It has been reported that the immune response due to alpha-Gal epitopes is an important factor in tissue valve failure. The elimination of the interaction between the natural anti-Gal antibodies and alpha-gal epitopes on the xenografts is a prerequisite to the success of xenografts in humans. Previously, we reported that the green coffee bean alpha-galactosidase could remove all alpha-Gal epitopes from cell surface of porcine aortic valve and pericardial tissue, but it has limitations on cost effectiveness. In this study we wanted to know whether the recently produced recombinant human alpha-galactosidase A has the same effective enzymatic activity as green coffee bean alpha-galactosidase in removing alpha-Gal epitopes from the same tissues. After treating fresh porcine aortic valve and pericardial tissue with recombinant alpha-galactosidase A, each sample was stained with Griffonia simplicifolia type I isolectin B4 indirect immunoperoxidase avidin-biotin technique. We then examined whether the alpha-Gal epitopes were reduced or abolished in each consecutive concentration of recombinant alpha-galactosidase A by comparing the degree of the Griffonia simplicifolia isolectin B4 staining. As a result, the recombinant alpha-galactosidase A could remove cell surface alpha-Gals on porcine aortic valve and pericardial tissue as effectively as green coffee bean alpha-galactosidase.
Adolescent ; Animals ; Aortic Valve/chemistry/cytology/*immunology ; Child ; Coffea/enzymology ; Epitopes/*immunology ; Heart Valve Prosthesis Implantation ; Humans ; Pericardium/chemistry/cytology/*immunology ; Recombinant Proteins/genetics/*immunology ; Swine ; Transplantation, Heterologous/immunology ; alpha-Galactosidase/genetics/*immunology

BACKGROUND: It is currently thought that tissue valve degeneration is related to an animal's immune response, which is mainly due to cell surface alpha-Gal epitopes. Cell surface alpha-Gal epitopes are known to be degraded by the enzyme called green coffee bean alpha-galactosidase. It is also well known that alpha-Gal epitopes are immunologically stained by Griffonia Simplicifolia isolectin type B4. We know that many commercially available tissue valves are made of aortic valves and pericardial tissue of pig. So, we investigated whether alpha-Gal epitopes of the aortic valve and pericardial tissue of a pig can be removed by green coffee bean alpha-galactosidase, and we did so by comparing immunologic staining of the tissues before and after the enzyme treatment. MATERIAL AND METHOD: After treating fresh porcine aortic valve and pericardial tissue with green coffee bean alpha-galactosidase at concentrations of 0.5 unit/mL, 1.0 unit/mL, 2.0 unit/mL, respectively, under the condition of pH 6.5, temperature 4degrees C and 24 hours of incubation, each sample was stained with Griffonia Simplicifolia isolectin type B4 immunofluorescent labeling. We then examined whether the alpha-Gal epitopes were reduced or abolished in each consecutive concentration of green coffee bean alpha-galactosidase by comparing the degree of the Griffonia Simplicifolia isolectin B4 staining in each sample. RESULT: In the pig aortic valve tissue, a 1.0 unit/mL concentration of green coffee bean alpha-galactosidase at pH 6.5, 4degrees C and reaction for 24 hours was enough for complete removal of alpha-Gal epitopes from the cell surface on the immunostaining with Griffonia Simplicifolia isolectin B4. On the other hand, more alpha-Gal epitopes were present in the pig pericardial tissue on Griffonia Simplicifolia isolectin B4 staining before the enzyme treatment, and 1.0 unit/mL of galactosidase was not sufficient for complete removal of alpha-Gal from the tissue. 2.0 units/mL of green coffee bean alpha-galactosidase was needed to completely remove the alpha-Gal epitopes from the pericardial tissue on immunostaining. CONCLUSION: The alpha-Gal epitopes of the pig's aortic valve and pericardial tissue were successfully stained with Griffonia Simplicifolia isolectin B4. We could remove nearly all the alpha-Gal epitopes using green coffee bean alpha-galactosidase at the concentration of 1.0 unit/mL in the aortic valve of pig, and 2.0 unit/mL was need to nearly completely remove all the alpha-Gal epitopes in the pericardial tissue of pig under the condition of pH 6.5, 4degrees C and 24 hours of reaction time. In the near future, removal of alpha-Gal epitopes in the pig's aortic valve and pericardial tissue will become a powerful tool for the improvement of the tissue valve durability. It needs to be determined if alpha-galactosidase treated pig tissue is immune to human anti-Gal antibody or anti-Gal monoclonal antibodies.
alpha-Galactosidase ; Aortic Valve ; Coffee ; Epitopes ; Galactosidases ; Griffonia ; Hand ; Humans ; Hydrogen-Ion Concentration ; Lectins ; Pericardium ; Plant Lectins ; Reaction Time ; Tissue Transplantation

