Differential diagnosis of an unusual shoulder articular lesion in an ancient domestic dog (Canis lupus familiaris L., 1758).

A proximal humeral articular surface from an ancient domestic dog deliberate burial was examined during survey of small mammal bones from a prehistoric early Late Woodland archeological site. An unusual lesion on the caudolateral articular surface prompted micro-computed tomography to define detailed structure. Results indicate cortical or immature woven bone arising subchondrally, replacing normal trabeculae, extending through a breach in the cortical surface, and having sharp transition with surrounding normal bone. Organized bone within the lesion indicates that the dog lived for months-to-years following insult. Differential diagnoses initially included: sharp penetrating trauma; intrinsic or extrinsic blunt fracturing force; osteochondrosis or complication of an osteochondral lesion; unusual osteoarthritis; and neoplasia. Computed tomography ruled out normal or unusual osteoarthritis, and neoplasia. The nature and small size of the lesion, relatively small size of the dog, and lack of evidence for complicating infection, suggest against sharp penetrating trauma as a sole cause. The most plausible differential diagnoses include: uncommon fracture-producing force in a companion animal, and blunt intrinsic or extrinsic force causing fracture at a weak point, such as an early osteochondral lesion, that was obliterated by healing. Combined gross examination, micro-computed tomography, and archeological-anthropological influences facilitated refinement of differential diagnosis.

Cathepsin D is partly endocytosed by the LRP1 receptor and inhibits LRP1-regulated intramembrane proteolysis.

The aspartic protease cathepsin-D (cath-D) is a marker of poor prognosis in breast cancer that is overexpressed and hypersecreted by human breast cancer cells. Secreted pro-cath-D binds to the extracellular domain of the ß-chain of the LDL receptor-related protein-1 (LRP1) in fibroblasts. The LRP1 receptor has an 85-kDa transmembrane ß-chain and a noncovalently attached 515-kDa extracellular α-chain. LRP1 acts by (1) internalizing many ligands via its α-chain, (2) activating signaling pathways by phosphorylating the LRP1ß-chain tyrosine and (3) modulating gene transcription by regulated intramembrane proteolysis (RIP) of its ß-chain. LRP1 RIP involves two cleavages: the first liberates the LRP1 ectodomain to give a membrane-associated form, LRP1ß-CTF, and the second generates the LRP1ß-intracellular domain, LRP1ß-ICD, that modulates gene transcription. Here, we investigated the endocytosis of pro-cath-D by LRP1 and the effect of pro-cath-D/LRP1ß interaction on LRP1ß tyrosine phosphorylation and/or LRP1ß RIP. Our results indicate that pro-cath-D was partially endocytosed by LRP1 in fibroblasts. However, pro-cath-D and ectopic cath-D did not stimulate phosphorylation of the LRP1ß-chain tyrosine. Interestingly, ectopic cath-D and its catalytically inactive (D231N)cath-D, and pro-(D231N)cath-D all significantly inhibited LRP1 RIP by preventing LRP1ß-CTF production. Thus, cath-D inhibits LRP1 RIP independently of its catalytic activity by blocking the first cleavage. As cath-D triggers fibroblast outgrowth by LRP1, we propose that cath-D modulates the growth of fibroblasts by inhibiting LRP1 RIP in the breast tumor microenvironment.

Distraction index as a risk factor for osteoarthritis associated with hip dysplasia in four large dog breeds.

OBJECTIVE: To determine if age, breed, gender, weight or distraction index (DI) influenced the risk of radiographic osteoarthritis (OA) of canine hip dysplasia (CHD) in four common dog breeds; the American bulldog, Bernese mountain dog, Newfoundland and standard poodle. MATERIALS AND METHODS: This was a cross sectional prevalence study with 4349 dogs. Canine hips were evaluated using 3 radiographic projections: the hip-extended view, the compression view and the distraction view. The hip-extended view was examined for the presence of OA. The PennHIP distraction view was utilized to calculate the DI. For all breeds, a multiple logistic regression model incorporating age, weight, gender, and DI was created. For each breed, disease-susceptibility curves grouping dogs on the basis of age were constructed. Receiver-operating characteristic (ROC) curves were developed for each breed regardless of age. RESULTS: For all breeds, DI was the most significant risk factor for the development of OA associated with CHD. Weight and age were also significant risk factors in all four breeds, but gender was not. CLINICAL SIGNIFICANCE: Results from this study support previous findings, that irrespective of breed, the probability of radiographic OA increases with hip joint laxity as measured by the DI. Breed-specific differences in this relationship, however, warrant investigation of all breeds affected by CHD to determine inherent dependency of hip OA on joint laxity. Such findings guide veterinarians in helping dog breeders to make evidence-based breeding decisions and in informing dog owners to implement preventative treatments for CHD for dogs found to be at risk.

