Early Detection of Coal Spontaneous Combustion by Complex Acoustic Waves in a Concealed Fire Source.

Prevention and control of coal spontaneous combustion are key to coal mining and storage. Existing technologies for the detection of coal spontaneous combustion have limitations, but coal spontaneous combustion creates some serious disasters in areas of the world where coal mining and/or storage exists. New technologies to detect coal spontaneous combustion are urgently needed to reduce the loss of life and resources. The article reviews the main techniques employed to detect coal spontaneous combustion and their advantages and disadvantages; it also reviews the good application prospect of acoustic temperature measurement technology on coal spontaneous combustion and introduces the basic principle of acoustic coal temperature measurement. The evolution of combustion sound and the propagation and attenuation of acoustic waves in quasi-porous media are discussed to form the basis for the development of acoustic thermometry technologies that can be used to accurately identify acoustic signals and temperature fields in loose coal. The concept of "single-source" coal temperature measurement to "dual-source" coal temperature measurement achieved by using combustion sound and an additional sound source device in the automatic combustion of loose coal in the mined area is discussed. The deep learning methods and correlation analyses are available to map the relationships between combustion sound, coal temperature, and sound velocity, and acquire coal temperature from dual source composite acoustic signals. The study lays the foundation for the development of acoustic thermometry technologies that have applications in different stages of combustion and applied to the early warning, prevention, and control of spontaneous combustion in coal, and it contributes to improving the environmental safety and efficiency of coal mining and storage.

Meticulous Graded and Early Warning System of Coal Spontaneous Combustion Based on Index Gases and Characteristic Temperature.

The accurate prediction of coal spontaneous combustion (CSC) in the goaf areas of coal mines is a vital aspect of the migration from passive to active fire prevention and control. However, CSC is highly complicated and existing technologies cannot accurately monitor coal temperatures over wide expanses. Thus, it may be beneficial to assess CSC based on various index gases produced by the reactions of coal. In the present study, the CSC process was simulated by temperature-programmed experiments, and the relationships between index gas concentrations with the coal temperature were determined using logistic fitting functions. CSC was divided into seven stages, and a coal seam spontaneous ignition early warning system involving six criteria was established. Field trials demonstrated that this system is a viable approach to predicting coal seam fires and meets the requirements for the active prevention and control of coal combustion. This work establishes an early warning system based on specific theoretical guidelines that permits the identification of CSC and the implementation of active fire prevention and extinguishing measures.

A membrane-associated metalloprotease of Schistosoma japonicum structurally related to the FACE-1/Ste24p protease family.

The CaaX proteases are closely related in the post-translational modification of many membrane-bound or secreted proteins and play a key role in the activation or stabilization of these molecules belonging to the CAAX family. In this study, a full-length cDNA putatively encoding a FACE-1/Ste24p CaaX protease (type I) of the Schistosoma japonicum was isolated. The cDNA, named SjSte24p, composed of 1646 bp and encoded 473 amino acids with predicted Mr/pI as 54.77 kDa/8.04. SjSte24p is a monoexonic gene constantly expressed in the parasite from cercariae to adult stages. It contained the characteristic of CaaX protease topology, including seven trans-membrane domains and a metallo-protease segment with a zinc-binding motif (HEXXH). SjSte24p shared a considerable degree of sequence identity with the type I CaaX proteases. A phylogenetic analysis showed that this protein family is tightly conserved from fungi to vertebrates. The expressed recombinant SjSte24p protein showed a proteolytic activity, which was inhibited by EDTA. Its activity was increased at low doses of the Zn2+ (0.001-0.01 mM); but was reversibly down-regulated at high doses (>0.1 mM). The native SjSte24p appeared to function in insoluble from. The protein was mainly localized in the tegument on the surface of adult worms. These results indicated that the SjSte24p is a practical zinc-dependent metalloprotease, which belongs to the FACE-1/Ste24p protease family.

Isolation and characterization of a novel serine protease inhibitor, SjSPI, from Schistosoma japonicum.

