The transcription factor AP2 and downstream genes shared by asexual reproduction and zooidal regeneration in the tunicate, Polyandrocarpa misakiensis.

Epithelial outpocketing, tunic softening, mesenchymal cell death, dedifferentiation/transdifferentiation, and resistance to environmental stress are major events that occur during asexual reproduction by budding in the tunicate, Polyandrocarpa misakiensis. To identify the molecules underlying these events and compare them with those operating in regeneration, differential gene expression profiles were developed in buds and zooids. Among approximately 40,000 contigs, 21 genes were identified as potentially being involved in asexual reproduction. Genes related to tunic softening, phagocytosis-stimulating opsonin, and stress resistance were activated in the very early stage of budding. At the later stage of budding when buds separated from the parent and entered the developmental stage, genes for cell adhesion, cell death, and differentiation were activated. The transcription factor AP2 was spatio-temporally expressed in a similar pattern to the tunic-softening gene endoglucanase (EndoG). AP2 mRNA activated EndoG when introduced into zooids by electroporation. Eight out of 21 budding-related genes were significantly activated by AP2 mRNA. Polyandrocarpa zooids possess regenerative potential other than budding. Zooidal regeneration accompanied cell death/phagocytosis, cell-cell adhesion/communication, and dedifferentiation/redifferentiation. Consistent with morphological features, eight related genes including SP8 transcription factor were activated during zooidal regeneration. Most of these genes were identical to those induced by AP2 mRNA, indicating that asexual reproduction in P. misakiensis shares AP2-regulated downstream genes with zooidal regeneration. The present results suggest that SP8 may be indispensable for both budding and regeneration and that the potential dedifferentiation-related gene SOXB1 plays a minor role in zooidal regeneration.

In Vitro Phagocytosis of Different Dinoflagellate Species by Coral Cells.

Coral-dinoflagellate symbiosis is a unique biological phenomenon, in which animal cells engulf single-celled photosynthetic algae and maintain them in their cytoplasm mutualistically. Studies are needed to reveal the complex mechanisms involved in symbiotic processes, but it is difficult to answer these questions using intact corals. To tackle these issues, our previous studies established an in vitro system of symbiosis between cells of the scleractinian coral Acropora tenuis and the dinoflagellate Breviolum minutum, and showed that corals direct phagocytosis, while algae are likely engulfed by coral cells passively. Several genera of the family Symbiodiniaceae can establish symbioses with corals, but the symbiotic ratio differs depending on the dinoflagellate clades involved. To understand possible causes of these differences, this study examined whether cultured coral cells show phagocytotic activity with various dinoflagellate strains similar to those shown by intact A. tenuis. We found that (a) A. tenuis larvae incorporate Symbiodinium and Breviolum, but not Cladocopium, and very few Effrenium, (b) cultured coral cells engulfed all four species but the ratio of engulfment was significantly higher with Symbiodinium and Breviolum than Cladocopium and Effrenium, (c) cultured coral cells also phagocytosed inorganic latex beads differently than they do dinoflagellates . It is likely that cultured coral cells preferentially phagocytose Symbiodinium and Breviolum, suggesting that specific molecular mechanisms involved in initiation of symbiosis should be investigated in the future.

Pronounced QT Prolongation During General Anesthesia in a Child with Left Ventricular Noncompaction Cardiomyopathy: A Case Report.

We report the case of an 8-year-old boy with left ventricular noncompaction cardiomyopathy (LVNC) and QT prolongation who experienced further prolongation of the QTc during general anesthesia for extraction of a maxillary mesiodens. Pronounced prolongation of the QTc was observed after induction of general anesthesia with thiamylal and during emergence. No notable fluctuations in blood pressure, heart rate, and estimated continuous cardiac output were observed. We considered it likely that the QT prolongation was triggered by thiamylal and increased sympathetic nervous system activity. During general anesthesia for children with LVNC and QT prolongation, it is necessary to monitor intraoperative hemodynamic fluctuations and prepare for the possible occurrence of arrhythmias.

