Allograft heart valve viability and valve-processing variables

BACKGROUND: The impact of allograft valve viability on valve durability remains controversial. Analyses of our clinical results have demonstrated the superiority of the cryopreserved valve viable at the time of implantation over the 4 degrees C stored valve nonviable at the time of implantation. In this study, we quantitatively assessed the effects on viability of current and past valve-processing protocols at The Prince Charles Hospital. METHODS: The viability of pulmonary valves was quantitatively analyzed by thin-layer autoradiography to assess the effects of donor type, antibiotics, and valve storage. RESULTS: Control valve segments obtained from beating-heart donor valves had a higher initial viability (0.92+/-0.02) than nonbeating-heart donor valves (0.66+/-0.03). Cryopreservation after low-dose antibiotic sterilization significantly reduced viability to 50% to 60% of the control, and in the presence of amphotericin B, viability dropped further to 10% to 36% of the control. After 7 days' storage at 4 degrees C, viability was reduced to 2% of control and to 0% viability after 21 days. CONCLUSIONS: For maximal preimplantation viability, valves should be procured as soon as possible after cessation of heart beat and should be cryopreserved if they are not to be clinically implanted within 1 to 2 days. Amphotericin B should not be used in conjunction with cryopreservation if viability is to be maximized.     

Influence of various doses of aspirin (in vivo) on platelet arachidonic acid metabolism (ex vivo) and function

It has been reported that aortic homografts that have been cryopreserved before transplantation remain viable longer as an allograft than tissue stored at 4 degrees C in an antibiotic solution. In the present study, we tested the hypothesis that storage of cardiac valve tissue by cryopreservation or by antibiotic preservation may alter the metabolic status of the tissue. Initially, we collected aortic valves composed of cardiac tissue, aortic root, and valvular tissue from cadaver donors. These specimens were divided into three equal portions, and one portion was analyzed before storage while the other two parts were stored for 3 weeks at either 4 degrees C in an antibiotic solution or at -196 degrees C in liquid nitrogen. All specimens were examined with regard to the following parameters: tissue structure, tissue viability, cell proliferative capacity, metabolic function, and identification of cell-specific antigens. We found no significant alterations in the structure of any of the three tissue components after antibiotic preservation or cryopreservation; however, cell viability and cell number were decreased in all three groups. All tissue samples grew in culture before storage. When we compared activities of the following organellar marker enzymes--lysosomal acid lipase, plasma membrane 5' nucleotidase, mitochondrial cytochrome oxidase, and microsomal neutral alpha-glucosidase--we observed no major differences between tissues stored by either technique. In addition, we observed no loss of enzymic activity as a result of storage. Finally, when cell lines isolated from each tissue specimen were incubated with monoclonal antibodies against cell-specific antigens in an immunoperoxidase assay, all the cell cultures proved to be endothelial cells. These results suggest that although cardiac valve tissue stored by cryopreservation or by antibiotic preservation retained its normal structure and metabolic capabilities, both storage techniques produced significant decreases in cell numbers and viability. However, only endothelial cells from tissue stored by cryopreservation retained the capacity to proliferate in vitro. These findings have important implications for the function of aortic homografts transplanted after storage.     

Should the aortic valve homograft be recryopreserved?

BACKGROUND: The number of homograft donors is limited and the once-thawed homograft may be unsuitable for the recipient and obliged to be wasted. The purpose of this study was to investigate the possibility of recryopreserving and using the once-thawed homograft for another patient. METHODS: Canine aortic valve leaflets were frozen to -80 degrees C by a programmed freezer, stored in liquid nitrogen, and thawed after 1 week. A subgroup of leaflets was left at 4 degrees C for 15 minutes, re-cryopreserved, and thawed after 1 week. Pathologic and flow cytometric evaluations were performed. RESULTS: After thawing, by pathology, alignment of the fibers was acceptably maintained but the membrane and cytoplasm of the fibroblast were damaged. These findings were not significantly aggravated even after rethawing. By flow cytometry, fibroblast viability was 90.7%+/-1.7% immediately after thawing, 87.6%+/-1.0% after thawing for 15 minutes at 4 degrees C, 63.7%+/-2.7% during refreezing at 0 degrees C, and 39.4%+/-4.3% after rethawing. CONCLUSIONS: From the standpoint of fibroblast viability, it is not possible to recryopreserve the once-cryopreserved and thawed aortic valve homograft.     

Successful thrombolysis for prosthetic pulmonary valve obstruction

Thrombosis is a serious complication of prosthetic heart valve operations. In recent years, systemic thrombolysis has emerged as a suitable alternative to surgery. Experience with thrombosis of pulmonary prosthetic valves is very limited. We report a case of successful administration of intravenous streptokinase for thrombosis of a St. Jude Medical prosthetic valve 3 weeks after pulmonary valve replacement.     

