A comparative experimental study of the efficacy and safety of Quality and Quantity media-cultured mononuclear cell transplantation in ischemic hindlimb mouse model

Abstract

Abstract Background: Up to 15-20% of revascularizations for chronic limb-threatening ischemia (CLTI) fail. Peripheral blood mononuclear cell (PBMNC) transplantation is an alternative treatment for CLTI patients, but a meta-analysis study found no significant clinical benefit of this treatment. Impaired function and/or an insufficient number of PBMNCs in CLTI patients might be the cause of unsatisfactory outcomes. Quality and Quantity media-cultured mononuclear cells (QQMNCs) were reported to enhance the number and function of PBMNCs. Accordingly, the aim of this study was to investigate the efficacy and safety of QQMNC transplantation in an ischemic hindlimb mouse model. Methods: PBMNCs from CLTI patients were cultured in Quality and Quantity (QQ) culture media or standard culture media. In vitro studies, including phenotypic analysis of progenitor cells (CD34+CD133+), M2 macrophages (CD206+), and inactivated T regulatory cells (CD4+CD25+CD127+); colony forming assay; and tube formation assay of QQMNCs and PBMNCs, were conducted. Intramuscular transplantation of QQMNCs or PBMNCs was performed in the ischemic hindlimb mouse model. The clinical appearance of ischemic limbs was observed, and blood flow in ischemic limbs was measured using a laser Doppler perfusion imager. Transplantation outcomes were compared between the QQMNC and PBMNC groups. Results: Twenty CLTI patients (mean age: 65.9±8.56 years) were included. Diabetes mellitus, hypertension, current smoker status, and chronic kidney disease was found in 12, 17, 11, and 6 patients, respectively. The mean percentages of CD34+ cells, CD133+ cells, CD34+CD133+ progenitor cells, CD206+ cells, colony-forming cells, and tube formation were significantly higher in the QQMNC group than in the PBMNC group. However, the mean percentage of CD4+CD25+CD127+ cells was significantly lower in QQMNCs than in PBMNCs. The colony-forming unit count and Dil-acetylated low-density lipoprotein uptake were both significantly greater in QQMNCs than in PBMNCs. The clinical appearance of post-QQMNC-injected limbs was less severe than the appearance of post-PBMNC-injected limbs. Limb perfusion was significantly better in the QQMNC group than in the PBMNC group. Conclusions: Proangiogenic and anti-inflammatory phenotypes of MNCs cultured in QQ culture media were successfully reproduced in vitro. Moreover, intramuscular QQMNC transplantation was found to be safe, and resulted in better reperfusion of ischemic mouse hindlimbs compared to PBMNCs.