Empowering powdered activated carbon (PAC) with 3D printing: Achieving highly efficient and reusable cationic dye removal.

Powdered activated carbon (PAC) has been extensively used as an effective adsorbent. Despite its excellent adsorption ability, PAC has drawbacks, including difficulty in filtration and reactivation after use, limitations of mass transfer in deeper areas because of its aggregated powder form, and limited applicability in high-flow systems. To overcome these limitations, we used a three-dimensional (3D) printing system to fabricate PAC into a 3D structure. Spectral and microscopic analyses indicated that PAC was embedded into 3D monolith and exhibited high porosity suitable for facile mass transfer. The designed 3D PAC filter effectively removed 200 ppm-methylene blue (MB) within 8 h and showed an adsorption efficiency of 93.4 ± 0.9%. The adsorption of MB onto the 3D PAC filter was described by the pseudo-first-order kinetic and Freundlich isotherm models. The negatively charged 3D PAC filter might attract the positively charged MB, thus favoring the physical adsorption of MB onto the 3D PAC filter. The adsorption performance of the 3D PAC filter was tested at various pH levels of 4-10 and against MB spiked in seawaters and freshwaters to evaluate its feasibility for use in real environments. Finally, the reproducibility and reusability of the 3D PAC filter were demonstrated through repeated adsorption and desorption processes against MB.

Epigenetic repression of CHCHD2 enhances survival from single cell dissociation through attenuated Rho A kinase activity.

During in vitro culture, human pluripotent stem cells (hPSCs) often acquire survival advantages characterized by decreased susceptibility to mitochondrial cell death, known as "culture adaptation." This adaptation is associated with genetic and epigenetic abnormalities, including TP53 mutations, copy number variations, trisomy, and methylation changes. Understanding the molecular mechanisms underlying this acquired survival advantage is crucial for safe hPSC-based cell therapies. Through transcriptome and methylome analysis, we discovered that the epigenetic repression of CHCHD2, a mitochondrial protein, is a common occurrence during in vitro culture using enzymatic dissociation. We confirmed this finding through genetic perturbation and reconstitution experiments in normal human embryonic stem cells (hESCs). Loss of CHCHD2 expression conferred resistance to single cell dissociation-induced cell death, a common stress encountered during in vitro culture. Importantly, we found that the downregulation of CHCHD2 significantly attenuates the activity of Rho-associated protein kinase (ROCK), which is responsible for inducing single cell death in hESCs. This suggests that hESCs may survive routine enzyme-based cell dissociation by downregulating CHCHD2 and thereby attenuating ROCK activity. These findings provide insights into the mechanisms by which hPSCs acquire survival advantages and adapt to in vitro culture conditions.

Elucidation of molecular basis of osteolytic bone lesions in advanced multiple myeloma.

Osteolytic bone lesion is a major cause of decreased quality of life and poor prognosis in patients with multiple myeloma (MM), but molecular pathogenesis of the osteolytic process in MM remains elusive. Fms-like tyrosine kinase 3 ligand (FLT3L) was reported to be elevated in bone marrow and blood of patients with advanced MM who often show osteolysis. Here, we investigated a functional link of FLT3L to osteolytic process in MM. We recruited 86, 306 and 52 patients with MM, acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL), respectively. FLT3L levels of patients with hematologic malignancies were measured in bone marrow-derived plasma and found to be significantly elevated in MM than in AML or ALL that rarely show osteolysis. FLT3L levels were further elevated in MM patients with bone lesion compared with patients without bone lesion. In vitro cell-based assays showed that the administration of FLT3L to HEK293T, HeLa and U2OS cells led to an increase in the DKK1 transcript level through STAT3 phosphorylation at tyrosine 705. WNT reporter assay showed that FLT3L treatment reduced WNT signaling, and nuclear translocation of ß-catenin. These results collectively show that FLT3L-STAT3-DKK1 pathway inhibits WNT signaling-mediated bone formation in MM, which can cause osteolytic bone lesion. Finally, transcriptomic profiles revealed that FLT3L and DKK1 were predominantly elevated in the hyperdiploidy subtype of MM. Taken together, FLT3L can serve as a promising biomarker for predicting osteolytic bone lesion and also a potential therapeutic target to prohibit the progression of osteolytic process in MM with hyperdiploidy.

