Optimizing conditions to electroporate Rothia mucilaginosa

Rothia mucilaginosa (Rm) is a gram-positive bacterium residing in the oral cavity. Recent studies in our laboratory have shown that this microorganism is able to cleave gluten, including immunogenic domains implicated in celiac disease. This can be beneficial to patients with celiac disease because exploitation of Rm can provide a novel mode of treatment. The enzymes responsible for this cleavage are as yet unknown. The purpose of this study was to optimize the transformation efficiencies of Rm cells through electroporation, with the ultimate goal to create knock-out mutants for enzyme activity. We have determined various aspects of Rm cells relevant for this project: (1) the growth curve characteristics of Rm; (2) the presence of endogenous restriction enzyme activities; and (3) the conditions facilitating Rm electroporation by varying electroporation voltages. Furthermore, electroporation and transformation of the plasmid pUC18 was conducted in Escherichia coli. The growth curve of Rm cells in BHI growth medium incubated at 37°C while shaking showed a doubling time of approximately 3 hours in the logarithmic growth phase. Using a cell sonicate of Rm cells incubated with Lambda DNA and four different restriction enzyme buffers, we found that there were no apparent endogenous restriction enzyme activities detectable. For the electroporation experiments, we used previously published protocols for the bacterium Staphylococcus aureus, as a standard condition to electroporate Rm cells. Those studies have shown that changing electrical parameters during the electroporation would yield a high efficiency rate of gram-positive bacterial transformation (Lofblom et. al., 2006; Metzler et. al., 1992). Therefore in our study, we increased the field strengths (kV*cm-1) to electroporate Rm cells. Rm cells could not be successfully transformed, and we observed that field strengths exceeding 18 kV*cm-1 destroyed Rm cells. On the other hand, the transformation of E. coli with pUC18 was successful. Our studies have laid the groundwork for investigating the transformation of Rm cells, and future studies can use the results obtained to further investigate ways to optimize transformation of Rm cells for potential utility in celiac patients

A zinc finger protein array for the visual detection of specific DNA sequences for diagnostic applications.

The visual detection of specific double-stranded DNA sequences possesses great potential for the development of diagnostics. Zinc finger domains provide a powerful scaffold for creating custom DNA-binding proteins that recognize specific DNA sequences. We previously demonstrated sequence-enabled reassembly of TEM-1 β-lactamase (SEER-LAC), a system consisting of two inactive fragments of β-lactamase each linked to engineered zinc finger proteins (ZFPs). Here the SEER-LAC system was applied to develop ZFP arrays that function as simple devices to identify bacterial double-stranded DNA sequences. The ZFP arrays provided a quantitative assay with a detection limit of 50 fmol of target DNA. The method could distinguish target DNA from non-target DNA within 5 min. The ZFP arrays provided sufficient sensitivity and high specificity to recognize specific DNA sequences. These results suggest that ZFP arrays have the potential to be developed into a simple and rapid point-of-care (POC) diagnostic for the multiplexed detection of pathogens

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Mitigating Health Inequalities of Socially Vulnerable in South Korea: Role for Social Work

The purpose of this study is to develop a social work education strategy for mitigating health inequalities among the socially vulnerable. The limitations of the healthcare approach to health promotion and health inequality, which we examined through the health belief model describing an individual’s health behavior, emphasize the prevention of diseases that have not yet occurred or are likely to occur, healthcare management for them, and the functions and role of the community in the process, based on the individual’s spontaneity to practice health behaviors. Therefore, to compensate for these limitations, it is deemed necessary to add healthcare curriculum, such as public health, social epidemiology, and etiology, to the existing curriculum of social work based on an in-depth understanding of social vulnerability and social environment as well as the importance of preventing and managing diseases

Estimating Tobins Q for Listed Firms in Korea (1980-2005): Comparing Alternative Approaches and an Experiment with Investment Functions

Tobins Q is the most common measurement of firm value and performance. However, estimating Tobins Q accurately is not easy. Researchers have used book values of debts or assets rather than the market values. We estimate Tobins Q for listed firms in Korea from 1980 to 2005 based on replacement costs of assets as well as market values of debts and common and preferred stocks. We compare the estimates using the modified annual average depreciation rates and economic depreciation rates. In sum, we present and compare four alternative series of Tobins Q measures. We then estimate investment functions with alternative Tobins Q values as regressors to compare the reliability of alternative estimates. We find that the simple measure of using book values of both debts and assets is the most unreliable

Epidemiology, burden and clinical spectrum of cluster headache: a global update

Disability; Economic and job-related burdens; Quality of lifeDiscapacidad; Cargas económicas y laborales; Calidad de vidaDiscapacitat; Càrregues econòmiques i laborals; Qualitat de vidaBackground This narrative review aims to broaden our understanding of the epidemiology, burden and clinical spectrum of cluster headache based on updated findings with a global perspective. Methods We conducted a literature search on the following topics: (a) epidemiology; (b) burden: quality of life, disability, economic burden, job-related burden and suicidality; and (c) clinical spectrum: male predominance and its changes, age, pre-cluster and pre-attack symptoms, aura, post-drome, attack characteristics (location, severity, duration and associated symptoms), bout characteristics (attack frequency, bout duration and bout frequency), circadian and seasonal rhythmicity and disease course. Results New large-scale population-based reports have suggested a lower prevalence than previous estimations. The impact of cluster headache creates a significant burden in terms of the quality of life, disability, economic and job-related burdens and suicidality. Several studies have reported decreasing male-to-female ratios and a wide age range at disease onset. The non-headache phases of cluster headache, including pre-cluster, pre-attack and postictal symptoms, have recently been revisited. The latest data regarding attack characteristics, bout characteristics, and circadian and seasonal rhythmicity from different countries have shown variability among bouts, attacks, individuals and ethnicities. Studies on the disease course of cluster headache have shown typical characteristics of attacks or bouts that decrease with time. Conclusions Cluster headache may be more than a “trigeminal autonomic headache” because it involves complex central nervous system phenomena. The spectrum of attacks and bouts is wider than previously recognised. Cluster headache is a dynamic disorder that evolves or regresses over time.This study was supported by the New Faculty Start-up Fund of Seoul National University and National Research Foundation of Korea (NRF) grant funded by the Korean Government (MSIP; No. 2020R1A2B5B01001826)

Rapid Hepatobiliary Excretion of Micelle-Encapsulated/Radiolabeled Upconverting Nanoparticles as an Integrated Form

In the field of nanomedicine, long term accumulation of nanoparticles (NPs) in the mononuclear phagocyte system (MPS) such as liver is the major hurdle in clinical translation. On the other hand, NPs could be excreted via hepatobiliary excretion pathway without overt tissue toxicity. Therefore, it is critical to develop NPs that show favorable excretion property. Herein, we demonstrated that micelle encapsulated Cu-64-labeled upconverting nanoparticles (micelle encapsulated Cu-64-NOTA-UCNPs) showed substantial hepatobiliary excretion by in vivo positron emission tomography (PET) and also upconversion luminescence imaging (ULI). Ex vivo biodistribution study reinforced the imaging results by showing clearance of 84% of initial hepatic uptake in 72 hours. Hepatobiliary excretion of the UCNPs was also verified by transmission electron microscopy (TEM) examination. Micelle encapsulated Cu-64-NOTA-UCNPs could be an optimal bimodal imaging agent owing to quantifiability of Cu-64, ability of in vivo/ex vivo ULI and good hepatobiliary excretion property.