Quantification of wild-type mitochondrial DNA and its 4.8-kb deletion in rat organs

Long-term durability of aortic valve allografts may be enhanced by cellular capacities for regeneration and repair. To evaluate aortic valve graft production of an important structural protein, rat aortic roots were implanted heterotopically into the abdominal aorta of recipient rats. Grafts were either syngeneic or strongly allogeneic, were implanted either fresh or after cryopreservation, and were left in place 2 to 21 days after implantation. A total of 80 aortic valve grafts and the corresponding native aortic valves were examined. The grafts were retrieved and immunocytochemically stained for the presence of procollagen, a precursor to collagen. Regardless of histocompatibility or preservation, grafts exhibited consistent procollagen presence that equaled or exceeded that seen in the corresponding native valves. Positive procollagen staining was predominantly in the aortic wall. The most prominent staining was near the hinge point of the valve leaflets, with no staining in the free portion of the leaflets. Staining with alpha-actin demonstrated vascular smooth muscle in sites remote from the areas positive for procollagen, which suggests that vascular smooth muscle was not responsible for the procollagen production. These findings indicate that cryopreservation is compatible with persistent fibroblast viability and in vivo protein synthesis by both syngeneic and allogeneic aortic valve grafts.     

Comparison of a 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-t etrazolium hydroxide (XTT) colorimetric method with the standardized National Committee for Clinical Laboratory Standards method of testing clinical yeast isolates for susceptibility to antifungal agents

We report here a case of concomitant aortic and tricuspid valve endocarditis occurring in a 26-year-old woman 2 weeks after she had given birth by cesarean delivery. Preoperative transthoracic echocardiography revealed a previously undetected aorta-right atrium fistula, which at operation appeared to be congenital in origin. Surgical treatment consisted of aortic valve replacement with a pulmonary autograft, tricuspid valve replacement with a cryopreserved mitral homograft, and closure of the fistulous communication. The postoperative recovery was uneventful.     

Atrial septal defect with tricuspid valve regurgitation and heart failure in early childhood: report of a case with successful surgical management consisting of special patch closure technique and use of polydioxanone suture for the tricuspid annuloplasty

A management of surgery for infant having a small left ventricular cavity associated with atrial septal defect and tricuspid valve regurgitation was presented. A right upper part of the defect was remained in open during a patch closure of ASD and this portion was temporarily sutured by prolene stayed extracardialy through Waterston's groove. During 20 minutes after weaning from the cardiopulmonary bypass, left heart failure did not appear and then it was completely closed. DeVega's method was employed for the tricuspid valve regurgitation using a absorbale Polydioxanone suture, because of a growth of the sutured annulus. Postoperative course was uneventful and trivial tricuspid valve regurgitation was recognized in angiocardiographic studies performed in 2 weeks and 6 months after operation.     

Isolated massive chylopericardium. Complication of open heart surgery for aortic valve replacement

Chylopericardium following open-heart surgery for aortic valve replacement in a 53-year-old woman is described. Five weeks after surgery, the chylous pericardial effusion was detected when the patient developed recurrent chest pain and cardiomegaly. Treatment included drainage of the fluid and partial pericardiectomy. No recurrence of the chylopericardium was observed in this patient up to 14 months after surgery.     

No-react detoxification process: a superior anticalcification method for bioprostheses

BACKGROUND: Glutaraldehyde pretreatment of bioprosthetic heart valves is the major pathogenic factor in their calcific degeneration. This comparative study investigates the merit of the No-React aldehyde detoxification process as an alternative modifier of xenograft tissues. METHODS: Glutaraldehyde- and No-React-pretreated porcine aortic valve cusps were implanted subcutaneously in 6-week-old rats (n = 20). At 3, 6, and 14 weeks, randomly selected animals were sacrificed and the explants underwent mineral and morphologic analyses. Glutaraldehyde- and No-React-treated bovine pericardium and porcine aortic valve cusp were incubated in fibroblast cell culture plates. Cell viability was observed under reversed microscope at 6, 24, 48, and 96 hours. Erythrosin B dye exclusion test was used to validate percent cell death. RESULTS: Pretreatment with No-React significantly inhibited calcification of aortic cusp subcutaneous implants throughout the 14-week period (mean tissue Ca2+ content = 1.3 +/- 0.7 micrograms/mg at 14 weeks.) Glutaraldehyde-treated cusps underwent protracted calcification (Ca2+ content = 190.6 +/- 89.5 micrograms/mg; p < 0.01). Morphologic findings correlated with mineral analyses. One-hundred percent of fibroblast cells survived in the presence of No-React-treated tissue, with a growth pattern indistinguishable from control cell culture (ie, in the presence of no tissue). The cells incubated with glutaraldehyde-treated tissue showed signs of nonviability by 6 hours, with 100% cell death by 48 hours. Dye exclusion tests validated these findings. CONCLUSIONS: The No-React detoxification process completely abolishes the cytotoxicity of the xenograft tissue and inhibits calcific degeneration.     

