Understanding Flexdispersion: Structure-Function Relationship Studies of Organic Amphiphilic Ligands.

Since inorganic nanoparticles have unique properties that differ from bulk materials, their material applications have attracted attention in various fields. In order to utilize inorganic nanoparticles for functional materials, they must be dispersed without agglomeration. Therefore, the surfaces of inorganic nanoparticles are typically modified with organic ligands to improve their dispersibility. Nevertheless, the relationship between the tail group structure in organic ligands and the dispersibility of inorganic nanoparticles in organic solvents remains poorly understood. We previously developed amphiphilic ligands that consist of ethylene glycol chains and alkyl chains to disperse inorganic nanoparticles in a variety of organic solvents. However, the structural requirements for amphiphilic ligands to "flexibly" disperse nanoparticles in less polar to polar solvents are still unclear. Here, we designed and synthesized several phosphonic acid ligands for structure‒function relationship studies of flexdispersion. Dynamic light scattering analysis and visible light transmittance measurements revealed that the ratio of alkyl/ethylene glycol chains in organic ligands alone does not determine the dispersibility of the nanoparticles in organic solvents, but the arrangement of the individual chains also has an effect. From a practical application standpoint, it is preferable to design ligands with ethylene glycol chains on the outside relative to the particle surface.

Mtc6/Ehg2 is a novel endoplasmic reticulum-resident glycoprotein essential for high-pressure tolerance.

Hydrostatic pressure is a common mechanical stressor that modulates metabolism and reduces cell viability. Eukaryotic cells have genetic programs to cope with hydrostatic pressure stress and maintain intracellular homeostasis. However, the mechanism underlying hydrostatic pressure tolerance remains largely unknown. We have recently demonstrated that Maintenance of telomere capping protein 6 (Mtc6) plays a protective role in the survival of the budding yeast Saccharomyces cerevisiae under hydrostatic pressure stress by supporting the integrity of nutrient permeases. The current study demonstrate that Mtc6 acts as an endoplasmic reticulum (ER) membrane protein. Mtc6 comprises two transmembrane domains, a C-terminal cytoplasmic domain, and a luminal region with 12 Asn (N)-linked glycans attached to it. Serial mutational analyses showed that the cytoplasmic C-terminal amino acid residues GVPS are essential for Mtc6 activity. Multiple N-linked glycans in the luminal region are involved in the structural conformation of Mtc6. Moreover, deletion of MTC6 led to increased degradation of the leucine permease Bap2 under hydrostatic pressure, suggesting that Mtc6 facilitates proper folding of nutrient permeases in the ER under the stress condition. We propose a novel model of molecular function in which the glycosylated luminal domain and cytoplasmic GVPS sequences of Mtc6 cooperatively support the nutrient permease activity.

Genetically stable kill-switch using "demon and angel" expression construct of essential genes.

Genetic instability of synthetic genetic devices is a key obstacle for practical use. This problem is particularly critical in kill-switches for conditional host killing. Here, we propose a genetically stable kill-switch based on a "demon and angel" expression construct of a toxic essential gene. The kill-switch conditionally overexpresses the toxic essential gene. Additionally, the identical essential gene is deleted in the genome. The essential gene is expressed at a low level to maintain host survival in the OFF state and kills the host by the overexpression in the ON state. The single expression construct is responsible for both killing the hosts and maintaining viability, reducing the emergence of loss-of-function mutants. We constructed the kill-switch using the toxic essential gene encoding tyrosyl-tRNA synthetase, tyrS, in Escherichia coli. The bacteria harboring the kill-switch were conditionally suicidal over 300 generations. Toxic overexpression of essential genes has also been found in other organisms, suggesting that the "demon and angel" kill switch is scalable to various organisms.

A novel vaccine strategy using quick and easy conversion of bacterial pathogens to unnatural amino acid-auxotrophic suicide derivatives.

