Metformin modulates autophagic pathway in renal fibrosis induced by carbon tetrachloride in adult male albino rats.

BACKGROUNDS: Chronic kidney disease (CKD) is a global public health problem. All progressive chronic kidney disease (CKD) is characterized by tubulointerstitial fibrosis. Exposure to high concentrations of carbon tetrachloride (including vapor) can destroy the kidneys. Autophagy played an important role in maintaining the homeostasis of organs. Impaired autophagy was frequently associated with renal damage and fibrosis. Recent data suggests that metformin protects against a variety of kidney disorders. AIM: To investigate the protective role of metformin on carbon tetrachloride induced renal damage via autophagy pathway. MATERIALS AND METHODS: Forty adult male albino rats were divided into four equal groups (10 rats, each); Group 1: control group. Group 2: olive oil group received olive oil 1.5 mg/kg twice weekly S.C for 12 weeks. Group 3: The ccl4 group, the rats were received ccl4 1.5 mg/kg twice weekly S.C for 12 weeks. Group 4: CCL4 and Metformin group received concomitant treatment of CCL4, 1.5 mg/kg twice weekly S.C and 100 mg/kg/day Metformin orally for 12 weeks. After sacrifice, kidneys were taken from all animal groups and processed for light and electron microscopy, immunological studies and biochemical tests. Statistical analysis was done. RESULTS: Administration of ccl4 resulted in histopathological changes in the kidney tissue in the form of areas of tissue destruction, inflammatory cell infiltration, congestion and fibrosis. Ultrastructurally, irregular thickening of GBM was observed. Improvement was noticed with concomitant treatment of ccl4 with metformin. CONCLUSION: Metformin administration can modulate histological and biochemical effects in the renal tissue induced by of ccl4.

Ultrastructural characterization of maturing iPSC-derived nephron structures upon transplantation.

Pluripotent stem cell-derived kidney organoids hold great promise as a potential auxiliary transplant tissue for individuals with end-stage renal disease and as a platform for studying kidney diseases and drug discovery. To establish accurate models, it is crucial to thoroughly characterize the morphological features and maturation stages of the cellular components within these organoids. Nephrons, the functional units of the kidney, possess distinct morphological structures that directly correlate with their specific functions. High spatial resolution imaging emerges as a powerful technique for capturing ultrastructural details that may go unnoticed with other methods such as immunofluorescent imaging and scRNA sequencing. In our study, we have applied software capable of seamlessly stitching virtual slides generated from electron microscopy, resulting in high-definition overviews of tissue slides. With this technology, we can comprehensively characterize the development and maturation of kidney organoids when transplanted under the renal capsule of mice. These organoids exhibit advanced ultrastructural developments upon transplantation, including the formation of the filtration barrier in the renal corpuscle, the presence of microvilli in the proximal tubule, and various types of cell sub-segmentation in the connecting tubule similarly to those seen in the adult kidney. Such ultrastructural characterization provides invaluable insights into the structural development and functional morphology of nephron segments within kidney organoids and how to advance them by interventions such as a transplantation. Research Highlights High-resolution imaging is crucial to determine morphological maturation of hiPSC-derived kidney organoids. Upon transplantation, refined ultrastructural development of nephron segments was observed, such as the development of the glomerular filtration barrier.

Acacia senegal inhibits diabetic nephropathy ultrastructure alteration and renal biomarkers changes in rats / Acacia senegal inhibe la alteración de la ultraestructura de la nefropatía diabética y los cambios en los biomarcadores renales en rata

SUMMARY: Diabetic nephropathy (DN) is the most common complication of diabetes. Several studies have been done in a trial to protect against this problem at the ultrastructure level. This study investigates the protective effect of oral administration of Acacia senegal (AS) against the development of DN. Sixty male albino rats were randomly divided into six groups: control, Acacia senegal control, Diabetic untreated, diabetic insulin-treated, Diabetic AS treated, and Diabetic insulin and AS combined treated groups. Plasma glucose, HbA1c, serum Albumin, creatinine, urine creatinine was measured using specific kits. Determinations of creatinine clearance and blood pressure were done. The renal tissues of both kidneys were prepared to investigate under both light (LM) and electron microscope (EM). Ultrastructure examination of renal rats tissue of diabetic untreated rats showed the destruction of the glomerular basement membrane and endothelial cells together with hemorrhage in glomerular capsules (Bowman's capsules). On the other side, both LM and EM revealed improving the endothelial cells and the other glomerular capsules structures, especially with the combined treated group, which confirmed the improvement of the biochemical investigation in the study. In conclusion, from the present study, using the oral AS together with SC insulin could be protected against the development of DN.

