Review of the Use of Animal Models of Human Polycystic Kidney Disease for the Evaluation of Experimental Therapeutic Modalities.

Autosomal dominant polycystic kidney disease, autosomal recessive polycystic kidney disease, and nephronophthisis are hereditary disorders with the occurrence of numerous cysts in both kidneys, often causing chronic and end-stage renal failure. Animal models have played an important role in recent advances in research not only on disease onset and progressive mechanisms but also on the development of therapeutic interventions. For a long time, spontaneous animal models have been used as the primary focus for human diseases; however, after the identification of the nucleotide sequence of the responsible genes, PKD1, PKD2, PKHD1, and NPHPs, various types of genetically modified models were developed by genetic and reproductive engineering techniques and played the leading role in the research field. In this review, we present murine models of hereditary renal cystic diseases, discussing their potential benefits in the development of therapeutic strategies.

Novel 3D capsule device to restrict kidney volume expansion on polycystic kidney progression: feasibility study in a rat model.

BACKGROUND: Cystogenesis in polycystic kidney disease (PKD) is likely accelerated by various renal insults, including crystal deposition, that activate renal tubule obstruction and dilation. We developed a capsule-based device that can be applied to cystic kidneys to restrict tubular lumen dilatation and cyst expansion. METHODS: Kidney capsule devices were designed from computed tomography images of wild-type and Cy/+ rats. Capsule devices were surgically implanted on kidneys in six surgical sessions over a period of 14 months in 7 wild-type rats of 6.5-8 weeks (3 sham operations, 2 right, 2 left) and 6 Cy/+ rats of 6.5 weeks (2 sham, 3 left, 1 bilateral). After surgery, the rats were followed for 5.4-12.4 weeks' growth and sacrificed to retrieve the kidneys. During the follow-up, serum creatinine was measured and retrieved kidneys were weighed. Histological analysis including cystic area measurement and immunohistochemistry was performed. RESULTS: Morphometric capsule devices were configured and developed by an image processing technique and produced using a 3D printer. Encapsulated Cy/+ kidneys (n = 5; mean weight 3.64 g) were consistently smaller in size (by 21-36%; p < 0.001) than unencapsulated Cy/+ kidneys (n = 7; mean weight 5.52 g). Encapsulated Cy/+ kidneys (mean %cyst area: 29.4%) showed smaller histological cystic area (by 28-58%; p < 0.001) than unencapsulated Cy/+ kidneys (mean %cyst area 48.6%). Cell proliferation and macrophages were also markedly reduced in encapsulated Cy/+ kidneys, compared to unencapsulated Cy/+ kidneys. CONCLUSIONS: We report a pilot feasibility study for the application of a novel morphometric 3D capsule device to the Cy/+ rat model showing restricted kidney volume expansion on polycystic kidney disease progression.

High Levels of Dietary Lard or Sucrose May Aggravate Lysosomal Renal Injury in Non-Obese, Streptozotocin-Injected CD-1 Mice Provided Isocaloric Diets.

Daily fat and sugar intake has increased in Japan, while total energy intake has decreased. However, the number of type 2 diabetes mellitus patients has increased, and this often causes renal injury characterized by autophagic vacuoles. Although many studies with comparisons of high fat or sugar versus a normal macronutrient balanced diet have been reported, there are few studies that equalized calorie intake and body weights. In the current study, AIN93M diets (CONT group) with matching energy content with lard derived high saturated fat (LARD group), soybean oil derived unsaturated fat (SOY OIL group) and sucrose (SUCROSE group) were provided to compare their effects on renal morphology in streptozotocin-injected CD-1 mice without causing obesity. The number of renal tubular vacuoles was higher in SUCROSE and slightly higher in LARD compared with CONT mice, and was higher in LARD and SUCROSE compared with SOY OIL mice. Most of those vacuoles were LAMP1-positive, a marker of lysosomal autophagy. These results suggest that despite identical energy contents, diets with high sucrose or saturated fat compared to unsaturated fat may aggravate lysosomal renal injury in a non-obese, streptozotocin-induced model of diabetes mellitus.

Increased salt intake does not worsen the progression of renal cystic disease in high water-loaded PCK rats.

