Blockade of the sympathetic nervous system degrades ligament in a rat MCL model.

We hypothesize that blockade of the sympathetic nervous system degrades ligament. We tested this hypothesis in a rat medial collateral ligament (MCL) model. Fifteen animals were treated for 10 days with the sympathetic chemotoxin guanethidine using osmotic pumps, whereas 15 control rats received pumps containing saline. A reduction in plasma concentrations of norepinephrine in the guanethidine rats indicated a significant decrease in sympathetic nerve activity. Vasoactive intestinal peptide and neuropeptide Y were decreased in MCLs from guanethidine animals, as quantified by radioimmunoassays. Tissue vascularity was substantially increased in guanethidine MCLs, whereas mechanical properties were significantly decreased. Proteases, such as matrix metalloproteinases (MMP) and cysteine proteases, play a major role in ligament degradation. The proteases MMP-13, cathepsin K, and tartrate-resistant acid phosphatase (TRAP) have collagenolytic activity and have been shown in rat ligament tissues. To determine whether the degradation seen in this study was due to protease activity, we determined the expression of these enzymes in control and treated MCLs. Real-time quantitative PCR revealed that guanethidine treatment increased expression of MMP-13 and cathepsin K mRNAs, although overall expression levels of MMP-13 and TRAP were relatively low. Histology also identified increases in TRAP and cathepsin K, but not MMP-13, in guanethidine-treated tissues. Results support our hypothesis that blockade of the sympathetic nervous system substantially degrades ligament.

Hindlimb unloading alters ligament healing.

We investigated the hypothesis that hindlimb unloading inhibits healing in fibrous connective tissue such as ligament. Male rats were assigned to 3- and 7-wk treatment groups with three subgroups each: sham control, ambulatory healing, and hindlimb-suspended healing. Ambulatory and suspended animals underwent surgical rupture of their medial collateral ligaments, whereas sham surgeries were performed on control animals. After 3 or 7 wk, mechanical and/or morphological properties were measured in ligament, muscle, and bone. During mechanical testing, most suspended ligaments failed in the scar region, indicating the greatest impairment was to ligament and not to bone-ligament insertion. Ligament testing revealed significant reductions in maximum force, ultimate stress, elastic modulus, and low-load properties in suspended animals. In addition, femoral mineral density, femoral strength, gastrocnemius mass, and tibialis anterior mass were significantly reduced. Microscopy revealed abnormal scar formation and cell distribution in suspended ligaments with extracellular matrix discontinuities and voids between misaligned, but well-formed, collagen fiber bundles. Hence, stress levels from ambulation appear unnecessary for formation of fiber bundles yet required for collagen to form structurally competent continuous fibers. Results support our hypothesis that hindlimb unloading impairs healing of fibrous connective tissue. In addition, this study provides compelling morphological evidence explaining the altered structure-function relationship in load-deprived healing connective tissue.