Desmosomes undergo dynamic architectural changes during assembly and maturation.

Desmosomes are macromolecular cell-cell junctions critical for maintaining adhesion and resisting mechanical stress in epithelial tissue. Desmosome assembly and the relationship between maturity and molecular architecture are not well understood. To address this, we employed a calcium switch assay to synchronize assembly followed by quantification of desmosome nanoscale organization using direct Stochastic Optical Reconstruction Microscopy (dSTORM). We found that the organization of the desmoplakin rod/C-terminal junction changed over the course of maturation, as indicated by a decrease in the plaque-to-plaque distance, while the plaque length increased. In contrast, the desmoplakin N-terminal domain and plakoglobin organization (plaque-to-plaque distance) were constant throughout maturation. This structural rearrangement of desmoplakin was concurrent with desmosome maturation measured by E-cadherin exclusion and increased adhesive strength. Using two-color dSTORM, we showed that while the number of individual E-cadherin containing junctions went down with the increasing time in high Ca2+, they maintained a wider desmoplakin rod/C-terminal plaque-to-plaque distance. This indicates that the maturation state of individual desmosomes can be identified by their architectural organization. We confirmed these architectural changes in another model of desmosome assembly, cell migration. Desmosomes in migrating cells, closest to the scratch where they are assembling, were shorter, E-cadherin enriched, and had wider desmoplakin rod/C-terminal plaque-to-plaque distances compared to desmosomes away from the wound edge. Key results were demonstrated in three cell lines representing simple, transitional, and stratified epithelia. Together, these data suggest that there is a set of architectural programs for desmosome maturation, and we hypothesize that desmoplakin architecture may be a contributing mechanism to regulating adhesive strength.

Steroid hormones and human choriogonadotropin influence the distribution of alpha6-integrin and desmoplakin 1 in gland-like endometrial epithelial spheroids.

In human glandular endometrial epithelial cells, desmosomal and adherens junction proteins have been shown to extend from a subapically restricted lateral position to the entire lateral membrane during the implantation window of the menstrual cycle. Similarly, a menstrual cycle stage-dependent redistribution of the extracellular matrix adhesion protein α6-integrin has been reported. These changes are believed to be important for endometrial receptiveness and successful embryo implantation. To prove the hypothesis that steroid hormones and human choriogonadotropin can induce the redistribution of these adhesion molecules, we used the human endometrial cell line Ishikawa in a 3D culture system. Gland-like spheroids were grown in reconstituted basement membrane (Matrigel™). The lumen-bearing spheroids were treated for 2 or 4 days with ovarian steroids or human choriogonadotropin and then assessed by immunofluorescence microscopy. In addition, human endometrial biopsies were obtained from patients, who were in therapy for assisted reproductive technology, and were examined in parallel. Lateral redistribution of the desmosomal plaque protein desmoplakin 1 was observed in the spheroids treated either with progesterone, medroxyprogesterone acetate or human choriogonadotropin. Furthermore, the extracellular matrix adhesion protein α6-integrin showed an increased lateral membrane localization upon gestagen stimulation in the 3D culture system. The results of this study demonstrate that the 3D endometrial Ishikawa cell culture might be suited as an experimental model system to prove the effect of hormonal changes like those occurring during the window of implantation.

Importancia de los hallazgos de la resonancia magnética cardiaca en el diagnóstico de la miocardiopatía arritmogénica del ventrículo izquierdo / Importance of cardiac magnetic resonance findings in the diagnosis of left dominant arrythmogenic cardiomyopathy

