Membrane Thickening Research Articles

Морфологічні особливості нирок плодів та новонароджених від матерів з підгострим інфекційно-запальним процесом у черевній порожнині, викликаним Escherichia coli (експериментальне дослідження)

Актуальність. В Україні з кожним роком відзначається збільшення кількості жінок, вагітність яких перебігає на тлі хронічних інфекційних захворювань. Escherichia coli є частим збудником бактеріальних інфекцій у жінок. Метою дослідження було виявлення морфологічних особливостей нирок плодів і новонароджених від матерів з експериментальним абдомінальним підгострим інфекційно-запальним процесом, викликаним Escherichia coli. Матеріали та методи. Авторами проведено експеримент на щурах лінії WAG, в ході якого сформовано дві групи: група I — 7 плодів та 11 новонароджених від 3 здорових самок; група II — 10 плодів і 13 новонароджених від 4 самок, яким моделювали абдомінальний інфекційно-запальний процес, викликаний Escherichia coli. Матеріалом дослідження послужили нирки плодів та новонароджених. Використовували гістологічні, гістохімічні, морфометричні та статистичні методи досліджень. Результати. Абдомінальний підгострий інфекційно-запальний процес в організмі матері, викликаний Escherichia coli, призводить до структурних змін у паренхіматозному та стромальному компонентах нирок, які наростали від плода до новонародженого. Гломерулярний апарат нирок характеризується нерівномірним розташуванням у кірковому шарі, затримкою розвитку, зміною форми, гемодинамічними змінами, розширенням сечового простору, відсутністю судинних клубочків, зменшенням кількості і компактності розташування капілярних петель в деяких молодих і зрілих ниркових тільцях; тубулярний апарат — затримкою розвитку, зміною форми та вогнищевим потовщенням базальних мембран окремих канальців, вогнищевими дистрофічними, некротичними та десквамативними змінами епітеліальної вистилки; стромальний компонент — склеротичними змінами, гемодинамічними порушеннями, більш вираженими в мозковому шарі, клітинною інфільтрацією, яка характеризується наявністю клітин фібробластичного ряду та імунних клітин. Висновки. Гістологічні та морфометричні зміни, що розвинулися в нирках плодів і новонароджених, зумовлені наявністю у матері експериментального підгострого інфекційно-запального процесу в черевній порожнині, викликаного Escherichia coli, призведуть в майбутньому до зниження функціональних можливостей нирок як гомеостатичного органу і розвитку різної нефрологічної патології у таких дітей.

Optimization of cone beam computed tomography for the assessment of alterations of the maxillary sinuses

Objective: To test the standard protocols of a CBCT unit in order to find lower-dose alternatives with diagnostically acceptable image quality for the maxillary sinuses visualization. Study design: An observational study was performed. Two dry skulls were used to simulate four conditions of the maxillary sinuses: normality, mucous retention pseudocyst, membrane thickening and bone graft. Cone beam computed tomography scans were obtained with an i-CAT classic unit using different acquisition protocols and a box of polystyrene to simulate soft tissue attenuation. All the protocols were established by the manufacturer, combining different energy parameters, fields of view and voxel sizes. Multiplanar reconstructions were presented to three Oral Radiologists through blind and randomized distribution. The specialists judged general image quality, sharpness, contrast, and the presence of noise and artifacts based on a 4-points scale. Results: Protocols with higher energy parameters had significant association with higher scores for general quality, sharpness and contrast (p<0.05). Protocols with intermediate level of radiation dose had also significant association with good and excellent image quality; for the presence of noise and artifacts the images were rated acceptable. Conclusion: i-CAT default protocols with lower dose of radiation were able to deliver acceptable image quality for the visualization of the maxillary sinuses.

Responses of Sinus Membrane and Antral Pseudocyst Following Lateral Window Sinus Augmentation with Bone Grafting: A Retrospective Study.

