In vivo study on the repair of rat Achilles tendon injury treated with non-thermal atmospheric-pressure helium microplasma jet.

Non-thermal atmospheric-pressure plasma (NTAPP) has been widely studied for clinical applications, e.g., disinfection, wound healing, cancer therapy, hemostasis, and bone regeneration. It is being revealed that the physical and chemical actions of plasma have enabled these clinical applications. Based on our previous report regarding plasma-stimulated bone regeneration, this study focused on Achilles tendon repair by NTAPP. This is the first study to reveal that exposure to NTAPP can accelerate Achilles tendon repair using a well-established Achilles tendon injury rat model. Histological evaluation using the Stoll's and histological scores showed a significant improvement at 2 and 4 weeks, with type I collagen content being substantial at the early time point of 2 weeks post-surgery. Notably, the replacement of type III collagen with type I collagen occurred more frequently in the plasma-treated groups at the early stage of repair. Tensile strength test results showed that the maximum breaking strength in the plasma-treated group at two weeks was significantly higher than that in the untreated group. Overall, our results indicate that a single event of NTAPP treatment during the surgery can contribute to an early recovery of an injured tendon.

Fracture healing on non-union fracture model promoted by non-thermal atmospheric-pressure plasma.

Non-thermal atmospheric-pressure plasma (NTAPP) is attracting widespread interest for use in medical applications. The tissue repair capacity of NTAPP has been reported in various fields; however, little is known about its effect on fracture healing. Non-union or delayed union after a fracture is a clinical challenge. In this study, we aimed to investigate how NTAPP irradiation promotes fracture healing in a non-union fracture model and its underlying mechanism, in vitro and in vivo. For the in vivo study, we created normal and non-union fracture models in LEW/SsNSlc rats to investigate the effects of NTAPP. To create a fracture, a transverse osteotomy was performed in the middle of the femoral shaft. To induce the non-union fracture model, the periosteum surrounding the fracture site was cauterized after a normal fracture model was created. The normal fracture model showed no significant difference in bone healing between the control and NTAPP-treated groups. The non-union fracture model demonstrated that the NTAPP-treated group showed consistent improvement in fracture healing. Histological and biomechanical assessments confirmed the fracture healing. The in vitro study using pre-osteoblastic MC3T3-E1 cells demonstrated that NTAPP irradiation under specific conditions did not reduce cell proliferation but did enhance osteoblastic differentiation. Overall, these results suggest that NTAPP is a novel approach to the treatment of bone fractures.

Preferential dissolution behaviour of the austenite phase in Fe-27Cr-xC high chromium cast iron.

Preferential dissolution behaviour of the austenite (γ) phase in Fe-27Cr-xC high chromium cast irons (HCCIs) immersed in 0.1 mol dm-3 H2SO4 + 0.05 mol dm-3 HCl was investigated. Potentiodynamic and potentiostatic polarisation revealed that the primary and eutectic γ phases dissolved preferentially at -0.35 and 0.00 VSilver Silverchloride Electrode potential in sat. KCl (SSE), respectively. The immersion of the HCCIs in the solution showed that the dissolution of the primary γ phase dominated for ca. 1 h, while the primary and eutectic γ phases dissolved after ca. 1 h. However, the carbide phases remained undissolved during the dissolution of the γ phases. Furthermore, the corrosion rate of the HCCIs increased with the increasing C content owing to the increase in the contact potential difference values of the γ and carbide phases. The change in electromotive force due to C addition was related to the accelerated corrosion rate of the γ phases.

Effect of carbon addition on the passivity of hypoeutectic high chromium cast irons.

The effect of adding C on the passivity of hypoeutectic high chromium cast iron (HCCI) was investigated in a pH 8.4 boric-borate buffer solution. The microstructure of HCCI is composed of austenite and carbide phases, whose fractions and chemical compositions are influenced by the amount of C added. Electrochemical and surface analyses revealed that the addition of C in the HCCI increased the defect densities in the n-type and p-type semiconductive oxide layers on the austenite and carbide phases, respectively.

Construction of a core collection of native Perilla germplasm collected from South Korea based on SSR markers and morphological characteristics.

