Cathepsin L Inhibition Prevents Murine Autoimmune Diabetes via Suppression of CD8+ T Cell Activity

Background: Type 1 diabetes (T1D) is an autoimmune disease resulting from defects in central and peripheral tolerance and characterized by T cell-mediated destruction of islet b cells. To determine whether specific lysosomal proteases might influence the outcome of a T cell–mediated autoimmune response, we examined the functional significance of cathepsin inhibition on autoimmune T1D-prone non-obese diabetic (NOD) mice. Methods and Findings: Here it was found that specific inhibition of cathepsin L affords strong protection from cyclophosphamide (CY)-induced insulitis and diabetes of NOD mice at the advanced stage of CD8 + T cell infiltration via inhibiting granzyme activity. It was discovered that cathepsin L inhibition prevents cytotoxic activity of CD8 + T cells in the pancreatic islets through controlling dipeptidyl peptidase I activity. Moreover, the gene targeting for cathepsin L with application of in vivo siRNA administration successfully prevented CY-induced diabetes of NOD mice. Finally, cathepsin L mRNA expression of peripheral CD8 + T cells from NOD mice developing spontaneous T1D was significantly increased compared with that from control mice. Conclusions: Our results identified a novel function of cathepsin L as an enzyme whose activity is essential for the progression of CD8 + T cell-mediated autoimmune diabetes, and inhibition of cathepsin L as a powerful therapeutic strateg

Search for dark matter produced in association with bottom or top quarks in √s = 13 TeV pp collisions with the ATLAS detector

A search for weakly interacting massive particle dark matter produced in association with bottom or top quarks is presented. Final states containing third-generation quarks and miss- ing transverse momentum are considered. The analysis uses 36.1 fb−1 of proton–proton collision data recorded by the ATLAS experiment at √s = 13 TeV in 2015 and 2016. No significant excess of events above the estimated backgrounds is observed. The results are in- terpreted in the framework of simplified models of spin-0 dark-matter mediators. For colour- neutral spin-0 mediators produced in association with top quarks and decaying into a pair of dark-matter particles, mediator masses below 50 GeV are excluded assuming a dark-matter candidate mass of 1 GeV and unitary couplings. For scalar and pseudoscalar mediators produced in association with bottom quarks, the search sets limits on the production cross- section of 300 times the predicted rate for mediators with masses between 10 and 50 GeV and assuming a dark-matter mass of 1 GeV and unitary coupling. Constraints on colour- charged scalar simplified models are also presented. Assuming a dark-matter particle mass of 35 GeV, mediator particles with mass below 1.1 TeV are excluded for couplings yielding a dark-matter relic density consistent with measurements

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

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

CROATIAN GUIDELINES FOR THE PHARMACOTHERAPY OF TYPE 2 DIABETES

Uvod: Hrvatsko društvo za dijabetes i bolesti metabolizma Hrvatskoga liječničkog zbora izradilo je 2011. godine prve nacionalne smjernice o prehrani, edukaciji i samokontroli te farmakološkom liječenju šećerne bolesti tipa 2. Sukladno povećanom broju dostupnih lijekova te novim spoznajama o učinkovitosti i sigurnosti primjene već uključenih lijekova, pokazala se potreba za obnovom postojećih smjernica za farmakološko liječenje šećerne bolesti tipa 2 u Republici ­Hrvatskoj. Sudionici: Kao koautori Smjernica navedeni su svi članovi Hrvatskog društva za dijabetes i bolesti metabolizma Hrvatskoga liječničkog zbora, kao i ostalih uključenih stručnih društava, koji su svojim komentarima i prijedlozima pridonijeli izradi Smjernica. Dokazi: Ove su Smjernice utemeljene na dokazima, prema sustavu GRADE (engl. Grading of Recommendations, Assessment, Development and Evaluation) koji uz razinu dokaza opisuje i snagu preporuke. Zaključci: Individualan pristup temeljen na fiziološkim principima regulacije glikemije nuždan je u liječenju osoba sa šećernom bolesti. Ciljeve liječenja i odabir medikamentne terapije treba prilagoditi oboljeloj osobi, uzimajući u obzir životnu dob, trajanje bolesti, očekivano trajanje života, rizik od hipoglikemije, komorbiditete, razvijene vaskularne i ostale komplikacije, kao i ostale čimbenike. Zbog svega navedenoga od nacionalnog je interesa imati praktične, racionalne i provedive smjernice za farmakološko liječenje šećerne bolesti tipa 2.Introduction: The Croatian Association for Diabetes and Metabolic Disorders of the Croatian Medical Association has issued in 2011 the first national guidelines for the nutrition, education, self-control, and pharmacotherapy of ­diabetes type 2. According to the increased number of available medicines and new evidence related to the effectiveness and safety of medicines already involved in the therapy there was a need for update of the existing guidelines for the ­pharmacotherapy of type 2 diabetes in the Republic of Croatia. Participants: as co-authors of the Guidelines there are listed all members of the Croatian Association for Diabetes and Metabolic Diseases, as well as other representatives of professional societies within the Croatian Medical Association, who have contributed with comments and suggestions to the development of the Guidelines. Evidence: These guidelines are evidence-based, according to the GRADE system (eng. Grading of Recommendations, Assessment, Development and Evaluation), which describes the level of evidence and strength of recommendations. Conclusions: An individual patient approach based on physiological principles in blood glucose control is essential for diabetes’ patients management. Glycemic targets and selection of the pharmacological agents should be tailored to the patient, taking into account the age, duration of disease, life expectancy, risk of hypoglycemia, comorbidities, developed vascular and other complications as well as other factors. Because of all this, is of national interest to have a practical, rational and applicable guidelines for the pharmacotherapy of type 2 diabetes

Hrvatske smjernice za farmakološko liječenje šećerne bolesti tipa 2 [Croatian guidelines for the pharmacotherapy of type 2 diabetes]

Introduction: The Croatian Association for Diabetes and Metabolic Disorders of the Croatian Medical Association has issued in 2011 the first national guidelines for the nutrition, education, self-control, and pharmacotherapy of diabetes type 2. According to the increased number of available medicines and new evidence related to the effectiveness and safety of medicines already involved in the therapy there was a need for update of the existing guidelines for the pharmacotherapy of type 2 diabetes in the Republic of Croatia. Participants: as co-authors of the Guidelines there are listed all members of the Croatian Association for Diabetes and Metabolic Diseases, as well as other representatives of professional societies within the Croatian Medical Association, who have contributed with comments and suggestions to the development of the Guidelines. Evidence: These guidelines are evidence-based, according to the GRADE system (eng. Grading of Recommendations, Assessment, Development and Evaluation), which describes the level of evidence and strength of recommendations. Conclusions: An individual patient approach based on physiological principles in blood glucose control is essential for diabetes' patients management. Glycemic targets and selection of the pharmacological agents should be tailored to the patient, taking into account the age, duration of disease, life expectancy, risk of hypoglyce- mia, comorbidities, developed vascular and other complications as well as other factors. Because of all this, is of national interest to have a practical, rational and applicable guidelines for the pharmacotherapy of type 2 diabetes