INTRODUCCIÓN Y OBJETIVOS: Recientemente, la miocardiopatía arritmogénica del ventrículo izquierdo (MCAVI) ha sido reconocida como parte del espectro de la miocardiopatía arritmogénica. Se caracteriza por el reemplazo fibroadiposo de la pared de dicho ventrículo. Se describen las formas de presentación clínica más frecuentes, hallazgos de imagen y eventos en el seguimiento, destacando la importancia de la resonancia magnética cardiaca (RMC). MÉTODOS: Registro prospectivo de pacientes con hallazgos compatibles con MCAVI. Se realizó análisis de imagen de RMC y seguimiento clínico. El objetivo primario fue la aparición de eventos cardiovasculares adversos mayores (MACE) durante el seguimiento, definidos como muerte súbita cardiaca, arritmias ventriculares sostenidas y trasplante cardiaco. RESULTADOS: Se incluyeron 74 pacientes consecutivos (edad media 48,6 años, 50 varones [67,6%]). Las indicaciones más frecuentes para la RMC fueron dolor torácico con coronariografía normal, arritmias ventriculares y sospecha de miocardiopatías. Los principales hallazgos de RMC fueron: realce tardío meso-subepicárdico (91,9%), infiltración grasa subepicárdica (83,8%) y anomalías segmentarias de la contractilidad del ventrículo izquierdo (47,9%). En un seguimiento medio de 3,74 años, 24 pacientes (32,4%) presentaron un MACE (muerte súbita cardiaca 8,1%, arritmias ventriculares sostenidas 21,6% y trasplante cardiaco 4,1%). La presencia en RMC de realce tardío grave, predice independientemente la aparición de MACE, además del hecho de ser varón y practicar deporte. CONCLUSIONES: La RMC es una herramienta clave para diagnosticar la MCAVI. La infiltración grasa subepicárdica y el realce tardío meso-subepicárdico son hallazgos característicos. El pronóstico de esta población es pobre con una alta incidencia de muerte súbita cardiaca y arritmias ventriculares

INTRODUCTION AND OBJECTIVES: Left dominant arrhythmogenic cardiomyopathy (LDAC) has recently been recognized as falling on the spectrum of arrhythmogenic cardiomyopathy. It is characterized by fibroadipose replacement of the left ventricle. The aim of this study was to describe the most frequent forms of clinical presentation of LDAC, imaging findings, and events at follow-up, highlighting the importance of cardiac magnetic resonance (CMR). METHODS: Prospective registry of patients with findings compatible with LDAC. CMR image analysis and clinical follow-up was performed. The primary endpoint was the appearance of major adverse cardiovascular events (MACE) during follow-up, defined as sudden cardiac death, sustained ventricular arrhythmias, and heart transplant. RESULTS: We included 74 consecutive patients (mean age, 48.6 years; 50 men [67.6%]). The most frequent CMR indications were chest pain with normal coronary angiography, ventricular arrhythmias, and suspicion of cardiomyopathies. The main CMR findings were midwall and/or subepicardial pattern of late gadolinium enhancement (91.9%), fatty epicardial infiltration (83.8%), and left ventricle segmental contractility abnormalities (47.9%). At a mean follow-up of 3.74 years, 24 patients (32.4%) had a MACE (sudden cardiac death 8.1%, sustained ventricular arrhythmias 21.6%, and heart transplant 4.1%). Independent predictors for the appearance for MACE were a CMR study showing severe late gadolinium enhancement, male sex, and practicing sports. CONCLUSIONS: CMR is a key tool for diagnosing LDAC. Characteristic findings are subepicardial fatty infiltration and midwall-subepicardial late gadolinium enhancement. The prognosis of this population is poor with a high incidence of sudden cardiac death and ventricular arrhythmias

Pattern-Specific Loss of Desmoplakin I and II Immunoreactivity in Erythema Multiforme and its Variants: A Possible Aid in Histologic Diagnosis.

Erythema multiforme (EM), Stevens-Johnson syndrome (SJS), and toxic epidermal necrolysis (TEN) comprise a family of mucocutaneous diseases associated with significant morbidity and mortality. Previous studies have confirmed the presence of autoantibodies to desmoplakin (Dp) I and II in patients with EM, SJS, and TEN. Truncated Dp production leads to characteristic changes visible on light microscopy: perinuclear clumping of keratin filaments and dyskeratotic keratinocyte. Based on these observations, the question arises as to whether a loss of Dp immunoreactivity in skin biopsies could serve as a diagnostic marker of EM, SJS, and TEN. This study analyzed Dp immunostaining patterns in 20 patients with EM or SJS/TEN. To assess the specificity of this approach, Dp immunostaining was also performed on specimens from patients with 5 potential histologic mimics of EM, SJS, and TEN. All of the samples from patients with EM, SJS, and TEN demonstrated absent or markedly diminished staining for Dp. A χ test demonstrated a statistically significant difference between the staining patterns in EM, SJS, and TEN and each of the other diagnostic groups that were investigated. This is the first report demonstrating statistically significant specificity of Dp staining patterns in EM/SJS/TEN as compared with other interface dermatitides.

