Publications by authors named "Jingxiu Bi"

52 Publications

Engineered Human Heavy-Chain Ferritin with Half-Life Extension and Tumor Targeting by PAS and RGDK Peptide Functionalization.

Pharmaceutics 2021 Apr 9;13(4). Epub 2021 Apr 9.

School of Chemical Engineering and Advanced Materials, The University of Adelaide, Adelaide SA5005, Australia.

Ferritin, one of the most investigated protein nanocages, is considered as a promising drug carrier because of its advantageous stability and safety. However, its short half-life and undesirable tumor targeting ability has limited its usage in tumor treatment. In this work, two types of functional peptides, half-life extension peptide PAS, and tumor targeting peptide RGDK (Arg-Gly-Asp-Lys), are inserted to human heavy-chain ferritin (HFn) at C-terminal through flexible linkers with two distinct enzyme cleavable sites. Structural characterizations show both HFn and engineered HFns can assemble into nanoparticles but with different apparent hydrodynamic volumes and molecular weights. RGDK peptide enhanced the internalization efficiency of HFn and showed a significant increase of growth inhibition against 4T1 cell line in vitro. Pharmacokinetic study in vivo demonstrates PAS peptides extended ferritin half-life about 4.9 times in Sprague Dawley rats. RGDK peptides greatly enhanced drug accumulation in the tumor site rather than in other organs in biodistribution analysis. Drug loaded PAS-RGDK functionalized HFns curbed tumor growth with significantly greater efficacies in comparison with drug loaded HFn.
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http://dx.doi.org/10.3390/pharmaceutics13040521DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8070472PMC
April 2021

Stability of Engineered Ferritin Nanovaccines Investigated by Combined Molecular Simulation and Experiments.

J Phys Chem B 2021 04 7;125(15):3830-3842. Epub 2021 Apr 7.

School of Chemical Engineering and Advanced Materials, The University of Adelaide, Adelaide, South Australia 5005, Australia.

Human ferritin is regarded as an attractive and promising vaccine platform because of its uniform structure, good plasticity, and desirable thermal and chemical stabilities. Besides, it is biocompatible and presumed safe when used as a vaccine carrier. However, there is a lack of knowledge of how different antigen insertion sites on the ferritin nanocage impact the resulting protein stability and performance. To address this question, we selected Epstein-Barr nuclear antigen 1 as a model epitope and fused it at the DNA level with different insertion sites, namely, the N- and C-termini of ferritin, to engineer proteins E1F1 and F1E1, respectively. Protein properties including hydrophobicity and thermal, pH, and chemical stability were investigated both by molecular dynamics (MD) simulation and by experiments. Both methods demonstrate that the insertion site plays an important role in protein properties. The C-terminus insertion (F1E1) leads to a less hydrophobic surface and more tolerance to the external influence of high temperature, pH, and high concentration of chemical denaturants compared to N-terminus insertion (E1F1). Simulated protein hydrophobicity and thermal stability by MD were in high accordance with experimental results. Thus, MD simulation can be used as a valuable tool to engineer nanovaccine candidates, cutting down costs by reducing the experimental effort and accelerating vaccine design.
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http://dx.doi.org/10.1021/acs.jpcb.1c00276DOI Listing
April 2021

Comparative evaluation of integrated purification pathways for bacterial modular polyomavirus major capsid protein VP1 to produce virus-like particles using high throughput process technologies.

J Chromatogr A 2021 Feb 21;1639:461924. Epub 2021 Jan 21.

The University of Adelaide, Division of Research and Innovation, Adelaide, SA 5005, Australia. Electronic address:

Modular virus-like particles and capsomeres are potential vaccine candidates that can induce strong immune responses. There are many described protocols for the purification of microbially-produced viral protein in the literature, however, they suffer from inherent limitations in efficiency, scalability and overall process costs. In this study, we investigated alternative purification pathways to identify and optimise a suitable purification pathway to overcome some of the current challenges. Among the methods, the optimised purification strategy consists of an anion exchange step in flow through mode followed by a multi modal cation exchange step in bind and elute mode. This approach allows an integrated process without any buffer adjustment between the purification steps. The major contaminants like host cell proteins, DNA and aggregates can be efficiently removed by the optimised strategy, without the need for a size exclusion polishing chromatography step, which otherwise could complicate the process scalability and increase overall cost. High throughput process technology studies were conducted to optimise binding and elution conditions for multi modal cation exchanger, Capto™ MMC and strong anion exchanger Capto™ Q. A dynamic binding capacity of 14 mg ml was achieved for Capto™ MMC resin. Samples derived from each purification process were thoroughly characterized by RP-HPLC, SEC-HPLC, SDS-PAGE and LC-ESI-MS/MS Mass Spectrometry analytical methods. Modular polyomavirus major capsid protein could be purified within hours using the optimised process achieving purities above 87% and above 96% with inclusion of an initial precipitation step. Purified capsid protein could be easily assembled in-vitro into well-defined virus-like particles by lowering pH with addition of calcium chloride to the eluate. High throughout studies allowed the screening of a vast design space within weeks, rather than months, and unveiled complicated binding behaviour for Capto MMC.
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http://dx.doi.org/10.1016/j.chroma.2021.461924DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7825977PMC
February 2021

Virus-like particle preparation is improved by control over capsomere-DNA interactions during chromatographic purification.

Biotechnol Bioeng 2021 Apr 19;118(4):1707-1720. Epub 2021 Feb 19.

Division of Research and Innovation, The University of Adelaide, Adelaide, South Australia, Australia.

Expression of viral capsomeres in bacterial systems and subsequent in vitro assembly into virus-like particles is a possible pathway for affordable future vaccines. However, purification is challenging as viral capsomeres show poor binding to chromatography media. In this study, the behavior of capsomeres in unfractionated bacterial lysate was compared with that for purified capsomeres, with or without added microbial DNA, to better understand reasons for poor bioprocess behavior. We show that aggregates or complexes form through the interaction between viral capsomeres and DNA, especially in bacterial lysates rich in contaminating DNA. The formation of these complexes prevents the target protein capsomeres from accessing the pores of chromatography media. We find that protein-DNA interactions can be modulated by controlling the ionic strength of the buffer and that at elevated ionic strengths the protein-DNA complexes dissociate. Capsomeres thus released show enhanced bind-elute behavior on salt-tolerant chromatography media. DNA could therefore be efficiently removed. We believe this is the first report of the use of an optimized salt concentration that dissociates capsomere-DNA complexes yet enables binding to salt-tolerant media. Post purification, assembly experiments indicate that DNA-protein interactions can play a negative role during in vitro assembly, as DNA-protein complexes could not be assembled into virus-like particles, but formed worm-like structures. This study reveals that the control over DNA-protein interaction is a critical consideration during downstream process development for viral vaccines.
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http://dx.doi.org/10.1002/bit.27687DOI Listing
April 2021

In vitro preparation of uniform and nucleic acid free hepatitis B core particles through an optimized disassembly-purification-reassembly process.

