How Does TGF-� Mediate Tubulointerstitial Fibrosis?

Leslie Gewin, MD,*,† and Roy Zent, MD, PhD†,‡,§,�

Summary: Tubulointerstitial fibrosis mediates the development of end-stage renal disease from renal injuries ofall etiologies and is considered an important predictor of renal survival. Transforming growth factor-� (TGF-�) isone of the most important growth factors that promotes tubulointerstitial fibrosis, but the mechanisms whereby thisoccurs are not well defined. This is because TGF-� has pleiotropic effects that depend on the target cell type. Thisreview discusses how TGF-� signaling in each of the relevant cell types (eg, tubular epithelium, fibroblasts) maycontribute to tubulointerstitial fibrosis progression and suggests ways in which future research can improve ourunderstanding of TGF-�–mediated tubulointerstitial fibrosis.Semin Nephrol 32:228-235 Published by Elsevier Inc.Keywords: TGF-�, tubulointerstitial fibrosis, extracellular matrix, chronic kidney disease

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Renal fibrosis, characterized by the accumulationof extracellular matrix (ECM), is the final com-mon pathway whereby all renal injuries lead to

end-stage renal disease. Although fibrosis in the kidneycan occur as either tubulointerstitial fibrosis or glomeru-losclerosis, injury to both the tubulointerstitial and glo-merular compartments ultimately results in tubulointer-stitial fibrosis. Exactly how glomerular injury causestubulointerstitial fibrosis is poorly understood, but it iswell established that the best predictor of renal survivalfrom kidney disease of any etiology is tubulointerstitialfibrosis. Fibrosis in the renal tubulointerstitium, similarto fibrosis in all organs, results from increased ECMsynthesis, reduced degradation, or a combination of both.Dysregulation of this equilibrium can occur during tissuerepair as a result of persistent inflammation, increasedreactive oxygen species (ROS) production, changes inprotease levels, as well as altered cell-ECM interactions.These cellular events are regulated at the molecular level

*Department of Research, Veterans Affairs Hospital, Tennessee ValleyHealthcare System, Nashville, TN.

†Division of Nephrology, Department of Medicine, Vanderbilt MedicalCenter, Nashville, TN.

‡Department of Medicine, Veterans Affairs Hospital, Tennessee ValleyHealthcare System, Nashville, TN.

§Cell and Developmental Biology, Vanderbilt Medical Center, Nash-ville, TN.

�Cancer Biology, Vanderbilt Medical Center, Nashville, TN.Financial support: Supported in part by a Veterans Affairs Career

Development Award, Biomedical Laboratory Research and Devel-opment (L.G.); the Vanderbilt Physician Scientist DevelopmentAward (L.G.); National Institutes of Health grants DK065123,DK075594, DK65123, and DK083187 (all to R.Z.); an AHA estab-lished investigator award (R.Z.); a Merit Award from the Depart-ment of Veterans Affairs (R.Z.); and the George O’Brien CenterGrant (R.Z.).

Conflict of interest statement: none.Address reprint requests to Leslie Gewin, Vanderbilt University Med-

ical Center, Room C3210 MCN, 1161 21st Ave. S., Nashville, TN37232-2372. E-mail: leslie.gewin@vanderbilt.edu

0270-9295/ - see front matterPublished by Elsevier Inc.

http://dx.doi.org/10.1016/j.semnephrol.2012.04.001

228

y numerous proteins including growth factors. One ofhe most important growth factors that plays a role inenal fibrosis is transforming growth factor-� (TGF-�).

