A culture method with berbamine, a plant alkaloid, enhances CAR-T cell efficacy through modulating cellular metabolism.

Memory T cells demonstrate superior in vivo persistence and antitumor efficacy. However, methods for manufacturing less differentiated T cells are not yet well-established. Here, we show that producing chimeric antigen receptor (CAR)-T cells using berbamine (BBM), a natural compound found in the Chinese herbal medicine Berberis amurensis, enhances the antitumor efficacy of CAR-T cells. BBM is identified through cell-based screening of chemical compounds using induced pluripotent stem cell-derived T cells, leading to improved viability with a memory T cell phenotype. Transcriptomics and metabolomics using stem cell memory T cells reveal that BBM broadly enhances lipid metabolism. Furthermore, the addition of BBM downregulates the phosphorylation of p38 mitogen-activated protein kinase and enhanced mitochondrial respiration. CD19-CAR-T cells cultured with BBM also extend the survival of leukaemia mouse models due to their superior in vivo persistence. This technology offers a straightforward approach to enhancing the antitumor efficacy of CAR-T cells.

Mini-TCRs: Truncated T cell receptors to generate T cells from induced pluripotent stem cells.

Allogeneic T cell platforms utilizing induced pluripotent stem cell (iPSC) technology exhibit significant promise for the facilitation of adoptive immunotherapies. While mature T cell receptor (TCR) signaling plays a crucial role in generating T cells from iPSCs, the introduction of exogenous mature TCR genes carries a potential risk of causing graft-versus-host disease (GvHD). In this study, we present the development of truncated TCRα and TCRß chains, termed mini-TCRs, which lack variable domains responsible for recognizing human leukocyte antigen (HLA)-peptide complexes. We successfully induced cytotoxic T lymphocytes (CTLs) from iPSCs by employing mini-TCRs. Combinations of TCRα and TCRß fragments were screened from mini-TCR libraries based on the surface localization of CD3 proteins and their ability to transduce T cell signaling. Consequently, mini-TCR-expressing iPSCs underwent physiological T cell development, progressing from the CD4 and CD8 double-positive stage to the CD8 single-positive stage. The resulting iPSC-derived CTLs exhibited comparable cytokine production and cytotoxicity in comparison to that of full-length TCR-expressing T lymphocytes when chimeric antigen receptors (CARs) were expressed. These findings demonstrate the potential of mini-TCR-carrying iPSCs as a versatile platform for CAR T cell therapy, offering a promising avenue for advancing adoptive immunotherapies.

Multi-modal and multi-postural assessment of platypnea-orthodeoxia syndrome triggered by stroke: A case report.

Platypnea-orthodeoxia syndrome (POS) is a rare condition where patients suffer from dyspnea and reduced oxygenation while in the sitting position. A 69-year-old man initially experienced dyspnea and hypoxemia in the sitting position after developing hemiplegia and postural instability secondary to a cerebral hemorrhage, but the symptoms improved in the supine position. Transesophageal echocardiography revealed a patent foramen ovale (PFO). In the sitting or semi-Fowler position, increased right-left shunt was observed using Swan-Ganz catheterization and pulmonary perfusion scintigraphy. The PFO closure was performed, which obliterated dyspnea and hypoxemia in the sitting position. In POS associated with PFO, comprehensive pre-operative evaluation using multi-modality tests in different postural settings critically delineates the etiology that guides appropriate treatment strategy. .

Impact of high-density mapping on outcome of the second ablation for atrial fibrillation.