The leaves and seed oil of Perilla crop (Perilla frutescens L.) have attracted interest as health foods in East Asia. This crop has been traditionally cultivated and used for a long time as a folk plant, especially in Korea. In our study, the 22 SSR markers and eight morphological traits were used to assess the genetic diversity and population structure, to select a core collection of 400 Perilla accessions conserved in the RDA-Genebank of South Korea. A total of 173 alleles were detected and the number of alleles per locus ranged from 4 to 15 (average = 7.9). Gene diversity and polymorphic information content ranged from 0.138 to 0.868 (average = 0.567) and 0.134 to 0.853 (average = 0.522), respectively. The 400 accessions were not clearly distinguished geographically by STRUCTURE and UPGMA analyses. A core collection (44 accessions) was selected from the entire collection by using PowerCore. The core collection accounted for 11.0% of the entire Perilla collection, including 100% of the number of alleles maintained in the whole collection and with similar or greater Shannon-Weaver and Nei diversity indices than the whole collection. The core collection selected by SSR markers was evenly distributed in three clusters on a scatter plot by eight morphological traits. The first core collection of Perilla accessions was constructed, and it maintained allelic richness. Further modification of the core collection is expected with the continuous addition of new accessions of the two cultivated types of Perilla crop and their weedy types.

In vivo study on the healing of bone defect treated with non-thermal atmospheric pressure gas discharge plasma.

Medical treatment using non-thermal atmospheric pressure plasma (NTAPP) is rapidly gaining recognition. NTAPP is thought to be a new therapeutic method because it could generate highly reactive species in an ambient atmosphere which could be exposed to biological targets (e.g., cells and tissues). If plasma-generated reactive species could stimulate bone regeneration, NTAPP can provide a new treatment opportunity in regenerative medicine. Here, we investigated the impact of NTAPP on bone regeneration using a large bone defect in New Zealand White rabbits and a simple atmospheric pressure plasma (helium microplasma jet). We observed the recovery progress of the large bone defects by X-ray imaging over eight weeks after surgery. The X-ray results showed a clear difference in the occupancy of the new bone of the large bone defect among groups with different plasma treatment times, whereas the new bone occupancy was not substantial in the untreated control group. According to the results of micro-computed tomography analysis at eight weeks, the most successful bone regeneration was achieved using a plasma treatment time of 10 min, wherein the new bone volume was 1.51 times larger than that in the plasma untreated control group. Using H&E and Masson trichrome stains, nucleated cells were uniformly observed, and no inclusion was confirmed, respectively, in the groups of plasma treatment. We concluded the critical large bone defect were filled with new bone. Overall, these results suggest that NTAPP is promising for fracture treatment.

Investigation of the Physicochemical and Biological Stability of the Adalimumab Biosimilar CT-P17.

INTRODUCTION: CT-P17 (Celltrion, Inc., Incheon, Republic of Korea) is a biosimilar of reference adalimumab (Humira®; AbbVie Inc., North Chicago, IL, USA), which has recently received regulatory approval from the European Medicines Agency. METHODS: This analysis was designed to evaluate the stability profile of CT-P17 compared with reference adalimumab and the currently licensed adalimumab biosimilars ABP 501 (Amjevita®/Amgevita®; Amgen Inc., Thousand Oaks, CA, USA) and SB5 (Imraldi®; Biogen Inc., Cambridge, MA, USA) when stored at low temperature (5 °C) or room temperature (25 °C) with 60% relative humidity for up to 28 days. RESULTS: Multiple orthogonal and complementary tests demonstrated that CT-P17 was stable for 28 days under all tested conditions, as well as for protein concentrations tested (50 vs 100 mg/mL), type of delivery device (autoinjector vs prefilled syringe), and manufacturing date (recently manufactured vs aged for 17 months). There were slight differences among products in terms of charge variants, oxidation level, purity, and number of subvisible particles; however, overall, the quality of each product was maintained over 28 days. CONCLUSION: Our data suggest that CT-P17 may be used without any significant loss of stability when stored at 5 °C or 25 °C with 60% relative humidity for up to 28 days, and was not impacted by protein concentration tested and delivery device. Comparative stability data suggest that the appropriate maximum storage period for CT-P17 may be up to 28 days at room temperature with 60% relative humidity.

Genetic Variation of Native Perilla Germplasms Collected from South Korea Using Simple Sequence Repeat (SSR) Markers and Morphological Characteristics.

