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Biomedical subjects

D J Ferguson

Publications and source records attributed to D J Ferguson.

At least 37 records · Page 2Linked to original sources
PubMed2000

Reconstitution of dimethylamine:coenzyme M methyl transfer with a discrete corrinoid protein and two methyltransferases purified from Methanosarcina barkeri.

Methyl transfer from dimethylamine to coenzyme M was reconstituted in vitro for the first time using only highly purified proteins. These proteins isolated from Methanosarcina barkeri included the previously unidentified corrinoid protein MtbC, which copurified with MtbA, the methylcorrinoid:Coenzyme M methyltransferase specific for methanogenesis from methylamines. MtbC binds 1.0 mol of corrinoid cofactor/mol of 24-kDa polypeptide and stimulated dimethylamine:coenzyme M methyl transfer 3.4-fold in a cell extract. Purified MtbC and MtbA were used to assay and purify a dimethylamine:corrinoid methyltransferase, MtbB1. MtbB1 is a 230-kDa protein composed of 51-kDa subunits that do not possess a corrinoid prosthetic group. Purified MtbB1, MtbC, and MtbA were the sole protein requirements for in vitro dimethylamine:coenzyme M methyl transfer. An MtbB1:MtbC ratio of 1 was optimal for coenzyme M methylation with dimethylamine. MtbB1 methylated either corrinoid bound to MtbC or free cob(I)alamin with dimethylamine, indicating MtbB1 carries an active site for dimethylamine demethylation and corrinoid methylation. Experiments in which different proteins of the resolved monomethylamine:coenzyme M methyl transfer reaction replaced proteins involved in dimethylamine:coenzyme M methyl transfer indicated high specificity of MtbB1 and MtbC in dimethylamine:coenzyme M methyl transfer activity. These results indicate MtbB1 demethylates dimethylamine and specifically methylates the corrinoid prosthetic group of MtbC, which is subsequently demethylated by MtbA to methylate coenzyme M during methanogenesis from dimethylamine.

Dimethylamines↗
PubMed2000

Toxoplasma gondii: identification of a developmentally regulated family of genes related to SAG2.

Previous studies have shown the surface of Toxoplasma gondii to be dominated by a family of proteins closely related to SAG1. In this study, we report the existence of a second family of genes defined by homology to SAG2. The predicted amino acid sequences of these three new proteins suggests that they are all glycosylphosphatidylinositol-linked surface antigens. All three also contain N-linked glycosylation sites, although their use has yet to be demonstrated. One of these SAG2-related antigens, SAG2B, is expressed in tachyzoites with an apparent size of 23 kDa. It is distinct, however, from the previously identified P23. In contrast to SAG2B, SAG2C and SAG2D appear to be expressed exclusively on the surface of bradyzoites. Analysis of the SAG2 family shows it to have weak but significant homology to the SAG1 family. Thus, all of the sequenced surface antigens of tachyzoites and many of those of bradyzoites fall into one large superfamily that can be divided into two subgroups defined by the prototypic and highly immunogenic SAG1 and SAG2, respectively.

Amino Acid Sequence↗
PubMed2000

Should internal mammary nodes be sampled in the sentinel lymph node era?

BACKGROUND: Controversy exists regarding internal mammary lymph nodes (IMNs) in the staging and treatment of breast cancer. Sentinel lymph node identification with radiocolloid can map drainage to IMNs and directed biopsy can be performed with minimal morbidity. Furthermore, recent studies suggest that IMN drainage of breast tumors may be underestimated. To gain further insight into the prognostic value of IMNs, we reviewed the outcome of patients in whom the IMN status was routinely assessed. METHODS: A retrospective review of 286 patients with breast cancer who underwent IMN dissection between 1956 and 1987 was conducted. RESULTS: Median follow-up is 186 months, age was 52 years (range, 21-85 years), tumor size was 2.5 cm, and number of IMNs removed was 5 (range, 1-22); 44% received chemotherapy, 16% endocrine therapy, and 5% radiotherapy. Presence of IMN metastases correlated with primary tumor size (P < .0001) and number of positive axillary nodes (P < .0001) but did not correlate with primary tumor location or age. Overall, the 20-year disease-free survival is significantly worse for the 25% of patients with IMN metastases (P < .0001). In patients with positive axillary nodes and tumors smaller than 2 cm, there was a significantly worse survival (P < .0001) in the patients with IMN metastases. This difference in survival was not seen in women with tumors larger than 2 cm. CONCLUSIONS: Patients with IMN metastases, regardless of axillary node status, have a highly significant decrease in 20-year disease-free survival. Treatment strategies based on knowledge of sentinel IMN status may lead to improvement in survival, especially for patients with small tumors. At present, sentinel IMN biopsies should be performed in a clinical trial setting.

