The circadian E3 ligase FBXL21 regulates myoblast differentiation and sarcomere architecture via MYOZ1 ubiquitination and NFAT signaling.

Numerous molecular and physiological processes in the skeletal muscle undergo circadian time-dependent oscillations in accordance with daily activity/rest cycles. The circadian regulatory mechanisms underlying these cyclic processes, especially at the post-transcriptional level, are not well defined. Previously, we reported that the circadian E3 ligase FBXL21 mediates rhythmic degradation of the sarcomere protein TCAP in conjunction with GSK-3ß, and Psttm mice harboring an Fbxl21 hypomorph allele show reduced muscle fiber diameter and impaired muscle function. To further elucidate the regulatory function of FBXL21 in skeletal muscle, we investigated another sarcomere protein, Myozenin1 (MYOZ1), that we identified as an FBXL21-binding protein from yeast 2-hybrid screening. We show that FBXL21 binding to MYOZ1 led to ubiquitination-mediated proteasomal degradation. GSK-3ß co-expression and inhibition were found to accelerate and decelerate FBXL21-mediated MYOZ1 degradation, respectively. Previously, MYOZ1 has been shown to inhibit calcineurin/NFAT signaling important for muscle differentiation. In accordance, Fbxl21 KO and MyoZ1 KO in C2C12 cells impaired and enhanced myogenic differentiation respectively compared with control C2C12 cells, concomitant with distinct effects on NFAT nuclear localization and NFAT target gene expression. Importantly, in Psttm mice, both the levels and diurnal rhythm of NFAT2 nuclear localization were significantly diminished relative to wild-type mice, and circadian expression of NFAT target genes associated with muscle differentiation was also markedly dampened. Furthermore, Psttm mice exhibited significant disruption of sarcomere structure with a considerable excess of MYOZ1 accumulation in the Z-line. Taken together, our study illustrates a pivotal role of FBXL21 in sarcomere structure and muscle differentiation by regulating MYOZ1 degradation and NFAT2 signaling.

Regulation of headache response and transcriptomic network by the trigeminal ganglion clock.

OBJECTIVE: To characterize the circadian features of the trigeminal ganglion in a mouse model of headache. BACKGROUND: Several headache disorders, such as migraine and cluster headache, are known to exhibit distinct circadian rhythms of attacks. The circadian basis for these rhythmic pain responses, however, remains poorly understood. METHODS: We examined trigeminal ganglion ex vivo and single-cell cultures from Per2::LucSV reporter mice and performed immunohistochemistry. Circadian behavior and transcriptomics were investigated using a novel combination of trigeminovascular and circadian models: a nitroglycerin mouse headache model with mechanical thresholds measured every 6 h, and trigeminal ganglion RNA sequencing measured every 4 h for 24 h. Finally, we performed pharmacogenomic analysis of gene targets for migraine, cluster headache, and trigeminal neuralgia treatments as well as trigeminal ganglion neuropeptides; this information was cross-referenced with our cycling genes from RNA sequencing data to identify potential targets for chronotherapy. RESULTS: The trigeminal ganglion demonstrates strong circadian rhythms in both ex vivo and single-cell cultures, with core circadian proteins found in both neuronal and non-neuronal cells. Using our novel behavioral model, we showed that nitroglycerin-treated mice display circadian rhythms of pain sensitivity which were abolished in arrhythmic Per1/2 double knockout mice. Furthermore, RNA-sequencing analysis of the trigeminal ganglion revealed 466 genes that displayed circadian oscillations in the control group, including core clock genes and clock-regulated pain neurotransmitters. In the nitroglycerin group, we observed a profound circadian reprogramming of gene expression, as 331 of circadian genes in the control group lost rhythm and another 584 genes gained rhythm. Finally, pharmacogenetics analysis identified 10 genes in our trigeminal ganglion circadian transcriptome that encode target proteins of current medications used to treat migraine, cluster headache, or trigeminal neuralgia. CONCLUSION: Our study unveiled robust circadian rhythms in the trigeminal ganglion at the behavioral, transcriptomic, and pharmacogenetic levels. These results support a fundamental role of the clock in pain pathophysiology. PLAIN LANGUAGE SUMMARY: Several headache diseases, such as migraine and cluster headache, have headaches that occur at the same time each day. We learned that the trigeminal ganglion, an important pain structure in several headache diseases, has a 24-hour cycle that might be related to this daily cycle of headaches. Our genetic analysis suggests that some medications may be more effective in treating migraine and cluster headache when taken at specific times of the day.

