A Missense Mutation of the Plasminogen Gene in a Japanese Family with Hereditary Angioedema with Normal C1 Inhibitor: Third Family Survey in Asia.

Hereditary angioedema (HAE) is a life-threatening disease associated with recurrent episodes of subcutaneous and mucosal swelling, painful abdominal cramping, and asphyxiation. HAE has long been thought to be caused by genetic defects of C1 inhibitors (C1-INH). Recently, HAE with a normal C1 inhibitor expression (HAEnCI) was reported, and the missense mutation p.Lys330Glu (K330E) in exon 9 of the plasminogen (PLG) gene was shown to be responsible for a subset of HAEnCI. HAE with the K330E mutation in the PLG gene-PLG (HAE-PLG) has been reported in only two Japanese families in Asia. We herein report a third family with HAE-PLG in Japan.

Can Blood Ammonia Level, Prehospital Time, and Return of Spontaneous Circulation Predict Neurological Outcomes of Out-of-Hospital Cardiac Arrest Patients? A Nationwide, Retrospective Cohort Study.

BACKGROUND: This study aimed to test if blood ammonia levels at hospital arrival, considering prehospital time and the patient's condition (whether return of spontaneous circulation [ROSC] was achieved at hospital arrival), can predict neurological outcomes after out-of-hospital cardiac arrest (OHCA). METHODS: This was a retrospective cohort study on data from a nationwide OHCA registry in Japan. Patients over 17 years old and whose blood ammonia levels had been recorded were included. The primary outcome was favorable neurological outcome at 30 days after OHCA. Blood ammonia levels, prehospital time, and the combination of the two were evaluated using the receiver operating characteristic curve to predict favorable outcomes. Then, cut-off blood ammonia values were determined based on whether ROSC was achieved at hospital arrival. RESULTS: Blood ammonia levels alone were sufficient to predict favorable outcomes. The overall cut-off ammonia value for favorable outcomes was 138 µg/dL; values were different for patients with ROSC (96.5 µg/dL) and those without ROSC (156 µg/dL) at hospital arrival. CONCLUSIONS: Our results using patient data from a large OHCA registry showed that blood ammonia levels at hospital arrival can predict neurological outcomes, with different cut-off values for patients with or without ROSC at hospital arrival.

Incidence and characteristics of medical emergencies related to dental treatment: a retrospective single-center study.

AIM: Although uncommon, medical emergencies arise in general dental practice. Inadequate data on their severity and frequency makes targeting medical education for general dental practitioners difficult. This also makes planning for unexpected events challenging for practitioners and makes collaborating with emergency physicians burdensome. We aimed to clarify the incidence and characteristics of a dental outpatient department's medical emergencies. METHODS: This single-center, retrospective, observational study was undertaken with patients who visited the dental outpatient department of Okayama University Hospital during the 8-year period. The primary outcome of the study was to identify the incidence and characteristics of medical emergencies in the dental outpatient department. Then we examined the timing of medical emergencies, administered medications, and final disposition (home/admission). RESULTS: During the period, 1,146,929 patients were enrolled. Forty-two patients (0.0037%) were consulted as medical emergencies. More than 60% of the incidents were vasovagal syncope, and dehydration and hypoglycemia were the second most prevalent at 9.5%. The most common types of dental treatments were tooth extraction (45.2%), followed by general dental treatment (28.6%), and other dental surgery such as implant placement (14.3%). Types of medical emergencies occurred equally before, during, and after dental treatment. Antihypertensive agents, sedatives, or glucose were used. For patients with emergencies, 90.5% recovered during the day and returned home, and 9.5% were hospitalized. CONCLUSION: The incidence of medical emergencies was low in our dental outpatient department. Knowledge of basic management principles, regular education for emergency care, and practicing first aid skills are mandatory for safe patient management.

Delay in Emergency Medical Service Transportation Responsiveness during the COVID-19 Pandemic in a Minimally Affected Region.