RESUMEN: La nefropatía diabética (ND) es la complicación más común de la diabetes. Se han realizado varios estudios de ensayo para abordar esta dificultad a nivel de ultraestructura. Este estudio investiga el efecto protector de la administración oral de Acacia senegal (AS) contra el desarrollo de la ND. Se dividieron sesenta ratas albinas machos aleatoriamente en seis grupos: control, control de Acacia senegal, diabéticos no tratados, diabéticos tratados con insulina, diabéticos tratados con AS y grupos tratados con compuesto de insulina diabética + AS. Se midieron utilizando kits específicos, glucosa plasmática, HbA1c, albúmina sérica, creatinina en sangre y en orina. Se registraron la creatinina y la presión arterial. Los tejidos renales de ambos riñones se prepararon para investigar tanto con microscopio óptico (MO) como electrónico (ME). El examen de la ultraestructura del tejido renal de ratas diabéticas no tratadas mostró la destrucción de la membrana basal glomerular y las células endoteliales junto con hemorragia en las cápsulas glomerulares (cápsulas de Bowman). Por otro lado, tanto MO como ME revelaron una mejora de las células endoteliales y las estructuras capsulares glomerulares, en el grupo tratado con el compuesto, lo que confirmó la mejora de la investigación bioquímica. En conclusión, el uso de AS oral en combinación con insulina podría proteger contra el desarrollo de ND.

An update on the evolutionary origin of aglomerular kidney with structural and ultrastructural descriptions of the kidney in three fish species.

The kidney of fish contains numerous nephrons, each of which is divided into the renal corpuscle and renal tubules. This glomerular structure is the filtration unit of the nephron and is important for the kidney function, but it has been reported that the renal corpuscle was lost in at least four independent linages of fish (i.e., aglomerular kidney). In this study, the authors newly described renal structures for three species by histological and ultrastructural observations: two aglomerular kidneys from a seahorse Hippocampus barbouri and a toadfish Allenbatrachus grunniens and a glomerular kidney from a snake eel Pisodonophis boro. The renal development of H. barbouri was also described during 1-35 days after birth. In all species tested, the anterior kidney was comprised of haematopoietic tissues and a few renal tubules, whereas the posterior kidney contained more renal tubules. Although the glomerular structure was present in P. boro, light microscopic observations identified no glomeruli in the kidney of H. barbouri and A. grunniens. Ultrastructurally, abundant deep basal infoldings with mitochondria in the renal tubules were observed in A. grunniens compared to H. barbouri and P. boro, suggesting the possible role of basal infoldings in maintaining the osmotic balance. By integrating the results from the three species and comprehensive literature search, the authors further showed that 56 species have been reported to be aglomerular, and that the aglomerular kidney has evolved at least eight times in bony fishes.

Spectrum of Kidney Injury Following COVID-19 Disease: Renal Biopsy Findings in a Single Italian Pathology Service.

The onset of coronavirus disease (COVID-19) as a pandemic infection, has led to increasing insights on its pathophysiology and clinical features being revealed, such as a noticeable kidney involvement. In this study, we describe the histopathological, immunofluorescence, and ultrastructural features of biopsy-proven kidney injury observed in a series of SARS-CoV-2 positive cases in our institution from April 2020 to November 2021. We retrieved and retrospectively reviewed nine cases (two pediatric and seven adults) that experienced nephrotic syndrome (six cases), acute kidney injury (two cases), and a clinically silent microhematuria and leukocyturia. Kidney biopsies were investigated by means of light microscopy, direct immunofluorescence, and electron microscopy. The primary diagnoses were minimal change disease (four cases), acute tubular necrosis (two cases), collapsing glomerulopathy (two cases), and C3 glomerulopathy (one case). None of the cases showed viral or viral-like particles on ultrastructural analysis. Novel and specific histologic features on kidney biopsy related to SARS-CoV-2 infection have been gradually disclosed and reported, harboring relevant clinical and therapeutic implications. Recognizing and properly diagnosing renal involvement in patients experiencing COVID-19 could be challenging (due to the lack of direct proof of viral infection, e.g., viral particles) and requires a proper integration of clinical and pathological data.

