UUO induces lung fibrosis with macrophage-myofibroblast transition in rats.

Progression of chronic kidney disease (CKD) to uremia is often accompanied by varying degrees of lung damage and this is also an important cause of death. Although there are many studies on the mechanism of lung injury, it is not clearly understood. Inflammatory macrophages may associated with fibrosis in the lungs. Here, we investigated the role of macrophage-myofibroblast transition (MMT) in lung fibrosis with unilateral ureteral obstruction (UUO) rats. We found that cells undergoing MMT accounted for an important part of the myofibroblast population, and correlated with lung fibrosis, MMT cells in lungs have a predominant M2 phenotype, and this process was attenuated after treatment with eplerenone. In conclusion, our studies provide a possible mechanism for UUO-induced kidney damage and lung injury, indicating the possibility of using eplerenone, a mineralocorticoid receptor blocker, to treat UUO to reduce kidney damage and protect lung function.

Dasatinib mitigates renal fibrosis in a rat model of UUO via inhibition of Src/STAT-3/NF-κB signaling.

This research aimed to investigate the reno-protective impact of the tyrosine kinase inhibitor dasatinib (DAS) against renal fibrosis induced by unilateral ureteral obstruction (UUO) in rats. DAS administration improved renal function and mitigated renal oxidative stress with paralleled reduction in the ligated kidney mass index, significant retraction in renal histopathological alterations and suppression of renal interstitial fibrosis. Nevertheless, DAS administration attenuated renal expression of phosphorylated Src (p-Src), Abelson (c-Abl) tyrosine kinases, nuclear factor-kappaB (NF-κB) p65, and phosphorylated signal transducer and activator of transcription-3 (p-STAT-3)/STAT-3 with paralleled reduction in renal contents of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and monocyte chemoattractant protein-1 (MCP-1). DAS diminished interstitial macrophage infiltration and decreased renal profibrotic transforming growth factor-ß1 (TGF-ß1) levels and suppressed interstitial expression of renal α-smooth muscle actin (α-SMA) and fibronectin. Collectively, DAS slowed the progression of renal interstitial fibrosis, possibly via attenuating renal oxidative stress, impairing Src/STAT-3/NF-κB signaling, and reducing renal inflammation.

Deletion of delta-like 1 homologue accelerates renal inflammation by modulating the Th17 immune response.

Preclinical studies have demonstrated that activation of the NOTCH pathway plays a key role in the pathogenesis of kidney damage. There is currently no information on the role of the Delta-like homologue 1 (DLK1), a NOTCH inhibitor, in the regulation of renal damage. Here, we investigated the contribution of DLK1 to experimental renal damage and the underlying molecular mechanisms. Using a Dlk1-null mouse model in the experimental renal damage of unilateral ureteral obstruction, we found activation of NOTCH, as shown by increased nuclear translocation of the NOTCH1 intracellular domain, and upregulation of Dlk2/hey-1 expression compared to wild-type (WT) littermates. NOTCH1 over-activation in Dlk1-null injured kidneys was associated with a higher inflammatory response, characterized by infiltration of inflammatory cells, mainly CD4/IL17A + lymphocytes, and activation of the Th17 immune response. Furthermore, pharmacological NOTCH blockade inhibited the transcription factors controlling Th17 differentiation and gene expression of the Th17 effector cytokine IL-17A and other related-inflammatory factors, linked to a diminution of inflammation in the injured kidneys. We propose that the non-canonical NOTCH ligand DLK1 acts as a NOTCH antagonist in renal injury regulating the Th17-mediated inflammatory response.

DAMPs in Unilateral Ureteral Obstruction.

