Internal carotid artery injury during endoscopic transsphenoidal pituitary surgery: risk factors, management.

BACKGROUND: Transsphenoidal surgeons should try to avoid internal carotid artery (ICA) injury but also be prepared to manage it. We analyzed our experience with ICA injury during endoscopic transsphenoidal pituitary surgery and present associated risk factors and a management protocol. METHODS: We retrospectively reviewed and analyzed the medical records of 1596 patients who underwent endoscopic transsphenoidal surgery for pituitary tumor resection in our institution from January 2009 to October 2022. RESULTS: Six patients experienced an ICA injury. All received timely and effective hemostasis with immediate direct tamponade followed by endovascular treatment. No serious postoperative complications occurred. CONCLUSIONS: We proposed a treatment plan for ICA injuries encountered during endoscopic transsphenoidal surgery and described our hemostasis process, methods of endovascular treatment, and means of postoperative follow-up in detail.

The Study on the Morphology and Compression Properties of Microcellular TPU/Nanoclay Tissue Scaffolds for Potential Tissue Engineering Applications.

Thermoplastic polyurethane (TPU) materials have shown promise in tissue engineering applications due to their mechanical properties and biocompatibility. However, the addition of nanoclays to TPU can further enhance its properties. In this study, the effects of nanoclays on the microstructure, mechanical behavior, cytocompatibility, and proliferation of TPU/nanoclay (TPUNC) composite scaffolds were comprehensively investigated. The dispersion morphology of nanoclays within the TPU matrix was examined using transmission electron microscopy (TEM). It was found that the nanoclays exhibited a well-dispersed and intercalated structure, which contributed to the improved mechanical properties of the TPUNC scaffolds. Mechanical testing revealed that the addition of nanoclays significantly enhanced the compressive strength and elastic resilience of the TPUNC scaffolds. Cell viability and proliferation assays were conducted using MG63 cells cultured on the TPUNC scaffolds. The incorporation of nanoclays did not adversely affect cell viability, as evidenced by the comparable cell numbers between nanoclay-filled and unfilled TPU scaffolds. The presence of nanoclays within the TPUNC scaffolds did not disrupt cell adhesion or proliferation. The incorporation of nanoclays improved the dispersion morphology, enhanced mechanical performance, and maintained excellent biocompatibility. These findings suggest that TPUNC composites have great potential for tissue engineering applications, providing a versatile and promising scaffold material for regenerative medicine.

The incremental significance of heart rate recovery as a predictor during exercise-stress myocardial perfusion SPECT imaging in individuals with suspected coronary artery disease.

Background: Major adverse cardiac events (MACE) are more likely to occur when abnormal heart rate recovery (HRR). This study aimed to assess the incremental predictive significance of HRR over exercise stress myocardial perfusion single-photon emission computed tomography (MPS) results for MACE in individuals with suspected coronary artery disease (CAD). Methods: Between January 2014 and December 2017, we continually gathered data on 595 patients with suspected CAD who received cycling exercise stress MPS. HRR at 1, 2, 3, and 4 min were used as study variables to obtain the optimal cut-off values of HRR for MACE. The difference between the peak heart rate achieved during exercise and the heart rate at 1, 2, 3, and 4 min was used to calculate the HRR, as shown in HRR3. Heart rate variations between two locations in time, such as HRR2 min-1 min, were used to establish the slope of HRR. All patients were followed for a minimum of 4 years, with MACE as the follow-up goal. The associations between HRR and MACE were assessed using Cox proportional hazards analyses. Results: Patients with MACE were older (P = 0.001), and they also had higher rates of hypertension, dyslipidemia, diabetes, abnormal MPS findings (SSS ≥ 5%), medication history (all P < 0.001), and lower HRR values (all P < 0.01). Patients with and without MACE did not significantly vary in their HRR4 min-3 min. The optimal cut-off of HRR1, 2, and 3 combined with SSS can stratify the risk of MACE in people with suspected CAD (all P < 0.001). HRR 1, 2, and 3 and its slope were linked to MACE in multivariate analysis, where HRR3 was the most significant risk predictor. With a global X2 increase from 101 to 126 (P < 0.0001), HRR3 demonstrated the greatest improvement in the model's predictive capacity, incorporating clinical data and MPS outcomes. Conclusions: HRR at 3 min has a more excellent incremental prognostic value for predicting MACE in patients with suspected CAD following cycling exercise stress MPS. Therefore, incorporating HRR at 3 min into known predictive models may further improve the risk stratification of the patients.

