Renal Inflammatory Infiltration Research Articles

HBx promotes glomerular podocyte-induced immune cell responses

Background Podocytes, as intrinsic renal cells, can also express MHC-II and costimulatory molecules under inflammatory conditions, suggesting that they may act as antigen-presenting cells (APCs) to activate immune cell responses and then lead to immune-mediated renal injury. They are already recognized as main targets in the pathogenic mechanism of hepatitis B virus (HBV)-associated glomerulonephritis (HBV-GN). Previous studies also have indicated that inflammatory cells infiltration and immune-mediated tissue injury are evident in the kidney samples of patients with HBV-GN. However, the role of podocytes immune disorder in the pathogenic mechanism of HBV-GN remains unclear. Methods Renal function and inflammatory cells infiltration were measured in HBV transgenic (HBV-Tg) mice. In vitro, podocytes/CD4+ T cells or macrophages co-culture system was established. Then, the expression of HBx, CD4, and CD68 was determined by immunohistochemistry, while the expression of MHC-II, CD40, and CD40L was determined by immunofluorescence. Co-stimulatory molecules expression was examined by flow cytometry. The levels of inflammatory factors were detected by ELISA. Results In vivo, renal function was obviously impaired in HBV-Tg mice. HBx was significantly upregulated and immune cells infiltrated in the glomerulus of HBV-Tg mice. Expression of MHC-II and costimulatory molecule CD40 increased in the podocytes of HBV-Tg mice; CD4+ T cells exhibited increased CD40L expression in glomerulus. In vitro, CD40 expression was markedly elevated in HBx-podocytes. In co-culture systems, HBx-podocytes stimulated CD4+ T cells activation and caused the imbalance between IFN-γ and IL-4. HBx-podocytes also enhanced the adhesion ability of macrophages and induced the release of proinflammatory mediators. Conclusion Taken together, these podocyte-related immune disorder may be involved in the pathogenic mechanism of HBV-GN.

Glomerular Elasticity and Gene Expression Patterns Define Two Phases of Alport Nephropathy.

To understand the early stages if Alport nephropathy, we characterize the structural, functional, and biophysical properties of glomerular capillaries and podocytes in Col4α3 -/- mice, analyze kidney cortex transcriptional profiles at three time points, and investigate the effects of the ER stress mitigation by TUDCA on these parameters. We use human FSGS associated genes to identify molecular pathways rescued by TUDCA. We define a disease progression timeline in Col4α3 -/- mice. Podocyte injury is evident by 3 months, with glomeruli reaching maximum deformability at 4 months, associated with 40% podocytes loss, followed by progressive capillary stiffening, increasing proteinuria, reduced renal function, inflammatory infiltrates, and fibrosis from months 4 to 7. RNA sequencing at 2, 4, and 7 months reveals increased cytokine and chemokine signaling, matrix and cell injury, and activation of the TNF pathway genes by 7 months, similar to NEPTUNE FSGS cohorts. These features are suppressed by TUDCA. We define two phases of Col4α3 -/- nephropathy. The first is characterized by podocytopathy, increased glomerular capillary deformability and accelerated podocyte loss, and the second by increased capillary wall stiffening and renal inflammatory and profibrotic pathway activation. Disease suppression by TUDCA treatment identifies potential therapeutic targets for treating Alport and related nephropathies.

Hedysarum polybotrys polysaccharide attenuates renal inflammatory infiltration and fibrosis in diabetic mice by inhibiting the HMGB1/RAGE/TLR4 pathway.

Diabetic kidney disease (DKD) is a leading cause of kidney failure. Previous studies demonstrated the therapeutic potential of Astragalus polysaccharide in treating diabetic nephropathy. Astragalus and Hongqi both come from the leguminous plant Astragalus, but their species and genera are different, belonging to the same family and different genera of traditional Chinese medicinal plants. However, the effects of Hedysarum polybotrys polysaccharide (HPS), a polysaccharide compound from Hongqi, on DKD, including its components and efficacy, have remained elusive. The present study utilized db/db mice as a DKD animal model administered with low (30 mg/kg) and high doses (60 mg/kg) of HPS, in addition to glyburide (7.2 mg/kg). Blood and urine samples were collected from mice and blood glucose, serum creatinine, urinary albumin excretion and urinary β2-microglobulin were measured. In addition, apoptosis and histological changes in kidney tissue were observed using TUNEL and HE staining, respectively, and the secretion and expression of inflammatory factors in kidney tissue were detected using EILSA and reverse transcription-quantitative PCR. Furthermore, we the expression of fibrosis-related proteins and NF-κB signaling pathway proteins was determined using western blot analysis. HPS was found to reduce the blood glucose concentration, serum creatinine levels, urinary albumin excretion rates and urinary β2-microglobulin in a dose-dependent manner. In addition, HPS treatment mitigated apoptosis and pathological damage in the kidney tissues of DKD mice. The expression levels of fibrosis-related proteins fibronectin, α-smooth muscle actin and TGF-β1 were observed to be decreased in kidney tissues of DKD mice following HPS treatment. The secretion levels of inflammatory factors (IL-6, TNF-α and IL-1β) were also reduced in kidney tissues, with high-dose HPS treatment found to be more effective, similar to the effects mediated by the glyburide. Further mechanistic analysis revealed that the therapeutic effects of HPS on DKD mice may be mediated by inhibiting the high mobility group box 1/receptor for advanced glycation end-products/toll-like receptor 4 pathway. In conclusion, the present findings could provide insight for the treatment of DKD.

