Renal Interstitium Research Articles

Expression and immunofluorescent localization of the Edwardsiella tarda EvpC protein in infected Pelteobagrus fulvidraco

To determine the expression and localization of immunofluorescent secretory proteins encoded by Edwardsiella tarda (E. tarda) EvpC gene in tissues of experimentally infected yellow catfish Pelteobagrus fulvidraco (P. fulvidraco), an indirect immunofluorescent staining method was performed for detecting EvpC protein. A rabbit anti-EvpC polyclonal antibody, raised against a recombinant 6× His-EvpC fusion protein expressed in Escherichia coli, was prepared, purified, and used as the primary antibody. Sixty fish (50 ± 2.2 g) were intraperitoneally administered with E. tarda (infected group). Tissues were collected and prepared for indirect immunofluorescent staining at 4, 8, 12, 24, 48, 72, and 96 h post-inoculation. E. tarda EvpC encoding protein antigens could be detected in the liver, spleen, and kidney at 4 h after challenge, in the intestine at 12 h after challenge, in brain tissue 24 h after challenge, and in renal interstitium and liver tissue at 48 h. In addition, the intensity of positive staining of E. tarda EvpCp antigen in various tissues increased sharply from 8 to 96 h, peaking around 48–72 h, and subsequently decreasing steadily after 96 h, thus suggesting the expression levels of E. tarda EvpCp in systemic organs to be closely correlated with disease progression. The distribution pattern of E. tarda EvpCp was more prevalent in the spleen, liver, and kidney after infection, than in the intestine and gills. The present study may be useful not only for describing the characteristics of EvpCp expression and distribution in the chosen tissue of P. fulvidraco, but also for understanding pathogenesis by E. tarda and providing a basis for further functional analysis of the gene.

Enfermedades renales tubulointersticiales. Conceptos generales. Formas agudas

Tubulointerstitial kidney diseases (TKD) encompass a heterogeneous group of disorders characterizing by tubular damage and, pathologically, inflammatory signs in the renal interstitium. Clinical classification is arbitrary and distinguishes between acute (ATKD) and chronic (CTKD) patterns. Its etiology is diverse and, sometimes, the same cause can provoke ATKD and CTKD. Acute forms present acute renal failure, meanwhile chronic one causes progressive chronic kidney disease. Commonly, pathological findings are unspecific. Oedema and acute inflammatory infiltration (polynuclear neutrophils and eosinophils) characterize ATKD. Infiltration of lymphocytes and monocytes and interstitial fibrosis highlights in CTKD. If possible, the therapeutic goal is to reverse the acute inflammatory process. In CTKD treatment looks for to eliminate the cause and to avoid the use of nephrotoxic substances.

PTEN improve renal fibrosis in vitro and in vivo through inhibiting FAK/AKT signaling pathway.

Renal fibrosis, the ultimate common pathway of progressive nephropathy, is characterized by excess accumulation and deposition of extracellular matrix (ECM) within the renal interstitium and glomeruli, finally resulting in end-stage kidney failure. TGFβ1 is not only abnormally increased during fibrosis but also involved in ECM induction and accumulation. Based on the bioinformative analyses, phosphatase and tensin homolog deleted on chromosome ten (PTEN) and focal adhesion kinase (FAK) signaling pathway might be involved in TGFβ1 functions on renal fibrosis development. In the present study, fibrosis was induced in HK-2 cells using TGFβ1 and PTEN expression was significantly suppressed by 24 or 48 hours TGFβ1 treatment. PTEN overexpression in HK-2 cells improved TGFβ1-induced fibrosis within α-SMA and E-cadherin. According to the KEGG signaling pathway annotation analyses on microarray profiles (GSE23338 and GSE20247) and immunoblotting validation, FAK signaling might be involved in PTEN functions in TGFβ1-induced fibrosis. PTEN overexpression significantly inhibited TGFβ1- or unilateral ureteral obstruction (UUO)-induced FAK signaling pathway activation both in vitro and in vivo; more importantly, PTEN silence enhanced TGFβ1- or UUO-induced fibrosis, while FAK inhibitor PF567721 significantly reversed the effects of PTEN silence, indicating that PTEN exerted its effects on TGFβ1- and UUO-induced fibrotic development in vitro and in vivo via inhibiting FAK signaling pathway. In summary, these findings indicate that PTEN could improve cellular fibrotic changes and renal fibrosis via inhibiting FAK/AKT signaling pathway. Restoring PTEN expression to target FAK/AKT signaling pathway might be a potent strategy for renal fibrosis treatment.

