VEGF, Ang-2 and SRIF associated abnormal postnatal development of the retinal capillary network in the Royal College of Surgeons rat

To examine retinal angiogenesis in the Royal College of Surgeons rat (RCS) serving as a model for ischemic proliferative retinopathies at morphological, proteomic and mRNA levels in order to evaluate the interplay of morphological and molecular changes in the course of the disease.Photoreceptor degeneration was confirmed by histological cross-sections and optical coherence tomography. The capillary retinal network was visualized in RCS rats aged between 14 and 45 days (P14–P45) by perfusion with high molecular weight fluorescein isothiocyanate-labeled dextran and compared with corresponding Sprague-Dawley rats. Vascular crosstalks within central areas to peripheral retinal eccentricities were analyzed. The expression of vascular growth-associated factors and their receptors in the course of the abnormal vascular development, namely vascular endothelial growth factor (VEGF), VEGF receptor subtype 1 (VEGF-R1) and -2 (VEGF-R2), somatostatin (SRIF), somatostatin receptor subtype 2 (Sstr-2), angiopoietin-2 (Ang-2) and tyrosine kinase with immunoglobulin-like and EGF-like domains 2 (Tie-2), was analyzed by immunohistochemistry, western blotting and quantitative real-time polymerase chain reaction.Underperfused areas without capillarization were found in the middle and peripheral retinal eccentricities of RCS rats until P29. Through the course of vascularization previously low perfused areas became completely perfused, but were characterized by significantly increased neovascularizations. Western blotting revealed specific expression of growth-associated factors and their receptors in the course of capillary development. VEGF was significantly increased until P29 in RCS rats, while SRIF was significantly upregulated at P21 and P29 at proteomic level in SD rats. At mRNA level Ang-2 was significantly upregulated in RCS rats at P29, VEGF-R1 and VEGF-R2 at P36 and SRIF at P36.Initial incomplete perfusion is followed by aberrant vessel formation. VEGF, SRIF, Ang-2 and their receptors are regulated at protein and mRNA levels, providing therapeutic possibilities for treating ischemic proliferative retinopathies in the course of the disease.     

Effects of crystallin-β-b2 on stressed RPE in vitro and in vivo

Background

Crystallins are thought to play a cytoprotective role in conditions of cellular stress. The aim of this study was to determine the effects of crystallin-β-b2 (cryβ-b2) and crystallin-β-b3 (cryβ-b3) on ARPE-19 cells in vitro and on the retinal pigment epithelium (RPE) in vivo.

Methods

The influence of cryβ-b2 and cryβ-b3 on the viability, proliferation and dying of ARPE-19 was measured by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium assay, bromo-2-deoxyuridine assay and life/death assay. The expressions of cryβ-b2, cryβ-b3, glial-derived neurotrophic factor (GDNF), and galectin-3 (Gal-3) in ARPE-19 cells were evaluated using immunohistochemistry (IHC), Western blotting (WB) and real-time-quantitative-PCR (qRT-PCR). To evaluate the response of cryβ-b2 and cryβ-b3 to stressed ARPE-19 cells, the cells were exposed to UV-light. In a rat model, cryβ-b2-expressing neural progenitor cells (cryβ-b2-NPCs) were injected intravitreally after retinal stress induced by optic nerve axotomy to examine whether they influence the RPE. Protein expression was examined 2 and 4 weeks postsurgery using IHC and WB.

Results

Detectable alterations of GDNF, and Gal-3 were found in ARPE-19 cells upon exposure to UV light. Adding the crystallins to the medium promoted proliferation and increased viability of ARPE-19 cells in vitro. The obtained data support the view that these crystallins possess epithelioprotective properties. Likewise, in vivo, intravitreally injected cryβ-b2 and transplanted cryβ-b2-NPCs protected RPE from indirectly induced stress.

Conclusions

The data suggest that the RPE response to retinal ganglion cell denegeration is mediated via crystallins, which may thus be used therapeutically.     

Noninvasive in vitro measurement of pig-blood d-glucose by using a microwave cavity sensor

We have developed an electromagnetic microwave cavity sensor based on the resonant frequency shift for real time measurement of the glycemia in pig blood. We could determine the concentration of d-glucose in pig blood in the range of 150-550mg/dl at the resonance frequency near 4.75GHz with a bandwidth of 300MHz. The change in the d-glucose concentration in blood brings microwave reflection coefficient S(11) changes of about 6.26dB and resonance frequency shifts of about 11.25MHz due to the electromagnetic interaction between the cavity resonator and the blood filled plastic tube inserted into the cavity. This proposed system provides a unique approach for real time noninvasive and contactless glucose monitoring.     

Implementing a decision-theoretic design in clinical trials: why and how?

