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2.  Cryo-electron tomography of motile cilia and flagella 
Cilia  2015;4:3.
Cryo-electron tomography has been a valuable tool in the analysis of 3D structures of cilia at molecular and cellular levels. It opened a way to reconstruct 3D conformations of proteins in cilia at 3-nm resolution, revealed networks of a number of component proteins in cilia, and has even allowed the study of component dynamics. In particular, we have identified the locations and conformations of all the regular inner and outer dyneins, as well as various regulators such as radial spokes. Since the mid 2000s, cryo-electron tomography has provided us with new knowledge, concepts, and questions in the area of cilia research. Now, after nearly 10 years of application of this technique, we are turning a corner and are at the stage to discuss the next steps. We expect further development of this technique for specimen preparation, data acquisition, and analysis. While combining this tool with other methodologies has already made cryo-electron tomography more biologically significant, we need to continue this cooperation using recently developed biotechnology and cell biology approaches.
In this review, we will provide an up-to-date overview of the biological insights obtained by cryo-electron tomography and will discuss future possibilities of this technique in the context of cilia research.
Electronic supplementary material
The online version of this article (doi:10.1186/s13630-014-0012-7) contains supplementary material, which is available to authorized users.
PMCID: PMC4313461  PMID: 25646146
3.  The Pathological Response to Anthracycline is Associated with Topoisomerase IIα Gene Amplification in the HER2 Breast Cancer Subset 
HER2-positive breast cancer sensitivity to anthracyclines is enhanced when topoisomerase IIα (TOP2A) is co-amplified under both adjuvant and metastatic settings. However, the relationship between anthracycline sensitivity and TOP2A amplification in HER2-positive breast cancers in neoadjuvant settings is not known.
The TOP2A gene status was examined by FISH in biopsies from 18 patients who received anthracycline and cyclophosphamide before surgery.
The TOP2A gene was amplified in 6/17 patients and was significantly associated with pathological response to the chemotherapy regimen.
TOP2A amplification could predict anthracycline-sensitivity. Thus, the HER2/TOP2A co-amplified subtype may be effectively treated by anthracycline-containing regimens alone.
PMCID: PMC4309011  PMID: 25642443
Breast Cancer; Neoadjuvant; Predictive factor; HER2; TOP2A; FISH
4.  Intrathoracic esophagojejunostomy using OrVil™ for gastric adenocarcinoma involving the esophagus 
AIM: To demonstrate a new surgical technique of lower mediastinal lymphadenectomy and intrathoracic anastomosis of esophagojejunostomy using OrVil™.
METHODS: After a total median phrenotomy, the supradiaphragmatic and lower thoracic paraesophageal lymph nodes were transhiatally dissected. The esophagus was cut off using a liner stapler and OrVil™was inserted. Finally, end-to-side esophagojejunostomy was created by using a circular stapler. From July 2009, we adopted this surgical technique for five patients with gastric cancer involving the lower esophagus.
RESULTS: The median operation time was 314 min (range; 210-367 min), and median blood loss was 210 mL (range; 100-838 mL). The median numbers of dissected lower mediastinal nodes were 3 (range; 1-10). None of the patients had postoperative complications including anastomotic leakage and stenosis. The median hospital stay was 16 d (range: 15-20 d). The median length of esophageal involvement was 14 mm (range: 6-48 mm) and that of the resected esophagus was 40 mm (range: 35-55 mm); all resected specimens had tumor-free margins.
CONCLUSION: This surgical technique is easy and safe intrathoracic anastomosis for the patients with gastric adenocarcinoma involving the lower esophagus.
