BRIEF REVIEW www.jasn.

org

Failed Tubule Recovery, AKI-CKD Transition, and Kidney

Disease Progression
† ‡ §
Manjeri A. Venkatachalam,* Joel M. Weinberg, Wilhelm Kriz, and Anil K. Bidani
†
*Department of Pathology, University of Texas Health Science Center, San Antonio, Texas; Department of Medicine,
Veterans Affairs Ann Arbor Healthcare System and University of Michigan Medical Center, Ann Arbor, Michigan;
‡
Medical Fakultät Mannheim, Abteilung Anatomie und Entwicklungsbiologie Mannheim, University of Heidelberg,
§
Baden-Wuerttemberg, Germany; and Department of Medicine, Loyola University and Hines Veterans Affairs Hospital,
Maywood, Illinois

ABSTRACT
The transition of AKI to CKD has major clinical significance. As reviewed here, Regardless of the diverse origins of
recent studies show that a subpopulation of dedifferentiated, proliferating tubules CKD in blood vessels, glomeruli, or tu-
recover-ing from AKI undergo pathologic growth arrest, fail to redifferentiate, and bules, tubulointerstitial fibrosis is the ma-
become atrophic. These abnormal tubules exhibit persistent, unregulated, and 26
jor pathway of progression to ESRD. In
progressively increasing profibrotic signaling along multiple pathways. Paracrine CKD caused by hypertension or GN, fi-
products derived therefrom perturb normal interactions between peritubular brosis develops around tubules made
capillary endothelium and pericyte-like fibroblasts, leading to myofibroblast atrophic by ischemia, misdirected glo-
transformation, proliferation, and fibrosis as well as capillary disintegration and 27,28
merular filtration, and disuse. After
rarefaction. Although signals from injured endothelium and inflammatory/immune primary tubule injury by AKI, glomeruli
cells also contribute, tubule injury alone is sufficient to produce the interstitial remain structurally normal over the short
pathology required for fibrosis. Localized hypoxia produced by microvascular term, even as tubules atrophy and fibrosis
pathology may also prevent tubule recovery. However, fibrosis is not intrinsically develops. However, over the long term,
progressive, and microvascular pathology develops strictly around damaged glomeruli in nephrons that emerge un-
tubules; thus, additional deterioration of kid-ney structure after the transition of AKI scathed from AKI can suffer hypertensive
to CKD requires new acute injury or other mechanisms of progression. Indeed, damage and foster progression if AKI had
experiments using an acute-on-chronic injury model suggest that additional loss of occurred in kidneys with reduced renal
29
parenchyma caused by failed repair of AKI in kidneys with prior renal mass reserve.
reduction triggers hemodynamically mediated pro-cesses that damage glomeruli to Why some tubules damaged by AKI
cause progression. Continued investigation of these pathologic mechanisms should become atrophic and then give rise to
reveal options for preventing renal disease pro-gression after AKI. subsequent long-term adverse effects,
whereas others recover completely is a
J Am Soc Nephrol 26: 1765–1776, 2015. doi: 10.1681/ASN.2015010006
fundamentally important unanswered
question. Particularly noteworthy in the
AKI-CKD transition and subsequent
Incomplete recovery from AKI can lead to adverse consequences attributable to im- progression are epithelial pathologies that
long-term functional deficits that are severe paired blood flow autoregulation, glomer- prevent tubule recovery, the cellular
and progressive in subpopulations of ular hypertension, glomerulosclerosis, and biology of fibrosis, and the significance of
patients with preexisting CKD.1–6 Kidneys tubulointerstitial fibrosis.21–25 Addi-tional
from patients recovering from AKI exhibit nephron loss by AKI in patients with CKD
chronic dysfunction, tubule atrophy, and could tip the balance of func-tional reserve Published online ahead of print. Publication date
7–20 4 available at www.jasn.org.
interstitial fibrosis (Figure 1, E and F). through hemodynamic ef-fects. Thus,
Correspondence: Dr. Manjeri A. Venkatachalam,
Incomplete recovery from AKI in patients failed recovery from AKI may have far-
Department of Pathology, University of Texas
with CKD not only adds to preexisting reaching significance. Recent re-search has Health Science Center, 7703 Floyd Curl Drive, San
pathology and dysfunction but also, may provided insights into the path-ologic basis Antonio, TX 78229. Email: venkatachal@uthscsa.
synergize with hemody-namic mechanisms for this failed recovery from AKI (i.e., edu
of progression. Severe loss of kidney mass tubule atrophy and renal fibrosis Copyright © 2015 by the American Society of
by CKD has long-term [tubulointerstitial fibrosis]). Nephrology

J Am Soc Nephrol 26: 1765–1776, 2015 ISSN : 1046-6673/2608-1765 1765

 BRIEF REVIEW www.jasn.org

renal mass reduction (RMR) by prior

CKD. Here, we review the pathology and
pathophysiology of the AKI-CKD tran-
sition and its potential over the long term
to degrade nephrons that had not been
severely injured by the initial AKI insult.

