Biomedical subjects
Publications and source records attributed to J H Wilson.
A 25-year-old Quarterhorse mare was euthanized for a variety of medical reasons. At necropsy, 7 liver flukes, identified as Fascioloides magna, were recovered from the liver. This is the first report of F. magna in a horse.
The objectives of this investigation were to assess the relationship between chemical measures and imaging estimates (radiographic photometry and dual-energy x-ray absorptiometry) of bone mineral content in dairy cows and to evaluate the effects of parity, stage of lactation, and site of measurement (fused third and fourth metacarpal bone vs. caudal vertebrae 14 and 15) on bone mineral content. In a preliminary study, the caudal vertebrae were excised from 33 cows following slaughter. Samples were analyzed by radiographic photometry and then analyzed for mineral content chemically. In a second experiment, the caudal vertebrae and right front metacarpal (sample pairs) were excised from 107 Holstein cull cows following slaughter. Parity and days in milk (DIM) of the donor animals were obtained for 43 pairs of samples. Samples were grouped by parity (1, 2, 3, and >or=4) stage of lactation (Stage 1: <90 DIM, Stage 2: 90 to 150 DIM, Stage 3: 151 to 250 DIM, and Stage 4: >250 DIM). Samples were analyzed by radiographic photometry and dual-energy x-ray absorptiometry and then analyzed for mineral content chemically. In both experiments, the relationship between mineral content estimated via the imaging techniques and mineral content measured chemically was poor, likely because of the relative maturity of animals in the sample set and lack of variation in mineral content. Ash content was higher in the metacarpal than in the caudal vertebrae, as were concentrations of Mg (expressed as a proportion of bone ash). No effects of stage of lactation were observed on bone mineral in the caudal vertebrae, but in the metacarpal, P content (proportion of total mineral) was highest in second lactation cows. Total bone mineral content (ash) was not affected by parity in the metacarpal or caudal vertebra, but Ca and P content of the metacarpal increased with parity. Noninvasive imaging techniques are not sufficiently sensitive to detect changes in mineral content or composition of mature cows, and only modest changes in bone mineral were observed with stage of lactation and parity.
This study evaluated the effects of maternal dietary lipids on chemical components and mechanical and histological properties of tibia in progeny of Japanese quail fed different dietary lipids. Laying hens were fed a basal diet containing either soybean oil (SBO), hydrogenated soybean oil (HSBO), chicken fat (CF), or menhaden fish oil (FO) at 50 g/kg of the diet. The various maternal dietary lipid treatments did not affect growth of progeny at any developmental stage. There were no differences in tibial length, diameter, or collagen content. Tibial percentage ash was significantly higher in newly hatched progeny from hens fed the FO and HSBO diets. The levels of tibial deoxypyridinoline and total pyridinium crosslinks were higher in the FO and HSBO groups at hatch. At 7 d of age, the tibial deoxypyridinoline links remained higher in the FO group compared to the CF and SBO groups. Likewise, progeny from hens consuming the FO or HSBO diet had higher tibial shear force and stiffness at 7 and 14 d of age. There were no pronounced differences in tibial fracture energy and deflection among treatments. Maternal FO or HSBO enlarged the cartilaginous proliferative and hypertrophic zones of the tibial proximal end in newly hatched quail, which was accompanied by a thicker cortical bone in the diaphysis. However, the width of the hypertrophic zones tended to be smaller in these two groups coupled with improvement in trabecular density and cortical thickness in the proximal end and cortical density in the diaphysis at 3 wk of age. These results suggest that maternal dietary lipids altered bone development by influencing organic matrix quality and mineralization in embryos.
