THE mitochondrial uniporter modulates neuronal regenerative outgrowth and calcium dynamics following axotomy in C. elegans

Following neuronal injury, calcium signaling plays a critical role in promoting repair processes. Injury produces an initial cytosolic calcium elevation mediated by calcium entry from the cut site, plasma membrane channels, and intracellular storage compartments. Subsequently, a variety of signaling factors are involved in promoting growth cone formation and axon outgrowth and guidance, some of which include DLK-1, CaMP, CED-3, CED-4, and calreticulin. Specific proteins mediating calcium transport have also been reported to significantly affect regenerative outgrowth, particularly inositol triphosphate receptors, voltage-gated calcium channels, and ryanodine receptors. Given that mitochondria can store intracellular calcium and regulate cytosolic calcium levels, we hypothesized that the mitochondrial uniporter (MCU) may play a significant role in neuronal regeneration. We found that inhibiting calcium entry into the mitochondria via a loss of function mutation in MCU significantly enhances axonal outgrowth following laser axotomy of single neurons in C. elegans. This effect is calcium-dependent, with the MCU mutant regenerative phenotype reverting to baseline levels when mutants are chronically treated with the calcium chelator EGTA. We also find that sub-cellular calcium signals at the axon cut site are significantly reduced in MCU mutants, while basal levels of calcium and axon guidance remain unaffected. These findings suggest that mitochondrial calcium regulation plays a significant role in the regeneration of single neurons, and that inhibition of MCU activity may be a promising avenue for the treatment of clinical syndromes derived from axonal injury, such as spinal cord injury.2017-11-03T00:00:00

Nuisance Suit Protection For Farms: North Carolina Law Takes A New Approach

The 1979 Session of General Assembly enacted farm operations from nuisance suits under certain circumstances. Representative Tom Ellis, Jr.(D-Vance) was the principal introducer of the law, along with thirty-five co-sponsors. The law, enacted by unanimous votes in both the House and Senate, is of interest to local government officials and planners because of its implications for urban-rural conflicts in land use, suburban growth patterns, and annexation of farm land by municipalities

Uptake and localisation of mTHPC (Foscan®) and its14C-labelled form in normal and tumour tissues of the hamster squamous cell carcinoma model: a comparative study

The aim of this study was to evaluate the pharmacokinetics of meta(tetrahydroxyphenyl)chlorin (mTHPC) on different tissues of interest in a hamster tumour model and to confirm our earlier animal studies on semi-quantitative fluorescence microscopy. The results obtained by three different evaluation methods were compared: in vivo spectrofluorometry, ex vivo fluorescence microscopy and chemical extraction of 14C-labelled mTHPC. Following intracardiac injection of 0.5 mg kg−1 mTHPC, groups of five tumour-bearing animals were used for in situ light-induced fluorescence spectroscopy. Afterwards, the biopsies were taken and snap frozen for fluorescence microscopy. The presence of radioactivity in serum and tissues was determined after chemical digestion in scintillation fluid using a scintillation counter. For each analysed tissue, a good correlation was observed between the three evaluation methods. The highest fluorescence intensity and quantities of mTHPC were observed between 12 and 24 h in liver, kidney, serum, vascular endothelium and advanced neoplasia. The majority of mTHPC was found at around 48 h in smooth muscle and at 96 h in healthy cheek pouch mucosa and early malignant lesions. The lowest level of mTHPC was noted in striated muscle at all times. No selectivity in dye localisation was observed between early squamous cell carcinoma and healthy mucosa. Soon after the injection, a significant selectivity was noted for advanced squamous cell carcinoma as compared to healthy cheek pouch mucosa or striated muscle. A significant difference in mTHPC localisation and quantity was also observed between striated and smooth muscle during the first 48 h following the injection. Finally, this study demonstrated the usefulness of non-invasive in situ spectroscopic measurements to be performed systematically prior to photodynamic therapy as a real-time monitoring for each treated patient in order to individualise and adapt the light dosimetry and avoid over or under treatments

Abstract 2875: NF-κB signaling in myeloid cells limits urethane-induced lung tumorigenesis.

