The effects of plasma rich in growth factors (PRGF-Endoret) on healing of medial collateral ligament of the knee.

PURPOSE: Platelet-rich plasma (PRP) has been increasingly used in sports-related injuries for therapeutic applications. However, there are numerous manufacturing procedures and treatment protocols of PRP use, which make difficult to assess its real efficacy for tissue healing. This study addressed to evaluate the therapeutic effects of locally delivered plasma rich in growth factors (PRGF-Endoret) on the early healing of medial collateral ligament (MCL) in rabbit knees. METHODS: Thirty-one Japanese white rabbits were subjected to a mop-end tear in the MCL of the left knee. PRGF-Endoret was prepared using Anitua's technique. Two groups were set up. In 17 knees, prepared 1.0 ml of PRGF-Endoret after clotting was applied on the tear site, while in 14 knees the tear site was untreated serving as a control. Quantitative aspects of PRGF-Endoret, the concentration of platelets, leukocytes and erythrocytes and therapeutic growth factors such as PDGF-BB and TGF-ß1 were measured. Rabbits were sacrificed at 3 and 6 weeks after the operation and histological and biomechanical evaluation were performed. RESULTS: No leukocytes were measured and certain amount of growth factors such as PDGF-BB and TGF-ß1 were confirmed in the PRGF-Endoret. PRGF-Endoret stimulated proliferation of fibroblasts and neovascularization, and induced statistically better structural properties in repaired MCL. CONCLUSIONS: Our findings provide evidence that local administration of PRGF-Endoret promotes early steps in ligament healing and the repair of structural properties in a rabbit model. PRGF-Endoret would be a useful product in clinical treatment of ligament injuries.

Role of angiogenesis after muscle derived stem cell transplantation in injured medial collateral ligament.

We performed this study to investigate the therapeutic role of vascular endothelial growth factor (VEGF) in medial collateral ligament (MCL) healing. Murine muscle derived stem cells (MDSCs) obtained via the preplate technique were retrovirally transduced to express: (1) VEGF and nLacZ (MDSC-VEGF), (2) soluble fms-like tyrosine kinase-1 (sFLT1, a VEGF-specific antagonist) and nLacZ (MDSC-sFLT1), and (3) nLacZ (MDSC-nLacZ). After transecting the MCL of immunodeficient rats, 5 × 10(5) cells of each of the transduction groups list above were transplanted into the MCL injury site. A control group was injected with phosphate-buffered saline (PBS) only. Immunohistochemical staining demonstrated that there were more Isolectin B4 and ß-galactosidase double positive cells in the rats transplanted with MDSC-VEGF transduced cells than the other groups at week 1. Capillary density was significantly higher in the MDSC-VEGF group than the other groups at week 2; however, there were no significant differences in the biomechanical assessment between the MDSC-VEGF and MDSC-nLacZ groups. On the other hand, the MDSC-sFLT1 group revealed a lower capillary density than the other two groups and the functional ligament healing of the MDSC-sFLT1 group was significantly decreased compared to the other groups when assessed biomechanically. The findings of the present study suggest that angiogenesis plays a critical role in the healing process of injured MCL.

Injury-induced changes in mRNA levels differ widely between anterior cruciate ligament and medial collateral ligament.

BACKGROUND: The drastic difference in healing capacity between the anterior cruciate ligament and the medial collateral ligament is still largely unexplained. Few studies have compared the profiles of messenger ribonucleic acid expression for healing-associated molecules in ligaments during the course of healing. HYPOTHESIS: Injury responses of the injured anterior cruciate ligament and medial collateral ligament are characterized by very different profiles of angiogenesis-promoting and repair-associated gene expression during the healing process. STUDY DESIGN: Controlled laboratory study. METHODS: Reverse-transcriptase polymerase chain reaction was used to assay expression of messenger ribonucleic acid for 11 healing- and angiogenesis-associated molecules at 3 days and 2, 6, and 16 weeks after anterior cruciate ligament or medial collateral ligament injury in adult female New Zealand White rabbits. RESULTS: Marked differences were found in the postinjury changes in messenger ribonucleic acid levels in the anterior cruciate ligament compared to the medial collateral ligament. Notably, messenger ribonucleic acid levels for the important repair-associated growth factor transforming growth factor-beta1 did not increase in injured anterior cruciate ligament at any time point. Similarly, unlike the injured medial collateral ligament, no statistically significant increases in messenger ribonucleic acid levels for the important scar matrix protein collagen III were detected in injured anterior cruciate ligament. In contrast, matrix metalloproteinase messenger ribonucleic acid levels were markedly elevated in injured anterior cruciate ligament but only modestly increased in medial collateral ligament. CONCLUSION: The results suggest that injury leads to an antifibrotic, catabolic response in the rabbit anterior cruciate ligament, possibly to prevent fibrosis and diminish the risk for loss of joint motion. CLINICAL RELEVANCE: The development of effective biologically based treatments for anterior cruciate ligament injuries will need to incorporate strategies to deal with the significant differences in the molecular responses to injury of these tissues.

