Monocarboxylate transporter 1-mediated lactate accumulation promotes nucleus pulposus degeneration under hypoxia in a 3D multilayered nucleus pulposus degeneration model.

During intervertebral disc degeneration (IVDD), due to endplate calcification, diminished oxygen and nutrient concentrations and accumulated lactate are present in the microenvironment of the nucleus pulposus (NP). The disadvantages of 3D layered culture include uneven oxygen and nutrient gradients. In the present study, to mimic the in vivo microenvironment of the NP, a 5-layered 3D culture was constructed using clinical haemostatic gelatine sponges and developed as a NP degeneration (NPD) model. Subsequently, cell distribution as well as expression of NP chondrogenic markers (type II collagen and aggrecan), glycosaminoglycan (GAG) and degeneration markers [e.g. matrix metalloproteinase (MMP) 3] were measured from the top to the bottom layer. However, in a single NP-cell-loaded disc model, the chondrogenic potency in the middle or bottom layer was higher than that in the top layer. To further study the mechanism underlying the degeneration of NP cells in this NPD model, the contribution of secreted metabolites was examined. Lactate identified in the supernatant modulated GAG accumulation and MMP3 expression. Inhibition of lactate influx by the monocarboxylate transporter (MCT)-1 inhibitor, AZD3965, reversed the effect of lactate on GAG accumulation and MMP3 expression and further improved NP cell degeneration in the NPD model. Thanks to the homogenous expression of lactate in the model, it was possible to further identified that the combination of lactate and hypoxia enhanced MMP3 expression. Taken together, multilayered cell-loaded sponges, with oxygen and nutrient gradients as well as lactate accumulation, can represent a 3D multilayered NPD model for exploring potential agents for IVDD.

Identification and functional analysis of the cytolethal distending toxin gene from Avibacterium paragallinarum.

Avibacterium paragallinarum is the causative agent of infectious coryza, an important respiratory disease of chickens. Cytolethal distending toxins (CDTs) are a family of protein cytotoxins that cause cell cycle arrest and apoptosis in eukaryotic cells. Whole-genome sequencing analysis showed that Av. paragallinarum contains cdtABC genes. Filter-sterilized lysates prepared from Av. paragallinarum or from recombinant Escherichia coli expressing cdtABC genes exhibited CDT activity on HeLa cells and chicken embryo fibroblast (DF-1) cells. In vitro DNase assays showed that purified recombinant CdtB has DNase activity. Polymerase chain reaction and sequencing analysis revealed that the cdtABC genes are present in all strains of Av. paragallinarum examined in this study. This is the first report of the identification and functional analysis of cdtABC genes from Av. paragallinarum. The gene products of cdtABC genes may be involved in the pathogenesis of the disease caused by Av. paragallinarum.

Electric Scooter Battery Detonation: A Case Series And Review Of Literature.

Since 2016 there has been a 20-fold increase in known burns injury from personal mobility device (PMD) related fires. The root cause is the failure of high-density lithium ion (Li-ion) battery packs powering the PMDs. This failure process, known as thermal runaway, is well documented in applied science journals. Importantly, the liberation of hydrogen fluoride from failing Li-ion batteries may contribute to unrecognized chemical burns. A clinical gap in knowledge exists in the understanding of the explosive nature of Li-ion batteries. We reviewed the electrochemical pathophysiology of a failing Li-ion cell as it impacts clinical management of burn injuries. This retrospective study was carried out in two major institutions in Singapore. All admitted PMD-related burns and follow up appointments were captured and reviewed from 2016 - 2020. Thirty patients were admitted to tertiary hospitals, 43% of patients were in the pediatric population and 57% were adult patients, aged from 0.3 to 77 years. TBSA of burns ranged from 0 to 80% with a mean 14.5%. 73% of cases presented with inhalation injury, 8 of whom did not suffer any cutaneous burns. 50% of patients sustained both cutaneous and inhalation burn injuries. 27% of patients sustained major burns of >20% TBSA, with 2 in the pediatric group. Mortali ty rate was 10% from PMD-related fires. This cause of burn injury has proven to be fa tal. Prevention of PMD-related fires by ensuring proper battery utilization, adherence to PMD sanctions for battery standards and public education is vital to reducing the morbidity and mortality of this unique type of thermal injury.