TGF-beta1 pathway activation and adherens junction molecular pattern in nonsyndromic mitral valve prolapse.

AIMS: Dysregulation of the transforming growth factor beta (TGF-ß) 1 pathway has been associated with either syndromic or isolated mitral valve (MV) prolapse due to myxoid degeneration (floppy MV). The activation of Smad receptor-mediated intracellular TGF-ß pathway and its effect on adherens junction (AJ) molecular pattern of activated valvular interstitial cells (VICs) in MV prolapse are herein investigated. METHODS: Floppy MV leaflets were obtained from 30 patients (24 males, mean age 55.5±12.7 years) who underwent surgical repair, and 10 age- and sex-matched Homograft Tissue Bank samples served as controls. MV leaflet cellular and extracellular matrix composition, including collagen I and III, was evaluated by histology and transmission electron microscopy. Smad2 active phosphorylated form (p-Smad2), α-smooth muscle actin (α-SMA), and junctional proteins (N-cadherin, cadherin-11, ß-catenin, plakoglobin, plakophilin-2) in VICs were assessed by immunohistochemistry and immunofluorescence and confirmed by immunoblotting. Quantitative real-time polymerase chain reaction was carried out for components of TGF-ß pathway cascade and filamin A (FLN-A). RESULTS: Floppy MV leaflets were thicker (P<.001) and had higher α-SMA+ cell density (P=.002) and collagen III expression (P<.001) than controls. Enhanced p-Smad2 nuclear immunoreactivity (P<.001) and TGF-ß1 gene (P=.045), TIMP1 (P=.020), and CTGF (P=.047) expression but no differences in FLN-A and total Smad2 gene expression levels were found between floppy MV and controls. Higher expression of cadherin-11, either exclusively or in colocalization with N-cadherin, and aberrant presence of plakophilin-2 at the AJ were found in floppy MV vs. CONCLUSIONS: TGF-ß1 pathway activation in nonsyndromic MV prolapse induces VICs differentiation into contractile myofibroblasts and is associated with changes in the molecular pattern of the AJ, with increased cadherin-11 and aberrant plakophilin-2 expression. AJ reinforcement might promote latent TGF-ß1 activation leading to extracellular matrix remodeling in floppy MV.

Disruption of Tgfbr2 in odontoblasts leads to aberrant pulp calcification.

Transforming growth factor ß (TGF-ß) signaling has been implicated in dentin formation and repair; however, the molecular mechanisms underlying dentin formation remain unclear. To address the role of TGF-ß signaling in dentin formation, we analyzed odontoblast-specific Tgfbr2 conditional knockout mice. The mutant mice had aberrant teeth with thin dysplastic dentin and pulpal obliteration, similar to teeth from human patients with dentinogenesis imperfecta type II and dentin dysplasia. In mutant, the odontoblasts lost their cellular polarity, and matrix secretion was disrupted after mantle dentin formation. As a consequence, the amount of predentin decreased significantly, and an ectopic fibrous matrix was formed below the odontoblast layer. This matrix gradually calcified and obliterated the pulp chamber with increasing age. Immunohistochemistry revealed decreased expression of alkaline phosphatase in mutant odontoblasts. In mutant dentin, Dsp expression was reduced, but Dmp1 expression increased significantly. Collagen type I, biglycan, and Dsp were expressed in the ectopic matrix. These results suggest that loss of responsiveness to TGF-ß in odontoblasts results in impaired matrix formation and pulpal obliteration. Our study indicates that TGF-ß signaling plays an important role in dentin formation and pulp protection. Furthermore, our findings may provide new insight into possible mechanisms underlying human hereditary dentin disorders and reparative dentin formation.

Plakoglobin: a diagnostic marker of arrhythmogenic right ventricular cardiomyopathy in forensic pathology?

