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A novel marching-on-time (MOT) based scheme for solving a time domain integral equation (TDIE) was discussed. It was observed that the scheme was pertinent to the analysis of plane wave scattering from nonplanar doubly periodic structures with unit cells comprising perfect electrically conducting (PEC) elements and inhomogeneous dielectric volumes. This scheme aimed at rendering the solver applicable to the nonplanar structures. The resulted scheme was applied without any difficulty to nonplanar doubly periodic structures comprising PEC and sculptured dielectric substrates.

Carbon quantum dots (CQDs) are garnering attention for their broad applications. This study offers a detailed evaluation of the biomedical safety and health risks of carbon quantum dots (CQDs) with different surface modifications, addressing a key gap in their safe application. It focuses on three CQD types: diammonium citrate-based (CQDs-A), spermidine trihydrochloride-based (CQDs-S), and a combination (CQDs-A/S), analyzing their physicochemical properties, cytotoxicity, oxidative stress, inflammatory responses, and nephrotoxicity. While all CQDs were under 10 nm, their biological impacts varied. Positively charged CQDs-S and CQDs-A/S showed significant cytotoxicity in HEK293 cells, inducing oxidative stress but not activating NLRP3 inflammasome, indicating a limited inflammatory response. Renal integrity remained unaffected, with stable zonula occludens 2 expression and unaltered renal markers. In vivo studies in BALB/c mice further supported the safety of CQDs, showing no organ damage or kidney pathology at high doses. The findings underscore the potential for safe biomedical use of CQDs, particularly when their retention time is minimized. This research makes a novel contribution by linking CQDs' surface charge to cytotoxic effects and oxidative stress, providing key insights into their safe use in biomedicine and filling a critical gap in nanoparticle toxicity studies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The early phases of clathrin mediated endocytosis are organized through a highly complex interaction network mediated by clathrin associated sorting proteins (CLASPs) that comprise long intrinsically disordered regions (IDRs). AP180 is a CLASP exclusively expressed in neurons and comprises a long IDR of around 600 residues, whose function remains partially elusive. Using NMR spectroscopy, we discovered an extended and strong interaction site within AP180 with the major adaptor protein AP2, and describe its binding dynamics at atomic resolution. We find that the 70 residue-long site determines the overall interaction between AP180 and AP2 in a dynamic equilibrium between its bound and unbound states, while weaker binding sites contribute to the overall affinity at much higher concentrations of AP2. Our data suggest that this particular interaction site might play a central role in recruitment of adaptors to the clathrin coated pit, whereas more transient and promiscuous interactions allow reshaping of the interaction network until cargo uptake inside a coated vesicle., (© 2024. The Author(s).)

Pollen germination is a crucial process in the life cycle of flowering plants, signifying the transition of quiescent pollen grains into active growth. This study delves into the dynamic changes within organelles and the pivotal role of autophagy during lily pollen germination. Initially, mature pollen grains harbor undifferentiated organelles, including amyloplasts, mitochondria, and the Golgi apparatus. However, germination unveils remarkable transformations, such as the redifferentiation of amyloplasts accompanied by starch granule accumulation. We investigate the self-sustained nature of amylogenesis during germination, shedding light on its association with osmotic pressure. Employing BODIPY 493/503 staining, we tracked lipid body distribution throughout pollen germination, both with or without autophagy inhibitors (3-MA, NEM). Typically, lipid bodies undergo polarized movement from pollen grains into elongating pollen tubes, a process crucial for directional growth. Inhibiting autophagy disrupted this essential lipid body redistribution, underscoring the interaction between autophagy and lipid body dynamics. Notably, the presence of tubular endoplasmic reticulum (ER)-like structures associated with developing amyloplasts and lipid bodies implies their participation in autophagy. Starch granules, lipid bodies, and membrane remnants observed within vacuoles further reinforce the involvement of autophagic processes. Among the autophagy inhibitors, particularly BFA, significantly impede germination and growth, thereby affecting Golgi morphology. Immunogold labeling substantiates the pivotal role of the ER in forming autophagosome-like compartments and protein localization. Our proposed speculative model of pollen germination encompasses proplastid differentiation and autophagosome formation. This study advances our understanding of organelle dynamics and autophagy during pollen germination, providing valuable insights into the realm of plant reproductive physiology., (© 2024. The Author(s).)

Crohn's disease (CD) is caused by immune, environmental, and genetic factors. It can involve the entire gastrointestinal tract, and although its prevalence is rapidly increasing its etiology remains unclear. Emerging biological and small-molecule drugs have advanced the treatment of CD; however, a considerable proportion of patients are non-responsive to all known drugs. To achieve a breakthrough in this field, innovations that could guide the further development of effective therapies are of utmost urgency. In this review, we first propose the innovative concept of pan-lymphatic dysfunction for the general distribution of lymphatic dysfunction in various diseases, and suggest that CD is the intestinal manifestation of pan-lymphatic dysfunction based on basic and clinical preliminary data. The supporting evidence is fully summarized, including the existence of lymphatic system dysfunction, recognition of the inside-out model, disorders of immune cells, changes in cell plasticity, partial overlap of the underlying mechanisms, and common gut-derived fatty and bile acid metabolism. Another benefit of this novel concept is that it proposes adopting the zebrafish model for studying intestinal diseases, especially CD, as this model is good at presenting and mimicking lymphatic dysfunction. More importantly, the ensuing focus on improving lymphatic function may lead to novel and promising therapeutic strategies for CD., Competing Interests: Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article., (©The Author(s) 2024. Published by Baishideng Publishing Group Inc. All rights reserved.)