TGF-� has 3 mammalian isoforms (TGF-�1, -�2, and-�3) that are a part of the TGF-� superfamily, which alsoncludes the bone morphogenic proteins and activins.GF-� is secreted in a latent form as a result of nonco-

valent binding to the latency-associated peptide (LAP),and is anchored to the ECM by latent TGF-� bindingroteins (Fig. 1).1 To become active, TGF-� is releasedrom the LAP by numerous mechanisms, including pro-eolytic cleavage (eg, matrix metalloproteinases [MMPs],lasmin), ROS, heat, or through conformational changes induced by thrombospondin and certain integrins.2

Once activated, TGF-� ligands bind to the TGF-� type IIreceptor (T�RII), which phosphorylates the TGF-� typeI receptor, leading to activation of multiple intracellularsignaling pathways.3,4 Canonical TGF-� signaling is me-iated by the Smads, but other downstream proteins (eg,itogen-activated protein kinase, TGF-beta activated ki-

ase) can modulate Smad signaling and exert Smad-ndependent effects.5,6 These complex signaling path-

ways allow TGF-� to regulate multiple cellular eventsnvolved in tissue development, immune function, woundepair, and cancer biology.

A large body of evidence suggests that TGF-� plays aritical role in the pathogenesis of tubulointerstitial fibro-is. The best direct evidence for these effects is thatverexpression of TGF-�1 by renal tubular epithelialells resulted in tubulointerstitial fibrosis in the absencef any injury,7 and, conversely, a blocking antibody toGF-� ameliorated interstitial matrix accumulation afternilateral ureteral obstruction (UUO), a model of severeubulointerstitial fibrosis.8 Despite this compelling evi-ence that TGF-� promotes tubulointerstitial fibrosis, the

mechanism whereby it does so remains unclear. This ispartly owing to the complexity of tubulointerstitial fibro-sis progression, which involves many different cell types:interstitial fibroblasts produce matrix components, in-

flammatory cells generate profibrotic growth factors, and

Seminars in Nephrology, Vol 32, No 3, May 2012, pp 228-235

TGF-� and tubulointerstitial fibrosis 229

Figure 1. The effects of TGF-� vary depending on target cell type. TGF-� ligands are secreted as mature homodimers noncovalently boundto the LAP. This latent complex is anchored to the ECM by LTBP, and the mature TGF-� homodimer must be separated from the LAP tobecome active and bind to the TGF-� type II receptor. TGF-� has pleiotropic effects that vary depending on the target cell type. Shown arethe different cellular responses to TGF-� by various cell types involved in tubulointerstitial fibrosis (tubular epithelium, renal fibroblast,macrophage, and endothelium). Many of these cellular responses were determined by in vitro studies, and validation of these effects in vivo

has yet to be confirmed. Abbreviations: CTGF, connective tissue growth factor; LTBP, latent TGF-� binding protein; PAI-1, plasminogenactivator inhibitor-1.

Figure 2. Proposed cellular interactions that lead to TGF-�–mediated tubulointerstitial fibrosis. Various forms of injury (eg, mechanical stretch,hypoxia) target the renal tubular epithelium, leading to production of inflammatory cytokines such as monocyte chemoattractant protein-1(MCP-1), which recruits macrophages. The infiltrating macrophages are potent sources of TGF-� that can signal on neighboring epithelial cellsor renal fibroblasts. TGF-� from either macrophages or injured tubular epithelium stimulates fibroblasts to produce matrix components such ascollagen I and fibronectin. The increased TGF-� production by injured epithelium can signal in an autocrine fashion, leading to further TGF-�

production, de-differentiation, and possibly increased collagen IV production. Tubular injury also may increase integrin �v�6 expression andactivation of latent TGF-�.

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230 L. Gewin and R. Zent

tubular epithelial cells are often the initial target of injury(Fig. 2). The lack of a clearly defined mechanismwhereby TGF-� promotes fibrosis is due to the fact thatTGF-� receptors are expressed ubiquitously and TGF-�has pleiotropic effects that are cell type–specific and varyin different forms of injury. In addition to these factors,the role of TGF-� in renal injury has been studied pri-marily by modulating TGF-� activity through systemicapproaches. This strategy only reveals the aggregate re-sponse of the kidney to TGF-� and does not elucidate themechanism whereby TGF-� increases tubulointerstitialfibrosis. In this review, we outline what is known aboutthe specific effects of TGF-� signaling on the response toinjury by each cell type involved in tubulointerstitialfibrosis (Fig. 1) to define some of the important questionsthat future research should address.

