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In CBA/CaJ mice, confocal analysis has shown that acoustic overexposure can immediately destroy synapses between auditory-nerve fibers (ANFs) and their peripheral targets, the inner hair cells (IHCs), and that years later, a corresponding number of ANF cell bodies degenerate. In guinea pig, post-exposure disappearance of pre-synaptic ribbons can be equally dramatic, however, post-exposure recovery to near-baseline counts has been reported. Since confocal counts are confounded by thresholding issues, the fall and rise of synaptic ribbon counts could represent “regeneration,” i.e., terminal retraction, re-extension and synaptogenesis, or “recovery,” i.e., down- and subsequent up-regulation of synaptic markers. To clarify, we counted pre-synaptic ribbons, assessed their juxtaposition with post-synaptic receptors, measured the extension of ANF terminals, and quantified the spatial organization and size gradients of these synaptic elements around the hair cell. Present results in guinea pigs exposed as adults (14 months), along with prior results in juveniles (1 month), suggest there is post-exposure neural regeneration in the guinea pig, but not the CBA/CaJ mouse, and that this regenerative capacity extends into adulthood. The results also show, for the first time, that the acute synaptic loss is concentrated on the modiolar side of IHCs, consistent with a selective loss of the high-threshold ANFs with low spontaneous rates. The morphological similarities between the post-exposure neurite extension and synaptogenesis, seen spontaneously in the guinea pig, and in CBA/CaJ only with forced overexpression of neurotrophins, suggest that the key difference may be in the degree of sustained or injury-induced expression of these signaling molecules in the cochlea.

Background Pneumonia is a common cause of illness and death of the elderly in Japan. Its prevalence is escalating globally with the aging of population. To describe the latest trends in pneumonia hospitalizations, especially aspiration pneumonia (AP) cases, we assessed the clinical records of pneumonia patients admitted to core acute care hospitals in Miyagi prefecture, Japan. Methods A retrospective multi-institutional joint research was conducted for hospitalized pneumonia patients aged ≥20 years from January 2019 to December 2019. Clinical data of patients were collected from the medical records of eight acute care hospitals. Results Out of the 1,800 patients included in this study, 79% of the hospitalized pneumonia patients were aged above 70 years. The most common age group was in the 80s. The ratio of AP to total pneumonia cases increased with age, and 692 out of 1,800 patients had AP. In univariate analysis, these patients had significantly older ages, lower body mass index (BMI), a lower ratio of normal diet intake and homestay before hospitalization, along with more AP recurrences and comorbidities. During hospitalization, AP patients had extended fasting periods, more swallowing assessments and interventions, longer hospitalization, and higher in-hospital mortality rate than non-AP patients. A total of 7% and 2% AP patients underwent video endoscopy and video fluorography respectively. In multivariate analysis, lower BMI, lower C-reactive protein, a lower ratio of homestay before hospitalization, a higher complication rate of cerebrovascular disease, dementia, and neuromuscular disease were noted as a characteristic of AP patients. Swallowing interventions were performed for 51% of the AP patients who had been hospitalized for more than two weeks. In univariate analysis, swallowing intervention improved in-hospital mortality. Lower AP recurrence before hospitalization and a lower ratio of homestay before hospitalization were indicated as characteristics of AP patients of the swallowing intervention group from multivariate analysis. Change in dietary pattern from normal to modified diet was observed more frequently in the swallowing intervention group. Conclusion AP accounts for 38.4% of all pneumonia cases in acute care hospitals in Northern Japan. The use of swallowing evaluations and interventions, which may reduce the risk of dysphagia and may associate with lowering mortality in AP patients, is still not widespread.

Overexposure to intense noise can destroy the synapses between auditory nerve fibers and their hair cell targets without destroying the hair cells themselves. In adult mice, this synaptopathy is immediate and largely irreversible, whereas, in guinea pigs, counts of immunostained synaptic puncta can recover with increasing post-exposure survival. Here, we asked whether this recovery simply reflects changes in synaptic immunostaining, or whether there is actual retraction and extension of neurites and/or synaptogenesis. Analysis of the numbers, sizes and spatial distribution of pre- and post-synaptic markers on cochlear inner hair cells, in guinea pigs surviving from 1 day to 6 months after a synaptopathic exposure, shows dramatic synaptic re-organization during the recovery period in which synapse counts recover from 16 to 91% of normal in the most affected regions. Synaptic puncta move all over the hair cell membrane during recovery, translocating far from their normal positions at the basolateral pole, and auditory-nerve terminals extend towards the hair cell’s apical end to re-establish contact with them. These observations provide stronger evidence for spontaneous neural regeneration in a mature mammalian cochlea than can be inferred from synaptic counts alone.

Objectives. This study aimed to evaluate the effect of Royal Jelly (RJ) at a dose of 800 mg/day on menopausal symptoms in healthy Japanese postmenopausal women with placebo-controlled design. Material and Methods. A total of 42 healthy Japanese postmenopausal women have been recruited for this study. The subjects were randomized to oral treatment with either 800 mg of protease-digested lyophilized powder of RJ (enzyme-treated RJ) or placebo (800 mg of dextrin) daily for 12 weeks. The level of menopausal symptoms has been evaluated every 4 weeks, using menopausal symptoms questionnaire of Japanese women. Independent t-test was used to evaluate statistical significance of the treatment effects between the two groups. Results and Conclusion. All of the 42 women have completed the trial. There were significant differences related to the anxiety score (P=0.046) and backache and low back pain score (P=0.040) between 800 mg/day enzyme-treated RJ and placebo-treated groups after 12 weeks of administration, and no significant differences were found between the two groups in 4 weeks after intervention. No side effects were observed in either group. This study demonstrates that enzyme-treated RJ supplementation with doses of 800 mg/day is effective in relieving menopausal symptoms such as anxiety, backache, and low back pain in Japanese postmenopausal women.

