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Simple Summary Mesenchymal triple negative breast cancer subtype expresses genes involved in proliferation, epithelial to mesenchymal transition, stromal interaction and cell motility. Moreover, this subgroup is characterized by an immunosuppressive microenvironment. This review focuses on the intracellular pathways involved in tumorigenesis and cancer progression, as well as in the immune evasion mechanisms. Furthermore, we provide an overview of current clinical trials investigating the efficacy and safety of different therapeutic molecules for this aggressive subtype of triple negative breast cancer. The challenge is to restore immunocompetence by overcoming the chemo and immune-resistance profile of mesenchymal triple negative breast cancer to achieve a lasting response to therapy. Abstract The pathologic definition of triple negative breast cancer (TNBC) relies on the absence of expression of estrogen, progesterone and HER2 receptors. However, this BC subgroup is distinguished by a wide biological, molecular and clinical heterogeneity. Among the intrinsic TNBC subtypes, the mesenchymal type is defined by the expression of genes involved in the epithelial to mesenchymal transition, stromal interaction and cell motility. Moreover, it shows a high expression of genes involved in proliferation and an immune-suppressive microenvironment. Several molecular alterations along different pathways activated during carcinogenesis and tumor progression have been outlined and could be involved in immune evasion mechanisms. Furthermore, reverting epithelial to mesenchymal transition process could lead to the overcoming of immune-resistance. This paper reviews the current knowledge regarding the mesenchymal TNBC subtype and its response to conventional therapeutic strategies, as well as to some promising molecular target agents and immunotherapy. The final goal is a tailored combination of cytotoxic drugs, target agents and immunotherapy in order to restore immunocompetence in mesenchymal breast cancer patients.

Deregulated Notch signaling is associated with T-cell Acute Lymphoblastic Leukemia (T-ALL) development and progression. Increasing evidence reveals that Notch pathway has an important role in the invasion ability of tumor cells, including leukemia, although the underlying molecular mechanisms remain mostly unclear. Here, we show that Notch3 is a novel target protein of the prolyl-isomerase Pin1, which is able to regulate Notch3 protein processing and to stabilize the cleaved product, leading to the increased expression of the intracellular domain (N3IC), finally enhancing Notch3-dependent invasiveness properties. We demonstrate that the combined inhibition of Notch3 and Pin1 in the Notch3-overexpressing human leukemic TALL-1 cells reduces their high invasive potential, by decreasing the expression of the matrix metalloprotease MMP9. Consistently, Pin1 depletion in a mouse model of Notch3-induced T-ALL, by reducing N3IC expression and signaling, impairs the expansion/invasiveness of CD4(+)CD8(+) DP cells in peripheral lymphoid and non-lymphoid organs. Notably, in in silico gene expression analysis of human T-ALL samples we observed a significant correlation between Pin1 and Notch3 expression levels, which may further suggest a key role of the newly identified Notch3-Pin1 axis in T-ALL aggressiveness and progression. Thus, combined suppression of Pin1 and Notch3 proteins may be exploited as an additional target therapy for T-ALL.

Mutations in mitochondrial (mt) genes coding for mt-tRNAs are responsible for a range of syndromes, for which no effective treatment is available. We recently showed that the carboxy-terminal domain (Cterm) of human mt-leucyl tRNA synthetase rescues the pathologic phenotype associated either with the m.3243A>G mutation in mt-tRNA(Leu(UUR)) or with mutations in the mt-tRNA(Ile), both of which are aminoacylated by Class I mt-aminoacyl-tRNA synthetases (mt-aaRSs). Here we show, by using the human transmitochondrial cybrid model, that the Cterm is also able to improve the phenotype caused by the m.8344A>G mutation in mt-tRNA(Lys), aminoacylated by a Class II aaRS. Importantly, we demonstrate that the same rescuing ability is retained by two Cterm-derived short peptides, β30_31 and β32_33, which are effective towards both the m.8344A>G and the m.3243A>G mutations. Furthermore, we provide in vitro evidence that these peptides bind with high affinity wild-type and mutant human mt-tRNA(Leu(UUR)) and mt-tRNA(Lys), and stabilize mutant mt-tRNA(Leu(UUR)). In conclusion, we demonstrate that small Cterm-derived peptides can be effective tools to rescue cellular defects caused by mutations in a wide range of mt-tRNAs.

Aim Arrhythmogenic cardiomyopathy (ACM) is a genetic disorder mainly due to mutations in desmosomal genes, characterized by progressive fibro-adipose replacement of the myocardium, arrhythmias, and sudden death. It is still unclear which cell type is responsible for fibro-adipose substitution and which molecular mechanisms lead to this structural change. Cardiac mesenchymal stromal cells (C-MSC) are the most abundant cells in the heart, with propensity to differentiate into several cell types, including adipocytes, and their role in ACM is unknown. The aim of the present study was to investigate whether C-MSC contributed to excess adipocytes in patients with ACM. Methods and results We found that, in ACM patients' explanted heart sections, cells actively differentiating into adipocytes are of mesenchymal origin. Therefore, we isolated C-MSC from endomyocardial biopsies of ACM and from not affected by arrhythmogenic cardiomyopathy (NON-ACM) (control) patients. We found that both ACM and control C-MSC express desmosomal genes, with ACM C-MSC showing lower expression of plakophilin (PKP2) protein vs. controls. Arrhythmogenic cardiomyopathy C-MSC cultured in adipogenic medium accumulated more lipid droplets than controls. Accordingly, the expression of adipogenic genes was higher in ACM vs. NON-ACM C-MSC, while expression of cell cycle and anti-adipogenic genes was lower. Both lipid accumulation and transcription reprogramming were dependent on PKP2 deficiency. Conclusions Cardiac mesenchymal stromal cells contribute to the adipogenic substitution observed in ACM patients' hearts. Moreover, C-MSC from ACM patients recapitulate the features of ACM adipogenesis, representing a novel, scalable, patient-specific in vitro tool for future mechanistic studies.

e18506 Background: In head and neck squamous cell carcinoma (HNSCC) only a small subset of patients (pts) really benefits from immunotherapy, suggesting the need of validated molecular and immunological predictive biomarkers. The aim of the study was to evaluate both the genomic and immune profile of HNSCC correlating it to early progression to immunotherapy. Methods: This is a pilot study evaluating immune and molecular profile in platinum-refractory HNSCC pts treated with Nivolumab. Blood samples were collected at baseline (T0) and at second cycle of therapy (T1). The immune profile was studied by multiplex assay, evaluating circulating: CD137, CTLA4, PD1, PDL1, PDL2, CD27, TIM3, LAG3, GITR, HVEM, BTLA, CD80, CD28 and IDO. The comprehensive genomic profile was evaluated at baseline on formalin-fixed paraffin-embedded samples of the tumor through Foundation One Cdx test. The results obtained were correlated with early progression (within 3 months from the start of therapy). Results: Ten pts were enrolled in the study. Early progression occurred in 6 pts (60%). Median progression free survival and overall survival were 3 months (1-11) and 6.5 months (1-12), respectively. In all patients ≥ 5 genetic mutations were detected; median number of mutation was 6 (5-14); 8/10 pts had a tumor mutational burden less than 10 Muts/mb. The more frequent genetic alterations involved the cycline-dependent-kynase pathway (9/10), the transcription factor associated gene TP53 (8/10) and the PI3K-Akt-PTEN signaling pathways (5/10). Less frequent alterations involved FGFR family (4/10), NOTCH signaling pathway (3/10) and TERT (4/10). Early progression was slightly associated with the number of mutations detected (p = 0.06). The rising or falling trend of circulating levels of biomarkers were detected in 9/10 pts and included modification in IDO (1/10), LAG3 (3/10), GITR (3/10), BTLA (3/10), CD137 (4/10), CTLA4 (4/10), PD1 (2/10), Tim3 (3/10), CD28 (3/10), HVEM (3/10), CD80 (2/10). No modifications were recorded in PDL1, PDL2 and CD27. Conclusions: Our results, although in a small cohort, highlighted the complexity and heterogeneity of landscape in HNSCC pts. A novel evaluation that combines molecular and immune profile is needed. In the context of a poor prognosis disease the combination of personalized molecular and immune approaches could represent a promising strategy to improve survival.

