Improvement of limit of detection in primer extension-based multiplexed mutation assay using capillary electrophoresis.

One of the challenges in liquid biopsy for early cancer detection is ascribed to the fact that mutation DNA often represents an extremely small ratio of less than 1% compared to wild-type genes in blood. However, in conventional fragment analysis with capillary electrophoresis (CE), the detectable allele frequency could be about 5%. In this work, we developed an original reagent-based fragment analysis with single base extension (SBE) reactions for cancer-associated mutation assay using a commercially available CE device, and investigated on a possibility of improvement of limit of detection (LOD) for genetic mutation. First, after adjustment of reagent conditions for the SBE reactions, the linear relationship between gene template concentration and fluorescence intensity was obtained from 1 to 100 fmol of target genes. Next, from the results of an experiment to detect mutation EGFR L858R at abundance ratios of mutant type to wild type (100-fmol template) of 0, 1, 5, and 10%, it was shown that the target gene can be detected with LOD of 0.33%. This high sensitivity was realized in part by separating fluorescently labeled substrates into an individual tube for an each-colored SBE reaction. Moreover, mutations EGFR L858R and KRAS G12V were simultaneously detected at sensitivities equivalent to LODs of 0.57 and 0.47%, respectively. These results indicate that < 1% of mutations in multiplex gene mutations can be simultaneously detected, and that possibility suggests that the developed method can be used in clinical practice for detecting cancers.

Simulation of the emergence of cell-like morphologies with evolutionary potential based on virtual molecular interactions.

This study explored the emergence of life using a simulation model approach. The "multiset chemical lattice model" allows the placement of virtual molecules of multiple types in each lattice cell in a two-dimensional space. This model was capable of describing a wide variety of states and interactions, such as the diffusion, chemical reaction, and polymerization of virtual molecules, in a limited number of lattice cell spaces. Moreover, this model was capable of describing a wide variety of states and interactions, even in the limited lattice cell space of 100 × 100 cells. In this study, I assumed 18 types of virtual molecules, i.e., 18 virtual numbers that do not correspond to real molecules with chemical reactions represented by transformation of the numbers that occur with a specified reaction rate probability. Furthermore, it considered the energy metabolism and energy resources in the environment, and was able to reproduce "evolution," in which a certain cell-like shape that adapted to the environment survived under conditions of decreasing amounts of energy resources in the environment. This enabled the simulation of the emergence of cell-like shapes with the four minimum cellular requirements, i.e., boundary, metabolism, replication, and evolution, based solely on the interaction of virtual molecules.

Anti-neutrophil Cytoplasmic Antibody-Associated Vasculitis With Periaortitis That Developed After mRNA COVID-19 Vaccination.

Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) has caused a global pandemic resulting in many deaths. As a result, vaccines to prevent the onset of coronavirus disease 2019 (COVID-19) have been developed and have demonstrated high efficacy in large-scale clinical trials. Adverse events that develop within a few days after vaccination are common, such as fever, malaise, body aches, and headaches, and have become widely known as transient reactions. However, as COVID-19 vaccines are administered worldwide, several studies have highlighted that long-term side effects associated with vaccines against SARS-CoV-2 may include serious adverse events. There has been an increase in reports of COVID-19 vaccinations being associated with the onset of autoimmune diseases, such as anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis. This is a report of ANCA-associated vasculitis with periaortitis following the second dose of COVID-19 mRNA vaccination, in which a 56-year-old man developed numbness and pain in his lower extremities three weeks after COVID-19 vaccination. Following the onset of sudden abdominal pain, a fluorodeoxyglucose-positron emission tomography scan revealed periaortic inflammation. Serum myeloperoxidase (MPO)-ANCA levels were significantly elevated, and renal biopsy revealed pauci-immune crescentic glomerulonephritis. Treatment with steroids and cyclophosphamide alleviated abdominal pain and numbness in the lower limbs, resulting in a decrease in MPO-ANCA titers. The side effects of COVID-19 vaccination are still unclear. This report has indicated that side effects associated with vaccines against COVID-19 may include ANCA-associated vasculitis. However, a causal relationship between COVID-19 vaccination and the development of ANCA-associated vasculitis has not yet been clearly demonstrated. COVID-19 vaccination will continue internationally, so it is necessary to accumulate similar case reports in the future.

