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关键词： cobia   methionine   PI3K   Akt   TOR pathway   primary muscle cells  

摘要： Methionine (Met) plays important roles in protein synthesis and in diverse cellular processes in eukaryotes. In the present study, primary muscle cells of cobia fish Rachycentron canadum were isolated and treated with Met at different concentrations (0.5, 1, 2, 5 or 10 mM) for 24 h in culture media. The results show that the 5 mM Met group had the highest cell viability and total protein content. The phosphorylation levels of phosphoinositide 3-kinase (PI3K) p85 on Tyr458 and of protein kinase B (Akt) on Ser371 were significantly affected by the Met concentration during incubation. Cells incubated with 5 or 10 mM Met showed higher mRNA levels of the target rapamycin (TOR) and higher phosphorylation levels of TOR on Ser2448. Cells incubated with 5 mM Met had the highest phosphorylation levels of ribosomal p70-S6 kinase (S6K1) on Ser371 and of 4E-binding protein 1 (4E-BP1) on Thr37/46. Cells incubated with PI3K and TOR inhibitors showed that LY294002 and rapamycin significantly inhibited the phosphorylation of Akt on Ser371 and of TOR on Ser2448 respectively. These results indicate that Met concentration affects cell viability and is associated with changes in the PI3K/Akt/TOR pathway.

摘要： Background Although pathological complete response (pCR) after multimodal treatment for esophageal cancer is associated to the best prognosis, recurrence may occur in 20-40% of cases. The present study investigated the recurrence pattern and predictive factors of recurrence after pCR in patients with esophageal cancer. Methods In this study, 427 patients received preoperative treatment for either esophageal squamous cell carcinoma (SCC) or adenocarcinoma at Verona University Hospital between 2000 and 2018. Of these, 145 patients (34%) achieved a pCR. Long-term prognosis, recurrence pattern, and risk factors for relapse in pCR patients were analysed. Results During a median follow-up of 52 months, 37 relapses (25.5%) occurred, mostly at distant level (n = 28). Nearly all locoregional relapses (8/9) were detected in SCC cases. The 5-year overall survival and cancer-related survival were 71.7% (95% confidence interval [CI] 62.6-78.9%) and 77.5% (95% CI 68.5-84.2%) respectively. Male sex, higher body mass index, and cT4 were significant risk factors for recurrence at univariate analysis. The multivariate analysis confirmed the role of cT4 as predictor of recurrence only in SCCs. Conclusions Esophageal cancer recurs in about one-fourth of pCR cases. A fair number of local recurrences occurs in SCCs, but the main problem is the systemic disease control. According to our analysis, SCCs patients with cT4 stage have an increased risk to recur, so they should be managed differently by a personalized approach in terms of adjuvant treatment and follow-up.

关键词： Berberine   Albumin   Nanoparticle   Thermodynamic   Neurons   Antioxidant   Alzheimer's Disease  

摘要： Berberine has shown an outstanding antioxidant activity, however the low bioavailability limits its applications in pharmaceutical platforms. Therefore, in this paper, after fabrication of the berberine-HSA nanoparticles by desolvation method, they were well characterized by TEM, SEM, DLS, and FTIR techniques. Afterwards the interaction of HSA and the berberine was evaluated by molecular docking analysis. Finally, the antioxidant activity of the berberine-HSA nanoparticles against H2O2-induced oxidative stress in cultured neurons as a model of AD was evaluated by cellular assays. The results showed that the prepared berberine- HSA nanoparticles have a spherical-shaped morphology with a size of around 100 nm and zeta potential value of -31.84 mV. The solubility value of nanoparticles was calculated to be 40.27%, with a berberine loading of 19.37%, berberine entrapment efficiency of 70.34%, and nanoparticles yield of 88.91%. Also, it was shown that the berberine is not significantly released from HSA nanoparticles within 24 hours. Afterwards, molecular docking investigation revealed that berberine spontaneously interacts with HSA through electrostatic interaction. Finally, cellular assays disclosed that the pretreatment of neuronal cultures with berberine-HSA nanoparticles decreased the H2O2-stimulated cytotoxicity and relevant morphological changes and enhanced the CAT activity. In conclusion, it can be indicated that the nanoformulation of the berberine can be used as a promising platform for inhibition of oxidative damage-induced Alzheimer's disease (AD).

摘要： 813

关键词： nanopore technology   RNA-seq   MinION   sequencing depth   multiplexed samples   barcoding   SQK-PCS109  

摘要： Transcript sequencing is a crucial tool for gaining a deep understanding of biological processes in diagnostic and clinical medicine. Given their potential to study novel complex eukaryotic transcriptomes, long-read sequencing technologies are able to overcome some limitations of short-read RNA-Seq approaches. Oxford Nanopore Technologies (ONT) offers the ability to generate long-read sequencing data in real time via portable protein nanopore USB devices. This work aimed to provide the user with the number of reads that should be sequenced, through the ONT MinION platform, to reach the desired accuracy level for a human cell RNA study. We sequenced three cDNA libraries prepared from poly-adenosine RNA of human primary cardiac fibroblasts. Since the runs were comparable, they were combined in a total dataset of 48 million reads. Synthetic datasets with different sizes were generated starting from the total and analyzed in terms of the number of identified genes and their expression levels. As expected, an improved sensitivity was obtained, increasing the sequencing depth, particularly for the non-coding genes. The reliability of expression levels was assayed by (i) comparison with PCR quantifications of selected genes and (ii) by the implementation of a user-friendly multiplexing method in a single run.

