The Transpeptidase Sortase A Binds Nucleic Acids and Mediates Mammalian Cell Labeling.

Wild-type sortase A is an important virulence factor displaying a diverse array of proteins on the surface of bacteria. This protein display relies on the transpeptidase activity of sortase A, which is widely engineered to allow protein ligation and protein engineering based on the interaction between sortase A and peptides. Here an unknown interaction is found between sortase A from Staphylococcus aureus and nucleic acids, in which exogenously expressed engineered sortase A binds oligonucleotides in vitro and is independent of its canonical transpeptidase activity. When incubated with mammalian cells, engineered sortase A further mediates oligonucleotide labeling to the cell surface, where sortase A attaches itself and is part of the labeled moiety. The labeling reaction can also be mediated by many classes of wild-type sortases as well. Cell surface GAG appears involved in sortase-mediated oligonucleotide cell labeling, as demonstrated by CRISPR screening. This interaction property is utilized to develop a technique called CellID to facilitate sample multiplexing for scRNA-seq and shows the potential of using sortases to label cells with diverse oligonucleotides. Together, the binding between sortase A and nucleic acids opens a new avenue to understanding the virulence of wild-type sortases and exploring the application of sortases in biotechnology.

Sternal wound infections following internal mammary artery grafts for a coronary bypass: A meta-analysis.

The meta-analysis aims to evaluate and compare the sternal wound infections following internal mammary artery grafts for a coronary bypass. Examinations comparing bilateral internal mammary artery to single internal mammary artery for coronary artery bypass grafting were among the meta-analyses from various languages that met the inclusion criteria. Using dichotomous random- or fixed-effect models, the results of these investigations were examined, and the Odd Ratio (OR) with 95% confidence intervals (CIs) was computed. A total of 31 examinations from 2001 to 2023 were recruited for the current analysis including 181 503 personals with coronary artery bypass grafting. Bilateral internal mammary artery had significantly higher sternal wound infection (OR, 1.51; 95% CI, 1.37-1.68, p < 0.001), superficial sternal wound infection (OR, 1.72; 95% CI, 1.16-2.56, p = 0.007), deep sternal wound infection (OR, 1.62; 95% CI, 1.41-1.86, p < 0.001), sternal wound infection in diabetics (OR, 1.48; 95% CI, 1.16-1.90, p = 0.002), sternal wound infection in elderly (OR, 1.38; 95% CI, 1.22-1.57, p < 0.001), sternal wound infection in pedicled preparation (OR, 1.70; 95% CI, 1.30-2.23, p < 0.001) and sternal wound infection in skeletonized preparation (OR, 1.40; 95% CI, 1.09-1.81, p = 0.009) compared to single internal mammary artery in personals with coronary artery bypass grafting. Bilateral internal mammary artery grafting is linked to a higher risk of impaired wound healing, particularly in diabetic individuals, elderly, pedicled preparation, and skeletonized preparation. Nevertheless, caution should be exercised while interacting with its values since examinations were performed by different surgeons with different skills on different types of personals.

Regulation Mechanism of Dopamine Receptor 1 in Low Temperature Response of Marsupenaeus japonicus.

Dopamine receptors (DARs) are important transmembrane receptors responsible for receiving extracellular signals in the DAR-mediated signaling pathway, and are involved in a variety of physiological functions. Herein, the D1 DAR gene from Marsupenaeus japonicus (MjDAD1) was identified and characterized. The protein encoded by MjDAD1 has the typical structure and functional domains of the G-protein coupled receptor family. MjDAD1 expression was significantly upregulated in the gills and hepatopancreas after low temperature stress. Moreover, double-stranded RNA-mediated silencing of MjDAD1 significantly changed the levels of protein kinases (PKA and PKC), second messengers (cyclic AMP (cAMP), cyclic cGMP, calmodulin, and diacyl glycerol), and G-protein effectors (adenylate cyclase and phospholipase C). Furthermore, MjDAD1 silencing increased the apoptosis rate of gill and hepatopancreas cells. Thus, following binding to their specific receptors, G-protein effectors are activated by MjDAD1, leading to DAD1-cAMP/PKA pathway-mediated regulation of caspase-dependent mitochondrial apoptosis. We suggest that MjDAD1 is indispensable for the environmental adaptation of M. japonicus.

