4-Iodobenzonitrile as a fluorogenic derivatization reagent for chromatographic analysis of L-p-boronophenylalanine in whole blood samples using Suzuki coupling reaction.

BACKGROUND: L-p-Boronophehylalanine (BPA) is used in boron neutron capture therapy (BNCT), which is a novel selective cancer radiotherapy technique. It is important to measure BPA levels in human blood for effective radiotherapy; a prompt gamma-ray spectrometer, ICP-AES, and ICP-MS have been used for this purpose. However, these methods require sophisticated and expensive apparatuses as well as experienced analysts. Herein, we propose an HPLC-FL method for the determination of BPA after precolumn derivatization. A new fluorogenic reagent for aryl boronic acid derivatives, namely, 4-iodobenzonitrile, was employed for the fluorogenic derivatization of BPA based on the Suzuki coupling reaction. RESULTS: After the fluorogenic derivatization, a fluorescent cyanobiphenyl derivative is formed with maximum fluorescence at 335 nm after excitation at 290 nm. The developed method showed good linearity (r2=0.997) over the concentration range of 0.5-1000 nmol/L, and the detection limit (S/N = 3) was 0.26 nmol/L. The proposed method is more sensitive than previously reported methods for the determination of BPA, including the ICP-MS. Finally, the proposed method was successively applied to the measurement of BPA in human whole blood samples with a good recovery rate (≥95.7 %) using only 10 µL of blood sample. The proposed method offers a simple and efficient solution for monitoring BPA levels in BNCT-treated patients. SIGNIFICANCE: 4-Iodobenzonitrile was investigated as a new fluorogenic reagent for BPA based on Suzuki coupling. A new HPLC-FL method for BPA in whole blood samples with ultrasensitivity was developed. The developed method is superior in sensitivity to all previously reported methods for BPA. The method requires only a very small sample volume, making it suitable for micro-blood analysis of BPA via fingerstick sampling.

Radiation proctitis after iodine-125 low-dose-rate prostate brachytherapy utilizing SpaceOAR hydrogel.

OBJECTIVE: We retrospectively evaluated the efficacy of combining the SpaceOAR (SOAR) hydrogel with prostate brachytherapy, using colonoscopy findings to assess for radiation proctitis. METHODS: Among 731 patients undergoing iodine-125 low-dose-rate prostate brachytherapy (LDR-BT), SOAR was utilized in 394 patients (53.9%). Colonoscopy was performed for 97 patients (13.3%) to assess the presence, location, condition, and treatment of radiation proctitis. We also investigated treatment factors associated with the occurrence of radiation proctitis. RESULTS: Radiation proctitis was observed in 57 patients (7.8%) and 17 (2.3%) were treated with argon plasma coagulation (APC). The incidence of radiation proctitis was 12.2% in the non-SOAR and 4.1% in the SOAR group (p < 0.001). In the non-SOAR group, the incidence of radiation proctitis was 6.6% for LDR-BT monotherapy and increased to 22.0% when combined with external beam radiation therapy (EBRT) (p = 0.001). However, in the SOAR group, these rates significantly decreased to 3.3% and 5.7% for monotherapy and combination therapy, respectively (p = 0.035, p < 0.001). With SOAR, inflammation was observed directly above the DL in most patients (87.5%), and only one patient (6.3%) required APC. The absence of SOAR (p < 0.001, HR = 0.29) and the concurrent use of EBRT (p = 0.018, HR = 2.87) were identified as significant risk factors for the occurrence of radiation proctitis. CONCLUSION: The use of SOAR significantly reduced the incidence of radiation proctitis in patients undergoing LDR-BT monotherapy and combined EBRT. Inflammation primarily occurred directly above the DL; further examination is necessary to clarify its cause.

Transportation of Medical Staff by Helicopter to Manage a Concealed Delivery at Home.

An unmarried pregnant woman felt lower abdominal pain. She rested in bed in her room on the second floor in her home. The next day she performed a delivery by herself. After the neonate cried, her parents noticed the birth and called an ambulance. After receiving the first call, the fire department decided to request the dispatch of a physician-staffed helicopter emergency medical service in Eastern Shizuoka, in addition to dispatching an ambulance. After receiving the request, the helicopter emergency medical service transported 1 neonatal intensive care unit physician along with the original medical staff members of the fire department. Then, the 3 medical staff members were transported to the home by another ambulance. When emergency medical technicians climbed up a steep narrow ladder to enter the room, both the mother and female neonate were connected by the umbilical cord. Their vital signs were stable. At 30 minutes after delivery, the medical staff reached the mother and neonate and cut the umbilical cord. The mother and neonate were evacuated separately from the room but transported in the same ambulance. The ambulance transported them with the medical staff members to our hospital directly. Their postadmission courses were uneventful, and they were discharged. This is the first case report to send medical staff members to the patient's home by helicopter and ambulance to provide medical intervention for the neonate and her mother. Further prospective studies are needed in the future to determine whether this action could lead to favorable outcomes in both neonates and maternal bodies.

