XTT assay of ex vivo saliva biofilms to test antimicrobial influences

Objective: Many dental diseases are attributable to biofilms. The screening of antimicrobial substances, in particular, requires a high sample throughput and a realistic model, the evaluation must be as quick and as simple as possible. For this purpose, a colorimetric assay of the tetrazolium salt XTT (sodium 3'-[1-[(phenylamino)-carbony]-3,4-tetrazolium]-bis(4-methoxy-6-nitro)benzene-sulfonic acid hydrate) converted by saliva biofilms is recommended. Cleavage of XTT by dehydrogenase enzymes of metabolically active cells in biofilms yields a highly colored formazan product which is measured photometrically.Materials and method: The suitability of the XTT assay for detecting the vitality of ex vivo saliva biofilms was tested to determine the efficacy of chlorhexidine and ozone versus saliva biofilms grown on titanium discs.Results: The XTT method lends itself to testing the vitality of microorganisms in saliva biofilms. The sensitivity of the arrays requires a specific minimum number of pathogens, this number being different for planktonic bacteria and those occurring in biofilms. The antibacterial effect after treatment with chlorhexidine or ozone was measured by XTT conversion that was significantly reduced. The antimicrobial efficacy of 60 s 0.5% and 0.1% chlorhexidine treatment was equal and comparable with 60 s ozone treatment. Conclusion: The XTT assay is a suitable method to determine the vitality in saliva biofilms, permitting assessment of the efficacy of antimicrobial substances. Its quick and easy applicability renders it especially suitable for screening.     

Feasibility of dynamic CT-based adenosine stress myocardial perfusion imaging to detect and differentiate ischemic and infarcted myocardium in an large experimental porcine animal model

The purpose of the study is feasibility of dynamic CT perfusion imaging to detect and differentiate ischemic and infarcted myocardium in a large porcine model. 12 Country pigs completed either implantation of a 75 % luminal coronary stenosis in the left anterior descending coronary artery simulating ischemia or balloon-occlusion inducing infarction. Dynamic CT-perfusion imaging (100 kV, 300 mAs), fluorescent microspheres, and histopathology were performed in all models. CT based myocardial blood flow (MBF_CT), blood volume (MBV_CT) and transit constant (K_trans), as well as microsphere’s based myocardial blood flow (MBF_Mic) were derived for each myocardial segment. According to histopathology or microsphere measurements, 20 myocardial segments were classified as infarcted and 23 were ischemic (12 and 14 %, respectively). Across all perfusion states, MBF_CT strongly predicted MBF_Mic (β 0.88 ± 0.12, p < 0.0001). MBF_CT, MBV_CT, and K_trans were significantly lower in ischemic/infarcted when compared to reference myocardium (all p < 0.01). Relative differences of all CT parameters between affected and non-affected myocardium were higher for infarcted when compared to ischemic segments under rest (48.4 vs. 22.6 % and 46.1 vs. 22.9 % for MBF_CT, MBV_CT, respectively). Under stress, MBF_CT was significantly lower in infarcted than in ischemic myocardium (67.8 ± 26 vs. 88.2 ± 22 ml/100 ml/min, p = 0.002). In a large animal model, CT-derived parameters of myocardial perfusion may enable detection and differentiation of ischemic and infarcted myocardium.     

Genetically determined heterogeneity in hemoglobin scavenging and susceptibility to diabetic cardiovascular disease

A major function of haptoglobin (Hp) is to bind hemoglobin (Hb) to form a stable Hp-Hb complex and thereby prevent Hb-induced oxidative tissue damage. Clearance of the Hp-Hb complex can be mediated by the monocyte/macrophage scavenger receptor CD163. We recently demonstrated that diabetic individuals homozygous for the Hp 2 allele (Hp 2-2) were at 500% greater risk of cardiovascular disease (CVD) compared with diabetic individuals homozygous for the Hp 1 allele (Hp 1-1). No differences in risk by Hp type were seen in individuals without diabetes. To understand the relationship between the Hp polymorphism and diabetic CVD, we sought to identify differences in antioxidant and scavenging functions between the Hp types and to determine how these functions were modified in diabetes. The scavenging function of Hp was assessed using rhodamine-tagged and 125I-Hp in cell lines stably transfected with CD163 and in macrophages expressing endogenous CD163. We found that the rate of clearance of Hp 1-1-Hb by CD163 is markedly greater than that of Hp 2-2-Hb. Diabetes is associated with an increase in the nonenzymatic glycosylation of serum proteins, including Hb. The antioxidant function of Hp was assessed with glycosylated and nonglycosylated Hb. We identified a severe impairment in the ability of Hp to prevent oxidation mediated by glycosylated Hb. We propose that the specific interaction between diabetes, CVD, and Hp genotype is the result of the heightened urgency of rapidly clearing glycosylated Hb-Hp complexes from the subendothelial space before they can oxidatively modify low-density lipoprotein to atherogenic oxidized low-density lipoprotein.     