Influenza is one of the important respiratory tract infections that require special attention for maintaining health and hygiene. The removal of influenza virus (IFV) by secretory IgA produced by the respiratory epithelium has been reported to be a critical host defense mechanism. Therefore, we isolated Lactobacillus kunkeei YB38 (YB38), the promoter of the salivary IgA secretion in humans, from honeybee pollen and studied the effect of heat-killed YB38 treatment for preventing IFV infection in a mouse model. Female BALB/c mice received YB38 orally for 21 consecutive days and were then inoculated nasally with IFV. The YB38-treated group with a daily dose of 100 mg/kg showed an increased survival rate after IFV infection relative to the control. IgA secretion in the respiratory epithelium in the YB38-treated group (100 mg/kg) was significantly increased after 6 days of infection, while IL-6 production in the same respiratory site and the number of cells infiltrating into alveoli were significantly decreased. Moreover, lung tissue damage that appeared after IFV infection was reduced. These results suggested that the YB38 dose induced early and local IgA secretion at the infection site, inhibited persistent IFV infection, and prevented the infiltration of inflammatory immune cells or production of excessive IL-6, resulting in less damage to lung tissues.

Connectin, also called titin, is the largest protein with a critical function as a molecular spring during contraction and relaxation of striated muscle; its mutation leads to severe myopathy and cardiomyopathy. To uncover the cause of this pathogenesis, zebrafish have recently been used as disease models because they are easier to genetically modify than mice. Although the gene structures and putative primary structures of zebrafish connectin have been determined, the actual primary structures of zebrafish connectin in heart and skeletal muscles remain unclear because of its large size and the PCR amplification-associated difficulties. In this research, using RT-PCR amplification from zebrafish heart and skeletal muscles, we determined the complete primary structures of zebrafish connectin in the I-band region at which mechanical property is modulated by alternative splicing. Our results showed that the domain structures of zebrafish connectins were largely similar to those of human connectins; however, the splicing pathways in the middle-Ig segment and the PEVK segment were highly diverse in every isoform. We also found that a set of 10 Ig domains in the middle-Ig segment of zebrafish connectin had been triplicated in human connectin. Because these triplicate regions are expressed in human leg and diaphragm, our findings may provide insight into the establishment of walking with limbs and lung respiration during tetrapod evolution.

Noise exposures causing only transient threshold shifts can destroy auditory-nerve synapses without damaging hair cells. Here, we asked whether virally mediated neurotrophin3 (NT3) overexpression can repair this damage. CBA/CaJ mice at 6 wks were injected unilaterally with adeno-associated virus (AAV) containing either NT3 or GFP genes, via the posterior semicircular canal, 3 wks prior to, or 5 hrs after, noise exposure. Controls included exposed animals receiving vehicle only, and unexposed animals receiving virus. Thresholds were measured 2 wks post-exposure, just before cochleas were harvested for histological analysis. In separate virus-injected animals, unexposed cochleas were extracted for qRT-PCR. The GFP reporter showed that inner hair cells (IHCs) were transfected throughout the cochlea, and outer hair cells mainly in the apex. qRT-PCR showed 4- to 10-fold overexpression of NT3 from 1–21 days post-injection, and 1.7-fold overexpression at 40 days. AAV-NT3 delivered prior to noise exposure produced a dose-dependent reduction of synaptopathy, with nearly complete rescue at some cochlear locations. In unexposed ears, NT3 overexpression did not affect thresholds, however GFP overexpression caused IHC loss. In exposed ears, NT3 overexpression increased permanent threshold shifts. Thus, although NT3 overexpression can minimize noise-induced synaptic damage, the forced overexpression may be harmful to hair cells themselves during cochlear overstimulation.

The origin of worm-like bodies and their relationship with Birbeck granules is poorly understood. To clarify the origin of worm-like bodies and their relationship with Birbeck granules. Over 800 electron micrographs of histiocytic disorders and several appendage tumours were reviewed in order to check for worm-like bodies and Birbeck granules. Worm-like bodies were most often encountered in mildly- to moderately-proliferative histiocytic tumours. Birbeck granules were observed in more malignant conditions. Narrow endoplasmic reticulum (Nrer), which resembles worm-like bodies, was abundant in worm-like body-rich cells and coexisted with Birbeck rods. Nrer is thought to be one of the candidates that gives rise to worm-like bodies. An ultrastructural similarity exists between worm-like bodies (and octopus body formation) and the so-called "lupus erythematosus virus". The presence of Nrer is often concomitant with other organelle markers, and could be a candidate for the origin of worm-like bodies.

Aging is associated with motor disorders that decrease the quality of life (QOL). Royal jelly (RJ), used as a dietary supplement, has shown various health benefits and, therefore, it has the potential to improve the QOL during aging. We have previously developed protease enzyme-treated RJ to avoid the anaphylactic response induced by RJ supplementation. However, the effects of a lifelong treatment with RJ on normal aging have not been fully clarified. In this study, we investigated the effects of enzyme-untreated RJ (NRJ) and enzyme-treated RJ (ERJ) on the aging process focusing on motor functions, by using a genetically heterogeneous (HET) mouse model experimentally endowed with genetic diversity. We performed four different physical performance tests (grip strength, wire hang, horizontal bar, and rotarod). We showed that the age-related impairment of the motor functions was significantly delayed in RJ-treated mice. Both NRJ and ERJ were similarly effective against these types of aging-associated declines. Histological analyses revealed that the RJ treatment affected the muscle fiber size at an advanced age. We also demonstrated that age-related changes in muscle satellite cell markers and catabolic genes were affected in RJ-treated mice. These results suggest that non-protein components of RJ improved the motor function in aging mice. These findings indicate that RJ has the potential to change the QOL during aging by regulating the motor function.

