Long-Term Fluorescent Tissue Marking Using Tissue-Adhesive Porphyrin with Polycations Consisting of Quaternary Ammonium Salt Groups.

Localization of tumors during laparoscopic surgery is generally performed by locally injecting India ink into the submucosal layer of the gastrointestinal tract using endoscopy. However, the location of the tumor is obscured because of the black-stained surgical field and the blurring caused by India ink. To solve this problem, in this study, we developed a tissue-adhesive porphyrin with polycations consisting of quaternary ammonium salt groups. To evaluate the ability of tissue-adhesive porphyrin in vivo, low-molecular-weight hematoporphyrin and tissue-adhesive porphyrin were injected into the anterior wall of the exposed stomach in rats. Local injection of low-molecular-weight hematoporphyrin into the anterior wall of the stomach was not visible even after 1 day because of its rapid diffusion. In contrast, the red fluorescence of the tissue-adhesive porphyrin was visible even after 7 days due to the electrostatic interactions between the positively-charged moieties of the polycation in the tissue-adhesive porphyrin and the negatively-charged molecules in the tissue. In addition, intraperitoneal injection of tissue-adhesive porphyrin in rats did not cause adverse effects such as weight loss, hepatic or renal dysfunction, or organ adhesion in the abdominal cavity. These results indicate that tissue-adhesive porphyrin is a promising fluorescent tissue-marking agent.

Elevated Production of Mitochondrial Reactive Oxygen Species via Hyperthermia Enhanced Cytotoxic Effect of Doxorubicin in Human Breast Cancer Cell Lines MDA-MB-453 and MCF-7.

Hyperthermia (HT) treatment is a noninvasive cancer therapy, often used with radiation therapy and chemotherapy. Compared with 37 °C, 42 °C is mild heat stress for cells and produces reactive oxygen species (ROS) from mitochondria. To involve subsequent intracellular accumulation of DOX, we have previously reported that the expression of ATP-binding cassette sub-family G member 2 (ABCG2), an exporter of doxorubicin (DOX), was suppressed by a larger amount of intracellular mitochondrial ROS. We then hypothesized that the additive effect of HT and chemotherapy would be induced by the downregulation of ABCG2 expression via intracellular ROS increase. We used human breast cancer cell lines, MCF-7 and MDA-MB-453, incubated at 37 °C or 42 °C for 1 h to clarify this hypothesis. Intracellular ROS production after HT was detected via electron spin resonance (ESR), and DOX cytotoxicity was calculated. Additionally, ABCG2 expression in whole cells was analyzed using Western blotting. We confirmed that the ESR signal peak with HT became higher than that without HT, indicating that the intracellular ROS level was increased by HT. ABCG2 expression was downregulated by HT, and cells were injured after DOX treatment. DOX cytotoxicity enhancement with HT was considered a result of ABCG2 expression downregulation via the increase of ROS production. HT increased intracellular ROS production and downregulated ABCG2 protein expression, leading to cell damage enhancement via DOX.

First discovery of Perkinsus beihaiensis in Mediterranean mussels (Mytilus galloprovincialis) in Tokyo Bay, Japan.

During analyses of the invasive Mediterranean mussel Mytilus galloprovincialis for pathologies in Tokyo Bay, infection by the protozoan parasite Perkinsus beihaiensis was found through histological examination, Ray's Fluid Thioglycollate Medium assays, and molecular analyses. Specific PCR assays for each Perkinsus species also revealed the presence of an indigenous congeneric species, Perkinsus olseni, but P. beihaiensis was dominant in M. galloprovincialis. Sequences of the ribosomal internal transcribed spacer region I of P. beihaiensis found in Japan were genetically more similar to those found in South American countries (Panama and Brazil) than in Asian countries (China and India). Though Mediterranean mussels have become widespread in Japanese waters since their invasion in the 1930s, epidemiological surveys show that mussels collected outside Tokyo Bay are free of any Perkinsus infections. Based on these results, it was strongly suggested that P. beihaiensis invaded Tokyo Bay by transportation of bivalves originating from South America but has not yet spread to other parts of Japan. The possibility is not ruled out, however, that the parasite is indigenous in Japan but the environment in Tokyo Bay favors its transmission to Mediterranean mussels.

Singlet oxygen-generating cell-adhesive glass surfaces for the fundamental investigation of plasma membrane-targeted photodynamic therapy.