To investigate the influence of lateral window sinus augmentation on sinus physiology, including sinus membrane thickness and the outcome of antral pseudocysts. This retrospective study was performed by reviewing all lateral window sinus augmentation procedures, which were done between the years 2013 and 2015. Each enrolled patient had CBCT images preoperatively (T0), immediately postoperatively (T1), and 6 months postoperatively (T2). The sinus membrane thickness, pseudocyst dimensions, and intraoperative perforation were evaluated. Patient-related factors such as age and sex that could influence the mucosal properties were also analyzed. Based on established inclusion criteria, 306 patients with 320 sinuses were included in this study. The overall mean thickness of the sinus membrane (T0) was 1.30 ± 1.08 mm, and membrane thickening (> 2 mm) was observed in 22.19% of the sinuses. Preexisting pseudocysts were identified in 24 sinuses (7.50%), most of which remained unchanged or disappeared after 6 months. The intraoperative membrane perforation rate was lowest (1.96%) when the membrane thickness was 1.0 to 1.5 mm, and the perforation rate was increased in patients with thickened (> 2 mm) or thinned (≤ 1 mm) membrane. Membranes swelled immediately after operation (T1 vs T0, P < .01) and gradually recovered at 6 months. No significant change in membrane thickness was shown after the sinus augmentation procedure in the perforation group and pseudocyst group. The mean thickness of the sinus membrane at T0 and T1 was significantly higher for male subjects (P < .01). Lateral window sinus augmentation has little or no impact on sinus membrane thickness and antral pseudocysts after a 6-month healing period, except for a transient mild membrane swelling. Thickened and thinned membrane were risk factors for intraoperative perforation. Small-sized perforation and pseudocysts might not contraindicate sinus augmentation from the standpoint of the surgical impact on the sinus membrane.

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.

An Nphp1 knockout mouse model targeting exon 2-20 demonstrates characteristic phenotypes of human nephronophthisis.

Nephronophthisis (NPH) is the most prevalent monogenetic disorder leading to end-stage renal failure (ESRD) in childhood. Mutations in Nphp1, encoding a cilia-localized protein, account for the majority of NPH cases. Despite its identification many years ago, Nphp1 deletions targeting exon 4 or exon 20 have not reproduced the histological features of human NPH in murine models. In this study, we deleted exon 2-20 of Nphp1 by CRISPR/Cas9 gene editing to create a near-total knockout (KO) mouse model (Nphp1del2-20/del2-20). Nphp1del2-20/del2-20 mice faithfully reproduced the renal and extrarenal phenotypes associated with human NPH, including renal cyst development, tubular basement membrane thickening, retinal degeneration and abnormal spermatogenesis. Importantly, Nphp1 re-expression using an adenoviral-associated-virus-9 vector could partially rescue both renal and retinal phenotypes in Nphp1del2-20/del2-20 mice. Our results reported the first relevant Nphp1 mouse model with renal phenotypes for human disease. It will be a valuable model for future studies of Nphp1 function and to develop novel treatments for this common childhood disease.

Excitatory postsynaptic calcium transients at Aplysia sensory-motor neuron synapses allow for quantal examination of synaptic strength over multiple days in culture.

A more thorough description of the changes in synaptic strength underlying synaptic plasticity may be achieved with quantal resolution measurements at individual synaptic sites. Here, we demonstrate that by using a membrane targeted genetic calcium sensor, we can measure quantal synaptic events at the individual synaptic sites of Aplysia sensory neuron to motor neuron synaptic connections. These results show that synaptic strength is not evenly distributed between all contacts in these cultures, but dominated by multiquantal sites of synaptic contact, likely clusters of individual synaptic sites. Surprisingly, most synaptic contacts were not found opposite presynaptic varicosities, but instead at areas of pre- and postsynaptic contact with no visible thickening of membranes. The release probability, quantal size, and quantal content can be measured over days at individual synaptic contacts using this technique. Homosynaptic depression was accompanied by a reduction in release site probability, with no evidence of individual synaptic site silencing over the course of depression. This technique shows promise in being able to address outstanding questions in this system, including determining the synaptic changes that maintain long-term alterations in synaptic strength that underlie memory.

