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1.  Metastasis of distal esophageal carcinoma to the thyroid with presentation simulating primary thyroid carcinoma: a case report and review of the literature 
Metastasis to the thyroid is extremely rare. There is a lack of awareness of and adequate preparation for this situation, especially in an individual without a past history of malignancy. We describe a rare case of a 61-year-old man in whom a primary distal esophageal carcinoma gave rise to a metastatic palpable mass in the thyroid gland. Palliative bilateral near-total thyroidectomy was performed with pathology showing squamous cell carcinoma and tracheostomy was carried out simultaneously due to airway compression with related symptoms. A review of the literature only reveals 4 similar cases. Secondary neoplasm of the thyroid mimicking a primary malignant lesion is seldom encountered, however, in order to make appropriate treatment, the most critical problem is to distinguish the difference between the above two and the final diagnosis can only be confirmed on pathologic examination. Although the prognosis of thyroid metastasis is commonly felt to be poor, improvement of living quality and prolongation of survival may be obtained in such patients through correct diagnosis and treatment.
doi:10.1186/1477-7819-12-106
PMCID: PMC4001107  PMID: 24755371
Thyroid metastasis; esophageal neoplasms; squamous cell carcinoma
2.  The Outcome of Ipsilateral Hemihepatectomy in Mucin-Producing Bile Duct Tumors 
PLoS ONE  2014;9(4):e92010.
Background
Mucin-producing bile duct tumors (MPBTs) are unusual, and we present our experience with nine surgically proven cases.
Methods
Between November 2002 and November 2012, 9 patients with surgically proven MPBTs (including history of relevant hepatobiliary surgery in 6 patients) were encountered. Their clinical, imaging, and surgical findings were reviewed.
Results
The most common symptom is intermittent jaundice, which occurs in seven patients. The diagnostic specificity was 77.8% by preoperative Magnetic Resonance Cholangiopancreatography (MRCP). All the patients underwent ipsilateral hemihepatectomy or remnant hemihepatectomy, accompanied caudate lobectomy in one case and concomitant Roux-en-Y choledochojejunostomy in four cases. Postoperative course was uneventful. One patient, who had intra-abdominal recurrence 59 months after surgery, was received reoperation without recurrence at the last follow-up. The remaining eight patients were alive without disease recurrence.
Conclusion
Based on our follow up of 9 cases that were surgically treated for MPBTs, we conclude that ipsilateral hemihepatectomy is a safe surgical procedure with an observed recurrence risk of 11.1% and all long-term survival.
doi:10.1371/journal.pone.0092010
PMCID: PMC3984073  PMID: 24727803
3.  Draft Genome Sequence of Clostridium ultunense Strain BS (DSMZ 10521), Recovered from a Mixed Culture 
Genome Announcements  2014;2(1):e01269-13.
Clostridium ultunense BS is the first isolated strain (type strain) of C. ultunense that was identified as a mesophilic syntrophic acetate-oxidizing bacterium (SAOB). Here, we report the draft genome sequence of this strain, which will help us to elucidate the mechanism of syntrophic acetate oxidization.
doi:10.1128/genomeA.01269-13
PMCID: PMC3916497  PMID: 24504003
4.  Proteome and Phosphoproteome Characterization Reveals New Response and Defense Mechanisms of Brachypodium distachyon Leaves under Salt Stress* 
Salinity is a major abiotic stress affecting plant growth and development. Understanding the molecular mechanisms of salt response and defense in plants will help in efforts to improve the salt tolerance of crops. Brachypodium distachyon is a new model plant for wheat, barley, and several potential biofuel grasses. In the current study, proteome and phosphoproteome changes induced by salt stress were the focus. The Bd21 leaves were initially treated with salt in concentrations ranging from 80 to 320 mm and then underwent a recovery process prior to proteome analysis. A total of 80 differentially expressed protein spots corresponding to 60 unique proteins were identified. The sample treated with a median salt level of 240 mm and the control were selected for phosphopeptide purification using TiO2 microcolumns and LC-MS/MS for phosphoproteome analysis to identify the phosphorylation sites and phosphoproteins. A total of 1509 phosphoproteins and 2839 phosphorylation sites were identified. Among them, 468 phosphoproteins containing 496 phosphorylation sites demonstrated significant changes at the phosphorylation level. Nine phosphorylation motifs were extracted from the 496 phosphorylation sites. Of the 60 unique differentially expressed proteins, 14 were also identified as phosphoproteins. Many proteins and phosphoproteins, as well as potential signal pathways associated with salt response and defense, were found, including three 14-3-3s (GF14A, GF14B, and 14-3-3A) for signal transduction and several ABA signal-associated proteins such as ABF2, TRAB1, and SAPK8. Finally, a schematic salt response and defense mechanism in B. distachyon was proposed.
doi:10.1074/mcp.M113.030171
PMCID: PMC3916659  PMID: 24335353
5.  Effect of Change in Spindle Structure on Proliferation Inhibition of Osteosarcoma Cells and Osteoblast under Simulated Microgravity during Incubation in Rotating Bioreactor 
PLoS ONE  2013;8(10):e76710.