Microtubule-associated protein tau in murine kidney: role in podocyte architecture.

Tau is a cytoskeletal protein that is expressed mainly in neurons and is involved in several cellular processes, such as microtubule stabilization, axonal maintenance, and transport. Altered tau metabolism is related to different tauopathies being the most important Alzheimer's disease where aberrant hyperphosphorylated and aggregated tau is found in the central nervous system. Here, we have analyzed that function in kidney by using tau knockout mice generated by integrating GFP-encoding cDNA into exon 1 of MAPT (here referred to as TauGFP/GFP). IVIS Lumina from PerkinElmer demonstrated GFP expression in the kidney. We then demonstrated by qPCR that the main tau isoform in the kidney is Tau4R. The GFP reporter allowed us to demonstrate that tau is found in the glomeruli of the renal cortex, and specifically in podocytes. This was further confirmed by immunohistochemistry. TauGFP/GFP mice present a podocyte cytoskeleton more dynamic as they contain higher levels of detyrosinated tubulin than wild-type mice. In addition, transmission electron microscopy studies demonstrated glomerular damage with a decrease in urinary creatinine. Our results prove that tau has an important role in kidney metabolism under normal physiological conditions.

Protective effect of H2S on LPS­induced AKI by promoting autophagy.

The present study explored the protective effect of exogenous hydrogen sulfide (H2S) on lipopolysaccharide (LPS)­induced acute kidney injury (AKI) and the underlying mechanisms. To establish an AKI injury mouse model, LPS (10 mg/kg) was intraperitoneally injected into mice pretreated with 0.8 mg/kg sodium hydrosulfide hydrate (NaHS), an H2S donor. The mouse survival rate and the degree of kidney injury were examined. To construct a cell damage model, HK­2 cells were pretreated with different concentrations (0.1, 0.3 and 0.5 mM) of NaHS, and then the cells were stimulated with LPS (1 µg/ml). The cell viability, autophagy, apoptosis levels and the release of inflammatory factors were examined in mouse kidney tissue and HK­2 renal tubular epithelial cells. It was found that pretreatment with NaHS significantly improved the survival rate of septic AKI mice, and reduced the renal damage, release of inflammatory factors and apoptosis. In HK­2 cells, NaHS protected cells from LPS caused damage via promoting autophagy and inhibiting apoptosis and the release of inflammatory factors. In order to clarify the relationship between autophagy and apoptosis and inflammatory factors, this study used 3­methyladenine (3­MA) to inhibit autophagy. The results revealed that 3­MA eliminated the protective effect of NaHS in HK­2 cells and AKI mice. Overall, NaHS can protect from LPS­induced AKI by promoting autophagy and inhibiting apoptosis and the release of inflammatory factors.

Evaluation of ultrastructural alterations of glomerular basement membrane and podocytes in glomeruli by low-vacuum scanning electron microscopy.

BACKGROUND: Low-vacuum scanning electron microscopy (LV-SEM) is applied to diagnostic renal pathology. METHODS: To demonstrate the usefulness of LV-SEM and to clarify the optimal conditions of pathology samples, we investigated the alterations of glomerular basement membrane (GBM) and podocytes in control and experimental active Heymann nephritis (AHN) rats by LV-SEM. RESULTS: On week 15 following induction of AHN, spike formation on GBM with diffuse deposition of IgG and C3 developed. Using LV-SEM, diffuse crater-like protrusions were clearly noted three-dimensionally (3D) on surface of GBM in the same specimens of light microscopy (LM) and immunofluorescence (IF) studies only after removal coverslips or further adding periodic acid-silver methenamine (PAM) staining. These 3D ultrastructural findings of GBM surface could be detected in PAM-stained specimens by LV-SEM, although true GBM surface findings could not be obtained in acellular glomeruli, because some subepithelial deposits remained on surface of GBM. Adequate thickness was 1.5-5 µm for 10% formalin-fixed paraffin-embedded (FFPE) and 5-10 µm for the unfixed frozen sections. The foot processes and their effacement of podocytes could be observed by LV-SEM using 10%FFPE specimens with platinum blue (Pt-blue) staining or double staining of PAM and Pt-blue. These findings were obtained more large areas in 2.5% glutaraldehyde-fixed paraffin-embedded (2.5%GFPE) specimens. CONCLUSION: Our findings suggest that LV-SEM is a useful assessment tool for evaluating the alterations of GBM and podocytes in renal pathology using routine LM and IF specimens, as well as 2.5%GFPE specimens.

