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This study is the first to demonstrate that deep ocean water (DOW) has physiological significant effects on squid. After 36 h of rearing squids, those reared with DOW had significantly higher total and free cholesterol levels and lower alanine transaminase activity in hemolymph as compared with those reared with surface water (SSW). SSW rearing also resulted in 6.95% weight loss, while DOW rearing caused only 2.5% weight loss, which might be due to liver metabolism suppression. Furthermore, both monovalent (sodium, chloride, and potassium ions) and divalent (calcium, inorganic phosphorus, and magnesium ions) ions in hemolymph were elevated when reared with DOW compared to those when reared with SSW. A study of genes expressed in the brain revealed that five genes were specifically remarked in DOW rearing. Most altered genes were neuropeptides, including those from vasopressin superfamily. These neuropeptides are involved in cholesterol and/or mineral metabolisms and physiological significant effects on squid. This study is the first report the effects of DOW on cholesterol and mineral metabolism of squid and will contribute to squid aquaculture using DOW.

We previously reported that the oral administration of melatonin from 4 to 20 months to male mice improved femoral bone strength and bone density during the aging. Additionally, melatonin receptor, MT2, was immunologically detected in both osteoblasts and osteoclasts of the mouse femoral bone. Thus, melatonin can act on both osteoblasts and osteoclasts to maintain bone strength during the aging process. Here, we analyzed plasma calcium (Ca2+), magnesium (Mg2+), and inorganic phosphorus ([PO4]3-) in 20-month-old male mice with or without administration melatonin (15-20 mg/kg/day) in drinking water. We found that plasma Ca2+ and Mg2+ levels in melatonin-treated mice increased significantly as compared with control mice. In [PO4]3-, melatonin administration tended to increase its plasma level, but did not reach statistical significance. The potential association between these divalent ions and metabolism markers of femoral bone was also examined. In the femoral diaphysis, the plasma Ca2+ and Mg2+ concentrations were positively correlated with periosteal and endosteal circumference which were significantly associated with the Strength Strain Index. Therefore, melatonin treatment enlarged femoral diaphysis and enhanced bone strength by increasing mineral depositions. In addition, the plasma melatonin levels were significantly positive correlation with total bone density and critical thickness in the femoral diaphysis. Since we had not observed the primary trabecular bone and osteoclasts in 20-month-old mice previously, it is suggested that plasma Ca2+ and Mg2+ are not elevated due to bone resorption. The increased plasma Ca2+ and Mg2+ by melatonin may originate from the intestinal absorption of these ions since melatonin binds to the vitamin D3 receptor, its activation is known to promote the intestinal absorption of Ca2+.

The high plasma glucose induced in glucose metabolism disorders leads to the non‐enzymatic glucose‐dependent modification (glycation) of type 1 collagen, which is an essential component of bone tissue. The glycation of proteins induces the formation of advanced glycation end‐products, such as carboxymethyl arginine, which is preferentially generated in glycated collagen. However, the effect of advanced glycation end‐product formation on the characteristics of type 1 collagen remains unclear due to the lack of suitable in vitro experimental systems analyzing type 1 collagen. Here, we show that the glycation of type 1 collagen can be analyzed in vitro using a goldfish‐scale bone model. Our study using these scales provides evidence that the advanced glycation end‐product formation in type 1 collagen induced by glyoxal, the carboxymethyl arginine inducer, facilitates the crosslinking of type 1 collagen, decreasing both its strength and flexibility., Here we show that the glycation of type 1 collagen can be analyzed in vitro using a goldfish‐scale bone model. Our study using these scales provides evidence that glycation of type I collagen alpha chains facilitates the crosslinking of type I collagen, decreasing both its strength and flexibility.

