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Supplementary Figure 1, Figure 2, Figure 3. Representative photomicrographs detailing immunohistochemical analysis of ANGPTL2 expression, Figure S2: ANGPTL2 expression in 5 colorectal cancer cell lines, Figure S3: Receiver operating characteristics curve analysis using serum ANGPTL2 for distinguishing patients with CRC with normal CEA levels from normal controls (NC).

Purpose: Angiopoietin-like protein 2 (ANGPTL2) is a mediator of chronic inflammation and inflammatory carcinogenesis. The biologic and clinical significance of ANGPTL2 remains unknown in human cancer. Therefore, we investigated the function of ANGPTL2 and evaluated its clinical significance in both primary tumors and matched sera in patients with colorectal cancer.Experimental Design: A colorectal cancer cell line was transfected with siRNA against ANGPTL2 for the assessment of its function. We examined ANGPTL2 expression in colorectal cancer tissues (n = 195) by immunohistochemistry. Finally, we screened serum ANGPTL2 levels from 32 colorectal cancers and 23 normal controls (NC), and validated these results in serum samples obtained from 195 colorectal cancers and 45 NCs by ELISA.Results: Knockdown of ANGPTL2 in vitro significantly inhibited cell proliferation, migration, and invasion, whereas it enhanced anoikis. ANGPTL2 was overexpressed in colorectal cancer tissues, and was significantly associated with advanced T stage, lymph node, and liver metastasis. Likewise, serum ANGPTL2 levels in colorectal cancers were significantly higher than NCs (P < 0.01), and allowed distinguishing of colorectal cancers from NCs with high accuracy (AUC = 0.837). The subsequent validation step confirmed that serum ANGPTL2 levels in colorectal cancers were significantly higher than in NCs (P < 0.0001), and had a high AUC value (0.885) for distinguishing colorectal cancers from NCs. High serum ANGPTL2 was significantly associated with advanced T stage, lymph node and liver metastasis, early relapse, and poor prognosis in colorectal cancers.Conclusion: Serum ANGPTL2 is a novel diagnostic and recurrence-predictive biomarker in patients with colorectal cancer. Clin Cancer Res; 20(23); 6175–86. ©2014 AACR.

Supplementary Tables S1-3. Table S1:Patient characteristics in the validation set, Table S2:Primer Sequences for KRAS and BRAF mutation analysis, Table S3:Comparison between ROC from screening step and validation step.

Supplementary material and methods. Detailed patient characteristics in the validation step, Cell culture condition, 3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, yellow tetrazole (MTT) assay, Anoikis assay, Invasion assay, Wound healing assay,Immunohistochemistry, Enzyme-Linked Immunosorbent Assay (ELISA).

Clarifying the mechanisms of shape alteration by insect metamorphosis is important for comprehending exoskeletal morphogenesis. The large horn of the Japanese rhinoceros beetleTrypoxylus dichotomusis the result of drastic metamorphosis, wherein it appears as a rounded shape via pupation and then undergoes remodeling into an angular adult shape. However, the mechanical mechanisms of this remodeling process remain unknown. In this study, we investigated the remodeling mechanisms of the Japanese rhinoceros beetle horn by developing a physical simulation. We identified three factors contributing to remodeling by biological experiments—ventral adhesion, uneven shrinkage, and volume reduction—which were demonstrated to be crucial to the transformation by a physical simulation. We also corroborated our findings by applying the simulation to the stag beetle’s mandibular remodeling. These results indicate that the physical simulation is applicable to pupal remodeling in other beetles, and the morphogenic mechanisms could explain various exoskeletal shapes.Significance statementThe metamorphosis in insects is a mysterious process. By metamorphosis, insects sometimes change their shape dramatically. The head horn of the Japanese rhinoceros beetle is one of the most famous examples of metamorphosis. In larva-to-pupa molting, the horn appears suddenly, caused by the “furrow formation and unfolding” mechanism. The unfolding process makes the pupal horn rounded. However, pupa-to-adult molting transforms the rounded shape into an angular shape. In this paper, we investigated the mechanisms of the transformation. We extracted factors contributing to it through observations and experiments and developed a physical simulation. It could reproduce the adult shape from the pupal shape and could be a general model for the pupa-adult transformation of beetles.

There are no established indications for facial nerve schwannoma treatment, including surgery, radiation and follow-up observation, and it is difficult to determine treatment policy uniformly. The treatment policy was examined from each treatment course. Data of patients with facial nerve schwannomas at our hospital from 1987 to 2018 were retrospectively examined. Their age, sex, clinical symptoms, tumor localization, treatment policies and outcomes were reviewed. In total, 22 patients underwent surgery and 1 patient underwent radiotherapy; 22 patients were followed up without treatment. After total resection, there were no tumor recurrences, and most patients had grade 3 or 4 postoperative facial paralysis. After subtotal resection, tumor regrowth was observed in four patients and reoperation was required in two patients. Facial nerve function was maintained in four patients and was decreased in two patients. During follow-up, six patients showed tumor growth. Only one patient had worsening facial nerve paralysis; four patients underwent facial nerve decompression owing to facial nerve paralysis during follow-up. If the tumor compresses the brain or it is prone to growth, surgery may be indicated, and when the preoperative facial nerve function is grade ≤ 3, consideration should be given to preserving facial nerve function and subtotal resection should be indicated. If the preoperative facial nerve function is grade ≥ 3, total resection with nerve grafting is an option to prevent regrowth. If there is no brain compression or tumor growth, the follow-up is a good indication, and decompression should be considered in facial nerve paralysis cases.