Background: Gut microbiota seems to be implicated in the athero-thrombotic process. Its relationship with coronary thrombosis has never been investigated. Methods: Serum levels of bacterial lipopolysaccharides (LPS) against Escherichia Coli (EC) and EC DNA amplification by polymerase chain reaction, plasma sP-selectin, a marker of platelet activation, and zonulin, a marker of gut permeability, were measured in patients with ST-elevation myocardial infarction (STEMI) (n=50) or stable coronary disease (SA) (n=50) and controls (n=50). Serum LPS and sP-selectin were also measured in coronary thrombi and coronary blood of patients with STEMI and stable coronary disease, respectively. Immuno-histochemical analysis was performed in coronary thrombi of 12 STEMI patients. Finally, in vitro study was undertaken to evaluate if LPS stimulate leucocyte-platelet interaction. Results: Compared to controls, patients with SA and STEMI had higher systemic levels of LPS (p Conclusion: Bacterial LPS is detected in coronary thrombi of STEMI patients and may be implicated in the thrombotic process via leucocyte-mediated platelet activation.

We evaluated the impact of breast cancer subtypes on pathologic complete response (pCR) in 181 patients with positive nodes undergoing neoadjuvant chemotherapy (NAC). After NAC, patients underwent surgery, with sentinel lymph node biopsy (SLNB) or axillary dissection (ALND). In 28.2% of cases a pCR was achieved, with the highest rate in Her2+ and triple negative tumors. Overall, nodal pCR was more frequent than breast pCR (P = 0.003) with higher percentages in Her2+ and LLB-Her2+ (P < 0.05). In the Her2+ group, nodal pCR was observed only with breast pCR. Thus, in Her2+ tumors, breast pCR predicts node pCR, supporting the use of SLNB in this subgroup to stage the axilla avoiding ALND.

This clinical case describes a 78-year-old female patient who underwent total thyroidectomy and parathyroidectomy due to thyroid cancer; surgery was followed by radioiodine therapy. The patient was treated with 20 µg of teriparatide once a day for 7 years and consistently showed above normal blood calcium levels. In addition, following the operation, the patient began taking 50 000 units of vitamin D orally every month and 1 g of calcium per os a day. A painful eschar had appeared on the front of her left leg 3 months before; it had developed into a deep, painful ulcer. The patient did not present arterial or venous diseases. Arterial hypertension was effectively managed with 5 mg of ramipril per day, renal function was normal, and the patient was not suffering from diabetes. A computed tomography scan gave evidence of extensive dermo-hypodermitis calcification in the ulcer area. A biopsy was performed and the histological examination revealed calciphylaxis. The patient was treated by suspending teriparatide, which actually had been suspended months before, and replacing it with bisphosphonate; suspension of calcium and vitamin D and the oral administration of 250 units of sulodexide every 12 hours. Local therapy was adapted according to the advancement of the ulcer’s stages. This case represents one of non-uremic calciphylaxis determined by an overly lengthy administration of a parathyroid hormone that caused drug-induced hyperthyroidism over a long period of time. A revision of the worldwide literature on the topic was carried out and an etiopathogenetic hypothesis concerning the clinical case is put forward.

Autophagy is usually a pro-survival mechanism in cancer cells, especially in the course of chemotherapy, thus autophagy inhibition may enhance the chemotherapy-mediated anti-cancer effect. However, since autophagy is strongly involved in the immunogenicity of cell death by promoting ATP release, its inhibition may reduce the immune response against tumors, negatively influencing the overall outcome of chemotherapy. In this study, we evaluated the in vitro and in vivo anti-cancer effect of curcumin (CUR) against Her2/neu overexpressing breast cancer cells (TUBO) in the presence or in the absence of the autophagy inhibitor chloroquine (CQ). We found that TUBO cell death induced by CUR was increased in vitro by CQ and slightly in vivo in nude mice. Conversely, CQ counteracted the Cur cytotoxic effect in immune competent mice, as demonstrated by the lack of in vivo tumor regression and the reduction of overall mice survival as compared with CUR-treated mice. Immunohistochemistry analysis revealed the presence of a remarkable FoxP3 T cell infiltrate within the tumors in CUR/CQ treated mice and a reduction of T cytotoxic cells, as compared with single CUR treatment. These findings suggest that autophagy is important to elicit anti-tumor immune response and that autophagy inhibition by CQ reduces such response also by recruiting T regulatory (Treg) cells in the tumor microenvironment that may be pro-tumorigenic and might counteract CUR-mediated anti-cancer effects.

Background— Endomyocardial biopsy (EMB) has a low sensitivity. Electroanatomic voltage mapping (EVM) is effective in guiding EMB thanks to its ability in identifying and locating low-voltage regions. The analysis of unipolar EVM can correlate with epicardial pathological involvement. We evaluated the unipolar EVM in EMB areas to determine whether it can increase EMB sensitivity in diagnosing epicardial diseases. Methods and Results— We performed endocardial bipolar EVM-guided EMBs in 29 patients and we analyzed unipolar EVM at withdrawal sites. Eighty myocardial samples were collected (mean, 2.8±0.9; median, 3 fragments per patient) and 60 were suitable for histological analysis. Ten specimens (17%) were collected from an area with discordant normal bipolar/low-voltage unipolar EVM and they were diagnostic or suggestive for arrhythmogenic right ventricular dysplasia/cardiomyopathy in 6 patients, for myocarditis and sarcoidosis in 1 patient each. Six samples (10%) were collected from an area with discordant low-voltage bipolar/normal unipolar EVM and they showed nonspecific features. The sensitivity of unipolar EVMs for a diagnostic biopsy finding EMB was significantly higher compared with bipolar EVMs analyzed according to samples ( P P =0.008). The specificity of unipolar EMB was better than bipolar EMB when analyzed for all samples ( P =0.0014) but the difference did not reach statistical significance when analyzed by patient ( P =0.083). The diagnostic yield was 63.3% for the bipolar and 83.3% for the unipolar EVM. Conclusions— These findings suggest that use of a combined bipolar/unipolar map may be able to improve the diagnostic yield of endomyocardial ventricular biopsy.