Exposure to the space environment induces a number of pathophysiological outcomes in astronauts, including bone demineralization, sleep disorders, circadian clock dysregulation, cardiovascular and metabolic dysfunction, and reduced immune system function. A recent report describing experiments aboard the Space Shuttle mission, STS-132, showed that the level of melatonin, a hormone that provides the biochemical signal of darkness, was decreased during microgravity in an in vitro culture model. Additionally, abnormal lighting conditions in outer space, such as low light intensity in orbital spacecraft and the altered 24-h light-dark cycles, may result in the dysregulation of melatonin rhythms and the misalignment of the circadian clock from sleep and work schedules in astronauts. Studies on Earth have demonstrated that melatonin regulates various physiological functions including bone metabolism. These data suggest that the abnormal regulation of melatonin in outer space may contribute to pathophysiological conditions of astronauts. In addition, experiments with high-linear energy transfer radiation, a ground-based model of space radiation, showed that melatonin may serve as a protectant against space radiation. Gene expression profiling using an in vitro culture model exposed to space flight during the STS-132 mission, showed that space radiation alters the expression of DNA repair and oxidative stress response genes, indicating that melatonin counteracts the expression of these genes responsive to space radiation to promote cell survival. These findings implicate the use of exogenous melatonin and the regulation of endogenous melatonin as countermeasures for the physiological consequences of space flight.

Nutritional and inflammatory marker ratios are known to predict response to chemotherapy in breast cancer, but whether they predict adverse effects caused by chemotherapy remains unclear. We investigated whether nutritional and inflammatory marker ratios before starting FEC therapy (5-fluorouracil, epirubicin, and cyclophosphamide) predict grade 4 neutropenia as a serious adverse effect.61 patients with breast cancer who started FEC therapy for the first time as preoperative or postoperative chemotherapy were studied. Relevant nutritional and inflammatory marker ratios were compared between patients who developed grade 4 neutropenia (n = 44) and those who did not (n = 17).In univariate analysis, occurrence of neutropenia was related significantly (p 0.05) to pre-FEC-therapy white blood cell count, platelet count, neutrophil count, lymphocyte-to-monocyte ratio (LMR), neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and modified Glasgow prognostic score. Analysis using cutoff values obtained from receiver operating characteristic curves showed that LMR, NLR, and PLR predicted grade 4 neutropenia. However, multivariate logistic regression analysis identified no independent factor associated with grade 4 neutropenia. A post-hoc power analysis revealed an inadequate sample size.Inflammatory marker ratios, especially PLR, may predict grade 4 neutropenia caused by FEC therapy for breast cancer. Although multivariate analysis identified no independent predictive markers in this study due to inadequate sample size, further prospective large-scale research is needed to examine the usefulness of nutritional and inflammatory marker ratios for predicting adverse effects.

A 66-year-old woman was diagnosed with stage IVb sigmoid colon cancer. Modified FOLFOX-6 (mFOLFOX-6; levofolinate‒fluorouracil‒oxaliplatin) plus panitumumab was selected as the chemotherapeutic regimen, but she was administered a regimen without oxaliplatin (L-OHP) or bolus 5-fluorouracil (5-FU) because of her general condition and concern about adverse effects. The patient had impaired consciousness on day 3 of chemotherapy. Computed tomography (CT) and magnetic resonance imaging (MRI) of the brain showed no findings of hemorrhage, infarction, brain metastasis, and leukoencephalopathy. Except for high blood ammonia concentration (353 µg/dL), there were no other findings that could have caused her condition. Impaired consciousness due to hyperammonemia was diagnosed. We started an intravenous drip supplemented with branched chain amino acids for liver protection. Approximately 6 hours later, blood ammonia level improved to 88 µg/dL, which approached the reference value. Consciousness level improved over time, reaching a level of alertness on day 5 after starting chemotherapy. 5-FU was suspected to be the cause of impaired consciousness due to hyperammonemia, but the exact cause could not be identified because most of the previously reported cases were given L-OHP, bolus 5-FU, and other concomitant medications. In this case, since there were no other concomitant medications, it is highly probable that continuous infusion of 5-FU alone caused impaired consciousness due to hyperammonemia. This is an important case that indicates the need to monitor carefully for the occurrence of hyperammonemia when 5-FU is administered continuously; it also proposes future issues for investigation.