BY ITSELF, RENAL FIBROSIS

IS NOT PROGRESSIVE;
PROGRESSION REQUIRES
ADDITIONAL INJURY

Although fibrosis in response to injury is

often perceived to be a pathologic and
destructive event, it is essentially a self-
limiting repair process that restricts in-
jury. After tissue damage, fibroblasts use
signaling and genetic/epigenetic pro-
grams to proliferate and make connec-
tive tissue but then, regress as scar tissue
matures and contracts. Therefore, such
fibrosis is not autonomous and does not
expand or invade normal tissue. Rather, it
shrinks. Damage to tubules is associ-ated
with fibrosis around them. Endo-thelial
injury and capillary loss around damaged
tubules may produce hypoxia that likely
prevents recovery of the af-fected
30–
segments and ensures tubule at-rophy.
38
However, such interactions are
confined to diseased tubulointersti-tial
microenvironments. Therefore, as in any Figure 1. Injured kidney tissue heals by fibrosis that does not extend to involve previously
other scar, fibrotic tissue that is formed nondiseased parenchyma. (A and C) Periodic acid–Schiff staining of the kidney from an
around damaged tubules shrinks over autopsy of a patient with human FSGS. (A) Advanced scar containing sclerotic glomerulus,
time as activated fibroblasts regress and atrophic tubules with greatly thickened basement membranes, and interstitial fibrosis is
collagens mature. That is, fibrosis sharply demarcated from completely normal parenchyma (block arrows with red outlines).
develops only around atrophic tubules, There is no indication of fibrosis spreading from the scar into the adjacent interstitium. (C)
and the surrounding tubulointerstitium Small scar from the same kidney showing a few atrophic tubules with thick basement
membranes and expanded interstitium adjacent to the atrophic tubules (red arrows) near a
remains normal. Pathologic observations
glomerulus with mild mesangial expansion. The histopathology is one of resolved injury to
bear this out—kidney surfaces in benign
tubules with development of a shrunken scar in relationship to an atrophic nephron with no
nephrosclerosis display shrunken scars
suggestion that the lesion is invasive or expansive in its nature. (B and D) Kidney of rat 14
alternating with areas of hypertrophic days after AKI was induced by proximal tubule selective toxin maleic acid stained with
cortex. Biopsies of patients with chronic Masson’s Trichrome. (B) Low-power micrograph showing a localized lesion containing
glomerular disease show clusters of atro- undifferentiated atrophic tubules surrounded by a florid early fibrotic response (yellow
phic tubules surrounded by fibrosis that is brackets). The lesion is sharply demarcated from the adjacent well differentiated proximal
confined to regions containing dis-eased tubules that had either recovered normally after AKI or had not been injured by the poison.
28
glomeruli (Figure 1, A and C). (D) High-power micrograph showing a single profile of an atrophic tubule with surrounding
Similarly, during repair of experimental fibroblastic response (yellow brackets). Adjacent proximal tubules are well differentiated. The
AKI, tubules that fail to recover normal interstitium between them is either normal or minimally expanded. Interstitial fibro-blastic
structure become atrophic, and fibrosis responses that occur after AKI resolve and regress as tubules recover and re-differentiate or
persist and undergo scarring if tubules fail to redifferentiate and become atrophic. (E and F)
develops around them in microenviron-
Kidney biopsy from a patient 10 months after post-transplant AKI with delayed graft function
ments sharply demarcated from tubules
stained with Masson’s Trichrome (provided by Robert B. Colvin, Massachusetts General
that had recovered normally or had not
Hospital and Harvard Medical School, Boston, MA). (E) Low-power
been injured (Figure 1, B and D) (AKI

