Genetic analysis of impaired healing responses after periodontal therapy in type 2 diabetes: Clinical and in vivo studies.

OBJECTIVE: This study aims to investigate the mechanisms underlying the impaired healing response by diabetes after periodontal therapy. BACKGROUND: Outcomes of periodontal therapy in patients with diabetes are impaired compared with those in patients without diabetes. However, the mechanisms underlying impaired healing response to periodontal therapy have not been sufficiently investigated. MATERIALS AND METHODS: Zucker diabetic fatty (ZDF) and lean (ZL) rats underwent experimental periodontitis by ligating the mandibular molars for one week. The gingiva at the ligated sites was harvested one day after ligature removal, and gene expression was comprehensively analyzed using RNA-Seq. In patients with and without type 2 diabetes (T2D), the corresponding gene expression was quantified in the gingiva of the shallow sulcus and residual periodontal pocket after non-surgical periodontal therapy. RESULTS: Ligation-induced bone resorption and its recovery after ligature removal were significantly impaired in the ZDF group than in the ZL group. The RNA-Seq analysis revealed 252 differentially expressed genes. Pathway analysis demonstrated the enrichment of downregulated genes involved in the peroxisome proliferator-activated receptor (PPAR) signaling pathway. PPARα and PPARγ were decreased in mRNA level and immunohistochemistry in the ZDF group than in the ZL group. In clinical, probing depth reduction was significantly less in the T2D group than control. Significantly downregulated expression of PPARα and PPARγ were detected in the residual periodontal pocket of the T2D group compared with those of the control group, but not in the shallow sulcus between the groups. CONCLUSIONS: Downregulated PPAR subtypes expression may involve the impaired healing of periodontal tissues by diabetes.

Improvement of periodontal parameters following intensive diabetes care and supragingival dental prophylaxis in patients with type 2 diabetes: A prospective cohort study.

AIM: This study aimed to investigate the effects of diabetes care on periodontal inflammation. MATERIALS AND METHODS: This prospective cohort study included 51 Japanese patients with type 2 diabetes who underwent intensive diabetes care including educational hospitalization and regular outpatient treatment for 6 months. Dental prophylaxis without subgingival scaling was provided three times during the observational period. Associations between changes in periodontal parameters and glycaemic control levels were evaluated using multiple regression analysis. RESULTS: Overall, 33 participants (mean age: 58.7 ± 12.9) were followed up for 6 months. At baseline examination, 82% were diagnosed with Stage III or IV periodontitis. Haemoglobin A1c (HbA1c) level changed from 9.6 ± 1.8% at baseline to 7.4 ± 1.3% at 6 months. The ratio of probing pocket depth (PPD) ≥4 mm, bleeding on probing (BOP), full-mouth plaque control record (PCR), periodontal epithelial surface area (PESA) and periodontal inflamed surface area (PISA) also significantly improved. The reduction in PPD and PESA was significantly associated with changes in both HbA1c and fasting plasma glucose (FPG) levels, and the reduction in PISA was significantly associated with an improvement in FPG after adjusting for smoking, change in body mass index and full-mouth PCR. CONCLUSIONS: This is the first study to report a significant improvement in PPD and BOP after intensive diabetes care and dental prophylaxis without subgingival scaling. CLINICAL TRIAL REGISTRATION NUMBER: UMIN000040218.

Contribution of traditional Japanese Kampo medicines, kakkonto with shosaikotokakikyosekko, in treating patients with mild-to-moderate coronavirus disease 2019: Further analysis of a multicenter, randomized controlled trial.