Breast neoplasms are one of the leading causes of morbidity and mortality in women. Even if surgery is the treatment of choice, other forms of less invasive radical treatment are desirable. High-intensity focused ultrasound is already established as a valid non-invasive technique that ensures tumor ablation in various organs. The use of ultrasound or magnetic resonance guidance allows having some advantages such as the capability to treat tumors in moving organs or the possibility to have a real-time monitoring of the temperature increase. The aim of this paper is to report the use of high-intensity focused ultrasound technique with ultrasound and magnetic resonance guidance for the ablation of breast tumors, including both benign and malignant lesions.

Background Minor salivary gland (MSG) biopsy represents an useful tool not only for the diagnosis of primary Sjogren9s Syndrome (pSS) but also to evaluate patients prognosis. Recognition of germinal centers (GCs) by hematoxilin eosin (HE) and/or IHC staining for follicular dendritic cells (FDC) detection is mandatory, representing a risk factor for lymphoma development. Focus score (FS) is one of the main instrument to quantify MSG impairment, nonetheless quality information regarding the type of infiltrate such as the entity, structure and localization, are lacking. Objectives Aim of this study is to find any association of specific histological features of MSG from patients with pSS with the principal clinical and laboratory features. Moreover, to investigate the utility of histological parameters, other than FS or GCs, for characterizing patients. Methods Patients with pSS were enrolled in our SS clinic, and clinical/laboratory data (table) referring to the time of MSG biopsy, gathered on a dedicated database. MSG, removed for diagnostic purposes, were preserved as paraffin embedded tissue, then cut and sequentially stained by HE monoclonal mouse anti-CD20 (lymphocytes B); monoclonal mouse anti-CD21 (FDC)]. Images were collected by Zeiss Axio Scan and analysed (ZEN software) as follows: FS calculation, mean foci area, percentage of infiltration, presence of segregated foci (SF) (specifically, clear evidence of T and B cells area by CD3-CD20 double staining), GCs and lymphoepithelial lesions (LELs) detection. Results 53 MSG from patients with pSS were collected and analysed. Patients clinical and laboratory data are reported in table. FS positively correlated with the percentage of infiltration (p Conclusions The FS was associated with the presence of GCs and LELs, as well as with more organized infiltrates characterized by segregation in T and B areas (SF), thus representing an useful tool which mirrors the risk of lymphoma. From our study, the qualitative characteristics of the biopsy, including SF, percentage of infiltration or the mean foci area, appear to be strictly linked. Moreover, their association with the presence of GCs and LELs supports the importance to consider also these features during histological examination. The lack of correlation between histological parameters and clinical/laboratory features might reveal a weaker connection between histological findings and specific SS phenotypes except for the relationship between glandular swelling and GCs which confirms how this clinical aspect should be considered as a risk factor for lymphoma development. Disclosure of Interest None declared

Loss-of-function mutations in TTC19 (tetra-tricopeptide repeat domain 19) have been associated with severe neurological phenotypes and mitochondrial respiratory chain complex III deficiency. We previously demonstrated the mitochondrial localization of TTC19 and its link with complex III biogenesis. Here we provide detailed insight into the mechanistic role of TTC19, by investigating a Ttc19?/? mouse model that shows progressive neurological and metabolic decline, decreased complex III activity, and increased production of reactive oxygen species. By using both the Ttc19?/? mouse model and a range of human cell lines, we demonstrate that TTC19 binds to the fully assembled complex III dimer, i.e., after the incorporation of the iron-sulfur Rieske protein (UQCRFS1). The in situ maturation of UQCRFS1 produces N-terminal polypeptides, which remain bound to holocomplex III. We show that, in normal conditions, these UQCRFS1 fragments are rapidly removed, but when TTC19 is absent they accumulate within complex III, causing its structural and functional impairment.

Background Symptoms and signs of myocardial ischemia in the absence of obstructive coronary disease are common in hypertensive patients. This can be explained by coronary microvascular dysfunction (CMD) due to adverse remodeling of coronary arterioles which have also been reported in the spontaneously hypertensive rat (SHR). Objective The aim of this study was to compare the effect of Ramipril, Perindopril, Candesartan, Atenolol, Amlodipine, Indapamide and HMR1766 on coronary microvascular dysfunction (CMD) in the spontaneously hypertensive rat (SHR). Methods Eight groups of 24 week old SHR were treated for 8 weeks. Blood pressure (BP) was measured invasively at the end of treatment. After sacrifice, hearts were mounted on a Langendorff apparatus for measurement of hyperemic coronary flow (CF). Hearts were then processed for histomorphometric analysis. Results All compounds, except HMR1766, induced a significant reduction of BP. Perindopril and Candesartan increased hyperemic CF whereas the other compounds had no significant effect. Perindopril, Ramipril, Atenolol, Indapamide and HMR1766 induced significant reverse arteriolar remodeling whereas Candesartan and Amlodipine did not. Conclusions The effect of anti-hypertensive treatment on CMD is not only dependent on BP reduction. Compounds with comparable anti-hypertensive efficacy may exert different effects on CF and induce different degrees of reverse arteriolar remodeling. This article is protected by copyright. All rights reserved.

Triple negative breast cancer (TNBC) has an aggressive clinical behaviour, with a poorer prognosis compared to other subtypes. Recently, tumor-infiltrating lymphocytes (TILs) have been proposed as a predictive biomarker for a better clinical outcome and pathological response (pR) after neoadjuvant chemotherapy (NACT) in TNBC. These data confirm the role of the immune system in the neoplastic progression and in the response to therapy. We performed a retrospective analysis of 54 pre-NACT biopsies of TNBC and compared both the percentage of stromal TILs and the degree of PD-L1 expression with the extent of pR to standard NACT. A pathological complete response (pCR) was achieved in 35% of cases. Univariate analysis showed (i) a significant association between PD-L1 expression in ≥25% of neoplastic cells and the achievement of a pCR (p=0.024); (ii) a significantly higher frequency of pCR in cases showing ≥50% stromal TILs (p<0.001). However in the multivariate analysis only PD-L1 expression on tumor cells remained significantly associated with pCR (OR = 1,13; 95% CI 1,01–1,27), suggesting that the expression of this biomarker could be associated with a subpopulation of TNBC more likely to respond to chemotherapy. These data need to be confirmed by larger studies.

Amphetamine congeners can be prescribed as anorexiant drugs despite their potential adverse effects, including cardiac toxicity. However, the morphologic features of cardiac damage related to protracted use of these compounds are unknown. We provide a detailed description of cardiac autopsy findings in 3 cases of sudden death associated with protracted use of high doses of phendimetrazine and/or phenylpropanolamine or bupropion prescribed as anorexiants, in association with other compounds. The main cardiac findings were similar in all 3 cases: (1) mild-moderate hypertrophy of the left ventricle and/or the septum; (2) myocardial nonischemic scarring (midmural and/or subepicardial) appearing as discrete foci or with a bandlike morphology; (3) mild-moderate intramural small vessel disease in the absence of significant epicardial coronary artery stenosis; and (4) acute/recent inflammatory lesions consistent with toxic myocarditis. In summary, the detailed pathology examination of the heart in these 3 cases revealed myocardial lesions identical to those reported in catecholamine myocardial damage in all their various stages of evolution. In the presence of a clinical history of long-term intake of anorexiants of this category, it is most important at autopsy to recognize and correctly interpret the acute and chronic myocardial lesions of the type herein described because they represent an anatomical substrate for arrhythmic death.

