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101.
Wei Feng Jian Wang Xin Yan Qianqian Zhang Limin Chai Qingting Wang Wenhua Shi Yuqian Chen Jin Liu Zhan Qu Shaojun Li Xinming Xie Manxiang Li 《Cell proliferation》2021,54(6)
ObjectivesHigh‐mobility group box‐1 (HMGB1) and aberrant mitochondrial fission mediated by excessive activation of GTPase dynamin‐related protein 1 (Drp1) have been found to be elevated in patients with pulmonary arterial hypertension (PAH) and critically implicated in PAH pathogenesis. However, it remains unknown whether Drp1‐mediated mitochondrial fission and which downstream targets of mitochondrial fission mediate HMGB1‐induced pulmonary arterial smooth muscle cells (PASMCs) proliferation and migration leading to vascular remodelling in PAH. This study aims to address these issues.MethodsPrimary cultured PASMCs were obtained from male Sprague‐Dawley (SD) rats. We detected RNA levels by qRT‐PCR, protein levels by Western blotting, cell proliferation by Cell Counting Kit‐8 (CCK‐8) and EdU incorporation assays, migration by wound healing and transwell assays. SD rats were injected with monocrotaline (MCT) to establish PAH. Hemodynamic parameters were measured by closed‐chest right heart catheterization.ResultsHMGB1 increased Drp1 phosphorylation and Drp1‐dependent mitochondrial fragmentation through extracellular signal‐regulated kinases 1/2 (ERK1/2) signalling activation, and subsequently triggered autophagy activation, which further led to bone morphogenetic protein receptor 2 (BMPR2) lysosomal degradation and inhibitor of DNA binding 1 (Id1) downregulation, and eventually promoted PASMCs proliferation/migration. Inhibition of ERK1/2 cascade, knockdown of Drp1 or suppression of autophagy restored HMGB1‐induced reductions of BMPR2 and Id1, and diminished HMGB1‐induced PASMCs proliferation/migration. In addition, pharmacological inhibition of HMGB1 by glycyrrhizin, suppression of mitochondrial fission by Mdivi‐1 or blockage of autophagy by chloroquine prevented PAH development in MCT‐induced rats PAH model.ConclusionsHMGB1 promotes PASMCs proliferation/migration and pulmonary vascular remodelling by activating ERK1/2/Drp1/Autophagy/BMPR2/Id1 axis, suggesting that this cascade might be a potential novel target for management of PAH. 相似文献
102.
The diameter, membrane thickness, and compression intensity of hollow Ca-alginate capsules were measured at different gelation conditions, such as the reactant concentration, dropping velocity, and gelation time. The optimum operation conditions for preparing capsules were determined at 100 g/L CaCl(2), 10 g/L sodium alginate (Na-alginate), a dropping velocity of 150 droplets/min, and a gelation time of 10 min. Diffusion of some saccharide and amino acid from bulk solution into capsules was investigated, and the diffusion coefficients were calculated by the developed mathematical model. All the tested substances can diffuse easily into the capsules. The combined diffusion coefficients of the capsule D(m) are 92-99% as large as their diffusion coefficients in pure water, while the diffusion coefficients in the capsule membrane D(1) are 60-95% as large as those. By employing polyethylene glycol (PEG) and bovine serum albumin (fraction V) (BSA(V)), the molecular weight cut-off of the capsule was determined. For linear macromolecules, hollow Ca-alginate capsules have a molecular weight cut-off of 4000. No diffusion of BSA(V) into the capsules was observed. 相似文献
103.
The tegumental ultrastructure of juvenile and adult Echinostoma cinetorchis (Trematoda: Echinostomatidae) was observed by scanning electron microscopy. Three-day (juvenile) and 16-day (adult) worms were harvested from rats (Sprague-Dawley) experimentally fed the metacercariae from the laboratory-infected fresh water snail, Hippeutis cantori. The worms were fixed with 2.5% glutaraldehyde, processed routinely, and observed by an ISI Korea DS-130 scanning electron microscope. The 3-day old juvenile worms were elongated and ventrally curved, with their ventral sucker near the anterior two-fifths of the body. The head crown was bearing 37-38 collar spines arranged in a zigzag pattern. The lips of the oral and ventral suckers had 8 and 5 type II sensory papillae respectively, and between the spines, a few type III papillae were observed. Tongue or spade-shape spines were distributed anteriorly to the ventral sucker, whereas peg-like spines were distributed posteriorly and became sparse toward the posterior body. The spines of the dorsal surface were similar to those of the ventral surface. The 16-day old adults were leaf-like, and their oral and ventral suckers were located very closely. Aspinous head crown, oral and ventral suckers had type II and type III sensory papillae, and numerous type I papillae were distributed on the tegument anterior to the ventral sucker. Scale-like spines, with broad base and round tip, were distributed densely on the tegument anterior to the ventral sucker but they became sparse posteriorly. At the dorsal surface, spines were observed at times only at the anterior body. The results showed that the tegument of E. cinetorchis is similar to that of other echinostomes, but differs in the number and arrangement of collar spines, shape and distribution of tegumenal spines, and type and distribution of sensory papillae. 相似文献
104.