Astragaloside IV improves renal function and alleviates renal damage and inflammation in rats with chronic glomerulonephritis.

From Astragalus membranaceus (Fisch.) Bge.var. mongholicus (Bge.) Hsiao, astragaloside IV (AS-IV), a saponin can be purified and is considered traditional Chinese medicine. The purpose of this study was to evaluate the AS-IV-mediated mechanism on chronic glomerulonephritis (CGN). A cationic bovine serum albumin-induced CGN rat model was established and 10, 15, or 20 mg/kg of AS-IV was administered to measure renal function and inflammatory infiltration. Influences of AS-IV on proliferation, cell cycle, and inflammation of LPS-induced rat mesangial cells (RMCs) were determined. The results demonstrated that AS-IV alleviated renal dysfunction, renal lesions, and inflammation in CGN rats. AS-IV prolonged the G0-G1 phase, shortened the S phase, and inhibited cell proliferation and inflammation in RMCs. AS-IV can promote miR-181d-5p expression to inhibit CSF1. miR-181d-5p promotion or CSF1 suppression could further enhance the therapeutic role of AS-IV in CGN rats, while miR-181d-5p silencing or CSF1 overexpression abolished the effect of AS-IV. In conclusion, AS-IV by mediating the miR-181d-5p/CSF1 axis protects against CGN.

Oxybaphus himalaicus Mitigates Lipopolysaccharide-Induced Acute Kidney Injury by Inhibiting TLR4/MD2 Complex Formation.

Acute kidney injury (AKI) is described as the abrupt decrease in kidney function always accompanied by inflammation. The roots of Oxybaphus himalaicus Edgew. have long been used in Tibetan folk medicine for the treatment of nephritis. Nevertheless, modern pharmacological studies, especially about the underlying mechanism of O. himalaicus medications, are still lacking. Here, in lipopolysaccharide (LPS)-induced RAW264.7 macrophages, the O. himalaicus extract (OE) showed significant anti-inflammatory activity with the dose dependently reducing the LPS-stimulated release of nitric oxide and the mRNA level and protein expression of inflammatory cytokines and reversed the activation of nuclear factor kappa B (NF-κB). Co-immunoprecipitation assay indicated that OE inhibited Toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD2) complex formation and further suppressed both myeloid differentiation factor 88 (MyD88)-dependent and TIR-domain-containing adapter-inducing interferon-β (TRIF)-dependent cascades activation. In addition, OE could restrain NADPH oxidase 2 (NOX2) endocytosis by blocking TLR4/MD2 complex formation to prevent reactive oxygen species production. In LPS-induced AKI mice, OE treatment mitigated renal injury and inflammatory infiltration by inhibiting TLR4/MD2 complex formation. UPLC-MS/MS analysis tentatively identified 41 components in OE. Our results indicated that OE presented significant anti-inflammatory activity by inhibiting TLR4/MD2 complex formation, which alleviated LPS-induced AKI in mice.

Ivabradine treatment lowers blood pressure and promotes cardiac and renal protection in spontaneously hypertensive rats

Hypertension is linked to hyperpolarization-activated cyclic nucleotide-gated (HCN) function, expressed in excitable and non-excitable cells. Considering that the reduction in heart rate (HR) improves coronary perfusion and cardiac performance, ivabradine (IVA) emerged as an important drug for the treatment of cardiovascular diseases. AimEvaluate if IVA chronic treatment effect can mitigate hypertension and reverse the cardiac and renal damage in SHR. Main methodsRats were divided into 4 groups treated for 14 days with PBS (1 ml/kg; i.p) or IVA (1 mg/kg; i.p): 1) WKY PBS; 2) SHR PBS; 3) WKY IVA; and 4) SHR IVA. The systolic blood pressure (SBP) was measured, indirectly, before and during the treatment period with IVA (day 0, 1, 7 and 11). Rats were subjected to artery cannulation for direct blood pressure (BP) measurement. Morphofunctional and gene expression were evaluated in the heart and kidneys. Key findingsIVA reduced SBP only in SHR on the 7th day. Direct blood pressure measurement showed that IVA chronic treatment reduced HR in the SHR. Interestingly, mean arterial pressure (MAP) was reduced in SHR IVA when compared to SHR PBS. Serum and urinary biochemical data were not altered by IVA. Moreover, IVA reduced the renal inflammatory infiltrates and increased glomerular density, besides preventing the cardiac inflammatory and hypertrophic responses. SignificanceIVA treatment lowered blood pressure, improved cardiac remodeling and inflammation, as well as decreasing renal damage in SHR. Further, IVA increased renal HCN2 mRNA and reduced cardiac HCN4 mRNA.

