Generation of conditional ALK F1174L mutant mouse models for the study of neuroblastoma pathogenesis.

Neuroblastoma, an embryonal tumor arising from the sympathetic ganglia and adrenal medulla, is among the most intractable pediatric cancers. Although a variety of genetic changes have been identified in neuroblastoma, how they contribute to its pathogenesis remains largely unclear. Recent studies have identified alterations of the anaplastic lymphoma kinase (ALK) gene in neuroblastoma; ALK F1174L (a phenylalanine-to-leucine substitution at codon 1174) represents one of the most frequent of these somatic mutations, and is associated with amplification of the MYCN gene, the most reliable marker for the poor survival. We engineered the mouse Alk locus so that ALK F1174L is expressed by its endogenous promoter and can be induced in a spatiotemporally controlled fashion using Cre-loxP system. Although expression of ALK F1174L resulted in enhanced proliferation of sympathetic ganglion progenitors and increased the size of the sympathetic ganglia, it was insufficient to cause neuroblastoma. However, lethal neuroblastoma frequently developed in mice co-expressing ALK F1174L and MYCN, even in a genetic background where MYCN alone does not cause overt tumors. These data reveal that physiological expression of ALK F1174L significantly potentiates the oncogenic ability of MYCN in vivo. Our conditional mutant mice provide a valuable platform for investigating the pathogenesis of neuroblastoma.

Risk factors for unsuccessful atropine therapy in hypertrophic pyloric stenosis.

BACKGROUND: I.v. atropine (IA) for infantile hypertrophic pyloric stenosis (IHPS) is a good alternative to pyloromyotomy but has not been broadly accepted. The lower success rate is one of the greatest disadvantages of IA. We investigated the risk factors for unsuccessful results following IA for IHPS. METHODS: Medical records of patients with IHPS who were admitted to Kimitsu Chuo Hospital between 2002 and 2016 and were initially given atropine sulfate were retrospectively reviewed. Atropine was given i.v. (0.1 mg/kg/day in eight divided doses). Oral feeding of milk was started with a small amount and increased in a stepwise fashion to full feed. IA therapy was defined as unsuccessful in the presence of projectile vomiting more than three times a day or intolerance to a predetermined amount of milk. RESULTS: Of the 48 patients with IHPS, 33 patients were successfully treated with IA and 15 patients needed surgical intervention. On univariate analysis the risk factors for unsuccessful IA therapy were younger age, lower bodyweight, and shorter duration of symptoms before diagnosis. On multivariate analysis age at diagnosis < 30 days was the only significant risk factor for unsuccessful IA therapy (OR, 5.7 l P = 0.03). CONCLUSIONS: Age at diagnosis < 30 days is a risk factor for unsuccessful IA therapy in IHPS. This might be considered when IA therapy is used for neonates with IHPS.

Subjective Symptoms Linked to Sleep Duration: An Analysis from Japanese National Statistics.

BACKGROUND: There is a high prevalence of sleep disorders in Japan, and they are a factor in a decreased quality of life. The main objective of this study was to clarify the background factors of sleep disorders that affect sleep duration, such as subjective symptoms and working hours. METHODS: We performed a cross-sectional study on the Japanese national statistics data. Answers from a household questionnaire were used to analyze risk factors for decreases in sleep duration. The subjects were a total of 3972 men and women aged 40-59 years, the age group that forms the core of the working population. For the analysis, a univariate analysis (contingency table) between sleep duration (two groups: sleep duration ≥ 6 h and <6 h) and 42 subjective symptoms was carried out. A multivariate analysis (binomial logistic regression) was conducted using sleep duration and subjective health assessment as objective variables, and odds ratios (ORs) adjusted for sex, working hours, and other factors were obtained. RESULTS: The univariate analysis by subjective symptom showed significant ORs for eight symptoms, including poor sleep quality (OR: 2.24), constipation (OR: 2.24), and dizziness (OR: 1.77). In the multivariate analysis, the model with sleep duration as the objective variable showed significantly adjusted ORs for four variables, including constipation (1.72) and poor sleep quality (1.66). The model with subjective health assessment as the objective variable showed significantly adjusted ORs for eight variables, including dizziness (4.18), while poor sleep quality (1.45) was not significant. CONCLUSIONS: The present results suggest the presence of subjective symptoms that may be inferred to be related to decreases in sleep duration.

