The Potential of Narrow-Band Imaging as a Novel Light Source for Photodynamic Therapy for Superficial Cancers via Endoscopes.

Background: Photodynamic therapy (PDT) has advanced through the utilization of photosensitizers and specific-wavelength light (≥ 600 nm). However, the widespread adoption of PDT is still impeded by high equipment costs and stringent laser safety requirements. Porphyrins, crucial in PDT, have another absorbance peak of blue light (λ = 380 - 500 nm). This peak corresponds to the wavelength of narrow-band imaging (NBI) (λ = 390 - 445 nm), an image-enhancement technology integrated into endoscopes by Olympus Medical Systems. The study aimed to investigate the potential of widely adopted NBI as a PDT light source for superficial cancers via endoscopes. Methods: Esophageal and biliary cancers were selected for investigation. Human esophageal cancer cell lines (KYSE30, KYSE70, KYSE170) and cholangiocarcinoma cell lines (HuCCT-1, KKU-213) were subjected to verteporfin-mediated PDT under NBI light (λ = 390 - 445 nm). Assessments included spectrometry, crystal violet staining, and fluorescein imaging of singlet oxygen generation and apoptosis. Results: Verteporfin exhibited a peak (λ = 436 nm) consistent with the NBI spectrum, suggesting compatibility with NBI light. NBI light significantly inhibited the growth of esophageal and biliary cancer cells. The half-maximum effective concentration (EC50) values (5 J/cm2) for KYSE30, KYSE70, KYSE170, HuCCT-1, and KKU-213 were calculated as 2.78 ± 0.37µM, 1.76 ± 1.20 µM, 0.77 ± 0.16 µM, 0.65 ± 0.18 µM, and 0.32 ± 0.04 µM, respectively. Verteporfin accumulation in mitochondria, coupled with singlet oxygen generation and observed apoptotic changes, suggests effective PDT under NBI light. Conclusions: NBI is a promising PDT light source for superficial cancers via endoscopes.

Docking Proteins Upregulate IL-1ß Expression in Lower Esophageal Sphincter Muscle in Esophageal Achalasia.

Background/Objectives: Esophageal achalasia is an archetypal esophageal motility disorder characterized by abnormal peristalsis of the esophageal body and impaired lower esophageal sphincter (LES) relaxation. Methods: In this study, the mRNA expression of docking proteins 1 and 2 (DOK1 and DOK2, respectively) were analyzed and the mechanisms underlying achalasia onset were investigated. Results:DOK1 and DOK2 mRNA levels significantly increased in the LES of patients with achalasia. Moreover, significant correlations were observed between IL-1ß and DOK1, IL-1ß and DOK2, ATG16L1 and DOK1, and HSV1-miR-H1-3p and DOK2 expression levels. However, a correlation between ATG16L1 and DOK2 or between HSV-miR-H1-3p and DOK1 expression was not observed. In addition, a positive correlation was observed between patient age and DOK1 expression. Microarray analysis revealed a significant decrease in the expression of hsa-miR-377-3p and miR-376a-3p in the LES muscle of patients with achalasia. Conclusions: These miRNAs possessed sequences targeting DOK. The upregulation of DOK1 and DOK2 expression induces IL-1ß expression in the LES of achalasia patients, which may contribute to the development of esophageal motility disorder.

Zinc deficiency induces hypertension by paradoxically amplifying salt sensitivity under high salt intake in mice.