Silencing long noncoding RNA-CES1P1 suppresses glomerular endothelial cell inflammation in diabetic nephropathy

Diabetic nephropathy (DN) has become the main cause of end-stage renal disease worldwide. Inflammation is associated with the occurrence and development of DN, and long noncoding RNAs (lncRNAs) are involved in the regulation of inflammatory processes. This study aims to determine the role and mechanism of lncRNA-CES1P1 in DN.C57BL/6 mice and human umbilical vein endothelial cells (HUVECs) were used for this experimental study. In vivo experimental intraperitoneal injection of streptozotocin (STZ) to construct a diabetes mellitus (DM) model in C57BL/6 mice caused increased expression of lncRNA-CES1P1, decreased expression of miR-214-3p in kidney tissue, and produced renal inflammation and proteinuria. Exogenous knockdown of lncRNA-CES1P1 expression decreased renal inflammatory infiltration. In vitro experiments using high glucose (HG) stimulation of HUVECs cell revealed increased expression of lncRNA-CES1P1, decreased expression of miR-214-3p, and increased expression of the inflammatory factors IL-17, IκB, NF-κB, and IL-6. Luciferase reporter assays showed direct targets of miR-214-3p interaction with lncRNA-CES1P1 and IL-17. These results suggest that hyperglycemia represses miR-214-3p by inducing lncRNA-CES1P1, which promotes the expression of the inflammatory factors IL-17, IκB, NF-κB and IL-6 ultimately leading to the development of DN. Interfering with lncRNA-CES1P1 can reduce hyperglycemia-induced DN.

RP105 protects against ischemic and septic acute kidney injury via suppressing TLR4/NF-κB signaling pathways

Acute kidney injury (AKI) is a critical and severe clinical disease caused by a variety of factors. Toll-like receptors (TLRs) play a crucial role in pathogenesis of AKI. Radioprotective 105 kDa protein (RP105) is a member of the TLR family, but the role of RP105 in AKI is unknown. In this study, we overexpressed RP105 in renal tissue and cultured proximal tubular cells in which we then induced ischemic and septic AKI. Renal structure injuries were examined by hematoxylin eosin staining, while renal function was assessed by measuring serum blood urea nitrogen (BUN) and creatinine (SCr) levels. The TUNEL assay was used to detect apoptosis induced changes in the expression of RP105, and nuclear factor κB (NF-κB) in renal tissue detected by Western blot. Inflammatory cytokines including iNOS, IL-1β, IL-6, and TNF-α were detected by quantitative real-time PCR. The inflammatory indicators, F4/80 and MPO, were identified by IHC staining. The results showed that expression of the TLR4/NF-kB signaling pathway was enhanced in renal ischemia–reperfusion injury and septic renal injury, and that overexpression of RP105 in renal tissue alleviated ischemic and septic AKI. Moreover, RP105 gene delivery was associated with reduced renal inflammatory cells infiltration and inflammatory cytokines after AKI. RP105 overexpression also inhibited nuclear translocation of NF-κB after AKI in both in vitro and in vivo, and blunted the interaction between Myeloid Differentiation factor 2 (MD2) and TLR4. These results indicated that RP105 protected against renal ischemic and septic AKI injury by suppressing inflammatory responses mediated by TLR4 signaling pathways. This study suggests that the anti-inflammatory roles of RP105 have potential for preventing and treating renal ischemic and septic AKI.