In order to study the effect of microgravity on the proliferation of mammalian osteosarcoma cells and osteoblasts, the changes in cell proliferation, spindle structure, expression of MAD2 or BUB1, and effect of MAD2 or BUB1 on the inhibition of cell proliferation is investigated by keeping mammalian osteosarcoma cells and osteoblasts under simulated microgravity in a rotating wall vessel (2D-RWVS) bioreactor. Experimental results indicate that the effect of microgravity on proliferation inhibition, incidence of multipolar spindles, and expression of MAD2 or BUB1 increases with the extension of treatment time. And multipolar cells enter mitosis after MAD2 or BUB1 is knocked down, which leads to the decrease in DNA content, and decrease the accumulation of cells within multipolar spindles. It can therefore be concluded that simulated microgravity can alter the structure of spindle microtubules, and stimulate the formation of multipolar spindles together with multicentrosomes, which causes the overexpression of SAC proteins to block the abnormal cells in metaphase, thereby inhibiting cell proliferation. By clarifying the relationship between cell proliferation inhibition, spindle structure and SAC changes under simulated microgravity, the molecular mechanism and morphology basis of proliferation inhibition induced by microgravity is revealed, which will give experiment and theoretical evidence for the mechanism of space bone loss and some other space medicine problems.
doi:10.1371/journal.pone.0076710
PMCID: PMC3792057  PMID: 24116140
6.  The Sex and Sex Determination in Pyropia haitanensis (Bangiales, Rhodophyta) 
PLoS ONE  2013;8(8):e73414.
Pyropia haitanensis has a biphasic life cycle with macroscopic gametophytic blade (n) and microscopic filamentous conchocelis (2n) phase. Its gametophytic blades have long been believed to be mainly dioecious. However, when crossing the red mutant (R, ♀) with the wild type (W, ♂), the parental colors were segregated in F1 blades, of which 96.1% were linearly sectored with 2–4 color sectors. When color sectors were excised from the color-sectored blades and cultured singly, 99.7% of the color sectors appeared to be unisexual with an equal sex ratio. Although the sex of color sector did not genetically link with its color, the boundaries of both sex and color sectors coincided precisely. About 87.9% of the examined color-sectored blades were monoecious and the percentage increased with the number of color sectors of a blade. The gametophytic blades from each conchocelis strain produced by parthenogenesis of the excised color sectors were unisexual and unicolor, showing the same sex and color as their original sectors. These results indicate that most of the sexually reproduced Py. haitanensis blades are monoecious, and their sex is controlled by segregation of a pair of alleles during meiosis of conchospore, forming a sex-sectored tetrad. During the subsequent development of blades, one or two lower cell(s) of the tetrad contribute mainly to rhizoid formation, and rarely show their sexual phenotype, leading to reduced frequency of full sex phenotype of the meiotic blades. Moreover, the aberrant segregations of sex genes or color genes in a few of F1 blades were probably due to gene conversions, but there was no sex transfer in Py. haitanensis.
doi:10.1371/journal.pone.0073414
PMCID: PMC3753276  PMID: 23991194
7.  Complete Genome Sequence of a Novel Deletion Porcine Reproductive and Respiratory Syndrome Virus Strain 
Genome Announcements  2013;1(4):e00486-13.
Porcine reproductive and respiratory syndrome virus HZ-31 strain is different from any other previously sequenced porcine reproductive and respiratory syndrome virus strains. It contains a 59-amino acid (aa) discontinuous deletion in aa 467 to 474, aa 498 to 519, and aa 533 to 561 of nsp2. Here, we report the complete genome sequence of this novel Chinese virulent PRRSV variant.
doi:10.1128/genomeA.00486-13
PMCID: PMC3735069  PMID: 23887914
8.  An In Vivo and In Vitro Evaluation of the Mutual Interactions between the Lung and the Large Intestine 
One of the most important theories of the traditional Chinese medicine is the exterior-interior relationship between the lung and the large intestine; so far, little direct experimental evidence has been reported to support such relationship. Here we for the first time investigated the mutual interactions between the lung and the large intestine by examining the relevancies between the pulmonary functions and the rectal resting pressure in the rat models of asthma and constipation. We also evaluated the effects of the lung homogenate and the large intestine homogenate on the isolated large intestine muscle strip and the isolated tracheal spiral, respectively. Our results showed that the pulmonary resistance and pulmonary compliance were closely related to the rectal resting pressure in the asthmatic rat model, while the rectal resting pressure was much correlated with the pulmonary resistance in the rat model of constipation. Moreover, it was shown that the lung homogenate could specifically contract the isolated large intestine muscle strip. Overall, this study provided new lines of evidence for the theory and highlighted the potential application in the treatment of the corresponding diseases.
doi:10.1155/2013/695641
PMCID: PMC3736455  PMID: 23970934
9.  Metagenomic Insights into Metabolic Capacities of the Gut Microbiota in a Fungus-Cultivating Termite (Odontotermes yunnanensis) 
PLoS ONE  2013;8(7):e69184.
