In this brief review, some key issues related to vitreous cryopreservation of living tissues (natural or engineered), including cells, embryos, tissues, organs, and engineered tissues, are outlined. The principle of vitreous cryopreservation for the biological activity and functionality is demonstrated. The procedures of cooling/ rewarming, composition and function of optimal cryoprotectants, and their effects on bioproducts are described. Vitrification could, therefore, prove to be a useful and effective method of bioproduct cryopreservation for a long period of time, particularly for organized tissues and organs. However, the toxicity of the cryoprotective agents and the devitrification occurring during the rewarming process need additional investigations. Several key areas of research on vitrification are also addressed.
Cryopreservation ; methods ; Cryoprotective Agents ; pharmacology ; Dimethyl Sulfoxide ; pharmacology ; Humans ; Organ Preservation ; methods ; Tissue Preservation ; methods

This study was purposed to evaluate the effect of different lyophilizing protectants including human albumin, glucan, polyvinyl pyrrolidone and glycerine on lyophilized trehalose-loading red blood cells (RBC), then to screen the optimal lyophilizing protectant. The RBC were incubated in 800 mmol/L concentration of trehalose solution at 37°C for 7 hours, and washed 3 times with PBS solution to obtain the trehalose-loading RBC. The trehalose-loading RBC in control group were directly lyophilized without lyophilizing protectants, the trehalose-loading RBC in the experimental group were mixed with Lyophilizing protectants. The samples of 2 groups were kept at room temperature for 30 minutes, pre-frozen at -80°C for 24 hours, then lyophilized in freeze-dryer for 24 hours. Finally the samples were quickly rehydrated by 6% HES at 37°C. The recovery rate and hemolysis rate of hemoglobin were detected by using cyanohemoglobin detection kit. The water content of unhydrated samples were detected at the same time. The results showed that when the moisture content of sample was 3% - 5%, the recovery rate of hemoglobin in control group was 33.57 ± 2.89%, and that in experimental group was 51.15 ± 1.98%, there was statistically significant difference between the control and experimental group (P < 0.05). When the different concentration of dextran solution was chosen as protectants, the recovery rate of hemoglobin of lyophilized RBC was obviously lower. The higher concentration of dextran, the better the recovery rate. The recovery rate of hemoglobin was 22.15 ± 4.12% when the concentration of dextran was 36%, there were statistically significant difference between the two groups (P < 0.05). When the different concentration of polyvinyl pyrrolidone (PVP) solutions was chosen as protectants, especially the concentration below 40%, the recovery rate of hemoglobin of lyophilized RBC was significantly belower than the control group, there was statistically significant difference between the two groups (P < 0.05). When 10% glycerol was used as protectants, the recovery rate of hemoglobin was 3.93 ± 1.80%. There was also statistically significant difference between the two groups (P < 0.05). It is concluded that human serum albumin shows an important protective effect on the lyophilization of the trehalose-loading red blood cells. The dextran and PVP at the concentration lower than 40% can decrease the protective effect of trehalose in cells. Glycerol can not be chosen as protectant for lyophilized trehalose-loading red blood cells.
Blood Preservation ; methods ; Cryoprotective Agents ; pharmacology ; Erythrocytes ; drug effects ; Freeze Drying ; methods ; Humans ; Trehalose ; pharmacology

Trehalose, as a nonreducing disaccharide, plays a role in protecting the cytoactivity when the cells is freezing, drying or lyophilization. It has been a biomembrane protectant applied to lyophilization of human blood cells (platelets and erythrocytes), and from which astonishing results have been obtained. Having powerful hydration, distinctive vitrification transform and crystal transform and unique resistance of high temperature and humidification, trehalose is thought of a preferred protectant in the study of cell preservation. In recent years, people concerned trehalose on its protective mechanism, experimental means of transit trehalose to mammal cells and the mechanism of loading in red blood cells. The above aspects were briefly summarized in this article.
Blood Preservation ; methods ; Cryoprotective Agents ; pharmacology ; Erythrocytes ; cytology ; drug effects ; Freeze Drying ; Humans ; Trehalose ; pharmacology

OBJECTIVETo investigate the effects of cryoprotectant with glycerol and the freezing-thawing procedure on the motility of

METHODSThe motion characteristics of human sperm from 18 selected specimens were assessed by computer-assisted human sperm. sperm analysis before and after adding hyper-osmolarity cryoprotectant with glycerol and the freeze-thaw procedure, and the data were evaluated in pairs.

RESULTSThe adding of cryoprotectant caused an increase in the proportion of rapid linear motile sperms (P < 0.05) and sperm velocities, including VCL, VSL and VAP (P < 0.005). But no changes were observed in the proportion of progressively motile sperms and sperm motility. Compared with the data from pre-frozen samples, velocities of post-thawed sperms and the percentage of motile sperms in each grade significantly declined (P < or = 0.01), so did ALH, while WOB, LIN and STR remained unchanged (P > 0.05). Significant differences in ALH, WOB, LIN and STR were observed only in comparison between the post-thawed and pre-treated samples.

CONCLUSIONThe number of Grade a sperms and sperm velocity increased after adding hyper-osmolarity cryoprotectant with glycerol. The sperm motile potential was impaired and even entirely destroyed in some cases by cryodamage. The influences on the sperm motion were differently induced in freezing-thawing procedures.