Coldwater, primary freshwater, fish such as the eight-barbel loach, Lefua nikkonis, are thought to have colonised Hokkaido from the continental Far East via Sakhalin Island during the Late Pleistocene. Lefua populations have been reported on southern Sakhalin, but detailed morphological and population structure analyses have not yet been completed. This information is important for reconstructing the colonisation history of L. nikkonis to Hokkaido. In this study, morphological analysis revealed that L. nikkonis and two continental congeners, Lefua pleskei and Lefua costata, are distinguishable from each other, and Lefua collected from southern Sakhalin is morphologically more similar to L. nikkonis. Random forest analysis, a machine learning classification method, classified all Sakhalin individuals as L. nikkonis. Haplotype analysis of mitochondrial DNA sequences revealed that all but one Sakhalin haplotype are shared with L. nikkonis and none is shared with the two continental congeners, which supports the hypothesis that Sakhalin Lefua are L. nikkonis. However, none of the Sakhalin haplotypes was distributed on northern Hokkaido. This discontinuous distribution of haplotypes across Sakhalin and Hokkaido suggests that the Sakhalin Lefua populations are not native. Some of the Sakhalin haplotypes were found only on Hokkaido’s Ishikari River system or the Tokachi River system, suggesting that they originated from these regions. Because previous field surveys reported wild Lefua only from northwestern Sakhalin, we concluded that native Lefua on southern Sakhalin may have gone extinct after they colonised Hokkaido in the Middle Pleistocene.

To study the toxicity of 3-hydroxybenzo[c]phenanthrene (3-OHBcP), a metabolite of benzo[c]phenanthrene (BcP), first we compared it with its parent compound, BcP, using an in ovo-nanoinjection method in Japanese medaka. Second, we examined the influence of 3-OHBcP on bone metabolism using goldfish. Third, the detailed mechanism of 3-OHBcP on bone metabolism was investigated using zebrafish and goldfish. The LC

It is known that the bone matrix plays an important role in the response to physical stresses such as hypergravity and microgravity. In order to accurately analyze the response of bone to hypergravity and microgravity, a culture system under the conditions of coexistence of osteoclasts, osteoblasts, and bone matrix was earnestly desired. The teleost scale is a unique calcified organ in which osteoclasts, osteoblasts, and the two layers of bone matrix, i.e., a bony layer and a fibrillary layer, coexist. Therefore, we have developed in vitro organ culture systems of osteoclasts and osteoblasts with the intact bone matrix using goldfish scales. Using the scale culture system, we examined the effects of hypergravity with a centrifuge and simulated ground microgravity (g-µG) with a three-dimensional clinostat on osteoclasts and osteoblasts. Under 3-gravity (3G) loading for 1 day, osteoclastic marker mRNA expression levels decreased, while the mRNA expression of the osteoblastic marker increased. Upon 1 day of exposure, the simulated g-µG induced remarkable enhancement of osteoclastic marker mRNA expression, whereas the osteoblastic marker mRNA expression decreased. In response to these gravitational stimuli, osteoclasts underwent major morphological changes. By simulated g-µG treatments, morphological osteoclastic activation was induced, while osteoclastic deactivation was observed in the 3G-treated scales. In space experiments, the results that had been obtained with simulated g-µG were reproduced. RNA-sequencing analysis showed that osteoclastic activation was induced by the down-regulation of Wnt signaling under flight-microgravity. Thus, goldfish scales can be utilized as a bone model to analyze the responses of osteoclasts and osteoblasts to gravity.

The activation of epidermal growth factor receptor (EGFR) involves the geometrical conversion of the extracellular domain (ECD) from the tethered to the extended forms with the dynamic rearrangement of the relative positions of four subdomains (SDs); however, this conversion process has not yet been thoroughly understood. We compare the two different forms of the X-ray crystal structures of ECD and simulate the ECD conversion process using adiabatic mapping that combines normal mode analysis of the elastic network model (ENM-NMA) and energy optimization. A comparison of the crystal structures reveals the rigidity of the intradomain geometry of the SD-I and -III backbone regardless of the form. The forward mapping from the tethered to the extended forms retains the intradomain geometry of the SD-I and -III backbone and reveals the trends to rearrange the relative positions of SD-I and -III and to dissociate the C-terminal tail of SD-IV from the hairpin loop in SD-II. The reverse mapping from the extended to the tethered forms complements the promotion of ECD conversion in the presence of epidermal growth factor (EGF).

