Then, Huh7 cells were infected with the Bac-Nt-ORF2 to product HEV-LPs. Nt-ORF2 expression click here was confirmed by western blot analysis. HEV-LPs produced from Huh7 cells were purified by sucrose gradient centrifugation and then the morphology of purified HEV-LPs was observed by electron microscopy (EM). To determine liver-specific

property of HEV-LPs, intracellular penetration of FITC-conjugated HEV-LPs was evaluated by flow cytometry and visualized by confocal microscopy. To establish HEV-LPs packing system to carry a therapeutic gene inside the VLP, HEV-LPs were disassembled using biochemical buffer containing DTT, low concentration of CaCl2 and reassembled by increasing concentration of CaCl2 up to 50 mM. Results: Nt-ORF2 expression and HEV-LPs assembly were observed in Huh7 cells infected with Bac-Nt-ORF2. The purified HEV-LPs particles were found 25 nm in diameter

in EM. Subsequently, intracellular penetration of FITC-conjugated HEV-LPs was observed with check details high frequency in hepatoma cell lines while that was not detected in the cell lines derived from other organs such as lung, colon, kidney, ovarian, and cervix. Furthermore, we confirmed that the morphology of HEV-LPs was well preserved after disassembly/reassembly procedure using biochemical buffer. Conclusions: We established HEV-LP as a liver-specific delivery system using baculovirus vector system and this system could be useful tool for a liver-specific target therapy in chronic liver disease. This research was supported by grants of Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2012-001941). Disclosures: The following people have nothing to disclose:

Jung-Hee Kim, Wonhee Hur, Eun Byul Lee, Jung Eun Choi, Seung Kew Yoon Development of HCC-specific adenoviral gene therapy inserted with cancer-specific human telomerase 上海皓元医药股份有限公司 reverse transcrip-tase(hTERT) RNA-targeting trans-splicing ribozyme, liver specific promoter, phosphoenolpyruvate carboxykinase(PEPCK) and suicidal HSV thymidine kinase(TK) gene, has been successfully performed, proving excellent efficacy followed by ganciclovir treatment. We developed next generation which overcomes possible TERT-targeting to non-neoplastic hepato-cytes via microRNA regulation. New construct designed with insertion of antisense target sequence of liver-specific microR-NA(miR122a), which will provide null expression in normal hepatocytes, and mTERT RNA-targeting ribozyme(Ad-PEP-CK-mTERT.Ribo-TK-mir122aT[PRT-mir122aT]) which enables immunocompetent animal study. Here, we studied mouse HCC-specific antitumor efficacy of PRT-mir122aT without non-neoplastic hepatocyte damage in syngeneic HCC model, and checked possible involvement of tumor-specific immune response by subsequent challenge with same tumor cells. Vec-tors(PRT-mir122aT, Ad-PEPCK-mTERT.Ribo-TK[PRT], Ad-PEP-CK-TK[PT], Ad-MOCK) were prepared. mTERT(+), miR122a(-) Hepa1-6 mouse HCC cell was used.

Increased hepatic CD1d has also been noted in patients

with severe NASH.24 Thus, factors that promote NKT cell recruitment, retention, and viability are induced in human and rodent livers ZD1839 molecular weight with NASH-related fibrosis. Hh-pathway activation plays an important role in this process because a genetic manipulation that increases Hh-pathway activity in murine livers exacerbates NASH-related enrichment of liver NKT cells. Hh-signaling may also mediate hepatic NKT cell accumulation in human NASH given the striking correlation between hepatic Hh-pathway activity and the level of enrichment of liver mononuclear cells with NKT cells in patients with NAFLD-related cirrhosis. Our work also suggests that NKT cells actively promote fibrogenesis in NASH because CD1d-deficient mice that lack NKT cells are protected from NASH-related fibrosis and treatment of mouse primary HSCs with conditioned medium from LMNC that contained αGalCer-activated NKT cells stimulated stellate cells to become myofibroblastic. Previously, we showed that primary liver NKT cells from mice produce Shh, and that Shh stimulates NKT cells to produce the profibrogenic cytokines, IL-4 and IL-13.29 Shh also directly stimulates

myofibroblastic activation of HSCs and promotes the proliferation and survival of liver myofibroblasts.36 Ptc+/− BAY 57-1293 datasheet mice develop worse fibrosis after either bile duct ligation27 or MCD diets.39 Others have reported that IL-13 production increases in mice with NASH-fibrosis, and shown that treatments that neutralize IL-13 reduce fibrogenesis.45 Likewise, inhibiting IL-4 activity is known to diminish hepatic fibrosis in mice.46 Our human studies demonstrate that hepatic enrichment with NKT cells is a feature of cirrhosis. Definitive NKT cell enrichment was observed in patients with NASH-related cirrhosis, in whom we detected a 4 to 5-fold relative increase

in liver NKT cells. The present study confirms the previous reports that increased hepatic expression of CD1d occurs in advanced NAFLD, raising the possibility that antigen presentation to NKT cells may medchemexpress be enhanced. This is intriguing because CD1d presents lipid antigens to NKT cells and lipid homeostasis is abnormal in NAFLD. However, further research is needed to determine if and why there might be disease-related differences in hepatic accumulation of NKT cells. Additional studies to address the possibility that NKT cells may interact with other types of innate immune cells to modulate fibrosis progression are also justified because CD56(+)/CD3(−) cells (i.e., NK cells) were relatively depleted in the human cirrhotic livers that we examined, and liver NK cells are thought to serve antifibrogenic functions.6 Also, it has been suggested that liver macrophages, which are a rich source of immunomodulatory cytokines, may be altered in NASH,18 and this could further influence fibrotic activity.

Methods: In this prospective study, from March 2011 to February 2013, six first-year GI fellows performed 500 colonoscopies respectively. Each fellow performed standard colonoscopy (SC) in the first

150 cases, then, 6 fellows were divided into 2 groups, which were CAP-ACE group and SC group. The 3 fellows in CAP-ACE group performed 30 procedures from the 150st case using a CAP-ACE with indigocarmine, and then, the rest 3 fellows performed additional 350 SCs. Six GI fellows made and fulfilled the “colonoscopy learning protocol” which includes all related parameters. Results: Six first-year GI fellows participated and a total of 3,000 colonoscopy procedures were analyzed. There were no significant differences in gender, selleck chemicals indications and bowel preparations between the two groups. Mean withdrawal time were only 1 minute longer in CAP-ACE group. In the first 150 cases, ADR, advanced ADR, number of patients with more than 3 adenomas (NMT3As) and mean number

of adenomas per patient (MNAPP) were similar. However, in the latter 350 cases, ADR, advanced ADR, NMT3As and MNAPP were significantly increased in CAP-ACE group. On per adenoma analysis, more flat and smaller adenomas were detected in CAP-ACE group than SC group. Conclusion: After technical competency of colonoscopy, the intervention of CAP-ACE significantly improved ADRs including advanced or multiple ones in the trainees, implicating the introduction of technological assistance in the colonoscopy training programs for quality improvement. Key Word(s): 1. Trainee; 2. Adenoma detection; 3. Colonoscopic cap; 4. Chromoendoscopy; Presenting Author: YING QI Additional

Authors: BINGXIA GAO Corresponding Author: BINGXIA Selleck BIBW2992 GAO Affiliations: Beijing Shijitan Hospital. CMU Objective: To study the clinicopathological features of primary gastrointestinal lymphoma (PGIL) in order to improve the early diagnosis. Methods: Retrospective analysis of clinical data of 23 pathologically or endoscopically confirmed PGIL cases in our hospital from January 1994 to December 2012. Results: The study comprised 12 patients with primary gastric lymphoma, 6 with primary small intestinal lymphoma and 5 with primary large intestinal lymphoma. The main clinical symptom was abdominal pain (91.30%), emaciation (47.83%), abdominal mass and anemia medchemexpress (43.48%) and anorexia (34.78%). Among them, mild anemia is quite common (66.67%). The common endoscopic findings of PGL was infiltration type (45.45%), while nodular protruding type was more common in colon lymphoma (80.0%), Multiple polypoid change was found in a colon lymphoma case. In pathological study, 5 cases (21.74%) were low-grade malignant lymphoma (MALT lymphoma), 15 cases (65.22%) high-grade lymphoma including 13 cases of diffused large B-cell type and 2 cases of T-cell lymphoma. 3 cases (13.04%) were not classified (all in 1994–1995). 11 cases were at stage I (47.83%), 9 cases at stage II (39.13%), including 6 cases at stage IIE, 1 case at stage III (4.

2 Patients present with depleted fat stores and varying degrees of muscle wasting and reduced muscle strength. Short-term survival is reduced in malnourished patients with cirrhosis, and PEM can adversely affect outcomes for patients Selleckchem AP24534 on the waiting list for transplantation, as well as post-transplantation morbidity and mortality.3 Malnutrition in cirrhosis is implicated in increased risk for infection,4 increased severity of ascites,5 and the development of hepatic encephalopathy.6 Repeated episodes of overt hepatic encephalopathy might result in recurrent

hospitalizations and persistent cumulative deficits in working memory, response inhibition, and learning.7 The mechanisms of PEM in cirrhosis are complex and multifactorial. They include reduced oral intake secondary

to disease-related Selleckchem 17-AAG anorexia, restrictive diets, including overzealous sodium-restricted diets, altered taste sensations, nausea, early satiety, particularly in the presence of marked ascites, portal-hypertension associated malabsorption, insulin resistance, reduced glycogen storage capacity, increased gluconeogenesis, and alterations in fuel utilization. Repeated episodes of infection and endotoxemia as a result of alterations in gut barrier function might also contribute to the increased energy requirements and reduced intakes in this group via the pro-inflammatory cytokine response.8 Despite studies demonstrating that there is no benefit to a low-protein diet in patients with episodic or chronic hepatic encephalopathy,9 a protein-restricted diet is commonly recommended by health-care practitioners under the misapprehension that it is beneficial

to patients. Protein restriction has further deleterious 上海皓元 impacts on the severity of malnutrition,10 while a higher protein intake has positive benefits both on overall nutrition and possibly the severity of hepatic encephalopathy.11 Hospitalization, with frequent prolonged periods of fasting for diagnostic or therapeutic procedures, is another major contributory factor; thus, clinicians should make every effort to minimize periods of fasting and maximize nutritional intake in patients with cirrhosis while they are in hospital. Accurate assessment of nutritional status might also be difficult in patients with cirrhosis. This is because many of the traditional markers of nutritional assessment are dependent on normal hepatic synthetic function. Weight is a poor indicator of nutritional status in the presence of ascites and/or peripheral edema. Nutritional assessment of the cirrhotic patient includes subjective global assessment—liver,12 anthropometrical measurements of mid-arm circumference, triceps skinfold thickness, and mid-arm muscle circumference. In addition, grip strength measurements are an accurate reflection of protein status in those with cirrhosis.

2 Patients present with depleted fat stores and varying degrees of muscle wasting and reduced muscle strength. Short-term survival is reduced in malnourished patients with cirrhosis, and PEM can adversely affect outcomes for patients selleck kinase inhibitor on the waiting list for transplantation, as well as post-transplantation morbidity and mortality.3 Malnutrition in cirrhosis is implicated in increased risk for infection,4 increased severity of ascites,5 and the development of hepatic encephalopathy.6 Repeated episodes of overt hepatic encephalopathy might result in recurrent

hospitalizations and persistent cumulative deficits in working memory, response inhibition, and learning.7 The mechanisms of PEM in cirrhosis are complex and multifactorial. They include reduced oral intake secondary

to disease-related Depsipeptide solubility dmso anorexia, restrictive diets, including overzealous sodium-restricted diets, altered taste sensations, nausea, early satiety, particularly in the presence of marked ascites, portal-hypertension associated malabsorption, insulin resistance, reduced glycogen storage capacity, increased gluconeogenesis, and alterations in fuel utilization. Repeated episodes of infection and endotoxemia as a result of alterations in gut barrier function might also contribute to the increased energy requirements and reduced intakes in this group via the pro-inflammatory cytokine response.8 Despite studies demonstrating that there is no benefit to a low-protein diet in patients with episodic or chronic hepatic encephalopathy,9 a protein-restricted diet is commonly recommended by health-care practitioners under the misapprehension that it is beneficial

to patients. Protein restriction has further deleterious 上海皓元 impacts on the severity of malnutrition,10 while a higher protein intake has positive benefits both on overall nutrition and possibly the severity of hepatic encephalopathy.11 Hospitalization, with frequent prolonged periods of fasting for diagnostic or therapeutic procedures, is another major contributory factor; thus, clinicians should make every effort to minimize periods of fasting and maximize nutritional intake in patients with cirrhosis while they are in hospital. Accurate assessment of nutritional status might also be difficult in patients with cirrhosis. This is because many of the traditional markers of nutritional assessment are dependent on normal hepatic synthetic function. Weight is a poor indicator of nutritional status in the presence of ascites and/or peripheral edema. Nutritional assessment of the cirrhotic patient includes subjective global assessment—liver,12 anthropometrical measurements of mid-arm circumference, triceps skinfold thickness, and mid-arm muscle circumference. In addition, grip strength measurements are an accurate reflection of protein status in those with cirrhosis.

After centrifuging at 13 000 × g for 30 min, pellets were washed with 70% (v/v) ethanol. After allowing the ethanol to evaporate completely, pellets were dissolved in 100 μL of diethylene pyrocarbonate-treated water (Invitrogen, Carlsbad, CA, USA). cDNA was prepared using reverse transcriptase originating from Murine-Moloney leukemia virus (Promega),

according to the manufacturer’s instructions. PCR primers for iNOS, COX-2, IL-1β, IL-6, ICAM-1, VCAM, HIF-1a, PDGF, HO-1, and GAPDH were shown in Table 2. PCR was performed over 28 cycles of 94°C for 30 s, 52°C for 30 s, and 72°C for 30 s. Oligonucleotide primers were purchased from Bioneer ICG-001 (Seoul, Korea). Cultured cells were washed twice with cold PBS on ice and harvested by scraping with a rubber scraper. Cells were sedimented by centrifugation at 4°C and resuspended in cell lysis buffer. After centrifugation (13 000 × g), the IKKβ activity in the supernatant was measured. The processes were performed using the IKKβ kinase assay kit (Cell Signaling Technology).

All samples were measured for their individual levels, and each sample was analyzed in triplicate manner, taking the mean of the three determinations. Cultured cells were washed twice with cold PBS on ice and harvested by scraping with a rubber scraper. Cells were sedimented check details by centrifugation at 4°C and resuspended in cell lysis buffer. After centrifugation (13 000 × g), the HDAC activity in the supernatant was measured. The processes were performed as Histone Deacetylase Assay Kit, Fluorometric kit