The anti-diuretic hormone arginine vasopressin is thought to be a detrimental factor in polycystic kidney disease (PKD). We previously reported that high water intake (HWI) reduced urine osmolality and urinary arginine vasopressin, improved renal function, and reduced the kidney/body weight ratio in PCK rats, an orthologous model of human PKD. In PKD patients, however, it is reported that HWI increases total kidney volume, urine volume, and urine sodium excretion, which could be a consequence of high salt intake. In the current study, we loaded PCK rats with high salt concurrently with HWI to determine whether this human-imitated condition exacerbates disease progression. PCK rats were assigned into 4 groups: control group (CONT: distilled water), HWI group (HWI: 5% glucose in water), HWI with 0.2% NaCl group (HWI+0.2%NaCl), and HWI with 0.45% NaCl group (HWI+0.45%NaCl). Total water intake during the experimental period was increased by 1.86-, 2.02-, and 2.42-fold in HWI, HWI+0.2%NaCl, and HWI+0.45%NaCl, and sodium intake was increased by 2.55- and 5.83-fold in HWI+0.2%NaCl and HWI+0.45%NaCl, respectively, compared with CONT. Systolic blood pressure was higher in HWI+0.2%NaCl and HWI+0.45%NaCl than in both CONT and HWI. Serum urea nitrogen, kidney/body weight ratio, cAMP, cystic area, and fibrosis index were significantly lower in HWI compared with CONT, and these ameliorative effects were not abrogated in either HWI+0.2%NaCl or HWI+0.45%NaCl. The amount of sodium excreted into the urine was increased by 2.50- and 8.38-fold in HWI+0.2%NaCl and HWI+0.45%NaCl, respectively, compared with HWI. Serum sodium levels were not different between the groups. These findings indicate that the beneficial effect of HWI against the progression of cystic kidney disease was not affected even by high salt-overload in this rodent model of PKD.

Distinct effects of dietary flax compared to fish oil, soy protein compared to casein, and sex on the renal oxylipin profile in models of polycystic kidney disease.

Oxylipins are bioactive lipids derived from polyunsaturated fatty acids (PUFA) that are important regulators of kidney function and health. Targeted lipidomic analyses of renal oxylipins from four studies of rodent models of renal disease were performed to investigate the differential effects of dietary flax compared to fish oil, soy protein compared to casein, and sex. Across all studies, dietary fish oil was more effective than flax oil in reducing n-6 PUFA derived oxylipins and elevating eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) derived oxylipins, whereas dietary flax oil resulted in higher α-linolenic acid (ALA) oxylipins. Dietary soy protein compared to casein resulted in higher linoleic acid (LA) derived oxylipins. Kidneys from females had higher levels of arachidonic acid (AA) oxylipins, but similar or lower levels of oxylipins from other PUFA. Modulation of the oxylipin profile by diet and sex may help elucidate their effects on renal physiology and health.

Distinct oxylipin alterations in diverse models of cystic kidney diseases.

Cystic kidney diseases are characterized by multiple renal cysts and are the leading cause of inherited renal disease. Oxylipins are bioactive lipids derived from fatty acids formed via cyclooxygenase, lipoxygenase and cytochrome P450 activity, and are important regulators of renal health and disease. Oxylipins are altered in nephronophthisis, a type of cystic kidney disease. To further investigate and to determine whether other cystic renal diseases share these abnormalities, a targeted lipidomic analysis of renal oxylipins was performed in orthologous models of autosomal dominant polycystic kidney disease 1 (Mx1Cre+Pkd1flox/flox mouse) and 2 (Pkd2ws25/- mouse), autosomal recessive polycystic kidney disease (PCK rat) and nephronophthisis (jck/jck mouse). Kidney cyclooxygenase oxylipins were consistently higher in all diseased kidneys, even in very early stage disease. On the other hand, cytochrome P450 epoxygenase derived oxylipins were lower only in the autosomal recessive polycystic kidney disease and nephronophthisis models, while lipoxygenase and cytochrome P450 hydroxylase derived oxylipins were lower only in nephronophthisis. Sex effects on renal oxylipin alterations were observed but they did not always coincide with sex effects on disease. For oxylipins with sex effects, arachidonic acid derived oxylipins formed via cyclooxygenases and lipoxygenases were higher in females, while oxylipins from other fatty acids and via cytochrome P450 enzymes were higher in males. The consistent and unique patterns of oxylipin alterations in the different models indicates the importance of these bioactive lipids in cystic renal diseases, suggesting that pharmacological agents (e.g. cyclooxygenase inhibitors) may be useful in treating these disorders, for which effective treatment remains elusive.