Loss of cardiomyocyte proliferative capacity after birth is a major obstacle for therapeutic heart regeneration in adult mammals. We and others have recently shown the importance of hypoxic in utero environments for active foetal cardiomyocyte proliferation. Here, we report the unexpected expression of novex-3, the short splice variant of the giant sarcomeric protein connectin (titin), in the cardiomyocyte nucleus specifically during the hypoxic foetal stage in mice. This nuclear localisation appeared to be regulated by the N-terminal region of novex-3, which contains the nuclear localisation signal. Importantly, the nuclear expression of novex-3 in hypoxic foetal cardiomyocytes was repressed at the postnatal stage following the onset of breathing and the resulting elevation of oxygen tension, whereas the sarcomeric expression remained unchanged. Novex-3 knockdown in foetal cardiomyocytes repressed cell cycle-promoting genes and proliferation, whereas novex-3 overexpression enhanced proliferation. Mechanical analysis by atomic force microscopy and microneedle-based tensile tests demonstrated that novex-3 expression in hypoxic foetal cardiomyocytes contributes to the elasticity/compliance of the nucleus at interphase and facilitates proliferation, by promoting phosphorylation-induced disassembly of multimer structures of nuclear lamins. We propose that novex-3 has a previously unrecognised role in promoting cardiomyocyte proliferation specifically at the hypoxic foetal stage.

Aims To identify lactic acid bacterial isolates, which promote immunoglobulin A (IgA) production in honeybee products and honeybees (Apis mellifera). Methods and Results Pyrosequencing analysis of the microbiota of honeybee products and honeybees revealed the predominance of Lactobacillus kunkeei in honey, bee pollen, bee bread and royal jelly. Lactobacillus kunkeei was isolated from bee pollen, bee bread and honey stomach, and its effect on IgA production was evaluated in vitro. Heat-killed YB38 and YB83 isolates from bee pollen promoted IgA production in mouse Peyer's Patch cells and had little mitogenic activity or effect on IL-2 production in mouse spleen cells in comparison with Listeria monocytogenes, which does exhibit mitogen activity. A pilot study in 11 healthy adults showed that 4-week intake of 1000 mg day−1 heat-killed YB38 increased secretory IgA (SIgA) concentrations and secretion in saliva with no adverse effects. Conclusion Heat-killed Lact. kunkeei YB38 from bee pollen increases IgA production and may safely improve immune responsiveness. Significance and Impact of the Study This is the first report of microbiota analysis of royal jelly and the immune efficacy of Lact. kunkeei from honeybee products in humans.

Fetal cardiomyocytes actively proliferate to form the primitive heart in utero in mammals, but they stop dividing shortly after birth. The identification of essential molecules maintaining this active cardiomyocyte proliferation is indispensable for potential adult heart regeneration. A recent study has shown that this proliferation depends on a low fetal oxygen condition before the onset of breathing at birth. We have established an isolation protocol for mouse fetal cardiomyocytes, performed under strict low oxygen conditions to mimic the intrauterine environment, that gives the highest proliferative activities thus far reported. Oxygen exposure during isolation/culture markedly inhibited cell division and repressed cell cycle-promoting genes, and subsequent genome-wide analysis identified Fam64a as a novel regulatory molecule. Fam64a was abundantly expressed in hypoxic fetal cardiomyocyte nuclei, but this expression was drastically repressed by oxygen exposure, and in postnatal cardiomyocytes following the onset of breathing and the resulting elevation of oxygen tension. Fam64a knockdown inhibited and its overexpression enhanced cardiomyocyte proliferation. Expression of a non-degradable Fam64a mutant suggested that optimum Fam64a expression and subsequent degradation by anaphase-promoting complex/cyclosome (APC/C) during the metaphase-to-anaphase transition are required for fetal cardiomyocyte division. We propose that Fam64a is a novel cell cycle promoter of hypoxic fetal cardiomyocytes in mice.

The use of viral vectors for inner ear gene therapy is receiving increased attention for treatment of genetic hearing disorders. Most animal studies to date have injected viral suspensions into neonatal ears, via the round window membrane. Achieving transduction of hair cells, or sensory neurons, throughout the cochlea has proven difficult, and no studies have been able to efficiently transduce sensory cells in adult ears while maintaining normal cochlear function. Here, we show, for the first time, successful transduction of all inner hair cells and the majority of outer hair cells in an adult cochlea via virus injection into the posterior semicircular canal. We used a “designer” AAV, AAV2/Anc80L65, in which the main capsid proteins approximate the ancestral sequence state of AAV1, 2, 8, and 9. Our injections also transduced ~10% of spiral ganglion cells and a much larger fraction of their satellite cells. In the vestibular sensory epithelia, the virus transduced large numbers of hair cells and virtually all the supporting cells, along with close to half of the vestibular ganglion cells. We conclude that this viral vector and this delivery route hold great promise for gene therapy applications in both cochlear and vestibular sense organs.

Royal jelly (RJ), the exclusive food for queen bees, is taken as a dietary supplement because it is highly rich in nutrients. However, RJ is known to induce an anaphylactic response in some individuals. We evaluated in the present study the hypoallergenicity of alkaline protease-treated RJ in vitro and in vivo. We first confirmed that this treated RJ contained the same levels of vitamins, minerals and specific fatty acid as in untreated RJ. We then showed that the IgE-binding capacity of the treated RJ was very significantly reduced by conducting in vitro assays of the blood from RJ-sensitive patients. An in vivo skin-prick test on the RJ-sensitive patients also showed that, in the majority of the patients (3 out of 4 tested), the treated RJ did not evoke any allergenic response. It is thus advantageous to prepare hypoallergenic RJ by a protease enzyme treatment for its safe consumption.

In Guatemala, the Ministry of Health (MoH) began a vector control project with Japanese cooperation in 2000 to reduce the risk of Chagas disease infection. Rhodnius prolixus is one of the principal vectors and is targeted for elimination. The control method consisted of extensive residual insecticide spraying campaigns, followed by community-based surveillance with selective respraying. Interventions in nine endemic departments identified 317 villages with R. prolixus of 4,417 villages surveyed. Two cycles of residual insecticide spraying covered over 98% of the houses in the identified villages. Fourteen villages reinfestated were all resprayed. Between 2000-2003 and 2008, the number of infested villages decreased from 317 to two and the house infestation rate reduced from 0.86% to 0.0036%. Seroprevalence rates in 2004-2005, when compared with an earlier study in 1998, showed a significant decline from 5.3% to 1.3% among schoolchildren in endemic areas. The total operational cost was US$ 921,815, where the cost ratio between preparatory, attack and surveillance phases was approximately 2:12:1. In 2008, Guatemala was certified for interruption of Chagas disease transmission by R. prolixus. What facilitated the process was existing knowledge in vector control and notable commitment by the MoH, as well as political, managerial and technical support by external stakeholders.