PMCID: PMC4278145  PMID: 25548608
Gastric cancer; Esophageal invasion; Lower mediastinal lymphadenectomy; OrVil™; Intrathoracic anastomosis
5.  Gastrointestinal stromal tumors with exon 8 c-kit gene mutation might occur at extragastric sites and have metastasis-prone nature 
Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the human gut. Most sporadic GISTs have somatic gain-of-function mutations of the c-kit gene. The mutations are frequently found at exon 11, sometimes at exon 9 and rarely at exon 13 or 17. Recently, exon 8 c-kit gene mutations were reported in very minor proportion of sporadic GISTs. We also found 3 GISTs with exon 8 c-kit gene mutations in approximately 1,000 sporadic GISTs examined. In the present report, we showed the clinicopathological data of those GISTs. One case had a deletion of codon 419 of aspartate, and 2 cases had a substitution of 3 amino acids of codon 417 to codon 419 to tyrosine. The former was the same mutation recently reported in 2 GIST cases, but the latter has not been reported in any GISTs. All three cases occurred at extragastric sites and two of three showed distant metastasis. Since the remaining case was regarded as high risk for recurrence, imatinib adjuvant treatment has been done without evidence of metastasis. Our results confirmed the idea that exon 8 mutations are minor but actually existing abnormalities in sporadic GISTs, and suggested that such GISTs have a feature of extragastric development and a metastasis-prone nature. Since the exon 8 mutations appeared to be really sensitive to imatinib as shown in the present case study, accurate genotyping including exon 8 of the c-kit gene is necessary in GISTs to predict response to imatinib in both the unresectable/metastatic and adjuvant settings.
PMCID: PMC4270532  PMID: 25550846
c-kit gene; exon 8; gain-of-function mutation; GIST; imatinib; sunitinib
6.  Protein tagging reveals new insights into signaling in flagella 
The Journal of Cell Biology  2014;204(5):631-633.
In this issue, Oda et al. (2014. J. Cell Biol. use mutant analysis, protein tagging, and cryoelectron tomography to determine the detailed location of components in flagellar radial spokes—a complex of proteins that connect the peripheral microtubule doublets to the central pair. Remarkably, this approach revealed an interaction between radial spokes and the central pair based on geometry rather than a specific signaling mechanism, highlighting the importance of studying a system in three dimensions.
PMCID: PMC3941048  PMID: 24590169
7.  Colorectal laterally spreading tumors show characteristic expression of cell polarity factors, including atypical protein kinase C λ/ι, E-cadherin, β-catenin and basement membrane component 
Oncology Letters  2014;8(3):977-984.
Colorectal flat-type tumors include laterally spreading tumors (LSTs) and flat depressed-type tumors. The former of which shows a predominant lateral spreading growth rather than an invasive growth. The present study examined the morphological characteristics of LSTs, in comparison with polypoid- or flat depressed-type tumors, along with the expression of atypical protein kinase C (aPKC) λ/ι, a pivotal cell polarity regulator, and the hallmarks of cell polarity, as well as with type IV collagen, β-catenin and E-cadherin. In total, 37 flat-type (24 LSTs and 13 flat depressed-type tumors) and 20 polypoid-type colorectal tumors were examined. The LSTs were classified as 15 LST adenoma (LST-A) and nine LST cancer in adenoma (LST-CA). An immunohistochemical examination was performed on aPKC λ/ι, type IV collagen, β-catenin and E-cadherin. The LST-A and -CA showed a superficial replacing growth pattern, with expression of β-catenin and E-cadherin in the basolateral membrane and type IV collagen along the basement membrane. In addition, 86.6% of LST-A and 55.6% of LST-CA showed aPKC λ/ι expression of 1+ (weak to normal intensity staining in the cytoplasm compared with the normal epithelium). Furthermore, ~45% of the polypoid-type adenomas showed 2+ (moderate intensity staining in the cytoplasm and/or nucleus) and 66.7% of the polypoid-type cancer in adenoma were 3+ (strong intensity staining in the cytoplasm and nucleus). A statistically significant positive correlation was observed between the expression of aPKC λ/ι and β-catenin (r=0.842; P<0.001), or type IV collagen (r=0.823; P<0.001). The LSTs showed a unique growth pattern, different from the expanding growth pattern presented by a polypoid tumor and invasive cancer. The growth characteristics of LST appear to be caused by adequate coexpression of β-catenin, type IV collagen and aPKC λ/ι.