Inhibition of microRNA-448 suppresses CD4+ T cell inflammatory activation via up-regulating suppressor of cytokine signaling 5 in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease. MicroRNA-448 (miR-448) has a pro-inflammatory effect in various inflammation-related diseases and is up-regulated in serum of patients with SLE. However, the role of miR-448 in SLE development remains elusive. In our study, we found high expression of miR-448 in peripheral blood mononuclear cells (PBMCs) of SLE patients, and miR-448 level was positively associated with disease severity. Besides, miR-448 level was up-regulated during the growth of MRL/lpr mice. To investigate the function of miR-448 in SLE, we subjected 8-week MRL/lpr mice to injection of lentivirus (LV)-mediated anti-miR-448. Inhibition of miR-448 reduced serum IgG and anti-dsDNA IgG contents, 24 h urine protein and blood urea nitrogen (BUN) levels, increased complement C3 concentration, and ameliorated splenomegaly and lymphadenectasis in MRL/lpr mice. MiR-448 inhibition alleviated renal inflammatory infiltration and glycogen deposition. Moreover, miR-448 inhibition promoted Treg cell activation and inhibited Th17 cell proportion in naïve CD4+ T cells from spleens, along with elevated interleukin (IL)-10 and reduced IL-17A levels. In vitro, miR-448 inhibition diminished CD4+ T cell polarization toward Th17 cells under Th17-polarizing conditions. Further, luciferase reporter assay revealed that miR-448 binds to the 3′UTR of suppressor of cytokine signaling 5 (SOCS5). SOCS5 expression was down-regulated in the spleens of MRL/lpr mice and induced Th17 cells. SOCS5 deficiency partially reversed the role of miR-448 in Th17 differentiation and IL-17A expression in SLE. Taken together, inhibition of miR-448 impedes Th17 cell activation and tissue damages via targeting SOCS5 in SLE.

Sodium-glucose cotransporter 2 inhibition prevents renal fibrosis in cyclosporine nephropathy

AimsSodium-glucose cotransporter 2 (SGLT2) inhibitors, a new class of antidiabetic drugs, are nephroprotective in case of diabetes, but whether a similar beneficial effect may be detectable also in case of chronic non-diabetic kidney diseases remains still unknown. The aim of this study was to evaluate the effects of empagliflozin, a SGLT-2 inhibitor, on the progression of cyclosporine nephropathy, in the absence of diabetes.MethodsSprague Dawley rats (n = 27) have been fed with low-salt diet starting 10 days before the beginning and finished at the end of the experimental period. Cyclosporine-A (CsA, 15 mg/kg/day, intraperitoneal injection, n = 8) and CsA plus empagliflozin (Empa, 10 mg/kg/day, per os, n = 7) were administered for 4 weeks. The control groups were treated with placebo (Control, n = 7) or empagliflozin (Control + Empa, n = 5). Blood pressure (plethysmographic method) was measured at the beginning and at the end of the experimental period. At the end of the experimental protocol, the kidneys were excised for histomorphometric analysis of renal fibrosis and for immunohistochemical evaluation of inflammatory infiltrates (monocytes/macrophages), type I and type IV collagen expression, and tyrosine hydroxylase expression, used as marker of sympathetic nerve activity.ResultsCsA-treated rats showed a significant increase (p < 0.01) in blood pressure, which was reduced by administration of empagliflozin (p < 0.05). CsA administration caused an increase in glomerular and tubulo-interstitial fibrosis (p < 0.05), renal inflammatory infiltrates (p < 0.05), type I and type IV collagen expression (p < 0.01), and tyrosine hydroxylase expression (p < 0.01) as compared to the control rats and control + Empa-treated rats. Treatment with empagliflozin in CsA-treated rats reduced glomerular (p < 0.01) and tubulo-interstitial fibrosis (p < 0.05), type I and type IV collagen expression (p < 0.01), inflammatory cell infiltration (p < 0.01) and tyrosine hydroxylase expression (p < 0.05), as compared to rats treated with CsA.ConclusionEmpagliflozin administration caused a reduction in blood pressure in CsA-treated rats and showed a protective effect on CsA nephropathy by decreasing renal fibrosis, type I and type IV collagen expression, macrophage infiltration and tyrosine hydroxylase expression. These data suggest that empagliflozin promotes nephroprotection also in non-diabetic kidney disease.

The profiles of biopsy-proven renal tubulointerstitial lesions in patients with glomerular disease.