Use of multi-detector row CT for postoperative follow-up of biliary atresia patients with sequelae.

BACKGROUND/AIMS: To evaluate the usefulness of multi-detector row computed tomography (MDCT) in the postoperative long-term follow-up of biliary atresia (BA) patients for detection of esophagogastric varices due to portal hypertension and intrahepatic bile duct (IHBD) dilatations. Well-developed esophagogastric varices can cause unexpected life-threatening rupture and contribute to the progression of hepatopulmonary vascular diseases. Dilated IHBDs can trigger cholangitis. METHODS: After corrective surgery, 29 BA patients (median age 14 years, range 6 months to 27 years) with suspected long-term sequelae underwent 16-slice MDCT. Esophagogastric varices and IHBD dilatations were evaluated using reconstructed images. Detection rates for esophagogastric varices were compared between MDCT and endoscopic findings. Differences in detection rates for IHBD dilatations between MDCT and ultrasonography (US) were also investigated. RESULTS: Detection rates for esophageal varices did not differ significantly between MDCT and endoscopy, while three cases positive only on MDCT, including one with hepatopulmonary syndrome, showed conspicuous peri-esophageal varices deep in the esophageal wall. MDCT showed a significantly higher detection rate than endoscopy (p = 0.03) for gastric fundal varices. MDCT with the maximum intensity projection disclosed IHBD dilatations in 17 postoperative BA cases, 15 out of which had past history of cholangitis, while US detected them in 6 cases (p = 0.003). In addition, MDCT was more advantageous for detecting IHBD dilatations, particularly those located in the posterior lobe. CONCLUSIONS: Among postoperative BA patients, compared to endoscopy, MDCT had comparable and superior detection rates for esophageal and gastric fundal varices, respectively. MDCT also precisely demonstrated the location and extent of IHBD dilatations. In long-term follow-up of BA patients, MDCT can help determine follow-up strategies.

Relationship between the Appearance of Symptoms and Hospital Visits in Childhood Based on Japanese Statistical Data.

BACKGROUND: Childhood health problems affect healthy growth. This study aimed to assess the symptoms and diseases requiring hospital visits commonly found in children in Japan and analyze their effects on health status. METHODS: Anonymized data on 1315 children aged 6-14 years were obtained from a national survey questionnaire. The survey items addressed symptoms, disease names, and hospital visits. Associations between symptoms and other factors were examined by means of a contingency table analysis and logistic regression. RESULTS: The proportions of responses for health status were compared for each question item; significant differences were found in age group (p < 0.01), subjective symptoms (p < 0.01), hospital visits (p < 0.01), and lifestyle (p < 0.01). The proportion of responses indicating "poor" perceived health status was high among those with subjective symptoms (4.8%) and hospital visits (4.7%). From the logistic regression, significant odds ratios were found for subjective symptoms (2.10, 95% confidence interval (C.I.) 1.15-3.83) and age group (1.98, 95% C.I. 1.05-3.72). CONCLUSION: Among measures to improve quality of life from childhood, comprehensive health guidance that emphasizes understanding symptoms and includes age and living conditions is important.

Factors Affecting Dietary Improvements in Elderly Residents of Long-Term Care Institutions Receiving Domiciliary Dental Care.

Background: Oral disabilities occur due to tooth loss. This study aimed to investigate oral and systemic factors related to diet in elderly residents receiving domiciliary dental care. Methods: The subjects were 74 consenting residents. Survey items included whether subjects could eat independently and diet type. Subjects were examined by the dentist for the number of teeth, occlusal support index, and wearing dentures. Contingency table analysis was performed to determine what levels of decline in general and oral functions led to difficulties eating a normal diet. Results: There was a significant difference in the mean number of activities of daily living (ADL) requiring assistance evident between subjects eating a normal diet and those eating fluid boiled rice (p < 0.01). A comparison of occlusal support and diet type showed that most subjects who ate a soft diet or gruel had no occlusal support. Almost all subjects who ate a normal diet wore dentures. However, only 38% of subjects eating a soft diet and 40% of those eating gruel did wear dentures; both group differences were significant (p < 0.01). Conclusions: Future studies need to further investigate oral factors related to the type of diet and their relationships to domiciliary dental care in older adults.