Serine proteinase inhibitor (Serpin, SPI) is a vital superfamily of endogenous inhibitors that monitor proteolytic events active in a number of biological functions. In this study, we isolated a full length gene encoding a novel serine protease inhibitor of Schistosoma japonicum (SjSPI) and characterized its molecular properties. Our result showed that SjSPI contained an open reading frame of 1,218 bp, which encoded 405 amino acid residues. Chromosomal structure analysis showed that SjSPI gene was comprised of six exons separated by five introns. It had essential structural motifs which were well conserved among the Serpin superfamily and showed 17-33% sequence identities with Serpins from other helminthic parasites. Trematode Serpin diverged separately into two different subclades and that the SjSPI clustered Subclade I. Exon-intron structures of trematode Serpins were highly conserved, closely with cestode Serpins. No signal peptide but a strongly transmembrane domain was predicted to exist in SjSPI, suggesting that the protein might be a soluble membrane-associated protein. Homology modeling predicted in silico confirmed that the SjSPI structure also belonged to the Serpin superfamily, containing nine α-helices and a reactive central loop. The bacterially expressed recombinant GST-SjSPI protein effectively inhibited the activities of chymotrypsin, trypsin and thrombin. Expression of SjSPI was detected throughout various developmental stages of the parasite in host and reached its maximal levels at the adult and egg stages, which suggests that SjSPI may be possibly involved in maintaining the physiology of eggs by regulating endogenous serine proteases.

Functionally Expression of Metalloproteinase in Taenia solium Metacestode and Its Evaluation for Serodiagnosis of Cysticercosis.

BACKGROUND: Parasite proteases have important roles in cleavage of host proteins during the invasion of host tissues and participate in the parasite's evasion from the host's immune response. The aim of the present study was to estimate a metalloproteinase properties of Taenia solium metacestode (TsMP) during host-parasite interactions, and evaluate its potential as a serodiagnostic antigen for cysticercosis. METHODS: The cDNA coding for the mature catalytic domain of TsMP was cloned into pGEX-6P-1 expression vector. A recombinant glutathione S-transferase and TsMP fusion protein was induced. After refolding and purification, enzymatic properties of the recombinant metalloproteinase were observed. Immunoblot assay was processed to evaluate its potential as a serodiagnostic antigen for cysticercosis. RESULTS: The recombinant TsMP protein showed proteolytic activity, which preferred host extracellular matrix proteins such as collagen and fibronectin as degradable substrates. In immunoblot assay, 87.5% of sera from patients with cysticercosis showed strong reactivity. In sera from patients with other parasitic infections and from normal controls, it showed high specificity. CONCLUSIONS: TsMP might be involved in the processing of numerous host proteins and play an important role in the parasite life cycle. A single recombinant TsMP antigen could have a potential value for serodiagnosis of cysticercosis.

Molecular cloning and characterization of a phospholipid hydroperoxide glutathione peroxidase gene from a blood fluke Schistosoma japonicum.

Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a major antioxidant enzyme and plays critical roles in the protection of cells against oxidative stress by catalysing reduction of lipid hydroperoxides. A full-length cDNA sequence corresponding to GPx gene from Schistosoma japonicum (designated SjGPx) was isolated and characterized. SjGPx contained an in-frame TGA codon for selenocysteine (Sec) and a concurrent Sec insertion sequence in its 3'-untranslated region. Protein encoded by SjGPx demonstrated a primary structure characteristic to the PHGPx family, including preservation of catalytic domains and absence of the subunit interaction domains. Phylogenetic analysis revealed that the SjGPx was highly related to the other PHGPx-related members, and clustered into the trematode subclade II. Semi-quantitative reverse transcription PCR and western blotting showed that the SjGPx was mainly expressed in the female adults and eggs. RNA interference was employed to investigate the effects of knockdown of SjGPx. SjGPx expression level was significantly reduced on the 5th day post-RNAi. We observed a 53.86% reduction in total GPx activity and the eggs severely deformed. Oxidative stimulation of viable worms with H2O2 or paraquat resulted in 1.6- to 2.1-fold induction of the GPx activity. Our results revealed that the SjGPx protein is selenium-dependent PHGPx, which might actively participate in the detoxification of oxidative damage during egg production.