PURPOSE: Identification of a conduction gap between the left atrium and pulmonary vein (LA-PV gap) and the circuit of atrial tachycardia after pulmonary vein isolation (PVI) is an important process during the second ablation for atrial fibrillation (AF). The high-density mapping system RHYTHMIA® is useful for identification of an LA-PV gap and the circuit of atrial tachycardia. Therefore, this study was performed to investigate the effect of RHYTHMIA® in terms of the outcome of the second ablation for AF. METHODS: One hundred patients underwent a second ablation for AF in our institute from April 2015 to December 2018. We retrospectively evaluated 49 patients using RHYTHMIA® (group 1) and 51 patients using the conventional method with additional anatomical guide by CARTO® system. RESULTS: In group 1, we performed redo PVI for 41 patients with 49 LA-PV countable gaps and ablation for other atrial arrhythmias in 7 patients. In group 2, we performed redo PVI in 40 patients with 33 LA-PV countable gaps and ablation for other atrial arrhythmias in 9 patients. Three and two unstable arrhythmias in each group were not ablated. The final recurrence of atrial arrhythmia was significantly lower in group 1 than 2 (8/49 (16%) vs. 17/51 (33%), respectively; P = 0.017). Atrial arrhythmias other than AF after the second ablation occurred in only one patient in group 1 but seven patients in group 2. CONCLUSION: Using high-density mapping for the second ablation of AF was found to be superior to the conventional ablation method in terms of the suppression of atrial events in this study. This technique warrants further investigation.

Myopotential Oversensing Is a Major Cause of Inappropriate Shock in Subcutaneous Implantable Defibrillator in Japan.

Previous study has identified marked differences in patient characteristics and causes of inappropriate shock (IAS) between Japan and the Western societies in terms of subcutaneous implantable cardioverter-defibrillator (S-ICD). However, evidence of IAS in Asian populations including Japan has been limited to one observational study.Thus, we conducted a single-center registry study that tracks the postoperative course of 61 consecutive patients who received S-ICD from February 2016 to January 2020. Our findings showed that IAS occurred in 9.8% of the study population (6/61), which is comparable to the previously reported incidence. Remarkably, T-wave oversensing did not result in an IAS (0/6). Instead, myopotential oversensing was determined to have caused the most IAS events (4/6), while atrial fibrillation ranked second (2/6). A provocation maneuver (e.g., abdominal clench, push-ups, lifting a heavy item) reproduced myopotential noise disguised as R-waves, which should potentially trigger an IAS if uninterrupted. R-wave amplitude of the IAS group appeared relatively low compared to that of the non-IAS group although this finding was not tested significant. Furthermore, no temporal changes were noted in R-wave amplitude between the time of implantation and IAS events, suggesting that it is neither constantly low nor acutely dropped R-wave amplitude but a relatively high noise level that drives IAS. All the myopotential-IAS patients were found to be male. Right-sided lead implantation was associated with a higher incidence of IAS.This study highlights the fact that IAS continues to occur due to myopotential noise oversensing instead of T-wave oversensing. To minimize the risk of IAS, it is desirable to search and secure high R-wave voltage.

Dendritic Spine Density is Increased in Arcadlin-deleted Mouse Hippocampus.

The neural network undergoes remodeling in response to neural activity and interventions, such as antidepressants. Cell adhesion molecules that link pre- and post-synaptic membranes are responsible not only for the establishment of the neural circuitry, but also for the modulation of the strength of each synaptic connection. Among the various classes of synaptic cell adhesion molecules, a non-clustered protocadherin, Arcadlin/Paraxial protocadherin/Protocadherin-8 (Acad), is unique in that it is induced quickly in response to neural activity. Although the primary structure of Arcadlin implies its cell adhesion activity, it weakens the adhesion of N-cadherin. Furthermore, Arcadlin reduces the dendritic spine density in cultured hippocampal neurons. In order to gain an insight into the function of Arcadlin in the brain, we examined the dendritic morphologies of the hippocampal neurons in Acad-/- mice. Acad-/- mice showed a higher spine density than wild-type mice. Following an electroconvulsive seizure (ECS), which strongly induces Arcadlin in the hippocampus, the spine density gradually decreased for 8 h. ECS did not reduce the spine density of CA1 apical dendrites in Acad-/- mice. Daily intraperitoneal injection of the antidepressant fluoxetine (25 mg/kg/day) for 18 days resulted in the induction of Arcadlin in the hippocampus. This treatment reduced spine density in the dentate gyrus and CA1. Chronic fluoxetine treatment did not suppress spine density in Acad-/- mice, suggesting that fluoxetine-induced decrease in spine density is largely due to Arcadlin. The present findings confirm the spine-repulsing activity of Arcadlin and its involvement in the remodeling of hippocampal neurons in response to antidepressants.

