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Background Interstitial lung disease (ILD) patients often develop lung cancer. However, previous studies on the incidences of lung cancer in ILD patients focused on specific aetiologies, such as idiopathic pulmonary fibrosis (IPF). The lung cancer incidences in these patients have not been investigated, and thus, we aimed to evaluate them here. Methods ILD patients at our hospital were retrospectively reviewed. The cumulative incidences of lung cancer in patients with various ILDs were estimated with Kaplan-Meier curves and compared between ILD groups using log-rank tests. The association between several variables at initial diagnosis and lung cancer development was assessed with Cox proportional hazards regression analysis to identify predictors. Results In all, 606 ILD patients, including 161 with IPF, 133 with non-IPF idiopathic interstitial pneumonias, 160 with chronic hypersensitivity pneumonitis, 87 with connective tissue disease-related ILDs, 19 with pulmonary sarcoidosis, and 46 with other ILDs, were included. Twenty-eight patients developed lung cancer. The cumulative incidences of lung cancer at 1, 3, and 5 years were: 1.9, 5.7, and 12.3% with IPF, respectively; 0.8, 0.8, and 4.0% in non-IPF idiopathic interstitial pneumonias; 2.0, 4.6, and 11.0% in chronic hypersensitivity pneumonitis; and 1.1, 1.1, and 2.9% in connective tissue disease-related ILDs. IPF patients had a higher incidence of lung cancer than non-IPF idiopathic interstitial pneumonia patients (p = 0.036). A radiological usual interstitial pneumonia pattern, forced vital capacity value, and pack-years were associated with lung cancer development (hazard ratios 2.959, 1.031, 1.011; 95% confidence intervals 1.257–6.963, 1.006–1.057, 1.002–1.020, p = 0.013, 0.017, 0.020, respectively). Conclusions The lung cancer incidence is higher in IPF patients than in non-IPF idiopathic interstitial pneumonia patients and is equally high in patients with chronic hypersensitivity pneumonitis and IPF.

Declines in mitochondrial functions are associated with aging. The combination of 5‐aminolevulinic acid (5‐ALA) and sodium ferrous citrate (SFC) improves mitochondrial functions in cultured cells. In this study, we investigated the effects of dietary supplementation with 5‐ALA and SFC (5‐ALA/SFC) on the healthspan and life span of Drosophila melanogaster. Adult Drosophila fruit flies were fed cornmeal food containing various concentrations of 5‐ALA/SFC. Locomotor functions, life span, muscle architecture, and age‐associated changes in mitochondrial function were analyzed. We found that feeding 5‐ALA/SFC mitigated age‐associated declines in locomotor functions and extended organismal life span. Moreover, 5‐ALA/SFC preserved muscle architecture and maintained the mitochondrial membrane potential in aged animals. Since 5‐ALA phosphate/SFC is used as a human dietary supplement, our results suggest that it could be used to slow the age‐related declines in muscle functions, prevent age‐associated clinical conditions such as frailty, and extend healthspan and life span., Age‐related declines in physiological systems cause problematic features such as frailty. In this study, we found that 5‐aminolevulinic acid hydrochloride and sodium ferrous citrate (5‐ALA/SFC) mitigated age‐associated declines in locomotor functions and the muscle architecture, preserved the mitochondrial membrane potential, and extended life span in Drosophila melanogaster. 5‐ALA/SFC might be used to prevent age‐associated clinical conditions.

Of the 80 solid tumor cases in which liquid biopsy (LB) was performed using Guardant360 in the PROFILE study, nine were lung cancer cases. Here, we review three cases in which LB was useful in diagnosing ALK fusion‐positive lung cancer, selecting sequential ALK‐tyrosine kinase inhibitors, confirming uncommon EGFR mutations, and receiving biomarker‐compatible therapy., For advanced lung cancer, the genome profiling test by minimally invasive liquid biopsy (LB) is gathering attention. Herein, we showed three cases who received target therapy based on LB. LB assisted in diagnosing EML‐ALK fusion‐positive lung cancer in case 1, detected resistance genes for ALK inhibitor, ALK E1210K and F1174C in case 2, and detected uncommon EGFR mutations that could not be detected by a conventional test in case 3. In these cases, target therapy based on genome profiling resulted in a favorable therapeutic effect.

