Biology subjects

Hampton, J.

Publications and source records attributed to Hampton, J..

4 recordsLinked to original sources

bioRxiv2024

AHR activation accelerates the resolution of TGF-β1 induced fibroblast activation and promotes alveolar type 1 cell regeneration in alveolar organoids

Regeneration of the alveolar epithelium is necessary to restore tissue architecture and gas exchange capabilities in chronic pulmonary diseases such as fibrosing interstitial lung disease. While it is known alveolar type 2 (AT2) cells give rise to alveolar type 1 (AT1) cells to repair the alveolar epithelium after injury, methods to promote this process under pathological settings are poorly understood. Here, using a complex 3D organoid culture with TGF-{beta}1 dependent impaired AT1 spheroid formation, we performed a high-throughput screen (HTS) with [~]16,800 compounds to identify small molecules that increase number of AT1 spheroids. Longitudinal single cell RNA sequencing (scRNA-seq) revealed that DB-11-BE87 increased AT1 regeneration by reducing TGF-{beta}1 induced fibroblast activation, concurrently with AHR activation in those cells. These studies highlight a novel HTS system to identify factors that can promote AT1 differentiation and suggest AHR activation as a method to counteract pathological TGF-{beta}1 signaling in pulmonary disease.

cell biology↗

bioRxiv2022

Aromatase Inhibitor Induced Musculoskeletal Inflammation is Observed Independent of Oophorectomy in a Novel Mouse Model

BackgroundAromatase Inhibitors (AIs) block physiological estrogen production in peripheral tissues and are used clinically to reduce disease recurrences and improve overall survival rates in hormone receptor-positive breast cancer patients. However, half of patients taking these drugs develop aromatase inhibitor induced arthralgia (AIIA), which is characterized by severe pain and inflammation in various joints and the surrounding musculoskeletal tissue. While the pathophysiology is not currently understood, it has been proposed to be associated with systemic estrogen deficiency resulting from AI treatment. Since AIIA leads to suspension of therapy in 20-30% of patients, reducing AIIA incidence may provide sustained AI treatment and enhance long-term survival. ObjectiveIn order to establish a better understanding of disease pathology and to create a platform that can be used to explore future interventional strategies, our objective in this study was to design a novel animal model of AIIA. MethodsFemale BALB/C-Tg(NF{kappa}B-RE-luc)-Xen mice, which have a firefly luciferase cDNA reporter transgene under the regulation of NF{kappa}B binding sites, were oophorectomized and treated with AI (letrozole) by daily subcutaneous injections for 5 weeks. Control groups included oophorectomized mice receiving vehicle injections and non-oophorectomized mice treated with AI. Knee joints and surrounding muscle tissue were imaged on the BioSpec 94/30 micro-MRI. The primary weight-bearing joint (hind limb) was examined histopathologically and NF{kappa}B activity was measured by bioluminescent imaging. Serum was collected for cytokine analysis. Additionally, healthy human PBMCs were treated with letrozole, estrogen, or both, and RNA sequencing was performed at 36 hrs. ResultsBioluminescent imaging showed significantly enhanced NF{kappa}B activation with AI treatment in the hind limbs compared to controls receiving vehicle treatment. Moreover, analysis of knee joints and legs by MRI showed enhanced signal detection in the joint space and surrounding tissue following daily AI injections. Surprisingly, the enhanced MRI detection and NF{kappa}B activation was observed with AI treatment independent of the oophorectomy procedure. This indicates that the induction of musculoskeletal-directed inflammation by AI is not mediated by changes in physiological estrogen levels, which is contrary to proposed mechanisms of disease pathogenesis. Similarly, histopathological analysis showed tenosynovitis and musculoskeletal infiltrates in all mice receiving AI with or without oophorectomy. IHC analysis of the infiltrates demonstrated a predominantly macrophage-mediated inflammatory response with scattered CD4+ T cells. Additionally, serum cytokine levels of IL-2, IL-4, IL-6, and CXCL1 were significantly elevated in mice with AI treatment. RNA sequencing of human PBMCs after in vitro AI stimulation did not demonstrate an AI-specific gene expression pattern associated with immune system activation directly, suggesting that the pathogenesis of AIIA may be mediated through cells in other tissues in vivo. ConclusionsCollectively, these data establish a novel mouse model of AIIA and identify an estrogen-independent stimulation of disease pathology via AI-mediated induction. This suggests that the pathogenesis of AIIA may not be mediated by estrogen deficiency, as previously hypothesized, and indicates that AI-induced inflammation may not be regulated directly through a pathogenic mechanism initially derived from circulating mononuclear cells. Future studies aim to characterize this inflammatory mechanism in vivo with a focus on other cells, including macrophages, synovial cells and chondrocytes, to provide insight into putative therapeutic strategies directed at mitigating disease pathology.