Macrotermitinae (fungus-cultivating termites) are major decomposers in tropical and subtropical areas of Asia and Africa. They have specifically evolved mutualistic associations with both a Termitomyces fungi on the nest and a gut microbiota, providing a model system for probing host-microbe interactions. Yet the symbiotic roles of gut microbes residing in its major feeding caste remain largely undefined. Here, by pyrosequencing the whole gut metagenome of adult workers of a fungus-cultivating termite (Odontotermes yunnanensis), we showed that it did harbor a broad set of genes or gene modules encoding carbohydrate-active enzymes (CAZymes) relevant to plant fiber degradation, particularly debranching enzymes and oligosaccharide-processing enzymes. Besides, it also contained a considerable number of genes encoding chitinases and glycoprotein oligosaccharide-processing enzymes for fungal cell wall degradation. To investigate the metabolic divergence of higher termites of different feeding guilds, a SEED subsystem-based gene-centric comparative analysis of the data with that of a previously sequenced wood-feeding Nasutitermes hindgut microbiome was also attempted, revealing that SEED classifications of nitrogen metabolism, and motility and chemotaxis were significantly overrepresented in the wood-feeder hindgut metagenome, while Bacteroidales conjugative transposons and subsystems related to central aromatic compounds metabolism were apparently overrepresented here. This work fills up our gaps in understanding the functional capacities of fungus-cultivating termite gut microbiota, especially their roles in the symbiotic digestion of lignocelluloses and utilization of fungal biomass, both of which greatly add to existing understandings of this peculiar symbiosis.
doi:10.1371/journal.pone.0069184
PMCID: PMC3714238  PMID: 23874908
10.  EFFECT OF IRRADIATION ON MICROVASCULAR ENDOTHELIAL CELLS OF PAROTID GLANDS IN THE MINIATURE PIG 
Purpose
To evaluate the effect of irradiation on microvascular endothelial cells in miniature pig parotid glands.
Methods and Materials
A single 25-Gy dose of irradiation (IR) was delivered to parotid glands of 6 miniature pigs. Three other animals served as non-IR controls. Local blood flow rate in glands was measured pre- and post-IR with an ultrasonic Doppler analyzer. Samples of parotid gland tissue were taken at 4 h, 24 h, 1 week, and 2 weeks after IR for microvascular density (MVD) analysis and sphingomyelinase (SMase) assay. Histopathology and immunohistochemical staining (anti-CD31 and anti-AQP1) were used to assess morphological changes. MVD was determined by calculating the number of CD31- or AQP1-stained cells per field. A terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis assay was used to detect apoptotic cells. The activity of acid and neutral Mg2+-dependent SMase (ASMase and NSMase, respectively) was also assayed.
Results
Local parotid gland blood flow rate decreased rapidly at 4 h post-IR and remained below control levels throughout the 14-day observation period. Parotid MVD also declined from 4 to 24 hours and remained below control levels thereafter. The activity levels of ASMase and NSMase in parotid glands increased rapidly from 4 to 24 h post-IR and then declined gradually. The frequency of detecting apoptotic nuclei in the glands followed similar kinetics.
Conclusions
Single-dose IR led to a significant reduction of MVD and local blood flow rate, indicating marked damage to microvascular endothelial cells in miniature pig parotid glands. The significant and rapid increases of ASMase and NSMase activity levels may be important in this IR-induced damage.
doi:10.1016/j.ijrobp.2010.05.048
PMCID: PMC3655408  PMID: 20832188
Salivary glands; Radiotherapy; Microvessel density; Endothelial cells; Sphingomyelinase
11.  Long-term strain improvements accumulate mutations in regulatory elements responsible for hyper-production of cellulolytic enzymes 
Scientific Reports  2013;3:1569.
