Post Mortem Leukocyte Scintigraphy in Juvenile Pigs with Experimentally Induced Osteomyelitis

We have previously demonstrated that 111In-labeled autologous leukocyte scintigraphy is able to detect osteomyelitis in living juvenile pigs. In animal research studies, it may well be an advantage if the animals could be scanned after euthanasia. Applying traditional scanning of living animals to euthanized animals will render anaesthesia unnecessary and be ideal for obtaining good and reliable scans for the correct interpretation of imaging afterwards, since the animals do not move. The autologous leukocytes of the pigs were collected, marked with 111In, and reinjected into the pigs and allowed for homing to the site of infections as usual while the pigs were alive. In this study, we demonstrate that it is possible to perform SPECT/CT with 111In-labelled autologous leukocytes almost 24 hrs after euthanasia with the same detectability of osteomyelitic lesions as in living pigs (78% versus 79%). The pigs in this study had exactly the same experimental conditions as the living pigs and were examined in parallel with the living pigs except for euthanasia prior to the leukocyte scan and that no PET/CT scans were performed

Attempts to Target <i>Staphylococcus aureus</i> Induced Osteomyelitis Bone Lesions in a Juvenile Pig Model by Using Radiotracers

Background [18F]FDG Positron Emission Tomography cannot differentiate between sterile inflammation and infection. Therefore, we, aimed to develop more specific radiotracers fitted for differentiation between sterile and septic infection to improve the diagnostic accuracy. Consequently, the clinicians can refine the treatment of, for example, prosthesis-related infection. Methods: We examined different target points; Staphylococcus aureus biofilm (68Ga-labeled DOTA-K-A9 and DOTA-GSGK-A11), bone remodeling ([18F]NaF), bacterial cell membranes ([68Ga]Ga-Ubiquicidin), and leukocyte trafficking ([68Ga]Ga-DOTA-Siglec-9). We compared them to the well-known glucose metabolism marker [18F]FDG, in a well-established juvenile S. aureus induced osteomyelitis (OM) pig model. Results: [18F]FDG accumulated in the OM lesions seven days after bacterial inoculation, but disappointingly we were not able to identify any tracer accumulation in OM with any of the supposedly more specific tracers. Conclusion: These negative results are, however, relevant to report as they may save other research groups from conducting the same animal experiments and provide a platform for developing and evaluating other new potential tracers or protocol instead

Preclinical Testing of Radiopharmaceuticals for the Detection and Characterization of Osteomyelitis:Experiences from a Porcine Model

The development of new and better radioactive tracers capable of detecting and characterizing osteomyelitis is an ongoing process, mainly because available tracers lack selectivity towards osteomyelitis. An integrated part of developing new tracers is the performance of in vivo tests using appropriate animal models. The available animal models for osteomyelitis are also far from ideal. Therefore, developing improved animal osteomyelitis models is as important as developing new radioactive tracers. We recently published a review on radioactive tracers. In this review, we only present and discuss osteomyelitis models. Three ethical aspects (3R) are essential when exposing experimental animals to infections. Thus, we should perform experiments in vitro rather than in vivo (Replacement), use as few animals as possible (Reduction), and impose as little pain on the animal as possible (Refinement). The gain for humans should by far exceed the disadvantages for the individual experimental animal. To this end, the translational value of animal experiments is crucial. We therefore need a robust and well-characterized animal model to evaluate new osteomyelitis tracers to be sure that unpredicted variation in the animal model does not lead to a misinterpretation of the tracer behavior. In this review, we focus on how the development of radioactive tracers relies heavily on the selection of a reliable animal model, and we base the discussions on our own experience with a porcine model

Lymph Nodes Draining Infections Investigated by PET and Immunohistochemistry in a Juvenile Porcine Model

Background: [(18)F]FDG and [(11)C]methionine accumulate in lymph nodes draining S. aureus -infected foci. The lymph nodes were characterized by weight, [(11)C]methionine- and [(18)F]FDG-positron emissions tomography (PET)/computed tomography (CT), and immunohistochemical (IHC)-staining. Methods: 20 pigs inoculated with S. aureus into the right femoral artery were PET/CT-scanned with [(18)F]FDG, and nine of the pigs were additionally scanned with [(11)C]methionine. Mammary, medial iliac, and popliteal lymph nodes from the left and right hind limbs were weighed. IHC-staining for calculations of area fractions of Ki-67, L1, and IL-8 positive cells was done in mammary and popliteal lymph nodes from the nine pigs. Results: The pigs developed one to six osteomyelitis foci. Some pigs developed contiguous infections of peri-osseous tissue and inoculation-site abscesses. Weights of mammary and medial iliac lymph nodes and their [(18)F]FDG maximum Standardized Uptake Values (SUV(FDGmax)) showed a significant increase in the inoculated limb compared to the left limb. Popliteal lymph node weight and their FDG uptake did not differ significantly between hind limbs. Area fractions of Ki-67 and IL-8 in the right mammary lymph nodes and SUV(Metmax) in the right popliteal lymph nodes were significantly increased compared with the left side. Conclusion: The PET-tracers [(18)F]FDG and [(11)C]methionine, and the IHC- markers Ki-67 and IL-8, but not L1, showed increased values in lymph nodes draining soft tissues infected with S. aureus. The increase in [(11)C]methionine may indicate a more acute lymph node response, whereas an increase in [(18)F]FDG may indicate a more chronic response

Refinement of a hematogenous localized osteomyelitis model in pigs

We have previously developed a model of localized osteomyelitis by injecting Staphylococcus aureus (S. aureus) unilaterally into the femoral artery of juvenile domestic pigs (Johansen et al., 2012; Nielsen et al., 2015). We used this model for the evaluation of bone-infection tracers applicable for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) (Nielsen et al., 2015). However, several of the 40 kg pigs were euthanized prior to PET and SPECT scanning due to lameness, shallow respiration, fever and anorexia; the last three clinical signs indicating dissemination of S. aureus to the lungs and other internal organs. We therefore decided to refine our model in order to improve the success rate. We speculated that younger pigs might respond differently to inoculations. A total of ten female domestic pigs were included in our study, three with a body weight of 40 kg