Mitochondria are key cytoplasmic organelles. Their activation is critical for the generation of T cell proliferation and cytotoxicity. Exhausted tumor‐infiltrating T cells show a decreased mitochondrial function and mass. 5‐Aminolevulinic acid (5‐ALA), a natural amino acid that is only produced in the mitochondria, has been shown to influence metabolic functions. We hypothesized that 5‐ALA with sodium ferrous citrate (SFC) might provide metabolic support for tumor‐infiltrating T cells. In a mouse melanoma model, we found that 5‐ALA/SFC with a programmed cell death‐ligand 1 (PD‐L1) blocking Ab synergized tumor regression. After treatment with 5‐ALA/SFC and anti‐PD‐L1 Ab, tumor infiltrating lymphocytes (TILs) were not only competent for the production of cytolytic particles and cytokines (granzyme B, interleukin‐2, and γ‐interferon) but also showed enhanced Ki‐67 activity (a proliferation marker). The number of activated T cells (PD‐1+Tim‐3−) was also significantly increased. Furthermore, we found that 5‐ALA/SFC activated the mitochondrial functions, including the oxygen consumption rate, ATP level, and complex V expression. The mRNA levels of Nrf‐2, HO‐1, Sirt‐1, and PGC‐1α and the protein levels of Sirt‐1 were upregulated by treatment with 5‐ALA/SFC. Taken together, our findings revealed that 5‐ALA/SFC could be a key metabolic regulator in exhausted T cell metabolism and suggested that 5‐ALA/SFC might synergize with anti‐PD‐1/PD‐L1 therapy to boost the intratumoral efficacy of tumor‐specific T cells. Our study not only revealed a new aspect of immune metabolism, but also paved the way to develop a strategy for combined anti‐PD‐1/PD‐L1 cancer immunotherapy., Exhausted T cells have decreased effector and proliferation function. We found that the mRNA levels of the Nrf‐2, HO‐1, Sirt‐1, and PGC‐1α and the protein levels of Sirt‐1 were upregulated by treatment with 5‐aminolevulinic acid (5‐ALA)/sodium ferrous ion (SFC). The enforced expression of PGC‐1α in activated T cells rescued their effector function and enhanced the efficacy of the mitochondrial function. Our findings revealed that 5‐ALA/SFC could be a key metabolic regulator in exhausted T cell metabolism and suggested that 5‐ALA/SFC might synergize with anti‐programmed cell death‐1/programmed cell death‐ligand 1 therapy to boost the intratumoral efficacy of tumor‐specific T cells. ​

Background Anticoagulant therapy for patients with severe coronavirus disease (COVID-19) pneumonia is considered to improve the hypercoagulable and inflammatory state. However, bleeding complications should also be considered. Case presentation A 77-year-old man with a history of falls was diagnosed with COVID-19. Owing to his severe condition, he was intubated and transferred to our hospital for intensive care. Favipiravir, tocilizumab, unfractionated heparin, and ART-123 were administered to treat COVID-19 and manage the antithrombotic prophylaxis for paroxysmal atrial fibrillation (Af). On the 6th day after admission, a hematoma was noted on the left chest wall. Computed tomography (CT) revealed multiple hematomas, including hematomas on his chest wall and obturatorius internus muscle. Emergency angiography transcatheter embolization (TAE) was performed. The patient was transferred to another hospital 23 days after TAE, without complications. Conclusion Our findings show that anticoagulation therapy and a history of falls induced multiple hematomas in a COVID-19 patient and that the condition was managed with TAE. When anticoagulants are considered in the management of Af and COVID-19 associated coagulopathy, it is necessary to closely monitor potential bleeding complications.

Background The prediction of COVID-19 disease behavior in the early phase of infection is challenging but urgently needed. MuLBSTA score is a scoring system that predicts the mortality of viral pneumonia induced by a variety of viruses, including coronavirus, but the scoring system has not been verified in novel coronavirus pneumonia. The aim of this study was to validate this scoring system for estimating the risk of disease worsening in patients with COVID-19. Methods This study included the patients who were treated between April 1 st and March 13 th , 2020. The patients were classified into mild, moderate, and severe groups according to the extent of respiratory failure. MuLBSTA score was applied to estimate the risk of disease worsening in each severity group and we validated the utility of the scoring system. Results A total of 72 patients were analyzed. Among the 46 patients with mild disease, 17 showed disease progression to moderate or severe disease after admission. The model showed a sensitivity of 100% and a specificity of only 34.5% with a cut-off value of 5 points. Among the 55 patients with mild or moderate disease, 6 deteriorated to severe disease, and the model showed a sensitivity of 83.3% and a specificity of 71.4% with a cut-off value of 11 points. Conclusions This study showed that MuLBSTA score is a potentially useful tool for predicting COVID-19 disease behavior. This scoring system may be used as one of the criteria to identify high-risk patients worsening to life-threatening status.

Hypersensitivity pneumonitis (HP) typically presents with interstitial inflammation and granulomas induced by an aberrant immune response to inhaled Ags in sensitized individuals. Although IL-17A is involved in the development of HP, the cellular sources of IL-17A and the mechanisms by which IL-17A contributes to granuloma formation remain unclear. Recent studies report that γδ T cells produce IL-17A and exhibit memory properties in various diseases. Therefore, we focused on IL-17A–secreting memory γδ T cells in the sensitization phase and aimed to elucidate the mechanisms by which IL-17A contributes to granuloma formation in HP. We induced a mouse model of HP using pigeon dropping extract (PDE) in wild-type and IL-17A knockout (IL-17A−/−) mice. IL-17A−/− mice exhibited reduced granulomatous areas, attenuated aggregation of CD11b+ alveolar macrophages, and reduced levels of CCL2, CCL4, and CCL5 in the bronchoalveolar lavage fluid. Among IL-17A+ cells, more γδ T cells than CD4+ cells were detected after intranasal PDE administration. Interestingly, the expansion of IL-17A–secreting Vγ4+ or Vγ1−Vγ4− cells of convalescent mice was enhanced in response to the sensitizing Ag. Additionally, coculture of macrophages with PDE and Vγ4+ cells purified from PDE-exposed convalescent mice produced significantly more IL-17A than coculture with Vγ4+ cells from naive mice. Our findings demonstrate that in the sensitization phase of HP, IL-17A–secreting memory γδ T cells play a pivotal role. Furthermore, we characterized the IL-17A/CCL2, CCL4, CCL5/CD11b+ alveolar macrophage axis, which underlies granuloma formation in HP. These findings may lead to new clinical examinations or therapeutic targets for HP.