Prognostic value of summed motion score assessed by gated SPECT myocardial perfusion imaging in patients with dilated cardiomyopathy.

Background: The prognosis of patients with dilated cardiomyopathy (DCM) is poor and new indicators are urgently needed to predict lethal cardiac events. This study aimed to investigate the value of summed motion score (SMS) in predicting cardiac death of DCM patients using gated single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI). Methods and results: Eighty-one patients with DCM who underwent 99mTc-MIBI gated SPECT MPI were retrospectively enrolled and were divided into cardiac death and survivor groups. The functional parameters of left ventricle including SMS were measured using quantitative gated SPECT software. During the follow-up period of 44 (25, 54) months, 14 (17.28%) cardiac deaths were observed. Compared with the survivor group, SMS was significantly higher in the cardiac death group. Multivariate cox regression analysis showed that SMS was an independent predictor for cardiac death (HR 1.34, 95% CI 1.02-1.77, P = 0.034). SMS also provided incremental prognostic value over other variables in the multivariate model as determined by likelihood ratio global chi-squared test. In the Kaplan-Meier survival analysis, the event-free survival rate was significantly lower in the high-SMS (HSMS) group than the low-SMS (LSMS) (log-rank P < 0.001). Furthermore, the area under curve (AUC) of SMS was larger than that of LVEF at the 12th month of follow-up (0.85 vs. 0.80, P = 0.045). Conclusion: SMS is an independent predictor of cardiac death in DCM patients and provides incremental prognostic value. SMS might have higher predictive value than LVEF for early cardiac death.

Effects of expandable graphite on the flame-retardant and mechanical performances of rigid polyurethane foams.

Polyurethane foams (PUFs) are found everywhere in our daily life, but they suffer from poor fire resistance. In this study, expansible graphite (EG) as flame retardant was incorporated into PUFs to improve material fire resistance. With the presence of EGs in the PU matrix, bubble size in PUF became smaller as confirmed by the scanning electron microscopy. The mass density of PUFs is directly proportional to the content of EG additive. The compression strengths of EG0/PUF and EG30/PUF decrease from 0.51 MPa to 0.29 MPa. The Fourier transform infrared spectroscopy (FTIR) analysis of RPUFs showed that the addition of EGs did not change the functional group structures of RPUFs. Thermo-gravimetric analysis (TGA) testing results showed that the carbon residue weight of EG30/PUF is higher than other PU composite foams. The combination of TGA and FTIR indicated that the EG addition did not change the thermal decomposition products of EG0/PUF, but effectively inhibited its thermal decomposition rate. Cone calorimeter combustion tests indicated that the peak of the heat release rate of EG30/PUF significantly decreased to 100.5 kW m-2compared to 390.6 kW m-2for EG0/PUF. The ignition time of EG/PUF composites also increased from 2 s to 11 s with incorporation of 30 wt% EGs. The limiting oxygen index (LOI) and UL-94 standard tests show that the LOI of EG30/PUF can reach 55 vol%, and go through V-0 level. This study showed that adding EG into PU foams could significantly improve the thermal stability and flame retardancy properties of EG/PUF composites without significantly sacrificing material compression strength. The research results provide useful guidelines on industrial production and applications of PUFs.

Metal-Organic Framework-Derived Fe-Doped Co1.11 Te2 Embedded in Nitrogen-Doped Carbon Nanotube for Water Splitting.

