The aim of this investigation was to determine the optimal storage medium for testicular hypothermic transportation and identify the ideal concentration for the application of the protective agent 5-aminolevulinic acid (5-ALA). Furthermore, this study aimed to explore the underlying mechanism of the protective effects of 5-ALA. First, we collected and stored mouse testicular fragments in different media, including Hank's balanced salt solution (HBSS; n = 5), Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12 (DMEM/F12; n = 5), and alpha-minimum essential medium (αMEM; n = 5). Storage of testicular tissue in HBSS preserved the integrity of testicular morphology better than that in the DMEM/F12 group ( P < 0.05) and the αMEM group ( P < 0.01). Testicular fragments were subsequently placed in HBSS with various concentrations of 5-ALA (0 [control], 1 mmol l -1 , 2 mmol l -1 , and 5 mmol l -1 ) to determine the most effective concentration of 5-ALA. The 2 mmol l -1 5-ALA group ( n = 3) presented the highest positive rate of spermatogonial stem cells compared with those in the control, 1 mmol l -1 , and 5 mmol l -1 5-ALA groups. Finally, the tissue fragments were preserved in HBSS with control ( n = 3) and 2 mmol l -1 5-ALA ( n = 3) under low-temperature conditions. A comparative analysis was performed against fresh testes ( n = 3) to elucidate the underlying mechanism of 5-ALA. Gene set enrichment analysis (GSEA) for WikiPathways revealed that the p38 mitogen-activated protein kinase (MAPK) signaling pathway was downregulated in the 2 mmol l -1 5-ALA group compared with that in the control group (normalized enrichment score [NES] = -1.57, false discovery rate [FDR] = 0.229, and P = 0.019). In conclusion, these data suggest that using 2 mmol l -1 5-ALA in HBSS effectively protected the viability of spermatogonial stem cells upon hypothermic transportation.
Male ; Animals ; Testis/cytology* ; Aminolevulinic Acid/pharmacology* ; Mice ; Organ Preservation/methods* ; Organ Preservation Solutions/pharmacology* ; Cryopreservation/methods*

OBJECTIVETo investigate the protective effect of luteolin on isolated rat heart in hypothermic preservation.

METHODSForty male SD rats were randomly divided into 4 groups (n = 10): control group, luteolin low-dose group (7.5 micromol/L), middle-dose group (15 micromol/L) and high dose group (30 micromol/L). Langendorff model of isolated rat heart was used. After 30 min basal perfusion, the hearts were stored in University of Wisconsin solution (UW solution) at 4 degrees C with luteolin (7.5, 15 and 30 micromol/L) or without luteolin for 12 h and followed by 60 min reperfusion. The recovery of cardiac contractile and diastolic function, coronary flow (CF), creatine kinase (CK) leakage in the coronary effluent, myocardial water content were determined. The myocardial ultrastructure was also observed.

RESULTSThe results revealed that luteolin improved the recovery of left ventricular peak systolic pressure and +/- dp/dtmax dose-dependently and increased coronary flow. The leakage of creatine kinase in the coronary effluent was significantly reduced in luteolin-added hearts. Impairment of myocardial ultrastructure after 12 h hypothermic preservation was obviously alleviated in hearts luteolin-added group compared with that in control group. There were no differences between the groups in myocardial water contents.

CONCLUSIONLuteolin as a supplementation in cardiac preservation solution can significantly improve the hypothermic preservation effects on rat heart and have myocardial protection effect, especially in luteolin-added with 30 micromol/L.

Animals ; Cryopreservation ; In Vitro Techniques ; Luteolin ; pharmacology ; Male ; Myocardium ; Organ Preservation ; methods ; Organ Preservation Solutions ; Rats

BACKGROUNDThe composition of the lung preservation solution used in lung graft procurement has been considered the key to minimize lung injury during the period of ischemia. Low-potassium dextran glucose (LPDG), an extracellular-type solution, has been adopted by most lung transplantation centers, due to the experimental and clinical evidences that LPDG is superior to intracellular-type solutions. Ulinastatin has been shown to attenuate ischemia-reperfusion (I/R) injury in various organs in animals. We supposed that the addition of ulinastatin to LPDG as a flushing solution, would further ameliorate I/R lung injury than LPDG solution alone.

METHODSTwelve male New Zealand white rabbits were randomly divided into 2 groups. Using an alternative in situ lung I/R model, the left lung in the control group was supplied and preserved with LPDG solution for 120 minutes. In the study group 50,000 U/kg of ulinastatin was added to the LPDG solution for lung preservation. Then re-ventilation and reperfusion of the left lung were performed for 90 minutes. Blood gas analysis (PaO₂, PaCO₂), mean pulmonary artery pressure (MPAP) and serum TNF-α level were measured intermittently. The pulmonary water index (D/W), tissue myeloperoxidase (MPO) activity, tissue malondialdehyde (MDA) content and morphologic changes were analyzed.

RESULTSThe study group showed significantly higher PaO₂ and lower MPAP at the end of reperfusion. Serum TNF-α level, left lung tissue MPO and MDA in the study group were significantly lower than those in the control group. D/W and pathologic evaluation were also remarkably different between the two groups.