We propose a novel strategy for quick and easy preparation of suicide live vaccine candidates against bacterial pathogens. This method requires only the transformation of one or more plasmids carrying genes encoding for two types of biological devices, an unnatural amino acid (uAA) incorporation system and toxin-antitoxin systems in which translation of the antitoxins requires the uAA incorporation. Escherichia coli BL21-AI laboratory strains carrying the plasmids were viable in the presence of the uAA, whereas the free toxins killed these strains after the removal of the uAA. The survival time after uAA removal could be controlled by the choice of the uAA incorporation system and toxin-antitoxin systems. Multilayered toxin-antitoxin systems suppressed escape frequency to less than 1 escape per 109 generations in the best case. This conditional suicide system also worked in Salmonella enterica and E. coli clinical isolates. The S. enterica vaccine strains were attenuated with a >105 fold lethal dose. Serum IgG response and protection against the parental pathogenic strain were confirmed. In addition, the live E. coli vaccine strain was significantly more immunogenic and provided greater protection than a formalin-inactivated vaccine. The live E. coli vaccine was not detected after inoculation, presumably because the uAA is not present in the host animals or the natural environment. These results suggest that this strategy provides a novel way to rapidly produce safe and highly immunogenic live bacterial vaccine candidates. IMPORTANCE: Live vaccines are the oldest vaccines with a history of more than 200 years. Due to their strong immunogenicity, live vaccines are still an important category of vaccines today. However, the development of live vaccines has been challenging due to the difficulties in achieving a balance between safety and immunogenicity. In recent decades, the frequent emergence of various new and old pathogens at risk of causing pandemics has highlighted the need for rapid vaccine development processes. We have pioneered the use of uAAs to control gene expression and to conditionally kill host bacteria as a biological containment system. This report proposes a quick and easy conversion of bacterial pathogens into live vaccine candidates using this containment system. The balance between safety and immunogenicity can be modulated by the selection of the genetic devices used. Moreover, the uAA-auxotrophy can prevent the vaccine from infecting other individuals or establishing the environment.

Weakly nonlinear analysis on synchronization and oscillation quenching of coupled mechanical oscillators.

Various oscillatory phenomena occur in the world. Because some are associated with abnormal states (e.g. epilepsy), it is important to establish ways to terminate oscillations by external stimuli. However, despite the prior development of techniques for stabilizing unstable oscillations, relatively few studies address the transition from oscillatory to resting state in nonlinear dynamics. This study mainly analyzes the oscillation-quenching of metronomes on a platform as an example of such transitions. To facilitate the analysis, we describe the impulsive force (escapement mechanism) of a metronome by a fifth-order polynomial. By performing both averaging approximation and numerical simulation, we obtain a phase diagram for synchronization and oscillation quenching. We find that quenching occurs when the feedback to the oscillator increases, which will help explore the general principle regarding the state transition from oscillatory to resting state. We also numerically investigate the bifurcation of out-of-phase synchronization and beat-like solution. Despite the simplicity, our model successfully reproduces essential phenomena in interacting mechanical clocks, such as the bistability of in-phase and anti-phase synchrony and oscillation quenching occurring for a large mass ratio between the oscillator and the platform. We believe that our simple model will contribute to future analyses of other dynamics of mechanical clocks.

Effect of False Lumen Occlusion Treatment With AFX VELATM, Candy-Plug Technique for Chronic Aortic Dissection.