We previously reported the finding of symptom relief in a randomized controlled trial with the combined use of kakkonto and shosaikotokakikyosekko added to conventional treatment in patients with coronavirus disease 2019 (COVID-19). For further evaluation, we performed post hoc analysis focused on symptom disappearance without recurrence, to determine a clearer effect of Kampo medicine. Patients with mild and moderate COVID-19 were randomly allocated to a control group receiving symptomatic therapy or a Kampo group receiving kakkonto (2.5 g) with shosaikotokakikyosekko (2.5 g) three times daily in addition to symptomatic therapy. The data of 161 patients (Kampo group, n = 81; control group, n = 80) were analyzed post hoc for the time to symptom disappearance. Kaplan-Meier and Cox proportional hazard estimates of disappearance of symptoms showed that all and each symptom targeted in this study disappeared faster in the Kampo group than in the control group, although not statistically significant (all symptomatic cases; hazard ratio [HR] 3.73, 95% confidence interval [CI] 0.46-29.98, log-rank p = 0.1763). In a supplemental assessment using covariate adjustment and competing risk analysis, fever disappeared faster in the Kampo group than in the control group (all symptomatic cases, HR 1.62, 95% CI 0.99-2.64, p = 0.0557; unvaccinated cases, HR 1.68, 95% CI 1.00-2.83, p = 0.0498) and shortness of breath disappeared significantly faster in Kampo group than in control group (all symptomatic cases, HR 1.92, 95% CI 1.07-3.42, p = 0.0278; unvaccinated cases, HR 2.15, 95% CI 1.17-3.96, p = 0.0141). These results demonstrate the advantages of Kampo treatment for acute COVID-19.

Predictors for the Development of Hypoxia or Prolonged Acute Symptoms among Non-Hospitalized Mild-to-Moderate Patients with Coronavirus Disease 2019.

The coronavirus disease 2019 (COVID-19) pandemic remains a global public health concern. The clinical course and risk of developing severe illness among patients with COVID-19 who are at low-risk of severe COVID-19 remain uncertain. This retrospective cohort study from an isolation facility for low-risk COVID-19 patients in Japan evaluated the potential risks for severe disease with hypoxia (SpO2 ≤ 93%) or experiencing prolonged isolation period longer than 14 days with persistent acute symptoms. The study was performed before the spread of the alpha variant in the country and before the start of a nationwide mass vaccination campaign against COVID-19. Among the 929 participants with reliable outcome data regarding the development of hypoxia, 63 (6.8%) developed severe disease with hypoxia during their stays at the facility. Higher age [adjusted odds ratio (aOR), 1.08; 95% confidence interval (CI), 1.06-1.10] and male sex (aOR, 4.70; 95% CI, 2.39-9.22) were associated with this outcome. As for the experience of prolonged isolation period, higher age (aOR, 1.02; 95% CI, 1.01-1.04), atopic diseases (aOR, 1.69, 95% CI, 1.09-2.64), presence of cough at onset (aOR, 1.64; 95% CI, 1.09-2.48), and prescription of oral antibiotics before positive test results for COVID-19 (aOR, 2.37; 95% CI, 1.33-4.22) were associated with this outcome. In summary, 5-10% of low-risk COVID-19 patients later develop hypoxia. Older age and male sex were associated with both the development of hypoxia and prolonged acute symptoms. The unnecessary prescription of antibiotics before COVID-19 diagnosis may prolong COVID-19 symptoms.

Income-related inequalities in the association of smoking with periodontitis: a cross-sectional analysis in Tokyo Metropolitan Districts.

OBJECTIVE: Socio-economic status (SES) and smoking are risk factors for periodontitis; however, their interaction has not been determined. We investigated the effect of modification of SES and smoking with periodontal conditions. MATERIALS AND METHODS: Data on the social background, smoking status, and dental examination of 1033 individuals residing in the Tokyo Metropolitan District were analyzed. The outcomes were the number of remaining teeth and the proportion of teeth with probing pocket depth (PPD) ≥ 4 mm and ≥ 6 mm. Multilevel linear and Poisson regression analyses were performed after adjusting for possible confounding factors, including SES, assessed by the average income of the residential area. RESULTS: The mean number of remaining teeth was 24.6 ± 4.8, and the proportion of teeth with PPD ≥ 4 mm and ≥ 6 mm was 31.2 ± 28.5% and 12.2 ± 18.1%, respectively. After adjusting for confounding factors, the lowest-income population had significantly lesser teeth (coefficient: - 0.46, 95% CI - 0.89, 0.02, p = 0.039) and a higher proportion of teeth with PPD ≥ 4 mm than the highest-income population (ratio of means: 1.22, 95% CI 1.03-1.44, p = 0.013). Significant interactions were observed; income inequalities in periodontitis were significant only among current smokers. CONCLUSION: Inequality in socio-economic status is associated with oral health inequalities. The adverse effects of smoking on periodontitis might be greater in the low-income population. CLINICAL RELEVANCE: The low-income population, especially current smokers, had significantly more compromised oral health than the high-income population. In addition to the emphasis on smoking cessation, the promotion of universal health coverage for dental care is necessary to reduce oral health inequalities.

