Biosafety Evaluation Research Articles

A high-dosage microneedle for programmable lidocaine delivery and enhanced local long-lasting analgesia

Considering the staggering global prevalence of local pain affecting hundreds of million individuals, it is of great significance to develop advanced dosage forms or delivery systems for analgesic therapy to fulfill clinical applicability. In this study, a hydrogel microneedles (MNs) system made out of gelatin-methacryloyl (GelMA) was designed to deliver lidocaine hydrochloride (LiH) in a sustained manner, and the drug loading capacity of the GelMA MNs was increased considerably by using the backing layer reservoir. The in vitro and in vivo tests showed that the fabricated GelMA MNs are strong enough for reliable skin application, and achieve high drug delivery efficiency as compared with the commercial lidocaine patches. The Spared-nerve injury (SNI) model of rats was also prepared to test behavioral pain sensitivity in response to mechanical stimuli, which proved that the LiH/GelMA MNs can enhance and prolong the anesthetic effect of LiH. In addition, with biosafety evaluation in rats, the MNs treated site restored to normal appearance within several hours of application and no dermatosis-related side effects or behavior disorders were observed during the experiment. Together these results indicated that the use of GelMA MNs for transdermal delivery of LiH is an effective, safe and simple treatment method to provide a better choice for local long-lasting analgesia.

Cell‐Derived Biomimetic 2D Nanoparticles to Improve Cell‐Specific Targeting and Tissue Penetration for Enhanced Magnetic Resonance Imaging

AbstractA critical challenge of nanoparticle‐based imaging diagnosis of cancers is limited accumulation of nanoparticles at the biological target. Here, a cancer cell derived membrane coated 2D layered double hydroxides nanoparticle (CM‐PEG/MnLDH) has been developed to realize both cell‐specific targeting and deep tissue penetration for enhanced magnetic resonance imaging (MRI) of tumors. Using 4T1 breast cancer cells as a model, the biomimetic CM‐PEG/MnLDH exhibited excellent cell‐specific targeting ability with significantly higher accumulation in homologous cancer cells than the nonhomologous cells. Using 3D tumor spheroid cultures, the CM‐PEG/MnLDH demonstrated highly selective binding and deep tissue penetration ability. In vivo imaging revealed that intravenous injection of CM‐PEG/MnLDH provides clear MRI of tumor tissues in a 4T1 mouse model. Particularly, the CM‐PEG/MnLDH showed enhanced imaging performance in the central region of tumors, owing to the cell homing and tissue penetration ability of the biomimetic nanoparticle. In vivo biosafety evaluation of the CM‐PEG/MnLDH showed a high degree of biocompatibility, signifying its suitability for further clinical applications. The biomimetic CM‐PEG/MnLDH nanoparticle featured with cell‐specific targeting ability and deep tissue penetration is a promising MRI contrast agent for precise cancer diagnosis.

Radiolytic synthesis and characterization of selenium nanoparticles: comparative biosafety evaluation with selenite and ionizing radiation.

The goal of this work is use a green chemistry route to synthesize selenium nanoparticles (SeNPs) that do not trigger oxidative stress, typical of metallic, oxide metallic and carbonaceous nanostructures, and supply the same beneficial effects as selenium nanostructures. SeNPs were synthesized using a radiolytic method involving irradiating a solution containing sodium selenite (Se4+) as the precursor in 1% Yeast extract, 2% Peptone, 2% Glucose (YPG) liquid medium with gamma-rays (60Cobalt). The method did not employ any hazardous reducing agents. Saccharomyces cerevisiae cells were incubated with 1mM SeNPs for 24h and/or then challenged with 400Gy of ionizing radiation were assessed for viability and biomarkers of oxidative stress: lipid peroxidation, protein carbonylation, free radical generation, and total sulfhydryl content. Spherical SeNPs with variable diameters (from 100 to 200nm) were formed after reactions of sodium selenite with hydrated electrons (eaq-) and hydrogen radicals (H·). Subsequent structural characterizations indicated an amorphous structure composed of elemental selenium (Se0). Compared to 1mM selenite, SeNPs were considered safe and less toxic to Saccharomyces cerevisiae cells as did not elicit significant modifications in cell viability or oxidative stress parameters except for increased protein carbonylation. Furthermore, SeNPs treatment afforded some protection against ionizing radiation exposure. SeNPs produced using green chemistry attenuated the reactive oxygen species generation after in vitro ionizing radiation exposure opens up tremendous possibilities for radiosensitizer development.