PURPOSE: The histopathological diagnosis of arrhythmogenic right ventricular cardiomyopathy (ARVC) can be challenging in forensic medicine. Immunohistochemical myocardial analysis for plakoglobin has been suggested as a new diagnostic test for ARVC. We examined this in the setting of forensic pathology, applying this method to forensic autopsy samples. METHODS: We performed immunohistochemical staining for plakoglobin on 40 myocardial samples with an autopsy diagnosis of ARVC. In addition, histopathological reevaluation was performed applying the revised 2010 task force criteria including morphometric analysis. Myocardial samples from 15 subjects without heart disease were used as controls. RESULTS: Based on the histopathological reevaluation, 38 out of 40 cases were categorized as ARVC. A marked reduction in the plakoglobin staining was seen in 26 out of 38 myocardial samples in the ARVC-group. Of the two samples categorized as not ARVC, one showed reduced plakoglobin staining and one sample had normal staining. No control samples showed reduced plakoglobin staining. CONCLUSIONS: In conclusion, our study displayed reduced plakoglobin staining in approximately 2/3 of myocardial samples with ARVC. Our data suggests that immunostaining for plakoglobin might serve as an additional diagnostic marker of ARVC in forensic pathology, but additional validation is required.

Salivary proteomics in bisphosphonate-related osteonecrosis of the jaw.

OBJECTIVE: The objective of this study was to identify differentially expressed salivary proteins in bisphosphonate-related osteonecrosis of the jaw (BRONJ) patients that could serve as biomarkers for BRONJ diagnosis. SUBJECTS AND METHODS: Whole saliva obtained from 20 BRONJ patients and 20 controls were pooled within groups. The samples were analyzed using iTRAQ-labeled two-dimensional liquid chromatography-tandem mass spectrometry. RESULTS: Overall, 1340 proteins were identified. Of these, biomarker candidates were selected based on P-value (<0.001), changes in protein expression (≥1.5-fold increase or decrease), and unique peptides identified (≥2). Three comparisons made between BRONJ and control patients identified 200 proteins to be differentially expressed in BRONJ patients. A majority of these proteins were predicted to have a role in drug metabolism and immunological and dermatological diseases. Of all the differentially expressed proteins, we selected metalloproteinase-9 and desmoplakin for further validation. Immunoassays confirmed increased expression of metalloproteinase-9 in individual saliva (P = 0.048) and serum samples (P = 0.05) of BRONJ patients. Desmoplakin was undetectable in saliva. However, desmoplakin levels tended to be lower in BRONJ serum than controls (P = 0.157). CONCLUSIONS: Multiple pathological reactions are involved in BRONJ development. One or more proteins identified by this study may prove to be useful biomarkers for BRONJ diagnosis. The role of metalloproteinase-9 and desmoplakin in BRONJ requires further investigation.

Clinical usefulness of immunohistochemistry for plakoglobin, N-cadherin, and connexin-43 in the diagnosis of arrhythmogenic right ventricular cardiomyopathy.

Diagnosing arrhythmogenic right ventricular cardiomyopathy (ARVC) is often challenging because no single diagnostic tool is available to detect the disease. We evaluated whether analysis of plakoglobin, N-cadherin, and connexin-43 immunoreactivity can be used as a significant test in diagnosis of ARVC. We selected subjects with suspicion of ARVC (n=22) in patients who underwent endomyocardial biopsy (EMB) in Kyungpook National University Hospital (n=1326). The patients (n=22) were classified into definite ARVC patients (n=17) and borderline ARVC (n=5). We selected control subjects (n=20) who were autopsied and died of non-cardiac disease. Hematoxylin-eosin, Masson's trichrome, and immunohistochemical stains for plakoglobin, N-cadherin, and connexin-43 were used for all specimens. Reduced immunoreactivity of plakoglobin was observed in 13 (76%) of the 17 patients with a definite ARVC and in 4 (80%) of the 5 patients with a borderline ARVC. All subjects displayed no significant reduction of the immunoreactivity for connexin-43 as well as for N-cadherin. Our investigation revealed that the immunohistochemical analysis for plakoglobin had an accuracy of 81%, 76% sensitivity, and 84% specificity in diagnosis of ARVC. Results of our study showed that the immunohistochemical analysis of plakoglobin had a relatively high sensitivity and specificity in ARVC, but immunohistochemistry for plakoglobin alone could not be relied upon as a diagnostic test for ARVC. We confirmed that N-cadherin and connexin-43 had no diagnostic value in ARVC.