BackgroundRenal tubules and interstitium are vulnerable to injury and play a central role in the progression of various chronic kidney diseases (CKDs). However, high quality epidemiologic study on the profiles of biopsy-proven tubulointerstitial lesions (TILs) is extremely limited.MethodsWe conducted a retrospective renal biopsy series including 62,569 native biopsies at 1,211 hospitals across China from 2015 to 2017. The TILs, including the shedding of tube epithelial, renal tubular atrophy, renal interstitial fibrosis, edema and inflammatory infiltration, were identified from the pathological report. We analyzed the severity and chronicity of TILs stratified by gender, age groups, biopsy indications, and concurrent glomerular diseases. We also examined the correlation between TIL and glomerulosclerosis.ResultsOf 56,880 patients with biopsy-proven glomerular disease, 79.5% had TILs. Renal interstitial inflammatory infiltration was the most common type of TIL (77.7%), followed by renal tubular atrophy (56.0%) and renal interstitial fibrosis (32.8%). Severe and chronic TILs were more common in adults than in children. The three glomerular diseases with the highest proportion of moderate-to-severe and chronic TIL were diabetic nephropathy, immunoglobulin A (IgA) nephropathy and focal segmental glomerulosclerosis. The severity of TILs was moderately correlated with glomerulosclerosis score (r=0.51). Moderate-to-severe and chronic TIL were more common in southern China. After adjusting for age, sex, hospital level, region, biopsy indication and type of concurrent glomerular diseases, adults with renal arteriole injury had a six-fold higher risk of moderate-to-severe TIL [odds ratio (OR), 7.12; 95% confidence interval (CI), 6.42 to 7.91] and a three-fold higher risk of chronic TIL (OR, 4.58; 95% CI, 4.37 to 4.79).ConclusionsTILs were common in patients with biopsy-proven glomerular disease. The type and severity of TILs varied with age, region and concurrent glomerular diseases. Renal arteriole injury and glomerulosclerosis was associated with a significantly increased risk of TIL.

Histone Deacetylase Inhibitor-loaded Calcium Alginate Microspheres for Acute Kidney Injury Treatment.

The protective effects of histone deacetylase (HDAC) inhibitors were highlighted in the treatment of kidney diseases, especially acute kidney injury (AKI). However, the currently available HDAC inhibitor cannot be delivered to the kidney properly because of its poor solubility in aqueous solutions. Therefore, calcium alginate (Ca-ALG) microspheres were proposed as microcarriers for the delivery of HDAC inhibitors in this study. First, Ca-ALG microspheres with high sphericity were obtained by a single-emulsion microfluidic strategy. Then, we selected suitable Ca-ALG microspheres for HDAC inhibitor loading by analyzing the swelling ratio and the release property using different parameters. Besides, thermal stimulation will change the drug release property of Ca-ALG microspheres in in vitro experiments. Furthermore, the HDAC inhibitor-loaded microspheres were delivered to the kidney by renal subcapsular injection for evaluating the treatment effects in mice with ischemia-reperfusion-induced AKI. The in vivo results showed that the HDAC inhibitor-loaded Ca-ALG microspheres could effectively reduce the renal regional inflammatory response and macrophage infiltration. Taken together, we proposed a promising therapy with an effective kidney-targeted drug delivery for the treatment of AKI.

Proliferation of HIV-infected renal epithelial cells following virus acquisition from infected macrophages.

HIV-1 can infect and persist in different organs and tissues, resulting in the generation of multiple viral compartments and reservoirs. Increasing evidence supports the kidney as such a reservoir. Previous work demonstrated that HIV-1 infected CD4 T-cells transfer virus to renal tubule epithelial (RTE) cells through cell-to-cell contact. In addition to CD4 T cells, macrophages represent the other major target of HIV-1. Renal macrophages induce and regulate inflammatory responses and are critical to homeostatic regulation of the kidney environment. Combined with their ability to harbour virus, macrophages may also play an important role in the spread of HIV-1 infection in the kidney. Multiparametric histochemistry analysis was performed on kidney biopsies from individuals with HIV-1 associated nephropathy (HIVAN). Primary monocyte-derived macrophages were infected with a GFP-expressing replication competent HIV-1. HIV-1 transfer from macrophages to RTE cells was carried out in a coculture system and evaluated by fluorescence-microscopy and flow-cytometry. Live imaging was performed to assess the fate of HIV-1 infected RTE cells over time. We show that macrophages are abundantly present in the renal inflammatory infiltrate of individuals with HIVAN. We observed contact-dependent HIV-1 transfer from infected macrophages to both primary and immortalized renal cells. Live imaging of HIV-1 infected RTE cells revealed four different fates: proliferation, hypertrophy, latency and cell death. Our study suggests that macrophages may play a role in the dissemination of HIV-1 in the kidney and that proliferation of infected renal cells may contribute to HIV-1 persistence in this compartment.

Melatonin pretreatment alleviates renal ischemia-reperfusion injury by promoting autophagic flux via TLR4/MyD88/MEK/ERK/mTORC1 signaling.

