Development and characterization of a humanized mouse model of osteoarthritis

Abstract

Purpose: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Osteoarthritis (OA) is mainly characterized by cartilage breakdown and synovial inflammation. While OA is one of the most common rheumatic disorders, no efficient disease-modifying drugs still exist today. Mesenchymal stem cells (MSC) have generated significant medical considerations since they exhibit tissue-regenerative properties through the secretion of bioactive factors. Phase I-II clinical trials have shown that intra-articular (IA) injection of MSC is well-tolerated and exhibited promising clinical results. However, MSC display a poor survival and engraftment rate in vivo. Using encapsulating systems could protect MSC and maintain their bioactivity after intra-articular injection. Considering the pivotal role of inflammation and immune system in OA and with regards to the differences between MSC-mediated immunomodulation in human and mice (12), the development of sophisticated animal models that could more closely mimic the human immune system, could be instrumental. Amongst these models, immunologically humanized mice may represent a preclinical bridge between in vitro studies and the in vivo evaluation of human stem cells in OA. Our objective is therefore to develop an experimental model of osteoarthritis by destabilizing the medial meniscus (DMM) in humanized mice.> Methods: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <4 weeks-old NOD/LtSz-scid IL2Rγnull (NSG) mice were humanized as follow. After irradiation (1,5Gy), 5.106 CD34+ human hematopoietic stem cells were injected intravenously. 120 days after the humanization procedure, the level of human immune cells CD45+ (leukocytes), CD3+ (T lymphocytes), CD19+ (B lymphocytes), CD56+ (NK cells) and CD14+ (macrophages) were evaluated by flow cytometry. Humanized NSG mice then had undergone a DMM or a SHAM surgery. 6 and 12 weeks after OA induction the development of osteoarthritis was assessed histologically by safranin-o staining, OA score and histomorphometric analysis of the knees. In addition, to evidence the presence of human immune cells in the joint, Immunohistochemicals stainings for human CD45+, CD3+, CD19+ et CD56+ and CD14+ cells have also been performed.> Results: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Our results showed that 120 days after the humanization procedure, NSG mice exhibit engraftment level (hCD45+) from 10% to 75 % and the presence of human T and B lymphocytes as well as macrophages and NK cells in blood and spleen. Our histological results show that the DMM procedure induce a significant increase in OA score at 12 weeks compared to sham mice. Our histomorphometric results also evidence a significant increase in bone volume of the anterior medial meniscus. However, no significant differences have been observed between DMM and Sham mice on the OA score at 6 weeks and on the subchondral bone morphometry at 6 and 12 weeks’ post-surgery. Immunohistochemical analyses have however revealed the presence of human B lymphocytes and macrophages in the subchondral bone marrow but not in the synovial membrane at 6 and 12 weeks post-surgery.> Conclusions: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Finally, using 2 methods of analysis (histology and histomorphometry), we have demonstrated that humanized NSG mice could develop post-traumatic OA after a DMM surgery. The presence of human immune cell in the spleen and subchondral bone marrow confirms the success of the hematopoietic transplant. However, we failed to detect any immune cells in the synovial membrane or articular cartilage although human immune cells were observed near the joint in the subchondral bone marrow. The search for these immune cells in the synovial membrane would be more appropriate during the inflammatory burst, meaning within the first 2 weeks following the DMM surgery. Our results therefore evidence the feasibility to induce OA by DMM in humanized mice. This model could constitute a useful tool to study the therapeutic efficacy of encapsulated human mesenchymal stem cells.>