Rowhammer Attacks in Dynamic Random-Access Memory and Defense Methods.

This paper provides a comprehensive overview of the security vulnerability known as rowhammer in Dynamic Random-Access Memory (DRAM). While DRAM offers many desirable advantages, including low latency, high density, and cost-effectiveness, rowhammer vulnerability, first identified in 2014, poses a significant threat to computing systems. Rowhammer attacks involve repetitive access to specific DRAM rows, which can cause bit flips in neighboring rows, potentially compromising system credentials, integrity, and availability. The paper discusses the various stages of rowhammer attacks, explores existing attack techniques, and examines defense strategies. It also emphasizes the importance of understanding DRAM organization and the associated security challenges.

Probing Cation Effects on *CO Intermediates from Electroreduction of CO2 through Operando Raman Spectroscopy.

Cations in an electrolyte modulate microenvironments near the catalyst surface and affect product distribution from an electrochemical CO2 reduction reaction, and thus, their interaction with intermediate states has been tried to be probed. Herein, we directly observed the cation effect on *CO intermediates on the Cu(OH)2-derived catalyst in real time through operando surface-enhanced Raman spectroscopy at high overpotentials (-1.0 VRHE). Atop *CO peaks are composed of low-frequency binding *CO (*COLFB) and high-frequency binding *CO (*COHFB) because of their adsorption sites. These two *CO intermediates are found to have different sensitivities to the cation-induced field, and each *CO is proposed to be suitably stabilized for efficient C-C coupling. The proportions between *COHFB and *COLFB are dependent on the type of alkali cations, and the increases in the *COHFB ratio have a high correlation with selective C2H4 production under K+ and Cs+, indicating that *COHFB is the dominant and fast active species. In addition, as the hydrated cation size decreases, *COLFB is more sensitively red-shifted than *COHFB, which promotes C-C coupling and suppresses C1 products. Through time-resolved operando measurements, dynamic changes between the two *CO species are observed, showing the rapid initial adsorption of *COHFB and subsequently reaching a steady ratio between *COLFB and *COHFB.

MutSα and MutSß as size-dependent cellular determinants for prime editing in human embryonic stem cells.

Precise genome editing in human pluripotent stem cells (hPSCs) has potential applications in isogenic disease modeling and ex vivo stem cell therapy, necessitating diverse genome editing tools. However, unlike differentiated somatic cells, hPSCs have unique cellular properties that maintain genome integrity, which largely determine the overall efficiency of an editing tool. Considering the high demand for prime editors (PEs), it is imperative to characterize the key molecular determinants of PE outcomes in hPSCs. Through homozygous knockout (KO) of MMR pathway key proteins MSH2, MSH3, and MSH6, we reveal that MutSα and MutSß determine PE efficiency in an editing size-dependent manner. Notably, MSH2 perturbation disrupted both MutSα and MutSß complexes, dramatically escalating PE efficiency from base mispair to 10 bases, up to 50 folds. Similarly, impaired MutSα by MSH6 KO improved editing efficiency from single to three base pairs, while defective MutSß by MSH3 KO heightened efficiency from three to 10 base pairs. Thus, the size-dependent effect of MutSα and MutSß on prime editing implies that MMR is a vital PE efficiency determinant in hPSCs and highlights the distinct roles of MutSα and MutSß in its outcome.

Gene editing with 'pencil' rather than 'scissors' in human pluripotent stem cells.

Owing to the advances in genome editing technologies, research on human pluripotent stem cells (hPSCs) have recently undergone breakthroughs that enable precise alteration of desired nucleotide bases in hPSCs for the creation of isogenic disease models or for autologous ex vivo cell therapy. As pathogenic variants largely consist of point mutations, precise substitution of mutated bases in hPSCs allows researchers study disease mechanisms with "disease-in-a-dish" and provide functionally repaired cells to patients for cell therapy. To this end, in addition to utilizing the conventional homologous directed repair system in the knock-in strategy based on endonuclease activity of Cas9 (i.e., 'scissors' like gene editing), diverse toolkits for editing the desirable bases (i.e., 'pencils' like gene editing) that avoid the accidental insertion and deletion (indel) mutations as well as large harmful deletions have been developed. In this review, we summarize the recent progress in genome editing methodologies and employment of hPSCs for future translational applications.

