Peptides derived from the copper‐binding region of lysyl oxidase exhibit antiangiogeneic properties by inhibiting enzyme activity: an in vitro study

Despite the rigorous research on abnormal angiogenesis, there is a persistent need for the development of new and efficient therapies against angiogenesis‐related diseases. The role of Lysyl oxidase (LOX) in angiogenesis and cancer has been established in prior studies. Copper is known to induce the synthesis of LOX, and hence regulates its activity. Hypoxia‐induced metastasis is dependent on LOX expression and activity. It has been believed that the inhibition of LOX would be a therapeutic strategy to inhibit angiogenesis. To explore this, we designed peptides (M peptides) from the copper‐binding region of LOX and hypothesized them to modulate LOX. The peptides were characterized, and their copper‐binding ability was confirmed by mass spectrometry. The M peptides were found to reduce the levels of intracellular copper when the cells were co‐treated with copper. The peptides showed promising effect on aortic LOX, recombinant human LOX and LOX produced by human umbilical vein endothelial cells (HUVECs). The study also explores the effect of these peptides on copper and hypoxia‐stimulated angiogenic response in HUVECs. It was found that the M peptides inhibited copper/hypoxia‐induced LOX activity and inhibited stimulated HUVEC tube formation and migration. This clearly indicated the potential of M peptides in inhibiting angiogenesis, highlighting their role in the formulation of drugs for the same. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.     

Analysis of physical parameters and determination of inflection point for Flattening Filter Free beams in medical linear accelerator

Background

Medical Linear accelerators manufactured without flattening filters are increasing popular in recent days. The removal of flattening filter results in increased dose rate, reduced mean energy, reduction in head leakage and lateral scattering, which have shown advantageous when used for special treatment procedures.

Aim

This study aims to analyze physical parameters of FFF beams and to determine the inflection point for standardizing the beam flatness and penumbra.

Materials and methods

The beam profiles and depth dose patterns were measured using Radiation Field Analyzer (RFA) with 0.13 cc cylindrical ion chamber. The beam energy characteristics, head scatter factor (Sc) were obtained for 6FFF and 10FFF beams and compared with 6 MV and 10 MV photons, respectively. The symmetry and stability of unflattened regions were also analyzed. In addition, the study proposes a simple physical concept for obtaining inflection point for FFF beams and results were compared using the Akima spline interpolation method. The inflection point was used to determine the field size and penumbra of FFF beams.

Results

The Sc varied from 0.922 to 1.044 for 6FFF and from 0.913 to 1.044 for 10FFF with field sizes from 3 cm × 3 cm to 40 cm × 40 cm which is much less than FF beams. The obtained value of field size and penumbra for both simple physical concept and Akima spline interpolation methods is within the ±1.0 mm for the field size and ±2 mm penumbra. The results indicate that FFF beams reduce Sc compared with FF beams due to the absence of a flattening filter.

Conclusion

The proposed simple method to find field size and penumbra using inflection point can be accepted as it is closely approximated to mathematical results. Stability of these parameters was ascertained by repeated measurements and the study indicates good stability for FFF beam similar to that of FF beams.     

Sinapic Acid Prevents Hypertension and Cardiovascular Remodeling in Pharmacological Model of Nitric Oxide Inhibited Rats

Objectives

Hypertensive heart disease is a constellation of abnormalities that includes cardiac fibrosis in response to elevated blood pressure, systolic and diastolic dysfunction. The present study was undertaken to examine the effect of sinapic acid on high blood pressure and cardiovascular remodeling.

Methods

An experimental hypertensive animal model was induced by L-NAME intake on rats. Sinapic acid (SA) was orally administered at a dose of 10, 20 and 40 mg/kg body weight (b.w.). Blood pressure was measured by tail cuff plethysmography system. Cardiac and vascular function was evaluated by Langendorff isolated heart system and organ bath studies, respectively. Fibrotic remodeling of heart and aorta was assessed by histopathologic analyses. Oxidative stress was measured by biochemical assays. mRNA and protein expressions were assessed by RT-qPCR and western blot, respectively. In order to confirm the protective role of SA on endothelial cells through its antioxidant property, we have utilized the in vitro model of H₂O₂-induced oxidative stress in EA.hy926 endothelial cells.

Results

Rats with hypertension showed elevated blood pressure, declined myocardial performance associated with myocardial hypertrophy and fibrosis, diminished vascular response, nitric oxide (NO) metabolites level, elevated markers of oxidative stress (TBARS, LOOH), ACE activity, depleted antioxidant system (SOD, CAT, GPx, reduced GSH), aberrant expression of TGF-β, β-MHC, eNOS mRNAs and eNOS protein. Remarkably, SA attenuated high blood pressure, myocardial, vascular dysfunction, cardiac fibrosis, oxidative stress and ACE activity. Level of NO metabolites, antioxidant system, and altered gene expression were also repaired by SA treatment. Results of in vitro study showed that, SA protects endothelial cells from oxidative stress and enhance the production of NO in a concentration dependent manner.

Conclusions

Taken together, these results suggest that SA may have beneficial role in the treatment of hypertensive heart disease by attenuating fibrosis and oxidative stress through its antioxidant potential.     

