The effect of an addition of sodium hydrogenphosphate to poly(D,L)lactide--results of in vitro examinations.

Aim of the study was to examine the influence of sodium hydrogenphosphate (NaP) on the pH value and the mechanical characteristics of degrading poly(D,L)lactide (PDLLA). Test rods of PDLLA with or without NaP amounting to 1, 10, 25 or 50 mol per 100 mol lactate, the degradation product of PDLLA, were produced by injection molding. Molecular weight and bending strength of the rods were measured before and after an accelerated in vitro-test (55 degrees C, Ringer's solution (RS)). For a long-time degradation test PDLLA-rods with or without 1 mol% NaP were placed in RS or Soerensen buffer solution (SB, pH 7.4) at 37 +/- 1 degrees C. Measurements of pH and determination of length, volume and weight of the samples were done in 2-4 week intervals up to the 52nd week after incubation. A pH-drop was measured in RS or SB containing pure PDLLA after 28 or 36 weeks respectively. Stabilization of the pH value due to admixed NaP delayed the degradation related pH drop for 8 weeks in RS or SB. A strong increase of length, volume and weight was measured in PDLLA + NaP-rods. In conclusion minimal stabilization of pH but also an increase of outer dimensions of the samples was found due to the admixture of NaP to PDLLA. Thus, an addition of substantially higher amounts than 1 mol% NaP in PDLLA can not be recommended, regardless of the positive effects on pH stabilization.

Degradation of poly(D,L)lactide implants with or without addition of calciumphosphates in vivo.

The study was aimed at examining the in vivo degradation of pure poly(D,L)lactide (PDLLA) or PDLLA with an admixture of calciumphosphates. One rod (20 x 3 x 2 mm) and one cube (3 x 2 x 2 mm) of pure PDLLA, PDLLA with tricalciumphosphate (PDLLA + TCP) or PDLLA with calciumhydrogenphosphate (PDLLA + CHP), respectively, were implanted into the dorsal muscles of 50 male Wistar Albino rats. After definite intervals (from 2nd to 72nd week), pH measurements were performed in the environment of the implants. Afterwards, the cubes with their surrounding tissues were excised for histological examinations, measurements of the outer dimensions and mechanical analyses of the explanted rods were performed. No drop of more than 0.1 pH units was detectable in the tissue surrounding any type of implants. No advantageous effect of the calciumphosphates could be proved. A mild foreign body reaction could be observed around PDLLA implants. After 72 weeks, pure PDLLA had been totally resorbed from the extracellular space, the degradation of calciumphosphate-enriched PDLLA was still in progress. A large amount of inflammations occurred in the tissues surrounding PDLLA with an admixture of slowly degrading TCP or CHP, leading to two abscesses and four fistulas at PDLLA + TCP, and two abscesses and three fistulas at PDLLA + CHP implantation site. Bending strength of pure PDLLA was constant up to the 4th week post-implantation and reduced to 60% of the initial value up to the 12th week. No traces of crystallinity could be observed during the degradation of PDLLA. As a conclusion of the study, complete resorption from the extracellular space and tissue tolerance of pure PDLLA is proved. An admixture of small calciumphosphate particles is not suitable to improve the biocompatibility of PDLLA but leads to a decrease in the mechanical characteristics.

[Cryosurgical ablation of bone tissue with a newly developed miniature cryoprobe--adaptation of the method for use in bones in vitro and in vivo]. / Die kryochirurgische Ablation von Knochengewebe mittels einer neuartigen Miniaturkryosonde--Adaptation der Methodik für die Anwendung am Knochen in vitro und in vivo.