TGF-� SIGNALING AND RENAL FIBROBLASTS

The fibroblast, considered the primary cell responsiblefor matrix synthesis, is thought to be the main mediatorof the profibrotic effects of TGF-�. This perception isbased on several in vitro studies showing increased fi-broblast proliferation and matrix production in responseto TGF-�. Specifically, TGF-� induces a morphologictransformation of human subcutaneous fibroblasts intomyofibroblasts, potent matrix-producing cells, by in-creasing expression of �-smooth muscle actin (�-SMA).9

TGF-� also stimulates fibroblast transcription of collagenI and fibronectin, important components of tubulointer-stitial fibrosis.10,11 In addition, TGF-� signaling in fibro-blasts promotes matrix accumulation by suppressing itsdegradation through down-regulation of certain MMPsand increased expression of tissue inhibitor of MMP.12,13

There are several reasons why validation of these invitro findings with in vivo studies is critical. First, not allfibroblasts respond equally to TGF-�. Thus, studies ondermal fibroblasts may not necessarily predict cellularbehavior of renal fibroblasts. Furthermore, althoughmany early in vitro studies were performed on renalfibroblasts (NRK-49F cell line), heterogeneic responsesto TGF-� by subclones of NRK-49F cells have beenreported.14 Perhaps this is because NRK-49F cells are amixture of cortical and medullary fibroblasts that havemorphologic and perhaps functional differences. Second,the response of cells to TGF-� is determined by the dose;for example, TGF-� can stimulate matrix production inrenal cortical fibroblasts at one dose (0.2 ng/mL) butsuppress collagen secretion at higher amounts (1 ng/mL).15 Whether the concentration of active TGF-� in theinjured renal tubulointerstitium results in fibroblast sup-pression or stimulation of ECM is unclear and an impor-tant reason why in vivo studies are necessary. Finally, theactions of TGF-� are not just cell-specific but also de-pendent on the surrounding microenvironment and canbe modulated by other growth factors and cytokines.16

Because the cell culture milieu is quite different from that d

f the injured kidney, the fibroblast’s response to TGF-�in vitro may poorly predict its actions after injury in vivo.

The intracellular signaling pathways whereby TGF-�mediates its profibrotic effects on renal fibroblasts arealso not well delineated. Both p21-activated kinase-2 andc-Abl kinases have been shown to mediate the prolifer-ative effects of TGF-� on renal fibroblasts but do nothave this effect on other renal cell types such as mesan-gial cells.17 The signaling pathways whereby TGF-�stimulates matrix production have been investigated inmesangial cells,18-20 but have not yet been defined inenal fibroblasts. This is an important area of study be-ause one potentially could identify fibroblast-specificGF-� signaling pathways that could be safer and more

specific pharmaceutical targets.

TGF-� SIGNALING ANDENAL TUBULAR EPITHELIAL CELLS

The renal tubular epithelium is often the cellular target ofinjury, and there is also increasing literature supportingits role as an effector of tubulointerstitial fibrosis. In-creased expression of TGF-� and its receptors have beenshown in the damaged epithelium in multiple models ofglomerular and renal tubular injury.21-23 These includehe following: (1) the UUO injury model, the classicodent model of renal tubulointerstitial fibrosis in whichhe collecting duct epithelium is directly injured by thencreased intraluminal pressure from obstruction; (2) di-betic models of injury resulting in proteinuria thatargets the proximal tubule epithelium as an importantxtraglomerular site of injury; (3) models of acuteidney injury using toxins or ischemia/reperfusion thatarget the proximal tubule because of its high meta-olic requirements and unique vascular supply. Inhese settings of injured renal tubules, epithelialGF-� signaling is thought to promote tubulointersti-

tial fibrosis by affecting tubular cell integrity andincreasing ECM production by both tubular epithelialcells and neighboring fibroblasts.