1766 Journal of the American Society of Nephrology J Am Soc Nephrol 26: 1765–1776, 2015

induced by maleate and tissue from the
work by Lan et al.39). Much later, such
lesions would shrink further and become
less perceptible (i.e., they do not prog-ress).
Fibrotic scars that develop after is-chemia-
reperfusion injury (IRI) show similarly
sharp demarcations from nor-mal kidney
tissue (figure 2 in the work by Goldfarb et
al.,40 figure 4g in the work by Yang et al.,41
and figures 10 and 11 in the work by Lan et
al.39). Indeed, small is-lands of pristine
kidney remain within areas of extensive
scarring after IRI (fig-ures 10 and 11 in the
work by Lan et al.39); fibrosis had not
involved these preserved tubules because
they either had not been injured or had
recovered normally after injury. Pathologic
features of healed hu-man post-transplant
AKI with delayed graft function bear this
out: 10 months after AKI, there is patchy
fibrosis contain-ing atrophic tubules, but the
fibrotic scars are clearly separated by
sharply demar-cated healthy parenchyma,
with only minimal focal increase of
interstitial con-nective tissue (Figure 1, E
and F).
Thus, insights from studies of kidney
pathology inform us that progressive renal
fibrosis requires additional, repet-itive,
and/or severe damage of previously normal
nephrons unless primary inter-stitial disease
is itself the instigating factor for fibrosis.
One possibility is that one or more episodes
of acute injury subsequent to initial AKI
cause progression. Re-peated but not single
episodes of myo-hemoglobinuric AKI
caused fibrotic CKD.42 Alternatively, single
episodes of massive AKI could produce
severe tubu-lointerstitial fibrosis and
consequent RMR, triggering adverse
hemodynamic events that foster
progression, particu-larly in settings where
functional renal mass had been reduced by
prior CKD (see below). These principles
apply to all forms of primary glomerular
and tu-bulointerstitial disease: progression
micrograph showing shrunken mature scars separated by healthy parenchyma with minimal
or no increase of interstitial connective tissue. (F) High-power micrograph with sharply
demarcated boundary between scar and healthy tubules. One healthy well differentiated
proximal tubule remains within the scar, suggesting that it had not been injured during the AKI
episode 10 months earlier or had recovered normal structure during regeneration and repair
after injury. Scale bars, 100 mm in A–D; 300 mm in E; 200 mm in F.
J Am Soc Nephrol 26: 1765–1776, 2015
requires serial recruitment of nephrons
to the injury process—be it slow and in-
dolent or rapid and severe. Fibrosis pro-
gresses incrementally, closely following
each nephron or clusters of nephrons
damaged by nascent injury.
CELLULAR ORIGIN OF
INTERSTITIAL FIBROSIS AFTER AKI
After AKI, humoral factors from regen-
erating tubules as well as inflammatory
cells, including monocytes, lymphocytes,
and dendritic cells, activate interstitial
precursor cells that become (myo)fibro-
blasts, which proliferate and make con-
nective tissue. The majority of resident
43,44
precursors—termed pericytes or fi-
28,45,46
broblasts —are cells with branch-
ing processes that contact capillaries and
28,43,47,48
tubules. They express peri-cyte
markers 59-ectonucleotidase (CD73) and
type I collagen and make contacts with
28,43,47,48
dendritic cells. By lineage
analysis, precursors termed peri-cytes are
derived from FoxD1–expressing
44
embryonic progenitors. Ephrin B sig-
naling between pericytes and capillary
endothelium maintains pericyte quies-
49
cence and endothelial integrity. Main-
tenance of endothelial integrity also
involves vascular endothelial growth fac-
tor (VEGF) produced by pericytes and
37,50
proximal tubules. Activating signals
from several sources disrupt these inter-
actions, inducing intercellular proteoly-sis
that is followed by dissociation from each
other. Two important effects ensue: (1)
PDGFRb-mediated migration and
transformation of fibroblasts/pericytes to
a-smooth muscle actin (a-SMA)
–expressing (myo)fibroblasts and (2)
dysangiogenic VEGF signaling, causing
37
loss of endothelial integrity attribut-able
to loss of the nursing function of pericytes
51
that stabilize capillaries.
www.jasn.org BRIEF REVIEW
With continued activation, the intersti-
tium becomes widened by proliferating
myofibroblasts and connective tissue,
and the injured endothelium regresses,
causing capillary rarefaction (Figure
51
2A).
Pathologic events at the endothelial–
pericyte/fibroblast interface that disrupt
capillaries and cause the detachment,
transformation, migration, and prolifer-
ation of fibroblasts include perturba-tions
of the activities of disintegrin ADAMTS-
1 and protease inhibitor TIMP3,
bidirectional Ephrin B2 signal-ing, and
37,43,48,52 –54
VEGFR2. These events are
reinforced by signaling through wnt,
TGF-b, PDGF-B, connec-tive tissue
growth factor (CTGF), and sonic
hedgehog pathways in myofibro-blasts,
with contributions from micro-RNA21,
37,52,54–63
PPARa, and NOX4. Although
triggered by tubule damage and loss of
endothelial–pericyte/fibro-blast
interactions, the subsequent prolif-eration
of (myo)fibroblasts, fibrosis, and capillary
rarefaction may depend also on cytokines
and growth factors from mon-ocytes that
infiltrate injured kidneys (Figure 2).
64,65
Although pericytes seem to be the major
source of fibroblasts driving fibro-sis, other
sources exist. Classic fibroblasts without
special relationship to capillaries are present
between tubules and around arterioles, and
they contribute to fibro-sis.47,66 Bone
marrow–derived precur-sors were reported
to infiltrate kidneys after injury and become
myofibro-blasts.67,68 However, these
invading cells did not express collagen or
proliferate after TGF-b stimulation,67,68
unlike fi-broblasts that do.69–71 Moreover,
hema-topoietic stem cells (cells reported to
become myofibroblasts in injured kidneys)
can fuse with tissue recipients and produce
tetraploid cells, thereby generating spuri-
ous signals of transdifferentiation72–74; also,
rare bone marrow monocytes/ macrophages
can express a-SMA.75,76 Moreover, other
investigators could not
identify transplantable bone marrow pre-
cursors of a-SMA–expressing cells in nor-
mal or injured kidneys.76,77 Bone marrow
derivation of kidney myofibroblasts re-
mains confounded by lineage issues and
AKI-CKD Transition 1767
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68,81
derivation from endothelium —is also
controversial owing to possible lin-eage
artifacts caused by nonspecificity of
78
endothelial markers, whereas sev-eral
publications have convincingly refuted a
role for epithelial-mesenchymal
46,66
transitions in kidney fibrosis.