BACKGROUND: Hypertension is one of the major etiologies that cause chronic kidney disease (CKD) and can exacerbate kidney dysfunction. Zinc is an essential trace element playing a role in blood pressure regulation, and zinc deficiency, a common comorbidity in patients with CKD, can cause hypertension. However, the precise mechanism underlying zinc deficiency-induced hypertension is unknown. Sodium (Na+) retention due to inappropriate Na+ reabsorption in the renal tubule is the principal pathophysiology of hypertension. Therefore, this study aimed to investigate the association between zinc deficiency and salt sensitivity. METHODS: Adult mice were fed a zinc-adequate (ZnA) or zinc-deficient (ZnD) diet combined with/without high salt in drinking water (HS) for 4 weeks (n = 6 each). Changes in blood pressure, urinary sodium excretion, and the expressions of the proximal tubular Na+ transporter, Na+/H+ exchanger 3 (NHE3), which mostly contributes to filtered Na+ reabsorption and the downstream Na+-Cl- transporter (NCC) were analyzed. RESULTS: Urinary Na+ excretion significantly increased in ZnD mice, indicating that zinc deficiency causes natriuresis. NHE3 expressions were significantly suppressed, whereas NCC was upregulated in ZnD mice. Interestingly, the combination of high salt and ZnD diet (HS-ZnD) reversed the urinary Na+ loss. The NCC remained activated and NHE3 expressions paradoxically increased in HS-ZnD mice compared with those fed the combination of high salt and ZnA diet. In addition, blood pressure significantly increased only in HS-ZnD mice. CONCLUSION: The combination of zinc deficiency and high salt causes hypertension. Zinc is associated with salt-sensitivity, potentially through NHE3 and NCC regulation.

ATG16L1 and ATG12 Gene Polymorphisms Are Involved in the Progression of Atrophic Gastritis.

Helicobacter pylori (H. pylori) infection causes a progression to atrophic gastritis and results in gastric cancer. Cytotoxin-associated gene A (CagA), a major virulence factor of H. pylori, is injected into gastric epithelial cells using the type IV secretion system. On the other hand, gastric epithelial cells degrade CagA using an autophagy system, which is strictly regulated by the autophagy-related (ATG) genes. This study aimed to identify SNPs in ATG5, ATG10, ATG12, and ATG16L1 associated with gastric mucosal atrophy (GMA). Here, two-hundred H. pylori-positive participants without gastric cancer were included. The degree of GMA was evaluated via the pepsinogen method. Twenty-five SNPs located in the four candidate genes were selected as tag SNPs. The frequency of each SNP between the GMA and the non-GMA group was evaluated. The rs6431655, rs6431659, and rs4663136 in ATG16L1 and rs26537 in ATG12 were independently associated with GMA. Of these four SNPs, the G/G genotype of rs6431659 in ATG16L1 has the highest odd ratio (Odds ratio = 3.835, 95% confidence intervals = 1.337-1.005, p = 0.008). Further functional analyses and prospective analyses with a larger sample size are required.

Polymorphism in autophagy-related genes LRP1 and CAPZA1 may promote gastric mucosal atrophy.

BACKGROUND: Helicobacter pylori secretes cytotoxin-associated gene A (CagA) into the gastric epithelium, causing gastric mucosal atrophy (GMA) and gastric cancer. In contrast, host cells degrade CagA via autophagy. However, the association between polymorphisms in autophagy-related genes and GMA must be fully elucidated. RESULTS: We evaluated the association between single nucleotide polymorphisms (SNPs) in autophagy-related genes (low-density lipoprotein receptor-related protein 1, LRP1; capping actin protein of muscle Z-line alpha subunit 1, CAPAZ1; and lysosomal-associated membrane protein 1, LAMP1) and GMA in 200 H. pylori-positive individuals. The frequency of the T/T genotype at rs1800137 in LRP1 was significantly lower in the GMA group than in the non-GMA group (p = 0.018, odds ratio [OR] = 0.188). The frequencies of the G/A or A/A genotype at rs4423118 and T/A or A/A genotype at rs58618380 of CAPAZ1 in the GMA group were significantly higher than those in the non-GMA group (p = 0.029 and p = 0.027, respectively). Multivariate analysis revealed that C/C or C/T genotype at rs1800137, T/A or A/A genotype at rs58618380, and age were independent risk factors for GMA (p = 0.038, p = 0.023, and p = 0.006, respectively). Furthermore, individuals with the rs1800137 C/C or C/T genotype of LRP1 had a 5.3-fold higher susceptibility to GMA. These genetic tests may provide future directions for precision medicine for individuals more likely to develop GMA. CONCLUSION: LRP1 and CAPZA1 polymorphisms may be associated with the development of GMA.