Long-term effects of sinus membrane perforation on dental implants placed with transcrestal sinus floor elevation: A case-control study.

There is a little comparative data on implants placed transcrestally with/without sinus membrane (SM) perforation. To compare the clinical and radiological outcomes of implants with maxillary sinus perforation and those without SM perforation. Among 560 transcrestally placed implants in 324 patients, the patients who underwent cone-beam computed tomographic radiography (CBCT) were included. The following groups were established: implants with SM perforation (group P) and implants without SM perforation based on postoperative panoramic radiographs and patient records (group NP). Group NP was further divided into subgroups based on CBCT taken at the last patient visit: group NP1 consisting of implants with no protrusion or <1 mm of protrusion and group NP2 consisting of implants with ≥1 mm of protrusion. Mixed linear regression was performed for the factors affecting SM thickening and marginal bone loss. Mixed survival analysis was also performed. A total of 379 implants in 221 patients were eligible. The mean follow-up period was 112.03 ± 54.2months. Twenty-six implants failed (2 and 24 implants in groups P and NP, respectively), mainly due to peri-implant bone loss. No statistically significant difference was noted between the groups in SM thickness (2.4 ± 2.8 mm, 2.1 ± 3.4 mm, and 2.5 ± 3.5 mm in groups P, NP1, and NP2, respectively, p > 0.05). Marginal bone loss in group NP1 was significantly greater than that in the other groups. In the mixed model, SM perforation was not a determinant of sinus membrane thickening and implant survival in the mixed models and the survival analysis, respectively. SM perforation in transcrestal sinus augmentation did not affect implant survival and SM thickening.

Short Term Pre-Operative Oral Corticosteroids-Tissue Remodeling in Chronic Rhinosinusitis with Nasal Polyps.

Chronic rhinosinusitis is a process involving a number of adverse changes in the mucosa of the paranasal sinuses and nasal polyps. The main histological features of tissue remodeling are changes in epithelial structure, oedema, degradation of ECM (extracellular matrix), angiogenesis, and subepithelial fibrosis. In this study, patients were divided into two groups: group 1—patients with CRSwNP (chronic rhinosinusitis with nasal polyps) taking a nasal steroid and an oral steroid in the preoperative period, and group 2—patients with CRSwNP taking only the nasal steroid in the preoperative period. All samples were subject to histopatologic evaluation. The aim of this study was to investigate the effect of oral corticosteroids and topical steroids on the tissue of paranasal sinuses. We have shown statistically significant decreases in tissue eosinophilia per 5HPF and decreased fibrosis in group 1. No significant differences were presented in the percentage of total tissue oedema, epithelium, neutrophils, basement membrane thickening and vessels. Using systemic administration of 40 mg of prednisone for seven days decreased the counts of eosinophils and decreased fibrosis in the nasal polyps tissue in CRSwNP.

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.

Testicular pathology in fatal COVID-19: A descriptive autopsy study.

BackgroundMulti‐organ damage is a common feature of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection, going beyond the initially observed severe pneumonia. Evidence that the testis is also compromised is growing.ObjectiveTo describe the pathological findings in testes from fatal cases of COVID‐19, including the detection of viral particles and antigens, and inflammatory cell subsets.Materials and methodsPostmortem testicular samples were obtained by percutaneous puncture from 11 deceased men and examined by reverse‐transcription polymerase chain reaction (RT‐PCR) for RNA detection and by light and electron microscopy (EM) for SARS‐CoV‐2. Immunohistochemistry (IHC) for the SARS‐CoV‐2 N‐protein and lymphocytic and histiocytic markers was also performed.ResultsEight patients had mild interstitial orchitis, composed mainly of CD68+ and TCD8+ cells. Fibrin thrombi were detected in five cases. All cases presented congestion, interstitial edema, thickening of the tubular basal membrane, decreased Leydig and Sertoli cells with reduced spermatogenesis, and strong expression of vascular cell adhesion molecule (VCAM) in vessels. IHC detected SARS‐Cov‐2 antigen in Leydig cells, Sertoli cells, spermatogonia, and fibroblasts in all cases. EM detected viral particles in the cytoplasm of fibroblasts, endothelium, Sertoli and Leydig cells, spermatids, and epithelial cells of the rete testis in four cases, while RT‐PCR detected SARS‐CoV‐2 RNA in three cases.Discussion and conclusionThe COVID‐19‐associated testicular lesion revealed a combination of orchitis, vascular changes, basal membrane thickening, Leydig and Sertoli cell scarcity, and reduced spermatogenesis associated with SARS‐CoV‐2 local infection that may impair hormonal function and fertility in men.

