Using Speckle to Measure Tissue Dispersion in Optical Coherence Tomography

Tissue dispersion could be used as a marker of early disease changes to further improve the diagnostic potential of Optical Coherence Tomography (OCT). However, most methods to measure dispersion, described in the literature, rely on the presence of distinct and strong reflectors and are, therefore, rarely applicable in vivo. A novel technique has been developed which estimates the dispersion-induced resolution degradation from the image speckle and, as such, is applicable in situ. This method was verified experimentally ex vivo and was applied to the classification of a set of normal and cancerous colon OCT images resulting in 96% correct classificatio

Altered metabolic pathways in clear cell renal cell carcinoma: A meta-analysis and validation study focused on the deregulated genes and their associated networks.

<p>Clear cell renal cell carcinoma (ccRCC) is the predominant subtype of renal cell carcinoma (RCC). It is one of the most therapy-resistant carcinomas, responding very poorly or not at all to radiotherapy, hormonal therapy and chemotherapy. A more comprehensive understanding of the deregulated pathways in ccRCC can lead to the development of new therapies and prognostic markers. We performed a meta-analysis of 5 publicly available gene expression datasets and identified a list of co-deregulated genes, for which we performed extensive bioinformatic analysis coupled with experimental validation on the mRNA level. Gene ontology enrichment showed that many proteins are involved in response to hypoxia/oxygen levels and positive regulation of the VEGFR signaling pathway. KEGG analysis revealed that metabolic pathways are mostly altered in ccRCC. Similarly, Ingenuity Pathway Analysis showed that the antigen presentation, inositol metabolism, pentose phosphate, glycolysis/gluconeogenesis and fructose/mannose metabolism pathways are altered in the disease. Cellular growth, proliferation and carbohydrate metabolism, were among the top molecular and cellular functions of the co-deregulated genes. qRT-PCR validated the deregulated expression of several genes in Caki-2 and ACHN cell lines and in a cohort of ccRCC tissues. <i>NNMT</i> and <i>NR3C1</i> increased expression was evident in ccRCC biopsies from patients using immunohistochemistry. ROC curves evaluated the diagnostic performance of the top deregulated genes in each dataset. We show that metabolic pathways are mostly deregulated in ccRCC and we highlight those being most responsible in its formation. We suggest that these genes are candidate predictive markers of the disease.</p&gt

COL4A3/COL4A4 Mutations Producing Focal Segmental Glomerulosclerosis and Renal Failure in Thin Basement Membrane Nephropathy

<p>Mutations in the COL4A3/COL4A4 genes of type IV collagen have been found in ~40% of cases of thin basement membrane nephropathy, which is characterized by microscopic hematuria and is classically thought to cause proteinuria and chronic renal failure rarely. Here we report our observations of 116 subjects from 13 Cypriot families clinically affected with thin basement membrane nephropathy. These families first came to our attention because they segregated microscopic hematuria, mild proteinuria, and variable degrees of renal impairment, but a dual diagnosis of focal segmental glomerulosclerosis (FSGS) and thin basement membrane nephropathy was made in 20 biopsied cases. Molecular studies identified founder mutations in both COL4A3 and COL4A4 genes in 10 families. None of 82 heterozygous patients had any extrarenal manifestations, supporting the diagnosis of thin basement membrane nephropathy. During follow-up of up to three decades, 31 of these 82 patients (37.8%) developed chronic renal failure and 16 (19.5%) reached end-stage renal disease. Mutations G1334E and G871C were detected in seven and three families, respectively, and were probably introduced by founders. We conclude that these particular COL4A3/COL4A4 mutations either predispose some patients to FSGS and chronic renal failure, or that thin basement membrane nephropathy sometimes coexists with another genetic modifier that is responsible for FSGS and progressive renal failure. The findings presented here do not justify the labelling of thin basement membrane nephropathy as a benign condition with excellent prognosis.</p&gt

Clinico-pathological correlations in 127 patients in 11 large pedigrees, segregating one of three heterozygous mutations in the COL4A3/ COL4A4 genes associated with familial haematuria and significant late progression to proteinuria and chronic kidney disease from focal segmental glomerulosclerosis

<p><strong>Background.</strong> Heterozygous mutations in the <i>COL4A3/ COL4A4</i> genes are currently thought to be responsible for familial benign microscopic haematuria and maintenance of normal long-term kidney function.</p><p><strong>Methods.</strong> We report on 11 large Cypriot pedigrees with three such mutations. A total of 236 at-risk family members were genetically studied, and 127 (53.8%) carried a heterozygous mutation. Clinico-pathological correlations were available in all of these patients. Renal biopsies in 21 of these patients all showed various stages of focal, segmental glomerulosclerosis (FSGS). Thirteen of these biopsies were also studied with EM and showed thinning of the glomerular basement membrane.</p><p><strong>Results.</strong> Mutation G1334E (<i>COL4A3</i>) was found in six pedigrees, mutation G871C (<i>COL4A3</i>) in four and mutation 3854delG (<i>COL4A4</i>) in one pedigree. Clinical and laboratory correlations in all 127 mutation carriers (MC) showed that microscopic haematuria was the only urinary finding in patients under age 30. The prevalence of 'haematuria alone' fell to 66% between 31 and 50 years, to 30% between 51 and 70 and to 23% over age 71. Proteinuria with CRF developed on top of haematuria in 8% of all MC between 31 and 50 years, to 25% between 51 and 70 years and to 50% over 71 years. Altogether 18 of these 127 MC (14%) developed ESRD at a mean age of 60 years. Two members with different mutations married, and two of their children inherited both mutations and developed adolescent, autosomal recessive Alport syndrome (ATS), confirming that these mutations are pathogenic.</p><p><strong>Conclusions.</strong> Our data confirm for the first time a definite association of heterozygous <i>COL4A3/COL4A4</i> mutations with familial microscopic haematuria, thin basement membrane nephropathy and the late development of familial proteinuria, CRF, and ESRD, due to FSGS, indicating that the term 'benign familial haematuria' is a misnomer, at least in this cohort. A strong hypothesis for a causal relationship between these mutations and FSGS is also made. Benign familial haematuria may not be so benign as commonly thought.</p&gt

