Rapid examination of lung tissues by nonlinear microscopy.

OBJECTIVES: Traditional histopathology is a time-intensive and labor-intensive process involving tissue formalin fixation, paraffin embedding, and microtoming into thin sections for H&E staining. Frozen section analysis is a modality used during surgery to quickly evaluate tissue, but it has limitations, such as the size and number of the specimens that can be analyzed as well as difficulties with fatty and bony tissues. Our objective was to investigate the performance of nonlinear microscopy, a fluorescence microscopy technique, for the rapid examination of resected lung tumors. METHODS: In this proof-of-principle study, nonlinear microscopy imaging of resected lung tissue was performed on a total of 73 tissue specimens collected from 13 patients who underwent lobectomy, segmentectomy, or wedge resection for pulmonary nodules. RESULTS: Two pathologists reviewed the digital nonlinear microscopy images in comparison to the corresponding histopathologic H&E slides from a variety of pulmonary pathologies. CONCLUSIONS: This study demonstrated that nonlinear microscopy readily replicates traditional H&E staining for both lung tumors and nonneoplastic pulmonary structures. Nonlinear microscopy provides many advantages over frozen section analysis and is an optical imaging platform that has the potential to augment rapid pathologic evaluation of resected tissues in the age of digital pathology.

Data retrieval from archival renal biopsies using nonlinear microscopy.

Thorough examination of renal biopsies may improve understanding of renal disease. Imaging of renal biopsies with fluorescence nonlinear microscopy (NLM) and optical clearing enables three-dimensional (3D) visualization of pathology without microtome sectioning. Archival renal paraffin blocks from 12 patients were deparaffinized and stained with Hoechst and Eosin for fluorescent nuclear and cytoplasmic/stromal contrast, then optically cleared using benzyl alcohol benzyl benzoate (BABB). NLM images of entire biopsy fragments (thickness range 88-660 µm) were acquired using NLM with fluorescent signals mapped to an H&E color scale. Cysts, glomeruli, exudative lesions, and Kimmelstiel-Wilson nodules were segmented in 3D and their volumes, diameters, and percent composition could be obtained. The glomerular count on 3D NLM volumes was high indicating that archival blocks could be a vast tissue resource to enable larger-scale retrospective studies. Rapid optical clearing and NLM imaging enables more thorough biopsy examination and is a promising technique for analysis of archival paraffin blocks.

Rapid Examination of Nonprocessed Renal Cell Carcinoma Using Nonlinear Microscopy.

CONTEXT.­: Histology, the traditional method of examining surgical tissue under a microscope, is a time-consuming process involving the fixation of tissue in formalin, dehydration, embedding in paraffin, and cutting into thin sections for hematoxylin-eosin (H&E) staining. Frozen section analysis is a faster alternative used in surgery to quickly evaluate tissue, but it has limitations, such as the size of the specimens that can be analyzed and difficulties with fatty and bony tissues. OBJECTIVE.­: To rapidly examine nonprocessed kidney tumors using nonlinear microscopy (NLM), a fluorescence microscopy technique that can rapidly visualize fresh or fixed, rapidly stained, nonprocessed tissue resembling H&E histology. This technology eliminates the need for fixation, embedding, microtome sectioning, or slide preparation. DESIGN.­: In this study, a total of 190 tissue specimens were collected from 46 patients who underwent partial or radical nephrectomy. RESULTS.­: Two genitourinary pathologists confirmed that diagnostically important features present in the H&E images could also be identified in the NLM images. CONCLUSIONS.­: The results of this study demonstrated that NLM had a high degree of correspondence with H&E staining for the classical variants of renal cell carcinoma. NLM offers several clinical benefits, such as facilitating rapid renal cell carcinoma diagnosis, assessment of targeted kidney biopsies for both tumor and medical kidney diseases, and collection of fresh renal cell carcinoma tissue for molecular studies.

Diffuse WT1 Positivity in Smooth Muscle Component Supports Dual Differentiation of Mesothelial Cells in the Histogenesis of Leiomyoadenomatoid Tumors-A Report of two Cases.