[Bleeding due to acquired factor V inhibitor successfully treated with platelet transfusion during treatment for recurrent prostate squamous cell carcinoma].

A 78-year-old man with prostate squamous cell carcinoma recurrence in his pelvis was admitted to our hospital. Rectal obstruction led to creation of an artificial anus on the transverse colon. Then, docetaxel and radiation therapies were started. A week later, severe hematuria and melena occurred. Activated partial thromboplastin time (APTT) and prothrombin time (PT) were extremely prolonged. Cross-mixing test for APTT and PT revealed an inhibitor pattern, which was diagnosed as acquired factor V inhibitor. Fresh frozen plasma and vitamin K infusions were ineffective, but platelet transfusion successfully stopped the bleeding. Platelet factor V derived from megakaryocytes may affect local hemostasis. The patient received prednisolone (PSL), and the inhibitor disappeared on day 70 and was in remission. PSL could be stopped on day 100. Later, we demonstrated APTT and PT shortening of factor V deficient plasma by the supernatant of activated platelets with collagen.

Minor ST-T changes on electrocardiograms are associated with reduced constructive myocardial work in hypertensive patients with a preserved ejection fraction.

Minor ST-T changes on electrocardiograms are observed in patients with hypertensive heart disease with a preserved left ventricular ejection fraction (LVEF); however, the relationship between minor ST-T changes and global myocardial work on echocardiograms remains unclear. We evaluated the global longitudinal strain (GLS), global work index (GWI), global constructive work (GCW), global wasted work (GWW), and global work efficiency (GWE) in 186 hypertensive patients with preserved LVEF (>50%) using an offline analysis system (View Pal, GE). Minor ST-T changes as well as major ST-T changes (depression in ST in lead V5 > 1 mV) were also evaluated by electrocardiography. The mean age was 79.4 ± 8.2 years (men 37.6%). Patients with minor and major ST-changes had a smaller absolute GLS (-20.3 ± 3.1, -18.2 ± 3.7, and -15.2 ± 5.0%, P < 0.001) and a smaller constructive workload [GWI (2148 ± 486, 1938 ± 462, and 1685 ± 701 mmHg%, P = 0.002) and GCW (2419 ± 510, 2185 ± 466, and 1865 ± 702 mmHg%, P < 0.001)] than those with no ST-T changes. Additionally, patients with minor and major ST-T changes had increased wasted myocardial work [GWW (87 ± 61, 105 ± 56, and 127 ± 75 mmHg%, P = 0.045)], which resulted in reductions in the myocardial work efficiency [GWE (95 ± 4, 94 ± 3, and 89 ± 12%, P < 0.001)]. Even after adjustments for confounding factors, including EF, diastolic function parameters, LV hypertrophy, history of heart failure and atrial fibrillation, patients with minor ST-T changes had significantly smaller LV global constructive work in comparison to those without [GWI (2260 ± 33 vs. 2025 ± 76 mmHg%, P = 0.007) and GCW (2501 ± 45 vs. 2210 ± 105 mmHg%, P = 0.003)]. In conclusion, in hypertensive patients with preserved LVEF, minor ST-T changes on electrocardiograms were associated with reduced constructive work in the LV.

Detection of low-mineral- and high-salt responsible caecal indigenous bacteria in ICR mice.

Both deficiency and overdose of minerals and salts negatively affect health. Changes in the dietary composition have immediate effects on the gut microbiota. This study was performed to clarify the presence of indigenous gut bacteria responsible for minerals and/or salts (MS-RIB). ICR mice were fed a diet containing 3.5% (w/w) mineral mix (control), 1% mineral mix (LM), or 3.5% mineral mix and 4% NaCl (HS) for 14 days. The caecal microbiota was examined using 16S rRNA gene (V4) amplicon sequencing. Consumption of drinking water was 2.5-fold higher in the HS group than in the other groups. Body weight gain was 55% lower in the HS group than in the other groups. At the family level, the relative abundance of Eryspelotrichaceae and Clostridiaceae was lower in the HS group than in the other groups. In contrast, the abundance of Bacteroidaceae was higher in the HS group. At the operational taxonomic unit level, Desulfovibrionaceaer-, Turicibacter sanguinis-, belonging to Eryspelotrichaceae, and Clostridium disporicum-like bacteria were dominant in the control group. Among these bacteria, T. sanguinis- and C. disporicum-like bacteria were markedly suppressed by HS. In the LM group, Bacteroides acidifaciens-like bacteria were suppressed. Suppression of C. disporicum and Turicibacter following consumption of the HS diet was the most notable effect, contrasting the results of previous studies.

