Protocol for the characterization of the pancreatic tumor microenvironment using organoid-derived mouse models and single-nuclei RNA sequencing.

Single-nuclei RNA sequencing (snRNA-seq) allows for obtaining gene expression profiles from frozen or hard-to-dissociate tissues at the single-nuclei level. Here, we describe a protocol to obtain snRNA-seq data of pancreatic tumors from orthotopically grafted organoid-derived mouse models. We provide details on the establishment of these mouse models, the isolation of single nuclei from pancreatic tumors, and the analysis of the snRNA-seq datasets. For complete details on the use and execution of this protocol, please refer to Mucciolo et al.1.

Cross-talk between ILC2 and Gata3high Tregs locally constrains adaptive type 2 immunity.

Regulatory T cells (Tregs) control adaptive immunity and restrain type 2 inflammation in allergic disease. Interleukin-33 promotes the expansion of tissue-resident Tregs and group 2 innate lymphoid cells (ILC2s); however, how Tregs locally coordinate their function within the inflammatory niche is not understood. Here, we show that ILC2s are critical orchestrators of Treg function. Using spatial, cellular, and molecular profiling of the type 2 inflamed niche, we found that ILC2s and Tregs engage in a direct (OX40L-OX40) and chemotaxis-dependent (CCL1-CCR8) cellular dialogue that enforces the local accumulation of Gata3high Tregs, which are transcriptionally and functionally adapted to the type 2 environment. Genetic interruption of ILC2-Treg communication resulted in uncontrolled type 2 lung inflammation after allergen exposure. Mechanistically, we found that Gata3high Tregs can modulate the local bioavailability of the costimulatory molecule OX40L, which subsequently controlled effector memory T helper 2 cell numbers. Hence, ILC2-Treg interactions represent a critical feedback mechanism to control adaptive type 2 immunity.

EGFR-activated myofibroblasts promote metastasis of pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis. Cancer-associated fibroblasts (CAFs) are recognized potential therapeutic targets, but poor understanding of these heterogeneous cell populations has limited the development of effective treatment strategies. We previously identified transforming growth factor beta (TGF-ß) as a main driver of myofibroblastic CAFs (myCAFs). Here, we show that epidermal growth factor receptor/Erb-B2 receptor (EGFR/ERBB2) signaling is induced by TGF-ß in myCAFs through an autocrine process mediated by amphiregulin. Inhibition of this EGFR/ERBB2-signaling network in PDAC organoid-derived cultures and mouse models differentially impacts distinct CAF subtypes, providing insights into mechanisms underpinning their heterogeneity. Remarkably, EGFR-activated myCAFs promote PDAC metastasis in mice, unmasking functional significance in myCAF heterogeneity. Finally, analyses of other cancer datasets suggest that these processes might operate in other malignancies. These data provide functional relevance to myCAF heterogeneity and identify a candidate target for preventing tumor invasion in PDAC.

Al0.1Ga0.9N p-i-n ultraviolet avalanche photodiodes with suppressed surface leakage current and uniform avalanche breakdown.

We report high-performance Al0.1Ga0.9N p-i-n ultraviolet (UV) avalanche photodiodes (APDs) based on sapphire substrates with stable breakdown voltages (VBR) around 113.4 V, low dark current densities (JBR) below 9 × 10-4 A/cm2 and a high avalanche gain over 2 × 106. A two-step deposition method was employed to reduce passivation-induced plasma damage while maintaining high dielectric film quality. Consistent JBR for various mesa sizes at the VBR are demonstrated, which reveals the suppression of the surface leakage current. Uniform electroluminescence (EL) distributions during the avalanche multiplication processes are displayed, which confirms the elimination of edge breakdown. Pure bulk leakage current distributions and uniform body avalanche breakdown behaviors are observed for the first time in AlGaN APDs. The emission spectra of the EL at various current levels are also presented.

Factors associated with cardiovascular events in systemic lupus erythematosus in a monocentric cohort with up to 40 years of follow-up.

OBJECTIVES: Systemic lupus erythematosus (SLE) is associated with an increased cardiovascular risk. Several traditional and disease-specific risk factors have been shown to correlate with the occurrence of cardiovascular events (CVE) in patients with SLE. However, results of previous studies are diverse. The objectives of this study were to report number, type and those factors associated with CVE in patients with SLE in a large, single-center, ethnically diverse cohort with a long follow-up duration. METHODS: Medical records of patients treated at the Lupus Clinic at University College London Hospital (UCLH) between 1979 and 2020 were retrospectively reviewed. Data about CVE, traditional cardiovascular risk factors, demographic and disease features, and treatment history were collected. Only patients with complete available information were included in the study. Regression analyses were performed to identify factors associated with CVE. RESULTS: Four hundred and nineteen patients were included in the study. Maximum follow-up length was 40 years. Seventy-one (17%) patients had at least one CVE. Multivariable analysis showed that only antiphospholipid antibody positivity (p-value<0.001) was associated with CVE. When analysing different types of CVE, antiphospholipid antibodies were specifically associated with both venous thromboembolic events (p-value<0.001) and cerebrovascular events (p-value=0.007). Dedicated subanalyses revealed that cumulative glucocorticoid dose (p-value=0.010) and a diagnosis of SLE before 2000 (p-value<0.001) were significantly associated with CVE. CONCLUSIONS: Cardiovascular disease is highly prevalent among patients with SLE and is associated with antiphospholipid antibodies, glucocorticoid therapy, and diagnosis before 2000.

