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Endothelial-to-mesenchymal transition (EndoMT) is a cellular process often initiated by the transforming growth factor β (TGF-β) family of ligands. Although required for normal heart valve development, deregulated EndoMT is linked to a wide range of pathological conditions. Here, we demonstrate that endothelial fatty acid oxidation (FAO) is a critical in vitro and in vivo regulator of EndoMT. We further show that this FAO-dependent metabolic regulation of EndoMT occurs through alterations in intracellular acetyl-CoA levels. Disruption of FAO via conditional deletion of endothelial carnitine palmitoyltransferase II (Cpt2) augments the magnitude of embryonic EndoMT, resulting in thickening of cardiac valves. Consistent with the known pathological effects of EndoMT, adult Cpt2 mice demonstrate increased permeability in multiple vascular beds. Taken together, these results demonstrate that endothelial FAO is required to maintain endothelial cell fate and that therapeutic manipulation of endothelial metabolism could provide the basis for treating a growing number of EndoMT-linked pathological conditions.
Copyright © 2018 Elsevier Inc. All rights reserved.
OBJECTIVES - We compared the utility of membrane-associated guanylate kinase, WW and PDZ domain-containing protein 2 (MAGI-2) and α-methylacyl CoA (AMACR) by immunohistochemistry in diagnosing prostatic adenocarcinoma.
METHODS - Seventy-eight radical prostatectomies were used to construct three tissue microarrays with 512 cores, including benign prostatic tissue, benign prostatic hyperplasia, high-grade prostatic intraepithelial neoplasia (HGPIN), and adenocarcinoma. AMACR and MAGI-2 immunohistochemistry were evaluated by visual and image analysis.
RESULTS - MAGI-2 and AMACR were significantly higher in adenocarcinoma and HGPIN compared with benign tissue. At H-score cutoffs of 300 and 200, MAGI-2 was more accurate in distinguishing benign from malignant glands than AMACR. Areas under the curve by image and visual analysis were 0.846 and 0.818 for MAGI-2 and 0.937 and 0.924 for AMACR, respectively. The accuracy of MAGI-2 in distinguishing benign from malignant glands on the same core was higher (95% vs 88%).
CONCLUSIONS - MAGI-2 could represent a useful adjunct for diagnosis of prostatic adenocarcinoma, especially when AMACR is not discriminatory.
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There is substantial evidence that NMDA receptor (NMDAR) hypofunction contributes to the pathophysiology of schizophrenia (SCZ). A recent large-scale genome-wide association study identified serine racemase (SR), the enzyme that produces the NMDAR co-agonist D-serine, as a risk gene for SCZ. Serine racemase knockout (SR-/-) mice, which lack D-serine, exhibit many of the neurochemical and behavioral abnormalities observed in SCZ. Metabotropic glutamate receptor 5 (mGlu5)-positive allosteric modulators (PAMs) are currently being developed to treat cognitive dysfunction. We used in vitro electrophysiology to determine whether the mGlu5 PAM VU0409551 directly enhances NMDAR function in hippocampal slices from adult male SR-/- mice. We administered VU0409551 systemically for 5 days to adult male wild-type C57BL/6 animals to determine the optimal dose to test in SR-/- mice. We used western blot analyses and trace-fear conditioning to determine whether 5 days of VU0409551 treatment could reverse the neuroplasticity and learning deficits, respectively, in SR-/- mice. We show that VU0409551 enhances NMDAR function and rescues long-term potentiation in hippocampal slices obtained from SR-/- mice. Systemic treatment with VU0409551 (10 and 30 mg/kg) to wild-type mice causes a dose-dependent increase in the Akt/GS3Kα/β signaling pathway, which is reduced in SR-/- mice and in SCZ. Furthermore, the administration of VU0409551 to SR-/- mice reverses their deficits in several neuroplasticity signaling pathways and improves their contextual fear memory. These results support positive allosteric modulation of mGlu5, particularly with VU0409551, as a viable mechanism to reverse the deficits in NMDAR function, synaptic plasticity, and memory that are known to be impaired in SCZ.