Familial C3 Glomerulopathy Associated with CFHR5 Mutations: Clinical Characteristics of 91 Patients in 16 Pedigrees

Background and objectives Complement factor H and related proteins (CFHR) are key regulators of the alternative complement pathway, where loss of function mutations lead to a glomerulopathy with isolated mesangial C3 deposits without immunoglobulins. Gale et al. (12) reported on 26 patients with the first familial, hematuric glomerulopathy caused by a founder mutation in the CFHR5 gene in patients of Cypriot descent living in the United Kingdom. CFHR5 nephropathy is clinically characterized by continuous microscopic hematuria whereas some patients present with additional episodes of synpharyngitic macrohematuria, associated with infection and pyrexia. A subgroup of patients, particularly men, develop additional proteinuria, hypertension, and chronic renal disease or ESRD.Design, setting, participants, & measurements We herewith expand significantly on the study by Gale et al., reporting on histologic, molecular, and clinical findings in 91 patients from 16 families with the same founder mutation.Results Eighty-two patients (90%) exhibited microscopic hematuria; 51(62%), exhibited only microscopic hematuria, whereas the remaining 31 additionally had proteinuria (38%); 28 proteinuric patients developed chronic renal failure (CRF). Among carriers of CFHR5 mutation aged >50 years, 80% of the men and 21% of the women developed CRF; 18 developed ESRD (14 men [78%], 4 women [22%]).Conclusions The diagnosis of CFHR5-related, isolated C3 glomerulopathy was established in 2009 using newly described mutation analysis after decades of follow-up with unclear diagnoses, occasionally confused with IgA nephropathy. This larger patient cohort establishes the clinical course, significant variable expressivity, and marked gender difference regarding the development of CRF and ESRD. Clin J Am Soc Nephrol 6: 1436-1446, 2011. doi: 10.2215/CJN.0954101

Evidence for activation of the unfolded protein response in collagen IV nephropathies

Thin-basement-membrane nephropathy (TBMN) and Alport syndrome (AS) are progressive collagen IV nephropathies caused by mutations in COL4A3/A4/A5 genes. These nephropathies invariably present with microscopic hematuria and frequently progress to proteinuria and CKD or ESRD during long-term follow-up. Nonetheless, the exact molecular mechanisms by which these mutations exert their deleterious effects on the glomerulus remain elusive. We hypothesized that defective trafficking of the COL4A3 chain causes a strong intracellular effect on the cell responsible for COL4A3 expression, the podocyte. To this end, we overexpressed normal and mutant COL4A3 chains (G1334E mutation) in human undifferentiated podocytes and tested their effects in various intracellular pathways using a microarray approach. COL4A3 overexpression in the podocyte caused chain retention in the endoplasmic reticulum (ER) that was associated with activation of unfolded protein response (UPR)–related markers of ER stress. Notably, the overexpression of normal or mutant COL4A3 chains differentially activated the UPR pathway. Similar results were observed in a novel knockin mouse carrying the Col4a3-G1332E mutation, which produced a phenotype consistent with AS, and in biopsy specimens from patients with TBMN carrying a heterozygous COL4A3-G1334E mutation. These results suggest that ER stress arising from defective localization of collagen IV chains in human podocytes contributes to the pathogenesis of TBMN and AS through activation of the UPR, a finding that may pave the way for novel therapeutic interventions for a variety of collagenopathies

Frequency of COL4A3/COL4A4 mutations amongst families segregating glomerular microscopic hematuria and evidence for activation of the unfolded protein response. Focal and segmental glomerulosclerosis is a frequent development during ageing.

Familial glomerular hematuria(s) comprise a genetically heterogeneous group of conditions which include Alport Syndrome (AS) and thin basement membrane nephropathy (TBMN). Here we investigated 57 Greek-Cypriot families presenting glomerular microscopic hematuria (GMH), with or without proteinuria or chronic kidney function decline, but excluded classical AS. We specifically searched the COL4A3/A4 genes and identified 8 heterozygous mutations in 16 families (28,1%). Eight non-related families featured the founder mutation COL4A3-p.(G1334E). Renal biopsies from 8 patients showed TBMN and focal segmental glomerulosclerosis (FSGS). Ten patients (11.5%) reached end-stage kidney disease (ESKD) at ages ranging from 37-69-yo (mean 50,1-yo). Next generation sequencing of the patients who progressed to ESKD failed to reveal a second mutation in any of the COL4A3/A4/A5 genes, supporting that true heterozygosity for COL4A3/A4 mutations predisposes to CRF/ESKD. Although this could be viewed as a milder and late-onset form of autosomal dominant AS, we had no evidence of ultrastructural features or extrarenal manifestations that would justify this diagnosis. Functional studies in cultured podocytes transfected with wild type or mutant COL4A3 chains showed retention of mutant collagens and differential activation of the unfolded protein response (UPR) cascade. This signifies the potential role of the UPR cascade in modulating the final phenotype in patients with collagen IV nephropathies