The purpose of this study was to investigate the effects of long-term supplementation of fat in the diets on the fatty acid composition, chemical, mechanical, and histological properties of tibial bone. Month-old male Japanese quail were fed a basal diet containing either soybean oil (SBO), hydrogenated soybean oil (HSBO), chicken fat (CF), or menhaden fish oil (FO) at 50 g/kg of the diet and maintained on these diets for 7 mo. Lipid treatments did not affect body weight, food intake, tibial length, or diameter. The FO diet group had the highest percentage of tibial ash, and both the FO and HSBO significantly increased tibial mineral content compared to those given SBO or CF. The type and amount of fatty acids in the diets had a profound influence on fatty acid composition of lipids in tibial cortical bones. Quail fed FO had the highest concentration of (n-3) fatty acids, and those fed SBO were highest in (n-6) fatty acids. The HSBO diet, containing high level of trans-fatty acids, led to the accumulation of these fatty acids in bone. In quail, long-term supplementation of FO or HSBO increased tibial shear force and shear stress and improved histological cortical thickness and density when compared to those given SBO or CF. These results suggest that long-term exposure to a FO or HSBO diet have a significant beneficial effect on bone metabolism.
Gene targeting in mammalian cells has proven invaluable in biotechnology, in studies of gene structure and function, and in understanding chromosome dynamics. It also offers a potential tool for gene-therapeutic applications. Two limitations constrain the current technology: the low rate of homologous recombination in mammalian cells and the high rate of random (nontargeted) integration of the vector DNA. Here we consider possible ways to overcome these limitations within the framework of our present understanding of recombination mechanisms and machinery. Several studies suggest that transient alteration of the levels of recombination proteins, by overexpression or interference with expression, may be able to increase homologous recombination or decrease random integration, and we present a list of candidate genes. We consider potentially beneficial modifications to the vector DNA and discuss the effects of methods of DNA delivery on targeting efficiency. Finally, we present work showing that gene-specific DNA damage can stimulate local homologous recombination, and we discuss recent results with two general methodologies--chimeric nucleases and triplex-forming oligonucleotides--for stimulating recombination in cells.
OBJECTIVE: To evaluate the effect of Ridogrel enemas (Janssen Research Foundation, Beerse, Belgium) on disease activity and mucosal inflammatory mediators in patients with active left-sided ulcerative colitis. DESIGN AND METHODS: Eleven patients with active left-sided ulcerative colitis were evaluated in an open non-placebo-controlled pilot study. All patients were treated with Ridogrel enemas (300 mg/40 ml once daily) over four weeks. A disease activity score based on clinical, endoscopic and histological criteria was obtained before and after treatment with Ridogrel. The concentrations of thromboxane B2 (TxB2), prostaglandin E2 (PGE2), interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF-alpha) were measured in mucosal biopsies before and after treatment. RESULTS: One patient discontinued treatment because of progression of disease, the other ten patients tolerated the Ridogrel enemas well. Mucosal TxB2 concentration decreased significantly in all patients. The mucosal concentrations of the other inflammatory mediators (PGE2, IL-6 and TNF-alpha) were unaltered. The disease score decreased in five patients. However, clinical improvement was not always associated with a decrease in endoscopic and/or histological scores. CONCLUSIONS: This pilot study shows that Ridogrel enemas selectively reduce mucosal TxB2 concentration.
BACKGROUND: Endoscopic cancer surveillance has been advocated for patients with Barrett's oesophagus. However, only a small minority of patients dies from adenocarcinoma in Barrett's oesophagus. It has been calculated that endoscopic cancer surveillance will only add to the quality of life of individuals in whom the incidence of adenocarcinoma in Barrett's oesophagus is greater than 1/200 patient-years. OBJECTIVE: To determine the proportion of a consecutive cohort of patients, in whom Barrett's oesophagus was diagnosed over a 5-year period, likely to benefit from endoscopic cancer surveillance. METHODS: All patients who had died during the observation period or were over 75 years old and those with diseases likely to impair survival were excluded. Next, all patients in whom the risk of developing adenocarcinoma in Barrett's oesophagus fell below 1/200 patient-years were excluded (including all women, all men under the age of 60 and all men with Barrett's oesophagus of < 3 cm in length). Patients with dysplasia of any degree and/or presence of an ulcer or stricture in Barrett's oesophagus were reinstated. RESULTS: Of 335 adult patients diagnosed with Barrett's oesophagus but without adenocarcinoma or high-grade dysplasia, 75 had died from unrelated causes, 47 had other diseases limiting survival and 59 were over 75 years old. After exclusion of all women, all men with Barrett's oesophagus of < 3 cm in length and all men under 60 years old, 15 patients were left. However, 32 were reinstated because of risk factors and another five because of insufficient data, resulting in 52 of the original 335 patients (15.5%) being eligible for endoscopic cancer surveillance. CONCLUSION: This study suggests that less than 20% of patients identified with Barrett's oesophagus at routine endoscopy would benefit from endoscopic cancer surveillance. Prospective surveillance programmes should be limited to patients with an increased cancer risk and a good health profile.
