Loss of Autophagy Disrupts Stemness of Ameloblast-Lineage Cells in Aging.

Autophagy is one of the intracellular degradation pathways and maintains cellular homeostasis, regulating the stress response, cell proliferation, and signal transduction. To elucidate the role of autophagy in the maintenance of dental epithelial stem cells and the subsequent enamel formation, we analyzed autophagy-deficient mice in epithelial cells (Atg7f/f;KRT14-Cre mice), focusing on the influence of aging and stress environments. We also performed in vitro cell and organ culture experiments with an autophagy inhibitor. In young Atg7f/f;KRT14-Cre mice, morphological change was not obvious in maxillary incisors, except for the remarkable cell death in the stratum intermedium of the transitional stage. However, under stress conditions of hyperglycemia, the incisor color changed to white in diabetes Atg7f/f;KRT14-Cre mice. Regarding dental epithelial stem cells, the shape of the apical bud region of the incisor became irregular with age, and odontoma was formed in aged Atg7f/f;KRT14-Cre mice. In addition, the shape of apical bud culture cells of Atg7f/f;KRT14-Cre mice became irregular and enlarged atypically, with epigenetic changes during culture, suggesting that autophagy deficiency may induce tumorigenesis in dental epithelial cells. The epigenetic change and upregulation of p21 expression were induced by autophagy inhibition in vivo and in vitro. These findings suggest that autophagy is important for the regulation of stem cell maintenance, proliferation, and differentiation of ameloblast-lineage cells, and an autophagy disorder may induce tumorigenesis in odontogenic epithelial cells.

Sample collection from asteroid (162173) Ryugu by Hayabusa2: Implications for surface evolution.

The near-Earth asteroid (162173) Ryugu is thought to be a primitive carbonaceous object that contains hydrated minerals and organic molecules. We report sample collection from Ryugu's surface by the Hayabusa2 spacecraft on 21 February 2019. Touchdown images and global observations of surface colors are used to investigate the stratigraphy of the surface around the sample location and across Ryugu. Latitudinal color variations suggest the reddening of exposed surface material by solar heating and/or space weathering. Immediately after touchdown, Hayabusa2's thrusters disturbed dark, fine grains that originate from the redder materials. The stratigraphic relationship between identified craters and the redder material indicates that surface reddening occurred over a short period of time. We suggest that Ryugu previously experienced an orbital excursion near the Sun.

Simple modification to improve reliability of copro-DNA examinations for diagnosing Echinococcus multilocularis infections in red foxes.

Epidemiological studies of Echinococcus multilocularis infections in definitive hosts require a reliable and economic diagnostic method. In this study, the current copro-DNA examination technique was modified by increasing the faecal amounts tested and adding a step to neutralize the faeces before DNA extraction. Reliability of the modified method was evaluated using rectal faecal samples from red foxes and comparing them with intestinal worms detected using the sedimentation and counting technique (SCT) following necropsy. The modified copro-DNA examination method demonstrated 93.9% sensitivity (138/147) on the SCT. Its detectability increased depending on the worm burden, and the sensitivity was 100% in cases harbouring over 1000 worms. From 111 SCT-negative cases, six (5.4%) were copro-DNA-positive, and all were confirmed as E. multilocularis via sequencing analysis. Five of the remaining 105 SCT-negative cases (4.8%) retained polymerase chain reaction (PCR) inhibitors in the extracted solution, suggesting that approximately 5% of the red fox faeces retained these inhibitors after treatment with the present copro-DNA extraction method. Although further evaluation is needed for faeces deposited in the wild, the present copro-DNA examination technique will help monitor the E. multilocularis prevalence in definitive hosts. When used for detailed evaluations of endemicity (e.g. changes in infection pressure or spread in non-endemic areas), the absence of PCR inhibitors should be confirmed, and multiple trials on faecal subsamples are recommended.

Single-center Experience of Living-donor Liver Transplantation in Patients Receiving Maintenance Renal Replacement Therapy.

BACKGROUND: For most patients with liver failure receiving maintenance renal replacement therapy (RRT), treatment with living-donor liver transplantation (LDLT) alone is indicated in Japan. MATERIAL AND METHODS: We retrospectively reviewed patients who underwent LDLT while receiving RRT in our hospital. RESULTS: Three of the 5 patients who underwent LDLT while on RRT died during the first year after transplantation. CONCLUSIONS: The indications for liver transplantation in patients on RRT require careful examination.

Portal Vein Stent Placement in Living-donor Liver Transplantation: A Single-center Experience.