A rational design is reported of Fe-doped cobalt telluride nanoparticles encapsulated in nitrogen-doped carbon nanotube frameworks (Fe-Co1.11 Te2 @NCNTF) by tellurization of Fe-etched ZIF-67 under a mixed H2 /Ar atmosphere. Fe-doping was able to effectively modulate the electronic structure of Co1.11 Te2 , increase the reaction activity, and further improve the electrochemical performance. The optimized electrocatalyst exhibited superior hydrogen evolution reaction (HER) and oxygen evolution reaction (OER) performances in an alkaline electrolyte with low overpotentials of 107 and 297 mV with a current density of 10 mA cm-2 , in contrast to the undoped Co1.11 Te2 @NCNTF (165 and 360 mV, respectively). The overall water splitting performance only required a voltage of 1.61 V to drive a current density of 10 mA cm-2 . Density function theory (DFT) calculations indicated that the Fe-doping not only afforded abundant exposed active sites but also decreased the hydrogen binding free energy. This work provided a feasible way to study non-precious-metal catalysts for an efficient overall water splitting.

Dissection of Chemical Composition and Associated Gene Expression in the Pigment-Deficient Tea Cultivar 'Xiaoxueya' Reveals an Albino Phenotype and Metabolite Formation.

The tea cultivar 'Xiaoxueya', a temperature-sensitive albino mutant, is a rare tea germplasm because of its highly enriched amino acid content and brisk flavour. In comparison with green leaf tissues of 'Xiaoxueya', albino leaves show significant deficiency in chlorophylls and carotenoids and severely disrupted chloroplasts. Furthermore, the accumulation of quality-related secondary metabolites is altered in 'Xiaoxueya' albino leaf, with significantly increased contents of total amino acids, theanine, and glutamic acid and significantly decreased contents of alkaloids, catechins, and polyphenols. To uncover the molecular mechanisms underlying albinism and quality-related constituent variation in 'Xiaoxueya' leaves, expression profiles of pivotal genes involved in the biosynthetic pathways of pigments, caffeine, theanine, and catechins were investigated by quantitative real-time PCR technology. The results revealed that suppressed expression of the chloroplast-localized 1-deoxy-D-xylulose-5-phosphate synthase genes DXS1 and DXS2 involved in the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway and protochlorophyllide oxidoreductase genes POR1 and POR2 involved in the chlorophyll biosynthetic pathway is responsible for the pigment deficiency in 'Xiaoxueya' albino leaf. Additionally, the low expression of the tea caffeine synthase gene (TCS) involved in caffeine biosynthesis and the chalcone synthase genes CHS1, CHS2, and CHS3, the chalcone isomerase gene CHI, the flavonoid 3',5'-hydroxylase genes F3'5'H1 and F3'5'H2, and the anthocyanidin reductase genes ANR1 and ANR2 involved in the flavonoid pathway is related to the reduction in alkaloid and catechin levels in 'Xiaoxueya' albino leaves.

Genome-wide identification and characterization of ALTERNATIVE OXIDASE genes and their response under abiotic stresses in Camellia sinensis (L.) O. Kuntze.

MAIN CONCLUSION: Four typical ALTERNATIVE OXIDASE genes have been identified in tea plants, and their sequence features and gene expression profiles have provided useful information for further studies on function and regulation. Alternative oxidase (AOX) is a terminal oxidase located in the respiratory electron transport chain. AOX catalyzes the oxidation of quinol and the reduction of oxygen into water. In this study, a genome-wide search and subsequent DNA cloning were performed to identify and characterize AOX genes in tea plant (Camellia sinensis (L.) O. Kuntze cv. Longjing43). Our results showed that tea plant possesses four AOX genes, i.e., CsAOX1a, CsAOX1d, CsAOX2a and CsAOX2b. Gene structure and protein sequence analyses revealed that all CsAOXs share a four-exon/three-intron structure with highly conserved regions and amino acid residues, which are necessary for AOX secondary structures, catalytic activities and post-translational regulations. All CsAOX were shown to localize in mitochondria using the green fluorescent protein (GFP)-targeting assay. Both CsAOX1a and CsAOX1d were induced by cold, salt and drought stresses, and with different expression patterns in young and mature leaves. Reactive oxygen species (ROS) accumulated strongly after 72 and 96 h cold treatments in both young and mature leaves, while the polyphenol and total catechin decreased significantly only in mature leaves. In comparison to AtAOX1a in Arabidopsis thaliana, CsAOX1a lost almost all of the stress-responsive cis-acting regulatory elements in its promoter region (1500 bp upstream), but possesses a flavonoid biosynthesis-related MBSII cis-acting regulatory element. These results suggest a link between CsAOX1a function and the metabolism of some secondary metabolites in tea plant. Our studies provide a basis for the further elucidation of the biological function and regulation of the AOX pathway in tea plants.