Comparative study between histochemical mucus volume, histopathological findings, and endocytoscopic scores in patients with ulcerative colitis.

Ulcerative colitis (UC) causes a reduction in goblet cells. However, there have been few reports on the relationship between endoscopic and pathological findings and mucus volume. In this study, we quantitatively evaluated histochemical colonic mucus volume by fixing biopsied tissue sections taken from patients with UC in Carnoy's solution and compared it with endoscopic and pathological findings to determine whether there is a correlation between them. Observational study. A single-center, university hospital in Japan. Twenty-seven patients with UC (male/female, 16/11; mean age, 48.4 years; disease median duration, 9 years) were included in the study. The colonic mucosa of the most inflamed area and the surrounding less inflamed area were evaluated separately by local MES and endocytoscopic (EC) classification. Two biopsies were taken from each area; one was fixed with formalin for histopathological evaluation, and the other was fixed with Carnoy's solution for the quantitative evaluation of mucus via histochemical Periodic Acid Schiff and Alcian Blue staining. The relative mucus volume was significantly reduced in the local MES 1-3 groups, with worsening findings in EC-A/B/C and in groups with severe mucosal inflammation, crypt abscess, and severe reduction in goblet cells. The severity of inflammatory findings in UC by EC classification correlated with the relative mucus volume suggesting functional mucosal healing. We found a correlation between the colonic mucus volume and endoscopic and histopathological findings in patients with UC, and a stepwise correlation with disease severity, particularly in EC classification.

Serum Activin A Is a Novel Biomarker of Endoscopic Activity in Ulcerative Colitis.

Background: Endoscopic healing (EH) is the long-term therapeutic goal for ulcerative colitis (UC). Since repeated colonoscopies are inconvenient and invasive, a surrogate biomarker for endoscopic activity is needed. Activin A is one of the transforming growth factor-ß superfamily of proteins and has been shown to be associated with intestinal inflammation. Methods: This single-center observational study included 27 Japanese patients with UC in clinical remission who underwent colonoscopy and blood sampling. We investigated the correlations between laboratory parameters, including serum activin A levels, and endoscopic activity, classified by the Mayo endoscopic subscore (MES) in these patients. Results: This study included 15 males and 12 females. The median age was 44.0 years. In terms of endoscopic activity, five patients were diagnosed with MES 0, 14 patients with MES 1, seven patients with MES 2, and one patient with MES 3. The median serum activin level was 134.8 pg/mL (interquartile range (IQR), 105.3 - 188.1). Serum activin A levels were significantly correlated with the MES (Spearman's rank correlation coefficient r = 0.591, P = 0.001), which was better than that of C-reactive protein (CRP) (r = 0.487, P = 0.010). In the comparison between the EH group (MES 0) and non-EH group (MES 1-3), patients without EH had significantly higher serum activin A levels (Mann-Whitney U test, P = 0.047). A cutoff value of 133.6 pg/mL indicated non-EH with a sensitivity and specificity of 0.682 and 1.000, respectively. The area under the curve (AUC) of serum activin A for detecting non-EH was 0.791 (95% confidence interval (CI), 0.618 - 0.964), while that of CRP was 0.723 (95% CI, 0.504 - 0.941). Conclusions: The serum activin A level is a potential novel biomarker of endoscopic activity in UC.

Intracellular glutathione levels affect the outcomes of verteporfin-mediated photodynamic therapy in esophageal cancer cells.

Photodynamic therapy (PDT) induces cancer cell death by generating reactive oxygen species (ROS). In this process, photosensitizers accumulate in cancer cells irradiated by laser light of a specific wavelength, leading to ROS generation. Verteporfin (VP), a second-generation photosensitizer, is used in PDT for age-related macular degeneration. However, the antitumor effects of VP-PDT remain poorly defined. This study investigated the antitumor effects of VP-PDT on esophageal cancer (EC) cell lines in vitro. Two types of EC cell lines, the KYSE30 cell line, derived from highly differentiated esophageal carcinoma, and the KYSE170 cell line, derived from moderately differentiated carcinoma, were used in this study. VP-PDT exerted effective anticancer effects in both cell lines. Our results revealed that the low-density lipoprotein receptor, albumin receptor, and heme carrier protein-1 in VP uptake were not involved in VP uptake. However, cells rich in intracellular glutathione were resistant to VP-PDT. Our study outcomes suggest that lowering intracellular glutathione via a glutathione synthesis inhibitor or sulfasalazine can increase the effectiveness of VP-PDT-mediated anticancer effects.

