Functional association of NR4A3 downregulation with impaired differentiation in myeloid leukemogenesis.

The coincident downregulation of NR4A1 and NR4A3 has been implicated in myeloid leukemogenesis, but it remains unknown how these two genes function in myeloid cells and how their combined downregulation promotes myeloid leukemogenesis. Since NR4A1 abrogation is thought to confer a survival and proliferation advantage to myeloid cells, we hypothesized that downregulation of NR4A3 may have a complementary effect on myeloid cell differentiation. First, we tested the association between differentiation status of leukemic cells and NR4A3 expression using two large clinical datasets from patients with different acute myeloid leukemia (AML) subtypes. The analysis revealed a close association between differentiation status and different subtypes of AML Then, we probed the effects of differentiation-inducing treatments on NR4A3 expression and NR4A3 knockdown on cell differentiation using two myeloid leukemia cell lines. Differentiation-inducing treatments caused upregulation of NR4A3, while NR4A3 knockdown prevented differentiation in both cell lines. The cell culture findings were validated using samples from chronic myeloid leukemia (CML) patients at chronic, accelerated and blastic phases, and in acute promyelocytic leukemia (APL) patients before and after all trans-retinoic acid (ATRA)-based differentiation therapy. Progressive NR4A3 downregulation was coincident with impairments in differentiation in patients during progression to blastic phase of CML, and NR4A3 expression was increased in APL patients treated with ATRA-based differentiating therapy. Together, our findings demonstrate a tight association between impaired differentiation status and NR4A3 downregulation in myeloid leukemias, providing a plausible mechanistic explanation of how myeloid leukemogenesis might occur upon concurrent downregulation of NR4A1 and NR4A3.

Genetic variants that influence SARS-CoV-2 receptor TMPRSS2 expression among population cohorts from multiple continents.

The World Health Organization recently announced that pandemic status has been achieved for coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Exponential increases in patient numbers have been reported around the world, along with proportional increases in the number of COVID-19-related deaths. The SARS-CoV-2 infection rate in a population is expected to be influenced by social practices, availability of vaccines or prophylactics, and the prevalence of susceptibility genes in the population. Previous work revealed that cellular uptake of SARS-CoV-2 requires Angiotensin Converting Enzyme 2 (ACE-2) and a cellular protease. The spike (S) protein on SARS-CoV-2 binds ACE-2, which functions as an entry receptor. Following receptor binding, transmembrane protease serine 2 (encoded by TMPRSS2) primes the S protein to allow cellular uptake. Therefore, individual expression of TMPRSS2 may be a crucial determinant of SARS-CoV-2 infection susceptibility. Here, we utilized multiple large genome databases, including the GTEx portal, SNP nexus, and Ensembl genome project, to identify gene expression profiles for TMPRSS2 and its important expression quantitative trait loci. Our results show that four variants (rs464397, rs469390, rs2070788 and rs383510) affect expression of TMPRSS2 in lung tissue. The allele frequency of each variant was then assessed in regional populations, including African, American, European, and three Asian cohorts (China, Japan and Taiwan). Interestingly, our data shows that TMPRSS2-upregulating variants are at higher frequencies in European and American populations than in the Asian populations, which implies that these populations might be relatively susceptible to SARS-CoV-2 infection.

miR-105/93-3p promotes chemoresistance and circulating miR-105/93-3p acts as a diagnostic biomarker for triple negative breast cancer.

