Study for the improvement of umbilical cord blood sampling using a new trial apparatus
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1 bs_bs_banner doi: /jog J. Obstet. Gynaecol. Res. Vol. 40, No. 2: , February 2014 Study for the improvement of umbilical cord blood sampling using a new trial apparatus Naoki Masaoka 1, Masako Morooka 1, Yoshiyuki Nakajima 1, Hajime Ogata 2, Hideki Kodo 3 and Shunichi Kato 4 1 Department of Obstetrics and Gynecology, Tokyo Women s Medical University Yachiyo Medical Center, Chiba, 2 Ogata Children s Clinic, Osaka, 3 Tokyo Cord Blood Bank, Tokyo, and 4 Department of Cell Transplantation, Tokai University School of Medicine, Isehara, Japan Abstract Aim: The aim of this study was to evaluate the usefulness of the trial umbilical cord for unrelated cord blood transplantation. Material and Methods: Data were obtained from 100 vaginal deliveries. In 50 cases, umbilical cord blood (UCB) was taken with the traditional Kawasumi type UCB sampling bag. In another 50 cases, UCB were taken with trial UCB sampling bag offered by NIPRO Co. We compared the sampling volume between the two groups. Furthermore, 10 cases in each group were matched by sampling volume; we examined the quality of UCB on the number and concentration of nucleated cells, mononuclear cells, CD34+ cells and colony-forming unit granulocyte macrophage and the numbers tested positive for bacteria. Results: Whereas there were no significant differences in gestational weeks at sampling, the ratio of primipara women to multipara women, maternal age, and neonatal weight between the two groups, the sampling UCB volumes with the trial sampling bag were significantly higher than those with traditional sampling bags (P < 0.05). In addition, this phenomenon was more significant in the latter part of the study period (P < 0.05). On the other hand, there were no significant differences in the quality of UCB between the two groups. Conclusion: Once clinicians have become accustomed to the trial UBC sampling bag, this method might be a useful method for collecting UCB for unrelated cord blood transplantation. Key words: cord blood sampling, placenta, stem cell, unrelated cord blood transplantation. Introduction Since 1989, 1 advances in umbilical cord blood (UCB) technology have allowed UCB transplants to become an increasingly desirable alternative to traditional bone marrow transplants in patients requiring a stem cell transplant, such as those suffering from leukemia, lymphoma and other life-threatening blood disorders. 2 5 UCB is one of three sources for the blood-forming cells used in transplants. The other two sources are bone marrow and peripheral (circulating) blood. UCB is collected from an umbilical cord and placenta after delivery, so that it may be tested, frozen and subsequently stored in a cord blood bank for future use. There are many advantages to using UCB compared with other sources of stem cells, such as bone marrow and peripheral blood: 6 (i) there is no risk involved during UCB collection to either mother or baby; (ii) UCB is much easier to collect and harvest without the risks of general anesthesia, which is required to harvest bone marrow; (iii) UCB is readily available when needed if it is properly collected and stored at birth; Received: December Accepted: May Reprint request to: Dr Naoki Masaoka, Department of Obstetrics and Gynecology, Tokyo Women s Medical University Yachiyo Medical Center, Owada-Shinden, Yachiyo, Chiba , Japan. masaoka@tymc.twmu.ac.jp 2013 The Authors 405
2 N. Masaoka et al. Figure 1 Umbilical cord. Right: trial umbilical cord. Left: traditional Kawasumi type UCB sampling bag. Figure 2 Fixing the apparatus to the appropriate position. Following umbilical venipuncture, the apparatus was clipped to affix the umbilical cord. After extracting the stylet, the tube to prevent external air from entering the system was freed, prior to venous sampling. Methods (iv) UCB is more proliferate in nature and often more compatible when used in transplants; (v) UCB has lower procurement costs; and (vi) UCB has demonstrated broader potential clinical applications for improving neural repair and bone and tissue growth. In Japan, more than 8000 unrelated umbilical cord blood transplantations (UCBT) have been performed through 11 UCB banks in the Japan Cord Blood Bank Network (JCBBN). As a component of quality management and promotion, the JCBBN established a common registry for studying the results of UCBT. 7 This investigation examined the usefulness of a trial UBC sampling bag designed for collecting UCB effectively and safely by NIPRO Co. (Osaka, Japan). This new apparatus for UBC collection has some features as listed below (Fig. 1,2): 1. This apparatus for umbilical blood sampling has an inner stylet, which is withdrawn after the umbilical vessel is entered. The outer tube is soft and pliable to prevent damage to the vascular wall. 2. The outer tube has multiple perforations to enable more efficient sampling. 3. Once the sampling tube has been affixed properly with the umbilical cord with a built-in clip, both hands become free for required additional manipulation. 