Background Ceftriaxone, a third-generation cephalosporin antibiotic with a long plasma half-life, is widely used to treat various infections. The use of ceftriaxone can sometimes induce biliary sludge or stone formation. Although most cases of ceftriaxone-induced pseudolithiasis are asymptomatic or mild and resolve with discontinuation of the drug, we experienced an elderly case of severe acute necrotizing calculous cholecystitis after administration of ceftriaxone. Case presentation A 72-year-old male patient was admitted to our hospital because of acute diverticulitis in ascending colon. Ceftriaxone was administered at a dose of 2 g/day for 6 days. Although he recovered after therapy, he was readmitted about 2 weeks later because of severe pain with rebound tenderness in the right upper quadrant. An abdominal imaging study revealed stones and sludge in the gallbladder that were not observed before starting ceftriaxone therapy. Therefore, antibiotic treatment with flomoxef 2 g/day was indicated. However, on the fifth day of readmission, the peritoneal irritation symptoms in the right upper quadrant worsened, and elevated inflammatory response and liver dysfunction were observed. Cholecystectomy was performed based on these findings. The resected inflamed gallbladder showed acute necrotizing cholecystitis with sand granular gallstones. A comparative analysis of the infrared spectroscopic pattern of the composition of gallstones collected during surgery with that of the ceftriaxone powder revealed that both have very similar infrared spectroscopic patterns. Conclusions Ceftriaxone-related pseudolithiasis is generally reversible and mainly observed in children. Here, we report a rare case of ceftriaxone-related acute necrotizing cholecystitis in an elderly patient. We confirmed that the stones in the gallbladder are composed of ceftriaxone. The older age, dehydration, fasting, and long-time bed rest during the administration of high-dose ceftriaxone were the potential risk factors for gallstone formation.

Bacterial leaf spot and bacterial leaf blight are global threats to the cultivation of cruciferous vegetables, and it is necessary to develop methods to easily detect, identify, and distinguish the causative pathogens Pseudomonas syringae pv. maculicola (Psm) and P. cannabina pv. alisalensis (Pca). Here, we used the sequence specificity of the exchangeable effector loci flanking the hrp gene cluster to design primers that can help detect and discriminate between Psm and Pca. Primers common to both bacteria (hrpK_fw1 and hrpK_fw2) were designed within hrpK at the end of the hrp gene cluster. Psm-specific primers (MAC_rv1 and MAC_rv2) were designed in hopPtoB1 and Pca-specific primers (ALS_rv1 and ALS_rv2) were designed in hopX1 adjacent to hrpK. PCR using hrpK_fw1 and MAC_rv1 or hrpK_fw2 and MAC_rv2 amplified DNA fragments of only Psm, P. syringae pv. tomato (causal agent of tomato bacterial speck), and P. syringae pv. spinaciae (causal agent of spinach bacterial leaf spot), among 76 strains of phytopathogenic bacteria. PCR using hrpK_fw1 and ALS_rv1 or hrpK_2 and ALS_rv2 amplified DNA fragments of only Pca. Multiplex PCR with these primers could easily distinguish Psm and Pca from bacterial colonies isolated on growth media and detect the pathogen in symptomatic leaves. Multiplex nested PCR with the primers detected contamination in one Psm- and/or one Pca-infected seeds in 1000 seeds. These results suggest that these PCR primers could help detect and discriminate Psm and Pca. KEY POINTS: • We investigated Pseudomonas syringae pv. maculicola and P. cannabina pv. alisalensis. • Novel primers common to both bacteria were designed following genome comparison. • Multiplex PCR with new primers could discriminate Psm and Pca.

Three semi-selective media, DTarTA, SPbc, and SPamt, were developed and tested to isolate Pseudomonas syringae pv. maculicola (Psm) and P. cannabina pv. alisalensis (Pca) from Raphanus sativus seeds. DTarTA contained D-tartaric acid as a carbon source and potassium tellurite, ampicillin sodium, and methyl violet as antibiotics. DTarTA suppressed growth in 19 of the 24 pathovars from the P. syringae complex, whereas Psm and Pca grew and formed gray to black colonies. SPamt contained sucrose and peptone as nutrient sources and was supplemented with bromothymol blue and the same antibiotics present in DTarTA and Psm and Pca formed yellowish to dark brown colonies on the SPamt medium. SPbc contained sucrose and peptone and was supplemented with cephalexin and boric acid as antibiotics and Psm and Pca formed semi-translucent to white colonies on the SPbc medium. SPamt and SPbc suppressed the growth of several plant-associated bacteria (except the P. syringae complex). The growth of saprophytic bacteria in seeds on the different media was compared with that on King's B medium, using five types of commercially available Raphanus sativus seeds. The suppression rate of DTarTA was 85-99% and was lower for seeds with more saprophytic bacteria. The suppression rates of SPamt and SPbc were 90-99%. In detection tests using 10,000 seed samples mixed with Pca or Psm-contaminated seeds, it was possible to selectively isolate Psm and Pca using SPamt and SPbc, even when the colony numbers of the target bacterium constituted less than 10% of the total colonies. KEY POINTS: • Bacterial leaf spot and blight pathogens were selectively isolated from seeds. • DTarTA medium distinguishes these pathogens from P. syringae complex pathovars. • SPamp and SPbc media have different selectivity for plant-associated bacteria.

Epithelial folding is a universal biological phenomenon in morphogenesis, typical examples being brain gyri, villi of the intestinal tract, and imaginal discs in invertebrates. During epithelial morphogenesis, the physical constraints imposed by the surrounding microenvironment on epithelial tissue play critical roles in folding morphology. In this study, we focused on the asymmetry of the environmental constraints sandwiching the epithelial sheet and introduced the degree of asymmetry, which indicates whether the basal or apical side of the epithelium is closer to the constraint wall. Then, we investigated the relationship between the degree of asymmetry and epithelial folding morphology using three-dimensional vertex simulations. The results show that the folding patterns of the epithelial sheets change from spot patterns to labyrinth patterns and then to hole patterns as the degree of asymmetry changes. Furthermore, we examined the pattern formation in terms of the equation of out-of-plane displacement of the sheet derived from the mechanical energy functional.

We designed two novel primer sets based on the sequence of the hrpZ gene to detect Pseudomonas syringae pv. syringae, the causal agent of bacterial black node in barley and wheat. A conventional PCR using these primers amplified only P. syringae group III strains. The result of a nested PCR with the novel primers showed that P. syringae pv. syringae was detectible even when only one culturable cell was used as the PCR template. This highly sensitive PCR detected P. syringae in diseased parts of wheat and barley plants and in asymptomatic plants.

