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Biomedical subjects

M T Campbell

Publications and source records attributed to M T Campbell.

17 recordsLinked to original sources
PubMed1998

A method for purification and characterisation of Mycobacterium avium subsp. paratuberculosis from the intestinal mucosa of sheep with Johne's disease.

Mycobacterium avium subsp. paratuberculosis, the cause of Johne's disease in ruminants, cannot be cultured in large quantities from affected sheep in Australia. A method is described for the purification of the organism from the intestinal mucosa of sheep with multibacillary Johne's disease in order to undertake restriction fragment length polymorphism (RFLP) analysis for epidemiological purposes. Using sucrose and potassium chloride as separation media for differential and density gradient centrifugation, yields of approximately 90 mg dry weight M. avium subsp. paratuberculosis per 5 g intestinal mucosa were obtained. The preparations of purified M. avium subsp. paratuberculosis were visually free of non-acid fast bacteria and contained 10(2)-10(3) aerobic/ facultatively anaerobic organisms per gram wet weight. DNA extracted from purified M. avium subsp. paratuberculosis was examined by hybridisation with an IS900 probe after digestion with BstEII and RFLP patterns distinct from isolates from cattle were obtained. The RFLP pattern of purified M. avium subsp. paratuberculosis from five sheep matched that obtained previously from organisms cultured from sheep in studies in New Zealand, indicating that the purification and RFLP method is robust.

Animals↗
PubMed1993

Analysis of the sheep trichohyalin gene: potential structural and calcium-binding roles of trichohyalin in the hair follicle.

Trichohyalin is a structural protein that is produced and retained in the cells of the inner root sheath and medulla of the hair follicle. The gene for sheep trichohyalin has been purified and the complete amino acid sequence of trichohyalin determined in an attempt to increase the understanding of the structure and function of this protein in the filamentous network of the hardened inner root sheath cells. Sheep trichohyalin has a molecular weight of 201,172 and is characterized by the presence of a high proportion of glutamate, arginine, glutamine, and leucine residues, together totaling more than 75% of the amino acids. Over 65% of trichohyalin consists of two sets of tandem peptide repeats which are based on two different consensus sequences. Trichohyalin is predicted to form an elongated alpha-helical rod structure but does not contain the sequences required for the formation of intermediate filaments. The amino terminus of trichohyalin contains two EF hand calcium-binding domains indicating that trichohyalin plays more than a structural role within the hair follicle. In situ hybridization studies have shown that trichohyalin is expressed in the epithelia of the tongue, hoof, and rumen as well as in the inner root sheath and medulla of the hair follicle.

Amino Acid Sequence↗
PubMed1992

Differential lectin binding to presumptive cortical cells of the wool follicle bulb.

An alpha-D-galactoside-specific lectin from Bandeiraea simplicifolia (BSLI) showed differential binding to cortical cells of the wool follicle bulb. The lectin bound to cells on one side only of the bulb and was completely blocked by alpha-D-galactose. The region of lectin binding extended from presumptive cortical cells at the base of the bulb to cortical cells at the top of the bulb, disappearing as cortical cells entered the fibre cortex. An orthocortex-specific monoclonal antibody was used to show that cortical cells recognised by the lectin lie directly below the fibre orthocortex and presumably give rise to the orthocortex. The results suggest that two distinct populations of presumptive cortical cells are present only two to three cell layers from the base of the bulb in a region where no morphological differences are detectable. The lectin-bound pre-cortical cells appear to give rise to orthocortical cells while cells not bound by lectin presumably give rise to paracortical cells. Electron microscopy showed that the lectin bound to sites on the plasma membrane, probably on the extracellular surface. This suggests that the lectin target may be a membrane glycoprotein or glycolipid. Two polypeptides recognised by BSLI were separated from wool follicle extracts by SDS-gel electrophoresis. These polypeptides migrated at approximately 69 kDa and 17 kDa. However, only the 69 kDa molecule showed the expected loss of binding by BSLI in the presence of alpha-D-galactose. Further work will be required to determine if this glycoprotein is the bulb cell molecule recognised by BSLI.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

A lens fibre differentiation factor from calf neural retina.