The involvement of autotaxin in renal interstitial fibrosis through regulation of fibroblast functions and induction of vascular leakage

The accumulation of fibroblasts is a critical step in the development of fibrosis, and lysophosphatidic acid (LPA) promotes fibrosis by regulating multiple fibroblast functions. Autotaxin (ATX) is a key LPA-producing enzyme, and we hypothesized that ATX contributes to the development of renal interstitial fibrosis through LPA-mediated effects on fibroblast functions. In a mouse model of renal interstitial fibrosis induced by unilateral ureteral obstruction (UUO), the levels of renal ATX protein and activity increased with the progression of fibrosis in ligated kidneys, despite concurrent reductions in renal ATX mRNA. UUO enhanced vascular permeability in the renal interstitium, and ATX protein localized to areas of vascular leak, suggesting that vascular leak allowed ATX to enter the renal interstitium. In vitro studies showed that ATX induces the migration and proliferation of renal fibroblasts and enhances the vascular permeability of endothelial monolayers. Finally, pharmacological inhibition of ATX partially attenuated renal interstitial fibrosis. These results suggest that during the development of renal fibrosis, ATX accumulates in the renal interstitium and drives fibroblast accumulation and promotes renal interstitial vascular leak, thereby partially contributing to the pathogenesis of renal interstitial fibrosis. Taken together, ATX inhibition may have the potential to be a novel therapeutic strategy to combat renal interstitial fibrosis.

APOL1 renal risk variants promote cholesterol accumulation in tissues and cultured macrophages from APOL1 transgenic mice.

Apolipoprotein L1 (APOL1) genetic variants G1 and G2, compared to the common allele G0, are major risk factors for non-diabetic kidney disease in African descent populations. APOL1 is a minor protein component of HDL, as well as being expressed in podocytes and vascular cells. Reverse cholesterol transport involves the transport of cholesterol to HDL by cellular ATP-binding cassette; ABCA1 and ABCG1 with subsequent delivery from peripheral tissues to the liver. With impaired reverse cholesterol transport, lipid accumulation occurs and macrophages morphologically transform into foam cells, releasing inflammatory factors. We asked whether the APOL1 risk variants alter peripheral cholesterol metabolism and specifically affect macrophage cholesterol efflux. Tissues and bone marrow (BM)-derived monocytes were isolated from wild-type mice (WT) and from BAC/APOL1 transgenic (APOL1-G0, APOL1-G1, and APOL1-G2) mice, which carry a bacterial artificial chromosome that contains the human APOL1 genomic region. Monocytes were differentiated into macrophages using M-CSF, and then polarized into M1 and M2 macrophages. Cholesterol content, cholesterol efflux, and ABCA1 and ABCG1 mRNA expression were measured. Kidney, spleen, and bone marrow-derived macrophages from APOL1-G1 and -G2 mice showed increased cholesterol accumulation and decreased ABCA1 and ABCG1 mRNA levels. BM-derived macrophages from APOL1-G1 and -G2 mice showed significantly reduced cholesterol efflux compared to WT or APOL1-G0 macrophages. Taken together, the evidence suggests that APOL1-G1 and -G2 risk variants impaired reverse cholesterol transport through decreased expression of cholesterol efflux transporters suggesting a possible mechanism to promote macrophage foam cell formation, driving inflammation in the glomerulus and renal interstitium.

Fibroblast-specific plasminogen activator inhibitor-1 depletion ameliorates renal interstitial fibrosis after unilateral ureteral obstruction.

