Cadmium(Cd)is a common environmental heavy metal pollutant known for its toxic effects on various plants and animals.Cd contamination also poses a threat to Pardosa pseudoannulata,a predator in rice paddies.Results in...Cadmium(Cd)is a common environmental heavy metal pollutant known for its toxic effects on various plants and animals.Cd contamination also poses a threat to Pardosa pseudoannulata,a predator in rice paddies.Results indicate that Cd pollution significantly reduced the levels of Metallothionein,phenoloxidase,and Ca^(2+).Furthermore,through metabolomics and 16S rDNA sequencing,alterations in the gut microbiota and immune-related metabolites of P.pseudoannulata were identified.The abundance of harmful bacteria such as Staphylococcus increased significantly,while levels of l-Tyrosine,glutathione disulfide,l-Dopa,among others,decreasedmarkedly.Pearson correlation analysis revealed significant associations between various gut microbes and immune-related metabolites,suggesting that Cd may alter the immune system of P.pseudoannulata by modulating its gut microbiota.展开更多
Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based pu...Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based purification(ClinProt system) and matrix-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) to characterize shrimp hemolymph peptides.Shrimp serum and plasma were used as the source of samples for comparative analysis,and it was found that serum was more suitable for shrimp hemolymph peptidomic analysis.To screen potential specific biomarkers in serum of immune-challenged shrimps,we applied magnetic bead-based MALDI-TOF MS to serum samples from 10 immune-challenged and 10 healthy shrimps.The spectra were analyzed using FlexAnalysis 3.0 and ClinProTools 2.1 software.Thirteen peptide peaks significantly different between the two groups were selected as candidate biomarkers of lipopolysaccharide(LPS)-infection.The diagnostic model established by genetic algorithm using five of these peaks was able to discriminate LPS-challenged shrimps from healthy control shrimps with a sensitivity of 90% and a specificity of 100%.Our approach in MALDITOF MS-based peptidomics is a powerful tool for screening bioactive peptides or biomarkers derived from hemolymph,and will help to enable a better understanding of the innate immune response of shrimps.展开更多
Experiments showed the agglutinin activity in the hemolymph of both spring wild parents and autumncultured individuals of the shrimp Penaeus chinensis Osbeck with the body length of 12 - 16 cm as all 32. Thehemolymph ...Experiments showed the agglutinin activity in the hemolymph of both spring wild parents and autumncultured individuals of the shrimp Penaeus chinensis Osbeck with the body length of 12 - 16 cm as all 32. Thehemolymph of the above shrimps showed agglutinant reaction to all A-, B- and O-type human erythrocytes, with thereaction titer equal to that of rabbit blood cells. However, the agglutinin activity level was enhanced after being in-duced by Vibro, which was 128 after 24 h of induction and reached the peak value 256 after 48 h of induction, thendecreased gradually.展开更多
Knowledge of dietary compositions is essential to the recovery of the dwindling populations of horseshoe crabs in China.The feeding habits of Tachypleus tridentatus and Carcinoscorpis rotundicauda of Pearl Bay in sout...Knowledge of dietary compositions is essential to the recovery of the dwindling populations of horseshoe crabs in China.The feeding habits of Tachypleus tridentatus and Carcinoscorpis rotundicauda of Pearl Bay in southwestern China were studied by stable isotope analysis.Hemolymph samples of the two species were obtained from two age groups living in different habitats(inner,central,and outer Bay).In addition,their potential food source samples were collected in May 2019.Results showed that the mean values ofδ^(13)C andδ^(15)N in C.rotundicauda individuals ranged from−19.01‰to−16.47‰and from 10.49‰to 13.5‰,respectively,and those of T.tridentatus ranged from−19.12‰to−14.96‰and from 8.78‰to 13.48‰,respectively.These values indicated that the horseshoe crabs have a wide variety of food sources and therefore are highly omnivorous.No remarkably correlations were found betweenδ^(13)C andδ^(15)N values and individual widths in the two species.Thirteen potential food sources were selected,in which Sipunculus nudus,Batillaria zonalisi,and Ulva lactuca contributed largely to the dietary compositions of the two species of horseshoe crabs.No significant correlation was found between the feeding habits and habitat geochemical characteristics of the horseshoe crabs.All these results possibly indicate the high diversity of their food sources and the indiscriminate dietary compositions of the two Asian horseshoe crabs.展开更多
Mixtures of hemolymph from Chlamys farreri with three different anticoagulant solutions were incu-bated for an hour in vitro, then the ultrastructural alterations of hemocytes were observed, and the aggre-gation rate ...Mixtures of hemolymph from Chlamys farreri with three different anticoagulant solutions were incu-bated for an hour in vitro, then the ultrastructural alterations of hemocytes were observed, and the aggre-gation rate was analyzed by using transmission electron microscropy and flow cytometry respectively. The results showed that Formula 3 （glucose 20.8 g L^-1; EDTA 20mM; sodium chloride 20 g L^-1; Tris-HCI 0.05M;pH 7.4）was the desirable anticoagulant solution for C. farreri hemocytes. Further phagocytosis assay showed that no obvious negative effect was given to the hemocyte phagocytic activity when using Formula 3 as the anticoagulant solution.展开更多
