Details of a Researcher - Fukunaga, Koichi

Papers - Fukunaga, Koichi

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Functional dissection of the KRAS G12C mutation by comparison among multiple oncogenic driver mutations in a lung cancer cell line model

Kobayashi K., Terai H., Yasuda H., Hamamoto J., Hayashi Y., Takeuchi O., Mitsuishi Y., Masuzawa K., Manabe T., Ikemura S., Kawada I., Suzuki Y., Soejima K., Fukunaga K.

Biochemical and Biophysical Research Communications 534 1 - 7 2021.01

ISSN 0006291X

The development of molecular targeted therapy has improved clinical outcomes in patients with life-threatening advanced lung cancers with driver oncogenes. However, selective treatment for KRAS-mutant lung cancer remains underdeveloped. We have successfully characterised specific molecular and pathological features of KRAS-mutant lung cancer utilising newly developed cell line models that can elucidate the differences in driver oncogenes among tissues with identical genetic backgrounds. Among these KRAS-mutation-associated specific features, we focused on the IGF2-IGF1R pathway, which has been implicated in the drug resistance mechanisms to AMG 510, a recently developed selective inhibitor of KRAS G12C lung cancer. Experimental data derived from our cell line model can be used as a tool for clinical treatment strategy development through understanding of the biology of lung cancer. The model developed in this paper may help understand the mechanism of anticancer drug resistance in KRAS-mutated lung cancer and help develop new targeted therapies to treat patients with this disease.

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Thymoma-associated T-cell immunodeficiency after radiotherapy: A case report

Nakagawara K., Chubachi S., Azekawa S., Otake S., Saito A., Okada M., Lee K., Masaki K., Koike N., Kamata H., Kawada I., Suzuki S., Ishii M., Fukunaga K.

Respiratory Medicine Case Reports 33 2021.01

Acquired immunodeficiency in thymoma (Good's syndrome) without hypogammaglobulinemia is a rare condition. Here we describe the case of a 29-year-old Japanese woman with thymoma-associated T cell immunodeficiency after radiation therapy. She was admitted to the hospital with refractory pneumonia, which resulted from as T cell immunodeficiency, as revealed through low peripheral lymphocytes and oral candidiasis triggered through radiotherapy and required long-term antimicrobial therapy. Although radiotherapy is commonly administered for thymoma, our findings suggest that physicians should consider carrying out lymphocyte counts during thymoma treatment.

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Clinical utility of SARS-CoV-2 whole genome sequencing in deciphering source of infection

Takenouchi T., Iwasaki Y.W., Harada S., Ishizu H., Uwamino Y., Uno S., Osada A., Abe K., Hasegawa N., Murata M., Takebayashi T., Fukunaga K., Saya H., Kitagawa Y., Amagai M., Siomi H., Kosaki K.

Journal of Hospital Infection 107 40 - 44 2021.01

ISSN 01956701

Coronavirus disease 2019 (COVID-19) caused by human severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a worldwide problem. From the standpoint of hospital infection control, determining the source of infection is critical. We conducted the present study to evaluate the efficacy of using whole genome sequencing to determine the source of infection in hospitalized patients who do not have a clear infectious contact history. Recently, we encountered two seemingly separate COVID-19 clusters in a tertiary hospital. Whole viral genome sequencing distinguished the two clusters according to the viral haplotype. However, the source of infection was unclear in 14 patients with COVID-19 who were clinically unlinked to clusters 1 or 2. These patients, who had no clear history of infectious contact within the hospital (‘undetermined source of infection’), had haplotypes similar to those in cluster 2 but did not have two of the mutations used to characterize cluster 2, suggesting that these 14 cases of ‘undetermined source of infection’ were not derived from cluster 2. Whole viral genome sequencing can be useful for confirming that sporadic COVID-19 cases with an undetermined source of infection are indeed not part of clusters at the institutional level.