During morphogenesis in development, multicellular tissues deform by mechanical forces induced by spatiotemporally regulated cellular activities, such as cell proliferation and constriction. Various morphologies are formed because of various spatiotemporal combinations and sequences of multicellular activities. Despite its potential to variations, morphogenesis is a surprisingly robust process, in which qualitatively similar morphologies are reproducibly formed even under spatiotemporal fluctuation of multicellular activities. To understand these essential characteristics of tissue morphogenesis, which involves the coexistence of various morphologies and robustness of the morphogenetic process, in this study, we propose a novel approach to capture the overall view of morphogenesis from mechanical viewpoints. This approach will enable visualization of the energy landscape, which includes morphogenetic processes induced by admissible histories of cellular activities. This approach was applied to investigate the morphogenesis of a sheet-like tissue with curvature, where it deformed to a concave or convex morphology depending on the history of growth and constriction. Qualitatively different morphologies were produced by bifurcation of the valley in the energy landscape. The depth and steepness of the valley near the stable states represented the degree of robustness to fluctuations of multicellular activities. Furthermore, as a realistic example, we showed an application of this approach to luminal folding observed in the initial stage of intestinal villus formation. This approach will be helpful to understand the mechanism of how various morphologies are formed and how tissues reproducibly achieve specific morphologies.

Cells require mechanical forces for their physiological functions. The forces are generated mainly from molecular interactions between actin filaments, cross-linking proteins, and myosin motors in the actin cytoskeleton. To better understand the molecular interactions, many studies employed myosin motility assays with actin filaments propelled by myosin heads fixed on a surface. Various interesting behaviors of actin filaments have been observed in the motility assay experiments. Despite the popularity of the motility assays, there were only a few computational models designed for simulating the motility assay systems. Most of the previous models have limitations which precluded full understanding of molecular origins for behaviors of actin filaments. In this study, we used an agent-based computational model based on Brownian dynamics for simulating the motility assay system. Our model rigorously describes the mechanics, dynamics, and interactions of actin filaments, cross-linking proteins, and molecular motors. Using the model, we first investigated how properties of actin filaments and motors affect gliding motions of actin filaments without volume-exclusion effects as a base study. We found that actin filaments can continuously glide at relative fast speed only when they are sufficiently longer than the average spacing between neighboring motors and that the gliding speed of F-actins shows a biphasic dependence on processivity of motors. Then, we showed that volume-exclusion effects between actin filaments can induce diverse collective movements and alignment of actin filaments, thus creating thick bundles and ring-like structures in the absence of cross-linking proteins. Lastly, we demonstrated that cross-linking proteins can lead to distinct contractile behaviors of actin networks depending on the density and kinetics of the cross-linking proteins. Results from our study show the ability of our model to simulate the motility assay system under various conditions and provide insights into understanding of different behaviors of actin filaments.

This retrospective study was conducted to clarify the morphological characteristics of colorectal cancer (CRC) in Japanese familial adenomatous polyposis (FAP) patients.This study was carried out by the study group for FAP of the Japanese Society for Cancer of the Colon and Rectum. FAP patients who underwent surgical resection between 2000 and 2012 were included in the study.Of the 303 patients enrolled, 119 patients without CRC were excluded. Of 523 lesions, 49 lesions with missing morphological information were excluded; hence, only 474 CRC lesions in 178 patients (328 superficial lesions in 122 patients and 146 non-superficial lesions in 92 patients) were included in the study. Depressed lesions accounted for 3.0% of superficial lesions and ulcerated lesions accounted for 84.9% of non-superficial lesions. The depressed superficial lesions were observed only in patients with sparse and attenuated FAP (In patients with FAP, depressed superficial CRC lesions rarely developed but were detected in our study group, and ulcerated non-superficial CRC lesions were also present with similar ratios. Clinicians should pay attention to depressed superficial lesions during endoscopic surveillance of FAP patients.

Acute cholecystitis is an infectious disease of the gallbladder caused mainly by Escherichia coli, Klebsiella, and Enterococcus species. Streptococcus gallolyticus subsp. pasteurianus, previously known as Streptococcus bovis biotype II/2, rarely causes endocarditis, meningitis, and septicemia, mainly in children. Biliary tract infections by Streptococcus gallolyticus subsp. pasteurianus are extremely rare. There have been no reports of cases in Japan. Here, we describe the first case in Japan of acute calculous cholecystitis caused by Streptococcus gallolyticus subsp. pasteurianus infection. A 63-year-old man was admitted to our hospital with epigastric pain and vomiting. He had moderate tenderness and a full sensation in the epigastrium. Abdominal imaging revealed multiple stones in the gallbladder. After admission, he had a high fever that did not improve with antibiotics. Percutaneous transhepatic gallbladder drainage was performed. The patient underwent open cholecystectomy. During surgery, several small stones in the gallbladder and an abscess were observed at the gallbladder base. Streptococcus gallolyticus subsp. pasteurianus was detected by bacterial culture of the bile juice. The gallstones were bilirubin calcium stones. The endoscopic study showed three adenomas in the colon, but the histopathological examination demonstrated no malignant cells. Although infection by this bacterium may not be rare, this is the first reported case in Japan of acute calculous cholecystitis caused by Streptococcus gallolyticus subsp. pasteurianus infection.

Pseudomonas cannabina pv. alisalensis and P. syringae pv. maculicola cause bacterial leaf blight and bacterial leaf spot of crucifers (Brassicaceae). Both pathogens are threats to the cultivation of cruciferous crops. Here, we sequenced two strains of each pathogen, which will contribute to the development of countermeasures for the above diseases., Pseudomonas cannabina pv. alisalensis and Pseudomonas syringae pv. maculicola cause bacterial leaf blight and bacterial leaf spot of crucifers (Brassicaceae). Both pathogens are threats to the cultivation of cruciferous crops. Here, we sequenced two strains of each pathogen, which will contribute to the development of countermeasures for the above diseases.