This paper addresses two main questions: first, why should Bayesian and other innovative, data-dependent design models be put into practice and, secondly, given the past dearth of actual applications, how might one example of such a design be implemented in a genuine example trial? Clinical trials amalgamate theory, practice and ethics, but this last point has become relegated to the background, rather than taking often a more appropriate primary role. Trial practice has evolved but has its roots in R. A. Fisher's randomized agricultural field trials of the 1920s. Reasons for, and consequences of, this are discussed from an ethical standpoint, drawing on an under-used dichotomy introduced by French authors Lellouch and Schwartz (Int. Statist. Rev. 1971; 39:27-36). Plenty of ethically motivated designs for trials, including Bayesian designs have been proposed, but have found little application thus far. One reason for this is a lack of awareness of such alternative designs among trialists, while another reason is a lack of user-friendly software to allow study simulations. To encourage implementation, a new C++ program called 'Daniel' is introduced, offering much potential to assist the design of today's randomized controlled trials. Daniel evaluates a particular decision-theoretic method suitable for coping with either two or three Bernoulli response treatments with input features allowing user-specified choices of: patient horizon (number to be treated before and after the comparative stages of the trial); an arbitrary fixed trial truncation size (to allow ready comparison with traditional designs or to cope with practical constraints); anticipated success rates and a measure of their uncertainty (a matter ignored in standard power calculations); and clinically relevant, and irrelevant, differences in treatment effect sizes. Error probabilities and expected trial durations can be thoroughly explored via simulation, it being better by far to harm 'computer patients' instead of real ones. Suppose the objective in a clinical trial is to select between two treatments using a maximum horizon of 500 patients, when the truly superior treatment is expected to yield a 40 per cent success rate, but is believed to really range between 20 and 60 per cent. Simulation studies show that to detect a clinically relevant, absolute difference of 10 per cent between treatments, simulation studies show the decision-theoretic procedure would treat a mean 68 pairs of patients (SD 37) before correctly identifying the better treatment 96.7 per cent of the time, an error rate of 3.3 per cent. Having made a recommendation based on these patients, the remaining, on average 364 individuals, could either be given the indicated treatment, knowing its choice is optimal for the chosen horizon, or, alternatively, they could be entered into another, separate clinical trial. For comparison, a fixed sample size trial, with standard 5 per cent level of significance and 80 per cent power to detect a 10 per cent difference, requires treating over 700 patients in two groups, with the half allocated to inferior treatment considerably outnumbering the 68 expected under the decision-theoretic design, and the overall number simply too high for realistic application. In brief, the keys to answering the above 'why?' and 'how?' questions are ethics and software, respectively. Wider implications, both pros and cons, of implementing the particular method described will be discussed, with the overall conclusion that, where appropriate, clinical trials are now ready to undergo modernization from the agricultural age to the information age.     

Dynamic PEEP Study: A Non-invasive Diagnostic Exam to Assess for Effective PEEP in Children with Severe BPD

Objective

Tracheobronchomalacia (TBM) is common in neonates with bronchopulmonary dysplasia (BPD) and is associated with higher morbidity. This study evaluates the value of a CT protocol to assess the degree of TBM and gauge the adequacy of prescribed PEEP.

Study Design

Four infants with severe BPD on invasive mechanical ventilation underwent a chest CT protocol, including limited reduced-dose expiratory scans with varying PEEP levels.

Results

Baseline PEEP was adjusted in all subjects after performing the Dynamic PEEP CT. In two infants, the PEEP was increased due to significant TBM and in the other two without signs of TBM PEEP was decreased. The clinical course improved in all patients after adjusting PEEP.

Conclusion

A “Dynamic PEEP” study may be reliable and non-invasive imaging modality for the evaluation of adequate ventilator settings in infants with severe BPD who are not optimal candidates for bronchoscopy.

     

New versatile dual‐support pediatric heart pump

Mechanical circulatory support (MCS) devices for pediatric patients continue to lag in development behind those for adults. There is no heart pump with the design innovation to support dysfunctional states of heart failure and the anatomic heterogeneity of cardiac defects in pediatric patients. To address this unmet need, we are developing a versatile MCS technology with 2 separate blood pumps under 1 housing, whereby a centrifugal pump rotates around an axial pump. In this study, we advanced the design with a new inducer for the axial pump component and flat inlet volute for the centrifugal pump component. We conducted computational modeling of the design iterations, built prototypes, and tested their performance. The axial pump component was able to generate pressure rises of 1–112 mm Hg for 2–5 L/min at 10 000–14 000 RPM, and the centrifugal pump component produced pressure rises of 1–184 mm Hg for 2–5 L/min at 1750–3000 RPM. Shear stresses and blood damage estimations were less than  490 Pa and 0.5%, respectively. Axial and radial forces were also estimated to be less than 5 N for the axially and radially centered impellers. Data sets were repeatable, and data trends followed theoretical expectations. The new designs for the axial and centrifugal pumps enabled us to reduce the height of the pump while maintaining performance expectations. These findings support the continued development of this new medical device for pediatric patients.