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Clinical characteristics of patients with not well-controlled severe asthma in Japan: Analysis of the Keio Severe Asthma Research Program in Japanese population (KEIO-SARP) registry

Tanosaki T., Kabata H., Matsusaka M., Miyata J., Masaki K., Mochimaru T., Okuzumi S., Kuwae M., Watanabe R., Suzuki Y., Sayama K., Izuhara K., Asano K., Fukunaga K.

Allergology International 70 ( 1 ) 61 - 67 2021.01

ISSN 13238930

Background: Multiple phenotypes exist within the classification of severe asthma. However, characteristics of patients with not well-controlled severe asthma have not been well identified. Methods: Japanese patients with asthma (age ≥ 20 years) were enrolled at the Keio University Hospital and its affiliated hospitals in this observational study (Keio Severe Asthma Research Program). Among them, patients with severe asthma (those undergoing Global Initiative for Asthma [GINA] 2018 step 4 or 5 treatment) were included in this analysis and investigated clinical characteristics based on asthma control status. Results: Of the 154 patients (men, 46.8%; age, 60.1 ± 14.9 years), 87 (56.5%) had not well-controlled (partly controlled and uncontrolled) asthma (GINA step 4, 42 patients; step 5, 45 patients). Overall, there were no significant differences in clinical characteristics between patients with well-controlled and not well-controlled asthma. However, cluster analysis revealed that distinct 5 clusters (cluster 1, well-controlled; cluster 2, eosinophilic; cluster 3, non–type 2 inflammation; cluster 4, high periostin; and cluster 5, late-onset type 2 inflammation), and clusters 2–5 were not well-controlled. Among them, cluster 3 was characterized by low eosinophil counts, low periostin levels, and less frequent olfactory disturbance, and this cluster had the worst asthma control. Conclusions: Japanese patients with severe asthma were divided into well-controlled and not-well controlled asthma, and we confirmed heterogeneity of not well-controlled severe asthma. These patients, especially non-type 2 phenotype, require a further therapeutic approach. (University Hospital Medical Information Network Clinical Trials Registry, UMIN000002980)

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An official JRS statement: The principles of fractional exhaled nitric oxide (FeNO) measurement and interpretation of the results in clinical practice

Matsunaga K., Kuwahira I., Hanaoka M., Saito J., Tsuburai T., Fukunaga K., Matsumoto H., Sugiura H., Ichinose M.

Respiratory Investigation 59 ( 1 ) 34 - 52 2021.01

ISSN 22125345

Nitric oxide (NO) is produced in the body and has been shown to have diverse actions in the abundance of research that has been performed on it since the 1970s, leading to Furchgott, Murad, and Ignarro receiving the Nobel Prize in Physiology or Medicine in 1998. NO is produced by nitric oxide synthase (NOS). NOS is broadly distributed, being found in the nerves, blood vessels, airway epithelium, and inflammatory cells. In asthma, inflammatory cytokines induce NOS activity in the airway epithelium and inflammatory cells, producing large amounts of NO. Measurement of fractional exhaled nitric oxide (FeNO) is a simple, safe, and quantitative method of assessing airway inflammation. The FeNO measurement method has been standardized and, in recent years, this noninvasive test has been broadly used to support the diagnosis of asthma, monitor airway inflammation, and detect asthma overlap in chronic obstructive pulmonary disease (COPD) patients. Since the normal upper limit of FeNO for healthy Japanese adults is 37 ppb, values of 35 ppb or more are likely to be interpreted as a signature of inflammatory condition presenting features with asthma, and this value is used in clinical practice. Research is also underway for clinical application of these measurements in other respiratory diseases such as COPD and interstitial lung disease. Currently, there remains some confusion regarding the significance of these measurements and the interpretation of the results. This statement is designed to provide a simple explanation including the principles of FeNO measurements, the measurement methods, and the interpretation of the measurement results.

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Left Ventricular Thrombus With COVID-19 Complication in a Patient With Dilated Cardiomyopathy

Imaeda S., Kabata H., Shiraishi Y., Kamata H., Tsuruta H., Yuasa S., Ishii M., Fukuda K., Fukunaga K.