Abstract The transcription factor Nuclear Factor κB (NF-κB) is a master regulator of inflammation, and its activation is associated with tumorigenesis in a variety of cancers. Our group has shown that NF-κB signaling in lung epithelial cells is critical for lung tumor formation. However, the role of NF-κB in other cell types, such as myeloid cells, during lung carcinogenesis is uncertain. We hypothesized that NF-κB signaling in myeloid cells (macrophages, monocytes, and neutrophils) promotes lung tumorigenesis by facilitating lung inflammation. Surprisingly, we found that mice with myeloid cell-targeted IKKβ deletion (IKKβΔmye mice) developed significantly more tumors than wild-type (WT) mice 4 months following a single intraperitoneal (IP) injection of the lung carcinogen urethane (1g/kg). IKKβΔmye mice exhibited a paradoxical increase in the proinflammatory cytokines IL-1β, TNFα, and CXCL1 at 1 and 2 weeks after urethane treatment, and a more than 3-fold higher influx of CD11b+Ly-6G+ cells into IKKβΔmye lungs compared to WT lungs was observed by flow cytometry 1 week post-urethane. By 6 weeks after urethane, IKKβΔmye mice developed more lung inflammation than WT mice, which was accompanied by an increased burden of atypical adenomatous hyperplasia lesions. Flow cytometric analysis of inflammatory cell populations at this time point revealed a sustained increase in granulocytic cells, indicated by significantly higher numbers of CD11b+Gr-1hi cells in the lungs of IKKβΔmye mice compared to WT mice. Depletion of granulocytic cells using Ly-6G antibody treatment (50ug/weekly IP injection) during the first 6 weeks of urethane-induced lung carcinogenesis reduced lung NF-κB activation and lung tumor incidence in IKKβΔmye mice to levels observed in urethane-treated WT mice, indicating that granulocytic cells play an important role in lung tumorigenesis during early tumor development and may do so by limiting NF-κB activation in lung epithelial cells. Together, our data suggest that the impact of NF-κB signaling on lung tumorigenesis is cell type specific and that myeloid NF-κB signaling limits urethane-induced inflammation and lung tumorigenesis. Granulocytes are important players during early lung tumor promotion, and we propose that they promote lung carcinogenesis through interactions that increase survival and/or proliferation of mutated lung epithelial cells. Citation Format: Allyson McLoed, Rinat Zaynagetdinov, Taylor Sherrill, Fiona Yull, Timothy Blackwell. NF-κB signaling in myeloid cells limits urethane-induced lung tumorigenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2875. doi:10.1158/1538-7445.AM2013-2875</jats:p

Abstract 3667: Attenuation of NF-κB signaling in myeloid cells enhances urethane-induced lung tumorigenesis via neutrophil-derived IL-1β

Abstract The transcription factor Nuclear Factor κB (NF-κB) is a master regulator of inflammation, and its activation is associated with tumorigenesis in a variety of cancers. Our group and others have shown that NF-κB signaling in lung epithelial cells is critical for lung carcinogenesis. However, pharmaceutical inhibition of NF-κB signaling has not been effective for the treatment of lung tumorigenesis in animals or humans for reasons that remain unknown. We hypothesized that attenuation of NF-κB signaling in myeloid cells contributes to inefficacy of NF-κB-targeted therapies by unmasking pro-tumorigenic characteristics of inflammatory cells. For our studies, we utilized mice with IKKβ deleted specifically in myeloid cells (IKKβΔmye) and induced lung tumorigenesis with a single intraperitoneal injection of the lung carcinogen urethane (1g/kg). We found that IKKβΔmye mice developed more atypical adenomatous hyperplasia (AAH) lesions at 6 weeks and more lung tumors at 16 weeks after urethane compared to WT mice. These histological differences were correlated with enhanced lung inflammation in IKKβΔmye mice which was observed 1 week after urethane and persisted for up to 6 weeks. Flow cytometric analysis of myeloid cell populations revealed that IKKβΔmye mice contained more pulmonary neutrophils at 1 and 6 weeks after urethane compared to WT mice. In bone marrow chimeras generated by transplantation of bone marrow from IKKβΔmye or WT donors into recipient NF-κB reporter mice, we showed that depletion of neutrophils using Ly6G antibody treatment (100 ug/twice weekly IP injection) during the first 6 weeks of lung carcinogenesis blocked NF-κB activation in stromal cells and reduced tumor multiplicity in IKKβΔmye mice to levels observed in urethane-treated WT mice. Thus, these studies indicated that neutrophils play an important role in lung tumorigenesis during early tumor development and may do so by limiting NF-κB activation in lung epithelial cells. Evaluation of cytokines in the lungs of WT and IKKβΔmye mice at 1 week after urethane revealed higher levels of IL-1β in IKKβΔmye mice compared to WT mice. Further examination of myeloid cells sorted 1 week after urethane identified neutrophils as the key producers of IL-1β in the lungs of IKKβΔmye mice. Neutralization of IL-1 signaling by IL-1 receptor antagonist (IL-1ra) treatment during the first 4 weeks of carcinogenesis decreased the number of AAH lesions in IKKβΔmye mice 6 weeks after urethane and reduced the number of tumors at 16 weeks in IKKβΔmye mice. Taken together, our data suggest that neutrophils can support tumor promotion through production and secretion of IL-1β, which may activate pro-tumorgenic NF-κB signaling in the lung epithelium. We speculate that the paradoxical increase in inflammation and IL-1β production resulting from NF-κB inhibition in myeloid cells contributes to the lack of effectiveness of NF-κB inhibitors in patients with lung cancer. Citation Format: Allyson McLoed, Rinat Zaynagetdinov, Taylor Sherrill, Fiona Yull, Timothy Blackwell. Attenuation of NF-κB signaling in myeloid cells enhances urethane-induced lung tumorigenesis via neutrophil-derived IL-1β. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3667. doi:10.1158/1538-7445.AM2014-3667</jats:p