Using morphological characteristics and simple sequence repeat (SSR) markers, we evaluated the morphological variation and genetic diversity of 200 Perilla accessions collected from the five regions of South Korea and another region. In morphological characteristics analysis, particularly leaf color, stem color, degree of pubescence, and leaf size have been found to help distinguish the morphological features of native Perilla accessions cultivated in South Korea. Twenty SSR primer sets confirmed a total of 137 alleles in the 200 Perilla accessions. The number of alleles per locus ranged from 3 to 13, with an average number of alleles per locus of 6.85. The average genetic diversity (GD) was 0.649, with a range of 0.290-0.828. From analysis of SSR markers, accessions from the Jeolla-do and Gyeongsang-do regions showed comparatively high genetic diversity values compared with those from other regions in South Korea. In the unweighted pair group method with arithmetic mean (UPGMA) analysis, the 200 Perilla accessions were found to cluster into three main groups and an outgroup with 42% genetic similarity, and did not show a clear geographic structure from the five regions of South Korea. Therefore, it is believed that landrace Perilla seeds are frequently exchanged by farmers through various routes between the five regions of South Korea. The results of this study are expected to provide interesting information on the conservation of these genetic resources and selection of useful resources for the development of varieties for seeds and leafy vegetables of cultivated Perilla frutescens var. frutescens in South Korea.

Discovering latent themes in traffic fatal crash narratives using text mining analytics and network topology.

The proliferation of digital textual archives in the transportation safety domain makes it imperative for the inventions of efficient ways of extracting information from the textual data sources. The present study aims at utilizing crash narratives complemented by crash metadata to discern the prevalence and co-occurrence of themes that contribute to crash incidents. Ten years (2009-2018) of Michigan traffic fatal crash narratives were used as a case study. The structural topic modeling (STM) and network topology analysis were used to generate and examine the prevalence and interaction of themes from the crash narratives that were mainly categorized into pre-crash events, crash locations and involved parties in the traffic crashes. The main advantage of the STM over the other topic modeling approaches is that it allows the researchers to discover themes from documents and estimate how the topic relates to the document metadata. Topics with the highest prevalence for the angle, head-on, rear-end, sideswipe and single motor vehicle crashes were crash at stop-sign, crossing the centerline, unable to stop, lane change maneuver and run-off-road crash, respectively. Eigenvector centrality measure in network topology showed that event-related topics were consistently central in articulating the crash occurrence. The centrality and association between topics varied across crash types. The efficacy of generated topics in classifying crashes by type was tested using a machine learning algorithm, Random Forest. The classification accuracy in the held-out sample ranged between 89.3 % for sideswipe crashes to 99.2 % for single motor vehicle crashes. High classification accuracy suggests that automation of crash typing and consistency checks can be accomplished effectively by using extracted latent themes from the crash narratives.

Assessment of genetic diversity and population structure among a collection of Korean Perilla germplasms based on SSR markers.

BACKGROUND: Information on the genetic variation of genetic resource collections is very important for both the conservation and utilization of crop germplasms in genebanks. Var. frutescens of Perilla crop is extensively cultivated in South Korea as both an oil crop and a vegetable crop. OBJECTIVES: We used SSR markers to evaluate the genetic diversity, genetic relationships, and population structure of 155 accessions of var. frutescens that have been selected as genetic resources for the development of leaf vegetable cultivars and preserved in the RDA-Genebank collection from South Korea. METHODS: A total of 155 accessions of var. frutescens of Perilla crop collected in South Korea were obtained from the RDA-Genebank of the Republic of Korea. We selected 20 SSR markers representing the polymorphism of and adequately amplifying all the Perilla accessions. RESULTS: The average GD and PIC values were 0.642 and 0.592, respectively, with ranges of 0.244-0.935 and 0.232- 0.931. The genetic variability in the southern region of South Korea was higher than that in the central region. The clustering patterns were not clearly distinguished between the accessions of var. frutescens from the central and southern regions of South Korea. CONCLUSION: These results regarding the genetic diversity and population structure of the 155 accessions of var. frutescens of South Korea provide useful information for understanding the genetic variability of this crop and selecting and managing core germplasm sets in the RDA-Genebank of the Republic of Korea.