Protein Expr Purif 2021 02 6;178:105747. Epub 2020 Sep 6.

National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, 100190, PR China. Electronic address:

Structure heterogeneity and host nucleic acids contamination are two major problems for virus-like particles (VLPs) produced by various host cells. In this study, an in vitro optimized disassembly-purification-reassembly process was developed to obtain uniform and nucleic acid free hepatitis B core (HBc) based VLPs from E. coli fermentation. The process started with ammonium sulfate precipitation of all heterogeneous HBc structures after cell disintegration. Then, dissolution and disassembly of pellets into basic subunits were carried out under the optimized disassembly condition. All contaminants, including host nucleic acids and proteins, were efficiently removed with affinity chromatography. The purified subunits reassembled into VLPs by final removal of the chaotropic agent. Two uniform and nucleic acid free HBc-based VLPs, truncated HBc and chimeric HBc-MAGE3 I, were successfully prepared. It was found that disassembly degree of HBc-based VLPs had a great influence on the protein yield, nucleic acid removal and reassembly efficiency. 4 M urea was optimal because lower concentration would not disassemble the particles completely while higher concentration would further denature the subunits into disordered aggregate and could not be purified and reassembled efficiently. For removal of strong binding nucleic acids such as in the case of HBc-MAGE3 I, benzonase nuclease was added to the disassembly buffer before affinity purification. Through the optimized downstream process, uniform and nucleic acid free HBc VLPs and HBc-MAGE3 I VLPs were obtained with purities above 90% and yields of 55.2 and 43.0 mg/L, respectively. This study would be a reference for efficient preparation of other VLPs.
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http://dx.doi.org/10.1016/j.pep.2020.105747DOI Listing
February 2021

Functional characterization of a special dicistronic transcription unit encoding histone methyltransferase su(var)3-9 and translation regulator eIF2γ in Tribolium castaneum.

Biochem J 2020 08;477(16):3059-3074

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210046, China.

Operons are rare in eukaryotes, where they often allow concerted expression of functionally related genes. While a dicistronic transcription unit encoding two unrelated genes, the suppressor of position-effect variegation su(var)3-9 and the gamma subunit of eukaryotic translation initiation factor 2 (eIF2γ) has been found in insecta, and its significance is not well understood. Here, we analyzed the evolutionary history of this transcription unit in arthropods and its functions by using model Coleoptera insect Tribolium castaneum. In T. castaneum, Tcsu(var)3-9 fused into the 80 N-terminal amino acids of TceIF2γ, the transcription of these two genes are resolved by alternative splicing. Phylogenetic analysis supports the natural gene fusion of su(var)3-9 and eIF2γ occurred in the ancestral line of winged insects and silverfish, but with frequent re-fission during the evolution of insects. Functional analysis by using RNAi for these two genes revealed that gene fusion did not invoke novel functions for the gene products. As a histone methyltransferase, Tcsu(var)3-9 is primarily responsible for H3K9 di-, and tri-methylation and plays important roles in metamorphosis and embryogenesis in T. castaneum. While TceIF2γ plays essential roles in T. castaneum by positively regulating protein translation mediated ecdysteroid biosynthesis. The vulnerability of the gene fusion and totally different role of su(var)3-9 and eIF2γ in T. castaneum confirm this gene fusion is a non-selected, constructive neutral evolution event in insect. Moreover, the positive relationship between protein translation and ecdysteroid biosynthesis gives new insights into correlations between translation regulation and hormonal signaling.
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http://dx.doi.org/10.1042/BCJ20200444DOI Listing
August 2020

Functional analysis of a novel orthologous small heat shock protein (shsp) hsp21.8a and seven species-specific shsps in Tribolium castaneum.

Genomics 2020 11 1;112(6):4474-4485. Epub 2020 Aug 1.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China. Electronic address:

Small heat shock proteins (sHSPs) are important modulators of insect survival. Previous research revealed that there is only one orthologous cluster of shsps in insects. Here, we identified another novel orthologous cluster of shsps in insects by comparative analysis. Multiple stress experiments and function investigation of Tchsp21.8a belonging to this orthologous cluster and seven species-specific shsps were performed in the stored-grain pest Tribolium castaneum. The results indicated that expression of Tchsp21.8a showed weak responses to different stresses. However, expressions of most species-specific shsps exhibited hyper-responses to heat stress, and expressions of all species-specific shsps displayed diverse responses during other stresses to protect beetles in a cooperative manner. Additionally, Tchsp21.8a and species-specific Tcshsp19.7 played important roles in the development of T. castaneum, and all Tcshsps had a certain impact on the fecundity. Our work created a comprehensive reliable scaffold of insect shsps that can further provide instructive insights to pest bio-control.
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http://dx.doi.org/10.1016/j.ygeno.2020.07.040DOI Listing
November 2020

3D bioprinting of cell-laden electroconductive MXene nanocomposite bioinks.

Nanoscale 2020 Aug;12(30):16069-16080

School of Chemical Engineering and Advanced Materials, The University of Adelaide, Adelaide, SA 5005, Australia.

MXenes, a new family of burgeoning two-dimensional (2D) transition metal carbides/nitrides, have been extensively explored in recent years owing to their outstanding properties such as a large specific surface area, high electrical conductivity, low toxicity, and biodegradability. Numerous efforts have been devoted to exploring MXenes for various biomedical applications such as cancer therapy, bioimaging, biosensing, and drug delivery. However, the potential application of MXene nanosheets in tissue engineering has been almost overlooked despite their excellent performance in other biomedical applications. The overarching goal of this paper is to demonstrate the potential of MXene cell-laden bioinks for tissue engineering and their ability to assemble functional scaffolds to regenerate damaged tissue via 3D bioprinting. We formulate a new electroconductive cell-laden bioink composed of Ti3C2 MXene nanosheets dispersed homogeneously within hyaluronic acid/alginate (HA/Alg) hydrogels and showed its performance for extrusion-based 3D bioprinting. The prepared hydrogel bioinks with MXenes display excellent rheological properties, which allows the fabrication of multilayered 3D structures with high resolution and shape retention. Moreover, the introduction of Ti3C2 MXene nanosheets within the HA/Alg hydrogel introduces electrical conductivity to the ink, addressing the poor electrical conductivity of the current bioinks that mismatch with the physico-chemical properties of tissue. In addition, the MXene nanocomposite ink with encapsulated Human Embryonic Kidney 293 (HEK-293) cells displayed high cell viability (>95%) in both bulk hydrogel and 3D bioprinted structures. These results suggest that MXene nanocomposite bioinks and their 3D bioprinting with high electrical conductivity, biocompatibility and degradability can synergize some new applications for tissue and neural engineering.
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http://dx.doi.org/10.1039/d0nr02581jDOI Listing
August 2020

3D printing of cell-laden electroconductive bioinks for tissue engineering applications.

