Bacterial DNA and serum IgG antibody titer assays for assessing infection of human-pathogenic and dog-pathogenic Porphyromonas species in dogs.

Periodontal disease is highly prevalent in both humans and dogs. Although there have been reports of cross-infection of periodontopathic bacteria, methods for assessing it have yet to be established. The actual status of cross-infection remains to be seen. The purpose of this study was to evaluate the utility of bacterial DNA and serum immunoglobulin G (IgG) antibody titer assays to assess infection of human-pathogenic and dog-pathogenic Porphyromonas species in dogs. Four experimental beagles were used for establishing methods. Sixty-six companion dogs at veterinary clinics visiting for treatment and prophylaxis of periodontal disease were used and divided into healthy, gingivitis, and periodontitis groups. Periodontal pathogens such as Porphyromonas gingivalis and Porphyromonas gulae were investigated as target bacteria. DNA levels of both bacteria were measured using species-specific primers designed for real-time polymerase chain reaction (PCR). Serum IgG titers of both bacteria were measured by enzyme-linked immunosorbent assay (ELISA). PCR primers were confirmed to have high sensitivity and specificity. However, there was no relationship between the amount of bacterial DNA and the severity of the periodontal disease. In addition, dogs with periodontitis had higher IgG titers against both bacteria compared to dogs in the healthy and gingivitis groups; there was cross-reactivity between the two bacteria. Receiver operating characteristic (ROC) analysis of IgG titers against both bacteria showed high sensitivity (>90 %) and specificity (>75 %). Since both bacteria were distinguished by DNA assays, the combination of these assays may be useful in the evaluation of cross-infection.

A novel type IIb L-asparaginase from Latilactobacillus sakei LK-145: characterization and application.

We succeeded in homogeneously expressing and purifying L-asparaginase from Latilactobacillus sakei LK-145 (Ls-Asn1) and its mutated enzymes C196S, C264S, C290S, C196S/C264S, C196S/C290S, C264S/C290S, and C196S/C264S/C290S-Ls-Asn1. Enzymological studies using purified enzymes revealed that all cysteine residues of Ls-Asn1 were found to affect the catalytic activity of Ls-Asn1 to varying degrees. The mutation of Cys196 did not affect the specific activity, but the mutation of Cys264, even a single mutation, significantly decreased the specific activity. Furthermore, C264S/C290S- and C196S/C264S/C290S-Ls-Asn1 almost completely lost their activity, suggesting that C290 cooperates with C264 to influence the catalytic activity of Ls-Asn1. The detailed enzymatic properties of three single-mutated enzymes (C196S, C264S, and C290S-Ls-Asn1) were investigated for comparison with Ls-Asn1. We found that only C196S-Ls-Asn1 has almost the same enzymatic properties as that of Ls-Asn1 except for its increased stability for thermal, pH, and the metals NaCl, KCl, CaCl2, and FeCl2. We measured the growth inhibitory effect of Ls-Asn1 and C196S-Ls-Asn1 on Jurkat cells, a human T-cell acute lymphoblastic leukemia cell line, using L-asparaginase from Escherichia coli K-12 as a reference. Only C196S-Ls-Asn1 effectively and selectively inhibited the growth of Jurkat T-cell leukemia, which suggested that it exhibited antileukemic activity. Furthermore, based on alignment, phylogenetic tree analysis, and structural modeling, we also proposed that Ls-Asn1 is a so-called "Type IIb" novel type of asparaginase that is distinct from previously reported type I or type II asparaginases. Based on the above results, Ls-Asn1 is expected to be useful as a new leukemia therapeutic agent.

A case of ovarian carcinosarcoma with a germline pathogenic variant of BRCA2 involving a perforated appendix with an abscess.