关键词： Kidney   Oncocytic renal cell carcinoma   Papillary   Unclassified   Copy number variation pattern   Oncocytoma   Overlapping  

摘要： So-called oncocytic papillary renal cell carcinoma (OPRCC) is a poorly defined variant of papillary renal cell carcinoma. Since its first description, several studies were published with conflicting results, and thus precise definition is lacking. A cohort of 39 PRCCs composed of oncocytic cells were analyzed. Cases were divided into 3 groups based on copy number variation (CNV) pattern. The first group consisted of 23 cases with CNV equal to renal oncocytoma. The second group consisted of 7 cases with polysomy of chromosomes 7 and 17 and the last group of 9 cases included those with variable CNV. Epidemiologic, morphologic and immunohistochemical features varied among the groups. There were not any particular histomorphologic features correlating with any of the genetic subgroups. Further, a combination of morphologic, immunohistochemical, and molecular-genetic features did not allow to precisely predict biologic behavior. Owing to variable CNV pattern in OPRCC, strict adherence to morphology and immunohistochemical profile is recommended, particularly in limited samples (i.e., core biopsy). Applying CNV pattern as a part of a diagnostic algorithm can be potentially misleading. OPRCC is a highly variable group of tumors, which might be misdiagnosed as renal oncocytoma. Using the term OPRCC as a distinct diagnostic entity is, thanks to its high heterogeneity, questionable.

摘要： 874

关键词： VANETs   SDVN   security   public key infrastructure (PKI)   digital signature   AVISPA  

摘要： Recently, by the rapid development of Vehicular Ad Hoc Networks (VANETs) and the advancement of Software Defined Networking (SDN) as an emerging technology, the Software-Defined Vehicular Network (SDVN) has a tremendous attraction in the academia and research community. SDN's unique properties and features, such as its flexibility, programmability, and centralized control, make the network scalable and straightforward. In VANETs, traffic management and secure communication of vehicle information using the public network are the main research dimensions in the current era for the researchers to be considered while designing an efficient and secure VANETs architecture. This paper highlights the possible identified threat vectors and efficiently resolves the network vulnerabilities to design a novel and secure hierarchic architecture for SDVN. To solve the above problem, we proposed a Public Key Infrastructure-based digital signature model for efficient and secure communication from Vehicle to Vehicle. We also used the public key authority infrastructure for Vehicle to Infrastructure and the three-way handshake method for secure session creation and secure data communication in the SDN controller. The proposed security is validated through the well-known simulation tool AVISPA. Additionally, a formal security model is applied to validate the design hierarchic architecture's fundamental security properties for SDVN in an efficient and desirable way. In a comparative analysis, we prove that our proposed scheme fulfills all the essential security properties compared to other states of the art schemes.

关键词： SARS-CoV-2   complement   anaphylatoxin receptors   primary human airway epithelial cells  

摘要： Background: Excessive inflammation triggered by a hitherto undescribed mechanism is a hallmark of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections and is associated with enhanced pathogenicity and mortality. Objective: Complement hyperactivation promotes lung injury and was observed in patients suffering from Middle East respiratory syndrome-related coronavirus, SARS-CoV-1, and SARS-CoV-2 infections. Therefore, we investigated the very first interactions of primary human airway epithelial cells on exposure to SARS-CoV-2 in terms of complement component 3 (C3)-mediated effects. Methods: For this, we used highly differentiated primary human 3-dimensional tissue models infected with SARS-CoV-2 patient isolates. On infection, viral load, viral infectivity, intracellular complement activation, inflammatory mechanisms, and tissue destruction were analyzed by real-time RT-PCR, high content screening, plaque assays, luminex analyses, and transepithelial electrical resistance measurements. Results: Here, we show that primary normal human bronchial and small airway epithelial cells respond to SARS-CoV-2 infection by an inflated local C3 mobilization. SARS-CoV-2 infection resulted in exaggerated intracellular complement activation and destruction of the epithelial integrity in monolayer cultures of primary human airway cells and highly differentiated, pseudostratified, mucus-producing, ciliated respiratory tissue models. SARS-CoV-2-infected 3-dimensional cultures secreted significantly higher levels of C3a and the proinflammatory cytokines IL-6, monocyte chemoattractant protein 1, IL-1a, and RANTES. Conclusions: Crucially, we illustrate here for the first time that targeting the anaphylotoxin receptors C3a receptor and C5a receptor in nonimmune respiratory cells can prevent intrinsic lung inflammation and tissue damage. This opens up the exciting possibility in the treatment of COVID-19.