The Improvement of Cell Viability due to Dilution and Removal of DMSO in Thawing of Stem Cells

PURPOSE: Cryopreserved stem cells from cord blood are usually infused with Dimethyl Sulfoxide (DMSO) immediately after thawing. However, this process may cause cell damage due to osmotic shock, and the administration of DMSO may also cause toxic effects. We studied a new method of increasing cell viability by stabilizing osmolarity by adding dextran 40 and washing out DMSO. METHODS: Thirty-five samples of cord blood were studied. RBCs were removed in 10% pentastarch. The cells were mixed with DMSO of 5, 10 and 20% each, and stored at -80 degree C. Cryopreserved cells were thawed and then diluted with dextran 40. DMSO was removed afterwards. The cell viability, osmolarity and colony forming capacity in this new thawing method were compared with the control group done by conventional methods. RESULTS: The recovery rate of WBC after RBCs separation was 92.06% but the contamination rate of RBC was still high(29.90%). The concentration of DMSO significantly affected the survival of WBCs(P<0.05). The osmolarity was 330+/-17.7mOsm/L before freezing, 1,457+/-508.7mOsm/L after thawing prior to dilution and 811+/-199.6mOsm/L after dilution, suggesting that the dilution process was effective in reducing osmolarity. The number of viable cells increased from 6.01+/-1.61(x103/L) to 7.16+/-1.48(x103/L) after dilution but was not significant. The number of CFU-C was increased from 5.82+/-4.19(/105) to 7.58+/-3.16(/105) after dilution but was not significant. CONCLUSION: Our method of removing DMSO during the thawing process yields a higher cell survival rate and less DMSO toxicities compared with the conventional method of direct injection with DMSO after thawing.

Expressions of c-fos and c-myc genes during 3'-methyl-4-dimethylaminoazobenzene (3'-MeDAB)-induced rat hepatocarcinoma

We investigated the expression of the growth-related nuclear proto-oncogenes, c-fos and c-myc, in early preneoplastic regions and tumor nodules of 3'-MeDAB induced rat hepatocarcinoma. To amplify the levels of these transcripts, we gave cycloheximide (100 mg/kg B.W. i.p.) to each group of rats. The elevated levels of the 2.2 kb c-fos and 2.4 kb c-myc transcripts appeared as early as the 2nd week after feeding on the 3'-MeDAB diet and lasted through the 4th; 6th weeks and tumor. Southern blot analysis indicated that gross amplification or rearrangements were not observed in DNA of the preneoplastic livers and hepatoma nodules. We also measured the rate of the incorporation of [3H] thymidine into hepatic DNA in order to monitor the rate of cell proliferation occurring at the early preneoplastic periods. We have found that the rate of [3H] thymidine incorporation corresponds to the elevated levels of c-fos and c-myc transcripts in the precancerous stages. This finding suggests that the elevated expressions of c-fos and c-myc may result from the continuous cell proliferative stimuli generated in the carcinogen altered cells, which is essential to the initiation and promotion of chemical hepatocarcinogenesis.

Angiographic Analysis of Ruptured Anterior Communicating artery Aneurysms

The authors reviewed angiographic findings of 96 cases of ruptured anterior communicating artery aneurysms that were operated at the Department of Neurosurgery of Hanyang University Hospital from Jan. 1986 to June 1990. The results were summarized as follows ; 1) Asymmetricity of caliber of A1 was observed in 71.9%(69/96) and which was 3 times more frequent than that of reported incidence of control group(2~23%). 2) The incidence of predominance of left A1 was 54.2%(53/96) and that of right A1 was 17.7%(17/96). Among them, predominant A1 was operated as afferent artery of aneurysm in 94.2%(65/69). 3) Of 69 cases in which a unilateral A1 showed predominance, the direction of aneurysm was opposite to the side of predominant A1 in 66.7%(46/69) and ipsilateral to the predominant A1 in only 2.9%(2/69). 4) Above findings suggest that it is desirable to approach the anterior communicating artery aneurysm via the side of predominant A1 when the direction of aneurysm is uncertain on cerebral angiogram.

Surgical Experience on directly treated 18 Intracranial Aneurysm

Eighteen intracranial aneurysm were operated upon with direct approach under the surgical microscope. The aneurysms arising in the anterior communicating artery(8) were most common, followed by internal carotid artery(6), middle cerebral artery(2), posterior communicating artery(1), anterior cerebral artery(1) in order. We preferred pterional approach. And the timing of operation was scheduled as soon as possible if other conditions permitted. Mannitol administration, hypotensive anesthesia, Sugita clip, biobond or aneurysmostat were utilizes during the operation. The aneurysmal neck was clipped with Sugita clip mostly, but wrapping was also carried out in certain condition. The elasticity of the Sugita clip is seemed to be better than others. In even Sugita clip, however, overopening at the time of clipping made it weak and resulted in incomplete occlusion of the aneurysm neck. To obtain successful aneurysmal surgical result, following surgical care should be taken. 1) lesser brain retraction, 2) complete removal of subarachnoid clots, 3) Meticulous hemostasis of micro-oozing from bridging vein and internal dural surface. Uneventful two surgical cases were found to be fatal due to hidden post-operative oozing clots from the internal surface of the dura.