(Sigma). All samples were measured for their individual levels, and each sample was analyzed in triplicate manner, taking 上海皓元 the mean of the three determinations. The data are presented as means ± SD. The statistical significance was assessed using one-way anova. Differences were considered to be significant for values of P < 0.05. Administration of 20 mg/kg indomethacin by gavage resulted in the development of significant gastric damages in all control mice, among which 80% mice showed definite gastric ulcer accompanied with brisk gastric hemorrhages (Fig. 1a). Microscopically, the exposure to indomethacin for 24 h provoked definite gastric mucosal lesions. However, pretreatment of SAC significantly decreased the incidence of gastric damage. As can be seen in Figure 1a, 3 mg/kg SAC significantly decreased the gastric mucosal lesions. Excavating ulcers and hemorrhagic necrosis of mucosa accompanied with inflammatory cell infiltration and focal hemorrhages were investigated in indomethacin-treated group. Significant pathologies, such as intense gastric inflammation and some mucosal erosive lesions, developed in indomethacin-treated group in all mice (Fig. 1a). Pathological lesion index of ulceration (Fig. 1b) and total pathologic score (Fig. 1c) were all increased in indomethacin-induced gastric damage group (P < 0.05).

However, the molecular mechanism of PTEN in hepatocellular carcinoma (HCC) metastasis is unclear. In this study, we found frequent (47.5%, n = 40) protein underexpression of PTEN in human HCCs compared with their corresponding nontumorous livers. Significantly, PTEN underexpression was associated with larger tumor size (P = 0.021), tumor microsatellite formation (P = 0.027), and shorter overall survival

of patients (P = 0.035). Using different cell models, we observed that PTEN-knockdown HCC cells and PTEN-knockout mouse embryonic fibroblasts (MEFs) had enhanced cell migratory and invasive abilities. In addition to activation of AKT, there was up-regulation of the Sp1 transcription factor (SP1) and matrix metalloproteinase 2 (MMP2), as well as MMP2 activation in PTEN-knockdown HCC cells and PTEN−/− GSI-IX cell line MEFs. With dual luciferase reporter A-769662 molecular weight assay, exogenous expression of SP1 in HCC cells led to enhanced MMP2 promoter activity by up to 74%, whereas deletion of the putative SP1 binding site on the MMP2 promoter led to reduced promoter activity by up to 65%. Using chromatin immunoprecipitation assay, we documented increased binding of SP1 to the MMP2 promoter in PTEN-knockdown HCC cells. Overexpression of SP1 and MMP2 was significantly but negatively associated with PTEN underexpression in human HCCs. Conclusion: Our results show that PTEN was underexpressed in HCCs,

and this underexpression was associated with more aggressive biological behavior and poorer patient survival. We have provided the first

evidence that MMP2 up-regulation upon PTEN loss is SP1-dependent. Our findings indicate that PTEN plays a significant role in down-regulating HCC cell invasion via the AKT/SP1/MMP2 pathway. (HEPATOLOGY 2011;) Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide and the second most common fatal cancer in Southeast Asia.1 HCC has a poor prognosis with high mortality. The majority of patients present late with advanced HCC,2 thereby limiting potentially curative 上海皓元 therapeutic options. Cancer metastases, both intrahepatic and extrahepatic, are major factors for the mortality of HCC patients. Nonetheless, the molecular mechanisms underlying HCC metastasis remain largely unclear. Phosphatase and tensin homolog (PTEN) is one of the most frequently mutated tumor suppressors, only second to p53. It is constitutively expressed, whereas p53 is a stress-responsive tumor suppressor. PTEN is an upstream negative regulator of the survival phosphoinositide 3-kinase (PI3K)/AKT cascade; activation of this signaling is frequently observed in multiple cancers due to loss of PTEN. AKT is a central regulator of various cellular processes—including cell survival, proliferation, growth, angiogenesis, and metabolism—via phosphorylation of various substrates.3 Hence, loss of PTEN gatekeeper function plays a pivotal role in promoting carcinogenesis.

