes involved in metabolic pathways, including the lipid metabolism pathway regulated by peroxisome proliferator-activated receptor-α (Pparα), a direct miR-21 target. Overexpression of Pparα prevented ureteral obstruction-induced injury and fibrosis. Pparα deficiency abrogated the antifibrotic effect of anti-miR-21 oligonucleotides. miR-21 also regulated the redox metabolic pathway. The mitochondrial inhibitor of reactive oxygen species generation Mpv17l was repressed by miR-21, correlating closely with enhanced oxidative kidney damage. These studies demonstrate that miR-21 contributes to fibrogenesis and epithelial injury in the kidney in two mouse models and is a candidate target for antifibrotic therapies.