Long-term strain improvements through repeated mutagenesis and screening have generated a hyper-producer of cellulases and hemicellulases from Penicillium decumbens 114 which was isolated 30 years ago. Here, the genome of the hyper-producer P. decumbens JU-A10-T was sequenced and compared with that of the wild-type strain 114-2. Further, the transcriptomes and secretomes were compared between the strains. Selective hyper-production of cellulases and hemicellulases but not all the secreted proteins was observed in the mutant, making it a more specific producer of lignocellulolytic enzymes. Functional analysis identified that changes in several transcriptional regulatory elements played crucial roles in the cellulase hyper-producing characteristics of the mutant. Additionally, the mutant showed enhanced supply of amino acids and decreased synthesis of secondary metabolites compared with the wild-type. The results clearly point out that we can target gene regulators and promoters with minimal alterations of the genetic content but maximal effects in genetic engineering.
doi:10.1038/srep01569
PMCID: PMC3610096  PMID: 23535838
12.  Establishment of transgene-free induced pluripotent stem cells reprogrammed from human stem cells of apical papilla for neural differentiation 
Introduction
Induced pluripotent stem cells (iPSCs) are a potent cell source for neurogenesis. Previously we have generated iPSCs from human dental stem cells carrying transgene vectors. These exogenous transgenes may affect iPSC behaviors and limit their clinical applications. The purpose of this study was to establish transgene-free iPSCs (TF-iPSCs) reprogrammed from human stem cells of apical papilla (SCAP) and determine their neurogenic potential.
Methods
A single lentiviral 'stem cell cassette' flanked by the loxP site (hSTEMCCA-loxP), encoding four human reprogramming factors, OCT4, SOX2, KLF4, and c-MYC, was used to reprogram human SCAP into iPSCs. Generated iPSCs were transfected with plasmid pHAGE2-EF1α-Cre-IRES-PuroR and selected with puromycin for the TF-iPSC subclones. PCR was performed to confirm the excision of hSTEMCCA. TF-iPSC clones did not resist to puromycin treatment indicating no pHAGE2-EF1α-Cre-IRES-PuroR integration into the genome. In vitro and in vivo analyses of their pluripotency were performed. Embryoid body-mediated neural differentiation was undertaken to verify their neurogenic potential.
Results
TF-SCAP iPSCs were generated via a hSTEMCCA-loxP/Cre system. PCR of genomic DNA confirmed transgene excision and puromycin treatment verified the lack of pHAGE2-EF1α-Cre-IRES-PuroR integration. Transplantation of the TF-iPSCs into immunodeficient mice gave rise to teratomas containing tissues representing the three germ layers -- ectoderm (neural rosettes), mesoderm (cartilage and bone tissues) and endoderm (glandular epithelial tissues). Embryonic stem cell-associated markers TRA-1-60, TRA-2-49 and OCT4 remained positive after transgene excision. After neurogenic differentiation, cells showed neural-like morphology expressing neural markers nestin, βIII-tubulin, NFM, NSE, NeuN, GRM1, NR1 and CNPase.
Conclusions
TF-SCAP iPSCs reprogrammed from SCAP can be generated and they may be a good cell source for neurogenesis.
doi:10.1186/scrt134
PMCID: PMC3580434  PMID: 23095454
13.  Transcriptomic Analysis of Neuropeptides and Peptide Hormones in the Barnacle Balanus amphitrite: Evidence of Roles in Larval Settlement 
PLoS ONE  2012;7(10):e46513.
The barnacle Balanus amphitrite is a globally distributed marine crustacean and has been used as a model species for intertidal ecology and biofouling studies. Its life cycle consists of seven planktonic larval stages followed by a sessile juvenile/adult stage. The transitional processes between larval stages and juveniles are crucial for barnacle development and recruitment. Although some studies have been conducted on the neuroanatomy and neuroactive substances of the barnacle, a comprehensive understanding of neuropeptides and peptide hormones remains lacking. To better characterize barnacle neuropeptidome and its potential roles in larval settlement, an in silico identification of putative transcripts encoding neuropeptides/peptide hormones was performed, based on transcriptome of the barnacle B. amphitrite that has been recently sequenced. Potential cleavage sites andstructure of mature peptides were predicted through homology search of known arthropod peptides. In total, 16 neuropeptide families/subfamilies were predicted from the barnacle transcriptome, and 14 of them were confirmed as genuine neuropeptides by Rapid Amplification of cDNA Ends. Analysis of peptide precursor structures and mature sequences showed that some neuropeptides of B. amphitrite are novel isoforms and shared similar characteristics with their homologs from insects. The expression profiling of predicted neuropeptide genes revealed that pigment dispersing hormone, SIFamide, calcitonin, and B-type allatostatin had the highest expression level in cypris stage, while tachykinin-related peptide was down regulated in both cyprids and juveniles. Furthermore, an inhibitor of proprotein convertase related to peptide maturation effectively delayed larval metamorphosis. Combination of real-time PCR results and bioassay indicated that certain neuropeptides may play an important role in cypris settlement. Overall, new insight into neuropeptides/peptide hormones characterized in this study shall provide a platform for unraveling peptidergic control of barnacle larval behavior and settlement process.
doi:10.1371/journal.pone.0046513
PMCID: PMC3462748  PMID: 23056329
14.  Proteome characterization of developing grains in bread wheat cultivars (Triticum aestivum L.) 
BMC Plant Biology  2012;12:147.