Imidazo[5,1-f][1,2,4]triazin-2-amines as novel inhibitors of polo-like kinase 1.

The synthesis and biological activities of imidazo[5,1-f][1,2,4]triazin-2-amines (imidazotriazines) as novel polo-like kinase 1 inhibitors are reported.

The history of spina bifida, hydrocephalus, paraplegia, and incontinence.

This paper covers the history since ancient times of spina bifida (SB) and its main associated conditions, viz., hydrocephalus, paraplegia and incontinence. Particular stress has been placed on the ancient authors who recognised these conditions. The article concludes with the history of some general aspects of SB and the dilemmas in its treatment.

Muscular dystrophy in female dogs.

The most common form of muscular dystrophy in dogs and humans is caused by mutations in the dystrophin gene. The dystrophin gene is located on the X chromosome, and, therefore, disease-causing mutations in dystrophin occur most often in males. Therefore, females with dystrophin deficiency or other forms of muscular dystrophy may be undiagnosed or misdiagnosed. Immunohistochemistry was used to analyze dystrophin and a number of other muscle proteins associated with muscular dystrophy in humans, including sarcoglycans and laminin alpha2, in muscle biopsy specimens from 5 female dogs with pathologic changes consistent with muscular dystrophy. The female dogs were presented with a variety of clinical signs including generalized weakness, muscle wasting, tremors, exercise intolerance, gait abnormalities, and limb deformity. Serum creatine kinase activity was variably high. One dog had no detectable dystrophin in the muscle; another was mosaic, with some fibers normal and others partly dystrophin-deficient. A 3rd dog had normal dystrophin but no detectable laminin alpha2. Two dogs could not be classified. This study demonstrates the occurrence of dystrophin- and laminin alpha2-associated muscular dystrophy and the difficulty in clinical diagnosis of these disorders in female dogs.

Pullout properties of 3.5-mm AO/ASIF self-tapping and cortex screws in a uniform synthetic material and in canine bone.

OBJECTIVES--To compare the pullout properties of 3.5-mm AO/ASIF self-tapping screws (STS) to corresponding standard cortex screws (CS) in a uniform synthetic test material and in canine femoral bone. The influence of screw-insertion technique, test material, and test-material thickness were also assessed. STUDY DESIGN--In vitro experimental study. SAMPLE POPULATION--Two independent studies: a uniform synthetic test material and paired femurs from mature dogs. METHODS-Mechanical testing was performed in accordance with standards established by the American Society for Testing and Materials for determination of axial pullout strength of medical bone screws. Completely inserted STS, completely inserted CS, and incompletely inserted STS were tested in 3 groups of 10 test specimens each in 4.96-mm and 6.8-mm thick sheets of synthetic material. In the bone study, group 1 consisted of 24 completely inserted STS compared with 24 completely inserted CS, and group 2 consisted of 24 incompletely inserted STS versus 24 completely inserted CS. Comparisons were made between paired femurs at corresponding insertion sites. Pullout data were normalized, thereby eliminating the effect of test-material thickness on pullout properties. Mean values were compared using 2-way ANOVA. Statistical significance was set at P <.05. RESULTS--In both the 4.96-mm and 6.8-mm synthetic material, pullout testing of the completely inserted STS demonstrated significantly greater yield strength and ultimate strength than completely inserted CS. There was no significant difference between incompletely inserted STS and completely inserted STS. The 6.8-mm test material significantly increased yield strength and ultimate strength for all test groups compared with the 4.96-mm test material. In canine bone, there was no significant difference in yield strength of completely inserted STS and completely inserted CS. Yield strength of completely inserted STS and completely inserted CS were significantly greater than incompletely inserted STS. CONCLUSIONS--Pullout properties of completely inserted STS were significantly greater than corresponding CS in a uniform test material. In canine bone, the pullout strength of STS and CS were not different. Incomplete STS insertion resulted in an 18% reduction in holding power as compared with completely inserted CS and STS in canine bone. CLINICAL RELEVANCE--The length of STS used in canine bone should be such that the cutting flutes extend beyond the trans cortex to maximize pullout strength.

