Deep learning models to predict the editing efficiencies and outcomes of diverse base editors.

Applications of base editing are frequently restricted by the requirement for a protospacer adjacent motif (PAM), and selecting the optimal base editor (BE) and single-guide RNA pair (sgRNA) for a given target can be difficult. To select for BEs and sgRNAs without extensive experimental work, we systematically compared the editing windows, outcomes and preferred motifs for seven BEs, including two cytosine BEs, two adenine BEs and three C•G to G•C BEs at thousands of target sequences. We also evaluated nine Cas9 variants that recognize different PAM sequences and developed a deep learning model, DeepCas9variants, for predicting which variants function most efficiently at sites with a given target sequence. We then develop a computational model, DeepBE, that predicts editing efficiencies and outcomes of 63 BEs that were generated by incorporating nine Cas9 variants as nickase domains into the seven BE variants. The predicted median efficiencies of BEs with DeepBE-based design were 2.9- to 20-fold higher than those of rationally designed SpCas9-containing BEs.

Augmented Decision-Making in wound Care: Evaluating the clinical utility of a Deep-Learning model for pressure injury staging.

BACKGROUND: Precise categorization of pressure injury (PI) stages is critical in determining the appropriate treatment for wound care. However, the expertise necessary for PI staging is frequently unavailable in residential care settings. OBJECTIVE: This study aimed to develop a convolutional neural network (CNN) model for classifying PIs and investigate whether its implementation can allow physicians to make better decisions for PI staging. METHODS: Using 3,098 clinical images (2,614 and 484 from internal and external datasets, respectively), a CNN was trained and validated to classify PIs and other related dermatoses. A two-part survey was conducted with 24 dermatology residents, ward nurses, and medical students to determine whether the implementation of the CNN improved initial PI classification decisions. RESULTS: The top-1 accuracy of the model was 0.793 (95% confidence interval [CI], 0.778-0.808) and 0.717 (95% CI, 0.676-0.758) over the internal and external testing sets, respectively. The accuracy of PI staging among participants was 0.501 (95% CI, 0.487-0.515) in Part I, improving by 17.1% to 0.672 (95% CI, 0.660-0.684) in Part II. Furthermore, the concordance between participants increased significantly with the use of the CNN model, with Fleiss' κ of 0.414 (95% CI, 0.410-0.417) and 0.641 (95% CI, 0.638-0.644) in Parts I and II, respectively. CONCLUSIONS: The proposed CNN model can help classify PIs and relevant dermatoses. In addition, augmented decision-making can improve consultation accuracy while ensuring concordance between the clinical decisions made by a diverse group of health professionals.

Sniper2L is a high-fidelity Cas9 variant with high activity.

Although several high-fidelity SpCas9 variants have been reported, it has been observed that this increased specificity is associated with reduced on-target activity, limiting the applications of the high-fidelity variants when efficient genome editing is required. Here, we developed an improved version of Sniper-Cas9, Sniper2L, which represents an exception to this trade-off trend as it showed higher specificity with retained high activity. We evaluated Sniper2L activities at a large number of target sequences and developed DeepSniper, a deep learning model that can predict the activity of Sniper2L. We also confirmed that Sniper2L can induce highly efficient and specific editing at a large number of target sequences when it is delivered as a ribonucleoprotein complex. Mechanically, the high specificity of Sniper2L originates from its superior ability to avoid unwinding a target DNA containing even a single mismatch. We envision that Sniper2L will be useful when efficient and specific genome editing is required.

Global burden of primary liver cancer and its association with underlying aetiologies, sociodemographic status, and sex differences from 1990-2019: A DALY-based analysis of the Global Burden of Disease 2019 study.