Damage-associated molecular patterns (DAMPs) are released from tubular and interstitial cells in the kidney after unilateral ureteral obstruction (UUO). DAMPs are recognized by pattern recognition receptors (PRRs), which mediate the initiation of an immune response and the release of inflammatory cytokines. The animal model of UUO is used for various purposes. UUO in adult mice serves as a model for accelerated renal fibrosis, which is a hallmark of progressive renal disease. UUO in adult mice enables to study cell death, inflammation, and extracellular matrix deposition in the kidney. Neonatal UUO is a model for congenital obstructive nephropathies. It studies inflammation, apoptosis, and interstitial fibrosis in the neonatal kidney, when nephrogenesis is still ongoing. Following UUO, several DAMPs as well as DAMP receptors are upregulated. In adult UUO, soluble uric acid is upregulated and activates the NOD-like receptor family, pyrin domain containing-3 (NLRP3) inflammasome, which promotes fibrosis, apoptosis, and reactive oxygen species (ROS) injury. Further DAMPs associated with UUO are uromodulin, members of the IL-1 family, and necrotic cell DNA, all of which promote sterile inflammation. In neonatal UUO, the receptor for advanced glycation endproducts (RAGE) is highly upregulated. RAGE is a ligand for several DAMPs, including high mobility group box 1 (HMGB1) and S100 proteins, which play an important role in renal fibrosis. Additionally, necroptosis is an important mechanism of cell death, besides apoptosis, in neonatal UUO. It is highly inflammatory due to release of cytokines and specific DAMPs. The release and recognition of DAMPs initiate sterile inflammation, which makes them good candidates to develop and improve diagnostic and therapeutic strategies in renal fibrosis and congenital obstructive nephropathies.

SerpinB2 Regulates Immune Response in Kidney Injury and Aging.

BACKGROUND: Expression of SerpinB2, a regulator of inflammatory processes, has been described in the context of macrophage activation and cellular senescence. Given that mechanisms for these processes interact and can shape kidney disease, it seems plausible that SerpinB2 might play a role in renal aging, injury, and repair. METHODS: We subjected SerpinB2 knockout mice to ischemia-reperfusion injury or unilateral ureteral obstruction. We performed phagocyte depletion to study SerpinB2's role beyond the effects of macrophages and transplanted bone marrow from knockout mice to wild-type mice and vice versa to dissect cell type-dependent effects. Primary tubular cells and macrophages from SerpinB2 knockout and wild-type mice were used for functional studies and transcriptional profiling. RESULTS: Cultured senescent tubular cells, kidneys of aged mice, and renal stress models exhibited upregulation of SerpinB2 expression. Functionally, lack of SerpinB2 in aged knockout mice had no effect on the magnitude of senescence markers but associated with enhanced kidney damage and fibrosis. In stress models, inflammatory cell infiltration was initially lower in knockout mice but later increased, leading to an accumulation of significantly more macrophages. SerpinB2 knockout tubular cells showed significantly reduced expression of the chemokine CCL2. Macrophages from knockout mice exhibited reduced phagocytosis and enhanced migration. Macrophage depletion and bone marrow transplantation experiments validated the functional relevance of these cell type-specific functions of SerpinB2. CONCLUSIONS: SerpinB2 influences tubule-macrophage crosstalk by supporting tubular CCL2 expression and regulating macrophage phagocytosis and migration. In mice, SerpinB2 expression seems to be needed for coordination and timely resolution of inflammation, successful repair, and kidney homeostasis during aging. Implications of SerpinB2 in human kidney disease deserve further exploration.

Kidney injury enhances renal G-CSF expression and modulates granulopoiesis and human neutrophil CD177 in vivo.