Autophagy is an important mechanism for cellular homeostasis and survival during pathologic stress conditions in the kidney, such as ischemia-reperfusion (IR) injury. In this study, renal IR was induced in female C57BL/6 mice after melatonin administration. Renal function, histological damage, inflammatory infiltration, cytokine production, oxidative stress, antioxidant capacity, autophagy changing, apoptosis levels, and autophagy-associated intracellular signaling pathway were assessed to evaluate the impact of antecedent melatonin treatment on IR-induced renal injury. The administration of melatonin resulted in significantly preserved renal function, and the protective effect was associated with ameliorated oxidative stress, limited pro-inflammatory cytokine production, and neutrophil and macrophage infiltration. Moreover, autophagic flux was increased after melatonin administration while the apoptosis levels were decreased in the melatonin-pretreated mice. Using TAK-242 and CRX-527, we confirmed that MyD88-dependent TLR4 and MEK/ERK/mTORC1 signaling participated in melatonin-induced autophagy in IR mice. Collectively, our results provide novel evidence that antecedent melatonin treatment provides protection for the kidney against IR injury by enhancing autophagy, as regulated by the TLR4/MyD88/MEK/ERK/mTORC1 signaling pathway. Therefore, melatonin preconditioning offers a potential therapeutic approach to prevent renal IR injury related to various clinical conditions.

P0984SGLT2 INHIBITION PREVENTS RENAL FIBROSIS IN CYCLOSPORINE NEPHROPATHY

Abstract Background and Aims Sodium glucose cotransporter 2 (SGLT2) inhibitors, a new class of antidiabetic drugs, showed nephroprotection in type 2 diabetic patients. The mechanisms underlying nephroprotection are not completely known and it is unclear whether the nephroprotective effects are present also in non-diabetic nephropathy. The aim of this study was to evaluate the effects of empagliflozin, a SGLT-2 inhibitor, in cyclosporine nephropathy in the absence of diabetes. Method Ten days before the beginning and then during the entire experimental periods, low-salt diet (Teklad 7034) was administered to Sprague Dawley rats. Cyclosporine-A (CsA, 15 mg/kg/day, intraperitoneal injection; n=6) and CsA plus empagliflozin (Empa, 10 mg/kg /day, per os; n=6) were administered for 4 weeks. Control group was treated with placebo (n=6). Blood pressure was measured by plethysmographic method at the beginning and at the end of the experimental period. At the end of the protocol, the kidneys were excised for histomorphometric analysis of renal fibrosis and for immunohistochemical evaluation of inflammatory infiltrates and tyrosine hydroxylase expression, used as marker of symphatetic nerve activity. Results The rats treated with CsA showed a significant increase (p &amp;lt;0.01) in blood pressure, which was slightly reduced by administration of empagliflozin. CsA administration caused an increase in glomerular and tubulo-interstitial fibrosis (p &amp;lt;0.05), renal inflammatory infiltrates (p &amp;lt;0.05) and tyrosine hydroxylase expression (p &amp;lt;0.01) as compared to the control rats. Treatment with empagliflozin reduced glomerular and tubulo-interstitial fibrosis (p &amp;lt;0.05), inflammatory cell infiltration (p &amp;lt;0.01) and tyrosine hydroxylase expression (p &amp;lt;0.01), as compared to CsA-treated rats. Conclusion Empagliflozin administration showed protective effects on cyclosporine nephropathy, decreasing renal fibrosis, macrophage infiltration and tyrosine hydroxylase expression. These data suggest that the nephroprotective role of empagliflozin could not be restricted only to diabetic nephropathy.

P0722VITAMIN D-DEFICIENT RATS RECOVER RENAL ALTERATIONS AFTER VITAMIN D REPLACEMENT FOLLOWING ISCHEMIA-REPERFUSION INJURY

Abstract Background and Aims Several studies have been linking vitamin D deficiency (VDD) as a non-traditional risk factor to acute kidney injury (AKI). It has been shown that VDD associated with AKI potentiates the injury and may accelerate the progression of kidney disease. Conversely, some lines of evidence have been describing that vitamin D sufficiency can be considered as a renal protective factor. Thus, we aimed to verify the effect of vitamin D replacement (R) on the renal disease progression post ischemia-reperfusion injury (IRI) in vitamin D deficient rats. Method We performed bilateral 45 min IRI on day 30 in all animal groups. Male Wistar rats were randomized into three groups: 1- IRI (fed a standard diet for 120 days); 2- VDD+IRI (fed a vitamin D-free diet for 120 days); and 3- VDD+IRI+R (fed a vitamin D-free diet for 30 days and just after IRI, on day 31, we reintroduced the standard diet for more 90 days). We evaluated inulin clearance (Cin); mean arterial pressure (MAP); renal blood flow (RBF); plasma levels of Vitamin D2+D3 [25(OH)D] and Parathormone (PTH) by ELISA. In addition, we performed histomorphometrical studies for CD3+ cells, fibronectin and vimentin as well as the fractional interstitial area (FIA). Moreover, we run qPCR studies and immunoblotted for vitamin D receptor (VDR) and Klotho. All the results are described in table 1. Results Vitamin D replacement restored the plasma levels of 25(OH)D and PTH in VDD+IRI+R group. Also, VDD+IRI+R presented an improvement of renal function and hemodynamic parameters. In addition, we observed more evident alterations in tubulointerstitial compartment, featuring interstitial expansion (renal fibrosis and inflammatory cell infiltrates) and fibronectin and vimentin expression in VDD+IRI rats. These alterations were recovered by vitamin D replacement. Concerning VDR and Klotho data, our results revealed a decreased expression of these targets in VDD+IRI group. On the other hand, vitamin D replacement retrieved the expression of VDR and Klotho. Conclusion Our study suggests that vitamin D replacement improved the recovery of renal function, hemodynamics, inflammatory and morphological alterations. In addition, vitamin D replacement was able to reestablish the expression of VDR and Klotho in IRI-AKI associated with vitamin D deficiency. Thus, it is recommendable that vitamin D levels should be observed as well as its reposition should be taken into account in renal patients.

