Automatic diagnosis and classification of breast surgical samples with dynamic full-field OCT and machine learning.

Purpose: The adoption of emerging imaging technologies in the medical community is often hampered when they provide a new unfamiliar contrast that requires experience to be interpreted. Dynamic full-field optical coherence tomography (D-FF-OCT) microscopy is such an emerging technique. It provides fast, high-resolution images of excised tissues with a contrast comparable to H&E histology but without any tissue preparation and alteration. Approach: We designed and compared two machine learning approaches to support interpretation of D-FF-OCT images of breast surgical specimens and thus provide tools to facilitate medical adoption. We conducted a pilot study on 51 breast lumpectomy and mastectomy surgical specimens and more than 1000 individual 1.3×1.3 mm2 images and compared with standard H&E histology diagnosis. Results: Using our automatic diagnosis algorithms, we obtained an accuracy above 88% at the image level (1.3×1.3 mm2) and above 96% at the specimen level (above cm2). Conclusions: Altogether, these results demonstrate the high potential of D-FF-OCT coupled to machine learning to provide a rapid, automatic, and accurate histopathology diagnosis with minimal sample alteration.

Missing Self-Induced Activation of NK Cells Combines with Non-Complement-Fixing Donor-Specific Antibodies to Accelerate Kidney Transplant Loss in Chronic Antibody-Mediated Rejection.

BACKGROUND: Binding of donor-specific antibodies (DSAs) to kidney allograft endothelial cells that does not activate the classic complement cascade can trigger the recruitment of innate immune effectors, including NK cells. Activated NK cells contribute to microvascular inflammation leading to chronic antibody-mediated rejection (AMR). Recipient NK cells can also trigger antibody-independent microvascular inflammation by sensing the absence of self HLA class I molecules ("missing self") on allograft endothelial cells. This translational study investigated whether the condition of missing self amplifies DSA-dependent NK cell activation to worsen chronic AMR. METHODS AND RESULTS: Among 1682 kidney transplant recipients who underwent an allograft biopsy at Lyon University Hospital between 2004 and 2017, 135 fulfilled the diagnostic criteria for AMR and were enrolled in the study. Patients with complement-fixing DSAs identified by a positive C3d binding assay (n=73, 54%) had a higher risk of transplant failure (P=0.002). Among the remaining patients with complement-independent chronic AMR (n=62, 46%), those in whom missing self was identified through donor and recipient genotyping exhibited worse allograft survival (P=0.02). In multivariable analysis, only proteinuria (HR: 7.24; P=0.01) and the presence of missing self (HR: 3.57; P=0.04) were independent predictors for transplant failure following diagnosis of chronic AMR. Cocultures of human NK cells and endothelial cells confirmed that addition of missing self to DSA-induced NK cell activation increased endothelial damage. CONCLUSIONS: The assessment of missing self at the time of diagnosis of chronic AMR identifies patients at higher risk for kidney transplant failure.

Transcriptional Changes in Kidney Allografts with Histology of Antibody-Mediated Rejection without Anti-HLA Donor-Specific Antibodies.

BACKGROUND: Circulating donor-specific anti-HLA antibodies (HLA-DSAs) are often absent in serum of kidney allograft recipients whose biopsy specimens demonstrate histology of antibody-mediated rejection (ABMR). It is unclear whether cases involving ABMR histology without detectable HLA-DSAs represent a distinct clinical and molecular phenotype. METHODS: In this multicenter cohort study, we integrated allograft microarray analysis with extensive clinical and histologic phenotyping from 224 kidney transplant recipients between 2011 and 2017. We used the term ABMR histology for biopsy specimens that fulfill the first two Banff 2017 criteria for ABMR, irrespective of HLA-DSA status. RESULTS: Of 224 biopsy specimens, 56 had ABMR histology; 26 of these (46.4%) lacked detectable serum HLA-DSAs. Biopsy specimens with ABMR histology showed overexpression of transcripts mostly related to IFNγ-induced pathways and activation of natural killer cells and endothelial cells. HLA-DSA-positive and HLA-DSA-negative biopsy specimens with ABMR histology displayed similar upregulation of pathways and enrichment of infiltrating leukocytes. Transcriptional heterogeneity observed in biopsy specimens with ABMR histology was not associated with HLA-DSA status but was caused by concomitant T cell-mediated rejection. Compared with cases lacking ABMR histology, those with ABMR histology and HLA-DSA had higher allograft failure risk (hazard ratio [HR], 7.24; 95% confidence interval [95% CI], 3.04 to 17.20) than cases without HLA-DSA (HR, 2.33; 95% CI, 0.85 to 6.33), despite the absence of transcriptional differences. CONCLUSIONS: ABMR histology corresponds to a robust intragraft transcriptional signature, irrespective of HLA-DSA status. Outcome after ABMR histology is not solely determined by the histomolecular presentation but is predicted by the underlying etiologic factor. It is important to consider this heterogeneity in further research and in treatment decisions for patients with ABMR histology.