Plasminogen activator inhibitor-1 (PAI-1) expression increases extracellular matrix deposition and contributes to interstitial fibrosis in the kidney after injury. While PAI-1 is ubiquitously expressed in the kidney, we hypothesized that interstitial fibrosis is strongly dependent on fibroblast-specific PAI-1 (fbPAI-1). Tenascin C Cre (TNC Cre) and fbPAI-1 knockdown (KD) mice with green fluorescent protein (GFP) expressed within the TNC construct underwent unilateral ureteral obstruction and were sacrificed 10 days later. GFP+ cells in fbPAI-1 KD mice showed significantly reduced PAI-1 expression. Interstitial fibrosis, measured by Sirius red staining and collagen I western blot, was significantly decreased in fbPAI-1 KD compared with TNC Cre mice. There was no significant difference in transforming growth factor β (TGF-β) expression or its activation between the two groups. However, GFP+ cells from fbPAI-1 KD mice had lower TGF β and connective tissue growth factor (CTGF) expression. The number of fibroblasts was decreased in fbPAI-1 KD compared with TNC Cre mice, correlating with decreased alpha smooth muscle actin (α-SMA) expression and less fibroblast cell proliferation. TNC Cre mice had decreased E-cadherin, a marker of differentiated tubular epithelium, in contrast to preserved expression in fbPAI-1 KD. F4/80-expressing cells, mostly CD11c+/F4/80+ cells, were increased while M1 macrophage markers were decreased in fbPAI-1 KD compared with TNC Cre mice. These findings indicate that fbPAI-1 depletion ameliorates interstitial fibrosis by decreasing fibroblast proliferation in the renal interstitium, with resulting decreased collagen I. This is linked to decreased M1 macrophages and preserved tubular epithelium.

Modulatory effects of fibronectin on calcium oxalate crystallization, growth, aggregation, adhesion on renal tubular cells, and invasion through extracellular matrix

Fibronectin, an extracellular matrix (ECM) protein, has been thought to be involved in pathogenic mechanisms of kidney stone disease, especially calcium oxalate (CaOx) type. Nevertheless, its precise roles in modulation of CaOx crystal remained unclear. We thus performed a systematic evaluation of effects of fibronectin on CaOx monohydrate (COM) crystal (the major causative chemical crystal in kidney stone formation) in various stages of kidney stone pathogenesis, including crystallization, crystal growth, aggregation, adhesion onto renal tubular cells, and invasion through ECM in renal interstitium. The data showed that fibronectin significantly decreased crystallization, growth and adhesive capability of COM crystals in a dose‐dependent manner. In contrast, COM crystal aggregation and invasion through ECM migration chamber were significantly enhanced by fibronectin in a dose‐dependent fashion. Sequence analysis revealed two and six calcium‐binding and oxalate‐binding domains, respectively. Saturation of fibronectin with oxalate prior to crystal modulation assays could abolish all the effects of fibronectin, whereas saturation with calcium had minimal interference. These data strongly indicate the dual functions of fibronectin, which serves as an inhibitor for COM crystallization, crystal growth and adhesion onto renal tubular cells, but on the other hand, acts as a promoter for COM crystal aggregation and invasion through ECM. Moreover, its crystal modulatory activities are mainly due to oxalate‐binding capability.Support or Funding InformationThis study was supported by Mahidol University research grant and the Thailand Research Fund (IRN60W0004 and IRG5980006).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Adefovir accumulation and nephrotoxicity in renal interstitium: Role of organic anion transporters of kidney

Common characteristics of drug induced nephrotoxicity are renal tubular and interstitial injury. Many studies have only focused on renal tubular injury. However, less is known about the effects of drugs in the renal interstitium on the nephrotoxicity. The aim of this study was to investigate the pharmacokinetics of adefovir (ADV) and the nephrotoxicity in the renal interstitium. Rats were treated with ADV alone or in combination with probenecid for 1, 7, 14, or 28 days. The renal interstitial fluid was collected by renal microdialysis. The concentration of ADV was determined by HPLC-MS/MS. Nephrotoxicity was evaluated by biochemical parameters or histological analysis. The results showed that organic anion transporters (OATs) inhibitor probenecid significantly increased the area under concentration-time curves (AUC) and peak concentration (Cmax) of ADV in the renal interstitium, while the clearance (CL) in the renal interstitium was decreased in the ADV plus probenecid group compared to the ADV groups. After long-term treatment, interstitial fibrosis was present in the ADV plus probenecid group, whereas no trace of that could be detected in the ADV groups. Furthermore, a decrease was observed in the expression of OATs/Oats, which was dependent upon the concentrations and time of ADV treatment. In conclusion, it is possible that ADV could be accumulated in the interstitium when Oats were inhibited, which could cause renal interstitial fibrosis. Simply reducing cell uptake in long-term treatment might not be an effective method to protect against chronic nephrotoxicity.