INTRODUCTION: Portal vein thrombosis (PVT) and portal vein stenosis (PVS) are rare complications after liver transplantation that can lead to graft failure and patient death. MATERIAL AND METHODS: The aim of this study was to evaluate the effect of interventional treatment for PVT and PVS occlusion after liver transplantation. Follow-up data of 7 patients who underwent stent replacement for PVT and/or PVS were analyzed. The clinical success, complications, and portal vein patency were analyzed. RESULTS: Clinical success was obtained in 6 of the 7 patients. No portal hypertension-related symptoms reoccurred in the 6 patients during the follow-up. CONCLUSIONS: Interventional radiologic treatment produced a high success rate and a favorable long-term outcome.

The epithelial zinc transporter ZIP10 epigenetically regulates human epidermal homeostasis by modulating histone acetyltransferase activity.

BACKGROUND: The skin is the first organ that manifests changes in response to zinc deficiency. However, the molecular mechanism underlying how zinc is involved in skin homeostasis, especially its epigenetic regulation, is largely unknown. OBJECTIVES: In this study we demonstrate the importance of zinc levels and the zinc transporter ZIP10 in the epigenetic maintenance of human epidermal homeostasis. METHODS: Adult human skin, including skin appendages, were stained with anti-ZIP10 antibody. Histone acetyltransferase (HAT) activity was assessed after treating human keratinocytes with ZIP10 small interfering (si)RNAs or the zinc chelator TPEN. ZIP10- or HAT-regulated genes were analysed based on limma bioinformatics analysis for keratinocytes treated with ZIP10 siRNAs or a HAT inhibitor, or using a public database for transcription factors. A reconstituted human skin model was used to validate the role of ZIP10 in epidermal differentiation and the functional association between ZIP10 and HAT. RESULTS: ZIP10 is predominantly expressed in the interfollicular epidermis, epidermal appendages and hair follicles. ZIP10 depletion resulted in epidermal malformations in a reconstituted human skin model via downregulation of the activity of the epigenetic enzyme HAT. This decreased HAT activity, resulting from either ZIP10 depletion or treatment with the zinc chelator TPEN, was readily restored by zinc supplementation. Through bioinformatics analysis for gene sets regulated by knockdown of SLC39A10 (encoding ZIP10) and HAT inhibition, we demonstrated that ZIP10 and HATs were closely linked with the regulation of genes related to epidermal homeostasis, particularly filaggrin and metallothionein. CONCLUSIONS: Our study suggests that ZIP10-mediated zinc distribution is crucial for epidermal homeostasis via HATs. Therefore, zinc-dependent epigenetic regulation could provide alternatives to maintaining healthy skin or alleviating disorders with skin barrier defects.

Regression of gastric de novo diffuse large B-cell lymphoma following Helicobacter pylori eradication: a case report.

We report a case of primary gastric diffuse large B-cell lymphoma (DLBCL), de novo DLBCL without the features of mucosa-associated lymphoid tissue (MALT) lymphoma, which regressed after Helicobacter pylori (HP) eradication. A 27-year-old Japanese female with epigastralgia was revealed to have ulcerated lesions in the angle and antral regions on gastroscopy. Biopsy specimen was consistent with a diagnosis of DLBCL without MALT lymphoma component, indicating de novo development. Her clinical staging on the Lugano system was Stage I. HP was positive on a rapid urease test, and she received HP eradication therapy twice, because the first therapy was not successful. On gastroscopy performed 1 month after the second HP eradication therapy, no ulcerated lesion was noted, and the lymphoma cells had regressed histopathologically. (Acta gastro-enterol. belg., 2016, 79, 367-369A).

The timing of worm exclusion in dogs repeatedly infected with the cestode Echinococcus multilocularis.

Experimental Echinococcus multilocularis infection and deworming was repeated three or five times in nine dogs at various re-infection schedules. The mean number of worms decreased more than 91% in dogs with repeated infection, compared to first infection controls (n= 6). The copro-antigen assay and the egg count in the faeces suggested that the worm burden gradually decreased each time the dogs were re-infected. To examine whether such worm exclusion was a non-specific response, five dogs were sequentially infected with the parasite four times and subsequently fed freely for 6 months. Even after the 6-month interval, the five dogs that were infected five times with the parasite were still able largely to exclude the adult worms. The results suggested that the ability of worm exclusion in dogs that developed a resistance did not become rapidly extinct. Observation of the condition of faeces and the excretion of hooks in the faeces of repeatedly infected dogs revealed that the exclusion of worms started at the first week after the re-infection, and it continued during the patent period. Serum antibodies specific to the parasite antigen increased gradually until the third infection and significantly decreased during the 6-month interval. There was little enhancement of serum antibodies after the fifth infection in most dogs, although no clear correlation was observed between the antibody response and the worm burden. These findings suggested the possibility of developing a vaccine.