Expression and characterization of a phospholipid hydroperoxide glutathione peroxidase gene in Schistosoma japonicum.

Phospholipid hydroperoxide glutathione peroxidase (PHGPx, GPx4) is a major antioxidant enzyme, which plays unique roles in the protection of cells against oxidative stress by catalysing reduction of lipid hydroperoxides. We isolated and characterized a full-length cDNA sequence encoding GPx gene from a blood fluke, Schistosoma japonicum (designated SjGPx), which contained an in-frame TGA codon for selenocysteine (Sec) and a concurrent Sec insertion sequence in its 3'-untranslated region. Protein encoded by SjGPx demonstrated a primary structure characteristic to the PHGPx family, including preservation of catalytic domains and absence of the subunit interaction domains. Semi-quantitative reverse transcription PCR and Western blotting showed that the SjGPx was mainly expressed in the female adults and eggs. RNA interference approach was employed to investigate the effects of knockdown of SjGPx. SjGPx expression level was significantly reduced on the 5th day post-RNAi. Significantly reduction in GPx enzyme activities, as well as obvious changes in morphology of intrauterine eggs followed the reduction in SjGPx transcript level. We observed a 63·04% reduction in GPx activity and the eggs severely deformed. Our results revealed that SjGPx protein might be involved in the provision of enzyme activity during egg production.

Reconstruction of upper lip scar using tissue expander advancement flap.

Some patients who previously underwent burns have upper lip deformities or extensive scars so they seek for treatments.We reported a patient who had extensive scars all over the upper lip. In the patient's case, we reconstructed the entire upper lip using a tissue expander advancement flap from the bilateral lip. This improved the upper lip contracture and appearance. The removal of the scar from the upper lip provided a satisfactory result.The use of 2 large tissue expander advancement flaps from the bilateral upper lip has several advantages, such as inconspicuous scar, nice skin texture, and color match.

Preparation, characterization and photocatalytic activity of AgBr/BiVO4 composite photocatalyst.

The AgBr/BiVO4 heterostructure were synthesized. The heterostructure materials were characterized by XRD, DRS, SEM-EDS, TEM, XPS and TG-DSC. In order to investigate the photocatalytic activity of AgBr/BiVO4 heterostructures, the methylene blue (MB) dye was used as a model compound. The result suggested that under visible light irradiation, the degradation efficiency of AgBr/BiVO4 reached 83.1% within 2.5 h, which was higher than that of the pure BiVO4. The presence of AgBr significantly increased the photocatalytic activity of the composites. The photocatalytic mechanism was also discussed.

Antimalarial and Structural Studies of Pyridine-containing Inhibitors of 1-Deoxyxylulose-5-phosphate Reductoisomerase.

1-Deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) in the non-mevalonate isoprene biosynthesis pathway is a target for developing antimalarial drugs. Fosmidomycin, a potent DXR inhibitor, showed safety as well as efficacy against P. falciparum malaria in clinical trials. Based on our previous quantitative structure activity relationship (QSAR) and crystallographic studies, several novel pyridine-containing fosmidomycin derivatives were designed, synthesized and found to be highly potent inhibitors of P. falciparum DXR (PfDXR) having Ki values of 1.9 - 13 nM, with the best one being ~11× more active than fosmidomycin. These compounds also potently block the proliferation of multi-drug resistant P. falciparum with EC50 values as low as 170 nM. A 2.3 Å crystal structure of PfDXR in complex with one of the inhibitors is reported, showing the flexible loop of the protein undergoes conformational changes upon ligand binding and a hydrogen bond and favorable hydrophobic interactions between the pyridine group and PfDXR account for the enhanced activity.

Expression pattern and substrate specificity of Clonorchis sinensis tyrosinases.