Background/aim We evaluated urinary levels of porphyrin metabolites, such as uroporphyrin (UP) and coproporphyrin (CP), after 5-Aminolevulinic acid (ALA) administration in patients with or without pancreatic cancer (PaC). Patients and methods Sixty-seven subjects with PaC, 11 with pancreatitis, and 9 with normal pancreas (NP) were enrolled. Urine samples from all subjects were collected prior to ALA administration and at more than 4 hours after ALA administration. We measured the urinary levels of UP and CP by high-performance liquid chromatography analysis. Results The PaC group showed significantly higher UP levels compared to NP groups (104.9 nmol/g Cre vs. 53.4 nmol/g Cre, p=0.014). Moreover, PaC patients with long-term survival had significantly lower urinary levels of UP at diagnosis (98.8 nmol/gCre) than the short-term survival group (125.2 nmol/gCre) (p=0.042). Conclusion The urinary levels of UP after ALA administration might serve as a promising biomarker for diagnosis and prognosis prediction of PaC.

Background and Purpose: In this study, we evaluated the effectiveness and safety of photodynamic diagnosis (PDD) of upper urinary tract carcinoma (UTUC) using oral 5-aminolevulinic acid (5-ALA). Ma...

Photodynamic therapy (PDT) is a clinically approved, minimally invasive treatment for malignant tumors. Protoporphyrin IX (PpIX), derived from 5-aminolevulinic acid (5-ALA) as the prodrug, is one of the photosensitizers used in PDT. Recently, we reported a significant difference in response to 5-ALA-mediated PDT treatment in two canine primary lung adenocarcinoma cell lines (sensitive to PDT: HDC cells, resistant to PDT: LuBi cells). This study aimed to examine the difference in cytotoxicity of 5-ALA-mediated PDT in these cells. Although intracellular PpIX levels before irradiation were similar between HDC and LuBi cells, the percentage of ROS-positive cells and apoptotic cells in LuBi cells treated with 5-ALA-mediated PDT was significantly lower than that in HDC cells treated with 5-ALA-mediated PDT. A high dosage of the NO donor, DETA NONOate, significantly increased the cytotoxicity of 5-ALA-mediated PDT against LuBi cells. These results suggest that the sensitivity of 5-ALA-mediated PDT might be correlated with NO.

5-Aminolevulinic acid, a natural amino acid, activates mitochondrial respiration and induces heme oxygenase-1 expression. Obesity and type 2 diabetes mellitus are associated with age-related mitochondrial respiration defect, oxidative stress and inflammation. The aim of this study is to investigate the effects of 5-aminolevulinic acid with sodium ferrous citrate on early renal damage and hepatic steatosis. 7-Month-old C57BL/6 mice were fed with a standard diet or high fat diet for 9 weeks, which were orally administered 300 mg/kg 5-aminolevulinic acid combined with 47 mg/kg sodium ferrous citrate (5-aminolevulinic acid/sodium ferrous citrate) or vehicle for the last 5 weeks. We observed that 5-aminolevulinic acid/sodium ferrous citrate significantly decreased body weight, fat weight, hepatic lipid deposits and improved levels of blood glucose and oral glucose tolerance test. In addition, 5-aminolevulinic acid/sodium ferrous citrate suppressed increased glomerular tuft area in high fat diet-fed mice, which was associated with increased heme oxygenase-1 protein expression. Our findings demonstrate additional evidence that 5-aminolevulinic acid/sodium ferrous citrate could improve glucose and lipid metabolism in diabetic mice. 5-Aminolevulinic acid/sodium ferrous citrate has potential application in obesity or type 2 diabetes mellitus-associated disease such as diabetic nephropathy and nonalcoholic fatty liver disease.

Decreased mitochondrial metabolism suppresses glucose metabolism, resulting in obesity and diabetes. The present study aimed to investigate mechanisms underlying the 5-aminolevulinic acid (5-ALA) hydrochloride-mediated increase in glucose uptake in high-fat diet (HFD)-fed mice in vivo and C2C12 myotube cells in vitro. C57BL/6N male mice (20 weeks old) were fed either HFD or normal diet (ND) for 4 weeks. A total of five HFD-fed mice were orally administered with 300 mg/kg 5-ALA hydrochloride and 47.1 mg/kg sodium ferrous citrate (SFC; HFD + 5-ALA/SFC), whereas ND and other HFD-fed mice were orally administered with saline. After 4 weeks, these mice were intraperitoneally administered with 2 g/kg glucose and 3.2 mg/kg 2-deoxyglucose (2DG) for intraperitoneal glucose tolerance test (IPGTT) and glucose uptake test. Body weights, plasma glucose levels and the area under the curve of IPGTT were lower in mice treated with HFD + 5-ALA/SFC compared with in those treated with HFD alone. 2DG uptake in the gastrocnemius muscle and heart were more significantly improved in the HFD + 5-ALA/SFC mice compared with the HFD-fed mice. Furthermore, 5-ALA/SFC increased 2DG uptake in C2C12 cells to a similar level to the insulin-treated group. Moreover, it increased glucose transport (GLUT)1 translocation in the plasma membrane by 2.5-fold relative to the controls without affecting GLUT1 expression; however, it had no effect on GLUT4 translocation. Therefore, 5-ALA/SFC enhanced gastrocnemius and cardiac glucose uptake in HFD-fed mice, and upregulated GLUT1 translocation to the plasma membrane, but not GLUT4 in C2C12 myotube cells. Therefore, it could potentially be used as a novel drug for the treatment of diabetes.