Impaired dental implant osseointegration in rat with streptozotocin-induced diabetes.

OBJECTIVE: Few studies have reported on the impact of oxidative stress on the dental implant failure. The aim of this study was to investigate the impact of hyperglycemia-induced oxidative stress on dental implant osseointegration in diabetes mellitus (DM). METHODS: Acid-treated titanium implants were bilaterally placed in the maxillary alveolar ridge of streptozotocin-induced diabetic (DM group) and control rats after extraction of first molars. Histological analysis and micro-push-out test were performed 4 weeks after surgery. Oxidative stress and osteogenic markers in the surrounding bone were quantified by real-time polymerase chain reaction. In the in vitro study, rat bone marrow-derived mesenchymal stem cells (BMMSCs) were cultured on acid-treated titanium discs in a high-glucose (HG) or normal environment. Intracellular reactive oxygen species (ROS), cell proliferation, alkaline phosphatase (ALP) activity, and extracellular calcification were evaluated following antioxidant treatment with N-acetyl-L-cysteine (NAC). RESULTS: The implant survival rate was 92.9% and 75.0% in control and DM group, respectively. Bone-implant contact and push-out loads were significantly lower in the DM group. Expression of superoxide dismutase 1 at the mRNA level and on immunohistochemistry was significantly lower in the DM group. In vitro experiments revealed that the HG condition significantly increased ROS expression and suppressed the proliferation and extracellular calcification of BMMSCs, while NAC treatment significantly restored ROS expression, cell proliferation, and calcification. The ALP activity of both groups was not significantly different. CONCLUSION: In diabetes, high-glucose-induced oxidative stress downregulates proliferation and calcification of BMMSCs, impairing osseointegration and leading to implant failure.

Refractory Chest Pain in Mild to Moderate Coronavirus Disease 2019 Successfully Treated with Saikanto, a Japanese Traditional Medicine.

Coronavirus disease 2019 (COVID-19) causes a variety of pain symptoms in the acute phase. Severe chest pain suddenly occurs even without abnormalities on examination and is sometimes refractory to analgesics. Such pain is a clinical concern in care facilities with limited resources, and this is the first report on the use of saikanto for its treatment. In Miyagi Prefecture, Japan, COVID-19 patients with mild symptoms were admitted to a hotel that operated as an isolation facility, and their symptoms were observed. In this article, we report four cases in which chest pain comorbid with mild to moderate COVID-19 was successfully treated with saikanto, a traditional Japanese (Kampo) medicine. The patients presented with chest pain and underwent medical examination at the facility. Two patients had severe chest pain refractory to acetaminophen. Critical cardiopulmonary diseases were ruled out in all the patients, and three patients had features of pneumonia on chest radiograph. Medications, including saikanto, were administered to the patients. The patients' chest pain and other symptoms improved 1-4 days after the administration of saikanto, and they left the care facility without hospitalization. The cause of the chest pain experienced by these patients is unclear, but we speculate that it could be minimal pleural inflammation or neuropathy. Previous pharmacological studies have suggested anti-inflammatory and analgesic properties of the crude drugs that constitute saikanto. This case report suggests that saikanto could be a treatment option for chest pain refractory to analgesics in patients with mild to moderate COVID-19.

Response to Glucocorticoid Therapy in Patients with Mild to Moderate Coronavirus Disease 2019 at a Japanese Care Facility.