Survival analysis of mortality and development of lupus nephritis in patients with systemic lupus erythematosus up to 40 years of follow-up.

OBJECTIVES: Patients with SLE have increased mortality compared with age- and sex-matched controls. LN is a severe manifestation of SLE and an important cause of death. We carried out a retrospective survival analysis to investigate factors that could influence the risk of mortality and LN in a large multi-ethnic cohort of patients with SLE. METHODS: By careful review of medical records, we identified 496 patients with SLE for whom we had complete information regarding the period of observation and occurrence of death and nephritis. Patients were stratified into groups according to sex, ethnicity, age at start of follow-up and time period of diagnosis. Kaplan-Meier analysis was used to investigate differences between the groups. RESULTS: Of the 496 patients in the study, 91 (18.3%) died, 165 (33.3%) developed LN and 33 (6.7%) developed end-stage renal failure. There was no difference between men and women in either mortality or development of LN. Caucasian patients were significantly less likely to develop LN than other ethnic groups (P < 0.0001) but not less likely to die. Patients diagnosed before the median age of 28 years were significantly more likely to develop LN (P < 0.0001) but significantly less likely to die (P = 0.0039) during the period of observation. There has been a significant improvement in survival in patients diagnosed between 1978 and 1989 and those diagnosed between 2006 and 2011 (P = 0.019). CONCLUSION: In our cohort, non-Caucasian ethnicity and younger age at diagnosis are associated with the risk of developing LN. There is evidence of improvement in survival of patients with SLE over time.

A Systematic Review of the safety of non-TNF inhibitor biologic and targeted synthetic drugs in rheumatic disease in pregnancy.

INTRODUCTION: Despite increasing evidence to support safe use of tumour necrosis factor inhibitors (TNFi) and other biologic disease modifying anti-rheumatic drugs (bDMARDs) during pre-conception/pregnancy, there remains a paucity of evidence regarding the safety and compatibility of other non-TNFi and novel targeted synthetic (ts)DMARDs during pre-conception/pregnancy. Therefore, we conducted a systematic review to determine the compatibility of these drugs in pre-conception, during pregnancy and post-partum period. METHOD: Databases including; EMBASE, Pubmed (MEDLINE), and Cochrane were searched up to 23rd October 2020 to find relevant peer-reviewed papers, using keywords including; rheumatic disease, pregnancy, conception/pre-conception, lactation/breastfeeding, childhood and vaccination/infection, and commonly prescribed non-TNFi drugs and tsDMARDs. RESULTS: Our search yielded 1483 papers that were screened independently by two authors, and 109 full-text papers were eligible for final analysis. These studies reported 1291 maternal pregnancies exposed to non-TNFi bDMARDs and tsDMARDs with known outcomes, including 721 live births, 219 spontaneous miscarriages and 27 congenital abnormalities. Paternal exposures in 174 pregnancies had reassuring outcomes. A total of 48 breastfed infants were exposed to non-TNFi bDMARDs and no adverse events reported upon long-term follow-up. Fifteen infants exposed to bDMARDs received normal vaccination regimes, including live vaccines, and had normal developmental outcomes, without any complications or infections. CONCLUSION: Overall, the findings are reassuring and do not suggest a cause for any major concerns or an increased risk of adverse pregnancy outcomes for maternal or paternal exposures to non-TNFi bDMARDs or tsDMARDs. There were no major concerns for breastfeeding exposures to non-TNFi bDMARDs.

Back-illuminated AlGaN heterostructure solar-blind avalanche photodiodes with one-dimensional photonic crystal filter.

AlGaN heterostructure solar-blind avalanche photodiodes (APDs) were fabricated on a double-polished AlN/sapphire template based on a separate absorption and multiplication (SAM) back-illuminated configuration. By employing AlGaN heterostructures with different Al compositions across the entire device, the SAM APD achieved an avalanche gain of over 1×105 at an operated reverse bias of 92 V and a low dark current of 0.5 nA at the onset point of breakdown. These excellent performances were attributed to the acceleration of holes by the polarization electric field with the same direction as the reverse bias and higher impact ionization coefficient of the low-Al-content Al0.2Ga0.8N in the multiplication region. However, the Al0.2Ga0.8N layer produced a photocurrent response in the out of the solar-blind band. To retain the solar-blind detecting characteristic, a periodic Si3N4/SiO2 photonic crystal was deposited on the back of the AlN/sapphire template as an optical filter. This significantly improved the solar-blind characteristic of the device.

High-quantum-efficiency ultraviolet solar-blind AlGaN photocathode detector with a sharp spectral sensitivity threshold at 300 nm.

At present, most of the existing practical ultraviolet (UV) solar-blind detectors are based on Te-Cs photocathode image intensifiers. However, limited by their photoemission characteristics, it is difficult for the Te-Cs-based photocathodes to achieve both high quantum efficiency in the application band and high cutoff ratio in the non-applied band simultaneously. In this paper, a high-quantum-efficiency UV solar-blind detector based on AlGaN photocathodes with a sharp spectral sensitivity threshold at 300 nm is reported. The proposed AlGaN photocathode has extremely high quantum efficiency (i.e., 20%) in the 210-275 nm band, while the efficiency curve steeply reduces to 2% at 300 nm, showing obviously superior performance than the existing Te-Cs photocathodes.