J Mater Chem B 2020 07;8(27):5862-5876

School of Chemical Engineering & Advanced Materials, The University of Adelaide, Adelaide, SA 5005, Australia.

Bioprinting is an emerging powerful fabrication method, which enables the rapid assembly of 3D bioconstructs with dispensing cell-laden bioinks in pre-designed locations. However, to translate this technology into real applications, there are still a number of challenges that need to be addressed. First, the current inks are generally composed of polymeric materials with poor electrical conductivity that mismatches with the native tissue environment. The second challenge associated with the 3D bioprinting of hydrogel-based bioinks is the fabrication of anatomical-size constructs without any loss of shape fidelity and resolution. To address these challenges, in this work, we introduced a biocompatible bioink associated with current 3D bioprinting by combining methylcellulose and kappa-carrageenan (MC/κCA) hydrogels with poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) conducting polymers. The prepared ink exhibited highly thixotropic behaviour, which could be tuned via changing the concentration of MC and κCA to obtain easy printing with high shape fidelity. The ink was able to fabricate physiological-scale constructs without requiring a secondary support bath. In addition, varying the concentration of PEDOT:PSS could control the electrical conductivity of the ink. Moreover, the encapsulated human embryonic kidney 293 (HEK-293) cells in bulk hydrogels and 3D bioprinted structures maintained high cell viability (>96%) over a week, confirming the in vitro biocompatibility of the ink. Overall, these findings indicate that the MC/κCA/PEDOT:PSS bioink can be promising in biomedical applications, which improved the electroconductivity of bioinks and can exploit the advantage of conductive polymers in the 3D bioprinting technology.
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http://dx.doi.org/10.1039/d0tb00627kDOI Listing
July 2020

A C-type lectin with dual-CRD from Tribolium castaneum is induced in response to bacterial challenge.

Pest Manag Sci 2020 Dec 25;76(12):3965-3974. Epub 2020 Jun 25.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing, China.

Background: C-type lectins (CTLs), a group of pattern recognition receptors, are involved in regulating the immune response of insects and could be used as potential targets for pest control. However, information about roles of CTLs in the innate immunity of Tribolium castaneum, a serious, worldwide pest that damages stored grain products, is relatively scarce.

Results: Here, a CTL with dual carbohydrate recognition domains (CRDs) containing a highly conserved WHD (Trp -His -Asp ) motif was identified in T. castaneum and named as TcCTL3. Spatiotemporal analysis showed that TcCTL3 was highly expressed in all developmental stages except early eggs, and mainly distributed in central nervous system and hemolymph. The transcript levels of TcCTL3 were significantly increased after lipopolysaccharide (LPS) and peptidoglycan (PGN) stimulation. Recombinant TcCTL3 was able to bind directly to LPS, PGN and all tested bacteria and induce a broad spectrum of microbial agglutination in the presence of Ca . The binding was shown mainly through CRD1 domain of TcCTL3. When TcCTL3 was knocked down by RNA interference, expression of nine antimicrobial peptides (AMPs) (attacin1, attacin2, attacin3, defensins1, defensins2, coleoptericin1, coleoptericin2, cecropins2 and cecropins3) and four transcription factors (TFs) (dif1, dif2, relish and jnk) were significantly decreased under LPS and PGN stimulation, leading to increased mortality of T. castaneum when infected with Gram-positive Staphylococcus aureus or Gram-negative Escherichia coli infection.

Conclusion: TcCTL3 could mediate the immune response in T. castaneum via the pattern recognition, agglutination and AMP expression. These findings indicate a potential mechanism of TcCTL3 in resisting bacteria and provide an alternative molecular target for pest control. © 2020 Society of Chemical Industry.
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http://dx.doi.org/10.1002/ps.5945DOI Listing
December 2020

A typical C-type lectin, perlucin-like protein, is involved in the innate immune defense of whiteleg shrimp Litopenaeus vannamei.

Fish Shellfish Immunol 2020 Aug 19;103:293-301. Epub 2020 May 19.

Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Life Sciences & College of Marine Science and Engineering, Nanjing Normal University, 1 Wenyuan Road, Nanjing, 210046, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu Province, Lianyungang, Jiangsu, 222005, China. Electronic address:

C-type lectins are a large group of the pattern-recognition proteins, and have been reported to be involved in invertebrate innate immunity, such as cell adhesion, bacterial clearance, phagocytosis, prophenoloxidase activation and encapsulation. Here, a perlucin-like protein (PLP), a typical C-type lectin, was identified from the cDNA library of the shrimp, Litopenaeus vannamei. LvPLP contains a 540 bp open reading frame, encoding a protein of 179 amino acids that includes a single carbohydrate-recognition domain. Phylogenetic analysis showed that LvPLP was clustered into a single group together with other perlucins from molluscs. Quantitative real-time PCR revealed that LvPLP was expressed mainly in the hemocytes, hemolymph, heart and gills. The transcription of LvPLP was significantly induced at 9 h by both Gram bacteria Vibrio parahaemolyticus and Vibrio anguillarum. Meanwhile, recombinant LvPLP (rLvPLP) bound directly to lipopolysaccharide and peptidoglycan with different affinity. rLvPLP showed a strong ability to bind to Gram (Staphylococcus aureus and Bacillus subtilis) and Gram bacteria (V. parahaemolyticus and V. anguillarum), and could induce agglutination of V. parahaemolyticus and V. anguillarum, but not S. aureus and B. subtilis in the presence Ca. Further study showed that when LvPLP was knocked down by RNAi, three phagocytosis-related genes (peroxinectin, mas-like protein and dynamin) and four antimicrobial peptide (AMP) genes (crustin, ALF1, ALF2 and ALF3) were significantly decreased. Altogether, these results demonstrated that LvPLP played a vital role in L. vannamei immune response towards bacterial challenge by binding and agglutinating bacteria and influencing phagocytosis and AMP expression.
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http://dx.doi.org/10.1016/j.fsi.2020.05.046DOI Listing
August 2020

Identification and functional characterization of methyl-CpG binding domain protein from Tribolium castaneum.