To identify cognitive function in Chinese breast cancer survivors. Research questions were: is cognitive function was associated with breast cancer and/or chemotherapy treatment and/or psychological functioning:? and did women with breast cancer experience more cognitive and psychological issues than age-matched women without cancer? Breast cancer survivors with chemotherapy (n = 106, mean age = 50.2 ± 9.5), breast cancer survivors without chemotherapy (n = 100, mean age = 50.5 ± 10.0) and matched healthy controls (n = 96, mean age = 47.9 ± 9.1) completed a battery of cognitive and psychosocial functioning. Demographic characteristics were also collected. The Perceived Cognitive Impairment score for cancer groups was significantly higher than for the healthy group (p = 0.04), but not between the cancer groups. Processing speed was significantly slower in the cancer groups than in the healthy group (both p < 0.001), but not between the cancer groups. Age, living status and education were significantly associated with the FACT-Cog (all p < 0.05). The correlations between the FACT-Cog score and BSI score were strong (r = 0.60 p < 0.01), and between the HADS anxiety and depression scales were strong (r = 0.53 and 0.50, p < 0.01) but correlations were weaker between performance based cognitive tests and measures of psychological functioning. Breast cancer groups indicated more cognitive impairment and reduced psychological functioning compared to the healthy group. However, there was no differences between the breast cancer groups. Chinese breast cancer survivors experienced excess cognitive impairment not associated with usual ageing. Assessment and intervention to address cognitive impairment should be made available to breast cancer survivors., (© 2023. The Author(s).)

Using unnatural amino acid mutagenesis, we made a mutant of CaMKII that forms a covalent linkage to Calmodulin upon illumination by UV light. Like wild-type CaMKII, the L308BzF mutant stoichiometrically binds to Calmodulin, in a calcium-dependent manner. Using this construct, we demonstrate that Calmodulin binding to CaMKII, even under these stochiometric conditions, does not perturb the CaMKII oligomeric state. Furthermore, we were able to achieve activation of CaMKII L308BzF by UV-induced binding of Calmodulin, which, once established, is further insensitive to calcium depletion. In addition to the canonical auto-inhibitory role of the regulatory segment, inter-subunit crosslinking in the absence of CaM indicates that kinase domains and regulatory segments are substantially mobile in basal conditions. Characterization of CaMKII L308BzF in vitro, and its expression in mammalian cells, suggests it could be a promising candidate for control of CaMKII activity in mammalian cells with light., (© 2023 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)

The dodecameric protein kinase CaMKII is expressed throughout the body. The alpha isoform is responsible for synaptic plasticity and participates in memory through its phosphorylation of synaptic proteins. Its elaborate subunit organization and propensity for autophosphorylation allow it to preserve neuronal plasticity across space and time. The prevailing hypothesis for the spread of CaMKII activity, involving shuffling of subunits between activated and naive holoenzymes, is broadly termed subunit exchange. In contrast to the expectations of previous work, we found little evidence for subunit exchange upon activation, and no effect of restraining subunits to their parent holoenzymes. Rather, mass photometry, crosslinking mass spectrometry, single molecule TIRF microscopy and biochemical assays identify inter-holoenzyme phosphorylation (IHP) as the mechanism for spreading phosphorylation. The transient, activity-dependent formation of groups of holoenzymes is well suited to the speed of neuronal activity. Our results place fundamental limits on the activation mechanism of this kinase., Competing Interests: IL, LH, PJ, AC, CW, AF, FH, MW, FL, AP No competing interests declared, (© 2023, Lučić et al.)

Stroke is a leading cause of adult disability worldwide, and better drugs are needed to promote functional recovery after stroke. Growing evidence suggests the critical role of network excitability during the repair phase for stroke recovery. Here, we show that β-hydroxybutyrate (β-HB), an essential ketone body (KB) component, is positively correlated with improved outcomes in patients with stroke and promotes functional recovery in rodents with stroke during the repair phase. These beneficial effects of β-HB depend on HDAC2/HDAC3-GABA transporter 1 (GAT-1) signaling-mediated enhancement of excitability and phasic GABA inhibition in the peri-infarct cortex and structural and functional plasticity in the ipsilateral cortex, the contralateral cortex, and the corticospinal tract. Together with available clinical approaches to elevate KB levels, our results offer a clinically translatable means to promote stroke recovery. Furthermore, GAT-1 can serve as a pharmacological target for developing drugs to promote functional recovery after stroke., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Cross-linking mass spectrometry (XL-MS) is an attractive method for the proteome-wide characterization of protein structures and interactions. Currently, the depth of in vivo XL-MS studies is lagging behind the established applications to cell lysates, because cross-linking reagents that can penetrate intact cells and strategies to enrich cross-linked peptides lack efficiency. To tackle these limitations, we have developed a phosphonate-containing cross-linker, tBu-PhoX, that efficiently permeates various biological membranes and can be robustly enriched using routine immobilized metal ion affinity chromatography. We have established a tBu-PhoX-based in vivo XL-MS approach that enables cross-links in intact human cells to be identified in high numbers with substantially reduced analysis time. Collectively, the developed cross-linker and XL-MS approach pave the way for the comprehensive XL-MS characterization of living systems., (© 2021 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Oil bodies are essential energy storage organelles that are generally present in the seeds of plants. Caleosin protein has been identified in the seed oil bodies of Cycas revoluta seed. In this study, mature Cycas revoluta pollen grains were collected from cycad elliptical cones. However, the isolation and identification of oil bodies protein from mature Cycas revoluta grains have never been experimentally determined. Ultrastructural studies have shown that the oil bodies were present in pollen Cycas revoluta grains. Lipid analysis showed that oil bodies are predominantly composed of triacylglycerol. Complete cDNA fragments encoding the caleosin were obtained by PCR cloning. Phylogenetic tree analyzes showed that cycad pollen caleosin is closely related to the caleosin of Cycas revoluta seeds. Fresh Cycas revoluta pollen grains were allowed to germinate for 48 h in a germination medium containing 2% sucrose and 0.01% H 3 BO 3. Pollen germination were recorded periodically from day one to day 10 and the results showed that the Cycas revoluta pollen elongate the tube and increasing of triacylglycerol(TAG) after 4 days.