Evaluation of risk factors for degenerative joint disease associated with hip dysplasia in German Shepherd Dogs, Golden Retrievers, Labrador Retrievers, and Rottweilers.

OBJECTIVE: To determine whether age, breed, sex, weight, or distraction index (DI) was associated with the risk that dogs of 4 common breeds (German Shepherd Dog, Golden Retriever, Labrador Retriever, Rottweiler) would have radiographic evidence of degenerative joint disease (DJD) associated with hip dysplasia. DESIGN: Cross-sectional prevalence study. ANIMALS: 15,742 dogs. PROCEDURE: Hips of dogs were evaluated radiographically by use of the ventrodorsal hip-extended view, the compression v ew, and the distraction view. The ventrodorsal hip-extended view was examined to determine whether dogs had DJD. For each breed, a multiple logistic regression model incorporating age, sex, weight, and DI was created. For each breed, disease-susceptibility curves were produced, using all dogs, regardless of age, and dogs grouped on the basis of age. RESULTS: Weight and DI were significant risk factors for DJD in all breeds. For German Shepherd Dogs, the risk of having DJD was 4.95 times the risk for dogs of the other 3 breeds combined. In all breeds, the probability of having DJD increased with age. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that the probability of having hip DJD increased with hip joint laxity as measured by use of DI. This association was breed-specific, indicating that breed-specific information on disease susceptibility should be incorporated when making breeding decisions and when deciding on possible surgical treatment of hip dysplasia.

Evaluation of the effect of limited food consumption on radiographic evidence of osteoarthritis in dogs.

OBJECTIVE: To determine prevalence of radiographic evidence of osteoarthritis in 4 diarthrodial joints of dogs with restricted feed intake, compared with dogs without restricted feed intake. DESIGN: Paired feeding study. ANIMALS: 48 Labrador Retrievers. PROCEDURE: Dogs in litters from 7 dams and 2 sires were paired by sex and weight within litters and randomly assigned to a control-fed group or a limit-fed group that received 25% less food than the control-fed group. Radiographic evaluation of prevalence and severity of osteoarthritis in the hip, shoulder, elbow, and stifle joints was performed when dogs were 8 years of age. RESULTS: Radiographic evidence of osteoarthritis that affected multiple joints was significantly more common in the control-fed group than in the limit-fed group. Prevalence of lesions in the hip joint was 15/22 in the control-fed group and 3/21 in the limit-fed group. Prevalence of lesions in the shoulder joint was 19/22 in the control-fed group and 12/21 in the limit-fed group; lesions in this joint were generally mild. Severity, but not prevalence, of osteoarthritis in the elbow joint was greater in the control-fed group than in the limit-fed group. CONCLUSIONS AND CLINICAL RELEVANCE: Prevalence and severity of osteoarthritis in several joints was less in dogs with long-term reduced food intake, compared with control dogs. Food intake is an environmental factor that may have a profound effect on development of osteoarthritis in dogs.

Effect of public symphysiodesis on pelvic development in the skeletally immature greyhound.

OBJECTIVE: To evaluate the effect of pubic symphysiodesis (PS) on pelvic development in skeletally immature dogs. STUDY DESIGN: Prospective randomized clinical trial. ANIMALS: Eight 4 month-old, sexually intact female Greyhounds. METHODS: Initial PS was performed at 4 months of age using a powered stapling device. Because of failure of the initial surgery, a second PS was performed 1 month later by resecting the pubic symphysis with a rongeur followed by placement of handmade bone staples in four dogs. Sham PS was performed in four control dogs at 4 months of age. Pubic growth rate and pelvic development were evaluated using standard plane radiography and computed tomography. Specific measurements included acetabular ventroversion, Norberg angle, lateral center-edge angle, and pelvic inlet dimensions. Hip distraction indices were determined as well. RESULTS: PS at 4 months of age using a stapling device failed. Pubic symphysiodesis using hand made staples was successful at 5 months of age and did not result in any clinically significant intraoperative or postoperative complications. Pubic symphysiodesis markedly decreased pubic symphysis growth in the treatment group. Hip distraction indices and pelvic inlet circumference, area, and width significantly decreased in treated dogs compared to those in the control group. Acetabular ventroversion was significantly increased in treated dogs compared to those in the control group. CONCLUSIONS: PS decreases pelvic canal size, increases acetabular ventroversion, and does not appear to have any clinically significant complications. CLINICAL RELEVANCE: PS performed in skeletally immature dogs with hip dysplasia may provide an effect similar to a triple pelvic osteotomy and warrants further investigation.

