Glomerular Basement Membrane Thickening Research Articles

Expert consensus on the application of rituximab in the treatment of membranous nephropathy

Membranous nephropathy is a histopathological type of renal disease, characterized by the thickening of glomerular basement membrane caused by an accumulation of immune deposits on the outer aspect of the glomerular basement membrane. Its main clinical manifestations are nephrotic syndrome or asymptomatic proteinuria. Rituximab is a murine/human chimeric anti-CD20 monoclonal antibody that depletes B cells. In recent years, rituximab has been applied to the treatment of membranous nephropathy and achieved good effects. The nephrology expert panel of the Peking University Health Science Center formed this consensus by searching PubMed, China National Knowledge Infrastructure database, Wanfang database, VIP database and other relevant literature, and combining the current situation in China, so as to standardize the clinical application of rituximab in the treatment of membranous nephropathy and provide practical guidance for clinicians.

Protective effects of Moutan Cortex polysaccharides components on renal injury in diabetic nephropathy rats

This study investigated the protective effects of Moutan Cortex polysaccharides components(MCPC) on the renal tissues of diabetic nephropathy(DN) rats and explored their regulation effect on inflammatory response and oxidative stress. The DN rat model was induced by high-glucose and high-fat diet combined with streptozotocin(STZ), and then the rats were randomly divided into control group, model group, positive group and MCPC high(120 mg·kg~(-1)·d~(-1)), low(60 mg·kg~(-1)·d~(-1)) dose groups. After 12 weeks treatment, blood was taken from the orbit of the rats, and then they were sacrificed before the kidney tissues were collected. The serum and tissues were detected for related biochemical indicators and pathological changes of the kidney. Immunohistochemical methods were used to determine the expression of FN and ColⅣ in the kidney tissue of DN rats. Compared with the model group, blood glucose, serum creatinine, blood urea nitrogen and 24 h urine protein in the MCPC high-dose group were significantly reduced(P<0.01). The results of HE, PAS, Masson staining showed that glomerular basement membrane thickening, Bowman's capsule narrowing and inflammatory cell infiltration in DN rats were improved in the MCPC high-dose group; the activity of T-SOD and GSH-Px in serum significantly increased(P<0.001), and the expression level of FN significantly decreased(P<0.001). The high-dose MCPC treatment could effectively inhibit the abnormal expression of Col Ⅳ(P<0.001) and significantly reduce the levels of AGEs and RAGE in serum(P<0.001), the content of VCAM-1 and IL-1β in serum(P<0.001), and the levels of IL-1β mRNA in kidney tissue(P<0.001), but failed to effectively reduce VCAM-1 mRNA levels in kidney tissues. The high-dose MCPC could significantly improve pathological injury of renal tissue and related renal indicators in DN rats, and achieve renal protection in DN rats mainly by regulating oxidative stress and inflammatory factors.

Pdx1 Knockout Leads to a Diabetic Nephropathy- Like Phenotype in Zebrafish and Identifies Phosphatidylethanolamine as Metabolite Promoting Early Diabetic Kidney Damage.

The pdx1-/- zebrafish mutant was recently established as a novel animal model of diabetic retinopathy. In this study, we investigate whether knockout of pdx1 also leads to diabetic kidney disease (DKD). pdx1-/- larvae exhibit several signs of early DKD, such as glomerular hypertrophy, impairments in the filtration barrier corresponding to microalbuminuria, and glomerular basement membrane (GBM) thickening. Adult pdx1-/- mutants show progressive GBM thickening in comparison with the larval state. Heterozygous pdx1 knockout also leads to glomerular hypertrophy as initial establishment of DKD similar to the pdx1-/- larvae. RNA sequencing of adult pdx1+/- kidneys uncovered regulations in multiple expected diabetic pathways related to podocyte disruption and hinting at early vascular dysregulation without obvious morphological alterations. Metabolome analysis and pharmacological intervention experiments revealed the contribution of phosphatidylethanolamine in the early establishment of kidney damage. In conclusion, this study identified the pdx1 mutant as a novel model for the study of DKD, showing signs of the early disease progression already in the larval stage and several selective features of later DKD in adult mutants.