VASOCONSTRICTION,
ENDOTHELIAL INJURY,
CAPILLARY COMPRESSION, AND
RAREFACTION: A ROLE FOR
POOR BLOOD FLOW AND
HYPOXIA AFTER AKI

Blood flow is persistently decreased after

IRI in deep cortex and outer medulla— the
regions most injured by ischemia. The
decrease is brought about by vaso-
constriction, tissue edema, endothelial
swelling, and capillary disintegration, lead-
ing to microvascular rarefaction.31,36,82–95
Reduced blood flow during the exten-sion
phase of IRI could cause hypoxia in
microenvironments of injured tu-bules and
promote tubulointerstitial fibrosis.31,34,36,82–
86,95,96
Microvascular defects and
tubulointerstitial fibrosis af-ter AKI occur in
close conjunction with tubule hypoxia
shown by pimonidazole adduct
formation.34,40,96 VEGF-A ex-pression in
proximal tubules is lost early after AKI
followed by decreased peritubular capillary
density, perfusion defects, and tubule
Figure 2. Pathologic events in tubules and interstitium interact to produce
hypoxia,30,31,33,34,36,96,97 con-sistent with a
tubulointerstitial fibrosis. (A) Schematic diagram of (left panel) normal tubule-interstitium
role for tubule VEGF-A for peritubular
and (right panel) early tubulointerstitial fibrosis. Resident fibroblasts in the interstitium
may or may not have intimate relationships to peritubular capillaries and basement capillary development50 and proliferation.98
membranes of tubules. The former type has also been termed pericyte. After injury, this Failure of endothelial cells to regenerate
type of fibroblast/pericyte detaches from capillaries, initiating pathologic events that after kidney injury may explain capillary
cause capillary disintegration and rarefaction as well as myofibroblasts transformation rarefaction.35 In-terventions that avert tissue
and proliferation. This process is aided and abetted by inflammatory cells, chiefly hypoxia by increasing blood flow or
monocytes, and resident immune cells, including dendritic cells. (B) Schematic diagram maintaining en-dothelial integrity mitigate
illustrating vicious cycle feedback inter-actions between tubule pathology and interstitial tubulointer-stitial fibrosis after kidney
pathology that potentiate tubule atrophy. Modified from reference 4, with permission. injury.34,37,99 These findings dovetail
findings that tu-
technical artifacts. There also are difficul- the large numbers of myeloid cells that do bule VEGF-A in tubules decreases after
78
ties in distinguishing markers in bone infiltrate injured kidneys. More-over, injury, and VEGF-A expression in peri-
marrow–derived precursor cells and my- the concept that fibrocytes of my-eloid cytes/fibroblasts and macrophages shifts
79 from predominantly VEGF164 isoform to
eloid cells in injured kidneys from those of lineage become myofibroblasts in the
fibroblasts/pericytes and myofibroblasts on kidney
80
is doubtful owing to un- dysangiogenic isoforms (VEGF120 and
37
account of complex branching processes in certainties of collagen expression versus -188). Concurrently with these al-
the latter. High-resolution microscopy is 78 terations, PDGF-B increases in tubules,
collagen internalization by these cells.
required to distinguish fibroblasts/ pericytes Endothelial-mesenchymal transition— a endothelium, and macrophages. Cru-
and their transformants from putative mechanism for myofibroblast cially, blockade of PDGFRb or VEGFR2

