Genome-wide analysis of ATP-binding cassette (ABC) transporter in Penaeus vannamei and identification of two ABC genes involved in immune defense against Vibrio parahaemolyticus by affecting NF-κB signaling pathway.

The ATP-binding cassette (ABC) transporters have crucial roles in various biological processes such as growth, development and immune defense in eukaryotes. However, the roles of ABC transporters in the immune system of crustaceans remain elusive. In this study, 38 ABC genes were systematically identified and characterized in Penaeus vannamei. Bioinformation analysis revealed that PvABC genes were categorized into ABC A-H eight subfamilies with 17 full-transporters, 11 half transporters and 10 soluble proteins, and multiple immunity-related cis-elements were found in gene promoter regions. Expression analysis showed that most PvABC genes were widely and highly expressed in immune-related tissues and responded to the stimulation of Vibrio parahaemolyticus. To investigate whether PvABC genes mediated innate immunity, PvABCC5, PvABCF1 and PvABCB4 were selected for dsRNA interference experiment. Knockdown of PvABCF1 and PvABCC5 not PvABCB4 increased the cumulative mortality of P. vannamei and bacterial loads in hepatopancreas after infection with V. parahaemolyticus. Further analysis showed that the PvABCF1 and PvABCC5 knockdown decreased expression levels of NF-κB pathway genes and antimicrobial peptides (AMPs). Collectively, these findings indicated that PvABCF1 and PvABCC5 might restrict V. parahaemolyticus challenge by positively regulating NF-κB pathway and then promoting the expression of AMPs, which would contribute to overall understand the function of ABC genes in innate immunity of invertebrates.

A smartphone-based fluorescent sensor for rapid detection of multiple pathogenic bacteria.

In this study, we developed a fluorescent sensor for the sensitive detection of multiple pathogenic bacteria based on magnetic separation, fluorescent probes, and smartphone image processing. A microchannel device was assembled using high-transparency resin and 3D printing technology. This device was combined with a smartphone and an external lens to develop a fluorescent sensor for autonomous detection of multiple pathogenic bacteria. Three fluorescence probes with different fluorescence were synthesized from highly specific aptamers and tetraphenylethylene derivatives. These fluorescent probes can make Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa emit different colors of fluorescence. Using the enrichment performance of molecularly imprinted materials, separation and detection of bacteria can be achieved simultaneously. Finally, with the Red-Green-Blue (RGB) analysis functionality of a smartphone, real-time field detection was realized with a sensitivity of 102 CFU/mL and a detection time of 40 min. This work provides a simple, inexpensive, and real-time sensor for the detection of multiple pathogens in medical diagnostics, food testing, and environmental analyses.

Transcriptomic analysis of gills in nitrite-tolerant and -sensitive families of Litopenaeus vannamei.

Nitrite stress is a major environmental factor that limits aquatic animal growth, reproduction and survival. Even so, some shrimps still can withstand somewhat high concentrations of nitrite environment. However, few studies have been conducted about the tolerance molecular mechanism of Litopenaeus vannamei in the high concentration nitrite. To identify the genes and pathways involved in the regulation of nitrite tolerance, we performed comparative transcriptomic analysis in the L. vannamei nitrite-tolerant (NT) and nitrite-sensitive (NS) families, and untreated shrimps were used as the control group. After 24 h of nitrite exposure (NaNO2, 112.5 mg/L), a total of 1521 and 868 differentially expressed genes (DEGs) were obtained from NT compared with NS and control group, respectively. Functional enrichment analysis revealed that most of these DEGs were involved in immune defense, energy metabolism processes and endoplasmic reticulum (ER) stress. During nitrite stress, energy metabolism in NT was significantly enhanced by activating the related genes expression of oxidative phosphorylation (OXPHOS) pathway and tricarboxylic acid (TCA) cycle. Meanwhile, some DEGs involved in innate immunity- related genes and pathways, and ER stress responses also were highly expressed in NT. Therefore, we speculate that accelerated energy metabolism, higher expression of immunity and ER related genes might be the important adaptive strategies for NT in relative to NS under nitrite stress. These results will provide new insights on the potential tolerant molecular mechanisms and the breeding of new varieties of nitrite tolerant L. vannamei.

Pheochromocytoma with abdominal aortic aneurysm presenting as recurrent dyspnea, hemoptysis, and hypotension: A case report.