Napsin A and α-methylacyl-coenzyme A racemase (AMACR, P504S) have recently been described as being frequently expressed in clear cell carcinomas (CCC) of the gynecological tract. The present study was conducted to assess the test performance of these newer markers relative to the more traditional marker, hepatocyte nuclear factor 1β (HNF1β), in a large and histotypically diverse dataset. A total of 279 ovarian tumours in tissue microarrays were immunohistochemically assessed for the expression of Napsin A, AMACR and HNF1β. HNF1β, Napsin A and AMACR were expressed in 92%, 82% and 63% of 65 CCC, 7%, 1% and 1% of 101 serous carcinomas, 37%, 5.3% and 0% of 19 endometrioid carcinomas, 60%, 0% and 0% of 45 mucinous tumours, 100%, 0% and 0% of seven yolk sac tumours, and 0%, 16.7% and 16.7% of six steroid cell tumours NOS, respectively. All other tumours, including 18 adult-type granulosa cell tumours, eight dysgerminomas and nine other miscellaneous tumour types were negative for all three markers. Using a benchmark of ≥1% of tumour cells for positivity and CCC as the diagnostic end-point, the sensitivity, specificity, negative predictive value and positive predictive value of Napsin A expression were 0.82, 0.99, 0.94, and 0.98, respectively (odds ratio 439, p < 0.0001). Respective parameters were 0.92, 0.79, 0.97, and 0.58 (odds ratio 44, p < 0.0001) for HNF1β and 0.63, 0.99, 0.89, and 0.5 (odds ratio 112, p < 0.0001) for AMACR. The combination of any two positive markers, irrespective of the staining pattern of the third, significantly predicted the CCC histotype in every analytic scenario. In summary, HNF1β is highly sensitive but is suboptimally specific in isolation, whereas AMACR is highly specific but is suboptimally sensitive. Napsin A is specific but of intermediate sensitivity. Napsin A, AMACR and HNF1β are all viable markers of CCC that can be deployed as components of larger panels when CCC is a diagnostic consideration.
Our aim was to compare the usefulness of fatty acid synthase (FASn) with that of α-methylacyl coenzyme-A racemase (AMACR) in the diagnosis of prostatic adenocarcinoma. The expression of these 2 markers was compared in a tissue microarray containing 62 foci of benign glands and 36 foci of prostatic adenocarcinoma. Similar to AMACR, there was significantly higher FASn expression in adenocarcinoma compared with that in benign glands. The optimal accuracy rate and area under curve (AUC) by receiver operating characteristic analysis for FASn were not significantly different from those for AMACR (accuracy, 80% vs 87%; AUC, 0.942 vs 0.956; P for both, > .05). Moreover, in cases with coexistent malignant and benign glands on the same core, FASn could selectively distinguish a proportion of cases (17/21 [81%]) similar to using AMACR. We conclude that FASn may aid in the diagnosis of prostatic adenocarcinoma, at least to supplement AMACR as another positive marker of carcinoma and potentially increase diagnostic accuracy.
Typically glands of prostatic adenocarcinoma have a single cell lining, although stratification can be seen in invasive carcinomas with a cribriform architecture, including ductal carcinoma. The presence and diagnostic significance of stratified cells within non-cribriform carcinomatous prostatic glands has not been well addressed. The histomorphological features and immunohistochemical profile of cases of non-cribriform prostatic adenocarcinoma with stratified malignant glandular epithelium were analyzed. These cases were identified from needle biopsy cases from the consultation files of one of the authors and from a review of 150 consecutive in-house needle biopsy cases of prostatic adenocarcinoma. Immunohistochemistry was performed utilizing antibodies reactive against high molecular weight cytokeratin (34betaE12), p63 and alpha-methylacyl-coenzyme-A racemase (AMACR). A total of 8 cases were identified, including 2 from the 150 consecutive in-house cases (1.3%). In 4 cases, the focus with glands having stratified epithelium was the sole carcinomatous component in the biopsy, while such a component represented 5-30% of the invasive carcinoma seen elsewhere in the remaining cases. The main attribute in all these foci was the presence of glandular profiles lined by several layers of epithelial cells with cytological and architectural features resembling flat or tufted high-grade prostatic intraepithelial neoplasia, but lacking basal cells as confirmed by negative 34betaE12 and/or p63 immunostains in all cases. The AMACR staining profile of the stratified foci was variable, with 4 foci showing positivity, and 3 foci being negative, including two cases that displayed AMACR positivity in adjacent non-stratified prostatic adenocarcinoma. Prostatic adenocarcinoma with stratified malignant glandular epithelium can be identified in prostate needle biopsy samples harboring non-cribriform prostatic adenocarcinoma and resembles glands with high-grade prostatic intraepithelial neoplasia. These 'PIN-like' carcinomas can present in pure form. Recognition of this pattern of prostatic adenocarcinoma is necessary to correctly diagnose such cases as invasive carcinoma.