Transcriptome sequencing dissection of the mechanisms underlying differential cold sensitivity in young and mature leaves of the tea plant (Camellia sinensis).

The tea plant originated in tropical and subtropical regions and experiences considerable challenges during cold winters and late spring frosts. After short-term chilling stress, young leaves of tea plants exhibit browning, a significant increase in electrolyte leakage and a marked decrease in the maximal photochemical efficiency of photosystem II (Fv/Fm) compared with mature leaves. To identify the mechanisms underlying the different chilling tolerance between young and mature leaves of the tea plant, we used Illumina RNA-Seq technology to analyse the transcript expression profiles of young and mature leaves exposed to temperatures of 20 °C, 4 °C, and 0 °C for 4 h. A total of 45.70-72.93 million RNA-Seq raw reads were obtained and then de novo assembled into 228,864 unigenes with an average length of 601 bp and an N50 of 867 bp. In addition, the differentially expressed unigenes were identified via Venn diagram analyses for paired comparisons of young and mature leaves. Functional classifications based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses revealed that the up-regulated differentially expressed genes were predominantly related to the cellular component terms of chloroplasts and cell membranes, the biological process term of oxidation-reduction process as well as the pathway terms of glutathione metabolism and photosynthesis, suggesting that these components and pathways may contribute to the cold hardiness of mature leaves. Conversely, the inhibited expression of genes related to cell membranes, carotenoid metabolism, photosynthesis, and ROS detoxification in young leaves under cold conditions might lead to the disintegration of cell membranes and oxidative damage to the photosynthetic apparatus. Further quantitative real-time PCR testing validated the reliability of our RNA-Seq results. This work provides valuable information for understanding the mechanisms underlying the cold susceptibility of young tea plant leaves and for breeding tea cultivars with superior frost resistance via the genetic manipulation of antioxidant enzymes.

Lentivirus­mediated RIG­I knockdown relieves cell proliferation inhibition, cell cycle arrest and apoptosis in ATRA­induced NB4 cells via the AKT­FOXO3A signaling pathway in vitro.

Retinoic acid inducible gene I (RIG­I) is upregulated during all­trans retinoic acid (ATRA)­induced terminal granulocytic differentiation of NB4 acute promyelocytic leukemia (APL) cells. However, the function and mechanism of RIG­I in NB4 cells remains to be fully elucidated. In the present study, lentivirus­mediated RIG­I­knockdown was used to investigate the proliferation, cell cycle and apoptotic processes of ATRA­induced NB4 cells in vitro using an MTT assay and flow cytometry, respectively. The roles of RIG­I and the AKT­FOXO3A signaling pathway were investigated using western blot analysis. The results showed that the ATRA­induced expression of RIG­I was specifically and effectively knocked down at the mRNA and protein levels by lentivirus mediated RIG­I short hairpin RNA. In addition, silencing of RIG­I reduced the ATRA­induced inhibition of NB4 cell proliferation, cell cycle arrest and apoptosis. Further investigations indicated that with ATRA­induced expression of RIG­I, levels of phosphorylated (p)AKT­Thr308 and pForkhead Box (FOX) O3A­Thr32 were decreased, the expression levels of cell cycle arrest protein p27 and the apoptotic protein, tumor necrosis factor­related apoptosis­inducing ligand (TRAIL), directly transcribed by FOXO3A were increased. By contrast, following the knockdown of ATRA­induced expression of RIG­I, the levels of pAKT­Thr308 and pFOXO3A­Thr32 were increased, and the protein expression levels of p27 and TRAIL were decreased. Taken together, these results showed that the knockdown of RIG­I reduced the inhibition of cell proliferation, cell cycle arrest and apoptosis in the ATRA­induced NB4 cells via the AKT­FOXO3A signaling pathway.