Biosafety evaluation of dual-responsive neutrobots.

The toxicity effects of paclitaxel (PTX)-loaded magnetic neutrophil-hybrid swimming microrobots ("neutrobots") in vivo were assessed after intravenous administration to mice. The mice after 72 hours exhibited minimal immunotoxicity and liver and kidney toxicity at an administration dose of 3 × 106 PTX-loaded neutrobots. The minor toxicity of drug-loaded neutrobots holds considerable promise for biomedical applications.

Optimization of Chlorella-Biomass Production Using Domestic and Restaurant Waste Water as a Potential Feedstock

The quest to invent a cost-effective media for commercial cultivation of microalgae biomass has remained a cause for motivation for quite some time now. In this study domestic and restaurant wastewater were obtained from Choba, Rivers State Nigeria. The biosafety of the wastewater was ascertained and prepared as a media for the cultivation of microalgae. The samples were prepared in 180:20, 160:40, 140:60, 120:80, and 100:100 ratios using pond water and mixture of domestic and restaurant wastewater respectively. The blooming process was observed for seven days and biomass was monitored by cell optical density and dry weight. The result revealed that biosafety evaluation saw a reduction from 120cfu/ml to 0cfu/ml on the third day. The optimal wavelength selected for growth monitoring was 620nm while growth media ratio selected was 120:80 for restaurant:domestic wastewater. The optimization revealed pH 6.0, temperature 300C, salinity 10ppm and photoperiod 12:12 day:night as optimal condition. Domestic wastewater can be a veritable medium for cultivation of Chlorella sp as a means of integrated waste management; the Chlorella biomass can be used as a feedstock for various biotechnological applications such as source of biochemical, nutraceuticals and for use in biofuel generation.

Non-enzymatic Ex-vitro Antioxidant, Biosafety, Analgesic and Anti-inflammatory Evaluation of Aqueous Polyherbal (ELNA) Extract in Mice.

The polyherbal formulation known as ELNA consists of Moringa oleifera, Crateva religiosa and Curcuma longa. It ethnomedicinal properties include; analgesic, inflammation, cardiac diseases, bacterial infection and immune suppressed diseases. The aim of this study was to evaluate the ex-vitro non-enzymatic anti-oxidant, biosafety, analgesic and anti-inflammatory effect of aqueous ELNA polyherbal extract. Ex-vitro anti-oxidant study was investigated in ELNA extract using standard radical scavenging protocol. Biosafety effect of ELNA extract was understudied with the aid of Locke method. Acetic acid-induced peripheral pain, hot plate-induced central pain and egg albumin-induced inflammatory models were investigated. Results from this study exhibited the scavenging effect of ELNA extract against 1, 1-diphenyl-2-picryhydroxyl with significant increase in concentration dependent manner. Acute toxicity study of the aqueous extract of ELNA elicited less or no toxic effect, with no mortality and side effect above LD50>5000 mg/kg body weight. Aqueous ELNA extract displayed no damaging effect of the sub-chronic toxicity study administered for 28 days, exhibited no significant difference (p= 0.05) in the organ and body weight. No significant different in the haematological parameters excluding 400 mg/kg with slight significant (p<0.05) increase in platelet count. Graded doses of the extract at 400, 800 and 1200 mg/kg body weight exhibited significant (p<0.05) reduction in central and peripheral pain. Significant reduction in paw edema volume. Specifically at higher doses, the extract showed an effective activities against the selected study. In conclusion, this study validated the ethnomedicianl evidence of ELNA aqueous extract of the formulation.