SNP rs2920280 in PSCA Is Associated with Susceptibility to Gastric Mucosal Atrophy and Is a Promising Biomarker in Japanese Individuals with Helicobacter pylori Infection.

Helicobacter pylori infection results in gastric cancer (GC) with gastric mucosal atrophy (GMA). Some single-nucleotide polymorphisms (SNPs) in the prostate stem cell antigen gene (PSCA) are associated with GC and duodenal ulcers. However, the relationship of other identified SNPs in PSCA with these diseases remains unclear. Herein, the association between PSCA SNPs and GMA among 195 Japanese individuals with H. pylori infection was evaluated. The definition of GMA or non-GMA was based on serum pepsinogen levels or endoscopic findings. Five tag PSCA SNPs were analyzed using PCR high-resolution melting curve analysis with nonlabelled probes. The frequencies of alleles and the genotypes of each tag SNP were compared between the GMA and non-GMA groups. Subsequently, a genetic test was performed using associated SNPs as biomarkers to detect patients developing GMA. Two tag PSCA SNPs (rs2920280 and rs2294008) were related to GMA susceptibility. Individuals with the rs2920280 G/G genotype or the rs2294008 T/T genotype in PSCA had 3.5- and 2.1-fold susceptibility to GMA, respectively. In conclusion, SNP rs2920280 is a possible biomarker for detecting individuals developing GMA. PSCA polymorphisms may be useful biomarkers for predicting GMA linked to GC risk and a screening endoscopy strategy to detect GC related to early stage H. pylori associated GMA.

Objective Methods of 5-Aminolevulinic Acid-Based Endoscopic Photodynamic Diagnosis Using Artificial Intelligence for Identification of Gastric Tumors.

Positive diagnoses of gastric tumors from photodynamic diagnosis (PDD) images after the administration of 5-aminolevulinic acid are subjectively identified by expert endoscopists. Objective methods of tumor identification are needed to reduce potential misidentifications. We developed two methods to identify gastric tumors from PDD images. Method one was applied to segmented regions in the PDD endoscopic image to determine the region in LAB color space to be attributed to tumors using a multi-layer neural network. Method two aimed to diagnose tumors and determine regions in the PDD endoscopic image attributed to tumors using the convoluted neural network method. The efficiencies of diagnosing tumors were 77.8% (7/9) and 93.3% (14/15) for method one and method two, respectively. The efficiencies of determining tumor region defined as the ratio of the area were 35.7% (0.0-78.0) and 48.5% (3.0-89.1) for method one and method two, respectively. False-positive rates defined as the ratio of the area were 0.3% (0.0-2.0) and 3.8% (0.0-17.4) for method one and method two, respectively. Objective methods of determining tumor region in 5-aminolevulinic acid-based endoscopic PDD were developed by identifying regions in LAB color space attributed to tumors or by applying a method of convoluted neural network.

Investigating the role of heat shock protein 47 in fibrosis in Crohn's disease.

Crohn's disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract that share similar genetic risk factors. However, while fibrotic stricture of the intestine is a major characteristic of CD; it is rarely observed in UC. Deposition of collagen in the extracellular matrix contributes to the formation of fibrotic strictures in CD, but the underlying mechanisms are unknown. In the present study, we found that heat shock protein 47 (HSP47), a stress-response protein that acts as a molecular chaperone during the processing and secretion of collagen, expressed in the intestinal tissue from patients with CD. Serum HSP47 levels and anti-HSP47 antibody titers were significantly higher in patients with CD than in those with UC. Furthermore, anti-HSP47 antibody levels correlated significantly with fibrosis in CD. In addition, HSP47 inhibition significantly suppressed collagen production in fibroblasts in vitro. These findings suggest that HSP47 is a biomarker for differentiating fibrotic from non-fibrotic forms of CD. Additionally, we propose that HSP47 could be a potential target for treating fibrosis in patients with CD.