Optimized minimal genome-wide human sgRNA library.

Genome-wide clustered regularly interspaced short palindromic repeats (CRISPR)-based knockout screening is revolting the genetic analysis of a cellular or molecular phenotype in question but is challenged by the large size of single-guide RNA (sgRNA) library. Here we designed a minimal genome-wide human sgRNA library, H-mLib, which is composed of 21,159 sgRNA pairs assembled based on a dedicated selection strategy from all potential SpCas9/sgRNAs in the human genome. These sgRNA pairs were cloned into a dual-gRNA vector each targeting one gene, resulting in a compact library size nearly identical to the number of human protein-coding genes. The performance of the H-mLib was benchmarked to other CRISPR libraries in a proliferation screening conducted in K562 cells. We also identified groups of core essential genes and cell-type specific essential genes by comparing the screening results from the K562 and Jurkat cells. Together, the H-mLib exemplified high specificity and sensitivity in identifying essential genes while containing minimal library complexity, emphasizing its advantages and applications in CRISPR screening with limited cell numbers.

Effect of hematoma volume on the 30-day mortality rate of patients with primary hypertensive brainstem hemorrhage: a retrospective cohort study.

Objective: The purpose of this study is to investigate the effect of hematoma volume on the 30-Day Mortality Rate of patients with Primary Hypertensive Brainstem Hemorrhage (PHBH). Methods: Retrospective analysis was done on the clinical information of 74 patients who underwent treatment for primary hypertensive brainstem hemorrhage at the Department of Neurosurgery of the 908th Hospital of the Joint Logistic Support Force of the Chinese People's Liberation Army between January 2018 and December 2021. Both univariate and multivariate logistic regression were used to assess clinical signs and risk factors that affect 30-day mortality. Results: In the 74 patients with primary hypertensive brainstem hemorrhage included in this investigation, 46 patients died and 28 patients survived. The mortality rate at 30 days was 62.16%. A statistically significant difference was seen (P < 0.001) in the results of the univariate analysis, which suggested that hematoma volume may be a factor affecting the prognosis of patients with hypertensive brainstem hemorrhage. Hematoma volume was further demonstrated to be a risk factor and an independent factor impacting death in patients with brainstem hemorrhage (P < 0.001) by multivariate logistic regression analysis (OR: 2.6, 95% CI: 1.7-3.9, P < 0.001 Crude Model, OR: 3.6, 95% CI: 1.7-7.7, P < 0.001 Multivariate-Adjusted Model). After adjusting for confounding variables such as age, body mass index, sex, history of diabetes mellitus, history of hypertension, admission GCS score, stereotactic aspiration, combined hydrocephalus, admission systolic and diastolic blood pressure, the hematoma volume was revealed to be an independent predictor of 30-day death in patients with brainstem hemorrhage. We discovered by smooth curve fitting that hematoma volume increased in a non-linear manner with 30-day mortality. The 30-day mortality rate did not alter significantly when the hematoma volume was less than 4 ml. When the hematoma volume was greater than 4 ml, the 30-day mortality rate increased rapidly, and when the hematoma volume was 10 ml, the 30-day mortality rate reached the maximum. Conclusions: Hematoma volume is an independent factor affecting 30-day mortality in patients with primary hypertensive brainstem hemorrhage. The severe and extensive neurological damage caused by primary hypertensive brainstem hemorrhage is highly unlikely to be fundamentally altered by a single protocol, and new avenues need to be explored scientifically and continuously.

The efficacy of low-frequency ultrasound as an added treatment for chronic wounds: A meta-analysis.

