Limitations and pitfalls of 99mTc-EDDA/HYNIC-TOC (Tektrotyd) scintigraphy

Tektrotyd kit was developed by Polatom company for 99mTc labeling to make an alternative tracer of somatostatin receptor scintigraphy available. Since 2005, 99mTc-EDDA/HYNIC-Tyr3-Octreotide has been used in clinical imaging and achieved high impact in management of patients with neuroendocrine tumors. Knowing the limitations and pitfalls is essential to provide ac­curate diagnosis. Therefore, the potential pitfalls associated with the use of 99mTc-EDDA/HYNIC-TOC are reviewed on the basis of own experience. Data were analyzed of 310 patients who underwent somatostatin receptor scintigraphy with 99mTc-Tektrotyd. Pitfalls during radiolabeling process or acquisition can worsen the sensitivity of SRS (somatostatin receptor scintigraphy). Recognizing physi­ological and clinical pitfalls, the diagnostic accuracy will improve

Staurosporine induces necroptotic cell death under caspase-compromised conditions in U937 cells

For a long time necrosis was thought to be an uncontrolled process but evidences recently have revealed that necrosis can also occur in a regulated manner. Necroptosis, a type of programmed necrosis is defined as a death receptor-initiated process under caspase-compromised conditions. The process requires the kinase activity of receptor-interacting protein kinase 1 and 3 (RIPK1 and RIPK3) and mixed lineage kinase domain-like protein (MLKL), as a substrate of RIPK3. The further downstream events remain elusive. We applied known inhibitors to characterize the contributing enzymes in necroptosis and their effect on cell viability and different cellular functions were detected mainly by flow cytometry. Here we report that staurosporine, the classical inducer of intrinsic apoptotic pathway can induce necroptosis under caspase-compromised conditions in U937 cell line. This process could be hampered at least partially by the RIPK1 inhibitor necrotstin-1 and by the heat shock protein 90 kDa inhibitor geldanamycin. Moreover both the staurosporine-triggered and the classical death ligand-induced necroptotic pathway can be effectively arrested by a lysosomal enzyme inhibitor CA-074-OMe and the recently discovered MLKL inhibitor necrosulfonamide. We also confirmed that the enzymatic role of poly(ADP-ribose)polymerase (PARP) is dispensable in necroptosis but it contributes to membrane disruption in secondary necrosis. In conclusion, we identified a novel way of necroptosis induction that can facilitate our understanding of the molecular mechanisms of necroptosis. Our results shed light on alternative application of staurosporine, as a possible anticancer therapeutic agent. Furthermore, we showed that the CA-074-OMe has a target in the signaling pathway leading to necroptosis. Finally, we could differentiate necroptotic and secondary necrotic processes based on participation of PARP enzyme

Refined heart failure detection algorithm for improved clinical reliability of OptiVol alerts in CRT-D recipients

   Background: The reliability of intrathoracic impedance monitoring for prediction of heart failure (HF) by implantable cardiac devices is controversial. Despite using additional device-based parameters described in the PARTNERS HF study, such as new onset of arrhythmias, abnormal autonomics, low biventricular pacing rate or patient activity level, the predictive power of device diagnostic algorithm is still in doubt. The objective of this study was to compare the device diagnostic algorithm described in the PARTNERS HF study to a newly developed algorithm applying refined diagnostic criteria. Methods: Fourty two patients were prospectively enrolled who had been implanted with an intrathoracic impedance and remote monitoring capable implantable cardiac defibrillator with a cardiac resychroniza­tion therapy (CRT-D) device in this observational study. If a remote OptiVolTM alert occurred, patients were checked for presence of HF symptoms. A new algorithm was derived from the original PARTNERS HF criteria, considering more sensitive cut-offs and changes of patterns of the device-based parameters. Results: During an average follow-up of 38 months, 722 remote transmissions were received. From the total of 128 transmissions with OptiVol alerts, 32 (25%) corresponded to true HF events. Upon multivariate discriminant analysis, low patient activity, high nocturnal heart rate, and low CRT pacing (< 90%) proved to be independent predictors of true HF events (all p < 0.01). Incorporating these three refined criteria in a new algorithm, the diagnostic yield of OptiVol was improved by increasing specific­ity from 37.5% to 86.5%, positive predictive value from 34.1% to 69.8% and area under the curve from 0.787 to 0.922 (p < 0.01), without a relevant loss in sensitivity (96.9% vs. 93.8%). Conclusions: A refined device diagnostic algorithm based on the parameters of low activity level, high nocturnal heart rate, and suboptimal biventricular pacing might improve the clinical reliability of OptiVol alerts.  

Suppression of AMPK/aak-2 by NRF2/SKN-1 down-regulates autophagy during prolonged oxidative stress.

