Biotin functionalization of 8-hydroxyquinoline anticancer organometallics: low toxicity but potent activity.

[M(arene)(HQ)Cl] complexes (M = Ru/Os/Rh/Ir; HQ = 8-hydroxyquinoline) have shown promise as anticancer agents. To assess the effect of conjugating biotin (vitamin B7) to such compounds and improve their tumor-targeting ability through interaction with the sodium-dependent multivitamin transporter (SMVT), the chlorido co-ligand was exchanged with biotinylated 6-aminoindazole. The complexes were characterized by NMR spectroscopy and mass spectrometry, and purity was determined by elemental analysis.

Microinjection of β-glucan Into Larval Zebrafish () for the Assessment of a Trained-Like Immunity Phenotype.

The innate immune system can previous inflammatory insults, enabling long-term heightened responsiveness to secondary immune challenges in a process termed "trained immunity." Trained innate immune cells undergo metabolic and epigenetic remodelling and, upon a secondary challenge, provide enhanced protection with therapeutic potential. Trained immunity has largely been studied in innate immune cells in vitro or following ex vivo re-stimulation where the primary insult is typically injected into a mouse, adult zebrafish, or human.

In vitro and in vivo accumulation of the anticancer Ru complexes [Ru(cym)(HQ)Cl] and [Ru(cym)(PCA)Cl]Cl.

The cellular accumulation and the underlying mechanisms for the two ruthenium-based anticancer complexes [Ru(cym)(HQ)Cl] 1 (cym = η-p-cymene, HQ = 8-hydroxyquinoline) and [Ru(cym)(PCA)Cl]Cl 2 (PCA = N-fluorophenyl-2-pyridinecarbothioamide) were investigated in HCT116 human colorectal carcinoma cells. The results showed that the cellular accumulation of both complexes increased over time and with higher concentrations, and that 2 accumulates in greater quantities in cells than 1. Inhibition studies of selected cellular accumulation mechanisms indicated that both 1 and 2 may be transported into the cells by both passive diffusion and active transporters, similar to cisplatin.

Infection-experienced HSPCs protect against infections by generating neutrophils with enhanced mitochondrial bactericidal activity.

Hematopoietic stem and progenitor cells (HSPCs) respond to infection by proliferating and generating in-demand neutrophils through a process called emergency granulopoiesis (EG). Recently, infection-induced changes in HSPCs have also been shown to underpin the longevity of trained immunity, where they generate innate immune cells with enhanced responses to subsequent microbial threats. Using larval zebrafish to live image neutrophils and HSPCs, we show that infection-experienced HSPCs generate neutrophils with enhanced bactericidal functions.

Platinum(terpyridine) complexes with N-heterocyclic carbene co-ligands: high antiproliferative activity and low toxicity .

Pt(terpyridine) complexes are well-known DNA intercalators. The introduction of an NHC co-ligand rendered such a complex highly antiproliferative in cancer cells compared to its chlorido derivative. Despite the high potency, zebrafish embryos tolerated the compound well, especially compared to cisplatin.

Uricase-Deficient Larval Zebrafish with Elevated Urate Levels Demonstrate Suppressed Acute Inflammatory Response to Monosodium Urate Crystals and Prolonged Crystal Persistence.

Gout is caused by elevated serum urate leading to the deposition of monosodium urate (MSU) crystals that can trigger episodes of acute inflammation. Humans are sensitive to developing gout because they lack a functional urate-metabolizing enzyme called uricase/urate oxidase (encoded by the gene). A hallmark of long-standing disease is tophaceous gout, characterized by the formation of tissue-damaging granuloma-like structures ('tophi') composed of densely packed MSU crystals and immune cells.

Plexin D1 negatively regulates zebrafish lymphatic development.

Lymphangiogenesis is a dynamic process that involves the directed migration of lymphatic endothelial cells (LECs) to form lymphatic vessels. The molecular mechanisms that underpin lymphatic vessel patterning are not fully elucidated and, to date, no global regulator of lymphatic vessel guidance is known. In this study, we identify the transmembrane cell signalling receptor Plexin D1 (Plxnd1) as a negative regulator of both lymphatic vessel guidance and lymphangiogenesis in zebrafish.

A New Transgenic Line for Rapid and Complete Neutrophil Ablation.