Insects are well recognized as a source of potentially useful compounds for modern medicine. Development of anticancer drugs from natural resources has been performed throughout the world. In the present study, antica...Insects are well recognized as a source of potentially useful compounds for modern medicine. Development of anticancer drugs from natural resources has been performed throughout the world. In the present study, anticancer activity of the hemolymph and fat body of Sarcophaga argyrostoma third larval instars is assayed against human breast cancer cell line (MDA-MB-231 cells). The cytotoxicity of the hemolymph and fat body samples were determined. The results showed that growth of MDA-MB-231 cells was inhibited at different concentrations upon 24 h of exposure. There is no inhibitory activity against Vero cells under these experimental conditions. Protein profile of the hemolymph and fat body were extracted and separated using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Protein analysis showed differences in number of electrophoretic protein bands with different molecular weights between treated and control larvae. The present work assumes that hemolymph and fat body tissue have cytotoxicity against MDA-MB-231 cells and these results exhibit that larvae from S. argyrostoma could be used as a good source for developing anti-cancer agents and knowledge of these anti-cancer compounds will lead to better control of human disease.展开更多
This study investigated the effects of silkworm hemolymph-derived storage protein 2 (SP2) on the whitening process in mouse B16F1 melanoma cells. After the cells were treated with various concentrations of SP2 (0.1 - ...This study investigated the effects of silkworm hemolymph-derived storage protein 2 (SP2) on the whitening process in mouse B16F1 melanoma cells. After the cells were treated with various concentrations of SP2 (0.1 - 1.0 mg/mL), cytotoxicity, melanin contents, and differences in mRNA and protein expression associated with melanogenesis were observed. No cytotoxicity was observed when cells were treated with SP2, even with increased SP2 concentrations of up to 2.0 mg/mL. When treated with various SP2 concentrations in the cells, the protein and mRNA expression of tyrosinase were dose-dependently decreased, respectively, and inhibition of tyrosinase was further increased by 50.0% with increasing SP2 concentration of 1.0 mg/mL. Expression mRNAs coding tyrosinase related protein-1 and protein-2 (TRP-1 and TRP-2) was also significantly decreased in a dose dependent manner. When measuring the melanin content in melanoma cells, SP2 at 1 mg/mL inhibited melanin synthesis by 73.5% compared with non-treated cells. The inhibitory effect was 2.8-fold higher than that obtained using arbutin as a positive control. This study demonstrates that SP2, as a whitening material, is capable of suppressing melanin synthesis through the downregulation of proteins and genes in the melanin biosynthesis pathway.展开更多
[Objective] The research aimed to study the activity and dynamic changes of phenoloxidase in Locusta migratoria manilensis（Meyen） .[Method] Two enzyme solutions of insect body and hemolymph of Locusta migratoria man...[Objective] The research aimed to study the activity and dynamic changes of phenoloxidase in Locusta migratoria manilensis（Meyen） .[Method] Two enzyme solutions of insect body and hemolymph of Locusta migratoria manilensis（Meyen） were prepared by different extract methods,the activity and dynamic changes of phenoloxidase in different sites were measured.[Result] The phenoloxidase activity in hemolymph was significantly higher than that in body fluid,and the activity in female individuals was higher than that in male individuals.The phenoloxidase activity in body fluid was gradually enhanced with the prolongation of laying time in air.The phenoloxidase activity in hemolymph of adult was also gradually enhanced with the prolongation of time.[Conclusion] Part of phenoloxidase was existed in tissues and cells of Locusta migratoria manilensis（Meyen） participating in insect development and construction,and the other part was existed in hemolymph playing an immunological and defensive role.The stability of phenoloxidase was very strong,which was the effective guarantee for wide adaptation ability and strong immunity of insect.展开更多
The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of suc...The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of successful cross breeding in silkworm. Protein samples from silk gland, hemolymph, and midgut were separated by 2-dimensional gel electrophoresis （2-DE）. In the three tissues the matched protein spots between Jinqiu and its cross parents were approximately 70% with approximately 30% specific protein spots. In the matched protein spots, 9-24% was differentially expressed representing up- and down-regulated expression. These specific protein spots might be either the newly appeared, which were produced from the genic interaction of cross parents＇ genes in cross breeding, or posttranscriptionally modified, which were produced from the different modifications on the same original proteins. These results indicate that it is important for a new successful breed, by cross breeding, relying on the actions of some newly produced functional proteins from genic interaction, in addition to marshaling excellent genes of cross parents.展开更多
Crassostrea nippona is a valuable species for aquaculture with considerable potential for commercial oyster farming.However,it is vulnerable to changes in salinity levels in coastal environments.In this study,we inves...Crassostrea nippona is a valuable species for aquaculture with considerable potential for commercial oyster farming.However,it is vulnerable to changes in salinity levels in coastal environments.In this study,we investigated the impacts of low salinity stress on the physiological responses of C.nippona.The hemolymph osmolality could not reach equilibrium with the surrounding environmental osmolality that was below salinity 15 within 1 week.Cell expansion,cellular valuocation,decrease of gill cilia,increased apoptotic cells under salinity 10 were observed through microscopic techniques.The activities of immunity-related enzymes,including alkaline phosphatase(AKP),acid phosphatase(ACP),superoxide dismutase(SOD),and catalase(CAT),were significantly increased at salinity 10 compared with the control group.These findings highlight the vulnerability of C.nippona to low salinity stress and provide insights into the physiological changes in response to fluctuating salinity levels.Understanding these physiological responses is crucial for effective aquaculture management and developing strategies to mitigate the negative impacts of low salinity stress on C.nippona populations in coastal areas.展开更多
