Saturday, February 6, 2016

Domestic Dog - Osteoporosis Research

The evolutionary origin of the domestic dog (Canis lupus familiaris) is not well understood. DNA evidence suggests that the domestic dog diverged from its closest relative, the grey wolf (Canis lupus), sometime between 11,000 and 16,000 years ago, although a recent report pushes this date back to at least 27,000 years ago, which is in closer accord to the archaeological evidence that suggests a divergence around 36,000 years ago. And of course, once we humans assert our dominance over another species, we begin to tinker with it, in order to improve it. Today, the World Canine Organization (Federation Cynologique Internationale) recognizes over 340 breeds of domestic dog.

The most common breed of dog used in biomedical research is the beagle, and our laboratory is no exception. The Organ Laboratory does not take animal research lightly. Like every other laboratory at Indiana University School of Medicine, before we begin a study we are required to submit an animal protocol for review by the Institutional Animal Care and Use Committee (IACUC). The IACUC is comprised of university leaders, other researchers, veterinarians, and members of the general public. Every single procedure we wish to perform in an experiment must be approved by the IACUC prior to the initiation of the experiment. THIS. IS. CRITICAL. It is critical because animals, unlike people, are not able to give their informed consent to a procedure. Therefore, the IACUC assures that these animals will experience the least amount of discomfort and stress during any procedure. In fact, it is not uncommon to have more difficulty getting an animal study approved than it is to get a study approved for humans. Again, this is because humans can refuse to participate in a study, whereas a laboratory animal cannot. Because we study the mechanical properties of bone and muscle in a biomedical context, our work could not proceed without the use of vertebrate animals. Typically, our studies are carried out using laboratory rodents such as mice and rats because a whole lot is known about the genetic makeup of these animals, and we can control more variables because we can measure them with species-specific tools like biomarkers and DNA. Unfortunately, and perhaps due in part to size constraints, mouse and rat bone are inferior models of human bone. Why is that? Allow us to digress for just a moment…

Bones are dynamic tissues, and they are constantly remodeling out the numerous micro-cracks that materialize from normal wear and tear. This remodeling occurs when osteoclast cells resorb the bone matrix and osteoblast cells synthesize and lay down new bone where the old bone was removed. There are multiple places (envelopes) on any given bone where this type of activity occurs. One of the most important areas in human bone is the cortical shell surrounding the marrow cavity. Human cortical bone experiences a lot of intra-cortical remodeling, activity within the middle of the cortex.  Contrast this with rats and mice which typically only remodel on the inner and outer surfaces of cortical bone. Therefore, rodents make a poor research model if the question to be answered focuses on intracortical remodeling. Beagles, and dogs in general, do experience intracortical remodeling, and thus they are suitable animal models of human cortical bone.

The ultimate goal of the research described here utilizing a dog model is to improve treatment of osteoporosis by treating the underlying mechanisms that lead to fracture. Our most recent studies have investigated whether combining different types of anti-osteoporosis drugs might be more beneficial than using either one of the drugs alone. In particular, we combined two different classes of anti-catabolic drugs, meaning that these drugs prevent the loss of bone. There are two different classes of anti-catabolic agents used to treat metabolic bone diseases: bisphosphonates and selective estrogen receptor modulators (SERMs). Through our experiments, we have made several significant findings. Below, we highlight two of them.

First, we showed that 6 months of daily raloxifene treatment improves bone material mechanical properties as measured with reference point indentation (RPI). RPI is a novel tool used to assess mechanical properties of bone in vivo and has shown some ability to distinguish between fracture and non-fracture patients when used clinically. We were very excited to discover that raloxifene was effective in enhancing skeletal mechanical properties because we know the drug has little effect on bone mineral density. Subsequent studies utilized the same set of dogs to demonstrate that raloxifene increases cortical bone matrix-bound water, essentially increasing the hydration of the bone, and therefore increasing its material toughness. We hypothesize that this altered hydration is related to the known anti-fracture efficacy of raloxifene. Voila! A mechanism of action!

Some of the skulls in our outreach collection.
Once The Eatles are finished cleaning this skull, we will add it to our growing collection of animal skulls that we use for outreach programming. Over the years we have developed a number of initiatives that bring skeletal biology and functional anatomy to the classroom, and we can aim our programs for any grade level from preschool to high school. If you are in central Indiana and are interested in having one of us visit your classroom, please get in touch on twitter: @eatlemania

Contributed by: Neil Jain, Kelly Biro, and Jason Organ, PhD


Allen MR, Territo PR, Lin C, Persohn S, Jiang L, Riley AA, McCarthy BP, Newman CL, Burr DB, & Hutchins GD (2015). In Vivo UTE-MRI Reveals Positive Effects of Raloxifene on Skeletal-Bound Water in Skeletally Mature Beagle Dogs. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research, 30 (8), 1441-4 PMID: 25644867

Allen MR, McNerny EM, Organ JM, & Wallace JM (2015). True Gold or Pyrite: A Review of Reference Point Indentation for Assessing Bone Mechanical Properties In Vivo. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research, 30 (9), 1539-50 PMID: 26235703 

Aref M, Gallant MA, Organ JM, Wallace JM, Newman CL, Burr DB, Brown DM, & Allen MR (2013). In vivo reference point indentation reveals positive effects of raloxifene on mechanical properties following 6 months of treatment in skeletally mature beagle dogs. Bone, 56 (2), 449-53 PMID: 23871851

Skoglund P, Ersmark E, Palkopoulou E, & Dalén L (2015). Ancient wolf genome reveals an early divergence of domestic dog ancestors and admixture into high-latitude breeds. Current biology : CB, 25 (11), 1515-9 PMID: 26004765