Aims Although increased Na+/Ca2+ exchanger 1 (NCX1) expression is observed during heart failure (HF), the pathological role of NCX1 during the progression of HF remains unclear. We examined alterations of NCX1 expression and activity in hearts after transverse aortic constriction (TAC) surgery and explored whether NCX1 influences pressure overload-induced pathological cardiac remodelling. Methods and results We generated novel transgenic mice in which NCX1 expression is controlled by a cardiac-specific, doxycycline (DOX)-dependent promoter. In the absence of DOX, TAC surgery caused substantial chamber dilation with a gradual decrease in contractility by 16 weeks. Cardiomyocytes showed a decline in contractility with abnormal Ca2+ handling during excitation–contraction (E–C) coupling. Reduced NCX1 activity was observed 8 weeks after TAC and was still apparent at 17 weeks. Induced NCX1 overexpression by DOX treatment starting 8 weeks after TAC returned NCX1 activity to pre-TAC levels and prevented chamber dilation with cardiac dysfunction. DOX treatment not only upregulated NCX1 expression in TAC-operated hearts but also returned L-type Ca2+ channel and sarcoplasmic reticulum (SR) Ca2+ ATPase expression levels to those in sham-operated hearts. In DOX-treated myocytes, contractility, T-tubule integrity, synchrony of Ca2+ release from the SR, and Ca2+ handling during E–C coupling was preserved 16 weeks after TAC surgery. In addition, DOX treatment attenuated the down-regulation of survival signalling and up-regulation of apoptosis signalling 16 weeks after TAC surgery. Conclusion Induced overexpression of NCX1 attenuated pressure overload-induced pathological cardiac remodelling. Thus, maintaining NCX1 activity may be a potential therapeutic strategy for preventing the progression of HF.

In response to atherogenic stimuli, blood monocytes transmigrate across the vascular endothelium not only through endothelial cell-cell junctions (para-cellular) but also through endothelial cells themselves (trans-cellular). The molecular mechanism of the latter is mostly unknown, because it rarely happens, especially in vitro. Although many reports have recognized trans-cellular migration from snapshot images of leukocytes halfway across the endothelium at non-junctional locations, it often produces a false-positive result, because some leukocytes that initiate trans-cellular migration withdraw and return to the apical endothelial surface. Thus, analyzing the entire process is essential. In this study, complete monocyte trans-cellular migration was successfully captured for live cells, with simultaneous visualization of endothelial PECAM-1. We suggest the possible existence of both PECAM-1-related migration at peri-junctional sites and PECAM-1-unrelated migration at sites remote from junctions. This is the first report to describe the entire process of monocyte trans-cellular migration for live cells and its relationship with endothelial PECAM-1.

Monocyte accumulation in the arterial intima is a hallmark in early atherosclerosis. Monocyte extravasation involves sequential processes including rolling, adhesion, and transmigration across vascular endothelial cells (ECs), where abundant mechanical interactions between these two cells exist. Platelet-endothelial cell adhesion molecule-1 (PECAM-1) is a junctional protein expressed on both cells, and participates in paracellular transmigration via homophilic binding between these cells, while PECAM-1 binds homophilically between neighboring ECs in the absence of monocytes. During monocyte transmigration, PECAM-1-bearing membrane can be recruited to the transmigration spot from intracellular pools. The mechanism by which PECAM-1 binding between ECs in control state is switched to that between monocytes and ECs during transmigration, and its relationship with recruited PECAM-1 remain unclear. In this study, we built an experimental model in vitro for studying molecular behavior of PECAM-1 during monocyte transmigration. A plasmid vector containing PECAM-1 tagged with green (GFP) or red (DsRed) fluorescent protein was constructed, and separately transfected into ECs. The mixture of both transfectants in culture achieved a monolayer that contains an intercellular PECAM-1 boundary between PECAM-1-GFP- and PECAM-1-DsRed-expressing cells which is visualized as yellow in merged image. Using this model together with confocal laser scanning microscope-based system, molecular behavior of PECAM-1 on neighboring ECs during monocyte transmigration was observed in live cells. This model can be essential to directly visualize binding/dissociation state of PECAM-1 between neighboring ECs during monocyte trans-endothelial migration in relation to mechanical monocyte-EC interactions.

Two new spirostanol glycosides (1, 2) and a new furostanol glycoside (3), together with nine known steroidal glycosides (4-12) were isolated from the leaves of Furcraea foetida (Agavaceae). The structures of the new compounds were determined by spectroscopic analysis and the results of hydrolytic cleavage. The isolated compounds were evaluated for their cytotoxic activities against HL-60 human leukemia cells, A549 human lung adenocarcinoma cells, HSC-2 human oral squamous carcinoma cells, and HSC-4 human oral squamous carcinoma cells.