1768 Journal of the American Society of Nephrology J Am Soc Nephrol 26: 1765–1776, 2015

signaling by soluble ectodomains of their
receptors prevents myofibroblast transfor-
mation and capillary damage and rarefac-
tion, maintains normal capillary–pericyte/
fibroblast interactions, and ameliorates fi-
brosis.37 These studies emphasize that
physiologic signaling between endothelial
cells and FoxD1+ pericytes/fibroblasts
maintains their quiescent and differenti-ated
states and keeps in abeyance disrup-tive
VEGF and PDGF-B signaling that causes
capillary rarefaction and myofi-broblast
transformation.
Kidneys are physiologically hypoxic in
vulnerable medullary regions, where pO2 is
normally as low as 4–5 mmHg.86,100–102 In
view of tenuous oxygen tensions in mi-
tochondria of parenchymal cells103 caused
by steep oxygen gradients from capillaries
across interstitial spaces and cytoplasm, 104
oxygen available for respiration could fall
further to precipitously low concentra-tions
when interstitial spaces are widened by
edema and inflammation and capillar-ies
regress during fibrosis. Indeed, patho-logic
hypoxia shown by pimonidazole technique
in deep cortex—outer medul-lary regions
during early reperfusion after IRI 105—
persists as fibrosis develops.34,40
Consequent to hypoxia, tubule recovery
after AKI could be impaired by oxidant
stress, protein synthesis inhibition, and
growth arrest—the known adverse effects
of hypoxia.106–109 However, such hypoxic
effects should remain confined to the in-
jured tubule-interstitial microenviron-
ments. In such locations but not beyond,
hypoxia could prevent epithelial recov-ery
through feedback effects that ensure tubule
atrophy (Figure 2). Apropos the effects of
hypoxia on tubule recovery, the actions of
TGF-b antagonism to promote tubule
differentiation during recovery from IRI 110
could be ascribed to not only direct effects
of TGF-b an-tagonism on regenerating
110
tubules but also, conceivably, effects that
preserve the renal microvasculature, thereby
mitigating hypoxia and averting tubule
110,111
atrophy and fibrosis. It is worth
noting, however, that hypoxic effects that
lead to tubule atrophy and fibro-sis may
involve other mechanisms as well,
including HIF-1–dependent and
101,112
–independent processes.
J Am Soc Nephrol 26: 1765–1776, 2015
ROLES OF CORTICAL VERSUS
MEDULLARY PATHOLOGY IN THE
AKI-CKD TRANSITION
As outlined above, falling oxygen ten-
sions in the renal medulla could injure
tubules after AKI; however, transition
from acute injury to chronic medullary
disease is only partially understood.
Being most prone to injury after AKI, S3
proximal tubule segments in medul-lary
rays of the inner cortex and the outer
stripe of outer medulla (OSOM) have
received most attention. In part, S3 seg-
ments are most injured because of cell-
113
specific susceptibility of S3 cells.
However, complexity of outer medullary
microcirculation, disproportionately poor
blood reflow to medulla after is-chemia,
and tubule hypoxia caused by oxygen
gradients attributable to coun-tercurrent
vascular systems also contrib-ute
86,87,114–116
substantially. Most of our
understanding of the AKI-CKD transi-
tion is, in fact, derived from research on
the progression of tubulointerstitial fi-
brosis in the OSOM. We note, however,
that such medullary pathology in the
OSOM and conceivably, the inner stripe
of outer medulla (see below), if exten-
sive, could give rise to secondary damage
in the cortex as a consequence of hemo-
dynamic injury mechanisms triggered by
significantly reduced renal mass (dis-
cussed below).
However, we have little knowledge
regarding the development of chronic
pathology after AKI in the inner stripe of
outer medulla—a region crucial for sev-
eral critical kidney functions and the site
of a dense grouping of collecting ducts in
the interbundle region most distant from
the vascular bundles and thus, most
117
vulnerable to hypoxia. This mi-
croanatomic feature suggests that pa-
thology of the inner stripe will affect the
integrity of large areas of the cortical
hinterland in proportion to the number of
collecting ducts that are involved.
Cortical damage caused by medullary
pathology would be particularly severe if
medullary tubules undergo atresia as the
result of tubulointerstitial fibrosis and
therefore, become obstructed. It is
surprising that this aspect of post-AKI
www.jasn.org BRIEF REVIEW
pathophysiology receives little attention.
The cortical consequences of tubule
obstruction in the medulla are exempli-
fied by clinical experience; human pap-
illary necrosis, such as that caused by
acetaminophen toxicity, causes damage to
the papilla, but this is followed by
cortical atrophy. Studies have empha-
sized acute damage in ascending thick
limbs of Henle caused by reduced blood
flow and hypoxia attributable to adverse
oxygen gradients,
40,86,114,115,117
but there
has been little focus on chronic pa-
thology. In this connection, we have
consistently noticed significant fibrosis
with reduced tubule numbers in the in-ner
stripe chronically after IRI (unpub-lished
observations), but we, also, have not
studied this pathology in detail. In-depth
investigation of this aspect of medullary
pathology after AKI is clearly in order.
FAILED DIFFERENTIATION OF
REGENERATING EPITHELIUM
LEADS TO PROFIBROTIC
SIGNALING THAT PERSISTS IN
TUBULES UNDERGOING
ATROPHY AFTER AKI
Some proximal tubule cells that dedif-
ferentiate during regeneration after IRI fail
to redifferentiate and regain normal
structure during recovery. 39,110,118 Such
abnormally undifferentiated but growth-
arrested epithelium occurred along entire
tubule segments, in small cell clusters, or as
single undifferentiated cells surrounded by
differentiated proximal tubule cells late
during recovery after IRI. 39 Remarkably,
the abnormal epithelium displayed in-tense
signaling activity and expressed pro-fibrotic
peptides, despite being growth arrested and
atrophic by morphologic criteria. 39 These
findings were similar to earlier observations
on proximal tubules undergoing atrophy in
a microembolic is-chemia model. 119 On the
basis of these observations,39,119 we
postulated that tu-bules undergoing
pathologic growth ar-rest during
regeneration after AKI fail to differentiate,
signal vicariously through multiple
profibrotic pathways, and secrete fibrogenic
peptides into the interstitium,
AKI-CKD Transition 1769
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instigating fibrosis (Figure 3).4 Underly-ing