The mucosal immune system is the host's first line of defense against invasion by foreign pathogens. Gelatin nanoparticles (GNPs) are suitable carriers for the delivery of antigens via various routes of administration. In the present study, GNPs were modified with polyethyleneimine (PEI), a positively charged polymer. Then, ovalbumin (OVA) and polyinosinic:polycytidylic acid (poly(I:C)), an immunostimulant, were adsorbed onto the surface of the positively charged GNPs. We assessed whether GNPs could act as an effective mucosal vaccine that is capable of inducing both mucosal and systemic immune responses. The results showed that GNPs effectively adsorbed OVA/poly(I:C), facilitated cellular uptake by RAW 264.7 macrophage cells and murine bone marrow-derived dendritic cells (BMDCs) in vitro, and led to increased expression of the maturation markers CD80 and CD86 on BMDCs. Furthermore, GNPs induced increased secretion of proinflammatory cytokines in both RAW 264.7 and BMDCs. C57BL/6 mice that were intranasally twice-immunized with OVA/poly(I:C)-loaded GNPs produced high levels of serum OVA-specific IgG antibodies and secretory IgA in nasal and lung lavage. Spleen cells from immunized mice were collected and re-stimulated with OVA, and results showed significantly augmented production of IFN-γ, IL-4, IL-5, and IL-6 in mice that received OVA/poly(I:C)-loaded GNPs. Moreover, intranasal immunization with OVA/poly(I:C)-loaded GNPs resulted in the inhibition of EG7 tumor growth in C57BL/6 mice. Taken together, these results indicate that nasal administration of OVA/poly(I:C)-loaded GNPs elicited effective mucosal and systemic immune responses, which might be useful for further applications of antigen delivery. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1228-1237, 2019., (© 2018 Wiley Periodicals, Inc.)

In this paper, nitrogen incorporated ultrananocrystalline diamond (NUNCD) films were fabricated for use as electrodes to detect dopamine. The NUNCD electrodes achieved high sensitivity, great selectivity, and excellent detection limits for dopamine sensing. The NUNCD electrode, fabricated as a potential sensitive biosensor for dopamine without any catalyst or mediators, demonstrated good activity for the direct detection of dopamine by simply putting the bare NUNCD electrode into a dopamine solution. Furthermore, the marked selectivity of the NUNCD electrode is very favorable for the determination of dopamine (DA) concentration (0.32 μM) in the presence of ascorbic acid (AA) and uric acid (UA). Considering dopamine detection in real biological fluid samples, the NUNCD electrode performed excellently with a detection limit of 0.39 μM and a high recovery ranging from 90-120%, revealing that NUNCD electrodes have promising use in the sensing of dopamine.

Objective: The aim of the study was to estimate breast cancer risk conferred by individual single-nucleotide polymorphisms of breast cancer susceptibility genes., Methods: We analyzed the 48 tagging single-nucleotide polymorphisms of 8 breast cancer susceptibility genes involved in the monoubiquitinated FANCD2-DNA damage repair pathway in 734 Chinese women with breast cancer and 672 age-matched healthy controls., Results: Forty-five tagging single-nucleotide polymorphisms were successfully genotyped by SNPscan, and the call rates for each tagging single-nucleotide polymorphisms were above 98.9%. We found that 13 tagging single-nucleotide polymorphisms of 5 genes ( Parter and localizer of Breast cancer gene2 ( PALB2), Tumour protein 53 ( TP53), Nijmegen breakage syndrome 1, Phosphatase and tensin homolog deleted from chromosome 10 ( PTEN), and Breast cancer gene 1 ( BRCA1-interacting protein 1)) were significantly associated with breast cancer risk. A total of 5 tagging single-nucleotide polymorphisms (rs2299941 of PTEN, rs2735385, rs6999227, rs1805812, and rs1061302 of Nijmegen breakage syndrome 1) were tightly associated with breast cancer risk in sporadic cases, and 5 other tagging single-nucleotide polymorphisms (rs1042522 of TP53, rs2735343 of PTEN, rs7220719, rs16945628, and rs11871753 of BRCA1-interacting protein 1) were tightly associated with breast cancer risk in familial and early-onset cases., Conclusions: Some of the tagging single-nucleotide polymorphisms of 5 genes ( PALB2, TP53, Nijmegen breakage syndrome 1, PTEN, and BRCA1-interacting protein 1) involved in the monoubiquitinated FANCD2-DNA damage repair pathway were significantly associated with breast cancer risk.

Objective: To investigate the oil body protein and function in seeds of mature seagrass, Thalassia hemprichii., Results: Seeds of mature seagrass T. hemprichii when stained with a fluorescent probe BODIPY showed the presence of oil bodies in intracellular cells. Triacylglycerol was the major lipid class in the seeds. Protein extracted from seagrass seeds was subjected to immunological cross-recognition with land plant seed oil body proteins, such as oleosin and caleosin, resulting in no cross-reactivity. An oleosin-like gene was found in seagrass seeds. Next generation sequencing and sequence alignment indicated that the deduced seagrass seed oleosin-like protein has a central hydrophobic domain responsible for their anchoring onto the surface of oil bodies. Phylogenetic analysis showed that the oleosin-like protein was evolutionarily closer to pollen oleosin than to seed oleosins., Conclusion: Oil body protein found in seagrass seeds represent a distinct class of land seed oil body proteins.

Unique proteins including steroleosin, caleosin, oleosin-L, and oleosin-G have been identified in seed oil bodies of pine (Pinus massoniana). In this study, mature pollen grains with wing-like bladders were collected from pine (Pinus elliottii). Ultrastructural studies showed that oil bodies were present in pollen grains, but not the attached bladders, and the presence of oil bodies was further confirmed by fluorescent staining with BODIPY 493/503. Stable oil bodies were successfully purified from pine pollen grains, and analyzed to be mainly composed of triacylglycerols. Putative oleosin and caleosin in pine pollen oil bodies were detected by immunoassaying with antibodies against sesame seed caleosin and lily pollen oleosin. Complete cDNA fragments encoding these two pollen oil-body proteins were obtained by PCR cloning. Sequence alignment showed that pine pollen caleosin (27 kDa) was highly homologous to pine seed caleosin (28 kDa) except for the lack of an appendix of eight residues at the C-terminus in accord with the 1 kDa difference in their molecular masses. Pine pollen oleosin (15 kDa) was highly homologous to pine seed oleosin-G (14 kDa) except for an insertion of eight residues at the N-terminus in accord with the 1 kDa difference in their molecular masses., (Copyright © 2016 Elsevier Masson SAS. All rights reserved.)