PMCID: PMC4114626  PMID: 25120645
laterally spreading tumor; atypical protein kinase C λ/ι
8.  High expression of ATP-binding cassette transporter ABCC11 in breast tumors is associated with aggressive subtypes and low disease-free survival 
ATP-binding cassette (ABC) transporters are membrane proteins that efflux various compounds from cells, including chemotherapeutic agents, and are known to affect multidrug resistance. Recent reports disagree on whether ABCC11 is a risk factor for breast tumorigenesis, but its expression in breast cancer is poorly investigated. We hypothesized that both frequency and expression levels of ABC transporters in breast tumors would vary by cancer subtype, and be associated with prognosis. Here, we constructed a tissue microarray breast tumor samples from 281 patients, and analyzed expressions of ABCB1, ABCC1, ABCC11, and ABCG2 immunohistochemically. Breast cancer subtypes were determined by immunohistochemistry of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2). Protein expression was correlated to clinicopathological characteristics, clinical follow-up, and pathological complete response to neoadjuvant chemotherapy. The tissue microarray comprised 191 luminal A (68.0%), 17 luminal B (6.0%), 27 HER2 (9.0%), and 46 triple-negative (16.4%) samples. ABCC1 and ABCC11 expressions were associated with significantly shorter disease-free survival (P = 0.027 and P = 0.003, respectively). ABCC1, ABCC11, and ABCG2, but not ABCB1, were expressed significantly more, and more frequently, in aggressive subtypes. Patients with HER2+ and triple-negative tumor subtypes that expressed high levels of ABCC11 had significantly worse disease-free survival (P = 0.017 and P < 0.001, respectively). We have shown, for the first time, that ABCC1, ABCC11 and ABCG2 are highly expressed in aggressive breast cancer subtypes, and that tumor ABCC11 expression is associated with poor prognosis.
PMCID: PMC3560367  PMID: 23288347
Breast cancer; ATP binding cassette transporters; ABCC11; tissue microarray; subtype
9.  Is Early Enteral Nutrition Initiated Within 24 Hours Better for the Postoperative Course in Esophageal Cancer Surgery? 
Early enteral nutrition within 24 h after surgery has become a recommended procedure. In the present study, we retrospectively examined whether initiating EN within 24 h after esophagectomy improves the postoperative course.
Among 103 patients who underwent thoracic esophagectomy for esophageal cancer, we enrolled the cases in which EN was initiated within 72 h after surgery. The patients were divided into two groups: EN started within 24 h (Group D1) and EN started at 24 - 72 h (Group D2-3). Clinical factors including days for first fecal passage, dose of postoperative albumin infusion, difference in serum albumin between pre- and postoperation, incidence of postoperative infection, and use of total parenteral nutrition were compared. Statistical analyses were performed by the Mann-Whitney U test and Chi square test, with significance defined as P < 0.05.
There was no significant difference between the groups in clinical factors. While pneumonia was significantly more frequent in Group D1 than in Group D2-3 (P = 0.0308), the frequency of infectious complications was comparable between the groups.
Initiating EN within 24 h showed no advantage for the postoperative course in esophageal cancer, and thus EN should be scheduled within 24 - 72 h, based on the patient condition.
PMCID: PMC3881990  PMID: 24400032
Early enteral nutrition; Esophageal cancer; Infectious complication; Pneumonia
10.  Two-Step Laparoscopic Surgery for a Patient with Synchronous Double Cancer of the Colon and Stomach Accompanied by Severe Chronic Obstructive Pulmonary Disease 
Case Reports in Surgery  2013;2013:246515.
Laparoscopic treatment strategies for synchronous intra-abdominal malignancies have not yet been standardized. We report a successful case of two-step laparoscopic surgery for synchronous double cancer of the colon and stomach accompanied by severe chronic obstructive pulmonary disease (COPD). A 66-year-old man with COPD was diagnosed as having advanced colon cancer and early gastric cancer. On admission, he could not go upstairs (Grade III according to the Hugh-Jones classification) and his forced expiratory volume in 1 second was 600 mL (35.9%). The patient initially underwent laparoscopy-assisted sigmoidectomy with D3 lymphadenectomy, followed by laparoscopy-assisted distal gastrectomy with D1 lymphadenectomy 68 days later. The patient's each postoperative course was uneventful with no pulmonary complications, and the patient was discharged 9 and 11 days after the first and second operations, respectively. The present case demonstrates that two-step laparoscopic surgery may be a safe and feasible surgical procedure for high-risk patients with synchronous intra-abdominal malignancies.
