Ida Elisabeth Holm

Object recognition as a measure of memory in 1-2 years old transgenic minipigs carrying the APPsw mutation for Alzheimer's disease.

Alzheimer’s disease (AD) is a disabling, fatal disease, where animal models potentially can enable investigation of aetiology and treatment. The first litter of Göttingen minipigs carrying a mutation for human AD was born in 2007, showing transgene expression. In human AD patients, memory impairment is the most striking and consistent feature. The aim of the present study was to examine effects of the APPsw transgene on memory of AD minipigs compared with non-transgenic controls at two ages (1–2xa0years) using the spontaneous object recognition test (SORT), which is based on behavioural discrimination of familiar and novel objects. No significant difference between AD minipigs and controls was found when comparing object recognition as a measure of memory. The minipigs did explore the novel object significantly more than the familiar, indicating the expected recognition of the familiar object. Two different inter-phase intervals were used (IPI: 10–40xa0min). For both ages, object recognition was evident using 10xa0min IPI. When using 40xa0min IPI, object recognition was evident only at age 1xa0year. Comparing memory of a relatively small group of AD minipigs and controls at two rather young ages using the SORT, we were not able to show memory impairment in APPsw carrying minipigs. Being an age-dependent disease, the transgene is expected to cause AD-like symptoms in this porcine model, and the SORT should be repeated at older ages.

Protein Replacement Therapy Partially Corrects the Cholesterol-Storage Phenotype in a Mouse Model of Niemann-Pick Type C2 Disease

Niemann-Pick type C2 (NPC2) disease is a fatal autosomal recessive neurovisceral degenerative disorder characterized by late endosomal-lysosomal sequestration of low-density lipoprotein derived cholesterol. The breach in intracellular cholesterol homeostasis is caused by deficiency of functional NPC2, a soluble sterol binding protein targeted to the lysosomes by binding the mannose-6-phosphate receptor. As currently there is no effective treatment for the disorder, we have investigated the efficacy of NPC2 replacement therapy in a murine gene-trap model of NPC2-disease generated on the 129P2/OlaHsd genetic background. NPC2 was purified from bovine milk and its functional competence assured in NPC2-deficient fibroblasts using the specific cholesterol fluorescent probe filipin. For evaluation of phenotype correction in vivo, three-week-old NPC2 −/− mice received two weekly intravenous injections of 5 mg/kg NPC2 until trial termination 66 days later. Whereas the saline treated NPC2 −/− mice exhibited massive visceral cholesterol storage as compared to their wild-type littermates, administration of NPC2 caused a marked reduction in cholesterol build up. The histological findings, indicating an amelioration of the disease pathology in liver, spleen, and lungs, corroborated the biochemical results. Little or no difference in the overall cholesterol levels was observed in the kidneys, blood, cerebral cortex and hippocampus when comparing NPC2 −/− and wild type mice. However, cerebellum cholesterol was increased about two fold in NPC2 −/− mice compared with wild-type littermates. Weight gain performance was slightly improved as a result of the NPC2 treatment but significant motor coordination deficits were still observed. Accordingly, ultrastructural cerebellar abnormalities were detected in both saline treated and NPC2 treated NPC2 −/− animals 87 days post partum. Our data indicate that protein replacement may be a beneficial therapeutic approach in the treatment of the visceral manifestations in NPC2 disease and further suggest that neurodegeneration is not secondary to visceral dysfunction.

Changes in lipoxygenase products from synovial fluid in carragheenan induced arthritis in dogs

A non‐suppurative chronic arthritis was induced in the juvenile dog knee by intra‐articular instillations with Carragheenan. Lipoxygenase products of arachidonic acid were separated from synovial fluid by reversed‐phase high‐performance liquid chromatography (RP‐HPLC). After ten weeks we observed an accumulation of leukotriene B4 (LTB4) in synovial fluid in five out of six experimental knees (0.94 to 5.5 ng/ml), as judged by integrated optical density. Biological activity of LTB4 was confirmed by chemokinesis. LTB4 was not detected in control knees. The 15‐lipoxygenase products, 15‐hydroxyeicosatetraenoic acid (15‐HETE) and 13‐hydroxy‐9, 11‐octadecadienoic acid (13‐HODD), being inhibitors of 5‐lipoxygenase, were found in relatively high levels in the control knees compared to the arthritic knees. The results denote LTB4 as a pro‐inflammatory local mediator during carragheenan‐induced arthritis; possibly, the decreased levels of 15‐HETE and 13‐HODD in the arthritic knees may have a regulatory function, thus facilitating LTB4 generation.