The effect of acute E. coli challenge on the anti-disease activity of scallops Argopecten irra-dians is examined. The treatments of scallop from which hemolymph samples were taken for study included (1) control scallo...The effect of acute E. coli challenge on the anti-disease activity of scallops Argopecten irra-dians is examined. The treatments of scallop from which hemolymph samples were taken for study included (1) control scallops, (2) sham-injected scallops, (3) PSW-injected scallops and (4) E. coli-injected scallops. From the beginning, the anti-disease activities of scallops are deter -mined at 12 hr and 24 hr.The concentrations of circulating hemocytes, the total serum protein concentrations and the activities of alkaline phosphatase, acid phosphatase and superoxide dismutase in the scallops Argopecten irradians are determined.Injection with E. coli results in a significant elevation in the concentration of circulating hemocytes and in the alkaline phosphatase activity and a significant decline in the total serum protein concentration and in the superoxide dismutase activity at 24 hr postchallenge. It shows that metabolism of bay scallop is expedited to adopt the challenge.展开更多
Insects in the Order Odonata are highly subject to infection by gregarine parasites. However, despite the important ecological roles that insects play in every ecosystem in which they exist, little research has been d...Insects in the Order Odonata are highly subject to infection by gregarine parasites. However, despite the important ecological roles that insects play in every ecosystem in which they exist, little research has been devoted to the description of insect immunity. Insects rely heavily on the rapid actions of innate immune mechanisms to prevent infection. We characterized the melanization response in the hemolymph of green darner dragonfly (Anax junius) nymphs. Incubation of chymotrypsin-activated hemolymph with L-DOPA resulted in volume- and time-dependent production of dopaquinone via the phenoloxidase (PO) enzyme, with biphasic accumulation of product. The PO activity was temperature-dependent, with a stepwise increase from 20℃ - 35℃ and maximum activity measured at 35℃ - 40℃. The formation of product was also inhibited in a concentration-dependent manner by diethylcarbonate, a specific inhibitor of PO activity, which indicated that the observed activity was due to the presence of PO enzyme. The rate of formation and quantity of melanin was dependent on exposure to different titers of bacteria. This is the first characterization of both PO activity and melanization response in green darner dragonflies.展开更多
There is an increasing interest in discovering new antibacterial agents derived from nature to enhance the treatment of various bacterial infections.Defensins and their derived peptide fragments exhibit significant an...There is an increasing interest in discovering new antibacterial agents derived from nature to enhance the treatment of various bacterial infections.Defensins and their derived peptide fragments exhibit significant antibacterial activity without any cytotoxic effects,making them attractive features for potential novel antibacterial therapeutics.Crassostrea gigas,a traditional seafood that has been used worldwide for centuries,has its shells applied in Chinese medicine as Ostreae concha.In this study,bioinformatics analysis was used to obtain a novel antibacterial peptide,CGD-1,derived from marine Chinese medicine Ostreae concha.The physicochemical characterization and circular dichroism analysis results demonstrated that CGD-1 assembled into anα-helical structure in a simulated membrane environment,and it displayed antibacterial action against Gram-negative bacteria.The minimal inhibitory concentrations against both Pseudomonas aeruginosa ATCC27853 and Escherichia coli ATCC25922 were 25μM.CGD-1 was able to efficiently permeate the cell membrane.Changes in bacterial cell morphology were evaluated using a field emission scanning electron microscope.The results suggested that CGD-1 exerted its antibacterial activity through permeabilizing and disrupting the bacterial cell membrane.Therefore,CGD-1 may have potential applications in fighting against pathogenic bacteria such as P.aeruginosa and E.coli.展开更多
Two-dimensional gel electrophoresis （2-DE） followed by matrix-assisted laser desorption ionization- time-of-flight/time-of-flight mass spectrometry （MS） analysis were used to charaterize the hemolymph proteomic pr...Two-dimensional gel electrophoresis （2-DE） followed by matrix-assisted laser desorption ionization- time-of-flight/time-of-flight mass spectrometry （MS） analysis were used to charaterize the hemolymph proteomic profiles of the silkworm, Bombyx mori. At days 4 （V4） and 5 （VS） of the fifth （final） instar, when the larvae were at the fast-growing stage, we found dramatic changes in spots representing proteins having an approximate molecular weight （MW） of 30 kDa. Of these spots, four 30K proteins were highly up- regulated, implying a close association with the growth and development ofB. mori larvae. To understand the molecular basis and underlying mechanisms involved in development and metamorphosis, the proteome of whole hemolymph at V5 was analyzed using shotgun liquid chromatography tandem mass spectrometry with an LTQ-Orbitrap. A total of 108 proteins were identified without any false discovery hits. These proteins were involved in a variety of cellular functions, including metabolism, development, nutrient transport and reserve, and defense response. Gene ontology analysis showed that 3.4% of these proteins had nutrient reservoir activities and 5.7% were involved in the response to stimulus. Pathway analysis revealed that 22 proteins with common targets were involved in various cellular processes such as immunity, differentiation, proliferation and metamorphosis. These results suggested that some key factors such as the 30K proteins in hemolymph play important roles in B. mori growth and development. Moreover, the multiple functions of hemolymph may be operated by a complex biological network.展开更多