Typical gut indigenous bacteria in ICR mice fed a soy protein-based normal or low-protein diet.

For patients with inflammatory bowel disease, cow's milk allergy, and lactose intolerance, soymilk is a potential alternative to cow's milk. In this study, we aimed to identify the effects of a soy protein-based low-protein diet on the body and organ weights and the gut microbiome of six-week-old mice fed a diet containing 20% (SP) or 5% (LP) soy protein for 14 days via 16S rRNA (V4) amplicon sequencing. Body weight gain (growth) and liver, spleen, and fat tissue weight were significantly suppressed by the LP diet. Operational taxonomic unit numbers and α-diversity were lower in the LP group than in the SP group. A principal coordinate analysis revealed differences in the gut microbiome compositions of SP and LP mice. The abundances of caecal Roseburia sp., Alistipes sp., and bacteria from the family Muribaculaceae were lower in the LP group than in the SP group. In contrast, the abundance of Desulfovibrionaceae, which is positively correlated with inflammation, was higher in the LP group than in the SP group. These results differed from the effects of a milk casein-based low-protein diet (reported previously). Based on these findings, we conclude that the undesirable effects of a low-protein diet and/or protein deficiency are related to changes in the gut microbiome composition and may differ depending on the kind of proteins used.

Detachment of a prosthetic valve due to infective endocarditis caused by Streptococcus pneumoniae.

The incidence of infective endocarditis (IE) due to S pneumoniae has decreased, thanks to antibiotics. However, when it does occur, it can be lethal. The present case provides a reminder of the potential lethality of this postoperative infection.

Detection and isolation of the typical gut indigenous bacteria from ddY mice fed a casein-beef tallow-based or egg yolk-based diet.

The effects of whole egg on the cecal microbiome of ddY mice has been reported. To investigate the existence of susceptible indigenous bacteria (SIB) to egg yolks (EY), mice were fed a diet containing either 20% (w/w) milk casein and 17% beef tallow (CT) or 12% milk casein and 27% EY for 14 days, and then, the cecal microbiome was analyzed by 16S rRNA (V4) amplicon sequencing. To isolate the typical species in each diet group, culture-dependent viable bacterial counts were determined on Blood Liver (BL) and Gifu Anaerobic Medium (GAM) agar plates. The amplicon sequencing analysis revealed typical CT-SIB, such as Lachnospiraceae-like bacteria, and EY-SIB, such as Allobaculum-, Lactobacillus murinus-, and Bacteroides vulgatus-like bacteria. Two of the detected SIB species, L. murinus- and B. vulgatus-like bacteria, were successfully isolated from the BL and GAM agar plates and defined using a 16S rDNA BLAST search. PRACTICAL APPLICATIONS: The SIB defined in the CT and EY groups might have some effects on the nutritional and functional chemical compounds in the milk casein, beef tallow, and/or EY. Analysis of its functional properties of the isolates might develop the new and unique probiotic strains.

Detection and isolation of low molecular weight alginate- and laminaran-susceptible gut indigenous bacteria from ICR mice.

Alginate and laminaran are the main water-soluble polysaccharides in edible brown algae such as arame Eisenia bicyclis. To determine the alginate- and/or laminaran-susceptible indigenous bacteria (SIB) in the gut, the caecal microbiomes of ICR mice fed a diet containing 2% low molecular weight (LMW ≒50 kDa) alginate or laminaran were analysed by 16S rRNA gene (V4) amplicon sequencing. At the phylum level abundances, compared to those in mice fed a no-fibre diet, Firmicutes was lower and Bacteroidetes was higher in both LMW alginate- or laminaran-fed mouse groups. At the operational taxonomic unit level, Bacteroides acidifaciens- and Bacteroides intestinalis-like bacteria were considered alginate- and laminaran-SIB, respectively. B. acidifaciens PS-4 isolated from the ICR mice fermented LMW alginate and laminaran and mainly produced succinate. B. intestinalis ALB-11 also isolated from these mice fermented laminaran and mainly produced lactate. These SIB might exert interactive effects with edible brown algal consumption and affect host health.

Female cotton rats (Sigmodon hispidus) develop chronic anemia with renal inflammation and cystic changes.

