Engineering a family of synthetic splicing ribozymes

Controlling RNA splicing opens up possibilities for the synthetic biologist. The Tetrahymena ribozyme is a model group I self-splicing ribozyme that has been shown to be useful in synthetic circuits. To create additional splicing ribozymes that can function in synthetic circuits, we generated synthetic ribozyme variants by rationally mutating the Tetrahymena ribozyme. We present an alignment visualization for the ribozyme termed as structure information diagram that is similar to a sequence logo but with alignment data mapped on to secondary structure information. Using the alignment data and known biochemical information about the Tetrahymena ribozyme, we designed synthetic ribozymes with different primary sequences without altering the secondary structure. One synthetic ribozyme with 110 nt mutated retained 12% splicing efficiency in vivo. The results indicate that our biochemical understanding of the ribozyme is accurate enough to engineer a family of active splicing ribozymes with similar secondary structure but different primary sequences

Quantitative assessment of heavy metal effects on sperm function using computer‐aided sperm analysis and cytotoxicity assays

One known environmental risk factor impacting on human reproduction is heavy metal pollution. Although some metals (e.g., Cu, Se and Zn) have protective effects on the male reproductive system in low doses, heavy metals can accumulate to toxic levels and result in poor semen quality and decreased sperm function. We investigated the effect of CuSO4 and CdCl2 (10, 50, 100 and 250 μg/ml or 500 μg/ml) on human sperm motility and vitality by using computer‐aided sperm analysis (CASA) and two cytotoxicity assays (WST‐1 and XTT). Several sperm motility parameters were significantly reduced after 5 hr of exposure to the highest concentrations of CuSO4 (250 μg/ml) and CdCl2 (500 μg/ml). The WST‐1 assay also revealed significantly lower absorbance values for 50, 100 and 250 μg/ml CuSO4 and for 500 μg/ml CdCl2; however, no significant effect was seen with XTT. The calculated average IC50 value was 50.31± 4.34 μg/ml for CuSO4 and 392.32 ±76.79 μg/ml for CdCl2. The effects of these metals were confirmed with MgCl2, a positive control. This study provides threshold concentrations for the harmful effect of CuSO4 and CdCl2 on human spermatozoa and recommends the use of WST‐1 as vitality assay in future in vitro studies

The role of glucose lowering agents on restenosis after percutaneous coronary intervention in patients with diabetes mellitus

Abstract Introduction The prevalence of diabetes is increasing rapidly, and individuals with diabetes are at high risk for cardiovascular disorders. Subsequently the percentage of patients with diabetes subjected to revascularisation, i.e. either percutaneous coronary intervention (PCI) or coronary artery bypass grafting (CABG) also rises rapidly. The outcome of patients with diabetes after PCI is worse than for patients without diabetes. Restenosis is the main limiting factor of the long-term success of PCI. Although stents and antithrombotics improved outcome after PCI in both diabetics and non-diabetics, diabetics still have a worse prognosis. This leads to the suggestion that the restenosis mechanism in diabetics might be different from that in non-diabetics. Conclusion Several glucose lowering agents have been shown to influence the restenosis process and thus the outcome after PCI. Current data of especially metformin and thiazolidinediones indicate beneficial results as compared to insulin and sulfonylurea on restenosis. However, no large trials have been undertaken in which the effect of glucose lowering agents on restenosis is associated with improved outcome. The purpose of this review is to summarize the effect of diabetes and glucose lowering agents on restenosis.</p

The UPHILL study: A nutrition and lifestyle intervention to improve quality of life for patients with pulmonary arterial hypertension

Abstract The aim of the UPHILL study (a nutrition and lifestyle intervention in patients with pulmonary arterial hypertension [PAH]: effect on quality of life [QoL]), was to determine the effect of innovative nutritional interventions on adjustments in nutritional intake and QoL. In this study a group of prevalent PAH patients at a single center in Amsterdam (the Netherlands) was informed about healthy nutrition using a newly designed video e‐learning. They were subsequently instructed to follow a healthy diet during dietary intervention. Nutritional intake was assessed using a food frequency questionnaire (HELIUS) and QoL by the short‐form (SF)‐36 questionnaire. Nutritional parameters were determined in blood samples. Seventeen patients stable under treatment, who had been diagnosed with PAH 7.0 [3.0−14.0] years before, started and completed the intervention (2 males, 15 females; 45.35 ± 13.57 years). Since all patients in the intervention group made behavioral changes in nutritional intake, during study and follow‐up, nutritional and lifestyle adaptations persisted. Despite the fact that patients had already high mean scores at baseline for both mental (74.10 [60.51−84.25]) and physical QoL (66.46 [50.21−73.84]), scores improved further during e‐learning. Furthermore, patients who realized most nutritional adaptations, had the best improvement in QoL. This pilot study showed that e‐learning modules on nutrition provide an unique opportunity to change nutritional intake in PAH patients and by that improve QoL

Nutritional status in pulmonary arterial hypertension

Abstract Nutritional deficiencies have been described in patients with pulmonary arterial hypertension (PAH), such as in iron and vitamin D. However, an extensive description of vitamin and mineral status is lacking and until now there is no data on dietary intake in PAH patients. We analyzed blood samples and determined nutritional intake using a food frequency questionnaire (HELIUS) in a cohort of prevalent PAH patients at a single center in Amsterdam, the Netherlands. Quality of life (QoL) was assessed by the SF‐36 questionnaire. In total, 37 patients were included (6 males, 31 females; 48 ± 16 years). The dietary intake of sugar was above 25 g in 87% of the patients and fluid intake was above 1500 ml in 78% of the patients. Sodium intake was below 1800 mg in the majority (56%) of the patients. Sugar and fluid intake were linear related. We confirm previously observed deficiencies of iron and vitamin D in our study population. In addition, we observed a functional vitamin B12 deficiency in 29% of patients, which coincided with an increased expression of methylmalonic acid. 60% of patients had a low vitamin K1 status (<0.8 nmol/L). Finally, 40% of patients had selenium levels below <100 μg/L and low selenium levels associated with reduced vitality in these patients. Besides the known deficiencies in iron and vitamin D levels, we observed in a subset of patients signs of vitamin B12, vitamin K1 and selenium deficiencies. There is room for improving dietary intake. Future research aims to demonstrate the clinical importance and reveal the effect of nutritional interventions