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Background: High intensity focused ultrasound (HIFU) can destroy tissue by thermal ablation which may be accompanied by acoustic cavitation and/or tissue water boiling, but the biological and histological effects of these treatments have not been fully documented. Here, detailed histological analysis over time using well characterized HIFU exposures in in vivo rat livers is described., Methods: Exposures used invoked either (i) thermal, with acoustic cavitation and/or tissue water boiling or (ii) predominantly thermal damage. Cavitation activity was detected using both active and passive methods. Histological assessment involved hematoxylin and eosin (H&E), picrosirius red and immunohistochemical staining., Results: Distinct concentric damage regions were identified after HIFU exposures. The outermost ring showed a red H&E-stained rim that was characterized by hemorrhage. The adjacent inner band appeared white due to increased extracellular spaces. The morphology of the next zone depended on the exposure. Where there was no tissue acoustic cavitation/water boiling, this was the lesion center, in which heat-fixed cells were seen. Where acoustic cavitation/boiling occurred, a centermost zone with irregular holes up to several hundred microns across was seen. Cleaved caspase-3 and Hsp70 staining in the periphery of both types of HIFU exposures was seen within the outermost ring of hemorrhage, where an inflammatory response was also observed. By day 7, a distinct acellular region in the center of the HIFU lesions had been created., Conclusions: These results identify the morphological effects and elucidate the similarities and differences of HIFU-induced thermal lesions in the presence or absence of acoustic cavitation/tissue water boiling.

Background: Sub-ablative heat induces pleiotropic biological effects in cancer cells, activating programmed cell death or survival processes. These processes decide the fate of the heated cell. This study investigates these and assesses whether heat, in combination with HSP90 inhibition, augments cell death and induces a pro-immune phenotype in these cells., Methods: HCT116 and HT29 cells were subjected to thermal doses (TID) of 60 and 120CEM 43 using a PCR thermal cycler. HSP90 was inhibited with NVP-AUY922. Viability was assessed using the MTT assay. Cellular ATP and HSP70 release were assessed using ATP and Enzyme-linked Immunosorbent assays, respectively. Flow cytometry and immunoblotting were used to study the regulation of biomarkers associated with the heat shock response, the cell cycle, and immunogenic and programmed cell death., Results: Exposure of HCT116 and HT29 cells to TIDs of 60 and 120CEM 43 decreased their viability. In addition, treatment with 120CEM 43 increased intracellular HSP70 and the percentage of HCT116/HT29 cells in the G2/M cell cycle phase, ATP release and Calreticulin/HSP70/HSP90 exposure in the plasma membrane, while downregulating CD47 compared to sham-exposed cells. When combined with NVP-AUY922, treatment of HCT116/HT29 cells with 120CEM 43 resulted in a synergistic decrease of cell viability associated with the induction of apoptosis. Also, the combined treatments increased Calreticulin exposure, CD47 downregulation, and HSP70 release compared to the sham-exposed cells., Conclusion: Sub-ablative heating can act synergistically with the clinically relevant HSP90 inhibitor NVP-AUY922 to induce a pro-immunogenic form of cell death in colon cancer cells.

Aim: Thermal isoeffective dose (TID) has not been convincingly validated for application to predict biological effects from rapid thermal ablation (e.g., using >55 °C). This study compares the classical method of quantifying TID (derived from hyperthermia data) with a temperature-adjusted method based on the Arrhenius model for predicting cell survival in vitro, after either 'rapid' ablative or 'slow' hyperthermic exposures., Methods: MTT assay viability data was obtained from two human colon cancer cell lines, (HCT116, HT29), subjected to a range of TIDs (120-720 CEM 43 ) using a thermal cycler for hyperthermic (>2 minutes, <50 °C) treatments, or a novel pre-heated water bath based technique for ablative exposures (<10 seconds, >55 °C). TID was initially estimated using a constant R CEM>43 ° C =0.5, and subsequently using R CEM (T), derived from temperature dependent cell survival (injury rate) Arrhenius analysis., Results: 'Slow' and 'rapid' exposures resulted in cell survival and significant regrowth (both cell lines) 10 days post-treatment for 240 CEM 43 (R CEM>43 ° C =0.5), while 340-550 CEM 43 (R CEM>43 ° C =0.5) delivered using 'rapid' exposures showed 12 ± 6% viability and 'slow' exposures resulted in undetectable viability. Arrhenius analysis of experimental data (activation energy ΔE = 5.78 ± 0.04 × 10 5  J mole -1 , frequency factor A = 3.27 ± 11 × 10 91  sec -1 ) yielded R CEM =0.42 * e 0.0041*T which better-predicted cell survival than using R  CEM> 43 ° C =0.5., Conclusions: TID calculated using an R CEM (T) informed by Arrhenius kinetic parameters provided a more consistent, heating strategy independent, predictor of cell viability, improving dosimetry of ablative thermal exposures. Cell viability was only undetectable above 305 ± 10 CEM 43 using this revised measure., (© 2019 The Author(s). Published with license by Taylor & Francis Group, LLC This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.)

Purpose: The pleiotropic effects of heat on cancer cells have been well documented. The biological effects seen depend on the temperature applied, and the heating duration. In this study we investigate the cytotoxic effects of heat on colon cancer cells and determine how different cell death processes such as autophagy, apoptosis and necroptosis play a role in cell response., Materials and Methods: The thermal dose concept was used to provide a parameter that will allow comparison of different thermal treatments. Two human colon cancer cell lines, HCT116 and HT29, were subjected to ablative temperatures using a polymerase chain reaction thermal cycler. Temperature was recorded using thermocouples. Cell viability was assessed using the MTT assay. Induction of apoptosis was estimated using an enzyme-linked immunosorbent assay that detects cleaved cytoplasmic nucleosomes. Protein regulation was determined using immunoblotting. The percentage of cells undergoing apoptosis and autophagy was determined with annexin V/propidium iodide staining and a cationic amphiphilic tracer using fluorescence-activated cell sorting analysis., Results: Exposure of colon cancer cells to ablative thermal doses results in decreased cell viability. The cytotoxic effect of heat is associated with induction of apoptosis and autophagy, the amount depending on both the thermal dose applied and on the time elapsed after treatment. Autophagy induction is mainly seen in live cells. RIPK3 protein levels are increased after exposure of cells to heat. A necroptosis inhibitor does not affect cell viability., Conclusions: Autophagy, apoptosis and necroptosis are associated with the response of these cancer cell lines to supra-normal temperatures., (© 2015 The Author(s). Published with license by Taylor & Francis Group, LLC This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/Licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited)