Neurovascular plasticity in the knee joint of an arthritic mouse model

Abstract

Lower numbers of neuropeptide-containing fibers in arthritic joints have been found as compared to control joints. This may be the result of fiber depletion, necrosis of fibers, or proliferation of soft tissues without neural sprouting. To discriminate between these possibilities, we studied the relationships between soft tissue proliferation, changes in vascularity of synovial tissues, and changes in joint innervation during arthritis. Arthritis was induced in the knee joint of mice by a single subpatellar injection of methylated bovine serum albumin after previous immunization. Antibodies to protein gene product 9.5, S-100, and growth-associated protein-43 (GAP-43) were used to study the general innervation pattern. Antibodies to calcitonin gene-related peptide (CGRP), vasointestinal polypeptide (VIP), substance P (SP), and tyrosine hydroxylase (TH) were used to localize sensory (SP, CGRP, VIP) and sympathetic (TH) fibers. Blood vessels of the joint were studied with ink perfusion, GAP-43, and a vascular marker (LF1). Directly after the induction of arthritis, the synovial cavity was enlarged and filled with leukocytes. From day 4 onward, small sprouting blood vessels penetrated the avascular mass of cells in the joint cavity. After 1 week, the vascular sprouting activity and GAP-43 immunoreactivity were maximal, and after 2 weeks, vascular sprouting activity diminished. In the subsequent period, the synovia slowly regained their prearthritic appearance and thickness. The most pronounced changes in the general staining pattern of CGRP, SP, VIP, and TH were found in the periosteum. From 2 days to 4 weeks after the induction of arthritis, the layer of SP, CGRP, and VIP fibers in the femoral periosteum was thicker and more irregular. GAP-43 staining showed many terminal varicosities, which suggested sprouting of nerve fibers. From 2 days to 2 weeks after the induction of arthritis, the SP and CGRP fibers in the periosteum showed gradual depletion. In the thickened subsynovial tissues that were revascularized, no ingrowth of neural elements was found. As the total number of nerve fibers in the synovial tissue did not change, large parts of the synovia directly facing the joint cavity were not innervated at 1 week after the induction of arthritis. These results strongly suggest that periosteal SP and CGRP fibers were depleted during arthritis. Synovial proliferation without concomitant fiber growth is the main cause of the reduced number of immunocytochemically detectable fibers in the mouse arthritic knee joint.

Quantitation of the changes in vascularity during arthritis in the knee joint of a mouse with a digital image analysis system

“…The relatively low vessel density on Day 4 and the increase in vessel density on Day 7 as compared with Day 4, strongly suggests that directly after the induction of arthritis, the size of the synovial area increases rapidly without vessel formation and that after Day 4, vessel formation increases. This was confirms previous preliminary findings in that blood vessels sprouted, particularly between Days 4 and 7 (Buma et al, 2000). The relative avascularity of the synovial membrane during the first few days after the induction of arthritis might induce hypoxia, which is a powerful inducer of angiogenesis (Blake et al, 1989;Fava et al, 1994;Paleolog, 1996;Stevens et al, 1991).…”

“…This is in agreement with our observation that sprouting vessels end blindly in the synovial tissue, making actual flow impossible (Buma et al, 2000). The perfused vessel density (VDp) on Day 7 was again comparable to that of the control knee on day seven, indicating that after a few days, newly formed vessels become functional.…”

“…In this experiment the mBSA-induced arthritis model exhibited highly reproducible swelling of the tissues of the synovial membrane on Days 4 and 7. Other experiments have shown that between 2 and 8 weeks the tissues of the synovial membrane will slowly return to their pre-arthritic configuration (van Beuningen et al, 1989;Buma et al, 2000).…”

“…On Day 4 after induction of arthritis, the swollen subsynovial membrane tissues were avascular (Buma et al, 2000). An important observation was that in the first stages of the inflammation process, the area of the synovial membrane (SA) increased significantly.…”

“…Cells in this tissue that may even grow out as pannus tissue over the surface of the cartilage, produce catabolic factors for cartilage (Ben et al, 1995;Colville-Nash and Scott, 1992;Fava et al, 1994;FitzGerald et al, 1991). The expansion of this hyperplastic synovial tissue is associated with an extensive prolif- eration of blood vessels (Blake et al, 1989;Buma et al, 2000;Colville-Nash and Scott, 1992). The regulation of neovascularization in the synovium of arthritic joints is complex and various growth factors and inflammatory mediators may be involved (Ben et al, 1995;Blake et al, 1989;Claffey and Robinson 1996;FitzGerald et al, 1991;Nagashima et al, 1995;Stevens et al, 1991).…”