Kidney injury significantly increases overall mortality. Neutrophilic granulocytes (neutrophils) are the most abundant human blood leukocytes. They are characterized by a high turnover rate, chiefly controlled by granulocyte colony stimulating factor (G-CSF). The role of kidney injury and uremia in regulation of granulopoiesis has not been reported. Kidney transplantation, which inherently causes ischemia-reperfusion injury of the graft, elevated human neutrophil expression of the surface glycoprotein CD177. CD177 is among the most G-CSF-responsive neutrophil genes and reversibly increased on neutrophils of healthy donors who received recombinant G-CSF. In kidney graft recipients, a transient rise in neutrophil CD177 correlated with renal tubular epithelial G-CSF expression. In contrast, CD177 was unaltered in patients with chronic renal impairment and independent of renal replacement therapy. Under controlled conditions of experimental ischemia-reperfusion and unilateral ureteral obstruction injuries in mice, renal G-CSF mRNA and protein expression significantly increased and systemic neutrophilia developed. Human renal tubular epithelial cell G-CSF expression was promoted by hypoxia and proinflammatory cytokine interleukin 17A in vitro. Clinically, recipients of ABO blood group-incompatible kidney grafts developed a larger rise in neutrophil CD177. Their grafts are characterized by complement C4d deposition on the renal endothelium, even in the absence of rejection. Indeed, complement activation, but not hypoxia, induced primary human endothelial cell G-CSF expression. Our data demonstrate that kidney injury induces renal G-CSF expression and modulates granulopoiesis. They delineate differential G-CSF regulation in renal epithelium and endothelium. Altered granulopoiesis may contribute to the systemic impact of kidney injury.

Delayed Rectifier K+-Channel Is a Novel Therapeutic Target for Interstitial Renal Fibrosis in Rats with Unilateral Ureteral Obstruction.

BACKGROUND: Delayed rectifier K+-channel, Kv1.3, is most predominantly expressed in T-lymphocytes and macrophages. In such leukocytes, Kv1.3-channels play pivotal roles in the activation and proliferation of cells, promoting cellular immunity. Since leukocyte-derived cytokines stimulate fibroblasts to produce collagen fibers in inflamed kidneys, Kv1.3-channels expressed in leukocytes would contribute to the progression of tubulointerstitial renal fibrosis. METHODS: Male Sprague-Dawley rats that underwent unilateral ureteral obstruction (UUO) were used at 1, 2, or 3 weeks after the operation. We examined the histological features of the kidneys and the leukocyte expression of Kv1.3-channels. We also examined the therapeutic effects of a selective channel inhibitor, margatoxin, on the progression of renal fibrosis and the proliferation of leukocytes within the cortical interstitium. RESULTS: In rat kidneys with UUO, progression of renal fibrosis and the infiltration of leukocytes became most prominent at 3 weeks after the operation, when Kv1.3-channels were overexpressed in proliferating leukocytes. In the cortical interstitium of margatoxin-treated UUO rat kidneys, immunohistochemistry revealed reduced expression of fibrosis markers. Additionally, margatoxin significantly decreased the numbers of leukocytes and suppressed their proliferation. CONCLUSIONS: This study clearly demonstrated that the numbers of T-lymphocytes and macrophages were markedly increased in UUO rat kidneys with longer postobstructive days. The overexpression of Kv1.3-channels in leukocytes was thought to be responsible for the proliferation of these cells and the progression of renal fibrosis. This study strongly suggested the therapeutic usefulness of targeting lymphocyte Kv1.3-channels in the treatment of renal fibrosis.

ATG5-mediated autophagy suppresses NF-κB signaling to limit epithelial inflammatory response to kidney injury.