INTRODUCTION: We sought to examine midterm results and remodeling effect of false-lumen occlusion treatment using AFX VELA in case of chronic dissection repair. MATERIAL AND METHODS: From June 2019 to May 2022, we performed false lumen occlusion treatment using a modified Candy-Plug technique with AFX VELA on 8 chronic aortic dissection patients with a patent false lumen. We collected operative data, short-term clinical outcomes, mid-term clinical outcomes and imaging test results. We conducted follow-up examinations at postoperative, 6-month and 1-, 2- and 3-year intervals, including contrast-enhanced computed tomography to evaluate the diameter, false lumen thrombosis and any events. RESULTS: The average time from the symptom onset to the thoracic endovascular repair was 81.5 (35-155) months. The aorta showed aneurysmal dilation with an average maximum short-axis diameter of 58.9 (41-91) mm. Two cases needed emergency surgery due to rupture and impending rupture. There were no postoperative deaths. Complete thrombosis within the false lumen was achieved in 6 cases (75%), but 2 cases had incomplete thrombosis, requiring additional treatment. The mean maximum diameter showed a significant decrease at 6 months, 1 year and 2 years postoperatively compared to preoperative measurements (P < .05). CONCLUSION: We showed the results of false lumen occlusion treatment using the AFX VELA cuff. We observed favorable clinical outcomes and remodeling effects. While the long-term durability and efficacy of this technique in aortic remodeling will need to be monitored with further observation, the use of this cuff is considered a reliable approach to false lumen occlusion treatment.

Impact of Accessory Renal Artery Embolization on Renal Deterioration Under Endovascular Aortic Repair.

OBJECTIVE: The optimal management strategy for patients with accessory renal arteries undergoing endovascular aortic repair is unclear. This study aimed to investigate the impact of accessory renal artery (aRA) embolization on postoperative renal deterioration and to identify the predictors of postoperative renal deterioration in patients who underwent endovascular aortic repair (EVAR). METHODS: A retrospective single-centre observational study was conducted at our hospital. Of 331 consecutive patients who underwent endovascular aortic repair between April 2011 and February 2021, 29 patients with an aRA were included in this study. Spearman's rank correlation coefficients of decrease in estimated glomerular filtration rate (eGFR), renal volume reduction rate, infarcted renal volume, and quantity of contrast use for postoperative renal deterioration were analyzed. The correlation coefficients of the correlations between infarcted renal volume, renal volume reduction rate, and decrease in eGFR and the rate of aRA diameter were also analyzed. Multivariable nominal logistic regression analyses were conducted to evaluate the odds of postoperative renal deterioration. RESULTS: The renal volume reduction rate and infarcted renal volume had a significant positive correlation with the decrease in eGFR. Body surface area and preoperative renal volume were significantly but negatively correlated with the decrease in eGFR. The infarcted renal volume, renal volume reduction rate, and decrease in eGFR were significantly and positively correlated with the aRA diameter. The odds ratio for decreased eGFR rate in preoperative renal volume was .96 (95% CI 0.930‒.996, P = .009). CONCLUSIONS: EVAR with aRA embolization impacts postoperative renal deterioration in patients with preoperative low renal volume, and the diameter of the embolized aRA might be a predictor of postoperative renal deterioration.

Comparison of clinical utility of deep learning-based systems for small-bowel capsule endoscopy reading.

BACKGROUND AND AIM: Convolutional neural network (CNN) systems that automatically detect abnormalities from small-bowel capsule endoscopy (SBCE) images are still experimental, and no studies have directly compared the clinical usefulness of different systems. We compared endoscopist readings using an existing and a novel CNN system in a real-world SBCE setting. METHODS: Thirty-six complete SBCE videos, including 43 abnormal lesions (18 mucosal breaks, 8 angioectasia, and 17 protruding lesions), were retrospectively prepared. Three reading processes were compared: (A) endoscopist readings without CNN screening, (B) endoscopist readings after an existing CNN screening, and (C) endoscopist readings after a novel CNN screening. RESULTS: The mean number of small-bowel images was 14 747 per patient. Among these images, existing and novel CNN systems automatically captured 24.3% and 9.4% of the images, respectively. In this process, both systems extracted all 43 abnormal lesions. Next, we focused on the clinical usefulness. The detection rates of abnormalities by trainee endoscopists were not significantly different across the three processes: A, 77%; B, 67%; and C, 79%. The mean reading time of the trainees was the shortest during process C (10.1 min per patient), followed by processes B (23.1 min per patient) and A (33.6 min per patient). The mean psychological stress score while reading videos (scale, 1-5) was the lowest in process C (1.8) but was not significantly different between processes B (2.8) and A (3.2). CONCLUSIONS: Our novel CNN system significantly reduced endoscopist reading time and psychological stress while maintaining the detectability of abnormalities. CNN performance directly affects clinical utility and should be carefully assessed.

