Toll-like receptor 5 tunes hepatic and pancreatic stellate cells activation.

OBJECTIVE: Stellate cells are responsible for liver and pancreas fibrosis and strictly correlate with tumourigenesis. Although their activation is reversible, an exacerbated signalling triggers chronic fibrosis. Toll-like receptors (TLRs) modulate stellate cells transition. TLR5 transduces the signal deriving by the binding to bacterial flagellin from invading mobile bacteria. DESIGN: Human hepatic and pancreatic stellate cells were activated by the administration of transforming growth factor-beta (TGF-ß). TLR5 was transiently knocked down by short-interference RNA transfection. Reverse Transcription-quantitativePCR and western blot were performed to analyse the transcript and protein level of TLR5 and the transition players. Fluorescence microscopy was performed to identify these targets in spheroids and in the sections of murine fibrotic liver. RESULTS: TGF-ß-activated human hepatic and pancreatic stellate cells showed an increase of TLR5 expression. TLR5 knockdown blocked the activation of those stellate cells. Furthermore, TLR5 busted during murine liver fibrosis and co-localised with the inducible Collagen I. Flagellin suppressed TLR5, COL1A1 and ACTA2 expression after the administration of TGF-ß. Instead, the antagonist of TLR5 did not block the effect of TGF-ß. Wortmannin, a specific AKT inhibitor, induced TLR5 but not COL1A1 and ACTA2 transcript and protein level. CONCLUSION: TGF-ß-mediated activation of hepatic and pancreatic stellate cells requires the over-expression of TLR5. Instead, its autonomous signalling inhibits the activation of the stellate cells, thus prompting a signalling through different regulatory pathways.

Radiofrequency and microwave ablation controls hepatic oligoprogression of advanced gastroenteropancreatic neuroendocrine tumors.

In progression of multifocal liver metastases of gastroenteropancreatic neuroendocrine tumors (GEP-NET) escalation of systemic therapy is indicated. The aim of this retrospective study was to investigate the potential of local thermal ablation in hepatic oligoprogression and otherwise stable disease in GEP-NET. Patients with hepatic oligoprogression and otherwise stable disease, who underwent radiofrequency ablation (RFA) or microwave ablation (MWA) for local control, were included in the study. Thermal ablation was performed while maintaining the ongoing systemic therapy or without addition of a systemic therapy. The effectiveness of this therapeutic approach was evaluated by determination of local treatment success, improvement of progression-free survival (PFS) and the safety. Seventeen thermal ablation procedures were performed in 13 patients with well differentiated NET including seven ileum NET, four pancreatic NET, one appendix NET and one rectum NET. RFA and MWA of liver metastases were well tolerated without major complications. Thermal ablation resulted in an estimated median PFS of 62.6 weeks (mean 50.5 weeks; range 10.1-78.9 weeks) per procedure. In four patients, two ablation procedures were performed throughout the course of their disease resulting in an estimated median PFS of 69.1 weeks (mean 71.6 weeks; range 10.1-123.1 weeks) per patient. Start or change of systemic therapy could be delayed up to 123.1 weeks by using thermal ablations for isolated progression of single liver metastases. 88% of thermal ablations prolonged PFS. Thermal ablation of liver metastases in a non-curative intent has the potential to provide focal growth control and to prolong PFS in GEP-NET patients with hepatic oligoprogression.

Hepatic perivascular epithelioid cell tumor (PEComa): contrast-enhanced ultrasound (CEUS) characteristics-a case report and literature review.

Perivascular epithelioid cell tumors (PEComa) are rare mesenchymal neoplasms that arise from soft tissue of various organs such as the stomach, intestines, and lungs. We report a rare case of a primary PEComa of the liver and its characteristics on contrast-enhanced ultrasound (CEUS) in a 51-year-old female patient with an incidental finding of a hypoechoic liver lesion with peripheral hypervascularization on Doppler ultrasound. CEUS showed homogenous hypervascularity in the arterial phase that was consistent in the portal phase. In the late phase, a central washout phenomenon was evident. Histopathologic findings on sonographic biopsy of the lesion revealed a mesenchymal tumor with positivity for melanocytic markers Human Melanin Black-45 (HMB45) and Melan-A consistent with a PEComa. Despite the absence of high-risk features for malignancy, surgical resection was recommended due to the uncertain malignant potential of PEComas. The patient refused the operation and preferred sonographic follow-up; the lesion was stable over a period of 2 years. CEUS can provide valuable information regarding PEComa. After histological confirmation, the choice between resection and a watchful waiting must be made on individual basis.

