Administration of Human Non-Diabetic Mesenchymal Stromal Cells to a Murine Model of Diabetic Fracture Repair: A Pilot Study

Abstract

Individuals living with type 1 diabetes mellitus may experience an increased risk of long bone fracture. These fractures are often slow to heal, resulting in delayed reunion or non-union. It is reasonable to theorize that the underlying cause of these diabetes-associated osteopathies is faulty repair dynamics as a result of compromised bone marrow progenitor cell function. Here it was hypothesized that the administration of non-diabetic, human adult bone marrow-derived mesenchymal stromal cells (MSCs) would enhance diabetic fracture healing. Human MSCs were locally introduced to femur fractures in streptozotocin-induced diabetic mice, and the quality of de novo bone was assessed eight weeks later. Biodistribution analysis demonstrated that the cells remained in situ for three days following administration. Bone bridging was evident in all animals. However, a large reparative callus was retained, indicating non-union. µCT analysis elucidated comparable callus dimensions, bone mineral density, bone volume/total volume, and volume of mature bone in all groups that received cells as compared to the saline-treated controls. Four-point bending evaluation of flexural strength, flexural modulus, and total energy to re-fracture did not indicate a statistically significant change as a result of cellular administration. An ex vivo lymphocytic proliferation recall assay indicated that the xenogeneic administration of human cells did not result in an immune response by the murine recipient. Due to this dataset, the administration of non-diabetic bone marrow-derived MSCs did not support fracture healing in this pilot study.

Regenerative and Anti-Inflammatory Potential of Regularly Fed, Starved Cells and Extracellular Vesicles In Vivo

“…To validate our main hypothesis in vivo, we adopted a well-established preclinical model of delayed fracture healing [ 21 , 32 ] using the experimental plan depicted in Figure 4 A. Diabetic induction with STZ was successful, resulting in a mean of plasma glucose level of 23.9 ± 5.3 mM in comparison to control animals (injected with HBSS) having 11.2 ± 0.57 mM ( p ≤ 0.05) ( Figure 4 B). Diabetic mice also showed a reduced but steady weight throughout the study, 26.34 ± 1.17 g for STZ-treated animals and 32.3 ± 1.92 g ( p ≤ 0.05) for control animals ( Figure 4 C).…”

“…All animals with no signs of pain and a glucose level ≥ 13 mM for three consecutive weeks were classified as hyperglycemic and enrolled in the study. After three weeks (day 0) animals were anesthetized with isoflurane, the joint surface was exposed, and an intramedullary stabilizing pin, a 27 G syringe needle, was inserted into the marrow cavity and confirmed using an X-ray machine, as previously described [ 21 ]. Mice femurs were positioned on the fracture device [ 35 ], a three-point bending guillotine, under the center of the sliding weight before being dropped onto the leg, causing the fracture [ 21 ].…”

“…After three weeks (day 0) animals were anesthetized with isoflurane, the joint surface was exposed, and an intramedullary stabilizing pin, a 27 G syringe needle, was inserted into the marrow cavity and confirmed using an X-ray machine, as previously described [ 21 ]. Mice femurs were positioned on the fracture device [ 35 ], a three-point bending guillotine, under the center of the sliding weight before being dropped onto the leg, causing the fracture [ 21 ]. An X-ray was taken immediately after the fracture, to ensure that the location of the intramedullary pin had not altered and that the transverse fracture had occurred.…”

“…An X-ray was taken immediately after the fracture, to ensure that the location of the intramedullary pin had not altered and that the transverse fracture had occurred. Following anesthesia, animals were given 0.05 mg/kg buprenorphine and were separately housed [ 21 ].…”

“…Diabetes is a disease that triggers chronic hyper-inflammatory conditions [ 1 , 21 , 22 , 23 ], impairing the activity and presence of neutrophils [ 1 ], monocytes and dendritic cells [ 24 ] that result in pro-inflammatory and chemotactic mediators’, such as IL-6, MCP-1 (CCL2), IL-1, TNF-α, IL-4 and IFN-γ, persistent secretion [ 25 , 26 , 27 , 28 , 29 ]. All these conditions contribute to induce anti-osteogenic activity, decreased soft callus formation and enhanced osteoclastic and bone turnover activity [ 30 ], thereby delaying fracture healing [ 1 , 22 , 31 ].…”