Depuis 2016, les rapports de brûlures après incendie de véhicules électriques personnels (VEP) ont été multipliés par 20. La cause essentielle en est le dysfonctionnement de la batterie lithium/ion (Li/ion) les motorisant. Ce dysfonctionnement est connu sous le terme d'emballement thermique, bien décrit dans les revues technologiques. La libération de fluorure d'hydrogène lors de cette réaction peut entraîner des brûlures chimiques ignorées et la physiopathologie exacte de ces brûlures reste largement méconnue des cliniciens. Nous avons revu les mécanismes physico- chimiques de l'emballement thermique des batteries Li/ion et leur conséquences sur la prise en charge des brûlures occasionnées. Cette étude rétrospective a été réalisée par 2 grosses structures singapouriennes. Tous les dossiers d'accidents de VEP survenus entre 2016 et 2020, comprenant le suivi à distance, ont été revus. Ils regroupaient 30 patients âgés de 3 mois à 77 ans, dont 43% d'enfants. La surface brûlée représentait 0 à 80% de SCT (moyenne 14,5%) et 27% des patients (dont 2 enfants) étaient brûlés sur plus de 20% SCT. Une inhalation était retrouvée dans 73% des cas (dont 8 sans brûlure cutanée). La moitié des patients avaient une brûlure et une inhalation. La mortalité s'élevait à 10%. La prévention de ces accidents par le contrôle- qualité des batteries (sanctions à l'appui) et l'éducation à l'utilisation correcte des VEP et de leur batterie est nécessaire pour éviter ces dysfonctionnements potentiellement létaux.

Pharmacokinetics and tissue depletion of florfenicol in Leghorn and Taiwan Native chickens.

Florfenicol (Ff) is a synthetic antibiotic with a broad antibacterial spectrum and high therapeutic effectiveness that was specifically developed for veterinary use. In the present study, tissue residual levels and the pharmacokinetics of Ff after oral administration of 30 mg/kg to Leghorn and Taiwan Native chicken were studied. Furthermore, differential pharmacokinetics between leg and breast muscles were compared using samples collected from an optimized microdialysis model designed for avian species. Significant differences in C(max) were detected between the plasma and muscle microdialysates, and between the breast and leg microdialysates of the Leghorn chickens by noncompartmental pharmacokinetic analysis. After a single oral dose of Ff at 30 mg/kg, the drug was quickly absorbed and widely distributed with tissue penetration factors significantly different between leg and breast muscles. The serum protein binding of Ff was estimated to be 16.8 ± 1.2%. Significant breed differences in tissue depletion were noted and characterized by higher Ff concentration in the brain, lung, kidney and at least 12 h longer resident times in kidney, heart and spleen for Taiwan Native chicken. Results from this investigation demonstrate the practicality of using in vivo microdialysis in chickens for pharmacokinetic studies and reveal significant time-dependent differences in the free concentrations of Ff in leg and breast muscles. The tissue depletion study signified breed differences in tissue residue concentration and detection times between Leghorn and Taiwan Native chickens. Therefore, currently used withdrawal times for Ff in chickens can not be assumed safe for Taiwan Native chickens.

Serotypes and Hemagglutinin Gene Sequences of Avibacterium paragallinarum Isolated in Taiwan.

Despite routine vaccine use, sporadic outbreaks of infectious coryza in poultry continue to occur in Taiwan. This study was designed to determine the serotypes and the complete nucleotide sequences of a hemagglutinin gene (HMTp210) of Avibacterium paragallinarum isolated in Taiwan between 1994 and 2017. Hemagglutination inhibition tests showed that these isolates belong to serogroups B and C. Sequence analyses of the HMTp210 gene showed that Taiwanese serogroup B isolates are most similar (94.7%-98.2% identity) to strain FARPER-174 isolated in Peru in 2015. In contrast, Taiwanese serogroup C isolates are most similar (96.3%-99.8% identity) to strain H-18 isolated in Japan in 1976. This is the first report showing the presence of A. paragallinarum of serogroup B in Taiwan. In addition, one Taiwanese isolate showed cross-reactivity with serogroup B and C antisera. This isolate contains a chimeric HMTp210 gene that might result from recombination between serogroups B and C. These findings could be valuable for the epidemiologic study and molecular serotyping of A. paragallinarum.

