BPB Reports

2022 - Vol. 5

2022 - Vol. 5

Amorphas Silica Nanoparticles Decrease Human Chorionic Gonadotropin β Expression During Syncytialization of BeWo Cell Vol.5, No.6, p.154-158
Yuji Sakahashi , Rena Yamamoto , Go Kitahara , Rina Izutani , Hirofumi Tsujino , Yuya Haga , Kazuma Higashisaka , Yasuo Tsutsumi
Received: October 24, 2022
Accepted: December 08, 2022
Released: December 27, 2022
Abstract Full Text PDF[852K]

Nanoparticles are used in many everyday products because of their innovative properties, but there are concerns that, being smaller than conventional materials, they have the potential to induce unexpected biological effects. Despite the need for the safety of nanoparticles to be assessed, action in this area is still lagging. Several studies, including those by us, have revealed that nanoparticles can reach the placenta and show placental toxicity, but it remains unclear how nanoparticles affect placental function. Here, we attempted to assess the effect of nanoparticles on placental hormone-producing function by using an in vitro forskolin-induced BeWo syncytialization model. BeWo cells were treated with amorphous silica nanoparticles with a diameter of 10 nm (nSP10) either during syncytialization or after being syncytialized by forskolin treatment. RT-PCR analysis showed that nSP10 inhibited forskolin-induced upregulation in the expression level of human chorionic gonadotropin β (hCGβ; gene name, CGB) during syncytialization, but not in syncytialized BeWo cells. Moreover, nSP10 downregulated forskolin-induced elevation in the expression of endogenous retrovirus group FRD member 1 (ERVFRD-1) and syndecan-1 (SDC1), which are involved in cell fusion during syncytialization. These results suggest that nSP10 could suppress trophoblast cell fusion, thus inhibiting the production of hCG in syncytialized BeWo cells.

Regular Article
Effects of Keishi-Bukuryo-gan-ka-Yokuinin on Rosacea Model Mice Vol.5, No.6, p.147-153
Natsu Watanabe , Wataru Matsunaga , Akinobu Gotoh
Received: October 31, 2022
Accepted: November 29, 2022
Released: December 15, 2022
Abstract Full Text PDF[2M]

Rosacea is an intractable skin disease and no effective treatment has been established, but there are cases of improvement with Kampo medicine. In this study, the efficacy of Keishi-Bukukryo-gan-ka-Yokuinin (KBY) in ameliorating the symptoms of rosacea was investigated in a steroid-induced rosacea mouse model. The mice were fed a KBY-containing diet for 28 d, after application of clobetasol propionate for 10 d; thereafter, the velocity of capillary blood flow, area of the skin purpura, histopathologic features, and gut microbiota were analyzed. The KBY-fed mice showed remarkable recovery in the capillary blood flow velocity and purpura area, as well as restoration of the epidermis features. Furthermore, gut microbiome analysis showed the enhancement of Lactobacillus population in the KBY-fed mice. The KBY-induced improvement of circulatory disturbances in skin might contribute to the alleviation of telangiectasia and erythema. Lactobacillus may be involved in alleviating the rosacea-like symptoms in association with KBY; however, this association should be investigated further by using other analysis methods. Our results provide evidence of the effectiveness of KBY in improving the pathology of rosacea.

Regular Article
Prevalence of Abnormal Blood Pressure and the Association between Blood Pressure and Anthropometric Measures or Body Indices in Japanese University Students- a Cross-Sectional Study Vol.5, No.6, p.140-146
Kaito Yamashiro , Kazunori Sagawa , Shigeharu Tanei , Fumihiko Ogata , Takehiro Nakamura , Naohito Kawasaki
Received: October 26, 2022
Accepted: November 21, 2022
Released: December 15, 2022
Abstract Full Text PDF[1M]

Adolescent blood pressure (BP) is associated with the prevalence of hypertension in later life, and anthropometric measures have been shown to be useful indicators for predicting hypertension. However, little is known about their associations in Japanese young adults. Here, this study aimed to analyze the association between BP and anthropometric measures in Japanese university students. The participants of this cross-sectional study included 17,512 university students (12,559 males and 4,953 females) in Japan between 2011 and 2019. Abnormal BP (ABP) was defined as systolic/diastolic BP above 120 and/or 80 mmHg. ABP was seen in 59.2% of males and 16.3% of females, with the proportion in male students increasing over the nine year study period. Areas under receiver operating characteristics curves analysis showed that the top three predictors (and their cut-off points) of ABP in descending order were body mass index (20.5 kg/m2), waist circumference (74.1 cm) and chest circumference (85.4 cm) in males, and body mass index (19.8 kg/m2), body fat rate (28.5%) and chest circumference (82.4 cm) in females. The odds ratio was 2.69 (2.45‒2.94) in males and 2.40 (1.96‒2.95) in females when all three measurements were higher than their cut-off points. The incidence of ABP has increased in male university students, suggesting the need for interventions for appropriate BP control. This study suggests that anthropometric measures in combination with body mass index might be useful for this intervention in Japanese university students.

Regular Article
Malnutrition-Induced Involution of Lymph Nodes in Mice Vol.5, No.6, p.133-139
Hiroshi Hasegawa , Kei Nakayama
Received: October 19, 2022
Accepted: November 04, 2022
Released: December 02, 2022
Abstract Full Text PDF[4M]

Immune suppression is one of the major factors affecting the pandemic outbreak of infectious diseases in societies where malnutrition is common. The thymus and spleen are known to respond to starvation via reductions in their size and functions, which is called thymic or splenic involution. However, almost no reports have been published on the response of lymph nodes, other secondary immune organs, to starvation. Therefore, we here examined the histological characters of lymph nodes of a mouse dietary restriction model. Dietary restriction for 48 h reduced the size of inguinal lymph nodes by 48%. Immunoreactivity to anti-immunoglobulin G antibody was reduced by the dietary restriction, along with a normal level of immunoglobulin M-immunoreactivity, suggesting inhibited immune capacity. Because splenic involution involves macrophages, we immunostained the lymph node sections to detect macrophages. The immunoreactivity to anti-ionized calcium-binding adaptor molecule 1 (IBA1)/allograft inflammatory factor 1 (AIF1) antibody was not changed by the dietary restriction. In contrast, anti-F4/80 antibody reactivities in medullary cord macrophages, interfollicular macrophages, and the macrophages located along trabeculae in the subcapsular sinus were reduced by the dietary restriction. These results indicate that, in addition to thymus and spleen, lymph nodes are also susceptible to starvation. Specific subpopulations of macrophages are reduced in the lymph nodes of starved mice.