Dementia with Lewy bodies can be well-differentiated from Alzheimer's disease by measurement of brain acetylcholinesterase activity-a [11C]MP4A PET study.

OBJECTIVE: To investigate the diagnostic performance of brain acetylcholinesterase (AChE) activity measurement using N-[(11) C]-methyl-4-piperidyl acetate (MP4A) and PET in patients with dementia with Lewy bodies (DLB) and Alzheimer's disease (AD). METHODS: Participants were 14 DLB patients, 25 AD patients and 18 age-matched healthy controls (HC). All subjects underwent PET scans and MP4A to measure regional brain AChE activity. We performed anatomical standardization of each brain image, and k3 values, an index of AChE activity, in each voxel were estimated by nonlinear least squares analysis. Volumes of interest (VOIs) were identified on parametric k3 images in frontal, temporal, parietal and occipital cortices, and in anterior and posterior cingulate gyri (ACG and PCG). In each VOI, the differential diagnostic performance between AD and DLB of k3 values was assessed by area under the curve (AUC) of the receiver-operating characteristic. Voxel-based statistical analyses were also performed. RESULTS: Mean cortical AChE activities in AD patients (-8.2% compared with normal mean) and DLB patients (-27.8%) were lower than HCs (p < 0.05, p < 0.001, respectively). There was a significant difference in mean cortical AChE activities between AD and DLB patients (p < 0.001). All regional brain AChE activities of defined VOIs except ACG were able to well discriminate DLB from AD, and notably performance was the most significant in PCG (AUC = 0.989, 95% CI: 0.965-1.000). CONCLUSIONS: Brain cholinergic deficit is consistently prominent in DLB compared with AD. PET measurement of brain AChE activity may be useful for the differential diagnosis between DLB and AD.

Increase of Clostridium perfringens in association with Eimeria in haemorrhagic enteritis in Japanese beef cattle.

A coprological survey with detailed clinical observation of naturally occurring haemorrhagic enteritis (HE) cases was conducted to understand the pathophysiology of HE by clarifying the infection status of Eimeria and enteropathogenic bacteria in cattle. Faecal samples from 55 cases of HE and 26 clinically normal animals were collected, and a quantitative examination of Eimeria and potential enteropathogenic bacteria was performed. The number of Eimeria species oocysts per gram of faeces (OPG) exceeded 10,000 in 69.1 per cent of HE cases with a maximum of 1,452,500 OPG and Eimeria zuernii was found to be overwhelmingly dominant. A significant increase in faecal coliform count was observed in HE cases compared with clinically normal animals. Among the animals shedding >10,000 OPG, 42.9 per cent showed a remarkable increase in Clostridium perfringens abundance (>104 CFU/g) in the faeces. In the cases with C. perfringens detected, its abundance was positively correlated with Eimeria OPG and high C. perfringens abundance was always accompanied by high Eimeria OPG. E. zuernii is likely to play a crucial role in massive multiplication of C. perfringens in HE in cattle.

Differences in the developmental origins of the periosteum may influence bone healing.

BACKGROUND AND OBJECTIVE: The jaw bone, unlike most other bones, is derived from neural crest stem cells, so we hypothesized that it may have different characteristics to bones from other parts of the body, especially in the nature of its periosteum. The periosteum exhibits osteogenic potential and has received considerable attention as a grafting material for the repair of bone and joint defects. MATERIAL AND METHODS: Gene expression profiles of jaw bone and periosteum were evaluated by DNA microarray and real-time polymerase chain reaction. Furthermore, we perforated an area 2 mm in diameter on mouse frontal and parietal bones. Bone regeneration of these calvarial defects was evaluated using microcomputed tomography and histological analysis. RESULTS: The DNA microarray data revealed close homology between the gene expression profiles within the ilium and femur. The gene expression of Wnt-1, SOX10, nestin, and musashi-1 were significantly higher in the jaw bone than in other locations. Microcomputed tomography and histological analysis revealed that the jaw bone had superior bone regenerative abilities than other bones. CONCLUSION: Jaw bone periosteum exhibits a unique gene expression profile that is associated with neural crest cells and has a positive influence on bone regeneration when used as a graft material to repair bone defects. A full investigation of the biological and mechanical properties of jaw bone as an alternative graft material for jaw reconstructive surgery is recommended.