To control the degradation rate of medical magnesium in body fluid environment, biocompatible films composed of Mussel Adhesive Protein (Mefp-1) and chitosan were electrodeposited on magnesium surface in cathodic constant current mode. The compositions and structures of the films were characterized by atomic force microscope (AFM), scanning electron microscope (SEM) and infrared reflection absorption spectroscopy (IRAS). And the corrosion protection performance was investigated using electrochemical measurements and immersion tests in simulated body fluid (Hanks' solution). The results revealed that Mefp-1 and chitosan successfully adhered on the magnesium surface and formed a protective film. Compared with either single Mefp-1 or single chitosan film, the composite film of chitosan/Mefp-1/chitosan (CPC (chitosan/Mefp-1/chitosan)) exhibited lower corrosion current density, higher polarization resistance and more homogenous corrosion morphology and thus was able to effectively control the degradation rate of magnesium in simulated body environment. In addition, the active attachment and spreading of MC3T3-E1 cells on the CPC film coated magnesium indicated that the CPC film was significantly able to improve the biocompatibility of the medical magnesium., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Objective: Otitis media (OM), as a common infectious disease, is a major cause of hearing impairment among the general population. OM remains a major public health threat in the Pacific islands, but the risks of OM have not been thoroughly explored in this region. The objective of this study is to investigate the prevalence, clinical features, and quality-of-life impacts of OM in Fiji., Methods: In the medical service trip entitled "Healing and Hope - Taiwan Cathay Heart and Hearing Medical Mission to Fiji" (TCHHMMF), we conducted a cross-sectional OM survey study in Suva and Sigatoka areas (Korolevu, Cuvu, and Lomawai) in the summer of 2015. The otitis media - 6 (OM-6) was used to survey the OM-related quality of life., Results: In the 467 pediatric patients (aged 0-18 years old) screened, 13 (2.78%) have acute otitis media (AOM), 37 (7.92%) have otitis media with effusion (OME), and 19 (4.1%) have chronic otitis media (COM). Age (OR 0.53, 95% CI: 0.36-0.77) is a significant predictor of AOM, whereas male gender (OR 2.46, 95% CI: 1.13-5.37), smoke exposure (OR 2.81, 95% CI: 1.01-7.82), and concomitant chronic sinusitis (OR 6.05, 95% CI: 2.31-15.88) are significant predictors of OME. The mean OM-6 item scores are highest in caregiver concerns (3.8), physical suffering (3.7), and hearing loss (3.4) domains., Conclusion: OM is an important primary care disease in Fiji that remains under-served. It is critical to educate professionals, parents, and patients to detect and to improve care for OM., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Three classes of integral proteins termed oleosin, caleosin and steroleosin have been identified in seed oil bodies of diverse angiosperm species. Recently, two oleosin isoforms and one caleosin were identified in megagametophyte oil bodies of pine (Pinus massoniana), a representative gymnosperm species. In this study, a putative steroleosin of approximately 41 kDa was observed in isolated oil bodies of pine megagametophytes, and its corresponding cDNA fragment was obtained by PCR cloning and further confirmed by mass spectrometric analysis. Phylogenetic tree analysis showed that pine steroleosin was evolutionarily more closely-related to steroleosin-B than steroleosin-A found in angiosperm seed oil bodies. As expected, artificial oil bodies constituted with recombinant steroleosin over-expressed in Escherichia coli were less stable and larger than native pine oil bodies. Filipin staining of artificial oil bodies sheltered by recombinant steroleosin with or without its sterol binding domain showed that the sterol binding domain was responsible for the sterol binding capability of steroleosin. Sterol-coupling dehydrogenase activity was demonstrated in artificial oil bodies constituted with recombinant steroleosin as well as in purified pine oil bodies., (Copyright © 2016 Elsevier Masson SAS. All rights reserved.)

Symbiodinium is a dinoflagellate that plays an important role in the physiology of the symbiotic relationships of Cnidarians such as corals and sea anemones. However, it is very difficult to cultivate free-living dinoflagellates after being isolated from the host, as they are very sensitive to environmental changes. How these symbiont cells are supported by the host tissue is still unclear. This study investigated the characteristics of Symbiodinium cells, particularly with respect to the morphological variability and distinct protein profiles of both cultured and endosymbiotic Symbiodinium which were freshly isolated from Exaiptasia pulchella. The response of the cellular morphology of freshly isolated Symbiodinium cells kept under a 12 h L:12 h D cycle to different temperatures was measured. Cellular proliferation was investigated by measuring the growth pattern of Symbiodinium cells, the results of which indicated that the growth was significantly reduced in response to the extreme temperatures. Proteomic analysis of freshly isolated Symbiodinium cells revealed twelve novel proteins that putatively included transcription translation factors, photosystem proteins, and proteins associated with energy and lipid metabolism, as well as defense response. The results of this study will bring more understandings to the mechanisms governing the endosymbiotic relationship between the cnidarians and dinoflagellates.