A Rationalized Overview of Diabetic Nephropathy

Diabetic nephropathy is a prime motive of end-level renal disease, and there was a non-stop boom in its occurrence global with inside the beyond decades. Diabetic nephropathy is characterized via way of means of microalbuminuria, renal and glomerular hypertrophy, mesangial growth with glomerular basement membrane thickening, arteriolar hyalinosis, and international glomerular sclerosis, which in the long run motive the development of proteinuria and renal failure. The time period diabetic kidney disease (DKD) is now normally used to embody the spectrum of human beings with diabetes who've both albuminuria and discounts in renal function. In this article, the medical presentation and method to prognosis of DKD might be discussed, as will its prognosis. The popular ideas of control of DKD can also be reviewed close to modern-day global guidelines. The present review deal with brief about DN pathophysiology, prevention measures along with treatment approaches.

A comparative study of human glomerular basement membrane thickness using direct measurement and orthogonal intercept methods

IntroductionHere we report estimates of glomerular basement membrane (GBM) thickness in the Brazilian population performed using direct (DM) and orthogonal interception methods (OIM), and comment on potential sources of variation among estimates made by different laboratories.MethodologyA total of 38 patients, ranging from 3 to 78 years of age, 26 (68%) males and 12 (32%) females, were submitted to kidney biopsy procedures for renal disease diagnosis. Glomeruli were diagnosed with minor histological changes by conventional, immunofluorescence and electron microscopy. GBM thickness was estimated using both DM and OIM methods.ResultsEstimates of GBM thickness obtained using DM were higher than those obtained by OIM. However, the application of a correction for non-perpendicular membrane sectioning to DM estimates yielded similar results to those obtained under OIM. The estimated GMB thickness using DM after correction was 289 + 44 nm, versus 287 + 48 nm by OIM. No statistically significant differences were detected in GMB thickness, nor with respect to patient age or sex.ConclusionsGBM thickness in the studied Brazilian population measured approximately 290 nm. The application of criteria for estimating the shortest distance between the endothelial and podocyte cell membranes with correction for non-perpendicular membrane sectioning can increase the accuracy of GBM thickness estimates using DM and OIM.

Therapeutic potential of pro-resolving mediators in diabetic kidney disease.

Renal microvascular disease associated with diabetes [Diabetic kidney disease - DKD] is the leading cause of chronic kidney disease. In DKD, glomerular basement membrane thickening, mesangial expansion, endothelial dysfunction, podocyte cell loss and renal tubule injury contribute to progressive glomerulosclerosis and tubulointerstitial fibrosis. Chronic inflammation is recognized as a major pathogenic mechanism for DKD, with resident and circulating immune cells interacting with local kidney cell populations to provoke an inflammatory response. The onset of inflammation is driven by the release of well described proinflammatory mediators, and this is typically followed by a resolution phase. Inflammation resolution is achieved through the bioactions of endogenous specialized pro-resolving lipid mediators (SPMs). As our understanding of SPMs advances 'resolution pharmacology' based approaches using these molecules are being explored in DKD.

Annexin 1 Mimetic Ac2-26 Holds Promise for the Treatment of Diabetic Nephropathy.