R A Childs J R Wright G F Ross C T Yuen A M Lawson W Chai K Drickamer T Feizi 《The Journal of biological chemistry》1992,267(14):9972-9979
Binding specificity of the major surfactant protein SP-A from human and dog lung has been investigated. Radiobinding experiments have shown that both proteins bind in a Ca(2+)-dependent manner to galactose, mannose, fucose, and glucose linked to bovine serum albumin. These results are in accord with a previous study in which monosaccharides were linked to agarose (Haagsman, H. P., Hawgood, S., Sargeant, T., Buckley, D., White, R. T., Drickamer, K., and Benson, B. J. (1987) J. Biol. Chem. 262, 13877-13880). Chromatogram overlays in conjunction with in situ liquid secondary ion mass spectrometry (TLC-LSIMS) of several purified glycosphingolipids and neoglycolipids as well as binding assays with glycolipids immobilized on plastic wells, demonstrate recognition of galactose (human and dog SP-A), glucose, and lactose (human SP-A) in association with specific lipids. In addition, the occurrence of several neutral and acidic glycosphingolipids in human and rat extracellular surfactants and rat alveolar type II cells is described. Selected components among the neutral glycolipids are bound by radiolabeled human SP-A; these are identified by TLC-LSIMS as predominantly ceramide mono- and disaccharides (human surfactant) and ceramide tri- and tetrasaccharides (rat surfactant and type II cells). A recombinant carbohydrate recognition domain (CRD) of human SP-A inhibits the binding of human SP-A to galactosyl ceramide and to galactose- and mannose-bovine serum albumin, indicating that the CRD is directly involved in the binding of SP-A to these ligands. These results provide evidence for a novel type of binding specificity for proteins that have Ca(2+)-dependent CRDs and raise the possibility that glycosphingolipids are endogenous ligands for SP-A. 相似文献
105.
Summary The electrochemical effect of a charged dextran derivative and the hydrophobic effect of hydrophobic chain PEG derivative on partitioning of six types of proteins in PEG/dextran aqueous two-phase systems were investigated- When 1. 6%(w/w)DEAE-dextran was present in the system,the partition coefficient decreased quickly with increasing pH value;when 0. 4% (w/w)PEG pentadecanoic acid ester was present in the system, the partition coefficient of protein with strong hydrophobicity was greatly increased. The experimental results show that the influence of hydrocarbon chain PEG derivative on partition coefficient is closely related to the hydrophobicity of proteins. 相似文献
106.
Kim SK Shahid S Kim SH Park JH Lee HT Jung KH Chai YG 《Letters in applied microbiology》2012,54(4):306-312
Aims: For the analysis of virulence factors produced and secreted by Bacillus anthracis vegetative cells during mammalian host infection, we evaluated the secretome of B. anthracis Sterne exposed to host‐specific factors specifically to host body temperature. Methods and Results: We employed a comparative proteomics‐based approach to analyse the proteins secreted by B. anthracis Sterne under host‐specific body temperature conditions. A total of 17 proteins encoded on a single chromosome and the pXO1 plasmid were identified by peptide mass fingerprinting. Multiple algorithms were used to predict the secretion mechanisms of the detected proteins in B. anthracis. Conclusions: Several putative virulence factors and known factors responsible for sporulation were differentially regulated, including CodY, pXO1‐130 and BA1952, revealing insights into temperature cues in the B. anthracis secretome. Significance and Impact of the Study: This study identified temperature‐regulated proteins. Further studies aimed at understanding the physical and functional roles of these proteins in infection and control by elevated temperatures will contribute to detection, diagnostics and prophylaxis. 相似文献
107.
