Phenotypes and ontogeny of senescent hepatic stellate cells in metabolic dysfunction-associated steatohepatitis.

BACKGROUND & AIMS: Hepatic stellate cells (HSCs) are the key drivers of fibrosis in metabolic dysfunction-associated steatohepatitis (MASH), the fastest growing cause of hepatocellular carcinoma (HCC) worldwide. HSCs are heterogenous, and a senescent subset of HSCs is implicated in hepatic fibrosis and HCC. Administration of anti-uPAR (urokinase-type plasminogen activator receptor) CAR T cells has been shown to deplete senescent HSCs and attenuate fibrosis in murine models. However, the comprehensive features of senescent HSCs in MASH, as well as their cellular ontogeny have not been characterized; hence, we aimed to comprehensively characterize and define the origin of HSCs in human and murine MASH. METHODS: To comprehensively characterize the phenotype and ontogeny of senescent HSCs in human and murine MASH, we integrated senescence-associated beta galactosidase activity with immunostaining, flow cytometry and single-nucleus RNA sequencing (snRNAseq). We integrated the immunohistochemical profile with a senescence score applied to snRNAseq data to characterize senescent HSCs and mapped the evolution of uPAR expression in MASH. RESULTS: Using pseudotime trajectory analysis, we establish that senescent HSCs arise from activated HSCs. While uPAR is expressed in MASH, the magnitude and cell-specificity of its expression evolve with disease stage. In early disease, uPAR is more specific to activated and senescent HSCs, while it is also expressed by myeloid-lineage cells, including Trem2+ macrophages and myeloid-derived suppressor cells, in late disease. Furthermore, we identify novel surface proteins expressed on senescent HSCs in human and murine MASH that could be exploited as therapeutic targets. CONCLUSIONS: These data define features of HSC senescence in human and murine MASH, establishing an important blueprint to target these cells as part of future antifibrotic therapies. IMPACT AND IMPLICATIONS: Hepatic stellate cells (HSCs) are the primary drivers of scarring in chronic liver diseases. As injury develops, a subset of HSCs become senescent; these cells are non-proliferative and pro-inflammatory, thereby contributing to worsening liver injury. Here we show that senescent HSCs are expanded in MASH (metabolic dysfunction-associated steatohepatitis) in humans and mice, and we trace their cellular origin from the activated HSC subset. We further characterize expression of uPAR (urokinase plasminogen activated receptor), a protein that marks senescent HSCs, and report that uPAR is also expressed by activated HSCs in early injury, and in immune cells as liver injury advances. We have integrated high-resolution single-nucleus RNA sequencing with immunostaining and flow cytometry to identify five other novel proteins expressed by senescent HSCs, including mannose receptor CD206, which will facilitate future therapeutic development.

Aptamer-based photonic crystals enable ultra-trace detection of staphylococcal enterotoxin B without labels.

Staphylococcal enterotoxin B (SEB) is one of the most common serotypes in staphylococcal food-poisoning cases. A rapid, sensitive, and simple method for SEB detection is crucial for public health. A photonic crystal (PC) sensing material for label-free detection of ultra-trace SEB was proposed in this study. Gold nanoparticle-doped silica microspheres were stacked to form an opal PC through self-assembly, and SEB aptamers, as the recognition element, were modified onto the PC. When the target protein of SEB came in contact with the PC sensing material, the reflection peak intensity of PCs decreased accordingly. The detection range was 1 × 10-6 to 1 ng mL-1, and the detection limit was 0.103 × 10-6 ng mL-1. Furthermore, the PC sensing material had great specificity and accuracy, which can be used for real sample monitoring. This PC sensing material achieved ultra-sensitive detection, which did not involve complicated preparation processes and reporter labelling.

Abundance and diversity of microbial arsenic biotransformation genes in the sludge of full-scale anaerobic digesters from a municipal wastewater treatment plant.

