Choline dehydrogenase interacts with SQSTM1 to activate mitophagy and promote coelomocyte survival in Apostichopus japonicus following Vibrio splendidus infection.

Previous studies have shown that Vibrio splendidus infection causes mitochondrial damage in Apostichopus japonicus coelomocytes, leading to the production of excessive reactive oxygen species (ROS) and irreversible apoptotic cell death. Emerging evidence suggests that mitochondrial autophagy (mitophagy) is the most effective method for eliminating damaged mitochondria and ROS, with choline dehydrogenase (CHDH) identified as a novel mitophagy receptor that can recognize non-ubiquitin damage signals and microtubule-associated protein 1 light chain 3 (LC3) in vertebrates. However, the functional role of CHDH in invertebrates is largely unknown. In this study, we observed a significant increase in the mRNA and protein expression levels of A. japonicus CHDH (AjCHDH) in response to V. splendidus infection and lipopolysaccharide (LPS) challenge, consistent with changes in mitophagy under the same conditions. Notably, AjCHDH was localized to the mitochondria rather than the cytosol following V. splendidus infection. Moreover, AjCHDH knockdown using siRNA transfection significantly reduced mitophagy levels, as observed through transmission electron microscopy and confocal microscopy. Further investigation into the molecular mechanisms underlying CHDH-regulated mitophagy showed that AjCHDH lacked an LC3-interacting region (LIR) for direct binding to LC3 but possessed a FB1 structural domain that binds to SQSTM1. The interaction between AjCHDH and SQSTM1 was further confirmed by immunoprecipitation analysis. Furthermore, laser confocal microscopy indicated that SQSTM1 and LC3 were recruited by AjCHDH in coelomocytes and HEK293T cells. In contrast, AjCHDH interference hindered SQSTM1 and LC3 recruitment to the mitochondria, a critical step in damaged mitochondrial degradation. Thus, AjCHDH interference led to a significant increase in both mitochondrial and intracellular ROS, followed by increased apoptosis and decreased coelomocyte survival. Collectively, these findings indicate that AjCHDH-mediated mitophagy plays a crucial role in coelomocyte survival in A. japonicus following V. splendidus infection.

ROS-mediated BNIP3-dependent mitophagy promotes coelomocyte survival in Apostichopus japonicus in response to Vibrio splendidus infection.

Organisms produce high levels of reactive oxygen species (ROS) to kill pathogens or act as signaling molecules to induce immune responses; however, excessive ROS can result in cell death. To maintain ROS balance and cell survival, mitophagy selectively eliminates damaged mitochondria via mitophagy receptors in vertebrates. In marine invertebrates, however, mitophagy and its functions remain largely unknown. In the current study, Vibrio splendidus infection damaged mitochondrial morphology in coelomocytes and reduced mitochondrial membrane potential (ΔΨm) and mitophagosome formation. The colocalization of mitochondria and lysosomes further confirmed that lipopolysaccharide (LPS) treatment increased mitophagy flux. To explore the regulatory mechanism of mitophagy, we cloned Bcl2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), a common mitophagy receptor, from sea cucumber Apostichopus japonicus (AjBNIP3) and confirmed that AjBNIP3 was significantly induced and accumulated in mitochondria after V. splendidus infection and LPS exposure. At the mitochondrial membrane, AjBNIP3 interacts with microtubule-associated protein 1 light chain 3 (LC3) on phagophore membranes to mediate mitophagy. After AjBNIP3 interference, mitophagy flux decreased significantly. Furthermore, AjBNIP3-mediated mitophagy was activated by ROS following the addition of exogenous hydrogen peroxide (H2O2), ROS scavengers, and ROS inhibitors. Finally, inhibition of BNIP3-mediated mitophagy by AjBNIP3 small interfering RNA (siRNA) or high concentrations of lactate increased apoptosis and decreased coelomocyte survival. These findings highlight the essential role of AjBNIP3 in damaged mitochondrial degradation during mitophagy. This mitophagy activity is required for coelomocyte survival in A. japonicus against V. splendidus infection.

An FMRFamide Neuropeptide in Cuttlefish Sepia pharaonis: Identification, Characterization, and Potential Function.

Neuropeptides are released by neurons that are involved in a wide range of brain functions, such as food intake, metabolism, reproduction, and learning and memory. A full-length cDNA sequence of an FMRFamide gene isolated from the cuttlefish Sepia pharaonis (designated as SpFMRFamide) was cloned. The predicted precursor protein contains one putative signal peptide and four FMRFamide-related peptides. Multiple amino acid and nucleotide sequence alignments showed that it shares 97% similarity with the precursor FMRFamides of Sepiella japonica and Sepia officinalis and shares 93% and 92% similarity with the SpFMRFamide gene of the two cuttlefish species, respectively. Moreover, the phylogenetic analysis also suggested that SpFMRFamide and FMRFamides from S. japonica and S. officinalis belong to the same sub-branch. Tissue expression analysis confirmed that SpFMRFamide was widely distributed among tissues and predominantly expressed in the brain at the three development stages. The combined effects of SpFMRFamide+SpGnRH and SpFLRFamide+SpGnRH showed a marked decrease in the level of the total proteins released in the CHO-K1 cells. This is the first report of SpFMRFamide in S. pharaonis and the results may contribute to future studies of neuropeptide evolution or may prove useful for the development of aquaculture methods for this cuttlefish species.

