Analysis of ischemia-reperfusion injury in a microcirculatory model of pressure ulcers

The aim of this study was to establish a pressure ulcer model that visualizes the microcirculation, and to examine the participation of ischemia-reperfusion injury in the pathophysiology of pressure ulcers. An original system composed of a new skin fold chamber and compression device allowed loading quantitative vertical stress to the skin. An intravital microscopic technique enabled direct visualization of the microcirculation in the physiological condition and in response to pressure application. To estimate the effect of ischemia-reperfusion injury, animals were divided into two groups: the compression-release group (n = 8), in which the animals received four cycles of compression-release which consisted of 2 hours of compression followed by 1 hour of pressure release; and the compression alone group (n = 8) in which the animals underwent continuous compression for 8 hours. Functional capillary density was quantified before the compression procedure and on day 1 (35 hours) after the first evaluation. The cyclic compression-release procedure significantly decreased functional capillary density as compared to continuous compression, indicating that in our experimental setting repetition of ischemia-reperfusion cycle more severely damaged the microcirculation than single prolonged ischemic insult. This finding supports the significant contribution of ischemia-reperfusion injury to the pathophysiology of pressure ulcers at the level of dynamic in vivo microcirculation.     

Efficacy of Crocus sativus L. on reduction of cadmium‐induced toxicity on spermatogenesis in adult rats

Cadmium is a toxic heavy metal element, which probably cause infertility by impairment in spermatogenesis. The present work aimed (i) to study the toxic effect of cadmium on spermatogenesis in rat, as well as (ii) the protective effect of Crocus sativus L. on cadmium‐intoxicated rats. Cadmium chloride was administered intraperitoneally during 16 days at intervals of 48 h between subsequent treatments. Crocus sativus L. was pre‐treated in both of control and cadmium‐injected rats. Animals were sacrificed on day 17 after the first treatment. The left cauda epididymis was removed and immediately immersed into Hank's balanced salt solution for the evaluation of sperm count and viability, and left testis was fixed in 10% formalin for histological evaluation. Following contamination with cadmium, a decrease was observed in the number and viability of cauda epididymis sperm, which were increased by Crocus sativus L. pre‐treatment (P < 0.05). In addition, cadmium decreased both cell proliferation and Johnsen Scores in the seminiferous tubules, which were reversed by Crocus sativus pre‐treatment (P < 0.05). Furthermore, cadmium‐induced decrease in the amount of free serum testosterone as well as an increase in lipid peroxidation activity in the testicular tissue was reversed by Crocus sativus L. (P < 0.05). These findings may support the concept that Crocus sativus L. can improve the cadmium toxicity on spermatogenesis.     

AlphaV beta3 (αvβ3) integrin inhibition reduces leukocyte–endothelium interaction in a pressure-induced reperfusion model

The leukocyte-endothelium interaction is known to contribute to reperfusion injury, which is considered to participate in the pathophysiology of pressure ulcers, and integrin alphaV beta3 (alphavbeta3) has been shown to mediate the processes of cellular adhesion in various types of cells. This study aims to clarify leukocyte behavior in our original microcirculatory pressure-induced reperfusion model, which can visualize the microcirculation in vivo. We also estimated the effect of alphavbeta3 integrin inhibition on the reduction of the leukocyte-endothelium interaction. Mice with dorsal skinfold chambers were divided into three groups: the baseline group (n=6), in which animals received no compression; the compression-reperfusion group (n=6), in which animals underwent 2-hour compression of the dorsal skin, followed by release, and the inhibitor-treated group (n=7), in which an alphavbeta3 inhibitor, CP4715, was administered in addition to the compression-release procedure. Staining with rhodamine 6G quantitatively visualized leukocyte behavior under the intravital fluorescent microscope. Compression-reperfusion induced a significant increase in rolling, sticking, and extravasation of the leukocytes. Treatment with the inhibitor strikingly reduced leukocyte sticking and extravasation. The present experiment has provided evidence that alphavbeta3 inhibition reduces leukocyte-endothelium interaction in our original pressure-induced reperfusion model.     