Steatosis is involved in the progression of kidney disease in a high-fat-diet-induced non-alcoholic steatohepatitis mouse model.

Chronic kidney disease (CKD) and non-alcoholic steatohepatitis (NASH) are major health issues associated with the metabolic syndrome. Although NASH is a known risk factor of CKD, the mechanisms linking these two diseases remain poorly understood. We aimed to investigate alterations in the kidney complicated with dyslipidemia in an established NASH mouse model. Male C57BL6/J mice were fed with control diet or high-fat diet (HFD), containing 40% fat, 22% fructose, and 2% cholesterol for 16 weeks. Metabolic characteristics, histological changes in the kidney, endoplasmic reticulum (ER) stress, apoptosis, and fibrosis were evaluated by histological analysis, immunoblotting, and quantitative reverse transcription-polymerase chain reaction. Levels of serum aspartate aminotransferase, alanine aminotransferase, alkali-phosphatase, total cholesterol, and urinary albumin were significantly higher in mice fed with HFD. Remarkable steatosis, glomerular hypertrophy, and interstitial fibrosis were also shown in in the kidney by leveraging HFD. Furthermore, HFD increased the mRNA expression levels of Casp3, Tgfb1, and Nfe2l2 and the protein level of BiP. We observed the early changes of CKD and speculate that the underlying mechanisms that link CKD and NASH are the induction of ER stress and apoptosis. Further, we observed the activation of Nfe2l2 in the steatosis-induced CKD mouse model. This NASH model holds implications in investigating the mechanisms linking dyslipidemia and CKD.

5-Aminolevulinic Acid Attenuates Glucose-Regulated Protein 78 Expression and Hepatocyte Lipoapoptosis via Heme Oxygenase-1 Induction.

Endoplasmic reticulum (ER) stress plays a pivotal role in the progression of steatohepatitis. 5-aminolevulinic acid (5-ALA), a precursor in the heme biosynthetic pathway, has recently been reported to induce heme oxygenase (HO)-1. HO-1 exerts important cytoprotective actions. In this study, we aimed to explore the therapeutic potential of 5-ALA on palmitate-induced ER stress and lipoapoptosis. Huh-7 cells were treated with palmitic acid (PA) (800 µM) to induce steatosis for eight hours. Steatosis was evaluated by Lipi-green staining. 5-ALA (200 µM) was added with PA. The gene expression levels of the nuclear factor erythroid 2-related factor 2 (NRF2), HO-1, Glucose-regulated protein 78 (GRP78), activating transcription factor 6 (ATF6), PKR-like endoplasmic reticulum kinase (PERK), inositol-requiring enzyme 1 (IRE1), C/EBP homologous protein (CHOP), and B-cell lymphoma 2 (BCL-2) were evaluated by RT-PCR. Caspase-3/7 activity was evaluated by fluorescein active Caspase-3/7 staining. Cell death was evaluated by Annexin V/SYTOX green staining. PA significantly induced steatosis and increased GRP78 expression in Huh-7 cells. 5-ALA significantly induced HO-1 and decreased GRP78 expression. ATF6 was subsequently decreased. However, NRF2 and CHOP expression were not altered. Anti-apoptotic BCL-2 expression significantly increased, and Caspase 3/7 activity and cell death also decreased. 5-ALA has a therapeutic potential on hepatic steatosis by suppressing ER stress and lipoapoptosis by attenuating GRP78 via HO-1 induction.

Current Status of Photodynamic Diagnosis for Gastric Tumors.