Background
The analyses of protein synthesis, accumulation and regulation during grain development in wheat are more complex because of its larger genome size compared to model plants such as Arabidopsis and rice. In this study, grains from two wheat cultivars Jimai 20 and Zhoumai 16 with different gluten quality properties were harvested at five development stages, and were used to displayed variable expression patterns of grain proteins.
Results
Proteome characterization during grain development in Chinese bread wheat cultivars Jimai 20 and Zhoumai 16 with different quality properties was investigated by 2-DE and tandem MALDI-TOF/TOF-MS. Identification of 117 differentially accumulated protein spots representing 82 unique proteins and five main expression patterns enabled a chronological description of wheat grain formation. Significant proteome expression differences between the two cultivars were found; these included 14 protein spots that accumulated in both cultivars but with different patterns and 27 cultivar-different spots. Among the cultivar-different protein spots, 14 accumulated in higher abundance in Jimai 20 than in Zhoumai 16, and included NAD-dependent isocitrate dehydrogenase, triticin precursor, LMW-s glutenin subunit and replication factor C-like protein. These proteins are likely to be associated with superior gluten quality. In addition, some proteins such as class II chitinase and peroxidase 1 with isoforms in developing grains were shown to be phosphorylated by Pro-Q Diamond staining and phosphorprotein site prediction. Phosphorylation could have important roles in wheat grain development. qRT-PCR analysis demonstrated that transcriptional and translational expression patterns of many genes were significantly different.
Conclusions
Wheat grain proteins displayed variable expression patterns at different developmental stages and a considerable number of protein spots showed differential accumulation between two cultivars. Differences in seed storage proteins were considered to be related to different quality performance of the flour from these wheat cultivars. Some proteins with isoforms were phosphorylated, and this may reflect their importance in grain development. Our results provide new insights into proteome characterization during grain development in different wheat genotypes.
doi:10.1186/1471-2229-12-147
PMCID: PMC3480910  PMID: 22900893
Wheat; Grain proteome; Phosphorproteins; 2-DE; Tandem MS; qRT-PCR
15.  Network Identification and Flux Quantification of Glucose Metabolism in Rhodobacter sphaeroides under Photoheterotrophic H2-Producing Conditions 
Journal of Bacteriology  2012;194(2):274-283.
The nonsulfur purple bacteria that exhibit unusual metabolic versatility can produce hydrogen gas (H2) using the electrons derived from metabolism of organic compounds during photoheterotrophic growth. Here, based on 13C tracer experiments, we identified the network of glucose metabolism and quantified intracellular carbon fluxes in Rhodobacter sphaeroides KD131 grown under H2-producing conditions. Moreover, we investigated how the intracellular fluxes in R. sphaeroides responded to knockout mutations in hydrogenase and poly-β-hydroxybutyrate synthase genes, which led to increased H2 yield. The relative contribution of the Entner-Doudoroff pathway and Calvin-Benson-Bassham cycle to glucose metabolism differed significantly in hydrogenase-deficient mutants, and this flux change contributed to the increased formation of the redox equivalent NADH. Disruption of hydrogenase and poly-β-hydroxybutyrate synthase resulted in a significantly increased flux through the phosphoenolpyruvate carboxykinase and a reduced flux through the malic enzyme. A remarkable increase in the flux through the tricarboxylic acid cycle, a major NADH producer, was observed for the mutant strains. The in vivo regulation of the tricarboxylic acid cycle flux in photoheterotrophic R. sphaeroides was discussed based on the measurements of in vitro enzyme activities and intracellular concentrations of NADH and NAD+. Overall, our results provide quantitative insights into how photoheterotrophic cells manipulate the metabolic network and redistribute intracellular fluxes to generate more electrons for increased H2 production.
doi:10.1128/JB.05624-11
PMCID: PMC3256653  PMID: 22056932
16.  Impaired Thymic Export and Apoptosis Contribute to Regulatory T-Cell Defects in Patients with Chronic Heart Failure 
PLoS ONE  2011;6(9):e24272.
Objective
Animal studies suggest that regulatory T (Treg) cells play a beneficial role in ventricular remodeling and our previous data have demonstrated defects of Treg cells in patients with chronic heart failure (CHF). However, the mechanisms behind Treg-cell defects remained unknown. We here sought to elucidate the mechanism of Treg-cell defects in CHF patients.
