NDT Advance Access originally published online on September 26, 2008
Nephrology Dialysis Transplantation 2008 23(12):3773-3775; doi:10.1093/ndt/gfn533
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Does TNF-
enhance cystogenesis in ADPKD?*
Division of Nephrology, Cliniques Universitaires Saint-Luc and Université catholique de Louvain Medical School, B-1200 Brussels, Belgium
Correspondence and offprint requests to: Prof. Y. Pirson, Service de Néphrologie, Cliniques Universitaires Saint-Luc, Av. Hippocrate, 10, B-1200 BRUXELLES. Tel: +32-2-7641857; Fax: +32-2-7642836; E-mail: yves.pirson{at}uclouvain.be
Keywords: autosomal dominant polycystic kidney disease; autosomal recessive polycystic kidney disease; ethanercept; FIP-2; TNF-
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Unravelling nongenetic factors contributing to cystogenesis in ADPKD |
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 Top Unravelling nongenetic factors... Exploring TNF-{alpha}-mediated...Revealing a pathway connecting...References |
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Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutation in either PKD1 (85%) or PKD2 (15%) genes. The resulting disease phenotypes are very similar, except that renal disease is typically less severe in PKD2 families [1].
The protein products of PDK1 and PKD2, polycystin-1 (PC-1) and polycystin-2 (PC-2), are membrane proteins that probably form a functional complex [1]. PC-1 is regarded as a receptor for an unidentified ligand while PC-2 has significant homology to the transient receptor potential (TRP) family of store-operated calcium channels and is likely to function similarly as a non-selective calcium channel [1–3]. Like many other proteins implicated in renal cystic diseases, PC-1 and PC-2 are located in the primary cilium, a single hair-like organelle projecting from the surface of most mammalian cells. In tubular epithelial cells, the cilium projects into the lumen and is thought to act as a mechanosensor, detecting changes in the urinary flow, with a resulting calcium influx through the PC-2 channel; the calcium influx in turn induces release of calcium from intracellular stores [1].
How mutations in either PKD1 or PKD2 lead to the cystic phenotype of ADPKD is the subject of intense investigation. The several potential mechanisms of cystogenesis have been recently reviewed. They include increased cell proliferation and apoptosis, enhanced fluid secretion, abnormal cell–matrix interactions, alterations in cell polarity and abnormal ciliary structure and function [3]. One of the most likely involved mechanisms is an increased tubular proliferative activity induced by elevated levels of cyclic AMP (cAMP). The latter could be related to the reduced intracellular concentration of calcium resulting from defective function of polycystins [1]. The understanding of these mechanisms and the availability of animal models have enabled the identification of distinct potential treatments targeting calcium signalling, cAMP, proliferation signalling and tubular secretion [3].
The mutational event initiating this chain of events remains controversial. Although there is evidence for a two-hit mechanism (germline and further somatic inactivation of two PKD alleles)—explaining the focal development of cysts—haploinsufficiency increasingly appears to be sufficient to elicit a switch to a cystic phenotype. In mice, haploinsufficiency of Pkd1 leads to decreased intracellular calcium level and abnormal water handling in the collecting duct [4] whereas further lowering of Pkd1 expression is able to initiate cystogenesis [5]. Also in mice, haploinsufficiency of Pkd2 leads to increased tubular cell proliferation [6]. Furthermore, human ADPKD is characterized by a large intrafamilial variability in the severity of renal and extrarenal manifestations, pointing to the existence of genetic and/or nongenetic modifying factors. Analysis of the variability in renal function between monozygotic twins and siblings lends support to the role of genetic modifiers [7]. However, even within monozygotic twins, the up to 6-year difference in the age at end-stage renal disease strongly suggests the intervention of nongenetic factors [7].
The study just reported by Li X and co-workers [8] suggests that tumour necrosis factor- (TNF-) may be one of these nongenetic modifiers.