Generic Drug Usage in Dentistry across Japan: Analysis Using a Japanese National Database.

The aim of this study was to identify regional disparities in generic drug usage and to examine related factors. The database used for the analysis was the 2018 national health insurance claims data published on the Japanese Ministry of Health, Labour, and Welfare. The drugs that were targeted were a combination of brand-name and generic tetracycline ointments for periodontal treatment and lidocaine injection solution used for dental anesthesia. The usage of generic drugs was calculated and compared by prefecture based on the number of health insurance claims. The comparison of related factors was conducted using data from other national statistical survey. The results showed that the mean generic drug usage of tetracycline for periodontal treatment in all prefectures was 71.2 ± 8.1%, ranging from 45.8% to 85.3%. The mean generic lidocaine used for dental anesthesia was 47.6 ± 10.0%, ranging from 30.5% to 66.2%. The rank correlation coefficient between the two was 0.359 (p < 0.05), and the tendency of using both generic drugs was low in major metropolitan areas. Generic drug usage in Japan is low; thus, in order to reduce healthcare costs, generic drugs need to be actively used in dentistry.

Pancreas divisum in pancreaticobiliary maljunction in children.

PURPOSE: Pancreaticobiliary maljunction (PBM) is defined as a congenital anomaly in which the main pancreatic and common bile ducts are joined outside the duodenal wall and forms the long common channel. Although PBM and pancreas divisum are congenital anomalies causing pancreatitides, distinct data about the incidence of pancreas divisum in pediatric PBM has not been reported to date. The present study was designed to reveal the incidence and clinical features of pancreas divisum in cases of PBM. METHODS: The configurations of pancreatic ducts of 78 pediatric cases of PBM were assessed by endoscopic retrograde cholangiopancreatography (ERCP) and/or intraoperative cholangiopancreatography. Additional cannulation of the minor papilla was performed when the entire length of the main pancreatic duct was not detected with cannulation of the major papilla alone. RESULTS: Clear pancreatography was obtained in 71 cases out of 78 cases of PBM. Abnormal fusion of the pancreatic duct was detected in 1 case (1.4%) with complete pancreas divisum. This case was asymptomatic preoperatively and for 10 years postoperatively. CONCLUSION: Pancreas divisum exists in 1.4% of PBM. Although pancreas divisum is one of the pathogenesis of pancreatitis in PBM, is rarely associated with PBM and not always causes pancreatitis.

Study of the Effects of Recognition of Stress on Symptoms and Regular Hospital Visits: An Analysis from Japanese National Statistics.

Stress is a risk factor for numerous lifestyle diseases, including dental diseases. The purpose of the present study was to investigate how sensitivity to psychological stress relates to subjective symptoms and regular hospital visits using information from the large-scale database of national statistics. Anonymized data from 10,584 respondents aged 30-79 of the Japanese 2013 Comprehensive Survey of Living Conditions were analyzed. Respondents were classified by age into a middle-aged group and an elderly group, and a contingency table analysis, rank correlation analysis, and binomial logistic regression analysis were performed. Contingency table analysis confirmed that eight items were related to the presence of a recognition of stress, including the presence of subjective symptoms (p < 0.001), the presence of regular hospital visits (p < 0.001), symptoms of periodontal disease (p < 0.001), and self-rated health (p < 0.001). Responses for symptoms and diseases requiring regular hospital visits were ranked in order for a stress group and a no stress group, and it was found that other than fatigue symptoms being ranked highly in the stress group, there were no particular differences. Logistic regression analysis results showed significant odds ratios for six items including: self-rated health (3.91, 95% CI, 3.23 to 4.73), lifestyle awareness (1.96, 95% CI, 1.68 to 2.28), and symptoms of periodontal disease (1.71, 95% CI 1.19 to 2.48). The present study showed that susceptibility to psychological stress is related to awareness of subjective symptoms and to regular hospital visits due to disease, suggesting that these have direct and indirect mutual effects.