Leiomyoadenomatoid tumors of the epididymis are exceedingly rare biphasic tumors composed of an adenomatoid component in the form of gland-like structures lined by single flat or cuboidal cells admixed with smooth muscle. Radiological and gross findings cannot distinguish leiomyoadenomatoid tumors from the more common classic adenomatoid tumors or leiomyomas, and careful microscopic examination is critical in the identification of this esoteric variant. The histogenesis of this entity remains ambiguous. Common hypotheses include a collision tumor, a variant of an adenomatoid tumor with a smooth muscle component, or an adenomatoid tumor arising in the background of reactive smooth muscle hyperplasia. We present 2 cases of leiomyoadenomatoid tumors with diffuse nuclear WT1 positivity in both the adenomatoid and smooth muscle components, supporting the mesothelial origin of these tumors.

Yolk sac differentiation in urothelial carcinoma - A rare variant originating from aberrant differentiation of sarcomatoid components.

Yolk sac differentiation occurs in somatic neoplasms of the gastrointestinal and gynecologic tracts; it has rarely been reported in urothelial carcinoma. Here, we report three cases of yolk sac differentiation in sarcomatoid urothelial carcinoma. The epithelioid component of the sarcomatoid urothelial carcinoma showed divergent differentiation, including squamous, conventional glandular, small cell carcinoma, and yolk sac components. The sarcomatoid component showed malignant spindle cells admixed with focal chondroid and rhabdoid elements. In all three cases, the yolk sac areas were admixed with the sarcomatoid component and showed a glandular pattern, with vacuolated, eosinophilic cytoplasm. These areas were positive for SALL4, variably positive for glypican 3 and AFP, and negative for the conventional urothelial markers GATA3, p63, and 34ßE12. Yolk sac differentiation is an extremely rare occurrence in sarcomatoid urothelial carcinoma.

Rapid histological imaging of bone without microtome sectioning using nonlinear microscopy.

Tissue preparation for histologic evaluation of bone is particularly lengthy, limiting its use in intraoperative or intraprocedural histological evaluation. Nonlinear microscopy (NLM) is an optical sectioning microscopy method that can visualize pathology in freshly excised tissue without requiring physical microtome sectioning. This study describes a rapid protocol for NLM imaging of bone and associated cartilage. NLM imaging was performed on 71 specimens of normal bone as well as arthritic, malignant and inflammatory bone tissue from 40 patients who underwent joint replacement, amputation, bone marrow biopsy or autopsy. Specimens ranged in size from core needle biopsies to transections of entire femoral heads. Specimens were stained with acridine orange and sulforhodamine 101, nuclear and cytoplasmic/stromal fluorescent dyes, for 5 min, then rinsed for 30 s. NLM fluorescent images were displayed using colors analogous to hematoxylin and eosin (H&E) to facilitate interpretation. Pathologists examined NLM images of the specimens in real time by rapidly translating the specimen to areas of interest, similar to a standard transmission light microscope. By adjusting the NLM focus depth, images from a few-µm-thick layer could be obtained down to ~100 µm below the tissue surface, analogous to serial sectioning. Following real-time NLM imaging, the tissue was processed for conventional paraffin histology, and H&E slides were compared to recorded NLM images. Similarities and differences between NLM and paraffin H&E were assessed. NLM enabled visualization of normal bone architecture, including the lamellar matrix and osteocytes of trabecular bone, articular cartilage, as well as pathological bone features such osteoarthritis, osteomyelitis, and malignancy with an appearance resembling the paraffin H&E. Differences such as changes in cell border sharpness, cellular and nucleolar size, and color patterns were noted, suggesting that training is required for accurate evaluation of bone pathology with NLM. Irregular surface contours and debris generated by gross tissue preparation of bone can make some regions difficult to evaluate with NLM, but the ability to perform rapid three-dimensional translation and sub-surface imaging reduced these problems. NLM is a promising technique for rapid evaluation of bone pathology. Further studies assessing diagnostic performance are warranted.

Real-time diagnosis and Gleason grading of prostate core needle biopsies using nonlinear microscopy.