Polyvinyl alcohol as a defined substitute for serum in vitrification and warming solutions to cryopreserve ovine embryos at different stages of development

The purpose of this study was to assess the viability of ovine embryos after replacing fetal calf serum (FCS) with polyvinyl alcohol (PVA) in vitrification and warming solutions. Ovine embryos were obtained from superovulated Sardinian breed ewes at 4, 5, 6, and 7 days after insemination. All vitrification and warming solutions were prepared using buffered saline solution with 20% FCS (group a) or 0.1% PVA (group b). Embryos were vitrified in 20 microliters of glycerol 3.4 M + ethylene glycol 4.6 M and loaded into the centre of 0.25 ml straws between two columns of sucrose solution (0.5 M), and plunged immediately into liquid nitrogen. After being warmed in a water bath at 35 degrees C for 10 s, the vitrified embryos were moved to 0.25 M sucrose solution for 3 min. Embryos were cultured in TCM-199 after washing with 10% FCS and sheep oviductal epithelial cells up to hatching or re-expansion of the blastocoelic cavity. No significant difference in the viability rates was observed between embryos vitrified/warmed in PVA or FCS solutions. In both groups, the rate of in vitro viability was (P < 0.01) lower at the precompacted and compacted morula stages than at the expanded, hatching or hatched blastocyst stage. In both groups, early blastocysts were less viable than expanded (P < 0.01), hatching or hatched blastocyst (P < 0.05). There was no significant difference in survival rates at days 14 (79 and 76%) and 45 (63 and 59%) after transfer into sychronised recipients between vitrified expanded blastocysts of groups a and b, respectively. These results suggest that it is possible replace serum with PVA in vitrification and warming solutions without reducing in vivo and in vitro viability.     

Molecular genetics of small bowel cancer

Streptococcus adjacens has never previously been reported as an etiologic organism of infective endocarditis in Taiwan. We describe a case of severe native valve endocarditis caused by S. adjacens, involving the mitral valve, the aortic valve, and the left atrium, in a 29-year-old woman with nephrotic syndrome on steroid therapy. Blood cultures yielded gram-positive cocci that grew poorly on blood agar but strongly on chocolate agar. Despite aggressive antibiotic treatment, the patients continued to have high fever and progressive congestive heart failure, which necessitated surgical intervention. Symptoms were alleviated after surgery; teicoplanin was continued for 4 weeks and the patient remained symptom-free at the 6 month follow up. To our knowledge, this is the first reported case of bacterial endocarditis involving the left atrium without preexisting myxoma.     

Rupture of asymptomatic mycotic aneurysm after valve replacement in infective endocarditis

Mycotic cerebral aneurysms (MCA) are one of the most serious complications of infective endocarditis. The rupture of MCA in patients under anticoagulant therapy following valve replacement carries high mortality. We encountered this serious complication in a patient who had no neurologic symptoms. A 12-year-old girl was scheduled for mitral valve replacement (MVR) 5 weeks after antibiotic therapy for infective endocarditis caused by Staphylococcus aureus. Before the surgery, she did not have any neurologic symptoms or abnormal findings in CT scanning examination. The surgery to remove her mitral valve with bacterial vegetations and replace it with an artificial valve proceeded smoothly and she appeared to begin an uneventful postoperative recovery. However, she suddenly began to complain of severe headache and became unconscious on the fifth days after MVR. A CT scan showed cerebral herniation due to a major subdural hematoma. A ruptured MCA was detected in the orbito-frontal artery and clipped in an emergency operation. She was transferred to the intensive care unit and given continuous infusion of barbiturate to prevent increase of her intracranial pressure. CT scanning and arteriography 10 days after the MCA clipping, revealed a new subdural hematoma and MCA just proximal to the previous clip. It is important to bear in mind that patients with infective endocarditis can have mycotic cerebral aneurysms without any clinical neurologic symptoms.     