Missing self triggers NK cell-mediated chronic vascular rejection of solid organ transplants.

Current doctrine is that microvascular inflammation (MVI) triggered by a transplant -recipient antibody response against alloantigens (antibody-mediated rejection) is the main cause of graft failure. Here, we show that histological lesions are not mediated by antibodies in approximately half the participants in a cohort of 129 renal recipients with MVI on graft biopsy. Genetic analysis of these patients shows a higher prevalence of mismatches between donor HLA I and recipient inhibitory killer cell immunoglobulin-like receptors (KIRs). Human in vitro models and transplantation of ß2-microglobulin-deficient hearts into wild-type mice demonstrates that the inability of graft endothelial cells to provide HLA I-mediated inhibitory signals to recipient circulating NK cells triggers their activation, which in turn promotes endothelial damage. Missing self-induced NK cell activation is mTORC1-dependent and the mTOR inhibitor rapamycin can prevent the development of this type of chronic vascular rejection.

Uncontrolled donation after circulatory death: comparison of two kidney preservation protocols on graft outcomes.

BACKGROUND: Kidney transplantation following uncontrolled donation after circulatory death (uDCD) presents a high risk of delayed graft function due to prolonged warm ischemia time. In order to minimise the effects of ischemia/reperfusion injury during warm ischemia, normothermic recirculation recently replaced in situ perfusion prior to implantation in several institutions. The aim of this study was to compare these preservation methods on kidney graft outcomes. METHODS: The primary endpoint was the one-year measured graft filtration rate (mGFR). We collected retrospective data from 64 consecutive uDCD recipients transplanted over a seven-year period in a single centre. RESULTS: Thirty-two grafts were preserved by in situ perfusion and 32 by normothermic recirculation. The mean ± SD mGFR at 1 year post-transplantation was 43.0 ± 12.8 mL/min/1.73 m2 in the in situ perfusion group and 53.2 ± 12.8 mL/min/1.73 m2 in the normothermic recirculation group (p = 0.01). Estimated GFR levels were significantly higher in the normothermic recirculation group at 12 months (p = 0.01) and 24 months (p = 0.03) of follow-up. We did not find any difference between groups regarding patient and graft survival, delayed graft function, graft rejection, or interstitial fibrosis. CONCLUSIONS: Function of grafts preserved by normothermic recirculation was better at 1 year and the results suggest that this persists at 2 years, although no difference was found in short-term outcomes. Despite the retrospective design, this study provides an additional argument in favour of normothermic recirculation.

Hypothermic pulsatile preservation of kidneys from uncontrolled deceased donors after cardiac arrest - a retrospective study.