BACKGROUND: Triple negative breast cancer (TNBC) lacks both early detection biomarkers and viable targeted therapeutics. Moreover, chemotherapy only produces 20-30% pathologic complete response. Because miRNAs are frequently dysregulated in breast cancer and have broad tissue effects, individual or combinations of circulating miRNAs may serve as ideal diagnostic, predictive or prognostic biomarkers, as well as therapeutic targets. Understanding the role and mechanism of dysregulated miRNAs in TNBC may help to develop novel diagnostic and prognostic strategy for TNBC patients. METHODS: The miRNA array profiles of 1299 breast cancer patients were collected from the Metabric database and subjected to analysis of the altered miRNAs between TNBC and non-TNBC. In Student's t-test and Kaplan-Meier analysis, four upregulated miRNAs correlated with poor survival in TNBC but not in non-TNBC. Four miRNAs were manipulated in multiple cell lines to investigate their functional role in carcinogenesis. From these results, we studied miR-105 and miR-93-3p in greater detail. The level of miR-105 and miR-93-3p were evaluated in 25 breast cancer tumor tissues. In addition, the diagnostic utility of circulating miR-105 and miR-93-3p were examined in 12 normal and 118 breast cancer plasma samples by ROC curve construction. RESULTS: miR-105 and miR-93-3p were upregulated and correlated with poor survival in TNBC patients. Both miR-105 and miR-93-3p were found to activate Wnt/ß-catenin signaling by downregulation of SFPR1. By this action, stemness, chemoresistance, and metastasis were promoted. Importantly, the combination of circulating miR-105/93-3p may serve as a powerful biomarker for TNBC, even in early-stage disease. CONCLUSIONS: miR-105/93-3p activates Wnt/ß-catenin signaling by downregulating SFRP1 and thereby promotes stemness, chemoresistance, and metastasis in TNBC cells. Most importantly, combined circulating miR-105/93-3p levels represent a prime candidate for development into a diagnostic biomarker for both early- and late-stage TNBC.

Mitochondrial defects arise from nucleoside/nucleotide reverse transcriptase inhibitors in neurons: Potential contribution to HIV-associated neurocognitive disorders.

The cornerstone of current HIV treatment is a class of drugs called nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs). However, patients who receive long term treatment with NRTIs often develop severe side effects, which are related to mitochondrial toxicity. The potential contribution of NRTI-mediated toxicity to HIV-associated neurocognitive disorders (HAND) has not been fully explored. NRTI toxicity is thought to be mediated through mitochondrial DNA polymerase γ (pol γ) inhibition, which impairs mitochondrial DNA (mtDNA) synthesis and leads to various mitochondrial dysfunctions. To evaluate the relationship between NRTI-mediated pol γ inhibition and mitochondrial toxicity in neurons, we systematically investigated mitochondrial regulation in NRTI-treated primary cortical neurons by measuring parameters related to mtDNA content, retrograde signaling responses and mitochondrial homeostasis. The effects of four different NRTIs with variable pol γ inhibitory activity and mitochondrial toxicity were assessed. The strong pol γ inhibitor, ddI, abolished mtDNA synthesis and greatly reduced mtDNA content. However, mtDNA transcription was not as severely affected, and no defects in oxidative phosphorylation were observed. Detrimental effects on mitochondrial respiration and motility were observed after AZT treatment in the absence of mtDNA depletion or inhibition of mtDNA synthesis. The results suggest that individual NRTIs, such as ddI and AZT, have the potential to cause mitochondrial toxicity in neurons. This mitochondrial toxicity would be expected to contribute to neurotoxicity in the central nervous system, and therefore, HAND etiology may be affected by NRTI treatment.

Huntingtin-Interacting Protein-1 Is an Early-Stage Prognostic Biomarker of Lung Adenocarcinoma and Suppresses Metastasis via Akt-mediated Epithelial-Mesenchymal Transition.

RATIONALE: Non-small cell lung cancer (NSCLC) carries a poor survival rate mainly because of metastasis. However, the molecular mechanisms that govern NSCLC metastasis have not been described. Because huntingtin-interacting protein-1 (HIP1) is known to play a role in tumorigenesis, we tested the involvement of HIP1 in NSCLC progression and metastasis. OBJECTIVES: HIP1 expression was measured in human NSCLC tumors, and correlation with survival outcome was evaluated. Furthermore, we investigated the ability of HIP1 to suppress metastasis. The molecular mechanism by which HIP1 contributes to suppress metastasis was investigated. METHODS: We used tissue arrays containing samples from 121 patients with NSCLC to analyze HIP1 expression by immunohistochemistry. To investigate the role of HIP1 expression on metastasis, we evaluated cellular mobility, migration, and invasion using lung adenocarcinoma (AdCA) cells with modified HIP1 expression levels. The human disease mouse models with the same cells were applied to evaluate the HIP1 suppressing metastasis and its mechanism in vivo. MEASUREMENTS AND MAIN RESULTS: HIP1 expression in AdCA progression was found to be an early-stage prognostic biomarker, with low expression correlated to poor prognosis. We also found HIP1 to be a metastatic suppressor in AdCA. HIP1 significantly repressed the mobility of lung cancer cells in vitro and in vivo and regulated the epithelial-mesenchymal transition by repressing AKT/glycogen synthase kinase-3ß/ß-catenin signaling. CONCLUSIONS: HIP1 serves as an early-stage prognostic biomarker and a metastatic suppressor. Reduced expression during AdCA progression can relieve HIP1 suppression of Akt-mediated epithelial-mesenchymal transition and thereby lead to development of late metastases and poor prognosis.

