Marked differences in local bone remodelling in response to different marrow stimulation techniques in a large animal

Abstract

Marrow stimulation, including subchondral drilling and microfracture, is the most commonly performed cartilage repair strategy, whereby the subchondral bone plate is perforated to release marrow-derived cells into a cartilage defect to initiate repair. Novel scaffolds and therapeutics are being designed to enhance and extend the positive short-term outcomes of this marrow stimulation. However, the translation of these newer treatments is hindered by bony abnormalities, including bone resorption, intralesional osteophytes, and bone cysts, that can arise after marrow stimulation. In this study, three different marrow stimulation approaches – microfracture, subchondral drilling and needle-puncture – were evaluated in a translationally relevant large-animal model, the Yucatan minipig. The objective of the study was to determine which method of marrow access (malleted awl, drilled Kirschner wire or spring-loaded needle) best preserved the underlying subchondral bone. Fluorochrome labels were injected at the time of surgery and 2 weeks post-surgery to capture bone remodelling over the first 4 weeks. Comprehensive outcome measures included cartilage indentation testing, histological grading, microcomputed tomography and fluorochrome imaging. Findings indicated that needle-puncture devices best preserved the underlying subchondral bone relative to other marrow access approaches. This may relate to the degree of bony compaction occurring with marrow access, as the Kirschner wire approach, which consolidated bone the most, induced the most significant bone damage with marrow stimulation. This study provided basic scientific evidence in support of updated marrow stimulation techniques for preclinical and clinical practice.

Promoting endogenous articular cartilage regeneration using extracellular matrix scaffolds

“…Recently, concerns with the microfracture procedure itself have centred on the detrimental effects on subchondral bone restoration from drilling/puncturing into the subchondral bone [ 79 , 80 ]. Translational models of bone marrow stimulation techniques have demonstrated the benefits of using awls, picks and wires of small-diameter over larger instruments [ [81] , [82] , [83] , [84] ]. In this study we employed the use of two different k-wires to perforate the subchondral bone.…”

Multiplexed tape-stabilized cryohistology of mineralized large animal specimens

“…Tissues harvested from larger species (e.g., human, cow, pig, goat, sheep, dog, horse) have a greater surface area for tearing and folding to occur, in comparison to rodent and rabbit tissues. Despite the utility of tape-stabilized cryohistology for large animal samples, only a few studies have applied this method to section such tissues [13][14][15].…”

“…NOTE: Heat at 60°C to dissolve. Narrow Excitation) Animals 9 Yucatan minipig bones were collected from a prior IACUC-approved study at the University of Pennsylvania [14]. The animals (12-month-old castrated male minipigs) received intravenous injections of the mineral labels xylenol orange (4 weeks before euthanasia) and calcein green (2 weeks before euthanasia).…”

“…A single tape-stabilized cryosection can be repeatedly stained, imaged, and washed, permitting the overlay of images from multiple staining rounds, as previously established with rodent tissues [ 3 ]. The overlaid images provide insight into the co-localization (or lack thereof) of deposited minerals, aligned collagen, matrix elements, and specific cell populations [ 6 ], which can be quantified [ 3 , 4 , 14 , 15 ].…”

“…The established workflow is validated using vertebral body, femoral head, and temporomandibular joint samples from adult Yucatan minipigs. Prior to euthanasia, these animals received intravenous injections of xylenol orange and calcein green bone fluorochrome labels, administered two weeks apart [ 14 ]. The developed protocol enables the overlay of the incorporated mineral labels, aligned collagen (polarized light), tartrate resistant phosphatase (TRAP), alkaline phosphatase (AP), and toluidine blue staining on a single histological section.…”