CONCLUSIONSThis study indicated that better lung preservation could be achieved with the use of an ulinastatin modified LPDG solution. Ulinastatin further attenuated lung I/R injury, at least partly by reducing oxidative reactions, inhibiting the release of inflammatory factors and neutrophils immigration.

Animals ; Glycoproteins ; pharmacology ; Lung ; drug effects ; metabolism ; Lung Transplantation ; Male ; Organ Preservation Solutions ; chemistry ; pharmacology ; Rabbits ; Random Allocation ; Reperfusion Injury ; prevention & control

OBJECTIVETo investigate the effect of diazoxide (DE) on the myocardial ultrastructure and opening of maitochondrial permeability transition pore (MPTP) in donor rat heart suffered from long-term hypothermic preservation.

METHODSThe Langendorff model of isolated rat heart was used. The hearts were stored in 4 degrees C Celsior solution containing different concentration of DE (15, 30, or 45 micromol/L) for 9 h followed by 60 min of reperfusion. The recovery of rate-pressure product (RPP) was observed. The opening of MPTP and myocardial mitochondria ultrastructure were also evaluated.

RESULTS(1) As compared with the celsior solution preserved group, DE (30 micromol/L) increased recovery of RPP during reperfusion and inhibited the opening of MPTP. DE also alleviated the myocardial mitochondrial ultrastucture damage induced by long-term hypothermic preservation. (2) The above effects of DE were attenuated by a mitoK(ATP) channel inhibitor 5-hydroxydecanoate and a MPTP opener atractyloside.

CONCLUSIONIn the donor rat heart, DE protects myocardial mitochondria ultrastructure against long-term hypothermic preservation injury via inhibiting the opening of MPIP.

Animals ; Cryopreservation ; Diazoxide ; pharmacology ; Heart ; In Vitro Techniques ; Male ; Mitochondria, Heart ; physiology ; ultrastructure ; Mitochondrial Membrane Transport Proteins ; drug effects ; metabolism ; Organ Preservation Solutions ; pharmacology ; Potassium Channels ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo evaluate the effects of tannic acid (TA) treatment on the physico-chemical properties of glutaraldehyde (Glut)-fixed bovine jugular vein (BJV).

METHODSFresh BJVs were treated with Glut or Glut/TA, respectively. The shrinkage temperature, resistance to collagenase or elastase digestion, bio-mechanical properties, and molecular structure of these prepared BJVs were evaluated by Fourier transform infrared spectroscopy.

RESULTSTA treatment resulted in higher shrinkage temperature (P < 0.01), higher resistance to collagenase or elastase digestion (P < 0.01), slightly increased tensile strength (P < 0.01), and elongation at break (P < 0.05) in Glut/TA BJV walls when compared with those of Glut group. Chemical bonds existed between TA and JBV tissue.

CONCLUSIONTA treatment can effectively improve the physicochemical properties of Glut-fixed BJVs.

Animals ; Cattle ; Chemical Phenomena ; drug effects ; Elasticity ; drug effects ; Glutaral ; pharmacology ; Jugular Veins ; pathology ; Organ Preservation Solutions ; pharmacology ; Tannins ; pharmacology ; Tensile Strength ; drug effects ; Tissue Fixation ; methods ; Tissue Preservation ; methods

BACKGROUNDThe University of Wisconsin colloid based preserving solution (UW solution) is the most efficient preserving solution for multiorgan transplantation. Unfortunately, unavailability of delayed organ preserving solutions hindered further progression of cardinal organ transplantation in China. In this study, we validated an organ preserving Changzheng Organ Preserving Solution (CZ-1 solution) and compared it with UW solution.

METHODSA series of studies were conducted on how and how long CZ-1 solution could preserve the kidneys, livers, hearts, lungs and pancreas of New Zealand rabbits and SD rats. Morphology of transplanted organs was studied by visible microscopy and electron microscopy; biochemical and physiological functions and the survival rate of the organs during prolonged cold storage were studied.

RESULTSThere was no significant difference between CZ-1 and UW solutions in preserving the kidneys, livers, hearts or lungs of rabbits; kidneys, livers, intestinal mucosa or pancreases of SD rats or five deceased donors' testicles. In some aspects, such as preserving rabbits' hearts, rats' intestinal mucosa and pancreases, the effect of CZ-1 solution was superior to UW solution. CZ-1 could safely preserve kidneys for 72 hours, livers for 24 hours, hearts for 18 hours and lungs for 8 hours for SD rats. Twelve kidneys preserved in cold CZ-1 solution for 22 - 31 hours were transplanted successfully and the mean renal function recovery time was (3.83 +/- 1.68) days.

CONCLUSIONSCZ-1 solution is as effective as UW solution for organ preservation. The development of CZ-1 solution not only reduces costs and improves preservation of organs, but also promotes future development of organ transplantation in China.