PMCID: PMC3777207  PMID: 24093070
11.  Is Early Enteral Nutrition Better for Postoperative Course in Esophageal Cancer Patients? 
Nutrients  2013;5(9):3461-3469.
We retrospectively examined esophageal cancer patients who received enteral nutrition (EN) to clarify the validity of early EN compared with delayed EN. A total of 103 patients who underwent transthoracic esophagectomy with three-field lymphadenectomy for esophageal cancer were entered. Patients were divided into two groups; Group E received EN within postoperative day 3, and Group L received EN after postoperative day 3. The clinical factors such as days for first fecal passage, the dose of postoperative albumin infusion, differences of serum albumin value between pre- and postoperation, duration of systematic inflammatory response syndrome (SIRS), incidence of postoperative infectious complication, and use of total parenteral nutrition (TPN) were compared between the groups. The statistical analyses were performed using Mann-Whitney U test and Chi square test. The statistical significance was defined as p < 0.05. Group E showed fewer days for the first fecal passage (p < 0.01), lesser dose of postoperative albumin infusion (p < 0.01), less use of TPN (p < 0.01), and shorter duration of SIRS (p < 0.01). However, there was no significant difference in postoperative complications between the two groups. Early EN started within 3 days after esophagectomy. It is safe and valid for reduction of albumin infusion and TPN, for promoting early recovery of intestinal movement, and for early recovery from systemic inflammation.
PMCID: PMC3798914  PMID: 24067386
early enteral nutrition; esophageal cancer; TPN; systematic inflammatory response syndrome
13.  Randomized controlled trial of toremifene 120 mg compared with exemestane 25 mg after prior treatment with a non-steroidal aromatase inhibitor in postmenopausal women with hormone receptor-positive metastatic breast cancer 
BMC Cancer  2013;13:239.
After the failure of a non-steroidal aromatase inhibitor (nsAI) for postmenopausal patients with metastatic breast cancer (mBC), it is unclear which of various kinds of endocrine therapy is the most appropriate. A randomized controlled trial was performed to compare the efficacy and safety of daily toremifene 120 mg (TOR120), a selective estrogen receptor modulator, and exemestane 25 mg (EXE), a steroidal aromatase inhibitor. The primary end point was the clinical benefit rate (CBR). The secondary end points were objective response rate (ORR), progression-free survival (PFS), overall survival (OS) and toxicity.
Initially, a total of 91 women was registered in the study and randomly assigned to either TOR120 (n = 46) or EXE (n = 45) from October 2008 to November 2011. Three of the 46 patients in the TOR120 arm were not received treatment, 2 patients having withdrawn from the trial by their preference and one having been dropped due to administration of another SERM.
When analyzed after a median observation period of 16.9 months, the intention-to-treat analysis showed that there were no statistical difference between TOR120 (N = 46) and EXE (n = 45) in terms of CBR (41.3% vs. 26.7%; P = 0.14), ORR (10.8% vs. 2.2%; P = 0.083), and OS (Hazard ratio, 0.60; P = 0.22). The PFS of TOR120 was longer than that of EXE, the difference being statistically significant (Hazard ratio, 0.61, P = 0.045). The results in treatment-received cohort (N = 88) were similar to those in ITT cohort. Both treatments were well-tolerated with no severe adverse events, although the treatment of 3 of 43 women administered TOR120 was stopped after a few days because of nausea, general fatigue, hot flush and night sweating.
TOR120, as a subsequent endocrine therapy for mBC patients who failed non-steroidal AI treatment, could potentially be more beneficial than EXE.
Trial registration number
PMCID: PMC3661372  PMID: 23679192
Refractory to aromatase inhibitor; Toremifene; Exemestane; Breast cancer
14.  Polarity and asymmetry in the arrangement of dynein and related structures in the Chlamydomonas axoneme 
The Journal of Cell Biology  2012;198(5):913-925.
Cryoelectron tomography and subtomogram averaging reveal a high degree of structural asymmetry and polarization in dynein localization in the Chlamydomonas flagella.