this hypothesis is the activation after AKI of
signaling pathways required for
dedifferentiation, migration, and prolifer-
ation. For this purpose, the surviving ep-
ithelium produces and secretes growth
factors, cytokines, and autacoids.82,120–122
These signaling and secretory events are
required for normal regeneration, but they
should cease when tubules recover.
Therefore, persistence of proliferative sig-
naling in growth–arrested undifferentiated
epithelium undergoing atrophy is inher-
ently abnormal. We showed that such atro-
phic tubules are engaged in pathologically
increased signaling through PI3K-Akt-
mTOR (mammalian target of rapamycin),
ERK-MAPK, JNK-MAPK, and TGF-b
pathways, with markedly increased expres-
sion of profibrotic peptides PDGF-B,
CTGF, and TGF-b.39,110,118,123
Experiments performed in tissue cul-ture
and in vivo showed that increased profibrotic
TGF-b signaling in tubules recovering from
AKI is, in part, attribut-able to autocrine
signaling by lysophos-phatidic acid.
Lysophosphatidic acid signaling through
separate G protein– coupled receptors triggers
avb6 integrin– dependent activation of latent
TGF-b as well as transactivation of EGFR and
ERK-MAPK. Although divergent when
initiated, the two pathways cooperatively
converge to increase TGF-b signaling and
thereby, increase the production and se-cretion
110,118,123
of PDGF-B and CTGF. Furthermore,
signaling intensities as well as signaling
protein and growth factor expression in these
abnormal tubules increased progressively with
time to strikingly high levels—far higher than
during the earlier stages of physiologic
39,110,118,123 Figure 3. Failed differentiation of proximal tubules regenerating after AKI leads to de-
regeneration. Interestingly, after
velopment of the atrophic abnormally signaling profibrotic tubule phenotype. (A) Sche-matic
119
microembolic kidney ischemia, fibrosis diagram illustrating (upper panel) the normal pathway of proximal tubule cell dedifferentiation
developed within interstitial spaces abutting and proliferation followed by redifferentiation and recovery of normal structure after AKI and
PDGF-B–expressing undifferentiated atro- (lower panel) the abnormal pathway of failure to redifferentiate after early dedifferentiation that
phic tubule epithelial cells but not differ- leads to tubule atrophy after AKI. (B, left panel) Immunohis-tochemistry and PHA lectin affinity
entiated cells without PDGF-B. In such cytochemistry of atrophic and normal proximal tubule cells 14 days after IRI in rats. Mosaic
fibrotic microenvironments, myofibro-blasts tubules showing well differentiated proximal tubule cells with brush border–bound PHA (lectin)
expressed PDGFRb, the cognate receptor for staining pink in close juxtaposition with atrophic epithelium without brush border that stains
PDGF-B, suggesting that PDGF-B from brown for the expression of vimentin, an in-termediate filament protein that is not present in
differentiated proximal tubule cells but is rapidly expressed after dedifferentiation during
atrophic epithelium had given rise to fibrosis
regeneration and retained after atrophy occurs. (B, right panel) Immunohistochemistry and
through paracrine effects.
lectin cytochemistry of proximal tubules 14 days after AKI induced by proximal tubule
selective toxin maleic acid. Well differenti-ated proximal tubule profiles and well differentiated
We have reported that atrophic tubules proximal tubule cells in a mosaic tu-bule (center) exhibit pink staining for brush border–bound
also exhibit near-total depletion of phos- PHA lectin but no nuclear staining for phospho-c-Jun, whereas atrophic tubule profiles and
atrophic cells in mosaic