Regular Article
Effectiveness of Community Pharmacist-Led Intervention for Patients with Dyslipidemia Using the Rubric-Based Intervention Program: A Randomized Controlled Trial Vol.5, No.6, p.125-132
Hisateru Ueki , Masayuki Ohbayashi , Toshinori Yamamoto , Tsuyoshi Inoue , Mari Kogo
Received: October 03, 2022
Accepted: November 01, 2022
Released: November 30, 2022
Abstract Full Text PDF[992K]

Background: A rubric-based intervention program was designed to reduce the risk of developing coronary artery disease (CAD) in patients with dyslipidemia, and the effectiveness of community pharmacist-led intervention using the program was evaluated. Methods: We conducted an open-label, multicenter, randomized controlled trial. Participants included patients with dyslipidemia on statin medication recruited from August 2020 to July 2021. The intervention group received instructions from community pharmacists using a rubric-based intervention program in addition to usual pharmacist care. Conversely, the control group only received the usual care. The primary outcome was the change in the Suita score at month 6, which was compared between the intervention and control groups. The secondary outcome was the change in the rubric score, which was compared between the post-intervention and baseline. Results: We analyzed 15 participants (mean age, 61.9 years), whose mean Suita score was 42.6. No significant difference was observed in the change in the Suita score between the two groups. However, in the intervention group, the post-intervention Suita score was significantly lower than the baseline (43.9 vs. 38.7, p = 0.01), and the CAD risk classification based on Suita score decreased from medium to low risk. The post-intervention rubric score increased significantly compared with the baseline (2.0 vs. 3.1, p < 0.01). Conclusion: The rubric was a useful tool in promoting behavior changes in patients. The community pharmacist-led intervention for patients with dyslipidemia using a rubric-based intervention program showed the possibility of reducing the risk of developing CAD.

Oxidative Stability and Antioxidant Activity of Crude Jojoba Oil Vol.5, No.6, p.121-124
Suguru Tsuchimoto , Hiroe Sakai , Kiichi Fukui
Received: August 04, 2022
Accepted: October 01, 2022
Released: November 24, 2022
Abstract Full Text PDF[942K]

The seeds of jojoba [Simmondsia chinensis (Link) Schneider] contain a unique oil known as jojoba oil. It mainly consists of liquid wax monoesters with structures similar to human sebum wax and is popular as a cosmetic ingredient. We previously observed that the oxidative stability and antioxidant activity of crude jojoba oil are higher than those of many other vegetable oils. In this study, these two parameters were determined and compared among crude jojoba oils from different companies, countries, and years of production, together with deodorized or refined oils. Oxidative stability and antioxidant activity vary among crude oils, and there was a negligible correlation between these two parameters. Both deodorized and refined oils showed lower antioxidant activities than crude oils. In addition to wax esters, triglycerides and squalene are other major oil components of human sebum. To compare the autooxidation of jojoba oil (wax ester), olive oil (triglyceride), and squalene, we measured their acid, peroxide, and carbonyl values after heat treatment at 60°C for 60 d. The acid value did not change in jojoba oil but increased in the other ones. In addition, the peroxide and carbonyl values were the lowest in jojoba oil following heat treatment. These results suggest that jojoba oil was the most stable in terms of autooxidation among these three investigated oils. Finally, we determined the cytotoxicity of olive and jojoba oils in human epidermal cells, and concluded that they were non-toxic after heating at 60°C for 30 or 60 d.

Evaluation of Conversion Factor for Rapid Quantification of Authorized Genetically Modified Maize and Soybean in Japan Vol.5, No.5, p.115-120
Norihito Shibata , Keisuke Soga , Miyu Sugino , Jumpei Narushima , Satoko Yoshiba , Tomohiro Egi , Reona Takabatake , Kazunari Kondo
Received: September 26, 2022
Accepted: September 28, 2022
Released: November 09, 2022
Abstract Full Text PDF[3M]

With the increasing development of genetically modified (GM) crops authorized for use in food, a rapid and accurate method of quantifying the weight-based amount of GM crops is needed to ensure consumers’ rights to choose. Conversion factor (Cf) value is the ratio of the copy number of a GM-specific sequence to an endogenous sequence in the GM crop and is used to convert a copy number ratio of the GM-specific sequence to the endogenous sequence of a sample into weight-based amount of GM crops. However, in the current Japanese official method for GM crops, determining Cf values using real-time PCR instruments capable of rapid measurements has not been established. In this study, Cf values for GM maize and soybean authorized for use as food in Japan were experimentally determined using an Applied Biosystems 7500 Fast Real-Time PCR System, which is capable of rapid measurement. The Cf values were almost the same as those of the PCR instruments described in the Japanese official method, and the weight-based amount of GM maize MON810 measured using this Cf value showed similar results. These results suggest that rapid quantification by this PCR instrument has the same performance as the recommended PCR instruments and may contribute to the labeling regulation of GM crops in Japan.