BACKGROUND: Various experimental trials for the development of bioprosthetic devices are actively underway, secondary to the limited supply of autologous and homograft tissue to treat cardiac diseases. In this study, porcine bioprostheses that were treated with glutaraldehyde (GA), ethanol, or sodium dodecylsulfate (SDS) were examined with light microscopy and transmission electron microscopy for mechanical and physical imperfections before implantation. MATERIAL AND METHOD: 1) Porcine pericardium, aortic valve, and pulmonary valve were examined using light microscopy and JEM-100CX II transmission electron microscopy, then compared with human pericardium and commercially produced heterografts. 2) Sections from six treated groups (GA-Ethanol, Ethanol-GA, SDS only, SDS-GA, Ethanol-SDS-GA and SDS-Ethanol-GA) were observed using the same methods. RESULT: 1) Porcine pericardium was composed of a serosal layer, fibrosa, and epicardial connective tissue. Treatment with GA, ethanol, or SDS had little influence on the collagen skeleton of porcine pericardium, except in the case of SDS pre-treatment. There was no alteration in the collagen skeleton of the porcine pericardium compared to commercially produced heterografts. 2) Porcine aortic valve was composed of lamina fibrosa, lamina spongiosa, and lamina ventricularis. Treatment with GA, ethanol, or SDS had little influence on these three layers and the collagen skeleton of porcine aortic valve, except in the case of SDS pre-treatment. There were no alterations in the three layers or the collagen skeleton of porcine aortic valve compared to commercially produced heterografts. CONCLUSION: There was little physical and mechanical damage incurred in porcine bioprosthesis structures during various glutaraldehyde fixation processes combined with anti-calcification or decellularization treatments. However, SDS treatment preceding GA fixation changed the collagen fibers into a slightly condensed form, which degraded during transmission electron micrograph. The optimal methods and conditions for sodium dodecylsulfate (SDS) treatment need to be modified.
Aortic Valve ; Bioprosthesis ; Collagen ; Connective Tissue ; Electrons ; Ethanol ; Glutaral ; Heart Diseases ; Humans ; Light ; Microscopy ; Microscopy, Electron ; Microscopy, Electron, Transmission ; Pericardium ; Pulmonary Valve ; Skeleton ; Sodium ; Transplantation, Heterologous ; Transplantation, Homologous

Purpose: We applied a low-dose fluoroscopic protocol in routine diagnostic cerebral angiography and evaluated the feasibility of the protocol. Materials and Methods: We retrospectively reviewed a total of 60 patients who underwent diagnostic cerebral angiography for various neurovascular diseases from September to November 2019. Routine protocols were used for patients in the first phase and low-dose protocols in the second phase. We compared radiation dose, fluoroscopy time, and complications between groups. Results: Age, diseases, and operators were not significantly different between the two groups. The mean fluoroscopy dose significantly decreased by 52% in the low-dose group (3.09 vs. 6.38 Gy·cm2 ); however, the total dose was not significantly different between the two groups (34.07 vs. 33.70 Gy·cm2 ). The total fluoroscopic time was slightly longer in the low-dose group, but the difference was not statistically significant (12.2. vs. 12.5 minutes). In all patients, angiography was successfully performed without complications. Conclusion The low-dose fluoroscopy protocol is feasible to apply for diagnostic cerebral angiography in that this protocol could significantly reduce the fluoroscopic dose.

Purpose: We present ultrasound-guided percutaneous low-dose thrombin (200–250 IU) injection for the treatment of iatrogenic femoral pseudoaneurysms. Second, we compared patient and procedure factors between subcutaneous hematoma and pseudoaneurysm groups. Materials and Methods: From April 2012 to May 2018, 8425 patients underwent neurointervention. Among these patients, 18 had small subcutaneous hematomas and 6 had pseudoaneurysms. Pseudoaneurysms in the neck and entire sac were visualized, and low-dose thrombins were injected while visualizing a “whirlpool” hyperechoic core in the pseudoaneurysm sac. Subcutaneous hematomas were treated with simple compression. We compared the following parameters between the subcutaneous hematoma group and pseudoaneurysm group: sex, age, body mass index (BMI), type of procedure, heparin usage, sheath size, procedure time, and number of previous neurointervention procedures with the Mann-Whitney U test. Results: Most of the pseudoaneurysms were successfully occluded with 200 IU of thrombin (n=5). Only 1 pseudoaneurysm required a slightly higher thrombin concentration (250 IU, n=1). During the short-term follow-up, no residual sac was observed and no surgical repair was necessary. Pain in the groin region was alleviated. During the 1-month follow-up, no evidence of pseudoaneurysm recurrence nor subcutaneous hematoma was noted. Patient factors (sex, age, and BMI) and procedure factors (heparin usage, sheath size, procedure time, number of previous procedures) were not statistically different between the subcutaneous hematoma and pseudoaneurysm groups. Conclusion Ultrasound-guided percutaneous low-dose thrombin injection (200–250 IU) is safe, effective, and less invasive for treating iatrogenic femoral pseudoaneurysm in neurointervention.