Mitochondrial autophagy eliminates damaged mitochondria and decreases reactive oxygen species (ROS). The autophagy inhibitor chloroquine (CQ) potentiates temozolomide (TMZ) cytotoxicity in glioma cells, but it is not known whether CQ does this by inhibiting mitochondrial autophagy. The effects of CQ and TMZ on MitoSOX Red fluorescence, a mitochondrial ROS indicator, and cell death were examined in rat C6 glioma cells. Mitochondrial autophagy was monitored by the colocalization of MitoTracker Red fluorescence and EGFP-LC3 dots. Mitochondrial content was measured by MitoTracker Green fluorescence and immunoblotting for a mitochondrial protein. Finally, CQ's effects on tumor cells derived from a glioblastoma patient and human U87-MG glioblastoma cells were assessed. TMZ (100-1,000 μM) alone did not affect mitochondrial ROS or cell death in C6 cells, but when administered with CQ (10 μM), it increased mitochondrial ROS and cell death. Antioxidants significantly suppressed the CQ-augmented cell death in TMZ-treated cells, indicating that mitochondrial ROS were involved in this cell death. TMZ treatment reduced MitoTracker Green fluorescence and mitochondrial protein levels, and these effects were inhibited by CQ. TMZ also increased the colocalization of EGFP-LC3 dots with mitochondria, and CQ enhanced this effect. CQ potentiated TMZ-induced cytotoxicity in patient-derived glioblastoma cells as well as human U87-MG glioblastoma cells. These results suggest that CQ increases cellular ROS and augments TMZ cytotoxicity in glioma cells by inhibiting mitochondrial autophagy.

Aims Cardiotoxicity by doxorubicin predicts worse prognosis of patients. Accumulation of damaged DNA has been implicated in doxorubicin-induced cardiotoxicity. SIRT1, an NAD+-dependent histone/protein deacetylase, protects cells by deacetylating target proteins. We investigated whether SIRT1 counteracts doxorubicin-induced cardiotoxicity by mediating Ser139 phosphorylation of histone H2AX, a critical signal of the DNA damage response. Methods and results Doxorubicin (5 mg/kg per week, x4) was administered to mice with intact SIRT1 (Sirt1f/f) and mice that lack SIRT1 activity in cardiomyocytes (Sirt1f/f; MHCcre/+). Reductions in left ventricular fractional shortening and ejection fraction by doxorubicin treatment were more severe in Sirt1f/f; MHCcre/+ than in Sirt1f/f. Myocardial expression level of type-B natriuretic peptide was 2.5-fold higher in Sirt1f/f; MHCcre/+ than in Sirt1f/f after doxorubicin treatment. Sirt1f/f; MHCcre/+ showed larger fibrotic areas and higher nitrotyrosine levels in the heart after doxorubicin treatment. Although doxorubicin-induced DNA damage evaluated by TUNEL staining was enhanced in Sirt1f/f; MHCcre/+, the myocardium from Sirt1f/f; MHCcre/+ showed blunted Ser139 phosphorylation of H2AX by doxorubicin treatment. In H9c2 cardiomyocytes, SIRT1 knockdown attenuated Ser139 phosphorylation of H2AX, increased DNA damage, and enhanced caspase-3 activation under doxorubicin treatment. Immunostaining revealed that acetylation level of H2AX at Lys5 was higher in hearts from Sirt1f/f; MHCcre/+. In H9c2 cells, acetyl-Lys5-H2AX level was increased by SIRT1 knockdown and reduced by SIRT1 overexpression. Ser139 phosphorylation in response to doxorubicin treatment was blunted in a mutant H2AX with substitution of Lys5 to Gln (K5Q) that mimics acetylated lysine compared with that in wild-type H2AX. Expression of K5Q-H2AX as well as S139A-H2AX, which cannot be phosphorylated at Ser139, augmented doxorubicin-induced caspase-3 activation. Treatment of mice with resveratrol, a SIRT1 activator, attenuated doxorubicin-induced cardiac dysfunction, which was associated with a reduction in acetyl-Lys5-H2AX level and a preserved phospho-Ser139-H2AX level. Conclusion These findings suggest that SIRT1 counteracts doxorubicin-induced cardiotoxicity by mediating H2AX phosphorylation through its deacetylation in cardiomyocytes. Translational perspective Doxorubicin-induced cardiotoxicity limits the further use of doxorubicin for cancer treatment and determines prognosis of patients. This work shows for the first time the protective effect of SIRT1, an NAD+-dependent deacetylase, on doxorubicin-induced cardiotoxicity using a genetically modified mouse model. We identified histone H2AX as a target of SIRT1 for proper DNA damage response. Therefore, DNA repair by SIRT1 could be a potential therapeutic target to attenuate doxorubicin cardiotoxicity. SIRT1 activity may also help to predict a risk of developing cardiotoxicity in patients treated with doxorubicin.