Efficient and accurate KRAS genotyping using digital PCR combined with melting curve analysis for ctDNA from pancreatic cancer patients.

A highly sensitive and highly multiplexed quantification technique for nucleic acids is necessary to predict and evaluate cancer treatment by liquid biopsy. Digital PCR (dPCR) is a highly sensitive quantification technique, but conventional dPCR discriminates multiple targets by the color of the fluorescent dye of the probe, which limits multiplexing beyond the number of colors of fluorescent dyes. We previously developed a highly multiplexed dPCR technique combined with melting curve analysis. Herein, we improved the detection efficiency and accuracy of multiplexed dPCR with melting curve analysis to detect KRAS mutations in circulating tumor DNA (ctDNA) prepared from clinical samples. The mutation detection efficiency was increased from 25.9% of the input DNA to 45.2% by shortening the amplicon size. The limit of detection of mutation was improved from 0.41 to 0.06% by changing the mutation type determination algorithm for G12A, resulting in a limit of detection of less than 0.2% for all the target mutations. Then, ctDNA in plasma from pancreatic cancer patients was measured and genotyped. The measured mutation frequencies correlated well with those measured by conventional dPCR, which can measure only the total frequency of KRAS mutants. KRAS mutations were detected in 82.3% of patients with liver or lung metastasis, which was consistent with other reports. Accordingly, this study demonstrated the clinical utility of multiplex dPCR with melting curve analysis to detect and genotype ctDNA from plasma with sufficient sensitivity.

Emergence Simulation of Biological Cell-like Shapes Satisfying the Conditions of Life Using a Lattice-Type Multiset Chemical Model.

Although numerous reports using methods such as molecular dynamics, cellular automata, and artificial chemistry have clarified the process connecting non-life and life on protocell simulations, none of the models could simultaneously explain the emergence of cell shape, continuous self-replication, and replication control solely from molecular reactions and diffusion. Herein, we developed a model to generate all three conditions, except evolution ability, from hypothetical chains of chemical and molecular polymerization reactions. The present model considers a 2D lattice cell space, where virtual molecules are placed in each cell, and molecular reactions in each cell are based on a multiset rewriting rule, indicating stochastic transition of molecular species. The reaction paths of virtual molecules were implemented by replacing the rules of cellular automata that generate Turing patterns with molecular reactions. The emergence of a cell-like form with all three conditions except evolution ability was modeled and demonstrated using only molecular diffusion, reaction, and polymerization for modeling the chemical reactions of 15 types of molecules and 2 types of polymerized molecules. Furthermore, controlling self-replication is possible by changing the initial arrangement of a specific molecule. In summary, the present model is capable of investigating and refining existing hypotheses on the emergence of life.

Percutaneous Decannulation for Venoarterial Extracorporeal Membrane Oxygenation Using a Perclose ProGlide Closure Device and a Balloon Catheter Without On-Site Cardiac Surgical Backup.

Surgical decannulation for venoarterial extracorporeal membrane oxygenation (VA-ECMO) is recommended as a standard weaning strategy considering large-sized cannulas (14-22 French) are inserted in VA-ECMO. However, we should be aware of complications such as bleeding and infection when removing an arterial cannula, especially in facilities without on-site cardiac surgical backup. Percutaneous closure devices for femoral arterial access sites are currently approved for the decannulation of a 10-French or smaller sheath. We reported a case of successful weaning off from ECMO using a combination method of a balloon catheter and a Perclose ProGlide closure device. We successfully removed the arterial cannula using this technique for four ECMO-treated patients without vascular complications or blood transfusion. Percutaneous decannulation by this method could reduce the procedural time and adverse events and be safely performed even in facilities without on-site cardiac surgical backup.