Characterization of tryptophan oxidation affecting D1 degradation by FtsH in the photosystem II quality control of chloroplasts.

Photosynthesis is one of the most important reactions for sustaining our environment. Photosystem II (PSII) is the initial site of photosynthetic electron transfer by water oxidation. Light in excess, however, causes the simultaneous production of reactive oxygen species (ROS), leading to photo-oxidative damage in PSII. To maintain photosynthetic activity, the PSII reaction center protein D1, which is the primary target of unavoidable photo-oxidative damage, is efficiently degraded by FtsH protease. In PSII subunits, photo-oxidative modifications of several amino acids such as Trp have been indeed documented, whereas the linkage between such modifications and D1 degradation remains elusive. Here, we show that an oxidative post-translational modification of Trp residue at the N-terminal tail of D1 is correlated with D1 degradation by FtsH during high-light stress. We revealed that Arabidopsis mutant lacking FtsH2 had increased levels of oxidative Trp residues in D1, among which an N-terminal Trp-14 was distinctively localized in the stromal side. Further characterization of Trp-14 using chloroplast transformation in Chlamydomonas indicated that substitution of D1 Trp-14 to Phe, mimicking Trp oxidation enhanced FtsH-mediated D1 degradation under high light, although the substitution did not affect protein stability and PSII activity. Molecular dynamics simulation of PSII implies that both Trp-14 oxidation and Phe substitution cause fluctuation of D1 N-terminal tail. Furthermore, Trp-14 to Phe modification appeared to have an additive effect in the interaction between FtsH and PSII core in vivo. Together, our results suggest that the Trp oxidation at its N-terminus of D1 may be one of the key oxidations in the PSII repair, leading to processive degradation by FtsH.

A specific plasma amino acid profile in the Insulin2Q104del Kuma mice at the diabetic state and reversal from hyperglycemia.

The metabolites in the plasma serve as potential biomarkers of disease. We previously established an early-onset diabetes mouse model, Ins2+/Q104del Kuma mice, under a severe immune-deficient (Rag-2/Jak3 double-deficient in BALB/c) background. Here, we revealed the differences in plasma amino acid profiles between Kuma and the wild-type mice. We observed an early reduction in glucogenic and ketogenic amino acids, a late increase in branched-chain amino acids (BCAAs) and succinyl CoA-related amino acids, and a trend of increasing ketogenic amino acids in Kuma mice than in the wild-type mice. Kuma mice exhibited hyperglucagonemia at high blood glucose, leading to perturbations in plasma amino acid profiles. The reversal of blood glucose by islet transplantation normalized the increases of the BCAAs and several aspects of the altered metabolic profiles in Kuma mice. Our results indicate that the Kuma mice are a unique animal model to study the link between plasma amino acid profile and the progression of diabetes for monitoring the therapeutic effects.

Synchronization and stability analysis of an exponentially diverging solution in a mathematical model of asymmetrically interacting agents.

This study deals with an existing mathematical model of asymmetrically interacting agents. We analyze the following two previously unfocused features of the model: (i) synchronization of growth rates and (ii) initial value dependence of damped oscillation. By applying the techniques of variable transformation and timescale separation, we perform the stability analysis of a diverging solution. We find that (i) all growth rates synchronize to the same value that is as small as the smallest growth rate and (ii) oscillatory dynamics appear if the initial value of the slowest-growing agent is sufficiently small. Furthermore, our analytical method proposes a way to apply stability analysis to an exponentially diverging solution, which we believe is also a contribution of this study. Although the employed model is originally proposed as a model of infectious disease, we do not discuss its biological relevance but merely focus on the technical aspects.