1770 Journal of the American Society of Nephrology J Am Soc Nephrol 26: 1765–1776, 2015

phatase and tensin homolog (PTEN), the
lipid phosphatase that inhibits PI3K sig-
39
naling. PTEN is normally low in pro-
liferating proximal tubule cells but highly
expressed in quiescent differentiated epi-
thelium, and PTEN decreases were shown
to be driven, at least in part, by upstream
TGF-b signaling.39 However, the most
proximal causes of unregulated signaling in
the growth–arrested atrophic proximal
tubules that develop after IRI are un-known,
including those for TGF-b and the related
PTEN abnormality. Recent studies have
emphasized the role of prema-ture growth
arrest in giving rise to the pro-fibrotic tubule
phenotype.41 We also have observed that
pathologically dedifferenti-ated profibrotic
atrophic tubules with sig-naling disorders
are growth arrested, which was inferred by
lack of expression of Ki67, a marker for
cycling cells.39 Whether tubule growth
arrest during regenera-tion after AKI is a
uniquely controlled pathologic event that
causes profibrotic signaling or part of a
spectrum of pathol-ogy controlled by a
common upstream abnormality that also
disrupts differenti-ation programs, causes
ongoing cell in-jury, and instigates
profibrotic signaling remains to be
determined.
TUBULE SELECTIVE INJURY IS
SUFFICIENT TO DRIVE FIBROSIS,
INFLAMMATION, AND CAPILLARY
RAREFACTION
Paracrine triggers that compromise cap-
illary integrity and activate myofibro-blast
precursors during AKI may derive from
inflammatory/immune cells and stressed
tubule epithelium. Because tu-bules and
capillaries can be simultaneously injured
during AKI produced by ischemia, it is
difficult in these contexts to distinguish the
most proximal activating factors that
drive endothelial–pericyte/fibroblast dis-
sociation, capillary rarefaction, and myo-
fibroblast proliferation. Interventions,
such as TGF-b antagonism, that protect
against fibrosis development after
110,111
IRI cannot distinguish between
benefits conferred by restored microvas-
cular integrity and those that promote
tubule recovery. Nevertheless, tubule
damage, by itself, can be sufficient to pro-
duce the full spectrum of interstitial pa-
thology. Infiltrates of myofibroblasts and
monocytes/macrophages developed ex-
clusively around proximal tubules selec-
tively damaged by uranyl ions, and images
of 3H-thymidine autoradiography in sec-
tions stained for a-SMA showed clear re-
lationships of regenerating epithelium to
myofibroblasts in surrounding intersti-
124
tium. Fibrosis developed around tu-
bules selectively injured by folic acid,
and affected tubules in hypoxic environ-
ments (indicated by pimonidazole adducts)
showed loss of VEGF-A corresponding to
fibrosis and capillary loss in adjoining in-
96
terstitium. We investigated tubulointer-
stitial fibrosis in rats after injections of
39
maleic acid, a proximal tubule–selective
125
poison. Two weeks after maleic acid,
patchy fibrosis with mononuclear cell in-
filtrates developed around atrophic tu-
39
bules together with capillary rarefaction
and myofibroblast increase in the intersti-
tium (unpublished observations). Fur-
thermore, selective proximal tubule injury
produced by diphtheria toxin treatment
and Kidney Injury Molecule-1 or Notch1
overexpression leads to interstitial inflam-
mation, fibrosis, and CKD
126–128
; recent
studies suggest that tubule–derived wnt
and sonic hedgehog ligands, tubule-specific
activity of ADAM17 protease, and
other pathologies strictly localized to tu-
bules are sufficient to drive interstitial fi-
41,58,62,129–133
brosis We note here that
such cross-talk between injured tubules,
www.jasn.org BRIEF REVIEW
peritubular capillaries, and interstitial
cells through locally activated mecha-
nisms has the advantage of circumscrib-
ing the fibrotic response and minimizing
dissociation of nephron function and
vascular perfusion.
THE CKD-AKI-CKD CONNECTION:
AKI ON CKD CAN TRIGGER
HEMODYNAMIC MECHANISMS
OF CKD PROGRESSION
Hospital-based AKI in patients with prior
CKD adversely affects their long-term
outcomes (Figure 4); there is in-complete
recovery, increased mortality, and
predilection for progression to ESRD
among survivors.
1–6
We modeled this
clinical paradigm experimentally by
inducing IRI in rats with prior RMR.
29
Rats with implanted BP transducers were
subjected to IRI 2 weeks after sham
surgery, unilateral nephrectomy (moderate
RMR), or unilateral nephrec-tomy with
surgical removal of 50% renal mass from
the other kidney (severe RMR). BPs
recorded by radiotelemetry in conscious
rats were normal before IRI in all groups,
although slightly higher in rats with severe
RMR. Rats in all groups showed
equivalently severe azotemia and tubule
necrosis 3 days after IRI and similar
fractional masses of dedifferenti-ated
regenerating proximal tubules 7 days after
IRI. However, 4 weeks after IRI,
disproportionately greater fractions of
regenerating tubules in the severe RMR
group failed to differentiate and became
atrophic. Correspondingly, tu-bules with
failed differentiation were surrounded by
fibrosis. We surmised that stresses
associated with renal hyper-trophy after
RMR had compromised the ability of
regenerating tubules to enter
differentiation programs required to regain
normal structure. Although the nature of
the epithelial defect that preven-ted
redifferentiation remained unknown, it
was clear that failed differentiation had
given rise to greater atrophic tubule mass
and proportionately severe fibrosis.
Functional effect of the tubule defect was
reflected by incomplete abatement of
29
azotemia 4 weeks after IRI.
AKI-CKD Transition
tubule (center) show nuclear staining for phospho-c-Jun, indicating the activation of the JNK-
MAPK signaling pathway. (C) The diverse abnormalities exhibited by atrophic tubules are
listed. These several alterations take place in vimentin–expressing atrophic tubules illustrated
in B, left panel. Scale bars, 100 mm. A is modified from reference 4, with per-mission. B is
modified from reference 39, with permission. GPCR, G-protein coupled receptor; JNK-MAPK,
Jun N-terminal kinase-mitogen activated protein kinase; LPA, lyso-phosphatidic acid; PHA,
phytohemagglutinin; PTEN, phosphatase and tensin homolog.

