Thesis--University of Wisconsin--Madison. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 81-96).

The single residue of cysteine in Cl. botulinum type E strain Iwanai toxin has been linked with toxicity, by chemical modification using p-chloromercuribenzoate. A peptide containing the oysteine residue has been isolated by exhaustive tryptic digestion of the toxin molecule tagged with N-(4-dimethylamino-3,5-dinitrophenyl) maleimide, and subsequent gel filtration with Sephadex G-25 and descending paper chromatography. The toxic peptide of trypsin-activated toxin was isolated by fractionation through a composite Sephadex G-75 and G-50 column. By chymotryptic and tryptic digestion of the toxin at pH 5.8, a toxic fragment has been isolated by gel filtration with Sephadex G-25. On the basis of quantitative amino acid analyses, the molecular weights of the intact toxin, the trypsin-activated toxin and the chymotrypsin-trypsin fragmented toxin have been estimated to be 14,000-16,000, 10,000-12,000 and 4,000-6,000 respectively. Although the mechanism of tryptic activation was found to involve chiefly the removal of at least 18 amino acid residues from the N-terminus of the toxin molecule, the manner of reduction by cleavage has not been determined for the chymotrypsin-trypsin fragmented toxin. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

Botulism is a foodborne intoxication caused by ingestion of Clostridium botulinum neurotoxin (BoNT). Preliminary studies focussed on the production of polyclonal antisera against BoNT/E by immunizing a rabbit with botulinal toxoid type E. The antiserum was subsequently used to detect BoNT/E using the slot blot immunoassay where samples were applied to a slot blot filtration manifold and drawn by vacuum through a membrane. The membrane was then immunologically processed before chemiluminescent detection. However, the antisera lacked specificity and cross-reacted with closely related clostridia strains. / The specificity of the antisera was increased by adsorbing cross-reactive antibodies from whole antisera with affinity columns made with total proteins from culture supernatants of closely related clostridia. Alternatively, specific antibodies were isolated with an affinity column prepared with C. botulinum type E toxoid. / Different methods of concentrating BoNT/E in each sample prior to testing them were evaluated to increase the sensitivity of the assay. / The slot blot immunoassay was then evaluated for detection of BoNT/E in mixed cultures and in food samples. (Abstract shortened by UMI.)

Thesis (M.S.)--University of Michigan, 2005. / Includes bibliographical references.

No description available.

Intraprostatic injection of BTX-A has demonstrated clinical improvement in men with bladder out let obstruction. Firstly, we investigated the mechanisms of action of BTX-A on the prostate. Secondly, an animal model for nonbacterial prostatitis in rats was developed using intraprostatic injection of capsaicin, an agent thought to excite c-afferent fibers and cause neurogenic inflammation. The analgesic and anti-inflammatory properties of Botulinum toxin type A (BoNT-A) was tested in this model. (1) Adult male Spragu-Dawley rats were injected with varying doses of BTX-A into the prostate, and the prostates were harvested after 1 or 2 weeks. The effects of BTX-A on prostate histology, and the proliferative and apoptotic indexes were determined using hematoxylin and eosin staining, proliferative cell nuclear antigen staining and TUNEL staining, respectively. Changes in a1A adrenergic receptor and androgen receptor were evaluated by Western blotting. (2) Adult male Spragu-Dawley rats were injected with varying doses of capsaicin into the prostate. The nociceptive effects of capsaicin were evaluated for 30 min by using a behavior approach; the prostate was removed for histology and cyclooxygenase (COX) 2 concentration measurement. Evans blue (50mg/kg) was also injected intravenously to assess for plasma protein extravasation. The other set of animals were injected with up to 20U of BoNT-A into the prostates 1 wk prior to intraprostatic injection of 1000umol/l capsaicin. (1) One week after BTX-A injection generalized prostate atrophy was observed. There was a significant increase in apoptotic cells (12, 16 and 22-fold), and decrease in proliferative cells (38%, 77% and 80%) and a1A adrenergic receptor (13%, 80% and 81%) for 5U, 10U and 20U, respectively. There was no significant change in androgen receptors. The effects were decreased 2 weeks after BTX-A treatment. (2) Capsaicin dose dependently induced modifications in pain behavior closing of the eyes, hypolocomotion, and inflammatory changes: increase of inflammatory cell accumulation, COX2 expression, and plasma extravasation at the acute stage, but completely recovered at 1 wk. BoNT-A pretreatment dose dependently reversed pain behavior and inflammation. BoNT-A 20U significantly decreased inflammatory cell accumulation, COX2 expression, and Evans blue extraction (82.1%, 93.0% and 50.4, respectively), and reduced pain behavior (66.% for eye score and 46.5% for locomotion score). Conclusion (1): BTX-A injection into the prostate alters cellular dynamics by inducing apoptosis, inhibiting proliferation and down-regulating a1A adrenergic receptors. BTX-A may potentially be the drug that has dual actions on the static and dynamic components of benign prostatic hyperplasia. (2): Intraprostatic capsaicin injection induced neurogenic prostatitis and prostatic pain, and may be a useful research model. BoNT-A produced anti-inflammatory and analgesic effects, and support clinical evaluation in prostatitis.

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Thesis (M.S.)--Virginia Polytechnic Institute and State University, 1988. / Vita. Abstract. Includes bibliographical references (leaves 88-100). Also available via the Internet.