Jong-Yil Chai Woon-Mok Sohn Byoung-Kuk Na Tai-Soon Yong Keeseon S. Eom Cheong-Ha Yoon Eui-Hyug Hoang Hoo-Gn Jeoung Duong Socheat 《The Korean journal of parasitology》2014,52(1):35-40
A survey was performed to investigate the infection status of freshwater fish with zoonotic trematode metacercariae in Phnom Penh and Pursat Province, Cambodia. All collected fish with ice were transferred to our laboratory and examined using the artificial digestion method. In fish from Phnom Penh, 2 kinds of metacercariae (Opisthorchis viverrini and Haplorchis yokogawai) were detected. O. viverrini metacercariae were positive in 37 (50.0%) of 74 fish in 11 species (average no. metacercariae/fish, 18.6). H. yokogawai metacercariae were detected in 23 (57.5%) of 40 fish in 5 species (average no. metacercariae/fish, 21.0). In fish from Pursat Province, 5 kinds of metacercariae (O. viverrini, H. yokogawai, Haplorchis pumilio, Centrocestus formosanus, and Procerovum sp.) were detected; O. viverrini metacercariae (n=3) in 2 fish species (Henicorhynchus lineatus and Puntioplites falcifer), H. yokogawai metacercariae (n=51) in 1 species (P. falcifer), H. pumilio metacercariae (n=476) in 2 species (H. lineatus and Pristolepis fasciata), C. formosanus metacercariae (n=1) in 1 species (H. lineatus), and Procerovum sp. metacercariae (n=63) in 1 species (Anabas testudineus). From the above results, it has been confirmed that various freshwater fish play the role of a second intermediate host for zoonotic trematodes (O. viverrini, H. yokogawai, H. pumilio, C. formosanus, and Procerovum sp.) in Cambodia. 相似文献
108.
Li Yu Dachuan Shi Junling Li Yingzhen Kong Yanchong Yu Guohua Chai Ruibo Hu Juan Wang Michael G. Hahn Gongke Zhou 《Plant physiology》2014,164(4):1842-1856
Mannans are hemicellulosic polysaccharides that are considered to have both
structural and storage functions in the plant cell wall. However, it is not yet known
how mannans function in Arabidopsis (Arabidopsis thaliana) seed
mucilage. In this study, CELLULOSE SYNTHASE-LIKE A2
(CSLA2; At5g22740) expression was observed in several seed
tissues, including the epidermal cells of developing seed coats. Disruption of
CSLA2 resulted in thinner adherent mucilage halos, although the
total amount of the adherent mucilage did not change compared with the wild type.
This suggested that the adherent mucilage in the mutant was more compact compared
with that of the wild type. In accordance with the role of CSLA2 in glucomannan
synthesis, csla2-1 mucilage contained 30% less mannosyl and glucosyl
content than did the wild type. No appreciable changes in the composition, structure,
or macromolecular properties were observed for nonmannan polysaccharides in mutant
mucilage. Biochemical analysis revealed that cellulose crystallinity was
substantially reduced in csla2-1 mucilage; this was supported by the
removal of most mucilage cellulose through treatment of csla2-1
seeds with endo-β-glucanase. Mutation in CSLA2 also resulted
in altered spatial distribution of cellulose and an absence of birefringent cellulose
microfibrils within the adherent mucilage. As with the observed changes in
crystalline cellulose, the spatial distribution of pectin was also modified in
csla2-1 mucilage. Taken together, our results demonstrate that
glucomannans synthesized by CSLA2 are involved in modulating the structure of
adherent mucilage, potentially through altering cellulose organization and
crystallization.Mannan polysaccharides are a complex set of hemicellulosic cell wall polymers that are
considered to have both structural and storage functions. Based on the particular chemical
composition of the backbone and the side chains, mannan polysaccharides are classified into
four types: pure mannan, glucomannan, galactomannan, and galactoglucomannan (Moreira and Filho, 2008; Wang et al., 2012; Pauly et al.,
2013). Each of these polysaccharides is composed of a β-1,4-linked
backbone containing Man or a combination of Glc and Man residues. In addition, the mannan
backbone can be substituted with side chains of α-1,6-linked Gal residues. Mannan
polysaccharides have been proposed to cross link with cellulose and other hemicelluloses
via hydrogen bonds (Fry, 1986; Iiyama et al., 1994; Obel et al., 2007; Scheller and
Ulvskov, 2010). Furthermore, it has been reported that heteromannans with
different levels of substitution can interact with cellulose in diverse ways (Whitney et al., 1998). Together, these observations
indicate the complexity of mannan polysaccharides in the context of cell wall