Mucosal surfaces contain specialized dendritic cells (DCs) that are able to recognize foreign pathogens and mount protective immunity. We previously demonstrated that intranasal administration of targeted galactosylated liposomes can elicit mucosal and systemic antibody responses. In the present study, we assessed whether galactosylated liposomes could act as an effective DC-targeted mucosal vaccine that would be capable of inducing systemic anti-tumor immunity as well as antibody responses. We show that targeted galactosylated liposomes effectively facilitated antigen uptake by DCs beyond that mediated by unmodified liposomes both in vitro and in vivo. Targeted galactosylated liposomes induced higher levels of pro-inflammatory cytokines than unmodified liposomes in vitro. C57BL/6 mice thrice immunized intranasally with ovalbumin (OVA)-encapsulated galactosylated liposomes produced high levels of OVA-specific IgG antibodies in their serum. Spleen cells from mice receiving galactosylated liposomes were restimulated with OVA and showed significantly augmented levels of IFN-γ, IL-4, IL-5 and IL-6. In addition, intranasal administration of OVA-encapsulated beta-galactosylated liposomes resulted in complete protection against EG7 tumor challenge in C57BL/6 mice. Taken together, these results indicate that nasal administration of a galactosylated liposome vaccine mediates the development of an effective immunity against tumors and might be useful for further clinical anti-tumoral applications., (Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Intracellular lipid droplets (LDs) have been proposed to play a key role in the mutualistic endosymbiosis between reef-building corals and the dinoflagellate endosymbiont Symbiodinium spp. This study investigates and identifies LD proteins in Symbiodinium from Euphyllia glabrescens. Discontinuous Percoll gradient centrifugation was used to separate Symbiodinium cells from E. glabrescens tentacles. Furthermore, staining with a fluorescent probe, Nile red, indicated that lipids accumulated in that freshly isolated Symbiodinium cells and lipid analyses further showed polyunsaturated fatty acids (PUFA) was abundant. The stable LDs were purified from endosymbiotic Symbiodinium cells. The structural integrity of the Symbiodinium LDs was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Protein extracts from the purified LDs revealed a major protein band with a molecular weight of 20 kDa, which was termed Symbiodinium lipid droplet protein (SLDP). Interestingly, immunological cross-recognition analysis revealed that SLDP was detected strongly by the anti-sesame and anti-cycad caleosin antibodies. It was suggested that the stable Symbiodinium LDs were sheltered by this unique structural protein and was suggested that SLDP might be homologous to caleosin to a certain extent.

Numerous oil bodies of 0.2-2 μm occupied approximately 80% of intracellular space in mature pine (Pinus massoniana) megagametophytes. They were stably isolated and found to comprise mostly triacylglycerols as examined by thin layer chromatography analysis and confirmed by both Nile red and BODIPY stainings. Fatty acids released from the triacylglycerols of pine oil bodies were mainly unsaturated, including linoleic acid (60%), adrenic acid (12.3%) and vaccenic acid (9.7%). Proteins extracted from pine oil bodies were subjected to immunological cross-recognition, and the results showed that three proteins of 28, 16 and 14 kDa were detected by antibodies against sesame seed caleosin, sesame oleosin-L and lily pollen oleosin-P, respectively. Complete cDNA fragments encoding these three pine oil-body proteins, tentatively named caleosin, oleosin-L and oleosin-G, were obtained by PCR cloning and further confirmed by mass spectrometric analysis. Consistently, phylogenetic tree analyses showed that pine caleosin was closely-related to the caleosin of cycad megagametophyte among known caleosin sequences. While pine oleosin-L was found clustered with seed oleosin isoforms of angiosperm species, oleosin-G was distinctively grouped with the oleosin-P of lily pollen. The oleosin-G identified in pine megagametophytes seems to represent a new class of seed oleosin isoform evolutionarily close to the pollen oleosin-P., (Copyright © 2014 Elsevier Masson SAS. All rights reserved.)

A red-eye colony was established in our laboratory in brown planthopper (BPH), Nilaparvata lugens (Stål), a major rice pest in Asia. Except for the red-eye phenotype, no other differences were observed between the wild-type (brown eye) and the mutant-type (red eye) in external characters. Genetic analysis revealed that the red-eye phenotype was controlled by a single autosomal recessive allele. Biological studies found that egg production and egg viability in the red-eye mutant colony were not significantly different from those in the wild-type BPH. Biochemical analysis and electronic microscopy examination revealed that the red-eye mutants contained decreased levels of both xanthommatin (brown) and pteridine (red) and reduced number of pigment granules. Thus, the changes of amount and ratio of the two pigments is the biochemical basis of this red-eye mutation. Our results indicate that the red-eye mutant gene (red) might be involved in one common gene locus shared by the two pigments in pigment transportation, pigment granule formation or some other processes., (© 2013 Institute of Zoology, Chinese Academy of Sciences.)

The stability of cnidarian-dinoflagellate (genus Symbiodinium spp.) endosymbioses depends on the regulation of nutrient transport between Symbiodinium populations and their hosts. Previously, we successfully induced the production of lipid droplets in the free-living cultured Symbiodinium (clade B) under the nitrogen-deprivation condition for 5 days. Therefore, the present study aimed at understanding the disruption of the endosymbiotic relationship between the cnidarians and dinoflagellates by nitrogen deprivation using Aiptasia pulchella as an example. Transmission electron micrographs revealed the formation of lipid droplets induced by nitrogen deprivation, and the lipid analyses further showed that polyunsaturated fatty acids were drastically enriched in Symbiodinium after 30 days of nitrogen deprivation, although these were unaffected after 5 days of nitrogen starvation. The present study also suggested that the host provided nitrogen to the symbiotic cells during short-term environmental stress. However, the relationship started to deteriorate after 30 days. These findings provide a more detailed understanding of the mechanisms of the symbiotic relationship between the symbiotic dinoflagellates in terms of the nitrogen source, which might provide more information for the explanation of the regulatory mechanism underlying endosymbiotic associations.

Stable cnidarian-dinoflagellate (genus Symbiodinium) endosymbioses depend on the regulation of nutrient transport between Symbiodinium populations and their hosts. It has been previously shown that the host cytosol is a nitrogen-deficient environment for the intracellular Symbiodinium and may act to limit growth rates of symbionts during the symbiotic association. This study aimed to investigate the cell proliferation, as well as ultrastructural and lipid compositional changes, in free-living Symbiodinium spp. (clade B) upon nitrogen (N)-deprivation. The cell proliferation of the N-deprived cells decreased significantly. Furthermore, staining with a fluorescent probe, boron dipyrromethane 493/503 (BODIPY 493/503), indicated that lipid contents progressively accumulated in the N-deprived cells. Lipid analyses further showed that both triacylglycerol (TAG) and cholesterol ester (CE) were drastically enriched, with polyunsaturated fatty acids (PUFA; i.e., docosahexaenoic acid, heneicosapentaenoic acid, and oleic acid) became more abundant. Ultrastructural examinations showed that the increase in concentration of these lipid species was due to the accumulation of lipid droplets (LDs), a cellular feature that have previously shown to be pivotal in the maintenance of intact endosymbioses. Integrity of these stable LDs was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Proteomic analyses of these LDs identified proteins putatively involved in lipid metabolism, signaling, stress response and energy metabolism. These results suggest that LDs production may be an adaptive response that enables Symbiodinium to maintain sufficient cellular energy stores for survival under the N-deprived conditions in the host cytoplasm.

