The effect of protocol for disinfection of extracted teeth recommended by center for disease control (CDC) on microhardness of enamel and dentin

Background: According to the guideline of the United States center for disease control (CDC), the extracted teeth should be sterilized by autoclaving or storage in 10% formalin before using for educational or research purposes. The objective of this study was to evaluate the effect of this protocol on microhardness of dentin and enamel. Material and Methods: Thirty extracted single-root teeth were used in this study. The crowns were resected, and the roots were longitudinally sectioned into two halves. The Vickers microhardness (VHN) of specimens was measured on polished canal dentin and buccal enamel surfaces. The crowns were randomly divided into three groups (n=10). Group 1 and 2 were sterilized using autoclave and formalin, respectively while group 3 (control) was stored in synthetic tissue fluid. The root halves were also randomly divided into 3 groups (n=20) which were treated as mentioned above for crown samples. Following sterilization, VHN of samples was measured again. ANOVA and paired samples t-tests were used to analyze the data. Results: Autoclaving caused a significant reduction in microhardness of dentin ( P <0.001, 12.04% decreases in VHN). However, there were no significant differences for before and after sterilization within other groups. Conclusions: Based on the results of this study, the CDC protocol is recommended in studies related to enamel microhardness. However, Autoclaving is not an appropriate sterilization method in studies related to dentin microhardness. In these studies, two-week immersion in 10% formalin is recommended

Characterisation and induction of tissue-resident gamma delta T-cells to target hepatocellular carcinoma

Immunotherapy is now the standard of care for advanced hepatocellular carcinoma (HCC), yet many patients fail to respond. A major unmet goal is the boosting of T-cells with both strong HCC reactivity and the protective advantages of tissue-resident memory T-cells (TRM). Here, we show that higher intratumoural frequencies of γδ T-cells, which have potential for HLA-unrestricted tumour reactivity, associate with enhanced HCC patient survival. We demonstrate that γδ T-cells exhibit bona fide tissue-residency in human liver and HCC, with γδTRM showing no egress from hepatic vasculature, persistence for >10 years and superior anti-tumour cytokine production. The Vγ9Vδ2 T-cell subset is selectively depleted in HCC but can efficiently target HCC cell lines sensitised to accumulate isopentenyl-pyrophosphate by the aminobisphosphonate Zoledronic acid. Aminobisphosphonate-based expansion of peripheral Vγ9Vδ2 T-cells recapitulates a TRM phenotype and boosts cytotoxic potential. Thus, our data suggest more universally effective HCC immunotherapy may be achieved by combining aminobisphosphonates to induce Vγ9Vδ2TRM capable of replenishing the depleted pool, with additional intratumoural delivery to sensitise HCC to Vγ9Vδ2TRM-based targeting

Isolation of human intrahepatic leukocytes for phenotypic and functional characterization by flow cytometry

With the growing appreciation of tissue-resident immunity, studying tissue-specific immune cells contributing to both homeostasis and disease is imperative. Here, we provide a protocol for the isolation of human intrahepatic leukocytes (IHL) maximizing viability, purity, and yield. Our protocol is scalable by tissue weight, allowing for reproducible and efficient IHL liberation suitable for functional characterization, cell isolation, and profiling by flow (or mass) cytometry. Furthermore, we provide a "guide" to determine an expected IHL yield per gram of tissue processed. For complete details on the use and execution of this protocol, please refer to Stegmann et al. (2016), Pallett et al. (2017), Easom et al. (2018), Swadling et al. (2020), Pallett et al. (2020), and Zakeri et al. (2022)

Targeting human Acyl-CoA:cholesterol acyltransferase as a dual viral and T cell metabolic checkpoint