Mutant huntingtin's interaction with mitochondrial protein Drp1 impairs mitochondrial biogenesis and causes defective axonal transport and synaptic degeneration in Huntington's disease.

The purpose of this study was to investigate the link between mutant huntingtin (Htt) and neuronal damage in relation to mitochondria in Huntington's disease (HD). In an earlier study, we determined the relationship between mutant Htt and mitochondrial dynamics/synaptic viability in HD patients. We found mitochondrial loss, abnormal mitochondrial dynamics and mutant Htt association with mitochondria in HD patients. In the current study, we sought to expand on our previous findings and further elucidate the relationship between mutant Htt and mitochondrial and synaptic deficiencies. We hypothesized that mutant Htt, in association with mitochondria, alters mitochondrial dynamics, leading to mitochondrial fragmentation and defective axonal transport of mitochondria in HD neurons. In this study, using postmortem HD brains and primary neurons from transgenic BACHD mice, we identified mutant Htt interaction with the mitochondrial protein Drp1 and factors that cause abnormal mitochondrial dynamics, including GTPase Drp1 enzymatic activity. Further, using primary neurons from BACHD mice, for the first time, we studied axonal transport of mitochondria and synaptic degeneration. We also investigated the effect of mutant Htt aggregates and oligomers in synaptic and mitochondrial deficiencies in postmortem HD brains and primary neurons from BACHD mice. We found that mutant Htt interacts with Drp1, elevates GTPase Drp1 enzymatic activity, increases abnormal mitochondrial dynamics and results in defective anterograde mitochondrial movement and synaptic deficiencies. These observations support our hypothesis and provide data that can be utilized to develop therapeutic targets that are capable of inhibiting mutant Htt interaction with Drp1, decreasing mitochondrial fragmentation, enhancing axonal transport of mitochondria and protecting synapses from toxic insults caused by mutant Htt.

Mitochondria-targeted catalase reduces abnormal APP processing, amyloid ß production and BACE1 in a mouse model of Alzheimer's disease: implications for neuroprotection and lifespan extension.

The purpose of this study was to investigate the protective effects of the mitochondria-targeted antioxidant catalase (MCAT) and lifespan extension in mice that express amyloid beta (Aß). Using immunoblotting and immunostaining analyses, we measured the production of full-length amyloid precursor protein (APP), soluble APPα, C-terminal fragments CTF99 and CTF83, monomeric and oligomeric Aß, Aß deposits and beta site amyloid precursor protein cleaving enzyme 1 (BACE1), in different stages of disease progression in MCAT/AßPP and AßPP mice. Using quantitative reverse transcriptase polymerase chain reaction and immunostaining analyses, we studied the expression of catalase, BACE1, the Alzheimer's disease (AD) markers, synaptophysin, APP, neprilysin, insulin-degrading enzyme and transthyretin in MCAT, AßPP, MCAT/AßPP and wild-type (WT) mice. Using the high pressure liquid chromatography analysis of 8-hydroxy-2-deoxyguanosine, we measured oxidative DNA damage in the cerebral cortical tissues from MCAT, AßPP, MCAT/AßPP and WT mice. We found that the AßPP transgenic mice that carried the human MCAT gene lived 5 months longer than did the AßPP mice. We also found that the overexpression of MCAT in the brain sections from the MCAT/AßPP transgenic mice significantly correlated with a reduction in the levels of full-length APP, CTF99, BACE1, Aß levels (40 and 42), Aß deposits and oxidative DNA damage relative to the brain sections from the AßPP mice. Interestingly, we found significantly increased levels of soluble APPα and CTF83 in the MCAT/AßPP mice, relative to the AßPP mice. These data provide direct evidence that oxidative stress plays a primary role in AD etiopathology and that in MCAT mice express Aß, MCAT prevents abnormal APP processing, reduces Aß levels and enhances Aß-degrading enzymes in mice at different ages, corresponding to different stages of disease progression. These findings indicate that mitochondria-targeted molecules may be an effective therapeutic approach to treat patients with AD.