Prognostic impact of malnutrition in elderly patients with acute myocardial infarction.

We aimed to investigate the prevalence and the prognostic impact of malnutrition using the geriatric nutritional risk index (GNRI) in elderly patients with acute myocardial infarction (AMI). We investigated 130 consecutive patients aged ≥ 75 years who were discharged following successful primary coronary intervention between March 2009 and December 2016. The mean age of the patients was 81.5 ± 5.0 years, and 55.4% of them were male. At discharge, the patients' nutritional statuses were evaluated using the GNRI, which were calculated as follows: 14.89 × serum albumin (g/dL) + 41.7 × body mass index/22. The patients were divided into two groups, a low GNRI group (GNRI < 92) with nutrition-related risk and a high GNRI group (GNRI ≥ 92) without nutrition-related risk. The mean GNRI score of all patients was 95.2 ± 11.0, and 46 patients (35.4%) had a low GNRI score. During the mean follow-up of 1030 ± 850 days, the all-cause mortality was significantly higher in the low GNRI group than in the high GNRI group (32.6% vs. 13.1%, p < 0.001), while the event rates of AMI recurrence and hospitalization due to heart failure did not differ significantly between the two groups. Multivariate Cox proportional analysis revealed that low GNRI was a modest but independent predictor of mortality (hazard ratio, 0.94; 95% confidence interval 0.90-0.98; p = 0.01). Among elderly patients with AMI, malnutrition assessed using the GNRI was often encountered and was an independent predictor of long-term mortality.

Use of oral hydromorphone in a patient with stage D heart failure.

Patients with advanced heart failure often experience dyspnea, fatigue, edema, and appetite loss. If these symptoms are refractory to treatment, palliative care via a team approach is necessary. We describe a patient with stage D heart failure whose dyspnea and overall condition improved with comprehensive medical treatments including conventional medications for heart failure, continuous infusions of catecholamine and diuretic, and oral hydromorphone. A 67-year-old man with a 12-year history of dilated cardiomyopathy was admitted to our hospital due to exacerbation of heart failure. Despite continuous infusion of catecholamine and diuretic, his dyspnea and liver and renal function continued to worsen. Oral hydromorphone was administered to relieve his refractory dyspnea, which also improved his conditions, continuous infusion of the catecholamine and diuretic could withdraw. Oral low-dose hydromorphone used in the present case might be a helpful agent for treating dyspnea in stage D heart failure patients with renal dysfunction.

A model of octopus epidermis pattern mimicry mechanisms using inverse operation of the Turing reaction model.

Many cephalopods such as octopi and squid can purposefully and rapidly change their skin color. Furthermore, it is widely known that some octopi have the ability to rapidly change the color and unevenness of their skin to mimic their surroundings. However, there has been little research published on the mechanisms by which an octopus recognizes its surrounding landscape and changes its skin pattern. We are unaware of any hypothetical model that explains this mimicry mechanism to date. In this study, the mechanism of octopus skin pattern change was assumed to be based on the Turing pattern model. Here, pattern formation using the Turing model was realized using an equivalent filter calculation model and a cellular automaton instead of directly solving the differential equations. It was shown that this model can create various patterns using two feature parameters. Furthermore, for visual recognition where two features are extracted from the Turing pattern image, a method that requires minimal calculation using the characteristics of the cellular Turing pattern model is proposed. These two calculations can be expressed in the same mathematical frame based on the cellular automaton model using a convolution filter. As a result, a model that is capable of extracting features from patterns and reconstructing those patterns rapidly can be created. This represents a basic model of the mimicry mechanism of octopi. Further, this study demonstrates the potential for creating a model with minimal learning calculation for application to machine learning.

The Multi-Domain Intervention Trial in Older Adults With Diabetes Mellitus for Prevention of Dementia in Japan: Study Protocol for a Multi-Center, Randomized, 18-Month Controlled Trial.

