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Biomedical subjects

A J Hamilton

Publications and source records attributed to A J Hamilton.

At least 19 recordsLinked to original sources
PubMed1996

Unusual early recurrence of a cerebellar pilocytic astrocytoma following complete surgical resection. Case report and review of the literature.

Pilocytic cerebellar astrocytomas are usually benign tumors with generally an excellent prognosis following complete surgical resection. The goal of surgery is total resection to minimize the risk of recurrence. In this case report, a 5-year old boy who had undergone total resection of a posterior fossa pilocytic cerebellar astrocytoma (as documented by a contrast-enhanced computed tomography (CT) scan within 24 hours following surgery), developed a massive recurrence of the tumor within four months. Both the initial histology and the sections examined after the second resection revealed features typical for a pilocytic astrocytoma with no suspicion of malignancy. This case is unusual in that it is contrary to other reports suggesting that CT-documented complete surgical resection of pilocytic astrocytomas is without recurrence, and suggests the need for vigilant radiographic and clinical follow-up of these patients even if apparent complete resection of the tumor has been achieved.

Astrocytoma↗
PubMed1996

Purification and characterisation of a novel 34,000-Mr cell-associated proteinase from the dermatophyte Trichophyton rubrum.

A novel cell-associated proteinase was purified to homogeneity from cytoplasmic antigen preparations of Trichophyton rubrum by sequential isoelectric focusing and gel filtration chromatography. The enzyme exhibited relative molecular masses of 34,000-Mr (non-reduced sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE)), 15,000-Mr (reduced SDS-PAGE) and 37,000-Mr (substrate SDS-PAGE). It had a pH optimum of 7.5 and a pI of 4.5. The proteinase exhibited broad substrate specificity and it was strongly inhibited by the serine proteinase inhibitors phenylmethylsulfonyl fluoride and chymostatin. The N-terminal amino acid sequence of the 34,000-Mr proteinase shared 50% homology with the deduced amino acid sequence of a Coccidioides immitis wall-associated chymotrypsin-type serine proteinase. This is the first cell-associated proteinase to be purified and characterised from T. rubrum and it would appear to be related to the chymotrypsin-type serine proteinases, a class of enzymes that have rarely been isolated from fungi. The function of the proteinase remains speculative although it may play a role in the development and subsequent proliferation of the fungus in vivo.

Amino Acid Sequence↗
PubMed1996

Differential expression of the 1-aminocyclopropane-1-carboxylate oxidase gene family of tomato.

The tomato ACC oxidase gene family is comprised of three members designated AC01, AC02 and AC03. These are highly homologous throughout the protein coding regions but do show a degree of sequence divergence within the 3' untranslated regions. These regions have been cloned and used as gene-specific probes to analyse the differential expression of the tomato ACC oxidase gene family in various tissues at different stages of development. Results indicate that all three genes are transcriptionally active and display a high degree of inducibility in a number of organs at various stages of the life cycle. Both AC01 and Ac03 transcripts accumulate during the senescence of leaves, fruit and flowers. In addition, it appears that AC01 is wound-inducible in leaves. All three ACC oxidase genes are expressed during flower development, with each showing a temporally distinct pattern of accumulation. In addition, the ACC oxidase transcripts are also spatially regulated throughout flower development; AC01 is predominantly expressed in the petals and the stigma and style, AC02 expression is mainly restricted to tissues associated with the anther cone whereas AC03 transcripts accumulate in all of the floral organs examined apart from the sepals. ACC oxidase enzyme assays and Western blot analysis indicate that both enzyme activity and ACC oxidase protein increase with transcript abundance in several tissues. The physiological role of the differential expression of the ACC oxidase gene family, in relation to the regulation of ethylene synthesis, during these various developmental processes is discussed.

Aging↗
PubMed1996

Purification and partial characterization of the Cu,Zn superoxide dismutase from the dermatophyte Trichophyton mentagrophytes var. interdigitale.

