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BackgroundThe preoperative characterization of thyroid nodules is a challenge for the clinicians. Fine-needle aspiration (FNA) is the commonly used pre-operative technique for diagnosis of malignant thyroid tumor. However, many benign lesions, with indeterminate diagnosis following FNA, are referred to surgery. There is an urgent need to identify biomarkers that could be used with the FNA to distinguish benign thyroid nodules from malignant tumors. The purpose of the study is to examine the level of expression of the helicase-like transcription factor (HLTF) in relation to neoplastic progression of thyroid carcinomas.MethodsThe presence of HLTF was investigated using quantitative and semi-quantitative immunohistochemistry in a series of 149 thyroid lesion specimens. Our first clinical series was composed of 80 patients, including 20 patients presenting thyroid adenoma, 40 patients presenting thyroid papillary carcinoma, 12 patients presenting thyroid follicular carcinoma and 8 patients presenting anaplastic carcinoma. These specimens were assessed quantitatively using computer assisted microscopy. Our initial results were validated on a second clinical series composed of 40 benign thyroid lesions and 29 malignant thyroid lesions using a semi-quantitative approach. Finally, the HLTF protein expression was investigated by Western blotting in four thyroid cancer cell lines.ResultsThe decrease of HLTF staining was statistically significant during thyroid tumor progression in terms of both the percentage of mean optical density (MOD), which corresponds to the mean staining intensity (Kruskall-Wallis: p < 0.0005), and the labelling index (LI), which corresponds to the percentage of immunopositive cells (Kruskall-Wallis: p < 10−6). Adenomas presented very pronounced nuclear HLTF immunostaining, whereas papillary carcinomas exhibited HLTF only in the cytoplasm. The number of HLTF positive nuclei was clearly higher in the adenomas group (30%) than in the papillary carcinomas group (5%).The 115-kDa full size HLTF protein was immunodetected in four studied thyroid cancer cell lines. Moreover, three truncated HLTF forms (95-kDa, 80-kDa and 70-kDa) were also found in these tumor cells.ConclusionsThis study reveals an association between HLTF expression level and thyroid neoplastic progression. Nuclear HLTF immunostaining could be used with FNA in an attempt to better distinguish benign thyroid nodules from malignant tumors.

The neuroprotective effect of hypothermia has been demonstrated in in vivo and in vitro models of cerebral ischemia. In regard to the hippocampus, previous studies have mainly focused on CA1 pyramidal neurons, which are very vulnerable to ischemia. But the dentate gyrus (DG), in which neuronal proliferation occurs, can also be damaged by ischemia. In this study, we explored the neuroprotective effect of postischemic hypothermia in different areas of the hippocampus after mild or severe ischemia. Organotypic hippocampal slice cultures were prepared from 6- to 8-day-old rats and maintained for 12 days. Cultures were exposed to 25 or 35 min of oxygen and glucose deprivation (OGD). Neuronal damage was quantified after 6, 24, 48, and 72 h by propidium iodide fluorescence. Mild hypothermia (33°C) was induced 1 h after the end of OGD and was maintained for a period of 24 h. Short OGD produced delayed neuronal damage in the CA1 area and in the DG and to a lesser extend in the CA3 area. Damage in CA1 pyramidal cells was totally prevented by hypothermia whereas neuroprotection was limited in the DG. Thirty-five-minute OGD induced more rapid and more severe cell death in the three regions. In this case, hypothermia induced 1 h after OGD was unable to protect CA1 pyramidal cells whereas hypothermia induced during OGD was able to prevent cell loss. This study provides evidence that neuroprotection by hypothermia is limited to specific areas and depends on the severity of the ischemia.

BackgroundHippocampal organotypic slices are used to improve the understanding of synaptic plasticity mechanisms because they allow longer term studies compared to acute slices. However, it is more delicate to keep cultures alive in the recording system outside in vitro conditions. Experiments from the organotypic cultures are common but the handling of slices is rarely described in the literature, even though tissue preservation is crucial. Instruments are sometimes required to extract the slices from the culture inserts but this approach is delicate and can lead to damage, given how strongly the slices are attached to the insert.MethodsA new configuration is proposed to secure the transfer of slices from the incubator to the recording chamber through an adaptor piece that can be designed for any model of chamber and/or insert. The adaptor is a Plexiglas ring in which a culture insert containing the slice can be easily introduced and stabilized. This system allows slices to be placed in the interface for electrophysiological investigations without having to detach them from the insert. That way, no damage is caused and the recording system can safely hold the slices, maintaining them close to culture conditions.ResultsIn addition to the description of the adaptation system, slices were characterized. Their viability was validated and microglial expression was observed. According to the experimental conditions, neuroprotective ramified microgliocytes are present. Dendritic spines studies were also performed to determine neuronal network maturity in culture. Moreover, SKF 83822 hydrobromide and three trains of 100 pulses at 100 Hz with a 10‐min inter‐train interval are suggested to induce long‐term potentiation and to record an increase of fEPSP amplitude and slope.ConclusionThis paper provides detailed information on the preparation and characterization of hippocampal organotypic slices, a new recording configuration more suitable for cultures, and a long‐term potentiation protocol combining SKF and trains.

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