Beneficial effect of combined treatment with octreotide and pasireotide in PCK rats, an orthologous model of human autosomal recessive polycystic kidney disease.

Increased intracellular cyclic AMP (cAMP) in renal tubular epithelia accelerates the progression of polycystic kidney disease (PKD). Thus, decreasing cAMP levels by an adenylyl cyclase inhibitory G protein activator is considered to be an effective approach in ameliorating PKD. In fact, pasireotide (PAS) was effective in reducing disease progression in animal models of PKD. However, hyperglycemia caused by the administration of PAS is an adverse effect in its clinical use. Whereas, co-administration of octreotide (OCT) with PAS did not increase serum glucose in normal rats. In the current study, we examined the efficacy of combined treatment with OCT and PAS in PCK rats, an autosomal recessive PKD model. Four-week-old PCK males were treated with the long-acting release type of OCT, PAS, or a combination of both (OCT/PAS) for 12 weeks. After termination, serum and renal tissue were used for analyses. Kidney weight, kidney weight per body weight, renal cyst area, renal Ki67 expression, and serum urea nitrogen were significantly decreased either in the PAS or OCT/PAS group, compared with vehicle. Renal tissue cAMP content was significantly decreased by PAS or OCT/PAS treatment, but not OCT, compared with vehicle. As a marker of cellular mTOR signaling activity, renal phospho-S6 kinase expression was significantly decreased by OCT/PAS treatment compared with vehicle, OCT, or PAS. Serum glucose was significantly increased by PAS administration, whereas no difference was shown between vehicle and OCT/PAS, possibly because serum glucagon was decreased either by the treatment of OCT alone or co-application of OCT/PAS. In conclusion, since serum glucose levels are increased by the use of PAS, its combination with OCT may reduce the risk of hyperglycemia associated with PAS monotherapy against PKD progression.

Lack of Benefit of Early Intervention with Dietary Flax and Fish Oil and Soy Protein in Orthologous Rodent Models of Human Hereditary Polycystic Kidney Disease.

Rationale for dietary advice in polycystic kidney disease (PKD) is based in part on animal studies that have examined non-orthologous models with progressive development of cystic disease. Since no model completely mimics human PKD, the purpose of the current studies was to examine the effects of dietary soy protein (compared to casein) or oils enriched in omega-3 fatty acids (fish or flax oil compared to soy oil) on early disease progression in two orthologous models of PKD. The models studied were Pkd2WS25/- mice as a model of autosomal dominant PKD, and PCK rats as a model of autosomal recessive PKD. After 13 weeks of feeding, dietary fish (but not flax) oil resulted in larger kidneys and greater kidney water content in female Pkd2WS25/- compared to control mice. After 12 weeks of feeding male PCK compared to control rats, both fish and flax compared to soy oil resulted in enlarged kidneys and livers, greater kidney water content and higher kidney cyst area in diseased rats. Dietary soy protein compared to casein had no effects in Pkd2WS25/- compared to control mice. In PCK rats, kidney and liver histology were not improved, but lower proteinuria and higher urine pH suggest that soy protein could be beneficial in the long term. Therefore, in contrast to studies in non-orthologous models during the progressive development phase, these studies in orthologous PKD models do not support dietary advice to increase soy protein or oils enriched in omega-3 oils in early PKD.

Cyclooxygenase product inhibition with acetylsalicylic acid slows disease progression in the Han:SPRD-Cy rat model of polycystic kidney disease.

Renal cyclooxygenase (COX) derived eicosanoids are elevated and lipoxygenase (LOX) products are reduced in the Han:SPRD-Cy rat model of polycystic kidney disease (PKD). Selective COX2 inhibition reduces kidney disease progression, but COX1 levels also are elevated in this model. Since the effect of reducing the products of both COX isoforms and the role of LOX products is not known, weanling normal and diseased Han:SPRD-cy littermates were given either low dose acetylsalicylic acid (ASA), nordihydroguaiaretic (NDGA) or no treatment for eight weeks. Renal eicosanoids were altered in the diseased compared to normal cortex, with COX products being higher and LOX products being lower. ASA reduced COX products, cyst growth and kidney water content, while NDGA reduced LOX products without altering disease progression or kidney function. Hence, a human equivalent ASA dose equal to less than one regular strength aspirin per day slowed disease progression, while further reduction of LOX products did not worsen disease progression.

