Biomedical subjects
Publications and source records attributed to G K Smith.
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.
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.
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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.
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.
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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.
Thymidylate synthase (TS) inhibition causes cell death, and this enzyme is the target for the important chemotherapy regime 5-fluorouracil/leucovorin. GW1843 (1843U89) is a potent and specific folate analog TS inhibitor in clinical development. Because of the importance of TS as a chemotherapy target, we are studying the mechanism of TS inhibition-induced cell death by GW1843. Ceramide is a regulatory lipid generated by the action of sphingomyelinase and is believed to signal apoptosis. The role of the ceramide in apoptotic signaling was studied in Molt-4 human T-cell leukemia cells undergoing cell death after treatment with GW1843. In response to GW1843, Molt-4 cells undergo apoptosis with both acidic pH, Mg2+-independent sphingomyelinase (ASMase) and neutral pH, Mg2+-dependent sphingomyelinase (NSMase) activities elevated as early steps in the initiation of apoptosis before Molt-4 commitment to death. These activities lead to ceramide production with kinetics consistent with a role as an effector molecule signaling the initiation of apoptosis in Molt-4 cells. These changes were found to be independent of caspase 3-like (CPP32/apopain) activity and DNA degradation, but were not separable from membrane blebbing or cell lysis in this cell line. In this report, kinetic evidence is provided for a role of ceramide in initiating GW1843-induced cell death of Molt-4 T-cell leukemia cells.
OBJECTIVE: To determine whether distraction index (DI), a measure of passive hip joint laxity, at 2 months of age was predictive of DI at 4 or 12 months of age in German Shepherd Dogs. DESIGN: Prospective cohort study. ANIMALS: 45 German Shepherd Dogs. PROCEDURE: DI was measured at 2, 4, and 12 months of age. At the same times, a standard ventrodorsal radiographic projection of the pelvis with the hip joints extended was obtained and examined for evidence of degenerative joint disease (DJD). To facilitate radiographic positioning, dogs were sedated or anesthetized. RESULTS: DI at 2 months of age was not significantly correlated with DI at 4 or 12 months of age. However, DI at 4 months of age was correlated with DI at 12 months of age. The proportion of dogs with DI > or = 0.3 at 12 months of age that had radiographic evidence of DJD by 12 months of age (13/22; 59%) was significantly greater than the proportion of dogs with DI < 0.3 at 12 months of age that had radiographic evidence of DJD by 12 months of age (1/9; 11%). CLINICAL IMPLICATIONS: For German Shepherd Dogs, DI at 2 months of age was not sufficiently reliable to predict DI at 4 and 12 months of age; however, DI at 4 and 12 months of age were comparable. We recommend that, for German Shepherd Dogs, DI not be measured before 4 months of age and that particularly for breeding dogs, DI be remeasured after maturity to confirm DI obtained at earlier ages. Studies including other breeds of dogs should be done to determine the youngest reliable age to initiate hip joint screening.
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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.