Arsenic (As) is a potential contaminant in sewage sludge that may affect waste treatment and limit the use of these waste materials as soil amendments. Anaerobic digestion (AD) is an important and effective process for the treatment of sewage sludge and the chemical speciation of As is particularly important in sludge AD. However, the biotransformation genes of As in sludge during AD has not been fully explored. In this study, the influent and effluent sludge of anaerobic digester in a wastewater treatment plant (WWTP) was collected to investigate the species transformations of As, the abundance and diversity of As biotransformation genes was explored by real-time PCR (qPCR) and metagenomic sequencing, separately. The results showed that arsenite [As(III)] and arsenate [As(V)] were predominant in the influent sludge, whereas the relative abundance of monomethylarsenic acid (MMA) increased by 25.7% after digestion. As biotransformation genes were highly abundant, and the As(III) S-adenosylmethionine methyltransferase (arsM) gene was the predominant which significantly increased after AD by qPCR analysis. Metagenomic analysis indicated that the diversity of the arsM-like sequences also increased significantly after AD. Most of the arsM-like sequences in all the influent and effluent sludge samples were related to Bacteroidetes and Alphaproteobacteria. Furthermore, co-occurrence network analysis indicated a strong correlation between the microbial communities and As. This study provides a direct and reliable reference on As biotransformation genes and microbial community in the AD of sludge.

Single injection of IL-12 coacervate as an effective therapy against B16-F10 melanoma in mice.

Melanoma is the deadliest type of skin cancer with one of the fastest increasing incidence rates among solid tumors. The use of checkpoint inhibitors (e.g. αPD-1 antibody) has recently emerged as a viable alternative to conventional modes of therapy. However, increasing evidence points towards the need for a tumor priming step to improve intratumoral immune cell infiltration. IL-12 is an immune-activating cytokine with such potential and was explored in earlier clinical trials as a highly concentrated systemic infusion. This unfortunately led to severe adverse effects. From this perspective, the localization and gradual release of such a potent immunotherapeutic agent in the tumor microenvironment is desired. This manuscript reports the use of a heparin-based complex coacervate to deliver IL-12, in which heparin-binding motifs on IL-12 allow for its effective encapsulation. IL-12-encapsulated complex coacervates significantly improved the bioactivity of IL-12 and provided protection from proteolytic cleavage in-vitro. Indeed, a single injection of IL-12 coacervate significantly inhibits the in-vivo growth of treated and untreated, contralateral tumor growth in a syngeneic B16F10 mouse melanoma model. Furthermore, tumors in mice receiving IL-12 complex coacervate treatment displayed increased infiltration by natural killer (NK) cells and CD8α+ T cells, and a decreased presence of CD4+Foxp3+ regulatory T cells. This study provides proof-of-concept data supporting the use of complex coacervates for sustained delivery of immunostimulatory proteins as an effective therapeutic strategy against disseminated tumors.

Performance of the ANAMMOX process using multi- and single-fed upflow anaerobic sludge blanket reactors.

The performance of the ANAMMOX process was investigated in two identical laboratory-scale multi- and single-fed upflow anaerobic sludge blanket (UASB) reactors (denoted R1 and R0) at different hydraulic residence times (HRTs) varying from 2.06 to 1.52 h and NH4(+)-N inf concentrations ranging from 70 to 266 mg L(-1). The substrate removal efficiencies of both reactors decreased as HRT decreased and NH4(+)-N inf increased. The kinetics of these reactions were analyzed, and the Stover-Kincannon model was appropriate to describe the process kinetics of the reactors. In addition, an empirical model incorporating the influent substrate concentration and HRT adequately described R1. Shock experiments were conducted in which the reactors were subjected to transient shock loads. The results showed that the operation of R1 was more stable than that of R0, especially in response to the substrate shocks. Subsequently, the properties of the ANAMMOX granules and the effects of the feeding protocol on those properties were investigated.