NF-E2-related factor 2 (NRF2) transcription factor has a fundamental role in cell homeostasis maintenance as one of the master regulators of oxidative and electrophilic stress responses. Previous studies have shown that a regulatory connection exists between NRF2 and autophagy during reactive oxygen species-generated oxidative stress. The aim of the present study was to investigate how autophagy is turned off during prolonged oxidative stress, to avoid overeating and destruction of essential cellular components. AMPK is a key cellular energy sensor highly conserved in eukaryotic organisms, and it has an essential role in autophagy activation at various stress events. Here the role of human AMPK and its Caenorhabditis elegans counterpart AAK-2 was explored upon oxidative stress. We investigated the regulatory connection between NRF2 and AMPK during oxidative stress induced by tert-butyl hydroperoxide (TBHP) in HEK293T cells and C. elegans. Putative conserved NRF2/protein skinhead-1 binding sites were found in AMPK/aak-2 genes by in silico analysis and were later confirmed experimentally by using EMSA. After addition of TBHP, NRF2 and AMPK showed a quick activation; AMPK was later down-regulated, however, while NRF2 level remained high. Autophagosome formation and Unc-51-like autophagy activating kinase 1 phosphorylation were initially stimulated, but they returned to basal values after 4 h of TBHP treatment. The silencing of NRF2 resulted in a constant activation of AMPK leading to hyperactivation of autophagy during oxidative stress. We observed the same effects in C. elegans demonstrating the conservation of this self-defense mechanism to save cells from hyperactivated autophagy upon prolonged oxidative stress. We conclude that NRF2 negatively regulates autophagy through delayed down-regulation of the expression of AMPK upon prolonged oxidative stress. This regulatory connection between NRF2 and AMPK may have an important role in understanding how autophagy is regulated in chronic human morbidities characterized by oxidative stress, such as neurodegenerative diseases, certain cancer types, and in metabolic diseases.-Kosztelnik, M., Kurucz, A., Papp, D., Jones, E., Sigmond, T., Barna, J., Traka, M. H., Lorincz, T., Szarka, A., Banhegyi, G., Vellai, T., Korcsmaros, T., Kapuy, O. Suppression of AMPK/aak-2 by NRF2/SKN-1 down-regulates autophagy during prolonged oxidative stress

Prognostic impact of reduced connexin43 expression and gap junction coupling of neoplastic stromal cells in giant cell tumor of bone

Missense mutations of the GJA1 gene encoding the gap junction channel protein connexin43 (Cx43) cause bone malformations resulting in oculodentodigital dysplasia (ODDD), while GJA1 null and ODDD mutant mice develop osteopenia. In this study we investigated Cx43 expression and channel functions in giant cell tumor of bone (GCTB), a locally aggressive osteolytic lesion with uncertain progression. Cx43 protein levels assessed by immunohistochemistry were correlated with GCTB cell types, clinico-radiological stages and progression free survival in tissue microarrays of 89 primary and 34 recurrent GCTB cases. Cx43 expression, phosphorylation, subcellular distribution and gap junction coupling was also investigated and compared between cultured neoplastic GCTB stromal cells and bone marow stromal cells or HDFa fibroblasts as a control. In GCTB tissues, most Cx43 was produced by CD163 negative neoplastic stromal cells and less by CD163 positive reactive monocytes/macrophages or by giant cells. Significantly less Cx43 was detected in alpha-smooth muscle actin positive than alpha-smooth muscle actin negative stromal cells and in osteoclast-rich tumor nests than in the adjacent reactive stroma. Progressively reduced Cx43 production in GCTB was significantly linked to advanced clinico-radiological stages and worse progression free survival. In neoplastic GCTB stromal cell cultures most Cx43 protein was localized in the paranuclear-Golgi region, while it was concentrated in the cell membranes both in bone marrow stromal cells and HDFa fibroblasts. In Western blots, alkaline phosphatase sensitive bands, linked to serine residues (Ser369, Ser372 or Ser373) detected in control cells, were missing in GCTB stromal cells. Defective cell membrane localization of Cx43 channels was in line with the significantly reduced transfer of the 622 Da fluorescing calcein dye between GCTB stromal cells. Our results show that significant downregulation of Cx43 expression and gap junction coupling in neoplastic stromal cells are associated with the clinical progression and worse prognosis in GCTB

PP13, Maternal ABO Blood Groups and the Risk Assessment of Pregnancy Complications

Placental Protein 13 (PP13), an early biomarker of preeclampsia, is a placenta-specific galectin that binds beta-galactosides, building-blocks of ABO blood-group antigens, possibly affecting its bioavailability in blood.We studied PP13-binding to erythrocytes, maternal blood-group effect on serum PP13 and its performance as a predictor of preeclampsia and intrauterine growth restriction (IUGR). Datasets of maternal serum PP13 in Caucasian (n = 1078) and Hispanic (n = 242) women were analyzed according to blood groups. In vivo, in vitro and in silico PP13-binding to ABO blood-group antigens and erythrocytes were studied by PP13-immunostainings of placental tissue-microarrays, flow-cytometry of erythrocyte-bound PP13, and model-building of PP13--blood-group H antigen complex, respectively. Women with blood group AB had the lowest serum PP13 in the first trimester, while those with blood group B had the highest PP13 throughout pregnancy. In accordance, PP13-binding was the strongest to blood-group AB erythrocytes and weakest to blood-group B erythrocytes. PP13-staining of maternal and fetal erythrocytes was revealed, and a plausible molecular model of PP13 complexed with blood-group H antigen was built. Adjustment of PP13 MoMs to maternal ABO blood group improved the prediction accuracy of first trimester maternal serum PP13 MoMs for preeclampsia and IUGR.ABO blood group can alter PP13-bioavailability in blood, and it may also be a key determinant for other lectins' bioavailability in the circulation. The adjustment of PP13 MoMs to ABO blood group improves the predictive accuracy of this test