Adult↗
PubMed2000

The microneme protein MIC4, or an MIC4-like protein, is expressed within the macrogamete and associated with oocyst wall formation in Toxoplasma gondii.

The expression and localisation of MIC4, or an immuno-cross reacting MIC4-like protein, was examined in the enteric forms of Toxoplasma gondii using immunocytochemistry. In addition to being located within the micronemes of the merozoites, MIC4 or the MIC4-like protein was present within the macrogamete and was associated with the developing oocyst wall. The macrogamete is characterised by two types of structurally distinct wall forming bodies (WFB1 and 2). However, by immuno-electron microscopy, it was possible to identify two populations of dense granules (WFB1) which appear to form sequentially during macrogamete development. The first granules to form (WFB1a) stained positively with anti-MIC4 and were followed by MIC4 negative granules (WFB1b). During oocyst wall formation, the WFB1a and b sequentially released their contents onto the surface with WFB1a material forming an anti-MIC4 positive outer veil, while the WFB1b forms the electron dense outer layer of the oocyst wall. The inner layer was formed by WFB2. Thus, for the first time, it was possible to identify two populations of dense granules (WFB1a and b) involved in the formation of different parts of the oocyst wall. It was not possible to analyse the contents of macrogametes by western blot to unequivocally identify the antigen recognised by the polyclonal antisera as MIC4.

Animals↗
PubMed2000

Inhibition of nitric oxide production exacerbates chronic ocular toxoplasmosis.

There is considerable controversy as to the roles of parasite proliferation and the inflammatory response in destruction of the retina during Toxoplasma gondii infection. A murine model was used to investigate the role of nitric oxide in pathogenesis of chronic ocular toxoplasmosis. Increased quantities of messenger RNA (mRNA) transcripts for iNOS were detected in the eyes of chronically infected C57BL/6 mice compared with noninfected control mice. Inhibition of nitric oxide (NO) by the addition of Lomega-nitro-L-arginine methyl ester (L-NAME) to the drinking water of infected mice between weeks 4-6 of infection, exacerbated ocular inflammation. The amount of inflammation was assessed semiquantitatively in histological sections of the eye. Eyes from L-NAME treated mice showed a significant increase in inflammation of the retina (P = 0.02), choroid (P = 0.03), and vitreous (P = 0.02) compared with control mice. These results demonstrate a protective role for NO in the control of chronic, ocular toxoplasmosis.

Animals↗
PubMed2000

What is normal black African hair? A light and scanning electron-microscopic study.

BACKGROUND: The hair of normal black Africans forms a mat of tightly interwoven hair shafts. The effect of this on the structure of the hair shaft and the response to grooming is unknown. OBJECTIVE: Our purpose was to use light and scanning electron microscopy (SEM) to examine the structure of Negroid-type hairs and effects of combing in black African volunteers. METHODS: Hair samples were collected, by combing, from Africans and compared with those from Caucasian and Asian volunteers. The volunteers had never used chemical treatments. Their hair had not been cut for at least 1 year and grooming had been limited to shampooing, drying, and combing. RESULTS: More than 2000 hairs in 12 African volunteers were examined by light microscopy. The hairs appear as a tight coiled springlike structure. Many shafts exhibited knots (10%-16% vs 0.15%) and appear broken compared with hair shafts from other ethnic groups. SEM of African hairs showed features consistent with repeated breaks of the shaft. Examination of hairs in situ showed interlocking of hair shafts. CONCLUSION: These observations provide an understanding of the physical nature of, and effect of combing on, black African hair.