During growth of the eye lens, epithelial cells differentiate into fibre cells under the influence of neural retina. The fibre differentiation factor (FDF) was partially characterized from calf retina-conditioned medium, using lens epithelial explants from young rats, to provide a bioassay for differentiation. FDF was associated with large-protein aggregates, the smallest of which eluted at approximately 500-600 kD on Sephacryl S-300 columns and migrated as a single protein band near 600 kD on gradient gels. This protein resolved into nine major peptides on SDS-polyacrylamide gels, ranging between 23 and 27 kD. Eight of these peptides were present oa four doublets, but did not appear to contain specific carbohydrate residues. The approximately 500-600 kD complex could be slightly disrupted by trypsin or heat treatment to release a less stable 90 kD component. Fractionation of FDF invariably led to loss of activity, possibly due to gradual dissociation into less active and/or less stable components. A working hypothesis suggested by these findings is that FDF is associated with a small group of peptides, each contributing an essential function to the process of fibre differentiation.

Animals↗
PubMed1985

Influence of neural retina on lens fibre differentiation in rats.

Lens epithelial explants grown in retina-conditioned medium (RCM) undergo structural and molecular changes characteristic of fibre differentiation in the intact lens. We suggest that in vivo neural retina releases a fibre differentiation factor (FDF) that interacts with equatorial lens epithelial cells and stimulates them to undergo fibre cell differentiation. According to this model, interaction with neural retina is essential for normal lens formation in embryos and for normal lens growth throughout life. Preliminary work on purification of the factor indicates that FDF activity is associated with a high molecular weight complex of 500 kd. The active component of this complex appears to be an 80 kd molecule.

Animals↗
PubMed1984

Onset of fibre differentiation in cultured rat lens epithelium under the influence of neural retina-conditioned medium.

In the intact rat lens, epithelial cells, which are cuboidal and contain alpha-crystallin, give rise to fibre cells, which are elongated and contain alpha-, beta- and gamma-crystallins. Epithelial explants cultured in medium conditioned by bovine neural retinas (BRCM) showed changes analogous to fibre cell differentiation in vivo. The cells became enlarged and elongated and accumulated beta- and gamma-crystallin as well as alpha-crystallin. Labelling studies with [35S]-methionine showed that sequential changes in the synthesis of all three classes of crystallin occurred during culture in BRCM. After two days, synthesis of alpha-crystallin subunits, particularly alpha A and alpha AINS, increased relative to overall protein synthesis. After four days in culture, synthesis of beta-crystallin subunits, identified as beta B1a, beta B4, beta B5 and possibly beta A2, was detected for the first time and between four and eight days gamma-crystallin synthesis became detectable. The time of onset of gamma-crystallin synthesis seemed to show greater experimental variability than did onset of beta-crystallin synthesis. In explants cultured for 10 days in BRCM approximately 25% of new protein synthesis could be attributed to alpha-, beta- and gamma-crystallins. These events were completely dependent on BRCM, suggesting that neural retina secretes a factor(s) which initiates fibre cell differentiation. This culture system appears to be a suitable one for investigating the control of fibre differentiation in vitro.

Animals↗
PubMed1982

The import of carbamoyl-phosphate synthase into mitochondria from foetal rat liver.

A putative precursor of carbamoyl-phosphate synthase was isolated from a microsomal wash fraction and purified by high-pressure liquid chromatography. Autolytic degradation and limited proteolysis were used to characterize the putative precursor of carbamoyl-phosphate synthase and to show its similarity to the processed enzyme. The carbamoyl-phosphate synthase precursor underwent a time-dependent and concentration-dependent conversion into a dimeric or polymeric form. When labelled with 125I and incubated with foetal rat liver mitochondria the precursor was bound to the mitochondria and about 30% of the label was imported into the matrix space. This labelling required the presence of ATP and was time-dependent. Mitoplasts also imported the carbamoyl-phosphate synthase precursor. After import of the precursor, increases in carbamoyl-phosphate synthase activity could be demonstrated in foetal rat liver mitochondria.