Participatory Video (PV) is emerging as a rich and valuable method for monitoring and evaluating (M & E) projects in the International Development sector. Although shown to be useful for engaging communities within short-term monitoring exercises or promotion, PV in these contexts presents signifcant complexity and logistical challenges for sustained uptake by Development organizations. In this paper, we present Our Story, a digitally mediated work fow iteratively designed and deployed on initiatives in Indonesia and Namibia. Developed in collaboration with the International Federation of Red Cross and Red Crescent (IFRC), it supports end-to-end PV production in the feld, and was specifcally developed to make PV a more sustainable tool for monitoring. We discuss and evaluate Our Story, reporting on how by lowering skills barriers for facilitators and leveraging consumer technology, PV can be delivered at scale. CCS CONCEPTS • Human-centered computing → Ubiquitous and mobile computing systems and tools.

Background: Cardiotoxicity of doxorubicin (DOX) is mediated through mitochondrial perturbations. SIRT1, an NAD + -dependent deacetylase, has been reported to promote cellular adaptation to metabolic imbalances in part via mitochondrial biogenesis. Hypothesis: SIRT1 in the cardiomyocyte counteracts DOX cardiotoxicity in vivo. Methods: Tamoxifen (Tam)-inducible cardiomyocyte-specific SIRT1 knockout mice (SIRT1 cKO) were used. At 2 months of age, mice received Tam to induce knockout, and mice without Cre recombinase served as wild type (WT). Both groups of mice were treated with DOX (4 IP injections of 5 mg/kg/week) starting at 3 months of age. Echocardiography was performed before and after DOX treatment. Cardiac tissues were sampled after the final DOX injection. H9c2 cardiomyocytes were used for in vitro analysis. Results: At baseline, there were no differences in left ventricular (LV) dimension, thickness, fractional shortening (FS), and ejection fraction (EF) between the groups. DOX significantly reduced FS, EF, and LV thickness in both groups of mice. However, FS and EF after DOX injections were lower in SIRT1 cKO than in WT (FS, 26% vs. 30%; EF, 59% vs. 65%, P<0.05). ANP and BNP mRNA levels were also higher in SIRT1 cKO (2.4-fold and 2.4-fold, respectively, P<0.05), confirming worsening of cardiac function. Compared with WT, SIRT1 cKO showed reduced cardiac mitochondrial content determined by qPCR analysis of mitochondrial DNA level (-16%, P<0.05). Unexpectedly, mRNA levels of nuclear-encoded mitochondria genes (Cox5b, Atp5a1, and citrate synthase) were not altered in the SIRT1 cKO heart, indicating preserved mitochondrial biogenesis in cKO. Instead, mRNA levels of parkin (+43% P<0.05), PINK1 (+18%), and Bnip3 (+40%) were higher in SIRT1 cKO than in WT, suggesting enhanced mitochondrial degradation via mitophagy. Finally, SIRT1 knockdown in H9c2 cells enhanced DOX (10 μM, 24 h)-induced cell death examined by LDH release compared with control cells (13.2±1.4% vs. 8.6±0.7%, P<0.05). Conclusions: The results suggested that SIRT1 in the cardiomyocyte counteracts DOX cardiotoxicity by preserving mitochondrial content in vivo. Enhanced degradation, but not impaired biogenesis, may be involved in mitochondrial depletion in SIRT1 cKO.