Structural Changes Induced by Potassium Dichromate in Renal Cortex of Adult Male Albino Rats and the Possible Protective Role of Selenium

AbstractBackground: Potassium Dichromate (Cr) is a common environmental agent that has been considered potentially carcinogenic, making it an issue of major environmental and search concern.Aim of Study: The aim of this study was to throw light on the possible structural changes in the renal cortex of adult male albino rats after administration of Cr and to elucidate the possibility of protection by selenium (Se) against the deleterious effects of Cr.Material and Methods: Thirty six adult male albino rats weighing 200-230g were divided equally into four groups as follow: Group I received balanced diet and tap water only; Group II received Se orally 0.25mg/kg b.w/daily; Group III rats were received Cr orally 67mg/kg/daily and Group IV rats were received oral Cr and Se at same previous doses. Animals were sacrificed on day 28.Results: The Cr-treated group showed damage of the renal cortex including capsule and renal interstitium. Masson's trichrome staining showed abundant collagen fibers. The expression of caspase-3 in the cytoplasm showed strong positive immune reaction in group III. Biochemical results showed Cr induced significant increase in blood urea nitrogen (BUN), serum creatinine (SC) and uric acid (UA). While morphometric measurement indicated that Cr induced a sta-tistical significant decrease in rat body weight and diameter of glomeruli when compared with other groups. The Cr-treated group also showed a statistical significant increase in kidney weight, surface area of Bowman space, diameter of the con-voluted tubules, area percentage of collagen fibers and ex-pression of caspase-3. Meanwhile, administration of Se showed some protection against the toxic effect of Cr on renal cortex.Conclusion: It is suggested that administration Se mod-erately improves the deleterious histological and biochemical changes induced by Cr on renal cortex.

N-Acetylcysteine Mitigates Histopathological and Ultrastructural Alterations Induced by Chronic Usage of Acetaminophen in the Rat Renal Cortex

Background: Acetaminophen or N-acetyl-p-aminophenol (APAP) is a potent analgesic and antipyretic drug that is widely used. N-Acetylcysteine (NAC) is a synthetic antioxidant, acting as a free radical scavenger.Aim of the work: This work was designed to demonstrate the effect of NAC on the histological changes induced by chronic usage of acetaminophen in the renal cortex.Materials and methods: Forty male albino rats distributed into four groups were utilized. Control group: given distilled water. NAC-group: given NAC (150 mg/kg/day), orally, once daily for 4 weeks. Aceta-group: given Acetaminophen (APAP 300 mg/kg/day) orally, once daily for 4 weeks. Aceta+NAC group: given Acetaminophen (APAP 300 mg/kg/day) orally then NAC (150 mg/kg) orally 2 hours after oral APAP, once daily for 4 weeks. Small specimens of the renal cortex were obtained and processed for light and electron microscopic studies. Blood urea nitrogen (BUN) and serum creatinine levels were assessed. Morphometric and statistical studies were also performed.Results: Chronic usage of acetaminophen resulted in distortion and shrinkage of some glomeruli and partial destruction of Bowman's capsule. Proximal convoluted tubules showed dilatations, degenerative changes and loss of the apical brush border. Mononuclear cellular infiltration and deposition of collagenous fibers in the renal interstitium were also demonstrated. By electron microscope, glomerular basement membrane showed irregular thickening and loss of its trilaminar appearance. Cells lining the proximal convoluted tubules showed dense irregular nucleus, cytolplasmic vacuoles and loss of apical microvilli. These changes were accompanied by significant elevations in the serum levels of BUN and creatinine. NAC administration provoked partial improvement in the renal histology and a significant decline in serum levels of BUN and creatinine.Conclusion: N-acetylcysteine could attenuate histological and biochemical changes induced by chronic usage of acetaminophen in the rat renal cortex.

Neuroendocrine Tumor Therapy with Lutetium-177: A Literature Review.

The worldwide incidence of neuroendocrine tumors (NETs) has been increasing. They are a very diverse group of tumors which are commonly found in the gastrointestinal and bronchopulmonary tracts. These tumors usually express somatostatin receptors. Therefore, somatostatin analogs are used for symptom relief as well as treatment. Of the many therapeutic options available, peptide receptor radionuclide therapy (PRRT) has been shown to be very promising. In January 2018, the Food Drug and Authority (FDA) approved 177Lu-Dotatate for use in gastroenteropancreatic neuroendocrine tumors (GEP-NETs). Lutetium is a lower energy beta-emitting radionuclide. The therapeutic use of lutetium-177 (177Lu) has shown better results in advanced gastroenteropancreatic and bronchial neuroendocrine tumors when compared with other therapies available. Adverse effects associated with this therapy include myelotoxicity and nephrotoxicity as the radiopeptides are reabsorbed and accumulate in the renal interstitium. Everolimus is a good and safe option in patients pretreated with 177Lu-Dotatate. Lutetium, in combination with somatostatin analogs, has proven efficacy to treat gastroenteropancreatic neuroendocrine tumors in candidates with somatostatin receptor-positive advanced tumors and normal renal function. This therapy has great potential as it decreases tumor size, improves symptoms, and improves quality of life.