Sodium alginate-based coating incorporated with Anemone vitifolia Buch.-Ham. extract: Application in peach preservation

The extract from Anemone vitifolia Buch.-Ham. was used for postharvest peach preservation for the first time. Different extracts of A. vitifolia were obtained by liquid-liquid extraction based on the gradient polarity, which were analyzed and characterized through HPLC, TLC NMR data and related literatures. Seven saprophytic fungi were isolated and identified from rotting peaches, thus clarifying the main saprophytic fungi. The extracts have certain antifungal effects on above identified fungi. It was found that n-butanol extract/sodium alginate coating of A. vitifolia, which mainly contains triterpenoid saponins and sterol saponins has a good effect on preserving the quality and repelling insects of peach fruit. Bio-safety evaluation result shows that To+SA, PE+SA, EA+SA, nB+SA and W+SA coating are relatively safe. Taken together, these results illuminated that A. vitifolia extract could be used as a promising alternative of peach natural preservatives.

Validation and Application of a Benchtop Cell Sorter in a Biosafety Level 3 Containment Setting.

Introduction: Fluorescent-activated cell sorting (FACS) is often the most appropriate technique to obtain pure populations of a cell type of interest for downstream analysis. However, aerosol droplets can be generated during the sort, which poses a biosafety risk when working with samples containing risk group 3 pathogens such as Francisella tularensis, Mycobacterium tuberculosis, Yersinia pestis, and severe acute respiratory syndrome coronavirus 2. For many researchers, placing the equipment required for FACS at biosafety level 3 (BSL-3) is often not possible due to expense, space, or expertise available.Methods: We performed aerosol testing as part of the biosafety evaluation of the MACSQuant Tyto, a completely closed, cartridge-based cell sorter. We also established quality control procedures to routinely evaluate instrument performance.Results: The MACSQuant Tyto does not produce aerosols as part of the sort procedure.Discussion: These data serve as guidance for other facilities with containment laboratories wishing to use the MACSQuant Tyto for cell sorting. Potential users should consult with their Institutional Biosafety Committees to perform in-house risk assessments of this equipment.Conclusion: The MACSQuant Tyto can safely be used on the benchtop to sort samples at BSL-3.

Evaluating the effects of Bt protein Vip3Aa on immature Propylea japonica (Thunberg) (Coleoptera: Coccinellidae)

BackgroundGenetically modified (GM) crops have become a common tool for managing insect pests of economic importance. However, before their commercialization, bio-safety evaluation on non-target arthropods such as pollinators and biological control agents is a pre-requisite to save services rendered by them in agro-ecosystem.ResultsIn the present experiment, the potential adverse effects of Bt protein (Vip3Aa) were studied on the life table parameters (pupation and emergence rates, 7-day larval weights, larval development time, and adult weight) of a predatory beetle, Propylea japonica (Thunberg) (Coleoptera: Coccinellidae) in the laboratory. P. japonica was exposed to Bt protein incorporated in synthetic diet at the rate of 200 µg/g of fresh weight of diet. Temporal stability, bioactivity and the uptake of insecticidal protein were confirmed by enzyme-linked immunosorbent assay and a sensitive insect bioassay. The results indicated that pupation and adult emergence rates did not differ significantly from that of negative control (pure synthetic diet). Similarly, the fresh weight of male and female adults emerged in Vip3Aa treatment (2.87 mg and 3.86 mg, respectively) did not vary significantly from that of the negative control treatment (2.66 mg, 3.75 mg). However, the time taken by the larvae to complete their development (12.70 days) was significantly extended than that of the negative control (10.21 days). In contrast, the tested life-table parameters of P. japonica were adversely affected when fed on synthetic diet containing boric acid (positive control) indicating experimental fitness to detect adverse effects.ConclusionsOn the basis of the results, it is suggested that GM crops expressing Bt protein (Vip3Aa) are safe for non-target insects such as P. japonica and are compatible with other biological programs for the integrated management of insect pests.

