Biology subjects

Storm, J.

Publications and source records attributed to Storm, J..

4 recordsLinked to original sources

bioRxiv2026

A Cellular Cytotoxicity Assay using Ready-to-Thaw Target Cells without Washing Steps

Functional testing of cytotoxic lymphocytes is essential for research and quality control (QC), but most assays require freshly prepared target cells and extensive handling. A ready-to-thaw, no-wash, flow cytometry-based cytotoxicity assay was developed using pre-labeled K562 targets cryopreserved in STEM-CELLBANKER(R) EX (SCB) as suitably sized aliquots. SCB tolerability was evaluated in K562, NK-92, and primary natural killer (NK) cells; post-cryopreservation label stability of CellTrace Violet (CTV) and carboxyfluorescein succinimidyl ester (CFSE) was assessed; freezing and thawing conditions were optimized; and wash versus no-wash workflows were compared using viability-based and absolute-count readouts, across effector-to-target (E:T) ratios with NK donors and NK-92 cells. Effector viability remained high at SCB concentrations up to 10%, and 5% SCB was selected for assay design. After cryopreservation, CTV labeling remained stable over the tested storage period, whereas CFSE showed substantial signal loss. Warm-medium thawing performed comparably to water-bath thawing, and the consolidated protocol (SCB plus fetal calf serum and thermal buffering) maintained high post-thaw target viability and recovery. In killing assays, lysis increased with increasing E:T ratios; omission of the post-thaw wash had minimal impact, and 5% SCB did not impair cytotoxic function. This ready-to-thaw workflow reduces hands-on time and sample manipulation, while improving standardization for reproducible results and enabling high-throughput functional testing and QC.

immunology↗

bioRxiv2024

Impact of Plasmodium falciparum patient-derived isolates on brain endothelium in vitro

A hallmark of cerebral malaria (CM) is sequestration of Plasmodium falciparum infected erythrocytes (IE) within the brain microvasculature. Binding of IE to endothelium reduces microvascular flow and combined with an inflammatory response perturbs endothelial barrier function, resulting in breakdown of the blood- brain barrier (BBB). Cytoadherence leads to activation of the endothelium and alters a range of cell processes affecting signalling pathways, receptor expression, coagulation, and disruption of BBB integrity. Here, we investigated whether CM derived parasites elicit differential effects on human brain microvascular endothelial cells (HBMECs), as compared to uncomplicated malaria (UM) derived parasites. Patient-derived IE from UM and CM clinical cases, as well as non-binding skeleton-binding protein 1 knockout parasites, were overlaid onto TNF-activated HBMECs. Gene expression analysis of endothelial responses was performed using probe-based assays of a panel of genes involved in inflammation, apoptosis, endothelial barrier function, and prostacyclin synthesis pathway. We observed a significant effect on endothelial transcriptional responses in the presence of IE, yet there was no significant correlation between HBMEC responses and type of clinical syndrome (UM or CM). Further, there was no correlation between HBMEC gene expression and both binding itself and level of IE binding to HBMEC, as we detected the same change in endothelial responses when employing both binding and non-binding parasites. Our results suggest that interaction of IE with endothelial cells in this co-culture model induce some endothelial responses that are independent of clinical origin and independent of the expression of the major variant antigen PfEMP1 on the IE surface.

cell biology↗

bioRxiv2019

Parasite histones mediate leak and coagulopathy in cerebral malaria

Coagulopathy and leak, specific to the brain vasculature, are central pathogenetic components of cerebral malaria (CM). It is unclear how the parasite, Plasmodium falciparum, triggers these processes. Extracellular histones, released from damaged host cells, bind to cell membranes and cause coagulation activation, platelet aggregation and vascular leak in diverse critical illnesses. In CM patients with P. falciparum, serum histones correlate with fibrin formation, thrombocytopenia, and endothelial activation and predict brain swelling on magnetic resonance imaging and fatal outcome. Post-mortem, histones bind to the luminal vascular surface, co-localizing with P. falciparum-infected erythrocytes (IE), and with thrombosis and leak. Purified P. falciparum histones cause toxicity and barrier disruption in cultured human brain microvascular endothelial cells, as does serum from CM patients, reversed by anti-histone antibodies and non-anticoagulant heparin. These data implicate parasite histones as a key trigger of fatal brain swelling in CM. Neutralizing histones with agents such as non-anticoagulant heparin warrant exploration to prevent brain swelling and improve outcome.

microbiology↗

bioRxiv2018

Glutamatergic and cholinergic metabotropic modulation induces plateau potentials in hippocampal OLM interneurons

Oriens-lacunosum moleculare (OLM) cells are hippocampal inhibitory interneurons that have been implicated in regulation of information flow and synaptic plasticity in the CA1 circuit. Since anatomical evidence indicate that OLM cells express metabotopic cholinergic (mAChR) and glutamatergic (mGluR) receptors, such modulation of these cells may contribute to switching between functional modes of the hippocampus. Using a transgenic mouse line to identify the Chrna2-positive OLM cells, we investigated metabotropic neuromodulation of intrinsic properties of OLM cells. We found that both mAChR and mGluR activation increased the spontaneous action potential rate and caused the cells to exhibit long-lasting depolarizing plateau potentials following evoked spike trains. Both the mAChR- and mGluR-induced increased spontaneous firing rate and plateau potentials were dependent on intracellular calcium, and were eliminated by blocking Ca2+-dependent transient receptor potential (TRP) cation channels. At the receptor level, Group I mGluRs were found to be responsible for the glutamatergic modulation of the plateau potentials. There was also a pronounced synergy between the cholinergic and glutamatergic modulation of the plateau potentials. Our findings provide insights into how OLM cells are modulated by different neurotransmitters, and are likely to have functional implications on how OLM cells regulate hippocampal information processing during different brain states.

neuroscience↗