RESUMEN
Stem cells from human exfoliated deciduous teeth (SHEDs) have great potential to treat various dental-related diseases in regenerative medicine. They are usually maintained with 10% fetal bovine serum (FBS) in vitro. Modified platelet-rich plasma (mPRP) would be a safe alternative to 10% FBS during SHEDs culture. Therefore, our study aimed to compare the proliferation and differentiation of SHEDs cultured in mPRP and FBS medium to explore an optimal concentration of mPRP for SHEDs maintenance. Platelets were harvested by automatic blood cell analyzer and activated by repeated liquid nitrogen freezing and thawing. The platelet-related cytokines were examined and analyzed by ELISA. SHEDs were extracted and cultured with different concentrations of mPRP or 10% FBS medium. Alkaline phosphatase (ALP) activity was measured. Mineralization factors, RUNX2 and OCN, were measured by real-time PCR. SHEDs were characterized with mesenchymal stem cells (MSCs) markers including vimentin, CD44, and CD105. mPRP at different concentrations (2, 5, 10, and 20%) enhanced the growth of SHEDs. Moreover, mPRP significantly stimulated ALP activity and promoted expression of RUNX2 and OCN compared with 10% FBS. mPRP could efficiently facilitate proliferation and differentiation of SHEDs, and 2% mPRP would be an optimal substitute for 10% FBS during SHEDs expansion and differentiation in clinical scale manufacturing.
Asunto(s)
Proliferación Celular , Pulpa Dental/citología , Células Madre Mesenquimatosas/citología , Plasma Rico en Plaquetas , Diente Primario/citología , Fosfatasa Alcalina/antagonistas & inhibidores , Análisis de Varianza , Animales , Bovinos , Técnicas de Cultivo de Célula/métodos , Diferenciación Celular/fisiología , Proliferación Celular/fisiología , Células Cultivadas , Subunidad alfa 1 del Factor de Unión al Sitio Principal/análisis , Medios de Cultivo , Ensayo de Inmunoadsorción Enzimática , Humanos , Factor de Crecimiento Derivado de Plaquetas/análisis , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los Resultados , Factores de Tiempo , Factor de Crecimiento Transformador beta1/análisisRESUMEN
Knowledge of genetic diversity is important to assist breeders in the selection of parental materials and in the design of breeding programs. In this study, we genotyped 348 inbred tomato lines, representing vintage and contemporary fresh-market varieties, by using 52 single nucleotide polymorphisms (SNPs); 45 of these were found to be polymorphic. The average minor allele frequency and unbiased expected heterozygosity were 0.315 and 0.356, respectively. Population structure analysis revealed that contemporary germplasm could be distinctly divided into six subpopulations representing three market classes and breeding programs (pink, green, and red). Vintage germplasm could be separated into at least two subpopulations, and more admixtures were found in vintage lines than in contemporary lines. These findings indicate that contemporary inbred lines are more diversified than vintage inbred lines. AMOVA of vintage and contemporary lines was performed. A significant difference was found (P < 0.01), which explained 17.4% of the total genetic variance. Subsequently, we constructed a core collection using 45 polymorphic SNP markers. The data showed that all alleles were captured by only 2% of lines, indicating that more alleles, as well as rare alleles, could enable more variation to be captured in the core collection. These data allow us to discard redundant inbred tomato lines and to select elite inbred lines, which will accelerate the breeding process.
Asunto(s)
Polimorfismo de Nucleótido Simple , Semillas/genética , Solanum lycopersicum/genética , Frecuencia de los Genes , Genes de Plantas , Estudios de Asociación Genética , Marcadores Genéticos , Genotipo , Fitomejoramiento , Análisis de Secuencia de ADNRESUMEN
This study evaluated the inhibitory effects of spironolac-tone, a non-selective aldosterone receptor antagonist, on hypertension-induced myocardial fibrosis. Collagen I and III contents was detected in the myocardial tissue of spontaneously hypertensive rats (SHRs) after spironolactone administration. Twenty male SHRs were assigned to the spironolactone group or control group (N = 10 each); 7 Wistar-Kyoto rats (WKY) were also used. Spironolactone dissolved in ddH2O was administered via gavage at a dosage of 20 mg·kg(-1)·day(-1). Meanwhile, the control and WKY groups were administered equivalent volumes of ddH2O for 16 weeks. Western blotting was used to detect the contents of collagen I in myocardial tissue; observations were performed using polarizing microscopy, and the area integration and ratio of collagen I/III were subsequently calculated. Compared to the WKY group, col-lagen I synthesis was significantly higher in the control group (1.87 ± 0.2 vs 1.21 ± 0.7, P < 0.05). After 16 weeks of treatment, collagen I contents were significantly lower in the spironolactone group than in the control group (1.42 ± 0.05 vs 1.87 ± 0.2, P < 0.05). The ar-eas of collagen I and collagen I/III ratio were significantly smaller in the spironolactone group than in the control group (6400 ± 259 vs 12,019 ± 734 pixels, 15.64 ± 1.34 vs 20.8 ± 3.04 pixels, respec-tively; P < 0.05). However, there were no significant differences in the area of collagen III among the three groups. In conclusion, spi-ronolactone improves myocardial collagen deposition, preventing myocardial fibrosis in SHRs.
Asunto(s)
Miocardio/patología , Espironolactona/farmacología , Animales , Western Blotting , Recuento de Células , Colágeno/metabolismo , Fibrosis , Masculino , Microscopía de Polarización , Miocitos Cardíacos/citología , Miocitos Cardíacos/efectos de los fármacos , Ratas Endogámicas SHR , Ratas Endogámicas WKYRESUMEN
Toll-like receptor 3 (TLR3) recognizes double-stranded RNA, which is a molecular signature of viruses, and plays a pivotal role in host defense against viral invasion. Polymorphisms in the human TLR3 gene have been shown to affect the receptor function and to be associated with a variety of diseases, suggesting correlations between TLR3 polymorphisms and the disease resistance/susceptibility in pigs. In this study, 5 known non-synonymous single nucleotide polymorphisms (SNPs) in the coding sequences of the porcine TLR3 gene - c.800C>T (p.T267M), c.933A>G (p.I311M), c.1116A>T (p.K372N), c.2129C>G (p.T710S), and c.2160T>G (p.I720M) - were analyzed for their effect on receptor function in transiently transfected PK-15 cells by using a luciferase reporter assay. In addition, the distribution of SNP c.933A>G was analyzed among pig populations. SNP c.933A>G significantly decreased the response to poly(I:C) (P < 0.05), as represented by the weaker induction of firefly luciferase relative to that achieved by wild-type TLR3. SNP c.933A>G results in the alteration of conserved amino acids in the highly conserved segment of the 12th leucine repeat region and is conserved among TLR3 orthologs from fishes to primates. Moreover, together with the results of previous studies, the results of the present study revealed that SNP c.933A>G is found solely in local Chinese pig breeds. These results suggested that SNP c.933A>G plays a role in porcine disease resistance/susceptibility.