Pantothenate kinase-associated neurodegeneration, caused by mutations in the PANK2 gene, is an autosomal recessive disorder characterized by dystonia, dysarthria, rigidity, pigmentary retinal degeneration and brain iron accumulation. PANK2 encodes the mitochondrial enzyme pantothenate kinase type 2, responsible for the phosphorylation of pantothenate or vitamin B5 in the biosynthesis of co-enzyme A. A Pank2 knockout (Pank2(-/-)) mouse model did not recapitulate the human disease but showed azoospermia and mitochondrial dysfunctions. We challenged this mouse model with a low glucose and high lipid content diet (ketogenic diet) to stimulate lipid use by mitochondrial beta-oxidation. In the presence of a shortage of co-enzyme A, this diet could evoke a general impairment of bioenergetic metabolism. Only Pank2(-/-) mice fed with a ketogenic diet developed a pantothenate kinase-associated neurodegeneration-like syndrome characterized by severe motor dysfunction, neurodegeneration and severely altered mitochondria in the central and peripheral nervous systems. These mice also showed structural alteration of muscle morphology, which was comparable with that observed in a patient with pantothenate kinase-associated neurodegeneration. We here demonstrate that pantethine administration can prevent the onset of the neuromuscular phenotype in mice suggesting the possibility of experimental treatment in patients with pantothenate kinase-associated neurodegeneration.

The molecular mechanisms responsible for the elevated metastatic potential of malignant melanoma are still not fully understood. In order to shed light on the molecules involved in the acquisition by melanoma of a highly aggressive phenotype, we compared the gene expression profiles of two cell clones derived from the human cutaneous metastatic melanoma cell line M14: a highly invasive clone (M14C2/MK18) and a clone (M14C2/C4) with low ability to invade the extracellular matrix (ECM). The highly invasive phenotype of M14C2/MK18 cells was correlated with overexpression of neuropilin-1, activation of a vascular endothelial growth factor (VEGF)-A/VEGFR-2 autocrine loop and secretion of matrix metalloprotease-2. Moreover, in an in vivo murine model, M14C2/MK18 cells displayed a higher growth rate as compared with M14C2/C4 cells, even though in vitro both clones possessed comparable proliferative potential. Microarray analysis in M14C2/MK18 cells showed a strong upregulation of platelet-derived growth factor (PDGF)-C, a cytokine that contributes to angiogenesis, and downregulation of calpain-3, a calcium-dependent thiol-protease that regulates specific signalling cascade components. Inhibition of PDGF-C with a specific antibody resulted in a significant decrease in ECM invasion by M14C2/MK18 cells, confirming the involvement of PDGF-C in melanoma cell invasiveness. Moreover, the PDGF-C transcript was found to be upregulated in a high percentage of human melanoma cell lines (17/20), whereas only low PDGF-C levels were detected in a few melanocytic cultures (2/6). By contrast, inhibition of calpain-3 activity in M14C2/C4 control cells, using a specific chemical inhibitor, markedly increased ECM invasion, strongly suggesting that downregulation of calpain-3 plays a role in the acquisition of a highly invasive phenotype. The results indicate that PDGF-C upregulation and calpain-3 downregulation are involved in the aggressiveness of malignant melanoma and suggest that modulators of these proteins or their downstream effectors may synergise with VEGF‑A therapies in combating tumour-associated angiogenesis and melanoma spread.

Surgical aortic specimens are usually examined in Pathology Departments as a result of treatment of aneurysms or dissections. A number of diseases, genetic syndromes (Marfan syndrome, Loeys–Dietz syndrome, etc.), and vasculopathic aging processes involved in vascular injury can cause both distinct and nonspecific histopathologic changes with degeneration of the media as a common denominator. Terminology for these changes has varied over time leading to confusion and inconsistencies. This consensus document has established a revised, unified nomenclature for the variety of noninflammatory degenerative aortic histopathologies seen in such specimens. Older terms such as cystic medial necrosis and medionecrosis are replaced by more technically accurate terms such as mucoid extracellular matrix accumulation (MEMA), elastic fiber fragmentation and/or loss, and smooth muscle cell nuclei loss. A straightforward system of grading is presented to gauge the extent of medial degeneration and synoptic reporting tables are provided. Herein we present a standardized nomenclature that is accessible to general pathologists and useful for future publications describing these entities.

Nonischemic left ventricular scar (NLVS) is a pattern of myocardial injury characterized by midventricular and/or subepicardial gadolinium hyperenhancement at cardiac magnetic resonance, in absence of significant coronary artery disease. We aimed to evaluate the prevalence of NLVS in juvenile sudden cardiac death and to ascertain its etiology at autopsy. We examined 281 consecutive cases of sudden death of subjects aged 1 to 35 years. NLVS was defined as a thin, gray rim of subepicardial and/or midmyocardial scar in the left ventricular free wall and/or the septum, in absence of significant stenosis of coronary arteries. NLVS was the most frequent finding (25%) in sudden deaths occurring during sports. Myocardial scar was localized most frequently within the left ventricular posterior wall and affected the subepicardial myocardium, often extending to the midventricular layer. On histology, it consisted of fibrous or fibroadipose tissue. Right ventricular involvement was always present. Patchy lymphocytic infiltrates were frequent. Genetic and molecular analyses clarified the etiology of NLVS in a subset of cases. Electrocardiographic (ECG) recordings were available in more than half of subjects. The most frequent abnormality was the presence of low QRS voltages (

Background Danon disease (DD) is a rare disorder characterized by cardiomyopathy, intellectual disability, and proximal myopathy. It is caused by mutations in the LAMP2 gene on X chromosome. Female patients most often present with late-onset cardiomyopathy and slow disease progression, but early-onset cases with unfavorable prognosis have been reported. Case report We describe the clinical, pathological, and molecular features of a novel LAMP2 c.453delT mutation in a female patient with severe hypertrophic cardiomyopathy, Wolff Parkinson White (WPW) syndrome and rapid progression to heart failure, requiring heart transplant. Immunohistochemical analysis of LAMP2 in the explanted heart revealed a mosaic pattern of distribution, with discrete clusters of either stained or unstained cardiac myocytes, the latter being more frequent in the septum. These findings paralleled X chromosome inactivation within the myocardium. Interestingly, multiple foci of microscarring were found on histology in the Left Ventricle (LV) free wall and septum, in a close spatial relationship with remodeling and severe stenosis of intramural coronary arterioles. Conclusions Our findings suggest that several features may contribute to the early and severe cardiac phenotype in female DD patients. The type of mutation may account for the early disease onset, while both the inhomogeneous distribution of LAMP2 loss and the presence of microvascular remodeling may be determinant in the rapid progression to heart failure.

The term idiopathic noncirrhotic portal hypertension (INCPH) has been recently proposed to replace terms, such as hepatoportal sclerosis, idiopathic portal hypertension, incomplete septal cirrhosis, and nodular regenerative hyperplasia, used to describe patients with a hepatic presinusoidal cause of portal hypertension of unknown etiology, characterized by features of portal hypertension (esophageal varices, nonmalignant ascites, porto-venous collaterals), splenomegaly, patent portal, and hepatic veins and no clinical and histological signs of cirrhosis. Physicians should learn to look for this condition in a number of clinical settings, including cryptogenic cirrhosis, a disease known to be associated with INCPH, drug administration, and even chronic alterations in liver function tests. Once INCPH is clinically suspected, liver histology becomes mandatory for the correct diagnosis. However, pathologists should be familiar with the histological features of INCPH, especially in cases in which histology is not only requested to exclude liver cirrhosis.