However, the molecular mechanism of PTEN in hepatocellular carcinoma (HCC) metastasis is unclear. In this study, we found frequent (47.5%, n = 40) protein underexpression of PTEN in human HCCs compared with their corresponding nontumorous livers. Significantly, PTEN underexpression was associated with larger tumor size (P = 0.021), tumor microsatellite formation (P = 0.027), and shorter overall survival

of patients (P = 0.035). Using different cell models, we observed that PTEN-knockdown HCC cells and PTEN-knockout mouse embryonic fibroblasts (MEFs) had enhanced cell migratory and invasive abilities. In addition to activation of AKT, there was up-regulation of the Sp1 transcription factor (SP1) and matrix metalloproteinase 2 (MMP2), as well as MMP2 activation in PTEN-knockdown HCC cells and PTEN−/− Apoptosis inhibitor MEFs. With dual luciferase reporter Daporinad mw assay, exogenous expression of SP1 in HCC cells led to enhanced MMP2 promoter activity by up to 74%, whereas deletion of the putative SP1 binding site on the MMP2 promoter led to reduced promoter activity by up to 65%. Using chromatin immunoprecipitation assay, we documented increased binding of SP1 to the MMP2 promoter in PTEN-knockdown HCC cells. Overexpression of SP1 and MMP2 was significantly but negatively associated with PTEN underexpression in human HCCs. Conclusion: Our results show that PTEN was underexpressed in HCCs,

and this underexpression was associated with more aggressive biological behavior and poorer patient survival. We have provided the first

evidence that MMP2 up-regulation upon PTEN loss is SP1-dependent. Our findings indicate that PTEN plays a significant role in down-regulating HCC cell invasion via the AKT/SP1/MMP2 pathway. (HEPATOLOGY 2011;) Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide and the second most common fatal cancer in Southeast Asia.1 HCC has a poor prognosis with high mortality. The majority of patients present late with advanced HCC,2 thereby limiting potentially curative MCE公司 therapeutic options. Cancer metastases, both intrahepatic and extrahepatic, are major factors for the mortality of HCC patients. Nonetheless, the molecular mechanisms underlying HCC metastasis remain largely unclear. Phosphatase and tensin homolog (PTEN) is one of the most frequently mutated tumor suppressors, only second to p53. It is constitutively expressed, whereas p53 is a stress-responsive tumor suppressor. PTEN is an upstream negative regulator of the survival phosphoinositide 3-kinase (PI3K)/AKT cascade; activation of this signaling is frequently observed in multiple cancers due to loss of PTEN. AKT is a central regulator of various cellular processes—including cell survival, proliferation, growth, angiogenesis, and metabolism—via phosphorylation of various substrates.3 Hence, loss of PTEN gatekeeper function plays a pivotal role in promoting carcinogenesis.

However, the molecular mechanism of PTEN in hepatocellular carcinoma (HCC) metastasis is unclear. In this study, we found frequent (47.5%, n = 40) protein underexpression of PTEN in human HCCs compared with their corresponding nontumorous livers. Significantly, PTEN underexpression was associated with larger tumor size (P = 0.021), tumor microsatellite formation (P = 0.027), and shorter overall survival

of patients (P = 0.035). Using different cell models, we observed that PTEN-knockdown HCC cells and PTEN-knockout mouse embryonic fibroblasts (MEFs) had enhanced cell migratory and invasive abilities. In addition to activation of AKT, there was up-regulation of the Sp1 transcription factor (SP1) and matrix metalloproteinase 2 (MMP2), as well as MMP2 activation in PTEN-knockdown HCC cells and PTEN−/− Osimertinib MEFs. With dual luciferase reporter this website assay, exogenous expression of SP1 in HCC cells led to enhanced MMP2 promoter activity by up to 74%, whereas deletion of the putative SP1 binding site on the MMP2 promoter led to reduced promoter activity by up to 65%. Using chromatin immunoprecipitation assay, we documented increased binding of SP1 to the MMP2 promoter in PTEN-knockdown HCC cells. Overexpression of SP1 and MMP2 was significantly but negatively associated with PTEN underexpression in human HCCs. Conclusion: Our results show that PTEN was underexpressed in HCCs,

and this underexpression was associated with more aggressive biological behavior and poorer patient survival. We have provided the first

