Measurement of bone blood flow using the hydrogen washout technique—part II: Validation by comparison to microsphere entrapment

Abstract

Accurate and reproducible measurement of bone blood flow has important clinical and experimental applications. Hydrogen washout is simple, safe, and widely used, but its use in bone tissue has not been validated. To this end, we have compared cortical bone blood flow measurements obtained by radioactive-labeled microsphere entrapment with those from hydrogen washout. Blood flow was measured in tibial cortical bone of 12 New Zealand White rabbits by radioactive microsphere entrapment and by hydrogen washout. Besides a control group (n ¼ 6), four animals were treated with systemic epinephrine (0.8 mg/kg/min) (group 2) and two with nitroprusside (100 mg/kg/min) (group 3). Furthermore, nine femora from seven rats were isolated on their vascular pedicles and repeated bone blood flow measurements were made at each location with the hydrogen washout method to confirm reproducibility of blood flow determinations by hydrogen washout. An average flow of 2.3 AE 2.0 mL/min/100 g was obtained with the microsphere method and 2.0 AE 0.5 mL/min/100 g with the hydrogen washout method. There was a significant correlation and agreement: R 2 ¼ 0.97 (p < 0.01). No consistent flow variations were found with systemic vasoactive drug administration. Hydrogen washout provided reproducible results and showed high sensitivity to flow changes. Hydrogen washout is both sensitive and reproducible in measuring bone blood flow. Results agree well with flow values obtained by labeled microsphere entrapment. ß

Evaluation of early cellular influences of bone morphogenetic proteins 12 and 2 on equine superficial digital flexor tenocytes and bone marrow–derived mesenchymal stem cells in vitro

“…Previous research in rats indicates that implants containing BMP12 at concentrations 20 times as high as necessary for bone induction with an osteogenic BMP did not induce bone formation; rather, the implants resulted in development of tendon with neonatal characteristics. 27 …”

“…The BMDMSCs were obtained via bone marrow aspiration from the sternum 27,41–43 from an additional 3 adult (age range, 10 to 18 years) clinically normal horses of various breeds. The bone marrow–aspiration procedure was approved by The Ohio State University Institutional Animal Use and Care Committee and was similar to that in previous studies.…”

“…24–26 Recombinant human BMP2 is well characterized and is the most studied BMP with potent osteoinductive capacity and ability to induce mineralization of BMDMSCs 24 in vivo and in vitro; furthermore, this has been determined in many species, including horses. 25,27 Bone morphogenetic protein 12, a human homologue of murine growth and differentiation factor-7, is in the BMP family and is related to other BMPs involved in the developmental processes of the musculoskeletal system, 24 including regulating tissue differentiation, 28,29 tendon healing, and tenogenesis. 30 Unlike other BMPs, however, BMP12 does not have an obvious osteoinduction effect on tendon cells 29–35 and is associated with accelerated healing and improved biomechanical quality of repairs in human patellar tendons, 32 tendon laceration models in chickens and rats, 31,35,36 gastrocnemius tendon models in rats and mice, 33,37 and periodontal ligaments in dogs.…”

“…39 Additionally, BMP12 could be delivered by gene therapy to a specific site, allowing for local expression and sustained release. 25,27,40 …”

Living Bone Allotransplants Survive by Surgical Angiogenesis Alone: Development of a Novel Method of Composite Tissue Allotransplantation

“…A patent vessel was observed to have both visible gross pulsation and a positive standard microsurgical patency test. Cortical blood flow was determined with use of the modified hydrogen washout method that we have previously described and validated 14,15 . Briefly, a shallow hole was drilled perpendicularly into the cortical bone.…”