The fifth wave of the coronavirus disease 2019 (COVID-19) pandemic caused by delta variant infection depleted medical resources, and the Japanese government announced glucocorticoid use for outpatients. An appropriate outpatient-glucocorticoid treatment for COVID-19 has not been established; therefore, we created treatment manuals with indications for glucocorticoid administration in a care facility adequately equipped to manage patients with mild to moderate COVID-19. Thirty-eight patients (24 males, 14 females; mean age 40.5 ± 11.8 years) were treated with glucocorticoids from August 1 to October 1, 2021 [COVID-19 staging, mild (n = 1), moderate I (n = 19), and moderate II (n = 18)]. Patients were treated with 6.6 mg/day d.i.v. or 6 mg/day p.o. dexamethasone, or 20-30 mg/day p.o. prednisolone. The median (25th-75th percentile) number of days from the date of onset to glucocorticoid administration was 8.0 days (7.0-11.25 days). While 24 patients were hospitalized, the condition of 14 improved without hospitalization. The median number of days from glucocorticoid administration to hospitalization was 1.0 day (range, 1.0-1.0 day). In the non-hospitalized patients, the median number of days of glucocorticoid administration was 5.0 days (5.0-5.25 days). The mean number of days from glucocorticoid administration to discharge from the care facility for non-hospitalized patients was 8.4 ± 3.3 days. The adverse reactions among non-hospitalized patients included insomnia (n = 1) and mild liver dysfunction (n = 3). The present method of glucocorticoid administration can be safely used for patients with COVID-19 in care facilities.

Impact of activation of neotrehalosadiamine/kanosamine biosynthetic pathway on the metabolism of Bacillus subtilis.

The pentose phosphate (PP) pathway is one of the major sources of cellular NADPH. A B. subtilis zwf mutant that lacks glucose-6-phosphate dehydrogenase (the enzyme that catalyzes the first step of the PP pathway) showed inoculum-dose-dependent growth. This growth defect was suppressed by glcP disruption, which causes the upregulation of an autoinducer neotrehalosadiamine (NTD)/kanosamine biosynthetic pathway. A metabolome analysis showed that the stimulation of NTD/kanosamine biosynthesis caused significant accumulation of TCA cycle intermediates and NADPH. Because the major malic enzyme YtsJ concomitantly generates NADPH through malate-to-pyruvate conversion, de novo NTD/kanosamine biosynthesis can result in an increase in the intracellular NADPH pool via the accumulation of malate. In fact, a zwf mutant grew in malate-supplemented medium. Artificial induction of glcP in the zwf mutant caused a reduction in the intracellular NADPH pool. Moreover, the correlation between the expression level of the NTD/kanosamine biosynthesis operon ntdABC and the intracellular NADPH pool was confirmed. Our results suggest that NTD/kanosamine has the potential to modulate the carbon-energy metabolism through an autoinduction mechanism.ImportanceAutoinducers enable bacteria to sense cell density and to coordinate collective behavior. NTD/kanosamine is an autoinducer produced by B. subtilis and several close relatives, although its physiological function remains unknown. The most important finding of this study was the significance of de novo NTD/kanosamine biosynthesis in the modulation of the central carbon metabolism in B. subtilis We showed that NTD/kanosamine biosynthesis caused an increase in the NADPH pool via the accumulation of TCA cycle intermediates. These results suggest a possible role for NTD/kanosamine in the carbon-energy metabolism. As Bacillus species are widely used for the industrial production of various useful enzymes and compounds, the NTD/kanosamine biosynthetic pathway might be utilized to control metabolic pathways in these industrial strains.

Two Cases of Multiple Chemical Sensitivity Successfully Treated With Kampo Medicine.

CONTEXT: Multiple chemical sensitivity (MCS) is a chronic disorder in which minimal but sustained exposure to particular chemicals causes headaches, vertigo, and multiple other symptoms. Up to 10% of cases are clinically unresponsive to treatment. Kampo medicines include multiple crude components with many applications for organ disorders. OBJECTIVE: The research team aimed to assess the efficacy of goreisan, a Japanese Kampo medicine, for 2 patients diagnosed with intractable cases of MCS. DESIGN: The research team conducted case studies with 2 female patients. SETTING: The study took place at the Kampo clinic in the Department of Gynecology and Obstetrics at the Japanese Red Cross Maebashi Hospital in Maebashi, Gunma, Japan. PARTICIPANTS: The participants were patients at the clinic. INTERVENTION: The first participant received the Kampo formula goreisan for her headaches and keigairengyoto for her mucosal discomfort; the second received goreisan for her headaches and kakkonto to reduce her shoulder-muscle stiffness. The participants received Kampo treatments for one month. OUTCOME MEASURES: The first participant's MCS symptoms were evaluated with the Quick Environment Exposure Sensitivity Inventory questionnaire (QEESI); the second using a numeric rating scale (NRS). RESULTS: After the Kampo treatments, the first participant's QEESI scores for chemical intolerance, symptom severity, and life impact were reduced from 47, 92, and 76 to 37, 39, and 55 points, respectively. The second participant's NRS scores were likewise reduced from 6, 8, and 8 out of 10 to 1, 2, and 1 out of 10. Symptoms were relieved in both patients, and didn't recur. CONCLUSIONS: These results highlight the potential benefits of Kampo medicine for the treatment of intractable MCS. Further investigations will be needed to confirm the mechanism of action, thereby improving the understanding of the effectiveness of Kampo medicine for MCS therapy.