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Exploring TNF--mediated pathway as an enhancer of ADPKD
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TopUnravelling nongenetic factors...Exploring TNF-{alpha}-mediated...Revealing a pathway connecting...References |
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TNF-
, originally characterized by its ability to induce tumour cell apoptosis and cachexia, is now considered as a central mediator of a broad range of biological activities, encompassing beneficial effects in inflammatory and immune responses against a variety of infectious pathogens, as well as damaging effects in sepsis and auto-immune diseases [9]. The latter explains why anti-TNF antibodies are now successfully used in the treatment of disorders such as rheumatoid arthritis and inflammatory bowel disease.
A potential role of TNF- in enhancing cyst development in ADPKD—as well as in autosomal recessive polycystic kidney disease (ARPKD)—could be suspected from several observations. Firstly, TNF- mRNA and protein are markedly increased after hypertensive stress, renal injury and urinary tract infection, conditions that are commonly encountered in this disease [8,10]. Secondly, pathological levels of TNF- were found in 72% of kidney cyst fluids obtained from 13 patients with ADPKD [11]. Thirdly, increased levels of TNF- mRNA were found in the kidneys of the cpk mouse, a model of ARPKD [12]. Fourth, an inhibitor of TNF--converting enzyme was shown to ameliorate the polycystic disease in the bpk mouse, another model of ARPKD [13].
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Revealing a pathway connecting TNF- signalling, polycystins and cystogenesis
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TopUnravelling nongenetic factors...Exploring TNF-{alpha}-mediated...Revealing a pathway connecting...References |
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To investigate the potential connection between TNF-
and ADPKD, Li et al. used successively inner medullary collecting duct (IMCD) cell lysates, mouse embryonic kidney culture assay, primary cultures of normal and ADPKD kidney cells and finally Pkd2+/– mice. They focused on the measurement of FIP-2 (for 14.7K-interacting protein 2), a TNF--induced protein. FIP-2 is a member of a recently described family of interacting proteins, thought to play a role in trafficking from the apical recycling compartment to plasma membrane [14–16] as well as in regulating epithelial cell polarity [17].
Using IMCD cell lysates, they first confirmed that treatment with TNF- elevated the expression of FIP-2 by three- to fourfold. After finding that PC-2 and FIP-2 coimmunoprecipitate, they observed that cells treated with TNF- show a striking loss of PC-2 staining from its normal locations (plasma membrane and cilium) whereas PC-2 is enriched within perinuclear regions. That this effect is FIP-2 dependent was established by restoring normal PC-2 location in TNF--treated cells using small interfering RNA (siRNA) to knock down FIP-2 expression. Even though the expression levels of PC-1 and PC-2 remained the same in TNF--treated versus -untreated cells, coimmunoprecipitation assays revealed that treatment disrupted the PC-1/PC-2 interaction. Briefly, TNF- disrupted the normal location of PC-2 as well as the interaction between PC-1 and PC-2. Of note, location of PC-1 was not affected by TNF- treatment.
Using mouse wild-type embryonic day 15.5 (E15.5) kidney cultures under a low concentration of cAMP (such that there was no cyst formation), Li et al. found that treatment for 5 days with TNF- resulted in the development of numerous cyst-like structures (arising from proximal tubules and collecting ducts). Remarkably, in Pkd2+/– embryonic kidneys, cyst formation occurred at much lower TNF- concentrations than in wild-type embryonic kidneys. Immunoblot analysis confirmed that TNF- increased the expression of FIP-2 by four- to fivefold in wild-type embryonic kidneys and that FIP-2 abundance in untreated Pkd2+/– kidneys was markedly higher than that in wild-type kidneys and further stimulated by TNF- treatment. In other words, Pkd2+/– heterozygosity exacerbated the FIP-2-mediated cystogenic effect of TNF-. Of note, the concentrations of TNF- necessary for cyst induction were similar (in the ng/ml range) to those found by Li et al. in human freshly collected cyst fluids.
To further test whether FIP-2 abundance was elevated in human cystic kidneys, they compared the amount of FIP-2 protein between primary cultures of normal human kidney cells and cyst-lining cells from humans with ADPKD: the amount of FIP-2 was approximately twofold higher in PKD than in normal cells, similar to the observed FIP-2 level difference between Pkd2+/– and wild-type mouse embryonic kidneys.