Expression of Inhibitory Receptors on T and NK Cells Defines Immunological Phenotypes of HCV Patients with Advanced Liver Fibrosis.

Chronic HCV can result in advanced liver disease, including cirrhosis. Patients with advanced fibrosis experience poor clinical outcomes and increased risk for hepatocellular carcinoma (HCC). These outcomes are, in part, a consequence of immune dysfunction. Increased inhibitory receptor and Galectin-9 (GAL-9) expression is a possible mechanism promoting lymphocyte dysfunction. In this study, we measured the expression of inhibitory receptors and GAL-9 on T/NK cells of patients with chronic HCV with no to moderate fibrosis (F0-F2) and advanced fibrosis (F3-F4). To analyze their co-expression, we employed t-SNE analysis. Notably, we found that F3-F4 patients had higher frequencies of >3 inhibitory receptor co-expression on NK cells. Moreover, F3-F4 patients manifest a higher frequency of NK cells co-expressing TIGIT and TIM-3, and CD4/NK cells co-expressing LAG-3 and GAL-9. In conclusion, we identified phenotypes of immune dysregulation that could explain the increased susceptibility to infection and HCC in patients with chronic HCV with advanced fibrosis.

Development of systemwide pedestrian safety performance function using stratified random sampling and a proxy measure of pedestrian exposure.

The lack of pedestrian counts at a systemwide level prompts the need to find other innovative ways of assessing pedestrian traffic crash risks using proxy measures of exposure. This study aims to formulate the methodology for developing pedestrian safety performance functions (SPF) using the proxy measure of pedestrian exposure and stratified random sampling. The case study was all urban intersections in Michigan State that comprise of collector and arterial roads. The stratified random sampling strategy was deployed to select the sample which is representative of all urban intersections in the state of Michigan. Factor analysis was used to develop a proxy measure of pedestrian exposure at urban intersections using a walkability measure (walk score), among other factors. The performances of various count models were compared using the goodness of fit measures based on the Akaike's Information Criterion (AIC), Bayesian Information Criterion (BIC), and Vuong test. The final pedestrian SPFs was formulated using the Zero-Inflated Poisson (ZIP) model with AADT at a major approach, AADT at the minor approach, and a proxy measure of pedestrian exposure. The proposed methodology in this study can benefit transportation agencies that have embarked on systemwide planning of pedestrian facilities to improve the safety of pedestrians but lack systemwide analytical tools and pedestrian counts to make data-driven decisions.

Assessing the safety impacts of raising the speed limit on Michigan freeways using the multilevel mixed-effects negative binomial model.

Objectives: The main objective of this study was to assess the change in crash patterns associated with speed limit changes from 55 mph to 70 mph that occurred on some of Michigan freeway segments between year 2005 and year 2010.Method: Many of the statistical methods used in the past to evaluate the safety impacts of raising the speed limit on freeways lack the ability to address one or more critical issues inherent in count data, such as omitted-sample bias, over-dispersion and regression-to-the-mean bias. This study used multilevel mixed-effects negative binomial regression to address these limitations, with an additional advantage of controlling for intra-cluster correlation of crashes on each freeway corridor and segments nested in the same corridor. Changes in the crash patterns between the year 2000 and year 2015 were investigated on test sites that had a change of speed limit from 55 mph to 70 mph, relative to control sites where the speed limit was maintained at 55 mph.Results: The inclusion of random effects improved the model's ability in explaining observed crash variations on the selected freeway segments, as indicated by test statistics such as the log-likelihood ratio test, the Akaike information criterion and Bayesian information criterion (BIC)values. Further, random effects improved the significant speed limit change fixed effects during model calibration. The final mixed-effects model indicated a significant increase in fatal and injury crashes (FI), total crashes (KABCO) and road departure crashes by 11.9 percent, 21.0 percent and 23.3 percent, respectively, on freeway segments where the speed limit was raised from 55 mph to 70 mph. The increase in road departure crashes was more pronounced on curved freeway segments with the raised speed limit compared to straight segments with no speed limit changes.Conclusions: The 15 mph increase in the speed limit on Michigan freeways had a significant association with the increase in fatal and injury crashes, total crashes and road departure crashes. The elevated crash risks associated with the speed limit increase suggest that further studies are needed to understand changes in drivers' behaviors following a speed limit increase.

How does an increased freeway speed limit influence the frequency of crashes on adjacent roads?