Background The antimalarial drug Pyrimethamine has been suggested to exert an antitumor activity by inducing apoptotic cell death in cancer cells, including metastatic melanoma cells. However, the dose of Pyrimethamine to be considered as an anticancer agent appears to be significantly higher than the maximum dose used as an antiprotozoal drug. Methods Hence, a series of Pyrimethamine analogs has been synthesized and screened for their apoptosis induction in two cultured metastatic melanoma cell lines. One of these analogs, the Methylbenzoprim, was further analyzed to evaluate cell-cycle and the mechanisms of cell death. The effects of Methylbenzoprim were also analyzed in a severe combined immunodeficiency (SCID)-mouse xenotransplantation model. Results Low dose of Methylbenzoprim was capable of inducing cytotoxic activity and a potent growth-inhibitory effect by arresting cell cycle in S-phase in melanoma cells. Methylbenzoprim was also detected as powerful antineoplastic agents in SCID-mouse although used at very low dose and as a single agent. Conclusions Our screening approach led to the identification of a “low cost” newly synthesized drug (methylbenzoprim), which is able to act as an antineoplastic agent in vitro and in vivo, inhibiting melanoma tumor growth at very low concentrations. Electronic supplementary material The online version of this article (doi:10.1186/s13046-016-0409-9) contains supplementary material, which is available to authorized users.

We here report on a sudden cardiac death during athletic competition in Italy, a country where annual pre-participation screening for sport activity is mandatory by law and proven to be life-saving. The case offers an opportunity to discuss current limitations of primary prevention through the early identification of patients at risk, and the persisting need of implementing secondary prevention measures by a proper treatment of cardiac arrest in sport arenas. A top-level football athlete, with regular annual pre-participation screening according to the Italian law, died suddenly during a football match of the Italian premier league. At 29′42′′ of the first half-time of the football match, while running in the football course, he fell down a first time on his knees, tried to get-up, and, after 5 s, collapsed again in a prone position. After 12 s, the team physician promptly intervened and found him unresponsive and unconscious. After 27 s a Red-Cross volunteer brought a semi-automated external defibrillator (sAED) to the resuscitation scene.While external cardiacmassagewas performed, the sAEDwasnever used. At 2′47′′ the athletewas put on a stretcher and rapidly transferred to an ambulance to the emergency room of the nearby hospital, where he arrived, with ongoing uninterrupted cardiopulmonary resuscitation (CPR) manoeuvres, 10 min after onset of the cardiac arrest. At the arrival, the ECG monitor documented ventricular fibrillation (VF). Despite multiple DC shocks, the athlete died.

Mitochondrial disease refers to a heterogenous group of genetic disorders that result from dysfunction of the final common pathway of energy metabolism. Mitochondrial DNA mutations affect key components of the respiratory chain and account for the majority of mitochondrial disease in adults. Owing to critical dependence of the heart on oxidative metabolism, cardiac involvement in mitochondrial disease is common and may occur as the principal clinical manifestation or part of multisystem disease. Recent advances in our understanding of the clinical spectrum and genetic aetiology of cardiac involvement in mitochondrial DNA disease have important implications for cardiologists in terms of the investigation and multi-disciplinary management of patients.

Neurodegeneration with brain iron accumulation (NBIA) comprises a group of neurodegenerative disorders characterized by high brain content of iron and presence of axonal spheroids. Mutations in the PANK2 gene, which encodes pantothenate kinase 2, underlie an autosomal recessive inborn error of coenzyme A metabolism, called pantothenate kinase-associated neurodegeneration (PKAN). PKAN is characterized by dystonia, dysarthria, rigidity and pigmentary retinal degeneration. The pathogenesis of this disorder is poorly understood and, although PANK2 is a mitochondrial protein, perturbations in mitochondrial bioenergetics have not been reported. A knock-out (KO) mouse model of PKAN exhibits retinal degeneration and azoospermia, but lacks any neurological phenotype. The absence of a clinical phenotype has partially been explained by the different cellular localization of the human and murine PANK2 proteins. Here we demonstrate that the mouse Pank2 protein localizes to mitochondria, similar to its human orthologue. Moreover, we show that Pank2-defective neurons derived from KO mice have an altered mitochondrial membrane potential, a defect further corroborated by the observations of swollen mitochondria at the ultra-structural level and by the presence of defective respiration. © The Author 2012.Published by Oxford University Press.

Mitochondrial dysfunction and altered proteostasis are central features of neurodegenerative diseases. The pitrilysin metallopeptidase 1 (PITRM1) is a mitochondrial matrix enzyme, which digests oligopeptides, including the mitochondrial targeting sequences that are cleaved from proteins imported across the inner mitochondrial membrane and the mitochondrial fraction of amyloid beta (Aβ). We identified two siblings carrying a homozygous PITRM1 missense mutation (c.548G>A, p.Arg183Gln) associated with an autosomal recessive, slowly progressive syndrome characterised by mental retardation, spinocerebellar ataxia, cognitive decline and psychosis. The pathogenicity of the mutation was tested in vitro , in mutant fibroblasts and skeletal muscle, and in a yeast model. A Pitrm1 +/− heterozygous mouse showed progressive ataxia associated with brain degenerative lesions, including accumulation of Aβ‐positive amyloid deposits. Our results show that PITRM1 is responsible for significant Aβ degradation and that impairment of its activity results in Aβ accumulation, thus providing a mechanistic demonstration of the mitochondrial involvement in amyloidotic neurodegeneration.

Patients and animal models of arterial hypertension are characterized by structural and functional abnormalities of the coronary microcirculation. Using a translational approach, we ascertained whether antihypertensive treatment can reverse microvascular remodelling and improve myocardial perfusion.In 20 hypertensive patients with left ventricular hypertrophy, blood pressure, left ventricular mass index and myocardial blood flow were measured at baseline and after 6 months of treatment with perindopril + indapamide. In spontaneously hypertensive rats, blood pressure, coronary flow and histomorphometry of intramural coronary arterioles were measured after 8 weeks of treatment with placebo or perindopril + indapamide.In patients, treatment decreased blood pressure (161 ± 10/96 ± 5 to 136 ± 12/81 ± 6 mmHg; P0.0001) and left ventricular mass index (93 ± 16 to 85 ± 17 g/m; P0.01) while increasing baseline (0.69 ± 0.13 to 0.88 ± 0.36 ml/min per g; P0.05) and hyperaemic myocardial blood flow (1.42 ± 0.32 to 1.94 ± 0.99 ml/min per g; P0.05). In rats treated with perindopril + indapamide (n = 11), blood pressure was 93 ± 18/55 ± 18 mmHg compared to 215 ± 18/161 ± 17 mmHg in placebo (n = 6; P0.001), baseline flow was unchanged whilst hyperaemic coronary flow was 19.89 ± 3.50 vs. 12.15 ± 0.99 ml/min per g, respectively (P0.01). The medial area of intramural arterioles was 1613 ± 409 with perindopril + indapamide and 8118 ± 901 μm with placebo (P0.001).In patients with arterial hypertension and left ventricular hypertrophy, perindopril + indapamide reduced blood pressure and left ventricular mass index and improved resting and hyperaemic myocardial blood flow. Data in rats provide evidence that the improvement in coronary flow observed after treatment is due to reverse remodelling of intramural coronary arterioles and improved microvascular function.