Tyrosinase (TYR) is a copper-containing glycoenzyme that mediates hydroxylation of tyrosine into dihydroxyphenylalanine and oxidation of dihydroxyphenylalanine into dihydroxyphenylalanine quinone. TYRs play pivotal roles in eggshell sclerotisation of trematode parasites, while their comprehensive biochemical properties remain elusive. We characterised genes encoding four TYRs (CsTYR1-4) of Clonorchis sinensis, a causative agent of human hepatobiliary disease. These genes shared tightly conserved amino acid residues, two copper binding catalytic motifs and a cysteine-rich epidermal growth factor-like domain. The native and recombinant CsTYRs showed high reactivity against diphenol compounds, especially those with hydroxyl groups in ortho-positions (catechol and l-dihydroxyphenylalanine), but showed minimal activity toward monophenol compounds. Diphenolase activity was enhanced by increased pH of the reaction buffer from 5.0 to 7.0. The temporal induction of CsTYR expression coordinated with the sexual maturation of the worm; enzyme activity was mainly in the vitelline glands and intrauterine immature eggs proximal to the ovary. The primary structures and functional domains of CsTYRs showed significant similarities to those of the vertebrate orthologs, whereas the amino acids shared with the nematode and insect proteins were largely restricted in the bicopper active center. Unlike highly diverged TYR homologs in vertebrates, multiple paralogs have not yet evolved into the separate lineages in trematode genomes, suggesting that duplication of TYR genes might relate to increased genic dosage/redundancy in trematodes. In vitro treatment of copper chelator, diethyldithiocarbamic acid, inhibited generation of phenotypically normal egg. TYR proteins are essential for C. sinensis reproduction, thus might be targeted for therapeutic and vaccine strategies against clonorchiasis, which is prevalent in several Asian countries and is one of the most important predisposing factors for human cholangiocarcinoma. The close phylogenetic relationships between trematode and vertebrate homologs also provide a molecular clue to understand the multifaceted evolutionary pathway of TYR homologs across animal taxa.

Visible-light-induced WO3/g-C3N4 composites with enhanced photocatalytic activity.

Novel WO3/g-C3N4 composite photocatalysts were prepared by a calcination process with different mass contents of WO3. The photocatalysts were characterized by thermogravimetric analysis (TG), powder X-ray diffraction (XRD), scanning electron microscopy (SEM) and energy dispersive X-ray spectrometry (EDS), high-resolution transmission electron microscopy (HRTEM), UV-vis diffuse reflection spectroscopy (DRS), X-ray photoelectron spectroscopy (XPS), photoluminescence (PL) and electrochemical impedance spectroscopy (EIS). The photocatalytic activity of the photocatalysts was evaluated by degradation of methylene blue (MB) dye and 4-chlorophenol (4-CP) under visible light. The results indicated that the WO3/g-C3N4 composite photocatalysts showed higher photocatalytic activity than both the pure WO3 and pure g-C3N4. The optimum photocatalytic activity of WO3/g-C3N4 at a WO3 mass content of 9.7% under visible light irradiation was up to 4.2 times and 2.9 times as high as that of the pure WO3 and pure g-C3N4, respectively. The remarkably increased performance of WO3/g-C3N4 was mainly attributed to the synergistic effect between the interface of WO3 and g-C3N4, including enhanced optical absorption in the visible region, enlarged specific surface areas and the suitable band positions of WO3/g-C3N4 composites.

Differential activation of diverse glutathione transferases of Clonorchis sinensis in response to the host bile and oxidative stressors.