Evaluation of Biosafety, Antiobesity, and Endothelial Cells Proliferation Potential of Basil Seed Extract Loaded Organic Solid Lipid Nanoparticle.

The present study aimed to synthesize solid lipid nanoparticles to enhance liposome-assisted intracellular uptake of basil seed active components in adipocytes and vascular smooth muscle cells to attain increased bioavailability. To obtain solid lipid nanoparticle (SLNp), the water phase containing basil seed extract (BSE) was encapsulated with lipid matrix containing chia seed phospholipids using homogenization and cold ultra-sonication method. The physicochemical characterization of BSE loaded solid lipid nanoparticles (BSE-SLNp) has been analyzed using Zetasizer, FT-IR, and TEM. The BSE-SLNp showed an average diameter of 20–110 nm on the day of preparation and it remains the same after 60 days of storage. The cytotoxicity assay confirmed that the BSE-SLNp did not produce toxicity in hMSCs, preadipocytes, or human umbilical vein endothelial cells (HUVECs) until the tested higher dose up to 64 μg/ml. During effective dose determination, 4 μg/ml of BSE-SLNp confirmed non-toxic and enhanced metabolic function in hMSCs, preadipocytes, and HUVECs. Biosafety assay confirmed normal nuclear morphology in PI staining and high mitochondrial membrane potential in JC-1 assay within 48 h in hMSCs. The maturing adipocyte treated with 4 μg/ml of BSE-SLNp significantly increased the mitochondrial efficiency and fatty acid beta-oxidation (PPARγC1α, UCP-1, and PRDM-16) related gene expression levels. Oxidative stress induced HUVECs treated with 4 μg/ml of BSE-SLNp potentially enhanced antioxidant capacity, cell growth, and microtubule development within 48 h H2O2 induced oxidative stressed HUVECs have shown 39.8% viable cells, but treatment with BSE-SLNp has shown 99% of viable cells within 48 h confirmed by Annexin-V assay. In addition, mitochondrial membrane potential (Δψm) increased to 89.4% confirmed by JC-1 assay. The observed DNA integrity, cell viability was confirmed by increased antioxidant and tumor suppressor-related gene expression levels. VEGF expression has been significantly increased and pro-inflammation-related mRNA levels were decreased in BSE-SLNp treated cells. In conclusion, enhanced adipocyte fatty acid oxidation is directly associated with decreased adipocytokine secretion which arrests obesity-associated comorbidities. In addition, suppressing vascular cell oxidative stress and metabolic inflammation supports vascular cell proliferation and arrests ageing-related vascular diseases.

Surface charge-dependent mitochondrial response to similar intracellular nanoparticle contents at sublethal dosages

BackgroundConsidering the inevitability for humans to be frequently exposed to nanoparticles (NPs), understanding the biosafety of NPs is important for rational usage. As an important part of the innate immune system, macrophages are widely distributed in vital tissues and are also a dominant cell type that engulfs particles. Mitochondria are one of the most sensitive organelles when macrophages are exposed to NPs. However, previous studies have mainly reported the mitochondrial response upon high-dose NP treatment. Herein, with gold nanoparticles (AuNPs) as a model, we investigated the mitochondrial alterations induced by NPs at a sublethal concentration.ResultsAt a similar internal exposure dose, different AuNPs showed distinct degrees of effects on mitochondrial alterations, including reduced tubular mitochondria, damaged mitochondria, increased reactive oxygen species, and decreased adenosine triphosphate. Cluster analysis, two-way ANOVA, and multiple linear regression suggested that the surface properties of AuNPs were the dominant determinants of the mitochondrial response. Based on the correlation analysis, the mitochondrial response was increased with the change in zeta potential from negative to positive. The alterations in mitochondrial respiratory chain proteins indicated that complex V was an indicator of the mitochondrial response to low-dose NPs.ConclusionOur current study suggests potential hazards of modified AuNPs on mitochondria even under sublethal dose, indicates the possibility of surface modification in biocompatibility improvement, and provides a new way to better evaluation of nanomaterials biosafety.