Background There are few agents that have been proven effective for COVID-19. Predicting clinical improvement as well as mortality or severity is very important. Objectives This study aimed to investigate the factors associated with the clinical improvement of COVID-19. Methods Overall, 74 patients receiving treatment for COVID-19 at Tokyo Medical and Dental University Hospital from April 6th to May 15th, 2020 were included in this study. Clinical improvement was evaluated, which defined as the decline of two levels on a six-point ordinal scale of clinical status or discharge alive from the hospital within 28 days after admission. The clinical courses were particularly investigated and the factors related to time to clinical improvement were analyzed with the log-rank test and the Cox proportional hazard model. Results Forty-nine patients required oxygen support during hospitalization, 22 patients required invasive mechanical ventilation, and 5 patients required extracorporeal membrane oxygenation. A total of 83% of cases reached clinical improvement. Longer period of time from onset to admission (≥10 days) (HR, 1.057; 95% CI, 1.002–1.114), no hypertension (HR, 2.077; 95% CI, 1.006–4.287), and low D-dimer levels (

Background 5-Aminolaevulinic acid (5ALA) is a precursor of the strong sensitizer protoporphyrin IX (PpIX) in the heme synthesis pathway. We conducted aclinical trial designed to evaluate the efficacy and safety of 5ALA photodynamic therapy (PDT) using a light-emitting diode (LED) in patients with cervical intraepithelial neoplasia (CIN). Methods Data for 51 CIN patients who underwent 5ALA-PDT between 2012 and 2017 were prospectively analysed. After a 20 % 5ALA jelly formulation was topically applied to the cervix, the region was irradiated with red light at approximately 633 nm to excite PpIX for treatment. We estimated outcomes by cytology, pathology, and human papilloma virus (HPV) testing after PDT. Results Patients underwent two PDT sessions at one-week intervals during outpatient treatment and achieved favourable results without photosensitivity and severe adverse events. Over a long follow-up period, 96.1 % of all patients showed some positive effects, including approximately 70 % with a complete response (CR), 10 % with a partial response, and 15 % with downgrades. The HPV clearance rate in patients with CR was 79.4 %. Recurrence occurred in five patients who mostly remained HPV-positive after PDT. Conclusions Based on our study, topical 5ALA-PDT using an LED light source potentially represents a safe treatment for CIN on an outpatient basis.

SMARCA4-deficient thoracic sarcoma is a rare tumor typically presenting as a mediastinal mass. The prognosis is estimated to be poor, and no effective treatment has been established. We present a case of a 76-year-old man who was diagnosed with SMARCA4-deficient thoracic sarcoma. The provisional diagnosis was carcinoma of unknown primary but subsequently corrected to SMARCA4-deficient thoracic sarcoma based on the panel-based cancer gene screening and immunohistochemistry. Cytotoxic chemotherapy as the first- and second-line did not reveal enough therapeutic effects but third-line therapy using nivolumab showed marked tumor regression, which was sustained. This is the first case report of SMARCA4-deficient thoracic sarcoma showing a good response to nivolumab. Immune checkpoint inhibitor might be therapeutic candidates for this type of tumor.

Background Bird-related hypersensitivity pneumonitis (BRHP) is the most common type of fibrotic hypersensitivity pneumonitis (HP). Antigen avoidance (AA) is a key step in the diagnosis and management of HP, but not all fibrotic HP patients improve through AA. Because of the poor prognosis of fibrotic HP, predictive biomarkers to identify patients showing rapid progression during AA are urgently needed. Methods From a retrospective review of the medical records of 1941 patients with interstitial lung disease, 75 cases of fibrotic BRHP confirmed by a provocation test or surgical lung biopsy were identified. To identify potential prognostic markers obtained at or around diagnosis, physiological and serological variables at diagnosis and the relative changes in those variables during strict AA were evaluated. Cox proportional hazards models with log-rank testing were used to compare the associations between these variables and survival. Results Univariate analyses showed that gender, smoking status, and the relative change in the serum levels of Krebs von den Lungen-6 (KL-6) were associated with prognosis (P = 0.02, 0.04, and 0.02, respectively), but the presence of honeycombing and the forced vital capacity were not associated with survival. The relative change in KL-6 levels (greater than vs. less than a 10% decrease) was significantly associated with survival in a stratified analysis (73.9 vs. 34.9 months; P = 0.04). Conclusions The relative change in KL-6 levels is associated with the prognosis of patients with fibrotic BRHP independent of previously identified prognostic biomarkers. This finding could help pulmonologists identify fibrotic BRHP patients that are likely to show rapid progression.