Objective: Patients with BRAF-mutant colorectal cancer (CRC) have a poor prognosis. Molecular status is not currently used to select which drug to use in combination with radiotherapy. Our aim was to identify drugs that radiosensitise CRC cells with known BRAF status. Methods: We screened 298 oncological drugs with and without ionising radiation in colorectal cancer cells isogenic for BRAF. Hits from rank product analysis were validated in a 16-cell line panel of human CRC cell lines, using clonogenic survival assays and xenograft models in vivo. Results: Most consistently identified hits were drugs targeting cell growth/proliferation or DNA damage repair. The most effective class of drugs that radiosensitised wild-type and mutant cell lines was PARP inhibitors. In clonogenic survival assays, talazoparib produced a radiation enhancement ratio of 1.9 in DLD1 (BRAF-wildtype) cells and 1.8 in RKO (BRAF V600E) cells. In DLD1 xenografts, talazoparib significantly increased the inhibitory effect of radiation on tumour growth (P ≤ 0.01). Conclusions: Our method for screening large drug libraries for radiosensitisation has identified PARP inhibitors as promising radiosensitisers of colorectal cancer cells with wild-type and mutant BRAF backgrounds.

Oscillatory flows of actomyosin play a key role in the migration of single cells in culture and in collective cell movements in Drosophila embryos. In vertebrate embryos undergoing convergent extension (CE), the Planar Cell Polarity (PCP) pathway drives the elongation of the body axis and shapes the central nervous system, and mutations of the PCP genes predispose humans to various malformations including neural tube defects. However, the spatiotemporal patterns of oscillatory actomyosin contractions during vertebrate CE and how they are controlled by the PCP signaling remain unknown. Here, we address these outstanding issues using a combination of in vivo imaging and mathematical modeling. We find that effective execution of CE requires alternative oscillations of cortical actomyosin across cell membranes of neighboring cells within an optimal frequency range. Intriguingly, temporal and spatial clustering of the core PCP protein Prickle 2 (Pk2) is correlated to submembranous accumulations of F-actin, and depletion of Pk2 perturbs the oscillation of actomyosin contractions. These findings shed light on the significance of temporal regulation of actomyosin contraction by the PCP pathway during CE, in addition to its well-studied spatial aspects.

Bacterial black node, a disease of barley and wheat, is caused by Pseudomonas syringae pv. syringae (synonym pv. japonica, hereafter Psj). Here a selective medium, serine-potassium tellurite-based Psj-selective agar (SPTPsjA) was developed to isolate and detect Psj. After 5–7 days on SPTPsjA at 25 °C, characteristic black Psj colonies formed with an efficiency equal to that on potato–peptone–glucose agar. Except for P. viridiflava, P. cichorii, and some strains of P. syringae group bacteria, SPTPsjA either inhibited growth of other phytopathogenic bacteria or enabled their discrimination. To facilitate diagnosis of Psj, we prepared three antisera against three strains; in a slide agglutination test, the antisera reacted to all tested strains of Psj and failed to react to P. viridiflava and P. cichorii although some strains of other P. syringae group bacteria were indistinguishable from Psj with SPTPsjA and antisera. When barley and wheat seeds from fields in which bacterial black node had occurred were placed on SPTPsjA in the dark at 25 °C for 7 days, black colonies grew on the medium around the seeds. Among these strains, only those responsive to the above antisera were pathogenic on barley. These results show that the combination of SPTPsjA and antisera can isolate Psj from seeds of barley and wheat.

Culturable bacteria were isolated from tomato seeds using media selective for the canker pathogen Clavibacter michiganensis subsp. michiganensis. Clustering analysis (>99% identity) revealed the presence of 16 operational taxonomic units (OTUs) among isolates detected on semi-selective media. Three OTUs belonged to the phylum Actinobacteria, including those of Micrococcus and Dermacoccus, and 13 OTUs belonged to the phylum Firmicutes, including Bacillus and related genera. These Gram-positive endophytic bacteria have the potential to provide false-positive results in seed health tests using media considered semi-selective for the cancer pathogen.

Black rot and bacterial spots threaten the cultivation of cruciferous vegetables worldwide, and the development of a method that can easily detect, identify, and distinguish their respective pathogens Xanthomonas campestris pv. campestris (Xcc) and X. campestris pv. raphani (Xcr) is required. Multiple whole-genome sequences of Xcc and Xcr were aligned to identify specific regions and subsequently design gene markers. A region present in Xcr, but absent in Xcc, was detected, which was approximately 11.5 kbp in length, sandwiched between the serine protease homolog (SPH) and nicotinate phosphoribosyltransferase gene (pncB). It contained putative cellulose synthesis-related genes, whereas Xcc only had a modified cellulose synthase gene. Designed primers were pncB_fw1 and pncB_fw2 (from the pncB gene), Xcc_rv1 and Xcc_rv2 (from the modified cellulose synthesis gene), and Xcr_rv1 and Xcr_rv2 (from the putative first and second open reading frames of the gene cluster). PCR using pncB_fw1 and Xcc_rv1, or pncB_fw2 and Xcc_rv2, amplified DNA fragments only in Xcc and X. campestris pv. incanae (Xci). Xci is the causal agent of black rot of garden stock and closely related to Xcc. PCR using pncB_fw1 and Xcr_rv1, or pncB_2 and Xcr_rv2, amplified DNA fragments only in Xcr. Multiplex PCR analysis easily distinguished Xcc and Xcr from bacterial colonies isolated on growth media and detected the pathogen in symptomatic leaves. Multiplex nested PCR detected the contamination of one seed with Xcc and/or Xcr infection from 1000 seeds. Therefore, the PCR primers designed in this study therefore helped detect and discriminate between Xcc and Xcr. KEY POINTS: • Xanthomonas campestris pv. campestris (Xcc) and pv. raphani (Xcr) were investigated. • Novel primers were designed following whole-genome comparison analyses. • Multiplex PCR with new primers distinguished Xcc and Xcr simultaneously.

Xanthomonas campestris pv. raphani and X. campestris pv. campestris are the causal bacteria of bacterial spot and black rot of crucifers ( Brassicaceae ), respectively. Both pathogens are threats in the cultivation of cruciferous crops such as cabbage. Here, we sequenced a strain of each of these pathogens.