Methods and Results
We performed flow cytometry analysis and demonstrated reduced numbers of peripheral blood CD4+CD25+FOXP3+CD45RO−CD45RA+ naïve Treg (nTreg) cells and CD4+CD25+FOXP3+CD45RO+CD45RA− memory Treg (mTreg) cells in CHF patients as compared with non-CHF controls. Moreover, the nTreg/mTreg ratio (p<0.01), CD4+CD25+FOXP3+CD45RO− CD45RA+CD31+ recent thymic emigrant Treg cell (RTE-Treg) frequency (p<0.01), and T-cell receptor excision circle levels in Treg cells (p<0.01) were lower in CHF patients than in non-CHF controls. Combined annexin-V and 7-AAD staining showed that peripheral Treg cells from CHF patients exhibited increased spontaneous apoptosis and were more prone to interleukin (IL)-2 deprivation- and CD95 ligand-mediated apoptosis than those from non-CHF individuals. Furthermore, analyses by both flow cytometry and real-time polymerase chain reaction showed that Treg-cell frequency in the mediastinal lymph nodes or Foxp3 expression in hearts of CHF patients was no higher than that of the non-CHF controls.
Conclusion
Our data suggested that the Treg-cell defects of CHF patients were likely caused by decreased thymic output of nascent Treg cells and increased susceptibility to apoptosis in the periphery.
doi:10.1371/journal.pone.0024272
PMCID: PMC3174174  PMID: 21935395
17.  Computational modelling elucidates the mechanism of ciliary regulation in health and disease 
BMC Systems Biology  2011;5:143.
Background
Ciliary dysfunction leads to a number of human pathologies, including primary ciliary dyskinesia, nephronophthisis, situs inversus pathology or infertility. The mechanism of cilia beating regulation is complex and despite extensive experimental characterization remains poorly understood. We develop a detailed systems model for calcium, membrane potential and cyclic nucleotide-dependent ciliary motility regulation.
Results
The model describes the intimate relationship between calcium and potassium ionic concentrations inside and outside of cilia with membrane voltage and, for the first time, describes a novel type of ciliary excitability which plays the major role in ciliary movement regulation. Our model describes a mechanism that allows ciliary excitation to be robust over a wide physiological range of extracellular ionic concentrations. The model predicts the existence of several dynamic modes of ciliary regulation, such as the generation of intraciliary Ca2+ spike with amplitude proportional to the degree of membrane depolarization, the ability to maintain stable oscillations, monostable multivibrator regimes, all of which are initiated by variability in ionic concentrations that translate into altered membrane voltage.
Conclusions
Computational investigation of the model offers several new insights into the underlying molecular mechanisms of ciliary pathologies. According to our analysis, the reported dynamic regulatory modes can be a physiological reaction to alterations in the extracellular environment. However, modification of the dynamic modes, as a result of genetic mutations or environmental conditions, can cause a life threatening pathology.
doi:10.1186/1752-0509-5-143
PMCID: PMC3224258  PMID: 21920041
18.  AAV2-mediated transfer of the human aquaporin-1 cDNA restores fluid secretion from irradiated miniature pig parotid glands 
Gene therapy  2010;18(1):38-42.
Previously (Shan et al, 2005), we reported that adenoviral vector-mediated transfer of the human aquaporin-1 (hAQP1) cDNA to minipig parotid glands following irradiation (IRti) transiently restored salivary flow to near normal levels. This study evaluated a serotype 2, adeno-associated viral (AAV2) vector for extended correction of IR (single dose; 20 Gy)-induced, parotid salivary hypofunction in minipigs. Sixteen weeks following IR, parotid salivary flow decreased by 85-90%. AAV2hAQP1 administration at week 17 transduced only duct cells and resulted in a dose-dependent increase in salivary flow to ∼35% of pre-IR levels (to ∼1ml/10min) after 8 weeks (peak response). Administration of a control AAV2 vector or saline, was without effect. Little change was observed in clinical chemistry and hematology values after AAV2hAQP1 delivery. Vector treated animals generated high anti-AAV2 neutralizing antibody titers by week 4 (∼1:1600) and significant elevations in salivary (∼15%), but not serum, GM-CSF levels. Following vector administration, salivary [Na+] was dramatically increased, from ∼10mM to ∼55 (at 4 weeks) and 39 mM (8 weeks). The findings demonstrate that localized delivery of AAV2hAQP1 to IR-damaged parotid glands leads to increased fluid secretion from surviving duct cells, and may be useful in providing extended relief of salivary hypofunction in previously irradiated patients.