Adolescent↗
PubMed2000

Developmental expression of a tandemly repeated, glycine- and serine-rich spore wall protein in the microsporidian pathogen Encephalitozoon cuniculi.

Microsporidia are intracellular organisms of increasing importance as opportunistic pathogens in immunocompromised patients. Host cells are infected by the extrusion and injection of polar tubes located within spores. The spore is surrounded by a rigid spore wall which, in addition to providing mechanical resistance, might be involved in host cell recognition and initiation of the infection process. A 51-kDa outer spore wall protein was identified in Encephalitozoon cuniculi with the aid of a monoclonal antibody, and the corresponding gene, SWP1, was cloned by immunoscreening of a cDNA expression library. The cDNA encodes a protein of 450 amino acids which displays no significant similarities to known proteins in databases. The carboxy-terminal region consists of five tandemly arranged glycine- and serine-rich repetitive elements. SWP1 is a single-copy gene that is also present in the genomes of Encephalitozoon intestinalis and Encephalitozoon hellem as demonstrated by Southern analysis. Indirect immunofluorescence and immunoelectron microscopy revealed that SWP1 is differentially expressed during the infection cycle. The protein is absent in replicative meronts until 24 h postinfection, and its expression is first induced in early sporonts at a time when organisms translocate from the periphery to the center of the parasitophorous vacuole. Expression of SWP1 appears to be regulated at the mRNA level, as was shown by reverse transcriptase PCR analysis. Further identification and characterization of stage-specific genes might help to unravel the complex intracellular differentiation process of microsporidia.

Amino Acid Sequence↗
PubMed2000

The trimethylamine methyltransferase gene and multiple dimethylamine methyltransferase genes of Methanosarcina barkeri contain in-frame and read-through amber codons.

Three different methyltransferases initiate methanogenesis from trimethylamine (TMA), dimethylamine (DMA) or monomethylamine (MMA) by methylating different cognate corrinoid proteins that are subsequently used to methylate coenzyme M (CoM). Here, genes encoding the DMA and TMA methyltransferases are characterized for the first time. A single copy of mttB, the TMA methyltransferase gene, was cotranscribed with a copy of the DMA methyltransferase gene, mtbB1. However, two other nearly identical copies of mtbB1, designated mtbB2 and mtbB3, were also found in the genome. A 6.8-kb transcript was detected with probes to mttB and mtbB1, as well as to mtbC and mttC, encoding the cognate corrinoid proteins for DMA:CoM and TMA:CoM methyl transfer, respectively, and with probes to mttP, encoding a putative membrane protein which might function as a methylamine permease. These results indicate that these genes, found on the chromosome in the order mtbC, mttB, mttC, mttP, and mtbB1, form a single transcriptional unit. A transcriptional start site was detected 303 or 304 bp upstream of the translational start of mtbC. The MMA, DMA, and TMA methyltransferases are not homologs; however, like the MMA methyltransferase gene, the genes encoding the DMA and TMA methyltransferases each contain a single in-frame amber codon. Each of the three DMA methyltransferase gene copies from Methanosarcina barkeri contained an amber codon at the same position, followed by a downstream UAA or UGA codon. The C-terminal residues of DMA methyltransferase purified from TMA-grown cells matched the residues predicted for the gene products of mtbB1, mtbB2, or mtbB3 if termination occurred at the UAA or UGA codon rather than the in-frame amber codon. The mttB gene from Methanosarcina thermophila contained a UAG codon at the same position as the M. barkeri mttB gene. The UAG codon is also present in mttB transcripts. Thus, the genes encoding the three types of methyltransferases that initiate methanogenesis from methylamine contain in-frame amber codons that are suppressed during expression of the characterized methyltransferases.