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.

Targeting the Pathobiology of Diabetic Kidney Disease

The pathobiology of diabetic kidney disease (DKD) involves an interplay between all the many different cell types that exist within the kidney and their shared and cumulative dysfunction in response to chronic hyperglycemia. DKD is characteriszed by morphological changes including tubular hypertrophy, podocyte dysfunction, mesangial expansion and mesangiolysis, endothelitis and capillary rarefaction, arteriolar hyalinosis, basement membrane thickening, and ultimately nephron dropout and tubulointerstitial fibrosis. These adaptive but ultimately maladaptive changes accelerate the progression of lesions in the diabetic kidney by increasing mechanical and oxidative stress, hypoxia, fibrogenesis, inflammation, senescence, and apoptosis. In particular, atrophy at the critical junction between Bowman's capsule and the proximal tubule likely represent the leading cause of nephron dropout and kidney function decline in DKD. Preventing, slowing, or reversing these changes should be the target of future "smart" therapies for patients with DKD, many of which are now under development.

Association between Odontogenic and Maxillary Sinus Conditions: A Retrospective Cone-Beam Computed Tomographic Study.

Odontogenic infections can directly trigger maxillary sinusitis. CBCT is an excellent choice for precise examination of maxillary sinuses and hard tissues within the oral cavity. The objective of this retrospective and the cross-sectional study was to analyze the influence of odontogenic conditions on the presence and intensity of maxillary sinus mucous membrane thickening using CBCT imaging. Moreover, periodontal bone loss and anatomic relationship between adjacent teeth and maxillary sinuses were assessed to evaluate its possible impact on creating maxillary thickening. The study sample consisted of 200 maxillary sinuses of 100 patients visible on CBCT examination with a field of view of 13 × 15 cm. The presented study revealed a significant influence of periapical lesions, inappropriate endodontic treatment, severe caries, and extracted teeth on the presence of increased thickening of maxillary sinus mucous membrane. In addition, an increase in the distance between root apices and maxillary sinus floor triggered a significant reduction of maxillary sinus mucous membrane thickening. The presence of periodontal bone loss significantly increases maxillary sinus mucous membrane thickening.

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.

Tight docking of membranes before fusion represents a metastable state with unique properties

Membrane fusion is fundamental to biological processes as diverse as membrane trafficking or viral infection. Proteins catalyzing membrane fusion need to overcome energy barriers to induce intermediate steps in which the integrity of bilayers is lost. Here, we investigate the structural features of tightly docked intermediates preceding hemifusion. Using lipid vesicles in which progression to hemifusion is arrested, we show that the metastable intermediate does not require but is enhanced by divalent cations and is characterized by the absence of proteins and local membrane thickening. Molecular dynamics simulations reveal that thickening is due to profound lipid rearrangements induced by dehydration of the membrane surface.

Histopathological Review on Discoid Lupus Erythematous Mimicking Granuloma Faciale

