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Korean Journal of Orthodontics Sep 2023To evaluate the effects of an electric toothbrush with vibrational frequencies of 125 Hz and 150 Hz on the orthodontic tooth movement (OTM) rate and the production of...
OBJECTIVE
To evaluate the effects of an electric toothbrush with vibrational frequencies of 125 Hz and 150 Hz on the orthodontic tooth movement (OTM) rate and the production of prostaglandin E2 (PGE2).
METHODS
Out of thirty patients (aged 18-25 years; 16 females and 14 males), ten patients each formed Group A and B, who used electric toothbrushes with 125 Hz and 150 Hz vibrations, respectively. The remaining ten patients (Group C) served as the control group and did not use electric toothbrushes. The rate of OTM and levels of PGE2 using microcapillary pipettes were calculated before the start of retraction (T0), on the 30th day (T1), on the 60th day (T2), and on the 90th day (T3) from the start of retraction in all the groups.
RESULTS
There was a statistically significant difference in the mean OTM values and PGE2 levels in all three groups at different time intervals, with the maximum difference seen in Group B compared to Group A and least in Group C at T1, T2 and T3.
CONCLUSIONS
The rate of OTM and levels of PGE2 were highest in patients who used an electric toothbrush with 150 Hz mechanical vibration compared to those who used an electric toothbrush with 125 Hz mechanical vibration and least in patients who did not use an electric toothbrush. Mechanical vibration led to an increase in the PGE2 levels and accelerated the OTM.
PubMed: 37746776
DOI: 10.4041/kjod23.076 -
Stress Biology Oct 2022The plant vacuole plays a fundamental role in cell homeostasis. The successful application of patch-clamp technique on isolated vacuoles allows the determination of the... (Review)
Review
The plant vacuole plays a fundamental role in cell homeostasis. The successful application of patch-clamp technique on isolated vacuoles allows the determination of the functional characteristics of tonoplast ion channels and transporters. The parallel use of a sensor-based fluorescence approach capable of detecting changes in calcium and proton concentrations opens up new possibilities for investigation. In excised patch, the presence of fura-2 in the vacuolar solution reveals the direct permeation of calcium in plant TPC channels. In whole-vacuole, the activity of non-electrogenic NHX potassium proton antiporters can be measured by using the proton sensitive dye BCECF loaded in the vacuolar lumen by the patch pipette. Both vacuolar NHXs and CLCa (chloride/nitrate antiporter) are inhibited by the phosphoinositide PI(3,5)P, suggesting a coordinated role of these proteins in salt accumulation. Increased knowledge in the molecular mechanisms of vacuolar ion channels and transporters has the potential to improve our understanding on how plants cope with a rapidly changing environment.
PubMed: 37676514
DOI: 10.1007/s44154-022-00064-z -
BMC Biotechnology Aug 2023The in-vitro scratch assay is a useful method in wound healing research to assess cell migration. In this assay, a scratch is created in a confluent cell layer by...
BACKGROUND
The in-vitro scratch assay is a useful method in wound healing research to assess cell migration. In this assay, a scratch is created in a confluent cell layer by mechanically removing cells through manual scraping with a sharp-edged tool. This step is traditionally done with pipette tips and is unsuitable for high-throughput assays, as the created scratches are highly variable in width and position. Commercially available solutions are often expensive, and require specific cultureware which might not be suitable for all studies.
RESULTS
In this study, we have developed a flexible cell scratch device comprising a single wounding tool, a guide and an imaging template for consistent and reproducible scratch assays in 96-well plates. Our results showed that the device produced a more consistent scratch profile compared to the conventional method of using pipette tips. The imaging template also allowed operators to easily locate and image the same region of interest at different time points, which potentially could be used for other assays.
CONCLUSIONS
Our flexible yet effective scratch device thus enables robust scratch assays that can be applied to different experimental needs, providing researchers with an easy and reliable tool for their studies.
Topics: Research Design; Biological Assay; High-Throughput Screening Assays; Wound Healing
PubMed: 37641063
DOI: 10.1186/s12896-023-00806-5 -
Plant Disease Aug 2023Quinoa (Chenopodium quinoa Willd.) is a traditional food originally from the Andes Mountains in South America. It was first planted in China in 1987 and is grown in...