Urban arterial roads carry the most traffic on urban road networks and experience the highest percentage of crashes in urban areas. Safety on urban arterials that are adjacent to a higher speed freeway may be impacted by speed spillover or adaptation. The objective of this study was to determine the effects of raising freeway speed limits on the frequency of crashes on urban arterial roads adjacent to freeways (spillover effects). Crash data within Michigan were collected on 1393 urban arterial road segments before and after freeway speed limits were altered. Before-and-after data was collected simultaneously on 1470 comparison segments of urban arterial where speed limits did not change to control for the regression-to-the-mean bias. The mixed effects negative binomial (MENB) regression model was developed to analyze crash frequency on urban arterials. The results indicate that raising speed limits of freeways by as little as five miles per hour had a likelihood of increasing crash frequency on adjacent arterial roads by as much as 13.9 percent. To investigate if the safety impact of speed spillover changes with the distance from the freeway, influence areas (0-1 mile, 1-2 mile, and 2-3 mile) were used. The findings of this study provide insights into the effects of speed spillover on crash occurrences, and it demonstrates that increasing freeway speed limit has a negative influence on driver compliance with the speed limit on adjacent arterial roads. Correspondingly, the influence of freeway speed on drivers' speeding behavior on adjacent urban arterials fades away with the distance from the freeway.

How membrane lipids influence plasma delivery of reactive oxygen species into cells and subsequent DNA damage: an experimental and computational study.

The mechanisms of plasma in medicine are broadly attributed to plasma-derived reactive oxygen and nitrogen species (RONS). In order to exert any intracellular effects, these plasma-derived RONS must first traverse a major barrier in the cell membrane. The cell membrane lipid composition, and thereby the magnitude of this barrier, is highly variable between cells depending on type and state (e.g. it is widely accepted that healthy and cancerous cells have different membrane lipid compositions). In this study, we investigate how plasma-derived RONS interactions with lipid membrane components can potentially be exploited in the future for treatment of diseases. We couple phospholipid vesicle experiments, used as simple cell models, with molecular dynamics (MD) simulations of the lipid membrane to provide new insights into how the interplay between phospholipids and cholesterol may influence the response of healthy and diseased cell membranes to plasma-derived RONS. We focus on the (i) lipid tail saturation degree, (ii) lipid head group type, and (iii) membrane cholesterol fraction. Using encapsulated molecular probes, we study the influence of the above membrane components on the ingress of RONS into the vesicles, and subsequent DNA damage. Our results indicate that all of the above membrane components can enhance or suppress RONS uptake, depending on their relative concentration within the membrane. Further, we show that higher RONS uptake into the vesicles does not always correlate with increased DNA damage, which is attributed to ROS reactivity and lifetime. The MD simulations indicate the multifactorial chemical and physical processes at play, including (i) lipid oxidation, (ii) lipid packing, and (iii) lipid rafts formation. The methods and findings presented here provide a platform of knowledge that could be leveraged in the development of therapies relying on the action of plasma, in which the cell membrane and oxidative stress response in cells is targeted.

In-Use Physicochemical and Biological Stability of the Trastuzumab Biosimilar CT-P6 Upon Preparation for Intravenous Infusion.

BACKGROUND: CT-P6 is a biosimilar of trastuzumab, a monoclonal antibody targeting human epidermal growth factor 2 (HER2), that is used in the treatment of breast and gastric cancers. OBJECTIVE: The aim of this study was to evaluate the in-use physicochemical and biological stability of CT-P6 following preparation for intravenous (IV) infusion. METHODS: One batch of CT-P6 within the final month of its 48-month shelf life was used to simulate sub-optimal administration conditions. CT-P6 dilutions of 0.4, 1.0, and 4.0 mg/mL, representative of actual use scenarios, were prepared in 0.9% saline solution in either polypropylene (PP) or polyvinylchloride (PVC) infusion bags. Following refrigeration at 2-8 °C for 1 month, samples were incubated at room temperature for 24 h. Physicochemical and biological stability were evaluated according to presence of sub-visible particles, pH, proportion of molecular weight variants, oxidation level of methionine residues 107, 255/256 and 432/433, and binding affinity to the Fc neonatal receptor and HER2. RESULTS: Analyses of CT-P6 preparations at all concentrations tested and in both PP and PVC infusion bags revealed no changes in sub-visible particles, pH, molecular weight variants, oxidation, or potency after 1 month at 2-8 °C followed by exposure to room temperature for 24 h. CONCLUSION: These analyses demonstrate the extended stability, after refrigerated storage for 1 month followed by 24-h exposure to room temperature, of CT-P6 under the dilution conditions required for IV infusion. This stability was sustained for all dilution factors and both infusion bag materials tested.