Increased Epithelial Cell Death

TGF-� signaling can impair renal tubule cell integrityleading to tubular atrophy, a hallmark of tubulointerstitialfibrosis, by inducing either epithelial apoptosis or au-tophagy. TGF-�, a strong promoter of epithelial apopto-is, has been shown to induce renal tubule cell apoptosisn vitro.24 Furthermore, the protective effects of a TGF-

�–blocking antibody (clone 1D11) on the response toUUO partially were attributed to reduced tubular apopto-sis in vivo.8 Recently, TGF-� was shown to cause tubulardestruction because overexpression of TGF-�1 in renalubules in vivo led to autophagy and tubular degenera-ion.7 One of the problems with this elegant model is thathe levels of TGF-�1 in these mice were very high (�200g/mL), and likely were much higher than those pro-

uced by most chronic disease states.

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TGF-� and tubulointerstitial fibrosis 231

In tubulointerstitial fibrosis, there is concurrent tubularatrophy that raises the question: does epithelial apoptosiscause increased matrix accumulation? In the pulmonaryliterature, the increased apoptosis induced in the lungs ofTGF-�1–overexpressing mice was mediated by the earlygrowth response gene-1, and inhibiting early growth re-sponse gene-1 not only rescued the mice from TGF-�–dependent apoptosis but also reduced fibrosis, establish-ing a link between the two.25 A similar connectionbetween TGF-�–dependent apoptosis and fibrosis hasnot yet been proven in the kidney. A putative mechanismwhereby apoptosis leads to fibrosis is through the phago-cytosis of apoptotic cells by macrophages.26 This stimu-lates the production of TGF-�, which may induce matrixproduction by surrounding fibroblasts. The tubular atro-phy induced by epithelial TGF-� signaling contributes tothe injury and impaired renal function observed in tubu-lointerstitial fibrosis even if a causal relationship withECM production has not yet been established.

Altered Production of ECM

TGF-� facilitates the de-differentiation of injured renalepithelial cells into more mesenchymal-like cells, and itis possible that this process results in increased ECMsynthesis. During this phenotypic change, tubular cellslose epithelial markers such as E cadherin and zonulaoccludens-1, express mesenchymal-like proteins such as�-SMA and vimentin, and undergo cytoskeletal altera-tions resulting in dissolution of tight junctions and adap-tation of a more migratory structure.27 Numerous mech-anisms are responsible for this change in phenotype.TGF-� mediated the loss of E cadherin in renal epithelialcells in vitro through induction of integrin-linked kinasein a Smad-dependent fashion.28 TGF-� also induced theexpression of �-SMA in renal epithelial cells throughpathways involving �-catenin as well as the guanosinetriphosphatase Rho.29,30 TGF-� can induce all of thesemorphologic changes in renal tubular epithelial cells aswell as increase the expression of collagens I and IV, andfibronectin in vitro, suggesting that the alteration in phe-notype may cause the increased matrix production.31,32

Alternatively, it is possible that these are two independenteffects induced by TGF-� and there is no causal relation-ship. TGF-� also may stimulate the de-differentiation ofinjured epithelial cells in vivo because Smad3 null epithelialcells had better preservation of epithelial markers afterUUO.33 Whether these de-differentiated tubular epithelialcells in vivo actually migrate through the tubular basementmembrane and populate the interstitium as matrix-produc-ing fibroblasts is controversial.34,35 Even if these mesenchy-mal-like epithelial cells never become bona fide interstitialfibroblasts, they still may contribute to peritubular fibrosis.Further research is necessary to determine how much auto-crine ECM production occurs in response to renal tubularTGF-� signaling in vivo.