CJC Open 3 ( 1 ) 124 - 126 2021.01

Thrombosis, especially venous thromboembolism, is a complication often associated with coronavirus disease 2019 (COVID-19). However, there have been relatively few reports of arterial thrombosis. Here, we describe a case of non-severe COVID-19 in a patient with dilated cardiomyopathy. After admission, symptoms, laboratory data, and imaging findings improved, but D-dimer levels gradually increased. Contrast computed tomography and echocardiography revealed a left ventricular thrombus. Anticoagulant treatment diminished the thrombus, and the patient recovered and was discharged. Although a left ventricular thrombus is a rare COVID-19 complication, performing appropriate diagnostic tests could improve COVID-19 mortality in patients with dilated cardiomyopathy.

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A prospective cohort study to assess obstructive respiratory disease phenotypes and endotypes in Japan: The trait study design

Hizawa N., Fukunaga K., Sugiura H., Nakano Y., Kato M., Sugiyama Y., Hanazawa T., Kaise T., Tal-Singer R., Jones P.W., Barnes N., Compton C., Ishii T.

International Journal of COPD 16 1813 - 1822 2021

ISSN 11769106

Background: Asthma, chronic obstructive pulmonary disease (COPD), and asthma–COPD overlap (ACO) are complex and heterogeneous diseases that share clinical characteristics (phenotypes) and molecular mechanisms (endotypes). Whilst physicians make clinical decisions on diagnostic groups, for some such as ACO there is no commonly accepted criteria. An alternative approach is to evaluate phenotypes and endotypes that are considered to respond well to a specific type of treatment (“treatable traits”) rather than diagnostic labels. Purpose: The prospective, longitudinal, and observational TRAIT study will evaluate disease characteristics, including both phenotypes and endotypes, in relation to the presentation of obstructive respiratory disease characteristics in patients diagnosed with asthma, COPD, or ACO in Japan, with the aim of further understanding the clinical benefit of a treatable traits-based approach. Patients and Methods: A total of 1500 participants will be enrolled into three cohorts according to their treating physician’s diagnosis of asthma, COPD, or ACO at screening. Part 1 of the study will involve cross-sectional phenotyping and endotyping at study enrollment. Part 2 of the study will evaluate the progression of clinical characteristics, biomarker profiles, and treatment over a 3-year follow-up period. The follow-up will involve three annual study visits and three telephone calls scheduled at 6-month intervals. A substudy involving 50 participants from the asthma cohort (in which the ratio will be approximately 1:1 including 25 participants with a smoking history of ≥10 pack-years and 25 participants with no smoking history), 100 participants from the ACO cohort, and 100 participants from the COPD cohort will evaluate disease phenotypes using inspiratory and expiratory computed tomography scans. Conclusion: TRAIT will describe clinical characteristics of patients with obstructive respiratory diseases to better understand potential differences and similarities between clinical diagnoses, which will support the improvement of personalized treatment strategies.

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Clinical characteristics and therapeutic outcomes of metastatic peritoneal carcinomatosis in non-small-cell lung cancer

Tani T., Nakachi I., Ikemura S., Nukaga S., Ohgino K., Kuroda A., Terai H., Masuzawa K., Shinozaki T., Ishioka K., Funatsu Y., Koh H., Fukunaga K., Soejima K.

Cancer Management and Research 13 7497 - 7503 2021

Background: Metastatic peritoneal carcinomatosis (MPC) is not common in patients with non-small cell lung cancer (NSCLC), and the clinical characteristics and treatment outcomes are still unclear. Patients and Methods: We recruited 46 NSCLC patients with MPC at Keio University and affiliated hospitals (Keio Lung Oncology Group) between January 2011 and December 2017, then retrospectively investigated their clinical characteristics and the impact of treatment interventions on their survival. Results: The profile of histological subtype was predominantly adenocarcinoma and 15 patients harbored driver oncogenes. Univariate and multivariate analysis demonstrated that performance status and the presence of a driver oncogene were significantly associated with the prolonged overall survival (OS). Regarding treatment, the median OS in the treatment group (9.3 months) was significantly longer than in the best supportive care group (1.3 months) (P <0.0001). Conclusion: The prognosis of MPC in NSCLC patients who receive only the best supportive care is poor, but therapeutic intervention may improve prognosis.