Leukocyte extravasation: an immunoregulatory role for alpha-L-fucosidase?

Fucosylated oligosaccharides and glycoconjugates have been implicated in several biological events, including the cell-cell adhesion processes that mediate inflammation. Alpha-L-fucosidase (ALF) is an exoglycosidase that is involved in the hydrolytic degradation of alpha-L-fucose from glycoconjugates. In this study, we investigated the potential role of ALF in regulation of leukocyte migration. Measurement of transendothelial migration in response to CCL5 demonstrated that pretreatment of monocytic cells with ALF reduced migration (p = 0.0004) to a greater extent than treatment of the endothelial monolayer (p = 0.0374). Treatment with ALF significantly reduced the adhesion of monocytic cells to immobilized P-selectin.Fc. A murine model of experimental autoimmune uveitis was then used to show that treatment of splenic cells with ALF produced an 8.6-fold decrease in rolling and a 3.2-fold decrease in cell migration across the retinal vasculature. Further in vitro studies demonstrated that treatment of monocytes with the chemokines CCL3 or CCL5 increased the level of mRNA encoding ALF; this was accompanied by the detection of significant increases in both the 51- and 56-kDa components of ALF by Western blotting. Treatment of monocytic cells with ALF for 2 h significantly reduced the cell surface expression of CD31, with a further decrease in expression observed after 5 h (p = 0.002). Thus, CD31 and fucosylated ligands of P-selectin seem to be the candidates through which ALF mediates its effect in vitro. These data identify a previously unrecognized immunoregulatory role for ALF in late stages of inflammation.     

Changes in Two Point Discrimination and the law of mobility in Diabetes Mellitus patients

Background

Though retrograde neuronal death and vascular insufficiency have been well established in plegics following intracerebral hemorrhage, the effects of plegia on arterial nervorums of peripheral nerves have not been reported. In this study, the histopathological effects of the intracerebral hemorrhage on the dorsal root ganglions and sciatic nerves via affecting the arterial nervorums were investigated.

Methods

This study was conducted on 13 male hybrid rabbits. Three animals were taken as control group and did not undergo surgery. The remaining 10 subjects were anesthetized and were injected with 0.50 ml of autologous blood into their right sensory-motor region. All rabbits were followed-up for two months and then sacrificed. Endothelial cell numbers and volume values were estimated a three dimensionally created standardized arterial nervorums model of lumbar 3. Neuron numbers of dorsal root ganglions, and axon numbers in the lumbar 3 nerve root and volume values of arterial nervorums were examined histopathologically. The results were analyzed by using a Mann-Whitney-U test.

Results

Left hemiplegia developed in 8 animals. On the hemiplegic side, degenerative vascular changes and volume reduction in the arterial nervorums of the sciatic nerves, neuronal injury in the dorsal root ganglions, and axonal injury in the lumbar 3 were detected. Statistical analyses showed a significant correlation between the normal or nonplegic sides and plegic sides in terms of the neurodegeneration in the dorsal root ganglions (p < 0.005), axonal degeneration in the lumbar 3 nerve roots (p < 0.005), endothelial cell degeneration in the arterial nervorums (p < 0.001), and volume reduction in the arterial nervorums (p < 0.001).

Conclusion

Intracerebral hemorrhage resulted in neurodegeneration in the dorsal root ganglion and axonolysis in the sciatic nerves, endothelial injury, and volume reduction of the arterial nervorums in the sciatic nerves. The interruption of the neural network connection in the walls of the arterial nervorums in the sciatic nerves may be responsible for circulation disorders of the arterial nervorums, and arterial nervorums degeneration could result in sciatic nerves injury.     

A naphthalocyanine-based EPR probe for localized measurements of tissue oxygenation.

A new electron paramagnetic resonance (EPR) oximetry probe, based on a naphthalocyanine macrocycle, is reported to exhibit high oxygen sensitivity and favorable EPR characteristics for biological applications. The free radical probe, lithium naphthalocyanine (LiNc), is synthesized as fine microcrystalline powder with particle size less than 1 microm and high spin density. It exhibits a single sharp EPR peak, whose width varies linearly with oxygen partial pressure (pO2). The EPR spectrum is nonsaturable at typical microwave power levels (< 25 mW at X-band). These unique characteristics make this probe ideal for measuring oxygen concentration in biological tissues, in vivo. The peak-to-peak width under anoxic conditions is 0.51 G (at X-band), and it increases linearly with increase in oxygen partial pressure and reaches 26.0 G for 100% oxygen (760 mmHg), showing an oxygen sensitivity of 34 mG/mmHg. The probe responds to changes in pO2 quickly and reproducibly, thus enabling dynamic measurements of regional oxygenation in real time. The application of this probe for oximetry is demonstrated in an in vivo biological system. The changes in pO2 were monitored in the leg muscle tissue of a living mouse breathing room air and carbogen (95% oxygen + 5% CO2), alternatively. The mean pO2 measured with this probe in muscle tissues was consistent with values reported previously using other methods. Overall, the probe shows very desirable characteristics for localized measurements of tissue oxygenation.