Retrospective analysis of local injection site adverse reactions associated with 230 allogenic administrations of bone marrow-derived mesenchymal stem cells in 164 horses.

BACKGROUND: Bone marrow-derived mesenchymal stem cells (BM-MSCs) are frequently used in the treatment of musculoskeletal injuries. Fully characterised cells that are readily available for use is optimum. Allogenic BM-MSCs can satisfy the need for rapid treatment, however, their safety has been questioned. OBJECTIVES: Objectives were to characterise BM-MSCs from an adult donor horse, in vitro, and to identify and describe adverse reactions that occurred following their injection into other horses. We hypothesised that BM-MSCs capable of proliferation, differentiation and lacking MHC II from one donor could be implanted into another individual without significant adverse reactions and the frequency of adverse reactions in clinical cases would be similar to that previously reported for autologous BM-MSCs. STUDY DESIGN: Retrospective clinical study. METHODS: BM-MSCs were proliferated and characterised from one donor and cryopreserved for clinical use. Medical records for horses injected with allogenic BM-MSCs from this donor at a single hospital were used. After routine lameness exam, lesions were identified using diagnostic ultrasound or MRI. Post injection reaction was defined as increased pain, swelling, or heat at or near injection site, or increased lameness. Treatments required for each reaction were noted. RESULTS: BM-MSCs proliferated and underwent differentiation. Cells were found to be negative for MHC-II (<2%) and were viable after cryopreservation and shipping. Ten of 230 (4.35%) injections were noted to be associated with an adverse reaction. Adverse reactions occurred in synovial structures (n = 3) and in soft tissues (n = 7). MAIN LIMITATIONS: This investigation could underestimate the number and severity of reactions. Mild reactions, such as synovitis, may have been missed. Also, anti-inflammatory drugs could overshadow mild reactions, making them less likely to be detected. CONCLUSIONS: Fully characterised allogenic BM-MSCs originating from a single donor horse can be administered to horses with soft tissue injuries with a low rate of adverse reaction. The Summary is available in Portuguese - see Supporting Information.

In vitro analysis of equine, bone marrow-derived mesenchymal stem cells demonstrates differences within age- and gender-matched horses.

REASONS FOR PERFORMING THE STUDY: Stem cell therapies are used routinely in equine practice. Most published reports characterise stem cells derived from younger horses; however, middle-aged horses are often in athletic performance, and experience degenerative medical conditions. Thus, mesenchymal stem cells (MSCs) from this group should be investigated. OBJECTIVE: To describe differences in in vitro adherence, proliferation and potential for differentiation of equine bone marrow-derived MSCs (equine BMMSCs) harvested from middle-aged (10-13 years old) female donors. STUDY DESIGN: Descriptive study of stem cell characteristics. METHODS: Equine BMMSCs from 6 horses were cultured in vitro and evaluated for viability, proliferation, osteogenesis, chondrogenesis, adipogenesis, cluster-of-differentiation markers and gene expression. RESULTS: Equine BMMSCs from all 6 donors demonstrated fibroblastic, cellular morphology, adherence to plastic and expression of cluster-of-differentiation markers. They varied in their rate of proliferation and trilineage differentiation. The equine BMMSCs of one of 6 donors demonstrated a higher rate of proliferation, enhanced ability for cell passaging and a more robust in vitro differentiation. Comparatively, equine BMMSCs from 2 donors demonstrated a lower rate of proliferation and lack of osteogenic and chondrogenic differentiation. CONCLUSION: The results of this study confirm that donor-to-donor variation in equine BMMSCs exists and this variation can be documented using in vitro assays. Subjective assessment suggests that the rate of proliferation tends to correlate with differentiation potential.

Loss of one but not two mdm2 null alleles alters the tumour spectrum in p53 null mice.

The transcriptional activity of the p53 tumour suppressor is inhibited by binding to MDM2. The in vivo significance of this interaction was established in mdm2 null mice. Embryonic lethality due to loss of mdm2 is completely rescued by deletion of p53, indicating that the lethality is due to inability to down-modulate p53 function. The production of mice null for both p53 and mdm2 led to an assessment of the role of MDM2 in tumour development. Tumour latency and spectrum in p53 null mice were monitored in the presence or absence of mdm2. Two unusual findings resulted: tumour latency in p53 null/mdm2 heterozygous mice was longer than in p53/mdm2 double-null mice; and the incidence of sarcomas was higher in p53 null/mdm2 heterozygous mice than in p53 null or p53/mdm2 double-null mice. These data raise the possibility that heterozygosity at the mdm2 locus in the absence of p53 affects the development of tumours of mesenchymal origin.

Overproduction of MDM2 in vivo disrupts S phase independent of E2F1.

Expression of a beta-lactoglobulin (BLG)/mdm2 transgene (BLGmdm2) in the epithelial cells of the mouse mammary gland causes an uncoupling of S phase from M phase, resulting in polyploidy and tumor formation. The cell cycle defects are independent of interactions with p53. Because MDM2 also binds and activates the S phase-specific transcription factor E2F1, we hypothesized that increased E2F1 activity causes the development of the BLGmdm2 phenotype. We, therefore, generated BLGmdm2 mice that were null for E2F1. We observed no notable differences in histology or cyclin gene expression between BLGmdm2 and BLGmdm2/E2F1-/- mice, indicating that endogenous E2F1 activity was not required for the BLGmdm2 phenotype. Because, depending on the experimental system, either loss of E2F1 function or overexpression of E2F1 results in transformation, we also tested whether overexpression of E2F1 augmented the severity of the BLGmdm2 phenotype by generating mice that were bitransgenic for BLGmdm2 and BLGE2F1. We observed a unique mixture of the two single transgenic phenotypes histologically and found no significant changes in cyclin levels, indicating that overexpression of E2F1 had no effect on the BLGmdm2 transgenic phenotype. Thus, increased expression or absence of E2F1 does not affect the ability of MDM2 to disrupt the cell cycle.