Rapid histologic assessment of fresh prostate biopsies may reduce patient anxiety, aid in biopsy sampling, and enable specimen triaging for molecular/genomic analyses and research that could benefit from fresh tissue analysis. Nonlinear microscopy (NLM) is a fluorescence microscopy technique that can produce high-resolution images of freshly excised tissue resembling formalin-fixed paraffin-embedded (FFPE) H&E. NLM enables evaluation of tissue up to ~100 µm below the surface, analogous to serial sectioning, but without requiring microtome sectioning. One hundred and seventy biopsies were collected from 63 patients who underwent in-bore MRI or MRI/ultrasound fusion biopsy procedures. Biopsies were stained in acridine orange and sulforhodamine 101, a nuclear and cytoplasmic/stromal fluorescent dye, for 45 s. Genitourinary pathologists evaluated the biopsies using NLM by translating the biopsies in real time to areas of interest and NLM images were recorded. After NLM evaluation, the biopsies were processed for standard FFPE H&E and similarities and differences between NLM and FFPE H&E were investigated. Accuracies of NLM diagnoses and Gleason scores were calculated using FFPE histology as the gold standard. Pathologists achieved a 92.4% sensitivity (85.0-96.9%, 95% confidence intervals) and 100.0% specificity (94.3-100.0%) for detecting carcinoma compared to FFPE histology. The agreement between the Grade Group determined by NLM versus FFPE histology had an unweighted Cohen's Kappa of 0.588. The average NLM evaluation time was 2.10 min per biopsy (3.08 min for the first 20 patients, decreasing to 1.54 min in subsequent patients). Further studies with larger patient populations, larger number of pathologists, and multiple institutions are warranted. NLM is a promising method for future rapid evaluation of prostate needle core biopsies.

Beyond the tubule: pathological variants of LRP2, encoding the megalin receptor, result in glomerular loss and early progressive chronic kidney disease.

Pathogenic variants in the LRP2 gene, encoding the multiligand receptor megalin, cause a rare autosomal recessive syndrome: Donnai-Barrow/Facio-Oculo-Acoustico-Renal (DB/FOAR) syndrome. Because of the rarity of the syndrome, the long-term consequences of the tubulopathy on human renal health have been difficult to ascertain, and the human clinical condition has hitherto been characterized as a benign tubular condition with asymptomatic low-molecular-weight proteinuria. We investigated renal function and morphology in a murine model of DB/FOAR syndrome and in patients with DB/FOAR. We analyzed glomerular filtration rate in mice by FITC-inulin clearance and clinically characterized six families, including nine patients with DB/FOAR and nine family members. Urine samples from patients were analyzed by Western blot analysis and biopsy materials were analyzed by histology. In the mouse model, we used histological methods to assess nephrogenesis and postnatal renal structure and contrast-enhanced magnetic resonance imaging to assess glomerular number. In megalin-deficient mice, we found a lower glomerular filtration rate and an increase in the abundance of injury markers, such as kidney injury molecule-1 and N-acetyl-ß-d-glucosaminidase. Renal injury was validated in patients, who presented with increased urinary kidney injury molecule-1, classical markers of chronic kidney disease, and glomerular proteinuria early in life. Megalin-deficient mice had normal nephrogenesis, but they had 19% fewer nephrons in early adulthood and an increased fraction of nephrons with disconnected glomerulotubular junction. In conclusion, megalin dysfunction, as present in DB/FOAR syndrome, confers an increased risk of progression into chronic kidney disease.

Nonlinear microscopy for detection of prostate cancer: analysis of sensitivity and specificity in radical prostatectomies.