Insecticidal activity of a proteinaceous α-glycosidase inhibitor isolated from an endophytic Aspergillus awamori and its biosafety evaluation

Agriculture Sector has been affected largely by biotic and abiotic stresses. Among biotic challenges faced by agriculture is the damage caused by insect pests. Insecticides are being widely used for rapid and effective management of insect pests. Despite of their usefulness, the excessive use of insecticides poses potential risks to food safety and environment. To reduce the usage of chemicals, biological control of insect pest can be effective alternative of insecticides for sustainable agriculture. The present study aims to evaluate the insecticidal potential of a protein obtained from an endophytic Aspergillus awamori against insect pest Spodoptera litura (Fab). Variable concentrations of the inhibitor induced high larval mortality ranging from 16.67 to 53%. Larval, pupal and total developmental period of S. litura increased significantly at higher concentrations. The LC50 value of the inhibitor was determined to be 34.34 μg/ml. A significant decrease in adult emergence was recorded at all the concentrations with maximum inhibitory effect at 30 μg/ml where only 65.94% adults emerged as compared to 100.00% in control. The in vivo effect of the inhibitor on gut α-amylase and α-glucosidase was also studied. The activity of α-amylase and α-glucosidase was suppressed to 33.84% and 38.13%, respectively after 72 h at highest concentration of 40 μg/ml. The biosafety evaluation of the extract revealed that the strain is ochratoxin negative and noncytotoxic which supports the above findings that it could be used as biocontrol agent.

Surface-modified ZrO2 nanoparticles with caffeic acid: Characterization and in vitro evaluation of biosafety for placental cells

The toxicity of hybrid nanoparticles, consisting of non-toxic components, zirconium dioxide nanoparticles (ZrO2 NPs), and caffeic acid (CA), was examined against four different cell lines (HTR-8 SV/Neo, JEG-3, JAR, and HeLa). Stable aqueous ZrO2 sol, synthesized by forced hydrolysis, consists of 3–4 nm in size primary particles organized in 30–60 nm in size snowflake-like particles, as determined by transmission electron microscopy and direct light scattering measurements. The surface modification of ZrO2 NPs with CA leads to the formation of an interfacial charge transfer (ICT) complex followed by the appearance of absorption in the visible spectral range. The spectroscopic observations are complemented with the density functional theory calculations using a cluster model. The ZrO2 NPs and CA are non-toxic against four different cell lines in investigated concentration range. Also, ZrO2 NPs promote the proliferation of HTR-8 SV/Neo, JAR, and HeLa cells. On the other hand, hybrid ZrO2/CA NPs induced a significant reduction of the viability of the JEG-3 cells (39 %) for the high concentration of components (1.6 mM ZrO2 and 0.4 mM CA).