Finally, to test the effect of TNF- on cyst formation in vivo, they intraperitoneally injected Pkd2+/– mice with TNF- (0.5 µg per gram body weight, one injection per week for 10 weeks). Whereas the control group developed cysts at the expected frequency (13/61, i.e. 21%), this frequency was higher (6/14, i.e. 43%) in the Pkd2+/– group. It is, however, noteworthy that only two mice developed multiple bilateral small cysts (while the 4 others had 
3 cysts). To test whether inhibition of TNF- could alleviate cyst formation, Pkd2+/– mice were treated with ethanercept, a TNF- inhibitor (one subcutaneous injection of 125 µg/ week for 10 weeks). At the end of treatment, none of the 50 treated mice developed cysts, as compared to 13/61 (21%, as expected) untreated animals.
Taken together, these findings suggest that TNF- is a potent factor that promotes kidney cyst development, especially in the genetic background associated with ADPKD.
Interestingly, Lee et al. recently found that TNF- can also activate the mammalian target of rapamycine (mTOR) pathway [18]. Since inhibition of mTOR has been shown to reverse cystogenesis in a mouse model of ADPKD [19], this pathway could also be involved in the TNF--mediated cyst development, as recognized by Li et al.
Weighing potential clinical implications of this work
Before considering potential implications of these findings in human ADPKD, their confirmation in other animal models of ADPKD (especially in Pkd1+/– and in more severely affected animals) would be welcome, and the mechanisms mediating the effect of TNF- on cystogenesis should be clarified.
If TNF- were definitely established as a promoter in the progression of human ADPKD, concurrent inflammatory and infectious states would be expected to accelerate cyst development and renal impairment. Such clinical observations are surprisingly lacking. The only available study is a retrospective analysis from Gabow et al. reported in 1992. Among the nongenetic factors potentially affecting the progression of renal disease in a cohort of patients with ADPKD, they showed that urinary tract infections in men (but not in women) was one of the variables independently associated with worsening of renal function [20]. Consistent with an enhancing role of infection, it was once demonstrated in mice that the development of cystic kidney disease was markedly slow in a germ-free as compared to an ambient environment [21]. Furthermore, a role of the immune system in the progression of polycystic kidney diseases has for long been suggested by the presence of inflammatory infiltrates and interstitial fibrosis in ADPKD (as well as ARPKD) human kidneys, and is now supported by recent studies using gene profiling [22]. One of them, a model for ARPKD performed in the cpk mouse, shows nicely in severely affected animals compared to mildly affected animals, the upregulation of genes expressed by activated macrophages and complement system factors including the central complement component 3 [23].
It would be most interesting that clinicians following cohorts of patients with ADPKD assess the progression of patients concurrently suffering from a chronic inflammatory disorder and those of them who would be receiving an anti-TNF treatment.
Finally, before considering a trial evaluating the therapeutic use of ethanercept in ADPKD, it is to remember that blockade of TNF- in human rheumatoid arthritis or Crohn's disease has led to the development of auto-antibodies, lupus-like syndrome and glomerulonephritis in some patients [10,24].
Take-home message
The large intrafamilial variability in the severity of manifestations of ADPKD points to the existence of nongenetic modifying factors. Using in vitro and animal experiments (mainly the Pkd2 +/– mouse), Li et al. show that one of these factors may be TNF-.
This could be an additional reason to aggressively treat infections and inflammatory disorders in ADPKD patients, and a starting point to consider the TNF- pathway as a potential therapeutic target in slowering the progression of this disease.
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Acknowledgments |
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The author thanks Olivier Devuyst and François Jouret for their very helpful comments and suggestions.
Conflict of interest statement. None declared.
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Notes |
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* This article is based on the science article from Li X, Magenheimer BS, Xia S et al. A tumor necrosis factor-
-mediated pathway promoting autosomal dominant polycystic kidney disease. Nat Med 2008; Aug; 14(8); 863-8. Epub 2008 Jun 15. 
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[Abstract/Free Full Text]
Accepted in revised form: 1. 9.08
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