Identification of Streptococcus parasanguinis DNA contamination in human buccal DNA samples.

BACKGROUND: The use of buccal swabs in clinical and scientific studies is a very popular method of collecting DNA, due to its non-invasive nature of collection. However, contamination of the DNA sample may interfere with analysis. FINDINGS: Here we report the finding of Streptococcus parasanguinis bacterial DNA contamination in human buccal DNA samples, which led to preferential amplification of bacterial sequence with PCR primers designed against human sequence. CONCLUSION: Contamination of buccal-derived DNA with bacterial DNA can be significant, and may influence downstream genetic analysis. One needs to be aware of possible bacterial contamination when interpreting abnormal findings following PCR amplification of buccal swab DNA samples.

Immortalization and characterization of human dental papilla cells with odontoblastic differentiation.

AIM: To establish a cell line of immortalized human dental papilla cells (hDPCs). METHODOLOGY: Primary hDPCs were cultured and infected with lentivirus containing the hTERT gene. Integration and transcription of the hTERT gene were verified by PCR. The characteristics of the cells, such as morphology, proliferation and mineralization, were analysed. Also, the expression of odontoblastic-related markers including ALP, DMP1, DLX3, OSX, DSP and Nestin, was detected by immunohistochemistry and real-time RT-PCR. RESULTS: hTERT gene was integrated into genomic DNA of immortalized cells (hDPC-TERT) and transcribed into mRNA. With long-time culture, hDPC-TERT bypassed senescence and grew over 120 population doublings. hDPC-TERT cells have a higher proliferation rate, but retain the phenotypic characteristics of the primary hDPCs, and so was ALP activity and mineralization activity. Furthermore, the hDPC-TERT cells express no DSP and Nestin with maintenance medium, but highly expressed DSP and Nestin after odontoblastic induction. CONCLUSIONS: A line of immortalized human dental papilla cells, which remains in an undifferentiated state and has odontoblastic differentiation potential, was established. This cell line can be used as a cell model for studying the mechanism of the initiation of odontoblast differentiation.

Miocardiopatía arritmogénica con afectación predominante del ventrículo izquierdo por una mutación nueva «sin sentido» en desmoplaquina / Left dominant arrhythmogenic cardiomyopathy caused by a novel nonsense mutation in desmoplakin

La miocardiopatía arritmogénica predominantemente izquierda (MCAI) presenta características fenotípicas y genotípicas reflejadas en la familia española de cinco miembros que aquí describimos. Durante un catarro, un joven presentó una taquicardia ventricular con origen ventricular izquierdo y realce tardío en dicha localización. Su ECG basal mostró bajos voltajes, retraso en la activación terminal del QRS (cara inferior y V4-V6) y trastorno de la conducción auriculoventricular. Su biopsia endomiocárdica evidenció pérdida miocitaria y fibrosis. Aunque inicialmente fue catalogado de miocarditis, la evaluación familiar fue decisiva para sospechar una MCAI. El estudio genético identificó una mutación nueva en desmoplaquina tipo «sin sentido» (Q1866X) congruente con la presencia de una desmoplaquina truncada en muestras de piel de los afectados (AU)

Left dominant arrhythmogenic cardiomyopathy (LDAC) exhibits characteristic phenotypic and genetic features which were found in the five Spanish family members described in this study. Triggered by a cold, a young man presented with a ventricular tachycardia of left ventricular origin and left ventricular late gadolinium enhancement. His resting ECG showed low potentials, delayed ventricular depolarization (inferior and V4-V6 leads) and atrioventricular conduction disturbances. His endomyocardial biopsy revealed myocyte loss with interstitial fibrosis. Despite the initial diagnosis of myocarditis, familial screening was pivotal in confirming the diagnosis of LDAC. A novel nonsense mutation in the desmoplakin gene (Q1866X) and the truncated protein which it produces were observed in skin samples (AU)

A novel kind of tumor type-characteristic junction: plakophilin-2 as a major protein of adherens junctions in cardiac myxomata.