ПОВЫШЕНИЕ ЭФФЕКТИВНОСТИ ОПЕРАТИВНОГО КОНТРОЛЯ ПРИ ЭКСПЕРТНОЙ ОЦЕНКЕ КАЧЕСТВА ЗЕРНА

Traditional methods for evaluating sowing and technological characteristics of grain do not sufficiently reveal the caryopsis internal structure damages. They can only be recorded using the microfocus soft X-ray radiography method. The aim of this work was, with the help of X-ray, to evaluate the hidden integrity of the caryopsis structure (including the period of the post-harvest maturation) to increase the efficiency of expert assessment of the degree of economic suitability of grain batches. Seeds of Hordеum vulgare L. variety ‘Suzdalets’ harvested in 2015 in the Gatchina and Vsevolozhsk districts of the Leningrad region, malting barley from the Morskoy Port St. Petersburg (2015–2016), as well as corn grain (Zea mais L.) harvested in 2010 and 2011 were the objects of the research. Grain, which is for feed use only, was provided by the poultry farm “Severnaya”. The poultry farm “Roskar” provided samples harvested in 2017. We used the microfocus radiography method combined with a visual analysis of digital X-ray images to detect hidden grain defects. Experimental data on the evaluation of quality and biosafety of barley and maize grain for various purposes determined by radiographic signs of hidden damage are presented. The survey highlighted several key points: the seed lots obtained in the Vsevolozhsk district had a low total level of hidden defects (39 %); in the seed lots obtained in the Gatchina district, this figure was significantly higher (> 140 %). The germination rate for these options, determined six months after harvesting, turned out to be 89 and 74 %, respectively; it was higher for batches of seeds with a small value of the total indicator of hidden defects. Samples, the total level of hidden defects of which was lower (29 %), had a higher germination index (97 %) compared to the same in seed samples with a total level of hidden defects 70 % or more (85–90 %). X-ray analysis of corn grain samples obtained at the poultry farm “Roskar” revealed that the amount of grain with defects in the embryo (hidden germination) compared with the same in grain lots from the poultry farm “Severnaya” was low and varied from 17 to 29 %. The index of the hidden fissuring rate in these samples was not reliably recorded. However, we detected such internal grain damage as enzyme mycosis depletion (64 to 98 %). Research results are the basis for the development of new engineering and technical solutions for the “gentle” agricultural technologies to obtain batches of high-quality non-injured grain that ensure high competitiveness in the global grain market.

Sex-related response in mice after sub-acute intraperitoneal exposure to silver nanoparticles

Silver nanoparticles (AgNPs) are among the most commercialized nanomaterials in biomedicine due to their antimicrobial and anti-inflammatory properties. Nevertheless, possible health hazards of exposure to AgNPs are yet to be understood and therefore raise public concern in regards of their safety.In this study, sex-related differences, role of steroidal hormones and influence of two different surface stabilizing agents (polymer vs. protein) on distribution and adverse effects of AgNPs were investigated in vivo. Intact and gonadectomised male and female mice were treated with seven AgNPs doses administered intraperitoneally during 21 days. After treatment, steroid hormone levels in serum, accumulation of Ag levels and oxidative stress biomarkers in liver, kidneys, brain and lungs were determined.Sex-related differences were observed in almost all tissues. Concentration of Ag was significantly higher in the liver of females compared to males. No significant difference was found for AgNP accumulation in lungs between females and males, while the lungs of intact males showed significantly higher Ag accumulation compared to gonadectomised group. Effect of surface coating was also observed, as Ag accumulation was significantly higher in kidneys and liver of intact females, as well as in kidneys and brain of intact males treated with protein-coated AgNPs compared to polymeric AgNPs. Oxidative stress response to AgNPs was the most pronounced in kidneys where protein-coated AgNPs induced stronger effects compared to polymeric AgNPs. Interestingly, protein-coated AgNPs reduced generation of reactive oxygen species in brains of females and gonadectomised males. Although there were no significant differences in levels of hormones in the AgNP-exposed animals compared to controls, sex-related differences in oxidative stress parameters were observed in all organs.Results of this study highlight the importance of including the sex-related differences and effects of protein corona in biosafety evaluation of AgNPs exposure.