Background Photodynamic therapy (PDT) is known as a minimally invasive treatment for cancer. 5-Aminolevulinic acid (ALA) is a precursor of the photosensitizing agent protoporphyrin IX (PpIX). Patients with ovarian clear-cell carcinoma (CCC) have poorer prognoses than those of patients with other histological CCC types. We evaluated the efficacy of ALA-PDT on CCC cells in vitro. Methods We used seven human CCC cell lines to measure the cytotoxicity of ALA-PDT. PpIX production in cancer cells was measured using a micro-plate reader. Quantitative real-time PCR was performed to assess the mRNA levels of genes involved in the accumulation of PpIX in cancer cells. Additionally, we measured the enhancement in cytotoxicity with the use of an ABCG2 inhibitor. Results We found that three cell lines were highly sensitive to ALA-PDT. In contrast, one cell line was resistant to ALA-PDT. The cytotoxicity of ALA-PDT varied among CCC cell lines. The IC50 values of ALA-PDT for the CCC cell lines had a wide range (30–882 μM). The cytotoxicity of ALA-PDT was correlated with the intracellular PpIX accumulation. The cell lines sensitive to ALA-PDT expressed PEPT1 (an ALA uptake transporter). The cell line resistant to ALA-PDT expressed ABCG2 (a PpIX export transporter). In the resistant cell line, a combination treatment with both ALA and an ABCG2 inhibitor resulted in the promotion of cytotoxic sensitivity. Conclusion The present study revealed the efficacy of ALA-PDT against CCC with chemoresistance in vitro.

5-aminolevulinic acid (5-ALA) has recently been employed for photodynamic diagnosis (ALA-PDD) and photodynamic therapy (ALA-PDT) of various types of cancer because hyperproliferating tumor cells do not utilize oxidative phosphorylation and do not efficiently produce heme; instead, they accumulate protoporphyrin IX (PpIX), which is a precursor of heme that is activated by violet light irradiation that results in the production of red fluorescence and singlet oxygen. The efficiencies of ALA-PDD and ALA-PDT depend on the efficient cellular uptake of 5-ALA and the inefficient excretion of PpIX. We employed the JFCR39 cell panel to determine whether tumor cells originating from different tissues can produce and accumulate PpIX. We also investigated cellular factors/molecules involved in PpIX excretion by tumor cells with the JFCR39 cell panel. Unexpectedly, the expression levels of ABCG2, which has been considered to play a major role in PpIX extracellular transport, did not show a strong correlation with PpIX excretion levels in the JFCR39 cell panel, although an ABCG2 inhibitor significantly increased intracellular PpIX accumulation in several tumor cell lines. In contrast, the expression levels of dynamin 2, which is a cell membrane-associated molecule involved in exocytosis, were correlated with the PpIX excretion levels. Moreover, inhibitors of dynamin significantly suppressed PpIX excretion and increased the intracellular levels of PpIX. This is the first report demonstrating the causal relationship between dynamin 2 expression and PpIX excretion in tumor cells.

5-Aminolevulinic acid (5-ALA), a commonly used photosensitizer in photodynamic detection (PDD) and therapy (PDT), is converted in situ to the established photosensitizer protoporphyrin IX (PpIX) via the heme biosynthetic pathway. To extend 5-ALA-PDT application, we evaluated the PpIX fluorescence induced by exogenous 5-ALA in various veterinary tumors and treated canine and feline tumors. 5-ALA-PDD sensitivity and specificity in the whole sample group for dogs and cats combined were 89.5 and 50%, respectively. Notably, some small tumors disappeared upon 5-ALA-PDT. Although single PDT application was not curative, repeated PDT+/&minus, chemotherapy achieved long-term tumor control. We analyzed the relationship between intracellular PpIX concentration and 5-ALA-PDT in vitro cytotoxicity using various primary tumor cells and determined the correlation between intracellular PpIX concentration and 5-ALA transporter and metabolic enzyme mRNA expression levels. 5-ALA-PDT cytotoxicity in vitro correlated with intracellular PpIX concentration in carcinomas. Ferrochelatase mRNA expression levels strongly negatively correlated with PpIX accumulation, representing the first report of a correlation between mRNA expression related to PpIX accumulation and PpIX concentration in canine tumor cells. Our findings suggested that the results of 5-ALA-PDD might be predictive for 5-ALA-PDT therapeutic effects for carcinomas, with 5-ALA-PDT plus chemotherapy potentially increasing the probability of tumor control in veterinary medicine.