Intraoperative evaluation of specimens during radical prostatectomy using frozen sections can be time and labor intensive. Nonlinear microscopy (NLM) is a fluorescence microscopy technique that can rapidly generate images that closely resemble H&E histology in freshly excised tissue, without requiring freezing or microtome sectioning. Specimens are stained with nuclear and cytoplasmic/stromal fluorophores, and NLM evaluation can begin within 3 min of grossing. Fluorescence signals can be displayed using an H&E color scale, facilitating pathologist interpretation. This study evaluates the accuracy of prostate cancer detection in a blinded reading of NLM images compared with the gold standard of formalin-fixed, paraffin-embedded H&E histology. A total of 122 freshly excised prostate specimens were obtained from 40 patients undergoing radical prostatectomy. The prostates were grossed, dissected into specimens of ~10 × 10 mm with 1-4 mm thickness, stained for 2 min for nuclear and cytoplasmic/stromal contrast, and then rinsed with saline for 30 s. NLM images were acquired and multiple images were stitched together to generate large field of view, centimeter-scale digital images suitable for reading. Specimens were then processed for standard paraffin H&E. The study protocol consisted of training, pretesting, and blinded reading phases. After a washout period, pathologists read corresponding paraffin H&E slides. Three pathologists achieved a 95% or greater sensitivity with 100% specificity for detecting cancer on NLM compared with paraffin H&E. Pooled sensitivity and specificity was 97.3% (93.7-99.1%; 95% confidence interval) and 100.0% (97.0-100.0%), respectively. Interobserver agreement for NLM reading had a Fleiss κ = 0.95. The high cancer detection accuracy and rapid specimen preparation suggest that NLM may be useful for intraoperative evaluation in radical prostatectomy.

Soluble fms-Like Tyrosine Kinase 1 Localization in Renal Biopsies of CKD.

INTRODUCTION: Soluble fms-like tyrosine kinase 1 (sFLT1) is a splice variant of the vascular endothelial growth factor (VEGF) receptor lacking the transmembrane and cytoplasmic domains and acts as a powerful antagonist of VEGF signaling. Plasma sFLT1 levels are higher in patients with chronic kidney disease (CKD) and correlate with renal dysfunction. The source of plasma sFLT1 in CKD is unclear. METHODS: Fifty-two renal biopsies were studied for sFLT1 expression using immunohistochemistry and evaluated on a 0-4 grading scale of positive cells within inflammatory infiltrates. These included drug-induced interstitial nephritis (6); allografts (12), with polyomavirus nephritis (3); diabetes mellitus (10); lupus glomerulonephritis (6); pauci-immune vasculitis (7); IgA nephropathy (6); and miscellaneous CKD (5). RESULTS: Forty-seven biopsies had inflammatory infiltrates of which 37 had sFLT1-positive cells: of these biopsies, 3 were grade 4, i.e., had cells that constituted more than 50% of the inflammatory infiltrate, 9 were grade 3 (25%-50%), 5 were grade 2 (10%-25%), 3 were grade 1 (10%), and 17 were grade 0.5 (<10%). There was a robust correlation (r2 = 0.89) between degree of inflammation and sFLT1-positive cells. CD68/sFLT1 co-immunostaining studies indicated that sFLT1-positive cells were histiocytes. The surrounding capillary network was reduced. CONCLUSION: sFLT1-positive histiocytes are generally part of the inflammatory infiltrates noted in CKD and are particularly abundant in forms of interstitial nephritis. Their presence promotes an anti-angiogenic state locally in the tubulointerstitium that could inhibit capillary repair, contribute to peritubular capillary loss, and enhance fibrosis in CKD.

Comparing histologic evaluation of prostate tissue using nonlinear microscopy and paraffin H&E: a pilot study.