Although the recent development and widespread use of image-enhanced endoscopy and magnifying endoscopy have improved endoscopic diagnosis of gastric cancer, it is somewhat complicated, requires a higher level of expertise, and is still subjective. Photodynamic endoscopic diagnosis (PDED) is based on the fluorescence of photosensitizers that accumulate in tumors, which enables objective evaluation independent of the endoscopist's experience, and is useful for tumor detection. The objective of this work was to perform a narrative review of PDED for gastric tumors and to introduce our approach to PDED in gastric tumors in our hospital. In our review there have been case reports of PDED for gastric cancer, but its usefulness has not been established because no prospective studies evaluating its usefulness have been performed. In our previous study, 85.7% (42/49) of gastric tumors exhibited fluorescence in PDED. PDED may be useful in the diagnosis of early gastric cancer. Our previous studies were pilot studies in cancer patients; therefore, future prospective studies are required to verify the usefulness of PDED.

Photosensitizer verteporfin inhibits the growth of YAP- and TAZ-dominant gastric cancer cells by suppressing the anti-apoptotic protein Survivin in a light-independent manner.

Yes-associated protein (YAP) positivity indicates a poor prognosis in gastric cancer. Transcriptional co-activator with a PDZ-binding domain (TAZ), a YAP paralog, is highly expressed in gastric signet ring cell carcinoma. Verteporfin (VP), a clinical photosensitizer, was recently shown to inhibit YAP/TAZ. In the present study, the therapeutic potential of VP treatment was explored using two gastric cancer cell lines: MKN-45 (TAZ-dominant) and MKN-74 (YAP-dominant). Cell proliferation was evaluated by MTS assay. Vascular mimicry was evaluated by the tube formation assay. Gene and protein expression levels of YAP/TAZ downstream effectors [such as Survivin, Cysteine-rich angiogenic inducer 61 (CYR61), and connective tissue growth factor (CTGF)] were measured. YAP or TAZ localization was evaluated by immunofluorescence. Cell death was assessed by immunofluorescent staining of Annexin V. YAP and TAZ expression were knocked down by small interfering RNA. The current results demonstrate that MKN-45, a poorly differentiated TAZ-dominant gastric cancer cell line, was more sensitive to VP than MKN-74, a moderately differentiated YAP-dominant gastric cancer cell line. VP changed the localization of YAP/TAZ, promoted its degradation and significantly decreased the protein level of Survivin in both cell lines. Cell death was induced by VP treatment in a dose-dependent manner. Vascular mimicry was inhibited in both cell lines. Proliferation in both cell lines decreased in response to YAP/TAZ knockdown. The present study indicated that VP has potential as a therapeutic agent in YAP- and TAZ-dominant gastric cancers due to its ability to suppress the anti-apoptotic protein Survivin via inhibition of YAP and TAZ.

New Closure Method Using Loop and Open-Close Clips after Endoscopic Submucosal Dissection of Stomach and Colon Lesions.

Endoscopic submucosal dissection (ESD) and en bloc resection of stomach and colon tumors have become common. However, mucosal defects resulting from ESD may cause delayed bleeding and perforation. To prevent adverse events, we developed a new clip closure technique, namely, the loop and open-close clip closure method (LOCCM), and aimed to examine its efficacy after ESD for stomach and colon tumors. The LOCCM uses loop and open-close clips. Here, the open-close clip was used to grasp the loop to bring it to the edge of the post-ESD mucosal defect. Another clip with a loop was then inserted into the opposite edge and clipped to the contralateral mucosa to pull both edges together. Once apposed, additional clips facilitated complete closure. The LOCCM was performed in 19 patients after ESD at Tottori University between October 2020 and March 2021. The outcomes retrospectively analyzed were the LOCCM success and adverse event rates. The complete closure rate using LOCCM was 89.5% and none of the patients had post-ESD bleeding or perforation. The results show that LOCCM is an effective and safe closure technique for mucosal defects after stomach and colon ESD to prevent bleeding and perforation.

Impact of endoscopic submucosal dissection and epithelial cell sheet engraftment on systemic cytokine dynamics in patients with oesophageal cancer.