Regular Article
Advanced Recombinant Cascade Reagent PyroSmart NextGen® for Bacterial Endotoxins Test as Described in the Pharmacopeias Vol.5, No.5, p.105-114
Ingrid Stevens , Norihiko Ogura , Madeline Kelley , Robert L. D’Ordine , Hikaru Mizumura , Toshio Oda , Junko Akiyoshi , Edwin G. Jahngen
Received: August 26, 2022
Accepted: September 22, 2022
Released: October 28, 2022
Abstract Full Text PDF[3M]


The bacterial endotoxins test (BET) is a method of detection and quantification of bacterial endotoxin in injectable drugs and medical devices using amoebocyte lysate reagents sourced from the horseshoe crab (Limulus polyphemus or Tachypleus tridentatus). Three manufacturers have launched three different types of recombinant reagents, and recently the United States, European and Japanese Pharmacopeias have been evaluating the equivalency of these reagents to natural amoebocyte lysate reagents. Several studies suggested that the recombinant reagents are very similar to amoebocyte lysate reagents, however there is potential for improvement. Previous reports indicate that recombinant reagents have two issues: extremely low endotoxin potency determination for Helicobacter pylori GU2 and high levels of interference from Heparin Calcium impacting endotoxin recovery. A new recombinant cascade reagent (rCR), PyroSmart NextGen®, recently introduced to the market has been developed to solve these issues. PyroSmart NextGen® has demonstrated higher reactivity to H. pylori GU2 and a lower level of interference from Heparin Calcium than other existing recombinant reagents. Additionally, the analytical capability and suitability of PyroSmart NextGen® has been demonstrated when applied to the BET as described in the US, European and Japanese pharmacopeias. PyroSmart NextGen® has also shown comparability to amoebocyte lysate reagents by demonstrating its ability to detect autochthonous endotoxin in water and in Escherichia coli culture. Overall, this study has verified that the rCR, PyroSmart NextGen® is a suitable alternative to amoebocyte lysate reagents.

Importance of Glycine Preceding Pivotal Tyrosine in the Lysosome-Targeting Signal GYQTI of Lysosome-Associated Membrane Protein-1 (LAMP-1) Vol.5, No.5, p.99-104
Fumiaki Yamaguchi , Hiroshi Sakane , Yosuke Morishita , Toshiyuki Hata , Kenji Akasaki
Received: June 24, 2022
Accepted: August 07, 2022
Released: September 21, 2022
Abstract Full Text PDF[2M]

Lysosome-associated membrane protein-1 (LAMP-1) is a type I membrane glycoprotein consisting of a large luminal domain, a membrane-anchoring domain, and a short cytoplasmic tail (CT). The tyrosine-based motif (G378Y379QTI382) in its CT exclusively binds to adapter protein complex-3 (AP-3), which may facilitate the incorporation of LAMP-1 into transport vesicles to late endosomes and lysosomes. Of this sequence, Y379 is critical, and hydrophobic I382 is optimal for the AP-3 binding and the efficient delivery to lysosomes. However, it is not clear how important G378 is in the AP-3 binding and lysosome transport. To clarify its importance, four mutants in which G378 was replaced with alanine, aspartic acid, glutamic acid, and asparagine (designated as G378A, G378D, G378E, and G378N, respectively) were prepared and their interaction strengths with AP-3 and lysosomal abundance were compared with those of wild-type (WT)-LAMP-1. A yeast two-hybrid system was applied to measure the interaction strengths of WT-, G378A-, G378D-, G378E-, and G378N-CTs with the medium subunit of AP-3 (μ3A). The G378A-, G378D-, and G378E-CTs as strongly interacted with μ3A as WT-CT, but the G378N-CT exhibited a very weak interaction with it. In the cell fractionation analyses, the lysosomal levels of G378A, G378D, and G378E were almost the same as that of WT whereas a lesser amount of G378N existed in the lysosomes. Taken together, it is considered that Y379 and I382 are essential for AP-3-mediated vesicular transport of LAMP-1 while the Y379-preceding amino acid is not restricted to glycine but asparagine at this position is less suitable for that.

Anti-Obesity Dendritic Cell Vaccine Therapy Targeting Neovascular Vessels in High-Fat-Diet-Fed Mice Vol.5, No.5, p.95-98
Yoshikazu Sawaguchi , Keiichi Hirata , Ryo Suzuki , Kazuo Maruyama , Naoki Utoguchi
Received: May 21, 2022
Accepted: August 05, 2022
Released: August 30, 2022
Abstract Full Text PDF[1M]

Obesity is a severe disease that causes diabetes, fatty liver, hypertension, and arteriosclerosis. It involves adipocytokines such as tumor necrosis factor-α, leptin, and resistin.The hypertrophic adipocytes induce the secretion of these adipocytokines. Regulating the growth of adipose tissue is an extremely important approach to prevent these diseases. Hypertrophy of adipose tissue is closely associated with angiogenesis, wherein neovascular vessels supply nutrients.Previous studies have shown that angiogenesis inhibitors can prevent the development of obesity in mice. In this study, we report an anti-obesity therapy targeting the neovascular vessels of adipose tissue via endothelial cell (EC) vaccination. EC vaccine was prepared using EC extract antigen, which was pulsed to dendritic cells, and was administered thrice intradermally. The anti-obesity effect was evaluated in high-fat diet-fed obesity model mice. EC vaccination significantly suppressed the increase in body weight and hyperplasia of adipocytes and improved the accumulation of hepatic lipid droplets in these mice. Adipose tissue-induced vascular-targeted vaccine therapy is a potential novel approach for obesity.