The Expression of Autophagy-Associated Genes Represents a Valid Footprint for Aggressive Pancreatic Neuroendocrine Neoplasms.

Pancreatic neuroendocrine neoplasms (pNEN) are rare and heterogeneous tumors. Previous investigations have shown that autophagy can be a target for cancer therapy. This study aimed to determine the association between the expression of autophagy-associated gene transcripts and clinical parameters in pNEN. In total, 54 pNEN specimens were obtained from our human biobank. The patient characteristics were retrieved from the medical record. RT-qPCR was performed to assess the expression of the autophagic transcripts BECN1, MAP1LC3B, SQSTM1, UVRAG, TFEB, PRKAA1, and PRKAA2 in the pNEN specimens. A Mann-Whitney U test was used to detect differences in the expression of autophagic gene transcripts between different tumor characteristics. This study showed that G1 sporadic pNEN have a higher expression of autophagic genes compared to G2. Lymphatic and distant metastasis occurred significantly more often in pNEN with a decreased expression of the autophagic genes. Within sporadic pNEN, the insulinomas express higher levels of autophagic transcripts than gastrinomas and non-functional pNEN. MEN1-associated pNEN show a higher expression of autophagic genes than sporadic pNEN. In summary, a decreased expression of autophagic transcripts distinguishes metastatic from non-metastatic sporadic pNEN. The significance of autophagy as a molecular marker for prognosis and therapy decisions needs to be further investigated.

Towards a COVID-19 symptom triad: The importance of symptom constellations in the SARS-CoV-2 pandemic.

Pandemic scenarios like SARS-Cov-2 require rapid information aggregation. In the age of eHealth and data-driven medicine, publicly available symptom tracking tools offer efficient and scalable means of collecting and analyzing large amounts of data. As a result, information gains can be communicated to front-line providers. We have developed such an application in less than a month and reached more than 500 thousand users within 48 hours. The dataset contains information on basic epidemiological parameters, symptoms, risk factors and details on previous exposure to a COVID-19 patient. Exploratory Data Analysis revealed different symptoms reported by users with confirmed contacts vs. no confirmed contacts. The symptom combination of anosmia, cough and fatigue was the most important feature to differentiate the groups, while single symptoms such as anosmia, cough or fatigue alone were not sufficient. A linear regression model from the literature using the same symptom combination as features was applied on all data. Predictions matched the regional distribution of confirmed cases closely across Germany, while also indicating that the number of cases in northern federal states might be higher than officially reported. In conclusion, we report that symptom combinations anosmia, fatigue and cough are most likely to indicate an acute SARS-CoV-2 infection.

Multicenter Analysis of Presacral Neuroendocrine Neoplasms-Clinicopathological Characterization and Treatment Outcomes of a Rare Disease.

Background and Aims: Neuroendocrine neoplasms (NENs) of the presacral space are an extremely rare disease entity with largely unknown outcome and no established standard of care treatment. Therefore, we wanted to analyze clinical presentation, histopathological findings, treatment outcomes, and prognosis in a multicentric patient cohort. Methods: We searched local databases of six German NEN centers for patients with presacral NEN. Retrospective descriptive analyses of age, sex, stage at diagnosis, symptoms, grade, immunohistochemical investigations, biomarkers, treatment, and treatment outcome were performed. Kaplan-Meier analysis was used to determine median overall survival. Results: We identified 17 patients (11 female, 6 male) with a median age of 50 years (range, 35-66) at diagnosis. Twelve cases presented initially with distant metastases including bone metastases in nine cases. On pathological review the majority of patients had well-differentiated G2 tumors. Immunohistochemical profile resembled rectal NENs. All but one patient had non-functioning tumors. Somatostatin receptor imaging was positive in 14 of 15 investigated cases. Eight patients were treated surgically including palliative resections; 14 patients received somatostatin analogs with limited efficacy. With 14 PRRTs completed, 79% showed clinical benefit, whereas only one patient with neuroendocrine carcinoma (NEC) responded to chemotherapy. Treatment with everolimus in three patients was not successful, whereas cabozantinib resulted in a disease stabilization in a heavily pretreated patient. During a median observation period of 44.5 months, 6 patients died. Median overall survival was not reached. Conclusion: Presacral NEN are histopathologically similar to rectal NENs. Presacral NEN should be considered as possible primary in NEN of unknown primary. The majority of tumors is non-functioning and somatostatin receptor positive. PRRT demonstrated promising activity; tyrosine kinase inhibitors warrant further investigations. Further molecular characterization and prospective evaluation of this rare tumor entity are needed.