We performed a meta-analysis to evaluate the effect of low-frequency ultrasound as an added treatment for chronic wounds. A systematic literature search up to May 2022 was performed and 838 subjects with chronic wounds at the baseline of the studies; 412 of them were using the low-frequency ultrasound (225 low-frequency high-intensity contact ultrasound for diabetic foot wound ulcers, and 187 low-frequency low-intensity non-contact ultrasound for a venous leg wound ulcers), and 426 were using standard care (233 sharp debridements for diabetic foot wound ulcers and 193 sham treatments for venous leg wound ulcers). Odds ratio (OR), and mean difference (MD) with 95% confidence intervals (CIs) were calculated to assess the effect of low-frequency ultrasound as an added treatment for chronic wounds using the dichotomous, and contentious methods with a random or fixed-effect model. The low-frequency high-intensity contact ultrasound for diabetic foot wound ulcers had significantly lower non-healed diabetic foot wound ulcers at ≥3 months (OR, 0.37; 95% CI, 0.24-0.56, P < .001), a higher percentage of diabetic foot wound ulcers area reduction (MD, 17.18; 95% CI, 6.62-27.85, P = .002) compared with sharp debridement for diabetic foot wound ulcers. The low-frequency low-intensity non-contact ultrasound for a venous leg wound ulcers had a significantly lower non-healed venous leg wound ulcers at ≥3 months (OR, 0.31; 95% CI, 0.15-0.62, P = .001), and higher percentage venous leg wound ulcers area reduction (MD, 18.96; 95% CI, 2.36-35.57, P = .03) compared with sham treatments for a venous leg wound ulcers. The low-frequency ultrasound as an added treatment for diabetic foot wound ulcers and venous leg wound ulcers had significantly lower non-healed chronic wound ulcers at ≥3 months, a higher percentage of chronic wound ulcers area reduction compared with standard care. The analysis of outcomes should be with caution because of the low sample size of all the 17 studies in the meta-analysis and a low number of studies in certain comparisons.

The coumarin-pyrazole dye for detection of hydrogen sulfide in cells.

Fluorescent probes for H2S are often interfered by other thiols. In this work, a coumarin-pyrazole dye with 2,4-dinitrosulfonyl group was designed for the detection of H2S. The probe exhibits weak fluorescence in water due to the photo induced electron transfer (PET) by 2,4-dinitrosulfonyl. After the sulfonyl group is cleaved off by H2S, strong fluorescence appears. The probe can specifically detect H2S without being interfered by other biological thiols, and shows a wide applicable pH range, low detection and wide detection range. The excellent detection properties of the probe can also be used to detect endogenous and exogenous H2S in cells. In addition, the probes can be made into portable test paper for the detection of H2S in solutions and can detect H2S in different water samples.

PEAC-seq adopts Prime Editor to detect CRISPR off-target and DNA translocation.

CRISPR technology holds significant promise for biological studies and gene therapies because of its high flexibility and efficiency when applied in mammalian cells. But endonuclease (e.g., Cas9) potentially generates undesired edits; thus, there is an urgent need to comprehensively identify off-target sites so that the genotoxicities can be accurately assessed. To date, it is still challenging to streamline the entire process to specifically label and efficiently enrich the cleavage sites from unknown genomic locations. Here we develop PEAC-seq, in which we adopt the Prime Editor to insert a sequence-optimized tag to the editing sites and enrich the tagged regions with site-specific primers for high throughput sequencing. Moreover, we demonstrate that PEAC-seq could identify DNA translocations, which are more genotoxic but usually overlooked by other off-target detection methods. As PEAC-seq does not rely on exogenous oligodeoxynucleotides to label the editing site, we also conduct in vivo off-target identification as proof of concept. In summary, PEAC-seq provides a comprehensive and streamlined strategy to identify CRISPR off-targeting sites in vitro and in vivo, as well as DNA translocation events. This technique further diversified the toolkit to evaluate the genotoxicity of CRISPR applications in research and clinics.

Lung-function trajectories in COVID-19 survivors after discharge: A two-year longitudinal cohort study.