During insect larval-pupal metamorphosis,proteins in the hemolymph are absorbed by the fat body for the maintenance of intracellular homeostasis;however,the type of proteins and how these proteins are internalized int...During insect larval-pupal metamorphosis,proteins in the hemolymph are absorbed by the fat body for the maintenance of intracellular homeostasis;however,the type of proteins and how these proteins are internalized into the fat body are unclear.In Bombyx mori,the developmental profiles o f total proteins in the hemolymph and fat body showed that hemolymph-decreased protein bands(55-100 kDa)were in accordance with those protein bands that increased in the fat body.Inhibition o f clathrin-dependent endocytosis predominantly blocked the transportation o f 55-100 kDa proteins from the hemolymph into the fat body,which was further verified by RNA interference treatment o f Bmclathrin.Six hexamerins were shown to comprise〜90%of the total identified proteins in both the hemolymph and fat body by mass spectrum(MS)analysis.In addition,hemolymphspecific proteins were mainly involved in material transportation,while fat body-specific proteins particularly participated in metabolism.In this paper,four hexamerins were found for the first time,and potential proteins absorbed by the fat body from the hemolymph through clathrin-dependent endocytosis were identified.This study sheds light on the protein absorption mechanism during insect metamorphosis.展开更多
Acute-phase response is documented to be a significant mechanism of innate immunity in vertebrates and invertebrates.In this study,proteomic methodologies were applied for different protein expressions in hemolymph of...Acute-phase response is documented to be a significant mechanism of innate immunity in vertebrates and invertebrates.In this study,proteomic methodologies were applied for different protein expressions in hemolymph of Scylla serrata challenged by Vibrio parahaemolyticus after immunization,and in muscles of the crabs separately chal-lenged by V.parahaemolyticus,V.anguillarum and Aeromonas hydrophila.Up-regulated cryptocyanin is documented in the hemolymph and up-regulated calexcitin,wingless(fragment)and tachykinin-related peptide in the muscle as acute-phase proteins.All the four altered proteins were responsible for bacterial stress,but cryptocyanin seemed to be a memory response protein against the chal-lenge by a live bacterium after immunization of the live cells.These up-regulated proteins can be indicative of an understanding of immunity of a crab.展开更多
Transglutaminase(TGase)is a key enzyme that mediates hemolymph coagulation and is thought to contribute to the elimination of pathogenic microorganisms in invertebrates.The objective of this study was to elucidate the...Transglutaminase(TGase)is a key enzyme that mediates hemolymph coagulation and is thought to contribute to the elimination of pathogenic microorganisms in invertebrates.The objective of this study was to elucidate the involvement of TGase in insect immune responses via functional analysis of this enzyme in the oriental armyworm,Mythimna separata,using recombinant proteins and RNA interference technique.We identified two TGase genes,mystgase1 and mystgase2,in Mythimna separata and found that both genes are expressed in all surveyed tissues in M.separata larvae.Significant changes were induced in hemocytes following Escherichia coli injection.Injection of Gram-positive bacteria(Micrococcus luteus)and Gram-negative bacteria(Escherichia coli and Serratia marcescens)into larvae triggered a time-specific induction of both mystgase1 and mystgase2 in hemocytes.Recombinant MysTGase1 and MysTGase2 proteins bound to both E.coli and M.luteus,localizing within bacterial clusters and resulting in agglutination in a Ca^(2+)-dependent manner.The hemocytes of larvae injected with recombinant MysTGase1 or MysTGase2 exhibited enhanced phagocytic ability against E.coli,improved in vivo bacterial clearance,and increased resistance to S.marcescens,decreasing larval mortality rate.Conversely,RNA interference targeting mystgase1 or mystgase2 significantly reduced hemocyte phagocytic capability,decreased bacterial clearance,and increased susceptibility to S.marcescens infection,thereby increasing larval mortality rate.The findings of this study are anticipated to expand our understanding of the function of TGases within insect immune responses and may contribute to developing new pest control strategies.展开更多
Innate immunity is essential for the wellbeing of vertebrates and invertebrates. Key components of this defense system include pattern recognition receptors that bind to infectious agents, extra-and intra-cellular pro...Innate immunity is essential for the wellbeing of vertebrates and invertebrates. Key components of this defense system include pattern recognition receptors that bind to infectious agents, extra-and intra-cellular proteins that relay signals, as well as molecules and cells that eliminate pathogens. We have been studying the defense mechanisms in a biochemical model insect, Manduca sexta. In this insect, hemolin, peptidoglycan recognition proteins, β-1,3-glucan recognition proteins and C-type lectins detect microbial surface molecules and induce immune responses such as phagocytosis, nodulation, encapsulation, melanization and production of antimicrobial peptides. Some of these responses are mediated by extracellular serine proteinase pathways. The proteolytic activation of prophenoloxidase （proPO） yields active phenoloxidase （PO） which catalyzes the formation of quinones and melanin for wound healing and microbe killing. M. sexta hemolymph proteinase 14 （HP 14） precursor interacts with peptidoglycan or β-1,3-glucan, autoactivates, and leads to the activation of other HPs including HP21 and proPO-activating proteinases （PAPs）. PAP-1, -2 and -3 cut proPO to generate active PO in the presence of two serine proteinase homologs. Inhibition of the proteinases by serpins and association of the proteinase homologs with bacteria ensure a localized defense reaction. M. sexta HP1, HP6, HP8, HP17 and other proteinases may also participate in proPO activation or processing of spatzle and plasmatocyte spreading peptide.展开更多
基金supported by the National Natural Science Foundation of China(No.42277280)the Research Foundation of the Education Bureau of Hunan Province,China(Nos.21A0048 and 22A0059)the Natural Science Foundation of Hunan Province of China(No.2022JJ50168).
