Biology subjects

Leo, P.

Publications and source records attributed to Leo, P..

4 recordsLinked to original sources

bioRxiv2023

Iron Oxide Nanoparticles Coated with Biodegradable Block-Copolymer PDMAEMA-b-PMPC and Functionalized with Aptamer for HER2 Breast Cancer Cell Identification

Hybrid nanoparticles have shown promise in biomedical applications; however, their seamless integration into clinical settings remains challenging. Here, we introduce a novel metal oxide polymer hybrid nanoparticle (NP) with a high affinity for nucleic acids. Iron oxide nanoparticles (IONP) were initially synthesized via the co-precipitation method and subjected to comprehensive characterization. Subsequently, block copolymers were synthesized using the Reversible Addition-Fragmentation Chain Transfer (RAFT) technique, employing the zwitterionic PMPC (Poly (2 Methacryloyloxyethyl Phosphorylcholine)) and the cationic PDMAEMA (Poly(2 (Dimethylamino) Ethyl Methacrylate)) with varying degrees of polymerization. In vitro cytotoxicity studies demonstrated the biocompatibility of the synthesized nanoparticles, with no observed toxicity up to a concentration of 150 {micro}g/mL. The cationic polymer PDMAEMA facilitated the facile coating of IONP, forming the IONPP complex, consisting of a 13.27 metal core and a 3.1 nm block-copolymer coating. Subsequently, the IONPP complex was functionalized with a DNA aptamer specifically targeting the human epidermal growth factor receptor 2 (HER2) in breast cancer, forming IONPPP. The block-copolymer exhibited an EC50 of 7.07 {micro}g/mL and demonstrated enhanced recognition efficiency in HER2-amplified SKBR3 cells. Our study presents a comprehensive IONPPP characterization capable of binding short DNA sequences and targeting proteins such as HER2. This newly developed nanoparticle holds significant potential for cancer cell identification and isolation, offering promising prospects in cancer research and clinical applications. 1. Statement of significanceDespite recent advancements in biomedical research, developing sensitive and specific tools for recognizing biological motifs, such as cell receptors and proteins in complex biological solutions, remains a challenge. Furthermore, current approaches often rely on complex biological derivatives like antibodies, lacking a cost-effective delivery strategy. Our study proposes creating and characterizing a novel hybrid metal oxide polymer nanoparticle named IONPPP, functionalized with a DNA aptamer designed to recognize HER2-positive cells. HER2 is a clinically actionable marker for gastric, gastroesophageal, and, particularly, breast cancers. This unique combination of a metal core with an external polymeric structure offers the potential for identification, isolation, and even theragnostic applications, benefiting from its low toxicity and high specificity. 2. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/543859v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@ff36a9org.highwire.dtl.DTLVardef@15da74corg.highwire.dtl.DTLVardef@1929c6aorg.highwire.dtl.DTLVardef@5afd2f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

Deficiencies of Runx3 and tissue-resident CD4+ intestinal epithelial lymphocytes link intestinal dysbiosis and inflammation in mouse and human spondyloarthropathy

ObjectiveDisturbances in immune regulation, intestinal microbial dysbiosis and intestinal inflammation characterize ankylosing spondylitis (AS), which is associated with RUNX3 loss-of-function variants. ZAP70W163C mutant (SKG) mice have reduced ZAP70 signaling, spondyloarthritis and ileitis. At intestinal epithelial interfaces, lamina propria Foxp3+ regulatory T cells (Treg) and intraepithelial CD4+CD8+TCR{beta}+ lymphocytes (CD4-IEL) control inflammation. TGF-{beta} and retinoic acid (RA)-producing dendritic cells are required for induction of Treg and for CD4-IEL differentiation from CD4+ conventional or Treg precursors, with upregulation of Runx3 and suppression of ThPOK. We investigated Treg, CD4-IEL, ZAP70 and Runx3 in SKG mice and AS patients. MethodsWe compared ileal Treg and CD4-IEL numbers and differentiation in BALB/c and SKG mice, and with ZAP70 inhibition, and related differentially-expressed genes in terminal ileum to ChIP-seq-identified Runx3-regulated genes. We compared proportions of CD4-IEL in ileum and CD4+8+ T cells in blood of AS patients and healthy controls. ResultsZAP70W163C or ZAP70 inhibition prevented intestinal CD4-IEL but not Foxp3+ Treg differentiation in context of TGF-{beta} and RA in vitro and in vivo, resulting in Runx3 and ThPOK dysregulation. CD4-IEL frequency and expression of tissue resident memory T-cell and Runx3-regulated genes was reduced in SKG intestine. Multiple under-expressed genes were shared with risk SNPs identified in human spondyloarthropathies. CD4-IEL were decreased in AS intestine. Double-positive T cells were reduced and Treg increased in AS peripheral blood. ConclusionHigh-affinity TCR-ZAP70 signalling is required for Runx3-mediated intestinal CD4-IEL differentiation from Treg. Genetically-encoded relative immunodeficiency of T cells underpins poor intestinal barrier control in mouse and human spondyloarthropathy. What is already known about this subject?Ankylosing spondylitis (AS) is associated with RUNX3 loss-of-function variants. Capacity of the AS T cell receptor repertoire to expand in response to infectious antigens is reduced. Foxp3+ regulatory T cells (Treg) are increased in AS intestine. ZAP70W163C mutant (SKG) mice have reduced ZAP70 signaling, spondyloarthritis (SpA) and ileitis. Intestinal epithelial Foxp3+ Treg and CD4+CD8+ cytotoxic lymphocytes (CD4-IEL) control local inflammation. CD4-IEL differentiate from Treg, with upregulation of Runx3 and suppression of ThPOK transcription factors. What does this study add?High-affinity TCR-ZAP70 signalling is required for Runx3-mediated intestinal CD4-IEL differentiation from Treg Intestinal CD4-IEL and circulating CD4+CD8+ T cells are reduced in AS while circulating Treg are increased. Impaired CD8 expression may be correctible by TNF inhibition in AS CD4+ T cells. Deficiencies of Runx3 and tissue-resident CD4-IEL link intestinal dysbiosis and inflammation in mouse and human SpA. How might this influence clinical practice or future developments?Genetically-encoded relative T immunodeficiency underpins poor intestinal barrier control in SpA