Using novel antibodies of high avidity to--and specificity for--the constitutive desmosomal plaque protein, plakophilin-2 (Pkp2), in a systematic study of the molecular composition of junctions connecting the cells of soft tissue tumors, we have discovered with immunocytochemical, biochemical and electron microscopical methods, a novel type of adherens junctions in all 32 cardiac myxomata examined. These junctions contain cadherin-11 as their major transmembrane glycoprotein, which we could repeatedly show in colocalization with N-cadherin, anchored in a cytoplasmic plaque formed by α- and ß-catenin, together with the further armadillo-type proteins plakoglobin, p120, p0071 and ARVCF. Surprisingly, all adherens junctions of these tumors contained, in addition, another major armadillo protein Pkp2, hitherto known as an obligatory and characteristic constituent of desmosomes in epithelium-derived tumors. We have not detected Pkp2 in a series of noncardiac myxomata studied in parallel. Therefore, we conclude that this acquisition of Pkp2, which we have recently also observed in some mesenchymally derived cells growing in culture, can also occur in tumorigenic transformations in situ. We propose to examine the marker value of Pkp2 in clinical diagnoses of cardiac myxomata and to develop Pkp2-targeted therapeutic reagents.

Cardiomyopathies: New test for arrhythmogenic right ventricular cardiomyopathy.

Arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C) is challenging to diagnose because of nonspecific findings, particularly in the early phases of the disease. Clinical diagnosis is made on the basis of several criteria, but these lack sensitivity. Asimaki et al. suggest that immunohistochemical analysis of myocardial desmosomal proteins is a highly sensitive and specific diagnostic test for ARVD/C.

Identification and quantification of preterm birth biomarkers in human cervicovaginal fluid by liquid chromatography/tandem mass spectrometry.

Spontaneous preterm birth (PTB) before 37 completed weeks of gestation resulting from preterm labor (PTL) is a leading contributor of perinatal morbidity and mortality. Early identification of at-risk women by reliable screening tests could alleviate this health issue; however, conventional methods such as obstetric history and clinical risk factors, uterine activity monitoring, biochemical markers, and cervical sonography for screening women at risk for PTB have proven unsuccessful in lowering the rate of PTB. Cervicovaginal fluid (CVF) might prove to be a useful, readily available biological fluid for identifying diagnostic PTB biomarkers. Human columnar epithelial endocervical-1 (End1) and vaginal (Vk2) cell secretomes were employed to generate a stable isotope labeled proteome (SILAP) standard to facilitate characterization and relative quantification of proteins present in CVF. The SILAP standard was prepared using stable isotope labeling by amino acids in cell culture (SILAC) of End1 and Vk2 through seven passages. The labeled secreted proteins from both cell lines were combined and characterized by liquid-chromatography-tandem mass spectrometry (LC-MS/MS). In total, 1211 proteins were identified in the End1-Vk2 SILAP standard, with 236 proteins being consistently identified in each of the replicates analyzed. Individual proteins were found to contain <0.5% of the endogenous unlabeled forms. Identified proteins were screened to provide a set of 15 candidates that have either previously been identified as potential PTB biomarkers or could be linked mechanistically to PTB. Stable isotope dilution LC-multiple reaction monitoring (MRM/MS) assays were then developed for conducting relative quantification of the 15 candidate biomarkers in human CVF samples from term and PTB cases. Three proteins were significantly elevated in PTB cases (desmoplakin isoform 1, stratifin, and thrombospondin 1 precursor), providing a foundation for further validation in larger patient cohorts.

A new diagnostic test for arrhythmogenic right ventricular cardiomyopathy.