Accumulating evidence suggests that overexpression of heat shock protein 47 (HSP47) increases cancer progression, and that HSP47 level in the tumor-associated stroma may serve as a diagnostic marker in various cancers. The present study aimed to evaluate whether HSP47 gene expression in colorectal cancer (CRC) tissues could be used to identify lymph node (LN) metastasis status preoperatively in patients with CRC. To do so, HSP47 gene expression was determined and its association with the clinicopathological characteristics of patients with CRC was analyzed. A total of 139 surgical specimens from patients with CRC and 36 patients with benign colonic disease undergoing surgery at Mie University Hospital were analyzed. HSP47 gene expression was determined by reverse transcription quantitative PCR using Power SYBR Green PCR methods. Expression level of HSP47 was significantly higher in CRC tissues compared with normal tissue from patients with benign colonic disease. Furthermore, high HSP47 expression was significantly associated with tumor progression, including high T stage, lymph node metastasis and venous invasion, and high TNM stage. High HSP47 expression may therefore serve as a novel predictive biomarker for determining patients with CRC and LN metastasis. According to Kaplan-Meier analysis, patients with high HSP47 expression level had significantly poorer overall survival than those with low HSP47 expression level. Furthermore, multivariate analyses identified HSP47 expression as an independent predictive marker for LN metastasis and poor overall survival in patients with CRC. In summary, the present study demonstrated that HSP47 expression may be considered as a novel biomarker for predicting LN metastasis status and prognosis in patients with CRC.

The beetle horn primordium is a complex and compactly folded epithelial sheet located beneath the larval cuticle. Only by unfolding the primordium the complete 3D shape of the horn appears, suggesting that the morphology of beetle horns is coded in the primordial folding pattern. To decipher the folding pattern, we have developed a method to manipulate the primordial local folding, reproduced it on a computer, and clarified the contribution of the folding of each primordium region to transformation. We found that the three major morphological changes (branching of distal tips, proximodistal elongation, and angular change) were caused by the folding of different regions, and that the folding mechanism was also different depending on the region. The computational methods we used are applicable to the morphological study of other exoskeletal animals.

Study design: The present study is a single-center retrospective cohort study. Objective: The objective of the study is to verify the hypothesis that sharp elevation of systolic blood pressure at extubation is a risk factor for development of symptomatic epidural hematoma after spinal surgery. Summary of background data: Postoperative symptomatic epidural hematoma (she) after spine surgery is a rare but potentially serious complication that may result in paralysis unless removed at an early stage. Methods: The subjects were 2611 patients treated with decompression and decompression/fusion of the cervical, thoracic, and lumbar vertebrae at our hospital. Twelve of these patients developed postoperative SEH and removal of hematoma was performed. To investigate the risk factors in these patients, data were analyzed for age at the time of surgery, sex, preoperative complications, medical history, body mass index, preoperative platelet count, surgical procedure, microscope use, operative time, blood loss, surgical site, systolic blood pressure (SBP) at extubation, difference between resting and extubation SBP, ratio of SBP at extubation to that at rest, blood pressure at discharge from the operating room, and use of a drain. Results: There was a significantly higher rate of SBP ratio (extubation/rest) ≥1.3 in patients with SEH ( p = 0.021, Fisher’s exact test). Among the preoperative complications and medical histories, the frequency of cerebrovascular disorder tended to be higher in SEH cases than in non-SEH cases ( p = 0.073). There was no significant difference for all other parameters listed above. In multivariate logistic analysis, the odds ratios were 3.98 ( p = 0.018) for an SBP ratio (extubation/rest) ≥1.3 and 4.75 ( p = 0.055) for cerebrovascular disorder, suggesting effects of these two items. With simultaneous input into a multivariate model, SBP ratio ≥1.3 had a significant independent association with postoperative SEH ( p = 0.021) and cerebrovascular disorder showed a tendency for this association ( p = 0.072). Conclusion: The risk for symptomatic epidural hematoma is significant in patients with SBP at extubation that is more than 1.3 times that of SBP at rest.

Insects can dramatically change their outer morphology at molting. To prepare for this drastic transformation, insects generate new external organs as folded primordia under the old cuticle. At molting, these folded primordia are physically extended to form their final outer shape in a very short time. Beetle horns are a typical example. Horn primordia are derived from a flat head epithelial sheet, on which deep furrows are densely added to construct the complex folded structure. Because the 3D structure of the pupa horn is coded in the complex furrow pattern, it is indispensable to know how and where the furrows are set. Here, we studied the mechanism of furrow formation using dachsous (ds) gene knocked down beetles that have shorter and fatter adult horns. The global shape of the beetle horn primordia is mushroom like, with dense local furrows across its surface. Knockdown of ds by RNAi changed the global shape of the primordia, causing the stalk region become apparently thicker. The direction of cell division is biased in wildtype horns to make the stalk shape thin and tall. However, in ds knocked down beetles, it became random, resulting in the short and thick stalk shape. On the other hand, a fine and dense local furrow was not significantly affected by the ds knockdown. In developing wildtype horn primordia, we observed that, before the local furrow is formed, the apical constriction signal emerged at the position of the future furrow, suggesting the pre-pattern for the fine furrow pattern. According to the results, we propose that development of complex horn primordia can be roughly divided to two distinct processes, 1) development of global primordia shape by anisotropic cell division, and 2) local furrow formation via actin-myosin dependent apical constriction of specific cells.

Isolated avulsion fracture of the peroneus longus tendon insertion at the base of the first metatarsal without injury of the tarsometatarsal joint is very rare. Similar to most avulsion fractures, this type of injury is caused by strong tension exerted by the peroneus longus tendon. The mechanism leading to this lesion and treatment options are not clearly defined. Several surgical techniques have been advocated for this fracture, including excision of an avulsion fragment and open reduction for internal fixation through the medial aspect of the foot or minimal plantar incision. We have described a method of percutaneous fixing of the avulsion fracture at the plantar lateral base of the first metatarsal using the ZipTight Fixation System (Zimmer Biomet Warsaw, Indiana, USA), which offers the advantage of allowing a rigid fixation and minimal invasive surgical technique for a small fragment.