Objectives The fluorescent dye, 5-aminolevulinic acid (5-ALA), is currently applied for fluorescence-guided resections of high-grade gliomas. Present limitations of this technique are qualitative and subjective analyses, which show little of the background structures. This paper describes the intraoperative quantitative analysis of fluorescence intensity, hot-spot enhancement by frame averaging, and observation of surrounding structures by using 1000-nm lighting in real time. Patients and methods A sample of diluted protoporphyrin IX (PpIX) in a bottle and 37 samples from nine patients with brain lesions were involved in this study. In this preliminary study, we determined appropriate conditions for image averaging and filters and selected the most sensitive spectrometer. In addition, we utilized a 1000-nm lighting system to visualize surrounding structures with no interference from PpIX fluorescence. Results The novel system permitted the real-time quantitative analysis of PpIX fluorescence in operative fields by illuminating structures with 1000-nm-lighting. The real-time quantification provided subjective evaluations for surgical decision-making. We found good correlations between the fluorescence and PpIX contents in brain tissue. Furthermore, 1000-nm lighting visualized the anatomical structures and PpIX fluorescence simultaneously. Conclusion The combination of spectroscopy and a 1000-nm lighting system could enable surgeons to create a spectrogram of targets of interest while observing background structures. The spectrometer that we selected is highly sensitive to PpIX fluorescence and enables us to perform intraoperative real-time tissue mapping. By using a real-time system, we can perform quantitative and objective evaluations to achieve maximal tumor resection.

Background Hypersensitivity pneumonitis (HP) is an interstitial lung disease caused by the inhalation of environmental antigens. The relationship between clinical, radiologic, and histopathologic findings of chronic HP remains unclear. Methods Sixteen patients with proven chronic bird-related HP with a usual interstitial pneumonia-like pattern were analyzed retrospectively. Histopathologic findings were semiquantitatively assessed and compared with clinical and radiologic findings. We also evaluated the histopathologic findings affecting prognosis. Results The extent of centrilobular fibrosis was negatively correlated with Pa o 2 ( r = –0.55, P = .03). The extent of bridging fibrosis was positively correlated with the ratio of maximal expiratory flow at 50% of forced vital capacity to that at 25% ( r = 0.60, P = .02). Patients with a greater extent of fibroblastic foci (FF) had more radiologic reticulation ( P = .01), honeycombing ( P = .01), and traction bronchiectasis ( P = .02), and had significantly shorter survival time ( P = .01) than patients with a lesser extent of FF. Multivariate analysis showed that the extent of FF was a significant prognostic factor (hazard ratio, 2.36; 95% confidence interval, 1.02-5.48; P = .04). Conclusions Our findings demonstrated that the extent of FF was significantly associated with reticulation, honeycombing, and traction bronchiectasis on high-resolution CT scanning. Moreover, the extent of FF could be a useful predictor of mortality in chronic HP with a usual interstitial pneumonia-like pattern.

Background Rheumatoid arthritis-associated interstitial lung disease (RA-ILD) occurs in 10%–30% of patients with RA, and interstitial lung disease (ILD) is associated with increased mortality in up to 10% of patients with RA. The pathogenesis of RA-ILD is virtually unknown. The aim of this study is to investigate the proteins related to UIP pattern by comparing to OP pattern in RA-ILD using proteome analysis of bronchoalveolar lavage fluid (BALF). Methods Proteomic differences in BALF were compared between the UIP pattern and OP pattern by examining BALF from 5 patients with the UIP pattern and 7 patients with the OP pattern by two-dimensional gel electrophoresis and mass spectrometry. Results In individual comparisons of BALF samples, the levels of the protein gelsolin and Ig kappa chain C region were significantly higher in the UIP pattern than in the OP pattern. In contrast, the levels of α-1 antitrypsin, CRP, haptoglobin β, and surfactant protein A (isoform number 5) were all significantly higher in the OP pattern than in the UIP pattern. Gelsolin was cleaved into two fragments, a C-terminal half and N-terminal half, and the levels of both were significantly higher in the UIP pattern than in the OP pattern. Conclusions Fragmented gelsolins may be associated with the pathogenesis of fibrosis in RA-ILD.

Background Photodynamic therapy (PDT) and diagnosis (PDD) using 5-aminolevulinic acid (ALA) to control the production of the intracellular photosensitizer protoporphyrin IX (PpIX) are commonly used clinically. Previously, we demonstrated that dormant and drug-induced dormancy-like cancer cells accumulated high PpIX levels, making them sensitive to ALA-PDT. Because EAU Guidelines awarded a level of evidence of 1a to mitomycin C, the drug is widely used to treat bladder cancer. In this study, we investigated that the effect of mitomycin C-induced cell cycle arrest on porphyrin metabolism, including that induced by ALA-PDT. Methods T24 human urinary bladder carcinoma cells were selected for this research. T24 cells were irradiated using a light-emitting diode emitting red light for the ALA-PDT assay. Cell cycle analysis was conducted by flow cytometry using bromodeoxyuridine. Cell viability was confirmed using the MTT or colony formation assay. Furthermore, mRNA gene expression analysis was performed using our previously reported methods. Results The cell cycle of T24 cells was arrested at G2/M phase by mitomycin C. PpIX accumulation was dramatically increased by mitomycin C treatment. Cell viability after ALA-PDT was remarkably decreased by mitomycin C pretreatment. The gene expression of porphyrin transporters was consistent with the metabolic and morphological results. Finally, we confirmed that ALA-PDT combined with mitomycin C treatment exerted a long-term inhibitory effect on cell proliferation. Conclusion This study demonstrated a new approach to enhance the effects of ALA-PDT using drugs that induce a dormancy-like status and upregulate porphyrin metabolism.