TGF-� signaling in injured tubular epithelium also

may promote tubulointerstitial fibrosis by increasing

ECM production of neighboring fibroblasts. TGF-� au-oinduces its own production in a feedforward processhat depends on the coordinated actions of Smad3, extra-ellular signal-regulated kinase (ERK) and p38 in renalroximal tubules.36,37 Therefore, injury to tubules resultsn increased epithelial synthesis of TGF-�, which can

signal on nearby fibroblasts. Such paracrine signalinglikely accounts for the proliferating fibroblasts and inter-stitial matrix accumulation observed in mice overex-pressing TGF-�1 in renal tubules.7

Epithelial TGF-� signaling also can increase the ex-pression of other profibrotic growth factors such as con-nective tissue growth factor, which has been shown tostimulate matrix production by increasing transcriptionof collagen I in both renal proximal tubules and fibro-blasts in vitro. Plasminogen activator inhibitor-1 is an-other protein downstream of TGF-� with clear profi-rotic actions in the kidney.38,39 Also, in nonrenal cells,GF-� signaling has been shown to modify the actions ofpidermal growth factor–receptor ligands, hepatocyterowth factor, and insulin-like growth factor.16,40,41 Fu-

ture research is required to define how TGF-� signalingalters these growth factors’ actions in the injured renaltubulointerstitium.

Epithelial Activation of TGF-�

Renal tubular epithelium also plays an important role inTGF-� activation. Abundant TGF-� is sequestered in itsLAP-bound latent form anchored to the matrix, so acti-vation is a critical rate-limiting step in controlling TGF-�signaling.1 The �v integrins are important in vivo acti-vators of TGF-�.42 In particular, integrin �v�6 has beenshown to mediate TGF-�–dependent tubulointerstitialfibrosis after UUO injury.43 The expression of integrin�v�6 is restricted to epithelial cells where it binds theLAP of TGF-�1 and TGF-�3, which are tethered to the

CM. For integrin �v�6 to induce the conformationalhange necessary to activate TGF-�, there must be anntact actin cytoskeleton as well as integrin activation,resumably causing traction on the matrix via binding toGF-�/LAP.44,45 Furthermore, latent TGF-� must be an-hored to the ECM through one of the latent TGF-�

binding proteins for integrin �v�6-mediated activation toccur.46 As such, integrin �v�6’s interaction with latent

TGF-� that is anchored to the ECM may be one way forpithelial cells to transmit mechanical stress and/or cy-oskeletal changes to the surrounding matrix.47 The cel-

lular stress from UUO-induced stretch imposed on col-lecting duct epithelial cells may activate integrin �v�6,eading to TGF-� activation. This mechanical strain fromncreased intraluminal pressure may not be limited tobstructive nephropathy. Reduction in nephron number,s occurs in the 5/6 nephrectomy model or chronic kid-ey disease, has been shown to increase single-nephronlomerular filtration rate, and, consequently, induce fluidhear stress to the tubules.48 This intratubular shear stress

can activate TGF-� independent of proteinuria or hyper-

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tension.49 Therefore, changes in intratubular fluid pres-sure caused by declining renal function may promoteTGF-�–mediated tubulointerstitial fibrosis through ef-fects on tubular epithelium.50

As discussed earlier, injured renal tubular epitheliumhas been shown to contribute to TGF-�–dependent tu-bulointerstitial fibrosis by inducing apoptosis/autophagy,increasing TGF-� and other growth factor production,and through augmented TGF-� activation. Paradoxically,when our group injured mice lacking TGF-� signaling inthe collecting duct tubular epithelium, the conditionalknockout mice sustained greater fibrosis and injury.51

These results suggest that, although excessive TGF-�signaling is deleterious, some TGF-� signaling is neces-sary for maintenance of the normal epithelial structure.Consistent with this, reduced epithelial TGF-� activity inlung epithelia resulted in emphysematous changes,52 anddominant-negative T�RII expression in the exocrine pan-creas perturbed epithelial homeostasis.53