Bacterial degradation kinetics of poly(Ɛ-caprolactone) (PCL) film by Aquabacterium sp. CY2-9 isolated from plastic-contaminated landfill.

Despite the identification of numerous bioplastic-degrading bacteria, the inconsistent rate of bioplastic degradation under differing cultivation conditions limits the intercomparison of results on biodegradation kinetics. In this study, we isolated a poly (Ɛ-caprolactone) (PCL)-degrading bacterium from a plastic-contaminated landfill and determined the principle-based biodegradation kinetics in a confined model system of varying cultivation conditions. Bacterial degradation of PCL films synthesized by different polymer number average molecular weights (Mn) and concentrations (% w/v) was investigated using both solid and liquid media at various temperatures. As a result, the most active gram-negative bacterial strain at ambient temperature (28 °C), designated CY2-9, was identified as Aquabacterium sp. Based on 16 S rRNA gene analysis. A clear zone around the bacterial colony was apparently exhibited during solid cultivation, and the diameter sizes increased with incubation time. During biodegradation processes in the PCL film, the thermal stability declined (determined by TGA; weight changes at critical temperature), whereas the crystalline proportion increased (determined by DSC; phase transition with temperature increment), implying preferential degradation of the amorphous region in the polymer structure. The surface morphologies (determined by SEM; electron optical system) were gradually hydrolyzed, creating destruction patterns as well as alterations in functional groups on film surfaces (determined by FT-IR; infrared spectrum of absorption or emission). In the kinetic study based on the weight loss of the PCL film (4.5 × 104 Da, 1% w/v), ∼1.5 (>±0.1) × 10-1 day-1 was obtained from linear regression for both solid and liquid media cultivation at 28 °C. The biodegradation efficiencies increased proportionally by a factor of 2.6-7.9, depending on the lower polymer number average molecular weight and lower concentration. Overall, our results are useful for measuring and/or predicting the degradation rates of PCL films by microorganisms in natural environments.

Circular RNAs in and out of Cells: Therapeutic Usages of Circular RNAs.

RNAs are versatile molecules that are primarily involved in gene regulation and can thus be widely used to advance the fields of therapeutics and diagnostics. In particular, circular RNAs which are highly stable, have emerged as strong candidates for use on next-generation therapeutic platforms. Endogenous circular RNAs control gene regulatory networks by interacting with other biomolecules or through translation into polypeptides. Circular RNAs exhibit cell-type specific expression patterns, which can be altered in tissues and body fluids depending on pathophysiological conditions. Circular RNAs that are aberrantly expressed in diseases can function as biomarkers or therapeutic targets. Moreover, exogenous circular RNAs synthesized in vitro can be introduced into cells as therapeutic molecules to modulate gene expression networks in vivo. Depending on the purpose, synthetic circular RNA sequences can either be identical to endogenous circular RNA sequences or artificially designed. In this review, we introduce the life cycle and known functions of intracellular circular RNAs. The current stage of endogenous circular RNAs as biomarkers and therapeutic targets is also described. Finally, approaches and considerations that are important for applying the available knowledge on endogenous circular RNAs to design exogenous circular RNAs for therapeutic purposes are presented.

Impaired Blastocyst Formation in Lnx2-Knockdown Mouse Embryos.

Ligand of Numb-protein X 2 (LNX2) is an E3 ubiquitin ligase that is known to regulate Notch signaling by participating in NUMB protein degradation. Notch signaling is important for differentiation and proliferation in mammals, and plays a significant role in blastocyst formation during early embryonic development. In this study, we investigated Lnx2 in mouse preimplantation embryos. Expression analysis showed that Lnx2 is expressed in oocytes and preimplantation embryos. Lnx2-knockdown embryos normally progress to the morula stage, but the majority of them do not develop into normal blastocysts. Transcript analysis revealed that the expression levels of genes critical for cell lineage specification, including octamer-binding transcription factor 4 (Oct4), are increased in Lnx2 knockdown embryos. Furthermore, the expression levels of Notch and Hippo signaling-related genes are also increased by Lnx2 knockdown. Collectively, our results show that Lnx2 is important for blastocyst formation in mice, suggest that this may act via lineage specification of inner cell mass, and further show that Lnx2 may be involved in transcriptionally regulating various genes implicated in early embryonic development.