G2/M-arrested proximal tubular epithelial cells (TECs) after renal injury are linked to increased cytokines production. ATG5-mediated autophagy in proximal TECs has recently been shown to protect against G2/M cell cycle arrest and renal fibrosis. However, the impacts of autophagy in regulating inflammatorily response mounted by injured TECs remains largely unknown. In the present study, we investigated whether ATG5 acts as an innate immune suppressor in proximal TECs during kidney injury. Using the unilateral ureteric obstruction model in proximal tubule-specific autophagy-deficient mice, we demonstrated that ablation of epithelial ATG5 genes markedly impaired autophagy, resulting in enhanced nuclear factor κB (NF-κB) activation, macrophage and lymphocyte infiltration, and proinflammatory cytokines production in obstructed kidneys, as compared with wild-type mice. Following stimulation with angiotensin II (Ang II), siRNA silencing of ATG5 in cultured HK-2 cells or ATG5-deficient primary proximal TECs produced more cytokines, including IL-1ß, IL-6, and TNF-α than did their control cells. Overexpressed ATG5, but not the autophagy-incompetent ATG5 mutant K130R in HK-2 cells, rendered resistant to Ang II-induced inflammatory response. Immunofluorescence assay indicated that ATG5 and p65 colocalized in the nucleus and cytoplasm, and their interaction was verified in immunoprecipitation assay from HEK-293T cell extracts. Genetic downregulation of endogenous ATG5 increased Ang II-induced phosphorylation and nuclear translocation of p65 and transcriptional activity of NF-κB, whereas the overexpressed ATG5, rather than ATG5 mutant K130R, hampered activation of NF-κB signaling, suggest an autophagy-dependent anti-inflammatory effect of ATG5. Further, pharmacological manipulation of autophagy yielded similar results both in vivo and in vitro. Additionally, JSH-23, a specific inhibitor of NF-κB nuclear translocation, rescued Ang II-driven IL-1ß production in ATG5 siRNA-treated cells and decreased the proportion of cells in G2/M phase. In conclusion, ATG5-mediated autophagy in tubules targets NF-κB signaling to protect against renal inflammation.

IL-33/ST2 axis mediates hyperplasia of intrarenal urothelium in obstructive renal injury.

The monolayered intrarenal urothelium covers the renal papilla and ureteropelvic junction (UPJ). In response to increased renal pressure during obstruction or ischemic injuries, intrarenal urothelial cells begin to proliferate and form a multilayered urothelium. Little is known regarding the mechanism and pathophysiological role of urothelium hyperplasia during renal obstruction. In this study, we investigated the expression of interleukin (IL)-33, an IL-1 family cytokine, in kidneys with unilateral ureteral obstruction (UUO)-induced obstructive injury. IL-33 levels in hydronephrotic urine and serum were upregulated 2 days after UUO. The number of ST2-expressing immune cells was increased in the UUO kidney. We found that IL-33 was upregulated in vimentin-positive cells in the cortical and medullar layers and the UPJ stroma. Moreover, IL-33 expression was predominantly induced in multilayered keratin 5-positive urothelial cells in the UPJ. IL-33 was not detected in terminally differentiated superficial umbrella cells expressing uroplakin 3a. In vivo, we confirmed that deficiency of IL33 or its receptor ST2 attenuated UUO-induced hyperplasia of the UPJ urothelium. Deficiency of IL33 attenuated the expression of UUO-induced type 2 inflammatory cytokines and upregulated uroplakins and urothelial differentiation signaling in UPJ tissues. Our results collectively suggest that the IL-33/ST2 axis mediates the activation of innate immune responses and contributes to urothelial hyperplasia by regulating urothelial differentiation in obstructive kidney injury.

Fluorofenidone attenuates interleukin-1ß production by interacting with NLRP3 inflammasome in unilateral ureteral obstruction.

AIM: We explored whether Fluorofenidone reduced interleukin-1ß (IL-1ß) production by interacting with NLRP3 inflammasome in unilateral ureteral obstruction (UUO). METHODS: Ureteral obstruction rats were treated with Fluorofenidone (500 mg/kg per day) for 3, 7 days. Morphologic analysis and leukocytes infiltration were assessed in ligated kidneys. Furthermore, plasmids of NLRP3, ASC, pro-Caspase-1, pro-IL-1ß were co-transfected into 293 T cells, and then treated with Fluorofenidone (2 mM). The expression of NLRP3, ASC, pro-caspase-1, cleavage caspase-1, pro-IL-1ß and cleavage IL-1ß were measured by Western blot or real-time PCR in vivo and in vitro. Moreover the interaction of NLRP3 inflammasome-assembly was detected by co-immunoprecipitation and confocal immunofluorescence. RESULTS: Fluorofenidone treatment significantly attenuated renal fibrosis and leukocytes infiltration in UUO model. Fluorofenidone had no effect on the expression of pro-IL-1ß. Interestingly, Fluorofenidone inhibited the activation of NLRP3 inflammasome, downregulated Caspase-1 levels and thereby decreased the cleavage of pro-IL-1ß into IL-1ß in vivo and in vitro. Fluorofenidone treatment distinctively weakened the interaction between NLRP3 and ASC, as well as ASC and pro-Caspase-1 in vivo. However, Fluorofenidone treatment only significantly weakened the interaction between ASC and pro-Caspase-1 in co-transfected 293 T cells. CONCLUSION: Fluorofenidone serves as a novel anti-inflammatory agent that attenuates IL-1ß production in UUO model by interacting with NLRP3 inflammasome.