We report a case of rare and aggressive ovarian carcinosarcoma with a germline pathogenic BRCA2 variant. A patient with a history of breast cancer who developed an inflammatory ovarian tumor with peritonitis carcinomatosis involving the appendix suffered from cachexia. Following three cycles of weekly paclitaxel and carboplatin chemotherapy, emergency surgery was required owing to sepsis. Bilateral salpingo-oophorectomy, total hysterectomy, appendectomy, and small intestine adhesiolysis were performed. Histologically, the tumor comprised an admixture of carcinomatous and sarcomatous components, with involvement of the appendix, which had caused perforation and abscess formation. The final diagnosis was ovarian carcinosarcoma with a germline pathogenic BRCA2 variant, c.658_659del (p.Val220fs). The patient responded completely to adjuvant chemotherapy. A combination of chemotherapy and surgery might be beneficial to patients with ovarian carcinosarcoma and germline pathogenic BRCA2 variants with a poor general condition. This is the first report of ovarian carcinosarcoma with a germline pathogenic BRCA2 variant that responded favorably to chemotherapy.

Relationship between Perceived Physical Competence and Outdoor Play among Children Aged 9-12 Years-Focused Sex-Specific Differences.

BACKGROUND: Outdoor play (OP), which is considered important for children's development, is declining every year. Perceived physical competence (PPC) is a vital factor that promotes physical activity such as OP, sports clubs, etc., but the relationship between PPC and OP was unknown. The purpose of this research was to investigate the relationship between PPC and OP in children and consider whether there were any sex-specific changes. METHODS: A cross-sectional study was conducted in Japan with 288 children (134 girls, age: 10.6 ± 1.01 years). OP was assessed using an original self-report questionnaire. Each weekday, the children reported the time of OP and were classified as "high" if they played outside for at least an hour three times. PPC was evaluated with a self-report questionnaire developed by Okazawa et al. (1996). It has 12 questions and was assessed on a 5-point Likert scale. After adjusting for age, sex, BMI, screen time, sports club participation, and the number of friends, logistic regression analyses were carried out. RESULTS: Children with better PPC were significantly more likely to be classified as "high" [crude odds ratio (OR): 1.04; 95% confidence interval (CI): 1.00-1.08; adjusted OR: 1.04; 95% CI: 1.00-1.08]. Only girls with better PPC were significantly more likely to be classified as "high" in a sex-based stratified analysis [crude OR: 1.08; 95% CI: 1.01-1.15, adjusted OR 1.09; 95% CI: 1.02-1.17]. CONCLUSIONS: Particularly among girls, OP could be promoted as a voluntary physical activity with improved PPC.

TRPA1-mediated repellency behavior in the red flour beetle Tribolium castaneum.

The sensory perception of irritant chemicals results in escape and repellency behavioral patterns in insects. Transient receptor potential channels are cation channels that function as sensor proteins for several types of signals, such as light, sound, temperature, taste, as well as chemical and physical stimuli; among these, the TRPA channel is widely conserved and activated by irritant chemicals. Certain plant-derived essential oils (EOs), produced by secondary metabolism, are mixtures of volatile compounds, which are used as repellents because they contain environmentally sustainable ingredients. Citronellal, which is present in citronella EO from Cymbopogon species, is a potentially viable insect repellent; however, the repellency capability against coleopteran beetles remains limited. We investigated the citronellal-derived repellency behavior for the red flour beetle Tribolium castaneum, in which TcTRPA1 and odorant receptor co-receptor (Orco) expressions were mediated by RNA interference. Area-preference tests showed dose-dependent repellency behavior for citronellal; additionally, both TcTRPA1 and TcOrco double-strand RNA (dsRNA) micro-injection showed clear TcTRPA1 and TcOrco transcript reductions, and only TcTRPA1 dsRNA treatment significantly impaired repellency behavior. The relative expression level of the TcTRPA1 transcripts, evaluated by quantitative reverse-transcription PCR (qRT-PCR), revealed dominant expression in the antennae, indicating the antennae-expressed TcTRPA1-mediated repellency behavior.

Association Between Workplace Social Capital and Neck Pain: A Cross-Sectional Study Among Workers.