Association between Drinking Habits and Oral Symptoms: A Cross-Sectional Study Based on Japanese National Statistical Data.

AIM: The purpose of this study was to investigate the association between drinking habits and subjective symptoms of the oral cavity based on Japanese national statistical data. METHODS: The subjects were 8,698 respondents of the Japan National Livelihood Survey 2013, in their 30s to 60s. The association between drinking habits and each survey item was investigated by contingency table analysis and binary logistic regression analysis. RESULTS: The proportion of people with drinking habits was 55.3% among men and 20.3% among women, and the proportion of men with drinking habits above the age of 40 years was high. Contingency table analysis indicated an association between drinking habits and the following items in men: subjective symptoms (p < 0.01), chewing difficulty (p < 0.05), subjective impression of health (p < 0.01), smoking habit (p < 0.01), and income level (p < 0.01). Analysis indicated an association between drinking habits and hospital visits (p < 0.01) and smoking habits (p < 0.01) in women. When comparing the response rates of symptoms and presence of disease stratified according to drinking habits, inflammatory and sensory system symptoms were common in those who had drinking habits, as were diabetes and gout. Binomial logistic regression analysis with drinking habit as the objective variable indicated statistically significant odds ratios for smoking habit (2.13; 95% CI: 1.65-2.75), difficulty in chewing (1.63; 95% CI: 1.01-2.62), and working hours (1.50; 95% CI: 1.10-2.04). This study identified a correlation between drinking habits and masticatory disorders, suggesting that the effects of drinking as a risk factor for dental diseases should be examined in greater detail in the future.

Circulating histone H3 levels are increased in septic mice in a neutrophil-dependent manner: preclinical evaluation of a novel sandwich ELISA for histone H3.

BACKGROUND: Nuclear histone proteins are released into the extracellular space during sepsis and act as major mediators of death. However, circulating histone levels have not been precisely quantified. METHODS: We developed a novel enzyme-linked immunosorbent assay (ELISA) for detection of circulating histone H3 levels and evaluated its performance. Using the ELISA, we measured plasma histone H3 levels in C57BL/6 J mice subjected to cecal ligation and puncture (CLP)-induced sepsis. RESULTS: The newly developed ELISA enabled reproducible measurement of histone H3 levels with a working range up to 250 ng/mL. Using the ELISA, we found that plasma histone H3 levels were elevated in septic mice compared with sham-operated mice (p < 0.01). The elevation of histone H3 levels was abrogated when neutrophils were depleted (p < 0.01). CONCLUSIONS: Our novel ELISA provides reproducible measurements of histone H3 levels. Circulating histone H3 levels are increased in septic mice in a neutrophil-dependent manner. Further studies are needed to evaluate the clinical utility of histone H3 levels in patients with sepsis.

Evidence for viral infection as a causative factor of human biliary atresia.

OBJECTIVES: To explore the evidence for viral infections triggering human biliary atresia (BA) by reviewing archival original articles that analyzed human samples via polymerase chain reaction (PCR) experiments, considering the recent experimental trend of extensive use of rotaviral BA animal models. METHODS: A PubMed search retrieved original articles that reported the results of PCR experiments for detecting viral DNA or RNA in patient samples as proof of past infection. Search terms included the often-debated DNA or RNA viruses and BA. Special focus was directed toward PCR analyses that targeted reovirus and rotavirus, where PCR accuracy, specimen characteristics and their interpretations were compared. RESULTS: Nineteen studies were conducted on 16 different kinds of viruses using PCR, with 5 studies on reovirus, 3 on rotavirus, 10 on cytomegalovirus, 5 on Epstein-Barr virus, 4 on parvovirus B19, and so on. Among the papers suggesting a possible viral link to only BA, there was no study on reovirus, 1 on rotavirus, 3 on cytomegalovirus, 1 on EB virus, and 1 on papillomavirus. Of the 6 PCR studies on Reoviridae, 3 on reovirus and 2 on rotavirus were evaluated rigorously for experimental accuracy, including their sensitivity. Two research groups analyzed preoperative stool samples in addition to generic hepatobiliary tissue obtained at surgery. Sample collection timing varied widely, with storage period prior to PCR experimentation not revealed in most reports on Reoviridae. CONCLUSION: Although a considerable number of PCR studies have sought to clarify a viral role in the pathogenesis of BA using human samples, the findings have been contradictory and have not succeeded in achieving an obvious differentiation between causative and accidental infection of the focused virus. Reproducible and convincing evidence for a causative Reoviridae infection has been lacking based on objective data from highly sensitive PCR experiments. Even though the possibility remains of viral disappearance at the timing of collection, to avoid further ambiguous interpretations of PCR results, rigorous and meticulous collection of large numbers of specimens at carefully planned timing, along with a strictly adjusted and finely tuned PCR system, is strongly recommended for obtaining more reliable and consistent results.