Background: The Japan-Multi-domain Intervention Trial for Prevention of Dementia in Older Adults with Diabetes (J-MIND-Diabetes) is an 18-month, multi-centered, open-labeled, randomized controlled trial designed to identify whether multi-domain intervention targeting modifiable risk factors for dementia could prevent the progression of cognitive decline among older adults with type 2 diabetes mellitus (T2DM). This manuscript describes the study protocol for the J-MIND-Diabetes trial. Materials and Methods: Subjects of this trial will comprise a total of 300 T2DM outpatients aged 70-85 years with mild cognitive impairment. Subjects will be centrally randomized into intervention and control groups at a 1:1 allocation ratio using the stratified permuted-block randomization methods. The intervention group will participate in multi-domain intervention programs aimed at: (1) management of metabolic and vascular risk factors; (2) physical exercise and self-monitoring of physical activity; (3) nutritional guidance; and (4) social participation. The control group will receive usual T2DM care and general instructions on dementia prevention. The primary and secondary outcomes will be assessed at baseline, at 6- and 18-month follow-up. The primary outcome is change from baseline at 18 months in a global composite score combining several neuropsychological domains, including global cognitive function, memory, attention, executive function, processing speed and language. Secondary outcomes include: (1) cognitive changes in neuropsychological tests; (2) changes in geriatrics assessments; (3) metabolic control and diabetic complications; (4) changes in blood and urinary markers. Discussion: This trial will be the first trial to demonstrate the effectiveness of multi-domain intervention in preventing cognitive decline in older adults with T2DM at increased risk of dementia in Japan. Trial Registration: UMIN000035911; Registered on the University Hospital Medical Information Network Clinical Trials Registry (UMIN-CTR) 18 February 2019. (https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000040908).

Pancreatic cancer secondary to intraductal papillary mucinous neoplasm with collision between gastric cancer and B-cell lymphoma: A case report.

BACKGROUND: Patients with intraductal papillary mucinous neoplasm (IPMN) have an increased risk of pancreatic and extrapancreatic malignancies. Lymphomas are rare extrapancreatic malignancies, and in situ collisions of early gastric cancer and diffuse large B-cell lymphoma (DLBCL) are even rarer. Here, we report the first case of pancreatic cancer comorbid with in situ collision of extrapancreatic malignancies (early gastric cancer and DLBCL) in a follow-up IPMN patient. Furthermore, we have made innovations in the treatment of such cases. CASE SUMMARY: An 81-year-old Japanese female diagnosed with IPMN developed elevated carbohydrate antigen (CA) 19-9 levels during follow-up. Because her CA19-9 levels continued to rise, endoscopic ultrasound (EUS) was performed and revealed a suspicious lesion at the pancreatic tail. However, lesions in the pancreas were not found by computed tomography, magnetic resonance imaging, or endoscopic retrograde cholangiopancreatography. To make an exact patho-logical diagnosis, EUS-guided fine needle aspiration was performed. To our supprise, early gastric cancer was found in preoperative gastroscopy. The gastric cancer was completely resected through endoscopic submucosal dissection before postoperative pathology identified early adenocarcinoma collided with DLBCL. Subsequent EUS-guided fine needle aspiration provided pathological support for the pancreatic cancer diagnosis, and then laparoscopic distal pancreatectomy and splenectomy were performed. CA19-9 levels returned to normal postoperatively. CONCLUSION: Endoscopic submucosal dissection is appropriate for submucosal lymphomas in patients intoleratant of chemotherapy. EUS can detect small IPMN-related pancreatic tumors.

Low-frequency noise induced by cation exchange fluctuation on the wall of silicon nitride nanopore.

Nanopore-based biosensors have attracted attention as highly sensitive microscopes for detecting single molecules in aqueous solutions. However, the ionic current noise through a nanopore degrades the measurement accuracy. In this study, the magnitude of the low-frequency noise in the ionic current through a silicon nitride nanopore was found to change depending on the metal ion species in the aqueous solution. The order of the low-frequency noise magnitudes of the alkali metal ionic current was consistent with the order of the adsorption affinities of the metal ions for the silanol surface of the nanopore (Li

Prognostic impact of malnutrition assessed using geriatric nutritional risk index in patients aged ⩾80 years with heart failure.