文摘Cadmium(Cd)is a common environmental heavy metal pollutant known for its toxic effects on various plants and animals.Cd contamination also poses a threat to Pardosa pseudoannulata,a predator in rice paddies.Results indicate that Cd pollution significantly reduced the levels of Metallothionein,phenoloxidase,and Ca^(2+).Furthermore,through metabolomics and 16S rDNA sequencing,alterations in the gut microbiota and immune-related metabolites of P.pseudoannulata were identified.The abundance of harmful bacteria such as Staphylococcus increased significantly,while levels of l-Tyrosine,glutathione disulfide,l-Dopa,among others,decreasedmarkedly.Pearson correlation analysis revealed significant associations between various gut microbes and immune-related metabolites,suggesting that Cd may alter the immune system of P.pseudoannulata by modulating its gut microbiota.
基金Supported by the National Natural Science Foundation of China(No.30600458)
文摘Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based purification(ClinProt system) and matrix-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) to characterize shrimp hemolymph peptides.Shrimp serum and plasma were used as the source of samples for comparative analysis,and it was found that serum was more suitable for shrimp hemolymph peptidomic analysis.To screen potential specific biomarkers in serum of immune-challenged shrimps,we applied magnetic bead-based MALDI-TOF MS to serum samples from 10 immune-challenged and 10 healthy shrimps.The spectra were analyzed using FlexAnalysis 3.0 and ClinProTools 2.1 software.Thirteen peptide peaks significantly different between the two groups were selected as candidate biomarkers of lipopolysaccharide(LPS)-infection.The diagnostic model established by genetic algorithm using five of these peaks was able to discriminate LPS-challenged shrimps from healthy control shrimps with a sensitivity of 90% and a specificity of 100%.Our approach in MALDITOF MS-based peptidomics is a powerful tool for screening bioactive peptides or biomarkers derived from hemolymph,and will help to enable a better understanding of the innate immune response of shrimps.
文摘Experiments showed the agglutinin activity in the hemolymph of both spring wild parents and autumncultured individuals of the shrimp Penaeus chinensis Osbeck with the body length of 12 - 16 cm as all 32. Thehemolymph of the above shrimps showed agglutinant reaction to all A-, B- and O-type human erythrocytes, with thereaction titer equal to that of rabbit blood cells. However, the agglutinin activity level was enhanced after being in-duced by Vibro, which was 128 after 24 h of induction and reached the peak value 256 after 48 h of induction, thendecreased gradually.
基金the Scientific Research Project of Huaqiao University(No.605-50X18005)Guangxi Bagui Youth Scholars ProgrammeGuangxi Recruitment Program of 100 Global Experts。
文摘Knowledge of dietary compositions is essential to the recovery of the dwindling populations of horseshoe crabs in China.The feeding habits of Tachypleus tridentatus and Carcinoscorpis rotundicauda of Pearl Bay in southwestern China were studied by stable isotope analysis.Hemolymph samples of the two species were obtained from two age groups living in different habitats(inner,central,and outer Bay).In addition,their potential food source samples were collected in May 2019.Results showed that the mean values ofδ^(13)C andδ^(15)N in C.rotundicauda individuals ranged from−19.01‰to−16.47‰and from 10.49‰to 13.5‰,respectively,and those of T.tridentatus ranged from−19.12‰to−14.96‰and from 8.78‰to 13.48‰,respectively.These values indicated that the horseshoe crabs have a wide variety of food sources and therefore are highly omnivorous.No remarkably correlations were found betweenδ^(13)C andδ^(15)N values and individual widths in the two species.Thirteen potential food sources were selected,in which Sipunculus nudus,Batillaria zonalisi,and Ulva lactuca contributed largely to the dietary compositions of the two species of horseshoe crabs.No significant correlation was found between the feeding habits and habitat geochemical characteristics of the horseshoe crabs.All these results possibly indicate the high diversity of their food sources and the indiscriminate dietary compositions of the two Asian horseshoe crabs.
基金Supported by the National Natural Science Foundation of China （No.30170742） and National Key Foundational Research Project of China （No.G1999012012）.
文摘Mixtures of hemolymph from Chlamys farreri with three different anticoagulant solutions were incu-bated for an hour in vitro, then the ultrastructural alterations of hemocytes were observed, and the aggre-gation rate was analyzed by using transmission electron microscropy and flow cytometry respectively. The results showed that Formula 3 （glucose 20.8 g L^-1; EDTA 20mM; sodium chloride 20 g L^-1; Tris-HCI 0.05M;pH 7.4）was the desirable anticoagulant solution for C. farreri hemocytes. Further phagocytosis assay showed that no obvious negative effect was given to the hemocyte phagocytic activity when using Formula 3 as the anticoagulant solution.