Recently, plasma membrane-targeted photodynamic therapy has attracted attention as an effective cancer immunotherapeutic strategy. However, the released photosensitizers do not only adhere to the plasma membrane but may also be internalized in the cytosol, in endosomes/lysosomes, hindering investigations of the effects of photosensitizers attached to the plasma membrane. In this study, we developed a cell culture dish with singlet oxygen-generating cell-adhesive glass surfaces that allows investigation of the effects of photosensitizers attached to the plasma membrane. For cell adhesion, poly[N-(3-aminopropyl)methacrylamide] conjugated with hematoporphyrin PA-HpD was immobilized on the glass surfaces. Singlet oxygen was produced from the PA-HpD-immobilized glass surface upon laser irradiation at 635 nm. When murine colon adenocarcinoma 26 (Colon-26) cells were cultured on the PA-HpD-immobilized surface, the cells were swollen and ruptured, leading to effective apoptotic cell death using laser irradiation at 635 nm. In addition, microvesicles of approximately 10 µm in diameter were released from the plasma membrane into the culture medium. These phenomena were due to the oxidation of lipids in the cellular membrane, caused by the plasma membrane-targeted photodynamic therapy. In contrast, these phenomena were not observed on poly[N-(3-aminopropyl)methacrylamide]-immobilized glass surfaces. These results indicate that cell culture dishes with singlet oxygen-generating cell-adhesive glass surfaces can be used to investigate fundamental mechanisms in plasma membrane-targeted photodynamic therapy.

Locally Administered Photodynamic Therapy for Cancer Using Nano-Adhesive Photosensitizer.

Photodynamic therapy (PDT) is a great potential anti-tumor therapy owing to its non-invasiveness and high spatiotemporal selectivity. However, systemically administered photosensitizers diffuse in the skin and the eyes for a long duration, which cause phototoxicity to bright light and sunlight. Therefore, following PDT, patients must avoid exposure of to light and sunlight to avoid this phototoxicity. In this study, we have developed a locally administered PDT using nano-adhesive porphyrin with polycations consisting of quaternary ammonium salt groups (aHP) as a photosensitizer. The aHP, approximately 3.0 nm in diameter, adhered the negatively charged cell membrane via electrostatic interaction. The aHP localized to the endosome via cell adhesion and induced apoptosis upon 635 nm light irradiation. On being administered subcutaneously on the tumor, 30% of the injected aHP remained in the administered sites. However, low-molecular-weight hematoporphyrin dihydrochloride (HP) disappeared due to rapid diffusion. PDT with locally administered aHP showed a higher anti-tumor effect after light irradiation at 635 nm for three days compared to low-molecular-weight HP. Intraperitoneal administration of HP caused severe phototoxicity upon irradiation with ultraviolet A at 10 J cm-2, whereas aHP did not cause phototoxicity because its diffusion into the skin could be suppressed, probably due to the high-molecular weight of aHP. Therefore, locally administered PDT with aHP is a potential PDT having high therapeutic efficacy without phototoxicity.

A case report: Long-term complete response of metastatic hepatocellular carcinoma obtained after discontinuation of 2-month sorafenib monotherapy.

A 69-year-old woman who had a history of chronic hepatitis C, autoimmune hemolytic anemia and myelodysplastic syndrome was treated with sorafenib at a daily dose of 400 mg for HCC with multiple lung metastases. Nonetheless, elevated serum tumor markers further increased (alpha fetoprotein from 121,100 to 348,660 ng/ml and protein induced by vitamin K absence/antagonist-II from 3435 to 29,357 mAU/ml), and lung metastatic lesions on chest X-ray showed no improvement after 2 months of sorafenib treatment. Sorafenib was discontinued because of adverse events with diarrhea, fatigue, and severe anemia due to bleeding from stomach telangiectasia. Hand-foot syndrome was mild. Thereafter, the tumor markers rapidly decreased to almost normal range, and the lung and liver tumors markedly shrunk and disappeared without any other cancer treatments. Her tumors remained in complete remission for 17 months until an intrahepatic recurrence occurred. This unique course of metastatic HCC indicated that antitumor mechanisms other than the direct anticancer effect of sorafenib contributed to tumor shrinkage.

Mediastinal Neuroendocrine Carcinoma Slowly Growing for 8 Years after Surgical Resection of Esophageal Squamous Cell Carcinoma.

A 70-year-old woman was referred to our department due to a solitary mediastinal tumor which gradually grew near the site of anastomosis for 8 years after radical surgery of esophageal squamous cell carcinoma. It was difficult to distinguish the lymph node recurrence of esophageal cancer from another tumor of unknown primary origin. Endoscopic ultrasound-guided fine-needle aspiration was performed, and the tumor was diagnosed to be neuroendocrine carcinoma. She received concurrent chemoradiotherapy with etoposide plus cisplatin. After the completion of chemoradiotherapy, the tumor disappeared. A solitary growing tumor which develops after radical resection of cancer would be better to be examined histologically in order to make an accurate diagnosis and select the most appropriate treatment.