BACKGROUND: Clonorchis sinensis causes chronic cumulative infections in the human hepatobiliary tract and is intimately associated with cholangiocarcinoma. Approximately 35 million people are infected and 600 million people are at risk of infections worldwide. C. sinensis excretory-secretory products (ESP) constitute the first-line effector system affecting the host-parasite interrelationship by interacting with bile fluids and ductal epithelium. However, the secretory behavior of C. sinensis in an environment close to natural host conditions is unclear. C. sinensis differs from Fasciola hepatica in migration to, and maturation in, the hepatic bile duct, implying that protein profile of the ESP of these two trematodes might be different from each other. METHODOLOGY/PRINCIPAL FINDINGS: We conducted systemic approaches to analyze the C. sinensis ESP proteome and the biological reactivity of C. sinensis glutathione transferases (GSTs), such as global expression patterns and induction profiles under oxidative stress and host bile. When we observed ex host excretion behavior of C. sinensis in the presence of 10% host bile, the global proteome pattern was not significantly altered, but the amount of secretory proteins was increased by approximately 3.5-fold. Bioactive molecules secreted by C. sinensis revealed universal/unique features in relation to its intraluminal hydrophobic residing niche. A total of 38 protein spots identified abundantly included enzymes involved in glucose metabolism (11 spots, 28.9%) and diverse-classes of glutathione transferases (GSTs; 10 spots, 26.3%). Cathepsin L/F (four spots, 10.5%) and transporter molecules (three spots, 7.9%) were also recognized. The universal secretory proteins found in other parasites, such as several enzymes involved in glucose metabolism and oxygen transporters, were commonly detected. C. sinensis secreted less cysteine proteases and fatty acid binding proteins compared to other tissue-invading or intravascular trematodes. Interestingly, secretion of a 28 kDa σ-class GST (Cs28σGST3) was significantly affected by the host bile, involving reduced secretion of the 28 kDa species and augmented secretion of Cs28σGST3-related high-molecular-weight 85 kDa protein. Oxidative stressors induced upregulated secretion of 28 kDa Cs28σGST3, but not an 85 kDa species. A secretory 26 kDa µ-class GST (Cs26µGST2) was increased upon treatment with oxidative stressors and bile juice, while another 28 kDa σ-class GST (Cs28σGST1) showed negligible responses. CONCLUSIONS/SIGNIFICANCE: Our results represent the first analysis of the genuine nature of the C. sinensis ESP proteome in the presence of host bile mimicking the natural host environments. The behavioral patterns of migration and maturation of C. sinensis in the bile ducts might contribute to the secretion of copious amounts of diverse GSTs, but a smaller quantity and fewer kinds of cysteine proteases. The Cs28σGST1 and its paralog(s) detoxify endogenous oxidative molecules, while Cs28σGST3 and Cs26µGST2 conjugate xenobiotics/hydrophobic substances in the extracellular environments, which imply that diverse C. sinensis GSTs might have evolved for each of the multiple specialized functions.

The CNT modified white C3N4 composite photocatalyst with enhanced visible-light response photoactivity.

A novel, multi-walled carbon nanotubes (CNT) modified white C3N4 composite (CNT/white C3N4) with enhanced visible-light-response photoactivity was prepared. The white C3N4 and CNT combined together and formed the CNT/white C3N4 composite due to electrostatically-driven self-assembly with the hydrothermal method. The as-prepared white C3N4 and CNT/white C3N4 composite photocatalyst were characterized by transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), UV-vis absorption spectra, X-ray photoelectron spectroscopy (XPS) and photoluminescence spectroscopy (PL). The photoelectrochemical i-t curves were tested using several on-off cycles of light irradiation. The photoactivity of the catalysts was evaluated by degrading methylene blue (MB) dye solution. The results showed that the photoactivity for the degradation of MB solution was in the following order: CNT/white C3N4 composite > C3N4 > the white C3N4. The photoactivity of the CNT/white C3N4 composite was 66.5% and 34.5% higher than that of the white C3N4 sample and that of the C3N4 at 1.5 h, respectively. The degradation rate of the CNT/white C3N4 photocatalyst was almost 8.1 times as high as that of the white C3N4. The results indicated that CNT played an important role, which led to the efficient separation of the photo-generated charge carriers. The reason why the photoactivity of the CNT/white C3N4 was much higher than that of C3N4 and the white C3N4 was discussed. A possible mechanism of CNT on the enhancement of composites' visible light performance was also proposed.