Low plasma tryptophan is associated with olfactory function in healthy elderly community dwellers in Japan.

BACKGROUND: Decreased circulating tryptophan (Trp) levels are frequently observed in elderly patients with neurodegenerative disease including Alzheimer's disease. Trp may serve as a potential biomarker for monitoring disease risk in elderly people. We aimed to investigate the association between low plasma Trp levels and olfactory function, which is known to predict age-related diseases including dementia in elderly people. METHODS: A total of 144 healthy elderly Japanese community (≥ 65 years old) dwellers from the Health, Aging and Nutritional Improvement study (HANI study) were the subjects of our analysis. Low Trp levels were classified using the lower limit values of the reference interval according to a previous report. Olfactory function was assessed using a card-type test called Open Essence, which includes 12 odour items that are familiar to Japanese people. The elderly subjects with low circulating Trp levels were compared to a control group with normal plasma Trp levels. RESULTS: We conducted the analyses using 144 people aged 65 years or older (mean age 73.7 ± 5.5 years; 36.1% men). The subjects showed normal serum albumin levels (4.4 ± 0.2 g/dL) and no daily living disabilities. Low plasma Trp levels (low Trp group) were found in 11.1% of the study population. The low Trp group showed a significantly lower correct-answer rate for the items india ink, perfume, curry and sweaty smelling socks than control group (P < 0.05). There was also a significant association between low Trp levels and low olfactory ability, after adjusting for age and sex. CONCLUSIONS: Lower plasma Trp levels were associated with a decrease in olfactory function in functionally competent older individuals. Because olfactory dysfunction predicts age-related diseases, low plasma Trp levels may represent a clinical sign of disease risk in elderly people.

A new voluntary blood collection method for the Andean bear (Tremarctos ornatus) and Asiatic black bear (Ursus thibetanus).

Various training methods have been developed for animal husbandry and health care in zoos and one of these trainings is blood collection. One training method, recently widely used for blood collection in Ursidae, requires setting up a sleeve outside the cage and gives access to limited blood collection sites. A new voluntary blood collection method without a sleeve was applied to the Andean bear (Tremarctos ornatus) and Asiatic black bear (Ursus thibetanus) with access to various veins at the same time. The present study evaluated the effectiveness of this new method and suggests improvements. Two Andean and two Asiatic black bears in Yokohama and Nogeyama Zoological Gardens, respectively, were trained to hold a bamboo pipe outside their cages. We could, thereby, simultaneously access superficial dorsal veins, the dorsal venous network of the hand, the cephalic vein from the carpal joint, and an area approximately 10 cm proximal to the carpal joint. This allowed us to evaluate which vein was most suitable for blood collection. We found that the cephalic vein, approximately 10 cm proximal to the carpal joint, was the most suitable for blood collection. This new method requires little or no modification of zoo facilities and provides a useful alternative method for blood collection. It could be adapted for use in other clinical examinations such as ultrasound examination.

FOLLICULAR CELL CARCINOMA OF THE THYROID GLAND IN THREE CAPTIVE AGED RACCOON DOGS (NYCTEREUTES PROCYONOIDES).

The clinical and histologic features of thyroid carcinoma in raccoon dogs have not been previously reported. Three of four raccoon dogs (Nyctereutes procyonoides) over 8 yr of age at the Nogeyama Zoological Gardens developed thyroid follicular cell carcinomas that were detected at necropsy. The affected raccoon dogs were rescued from the wild and were housed at the Nogeyama Zoological Gardens for 8 yr 8 mo, 8 yr 10 mo, and 10 yr 3 mo, respectively. Although all of them appeared lethargic and developed partial alopecia or desquamation of their skin, they did not display any other specific clinical signs associated with a thyroid lesion. Serum thyroid hormone values were examined in two of the affected raccoon dogs and the average and standard deviation values (free-thyroxin [FT4]: 0.078 ± 0.077 pM/L and 0.062 ± 0.0039 pM/L; free-triiodothyronine [FT3]: 3.261 ± 0.765 pM/L and 3.407 ± 0.919 pM/L) were lower than the reference range (FT4: 0.141 ± 0.117 pM/L; FT3: 5.139 ± 2.412 pM/L) derived from a clinically normal raccoon dog. On necropsy, the thyroid lobes were markedly enlarged bilaterally. Histopathologically, the neoplastic cells in the thyroid gland appeared round or oval and columnar or cuboidal with minimal heteromorphism. Moreover, mostly small (but occasionally large) follicles were identified, and the neoplastic cells had infiltrated into the surrounding capsule and blood vessels. The histopathologic features of the thyroid tumors in the raccoon dogs revealed that the tumors were derived from follicular cells.