Few studies have investigated the influence of the Coronavirus Disease 2019 (COVID-19) pandemic on emer-gency medical service (EMS) systems, especially in areas less affected or unaffected by COVID-19. In this study, we investigated changes in prehospital EMS activity and transport times during the COVID-19 pandemic. All patients transported by EMS in the city of Okayama from March-May 2019 or March-May 2020 were included. Interfacility transports were excluded. The primary outcome was the time from a patient's first emergency call until hospital arrival (total prehospital time). Secondary outcomes included three segments of total prehospital time: the response time, on-scene time, and transportation time. Total prehospital time and the durations of each segment were compared between corresponding months in 2020 (COVID19-affected) and 2019 (control). The results showed that total prehospital times in April 2020 were significantly higher than those in 2019 (33.8 ± 11.6 vs. 32.2 ± 10.8 min, p < 0.001). Increases in total prehospital time were caused by longer response time (9.3 ± 3.8 vs. 8.7 ± 3.7 min, p < 0.001) and on-scene time (14.4 ± 7.9 vs. 13.5 ± 6.2min, p < 0.001). The COVID-19 pandemic was thus shown to affect EMS and delayed arrival/response even in a minimally affected region. A system to minimize transportation delays should be developed for emerging pandemics.

Novel single-chain variant of antibody against mesothelin established by phage library.

Mesothelin (MSLN) shows increased expression in various cancer cells. For clinical application of antibodies as a positron emission tomography (PET) imaging reagent, a human shortened antibody is essential both for avoiding redundant immune responses and for providing rapid imaging. Therefore, we cloned a single-chain fragment of variable regions (scFv) from a human-derived gene sequence. This was achieved through the construction of a naïve phage library derived from human tonsil lymphocytes. Using a column with human recombinant MSLN, we carried out bio-panning of phage-variants by colony formation. We first obtained 120 clones that were subjected to selection in an ELISA using human recombinant MSLN as a solid phase antigen, and 15 phage clones of scFv with a different sequence were selected and investigated by flow cytometry (FCM). Then, six variants were selected and the individual scFv gene was synthesized in the VL and VH domains and expressed in Chinese hamster ovary cells. Mammalian cell-derived human-origin scFv clones were analyzed by FCM again, and one MSLN highly specific scFv clone was established. PET imaging by 89 Zr-labeled scFv was done in mice bearing xenografts with MSLN-expressing cancer cells, and tumor legions were successfully visualized. The scFv variant established in the present study may be potentially useful for cancer diagnosis by PET imaging.

Associations of obesity with tracheal intubation success on first attempt and adverse events in the emergency department: An analysis of the multicenter prospective observational study in Japan.

Obesity is deemed to increase the risk of difficult tracheal intubation. However, there is a dearth of research that examines the relationship of obesity with intubation success and adverse events in the emergency department (ED). We analyzed the data from a prospective, observational, multicenter study-the Japanese Emergency Airway Network (JEAN) 2 study from 2012 through 2016. We included all adults (aged ≥18 years) who underwent tracheal intubation in the ED. Patients were categorized into three groups according to their body mass index (BMI): lean (<25.0 kg/m²), overweight (25.0-29.9 kg/m²), and obesity (≥30.0 kg/m²). Outcomes of interest were intubation success on the first attempt and intubation-related adverse events. Of 6,889 patients who are eligible for the analysis, 5,370 patients (77%) were lean, 1,177 (17%) were overweight, and 342 (4%) were obese. Compared to the lean patients, the intubation success rates were significantly lower in the overweight and obese patients (70.9% in lean, 66.4% in overweight, and 59.3% in obese patients; P<0.001). In the multivariable analysis, compared to the lean patients, overweight (adjusted odds ratio [OR], 0.85; 95%CI, 0.74-0.98) and obese (adjusted OR, 0.62; 95%CI, 0.49-0.79) patients had a significantly lower success rate on the first attempt. Additionally, obesity was significantly associated with a higher risk of adverse events (adjusted OR, 1.62; 95%CI, 1.23-2.13). Based on the data from a multicenter prospectively study, obesity was associated with a lower success rate on the first intubation attempt and a higher risk of adverse event in the ED.