AIM: Up to now, modern miniature cryoprobes have been used successfully for local destruction of soft tissue tumors without damaging adjacent healthy tissue. In this study, the methodology of cryoablation was applied to bone and the freezing effect as well as the cooling capacity of the probe was examined in vivo and in vitro. METHODS: Freezing was performed by cooling one or two probes, with a diameter of 3.2 mm to -180 degrees C with liquid nitrogen. The cooling capacity of the probes was determined under thermic control by an in vitro measurement on human bone, followed by an in vivo measurement on femoral and tibia bones of a sheep. RESULTS: The in vitro freezings achieved a sufficient tissue cooling using one or two cryoprobes. The simultaneous use of 2 probes resulted in a synergistic effect between the probes. According to the body heat, the registered temperature curves, during the in vivo freezings, showed a more flat trend. Nevertheless, temperatures below -50 degrees C were achieved at a distance of 1 cm from the probe due to the synergistic effect. Local or systematical intraoperative complications have not been observed. CONCLUSION: An adequate tissue cooling of bone matrix can be achieved within in vivo freezings through the use of one or more miniature cryoprobes so that the use of this probe could possibly become an alternative or supplement to the surgical resection of pathologic bone processes.

Coronary oxygen persufflation combined with HTK cardioplegia prolongs the preservation time in heart transplantation.

BACKGROUND: One of the most restricting factors remaining in heart transplantation is the limited myocardial ischemia time. A new approach towards the prolongation of this time is the combination of primary cardioplegic arrest followed by continuous coronary oxygen persufflation (COP) with gaseous oxygen. METHODS: This technique was applied in pig hearts, which we transplanted orthotopically after cardioplegic arrest by original (n = 5) and modified (addition of hyaluronidase: n = 11) Bretschneider HTK solution and 14 h of hypothermic preservation. Depending on the different preservation techniques, we created four groups: (1), original HTK (HTK), n = 5; (2), modified HTK (mHTK), n = 5; (3), modified HTK solution plus COP (mHTK + COP), n = 6; and (4), as a control five hearts were transplanted after cardioplegic arrest by the original HTK solution and a cold ischemia time of 3 h comparable to clinical routine procedure. RESULTS: After 14 h of preservation and orthotopic transplantation, cardiac functional recovery in mHTK + COP hearts was similar to control hearts, and improved compared to hearts of both other groups. Hemodynamics were significantly better in hearts preserved by mHTK + COP and in the control group compared to the HTK-hearts (P < 0.05), not significant compared to mHTK hearts (dp/dt(max) in % of preoperative +/- standard error of mean (SEM): mHTK + COP, 85 +/- 9; control, 85 +/- 10.5; mHTK, 59 +/- 14; HTK, 50 +/- 4). The cardiac output (CO) in % of preoperative was: mHTK + COP, 68 +/- 5.4; control, 64 +/- 4; mHTK, 44 +/- 2.7; HTK, 25 +/- 11. The ATP of left ventricular myocardium in mHTK + COP hearts at 14.7 +/- 1 micromol/g dry weight (DW) and in the control at 14.59 +/- 1.8 was higher compared to that in mHTK at 12.2 +/- 2.8 (P is non-significant (n.s.) versus mHTK + COP and control) and in HTK-hearts at 7.0 +/- 0.5 (P < 0.05 versus mHTK + COP and control). CK-MB in percent of CK showed no increase in either group. CONCLUSIONS: These data show that COP combined with a mHTK solution represents a potential alternative to complement currently used cold storage techniques for prolonged preservation periods.

Intracoronary shunt insertion prevents myocardial stunning in a juvenile porcine MIDCAB model absent of coronary artery disease.