Abnormal mitochondrial dynamics and synaptic degeneration as early events in Alzheimer's disease: implications to mitochondria-targeted antioxidant therapeutics.

Synaptic pathology and mitochondrial oxidative damage are early events in Alzheimer's disease (AD) progression. Loss of synapses and synaptic damage are the best correlates of cognitive deficits found in AD patients. Recent research on amyloid beta (Aß) and mitochondria in AD revealed that Aß accumulates in synapses and synaptic mitochondria, leading to abnormal mitochondrial dynamics and synaptic degeneration in AD neurons. Further, recent studies using live-cell imaging and primary neurons from amyloid beta precursor protein (AßPP) transgenic mice revealed reduced mitochondrial mass, defective axonal transport of mitochondria and synaptic degeneration, indicating that Aß is responsible for mitochondrial and synaptic deficiencies. Tremendous progress has been made in studying antioxidant approaches in mouse models of AD and clinical trials of AD patients. This article highlights the recent developments made in Aß-induced abnormal mitochondrial dynamics, defective mitochondrial biogenesis, impaired axonal transport and synaptic deficiencies in AD. This article also focuses on mitochondrial approaches in treating AD, and also discusses latest research on mitochondria-targeted antioxidants in AD. This article is part of a Special Issue entitled: Antioxidants and Antioxidant Treatment in Disease.

Impaired mitochondrial dynamics and abnormal interaction of amyloid beta with mitochondrial protein Drp1 in neurons from patients with Alzheimer's disease: implications for neuronal damage.

The purpose of our study was to better understand the relationship between mitochondrial structural proteins, particularly dynamin-related protein 1 (Drp1) and amyloid beta (Aß) in the progression of Alzheimer's disease (AD). Using qRT-PCR and immunoblotting analyses, we measured mRNA and protein levels of mitochondrial structural genes in the frontal cortex of patients with early, definite and severe AD and in control subjects. We also characterized monomeric and oligomeric forms of Aß in these patients. Using immunoprecipitation/immunoblotting analysis, we investigated the interaction between Aß and Drp1. Using immunofluorescence analysis, we determined the localization of Drp1 and intraneuronal and oligomeric Aß in the AD brains and primary hippocampal neurons from Aß precursor protein (AßPP) transgenic mice. We found increased expression of the mitochondrial fission genes Drp1 and Fis1 (fission 1) and decreased expression of the mitochondrial fusion genes Mfn1 (mitofusin 1), Mfn2 (mitofusin 2), Opa1 (optic atrophy 1) and Tomm40. The matrix gene CypD was up-regulated in AD patients. Results from our qRT-PCR and immunoblotting analyses suggest that abnormal mitochondrial dynamics increase as AD progresses. Immunofluorescence analysis of the Drp1 antibody and the Aß antibodies 6E10 and A11 revealed the colocalization of Drp1 and Aß. Drp1 immunoprecipitation/immunoblotting analysis of Aß antibodies 6E10 and A11 revealed that Drp1 interacts with Aß monomers and oligomers in AD patients, and these abnormal interactions are increased with disease progression. Primary neurons that were found with accumulated oligomeric Aß had lost branches and were degenerated, indicating that oligomeric Aß may cause neuronal degeneration. These findings suggest that in patients with AD, increased production of Aß and the interaction of Aß with Drp1 are crucial factors in mitochondrial fragmentation, abnormal mitochondrial dynamics and synaptic damage. Inhibiting, these abnormal interactions may be a therapeutic strategy to reduce mitochondrial fragmentation, neuronal and synaptic damage and cognitive decline in patients with AD.

Impaired mitochondrial biogenesis, defective axonal transport of mitochondria, abnormal mitochondrial dynamics and synaptic degeneration in a mouse model of Alzheimer's disease.