Treatment of COVID-19-Related Olfactory Disorder Promoted by Kakkontokasenkyushin'i: A Case Series.

Olfactory disorders are one of the characteristic symptoms of the coronavirus disease of 2019 (COVID-19), which causes infection and inflammation of the upper and lower respiratory tract. To our knowledge, there are no treatments for COVID-19-related olfactory disorder. Here, we report five olfactory disorder cases in COVID-19, treated using the Japanese traditional (Kampo) medicine, kakkontokasenkyushin'i. We treated five patients with mild COVID-19 at an isolation facility using Kampo medicine, depending on their symptoms. Patients with the olfactory disorder presented with a blocked nose, nasal discharge or taste impairment. Physical examination using Kampo medicine showed similar findings, such as a red tongue with red spots and sublingual vein congestion, which presented as blood stasis and inflammation; thus, we prescribed the Kampo medicine, kakkontokasenkyushin'i. After administration, the numeric rating scale scores of the smell impairment improved within 3 days from 9 to 3 in case 1, from 10 to 0 in case 2, from 9 to 0 in case 3, from 5 to 0 in case 4, and from 9 to 0 within 5 days in case 5. Following the treatment, other common cold symptoms were also alleviated. Kakkontokasenkyushin'i can be used for treating nasal congestion, rhinitis, and inflammation in the nasal mucosa. The olfactory disorder in COVID-19 has been reportedly associated with inflammation and congestion, especially in the olfactory bulb and olfactory cleft. Kakkontokasenkyushin'i may be one of the treatment alternatives for the olfactory disorder with rhinitis in patients with COVID-19.

Sodium Alginate as a Potential Therapeutic Filler: An In Vivo Study in Rats.

Filler injection demand is increasing worldwide, but no ideal filler with safety and longevity currently exists. Sodium alginate (SA) is the sodium salt of alginic acid, which is a polymeric polysaccharide obtained by linear polymerization of two types of uronic acid, d-mannuronic acid (M) and l-guluronic acid (G). This study aimed to evaluate the therapeutic value of SA. Nine SA types with different M/G ratios and viscosities were tested and compared with a commercially available sodium hyaluronate (SH) filler. Three injection modes (onto the periosteum, intradermally, or subcutaneously) were used in six rats for each substance, and the animals were sacrificed at 4 or 24 weeks. Changes in the diameter and volume were measured macroscopically and by computed tomography, and histopathological evaluations were performed. SA with a low M/G ratio generally maintained skin uplift. The bulge gradually decreased over time but slightly increased at 4 weeks in some samples. No capsule formation was observed around SA. However, granulomatous reactions, including macrophage recruitment, were observed 4 weeks after SA implantation, although fewer macrophages and granulomatous reactions were observed at 24 weeks. The long-term volumizing effects and degree of granulomatous reactions differed depending on the M/G ratio and viscosity. By contrast, SH showed capsule formation but with minimal granulomatous reactions. The beneficial and adverse effects of SA as a filler differed according to the viscosity or M/G ratio, suggesting a better long-term volumizing effect than SH with relatively low immunogenicity.

Synthetic Biomarker Design by Using Analyte-Responsive Acetaminophen.

