Advances and challenges in investigating B-cells via single-cell transcriptomics.

Single-cell RNA sequencing (scRNAseq) and Variable, Diversity, Joining (VDJ) profiling have improved our understanding of B-cells. Recent scRNAseq-based approaches have led to the discovery of intermediate B-cell states, including preplasma cells and pregerminal centre B-cells, as well as unveiling protective roles for B-cells within tertiary lymphoid structures in respiratory infections and cancers. These studies have improved our understanding of transcriptional and epigenetic control of B-cell development and of atypical and memory B-cell differentiation.

CD4 T cells display a spectrum of recall dynamics during re-infection with malaria parasites.

Children in malaria-endemic regions can experience repeated Plasmodium infections over short periods of time. Effects of re-infection on multiple co-existing CD4 T cell subsets remain unresolved. Here, we examine antigen-experienced CD4 T cells during re-infection in mice, using scRNA-seq/TCR-seq and spatial transcriptomics.

Plasmodium infection induces phenotypic, clonal, and spatial diversity among differentiating CD4 T cells.

Naive CD4 T cells must differentiate in order to orchestrate immunity to Plasmodium, yet understanding of their emerging phenotypes, clonality, spatial distributions, and cellular interactions remains incomplete. Here, we observe that splenic polyclonal CD4 T cells differentiate toward T helper 1 (Th1) and T follicular helper (Tfh)-like states and exhibit rarer phenotypes not elicited among T cell receptor (TCR) transgenic counterparts. TCR clones present at higher frequencies exhibit Th1 skewing, suggesting that variation in major histocompatibility complex class II (MHC-II) interaction influences proliferation and Th1 differentiation.

Systemic host inflammation induces stage-specific transcriptomic modification and slower maturation in malaria parasites.

Maturation rates of malaria parasites within red blood cells (RBCs) can be influenced by host nutrient status and circadian rhythm; whether host inflammatory responses can also influence maturation remains less clear. Here, we observed that systemic host inflammation induced in mice by an innate immune stimulus, lipopolysaccharide (LPS), or by ongoing acute infection, slowed the progression of a single cohort of parasites from one generation of RBC to the next. Importantly, plasma from LPS-conditioned or acutely infected mice directly inhibited parasite maturation during in vitro culture, which was not rescued by supplementation, suggesting the emergence of inhibitory factors in plasma.

Increased core body temperature exacerbates defective protein prenylation in mouse models of mevalonate kinase deficiency.

Mevalonate kinase deficiency (MKD) is characterized by recurrent fevers and flares of systemic inflammation, caused by biallelic loss-of-function mutations in MVK. The underlying disease mechanisms and triggers of inflammatory flares are poorly understood because of the lack of in vivo models. We describe genetically modified mice bearing the hypomorphic mutation p.

Bisphosphonate drugs have actions in the lung and inhibit the mevalonate pathway in alveolar macrophages.

Bisphosphonates drugs target the skeleton and are used globally for the treatment of common bone disorders. Nitrogen-containing bisphosphonates act by inhibiting the mevalonate pathway in bone-resorbing osteoclasts but, surprisingly, also appear to reduce the risk of death from pneumonia. We overturn the long-held belief that these drugs act only in the skeleton and show that a fluorescently labelled bisphosphonate is internalised by alveolar macrophages and large peritoneal macrophages in vivo.

The potential of microdialysis to estimate rifampicin concentrations in the lung of guinea pigs.

Optimised pre-clinical models are required for TB drug development to better predict the pharmacokinetics of anti-tuberculosis (anti-TB) drugs to shorten the time taken for novel drugs and combinations to be approved for clinical trial. Microdialysis can be used to measure unbound drug concentrations in awake freely moving animals in order to describe the pharmacokinetics of drugs in the organs as a continuous sampling technique. The aim of this work was to develop and optimise the microdialysis methodology in guinea pigs to better understand the pharmacokinetics of rifampicin in the lung.

Dose-dependent response to infection with SARS-CoV-2 in the ferret model and evidence of protective immunity.

There is a vital need for authentic COVID-19 animal models to enable the pre-clinical evaluation of candidate vaccines and therapeutics. Here we report a dose titration study of SARS-CoV-2 in the ferret model. After a high (5 × 10 pfu) and medium (5 × 10 pfu) dose of virus is delivered, intranasally, viral RNA shedding in the upper respiratory tract (URT) is observed in 6/6 animals, however, only 1/6 ferrets show similar signs after low dose (5 × 10 pfu) challenge.

Use of high frequency jet ventilation as a refinement for imaging macaques with respiratory disease.

Imaging is used in human medicine to diagnose disease and monitor treatment efficacy. Computed tomography (CT) positron emission tomography (PET) and magnetic resonance (MR) are applied to animal models of infectious diseases to increase data quality, enhance their relevance to the clinical situation, and to address ethical issues through reduction of numbers and refinement of study designs. The time required for collection of MR and PET-CT scans means that normal breathing produces motion artefacts that can render images unacceptable.

Mevalonate kinase deficiency leads to decreased prenylation of Rab GTPases.

Mevalonate kinase deficiency (MKD) is caused by mutations in a key enzyme of the mevalonate-cholesterol biosynthesis pathway, leading to recurrent autoinflammatory disease characterised by enhanced release of interleukin-1β (IL-1β). It is currently believed that the inflammatory phenotype of MKD is triggered by temperature-sensitive loss of mevalonate kinase activity and reduced biosynthesis of isoprenoid lipids required for the prenylation of small GTPase proteins. However, previous studies have not clearly shown any change in protein prenylation in patient cells under normal conditions.

A highly sensitive prenylation assay reveals in vivo effects of bisphosphonate drug on the Rab prenylome of macrophages outside the skeleton.

Bisphosphonate drugs such as zoledronic acid (ZOL), used for the treatment of common bone disorders, target the skeleton and inhibit bone resorption by preventing the prenylation of small GTPases in bone-destroying osteoclasts. Increasing evidence indicates that bisphosphonates also have pleiotropic effects outside the skeleton, most likely via cells of the monocyte/macrophage lineage exposed to nanomolar circulating drug concentrations. However, no effects of such low concentrations of ZOL have been reported using existing approaches.