OBJECTIVE: We examined the association between workplace social capital (WSC) and presence of neck pain (NP) among workers. METHODS: This cross-sectional study included 595 workers. Presence of NP was assessed using Numerical Rating scale. WSC (overall, bonding, bridging, and linking) was divided into three categories (low, middle, and high) based on tertile distributions. We used logistic regression analysis to investigate the association between WSC and presence of NP. RESULTS: Low overall, bonding, and linking social capital were significantly associated with presence of NP in comparison with each high social capital (overall: adjusted odds ratio [aOR] = 1.76, 95% confidence interval [CI] = 1.14 to 2.73; bonding: aOR = 1.78, 95% CI = 1.19 to 2.67, linking: aOR = 2.18, 95% CI = 1.32 to 3.63). CONCLUSION: Lower WSC had an association with higher prevalence of NP among workers.

Estimation of periodontal pocket surface area in small to medium dogs: a proof-of-concept study.

BACKGROUND: Periodontal disease is the most common dental disease in dogs. Although the systemic effects of periodontal disease have not been clarified in veterinary science, it is necessary to evaluate the effects of periodontal disease in clinical trials in the future. There have been a few clinical attempts made, however, to assess the severity of periodontal inflammation and its impact on the systemic health of dogs. Meanwhile, in the field of dentistry for humans, the periodontal inflamed surface area (PISA) and periodontal epithelial surface area (PESA) have been used to quantitatively assess the degree of periodontal disease affecting a single tooth as well as the overall extent of periodontitis. Recent studies have also suggested the use of these assessments to examine the relationship between periodontal inflammation and systemic health. RESULTS: The estimation formula for a dog's periodontal pocket surface area (PPSA), an alternative to PISA and PESA in humans, was established using body weight and periodontal pocket depth. Actual values were measured using extracted teeth from various dog breeds and sizes (2.3-25.0 kg of body weight) to obtain universal regression equations for PPSA. Altogether, 625 teeth from 73 dogs of 16 breeds were extracted and subsequently analyzed for morphological information. PPSA was measured in 61 dogs of 10 breeds with periodontal disease using the established estimation formulas, and the correlation between PPSA and preoperative blood chemistry data was analyzed accordingly. A strong correlation was found between PPSA and serum globulin (r = 0.71) while moderate correlations were found for C-reactive protein (r = 0.54) and serum albumin (r = -0.51). CONCLUSIONS: Estimation formulas using body weight and the 6-point probing depth were established for determining PPSA. Direct correlations between PPSA and several blood test results were observed in the study sample. Taken together, these results suggest that PPSA could be useful for evaluating the effects of periodontitis on systemic conditions in dogs.

Evaluation of Internal Distribution and Extracellular Action of the Cell via TOF-SIMS.

Cisplatin is one of the most popular and traditional platinum-based anti-cancer drugs. Additionally, it is known for its effect on different types of cancers. To clarify the reaction mechanism of anti-cancer drugs in a cell, the visualization of drugs in a single cell is required. In this study, we investigated a secondary ion species obtained from cisplatin, which was bounded to the nucleus in a cell and its intensity. PtCl2- was mainly detected via SIMS during an analysis of pure cisplatin reagent. In contrast, a high-intensity signal for PtCN- was detected from cultured cells that were administered cisplatin. However, this signal was not detected from cisplatin in the reagent state. Chlorine in the cisplatin structure is replaced with water when it is combined with the cell nucleus. Therefore, PtCN- was mainly detected from the intracellular region because the structure was changed by cisplatin binding to the nucleus and which exhibits anti-cancer activity. The results showed that the cisplatin selectively combined with the nucleus. Through TOF-SIMS, we achieved a visual distribution of the cisplatin intracellular nucleus.

Novel Methods to Generate Active Ingredients-Enriched Ashwagandha Leaves and Extracts.

