The role of cytology in patients undergoing pressurized intraperitoneal aerosol chemotherapy (PIPAC) treatment for peritoneal carcinomatosis.

Cytology of ascites or peritoneal washing is a routine part of staging of peritoneal metastases (PM). We aim to determine value of cytology in patients undergoing pressurized intraperitoneal aerosol chemotherapy (PIPAC). Single-center retrospective cohort study included consecutive patients having PIPAC for PM of different primary between January 2015 and January 2020. A total of 75 patients (median 63 years (IQR 51-70), 67 % female) underwent a total of 144 PIPAC. At PIPAC 1 59 % patients had positive and 41 % patients had negative cytology. Patients with negative and positive cytology only differed in terms of symptoms of ascites (16% vs. 39 % respectively, p=0.04), median ascites volume (100 vs. 0 mL, p=0.01) and median PCI (9 vs. 19, p<0.01). Among 20 patients who completed 3 PIPACs (per protocol), cytology changed in one from positive to negative, and in two from negative to positive. Median overall survival was 30.9 months in the per protocol group and 12.9 months in patients having <3 PIPACs (=0.519). Positive cytology under PIPAC treatment is more frequently encountered in patients with higher PCI and symptomatic ascites. Cytoversion was rarely observed and cytology status had no impact on treatment decisions in this cohort

Cannabidiol causes activated hepatic stellate cell death through a mechanism of endoplasmic reticulum stress-induced apoptosis

The major cellular event in the development and progression of liver fibrosis is the activation of hepatic stellate cells (HSCs). Activated HSCs proliferate and produce excess collagen, leading to accumulation of scar matrix and fibrotic liver. As such, the induction of activated HSC death has been proposed as a means to achieve resolution of liver fibrosis. Here we demonstrate that cannabidiol (CBD), a major non-psychoactive component of the plant Cannabis sativa, induces apoptosis in activated HSCs through a cannabinoid receptor-independent mechanism. CBD elicits an endoplasmic reticulum (ER) stress response, characterized by changes in ER morphology and the initiation of RNA-dependent protein kinase-like ER kinase-, activating transcription factor-6-, and inositol-requiring ER-to-nucleus signal kinase-1 (IRE1)-mediated signaling cascades. Furthermore, CBD induces downstream activation of the pro-apoptotic IRE1/ASK1/c-Jun N-terminal kinase pathway, leading to HSC death. Importantly, we show that this mechanism of CBD-induced ER stress-mediated apoptosis is specific to activated HSCs, as it occurs in activated human and rat HSC lines, and in primary in vivo-activated mouse HSCs, but not in quiescent HSCs or primary hepatocytes from rat. Finally, we provide evidence that the elevated basal level of ER stress in activated HSCs has a role in their susceptibility to the pro-apoptotic effect of CBD. We propose that CBD, by selectively inducing death of activated HSCs, represents a potential therapeutic agent for the treatment of liver fibrosis

Hepatitis C Virus Induces the Cannabinoid Receptor 1

BACKGROUND: Activation of hepatic CB(1) receptors (CB(1)) is associated with steatosis and fibrosis in experimental forms of liver disease. However, CB(1) expression has not been assessed in patients with chronic hepatitis C (CHC), a disease associated with insulin resistance, steatosis and metabolic disturbance. We aimed to determine the importance and explore the associations of CB(1) expression in CHC. METHODS: CB(1) receptor mRNA was measured by real time quantitative PCR on extracted liver tissue from 88 patients with CHC (genotypes 1 and 3), 12 controls and 10 patients with chronic hepatitis B (CHB). The Huh7/JFH1 Hepatitis C virus (HCV) cell culture model was used to validate results. PRINCIPAL FINDINGS: CB(1) was expressed in all patients with CHC and levels were 6-fold higher than in controls (P<0.001). CB(1) expression increased with fibrosis stage, with cirrhotics having up to a 2 fold up-regulation compared to those with low fibrosis stage (p<0.05). Even in mild CHC with no steatosis (F0-1), CB(1) levels remained substantially greater than in controls (p<0.001) and in those with mild CHB (F0-1; p<0.001). Huh7 cells infected with JFH-1 HCV showed an 8-fold upregulation of CB(1), and CB(1) expression directly correlated with the percentage of cells infected over time, suggesting that CB(1) is an HCV inducible gene. While HCV structural proteins appear essential for CB(1) induction, there was no core genotype specific difference in CB(1) expression. CB(1) significantly increased with steatosis grade, primarily driven by patients with genotype 3 CHC. In genotype 3 patients, CB(1) correlated with SREBP-1c and its downstream target FASN (SREBP-1c; R=0.37, FASN; R=0.39, p<0.05 for both). CONCLUSIONS/SIGNIFICANCE: CB(1) is up-regulated in CHC and is associated with increased steatosis in genotype 3. It is induced by the hepatitis C virus

Identification and Structural Characterization of a New Three-Finger Toxin Hemachatoxin from Hemachatus haemachatus Venom

10.1371/journal.pone.0048112PLoS ONE710

Obesity, the Endocannabinoid System, and Bias Arising from Pharmaceutical Sponsorship

Previous research has shown that academic physicians conflicted by funding from the pharmaceutical industry have corrupted evidence based medicine and helped enlarge the market for drugs. Physicians made pharmaceutical-friendly statements, engaged in disease mongering, and signed biased review articles ghost-authored by corporate employees. This paper tested the hypothesis that bias affects review articles regarding rimonabant, an anti-obesity drug that blocks the central cannabinoid receptor.A MEDLINE search was performed for rimonabant review articles, limited to articles authored by USA physicians who served as consultants for the company that manufactures rimonabant. Extracted articles were examined for industry-friendly bias, identified by three methods: analysis with a validated instrument for monitoring bias in continuing medical education (CME); analysis for bias defined as statements that ran contrary to external evidence; and a tally of misrepresentations about the endocannabinoid system. Eight review articles were identified, but only three disclosed authors' financial conflicts of interest, despite easily accessible information to the contrary. The Takhar CME bias instrument demonstrated statistically significant bias in all the review articles. Biased statements that were nearly identical reappeared in the articles, including disease mongering, exaggerating rimonabant's efficacy and safety, lack of criticisms regarding rimonabant clinical trials, and speculations about surrogate markers stated as facts. Distinctive and identical misrepresentations regarding the endocannabinoid system also reappeared in articles by different authors.The findings are characteristic of bias that arises from financial conflicts of interest, and suggestive of ghostwriting by a common author. Resolutions for this scenario are proposed

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field