Telomere repeat sequences cap the ends of eucaryotic chromosomes and help stabilize them. At interstitial sites, however, they may destabilize chromosomes, as suggested by cytogenetic studies in mammalian cells that correlate interstitial telomere sequence with sites of spontaneous and radiation-induced chromosome rearrangements. In no instance is the length, purity, or orientation of the telomere repeats at these potentially destabilizing interstitial sites known. To determine the effects of a defined interstitial telomere sequence on chromosome instability, as well as other aspects of DNA metabolism, we deposited 800 bp of the functional vertebrate telomere repeat, TTAGGG, in two orientations in the second intron of the adenosine phosphoribosyltransferase (APRT) gene in Chinese hamster ovary cells. In one orientation, the deposited telomere sequence did not interfere with expression of the APRT gene, whereas in the other it reduced mRNA levels slightly. The telomere sequence did not induce chromosome truncation and the seeding of a new telomere at a frequency above the limits of detection. Similarly, the telomere sequence did not alter the rate or distribution of homologous recombination events. The interstitial telomere repeat sequence in both orientations, however, dramatically increased gene rearrangements some 30-fold. Analysis of individual rearrangements confirmed the involvement of the telomere sequence. These studies define the telomere repeat sequence as a destabilizing element in the interior of chromosomes in mammalian cells.
AIM: To investigate whether mutations in the STK11/LKB1 gene and genes implicated in the colorectal adenoma-carcinoma sequence are involved in Peutz-Jeghers syndrome (PJS) related tumorigenesis. METHODS: Thirty nine polyps and five carcinomas from 17 patients (from 13 families) with PJS were analysed for loss of heterozygosity (LOH) at 19p13.3 (STK11/LKB1 gene locus), 5q21 (APC gene locus), 18q21-22 (Smad4 and Smad2 gene locus), and 17p13 (p53 gene locus), and evaluated for immunohistochemical staining of p53. In addition, mutational analysis of K-ras codon 12, APC, and p53 and immunohistochemistry for Smad4 expression were performed on all carcinomas. RESULTS: LOH at 19p was seen in 15 of the 39 polyps and in all carcinomas (n = 5). Interestingly, six of the seven polyps from patients with cancer had LOH, compared with nine of the 31 polyps from the remaining patients (p = 0.01). In one polyp from a patient without a germline STK11/LKB1 mutation, no LOH at 19p or at three alternative PJS candidate loci (19q, 6p, and 6q) was found. No LOH at 5q was observed. However, mutational analysis revealed an APC mutation in four of the five carcinomas. LOH at 17p was not seen in polyps or carcinomas; immunohistochemistry showed expression of p53 in one carcinoma and focal expression in three polyps. At subsequent sequence analysis, no p53 mutation was found. One carcinoma had an activating K-ras codon 12 mutation and another carcinoma showed 18q LOH; however, no loss of Smad4 expression was seen. CONCLUSIONS: These results provide further evidence that STK11/LKB1 acts as a tumour suppressor gene, and may be involved in the early stages of PJS tumorigenesis. Further research is needed to see whether LOH in PJS polyps could be used as a biomarker to predict cancer. Differences in molecular genetic alterations noted between the adenoma-carcinoma sequence and PJS related tumours suggest the presence of a distinct pathway of carcinogenesis.
The effects of dietary nickel (0, 25, 50, 75, 100, and 150 mg/kg) on the bone strength characteristics and performance parameters of male broilers were investigated. Broilers were housed in either cages or floor pens. At 6 wk of age, the shear fracture energy of the tibia from the caged birds increased when the basal diet was supplemented with 25 mg of dietary nickel per kilogram of feed. The shear force, stress, and fracture energy of the radius from the caged birds also increased at 25 mg/kg nickel. Dietary nickel had no effect on bird body weight, but the caged broilers (2,161 g) were heavier than the floor birds (2,005 g). Nickel had no effect on the strength characteristics of the tibia from the floor birds. Percent tibia bone ash, a measure of bone density, was not influenced by dietary nickel, but the tibia ash of the floor birds was greater than that of the caged birds. Overall, the data indicates that adding 25 mg/kg of dietary nickel to a poultry diet will have a positive influence on bone strength characteristics and performance.