Activation of cAMP (EPAC2) signaling pathway promotes hepatocyte attachment.

Primary Human Hepatocyte (PHH) remains undefeated as the gold standard in hepatic studies. Despite its valuable properties, partial attachment loss due to the extraction process and cryopreservation remained the main hurdle in its application. We hypothesized that we could overcome the loss of PHH cell attachment through thawing protocol adjustment and medium composition. We reported a novel use of a medium designed for iPSC-derived hepatocytes, increasing PHH attachment on the collagen matrix. Delving further into the medium composition, we discovered that removing BSA and exposure to cAMP activators such as IBMX and Forskolin benefit PHH attachment. We found that activating EPAC2, the cAMP downstream effector, by S-220 significantly increased PHH attachment. We also found that EPAC2 activation induced bile canaliculi formation in iPS-derived hepatocytes. Combining these factors in studies involving PHH or iPS-hepatocyte culture provides promising means to improve cell attachment and maintenance of hepatic function.

Disease surveillance evaluation of primary small-bowel follicular lymphoma using capsule endoscopy images based on a deep convolutional neural network (with video).

BACKGROUND AND AIMS: Capsule endoscopy (CE) is useful in evaluating disease surveillance for primary small-bowel follicular lymphoma (FL), but some cases are difficult to evaluate objectively. This study evaluated the usefulness of a deep convolutional neural network (CNN) system using CE images for disease surveillance of primary small-bowel FL. METHODS: We enrolled 26 consecutive patients with primary small-bowel FL diagnosed between January 2011 and January 2021 who underwent CE before and after a watch-and-wait strategy or chemotherapy. Disease surveillance by the CNN system was evaluated by the percentage of FL-detected images among all CE images of the small-bowel mucosa. RESULTS: Eighteen cases (69%) were managed with a watch-and-wait approach, and 8 cases (31%) were treated with chemotherapy. Among the 18 cases managed with the watch-and-wait approach, the outcome of lesion evaluation by the CNN system was almost the same in 13 cases (72%), aggravation in 4 (22%), and improvement in 1 (6%). Among the 8 cases treated with chemotherapy, the outcome of lesion evaluation by the CNN system was improvement in 5 cases (63%), almost the same in 2 (25%), and aggravation in 1 (12%). The physician and CNN system reported similar results regarding disease surveillance evaluation in 23 of 26 cases (88%), whereas a discrepancy between the 2 was found in the remaining 3 cases (12%), attributed to poor small-bowel cleansing level. CONCLUSIONS: Disease surveillance evaluation of primary small-bowel FL using CE images by the developed CNN system was useful under the condition of excellent small-bowel cleansing level.

Activation of CWI pathway through high hydrostatic pressure, enhancing glycerol efflux via the aquaglyceroporin Fps1 in Saccharomyces cerevisiae.

The fungal cell wall is the initial barrier for the fungi against diverse external stresses, such as osmolarity changes, harmful drugs, and mechanical injuries. This study explores the roles of osmoregulation and the cell-wall integrity (CWI) pathway in response to high hydrostatic pressure in the yeast Saccharomyces cerevisiae. We demonstrate the roles of the transmembrane mechanosensor Wsc1 and aquaglyceroporin Fps1 in a general mechanism to maintain cell growth under high-pressure regimes. The promotion of water influx into cells at 25 MPa, as evident by an increase in cell volume and a loss of the plasma membrane eisosome structure, activates the CWI pathway through the function of Wsc1. Phosphorylation of Slt2, the downstream mitogen-activated protein kinase, was increased at 25 MPa. Glycerol efflux increases via Fps1 phosphorylation, which is initiated by downstream components of the CWI pathway, and contributes to the reduction in intracellular osmolarity under high pressure. The elucidation of the mechanisms underlying adaptation to high pressure through the well-established CWI pathway could potentially translate to mammalian cells and provide novel insights into cellular mechanosensation.