The effect of bee pollen cistus extract on serum and bone biochemical components in streptozotocin (STZ)diabetic rats was investigated. The water-solubilized extracts were obtained from the bee pollen of Cistus ladaniferus .R ats received a single subcutaneous administration of STZ (6.0 mg/100 g body weight), and then the animals were orally administered water-solubilized extract (5, 10, or 20 mg/100 g body weight) of bee pollen cistus once daily for 14 days. The administration of STZ caused a significant decrease in body weight and a significant increase in serum glucose, triglyceride, and calcium levels, indicating a diabetic state. These alterations were significantly prevented by the administration of the extract (5, 10, or 20 mg/100 g). Serum inorganic phosphorus concentration was significantly decreased in STZ-diabetic rats, and the decrease was significantly prevented after administration of the extract of 10 or 20 mg/100 g. Calcium content and alkaline phosphatase activity in the femoral-diaphyseal and -metaphyseal tissues were significantly decreased in STZ-diabetic rats. These decrease were significantly prevented after administration of the extract of 10 or 20 mg/100 g. The disphyseal DNA content was also significantly decreased in STZ-diabetic rats. This decrease was significantly prevented after the administration of the extract of 10 or 20 mg/100 g. This study demonstrates that the intake of bee pollen cistus extract has preventive effects on bone loss in STZ-diabetic rats, and that the intake has partial restorative effects on serum biochemical findings with the diabetic state.

Four phenolic compounds were isolated from Japanese butterbur (Petasites japonicus) by preparative RP-HPLC technique, and were identified as chlorogenic acid (CGA), fukinolic acid (FA), 3,5-dicaffeoyl quinic acid, and 3,4,5-tricaffeoyl quinic acid. Since the antioxidant activity of FA among these four compounds has not been reported in detail, we investigated the scavenging activity of FA against several active oxygens and radicals, compared with CGA, gallic acid (GA) and epigallocatechin gallate (EGCG), using ESR spectroscopy. The order of potency of scavenging superoxide anion, nitric oxide (NO) and DPPH was EGCG>FA>CGA. The order of potency of scavenging hydroxyl radical was EGCG>FA>GA. FA inhibited the lipopolysaccharide-stimulated NO production and inducible NO synthase protein and mRNA expression by mouse macrophage-like cells Raw 264.7 more potently than CGA. The present study suggests the usefulness of FA as an antioxidant.

Melinjo (Gnetum gnemon L.) seed extracts (MSEs) are rich in resveratrol dimers (gnemonoside A, C, D, gnetin C), trans-resveratrol, and other resveratrol derivatives. trans-Resveratrol is a widely studied caloric restriction mimetic. In mice fed a high-fat diet (HFD), trans-resveratrol protects against obesity, type 2 diabetes, and premature death. Here, treatment of HFD-fed mice with 2.0% MSE significantly reduced body weight gain (p

On March 11, 2011, Japan's northeast Pacific coast was hit by a gigantic earthquake and subsequent tsunami. Soma City in Fukushima Prefecture is situated approximately 44 km north of Fukushima Daiichi Nuclear Power Plant. Soma General Hospital is the only hospital in Soma City that provides full-time otolaryngological medical care. We investigated the changes in new patients from one year before to three years after the disaster. We investigated 18,167 new patients treated at our department during the four years from April 1, 2010 to March 31, 2014. Of the new patients, we categorized the diagnoses into Meniere's disease, acute low-tone sensorineural hearing loss, vertigo, sudden deafness, tinnitus, and facial palsy as neuro-otologic symptoms. We also investigated the changes in the numbers of patients whom we examined at that time concerning other otolaryngological disorders, including epistaxis, infectious diseases of the laryngopharynx, and allergic rhinitis. The total number of new patients did not change remarkably on a year-to-year basis. Conversely, cases of vertigo, Meniere's disease, and acute low-tone sensorineural hearing loss increased in number immediately after the disaster, reaching a plateau in the second year and slightly decreasing in the third year. Specifically, 4.8% of patients suffering from these neuro-otologic diseases had complications from depression and other mental diseases. With regard to new patients in our department, there was no apparent increase in the number of patients suffering from diseases other than neuro-otologic diseases, including epistaxis, and allergic rhinitis. Patients suffering from vertigo and/or dizziness increased during the first few years after the disaster. These results are attributed to the continuing stress and tension of the inhabitants. This investigation of those living in the disaster area highlights the need for long-term support.

Background External auditory canal cholesteatoma (EACC) is a rare otologic disease, characterized by focal osteonecrosis, sequestration and overlying epithelial loss of the bony external auditory canal (EAC). The etiology and pathogenesis of EACC remain controversial. There are only 2 reports on the association between EACC and chronic renal failure (CRF)/hemodialysis (HD). METHOD & RESULT: (1) This study reviewed seven EACC cases with CRF. The mean age was 68.4 years (range: 56 -81 years), and the male-female ratio was 5:2. There were 12 ears with EACC (5 cases were bilateral and 2 cases were unilateral). The EACCs were found in the inferior or posterior inferior part of the EAC in 11 ears. The number of the ears in stage III or IV was 6. Five cases were on HD. (2) Seventy-six cases with CRF on HD were examined for EACC, and 2 out of those 70 cases were diagnosed as having EACC. (3) A comparative study of the 7 EACC cases on HD and 68 non-EACC cases on HD revealed no significant differences in the sex, age, period on CRF/HD, complications (diabetes mellitus or skin disease), smoking, ear cleaning and the use of an earphone or a hearing aid. Conclusion & discussion Six out of all 9 EACC cases with CRF developed bilaterally, and in the inferior or posterior inferior part of the EAC, which implies a common pathological condition that contributes to the development of EACC. The mean age of 9 patients with EACC was relatively older (66.7 years), therefore age-related changes in the EAC are suspected in the cases with CRF on HD. The patients with CRF on HD have a high incidence of EACC. This strongly suggests the association between EACC and CRF/HD, but the mechanism of this pathogenesis has not been revealed.