Understanding the molecular architecture of the flagellum is crucial to elucidate the bending mechanism produced by this complex organelle. The current known structure of the flagellum has not yet been fully correlated with the complex composition and localization of flagellar components. Using cryoelectron tomography and subtomogram averaging while distinguishing each one of the nine outer doublet microtubules, we systematically collected and reconstructed the three-dimensional structures in different regions of the Chlamydomonas flagellum. We visualized the radial and longitudinal differences in the flagellum. One doublet showed a distinct structure, whereas the other eight were similar but not identical to each other. In the proximal region, some dyneins were missing or replaced by minor dyneins, and outer–inner arm dynein links were variable among different microtubule doublets. These findings shed light on the intricate organization of Chlamydomonas flagella, provide clues to the mechanism that produces asymmetric flagellar beating, and pose a new challenge for the functional study of the flagella.
PMCID: PMC3432765  PMID: 22945936
15.  Pearl Microstructure and Expression of Shell Matrix Protein Genes MSI31 and MSI60 in the Pearl Sac Epithelium of Pinctada fucata by In Situ Hybridization 
PLoS ONE  2013;8(1):e52372.
Expression patterns of the shell matrix protein genes MSI31 and MSI60 in the pearl sac epithelium were examined by in situ hybridization 38 days after implantation, and related to pearl quality. A pearl sac that produced a nacreous pearl showed very weak expression of MSI31 and strong expression of MSI60. A pearl sac, which yielded a prismatic pearl, strongly expressed MSI31 and very weakly expressed MSI60. In a complex pearl, whose surface consisted of a mosaic of both nacreous and prismatic layers, the expression pattern of MSI31 and MSI60 similarly corresponded to the underlying surface structures of the pearl. A nacreous pearl whose pearl sac showed strong MSI31 expression had an entirely nacreous surface composed of a laminar structure with unusual tablet growth at the corresponding site. MSI31 and MSI60 are the major components of the shell matrix proteins of the nacreous and prismatic layers. Clearly, high expression of MSI31 does not always result in prismatic secretion. These observations cannot be explained solely on the basis of the expression patterns of MSI31 and MSI60. We propose that, in addition to the MSI genes that form the prismatic and nacreous layers, upstream from these genes there are regulatory master genes that determine whether a nacreous layer (aragonite) or a prismatic layer (calcite) is formed.
PMCID: PMC3544836  PMID: 23341897
16.  Breast cancer manifested by hematologic disorders 
Journal of Thoracic Disease  2012;4(6):650-654.
Breast cancer is the most common type of cancer in women. However, it is very rarely manifested as hematologic disorders. A 35-year-old woman was admitted because of disseminated intravascular coagulation. Examinations revealed the presence of breast cancer in her left breast; therefore, paclitaxel was administered weekly. Although disseminated intravascular coagulation was controlled, pulmonary dysfunction due to lymphangitis carcinomatosa suddenly occurred 10 weeks after treatment. Pulmonary dysfunction was effectively treated with epirubicin and cyclophosphamide. Twenty-three weeks after treatment, the patient developed liver dysfunction accompanied with jaundice due to progressive metastatic lesions in the liver; liver dysfunction improved after the administration of vinorelbine. Subsequently, because of the recurrence of pulmonary dysfunction, rechallenge with epirubicin and cyclophosphamide was performed and was effective; however, this therapy was discontinued because of its adverse effects. She expired of liver failure 33 weeks after the occurrence of disseminated intravascular coagulation. Metastatic tumors in the bone marrow, lung, and liver showed different sensitivities to different anti-cancer agents. We report a case of breast cancer manifested by hematologic disorders which was treated by a sequential chemotherapy.
PMCID: PMC3506789  PMID: 23205295
Breast cancer; disseminated intravascular coagulation; multiple organ metastases
17.  Reexpansion Pulmonary Edema following Laparoscopy-Assisted Distal Gastrectomy for a Patient with Early Gastric Cancer: A Case Report 
Case Reports in Surgery  2012;2012:863163.