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Lorlatinib-induced visual and auditory hallucinations: A case report

Hakamata J., Nakada H., Muramatsu H., Masuzawa K., Terai H., Ikemura S., Fukunaga K., Aomori T.

Clinical Case Reports 2021

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PATHOPHYSIOLOGY AND TREATMENT OF ASTHMA IN PATIENTS WITH EOSINOPHILIC RHINOSINUSITIS AND OTITIS MEDIA

Fukunaga K., Miyata J.

Japanese Journal of Allergology 70 ( 10 ) 1343 - 1349 2021

ISSN 00214884

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Author's reply: Atrial fibrillation and sleep apnea: A chicken and egg situation

Kimura, T., Kohno, T., Fukunaga, K., Yamasawa, W., Fujisawa, T., Fukuoka, R., Nakajima, K., Kashimura, S., Kunitomi, A., Katsumata, Y., Nishiyama, T., Nishiyama, N., Aizawa, Y., Fukuda, K. and Takatsuki, S.

Int J Cardiol 270 187 2018.11

ISSN 1874-1754

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Intermittent Exposure to Cigarette Smoke Increases Lung Tumors and the Severity of Emphysema More than Continuous Exposure

Kameyama, N., Chubachi, S., Hegab, A. E., Yasuda, H., Kagawa, S., Tsutsumi, A., Fukunaga, K., Shimoda, M., Kanai, Y., Soejima, K. and Betsuyaku, T.

Am J Respir Cell Mol Biol 59 ( 2 ) 179 - 188 2018.08

ISSN 1535-4989

Lung cancer and chronic obstructive pulmonary disease are leading causes of morbidity and mortality worldwide, and cigarette smoking is a main risk factor for both. The presence of emphysema, an irreversible lung disease, further raises the risk of lung cancer in patients with chronic obstructive pulmonary disease. The mechanisms involved in smoke-induced tumorigenesis and emphysema are not fully understood, attributable to a lack of appropriate animal models. Here, we optimized a model of cigarette smoke (CS)-induced lung cancer and emphysema in A/J mice treated with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, a potent carcinogen. We investigated whether variations in CS exposure patterns with the same total amount and duration of exposure affect tumorigenesis and/or development of emphysema. Continuous CS exposure for 3 months significantly suppressed 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced development of adenomas and adenocarcinomas; however, emphysema independently developed during this period. Surprisingly, intermittent CS exposure increased the severity of emphysema and resulted in a higher incidence of adenocarcinomas. Furthermore, intermittent CS exposure elicited a marked increase in M2-polarized macrophages within and near the developed tumors. By employing a CS exposure protocol with repeated cycles of cessation and relapse, we provide evidence that intermittent CS exposure enhances tumorigenesis and emphysema progression more than that of continuous CS exposure.

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Development of Necrotizing Myopathy Following Interstitial Lung Disease with Anti-signal Recognition Particle Antibody

Kusumoto, T., Okamori, S., Masuzawa, K., Asakura, T., Nishina, N., Chubachi, S., Naoki, K., Fukunaga, K. and Betsuyaku, T.

Intern Med 57 ( 14 ) 2045 - 2049 2018.07

ISSN 1349-7235

A 72-year-old man was admitted due to dyspnea on exertion with interstitial shadows and elevated serum creatinine kinase (CK). Despite a close examination, which included magnetic resonance imaging (MRI), we could not diagnose myopathy. Prednisolone was administered and gradually tapered. One year later, anti-signal recognition particle (SRP) antibody was confirmed and he was re-admitted for hypoxemia with elevated CK. MRI revealed muscle edema and a histopathological examination of a muscle biopsy specimen showed necrotizing myopathy. Prednisolone, cyclosporine, and intravenous immunoglobulin were administered. Physicians should carefully monitor muscle symptoms and serum CK levels in cases of interstitial lung disease with anti-SRP antibodies.

