T Helper 2 Cytokine Signaling in Bone Marrow–Derived Fibroblasts

Fibrosis is a pathologic characteristic in many chronic diseases that result in end stage organ failure, and consequently, it is associated with major morbidity and mortality. The pathogenesis of fibrosis in many of these diseases seems to result from aberrant wound–healing responses to injurious stimuli.1 Dysregulated repair processes can result in excessive accumulation of matrix-producing cells and excessive deposition of extracellular matrix, such as collagen and fibronectin, causing the disruption of normal tissue homeostasis. Accumulation of matrix-producing cells is a critical component of profibrotic injury responses,1 but the precise mechanisms driving this expansion in vivo have not yet been fully identified. Elucidation of these mechanisms will lead to discovery of new therapeutic targets for fibrotic diseases. Among matrix-producing cells, myofibroblasts showing high contractile properties and strongly expressing α–smooth muscle actin (αSMA) are a major component in fibrotic organs.2 Recent studies have shown that myofibroblasts consist of heterogeneous cells, which are derived from different kinds of cells, such as resident fibroblasts, pericytes, epithelial/endothelial mesenchymal transition–derived cells, and bone marrow–derived fibroblasts (termed fibrocytes).3,4 Of these cells, accumulating evidence suggest that bone marrow–derived fibroblasts are significantly involved in the pathogenesis of renal fibrosis through (1) recruitment into kidney and (2) differentiation into myofibroblasts.3 Bone marrow–derived fibroblasts were first identified in the wound chamber model by Bucala et al.5 Bone marrow–derived fibroblasts comprise a minor fraction of the circulating pool of leukocytes (<1%) and share the markers of hematopoietic cells, such as CD45, CD11b, and CD34, as well as mesenchymal cells, including type I collagen (COLI), fibronectin, and platelet–derived growth factor receptor–β.5,6 In addition, bone marrow–derived fibroblasts express chemokine receptors, such as CC chemokine receptor 2 (CCR2), CCR5, CCR7, CXC chemokine receptor 4 (CXCR4), and CXCR6.7–11 Recent studies have shown that chemokine/chemokine receptor systems on bone marrow–derived fibroblasts regulate the recruitment of bone marrow–derived fibroblasts to sites of renal fibrosis.7–15 However, differentiation of bone marrow–derived fibroblasts has been thought to be dependent on various cytokines produced in the local environment. Interestingly, T helper 2 (Th2) cytokines IL-4 and IL-13 have been known to stimulate bone marrow–derived monocytes to differentiate into bone marrow–derived fibroblasts, whereas Th1 cytokines IL-12 and IFN-γ inhibit it.16 However, the precise molecular mechanisms by which bone marrow–derived monocytes are activated to differentiate into bone marrow–derived fibroblasts, resulting in myofibroblast accumulation, are not fully investigated. In this issue of JASN, Yan et al.17 clarified the importance of Janus kinase 3 (JAK3) -signal transducer and activator of transcription 6 (STAT6) signaling in the pathogenesis of renal fibrosis through the activation of bone marrow–derived fibroblasts. Yan et al.17 showed that IL-4 or IL-13 stimulated bone marrow–derived monocytes to express αSMA and extracellular matrix, such as COLI and fibronectin, suggesting that bone marrow–derived monocytes could differentiate into bone marrow–derived fibroblasts (COLI+) and myofibroblasts (COLI+/αSMA+) in response to Th2 cytokines IL-4 or IL-13. JAK3 inhibitor CP690550 (tofacitinib) suppressed IL-4– or IL-13–induced expression of αSMA and extracellular matrix as well as phosphorylation of STAT6 in bone marrow–derived monocytes. STAT6 deficiency also abolished the expression of αSMA and extracellular matrix in bone marrow–derived monocytes. These results suggest the important role of JAK3-STAT6 signaling in Th2 cytokine–induced myeloid fibroblast differentiation into myofibroblasts, which are originally from monocytes. Other than JAK3-STAT6 signaling, TGF-β1 is a well known inducer of fibroblast differentiation into myofibroblasts.18 TGF-β1 is a strong profibrotic cytokine and capable of enhancing the production of COLI, fibronectin, and αSMA in resident fibroblasts. TGF-β1 signaling is mediated by the Smad protein family through types I and II serine/threonine kinase receptors. Among the Smad protein family, Smad3 is a major downstream signaling molecule of TGF-β1 in mediating the pathogenesis of fibrosis. In addition, the blockade of the TGF-β1-Smad3 pathway clearly protects from various organ fibrosis, suggesting that the TGF-β1-Smad3 pathway is one of the fundamental pathways to induce organ fibrosis.19 A recent study has revealed that the TGF-β1-Smad3 pathway induces the transition of monocytes to bone marrow–derived fibroblasts in the course of renal fibrosis.20 Yan et al.17 showed in this study that TGF-β1–induced expression of αSMA, fibronectin, and COLI was not affected by the pretreatment with CP690550 in rat kidney fibroblasts. These results indicate that the effect of CP690550 on the transition of monocytes to bone marrow–derived fibroblasts could be independent of the TGF-β1-Smad3 pathway; however, an experiment using TGF-β1–treated bone marrow–derived monocytes and CP690550 will be required to clarify it. Thus far, JAK3-STAT6 signaling has been reported to suppress the amount of renal collagen accumulation induced by unilateral ureteral obstruction (UUO); however, the mechanism