AIMS: Malnutrition in elderly patients is one of the important issues in an aging society. We aimed to investigate the prevalence and prognostic impact of malnutrition assessed using the geriatric nutritional risk index in very elderly patients hospitalized owing to heart failure. METHODS: We enrolled 213 consecutive patients aged ⩾80 years who were hospitalized with heart failure. The mean age was 87.2 ± 4.9 years, and 43.7% of them were male. The nutritional status on admission was evaluated using the geriatric nutritional risk index, which was calculated as follows: 14.89 × serum albumin (g/dL) + 41.7 × body mass index/22. The patients were divided into two groups, a low geriatric nutritional risk index group (<92) with malnutrition risk and a high geriatric nutritional risk index group (⩾92) without malnutrition risk. RESULTS: The mean geriatric nutritional risk index of all patients was 90.7 ± 10.6, and 108 patients (50.7%) had low geriatric nutritional risk index. During the 540-day follow-up, the all-cause mortality was significantly higher in the low geriatric nutritional risk index group than in the high geriatric nutritional risk index group (35.7% vs. 12.9%, p < 0.001). The Cox proportional-hazards regression analysis revealed that low geriatric nutritional risk index was an independent predictor of mortality (hazard ratio, 2.28; 95% confidence interval, 1.15-4.49; p = 0.02). CONCLUSIONS: Low geriatric nutritional risk index on admission was common and was associated with poor prognosis in patients aged ⩾80 years who were hospitalized owing to heart failure.

Possibility of Controlling Self-Organized Patterns with Totalistic Cellular Automata Consisting of Both Rules like Game of Life and Rules Producing Turing Patterns.

The basic rules of self-organization using a totalistic cellular automaton (CA) were investigated, for which the cell state was determined by summing the states of neighboring cells, like in Conway's Game of Life. This study used a short-range and long-range summation of the cell states around the focal cell. These resemble reaction-diffusion (RD) equations, in which self-organizing behavior emerges from interactions between an activating factor and an inhibiting factor. In addition, Game-of-Life-type rules, in which a cell cannot survive when adjoined by too many or too few living cells, were applied. Our model was able to mimic patterns characteristic of biological cells, including movement, growth, and reproduction. This result suggests the possibility of controlling self-organized patterns. Our model can also be applied to the control of engineering systems, such as multirobot swarms and self-assembling microrobots.

Anaphylactoid Purpura Associated with Streptococcal Cellulitis: A Case Report and Literature Review.

A 54-year-old Japanese man noticed painful swelling and redness of his left leg. He was admitted for treatment of cellulitis, which was accompanied with increased anti-streptolysin O and anti-streptokinase titers in his clinical course. After Piperacillin/Tazobactam administration, the skin lesion resolved. However, the patient then developed arthritis, palpable purpura, and intermittent abdominal pain, later found to be secondary to a severe duodenal ulcer. He was diagnosed with cellulitis-associated anaphylactoid purpura and was given prednisolone, which dramatically improved his symptoms. The anaphylactoid purpura was likely caused by Streptococcus-induced cellulitis, which was successfully treated with prednisolone. Association between these diseases is rare.

Side-gated ultrathin-channel nanopore FET sensors.

A side-gated, ultrathin-channel nanopore FET (SGNAFET) is proposed for fast and label-free DNA sequencing. The concept of the SGNAFET comprises the detection of changes in the channel current during DNA translocation through a nanopore and identifying the four types of nucleotides as a result of these changes. To achieve this goal, both p- and n-type SGNAFETs with a channel thicknesses of 2 or 4 nm were fabricated, and the stable transistor operation of both SGNAFETs in air, water, and a KCl buffer solution were confirmed. In addition, synchronized current changes were observed between the ionic current through the nanopore and the SGNAFET's drain current during DNA translocation through the nanopore.