Corrigendum: Distribution of 58 Semi-Volatile Organic Chemicals in the Gas Phase and Three Particle Sizes in Indoor Air and House Dust in Residential Buildings During the Hot Season in Japan
[Notice] The Original Article was published on December 04, 2019
Vol.5, No.4, p.94-94
Shinji Takeuchi , Toshiko Tanaka-Kagawa , Ikue Saito , Hiroyuki Kojima , Hideto Jinno
Received: July 14, 2022
Accepted: July 28, 2022
Released: July 29, 2022
Abstract Full Text PDF[596K]

Interleukin-3 Potentiates Murine Basophils for Protease Allergen-Induced Interleukin-4 Production Vol.5, No.4, p.88-93
Arisa Morikawa , Yuka Matsui , Takuma Kitano , Saotomo Itoh , Shinsuke Taki , Shigeaki Hida
Received: June 23, 2022
Accepted: June 24, 2022
Released: July 29, 2022
Abstract Full Text PDF[1M]

The cysteine protease papain was shown to induce production of IL-4 and other cytokines in murine basophils, a critical event for initiating and promoting type 2 immune responses. However, the papain ‘sensor(s)’ and the intracellular signaling pathway for IL-4 production triggered by the protease remained unknown. Here we show that basophils lacking FcRγ or expressing dominant negative Syk failed to produce IL-4 in response to papain, indicating the involvement of the FcRγ-Syk pathway that was essential also for IL-3-induced IL-4 production. While IL-3, IgE cross-linkage and papain induced production of IL-4, IFN-β production was induced only by papain, indicating that papain sensor(s) or its downstream signaling pathways for production of IL-4 seemed to be distinct from those for IFN-β production. We further showed using the artificial papain sensor CD8-FcRγ fusion protein that IL-3 potentiated, independently of FcRγ, basophil response to FcRγ-dependent papain signals. Thus, our current study showed that papain sensing and/or downstream signaling are unique for cytokines to be induced and regulated through cross-talk with other signals such as IL-3 signals.

Comparison of Helium-Alternative Carrier Gases for Gas Chromatography/Mass Spectrometry of Standard Test Methods for Indoor Air Quality Guidelines in Japan Vol.5, No.4, p.84-87
Naohiro Oshima , Mihono Takagi , Shinobu Sakai , Yoshiaki Ikarashi
Received: June 14, 2022
Accepted: July 04, 2022
Released: July 20, 2022
Abstract Full Text PDF[1M]

Helium is the most frequently used carrier gas for GC/MS, which is the official standardized test method in Japan to assess chemical substances in indoor air. However, recent global challenges in the supply chain for helium have led to a need to validate GC/MS using alternative carrier gases. In this study, we examined the applicability of hydrogen and nitrogen as helium-alternative carrier gases in the standardized GC/MS analytical test method for volatile organic compounds (VOC) and phthalate esters in indoor air. Comparison of the signal-to-noise ratios of standard solutions showed that detection sensitivities of hydrogen and nitrogen analysis were enough for the standard test method, although these gases, especially nitrogen, were less sensitive than helium. Measurements using these alternative carrier gases showed good linearity and could quantify around 1/100th of Japanese guideline values for indoor air concentrations. Therefore, hydrogen and nitrogen gases can be applied to the standard GC/MS analysis test method for VOC and phthalate esters in indoor air as alternative carrier gases to helium.

Establishment of an Easy and Convenient Anti-SARS-CoV-2 Neutralizing Antibodies Test Flow Starting from Community Pharmacies: Expansion of Pharmacy Functions Suitable During the Novel Coronavirus Era Vol.5, No.4, p.80-83
Kazuyuki Niki , Kozo Tada , Katsumi Suezawa , Tomoko Kasashima , Shin Fujisue , Eri Seichi , Miki Nakahara , Yoshiko Ohara , Akiko Yoshida , Mikiko Ueda
Received: June 14, 2022
Accepted: June 23, 2022
Released: July 12, 2022
Abstract Full Text PDF[977K]

The third vaccination for coronavirus disease 2019 is recommended; however, the vaccination rate is not increasing. This is due to a lack of information on the effectiveness of the booster vaccination and the fact that some people assume that they have acquired sufficient immunity after the second vaccination. Therefore, this study examined whether a system could be established in which a community pharmacy could act as a starting point to conduct antibody tests. The study was conducted between November 18, 2021 and February 28, 2022. Thirty-eight subjects collected their own samples in the Greenmedic pharmacy or at home using a self-blood collection kit and submitted them to the pharmacy. Samples were transported from the pharmacy to the AC clinic within 2 days for measuring anti-SARS-CoV-2 neutralizing antibodies (NAbs). There were eight men and 30 women with a mean age (mean ± standard deviation) of 33.6 ± 15.3 years. Of the subjects, 12 had completed the third vaccination; the median (interquartile range) blood NAbs after the second vaccination was 34.9 (23.3–59.4) AU/mL, which increased to 994.1 (974.7–1042.1) AU/mL after the third vaccination (p = 0.0022). Significant negative correlations were also observed for both blood NAbs and age after the second vaccination (rs = −0.579, p = 0.00014). Since these results were similar to those of previous studies conducted at hospitals, this study seems to be the first to demonstrate that anti-SARS-CoV-2 neutralizing antibody test flow can be established at a community pharmacy.

Effects of the Interplay between Selenocystine and Methylmercury on Their Cytotoxicity and Glucose-Driven Insulin Secretion from Mouse Insulinoma Cells Vol.5, No.4, p.74-79
Daichi Chida , Takashi Toyama , Takanori Chiba , Takayuki Kaneko , Kotoko Arisawa , Yoshiro Saito
Received: April 26, 2022
Accepted: June 27, 2022
Released: July 12, 2022
Abstract Full Text PDF[1M]

We’ve previously shown that high levels of selenoprotein P (SeP), a major selenoprotein in plasma, can be a risk factor of type 2 diabetes. It was also thought that inhibition of insulin secretion caused by over-supplementation of selenium by SeP to pancreatic β cells contributed to the progress of diabetes. On the other hand, methylmercury, which is an environmental pollutant, is known to cancel the action of selenium via the covalent modification. Therefore, we thought that the interaction between selenium and methylmercury could be associated with the pathogenesis of diabetes. To address the hypothesis, MIN6 cells, a mouse pancreatic β-cell line, were treated with selenocystine (as a selenium donner) and methylmercury then examined insulin release from the cells. Selenocystine (400–1200 nM), which corresponds to the concentration of selenium in SeP of diabetic patients, shows cytotoxicity and inhibited glucose-driven insulin secretion. Methylmercury rescued the cytotoxicity that induced by selenocystine, however it affected the insulin secretion that is depressed by selenocystine at little intense. These data indicate that the mechanisms underlying inhibition of insulin secretion by selenocystine are independent of cytotoxicity, and methylmercury cannot be expected to restore insulin secretion or suppress diabetes as selenium neutlizer.