Genomics 2020 05 26;112(3):2223-2232. Epub 2019 Dec 26.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210046, China. Electronic address:

Methyl-CpG binding domain proteins (MBD) can specifically bind to methylated CpG sites and play important roles in epigenetic gene regulation. Here, we identified and functionally characterized the MBD protein in Tribolium castaneum. T. castaneum genome encodes only one MBD protein: TcMBD2/3. RNA interference targeting this gene at different developmental stages caused lethal phenotypes including metamorphosis deficiency in larvae and pupae, gastrointestinal system problems and fecundity deficiency in adult. Moreover, Tcmbd2/3 knockdown adult showed progressive reduced locomoter activity, a typical neurodegeneration phenotype. This is a common feature of DNA methylation in mammals and has not been found in other insects. However, band shift assays demonstrated that TcMBD2/3 could not bind to methylated DNA, indicating the essential roles of TcMBD2/3 is independent of DNA methylation. Our study provides Tcmbd2/3 plays important roles in T. castaneum and gives new insights into the potential mechanism of action of MBD proteins in insect.
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http://dx.doi.org/10.1016/j.ygeno.2019.12.018DOI Listing
May 2020

Invasion Into Hemocytes Is Mediated by Pathogen Enolase and Host Lipopolysaccharide and β-1, 3-Glucan Binding Protein.

Front Immunol 2019 8;10:1852. Epub 2019 Aug 8.

Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Marine Science and Engineering, Nanjing Normal University, Nanjing, China.

is a crustacean pathogen, without a cell wall, that causes enormous economic loss. hemocytes are the major targets during infection. As wall-less bacteria, , its membrane protein should interact with host membrane protein directly and firstly when invaded in host cell. In this investigation, six potential hemocyte receptor proteins were identified firstly that mediate interaction between and . Among these proteins, lipopolysaccharide and β-1, 3-glucan binding protein (MrLGBP) demonstrated to bind to using bacterial binding assays and confocal microscopy. Four spiroplasma ligand proteins for MrLGBP were isolated and identified. But, competitive assessment demonstrated that only enolase of (SeEnolase) could be a candidate ligand for MrLGBP. Subsequently, the interaction between MrLGBP and SeEnolase was confirmed by co-immunoprecipitation and co-localization . After the interaction between MrLGBP and SeEnolase was inhibited by antibody neutralization test, the virulence ability of was effectively reduced. The quantity of decreased in S2 cells after overexpression of , compared with the controls. In addition, RNA interference (RNAi) knockdown of made more sensitive to infection. Further studies found that the immune genes, including and (), , and α were significantly up-regulated by SeEnolase stimulation. After SeEnolase pre-stimulation, the ability of resistance to was significantly improved. Collectively, this investigation demonstrated that MrLGBP and pathogen SeEnolase involved in mediating invasion into hemocytes.
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http://dx.doi.org/10.3389/fimmu.2019.01852DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6694788PMC
October 2020

Latrophilin participates in insecticide susceptibility through positively regulating CSP10 and partially compensated by OBPC01 in Tribolium castaneum.

Pestic Biochem Physiol 2019 Sep 6;159:107-117. Epub 2019 Jun 6.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China. Electronic address:

Latrophilin (LPH) is an adhesion G protein-coupled receptor (aGPCR) that participates in multiple essential physiological processes. Our previous studies have shown that lph is not only indispensable for the development and reproduction of red flour beetles (Tribolium castaneum), but also for their resistance against dichlorvos or carbofuran insecticides. However, the regulatory mechanism of lph-mediated insecticide susceptibility remains unclear. Here, we revealed that knockdown of lph in beetles resulted in opposing changes in two chemoreception genes, chemosensory protein 10 (CSP10) and odorant-binding protein C01 (OBPC01), in which the expression of TcCSP10 was downregulated, whereas the expression of TcOBPC01 was upregulated. TcCSP10 and TcOBPC01 were expressed at the highest levels in early pupal and late larval stages, respectively. High levels of expression of both these genes were observed in the heads (without antennae) of adults. TcCSP10 and TcOBPC01 were significantly induced by dichlorvos or carbofuran between 12 and 72 h (hrs) after exposure, suggesting that they are likely associated with increasing the binding affinity of insecticides, leading to a decrease in sensitivity to the insecticides. Moreover, once these two genes were knocked down, the susceptibility of the beetles to dichlorvos or carbofuran was enhanced. Additionally, RNA interference (RNAi) targeting of lph followed by exposure to dichlorvos or carbofuran also caused the opposing expression levels of TcCSP10 and TcOBPC01 compared to the expression levels of wild-type larvae treated with insecticides alone. All these results indicate that lph is involved in insecticide susceptibility through positively regulating TcCSP10; and the susceptibility could also further partially compensated for through the negative regulation of TcOBPC01 when lph was knockdown in the red flour beetle. Our studies shed new light on the molecular regulatory mechanisms of lph related to insecticide susceptibility.
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http://dx.doi.org/10.1016/j.pestbp.2019.06.005DOI Listing
September 2019

CYP4BN6 and CYP6BQ11 mediate insecticide susceptibility and their expression is regulated by Latrophilin in Tribolium castaneum.

Pest Manag Sci 2019 Oct 21;75(10):2744-2755. Epub 2019 Mar 21.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing, China.

Background: Many insect cytochrome P450 proteins (CYPs) are involved in the metabolic detoxification of exogenous compounds such as plant toxins and insecticides. Tribolium castaneum, the red flour beetle, is a major agricultural pest that damages stored grains and cereal products. With the completion of the sequencing of its genome, two T. castaneum species-specific CYP genes, CYP4BN6, and CYP6BQ11, were identified. However, it is unknown whether the functions of most CYPs are shared by TcCYP4BN6 and TcCYP6BQ11, and the upstream regulatory mechanism of these two CYPs remains elusive.

