Felícito García-Alvarez

Phenotype and chondrogenic differentiation of mesenchymal cells from adipose tissue of different species.

Mesenchymal stem cells (MSCs) are multipotent cells capable of differentiating into several mesoderm lineages. They have been isolated from different tissues, such as bone marrow, adult peripheral blood, umbilical cord blood, and adipose tissue. The aim of this study was to analyze the differences in proliferation and phenotype of adipose tissue‐derived MSCs from three different species, and to evaluate their capacity to differentiate into chondrocytes in vitro. A comparative study of cultured human, rabbit, and sheep mesenchymal cells from adipose tissue was carried out, and the main morphological parameters, proliferative activity, and expression of surface markers were characterized. Proliferation and flow cytometry data showed species‐related differences between animal and human MSCs. Histological staining suggested that rabbit and sheep mesenchymal cells were able to differentiate into chondrocytic lineages. Human mesenchymal cells, though they could also differentiate, accomplished it with more difficulty than animal MSCs. These results could help to explain the differences in the chondrogenic capacity of sheep and rabbit MSCs when they are used as animal models compared to human mesenchymal cells in a clinical assay.

Musculoskeletal hydatid disease: A report of 13 cases

This is a retrospective study of 13 patients with muscular hydatidosis - i.e., 4% of the 309 cases of hydatid disease treated in our department during 1983-1999. The commonest clinical finding was an asymptomatic and slowly growing mass (7). Puncture or incision of the mass was followed by an infection of the cystic cavity with fistulization in 2 patients. The immunological findings were false negative in 4 patients. MR images were obtained in 4 patients before diagnosis, and were highly suggetive of hydatid disease. The cystic cavities in all 9 patients subjected to radical surgery healed without chemotherapy. Radical surgery was not possible in 4 cases, in 3 of whom the sacrum was involved. Medical treatment of these patients did not eliminate the disease and new operations were necessary.

Evaluation of four experimental osteomyelitis infection models by using precolonized implants and bacterial suspensions.

Staphylococcus aureus osteomyelitis, a major problem in orthopedic surgery, often involves biofilm bacteria adhering to implants and surrounding bone and tissues. The inadequacy of therapy or immunological surveillance has encouraged studies using animal models which simulate natural osteomyelitic infections, ensure the development of infections and avoid mortality. We evaluated 4 models for infection (8 animals/model) in rats, using stainless-steel implants in tibiae and a very adherent slime-producing bacterial strain. Each animal received: an implant containing a 12 h-biofilm with about 10 6 cfu (Model 1); an implant containing this biofilm and a suspension with about 10 4 cfu (Model 2); a sterile implant and a suspension with about 10 5 cfu (Model 3); or a sterile implant and a suspension with about 10 6 cfu (Model 4). 63 days after surgery we found 100% rat survival, colonization of bone by implant biofilm bacteria in some animals and local, but not systemic infections. Model 1 (but not Models 2-4) reproduced an infection in both, tibiae and implants, most reliably (in 100% of the animals). Model 3 was the least reliable (p < 0.01, 25% infected implants, 12% infected tibiae).

Risk factors for postoperative infections in patients with hip fracture treated by means of Thompson arthroplasty

Specific conditions associated with surgery may predispose elderly people to septic complications after hip fracture surgery. This study investigated the risk factors predisposing infection in aged patients with subcapital hip fracture. We performed a prospective study of 290 patients with displaced subcapital hip fracture, operated by means of Thompson hip hemi-arthroplasty (83.5% fractures in women). The mean age was 85.42+/-6.06 years (ranging from 69 to 104). Follow-up was realized until death or at least for 2 years. The chi(2) test, analysis of variance, Kruskal-Wallis test, correlation analysis and the Spearman test were applied. Odds ratios (OR) were calculated. During the hospital stay, there were diagnosed 94 urinary tract infections, 25 pneumonias, 50 superficial wound infections, 11 deep wound infections. Transfusions were made in 120 patients (in average: 2.54+/-1.45 units of red cell concentrate/transfused patient). Transfusion appeared to be correlated with superficial wound infection (OR=1.96), urinary infection (OR=1.76) and pneumonia (OR=2.85). Higher number of days waiting for surgery were related significantly with pneumonia (9.8+/-7.44 days vs. 6.39+/-3.75), or urinary tract infection (7.76+/-4.39 days vs. 6.17+/-4.14). We concluded that the transfusion and longer waiting time for surgery have been associated with the septic complications in elderly patients treated surgically for hip fracture.