Isolation and expression features of hexose kinase genes under various abiotic stresses in the tea plant (Camellia sinensis).

Hexokinases (HXKs, EC 2.7.1.1) and fructokinases (FRKs, EC 2.7.1.4) play important roles in carbohydrate metabolism and sugar signaling during the growth and development of plants. However, the HXKs and FRKs in the tea plant (Camellia sinensis) remain largely unknown. In this manuscript, we present the molecular characterization, phylogenetic relationships, conserved domains and expression profiles of four HXK and seven FRK genes of the tea plant. The 11 deduced CsHXK and CsFRK proteins were grouped into six main classes. All of the deduced proteins, except for CsFKR7, possessed putative ATP-binding motifs and a sugar recognition region. These genes exhibited tissue-specific expression patterns, which suggests that they play different roles in the metabolism and development of source and sink tissues in the tea plant. There were variations in CsHXKs and CsFRKs transcript abundance in response to four abiotic stresses: cold, salt, drought and exogenous abscisic acid (ABA). Remarkably, CsHXK3 and CsHXK4 were significantly induced in the leaves and roots under cold conditions, CsHXK1 was apparently up-regulated in the leaves and roots under salt and drought stresses, and CsHXK3 was obviously stimulated in the leaves and roots under short-term treatment with exogenous ABA. These findings demonstrate that CsHXKs play critical roles in response to abiotic stresses in the tea plant. Our research provides a fundamental understanding of the CsHXK and CsFRK genes of the tea plant and important information for the breeding of stress-tolerant tea cultivars.

Diverse Colletotrichum species cause anthracnose of tea plants (Camellia sinensis (L.) O. Kuntze) in China.

Anthracnose caused by Colletotrichum is one of the most severe diseases that can afflict Camellia sinensis. However, research on the diversity and geographical distribution of Colletotrichum in China remain limited. In this study, 106 Colletotrichum isolates were collected from diseased leaves of Ca. sinensis cultivated in the 15 main tea production provinces in China. Multi-locus phylogenetic analysis coupled with morphological identification showed that the collected isolates belonged to 11 species, including 6 known species (C. camelliae, C. cliviae, C. fioriniae, C. fructicola, C. karstii, and C. siamense), 3 new record species (C. aenigma, C. endophytica, and C. truncatum), 1 novel species (C. wuxiense), and 1 indistinguishable strain, herein described as Colletotrichum sp. Of these species, C. camelliae and C. fructicola were the dominant species causing anthracnose in Ca. sinensis. In addition, our study provided further evidence that phylogenetic analysis using a combination of ApMat and GS sequences can be used to effectively resolve the taxonomic relationships within the C. gloeosporioides species complex. Finally, pathogenicity tests suggested that C. camelliae, C. aenigma, and C. endophytica are more invasive than other species after the inoculation of the leaves of Ca. sinensis.

Metabolic Changes of Caffeine in Tea Plant (Camellia sinensis (L.) O. Kuntze) as Defense Response to Colletotrichum fructicola.