4. The tube connecting the needle and collecting bag has been designed to prevent external air from entering the system. Prior to the study, approval for the experimental design was obtained from the Experimental Ethics Committee of Tokyo Women s Medical University. Data were obtained from 100 normal singleton vaginal deliveries at Tokyo Women s Medical University Yachiyo Medical Center from 1 February to 31 December In 50 cases, UCB was taken with the traditional Kawasumi type UCB sampling bag made by NIPRO Co. In another 50 cases, UCB were taken with the trial UCB sampling bag developed by NIPRO Co. Informed consent was obtained from all participating women. It was explained as follows: UCB collected by the trial UCB sampling bag would not be stored for supplying for clinical application, even if it had sufficient volume and quality. It would only be used for the investigation of the improvement of UCB sampling. UCB was collected by three doctors with almost equal ability and experience in UCB sampling. On odd days we collected UCB by the traditional Kawasumi type bag and on even-numbered days we collected UCB with the trial UCB sampling bag. We were anxious about deterioration in the quality of umbilical cord blood (number of stem cells and contamination) by changing sampling method. To examine the UCB quality, 10 cases of similar volume from each group were compared by the number and concentration of nucleated cells, mononuclear cells, CD34 + cells and colony-forming unit granulocyte macrophage (CFU-GM) and assessed for bacterial contamination. We felt that 10 samples of similar volume from both groups would provide sufficient data for comparison The Authors
3 Umbilical cord blood sampling All measurements and analyses were performed by the Tokyo Cord Blood Bank as a third participating party. All values are expressed as the mean standard deviation. Statistical comparisons between groups were made using the Student s t-test. A value of P < 0.05 was considered significant. Results Backgrounds of cases (Table 1) There were no significant differences about gestational weeks at sampling, the ratio of primipara women to multipara women, maternal age and neonatal weight between the two groups. UCB sampling volume (Table 2) Table 2 shows that the UCB sampling volume with trial sampling bag was significantly higher than that with the traditional sampling bag ( g vs g) (P < 0.05). In addition, Table 2 shows the changes of UCB sampling volume during the study protocol divided into two periods as the former term and the latter term. Increasing UCB sampling with trial sampling bag was more significant during the latter term of the study period (P < 0.05). Comparison of UCB quality between two groups (Table 3) There were no significant differences in number and concentration of nucleated cells, mononuclear cells, CD34 + cells and CFU-GM and occurrence of bacterial contamination of UCB between the two groups. Discussion In this study we demonstrated that the trial sampling bag was able to collect UCB more effectively than the traditional sampling bag. Initially, umbilical cord blood transplant (UCBT) had indication for child patients, 8 10 however the number of UCBT for adult patients has increased dramatically In cases of UCBT for adult patients, we need a sufficient number of nucleated cells. Namely we require a sufficient volume of UCB. One problem associated with UCB is that its collection is a one-time possibility and the amount of umbilical cord blood that can be collected is limited using conventional current ways of blood collection, which include syringe-assisted and gravity-assisted methods. 14 Elchalal et al. reported that the syringe-assisted sodium chloride solution flush collection method with a blood bag was found to be the most effective method for human UCB collection. 15 Table 1 Backgrounds of cases Traditional Kawasumi type umbilical cord n = 50 Trial umbilical cord n = 50 Gestational ages at sampling Primipara/multipara 28/22 31/19 Maternal age Neonatal weight (g) Table 2 Umbilical cord blood sampling volume 2013 The Authors 407
4 N. Masaoka et al. Table 3 The comparison of umbilical cord blood quality between two groups Traditional Kawasumi type umbilical cord n = 10 Trial umbilical cord n = 10 Umbilical cord blood sampling volume (g) Concentration of nucleated cells (10 2 /ml) Number of nucleated cells ( 10 8 ) Concentration of mononuclear cells (10 2 /ml) Number of mononuclear cells ( 10 8 ) Concentration of CD34 + cells (/ml) Number of CD34 + cells ( 10 6 ) Concentration of CFU-GM (/ml) Number of CFU-GM ( 10 5 ) Number of bacterial contamination 0 0 CFU-GM, colony-forming unit granulocyte macrophage. The predominant collection procedure currently practiced involves a relatively simple venipuncture, followed by gravity drainage into a standard sterile anti-coagulant-filled blood bag, using a closed system, similar to the one utilized in whole blood collection. In ordinary circumstances, transplant requires nucleated cells/kg. Thus we need nucleated cells for a 50-kg adult patient. A previous study demonstrated that the necessary volume of UCB for obtaining nucleated cells was more than 70 g. The average sampling UCB volume with the trial sampling bag was about 100 g. In a strict sense, UCB sampling volume is affected by the umbilical cord s strength, size and placental weight, etc. However, we demonstrated that the trial sampling bag could collect sufficient volume for a UCB transplant to an adult