Gitelman-Like Syndrome Caused by Pathogenic Variants in mtDNA.

BACKGROUND: Gitelman syndrome is the most frequent hereditary salt-losing tubulopathy characterized by hypokalemic alkalosis and hypomagnesemia. Gitelman syndrome is caused by biallelic pathogenic variants in SLC12A3, encoding the Na+-Cl- cotransporter (NCC) expressed in the distal convoluted tubule. Pathogenic variants of CLCNKB, HNF1B, FXYD2, or KCNJ10 may result in the same renal phenotype of Gitelman syndrome, as they can lead to reduced NCC activity. For approximately 10 percent of patients with a Gitelman syndrome phenotype, the genotype is unknown. METHODS: We identified mitochondrial DNA (mtDNA) variants in three families with Gitelman-like electrolyte abnormalities, then investigated 156 families for variants in MT-TI and MT-TF, which encode the transfer RNAs for phenylalanine and isoleucine. Mitochondrial respiratory chain function was assessed in patient fibroblasts. Mitochondrial dysfunction was induced in NCC-expressing HEK293 cells to assess the effect on thiazide-sensitive 22Na+ transport. RESULTS: Genetic investigations revealed four mtDNA variants in 13 families: m.591C>T (n=7), m.616T>C (n=1), m.643A>G (n=1) (all in MT-TF), and m.4291T>C (n=4, in MT-TI). Variants were near homoplasmic in affected individuals. All variants were classified as pathogenic, except for m.643A>G, which was classified as a variant of uncertain significance. Importantly, affected members of six families with an MT-TF variant additionally suffered from progressive chronic kidney disease. Dysfunction of oxidative phosphorylation complex IV and reduced maximal mitochondrial respiratory capacity were found in patient fibroblasts. In vitro pharmacological inhibition of complex IV, mimicking the effect of the mtDNA variants, inhibited NCC phosphorylation and NCC-mediated sodium uptake. CONCLUSION: Pathogenic mtDNA variants in MT-TF and MT-TI can cause a Gitelman-like syndrome. Genetic investigation of mtDNA should be considered in patients with unexplained Gitelman syndrome-like tubulopathies.

C4d Deposition after Allogeneic Renal Transplantation in Rats Is Involved in Initial Apoptotic Cell Clearance.

In the context of transplantation, complement activation is associated with poor prognosis and outcome. While complement activation in antibody-mediated rejection is well-known, less is known about complement activation in acute T cell-mediated rejection (TCMR). There is increasing evidence that complement contributes to the clearance of apoptotic debris and tissue repair. In this regard, we have analysed published human kidney biopsy transcriptome data clearly showing upregulated expression of complement factors in TCMR. To clarify whether and how the complement system is activated early during acute TCMR, experimental syngeneic and allogeneic renal transplantations were performed. Using an allogeneic rat renal transplant model, we also observed upregulation of complement factors in TCMR in contrast to healthy kidneys and isograft controls. While staining for C4d was positive, staining with a C3d antibody showed no C3d deposition. FACS analysis of blood showed the absence of alloantibodies that could have explained the C4d deposition. Gene expression pathway analysis showed upregulation of pro-apoptotic factors in TCMR, and apoptotic endothelial cells were detected by ultrastructural analysis. Monocytes/macrophages were found to bind to and phagocytise these apoptotic cells. Therefore, we conclude that early C4d deposition in TCMR may be relevant to the clearance of apoptotic cells.

Molecular Studies on the Nephroprotective Potential of Celastrus paniculatus against Lead-Acetate-Induced Nephrotoxicity in Experimental Rats: Role of the PI3K/AKT Signaling Pathway.