Increasing evidence suggests that the accumulation of amyloid beta (Aß) in synapses and synaptic mitochondria causes synaptic mitochondrial failure and synaptic degeneration in Alzheimer's disease (AD). The purpose of this study was to better understand the effects of Aß in mitochondrial activity and synaptic alterations in neurons from a mouse model of AD. Using primary neurons from a well-characterized Aß precursor protein transgenic (AßPP) mouse model (Tg2576 mouse line), for the first time, we studied mitochondrial activity, including axonal transport of mitochondria, mitochondrial dynamics, morphology and function. Further, we also studied the nature of Aß-induced synaptic alterations, and cell death in primary neurons from Tg2576 mice, and we sought to determine whether the mitochondria-targeted antioxidant SS31 could mitigate the effects of oligomeric Aß. We found significantly decreased anterograde mitochondrial movement, increased mitochondrial fission and decreased fusion, abnormal mitochondrial and synaptic proteins and defective mitochondrial function in primary neurons from AßPP mice compared with wild-type (WT) neurons. Transmission electron microscopy revealed a large number of small mitochondria and structurally damaged mitochondria, with broken cristae in AßPP primary neurons. We also found an increased accumulation of oligomeric Aß and increased apoptotic neuronal death in the primary neurons from the AßPP mice relative to the WT neurons. Our results revealed an accumulation of intraneuronal oligomeric Aß, leading to mitochondrial and synaptic deficiencies, and ultimately causing neurodegeneration in AßPP cultures. However, we found that the mitochondria-targeted antioxidant SS31 restored mitochondrial transport and synaptic viability, and decreased the percentage of defective mitochondria, indicating that SS31 protects mitochondria and synapses from Aß toxicity.

Abnormal mitochondrial dynamics, mitochondrial loss and mutant huntingtin oligomers in Huntington's disease: implications for selective neuronal damage.

The purpose of our study was to determine the relationship between mutant huntingtin (Htt) and mitochondrial dynamics in the progression of Huntington's disease (HD). We measured the mRNA levels of electron transport chain genes, and mitochondrial structural genes, Drp1 (dynamin-related protein 1), Fis1 (fission 1), Mfn1 (mitofusin 1), Mfn2 (mitofusin 2), Opa1 (optric atrophy 1), Tomm40 (translocase of outermembrane 40) and CypD (cyclophilin D) in grade III and grade IV HD patients and controls. The mutant Htt oligomers and the mitochondrial structural proteins were quantified in the striatum and frontal cortex of HD patients. Changes in expressions of the electron transport chain genes were found in HD patients and may represent a compensatory response to mitochondrial damage caused by mutant Htt. Increased expression of Drp1 and Fis1 and decreased expression of Mfn1, Mfn2, Opa1 and Tomm40 were found in HD patients relative to the controls. CypD was upregulated in HD patients, and this upregulation increased as HD progressed. Significantly increased immunoreactivity of 8-hydroxy-guanosine was found in the cortical specimens from stage III and IV HD patients relative to controls, suggesting increased oxidative DNA damage in HD patients. In contrast, significantly decreased immunoreactivities of cytochrome oxidase 1 and cytochrome b were found in HD patients relative to controls, indicating a loss of mitochondrial function in HD patients. Immunoblotting analysis revealed 15, 25 and 50 kDa mutant Htt oligomers in the brain specimens of HD patients. All oligomeric forms of mutant Htt were significantly increased in the cortical tissues of HD patients, and mutant Htt oligomers were found in the nucleus and in mitochondria. The increase in Drp1, Fis1 and CypD and the decrease in Mfn1 and Mfn2 may be responsible for abnormal mitochondrial dynamics that we found in the cortex of HD patients, and may contribute to neuronal damage in HD patients. The presence of mutant Htt oligomers in the nucleus of HD neurons and in mitochondria may disrupt neuronal functions. Based on these findings, we propose that mutant Htt in association with mitochondria imbalance and mitochondrial dynamics impairs axonal transport of mitochondria, decreases mitochondrial function and damages neurons in affected brain regions of HD patients.

Amyloid beta impairs mitochondrial anterograde transport and degenerates synapses in Alzheimer's disease neurons.