OBJECTIVE: The relevance of regional LV myocardial ischemia/reperfusion induced by temporary left anterior descending (LAD) coronary artery occlusion during minimally invasive direct coronary artery bypass (MIDCAB) grafting is controversial. The purpose of our study was (1) to determine the impact of conventional LAD occlusion during left internal thoracic artery (LITA)-LAD anastomosis on regional LV myocardial ischemia and function, and (2) to evaluate if intra-LAD shunt insertion during LITA-LAD anastomosis prevents potential regional LV ischemia and dysfunction in a pig model. METHODS: In 20 anesthetized, mechanically ventilated pigs we performed LITA-LAD anastomosis on the beating heart without cardiopulmonary bypass during either 15 min LAD occlusion (occlusion-group; n = 10) or 15 min intra-LAD shunt insertion to maintain blood supply to the myocardium beyond the anastomosis (shunt-group; n = 10). Besides standard hemodynamics we determined the global and regional LV wall motion score index (WMSI) using epimyocardial echocardiography. To quantitate structural myocardial alteration we determined the inducible heat-shock protein-70 (HSP-70) in LV anterior wall myocardial biopsies. Data were recorded at baseline, at 15 min of LAD occlusion or shunt insertion, respectively, and at 30 min of reperfusion. At the end of the experiments we determined myocardial adenine nucleotide (ATP, ADP, AMP) and glycogen content. RESULTS: In both groups WMSI was not significantly different at 15 min LAD occlusion or shunt insertion, respectively, as compared to baseline. However, at 30 min reperfusion both global and regional WMSI demonstrated significant LV dysfunction in the occlusion-group, whereas LV function in the shunt-group remained normal. This was associated with higher myocardial HSP-70 expression in the occlusion-group (P < 0.05). Myocardial adenine nucleotide and glycogen contents were significantly better preserved in the shunt-group. CONCLUSIONS: Our data show that in a porcine MIDCAB model 15 min LAD occlusion and 30 min reperfusion result in significant myocardial stunning. In contrast, maintenance of LAD perfusion using intracoronary shunt insertion minimizes ischemia/reperfusion injury and prevents regional LV dysfunction. Although our experiments were conducted in healthy pig hearts absent from coronary artery disease, similar results may--at least partially--be expected in humans, and thus, intracoronary shunts could be a useful tool for myocardial protection during 'off-pump revascularization'.

Excellent recovery after prolonged heart storage by preservation with coronary oxygen persufflation: orthotopic pig heart transplantations after 14-hr storage.

BACKGROUND: Improvement of heart preservation is still the greatest challenge in preservation research. The unchanged severe restriction of acceptable storage periods of heart grafts since the beginning of clinical heart transplantation indicates that technical innovations are necessary if a substantial improvement is to be achieved. METHODS: Here, we present the results of hypothermic preservation using the innovative technique of coronary oxygen persufflation (COP). COP simply adds gaseous oxygen to hypothermic graft storage and requires only a "valve guard" for reversible closure of the aortic valve. Fourteen-hr preservation was followed by orthotopic transplantation and evaluations of functional as well as metabolic recovery. Mature pig hearts, a model with restricted preservation tolerance similar to the human heart, were used to guarantee the clinical relevance of this study. RESULTS: After 14-hr hypothermic storage, COP-preserved hearts were able to recover within 2 hr of cardiopulmonary bypass to a steady cardiovascular function without mechanical or pharmacologic support. The left ventricular pressure amplitude of mHTK-COP-preserved hearts as well as energy charge potential recovered to pregrafting values and the ventricular power output to 66%. Hearts simply stored in University of Wisconsin (UW), modified Bretschneider's histidine-tryptophan-ketoglurate (mHTK), or Euro-Flush with glutathione (EFG) solution had only limited recovery, with significantly lower ventricular power output of 18%, 29% or 30% of pregrafting controls on average. CONCLUSIONS: Fourteen-hr oxygenated pig heart preservation using COP results in optimal recovery. Storage preservation in solutions containing hyaluronidase (mHTK and EFG) results in higher recoveries as compared to UW solution, an effect that may support the excellent recovery after mHTK-COP preservation.

Coronary oxygen persufflation for long-term myocardial protection.