The Effect of Vitamin D3 and Valproic Acid on the Maturation of Human-Induced Pluripotent Stem Cell-Derived Enterocyte-Like Cells.

Cytochrome P450 3A4 (CYP3A4) is involved in first-pass metabolism in the small intestine and is heavily implicated in oral drug bioavailability and pharmacokinetics. We previously reported that vitamin D3 (VD3), a known CYP enzyme inducer, induces functional maturation of iPSC-derived enterocyte-like cells (iPSC-ent). Here, we identified a Notch activator and CYP modulator valproic acid (VPA), as a promotor for the maturation of iPSC-ent. We performed bulk RNA sequencing to investigate the changes in gene expression during the differentiation and maturation periods of these cells. VPA potentiated gene expression of key enterocyte markers ALPI, FABP2, and transporters such as SULT1B1. RNA-sequencing analysis further elucidated several function-related pathways involved in fatty acid metabolism, significantly upregulated by VPA when combined with VD3. Particularly, VPA treatment in tandem with VD3 significantly upregulated key regulators of enterohepatic circulation, such as FGF19, apical bile acid transporter SLCO1A2 and basolateral bile acid transporters SLC51A and SLC51B. To sum up, we could ascertain the genetic profile of our iPSC-ent cells to be specialized toward fatty acid absorption and metabolism instead of transporting other nutrients, such as amino acids, with the addition of VD3 and VPA in tandem. Together, these results suggest the possible application of VPA-treated iPSC-ent for modelling enterohepatic circulation.

A single-center prospective study evaluating the usefulness of artificial intelligence for the diagnosis of esophageal squamous cell carcinoma in a real-time setting.

BACKGROUND: Several pre-clinical studies have reported the usefulness of artificial intelligence (AI) systems in the diagnosis of esophageal squamous cell carcinoma (ESCC). We conducted this study to evaluate the usefulness of an AI system for real-time diagnosis of ESCC in a clinical setting. METHODS: This study followed a single-center prospective single-arm non-inferiority design. Patients at high risk for ESCC were recruited and real-time diagnosis by the AI system was compared with that of endoscopists for lesions suspected to be ESCC. The primary outcomes were the diagnostic accuracy of the AI system and endoscopists. The secondary outcomes were sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and adverse events. RESULTS: A total of 237 lesions were evaluated. The accuracy, sensitivity, and specificity of the AI system were 80.6%, 68.2%, and 83.4%, respectively. The accuracy, sensitivity, and specificity of endoscopists were 85.7%, 61.4%, and 91.2%, respectively. The difference between the accuracy of the AI system and that of the endoscopists was - 5.1%, and the lower limit of the 90% confidence interval was less than the non-inferiority margin. CONCLUSIONS: The non-inferiority of the AI system in comparison with endoscopists in the real-time diagnosis of ESCC in a clinical setting was not proven. TRIAL REGISTRATION: Japan Registry of Clinical Trials (jRCTs052200015, 18/05/2020).

A new evaluation system of iron bioavailability in seaweed.