TGF-� SIGNALING AND INFLAMMATORY CELLS

Inflammation plays an important role in the pathophysi-ology of tubulointerstitial fibrosis.54 Most renal injurieshave some component of an inflammatory infiltrate com-posed of T cells, macrophages, and neutrophils, withvarying distribution based on the nature of injury. Inchronic renal injury, the macrophage is the predominantimmune cell and although it exerts beneficial effects byphagocytosis of cellular debris and apoptotic cells, mac-rophages also are associated with worsened tubulointer-stitial injury.55 The detrimental actions of macrophagesmay be mediated, in part, by augmenting TGF-� activity.Macrophages directly increase TGF-� activity by bothproducing large amounts of TGF-� as well as generatingROS, which activate latent TGF-�. Also, macrophages

Table 1. TGF-� Modulates the Immune Response

TGF-� as Proinflammatory

TGF-�1 increased inflammatory cytokine (MCP-1, IL-8) production by proximal tubule cells invitro

Qi et al61

Smad3�/� mice had attenuated macrophageinfiltration after UUO

Sato et al3

Smad3�/� mice had less interleukin-6 after acutekidney injury

Nath et al7

ALK5 inhibitor targeted to proximal tubule reducedinflammation after UUO

Prakash et

NOTE. TGF-� has been described as a proinflammatory and anti-icell and microenvironment. This table shows seemingly contrahave immunosuppressive effects on immune cells but proinflamby Cao et al,74 in which only macrophages ex vivo were treateinflammation, likely as a result of altered signaling in inflammatoto how their models differentially affected the activity of SmaOverexpressing latent TGF-�1 as well as deleting Smad4 both athe inflammatory effects.74,77

produce inflammatory cytokines such as IL-6, which

lters TGF-�–receptor trafficking, leading to heightenedsignaling.56

Not only do inflammatory cells modulate TGF-� ac-ivity, but TGF-� affects immune responses as evidenced

by the early death of TGF-�1 null mice from overwhelm-ing inflammation.57 Although this finding implies that

GF-� has an immunosuppressive effect, the preciseeffect of TGF-� on immune responses is complicated andlikely depends on the cellular milieu (Table 1). Theeffects of TGF-� on monocytes/macrophages vary, de-ending on cellular differentiation: this growth factortimulates monocyte migration and adhesion but sup-resses inflammatory cytokine production by tissue mac-ophages (Fig. 3).58 The effect of TGF-� signaling on

inflammation also may vary, depending on the cell typein which signaling is altered: TGF-� signaling in mac-rophages may decrease the production of proinflamma-tory growth factors and cytokines whereas TGF-� sig-naling within injured epithelium likely heightens theinflammatory response.59,60 For example, Smad3-defi-cient mice have attenuated macrophage infiltration afterinjury by UUO,33 suggesting that TGF-� may promoteinflammation in injured kidneys. One possible explana-tion for this increased macrophage infiltration is thatTGF-� can stimulate monocyte chemoattractant pro-ein-1 expression in renal proximal tubules.61 Thus, it is

likely that TGF-� signaling in injured tubular epitheliumresults in increased macrophage infiltration that, if pro-longed, can lead to synthesis of growth factors such asTGF-�, which stimulates neighboring fibroblasts to pro-duce matrix components (Figs. 2 and 3).