Modulatory effects of fibronectin on calcium oxalate crystallization, growth, aggregation, adhesion on renal tubular cells, and invasion through extracellular matrix.

Fibronectin, an extracellular matrix (ECM) protein, has been thought to be involved in pathogenic mechanisms of kidney stone disease, especially calcium oxalate (CaOx) type. Nevertheless, its precise roles in modulation of CaOx crystal remained unclear. We thus performed a systematic evaluation of effects of fibronectin on CaOx monohydrate (COM) crystal (the major causative chemical crystal in kidney stone formation) in various stages of kidney stone pathogenesis, including crystallization, crystal growth, aggregation, adhesion onto renal tubular cells, and invasion through ECM in renal interstitium. The data showed that fibronectin significantly decreased crystallization, growth and adhesive capability of COM crystals in a dose-dependent manner. In contrast, COM crystal aggregation and invasion through ECM migration chamber were significantly enhanced by fibronectin in a dose-dependent fashion. Sequence analysis revealed three calcium-binding and six oxalate-binding domains in fibronectin. Immunofluorescence study confirmed binding of fibronectin to COM crystals. Additionally, calcium- and oxalate-affinity assays confirmed depletion of both calcium and oxalate ions after incubation with fibronectin. Moreover, calcium-saturated and oxalate-saturated forms of fibronectin markedly reduced the modulatory activities of fibronectin on COM crystallization, crystal growth, aggregation, and adhesion onto the cells. These data strongly indicate the dual functions of fibronectin, which serves as an inhibitor for COM crystallization, crystal growth and adhesion onto renal tubular cells, but on the other hand, acts as a promoter for COM crystal aggregation and invasion through ECM. Finally, its COM crystal modulatory activities are most likely mediated through binding with calcium and oxalate ions on the crystals and in their environment.

Involvement of the NADPH oxidase 2 pathway in renal oxidative stress in Aqp11-/- mice

Aquaporin-11 (AQP11) is an intracellular AQP. Several studies with Aqp11-/- mice have shown that AQP11 has a role in normal development of the kidney after birth. Our previous studies have suggested that alteration of oxygen homeostasis may be involved in the kidney injury caused by AQP11 deficiency, although the underlying mechanism is largely unknown. To clarify this issue, we examined genes that are related to oxygen homeostasis in Aqp11-/- mice. Among 62 genes that are involved in oxygen homeostasis, 35 were upregulated by more than 2-fold in Aqp11-/- mice in comparison with wild-type mice. Pathway analysis using these genes extracted the pathway responsible for production of reactive oxygen species in macrophages. As expression of the genes involved in the NADPH oxidase 2 (NOX2) complex was dramatically increased by more than 14-fold, we further analyzed NOX2 at the protein level. Immunoblotting analysis demonstrated a dramatic increase of NOX2 protein in the kidney of Aqp11-/- mice, and immunohistochemistry showed that NOX2 protein and a marker protein for macrophages were increased in the renal interstitium. These results indicate that NOX2-induced oxidative stress accompanied by macrophage infiltration plays an important role in alteration of oxygen homeostasis in Aqp11-/- mice.