Ashwagandha (Withania somnifera) is an Ayurvedic herb commonly used in world-renowned traditional Indian home medicine system. Roots of Ashwagandha have been traditionally known to possess a variety of therapeutic and health promoting potentials that have not been sufficiently supported by laboratory studies. Nevertheless, most, if not all, of the preventive and therapeutic potentials have been assigned to its bioactive components, steroidal alkaloids and lactones. In contrast to the traditional use of roots, we have been exploring bioactivities in leaves of Ashwagandha. Here, we report that the leaves possess higher content of active Withanolides, Withaferin-A (Wi-A) and Withanone (Wi-N), as compared to the roots. We also established, for the first time, hydroponic cultivation of Ashwagandha and investigated the effect of various cultivation conditions on the content of Wi-A and Wi-N by chemical analysis and bioassays. We report that the Withanone/Withaferin A-rich leaves could be obtained by manipulating light condition during hydroponic cultivation. Furthermore, we recruited cyclodextrins to prepare extracts with desired ratio of Wi-N and Wi-A. Hydroponically grown Ashwagandha and its extracts with high ratio of withanolides are valuable for cancer treatment.

Cholesterol enrichment of rabbit platelets enhances the Ca(2+) entry pathway induced by platelet-derived secondary feedback agonists.

AIMS: Hypersensitivity of platelets due to increased platelet cholesterol levels has been reported in hypercholesterolemia. However, the signaling pathways linking increased platelet reactivity and cholesterol contents are not fully understood. This study aims to determine the direct effect of cholesterol enrichment of platelets on the pathways including Ca(2+) mobilization and secondary feedback agonists such as adenosine diphosphate (ADP) and thromboxane A2 (TXA2). MAIN METHODS: In vitro cholesterol enrichment of rabbit platelets was performed by incubation with cholesterol complexed with methyl-ß-cyclodextrin. Ca(2+) mobilization was monitored using platelets loaded with fura-PE3/AM, a fluorescent calcium indicator. Released ATP and TXB2 from platelets were measured by a luciferin-luciferase ATP assay system and a TXB2 ELISA Kit, respectively. KEY FINDINGS: Cholesterol enrichment of rabbit platelets significantly enhanced Ca(2+) mobilization induced by thrombin, accompanying an augmented Ca(2+) entry. The augmentation of Ca(2+) entry by cholesterol enrichment was significantly suppressed by treatment with inhibitors for secondary feedback agonists. In cholesterol-enriched platelets, the amount of released ATP or TXB2 induced by thrombin was not significantly altered in comparison with control platelets, whereas an increase in [Ca(2+)]i induced by ADP or U46619, a TXA2 mimetic, was significantly enhanced. SIGNIFICANCE: These results suggest that cholesterol enrichment of rabbit platelets results in enhanced Ca(2+) mobilization via ADP/TXA2-dependent augmentation of the Ca(2+) entry pathway. The results reveal a novel mechanism by which platelet hypersensitivity is regulated by cholesterol contents.

Identification of two phytotoxins, blumenol A and grasshopper ketone, in the allelopathic Japanese rice variety Awaakamai.

Aqueous methanol extracts of the traditional rice (Oryza sativa) variety Awaakamai, which is known to have the greatest allelopathic activity among Japanese traditional rice varieties, inhibited the growth of roots and shoots of cress (Lepidium sativum), lettuce (Lactuca sativa), timothy (Phleum pratense), Digitaria sanguinalis, Lolium multiflorum and Echinochloa crus-galli. Increasing the extract concentration increased the inhibition, suggesting that the extract of Awaakamai contains growth inhibitory substances. The extract of Awaakamai was purified and two main growth inhibitory substances were isolated and determined by spectral data as blumenol A and grasshopper ketone. Blumenol A and grasshopper ketone, respectively, inhibited the growth of cress shoots and roots at concentrations greater than 10 and 30 µmol/L. The concentrations required for 50% growth inhibition on cress roots and shoots were 84 and 27 µmol/L, respectively, for blumenol A, and 185 and 76 µmol/L, respectively, for grasshopper ketone. These results suggest that blumenol A and grasshopper ketone may contribute to the growth inhibitory effect of Awaakamai and may play an important role in the allelopathy of Awaakamai.