Results: QRT-PCR analysis indicated that TcCYP4BN6 and TcCYP6BQ11 were both most highly expressed at the late pupal stage and were mainly observed in the head and gut, respectively, of adults. Moreover, the transcripts of these two CYPs were significantly induced by dichlorvos and carbofuran, and RNA interference (RNAi) targeting of each of them enhanced the susceptibility of beetles to these two insecticides. Intriguingly, knockdown of the latrophilin (lph) gene, which has been reported to be related to the insecticide susceptibility, reduced the expression of TcCYP4BN6 and TcCYP6BQ11 after insecticide treatment, suggesting that these two CYP genes are regulated by lph to participate in insecticide susceptibility in T. castaneum.

Conclusion: These results shed new light on the function and mechanism of CYP genes associated with insecticide susceptibility and could facilitate research on appropriate and sustainable pest control management. © 2019 Society of Chemical Industry.
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http://dx.doi.org/10.1002/ps.5384DOI Listing
October 2019

Tuning microenvironment for multicellular spheroid formation in thermo-responsive anionic microgel scaffolds.

J Biomed Mater Res A 2018 11 26;106(11):2899-2909. Epub 2018 Oct 26.

School of Chemical Engineering, the University of Adelaide, Adelaide, Australia.

Tumor spheroids are considered to be effective in drug screening and evaluation. Three-dimensional scaffold-based cell culture becomes very promising in producing multicellular spheroids. Different from other approaches, 3D scaffolds mimic in vivo cellular microenvironment which encourages intercellular and extracellular interactions. The properties of the cellular microenvironment include the surface wettability, chemistry, and charge of the scaffolds which may influence cell attachment, proliferation as well as migration and these properties are essential for multicellular spheroids formation. Through co-polymerization with different carboxylic acids, we demonstrate that the surface charge density and hydrophobicity of the microenvironment have a great impact on the tumor spheroids formation progress and their size distribution. Our results show that a scaffold with a moderate negative charge density and a highly hydrophilic surface promotes cell proliferation, resulting in quicker and larger spheroids formation. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2899-2909, 2018.
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http://dx.doi.org/10.1002/jbm.a.36479DOI Listing
November 2018

NIPAM-based Microgel Microenvironment Regulates the Therapeutic Function of Cardiac Stromal Cells.

ACS Appl Mater Interfaces 2018 Nov 29;10(44):37783-37796. Epub 2018 Oct 29.

School of Chemical Engineering , The University of Adelaide , Adelaide 5000 , Australia.

To tune the chemical, physical, and mechanical microenvironment for cardiac stromal cells to treat acute myocardial infarction (MI), we prepared a series of thermally responsive microgels with different surface charges (positive, negative, and neutral) and different degrees of hydrophilicity, as well as functional groups (carboxyl, hydroxyl, amino, and methyl). These microgels were used as injectable hydrogels to create an optimized microenvironment for cardiac stromal cells (CSCs). Our results indicated that a hydrophilic and negatively charged microenvironment created from poly( N-isopropylacrylamide- co-itaconic acid) was favorable for maintaining high viability of CSCs, promoting CSC proliferation and facilitating the formation of CSC spheroids. A large number of growth factors, such as vascular endothelial growth factor (VEGF), insulin-like growth factor I (IGF-1), and stromal-derived factor-1 (SDF-1) were released from the spheroids, promoting neonatal rat cardiomyocyte activation and survival. After injecting the poly( N-isopropylacrylamide- co-itaconic acid) microgel into mice, we examined their acute inflammation and T-cell immune reactions. The microgel itself did not elicit obvious immune response. We then injected the same microgel-encapsulated with CSCs into MI mice. The result revealed the treatment-promoted MI heart repair through angiogenesis and inhibition of apoptosis with an improved cell retention rate. This study will open a door for tailoring poly( N-isopropylacrylamide)-based microgel as a delivery vehicle for CSC therapy.
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http://dx.doi.org/10.1021/acsami.8b09757DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7034655PMC
November 2018

[Study on the 3,4-Dihydroxyphenylalanine Redox State Characterization Method of Mussel Adhesive Protein].

Zhongguo Yi Liao Qi Xie Za Zhi 2018 Sep;42(5):365-367

National Institutes for Food and Drug Control(Center for Medical Device Standardization Administration, CFDA), Beijing, 102629.

Objective: To investigate the feasibility of using liquid chromatography (HPLC) to characterize the 3, 4-Dihydroxyphenylalanine (DOPA) redox state of mussel adhesive protein (MAP).

Methods: The DOPA and protein contents of MAP were determined by HPLC, Arnow and Bradford methods respectively.

Results: With extended oxidation time, the protein contents of MAP samples remained unchanged whereas the DOPA contents declined. The retention times of main peaks in HPLC for both the accelerated oxidation and retained samples shifted as the storage time extended, which could be related to the changes of sample redox state.

Conclusions: The redox state of MAP can be characterized by the change of HPLC peak retention time. HPLC can be used in the research on the MAP redox state, which is beneficial to the product development and quality control.
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http://dx.doi.org/10.3969/j.issn.1671-7104.2018.05.015DOI Listing
September 2018

Transcriptome profiling analysis reveals the role of latrophilin in controlling development, reproduction and insecticide susceptibility in Tribolium castaneum.

Genetica 2018 Jun 24;146(3):287-302. Epub 2018 May 24.

Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing, 210023, China.

Latrophilin of Tribolium castaneum (Tclph) has been reported to play crucial roles in growth, development and reproduction. However, the regulatory mechanism of Tclph associated with these physiology processes is unknown. Thus, the global transcriptome profiles between RNAi treated (ds-Tclph) and control larvae of T. castaneum were analyzed by RNA-sequencing. Totally, 274 differentially expressed genes (DEGs) were identified between the ds-Tclph and control samples. These DEGs were classified into 42 GO functional groups, including developmental process, reproduction and stress response. The results indicated that knockdown of Tclph disturbed the antioxidant activity process, and partially inhibited the serine protease (SP) and lipase signaling pathways to regulate the development and reproduction as well as the decreasing of the stress response in T. castaneum. Additionally, knockdown of Tclph suppressed IMD immunity pathways which likely modulated the effects of Tclph on stress response. Interestingly, CSPs, ESTs, CYPs, AOXs and BGs were significantly down-regulated in ds-Tclph larvae, implying that they cooperated with Tclph to reduce the activity of cellular metabolism system. FMOs was up-regulated in ds-Tclph insects suggested it may be involved in detoxifying alkaloid of insect metabolism system. These results implied that Tclph participated in phase 0, I and II cellular detoxification. Furthermore, RNAi against Tclph increased larval susceptibility to carbamates and organophosphates insecticides, supporting that Tclph was indeed involved into the insecticide susceptibility in T. castaneum.
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http://dx.doi.org/10.1007/s10709-018-0020-4DOI Listing
June 2018

Enhanced multi-lineage differentiation of human mesenchymal stem/stromal cells within poly(N-isopropylacrylamide-acrylic acid) microgel-formed three-dimensional constructs.