To elucidate the physiological role of calcitonin (CT) in stingrays (cartilaginous fish), an enzyme-linked immunosorbent assay (ELISA) system using a specific antibody against stingray CT has been developed. Synthetic stingray CT was subcutaneously injected into mice four times—once every 2 weeks—together with an adjuvant. We purified the IgG antibody fraction using the protein A affinity chromatography from collected antiserum. Evaluating the antibody titer, we found the antibody’s optimum dilution ratio to be 600 times. Competitive ELISA has been developed using the antibody diluted 600 times. Our antibody did not cross-react with teleost CTs and muscle extraction, but cross-reacted with stingray plasma and the extract of the ultimobranchial gland, the secretary organ of stingray CT. Using this ELISA, we measured the plasma CT level in stingrays and examined its correlation with several mineral concentrations. Plasma CT did not show significant correlation to calcium, magnesium, inorganic phosphorus, sodium, chlorine, or urea, although there was a correlation among the factors involved in osmoregulation, such as sodium, chlorine, and urea. On the other hand, plasma CT was significantly correlated to body weight and length. Furthermore, there was a significant correlation between plasma CT and gonad weight. Since plasma CT was correlated with the weight of liver, which is involved in the synthesis of egg yolk protein, we examined the influence of 17β-estradiol (E2) on CT secretion. After E2 injection, the plasma CT level increased significantly. This is the first study to demonstrate that E2 induced plasma CT secretion in cartilaginous fish.

For determining the effect of environmental pollutants on fish bone metabolism, we have developed an in vitro bioassay system using teleost scales that has osteoclasts, osteoblasts and bone matrix as markers: alkaline phosphatase (ALP) for osteoblasts and tartrate-resistant acid phosphatase (TRAP) for osteoclasts. Using this bioassay, the influence of gadolinium (Gd) on osteoclasts and osteoblasts of goldfish scales was examined in the present study. Gd sensitively inhibited TRAP activity. Even Gd at 10-13 M suppressed TRAP activity at 6 hours of incubation. At 18 hours of incubation, this inhibition occurred only at 10-7 and 10-6 M. After 36 hours of incubation, Gd did not influence TRAP activity. In osteoblasts, ALP activity was also suppressed by Gd in the range of 10-10 to 10-6 M for 6 to 18 hours of incubation. At 36 and 64 hours of incubation, ALP activity was significantly suppressed by Gd (36 hours: 10-9 to 10-6 M; 64 hours: 10-7 and 10-6 M). At 96 hours of incubation, however, Gd did not influence ALP activity. This is the first report to indicate the toxicity of Gd on fish bone metabolism using TRAP and ALP enzyme activities. The toxicity of Gd to osteoblasts is comparable to that of tributyltin, an aquatic environmental pollutant used as a biocide in anti-fouling paint. Gd is used in Magnetic Resonance Imaging (MRI) for clinical diagnoses. To avoid the toxicity of Gd ions, chelated forms, known as Gd-based contrast agents, are used for MRI diagnosis. Without a specific recycling process, these compounds are quickly released by urinary excretion and released into environmental waters. Therefore, it is possible that anthropogenic Gd influences aquatic animals. Considering our present data together with that of anthropogenic Gd pollution, we should conduct a Gd risk assessment to protect the ecosystem in the aquatic environment.

Objective In 2018, we conducted a retrospective survey using the medical records of 484 patients with type 2 diabetes. The observed value of coronary heart disease (CHD) incidence after 5 years and the predicted value by the JJ risk engine as of 2013 were compared and verified using the discrimination and calibration values. Results Among the total cases analyzed, the C-statistic was 0.588, and the calibration was p

Objective: The JJ risk engine, developed in 2012, is an algorithm that predicts the 19 incidence of diabetes complications that may develop after 5 to 10 years. However, 20 studies validating the JJ risk engine have not yet been reported;we aimed to verify the 21 JJ risk engine. In 2013, we conducted a retrospective survey using medical records of 22 484 patients with type 2 diabetes. The observed value of coronary heart disease (CHD) 23 complicationsafter 5 years and the predicted value by the JJ risk engine as of 2013 were 24 compared and verified using the discrimination and calibration values.25Results: Among the total cases analyzed, the C-statistic was 0.588,and the calibration 26 was p