Background: Immune checkpoint inhibitors have revolutionized treatment and outcome of severe and often fatal disease, as metastatic lung cancer, demonstrating long-term tumor control and extended patient survival. Unfortunately, only 25-30% of patients have a long-term benefit from immunotherapy, while the remaining 70-75% demonstrate primary or acquired resistance. Recently, indoleamine 2,3-dioxygenase (IDO) has been proposed as a possible mechanism of resistance to anti-PD1 treatment. Indeed, IDO catalyzes the degradation of tryptophan (Trp) into kynurenine (Kyn), which seems to enhance the activity of Treg, leading to an immunosuppressive microenvironment. Methods: The serum concentrations of Trp and Kyn were measured by high-performance liquid chromatography tandem mass spectrometry in 26 patients affected by non-small cell lung cancer (NCLSC) before the start of the second-line therapy with nivolumab. The IDO activity was expressed with Kyn/Trp ratio. The associations between Kyn/Trip ratio and early progression, PS, age, sex, brain metastases and pleural effusion were analyzed using Spearman test and Mann Whitney test. Results: 14 out of 26 patients (54%) presented early progression (defined as progression of the disease within 6 months from the beginning of nivolumab treatment). The median value of Kyn/Trp ratio was 0.073 (0.024-0.18). We found a significant association between Kyn/Trp ratio and early progression (p= 0.009), while no statistical associations were found between Kyn/Trp ratio and PS, age, sex, brain metastases and pleural effusion. Indeed, patients with early progressive disease presented a median value of Kyn/Trp ratio significantly higher than other patients (0.094 vs 0.052; p= 0.01). Conclusion: The pretreatment evaluation of IDO activity, expressed as Kyn/Trp ratio, seems to be associated with response to immunotherapy. In particular, higher Kyn/Trp ratio could predict resistance to anti-PD-1 treatment. These preliminary results suggest the possibility of using anti-PD-1 plus IDO inhibitor in patients with high level of Kyn/Trp ratio. Citation Format: Andrea Botticelli, Bruna Cerbelli, Luana Lionetto, Ilaria Zizzari, Annalina Pisano, Michela Roberto, Elisa Onesti, Francesca Romana DI Pietro, Chiara Napoletano, Laura Pizzuti, Patrizia Vici, Giulia D'Amati, Federica Mazzuca, Maurizio Simmaco, Marianna Nuti, Paolo Marchetti. The key role of kynurenine in anti-PD-1 failure [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5705.

e15021Background: Immune checkpoint inhibitors have revolutionized treatment and outcome of NSCLC. Unfortunately, only 25-30% of patients have a long-term benefit, while the remaining 70-75% demons...

UNLABELLED Voltage Mapping-Guided Biopsy in ARVC/D. INTRODUCTION To improve the endomyocardial biopsy (EMB) diagnostic sensitivity for arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D), we hypothesized a biopsy sampling focused on selected right ventricle (RV) low-voltage areas identified by electroanatomic voltage mapping. METHODS AND RESULTS The study population (22 patients, 10 men; mean age 34 +/- 10 years) included 11 patients with overt ARVC/D (group A) and 11 patients with suspected ARVC/D (group B), according to both arrhythmic profile and standardized noninvasive diagnostic criteria. In all 22 patients, an RV bipolar voltage mapping was performed with CARTO system sampling multiple endocardial sites (262 +/- 61), during sinus rhythm, with a 0.5-1.5 mV color range setting of voltage display. All 11 (100%) group A patients and 8 of the 11 (73%) group B patients (P = nonsignificant [NS]) presented RV low-voltage areas (

Objectives The purpose of this study was to clarify the molecular mechanisms linking human mitochondrial deoxyribonucleic acid (mtDNA) dysfunction to cardiac remodeling. Background Defects of the mitochondrial genome cause a heterogeneous group of clinical disorders, including mitochondrial cardiomyopathies (MIC). The molecular events linking mtDNA defects to cardiac remodeling are unknown. Energy derangements and increase of mitochondrial-derived reactive oxygen species (ROS) could both play a role in the development of cardiac dysfunction in MIC. In addition, mitochondrial proliferation could interfere with sarcomere alignment and contraction. Methods We performed a detailed morphologic and molecular analysis on failing hearts from 3 patients with MIC, failing human hearts due to ischemic heart disease (IHD) or dilated cardiomyopathies (DCM), and nonfailing hearts. Results The MIC hearts showed marked mitochondrial proliferation with myofibril displacement. Consistent with morphologic features, increase in mtDNA content per cell and induction of genes involved in mitochondrial biogenesis, fatty acid metabolism, and glucose transport were observed. Down-regulation of these genes characterized DCM and IHD hearts. A pronounced increase in mitochondrial-derived ROS was observed in MIC hearts compared with failing hearts due to other causes. This was paralleled by up-regulation of genes encoding for uncoupling proteins and antioxidant enzymes. However, there was not a significant increase in antioxidant enzyme activity. Conclusions Our results suggest that besides energy deficiency, mitochondrial biogenesis per se is a maladaptive response in MIC and, possibly, in other metabolic cardiomyopathies.

Studies aimed at elucidating the pathogenetic mechanisms underlying the initiation and progression of human atherosclerosis have emphasized the central role of inflammatory and immune cells. Atherosclerotic plaques are infiltrated by activated macrophages, T and B lymphocytes, plasma cells, and mast cells, releasing inflammatory molecules, which amplify the severity of the disease. Endothelial cells subjected to various stress conditions express increased amounts of heat shock proteins (HSPs), some of the most successfully conserved proteins throughout evolution. Many experimental observations reviewed in this article draw attention to several HSPs targeted by a specific cellular and humoral immune response in patients with atherosclerotic disease. The review also reports preliminary data obtained by our group on the possible role of HSP90 as a candidate autoantigen in carotid atherosclerosis. Our study deals with the presence of specific antibodies and T cells directed against HSP90 in patients with carotid atherosclerotic plaques. In 60% of these subjects' sera but in none of the sera from healthy controls immunoblotting (IB) detected the presence of specific antibodies. Moreover, 20% of peripheral blood mononuclear cells (PBMC) samples from patients but none from healthy subjects proliferated in response to human purified HSP90. In vitro experiments showed an upregulation of HSP90 expression in endothelial cells exposed to oxidative stress by treatment with H(2)O(2) and greater release of soluble HSP90 in culture supernatants from H(2)O(2)-treated cells than from untreated cells.

This report describes the case of two siblings who underwent heart transplantation with a clinical diagnosis of mildly dilated cardiomyopathy. Pathological examination of the hearts revealed arrhythmogenic (right ventricular) cardiomyopathy, adipose type, associated with biventricular myocarditis in both the recipients' hearts. Family history revealed the occurrence of dilated cardiomyopathy and myocarditis in their father and his sister. To our knowledge, this is the first pathological demonstration of arrhythmogenic cardiomyopathy and myocarditis in siblings. We think this report substantiates a genetic ground for the relationship between these two heart conditions.