This study demonstrates computational simulations of multicellular deformation coupled with chemical patterning in the three-dimensional (3D) space. To address these aspects, we proposes a novel mathematical model, where a reaction–diffusion system is discretely expressed at a single cell level and combined with a 3D vertex model. To investigate complex phenomena emerging from the coupling of patterning and deformation, as an example, we employed an activator–inhibitor system and converted the activator concentration of individual cells into their growth rate. Despite the simplicity of the model, by growing a monolayer cell vesicle, the coupling system provided rich morphological dynamics such as undulation, tubulation, and branching. Interestingly, the morphological variety depends on the difference in time scales between patterning and deformation, and can be partially understood by the intrinsic hysteresis in the activator-inhibitor system with domain growth. Importantly, the model can be applied to 3D multicellular dynamics that couple the reaction–diffusion patterning with various cell behaviors, such as deformation, rearrangement, division, apoptosis, differentiation, and proliferation. Thus, the results demonstrate the significant advantage of the proposed model as well as the biophysical importance of exploring spatiotemporal dynamics of the coupling phenomena of patterning and deformation in 3D space.

Ralstonia solanacearum, the causal agent of bacterial wilt of tomato, grows in infected plants and migrates from the roots into the soil. We investigated the effectiveness of bacterial wilt-resistant tomato rootstock in reducing the migration of R. solanacearum from susceptible scions into the soil. Rootstock stems were either 3–5 cm tall (low-grafted, LG) or ≥ 10 cm tall (high-grafted, HG). After inoculation of scions of the susceptible cultivar (SC) with R. solanacearum below the first flower, there was no difference in disease progression among LG, HG, and ungrafted SC plants, and plants had wilted by 2 weeks. However, the rate of detection of R. solanacearum in the soil of wilted plants was reduced by grafting. The size of the R. solanacearum population in the soil of fully wilted plants increased in the order of HG

Summary Background Preoperative hypoalbuminemia is a well-known risk factor for anastomotic leakage after colorectal surgery, but the association between perioperative albumin level and anastomotic leakage has not been fully investigated in curative colorectal cancer (CRC) patients. Methods In total, 200 CRC patients (Stage I–III) undergoing curative laparoscopic surgery between January 2004 and December 2013 were enrolled in this study. We collected data on surgical factors, perioperative levels of serum albumin and inflammatory markers, and perioperative factors affecting hypoalbuminemia from 196 CRC patients to assess the relation to anastomotic leakage. Results Anastomotic leakage occurred in 11 cases (5.6%) and the frequency was higher in rectal cancer patients ( p = 0.0044). There was no significant difference of preoperative serum albumin level between the anastomotic leakage group (AL) and the nonanastomotic leakage group (NAL). Postoperative serum albumin levels in AL were significantly lower than in NAL [postoperative day (POD) 0, p = 0.0004; POD1, p = 0.0001; POD3, p = 0.0004; and POD7, p = 0.0021]. On multivariate analysis, lower average level of serum albumin on POD1 and POD3 {odds ratio (OR) [95% confidence interval (CI)] = 7.53 (1.60–55.80), p = 0.0095}, higher average level of serum white blood cells on POD1 and POD3 [OR (95% CI) = 7.24 (1.40–59.25), p = 0.0165], and surgery for rectal cancer [OR (95% CI) = 15.18 (3.26–93.99), p = 0.0004] were independent risk factors for anastomotic leakage. Conclusion Lower early postoperative serum albumin levels are a potentially valuable indicator of anastomotic leakage in CRC patients undergoing curative surgery.

We examined the potential for biological control of black rot of broccoli, caused by Xanthomonas campestris pv. campestris (Xcc), using nonpathogenic Xanthomonas sp. strain 11-100-01 (npX) mixed with bacteriophage XcpSFC211 (pXS). Inoculation of intact broccoli plants in greenhouse trials with either npX or pXS did not control black rot. After injured plant inoculation, however, npX alone or npX with pXS significantly controlled black rot. When a mixed suspension of npX with pXS was placed on a membrane filter, then washed with distilled water and air-dried, a substantial amount of pXS adsorbed to the surface of npX. In a field trial, broccoli plants were sprayed with a suspension of npX with pXS, then inoculated with Xcc. A meta-analysis of the results from five field trials showed an integrated risk ratio (IRR, the ratio of disease incidence in inoculated broccoli plants to the incidence in control plants) of 0.69 after treatment with only npX and 0.59 with npX with pXS, indicating that black rot incidence was significantly reduced by each treatment. The difference between these two treatments was also significant. IRR was 1.24 when comparing suppression by npX with pXS and that by basic copper sulfate wettable powder; thus, their control was comparable. The combination of npX with pXS improved the preventive effect against black rot. This is the first report describing that a nonpathogenic Xanthomonas sp. strain mixed with a bacteriophage effectively controlled black rot of broccoli in field trials.

Solitary mediastinal lymph node metastasis of hepatocellular carcinoma (HCC) is rare. We report a case of metachronically solitary mediastinal metastases of HCC treated by video-assisted thoracic surgery (VATS) twice. A 66-year-old man underwent repeated laparoscopic radiofrequency ablation or trans-arterial catheter chemo-embolization against HCC for more than 10 years. The level of alpha fetoprotein protein was elevated, and radiological modalities including FDG-PET revealed solitary mediastinal tumor metachronically. VATS was performed bilaterally twice. The postoperative course was uneventful and there had no recurrence of extra-hepatic metastases and tumor markers are within normal limits at 18 months after second VATS. VATS is a minimally invasive and useful procedure for solitary mediastinal lymph node metastasis of HCC. If primary HCC was controlled and lymph node metastasis was solitary, mediastinum lymphadenectomy using VATS might give good short and long term results.