Cell survival following bone-anterior cruciate ligament-bone allograft transplantation: DNA fingerprints, segregation, and collagen morphological analysis of multiple markers in the canine model

Bone-anterior cruciate ligament-bone allograft transplantation has become recognized as a potential solution to reconstruction of the anterior cruciate ligament (ACL). The purpose of this study was to determine the time-dependent fibrocyte donor cell survival rate after cryopreserved bone-ACL-bone allograft transplantation. Additionally, bony incorporation of the pediculated bone plugs was examined. The ability to successfully transplant allogenous ACL fibrocytes and have them survive has not previously been documented. In this study, DNA fingerprints identified and documented the survival rate of the cellular DNA in transplanted ACL allografts for ACL re-construction in the knee joints of 10 skeletally mature dogs. At 4, 8, 26 and 52 weeks after ACL allograft transplantation, DNA probes, H & E, Giemsa, Goldner, PAS and polarized light staining was done to demonstrate the time-dependent changes in the allografts after transplantation. At 4 weeks host fibrocytes began to grow into the graft; however, histologically the cells could not be distinguished as to host or donor origin. After 4 weeks the DNA pattern reflected only the band pattern of the host. This reveals the early cellular infiltration activity of the host into the ACL allograft, also demonstrated in the light microscopy stainings. The survival rate of transplanted allogenous ACL fibrocytes had not been documented before this study. There is no evidence that ACL allograft cells survive in the intra-articular environment of the host's knee. Within 4 weeks ACL allografts became completely repopulated with host cells. The cells that migrate early into the ACL allografts are probably of synovial origin because they are present before revascularization and collagen reorganization occur. We conclude from this study that viable cells in transplanted ACL allografts did not survive longer than 4 weeks after intra-articular transplantation. Advances in molecular biology may offer new approaches to alter or stimulate fibrocyte population and function in the transplanted ACL allograft used for ACL reconstruction. New methods to maintain the viability of donor cells may be necessary to improve the biomechanical and histological properties of autografts or allografts for ACL reconstruction.     

Protective effects of various preservation solutions on cultured endothelial cells

Vascular endothelium represents the first target in organ preservation and plays an important role in reperfusion injury. Bovine aortic endothelial cells were cultivated and the most commonly used preservation solutions, such as University of Wisconsin HTK (Brettschneider's histidine-tryptophane-ketoglutarate), and Euro-Collins solutions were tested on the endothelial monolayer. In addition, one group of cultivated cells was preserved with cold saline solution, and endothelial monolayers grown in culture medium were used as controls. The quality of preservation was assessed after 24, 48, and 72 hours of cold storage. Reperfusion was simulated and its effects were observed by reincubation in culture medium at 37 degrees C for 6 hours. The total number of cells, cell viability (determined using trypan blue exclusion), and morphologic alterations were determined. Prostacyclin release was evaluated as a biochemical marker. University of Wisconsin solution maintains more than 99% cell viability after rewarming after both 24 and 48 hours of cold storage. After 72 hours, 86.7% of cells were still viable. Preservation with HTK and Euro-Collins solution allowed cell survival for only 24 hours (96.7%, HTK; 49.9%, Euro-Collins), with no viable cells seen after 48 hours. The cold saline-preserved sample showed 57.8% viable cells after 24 hours and 29.7% after 48 hours. No viable cells were detectable after 72 hours. Light microscopy revealed several patterns of both structural damage and intracellular change (nucleus and cytoplasm) in the endothelial monolayer after preservation with HTK, Euro-Collins solution, and cold saline solution. No morphologic alterations were seen in the University of Wisconsin solution group for as long as 72 hours.(ABSTRACT TRUNCATED AT 250 WORDS)     

Donor-specific anti-human leukocyte antigen class I antibodies after implantation of cardiac valve allografts

The allospecific humoral immune response was examined in 31 patients in the first year after implantation of cryopreserved human cardiac valves. We determined the percentage antibodies against human leucocyte antigens (HLA) class I in a complement-dependent microlymphocytotoxicity test against a panel of 50 selected donors carrying most of the defined HLA-A and HLA-B specificities (panel-reactive antibodies). In blood samples taken immediately before implantation, no antibodies could be detected. Thereafter, antibodies were present in 23 of 31 (74%) patients (median panel-reactive antibodies: 57%, range 9% to 91%). In 21 patients the HLA-type of the valve donor was available. In four patients no blood samples taken after 4 weeks were available. In 14 of 17 patients (82%) with a follow-up of more than 1 month antibodies were present, and in 12 of these 14 (86%) antibodies were specifically directed against HLA class I of the donor. In conclusion, the formation of donor-specific antibodies is frequently observed after human cardiac valve replacement. It could be one of the factors leading to valve destruction and dysfunction.     