BACKGROUND: Pheochromocytomas are rare endocrine tumors with various clinical manifestations, and few of them might present with profound, life-threatening conditions. CASE SUMMARY: We report the case of a 65-year-old man who complained of sudden dyspnea and hemoptysis for half a day. There was no obvious cause for the patient to have dyspnea, coughing, or coughing up to approximately 100 mL of fresh blood. Finally, he was diagnosed with pheochromocytoma crisis (PCC), coexisting with an abdominal aortic aneurysm (AAA). CONCLUSION: We report a case of pheochromocytoma presenting with recurrent hemoptysis, dyspnea and hypotension coexisting with an AAA. It not only proved the uncommon manifestations of pheochromocytoma but also directed clinicians to consider PCC among the possible diagnoses when meeting similar cases. Moreover, surgical excision is the most beneficial method for the treatment of pheochromocytoma coexisting with AAA when the situation is stable.

Integrated analysis of physiological, transcriptomic and metabolomic responses and tolerance mechanism of nitrite exposure in Litopenaeus vannamei.

Nitrite accumulation in aquatic environments is a potential risk factor that disrupts multiple physiological functions in aquatic animals. In this study, the physiology, transcriptome and metabolome of the control group (LV-C), nitrite-tolerance group (LV-NT) and nitrite-sensitive group (LV-NS) were investigated to identify the stress responses and mechanisms underlying the nitrite tolerance of Litopenaeus vannamei. After LV-NT and LV-NS were subjected to nitrite stress, the hemocyanin contents were significantly decreased, and hepatopancreas showed severe histological damage compared with LV-C. Likewise, the antioxidant enzymes were also significantly changed after nitrite exposure. The transcriptome data revealed differentially expressed genes associated with immune system, cytoskeleton remodeling and apoptosis in LV-NT and LV-NS. The combination of transcriptomic and metabolomic analysis revealed nitrite exposure disturbed metabolism processes in L. vannamei, including amino acid metabolism, nucleotide metabolism and lipid metabolism. The multiple comparative analysis implicated that higher nitrite tolerance of LV-NT than LV-NS may be attributed to enhanced hypoxia inducible factor-1α expression to regulate energy supply and gaseous exchange. Moreover, LV-NT showed higher antioxidative ability, detoxification gene expression and enhanced fatty acids contents after nitrite exposure in relative to LV-NS. Collectively, all these results will greatly provide new insights into the molecular mechanisms underlying the stress responses and tolerance of nitrite exposure in L. vannamei.

Stress response and tolerance mechanisms of ammonia exposure based on transcriptomics and metabolomics in Litopenaeus vannamei.

Ammonia, one of the major limiting environment factors in aquaculture, may pose a threat to the shrimp growth, reproduction and survival. In this study, to understand molecular differences of transcriptomic and metabolomic responses and investigate the tolerance mechanisms underlying ammonia stress in Litopenaeus vannamei, ammonia-tolerant family (LV-AT) and ammonia-sensitive family (LV-AS) of these two extreme families were exposed to high-concentration (NH4Cl, 46 mg/L) ammonia for 24 h. The comparative transcriptome analysis between ammonia-treated and control (LV-C) groups revealed involvement of immune defense, cytoskeleton remodeling, antioxidative system and metabolic pathway in ammonia-stress response of L. vannamei. Likewise, metabolomics analysis showed that ammonia exposure could disturb amino acid metabolism, nucleotide metabolism and lipid metabolism, with metabolism related-genes changed according to RNA-seq analysis. The comparison of metabolite and transcript profiles between LV-AT and LV-AS indicated that LV-AT used the enhanced glycolysis and tricarboxylic acid (TCA) cycle strategies for energy supply and ammonia excretion to adapt high-concentration ammonia. Furthermore, some of genes involved in the detoxification and ammonia excretion were highly expressed in LV-AT. We speculate that the higher ability of ammonia excretion and detoxification and the accelerated energy metabolism for energy supplies might be the adaptive strategies for LV-AT relative to LV-AS after ammonia stress. Collectively, the combination of transcriptomics and metabolomics results will greatly contribute to incrementally understand the stress responses on ammonia exposure to L. vannamei and supply molecular level support for evaluating the environmental effects of ammonia on aquatic organisms. The results further constitute new sights on the potential molecular mechanisms of ammonia adaptive strategies in shrimps at the transcriptomics and metabolomics levels.

Effects of Mild Hypothermia on Cardiac and Neurological Function in Piglets Under Pathological and Physiological Stress Conditions.

