RESUMO
Eleven neo-clerodane diterpenoids (1-11) including the new analogues 1, 2, and 10, and 3',5,6,7-tetrahydroxy-4'-methoxyflavone (12) were isolated from the aerial parts of Salvia polystachya. Polystachyne G (1) and 15-epi-polystachyne G (2) were isolated as an epimeric mixture, containing a 5-hydroxyfuran-2(5H)-one unit in the side chain at C-12 of the neo-clerodane framework. Polystachyne H (10) contains a 1(10),2-diene moiety and a tertiary C-4 hydroxy group. The structures of these compounds were established by analysis of their NMR spectroscopic and MS spectrometric data. The absolute configurations of compounds 3, 4, and 10 were determined through single-crystal X-ray diffraction analysis. The antibacterial, antifungal, and phytotoxic activities of the diterpenoids were determined. In addition, the stimulatory effect of the expression of extracellular matrix components of nine of the isolates (1-8 and 11) was assayed. Compounds 1-4, 8, and 11 increased the expression of the genes codifying for type I, type III, and type V collagens and for elastin.
Assuntos
Diterpenos Clerodânicos/isolamento & purificação , Diterpenos Clerodânicos/farmacologia , Matriz Extracelular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Salvia/química , Bacillus/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Colágeno/efeitos dos fármacos , Colágeno/genética , Cristalografia por Raios X , Diterpenos Clerodânicos/química , Elastina/efeitos dos fármacos , Elastina/genética , Escherichia coli/efeitos dos fármacos , Flavonoides/química , Flores/química , Humanos , México , Testes de Sensibilidade Microbiana , Conformação Molecular , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Folhas de Planta/química , Reação em Cadeia da PolimeraseRESUMO
Birth weight is an early predictor for metabolic diseases and microRNAs (miRNAs) are proposed as fetal programming participants. To evaluate the use of dried blood spots (DBS) on newborn screening cards (NSC) as a source of analyzable miRNAs, we optimized a commercial protocol to recover total miRNA from normal birth weight (NBW, n = 17-20), low birth weight (LBW, n = 17-20) and high birth weight (macrosomia, n = 17-20) newborns and analyzed the relative expression of selected miRNAs by stem-loop RT-qPCR. The possible role of miRNAs on the fetal programming of metabolic diseases was explored by bioinformatic tools. The optimized extraction of RNA resulted in a 1.2-fold enrichment of miRNAs respect to the commercial kit. miR-33b and miR-375 were overexpressed in macrosomia 9.8-fold (p < 0.001) and 1.7-fold, (p < 0.05), respectively and miR-454-3p was overexpressed in both LBW and macrosomia (19.7-fold, p < 0.001 and 10.8-fold, p < 0.001, respectively), as compared to NBW. Potential target genes for these miRNAs are associated to cyclic-guanosine monophosphate (cGMP)-dependent protein kinase (PKG), mitogen-activated protein kinase (MAPK), type 2 diabetes, transforming growth factor-ß (TGF-ß)and Forkhead box O protein (FoxO) pathways. In summary, we improved a protocol for analyzing miRNAs from NSC and provide the first evidence that birth weight modifies the expression of miRNAs associated to adult metabolic dysfunctions. Our work suggests archived NSC are an invaluable resource in the search for fetal programming biomarkers.
Assuntos
Peso ao Nascer/fisiologia , MicroRNAs/fisiologia , Peso ao Nascer/genética , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Feminino , Macrossomia Fetal/genética , Macrossomia Fetal/metabolismo , Perfilação da Expressão Gênica , Humanos , Recém-Nascido , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Triagem Neonatal/métodosRESUMO
Murine 3T3 cell lines constitute a standard model system for in vitro study of mammalian adipogenesis although they do not faithfully reflect the biology of the human adipose cells. Several human adipose cell lines and strains have been used to recapitulate human adipogenesis in vitro, but to date there is no generally accepted in vitro model for human adipogenesis. We obtained a clonal strain of human subcutaneous adipose stromal cells, IPI-SA3-C4, and characterized its utility as an in vitro model for human subcutaneous adipogenesis. IPI-SA3-C4 cells showed a high proliferative potential for at least 30 serial passages, reached 70 cumulative population doublings and exhibited a population doubling time of 47 h and colony forming efficiency of 12% at the 57th cumulative population doublings. IPI-SA3-C4 cells remained diploid (46XY) even at the 56th cumulative population doublings and expressed the pluripotency markers POU5F1, NANOG, KLF4, and MYC even at 50th cumulative population doublings. Under specific culture conditions, IPI-SA3-C4 cells displayed cellular hallmarks and molecular markers of adipogenic, osteogenic, and chondrogenic lineages and showed adipogenic capacity even at the 66th cumulative population doublings. These characteristics show IPI-SA3-C4 cells as a promising potential model for human subcutaneous adipogenesis in vitro.