Rapid histological assessment of large areas of prostate tissue is required for many intraoperative consultation scenarios such as margin evaluation. Nonlinear microscopy (NLM) enables imaging of large (whole mount) specimens without freezing or cryotoming. This study demonstrates rapid histological imaging of unsectioned prostate cancer surgical specimens using nonlinear microscopy and compares features of prostate pathology to standard paraffin embedded H&E histology. Fresh or formalin fixed specimens were stained in 2.5 min with fluorescent nuclear and stromal dyes. Nonlinear microscopy images of unsectioned tissues were generated by nonlinear (two-photon) excitation of the fluorophores, where fluorescence is only emitted from tissue at the microscope focus, avoiding the need for physical sectioning. The images were displayed in real time using a color scale similar to H&E, then tissues were processed for standard paraffin embedded H&E histology. Seventy nonlinear microscopy and corresponding paraffin H&E images of fresh and fixed prostate specimens (15 cancer, 55 benign) from 24 patients were read by genitourinary pathologists to assess if nonlinear microscopy could achieve an equivalent evaluation to paraffin embedded H&E histology. Differences between nonlinear microscopy images and paraffin H&E slides, including cytoplasmic color and stromal density, were observed, however nonlinear microscopy images could be interpreted with minimal training. Nonlinear microscopy enabled visualization of benign, atrophic and hyperplastic glands and stroma, ejaculatory ducts, vasculature and inflammatory changes. Nonlinear microscopy enabled identification of typical and variants of adenocarcinoma, as well as Gleason patterns. Perineural invasion and extraprostatic extension could also be assessed. Nonlinear microscopy images closely resemble paraffin H&E slides and enable rapid assessment of normal prostate architecture, benign conditions, and carcinoma in freshly excised and fixed specimens. Nonlinear microscopy can image large regions of tissue, equivalent to multiple frozen section tissue blocks, within minutes because cryotoming/microtoming are not required, making it a promising technique for intraoperative consultation.

The Convergence of Vasculopathy and Vasculitis: Computer Mapping Analysis of 2 Renal Biopsies in a Patient with both Systemic Sclerosis and ANCA-Related Vasculitis.

Scleroderma vasculopathy and ANCA (antineutrophil cytoplasmic antibodies)-associated glomerulonephritis have rarely been reported to occur simultaneously in one patient. Herein, we report a patient who presented with a classic constellation of clinical and laboratory findings of systemic scleroderma and was subsequently found to be positive for p-ANCA. Two renal biopsies, performed 5 months apart, demonstrated typical changes of the two entities in both acute and "healed" phases, which were analyzed by computer mapping techniques. The two renal biopsies were serially sectioned and stained routinely, and with CD31 and CD34 as endothelial markers. The slides were digitized, aligned and analyzed. Each glomerular tuft was sequentially studied in terms of total area (µm2) and each biopsy was individually profiled. All arterial vessels were sequentially studied with whole vessel and luminal areas delineated and ratios calculated. The initial biopsy contained 32 glomeruli almost all with extensive fibrinoid necrosis and destruction of the capillary network. The arterial vessels (interlobular and arcuate) showed intimal edema with luminal occlusion. CD31/CD34 stains showed variable endothelial intactness but demonstrated the luminal size shifts. The second biopsy had 37 glomeruli that were either segmentally or globally sclerotic with no active changes. The vessels were now normally patent. Each glomerular tuft and arterial vessel in both biopsies was analyzed as a serial section histogram documenting these changes. These studies depict the rare occurrence of two entities together, the scleroderma kidney vasculopathy and the glomerulonephritis of ANCA-associated vasculitis syndrome both in an acute and healing phase, profiled by computer mapping techniques.

Disruption of Renal Arginine Metabolism Promotes Kidney Injury in Hepatorenal Syndrome in Mice.

Tubular dysfunction is an important feature of renal injury in hepatorenal syndrome (HRS) in patients with end-stage liver disease. The pathogenesis of kidney injury in HRS is elusive, and there are no clinically relevant rodent models of HRS. We investigated the renal consequences of bile duct ligation (BDL)-induced hepatic and renal injury in mice in vivo by using biochemical assays, real-time polymerase chain reaction (PCR), Western blot, mass spectrometry, histology, and electron microscopy. BDL resulted in time-dependent hepatic injury and hyperammonemia which were paralleled by tubular dilation and tubulointerstitial nephritis with marked upregulation of lipocalin-2, kidney injury molecule 1 (KIM-1) and osteopontin. Renal injury was associated with dramatically impaired microvascular flow and decreased endothelial nitric oxide synthase (eNOS) activity. Gene expression analyses signified proximal tubular epithelial injury, tissue hypoxia, inflammation, and activation of the fibrotic gene program. Marked changes in renal arginine metabolism (upregulation of arginase-2 and downregulation of argininosuccinate synthase 1), resulted in decreased circulating arginine levels. Arginase-2 knockout mice were partially protected from BDL-induced renal injury and had less impairment in microvascular function. In human-cultured proximal tubular epithelial cells hyperammonemia per se induced upregulation of arginase-2 and markers of tubular cell injury. CONCLUSION: We propose that hyperammonemia may contribute to impaired renal arginine metabolism, leading to decreased eNOS activity, impaired microcirculation, tubular cell death, tubulointerstitial nephritis and fibrosis. Genetic deletion of arginase-2 partially restores microcirculation and thereby alleviates tubular injury. We also demonstrate that BDL in mice is an excellent, clinically relevant model to study the renal consequences of HRS. (Hepatology 2018; 00:000-000).