Regulators specifying cell fate activate cell cycle regulator genes to determine cell numbers in ascidian larval tissues.

In animal development, most cell types stop dividing before terminal differentiation; thus, cell cycle control is tightly linked to cell differentiation programmes. In ascidian embryos, cell lineages do not vary among individuals, and rounds of the cell cycle are determined according to cell lineages. Notochord and muscle cells stop dividing after eight or nine rounds of cell division depending on their lineages. In the present study, we showed that a Cdk inhibitor, Cdkn1.b, is responsible for stopping cell cycle progression in these lineages. Cdkn1.b is also necessary for epidermal cells to stop dividing. In contrast, mesenchymal and endodermal cells continue to divide even after hatching, and Myc is responsible for maintaining cell cycle progression in these tissues. Expression of Cdkn1.b in notochord and muscle is controlled by transcription factors that specify the developmental fate of notochord and muscle. Likewise, expression of Myc in mesenchyme and endoderm is under control of transcription factors that specify the developmental fate of mesenchyme and endoderm. Thus, cell fate specification and cell cycle control are linked by these transcription factors.

Total Intravenous Anesthesia Using Remimazolam and Continuous Cardiac Output Monitoring for Dental Anesthesia in a Patient With Takayasu's Arteritis: A Case Report.

Takayasu's arteritis is a persistent chronic progressive inflammation of the large- and medium-caliber arteries. Controlling cardiovascular variability during anesthesia and overcoming difficulties of cardiovascular monitoring due to the impaired vessels are important in patients with Takayasu's arteritis. Remimazolam is a novel short-acting benzodiazepine with mild effects on hemodynamics. We report the case of a patient with Takayasu's arteritis who underwent oral surgery under general anesthesia. This report highlights the use of remimazolam and remifentanil to reduce hemodynamic perturbations using estimated continuous cardiac output monitoring.

A Retrospective Case Series of Anesthetic Patients With Epiglottic Cysts.

Previously undiagnosed or asymptomatic epiglottic cysts may be coincidentally detected during intubation. This retrospective case series identified undiagnosed epiglottic cysts that were discovered during intubation in 4 patients who underwent oral surgery under general anesthesia at our hospital during a 6-year period. Including 2 additional cases, 1 previously diagnosed and 1 detected during preoperative imaging, epiglottic cysts were observed in 6 of 1112 cases (0.54%) total. Among the undiagnosed epiglottic cyst cases, mild dyspnea on effort or snoring was reported in 2 patients, but all others were asymptomatic. Upon discovering previously undiagnosed epiglottic cysts during intubation, it is essential to proceed cautiously, remain alert for potential airway management difficulties, and avoid injuring or rupturing the cysts. In addition, any available preoperative imaging should be reviewed as information pertinent to the airway and any abnormalities may be useful. This report discusses the anesthetic care of 6 patients with epiglottic cysts that were previously known or initially discovered during intubation.

Establishing Sustainable Cell Lines of a Coral, Acropora tenuis.

Planula larvae of the scleractinian coral, Acropora tenuis, consist of elongated ectodermal cells and developing inner endodermal cells. To establish in vitro cell lines for future studies of cellular and developmental potential of coral cells, larvae were successfully dissociated into single cells by treating them with a tissue dissociation solution consisting of trypsin, EDTA, and collagenase. Brown-colored cells, translucent cells, and pale blue cells were the major components of dissociated larvae. Brown-colored cells began to proliferate transiently in the culture medium that was devised for the coral, while translucent cells and pale blue cells decreased in number about 1 week after cell dissociation. In addition, when a modular protease, plasmin, was added to the cell culture medium, brown-colored cells extended pseudopodia and assumed amorphous shapes. They then continued to proliferate in clumps for more than 6 months with a doubling time of approximately 4-5 days. From 3 weeks of cell culture onward, brown-colored cells often aggregated and exhibited morphogenesis-like behavior to form flat sheets, and blastula-like clusters or gastrula-like spheres. Single cells or cell-clusters of the cell lines were analyzed by RNA-seq. This analysis showed that genes expressed in these cells in vitro were A. tenuis genes. Furthermore, each cell line expressed a specific set of genes, suggesting that their properties include gastroderm, secretory cells, undifferentiated cells, neuronal cells, and epidermis. All cell properties were maintained stably throughout successive cell cultures. These results confirm the successful establishment of a coral in vitro cell line.