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Prevalence and clinical characteristics of obstructive- and central-dominant sleep apnea in candidates of catheter ablation for atrial fibrillation in Japan

Kohno, T., Kimura, T., Fukunaga, K., Yamasawa, W., Fujisawa, T., Fukuoka, R., Nakajima, K., Kashimura, S., Kunitomi, A., Katsumata, Y., Nishiyama, T., Nishiyama, N., Aizawa, Y., Fukuda, K. and Takatsuki, S.

Int J Cardiol 260 99 - 102 2018.06

ISSN 1874-1754

INTRODUCTION: We aimed to study the prevalence and types of sleep apnea (SA) as well as their clinical characteristics in atrial fibrillation (AF) ablation candidates in Japan. METHODS: Before catheter ablation, 197 consecutive AF patients (age: 60+/-9years, body mass index; 25.0+/-3.0) were evaluated with portable polygraphy. We compared the clinical characteristics, according to the severity of SA as well as its types, as defined by the presence of obstruction and the mixed vs. central apnea indices. RESULTS: The mean apnea-hypopnea index (AHI) was 17.7+/-11.9, with 135 AF patients having an AHI >/=10 (68.5%). Patients with an AHI >/=10 had a significantly higher body mass index, plasma brain natriuretic peptide (BNP) level, prevalence of hypertension, and larger left atrial size. Among patients with an AHI >/=10, the incidence of obstructive-dominant SA was 60.9% and that of central-dominant SA was 7.6%. The prevalence of hypertension was significantly higher in obstructive-dominant SA patients (obstructive vs. central: 48.3% vs. 20.0%, P=0.038). The obstructive apnea index correlated with plasma BNP level and age, but the central and mixed apnea indices did not. CONCLUSIONS: The prevalence of SA was common in AF ablation candidates, even without an obesity epidemic, and the SA type was predominantly obstructive. Portable polygraphy was useful for detecting undiagnosed SA patients in AF ablation candidates.

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12-OH-17,18-Epoxyeicosatetraenoic acid alleviates eosinophilic airway inflammation in murine lungs

Mochimaru, T., Fukunaga, K., Miyata, J., Matsusaka, M., Masaki, K., Kabata, H., Ueda, S., Suzuki, Y., Goto, T., Urabe, D., Inoue, M., Isobe, Y., Arita, M. and Betsuyaku, T.

Allergy 73 ( 2 ) 369 - 378 2018.02

ISSN 1398-9995

BACKGROUND: Asthma is characterized by airway inflammation and obstruction with eosinophil infiltration into the airway. Arachidonic acid, an omega-6 fatty acid, is metabolized into cysteinyl leukotriene with pro-inflammatory properties for allergic inflammation, whereas the omega-3 fatty acid eicosapentaenoic acid (EPA) and its downstream metabolites are known to have anti-inflammatory effects. In this study, we investigated the mechanism underlying the counter-regulatory roles of EPA in inflamed lungs. METHODS: Male C57BL6 mice were sensitized and challenged by ovalbumin (OVA). After EPA treatment, we evaluated the cell count of Bronchoalveolar lavage fluid (BALF), mRNA expressions in the lungs by q-PCR, and the amounts of lipid mediators by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based lipidomics. We investigated the effect of the metabolite of EPA by in vivo and in vitro studies. RESULTS: Eicosapentaenoic acid treatment reduced the accumulation of eosinophils in the airway and decreased mRNA expression of selected inflammatory mediators in the lung. Lipidomics clarified the metabolomic profile in the lungs. Among EPA-derived metabolites, 12-hydroxy-17,18-epoxyeicosatetraenoic acid (12-OH-17,18-EpETE) was identified as one of the major biosynthesized molecules; the production of this molecule was amplified by EPA administration and allergic inflammation. Intravenous administration of 12-OH-17,18-EpETE attenuated airway eosinophilic inflammation through downregulation of C-C chemokine motif 11 (CCL11) mRNA expression in the lungs. In vitro, this molecule also inhibited the release of CCL11 from human airway epithelial cells stimulated with interleukin-4. CONCLUSION: These results demonstrated that EPA alleviated airway eosinophilic inflammation through its conversion into bioactive metabolites. Additionally, our results suggest that 12-OH-17,18-EpETE is a potential therapeutic target for the management of asthma.