In this issue of Diabetes , Wu et al. (1) report that in a high-fat diet/streptozotocin–induced diabetes model, global annexin 1 (ANAX1) deletion exacerbates diabetic nephropathy (DN), as indicated by increased albuminuria and macrophage infiltration, increased glomerular size, mesangial expansion, glomerular basement membrane thickness, and foot process effacement, increased tubulointerstitial injury, and fibrosis. They also detected lipid accumulation and mitochondrial abnormalities in the proximal tubule epithelial cells. RNA-sequencing analysis revealed a role of ANAX1 in lipid metabolism in the HFD/STZ-induced DN mice. Using a human immortalized proximal tubule epithelial cell line, HK-2 cells, they found that ANAX1 knockdown impaired mitochondrial fatty acid oxidation via a FPR2/AMPK/PPARα/CPT2b signaling pathway, leading to reduced intracellular lipid accumulation. ANAX1 knockdown also aggravated the lipid disorders in podocytes. Finally, administration of Ac2-26, an ANAX1 mimetic, ameliorated renal …

LAMA2 and LOXL4 are candidate FSGS genes

BackgroundFocal 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.MethodsIn 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.ResultsPathogenic 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.ConclusionsBased 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.

Transforming growth factor β3 deficiency promotes defective lipid metabolism and fibrosis in murine kidney.

ABSTRACTGlomerulosclerosis and tubulointerstitial fibrosis are pathological features of chronic kidney disease. Transforming growth factor β (TGFβ) is a key player in the development of fibrosis. However, of the three known TGFβ isoforms, only TGFβ1 has an established role in fibrosis, and the pathophysiological relevance of TGFβ2 and TGFβ3 is unknown. Because Tgfb3 deficiency in mice results in early postnatal lethality, we analyzed the kidney phenotype of heterozygous Tgfb3-knockout mice (Tgfb3+/−) and compared it with that of matched wild-type mice. Four-month-old Tgfb3+/− mice exhibited incipient renal fibrosis with epithelial–mesenchymal transition, in addition to glomerular basement membrane thickening and podocyte foot process effacement associated with albuminuria. Also evident was insulin resistance and oxidative stress at the renal level, together with aberrant renal lipid metabolism and mitochondrial function. Omics analysis revealed toxic species, such as diacylglycerides and ceramides, and dysregulated mitochondrial metabolism in Tgfb3+/− mice. Kidneys of Tgfb3+/− mice showed morphological alterations of mitochondria and overactivation of non-canonical MAPK ERK1/2 and JNK cascades. Our study indicates that renal TGFβ3 might have antifibrotic and renoprotective properties, opposing or counteracting the activity of TGFβ1. This article has an associated First Person interview with the first author of the paper.

Reduced Acrolein Detoxification in akr1a1a Zebrafish Mutants Causes Impaired Insulin Receptor Signaling and Microvascular Alterations.

Increased acrolein (ACR), a toxic metabolite derived from energy consumption, is associated with diabetes and its complications. However, the molecular mechanisms are mostly unknown, and a suitable animal model with internal increased ACR does not exist for in vivo studying so far. Several enzyme systems are responsible for acrolein detoxification, such as Aldehyde Dehydrogenase (ALDH), Aldo‐Keto Reductase (AKR), and Glutathione S‐Transferase (GST). To evaluate the function of ACR in glucose homeostasis and diabetes, akr1a1a−/− zebrafish mutants are generated using CRISPR/Cas9 technology. Accumulated endogenous acrolein is confirmed in akr1a1a−/− larvae and livers of adults. Moreover, a series of experiments are performed regarding organic alterations, the glucose homeostasis, transcriptome, and metabolomics in Tg(fli1:EGFP) zebrafish. Akr1a1a−/− larvae display impaired glucose homeostasis and angiogenic retina hyaloid vasculature, which are caused by reduced acrolein detoxification ability and increased internal ACR concentration. The effects of acrolein on hyaloid vasculature can be reversed by acrolein‐scavenger l‐carnosine treatment. In adult akr1a1a−/− mutants, impaired glucose tolerance accompanied by angiogenic retina vessels and glomerular basement membrane thickening, consistent with an early pathological appearance in diabetic retinopathy and nephropathy, are observed. Thus, the data strongly suggest impaired ACR detoxification and elevated ACR concentration as biomarkers and inducers for diabetes and diabetic complications.