was unclear.21 Yan et al.17 investigated the role of JAK3-STAT6 signaling in UUO–induced renal fibrosis as well. CP690550 administration or STAT6 deficiency significantly attenuated renal fibrosis as estimated by renal amounts of collagen and αSMA. Furthermore, the infiltration of bone marrow–derived fibrobalsts (identified by CD11b+/platelet–derived growth factor receptor–β+ or CD45+/pro-COLI+) increased with the progression of renal fibrosis, whereas the blockade of JAK3-STAT6 signaling using CP690550 or STAT6 knockout mice inhibited bone marrow–derived fibroblast accumulation. These results suggest that JAK3-STAT6 signaling regulates renal fibrosis through the accumulation of bone marrow–derived fibroblasts/myofibroblasts. However, the cells surely involved in JAK3-STAT6 signaling in the development of renal fibrosis are still unclear. Several studies22,23 have shown that JAK3 is also expressed in epithelial cells, including those in the kidney, and JAK3 signaling is thought to be involved in renal phosphate metabolism through 25–hydroxyvitamin D 1α–hydroxylase expression.24 To clarify the importance of bone marrow JAK3-STAT6 signaling in the pathogenesis of renal fibrosis, Yan et al.17 performed bone marrow transplantation of wild-type mice with STAT6+/+ or STAT6−/− bone marrow cells. Wild-type mice transplanted with STAT6−/− bone marrow cells showed fewer bone marrow–derived fibroblasts and a lesser degree of renal collagen content. These data indicate that JAK3-STAT6 signaling in bone marrow cells has a fundamental role in the development of renal fibrosis through accumulation of bone marrow–derived fibroblasts/myofibroblasts. Combining in vivo data with in vitro results, JAK3-STAT6 signaling in bone marrow–derived monocytes might induce the transition to bone marrow–derived fibroblasts and myofibroblasts, thereby contributing to the development of renal fibrosis. Despite the fact that JAK3-STA6 signaling significantly contributes to the pathogenesis of renal fibrosis, which was shown in the study Yan et al.,17 there seems to be several issues to be clarified. First, the specificity of JAK3-STAT6 signaling is not restricted to bone marrow–derived monocytes. JAK3 is a nonreceptor protein tyrosine kinase mainly expressed on hematopoietic cells, and it has been recognized to be essential to lymphocyte differentiation and proliferation as well as dendritic cell maturation.25,26 A recent study has revealed that myeloid dendritic cells are involved in the progression of renal fibrosis through TGF-β production.27 Therefore, the effects of JAK3-STAT6 signaling on dendritic cell maturation should be clarified. Second, Yan et al.17 clearly showed the importance of Th2 cytokines IL-4 and IL-13 through JAK3-STAT6 signaling to induce bone marrow–derived fibroblast/myofibroblast differentiation. However, the mechanisms by which renal expression of IL-4 and IL-13 as well as renal infiltration of Th2 lymphocytes are regulated by JAK3-STAT6 signaling are lacking in this study. In general, Th2 cytokine IL-4 and IL-13 are thought to be produced by Th2 lymphocytes. Recently, Th2 lymphocytes have been reported to contribute to the development of renal fibrosis, because reconstituted mice with Th2 lymphocytes induced renal fibrosis more easily than Th1 reconstituted mice.28 Therefore, the effects of JAK3-STAT6 signaling on the behavior of Th2 lymphocytes also need clarification. Third, Yan et al.17 did not show direct evidence of where bone marrow–derived fibroblasts come from in the course of a renal fibrosis model. There are at least two pathways of how bone marrow–derived fibroblasts accumulate in the fibrotic kidney: (1) the transition of migrated monocytes and (2) the migration of circulating bone marrow–derived fibroblasts. Yan et al.17 mentioned the first possibility regulated by JAK3-STAST6 signaling in this study; however, bone marrow–derived fibroblasts have been reported to be detected in the peripheral blood of patients with various fibrotic diseases, indicating that bone marrow–derived fibroblasts migrate into organs as already differentiated collagen–producing cells.29,30 A recent study has also reported that the depletion of monocytes in the peripheral blood did not inhibit bone marrow–derived fibroblast accumulation in fibrotic kidney after UUO.14 In addition, chemokine/chemokine receptor systems are known to play important roles for renal fibrosis through the migration of circulating bone marrow–derived fibroblasts as described before; however, the effect of JAK3-STAT6 signaling on renal expression of chemokines was not determined in this study. Finally, the interaction between bone marrow–derived fibroblasts and resident cells in kidneys may be involved in the progression of fibrotic events, but it remains to be investigated. More detailed study to address the pathways to accumulate bone marrow–derived fibroblasts would provide better understanding of the relationship of JAK3-STAT6 signaling with bone marrow–derived fibroblasts in the pathogenesis of renal fibrosis. Although there are still unsolved questions regarding the biology of bone marrow–derived fibroblasts, Yan et al.17 have found strong evidence of the importance of JAK3-STAT6 signaling in bone marrow–derived fibroblast activation in renal fibrosis. Additional investigations to clarify the precise mechanisms of matrix–producing cell accumulation, including bone marrow–derived fibroblasts, would open novel therapeutic strategies for combatting renal fibrosis. Disclosures None.