LUFFACHITIN obtained from the residue of the sponge-like dried fruit of Luffa aegyptiaca was developed as a weavable skin substitute in this study. A chemical analysis revealed that LUFFACHITIN was composed of a copolymer containing N-acetyl-glucosamine (~40%) as a major monomer with a filamentary structure as demonstrated by both optical and scanning electron microscopy. The pulp-like white residue of the sponge-like dried fruit of Luffa aegyptiaca after treatment was then woven into a thin, porous membrane by filtration and lyophilization as a skin substitute for conducting wound-healing study on rats. The results indicated that the LUFFACHITIN membrane showed significant wound-healing enhancement (25 days to complete healing) compared to cotton gauze (>30 days), but not inferior to that of SACCHACHITIN. Furthermore, the LUFFACHITIN membrane had advantages of having a high yield, better physical properties for fabrication, and a more attractive appearance.

Oil bodies formed in Auxenochlorella protothecoides induced during limited nutrition had a coating of caleosin. The total lipid content obtained from A. protothecoides in unstressed cultures (first week) was ~210 mg/g compared to the 231 mg/g obtained in the third week (nutrient limited) and 290 mg/g obtained in the fourth week (nutrient limited). The proportion of total saturated fatty acids increased from 28 to 46 %, whereas that of total polyunsaturated fatty acids decreased from 52 to 35 %. The expression levels of the 28 kDa caleosin protein in A. protothecoides rose to a maximum up to 4 weeks; immunolocalization studies showed that caleosin was predominantly associated with the membranes of oil bodies.

Background: This study aimed to explore the relationship between cancer-related fatigue (CRF) and personality in patients with breast cancer after chemotherapy., Methods: A cross-sectional study was conducted to study the relationship between CRF and personality in breast cancer patients after chemotherapy. CRF and personality were measured by the cancer fatigue score and the Eysenck Personality Questionnaire, respectively., Results: A total of 300 breast cancer patients who had received chemotherapy were recruited to this study. Eysenck Personality Questionnaire scores of psychoticism, introversion, and extroversion in the patients were lower than the norm level (p < 0.01), but those of neuroticism and lie were higher than the norm level (p < 0.01). Multivariate analyses showed positive correlation between psychoticism and affective fatigue, neuroticism and total fatigue, and physical fatigue and cognitive fatigue. Multivariate analyses also showed negative correlation between introversion or extroversion and total fatigue, physical fatigue or affective fatigue, and lie and total fatigue or cognitive fatigue., Conclusions: There was CRF in patients with breast cancer after chemotherapy. Psychoticism, extroversion/introversion, neuroticism, and lie are correlated with CRF in breast cancer patients after chemotherapy., (Copyright © 2013 John Wiley & Sons, Ltd.)

The mucosal immune system produces secretory IgA (sIgA) as the first line of defense against invasion by foreign pathogens. Our aim was to develop a galactose-modified liposome as a targeted carrier which can be specifically recognized by macrophage, one of the most important antigen presenting cells. First, galactose was covalently conjugated with 1,2-didodecanoyl-sn-glycero-3-phosphoethanolamine (DLPE) to give a targeted ligand, a galactosyl lipid. The galactosyl lipid was then incorporated into a liposomal bilayer to form a galactosylated liposome carrier. Further, the ovalbumin (OVA) was encapsulated into the galactosylated liposome carriers and mice were intranasally immunized. Confocal laser scanning microscopy and flow cytometry analysis showed that the targeted galactosylated liposome carrier had a higher uptake rate than unmodified liposomes. The targeted galactosylated liposome induced higher levels of tumor necrosis factor-α and interleukin-6 production than unmodified liposomes (P<0.05). Furthermore, 6-week-old BALB/c female mice immunized with the OVA-encapsulated targeted galactosylated liposome had significantly higher OVA-specific s-IgA levels in the nasal and lung wash fluid (P<0.05). In addition, the targeted galactosylated liposome simultaneously augmented the serum IgG antibody response. In summary, the OVA-encapsulated targeted galactosylated liposome induced significantly higher mucosal IgA and systemic IgG antibody titers and is a potential antigen delivery carrier for further clinical applications., (Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Accumulation of oil bodies was successfully induced in a microalga, Chlorella sp., cultured in a nitrogen-limited medium. The oil bodies were initially assembled as many small entities (mostly 0.1-1 μm), and lately found as a major irregular compartment (>3 μm) occupying more than half of the cell space. Approximately, two thirds of oil bodies isolated from Chlorella cells were broken and formed a transparent oil layer on top of the milky compact layer of the remaining stable oil bodies after being washed with 0.1% triton X-100. The stable oil bodies mainly comprised triacylglycerols as examined by thin layer chromatography analysis and confirmed by both Nile red and BODIPY stainings. Integrity of these stable oil bodies was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Immunological cross-recognition revealed that a major protein of 29 kDa, tentatively identified as caleosin, was exclusively present in Chlorella oil bodies. Mass spectrometric analysis showed that the putative caleosin possessed a trypic fragment of 13 residues matching to that of a hypothetical caleosin in Picea sitchensis. With the aid of a degenerate primer designed according to the tryptic peptide, a complete cDNA fragment encoding this putative caleosin was obtained by PCR. Phylogenetic tree analysis supports that Chlorella caleosin is the most primitive caleosin found in oil bodies to date., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

Oil bodies of similar sizes were observed in the cells of embryo and aleurone layer of rice seeds, and remained their structural integrity in vitro after isolation. Comparably, two abundant oleosin isoforms were found in both preparations of oil bodies isolated from the embryo and the aleurone layer. Immunological detection and mass spectrometric analyses indicated that the two oleosin isoforms, termed oleosin-H and oleosin-L, in the embryo and those in the aleurone layer were identical proteins encoded by the same genes (BAF12898.1 and BAF15387.1 for oleosin-H and oleosin-L, respectively). In contrast, one caleosin was found in oil bodies isolated from the embryo but not those isolated from the aleurone layer. Immunological staining of rice seeds confirms that oleosin is present in both embryo and aleurone layer while caleosin exists only in embryo. Caleosin extracted from oil bodies of rice embryo migrated faster on SDS-PAGE in the presence of Ca(2+), in a manner identical to caleosin extracted from sesame oil bodies. Similar to other known monocot caleosins, the rice caleosin possesses an N-terminal appendix that is absent in dicotyledonous caleosins., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