Background: Data on the long-term trajectories of lung function are scarce in COVID-19 survivors. Methods: We re-analyzed the data from a prospective longitudinal cohort follow-up study of COVID-19 survivors over 2 years after infection. All participants were divided into scale 3, scale 4 and scale 5-6 groups according to seven-category ordinal scale. The changes of pulmonary function tests (PFTs), the Modified Medical Research Council (mMRC) Dyspnea Scale, 6-min walking test health-related quality of life (HRQoL) across the three serial follow-up visits were evaluated, and compared among three groups. We performed liner regression to determine potential factors that were associated with changes of PFTs and distance walked in 6 minutes (6MWD). Findings: In this study, 288 participants generally presented an improvement of PFTs parameters from 6 months to 1 year after infection. The scale 5-6 group displayed a significantly higher increase of PFTs compared with scale 3 and scale 4 groups (all p<0.0167), and corticosteroids therapy was identified as a protective factor for the PFTs improvement with a correlation coefficient of 2.730 (0.215-5.246) for forced vital capacity (FVC), 2.909 (0.383-5.436) for total lung capacity (TLC), and 3.299 (0.211-6.387) for diffusion capacity for carbon monoxide (DLco), respectively. From 1-year to 2-year follow-up, the PFTs parameters generally decreased, which was not observed to be associated with changes of 6MWD and HRQoL. Dyspnea (mMRC≥1) generally decreased over time (23.3% [61/262] for 6-month, 27.9% [67/240] for 1-year, 13.4% [35/261] for 2-year), and 6MWD increased continuously (500.0 m vs 505.0 m vs 525.0 m). Interpretation: Corticosteroids therapy during hospitalization was a protective factor for PFTs improvement from 6 months to 1 year. The relatively fast decline trend of PFTs from 1 year to 2 years needs to be paid attention and further validated in the future follow-up study. Fundings: This work was supported by Chinese Academy of Medical Sciences Innovation Fund for Medical Sciences (CIFMS 2021-I2M-1-048) and the National Key Research and Development Program of China (2021YFC0864700).

Effects of general anesthesia and epidural anesthesia on deep vein thrombosis and perioperative cognitive function of patients undergoing total knee arthroplasty.

OBJECTIVE: To explore the effects of general anesthesia and epidural anesthesia on deep vein thrombosis (DVT) and perioperative cognitive function of patients undergoing total knee arthroplasty. METHODS: A retrospective analysis was conducted on 110 orthopedic patients who underwent total knee arthroplasty at Zhangzhou Affiliated Hospital of Fujian Medical University from March 2018 to March 2021. According to different anesthesia schemes, 56 cases with epidural anesthesia were included in the observation group and 54 patients with general anesthesia were assigned to the control group. The following items were recorded and compared between the two groups: postoperative coagulation indicators; operation duration, total dosage of anesthetics; postoperative recovery time; heart rate (HR) and mean arterial pressure (MAP) before anesthesia induction (T0), intubation (T1), and completion of the operation (T2); cognitive function before surgery and 1 d and 3 d after surgery; postoperative incidence of DVT; pain at postoperative 24 h; stress-related factors before and 2 h after surgery, and incidence of adverse reactions during hospitalization. RESULTS: Compared to the control group, the levels of fibrinogen (Fbg) and platelets (PLC) in the observation group 24 hours after surgery were significantly lower, with longer thrombin time (TT) and prothrombin time (PT) (all P<0.05); no significant difference was found in operation duration between the two groups (P>0.05). Total dosage of anesthetics and postoperative recovery time were less in the observation group (P<0.05); the HR and MAP fluctuations were lower in the observation group (P<0.05). The postoperative cognitive function score of patients in the observation group was significantly higher (P<0.05), with a lower incidence of postoperative DVT (P<0.05) and better relief of pain (P<0.05). The expression levels of stress-related factors 2 h after surgery and the incidence of adverse reactions were lower in the observation group compared to the control group (P<0.05). CONCLUSION: In total knee arthroplasty, epidural anesthesia, compared with general anesthesia, can reduce the incidence of DVT in patients and has less impact on patients' cognitive function and stress state with a higher safety profile.

The clinical significance of thyroid hormone-responsive in thyroid carcinoma and its potential regulatory pathway.