immunology↗

bioRxiv2022

Isolation, purification, and characterisation of a phage tail-like bacteriocin from the insect pathogenic bacterium Brevibacillus laterosporus

The gram-positive and spore-forming bacterium Brevibacillus laterosporus (Bl) belongs to the Brevibacillus brevis phylogenetic cluster. Isolates of the species have demonstrated pesticidal potency against a wide range of invertebrate pests and plant diseases. Two New Zealand isolates, Bl 1821L and Bl 1951, are under development as biopesticides for control of diamondback moth and other pests. However, due to often restricted growth of these endemic isolates, production can be an issue. During investigations of the cause of the disrupted growth, electron micrographs of crude lysate of Bl 1821L showed the presence of phages tail-like structures. PEG 8000 precipitated lysate harbouring the putative phage tail-like particles demonstrated broad-spectrum activity against several gram-positive bacteria. SDS-PAGE of purified and concentrated lysate showed a prominent protein band of [~]48 kD from where transmission electron microscopy revealed the presence of polysheath-like structures. N-terminal sequencing of the [~]48 kD protein mapped to a gene with weak predicted amino acid homology to a Bacillus PBSX phage-like element xkdK, the translated product of which shared >90% amino acid similarity to the phage tail-sheath protein of another Bl published genome, LMG15441. An xkdK homolog was also identified in the Bl 1951 genome. However, genome comparison of the region around the xkdK gene between Bl 1821L and Bl 1951 found differences including two glycine rich protein encoding genes which contain imperfect repeats (1700 bp) in Bl 1951, while a putative phage region resides in the analogous Bl 1821L region. Although comparative analysis of the genomic organisation of Bl 1821L and Bl 1951 PBSX-like region with the defective phages PBSX, PBSZ, and PBP 180 of Bacillus subtilis isolates 168 and W23, and Bacillus phage PBP180 revealed low amino acids similarity, the genes encode similar functional proteins in similar arrangements, including phage tail-sheath (XkdK), tail (XkdO), holin (XhlB), and N-acetylmuramoyl-L-alanine (XlyA). AMPA analysis identified a bactericidal stretch of 13 amino acids in the [~]48 kD sequenced protein of Bl 1821L. Assays of purified [~]48 kD protein of Bl 1821L caused a decrease of 34.2% in the number of viable cells of Bl 1951, 18 hours after treatment as compared to the control. Significance of the studyThis study for the first time isolated, purified, and characterised putative phage tail-like bacteriocins (PTLBs) from the insect pathogenic isolates of Brevibacillus laterosporus. Identified PTLBs caused a decrease in the number of viable cells of Bl 195, 18 hours after treatment as compared to control. Therefore, it is likely that the putative PTLBs might have implications in harnessing the insecticidal potential of this useful bacterium.

microbiology↗

bioRxiv2020

Cooperative Policing Behavior Regulates Reproductive Division of Labor in a Termite

Reproductive conflicts are common in insect societies where helping castes retain reproductive potential. One of the mechanisms regulating the conflicts is policing, a coercive behavior that reduces direct reproduction by other individuals. In eusocial Hymenoptera (ants, bees, and wasps), workers or the queen act aggressively toward fertile workers, or destroy their eggs. In many termite species (order Blattodea), upon the death of primary queen and king, workers or nymphs can differentiate into neotenic reproductives and inherit the breeding position. During this process, competition among neotenics is inevitable, but how this conflict is resolved remains unclear. Here, we report a policing behavior that regulates reproductive division of labor in the eastern subterranean termite, Reticulitermes flavipes. Our results demonstrate that the policing behavior is a cooperative effort performed sequentially by successful neotenics and workers. A neotenic reproductive initiates the attack of the fellow neotenic by biting and displays alarm behavior. Workers are then recruited to cannibalize the injured neotenic. Furthermore, the initiation of policing is age-dependent, with older reproductives attacking younger ones, thereby inheriting the reproductive position. This study provides empirical evidence of policing behavior in termites, which represents a convergent trait shared between eusocial Hymenoptera and Blattodea.

animal behavior and cognition↗