Kidneys from uncontrolled donors after cardiac arrest (uDCD) suffer from a period of warm ischemia between cardiac arrest and cold flushing. Aim of the study was to evaluate renal outcomes of uDCD kidneys selected on the basis of renal Resistance Index (RI) and its influence on graft function and survival. The study included 44 kidneys procured from 26 uDCD starting 1.1.2006 until 12.31.2013. The donors (Maastricht category II) underwent cardiopulmonary resuscitation by assisted ventilation and chest compression; the organs were preserved with in situ cold perfusion or a normothermic regional perfusion. All kidneys were perfused on hypothermic (1-4 °C) pulsatile perfusion machine (RM3; Waters Medical System) and discarded when RI ≥0.5 mmHg/ml/min after 6 h of perfusion. There was one (2.2%) primary non function, while 37 recipients (84.1%) experienced delayed graft function. Graft survival was 97.6% at 1 and 3 post-transplantation years. Linear regression models showed that lower values of RI at the end of perfusion were associated with higher values of Modification of Diet in Renal Disease at 3 (P = 0.049) and 6 months after transplantation (P = 0.010) and with higher values of inulin clearance at 1 year (P = 0.030). RI showed to be a useful tool to select uDCD kidneys allowing to achieve good clinical results.

Computer-assisted topological analysis of renal allograft inflammation adds to risk evaluation at diagnosis of humoral rejection.

Antibody-mediated rejection is associated with heterogeneous kidney allograft outcomes. Accurate evaluation of risk for graft loss at time of diagnosis is necessary to offer personalized treatment. In contrast with serological and molecular assessment, morpho-histological evaluation of antibody-mediated rejection lesions has not significantly evolved. This relies on Banff classifications designed to be of diagnostic discriminatory power rather than prognostic and face quantitative and qualitative limitations. Here we developed a method of Computer-assisted Analysis of Graft Inflammation (CAGI) to improve the classification of allograft inflammation. Digitization of immunostained biopsy sections, image processing and algorithm-driven analysis allowed quantification of macrophages, T cells, B cells, and granulocytes per unit surface of interstitium, capillaries or glomeruli. CAGI was performed on biopsy specimens of 52 patients with extensively phenotyped antibody-mediated rejection. Macrophage numbers in capillaries and interstitium, but not Banff scores or the amount of other immune cell subsets, correlated with donor-specific antibody (DSA) mean fluorescence intensity and DSA-C3d status. The quantity of macrophages in the interstitium and DSA-C3d status were the only independent predictors for significant allograft loss at the time of antibody-mediated rejection diagnosis (hazard ratio 3.71 and 2.34, respectively). A significant strategy integrating the DSA-C3d assay and the quantification of interstitial macrophages allowed identification of three groups with distinct renal prognosis: DSA-C3d-, DSA-C3d+/macrophages-low and DSAC3d+/macrophages-high. Thus, CAGI brings a missing piece to the antibody-mediated rejection puzzle by identifying morpho-histological processes that bridge in vitro parameters of DSA pathogenicity and graft loss. Hence, this approach could be useful in future integrated strategies of risk evaluation.

Essential role of immobilized chemokine CXCL12 in the regulation of the humoral immune response.

Chemokines control the migration of a large array of cells by binding to specific receptors on cell surfaces. The biological function of chemokines also depends on interactions between nonreceptor binding domains and proteoglycans, which mediate chemokine immobilization on cellular or extracellular surfaces and formation of fixed gradients. Chemokine gradients regulate synchronous cell motility and integrin-dependent cell adhesion. Of the various chemokines, CXCL12 has a unique structure because its receptor-binding domain is distinct and does not overlap with the immobilization domains. Although CXCL12 is known to be essential for the germinal center (GC) response, the role of its immobilization in biological functions has never been addressed. In this work, we investigated the unexplored paradigm of CXCL12 immobilization during the germinal center reaction, a fundamental process where cellular traffic is crucial for the quality of humoral immune responses. We show that the structure of murine germinal centers and the localization of GC B cells are impaired when CXCL12 is unable to bind to cellular or extracellular surfaces. In such mice, B cells carry fewer somatic mutations in Ig genes and are impaired in affinity maturation. Therefore, immobilization of CXCL12 is necessary for proper trafficking of B cells during GC reaction and for optimal humoral immune responses.

A multicentre study of 95 biopsy-proven cases of renal disease in primary Sjögren's syndrome.