J Mater Chem B 2018 Mar 9;6(12):1799-1814. Epub 2018 Mar 9.

School of Chemical Engineering, The University of Adelaide, Adelaide, SA 5005, Australia.

Human mesenchymal stem/stromal cells (hMSCs) are a potential cell source of stem cell therapy for many serious diseases and hMSC spheroids have emerged to replace single cell suspensions for cell therapy. Three-dimensional (3D) scaffolds or hydrogels which can mimic properties of the extracellular matrix (ECM) have been widely explored for their application in tissue regeneration. However, there are considerably less studies on inducing differentiation of hMSC spheroids using 3D scaffolds or hydrogels. This study is the first to explore multi-lineage differentiation of a stem cell line and primary stem cells within poly(N-isopropylacrylamide) (p(NIPAAm))-based thermosensitive microgel-formed constructs. We first demonstrated that poly(N-isopropylacrylamide-co-acrylic acid) (p(NIPAAm-AA)) was not toxic to hMSCs and the microgel-formed constructs facilitated formation of uniform stem cell spheroids. Due to functional enhancement of cell spheroids, hMSCs within the 3D microgel-formed constructs were induced for multi-lineage differentiation as evidenced by significant up-regulation of messenger RNA (mRNA) expression of chondrogenic and osteogenic genes even in the absence of induction media on day 9. When induction media were in situ supplied on day 9, mRNA expression of chondrogenic, osteogenic and adipogenic genes within the microgel-formed constructs were significantly higher than that in the pellet and 2D cultures, respectively, on day 37. In addition, histological and immunofluorescent images also confirmed successful multi-lineage differentiation of hMSCs within the 3D microgel-formed constructs. Hence, the thermosensitive p(NIPAAm-AA) microgel can be potentially used in an in vitro model for cell differentiation or in vivo transplantation of pre-differentiated human mesenchymal stromal cells into patients for specific lineage differentiation.
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http://dx.doi.org/10.1039/c8tb00376aDOI Listing
March 2018

Preparation, characterization and antioxidant activity of polysaccharide from spent Lentinus edodes substrate.

Int J Biol Macromol 2018 Jun 9;112:976-984. Epub 2018 Feb 9.

Key Laboratory of Systems Bioengineering, Ministry of Education (Tianjin University), Tianjin 300072, China; School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin 300072, China. Electronic address:

This study explored the potential of spent Lentinus edodes substrate, a by-product of mushroom industries causing environmental pollution, serving as materials to produce antioxidant polysaccharide. The extraction process of spent Lentinus edodes substrate polysaccharide (SLSP) was optimized and the effects of drying methods on chemical composition, morphological property and antioxidant activity were investigated. Results showed that freeze-dried SLSP (SLSP-F) exhibited the best quality in terms of the polysaccharide yield (13.00%) and antioxidant activity. The EC values of SLSP-F on DPPH, ABTS and superoxide anion radicals was 0.051mg/mL, 0.379mg/mL, 0.719mg/mL, respectively, which was significantly lower than that of freeze-dried Lentinus edodes polysaccharide (LP-F). After purification by Sephadex G-150, the purified SLSP-F (PSP) has a molecular weight of 16.77kDa. Compared with LP-F, PSP has more reducing sugars and uronic acids in chemical composition and higher contents of xylose, glucose and galactose in monosaccharide composition. FT-IR and NMR spectroscopy analysis revealed that PSP has both α and β glycosidic bonds and massive acetyl groups, which is different from LP-F mainly composed of 1, 3 linked α-D-Manp residue with some acetyl groups. The findings provided a reliable approach for the development of antioxidant polysaccharide from spent Lentinus edodes substrate.
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http://dx.doi.org/10.1016/j.ijbiomac.2018.01.196DOI Listing
June 2018

Albumin Binding Domain Fusing R/K-X-X-R/K Sequence for Enhancing Tumor Delivery of Doxorubicin.

Mol Pharm 2017 11 9;14(11):3739-3749. Epub 2017 Oct 9.

State Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences , Beijing 100190, China.

For the purpose of improving the tumor delivery of doxorubicin (DOX), a kind of peptide-DOXO conjugate was designed and prepared, in which the peptide composed of an albumin-binding domain (ABD) and a tumor-specific internalizing sequence (RGDK or RPARPAR) was conjugated to a (6-maleimidocaproyl) hydrazone derivative of doxorubicin (DOXO-EMCH). The doxorubicin uptake by lung cancer cell line of A549 evidenced that the conjugates are capable of being internalized through a tumor-specific sequence mediated manner, and the intracellular imaging of distribution in A549 cell demonstrated that the conjugated doxorubicin can be delivered to the cell nucleus. The A549 cell cytotoxicity of peptide-DOXO conjugates was presented with IC values and shown in the range of about 9-11 μM. Pharmacokinetics study revealed that both conjugates exhibited nearly 5.5 times longer half-time than DOX, and about 4 times than DOXO-EMCH. The in vivo growth inhibitions of the two peptide-DOXO conjugates on BALB/c nude mice bearing A549 tumor (47.78% for ABD-RGDK-DOXO and 47.09% for ABD-RPARPAR-DOXO) were much stronger than that of doxorubicin and DOXO-EMCH (24.28% and 25.67% respectively) at a doxorubicin equivalent dose. Besides, the in vivo fluorescence imaging study confirmed that the peptide markedly increased the payload accumulation in tumor tissues and indicated that albumin binding domain fusing tumor-specific sequence effectively enhanced the tumor delivery of doxorubicin and thus improved its therapeutic potency.
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http://dx.doi.org/10.1021/acs.molpharmaceut.7b00497DOI Listing
November 2017

Identification and function analysis of ras-related nuclear protein from Macrobrachium rosenbergii involved in Spiroplasma eriocheiris infection.

Fish Shellfish Immunol 2017 Nov 19;70:583-592. Epub 2017 Sep 19.