FTY720 ameliorates renal fibrosis by simultaneously affecting leucocyte recruitment and TGF-ß signalling in fibroblasts.

Renal fibrosis is the common final manifestation of chronic kidney diseases and usually results in end-stage renal failure. In this study, we evaluated the effect of fingolimod (FTY720), an analogue of sphingosine 1-phosphate (S1P), as a treatment for the unilateral ureteral obstruction (UUO)-induced renal fibrosis animal model. We treated mice with FTY720 at a dosage of 1 mg/kg/day by intragastric administration from day 1 until day 7. The control group received the same amount of saline. FTY720 reduced significantly the urine albumin/creatinine ratio (UACR) in treated UUO mice. FTY720 treatment also caused a significant decrease in interstitial expansion and collagen deposition in the kidney, accompanied by reduced mononuclear cell recruitment and inflammatory cytokine expression. In addition, the expression levels of the endothelial cell adhesion molecules P-selectin and vascular cell adhesion protein 1 (VCAM-1) were suppressed in the ligated kidney by FTY720 administration, suggesting reduced renal endothelial cell activation. Furthermore, in renal interstitial fibroblast normal rat kidney (NRK)-49F cells, FTY720 significantly affected transforming growth factor (TGF)-ß-induced α-smooth muscle actin (SMA) expression and collagen synthesis by inhibiting both the Mothers against decapentaplegic homologue (Smad)2/3 and phosphatidylinositol 3-kinase/protein kinase B/glycogen synthase kinase 3 beta (PI3K/AKT/GSK3ß) signalling pathways. S1P1 knock-down by siRNA reversed this effect significantly in our fibroblast cell culture model. Therefore, FTY720 attenuates renal fibrosis via two different mechanisms: first, FTY720 suppresses the synthesis of extracellular matrix in interstitial fibroblasts by interfering with TGF-ß signalling; and secondly, FTY720 affects endothelial cell activation and chemokine expression, thereby reducing immune cell recruitment into the kidney.

Generation and characterization of an antagonistic monoclonal antibody against an extracellular domain of mouse DP2 (CRTH2/GPR44) receptors for prostaglandin D2.

Prostaglandin D2 (PGD2) is a lipid mediator involved in sleep regulation and inflammation. PGD2 interacts with 2 types of G protein-coupled receptors, DP1 and DP2/CRTH2 (chemoattractant receptor homologous molecule expressed on T helper type 2 cells)/GPR44 to show a variety of biological effects. DP1 activation leads to Gs-mediated elevation of the intracellular cAMP level, whereas activation of DP2 decreases this level via the Gi pathway; and it also induces G protein-independent, arrestin-mediated cellular responses. Activation of DP2 by PGD2 causes the progression of inflammation via the recruitment of lymphocytes by enhancing the production of Th2-cytokines. Here we developed monoclonal antibodies (MAbs) against the extracellular domain of mouse DP2 by immunization of DP2-null mutant mice with DP2-overexpressing BAF3, murine interleukin-3 dependent pro-B cells, to reduce the generation of antibodies against the host cells by immunization of mice. Moreover, we immunized DP2-KO mice to prevent immunological tolerance to mDP2 protein. After cell ELISA, immunocytochemical, and Western blot analyses, we successfully obtained a novel monoclonal antibody, MAb-1D8, that specifically recognized native mouse DP2, but neither human DP2 nor denatured mouse DP2, by binding to a particular 3D receptor conformation formed by the N-terminus and extracellular loop 1, 2, and 3 of DP2. This antibody inhibited the binding of 0.5 nM [3H]PGD2 to mouse DP2 (IC50 = 46.3 ± 18.6 nM), showed antagonistic activity toward 15(R)-15-methyl PGD2-induced inhibition of 300 nM forskolin-activated cAMP production (IC50 = 16.9 ± 2.6 nM), and gave positive results for immunohistochemical staining of DP2-expressing CD4+ Th2 lymphocytes that had accumulated in the kidney of unilateral ureteral obstruction model mice. This monoclonal antibody will be very useful for in vitro and in vivo studies on DP2-mediated diseases.