Background: Hypersensitivity pneumonitis (HP) is an immune-mediated lung disease induced by the inhalation of a wide variety of antigens and a persistent antigen exposure induces inevitably pulmonary fibrosis in chronic HP. Although neutrophils, Th1 and Th17 cells contribute to lung inflammation in acute phase of HP, there is no clear explanation as to how the immunological reaction occurs just after the inhalation of causative antigens in the chronic phase of HP. Methods: We examined the inflammatory and immunologic profiles before and after the inhalation provocation test (IPT) in serum and bronchoalveolar lavage fluid (BALF) from patients with chronic bird-related HP (BRHP) and other interstitial lung diseases (ILDs). We analyzed BALF samples from 39 patients (19 BRHP and 20 other ILDs) and serum samples from 25 consecutive patients (20 BRHP and 5 other ILDs) who underwent the IPT. Results: A significant increase of neutrophils was observed in the BALF from the BRHP patients following the IPT. Neutrophil chemoattractants, namely, granulocyte colony-stimulating factor, IL-6, IL-8, IL-17, and CXCL2 significantly increased in both the serum and BALF of the BRHP patients after the IPT. Serum IFN-γ and CXCL10, cytokines/chemokines that contributed to Th1 inflammation, were also significantly increased in BRHP following the IPT. Conclusions: This study demonstrated the exposure to the causative antigen provoked acute neutrophilic and Th1 immunologic responses similar to acute HP even in the chronic phase of HP. Keywords: Bird-related hypersensitivity pneumonitis, Chemokine, Cytokine, Inhalation provocation test, Neutrophil

Exogenously administered 5-aminolevulinic acid (ALA) is metabolized to protoporphyrin IX (PpIX), which specifically accumulates in cancer cells and emits red fluorescence by blue light irradiation. These phenomena are applied for the intraoperative diagnosis of cancer. Based on the fact that accumulated PpIX in cancer cells is exported extracellularly via the ATP-binding cassette transporter G2, we hypothesized that the measurement of plasma PpIX concentrations could be applied as a tumor marker for cancer screening. In the present study, the use of plasma samples from bladder cancer patients were evaluated as a tumor marker. ALA, 1.0 g, was orally administered to bladder cancer patients and healthy adults. The plasma concentration of PpIX was measured using a high-performance liquid chromatography system. The plasma PpIX concentration following ALA administration was significantly higher in bladder cancer patients than that in the healthy adults, suggesting the effectiveness of plasma PpIX analysis following ALA administration for cancer screening. Additionally, 4 h after ALA administration, plasma PpIX showed high sensitivity (94.4%) and high specificity (80.0%).

Summary Background Currently, 5-aminolevulinic acid-based photodynamic diagnosis (ALA-PDD) is used to detect tumors during surgery and exploit tumor-specific accumulation of protoporphyrin IX (PpIX) after administration of ALA. In a recent study, we showed that the human ATP-binding cassette transporter ABCG2 plays a key role in the regulation of PpIX as a specific exporter. However, coproporphyrin III (CPIII) was also detected in urine after ALA administration in patients with tumor, indicating the presence of a CPIII transporter. Methods We used two lines of human gastric cancer cells to measure the ALA-induced porphyrin metabolism. Intracellular and extracellular porphyrin levels and expressions of transporter were determined. Results In the present study, we showed that although ABCG2 did not transport CPIII, plasma membrane ABCB6 did. Moreover, under conditions of hypoxia, the expression of ABCB6 in plasma membrane was upregulated, resulting in increased extracellular CPIII concentrations. Conclusion These data indicate that the expression of ABCB6 in plasma membrane is important for porphyrin accumulation after ALA administration, including hypoxic conditions.

Hypersensitivity pneumonitis (HP) is an immunologically mediated lung disease induced by the inhalation of any of a wide variety of antigens. For example, bird-related HP (BRHP) results from the inhalation of avian antigens. The clinical features of chronic HP, including imaging and histological findings, are similar to those of idiopathic pulmonary fibrosis. Despite its status as the "gold standard," the inhalation provocation test (IPT) is rarely performed, because the methods and the criteria are not standardized. In 2000, we reported the utility of IPT for pigeon dropping extracts.The purpose of the current study was to validate the utility and safety of the test, and to differentiate chronic HP from other interstitial lung diseases.A total of 28 patients with chronic BRHP and 19 control subjects were evaluated in this retrospective study. We validated the previous criteria and proposed new criteria using prediction scores.In the IPT using pigeon dropping extracts, the previous criteria showed a sensitivity of 78.6% and a specificity of 94.7% in this retrospective study. The increases in the peripheral white blood cell count and C-reactive protein levels are good indicators of a positive response to the inhalation challenge. We propose the use of the IPT prediction score (∆WBC [%] + 2 × ∆P[a - a]O2 [mm Hg], where WBC is white blood cell) and the prediction rule, which showed high sensitivity and specificity values of 92.9 and 94.7%, respectively. Two (1.5%) out of a total of 130 subjects who underwent the tests required treatment after the challenge.The IPT is a useful and safe tool for the diagnosis of chronic HP. The IPT prediction score that we have proposed has high sensitivity and specificity in the diagnosis of chronic BRHP.