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Association between severity of obstructive sleep apnea and glycated hemoglobin level in Japanese individuals with and without diabetes

Kurosawa, H., Saisho, Y., Fukunaga, K., Haraguchi, M., Yamasawa, W., Kurihara, I., Betsuyaku, T. and Itoh, H.

Endocr J 65 ( 1 ) 121 - 127 2018.01

ISSN 1348-4540

Aim of this study was to examine the association between the severity of obstructive sleep apnea (OSA) and dysglycemia in Japanese individuals with and without type 2 diabetes (T2DM). We enrolled 115 individuals diagnosed with OSA with an apnea hypopnea-index (AHI) >/= 20 in whom continuous positive airway pressure (CPAP) therapy was introduced (N = 115, 44 with T2DM, age 62 +/- 11 years, BMI 27.0 +/- 4.4 kg/m(2) and AHI median 36.1; interquartile range 27.2-48.1). During admission, the severity of OSA was evaluated by polysomnography, and its association with glycated hemoglobin (HbA1c) level was examined. Continuous glucose monitoring (CGM) was also conducted during the admission in 94 individuals. Apnea-hypopnea index (AHI), non-rapid eye movement (REM) AHI, minimum peripheral capillary oxygen saturation (SpO2) and percentage of sleep time (%TST) with SpO2 < 90% were significantly associated with HbA1c level in total and non-diabetic individuals (all p < 0.05) but not in those with T2DM, the majority of whom were treated with anti-diabetic medications. The associations of the non-REM AHI and %TST with SpO2 < 90% with HbA1c level remained significant after adjustment for age, sex and BMI in non-diabetic and T2DM subjects treated with dietary therapy only. Mean glucose level, but not SD or coefficient of variation of glucose, assessed by CGM was significantly associated with AHI and non-REM AHI in non-diabetic subjects after adjustment for age, sex and BMI. In conclusion, the severity of OSA was associated with increased HbA1c level independently of BMI in Japanese individuals, especially in those without diabetes.

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Allergic bronchopulmonary aspergillosis in Japan: A nationwide survey

Oguma, T., Taniguchi, M., Shimoda, T., Kamei, K., Matsuse, H., Hebisawa, A., Takayanagi, N., Konno, S., Fukunaga, K., Harada, K., Tanaka, J., Tomomatsu, K. and Asano, K.

Allergol Int 67 ( 1 ) 79 - 84 2018.01

ISSN 1440-1592

BACKGROUND: Allergic bronchopulmonary aspergillosis (ABPA) is an allergic pulmonary disease characterized by a hypersensitivity reaction to Aspergillus species colonizing the airways. The clinical characteristics of ABPA may differ depending on genetic and environmental background. We performed a nationwide survey to determine the clinical characteristics of ABPA in Japan. METHODS: In 2013, a questionnaire on physician-diagnosed ABPA/allergic bronchopulmonary mycosis was sent to 903 medical centers specializing in respiratory or allergic diseases. Cases fulfilling the following criteria were categorized as possible ABPA-central bronchiectasis (ABPA-CB): 1) presence of specific serum immunoglobulin E (IgE) antibodies or a positive skin reaction to Aspergillus, and 2) bronchiectasis or mucoid impaction in the central bronchi. RESULTS: Of 499 physician-diagnosed cases reported by 132 clinical centers, 358 cases met the criteria for possible ABPA-CB. Median age of ABPA-CB onset was 57 (interquartile range, 44-68) years; later-onset disease, developing >/=50 years of age, accounted for 66% of the cases and was associated with female sex, delayed onset of asthma, and lower levels of serum IgE. A third of the patients (120 patients, 34%) exhibited low levels of serum total IgE (<1000 IU/mL). Aspergillus species were isolated from sputum in 126/213 cases (59%), and Schizophyllum commune was identified in 12 (6%) patients. During the course of the treatment, ABPA recurred in 169 (48%) cases. CONCLUSIONS: This nationwide survey identified several unique clinical characteristics of ABPA in Japan, such as late-onset, relatively lower serum IgE levels, and frequent recurrences/flares.

