Although Holder pasteurization is the recommended method for processing breast milk, it does affect some of its nutritional and biological properties and is ineffective at inactivating spores. The aim of this study was to find and validate an alternative methodology for processing breast milk to increase its availability for newborn babies and reduce the financial loss associated with discarding milk that has become microbiologically positive. We prepared two series of breast milk samples inoculated with the Bacillus cereus (B. cereus) strain to verify the effectiveness of two high-pressure treatments: (1) 350 MPa/5 min/38 °C in four cycles and (2) cumulative pressure of 350 MPa/20 min/38 °C. We found that the use of pressure in cycles was statistically more effective than cumulative pressure. It reduced the number of spores by three to four orders of magnitude. We verified that the method was reproducible. The routine use of this method could lead to an increased availability of milk for newborn babies, and at the same time, reduce the amount of wasted milk. In addition, high-pressure treatment preserves the nutritional quality of milk.

• Publikační typ
• časopisecké články MeSH

Bacillus cereus is relatively resistant to pasteurization. We assessed the risk of B. cereus growth during warming and subsequent storage of pasteurized banked milk (PBM) in the warmed state using a predictive mathematical model. Holder pasteurization followed by storage below -18 °C was used. Temperature maps, water activity values, and B. cereus growth in artificially inoculated PBM were obtained during a simulation of manipulation of PBM after its release from a Human Milk Bank. As a real risk level, we chose a B. cereus concentration of 100 CFU/mL; the risk was assessed for three cases: 1. For an immediate post-pasteurization B. cereus concentration below 1 CFU/mL (level of detection); 2. For a B. cereus concentration of 10 CFU/mL, which is allowed in some countries; 3. For a B. cereus concentration of 50 CFU/mL, which is approved for milk formulas. In the first and second cases, no risk was detected after 1 h of storage in the warmed state, while after 2 h of storage, B. cereus concentrations of 102 CFU/mL were occasionally encountered. In the third case, exceeding the B. cereus concentration of 102 CFU/mL could be regularly expected after 2 h of storage. Based on these results, we recommend that post-pasteurization bacteriological analysis be performed as recommended by the European Milk Bank Association (EMBA) and using warmed PBM within 1 h after warming (no exceptions).

• Publikační typ
• časopisecké články MeSH

The authors present their contribution to the improvement of methods suitable for the detection of the freezing and thawing damage of cells of cryopreserved venous grafts used for lower limb revascularization procedures. They studied the post-thaw viability of cells of the wall of cryopreserved venous grafts (CVG) immediately after thawing and after 24 and 48 h culture at +37 °C in two groups of six CVG selected randomly for slow thawing in the refrigerator and rapid thawing in a water bath at +37 °C. The grafts were collected from multi-organ and tissue brain-dead donors, cryopreserved, and stored in a liquid nitrogen vapor phase for five years. The viability was assessed from tissue slices obtained by perpendicular and longitudinal cuts of the thawed graft samples using in situ staining with fluorescence vital dyes. The mean and median immediate post-thaw viability values above 70% were found in using both thawing protocols and both types of cutting. The statistically significant decline in viability after the 48-h culture was observed only when using the slow thawing protocol and perpendicular cutting. The possible explanation might be the "solution effect damage" during slow thawing, which caused a gentle reduction in the graft cellularity. The possible influence of this phenomenon on the immunogenicity of CVG should be the subject of further investigations.