The study aimed to evaluate the clinical significance of thyroid hormone-responsive (THRSP) and explore its relevant pathways in thyroid carcinoma (THCA). The gene expression data of THRSP were obtained and the prognostic significance of THRSP in THCA was analyzed through various bioinformatics databases. Then, the factors influencing THRSP mRNA expression were explored, and the function of THRSP in predicting the lymph node metastasis (LNM) stage was determined. We further performed the enrichment analysis and constructed a protein-protein interaction (PPI) network to examine potential regulatory pathways associated with THRSP. THRSP gene expression was significantly increased in THCA compared with the normal tissues. High THRSP mRNA expression had a favorable overall survival (OS) in THCA patients (P < .05). Additionally, the mRNA expression of THRSP was related to stage, histological subtype, and methylation among THCA patients (all P < .05). Besides, THRSP served as a potent predictor in discriminating the LNM stage of thyroid cancer patients. According to Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene set enrichment analysis (GSEA) on THRSP-associated genes, THRSP was positively related to metabolic pathways. The upregulation of THRSP predicted a good OS in THCA patients. Furthermore, THRSP might inhibit THCA progression through positive regulation of metabolism-associated pathways.

An in-library ligation strategy and its application in CRISPR/Cas9 screening of high-order gRNA combinations.

Simultaneous targeting multiple genes is a big advantage of CRISPR (clustered regularly interspaced short palindromic repeats) genome editing but challenging to achieve in CRISPR screening. The crosstalk among genes or gene products is a common and fundamental mechanism to ensure cellular stability and functional diversity. However, the screening approach to map high-order gene combinations to the interesting phenotype is still lacking. Here, we developed a universal in-library ligation strategy and applied it to generate multiplexed CRISPR library, which could perturb four pre-designed targets in a cell. We conducted in vivo CRISPR screening for potential guide RNA (gRNA) combinations inducing anti-tumor immune responses. Simultaneously disturbing a combination of three checkpoints in CD8+ T cells was demonstrated to be more effective than disturbing Pdcd1 only for T cell activation in the tumor environment. This study developed a novel in-library ligation strategy to facilitate the multiplexed CRISPR screening, which could extend our ability to explore the combinatorial outcomes from coordinated gene behaviors.

Artificial Intelligence Algorithm-Based CTA Imaging for Diagnosing Ischemic Type Biliary Lesions after Orthotopic Liver Transplantation.

The study focused on the clinical application value of artificial intelligence-based computed tomography angiography (CTA) in the diagnosis of orthotopic liver transplantation (OLT) after ischemic type biliary lesions (ITBL). A total of 66 patients receiving OLT in hospital were selected. Convolutional neural network (CNN) algorithm was used to denoise and detect the edges of CTA images of patients. At the same time, the quality of the processed image was subjectively evaluated and quantified by Hmax, Ur, Cr, and other indicators. Then, the digital subtraction angiography (DSA) diagnosis and CTA diagnosis based on CNN were compared for the sensitivity, specificity, positive predictive value, negative predictive value, and patient classification results. It was found that CTA can clearly reflect the information of hepatic aorta lesions and thrombosis in patients with ischemic single-duct injury after liver transplantation. After neural network algorithm processing, the image quality is obviously improved, the lesions are more prominent, and the details of lesion parts are also well displayed. ITBL occurred in 40 (71%) of 56 patients with abnormal CTA at early stage. ITBL occurred in only 8 (12.3%) of 65 patients with normal CTA at early stage. Early CTA manifestations had high sensitivity (72.22%), specificity (87.44%), positive predictive value (60.94%), and negative predictive value (92.06%) for the diagnosis of ITBL. It was concluded that artificial intelligence-based CTA had high clinical application value in the diagnosis of ITBL after OLT.

Nicorandil Inhibits Osteoclast Formation Base on NF-κB and p-38 MAPK Signaling Pathways and Relieves Ovariectomy-Induced Bone Loss.

