Reduced Mitochondrial Protein Translation Promotes Cardiomyocyte Proliferation and Heart Regeneration.

BACKGROUND: The importance of mitochondria in normal heart function are well recognized and recent studies have implicated changes in mitochondrial metabolism with some forms of heart disease. Previous studies demonstrated that knockdown of the mitochondrial ribosomal protein S5 (MRPS5) by small interfering RNA (siRNA) inhibits mitochondrial translation and thereby causes a mitonuclear protein imbalance. Therefore, we decided to examine the effects of MRPS5 loss and the role of these processes on cardiomyocyte proliferation. METHODS: We deleted a single allele of MRPS5 in mice and used left anterior descending coronary artery ligation surgery to induce myocardial damage in these animals. We examined cardiomyocyte proliferation and cardiac regeneration both in vivo and in vitro. Doxycycline treatment was used to inhibit protein translation. Heart function in mice was assessed by echocardiography. Quantitative real-time polymerase chain reaction and RNA sequencing were used to assess changes in transcription and chromatin immunoprecipitation (ChIP) and BioChIP were used to assess chromatin effects. Protein levels were assessed by Western blotting and cell proliferation or death by histology and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assays. Adeno-associated virus was used to overexpress genes. The luciferase reporter assay was used to assess promoter activity. Mitochondrial oxygen consumption rate, ATP levels, and reactive oxygen species were also analyzed. RESULTS: We determined that deletion of a single allele of MRPS5 in mice results in elevated cardiomyocyte proliferation and cardiac regeneration; this observation correlates with improved cardiac function after induction of myocardial infarction. We identified ATF4 (activating transcription factor 4) as a key regulator of the mitochondrial stress response in cardiomyocytes from Mrps5+/- mice; furthermore, ATF4 regulates Knl1 (kinetochore scaffold 1) leading to an increase in cytokinesis during cardiomyocyte proliferation. The increased cardiomyocyte proliferation observed in Mrps5+/- mice was attenuated when one allele of Atf4 was deleted genetically (Mrps5+/-/Atf4+/-), resulting in the loss in the capacity for cardiac regeneration. Either MRPS5 inhibition (or as we also demonstrate, doxycycline treatment) activate a conserved regulatory mechanism that increases the proliferation of human induced pluripotent stem cell-derived cardiomyocytes. CONCLUSIONS: These data highlight a critical role for MRPS5/ATF4 in cardiomyocytes and an exciting new avenue of study for therapies to treat myocardial injury.

A defect in mitochondrial protein translation influences mitonuclear communication in the heart.

The regulation of the informational flow from the mitochondria to the nucleus (mitonuclear communication) is not fully characterized in the heart. We have determined that mitochondrial ribosomal protein S5 (MRPS5/uS5m) can regulate cardiac function and key pathways to coordinate this process during cardiac stress. We demonstrate that loss of Mrps5 in the developing heart leads to cardiac defects and embryonic lethality while postnatal loss induces cardiac hypertrophy and heart failure. The structure and function of mitochondria is disrupted in Mrps5 mutant cardiomyocytes, impairing mitochondrial protein translation and OXPHOS. We identify Klf15 as a Mrps5 downstream target and demonstrate that exogenous Klf15 is able to rescue the overt defects and re-balance the cardiac metabolome. We further show that Mrps5 represses Klf15 expression through c-myc, together with the metabolite L-phenylalanine. This critical role for Mrps5 in cardiac metabolism and mitonuclear communication highlights its potential as a target for heart failure therapies.

Optimizing DUS testing for Chimonanthus praecox using feature selection based on a genetic algorithm.

Chimonanthus praecox is a famous traditional flower in China with high ornamental value. It has numerous varieties, yet its classification is highly disorganized. The distinctness, uniformity, and stability (DUS) test enables the classification and nomenclature of various species; thus, it can be used to classify the Chimonanthus varieties. In this study, flower traits were quantified using an automatic system based on pattern recognition instead of traditional manual measurement to improve the efficiency of DUS testing. A total of 42 features were quantified, including 28 features in the DUS guidelines and 14 new features proposed in this study. Eight algorithms were used to classify wintersweet, and the random forest (RF) algorithm performed the best when all features were used. The classification accuracy of the outer perianth was the highest when the features of the different parts were used for classification. A genetic algorithm was used as the feature selection algorithm to select a set of 22 reduced core features and improve the accuracy and efficiency of the classification. Using the core feature set, the classification accuracy of the RF model improved to 99.13%. Finally, K-means was used to construct a pedigree cluster tree of 23 varieties of wintersweet; evidently, wintersweet was clustered into a single class, which can be the basis for further study of genetic relationships among varieties. This study provides a novel method for DUS detection, variety identification, and pedigree analysis.