Serotipos y secuencias de genes de hemaglutinina de Avibacterium paragallinarum aislados en Taiwán. A pesar del uso rutinario de vacunas, en Taiwán continúan ocurriendo brotes esporádicos de coriza infecciosa en avicultura. Este estudio fue diseñado para determinar los serotipos y las secuencias de nucleótidos completas de un gene de hemaglutinina (HMTp210) de Avibacterium paragallinarum aislado en Taiwán entre 1994 y 2017. Las pruebas de inhibición de la hemaglutinación mostraron que estos aislamientos pertenecen a los serogrupos B y C. El análisis de secuencias del gene HMTp210 mostró que los aislamientos del serogrupo B taiwaneses son más similares (94.7% ­98.2% de identidad) a la cepa FARPER-174 aislada en Perú en el año 2015. En contraste, los aislamientos del serogrupo C taiwaneses son más similares (96.3% ­99.8% de identidad) a la cepa H -18 aislada en Japón en 1976. Este es el primer reporte que muestra la presencia de A. paragallinarum del serogrupo B en Taiwán. Además, un aislado taiwanés mostró reactividad cruzada con los antisueros del serogrupo B y C. Este aislado contiene un gene HMTp210 quimérico que podría resultar de la recombinación entre los serogrupos B y C. Estos hallazgos podrían ser valiosos para el estudio epidemiológico y la serotipificación molecular de A. paragallinarum.

White Matter Lesion Penumbra Shows Abnormalities on Structural and Physiologic MRIs in the Coronary Artery Risk Development in Young Adults Cohort.

BACKGROUND AND PURPOSE: White matter lesions are 1 age-related manifestation of cerebrovascular disease, but subthreshold abnormalities have been identified in nonlesional WM. We hypothesized that structural and physiologic MR imaging findings of early cerebrovascular disease can be measured in middle-aged subjects in tissue adjacent to WM lesions, termed "penumbra." MATERIALS AND METHODS: WM lesions were defined using automated segmentation in 463 subjects, 43-56 years of age, from the Coronary Artery Risk Development in Young Adults (CARDIA) longitudinal observational cohort study. We described 0- to 2-mm and 2- to 4-mm-thick spatially defined penumbral WM tissue ROIs as rings surrounding WM lesions. The remaining WM was defined as distant normal-appearing WM. Mean signal intensities were measured for FLAIR, T1-, and T2-weighted images, and from fractional anisotropy, mean diffusivity, CBF, and vascular reactivity maps. Group comparisons were made using Kruskal-Wallis and pair-wise t tests. RESULTS: Lesion volumes averaged 0.738 ± 0.842 cm3 (range, 0.005-7.27 cm3). Mean signal intensity for FLAIR, T2, and mean diffusivity was increased, while T1, fractional anisotropy, and CBF were decreased in white matter lesions versus distant normal-appearing WM, with penumbral tissues showing graded intermediate values (corrected P < .001 for all group/parameter comparisons). Vascular reactivity was significantly elevated in white matter lesions and penumbral tissue compared with distant normal-appearing white matter (corrected P ≤ .001). CONCLUSIONS: Even in relatively healthy 43- to 56-year-old subjects with small white matter lesion burden, structural and functional MR imaging in penumbral tissue reveals significant signal abnormalities versus white matter lesions and other normal WM. Findings suggest that the onset of WM injury starts by middle age and involves substantially more tissue than evident from focal white matter lesions visualized on structural imaging.

The Fimbrial Protein is a Virulence Factor and Potential Vaccine Antigen of Avibacterium paragallinarum.