The Cases on the Application of Touch Panel Type Screening Check for Early Detection of Dementia Conducted at a Community Pharmacy Near a Community Medical Hospital to an Advanced Treatment Hospital Vol.5, No.3, p.70-73
Hiroshi Akura , Mamiko Kai , Yoshio Soda , Takafumi Nakano , Toshinobu Hayashi , Hitoshi Iida , Koji Ogomori , Hiroaki Kawasaki , Masanobu Sonoda , Masashi Takahashi , Takashi Egawa
Received: May 01, 2022
Accepted: June 22, 2022
Released: July 06, 2022
Abstract Full Text PDF[1M]

The purpose of this study is to construct the consultation system based on the touch panel type screening check for dementia (TPCD) results from a pharmacy to psychiatric specialists at an advanced treatment hospital, and to evaluate the effectiveness of resulting in dementia diagnosis. We herein report three cases that demonstrate the significance of TPCD in the course of study. Cases 1 was successful case from a 78-year-old man in which pharmacists recommended individuals scored 11 on the TPCD to visit psychiatrists at Fukuoka University Hospital, an advanced medical hospital, based on the results of TPCD. He was practically diagnosed with mild cognitive impairment (MCI). Cases 2 was partially successful case from a 78-year-old woman. Her TPCD score resulted only 9. The pharmacist effectively utilized tracing report to notify the patient’s cardiologist about the results of the TPCD performed at the pharmacy as she wished. On the other hand, Case 3 showed that patient whose TPCD score resulted 2 could receive no treatment due to advanced stage of dementia despite our consulting with family doctor based on the TPCD scores. These cases suggest that the pharmacy near an advanced medical hospital should play a role to provide accessibility between the patients with suspected dementia and psychiatric specialists and make opportunities to consult specialists earlier before progressing towards a dementia following MCI collaborated with community interprofessionals.

Critical Residues of Clostridium perfringens Delta-Toxin for Oligomerization and Receptor Binding Vol.5, No.3, p.66-69
Soshi Seike , Masaya Takehara , Keiko Kobayashi , Masahiro Nagahama
Received: May 16, 2022
Accepted: June 21, 2022
Released: July 06, 2022
Abstract Full Text PDF[1M]

Delta-toxin produced by Clostridium perfringens types B and C is a β-pore-forming cytotoxin. Here, using site-directed mutagenesis, we identified the amino acid residues that contribute to delta-toxin oligomerization and binding. We replaced Lys-43 and Ser-109 located in the β-sandwich domain and Arg-200 located in the rim domain. Substitution of alanine for Ser-109 caused reductions in both cytotoxicity and oligomerization. However, exchange of Lys-43 for alanine resulted in a reduction in the cytotoxicity. Replacement of Arg-200 with alanine led to drastic reductions cytotoxicity and cell binding. Our results demonstrate that Ser-109 plays a role in oligomerization, and that Arg-200 is critical for binding of the toxin.

Regular Article
Potential Ameliorative Effects of Kampo Medicines Ninjin’yoeito and Kamikihito on Frailty-Like Behavior in Naturally Aged C57BL/6J Mice Vol.5, No.3, p.59-65
Masahiro Tabuchi , Yosuke Matsubara , Shinji Miyazaki , Keita Mizuno , Yuji Omiya , Yoshiki Ikeda
Received: March 30, 2022
Accepted: June 03, 2022
Released: June 22, 2022
Abstract Full Text PDF[1M]

Frailty is defined as an age-related decline in physiological reserve and increased vulnerability to stress. As frailty is a multifaceted condition, there is no effective pharmacotherapy for it yet. Ninjin’yoeito (NYT) and Kamikihito (KKT), traditional Japanese medicines (Kampo medicines), are promising in treating multifaceted conditions of frailty including fatigue and mental anxiety. However, their effects are still unclear. In this study, the effects of NYT and KKT on different types of frailty in naturally aged mice were explored by survival, physical aspects (Frailty Assessment Scores and muscle strength measurements), psychological aspects (sucrose splash test for motivation-related behavior), and social aspects (rescue-like behavior test for prosocial behavior). Mice aged 22-months were fed a diet containing 3% NYT or 3%KKT for 13 weeks until the age of 25 months. Behavioral alterations in old mice were compared with those in adult mice (five months old). Throughout the study period, Old-control mice showed frailty-like symptoms, including elevation of frailty assessment score, reduction of muscle strength and motivation for self-care, and rescue-like behavior compared to adult mice. NYT increased the survival rate of old mice and suppressed the declines in their frailty assessment score, muscle strength, and motivation for self-care. KKT reduced decreases in the frailty assessment score, motivation for self-care, and rescue-like behavior in old mice. These results suggest that NYT and KKT alleviate general frailty-like symptoms in old mice. Additionally, NYT may extend lifespan. These findings suggest that NYT and KKT may be helpful for improving the multifaceted symptoms of frailty.

