23 research outputs found
Correction of high amounts of astigmatism through orthokeratology. A case report
AbstractThe purpose of this case report is to introduce a method for a successful treatment of high astigmatism with a new orthokeratology design, called FOKX (Falco Kontaktlinsen, Switzerland). This novel toric orthokeratology contact lens design, the fitting approach and the performance of FOKX lenses will be illustrated in the form of a case report. Correcting astigmatism with orthokeratology offers a new perspective for all patients suffering astigmatism
Investigation of the effect of nicotinamide riboside on primary human bone-marrow derived mesenchymal stromal cells in vitro
INTRODUCTION: Mesenchymal stromal cells (MSC) have been identified as important cell-based therapy candidates for cartilage, bone and intervertebral disc diseases [1]. During prolonged in vitro expansion prior to administration, MSC become senescent which impairs their therapeutic potential [2]. Here, we aimed to investigate whether extracellular nicotinamide riboside (NR) is beneficial for MSC expansion with respect to delaying the senescence, improving cellular activity and growth kinetics.
METHODS: MSC were isolated from human bone marrow aspirates by gradient centrifugation of the bone marrow and subsequent expansion in α-MEM + 10% fetal bovine serum + 2.5 ng/ml bFGF2. The cytotoxicity of NR was measured at day 4 for 29 concentrations in a range from 5 nM to 4 mM. The long-term effects of NR were tested at concentrations of 10, 100 and 1,000 μM by measuring the population doubling level (PDL), relative confluency (real-time live-cell imaging with IncuCyte S3 ® ), mitochondrial activity by resazurin reduction, senescence-associated β-galactosidase assay (SA-β-gal) and NAD/NADH ratio.
RESULTS & DISCUSSION: MSC treated with 3 and 4 mM NR had a significantly higher mitochondrial activity at day 4 than the negative control (p=0.0027 and p<0.0001 respectively, N=3). However, in the weeks 3 to 8, cells treated with ≥100 μM NR died reaching a maximum PDL of 13.43 (N=4). In two donors, the experimental group with 10 μM NR reached a 2-fold higher PDL than the negative control. The relative confluency at passage (P) 2 after 6 days in culture was higher with 10 μM NR compared to the negative control (35.00 ± 9.29% and 26.19 ± 5.41% respectively, mean ± SD, N=2). The mitochondrial activity was significantly higher with 10 μM NR at P4, P8 and P10 (p<0.01, N=4). At all passages, the percentage of SA-β-gal positive cells was under 5%, except in the negative control medium at P11 (18.17% ± 18.18%, mean ± SD, N=1). All experimental groups treated with NR had a higher NAD/NADH ratio which exhibited a dose-dependent trend (N=1).
CONCLUSIONS: Extracellular NR elevated the intracellular NAD/NADH ratio. NR is not cytotoxic within 4 days of culture at concentrations up to 4 mM. Long-term culture with 10 μM NR improved the growth kinetics markedly in two donors.
ACKNOWLEDGEMENTS: Financial support was received by the Competence Center for Applied Biotechnology and Molecular Medicine (CABMM) start-up grant to BG.
REFERENCES
[1] Vedicherla S et al. J Orthop Res. 2017; 35(1):8-22
[2] Turinetto V et al. Int J Mol Sci. 2016;17(7):1164
Submucosal enteric neurons of the cavine distal colon are sensitive to hypoosmolar stimuli
Direct Evidence of 2-Cyano-2-Propoxy Radical Activity During AIBN-Based Oxidative Stress Testing in Acetonitrile–Water Solvent Systems
Prolonged retention after aggregation into secretory granules of human R183H-growth hormone (GH), a mutant that causes autosomal dominant GH deficiency type II
Human R183H-GH causes autosomal dominant GH deficiency type II. Because we show here that the mutant hormone is fully bioactive, we have sought to locate an impairment in its progress through the secretory pathway as assessed by pulse chase experiments. Newly synthesized wild-type and R183H-GH were stable when expressed transiently in AtT20 cells, and both formed equivalent amounts of Lubrol-insoluble aggregates within 40 min after synthesis. There was no evidence for intermolecular disulfide bond formation in aggregates of wild-type hormone or the R183H mutant. Both wildtype and R183H-GH were packaged into secretory granules, assessed by the ability of 1 mm BaCl2 to stimulate release and by immunocytochemistry. The mutant differed from wildtype hormone in its retention in the cells after packaging into secretory granules; 50% more R183H-GH than wild-type aggregates were retained in AtT20 cells 120 min after synthesis, and stimulated release of R183H-GH or a mixture of R183H-GH and wild-type that had been retained in the cell was reduced. The longer retention of R183H-GH aggregates indicates that a single point mutation in a protein contained in secretory granules affects the rate of secretory granule release