Clinical Variables, Deep Learning and Radiomics Features Help Predict the Prognosis of Adult Anti-N-methyl-D-aspartate Receptor Encephalitis Early: A Two-Center Study in Southwest China.

Objective: To develop a fusion model combining clinical variables, deep learning (DL), and radiomics features to predict the functional outcomes early in patients with adult anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis in Southwest China. Methods: From January 2012, a two-center study of anti-NMDAR encephalitis was initiated to collect clinical and MRI data from acute patients in Southwest China. Two experienced neurologists independently assessed the patients' prognosis at 24 moths based on the modified Rankin Scale (mRS) (good outcome defined as mRS 0-2; bad outcome defined as mRS 3-6). Risk factors influencing the prognosis of patients with acute anti-NMDAR encephalitis were investigated using clinical data. Five DL and radiomics models trained with four single or combined four MRI sequences (T1-weighted imaging, T2-weighted imaging, fluid-attenuated inversion recovery imaging and diffusion weighted imaging) and a clinical model were developed to predict the prognosis of anti-NMDAR encephalitis. A fusion model combing a clinical model and two machine learning-based models was built. The performances of the fusion model, clinical model, DL-based models and radiomics-based models were compared using the area under the receiver operating characteristic curve (AUC) and accuracy and then assessed by paired t-tests (P < 0.05 was considered significant). Results: The fusion model achieved the significantly greatest predictive performance in the internal test dataset with an AUC of 0.963 [95% CI: (0.874-0.999)], and also significantly exhibited an equally good performance in the external validation dataset, with an AUC of 0.927 [95% CI: (0.688-0.975)]. The radiomics_combined model (AUC: 0.889; accuracy: 0.857) provided significantly superior predictive performance than the DL_combined (AUC: 0.845; accuracy: 0.857) and clinical models (AUC: 0.840; accuracy: 0.905), whereas the clinical model showed significantly higher accuracy. Compared with all single-sequence models, the DL_combined model and the radiomics_combined model had significantly greater AUCs and accuracies. Conclusions: The fusion model combining clinical variables and machine learning-based models may have early predictive value for poor outcomes associated with anti-NMDAR encephalitis.

LncRNA LncHrt preserves cardiac metabolic homeostasis and heart function by modulating the LKB1-AMPK signaling pathway.

Metabolic modulation is a promising therapeutic approach to prevent adverse remodeling of the ischemic heart. Because little is known about the involvement of long non-coding RNAs (lncRNAs) in regulating cardiac metabolism, we used unbiased transcriptome profiling in a mouse model of myocardial infarction (MI). We identified a novel cardiomyocyte-enriched lncRNA, called LncHrt, which regulates metabolism and the pathophysiological processes that lead to heart failure. AAV-based LncHrt overexpression protects the heart from MI as demonstrated by improved contractile function, preserved metabolic homeostasis, and attenuated maladaptive remodeling responses. RNA-pull down followed by mass spectrometry and RNA immunoprecipitation (RIP) identified SIRT2 as a LncHrt-interacting protein involved in cardiac metabolic regulation. Mechanistically, we established that LncHrt interacts with SIRT2 to preserve SIRT2 deacetylase activity by interfering with the CDK5 and SIRT2 interaction. This increases downstream LKB1-AMPK kinase signaling, which ameliorates functional and metabolic deficits. Importantly, we found the expression of the human homolog of mouse LncHrt was decreased in patients with dilated cardiomyopathy. Together, these studies identify LncHrt as a cardiac metabolic regulator that plays an essential role in preserving heart function by regulating downstream metabolic signaling pathways. Consequently, LncHrt is a potentially novel RNA-based therapeutic target for ischemic heart disease.

Long noncoding RNA Cfast regulates cardiac fibrosis.