Co‐Delivery of Anti‐Inflammatory and Proliferative Agents by Injectable Hydrogel to Promote Tissue Repair after Acute Kidney Injury

ObjectIschemia‐reperfusion (I/R)‐induced acute renal injury has become a serious issue in clinical. The increasingly mortality, induced by chronic fibrosis and further renal dysfunction which caused by poor recovery of acute injury, has made it urgent to address kidney tissue repair. By using a renal I/R injury model, we aimed to investigate the role of a hydrogel‐based dual‐drug delivery platform on promoting tissue repair.MethodsAn injectable, self‐assembling peptide/heparin (SAP/Hep) hydrogel was employed to co‐deliver TNF‐α neutralizing antibody (anti‐TNF‐α) and hepatocyte growth factor (HGF). We analyzed the microstructure and controlled release properties of KLD2R/Hep hydrogel in vivo. The effects of the drug‐loaded hydrogel (SAP‐drug) on renal injury were evaluated in a rodent renal I/R injury model.ResultsIn vitro, the SAP/Hep hydrogel exhibited a faster release of anti‐TNF‐α with a sustained release of HGF, and both drugs could maintain better bioactivities after release. In vivo, combined anti‐TNF‐α/HGF showed better renal protective potential than anti‐TNF‐α or HGF alone. Compared with free‐drug (anti‐TNF‐α/HGF in solution) or SAP alone , SAP‐drug treatment significantly reduced the levels of serum creatinine (Scr)/blood urea nitrogen (BUN), tubular apoptosis, renal inflammatory factors and macrophage infiltration. Moreover, the SAP‐drug group exhibited better efficacy on promoting tubular cell proliferation and dedifferentiation than SAP or free‐drug alone, and thus reduced chronic renal fibrosis in I/R injury mice.ConclusionsWe confirmed that SAP could sequentially deliver anti‐TNF‐α and HGF to achieve anti‐inflammatory and pro‐proliferative effects simultaneously with only once injection, and thus can be a promising delivery platform for tissue repair.Support or Funding InformationThis work was supported by grants from National Natural Science Foundation of China (81571808, 31871001, 31200754) and Sichuan Science and Technology Program (2019YJ0069)Design of dual‐drug delivery KLD2R/Hep hydrogel for enhancing renal repair. The formation of the cationic KLD2R/anionic heparin hybrid hydrogel via electrostatic interaction, and heparin further had high affinity to HGF. The encapsulated anti‐TNF‐α and HGF were released from hydrogel in a sequential manner, and thus synergistically promoted tissue repair.Figure 1

SAT-393 The profile of biopsy-proven renal tubulointerstitial lesions in patients with Glomerular disease

Renal tubules and interstitium are vulnerable to injury and play a central role in the progression of various CKDs. However, high quality study on the profiles of biopsy-proven tubulointerstitial lesions (TIL) is extremely limited. We conducted a retrospective renal biopsy series including 62,521native biopsies at 1211 hospitals across China from 2015 to 2017. The TILs, including the shedding of tube epithelial, renal tubular atrophy, renal interstitial fibrosis, edema and inflammatory infiltration, were identified from the pathological report. We analyzed the severity and chronicity of TILs stratified by gender, age groups, biopsy indications, and concurrent glomerular diseases. We also examined the effect of intrarenal arterial lesion on the risk of TIL. Of the 56,701 patients with exact pathological diagnosis, 79.95% of the patients had tubulointerstitial lesions, renal interstitial inflammatory infiltration was the most common type of TIL (77.70%), followed by renal tubular atrophy (56.36%) and renal interstitial fibrosis (27.84%). Severe and chronic TIL was more common in adults than in children. The prevalence of moderate to severe TIL was 74.14%, 26.00%, and 18.84% in patients with diabetic nephropathy (DN), IgA nephropathy (IgAN) and focal segmental glomerulosclerosis (FSGS), respectively. Similarly, patients with DN, IgAN, focal segmental glomerulosclerosis (FSGS) were more likely to have chronic TIL. Moderate to severe TIL was more common in the south of China. After adjusting for age, sex, hospital level, region, biopsy indication and type of concurrent glomerular diseases, patients with renal arteriole injury had a six-fold higher risk of TIL (OR 7.22, 95% CI, 6.48 to 8.05). In this large, multicenter renal biopsy series, the type and severity of TILs varied with age, region and concurrent glomerular diseases. Renal arteriole injury was associated with a significantly increased risk of TIL.