Discoid lupus erythematosus is the most common forms of chronic cutaneous lupus erythematosus. It is characterized by clinical manifestations of erythematous macules, papules, or plaques with a coin-like shape and the face is the most common predilection site. Clinical features often resemble granuloma faciale. This case report aimed to distinguish discoid lesions on the face based on the histopathological examination. A 71-year-old male with a few reddish lumps appeared on his face since three months ago. Physical examination showed multiple discrete erythematous plaques with overlying squamous. Hematoxylin and eosin staining on the epidermis demonstrated basket weave type orthokeratosis, basal vacuolar cell degeneration, epidermal atrophy with flat rete ridges, and follicular plugging while in the dermis obtained inflammatory cell infiltrates, especially in periadnexal areas. Histopathological features of DLE are hyperkeratosis, pilosebasea gland atrophy, follicular plugging, basal membrane thickening, and cellular infiltrate in periadnexa or perivascular areas more visible than in other types of CLE. In DLE, no subepidermal gren zone and eosinophil infiltrate were found, like histological features of granuloma faciale. Histopathological examination can be used to establish a diagnosis for discoid lesions on the face, although serology examination remains as the gold standart. Keywords: Discoid lupus erythematosus; Granuloma faciale; Histopathology

Anti-IgE therapy inhibits chemotaxis, proliferation and transformation of circulating fibrocytes in patients with severe allergic asthma.

Circulating fibrocytes act as precursors of myofibroblasts, contribute to airway remodelling in chronic asthma and migrate to injured tissues by expressing CXCR4 and CCR7. Anti-IgE therapy improves severe allergic asthma (SAA) control and airway remodelling in T2-high SAA. The effects of anti-IgE therapy on fibrocyte activities were investigated in this study. The expression of CCR7, CXCR4, ST2 and α-SMA (α-smooth muscle actin) in both circulating and cultured fibrocytes from all patients with asthma was measured, and was repeated after omalizumab treatment in SAA. Fibrocytes recruitment, proliferation and transformation were also measured in response to anti-IgE therapy. Omalizumab effectively improved asthma control and pulmonary function in T2-high SAA, associated with a decline in serum levels of IL-33 and IL-13. Omalizumab down-regulates CXCR4 and CCR7 expression of fibrocytes, which could suppress fibrocyte recruitment into the lungs. Omalizumab also suppressed the increased number of fibrocytes and α-SMA+ fibrocytes within the cultured non-adherent non-T (NANT) cells after 3-7 days of culture. The decrease in serum levels of IL-33 by omalizumab contributed to the effectiveness in inhibiting fibrocyte recruitment, proliferation and myofibroblast transformation through IL-33/ST2 axis. The elevated IL-13 expression in SAA patients potentiated the effects of IL-33 by increasing ST2 expression. Omalizumab reduced the number of circulating fibrocytes, cell and number of fibrocytes as well as α-SMA+ fibrocytes after 3-7 days of culture in SAA patients. IL-33 and IL-13 may be implicated in the effectiveness of omalizumab in inhibiting fibrocyte activation contributing partly to the clinical benefits in reducing lamina propria and basement membrane thickening.

182-OR: Genetic Susceptibility to Diabetic Kidney Disease Is Linked to C/EBP-XOR Axis

Diabetic kidney disease (DKD) is the leading single cause of end-stage renal disease in the United States. Approximately 30% of diabetics develop DKD with comparable blood glucose levels, indicating a significant genetic contribution for susceptibility. However, the underlying mechanisms that contribute to differential susceptibility are poorly understood. The glomerulus is the primary site of injury with hypertrophy and podocyte depletion being the hallmarks for progressive DKD. We have demonstrated that mitochondrial oxidative damage accumulation in glomerular endothelial cells, leads to podocyte loss via endothelial-to-podocyte crosstalk in DKD susceptible DBA/2J (D2) mice compared to DKD resistant C57BL/6J (B6) mice. To map genetic loci associated with podocyte depletion after long-term diabetes (6mth), we used the 39 strains of BXD, and parental B6 and D2 mice. We identified a significant cis-acting variant in the promoter of xanthine oxidoreductase (XOR). XORs catalyze the oxidation of purine substrates produce uric acid and reactive oxygen species. XOR expression in the kidney and circulating activity were significantly higher in diabetic D2 compared to B6. XOR inhibition significantly reduced albuminuria, oxidative damage in glomeruli and prevented podocyte depletion in diabetic D2 mice. We used CRISPR/Cas9 to knock-in the XOR risk variant into DKD-resistant B6 mice and generated mutant B6-XORem1 mice with significantly higher XOR activity than B6. This risk variant is a transcription factor binding site to C/EBPβ, we show higher C/EBPβ expression in glomerular endothelium from diabetic B6-XORem1 mice but not in diabetic B6. B6-XORem1 mice had increased mitochondrial oxidative stress in endothelial cells, podocyte depletion, basement membrane thickening, albuminuria, glomerulosclerosis and tubular injury, and these changes were prevented with XOR inhibition. Our data suggest that the identified promoter variant is causal for DKD susceptibility via the C/EBPβ-XOR axis. Disclosure Q. Wang: None. H. Qi: None. S. Li: None. G. Casalena: None. L. Yu: None. I. S. Daehn: None. Funding National Institutes of Health