Nerve growth factor improves ligament healing.

Previous work has shown that innervation participates in normal ligament healing. The present study was performed to determine if exogenous nerve growth factor (NGF) would improve the healing of injured ligament by promoting reinnervation, blood flow, and angiogenesis. Two groups of 30 Sprague-Dawley rats underwent unilateral medial collateral ligament transection (MCL). One group was given 10 microg NGF and the other was given PBS via osmotic pump over 7 days after injury. After 7, 14, and 42 days, in vivo blood flow was measured using laser speckle perfusion imaging (LSPI). Morphologic assessments of nerve density, vascularity, and angiogenesis inhibitor production were done in three animals at each time point by immunohistochemical staining for the pan-neuronal marker PGP9.5, the endothelial marker vWF, and the angiogenesis inhibitor thrombospondin-2 (TSP-2). Ligament scar material and structural mechanical properties were assessed in seven rats at each time point. Increased nerve density was promoted by NGF at both 14 and 42 days. Exposure to NGF also led to increased ligament vascularity, as measured by histologic assessment of vWF immunohistochemistry, although LSPI-measured blood flow was not significantly different from controls. NGF treatment also led to decreased expression of TSP-2 at 14 days. Mechanical testing revealed that exposure to NGF increased failure load by 40%, ultimate tensile strength by 55%, and stiffness by 30% at 42 days. There were no detectable differences between groups in creep properties. The results suggest that local application of NGF can improve ligament healing by promoting both reinnervation and angiogenesis, and results in scars with enhanced mechanical properties.

Physiological and mechanical adaptations of rabbit medial collateral ligament after anterior cruciate ligament transection.

Progressive physiological and mechanical changes in the medial collateral ligament of the adult rabbit were investigated for as long as 48 weeks after disruption of the anterior cruciate ligament. Eighty-one New Zealand White rabbits were separated into experimental, sham-operated control, and normal control groups. The experimental group underwent unilateral transection of the right anterior cruciate ligament, sham-operated animals served as controls for comparison, and normal animals were evaluated as age-matched, undisturbed (no surgery) controls. Blood flow to the medial collateral ligament (as a physiological measure) and mechanical function (structural and material properties) were assessed at 6, 14, and 48 weeks. The results indicated that loss of the anterior cruciate ligament leads to early mechanical deterioration of the medial collateral ligament with a corresponding loss of physiological homeostasis. Six to 14 weeks after the transection, values for cross-sectional area of the medial collateral ligaments rapidly increased to 1.5 times control values. The ligament became twice as large as the control ligament by 48 weeks. Concomitantly, medial collateral ligament stress at failure of the medial collateral ligament complex decreased rapidly 6-14 weeks after the transection and eventually fell to one-half that of controls by 48 weeks. In terms of low-load behaviour, laxity and load relaxation were significantly greater than that of controls 6 weeks after transection and were further increased by 14 weeks. By 48 weeks, laxity values had recovered somewhat and load-relaxation measures had recovered to near control values. At both 6 and 14 weeks, a statistically significant elevation in blood flow was demonstrated compared with controls. By 48 weeks, however, blood flow was no different from that of the sham-operated control. Thus, early after transection of the anterior cruciate ligament, both low-load and high-load mechanical properties of the medial collateral ligament deteriorated and the rate of blood flow was temporarily elevated. By 48 weeks, blood flow declined to near control values, with a corresponding recovery in viscoelastic behaviour. These findings suggest that, after transection of the anterior cruciate ligament, viscoelastic behaviour of the medial collateral ligament may be related to changes in blood flow and that restoration of normal flow patterns and vascular responses may be linked to the recovery of some low-load mechanical properties in the anterior cruciate ligament-deficient medial collateral ligament.