Loss of synapses and synaptic damage are the best correlates of cognitive decline identified in patients with Alzheimer's disease (AD), and mitochondrial oxidative damage and synaptic pathology have been identified as early events in the progression of AD. The progressive accumulation of amyloid beta (Aß) in synapses and synaptic mitochondria are hypothesized to cause synaptic degeneration and cognitive decline in patients with AD. However, the precise mechanistic link between Aß and mitochondria is not well understood. The purpose of this study was to better understand the effects of Aß on mitochondrial axonal transport and synaptic alterations in AD. Using mouse hippocampal neurons and Aß(25-35) peptide, we studied axonal transport of mitochondria, including mitochondrial motility, mitochondrial length and size, mitochondrial index per neurite, and synaptic alterations of the hippocampal neurons. In the PBS-treated neurons, 36.4±4.7% of the observed mitochondria were motile, with 21.0±1.3% moving anterograde and 15.4±3.4% moving retrograde and the average speed of movement was 12.1±1.8µm/min. In contrast, in the Aß-treated neurons, the number of motile mitochondria were significantly less, at 20.4±2.6% (P<0.032), as were those moving anterograde (10.1±2.6%, P<0.016) relative to PBS-treated neurons, suggesting that the Aß(25-35) peptide impairs axonal transport of mitochondria in AD neurons. In the Aß-treated neurons, the average speed of motile mitochondria was also less, at 10.9±1.9µm/min, and mitochondrial length was significantly decreased. Further, synaptic immunoreactivity was also significantly less in the Aß-treated neurons relative to the PBS-treated neurons, indicating that Aß affects synaptic viability. These findings suggest that, in neurons affected by AD, Aß is toxic, impairs mitochondrial movements, reduces mitochondrial length, and causes synaptic degeneration.

Assessment of newly synthesized mitochondrial DNA using BrdU labeling in primary neurons from Alzheimer's disease mice: Implications for impaired mitochondrial biogenesis and synaptic damage.

The purpose of our study was to assess mitochondrial biogenesis and distribution in murine primary neurons. Using 5-bromo-2-deoxyuridine (BrdU) incorporation and primary neurons, we studied the mitochondrial biogenesis and mitochondrial distribution in hippocampal neurons from amyloid beta precursor protein (AßPP) transgenic mice and wild-type (WT) neurons treated with oxidative stressors, rotenone and H(2)O(2). We found that after 20h of labeling, BrdU incorporation was specific to porin-positive mitochondria. The proportion of mitochondrial area labeled with BrdU was 40.3±6.3% at 20h. The number of mitochondria with newly synthesized DNA was higher in AßPP neuronal cell bodies than in the cell bodies of WT neurons (AßPP, 45.23±2.67 BrdU-positive/cell body; WT, 32.92±2.49 BrdU-positive/cell body; p=0.005). In neurites, the number of BrdU-positive mitochondria decreased in AßPP cultures compared to WT neurons (AßPP, 0.105±0.008 BrdU-positive/µm neurite; WT, 0.220±0.036 BrdU-positive/µm neurite; p=0.010). Further, BrdU in the cell body increased when neurons were treated with low doses of H(2)O(2) (49.6±2.7 BrdU-positive/cell body, p=0.0002 compared to untreated cells), while the neurites showed decreased BrdU staining (0.122±0.010 BrdU-positive/µm neurite, p=0.005 compared to the untreated). BrdU labeling was increased in the cell body under rotenone treatment. Additionally, under rotenone treatment, the content of BrdU labeling decreased in neurites. These findings suggest that Aß and mitochondrial toxins enhance mitochondrial fragmentation in the cell body, and may cause impaired axonal transport of mitochondria leading to synaptic degeneration.

Endonuclease VIII-like 1 deficiency impairs survival of newly generated hippocampal neurons and memory performance in young-adult male mice.

AIMS: Efficient memory formation in rodents depends on adult neurogenesis in the subgranular zone of the hippocampus, and mounting evidence suggests that deficiencies in initiating repair of oxidatively induced DNA damage may impair neurogenesis. Hence, we aimed to determine whether loss of the DNA glycosylase, endonuclease VIII-like 1 (Neil1), affects hippocampal neurogenesis and memory performance in young-adult mice. MAIN METHODS: Eight-week-old male wild-type and Neil1-deficient (Neil1-/-) mice were treated with bromodeoxyuridine to track neuronal proliferation and differentiation. A neurosphere formation assay was further used to measure neuroprogenitor proliferative capacity. Hippocampus-related memory functions were assessed with Y-maze spontaneous alternation and novel object recognition tests. KEY FINDINGS: Young-adult male Neil1-/- mice exhibited diminished adult hippocampal neurogenesis in the dentate gyrus, probably as a result of poor survival of newly proliferated neurons. Furthermore, the Y-maze spontaneous alternation and novel object recognition tests respectively revealed that Neil1 deficiency impairs spatial and non-spatial hippocampus-related memory functions. We also found that expression of p53, a central regulator of apoptosis, was upregulated in the dentate gyrus of Neil1-/- mice, while the level of ß-catenin, a key cell survival molecule, was downregulated. SIGNIFICANCE: The DNA glycosylase, Neil1, promotes successful hippocampal neurogenesis and learning and memory in young-adult mice.