Low concentrations of ethanol stimulate biofilm and pellicle formation in Pseudomonas aeruginosa.

Biofilms are communities of surface-attached microbial cells that resist environmental stresses. In this study, we found that low concentrations of ethanol increase biofilm formation in Pseudomonas aeruginosa PAO1 but not in a mutant of it lacking both Psl and Pel exopolysaccharides. Low concentrations of ethanol also increased pellicle formation at the air-liquid interface.

Effective Oral Indicators With Medical and Dental Collaboration in Open Heart Surgery: A Representative Survey.

Purpose Postoperative infections pose an important problem for patients with cardiac disease. Moreover, oral health status is associated with the risk of longer hospital stays. Therefore, the oral health status of patients was assessed before open-heart surgery. This study aimed to determine the relationship between oral health status and postoperative status. Methods The study included 25 patients who underwent open-heart surgery at our university hospital in 2020. Upon admission, dentists conducted an oral examination and assessed the oral health status of the patients, also using the Japanese version of the Oral Health Assessment Tool (OHAT-J), Revised Oral Assessment Guide (ROAG), oral moisture level, oral bacteria, and other relevant factors. The study investigated the association with postoperative status. Findings Significant postoperative infections were found in patients aged ≥70 years, with an OHAT-J score of ≥5, OHAT-J lip score of ≥1, Streptococcus γ count of 1.0 × 10^6 or higher (CFU/mL), and increased Streptococcus γ before and after surgery. The duration of hospitalization correlated with the OHAT-J, OHAT-J gum and tissue, and ROAG scores. The duration of intensive care unit (ICU) stays correlated with the OHAT-J score. Conclusions The study demonstrates that OHAT-J scores are linked with predicting not just postoperative infection but also the length of hospitalization and ICU stay. As OHAT-J scores do not necessitate specialized dental instruments, they are straightforward and beneficial for healthcare professionals outside of dentistry.

The red ruffed lemur, Varecia rubra (É. Geoffroy Saint-Hilaire, 1812): a comparative morphology investigation of lingual papillae and connective tissue cores.

The morphology of the lingual papillae (filiform, foliate, fungiform, and vallate papillae) and the underlying connective tissue core of the red ruffed lemur (Varecia rubra) of a strepsirrhines species were studied using light and scanning electron microscopy. The filiform papillae distributed at the root of the tongue were larger than the structures distributed at the body and apex. Six to eight vallate papillae were arranged in a Y-shape at the border between the lingual body and the lingual root. Foliate papillae were observed at the posterior lateral border of the tongue. Scanning electron microscopy revealed a primary process and numerous auxiliary processes in the epithelial layer of filiform papillae. After epithelial removal, the connective tissue core of the filiform papilla showed several protrusions surrounding an oval-shaped depression that extended slightly posteriorly, and a large, maple-shaped filiform papilla was seen in the posterior portion of the tongue. The connective tissue cores of the fungiform papillae exhibited a longitudinally ridged cylindrical structure. The connective tissue core of the foliate papillae had numerous tubular projections arranged along a groove with a salivary gland conduit at the base. As a Lemuridae species, the appearance of the fungiform and filiform papillae of the red ruffed lemur is similar to that reported in previous studies of the ring-tailed lemur, with some differences, especially in the filiform papillary connective tissue core at the base and tongue body border. These findings suggest the taxonomic and phylogenetic origins of the lemurs as well as the influence of dietary diversity.