The endosymbiotic relationship between coral hosts and dinoflagellates of the genus Symbiodinium is critical for the growth and productivity of coral reef ecosystems. Here, synchrotron radiation-based infrared microspectroscopy was applied to examine metabolite concentration differences between endosymbiotic (within the anemone Aiptasia pulchella) and free-living Symbiodinium over the light-dark cycle. Significant differences in levels of lipids, nitrogenous compounds, polysaccharides and putative cell wall components were documented. Compared with free-living Symbiodinium, total lipids, unsaturated lipids and polysaccharides were relatively enriched in endosymbiotic Symbiodinium during both light and dark photoperiods. Concentrations of cell wall-related metabolites did not vary temporally in endosymbiotic samples; in contrast, the concentrations of these metabolites increased dramatically during the dark photoperiod in free-living samples, possibly reflecting rhythmic cell-wall synthesis related to light-driven cell proliferation. The level of nitrogenous compounds in endosymbiotic cells did not vary greatly across the light-dark cycle and in general was significantly lower than that observed in free-living samples collected during the light. Collectively, these data suggest that nitrogen limitation is a factor that the host cell exploits to induce the biosynthesis of lipids and polysaccharides in endosymbiotic Symbiodinium.

Oleosin, the most abundant protein in oil bodies of all examined angiosperm seeds, has been demonstrated to serve as a structural protein to maintain the integrity of these lipid storage organelles. Caleosin, a minor protein in oil bodies of angiosperm seeds, is assumed to anchor to the organelles in a manner similar to oleosin, i.e., via its central hydrophobic domain with a unique proline-knot motif presumably responsible for the targeting to nascent oil bodies. Recently, we found that stable oil bodies in cycad seeds were mainly sheltered by caleosin in the absence of oleosin. This finding suggests that caleosin is competent as the major structural protein, just like oleosin, on the surface of seed oil bodies. A search of literature shows that genes putatively encoding for caleosin-, but not oleosin-like proteins, are present in more primitive species, such as algae and fungi. We hypothesize that oleosin may have been derived from caleosin after a long term of divergent evolution.

Oil bodies were observed in cells of both embryo and aleurone layers of mature adlay grains (Coix lachryma-jobi L. var. ma-yuen Stapf). Stable oil bodies were successfully isolated from the adlay grains. Thin-layer chromatography revealed that the contents stored in the adlay oil bodies were mainly neutral lipids (>90% triacylglycerols and about 5% diacylglycerols). The integrity of the isolated oil bodies was presumably maintained via electronegative repulsion and steric hindrance provided by their surface proteins. Immunological cross-recognition using antibodies against sesame oil-body proteins indicated that two oleosin isoforms (termed oleosin-H and oleosin-L) and one caleosin were present in the adlay oil bodies. Full-length cDNA fragments encoding these three unique oil-body proteins were obtained by PCR cloning. MALDI-MS analyses confirmed that the three full-length cDNA fragments encoded the two oleosin isoforms and one caleosin observed in the oil bodies isolated from the adlay grains.

Stable oil bodies of smaller sizes and higher thermostability were isolated from mature cycad (Cycas revoluta) megagametophytes compared with those isolated from sesame seeds. Immunological cross-recognition revealed that cycad oil bodies contained a major protein of 27 kDa, tentatively identified as caleosin, while oleosin, the well-known structural protein, was apparently absent. Mass spectrometric analysis showed that the putative cycad caleosin possessed a tryptic fragment of 15 residues matching to that of a theoretical moss caleosin. A complete cDNA fragment encoding this putative caleosin was obtained by PCR cloning using a primer designed according to the tryptic peptide and another one designed according to a highly conservative region among diverse caleosins. The identification of this clone was subsequently confirmed by immunodetection and MALDI-MS analyses of its recombinant fusion protein over-expressed in Escherichia coli and the native form from cycad oil bodies. Stable artificial oil bodies were successfully constituted with triacylglycerol, phospholipid and the recombinant fusion protein containing the cycad caleosin. These results suggest that stable oil bodies in cycad megagametophytes are mainly sheltered by a unique structural protein caleosin.

In view of the recent isolation of stable oil bodies as well as a unique oleosin from lily pollen, this study examined whether other minor proteins were present in this lipid-storage organelle. Immunological cross-recognition using antibodies against three minor oil-body proteins from sesame suggested that a putative caleosin was specifically detected in the oil-body fraction of pollen extract. A cDNA fragment encoding this putative pollen caleosin, obtained by PCR cloning, was confirmed by immunodetection and MALDI-MS analyses of the recombinant protein over-expressed in Escherichia coli and the native form. Caleosin in lily pollen oil bodies seemed to be a unique isoform distinct from that in lily seed oil bodies.

Thin-layer chromatography analysis revealed that the contents stored in oil bodies isolated from jelly fig (Ficus awkeotsang Makino) achenes were mainly neutral lipids (>90% triacylglycerols and approximately 5% diacylglycerols). Fatty acids released from the neutral lipids of achene oil bodies were highly unsaturated (62.65% alpha-linolenic acid, 18.24% linoleic acid, and 10.62% oleic acid). The integrity of isolated oil bodies was presumably maintained via electronegative repulsion and steric hindrance provided by their surface proteins. Immunological cross-recognition using antibodies against sesame oil-body proteins indicated that two oleosin isoforms and one caleosin were present in these oil bodies. MALDI-MS analyses confirmed that the three full-length cDNA fragments obtained by PCR cloning from maturing achenes encoded the two jelly fig oleosin isoforms and one caleosin identified by immunological screening.