Adipose-derived stem cells ameliorate renal interstitial fibrosis through inhibition of EMT and inflammatory response via TGF-ß1 signaling pathway.

Adipose-derived stem cells (ADSCs) have been successfully used to treat acute kidney injury or acute renal failure. However, the effect of ADSCs on treating renal interstitial fibrosis remains unknown. Here, we assessed the therapeutic efficacy of ADSCs on renal interstitial fibrosis induced by unilateral ureter obstruction (UUO) and explored the potential mechanisms. After 7days of UUO, rats were injected with ADSCs (5×106) or vehicle via tail vein. We found that ADSCs administration significantly ameliorated renal interstitial fibrosis, the occurrence of epithelial-mesenchymal transition (EMT) and inflammatory response. Furthermore, ADSCs administration could inhibit the activation of transforming growth factor-ß1 (TGF-ß1) signaling pathway, which might play a crucial role in renal interstitial fibrosis of the UUO model rats. These results suggested that ADSCs treatment attenuates renal interstitial fibrosis possibly through inhibition of EMT and inflammatory response via TGF-ß1 signaling pathway. Therefore, ADSCs may be an effective therapeutic strategy for the treatment of renal interstitial fibrosis.

Renal recruitment of B lymphocytes exacerbates tubulointerstitial fibrosis by promoting monocyte mobilization and infiltration after unilateral ureteral obstruction.

Renal fibrosis is a significant threat to public health globally. Diverse primary aetiologies eventually result in chronic kidney disease (CKD) and immune cells influence this process. The roles of monocytes/macrophages, T cells, and mast cells have been carefully examined, whilst only a few studies have focused on the effect of B cells. We investigated B-cell function in tubulointerstitial fibrosis induced by unilateral ureteral obstruction (UUO), using genetic B-cell-deficient µMT mice or CD20 antibody-mediated B-cell-depleted mice. Obstructed kidneys of µMT and anti-CD20-treated mice showed lower levels of monocyte/macrophage infiltration and collagen deposition compared to wild-type mice. Mechanistically, anti-CD20 attenuated UUO-induced alterations of renal tumour necrosis factor-α (TNF-α), vascular cell adhesion molecule 1 (VCAM-1) pro-inflammatory genes, and CC chemokine ligand-2 (CCL2) essential for monocyte recruitment; B cells were one of the main sources of CCL2 in post-UUO kidneys. Neutralization of CCL2 reduced monocyte/macrophage influx and fibrotic changes in obstructed kidneys. Therefore, early-stage accumulation of B cells in the kidney accelerated monocyte/macrophage mobilization and infiltration, aggravating the fibrosis resulting from acutely induced kidney nephropathy. © 2016 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Danggui Buxue Tang Attenuates Tubulointerstitial Fibrosis via Suppressing NLRP3 Inflammasome in a Rat Model of Unilateral Ureteral Obstruction.

