Biology subjects

Howe, C. L.

Publications and source records attributed to Howe, C. L..

3 recordsLinked to original sources

bioRxiv2018

Surface Plasmon Resonance Imaging of Excitable Cells

Surface plasmons are highly sensitive to refractive index variations adjacent to the surface. This sensitivity has been exploited successfully for chemical and biological assays. In these systems, a surface plasmon resonance (SPR)-based sensor detects temporal variations in the refractive index at a point. SPR has also been used in imaging systems where the spatial variations of refractive index in the sample provide the contrast mechanism. A high numerical aperture objective lens has been used to design SPR microscopy systems with the ability to image adherent live cells. Addressing research questions in cell physiology and pharmacology often requires the development of a multimodal microscope where complementary information can be obtained.\n\nIn this paper, we present the development of a multimodal microscope that combines surface plasmon resonance imaging with a number of additional imaging modalities including bright-field, epi-fluorescence, total internal reflection microscopy (TIRM) and SPR fluorescence microscopy. We used a high numerical aperture objective lens to achieve SPR and TIR microscopy with the ability to image adherent live cells non-invasively. The platform has been used to image live cell cultures demonstrating both fluorescent and label-free techniques. The SPR and TIR imaging systems feature a wide field of view (300 {micro}m) that allows measurements from multiple cells while the resolution is sufficient to image fine cellular processes. The ability of the platform to perform label-free functional imaging of living cell was demonstrated by imaging the spatial variations in contraction of stem cell-derived cardiomyocytes. This technique has a promise for non-invasive imaging of the development of cultured cells over very long periods of time.

bioengineering↗

bioRxiv2017

Neuronal CCL2 expression drives inflammatory monocyte infiltration into the brain during acute virus infection

BackgroundViral encephalitis is a dangerous compromise between the need to robustly clear pathogen from the brain and the need to protect neurons from bystander injury. Theiler's murine encephalomyelitis virus (TMEV) infection of C57Bl/6 mice is a model of viral encephalitis in which the compromise results in hippocampal damage and permanent neurological sequelae. We previously identified brain infiltrating inflammatory monocytes as the primary driver of this hippocampal pathology, but the mechanisms involved in recruiting these cells to the brain were unclear.\n\nMethodsChemokine expression levels in the hippocampus were assessed by microarray, ELISA, RT-PCR, and immunofluorescence. Monocyte infiltration during acute TMEV infection was measured by flow cytometry. CCL2 levels were manipulated by immunodepletion and by specific removal from neurons in mice generated by crossing a line expressing the Cre recombinase behind the synapsin promoter to animals with floxed CCL2.\n\nResultsInoculation of the brain with TMEV induced hippocampal production of the proinflammatory chemokine CCL2 that peaked at 6 hours postinfection, whereas inoculation with UV-inactivated TMEV did not elicit this response. Immunofluorescence revealed that hippocampal neurons expressed high levels of CCL2 at this timepoint. Genetic deletion of CCR2 and systemic immunodepletion of CCL2 abrogated or blunted the infiltration of inflammatory monocytes into the brain during acute infection. Specific genetic deletion of CCL2 from neurons reduced serum and hippocampal CCL2 levels and inhibited inflammatory monocyte infiltration into the brain.\n\nConclusionsWe conclude that intracranial inoculation with infectious TMEV rapidly induces the expression of CCL2 in neurons, and this cellular source is necessary for CCR2-dependent infiltration of inflammatory monocytes into the brain during the most acute stage of encephalitis. These findings highlight a unique role for neuronal production of chemokines in the initiation of leukocytic infiltration into the infected central nervous system.

neuroscience↗

bioRxiv2017

Systemic evidence of acute seizure-associated neuronal injury in patients with temporal lobe epilepsy

Patients with drug refractory temporal lobe epilepsy frequently accumulate cognitive impairment over time, suggesting neuronal loss induced by seizures. We measured serum levels of neuron-specific enolase (NSE), a neuronal injury marker, relative to levels of S100{beta}, a marker of glial injury, at 6 AM, 9 AM, noon, 3 PM, and 6 PM over the course of several days in 7 epilepsy patients and 4 healthy controls. All epilepsy patients exhibited significant deviations in NSE levels through time, and 4 of the epilepsy patients exhibited large sample entropy values and large signal variation metrics for NSE relative to S100{beta}. Controls did not exhibit such changes. Correlation analysis revealed that NSE levels were significantly elevated after clinical seizure events. There was also a highly significant relationship between increased EEG spike frequency and an increase in serum NSE levels measured 24 hours later. The detection of large but transient post-ictal increases in NSE suggests that even self-limited seizures may cause an injury to neurons that underlies cognitive decline in some patients. Post-ictal assessment of serum NSE may serve as a biomarker for measuring the efficacy of future acute neuroprotective strategies in epilepsy patients.

neuroscience↗