The use of synthetic biomarkers is an emerging technique to improve disease diagnosis. Here, we report a novel design strategy that uses analyte-responsive acetaminophen (APAP) to expand the catalogue of analytes available for synthetic biomarker development. As proof-of-concept, we designed hydrogen peroxide (H2 O2 )-responsive APAP (HR-APAP) and succeeded in H2 O2 detection with cellular and animal experiments. In fact, for blood samples following HR-APAP injection, we demonstrated that the plasma concentration ratio [APAP+APAP conjugates]/[HR-APAP] accurately reflects in vivo differences in H2 O2 levels. We anticipate that our practical methodology will be broadly useful for the preparation of various synthetic biomarkers.

Interstitial fluid flow-induced growth potential and hyaluronan synthesis of fibroblasts in a fibroblast-populated stretched collagen gel culture.

BACKGROUND: Tensioned collagen gels with dermal fibroblasts (DFs) as a dermis model are usually utilized in a static culture (SC) that lacks medium flowing. To make the model closer to its in vivo state, we created a device to perfuse the model with media flowing at a physiological velocity and examined the effects of medium flow (MF) on the cultures. METHODS: We constructed a medium perfusion device for human DF-embedded stretched collagen gels (human dermis model), exposed the model to media that flows upwardly at ~1mL/day, and examined water retention of the gels, cells' growth ability, metabolic activity, expression profiles of nine extracellular matrix (ECM)-related genes. The obtained data were compared with those from the model in SC. RESULTS: MF increases the gels' water retention and cells' growth potential but had little effect on their metabolic activities. MF robustly enhanced hyaluronan synthase 2 (HAS2) and matrix metalloprotease 1 (MMP1) gene expressions but not of the other genes (MMP2, HYAL1, HYAL2, HYAL3, COL1A1, COL3A1, and CD44). MF significantly increased the amounts of cellular hyaluronan and adenosine triphosphate. CONCLUSIONS: The MF at a physiological speed significantly influences the nature of ECMs and their resident fibroblasts and remodels ECMs by regulating hyaluronan metabolism. GENERAL SIGNIFICANCE: Fibroblasts in tensioned collagen gels altered their phenotypes in a MF rate-dependent manner. Collagen gel culture with tension and MF could be utilized as an appropriate in vitro model of interstitial connective tissues to evaluate the pathophysiological significance of mechanosignals generated by fluid flow and cellular/extracellular tension.

Improving the Quality of Postgraduate Education in Traditional Japanese Kampo Medicine for Junior Residents: An Exploratory Survey Conducted in Five Institutions in the Tohoku Area.

Traditional Japanese (Kampo) medicine has been widely applied in general medicine in Japan. In 2001, the model core curriculum for Japanese medical education was revised to include Kampo medicine. Since 2007, all 80 Japanese medical schools have incorporated it within their programs. However, postgraduate training or instruction of Kampo medicine has not been recognized as a goal for the clinical training of junior residents by Japan's Ministry of Health, Labour and Welfare; little is known about postgraduate Kampo medicine education. This exploratory study investigated attitudes about Kampo medicine among junior residents in Japanese postgraduate training programs. A questionnaire survey was administered to junior residents at five institutions in the Tohoku area of Japan. Questions evaluated residents' experiences of prescribing Kampo medicines and their expectations for postgraduate Kampo education and training. As a result, 121 residents responded (response rate = 74%). About 96% of participants had previously received Kampo medicine education at their pre-graduate medical schools and 64% had prescribed Kampo medications. Specifically, daikenchuto was prescribed to prevent ileus and constipation after abdominal surgery and yokukansan was prescribed to treat delirium in the elderly. Residents received on-the-job instruction by attending doctors. Over 70% of participants indicated that there was a need for postgraduate Kampo medicine education opportunities and expected lectures and instruction on how to use it to treat common diseases. In conclusion, we have revealed that junior residents require Kampo medicine education in Japanese postgraduate training programs. The programs for comprehensive pre-graduate and postgraduate Kampo education are expected.

Questionnaire-Based Development of an Educational Program of Traditional Japanese Kampo Medicine.

