Biological evaluation of intervertebral disc cells in different formulations of gellan gum‐based hydrogels

Gellan gum (GG)‐based hydrogels are advantageous in tissue engineering not only due to their ability to retain large quantities of water and provide a similar environment to that of natural extracellular matrix (ECM), but also because they can gelify in situ in seconds. Their mechanical properties can be fine‐tuned to mimic natural tissues such as the nucleus pulposus (NP). This study produced different formulations of GG hydrogels by mixing varying amounts of methacrylated (GG‐MA) and high‐acyl gellan gums (HA‐GG) for applications as acellular and cellular NP substitutes. The hydrogels were physicochemically characterized by dynamic mechanical analysis. Degradation and swelling abilities were assessed by soaking in a phosphate buffered saline solution for up to 170 h. Results showed that as HA‐GG content increased, the modulus of the hydrogels decreased. Moreover, increases in HA‐GG content induced greater weight loss in the GG‐MA/HA‐GG formulation compared to GG‐MA hydrogel. Potential cytotoxicity of the hydrogel was assessed by culturing rabbit NP cells up to 7 days. An MTS assay was performed by seeding rabbit NP cells onto the surface of 3D hydrogel disc formulations. Viability of rabbit NP cells encapsulated within the different hydrogel formulations was also evaluated by Calcein‐AM and ATP assays. Results showed that tunable GG‐MA/HA‐GG hydrogels were non‐cytotoxic and supported viability of rabbit NP cells. Copyright © 2012 John Wiley & Sons, Ltd.     

Platelet glycoproteins IIb and IIIa as a calcium channel in liposomes

Human platelet membrane glycoproteins IIb and IIIa (GPIIb and IIIa) were incorporated into phospholipid vesicles by the reverse-phase technique to assess the ability of GPIIb and IIIa to function as a Ca2+ channel. Movement of Ca2+ across the lipid bilayer was quantitated by injection of proteoliposomes with encapsulated Fura-2 into Ca2+ buffers and measurement of Fura-2 fluorescence as an indicator of Ca2+ influx. Reciprocally, to assess the function of proteins in an inside-out orientation, Ca2+-loaded vesicles were injected into Ca2+-free buffer and Ca2+ efflux monitored by a calcium electrode. Incorporation of the IIb-IIIa complex produced significant facilitation of Ca2+ movement across the lipid bilayer. No net transmembrane Ca2+ movement was seen with dissociated IIb and IIIa. Movement of Ca2+ was proportional to the transmembrane Ca2+ gradient. Ca2+ movement into the vesicles was inversely proportional to extravesicular NaCl from 25 to 150 mmol/L, analogous to several studies in the intact platelet. Adenosine triphosphate had no effect on Ca2+ movement into or out of the vesicles. Specific inhibition of a Ca2+ shift into the vesicles was seen with M148, a monoclonal antibody to IIb/IIIa, while no inhibition was observed with a panel of other anti-IIb/IIIa monoclonal antibodies. This suggests that a specific site on the complex or orientation of the complex is essential for calcium channel function. These data demonstrate that the GPIIb/IIIa complex can serve as a passive Ca2+ channel across a phospholipid bilayer and has the potential to play a role in Ca2+ flux across the platelet plasma membrane.     

Factor V deficiency in Philadelphia-positive chronic myelogenous leukemia

Factor V deficiency has been identified in 8 of 8 patients 7--20 yr of age, with Philadelphia-positive (Ph1+) chronic myelogenous leukemia (CML). In these 8 patients, factor V deficiency was not due to hepatic dysfunction, factor V inhibitors, or disseminated intravascular coagulation. In 3 patients, factor V activity rose 10%--12% (0.10--0.12 U/ml) after the infusion of 28--31 ml/kg body weight of fresh frozen plasma (FFP). The rise persisted less than 14 hr. The mean measured postinfusion rise in factor V was 18% of the expected rise calculated from the volume of FFP infused in the patients' plasma volume. In 4 patients, a small transient rise in factor V activity occurred after splenectomy or plateletpheresis. Factor V deficiency was completely corrected after a marked reduction in bone marrow cellularity in 2 patients with Ph1+ CML treated with extensive chemotherapy, total body irradiation, and bone marrow transplantation. Factor V deficiency was retrospectively observed in 6 of 20 patients, ages 20--80 yr, with Ph1+ CML and 3 of 6 patients with other myeloproliferative disorders. The factor V deficiency appears to be associated with the large myeloid- megakaryocytic cell mass characteristic of CML and other myeloproliferative disorders.     

Coronary computed tomographic angiography for prediction of procedural and intermediate outcome of bypass grafting to left anterior descending artery occlusion with failed visualization on conventional angiography

Conventional coronary angiography (CCA) has considerable limitations regarding visualization of distal vessel segments in chronic total occlusion. We assessed the ability of coronary computed tomographic angiography (CCTA) to predict the success of coronary artery bypass grafting (CABG) to the chronically occluded left anterior descending coronary artery (LAD) incompletely visualized on CCA. Thirty symptomatic patients rejected for CABG on the basis of the CCA findings underwent preoperative CCTA before intended transmyocardial laser revascularization. The LAD was explored operatively in all patients, and CABG to the LAD was attempted if the distal vessel was suitable for anastomosis. The procedural outcome of CABG and the 6-month patency of the left internal mammary artery graft at follow-up CCTA were defined as the primary and secondary end point, respectively. The primary and secondary end points were achieved in 80% and 77% of patients, respectively. We found a significant correlation between the intraoperative and computed tomographic measurement of distal LAD diameter (R = 0.428, p = 0.037). On multivariate analysis, the maximum diameter of the distal LAD by CCTA (odds ratio 8.16, p = 0.043) was the only independent correlate of procedural success of CABG. A cutoff value of 1.5 mm for the mean distal LAD diameter predicted left internal mammary artery graft patency with 100% specificity and 83% sensitivity. Successful CABG resulted in significant improvements in angina class and left ventricular function in LAD segments at 6 months of follow-up. In conclusion, CCTA predicted both the procedural and the intermediate outcome of CABG to chronic LAD occlusion with failed visualization on CCA.     

Single coronary artery with anomalous origin from the pulmonary trunk: congenital lack of coronary arteries originating from the aortic bulb

A 23-year-old woman with an initial diagnosis of dilated cardiomyopathy for several years was referred to our center because of progressive congestive heart failure. Echocardiography showed a hypokinetic dilated left ventricle with globally impaired systolic function and an ejection fraction of 30%. We performed multislice cardiac computed tomography that revealed abnormal origin of the single coronary artery from the pulmonary trunk, lack of coronary arteries arising from the aorta, and abundant collateral flow from the bronchial artery to the anomalous coronary artery. This case report shows that cardiac computed tomography can be a valuable tool in the diagnosis of rare coronary anomalies.