Hydride Transfer Initiated Redox-Neutral Cascade Cyclizations of Aurones: Facile Access to [6,5] Spirocycles.

Reported herein is the hydride transfer initiated redox-neutral cascade cyclizations of aurones, providing a variety of [6,5] spiro-heterocycles in satisfactory yields and good diastereoselectivities.

Blood urea nitrogen in the prediction of in-hospital mortality of patients with acute aortic dissection.

BACKGROUND: Blood urea nitrogen (BUN) has been shown to be associated with adverse cardiovascular disease outcomes. The aim of the present study was to evaluate the prognostic role of BUN in patients with acute aortic dissection (AAD). HYPOTHESIS: BUN has correlation with in-hospital mortality of patients with AAD. METHODS: Patients admitted to the emergency room within the first 24 h of onset of AAD were included in the study. BUN levels were measured on admission and the endpoints were mortality during hospi-talization after receiving surgical or endovascular repair. RESULTS: A total of 192 patients with AAD were enrolled. During hospitalization, 19 patients died and 173 patients survived. Increased levels of BUN (8.9 [7.0-9.7] vs. 6.0 [5.1-7.2] mmol/L, p < 0.001) were found in non-survivors compared with those survived. Using multivariable logistic analysis, BUN was an independent predictor of in-hospital mortality in patients with AAD (OR 1.415, 95% CI 1.016-1.971, p = 0.040). Furthermore, using receiver operating characteristic analysis, the optimal cutoff value for BUN was 6.95 mmol/L. Under this value, the area under the curve was 0.785 (95% CI 0.662-0.909, p < 0.001) and the sensitivity and specificity to predict in-hospital mortality was 78.9%, and 72.2%, respectively. CONCLUSIONS: Admission BUN levels were an independent predictor for in hospital mortality in pa-tients with AAD.

The relationship between fibrinogen and in-hospital mortality in patients with type A acute aortic dissection.

BACKGROUND AND PURPOSE: Fibrinogen plays an important role in hemostasis and thrombosis and is proven to have prognostic significance in patients with cardiovascular disease. We examined the utility of fibrinogen as a prognostic indicator for patients with type A acute aortic dissection (AAD). METHODS: This study was performed in consecutive patients with type A AAD admitted to our hospital within 24 hours after onset of symptoms. Fibrinogen levels were measured on admission. Baseline clinical characteristics and laboratory test results were collected. The endpoint was in-hospital mortality. RESULTS: A total of 143 patients with type A AAD were enrolled. Compared with the survivors, the nonsurvivors had significant lower fibrinogen levels (1.95(1.37, 2.38) vs. 2.37(1.85, 3.15) g/L, p=0.001). The cutoff level of fibrinogen determined by ROC curve analysis was 2.17 g/L, with a sensitivity, specificity of 71.9%, 60.4% respectively, and the area under the ROC curve was 0.686 (95% CI, 0.585-0.768; p=0.001). After controlling for potentially relevant confounding variables, we found an admission fibrinogen level less than 2.17g/L was associated with an increased risk of in-hospital mortality (odds ratio, 5.527; 95% CI, 1.660-18.401; p=0.005) compared with those with fibrinogen greater than 2.17g/L. CONCLUSION: Low fibrinogen level on admission is an independent predictor of in-hospital mortality in patients with type A AAD.

Molecular cloning, expression analysis and cellular localization of an LFRFamide gene in the cuttlefish Sepiella japonica.

Neuropeptides are important regulators of physiological processes in metazoans, such as feeding, reproduction, and heart activities. In this study, an LFRFamide gene was identified from the cuttlefish Sepiella japonica (designated as SjLFRFamide). The full-length sequence of SjLFRFamide cDNA has 841bp, and the open reading frame contains 567bp encoding 188 amino acids, which shared high similarity with precursor SOFaRP2 from Sepia officinalis. The deduced SjLFRFamdie precursor protein contains a signal peptide and four different FLPs (FMRFamide-like peptides): one pentapeptide (TIFRFamide), two hexapeptides (NSLFRFamide and GNLFRFamide) and one heptapeptide (PHTPFRFamide). Multiple sequence alignment showed that SjLFRFamide contains rather conserved mature peptides, which all ended in FRF. The phylogenetic analysis suggests that SjLFRFamide belongs to the LFRFamide subfamily. The tissue distribution analysis through quantitative real-time PCR method showed that SjLFRFamide mRNA is significantly expressed in the brain, and slight trace are detected in female nidamental gland and accessory nidamental gland. In situ hybridization assay of the brain indicated that SjLFRFamide is transcribed in several different functional lobes, suggesting SjLFRFamide might associate with multiple physiological regulations, such as feeding, chromatophore regulation and reproduction. This is the first study describing LFRFamide in S. japonica, which might have great importance for cuttlefish artificial breeding.