Cell homogenization, isoelectric focusing and gel filtration FPLC have been used to purify a superoxide dismutase (SOD) from the dermatophyte Trichophyton mentagrophytes var. interdigitale (T. interdigitale). N-terminal amino acid sequencing identified this enzyme as a Cu,ZnSOD, with a pH of 5.1, a reduced molecular mass of 18 kDa, and a non-reduced molecular mass of 59 kDa. SOD activity was detectable in culture filtrates, as early as the mid-log phase of growth. The known Cu,Zn inhibitor potassium cyanide caused some inhibition of the purified enzyme, whereas the inhibitors sodium azide, guanidinium hydrochloride, EDTA and chloroform/ethanol had no discernible effect. The T. interdigitale SOD was pH insensitive in the range 7.0-10.5 and remained active after prolonged incubation at 50 degrees C. The purification and characterization of this enzyme represents the first step in determining whether SOD plays any part in protecting T. interdigitale from free radicals generated by the oxidative burst of immune effector cells.

Amino Acid Sequence↗
PubMed1996

Expression of the Cu,Zn superoxide dismutase of Aspergillus fumigatus as determined by immunochemistry and immunoelectron microscopy.

A polyclonal antibody against purified Cu,Zn superoxide dismutase (SOD) from the pathogen Aspergillus fumigatus was raised in a sheep. This antibody recognised purified A. fumigatus SOD, together with a single band of 19 kDa in A. fumigatus cytoplasmic antigen, by immunodevelopment of Western blots. The polyclonal serum did not recognise either the manganese or iron containing forms of the enzyme; however, it was reactive against putative Cu,Zn SODs in other members of the genus Aspergillus. Immunofluorescent staining of A. fumigatus cultures demonstrated expression of the Cu,Zn SOD in conidia and hyphae, with the cell wall staining particularly intensely. Conidiophores were stained in an uniformly intense pattern. Immunoelectron microscopy confirmed that the SOD was present within the hyphal cell wall, although there was also labelling in the cytoplasm. SOD may protect Aspergillus against oxidants produced by immune effector cells and these observations demonstrate that the enzyme is available to perform its antioxidant function within the cell wall.

Antibodies, Fungal↗
PubMed1996

The Cu,Zn superoxide dismutases of Aspergillus flavus, Aspergillus niger, Aspergillus nidulans, and Aspergillus terreus: purification and biochemical comparison with the Aspergillus fumigatus Cu,Zn superoxide dismutase.

Cu,Zn superoxide dismutases (SODs) have been purified to homogeneity from Aspergillus flavus and A. niger, which are significant causative agents of aspergillosis, and from A. nidulans and A. terreus, which are much rarer causative agents of disease, using a combination of isoelectric focusing and gel filtration fast protein liquid chromatography. The purified enzymes have been compared with the previously described SOD from the most important pathogen in the genus, A. fumigatus (M. D. Holdom, R. J. Hay, and A. J. Hamilton, Free Radical Res. 22:519-531, 1995). The N-terminal amino acid sequences of the four newly purified enzymes were almost identical and demonstrated homology to known Cu,Zn SODs from a range of organisms including that from the previously described SOD from A. fumigatus. SOD activity was detectable in the culture filtrates of all species, and intracellular Cu,Zn SOD activity as a proportion of total protein was highest in early-log-phase cultures. The specific activities of the purified enzymes were similar, and all four of the newly described enzymes were inhibited by potassium cyanide and diethyldithiocarbamate, known Cu,Zn SOD inhibitors. Sodium azide and o-phenanthroline demonstrated inhibition at concentrations from 5 to 30 mM, and EDTA also exhibited a varying degree of inhibition of SOD activity. However, there were differences in the nonreduced molecular masses, the reduced molecular masses, and the isoelectric points of the four newly described SODs and the A. fumigatus enzyme; these varied from 55 to 123 kDa, 17.5 to 19.5 kDa, and 5.0 to 5.9, respectively. Of particular note was the observation that the A. fumigatus enzyme was thermostable compared with the SODs from the other species; in addition, the A.fiumigatus enzyme retained all of its activity at 37 degrees C relative to 20 degrees C, whereas the SODs of A. nidulans and A. terreus lost significant activity at the higher temperature. Aspergillus Cu,Zn SOD plays a hypothetical role in the avoidance of oxidative killing mechanisms, and our data suggest that the thermotolerant A. fumigatus Cu,Zn SOD would be more effective in such a protective system than, for example, the equivalent enzyme from the more rarely pathogenic A. nidulans.