Mutations in mitochondrial (mt) DNA determine important human diseases. The majority of the known pathogenic mutations are located in transfer RNA (tRNA) genes and are responsible for a wide range of currently untreatable disorders. Experimental evidence both in yeast and in human cells has shown that the detrimental effects of mt-tRNA point mutations can be attenuated by increasing the expression of the cognate mt-aminoacyl-tRNA synthetases (aaRSs). In addition, constitutive high levels of isoleucyl-tRNA syntethase have been shown to reduce the penetrance of a homoplasmic mutation in mt-tRNA(Ile) in a small kindred. More recently, we showed that the isolated carboxy-terminal domain of human mt-leucyl tRNA synthetase (LeuRS-Cterm) localizes to mitochondria and ameliorates the energetic defect in transmitochondrial cybrids carrying mutations either in the cognate mt-tRNA(Leu(UUR)) or in the non-cognate mt-tRNA(Ile) gene. Since the mt-LeuRS-Cterm does not possess catalytic activity, its rescuing ability is most likely mediated by a chaperon-like effect, consisting in the stabilization of the tRNA structure altered by the mutation. All together, these observations open potential therapeutic options for mt-tRNA mutations-associated diseases.

Inflammatory diseases of the aorta include routine atherosclerosis, aortitis, periaortitis, and atherosclerosis with excessive inflammatory responses, such as inflammatory atherosclerotic aneurysms. The nomenclature and histologic features of these disorders are reviewed and discussed. In addition, diagnostic criteria are provided to distinguish between these disorders in surgical pathology specimens. An initial classification scheme is provided for aortitis and periaortitis based on the pattern of the inflammatory infiltrate: granulomatous/giant cell pattern, lymphoplasmacytic pattern, mixed inflammatory pattern, and the suppurative pattern. These inflammatory patterns are discussed in relation to specific systemic diseases including giant cell arteritis, Takayasu arteritis, granulomatosis with polyangiitis (Wegener's), rheumatoid arthritis, sarcoidosis, ankylosing spondylitis, Cogan syndrome, Behcet's disease, relapsing polychondritis, syphilitic aortitis, and bacterial and fungal infections. (C) 2015 Elsevier Inc. All rights reserved

Mutations in cardiac sarcomere protein genes are associated with a variety of clinical phenotypes, including hypertrophic (HCM), dilated (DCM), and restrictive (RCM) cardiomyopathy as well as left ventricular non-compaction, with the overlap of morpho-functional manifestations in individual patients and families. Over time, initial phenotypes may undergo profound changes which determine clinical course and disease progression. Although genetic defects causing HCM and DCM have opposite effects at the myofilament level, a number of downstream maladaptive mechanisms, ranging from microvascular dysfunction and ischaemia to myocardial fibrosis and from diastolic dysfunction to abnormal sympathetic activation and arrhythmogenesis, seem to recur in sarcomeric cardiomyopathies, independent of the presenting phenotype. The extent and rate at which each of these features occur and evolve may be radically different in each form of cardiomyopathy, determining a clinical heterogeneity that is not only cross-sectional, but also longitudinal, i.e. time-related. Timely and sensitive detection of these long-term modifications in the clinical setting is a key to preventing advanced disease and identifying novel therapeutic targets. The present review evaluates the contribution of contemporary technology to pre-clinical diagnosis, characterization of phenotypes, and assessment of disease progression in sarcomere cardiomyopathies, including echocardiography, positron emission tomography, magnetic resonance, pathology, and circulating biomarkers.

none 14 si Leber's hereditary optic neuropathy (LHON) is a maternally inherited blinding disease characterized by degeneration of retinal ganglion cells (RGCs) and consequent optic nerve atrophy. Peculiar features of LHON are incomplete penetrance and gender bias, with a marked male prevalence. Based on the different hormonal metabolism between genders, we proposed that estrogens play a protective role in females and showed that these hormones ameliorate mitochondrial dysfunction in LHON through the estrogen receptors (ERs). We also showed that ERβ localize to the mitochondria of RGCs. Thus, targeting ERβ may become a therapeutic strategy for LHON specifically aimed at avoiding or delaying the onset of disease in mutation carriers. Here, we tested the effects of ERβ targeting on LHON mitochondrial defective metabolism by treating LHON cybrid cells carrying the m.11778G>A mutation with a combination of natural estrogen-like compounds that bind ERβ with high selectivity. We demonstrated that these molecules improve cell viability by reducing apoptosis, inducing mitochondrial biogenesis and strongly reducing the levels of reactive oxygen species in LHON cells. These effects were abolished in cells with ERβ knockdown by silencing receptor expression or by using specific receptor antagonists. Our observations support the hypothesis that estrogen-like molecules may be useful in LHON prophylactic therapy. This is particularly important for lifelong disease prevention in unaffected LHON mutation carriers. Current strategies attempting to combat degeneration of RGCs during the acute phase of LHON have not been very effective. Implementing a different and preemptive approach with a low risk profile may be very helpful. none Pisano, Annalinda; Preziuso, Carmela; Iommarini, Luisa; Perli, Elena; Grazioli, Paola; Campese, Antonio F; Maresca, Alessandra; Montopoli, Monica; Masuelli, Laura; Sadun, Alfredo A; D'Amati, Giulia; Carelli, Valerio; Ghelli, Anna; Giordano, Carla Pisano, Annalinda; Preziuso, Carmela; Iommarini, Luisa; Perli, Elena; Grazioli, Paola; Campese, Antonio F; Maresca, Alessandra; Montopoli, Monica; Masuelli, Laura; Sadun, Alfredo A; D'Amati, Giulia; Carelli, Valerio; Ghelli, Anna; Giordano, Carla

The aim of this study was to assess whether lonidamine (LND) interferes with some steps in angiogenesis progression. We report here, for the first time, that LND inhibited angiogenic-related endothelial cell functions in a dose-dependent manner (1-50 microg/ml). In particular, LND decreased proliferation, migration, invasion, and morphogenesis on matrigel of different endothelial cell lines. Zymographic and Western blot analysis assays showed that LND treatment produced a reduction in the secretion of matrix metalloproteinase-2 and metalloproteinase-9 by endothelial cells. Vessel formation in a matrigel plug was also reduced by LND. The viability, migration, invasion, and matrix metalloproteinase production of different tumor cell lines were not affected by low doses of LND (1-10 microg/ml), whereas 50 microg/ml LND, which corresponds to the dose used in clinical management of tumors, triggered apoptosis both in endothelial and tumor cells. Together, these data demonstrate that LND is a compound that interferes with endothelial cell functions, both at low and high doses. Thus, the effect of LND on endothelial cell functions, previously undescribed, may be a significant contributor to the antitumor effect of LND observed for clinical management of solid tumors.