Inflammation significantly contributes to the progression of chronic kidney disease (CKD). This study aimed to characterize Danggui Buxue Tang (DBT) renoprotection and relationship with NOD-like receptors family pyrin domain-containing 3 (NLRP3) inflammasome expression in rats with unilateral ureteral obstruction (UUO). Sprague-Dawley rats were subjected to UUO and randomly assigned to untreated UUO, enalapril-treated (10 mg/kg/day), and DBT-treated (9 g/kg/day) groups. Sham-operated rats served as controls, with 8 rats in each group. All rats were sacrificed for blood and renal specimen collection at 14 days after UUO. Untreated UUO rats exhibited azotemia, intense tubulointerstitial collagen deposition, upregulations of tubulointerstitial injury index, augmentation levels of collagen I (Col I), α-smooth muscle actin (α-SMA), NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), pro-caspase-1, caspase-1, IL-1ß, and pro-IL-1ß. DBT treatment significantly attenuated interstitial collagen deposition and tubulointerstitial injury, lowering Col I and α-SMA levels. Synchronous expressions of NLRP3, ASC, pro-caspase-1, caspase-1, pro-IL-1ß, and IL-1ß decreased in renal tissue. In comparison to enalapril, DBT significantly reduced tubulointerstitial injury, interstitial collagen deposition, and expressions of Col I and IL-1ß. Thus, DBT offers renoprotection in UUO rats, which was associated with suppressing NLRP3 inflammasome expression and following reduction of the secretion of cytokine IL-1ß. The mechanisms of multitargets of traditional Chinese medicine can be better used for antifibrotic treatment.

Sorafenib Inhibits Renal Fibrosis Induced by Unilateral Ureteral Obstruction via Inhibition of Macrophage Infiltration.

AIMS: Sorafenib, which has been used extensively for the treatment of renal cell cancer and advanced hepatocellular carcinoma (HCC), has also been shown to have antifibrotic effects in liver fibrosis. However, the effects of sorafenib on renal fibrosis are unknown. Therefore, we investigated whether sorafenib inhibited renal fibrosis in a mouse model of unilateral ureteral obstruction (UUO) and further explored the potential mechanism. METHODS: Mice underwent UUO followed by vehicle or sorafenib treatment. The expression of CD68, a macrophage marker, and the pro-inflammatory cytokines, MCP1 and CXCR3, were immunohistochemically analyzed. The involvement of macrophages in the formation of renal fibrosis was studied using confocal microscopy. RESULTS: Renal histopathology improved in the UUO-sorafenib mice. Sorafenib notably suppressed TGF-ß1-mediated renal fibrogenic effects. The mRNA and protein expressions of CD68, MCP1, and CXCR3 in the obstructed kidney were significantly decreased by sorafenib. Immunohistochemistry showed that CD68 and CXCR3 had a similar distribution, whereas MCP1 was observed predominantly in the tubular epithelial cells. Double immunofluorescence demonstrated that CD68-positive macrophages could co-localize with CXCR3. It also revealed that CXCR3 interacted with CXCL11 in the UUO mouse kidneys. Widespread adhesion of macrophages to myofibroblasts was markedly inhibited in UUO-sorafenib mouse kidneys. CONCLUSIONS: Taken together, the results indicated that sorafenib had protective effects against renal fibrosis; its mechanism of action was associated with inhibition of macrophage infiltration via the CXCR3/CXCL11 pathway. These data suggest the clinical potential of sorafenib for treatment of renal fibrosis and illustrate the immunological mechanisms underlying the protective effects of sorafenib.

Antagonism of scavenger receptor CD36 by 5A peptide prevents chronic kidney disease progression in mice independent of blood pressure regulation.

Scavenger receptor CD36 participates in lipid metabolism and inflammatory pathways important for cardiovascular disease and chronic kidney disease (CKD). Few pharmacological agents are available to slow the progression of CKD. However, apolipoprotein A-I-mimetic peptide 5A antagonizes CD36 in vitro. To test the efficacy of 5A, and to test the role of CD36 during CKD, we compared wild-type to CD36 knockout mice and wild-type mice treated with 5A, in a progressive CKD model that resembles human disease. Knockout and 5A-treated wild-type mice were protected from CKD progression without changes in blood pressure and had reductions in cardiovascular risk surrogate markers that are associated with CKD. Treatment with 5A did not further protect CD36 knockout mice from CKD progression, implicating CD36 as its main site of action. In a separate model of kidney fibrosis, 5A-treated wild-type mice had less macrophage infiltration and interstitial fibrosis. Peptide 5A exerted anti-inflammatory effects in the kidney and decreased renal expression of inflammasome genes. Thus, CD36 is a new therapeutic target for CKD and its associated cardiovascular risk factors. Peptide 5A may be a promising new agent to slow CKD progression.