YAF2-Mediated YY1-Sirtuin6 Interactions Responsible for Mitochondrial Downregulation in Aging Tunicates.

In budding tunicates, aging accompanies a decrease in the gene expression of mitochondrial transcription factor A (Tfam), and the in vivo transfection of Tfam mRNA stimulates the mitochondrial respiratory activity of aged animals. The gene expression of both the transcriptional repressor Yin-Yang-1 (YY1) and corepressor Sirtuin6 (Sirt6) increased during aging, and the cotransfection of synthetic mRNA of YY1 and Sirt6 synergistically downregulated Tfam gene expression. Pulldown assays of proteins indicated that YY1-associated factor 2 (YAF2) was associated with both YY1 and SIRT6. Protein cross-linking confirmed that YAF2 bound YY1 and SIRT6 with a molar ratio of 1:1. YY1 was bound to CCAT- or ACAT-containing oligonucleotides in the 5' flanking region of the Tfam gene. Chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) showed that SIRT6 specifically induced the histone H3 lysine 9 (H3K9) deacetylation of the Tfam upstream region. YY1 and YAF2 accelerated SIRT6-induced H3K9 deacetylation. YY1 and Sirt6 mRNA transfection attenuated mitochondrial respiratory gene expression and blocked MitoTracker fluorescence. In contrast, the SIRT6 inhibitor and Tfam mRNA antagonized the inhibitory effects of YY1 and Sirt6, indicating that Tfam acts on mitochondria downstream of YY1 and Sirt6. We concluded that in the budding tunicate Polyandrocarpa misakiensis, YY1 recruits SIRT6 via YAF2 to the TFAM gene, resulting in aging-related mitochondrial downregulation.

ANISEED 2019: 4D exploration of genetic data for an extended range of tunicates.

ANISEED (https://www.aniseed.cnrs.fr) is the main model organism database for the worldwide community of scientists working on tunicates, the vertebrate sister-group. Information provided for each species includes functionally-annotated gene and transcript models with orthology relationships within tunicates, and with echinoderms, cephalochordates and vertebrates. Beyond genes the system describes other genetic elements, including repeated elements and cis-regulatory modules. Gene expression profiles for several thousand genes are formalized in both wild-type and experimentally-manipulated conditions, using formal anatomical ontologies. These data can be explored through three complementary types of browsers, each offering a different view-point. A developmental browser summarizes the information in a gene- or territory-centric manner. Advanced genomic browsers integrate the genetic features surrounding genes or gene sets within a species. A Genomicus synteny browser explores the conservation of local gene order across deuterostome. This new release covers an extended taxonomic range of 14 species, including for the first time a non-ascidian species, the appendicularian Oikopleura dioica. Functional annotations, provided for each species, were enhanced through a combination of manual curation of gene models and the development of an improved orthology detection pipeline. Finally, gene expression profiles and anatomical territories can be explored in 4D online through the newly developed Morphonet morphogenetic browser.

Reporter Analyses Reveal Redundant Enhancers that Confer Robustness on Cis-Regulatory Mechanisms.

