Biology subjects

Ali Awadelkareem, M.

Publications and source records attributed to Ali Awadelkareem, M..

3 recordsLinked to original sources

bioRxiv2026

LIF is a pro-algesic mediator in human and mouse models of arthritis-related pain

Purpose: Emerging clinical and experimental data suggest that JAK inhibitors (JAKi) are effective in reducing pain in Rheumatoid Arthritis (RA). We recently showed that JAK/STAT signalling cytokines, specifically IL-6, LIF and IL-11, are present in the RA joint and can act directly on human induced pluripotent stem cell (iPSC) derived sensory neurons by increasing their STAT3 phosphorylation (pSTAT3) levels and expression of select pain-relevant genes. In this study, we determined that LIF is the most consequential cytokine out of the three, altering neuronal signalling, outgrowth and pain-like behaviours in in vitro and in vivo models of arthritis. Methods: Human iPSC-derived sensory neurons were stimulated with RA synovial fluid (SF) or with cytokine-treated RA synovial fibroblast conditioned media (CM) in the absence or presence of anti-cytokine antibodies. pSTAT3 and pSTAT1 levels, and gene expression patterns, were assessed by Western blot and RNA-sequencing (RNA-seq). The immortalised dorsal root ganglion (DRG)-derived cell line (HD 10.6) was assessed for LIF-induced JAK/STAT signalling in addition to neurite outgrowth assays as a functional readout of the sensory neurons. Lastly, mouse models were used to assess pain-related behaviours in vivo. Results: LIF was found to be consistently present in the SF from painful joints from individuals with inflammatory arthritis. Western blotting revealed that the addition of RA SF to iPSC-derived sensory neurons led to LIF-dependent induction of pSTAT3 and pSTAT1, while RNA-seq demonstrated that LIF alters the transcriptome of sensory neurons more prominently than IL-6 and IL-11, up-regulating signature genes related to peripheral neuron sensitization, such as TRPV1, NTRK1, BDNF, HRH1 and SCN10A. Similarly, cytokine-treated fibroblast conditioned media was shown to induce neuronal pSTAT3 and pSTAT1 and upregulate expression of BDNF, HRH1 and VGF in a LIF-dependent manner. Additionally, the immortalised dorsal root ganglion (DRG)-derived cell line (HD 10.6), which also exhibited LIF-induced pSTAT1 and pSTAT3, showed increased neurite outgrowth after axotomy when treated with LIF. Finally, in a mouse model of inflammatory arthritis, LIF was shown to be significantly increased in the affected joint. Furthermore, intraarticular injection of LIF into a healthy mouse knee resulted in increased responsiveness to painful pressure stimuli in that joint. Conclusions: Our data indicate that LIF may be a pro-algesic mediator and a possible therapeutic target to improve pain management in individuals living with inflammatory arthritis.

neuroscience↗

bioRxiv2026

Extended maturation of the HD10.6 immortalised human dorsal root ganglion cell line enables modelling of nociceptive responses and neural injury

Human sensory neuron models are an important resource for studying pain mechanisms and axon injury and repair. Current systems are limited by accessibility, scalability, or incomplete functional maturation. The HD10.6 human dorsal root ganglion-derived immortalised cell line represents a promising alternative; however, its maturation trajectory and suitability for disease modelling remain incompletely defined. Here, we performed a longitudinal, multi-modal characterisation of HD10.6 cells during differentiation over 28 days. Bulk RNA sequencing revealed progressive transcriptional remodelling, with temporal up-regulation of neuronal and nociceptor-associated gene programmes, including ion channels implicated in pain signalling. Protein-level analyses confirmed increased expression of key nociceptor markers and neuropeptides, including TRPV1, Nav1.7, Nav1.8 and CGRP. Functional assays demonstrated the emergence of sensory neuron-like properties over time. Calcium imaging revealed increasing responsiveness to capsaicin, allyl isothiocyanate, ,{beta}-MeATP, and prostaglandin E2, while patch-clamp electrophysiology at DIV 21 after maturation showed repetitive firing of action potential and, most importantly, exhibited TTX-Resistant sodium currents. These findings establish a temporal relationship between transcriptional changes and functional competence. Finally, we evaluated the utility of HD10.6 neurons for modelling axon degeneration. Treatment with vacor-induced robust neurite degeneration, which was attenuated by pharmacological inhibition of SARM1, demonstrating engagement of conserved axon degeneration pathways. Together, our findings define the progressive maturation of HD10.6 sensory neurons and establish this system as a scalable human platform for studying nociceptor biology and SARM1-dependent axon degeneration.

neuroscience↗

bioRxiv2021

Stress-induced protein disaggregation in the Endoplasmic Reticulum catalysed by BiP

Protein synthesis is supported by cellular machineries that ensure polypeptides fold to their native three-dimensional conformation with high fidelity whilst eliminating misfolded, aggregation-prone species. While protein aggregates can contribute to pathologies exemplified by Alzheimers and Parkinsons diseases, their abundance is normally minimised by molecular chaperones such as HSP70/90, which promote native folding and drive the recycling of aberrantly folded species. Cytoplasmic chaperones (e.g. HSP70/40) can resolve insoluble protein aggregates should the preventive mechanisms falter. However, it is unknown whether an analogous disaggregation system is needed and exists in the Endoplasmic Reticulum (ER), where [~]30% of the proteome is synthesised. Here we show that the ER of a variety of mammalian cell types, including neurons, is endowed with the capability to resolve protein aggregates that accumulate upon expression of metastable proteins. Utilising a purpose-developed protein aggregation probing system with a sub-organellar resolution, we observed steady-state aggregate accumulation in the ER. Strikingly, pharmacological induction of ER stress did not augment aggregates but rather stimulated their clearance within hours. We found that this disaggregation activity was catalysed by the stress-responsive ER molecular chaperone - BiP. Its elimination abolished the disaggregation activity in cells. Further, we reconstructed the disaggregation-reaction in-vitro by a minimal system of ATP-fuelled BiP and its J-protein cofactor. These data reveal a hitherto unknown, non-redundant function of the ER stress response. Thus, our findings may facilitate the identification of aggregation-antagonising strategies and rationalising the age-dependent protein misfolding pathology. SummaryER stress induction activates a protein disaggregation machinery, powered by BiP - an abundant ER chaperone, revealed by a FLIM-based protein aggregation monitoring in live cells.

molecular biology↗