BACKGROUND: The diagnosis of arrhythmogenic right ventricular cardiomyopathy (ARVC) can be challenging because the clinical presentation is highly variable and genetic penetrance is often low. METHODS: To determine whether a change in the distribution of desmosomal proteins can be used as a sensitive and specific diagnostic test for ARVC, we performed immunohistochemical analysis of human myocardial samples. RESULTS: We first tested myocardium from 11 subjects with ARVC; of these samples, 8 had desmosomal gene mutations. We also tested myocardium obtained at autopsy from 10 subjects with no clinical or pathological evidence of heart disease as control samples. All ARVC samples but no control samples showed a marked reduction in immunoreactive signal levels for plakoglobin (also known as gamma-catenin), a protein that links adhesion molecules at the intercalated disk to the cytoskeleton. Other desmosomal proteins showed variable changes, but signal levels for the nondesmosomal adhesion molecule N-cadherin were normal in all subjects with ARVC. To determine whether a diminished plakoglobin signal level was specific for ARVC, we analyzed myocardium from 15 subjects with hypertrophic, dilated, or ischemic cardiomyopathies. In every sample, levels of N-cadherin and plakoglobin signals at junctions were indistinguishable from those in control samples. Finally, we performed blinded immunohistochemical analysis of heart-biopsy samples from the Johns Hopkins ARVC registry. We made the correct diagnosis in 10 of 11 subjects with definite ARVC on the basis of clinical criteria and correctly ruled out ARVC in 10 of 11 subjects without ARVC, for a sensitivity of 91%, a specificity of 82%, a positive predictive value of 83%, and a negative predictive value of 90%. The plakoglobin signal level was reduced diffusely in ARVC samples, including those obtained in the left ventricle and the interventricular septum. CONCLUSIONS: Routine immunohistochemical analysis of a conventional endomyocardial-biopsy sample appears to be a highly sensitive and specific diagnostic test for ARVC.

Reduced expression of E-cadherin/catenin complex in hepatocellular carcinomas.

AIM: To examine the immunoreactivity of E-cadherin and four subtypes of catenin family in human hepatocellular carcinomas (HCCs) and to investigate the correlation between expression of E-cadherin/catenin complex and clinicopathologic parameters of HCC patients. METHODS: An immunohistochemical study for E-cadherin and catenins was performed on 97 formalin-fixed, paraffin-embedded specimens of HCC. RESULTS: Reduced expression of E-cadherin, alpha-, beta-, gamma-catenin and p120 was observed in 69%, 76%, 63%, 71% and 73%, respectively. Both expressions of E-cadherin and catenin components were significantly correlated with tumor grade (P = 0.000). It showed significant difference between expression of catenin members and tumor stage (P = 0.003, P = 0.017, P = 0.007 and P = 0.000, respectively). The reduced expression of E-cadherin in HCCs was significantly correlated with intrahepatic metastasis (IM) and capsular invasion (P = 0.008, P = 0.03, respectively). A close correlation was also observed between the expression of catenins and the tumor size (P = 0.002, P = 0.034, P = 0.016 and P = 0.000, respectively). In addition, the expression of each catenin was found correlated with IM (P = 0.012, P = 0.049, P = 0.026 and P = 0.014, respectively). No statistically significant difference was observed between the expression level of E-cadherin/catenin complex and lymph node permission, vascular invasion and satellite nodules. Interestingly, only expression of p120 showed correlation with AFP value (P = 0.035). The expression of E-cadherin was consistent with alpha-, beta-, gamma-catenin and p120 expression (P = 0.000). Finally, the abnormal expression of E-cadherin/catenin complex was significantly associated with patients' survival (P = 0.0253, P = 0.0052, P = 0.003, P = 0.0105 and P = 0.0016, respectively). Nevertheless, no component of E-cadherin/catenin complex was the independent prognostic factor of HCC patients. CONCLUSION: Down-regulated expressions of E-cadherin, catenins and p120 occur frequently in HCCs and contribute to the progression and development of tumor. It may be more exact and valuable to detect the co-expression of E-cadherin/catenin complex than to explore one of them in predicting tumor invasion, metastasis and patient's survival.

The cell adhesion molecule nectin-1 is critical for normal enamel formation in mice.

Nectin-1 is a member of a sub-family of immunoglobulin-like adhesion molecules and a component of adherens junctions. In the current study, we have shown that mice lacking nectin-1 exhibit defective enamel formation in their incisor teeth. Although the incisors of nectin-1-null mice were hypomineralized, the protein composition of the enamel matrix was unaltered. While strong immunostaining for nectin-1 was observed at the interface between the maturation-stage ameloblasts and the underlying cells of the stratum intermedium (SI), its absence in nectin-1-null mice correlated with separation of the cell layers at this interface. Numerous, large desmosomes were present at this interface in wild-type mice; however, where adhesion persisted in the mutant mice, the desmosomes were smaller and less numerous. Nectins have been shown to regulate tight junction formation; however, this is the first report showing that they may also participate in the regulation of desmosome assembly. Importantly, our results show that integrity of the SI-ameloblast interface is essential for normal enamel mineralization.