Diets containing 0.31, 0.39, or 0.47% P on a DM basis were fed to 10, 14, and 13 multiparous Holstein cows, respectively, for a full lactation. Most of the cows (33) were fed similar amounts of P in the previous one or two lactations. The objective was to obtain information on bone integrity after prolonged feeding of different amounts of P. At the end of the experiment, a section (-20 cm) of the 12th rib was surgically removed. The bone was tested for strength and analyzed for ash and P content. The shear strength and the energy required to deform the bone to the point of fracture did not differ among treatments. Bone specific gravities were 1.50, 1.57, and 1.55 for the three treatments. Ash and P content of the bone, measured in wet weight, dry weight, and wet bone volume, were similar for the 0.39 and 0.47% P treatments, but tended to be lower with the 0.31% P treatment. Milk production of cows in all groups was high, averaging >11,900 kg for the 308-d lactation. Feeding P at 0.31% of dietary DM over two to three lactations appeared to decrease P concentration of bone, but the decrease was not severe enough to affect bone strength. Dietary P at 0.39% did not affect bone P content or strength. Dietary P content of 0.31% appears to be borderline deficient for cows producing >11,900 kg/308 d.
Prolactin is released in response to physical and psychological stress, the latter of which involves an animal's interpretation of its environment as potentially dangerous. This interpretation might be altered by social buffering, as defined by the presence of a social partner. In support of this claim, I previously found that the presence of a conspecific during exposure to the open field caused lower levels of prolactin in juvenile rats than did exposure to the open field alone. Because testing with a conspecific allowed the animals to touch, physical contact may have been necessary for the reduction of prolactin. To test this hypothesis, juvenile male and female rats in the present study were exposed to the open field (1) alone, (2) with a conspecific separated by a Plexiglas partition, or (3) with a conspecific without a partition. Touch was necessary to reduce stress as measured by prolactin levels.
To explore the ability of triplex-forming oligodeoxyribonucleotides (TFOs) to inhibit genes responsible for dominant genetic disorders, we used two TFOs to block expression of the human rhodopsin gene, which encodes a G protein-coupled receptor involved in the blinding disorder autosomal dominant retinitis pigmentosa. Psoralen-modified TFOs and UVA irradiation were used to form photoadducts at two target sites in a plasmid expressing a rhodopsin-EGFP fusion, which was then transfected into HT1080 cells. Each TFO reduced rhodopsin-GFP expression by 70-80%, whereas treatment with both reduced expression by 90%. Expression levels of control genes on either the same plasmid or one co-transfected were not affected by the treatment. Mutations at one TFO target eliminated its effect on transcription, without diminishing inhibition by the other TFO. Northern blots indicated that TFO-directed psoralen photoadducts blocked progression of RNA polymerase, resulting in truncated transcripts. Inhibition of gene expression was not relieved over a 72 h period, suggesting that TFO-induced psoralen lesions are not repaired on this time scale. Irradiation of cells after transfection with plasmid and psoralen-TFOs produced photoadducts inside the cells and also inhibited expression of rhodopsin-EGFP. We conclude that directing DNA damage with psoralen-TFOs is an efficient and specific means for blocking transcription from the human rhodopsin gene.