Dietary flax oil rich in α-linolenic acid reduces renal disease and oxylipin abnormalities, including formation of docosahexaenoic acid derived oxylipins in the CD1-pcy/pcy mouse model of nephronophthisis.

The CD1-pcy/pcy mouse model of nephronophthisis displays reduced renal docosahexaenoic acid (DHA) levels and alterations in renal cyclooxygenase and lipoxygenase oxylipins derived from n-6 fatty acids. Since dietary flax oil ameliorates disease progression, its effect on renal fatty acids and oxylipins was examined. Sixteen weeks of feeding resulted in reduced disease progression and enrichment of renal phospholipid α-linolenic acid (ALA) and eicosapentaenoic acid, reduction in arachidonic acid (AA), but no change in linoleic acid (LA) or DHA. In diseased kidneys, flax oil feeding mitigated the elevated levels of renal cyclooxygenase derived oxylipins formed from AA and the lowered lipoxygenase and cytochrome P450 derived oxylipins formed from ALA and DHA. Increased DHA oxylipins occurred with flax feeding despite not altering DHA levels. Dietary flax oil may therefore reduce disease progression via mitigation of oxylipin abnormalities. This study also provides evidence of in vivo ALA conversion to DHA in amounts necessary to restore DHA oxylipin levels.

Renal cyclooxygenase and lipoxygenase products are altered in polycystic kidneys and by dietary soy protein and fish oil treatment in the Han:SPRD-Cy rat.

SCOPE: Dietary fish oil (FO) and soy protein (SP) are two interventions that slow disease progression in the Han:SPRD-Cy rat model of polycystic kidney disease (PKD). Inhibition of cyclooxygenase (COX)-derived eicosanoids also reduces disease progression, but the role of lipoxygenase (LOX) products in this disease is not known. METHODS AND RESULTS: Since dietary FO and SP have been shown to alter eicosanoid formation via differing mechanisms, Han:SPRD-Cy rats were given diets containing either casein protein (CP) or SP, and soy oil (SO) or FO. Analysis of eicosanoids revealed that renal COX products were higher and LOX products were lower in diseased kidneys. SP feeding resulted in lower COX products, activity and COX1 protein and higher LOX products in the diseased kidneys in parallel with reduced renal cyst growth and fibrosis. By comparison, FO reduced both COX and LOX products produced from n-6 fatty acids and increased 3-series prostanoids in both normal and diseased cortex and medulla, but these differences did not parallel effects on disease. CONCLUSION: Renal COX-derived eicosanoids are elevated and LOX products are reduced in this model of kidney disease. The effects of dietary SP, but not FO, on renal eicosanoids parallel the effects on disease.

Renal cyclooxygenase products are higher and lipoxygenase products are lower in early disease in the pcy mouse model of adolescent nephronophthisis.

Nephronophthisis (NPHP) is a pediatric form of hereditary polycystic kidney disease (PKD), and is the leading cause of end stage renal disease in children. The pcy mouse is an orthologous model of human NPHP, with a mutation in the Nphp3 gene. Renal phospholipase A2, cyclooxygenase (COX) 1 and cyclic AMP are elevated in this model, suggesting that eicosanoid formation may be altered. In another type of PKD observed in the Han:SPRD-Cy rat, inhibition of eicosanoid production slows disease progression. If renal eicosanoids are similarly altered in NPHP, potential for pharmacologic intervention also may exist for this disorder. Therefore, renal fatty acids and eicosanoids were determined in pcy and normal mice at 15, 30 and 60 days of age by gas chromatography and HPLC-tandem mass spectrometry, respectively. Renal cysts in enlarged kidneys were observed in pcy mice by 15 days of age and increased over time. Renal phospholipid ARA levels were higher in pcy compared to normal mice at 15 and 30 days. Eicosanoid differences were observed starting at 30 days, when the COX products 6-keto-prostaglandin (PG) F1α, thromboxane B2 and PGE2 were higher in pcy compared to normal kidneys. Overall, total COX products were elevated at 30 and 60 days. In contrast, the levels of the lipoxygenase (LOX) products were not altered until 60 days of age and these were lower in pcy kidneys compared to normal. These findings suggest that altered eicosanoids play a role in NPHP, and that manipulating these levels with pharmacologic agents may have therapeutic potential.