Lin28a is a putative factor in regulating cancer stem cell-like properties in side population cells of oral squamous cell carcinoma.

Cancer stem cells (CSCs) are among the target cells of cancer therapy because they are uniquely involved in both cancer progression and sensitivity to chemotherapeutic agents. We identified side population (SP) cells, which are known to be an enriched population of CSC, in five oral squamous cell carcinoma (OSCC) cells (SCC9, SCC25, TOSCC7, TOSCC17, and TOSCC23). The percentages of SP cells ranged from 0% to 3.3%, with TOSCC23 cells showing the highest percentages of SP cells (3.3% of the total cell population). The SP cells isolated from TOSCC23 cells also showed greater cell proliferation and invasion compared to non-SP (MP) cells. Therefore, our initial findings suggested that SP cells were enriched for CSC-like cells. Furthermore, DNA microarray analysis revealed that the expression of cell proliferation-related and anti-apoptotic genes was greater in SP cells compared to MP cells. We focused on Lin28a, which showed the highest expression (approximately 22-fold) among the upregulated genes. The overexpression of Lin28a in TOSCC23 cells increased their proliferation, colony formation, and invasion. These findings suggest that Lin28a is an appropriate CSC target molecule for OSCC treatment.

PET imaging of MRP1 function in the living brain: method development and future perspectives.

Multidrug resistance-associated protein 1 (MRP1) functions as a primary active transporter utilizing energy from ATP hydrolysis. In the central nervous system (CNS), MRP1 plays an important role in limiting the permeation of xenobiotic and endogenous substrates across the blood-brain and blood-cerebrospinal fluid barriers, and across brain parenchymal cells. While MRP1 contributes to minimizing the neurotoxic effects of drugs, it may also restrict the distribution of drugs for the treatment of CNS diseases. Moreover, neurodegenerative disease may be associated with abnormal expression of efflux transporters in the brain. Noninvasive measurement of MRP1 function will therefore be useful for directly evaluating the effect of modulators on enhancing the penetration of drugs into the brain and for examining the pathophysiological role of MRP1 in the brain. Positron emission tomography (PET) is a powerful molecular imaging technique. While several PET probes have been proposed for imaging function of the efflux transporter P-glycoprotein, few reports discuss the probes for imaging MRP1 function in the brain. Ideally, brain radioactivity should consist of a single radioactive compound that is selectively transported by the efflux transporter of interest, without other efflux routes. However, most PET probes for MRP1 or P-glycoprotein are eliminated by both a transporter and simple diffusion, resulting in inaccurate measurement of pump function. This review addresses a new strategy to avoid this problem, and suggests the design of a PET probe based on this strategy, particularly for MRP1 imaging. Several published reports on imaging MRP1 function with PET are also discussed.

Surgical strategies for hepatocellular carcinoma with special reference to anatomical hepatic resection and intraoperative contrast-enhanced ultrasonography.

Here we described our strategies to attain a better prognosis for hepatocellular carcinoma (HCC) patients by surgery. Among a variety of attempts conducted to date, we focused on anatomical resection and intraoperative contrast-enhanced ultrasonography. There are still controversies with respect to the significance of anatomical resection. We analyzed the significance of this surgical procedure in 207 patients without macrovascular invasion. These patients underwent either anatomical resection or non-anatomical resection. We found that the patients with anatomical resection had higher recurrence-free survival rate than those with non-anatomical resection. Univariable analysis showed that liver damage, the serum level of alpha-fetoprotein, tumor number, surgical margin, and type of surgery (anatomical or non-anatomical resection) were significant predictive factors for intrahepatic recurrence. Multivariable analysis revealed that multiple tumors, alpha-fetoprotein, and non-anatomical resection were independent risk factors for recurrence. We conclude that anatomical resection is a recommendable surgical procedure in patients without macrovascular invasion. A recent innovation is the development of contrast-enhanced ultrasonography. Then we have applied this to liver surgery intraoperatively. We confirm that vascular images contribute to a precise diagnosis and the detection of small portal tumor thrombi, and that Kupffer images are useful to discover the minute tumors. In addition, by clarifying the relationship between tumors and the vascular architecture, real-time 3-dimensional images using Kupffer imaging are a promising guide during the surgical procedures, although further development is needed.

Mapping of brain acetylcholinesterase alterations in Lewy body disease by PET.