The cotton rat (Sigmodon hispidus) is a laboratory rodent that has been used for studies on human infectious diseases. In the present study, we observed that female cotton rats, not the male cotton rats, developed chronic anemia characterized by reduced red blood cell, hemoglobin, and hematocrit levels from 5 to 9 months of age without any changes in the mean corpuscular hemoglobin and volume levels. In peripheral blood, the reticulocyte count did not increase in response to anemia in female cotton rats, and no extramedullary hematopoiesis was observed in the liver or spleen. Further, the serum levels of urea nitrogen and creatinine increased from 5 to 9 months of age in female cotton rats compared to male cotton rats, and these increases became more prominent from 10 months of age onward, indicating chronic kidney disease. Histopathologically, female cotton rats manifested tubulointerstitial lesions characterized by the infiltration of mononuclear cells, including plasma cells and CD3(+) T-cells, as well as the dilation of calbindin-D28k(+) distal tubules from 5 to 9 months of age. The severity of these lesions progressed from 10 months of age onward, and renal fibrotic features and numerous tubular cysts appeared without any obvious glomerular lesions. A significant decrease in the erythropoietin protein levels was observed in the kidney of aged female cotton rats, and significant correlations were detected between anemia and tubulointerstitial damage. These results suggest that aged female cotton rats chronically develop renal anemia, and this rodent may serve as a novel model to elucidate its pathogenesis.

Usefulness of an anesthetic mixture of medetomidine, midazolam, and butorphanol in cotton rats (Sigmodn hispidus).

Tne cotton rat (Sigmodon hispidus) is a laboratory rodent used for studying human infectious diseases. However, a lack of suitable anesthetic agents inconveniences the use of cotton rats in surgical manipulation. This study demonstrated that subcutaneous injection of the mixture of medetomidine, midazolam, and butorphanol (0.15, 2.0, and 2.5 mg/kg, respectively), which is a suitable anesthetic agents for mice and rats, produced an anesthetic duration of more than 50 min in cotton rats. We also demonstrated that 0.15 mg/kg of atipamezole, an antagonist of medetomidine, produced a quick recovery from anesthesia in cotton rats. This indicated that the anesthetic mixture of medetomidine, midazolam, and butorphanol, functioned as a useful and effective anesthetic for short-term surgery in cotton rats.

Induced pluripotent stem cells from CINCA syndrome patients as a model for dissecting somatic mosaicism and drug discovery.

Chronic infantile neurologic cutaneous and articular (CINCA) syndrome is an IL-1-driven autoinflammatory disorder caused mainly by NLRP3 mutations. The pathogenesis of CINCA syndrome patients who carry NLRP3 mutations as somatic mosaicism has not been precisely described because of the difficulty in separating individual cells based on the presence or absence of the mutation. Here we report the generation of NLRP3-mutant and nonmutant-induced pluripotent stem cell (iPSC) lines from 2 CINCA syndrome patients with somatic mosaicism, and describe their differentiation into macrophages (iPS-MPs). We found that mutant cells are predominantly responsible for the pathogenesis in these mosaic patients because only mutant iPS-MPs showed the disease relevant phenotype of abnormal IL-1ß secretion. We also confirmed that the existing anti-inflammatory compounds inhibited the abnormal IL-1ß secretion, indicating that mutant iPS-MPs are applicable for drug screening for CINCA syndrome and other NLRP3-related inflammatory conditions. Our results illustrate that patient-derived iPSCs are useful for dissecting somatic mosaicism and that NLRP3-mutant iPSCs can provide a valuable platform for drug discovery for multiple NLRP3-related disorders.

ONO-8130, a selective prostanoid EP1 receptor antagonist, relieves bladder pain in mice with cyclophosphamide-induced cystitis.

Given the previous evidence for involvement of prostanoid EP1 receptors in facilitation of the bladder afferent nerve activity and micturition reflex, the present study investigated the effect of ONO-8130, a selective EP1 receptor antagonist, on cystitis-related bladder pain in mice. Cystitis in mice was produced by intraperitoneal administration of cyclophosphamide at 300mg/kg. Bladder pain-like nociceptive behavior and referred hyperalgesia were assessed in conscious mice. Phosphorylation of extracellular signal-regulated kinase (ERK) in the L6 spinal cord was determined by immunohistochemistry in anesthetized mice. Cyclophosphamide treatment caused bladder pain-like nociceptive behavior and referred hyperalgesia accompanying cystitis symptoms, including increased bladder weight and vascular permeability and upregulation of cyclooxygenase-2 in the bladder tissue. Oral preadministration of ONO-8130 at 0.3-30 mg/kg strongly prevented both the bladder pain-like behavior and referred hyperalgesia in a dose-dependent manner, but had slight effect on the increased bladder weight and vascular permeability. Oral ONO-8130 at 30 mg/kg also reversed the established cystitis-related bladder pain. Intravesical administration of prostaglandin E2 caused prompt phosphorylation of ERK in the L6 spinal cord, an effect blocked by ONO-8130. Our findings strongly suggest that the prostaglandin E2/EP1 system participates in processing of cystitis-related bladder pain, and that EP1 antagonists including ONO-8130 are useful for treatment of bladder pain, particularly in interstitial cystitis. Prostaglandin E2 contributes to cystitis-related bladder pain via EP1 receptors in mice, indicating possible therapeutic usefulness of selective EP1 antagonists.