Background/Aims: Pouchitis is one of the main complications after ileal pouch-anal anastomosis in patients with ulcerative colitis. The aim of this study was to determine whether the use of colonic histological criteria can predict the development of pouchitis. Methodology: We retrospectively reviewed 147 patients' clinical data and performed a histological evaluation of the resected total colon using Tanaka's criteria, which comprise the following 6 factors: ulceration (H1), crypt abscesses (H2), degree of mononuclear cell infiltration (MNCI) (H3), segmental distribution of MNCI (H4), eosinophil infiltration (H5), and extent of disease of resected colon (H6). Results: The development of pouchitis and chronic pouchitis within 3 years after restoration of gastrointestinal continuity was recognized in 52 (35.4%) and 26 (17.7%) of the 147 patients, respectively. Using various combinations of each score, the H3 + H4 - H5 scores of patients with pouchitis or chronic pouchitis were significantly higher than those of patients without. A H3 + H4 - H5 score of >0.4 was a statistically significant risk factor for the development of both pouchitis and chronic pouchitis. Conclusions: The combination of the degree of MNCI, segmental distribution of MNCI, and eosinophil infiltration from histological criteria has utility in predicting the future development of pouchitis, especially chronic pouchitis.

Ileal pouch‐anal anastomosis (IPAA) after total proctocolectomy (TPC) can be conducted with either hand‐sewn or stapled anastomosis for patients with familial adenomatous polyposis (FAP). Although stapled IPAA without mucosectomy has a higher risk for developing adenomas in the remnant mucosa, it is the simpler procedure with potential benefit in short‐term outcomes. However, it remains controversial as to whether stapled IPAA has any advantages in reducing postoperative complications. The aim of the present study was to compare the postoperative complications and short‐term outcomes of stapled and hand‐sewn IPAA for patients with FAP, using a multicenter cohort sample in Japan. Data of 143 patients with FAP who underwent TPC with stapled IPAA (n=37) and hand‐sewn IPAA (n=106) at 23 institutions between 2000 and 2012 were collected. Postoperative complications, proportion of ostomy, fecal continence and overall survival were compared. Overall rates of the Clavien‐Dindo grade II‐IV complications were not different between the two groups (19% in stapled vs 25% in hand‐sewn, P=.42), with significantly fewer pouch‐related complications including leakage, pelvic abscess, vaginal fistula and anastomotic stricture in stapled IPAA (none in stapled vs 11% in hand‐sewn, P=.036). There was no mortality. Proportion of ostomy at 12 months was similar (2.7% in stapled vs 4.3% in hand‐sewn, P=.26). Mean Wexner score was similar. (0.47 in stapled vs 2.0 in hand‐sewn, P=.12). Five‐year overall survival excluding Stage IV patients was 96% in both groups. Stapled IPAA is a safe option in patients with FAP with a potential benefit in reducing pouch‐related complications.

Aim To identify predictors of poor prognosis of patients with colon cancer (CC) who underwent surgery with curative intent, we investigated the association between the albumin to globulin ratio (AGR) with clinicopathological findings such as overall (OS) and disease-free (DFS) survival. Patients and methods We conducted a retrospective study of clinicopathological findings, including preoperative laboratory data, for 248 patients with stage I-III CC. Results Patients with low AGR had shorter DFS and OS compared to those with high AGR. Multivariate analyses identified low AGR as an independent variable independently associated with recurrence and poor prognosis of patients with CC who underwent surgery with curative intent regardless of lymphnode metastasis. Conclusion The preoperative AGR was an independent predictor of recurrence and poor prognosis of patients with CC who underwent surgery with curative intent. The AGR indicates that these patients may benefit from intensive adjuvant therapy.

Although patients with metastatic colorectal cancer (CRC) are often unable to undergo treatment after resection of primary tumors, identifying such patients before surgery is not easy. In this study, we evaluated the association among clinicopathological findings, survival outcomes, and ability to undergo multimodal therapy after primary tumor resection in patients with Stage IV CRC. We collected clinicopathological findings and preoperative laboratory data, including carcinoembryonic antigen (CEA) and systemic inflammatory response markers for 92 patients who were treated for Stage IV CRC between 2005 and 2014. We used multivariate analysis on factors that affect prognosis and ability to undergo postoperative treatment. Postoperative multimodal therapy improved overall survival (OS) significantly. Among serum markers, elevated CEA, neutrophil-to-lymphocyte ratio, and modified Glasgow prognosis score (mGPS) were significant indicators of shorter OS. In multivariate analysis, low performance status (P = 0.003), undifferentiated histology type (P = 0.019), and elevated mGPS (P = 0.042) were independent predictors of worse prognosis; and older age (P = 0.016), right-sided colon cancer (P = 0.043), and elevated mGPS (P = 0.031) were independent risk factors for difficulty of introducing postoperative multimodal therapy. Preoperative mGPS is a useful objective indicator for CRC patients with multiple metastases who are able to undergo primary site resection followed by postoperative multimodal therapy.

Surgical site infection (SSI) is a potentially morbid and costly complication of surgery. The purpose of this study was to determine preoperative and operative predictors of SSIs after gastric resection with lymphadenectomy in patients with gastric cancer (GC). Data on clinicopathologic factors, including operative and preoperative laboratory factors, for 384 patients with GC who had undergone curative surgery were analyzed in this retrospective study to assess their associations with SSIs. Superficial/deep incisional SSIs (iSSIs) and organ/space SSIs (o/sSSIs) occurred in 18 (4.6%), and 27 (7.0%), respectively. The o/sSSIs were significantly associated with surgery-related factors such as duration of operation, blood loss, and extent of tumor. Additionally, high levels of preoperative indicators of systemic inflammation, including neutrophil counts, neutrophil/lymphocyte ratio, and C-reactive protein concentrations, were significantly associated with o/sSSIs. Multivariate analyses demonstrated that preoperative neutrophil counts and duration of surgery were independent predictors for o/sSSIs, whereas only preoperative serum albumin concentration was predicted for iSSIs. In patients with GC undergoing curative surgery, preoperative neutrophil count and operation time are potentially valuable predictors of o/sSSIs, whereas only preoperative serum albumin predicts iSSIs.