Tea plant (Camellia sinensis) is one of the most economically valuable crops in the world. Anthracnose can affect the growth of leaves and cause serious yield losses of tea. Tea plants are rich in secondary metabolites; however, their roles in resistance to anthracnose are unclear. Herein we compared the contents of total phenolics, catechins, and caffeine in two cultivars with different resistances to anthracnose during Colletotrichum fructicola infection. (-)-Epigallocatechin-3-gallate (EGCG), (+)-catechin (C), caffeine, and critical regulatory genes were induced in C. fructicola-resistant tissues. In vitro antifungal tests showed that caffeine more strongly inhibited mycelial growth than tea polyphenols and catechins. Both electron microscopy and bioactivity analysis results showed that caffeine can affect mycelial cell walls and plasma membranes. Through promoter sequences analysis, a number of stress response-related cis-acting elements were identified in S-adenosylmethionine synthetase and tea caffeine synthase. These results demonstrated that (-)-EGCG, (+)-C, and caffeine may be involved in the resistance of tea plants to anthracnose.

Effects of cold acclimation on sugar metabolism and sugar-related gene expression in tea plant during the winter season.

Sugar plays an essential role in plant cold acclimation (CA), but the interaction between CA and sugar remains unclear in tea plants. In this study, during the whole winter season, we investigated the variations of sugar contents and the expression of a large number of sugar-related genes in tea leaves. Results indicated that cold tolerance of tea plant was improved with the development of CA during early winter season. At this stage, starch was dramatically degraded, whereas the content of total sugars and several specific sugars including sucrose, glucose and fructose were constantly elevated. Beyond the CA stage, the content of starch was maintained at a low level during winter hardiness (WH) period and then was elevated during de-acclimation (DC) period. Conversely, the content of sugar reached a peak at WH stage followed by a decrease during DC stage. Moreover, gene expression results showed that, during CA period, sugar metabolism-related genes exhibited different expression pattern, in which beta-amylase gene (CsBAM), invertase gene (CsINV5) and raffinose synthase gene (CsRS2) engaged in starch, sucrose and raffinose metabolism respectively were solidly up-regulated; the expressions of sugar transporters were stimulated in general except the down-regulations of CsSWEET2, 3, 16, CsERD6.7 and CsINT2; interestingly, the sugar-signaling related CsHXK3 and CsHXK2 had opposite expression patterns at the early stage of CA. These provided comprehensive insight into the effects of CA on carbohydrates indicating that sugar accumulation contributes to tea plant cold tolerance during winter season, and a simply model of sugar regulation in response to cold stimuli is proposed.

Global transcriptome and gene regulation network for secondary metabolite biosynthesis of tea plant (Camellia sinensis).

BACKGROUND: Major secondary metabolites, including flavonoids, caffeine, and theanine, are important components of tea products and are closely related to the taste, flavor, and health benefits of tea. Secondary metabolite biosynthesis in Camellia sinensis is differentially regulated in different tissues during growth and development. Until now, little was known about the expression patterns of genes involved in secondary metabolic pathways or their regulatory mechanisms. This study aimed to generate expression profiles for C. sinensis tissues and to build a gene regulation model of the secondary metabolic pathways. RESULTS: RNA sequencing was performed on 13 different tissue samples from various organs and developmental stages of tea plants, including buds and leaves of different ages, stems, flowers, seeds, and roots. A total of 43.7 Gbp of raw sequencing data were generated, from which 347,827 unigenes were assembled and annotated. There were 46,693, 8446, 3814, 10,206, and 4948 unigenes specifically expressed in the buds and leaves, stems, flowers, seeds, and roots, respectively. In total, 1719 unigenes were identified as being involved in the secondary metabolic pathways in C. sinensis, and the expression patterns of the genes involved in flavonoid, caffeine, and theanine biosynthesis were characterized, revealing the dynamic nature of their regulation during plant growth and development. The possible transcription factor regulation network for the biosynthesis of flavonoid, caffeine, and theanine was built, encompassing 339 transcription factors from 35 families, namely bHLH, MYB, and NAC, among others. Remarkably, not only did the data reveal the possible critical check points in the flavonoid, caffeine, and theanine biosynthesis pathways, but also implicated the key transcription factors and related mechanisms in the regulation of secondary metabolite biosynthesis. CONCLUSIONS: Our study generated gene expression profiles for different tissues at different developmental stages in tea plants. The gene network responsible for the regulation of the secondary metabolic pathways was analyzed. Our work elucidated the possible cross talk in gene regulation between the secondary metabolite biosynthetic pathways in C. sinensis. The results increase our understanding of how secondary metabolic pathways are regulated during plant development and growth cycles, and help pave the way for genetic selection and engineering for germplasm improvement.