Percutaneous Simple Lead Traction Is a Feasible and Effective Method for Right Ventricular Lead Perforations.

Right ventricular (RV) lead perforations are relatively rare but a potentially life-threatening complication of surgical implantations of cardiac implantable electronic devices (CIEDs). The result of percutaneous simple lead traction after lead perforations in the Japanese population has not been well clarified.We retrospectively studied 1359 patients (pacemakers [PMs], 973 patients; implantable cardioverter defibrillators [ICD], 386 patients) from April 2007 to December 2018 who underwent initial CIED implantation. Fifteen patients (1.1%) were diagnosed with RV lead perforations. The clinical data were evaluated in those patients, and the baseline characteristics and echocardiographic data were compared between the lead perforation group and the non-perforation group. The success and complication rates of the simple traction and repositioning of the RV lead were also assessed.The number of perforated RV leads was seven ICD leads (1.8%) and eight PM leads (0.82%). They were diagnosed on a median seven days (5.5-36.0) after the CIED implantation. Twelve patients were asymptomatic but were detected by an increased capture threshold. Three patients had pericarditis and stimulation of the diaphragm. Only one patient in the ICD lead group who took anticoagulants had a cardiac tamponade and needed an urgent pericardiocentesis (0.07%). No one required a thoracotomy or other devices related to complications after repositioning the RV lead. There was no significant difference in the baseline characteristics and echocardiographic parameters between the groups.RV lead perforations were relatively rare complications of CIED implantations. Percutaneous simple lead traction and repositioning the perforated lead was feasible and effective if the patients did not receive anticoagulants.

Ectopic expression of DNA methyltransferases DNMT3A2 and DNMT3L leads to aberrant hypermethylation and postnatal lethality in mice.

DNA methylation is generally known to inactivate gene expression. The DNA methyltransferases (DNMTs), DNMT3A and DNMT3B, catalyze somatic cell lineage-specific DNA methylation, while DNMT3A and DNMT3L catalyze germ cell lineage-specific DNA methylation. How such lineage- and gene-specific DNA methylation patterns are created remains to be elucidated. To better understand the regulatory mechanisms underlying DNA methylation, we generated transgenic mice that constitutively expressed DNMT3A and DNMT3L, and analyzed DNA methylation, gene expression, and their subsequent impact on ontogeny. All transgenic mice were born normally but died within 20 weeks accompanied with cardiac hypertrophy. Several genes were repressed in the hearts of transgenic mice compared with those in wild-type mice. CpG islands of these downregulated genes were highly methylated in the transgenic mice. This abnormal methylation occurred in the perinatal stage. Conversely, monoallelic DNA methylation at imprinted loci was faithfully maintained in all transgenic mice, except H19. Thus, the loci preferred by DNMT3A and DNMT3L differ between somatic and germ cell lineages.

CRISPR-Cas9-based photoactivatable transcription systems to induce neuronal differentiation.

Our improved CRISPR-Cas9-based photoactivatable transcription systems, CPTS2.0 and Split-CPTS2.0, enable high blue-light-inducible activation of endogenous target genes in various human cell lines. We achieved reversible activation of target genes with CPTS2.0 and induced neuronal differentiation in induced pluripotent stem cells (iPSCs) by upregulating NEUROD1 with Split-CPTS2.0.

Assessment of in vitro antiovulatory activities of nonsteroidal anti-inflammatory drugs and comparison with in vivo reproductive toxicities of medaka (Oryzias latipes).

Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used therapeutic agents; however, their pharmacological actions raise concerns about potential risks to the reproductive health of aquatic vertebrates. In the present study, a medaka ovulation assay was applied as an in vitro model to evaluate NSAID-induced antiovulatory activity. We first tested five NSAIDs, including diclofenac sodium (DCF), ketoprofen (KP), salicylic acid (SA), mefenamic acid (MA), and acetylsalicylic acid (ASA) for their antiovulatory activities toward the follicles isolated from the ovaries of spawning females. Of all the chemicals tested, DCF had the highest antiovulatory activity, with the concentration that caused 50% inhibition (IC50) (101 µM). MA was the second most potent inhibitor following DCF, but KP, SA, or ASA had little inhibitory effect on the ovulation of the follicles. The in vitro antiovulatory activity of five NSAIDs showed good correlation with data published on the inhibitory activity on human COX-2. Second, we selected DCF and SA as the most and least potent NSAIDs, respectively, and examined the effects on reproduction of intact fish in order to evaluate whether the ovulation assay was a reasonable predictor of potential reproductive effects in fish. Females exposed to DCF showed a concentration-dependent decrease in the number of spawned eggs and an increment in the gonadosomatic index (GSI), possibly due to an anovulation in the females. In contrast, neither fecundity nor the GSI of females decreased at up to 20 mg/L of SA, at which acute lethality to medaka was induced. In conclusion, the medaka ovulation assay reflected the potency of NSAID-induced antiovulatory activity and may thus serve as an in vitro model for the prediction of NSAID-induced reproductive toxicity. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1710-1719, 2016.

5-N-Arylaminothiazoles as Highly Twisted Fluorescent Monocyclic Heterocycles: Synthesis and Characterization.

A series of 5-N-arylaminothiazoles was prepared by reacting thioamide dianions derived from secondary thioamides with thioformamides, followed by sequential oxidation with iodine. X-ray analyses demonstrated that they adopt structures that are highly twisted from planar conformations. Their orientations were tuned by the steric and/or electronic interactions of the substituents at their 2-, 4-, and 5-positions. The 5-aminothiazoles exhibited a range of fluorescent emissions, from blue to orange. Although the absorption spectra were independent of the polarity of the solvent, fluorescent emissions were influenced by the polarity of the solvent: in more polar solvents, the emissions were red-shifted. These phenomena were examined in terms of Lippert-Mataga plots and the change in the dipole moment between the ground and excited states. They also exhibited emissions in the solid state, again from blue to orange. Cyclic voltammetry of the 5-aminothiazoles showed reversible waves of one-electron oxidation. The half-potential of the oxidation was reduced by the introduction of electron-donating groups to the phenyl groups on the nitrogen atom at the 5-position. DFT calculations were carried out to determine the energy levels of the HOMO and LUMO. Finally, the results of TG-DTA showed that they are thermally stable.

Speciation and identification of tellurium-containing metabolites in garlic, Allium sativum.

Tellurium (Te) is a widely used metalloid in industry because of its unique chemical and physical properties. However, information about the biological and toxicological activities of Te in plants and animals is limited. Although Te is expected to be metabolized in organisms via the same pathway as sulfur and selenium (Se), no precise metabolic pathways are known in organisms, particularly in plants. To reveal the metabolic pathway of Te in plants, garlic, a well-known Se accumulator, was chosen as the model plant. Garlic was hydroponically cultivated and exposed to sodium tellurate, and Te-containing metabolites in the water extract of garlic leaves were identified using HPLC coupled with inductively coupled plasma mass spectrometry (ICP-MS) or electrospray tandem mass spectrometry (ESI-MS-MS). At least three Te-containing metabolites were detected using HPLC-ICP-MS, and two of them were subjected to HPLC-ESI-MS-MS for identification. The MS spectra obtained by ESI-MS-MS indicated that the metabolite was Te-methyltellurocysteine oxide (MeTeCysO). Then, MeTeCysO was chemically synthesized and its chromatographic behavior matched with that of the Te-containing metabolite in garlic. The other was assigned as cysteine S-methyltellurosulfide. These results suggest that garlic can assimilate tellurate, an inorganic Te compound, and tellurate is transformed into a Te-containing amino acid, the so-called telluroamino acid. This is the first report addressing that telluroamino acid is de novo synthesized in a higher plant.