OBJECTIVE.: Renal involvement is a rare event during primary SS (pSS). We aimed to describe the clinico-biological and histopathological characteristics of pSS-related nephropathy and its response to treatment. METHODS.: We conducted a French nationwide, retrospective, multicentre study including pSS patients fulfilling American-European Consensus Group criteria or enlarged American-European Consensus Group criteria, and with biopsy-proven renal involvement. RESULTS.: A total of 95 patients were included (median age 49 years). An estimated glomerular filtration rate (eGFR) of <60 ml/min was found in 82/95 patients (86.3%). Renal biopsy demonstrated tubulointerstitial nephritis (TIN) in 93 patients (97.9%), and frequent (75%) plasma cell infiltrates. Glomerular lesions were found in 22 patients (23.2%), mainly related to cryoglobulin. The presence of anti-SSA (76.8%) and anti-SSB (53.8%) antibodies was particularly frequent among patients with TIN and was associated with a worse renal prognosis. Eighty-one patients (85.3%) were treated, with CSs in 80 (98.8%) and immunosuppressive agents (mostly rituximab) in 21 cases (25.9%). Despite marked interstitial fibrosis at initial biopsy, kidney function improved significantly during the 12-month period following diagnosis (final eGFR 49.9 vs 39.8 ml/min/1.73 m 2 at baseline, P < 0.001). No proven benefit of immunosuppressive agents over steroid therapy alone was found in this study. CONCLUSION.: Renal involvement of pSS is mostly due to TIN with marked T, B and especially plasma cell infiltration. Renal dysfunction is usually isolated but can be severe. Use of CSs can improve the eGFR, but further studies are needed to define the best therapeutic strategy in this disease.

The intracellular bacteria Chlamydia hijack peroxisomes and utilize their enzymatic capacity to produce bacteria-specific phospholipids.

Chlamydia trachomatis is an obligate intracellular pathogen responsible for loss of eyesight through trachoma and for millions of cases annually of sexually transmitted diseases. The bacteria develop within a membrane-bounded inclusion. They lack enzymes for several biosynthetic pathways, including those to make some phospholipids, and exploit their host to compensate. Three-dimensional fluorescence microscopy demonstrates that small organelles of the host, peroxisomes, are translocated into the Chlamydia inclusion and are found adjacent to the bacteria. In cells deficient for peroxisome biogenesis the bacteria are able to multiply and give rise to infectious progeny, demonstrating that peroxisomes are not essential for bacterial development in vitro. Mass spectrometry-based lipidomics reveal the presence in C. trachomatis of plasmalogens, ether phospholipids whose synthesis begins in peroxisomes and have never been described in aerobic bacteria before. Some of the bacterial plasmalogens are novel structures containing bacteria-specific odd-chain fatty acids; they are not made in uninfected cells nor in peroxisome-deficient cells. Their biosynthesis is thus accomplished by the metabolic collaboration of peroxisomes and bacteria.

Urinary retinol binding protein is a marker of the extent of interstitial kidney fibrosis.

Currently, a non-invasive method to estimate the degree of interstitial fibrosis (IF) in chronic kidney disease is not available in routine. The aim of our study was to evaluate the diagnostic performance of the measurement of urinary low molecular weight (LMW) protein concentrations as a method to determine the extent of IF. The urines specimen from 162 consecutive patients who underwent renal biopsy were used in the analysis. Numerical quantification software based on the colorimetric analysis of fibrous areas was used to assess the percentage IF. Total proteinuria, albuminuria, and the urinary levels of retinol binding protein (RBP), alpha1-microglobulin (α1MG), beta 2-microglobulin (ß2MG), transferrin, and IgG immunoglobulins were measured. There was a significant correlation between the degree of IF and the RBP/creatinine (creat) ratio (R2: 0.11, p<0.0001). IF was associated to a lesser extent with urinary ß2MG and α1MG; however, there was no association with total proteinuria or high molecular weight (HMW) proteinuria. The correlation between IF and RBP/creat remained significant after adjustment to the estimated glomerular filtration rate, age, body mass index, α1MG, and ß2MG. The specificity of the test for diagnosing a fibrosis score of >25% of the parenchyma was 95% when using a threshold of 20 mg/g creat. In conclusion, RBP appears to be a quantitative and non-invasive marker for the independent prediction of the extent of kidney IF. Because methods for the measurement of urinary RBP are available in most clinical chemistry departments, RBP measurement is appealing for implementation in the routine care of patients with chronic kidney disease.