architecture.CELLULOSE SYNTHASE-LIKE A (CSLA) enzymes have been shown to have mannan synthase activity
in vitro. These enzymes polymerize the β-1,4-linked backbone of mannans or
glucomannans, depending on the substrates (GDP-Man and/or GDP-Glc) provided (Richmond and Somerville, 2000; Liepman et al., 2005, 2007;
Pauly et al., 2013). In Arabidopsis
(Arabidopsis thaliana), nine CSLA genes have been
identified; different CSLAs are responsible for the synthesis of different
mannan types (Liepman et al., 2005, 2007). CSLA7 has mannan synthase activity in vitro
(Liepman et al., 2005) and has been shown to
synthesize stem glucomannan in vivo (Goubet et al.,
2009). Disrupting the CSLA7 gene results in defective pollen
growth and embryo lethality phenotypes in Arabidopsis, indicating structural or signaling
functions of mannan polysaccharides during plant embryo development (Goubet et al., 2003). A mutation in CSLA9 results in
the inhibition of Agrobacterium tumefaciens-mediated root transformation
in the rat4 mutant (Zhu et al.,
2003). CSLA2, CSLA3, and CSLA9 are proposed to play nonredundant roles in the
biosynthesis of stem glucomannans, although mutations in CSLA2,
CSLA3, or CSLA9 have no effect on stem development or
strength (Goubet et al., 2009). All of the
Arabidopsis CSLA proteins have been shown to be involved in the biosynthesis of mannan
polysaccharides in the plant cell wall (Liepman et al.,
2005, 2007), although the precise
physiological functions of only CSLA7 and CSLA9 have been conclusively demonstrated.In Arabidopsis, when mature dry seeds are hydrated, gel-like mucilage is extruded to
envelop the entire seed. Ruthenium red staining of Arabidopsis seeds reveals two different
mucilage layers, termed the nonadherent and the adherent mucilage layers (Western et al., 2000; Macquet et al., 2007a). The outer, nonadherent mucilage is loosely
attached and can be easily extracted by shaking seeds in water. Compositional and linkage
analyses suggest that this layer is almost exclusively composed of unbranched
rhamnogalacturonan I (RG-I) (>80% to 90%), with
small amounts of branched RG-I, arabinoxylan, and
high methylesterified homogalacturonan (HG). By
contrast, the inner, adherent mucilage layer is tightly attached to the seed and can only
be removed by strong acid or base treatment, or by enzymatic digestion (Macquet et al., 2007a; Huang et al., 2011; Walker et al.,
2011). As with the nonadherent layer, adherent mucilage is also mainly composed
of unbranched RG-I, but with small numbers of
arabinan and galactan ramifications (Penfield et al.,
2001; Willats et al., 2001; Dean et al., 2007; Macquet et al., 2007a, 2007b; Arsovski et al., 2009; Haughn and Western, 2012). There are also minor amounts of pectic
HG in the adherent mucilage, with high
methylesterified HG in the external domain compared
with the internal domain of the adherent layer (Willats
et al., 2001; Macquet et al., 2007a;
Rautengarten et al., 2008; Sullivan et al., 2011; Saez-Aguayo et al., 2013). In addition, the adherent mucilage
contains cellulose (Blake et al., 2006; Macquet et al., 2007a), which is entangled with RG-I and is thought to anchor the pectin-rich mucilage
onto seeds (Macquet et al., 2007a; Harpaz-Saad et al., 2011, 2012; Mendu et al., 2011;
Sullivan et al., 2011). As such, Arabidopsis
seed mucilage is considered to be a useful model for investigating the biosynthesis of cell
wall polysaccharides and how this process is regulated in vivo (Haughn and Western, 2012).Screening for altered seed coat mucilage has led to the identification of several genes
encoding enzymes that are involved in the biosynthesis or modification of mucilage
components. RHAMNOSE SYNTHASE2/MUCILAGE-MODIFIED4 (MUM4) is responsible for the synthesis
of UDP-l-Rha (Usadel et al., 2004; Western et al., 2004; Oka et al., 2007). The putative GALACTURONSYLTRANSFERASE11 can
potentially synthesize mucilage RG-I or HG pectin from UDP-d-GalUA (Caffall et al., 2009). GALACTURONSYLTRANSFERASE-LIKE5
appears to function in the regulation of the final size of the mucilage RG-I (Kong et al.,
2011, 2013). Mutant seeds defective in
these genes display reduced thickness of the extruded mucilage layer compared with
wild-type Arabidopsis seeds.RG-I deposited in the apoplast of seed coat
epidermal cells appears to be synthesized in a branched form that is subsequently modified
by enzymes in the apoplast. MUM2 encodes a β-galactosidase that
removes Gal residues from RG-I side chains (Dean et al., 2007; Macquet et al., 2007b). β-XYLOSIDASE1 encodes an