Reporter analyses of Hox1 and Brachyury (Bra) genes have revealed examples of redundant enhancers that provide regulatory robustness. Retinoic acid (RA) activates through an RA-response element the transcription of Hox1 in the nerve cord of the ascidian Ciona intestinalis. We also found a weak RA-independent neural enhancer within the second intron of Hox1. The Hox1 gene in the larvacean Oikopleura dioica is also expressed in the nerve cord. The O. dioica genome, however, does not contain the RA receptor-encoding gene, and the expression of Hox1 has become independent of RA. We have found that the upstream sequence of the O. dioica Hox1 was able to activate reporter gene expression in the nerve cord of the C. intestinalis embryo, suggesting that an RA-independent regulatory system in the nerve cord might be common in larvaceans and ascidians. This RA-independent redundant regulatory system may have facilitated the Oikopleura ancestor losing RA signaling without an apparent impact on Hox1 expression domains. On the other hand, vertebrate Bra is expressed in the ventral mesoderm and notochord, whereas its ascidian ortholog is exclusively expressed in the notochord. Fibroblast growth factor (FGF) induces Bra in the ventral mesoderm in vertebrates, whereas it induces Bra in the notochord in ascidians. Disruption of the FGF signal does not completely silence Bra expression in ascidians, suggesting that FGF-dependent and independent enhancers might comprise a redundant regulatory system in ascidians. The existence of redundant enhancers, therefore, provides regulatory robustness that may facilitate the acquisition of new expression domains.

Influence of the marvelous™ three-way stopcock on the natural frequency and damping coefficient in blood pressure transducer kits.

Two types of Planecta™ ports are commonly used as sampling ports in blood pressure transducer kits: a flat-type port (FTP) and a port with a three-way stopcock (PTS). Recently, a new type of three-way stopcock (Marvelous™) has been released as a Planecta™ counterpart, but its effects on the frequency characteristics and reliability of blood pressure monitoring have not been investigated. We assessed the influence of the Marvelous™ stopcock on the frequency characteristics of the pressure transducer kit. The basic pressure transducer kit, DT4812J, was modified by replacing one or two of the original three-way stopcocks with Marvelous™ stopcocks. The frequency characteristics (i.e., natural frequency and damping coefficient) of each kit were determined using wave parameter analysis software, and subsequently evaluated on a Gardner chart. Replacement of the original blood pressure transducer kit stopcocks with Marvelous™ stopcocks decreased the natural frequency (48.3 Hz) to 46.3 Hz or 44.8 Hz, respectively; the damping coefficient was not significantly changed. Plotting the data on a Gardner chart revealed that the changes fell within the adequate dynamic response region, indicating they were within the allowable range. Insertion of Marvelous™ stopcocks slightly affects the natural frequency of the pressure transducer kit, similar to inserting a PTS. The results indicate that the Marvelous™ stopcock is useful for accurate monitoring of arterial blood pressure, and may be recommended when insertion of two or more closed-loop blood sampling systems is necessary.

Frequency characteristics of pressure transducer kits with inserted pressure-resistant extension tubes.

The accurate monitoring of arterial blood pressure is important for cardiovascular management. However, the frequency characteristics of pressure transducer kits are influenced by the length of the pressure-resistant tube. To date, there have been few studies addressing the frequency characteristics of pressure transducer kits with inserted pressure-resistant extension tubes (pressure-resistant extension tube (ET) circuits). In this study, we examine ET circuits from the viewpoint of the frequency characteristics of pressure transducer kits. DT4812J transducer kits (length 150 cm; Argon Medical Devices, TX, USA) were used. Three original ET circuits were prepared, with the pressure-resistant tube of the DT4812J being extended with a 30-cm length of pressure-resistant tube (180ET circuit), a 60-cm length of pressure-resistant tube (210ET circuit), and a 90-cm length of pressure-resistant tube (240ET circuit). Each of these circuits was evaluated as part of this study. The natural frequency of the original DT4812J circuit was 45.90 Hz while the damping coefficient was 0.160. For the 180 ET circuit, the natural frequency and damping coefficient were 36.4 Hz and 0.162, respectively. For the ET210 circuit, the natural frequency and damping coefficient were 30.3 Hz and 0.175, respectively. For the ET210 circuit, the natural frequency and damping coefficient were 25.3 Hz and 0.180, respectively. As a result of extending the circuit, it was found that the natural frequency decreased drastically, while the damping coefficient increased slightly. When the extension of a pressure transducer kit is required, we should pay careful attention to the major decrease in the natural frequency, which may influence the pressure monitoring.