Sinigrin attenuates angiotensin II‑induced kidney injury by inactivating nuclear factor‑κB and extracellular signal‑regulated kinase signaling invivo and invitro.

The present study investigated the function of sinigrin in angiotensin II (Ang II)-induced renal damage. The results demonstrated that systolic blood pressure (SBP) and diastolic blood pressure (DBP) were increased in Ang II-challenged rats, and sinigrin treatment inhibited their increase. The levels of blood urea nitrogen (BUN) and serum creatinine (SCR) were increased by Ang II in the rats, and these were reversed by sinigrin in a dose-dependent manner. In addition, the Ang II-induced elevation of urinary protein levels was inhibited by sinigrin treatment. Glomerular basement membrane thickness and ECM degradation markers, such as collagen I, collagen IV and fibronectin, were suppressed by sinigrin in the Ang II-challenged rats. Moreover, the levels of inflammatory regulators, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1), were reduced following sinigrin treatment of the Ang II-challenged rats and in Ang II-exposed proximal tubule epithelial cells. Furthermore, the superoxide dismutase (SOD) and catalase (CAT) levels were downregulated, whereas the malondialdehyde (MDA) levels were upregulated by Ang II; these effects were reversed by sinigrin treatment in vivo and in vitro. Mechanistically, sinigrin inhibited the Ang II-induced phosphorylation of ERK, p65 and IκBα. Thus, sinigrin attenuated Ang II-induced renal injury by inactivating ERK and NF-κB signaling. Sinigrin may thus prove to be a potential candidate for the treatment of hypertension-induced kidney damage.

Potential relationship between eGFRcystatin C /eGFRcreatinine -ratio and glomerular basement membrane thickness in diabetic kidney disease.

Diabetic kidney disease (DKD) is a leading cause of end‐stage renal disease and renal replacement therapy worldwide. A pathophysiological hallmark of DKD is glomerular basal membrane (GBM) thickening, whereas this feature is absent in minimal change disease (MCD). According to fundamental transport physiological principles, a thicker GBM will impede the diffusion of middle‐molecules such as cystatin C, potentially leading to a lower estimated GFR (eGFR) from cystatin C compared to that of creatinine. Here we test the hypothesis that thickening of the glomerular filter leads to an increased diffusion length, and lower clearance, of cystatin C. Twenty‐nine patients with a kidney biopsy diagnosis of either DKD (n = 17) or MCD (n = 12) were retrospectively included in the study. GBM thickness was measured at 20 separate locations in the biopsy specimen and plasma levels of cystatin C and creatinine were retrieved from health records. A modified two‐pore model was used to simulate the effects of a thicker GBM on glomerular water and solute transport. The mean age of the patients was 52 years, and 38% were women. The mean eGFRcystatin C/eGFRcreatinine‐ratio was 74% in DKD compared to 98% in MCD (p < 0.001). Average GBM thickness was strongly inversely correlated to the eGFRcystatin C/eGFRcreatinine‐ratio (Pearson's r = −0.61, p < 0.01). Two‐pore modeling predicted a eGFRcystatin C/eGFRcreatinine‐ratio of 78% in DKD. We provide clinical and theoretical evidence suggesting that thickening of the glomerular filter, increasing the diffusion length of cystatin C, lowers the eGFRcystatin C/eGFRcreatinine‐ratio in DKD.

The AGE receptor, OST48 drives podocyte foot process effacement and basement membrane expansion (alters structural composition).