The XpF/Ercc1 structure-specific endonuclease performs the 5' incision in nucleotide excision repair and is the apparent mammalian counterpart of the Rad1/Rad10 endonuclease from Saccharomyces cerevisiae. In yeast, Rad1/Rad10 endonuclease also functions in mitotic recombination. To determine whether XpF/Ercc1 endonuclease has a similar role in mitotic recombination, we targeted the APRT locus in Chinese hamster ovary ERCC1(+) and ERCC1(-) cell lines with insertion vectors having long or short terminal non-homologies flanking each side of a double-strand break. No substantial differences were evident in overall recombination frequencies, in contrast to results from targeting experiments in yeast. However, profound differences were observed in types of APRT(+) recombinants recovered from ERCC1(-) cells using targeting vectors with long terminal non-homologies-almost complete ablation of gap repair and single-reciprocal exchange events, and generation of a new class of aberrant insertion/deletion recombinants absent in ERCC1(+) cells. These results represent the first demonstration of a requirement for ERCC1 in targeted homologous recombination in mammalian cells, specifically in removal of long non-homologous tails from invading homologous strands.
Spontaneous recombination between direct repeats at the adenine phosphoribosyltransferase (APRT) locus in ERCC1-deficient cells generates a high frequency of rearrangements that are dependent on the process of homologous recombination, suggesting that rearrangements are formed by misprocessing of recombination intermediates. Given the specificity of the structure-specific Ercc1/Xpf endonuclease, two potential recombination intermediates are substrates for misprocessing in ERCC1(-) cells: heteroduplex loops and heteroduplex intermediates with non-homologous 3' tails. To investigate the roles of each, we constructed repeats that would yield no heteroduplex loops during spontaneous recombination or that would yield two non-homologous 3' tails after treatment with the rare-cutting endonuclease I-SCE:I. Our results indicate that misprocessing of heteroduplex loops is not the major source of recombination-dependent rearrangements in ERCC1-deficient cells. Our results also suggest that the Ercc1/Xpf endonuclease is required for efficient removal of non-homologous 3' tails, like its Rad1/Rad10 counterpart in yeast. Thus, it is likely that misprocessing of non-homologous 3' tails is the primary source of recombination-dependent rearrangements in mammalian cells. We also find an unexpected effect of ERCC1 deficiency on I-SCE:I-stimulated rearrangements, which are not dependent on homologous recombination, suggesting that the ERCC1 gene product may play a role in generating the rearrangements that arise after I-SCE:I-induced double-strand breaks.
A 2.5-year-old female Thoroughbred was examined because of lethargy, anorexia, and weight loss. Analysis of a CBC revealed erythrocytosis and an increase in PCV. Serum biochemical analysis revealed increases in activities of several hepatic enzymes. Ultrasonography revealed hepatomegaly and a heterogeneous appearance of the hepatic parenchyma. The horse did not improve despite supportive care, and it was euthanatized. Necropsy revealed numerous raised white to gray foci in the liver. Histologically, these foci consisted of neoplastic cells that resembled fetal hepatocytes, embryonal-type cells, and cells with features intermediate between those 2 cell types. Immunohistochemical staining revealed that hepatocytes stained strongly with anti-alpha-fetoprotein. On the basis of these results, hepatoblastoma was diagnosed. Diagnosis of hepatoblastoma is difficult, because it can appear histologically similar to other hepatic tumors, such as hepatocellular carcinomas. Definitive diagnosis requires histologic evaluation of tumor architecture and cell morphology. Immunohistochemical staining for alpha-fetoprotein in tumor cells may serve as a tumor marker but is not pathognomonic of hepatoblastoma. Paraneoplastic syndromes, such as erythrocytosis, can accompany hepatoblastoma. The prognosis for horses with hepatoblastoma is grave.