Biosafety evaluation and quantitative determination of poly(hexamethylene biguanide) (PHMB) coated on cellulosic fabrics by Kubelka–Munk equation

It is difficult to directly determine the quantity of cationic finishing agents on the surface of cellulosic fabrics. Herein, we report a direct and feasible method by Kubelka–Munk equation to quantify the cationic poly (hexamethylene biguanide) hydrochloride (PHMB) adsorbed onto cotton fabrics based on the principle of formation of a stable blue dye between PHMB and bromophenol blue sodium (BPB). The adsorption of PHMB onto cotton fabrics was first investigated and the maximum adsorption of PHMB was found to be around 8 mg per gram of cotton fabric. After being dyed with BPB, colour strength shows a positive correlation with PHMB at low concentrations (< 2400 mg/L). A linear relationship with a high correlation (C(PHMB) = (K/S—0.7411)/3.4517, R2 = 0.9983) was thus established between colour strength and PHMB concentration. However, this equation should fulfill four requirements for quantifying PHMB: (1) the distribution of PHMB on the surface of cellulosic fabric should be in the form of a monolayer with the content less than 5.3 mg/g; (2) an excess of BPB dyebath should be applied; (3) the dyeing should come to equilibrium; and (4) the fabrics should be evenly dyed. Moreover, methyl thiazolyl tetrazolium (MTT) assay results indicate that excess PHMB coated on cotton fabrics can cause cytotoxicity and the quantity of PHMB should not exceed 4.62 mg per gram of cotton fabrics (0.462% w/w) for the purpose of biosafety. The sample can be considered non-cytotoxic if the K/S value is lower than 13.2 after dyeing with BPB.

Biosafety evaluation and detection of shrimp viruses on field samples using dual priming oligonucleotide (DPO) system based multiplex PCR assay

Disease is the current major challenge and issue in shrimp aquaculture as it can be spread easily causing global outbreak. The risk of disease outbreak in shrimp populations can be further minimized with rapid and accurate diagnostic method. This study aims to develop a dual priming oligonucleotide (DPO) based multiplex PCR system for simultaneous detection of shrimp DNA and RNA viruses. This system targets Baculovirus Penaei (BP), Monodon Baculovirus (MB), Infectious Hypodermal, Haematopoietic Necrosis Virus (IHHNV) and White Spot Syndrome Virus (WSSV), Macrobrachium rosenbergii nodavirus (MrNV), Infectious Myonecrosis Virus (IMNV), Taura Syndrome Virus (TSV) and Yellow Head Virus (YHV). The shrimp samples collected from shrimp farm were diagnosed to be infected with YHV infection in the 2nd cycle in year 2018 with 80% screening rate (4/5). DPO based multiplex PCR system may facilitate the rapid and effective screening of shrimp viruses in laboratory.

Biosafety and efficacy evaluation of a biodegradable magnesium-based drug-eluting stent in porcine coronary artery

Although the drug-eluting stent (DES) has become the standard for percutaneous coronary intervention (PCI)-based revascularization, concerns remain regarding the use of DES, mainly due to its permanent rigid constraint to vessels. A drug-eluting bioresorbable stent (BRS) was thus developed as an alternative to DES, which can be absorbed entirely after its therapeutic period. Magnesium (Mg)-based BRSs have attracted a great deal of attention due to their suitable mechanical properties, innovative chemical features, and well-proven biocompatibility. However, the primary disadvantage of Mg-based BRSs is the rapid degradation rate, resulting in the early loss of structural support long before the recovery of vascular function. Recently, a new type of patented Mg–Nd–Zn-Zr alloy (JDBM) was developed at Shanghai Jiao Tong University to reduce the degradation rate compared to commercial Mg alloys. In the present investigation, a poly(d,l-lactic acid)-coated and rapamycin eluting (PDLLA/RAPA) JDBM BRS was prepared, and its biosafety and efficacy for coronary artery stenosis were evaluated via in vitro and in vivo experiments. The degree of smooth muscle cell adhesion to the PDLLA/RAPA coated alloy and the rapamycin pharmacokinetics of JDBM BRS were first assessed in vitro. JDBM BRS and commercial DES FIREHAWK were then implanted in the coronary arteries of a porcine model. Neointimal hyperplasia was evaluated at 30, 90, and 180 days, and re-endothelialization was evaluated at 30 days. Furthermore, Micro-CT and optical coherence tomography (OCT) analyses were performed 180 days after stent implantation to evaluate the technical feasibility, biocompatibility, and degradation characteristics of JDBM BRS in vivo. The results show the ability of a PDLLA/RAPA coated JDBM to inhibit smooth muscle cell adhesion and moderate the drug release rate of JDBM BRS in vitro. In vivo, low local and systemic risks of JDBM BRS were demonstrated in the porcine model, with preserved mechanical integrity after 6 months of implantation. We also showed that this novel BRS was associated with a similar efficacy profile compared with standard DES and high anti-restenosis performance. These findings may confer long term advantages for using this BRS over a traditional DES.