Trends in In-Hospital Mortality,Length of Stay,Nonroutine Discharge,and Cost Among End-Stage Renal Disease Patients on Dialysis Hospitalized With Heart Failure (2001–2014)

BackgroundHeart failure (HF) is a common cause of morbidity and mortality among end-stage renal disease (ESRD) patients on dialysis. We aimed to assess the trends and outcomes in primary and secondary HF hospitalizations among ESRD patients with the use of a nationally representative database.Methods and ResultsWe analyzed data from the National Inpatient Sample and the US Census Bureau to calculate annual national rates of in-hospital mortality, length of stay, disposition with a focus on nonroutine discharge (discharge to a health care facility rather than to home), and adjusted median cost among patients with ESRD on dialysis with primary or secondary HF admissions from 2001 to 2014. An estimated 812,090 primary and 2,887,432 secondary HF admissions occurred from 2001 to 2014. The prevalence of comorbidities increased during the study period. Primary HF admission rates increased from 2001 to 2006 and decreased from 2007 to 2014, whereas secondary HF admissions increased significantly during the study period (P < .001). We found statistically significant declines of primary and secondary admission in-hospital mortality, with annual percentage changes of ?3.1% and ?2.6% respectively (P < .001 for both). In addition, the lengths of stay decreased significantly for primary and secondary HF admissions (P < .001 for both). However, nonroutine discharges increased significantly for both. Subgroup analysis showed higher in-hospital mortality for men, patients >65 years of age, whites, and those on peritoneal dialysis. The cost of hospitalization did not change significantly for primary and secondary HF admissions.ConclusionAmong ESRD patients on dialysis with primary or secondary HF admission diagnosis, comorbidity prevalence increased but in-hospital mortality and length of stay decreased significantly from 2001 to 2014.     

Myocardial gene transfer by selective pressure-regulated retroinfusion of coronary veins: comparison with surgical and percutaneous intramyocardial gene delivery.

OBJECTIVES: We sought to study adenoviral gene delivery using percutaneous selective pressure-regulated retroinfusion and to compare it directly with surgical and percutaneous intramyocardial delivery (PIMD) for the first time. BACKGROUND: Intramyocardial delivery (IMD) has been recommended to be the preferred gene delivery strategy so far. However, surgical and percutaneous intramyocardial injection lead to incomplete retention of the injected viral vectors and to limited spatial myocardial distribution. Percutaneous selective pressure-regulated retroinfusion of the coronary veins was developed recently to provide an effective and more homogenous regional myocardial gene transfer. METHODS: In 15 pigs, adenoviral vectors (Ad2-CMV beta-galactosidase [beta-gal] 5 x 10(9) pfu) were applied via surgical IMD (n = 5), PIMD (n = 5), and selective pressure-regulated retroinfusion (n = 5). Seven days after gene transfer, myocardial beta-gal expression was measured by ELISA. RESULTS: Selective retroinfusion into the anterior cardiac vein substantially increased reporter gene expression (1,039 +/- 79 pg beta-gal/mg protein) in the targeted left anterior descending coronary artery territory when compared with surgical (448 +/- 127, p < 0.05) and PIMD (842 +/- 145, p < 0.05). Both IMD approaches showed an inhomogenous beta-gal expression, particularly along the injection sites, while retroinfusion resulted in a more homogenous transmural gene expression. CONCLUSIONS: Percutaneous selective pressure-regulated retroinfusion compares favorably with surgical and percutaneous intramyocardial injection techniques by providing a more homogenous and even more efficient adenoviral gene delivery.