Hsiao-Li Ma

Isolation and Characterization of Size‐Sieved Stem Cells from Human Bone Marrow

Bone marrow mesenchymal stem cells (MSCs) have the capacity for renewal and the potential to differentiate into multiple lineages of mesenchymal tissues. In the laboratory, MSCs have the tendency to adhere to culture dish plastic and are characterized by fibroblastic morphology, but possess no specific markers to select them. To isolate and purify MSCs from bone marrow, we use a culture device—a plastic culture dish comprising a plate with 3‐μm pores—to sieve out a homogeneous population of cells (termed size‐sieved [SS] cells) from bone marrow aspirates. SS cells that adhered to the upper porous plate surface were a relatively homogeneous population as indicated by morphology and other criteria, such as surface markers. They had the capacity for self‐renewal and the multilineage potential to form bone, fat, and cartilage, and satisfy the characteristics of MSCs. In addition, if all the cells from each passage had been plated and cultured in our defined conditions, over 1014 SS cells would have been obtained from each 10‐ml aspirate in 15 additional weeks of culture. This technically simple method leads to an efficient isolation and purification of cells with the characteristics of MSCs.

In vitro differentiation of size-sieved stem cells into electrically active neural cells.

Size‐sieved stem (SS) cells isolated from human bone marrow and propagated in vitro are a population of cells with consistent marker typing, and can form bone, fat, and cartilage. In this experiment, we demonstrated that SS cells could be induced to differentiate into neural cells under experimental cell culture conditions. Five hours after exposure to antioxidant agents (β‐mercaptoethanol ± retinoic acid) in serum‐free conditions, SS cells expressed the protein for nestin, neuron‐specific enolase (NSE), neuron‐specific nuclear protein (NeuN), and neuron‐specific tubulin‐1 (TuJ‐1), and the mRNA for NSE and Tau. Immunofluorescence showed that almost all the cells (>98%) expressed NeuN and TuJ‐1. After 5 days of β‐mercaptoethanol treatment, the SS cells expressed neurofilament high protein but not mitogen‐activated protein‐2, glial filament acidic protein, and galactocerebroside. For such long‐term‐treated cells, voltage‐sensitive ionic current could be detected by electrophysiological recording, and the intracellular calcium ion, Ca2+, concentration can be elevated by high potassium (K+) buffer and glutamate. These findings suggest that SS cells may be an alternative source of undifferentiated cells for cell therapy and gene therapy in neural dysfunction.

Is fusion necessary for surgically treated burst fractures of the thoracolumbar and lumbar spine? : A prospective, randomized study

Study Design. A prospective clinical trial was conducted. Objectives. To compare the results of fusion versus nonfusion for surgically treated burst fractures of the thoracolumbar and lumbar spine. Summary of Background Data. The operative results of surgically treated burst fractures with short segmental fixation have been well documented. There is no report comparing the results of fusion and nonfusion. Methods. Fifty-eight patients were included in this study, with the inclusion criteria as follows: neurologically intact spine with a kyphotic angle ≥20°, decreased vertebral body height ≥50% or a canal compromise ≥50%, incomplete neurologic deficit with a canal compromise <50%, complete neurologic deficit, and multilevel spinal injury or multiple traumas. All patients were randomly assigned to fusion or nonfusion groups, and operative treatment with posterior reduction and instrumentation was carried out. Posterior fusion with autogenous bone graft was performed for the fusion group (n = 30), and no fusion procedure was done for the nonfusion group (n = 28). The average follow-up period was 41 months (range, 24–71 months). Results. The average loss of kyphotic angle was not statistically significant between these 2 groups. The radiographic parameters were statistically significantly better in the nonfusion group, including angular change in the flexion-extension lateral view (4.8° vs. 1.0°), lost correction of decreased vertebral body height (3.6% vs. 8.3%), intraoperative estimated blood loss (303 mL vs. 572 mL), and operative time (162 minutes vs. 224 minutes). The scores on the low back outcome scale were not statistically significant for these 2 groups. Conclusions. The short-term results of short segmental fixation without fusion for surgically treated burst fractures of the thoracolumbar spine were satisfactory. The advantages of instrumentation without fusion are the elimination of donor site complications, saving more motion segments, and reducing blood loss and operative time.