To investigate the different effects of mild hypothermia on pathological and physiological stress conditions in piglets, 30 pigs were randomized into four groups: cardiac arrest and mild hypothermia (CA-MH group), cardiac arrest and normothermia (CA-NH group), non-CA-MH (NCA-MH group), and a sham operation. The same hypothermia intervention was implemented in both CA-MH and NCA-MH groups. The CA-NH group did not undergo therapeutic hypothermia after resuscitation. The hemodynamic parameters were recorded. Cerebral metabolism variables and neurotransmitters in the extracellular fluid were collected through microdialysis tubes. The serum of venous blood was used to detect levels of inflammatory factors. The cerebral function was evaluated. At 24 and 72 hours after resuscitation, the cerebral performance category and neurological deficit score in the CA-NH group had higher values. Heart rate and cardiac output (CO) in the CA-MH group during cooling were lower than that of the CA-NH group, but CO was higher after rewarming. Glucose was higher during cooling, and extracellular lactate and lactate/pyruvate ratio in the CA-MH group were lower than that of the CA-NH group. Noradrenaline and 5-hydroxytryptamine in the CA-MH and NCA-MH groups were lower than that of the CA-NH group and sham group during cooling, respectively. Inflammatory factor levels, including interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, and tumor necrosis factor-α, in the CA-MH group were lower than that of the CA-NH group at cooling for 12 hours. These values in the NCA-MH group were higher than that of the sham group. Under a light and an electron microscope, the worse pathological results of heart and brain were observed in the two cardiac arrest groups. Mild hypothermia can provide limited organ protection in the specific pathological condition caused by ischemia-reperfusion, but it may produce a negative effect in a normal physiological state.

Proteomic Responses Under Cold Stress Reveal Unique Cold Tolerance Mechanisms in the Pacific White Shrimp (Litopenaeus vannamei).

The Pacific white shrimp (Litopenaeus vannamei), one of the most widely cultured shrimp species in the world, often suffers from cold stress. To understand the molecular mechanism of cold tolerance in Pacific white shrimp, we conducted a proteomic analysis on two contrasting shrimp cultivars, namely, cold-tolerant Guihai2 (GH2) and cold-sensitive Guihai1 (GH1), under normal temperature (28°C), under cold stress (16°C), and during recovery to 28°C. In total, 3,349 proteins were identified, among which 2,736 proteins were quantified. Based on gene ontology annotations, differentially expressed proteins largely belonged to biological processes, cellular components, and molecular functions. KEGG pathway annotations indicated that the main changes were observed in the lysosome, ribosomes, and oxidative phosphorylation. Subcellular localization analysis showed a significant increase in proteins present in cytosol, extracellular regions, and mitochondria. Combining enrichment-based clustering analysis and qRT-PCR analysis, we found that glutathione S-transferase, zinc proteinase, m7GpppX diphosphatase, AP2 transcription complex, and zinc-finger transcription factors played a major role in the cold stress response in Pacific white shrimp. Moreover, structure proteins, including different types of lectin and DAPPUDRAFT, were indispensable for cold stress tolerance of the Pacific white shrimp. Results indicate the molecular mechanisms of the Pacific white shrimp in response to cold stress and provide new insight into breeding new cultivars with increased cold tolerance.

Effect of Low Temperature on Globin Expression, Respiratory Metabolic Enzyme Activities, and Gill Structure of Litopenaeus vannamei.

Low temperature frequently influences growth, development, and even survival of aquatic animals. In the present study, physiological and molecular responses to low temperature in Litopenaeus vannamei were investigated. The cDNA sequences of two oxygen-carrying proteins, cytoglobin (Cygb) and neuroglobin (Ngb), were isolated. Protein structure analysis revealed that both proteins share a globin superfamily domain. Real-time PCR analysis indicated that Cygb and Ngb mRNA levels gradually increased during decrease in temperatures from 25 to 15°C and then decreased at 10°C in muscle, brain, stomach, and heart, except for a continuing increase in gills, whereas they showed a different expression trend in the hepatopancreas. Hemocyanin concentration gradually reduced as the temperature decreased. Moreover, the activities of respiratory metabolic enzymes including lactate dehydrogenase (LDH) and succinate dehydrogenase (SDH) were measured, and it was found that LDH activity gradually increased while SDH activity decreased after low-temperature treatment. Finally, damage to gill structure at low temperature was also observed, and this intensified with further decrease in temperature. Taken together, these results show that low temperature has an adverse influence in L. vannamei, which contributes to systematic understanding of the adaptation mechanisms of shrimp at low temperature.