Amino Acid Sequence↗
PubMed1996

LINAC-based spinal stereotactic radiosurgery.

The authors' report on the use of a prototype spinal stereotactic radiosurgery frame which was employed for the treatment of 9 patients who presented with recurrent neoplastic involvement of the spinal column. All patients had failed standard therapy consisting of surgery, external fractionated radiation therapy, and/or chemotherapy. Eight of the lesions represented metastatic tumors in the vertebral column, one of the lesions was a primary osteosarcoma involving multiple vertebral bodies. The lesions were found at multiple levels, from the cervical through the sacral region. Six out of the 9 patients presented with epidural compression: 4 of the 9 patients with evidence of myelopathy: 2 of the 9 patients with radicular symptoms secondary to compression from the tumor, and 1 patient was free of any compressive symptoms. All patients had pain requiring narcotics. Patients were treated with a median radiosurgical dose of 800 cGy (range 800-1.000) with a median of 1 isocenter (range 1-7 isocenters) and median normalization of 80% to the isodose contour (range 80-160). Median dose delivered to the already prior irradiated spinal cord was 179 cGy (range 52-320 cGy) with a median spinal cord dose of 34 (range 4-68). To date, there have been three minor complications: one radiation-induced esophagitis which was treated medically: one wound infection, and 1 patient requiring an additional 24 h of hospitalization stay. There have been no major complications. To date, 5 of the 9 patients have died, all from causes unrelated to the spinal radiosurgery. Three out of the 9 patients have been followed for more than 1 year. In all 3, there was radiographic regression of the tumor and epidural compression. In 2 patients, there was histologic confirmation of absence of tumor in the treated site: in 1 patient. no tumor was found at postmortem. 12 months after treatment, when the patient died of unrelated causes. Although the number of patients followed is limited, the phase I study clearly shows the technical feasibility of spinal radiosurgery for the control of metastatic involvement of the vertebral column even in the face of epidural compression.

Adult↗
PubMed1995

The use of gold foil wrapping for radiation protection of the spinal cord for recurrent tumor therapy.

PURPOSE: The development of a technique to provide sufficient radiation protection to previously irradiated spinal cord in such a manner that interstitial brachytherapy can be conducted after resection of a recurrent tumor and decompression of the cord. METHODS AND MATERIALS: A technique was developed that uses multiple layers of gold foil that are applied around the thecal sac and nerve root sleeves to produce an enveloping radiation shield after resection of recurrent tumor. Once the layers of gold foil are in place, interstitial I125 seeds are permanently placed in the bed of the tumor resection to prevent any recurrence from microcellular disease. The technique is described and its application in the case of a 28-year-old with a third recurrence of chondrosarcoma after external fractionated radiation therapy at the second to the fourth thoracic segments is reviewed. RESULTS: This technique has been used in this first patient. An additional tumor dose of 120.0 Gy was delivered to the tumor bed while the spinal cord was calculated to receive only 1% of the dose over the life span of the implant. To date, this dose of radiation has prevented tumor recurrence for more than 18 months of follow-up. CONCLUSION: This technique of multiple layers of gold foil shielding over the spinal cord and nerve roots has the potential to be a useful tool for the shielding of a previously irradiated spinal cord in the setting of resection of recurrent tumor. It may also have a wider application to a number of other radiosensitive tumors where interstitial brachytherapy may be useful to provide additional treatment after external fractionated radiation therapy.

Adult↗
PubMed1995

A prototype device for linear accelerator-based extracranial radiosurgery.