99mTc-DPD uptake in juvenile arthritis: Scintimetry and autoradiography of the knee in dogs

Unilateral arthritis of the knee was induced in mongrel puppies by intraarticular injections of 1% Carragheenan. Bone metabolism was studied by a scintimetric technique on static 99mTc-diphosphonate bone scans every 2nd week during the induction of arthritis for 3 months and monthly in a postarthritic phase of another 3 months. Changes in uptake of radionuclide were present after 2 weeks. The induction phase was characterized by a decreased uptake in the calcification layer of the juxta-articular growth plates and a moderately increased epiphyseal uptake. The postarthritic phase was characterized by normalization of growth plate uptake and a marked increase in epiphyseal uptake. Using contact autoradiography, the epiphyseal uptake was seen mainly in a narrow subchondral and subsynovial bone layer, around bone cysts and osteophytes, whereas central epiphyseal bone was osteopenic with decreased uptake of tracer. The study suggests that the early scintigraphic appearance of juvenile non-suppurative arthritis may be an overall decrease in uptake of 99mTc-diphosphonate due to a depression of growth plate metabolism.

Effect of indomethacin on regulation of juxta-articular bone blood flow during joint tamponade. An experimental study in puppies

Prostaglandins are vasoactive substances which are assumed to play a major role in bone metabolism and bone repair. The purpose of the present study was to investigate the effect of indomethacin on the control of epiphyseal bone blood-flow. By means of simultaneous intra-osseous pressure (IOP) and regional blood flow (RBF) measurements in the distal femoral epiphysis (DFE), aspects of vascular control mechanisms in the distal femoral epiphysis were investigated during knee joint tamponade (50% of mean arterial pressure) before and after administration of indomethacin 7.5 mg/kg. Six dogs aged 3-4 months were investigated in fentanyl anaesthesia. Knee joint tamponade resulted in a significant increase in IOP and calculated venous resistance in the DFE, while no significant changes in regional blood-flow or arterial resistance were encountered. Administration of indomethacin did not affect this reaction. The results suggests that indomethacin 7.5 mg/kg does not influence the regulation of epiphyseal blood-flow during elevation of joint pressure indicating that prostaglandins play only a minor or no role in this regulation.

^sup 99m^Tc-DPD Uptake in Juvenile Hemarthrosis Scintimetry and Autoradiography of the Knee in Dogs

The pathogenesis of subchondral bone lesions and growth plate affection in hemophilic arthropathy was studied in puppies by means of repeated regional 99mTc-diphosphonate scintimetry and contact autoradiography. Unilateral hemarthrosis of the knee was induced by biweekly intraarticular injections of autologous blood for 12 weeks. Hemarthrosis caused an early (2 to 4 weeks) decrease in uptake of 99mTc-diphosphonate in the juxtaarticular growth plates (ratio 0.7) and a delayed (8 to 10 weeks) increase in epiphyseal uptake (ratio 1.5). In a recovery phase after hemarthrosis, growth plate uptake returned to normal, while the epiphyseal uptake remained elevated for 8 to 10 weeks. By contact autoradiography, the growth plate uptake was localized to the calcification layer at the metaphyseal aspect of the growth plates, while the epiphyseal uptake mainly was seen in the thin subchondral and subsynovial bone layer and around osteophytes. The changes in uptake of 99mTc-diphosphonate following hemarthrosis for 3 months were reversible and could be ascribed to the presence of synovial inflammation.

Tissue vitality in septic gonitis: 99mTc-DPD scintimetry in puppies

After a single intraarticular injection of 10(9) Staphylococcus aureus in 12 puppies, septic arthritis developed in all the experimental knees after 48 hours. A considerable variability in scintigraphic appearance was observed. The juxtaarticular growth plates showed either unchanged or slightly decreased uptake except in 1 dog exhibiting a definite increase in tracer uptake. The epiphyseal uptake showed no consistent pattern. The intraarticular pressure of the arthritic joints increased significantly, but was not related to the tracer uptake pattern. We conclude that delayed joint scintigraphy as a single investigation in early septic arthritis does not provide diagnostic information and may be misleading.