Biology subjects

Cohen, A. J.

Publications and source records attributed to Cohen, A. J..

4 recordsLinked to original sources

bioRxiv2025

EVALUATION OF STAINING FOR FIBRONECTIN ISOFORMS IN LIVER DISEASES

Fibronectin is an ubiquitous extracellular matrix protein, that is produced by the hepatocytes contributing to circulating fibronectin, and by the hepatic stellate cells contributing to fibrotic bundles. Fibronectin is required for the accumulation of collagen type I, the main fibrotic collagen in liver disease. It is presumed that cellular fibronectins including EDA-containing, EDB-containing and oncofetal fibronectin are the ones localized to the fibrotic bundles, since these isoforms are increased in the presence of fibrosis and cirrhosis in patients with liver disease. The aim of this work was therefore to evaluate the distribution of the various isoforms and determine whether they are detected in the hepatocytes too or whether they are indeed limited in their expression to fibrotic bundles. Total fibronectin was detected in hepatocytes particularly in healthy liver tissue. In advanced disease, fibronectin staining seemed diminished. All three isoforms, EDA-, EDB-, and oFN were detected in hepatocytes or the fibrotic bundles, with EDB being the most abundant. Interestingly, the isoforms did not fully account for all of fibronectin. Staining patterns confirmed that the three isoforms are not limited to the fibrotic bundles, but are also found in the hepatocytes. This contradicts the common notion that cellular fibronectins precipitate and are localized in the matrix, and explains why they can be measured in the circulation. Finally, there is some convergence between the presence of the isoforms in the fibrotic bundles and the elevation of their levels in the blood stream at least in chronic hepatitis C and cholestatic liver disease. This is in line with a high degree of complexity of fibronectin distribution and effects.

cell biology↗

bioRxiv2023

Development and Implementation of Novel Chatbot-based Genomic Research Consent

ObjectiveTo conduct a retrospective analysis comparing traditional human-based consenting to an automated chat-based consenting process. Materials and MethodsWe developed a new chat-based consent using our IRB-approved consent forms. We leveraged a previously developed platform (Gia(R), or "Genetic Information Assistant") to deliver the chat content to candidate participants. The content included information about the study, educational information, and a quiz to assess understanding. We analyzed 144 families referred to our study during a 6-month time period. A total of 37 families completed consent using the traditional process, while 35 families completed consent using Gia. ResultsEngagement rates were similar between both consenting methods. The median length of the consent conversation was shorter for Gia users compared to traditional (44 vs. 76 minutes). Additionally, the total time from referral to consent completion was faster with Gia (5 vs. 16 days). Within Gia, understanding was assessed with a 10-question quiz that most participants (96%) passed. Feedback about the chat consent indicated that 86% of participants had a positive experience. DiscussionUsing Gia resulted in time savings for both the participant and study staff. The chatbot enables studies to reach more potential candidates. We identified five key features related to human-centered design for developing a consent chat. ConclusionThis analysis suggests that it is feasible to use an automated chatbot to scale obtaining informed consent for a genomics research study. We further identify a number of advantages when using a chatbot.

genetics↗

bioRxiv2022

Dual function of the O-antigen WaaL ligase of Aggregatibacter actinomycetemcomitans

Protein glycosylation is critical to the quaternary structure and collagen binding activity of the extracellular matrix protein adhesin A (EmaA) associated with Aggregatibacter actinomycetemcomitans. The glycosylation of this large, trimeric autotransporter adhesin is postulated to be mediated by WaaL, an enzyme with the canonical function to ligate the O-polysaccharide (O-PS) antigen with a terminal sugar of the lipid A-core oligosaccharide of lipopolysaccharide (LPS). In this study, we have determined that the Escherichia coli waaL ortholog (rflA) does not restore collagen binding of a waaL mutant strain of A. actinomycetemcomitans but does restore O-PS ligase activity following transformation of a plasmid expressing waaL. Therefore, a heterologous E. coli expression system was developed constituted of two independently replicating plasmids expressing either waaL or emaA of A. actinomycetemcomitans to directly demonstrate the necessity of ligase activity for EmaA collagen binding. Proper expression of the protein encoded by each plasmid was characterized, and the individually transformed strains did not promote collagen binding. However, co-expression of the two plasmids resulted in a strain with a significant increase in collagen binding activity and a change in the biochemical properties of the protein. These results provide additional data supporting the novel hypothesis that the WaaL ligase of A. actinomycetemcomitans shares a dual role as a ligase in LPS biosynthesis and is required for collagen binding activity of EmaA. ImportanceThe human oral pathogen A. actinomycetemcomitans is a causative agent of periodontal and several systemic diseases. The organism expresses an adhesin, EmaA, important for the colonization of this pathobiont via collagen binding and biofilm formation. EmaA is suggested to be modified with sugars and the modification is mediated using the same enzymes involved in lipopolysaccharide (LPS) biosynthesis. In this study, evidence is presented which suggests that the WaaL ligase, the enzyme that ligates the O-polysaccharide (O-PS) antigen with a terminal sugar of the lipid A-core oligosaccharide of LPS, is required for the collagen binding activity of EmaA. This finding represents a new paradigm for the posttranslational modification of this type of autotransporter protein.

microbiology↗

bioRxiv2022

A dual role for CRTH2 in acute lung injury

Acute respiratory distress syndrome (ARDS) is a life-threatening clinical condition defined by rapid-onset respiratory failure following acute lung injury (ALI). The high mortality rate and rising incidence of ARDS due to COVID-19 make it an important research priority. Here we sought to investigate the role of chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) in ARDS. CRTH2 is a G protein-coupled receptor best studied in the context of type 2 immunity, but it also exerts effects on neutrophilic inflammation. To evaluate its role in mouse models of ARDS, we first examined its expression pattern on murine neutrophils. We found it is expressed on neutrophils, but only after extravasation into the lung. Next, we showed that CRTH2 expression on extravasated lung neutrophils promotes cell survival, as genetic deletion of CRTH2 and pharmacologic inhibition of CRTH2 using fevipiprant both led to increased apoptosis in vitro. We then evaluated the role of CRTH2 in vivo using a murine model of LPS-induced ALI. In line with the pro-inflammatory effects of CRTH2 in vitro, we observed improvement of lung injury in CRTH2-deficient mice in terms of vascular leak, weight loss and survival after LPS administration. However, neutrophilic inflammation was elevated, not suppressed in the CRTH2 KO. This finding indicated a second mechanism offsetting the pro-survival effect of CRTH2 on neutrophils. Bulk RNAseq of lung tissue indicated impairments in type 2 immune signaling in the CRTH2 KO, and qPCR and ELISA confirmed downregulation of IL-4, which is known to suppress neutrophilic inflammation. Thus, CRTH2 may play a dual role in ALI, directly promoting neutrophil cell survival, but indirectly suppressing neutrophil effector function via IL-4.

immunology↗