Clinical Outcome and Imaging of Arthroscopic Single-Row and Double-Row Rotator Cuff Repair: A Prospective Randomized Trial

PURPOSE The purpose of this study was to compare the clinical and imaging outcomes of single-row and double-row suture anchor fixation in arthroscopic rotator cuff repair with emphasis on analysis of the effect of various tear size on repair integrity. METHODS Fifty-three patents were randomized to either single-row or double-row rotator cuff repair at the time of surgical intervention. The clinical results were evaluated by applying the UCLA score and the ASES index and assessing muscle strength in abduction and external rotation with a minimum 2-year follow-up. The postoperative rotator cuff integrity was evaluated by magnetic resonance arthrography at 6-month and minimum 2-year follow-up. RESULTS We enrolled 27 patients in the single-row group and 26 patients in the double-row group. Statistically, the UCLA score; the ASES index; and muscle strength were significantly increased in both groups after surgery, but there was no significant difference between the 2 groups. At minimum 2-year follow-up, intact rotator cuffs were found in 17 patients in the single-row group and 20 in the double-row group, based on magnetic resonance arthrography results. Overall, there was no significant difference in postoperative structural integrity between the 2 groups at 6-month and 2-year follow-up. In patients with tear size larger than 3 cm, the muscle strength of the shoulder was significantly better in the double-row group. For the final imaging results, regardless of the tear size, there was no difference between the single-row and double-row groups. CONCLUSIONS Arthroscopic rotator cuff repair with double-row fixation showed better shoulder strength in patients with larger tear size (>3 cm) in comparison with single-row fixation. However, the imaging results showed no significant difference in cuff integrity in both groups in patients with any tear size at 6-month and minimum 2-year follow-up. LEVEL OF EVIDENCE Level II, lesser-quality randomized control trial.

Comparison of shoulder ultrasound and MR imaging in diagnosing full-thickness rotator cuff tears

Ultrasound (US) and magnetic resonance imaging (MRI) of 422 cases were evaluated to compare the feasibility in diagnosing full-thickness rotator cuff tears (FTRCTs). On the basis of different US performers, they were divided into two groups: Group 1 performed by a 5-year experience technician and Group 2 performed by a 10-year experience radiologist. Sensitivity, negative predictive value (NPV), accuracy of US, and correlation between the two modalities were better in Group 2. When an expert is available, US can be used for diagnosing FTRCTs; otherwise, MRI should be performed.

Difference in femoral head and neck material properties between osteoarthritis and osteoporosis.

BACKGROUND Osteoarthritis and osteoporosis are the two most common musculoskeletal diseases found in the aged population. It is of interest to measure and study the material properties of the femoral head and neck of these two groups, and hopefully to offer explanation of the observed phenomenon that most patients suffer from one of the two disorders, not both. METHODS Seven osteoarthritic and seven osteoporotic femoral heads were used for this study. The principal compressive region of the femoral heads were cut to determine the Youngs modulus and yielding stress by a material testing machine. Comparisons between these two groups were conducted by using material properties and the properties normalized by individual patient physical parameters, including body weight, body height and femoral head diameter, respectively. The finite element model of femoral neck cuboid in OA and OP were obtained based on the micro-CT-scan cross-section. The intrinsic material properties were calculated from the solid FE models. FINDINGS The results showed significant differences in density, modulus and strength between the osteoarthritic and osteoporotic femoral heads as measured, with the former having 2-3 times the values of the latter. Femoral head diameter has stronger influence in mechanical properties than patients body weight and body height. Regarding to bone volume (BV), bone surface (BS), bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), and true trabecular elastic modulus, the intrinsic material properties of femoral neck with OA were higher than OP. INTERPRETATION It is still unknown why patients do not suffer from both osteoporosis and osteoarthritis at the same time. Many studies aimed to investigate the mechanical property of two groups. However, individual difference of the femoral head and neck is too difficult to obtain a reasonable comparison between these two groups. This study investigated the two groups more quantitatively and further estimated the factors which influence mechanical properties from a biomechanical point of view.

Mesenchymal Stem Cells From a Hypoxic Culture Improve and Engraft Achilles Tendon Repair