Distinct Bacterial and Fungal Communities Colonizing Waste Plastic Films Buried for More Than 20 Years in Four Landfill Sites in Korea.

Plastic pollution has been recognized as a serious environmental problem, and microbial degradation of plastics is a potential, environmentally friendly solution to this. Here, we analyzed and compared microbial communities on waste plastic films (WPFs) buried for long periods at four landfill sites with those in nearby soils to identify microbes with the potential to degrade plastics. Fourier-transform infrared spectroscopy spectra of these WPFs showed that most were polyethylene and had signs of oxidation, such as carbon-carbon double bonds, carbon-oxygen single bonds, or hydrogen-oxygen single bonds, but the presence of carbonyl groups was rare. The species richness and diversity of the bacterial and fungal communities on the films were generally lower than those in nearby soils. Principal coordinate analysis of the bacterial and fungal communities showed that their overall structures were determined by their geographical locations; however, the microbial communities on the films were generally different from those in the soils. For the pulled data from the four landfill sites, the relative abundances of Bradyrhizobiaceae, Pseudarthrobacter, Myxococcales, Sphingomonas, and Spartobacteria were higher on films than in soils at the bacterial genus level. At the species level, operational taxonomic units classified as Bradyrhizobiaceae and Pseudarthrobacter in bacteria and Mortierella in fungi were enriched on the films. PICRUSt analysis showed that the predicted functions related to amino acid and carbohydrate metabolism and xenobiotic degradation were more abundant on films than in soils. These results suggest that specific microbial groups were enriched on the WPFs and may be involved in plastic degradation.

Horticoccus luteus gen. nov., sp. nov., A Novel Member of the Phylum Verrucomicrobia Isolated from a Dichlorodiphenyltrichloroethane (DDT)-Contaminated Orchard Soil.

Strain KSB-15 T was isolated from an orchard soil that had been contaminated with the insecticide dichlorodiphenyltrichloroethane for about 60 years. The 16S rRNA gene sequence of this strain showed the highest sequence similarities with those of Oleiharenicola alkalitolerans NVTT (95.3%), Opitutus terrae PB90-1 T (94.8%), and Oleiharenicola lentus TWA-58 T (94.7%) among type strains, which are members of the family Opitutaceae within the phylum Verrucomicrobia. Strain KSB-15 T was an obligate aerobe, Gram-negative, non-motile, coccoid or short rod with the cellular dimensions of 0.37-0.62 µm width and 0.43-0.72 µm length. The strain grew at temperatures between 15-37 °C (optimum, 25 °C), at a pH range of 5.0-11.0 (optimum, pH 6.0), and at a NaCl concentration of 0-3% (w/v) (optimum, 0%). It contained menaquinone-7 (MK-7) as the major isoprenoid quinone (94.1%), and iso-C15:0 (34.9%) and anteiso-C15:0 (29.0%) as the two major fatty acids. The genome of strain KSB-15 T was composed of one chromosome with a total size of 4,320,198 bp, a G + C content of 64.3%, 3,393 coding genes (CDS), 14 pseudogenes, and 52 RNA genes. The OrthoANIu values, In silico DDH values and average amino acid identities between strain KSB-15 T and the members of the family Opitutaceae were 71.6 ~ 73.0%, 19.0 ~ 19.9%, and 55.9 ~ 62.0%, respectively. On the basis of our polyphasic taxonomic study, we conclude that strain KSB-15 T should be classified as a novel genus of the family Opitutaceae, for which the name Horticcoccus luteus gen. nov., sp. nov. is proposed.The type strain is KSB-15 T (= KACC 22271 T = DSM 113638 T).

Gene Expression Profiles Identify Biomarkers of Resistance to Decitabine in Myelodysplastic Syndromes.

