Cell-based therapies offer a promising strategy to treat tendon injuries and diseases. Both mesenchymal stromal cells (MSCs) and pluripotent stem cells (PSCs) are good candidates for such applications due to their self-renewing and differentiation capacity. However, the translation of cell-based therapies from bench to bedside can be hindered by the use of animal-derived components in ancillary materials and by the lack of standardised media and protocols for in vitro tenogenic differentiation. To address this, we have optimized animal component-free (ACF) workflows for differentiating human MSCs and PSCs to tenocyte-like cells (TLCs) respectively. MSCs isolated from bone marrow (n = 3) or adipose tissue (n = 3) were expanded using MesenCult™-ACF Plus Culture Kit for at least 2 passages, and differentiated to TLCs in 21 days using a step-wise approach. Briefly, confluent cultures were treated with an ACF tenogenic induction medium for 3 days, followed by treatment with an ACF maturation medium for 18 days. Monolayer cultures were maintained at high density without passaging for the entire duration of the protocol, and the medium was changed every 2 – 3 days. In a similar fashion, embryonic (n = 3) or induced PSCs (n = 3) were first differentiated to acquire a mesenchymal progenitor cell (MPC) phenotype in 21 days using STEMdiff™ Mesenchymal Progenitor Kit, followed by the aforementioned tenogenic protocol for an additional 21 days. In all cases, the optimized workflows using ACF formulations consistently activated a tenogenic transcriptional program, leading to the generation of elongated, spindle-shaped tenomodulin-positive (TNMD+) cells and deposition of an extracellular matrix predominantly composed of collagen type I. In summary, here we describe novel workflows that can robustly generate TLCs from MSCs and hPSC-derived MPCs for potential translational applications.
Mesenchymal stromal cells (MSCs) have been one of the most widely studied cell types in preclinical and clinical trials, due to their self-renewing, multipotent capacity, immunomodulatory properties and relative ease of isolation from multiple tissues. Despite limitations and safety concerns, fetal bovine serum (FBS) is still predominantly used for MSC expansion in clinical protocols. In addition, the undefined nature of serum composition and lot-to-lot variability have been linked to reduced reproducibility and efficiency of MSC bioprocessing. Moreover, use of animal serum in human cell culture increases the risk of contamination with adventitious pathogenic microorganisms, such as viruses, prions and bacteria. Hence, a defined serum-free formulation can provide increased safety, better control over physiological responsiveness, consistent performance and reproducible results. Here we present preliminary data on a prototype serum-free medium optimized for
We aimed to determine the impact of anaesthetic techniques on post-operative analgesic utilisation in people undergoing total knee arthroplasty. We conducted a prospective study at a district general hospital to assess post-operative analgesia use via patient controlled analgesia (PCA) in patients undergoing a elective total knee arthroplasty. Subjects were divided into three groups; group 1 (general anaesthetic and local nerve block, n = 39), group 2 (spinal only, n= 39) and group 3 (general anaesthetic only, n = 38). The primary outcome measure was post-operative morphine consumption through a PCA within the first 24 hours. All subjects were followed up post-operatively by a dedicated pain control nurse. Data was analysed using SPSS version 17 for Windows (SPSS Inc, Chicago, IL, USA). Results are reported as mean (SD) unless stated otherwise. No significant difference in the age or sex distribution between groups was demonstrable. There was no statistical difference in the average usage of morphine when comparing Group 1 compared to group 2, (77.57 (49.56) vs 65.80 (44.71), p=0.27), group 1 compared to group 3, (77.57 (49.56) vs 77.80 (45.52) p=0.98) and finally group 2 compared to group 3, (65.80 (44.71) vs 77.80 (45.52), p=0.25). In this cohort of patients undergoing total knee arthroplasty the anaesthetic technique used does not appear to have an influence on the post-operative pain management.