5-Aminolevulinic acid (ALA), a precursor of protoporphyrin IX (PpIX), is now widely used for photodynamic diagnosis (ALA-PDD) and photodynamic therapy (ALA-PDT) of various cancers. Recently, we found that treatment of cancer cells with the Schiff base derivative TX-816 along with ALA could significantly increase the efficacy of ALA-PDT. This enhancing effect of TX-816 on ALA-PDT is attributed to 3,5-dichlorosalicylaldehyde (DCSA), a molecule produced by the degradation of TX-816. Similar to TX-816, DCSA significantly enhances the effect of ALA-PDT. Furthermore, DCSA could restore the sensitivity of cancer cells that acquired resistance to ALA-PDT. These results indicate that DCSA, as well as TX-816, is a potent lead compound for the development of an ALA-PDT sensitizer. TX-816 might be a useful compound for designing prodrug-type ALA-PDT enhancers.

Background/Aim: Previous studies on the accuracy of 5-aminolevulinic-acid-mediated photodynamic diagnosis (5-ALA PDD) have been reported for various cancers and brain surgery. However, biliary tract cancer is rare. Therefore, 5-ALA PDD has not been fully evaluated in biliary tract cancers. Small biliary tract cancer lesions such as peritoneal dissemination, liver metastases, and lymph node metastases are negative prognosticators in patients with biliary cancer. The purpose of this exploratory study was to determine if 5-ALA PDD could detect small biliary tract cancer lesions in murine models of biliary cancers. Materials and Methods: Biliary cancer cell lines (TFK-1, HuCCT-1, G415, HuH28, SSP25, RBE, KKU055 and KKU100) and Normal human dermal fibroblast cells were used to evaluate protoporphyrin IX (PpIX) accumulation in vitro. Subcutaneous tumor mice were established using two cell lines (TFK-1 and HuCCT-1). 5-ALA (250 mg/kg) was administered intraperitoneally, and fluorescent 5ALA-PDD was performed 3 h later to evaluate tumoral PpIX accumulation. A murine peritoneal disseminated nodule model was established by intraperitoneal injection of TFK-1 cells. Four weeks later, 5-ALA was administered intraperitoneally, and 5-ALA-PDD was performed 3 h post administration to evaluate PpIX accumulation in the disseminated nodules. The presence of tumor cells in tumors and nodules was confirmed by haematoxylin and eosin staining. Results: Compared TO non-cancerous cell lines, PpIX accumulation was increased in biliary tract cancer cell lines. PpIX accumulation led to a strong fluorescent signal in all subcutaneous tumors. In the murine model of peritoneal dissemination, microdisseminated nodules (

The biguanide metformin is a drug widely used for the treatment of type 2 diabetes. Metformin enhances the cytotoxicity of chemotherapy by promoting the adenosine monophosphate-activated protein kinase (AMPK) autophagy signaling pathway. Photodynamic therapy (PDT) with 5-aminolevulinic acid (5-ALA), a precursor of protoporphyrin IX (PpIX), leads to apoptosis when PpIX accumulates in the mitochondria, and also leads to autophagy through activation of AMPK. In the present study, the effect of metformin in combination with 5-ALA-PDT was evaluated in vitro in KLN205 lung cancer cells. At a fluence of 5 J/cm2, 5-ALA-PDT in combination with 5 mM metformin exhibited significantly increased cytotoxicity compared with that observed with 0 and 0.1 mM metformin (P=0.0197 and P=0.0423, respectively). The cells treated with 5-ALA-PDT and metformin exhibited condensation of nuclear chromatin and the presence of autophagosomes. These results indicate that apoptosis and autophagy occur in KLN205 cells following combined treatment with 5-ALA-PDT and metformin. The results from the present study are the first to indicate, to the best of our knowledge, that metformin potentiates the efficacy of 5-ALA-PDT.

5-Aminolevulinic acid (ALA) is a precursor for the biosynthesis of porphyrins and heme. Although the oral administration of ALA has been widely applied in clinical settings, the dynamics of its absorption, metabolism, and excretion within enterocytes remain unknown. In this study, after enterocytic differentiation, Caco-2 cells were incubated with 200 µM ALA and/or 100 µM sodium ferrous citrate (SFC) for up to 72 h. Both ALA and the combination of ALA and SFC promoted the synthesis of heme, without affecting the expression of genes involved in intestinal iron transport, such as DMT1 and FPN. The enhanced heme synthesis in Caco-2 cells was more pronounced under the effect of the combination of ALA and SFC than under the effect of ALA alone, as reflected by the induced expression of heme oxygenase 1 (HO-1), as well as a reduced protein level of the transcriptional corepressor Bach1. Chromatin immunoprecipitation analysis confirmed Bach1 chromatin occupancy at the enhancer regions of HO-1, which were significantly decreased by the addition of ALA and SFC. Finally, Transwell culture of Caco-2 cells suggested that the administered ALA to the intestinal lumen was partially transported into vasolateral space. These findings enhance our understanding of the absorption and metabolism of ALA in enterocytes, which could aid in the development of a treatment strategy for various conditions such as anemia., Highlights • Combination of ALA and SFC promotes heme synthesis than ALA alone in Caco-2 cells. • Heme induces HO-1 by inhibiting transcriptional corepressor Bach1 in Caco-2 cells. • Addition of ALA to intestinal lumen was partially transported to vasolateral space. • Our data enhance the understanding of the dynamics of ALA in enterocytes.