Quinoa (Chenopodium quinoa Willd.) is a traditional food originally from the Andes Mountains in South America. It was first planted in China in 1987 and is grown in Tibet, Gansu, and Qinghai provinces. In May 2021, 40% of 2-month-old quinoa plants in the 3.4 hm² experimental base of Qinghai University (36.7262° N, 101.7487° E) were found to have leaves with grey-brown subcircular spots (about 0.4 to 0.7 cm) with black dots (acervuli). Severely infected plants exhibited symptoms such as withered and stunted growth. The diseased-healthy junctions of infected leaves (0.5 cm) were cut out, disinfected with 3% NaClO for 1.5 min, washed three times with sterile water, dried, placed on water agar, and incubated at 25°C for 48 h. After sporulation was seen on the leaf surface, spore suspensions were prepared by placing conidia in sterile water using a pipette. Next, 200 μl of each spore suspension was spread on the surface of water agar and incubated at 25°C for 12 h. Single spores were selected under a stereomicroscope and cultured on potato dextrose agar (PDA) (Qi et al. 2022). The mycelium of two representative isolates (20DLMF-5-4-1 and 20DLMF-7-4-1) was grey-black with white edges and included a fluffy aerial mycelium. Conidia were unicellular, colorless, long ellipsoid or curved moon shaped, averaging 14.3 × 1.8 to 20.2 × 2.2 μm (n=100). The light brown appressoria were ovoid, averaging 8.5 × 5.2 to 7.7 × 4.1 μm (n=20). Spherical, dark brown acervuli were observed on the leaves, averaging 160 to 200 μm (n=20), and there were dark brown spiny bristles. The ITS, partial ACT, CHS, GAPDH and TUB2 genes were amplified from genomic DNA of the two isolates (Weir et al. 2012). Sequences were deposited in GenBank (accession no. OQ871595 to OQ871602 for ACT, CHS, GAPDH, and TUB2, and OQ860235 to OQ860236 for ITS) and showed over 99% identities with the corresponding sequences of C. spinaciae CBS125347 and CBS128.57 (Vu et al. 2019; Damm et al. 2009). Both isolates clustered with the type culture of C. spinaciae (CBS125347, CBS128.57), with 100% bootstrap support in the phylogenetic tree. Thus, according to the morphological and molecular characteristics, the two isolates were identified as C. spinaciae. Pathogenicity tests were conducted on 24 healthy, tender leaves of six 1-month-old quinoa plants, with three replicates (Yang et al. 2021). The leaves were gently scratched in 3-4 areas with a sterile needle. A conidial suspension (105 conidia/ml) of the two isolates was sprayed on these wounds. The control group was unscratched and sprayed with sterile water. The plants were incubated in a greenhouse at 25°C for 24 h in the dark and 7 days in the light. Tiny grey-brown spots appeared on day 3 (about 0.4 to 0.6 cm) and continued to enlarge until perforations and ruptures developed on day 7. Subsequently, acervuli were observed on the surface of the leaves. The control leaves remained healthy. Isolates were reisolated from the symptomatic leaves and they had the same morphological and molecular characteristics as the original isolates, confirming Koch's postulates. To our knowledge, this is the first report of C. spinaciae causing quinoa leaf anthracnose in China. C. spinaciae seriously affects the yield and quality of quinoa and has been previously reported to cause anthracnose of Vicia sativa in China (Wang et al. 2019). The results provide a basis for the study and control of quinoa leaf anthracnose.
PubMed: 37622274
DOI: 10.1094/PDIS-07-23-1285-PDN -
Frontiers in Microbiology 2023Swab pooling may allow for more efficient use of point-of-care assays for SARS-CoV-2 detection in settings where widespread testing is warranted, but the effects of...
INTRODUCTION
Swab pooling may allow for more efficient use of point-of-care assays for SARS-CoV-2 detection in settings where widespread testing is warranted, but the effects of pooling on assay performance are not well described.
METHODS
We tested the Thermo-Fisher Accula rapid point-of-care RT-PCR platform with contrived pooled nasal swab specimens.
RESULTS
We observed a higher limit of detection of 3,750 copies/swab in pooled specimens compared to 2,250 copies/swab in individual specimens. Assay performance appeared worse in a specimen with visible nasal mucous and debris, although performance was improved when using a standard laboratory mechanical pipette compared to the transfer pipette included in the assay kit.