Summary of Background Data: Spinal deformities, occurring in patients with cerebral palsy and good ambulatory capacity, are infrequently associated with pelvic obliquity, and so instrumented spinal fusions traditionally do not extend to the pelvis.
Purpose. The aim of this study was to document rate of survival among 288 severely affected pediatric patients with spasticity and neuromuscular scoliosis who underwent spinal fusion and to identify exposure variables that could significantly predict survival times. Methods. Kaplan-Meier survivorship analysis was performed and Cox’s proportional hazards model was used to evaluate predictive efficacy of exposure variables such as gender, age at surgery, level of ambulation, mental ability, degree of coronal and sagittal plane spinal deformity, intraoperative blood loss, surgical time, days in the hospital, and days in the intensive care unit (ICU). Results. The statistical analysis demonstrated a mean predicted survival of 134.3 months (11.2 years) after spine surgery for this group of globally involved children with cerebral palsy (Figure 1). The number of days in the ICU after surgery and the presence of severe preoperative thoracic hyperkyphosis were the only factors affecting survival rates. ICU stay of greater than five days, which was usually associated with respiratory problems, substantially increased the risk of death. Thoracic hyperkyphosis of greater than 70° caused a considerable increase in the predicted mortality rate. Conclusions. Our study demonstrated a relatively long mean predicted survivorship for children and adolescents with severe spastic cerebral palsy and neuromuscular scoliosis who underwent spine surgery, which is consistent with the current concept of increased life expectancy even for the total-body involved patients. The most accurate determinants for survival rates among this population group were the number of days the patient had to spend postoperatively in the intensive care unit, and the presence of excessive thoracic hyper-kyphosis.
Purpose: To evaluate the outcome of spinal fusion with unit rod in pediatric patients with cerebral palsy who were treated by the two senior authors using the same operative technique. Methods: This is a retrospective study of 288 patients with mean age at surgery 13.9 years (SD: 3.26), whose medical charts and radiographs were reviewed. A questionnaire including 14 questions assessing patients’ functional improvement was given to the caretakers. Results: Mean radiographic follow up of 3.2 years (range: 1-9.9) was available in 213 patients. In 46 patients anterior-posterior fusion was performed and in 242 only posterior. The preop. Cobb angle was 740(range: 6-176°) corrected by 68% to 230 (p<
.01) and increased by 20 in the last follow-up (p<
.01). The preop. pelvic obliquity was 170(range: 0-57°), corrected by 73% to 4.70 (p<
.01) and increased by 0.40 at follow-up. The preop. kyphosis angle was 560(range: -44-130°), corrected by 37% to 350 (p<
.01) and increased by 20 at follow-up. The preop. lordosis angle was 380(range: -50-140°), corrected by 14% to 430 (p<
.05) and increased by 1.40 at follow-up. There was a strong correlation between hyperlordosis and days of hospitalization, blood loss and surgical time (r= -.22, .23, .24). Patients with lordotic angle >
600 developed 15.1% technical problems related to pelvic fixation, whereas only 3.4% of those with <
600. The mean days of hospitalization were 19.6, the mean blood loss 2.9lt (1.2BV), the mean surgical time 4.4h (Anterior-Posterior: 7h, Posterior only: 3.9h) and the mean ICU stay 5 days. There was a difference only in surgical time (p<
.01) if an anterior procedure was required. The major complications included 3 perioperative deaths, 14 deep infections, 13 (4.5%) reop-erations for mechanical problems and no detected pseud-arthrosis. The answers received by the caretakers were 99% positive, emphasizing the improvement in child’s appearance, sitting ability and respiratory function. Conclusion: Spine surgery in patients with cerebral palsy was accompanied by a considerable blood loss and multiple medical complications. A very satisfactory correction of spinal curvatures was achieved and maintained in follow up. Excessive lumbar lordosis was associated with a high incidence of technical problems and an increased morbidity. There were no pseudarthrosis and the overall number of reoperations for technical reasons was very low. The caretakers were extremely pleased with the outcome of this procedure.