文摘Insects are well recognized as a source of potentially useful compounds for modern medicine. Development of anticancer drugs from natural resources has been performed throughout the world. In the present study, anticancer activity of the hemolymph and fat body of Sarcophaga argyrostoma third larval instars is assayed against human breast cancer cell line (MDA-MB-231 cells). The cytotoxicity of the hemolymph and fat body samples were determined. The results showed that growth of MDA-MB-231 cells was inhibited at different concentrations upon 24 h of exposure. There is no inhibitory activity against Vero cells under these experimental conditions. Protein profile of the hemolymph and fat body were extracted and separated using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Protein analysis showed differences in number of electrophoretic protein bands with different molecular weights between treated and control larvae. The present work assumes that hemolymph and fat body tissue have cytotoxicity against MDA-MB-231 cells and these results exhibit that larvae from S. argyrostoma could be used as a good source for developing anti-cancer agents and knowledge of these anti-cancer compounds will lead to better control of human disease.
文摘This study investigated the effects of silkworm hemolymph-derived storage protein 2 (SP2) on the whitening process in mouse B16F1 melanoma cells. After the cells were treated with various concentrations of SP2 (0.1 - 1.0 mg/mL), cytotoxicity, melanin contents, and differences in mRNA and protein expression associated with melanogenesis were observed. No cytotoxicity was observed when cells were treated with SP2, even with increased SP2 concentrations of up to 2.0 mg/mL. When treated with various SP2 concentrations in the cells, the protein and mRNA expression of tyrosinase were dose-dependently decreased, respectively, and inhibition of tyrosinase was further increased by 50.0% with increasing SP2 concentration of 1.0 mg/mL. Expression mRNAs coding tyrosinase related protein-1 and protein-2 (TRP-1 and TRP-2) was also significantly decreased in a dose dependent manner. When measuring the melanin content in melanoma cells, SP2 at 1 mg/mL inhibited melanin synthesis by 73.5% compared with non-treated cells. The inhibitory effect was 2.8-fold higher than that obtained using arbutin as a positive control. This study demonstrates that SP2, as a whitening material, is capable of suppressing melanin synthesis through the downregulation of proteins and genes in the melanin biosynthesis pathway.
基金Supported by Science and Technology Development Program of Jilin Province(201105087)~~
文摘[Objective] The research aimed to study the activity and dynamic changes of phenoloxidase in Locusta migratoria manilensis（Meyen） .[Method] Two enzyme solutions of insect body and hemolymph of Locusta migratoria manilensis（Meyen） were prepared by different extract methods,the activity and dynamic changes of phenoloxidase in different sites were measured.[Result] The phenoloxidase activity in hemolymph was significantly higher than that in body fluid,and the activity in female individuals was higher than that in male individuals.The phenoloxidase activity in body fluid was gradually enhanced with the prolongation of laying time in air.The phenoloxidase activity in hemolymph of adult was also gradually enhanced with the prolongation of time.[Conclusion] Part of phenoloxidase was existed in tissues and cells of Locusta migratoria manilensis（Meyen） participating in insect development and construction,and the other part was existed in hemolymph playing an immunological and defensive role.The stability of phenoloxidase was very strong,which was the effective guarantee for wide adaptation ability and strong immunity of insect.
基金supported by the National Basic Re-search Program of China (973 Program,2005CB121003)the National High-Tech Research and Development Program of China (863 Program,2006AA10A118)the Doctoral Fund of Ministry of Education of China (20070335148)
文摘The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of successful cross breeding in silkworm. Protein samples from silk gland, hemolymph, and midgut were separated by 2-dimensional gel electrophoresis （2-DE）. In the three tissues the matched protein spots between Jinqiu and its cross parents were approximately 70% with approximately 30% specific protein spots. In the matched protein spots, 9-24% was differentially expressed representing up- and down-regulated expression. These specific protein spots might be either the newly appeared, which were produced from the genic interaction of cross parents＇ genes in cross breeding, or posttranscriptionally modified, which were produced from the different modifications on the same original proteins. These results indicate that it is important for a new successful breed, by cross breeding, relying on the actions of some newly produced functional proteins from genic interaction, in addition to marshaling excellent genes of cross parents.
基金supported by the National Key R&D Program of China(No.2022YFD2400305)the Earmarked Fund for Agriculture Seed Improvement Project of Shandong Province(Nos.2021ZLGX03,2022LZGCQY010 and 2021LZGC027)the China Agriculture Research System Project(No.CARS-49)。
文摘Crassostrea nippona is a valuable species for aquaculture with considerable potential for commercial oyster farming.However,it is vulnerable to changes in salinity levels in coastal environments.In this study,we investigated the impacts of low salinity stress on the physiological responses of C.nippona.The hemolymph osmolality could not reach equilibrium with the surrounding environmental osmolality that was below salinity 15 within 1 week.Cell expansion,cellular valuocation,decrease of gill cilia,increased apoptotic cells under salinity 10 were observed through microscopic techniques.The activities of immunity-related enzymes,including alkaline phosphatase(AKP),acid phosphatase(ACP),superoxide dismutase(SOD),and catalase(CAT),were significantly increased at salinity 10 compared with the control group.These findings highlight the vulnerability of C.nippona to low salinity stress and provide insights into the physiological changes in response to fluctuating salinity levels.Understanding these physiological responses is crucial for effective aquaculture management and developing strategies to mitigate the negative impacts of low salinity stress on C.nippona populations in coastal areas.