Gas7 knockout affects PINK1 expression and mitochondrial dynamics in mouse cortical neurons.

Dynamic fission and fusion events regulate mitochondrial shape, distribution, and rejuvenation, and proper control of these processes is essential for neuronal homeostasis. Here, we report that Gas7, a known cytoskeleton regulator, controls mitochondrial dynamics within neurons of the central nervous system. In this study, we generated an improved Gas7-knockout mouse and evaluated its mitochondrial phenotype. We first identified Gas7 in mitochondrial fractions from wild-type brain tissue, and observed Gas7 colocalization with mitochondria in primary cortical neurons. In Gas7-deficient brain tissue and neuronal cultures mitochondria were elongated with perinuclear clustering. These morphological abnormalities were associated with increased levels mitochondrial fusion proteins and increased PKA-dependent phosphorylation of Drp-1 in brain tissues, suggesting an imbalance of mitochondrial fusion and fission. Moreover, expression of mitochondrial quality control kinase, PINK1, and PINK1-specific phosphorylation of Mfn-2 (S442), Parkin (S65), and ubiquitin (S65) were all reduced in the knockout cells. Ectopic expression of Gas7 restored mitochondrial morphology and distribution, as well as PINK1 expression in Gas7-null cortical neurons. Collectively, our results introduce a novel role of mouse Gas7 in determining the dynamics, morphology, and intracellular distribution of neuronal mitochondria, which are expected to be required for normal neuronal function.

High-CLDN4 ESCC cells harbor stem-like properties and indicate for poor concurrent chemoradiation therapy response in esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is the major type of esophageal cancer in Asia and demonstrates poor survival rates following a therapeutic regimen. METHODS: Cancer stem cells (CSCs) are responsible for tumor initiation, progression, and treatment failure in cancers. Therefore, identification and characterization of CSCs may help to improve clinical outcomes for ESCC patients. Tumor sphere formation assay are performed to isolate cancer stem-like ESCC cells. QRT-PCR, tumor initiation, metastasis, CCRT treatment are used to evaluate ESCC cells' stemness properties in vitro and in vivo. RESULTS: The authors' data demonstrates that cancer stem-like ESCC cells harbored stemness characteristics including self-renewal, differentiation, and transdifferentiation, and possess tumor initiation, metastasis, and treatment inefficiency properties. For the identification of useful biomarkers of cancer stem-like ESCC cells, the authors further identified that CLDN4 was upregulated in cancer stem-like ESCC cells when compared with bulk cancer cells. High-CLDN4 cells harbored stemness and cisplatin/concurrent chemoradiation therapy (CCRT) resistance properties and a high level of CLDN4 was correlated with poor prognosis and poor CCRT response in ESCC patients. Importantly, thiamine tetrahydrofurfuryl disulfide (TTFD) decreased CLDN4 and attenuated stemness in ESCC cells, and TTFD combined with CCRT improved CCRT response in vivo. CONCLUSIONS: CLDN4 was suggested as prognostic and a CCRT response indicator for ESCC patients. TTFD combined with CCRT has potential to improve ESCC patient's clinical outcomes in the future.

Modulation of UVB-induced Carcinogenesis by Activation of Alternative DNA Repair Pathways.

The molecular basis for ultraviolet (UV) light-induced nonmelanoma and melanoma skin cancers centers on cumulative genomic instability caused by inefficient DNA repair of dipyrimidine photoproducts. Inefficient DNA repair and subsequent translesion replication past these DNA lesions generate distinct molecular signatures of tandem CC to TT and C to T transitions at dipyrimidine sites. Since previous efforts to develop experimental strategies to enhance the repair capacity of basal keratinocytes have been limited, we have engineered the N-terminally truncated form (Δ228) UV endonuclease (UVDE) from Schizosaccharomyces pombe to include a TAT cell-penetrating peptide sequence with or without a nuclear localization signal (NLS): UVDE-TAT and UVDE-NLS-TAT. Further, a NLS was engineered onto a pyrimidine dimer glycosylase from Paramecium bursaria chlorella virus-1 (cv-pdg-NLS). Purified enzymes were encapsulated into liposomes and topically delivered to the dorsal surface of SKH1 hairless mice in a UVB-induced carcinogenesis study. Total tumor burden was significantly reduced in mice receiving either UVDE-TAT or UVDE-NLS-TAT versus control empty liposomes and time to death was significantly reduced with the UVDE-NLS-TAT. These data suggest that efficient delivery of exogenous enzymes for the initiation of repair of UVB-induced DNA damage may protect from UVB induction of squamous and basal cell carcinomas.