Open-heart correction of tetralogy of Fallot in an acyanotic dog

Tetralogy of Fallot was diagnosed in an acyanotic 11-month-old dog. Predicted pressure gradient across the pulmonic valve, as assessed by use of continuous wave Doppler echocardiography, was 94.5 mm Hg. Bidirectional shunting was identified by means of selective angiography. Open-heart correction was performed, using a transatrial approach with limited ventriculotomy and cardiopulmonary bypass. The hypertrophied infundibulum was resected, the ventricular septal defect was closed primarily, and a transannular pericardial patch graft was applied. Pressure gradients across the pulmonic valve were 52.9 and 22.8 mm Hg 2 weeks and 4 months after surgery, respectively. Advances in cardiopulmonary bypass, anesthetic management, and use of the transatrial approach may improve the success of open-heart correction of tetralogy of Fallot in dogs.     

The relationship of eosinophil viability enhancing activity in sputum and clinical symptoms in asthma

We studied the correlation of eosinophil viability enhancing activity (EVEA) in sputum from asthmatic children and clinical symptoms. Sputum from asthmatic children and equal volume of saline were mixed with a Vortex mixer and centrifuged at 40000 G. Clear supernatants were obtained and filtered with 0.22 micron membrane filter. Periphral blood eosinophils purified by Percoll density gradient centrifugation and CD16 negative selection/ immunomagnetic beads technique were incubated with sputum extract for 4 days. Eosinophil viability was examined by staining the cells with fluorescen diacetate and propidium iodide and expressed as eosinophil viability enhancing activity (EVEA). Eosinophil cationic protein (ECP) concentrations in sputum were also measured by a radioimmunoassay. Correlation between EVEA in sputum and pulmonary functions, attack score, treatment score, or sputum ECP was determined by Spearman correlation test. Sputum EVEA was correlated 1) inversely with pulmonary functions on sampling, 2) with attack score of 2 weeks period before sampling, 3) with treatment score of 1 month to 1 week before and 1 to 2 weeks after sampling, and 4) with sputum ECP in individual cases. Sputum EVEA may serve as a suitable parameter for monitoring airway inflammation in asthma.     

Infective endocarditis with a huge mitral vegetation related to atopic dermatitis and high serum level of infection-related antiphospholipid antibody: a case report]

A 24-year-old woman with atopic dermatitis was admitted to our hospital with fever. Echocardiography showed a huge vegetation attached to the posterior mitral commissure without mitral valve dysfunction. Blood culture identified methicillin-sensitive Staphylococcus aureus. The serum level of antiphospholipid antibody was elevated. A splenic infarction occurred on the second hospital day. Surgery to resect the residual mobile vegetation was performed uneventfully on the 6th hospital day. The postoperative course was uneventful, and the patient was discharged after 4 weeks of antibiotic therapy. Preservation of the mitral valve is rare in the face of virulent Staphylococcus infection and the presence of a huge mobile vegetation. These findings were apparently related to the high serum level of infection-related antiphospholipid antibody and atopic dermatitis.     

Assessment of coenzyme Q10 tolerability in Huntington''s disease

BACKGROUND: Maintaining viability of periodontal ligament cells is important after an avulsion injury occurs. OBJECTIVE: This study examined cell death and compared different transport media at varying times. Lactate dehydrogenase (LD) measurements were made on root surfaces of extracted teeth for evaluating the results of breakdown and necrosis of periodontal ligament cells. STUDY DESIGN: Teeth were divided into three groups. Each was immersed in one of the following storage media: Hanks balanced salt solution, Custodiol solution (an organ storage medium), or sterile saline solution. I.D. measurements (an indicator of cell death) were made at 2, 6, 24, 72, and 168 hours after extraction. RESULTS: When the LD levels were compared according to the time periods, a progressive increase of cell death was found to be statistically significant (p < 0.001). However, when the effects of both time periods and the kind of storage medium on the LD levels were examined, the LD levels of the saline solution storage medium were found to be significantly higher (p < 0.001) than that of the other two groups. CONCLUSIONS: Both Hanks balanced salt solution and Custodiol appeared to be suitable transport media for maintaining cell viability, whereas saline solution was not.     

连续碳酸化分解过程进料流量软测量

维持进料量稳定,是保证碳酸化分解过程生产优化稳定的关键。但由于分解槽进料管较短、管径比较大、容易结疤,进料量难以用常规的过程检测仪表进行检测。本文建立了汇流的伯努利方程,总结出进料阀门的局部阻力系数与开度的关系,推导出高位槽液位、进料阀门及进料流量的函数关系,从而建立了进料流量的软测量模型。现场实验数据表明,进料流量软测量模型的预报精度在2 06%以内,可满足现场实际需要。