Jiangsu Key Laboratory for Biodiversity & Biotechnology, Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Life Sciences, Nanjing Normal University, 1 Wenyuan Road, Nanjing 210023, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu Province, Lianyungang, Jiangsu 222005, China. Electronic address:

A ras-related nuclear protein (Ran) protein was obtained from Macrobrachium rosenbergii, named MrRan. Phylogenetic analysis results showed that MrRan was clustered in one group together with other crustaceans. Tissue distribution analysis revealed that MrRan was expressed mainly in gill, intestine and stomach, and expressed weakly in muscle. The MrRan expression levels in gill and hemocyte of prawns were significantly up-regulated after challenged by Spiroplasma eriocheiris. The copy number of S. eriocheiris in MrRan dsRNA injection group was significantly less than control groups during infection. Meanwhile, silencing MrRan obviously increased the survival rate of prawns. The subcellular localization experiment suggested that recombinant MrRan was mainly located in the nucleus, and relatively weak in the cytoplasm. Finally, over-expression in Drosophila S2 cell indicated that MrRan could increase copies of S. eriocheiris and decrease of cell viability. The present study suggested that MrRan participated in regulating the phagocytosis of S. eriocheiris in M. rosenbergii.
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http://dx.doi.org/10.1016/j.fsi.2017.09.046DOI Listing
November 2017

Prompt and Robust Humoral Immunity Elicited by a Conjugated Chimeric Malaria Antigen with a Truncated Flagellin.

Bioconjug Chem 2018 03 22;29(3):761-770. Epub 2017 Aug 22.

National Key Laboratory of Biochemical Engineering, Institute of Process Engineering , Chinese Academy of Sciences , Beijing 100190 , PR China.

As one of the pathogen-associated molecular patterns (PAMPs), flagellin is recently utilized as a potent adjuvant for many subunit vaccines. In this study, a truncated flagellin (tFL) with deletion of the hypervariable regions was adopted as a carrier-adjuvant by chemical conjugation with a chimeric malaria antigen M.RCAg-1 (M312) via a heterobifunctional polyethylene glycol (PEG) linker. After booster immunization in mice without any extra adjuvants, the M312-PEG-tFL conjugates elicited M312-specific antibody titers 100-1000 times higher than M312 and 10-100 times higher than the physical mixture of M312 and tFL. The elicited specific antibodies could recognize the native parasites, and the immunofluorescence assay (IFA) titer was 2100 for M312-P5k-tFL, which was about 7 times higher than M312. Furthermore, the IFA titers of the conjugates were comparable to the positive control of complete Freund's adjuvant (CFA). Compared to M312, the M312-PEG-tFL conjugates enhanced the proliferation index, lymphocyte activation, and memory T-cell generation. IgG subclasses of sera and cytokines analysis of splenocytes showed that conjugation with tFL could slightly trigger the Th1 polarization, while the antigen alone predominantly induced a Th2-biased immune response. Furthermore, a more-efficient innate immune response was provoked by the M312-PEG-tFL conjugates, as determined by the detection of antigen-specific TNF-α secretion by splenocytes. Our results indicated that tFL mainly retained the function as an agonist of TLR5. Conjugation of antigen to tFL could induce strong humoral and moderate cellular immune responses. Thus, conjugation of antigen to tFL as a potent carrier-adjuvant is an effective strategy for developing a promising protein-based vaccine.
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http://dx.doi.org/10.1021/acs.bioconjchem.7b00320DOI Listing
March 2018

Adhesin-Like Protein (ALP) Interacts with Epidermal Growth Factor (EGF) Domain Proteins to Facilitate Infection.

Front Cell Infect Microbiol 2017 26;7:13. Epub 2017 Jan 26.

Jiangsu Key Laboratory for Biodiversity and Biotechnology and Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Life Sciences, Nanjing Normal UniversityNanjing, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu ProvinceLianYungang, China.

is a novel pathogen found in recent years, causing the tremor disease (TD) of Chinese mitten crab . Like infects the newborn mouse (adult mice are not infected) and can cause cataract. Adhesion-related protein is an important protein involved in the interaction between pathogen and host. In this study, the Adhesin-like Protein (ALP) of was detected on its outer membrane by using immune electron microscopy, and was found to be involved in the bacterium's infection of mouse embryo fibroblasts (3T6-Swiss albino). Yeast two-hybrid analysis demonstrated that ALP interacts with a diverse group of mouse proteins. The interactions between recombinant partial fibulin7 (FBLN7; including two epidermal growth factor [EGF] domains) and ALP were confirmed by Far-western blotting and colocalization. We synthetized the domains of FBLN7 [EGF domain: amino acids 136-172 and complement control protein (CCP) domain: 81-134 amino acids], and demonstrated that only EGF domain of FBLN7 can interact with ALP. Because the EGF domain has high degree of similarity to EGF, it can activate the downstream EGFR signaling pathway, in key site amino acids. The EGFR pathway in 3T6 cells was restrained after rALP stimulation resulting from competitive binding of ALP to EGF. The unborn mouse, newborn mouse, and the adult mouse with cataract have a small amount of expressed FBLN7; however, none was detected in the brain and very little expression was seen in the eye of normal adult mice. In short, ALP as a surface protein, is critical for infection and further supports the role of ALP in infection by competitive effection of the EGF/EGFR axis of the target cells.
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http://dx.doi.org/10.3389/fcimb.2017.00013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5266718PMC
September 2017

Cell-penetrating peptide-labelled smart polymers for enhanced gene delivery.

Eng Life Sci 2017 Feb 4;17(2):193-203. Epub 2016 Oct 4.

School of Chemical Engineering University of Adelaide Adelaide Australia.

Highly efficient gene delivery vehicles are pursued to progress gene therapy. In this study, we developed the cell-penetrating peptide-labelled and degradable gene carriers for efficient external gene transfection. The cationic carriers were prepared by coupling low-molecular-weight polyethylenimine (PEI800) with 4'4-dithiodibutyric acid (DA), and HIV-1 Trans-Activator of Transcription (TAT) was conjugated to the carriers as a penetrating peptide. The resulted PEI-DA-TAT was able to condense plasmid DNA (pDNA) into a complex with a hydrodynamic size of around 150 nm under a neutral condition. PEI-DA-TAT showed negligible cytotoxicity to both Hela and HEK 293 cells with the cell viability of more than 80% beyond the carrier concentration of 50 μg/mL. This new carrier displayed better performance in regard to DNA transfection efficiency in comparison with the carriers of non-TAT labelled PEI-DA, commercial PEI25K and low-molecular-weight PEI (PEI800). The transfection efficiency of PEI-DA-TAT was increased by 8% compared with PEI-DA and PEI25K. The experimental findings suggested that the developed PEI-DA-TAT is a promising carrier for efficient DNA delivery with low cytotoxicity for gene therapy.
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http://dx.doi.org/10.1002/elsc.201600069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6999197PMC
February 2017

Microengineered 3D cell-laden thermoresponsive hydrogels for mimicking cell morphology and orientation in cartilage tissue engineering.