Regular Article
Distribution, Metabolism, Excretion, and Lactational Transfer to Pups of Tetrabromobisphenol A and Its Metabolites in C57BL/6 Mice Vol.5, No.3, p.50-58
Teruyuki Nakao , Hideki Kakutani , Tomohiro Yuzuriha , Souichi Ohta
Received: December 24, 2021
Accepted: May 20, 2022
Released: June 07, 2022
Abstract Full Text PDF[2M]

We characterized the absorption, distribution, metabolism, and excretion of tetrabromobisphenol A (TeBBPA) in C57BL/6 mice. After TeBBPA dosing, we monitored the mice and collected samples for 24 h. Most TeBBPA was excreted in the feces at 12 h. At 24 h, 71% of administered TeBBPA was found in the feces and TeBBPA-conjugates were detected at 17.8%. We also detected debrominated metabolites of TeBBPA, including tribromobisphenol A (TriBBPA), dibromobisphenol A (DiBBPA), and monobromobisphenol A (MoBBPA). TeBBPA, TriBBPA, 2,2′-DiBBPA and MoBBPA were detected in dam’s blood between 30 min and 1 h. Moreover, TriBBPA and 2,2′-DiBBPA were detected in milk at 30 min. The primary metabolites of TeBBPA were formed by conjugation, whereas debromination represented a minor metabolism pathway. Moreover, our findings demonstrated that the parent TeBBPA compound and its debrominated metabolites were distributed into the maternal milk of mice and transferred to the nursing pups following TeBBPA oral administration.

Regular Article
Optimization of Milling Parameters for Low Metal Contamination in Bead Milling Technology Vol.5, No.3, p.45-49
Hironori Tanaka , Yuya Ochii , Yasushi Moroto , Daisuke Hirata , Tetsuharu Ibaraki , Ken-ichi Ogawara
Received: March 31, 2022
Accepted: May 20, 2022
Released: June 07, 2022
Abstract Full Text PDF[767K]

This study aimed to develop a contamination-less bead milling technology using zirconia beads, by optimizing milling parameters. We evaluated the effects of bead milling parameters on milling time and metal contamination; focusing on bead diameter, rotation speed, and bead filling rate as they are the critical parameters determining bead milling efficiency. We studied the milling time required to grind a drug to 0.2 μm size and quantified the extent of metal contamination that arises from the grinding procedure. With optimal bead milling parameters, the minimum concentration of the metal contaminants was 1.27 ± 0.08 μg/mL (zirconium: 0.73 ± 0.09 μg/mL; yttrium: 0.35 ± 0.03 μg/mL; aluminum: 0.19 ± 0.05 μg/mL), when the rotation speed, bead diameter, and bead filling rate were set to 2 m/s, 0.3 mm, and 75% (v/v), respectively. Additionally, under optimized conditions, it was possible to reduce the metal contamination per weight of the drug by increasing the drug concentration to up to 40% (w/w). Under the optimized conditions for drug concentrations of 30–40% (w/w), metal contamination from the grinding process was minimized and reduced to < 10 μg/g, which is comparable to that achieved by the NanoCrystal® technology. These results indicate that contamination-less bead milling using zirconia beads can be achieved when bead milling parameters are optimized.

The Potential of Virgin and Calcined Gibbsite for the Removal of Dyes from Aqueous Media Vol.5, No.3, p.42-44
Fumihiko Ogata , Yugo Uematsu , Ayako Tabuchi , Takehiro Nakamura , Naohito Kawasaki
Received: May 09, 2022
Accepted: May 10, 2022
Released: June 07, 2022
Abstract Full Text PDF[760K]

Virgin gibbsite (GB) and calcined gibbsite (denoted as GB200, GB400, GB600, GB800, and GB1000), were prepared at 200°C, 400°C, 600°C, 800°C, and 1000°C, and the characteristics of prepared GB samples were investigated. The amount of the hydroxyl group, the specific surface area, and the micropore volume of GB400 were higher than those of other GB samples. Additionally, the crystal structure of GB changed significantly from the gibbsite phase to transitional states at our calcination temperatures. Furthermore, we showed the adsorption capacity of various dyes using GB samples. The prepared GB400, GB600, and GB800 showed adsorption capacity of Red, Blue, and Green, respectively. However, other GB samples did not show a dye adsorption capacity. Moreover, the adsorption mechanism of Red and Blue using GB400 and GB600 was related to the number of hydroxyl groups, specific surface area, and micropore volumes in this study. Finally, the relationship between the amount of Red and Blue adsorbed and the number of total organic carbons decreased was positively correlated with the values of 0.974 and 0.935, respectively. In our prepared GB sample, both dyes and organic carbons were simultaneously removed during the adsorption treatment.

Analysis of the Possibility of Drug-Induced Aspiration Pneumonia Due to Anxiolytics by Using the Japanese Adverse Drug Event Report Database Vol.5, No.3, p.39-41
Rei Nakayama , Hideki Nawa , Takayoshi Maiguma , Yuichi Tasaka
Received: March 06, 2022
Accepted: May 02, 2022
Released: June 01, 2022
Abstract Full Text PDF[674K]

With societal aging, the number of patients with aspiration pneumonia is increasing. However, because the relationship between drug use and the development of aspiration pneumonia is not fully understood, improvements in information on the possibility of drug-induced aspiration pneumonia are urgently needed. Hence, in this study, we investigated the relationship between the use of anxiolytics and the development of aspiration pneumonia by using data from the Japan Adverse Drug Event Report (JADER) database. We found that anxiolytics had a signal for the development of aspiration pneumonia with a reporting odds ratio (ROR) of 3.2 (95% confidence interval: 2.5–4.1), and seven of eight anxiolytics for which the development of aspiration pneumonia was reported in the JADER database had a signal as well. Of note, the possibility of the development of aspiration pneumonia was mentioned only in the package inserts of clobazam among the package inserts or risk management plans (RMPs) of these anxiolytics. These results suggest the need for including information on the possibility of aspiration pneumonia development in drug package inserts and RMPs so as to prevent anxiolytic-associated aspiration pneumonia or facilitate its early detection.