ROR activation by Nobiletin enhances antitumor efficacy via suppression of IκB/NF-κB signaling in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a heterogeneous disease characterized by poor response to standard therapies and therefore unfavorable clinical outcomes. Better understanding of TNBC and new therapeutic strategies are urgently needed. ROR nuclear receptors are multifunctional transcription factors with important roles in circadian pathways and other processes including immunity and tumorigenesis. Nobiletin (NOB) is a natural compound known to display anticancer effects, and our previous studies showed that NOB activates RORs to enhance circadian rhythms and promote physiological fitness in mice. Here, we identified several TNBC cell lines being sensitive to NOB, by itself or in combination. Cell and xenograft experiments showed that NOB significantly inhibited TNBC cell proliferation and motility in vitro and in vivo. ROR loss- and gain-of-function studies showed concordant effects of the NOB-ROR axis on MDA-MB-231 cell growth. Mechanistically, we found that NOB activates ROR binding to the ROR response elements (RRE) of the IκBα promoter, and NOB strongly inhibited p65 nuclear translocation. Consistent with transcriptomic analysis indicating cancer and NF-κB signaling as major pathways altered by NOB, p65-inducible expression abolished NOB effects, illustrating a requisite role of NF-κB suppression mediating the anti-TNBC effect of NOB. Finally, in vivo mouse xenograft studies showed that NOB enhanced the antitumor efficacy in mammary fat pad implanted TNBC, as a single agent or in combination with the chemotherapy agent Docetaxel. Together, our study highlights an anti-TNBC mechanism of ROR-NOB via suppression of NF-κB signaling, suggesting novel preventive and chemotherapeutic strategies against this devastating disease.

The first-line cluster headache medication verapamil alters the circadian period and elicits sex-specific sleep changes in mice.

Verapamil is the first-line preventive medication for cluster headache, an excruciating disorder with strong circadian features. Whereas second- and third-line preventives include known circadian modulators, such as melatonin, corticosteroids, and lithium, the circadian effects of verapamil are poorly understood. Here, we characterize the circadian features of verapamil using both in vitro and in vivo models. In Per2::LucSV reporter fibroblasts, treatment with verapamil (0.03-10 µM) showed a dose-dependent period shortening of the reporter rhythm which reached a nadir at 1 µM, and altered core clock gene expression at 10 µM. Mouse wheel-running activity with verapamil (1 mg/mL added to the drinking water) also resulted in significant period shortening and activity reduction in both male and female free-running wild-type C57BL6/J mice. The temporal patterns of activity reduction, however, differ between the two sexes. Importantly, piezo sleep recording revealed sexual dimorphism in the effects of verapamil on sleep timing and bout duration, with more pronounced adverse effects in female mice. We also found altered circadian clock gene expression in the cerebellum, hypothalamus, and trigeminal ganglion of verapamil-treated mice. Verapamil did not affect reporter rhythms in ex vivo suprachiasmatic nucleus (SCN) slices from Per2:Luc reporter mice, perhaps due to the exceptionally tight coupling in the SCN. Thus, verapamil affects both peripheral (trigeminal ganglion) and central (hypothalamus and cerebellum) nervous system structures involved in cluster headache pathophysiology, possibly with network effects instead of isolated SCN effects. These studies suggest that verapamil is a circadian modulator in laboratory models at both molecular and behavioral levels, and sex is an important biological variable for cluster headache medications. These observations highlight the circadian system as a potential convergent target for cluster headache medications with different primary mechanisms of action.

Pilot study for in vivo cellular imaging of the muscularis propria and ex vivo molecular imaging of myenteric neurons (with video).

BACKGROUND: It is challenging to optimally sample the muscularis propria endoscopically for the diagnosis of muscle layer diseases, especially for motility disorders resulting from neuroenteric dysfunction. OBJECTIVES: Ultramagnification in vivo imaging of the muscularis mucosa and ex vivo identification of myenteric neuronal elements by confocal microscopy. DESIGN: Ex vivo and in vivo porcine animal studies. SETTING: Short-term study in an animal laboratory. INTERVENTIONS: The muscularis propria in the stomach and esophagus was accessed by resecting the mucosal layer with endoscopic submucosal dissection or cap EMR techniques or by creating a submucosal space by the submucosal endoscopy with mucosal flap technique. The muscularis propria was stained with Nissl stains and 2 types of neuronal molecular stains. The muscular layer was imaged with the endocytoscope in vivo. The muscularis stained with molecular-based stains was also evaluated with a confocal microscope. RESULTS: Cellular microstructures resembling spindle-shaped smooth muscle cells were visualized by endocytoscopy in vivo. Confocal endoscopic microscopy demonstrated that in vivo topical application of neuronal molecular stains successfully stained the muscularis and specifically highlighted neuron-like cells. LIMITATION: Animal model pilot study. CONCLUSIONS: In vivo endoscopic histologic evaluation of the muscularis propria is technically feasible and easy. Minimally invasive advanced endoscopic imaging may be useful for the diagnosis and study of neuroenteric disorders at the level of the muscularis propria, avoiding surgical full-thickness tissue sampling.