Renal Anti-Fibrotic Effect of Sodium Glucose Cotransporter 2 Inhibition in Angiotensin II-Dependent Hypertension

Background: Clinical trials have shown that empagliflozin (Empa), a sodium-glucose cotransporter 2 (SGLT2) inhibitor, promotes nephroprotective effects in diabetic patients. The mechanisms underlying nephroprotection are not completely known and it is not known whether the renal beneficial action is present even in non-diabetic kidney disease. The aim of this study was to evaluate the effect of Empa administration on the development of renal fibrosis in an experimental model of angiotensin II (Ang II)-dependent hypertension. Methods: Sprague Dawley rats (n = 31) were divided into 4 experimental groups. Ang II (200 ng/kg/min, osmotic minipumps, s.c., n = 9) or Ang II + Empa (10 mg/kg/day, per os, n = 10) were administered for 2 weeks. Control rats were treated with placebo (physiological saline, n = 6), and another group was treated with placebo plus Empa (n = 6) for the same period. Blood pressure (plethysmographic method) was measured at the beginning and at the end of the experimental protocol. After 2 weeks, the rats were euthanized and the kidneys were excised for histomorphometric evaluation of glomerular and tubulo-interstitial fibrosis and for the immunohistochemical evaluation of inflammatory infiltrates (monocytes/macrophages) and types I and IV collagen expression. Results: The administration of Ang II resulted in an increase in blood pressure (p < 0.01), glomerular (p < 0.05) and tubulo-interstitial (p < 0.01) fibrosis, renal inflammatory infiltrates (p < 0.01) and type I (p < 0.01) and type IV collagen expression (p < 0.05) compared to the control group. Treatment with Empa did not significantly modify the increase in blood pressure due to Ang II, but prevented the development of renal glomerular and tubulo-interstitial fibrosis, and the increase in inflammatory infiltrates and types I and IV collagen expression in Ang II-treated rats (p < 0.01). Conclusions: These data demonstrate that the treatment with Empa prevents the development of renal fibrosis in Ang II-dependent hypertension. In Ang II-dependent hypertension, the anti-fibrotic effect due to SGLT2 inhibition is caused by the reduction of inflammatory infiltrates and it is independent on the modulation of blood pressure increase.

Targeted VEGF (Vascular Endothelial Growth Factor) Therapy Induces Long-Term Renal Recovery in Chronic Kidney Disease via Macrophage Polarization.

Chronic kidney disease (CKD) universally associates with renal microvascular rarefaction and inflammation, but whether a link exists between these 2 processes is unclear. We designed a therapeutic construct of VEGF (vascular endothelial growth factor) fused to an ELP (elastin-like polypeptide) carrier and show that it improves renal function in experimental renovascular disease. We test the hypothesis that ELP-VEGF therapy will improve CKD, and that recovery will be driven by decreasing microvascular rarefaction partly via modulation of macrophage phenotype and inflammation. CKD was induced in 14 pigs, which were observed for 14 weeks. At 6 weeks, renal blood flow and filtration were quantified using multidetector computed tomography, and then pigs received single intrarenal ELP-VEGF or placebo (n=7 each). Renal function was quantified again 4 and 8 weeks later. Pigs were euthanized and renal microvascular density, angiogenic and inflammatory markers, fibrosis, macrophage infiltration, and phenotype were quantified. Loss of renal hemodynamics in CKD was progressively recovered by ELP-VEGF therapy, accompanied by improved renal microvascular density, fibrosis, and expression of inflammatory mediators. Although renal macrophage infiltration was similar in both CKD groups, ELP-VEGF therapy distinctly shifted their phenotype from proinflammatory M1 to VEGF-expressing M2. Our study unravels potential mechanisms and feasibility of a new strategy to offset progression of CKD using drug-delivery technologies. The results indicate that renal recovery after ELP-VEGF therapy was largely driven by modulation of renal macrophages toward VEGF-expressing M2 phenotype, restoring VEGF signaling and sustaining improvement of renal function and microvascular integrity in CKD.