We report here a case of reexpansion pulmonary edema following laparoscopy-assisted distal gastrectomy (LADG) for early gastric cancer. A 57-year-old Japanese woman with no preoperative comorbidity was diagnosed with early gastric cancer. The patient underwent LADG using the pneumoperitoneum method. During surgery, the patient was unintentionally subjected to single-lung ventilation for approximately 247 minutes due to intratracheal tube dislocation. One hour after surgery, she developed severe dyspnea and produced a large amount of pink frothy sputum. Chest radiography results showed diffuse ground-glass attenuation and alveolar consolidation in both lungs without cardiomegaly. A diagnosis of pulmonary edema was made, and the patient was immediately intubated and received ventilatory support with high positive end-expiratory pressure. The patient gradually recovered and was weaned from the ventilatory support on the third postoperative day. This case shows that single-lung ventilation may be a risk factor for reexpansion pulmonary edema during laparoscopic surgery with pneumoperitoneum.
PMCID: PMC3514816  PMID: 23227412
18.  WallFlex™ Duodenal Stent Placement in a Gastric Cancer Patient with Malignant Stenosis of a Roux-en-Y Gastrojejunostomy following Distal Gastrectomy 
Case Reports in Oncology  2012;5(3):554-560.
A 69-year-old Japanese woman with a history of distal gastrectomy with a Roux-en-Y reconstruction for advanced gastric cancer was admitted to our hospital complaining of severe dysphagia. On admission, the patient was only able to take liquids, and a firm, fist-sized tumor was palpable in her left upper abdomen. An endoscopic examination disclosed stenosis of the jejunal limb of the gastrojejunostomy. Abdominal computed tomography revealed that a recurrent tumor, 5.0 cm in diameter, was compressing the jejunal limb of the gastrojejunostomy. A knitted nitinol self-expandable metallic stent (WallFlex™ duodenal stent) was placed endoscopically at the stenotic jejunum from the gastrojejunostomy. The time required for stenting and total endoscopic manipulation was 12 and 35 minutes, respectively. No stent-related complications were observed. The patient could resume oral ingestion 1 day after endoscopic stenting and was discharged on the fifth day after treatment. She survived for 201 days after stenting. She continued oral ingestion for 194 days and stayed at home for 165 days. The WallFlex duodenal stent allows safe endoscopic stenting, even in cases of malignant stenosis of a gastrojejunostomy following distal gastrectomy. This stenting device will extend the indications for endoscopic palliation of gastric cancer patients with gastric outlet stenosis.
PMCID: PMC3492972  PMID: 23139671
Gastric cancer; Roux-en-Y gastrojejunostomy; Recurrence; WallFlex™ duodenal stent
19.  Cryoelectron tomography of radial spokes in cilia and flagella 
The Journal of Cell Biology  2011;195(4):673-687.
Cryo-EM tomography of wild-type and mutant cilia and flagella from Tetrahymena and Chlamydomonas reveals new information on the substructure of radial spokes.
Radial spokes (RSs) are ubiquitous components in the 9 + 2 axoneme thought to be mechanochemical transducers involved in local control of dynein-driven microtubule sliding. They are composed of >23 polypeptides, whose interactions and placement must be deciphered to understand RS function. In this paper, we show the detailed three-dimensional (3D) structure of RS in situ in Chlamydomonas reinhardtii flagella and Tetrahymena thermophila cilia that we obtained using cryoelectron tomography (cryo-ET). We clarify similarities and differences between the three spoke species, RS1, RS2, and RS3, in T. thermophila and in C. reinhardtii and show that part of RS3 is conserved in C. reinhardtii, which only has two species of complete RSs. By analyzing C. reinhardtii mutants, we identified the specific location of subsets of RS proteins (RSPs). Our 3D reconstructions show a twofold symmetry, suggesting that fully assembled RSs are produced by dimerization. Based on our cryo-ET data, we propose models of subdomain organization within the RS as well as interactions between RSPs and with other axonemal components.