Comparative morphology of the lingual papillae and their connective tissue cores in the tongue of the Abyssinian black-and-white colobus (Colobus guereza).

We observed the morphology of the lingual papillae (filiform, fungiform, foliate, and vallate) and their underlying connective tissue cores (CTCs) in Abyssinian black-and-white colobus monkeys using light and scanning electron microscopy. The tongues of both juvenile and senescent individuals were relatively short in the rostro-caudal direction, with a rounded apex. Lingual tori were absent. Numerous filiform papillae were distributed over the entire tongue, except at the lingual root. A pair of foliate papillae was present on both the lateral and caudal margins of the corpus. Three vallate papillae were distributed on the boundary between the caudal part of the body and the root in both juvenile and senescent individuals. Based on scanning electron microscopy observations, the morphologies of the filiform papillae differed between juvenile and senescent individuals. The epithelial surface of juvenile filiform papillae had a main process, but the associated processes were weak and the underlying CTCs displayed immature morphology. In contrast, the epithelial surface of senescent filiform papillae was associated with several accessory processes, and their underlying CTCs consisted of several auxiliary cores that nearly encircled the main core, forming a concavity in the papilla. CTCs of the filiform papillae showed variable morphology. Juvenile filiform CTCs exhibited a rather primitive morphology, resembling those of the hamster, mole, and Cape hyrax while, conversely, despite the basically folivorous diet of the Abyssinian black-and-white colobus, senescent filiform CTCs resembled those found in omnivorous primates, including members of the Callitrichinae and Homoidea, and also those in Carnivora (e.g., Canidae and Felidae).

Discovery of a Potent, Selective, and Orally Available MTHFD2 Inhibitor (DS18561882) with in Vivo Antitumor Activity.

We report the discovery of a potent and isozyme-selective MTHFD2 inhibitor, DS18561882 (2). Through investigation of the substituents on our tricyclic coumarin scaffold (1,2,3,4-tetrahydrochromeno[3,4-c]pyridin-5-one), MTHFD2 inhibitory activity was shown to be elevated by incorporating an amine moiety at the 8-position and a methyl group at the 7-position of the initial lead 1. X-ray structure analysis revealed that a key interaction for enhanced potency was salt bridge formation between the amine moiety and the diphosphate linker of an NAD+ cofactor. Furthermore, ortho-substituted sulfonamide in place of benzoic acid of 1 significantly improved cell permeability and cell-based growth inhibition against a human breast cancer cell line. The thus-optimized DS18561882 showed the strongest cell-based activity (GI50 = 140 nM) in the class, a good oral pharmacokinetic profile, and thereby tumor growth inhibition in a mouse xenograft model upon oral administration.

Extensive expansion and diversification of the chemokine gene family in zebrafish: identification of a novel chemokine subfamily CX.