Telmisartan ameliorates fibrocystic liver disease in an orthologous rat model of human autosomal recessive polycystic kidney disease.

Human autosomal recessive polycystic kidney disease (ARPKD) produces kidneys which are massively enlarged due to multiple cysts, hypertension, and congenital hepatic fibrosis characterized by dilated bile ducts and portal hypertension. The PCK rat is an orthologous model of human ARPKD with numerous fluid-filled cysts caused by stimulated cellular proliferation in the renal tubules and hepatic bile duct epithelia, with interstitial fibrosis developed in the liver. We previously reported that a peroxisome proliferator activated receptor (PPAR)-γ full agonist ameliorated kidney and liver disease in PCK rats. Telmisartan is an angiotensin receptor blocker (ARB) used widely as an antihypertensive drug and shows partial PPAR-γ agonist activity. It also has nephroprotective activity in diabetes and renal injury and prevents the effects of drug-induced hepatotoxicity and hepatic fibrosis. In the present study, we determined whether telmisartan ameliorates progression of polycystic kidney and fibrocystic liver disease in PCK rats. Five male and 5 female PCK and normal control (+/+) rats were orally administered 3 mg/kg telmisartan or vehicle every day from 4 to 20 weeks of age. Treatment with telmisartan decreased blood pressure in both PCK and +/+ rats. Blood levels of aspartate amino transferase, alanine amino transferase and urea nitrogen were unaffected by telmisartan treatment. There was no effect on kidney disease progression, but liver weight relative to body weight, liver cystic area, hepatic fibrosis index, expression levels of Ki67 and TGF-ß, and the number of Ki67- and TGF-ß-positive interstitial cells in the liver were significantly decreased in telmisartan-treated PCK rats. Therefore, telmisartan ameliorates congenital hepatic fibrosis in ARPKD, possibly through the inhibition of signaling cascades responsible for cellular proliferation and interstitial fibrosis in PCK rats. The present results support the potential therapeutic use of ARBs for the treatment of fibrocystic liver disease in ARPKD patients.

Animal models for human polycystic kidney disease.

Polycystic kidney disease (PKD) is a hereditary disorder with abnormal cellular proliferation, fluid accumulation in numerous cysts, remodeling of extracellular matrix, inflammation, and fibrosis in the kidney and liver. The two major types of PKD show autosomal dominant (ADPKD) or autosomal recessive inheritance (ARPKD). ADPKD is one of the most common genetic diseases, with an incidence of 1:500-1,000. Approximately 50% of patients with ADPKD develop end-stage renal disease (ESRD) by the age of 60. On the other hand, ARPKD is relatively rare, with an incidence of approximately 1:20,000-40,000. ARPKD is diagnosed early in life, often prenatally. The gene products responsible for ADPKD and ARPKD distribute in primary cilia and are thought to control intercellular Ca(2+). Two types of animal model of PKD have been established: spontaneous hereditary models identified by the typical manifestations of PKD and gene-engineered models established by modification of human orthologous genes. Both types of animal models are used to study the mechanism of cystogenesis and efficacy of medical treatments. In PKD progression, critical roles of signaling pathways including MAPK, mTOR, and PPAR-γ have been discovered with these models. Therefore, experimental animal models are indispensable for investigating molecular mechanisms of PKD onset and progression as well as potential therapeutic treatments.

Global Gene Expression Profiling in PPAR-γ Agonist-Treated Kidneys in an Orthologous Rat Model of Human Autosomal Recessive Polycystic Kidney Disease.

Kidneys are enlarged by aberrant proliferation of tubule epithelial cells leading to the formation of numerous cysts, nephron loss, and interstitial fibrosis in polycystic kidney disease (PKD). Pioglitazone (PIO), a PPAR-γ agonist, decreased cell proliferation, interstitial fibrosis, and inflammation, and ameliorated PKD progression in PCK rats (Am. J. Physiol.-Renal, 2011). To explore genetic mechanisms involved, changes in global gene expression were analyzed. By Gene Set Enrichment Analysis of 30655 genes, 13 of the top 20 downregulated gene ontology biological process gene sets and six of the top 20 curated gene set canonical pathways identified to be downregulated by PIOtreatment were related to cell cycle and proliferation, including EGF, PDGF and JNK pathways. Their relevant pathways were identified using the Kyoto Encyclopedia of Gene and Genomes database. Stearoyl-coenzyme A desaturase 1 is a key enzyme in fatty acid metabolism found in the top 5 genes downregulated by PIO treatment. Immunohistochemical analysis revealed that the gene product of this enzyme was highly expressed in PCK kidneys and decreased by PIO. These data show that PIO alters the expression of genes involved in cell cycle progression, cell proliferation, and fatty acid metabolism.