PMCID: PMC3257535  PMID: 22065640
20.  Draft Genome of the Pearl Oyster Pinctada fucata: A Platform for Understanding Bivalve Biology 
The study of the pearl oyster Pinctada fucata is key to increasing our understanding of the molecular mechanisms involved in pearl biosynthesis and biology of bivalve molluscs. We sequenced ∼1150-Mb genome at ∼40-fold coverage using the Roche 454 GS-FLX and Illumina GAIIx sequencers. The sequences were assembled into contigs with N50 = 1.6 kb (total contig assembly reached to 1024 Mb) and scaffolds with N50 = 14.5 kb. The pearl oyster genome is AT-rich, with a GC content of 34%. DNA transposons, retrotransposons, and tandem repeat elements occupied 0.4, 1.5, and 7.9% of the genome, respectively (a total of 9.8%). Version 1.0 of the P. fucata draft genome contains 23 257 complete gene models, 70% of which are supported by the corresponding expressed sequence tags. The genes include those reported to have an association with bio-mineralization. Genes encoding transcription factors and signal transduction molecules are present in numbers comparable with genomes of other metazoans. Genome-wide molecular phylogeny suggests that the lophotrochozoan represents a distinct clade from ecdysozoans. Our draft genome of the pearl oyster thus provides a platform for the identification of selection markers and genes for calcification, knowledge of which will be important in the pearl industry.
PMCID: PMC3325083  PMID: 22315334
pearl oyster; Pinctada fucata; draft genome
21.  Axonemal radial spokes 
Bioarchitecture  2012;2(2):50-58.
The radial spoke (RS) is a complex of at least 23 proteins that works as a mechanochemical transducer between the central‐pair apparatus and the peripheral microtubule doublets in eukaryotic flagella and motile cilia. The RS contributes to the regulation of the activity of dynein motors, and thus to flagellar motility. Despite numerous biochemical, physiological and structural studies, the mechanism of the function of the radial spoke remains unclear. Detailed knowledge of the 3D structure of the RS protein complex is needed in order to understand how RS regulates dynein activity. Here we review the most important findings on the structure of the RS, including results of our recent cryo‐electron tomographic analysis of the RS protein complex.
PMCID: PMC3383722  PMID: 22754630
axoneme; cilia; cryo‐electron tomography; dynein; flagella; motility; radial spokes
22.  Plasma Free Amino Acid Profiling of Five Types of Cancer Patients and Its Application for Early Detection 
PLoS ONE  2011;6(9):e24143.
Recently, rapid advances have been made in metabolomics-based, easy-to-use early cancer detection methods using blood samples. Among metabolites, profiling of plasma free amino acids (PFAAs) is a promising approach because PFAAs link all organ systems and have important roles in metabolism. Furthermore, PFAA profiles are known to be influenced by specific diseases, including cancers. Therefore, the purpose of the present study was to determine the characteristics of the PFAA profiles in cancer patients and the possibility of using this information for early detection.
Methods and Findings
Plasma samples were collected from approximately 200 patients from multiple institutes, each diagnosed with one of the following five types of cancer: lung, gastric, colorectal, breast, or prostate cancer. Patients were compared to gender- and age- matched controls also used in this study. The PFAA levels were measured using high-performance liquid chromatography (HPLC)–electrospray ionization (ESI)–mass spectrometry (MS). Univariate analysis revealed significant differences in the PFAA profiles between the controls and the patients with any of the five types of cancer listed above, even those with asymptomatic early-stage disease. Furthermore, multivariate analysis clearly discriminated the cancer patients from the controls in terms of the area under the receiver-operator characteristics curve (AUC of ROC >0.75 for each cancer), regardless of cancer stage. Because this study was designed as case-control study, further investigations, including model construction and validation using cohorts with larger sample sizes, are necessary to determine the usefulness of PFAA profiling.
These findings suggest that PFAA profiling has great potential for improving cancer screening and diagnosis and understanding disease pathogenesis. PFAA profiles can also be used to determine various disease diagnoses from a single blood sample, which involves a relatively simple plasma assay and imposes a lower physical burden on subjects when compared to existing screening methods.
PMCID: PMC3168486  PMID: 21915291
23.  Early Passenger Leukocyte Migration and Acute Immune Reactions in the Rat Recipient Spleen During Liver Engraftment: With Particular Emphasis on Donor Major Histocompatibility Complex Class II+ Cells1 
Transplantation  2002;74(1):103-111.