Evaluation of the association between medial patellar luxation and hip dysplasia in cats.

OBJECTIVE: To investigate the association between hip dysplasia (HD) and medial patellar luxation (MPL) in cats. DESIGN: Cross-sectional prevalence study. ANIMALS: 78 cats. PROCEDURE: A complete history was obtained. Cats were examined to detect MPL and HD. Radiographs of the stifle and hip joints were obtained. Hip joints were evaluated by use of Norberg angle, distraction index, and scoring consistent with that established by the Orthopedic Foundation for Animals. RESULTS: There were 43 male and 35 female cats mean age, 2.5 years). Eleven cats had clinical signs of disease in the pelvic limbs. Medial subluxation of the patella (subgrade 1) was seen in 31 of 33 cats with otherwise normal stifle joints. Medial patellar luxation was found in 45 of 78 (58%) cats, and 35 of 45 (78%) had grade-1 MPL. Bilateral MPL was seen in 32 of 45 (71%) cats. A weak association existed between MPL and HD, because cats were 3 times more likely to have HD and patellar luxation than to have either condition alone. Concurrent MPL and HD were detected in 19 of 78 (24%) cats, and HD was diagnosed radiographically in 25 of 78 (32%) cats (19 mild, 4 moderate, 2 severe). Eighteen of the 25 cats with HD had bilateral HD. CONCLUSIONS AND CLINICAL RELEVANCE: Clinically normal cats may have a certain degree of laxity in the stifle joint, evident as medial patellar subluxation (< grade 1). There is a weak association between MPL and HD, and both conditions may develop, alone or in combination, more frequently than has been reported.

Relationships between results of the Ortolani method of hip joint palpation and distraction index, Norberg angle, and hip score in dogs.

OBJECTIVE: To determine whether results of the Ortolani method of hip joint palpation in dogs were related to distraction index (DI), Norberg angle, or radiographic hip score. DESIGN: Cross-sectional study. ANIMALS: 459 clinically normal dogs. PROCEDURE: Dogs were sedated for radiography and palpation of the hip joints. Results of hip joint palpation were classified as negative, mild positive, moderate positive, or severe positive. Distraction indices were measured for all dogs. Norberg angles were measured for 380 dogs for which ventrodorsal hip-extended radiographic projections were available. Hip scores assigned by the Orthopedic Foundation for Animals (OFA) were available for 95 dogs. RESULTS: Age, weight, and sex were not significantly associated with results of hip joint palpation. There was moderate correlation between results of hip joint palpation and DI (r = 0.636), low-moderate correlation between results of hip joint palpation and OFA hip scores (rs = 0.437), and weak negative correlation (r = -0.236) between results of hip joint palpation and Norberg angle. For joints without degenerative joint disease (DJD), there was a significant linear relationship between results of hip palpation and DI; however, for joint with DJD, there was not. Results of hip joint palpation were 5.3-fold as likely to be negative for dogs with DJD as for dogs without. CLINICAL IMPLICATIONS: Results of hip joint palpation were at best moderately correlated with radiographic measures of hip joint laxity. Therefore, hip joint palpation should be combined with hip-extended and stress radiography when assessing hip joint quality.

Antibody-directed enzyme prodrug therapy with the T268G mutant of human carboxypeptidase A1: in vitro and in vivo studies with prodrugs of methotrexate and the thymidylate synthase inhibitors GW1031 and GW1843.

