We have invented mobile HEPA filter unit of hemopoietic stem cell transplantation clean room (class 100), which is a capsule type to protect the patient from being infected by harmful Aspergillus spore. This equipment is based on the basic structure of US Federal Standard 209D, biological clean room standard of NASA NHB 5340-2 and measurement standard of Japanese JACA No. 24-1989. Performance of HEPA filter was evaluated by dioctyl phthalate (DOP) test method, resulting in 99.97% efficiency for 0.3 ㎛ particles. HEPA filter unit used only 98 W electricity with 24 V storage cell for the patient's treatment for over three hours. When the wind speed was 0.2 to 0.3 m/sec as in bone marrow transplantation clean room, the air flow was constant laminar vertical type. When the amount of dust was measured in the hospital, the equipment maintained the grade of clean class 100 and microbe was not detected. The noise level was less than 52 dB(A). The amounts of microbe inside and outside the building were almost same. The amount was even more in cancer sick room, serious case room and X-rays room than in the general rooms. Also, elevators and stair room were found to transport dust to other areas. Even though the patient with low immunity can be protected from Aspergillus spore by a HEPA filter unit, the system preventing microbe system such as BMT clean room should be placed in rooms such as PET, CT, MRI, and radiation room so that the patients can be protected from having infected by the extremely harmful Aspergillus spore.

Bisphosphonates have been widely used to treat metabolic bone diseases, although the . mechanism of bisphosphonate action on bone has not been fully understood. This study aimed to examine the direct action of pamidronate on cell proliferation and differentiation of cultured human mesenchymal stem cells(hMSC). Four experimental groups and two control groups were designed; Experimental groups included both osteogenic supplement(OS) and pamidronate-treated group, pamidronate-treated group after 1 week OS treatment, only pamidronate-treated group, OS-treated group after 1week pamidronate treatrnent. Control gr。니ps included DMEMtreated group and OS-treated group. Human MSCs were isolate from bone maπow ， and cultured for 7, 14, 21 days. For the detection of osteoblastic differentiation, AI.Pase activity was measured and the expression of type 1 collagen and osteocalcin were evaluated. Von Kossa’s silver stain was performed for the examination of calcification. As results, the proliferation rate of 바1SC was maintained to be more than 90% by 1uglml of pamidronate. AI.Pase activity showed the highest value at the concentration of 100nglml of pamidronate. In pamidronate-treated group, ALPase activity reached a peak at the third week and the expression of type 1 collagen mRNA and protein was enhanced compared to other experimental and control groups, whereas osteocalcin expression was found only in OStreated group. Calcification was decreased by a dose dependent manner followed by pamidronate treatment. This study su잃，est that pamidronate treatrnent may be able to enhance the osteoblastic differentiation of hMSC at the early stage. On the other hand, calcification appeared to be inhibited by pamidronate treatrnent.

In mammals, male and female germline stem cells are derived from primodial germ cells. Despite many efforts to identify stem cells from gonads, there has been little successe to identify germline stem cells yet. In this study, we isolate and characterized porcine germline stem cells using only stem cell markers that are prevalently expressed in various tissues. Gonadal cells derived from both male and female formed colonies and showed AP activities and different lectin binding properties. Pluripotency of germline stem cells was also identified by positive signals against putative stem cells markers such as SSEA-1 and SSEA-3. In addition, nestin was also found in primary gonad cells that have a similar morphology to the AP-positive cells. The nestin expression suggests that the germline stem cells may have similar expression of the prevalent stem cell markers found in other tissues. The demonstration of nestin expression together with pluripotent cell markers calls further investigation of the possible differentiation of nestin-positive cells into neurons.

Human embryonic stem (hES) cell lines have been derived from human blastocysts and are expected to have far-reaching applications in regenerative medicine. The objective of this study is to improve freezing method with less cryo-injuries and best survival rates in hES cells by comparing various vitrification conditions. For the vitrifications, ES cells are exposed to the 4 different cryoprotectants, ethylene glycol (EG), 1,2-propanediol (PROH), EG with dime-thylsulfoxide (DMSO) and EG with PROH. We compared to types of vehicles, such as open pulled straw (OPS) or electron microscopic cooper grids (EM grids). Thawed hES cells were dipped into sequentially holding media with 0.2 M sucrose for 1 min, 0.1 M sucrose for 5 min and holding media for 5 min twice and plated onto a fresh feeder layer. Survival rates of vitrified hES cells were assessed by counting of undifferentiated colonies. It shows high survival rates of hES cells frozen with EG and DMSO (60.8％), or EG and PROH(65.8％) on EM grids better than those of OPS, compared to those frozen with EG alone (2.4％) or PROH alone (0％) alone. The hES cells vitrified with EM grid showed relatively constant colony forming efficiency and survival rates, compared to those of unverified hES cells. The vitrified hES cells retained the normal morphology, alkaline phosphates activity, and the expression of SSEA-3 and 4. Through RT-PCR analysis showed Oct-4 gene expression was down-regulated and embryonic germ layer markers were up-regulated in the vitrified hES cells during spontaneous differentiation. These results show that vitrification method by using EM grid supplemented with EG and PROH in hES cells may be most efficient at present to minimize cyto-toxicity and cellular damage derived by ice crystal formation and furthermore may be employed for clinical application.