After a short course of tacrolimus, Lewis rat liver allografts induce donor-specific nonreactivity in Brown Norway recipients that is immunosuppression-independent after 28 days. To clarify the role of donor major histocompatibility complex (MHC) class II+ cells, we investigated the migration to the recipient splenic T- and B-cell compartments of different subsets of Lewis MHC class II+ passenger leukocytes. The rise and decline of immune activation were monitored in the hepatic allograft and in the host spleen by analyses of BrdU+ (proliferating) leukocytes, TUNEL+ (apoptotic) cells, apoptosis-associated molecules, TH1/ TH2 cytokine profiles, and histoimmunocytochemical examination of graft and splenic tissues. Serial flow cytometry studies during the 28-day period of drug-assisted “hepatic tolerogenesis” showed that migratory MHC class II+ cells accounted for less than half of the donor cells in the host spleen. The class II+ cells consisted mostly of B cells that homed to splenic B-cell follicles with only a sparse representation of dendritic cells that were exclusively found in the splenic periarteriolar lymphoid sheath. In parallel studies, transplantation of the less tolerogenic heart produced a diminutive version of the same events, but with far fewer donor cells in the host spleen, evidence of sustained immune activation, and the development of chronic rejection by 100 days. The data are consistent with the paradigm that migration of donor leukocytes is the prime determinant of variable tolerance induction induced by transplantation of the liver and other organs, but without regard for donor MHC class II+ expression.
PMCID: PMC3154775  PMID: 12134107
24.  Pedunculated gastric tube interposition in an esophageal cancer patient with prepyloric adenocarcinoma 
Gastric carcinoma is one of the malignancies that are most frequently associated with esophageal carcinoma. We describe herein our device for advanced esophageal cancer associated with early gastric cancer in the antrum. A 57-year-old man presenting with dysphagia and upper abdominal pain was admitted to our hospital. Preoperative examinations revealed locally advanced squamous cell carcinoma (SCC) of the middle thoracic esophagus (T3N0M0 Stage IIA) and mucosal signet-ring cell carcinoma of the gastric antrum (T1N0M0 Stage IA). Although the gastric tumor appeared to be an intramucosal carcinoma, its margin was obscure, so endoscopic en-bloc resection was considered inadequate. We chose surgical resection of the gastric tumor as well as the esophageal SCC after neoadjuvant chemotherapy with 5-fluorouracil and cisplatin for advanced esophageal cancer. Following transthoracic esophagectomy with three-field lymph node dissection, the gastric carcinoma was removed by gastric antrectomy, which preserved the right gastroepiploic vessels, and a pedunculated short gastric tube was used as the esophageal substitute. Twenty-eight months after the surgery, the patient is well with no evidence of cancer recurrence. Because it minimizes surgical stress and organ sacrifice, gastric tube interposition is a potentially useful technique for esophageal cancer associated with localized early gastric cancer.
PMCID: PMC3098435  PMID: 21603033
Antrectomy; Early gastric cancer; Esophageal cancer; Esophageal reconstruction; Gastric tube
25.  Local and Global Mobility in the ClpA AAA+ Chaperone Detected by Cryo-electron Microscopy: Functional Connotations 
The ClpA chaperone combines with the ClpP peptidase to perform targeted proteolysis in the bacterial cytoplasm. ClpA monomer has an N-terminal substrate-binding domain and two AAA+ ATPase domains (D1 and D2). ClpA hexamers stack axially on ClpP heptamers to form the symmetry-mismatched protease. We used cryo-electron microscopy to visualize the ClpA-ATPγS hexamer, in the context of ClpAP complexes. Two segments lining the axial channel show anomalously low density, indicating that these motifs, which have been implicated in substrate translocation, are mobile. We infer that ATP hydrolysis is accompanied by substantial structural changes in the D2 but not the D1 tier. The entire N-domain is rendered invisible by large-scale fluctuations. When deletions of 10 and 15-residues were introduced into the linker, N-domain mobility was reduced but not eliminated and changes were observed in enzymatic activities. Based on these observations, we present a pseudo-atomic model of ClpAP holoenzyme, a dynamic proteolytic nanomachine.
PMCID: PMC2871031  PMID: 20462489
chaperone; AAA+ ATPase; unfoldase; pseudo-atomic model; image reconstruction; conformational change

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