Myelodysplastic syndrome (MDS) is a clonal hematopoietic stem cell disease characterized by inefficient hematopoiesis and the potential development of acute leukemia. Among the most notable advances in the treatment of MDS is the hypomethylating agent, decitabine (5-aza-2'deoxycytidine). Although decitabine is well known as an effective method for treating MDS patients, only a subset of patients respond and a tolerance often develops, leading to treatment failure. Moreover, decitabine treatment is costly and causes unnecessary toxicity. Therefore, clarifying the mechanism of decitabine resistance is important for improving its therapeutic efficacy. To this end, we established a decitabine-resistant F-36P cell line from the parental F-36P leukemia cell line, and applied a genetic approach employing next-generation sequencing, various experimental techniques, and bioinformatics tools to determine differences in gene expression and relationships among genes. Thirty-eight candidate genes encoding proteins involved in decitabine-resistant-related pathways, including immune checkpoints, the regulation of myeloid cell differentiation, and PI3K-Akt signaling, were identified. Interestingly, two of the candidate genes, AKT3 and FOS, were overexpressed in MDS patients with poor prognoses. On the basis of these results, we are pursuing development of a gene chip for diagnosing decitabine resistance in MDS patients, with the goal of ultimately improving the power to predict treatment strategies and the prognosis of MDS patients.

Endophytic Isaria javanica pf185 Persists after Spraying and Controls Myzus persicae (Hemiptera: Aphididae) and Colletotrichum acutatum (Glomerellales: Glomerellaceae) in Pepper.

This study endeavored to sustainably control aphids and anthracnose after spraying endophytic Isaria javanica pf185 under field conditions. Under two different tents; one batch of seedlings was sprayed with a 107 conidia/mL I. javanica pf185 suspension; while another was sprayed with 0.05% Tween 80® in distilled water. Six leaf discs from the top; middle; and bottom part of the plant canopy were weekly collected and placed on moistened filter paper in a Petri dish for insecticidal and antifungal bioassays against Myzus persicae and Colletotrichum acutatum. Differences were noticed from the 18th day after spraying with mortality (86.67 ± 0.57% versus 36.67 ± 0.64%) and leaf damage (13.45 ± 0.03% versus 41.18 ± 0.06%) on fungus-treated and controlled, respectively. The corrected insecticidal efficacy was 20.43, 39.82, 72.32, 66.43 and 70.04%, while the corrected fungicidal efficacy was 26.07, 38.01, 53.35, 29.08 and 41.81% during five successive weeks. A positive correlation was evident between insecticidal efficacy and relative humidity (r2 = 0.620) and temperature (r2 = 0.424), respectively. No correlation was found between antifungal activity and relative humidity (r2 = 0.061) and temperature (r2 = 0), respectively. The entomopathogenic fungus survived on leaf surface area and in tissues after spraying.

Promyelocytic Leukemia Proteins Regulate Fanconi Anemia Gene Expression.

Promyelocytic leukemia (PML) protein is the core component of subnuclear structures called PML nuclear bodies that are known to play important roles in cell survival, DNA damage responses, and DNA repair. Fanconi anemia (FA) proteins are required for repairing interstrand DNA crosslinks (ICLs). Here we report a novel role of PML proteins, regulating the ICL repair pathway. We found that depletion of the PML protein led to the significant reduction of damage-induced FANCD2 mono-ubiquitination and FANCD2 foci formation. Consistently, the cells treated with siRNA against PML showed enhanced sensitivity to a crosslinking agent, mitomycin C. Further studies showed that depletion of PML reduced the protein expression of FANCA, FANCG, and FANCD2 via reduced transcriptional activity. Interestingly, we observed that damage-induced CHK1 phosphorylation was severely impaired in cells with depleted PML, and we demonstrated that CHK1 regulates FANCA, FANCG, and FANCD2 transcription. Finally, we showed that inhibition of CHK1 phosphorylation further sensitized cancer cells to mitomycin C. Taken together, these findings suggest that the PML is critical for damage-induced CHK1 phosphorylation, which is important for FA gene expression and for repairing ICLs.

Gene Expression Profiles Associated with Radio-Responsiveness in Locally Advanced Rectal Cancer.

LARC patients were sorted according to their radio-responsiveness and patient-derived organoids were established from the respective cancer tissues. Expression profiles for each group were obtained using RNA-seq. Biological and bioinformatic analysis approaches were used in deciphering genes and pathways that participate in the radio-resistance of LARC. Thirty candidate genes encoding proteins involved in radio-responsiveness-related pathways, including the immune system, DNA repair and cell-cycle control, were identified. Interestingly, one of the candidate genes, cathepsin E (CTSE), exhibited differential methylation at the promoter region that was inversely correlated with the radio-resistance of patient-derived organoids, suggesting that methylation status could contribute to radio-responsiveness. On the basis of these results, we plan to pursue development of a gene chip for diagnosing the radio-responsiveness of LARC patients, with the hope that our efforts will ultimately improve the prognosis of LARC patients.