Cardiac fibrosis occurs in most cardiac diseases, which reduces cardiac muscle compliance, impairs both systolic and diastolic heart function and, ultimately, leads to heart failure. Long noncoding RNAs (lncRNAs) have recently emerged as important regulators of a variety of biological processes; however, little is known about the expression and function of lncRNAs in cardiac fibrosis. Using unbiased transcriptome profiling in a mouse model of myocardial infarction (MI), we identified a cardiac fibroblast-enriched lncRNA (AK048087) named cardiac fibroblast-associated transcript (Cfast), which is significantly elevated after MI. Silencing Cfast expression by small interfering RNAs (siRNAs) or lentiviral short hairpin RNAs (shRNAs) resulted in suppression of fibrosis-related gene expression and transdifferentiation of myofibroblasts into cardiac fibroblasts. Depletion of Cfast by lentiviral shRNAs in mouse hearts significantly attenuated cardiac fibrosis induced by MI or isoproterenol-infusion. Importantly, inhibition of Cfast ameliorated cardiac function following cardiac injury. RNA pull-down followed by mass spectrometry analyses identified COTL1 (coactosin-like 1) as one of the Cfast interacting proteins. Mechanistically, Cfast competitively inhibits the COTL1 interaction with TRAP1 (transforming growth factor-ß receptor-associated protein 1), which enhances TGF-ß signaling by augmenting SMAD2/SMAD4 complex formation. Therefore, our study identifies Cfast as a novel cardiac fibroblast-enriched lncRNA that regulates cardiac fibroblast activation in response to pathophysiological stress. Cfast could serve as a potential therapeutic target for the prevention of cardiac fibrosis and cardiac diseases.

Non-coding RNAs in cardiomyocyte proliferation and cardiac regeneration: Dissecting their therapeutic values.

Cardiovascular diseases are associated with high incidence and mortality, contribute to disability and place a heavy economic burden on countries worldwide. Stimulating endogenous cardiomyocyte proliferation and regeneration has been considering as a key to repair the injured heart caused by ischaemia. Emerging evidence has proved that non-coding RNAs participate in cardiac proliferation and regeneration. In this review, we focus on the observation and mechanism that microRNAs (or miRNAs), long non-coding RNAs (or lncRNAs) and circular RNA (or circRNAs) regulate cardiomyocyte proliferation and regeneration to repair a damaged heart. Furthermore, we highlight the potential therapeutic role of some non-coding RNAs used in stimulating CMs proliferation. Finally, perspective on the development of non-coding RNAs therapy in cardiac regeneration is presented.

LPA3-mediated lysophosphatidic acid signaling promotes postnatal heart regeneration in mice.

Background: Lysophosphatidic acid (LPA) is a small glycerophospholipid that acts as a potent extracellular signal in various biological processes and diseases. Our previous work demonstrated that the expression of the LPA receptors LPA1 and LPA3 is elevated in the early postnatal heart. However, the role of this stage-specific expression of LPA1 and LPA3 in the heart is unknown. Methods and Results: By using LPA3 and LPA1 knockout mice, and neonatal SD rats treated with Ki16425 (LPA1/LPA3 inhibitor), we found that the number of proliferating cardiomyocytes, detected by coimmunostaining pH3, Ki67 or BrdU with cardiac troponin T, was significantly decreased in the LPA3 knockout mice and the Ki16425-treated rats but not in the LPA1 knockout mice during the first week of postnatal life. Using a myocardial infarction (MI) model, we found that cardiac function and the number of proliferating cardiomyocytes were decreased in the neonatal LPA3 KO mice and increased in the AAV9-mediated cardiac-specific LPA3 overexpression mice. By using lineage tracing and AAV9-LPA3, we further found that LPA3 overexpression in adult mice enhances cardiac function and heart regeneration as assessed by pH3-, Ki67-, and Aurora B-positive cardiomyocytes and clonal cardiomyocytes after MI. Genome-wide transcriptional profiling and additional mechanistic studies showed that LPA induces cardiomyocyte proliferation through the PI3K/AKT, BMP-Smad1/5, Hippo/YAP and MAPK/ERK pathways in vitro, whereas only ERK was confirmed to be activated by LPA-LPA3 signaling in vivo. Conclusion: Our study reports that LPA3-mediated LPA signaling is a crucial factor for cardiomyocyte proliferation in the early postnatal heart. Cardiac-specific LPA3 overexpression improved cardiac function and promoted cardiac regeneration after myocardial injury induced by MI. This finding suggested that activation of LPA3 potentially through AAV-mediated gene therapy might be a therapeutic strategy to improve the outcome after MI.