Zebrafish lines expressing nitroreductase (NTR) in specific cell compartments, which sensitizes those cells to metronidazole (MTZ)-mediated ablation, have proven extremely useful for studying tissue regeneration and investigating cell function. In contrast to many cells, neutrophils are comparatively resistant to the NTR/MTZ targeted ablation strategy. Recently, a rationally engineered variant of NTR (NTR 2.

Towards a new model of trained immunity: Exposure to bacteria and β-glucan protects larval zebrafish against subsequent infections.

Once thought to be a feature exclusive to lymphocyte-driven adaptive immunity, immune memory has also been shown to operate as part of the innate immune system following infection to provide an elevated host response to subsequent pathogenic challenge. This evolutionarily conserved process, termed 'trained immunity', enables cells of the innate immune system to 'remember' previous pathogen encounters and mount stronger responses to the same, or different, pathogens after returning to a non-activated state. Here we show that challenging larval zebrafish, that exclusively rely on innate immunity, with live or heat-killed Salmonella typhimurium provides protection to subsequent infection with either Salmonella typhimurium or Streptococcus iniae, that lasts for at least 12 days.

3,4-Difluorobenzocurcumin Inhibits Vegfc-Vegfr3-Erk Signalling to Block Developmental Lymphangiogenesis in Zebrafish.

Lymphangiogenesis, the formation of new lymphatic vessels from pre-existing vasculature, plays critical roles in disease, including in cancer metastasis and chronic inflammation. Preclinical and recent clinical studies have now demonstrated therapeutic utility for several anti-lymphangiogenic agents, but optimal agents and efficacy in different settings remain to be determined. We tested the anti-lymphangiogenic property of 3,4-Difluorobenzocurcumin (CDF), which has previously been implicated as an anti-cancer agent, using zebrafish embryos and cultured vascular endothelial cells.

A Multitargeted Approach: Organorhodium Anticancer Agent Based on Vorinostat as a Potent Histone Deacetylase Inhibitor.

The combination of more than one bioactive moiety in a multitargeted anticancer agent may result in synergistic activity of its components. Using this concept, bioorganometallic compounds were designed to feature a metal center, a 2-pyridinecarbothioamide (PCA), and a hydroxamic acid, which is found in the anticancer drug vorinostat (SAHA). The organometallics showed inhibitory activity in the nanomolar range against histone deacetylases (HDACs) as the key target for SAHA.

Targeting Drugs to Larval Zebrafish Macrophages by Injecting Drug-Loaded Liposomes.

Zebrafish (Danio rerio) larvae have developed into a popular model to investigate host-pathogen interactions and the contribution of innate immune cells to inflammatory disease due to their functionally conserved innate immune system. They are also widely used to examine how innate immune cells help guide developmental processes. By taking advantage of the optical transparency and genetic tractability of larval zebrafish, these studies often focus on live imaging approaches to functionally characterize fluorescently marked macrophages and neutrophils within intact animals.

Isolation of Neutrophils from Larval Zebrafish and Their Transplantation into Recipient Larvae for Functional Studies.

Live imaging of neutrophils within optically transparent larval zebrafish has proved a powerful technique to investigate how specific gene products control neutrophil function. To resolve whether a gene contributes to neutrophil function in a cell-autonomous manner necessitates a way to examine gene-deficient neutrophils in an otherwise wild type background. To this end, here we describe methods to harvest fluorescent neutrophils from larval donor zebrafish and transplant them into age-matched recipients.

Anticancer organorhodium and -iridium complexes with low toxicity in vivo but high potency in vitro: DNA damage, reactive oxygen species formation, and haemolytic activity.

Redox-modulating anticancer drugs allow the exploitation of altered redox biology observed in many cancer cells. We discovered dinuclear RhIII(Cp*) and IrIII(Cp*) complexes that have in vitro anticancer activity superior to cisplatin and the investigational drug IT-139, while being less toxic in haemolysis and in vivo zebrafish models. The mode of action appears to be related to DNA damage and ROS-mediated stress pathways.

In Vivo Imaging and Quantitation of the Host Angiogenic Response in Zebrafish Tumor Xenografts.