Traditional Japanese Kampo medicine has been widely used in clinical practice in Japan. Though it is a compulsory subject in Japanese medical schools, a standard educational program in Kampo medicine does not exist. Tohoku University has incorporated Kampo medicine into clinical education via didactic lectures since 2003; however, student evaluations have been lower for Kampo than for all other clinical specialties. We administered a questionnaire about a Kampo medicine course for fifth-year students from 2009 to 2012 and developed an educational program based on feedback obtained. The questionnaire consisted of nine questions (a clear training plan; opportunities for learning, practice, and patient contact; acquisition of medical knowledge and physical examination; learning professionalism; understanding the specialty; overall assessment) that were rated on a 5-point Likert scale along with open-ended questions about the course's strengths and weaknesses. The students responded to the questionnaire after clinical practice in Kampo medicine and other clinical specialty courses. Scores for Kampo medicine and the average of other clinical specialties were compared. All 389 students who participated in Kampo clinical practice answered the questionnaire. In 2009, scores for Kampo medicine for nine questions were lower than for the average of the other clinical specialties. After curriculum reformation involving hands-on training in 2012, all scores except "opportunities to learn about clinical cases" and "opportunities to practice involvement" were higher than the average of all other clinical specialties. In conclusion, we have successfully developed a Kampo medicine educational program for our university through this survey study.

PHYSIOLOGIC AIRWAY RESPONSES TO INHALED HISTAMINE AND ACETYLCHOLINE IN PATIENTS WITH MILD ASTHMA AS ANALYZED BY FORCED OSCILLATION.

BACKGROUND AND AIMS: Asthma is a chronic disease characterized by airway inflammation; it is sometimes difficult to diagnose. For clinical diagnosis, forced oscillation technique (FOT) measures airway reactance and resistance. By FOT, we investigated respiratory resistance and ventilation perfusion ratio inequality in adults with mild asthma. METHODS: We examined 58 adult patients with mild asthma having no inhaled corticosteroid treatment, and 10 adult patients with post-infectious prolonged cough. Using a MostGraph-01 FOT instrument, we evaluated these patients before and after bronchial hyperresponsiveness to acetylcholine (ACh) or histamine (Hist). We measured the following conditions: change of resistance at 5Hz (R5) and 20Hz (R20), R5-R20, reactance at 5Hz, frequency of resonance (Fres), low-frequency reactance area (ALX), and forced expiratory volume in 1 second (FEV1). RESULTS: There were significant changes of R5, R20, R5-R20, X5, Fres, ALX after provocations for ACh or Hist in all patients with asthma, but not in patients with post-infectious prolonged cough. We calculated the percent decrease in FEV1 after provocation with ACh or Hist. For Ach, this decrease in FEV1 correlated with changes in R20 and Fres for all patients. For Hist, the percent decrease in FEV1 correlated with changes in R5, R20, Fres, and ALX for all patients. Furthermore, we investigated these correlations in patients with normalized bronchial hyperresponsiveness to ACh or Hist. For Ach, the percent decrease in FEV1 correlated with changes in Fres or R5-R20. For Hist, this decrease in FEV1 correlated with changes in R5, R20, and Fres. ROC analysis was used to evaluate the diagnostic value of the ratio of change of Fres in BHR to Hist. The area under the curve was 0.7808 (95% CI=0.657-0.904). A reasonably high specificity (100.0%) and a high sensitivity (53.8%) with a cut-off point of 1.5 in the ratio before and after of Fres were obtained. CONCLUSION: The changes in FOT parameters (before and after bronchial airway responses) may detect airway resistance and ventilation perfusion ratio inequality even in adult patients with asthma having normalized bronchial hyperresponsiveness to ACh or Hist. That results may be useful for an early diagnosis of asthma.

Human Adipose-Derived Endothelial Progenitor Cells Accelerate Epithelialization of Radiation Ulcers in Nude Mice.