SUMOylation and deacetylation affect NF-κB p65 activity induced by high glucose in human lens epithelial cells.

AIM: To explore the effects of IκBα SUMOylation and NF-κB p65 deacetylation on NF-κB p65 activity induced by high glucose in cultured human lens epithelial cells (HLECs). METHODS: HLECs (SRA01/04) were cultured with 5.5, 25, and 50 mmol/L glucose media for 24h, and with 50 mmol/L glucose media for 0, 12, and 24h respectively. SUMO1 and SIRT1 expressions were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot (WB). IκBα and NF-κB p65 expressions were detected by WB. With NAC, DTT, MG132 or Resveratrol (RSV) treatment, SUMO1 and SIRT1 expressions were detected by WB. Protein expression localizations were examined by immunofluorescence and co-immunofluorescence. The effects of SUMO1 or SIRT1 overexpression, as well as MG132 and RSV, on the nuclear expression and activity of IκBα and NF-κB p65 were analyzed by immunoblot and dual luciferase reporter gene assay. RESULTS: SUMO1 and SIRT1 expressions were influenced by high glucose in mRNA and protein levels, which could be blocked by NAC or DTT. SUMO1 was down-regulated by using MG132, and SIRT1 was up-regulated under RSV treatment. IκBα nuclear expression was attenuated and NF-κB p65 was opposite under high glucose, while IκBα and NF-κB p65 location was transferred to the nucleus. SUMO1 or SIRT1 overexpression and MG132 or RSV treatment affected the nuclear expression and activity of IκBα and NF-κB p65 under high glucose condition. CONCLUSION: IκBα SUMOylation and NF-κB p65 deacetylation affect NF-κB p65 activity in cultured HLECs under high glucose, and presumably play a significant role in controlling diabetic cataract.

Multiscale Micro-Nano Nested Structures: Engineered Surface Morphology for Efficient Light Escaping in Organic Light-Emitting Diodes.

Various micro-to-nanometer scale structures are extremely attractive for light escaping in organic light-emitting diodes. To develop and optimize such structures, an innovative approach was demonstrated for the first time to fabricate multiscale micro-nano nested structures by photolithography with a well-designed mask pattern followed by a controllable thermal reflow process. The experimental and theoretical characterizations verify that these unique nested structures hold the capability of light concentration, noticeable low haze, and efficient antireflection. As a proof-of-concept, the incorporation of this pattern onto the glass substrate efficiently facilitates light escaping from the device, resulting in current efficiency 1.60 times and external quantum efficiency 1.63 times that of a control flat device, respectively. Moreover, compared to a hexagonally arranged microlens array and quasi-random biomimetic moth eye nanostructures, the nested structures proposed here can magically tune the spatial emission profile to comply with the Lambertian radiation pattern. Hence, this novel structure is expected to be of great potential in related ubiquitous optoelectronic applications and provide scientific inspiration to other novel multiscale micro-nanostructure research.

HIF-1α SUMOylation affects the stability and transcriptional activity of HIF-1α in human lens epithelial cells.