基金Funded by Projects under the Major State Basic Research Developmet Program(G1999012010)
文摘The effect of acute E. coli challenge on the anti-disease activity of scallops Argopecten irra-dians is examined. The treatments of scallop from which hemolymph samples were taken for study included (1) control scallops, (2) sham-injected scallops, (3) PSW-injected scallops and (4) E. coli-injected scallops. From the beginning, the anti-disease activities of scallops are deter -mined at 12 hr and 24 hr.The concentrations of circulating hemocytes, the total serum protein concentrations and the activities of alkaline phosphatase, acid phosphatase and superoxide dismutase in the scallops Argopecten irradians are determined.Injection with E. coli results in a significant elevation in the concentration of circulating hemocytes and in the alkaline phosphatase activity and a significant decline in the total serum protein concentration and in the superoxide dismutase activity at 24 hr postchallenge. It shows that metabolism of bay scallop is expedited to adopt the challenge.
文摘Insects in the Order Odonata are highly subject to infection by gregarine parasites. However, despite the important ecological roles that insects play in every ecosystem in which they exist, little research has been devoted to the description of insect immunity. Insects rely heavily on the rapid actions of innate immune mechanisms to prevent infection. We characterized the melanization response in the hemolymph of green darner dragonfly (Anax junius) nymphs. Incubation of chymotrypsin-activated hemolymph with L-DOPA resulted in volume- and time-dependent production of dopaquinone via the phenoloxidase (PO) enzyme, with biphasic accumulation of product. The PO activity was temperature-dependent, with a stepwise increase from 20℃ - 35℃ and maximum activity measured at 35℃ - 40℃. The formation of product was also inhibited in a concentration-dependent manner by diethylcarbonate, a specific inhibitor of PO activity, which indicated that the observed activity was due to the presence of PO enzyme. The rate of formation and quantity of melanin was dependent on exposure to different titers of bacteria. This is the first characterization of both PO activity and melanization response in green darner dragonflies.
基金This research work was financially supported National Natural Science Foundation of China(No.82003910)Guangdong Basic and Applied Basic Research Foundation(2020A1515110550)+2 种基金Funding Key R&D Program of Shandong Province(2022SFGC0105)Exploration Innovation Team(2021GXRC062),Jinan Talent Project for Universities(202228088)Key innovation Project of Qilu University of Technology(Shandong Academy of Sciences)(2022JBZ01-06).
文摘There is an increasing interest in discovering new antibacterial agents derived from nature to enhance the treatment of various bacterial infections.Defensins and their derived peptide fragments exhibit significant antibacterial activity without any cytotoxic effects,making them attractive features for potential novel antibacterial therapeutics.Crassostrea gigas,a traditional seafood that has been used worldwide for centuries,has its shells applied in Chinese medicine as Ostreae concha.In this study,bioinformatics analysis was used to obtain a novel antibacterial peptide,CGD-1,derived from marine Chinese medicine Ostreae concha.The physicochemical characterization and circular dichroism analysis results demonstrated that CGD-1 assembled into anα-helical structure in a simulated membrane environment,and it displayed antibacterial action against Gram-negative bacteria.The minimal inhibitory concentrations against both Pseudomonas aeruginosa ATCC27853 and Escherichia coli ATCC25922 were 25μM.CGD-1 was able to efficiently permeate the cell membrane.Changes in bacterial cell morphology were evaluated using a field emission scanning electron microscope.The results suggested that CGD-1 exerted its antibacterial activity through permeabilizing and disrupting the bacterial cell membrane.Therefore,CGD-1 may have potential applications in fighting against pathogenic bacteria such as P.aeruginosa and E.coli.
文摘Two-dimensional gel electrophoresis （2-DE） followed by matrix-assisted laser desorption ionization- time-of-flight/time-of-flight mass spectrometry （MS） analysis were used to charaterize the hemolymph proteomic profiles of the silkworm, Bombyx mori. At days 4 （V4） and 5 （VS） of the fifth （final） instar, when the larvae were at the fast-growing stage, we found dramatic changes in spots representing proteins having an approximate molecular weight （MW） of 30 kDa. Of these spots, four 30K proteins were highly up- regulated, implying a close association with the growth and development ofB. mori larvae. To understand the molecular basis and underlying mechanisms involved in development and metamorphosis, the proteome of whole hemolymph at V5 was analyzed using shotgun liquid chromatography tandem mass spectrometry with an LTQ-Orbitrap. A total of 108 proteins were identified without any false discovery hits. These proteins were involved in a variety of cellular functions, including metabolism, development, nutrient transport and reserve, and defense response. Gene ontology analysis showed that 3.4% of these proteins had nutrient reservoir activities and 5.7% were involved in the response to stimulus. Pathway analysis revealed that 22 proteins with common targets were involved in various cellular processes such as immunity, differentiation, proliferation and metamorphosis. These results suggested that some key factors such as the 30K proteins in hemolymph play important roles in B. mori growth and development. Moreover, the multiple functions of hemolymph may be operated by a complex biological network.