In marine ecosystems, the avid binding of iron (Fe) to organic ligands influences Fe bioavailability in seaweed. This study aimed to elucidate Fe's biological availability to seaweed and develop a simple and rapid bioassay method as a new evaluation system. Undaria pinnatifida was used as a model seaweed species and the actual seaweed samples were collected using the 0.5 m × 0.5 m quadrat from the Mashike Bay area of Hokkaido, Japan. Chlorophyll fluorescence measurements were utilized as an index to evaluate the biological -effectiveness of Fe and compared with the results of culture tests based on growth. The effect of Fe content on media, pre-culture, concentrations and types of chelating and reducing agents in clearing solutions, cleaning time, Fe removal effect, and resistance to seaweed were systematically optimized to obtain the maximum efficacy of the washing solution. A bioassay was developed to evaluate the Fe environment by combining chlorophyll fluorescence measurements. The findings suggest that the tolerance of seaweeds to the wash solution is strongly influenced by the concentrations of the chelating and reducing agents than their types. Washing with 0.02 M Ti-Citrate/EDTA solution for 80 s was the most effective in terms of maximum Fe removal with minimum cell damage. The application of pre-culture and chemical pre-treatment methods under Fe deficiency to the culture strain confirmed the maximum reproducibility in the culture test. Finally, the developed method was applied to actual seaweed samples and was found to be applicable to many seaweed species. However, the method was less robust for some seaweed species and depended on the seaweed growth stage.

Liquid-based cytology for differentiating two cases of pemphigus vulgaris from oral squamous cell carcinoma.

Pemphigus vulgaris (PV) is a rare autoimmune disease characterized by blisters on the skin and mucous membrane. Since it often appears in the oral mucosa first, it may be diagnosed by oral mucosal cytology. Although the cytologic finding is characterized by acantholytic cells, that is, Tzanck cells, it is important to distinguish PV from neoplastic lesions of the oral mucosal epithelium, including differentiation from atypical parabasal/basal cells, which appear in squamous cell carcinoma (SCC). In this study, we examined the cellular findings in two cases of PV and a case of well-differentiated SCC with loss of epithelial cell cohesion. The samples were prepared using liquid-based cytology, which showed small round-shaped and deeply stained atypical, orangeophilic keratinocytes not only in SCC but also in PV, which made differentiation between the two difficult. However, Tzanck cells found in PV differ from the deep atypical parabasal/basal cells of SCC, suggesting that the cell outline is indistinct and small protrusions and brush-like structures are observed. This feature of Tzanck cells may be useful in cytological judgment.

A strategy for addicting transgene-free bacteria to synthetic modified metabolites.

Biological containment is a safeguard technology to prevent uncontrolled proliferation of "useful but dangerous" microbes. Addiction to synthetic chemicals is ideal for biological containment, but this currently requires introduction of transgenes containing synthetic genetic elements for which environmental diffusion has to be prevented. Here, I designed a strategy for addicting transgene-free bacteria to synthetic modified metabolites, in which the target organism that can neither produce an essential metabolite nor use the extracellularly supplied metabolite, is rescued by a synthetic derivative that is taken up from a medium and converted into the metabolite in the cell. Because design of the synthetic modified metabolite is the key technology, our strategy differs distinctly from conventional biological containment, which mainly depends on genetic manipulation of the target microorganisms. Our strategy is particularly promising for containment of non-genetically modified organisms such as pathogens and live vaccines.

Generation of induced pluripotent stem cell-derived beta-cells in blood amino acids-like medium.

Traditional cell culture media do not accurately represent the availability of the nutrients in plasma. They usually contain a supraphysiological concentration of nutrients such as glucose, amino acids, etc. These high nutrients can alter the metabolism of cultured cells and induce metabolic phenotypes that do not reflect in vivo conditions. We demonstrate that the supraphysiological levels of nutrients interfere with endodermal differentiation. Refinement of media formulations has a potential application in maturity modulation of stem cell-derived ß-cells (SC-ß) generation in vitro. To address these issues, we established a defined culture system to derive SC-ß-cells using a blood amino acids-like medium (BALM). Human induced pluripotent stem cells (hiPSCs) can be efficiently differentiated into the definitive endoderm, pancreatic progenitors, endocrine progenitors, and SC-ß in BALM-based med. The differentiated cells secreted C-peptide in vitro in response to high glucose levels and expressed several pancreatic ß-cell markers. In conclusion, amino acids at the physiological levels are sufficient for deriving functional SC-ß cells.