Chemicals can induce nephrotoxicity, with damage to different segments of the nephron and deterioration of renal function. Nephrotoxicity due to exposure to a toxin such as carbon tetrachloride, sodium oxalate, or heavy metals is the most common cause of kidney injury. The current study aimed to evaluate the protective effects of Celastrus paniculatus seed extract against lead-acetate-induced nephrotoxicity by evaluating the histopathology, immunohistochemistry, ultrastructure, and phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway. Twenty-four rats were divided into four groups (n = 6 per group): group 1 contained normal animals and served as the control; group 2 received lead acetate (30 mg/kg body weight (b.w.)/day, oral); group 3 received lead acetate and the standard drug N-acetylcysteine (NAC, 200 mg/kg b.w./day, oral); and group 4 received lead acetate and the ethanolic extract of C. paniculatus seed (EECP; 800 mg/kg b.w./day, oral). Treatment was given for 28 consecutive days. The data were analyzed using one-way analysis of variance with SIGMA PLOT 13 using SYSTAT software followed by Newman-Keul's test for comparison between the groups. EECP ameliorated the adverse changes caused by lead acetate. PI3K and AKT messenger RNA (mRNA) levels were diminished in lead-acetate-treated rats. Treatment with EECP inhibited the occurrence of shrunken cells, the atrophy of glomeruli, and degenerative changes in renal tubules caused by lead acetate. Interestingly, the PI3K and AKT mRNA levels were significantly increased in EECP-treated animals. Our results clearly evidence for the first time that C. paniculatus seed extract inhibits lead-acetate-induced detrimental changes in kidneys by regulating PI3K/AKT signaling pathways.

AMPK agonist alleviate renal tubulointerstitial fibrosis via activating mitophagy in high fat and streptozotocin induced diabetic mice.

Renal tubulointerstitial fibrosis was a crucial pathological feature of diabetic nephropathy (DN), and renal tubular injury might associate with abnormal mitophagy. In this study, we investigated the effects and molecular mechanisms of AMPK agonist metformin on mitophagy and cellular injury in renal tubular cell under diabetic condition. The high fat diet (HFD) and streptozotocin (STZ)-induced type 2 diabetic mice model and HK-2 cells were used in this study. Metformin was administered in the drinking water (200 mg/kg/d) for 24 weeks. Renal tubulointerstitial lesions, oxidative stress and some indicators of mitophagy (e.g., LC3II, Pink1, and Parkin) were examined both in renal tissue and HK-2 cells. Additionally, compound C (an AMPK inhibitor) and Pink1 siRNA were applied to explore the molecular regulation mechanism of metformin on mitophagy. We found that the expression of p-AMPK, Pink1, Parkin, LC3II, and Atg5 in renal tissue of diabetic mice was decreased obviously. Metformin reduced the levels of serum creatinine, urine protein, and attenuated renal oxidative injury and fibrosis in HFD/STZ induced diabetic mice. In addition, Metformin reversed mitophagy dysfunction and the over-expression of NLRP3. In vitro pretreatment of HK-2 cells with AMPK inhibitor compound C or Pink1 siRNA negated the beneficial effects of metformin. Furthermore, we noted that metformin activated p-AMPK and promoted the translocation of Pink1 from the cytoplasm to mitochondria, then promoted the occurrence of mitophagy in HK-2 cells under HG/HFA ambience. Our results suggested for the first time that AMPK agonist metformin ameliorated renal oxidative stress and tubulointerstitial fibrosis in HFD/STZ-induced diabetic mice via activating mitophagy through a p-AMPK-Pink1-Parkin pathway.

LAMA2 and LOXL4 are candidate FSGS genes.

BACKGROUND: Focal and segmental glomerulosclerosis (FSGS) is a histologic pattern of injury that characterizes a wide spectrum of diseases. Many genetic causes have been identified in FSGS but even in families with comprehensive testing, a significant proportion remain unexplained. METHODS: In a family with adult-onset autosomal dominant FSGS, linkage analysis was performed in 11 family members followed by whole exome sequencing (WES) in 3 affected relatives to identify candidate genes. RESULTS: Pathogenic variants in known nephropathy genes were excluded. Subsequently, linkage analysis was performed and narrowed the disease gene(s) to within 3% of the genome. WES identified 5 heterozygous rare variants, which were sequenced in 11 relatives where DNA was available. Two of these variants, in LAMA2 and LOXL4, remained as candidates after segregation analysis and encode extracellular matrix proteins of the glomerulus. Renal biopsies showed classic segmental sclerosis/hyalinosis lesion on a background of mild mesangial hypercellularity. Examination of basement membranes with electron microscopy showed regions of dense mesangial matrix in one individual and wider glomerular basement membrane (GBM) thickness in two individuals compared to historic control averages. CONCLUSIONS: Based on our findings, we postulate that the additive effect of digenic inheritance of heterozygous variants in LAMA2 and LOXL4 leads to adult-onset FSGS. Limitations to our study includes the absence of functional characterization to support pathogenicity. Alternatively, identification of additional FSGS cases with suspected deleterious variants in LAMA2 and LOXL4 will provide more evidence for disease causality. Thus, our report will be of benefit to the renal community as sequencing in renal disease becomes more widespread.