The effects of bee pollen extract on bone components in the femoral-diaphyseal (cortical bone) and -metaphyseal (trabecular bone) tissues of rats in vitro and in vivo were investigated. Bone tissues were cultured for 48 hr in serum-free Dulbecco's modified Eagle's medium containing either vehicle or water- or ethanol-solubilized extracts (10, 100, or 1000 μg/ml of medium) obtained from the bee pollen of Cistus ladaniferus. Calcium content in the femoral-diaphyseal or -metaphyseal tissues was significantly increased in the presence of water-solubilized extract (100 or 1000 μg/ml) and ethanol-solubilized extract (1000 μg/ml). An increase was also observed in the presence of water-solubilized extract (100 μg/ml) obtained from Fagopyrum esculentum, Camellia sinesis, or Brassica napus L. Alkaline phosphatase activity and DNA content in the femoral-diaphyseal or -metaphyseal tissues in vitro were significantly increased in the presence of water-solubilized extract (100 or 1000 μg/ml) obtained from the bee pollen. The effects of the bee pollen extract (100 μg/ml) in increasing bone components were completely inhibited in the presence of cycloheximide (10-6 M), an inhibitor of protein synthesis, in vitro. Moreover, the calcium content and alkaline phosphatase activity in the femoral-diaphyseal or -metaphyseal tissues were significantly increased by the oral administration of water-solubilized extracts (5 or 10 mg/100 g body weight) obtained from the bee pollen of Cistus ladaniferus once daily for 7 days. The DNA content in the diaphyseal or metaphyseal tissues was significantly increased by the oral administration of water-solubilized extract (10 mg/100 g) of bee pollen cistus. The dose of 1.0 mg/100 g caused a significant increase in the diaphyseal and metaphyseal alkaline phosphatase activity or the metaphyseal DNA content in vivo. This study demonstrates that the extract of bee pollen has an anabolic effect on bone components in rats in vitro and in vivo.

Royal jelly (RJ) has been used worldwide for many years as medical products, health foods and cosmetics. Since RJ contains testosterone and has steroid hormone-type activities, we hypothesized that it may have beneficial effects on osteoporosis. We used both an ovariectomized rat model and a tissue culture model. Rats were divided into eight groups as follows: sham-operated (Sham), ovariectomized (OVX), OVX given 0.5% (w/w) raw RJ, OVX given 2.0% (w/w) RJ, OVX given 0.5% (w/w) protease-treated RJ (pRJ), OVX given 2.0% (w/w) pRJ, OVX given 17β-estradiol and OVX given its vehicle, respectively. The Ovariectomy decreased tibial bone mineral density (BMD) by 24%. Administration of 17β-estradiol to OVX rats recovered the tibial BMD decrease by 100%. Administration of 2.0% (w/w) RJ and 0.5–2.0% (w/w) pRJ to OVX rats recovered it by 85% or more. These results indicate that both RJ and pRJ are almost as effective as 17β-estradiol in preventing the development of bone loss induced by ovariectomy in rats. In tissue culture models, both RJ and pRJ increased calcium contents in femoral-diaphyseal and femoral-metaphyseal tissue cultures obtained from normal male rats. However, in a mouse marrow culture model, they neither inhibited the parathyroid hormone (PTH)-induced calcium loss nor affected the formation of osteoclast-like cells induced by PTH in mouse marrow culture system. Therefore, our results suggest that both RJ and pRJ may prevent osteoporosis by enhancing intestinal calcium absorption, but not by directly antagonizing the action of PTH.

The bee pollen Cistus ladaniferus (C. ladaniferus) extract has an anabolic effect on bone metabolism. The effects of the fractionated extracts obtained from bee pollen on bone calcium content and osteoclast-like cell formation in vitro were investigated. Rat femoral-diaphyseal and -metaphyseal tissues were cultured for 48 hr in a medium containing either vehicle or a water-solubilized extract with the membrane fractions obtained from bee pollen. The active component of bee pollen in increasing calcium content in diaphyseal tissues was seen in the fraction of molecular weight (MW) of less than 1000, and it was not observed in fractions of greater than MW 1000. Culture with the fractionated bee pollen extract (25 or 50 µg/ml of medium) of less than MW 1000 caused a significant increase in calcium content in the diaphyseal or metaphyseal tissues. The parathyroid hormone (PTH; 10 ‐6 M)induced decrease in diaphyseal calcium content was significantly prevented in the presence of the fractionated bee pollen extracts (10 µg/ml) of less than MW 1000 or greater than MW 1000. Mouse marrow cells were cultured for 7 days in a medium containing PTH (10 ‐6 M) in the presence or absence of the fractionated bee pollen extract (10 or 50 µg/ml). The PTH-induced increase in osteoclast-like cell formation was markedly suppressed in the presence of extracts of less than MW 1000 as compared with that in the presence of fractions of greater than MW 1000. The effects of the fractionated bee pollen extracts of less than MW 1000 in increasing diaphyseal calcium content and in inhibiting PTH-induced osteoclastic cell formation were significantly decreased upon heat treatment for 20 and 60 min at 80°C. This study demonstrates that the active component of bee pollen C. ladaniferus extract, which stimulates bone formation and inhibits osteoclastic bone resorption, is the fraction with MW less than 1000.

We demonstrated recently that chronic administration of aldosterone to rats induces glomerular mesangial injury and activates mitogen-activated protein kinases including extracellular signal-regulated kinases 1/2 (ERK1/2). We also observed that the aldosterone-induced mesangial injury and ERK1/2 activation were prevented by treatment with a selective mineralocorticoid receptor (MR) antagonist, eplerenone, suggesting that the glomerular mesangium is a potential target for injuries induced by aldosterone via activation of MR. In the present study, we investigated whether MR is expressed in cultured rat mesangial cells (RMCs) and involved in aldosterone-induced RMC injury. MR expression and localization were evaluated by Western blotting analysis and fluorolabeling methods. Cell proliferation and micromechanical properties were determined by [ 3 H]-thymidine uptake measurements and a nanoindentation technique using an atomic force microscope cantilever, respectively. ERK1/2 activity was measured by Western blotting analysis with an anti-phospho–ERK1/2 antibody. Protein expression and immunostaining revealed that MR was abundant in the cytoplasm of RMCs. Aldosterone (1 to 100 nmol/L) dose-dependently activated ERK1/2 in RMCs with a peak at 10 minutes. Pretreatment with eplerenone (10 μmol/L) significantly attenuated aldosterone-induced ERK1/2 phosphorylation. Aldosterone (100 nmol/L) treatment for 30 hours increased [ 3 H]-thymidine incorporation and decreased the elastic modulus, indicating cellular proliferative and deforming effects of aldosterone, respectively. These aldosterone-induced changes in cellular characteristics were prevented by pretreatment with eplerenone or an ERK (MEK) inhibitor, PD988059 (100 μmol/L). The results indicate that aldosterone directly induces RMC proliferation and deformability through MR and ERK1/2 activation, which may contribute to the pathogenesis of glomerular mesangial injury.