CONCLUSION
Clinicians and public health officials overseeing mass testing efforts must understand limitations and benefits of swab or sample pooling, including reduced assay performance from pooled specimens. We conclude that the Accula RT-PCR platform remains an attractive candidate assay for pooling strategies owing to the superior analytical sensitivity compared to most home use and point-of-care tests despite the inhibitory effects of pooled specimens we characterized.
PubMed: 37608952
DOI: 10.3389/fmicb.2023.1219214 -
Journal of Ophthalmic & Vision Research 2023Animal models are necessary in understanding the pathogenesis of endophthalmitis and are also necessary to assist the development of new therapeutics for this...
PURPOSE
Animal models are necessary in understanding the pathogenesis of endophthalmitis and are also necessary to assist the development of new therapeutics for this sight-threatening ocular inflammation. Hamilton syringes are usually preferred to inject pathogens when performing experiments on test subjects, however, this method has technical and financial disadvantages. In this study, we report the findings and assess the related benefits of applying a novel low-cost intravitreal injection technique to initiate endophthalmitis in a mouse model while using the Eppendorf tip and a 26G needle.
METHODS
The 18-hr culture of clinical isolates of bacteria ( and ) and fungus ( and ) were resuspended to a final concentration of 10,000 colony forming units (CFU)/1 µL which were separately injected intravitreally into C57BL/6 mice (6-8 weeks) using a 0.1-2.5µL pipette attached to the modified Eppendorf tip with a 26G needle. The contralateral eye served as vehicle/uninjected control. Disease progression was determined by assessing the corneal haze, opacity, bacterial burden, and retinal histology of the eyes used in the model. Following euthanization, bacteria-infected mice were enucleated after 24 hr of the initial injection, and fungus-infected mice after 72 hr.
RESULTS
Of the 50 mice injected, the modified technique was successful in 48 mice. Two mice were excluded due to cataract formed by accidental injury to the lens. The experimental endophthalmitis mice model successfully mimicked the natural clinical course. Clinical assessment and histopathology confirmed the influx of inflammatory cells into the posterior segment of the eye along with dissolution of retinal architecture.
CONCLUSION
Our novel method of injection using a modified Eppendorf tip and 26G needle yielded a cost-effective mouse model of clinical endophthalmitis, resulting in reproducible infection for understanding various aspects of its pathobiology.
PubMed: 37600911
DOI: 10.18502/jovr.v18i3.13775 -
Light, Science & Applications Aug 2023The dynamics and structure of mixed phases in a complex fluid can significantly impact its material properties, such as viscoelasticity. Small-angle X-ray Photon...
The dynamics and structure of mixed phases in a complex fluid can significantly impact its material properties, such as viscoelasticity. Small-angle X-ray Photon Correlation Spectroscopy (SA-XPCS) can probe the spontaneous spatial fluctuations of the mixed phases under various in situ environments over wide spatiotemporal ranges (10-10 s /10-10 m). Tailored material design, however, requires searching through a massive number of sample compositions and experimental parameters, which is beyond the bandwidth of the current coherent X-ray beamline. Using 3.7-μs-resolved XPCS synchronized with the clock frequency at the Advanced Photon Source, we demonstrated the consistency between the Brownian dynamics of ~100 nm diameter colloidal silica nanoparticles measured from an enclosed pendant drop and a sealed capillary. The electronic pipette can also be mounted on a robotic arm to access different stock solutions and create complex fluids with highly-repeatable and precisely controlled composition profiles. This closed-loop, AI-executable protocol is applicable to light scattering techniques regardless of the light wavelength and optical coherence, and is a first step towards high-throughput, autonomous material discovery.
PubMed: 37596264
DOI: 10.1038/s41377-023-01233-z -
Plant Disease Aug 2023Multiple Diaporthe spp. cause root and fruit rots or stem lesions on Cucumis spp.: (syn. ), , and (Broge et al. 2020; Fukada et al. 2018; Udayanga et al. 2012, 2015)....