Impact of cypermethrin in nephrocytes of freshwater fish Catla catla.

The kidney of Catla catla, chronically exposed to sub-lethal concentrations (0.24 µg/L and 0.41 µg/L) of cypermethrin revealed a significant elevation in the activity of antioxidants superoxide dismutase (SOD), catalase (CAT), glutathione-s-transferase (GST) and reduced glutathione (GSH) after 15 days, followed by a decline of up to 45 days. Lipid peroxidation (LPO) remained elevated throughout the exposure duration. Histology presented proliferated haematopoietic tissue, tubular and glomerular degeneration. The maximum increase in the mean degree of tissue change (DTC) was observed on the 45th day of treatment. Ultra-structure study depicted cytoplasmic vacuolation, fragmented RER, the proliferation of lysosomes, mitochondrial degeneration, and degenerative changes in the epithelial lining of renal tubules. Principal component analysis (PCA) of various biomarkers generated two components PCI (SOD, GST, GSH, LPO and DTC) and PCII (CAT). These findings suggest that long term exposure to cypermethrin can lead to various pathological alterations in the fish kidney which in turn might interfere with normal renal excretory mechanism.

Nephroprotective effect of naringin in methotrexate induced renal toxicity in male rats.

The current work aims to study the nephroprotective potential of naringin (NG), a flavanone derived from citrus fruits, in methotrexate (MTX)-induced renal toxicity. Thirty male rats were divided into five groups; control group (IP saline), MTX group (IP single dose, 20 mg/kg), and three groups co-treated with MTX and naringin (IP daily dose; 20, 40, and 80 mg/kg, respectively). Kidney tissues were used to investigate renal function, oxidative stress, lipid peroxidation, and caspase-3 activity. Biochemical cytokine analysis was performed in addition to ultrastructural examinations of kidney tissue. When compared to the MTX-treated rats, MTX+NG significantly reduced the levels of urea, creatinine, MDA, NO, TNFα, IL-6, and caspase-3 activity. A significant increase in the levels of the antioxidant enzymes and GSH were also noted. Additionally, naringin ameliorated the apparent ultrastructural changes observed in the glomeruli and renal tubules of MTX-intoxicated rats. Noticeable structural improvements of glomerular lesions, proximal, and distal convoluted tubular epithelium were observed in MTX+NG treated animals, including podocytes with regular foot processes, perfectly organized filtration barrier, no signs of GBM thickening, organized brush border, and normal architecture of microvilli. Naringin (80 mg/kg) had the maximum amelioration effect. To the best of our knowledge, this is the first study to investigate the ultrastructural manifestations of naringin and/or MTX on the kidney of rats. Taken all, naringin has a potent therapeutic effect and can be used in adjuvant therapy to prevent MTX-induced nephrotoxicity. Nevertheless, the molecular mechanism underlying the nephroprotective capacity of naringin needs further investigation.

Gaia Pigino: Inside the cell.

Gaia Pigino studies the molecular mechanisms and principles of self-organization in cilia using 3D cryo-EM.

Decellularization of kidney tissue: comparison of sodium lauryl ether sulfate and sodium dodecyl sulfate for allotransplantation in rat.

An automatic decellularization device was developed to perfuse and decellularize male rats' kidneys using both sodium lauryl ether sulfate (SLES) and sodium dodecyl sulfate (SDS) and to compare their efficacy in kidney decellularization and post-transplantation angiogenesis. Kidneys were perfused with either 1% SDS solution for 4 h or 1% SLES solution for 6 h. The decellularized scaffolds were stained with hematoxylin and eosin, periodic acid Schiff, Masson's trichrome, and Alcian blue to determine cell removal and glycogen, collagen, and glycosaminoglycan contents, respectively. Moreover, scanning electron microscopy was performed to evaluate the cell removal and preservation of microarchitecture of both SDS and SLES scaffolds. Additionally, DNA quantification assay was applied for all groups in order to measure residual DNA in the scaffolds and normal kidney. In order to demonstrate biocompatibility of the decellularized scaffolds, human umbilical cord mesenchymal stromal/stem cells (hUC-MSCs) were seeded on the scaffolds. In addition, the allotransplantation was performed in back muscle and angiogenesis was evaluated. Complete cell removal in both SLES and SDS groups was observed in scanning electron microscopy and DNA quantification assays. Moreover, the extracellular matrix (ECM) architecture of rat kidney in the SLES group was significantly preserved better than the SDS group. The hUC-MSCs were successfully migrated from the cell culture plate surface into the SDS and SLES decellularized scaffolds. The formation of blood vessels was observed in the kidney in both SLES and SDS decellularized kidneys. The better preservation of ECM than SDS introduces SLES as the solvent of choice for kidney decellularization.