PURPOSE: High blood glucose can induce oxidative damage and result in diabetic cataract. Oxidative stress induces various signal pathways including HIF-1α transcriptional signal to attenuate the damage of lenses. Whether HIF-1α SUMOylation can increase the activation of HIF-1α or if high glucose can affect the SUMOylation of HIF-1α in cultured human lens epithelial cells (HLECs) is still unknown, as well as the function of HIF-1α SUMOylation in oxidative damage induced by high glucose in HLECs. In the present study, we examined SUMO and SUMO E3 (Cbx4 and PIASy) expression induced by high glucose, and investigated SUMO or SUMO E3 overexpression that enhanced HIF-1α SUMOylation in HLECs. METHODS: SRA01/04 cells, one kind of human lens epithelial cell line, were addressed in media with 5.5 mmol/l (normal control group), 25 mmol/l (high glucose1 group) and 50 mmol/l (high glucose2 group) final glucose respectively. Expression of SUMO1 ~ 4, Cbx4, PIASy, HIF-1α, GLUT1, and VEGFA were detected in the mRNA and protein levels by RT-PCR and Western blot analysis. Protein expression localization and co-localization were examined by immunofluorescence and co-immunofluorescence. The effects of SUMO overexpression, SUMO E3 overexpression, and Proteasome inhibitor MG132 respectively on the stability and transcriptional activity of HIF-1α were analyzed by immunoblot. RESULTS: High glucose treatment induced SUMO1-4 expression and enhanced the expression of Cbx4 and PIASy. It also increased the expression of HIF-1α, GLUT1, and VEGFA. The co-localization of HIF-1α and SUMO was mainly in the nucleus induced by high glucose. Further studies showed that SUMO overexpression or SUMO E3 overexpression could enhance HIF-1α stability and transcriptional activity in HLECs. Proteasome inhibitor MG132 protected the stability and transcriptional activity of HIF-1α in the SRA01/04 cells. CONCLUSIONS: HIF-1α SUMOylation affected the stability and transcriptional activity of HIF-1α in cultured human lens epithelial cells; SUMO overexpression or SUMO E3 overexpression enhanced the expression of HIF-1α, which is involved in inhibiting cell apoptosis and protecting lens opacification, and presumably plays a key role in protecting lenses from diabetic cataract.

Handling of Diskus dry powder inhaler in Chinese chronic obstructive pulmonary disease patients.

BACKGROUND: The incorrect handling of Diskus inhalers in Chinese patients with chronic obstructive pulmonary disease (COPD) is not well documented. OBJECTIVE: The present study was conducted to evaluate in detail the handling errors related to the Diskus device, and to elucidate the importance of educating COPD patients on the proper use of the device. METHODS: A total of 384 COPD patients from a pulmonary clinic in China over a period of 5 years were included in the study. The compliance of COPD patients to the 13 discrete steps of Diskus usage were scored and analyzed by three measures: (1) On day 0, patients were given only a package insert on Diskus, and the handling error rate was assessed. Then the patients were given instruction on the 13-step Diskus procedure until they could demonstrate the proper technique. (2) On days 1, 2, and 3, the observation group was continuously educated on a 13-step procedure, and the percentage of patients who scored 100% for each step was recorded. The control group had no such training. (3) On days 10, 20, and 30, the percentage of all subjects correctly performing the Diskus 13-step inhalation procedure was assessed. RESULTS: Incorrect handling techniques on Diskus usage were widely distributed among Chinese COPD patients. Step 8 ("Inhale forcefully from the beginning, slowly, deeply, and uniformly during the inspiratory phase until the lungs are full") was the most commonly mishandled step (93.8%). The total score and individual step scores of the patients from the observation group were significantly improved during 3-day continuous education. There was also a significantly higher percentage of correctly performed steps in the observation group than in the control group upon assessment on day 10 (96.24% vs. 85.63%, respectively; p<0.01), day 20 (97.31% vs. 86.09%, respectively; p<0.01), and day 30 (98.19% vs. 87.39%, respectively; p<0.01). CONCLUSION: Handling errors of the Diskus 13-step inhalation procedure were commonly observed in Chinese COPD patients. Continuous educational interventions and regular supervision by health-care providers are therefore crucial for the optimum use of the Diskus inhaler.

Lactulose biosynthesis by ß-galactosidase from a newly isolated Arthrobacter sp.

Lactulose, a ketose disaccharide, is used in both pharmaceutical and food industries. This study was undertaken to screen and isolate potent ß-galactosidase-producing bacteria and to evaluate their enzymatic production of lactulose. Soil samples from fruit gardens were collected. One isolate designated LAS was identified whose cell extract could convert lactose and fructose into lactulose. The 16S rDNA gene analysis of LAS revealed its phylogenetic relatedness to Arthrobacter sp. The ß-galactosidase produced by LAS was purified 15.7-fold by ammonium sulfate precipitation and subsequent Phenyl-Sepharose hydrophobic chromatography. The optimum pH and temperature for lactulose synthesis by this ß-galactosidase were 6.0 and 20 °C, respectively. The low optimum temperature of this enzyme compared to the currently used ones for lactulose production has the advantage of reducing the nonenzymatic browning in biotransformations. The results indicated that Arthrobacter could be used as a novel bacterial ß-galactosidase source for lactulose production.