immunology↗

bioRxiv2020

A novel mutation in KCNK16 causing a gain-of-function in the TALK-1 potassium channel: a new cause of maturity onset diabetes of the young.

BackgroundMaturity-onset diabetes of the young (MODY) is a heterogeneous group of monogenic disorders of impaired glucose-stimulated insulin secretion (GSIS). Mechanisms include {beta}-cell KATP channel dysfunction (e.g., KCNJ11 (MODY13) or ABCC8 (MODY12) mutations); however, no other {beta}-cell channelopathies have been identified in MODY. MethodsA four-generation family with autosomal dominant non-obese, non-ketotic antibody-negative diabetes, without mutations in known MODY genes, underwent exome sequencing. Whole-cell and single-channel K+ currents, Ca2+ handling, and GSIS were determined in cells expressing either mutated or wild-type (WT) protein. ResultsWe identified a novel non-synonymous genetic mutation in KCNK16 (NM_001135105: c.341T>C, p.Leu114Pro) segregating with MODY. KCNK16 is the most abundant and {beta}-cell-restricted K+ channel transcript and encodes the two-pore-domain K+ channel TALK-1. Whole-cell K+ currents in transfected HEK293 cells demonstrated drastic (312-fold increase) gain-of-function with TALK-1 Leu144Pro vs. WT, due to greater single channel activity. Glucose-stimulated cytosolic Ca2+ influx was inhibited in mouse islets expressing TALK-1 Leu114Pro (area under the curve [AUC] at 20mM glucose: Leu114Pro 60.1 vs. WT 89.1; P=0.030) and less endoplasmic reticulum calcium storage (cyclopiazonic acid-induced release AUC: Leu114Pro 17.5 vs. WT 46.8; P=0.008). TALK-1 Leu114Pro significantly blunted GSIS compared to TALK-1 WT in both mouse (52% decrease, P=0.039) and human (38% decrease, P=0.019) islets. ConclusionsOur data identify a novel MODY-associated gene, KCNK16; with a gain-of-function mutation limiting Ca2+ influx and GSIS. A gain-of-function common polymorphism in KCNK16 is associated with type 2 diabetes (T2DM); thus, our findings have therapeutic implications not only for KCNK16-associated MODY but also for T2DM.

genetics↗

bioRxiv2019

A rare SMAD9 mutation identifies the BMP signalling pathway as a potential osteoanabolic target

To identify targets for novel anabolic medicines for osteoporosis, we recruited a large cohort with unexplained high bone mass (HBM). Exome sequencing identified a rare (minor allele frequency 0.0014) missense mutation in SMAD9 (c.65T>C, p.Leu22Pro) segregating with HBM in an autosomal dominant family. The same mutation was identified in another two unrelated individuals with HBM. In-silico protein modelling predicts the mutation severely disrupts the MH1 DNA-binding domain of SMAD9. Affected individuals have bone mineral density [BMD] Z-Scores +3 to +5, with increased volumetric cortical and trabecular BMD, increased cortical thickness, and low/normal bone turnover. Fractures and nerve compressions are not seen. Both genome-wide, and gene-based association testing of heel estimated-BMD in >362,924 UK-Biobank British subjects showed strong associations with SMAD9 (PGWAS=6x10-16; PGENE =8x10-17). Smad9 is highly expressed in murine osteocytes and zebrafish bone tissue. Our findings support SMAD9 as a novel HBM gene, and a potential novel osteoanabolic target.

genetics↗