Hereditary Angioedema with Recurrent Abdominal Pain in a Patient with a Novel Mutation.

We describe a patient with hereditary angioedema type I. The patient had experienced recurrent abdominal pain around the time of her menstrual period for 13 years. A laboratory examination showed reduced functional and antigenic levels of C4 and C1 inhibitor (C1-INH). To establish a diagnosis, we carried out a DNA analysis of the patient's C1-INH gene. We determined that the patient was heterozygous for a single base pair transposition of T to C at nucleotide 4429 in exon 4, which had not been reported in the literature. As the patient had no family history of hereditary diseases, it was considered to be a de novo mutation.

A novel PET imaging using 64Cu-labeled monoclonal antibody against mesothelin commonly expressed on cancer cells.

Mesothelin (MSLN) is a 40-kDa cell differentiation-associated glycoprotein appearing with carcinogenesis and is highly expressed in many human cancers, including the majority of pancreatic adenocarcinomas, ovarian cancers, and mesotheliomas, while its expression in normal tissue is limited to mesothelial cells lining the pleura, pericardium, and peritoneum. Clone 11-25 is a murine hybridoma secreting monoclonal antibody (mAb) against human MSLN. In this study, we applied the 11-25 mAb to in vivo imaging to detect MSLN-expressing tumors. In in vitro and ex vivo immunochemical studies, we demonstrated specificity of 11-25 mAb to membranous MSLN expressed on several pancreatic cancer cells. We showed the accumulation of Alexa Fluor 750-labeled 11-25 mAb in MSLN-expressing tumor xenografts in athymic nude mice. Then, 11-25 mAb was labeled with (64)Cu via a chelating agent DOTA and was used in both in vitro cell binding assay and in vivo positron emission tomography (PET) imaging in the tumor-bearing mice. We confirmed that (64)Cu-labeled 11-25 mAb highly accumulated in MSLN-expressing tumors as compared to MSLN-negative ones. The (64)Cu-labeled 11-25 mAb is potentially useful as a PET probe capable of being used for wide range of tumors, rather than (18)F-FDG that occasionally provides nonspecific accumulation into the inflammatory lesions.

Tumorigenesis in cells derived from induced pluripotent stem cells.

Induced pluripotent stem (iPS) cells are an attractive source for potential cell-replacement therapy. However, transplantation of differentiated products harbors the risk of teratoma formation, presenting a serious health risk. Thus, we characterized Nanog-expressing (undifferentiated) cells remaining after induction of differentiation by cytological examination. To induce differentiation of iPS cells, we generated embryoid bodies (EBs) derived from iPS cells carrying a Nanog­green fluorescent protein(GFP) reporter and then injected GFP-positive and GFP negative EBs into nude mice. GFP-positive EB transplantation resulted in the formation of immature teratoma grade 3, but no tumors were induced by GFP-negative EB. GFP positive cells revealed significantly lower cytoplasmic area and higher nucleus/cytoplasm ratio than those of GFP negative cells. Our results suggest that morphological analysis might be a useful method for distinguishing between tumorigenic and nontumorigenic iPS cells.

Isolation of side population cells from endometrial cancer cells using a violet laser diode.

Cancer stem cells (CSCs) possess the ability for self-renewal, differentiation, and tumorigenesis and play a role in cancer recurrence and metastasis. CSCs are usually sorted in analysis into side population (SP) cells using ultraviolet (UV) laser (350 nm) excitation; they cannot be stained with Hoechst 33342 because of their efflux ability. However, it is difficult to avoid cell damage using a UV laser. Therefore, we attempted to isolate CSCs using a violet laser (407 nm) excitation to avoid cellular DNA damage. We sorted SP cells and main population (MP) cells from a human endometrial cancer cell line using the FACSAria system equipped with a violet laser and analyzed the biological properties of these cells. SP cells exhibited drug efflux, self-renewal, differentiation abilities, and tumorigenicity. It was found that v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) expression was significantly higher in SP cells than in MP cells. Our results suggest that CSCs exist in the SP fraction sorted using the FACSAria system equipped with a violet laser, which presents a useful tool to isolate small populations of viable putative CSCs from solid tumors and can be used to identify and characterize CSCs.