Expression, characterization and inhibition of Toxoplasma gondii 1-deoxy-D-xylulose-5-phosphate reductoisomerase.

The apicomplexan parasite Toxoplasma gondii, the causative agent of toxoplasmosis, is an important human pathogen. 1-Deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) in the non-mevalonate isoprene biosynthesis pathway is essential to the organism and therefore a target for developing anti-toxoplasmosis drugs. In order to find potent inhibitors, we expressed and purified recombinant T. gondii DXR (TgDXR). Biochemical properties of this enzyme were characterized and an enzyme activity/inhibition assay was developed. A collection of 11 compounds with a broad structural diversity were tested against TgDXR and several potent inhibitors were identified with Ki values as low as 48 nM. Analysis of the results as well as those of Escherichia coli and Plasmodium falciparum DXR enzymes revealed a different structure-activity relationship profile for the inhibition of TgDXR.

Thermodynamic Investigation of Inhibitor Binding to 1-Deoxy-D-Xylulose-5-Phosphate Reductoisomerase.

Isothermal titration calorimetry (ITC) was used to investigate the binding of six inhibitors to 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR), a target for developing novel anti-infectives. The binding of hydroxamate inhibitors to E. coli DXR is Mg(2+)-dependent, highly endothermic (ΔH: 22.7-24.3 kJ/mol) and entropy-driven, while that of non-hydroxamate compounds is metal ion independent and exothermic (ΔH: -19.4- -13.8 kJ/mol), showing hydration/dehydration of the enzyme metal ion binding pocket account for the drastic ΔH change. However, for DXRs from Plasmodium falciparum and Mycobacterium tuberculosis, the binding of all inhibitors is exothermic (ΔH: -24.9 - -9.2 kJ/mol), suggesting the metal ion binding sites of these two enzymes are considerably less hydrated. The dissociation constants measured by ITC are well correlated with those obtained by enzyme inhibition assays (R(2) = 0.75). Given the rapid rise of antibiotic resistance, this work is of interest since it provides novel structural implications for rational development of potent DXR inhibitors.

siRNA-mediated knockdown of two tyrosinase genes from Schistosoma japonicum cultured in vitro.

The cross-linking process of eggshell proteins in helminths is dependent on the activities of tyrosinases (TYRs), which can be inhibited by phenol oxidase inhibitors. Two genes encoding TYRs, SjTYR1 and SjTYR2, have been identified in Schistosoma japonicum. In this study, siRNA-mediated RNA interference (RNAi) was performed to silence these two SjTYR genes to evaluate their roles in eggshell formation. The effects of individual or double knockdown of the SjTYR genes were compared by determining SjTYR1/SjTYR2 transcript levels, enzyme activities, and by observing the morphology and amounts of intrauterine eggs. Results showed that SjTYR transcript levels were significantly reduced on the 3rd day post-RNAi. Significant reductions in TYR enzyme activities, as well as obvious changes in morphology and the number of intrauterine eggs followed the reductions in SjTYR transcript levels. On the 8th day after simultaneous knockdown of both SjTYR genes, which effected a 40% reduction in SjTYR1 transcript level and a 59% reduction in SjTYR2 transcript level, we observed an 80% reduction in diphenol oxidase (DPO) activity of TYRs, and a 74% reduction in the number of normal eggs in female uteri. Knockdown of both SjTYR genes has a greater effect than single knockdown of the SjTYR genes. These results demonstrate that both SjTYRs play an important role in eggshell sclerotization of S. japonicum, and that their enzyme activities depend on the transcript levels of two SjTYR genes.

Synthesis and structure-activity relationship investigation of adenosine-containing inhibitors of histone methyltransferase DOT1L.