BACKGROUND: The chemokine family plays important roles in cell migration and activation. In humans, at least 44 members are known. Based on the arrangement of the four conserved cysteine residues, chemokines are now classified into four subfamilies, CXC, CC, XC and CX3C. Given that zebrafish is an important experimental model and teleost fishes constitute an evolutionarily diverse group that forms half the vertebrate species, it would be useful to compare the zebrafish chemokine system with those of mammals. Prior to this study, however, only incomplete lists of the zebrafish chemokine genes were reported. RESULTS: We systematically searched chemokine genes in the zebrafish genome and EST databases, and identified more than 100 chemokine genes. These genes were CXC, CC and XC subfamily members, while no CX3C gene was identified. We also searched chemokine genes in pufferfish fugu and Tetraodon, and found only 18 chemokine genes in each species. The majority of the identified chemokine genes are unique to zebrafish or teleost fishes. However, several groups of chemokines are moderately similar to human chemokines, and some chemokines are orthologous to human homeostatic chemokines CXCL12 and CXCL14. Zebrafish also possesses a novel species-specific subfamily consisting of five members, which we term the CX subfamily. The CX chemokines lack one of the two N-terminus conserved cysteine residues but retain the third and the fourth ones. (Note that the XC subfamily only retains the second and fourth of the signature cysteines residues.) Phylogenetic analysis and genome organization of the chemokine genes showed that successive tandem duplication events generated the CX genes from the CC subfamily. Recombinant CXL-chr24a, one of the CX subfamily members on chromosome 24, showed marked chemotactic activity for carp leukocytes. The mRNA was expressed mainly during a certain period of the embryogenesis, suggesting its role in the zebrafish development. CONCLUSION: The phylogenic and genomic organization analyses suggest that a substantial number of chemokine genes in zebrafish were generated by zebrafish-specific tandem duplication events. During such duplications, a novel chemokine subfamily termed CX was generated in zebrafish. Only two human chemokines CXCL12 and CXCL14 have the orthologous chemokines in zebrafish. The diversification observed in the numbers and sequences of chemokines in the fish may reflect the adaptation of the individual species to their respective biological environment.

Gender and Smoking Correlations of Surfactant Lipids and Proteins in the Saliva of Dental Patients.

We sought to determine the effects of smoking on surfactant lipids and proteins in saliva. Levels of sphingomyelin (Sph) phosphatidylcholine (PC) and lyso-PC (LPC) were determined by thin layer chromatography. Levels of surfactant protein A (SP-A) were determined by western analysis using antibodies specific for SP-A. Significance of the results was determined by the student's t-test. The LPC/PC ratio had a tendency to be much higher in smokers compared to nonsmokers. LPC levels were significantly higher in females smokers compared to male smokers. Additionally, levels of SP-A were significantly reduced in females smokers compared to non-smokers. Smoking alters surfactant protein and LPC/PC ratios in saliva. There is a significant difference in the effects in females compared to males. Findings suggest smoking alters the composition of saliva that may reduce protection of the oral cavity, which may explain why women smokers are at greater risk of developing oral mucositis.

Identification of a novel CXCL1-like chemokine gene in macaques and its inactivation in hominids.

Chemokines are a rapidly evolving cytokine gene family. Because of various genome rearrangements after divergence of primates and rodents, humans and mice have different sets of chemokine genes, with humans having members outnumbering those of mice. Here, we report the occurrence of lineage-specific chemokine gene generation or inactivation events within primates. By using human chemokine sequences as queries, we isolated a novel cynomolgus macaque CXC chemokine cDNA. The encoded chemokine, termed CXCL1L (from CXCL1-like) showed the highest similarity to human CXCL1. A highly homologous gene was also found in the rhesus macaque genome. By comparing the genome organization of the major CXC chemokine clusters among the primates, we found that one copy of the duplicated CXCL1 genes turned into a pseudogene in the hominids, whereas the gene in macaques has been maintained as a functionally active CXCL1L. In addition, cynomolgus macaque was found to contain an additional CXC chemokine highly homologous to CXCL3, termed CXCL3L (from CXCL3-like). These results demonstrate the birth-and-death process of a new gene in association with gene duplication within the primates.

The nucleoside and nucleotide mixture (OG-VI) rescues intestinal-like epithelial cells from the cytotoxicity of chemotherapeutic agents.

Immune cells and cells undergoing rapid turn-over can obtain exogenous nucleotides via salvage synthesis. We evaluated whether or not the balanced nucleoside and nucleotide mixture OG-VI, could rescue intestinal epithelial-like Caco-2 cells from the cytotoxic effects of several chemotherapeutic agents, in the presence and absence of glutamine (Gln). Cells were exposed to 5-fluorouracil (5FU), methotrexate (MTX) or 6-mercaptopurine (6MP), after which proliferation and cell cycle analyses were performed. Following exposure to the chemotherapeutic agents, we observed that cells treated with OG-VI proliferated well, whereas those without the supplement did not proliferate. Furthermore, following treatment with either 5FU or MTX, we observed that the number of cells in the G0/G1 phase decreased and those in the S phases increased. However, these cell cycle alterations were prevented by the addition of OG-VI. With the exception of 6MP-treated cells, we did not observe any effects on proliferation or cell cycle regulation that could be ascribed to the presence of Gln. Thus, we have demonstrated that OG-VI rescues cells from the cytotoxic effects of several chemotherapeutic agents.