Novel management of acute or secondary biliary liver conditions using hepatically differentiated human dental pulp cells.

The current definitive treatment for acute or chronic liver condition, that is, cirrhosis, is liver transplantation from a limited number of donors, which might cause complications after donation. Hence, bone marrow stem cell transplantation has been developed, but the risk of carcinogenesis remains. We have recently developed a protocol for hepatic differentiation of CD117(+) stem cells from human exfoliated deciduous teeth (SHED). In the present study, we examine whether SHED hepatically differentiated (hd) in vitro could be used to treat acute liver injury (ALI) and secondary biliary cirrhosis. The CD117(+) cell fraction was magnetically separated from SHED and then differentiated into hepatocyte-like cells in vitro. The cells were transplanted into rats with either ALI or induced secondary biliary cirrhosis. Engraftment of human liver cells was determined immunohistochemically and by in situ hybridization. Recovery of liver function was examined by means of histochemical and serological tests. Livers of transplanted animals were strongly positive for human immunohistochemical factors, and in situ hybridization confirmed engraftment of human hepatocytes. The tests for recovery of liver function confirmed the presence of human hepatic markers in the animals' blood serum and lack of fibrosis and functional integration of transplanted human cells into livers. No evidence of malignancy was found. We show that in vitro hdSHED engraft morphologically and functionally into the livers of rats having acute injury or secondary biliary cirrhosis. SHED are readily accessible adult stem cells, capable of proliferating in large numbers before differentiating in vitro. This makes SHED an appropriate and safe stem cell source for regenerative medicine.

Reevaluation of trkA expression as a biological marker of neuroblastoma by high-sensitivity expression analysis--a study of 106 primary neuroblastomas treated in a single institute.

BACKGROUND/PURPOSE: It has previously been shown that neuroblastomas with favorable prognosis often express a high level of nerve growth factor receptor trkA. We performed an expression analysis of trkA in 106 NB samples based on the quantitative real-time polymerase chain reaction (PCR) and reevaluated the prognostic power of trkA. MATERIALS AND METHODS: A total of 106 primary tumors from NB patients treated from 1988 to 2009 were analyzed. MYCN was amplified in 13 cases. TaqMan probe method was used for quantitative PCR. Primers and probes were designed to detect trkA I and trkA II, but not the oncogenic splice variant trkA III. RESULTS: Expression analysis by real-time PCR revealed a wide range of expression levels of trkA within neuroblastoma tissues. Extremely low levels of trkA that were undetectable by semiquantitative PCR were able to be quantified by this method. trkA was predominantly expressed in tumors with favorable outcome. Further analysis of trkA expression was performed in a cohort excluding mass-screened neuroblastomas. Strikingly, multivariate analysis containing age, MYCN status, and trkA expression identified trkA as the only variable that independently predicts the prognosis of the 44 patients who presented clinically. CONCLUSION: High-resolution expression analysis targeting trkA and trkA II may add more statistical power on trkA as a biological marker.

Kaposiform hemangioendothelioma of the choledochus.

Kaposiform hemangioendothelioma (KHE) is a rare, locally aggressive vascular neoplasm that mainly occurs during childhood. Although KHE may involve various organs, involvement of the choledochus has not been reported. We report a case of KHE in a 5-month-old male infant. The patient was admitted with icterus and acholic stool. Contrast computed tomography revealed a vascular tumor in the hepatic portal region causing biliary obstruction. Excision of the extrahepatic duct and hepatoportoenterostomy were performed successfully, and he has been well during 3 years of postoperative follow-up.