Antibody-directed enzyme prodrug therapy (ADEPT) has the potential of greatly enhancing antitumor selectivity of cancer therapy by synthesizing chemotherapeutic agents selectively at tumor sites. This therapy is based upon targeting a prodrug-activating enzyme to a tumor by attaching the enzyme to a tumor-selective antibody and dosing the enzyme-antibody conjugate systemically. After the enzyme-antibody conjugate is localized to the tumor, the prodrug is then also dosed systemically, and the previously targeted enzyme converts it to the active drug selectively at the tumor. Unfortunately, most enzymes capable of this specific, tumor site generation of drugs are foreign to the human body and as such are expected to raise an immune response when injected, which will limit their repeated administration. We reasoned that with the power of crystallography, molecular modeling and site-directed mutagenesis, this problem could be addressed through the development of a human enzyme that is capable of catalyzing a reaction that is otherwise not carried out in the human body. This would then allow use of prodrugs that are otherwise stable in vivo but that are substrates for a tumor-targeted mutant human enzyme. We report here the first test of this concept using the human enzyme carboxypeptidase A1 (hCPA1) and prodrugs of methotrexate (MTX). Based upon a computer model of the human enzyme built from the well known crystal structure of bovine carboxypeptidase A, we have designed and synthesized novel bulky phenylalanine- and tyrosine-based prodrugs of MTX that are metabolically stable in vivo and are not substrates for wild type human carboxypeptidases A. Two of these analogs are MTX-alpha-3-cyclobutylphenylalanine and MTX-alpha-3-cyclopentyltyrosine. Also based upon the computer model, we have designed and produced a mutant of human carboxypeptidase A1, changed at position 268 from the wild type threonine to a glycine (hCPA1-T268G). This novel enzyme is capable of using the in vivo stable prodrugs, which are not substrates for the wild type hCPA1, as efficiently as the wild type hCPA1 uses its best substrates (i.e. MTX-alpha-phenylalanine). Thus, the kcat/Km value for the wild type hCPA1 with MTX-alpha-phenylalanine is 0.44 microM-1 s-1, and kcat/Km values for hCPA1-T268G with MTX-alpha-3-cyclobutylphenylalanine and MTX-alpha-3-cyclopentyltyrosine are 1.8 and 0.16 microM-1 s-1, respectively. The cytotoxic efficiency of hCPA1-268G was tested in an in vitro ADEPT model. For this experiment, hCPA1-T268G was chemically conjugated to ING-1, an antibody that binds to the tumor antigen Ep-Cam, or to Campath-1H, an antibody that binds to the T and B cell antigen CDw52. These conjugates were then incubated with HT-29 human colon adenocarcinoma cells (which express Ep-Cam but not the Campath 1H antigen) followed by incubation of the cells with the in vivo stable prodrugs. The results showed that the targeted ING-1:hCPA1-T268G conjugate produced excellent activation of the MTX prodrugs to kill HT-29 cells as efficiently as MTX itself. By contrast, the enzyme-Campath 1H conjugate was without effect. These data strongly support the feasibility of ADEPT using a mutated human enzyme with a single amino acid change.
The DI method was developed, at least in part, because of perceived variations among radiologists in regard to subjective hip scores assigned to radiographs. It also was recognized that the prevalence of CHD among many dog breeds was disturbingly high, despite efforts to lower the prevalence using subjective score as a selection criterion. The DI method was developed on the basis of results of mechanical testing of cadaver hip joints, which accomplished 3 principal objectives. First, it showed that when the hip joint is in the standard, hip-extended, internally rotated position, passive laxity is minimized. Second, it showed that passive laxity was maximized when the hip joint is in the neutral position. Third, it demonstrated the sigmoidal nature of the load/displacement curve for the hip joint in the neutral position, which suggested that high repeatability from examiner to examiner in regard to DI could be expected without the need to standardize applied force. The mean and range of DI were shown to vary from 1 breed to the next, but within an individual dog, DI appears to remain constant (within limits of scientific acceptability and clinical applicability) from 16 weeks of age. Passive hip laxity measured on the distraction view is, on average, 2.5 to 11 times greater than that measured on the standard, hip-extended radiographic view. Performance Borzois and Greyhounds, which have an extremely low prevalence of CHD, uniformly have tight hip joints (DI < 0.3), and mean DI for dog breeds that have a high prevalence of CHD is significantly greater than mean DI for Borzois and Greyhounds. However, individual dogs that have DI < 0.3, even though members of breeds prone to CHD, have a low risk of developing DJD. Some, but not all, dogs with DI > 0.3 will develop DJD by 3 years of age, and DI has been shown to be the principal risk factor for development of DJD. Susceptibility to development of DJD appears to be breed-specific. For example, given equivalent DI, German Shepherd Dogs are more at risk for developing DJD than are Rottweilers. Heritability of DI is higher than that of the subjective hip score; thus, selection pressure based on DI should result in faster genetic change than selection pressure based on subjective hip scores. Finally, the DI method has been performed on approximately 14,000 dogs, some of which were evaluated multiple times. It is apparent that this method is no more harmful than the standard, hip-extended radiographic method or palpation performed as part of a routine orthopedic examination.