Shish-kebab-structured poly(ε-caprolactone) nanofibers hierarchically decorated with chitosan-poly(ε-caprolactone) copolymers for bone tissue engineering.

In this work, scaffolds with a shish-kebab (SK) structure formed by poly(ε-caprolactone) (PCL) nanofibers and chitosan-PCL (CS-PCL) copolymers were prepared via electrospinning and subsequent crystallization for bone tissue engineering applications. The aim of this study was to introduce nanosized topography and the high biocompatibility of chitosan onto PCL nanofibers to enhance cell affinity to PCL scaffolds. CS-PCL copolymers with various ratios were synthesized, and then spontaneously crystallized as kebabs onto the electrospun PCL fibers, which acted as shishes. Scanning electron microscopy (SEM) results demonstrated that the copolymer with PCL to chitosan ratio of 8.8 could hierarchically decorate the PCL nanofibers and formed well-shaped kebabs on the PCL nanofiber surface. Water contact angle tests and biomimetic activity experiments revealed that the shish-kebab scaffolds with CS-PCL kebabs (PCL-SK(CS-PCL(8.8))) showed enhanced hydrophilicity and mineralization ability compared with smooth PCL and PCL-SK(PCL) shish-kebab scaffolds. Osteoblast-like MG63 cells cultured on the PCL-SK(CS-PCL(8.8)) scaffolds showed optimizing cell attachment, cell viability, and metabolic activity, demonstrating that this kind of scaffold has potential applications in bone tissue engineering.

Characterization of a novel antimicrobial peptide with chitin-biding domain from Mytilus coruscus.

Using reverse phase high performance liquid chromatography (RP-HPLC), a novel antimicrobial peptide with 55 amino acid residues was isolated from the hemolymph of Mytilus coruscus. This new antimicrobial peptide displays predominant antimicrobial activity against fungi and Gram-positive bacteria. The molecular mass and the N-terminal sequence of this peptide were analyzed by Mass Spectrometry and Edman degradation, respectively. This antimicrobial peptide, with molecular mass of 6621.55 Da, is characterized by a chitin-biding domain and by 6 Cysteine residues engaged in three intra-molecular disulfide bridges. The full-length of cDNA sequence of this new peptide was obtained by rapid amplification of cDNA ends (RACE) and the encoded precursor was turn out to be a chitotriosidase-like protein. Therefore, we named the precursor with mytichitin-1 and the new antimicrobial peptide (designated as mytichitin-CB) is the carboxyl-terminal part of mytichitin-1. The mRNA transcripts of mytichitin-1 are mainly detected in gonad and the expression level of mytichitin-1 in gonad was up-regulated and reached the highest level at 12 h after bacterial challenge, which was 9-fold increase compared to that of the control group. These results indicated that mytichitin-1 was involved in the host immune response against bacterial infection and might contribute to the clearance of invading bacteria.

Construction of a SSR-based genetic map and identification of QTLs for catechins content in tea plant (Camellia sinensis).