Background: Bone marrow–derived mesenchymal stem cells (MSCs) from humans cultured under hypoxic conditions increase bone healing capacity. Hypothesis: Rat MSCs cultured under hypoxic conditions increase the tendon healing potential after transplantation into injured Achilles tendons. Study Design: Controlled laboratory study. Methods: Biomechanical testing, histological analysis, and bromodeoxyuridine (BrdU) labeling/collagen immunohistochemistry were performed to demonstrate that augmentation of an Achilles tendon rupture site with hypoxic MSCs increases healing capacity compared with normoxic MSCs and controls. Fifty Sprague-Dawley rats were used for the experiments, with 2 rats as the source of bone marrow MSCs. The cut Achilles tendons in the rats were equally divided into 3 groups: hypoxic MSC, normoxic MSC, and nontreated (vehicle control). The uncut tendons served as normal uncut controls. Outcome measures included mechanical testing in 24 rats, histological analysis, and BrdU labeling/collagen immunohistochemistry in another 24 rats. Results: The ultimate failure load in the hypoxic MSC group was significantly greater than that in the nontreated or normoxic MSC group at 2 weeks after incision (2.1 N/mm2 vs 1.1 N/mm2 or 1.9 N/mm2, respectively) and at 4 weeks after incision (5.5 N/mm2 vs 1.7 N/mm2 or 2.7 N/mm2, respectively). The ultimate failure load in the hypoxic MSC group at 4 weeks after incision (5.5 N/mm2) was close to but still significantly less than that of the uncut tendon (7.2 N/mm2). Histological analysis as determined by the semiquantitative Bonar histopathological grading scale revealed that the hypoxic MSC group underwent a significant improvement in Achilles tendon healing both at 2 and 4 weeks when compared with the nontreated or normoxic MSC group via statistical analysis. Immunohistochemistry further demonstrated that the hypoxic and normoxic MSC groups had stronger immunostaining for type I and type III collagen than did the nontreated group both at 2 and 4 weeks after incision. Moreover, BrdU labeling of MSCs before injection further determined the incorporation and retention of transplanted cells at the rupture site. Conclusion: Transplantation of hypoxic MSCs may be a better and more readily available treatment than normoxic MSCs for Achilles tendon ruptures. Clinical Relevance: The present study provides evidence that transplantation of hypoxic MSCs may be a promising therapy for the treatment of Achilles tendon ruptures.

Isolation of mesenchymal stem cells from shoulder rotator cuff: a potential source for muscle and tendon repair.

The self-healing potential of each tissue belongs to endogenous stem cells residing in the tissue; however, there are currently no reports mentioned for the isolation of human rotator cuff-derived mesenchymal stem cells (RC-MSCs) since. To isolate RC-MSCs, minced rotator cuff samples were first digested with enzymes and the single cell suspensions were seeded in plastic culture dishes. Twenty-four hours later, nonadherent cells were removed and the adherent cells were further cultured. The RC-MSCs had fibroblast-like morphology and were positive for the putative surface markers of MSCs, such as CD44, CD73, CD90, CD105, and CD166, and negative for the putative markers of hematopoietic cells, such as CD34, CD45, and CD133. Similar to BM-MSCs, RC-MSCs were demonstrated to have the potential to undergo osteogenic, adipogenic, and chondrogenic differentiation. Upon induction in the defined media, RC-MSCs also expressed lineage-specific genes, such as Runx 2 and osteocalcin in osteogenic induction, PPAR-γ and LPL in adipogenic differentiation, and aggrecan and Col2a1 in chondrogenic differentiation. The multipotent feature of RC-MSCs in the myogenic injury model was further strengthened by the increase in myogenic potential both in vitro and in vivo when compared with BM-MSCs. These results demonstrate the successful isolation of MSCs from human rotator cuffs and encourage the application of RC-MSCs in myogenic regeneration.

Isolation and characterization of mesenchymal stromal cells from human anterior cruciate ligament

BACKGROUND The anterior cruciate ligament (ACL) is one of the most commonly injured ligaments of the knee. Because the torn ACL is always discarded during ACL reconstruction, it may be a potential source for isolating mesenchymal stromal cells (MSC). METHODS To characterize MSC from human ACL, cells were enzymatically released from the ACL of adult human donors and seeded in plastic dishes with serial passages at confluence. At different passages, ACL-derived cells were subjected to in vitro assays to investigate their multilineage potential. Upon treatment, the phenotypes of the cell cultures were analyzed by histo- and immunohistochemistry and semi-quantitative reverse transcription-polymerase chain reaction for the expression of lineage-specific genes. RESULTS Six independent cell lines from individual donors showed diversity in multilineage potential. Interestingly, five of the six lines displayed adipogenic potential, four had osteogenic and adipogenic potential, and only one cell line was tripotent. Both bone marrow (BM)- and ACL-derived MSC expressed marker genes for ligament fibroblasts, whereas the mRNA levels of collagen I and III were more abundant in ACL-derived MSC. DISCUSSION Our study demonstrates that human MSC can be isolated from ACL with diversity in the potential to form bone, fat and cartilage and an increase as compared to BM MSC, in the potential to form ligament fibroblasts.

Imaging evaluation of meniscal injury of the knee joint: A comparative MR imaging and arthroscopic study

To evaluate the efficacy of MR imaging in the diagnosis and classification of meniscal tear of the knee joint, we retrospectively characterized the MR features of 78 meniscal tears in 148 patients according to the Mesgarzadehs criteria. The results showed that the sensitivity and specificity for meniscal tears were 92% and 87%, respectively. Type VI meniscal tear was the most common type, especially in displaced meniscal tear. MR is a reliable diagnostic tool for meniscal tears and associated cruciate ligament injury.