doi:10.1038/gt.2010.128
PMCID: PMC3015016  PMID: 20882054
salivary gland; irradiation; aquaporin-1; AAV2; gene therapy
19.  Microbiome of Fungus-Growing Termites: a New Reservoir for Lignocellulase Genes▿ †  
Fungus-growing termites play an important role in lignocellulose degradation and carbon mineralization in tropical and subtropical regions, but the degradation potentiality of their gut microbiota has long been neglected. The high quality and quantity of intestinal microbial DNA are indispensable for exploring new cellulose genes from termites by function-based screening. Here, using a refined intestinal microbial DNA extraction method followed by multiple-displacement amplification (MDA), a fosmid library was constructed from the total microbial DNA isolated from the gut of a termite growing in fungi. Functional screening for endoglucanase, cellobiohydrolase, β-glucosidase, and xylanase resulted in 12 β-glucosidase-positive clones and one xylanase-positive clone. The sequencing result of the xylanase-positive clone revealed an 1,818-bp open reading frame (ORF) encoding a 64.5-kDa multidomain endo-1,4-β-xylanase, designated Xyl6E7, which consisted of an N-terminal GH11 family catalytic domain, a CBM_4_9 domain, and a Listeria-Bacteroides repeat domain. Xyl6E7 was a highly active, substrate-specific, and endo-acting alkaline xylanase with considerably wide pH tolerance and stability but extremely low thermostability.
doi:10.1128/AEM.01521-10
PMCID: PMC3019695  PMID: 21057022
20.  iPS Cells Reprogrammed From Human Mesenchymal-Like Stem/Progenitor Cells of Dental Tissue Origin 
Stem Cells and Development  2010;19(4):469-480.
Generation of induced pluripotent stem (iPS) cells holds a great promise for regenerative medicine and other aspects of clinical applications. Many types of cells have been successfully reprogrammed into iPS cells in the mouse system; however, reprogramming human cells have been more difficult. To date, human dermal fibroblasts are the most accessible and feasible cell source for iPS generation. Dental tissues derived from ectomesenchyme harbor mesenchymal-like stem/progenitor cells and some of the tissues have been treated as biomedical wastes, for example, exfoliated primary teeth and extracted third molars. We asked whether stem/progenitor cells from discarded dental tissues can be reprogrammed into iPS cells. The 4 factors Lin28/Nanog/Oct4/Sox2 or c-Myc/Klf4/Oct4/Sox2 carried by viral vectors were used to reprogram 3 different dental stem/progenitor cells: stem cells from exfoliated deciduous teeth (SHED), stem cells from apical papilla (SCAP), and dental pulp stem cells (DPSCs). We showed that all 3 can be reprogrammed into iPS cells and appeared to be at a higher rate than fibroblasts. They exhibited a morphology indistinguishable from human embryonic stem (hES) cells in cultures and expressed hES cell markers SSEA-4, TRA-1-60, TRA-1-80, TRA-2-49, Nanog, Oct4, and Sox2. They formed embryoid bodies in vitro and teratomas in vivo containing tissues of all 3 germ layers. We conclude that cells of ectomesenchymal origin serve as an excellent alternative source for generating iPS cells.
doi:10.1089/scd.2009.0314
PMCID: PMC2851830  PMID: 19795982
21.  iPS Cells Reprogrammed From Mesenchymal-Like Stem/Progenitor Cells of Dental Tissue Origin 
Stem cells and development  2010;19(4):469-480.
Generation of induced pluripotent stem (iPS) cells holds a great promise for regenerative medicine and other aspects of clinical applications. Many types of cells have been successfully reprogrammed into iPS cells in the mouse system; however, reprogramming human cells have been more difficult. To date, human dermal fibroblasts are the most accessible and feasible cell source for iPS generation. Dental tissues derived from ectomesenchyme harbor mesenchymal-like stem/progenitor cells and some of the tissues have been treated as biomedical wastes, for example, exfoliated primary teeth and extracted third molars. We asked whether stem/progenitor cells from discarded dental tissues can be reprogrammed into iPS cells. The 4 factors Lin28/Nanog/Oct4/Sox2 or c-Myc/Klf4/Oct4/Sox2 carried by viral vectors were used to reprogram 3 different dental stem/progenitor cells: stem cells from exfoliated deciduous teeth (SHED), stem cells from apical papilla (SCAP), and dental pulp stem cells (DPSCs). We showed that all 3 can be reprogrammed into iPS cells and appeared to be at a higher rate than fibroblasts. They exhibited a morphology indistinguishable from human embryonic stem (hES) cells in cultures and expressed hES cell markers SSEA-4, TRA-1-60, TRA-1-80, TRA-2-49, Nanog, Oct4, and Sox2. They formed embryoid bodies in vitro and teratomas in vivo containing tissues of all 3 germ layers. We conclude that cells of ectomesenchymal origin serve as an excellent alternative source for generating iPS cells.
doi:10.1089/scd.2009.0314
PMCID: PMC2851830  PMID: 19795982
22.  Succession of the Bacterial Community and Dynamics of Hydrogen Producers in a Hydrogen-Producing Bioreactor▿ †  
Applied and Environmental Microbiology  2010;76(10):3387-3390.