J Am Soc Nephrol 26: 1765–1776, 2015 1771

 BRIEF REVIEW www.jasn.org
Figure 4. Failed tubule differentiation and RMR after AKI lead to hemodynamic abnor-
malities that cause progression. Schematic diagram illustrating the effects of AKI that
lead to tubulointerstitial fibrosis, the RMR that retards recovery of tubules regenerating
after AKI, and the resulting disproportionate further reduction of renal mass that triggers
hemody-namic mechanisms of renal disease progression.
Failed tubule recovery after AKI in rats
with severe RMR was attended by hy-
pertension.29 Severe RMR impairs renal
blood flow autoregulation.134,135 Severe
RMR, impaired blood flow autoregula-tion,
hypertension, and glomeruloscle-rosis are
closely related.134,136 Therefore, we
surmised that high systemic arterial
pressures in RMR rats recovering poorly
from AKI would be transmitted through less
responsive arterioles and damage glomeruli.
Rats with IRI after severe RMR not only
became hypertensive but also proteinuric; in
the most hyperten-sive animals, proteinuria
was severe, and glomeruli showed
focal/segmental hyalinosis, necrosis, and
sclerosis.29 These findings showed that, in a
normo-tensive setting with impaired blood
flow autoregulation (caused by RMR), poor
re-covery from superimposed AKI was
suffi-cient in itself to cause hypertension. 29
Rats with severe RMR were not
hypertensive before IRI. Without IRI,
control rats with severe RMR remained
normotensive for
29
6 weeks. Furthermore, the majority
of such rats with 75% RMR produced
by surgical excision without IRI are
known to remain normotensive up to
6 months,
137,138
unlike rats with 75% RMR
produced by unilateral nephrectomy and
1772 Journal of the American Society of Nephrology
partial renal infarction that develop hy-
pertension, vascular damage, and pro-
gressively increasing glomerulosclerosis,
tubule atrophy, and fibrosis by 16 weeks.135
Even lesser degrees of RMR may
predispose for such pathologies in the
aftermath of AKI. Uninephrectomized rats
(50% RMR) but not rats with intact kidneys
(0% RMR) developed hy-pertension,
proteinuria, and glomerulo-sclerosis over
several months after IRI.34,139–141 It is
unclear how incom-plete tubule healing and
fibrosis after AKI in rats with RMR give rise
to hyper-tension. Prior observations suggest
that hypertension after AKI may be volume
dependent. Kidneys develop microvascu-lar
defects after AKI,30,31,33–36,96 a pathol-ogy
that is particularly severe in kidneys with
RMR and superimposed IRI.29 Mi-
crovascular injury with reduction of per-
itubular capillary capacitance after AKI or
other tubulointerstitial disease may
predispose to impaired pressure natri-uresis,
reduced capacity of tubules to excrete salt,
and volume-dependent hy-
pertension.30,34,36,99,142–146 Notably, even
without RMR, rats remain normo-tensive
after IRI, but they become hyper-tensive
after salt loading—unlike normal rats,
which remain normotensive despite
144
high salt loads. Although additional
work is necessary to delineate the patho-
physiology of hypertension development
after AKI in rats with RMR, there can be
no doubt that increased BP bodes poorly
for kidneys in such contexts. Patients
with CKD with compromised kidney re-
serve who also undergo AKI are likely to
be at risk for long-term deterioration of
kidney structure and function through
hemodynamic mechanisms if they are
also hypertensive. Indeed, it is the most
plausible mechanism for progression to
ESRD in patients with CKD who experi-
ence superimposed AKI. The relation-
ships of preexisting hypertension in
cohorts of patients with CKD with addi-
tional AKI as well as hypertension devel-
opment after AKI in such cohorts need
investigation.
SUMMARY
In summary, recent studies have pro-vided
crucial connections between AKI and CKD
in terms of understanding how AKI
contributes to the progression of renal
disease. AKI by itself is a self-healing
process, but if severe, it leaves behind
tubule atrophy, interstitial fibro-sis, and
long-term dysfunction. How-ever, these
pathologies cannot and do not progress
without additional AKI epi-sodes. However,
if AKI is massive or superimposed on CKD
with compro-mised renal reserve, injured
tubules heal poorly and cause
disproportionately se-vere scarring with loss
of peritubular capillaries, setting in motion
a pathophys-iology that produces volume–
dependent salt–sensitive hypertension.
Because hy-pertension occurs in the setting
of im-paired renal blood flow
autoregulation, increased transmission of
arterial pres-sure damages glomeruli. Serial
glo-merular damage and consequent tubule
atrophy cause progression. On the basis of
these considerations, rigorous control of BP
and calorie restriction—an approach that
also decreases BP—still remain staple
strategies to delay CKD progression.
However, it is unclear why tubules regen-
erating after AKI sometimes fail to
differentiate, a pathology that instigates
J Am Soc Nephrol 26: 1765–1776, 2015

fibrosis. Therefore, investigation of the
conundrum of failed tubule differentia-
tion after acute injury has the potential
to uncover strategies that prevent AKI-
CKD transitions, preserve renal mass,
and thereby, delay hemodynamically
medi-ated progression.
ACKNOWLEDGMENTS
We thank Dr. Robert B. Colvin for
providing renal biopsy illustrations of
healed post-transplant AKI. Illustrations
from experi-mental models of AKI-CKD
transition are from the work by Lan et al.39
DISCLOSURES
None.
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