Effects of N-[N-(3, 5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT) on cell proliferation and apoptosis in Ishikawa endometrial cancer cells.

Endometrial cancer is one of the most common gynecological malignancies in Japan, where the disease shows an increasing morbidity. However, surgical therapy remains the treatment of choice for endometrial cancers that tend to be insensitive to radiation therapy and chemotherapy. Therefore, novel therapeutic strategies are required. The Notch signaling pathway regulates embryogenesis and cellular development, but deregulated Notch signaling may contribute to tumorigenesis in several cancers. Moreover, γ-secretase inhibitors have been shown to be potent inhibitors of the Notch signaling pathway; they suppress cellular proliferation and induce apoptosis in several cancer cells. In the present study, we investigated the effect of N-[N-(3, 5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT, γ-secretase inhibitor) on the cell proliferation and apoptosis in Ishikawa endometrial cancer cells. Real-time PCR detected mRNA derived from NOTCH1 and HES1, which are target genes of the Notch signaling pathway, in Ishikawa endometrial cancer cells. After blocking Notch signaling, cellular proliferation decreased, accompanied by increased expression of p21 mRNA and decreased expression of the cyclin A protein. Furthermore, blockade of Notch signaling induced apoptosis. These results suggest that the Notch signaling pathway may be involved in cell proliferation through cell cycle regulation and apoptosis in Ishikawa endometrial cancer cells. Inhibition of the Notch signaling pathway by γ-secretase inhibitors is expected to be a potential target of novel therapeutic strategies for endometrial cancer.

Mucinous breast carcinoma with myoepithelial-like spindle cells.

Appearance of spindle cells has been believed as a benign index of breast cytology. But, we have frequently observed the spindle cells in smears from mucinous carcinoma of the breast. Here, we characterized the biochemical nature of the spindle cells, so as to clarify their identity in cytology. Nineteen cases of breast mucinous carcinoma were used for cytological examination. The spindle cells were located at edges of tumor cell nests and in the backgrounds of cytological specimens. Immunohistological examination revealed that the spindle cells exhibited both immunoreactivity against carcinoembryonic antigen (CEA) and epithelial membrane antigen (EMA). Immunoreactivity against vimentin, cytokeratin, or alpha-smooth muscle actin was, however, not observed. The mode of distribution of biochemical markers suggests that the positive cells for anti-CEA antibody and anti-EMA antibody are tumor cells compressed by mucin, while the vimentin-positive cells are fibroblasts. We assert that the presence of spindle cells can be a characteristic feature of mucinous carcinoma of the breast. Discrimination of the spindle cells in mucinous carcinoma from myoepithelial cells and naked bipolar nuclei in benign lesions was established here. It should facilitate precise diagnosis of breast cancer.

[Unique system for education of medical technology and cytotechnology students at Kurashiki University of Science and the Arts].

For students orienting medical technologist (MT) and cytotechnologist (CT), a peculiar system of education is operated in Department of Life Science, Kurashiki University of Science and the Arts. Forty candidates for MT are selected from 90 students at end of the university first year. They study from lectures and practices specialized in medicine and face another selection at end of the third year. Selected candidates (ten and some in number) will challenge examination for CT license as well as that for MT license in the forth year in "double-license course". Prior to these selections, education for CT is started at the first year in order to clarify suitability of each student as a CT from earlier stage. In parallel, education for basic medical research is opened to all students at the first year for developing another suitability in them. Topics in cytopathology, neuroscience, cellular culture and regenerative medicine are presented to stimulate their interest in graduate school. Kake Educational Institution Cytopathological Center is located in the university campus and provides excellent facility for basic medical research. The university develops talent of students both for MT and CT through license acquisition and basic researcher for modern medicine in collaboration with this research center. The premier group of students under application of this system is active at the third year at present. They understand their own suitability and spend their time in the best manner toward their goal. We are confident that this system activates talent of students without overload.