Islets Transplantation at a Crossroads - Need for Urgent Regulatory Update in the United States: Perspective Presented During the Scientific Sessions 2021 at the American Diabetes Association Congress

Clinical islet allotransplantation has been successfully regulated as tissue/organ for transplantation in number of countries and is recognized as a safe and efficacious therapy for selected patients with type 1 diabetes mellitus. However, in the United States, the FDA considers pancreatic islets as a biologic drug, and islet transplantation has not yet shifted from the experimental to the clinical arena for last 20 years. In order to transplant islets, the FDA requires a valid Biological License Application (BLA) in place. The BLA process is costly and lengthy. However, despite the application of drug manufacturing technology and regulations, the final islet product sterility and potency cannot be confirmed, even when islets meet all the predetermined release criteria. Therefore, further regulation of islets as drugs is obsolete and will continue to hinder clinical application of islet transplantation in the US. The Organ Procurement and Transplantation Network together with the United Network for Organ Sharing have developed separately from the FDA and BLA regulatory framework for human organs under the Human Resources & Services Administration to assure safety and efficacy of transplantation. Based on similar biologic characteristics of islets and human organs, we propose inclusion of islets into the existing regulatory framework for organs for transplantation, along with continued FDA oversight for islet processing, as it is for other cell/tissue products exempt from BLA. This approach would reassure islet quality, efficacy and access for Americans with diabetes to this effective procedure.

RETINOBLASTOMA RELATED (RBR) interaction with key factors of the RNA-directed DNA methylation (RdDM) pathway

● Transposable elements and other repetitive elements are silenced by the RNA-directed DNA methylation pathway (RdDM). In RdDM, POLIV-derived transcripts are converted into double stranded RNA (dsRNA) by the activity of RDR2 and subsequently processed into 24 nucleotide short interfering RNAs (24-nt siRNAs) by DCL3. 24-nt siRNAs are recruited by AGO4 and serve as guides to direct AGO4-siRNA complexes to chromatin bound POLV-derived transcripts generated from the template/target DNA. The interaction between POLV, AGO4, DMS3, DRD1, RDM1 and DRM2 promotes DRM2-mediated de novo DNA methylation. In silico exploration of Arabidopsis RBR protein partners revealed that several members of the RdDM pathway contain a motif that confers high affinity binding to RBR, including the largest subunits of POLIV and POLV (NRPD1 and NRPE1), the shared second largest subunit of POLIV and POLV (NRPD/E2), RDR1, RDR2, DCL3, DRM2 and SUVR2. We demonstrate that RBR binds to DRM2, DRD1 and SUVR2. We also report that seedlings from loss-of-function mutants in RdDM and in RBR show similar phenotypes in the root apical meristem. Furthermore, we show that RdDM and SUVR2 targets are up-regulated in the 35S::AmiGO-RBR background. Our results suggest a novel mechanism for RBR function in transcriptional gene silencing based on the interaction with key players of the RdDM pathway and opens several new hypotheses, including the convergence of RBR-DRM2 on the transcriptional control of TEs and several cell/tissue and stage-specific target genes.

FROM BASIC SCIENCES TO CLINICAL APPLICATIONS WITH A SYMPOSIA SERIES: A GLOBAL OUTREACH

Dissemination of research results between scientists usually happens via publications of original papers & review articles published in strictly controlled, peer-reviewed scientific journals. This is certainly a beneficial and useful way of communication, but it does not cover all the needs in scientific interactions. This manuscript aims to highlight the unmet need for effective creative communication among scientists during international conferences and congresses. A brief history of the 35-years symposia series “International Symposia on Cell/Tissue Injury & Cytoprotection/Organoprotection” (ISCTICO) are presented

Microfluidic-Based Oxygen (O2) Sensors for On-Chip Monitoring of Cell, Tissue and Organ Metabolism

Oxygen (O2) quantification is essential for assessing cell metabolism, and its consumption in cell culture is an important indicator of cell viability. Recent advances in microfluidics have made O2 sensing a crucial feature for organ-on-chip (OOC) devices for various biomedical applications. OOC O2 sensors can be categorized, based on their transducer type, into two main groups, optical and electrochemical. In this review, we provide an overview of on-chip O2 sensors integrated with the OOC devices and evaluate their advantages and disadvantages. Recent innovations in optical O2 sensors integrated with OOCs are discussed in four main categories: (i) basic luminescence-based sensors; (ii) microparticle-based sensors; (iii) nano-enabled sensors; and (iv) commercial probes and portable devices. Furthermore, we discuss recent advancements in electrochemical sensors in five main categories: (i) novel configurations in Clark-type sensors; (ii) novel materials (e.g., polymers, O2 scavenging and passivation materials); (iii) nano-enabled electrochemical sensors; (iv) novel designs and fabrication techniques; and (v) commercial and portable electrochemical readouts. Together, this review provides a comprehensive overview of the current advances in the design, fabrication and application of optical and electrochemical O2 sensors.