Familial adenomatous polyposis (FAP)-associated desmoid tumor (DT) is sometimes life threatening. However, the optimal treatment for DTs has not been established. The aim of this study was to analyze the outcomes of surgical and pharmacological treatments for DT in Japanese FAP patients.We retrospectively reviewed the data of 303 patients who underwent colectomy for FAP between 2000 and 2012. We analyzed 41 patients with DTs in which the location was apparent. The selection of treatment for intra-abdominal DTs was also evaluated according to Church's classification.Surgery was frequently used to treat extra-abdominal DTs. Multimodal treatments, including surgery, and the administration of non-steroidal anti-inflammatory drugs, hormonal therapy, and chemotherapy were widely used for intra-abdominal DTs. The most effective pharmacological treatment was cytotoxic chemotherapy, which was associated with a response rate of 45.5% and a disease control rate of 72.7%. After a median follow-up period of 53.0 months, the 5-year DT-specific survival rate in patients with stage IV disease was 71.4%; in contrast, the rate in patients with other stages was 100%. Four-stage IV patients died of DT due to uncontrollable rapid progression. No cytotoxic chemotherapy was administered; however, incomplete resection was performed in three cases.Our findings will provide clues that may help physicians in selecting the optimal strategy for this rare disease.

We report a patient who experienced colonic necrosis after laparoscopic high anterior resection for sigmoid colon cancer, and review the literature to evaluate the clinical features of colonic necrosis following surgery for sigmoid colon and rectal cancer. A 76-year-old man with sigmoid colon cancer underwent laparoscopic high anterior resection. The operation included high ligation of the inferior mesenteric artery and end-to-end anastomosis using circular staples. Pathology findings revealed a pT4N2M0 lesion. Beginning on postoperative day (POD) 1, the patient experienced a high, spiking fever, and gradually developed leukocytosis and high inflammatory condition. The patient complained of abdominal distention, but had no signs of peritonitis. Abdominal computed tomography on POD4 showed wall thickness of the proximal colon from the anastomosis site and ascites with free air. An anastomotic leakage was suspected. Emergency laparotomy revealed a disrupted anastomosis without feces in the abdomen and a gangrenous 15 cm segment of the colon proximal to the anastomosis. The affected area of the colon was excised and Hartmann's procedure was performed. His postoperative period was uneventful. Our review of the literature demonstrates that elderly male patients with cardiovascular and pulmonary complications undergoing laparoscopic sigmoid and rectal cancer surgery with high ligation have high risk of postoperative colonic necrosis. We experienced colonic necrosis following laparoscopic high anterior resection for sigmoid colon cancer and required immediate resection. Elderly male patients with cardiovascular and pulmonary complications undergoing laparoscopic sigmoid and rectal cancer surgery with high ligation should be carefully monitored for postoperative colonic necrosis.

The discovery of biomarkers to predict the potential for lymph node (LN) metastasis in patients with colorectal cancer (CRC) is essential for developing improved strategies for treating CRC. In the present study, they used isobaric tags for relative and absolute quantitation to conduct a proteomic analysis designed to identify novel biomarkers for predicting LN metastasis in patients with CRC. They identified 60 differentially expressed proteins specifically associated with LN metastasis in CRC patients and classified the molecular and functional characteristics of these proteins by bioinformatic approaches. A literature search led them to select heat shock protein 47 (HSP47) as the most suitable candidate biomarker for predicting LN metastasis. Validation analysis by immunohistochemistry showed that HSP47 expression in patients with CRC and the number of HSP47-positive spindle cells in the tumor stroma were significantly higher compared with those in adjacent normal colonic mucosa, and the number of the latter cells increased with tumor progression. Further, the number of HSP47-positive spindle cells in stroma was a more informative marker for identifying LN metastasis than HSP47expression. Multivariate analysis identified spindle cells that expressed elevated levels of HSP47 as an independent predictive biomarker for CRC with LN metastasis. Moreover, these cells served as an independent marker of disease-free and overall survival of patients with CRC. Their data indicate that the number of HSP47-positive spindle cells in the stroma of CRC may serve as a novel predictive biomarker of LN metastasis, early recurrence and poor prognosis.

This pilot study aimed to develop a new technique, complete laparoscopic total mesorectal excision (TME) with an intersphincteric resection (ISR) and coloplasty pouch anal anastomosis to avoid any further abdominal incision other than laparoscopic port sites, and to assess the impact on short-quality of life and oncological outcomes of this technique. After laparoscopic TME, large bowel was dissected at the level of the promontory. Then, laparoscopic construction of the coloplasty pouch was performed. Simultaneously, a rectal specimen with ISR was excised using the transanal approach. Coloplasty pouch was gently pulled from pelvic thorough anal and a hand-sewn coloplasty pouch anal anastomosis was created. We had performed 8 surgeries using the new technique. Though one patient developed pelvic infections, but intestinal continuity could be maintained and no local and distant recurrence was recognized in other patients. We foresee this novel approach to have significant clinical potential for lower rectal cancer patients with ISR.

Objectives: Anti-tumor necrosis factor (TNF) antibodies have shown efficacy in the prevention of recurrence of Crohn's disease after intestinal resection. However, some patients develop surgical recurrence despite this therapy. We aimed to evaluate the risk factors for recurrence of Crohn's disease requiring surgery while receiving post-operative anti-TNF therapy. Methods: We performed a retrospective evaluation of 164 patients who had received post-operative anti-TNF maintenance therapy between 2002 and 2016. We classified Crohn's disease-related re-operation as surgical recurrence and analyzed its risk factors using the Cox proportional hazard model. Results: Of the 164 participants, 128 had received infliximab and 36 had received adalimumab maintenance therapy. We obtained follow-up data over a mean of 60.2 months. The proportion of patients with surgical recurrence at 5 years was 14.9%. The only independent risk factor for surgical recurrence, which we identified was post-operative smoking habit (odds ratio, 5.03; 95% CI, 1.14-12.8; P=0.033). Conclusions: Post-operative smoking may be a significant risk factor for post-operative surgical recurrence of Crohn's disease while receiving anti-TNF maintenance therapy.