PPAR-γ agonists in polycystic kidney disease with frequent development of cardiovascular disorders.

Autosomal dominant polycystic kidney disease (ADPKD) is the most common of the monogenic disorders and is characterized by bilateral renal cysts; cysts in other organs including liver, pancreas, spleen, testis and ovary; vascular abnormalities including intracranial aneurysms and subarachnoid hemorrhage; and cardiac disorders such as left ventricular hypertrophy (LVH), mitral valve regurgitation, mitral valve prolapse and aortic regurgitation. Autosomal recessive polycystic kidney disease (ARPKD) is an early-onset multisystem disorder characterized by polycysts divided from the renal collecting ducts, congenital hepatic fibrosis, and ductal plate malformation complicated by pulmonary hyperplasia and systemic hypertension. In these polycystic kidney diseases (PKD), progressive enlargement of the cysts results from the aberrant proliferation of tubule epithelial cells and trans-epithelial fluid secretion leading to extensive nephron loss and interstitial fibrosis. Peroxisome proliferator-activated receptor-γ (PPAR-γ), a member of the ligand-dependent nuclear receptor superfamily, is expressed in a variety of tissues, including kidneys and liver, and plays important roles in cell proliferation, fibrosis, and inflammation. PPAR-γ agonists ameliorate polycystic kidney, polycystic liver and cardiac defects through ß-catenin, c-Myc, CFTR, MCP-1, S6, ERK, and TGF-ß signaling pathways in animal models of PKD. In this review, we describe the possible therapeutic value of PPAR-γ agonists in the treatment of renal and hepatic manifestations, and cardiac defects in progressive PKD.

Tolvaptan inhibits ERK-dependent cell proliferation, Cl⁻ secretion, and in vitro cyst growth of human ADPKD cells stimulated by vasopressin.

In autosomal dominant polycystic kidney disease (ADPKD), arginine vasopressin (AVP) accelerates cyst growth by stimulating cAMP-dependent ERK activity and epithelial cell proliferation and by promoting Cl(-)-dependent fluid secretion. Tolvaptan, a V2 receptor antagonist, inhibits the renal effects of AVP and slows cyst growth in PKD animals. Here, we determined the effect of graded concentrations of tolvaptan on intracellular cAMP, ERK activity, cell proliferation, and transcellular Cl(-) secretion using human ADPKD cyst epithelial cells. Incubation of ADPKD cells with 10(-9) M AVP increased intracellular cAMP and stimulated ERK and cell proliferation. Tolvaptan caused a concentration-dependent inhibition of AVP-induced cAMP production with an apparent IC(50) of ∼10(-10) M. Correspondingly, tolvaptan inhibited AVP-induced ERK signaling and cell proliferation. Basolateral application of AVP to ADPKD cell monolayers grown on permeable supports caused a sustained increase in short-circuit current that was completely blocked by the Cl(-) channel blocker CFTR(inh-172), consistent with AVP-induced transepithelial Cl(-) secretion. Tolvaptan inhibited AVP-induced Cl(-) secretion and decreased in vitro cyst growth of ADPKD cells cultured within a three-dimensional collagen matrix. These data demonstrate that relatively low concentrations of tolvaptan inhibit AVP-stimulated cell proliferation and Cl(-)-dependent fluid secretion by human ADPKD cystic cells.

Global gene expression profiling in early-stage polycystic kidney disease in the Han:SPRD Cy rat identifies a role for RXR signaling.