AimsThe accumulation of advanced glycation end products is implicated in the development and progression of diabetic kidney disease. No study has examined whether stimulating advanced glycation clearance via receptor manipulation is reno‐protective in diabetes. Podocytes, which are early contributors to diabetic kidney disease and could be a target for reno‐protection.Materials and methodsTo examine the effects of increased podocyte oligosaccharyltransferase‐48 on kidney function, glomerular sclerosis, tubulointerstitial fibrosis and proteome (PXD011434), we generated a mouse with increased oligosaccharyltransferase‐48kDa subunit abundance in podocytes driven by the podocin promoter.ResultsDespite increased urinary clearance of advanced glycation end products, we observed a decline in renal function, significant glomerular damage including glomerulosclerosis, collagen IV deposition, glomerular basement membrane thickening and foot process effacement and tubulointerstitial fibrosis. Analysis of isolated glomeruli identified enrichment in proteins associated with collagen deposition, endoplasmic reticulum stress and oxidative stress. Ultra‐resolution microscopy of podocytes revealed denudation of foot processes where there was co‐localization of oligosaccharyltransferase‐48kDa subunit and advanced glycation end‐products.ConclusionsThese studies indicate that increased podocyte expression of oligosaccharyltransferase‐48 kDa subunit results in glomerular endoplasmic reticulum stress and a decline in kidney function.

Vaccination Against Receptor for Advanced Glycation End Products Attenuates the Progression of Diabetic Kidney Disease.

Effective treatment of diabetic kidney disease (DKD) remains a large unmet medical need. Within the disease's complicated pathogenic mechanism, activation of the advanced glycation end products (AGEs)-receptor for AGE (RAGE) axis plays a pivotal role in the development and progression of DKD. To provide a new therapeutic strategy against DKD progression, we developed a vaccine against RAGE. Three rounds of immunization of mice with the RAGE vaccine successfully induced antigen-specific serum IgG antibody titers and elevated antibody titers were sustained for at least 38 weeks. In addition, RAGE vaccination significantly attenuated the increase in urinary albumin excretion in streptozotocin-induced diabetic mice (type 1 diabetes model) and leptin-receptor-deficient db/db mice (type 2 diabetes model). In microscopic analyses, RAGE vaccination suppressed glomerular hypertrophy and mesangial expansion in both diabetic models and significantly reduced glomerular basement membrane thickness in streptozotocin-induced diabetic mice. Results of an in vitro study indicated that the serum IgG antibody elicited by RAGE vaccination suppressed the expression of AGE-induced vascular cell adhesion molecule 1 and intracellular adhesion molecule 1 in endothelial cells. Thus, our newly developed RAGE vaccine attenuated the progression of DKD in mice and is a promising potential therapeutic strategy for patients with DKD.

MO052COL4A3/COL4A4 AS A CAUSE OF MULTICYSTIC KIDNEY DISEASE

Abstract Background and Aims Thin basement membrane nephropathy (TBMN), the most common cause of persistent microhaematuria (mH), is due to mutations in genes codifying alfa-3 and alfa-4 collagen IV chains (COL4A4/COL4A3). Initially considered as a benign condition, subsequent studies have shown that an important number of patients develop proteinuria and CKD. We reported in a previous small study the presence of multicystic kidney disease (MCD) in some TBMD patients. In this study we aimed to evaluate the presence of MCD in a larger cohort of TBMD patients and analyze its association with renal outcomes. Method We collected 50 patients with a diagnosis of TBMD based on the presence of persistent mH (&amp;gt;5 erythocytes per high power field in more than 90% of urinary sediments and radiological examinations to exclude other causes of mH) and at least one first-degree relative with persistent mH. TBMD diagnosis was confirmed by renal biopsy (glomerular basement membrane thickness less than 150nm) in 18 patients and by genetic test (pathogenic mutations in COL4A3/COL4A4) in 6 patients. MCD was diagnosed by the presence of uncountable cysts on renal ultrasonography. Results Mean age at diagnosis was 43.7 years, 34% were males and 18% had hypertension. At baseline, serum creatinine (SCr) was 0.9 mg/dL, proteinuria 0.48 gr/24h and 9 patients (18%) had CKD (estimated glomerular filtration rate -eGFR- lower than &amp;lt;60 mL/min/1.73m2). 7 patients (14%) had CKD G3 and 2 (4%) CKD G4. Kidney cysts were found in 34 patients (68%) and 19 (38%) met MCD criteria. After a mean follow-up of 14.7±11.5 years, 23 patients (46%) had CKD. Among them, 17 patients (34%) had CKD G3, 2 (4%) CKD G4, and 4 (8%) CKD G5. Hypertension was more frequent among CKD patients as compared with no-CKD patients (39 vs 0%, p 0.00), proteinuria was higher (0.58±0.68 vs 0.39±0.58 g/24h, p 0.05) and MCD more frequent (65.2% vs 14.8%, p 0.00). Patients with MCD had higher SCr (2.1 vs 1.1 mg/dL, p 0.004) and lower eGFR (41.7 vs 77.2 mL/min/1.73m2, p 0.00) at the end of follow-up, and MCD was the only risk factor for the occurrence of CKD (OR 6.49, 95% CI 1.3-31.6) by multivariable analysis that included age, hypertension and proteinuria. Conclusion MCD is frequently observed in TBMD patients and is a risk factor for the progression of CKD.