Tumor angiogenesis is a key target of anti-cancer therapy and this method has been developed to provide a new model to study this process in vivo. A zebrafish xenograft is created by implanting mammalian tumor cells into the perivitelline space of two days-post-fertilization zebrafish embryos, followed by measuring the extent of the angiogenic response observed at an experimental endpoint up to two days post-implantation. The key advantage to this method is the ability to accurately quantitate the zebrafish host angiogenic response to the graft.

Glycogen synthase kinase 3-β inhibition induces lymphangiogenesis through β-catenin-dependent and mTOR-independent pathways.

Lymphatic vessels play an important role in health and in disease. In this study, we evaluated the effects of GSK3-β inhibition on lung lymphatic endothelial cells in vitro. Pharmacological inhibition and silencing of GSK3-β resulted in increased lymphangiogenesis of lung lymphatic endothelial cells.

Zebrafish facial lymphatics develop through sequential addition of venous and non-venous progenitors.

Lymphatic vessels are known to be derived from veins; however, recent lineage-tracing experiments propose that specific lymphatic networks may originate from both venous and non-venous sources. Despite this, direct evidence of a non-venous lymphatic progenitor is missing. Here, we show that the zebrafish facial lymphatic network is derived from three distinct progenitor populations that add sequentially to the developing facial lymphatic through a relay-like mechanism.

Liposome-Mediated Drug Delivery in Larval Zebrafish to Manipulate Macrophage Function.

Chemical interventions are regularly used to examine and manipulate macrophage function in larval zebrafish. Given chemicals are typically administered by simple immersion or injection, it is not possible to resolve whether their impact on macrophage function is direct or indirect. Liposomes provide an attractive strategy to target drugs to specific cellular compartments, including macrophages.

Macrophages enhance Vegfa-driven angiogenesis in an embryonic zebrafish tumour xenograft model.

Tumour angiogenesis has long been a focus of anti-cancer therapy; however, anti-angiogenic cancer treatment strategies have had limited clinical success. Tumour-associated myeloid cells are believed to play a role in the resistance of cancer towards anti-angiogenesis therapy, but the mechanisms by which they do this are unclear. An embryonic zebrafish xenograft model has been developed to investigate the mechanisms of tumour angiogenesis and as an assay to screen anti-angiogenic compounds.

Characterization of Zebrafish Facial Lymphatics.

Zebrafish are well-established as a model of vascular development. The genetic tractability, external development, permeability to small molecules and optical transparency of zebrafish larvae are all attributes that make this model attractive to the vascular biologist. There are an increasing number of lymphatic reporter lines that enable the visualization of zebrafish lymphatic vessel growth in vivo; these tools, coupled with either forward or reverse genetics, have provided new insights into the process of lymphatic specification and development.

Blocking fatty acid-fueled mROS production within macrophages alleviates acute gouty inflammation.

Gout is the most common inflammatory arthritis affecting men. Acute gouty inflammation is triggered by monosodium urate (MSU) crystal deposition in and around joints that activates macrophages into a proinflammatory state, resulting in neutrophil recruitment. A complete understanding of how MSU crystals activate macrophages in vivo has been difficult because of limitations of live imaging this process in traditional animal models.

The innate immune cell response to bacterial infection in larval zebrafish is light-regulated.

The circadian clock, which evolved to help organisms harmonize physiological responses to external conditions (such as the light/dark cycle, LD), is emerging as an important regulator of the immune response to infection. Gaining a complete understanding of how the circadian clock influences the immune cell response requires animal models that permit direct observation of these processes within an intact host. Here, we investigated the use of larval zebrafish, a powerful live imaging system, as a new model to study the impact of a fundamental zeitgeber, light, on the innate immune cell response to infection.

Lymphatics, Cancer and Zebrafish.

Many solid tumors are known to metastasize through the lymphatic vasculature. This process is facilitated by the generation of new lymphatic vessels (tumor lymphangiogenesis) and also by the remodelling of existing lymphatics. Together these processes enable the spread of tumor cells to distant sites.

A zebrafish model of inflammatory lymphangiogenesis.

Inflammatory bowel disease (IBD) is a disabling chronic inflammatory disease of the gastrointestinal tract. IBD patients have increased intestinal lymphatic vessel density and recent studies have shown that this may contribute to the resolution of IBD. However, the molecular mechanisms involved in IBD-associated lymphangiogenesis are still unclear.