Quality of thawed hematopoietic progenitor cell grafts

• Main Authors: Metka Krašna, Elvira Maličev, Mojca Jež, Andreja Nunar Perko, Marko Cukjati
• Format: Article
• Language: English
• Published: Slovenian Medical Association 2015-06-01
• Series: Zdravniški Vestnik
• Subjects: Viability; CD34+; transplantation; hematopoietic progenitor cells; cryopreservation
• Online Access: http://vestnik.szd.si/index.php/ZdravVest/article/view/1269

<p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span class="highlight2"><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">Background</span></span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">:</span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">The appropriate dose of compatible and viable hematopoietic progenitor cells (HPC) is crucial for successful engraftment of transplanted cells within patients. In </span><span class="hps"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">the study we</span></span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> <span class="hps">therefore</span> <span class="hps">checked</span> <span class="hps">the viability of</span> HPC in thawed HPC grafts which were <span class="hps">transplanted into</span> <span class="hps">haematology</span> <span class="hps">patients</span> <span class="hps">in one year.</span> </span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">Methods:</span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span class="hps"><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">We acquired HPC</span></span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> <span class="hps">by apheresis</span> <span class="hps">and</span> <span class="hps">froze them in</span> <span class="hps">a programmable</span> <span class="hps">freezer</span> <span class="hps">in</span> <span class="hps">autologous</span> <span class="hps">plasma</span> <span class="hps">with</span> 10 % <span class="hps">DMSO</span> <span class="hps">and stored them in</span> <span class="hps">liquid nitrogen</span> <span class="hps">vapour</span>. </span><span class="hps"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">We thawed the bags with HPC</span></span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> <span class="hps">in a water bath</span> <span class="hps">at the patient's bedside</span> and <span class="hps">in the eluate residues from the bags after the addition of saline solution we determined the viability</span> <span class="hps">(</span>7<span class="atn">-</span>AAD) of HPC <span class="hps">(</span>CD34 <span class="hps">+</span>) and <span class="hps">lymphocytes by flow cytometry.</span></span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">Results:</span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">After the transplantation of HPC, the viability of HPC was 79.9 ± 12.4 % and of leukocytes 62.2 ± 12.2 % (paired t-test, P &lt; 0.001) in cell suspension residues (n = 88). </span><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">The average viability of HPC in samples which were frozen immediately after collection (n = 58) was 79.9 ± 9.2 % and in samples frozen the next day after 18 hours in the fridge, the viability was 80.5 ± 11.3 % (n = 67).</span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> Storing samples overnight did not significantly affect the viability of cells in comparison with samples which were frozen immediately (t-test, P &gt; 0.5). </span><span class="hps"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">Rinsing</span></span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> of HPC bags and <span class="hps">storing cells</span> <span class="hps">after thawing</span> <span class="hps">in saline solution</span> <span class="hps">significantly (</span>n <span class="hps">= 7</span>; paired<span class="hps"> t</span><span class="atn">-</span>test, <span class="hps">P</span> <span class="hps">&lt;</span>0.01) <span class="hps">increased the mortality of HPC</span>, since the viability <span class="hps">immediately</span> <span class="hps">after thawing</span> was<span class="hps"> 84.1</span> <span class="hps">± 4.5 </span>% and in samples <span class="hps">diluted</span> <span class="hps">with saline solution</span> <span class="hps">60.1</span> <span class="hps">±</span> <span class="hps">15.7 %</span>.</span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">Conclusions:</span></p><p class="MsoNormal" style="text-align: justify; line-height: 116%;"><span style="font-size: 12.0pt; line-height: 116%; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-GB;" lang="EN-GB">In our laboratory, cca 20 % of HPC are lost during freezing and thawing of HPC grafts. </span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB">Freezing HPC grafts 18 hours after cell collection does not affect the viability of thawed cells in comparison with grafts which are frozen immediately.</span><span style="font-size: 12.0pt; line-height: 116%; mso-ansi-language: EN-GB;" lang="EN-GB"> Saline solution is not suitable for rinsing bags and storing HPC residuals after thawing HPC products.</span></p><br /><br />