Table_1_The Role of SIRT3 in the Brain Under Physiological and Pathological Conditions.docx

<p>Sirtuin enzymes are a family of highly seven conserved protein deacetylases, namely SIRT1 through SIRT7, whose enzymatic activities require the cofactor nicotinamide adenine dinucleotide (NAD⁺). Sirtuins reside in different compartments within cells, and their activities have been shown to regulate a number of cellular pathways involved in but not limited to stress management, apoptosis and inflammatory responses. Given the importance of mitochondrial functional state in neurodegenerative conditions, the mitochondrial SIRT3 sirtuin, which is the primary deacetylase within mitochondria, has garnered considerable recent attention. It is now clear that SIRT3 plays a major role in regulating a host of mitochondrial molecular cascades that can contribute to both normal and pathophysiological processes. However, most of the currently available knowledge on SIRT3 stems from studies in non-neuronal cells, and the consequences of the interactions between SIRT3 and its targets in the CNS are only beginning to be elucidated. In this review, we will summarize current advances relating to SIRT3, and explore how its known functions could influence brain physiology.</p

ALA-PpIX mediated photodynamic therapy of malignant gliomas augmented by hypothermia.

Malignant gliomas are highly invasive, difficult to treat, and account for 2% of cancer deaths worldwide. Glioblastoma Multiforme (GBM) comprises the most common and aggressive intracranial tumor. The study hypothesis is to investigate the modification of Photodynamic Therapy (PDT) efficacy by mild hypothermia leads to increased glioma cell kill while protecting normal neuronal structures.Photosensitizer accumulation and PDT efficacy in vitro were quantified in various glioma cell lines, primary rat neurons, and astrocytes. In vivo studies were carried out in healthy brain and RG2 glioma of naïve Fischer rats. Hypothermia was induced at 1 hour pre- to 2 hours post-PDT, with ALA-PpIX accumulation and PDT treatments effects on tumor and normal brain PDT quantified using optical spectroscopy, histology, immunohistochemistry, MRI, and survival studies, respectively.In vitro studies demonstrated significantly improved post-PDT survival in primary rat neuronal cells. Rat in vivo studies confirmed a neuroprotective effect to hypothermia following PpIX mediated PDT by T2 mapping at day 10, reflecting edema/inflammation volume reduction. Mild hypothermia increased PpIX fluorescence in tumors five-fold, and the median post-PDT rat survival time (8.5 days normothermia; 14 days hypothermia). Histology and immunohistochemistry show close to complete cellular protection in normal brain structures under hypothermia.The benefits of hypothermia on both normal neuronal tissue as well as increased PpIX fluorescence and RG2 induced rat survival strongly suggest a role for hypothermia in photonics-based surgical techniques, and that a hypothermic intervention could lead to considerable patient outcome improvements

Mild hypothermia modulates PDT responsivity of cell lines and primary neuronal cells <i>in vitro</i>.

<p>A) LD₅₀ values of 5 glioma cell lines, one purported glioma cancer stem cell line (GS2), as well as primary murine neurons (DIV 13) and astrocytes of PDT with and without hypothermia, note LD₅₀ of ALA, was chosen as a surrogate for PDT dose in these studies. Hypothermia altered LD₅₀ significantly from primary neurons and astrocytes, as well as GS2 cells (p<0.05, n = 3). B) Normalized dose-response curve of primary astrocytes and C) of primary neurons.</p

Hypothermia reduces lesion volume on the healthy brain following PDT treatment.

<p>A) Extracted regions of GFAP staining used in the analysis. For each subgroup, 2 animals were subjected to PDT and followed for 10 days using MRI. Note: Normothermia animals demonstrated some necrosis that was not seen in the other subgroups (The top and bottom panels represent different animals) (blue arrows mark out increased GFAP staining, red arrows mark out cell death).</p

Hypothermia reduces volume and intensity of edema/inflammation on the healthy brain following PDT.

<p>A) T2 weighted images of central slice regions (areas with the highest amount of inflammation) for each of the treatment conditions. B) Mean T2 of the ROIs of T2 maps for each of the treatment conditions (p>0.3, n = 4 animals) C) Number of voxels above baseline in each treatment group versus the contralateral side control (no PDT). (p<0.05, n = 4 animals).</p

PpIX fluorescence increased in tumor cells following mild hypothermia treatment.

<p>A) Point spectroscopy of PpIX fluorescence in live animals. Capital C stands for contralateral (non-tumor bearing hemisphere), while capital T is for the tumor-bearing hemisphere (p<0.05, n = 4). B) Tissue solubilization data of PpIX fluorescence between the same animals from panel A, following euthanasia (p<0.05, n = 4 animals).</p

Mild hypothermia leads to increases in edema/inflammation through the tumor volume following PDT treatment.

<p>A)T2 maps of 3 animals for hypothermia and normothermia PDT-treated at day 3. B) Mean T2 of the ROIs of T2 maps for each of the treatment conditions around the tumor treated area (p<0.01, n = 7 animals per group). C) The volume of inflammation reported as some voxels above baseline T2 in the PDT-treated area (p<0.05, n = 7 animals per group).</p