evidence that MMP2 up-regulation upon PTEN loss is SP1-dependent. Our findings indicate that PTEN plays a significant role in down-regulating HCC cell invasion via the AKT/SP1/MMP2 pathway. (HEPATOLOGY 2011;) Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide and the second most common fatal cancer in Southeast Asia.1 HCC has a poor prognosis with high mortality. The majority of patients present late with advanced HCC,2 thereby limiting potentially curative 上海皓元 therapeutic options. Cancer metastases, both intrahepatic and extrahepatic, are major factors for the mortality of HCC patients. Nonetheless, the molecular mechanisms underlying HCC metastasis remain largely unclear. Phosphatase and tensin homolog (PTEN) is one of the most frequently mutated tumor suppressors, only second to p53. It is constitutively expressed, whereas p53 is a stress-responsive tumor suppressor. PTEN is an upstream negative regulator of the survival phosphoinositide 3-kinase (PI3K)/AKT cascade; activation of this signaling is frequently observed in multiple cancers due to loss of PTEN. AKT is a central regulator of various cellular processes—including cell survival, proliferation, growth, angiogenesis, and metabolism—via phosphorylation of various substrates.3 Hence, loss of PTEN gatekeeper function plays a pivotal role in promoting carcinogenesis.

0000, A<->G = 4.0364, A<->T = 1.0000, C<->G = 1.0000, C<->T = 7.4185; proportion of sites assumed to be invariable=0.3824; rates for variable sites assumed to follow a gamma Pembrolizumab distribution with shape parameter=0.6037; and number of rate categories=4. Bootstrap (BS) values were calculated to 1,000 pseudoreplicates by full heuristic

search which was performed with an NJ starting tree option with a TBR swapping algorithm. Six of the species with the chlorophyll a derivative were encountered on the seafloor at a depth of about 30–40 m (No. 1–4) or on a sandy beach (No. 5 and 6) and so are benthic in habit. Alexandrium hiranoi (HG3) used for comparison was collected from tide pool sample. Using light Buparlisib microscope characteristics only, these six species were identified as B. angelaceum (Fig. S1A;

No. 1, Yamada et al. 2013), A. gibbosum (Fig. S1B; No.2, Murray et al. 2004) and Symbiodinium spp. (Fig. S1, E and F; No. 5 and 6), and the other two remained unidentified (Fig. S1, C and D; No. 3 and 4). The two unidentified dinoflagellates exhibit a similar type of life cycle consisting of three forms: rare swimming cells, nonmotile, with flagella, attached cells (the stage during which cell division takes place), and floating, nonmotile cells (a stage during which no cell division takes place). These two benthic dinoflagellates were here treated as unidentified athecate dinoflagellate 1 and 2 because their identity could not be resolved. We analyzed the photosynthetic pigments using HPLC which enabled the determination of the chlorophyll and carotenoid content. The photosynthetic pigments were extracted with acetone and subjected to HPLC and the elution profiles were monitored by measuring the absorbance at 450 nm. Typical profiles are shown in Figure 1. All the dinoflagellates, including Alexandrium hiranoi, examined contained the major carotenoid of dinoflagellates, peridinin. They also have diadinoxanthin and diatoxanthin of the diadinoxanthin cycle, which is involved

in the dissipation 上海皓元医药股份有限公司 of surplus absorbed light energy. β-Carotene was also common to all stains examined, but many unidentified carotenoids were found in some (Fig. 1, A, C and E). The strains universally contained chlorophyll a and chlorophyll c2 as reported previously, and chlorophyll c1 also was found in all except B. angelaceum (No. 1). A new peak (X) was found at a retention time of 21 min. We also examined the pigment compositions of B. angelaceum (No. 1) for various culture periods (1–5 months) and various light intensities (20, 60, 100 μmol photons · m−2 · s−1). However, we did not detect any effects of culture periods and light intensities (data not shown); the pigment composition of B. angelaceum were the same in all conditions, and peak X was detected under any cellular conditions. The Fv/Fm ratio using PAM method of both culture periods in 3 and 5 months was around 0.30–0.35.