Variation in the hydrogen production rate was consistent with the succession of dominant bacteria during the batch fermentation process. Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes and quantitative analysis of the hydA genes at both the DNA and mRNA levels confirmed that Clostridium perfringens was the most dominant hydrogen producer in the bioreactor.
doi:10.1128/AEM.02444-09
PMCID: PMC2869144  PMID: 20305018
23.  Follow-Up of Patients with Multidrug Resistant Tuberculosis Four Years after Standardized First-Line Drug Treatment 
PLoS ONE  2010;5(5):e10799.
Background
In 2004, an anti-tuberculosis (TB) drug resistance survey in Heilongjiang province, China, enrolled 1574 (79%) new and 421 (21%) retreatment patients. Multi-drug resistant (MDR) TB was detected in 7.2% of new and 30.4% of retreatment patients. All received treatment with standardized first-line drug (FLD) regimens.
Methodology/Principal Findings
We report treatment outcomes of the 2004 cohort, and long-term outcomes as assessed in the second half of 2008. The reported cure rate for MDR-TB patients was 83% (94/113) among new and 66% (85/128) among retreatment patients (P<0.001). Ten of the 241 MDR-TB patients died during treatment. Of the remaining 231, 129 (56%) could be traced in 2008. The overall recurrence rates among new and retreatment cases were 46% and 66%, respectively (P = 0.03). The overall death rates among new and retreatment cases were 25% and 46%, respectively (P = 0.02). Forty percent of the traced new cases and 24% of the retreatment cases were alive and without recurrent TB (P = 0.01). Of the 16 patients who failed or defaulted from treatment in 2004, only two patients were not re-diagnosed with TB by 2008. Of the 111 (86%) patients with an initial successful treatment outcome 63 (57%) had developed recurrent TB, 40 (36%) had died, 27 (24%) of them died of TB. The follow-up period of four years precluded follow-up of all patients. In a highly conservative sensitivity analysis in which we assumed that all non-included patients were alive and did not have recurrent TB, the recurrence and death rate were 33% and 21%.
Conclusions/Significance
Documentation of cure based on conventional smear microscopy was a poor predictor of long term outcomes. MDR-TB patients in Heilongjiang province in China had high recurrence and death rates four years after treatment with standardized FLD regimens, reinforcing the need for early diagnosis and treatment of MDR-TB, including assessment of treatment outcomes with more sensitive laboratory methods.
doi:10.1371/journal.pone.0010799
PMCID: PMC2875387  PMID: 20520720
24.  Effect of Same-dose Single or Dual Field Irradiation on Damage to Miniature Pig Parotid Glands 
Aim
To evaluate the effect of single or dual field irradiation (IR) with the same dose on damage to miniature pig parotid glands.
Methodology
Sixteen miniature pigs were divided into two IR groups (n=6) and a control group (n=4). The irradiation groups were subjected to 20 Gy X-radiation to one parotid gland using single-field or dual-field modality by linear accelerator. The dose-volume distributions between two IR groups were compared. Saliva from parotid glands and blood were collected at 0, 4, 8 and 16 weeks after irradiation. Parotid glands were removed at 16 weeks to evaluate tissue morphology.
Results
The irradiation dose volume distributions were significantly different between single and dual field irradiation groups (t=4.177, P=0.002), although dose volume histogramin (DVH) indicated the equal maximal dose in parotid glands. Saliva flow rates from IR side decreased dramatically at all time points in IR groups, especially in dual field irradiation group. The radiation caused changes of white blood cell count in blood, lactate dehydrogenase and amylase in serum, calcium, potassium and amylase in saliva. Morphologically, more severe radiation damage was found in irradiated parotid glands from dual field irradiation group than that from single field irradiation group.
Conclusion
Data from this large animal model demonstrated that the radiation damage from the dual field irradiation was more severe than that of the single field irradiation at the same dose, suggesting that dose-volume distribution is an important factor in evaluation of the radiobiology of parotid glands.
doi:10.4248/ijos.08001
PMCID: PMC3735788  PMID: 20690500
irradiation damage; miniature pig; parotid gland
25.  catena-Poly[[(2,2′-bipyridine-κ2 N,N′)nickel(II)]-μ-oxalato-κ4 O 1,O 2:O 1′,O 2′] 
The title compound, [Ni(C2O4)(C10H8N2)]n, is isostructural with its MnII, FeII, CuII and ZnII analogues. Each NiII atom is chelated by two oxalate ligands and one 2,2′-bipyridine, forming a slightly distorted octa­hedral geometry. Oxlate acts as a bridge to link neighbouring pairs of NiII cations, forming a one-dimensional wave-like chain. The crystal showed partial inversion twinning.
doi:10.1107/S1600536808028389
PMCID: PMC2959270  PMID: 21201013

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