In human heart transplantation limited myocardial ischemia duration remains one of the most restricting factors. A new approach towards prolongation of this duration is the combination of cardioplegic arrest and continuous Coronary Oxygen Persufflation (COP) with gaseous oxygen. This technique, which is based on former experiments, was applied in pig hearts which we transplanted orthotopically after a hypothermic preservation time of 14 hours. For cardioplegic arrest we used either Euro-Flush glutathion solution (EFG; n=5), University of Wisconsin solution (UW; n=5), modified Bretschneider HTK cardioplegic solution (mHTK; n=6). In preliminary experiments all three solutions had shown equal cardioprotective qualities. Hearts of the mHTK group were submitted to continuous COP during storage (mHTK+COP). After 14 hours of preservation and orthotopic transplantation the mHTK+COP hearts showed significantly improved cardiac functional recovery compared to hearts preserved by simple cold storage techniques. Hemodynamics measured after 3 hours reperfusion were significantly better in the mHTK+COP group compared to EFG and UW: dp/dtmax in % of baseline+/-standard deviation (SD): 85+/-22, 65+/-26, 36+/-15, CO in % of baseline: 68+/-13, 35+/-8, 39+/-8. Postoperative preload recruitable stroke work in the mHTK+COP hearts was: 51.4+/-23.1 mmHg compared to preoperative: 57.3+/-17.2. ATP of left-ventricular myocardium in the mHTK+COP group: 14.7+2.1 micromol/g dry weight was significantly higher compared to EFG: 10.3+/-4.5 and UW: 5.9+/-3.2. CK-MB in percent of CK in all groups showed no increase during postoperative reperfusion. This study suggests that COP may present an effective complement to cold storage techniques currently used in heart transplantation. Prior to clinical application further investigations regarding long-term survival and endothelial function are required.

Minimal amounts of hyaluronidase in HTK or UW solution substantially improve the recovery of preserved hearts.

Rat hearts were preserved by simple storage for 18 h at 0-1 degree C and reperfused parabiotically with whole blood from a host rat. The preservation solutions used for flush perfusion and storage were the commercial solutions EuroCollins, HTK, or UW with or without adding 40 mg/l hyaluronidase or EuroFlush-Glutathione (EFG) solution, especially designed for prolonged heart storage. All solutions were filtered (0.45 micron) before use. The functional recovery was measured using a latex balloon in the left ventricle for LVP, dp/dt, and isotonic stroke volume. The metabolic recovery as well as the edema formation was determined from freeze-clamped myocardium at the end of reperfusion. In hearts preserved with hyaluronidase-containing solutions, the edema formation during reperfusion was reduced combined with an improvement in the coronary flow. Functional and metabolic recovery were improved in these hearts with significant increase in the stroke volume and ECP in all groups versus hearts preserved in the hyaluronidase-free basic solutions. The effectiveness of HTK preservation was significantly improved by hyaluronidase in all parameters measured in our study. The best functional and metabolic recovery was found in hearts preserved by HTK + H- or EFG-solution. Thus, preservation solutions containing hyaluronidase, especially HTK + H and EFG, seem best suited for the prolonged storage preservation of the heart.

Effectivity of freshly prepared or refreshed solutions for heart preservation versus commercial EuroCollins, Bretschneider's HTK or University of Wisconsin solution.

Rat hearts were preserved for 18 hr under totally ischemic storage conditions at 0-1 degree C with the commercially supplied EuroCollins, Bretschneider's HTK, and University of Wisconsin (UW) preservation solutions compared with our new preservation solution, Euro-Flush-glutathione solution, and a "refreshed" UW solution (UWG) with 3 mmol/L reduced glutathione added before use. Recovery of the organs was measured during 30 min of parabiotic reperfusion with whole blood of a host rat of the same inbred LEW strain, following an initial warm reflush for 5 min. Functional measurements were performed using a latex balloon in the left ventricle. The metabolic recovery was determined from the myocardium freeze-clamped at the end of reperfusion. The left ventricular pressure (LVP) amplitude during pacing to a heart rate of 300/min, as well as +dp/dtmax, -dp/dtmax, isotonic stroke volume, coronary flow, ATP, and ECP values, recovered significantly better after storage in Euro-Flush-glutathione solution (LVP: 63% of controls on average) compared with when the commercially available solutions were used (EuroCollins: 20%, HTK: 42% of controls in LVP). Hearts preserved in UW solution ViaSpan did not recover during the reperfusion period, when unfiltered solution was used. Filtered ViaSpan resulted in LVP recoveries of 38% of controls, while addition of reduced glutathione immediately before use (UWG) improved the effectivity of this solution significantly (LVP 63% of controls). Similar improvements were found for all other functional and metabolic parameters. Thus, the effectivity of UW solution ViaSpan depends upon extraction of the typical particles by a filtering procedure. Effectivity can be improved by a refreshment procedure with reduced glutathione immediately before use.