α-l-arabinfuranosidase that removes Ara residues from RG-I side chains (Arsovski et al., 2009). Disruptions of these genes lead to defective hydration
properties and affect the extrusion of mucilage. Furthermore, correct methylesterification
of mucilage HG is also required for mucilage
extrusion. HG is secreted into the wall in a high
methylesterified form that can then be enzymatically demethylesterified by pectin
methylesterases (PMEs; Bosch and Hepler, 2005). PECTIN METHYLESTERASE INHIBITOR6 (PMEI6)
inhibits PME activities (Saez-Aguayo et al., 2013). The subtilisin-like Ser protease (SBT1.7)
can activate other PME inhibitors, but not PMEI6
(Rautengarten et al., 2008; Saez-Aguayo et al., 2013). Disruption of either
PMEI6 or SBT1.7 results in the delay of mucilage
release.Although cellulose is present at low levels in adherent mucilage, it plays an important
adhesive role for the attachment of mucilage pectin to the seed coat epidermal cells. The
orientation and amount of pectin associated with the cellulose network is largely
determined by cellulose conformation properties (Macquet
et al., 2007a; Haughn and Western,
2012). Previous studies have demonstrated that CELLULOSE SYNTHASE A5 (CESA5) is
required for the production of seed mucilage cellulose and the adherent mucilage in the
cesa5 mutant can be easily extracted with water (Harpaz-Saad et al., 2011, 2012; Mendu et al., 2011; Sullivan et al., 2011).Despite all of these discoveries, large gaps remain in the current knowledge of the
biosynthesis and functions of mucilage polysaccharides in seed coats. In this study, we
show that CSLA2 is involved in the biosynthesis of mucilage glucomannan. Furthermore, we
show that CSLA2 functions in the maintenance of the normal structure of the adherent
mucilage layer through modifying the mucilage cellulose ultrastructure. 相似文献
109.
Mario Gimona Maria Felice Brizzi Andre Boon Hwa Choo Massimo Dominici Sean M. Davidson Johannes Grillari Dirk M. Hermann Andrew F. Hill Dominique de Kleijn Ruenn Chai Lai Charles P. Lai Rebecca Lim Marta Monguió-Tortajada Maurizio Muraca Takahiro Ochiya Luis A. Ortiz Wei Seong Toh Yong Weon Yi Sai Kiang Lim 《Cytotherapy》2021,23(5):373-380
Mesenchymal stromal/stem cells (MSCs) have been widely tested against many diseases, with more than 1000 registered clinical trials worldwide. Despite many setbacks, MSCs have been approved for the treatment of graft-versus-host disease and Crohn disease. However, it is increasingly clear that MSCs exert their therapeutic functions in a paracrine manner through the secretion of small extracellular vesicles (sEVs) of 50–200 nm in diameter. Unlike living cells that can persist long-term, sEVs are non-living and non-replicative and have a transient presence in the body. Their small size also renders sEV preparations highly amenable to sterilization by filtration. Together, acellular MSC-sEV preparations are potentially safer and easier to translate into the clinic than cellular MSC products. Nevertheless, there are inherent challenges in the development of MSC-sEV drug products. MSC-sEVs are products of living cells, and living cells are sensitive to changes in the external microenvironment. Consequently, quality control metrics to measure key identity and potency features of MSC-sEV preparations have to be specified during development of MSC-sEV therapeutics. The authors have previously described quantifiable assays to define the identity of MSC-sEVs. Here the authors discuss requirements for prospective potency assays to predict the therapeutic effectiveness of the drug substance in accordance with International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use guidelines. Although potency assays should ideally reflect the mechanism of action (MoA), this is challenging because the MoA for the reported efficacy of MSC-sEV preparations against multiple diseases of diverse underlying pathology is likely to be complex and different for each disease and difficult to fully elucidate. Nevertheless, robust potency assays could be developed by identifying the EV attribute most relevant to the intended biological activity in EV-mediated therapy and quantifying the EV attribute. Specifically, the authors highlight challenges and mitigation measures to enhance the manufacture of consistent and reproducibly potent sEV preparations, to identify and select the appropriate EV attribute for potency assays despite a complex “work-in-progress” MoA and to develop assays likely to be compliant with regulatory guidance for assay validation. 相似文献
110.