Progression of pulse pressure in kidney recipients durably exposed to CsA is a risk factor for epithelial phenotypic changes: an ancillary study of the CONCEPT trial.

In this ancillary study of the CONCEPT trial, we studied the role of CsA withdrawal at 3 months (3M) post-transplant on the intensity of epithelial phenotypic changes (EPC, an early marker for kidney fibrogenesis) on the 12 M surveillance biopsy. Although conversion from CsA to sirolimus (SRL) at 3M was reported to have improved mean graft function at 12 M, it did not reduce the score of EPC (1.73 ± 1.15 in the SRL group vs. 1.87 ± 1 in the CsA group, P = 0.61). Acute rejection, which had occurred twice more frequently in SRL-converted patients included here, was associated with 12 M EPC. Interestingly, we observed that the patients durably exposed to CsA and who developed 12 M EPC had a significant progression of blood pulse pressure (pp) from 1 to 6M post-transplantation (Δpp = +12.3 mmHg, P = 0.0035). Pulse pressure at 4, 6, and 9 M and pp progression from 1 to 6M were significantly associated with the development of EPC at 12 M in renal grafts. Logistic regression analysis revealed that a high 6M pp (≥ 60 mmHg) was an independent risk factor for 12 M EPC with an odds ratio of 2.25 per additional 10 mmHg pp (95%CI: 1.14-4.4, P = 0.02) after adjustment with recipient's and donor's age, acute rejection incidence and immunosuppressive regimen. A post hoc analysis of the data collected in the whole population CONCEPT study revealed that pp was significantly higher at 6 months in patients maintained on CsA and that at this time point pp correlated negatively with GFR at 1 year.

Dual kidney transplantation from uncontrolled deceased donors after cardiac arrest: a possible option.

Organ shortage is a major problem in organ transplantation. For this reason, transplantation teams have found it necessary to revisit their organ acceptance criteria. Uncontrolled deceased donors after cardiac arrest could increase the donor pool by 20%, but at the same time there is a greater risk of delayed graft function and primary non-function. Dual kidney transplantation is an option when single kidney transplantation cannot be carried out because of lack of organ quality. We report for the first time our four first dual kidney transplantation from uncontrolled deceased donors after cardiac arrest with a follow up longer than 1 year. We described graft outcomes until 5 years, and histology at 3 and 12 months after transplantation. All organs were machine perfused in order to assess their quality leading to a single kidney transplantation or dual kidney transplantation decision. After 1 year of follow up, all grafts were functional with a mean estimated glomerular filtration rate of 44.5 ± 3.3 mL/min/1.73 m(2), and a mean inulin clearance of 43.7 ± 13.6 mL/mn/1.73 m(2). These findings suggest that dual kidney transplantation can represent a viable option for kidneys unsuitable for single kidney transplantation without increasing the rate of surgical complications. Successful transplantation is linked to histological, biological and donor clinical criteria, as well as perfusion parameters.

αTAT1 catalyses microtubule acetylation at clathrin-coated pits.

In most eukaryotic cells microtubules undergo post-translational modifications such as acetylation of α-tubulin on lysine 40, a widespread modification restricted to a subset of microtubules that turns over slowly. This subset of stable microtubules accumulates in cell protrusions and regulates cell polarization, migration and invasion. However, mechanisms restricting acetylation to these microtubules are unknown. Here we report that clathrin-coated pits (CCPs) control microtubule acetylation through a direct interaction of the α-tubulin acetyltransferase αTAT1 (refs 8, 9) with the clathrin adaptor AP2. We observe that about one-third of growing microtubule ends contact and pause at CCPs and that loss of CCPs decreases lysine 40 acetylation levels. We show that αTAT1 localizes to CCPs through a direct interaction with AP2 that is required for microtubule acetylation. In migrating cells, the polarized orientation of acetylated microtubules correlates with CCP accumulation at the leading edge, and interaction of αTAT1 with AP2 is required for directional migration. We conclude that microtubules contacting CCPs become acetylated by αTAT1. In migrating cells, this mechanism ensures the acetylation of microtubules oriented towards the leading edge, thus promoting directional cell locomotion and chemotaxis.