BACKGROUND/AIMS: Global distribution of dominant liver cancer aetiologies has significantly changed over the past decades. This study analyzed the updated temporal trends of liver cancer aetiologies and sociodemographic status in 204 countries and territories from 1990 to 2019. METHODS: The Global Burden of Disease 2019 report was used for statistical analysis. In addition, we performed stratification analysis to five quintiles using sociodemographic index and 21 geographic regions. RESULTS: The crude numbers of liver cancer disease-adjusted life years (DALYs) and deaths significantly increased during the study period (DALYs; 11,278,630 in 1990 and 12,528,422 in 2019, deaths; 365,215 in 1990 and 484,577 in 2019). However, the Age-standardized DALY and mortality rates decreased. Hepatitis B virus (HBV) remains the leading cause of liver cancer DALYs and mortality, followed by hepatitis C virus (HCV), alcohol consumption, and non-alcoholic steatohepatitis/non-alcoholic fatty liver disease (NASH/NAFLD). Although Age-standardized DALY and mortality rates of liver cancer due to HBV and HCV have decreased, the rates due to alcohol consumption and NASH/NAFLD have increased. In 2019, the population of the East Asia region had the highest Age-standardized DALY and mortality rates, followed by high-income Asia-Pacific and Central Asia populations. Although East Asia and high-income Asia-Pacific regions showed a decrease during the study period, Age-standardized DALY rates increased in Central Asia. High-income North American and Australasian populations also showed a significant increase in Age-standardized DALY. CONCLUSION: Liver cancer remains an ongoing global threat. The burden of liver cancer associated with alcohol consumption and NASH/NAFLD is markedly increasing and projected to continuously increase.

Very early environmental enrichment protects against apoptosis and improves functional recovery from hypoxic-ischemic brain injury.

Appropriate rehabilitation of stroke patients at a very early phase results in favorable outcomes. However, the optimal strategy for very early rehabilitation is at present unclear due to the limited knowledge on the effects of very early initiation of rehabilitation based on voluntary exercise (VE). Environmental enrichment (EE) is a therapeutic paradigm for laboratory animals that involves complex combinations of physical, cognitive, and social stimuli, as well as VE. Few studies delineated the effect of EE on apoptosis in very early stroke in an experimental model. Although a minimal benefit of early rehabilitation in stroke models has been claimed in previous studies, these were based on a forced exercise paradigm. The aim of this study is to determine whether very early exposure to EE can effectively regulate Fas/FasL-mediated apoptosis following hypoxic-ischemic (HI) brain injury and improve neurobehavioral function. C57Bl/6 mice were housed for 2 weeks in either cages with EE or standard cages (SC) 3 h or 72 h after HI brain injury. Very early exposure to EE was associated with greater improvement in motor function and cognitive ability, reduced volume of the infarcted area, decreased mitochondria-mediated apoptosis, and decreased oxidative stress. Very early exposure to EE significantly downregulated Fas/FasL-mediated apoptosis, decreased expression of Fas, Fas-associated death domain, cleaved caspase-8/caspase-8, cleaved caspase-3/caspase-3, as well as Bax and Bcl-2, in the cerebral cortex and the hippocampus. Delayed exposure to EE, on the other hand, failed to inhibit the extrinsic pathway of apoptosis. This study demonstrates that very early exposure to EE is a potentially useful therapeutic translation for stroke rehabilitation through effective inhibition of the extrinsic and intrinsic apoptotic pathways.

Environmental Enrichment Enhances Cav 2.1 Channel-Mediated Presynaptic Plasticity in Hypoxic-Ischemic Encephalopathy.

Hypoxic-ischemic encephalopathy (HIE) is a devastating neonatal brain condition caused by lack of oxygen and limited blood flow. Environmental enrichment (EE) is a classic paradigm with a complex stimulation of physical, cognitive, and social components. EE can exert neuroplasticity and neuroprotective effects in immature brains. However, the exact mechanism of EE on the chronic condition of HIE remains unclear. HIE was induced by a permanent ligation of the right carotid artery, followed by an 8% O2 hypoxic condition for 1 h. At 6 weeks of age, HIE mice were randomly assigned to either standard cages or EE cages. In the behavioral assessments, EE mice showed significantly improved motor performances in rotarod tests, ladder walking tests, and hanging wire tests, compared with HIE control mice. EE mice also significantly enhanced cognitive performances in Y-maze tests. Particularly, EE mice showed a significant increase in Cav 2.1 (P/Q type) and presynaptic proteins by molecular assessments, and a significant increase of Cav 2.1 in histological assessments of the cerebral cortex and hippocampus. These results indicate that EE can upregulate the expression of the Cav 2.1 channel and presynaptic proteins related to the synaptic vesicle cycle and neurotransmitter release, which may be responsible for motor and cognitive improvements in HIE.

Doxycycline-Loaded Nitric Oxide-Releasing Nanomatrix Gel in Replanted Rat Molar on Pulp Regeneration.