A prototype frame for accurate stereotactic localization and linear accelerator (LINAC)-based treatment of extracranial targets was developed. The ECRSF is designed to employ either spinal or skeletal osseous fixation to immobilize the area of interest and then encircle the targeted region with a traditional orthogonal, three-axis system. A series of experiments (n = 5) with semi-radiolucent calibration targets (n = 15) and computed tomography (CT) scanning using the EC showed that a mean localization error of 0.98 +/- 0.22 mm was obtainable in the last two and most accurate series of experiments with these targets (n = 8). Using the LINAC to irradiate these same targets demonstrated an overall radiation treatment accuracy ranging from 1.4 to 2.0 mm. This discrepancy between localization error and overall radiation treatment error can be explained by a lack of isocentricity of the LINAC treatment which is typically less than 1 mm and can be as low as 0.5 mm. These data demonstrate that extracranial stereotactic radiosurgery is now technically feasible and that the accuracy of such treatment would be acceptable for clinical treatment.

Equipment Design↗
PubMed1995

Specific recognition of purified Cu,Zn superoxide dismutase from Aspergillus fumigatus by immune human sera.

Sera from 22 (84.6%) of 26 patients with confirmed Aspergillus infections recognized a purified Cu,Zn superoxide dismutase from A. fumigatus on Western blots (immunoblots). Sera from 32 (71%) of 45 patients with suspected Aspergillus infections were reactive to the superoxide dismutase. Normal human sera and sera from patients with other infections were unreactive.

Antibodies, Fungal↗
PubMed1995

Preliminary clinical experience with linear accelerator-based spinal stereotactic radiosurgery.

A prototype device called an extracranial stereotactic radiosurgery frame was used to deliver stereotactic radiosurgery, with a modified linear accelerator, to metastatic neoplasms in the cervical, thoracic, and lumbar regions in five patients. In all patients, the neoplasms had failed to respond to spinal cord tolerance doses delivered by standard external fractionated radiation therapy to a median dose of 45 Gy (range, 33-65 Gy/11-30 fractions). The tumors were treated with single-fraction stereotactic radiosurgery with the spinal stereotactic frame for immobilization, localization, and treatment. The median number of isocenters was one (range, one to five) with a median single fraction dose of 10 Gy (range, 8-10 Gy) with median normalization to 80% isodose contour (range, 80-160%). There has been a single complication of esophagitis to date from radiosurgery of a tumor involving the C6-T1 segments; the esophagitis resolved with medical therapy. Median follow-up in this group of patients has been 6 months (range, 1-12 mo). To date, there has been no radiographic or clinical progression of the treated tumor in any patient. Two patients have died from systemic metastatic disease. In the three surviving patients, there has been computed tomographic- or magnetic resonance-documented regression of the treated tumor with a decrease of thecal sac compression with a median follow-up of 6 months (range, 3-14 mo). These five patients represent the first clinical application of stereotactic radiosurgery in the spine. The results suggest that extracranial radiosurgery may be suitable for the treatment of paraspinal neoplasms after external fractionated radiation therapy, even in the face of spinal cord compression.

Adolescent↗
PubMed1995

Continuous regional cerebral cortical blood flow monitoring in head-injured patients.

Continuous regional cerebral cortical blood flow (rCoBF) was monitored with thermal diffusion flowmetry in 56 severely head-injured patients. Adequate, reliable data were accumulated from 37 patients (21 acute subdural hematomas, 10 cerebral contusions, 4 epidural hematomas, and 2 intracerebral hematomas). The thermal sensor was placed at the time of either craniotomy or burr hole placement. In 15 patients, monitoring was initiated within 8 hours of injury. One-third of the comatose patients monitored within 8 hours had rCoBF measurements of 18 ml per 100 g per minute or less, consistent with previous reports of significant ischemia in the early postinjury period. Initial rCoBF measurements were similar in the patients with Glasgow Coma Scale scores of 3 to 7 and in those with scores of 8 or greater. In patients with poor outcomes, rCoBF measurements did not change significantly from initial measurements; however, in those patients who had better outcomes, final rCoBF measurements were higher than initial rCoBF measurements. The patients who had better outcomes experienced normalization of rCoBF during the period of monitoring, and patients with poor outcomes had markedly reduced final rCoBF. These changes were statistically significant. When management was based strictly upon the intracranial pressure, examples of inappropriate treatment were found. For example, hyperemia and increased intracranial pressure treated with mannitol caused further rCoBF increase, and elevated intracranial pressure with low cerebral blood flow treated with hyperventilation increased the severity of ischemia. In 3 (5%) of 56 patients, wound infections developed. Continuous rCoBF monitoring in head-injured patients offers new therapeutic and prognostic insights into their management.