PROKR1 delivery by cell-derived vesicles restores the myogenic potential of Prokr1-deficient C2C12 myoblasts.

Cell-derived vesicles (CDVs) have been investigated as an alternative to exosomes. Here, we generated CDVs from Prokineticin receptor 1 (PROKR1) overexpressing HEK293T cells using micro-extrusion. More than 60 billion PROKR1-enriched CDV (PROKR1Tg CDVs) particles with canonical exosome properties were recovered from 107 cells. With 25 µg/mL of PROKR1Tg CDVs, we observed delivery of PROKR1, significant reduction of apoptosis, and myotube formation in C2C12Prokr1-/- myoblasts that have lost their myogenic potential but underwent apoptosis following myogenic commitment. Expression levels of early and late myogenic marker genes and glucose uptake capacity were restored to equivalent levels with wild-type control. Furthermore, PROKR1Tg CDVs were accumulated in soleus muscle comparable to the liver without significant differences. Therefore, CDVs obtained from genetically engineered cells appear to be an effective method of PROKR1 protein delivery and offer promise as an alternative therapy for muscular dystrophy.

Tanc2-mediated mTOR inhibition balances mTORC1/2 signaling in the developing mouse brain and human neurons.

mTOR signaling, involving mTORC1 and mTORC2 complexes, critically regulates neural development and is implicated in various brain disorders. However, we do not fully understand all of the upstream signaling components that can regulate mTOR signaling, especially in neurons. Here, we show a direct, regulated inhibition of mTOR by Tanc2, an adaptor/scaffolding protein with strong neurodevelopmental and psychiatric implications. While Tanc2-null mice show embryonic lethality, Tanc2-haploinsufficient mice survive but display mTORC1/2 hyperactivity accompanying synaptic and behavioral deficits reversed by mTOR-inhibiting rapamycin. Tanc2 interacts with and inhibits mTOR, which is suppressed by mTOR-activating serum or ketamine, a fast-acting antidepressant. Tanc2 and Deptor, also known to inhibit mTORC1/2 minimally affecting neurodevelopment, distinctly inhibit mTOR in early- and late-stage neurons. Lastly, Tanc2 inhibits mTORC1/2 in human neural progenitor cells and neurons. In summary, our findings show that Tanc2 is a mTORC1/2 inhibitor affecting neurodevelopment.

SLC6A20 transporter: a novel regulator of brain glycine homeostasis and NMDAR function.

Glycine transporters (GlyT1 and GlyT2) that regulate levels of brain glycine, an inhibitory neurotransmitter with co-agonist activity for NMDA receptors (NMDARs), have been considered to be important targets for the treatment of brain disorders with suppressed NMDAR function such as schizophrenia. However, it remains unclear whether other amino acid transporters expressed in the brain can also regulate brain glycine levels and NMDAR function. Here, we report that SLC6A20A, an amino acid transporter known to transport proline based on in vitro data but is understudied in the brain, regulates proline and glycine levels and NMDAR function in the mouse brain. SLC6A20A transcript and protein levels were abnormally increased in mice carrying a mutant PTEN protein lacking the C terminus through enhanced ß-catenin binding to the Slc6a20a gene. These mice displayed reduced extracellular levels of brain proline and glycine and decreased NMDAR currents. Elevating glycine levels back to normal ranges by antisense oligonucleotide-induced SLC6A20 knockdown, or the competitive GlyT1 antagonist sarcosine, normalized NMDAR currents and repetitive climbing behavior observed in these mice. Conversely, mice lacking SLC6A20A displayed increased extracellular glycine levels and NMDAR currents. Lastly, both mouse and human SLC6A20 proteins mediated proline and glycine transports, and SLC6A20 proteins could be detected in human neurons. These results suggest that SLC6A20 regulates proline and glycine homeostasis in the brain and that SLC6A20 inhibition has therapeutic potential for brain disorders involving NMDAR hypofunction.