Osteocytes, osteoblasts (bone-forming cells), and osteoclasts (bone-resorbing cells) are the primary types of cells that regulate bone metabolism in mammals. Sclerostin produced in bone cells activates osteoclasts, inhibiting bone formation; excess production of sclerostin, therefore, leads to the loss of bone mass. Fish scales have been reported to have morphological and functional similarities to mammalian bones, making them a useful experimental system for analyzing vertebrate bone metabolism in vitro. However, whether fish scales contain cells producing sclerostin and/or osteocytes has not been determined. The current study demonstrated, for the first time, that sclerostin-containing cells exist in goldfish scales. Analysis of the distribution and shape of sclerostin-expressing cells provided evidence that osteoblasts produce sclerostin in goldfish scales. Furthermore, our results found that osteocyte-like cells exist in goldfish scales, which also produce sclerostin. Finally, we demonstrated that microgravity in outer space increased the level of sclerostin in the scales of goldfish, a finding suggesting that the induction of sclerostin is the mechanism underlying the activation of osteoclasts under microgravity.

Background/aims Oral mesalazine is an important treatment for ulcerative colitis (UC), and non-adherence to mesalazine increases the risk of relapse. Controlled-release (CR) mesalazine has 2 formulations: tablets and granules. The relative acceptabilities of these formulations may influence patient adherence; however, they have not been compared to date. This study aimed to evaluate the acceptabilities of the 2 formulations of CR mesalazine in relation to patient adherence using a crossover questionnaire survey. Methods UC patients were randomly assigned to 2 groups in a 1:1 ratio. Patients in each group took either 4 g of CR mesalazine tablets or granules for 6 to 9 weeks, and then switched to 4 g of the other formulation for a further 6 to 9 weeks. The acceptability and efficacy were evaluated by questionnaires, and adherence was assessed using a visual analog scale. The difference in acceptabilities between the 2 formulations and its impact on adherence were assessed. Results A total of 49 patients were prospectively enrolled and 33 patients were included in the analysis. Significantly more patients found the tablets to be less acceptable than the granules (76% vs. 33%, P=0.0005). The granules were preferable to the tablets when the 2 formulations were compared directly (73% vs. 21%, P=0.004), for their portability, size, and numbers of pills. The adherence rate was slightly better among patients taking the granules (94% vs. 91%) during the observation period, but the difference was not significant (P=0.139). Conclusions CR mesalazine granules are more acceptable than tablets, and may therefore be a better option for long-term medication.

Tiger puffer, Takifugu rubripes, a commercially important long-distance migratory fish, return to specific spawning grounds for reproduction. To clarify reproductive neuroendocrine system of the tiger puffer, the changes in the expression levels of the genes encoding three gonadotropin-releasing hormones (GnRHs), gonadotropin-inhibitory hormone (GnIH), GnIH receptor (GnIH-R), kisspeptin and kisspeptin receptor in the brain and gonadotropin (GTH) subunits, growth hormone (GH) and prolactin (PRL) in the pituitary were examined in the tiger puffer captured in the wild at different reproductive stages, namely immature and mature fish of both sexes, and post-ovulatory females that were obtained by hormonal treatment. The amounts of three gnrh mRNAs, gnih, gnih-r, fshb and lhb were substantially increased in the mature fish compared to the immature fish, especially in the females, and these augmented expressions were drastically decreased in the post-ovulatory females. gh expression showed a slight increase in the mature males. In contrast, kiss2, kiss2r and prl did not show significant changes in the males but significantly decreased in the post-ovulatory females. The present results demonstrate the expression dynamics of the hypothalamo-pituitary-gonadal axis genes associated with the reproductive conditions and the possible involvement of the GnRH/GnIH/GTH system in the regulation of the sexual maturation and spawning in the wild tiger puffer.

Melatonin has recently been found to be a possible new regulator of bone metabolism. However, the influence of melatonin in natural age-related osteoporosis has not been fully elucidated yet, although there have been some reports regarding postmenopausal osteoporosis with melatonin treatments. The present study investigated the effects of long-term melatonin administration during the aging process on bone metabolism. Using quantitative computed tomography methods, we found that the total bone density of both the femur metaphysis and diaphysis decreased significantly in 20-month-old male mice. In the metaphysis, both trabecular bone mass and Polar-Strength Strain Index (SSI), which is an index of bone strength, decreased significantly. Judging from bone histomorphometry analysis, trabecular bone in 20-month-old male mice decreases significantly with age and is small and sparse, as compared to that of 4-month-old male mice. Loss of trabecular bone is one possible cause of loss of bone strength in the femoral bone. In the metaphysis, the melatonin administration group had significantly higher trabecular bone density than the non-administration group. The Polar-SSI, cortical area, and periosteal circumference in the diaphysis was also significantly higher with melatonin treatments. Since the melatonin receptor, MT2, was detected in both osteoblasts and osteoclasts of the femoral bone of male mice, we expect that melatonin acts on osteoblasts and osteoclasts to maintain the bone strength of the diaphysis and metaphysis. Thus, melatonin is a potential drug for natural age-related osteoporosis.