Catechins are the most important bioactive compounds in tea, and have been demonstrated to possess a wide variety of pharmacological activities. To characterize quantitative trait loci (QTLs) for catechins content in the tender shoots of tea plant, we constructed a moderately saturated genetic map using 406 simple sequence repeat (SSR) markers, based on a pseudo-testcross population of 183 individuals derived from an intraspecific cross of two Camellia sinensis varieties with diverse catechins composition. The map consisted of fifteen linkage groups (LGs), corresponding to the haploid chromosome number of tea plant (2n = 2x = 30). The total map length was 1,143.5 cM, with an average locus spacing of 2.9 cM. A total of 25 QTLs associated with catechins content were identified over two measurement years. Of these, nine stable QTLs were validated across years, and clustered into four main chromosome regions on LG03, LG11, LG12 and LG15. The population variability explained by each QTL was predominantly at moderate-to-high levels and ranged from 2.4% to 71.0%, with an average of 17.7%. The total number of QTL for each trait varied from four to eight, while the total population variability explained by all QTLs for a trait ranged between 38.4% and 79.7%. This is the first report on the identification of QTL for catechins content in tea plant. The results of this study provide a foundation for further cloning and functional characterization of catechin QTLs for utilization in improvement of tea plant.

Global transcriptome profiles of Camellia sinensis during cold acclimation.

BACKGROUND: Tea is the most popular non-alcoholic health beverage in the world. The tea plant (Camellia sinensis (L.) O. Kuntze) needs to undergo a cold acclimation process to enhance its freezing tolerance in winter. Changes that occur at the molecular level in response to low temperatures are poorly understood in tea plants. To elucidate the molecular mechanisms of cold acclimation, we employed RNA-Seq and digital gene expression (DGE) technologies to the study of genome-wide expression profiles during cold acclimation in tea plants. RESULTS: Using the Illumina sequencing platform, we obtained approximately 57.35 million RNA-Seq reads. These reads were assembled into 216,831 transcripts, with an average length of 356 bp and an N50 of 529 bp. In total, 1,770 differentially expressed transcripts were identified, of which 1,168 were up-regulated and 602 down-regulated. These include a group of cold sensor or signal transduction genes, cold-responsive transcription factor genes, plasma membrane stabilization related genes, osmosensing-responsive genes, and detoxification enzyme genes. DGE and quantitative RT-PCR analysis further confirmed the results from RNA-Seq analysis. Pathway analysis indicated that the "carbohydrate metabolism pathway" and the "calcium signaling pathway" might play a vital role in tea plants' responses to cold stress. CONCLUSIONS: Our study presents a global survey of transcriptome profiles of tea plants in response to low, non-freezing temperatures and yields insights into the molecular mechanisms of tea plants during the cold acclimation process. It could also serve as a valuable resource for relevant research on cold-tolerance and help to explore the cold-related genes in improving the understanding of low-temperature tolerance and plant-environment interactions.

Streptamer versus tetramer-based selection of functional cytomegalovirus-specific T cells.

BACKGROUND/PURPOSE: Cytomegalovirus (CMV) disease constitutes a serious complication after stem cell transplantation and has been treated by adoptive transfer of donor-derived CMV-specific CD8(+) T cells. CMV-specific CD8(+) T cells were selected by multimers, and the technologies may alter the function of these T cells. Therefore, here we evaluated the impact of multimer reagents on the function of CD8(+) T lymphocytes. METHODS: CMV-specific CD8(+) T cells were purified from the peripheral blood of donors using tetra- and streptamer technologies. The functional status of purified CMV-specific CD8(+) T cells was assessed by multiparametric immunophenotyping and carboxyfluorescein succinimidyl ester proliferation assays as well as by enzyme-linked immunospot assays. RESULTS: A similar percentage of CMV-specific CD8(+) T cells could be purified by both tetra- (90%) and streptamer (92%) technologies. That constitutes a 30- to 50-fold concentration of CMV-specific CD8(+)CD45RA(+)CCR7-effector T cells. Selected cells secreted interferon-gamma and granzyme B upon stimulation with CMVpp65 peptide, thus demonstrating their functionality. CONCLUSION: Our study demonstrated that both tetra- and streptamer technologies can be used to purify CMV-specific cytotoxic CD8(+) T cells for adoptive T-cell transfer. Both multimer technologies did not have any negative influence on the proliferation of selected T cells. Importantly, streptamer technology is available at good manufacturing practice level.