Antibody-directed enzyme prodrug therapy (ADEPT) is a technique to increase antitumor selectivity in cancer chemotherapy. Our approach to this technology has been to design a mutant of human carboxypeptidase A (hCPA1-T268G) which is capable of hydrolyzing in vivo stable prodrugs of MTX and targeting this enzyme to tumors on an Ep-CAM1-specific antibody, ING1. Through the use of this >99% human enzyme which is capable of catalyzing a completely nonhuman reaction, we hope to increase ADEPT selectivity while decreasing overall immunogenicity of the enzyme-antibody conjugate. In the current report, prodrugs of the thymidylate synthase inhibitors GW1031 and GW1843 and the dihydrofolate reductase inhibitor methotrexate were studied for their wild-type and mutant hCPA enzyme hydrolysis, their in vivo stability, and their use in therapy. Prodrugs with high kcat/Km ratios for mutated versus wild-type hCPA1 were examined in vitro for their stability in human pancreatic juice, and in vivo for their stability in mouse plasma and tissues. In addition, targeting and in vivo enzyme activity studies were performed with an ING1 antibody conjugate of the mutant enzyme (ING1-hCPA1-T268G). Finally, in vivo therapy studies were performed with LS174T tumors to demonstrate proof of principle. Results indicate that prodrugs can be synthesized that are selective and efficient substrates of hCPA1-T268G and not substrates of the endogenous CPA activities; this leads to excellent in vivo stability for these compounds. In vivo conjugate targeting studies showed that the antibody-enzyme conjugate was targeted to the tumor and enzyme was initially active in vivo at the site. Unfortunately therapeutic studies did not demonstrate tumor reduction. Experiments to determine reasons for the lack of antitumor activity showed that the enzyme activity decreased as a result of enzyme instability. The results offer encouragement for additional novel mutant enzyme improvements and additional in vivo studies on this unique approach to ADEPT.

Purification and characterization of a membrane bound neutral pH optimum magnesium-dependent and phosphatidylserine-stimulated sphingomyelinase from rat brain.

Sphingomyelin hydrolysis and ceramide generation catalyzed by sphingomyelinases (SMase) are key components of the signaling pathways in cytokine- and stress-induced cellular responses. In this study, we report the partial purification and characterization of the membrane bound, neutral pH optimal, and magnesium-dependent SMase (N-SMase) from rat brain. Proteins from Triton X-100 extract of brain membrane were purified sequentially with DEAE-Sephacel, heparin-Sepharose, ceramic hydroxyapatite, Mono Q, phenyl-Superose, and Superose 12 column chromatography. After eight purification steps, the specific activity of the enzyme increased by 3030-fold over the brain homogenate. The enzyme hydrolyzed sphingomyelin but not phosphatidylcholine and its activity was dependent upon magnesium with an optimal pH of 7.5 and a native pI of 5.2. Delipidation of the enzyme through chromatographic purification or by extraction with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid followed by gel filtration revealed that the enzyme became increasingly dependent on phosphatidylserine (PS). Up to 20-fold stimulation was observed with PS whereas other lipids examined were either ineffective or only mildly stimulatory. The Km of the enzyme for substrate sphingomyelin (3.4 mol %) was not affected by PS. The highly purified enzyme was inhibited by glutathione with a >95% inhibition observed with 3 mM glutathione and with a Hill number calculated at approximately 8. The significance of these results to the regulation of N-SMase is discussed.

Correlation between longitudinal bone growth, growth hormone, and insulin-like growth factor-I in prepubertal dogs.

OBJECTIVE: To determine the association between longitudinal bone growth and concentrations of growth hormone (GH) and insulin-like growth factor-I (IGF-I) in serum from prepubertal dogs. Animals-6 male 14-week-old German Shepherd Dogs. PROCEDURE: Blood was obtained every 30 minutes for 14 consecutive days. Concentrations of GH and IGF-I in serum were determined, using a canine-specific radioimmunoassay and conventional radioimmunoassay after acid-ethanol extraction, respectively. Simultaneous biplanar radiography was performed daily to measure bone growth. Spectral analysis was used to estimate specific features of GH secretion during an extended period. Multiple linear regression with different lag times between independent and dependent variables was used to determine the strongest predictors of bone growth. RESULTS: The power spectra of GH concentrations in serum had a primary peak at a frequency of 0.02 cycles/h or a periodicity of 50 h/cycle. A significant determinant of longitudinal bone growth was a lag time of 1 day in concentration of GH in serum. The relationship between IGF-I concentration in serum and bone growth was not significant. CONCLUSIONS: The primary frequency of GH secretion is outside the time frame of a single day and the concentration of GH in serum is a primary determinant of bone growth. CLINICAL RELEVANCE: A better understanding of the components of bone growth provide discernment to improved diagnosis and treatment of abnormal bone growth.