Acoustic method respiratory rate monitoring is useful in patients under intravenous anesthesia.

Respiratory depression can occur during intravenous general anesthesia without tracheal intubation. A new acoustic method for respiratory rate monitoring, RRa® (Masimo Corp., Tokyo, Japan), has been reported to show good reliability in post-anesthesia care and emergency units. The purpose of this study was to investigate the reliability of the acoustic method for measurement of respiratory rate during intravenous general anesthesia, as compared with capnography. Patients with dental anxiety undergoing dental treatment under intravenous anesthesia without tracheal intubation were enrolled in this study. Respiratory rate was recorded every 30 s using the acoustic method and capnography, and detectability of respiratory rate was investigated for both methods. This study used a cohort study design. In 1953 recorded respiratory rate data points, the number of detected points by the acoustic method (1884, 96.5 %) was significantly higher than that by capnography (1682, 86.1 %) (P < 0.0001). In the intraoperative period, there was a significant difference in the LOA (95 % limits of agreement of correlation between difference and average of the two methods)/ULLOA (under the lower limit of agreement) in terms of use or non-use of a dental air turbine (P < 0.0001). In comparison between capnography, the acoustic method is useful for continuous monitoring of respiratory rate in spontaneously breathing subjects undergoing dental procedures under intravenous general anesthesia. However, the acoustic method might not accurately detect in cases in with dental air turbine.

Transcriptional regulation of a horizontally transferred gene from bacterium to chordate.

The horizontal transfer of genes between distantly related organisms is undoubtedly a major factor in the evolution of novel traits. Because genes are functionless without expression, horizontally transferred genes must acquire appropriate transcriptional regulations in their recipient organisms, although the evolutionary mechanism is not known well. The defining characteristic of tunicates is the presence of a cellulose containing tunic covering the adult and larval body surface. Cellulose synthase was acquired by horizontal gene transfer from Actinobacteria. We found that acquisition of the binding site of AP-2 transcription factor was essential for tunicate cellulose synthase to gain epidermal-specific expression. Actinobacteria have very GC-rich genomes, regions of which are capable of inducing specific expression in the tunicate epidermis as the AP-2 binds to a GC-rich region. Therefore, the actinobacterial cellulose synthase could have been potentiated to evolve its new function in the ancestor of tunicates with a higher probability than the evolution depending solely on a spontaneous event.

[Perioperative Management of a Child with Vocal Adhesion Leading to Unexpected Difficult Airway].

We report a child with vocal cord adhesion encountered during induction of anesthesia. A 4-month-old girl was scheduled for bilateral lip plasty. She was intubated for one week due to pneumonia at the age of 3 days. Hoarseness and stridor appeared just after extubation. Although laryngo-fiberoptic examination had been tried several times, otorhinologists could not find any abnormality. We once decided to postpone the operation because of severe stridor. However, laryngofiberoptic examination could not reveal any abnormality, and we rescheduled the operation. Tracheal intubation using laryngoscope was not possible due to vocal cord adhesion. Finally, 2.5 mm ID tracheal tube was intubated by using a fiberscope, and lip plasty was performed. The patient stayed in the ICU for 7 days after surgery. Tracheotomy was performed 3 weeks after the operation. We should pay attention to stridor in an infant before general anesthesia, since it suggests severe airway narrowing although laryngo-fiberoptic examination could not find any abnormalities.

[Anesthesia Management of a Patient with Pulmonary Atresia, Intact Ventricular Septum, Major Aortopulmonary Collateral Artery and Tetralogy of Fallot].