The Use of Cytochrome C Oxidase Enzyme Activity and Immunohistochemistry in Defining Mitochondrial Injury in Kidney Disease.

The renal biopsy is a dynamic way of looking at renal disease, and tubular elements are an important part of this analysis. The mitochondria in 20 renal biopsies were examined by immunohistochemical (electron transport chain enzyme: cytochrome C oxidase IV [COX IV]) and enzyme histochemical methods (COX), both by light and electron microscopy. The distal convoluted tubules and thick ascending limbs showed the greatest intensity in the COX immunostains and enzyme activity in controls. The degree of mitochondrial COX protein and enzyme activity diminished as the tubules became atrophic. With proximal hypertrophic changes, there was great variation in both COX activity and protein expression. In contrast, in three cases of systemic lupus erythematosus, biopsied for high-grade proteinuria, the activity was consistently upregulated, whereas protein expression remained normal. These unexpected findings of heterogeneous upregulation in hypertrophy and the dyssynchrony of protein expression and activity may indicate mitochondrial dysregulation. Functional electron microscopy showed COX activity delineated by the intense mitochondrial staining in normal or hypertrophic proximal tubules. With atrophic changes, residual small mitochondria with diminished activity could be seen. With mitochondrial size abnormalities (enlargement and irregularity, adefovir toxicity), activity persisted. In the renal biopsy, mitochondrial analysis is feasible utilizing immunohistochemical and enzyme histochemical techniques.

Gleason Misclassification Rate Is Independent of Number of Biopsy Cores in Systematic Biopsy.

OBJECTIVE: To compare the utility of saturation core biopsy and 12-core biopsy in detecting true Gleason grades, using final pathology in prostatectomy specimens as outcome measures, with a particular interest in Gleason upgrading. PATIENTS AND METHODS: We compared the concordance rates of Gleason grades diagnosed on biopsies and prostatectomy specimens in 375 consecutive patients, including 106 saturation biopsies (18-33 cores, median = 20 cores) and 269 12-core biopsies. Grading bias was addressed by a central rereview of all cases that had discordance in reporting high Gleason grades (Gleason grade ≥ 4) on biopsies and prostatectomy specimens. RESULTS: For patients with high Gleason grades on final pathology, saturation and 12-core biopsy schemes had a comparable sensitivity, specificity, negative and positive predictive values (72.5% vs 69.5%, 91.9% vs 97.6%, 64.2% vs 58.4%, and 94.3% vs 98.5%, respectively) in detecting high Gleason grades. On multivariate analysis, prebiopsy serum prostate-specific antigen and clinical T stage independently predicted Gleason upgrading; saturation biopsy was not a significant predictor. Approximately one-third of cases where high Gleason grade was not present in the biopsy were attributed to the confinement of high-grade tumors to unusual anatomic locations such as anterior lobes, apex, bladder neck, and parasagittal zones. CONCLUSION: Our study showed that Gleason misclassification rate is independent of the number of biopsy cores in systematic biopsy. One of the reasons for missing high Gleason grade tumors on systematic biopsy was unusual tumor location outside of the biopsy grid, supporting the need for improved detection technique such as magnetic resonance imaging-guided targeted biopsies.

Renal involvement in the immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) disorder.