Effect of mild hypothermia on renal ischemia/reperfusion injury after cardiopulmonary resuscitation in a swine model.

PURPOSE:: To investigate the effect of intravascular cooling on renal function after resuscitation. METHODS:: Twenty four pigs were randomized into three groups (n=8 in each group): therapeutic hypothermia group (TH group), normothermia group (NH group) and sham operation group (SHAM group). After 6 minutes of untreated VF, CPR was performed. Upon ROSC, the TH group received the intravascular cooling. The NH and SHAM group did not undergo therapeutic hypothermia. Haemodynamic parameters were recorded. The bloods were analyzed for serum creatinine (sCr), CysC and NGAL. The kidney was surgically removed observe pathologic changes under a light microscope. RESULTS:: The sCr increased in both TH and NH groups after ROSC, compared to baseline. Between two groups, the sCr and creatinine clearance (Cc) showed lower level in the TH group. The urine volume per hour in the TH group were higher during cooling. After resuscitation, NGAL and CysC in the NH group were higher than in the TH group. Under the light microscope, compared with the TH group, the renal injury was prominent in the NH group. CONCLUSION:: Mild hypothermia had a protection to renal ischemia reperfusion injury after resuscitation.

Effect of mild hypothermia on renal ischemia/reperfusion injury after cardiopulmonary resuscitation in a swine model

Abstract Purpose: To investigate the effect of intravascular cooling on renal function after resuscitation. Methods: Twenty four pigs were randomized into three groups (n=8 in each group): therapeutic hypothermia group (TH group), normothermia group (NH group) and sham operation group (SHAM group). After 6 minutes of untreated VF, CPR was performed. Upon ROSC, the TH group received the intravascular cooling. The NH and SHAM group did not undergo therapeutic hypothermia. Haemodynamic parameters were recorded. The bloods were analyzed for serum creatinine (sCr), CysC and NGAL. The kidney was surgically removed observe pathologic changes under a light microscope. Results: The sCr increased in both TH and NH groups after ROSC, compared to baseline. Between two groups, the sCr and creatinine clearance (Cc) showed lower level in the TH group. The urine volume per hour in the TH group were higher during cooling. After resuscitation, NGAL and CysC in the NH group were higher than in the TH group. Under the light microscope, compared with the TH group, the renal injury was prominent in the NH group. Conclusion: Mild hypothermia had a protection to renal ischemia reperfusion injury after resuscitation.

Comparison of early sequential hypothermia and delayed hypothermia on neurological function after resuscitation in a swine model.

BACKGROUND: We utilized a porcine cardiac arrest model to compare early sequential hypothermia (ESH) with delayed hypothermia (DH) and no hypothermia (NH) to investigate the different effects on cerebral function after resuscitation. METHODS: After return of spontaneous circulation (ROSC), resuscitated 24 pigs divided into three groups. The ESH group implemented early sequential hypothermia immediately, and the DH group implemented delayed hypothermia at 1 h after ROSC. The core temperature, hemodynamic parameters and oxygen metabolism were recorded. Cerebral metabolism variables and neurotransmitter in the extracellular fluid were collected through the microdialysis tubes. The bloods were analyzed for venous jugular bulb oxygen saturation, lactate and neuron specific nolase. The cerebral function was evaluated using the cerebral performance category and neurologic deficit score at 72h after ROSC and cerebral histology in the right posterior frontal lobe were collected. RESULTS: ESH reached the target temperature earlier and showed more favorable outcomes of neurological function than DH. Specifically, early sequential hypothermia reduced cerebral oxygen and energy consumption and decreased extracellular accumulation of neurotransmitters after resuscitation and protected the integrity of the BBB during reperfusion. CONCLUSIONS: Early sequential hypothermia could increase the protection of neurological function after resuscitation and produce better neurological outcomes. The institutional protocol number: 2010-D-013.

[Establishment of a new method for early embryo culture in vitro].

A new method, nest culture method, was developed in this study. The culture effects of different methods for embryo culture in vitro were compared. The results showed that whether the suspension in phi35mm dish in nest culture method was covered with mineral oil or not, the developmental rates of embryos had no significant difference. Compared to the nest culture method, the developmental rates of embryos in Brinster's method were significant lower. However, all embryos cultured in the single dish in which the suspensions were not covered with mineral oil were blocked at 2-cell stage. The nest culture method is an effective method for early embryo culture in vitro.