Sex steroid hormones including estrogens and androgens play fundamental roles in regulating reproductive activities and they act through estrogen and androgen receptors (ESR and AR). These steroid receptors have evolved from a common ancestor in association with several gene duplications. In most vertebrates, this has resulted in two ESR subtypes (ESR1 and ESR2) and one AR, whereas in teleost fish there are at least three ESRs (ESR1, ESR2a and ESR2b) and two ARs (ARα and ARβ) due to a lineage-specific whole genome duplication. Functional distinctions have been suggested among these receptors, but to date their roles have only been characterized in a limited number of species. Sexual differentiation and the development of reproductive organs are indispensable for all animal species and in vertebrates these events depend on the action of sex steroid hormones. Here we review the recent progress in understanding of the functions of the ESRs and ARs in the development and expression of sexually dimorphic characteristics associated with steroid hormone signaling in vertebrates, with representative fish, amphibians, reptiles, birds and mammals.

Objective This study was designed to clarify the factors affecting the efficacy, adverse events, and pharmacokinetics of fondaparinux in Japanese patients undergoing artificial knee replacement surgery. Materials and methods Fondaparinux (1.5 mg/d) was administered subcutaneously to patients (n = 30) at 24 hours after surgery, and blood samples were taken at various time points thereafter. Venous thromboembolism (VTE), presence of bleeding, and pharmacokinetics were evaluated. Multivariate analysis and population pharmacokinetic analysis were performed to detect factors that necessitated withdrawal of fondaparinux and individual differences in its pharmacokinetics. Results VTE was observed in 9 patients (3 for whom administration was continued and 6 for whom withdrawal was necessary). The maximum plasma concentration of fondaparinux was found to be a significant factor determining withdrawal of the drug. Population pharmacokinetic analysis demonstrated that individual renal function and body weight were significant factors associated with apparent clearance and volume of distribution, respectively. Conclusions A high maximum plasma concentration of fondaparinux may result in subcutaneous hemorrhage, necessitating withdrawal of fondaparinux administration. The patient's kidney function and body weight also contribute to individual differences in pharmacokinetics. We recommend considering an adjustment to the dose of fondaparinux based on body weight in patients undergoing artificial knee replacement surgery. .

Acanthocephalans of the genus Corynosoma are known as intestinal parasites, mainly of pinnipeds. Human corynosomiasis has been reported as an infrequent foodborne disease in Hokkaido, the northernmost island of Japan. Potential sources of the human infection are marine fish, because they are paratenic hosts of these parasites. In this study, the prevalence and intensity of larval Corynosoma in commercial fish from 17 fishing ports of Hokkaido were examined from April 2016 to January 2019. Out of a total of 1217 fish examined, 122 (10.0%) were infected with cystacanth larvae. The infected fish assemblage was composed of 7 families and 13 species from all the coastal seas of Hokkaido (the Pacific Ocean, Okhotsk Sea, and Japan Sea), showing that commercial fish can be source of human infection when eaten raw. Flatfish of the family Pleuronectidae showed the highest intensity of cystacanths, ranging from 1 to 56. A DNA barcoding system was developed in this study, based on the standard mitochondrial cox1 sequences of morphologically identified adults of Corynosoma spp. from pinnipeds in Hokkaido. By using the DNA barcoding, most of the fish-derived cystacanths were identified as either C. strumosum or C. villosum, and furthermore, a clinical isolate from human as C. villosum. Both of the species were commonly detected from various fish of Hokkaido, irrespective of the coastal seas. Flatfish frequently harbored C. villosum. Considering the wide range of commercial fish in Hokkaido and the advanced transportation system of fresh fish, there is a possibility that human corynosomiasis will occur everywhere in Japan.