Fimbriae are recognized as virulence factors and potential vaccine antigens of several pathogenic bacteria, but the function of the fimbriae from Avibacterium paragallinarum is not well known. In this study, a gene encoding the fimbrial protein FlfA was identified in A. paragallinarum . Sequencing analysis of the putative promoter region of flfA suggests that flfA expression in A. paragallinarum might be controlled by phase variation. The flfA gene from A. paragallinarum was expressed as a recombinant protein (r-FlfA) in Escherichia coli . Immunization with r-FlfA conferred chickens protection against challenge infection with A. paragallinarum . Virulence assays showed that the flfA-deficient mutants of A. paragallinarum were less virulent than their parental wild-type strains. These results indicated that the fimbrial protein FlfA is a virulence factor and potential vaccine antigen from A. paragallinarum .

Cell sorting of formalin-treated pathogenic Mycobacterium paratuberculosis expressing GFP.

GFP is widely used as a molecular tool for the study of microbial pathogens. However, the manipulation of these pathogenic microorganisms poses a health threat to the laboratory worker, requiring biosafety level II or III containment. Although the GFPfluorophore is tolerant toformalin, a thorough analysis of this treatment on fluorescent output in prokaryotic systems has not been described. In addition, the analysis of microorganisms expressing GFP often depends on specialized equipment, which may not be housed in biosafety level II or III laboratories. Therefore, we sought to develop a safe and effective method for manipulating the GFP-expressing pathogenic bacterium Mycobacterium avium subsp, paratuberculosis (M. paratuberculosis) utilizing a formalin treatment that would permit the analysis of GFP fluorescence without requiring stringent biosafety containment. We demonstrate that formalin-treated M. paratuberculosis expresses 50% less fluorescence than viable cells, but this reduction is still compatible with spectrofluorometry and cell sorting. Furthermore, plasmid DNA that expresses GFP can be recovered efficiently from nonviable, sorted fluorescent cells. This approach is flexible, provides an additional margin of safety for laboratory personnel, and can be easily applied to other pathogenic microorganisms expressing GFP.

Molecular identification and characterization of turkey IFN-gamma gene.

The cDNAs of turkey and chicken interferon gamma (IFN-gamma) were cloned and the functional activity of turkey and chicken IFN-gamma was compared. The coding region of turkey IFN-gamma gene encodes a predicted mature protein of 145 amino acids with a molecular weight at 16.8 kDa. Compared with type I IFN, the IFN-gamma between turkey and chicken also had the same size and high degree of identity at the nucleotide (96.0%) and amino acid (96.4%) sequence. Turkey IFN-gamma was cross-reactive with chicken cells. Both turkey and chicken IFN-gamma could induce production of nitric oxide by turkey or chicken macrophages. Turkey IFN-gamma also had similar degree of sensitivity to heat and pH 2.0 as chicken IFN-gamma. The functional activity of both turkey and chicken IFN-gamma could be neutralized by a monoclonal antibody specific to chicken IFN-gamma to a similar extent. These results indicated that IFN-gamma protein was cross-reactive between turkey and chicken.

Pilot-scale cooling tower to evaluate corrosion, scaling, and biofouling control strategies for cooling system makeup water.

Pilot-scale cooling towers can be used to evaluate corrosion, scaling, and biofouling control strategies when using particular cooling system makeup water and particular operating conditions. To study the potential for using a number of different impaired waters as makeup water, a pilot-scale system capable of generating 27,000 kJ∕h heat load and maintaining recirculating water flow with a Reynolds number of 1.92 × 10(4) was designed to study these critical processes under conditions that are similar to full-scale systems. The pilot-scale cooling tower was equipped with an automatic makeup water control system, automatic blowdown control system, semi-automatic biocide feeding system, and corrosion, scaling, and biofouling monitoring systems. Observed operational data revealed that the major operating parameters, including temperature change (6.6 °C), cycles of concentration (N = 4.6), water flow velocity (0.66 m∕s), and air mass velocity (3660 kg∕h m(2)), were controlled quite well for an extended period of time (up to 2 months). Overall, the performance of the pilot-scale cooling towers using treated municipal wastewater was shown to be suitable to study critical processes (corrosion, scaling, biofouling) and evaluate cooling water management strategies for makeup waters of complex quality.