TGF-� AND THEMICROVASCULATURE

Capillary degeneration is linked integrally to tubulointer-stitial fibrosis, and endothelial cell drop-out may play a

TGF-� as Anti-Inflammatory

TGF-�1–treated macrophages attenuatedinflammation after adriamycin nephrosis

Cao et al74

Overexpression of latent TGF-�1prevented inflammation in UUO model

Wang et al77

Deletion of Smad4 in renal tubulesenhanced renal inflammation after UUO

Meng et al79

Deletion of T�RII caused increasedinflammation in kidney and other organs

Leveen et al81

matory growth factor, and its specific effect depends on the targety effects of TGF-� in modulating renal inflammation. TGF-� canry effects on renal parenchyma.73 This is consistent with the study

TGF-�. Also, the global deletion of T�RII produced multi-organlls. The conflicting reports of the other studies listed may be owinga known modulator of inflammation and nuclear factor-�B.75,76

red to have dramatic effects on Smad7 activity, which may explain

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TGF-� and tubulointerstitial fibrosis 233

glomerular capillaries creates a hypoxic environment thatfurther stimulates fibrosis.63 Exactly how TGF-� signal-ing in the endothelium promotes fibrosis in vivo is un-clear. Endothelial injury can stimulate TGF-� productionas shown in animal models of increased salt intake andsalt-sensitive hypertension in which TGF-� productionwas up-regulated by endothelial cells of renal arteri-oles.64,65 Conversely, TGF-� may have adverse effects onthe renal microvasculature by promoting both endothelialapoptosis and de-differentiation, leading to increased ma-trix production.66,67 TGF-� also may modulate angiogen-esis, a pivotal event in the restoration of healthy renalvasculature after injury. TGF-� signaling on endothelialcells in vitro has been shown to have a net anti-angio-genic effect.68 However, the in vivo effect of TGF-� onangiogenesis is complicated because this growth factorinduces renal proximal tubular expression of both vascu-lar endothelial growth factor-A and thrombospondin-1,which have pro-angiogenic and anti-angiogenic effects,respectively, in vivo.69-71 These findings suggest thatTGF-� signaling on injured epithelium may affect theadjacent vasculature and that divergent effects on angio-genesis may occur. Interestingly, a neutralizing TGF-�antibody protected against the vascular loss that occurred5 weeks after ischemia-reperfusion, implying that TGF-�may mediate chronic microvascular changes that resultfrom acute kidney injury.72 Whether these changes werecaused by TGF-�–mediated anti-angiogenesis or causedby other effects on the endothelium (eg, apoptosis) isunclear. To better define the role of TGF-� in the endo-thelial response to injury, future studies should examineendothelial-specific TGF-� signaling in vivo as well asanalyze the TGF-�–mediated cross-talk between injured

Figure 3. The effects of TGF-� on monocytes/macrophages. TGF-�such as macrophage inflammatory protein-1� (MIP-1�) and monocytof monocytes also are enhanced by TGF-� through up-regulated expTGF-� suppresses activation of macrophages as evidenced by decreproduction of inflammatory cytokines such as IFN-� but increases tinterferon-�; iNOS, inducible nitric oxide synthase; TNF-�, tumor ne

tubular epithelium and the surrounding vasculature.

CONCLUSIONS

TGF-� clearly plays a key role in the progression ofubulointerstitial fibrosis, but the mechanisms wherebyhis occurs remain undefined. Understanding how TGF-�

mediates tubulointerstitial fibrosis is challenging becausemultiple cell types are involved in fibrosis and the re-sponses to TGF-� vary, depending on cell type andellular milieu. Fortunately, the advent of Cre/lox tech-ology allows for selective inhibition of TGF-� signalingn specific compartments of the kidney. This tool willmprove our understanding of cell-autonomous TGF-�

signaling, providing important information regarding theeffects of inhibiting TGF-� activity in targeted cell typesin vivo. Another important direction for future studies iselucidating cell-specific downstream pathways ofTGF-�. These pathways likely mediate the divergenteffects of TGF-�, and defining how these pathways differin epithelial cells and fibroblasts may identify pharma-cologic targets with safer profiles than TGF-�. Also, abetter understanding of how inhibiting TGF-� signalinglters the activity of other growth factors after renalnjury is crucial and could have important therapeuticmplications. As the population with chronic kidney dis-ase continues to expand, a safe and effective way toodulate TGF-�–dependent tubulointerstitial fibrosisould make a huge medical and economic impact.

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