Histone3-lysine79 (H3K79) methyltransferase DOT1L has been found to be a drug target for acute leukemia with MLL (mixed lineage leukemia) gene translocations. A total of 55 adenosine-containing compounds were designed and synthesized, among which several potent DOT1L inhibitors were identified with K(i) values as low as 0.5 nM. These compounds also show high selectivity (>4500-fold) over three other histone methyltransferases. Structure-activity relationships (SAR) of these compounds for their inhibitory activities against DOT1L are discussed. Potent DOT1L inhibitors exhibit selective activity against the proliferation of MLL-translocated leukemia cell lines MV4;11 and THP1 with EC(50) values of 4-11 µM. Isothermal titration calorimetry studies showed that two representative inhibitors bind with a high affinity to the DOT1L:nucleosome complex and only compete with the enzyme cofactor SAM (S-adenosyl-L-methionine) but not the substrate nucleosome.

[Construction and transplantation of tissue-engineered skin with mouse embryonic fibroblasts in SD mice].

OBJECTIVE: To investigate the application and mechanism of tissue-engineered skin with mouse embryonic fibroblasts (MEFs) for the full-thickness skin defects on mice. METHODS: The MEFs and fibroblasts were cultured and seeded in scaffold made of rat tail collage. ELISA method was used for detection of secretory function. The full-thickness skin defects were created on mice and covered by MEFs-scaffold complex (experimental group), or FBs-scaffold complex (control group 1), or scaffold only (control group 2). The process of wound healing was evaluated by observation of the re-epithelization rate. Microvessel density (MVD) and vimentin within the wound sites were also detected with immunohistochemistry staining technique to describe the characteristics of wound healing. Hoechst 33342 staining was performed to trace MEFs'fate. RESULTS: MEFs scaffold group had higher level secretion of IL-6 and lower of TGF-beta1 than FBs scaffold group (P<0.05). Compared with wounds in control groups, the wounds in MEFs group healed markedly fast (P<0.05) and the MVD was significantly higher (P <0.05). The fibroblasts in the wounds of MEFs group were arranged regularly and the MEFs decreased during the healing process. CONCLUSIONS: The MEFs-scaffold complex can promote wound healing with less scar formation. MEFs may have an inducing effect on the wound healing.

Characterization and expression of a novel cystatin gene from Schistosoma japonicum.

Cystatins are a family of cysteine protease inhibitors that play a crucial role in the immune evasion from their host and in the adaptation to host defence. Here, we isolated a full-length cDNA sequence inferred to encode a novel cystatin gene from a blood fluke, Schistosoma japonicum. The cDNA, designated SjCystatin, comprised an open reading frame (ORF) of 306 bp, and encoded 101 amino acids with a predicted molecular weight of 11.3 kDa. This predicted protein shared a significant degree of sequence identity with the type I cystatin (stefin) of Schistosoma mansoni and Homo sapiens. These proteins exhibited a typical cystatin topology, including the absence of disulfide bonds and three conserved catalytic motifs, Gly at the N-terminus (Gly(6)), Gln-X-Val-X-Gly motif (Q(49)VVAG(53)) and an LP pair at the C-terminus (L(76)P(77)). The SjCystatin gene spanned 376 bp and contained three exons. The positions of two introns were conserved between the cystatin genes of trematodes and their vertebrate hosts. Reverse transcription polymerase chain reaction confirmed the transcription of SjCystatin in the egg, schistosomula and adult stages of S. japonicum. The encoding ORF region was cloned into pET-28a (+) prokaryotic expression vector. After purification, the recombinant protein SjCystatin (recSjCystatin), expressed in Escherichia coli, was used to immunize animals and produce its specific polyclonal antibody. Western blot analysis revealed that the native SjCystatin was expressed in the egg and adult stages. The enzyme activity assay of the recSjCystatin showed that it inhibited the proteolytic activity of papain. SjCystatin protein was mainly localized on the miracidium within eggs. Immunohistochemistry revealed that SjCystatin mainly localized in the epithelial cells lining the gut as well as the tegument on the surface of adult worms. The conserved genomic DNA structure among cystatin homologues of trematode and their vertebrate host emphasized the characteristics of compatibility between parasites and their hosts. This study provides the first insight into the gene and protein of S. japonicum cystatin and a basis for a further understanding the functions of this gene.