Adenosine ; pharmacology ; Allopurinol ; pharmacology ; Animals ; China ; Glutathione ; pharmacology ; Heart ; drug effects ; physiology ; Heart Transplantation ; methods ; Insulin ; pharmacology ; Intestine, Small ; drug effects ; physiology ; Kidney ; drug effects ; physiology ; Kidney Transplantation ; methods ; Liver ; drug effects ; physiology ; Liver Transplantation ; methods ; Lung ; drug effects ; physiology ; Lung Transplantation ; methods ; Male ; Organ Preservation ; economics ; methods ; Organ Preservation Solutions ; pharmacology ; Pancreas ; drug effects ; physiology ; Pancreas Transplantation ; methods ; Pharmaceutical Solutions ; pharmacology ; Rabbits ; Raffinose ; pharmacology ; Testis ; drug effects ; physiology

The effects of oxygen partial pressure on cryopreservation of the cells with organ preservation solution were explored. Hypoxic UW solution was made by purging the UW solution with argon. The pig proximal tubule epithelial cells (LLC-PK1 cells) were cryopreserved in hypoxic UW solution (Ar-UW group) or standard UW solution (UW group) at 4 degrees C for 48 h. Trypan blue staining and LDH detection were performed to evaluate the injury of the cells. The results showed that the oxygen partial pressure in Ar-UW group was significantly declined from 242+/-6 mmHg to 83+/-10 mmHg. After cryopreservation at 4 degrees C for 48 h, LDH leakage rate and Trypan blue-stained rate in Ar-UW group were (11.3+/-3.4)% and (10.5+/-4.7)%, respectively, which were significantly lower than in UW group [(49.5+/-6.9)% and (47.6+/-9.3)% respectively, both P<0.01]. It was concluded that lower oxygen partial pressure of UW solution was more beneficial to the cryopreservation of LLC.
Adenosine ; Allopurinol ; Cell Hypoxia ; Cell Line ; Cryopreservation ; Cryoprotective Agents ; Epithelial Cells/*cytology ; Glutathione ; Insulin ; Kidney Tubules, Proximal/cytology ; Organ Preservation Solutions ; Oxygen/pharmacology ; Raffinose ; Swine ; Tissue Preservation/methods

OBJECTIVETo investigate whether the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) opener diazoxide as an additive to cardioplegia solution could enhance myocardial protection during hypothermic preservation of the rat heart.

METHODSThe Langendorff model of isolated rat heart was used. After equilibrium, the hearts were stored in Celsior cardioplegia solution at 4 degree with or without supplement of diazoxide for 3 or 8 h followed by 60 minutes reperfusion. The recovery of cardiac contractile function, myocardial enzyme leakage in the coronary effluent, and myocardial water content were determined. The myocardial ultrastructure was also observed.

RESULT(1) Treatment of diazoxide improved the recovery of left ventricular developed pressure and decreased the leakage of myocardial enzymes, lactate dehydrogenase (LDH) and creatine kinase (CK), at the 2nd and 4th minute of reperfusion of rat heart after hypothermic preservation for 3 h. (2) After hypothermic preservation for 8 h, diazoxide improved the recovery of left ventricular developed pressure and decreased the leakage of myocardial enzymes (LDH, CK and glutamic oxalic transaminase) during reperfusion. Moreover, left ventricular end-diastolic pressure was significantly lower in diazoxide-treated hearts than that of hearts in Celsior solution. (3) Diazoxide significantly decreased the water content of myocardium and increased coronary flow of the hearts compared with those in control after hypothermic preservation for 8 h. (4) Impairment of myocardial ultrastructure after 8 h hypothermic preservation was alleviated in hearts treated with 30 mol/L diazoxide. (5) The cardiac effects of 30 mol/L diazoxide were attenuated by a mitoK(ATP) blocker 5-hydroxydecanoate (100 micromol/L).

CONCLUSIONDiazoxide as a supplementation in cardioplegia solution could enhance myocardial protection during hypothermic heart preservation via opening of mitochondrial K(ATP) channel.

Animals ; Cardioplegic Solutions ; Cryopreservation ; Diazoxide ; pharmacology ; Heart ; Male ; Organ Preservation ; Organ Preservation Solutions ; pharmacology ; Potassium Channels ; drug effects ; Rats ; Rats, Sprague-Dawley

Drugs, Chinese Herbal ; pharmacology ; Heart ; Humans ; Liver ; Organ Preservation Solutions ; Panax ; Pancreas ; Salvia miltiorrhiza

Generally, mammalian cells and living tissues can be cryopreserved in a frozen state at very low temperatures over a long storage term. The survival rate of cell suspensions is often acceptable however, living tissues suffer a variety of injuries. In this paper, it was demonstrated that the addition of polyphenols extracted from green tea to conventional cell culture medium and tissue compatible liquid, can control cell proliferation and also preserve tissues for several months at ordinary room temperature, including such tissues as blood vessels, cartilage, islet cells and corneas. This protocol allows a non-frozen living tissue bank to be established using the preservation fluid described.
Animals ; Flavonoids/*pharmacology ; Humans ; Organ Preservation Solutions/*pharmacology ; Phenols/*pharmacology ; Tea/*chemistry ; *Tissue Banks ; *Tissue Preservation ; *Tissue Transplantation ; Transplantation, Homologous