Proteolytic processing of osteopontin by PHEX and accumulation of osteopontin fragments in Hyp mouse bone,the murine model of X‐linked hypophosphatemia

X‐linked hypophosphatemia (XLH/HYP)—with renal phosphate wasting, hypophosphatemia, osteomalacia, and tooth abscesses—is caused by mutations in the zinc‐metallopeptidase PHEX gene (phosphate‐regulating gene with homologies to endopeptidase on the X chromosome). PHEX is highly expressed by mineralized tissue cells. Inactivating mutations in PHEX lead to distal renal effects (implying accumulation of a secreted, circulating phosphaturic factor) and accumulation in bone and teeth of mineralization‐inhibiting, acidic serine‐ and aspartate‐rich motif (ASARM)‐containing peptides, which are proteolytically derived from the mineral‐binding matrix proteins of the SIBLING family (small, integrin‐binding ligand N‐linked glycoproteins). Although the latter observation suggests a local, direct matrix effect for PHEX, its physiologically relevant substrate protein(s) have not been identified. Here, we investigated two SIBLING proteins containing the ASARM motif—osteopontin (OPN) and bone sialoprotein (BSP)—as potential substrates for PHEX. Using cleavage assays, gel electrophoresis, and mass spectrometry, we report that OPN is a full‐length protein substrate for PHEX. Degradation of OPN was essentially complete, including hydrolysis of the ASARM motif, resulting in only very small residual fragments. Western blotting of Hyp (the murine homolog of human XLH) mouse bone extracts having no PHEX activity clearly showed accumulation of an ～35 kDa OPN fragment that was not present in wild‐type mouse bone. Immunohistochemistry and immunogold labeling (electron microscopy) for OPN in Hyp bone likewise showed an accumulation of OPN and/or its fragments compared with normal wild‐type bone. Incubation of Hyp mouse bone extracts with PHEX resulted in the complete degradation of these fragments. In conclusion, these results identify full‐length OPN and its fragments as novel, physiologically relevant substrates for PHEX, suggesting that accumulation of mineralization‐inhibiting OPN fragments may contribute to the mineralization defect seen in the osteomalacic bone characteristic of XLH/HYP. © 2013 American Society for Bone and Mineral Research.     

Digital subtraction angiography of the thoracic aorta

Forty-three patients with acquired and congenital abnormalities of the thoracic aorta were studied using digital subtraction angiography (DSA) after an intravenous bolus injection of 40 ml of contrast material. Abnormalities studied included coarctation, pseudocoarctation, Marfan syndrome, cervical aorta, double aortic arch, aneurysm, dissection, and tumor. Twenty-four patients also had conventional angiography. DSA was accurate in 95% of cases; in the other 5%, involving patients with acute type I dissection, the coronary arteries could not be seen. The authors concluded that in 92% of their patients, DSA could have replaced the standard aortogram.     

Chondromalacia patellae: assessment with MR imaging

Magnetic resonance (MR) images of the posterior patellar hyaline articular cartilage were obtained in 23 subjects to determine if MR imaging could accurately demonstrate the patellar cartilage. Arthroscopy was used as the standard of reference. Three subjects were asymptomatic volunteers. In the remaining 20 who had patellofemoral pain, arthroscopy was performed before MR imaging in seven and afterward in 12; one did not undergo arthroscopy. MR imaging showed focal areas of swelling of the patellar cartilage, focal hypointensity, surface irregularity, areas of thinning, and areas of cartilage loss with exposure of subchondral bone. The surgical findings agreed with those from MR images in all seven patients who underwent arthroscopy before MR imaging and in ten of the 12 who underwent surgery afterward. MR imaging is an accurate means of examining the posterior patellar cartilage and should be considered as an alternative to diagnostic arthroscopy when chondromalacia patellae is suspected.     

Acute experimental cerebral ischemia: MR enhancement using Gd-DTPA

The effects of a paramagnetic contrast agent, gadolinium-DTPA (Gd-DTPA), on magnetic resonance (MR) imaging of acute cerebral ischemia was investigated in a feline model of middle cerebral artery occlusion. Imaging was performed both before and after administration of an intravenous dose of 0.2 mmol/kg of Gd-DTPA. The animals were then sacrificed for pathologic correlation. No changes in intensity or relaxation times were noted before or after Gd-DTPA administration in two animals with 2 hours of occlusion. Infarcts were noted before and after contrast enhancement in all six cats with ischemia of greater than 16-hours duration. Gd-DTPA caused significant increase in intensity of infarct but not in that of normal cerebral tissue. Rapid enhancement was visible in infarcts of 16-24 hours, but such enhancement was slower in infarcts of 72-168 hours, presumably owing to slowed inflow caused by increased vasogenic edema in the latter group. Contrast enhancement of acute cerebral ischemic lesions with Gd-DTPA offers no improvement in sensitivity of MR imaging, although the conspicuity of the lesion may be improved. Additionally, contrast media may provide potential temporal and pathophysiological data for better characterization of cerebral ischemia.