DMDD, Isolated from the Root of <i> Averrhoa carambola</i> L., Protects Against Diabetic Kidney Disease by Inhibiting TLR4/TGFβ Signaling Pathway

Objective: Diabetic kidney disease (DKD) is the leading cause of death and disability of diabetes mellitus. However, there is still a lack of specific drugs for the treatment of DKD. Objective: The chief aim of this research is to investigate the role and mechanism of DMDD for DKD. Methods: Wild type and TLR4 knockout mice were induced to diabetes. After 4-week treatment with DMDD, blood sugar, renal function, blood lipid and pathological changes were assessed. Real-time PCR, western blotting, and immunohistochemistry were employed to detect the expressions of TLR4, TGFβ1 and Smad2/3 in the renal tissue. Results: DMDD improved the serum lipid and decreased fasting blood glucose levels in diabetic mice. CysC and urinary albumin levels increased markedly in the diabetic group, and they were obviously decreased after 4 weeks of DMDD treatment. Compared with the WT diabetic mice, the urinary albumin and CysC in the TLR4-/- mice were expressed at lower levels. HE and Masson's staining revealed that DMDD clearly ameliorated pathological changes and renal fibrosis. When TLR4 gene was knock out, the pathological was improved. Mechanistically, TLR4, TGF-β1 and Smad2/3 were obvious up-regulation in the renal tissues of diabetic mice. The expressions of these proteins were significantly down-regulated after DMDD treatment (p< 0.05). In the TLR4-/- mice, mRNA and protein levels of TGF-β1 and Smad2/3 were obviously lower than those in the WT mice. In addition, IHC revealed that a strong in situ expressions of TLR4, TGF-β1 and Smad2/3 were seen in the kidney tissues of diabetic mice, which were distinctly weakened in the DMDD-treated mice. In the TLR4-/- mice, however, expressions of TGF-β1 and Smad2/3 were not remarkable increase in the diabetic mice compared with normal mice. Conclusions: These results strongly indicate that TLR4 is essential for DMDD protection against renal dysfunction in diabetic mice. Its hypoglycemic and anti-fibrosis effects were likely mediated by inhibiting of the TLR4/TGFβ signaling pathway, and the downregulation of Smads in the renal interstitium. Funding Statement: This research was supported by the National Natural Science Foundation of China (No. 81760665, 81460205), and Guangxi Natural Science Foundation under Grant No. 2017GXNSFBA198177. In addition, this project was financially supported by the project of improving the basic ability of young teachers in colleges and universities in Guangxi (KY2016YB098). Declaration of Interests: The authors declare no conflicts of interest in this work. Ethics Approval Statement: All experimental procedures were approved by the Institutional Animal Care and Use Committee of Guangxi Medical University.

Nonselective Cyclooxygenase Inhibition Retards Cyst Progression in a Murine Model of Autosomal Dominant Polycystic Kidney Disease.

Aim: Autosomal dominant polycystic kidney disease is one of the most common genetic renal diseases. Cyclooxygenase plays an important role in epithelial cell proliferation and may contribute to the mechanisms underlying cyst formation. The aim of the present study was to evaluate the role of cyclooxygenase inhibition in the cyst progression in polycystic kidney disease.Method: Pkd2WS25/- mice, a murine model which harbors a compound cis-heterozygous mutation of the Pkd2 gene were used. Cyclooxygenase expression was assessed in both human and murine kidney specimens. Pkd2WS25/- mice were treated with Sulindac (a nonselective cyclooxygenase inhibitor) or vehicle for 8 months starting at three weeks age, and then renal cyst burden was assessed by kidney weight and volume.Results: Cyclooxygenase-2 expression was up-regulated compared to control kidneys as shown by RNase protection in human polycystic kidneys and immunoblot in mouse Pkd2WS25/- kidneys. Cyclooxygenase-2 expression was up-regulated in the renal interstitium as well as focal areas of the cystic epithelium (p<0.05). Basal Cyclooxygenase-1 levels were unchanged in both immunohistochemistry and real-time PCR. Administration of Sulindac to Pkd2WS25/- mice and to control mice for 8 months resulted in reduced kidney weights and volume in cystic mice. Renal function and electrolytes were not significantly different between groups.Conclusion: Thus treatment of a murine model of polycystic kidney disease with Sulindac results in decreased kidney cyst burden. These findings provide additional implications for the use of Cyclooxygenase inhibition as treatment to slow the progression of cyst burden in patients with polycystic kidney disease.

Morphology and Evaluation of Renal Fibrosis.