Differences in Surface Marker Expression and Chondrogenic Potential among Various Tissue-Derived Mesenchymal Cells from Elderly Patients with Osteoarthritis

Mesenchymal stem cells (MSCs) are self-renewing, multipotent cells that could potentially be used to repair injured cartilage in diseases such as osteoarthritis (OA). In this study we used bone marrow, adipose tissue from articular and subcutaneous locations, and synovial fluid samples from 18 patients with knee OA to find a suitable alternative source for the isolation of MSCs with high chondrogenic potential. MSCs from all tissues analysed had a fibroblastic morphology, but their rates of proliferation varied. Subcutaneous fat-derived MSCs proliferated faster than bone marrow- and Hoffa’s fat pad-derived MSCs, while synovial fluid-derived MSCs grew more slowly. CD36 and CD54 expression was similar across all groups of MSCs with several minor differences. High expression of these surface markers in subcutaneous fat-derived MSCs was correlated with poor differentiation into hyaline cartilage. Synovial fluid-derived MSCs presented a relatively small chondrogenic differentiation capacity while Hoffa’s fat pad-derived MSCs had strong chondrogenic potential. In conclusion, MSCs from elderly patients with OA may still display significant chondrogenic potential, depending on their origin.

Chondrogenic differentiation in femoral bone marrow-derived mesenchymal cells (MSC) from elderly patients suffering osteoarthritis or femoral fracture

This study analyzed the phenotype and the chondrogenic differentiation of bone marrow-derived MSCs from old patients undergoing knee osteoarthritis or femoral fracture surgery. Twenty patients (12 females), with a mean age of 77.35±8.76 years, were studied. Ten patients suffered of knee osteoarthritis (OA) pathology and underwent surgery for arthroplasty, and the other 10 patients suffered femoral fracture. A comparative study of bone marrow-derived cultured human MSCs was carried out, and the main morphological parameters, proliferative activity and expression of surface markers were characterized. Bone marrow was obtained from the femur in all cases. The χ2-test, Mann-Whitney U-test, correlation coefficient and the Spearman test were applied. Bone marrow MSCs from old patients were able to differentiate into chondrocytic lineages. Proliferation and flow cytometry data showed no difference associated to the gender. No significant differences between the knee arthroplasty group or the femoral fracture group were found, except for higher CD49d % in MSC from fracture, and higher CD49f % in MSC from knee OA patients at passage one. MSCs from old patients suffering knee OA can be differentiated into chondrocytic lineages, and these present no differences with MSCs from femoral fracture patients.

Pharmacological Immunomodulation of Surgical Trauma

Surgery and accidental trauma induce changes in the immune response, showing a predominant pattern of activation through the Th2 cell pathway to the detriment of Th1 cell pathway activation. Anapsos is a hydrosoluble extract obtained from Polypodium leucotomos. Anapsos has shown immunomodulating effects in vitro. On a rat experimental model (tibia and fibula fracture), cytokines (interleukin [IL]-2, IL-4, IL-6, IL-10, and IL-12) (enzyme-linked immunosorbent assay, ELISA) and cell percentages of CD4, CD8 CD25, CD122, and CD132 (monoclonal antibodies, MoAb) were determined in peripheral blood 7 days before surgery (PRE), 1 day after surgery (1PO), and 7 days after surgery (7PO). On postoperative day 1, rats undergoing fracture show an increase of CD8 percent expression and IL-6 and IL-10 levels, in contrast to rats undergoing fracture plus anapsos treatment. On postoperative day 7, rats undergoing fracture show an increase of IL-6 levels, whereas rats undergoing fracture plus anapsos do not. The IL-12 level decreases on postoperative day 7 in the group with fracture but not in the fracture plus anapsos group. Thus, we conclude that anapsos is able to modulate the immune response after trauma, inhibiting Th2 pathway activation with no effect on Th1 pathway activation. In trauma, Anapsos could prevent the shifting Th1/Th2 balance.

INFLUENCE OF PLATELET TIME ACTIVATION ON ARTICULAR CARTILAGE GROWTH IN THE RABBIT KNEE: PRELIMINARY STUDY

PURPOSE The aim of the present article was to study the influence of platelets and different time activation on cartilage growth in articular defects in the rabbit knee. METHODS Twelve male New Zealand rabbits (12 weeks) were divided in two groups. Under general anaesthesia, a 4 mm diameter and 2 mm deep defect was performed in medial condyles in both knees. The right knee defect was filled with platelet concentrate 5 min after being activated with ClCa in group A, and 2 min afterwards in group B. Platelets were obtained by centrifuging 10 ml arterial blood from the rabbit prior to the surgical procedure. The left knee defect was not filled. Rabbits were sacrificed 6 weeks after surgery. Macroscopic and microscopic studies were performed. RESULTS In group A, hyaline cartilage was observed in the right knee defect at the end of the experiment in five rabbits. None of the defects of the left knees showed hyaline cartilage growth. In group B, hyaline cartilage was observed in the right knee defect in only one rabbit. Nevertheless, in group B, all rabbits presented better chondral cellularity and regeneration and lower fibrosis in defects treated with platelets than in non-treated ones. CONCLUSIONS This technique for articular defect reconstruction with platelets is simple and easy, and has shown satisfactory results in our study. Platelets may be useful as an autologous source of multiple growth factors for articular defect reconstruction. Nevertheless, this is a preliminary study and further research is required.