Adenosine Triphosphate↗
PubMed1980

The effect of premature and delayed birth on the development of UDP-glucuronosyltransferase activities towards bilirubin, morphine and testosterone in the rat.

In the rat, UDP-glucuronosyltransferase activities towards bilirubin, morphine and testosterone increase markedly after normal or premature birth. This rapid development is superimposed upon a much slower maturation of activity which occurs in utero during the last 2 days of normal gestation and gestation when birth is delayed. Development of all three activities is similar under these different conditions, suggesting a common developmenpal regulatory mechanism.

Animals↗
PubMed1979

The formation and distribution of bilirubin monoglucuronide and diglucuronide in rat liver slices.

1. Bilirubin conjugation in rat liver slices was reassessed by using analysis of ethyl anthranilate azopigments to estimate separately the formation of bilirubin mono- and di-glucuronides. 2. Conjugation in slices resembles the situation in vivo more closely than does microsomal conjugation, in that diglucuronide is formed in appreciable quantity. 3. Both bilirubin mono- and di-glucuronides were present in slices in approximately equal amounts, but the monoglucuronide was the major product found in the incubation medium. 4. These results are discussed in relation to recent theories on the relationship between bilirubin mono- and di-glucuronide formation in vivo.

Animals↗
PubMed1979

Demonstration of two functionally heterogenous groups within the activities of UDP-glucuronosyltransferase towards a series of 4-alkyl-substituted phenols.

1. A simple colorimetric assay for UDP-glucuronosyltransferase activities towards phenolic substrates, using Folin & Ciocalteu's phenol reagent, is described. The assay is used to measure rat liver transferase activities towards substrates from a series of 4-alkyl-substituted phenols. 2. Activities towards phenol, 4-methylphenol and 4-ethylphenol develop near-adult values before birth, are precociously stimulated by dexa methasone in utero and are stimulated 3--4-fold by 3-methylcholanthrene in adult liver. These are assigned to a "late-foetal" group of transferase activities. 3. Activities towards 4-n-propylphenol, 4-s-butylphenol and 4-t-butylphenol are negligible in late-foetal liver, developing to near-adult values in the first 4 postnatal days, and are not affected by dexamethasone or 3-methylcholanthrene. They are assigned to a "neonatal" group of transferase activities. 4. Although 4-ethylphenol and 4-n-propylphenol differ only by a single --CH2-- moiety, this is sufficient to change the acceptability of these substrates respectively from the late-foetal to the neonatal group of transferase activities. The change is distinct, with no overlapping of substrate acceptability between the two groups of transferase activities. 5. From consideration of the above and other substrates, the two groups of transferase activities do not distinguish substrates on the basis of their molecular weights or lipophilicity. The distinguishing feature appears to be the specific molecular configurations of the substrates.

Animals↗
PubMed1977

Synthesis and antiviral activity of certain thiazole C-nucleosides.

A general reaction of glycosyl cyanides with liquid hydrogen sulfide in the presence of 4-dimethylaminopyridine to provide the corresponding glycosylthiocarboxamides is described. These glycosylthiocarboxamides were utilized as the precursors for the synthesis of 2-D-ribofuranosylthiazole-4-carboxamide and 2-beta-D-ribofuranosylthiazole-5-carboxamide (23). The structural modification of 2-beta-D-ribofuranosylthiazole-4-carboxamide (12) into 2-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)thiazole-4-carboxamide (15), 2-beta-D-ribofuranosylthiazole-4-thiocarboxamide (17), and 2-(5-deoxy-beta-D-ribofuranosyl)thiazole-4-carboxamide (19) is also described. These thiazole nucleosides were tested for in vitro activity against type 1 herpes virus, type 3 parainfluenza virus, and type 13 rhinovirus and an in vivo experiment was run against parainfluenza virus. They were also evaluated as potential inhibitors of purine nucleotide biosynthesis. It was shown that the compounds (12 and 15) which possessed the most significant antiviral activity were also active inhibitors (40-70%) of guanine nucleotide biosynthesis.

Animals↗