Determining divergent metabolic requirements of T cells, and the viruses and tumours they fail to combat, could provide new therapeutic checkpoints. Inhibition of acyl-CoA:cholesterol acyltransferase (ACAT) has direct anti-carcinogenic activity. Here, we show that ACAT inhibition has antiviral activity against hepatitis B (HBV), as well as boosting protective anti-HBV and anti-hepatocellular carcinoma (HCC) T cells. ACAT inhibition reduces CD8+ T cell neutral lipid droplets and promotes lipid microdomains, enhancing TCR signalling and TCR-independent bioenergetics. Dysfunctional HBV- and HCC-specific T cells are rescued by ACAT inhibitors directly ex vivo from human liver and tumour tissue respectively, including tissue-resident responses. ACAT inhibition enhances in vitro responsiveness of HBV-specific CD8+ T cells to PD-1 blockade and increases the functional avidity of TCR-gene-modified T cells. Finally, ACAT regulates HBV particle genesis in vitro, with inhibitors reducing both virions and subviral particles. Thus, ACAT inhibition provides a paradigm of a metabolic checkpoint able to constrain tumours and viruses but rescue exhausted T cells, rendering it an attractive therapeutic target for the functional cure of HBV and HBV-related HCC

Human liver memory CD8(+) T cells use autophagy for tissue residence

Tissue-resident memory T cells have critical roles in long-term pathogen and tumor immune surveillance in the liver. We investigate the role of autophagy in equipping human memory T cells to acquire tissue residence and maintain functionality in the immunosuppressive liver environment. By performing ex vivo staining of freshly isolated cells from human liver tissue, we find that an increased rate of basal autophagy is a hallmark of intrahepatic lymphocytes, particularly liver-resident CD8(+) T cells. CD8(+) T cells with increased autophagy are those best able to proliferate and mediate cytotoxicity and cytokine production. Conversely, blocking autophagy induction results in the accumulation of depolarized mitochondria, a feature of exhausted T cells. Primary hepatic stellate cells or the prototypic hepatic cytokine interleukin (IL)-15 induce autophagy in parallel with tissue-homing/retention markers. Inhibition of T cell autophagy abrogates tissue-residence programming. Thus, upregulation of autophagy adapts CD8(+) T cells to combat mitochondrial depolarization, optimize functionality, and acquire tissue residence

An overview of progress in electrolytes for secondary zinc-air batteries and other storage systems based on zinc

The revived interest and research on the development of novel energy storage systems with exceptional inherent safety, environmentally benign and low cost for integration in large scale electricity grid and electric vehicles is now driven by the global energy policies. Within various technical challenges yet to be resolved and despite extensive studies, the low cycle life of the zinc anode is still hindering the implementation of rechargeable zinc batteries at industrial scale. This review presents an extensive overview of electrolytes for rechargeable zinc batteries in relation to the anode issues which are closely affected by the electrolyte nature. Widely studied aqueous electrolytes, from alkaline to acidic pH, as well as non-aqueous systems including polymeric and room temperature ionic liquids are reported. References from early rechargeable Zn-air research to recent results on novel Zn hybrid systems have been analyzed. The ambition is to identify the challenges of the electrolyte system and to compile the proposed improvements and solutions. Ultimately, all the technologies based on zinc, including the more recently proposed novel zinc hybrid batteries combining the strong points of lithium-ion, redox-flow and metal-air systems, can benefit from this compilation in order to improve secondary zinc based batteries performance.Basque Country University (ZABALDUZ2012 program), and the Basque Country Government (Project: CIC energiGUNÉ16 of the ELKARTEK program) and the European Commission through the project ZAS: “Zinc Air Secondary innovative nanotech based batteries for efficient energy storage” (Grant Agreement 646186

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Dual Role of Toll-like Receptors in Human and Experimental Asthma Models

Asthma is a chronic airway inflammatory disease that is influenced by the interplay between genetic factors and exposure to environmental allergens, microbes, or microbial products where toll-like receptors (TLRs) play a pivotal role. TLRs recognize a wide range of microbial or endogenous molecules as well as airborne environmental allergens and act as adjuvants that influence positively or negatively allergic sensitization. TLRs are qualitatively and differentially expressed on hematopoietic and non-hematopoietic stromal or structural airway cells that when activated by TLRs agonists exert an immune-modulatory role in asthma development. Therefore, understanding mechanisms and pathways by which TLRs orchestrate asthma outcomes may offer new strategies to control the disease. Here, we aim to review and critically discuss the role of TLRs in human asthma and murine models of allergic airway inflammation, highlighting the complexity of TLRs function in development, exacerbation, or control of airway allergic inflammation