Α-arrestin 1 (ARRDC1) and ß-arrestins cooperate to mediate Notch degradation in mammals.

Notch signaling is a conserved signaling pathway implicated in embryogenesis and adult tissue maintenance. Notch signaling strength is strictly regulated, notably by maintaining a controlled pool of functional receptor at the cell surface. Mammalian non-activated Notch receptor is internalized, ubiquitylated by the Itch E3 ubiquitin ligase and degraded in the lysosomes. Here, we show that ß-arrestins are necessary for Itch-Notch interaction and for Itch-driven ubiquitylation and degradation of Notch. Interestingly, ß-arrestins do not directly bind Itch but heterodimerize with a member of another subfamily of arrestins called ARRDC1 or α-arrestin 1, which harbors PPxY motifs that allow direct interaction with Itch. Cells transfected with ARRDC1 mutated in PPxY motifs show reduced Itch-mediated Notch ubiquitylation and impaired lysosomal degradation of Notch, as observed in ß-arrestin(-/-) or Itch(-/-) cells. Our data show for the first time that ARRDC1 and ß-arrestins heterodimerize and cooperate in the same complex to promote non-activated Notch receptor degradation, thus acting as negative regulators of Notch signaling.

The signalling factor PI3K is a specific regulator of the clathrin-independent dynamin-dependent endocytosis of IL-2 receptors.

Receptor-mediated endocytosis is an essential process used by eukaryotic cells to internalise many molecules. Several clathrin-independent endocytic routes exist, but the molecular mechanism of each pathway remains to be uncovered. The present study focuses on a clathrin-independent dynamin-dependent pathway used by interleukin 2 receptors (IL-2R), essential players of the immune response. Ras-related C3 botulinum toxin substrate (Rac1) and its targets, the p21-activated kinases (Pak), are specific regulators of this pathway, acting on cortactin and actin polymerization. The present study reveals a dual and specific role of phosphatidylinositol 3-kinase (PI3K) in IL-2R endocytosis. Inhibition of the catalytic activity of PI3K strongly affects IL-2R endocytosis, in contrast to transferrin (Tf) uptake, a marker of the clathrin-mediated pathway. Moreover, Vav2, a GTPase exchange factor (GEF) induced upon PI3K activation, is specifically involved in IL-2R entry. The second action of PI3K is through its regulatory subunit, p85α, which binds to and recruits Rac1 during IL-2R internalisation. Indeed, the overexpression of a p85α mutant missing the Rac1 binding motif leads to the specific inhibition of IL-2R endocytosis. The inhibitory effect of this p85α mutant could be rescued by the overexpression of either Rac1 or the active form of Pak, indicating that p85α acts upstream of the Rac1-Pak cascade. Finally, biochemical and fluorescent microscopy techniques reveal an interaction between p85α, Rac1 and IL-2R that is enhanced by IL-2. In summary, our results indicate a key role of class I PI3K in IL-2R endocytosis that creates a link with IL-2 signalling.

Do kidney histology lesions predict long-term kidney function after liver transplantation?

Histological renal lesions observed after liver transplantation are complex, multifactorial, and interrelated. The aims of this study were to determine whether kidney lesions observed at five yr after liver transplantation can predict long-term kidney function. Ninety-nine liver transplant patients receiving calcineurin inhibitor (CNI)-based immunosuppression, who had undergone a kidney biopsy at 60±48 months post-transplant, were included in this follow-up study. Kidney biopsies were scored according to the Banff classification. Estimated glomerular filtration rate (eGFR) was assessed at last follow-up, that is, 109±48 months after liver transplantation. eGFR decreased from 92±33 mL/min at transplantation to 63±19 mL/min after six months, to 57±17 mL/min at the kidney biopsy, to 54±24 mL/min at last follow-up (p<0.0001). At last follow-up, only three patients required renal replacement therapy. After the kidney biopsy, 13 patients were converted from CNIs to mammalian target of rapamycin inhibitors, but no significant improvement in eGFR was observed after conversion. Elevated eGFR at six months post-transplant and a lower fibrous intimal thickening score (cv) observed at five yr post-transplant were the two independent predictive factors for eGFR≥60 mL/min at nine yr post-transplant. Long-term kidney function seems to be predicted by the kidney vascular lesions.