Receptor activator of NF-κB mediates podocyte injury in diabetic nephropathy

Receptor activator of NF-κB (RANK) expression is increased in podocytes of patients with diabetic nephropathy. However, the relevance of RANK to diabetic nephropathy pathobiology remains unclear. Here, to evaluate the role of podocyte RANK in the development of diabetic nephropathy, we generated a mouse model of podocyte-specific RANK depletion (RANK-/-Cre T), and a model of podocyte-specific RANK overexpression (RANK TG), and induced diabetes in these mice with streptozotocin. We found that podocyte RANK depletion alleviated albuminuria, mesangial matrix expansion, and basement membrane thickening, while RANK overexpression aggravated these indices in streptozotocin-treated mice. Moreover, streptozotocin-triggered oxidative stress was increased in RANK overexpression but decreased in the RANK depleted mice. Particularly, the expression of NADPH oxidase 4, and its obligate partner, P22phox, were enhanced in RANK overexpression, but reduced in RANK depleted mice. In parallel, the transcription factor p65 was increased in the podocyte nuclei of RANK overexpressing mice but decreased in the RANK depleted mice. The relevant findings were largely replicated with high glucose-treated podocytes invitro. Mechanistically, p65 could bind to the promoter regions of NADPH oxidase 4 and P22phox, and increased their respective gene promoter activity in podocytes, dependent on the levels of RANK. Taken together, these findings suggested that high glucose induced RANK in podocytes and caused the increase of NADPH oxidase 4 and P22phox via p65, possibly together with the cytokines TNF- α, MAC-2 and IL-1 β, resulting in podocyte injury. Thus, we found that podocyte RANK was induced in the diabetic milieu and RANK mediated the development of diabetic nephropathy, likely by promoting glomerular oxidative stress and proinflammatory cytokine production.

Pathophysiological mechanisms of diabetic retinopathy

Aim. To evaluate pathophysiological mechanisms in diabetic retinopathy.Material and methods. Factors of diabetic retinopathy pathogenesis, scientific reviews.Results. The evaluation of pathophysiological mechanisms in diabetic retinopathy found that early stages are characterized by histopathological changes, which include loss of pericytes, basement membrane thickening, haemodynamic alterations leading to reduced vascular integrity. The later stages of diabetic retinopathy are characterized by complications, which include visual impairment, primarily due to macular edema and proliferative diabetic retinopathy. Also, the severity of retinopathy was associated with poorer metabolic control, demonstrated by elevated HbA1c. Diabetic complications accompany the accumulation of advanced glycation end products in diabetic tissues. Increased accumulation of these products has been reported in epiretinal membranes by the use of the immunohistochemical technique. The binding of advanced glycation end products to a high-affinity receptor in pericytes exerts selective toxicity resulting in their death. Vascular endothelial growth factor exerts an important role in intraocular neovascularization due to ischemic retinopathy.Conclusions. Early stages of diabetic retinopathy are characterized by his topathological changes which include loss of pericytes, basement membrane thickening, haemodynamic alterations leading to reduced vascular integrity. The later stages of diabetic retinopathy are characterized by complications, which include visual impairment, primarily due to macular edema and proliferative diabetic retinopathy. The binding of advanced glycation end products to a high-affinity receptor in pericytes exerts selective toxicity resulting in their death.