基金supported by the National Natu-ral Science Foundation of China(grants 31472042 and 31672368 to LT,31702053 to KL and 31702065 to HY)the Natural Science Foundation of Guangdong Province(grant 2017A030311024 to LT)。
文摘During insect larval-pupal metamorphosis,proteins in the hemolymph are absorbed by the fat body for the maintenance of intracellular homeostasis;however,the type of proteins and how these proteins are internalized into the fat body are unclear.In Bombyx mori,the developmental profiles o f total proteins in the hemolymph and fat body showed that hemolymph-decreased protein bands(55-100 kDa)were in accordance with those protein bands that increased in the fat body.Inhibition o f clathrin-dependent endocytosis predominantly blocked the transportation o f 55-100 kDa proteins from the hemolymph into the fat body,which was further verified by RNA interference treatment o f Bmclathrin.Six hexamerins were shown to comprise〜90%of the total identified proteins in both the hemolymph and fat body by mass spectrum(MS)analysis.In addition,hemolymphspecific proteins were mainly involved in material transportation,while fat body-specific proteins particularly participated in metabolism.In this paper,four hexamerins were found for the first time,and potential proteins absorbed by the fat body from the hemolymph through clathrin-dependent endocytosis were identified.This study sheds light on the protein absorption mechanism during insect metamorphosis.
基金supported by Hi-Tech Research and Development Program of China (No.2002AA603013)the Natural Science Foundation of Fujian Province (No.B0110005).
文摘Acute-phase response is documented to be a significant mechanism of innate immunity in vertebrates and invertebrates.In this study,proteomic methodologies were applied for different protein expressions in hemolymph of Scylla serrata challenged by Vibrio parahaemolyticus after immunization,and in muscles of the crabs separately chal-lenged by V.parahaemolyticus,V.anguillarum and Aeromonas hydrophila.Up-regulated cryptocyanin is documented in the hemolymph and up-regulated calexcitin,wingless(fragment)and tachykinin-related peptide in the muscle as acute-phase proteins.All the four altered proteins were responsible for bacterial stress,but cryptocyanin seemed to be a memory response protein against the chal-lenge by a live bacterium after immunization of the live cells.These up-regulated proteins can be indicative of an understanding of immunity of a crab.
基金supported by the Japan Society for the Promotion of Science,KAKENHI[Grant Number JP17K08157].
文摘Transglutaminase(TGase)is a key enzyme that mediates hemolymph coagulation and is thought to contribute to the elimination of pathogenic microorganisms in invertebrates.The objective of this study was to elucidate the involvement of TGase in insect immune responses via functional analysis of this enzyme in the oriental armyworm,Mythimna separata,using recombinant proteins and RNA interference technique.We identified two TGase genes,mystgase1 and mystgase2,in Mythimna separata and found that both genes are expressed in all surveyed tissues in M.separata larvae.Significant changes were induced in hemocytes following Escherichia coli injection.Injection of Gram-positive bacteria(Micrococcus luteus)and Gram-negative bacteria(Escherichia coli and Serratia marcescens)into larvae triggered a time-specific induction of both mystgase1 and mystgase2 in hemocytes.Recombinant MysTGase1 and MysTGase2 proteins bound to both E.coli and M.luteus,localizing within bacterial clusters and resulting in agglutination in a Ca^(2+)-dependent manner.The hemocytes of larvae injected with recombinant MysTGase1 or MysTGase2 exhibited enhanced phagocytic ability against E.coli,improved in vivo bacterial clearance,and increased resistance to S.marcescens,decreasing larval mortality rate.Conversely,RNA interference targeting mystgase1 or mystgase2 significantly reduced hemocyte phagocytic capability,decreased bacterial clearance,and increased susceptibility to S.marcescens infection,thereby increasing larval mortality rate.The findings of this study are anticipated to expand our understanding of the function of TGases within insect immune responses and may contribute to developing new pest control strategies.
文摘Innate immunity is essential for the wellbeing of vertebrates and invertebrates. Key components of this defense system include pattern recognition receptors that bind to infectious agents, extra-and intra-cellular proteins that relay signals, as well as molecules and cells that eliminate pathogens. We have been studying the defense mechanisms in a biochemical model insect, Manduca sexta. In this insect, hemolin, peptidoglycan recognition proteins, β-1,3-glucan recognition proteins and C-type lectins detect microbial surface molecules and induce immune responses such as phagocytosis, nodulation, encapsulation, melanization and production of antimicrobial peptides. Some of these responses are mediated by extracellular serine proteinase pathways. The proteolytic activation of prophenoloxidase （proPO） yields active phenoloxidase （PO） which catalyzes the formation of quinones and melanin for wound healing and microbe killing. M. sexta hemolymph proteinase 14 （HP 14） precursor interacts with peptidoglycan or β-1,3-glucan, autoactivates, and leads to the activation of other HPs including HP21 and proPO-activating proteinases （PAPs）. PAP-1, -2 and -3 cut proPO to generate active PO in the presence of two serine proteinase homologs. Inhibition of the proteinases by serpins and association of the proteinase homologs with bacteria ensure a localized defense reaction. M. sexta HP1, HP6, HP8, HP17 and other proteinases may also participate in proPO activation or processing of spatzle and plasmatocyte spreading peptide.