We assessed the utility of using a p63/a-methylacyl-coenzyme-A racemase (AMACR) antibody cocktail on destained H&E-stained sections. We transferred 61 stored (7-11 months old) and 10 recent (<1 month old) H&E-stained sections of prostate needle biopsy tissues to charged slides and subsequently stained them with a p63/AMACR immunohistochemical antibody cocktail. The AMACR and p63 staining intensities were compared with those obtained with the same antibody cocktail performed on sections recut directly from the paraffin block. Transfer of sections and subsequent immunohistochemical staining was successful in 69 (97%) of 71 cases. For stored cases, there were similar AMACR and p63 staining intensities in destained and recut sections in 55 (90%) and 11 (18%) of 61 cases, respectively. In recent sections, AMACR and p63 staining intensities were almost identical by both methods. We conclude that p63/AMACR cocktail immunostaining of destained H&E-stained sections is a viable approach in the workup of small "suspicious" foci in recently sectioned prostate needle biopsy tissues. This approach is best used when 2 or more H&E-stained sections harbor the suspicious focus, as we always recommended preservation of at least 1 H&E-stained section.
The diagnosis of prostatic carcinoma and especially minimal prostatic carcinoma can sometimes be challenging on needle core biopsy and occasionally immunohistochemistry is an aid in the diagnosis. Immunostains, such as those directed against the basal cell marker p63 and, more recently, employing antibodies reactive with alpha-methylacyl-CoA racemase (AMACR), can be useful in this situation. The aim of this investigation was to assess the diagnostic utility of a p63/AMACR antibody cocktail and compare the staining pattern it produces with that using the individual antibodies alone. A retrospective review of 31 consecutive radical prostatectomy specimens and 150 prostate needle biopsy samples was performed to select histologic sections showing foci of prostatic carcinoma and/or minimal prostatic carcinoma, high-grade prostatic intraepithelial neoplasia (HGPIN), as well as common benign mimickers of prostatic carcinoma, to include atrophy and basal cell hyperplasia, especially with prominent nucleoli. Serial histologic sections from the corresponding paraffin blocks were stained with hematoxylin and eosin and by immunostains for p63, AMACR, and a prediluted antibody cocktail comprising both. The diagnostic utility of the cocktail was assessed, and the staining characteristics it produced were compared with those using the individual immunostains. In 430 foci, the cocktail produced a p63 staining profile identical to that using the single p63 antibody. Distinction of the nuclear p63 signal from the cytoplasmic AMACR localization was readily accomplished. There was an excellent agreement (kappa=0.91; P<0.0001) between the AMACR staining profile using the cocktail and the single AMACR antibody alone. The cocktail was very useful in highlighting prostatic carcinoma associated with HGPIN, flat and cribriform HGPIN, and distorted foci of minimal prostatic carcinoma. These data indicate that use of a p63/AMACR cocktail is essentially equivalent to use of each antibody separately for immunohistochemical confirmation of a diagnosis of prostatic carcinoma in needle biopsy. This cocktail would be of diagnostic utility when only limited tissue is available for immunohistochemical evaluation of small, diagnostically difficult foci in prostate needle biopsy tissue.