With continuing damage, both the indigenous cells of the cortex and medulla, and inflammatory cells are involved in the formation and development of renal fibrosis. Furthermore, interactions among the glomerular, tubular, and interstitial cells contribute to the process by excessive synthesis and decreased degradation of extracellular matrix. The morphology of kidney is different from pathological stages of diseases and changes with various causes. At the end stage of the disease, the kidneys are symmetrically contracted with diffuse granules. Most glomeruli show diffuse fibrosis and hyaline degeneration, and intervening tubules become atrophied. Renal interstitium shows obvious hyperplasia of fibrous tissues with marked infiltration of lymphocytes, mononuclear cells, and plasma cells. The renal arterioles are wall thickening frequently because of hyaline degeneration. Morphologic analysis based on Masson staining of the kidney tissues has been regarded as the golden standard to evaluate the visual fibrosis. However, the present studies have found that the evaluation system has poor repeatability. Several computer-aided image analysis techniques have been used to assess interstitial fibrosis. It is possible that the evaluation of renal fibrosis is carried out by the artificial intelligence renal biopsy pathological diagnosis system in the near future.

Functional roles of Grainyhead-like transcription factors in renal development and disease.

Proper renal function relies on the tightly regulated development of nephrons and collecting ducts. This process, known as tubulogenesis, involves dynamic cellular and molecular changes that instruct cells to form highly organized tubes of epithelial cells which compartmentalize the renal interstitium and tubular lumen via assembly of a selective barrier. The integrity and diversity of the various renal epithelia is achieved via formation of intercellular protein complexes along the apical-basal axis of the epithelial cells. In recent years, the evolutionarily conserved family of Grainyhead-like (GRHL) transcription factors which encompasses three mammalian family members (Grainyhead-like 1, 2, 3) has emerged as a group of critical regulators for organ development, epithelial differentiation, and barrier formation. Evidence from transgenic animal models supports the presence of Grainyhead-like-dependent transcriptional mechanisms that promote formation and maintenance of epithelial barriers in the kidney. In this review, we highlight different Grhl-dependent mechanisms that modulate epithelial differentiation in the kidney. Additionally, we discuss how disruptions in these mechanisms result in impaired renal function later in life.

Nudel involvement in the high-glucose-induced epithelial-mesenchymal transition of tubular epithelial cells.

Nudel is a newly discovered factor related to cell migration. The tubular epithelial-mesenchymal transition (EMT) includes four steps: the loss of the adhesive properties of epithelial cells, the acquisition of a mesenchymal cell phenotype, the destruction of the tubular basal membrane, and the migration into the renal interstitium. The purpose of this study was to investigate the role of Nudel in the high-glucose-induced EMT of tubular epithelial cells. Human renal proximal tubular epithelial cells (HKCs) were treated with Nudel shRNA to clarify the role and mechanism of Nudel in tubular EMT induced by high glucose. We found that Nudel was expressed at a high level in high-glucose-stimulated HKCs, and the expression of Nudel was associated with the activation of signal transducer and activator of transcription 3. After transfection with Nudel shRNA, we detected the expression levels of E-cadherin, α-smooth muscle actin (α-SMA), and the Wiskott-Aldrich syndrome family of proteins (including WASP, N-WASP, WAVE1, WAVE2, and WAVE3) via assay. Cell migration was analyzed by the scratching method. The results showed that high glucose downregulated E-cadherin expression, upregulated α-SMA expression, and promoted the migration of HKCs. The expression levels of N-WASP, WAVE1, and WAVE2 were also elevated in HKCs treated with high glucose. All changes induced by high glucose were ameliorated by Nudel depletion. We conclude that Nudel participates in the transition and the migration of tubular epithelial cells via the regulation of WASP family proteins.

Exploration about the protection mechanism of 5-hydroxy-1-methylhydantoin on paraquat poisoning model