OBJECTIVE: To characterize brain cholinergic deficits in Parkinson disease (PD), PD with dementia (PDD), and dementia with Lewy bodies (DLB). METHODS: Participants included 18 patients with PD, 21 patients with PDD/DLB, and 26 healthy controls. The PD group consisted of nine patients with early PD, each with a disease duration of less than 3 years, five of whom were de novo PD patients, and nine patients with advanced PD, each with a disease duration greater than or equal to 3 years. The PDD/DLB group consisted of 10 patients with PDD and 11 patients with DLB. All subjects underwent PET scans with N-[11C]-methyl-4-piperidyl acetate to measure brain acetylcholinesterase (AChE) activity. Brain AChE activity levels were estimated voxel-by-voxel in a three-compartment analysis using the arterial input function, and compared among our subject groups through both voxel-based analysis using the statistical parametric mapping software SPM5 and volume-of-interest analysis. RESULTS: Among patients with PD, AChE activity was significantly decreased in the cerebral cortex and especially in the medial occipital cortex (% reduction compared with the normal mean = -12%) (false discovery rate-corrected p value <0.01). Patients with PDD/DLB, however, had even lower AChE activity in the cerebral cortex (% reduction = -27%) (p < 0.01). There was no significant difference between early PD and advanced PD groups or between DLB and PDD groups in the amount by which regional AChE activity in the brain was reduced. CONCLUSIONS: Brain cholinergic dysfunction occurs in the cerebral cortex, especially in the medial occipital cortex. It begins in early Parkinson disease, and is more widespread and profound in both Parkinson disease with dementia and dementia with Lewy bodies.

Cost benefit and clinical efficacy of low-dose granulocyte colony-stimulating factor after standard chemotherapy in patients with non-Hodgkin's lymphoma.

High costs of molecule-targeted drugs, such as rituximab, ibritumomab, and tositumomab have given rise to an economical issue for treating patients with non-Hodgkin's lymphoma (NHL). Granulocyte colony-stimulating factors (G-CSFs), which are also expensive, are widely used for treating neutropenia after chemotherapy. In Japan, lenograstim at 2 microg/kg (about 100 microg/body) or filgrastim at 50 microg/m(2) (about 75 microg/body) is commonly administered for patients with NHL after chemotherapy. Therefore, cost-effectiveness is an important issue in treatment for NHL. Patients with advanced-stage NHL who needed chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) or a CHOP-like regimen with or without rituximab were enrolled in this randomized cross-over trial to investigate the efficacy and safety of low-dose G-CSF. Half of the patients were administered 75 microg filgrastim in the first course after neutropenia and 50 microg lenograstim in the second course, and the other half were crossed over. Forty-seven patients were enrolled in this cross-over trial, and 24 patients completed the trial. Frequencies and durations of grade 4 leukocytopenia and neutropenia were similar in the two groups. Severe infection was rare and was observed at similar frequency. Frequencies of antibiotics use were also similar. The total cost of G-CSF (cost/drug x duration of administration) was significantly lower in patients who received 50 microg lenograstim. Hence, a low dose of lenograstim might be safe, effective and pharmaco-economically beneficial in patients with advanced-stage NHL.

[Lung abscess which ruptured during the medical treatment of lung abscess; report of a case].

63-year-old man was admitted to our hospital with fever and cough for about 2 months. Laboratory data showed marked inflammatory changes, and chest computed tomography (CT) scans revealed right-sided hydrothorax, atelectasis of the right middle lobe, and a cystic mass in the right middle lobe. We diagnosed the patients as having lung abscess and empyema. Following the intravenous antibiotic chemotherapy, symptoms and laboratory data showed the improvement, however, on the 11th hospital day, he developed high fever again. A chest CT showed pneumopyothorax suggesting the rupture of lung abscess. Since the chest tube drainage was ineffective, open chest surgery was performed. Curettage of both thoracic and abscess cavity with closure of air leakage successfully cured the pyothorax.

Chitosan-based hyaluronan hybrid polymer fibre scaffold for ligament and tendon tissue engineering.

To establish medical use of tissue engineering technology for ligament and tendon injuries, a scaffold was developed which has sufficient ability for cell growth, cell differentiation, and mechanical properties. The scaffold made from chitosan and 0.1 per cent hyaluronic acid has adequate biodegradability and biocompatibility. An animal experiment showed that the scaffold has less toxicity and less inflammation induction. Furthermore, in-vivo animal experiments showed that the mechanical properties of the engineered ligament or tendon had the possibility to stabilize the joint. It was shown that newly developed hybrid-polymer fibre scaffold has feasibility for joint tissue engineering.