The increasing frequency of penicillin-resistant pneumococcus continues to be of concern throughout the world. Newer fluoroquinolone antibiotics, such as levofloxacin, have shown enhanced in vitro activity against Streptococcus pneumoniae. In this study, the bactericidal characteristics and pharmacodynamic profiles of levofloxacin, ciprofloxacin, and ampicillin against four isolates of S. pneumoniae were compared by using an in vitro model of infection. Standard antibiotic dosing regimens which simulated the pharmacokinetic profile observed in humans were used. Control and treatment models were sampled for bacterial CFU per milliliter over the duration of each 24-or 48-h experiment. In addition, treatment models were sampled for MIC determinations and drug concentration. Regrowth of all isolates as well as an increase in MICs throughout the study period was observed in the ciprofloxacin experiments. A limited amount of regrowth was noted during levofloxacin therapy for one isolate; however, no change in MIC was detected for any isolate. Ampicillin showed rapid and sustained bactericidal activity against all isolates. In this study, ratios of effective fluoroquinolone area under the concentration-time curve (AUC):MIC values ranged from 30 to 55. Levofloxacin, owing to its larger AUC 0-24 values, has excellent and sustained activity against different pneumococcal strains superior to that of ciprofloxacin.

It is now known that over-consumption of caffeine by pregnant mothers could have detrimental effects on normal fetal development. However, it remains obscure how caffeine's harmful effect impacts directly or indirectly on the developing embryo/fetus through damaging placenta development. In this study, we demonstrated the morphological similarities between the yolk sac and chorioallantoic membranes (CAM) of chick embryos and the villi of the mammalian placenta. Using the chick yolk sac and the CAM as a model, we found that 5-15 µmol per egg of caffeine exposure inhibited angiogenesis. Under the same condition, cell proliferation in extraembryonic mesoderm was reduced while apoptosis was enhanced. Semi-quantitative RT-PCR analysis revealed that caffeine treatment down-regulated VEGF, VEGFR2, PIGF, IGF2 and NRP1 expression, but up-regulated Ang1 and Ang2 expression. We performed in situ hybridization to show VE-cadherin expression and as to demonstrate the blood vessels in the CAM and yolk sac membranes. This distribution of the VE-cadherin(+) blood vessels was determined to be reduced after caffeine treatment. Furthermore, MDA activity was induced after caffeine exposure, but GSH-PX activity was inhibited after caffeine exposure; SOD activity was unchanged as compared with the control. In summary, our results suggest that caffeine exposure could negatively impact on angiogenesis in the chick yolk sac and CAM by targeting angiogenesis-related genes. Some of these genes are also involved in regulating excess ROS generation. The results implied that the negative impact of caffeine on fetal development was partly attributed to impaired placental angiogenesis.

Excess alcohol consumption during pregnancy has been acknowledged to increase the incidence of congenital disorders, especially the cardiovascular system. However, the mechanism involved in ethanol-induced cardiac malformation in prenatal fetus is still unknown. We demonstrated that ethanol exposure during gastrulation in the chick embryo increased the incidence of cardia bifida. Previously, we reported that autophagy was involved in heart tube formation. In this context, we demonstrated that ethanol exposure increased ATG7 and LC3 expression. mTOR was found to be inhibited by ethanol exposure. We activated autophagy using exogenous rapamycin (RAPA) and observed that it induced cardiac bifida and increased GATA5 expression. RAPA beads implantation experiments revealed that RAPA restricted ventricular myosin heavy chain (VMHC) expression. In vitro explant cultures of anterior primitive streak demonstrated that both ethanol and RAPA treatments could reduce cell differentiation and the spontaneous beating of cardiac precursor cells. In addition, the bead experiments showed that RAPA inhibited GATA5 expression during heart tube formation. Semiquantitative RT-PCR analysis indicated that BMP2 expression was increased while GATA4 expression was suppressed. In the embryos exposed to excess ethanol, BMP2, GATA4 and FGF8 expression was repressed. These genes are associated with cardiomyocyte differentiation, while heart tube fusion is associated with increased Wnt3a but reduced VEGF and Slit2 expression. Furthermore, the ethanol exposure also caused the production of excess ROS, which might damage the cardiac precursor cells of developing embryos. In sum, our results revealed that disrupting autophagy and excess ROS generation are responsible for inducing abnormal cardiogenesis in ethanol-treated chick embryos.

People have known that autophagy plays a very important role in many physiological and pathological events. But the role of autophagy on embryonic angiogenesis still remains obscure. In this study, we demonstrated that Atg7, Atg8 and Beclin1 were expressed in the plexus vessels of angiogenesis at chick yolk sac membrane and chorioallantoic membrane. Interfering in autophagy with autophagy inducer or inhibitor could restrict the angiogenesis in vivo, which might be driven by the disorder of angiogenesisrelated gene expressions, and also lead to embryonic hemorrhage, which was due to imperfection cell junctions in endothelial cells including abnormal expressions of tight junction, adheren junction and desmosome genes. Using HUVECs, we revealed that cell viability and migration ability changed with the alteration of cell autophagy exposed to RAPA or 3-MA. Interestingly, tube formation assay showed that HUVECs ability of tube formation altered with the change of Atg5, Atg7 and Atg8 manipulated by the transfection of their corresponding siRNA or plasmids. Moreover, the lost cell polarity labeled by F-actin and the absenced b-catenin in RAPA-treated and 3-MA-treated cell membrane implied intracellular cytoskeleton alteration was induced by the activation and depression of autophagy. Taken together, our current experimental data reveal that autophagy is really involved in regulating angiogenesis during embryo development.

Extracellular matrix (ECM) accumulation is considered an important pathological feature of diabetic kidney disease (DKD). Histone deacetylase (HDAC) inhibitors protect against kidney injury. However, the potential mechanisms of HDACs in DKD are still largely unknown. Here, we describe a novel feedback loop composed of HDAC2 and miR-205 that regulates ECM production in tubular epithelial cells in individuals with DKD. We found that HDAC2 mRNA expression in peripheral blood was markedly higher in patients with DKD than in patients with diabetes. Nuclear HDAC2 protein expression was increased in TGFβ1-stimulated tubular epithelial cells and db/db mice. We also found that miR-205 was regulated by HDAC2 and downregulated in TGFβ1-treated HK2 cells and db/db mice. In addition, HDAC2 reduced histone H3K9 acetylation in the miR-205 promoter region to inhibit its promoter activity and subsequently suppressed miR-205 expression through an SP1-mediated pathway. Furthermore, miR-205 directly targeted HDAC2 and inhibited HDAC2 expression. Intriguingly, miR-205 also regulated its own transcription by inhibiting HDAC2 and increasing histone H3K9 acetylation in its promoter, forming a feedback regulatory loop. Additionally, the miR-205 agonist attenuated ECM production in HK2 cells and renal interstitial fibrosis in db/db mice. In conclusion, the HDAC2/SP1/miR-205 feedback loop may be crucial for the pathogenesis of DKD.