MO070NEW ASPECTS OF THE PATHOMORPHOLOGIC SEQUENCE OF NEPHRON LOSS IN DIABETIC NEPHROPATHY

Abstract Background and Aims Diabetic nephropathy (DN) is the leading cause of end-stage-renal disease in western countries. Despite of innumerable studies undertaken to elucidate the pathogenesis of DN the underlying morphologic alterations have been insufficiently analyzed. Method Re-evaluation of more than 800 biopsies was done showing several unknown features. Results: 1. Matrix accumulation in the mesangium: Thickening of the glomerular basement membrane (GBM) and expansion of the mesangial matrix are hallmarks of DN, generally considered to emerge from different sites of overproduction: GBM components from podocytes and mesangial matrix from mesangial cells. We show, that the accumulation of matrix in the mesangium emerges from an overproduction of GBM material by podocytes and endothelial cells and an impaired degradation by mesangial cells. The progressing deposition of worn-out GBM material into the mesangium accounts for the advancement from diffuse mesangial sclerosis (DMS) to nodular sclerosis (NS) and to the herniation of the tuft through the glomerular vascular pole to the outside; the latter is associated with the outgrowth of glomerular capillaries into the peri-glomerular space leading to the destruction of the juxtaglomerular apparatus. 2.The role of podocytes Podocytes have frequently been accused to play a central role in DN. This is correct, but in another way than generally assumed. Damage to podocytes cannot be seen in DMS. The albuminuria regularly seen during this stage derives, as previously suggested by others, from an increased leakiness of the glomerular capillary endothelium based on a deranged glycocalyx. Podocyte detachments start at the transition from DMS to NS, based on the loss of cross talk signals with the capillary endothelium: the increasing deposition of matrix leads to the collapse of many capillaries. These podocytes contribute little to the further progression of the damage: they are lost into primary urine or they undergo cell lysis.In addition to their role in increased matrix production, podocytes take an active role in the formation of tuft adhesions to Bowman’s capsule (BC), starting the progression to NS. Expansion of the matrix within the mesangium has led to expansion of the tuft (frequently associated with nodules) towards Bowman’s capsule (BC) or towards the urinary orifice. Podocytes on the surface of these expansions are in their majority structurally intact, exhibiting an intact pattern of foot processes. These podocytes come into contact with parietal epithelial cells and initiate DN-specific tuft adhesions to BC allowing the proliferation of glomerular capillaries into BC. There they deliver an exudate into BC that spreads around the entire circumference of the glomerulus presenting as giant insudative spaces. Moreover, this process encroaches via the glomerulo-tubular junction onto the tubule constituting the major pathway of glomerular damage extending to the tubulointerstitium. 3. Tubulointerstitial fibrosisIt is current opinion that the tubulointerstitial fibrosis may start from tubular damage resulting in an own, glomerular-independent pathway to nephron loss. However, there is scant evidence for such a mechanism. Studying 162 glomerulo-tubular transitions, we did not see a tubular epithelial or interstitial damage in those biopsies without any evidence of a glomerulo-tubular damage transfer. The only exception consists of the well-known prominent thickening of the tubular basement membrane, which may result in functional loss but does not lead to structural epithelial damage. Conclusion We consistently found that tubulo-interstitial damage develops after encroachment of the glomerular damage onto the tubule, leading first to a gradual degeneration of tubules which subsequently initiate the process of interstitial fibrosis.