Reconceptualization of Hormetic Responses in the Frame of Redox Toxicology

Cellular adaptive mechanisms emerging after exposure to low levels of toxic agents or stressful stimuli comprise an important biological feature that has gained considerable scientific interest. Investigations of low-dose exposures to diverse chemical compounds signify the non-linear mode of action in the exposed cell or organism at such dose levels in contrast to the classic detrimental effects induced at higher ones, a phenomenon usually referred to as hormesis. The resulting phenotype is a beneficial effect that tests our physiology within the limits of our homeostatic adaptations. Therefore, doses below the region of adverse responses are of particular interest and are specified as the hormetic gain zone. The manifestation of redox adaptations aiming to prevent from disturbances of redox homeostasis represent an area of particular interest in hormetic responses, observed after exposure not only to stressors but also to compounds of natural origin, such as phytochemicals. Findings from previous studies on several agents demonstrate the heterogeneity of the specific zone in terms of the molecular events occurring. Major factors deeply involved in these biphasic phenomena are the bioactive compound per se, the dose level, the duration of exposure, the cell, tissue or even organ exposed to and, of course, the biomarker examined. In the end, the molecular fate is a complex toxicological event, based on beneficial and detrimental effects, which, however, are poorly understood to date.

Aurora A and AKT Kinase Signaling Associated with Primary Cilia

Dysregulation of kinase signaling is associated with various pathological conditions, including cancer, inflammation, and autoimmunity; consequently, the kinases involved have become major therapeutic targets. While kinase signaling pathways play crucial roles in multiple cellular processes, the precise manner in which their dysregulation contributes to disease is dependent on the context; for example, the cell/tissue type or subcellular localization of the kinase or substrate. Thus, context-selective targeting of dysregulated kinases may serve to increase the therapeutic specificity while reducing off-target adverse effects. Primary cilia are antenna-like structures that extend from the plasma membrane and function by detecting extracellular cues and transducing signals into the cell. Cilia formation and signaling are dynamically regulated through context-dependent mechanisms; as such, dysregulation of primary cilia contributes to disease in a variety of ways. Here, we review the involvement of primary cilia-associated signaling through aurora A and AKT kinases with respect to cancer, obesity, and other ciliopathies.

The Cytotoxic Necrotizing Factors (CNFs)—A Family of Rho GTPase-Activating Bacterial Exotoxins

The cytotoxic necrotizing factors (CNFs) are a family of Rho GTPase-activating single-chain exotoxins that are produced by several Gram-negative pathogenic bacteria. Due to the pleiotropic activities of the targeted Rho GTPases, the CNFs trigger multiple signaling pathways and host cell processes with diverse functional consequences. They influence cytokinesis, tissue integrity, cell barriers, and cell death, as well as the induction of inflammatory and immune cell responses. This has an enormous influence on host–pathogen interactions and the severity of the infection. The present review provides a comprehensive insight into our current knowledge of the modular structure, cell entry mechanisms, and the mode of action of this class of toxins, and describes their influence on the cell, tissue/organ, and systems levels. In addition to their toxic functions, possibilities for their use as drug delivery tool and for therapeutic applications against important illnesses, including nervous system diseases and cancer, have also been identified and are discussed.

Proteomics

Proteomics is a branch of molecular biology that deals with the identification and quantification of proteins in living objects, as well as the analysis of protein functions and their interactions. Proteomics is studied by proteins that are expressed in a given cell, tissue or organism over a period of time (under certain conditions). It is known that information about the primary structure of a protein (the sequence of amino acid residues in a protein) is contained in a structural gene in the form of a codon sequence (genetic code). On the other hand, less than 10% of genes are functionally active (expressed) in the somatic cells of our body. Moreover, a distinct tissue-specific expression of genes is observed. This, in turn, leads to the peculiarities of the qualitative composition of the synthesized proteins in various tissues. No less important is the fact that the total amount of proteins synthesized by our tissues is much greater than the total number of structural genes containing information about their original structure. This phenomenon is explained by the activity of such mechanisms as alternative splicing and a wide variety of post-translational peptide processing pathways (covalent modification of a polypeptide synthesized on the ribosome) in health and disease. Thus, even a brief review of the semantic content of the term "proteomics" indicates an extremely complex system of protein molecules in our body, which plays a fundamental role in maintaining homeostasis and is involved in the formation of adaptive responses in response to adverse changes in the internal and external environment.

Design and Fabrication of Low-Cost Microfluidic Chips and Microfluidic Routing System for Reconfigurable Multi-(Organ-on-a-Chip) Assembly

A low-cost, versatile, and reconfigurable fluidic routing system and chip assembly have been fabricated and tested. The platform and its accessories were fabricated in-house without the need for costly and specialized equipment nor specific expertise. An agarose-based artificial membrane was integrated into the chips and employed to test the chip-to-chip communication in various configurations. Various chip assemblies were constructed and tested which demonstrate the versatile utility of the fluidic routing system that enables the custom design of the chip-to-chip communication and the possibility of fitting a variety of (organ-on-a-chip)-based biological models with multicell architectures. The reconfigurable chip assembly would enable selective linking/isolating the desired chip/compartment, hence allowing the study of the contribution of specific cell/tissue within the in vitro models.