BACKGROUND: Extracellular adenosine 5'-triphosphate (ATP) is involved in the regulation of a variety of biologic processes, including neurotransmission, muscle contraction, and liver glucose metabolism, via purinergic receptors. In nonrandomized studies involving patients with different tumor types including non-small-cell lung cancer (NSCLC), ATP infusion appeared to inhibit loss of weight and deterioration of quality of life (QOL) and performance status. We conducted a randomized clinical trial to evaluate the effects of ATP in patients with advanced NSCLC (stage IIIB or IV). METHODS: Fifty-eight patients were randomly assigned to receive either 10 intravenous 30-hour ATP infusions, with the infusions given at 2- to 4-week intervals, or no ATP. Outcome parameters were assessed every 4 weeks until 28 weeks. Between-group differences were tested for statistical significance by use of repeated-measures analysis, and reported P values are two-sided. RESULTS: Twenty-eight patients were allocated to receive ATP treatment and 30 received no ATP. Mean weight changes per 4-week period were -1.0 kg (95% confidence interval [CI] = -1.5 to -0.5) in the control group and 0.2 kg (95% CI = -0.2 to +0.6) in the ATP group (P =.002). Serum albumin concentration declined by -1.2 g/L (95% CI= -2.0 to -0.4) per 4 weeks in the control group but remained stable (0.0 g/L; 95% CI = -0.3 to +0.3) in the ATP group (P =.006). Elbow flexor muscle strength declined by -5.5% (95% CI = -9.6% to -1. 4%) per 4 weeks in the control group but remained stable (0.0%; 95% CI= -1.4% to +1.4%) in the ATP group (P =.01). A similar pattern was observed for knee extensor muscles (P =.02). The effects of ATP on body weight, muscle strength, and albumin concentration were especially marked in cachectic patients (P =.0002, P =.0001, and P =. 0001, respectively, for ATP versus no ATP). QOL score changes per 4-week period in the ATP group showed overall less deterioration than in the control group-physical scores (-0.2% versus -2.4%; P =. 0002); functional scores (+0.4% versus -5.5%; P =.02); psychologic scores (-0.7% versus -2.4%; P =.11); overall QOL score (+0.1% versus -3.5%; P =.0001). CONCLUSIONS: This randomized trial demonstrates that ATP has beneficial effects on weight, muscle strength, and QOL in patients with advanced NSCLC.
Profound alterations in host metabolism in lung cancer patients with weight loss have been reported, including elevated phosphomonoesters (PMEs) as detected by 31P magnetic resonance spectroscopy (MRS). In healthy subjects, infusion of L-alanine induced significant increases in hepatic PMEs and phosphodiesters (PDEs) due to rising concentrations of 3-phosphoglycerate and phosphoenolpyruvate, respectively. The aim of the present study was to monitor these changes in the tumor-free liver of lung cancer patients during L-alanine infusion by means of simultaneous 31P MRS and turnover measurements. Twenty-one lung cancer patients without liver metastases with (CaWL) or without weight loss (CaWS), and 12 healthy control subjects were studied during an i.v. L-alanine challenge of 1.4-2.8 mmol/kg followed by 2.8 mmol/kg/h for 90 min. Plasma L-alanine concentrations increased during alanine infusion, from 0.35-0.37 mM at baseline to 5.37 +/- 0.14 mM in the CaWL patients, 6.67 +/- 0.51 mM in the CaWS patients, and 8.47 +/- 0.88 mM in the controls (difference from baseline and between groups during alanine infusion, all P < 0.001). Glucose turnover and liver PME levels at baseline were significantly elevated in the CaWL patients. Alanine infusion increased whole-body glucose turnover by 8 +/- 3% in the CaWS patients (P = 0.03), whereas no significant change occurred in the CaWL and controls. PME levels increased by 50 +/- 16% in controls (area under the curve, P < 0.01) and by 87 +/- 31% in the CaWS patients (P < 0.05) after 45-90 min. In contrast, no significant changes in PME levels were observed in the CaWL patients. Plasma insulin concentrations increased during L-alanine infusion in all groups to levels that were lower in the CaWL patients than in the CaWS patients and controls (P < 0.05). In lung cancer patients, but not in controls, changes in PME and PDE levels during alanine infusion were inversely correlated with their respective baseline levels (r = -0.82 and -0.86, respectively; P < 0.001). In addition, changes in PMEs during alanine infusion in lung cancer patients were inversely correlated with the degree of weight loss (r = -0.54; P < 0.05). This study demonstrates the presence of major alterations in the pathway of hepatic gluconeogenesis in weight-losing lung cancer patients, as shown by elevated glucose flux before and during L-alanine infusion, and by the increased PME and PDE levels, which reflect accumulation of gluconeogenic intermediates in these patients. Weight-stable lung cancer patients show accelerated increases in PME and PDE levels during L-alanine infusion, suggesting enhanced induction of the gluconeogenic pathway. Our results suggest altered gluconeogenic enzyme activities and elevated alanine uptake within the livers of weight-losing/weight-stable lung cancer patients.