OBJECTIVE: To determine risk factors for, and prevalence and short- and long-term effects of synovial fluid cavitation during distraction radiography. DESIGN: Multicenter prevalence survey. ANIMALS: 6,649 purebred dogs comprising 129 breeds. PROCEDURE: Radiographs from the PennHIP (University of Pennsylvania Hip Improvement Program) Laboratory were subjectively evaluated for evidence of cavitation. Multiple logistic regression was used to determine whether sex, breed, age, weight, distraction index (DI), or examining veterinarian was associated with cavitation. Short-term effects of cavitation were assessed by comparing DI for the hip with cavitation with DI for the contralateral hip in dogs with unilateral cavitation. Long-term effects of cavitation were assessed by comparing DI before and after cavitation was detected. RESULTS: Cavitation was detected in 279 (4.2%) of the radiographs analyzed. Male dogs, Golden Retrievers, and heavier dogs were at a decreased risk for cavitation. Irish Wolfhounds, Irish Setters, Rhodesian Ridgebacks, and Weimaraners had an increased risk for cavitation. Age and DI were not risk factors for cavitation. Mean DI was 0.08 greater in hips with cavitation than in paired hips without cavitation. Significant differences were not detected between DI before and after cavitation, but only 7 dogs were included in this analysis. CLINICAL IMPLICATIONS: Cavitation is rare during distraction radiography and can increase measured DI. Radiographs should be routinely examined to ensure accurate reporting of DI.
OBJECTIVE: To examine the effects of limited food intake on frequency and severity of osteoarthritis in coxofemoral joints of labrador Retrievers. DESIGN: Dogs were paired according to gender and body weight, within each litter at 8 weeks of age. One dog of each pair was fed ad libitum. The limit-fed pairmate was fed 75% of the amount eaten the previous day by the ad libitum-fed counterpart. ANIMALS: 48 Labrador Retrievers. PROCEDURE: All dogs received the same diet. Radiographic evaluation of coxofemoral joints for frequency and severity of osteoarthritis were made when dogs were 4 and 6 months and 1, 2, 3, and 5 years old. RESULTS: Radiographic evaluation for osteoarthritis indicated greater frequency and more severity of osteoarthritis in the ad libitum-fed group of dogs. CLINICAL IMPLICATIONS: Analysis of data suggested that limit feeding of dogs over a 5-year period minimizes development of osteoarthritis in the coxofemoral joints.
OBJECTIVE: To evaluate in vivo repeatability of the distraction index method of evaluating hip joint laxity in dogs. ANIMALS: 31 two-year-old Labrador Retrievers. PROCEDURE: Each dog was anesthetized and radiographically evaluated for hip joint laxity 4 times: twice by an experienced examiner and twice by an examiner who had no previous knowledge of or training in the technique prior to the first day of testing. Distraction indices (DI) were determined from the radiographs and intraclass correlation coefficients were calculated to evaluate the repeatability of DI measurements between and within examiners. RESULTS: Intraclass correlation coefficients were high (range, 0.85 to 0.94). Lower limits of the 95% confidence intervals for the intraclass correlation coefficients ranged from 0.75 to 0.89. CONCLUSIONS: Between- and within-examiner repeatabilities of DI measurements were high, suggesting that the technique is clinically reliable. CLINICAL RELEVANCE: Distraction index is a reliable measure of hip joint laxity and a good predictor of the risk of development of degenerative joint disease associated with hip dysplasia in dogs. Establishment of high repeatability of DI measurements suggests that the stress-radiographic method may be used by multiple examiners with the expectation of comparable and consistent results.