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Subphenotypes of type 2 severe asthma in adults

Matsusaka, M., Fukunaga, K., Kabata, H., Izuhara, K., Asano, K. and Betsuyaku, T.

J Allergy Clin Immunol Pract 6 ( 1 ) 274 - 276 e2 2018.01

ISSN 2213-2201

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Construction of a system using a deep learning algorithm to count cell numbers in nanoliter wells for viable single-cell experiments

Kamatani, T., Fukunaga, K., Miyata, K., Shirasaki, Y., Tanaka, J., Baba, R., Matsusaka, M., Kamatani, N., Moro, K., Betsuyaku, T. and Uemura, S.

Sci Rep 7 ( 1 ) 16831 2017.12

ISSN 2045-2322

For single-cell experiments, it is important to accurately count the number of viable cells in a nanoliter well. We used a deep learning-based convolutional neural network (CNN) on a large amount of digital data obtained as microscopic images. The training set consisted of 103 019 samples, each representing a microscopic grayscale image. After extensive training, the CNN was able to classify the samples into four categories, i.e., 0, 1, 2, and more than 2 cells per well, with an accuracy of 98.3% when compared to determination by two trained technicians. By analyzing the samples for which judgments were discordant, we found that the judgment by technicians was relatively correct although cell counting was often difficult by the images of discordant samples. Based on the results, the system was further enhanced by introducing a new algorithm in which the highest outputs from CNN were used, increasing the accuracy to higher than 99%. Our system was able to classify the data even from wells with a different shape. No other tested machine learning algorithm showed a performance higher than that of our system. The presented CNN system is expected to be useful for various single-cell experiments, and for high-throughput and high-content screening.

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The alpha-lipoic acid derivative DHLHZn: a new therapeutic agent for acute lung injury in vivo

Shoji, Y., Takeuchi, H., Fukuda, K., Fukunaga, K., Nakamura, R., Takahashi, T., Wada, N., Kawakubo, H., Miyasho, T., Hiratsuka, T., Inomata, M., Betsuyaku, T. and Kitagawa, Y.

Inflamm Res 66 ( 9 ) 803 - 811 2017.09

ISSN 1420-908X

OBJECTIVE AND DESIGN: An animal experiment was performed to demonstrate the anti-inflammatory effects of an alpha-lipoic acid (ALA) derivative, dihydrolipoyl histidinate zinc complex (DHLHZn) for acute lung injury (ALI) and to investigate the mechanism of action. MATERIAL: Rats were randomly divided into three experimental groups: control group (n = 17), DHLHZn(-) group (n = 11, ALI model rats), and DHLHZn(+) group (n = 12, ALI model rats treated by DHLHZn). TREATMENT: Lipopolysaccharides (LPS, 10 mg/kg) were administered intratracheally in the DHLHZn(-) group and the DHLHZn(+) group. For the DHLHZn(+) group, DHLHZn (100 mg/kg) was administered intraperitoneally 2 h prior to LPS administration. METHODS: Four hours after LPS administration, bronchoalveolar lavage fluid (BALF) and lung tissue were collected. The findings were analyzed using the Mann-Whitney U test. RESULTS: Total number of cells, number of neutrophils and lymphocytes, levels of various inflammatory cytokines, and NF-kB p65 concentration of BALF were significantly lower in the DHLHZn(+) group than in the DHLHZn(-) group (p < 0.05). ALI pathology scores were significantly lower in the DHLHZn(+) group than in the DHLHZn(-) group (p < 0.001). CONCLUSIONS: Anti-inflammatory effects of DHLHZn for ALI were demonstrated by BALF and histopathological findings. The mechanism of action of DHLHZn was considered to be via inhibition of the NF-kB signaling pathway. DHLHZn is thus suggested to be a new prophylactic agent for ALI.

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