The aim of the present study was to evaluate the effect of doxycycline-loaded NO-releasing nanomatrix gel on pulp regeneration in replantation of avulsed rat teeth. A total of 28 maxillary first molars extracted from rats were replanted. The rats were divided into two groups based on the use of root surface treatment: doxycycline-loaded NO-releasing nanomatrix group and no treatment. Eight weeks after replantation, the rats were sacrificed, and the teeth were evaluated using histomorphometric analysis. On histomorphometric analysis, the NO-releasing nanomatrix group demonstrated a significantly lower grade of pulp inflammation (1.00 ± 1.11, mean ± standard deviation) compared to the no treatment group (2.21 ± 1.25, p = 0.014). NO-releasing nanomatrix group showed a significantly higher grade of pulp regeneration (2.57 ± 0.85, p = 0.012) and significantly lower grade of pulp inflammation (1.00 ± 0.68, p = 0.025) compared to the no treatment group. In conclusion, NO-releasing nanomatrix gel improved pulp regeneration of replanted teeth, though the sample size of this study was rather small. Within the limits of this study, NO-releasing nanomatrix gel can provide more favorable pulpal regeneration despite replantation.

Association between Blood Heavy Metal Levels and Predicted 10-Year Risk for A First Atherosclerosis Cardiovascular Disease in the General Korean Population.

BACKGROUND: Atherosclerotic cardiovascular disease (ASCVD) is a preventable type of disease, thus, specifying factors that increase the occurrence of this type of disease is needed. Heavy metals such as cadmium (Cd), lead (Pb), and mercury (Hg) have been suggested as possible factors influencing the development of cardiovascular disease. We aimed to link blood heavy metal levels (Cd, Pb, Hg) with 10-year ASCVD risk scores. METHODS: A population of 993 men and 1431 women who participated in the Korea National Health and Nutrition Examination Survey (KNHANES) were included. The 2013 American College of Cardiology/American Heart Association (ACC/AHA) pooled cohort equations risk prediction model and Korean Risk Prediction Model (KRPM) were used as means for risk prediction. Following multivariate adjustment; blood Cd; Pb; and Hg levels were divided into quartiles for analysis using linear trends estimation and multiple regression models. RESULTS: There was an overall positive trend between blood Cd, Pb, and Hg levels and 10-year ASCVD risk scores; KRPM risk score increasing by quartile for blood Cd (men p < 0.0001, women p = 0.0024), Pb (men p = 0.0097, women p = 0.0330), Hg (men p = 0.0096, women p = 0.0030) rates and pooled cohort equations risk score increasing by quartile for Cd (men p < 0.0001, women p = 0.0034) and Hg (men p = 0.0099, women p = 0.0010) with linear trends. Urban population showed a stronger relationship between blood Cd, Pb, and Hg levels and 10-year ASCVD risk score especially among men with multiple regression analysis. CONCLUSION: Blood Cd, Pb, and Hg levels are associated with ASCVD risk. Thus, they should be considered while developing preventive measures for ASCVD.

Parasitic infections based on 320 clinical samples submitted to Hanyang University, Korea (2004-2011).

We analyzed 320 clinical samples of parasitic infections submitted to the Department of Environmental Biology and Medical Parasitology, Hanyang University from January 2004 to June 2011. They consisted of 211 nematode infections, 64 trematode or cestode infections, 32 protozoan infections, and 13 infections with arthropods. The nematode infections included 67 cases of trichuriasis, 62 of anisakiasis (Anisakis sp. and Pseudoterranova decipiens), 40 of enterobiasis, and 24 of ascariasis, as well as other infections including strongyloidiasis, thelaziasis, loiasis, and hookworm infecions. Among the cestode or trematode infections, we observed 27 cases of diphyllobothriasis, 14 of sparganosis, 9 of clonorchiasis, and 5 of paragonimiasis together with a few cases of taeniasis saginata, cysticercosis cellulosae, hymenolepiasis, and echinostomiasis. The protozoan infections included 14 cases of malaria, 4 of cryptosporidiosis, and 3 of trichomoniasis, in addition to infections with Entamoeba histolytica, Entamoeba dispar, Entamoeba coli, Endolimax nana, Giardia lamblia, and Toxoplasma gondii. Among the arthropods, we detected 6 cases of Ixodes sp., 5 of Phthirus pubis, 1 of Sarcoptes scabiei, and 1 of fly larva. The results revealed that trichuriasis, anisakiasis, enterobiasis, and diphyllobothriasis were the most frequently found parasitosis among the clinical samples.