BACKGROUND: Immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) disorder is an autoimmune disease caused by loss-of-function mutations in the gene encoding the forkhead box P3 (FOXP3) transcription factor. These mutations affect the normal function of circulating regulatory T cells. IPEX is characterized by profound immune dysregulation leading to dermatitis, enteropathy, multiple endocrinopathies and failure to thrive. Different forms of renal injury have also been noted in these patients but these have been described to a very limited extent. CASE-DIAGNOSIS: Three patients with IPEX with characteristic renal findings and mutations in FOXP3, including one novel mutation, are described. Case presentations are followed by a review of the renal manifestations noted in IPEX and the range of therapeutic options for this disorder. CONCLUSIONS: We recommend that IPEX be considered in the differential diagnosis of young children who present with signs of immune dysregulation with a concomitant renal biopsy demonstrating immune complex deposition in a membranous-like pattern and/or interstitial nephritis.

Endothelin-converting enzyme is a plausible target gene for hypoxia-inducible factor.

Renal endothelin-converting enzyme (ECE)-1 is induced in experimental diabetes and following radiocontrast administration, conditions characterized by renal hypoxia, hypoxia-inducible factor (HIF) stabilization, and enhanced endothelin synthesis. Here we tested whether ECE-1 might be a HIF-target gene in vitro and in vivo. ECE-1 transcription and expression increased in cultured vascular endothelial and proximal tubular cell lines, subject to hypoxia, to mimosine or cobalt chloride. These interventions are known to stabilize HIF signaling by inhibition of HIF-prolyl hydroxylases. In rats, HIF-prolyl-hydroxylase inhibition by mimosine or FG-4497 increased HIF-1α immunostaining in renal tubules, principally in distal nephron segments. This was associated with markedly enhanced ECE-1 protein expression, predominantly in the renal medulla. A progressive and dramatic increase in ECE-1 immunostaining over time, in parallel with enhanced HIF expression, was also noted in conditional von Hippel-Lindau knockout mice. Since HIF and STAT3 are cross-stimulated, we triggered HIF expression by STAT3 activation in mice, transfected by or injected with a chimeric IL-6/IL-6-receptor protein, and found a similar pattern of enhanced ECE-1 expression. Chromatin immunoprecipitation sequence (ChIP-seq) and PCR analysis in hypoxic endothelial cells identified HIF binding at the ECE-1 promoter and intron regions. Thus, our findings suggest that ECE-1 may be a novel HIF-target gene.

Granulomatosis with polyangiitis presenting as a renal tumor.

Inflammatory tumors of the kidney are uncommon and include primary inflammatory processes and systemic diseases such as sarcoidosis, IgG4 disease, and granulomatosis with polyangiitis (GPA). There are approximately 15 cases of the latter in the literature. Tumors in GPA are well described and have been reported in several organs including breast, orbit, mediastinum, central nervous system, and especially the lung. We report the case of a 48-year-old woman who presented with diffuse frontal headaches. Imaging showed both a cranial/sinus and renal mass. The pathology of the dura and nasal sinus biopsies were unrevealing. A nephrectomy was performed that demonstrated a discrete lesion with extensive necrosis, granulomatous inflammation, and crescentic pauci-immune glomerulonephritis, findings consistent with GPA.

Why is diabetes mellitus a risk factor for contrast-induced nephropathy?

Contrast-induced nephropathy (CIN) remains a leading cause of iatrogenic acute kidney injury, as the usage of contrast media for imaging and intravascular intervention keeps expanding. Diabetes is an important predisposing factor for CIN, particularly in patients with renal functional impairment. Renal hypoxia, combined with the generation of reactive oxygen species, plays a central role in the pathogenesis of CIN, and the diabetic kidney is particularly susceptible to intensified hypoxic and oxidative stress following the administration of contrast media. The pathophysiology of this vulnerability is complex and involves various mechanisms, including a priori enhanced tubular transport activity, oxygen consumption, and the generation of reactive oxygen species. The regulation of vascular tone and peritubular blood flow may also be altered, particularly due to defective nitrovasodilation, enhanced endothelin production, and a particular hyperresponsiveness to adenosine-related vasoconstriction. In addition, micro- and macrovascular diseases and chronic tubulointerstitial changes further compromise regional oxygen delivery, and renal antioxidant capacity might be hampered. A better understanding of these mechanisms and their control in the diabetic patient may initiate novel strategies in the prevention of contrast nephropathy in these susceptible patients.