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1.
Pol J Vet Sci ; 23(2): 197-202, 2020 Jun.
Article in English | MEDLINE | ID: mdl-32627977

ABSTRACT

In this study, a SYBR Green-based real-time quantitative polymerase chain reaction (qPCR) assay was developed for rapid detection of porcine parvovirus (PPV) 6. Primer pairs targeting the conserved regions of PPV6 Capsid gene were designed. Sensitivity analyses revealed the lowest detection limit of the SYBR Green-based real-time PCR assay to be 47.8 copies/µL, which indicated it was 1000 times higher than that found in the conventional PCR investigations. This assay was specific and showed no cross-species amplification with other six porcine viruses. The assay demonstrated high repeatability and reproducibility; the intra- and inter-assay coefficients of variation were 0.79% and 0.42%, respectively. The positive detection rates of 180 clinical samples with SYBR Green-based real-time PCR and conventional PCR were 12.22% (22/180) and 4.44% (8/180), respectively. Our method is sensitive, specific, and reproducible. The use of SYBR Green-based real-time PCR may be suitable for the clinical detection and epidemiological investigation of PPV6.


Subject(s)
Benzothiazoles , Diamines , Parvoviridae Infections/veterinary , Parvovirus, Porcine/isolation & purification , Quinolines , Real-Time Polymerase Chain Reaction/veterinary , Swine Diseases/virology , Animals , Parvoviridae Infections/diagnosis , Parvoviridae Infections/virology , Parvovirus, Porcine/classification , Real-Time Polymerase Chain Reaction/methods , Reproducibility of Results , Sensitivity and Specificity , Swine , Swine Diseases/diagnosis
2.
Plant Biol (Stuttg) ; 22(5): 794-804, 2020 Sep.
Article in English | MEDLINE | ID: mdl-32501628

ABSTRACT

Heat stress decreases crop growth and yield worldwide. Spermidine (Spd) is a small aliphatic amine and acts as a ubiquitous regulator for plant growth, development and stress tolerance. Objectives of this study were to determine effects of exogenous Spd on changes in endogenous polyamine (PA) and γ-aminobutyric acid (GABA) metabolism, oxidative damage, senescence and heat shock protein (HSP) expression in white clover subjected to heat stress. Physiological and molecular methods, including colorimetric assay, high performance liquid chromatography and qRT-PCR, were applied. Results showed that exogenous Spd significantly alleviated heat-induced stress damage. Application of Spd not only increased endogenous putrescine, Spd, spermine and total PA accumulation, but also accelerated PA oxidation and improved glutamic acid decarboxylase activity, leading to GABA accumulation in leaves under heat stress. The Spd-pretreated white clover maintained a significantly higher chlorophyll (Chl) content than untreated plants under heat stress, which could be related to the roles of Spd in up-regulating genes encoding Chl synthesis (PBGD and Mg-CHT) and maintaining reduced Chl degradation (PaO and CHLASE) during heat stress. In addition, Spd up-regulated HSP70, HSP70B and HSP70-5 expression, which might function in stabilizing denatured proteins and helping proteins to folding correctly in white clover under high temperature stress. In summary, exogenous Spd treatment improves the heat tolerance of white clover by altering endogenous PA and GABA content and metabolism, enhancing the antioxidant system and HSP expression and slowing leaf senescence related to an increase in Chl biosynthesis and a decrease in Chl degradation during heat stress.


Subject(s)
Medicago , Polyamines , Spermidine , Thermotolerance , gamma-Aminobutyric Acid , Gene Expression Regulation, Plant/drug effects , HSP70 Heat-Shock Proteins/genetics , Heat-Shock Response/drug effects , Medicago/drug effects , Medicago/metabolism , Oxidative Stress/drug effects , Polyamines/metabolism , Spermidine/pharmacology , Thermotolerance/drug effects , gamma-Aminobutyric Acid/metabolism
3.
Eur Rev Med Pharmacol Sci ; 22(19): 6500-6506, 2018 10.
Article in English | MEDLINE | ID: mdl-30338820

ABSTRACT

OBJECTIVE: The occurrence rate of delayed fracture healing in diabetes mellitus patients is high. Transforming growth factor ß1 (TGF-ß1) is an important regulatory factor in bone tissue repair and regeneration. However, TGF-ß1 expression and its function in diabetic patient fracture have not been fully elucidated. PATIENTS AND METHODS: Type 2 diabetes fracture patients (T2DM group), fracture patients without diabetes (non-T2DM group), and healthy volunteers (Control group) were selected for the research. TGF-ß1 expression in peripheral blood was detected by using enzyme-linked immunosorbent assay (ELISA). Osteoblast cell line, MG-63 cells, were randomly divided into Control, high glucose group, and TGF-ß1 group. TGF-ß1 expression was evaluated by using Real Time-PCR (RT-PCR). Cell proliferation was evaluated by using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Cell apoptosis activity was determined with caspase-3 activity and flow cytometry assay. The effect of TGF-ß1 on NF-κB was detected by using Western blot. RESULTS: TGF-ß1 was significantly reduced in patients of T2DM and non-T2DM groups compared with Control (p<0.05), while it was lower in T2DM group (p<0.05). TGF-ß1 expression was declined, cell proliferation was inhibited, caspase-3 activity was enhanced, cell apoptosis was elevated, and NF-κB expression was reduced in MG-63 cells of high glucose group compared to Control group (p<0.05). TGF-ß1 significantly promoted cell proliferation, suppressed caspase-3 activity, alleviated cell apoptosis, and elevated NF-κB expression in MG-63 cells compared with high glucose group (p<0.05). CONCLUSIONS: TGF-ß1 decreased in diabetes fracture patients. Up-regulation of TGF-ß1 regulates cell apoptosis and caspase-3 activity, and it facilitates osteoblasts proliferation.


Subject(s)
Blood Glucose/metabolism , Diabetes Mellitus, Type 2/blood , Fracture Healing , Fractures, Bone/metabolism , Osteoblasts/metabolism , Transforming Growth Factor beta1/blood , Adult , Aged , Apoptosis , Case-Control Studies , Caspase 3/metabolism , Cell Line, Tumor , Cell Proliferation , Female , Fracture Healing/drug effects , Fractures, Bone/pathology , Fractures, Bone/physiopathology , Humans , Male , Middle Aged , NF-kappa B/metabolism , Osteoblasts/drug effects , Osteoblasts/pathology , Signal Transduction , Time Factors , Transforming Growth Factor beta1/genetics , Transforming Growth Factor beta1/pharmacology
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