Stable oil bodies were purified from mature lily (Lilium longiflorum Thunb.) pollen. The integrity of pollen oil bodies was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Immunodetection revealed that a major protein of 18 kDa was exclusively present in pollen oil bodies and massively accumulated in late stages of pollen maturation. According to mass spectrometric analyses, this oil body protein possessed a tryptic fragment of 13 residues matching that of a theoretical rice oleosin. A complete cDNA fragment encoding this putative oleosin was obtained by PCR cloning with primers derived from its known 13-residue sequence. Sequence analysis as well as immunological non-cross-reactivity suggests that this pollen oleosin represents a distinct class in comparison with oleosins found in seed oil bodies and tapetum. In pollen cells observed by electron microscopy, oil bodies were presumably surrounded by tubular membrane structures, and encapsulated in the vacuoles after germination. It seems that pollen oil bodies are mobilized via a different route from that of glyoxysomal mobilization of seed oil bodies after germination.

Objective: To study the safety, immunogenicity on the enterotoxige Escherichia coli (E. coli) recombinant active vaccine FE3 and FE16., Methods: Toxicity and immunogenicity of the vaccine were determined by experiments on enterotoxigenic E. coli toxicity and immunological experiments on rabbits and mice., Results: The results of an toxicological experiments were negative. The agglutination titer of antibodies against the S. flexneri 2a and enterotoxigenic E. coli plamid antigen were all higher than 1:640 and 1:1280 in the sera of rabbits. IgG in the serum went up remarkably, while sIgA against CFA/I was also decteted in the dejecta of mice immunized with active bacteria either orogastrically or intranasally. Simultaneously, sIgA was not detected in the dejecta of mice immunized with inactive bacteria either orogastrically or intranasally., Conclusion: The enterotoxigenic E. coli recombinant active vaccine showed good safety and immunogenicity, inducing both humoral and mucosal immunity in mice.

Objective: To study the correlation between DNA fingerprinting of Mycobacterium tuberculosis (MTB) stains isolated from the Chinese army in the south and from local residents, and to investigate the molecular epidemiological characteristics of tuberculosis (TB) in the army, for the sake of TB prevention in the army., Methods: MTB DNA was digested with restriction endonuclease PvuII and electrophoresed in agarose gel, after Southern Blotting, the membrane was hybridized with a 245 bp fragment of IS6110 which labeled [alpha(32)P]-dCTP as probe. Finally, a restriction fragment length polymorphism (RFLP) patterns was shown, and analyzed logestic with epidemiological data from the patients., Results: A total number of 185 TB strains were detected and the IS6110 copy numbers ranged from 1 - 22. No significant difference was found in the IS6110 copy numbers between patients from army and local patients. IS6110 copy numbers of TB strains in army patients were centered in 6 - 20, however, with 7 - 20 copies in local TB patients. The TB strains were dispersed into 8 groups and the majority of TB strains in both army and local patients was centered in groups I, II, III. The distribution of DNA fingerprint for drug resistance TB strains was significantly different from those for sensitive strains. No different distribution of among groups was found regarding BCG history., Conclusions: The genetics of TB stains were roughly the same between the army patients and local ones, but there was a strong correlation in the gene levels. Data suggested that a close connection should be considered on TB prevention and treatment for TB patients in the army and local residents.

Objective: Typing of Mycobacterium tuberculosis strains and epidemiological studies in the army of southern China to provide scientific basis for prevention of pulmonary tuberculosis., Methods: A rapid fingerprinting of M. tuberculosis strains method by polymerase chain reaction (PCR) with outward-directed primers that designed to the ends of the insertion sequence IS6110 was developed, and to analyze the relationship between the polymorphism of DNA fingerprinting and epidemiology of M. tuberculosis., Results: One hundred and fifty-four M. tuberculosis detected were classified into eight types according to their characters of PCR amplified fingerprints. The main types were type I (36.4%), type II (31.8%), and type III (21.4%), while other types were less than 4 percentage. In those main type groups, patients aged 20 to 29 and 30 to 39 took up 31.8% and 27.9% respectively. For those main types, the distribution of those types in the first treated patients showed significant difference compared with that in the retreated patients, and the rate of drug-resistance was also statistically different. However, the distribution was not statistically significant to history of BCG vaccination and patients living in urban or rural area. The main drug-resistant strains were only Isoniazid-resistant or Rifampin-resistant strains, while the drug-resistant strains were 44.4%, 29.6% and 14.8% respectively in type I, type II and type III., Conclusion: PCR fingerprinting was a rapid, precise, sensitive, specific method to type M. tuberculosis, and could be used to study the epidemiology of tuberculosis; The prevalence of tuberculosis was primarily due to the transmission of type I, type II and type III in the army being studied from Southern China, to suggest that surveillance needs to be strengthened.

A comparison was made of two series of consecutive outpatients with a presumptive diagnosis of rheumatoid arthritis (RA) attending referral centres in Melbourne and Shanghai. No significant differences were observed in disease onset, course, presence of antinuclear antibodies (ANA), or seropositivity. In the Australian series there was a higher frequency of nodules, Raynaud's phenomenon, carpal tunnel syndrome, and 'classical' in comparison with 'definite' disease, and a lower frequency of lymphadenopathy and hepatomegaly. Joint tenderness and soft tissue swelling tended to be more marked in the Chinese series, while deformity and limited range of movement were less severe. Drug therapy was similar overall but influenced by drug availability. Peptic ulceration was recorded in 28% of the Australian series but in only 6% of the Chinese; although 25% of the Chinese were receiving antacids and 6% antiulcerants. X-rays of hands and feet showed more severe disease in the Australian series. The older age group and longer duration of the disease in the Australian patients, who had more chronic and less active disease, may have influenced some of these results.

To test the hypothesis that rheumatoid arthritis in patients in China is milder in severity than that in patients in a western country, a comparison was made of 2 series of consecutive outpatients with definite or classic rheumatoid arthritis who were attending referral centers in Melbourne, Australia and Shanghai, China. This paper reports the findings on radiographs of the wrists and hands. Making allowances for age and duration of disease, changes were more frequent and severe in the Australian patient series, particularly at the metacarpophalangeal joints.