A comparison of safety benefits of pedestrian countdown signals with and without pushbuttons in Michigan.

OBJECTIVE: This study evaluated the safety impacts of pedestrian countdown signals (PCSs) with and without pushbuttons based on pedestrian crashes and pedestrian injuries in Michigan. METHODOLOGY: This study used 10 years of intersection data-5 years before PCSs were installed and 5 years after they were installed-along with a comparison group, to evaluate the crash impacts of PCSs; at 107 intersections the PCS had a pushbutton and at 96 it did not. At these intersections, and at their comparison sites (where no PCS was installed), crash data (from 2004 to 2016) were examined, along with traffic and geometric characteristics, population, education, and poverty level data. RESULTS: Intersections where PCSs with pushbuttons have been installed showed a 29% reduction in total pedestrian crashes and a 30% reduction in fatal/injury pedestrian crashes. Further, when considering only pedestrians age 65 and below, these respective reductions are 33 and 35%. Intersections with PCSs but without pushbuttons did not show any significant change in any type of pedestrian crash. CONCLUSIONS: Although the Manual on Uniform Traffic Control Devices (Federal Highway Administration [FHWA] 2009 ) requires the use of PCSs at new traffic signal installations, this study suggests a safety benefit of installing PCSs with pushbutton at signals where a PCS without a pushbutton is present.

Tracking the Penetration of Plasma Reactive Species in Tissue Models.

Electrically generated cold atmospheric plasma is being intensively researched for novel applications in biology and medicine. Significant attention is being given to reactive oxygen and nitrogen species (RONS), initially generated upon plasma-air interactions, and subsequently delivered to biological systems. Effects of plasma exposure are observed to millimeter depths within tissue. However, the exact nature of the initial plasma-tissue interactions remains unknown, including RONS speciation and delivery depth, or how plasma-derived RONS intervene in biological processes. Herein, we focus on current research using tissue and cell models to learn more about the plasma delivery of RONS into biological environments. We argue that this research is vital in underpinning the knowledge required to realize the full potential of plasma in biology and medicine.

Oxygen-radical pretreatment promotes cellulose degradation by cellulolytic enzymes.

BACKGROUND: The efficiency of cellulolytic enzymes is important in industrial biorefinery processes, including biofuel production. Chemical methods, such as alkali pretreatment, have been extensively studied and demonstrated as effective for breaking recalcitrant lignocellulose structures. However, these methods have a detrimental effect on the environment. In addition, utilization of these chemicals requires alkali- or acid-resistant equipment and a neutralization step. RESULTS: Here, a radical generator based on non-thermal atmospheric pressure plasma technology was developed and tested to determine whether oxygen-radical pretreatment enhances cellulolytic activity. Our results showed that the viscosity of carboxymethyl cellulose (CMC) solutions was reduced in a time-dependent manner by oxygen-radical pretreatment using the radical generator. Compared with non-pretreated CMC, oxygen-radical pretreatment of CMC significantly increased the production of reducing sugars in culture supernatant containing various cellulases from Phanerochaete chrysosporium. The production of reducing sugar from oxygen-radical-pretreated CMC by commercially available cellobiohydrolases I and II was 1.7- and 1.6-fold higher, respectively, than those from non-pretreated and oxygen-gas-pretreated CMC. Moreover, the amount of reducing sugar from oxygen-radical-pretreated wheat straw was 1.8-fold larger than those from non-pretreated and oxygen-gas-pretreated wheat straw. CONCLUSIONS: Oxygen-radical pretreatment of CMC and wheat straw enhanced the degradation of cellulose by reducing- and non-reducing-end cellulases in the supernatant of a culture of the white-rot fungus P. chrysosporium. These findings indicated that oxygen-radical pretreatment of plant biomass offers great promise for improvements in lignocellulose-deconstruction processes.