Adolescent↗
PubMed1995

Purification, N-terminal amino acid sequence and partial characterization of a Cu,Zn superoxide dismutase from the pathogenic fungus Aspergillus fumigatus.

A superoxide dismutase (SOD) has been purified to homogeneity from the fungal pathogen Aspergillus fumigatus using a combination of cell homogenization, isoelectric focusing and gel filtration FPLC. The N-terminal amino acid sequence of the purified enzyme demonstrated substantial homology to known Cu,Zn superoxide dismutases for a range of organisms, including Neurospora crassa and Saccharomyces cerevisiae. The enzyme subunit has a pI of 5.9, a relative molecular mass of 19 kDa and a spectral absorbance maximum of 550nm. The non reduced enzyme has a relative molecular mass of 95 kDa. The enzyme remained active after prolonged incubation at 70 degrees C and was pH insensitive in the range 7-11. Potassium cyanide and diethyldithiocarbamate, known Cu,Zn SOD inhibitors, caused inhibition of the purified enzyme at working concentrations of 0.25 mM, whilst sodium azide and o-phenanthroline demonstrated inhibition at higher concentrations (10-30 mM). SOD activity was also detectable in culture filtrate of A. fumigatus. This enzyme may have a potential role as a virulence factor in the avoidance of neutrophil and phagocyte oxidative burst killing mechanisms.

Amino Acid Sequence↗
PubMed1993

Purification of the 115-kilodalton exoantigen of Cryptococcus neoformans and its recognition by immune sera.

A 115-kDa exoantigen produced by Cryptococcus neoformans recognized by the previously described murine monoclonal antibody 7C9 has been purified from culture filtrate by a combination of membrane ultrafiltration, isoelectric focusing, and preparative gel electrophoresis. It is produced in late-log-phase cultures and is present in greater amounts in cultures grown at 25 degrees C than in those grown at 37 degrees C. Recognition of the antigen by 7C9 on immunoblots is abolished by the proteolytic enzymes papain and trypsin. The antigen is a glycoprotein bearing N-linked oligosaccharides, of which mannose is an important constituent. It does not appear to have proteolytic activity and is acidic, with a pI of 3 to 3.2. Its relationship to previously described C. neoformans mannoprotein is unclear since 7C9 shows only very weak cross-reactivity with a purified sample of the latter. Sera from patients infected with C. neoformans exhibited strong recognition of the glycoprotein as shown by immunoenzyme development of Western immunoblots, indicating its possible significance as a marker of disease.

Antibodies, Bacterial↗
PubMed1992

Toxocara canis: monoclonal antibodies to carbohydrate epitopes of secreted (TES) antigens localize to different secretion-related structures in infective larvae.

The major secreted glycoproteins of Toxocara canis larvae appear to be derived from two specialized organs within the nematode organism. Using immunogold electron microscopy, we have analyzed the binding patterns of a panel of monoclonal antibodies (Tcn-1 to Tcn-8) reactive with Toxocara excretory-secretory (TES) antigens. We find, first, that the esophageal gland and lumen are strongly reactive with monoclonals Tcn-4, Tcn-5, and Tcn-8, and because the posterior portion of the gut is closed, we hypothesize that products of this gland are released through the oral aperture. Second, a distinct anti-TES antibody (Tcn-2) localizes solely to the midbody secretory column, which opens onto the cuticle at a secretory pore. Thus, the secretory apparatus is probably functional in this stage of parasite as an important source of TES products. Only one monoclonal, Tcn-7, can bind to both esophageal and secretory structures. In addition, another antibody, Tcn-3, binds both to the epicuticle and to a TES antigen, but our data do not directly determine whether antigens located in the cuticle are subsequently released. Thus there are at least two, and possibly three, independent sources of TES antigens within Toxocara larvae.

Animals↗