The tumour microenvironment (TME) plays an important role in cancer development, progression, and metastasis. Various cytokines are present in the TME in oesophageal cancer. Oesophageal stricture is a major complication of endoscopic submucosal dissection (ESD) for oesophageal cancer, and inflammatory cytokines are closely related to its pathogenesis. However, the cytokine crosstalk involved in the oesophageal cancer TME and post-ESD stricture has not been fully elucidated. This study investigated the comprehensive cytokine dynamics following ESD in patients with oesophageal cancer. In addition, the effect of a novel preventive technique for post-ESD stricture, autologous cell sheet engraftment, on cytokine levels was evaluated. Various pro-inflammatory and anti-tumorigenic cytokines were elevated in patients with oesophageal cancer, and ESD transiently influenced cytokine concentrations. IL-1ß and TNF-α, two major pro-inflammatory cytokines that induce oesophageal stricture, were significantly suppressed by cell sheet engraftment. In conclusion, this study revealed the distinct cytokine dynamics after ESD in patients with oesophageal cancer, together with the effect of autologous cell sheet engraftment on cytokine fluctuation. These results can accelerate research on the TME and therapeutic strategies for oesophageal cancer.

Detection of cytokine storm in patients with achalasia using ELISA.

Esophageal achalasia is characterized by abnormal peristaltic movements of the esophageal body and impaired relaxation of the lower esophageal sphincter (LES). However, its etiology remains unknown. In our previous study, it was shown that in the LES of patients with achalasia, hsv1-miR-H1 was overexpressed, ATG16L1 expression was downregulated and interleukin (IL)-1ß levels were upregulated. However, systemic features were not evaluated. Herein, the plasma cytokine levels in patients with achalasia were determined. Plasma was collected from patients at Nagasaki University Hospital between February 2013 and March 2016, both before and after peroral endoscopic myotomy (POEM). Cytokine analysis was performed using plasma collected from 10 healthy individuals (control group) and 12 patients with achalasia using the Bio-Plex Pro™ Human Cytokine 27-plex assay kit. The levels of IL-17, IL-1ß, C-C motif chemokine ligand 2, IL-4, IL-5, IL-1ra, IL-7, IL-12, interferon-γ, IL-2, fibroblast growth factor-2, colony-stimulating factor (CSF)2 and CSF3 were significantly higher in patients with achalasia compared with the control subjects. However, the levels did not differ between plasma samples collected before and after POEM. Thus, the occurrence of a cytokine storm was confirmed in the patients with achalasia.

Newly Invented Micellized Vitamin K2 Recovered Prolonged Prothrombin Time under Obstructive Jaundice in Rats with Bile Duct Ligation.

In cholestatic liver diseases, coagulopathy is induced by malabsorption of vitamin K. Supplementation of vitamin K has previously been shown to prevent coagulopathy. In this study, we tested the efficacy of a newly invented micellized vitamin K2 (m-vitK2) in treating coagulopathy, using a rat bile duct ligation (BDL) model. Experiment 1: m-vitK2 (0.3 mg/kg) or m-vitK2 (0.3 mg/kg) mixed with taurocholic acid (TA) (10 mg/body) was orally administrated every day for 7 d from the fourth day after BDL (n=6 for each). Experiment 2: To evaluate absorption, m-vitK2 (0.3 mg/kg) with or without TA (10 mg/body) was orally administered on the fourth day after BDL and compared with the untreated control BDL (n=2 for each). These data were compared with sham-operated (n=6) and untreated control BDL rats (n=6). The m-vitK2 recovered prothrombin time (PT) in Experiment 1 (control 42.7±5.7 s vs. m-vitK2 24.0±9.3 s, p<0.05). Experiment 2 demonstrated that the mixture of m-vitK2 and TA enhanced absorption compared to m-vitK2 alone. Moreover, in Experiment 1, m-vitK2 mixed with TA completely recovered PT (control 42.7±5.7 s vs. m-vitK2+TA 14.9±1.2 s, p<0.01). Micelle sizes decreased with the m-vitK2 and TA treatment (m-vitK2 86.3±5.6 nm vs. m-vitK2+TA 71.9±4.7 nm, p<0.05). Orally administered, newly invented m-vitK2 recovered coagulopathy even under obstructive jaundice. TA decreased the mean micelle size and improved m-vitK2 absorption.