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Biomedical subjects

R J Higgins

Publications and source records attributed to R J Higgins.

At least 55 records · Page 3Linked to original sources

The effect of organophosphates on a chicken brain or sea urchin egg kinesin-driven microtubule motility assay.

The effect of neuropathic and non-neuropathic organophosphates (OPs) and acrylamide on an in vitro kinesin-driven microtubule (MT) motility assay was compared. The goal of the study was to determine whether this in vitro assay could confirm that a mechanism of action of neuropathic OPs was to impair kinesin activity and, therefore, possibly fast axonal anterograde transport (FAAT) in vivo. For our study, kinesin from chicken brain (CK) and sea urchin egg (SUK) was initially purified. Western immunoblotting confirmed the close antigenic homology between CK and SUK, using a mouse monoclonal sea urchin kinesin heavy chain-specific antibody (SUK 4). In the presence of microtubules (MTs) and MgATP, both CK- and SUK-driven MT movement was measured using a video-enhanced differential interference contrast microscope system with computer-assisted analysis. Using this assay system, we then tested separately the effect of two neuropathic OPs (diisopropylfluorophosphate (DFP) and phenyl saligenin phosphate (PSP)) and a non-neuropathic OP (paraoxon (PO)) each at a concentration of 10(-2) M at 27 degrees C. Additionally, we tested acrylamide (10(-2) M), since it is one of the best-characterized neurotoxins impairing FAAT in vivo. Our results demonstrated that none of these compounds significantly affected kinesin-driven MT motility in vitro compared to the standard controls. Further, this assay system was thus not able to discriminate between the neuropathic and non-neuropathic effect of these OPs.

Acrylamide↗

In vitro photodynamic effects of lysyl chlorin p6: cell survival, localization and ultrastructural changes.

The in vitro cell survival, localization and ultrastructural changes following irradiation were examined in 9L glioma cells sensitized with a new photosensitizer, lysyl chlorin p6 (LCP). In clonogenic assays, LCP was 10-100-fold more phototoxic than photofrin II on a microgram/mL basis. Lysyl chlorin p6 uptake was blocked when cells were incubated at 2 degrees C. In view of the chemical properties of LCP, this finding indicates that uptake probably occurred through the endocytic pathway. Fluorescence studies showed LCP localized in a region of the endocytic compartment similar in size, shape and distribution to that labeled by lucifer yellow CH (LY), as well as localizing diffusely throughout the perinuclear cytoplasm. Cells stained with both LY and LCP, however, had distinctly separate regions of staining. Lysyl chlorin p6 localization differed from that of fluorescent probes labeling the mitochondria, Golgi apparatus and endoplasmic reticulum. Ultrastructural changes at both 2 and 30 min after laser irradiation were similar. Mitochondria were often condensed or swollen and also had constrictions and cytoplasmic invaginations. The Golgi apparatus, perinuclear space and rough endoplasmic reticulum (RER) were dilated. These data demonstrate that LCP localizes in a portion of the endosomal compartment, but that morphologic damage initially occurs in the mitochondria, Golgi apparatus and RER.

Animals↗

Immunohistochemical reactivity of basal and luminal epithelium-specific cytokeratin antibodies within normal and neoplastic canine mammary glands.