BACKGROUND: Cotransplantation of adipose-derived stem cells (ASCs) and endothelial progenitor cells has shown superior angiogenic effects compared with ASCs alone in recent animal studies. However, endothelial progenitor cells could only be collected from blood vessels or bone marrow. Thus, the authors have established a method for purifying adipose-derived endothelial progenitor cells (AEPCs). The authors hypothesized that AEPCs would enhance the therapeutic effect of ASCs on radiation ulcers. METHODS: Seven-week-old male nude mice (BALB/cAJcl-nu/nu) were irradiated on the dorsal skin (total 40 Gy); 12 weeks later, 6-mm-diameter wounds were created. The mice were then treated with subcutaneous injection of human ASCs [1 × 10 5 ( n = 4)], human AEPCs [2 × 10 5 or 5 × 10 5 ( n = 5)], combinations of those [ASCs 1 × 10 5 plus AEPCs 2 × 10 5 ( n = 4) or 5 × 10 5 ( n = 5)], or only vehicle ( n = 7). The nonirradiated group was also prepared as a control ( n = 6). The days required for macroscopic epithelialization was compared, and immunostaining for human-derived cells and vascular endothelial cells was performed at day 28. RESULTS: AEPC-ASC combination-treated groups healed faster than the ASC-treated group (14 ± 0 days versus 17 ± 2 days; P < 0.01). Engraftment of the injected cells could not be confirmed. Only the nonirradiated mice had significantly higher vascular density (0.988 ± 0.183 × 10 -5 /µm -2 versus 0.474 ± 0.092 × 10 -5 /µm 2 ; P = 0.02). CONCLUSION: The results suggested therapeutic potentials of AEPCs and an enhanced effect of combination with ASCs. This study is a xenogenic transplantation model, and further validation in an autologous transplantation model is needed. CLINICAL RELEVANCE STATEMENT: Human AEPCs and their combination with ASCs accelerated epithelialization of radiation ulcers in nude mice. The authors suggest that administration of humoral factors secreted from AEPCs (eg, treatment with culture-conditioned media) could be used for the same purpose.

The tRNA thiolation pathway modulates the intracellular redox state in Escherichia coli.

We have performed a screening of hydroxyurea (HU)-sensitive mutants using a single-gene-deletion mutant collection in Escherichia coli K-12. HU inhibits ribonucleotide reductase (RNR), an enzyme that catalyzes the formation of deoxyribonucleotides. Unexpectedly, seven of the mutants lacked genes that are required for the incorporation of sulfur into a specific tRNA modification base, 5-methylaminomethyl-2-thiouridine (mnm(5)s(2)U), via persulfide relay. We found that the expression of RNR in the mutants was reduced to about one-third both in the absence and presence of HU, while sufficient deoxynucleoside triphosphate (dNTP) was maintained in the mutants in the absence of HU but a shortage occurred in the presence of HU. Trans-supply of an RNR R2 subunit rescued the HU sensitivity of these mutants. The mutants showed high intracellular ATP/ADP ratios, and overexpression of Hda, which catalyzes the conversion of DnaA-ATP to DnaA-ADP, rescued the HU sensitivity of the mutants, suggesting that DnaA-ATP represses RNR expression. The high intracellular ATP/ADP ratios were due to high respiration activity in the mutants. Our data suggested that intracellular redox was inclined toward the reduced state in these mutants, which may explain a change in RNR activity by reduction of the catalytically formed disulfide bond and high respiration activity by the NADH reducing potential. The relation between persulfide relay and intracellular redox is discussed.

Characterization of a novel lipolytic enzyme from Aspergillus oryzae.

In this study, we report the characterization of a protein from Aspergillus oryzae, exhibiting sequence identity with paraben esterase from the genus Aspergillus. The coding region of 1,586 bp, including a 77-bp intron, encoded a protein of 502 amino acids. The gene without the signal peptide of 19 amino acids was cloned into a vector, pPICZαC, and expressed successfully in Pichia pastoris as an active extracellular protein. The purified recombinant protein had pH and temperature optima of 7.0-8.0 and 30 °C, respectively, and was stable at the pH range of 7.0-10.0 and up to 40 °C. The optimal substrate for hydrolysis by the purified recombinant protein, among a panel of α-naphthyl esters (C2-C16), was α-naphthyl butyrate (C4), with activity of 0.16 units/mg protein. The considerable hydrolytic activity of the purified recombinant enzyme toward tributyrin was determined. However, no paraben esterase activity was detected toward the ethyl, propyl, and butyl esters of 4-hydroxybenzoic acid. In addition, no activity was detected toward the methyl esters of ferulic, p-coumaric, caffeic, and sinapic acids that would indicate feruloyl esterase activity.