Icy: an open bioimage informatics platform for extended reproducible research.

Current research in biology uses evermore complex computational and imaging tools. Here we describe Icy, a collaborative bioimage informatics platform that combines a community website for contributing and sharing tools and material, and software with a high-end visual programming framework for seamless development of sophisticated imaging workflows. Icy extends the reproducible research principles, by encouraging and facilitating the reusability, modularity, standardization and management of algorithms and protocols. Icy is free, open-source and available at http://icy.bioimageanalysis.org/.

Quantitative elastography of renal transplants using supersonic shear imaging: a pilot study.

PURPOSE: To evaluate the reliability of quantitative ultrasonic measurement of renal allograft elasticity using supersonic shear imaging (SSI) and its relationship with parenchymal pathological changes. MATERIALS AND METHODS: Forty-three kidney transplant recipients (22 women, 21 men) (mean age, 51 years; age range, 18-70 years) underwent SSI elastography, followed by biopsy. The quantitative measurements of cortical elasticity were performed by two radiologists and expressed in terms of Young's modulus (kPa). Intra- and inter-observer reproducibility was assessed (Kruskal-Wallis test and Bland-Altman analysis), as well as the correlation between elasticity values and clinical, biological and pathological data (semi-quantitative Banff scoring). Interstitial fibrosis was evaluated semi-quantitatively by the Banff score and measured by quantitative image analysis. RESULTS: Intra- and inter-observer variation coefficients of cortical elasticity were 20 % and 12 %, respectively. Renal cortical stiffness did not correlate with any clinical parameters, any single semi-quantitative Banff score or the level of interstitial fibrosis; however, a significant correlation was observed between cortical stiffness and the total Banff scores of chronic lesions and of all elementary lesions (R = 0.34, P = 0.05 and R = 0.41, P = 0.03,respectively). CONCLUSION: Quantitative measurement of renal cortical stiffness using SSI is a promising non-invasive tool to evaluate global histological deterioration. KEY POINTS : • Supersonic shear imaging elastography can measure cortical stiffness in renal transplants • The level of cortical stiffness is correlated with the global degree of tissue lesions • The global histological deterioration of transplanted kidneys can be quantified using elastography.

New computerized color image analysis for the quantification of interstitial fibrosis in renal transplantation.

BACKGROUND: Chronic allograft injury, the primary cause of late allograft failure in renal transplantation, can be diagnosed early at a preclinical stage by histopathological changes such as interstitial fibrosis (IF). Currently, assessed by semiquantitative analysis in the Banff classification, IF quantification is limited by pathologist's subjective interpretation. METHODS: We have designed algorithms dedicated to quantify IF by computerized color image analysis. This innovative and objective software automatically extracts the green areas characteristic of IF in Masson's trichrome based on color image segmentation followed by removal of nonspecific IF staining (capsula, sclerosis glomeruli and normal glomeruli, normal basement membrane) and computes an index. It also counts automatically the number of glomeruli. Sixty-seven Masson stained renal transplant biopsies at various IF stages were imaged using a digital color camera mounted on a microscope. We tested the robustness of the method against varying acquisition parameters. RESULTS: We demonstrated that the parameters do not have an impact on this quantification and that the algorithm is able to handle biopsy color variations. The intra- and interobserver reproducibility was good (P<0.003). The kappa coefficient that was performed on another set of 90 biopsies to evaluate the concordance of our method with an expert Banff quantification was 0.68, indicating a substantial agreement. Finally, the computerized IF correlated with renal function. CONCLUSION: This study demonstrates that computerized color image analysis is a reliable and reproducible method to evaluate renal IF in routine practice and in multi-centric studies.