Amino Acid Sequence↗
PubMed2000

Ovine male genital duct epithelial cells differentiate in vitro and express functional CFTR and ENaC.

To investigate the biology of the male genital duct epithelium, we have established cell cultures from the ovine vas deferens and epididymis epithelium. These cells develop tight junctions, high transepithelial electrical resistance, and a lumen-negative transepithelial potential difference as a sign of active transepithelial ion transport. In epididymis cultures the equivalent short-circuit current (I(sc)) averaged 20.8+/-0.7 microA/cm(2) (n = 150) and was partially inhibited by apical application of amiloride with an inhibitor concentration of 0.64 microM. In vas deferens cultures, I(sc) averaged 14.4+/-1.1 microA/cm(2) (n = 18) and was also inhibited by apical application of amiloride with a half-maximal inhibitor concentration (K(i)) of 0.68 microM. The remaining amiloride-insensitive I(sc) component in epididymis and vas deferens cells was partially inhibited by apical application of the Cl(-) channel blocker diphenylamine-2-carboxylic acid (1 mM). It was largely dependent on extracellular Cl(-) and, to a lesser extent, on extracellular HCO(-)(3). It was further stimulated by basolateral application of forskolin (10(-5) M), which increased I(sc) by 3.1+/-0.3 microA/cm(2) (n = 65) in epididymis and 0.9+/-0.1 microA/cm(2) (n = 11) in vas deferens. These findings suggest that cultured ovine vas deferens and epididymis cells absorb Na(+) via amiloride-sensitive epithelial Na(+) channels (ENaC) and secrete Cl(-) and HCO(-)(3) via apical cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channels. This interpretation is supported by RT-PCR data showing that vas deferens and epididymis cells express CFTR and ENaC mRNA.

Amiloride↗
PubMed2000

Rethinking knowledge and pedagogy in dental education.

Dentistry as a profession has often been considered both art and science. Traditional dental education has attempted to address both; however, in many places only the science of dentistry is emphasized. The move toward competency-based curricula in dental education requires an expansion of what constitutes meaningful knowledge in the curriculum and what pedagogies best support that curriculum. The scientific and technical knowledge considered foundational to clinical practice is not sufficient to teach competencies associated with the art of dentistry. Habermas, a social scientist, offers a way of looking beyond technical knowledge to consider two other forms of knowledge: practical and emancipatory. Pedagogy that supports development of practical and emancipatory knowledge includes problem-based learning and case methods, heuristics, reflective practica, journals, storytelling, and performance-based assessment methods. These important teaching strategies are being integrated into various dental curricula including a new competency-based dental curriculum at Marquette University's School of Dentistry. It will be critical for dental educators to continue developing these methods to provide efficient and effective education for future practitioners in both the art and science of dentistry.

Clinical Clerkship↗
PubMed1999

Plasmodium falciparum-infected erythrocytes modulate the maturation of dendritic cells.

The malaria parasite Plasmodium falciparum is one of the most successful human pathogens. Specific virulence factors remain poorly defined, although the adhesion of infected erythrocytes to the venular endothelium has been associated with some of the syndromes of severe disease. Immune responses cannot prevent the development of symptomatic infections throughout life, and clinical immunity to the disease develops only slowly during childhood. An understanding of the obstacles to the development of protective immunity is crucial for developing rational approaches to prevent the disease. Here we show that intact malaria-infected erythrocytes adhere to dendritic cells, inhibit the maturation of dendritic cells and subsequently reduce their capacity to stimulate T cells. These data demonstrate both a novel mechanism by which malaria parasites induce immune dysregulation and a functional role beyond endothelial adhesion for the adhesive phenotypes expressed at the surface of infected erythrocytes.

Animals↗
PubMed1999

Identification of CD22 ligands on bone marrow sinusoidal endothelium implicated in CD22-dependent homing of recirculating B cells.