Multiple Diaporthe spp. cause root and fruit rots or stem lesions on Cucumis spp.: (syn. ), , and (Broge et al. 2020; Fukada et al. 2018; Udayanga et al. 2012, 2015). From May-August 2021, cucumbers () 'Katrina' and 'Alcazar' were grown in a 24-plant, commercial Bato bucket system with rockwool blocks on a perlite substrate in a research greenhouse in Wooster, Ohio. At maturity, plants collapsed rapidly from stem lesions without foliar chlorosis (25% of 'Katrina' and 17% of 'Alcazar'). Lesions were 7.5 to 15 cm in length, tan to golden-brown with black pycnidia and located 5 to 15 cm above the crown. Stems shredded easily with vascular discoloration around the lesion. Two identical fungal strains were isolated on ½ acidified potato dextrose agar (APDA) following surface disinfestation with 0.6% sodium hypochlorite for 30 s and sterile water rinse. Fungal cultures were floccose, white to tan mycelia with pycnidia. Oblong, elliptical, biguttulate, aseptate alpha conidia were observed with mean dimensions: 8.0 µm (5.2-9.8 µm) by 3.1 µm (2.5-3.8 µm) on ½ APDA and 9.8 µm (6.6-12.4 µm) by 3.0 µm (1.9-5.3 µm) on petioles. On prune extract agar, beta conidia mean dimensions were: 19.7 µm (12.0-27.7 µm) by 1.2 µm (0.8-1.8 µm). Fungal DNA was amplified and sequenced bidirectionally with ITS (ITS4/ITS5), CAL (CAL228F/737R), HIS (CYLH3F/H3-1B), TEF1 (EF1-728f/EF1-986R), and TUB2 (Bt1a/Bt1b) primers (Carbone and Kohn 1999; Glass and Donaldson 1995) (GenBank: OP265712-13, OP288460-65, OQ418506-07). Based on a maximum likelihood phylogenetic tree of concatenated genes, this novel sp., most closely related to D. stewartii, has not been reported on Cucumis spp. Strains were deposited in the USDA-ARS Culture Collection (NRRL# 64461-62). Koch's postulates were conducted in a greenhouse with mean day temperature of 25°C and 12 hr supplemental lighting. One-month old cucumbers 'Katrina,' grown in rockwool cubes (5 plants per isolate) and potting mix (6 plants per isolate), were inoculated with a one-week-old culture of either strain. The second true leaf was cut and a pipette tip containing an inoculated plug of ½ APDA was placed on the remaining petiole (Mathew et al. 2018). Non-inoculated ½ APDA was used for controls. Plants were tarped for 24 hours to increase humidity and pipette tips removed after one week. After two weeks, petioles were shrunken, tan to golden brown with pycnidia. After 3-4 weeks, stem lesions matching those above were observed on inoculated plants, and plants collapsed. For fruit rot, three Beit Alpha cucumbers were rinsed with tap water, dried, a 5 mm plug was removed from the fruit and replaced with a 5 mm plug of one-week-old fungus on ½ APDA. After 3 days, fruits were water soaked and soft. For root rot, two plates of one-week-old cultures were macerated in 500 mL of sterile water and mixed with 1500 mL of vermiculite. Two seeds of cucumber 'Katrina' were planted into three reps of each isolate and control. All control seeds germinated, but all inoculated seeds experienced pre- or post-emergence damping off. No symptoms were ever observed on any controls. Fungi were isolated from all inoculated tissues as described above. Based on morphology, Diaporthe sp. was isolated from all inoculated plants but never from controls. This Diaporthe sp. may be a new constraint to hydroponic cucumber production, but incidence needs to be determined globally.
PubMed: 37578364
DOI: 10.1094/PDIS-06-23-1214-PDN -
Longitudinal diffusion barriers imposed by myofilaments and mitochondria in murine cardiac myocytes.The Journal of General Physiology Oct 2023Using optical and electrical methods, we document that diffusion in the cytoplasm of BL6 murine cardiomyocytes becomes restricted >20-fold as molecular weight increases...