• MeSH
• alografty diagnostické zobrazování   účinky léků   MeSH
• apoptóza účinky léků   MeSH
• dárci tkání MeSH
• dimethylsulfoxid farmakologie   MeSH
• fluorescenční barviva * MeSH
• konfokální mikroskopie metody   MeSH
• kryoprezervace metody   MeSH
• kryoprotektivní látky farmakologie   MeSH
• lidé MeSH
• optické zobrazování metody   MeSH
• transplantace cév metody   MeSH
• vena femoralis diagnostické zobrazování   účinky léků   MeSH
• vena saphena diagnostické zobrazování   účinky léků   MeSH
• viabilita buněk účinky léků   MeSH
• zmrazování * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: The aim of our study was to assess the impact of different thawing protocols on morphological changes arising in cryopreserved human saphenous vein grafts. METHODS: The study was performed in 12 saphenous vein grafts harvested in brain death donors. Storage in the vapor phase of liquid nitrogen for 3 or 5 years followed. Two thawing protocols were tested: slow thawing in a refrigerator at temperature +4°C for 2 hr and rapid thawing-in a water bath at +37°C. Grafts were processed for scanning electron microscopy. Comparisons of continuous parameters under study between experimental groups were performed using the t-test (age, cold ischemia time, exposure to cryoprotectant, time of storage, total thawing time, mean thawing rate, morphology scoring of thawed HSVG) and the median test (HSVG length). Categorical parameters (sex and blood group) were formally tested using the chi-square test. RESULTS: All samples were evaluated according to morphological changes and scored in terms of morphologically intact endothelium, confluent endothelium with structural inhomogeneity, disruption of the intercellular contacts, separation of the endothelial cells, complete loss of the endothelium, and damage of the subendothelial layers. There is no statistically significant difference between the sample sets at the significance level of 0.05. There was no association with donors' age, sex, and time of storage. CONCLUSIONS: Human cryopreserved saphenous vein grafts in our experimental work showed no difference in terms of structural deterioration of the endothelial surface and basal membrane depending on different thawing protocols used.

• MeSH
• časové faktory MeSH
• dospělí MeSH
• endoteliální buňky účinky léků   transplantace   ultrastruktura   MeSH
• kryoprezervace * MeSH
• kryoprotektivní látky farmakologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• odběr tkání a orgánů MeSH
• přežití tkáně MeSH
• vena saphena účinky léků   transplantace   ultrastruktura   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Introduction: The rate of thawing of cryopreserved human iliac arteries allografts (CHIAA) directly affects the severeness of structural changes that occur during this process. Method: The experiment was performed on ten CHIAA. The 10% dimethylsulphoxide in 6% hydroxyethyl starch solution was used as the cryoprotectant; all CHIAA were cooled at a controlled rate and stored in the vapor phase of liquid nitrogen (-194°C). Two thawing protocols were tested: (1) placing the CHIAA in a water bath at 37°C, and (2) the CHIAA were thawed in a controlled environment at 5°C. All samples underwent analysis under a scanning electron microscope. Testing of the mechanical properties of the CHIAA was evaluated on a custom-built single axis strain testing machine. Longitudinal and circumferential samples were prepared from each tested CHIAA. Results: Ultrastructural analysis revealed that all five CHIAA thawed during the thawing protocol 1 which showed significantly more damage to the subendothelial structures when compared to the samples thawed in protocol 2. Mechanical properties: Thawing protocol 1-longitudinal UTS 2, 53 ± 0, 47 MPa at relative strain 1, 27 ± 0, 12 and circumferential UTS 1, 94 ± 0, 27 MPa at relative strain 1, 33 ± 0, 09. Thawing protocol 2-longitudinal ultimate tensile strain (UTS) 2, 42 ± 0, 34 MPa at relative strain 1, 32 ± 0, 09 and circumferential UTS 1, 98 ± 0, 26 MPa at relative strain 1, 29 ± 0, 07. Comparing UTS showed no statistical difference between thawing methods. Conclusion: Despite the significant differences in structural changes of presented thawing protocols, the ultimate tensile strain showed no statistical difference between thawing methods.