Regular Article
Effect of Using Concomitant Drugs on the Efficacy of Sodium Polystyrene Sulfonate Vol.5, No.3, p.33-38
Yugo Uematsu , Masashi Yanae , Manabu Akegami , Fumihiko Ogata , Takehiro Nakamura , Naohito Kawasaki
Received: April 08, 2022
Accepted: May 06, 2022
Released: June 01, 2022
Abstract Full Text PDF[838K]

Sodium polystyrene sulfonate (SPS) is well used for hyperkalemia. A recent study has shown that SPS may bind to other drugs in the digestive tract. However, there are few reports about the effect of concomitant drug use on serum potassium level variations. Therefore, this study aimed to investigate the effect of concomitant drug use on the variation of serum potassium levels among patients taking SPS. In total, 632 patients were newly prescribed with SPS from 2017 to 2019, and 186 patients were evaluated in this study. Further, the association between increase in serum potassium levels and concomitant drug use was investigated. We classified patients into the Grade 1 (G1) group and the Grade 2–4 (G2–4) group according to baseline serum potassium level by Common Terminology Criteria for Adverse Events (CTCAE) v5.0. There was the significant decrease in serum potassium and chloride levels and the increase in serum sodium levels after SPS treatment. In addition, therapy with SPS might improve renal function. The concomitant use of imidapril in the G1 group (odds ratio: 4.4, 95% confidence interval: 1.1–11.7, p = 0.0394) and nifedipine in the G2–4 group (odds ratio: 7.3, 95% confidence interval: 1.5–35.5, p = 0.0139) were significantly associated with the increase in serum potassium levels after SPS treatment. These associations might be affected by not only adverse drug reactions but also binding of other drugs to SPS. Hence, concomitant drug use may affect the efficacy of SPS.

Regular Article
Effect of Acylated Sugar Moieties on Gastrointestinal Absorption of Mono-Acylated Anthocyanins in Rats Vol.5, No.3, p.24-32
Takashi Ichiyanagi , Masayuki Nashimoto , Hiroki Hayashi , Norihiko Terahara
Received: February 01, 2022
Accepted: April 23, 2022
Released: May 20, 2022
Abstract Full Text PDF[1M]

In this study, five types of mono-acylated anthocyanins were isolated from colored-fleshed potato (Solanum tuberosum L.) and black carrot (Daucus carota subsp. sativus), and their gastrointestinal absorption was evaluated based on the plasma profiles of rats. The absorption amounts of mono-acylated anthocyanins were slightly higher or at least similar to those of non-acylated anthocyanins, which carry the same aglycone, when the purified anthocyanins were orally administered. The absorption amounts of petunidin 3-O-(6′′-O-(4′′′-O-p-coumaroyl-α-L-rhamnopyranosyl)-β-D-glucopyranoside)-5-O-β-D-glucopyranoside isolated from purple-fleshed potato were 1.3 to 2.3-fold higher than those of three types of cyanidin 3-O-(2′′-xylopyranosyl-6′′-O-(6′′′-O-acyl-β-D-glucopyranosyl))- β-D-galactopyranosides isolated from black carrot, whereas pelargonidin 3-O-(6′′-O-(4′′′-O-p-coumaroyl-α-L-rhamnopyranosyl)-β-D-glucopyranoside)-5-O-β-D-glucopyranoside isolated from red-fleshed potato exhibited an exceptionally poor absorption profile, as demonstrated by the comparison of the area under the plasma concentration curves during 8 h after oral administration normalized to the orally administered dose. The gastrointestinal absorption of mono-acylated anthocyanins was tended to determine by their structures, including aglycone moieties, attached sugars, and the balance of the entire molecule. Our findings suggested that the branched acylated sugars attached to position 3 tended to suppress the absorption of mono-acylated anthocyanins from the gastrointestinal tract. Furthermore, the poor intestinal absorption of mono-acylated anthocyanins in the mixtures was likely due to the competitive absorption of these molecules with contaminants that co-occur in plant materials such as organic acids.

Effect of Fluconazole on Blood Pressure in Patients Treated with Dihydropyridine Calcium Channel Blockers: A Retrospective Study Vol.5, No.2, p.21-23
Takeru Takahashi , Yasuharu Goto
Received: February 10, 2022
Accepted: March 26, 2022
Released: April 05, 2022
Abstract Full Text PDF[790K]

Fluconazole and dihydropyridine calcium channel blockers (DCCBs) are simultaneously used in clinical practice. Although fluconazole can increase the blood levels of DCCBs by inhibiting CYP3A4, there are only a few reports regarding the effects of interaction of these drugs on blood pressure. Based on electronic medical records, we conducted a retrospective study of blood pressure in hospitalized patients treated with fluconazole while receiving amlodipine and nifedipine. The mean blood pressure over 3 days, specifically 2 days before starting fluconazole treatment and the first day of fluconazole treatment (day 1), was used as the reference value to compare the mean blood pressure calculated every 3 days after day 1 until day 13. Most of the 26 patients included in the study had underlying hematologic malignancies. The average age of patients was 71.8 years. Twenty-one patients received amlodipine and five received nifedipine. The combination of DCCBs and fluconazole was associated with a significant decrease in blood pressure on day 11–13 (p < 0.01). The mean difference in overall systolic blood pressure was −15.8 mmHg (95% CI: −21.1 to −10.4). Therefore, the combination of fluconazole and DCCBs might potentiate the antihypertensive effect of DCCBs, and caution should be exercised when using them for lowering blood pressure.