Biotechnol Bioeng 2017 01 9;114(1):217-231. Epub 2016 Aug 9.

School of Chemical Engineering, The University of Adelaide, Adelaide, SA, 5005, Australia.

Mimicking the zonal organization of native articular cartilage, which is essential for proper tissue functions, has remained a challenge. In this study, a thermoresponsive copolymer of chitosan-g-poly(N-isopropylacrylamide) (CS-g-PNIPAAm) was synthesized as a carrier of mesenchymal stem cells (MSCs) to provide a support for their proliferation and differentiation. Microengineered three-dimensional (3D) cell-laden CS-g-PNIPAAm hydrogels with different microstripe widths were fabricated to control cellular alignment and elongation in order to mimic the superficial zone of natural cartilage. Biochemical assays showed six- and sevenfold increment in secretion of glycosaminoglycans (GAGs) and total collagen from MSCs encapsulated within the synthesized hydrogel after 28 days incubation in chondrogenic medium. Chondrogenic differentiation was also verified qualitatively by histological and immunohistochemical assessments. It was found that 75 ± 6% of cells encapsulated within 50 μm wide microstripes were aligned with an aspect ratio of 2.07 ± 0.16 at day 5, which was more organized than those observed in unpatterned constructs (12 ± 7% alignment and a shape index of 1.20 ± 0.07). The microengineered constructs mimicked the cell shape and organization in the superficial zone of cartilage whiles the unpatterned one resembled the middle zone. Our results suggest that microfabrication of 3D cell-laden thermosensitive hydrogels is a promising platform for creating biomimetic structures leading to more successful multi-zonal cartilage tissue engineering. Biotechnol. Bioeng. 2017;114: 217-231. © 2016 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/bit.26061DOI Listing
January 2017

Poly(N-isopropylacrylamide) hydrogel/chitosan scaffold hybrid for three-dimensional stem cell culture and cartilage tissue engineering.

J Biomed Mater Res A 2016 11 8;104(11):2764-74. Epub 2016 Jul 8.

School of Chemical Engineering, The University of Adelaide, Adelaide, SA5005, Australia.

Providing a controllable and definable three-dimensional (3D) microenvironment for chondrogenic differentiation of mesenchymal stem cells (MSCs) remains a great challenge for cartilage tissue engineering. In this work, poly(N-isopropylacrylamide) (PNIPAAm) polymers with the degrees of polymerization of 100 and 400 (NI100 and NI400) were prepared and the polymer solutions were introduced into the preprepared chitosan porous scaffolds (CS) to form hybrids (CSNI100 and CSNI400, respectively). SEM images indicated that the PNIPAAm gel partially occupied chitosan pores while the interconnected porous structure of chitosan was preserved. MSCs were incorporated within the hybrid and cell proliferation and chondrogenic differentiation were monitored. After 7-day incubation of the cell-laden constructs in a growth medium, the cell viability in CSNI100 and CSNI400 were 54 and 108% higher than that in CS alone, respectively. Glycosaminoglycan and total collagen contents increased 2.6- and 2.5-fold after 28-day culture of cell-laden CSNI400 in the chondrogenic medium. These results suggest that the hybrid structure composed of the chitosan porous scaffold and the well-defined PNIPAAm hydrogel, in particular CSNI400, is suitable for 3D stem cell culture and cartilage tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2764-2774, 2016.
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http://dx.doi.org/10.1002/jbm.a.35810DOI Listing
November 2016

A novel C-type lectin is involved in the innate immunity of Macrobrachium nipponense.

Fish Shellfish Immunol 2016 Mar 22;50:117-26. Epub 2016 Jan 22.

Jiangsu Key Laboratory for Biodiversity & Biotechnology and Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Life Sciences, Nanjing Normal University, 1 Wenyuan Road, Nanjing 210023, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu Province, Lianyungang, Jiangsu 222005, China. Electronic address:

C-type lectins (CTLs) play important roles in invertebrate innate immunity by recognizing and eliminating pathogens. In the present study, a low-density lipoprotein receptor class A (LDLa) domain-containing CTL was identified from the oriental river prawn Macrobrachium nipponense, designated as MnCTLDcp1. The full-length cDNA of MnCTLDcp1 was composed of 1462 bp, with a 999-bp ORF encoding a 332-aa protein. An LDLa and a single C-type lectin-like domain (CTLD) were found. The mRNA transcripts of MnCTLDcp1 was expressed the highest in heart. After the prawns were challenged by Aeromonas hydrophila and Staphylococcus aureus, the expression level of MnCTLDcp1 in heart and hemocytes were all significantly up-regulated. Sugar binding assay revealed that the MnCTLDcp1 could bind to the glycoconjugates of bacteria surface, such as LPS, PGN and they can compete with bacterial as competitors. The recombinant MnCTLDcp1 agglutinates Gram-positive (S. aureus and Bacillus subtilis) and Gram-negative bacteria (A. hydrophila, Vibrio parahaemolyticus, Escherichia coli and Pseudomonas aeruginosa) in the presence of calcium and also could bind to these bacteria. These results clearly suggested that MnCTLDcp1 functions as a pattern-recognition receptor involved in the innate immunity of M. nipponense.
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http://dx.doi.org/10.1016/j.fsi.2016.01.026DOI Listing
March 2016

Influence of polymer molecular weight on the in vitro cytotoxicity of poly (N-isopropylacrylamide).

Mater Sci Eng C Mater Biol Appl 2016 Feb 22;59:509-513. Epub 2015 Oct 22.

School of Chemical Engineering, The University of Adelaide, Adelaide SA5005, Australia. Electronic address:

Poly (N-isopropylacrylamide) (PNIPAAm) is a thermosensitive polymer with various biomedical applications. We examined molecular weight (MW)-dependent cytotoxicity of PNIPAAm. Our results indicated that low-MW PNIPAAm (degree of polymerization (DP)=35) is inherently toxic to cells. Moderate-MW PNIPAAms with their DP between 100 and 200 are non-cytotoxic. When cells are seeded on top of a polymer-coated surface, PNIPAAm with a higher MW (DP=400) shows non/low cytotoxicity, while when monolayer cells are exposed to the polymer solution, cell viability drops drastically. This may be due to lack of nutrient and oxygen rather than intrinsic toxicity of the polymer.
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http://dx.doi.org/10.1016/j.msec.2015.10.043DOI Listing
February 2016