• MeSH
• alografty účinky léků   fyziologie   MeSH
• arteria iliaca účinky léků   fyziologie   MeSH
• dimethylsulfoxid farmakologie   MeSH
• dospělí MeSH
• kryoprezervace metody   MeSH
• kryoprotektivní látky farmakologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Dimethyl sulfoxide (DMSO) is the cryoprotectant of choice for most animal cell systems since the early history of cryopreservation. It has been used for decades in many thousands of cell transplants. These treatments would not have taken place without suitable sources of DMSO that enabled stable and safe storage of bone marrow and blood cells until needed for transfusion. Nevertheless, its effects on cell biology and apparent toxicity in patients have been an ongoing topic of debate, driving the search for less cytotoxic cryoprotectants. This review seeks to place the toxicity of DMSO in context of its effectiveness. It will also consider means of reducing its toxic effects, the alternatives to its use and their readiness for active use in clinical settings.

• MeSH
• dimethylsulfoxid farmakologie   MeSH
• kryobiologie MeSH
• kryoprezervace metody   MeSH
• kryoprotektivní látky farmakologie   MeSH
• lidé MeSH
• tkáňové inženýrství MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

• Publikační typ
• abstrakt z konference MeSH

OBJECTIVES AND DESIGN: At the present time there are two waiting list for patients with vascular prosthetic infection indicated for arterial transplantation in the Czech Republic. The inclusion of each patient for cold-stored or cryopreserved arterial transplantation is the preference of indicating surgeon. In this experimental work we studied the immunogenicity of rat aortal allografts treated by our new clinical cryopreservation/slow thawing protocol. MATERIAL AND METHODS: Brown-Norway (BN) (N = 6, 203-217 g) or Lewis (LEW) (N = 6, 248-254 g) abdominal aortal grafts treated in accordance with our new clinical cryopreservation/slow thawing protocol were orthotopically transplanted to Lewis recipients (N = 12, 191-245 g). Aortal wall histology and infiltration by recipient immune cells, as well as donor specific anti MHC class I and II antibodies in recipient serum were studied in both isografts and allografts on day 30 postransplant. Core data of cryopreserved allografts were compared to our previous data of cold-stored aortal allografts treated in accordance with our clinical cold-storage protocol. RESULTS: Cryopreserved allografts showed regular morphology of aortal wall with clear differentiation of all three basic anatomical layers on day 30 postransplant. Intimal layer showed no hyperplasia, luminal surface was covered by endothelial cells. No statistical difference was observed in tunica media thickness between isografts and allografts. The medial layer showed no necrosis, shrinkage or immunoglobuline G deposition in any experimental group. The adventitial infiltration by immune cells was significantly higher (P<0.05) in allografts. Cryopreserved allografts showed significant lower activation of both cell- and antibody mediated immunity compared to historical data of cold-stored allografts. CONCLUSION: Aortal wall histology of rat allografts treated by our new standardized clinical cryopreservation/slow thawing protocol was comparable to that of the cryopreserved isografts on day 30 posttranspant. The immunogenicity of cryopreserved aortal allografts was significantly lower compared to that of cold-stored aortal allografts.

• MeSH
• alografty fyziologie   MeSH
• aorta transplantace   MeSH
• arterie transplantace   MeSH
• homologní transplantace metody   MeSH
• kryoprezervace metody   normy   MeSH
• krysa rodu rattus MeSH
• modely u zvířat MeSH
• potkani inbrední BN MeSH
• potkani inbrední LEW MeSH
• rejekce štěpu imunologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH

BACKGROUND: Vascular allotransplantations are performed worldwide in selected patients suffering from vascular prosthesis infection or critical limb ischemia. Either fresh or cryopreserved vascular allograft may be used. OBJECTIVES: In various points, we address several aspects (allograft procurement, cryopreservation and transplantation technique) of the program of vascular allotransplantations in the Czech Republic. MATERIAL AND METHODS: Vascular grafts retrieval has been done within multiorgan harvests using no-touch technique. Very short time of cold ischemia is achieved due to close cooperation with Tissue Establishment where the following processing of cryopreservation is performed. Meeting all necessary quality criteria is a prerequisity for releasing grafts for clinical application. Standardized thawing protocol and surgical handling aims to minimize microfractures before implantation. RESULTS: Based on experimental and clinical work, the first validation of cryopreserved arterial and venous grafts for clinical use was performed between 2011 and 2013 in the Czech Republic. The developement of storage of vascular tissue in banks was stimulated in 2000-2010 by the issue of EU directives and national harmonized norms, aimed at assurance of high quality and safety of cells and tissues used for transplantations in humans. CONCLUSIONS: There are several crucial moments affecting final quality, including graft retrieval within a multiorgan harvest, short ischemic time, cryopreservation and thawing technique used. The recommended surgical handling during implantation may also affect results and graft-related complications.

