Biology subjects

Alharbi, R.

Publications and source records attributed to Alharbi, R..

3 recordsLinked to original sources

bioRxiv2026

Feline calicivirus encoding NanoLuc luciferase as a tool for assessing antibody neutralisation and antivirals

Feline calicivirus (FCV) is among the most common viruses to infect cats worldwide, with prevalence estimated to range from 10-90% depending on the population sampled. Typical FCV infection presents with oral ulcerations, fever and in some cases can also lead to clinical signs such as pneumonia or "limping syndrome". However, some FCV strains have been isolated from cats exhibiting virulent systemic (VS) disease, which is associated with high morbidity and mortality. Breakthrough VS-FCV infections have been recorded in vaccinated cats and, therefore, there is considerable interest in developing novel therapeutics for use in the face of VS-FCV outbreaks. However, to design effective therapeutics, a tractable system to systematically assess the efficacy of novel vaccine candidates or antivirals is required. Here, we used reverse genetics to develop an FCV reporter virus, inserting NanoLuc luciferase into the LC protein of FCV-Urbana (FCV-UrbanaNL). We characterised the replication kinetics of FCV-UrbanaNL in comparison to its parent virus and assessed the stability of the reporter over multiple passages. Subsequently, we developed virus neutralisation assays to assess a range of monoclonal antibodies that recognise FCV Urbana. We then assessed the breadth of neutralisation by exchanging the major capsid protein, VP1, of FCV Urbana with VP1 from the vaccine strain F9 and the VS-FCV strain NSW-E1. Finally, we evaluated the utility of the FCVNL reporter system to screen candidate antiviral compounds, identifying GS-441524 (the active metabolite of the parent nucleoside remdesivir) as having therapeutic potential against FCV. These findings highlight the potential of this reporter virus as a powerful molecular tool to accelerate the discovery and development of novel therapeutics.

microbiology↗

bioRxiv2024

KLF6 in Pulmonary Hypertension: The Dual Role of Friend and Foe

BackgroundPulmonary arterial hypertension (PAH) is a severe lung condition with unmet clinical needs, marked by endothelial damage, excessive repair, and arterial narrowing, though mechanisms remain unclear. MethodsThis study investigates Kruppel-like transcription factor 6 (KLF6), known for its role in tissue injury response and cancer onset, in PAH through functional and expression analyses in human pulmonary artery endothelial cells (HPAECs) and human and rodent PAH lung tissues. FindingsKLF6 expression increased in early experimental PAH in response to hypoxia and inflammation, while the expression of endothelial homeostasis regulators KLF2 and KLF4, previously linked to PAH, decreased. KLF6 overexpression enhanced pulmonary endothelial survival and angiogenesis through broad transcriptomic remodelling, including changes in genes governing endothelial homeostasis and arterial identity (e.g., SOX17, ERG, BMPR2) and promoted human pulmonary artery smooth muscle cells (HPASMCs) proliferation, which was inhibited by bosentan and imatinib. KLF6 functional and transcriptomic responses differed from those of KLF2 and KLF4. Comparative analysis of RNA-seq PAH databases and spatial transcriptomic analysis of human idiopathic PAH (IPAH) tissues highlighted strong association of KLF6 with vascular remodelling, especially with the formation of angioproliferative (plexiform) lesions. High KLF6 expression was observed in IPAH vascular endothelium and IPAH blood-derived endothelial progenitor cells. Single nucleus RNA-seq in PAH associated with Alveolar Capillary Dysplasia confirmed disease-related elevated KLF6 expression in arterial endothelial cells. InterpretationAccumulation and reorganization of KLF6+ endothelial cells characterize human PAH. KLF6 drives endothelial repair and an apoptosis-resistant, angioproliferative endothelial phenotype. Targeting KLF6 could be a novel therapeutic approach for PAH. RESEARCH IN CONTEXTO_ST_ABSEvidence before this studyC_ST_ABSPulmonary arterial hypertension (PAH) is a progressive and life-shortening lung disease with no cure. In PAH development, endothelial damage is believed to initiate an abnormal repair process, leading to extensive vascular remodelling and the formation of complex angio-proliferative (plexiform) lesions. We conducted a systematic search of the PubMed database to identify transcription factors potentially involved in driving endothelial repair and promoting an apoptosis-resistant, angio-proliferative vascular phenotype. Previous research has linked the loss of endothelial homeostasis in PAH to the inhibition of transcription factors KLF2 and KLF4. While KLF6 is known to play a vital role in vascular development and supports endothelial repair, its specific role in PAH remains unexplored. Added value of this studyThis study is the first to establish a connection between KLF6 and PAH pathogenesis. Our findings reveal that KLF6 activation is a key feature of an apoptosis-resistant, angio-proliferative endothelial phenotype characteristic of human PAH-associated plexogenic arteriopathy. Furthermore, we identify both overlapping and unique activation patterns and transcriptional programs regulated by KLF2, KLF4, and KLF6 in lung endothelial cells, highlighting KLF6s unique role in driving endothelial dysfunction in PAH. Implications of all the available evidenceTargeting KLF6 offers a promising therapeutic strategy to counteract excessive vascular repair and prevent the vascular remodelling in PAH. FUNDINGPhD studentship from the University of Hafr Al Batin, KSA, and the Saudi Cultural Bureau in London (UKSACB) (Rehab Alharbi). Spatial transcriptomic reagents were funded by the British Heart Foundation Centre of Research Excellence Award and Senior BHF Fellowship FS/18/52/33808 (Allan Lawrie). Human samples used in this research project were obtained from the Imperial College Healthcare Tissue Bank (ICHTB) supported by the National Institute for Health Research (NIHR) Biomedical Research Centre based at Imperial College Healthcare NHS Trust and Imperial College London.