CD22 is a B cell-specific transmembrane protein known to function as a negative regulator of B cell signaling. It has also been implicated in cell adhesion through recognition of alpha2,6-linked sialic acids on glycans of target cells. Previous studies showed that CD22-deficient mice had a strongly reduced population of mature recirculating B cells in the bone marrow despite normal B cell development. Using a soluble recombinant form of the receptor (CD22-Fc), we demonstrate here that sialylated ligands for CD22 are expressed on sinusoidal endothelial cells of murine bone marrow but not on endothelial cells in other tissues examined. Injection of CD22-Fc revealed that the CD22 ligands in the bone marrow were accessible to the circulation. Treatment of mice with either CD22-Fc or affinity-purified anti-CD22 antibody led to an approximately 50% reduction in mature recirculating B cells in the bone marrow without affecting numbers in the spleen. Finally, consistent with the notion that CD22 is a homing receptor, we show that compared with wild-type mice, CD22-deficient animals have a lower number of immunoglobulin M-secreting plasma cells in the bone marrow.

Animals↗
PubMed1999

Dormancy of mammary carcinoma after mastectomy.

BACKGROUND: The longest interval between primary treatment of breast cancer and tumor recurrence, i.e., the limit of breast cancer dormancy, defines the appropriate length of follow-up, the effectiveness of treatment, and curability (no excess mortality risk for patients relative to the general population) for the disease. To determine this limit, we analyzed long-term follow-up data from patients who underwent a radical mastectomy during a four-decade period at the University of Chicago Hospitals. METHODS: For 1547 patients operated on during the period from 1945 through mid-1987, the number of recurrences and deaths occurring within each postoperative year were tabulated, and the hazard rate for first recurrence or death from breast cancer was estimated by use of the actuarial method. The excess mortality rate was calculated for successive 5-year intervals, beginning at the time of mastectomy, by use of U.S. life tables and matching on the basis of age, race, and sex. RESULTS: Most recurrences occurred within the first 10 years after mastectomy. Recurrences were rare after 20 years; only one recurrence was reported among 192 patients followed for 26-45 years. Patients who had a recurrence within 5 years following mastectomy had shorter subsequent survival times than those whose recurrence appeared after 5 years (two-sided P = .0001). The excess death rate increased with pathologic stage of the primary tumor. Overall, there was evidence of excess mortality up to 20 years postsurgery (two-sided P = .009). CONCLUSIONS: The limit of breast cancer dormancy in this patient population appears to be between 20 and 25 years. After this time, recurrences were rare, and the mortality rate was no longer statistically significantly different from that of the general population. Patients surviving to this time without evidence of recurrence or contralateral breast cancer are probably cured.

Breast Neoplasms↗
PubMed1999

The expression and distribution of dense granule proteins in the enteric (Coccidian) forms of Toxoplasma gondii in the small intestine of the cat.

The expression and distribution of dense granule proteins in the enteric (coccidian) forms of Toxoplasma gondii in the small intestine of the cat. Experimental Parasitology 91, 203-211. The expression and location of the dense granule proteins (GRA1-6 and NTPase) in the merozoite and during asexual and sexual development of Toxoplasma gondii in the small intestine of the cat (definitive host) was examined by immuno-light and electron microscopy. This was compared with that of tachyzoites and bradyzoites present in the intermediate host. It was found that the merozoite contained the characteristic apical organelles plus a few large dense granules. By immunocytochemistry, dense granules in merozoites were negative for GRA proteins 1 to 6 in contrast to both tachyzoites and bradyzoites in which dense granules were positive for all six proteins. The GRA proteins were associated with the parasitophorous vacuole (PV) during tachyzoite and bradyzoite development but were absent from the PV of the enteric stages. However, the merozoite dense granules were positive for NTPase, which was similar to the tachyzoite while this antigen was down regulated in the bradyzoite. The apparent release of the NTPases into the PV formed by merozoites was also similar to that described for the tachyzoite, possibly reflecting the relative metabolic activity of the various stages. This study shows that the majority of GRA proteins have a similar stage-specific expression, which is independent of NTPases expression. These observations are consistent with T. gondii having a different host parasite relationship in the enteric forms, which does not involve the GRA proteins 1-6.