Isolation, Characterization and Biosafety Evaluation of Lactobacillus Fermentum OK with Potential Oral Probiotic Properties.

It has been reported that certain probiotic bacteria have inhibitory effects against oral pathogens. Lactobacillus spp. have been studied and used as probiotics globally, but due to difficulties with laboratory cultivation and experimentation with oral microorganisms, there are few studies on Lactobacillus spp. isolated from the oral cavity being used against oral pathogens. The purpose of this study was to evaluate the biosafety and inhibitory effects of Lactobacillus fermentum OK as a potential oral biotherapeutic probiotic against oral pathogens. L. fermentum OK was evaluated based on microbial and genetic characteristics. A 5% dilution of L. fermentum OK culture supernatant showed that 60% inhibition against the growth of S. mutans and L. fermentum OK displayed significant inhibitory effects against the growth of Fusobacterium nucleatum, Porphyromonas gingivalis, Streptococcus gordonii, and Streptococcus sanguinis. However, proliferation of L. fermentum OK, when co-cultured with harmful oral bacteria, was retarded. L. fermentum OK was shown to produce 1130μmol/L hydrogen peroxide, aggregate efficiently with Streptococcus sobrinus, S. gordonii, S. mutans, S. sanguinis, and P. gingivalis, and reduce S. mutans that produced artificial dental plaque by 97.9%. The in vitro cell adhesion capacity of L. fermentum OK to an oral epithelial cell line was 3.1 cells per cell and the cell adhesion of F. nucleatum and S. mutans decreased strongly in protection and displacement assays. L. fermentum OK was evaluated for safety using ammonia production, biogenic amine production, hemolytic property, mucin degradation testing, antibiotic susceptibility, and whole genome sequencing (WGS). Based on this study, L. fermentum OK appears to be a safe and bioactive lactobacterial food ingredient, starter culture, and/or probiotic microorganism for human oral health.

A rare transgenic event of rice with Agrobacterium binary vector backbone integration at the right T-DNA border junction

Agrobacterium tumefaciens precisely transfers the transferred DNA (T-DNA) between the right border (RB) and the left border (LB) and integrates it in the plant genome. However, transfer of the binary vector backbone (BVB) at the LB junction has been frequently reported. Here, we describe a complex T-DNA integration in the rice transgenic event CG27 with BVB at the RB junction. Amplification of an RB-plant DNA junction fragment by genome walking and its sequencing revealed that the T-DNA portion ended at the 1st bp of RB, but is followed by a binary vector backbone (BVB) along with an RB-flanking T-DNA portion, both in an inverse orientation. This is followed by a 5-bp filler sequence and the rice chromosome 1 sequence. The configuration of the BVB integration at the RB junction was independently confirmed by PCR. Disruption of native chromosome 1 sequence was confirmed by PCR with chromosome 1-specific primers designed from either side of the T-DNA integration site. BVB sequence close to the RB is transferred and integrated into the CG27 plant genome by a mechanism that does not involve LB-skipping and RB read-through. Therefore, a check for BVB sequences at both LB and RB junctions in a transgenic crop is important in biosafety evaluation.