patient. Furthermore, the quality of UCB collected by the trial sampling bag in regards to number and concentration of nucleated cells, mononuclear cells, CD34 + cells, CFU-GM and tested bacterial contamination were not significantly different from the UBC collected by the traditional sampling bag. The volumes obtained and quality were sufficient to obtain stem cells for adult patients requiring a stem cell transplant. As far as traditional sampling bag was concerned, we were used to this method; on the other hand we were inexperienced with the new sampling bag at the beginning of this study. As a matter of fact, there were some troublesome problems in sampling UBC with the new sampling bag, such as venipuncture connecting the apparatus effectively removal of stylet freeing the connecting tube sampling. However, the more we became accustomed to the new sampling bag, the more UCB volume we could obtain. Therefore, we now believe that after becoming accustomed to using the trial UBC sampling bag, this new sampling bag might be a useful apparatus for collecting UCB for unrelated cord blood transplantation. The new sampling bag is manufactured for trial usage, thus unit price is expensive. But this bag is an improved model of the traditional bag that had obtained approval from the Ministry of Health, Labor and Welfare (MHLW) in Japan, thus we do not require the extensive clinical trials and are able to supply the new sampling bags to the market with only an announcement of modification. We experienced an increased volume of UCB obtained with the improved collecting bag, sufficient to allow the transplant to an adult from one normal childbirth. Thus, making it more potentially utile and, consequently, with increased demand, would force the price per unit downward. We plan to introduce this product to acquire sufficient popularity for its use in clinical medicine and we are in the process of appealing our finding to the MHLW in order to allow its utilization. Another future development of the UCB system may lead to an integrated and comprehensive computercontrolled system for the whole process from placenta harvesting to cell separation and storage. Acknowledgment This work was supported in part by a Health and Labor Science Research Grant for Allergic Disease and Immunology (H20-015) from the Japanese Ministry of Health, Labor and Welfare of Japan The Authors
5 Umbilical cord blood sampling Disclosure The authors have no potential conflicts of interest to closure. References 1. Gluckman E, Broxmyer HA, Auerbach AD et al. Hematopoietic reconstitution in a patient with Fanconi s anemia by means of umbilical cord blood from an HLA-identical sibling. N Engl J Med 1989; 321: Broxmeyer HE, Douglas GW, Hangoc G et al. Human umbilical cord blood as a potential source of transplantable hematopoietic stem/progenitor cells. Proc Natl Acad Sci USA 1989; 86: Rubinstein P, Carrier C, Scaradavou A et al. Outcomes among 562 recipient of placental-blood transplants from unrelated donors. N Engl J Med 1998; 339: Gluckman E, Rocha V, Arcese W et al. Factors associated with outcomes of unrelated cord blood transplant: Guidelines for donor choice. Exp Hematol 2004; 32: Rocha V, Labopin M, Sanz G et al. Transplants of umbilicalcord blood or bone marrow from unrelated donors in adults with acute leukemia. N Engl J Med 2004; 351: Tan KK, Tang KZ, Huang S. Devices for umbilical cord blood collection. Recent Pat Eng 2007; 1: Nagamura-Inoue T, Kai S, Azuma H et al. Unrelated cord blood transplantation in CML: Japan Cord Blood Bank Network analysis. Bone Marrow Transplant 2008; 42: Gluckman E, Rocha V, Boyer-Chmmard A et al. Outcome of cord-blood transplantation from related and unrelated donors. Eurocord Transplant Group and the European Blood and Marrow Transplantation Group. N Engl J Med 1997; 337: Rocha V, Wagner JE, Sobocinski KA et al. Graft-versus-host disease in children who have received a cord-blood or bone marrow transplant from an HLA-identical sibling. Eurocord and International Bone Marrow Transplant Registry Working Committee on Alternative Donor and Stem Cell Sources. N Engl J Med 2000; 342: Tse W, Laughlin MJ. Umbilical cord blood transplantation: A new alternative option. Hematology 2005; 2005: Barker JN, Davies SM, DeFor T et al. Survival after transplantation of unrelated donor umbilical cord blood is comparable to that of human leukocyte antigen-matched unrelated donor bone marrow; results of a matched pair analysis. Blood 2001; 97: Laughlin MJ, Eapen M, Rubinstein P et al. Outcomes after transplantation of cord blood or bone marrow from unrelated donors in adults with leukemia. N Engl J Mesd 2004; 351: Rocha V, Labopin M, Sanz G et al.; Acute Leukemia Working Party of European Blood and Marrow Transplant Group; Eurocord-Netcord Registry. Transplants of umbilical-cord blood or bone marrow from unrelated donors in adults with acute leukemia. N Engl J Med 2004; 351: Bertolini F, Lazzari L, Lauri E. Comparative study of different procedures for the collection and banking of umbilical cord blood. J Hematother Stem Cell Res 1995; 4: Elchalal U, Fasouliotis SJ, Shtockheim D et al. Postpartum umbilical cord blood collection for transplantation: A comparison of three methods. Am J Obstet Gynecol 2000; 182: The Authors 409
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