Hematological and pathological features of massive hepatic necrosis in two radiated tortoises (Astrochelys radiata).

Two radiated tortoises (Astrochelys radiata) exhibited anorexia and hypokinesia. In both cases, hematological and serum biochemical examinations revealed high alkaline phosphatase levels, moderately high aspartate aminotransferase levels and white blood cell counts approximately within the normal range. Despite being treated, the tortoises died 9 and 43 days after the first clinical examination. Gross pathological examinations revealed that the livers of both animals were extremely swollen and contained pale yellow necrotic tissue. Histopathological assessment revealed that the livers contained a massive area of hepatic necrosis surrounded by migration of macrophages and multinucleated giant cells. In one of the cases, severe fibrosis was observed. The present study provides reference information for similar cases in the future.

Contributions and source identification of biogenic and anthropogenic hydrocarbons to secondary organic aerosols at Mt. Tai in 2014.

Ambient fine particulate matter (PM2.5) and volatile organic compounds (VOCs) collected at Mt. Tai in summer 2014 were analysed and the data were used to identify the contribution of biogenic and anthropogenic hydrocarbons to secondary organic aerosols (SOA) and their sources and potential source areas in high mountain regions. Compared with those in 2006, the 2014 anthropogenic SOA tracers in PM2.5 aerosols and VOC species related to vehicular emissions exhibited higher concentrations, whereas the levels of biogenic SOA tracers were lower, possibly due to decreased biomass burning. Using the SOA tracer and parameterisation method, we estimated the contributions from biogenic and anthropogenic VOCs, respectively. The results showed that the average concentration of biogenic SOA was 1.08 ± 0.51 µg m-3, among which isoprene SOA tracers were dominant. The anthropogenic VOC-derived SOA were 7.03 ± 1.21 µg m-3 and 1.92 ± 1.34 µg m-3 under low- and high-NOx conditions, respectively, and aromatics made the greatest contribution. However, the sum of biogenic and anthropogenic SOA only contributed 18.1-49.1% of the total SOA. Source apportionment by positive matrix factorisation (PMF) revealed that secondary oxidation and biomass burning were the major sources of biogenic SOA tracers. Anthropogenic aromatics mainly came from solvent use, fuel and plastics combustion and vehicular emissions. However, for > C6 alkanes and cycloalkanes, vehicular emissions and fuel and plastics combustion were the most important contributors. The potential source contribution function (PSCF) identified the Bohai Sea Region (BSR) as the major source area for organic aerosol compounds and VOC species at Mt. Tai.

Protein deficiency impairs DNA vaccine-induced antigen-specific T cell but not B cell response in C57BL/6 mice.

DNA vaccination is a simple method to induce antigen (Ag)-specific immunoresponse and has many potential advantages over other vaccines. Although people who need to receive vaccines often suffer undernutrition, there has been no study on a how nutritional status affects the immunoresponse induced by DNA vaccination. The aim of this study was to determine the relationship between protein deficiency and DNA vaccine-induced immunoresponses. C57BL/6 mice were fed a 5% or 20% casein diet for 30 d. The mice were immunized with an ovalubumin (OVA)-expression plasmid by the gene gun-based method three times at 10-d intervals. Body weight and serum albumin concentration in protein-deficient mice were significantly lower than those in mice fed the 20% casein diet (p<0.01, p<0.05). The percentage of OVA-specific CD8+ T cells was significantly decreased in the 5% casein group compared to that in the 20% casein group (p<0.05). Furthermore, CD4+ T cells from mice fed the low-protein diet showed lower interleukin (IL)-2 production than did those from the 20% group. In contrast to the T-cell function, protein deficiency did not affect OVA-specific Ab responses (p>0.05). These results suggest that protein deficiency impairs the induction of Ag-specific T-cell but not B-cell response in DNA-immunized mice. Our observation indicates that, in addition to development of an effective of DNA vaccine, the management of nutritional state is important for the prevention of infectious disease by DNA vaccination.