胃切除後21年目に発症した空腸残胃腸重積症の1例を経験したので報告する.症例は71歳,男性,コーヒー残渣様嘔吐のため当院紹介,入院となった.既往歴は50歳時に胃潰瘍のため胃切除術を施行されていた.現症は腹部は平坦で上腹部に軽い疼痛と腹膜刺激症状を認めた.血液検査では特記すべき異常所見は認めなかった. X線透視では残胃内に陰影欠損像がみられ, CTでは上腹部に重層構造を示す腸管像を認めた.内視鏡では残胃内に暗赤色蛇腹様腫瘍所見を認めた.以上より空腸残胃重積症と診断,手術を施行した.開腹すると初回手術はビルロートII法で行われており残胃内への輸出脚小腸の重積と,さらにその小腸内にも小腸の重積があり三筒性の腸重積であった.用手整復後,壊死腸管を切除,ブラウン吻合,腸間膜への輸出脚固定を付加し手術を終了した.術後5年経過後の現在,再発所見はなく良好である.

Background There are relatively few studies about the antiproliferative effects of codeine-related compounds on human cancer cell lines, compared with those of morphine-related compounds. The authors previously found that codeinone, an oxidation metabolite of codeine, among 10 opioids, showed the highest cytotoxic activity (DNA fragmentation-inducing activity) against human promyelocytic leukemic cell lines (HL-60). This was counteracted by an antioxidant, N-acetyl-L-cysteine (NAC). These findings prompted us to perform a more detailed study of apoptosis induction after codeinone treatment. Methods Apoptosis was induced by treating HL-60 cells for 1-6 h with codeine or codeinone. DNA fragmentation was assessed by both agarose gel electrophoresis and fluorometric determination of the fragmented DNA after staining with diamidinophenylindole (DAPI). The appearance of apoptotic cells was monitored by microscopic observation after staining with Hoechst (H)-33342, and fluorescence activated cell sorter (FACS) after staining with Annexin. The release of cytochrome c and cytochrome oxidase from mitochondria and activation of caspase 3 were monitored by Western blot analysis. Intracellular caspase 3-like activity was confirmed by FACS, using cell permeable substrate. Mitochondrial manganese-containing superoxide dismutase (MnSOD) activity and mRNA expression were assayed by activity staining after separation on the polyacrylamide gel electrophoresis, and reverse transcriptase-polymerase chain reaction (RT-PCR), respectively. Results Codeinone induced internucleosomal DNA fragmentation and production of Annexin-positive apoptotic cells more potently than codeine in HL-60 cells. Codeinone stimulated the release of both cytochrome c and cytochrome oxidase, and cleavage of procaspase 3 without significant changes in both the activity and expression of MnSOD. Conclusions Codeinone was found to possess both apoptosis and necrosis-inducing activity, in addition to the reported antinociceptive activity, further substantiating its antitumor potential.

The interaction between monocytes and endothelial cells is considered to play a major role in the early stage of atherosclerosis, and the involved endothelial cell micromechanics may provide us with important aspects of atherogenesis. In the present study, we evaluated (i) the endothelial cell-to-cell and cell-to-substrate gaps with the electric cell-substrate impedance sensing system, which can detect the nanometer order changes of cell-to-cell and cell-to-substrate distances separately, and (ii) the endothelial cell micromechanical properties with an atomic force microscope after application of monocytes to endothelial cells. Application of monocytic THP-1 cells to IL-1β-stimulated human umbilical vein endothelial cells immediately decreased the electrical resistance of the endothelial cell-to-substrate (increase of the cell-to-substrate gap), whereas the endothelial cell-to-cell resistance (cell-to-cell gap) did not change. The elastic modulus of the endothelial cells decreased after 2-h monocyte application, indicating an increase of endothelial cell deformability. In conclusion, the interaction of the monocytes to the endothelial cells reduced the adhesiveness to the substrate and increased the deformability of endothelial cells. These changes in the adhesiveness and the deformability may facilitate migration of monocytes, a key process of atherogenesis in the later stage.

As an evaluation scheme, we propose certifying for "control", as alternative to "interruption", of Chagas disease transmission by native vectors, to project a more achievable and measurable goal and sharing good practices through an "open online platform" rather than "formal certification" to make the key knowledge more accumulable and accessible.

Sequences of the folP1 , rpoB , and gyrA genes were analyzed for 88 isolates of Mycobacterium leprae from leprosy patients in Japan, Haiti, Indonesia, Pakistan, and the Philippines. Thirteen isolates (14.8%) showed representative mutations in more than two genes, suggesting the emergence of multidrug-resistant M. leprae .

A 67-year-old man was seen at the hospital because of general fatigue. Examination of the blood disclosed macrocytic and hyperchromic anemia, increases in LDH isozyme I and II, an increase in serum gastrins level, and a decrease in vitamin B12. Based on these findings together with positive reactions to anti-gastric wall antibody and anti-intrinsic factor antibody, the patient was diagnosed as having pernicious anemia. Fluoroscopic and endoscopic examinations of the stomach revealed a group of irregular shaped small elevations covering from the posterior wall of the middle portion to angular notch, and it was diagnosed as early gastric cancer of IIa grouped type. Operative procedures included distal gastrectomy and D 1+No 7 lymph nodes dissection. It is believed that gastric cancer with pernicious anemia occurs at a relatively low frequency in Japan compared with those in Western countries. This paper presents a recent case of early gastric cancer associated with perinicious anemia, together with a review of 61 cases seen in the Japanese literature. Gastric cancers associated with perinicious anemia are characterized by elevated lesion, a dominant occurrence of well differentiated adenocarcinoma, no particular common site, and a tendency of multiple occurrence.