Han:SPRD Cy is a spontaneous rat model of polycystic kidney disease (PKD) caused by a missense mutation in Pkdr1. Cystogenesis in this model is not clearly understood. In the current study, we performed global gene expression profiling in early-stage PKD cyst development in Cy/Cy kidneys and normal (+/+) kidneys at 3 and 7 days of postnatal age. Expression profiles were determined by microarray analysis, followed by validation with real-time RT-PCR. Genes were selected with over 1.5-fold expression changes compared with age-matched +/+ kidneys for canonical pathway analysis. We found nine pathways in common between 3- and 7-day Cy/Cy kidneys. Three significantly changed pathways were designated "Vitamin D Receptor (VDR)/Retinoid X Receptor (RXR) Activation," "LPS/IL-1-Mediated Inhibition of RXR Function," and "Liver X Receptor (LXR)/RXR Activation." These results suggest that RXR-mediated signaling is significantly altered in developing kidneys with mutated Pkdr1. In gene ontology analysis, the functions of these RXR-related genes were found to be involved in regulating cell proliferation and organ morphogenesis. With real-time RT-PCR analysis, the upregulation of Ptx2, Alox15b, OSP, and PCNA, major markers of cell proliferation associated with the RXR pathway, were confirmed in 3- and 7-day Cy/Cy kidneys compared with 3-day +/+ kidneys. The increased RXR protein was observed in both the nucleus and cytoplasm of cystic epithelial cells in early-stage Cy/Cy kidneys, and the RXR-positive cells were strongly positive for PCNA staining. Taken together, cell proliferation and organ morphogenesis signals transduced by RXR-mediated pathways may have important roles for cystogenesis in early-stage PKD in this Pkdr1-mutated Cy rat.

PPAR-gamma agonist ameliorates kidney and liver disease in an orthologous rat model of human autosomal recessive polycystic kidney disease.

In autosomal recessive polycystic kidney disease (ARPKD), progressive enlargement of fluid-filled cysts is due to aberrant proliferation of tubule epithelial cells and transepithelial fluid secretion leading to extensive nephron loss and interstitial fibrosis. Congenital hepatic fibrosis associated with biliary cysts/dilatations is the most common extrarenal manifestation in ARPKD and can lead to massive liver enlargement. Peroxisome proliferator-activated receptor γ (PPAR-γ), a member of the ligand-dependent nuclear receptor superfamily, is expressed in a variety of tissues, including the kidneys and liver, and plays important roles in cell proliferation, fibrosis, and inflammation. In the current study, we determined that pioglitazone (PIO), a PPAR-γ agonist, decreases polycystic kidney and liver disease progression in the polycystic kidney rat, an orthologous model of human ARPKD. Daily treatment with 10 mg/kg PIO for 16 wk decreased kidney weight (% of body weight), renal cystic area, serum urea nitrogen, and the number of Ki67-, pERK1/2-, and pS6-positive cells in the kidney. There was also a decrease in liver weight (% of body weight), liver cystic area, fibrotic index, and the number of Ki67-, pERK1/2-, pERK5-, and TGF-ß-positive cells in the liver. Taken together, these data suggest that PIO inhibits the progression of polycystic kidney and liver disease in a model of human ARPKD by inhibiting cell proliferation and fibrosis. These findings suggest that PPAR-γ agonists may have therapeutic value in the treatment of the renal and hepatic manifestations of ARPKD.

Sorafenib inhibits cAMP-dependent ERK activation, cell proliferation, and in vitro cyst growth of human ADPKD cyst epithelial cells.

In autosomal dominant polycystic kidney disease (ADPKD), aberrant proliferation of the renal epithelial cells is responsible for the formation of numerable fluid-filled cysts, massively enlarged kidneys, and progressive loss of renal function. cAMP agonists, including arginine vasopressin, accelerate cyst epithelial cell proliferation through protein kinase A activation of the B-Raf/MEK/extracellular signal-regulated kinase (ERK) signaling pathway. The mitogenic effect of cAMP is equally potent and additive to growth factor stimulation. Here, we determined whether Sorafenib (BAY 43-9006), a small molecule Raf inhibitor, inhibits proliferation of cells derived from the cysts of human ADPKD kidneys. We found that nanomolar concentrations of Sorafenib reduced the basal activity of ERK, inhibited cAMP-dependent activation of B-Raf and MEK/ERK signaling, and caused a concentration-dependent inhibition of cell proliferation induced by cAMP, epidermal growth factor, or the combination of the two agonists. Sorafenib completely blocked in vitro cyst growth of human ADPKD cystic cells cultured within a three-dimensional collagen gel. These data demonstrate that cAMP-dependent proliferation of human ADPKD cyst epithelial cells is blocked by Sorafenib and suggest that small molecule B-Raf inhibitors may be a therapeutic option to reduce the mitogenic effects of cAMP on cyst expansion.