Osteolytic bone disorders are characterized by an overall reduction in bone mineral density which enhances bone ductility and vulnerability to fractures. This disorder is primarily associated with superabundant osteoclast formation and bone resorption activity. Nicorandil (NIC) is a vasodilatory anti-anginal drug with ATP-dependent potassium (KATP) channel openings. However, NIC is adopted to manage adverse cardiovascular and coronary events. Recent research has demonstrated that NIC also possesses anti-inflammatory peculiarity through the regulation of p38 MAPK and NF-κB signaling pathways. Both MAPK and NF-κB signaling pathways play pivotal roles in RANKL-induced osteoclast formation and bone resorption function. Herein, we hypothesized that NIC may exert potential biological effects against osteoclasts, and revealed that NIC dose-dependently suppressed bone marrow macrophage (BMM) precursors to differentiate into TRAP + multinucleated osteoclasts in vitro. Furthermore, osteoclast resorption assays demonstrated anti-resorptive effects exhibited by NIC. NIC had no impact on osteoblast differentiation or mineralization function. Based on Biochemical analyses, NIC relieved RANKL-induced ERK, NF-κB and p38 MAPK signaling without noticeable effects on JNK MAPK activation. However, the attenuation of NF-κB and p38 MAPK activation was sufficient to hamper the downstream induction of c-Fos and NFATc1 expression. Meanwhile, NIC administration markedly protected mice from ovariectomy (OVX)-induced bone loss through in vivo inhibition of osteoclast formation and bone resorption activity. Collectively, this work demonstrated the potential of NIC in the management of osteolytic bone disorders mediated by osteoclasts.

The role of Nrf2 in mitigating cadmium-induced oxidative stress of Marsupenaeus japonicus.

Nuclear factor-erythroid 2-related factor-2 (Nrf2) is an important modulator of cellular responses against Cd in mammalian cells. However, whether such modulation is conserved in Marsupenaeus japonicas remains unknown.In our study, the shrimps were injected with dsRNA targeting Nrf2 at 4 µg g-1 body weight (b.w.) or sulforaphane (SFN) at 5 µg g-1 b.w., and then were exposed to 40 mg L-1 CdCl2 for 48 h. After Nrf2 knockdown, the Cd content increased, but decreased in the SFN group. This suggested that Nrf2 could promote Cd excretion. A terminal deoxynulceotidyl transferase nick-end-labeling (TUNEL) assay revealed that the Nrf2 knockdown increased the number of apoptotic cells in M. japonicas, while SFN decreased the number of apoptotic cells. After Nrf2 knockdown, the total antioxidant capacity (T-AOC), superoxide dismutase (Sod) activity, and related gene expression decreased significantly, while the malondialdehyde (MDA) content increased remarkably. By contrast, SFN injection alleviated the oxidative stress, as evidenced by increased T-AOC, Sod activity, sod mRNA expression and a reduced MDA content. Similarly, detoxification related enzyme activities (ethoxyresorufin O-deethylase and glutathione-S-transferase (GST)) and their corresponding gene expressions (cyp3a (cytochrome P450 family 3 subfamily A) and gst) were suppressed in the ds-Nrf2 injection group, while they were elevated in the SFN group. In addition, ds-Nrf2 activated mitochondrial apoptotic pathway, as evidenced the mRNA and protein levels of caspase-3, Bcl2 associated X protein (Bax), and p53, while SFN treatment suppressed them. These results displayed that in M. japonicus Cd-induced cellular oxidative damage probably acts via the Nrf2 pathway.

Integrated transcriptomic and metabolomic responses in the hepatopancreas of kuruma shrimp (Marsupenaeus japonicus) under cold stress.

In aquatic ecosystems, the temperature of the water is an important ecological factor that modulates aquatic organisms' metabolism, growth, development, and reproduction. In this study, the morphological, transcriptomic, and metabolomic analyses of response of Marsupeneus japonicus to acute cold stress was investigated. The results revealed that low temperature caused profound morphological damage to the hepatopancreas. Transcriptomic responses suggested that energy and primary metabolism, cytoskeleton structure, and apoptosis signaling were altered. The metabolic responses to cold stress included changes of multiple amino acids and unsaturated fatty acids. Combined transcriptomic and metabolomic data indicated that energy metabolism pathways were downregulated in the hepatopancreas under cold stress. However, M. japonicus increased ATP and unsaturated fatty acids production to ameliorate. Moreover, cold stress caused significant attenuation of macrophage apoptosis. This study provides key information to increase our understanding of low-temperature tolerance in shrimp.