A case with an alpha-fetoprotein (AFP)-producing carcinoma originating from the rectum is described. A 71-year-old male patient, who underwent a rectectomy for rectal carcinoma, developed space occupying lesions in the liver and a remarkable AFP elevation (220,000 ng/ml) in the 6th month postoperatively, and then expired one year later. Histologically, the rectal carcinoma consisted of well-differentiated adenocarcinomas, and contiguous cancerous cells proliferating in either a medullary or trabecular pattern. In the "trabecular" areas, localization of AFP was confirmed immunohistochemically. Results of concanavalin A or lens culinaris agglutinin affinity chromatography demonstrated that it was between a hepatic type and a yolk sac type, and was considered to be an intestinal type. This could have been an AFP-producing rectal carcinoma, in which the patient experienced liver metastasis at a relatively early postoperative period and died. This shows that AFP-producing rectal carcinomas are highly malignant, biologically, similar to AFP-producing gastric cancers.

Melinjo (Gnetum gnemon L.) is widely cultivated in Southeast Asia. Its fruit and seeds are common ingredients in Indonesian foods. The seeds are very rich in resveratrol dimers such as gnetin C and its glucosides, gnemonoside A and gnemonoside D, and also contain trans-resveratrol and its glucoside, trans-piceid. The safety of melinjo seeds is assured, since people in Southeast Asia have consumed them for a long time; however, their safety has not been scientifically verified. In this study, the safety of melinjo seed extract (MSE) powder was assessed in an acute oral toxicity study, a 4-week repeated dose toxicity study, and in a micronucleus test in rats. In the acute and subchronic toxicity studies, the group administered the powder did not show any toxicologically significant MSE-related changes, compared with the control group. The no observed adverse effect level (NOAEL) was determined as 1000 mg/kg/day. A genotoxicity test (rat bone marrow micronucleus test) was negative for MSE powder at levels up to 4000 mg/kg/day. These results might provide supportive evidence of safety of melinjo seeds, which has been used as food ingredients for a long time.

Background: Left ventricular (LV) hypertrophy is often present in patients with diastolic heart failure. However, stiffness of hypertrophied cardiomyocytes in the transverse direction has not been fully elucidated. The aim of this study was to assess passive cardiomyocyte stiffness of hypertrophied hearts in the transverse direction and the influence of actin-myosin cross-bridge formation on the stiffness. Methods and Results: Wistar rats received a vehicle (control) or isoproterenol (ISO) subcutaneously. After 7 days, compared with the controls, ISO administration had significantly increased heart weight and LV wall thickness and had decreased peak early annular relaxation velocity (e’) assessed by echocardiography. Elastic modulus of living cardiomyocytes in the transverse direction assessed by an atomic force microscope was significantly higher in the ISO group than in controls. We added butanedione monoxime (BDM), an inhibitor of actin-myosin interaction, and blebbistatin, a specific myosin II inhibitor, to the medium. BDM and blebbistatin significantly reduced the elastic modulus of cardiomyocytes in the ISO group. X-ray diffraction analysis showed that the reflection intensity ratio (I(1,0)/I(1,1)) at diastole was not different before and after treatment with BDM, which induces complete relaxation, in control hearts, but that I(1,0)/I(1,1) was significantly increased after BDM treatment in the ISO group, indicating residual cross-bridge formation in hypertrophied hearts. Conclusions: Passive cardiomyocyte stiffness in the transverse direction is increased in hearts with ISO-induced hypertrophy and this is caused by residual actin-myosin cross-bridge formation. (Circ J 2013; 77: 741–748)

Rhodnius prolixus is one of the main vectors of Trypanosoma cruzi, causative agent of Chagas disease. In Central America, it was first discovered in 1915 in El Salvador, from where it spread northwest to Guatemala and Mexico, and southeast to Nicaragua and Costa Rica, arriving also in Honduras in the late 1950s. Indoor residual spraying (IRS) by the antimalaria services of Costa Rica prevented its spread southwards, and similar IRS programmes appear to have eliminated it from El Salvador by the late 1970s. In 1997, by resolution of the Ministers of Health of the seven Central American countries, a multinational initiative against Chagas disease (IPCA) was launched with one of the specific objectives being the elimination of R. prolixus from the region. As a result, more and more infested areas were encountered, and progressively sprayed using an IRS strategy already deployed against Triatoma infestans in the southern cone countries of South America. In 2008, Guatemala became the first of these countries to be formally certified as free of Chagas disease transmission due to R. prolixus. The other infested countries have since been similarly certified, and none of these has reported the presence of R. prolixus since June 2010. Further surveillance is required, but current evidence suggests that R. prolixus may now been eliminated from throughout the mesoamerican region, with a corresponding decline in the incidence of T. cruzi infections.

Human scalp skins were treated with 20 mM ethylenediaminetetraacetic acid and terminal hair follicles were extracted with the epidermis. Some terminal hair follicles were morphologically preserved well and provided opportunity to examine three-dimensional distribution of CAM5.2 (K8, 52.5 kD) reactive Merkel cells. In anagen terminal hair of the scalp numerous immunoreactive Merkel cells were distributed in the presumptive bulge area. Distinct swelling as in the bulge of human vellus hair was usually absent; however, in rare instances anagen terminal hair demonstrated unilateral prominent swelling with dense aggregation of Merkel cells. In telogen hair the bulge becomes indistinguishable from the regressed end of the club hair follicle but Merkel cells continued to be abundant. We found morphologic variation of the bulge such as formation of knoblike swellings and villous projections. Interestingly, Merkel cells were also located in these structures. Palisading stockade-like nerve endings were observed surrounding the follicular epithelium at the sebaceous gland level. Merkel cells were sparse in this follicular segment. Variable number of Merkel cells were also scattered in the infundibulum of terminal hair in no association with peripheral nerves.