Frequent mutations of coding nucleotide repeats are thought to contribute significantly to carcinogenesis associated with microsatellite instability (MSI). We have shown that shortening of the poly(T)11 within the polypyrimidine stretch/accessory splicing signal of human MRE11 leads to the reduced expression and functional impairment of the MRE11/NBS1/RAD50 complex. This mutation was selectively found in mismatch repair (MMR) defective cell lines and potentially identifies MRE11 as a novel target for MSI. Here, we examined 70 microsatellite unstable primary human cancers and we report that MRE11 mutations occur in 83.7 and 50% of the colorectal and endometrial cancers, respectively. In the colorectal cancer series, mutated MRE11 is more frequently associated with advanced age at diagnosis and A/B stages. Biallelic mutations were present in 38.8% of the cases and more frequently associated with lower (G1/G2) grade tumors. Impaired MRE11 expression was prevalent in primary colorectal tumors with larger and biallelic shortening of the poly(T)11. Immunohistochemistry confirmed the impaired MRE11 expression and revealed NBS1-defective expression in MRE11 mutated cancers. Together with the observation that perturbation of the MRE11/NBS1/RAD50 complex predisposes to cancer, our work highlights MRE11 as a new common target in the MMR deficient tumorigenesis and suggests its role in colorectal carcinogenesis.

A novel 4949-base pair mitochondrial DNA (mtDNA) deletion was detected in various tissues in a postmortem study of a patient with Kearns-Sayre syndrome (KSS). Deleted mtDNA levels were higher in skeletal muscle and brain and lower in kidney, working myocardium, and endocrine tissues (thyroid, parathyroids, pancreas, and adrenal glands). The distribution of the deletion in skeletal muscle and conducting myocardium was analyzed by means of laser capture microdissection (LCM). In skeletal muscle, the abundance of deleted mtDNA was slightly higher in cytochrome c oxidase (COX)-negative fibers (70%) than in COX-positive fibers (64%), whereas in the conducting myocardium it was lower in the atrioventricular node (9%) than in the sinus node and bundle of His (30% and 32%, respectively). In this study, LCM proved to be a reliable technique for a more accurate assessment of genotype/phenotype correlation when investigating mtDNA-related disorders.

An infant with normal facies and none of the extracardiac anomalies usually associated with Williams syndrome presented at birth with an echocardiographic pattern of supravalvular pulmonary stenosis and displastic pulmonary valve. A clinical reappraisal was planned at 3 months of age, but the girl died suddenly at home at 2 months of age. At autopsy, both ventricles were hypertrophic, and the valves showed mild dysplasia. The walls of the great arteries were thick, with a “washed leather” consistency, but there was no gross evidence of discrete stenosis. The histologic mosaic appearance of the media of the great arteries, due to elastosis and extreme disarray of the elastic lamellae, prompted a postmortem diagnosis of supravalvar aortic stenosis and suggested a diagnosis of Williams syndrome, which was subsequently confirmed by fluorescence in situ hybridization. Pediatricians and pathologists should be alerted that Williams syndrome in the newborn may present as an isolated supravalvular pulmonary stenosis, whereas supravalvular aortic stenosis becomes clinically significant only a few months later.

ObjectivesThe purpose of this study was to understand the clinical and molecular features of familial hypertrophic cardiomyopathy (HCM) in which a mitochondrial abnormality was strongly suspected.BackgroundDefects of the mitochondrial genome are responsible for a heterogeneous group of clinical disorders, including cardiomyopathy. The majority of pathogenic mutations are heteroplasmic, with mutated and wild-type mitochondrial deoxyribonucleic acid (mtDNA) coexisting within the same cell. Homoplasmic mutations (present in every copy of the genome within the cell) present a difficult challenge in terms of diagnosis and assigning pathogenicity, as human mtDNA is highly polymorphic.MethodsA detailed clinical, histochemical, biochemical, and molecular genetic analysis was performed on two families with HCM to investigate the underlying mitochondrial defect.ResultsCardiac tissue from an affected child in the presenting family exhibited severe deficiencies of mitochondrial respiratory chain enzymes, whereas histochemical and biochemical studies of the skeletal muscle were normal. Mitochondrial DNA sequencing revealed an A4300G transition in the mitochondrial transfer ribonucleic acid (tRNA)Ilegene, which was shown to be homoplasmic by polymerase chain reaction/restriction fragment length polymorphism analysis in all samples from affected individuals and other maternal relatives. In a second family, previously reported as heteroplasmic for this base substitution, the mutation has subsequently been shown to be homoplasmic. The pathogenic role for this mutation was confirmed by high-resolution Northern blot analysis of heart tissue from both families, revealing very low steady-state levels of the mature mitochondrial tRNAIle.ConclusionsThis report documents, for the first time, that a homoplasmic mitochondrial tRNA mutation may cause maternally inherited HCM. It highlights the significant contribution that homoplasmic mitochondrial tRNA substitutions may play in the development of cardiac disease. A restriction of the biochemical defect to the affected tissue has important implications for the screening of patients with cardiomyopathy for mitochondrial disease.

Objectives. It has been demonstrated that clitoral and vaginal tissues express nitric oxide synthase isoforms in a way that parallels that of the penile corpus cavernosum. Considering the role of the vagina in the female sexual response and the anatomic connection between the clitoris and the anterosuperior vaginal wall, our aim was to study the distribution of type 5 phosphodiesterase (PDE5) in the anterosuperior wall of the human vagina. Methods. Immunohistochemistry was performed on the vaginal tissue of 14 women obtained at autopsy and on exfoliated cells of the vaginal epithelium obtained from 5 healthy female donors. Specific antibodies against PDE5 were tested on both paraffin sections and cytologic smears. Immunoblotting experiments were performed in parallel with the same antibodies. Results. The histologic analysis of human cadaveric vaginal tissue revealed that PDE5 immunoreactivity was mostly localized in the smooth muscle of vessels, forming a pseudocavernous tissue in the vaginal wall and endothelium. The Skene periurethral glands and vaginal epithelium were also positive for the antibody. The latter finding was confirmed using exfoliated cells of the vaginal epithelium harvested in vivo. Conclusions. The presence and tissue distribution of PDE5 in the human vagina suggest that the integrated system of nitric oxide synthase-PDE5 may play a physiologic role not only in the male sexual response but also in female sexual arousal.

Background: A persistent left superior vena cava (PLSVC) is a relatively frequent systemic venous anomaly associated with congenital heart defects. This anomaly has been explained with the persistence of the left superior cardinal vein. PLSVC usually drains into the right atrium, via coronary sinus, but it joins the left atrium in approximately 8% of the cases either directly in the setting of atrial isomerism, or via an unroofed coronary sinus, or through a coronary sinus type atrial septal defect. Case report: We describe a case of an adult patient with atria in the situs solitus, PLSVC draining into the left atrium, atresia of coronary sinus without atrial septal defect, and with additional cardiac anomalies (ventricular septal defect and discrete subaortic stenosis). Conclusion: A possible embryological explanation to this case rises from a right partial isomerism of the superior cardinal veins, which gives reason for both the coexistence of the PLSVC draining into the left atrium and the absence of coronary sinus, atrial septal defect, or coronary sinus ostium.

Obstructive disease of the epicardial coronary arteries was recognized as the cause of angina pectoris >2 centuries ago, and sudden thrombotic occlusion of an epicardial coronary artery has been established as the cause of acute myocardial infarction for >100 years. In the past 2 decades, dysfunction of the coronary microvasculature emerged as an additional mechanism of myocardial ischaemia that bears important prognostic implications. The coronary microvasculature (vessels