• MeSH
• alografty MeSH
• cévní protézy * MeSH
• cévy fyziologie   transplantace   MeSH
• homologní transplantace metody   MeSH
• kryoprezervace * metody   MeSH
• lidé MeSH
• tkáňové banky MeSH
• transplantace cév metody   MeSH
• získávání tkání a orgánů * statistika a číselné údaje   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH

Základní informace: Popisujeme soubor čtyř pacientů s kritickou ischemií končetiny a absencí autologních náhrad (konduitů), kteří byli ošetřeni vytvořením kompozitního (s anastomózou „end-to-end“) alogenního bypassu z čerstvého nebo kryoprezervovaného štěpu z vena saphena (saféna). Použití této techniky může být nezbytné při vytváření femoropedálního bypassu, kdy jde o extrémní situaci charakterizovanou příliš krátkými či nedostatečně kvalitními žilními štěpy. Podobně dlouhá rekonstrukce vyžaduje dvě dárcovské safény. Kasuistiky: U čtyř pacientů bylo indikováno vytvoření „I-kompozitního“ femoropedálního bypassu s využitím čerstvého žilního alograftu. Jeden kompozitní štěp se uzavřel s odstupem čtyř měsíců od operace, jeden za 21 měsíců od operace a další dva zůstaly průchodné po dobu sledování s mediánem 23 měsíců. U žádného z pacientů nebylo po dobu sledování nutno provést amputaci. Závěr: Střednědobého udržení průchodnosti rekonstrukce lze uspokojivě dosáhnout za předpokladu kompatibility v systému ABO, krátké chladné ischemie štěpu, adekvátní imunosupresivní terapie a dodržení správného protokolu pro sledování nemocných po transplantaci cévního alograftu. Cévní chirurgové by měli mít na paměti možnost uplatnění této techniky zejména u diabetiků s kritickou ischemií končetiny a okluzí bércových tepen.

Background: We report a series of four patients with critical limb ischaemia and lack of autologous conduits, treated with composite (end-to-end anastomosis) allogenic fresh/or cryopreserved saphenous vein bypass grafting. This technique may be necessary in case of femoro-pedal artery bypass grafting, which is an extreme situation if there is shortage in length or inadequate quality of the venous allograft. Such a long reconstruction requires two donor saphenous veins. Case report: Four patients were indicated for “I-composite” fresh venous allograft for femoro-pedal bypass grafting. One composite graft occluded 4 months postoperatively, one 21 months postoperatively, two other remained patent with median follow-up 23 months. No amputation was required in any of the patients during the follow-up. Conclusion: Midterm patency of the reconstruction may be satisfactory provided that the ABO compatibility, short cold ischaemia time of the graft, adequate immunosuppressive therapy and proper follow-up protocol of the patient after vascular allograft transplantation are observed. Surgeons should keep in mind possibility of this technique mainly in diabetic patients with critical limb ischaemia and occluded crural vessels.

• MeSH
• alografty chirurgie   MeSH
• bérec chirurgie   krevní zásobení   MeSH
• cévy - implantace protéz metody   MeSH
• kryoprezervace MeSH
• lidé MeSH
• noha (od hlezna dolů) chirurgie   krevní zásobení   MeSH
• onemocnění periferních arterií * chirurgie   klasifikace   MeSH
• senioři MeSH
• vena saphena transplantace   MeSH
• výsledek terapie MeSH
• získávání tkání a orgánů MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• Publikační typ
• kazuistiky MeSH