Human basal epithelium (myoepithelium)-specific (312C8-1) and luminal epithelium-specific (13H5) cytokeratin antibodies were applied to frozen sections of normal canine mammary tissues (seven), benign adenomas and hyperplasias (five), mixed tumors (12), and adenocarcinomas (18) to determine if epithelial subsets could be discriminated by the use of an avidin biotin peroxidase complex immunohistochemical procedure. The 312C8-1 and 13H5 antibodies were consistently reactive with basal and luminal epithelium, respectively, in the normal mammary gland (7/7) and in benign adenomas and hyperplasias (5/5). Mixed mammary tumors had similar basal and luminal epithelial reactivity and also had proliferating spindle-shaped stromal cells that were reactive with 312C8-1 (10/12) and 13H5 (4/12). The adenocarcinomas were subclassified into basal, luminal, and basal/luminal on the basis of 312C8-1 reactivity (4/18), 13H5 reactivity (2/18), and dual reactivity with mutually exclusive anatomic distribution (11/18), respectively. Those tumors with dual immunoreactivity were indicative of noninvasive carcinomas. Dogs with neoplasms that were reactive with 312C8-1 and nonreactive with 13H5 had local recurrence or distant metastasis within 2 weeks to 6 months after diagnosis. Other antibodies used for comparison were pan cytokeratin AE1/AE3, actin HHF35, and vimentin. 312C8-1 and 13H5 antibodies are specific for canine mammary basal and luminal epithelium, respectively, and by employing these antibodies, the origin and differentiation of canine mammary neoplasms can be determined more accurately than on the basis of hematoxylin and eosin-stained tissue alone.

Actins↗

A study of radiation necrosis and edema in the canine brain using positron emission tomography and magnetic resonance imaging.

Radiation injury, a major hazard of central nervous system (CNS) radiotherapy, was investigated using sequential studies with positron emission tomography (PET) and magnetic resonance imaging (MRI) in beagle dogs with both helium and neon-ion hemibrain irradiation. All dogs receiving 7.5-11 Gy of neon showed no signs of radiation injury to 3 years after irradiation. Dogs receiving > or = 13 Gy neon or helium succumbed to radiation necrosis and died 21-32 weeks after irradiation. The findings of imaging studies for all dogs who succumbed to radiation necrosis were normal until 3-6 weeks before death. Sequential studies were performed using 0.5 T MRI spin-echo and inversion recovery imaging sequences, and high-resolution (2-3 mm) PET with 18F deoxyglucose and 82Rb. The same axial slices (within 1-2 mm) were imaged repeatedly (weekly) after irradiation until death. The earliest CNS changes were seen as decreased metabolic activity in the cortex of the irradiated hemisphere with PET or an increase in signal intensity in the periventricular white matter on T2-weighted spin-echo imaging on MRI. From the time this increase in signal intensity was first observed, T1 and T2 values increased steadily in both the gray and white matter until death. The changes in white matter were consistently greater than those in gray matter. The results of PET, MRI, and histopathological examinations support the theory that both cellular and vascular mechanisms are involved in radiation necrosis.

Animals↗

Feline spongiform encephalopathy: fibril and PrP studies.

The brains from 18 cats were examined for the presence of the fibrils and modified PrP protein which are molecular diagnostic markers for scrapie-like diseases. Thirteen cats were referred with clinical neurological signs potentially indicative of feline spongiform encephalopathy (FSE). Of these, five had histopathological changes of FSE, five had other lesions of the central nervous system, and in three the brain was normal. The remaining five cats had no clinical neurological signs and were selected as controls. Fibrils and modified PrP protein were found in the brains of the five cats with FSE and in one of the cats with neurological signs but no histopathological changes in the central nervous system. Fibrils were present in the absence of modified PrP in the brains of two cats, one with neurological signs and a histologically confirmed meningioma, and one with no neurological signs and a histologically normal brain.

Animals↗

Effectiveness of a lysyl chlorin p6/chlorin p6 mixture in photodynamic therapy of the subcutaneous 9L glioma in the rat.