cell biology↗

bioRxiv2021

Enhanced immunogenicity of a synthetic DNA vaccine expressing consensus SARS-CoV-2 Spike protein using needle-free immunization

The ongoing global pandemic of Coronavirus Disease 2019 (COVID-19) calls for an urgent development of effective and safe prophylactic and therapeutic measures. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) glycoprotein is a major immunogenic and protective protein, and plays a crucial role in viral pathogenesis. In this study, we successfully constructed a synthetic codon-optimized DNA-based vaccine as a countermeasure against SARS-CoV-2; denoted as VIU-1005. The design was based on the synthesis of codon-optimized coding sequence for optimal mammalian expression of a consensus full-length S glycoprotein. The successful construction of the vaccine was confirmed by restriction digestion and sequencing, and the protein expression of the S protein was confirmed by western blot and immunofluorescence staining in mammalian cells. The immunogenicity of the vaccine was tested in two mouse models (BALB/c and C57BL/6J). Th1-skewed systemic S-specific IgG antibodies and neutralizing antibodies (nAbs) were significantly induced in both models four weeks post three injections with 100 g of the VIU-1005 vaccine via intramuscular needle injection but not intradermal or subcutaneous routes. Importantly, such immunization induced long-lasting IgG response in mice that lasted for at least 6 months. Interestingly, using a needle-free system, we showed an enhanced immunogenicity of VIU-1005 in which lower doses such as 25-50 g or less number of doses were able to elicit significantly high levels of Th1-biased systemic S-specific IgG antibodies and nAbs via intramuscular immunization compared to needle immunization. Compared to the intradermal needle injection which failed to induce any significant immune response, intradermal needle-free immunization elicited robust Th1-biased humoral response similar to that observed with intramuscular immunization. Furthermore, immunization with VIU-1005 induced potent S-specific cellular response as demonstrated by the significantly high levels of IFN-{gamma}, TNF and IL-2 cytokines production in memory CD8+ and CD4+ T cells in BALB/c mice. Together, our results demonstrate that the synthetic VIU-1005 candidate DNA vaccine is highly immunogenic and capable of inducing long-lasting and Th1-skewed immune response in mice. Furthermore, we show that the use of needle-free system could enhance the immunogenicity and minimize doses needed to induce protective immunity in mice, supporting further preclinical and clinical testing of this candidate vaccine.

immunology↗