Involvement of cytokines in the modulation and progression of renal fibrosis induced by unilateral ureteral obstruction in C57BL/6 mice: effects of thalidomide and dexamethasone.

This study investigated the kinetics of cytokines that are involved in the development of interstitial fibrosis in mice that were subjected to UUO, the interstitial type I and III collagen deposition, and the effects of Thalido and Dexa treatment on these parameters. Inbred C57BL/6 mice were divided into the groups: Normal (not submitted surgery), Sham (sham surgery), Control (UUO treated with 0.5% carboxymethyl cellulose), Thalido (UUO treated with 5 mg/kg thalidomide), and Dexa (UUO treated with 1 mg/kg dexamethasone). The treatments began the day before surgery and were administered once daily by gavage for 1, 7, or 14 days. At the end of each treatment period, blood samples were collected for the determination of creatinine, urea, cytokines. The Control group exhibited a increase in creatinine concentration compared with the Normal and Sham groups within the first 24 h after UUO, which remained high until days 7 and 14. The urea concentration was higher on days 7 and 14 in the Control group compared with the Sham group. In the Thalido and Dexa groups, a reduction of serum creatinine concentration was seen on day 14. Treatment with Dexa reduced the serum concentration of urea on day 7. The serum concentrations of cytokines (TNF-α, IL-1ß, IL-6, IL-10 and IL-17) and chemokines (KC, MIG, bFGF) increased in UUO mice at all of the sampling times. The Dexa and Thalido groups exhibited alterations in the concentrations of these cytokines, suggesting the involvement of anti-inflammatory and immunomodulatory mechanisms that may have modified the fibrosis framework.

Effect of salvianolic acid A and C compatibility on inflammatory cytokines in rats with unilateral ureteral obstruction.

OBJECTIVE: To investigate the effect of salvianolic acid A and C component molecules, which are involved in drug compatibility, on inflammatory cytokine expression that affects human chemokine ligand 5 (CCL5) and chemokine ligand 10 (CXCL10) levels in rats with unilateral ureteral obstruction (UUO). METHODS: Fifty Sprague Dawley rats were randomly divided into five groups: normal, model, salvianolic acid A, salvianolic acid C and salvianolic acid A and C groups. The normal group was used as the control, and the other groups of rats had a UUO model established. The control group had free access to food and water, and the other groups received the corresponding drugs for 2 weeks. After the last administration, urine ß2-microglobulin (ß 2-MG) and N-acetyl-ß-D-glucosaminidase (NAG) levels were analyzed. After 24 h, all rats were sacrificed and the serum was analyzed for creatinine (Cr) and blood urea nitrogen (BUN) levels. Rat kidneys were removed, and CCL5 and CXCL10 inflammatory cytokine mRNA expression was measured using real-time fluorescent quantitative reverse transcription-polymerase chain reaction (RT-PCR). Kidney fibrosis was observed using hematoxylin-eosin (HE) staining and Masson trichrome staining. RESULTS: In the salvianolic acid A and salvianolic acid C treatment groups, serum Cr and urine NAG levels were significantly lower than in the model group (both P < 0.05). In all treatment groups, urine ß2-MG levels were significantly lower than in the model group (all P < 0.05). Compared with model group, the pathological changes and collagen deposition improved to varying degrees (both P < 0.05). CCL5 and CXCL10 mRNA expression decreased to different degrees compared with the model group (both P < 0.05). CONCLUSION: Salvianolic acid A and C are component molecules of drug compatibility, and they may protect renal function and improve tubular function and renal pathology to a certain degree in UUO. This improvement may be related to a reduction in inflammatory cytokines CCL5 and CXCL10 secretion in the UUO rat kidney.