Acute Kidney Injury in a Patient with Trisomy 21.

1. Hannah S. Kim, MD* 2. Francesca Costigliolo, MD† 3. Serena Bagnasco, MD† 4. Jeffrey Fadrowski, MD, MHS* 5. Rebecca L. Ruebner, MD, MSCE* 1. *Division of Pediatric Nephrology and 2. †Division of Pathology, Johns Hopkins University, Baltimore, MD A 5-year-old former 34-week girl with trisomy 21, hypothyroidism, and obstructive sleep apnea presents to the emergency department with a chief complaint of fever. She initially presented to her pediatrician 3 weeks earlier with sore throat and was diagnosed as having group A streptococcal pharyngitis. She completed 10 days of amoxicillin with resolution of sore throat. Now for the past 3 days she has had fever, chest pain, decreased enteral intake, and emesis. Review of systems is negative for diarrhea, gross hematuria, dysuria, polyuria, polydipsia, edema, weight loss, rash, arthralgia, arthritis, oral ulcers, epistaxis, and hemoptysis. She has had normal growth. She takes levothyroxine and took several doses of ibuprofen for fever over the past several weeks. There is no family history of kidney disease. She had no recent travel or sick contacts. On physical examination her weight is 45.2 lb (20.5 kg) (65th percentile); height, 46.1 in (117 cm) (62% percentile); temperature, 100.8°F (38.2°C); heart rate, 110 beats/min; respiratory rate, 24 breaths/min; oxygen saturation, 100% on room air; and blood pressure, 92/42 mm Hg (systolic 31st and diastolic 9th percentiles for age and sex). There is no pharyngeal injection or exudate. She appears well-hydrated, including moist mucous membranes. Lungs are clear to auscultation. Her abdomen is soft, nontender, and nondistended. There is no edema, and she has brisk capillary refill. She has no rash or joint swelling, erythema, or warmth. Laboratory evaluation includes a white blood cell count of 15,310/µL (15.31×109/L) with 83.6% neutrophils and 0.8% bands; hemoglobin …

Recovering histological sections for ultrastructural diagnosis of glomerular diseases through the pop-off technique.

Electron microscopy (EM) represents an indispensable technique for the diagnosis of kidney glomerular diseases. When dedicated tissue is not available, histological and cryostat sections can be reprocessed for EM using the pop-off technique. Here the practical value of this technique is analysed with emphasis on its accuracy in measuring basement membrane thickness and detecting immune deposits. Ninety-four histological sections of kidney tissues fixed in Serra's solution, stained with H&E, PAS, and Masson's Trichrome; for EM analysis, the sections were recovered from either treated or untreated microscope slides through the pop-off technique. Some sections were recovered from cryosections allocated for immunofluorescence. The ultrastructural details were sufficiently maintained on tissues fixed with Serra's solution despite being considered disadvantageous for EM. The type of microscope slides and the time of biopsy storage did not affect the quality of section recovery. The histological stains had only moderate effects on the electron-density of the glomerular basement membrane (GBM). The pop-off technique reduced the GBM thickness when compared to the conventional EM processing but preserved the electron density of immune deposits. The application of the pop-off method to renal biopsy is a useful recovery method that produces limited but satisfactory results when there is no suitable material for EM. The ultrastructural morphology was retained even from tissues fixed with Serra's solution, and deposits maintained the expected electron density, however, we observed an overall thickness reduction of the GBM that could have a potential impact on thin membrane disease diagnosis.