A new photosensitizer, LCP, a combination of lysyl chlorin p6 and chlorin p6, was synthesized and tested for effectiveness in photodynamic therapy using s.c. implanted 9L glioma tumors in rats. Tumors were irradiated with 664-nm light 4 h after LCP injection. Mean intratumoral temperature elevations were less than 4 degrees C using a power density of 50 mW/cm2 for 33.3 min (100 J/cm2). Subsequent experiments examining histological changes and tumor regrowth used a power density of 50 mW/cm2 and total energy densities of 25, 50, and 100 J/cm2. Microscopically, an energy density-dependent coagulation necrosis of tumor cells occurred in treated tumors. Long term inhibition of tumor growth was achieved only at an energy density of 100 J/cm2. Side effects of treatment were seen only in the irradiated area and consisted of coagulation necrosis of normal tissues in rats treated at 50 and 100 J/cm2, including severe skin necrosis. Exposure of rats to fluorescent room light did not cause any macroscopically detectable skin damage. Our data indicate that photodynamic destruction of s.c. 9L glioma tumors using LCP as a photosensitizer results in significant tumor growth inhibition and that further study of LCP is warranted.

Animals↗

Neurotoxicity of acute and repeated treatments of tabun, paraoxon, diisopropyl fluorophosphate and isofenphos to the hen.

The neuropathic potential of acute and repeated exposures of the phosphoramidates tabun (GA) and isofenphos (IFP), of diisopropyl fluorophosphate (DFP) and paraoxon (PO) were examined in the hen with treatments for up to 90 days via intramuscular injections of the highest tolerated doses with atropine protection. Plasma acetylcholinesterase (AChE), non-specific butyrylcholinesterase (BChE) and creatine kinase (CK) activities were measured in order to monitor whether the compounds were present at biologically active concentrations. Locomotor behavior was observed and tissues from the peripheral and central nervous systems were examined for signs of organophosphate-induced delayed neuropathy (OPIDN). No behavioral or histological evidence of OPIDN was observed after treatments with GA, IFP, PO, saline or atropine sulfate. DFP-treated birds displayed locomotor and neuropathological signs of OPIDN with a no effect level (NOEL) between 25 and 50 micrograms/kg.

Acetylcholinesterase↗

Brain lesions of naturally occurring pregnancy toxemia of sheep.

The neuropathology and biochemical features of 17 sheep with clinical signs and gross necropsy features of naturally occurring pregnancy toxemia were retrospectively evaluated. The sheep ranged in age from 3 to 6 years and were of seven different breeds and three breed crosses. Thirteen sheep (case Nos. 1-4, 6-9, 11-14, 16) showed astrocytic nuclear swelling, hypertrophy and proliferation, and cerebrocortical neuronal necrosis. Seven of these sheep had Purkinje cell necrosis (case Nos. 2, 3, 6, 11, 12, 14, 16), and seven had vacuolation of cerebral and cerebellar sub-cortical white matter (case Nos. 1-4, 9, 12, 13). The neuropathologic features were similar to those of naturally occurring hypoglycemia of human beings and experimentally induced hypoglycemia of primates and the rat. The lesions seen in the sheep studied may have been caused by cerebral hypoglycemia, but data for blood or cerebral glucose concentrations were not available.

Animals↗

Evaluation of commercially available antibodies to cytokeratin intermediate filaments and laminin in normal cat pinna.

The pattern of distribution of cytokeratin (CK) intermediate filaments can be used to characterize subsets of epithelial tissues. The purpose of the study was to examine the CK expression of feline pinna skin. Six normal feline pinnae were routinely processed in formalin. An immunohistochemical method was used to stain the pinnae with 8 commercially available anti-human CK antibodies (Abs) (PKK1, CAM 5.2, UCD 10/11, 35BH11, 34BE12, AE1/AE3, MAK 6, A575) and an anti-human laminin Ab. All the CK Abs selectively localized to epithelium except 35BH11, which did not react with any part of the pinna. Some epithelial subsets were identified by their unique staining pattern with CK Abs. Basal cells but not suprabasal cells of the epidermis stained with PKK1; basal but not lumenal cells of apocrine glands stained with 34BE12. Apocrine glands stained with all CK Abs except 35BH11. All epithelial structures were stained with A575. Basal lamina of epithelial and mesenchymal tissues was clearly identified by the anti-laminin Ab. The results indicate that in cat pinna some commercially available anti-human CK Abs selectively stain subsets of epithelium and adnexa. PKK1, 34BE12, and A575 were the CK Abs with the most consistent staining patterns, the other Abs stained more variably from pinna to pinna. The pattern of epithelial and adnexal staining was similar but not identical to that reported for humans.