Direct-seq: programmed gRNA scaffold for streamlined scRNA-seq in CRISPR screen.

CRISPR-based genome perturbation provides a new avenue to conveniently change DNA sequences, transcription, and epigenetic modifications in genetic screens. However, it remains challenging to assay the complex molecular readouts after perturbation at high resolution and at scale. By introducing an A/G mixed capture sequence into the gRNA scaffold, we demonstrate that gRNA transcripts could be directly reverse transcribed by poly (dT) primer together with the endogenous mRNA, followed by high-content molecular phenotyping in scRNA-seq (Direct-seq). With this method, the CRISPR perturbation and its transcriptional readouts can be profiled together in a streamlined workflow.

Molecular Cloning and Bioinformatics Analysis of DQA Gene from Mink (Neovison vison).

In the present study, we cloned, sequenced, and explored the structural and functional characteristics of the major histocompatibility complex (MHC)-DQA gene from mink (Neovison vison) for the first time. The full-length sequence of DQA gene was 1147-bp-long, contained a coding region of 768-bp, which was predicted to encoding 255 amino acid residues. The comparison between DQA from mink (Neovison vison) and other MHC-DQA molecules from different animal species showed that nucleotide and encoded amino acid sequences of the mink DQA gene exhibited high similarity with the ferret (Mustela pulourius furo). Phylogenetic analysis revealed that mink (Neovison vison) DQA is grouped with that of ferret (Mustela pulourius furo). The cloned sequence contained a 23-amino acid NH2-terminal signal sequence with the signal peptide cutting site located in amino acids 23⁻24, and had three Asn-Xaa-Ser/Thr sequons. Three cysteine residues were also identified (Cys-85, Cys-121, and Cys-138). The 218 to 240 amino acids were predicted to be the transmembrane domains. The prediction of the secondary structure revealed three α-helixes and fourteen ß-sheets in Neovison vison DQA protein, while random coil was a major pattern. In this study, the whole CDS sequence of Neovison vison DQA gene was successfully cloned, which was valuable for exploring the function and antiviral molecular mechanisms underlying the molecule. The findings of the present study have laid the foundation for the disease resistance and breeding of mink.

The impact of acute thermal stress on green mussel Perna viridis: Oxidative damage and responses.

Examining the physiological responses of mussels to thermal stress is crucial to evaluate their biogeographic distribution and ability to adapt to a changing climate. In the present study, we investigated the effects of acute cold (8 °C and 15 °C) and heat (35 °C and 42 °C) stress on the mortality rate, reactive oxygen species (ROS) production, malondialdehyde (MDA) content, mitochondrial membrane potential (MMP) and antioxdative responses in the gill tissue of the green mussel species Perna viridis. Our results showed that cold and heat stress induced a temperature-dependent increase in mortality rate. ROS production increased significantly (p < 0.01) after both cold and heat stress. However, the activities of antioxidant enzymes, including SOD, CAT and GSH-Px, were greatly enhanced only after heat stress. In addition, MDA content and MMP increased significantly under both cold and heat stress. The up-regulation of Hsp70 transcripts was only detected after acute stress at 35 °C. However, p38-MAPK phosphorylation levels increased after both cold and heat stress. In addition, a moderate activation of caspase-3 was found after mussels were exposed to 8 °C and 42 °C stress. Our results suggest that both extreme cold and heat stress could induce ROS production in the gill tissue of P. viridis, which might result in lipid peroxidation and mitochondria dysfunction. Antioxidative enzymes and Hsp70 might be important in the heat stress response of animals, whereas p38-MAPK might be crucial in the acute response to both cold and heat stress. However, caspase-3 activation might be very weak under both cold and heat stress.