To investigate the effects of 5-hydroxy-1-methylhydantoin (HMH) on kidney injury induced by paraquat (PQ). Fifteen SPF healthy Kunming mice were randomly divided into normal saline (NS) control group, PQ poisoning model group and HMH intervention group, with 5 mice in each group. PQ poisoning model was challenged by one-time gavage of 30 mg/kg PQ solution. The NS group received the same amount of NS by gavage. The HMH group was given 100 mg/kg of HMH immediately after the model was made and continued to be gavaged. Mice in each group were sacrificed 1 day after HMH gavage and heart blood and renal tissue were harvested for examination. The morphological changes of renal tissue were observed under light microscope by hematoxylin-eosin (HE) staining. The content of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in renal tissue were detected according to the instructions of the kit. The expression of heme oxygenase-1 (HO-1) and interleukin-1β (IL-1β) in renal tissues were detected by Western Blot. The serum metabolites were detected by gas chromatography time-of-flight mass spectrometry (GC-TOF-MS), the overall distribution of each sample was observed by principal component analysis (PCA), the accuracy of the model was evaluated by multidimensional analysis orthogonal partial least squares-discriminant analysis (OPLS-DA), and the difference metabolites were screened by variable importance in the projection (VIP) value > 1. Light microscopic observation showed that: glomerular structure in NS group was clear, there was no hyperemia and inflammatory cell infiltration in renal interstitium and blood vessels. In PQ group, some glomeruli atrophy and necrosis, capillary congestion in glomeruli, infiltration of inflammatory cells around glomeruli, swelling of renal tubular epithelial cells, slight stenosis of lumen, and occasional necrosis and exfoliation of epithelial cells occurred. The degree of kidney injury in HMH group was significantly less than that in PQ group. Compared with the NS group, the content of MDA in the PQ group was significantly increased (nmol/g: 6.70±0.84 vs. 2.70±0.43, P < 0.01) and the activity of SOD was significantly decreased (kU/L: 33.30±4.66 vs. 50.20±3.23, P < 0.05), the protein expression of HO-1 and IL-1β were significantly increased (HO-1/β-actin: 1.11±0.12 vs. 0.61±0.13, IL-1β/β-actin: 0.93±0.13 vs. 0.32±0.06, both P < 0.05). Compared with the PQ group, the content of MDA in the HMH group was significantly decreased (nmol/g: 5.10±0.93 vs. 6.70±0.84, P < 0.05) and the activity of SOD was significantly increased (kU/L: 61.00±9.02 vs. 33.30±4.66, P < 0.05), the protein expression of HO-1 was significantly decreased (HO-1/β-actin: 0.77±0.07 vs. 1.11±0.12, P < 0.05), however, there was no significant difference in the protein expression of IL-1β (IL-1β/β-actin: 0.87±0.13 vs. 0.93±0.13, P > 0.05). Metabolite detection results showed that: compared with NS group, the levels of creatinine, glycine, succinic acid, fumaric acid and citric acid were significantly increased in the PQ group (VIP value was 1.50, 1.58, 1.64, 1.74 and 1.95 respectively, all P < 0.05), while the levels of palmitic acid,α-tocopherol and 6-phosphogluconic acid were significantly decreased (VIP value was 1.10, 1.55 and 1.56 respectively, all P < 0.05). Compared with the PQ group, the levels of creatinine and citric acid were significantly decreased in the HMH group (VIP value was 1.50 and 1.86, both P < 0.05), while trans-4-hydroxy-proline, D-glyceric acid, 2, 6-fructose phosphate, 6-phosphate gluconic acid and aminomalonic acid were significantly increased (VIP value was 1.36, 1.55, 1.63, 1.68 and 1.76 respectively, all P < 0.05). HMH protects kidney injury caused by PQ poisoning by correcting tricarboxylic acids cycle disturbance, lipid peroxidation and energy metabolism disturbance, and its mechanism is related to the regulation of HO-1 protein expression through Nrf2 pathway.

Tenascin-C protects against acute kidney injury by recruiting Wnt ligands

The development of acute kidney injury (AKI) is a complex process involving tubular, inflammatory, and vascular components, but less is known about the role of the interstitial microenvironment. We have previously shown that the extracellular matrix glycoprotein tenascin-C (TNC) is induced in fibrotic kidneys. In mouse models of AKI induced by ischemia-reperfusion injury (IRI) or cisplatin, TNC was induced de novo in the injured sites and localized to the renal interstitium. The circulating level of TNC protein was also elevated in AKI patients after cardiac surgery. Knockdown of TNC by shRNA invivo aggravated AKI after ischemic or toxic injury. This effect was associated with reduced renal β-catenin expression, suggesting an impact on Wnt signaling. Invitro, TNC protected tubular epithelial cells against apoptosis and augmented Wnt1-mediated β-catenin activation. Co-immunoprecipitation revealed that TNC physically interacts with Wnt ligands. Furthermore, a TNC-enriched kidney tissue scaffold prepared from IRI mice was able to recruit and concentrate Wnt ligands from the surrounding milieu exvivo. The ability to recruit Wnt ligands in this exvivo model diminished after TNC depletion. These studies indicate that TNC is specifically induced at sites of injury and recruits Wnt ligands, thereby creating a favorable microenvironment for tubular repair and regeneration after AKI.