Animals↗

DNA flow cytometric study of the hyperplastic and neoplastic canine prostate.

Flow cytometric DNA measurements were carried out on 45 formalin-fixed, paraffin-embedded canine prostate tissues. The tissues were categorized as normal, hyperplastic, or neoplastic on the basis of light microscopic examination, and DNA ploidy was compared with histologic classification. Ten normal prostate samples showed diploid DNA histograms, but with proliferation greater than that found in the normal human prostate. Thirteen specimens of benign prostate hyperplasia showed diploid or near-diploid DNA histograms. Of 22 prostatic carcinomas, 12 were diploid and 10 were aneuploid, with the majority of the aneuploidy being near-triploid. The frequency of DNA aneuploidy recognized in canine prostatic carcinoma is similar to findings in human prostatic carcinoma if all their grades of malignancy are included.

Aneuploidy↗

Type C retroviral expression in spontaneous feline olfactory neuroblastomas.

Three cases of spontaneous olfactory neuroblastoma (ONB) in domestic cats were morphologically and immunocytochemically characterized. Diagnostic light microscopic features included Flexner and Homer-Wright rosettes, while ultrastructurally the cells had neuritic processes, intracellular intermediate filaments, and intercellular junctions. Immunocytochemically, the tumors stained positively for neuron-specific enolase, cytokeratins, and S-100 protein antigens. In each case, a key finding was the identification of numerous mature type C retroviral particles within the tumors. In one case, budding of viral particles from the plasmalemma of tumor cells suggested the source of mature particles. This cat and one other were tested, and both were serologically positive for feline leukemia virus (FeLV). The virus in the tumors was identified as FeLV by polymerase chain reaction and immunocytochemistry. No other neoplasms were found in any of the cats, nor was there similar evidence of active viral infection in other non-tumor tissues, including the brain. Although the relationship between FeLV infection and ONB is uncertain, our findings indicate that FeLV should be investigated as an etiologic agent of ONB.

Animals↗

Activation of retrovirus in transgenic mice: association with development of olfactory neuroblastoma.

A line of transgenic mice that express the human adenovirus type 12 E1A and E1B genes under the regulatory control of the mouse mammary tumor virus long terminal repeat was studied. Mice from this line develop olfactory neuroblastomas at approximately 6 months of age. Large numbers of type C retrovirus (ecotropic murine leukemia virus) particles were found in the tumor rosettes. No similar examples of virus activation were identified in tumors from other transgenic experiments. Examination of spontaneous olfactory neuroblastomas from three domestic cats also demonstrated retrovirus in tumor rosettes.

Adenovirus Early Proteins↗

Bovine spongiform encephalopathy: diagnostic significance of vacuolar changes in selected nuclei of the medulla oblongata.

The adequacy of a histopathological diagnosis of bovine spongiform encephalopathy (BSE) based exclusively on observations of neuroparenchymal vacuolation in three specific neuroanatomic nuclei was tested by using a standard coronal section of medulla oblongata cut at the obex. The agreement between the observations and the definitive histopathological diagnosis was assessed in each of 684 bovine brains - 563 confirmed cases of BSE, 20 with changes which did not diagnose BSE conclusively and 101 in which the lesions of BSE were not detected. When the assessment was confined to the solitary tract nucleus and the spinal tract nucleus of the trigeminal nerve a positive result was obtained in 99.6 per cent of confirmed cases of BSE and only 1 per cent of brains in which lesions of BSE were not detected gave a false positive result. Thus an initial examination of the single section, together with an examination of representative areas of the rest of the brain when no unequivocal lesion was found, provided a satisfactory method for the routine diagnosis of BSE.

Animals↗