Betacyanins attenuates diabetic nephropathy in mice by inhibiting fibrosis and oxidative stress via the improvement of Nrf2 signaling

Abstract Diabetic nephropathy (DN) is a major cause of mortality in patients with diabetes, but there is a lack of effective therapeutic drugs for the disease treatment. Betacyanins (BCN) derived from plants exhibits anti-oxidative, anti-tumor and anti-inflammatory activities. However, its effects on DN have not been reported. In this study, we found that BCN supplementation markedly alleviated metabolic disorder and renal dysfunction in db/db mice. Additionally, BCN treatment obviously mitigated podocyte damage in diabetic mice, as evidenced by the decreased glomerular basement membrane thickness and the foot process width. Pathological analysis demonstrated that renal fibrosis was significantly inhibited by BCN through suppressing transforming growth factor (TGF)-β1/SMADs signaling in diabetic mice. Furthermore, diabetes-induced oxidative stress in kidney samples was evidently reduced in mice supplemented with BCN by improving nuclear factor erythroid-derived 2-related factor 2 (Nrf2) nuclear expression. The anti-fibrotic and anti-oxidant activities of BCN were confirmed in high glucose (HG)-stimulated podocytes in vitro. Importantly, we found that Nrf2 suppression could further exacerbate HG-induced collagen accumulation and reactive oxygen species (ROS) production in podocytes. Meanwhile, BCN-ameliorated fibrosis and oxidative stress were almost abrogated by the Nrf2 knockdown in HG-incubated podocytes. These findings elucidated that BCN could alleviate DN through reducing collagen and ROS deposition mainly through improving the Nrf2 signaling, subsequently alleviating renal dysfunction. Thus, BCN could be considered as a promising therapeutic agent for DN treatment.

ACTIVE IMMUNOTHERAPY AGAINST RECEPTOR FOR ADVANCED GLYCATION END PRODUCTS ATTENUATES THE PROGRESSION OF DIABETIC KIDNEY DISEASE

Objective: To combat global increases in the prevalence of diabetic kidney disease (DKD), we aimed to develop a new therapeutic vaccine targeting receptor for advanced glycation end products (RAGE). Design and method: RAGE vaccine was a water-in-oil emulsion composed of Freund's adjuvant and RAGE partial peptide that was conjugated to the carrier protein KLH. Male DBA2/J mice were subcutaneously immunized with three doses of KLH (vehicle) or RAGE vaccine and subsequently induced diabetes by multiple low-dose streptozotocin. To investigate the renal protective effect of RAGE vaccine, we measured urine albumin to creatinine ratio (UACR) and analyzed renal histology using light and electron microscopy. Results: There was no difference in blood glucose between vehicle-treated mice and vaccinated mice. On the other hand, immunization with the RAGE vaccine significantly attenuated the increase in UACR (vehicle group 850.6 ± 208.6 vs vaccine group 333.6 ± 104.7 ug/mg, n = 10/group, p &lt; 0.05). In the histological analyses, total glomerular volume of vaccinated mice was siginificantly less than that of vehicle-treated mice (vehicle group 5.28 ± 0.52 vs vaccine group 3.18 ± 0.17 um3, p &lt; 0.01) and glomerular mesangial matrix expansion was attenuated in vaccine-treated mice (vehicle group 23.6 ± 1.6 vs vaccine group 18.8 ± 0.8%, p &lt; 0.01). Ultrastructural morphometric study (n = 3/group) also indicated that glomerular basement membrane thickness was significantly suppressed in vaccinated mice compared with vehicle-treated mice (vehicle group 215.3 ± 1.9 vs vaccine group 181.3 ± 1.8 nm, p &lt; 0.01). Conclusions: These results suggest that active immunotherapy targeting RAGE is effective for the attenuation of DKD in streptozotocin-induced diabetic mice.