Using optical and electrical methods, we document that diffusion in the cytoplasm of BL6 murine cardiomyocytes becomes restricted >20-fold as molecular weight increases from 30 to 2,000, roughly as expected for pores with porin channel dimensions. Bodipy-FL ATP diffuses >40-fold slower than in free water at 25°C. From several fluorophores analyzed, bound fluorophore fractions range from 0.1 for a 2 kD FITC-labeled polyethylene glycol to 0.93 for sulforhodamine. Unbound fluorophores diffuse at 0.5-8 × 10-7 cm2/s (5-80 μm2/s). Analysis of Na/K pump and veratridine-modified Na channel currents suggests that Na diffusion is nearly unrestricted at 35°C (time constant for equilibration with the pipette tip, ∼20 s). Using multiple strategies, we estimate that at 35°C, ATP diffuses four to eight times slower than in free water. To address whether restrictions are caused more by protein or membrane networks, we verified first that a protein gel, 10 g% gelatin, restricts diffusion with strong dependence on molecular weight. Solute diffusion in membrane-extracted cardiac myofilaments, confined laterally by suction into large-diameter pipette tips, is less restricted than in intact myocytes. Notably, myofilaments extracted similarly from skeletal (diaphragm) myocytes are less restrictive. Solute diffusion in myocytes with sarcolemma permeabilized by β-escin (80 µM) is similar to diffusion in intact myocytes. Restrictions are strain-dependent, being twofold greater in BL6 myocytes than in CD1/J6/129svJ myocytes. Furthermore, longitudinal diffusion is 2.5-fold more restricted in CD1/J6/129svJ myocytes lacking the mitochondrial porin, VDAC1, than in WT CD1/J6/129svJ myocytes. Thus, mitochondria networks restrict long-range diffusion while presumably optimizing nucleotide transfer between myofilaments and mitochondria. We project that diffusion restrictions imposed by both myofilaments and the outer mitochondrial membrane are important determinants of total free cytoplasmic AMP and ADP (∼10 μM). However, the capacity of diffusion to deliver ATP to myofilaments remains ∼100-fold greater than ATP consumption.
Topics: Mice; Animals; Myocytes, Cardiac; Myofibrils; Mitochondria; Diffusion; Voltage-Dependent Anion Channels; Adenosine Triphosphate; Water
PubMed: 37555782
DOI: 10.1085/jgp.202213329 -
Forensic Toxicology Jan 2024We developed and validated a method for quantitative analysis of ten synthetic cathinones in oral fluid (OF) samples, using microextraction by packed sorbent (MEPS) for...
Green analytical toxicology method for determination of synthetic cathinones in oral fluid samples by microextraction by packed sorbent and liquid chromatography-tandem mass spectrometry.
PURPOSE
We developed and validated a method for quantitative analysis of ten synthetic cathinones in oral fluid (OF) samples, using microextraction by packed sorbent (MEPS) for sample preparation followed by liquid chromatography‒tandem mass spectrometry (LC‒MS/MS).
METHOD
OF samples were collected with a Quantisal™ device and 200 µL was extracted using a C18 MEPS cartridge installed on a semi-automated pipette and then analyzed using LC‒M/SMS.
RESULTS
Linearity was achieved between 0.1 and 25 ng/mL, with a limit of detection (LOD) of 0.05 ng/mL and a limit of quantification (LOQ) of 0.1 ng/mL. Imprecision (% relative standard deviation) and bias (%) were better than 11.6% and 7.5%, respectively. The method had good specificity and selectivity against 9 different blank OF samples (from different donors) and 68 pharmaceutical and drugs of abuse with concentrations varying between 400 and 10,000 ng/mL. No evidence of carryover was observed. The analytes were stable after three freeze/thaw cycles and when kept in the autosampler (10 °C) for up to 24 h. The method was successfully applied to quantify 41 authentic positive samples. Methylone (mean 0.6 ng/mL, median 0.2 ng/mL), N-ethylpentylone (mean 16.7 ng/mL, median 0.35 ng/mL), eutylone (mean 39.1 ng/mL, median 3.6 ng/mL), mephedrone (mean 0.5 ng/mL, median 0.5 ng/mL), and 4-chloroethcathinone (8.1 ng/mL) were quantified in these samples.
CONCLUSION
MEPS was an efficient technique for Green Analytical Toxicology purposes, which required only 650 µL organic solvent and 200 µL sodium hydroxide, and the BIN cartridge had a lifespan of 100 sample extractions.
Topics: Chromatography, Liquid; Synthetic Cathinone; Tandem Mass Spectrometry; Solid Phase Microextraction; Limit of Detection
PubMed: 37505359
DOI: 10.1007/s11419-023-00671-z