Modulation of Pancreatic Neuroendocrine Neoplastic Cell Fate by Autophagy-Mediated Death.

INTRODUCTION: Autophagic cell death in cancer cells can be mediated by inhibition of deacetylases. Although extensive studies have focused on the autophagic process in cancer, little is known about the role of autophagy in degrading cytosolic and nuclear components of pancreatic neuroendocrine neoplastic (pNEN) cells leading to cell death, thus improving the therapy of patients affected by pNEN. METHODS: 2D and 3D human pNEN and pancreatic stellate cells were treated with panobinostat and bafilomycin. Autophagy markers were detected by RT-qPCR, immunofluorescence, and Western blot. Autophagosomes were detected by electron microscopy and their maturation by real-time fluorescence of LC3B stable transfected cells. ChIP was performed at the cAMP responsive element. Immunofluorescence was performed in murine pancreatic tissue. RESULTS: We observed that pan-deacetylase inhibitor panobinostat treatment causes autophagic cell death in pNEN cells. We also found that although AMPK-α phosphorylation is counterbalanced by phosphorylated AKT, it is not capable to inhibiting autophagic cell death. However, the binding activity of the cAMP responsive element is prompted by panobinostat. Although autophagy inhibition prevented autophagosome synthesis, maturation, and cell death, panobinostat treatment induced the accumulation of mature autophagosomes in the cytosol and the nucleus, leading to disruption of the organelles, cellular digestion, and decay. Observation of autophagosome membrane proteins Beclin1 and LC3B aggregation in murine pancreatic islets indicates that autophagy restoration may also lead to autophagosome aggregation in murine insulinoma cells. A basal low expression of autophagy markers was detectable in patients affected by pNEN, and, interestingly, the expression of these markers was significantly lower in metastatic pNEN. DISCUSSION/CONCLUSION: Our study highlights that the autophagy functional restoration and prolongation of this catabolic process, mediated by inhibition of deacetylase, is responsible for the reduction of pNEN cells. Prompting of autophagy cell death could be a promising strategy for the therapy of pNEN.

Targeting autophagy in liver cancer.

Autophagy is a catabolic cellular process conserved in animals. It is characterized by the main role of recycling all the non-functional products of the cells. Once, autophagy players detect non-functioning sub-cellular organelles and proteins, they start the so-called nucleation process. The organelles will be surrounded by a double membrane vesicle mainly constituted by endoplasmic reticulum (ER) membrane and autophagy proteins, e.g., MAP1LC3B, Beclin-1, VPS34, Unc-51 like autophagy activating kinase (ULK1) and ubiquitination-related proteins. Then the autophagic membrane will go through an elongation phase involving additional autophagy players. Once the autophagic vesicle is complete, the sub-cellular organelles will be isolated from the rest of the cytosol and driven to the final fusion with lysosomes. Here, the digestion process will end. Alteration and or impairment of autophagy have been shown to be correlated with development of diseases affecting the central nervous system, e.g., Alzheimer and other neurodegenerative diseases. Nonetheless, autophagy defect is responsible for tumorigenesis in blood and solid malignancies, in particular liver cancer. Malignancies of the liver are determined by several genetics and epigenetics mechanisms triggering the up-regulation of survival mechanisms and resistance to cell death. Furthermore, liver cancer could result from pathologic conditions like cirrhosis and fibrosis related to virus infection, aflatoxin, alcohol consumption and high fat diet together with insulin resistance. The role exerted by autophagy in the pathogenesis of the liver and tumor development has been evidenced in recent years. The alteration of autophagy assumes a fundamental role for liver tumorigenesis determining an accumulation of non-functional proteins and organelles that trigger oxidative stress leading to genotoxic stress and gene alterations. Furthermore, the absence of this degradation mechanism could prompt the cells to alter their metabolic status and turn into malignant cells. Interestingly, the heterozygous loss of function of Beclin-1 is able to trigger liver tumorigenesis or even the simple accumulation of proteins caused by the block of the final autolysosome fusion and degradation process is responsible for liver cancer development. This review highlights the importance of targeting the autophagy process in liver cancer in order to restore its function and to promote autophagy-mediated cell demise.