Melatonin Treatment Improves Renal Fibrosis via miR-4516/SIAH3/PINK1 Axis.

Dysregulation in mitophagy, in addition to contributing to imbalance in the mitochondrial dynamic, has been implicated in the development of renal fibrosis and progression of chronic kidney disease (CKD). However, the current understanding of the precise mechanisms behind the pathogenic loss of mitophagy remains unclear for developing cures for CKD. We found that miR-4516 is downregulated and its target SIAH3, an E3 ubiquitin protein ligase that reduces PINK1 accumulation to damaged mitochondria, is upregulated in the renal cortex of CKD mice. Here, we demonstrated that melatonin injection induces miR-4516 expression and suppresses SIAH3, and promotes PINK1/Parkin-mediated mitophagy. Furthermore, we demonstrated that melatonin injection attenuates the pathological features of CKD by improving mitochondrial homeostasis. Our data supports that mitochondrial autophagy regulation by activating miR-4516/SIAH3/PINK1 mitophagy signaling axis can be a viable new strategy for treating CKD.

Kidney involvement and histological findings in two pediatric COVID-19 patients.

BACKGROUND: Histological findings of kidney involvement have been rarely reported in pediatric patients with SARS-CoV-2 infection. Here, we describe clinical, laboratory, and histological findings of two pediatric cases with almost exclusive kidney involvement by SARS-CoV-2. RESULTS: A 10-year-old girl with IgA vasculitis nephritis underwent kidney biopsy, showing diffuse and segmental mesangial-proliferative glomerulonephritis, and steroid therapy was initiated. After the worsening of the clinical picture, including an atypical skin rash, she was diagnosed with SARS-CoV-2. The re-evaluation of initial biopsy showed cytoplasmatic blebs and virus-like particles in tubular cells at electron microscopy. Despite SARS-CoV-2 clearance and the intensification of immunosuppression, no improvement was observed. A second kidney biopsy showed a crescentic glomerulonephritis with sclerosis, while virus-like particles were no longer evident. The second patient was a 12-year-old girl with a 3-week history of weakness and weight loss. Rhinitis was reported the month before. No medications were being taken. Blood and urine analysis revealed elevated serum creatinine, hypouricemia, low molecular weight proteinuria, and glycosuria. A high SARS-CoV-2-IgG titre was detected. Kidney biopsy showed acute tubular-interstitial nephritis. Steroid therapy was started with a complete resolution of kidney involvement. CONCLUSION: We can speculate that in both cases SARS-CoV-2 played a major role as inflammatory trigger of the kidney damage. Therefore, we suggest investigating the potential kidney damage by SARS-CoV-2 in children. Moreover, SARS-CoV-2 can be included among infectious agents responsible for pediatric acute tubular interstitial nephritis.

miR­214 ameliorates sepsis­induced acute kidney injury via PTEN/AKT/mTOR­regulated autophagy.

Previous studies have suggested that oxidative stress and autophagy results in acute kidney injury (AKI) during sepsis and microRNA (miR)­214 serves a vital role in the protection of kidneys subjected to oxidative stress. The present study aimed to test whether the renoprotection of miR­214 is related to autophagy in sepsis. The role of autophagy was investigated in a mouse model of cecal ligation and puncture (CLP). Reverse transcription­quantitative polymerase chain reaction (RT­qPCR) was used to analyze the expression of miR­214. The structure and function of kidneys harvested from the mice were evaluated. Kidney autophagy levels were detected with immunohistochemical, immunofluorescent and western blotting. It was found that miR­214 could alleviate AKI in septic mice by inhibiting the level of kidney autophagy. Furthermore, miR­214 inhibited autophagy by silencing PTEN expression in the kidney tissues of septic mice. These findings indicated that miR­214 ameliorated CLP­induced AKI by reducing oxidative stress and inhibiting autophagy through the regulation of the PTEN/AKT/mTOR pathway.