The patient was a 6-year-old girl with pulmonary atresia, intact ventricular septum and major aortopulmonary collateral artery with tetralogy of Fallot Her Sp(O2) was around 60% under room air, and she could not walk long. She underwent dental treatment under general anesthesia. Invasive monitoring using pulmonary artery catheter should have been avoided, since the risk of monitoring greatly exceeds that of the treatment. The patient entered the operating room with her mother, and anesthesia was induced with intravenous midazolam, propofol and vecuronium. She was intubated orally first and impedance cardiography monitoring was started. FI(O2) was maintained at 0.5-1.0. Increases in airway pressure and Pa(CO2) were appropriately avoided. Dental treatment is important for infants with cardiac disease not only to reduce their pain, but also to reduce the risk of infection. It often requires general anesthesia. We have to conduct it with less invasiveness and less stress.

[Intravenous Sedation and Repeated "the Same Day General Anesthesia" for a School-age Boy with Dandy-Walker Syndrome and Dentinogenesis].

Dandy-Walker syndrome (DWS) is characterized by perfect or partial defect of the cerebellum vermis and cystic dilatation of the posterior fossa communicating with the fourth ventricle. Common clinical signs are mental retardation, cerebellar ataxia, and those of increased intracranial pressure (ICP). Associated congenital anomalies are craniofacial, cardiac, renal, and skeletal abnormalities. We experienced a case of intravenous sedation and six times of "the same day" general anesthesia for a school-aged boy (10-13 years old) with DWS and hypodentinogenesis. The patient underwent an examination and dental treatments. We had to pay attention to airway management tracheal tube selection and control of ICP. In addition, we should prevent tooth injuries through mishaps during tracheal intubations, since all-tooth-hypoplasia with fragile dental crowns was strongly suggested in this case. Detailed postoperative care is also required for general anesthesia afflicted with DWS.

[Repeated Anesthesia Management in a Patient with Aicardi Syndrome].

Aicardi syndrome is a rare hereditary disorder that develops in only girls with the trilogy of nutatory epilepsy, callosal agenesis and chorioretinopathy. We experienced general anesthesia twice for a patient with Aicardi syndrome in addition to heavy mental retardation. She underwent surgical correction for cleft lip and palate at 6 months of age and at 2 years of age, respectively. Anesthesia was induced slowly with inhalation of nitrous oxide, oxygen and sevoflurare. After securing an intravenous route, midazolam, thiopental and vecuronium were administered and intubated orally. Anesthesia was maintained with isoflurane safely. Patients with Aicardi syndrome have a high risk of aspiration pneumonia caused by underdeveloped swallowing ability due to callosal agenesis. We should, therefore, pay attention to prevention of seizure and aspiration pneumonia during the perioperative period.

Effect of using a Planecta™ port with a three-way stopcock on the natural frequency of blood pressure transducer kits.

Blood pressure transducer kits are equipped with two types of Planecta™ ports-the flat-type Planecta™ port (FTP) and the Planecta™ port with a three-way stopcock (PTS). We reported that FTP application decreased the natural frequency of the kits. However, Planecta™ is an invaluable tool as it prevents infection, ensures technical simplicity, and excludes air. Hence, an ideal Planecta™ port that does not decrease the frequency characteristics is required. As a first step in this direction, we aimed to assess the influence of PTSs on the natural frequency of blood transducer kits. A DTXplus transducer kit (DT4812J; Argon Medical Devices, TX, USA) was used along with ≥1 PTSs (JMS, Hiroshima, Japan), and the frequency characteristics were assessed. The natural frequency and damping coefficient of each kit were obtained by using frequency characteristics analysis software, and these parameters were evaluated by plotting them on Gardner's chart. Regardless of whether one or two PTSs were inserted, the natural frequency of the kits only slightly decreased (from 42.5 to 41.1 Hz, when 2 PTSs were used). Thus, the frequency characteristics of the kits with PTSs were adequate for pressure monitoring. The insertion of ≥2 FTPs in pressure transducer kits should be avoided, as they markedly decrease the natural frequency and lead to underdamping. However, the effect of PTS insertion in pressure transducer kits on the frequency characteristics is minimal. Thus, we found that the use of PTS markedly improved the frequency characteristics as compared to the use of FTP.