Temporal analysis of equine bone marrow aspirate during establishment of putative mesenchymal progenitor cell populations.

Mesenchymal progenitor cells (MPCs) are often characterized using surface markers after expansion and treatment in culture. There are no studies directly comparing gene and protein markers in undifferentiated samples during the very early phases of culture. The goal of this study was to evaluate temporal gene and protein expression changes during establishment of equine MPC cultures. Bone marrow aspirate was obtained from 35 horses and processed by density gradient centrifugation. In freshly isolated bone marrow, mononuclear cells had variable expression of CD44, CD11a/CD18, CD90, and CD45RB cell surface molecules. After 2 h of culture, bone marrow mononuclear cells had a phenotype of CD44(hi), CD29(hi), CD90(lo), CD11a/CD18(hi), and CD45RB(lo). Isolated mononuclear cells were analyzed by flow cytometry and RT-qPCR at 2, 7, 14, 21, and 30 days of culture. At all culture time points, gene expression was in agreement with cell surface protein expression. In established cultures of MPCs, cells remained robustly positive for CD44 and CD29. The proportion of positive cells and the mean fluorescence intensity of positive cells increased in CD90 expression as MPC cultures became more homogeneous. Inversely, the population of cells in culture decreased expression of CD11a/CD18 and CD45RB molecules over time. The decreased expression of the latter molecules makes these useful negative markers of established MPC cultures under normal expansion conditions. The results of this study demonstrate numerous dynamic changes in cell surface molecule expression during early establishment of MPC populations, which may aid to improve MPC isolation methods for research or therapeutic applications.

A comparison of laryngoplasty and modified partial arytenoidectomy as treatments for laryngeal hemiplegia in exercising horses.

OBJECTIVE: To compare upper airway mechanics, arterial blood gases, and tracheal contamination in horses with induced left laryngeal hemiplegia (recurrent laryngeal neuropathy [RLN]) treated by laryngoplasty/vocal cordectomy (LPVC) or modified partial arytenoidectomy (MPA). STUDY DESIGN: Repeated measures under the following conditions: Control, RLN, LPVC, and MPA. ANIMALS: Six horses. METHODS: Two trials were conducted under all conditions at 80% and 100% of maximal heart rate (HR(max)). In Trial 1, arterial blood gases, tracheal and pharyngeal pressures, and laryngeal videoendoscopy were recorded. In Trial 2, upper airway pressure and airflow were determined. Tracheobronchial aspirates were performed after exercise to quantify airway contamination. RESULTS: Compared with control, RLN significantly increased inspiratory impedance and worsened exercise-induced hypoxemia. At 80% HR(max), LPVC restored most variables to control values. At 100% HR(max), LPVC improved all variables, but did not restore minute volume, arterial pH, and PaCO(2). At 80% HR(max), MPA restored all variables except bicarbonate to control values. At 100% HR(max), MPA improved all variables, but did not statistically restore minute ventilation or bicarbonate level. Only minor differences were noted between LPVC and MPA. Both resulted in equivalent tracheal contamination. CONCLUSIONS: Airway mechanics and arterial blood gas values were not restored to normal after either LPVC or MPA in horses exercising at HR(max). This does not affect ventilation at sub-maximal exercise, but has clinical implications at HR(max). Both procedures diminish normal laryngeal protective mechanisms. CLINICAL RELEVANCE: At sub-maximal exercise intensities both LPVC and MPA restore airway ventilation to normal. At maximal exercise the superiority of LPVC over MPA is slight.