Purification and biocompatibility of fermented hyaluronic acid for its applications to biomaterials.

BACKGROUND: Hyaluronic acid (HA) is of importance due to its diverse applications in pharmaceuticals and medical devices such as dermal filler, adhesion barriers, carrier for cells and bioactive molecules as well as scaffold biomaterials for tissue engineering. Evaluations of purification and biocompatibility of HA are required for its applications to biomaterials. RESULTS: After synthesizing HA by fermentation of streptococcus zooepidemicus for 25 hr, extensively purification of the fermented broth was performed to remove impurities using a filtration process for insoluble components and cells, and diverse adsorbents for soluble impurities. Its in vitro biocompatibility has been evaluated by measurement of cell counting and assay of cell live and dead. 60% yield of white HA powder was obtained, having 15-17 dL/g intrinsic viscosity with a molecular weight of approximately 1,000 kDa. While low molecular weight impurities and insoluble impurities were successfully removed using a ultrafiltration membrane with 50 KDa molecular weight cut, endotoxins, high molecular weight proteins and nucleic acids were removed from the broth by employing adsorbents such as alumina and activated carbons. Alumina showed the best results for the removal of endotoxins, all of the activated carbons were very effective in the removal of high molecular weight proteins and nucleic acids. The purified HA solution showed excellent cell compatibility with no cell damages as observed by both measurement of cell proliferation and observation of cell viability. CONCLUSIONS: We obtained high molecular weight HA with excellent biocompatibility as judged by both measurement of cell proliferation and viability, indicating high possibility of its applications to biomaterials.

Sodium hydrogen sulfide inhibits nicotine and lipopolysaccharide-induced osteoclastic differentiation and reversed osteoblastic differentiation in human periodontal ligament cells.

Although previous studies have demonstrated that hydrogen sulfide (H(2)S) stimulated or inhibited osteoclastic differentiation, little is known about the effects of H(2)S on the differentiation of osteoblasts and osteoclasts. To determine the possible bioactivities of H(2)S on bone metabolism, we investigated the in vitro effects of H(2)S on cytotoxicity, osteoblastic, and osteoclastic differentiation as well as the underlying mechanism in lipopolysaccharide (LPS) and nicotine-stimulated human periodontal ligament cells (hPDLCs). The H(2)S donor, NaHS, protected hPDLCs from nicotine and LPS-induced cytotoxicity and recovered nicotine- and LPS-downregulated osteoblastic differentiation, such as alkaline phosphatase (ALP) activity, mRNA expression of osteoblasts, including ALP, osteopontin (OPN), and osteocalcin (OCN), and mineralized nodule formation. Concomitantly, NaHS inhibited the differentiation of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts in mouse bone marrow cells and blocked nicotine- and LPS-induced osteoclastogenesis regulatory molecules, such as RANKL, OPG, M-CSF, MMP-9, TRAP, and cathepsin K mRNA. NaHS blocked nicotine and LPS-induced activation of p38, ERK, MKP-1, PI3K, PKC, and PKC isoenzymes, and NF-κB. The effects of H(2)S on nicotine- and LPS-induced osteoblastic and osteoclastic differentiation were remarkably reversed by MKP-1 enzyme inhibitor (vanadate) and expression inhibitor (triptolide). Taken together, we report for the first time that H(2)S inhibited cytotoxicity and osteoclastic differentiation and recovered osteoblastic differentiation in a nicotine- and periodontopathogen-stimulated hPDLCs model, which has potential therapeutic value for treatment of periodontal and inflammatory bone diseases.

Application of pulsed buffer gas jets for the signal enhancement of laser-induced breakdown spectroscopy.

We report a new simple method for the signal enhancement of laser-induced breakdown spectroscopy using a pulsed buffer gas jet. The signal is enhanced up to more than 10 fold by using argon gas jets, which are injected through a pulsed nozzle onto the sample area to be analyzed. By synchronizing the buffer gas pulse with the laser pulse and optimizing the spatial arrangements between the gas jet and the sample surface, we have successfully exploited the useful properties of the buffer gas in open atmosphere. The signal-enhancement mechanism in our buffer gas jet has been discussed. Also, applications to various samples (metal, glass, and paper) have been demonstrated.