Role of Nrf2-dependent ARE-driven antioxidant pathway in neuroprotection.

The promoter regions of many detoxification enzymes contain a cis-acting enhancer known as the antioxidant response element (ARE). NF-E2-related factor 2 (Nrf2) is considered as one of the major transcription factors for the ARE. Nrf2-dependent transcriptional activation by means of the ARE is known to coordinate the upregulation of these antioxidant enzymes involved in combating oxidative stress and has been shown to be protective against neural toxicants. The mitochondrial complex II inhibitor malonate causes striatal damage reminiscent of Huntington's disease and is known to involve oxidative stress in its pathogenesis. In order to achieve a systemic upregulation of antioxidant potential in local striatal region, a cell-based, Nrf2-dependent antioxidant gene therapy is performed to attenuate malonate-induced neuronal cell death. The details for generating Nrf2-overexpressing astrocytes and grafting them onto the lesion model are described in this chapter.

CTMP, a predictive biomarker for trastuzumab resistance in HER2-enriched breast cancer patient.

Trastuzumab is regarded as the primary therapy for patients with HER2-enriched breast cancer, but the pathological complete response for advanced cases is less than 30%. The underlying mechanism of trastuzumab resistance remains unclear and there are currently no conclusive biomarkers for patient response to trastuzumab. Identifying predictive biomarkers for trastuzumab response may allow treatments to be individually tailored and optimized multi-target therapies may be developed. CTMP activates AKT signaling in breast cancer and over-activation of AKT has been reported to contribute to trastuzumab resistance. In this study, we examined samples from 369 patients to investigate the correlation between CTMP expression level and patient outcome. Elevated CTMP expression was correlated with adverse outcomes in HER2-enriched patients including overall and disease-free survival as well as trastuzumab resistance. Ectopic expression of varying levels of CTMP in SkBR3 cells dose-dependently attenuated trastuzumab-mediated growth inhibition through AKT activation. In addition, inhibition of AKT signaling by AKT inhibitor IV and Rapamycin reversed CTMP-mediated trastuzumab resistance. In clinical samples, the high expression of CTMP was showed in trastuzumab non-responders and positively correlated with AKT activity. Taken together, we demonstrated that CTMP promotes AKT activation resulting in trastuzumab resistance in patients with HER2-enriched breast cancer. High CTMP expression not only predicted poor prognosis, but may also predict resistance to trastuzumab in HER2-enriched patients. Therefore, CTMP expression may be considered as a prognostic biomarker in HER2-enriched breast cancer and high expression may indicate a utility for AKT-inhibition in these patients.

Nrf2, a multi-organ protector?

NF-E2-related factor 2 (Nrf2) is a basic leucine zipper transcription factor that binds to the promoter sequence "antioxidant responsive element (ARE)" leading to coordinated up-regulation of ARE-driven detoxification and antioxidant genes. Since the expression of a wide array of antioxidant and detoxification genes are positively regulated by the ARE sequence, Nrf2 may serve as a master regulator of the ARE-driven cellular defense system against oxidative stress. In support of this, numerous studies have shown that Nrf2 protects many cell types and organ systems from a broad spectrum of toxic insults and disease pathogenesis. This Nrf2-conferred, multi-organ protection phenomenon raises an interesting question about how a single protein can protect many different organs from various toxic insults. A possible molecular mechanism explaining this phenomenon is that Nrf2 protects many different cell types by coordinately up-regulating classic ARE-driven genes as well as cell type-specific target genes that are required for the defense system of each cell type in its unique environment. This hypothesis is supported by microarray data indicating the protective role of Nrf2 is conveyed through both known ARE-driven genes and novel cell type-specific genes. The widespread nature of Nrf2 may have an important therapeutic potential, allowing prevention of carcinogenesis and neurodegenerative diseases.