Expression and Distribution of GPR55, a Receptor for Lysophosphatidylinositol, in Mouse Tissues and Cells Vol.5, No.2, p.16-20
Takashi Tanikawa , Saori Oka , Keisuke Nakajima , Yasuhiro Hayashi , Yoko Nemoto-Sasaki , Yoichiro Arata , Takayuki Sugiura , Atsushi Yamashita
Received: October 04, 2021
Accepted: February 15, 2022
Released: March 11, 2022
Abstract Full Text PDF[907K]

GPR55 is a G protein-coupled receptor that is proposed as a novel type of cannabinoid receptor. Lysophosphatidylinositol is an endogenous ligand for GPR55. The physiological roles of GPR55 have not yet been elucidated in detail. In the present study, the expression of Gpr55 mRNA was evaluated in various mouse tissues and organs using real-time RT-PCR. Gpr55 mRNA expression was highest in testis, the male reproductive system, among mouse tissues. Gpr55 mRNA expression was high in immune organs such as the spleen, lymph nodes, and thymus. Gpr55 mRNA was also detected in the small and large intestines. The expression of Gpr55 mRNA was relatively low in a mouse brain. The distribution of Gpr55 in mice is very similar to that in humans, however, the rank order was somewhat different. The sub-fractionation revealed that Gpr55 mRNA was expressed in both germinal cells and somatic cells in the testis. In the small intestine, Gpr55 was expressed in the duodenum, jejunum, and ileum. Gpr55 was highly expressed in B and T lymphocytes and dendritic cells in the mouse spleen.

Regular Article
Addition of Malonyl Groups Enhances Intestinal Absorption of Anthocyanins Derived from Edible Red Chrysanthemum (Dendranthema grandiflorum) in Rats Vol.5, No.1, p.9-15
Takashi Ichiyanagi , Yoshiki Kashiwada , Masayuki Nashimoto , Norihiko Terahara
Received: December 23, 2021
Accepted: February 01, 2022
Released: February 10, 2022
Abstract Full Text PDF[1017K]

Petals of red chrysanthemum (Dendranthema grandiflorum) are habitually eaten in Japan. In the present study, plasma concentration profiles of two major acylated anthocyanins derived from the petals of red chrysanthemum were evaluated in rats after oral administration of an anthocyanin-rich fraction obtained from red chrysanthemum. The structures of the two major anthocyanins in the petals of red chrysanthemum were determined to be cyanidin 3-O-β-D-(3″, 6″-di-O-malonyl)-glucopyranoside and cyanidin 3-O-β-D-(6″-mono-O-malonyl)- glucopyranoside. Both malonyl anthocyanins were quickly absorbed from the gastrointestinal tract and were detected in rat blood plasma in their original acylated forms 15 min after the oral administration of the anthocyanin fraction obtained from the petals of red chrysanthemum. The absorption amounts of anthocyanins evaluated from the area under the plasma concentration curves during 8 h normalized to the orally administered dose were in the following order: cyanidin 3-O-β-D-(3″, 6″-di-O-malonyl)-glucopyranoside > cyanidin 3-O-β-D-(6″-mono-O-malonyl)-glucopyranoside > cyanidin 3-O-β-D-glucopyranoside. The present results demonstrated that the additional malonylation of the glucopyranosyl moiety of position 3″ and 6″ of cyanidin 3-O-β-D-glucopyranoside enhanced the intestinal absorption of anthocyanins.

Selenium Toxicity Accelerated by Out-of-Control Response of Nrf2-xCT Pathway Vol.5, No.1, p.5-8
Koji Ueda , Yoshinori Okamoto , Akira Aoki , Hideto Jinno
Received: December 14, 2021
Accepted: January 07, 2022
Released: January 20, 2022
Abstract Full Text PDF[1M]

Selenium (Se) is an essential biological element and selenite is used to supplement malnutrition of Se and may also have anticancer activity. However, Se is also known as a delicate micronutrient with a narrow window of useful dose and the mechanisms for the sudden toxic effect remain unclear. Recently, we reported elsewhere that selenite was incorporated into cells via xCT, a cystine/glutamate antiporter. xCT is regulated by a stress responsive nuclear factor erythroid 2-related factor 2 (Nrf2). Therefore, we hypothesized and preliminary substantiated that the Nrf2-xCT pathway underlies the toxicity mechanism of Se. Expression of xCT mRNA was remarkably increased in MCF-7 cells after Se treatment, which may further increase Se uptake and oxidative stress. Pretreatment with xCT or Nrf2 inhibitors prevented morphological changes by releasing cells from uncontrolled feedback on Se uptake. Paradoxically, Se-induced oxidative toxicity is promoted by a runaway of stress response in Nrf2-xCT pathway. The results imply a novel mechanism by which Se accelerates its oxidative toxicity through feedback via Nrf2-xCT. This mechanism explain the elusive toxicological properties of inorganic Se, including strong cytotoxicity, high sensitivity in cancer cells, and narrowness of pharmacological dose range.

Silica Particles with Human Protein Corona Shows Sensitization Potential in the Human Cell Line Activation Test Vol.5, No.1, p.1-4
Shun-ichi Eto , Aoi Koshida , Hirofumi Tsujino , Kazuya Nagano , Kazuma Higashisaka , Yasuo Tsutsumi
Received: December 20, 2021
Accepted: December 24, 2021
Released: January 12, 2022
Abstract Full Text PDF[1M]

Nanoparticles are concerned to show adverse biological effects despite their unique functions. Their physicochemical properties and status are widely diverse; this makes safety analysis of nanoparticles complicated. Some reports showed that nanoparticles could disturb acquired immunity, while it is still unclear what is the inducer of that effects. Here, we tried to explore the relationship among nanoparticles' physicochemical properties and sensitizing potential by using the human cell line activation test based in vitro method; that uses in expression of CD86 and CD54 as an index of cellular activation. As a model of nanoparticles, we examined sensitization potential of silica particles with or without a human protein corona. Of the cells treated with silica particles (diameter: 50 or 300 nm) only, none of them showed activation. On the other hand, silica particles with human protein corona showed activation. Moreover, protein corona that forms around 50 nm silica nanoparticles have a higher sensitization potential than that of protein corona that forms around 300 nm silica particles. Our findings indicated that silica particles with human protein corona showed sensitization potential, and that sensitization potential could depend on the amount or kind of proteins within the corona.