Cryopreservation ; Cryoprotective Agents ; pharmacology ; Freezing ; Glycerol ; pharmacology ; Humans ; Male ; Semen Preservation ; Sperm Motility ; drug effects

Ovarian cryopreservation can save a large number of germ cells during the cryogenic process, and can restore ovarian endocrine function and ovulation potential after thawing and transplantation, which is an ideal way to retain fertility for patients with cancer. Many factors influence the effect of ovarian cryopreservation, like cryoprotectant (CPA), frozen carrier, cortical block size and freezing procedure. An efficient and standard transplantation procedure is needed to develop for further clinical application and scientific research. In order to optimize this technology, we analyzed different factors to improve the recovery of ovarian function after freezing during ovarian cryopreservation.
Cryopreservation ; methods ; Cryoprotective Agents ; pharmacology ; Female ; Humans ; Ovary ; anatomy & histology ; physiology

Organ transplantation is an effective approach for the treatment of end-stage organ failures. Currently, the donor organs used for clinical transplantation are all preserved at above-zero temperatures. These preservation methods are well-established and simple but the storage time lasts for only 4-12 h. Some researchers tried to extend the organ storage time by improving protectant and HLA matching to raise the use of stored organs and prolong the long-term survival of organs. These efforts still fall short of the clinical demand for organ transplantation. Moreover, a great many organs were wasted due to limited storage time, HLA mismatch, patients' conditions or distance involved. Therefore, preserving organs for several weeks or even months and establishing Organ Bank are the tough challenges and have become a shared goal of global scholars. This article reviews some issues involved in the cryopreservation of organs, such as use of cryoprotecting agents, freezing and thawing methods in the cryopreservation of hearts, kidneys and other organs.
Cryopreservation ; methods ; Cryoprotective Agents ; adverse effects ; pharmacology ; Humans ; Organ Preservation ; methods

OBJECTIVETo investigate the effects of different proportions of cryoprotectant to seminal plasma on the motility of post-thaw human sperm.

METHODSDifferent proportions of cryoprotectant to seminal plasma (1:1 and 1:3) were used for freezing sperm, and the forward movement and total motility rates of the frozen-thawed sperm were compared.

RESULTSThe forward movement and total motility rates were (58.60 +/- 5.57)% and (66.17 +/- 5.24)% before cryopreservation. The 1:1 proportion achieved post-thaw forward movement and total motility rates of (40.53 +/- 8.97)% and (51.23 +/- 9. 30)%, while the 1:3 (44.7 +/- 8.67)% and (51.50 +/- 7.40)%, respectively. Significantly decreased sperm motility was observed after cryopreservation (P < 0.05). Statistically significant differences were found in the forward movement but not in the total motility of the frozen-thawed sperm between the two proportions.

CONCLUSIONCryopreservation causes obvious damage to human sperm. Higher proportion of cryoprotectant to seminal plasma (1:3) can improve the forward movement of post-thaw sperm as compared with the lower one (1:1).

Adult ; Cryoprotective Agents ; adverse effects ; pharmacology ; Freezing ; Humans ; Male ; Semen ; drug effects ; Sperm Motility ; drug effects

Lyophilization is the best preservation of platelets at present. Lyophilized platelets could be stored in circumstance temperature, small space and be transported with portability, but lyophilizing may damage platelet membrane, denature protein and induce platelet activation. The chain of events including freezing and drying that lead to activation are related to the phase transition of platelet membrane lipid. Application of protectants in accordance with damage mechanism of lyophilization could alleviate the damnification. Human platelets can be preserved using lyophilization successfully in the presence of trehalose in laboratory presently, that retains normal functions and activities after rehydration. These findings suggest that the application of lyophilized platelets in transfusion medicine will be carried out in future. In this paper damage effect of lyophilization on platelets, application of cryoprotectants in the platelet lyophilization and experimental studies on platelet cryopreservation were reviewed.
Blood Platelets ; physiology ; Blood Preservation ; Cryoprotective Agents ; pharmacology ; Freeze Drying ; Humans

In the 1960s, sperm cryopreservation was developed as a method to preserve fertility. Currently, techniques for the cryopreservation of human spermatozoa have been widely used in assisted reproduction. However, although sperm cryobiology has made notable achievements, the optimal method for the recovery of viable spermatozoa after cryopreservation remains elusive. Postthawing sperm quality can be affected by cryoprotectants, ice formation, storage conditions, and osmotic stress during the freezing process. This review discusses recent advances in different cryopreservation techniques, cryoprotectants, and freezing and thawing methods during cryopreservation and new indications for the use of cryopreserved spermatozoa.
Humans ; Male ; Semen Preservation/methods* ; Sperm Motility ; Semen ; Cryopreservation/methods* ; Spermatozoa ; Cryoprotective Agents/pharmacology*

This study was aimed to search for effective cryoprotectants and freezing methods used in cord blood bank (CBB) for cryopreservation of cord blood hematopoietic stem cells. The non-programmed group using 8% final concentration of dimethyl sulfoxide (DMSO) and 5% final concentration hydroxyethyl starch (HES) (molecular weight 120,000) as protectants and group of conventional of programmed controller method using 10% DMSO only as cryoprotectant in cryopreservation of cord blood hematopoietic stem cells were compared. In each of the two groups, 15 cord blood units were used. In non-programmed group, cord blood units put in -80 degrees C refrigerator for 24 hours as a transitional step before deep-freezing in liquid nitrogen, when both of DMSO and HES had been added. The recoveries of the nuclear cells number, the yield of granulocyto-macrophage colony forming units (CFU-GM) and the cells viability in cord blood units before preservation and after thawing were tested for both methods. The results showed that no significant difference was found in above assays between two groups. The clinical application results also showed that hematopoietic engraftment rates after infusion were similar in both groups. It is concluded that the non-programmed method by -80 degrees C refrigerator as a transitional step and using the combined two protectants seems simple in operation and effective in clinical transplantation as well as the conventional programmed method.
Cryopreservation ; Cryoprotective Agents ; pharmacology ; Dimethyl Sulfoxide ; pharmacology ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans ; Hydroxyethyl Starch Derivatives ; pharmacology