Acid Anhydride Hydrolases↗
PubMed1999

In vivo expression and distribution of dense granule protein 7 (GRA7) in the exoenteric (tachyzoite, bradyzoite) and enteric (coccidian) forms of Toxoplasma gondii.

The in vivo expression and distribution of the dense granule protein GRA7 was examined in both the exoenteric (tachyzoite and bradyzoite) and enteric (coccidian) forms of Toxoplasma gondii by immunocytochemistry. There was strong staining of GRA7 in granules within all the infectious stages (tachyzoite, bradyzoite, merozoite and sporozoite). During tachyzoite development, GRA7 was secreted and was associated with the parasitophorous vacuole. In contrast, although there was staining of granules within the bradyzoites of more mature cysts, there appeared to be little staining of the tissue cyst wall or host cell. The apparent stage-specific variation in secretion of GRA7 between tachyzoites and bradyzoites was confirmed by double labelling using stage-specific markers (SAG1 and BAG1). In the enteric forms in the cat gut there was strong labelling of the PV containing early asexual and sexual stages and staining of a few granules in the apical cytoplasm of the merozoite. The positive enteric staining pattern differentiates GRA7 from the other GRA proteins (GRA1-6) which were absent in the merozoites and enteric stages. The staining pattern of GRA7 with strong staining during tachyzoite and enteric development and reduced staining in the tissue cysts is similar to that seen for NTPases. The function of GRA7 is unknown but it is unique among the dense granule proteins in being expressed in all the infectious forms of T. gondii which would point to a basic role in the vacuolar adaptations required for active parasite development.

Animals↗
PubMed1999

C-axis: a growth vector for the maxilla.

Based on M-point, defined as the center of the largest circle that is tangent to the superior, anterior, and palatal outlines of the maxilla in the sagittal view, a growth axis and its direction are described for each gender from age 7.4 to 18.75 years. Growth increments along the C-axis, defined by sella-M-point, are described by regression formulas with correlation coefficients of 0.618 for females and 0.669 for males. The vector (direction) of the growth axis, defined by the angle S-M-C-axis, increases in females from 42.21 degrees at 7.4 years to 44.47 degrees at 18.75 years, with a standard deviation of 3.55 degrees. In males, it increases from a mean of 41.69 degrees at 7.6 years to 45.55 degrees at 18.6 years, with a standard deviation of 2.74 degrees. This is based on 172 serial lateral cephalograms of 20 females and 174 serial lateral cephalograms of 19 males. The C-axis incremental growth change and its vector offer a means of quantifying complex maxillary growth in the sagittal plane via cephalometric measurements relative to and coordinated with other craniofacial structures.

Adolescent↗
PubMed1999

Netherton's syndrome: increased likelihood of diagnosis by examining eyebrow hairs.

Netherton's syndrome is a rare autosomal recessive condition with variable expression. It comprises an ichthyosiform dermatitis and erythroderma of variable intensity and manifestations, associated with hair abnormalities. The pathognomonic finding (required for diagnosis) is that of trichorrhexis invaginata identified by light and scanning electron microscopic examination of hair shafts. This may be difficult to establish because the hair is sparse and not all hairs exhibit abnormalities. In one patient, cutaneous and hair problems had existed since infancy, and despite repeated examination of scalp hairs, the definitive diagnosis was made only by examining eyebrow hairs at the age of 30 years. We subsequently compared the number of diagnostic lesions found on scalp and eyebrow hairs from two other patients with previously diagnosed Netherton's syndrome. The density of lesions was greater in eyebrow than scalp hair, and furthermore, all eyebrow hairs had at least one lesion. It is proposed that microscopic examination, if possible, of both scalp and eyebrow hair from patients in whom Netherton's syndrome is suspected would increase the chance of a positive diagnosis.

Adult↗