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J D Wilson

Publications and source records attributed to J D Wilson.

At least 181 records · Page 10Linked to original sources
PubMed1991

Technique of venous catheterization for sequential blood sampling from the pig.

A surgical technique for catheter implantation for sequential blood sampling in the pig is described. This methodology minimizes stress to the animal and chances of catheter dislodgement, both of which are detrimental to accurate long-term metabolic assessment of the animal model. The technique involves implantation of a vinyl catheter via the external jugular vein into the vena cava. To ensure catheter retention, the method uses a silicone plug fitted to the catheter and located cranial to its entry to the vein, double ligation at the venous insertion, subsequent passage of the catheter to exit 10 cm cranial to the scapula on the dorsal midline, and a denim vest fitted to the thorax. With this technique seven out of nine catheters have functioned more than 50 days, two functioned for approximately 100 days, and one for over 150 days.

Animals↗
PubMed1991

Aromatase expression in extragonadal tissues of the Sebright chicken is controlled by a retroviral promoter.

These studies have clarified a number of issues regarding the genetic control of extragonadal aromatase expression in birds carrying the henny-feathering trait. The marked increased in extragonadal aromatase expression of a unique species of aromatase mRNA is apparently controlled by a retrovirus. We have not yet established whether this promoter is the consequence of a new retroviral insertion or is due to a mutation in a preexisting retrovirus. It is of interest in this regard that the chicken aromatase gene is located in a region of chromosome 1 known to be rich in retroviral insertions. Second, the relationship of the henny-feathering trait to normal and abnormal extragonadal aromatase expression in man remains to be determined. At one extreme, it is possible that a similar mechanism may underlie syndromes typified by increases in extraglandular aromatase expression. Even if the parallel proves to be less exact, the Sebright chicken provides important insight into how aromatase expression may be modulated differently in different tissues. Finally, the discovery that retroviral sequences appear to modulate the expression of aromatase in extragonadal tissues represents an unusual method by which the genetic control of genes can be altered. In the case of the henny-feathering trait, the alteration in the dimorphic feathering pattern was the principal clue to the altered regulation of extragonadal aromatase. This relation is in keeping with the other examples of altered gene expression caused by retroviruses that have also caused morphologic alterations such as changes in coat color or the appearance of neoplasms (for examples, see Refs. 14-16).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

A two-channel hypothesis for regulation of cell division and differentiation.

We propose the hypothesis that extracellular regulation of cell division and differentiation acts through just two communications channels. These channels consist of a series of redundant components: extracellular messenger hormones; these hormones' receptors; cytoplasmic proteins activated by the hormone-receptor complex; and trans-activating nuclear regulatory proteins. One channel, here labeled "D" ("differentiate"), includes transforming growth factor-beta as one of its hormones; the other, labeled "G" ("growth") includes epidermal growth factor. We postulate that signal reception occurs as a result of competition between different actuating proteins for equilibrium-controlled binding to critical DNA sites. Stem cells commit to mitosis when some high proportion of critical sites is occupied by actuating proteins of the G class, and to terminal differentiation when a high proportion is occupied by "D" actuators. Intermediate occupancy can either lead to division into one differentiated and one stem cell, or to maintenance of cells in the reference state, quiescence. Equilibrium control of binding implies that critical site occupancy will be proportional to the relative concentrations of "D" and "G" actuating proteins in the nuclear fluid. These concentrations depend on the external hormone concentrations, the numbers of receptors on the cell membrane, and the coefficients of the rate-determining steps between internalization of the hormone-receptor complexes and activation of the actuating proteins. All of these quantities can be affected by various factors, including endocrine hormones. This model is consistent with most reported behavior of various growth factors, interferons, etc, toward a variety of cells in culture. It predicts that under otherwise constant conditions, high relative concentrations of a D-hormone (e.g. transforming growth factor-beta) will induce commitment to terminal differentiation, while high relative concentrations of a "G" hormone (e.g. epidermal growth factor) will induce mitosis. We have seen no report of an experiment which adequately tests this prediction. The model implies that cancer causing mutations are those which increase the relative intensity of the "G" signal; this can occur via changes in components of either channel. Such mutant cells should be both more likely to divide and less likely to differentiate than normal stem cells. Conversely, mutations which increase relative sensitivity to the "D" signal during embryonal development can lead to premature differentiation, cessation of growth, and structural abnormalities (terata).

Animals↗
PubMed1990

Implications of inflammatory changes on cervical cytology.

OBJECTIVE: To assess premenopausal women with inflammatory changes on cervical cytology for genital infections and cervical abnormalities. DESIGN: Prospective study of women attending general practice and family planning clinics who had a recent cervical cytology result with inflammatory changes. SETTING: Department of genitourinary medicine. PATIENTS: 102 Premenopausal women with recent cytology result showing inflammatory changes and with no history of antibiotic or antifungal treatment since their smear. INVESTIGATIONS: Genital examination and microbiological screening for genital infections; colposcopic examination about six weeks later. MAIN OUTCOME MEASURES: Detection of genital infections, particularly those sexually acquired, and abnormalities on colposcopy. RESULTS: Genital infections were isolated in 77 patients, and one or more sexually acquired infections were found in 22. Prevalence of sexually acquired infections was significantly correlated with younger age (particularly being under 25), being single, separated, or divorced; using non-barrier contraception; and recent change of sexual partner. An abnormality on colposcopy was found in 36 women. There was a strong correlation of a sexually acquired infection with an abnormality at colposcopy; hence younger women were more likely to have a colposcopic abnormality. CONCLUSIONS: Inflammatory changes on cytology are often associated with the presence of a sexually acquired infection and premalignant disease of the cervix, particularly in younger, single women using non-barrier contraception.

Adolescent↗
PubMed1990

Androgen resistance due to decreased amounts of androgen receptor: a reinvestigation.

To provide insight into the pathogenesis of the androgen resistance in a previously described family with X-linked Reifenstein syndrome, we have systematically assessed a variety of parameters of androgen receptor function in fibroblasts cultured from scrotal skin biopsies. We assessed the amount of high affinity binding, the affinity of ligand binding to the receptor, the upregulation of androgen receptor levels by androgen, the stability of ligand binding in intact fibroblasts at high temperature, the dissociation of ligand from the receptor, the intranuclear localization and salt elution profiles of ligand-receptor complexes, and the ultracentrifugation characteristics of the ligand-receptor complexes. Since the only parameter found to be abnormal is a decreased amount of receptor, we conclude that the underlying mutation in this family influences the amount rather than the structure of the androgen receptor protein.

Androgens↗
PubMed1990

Functional properties of proislet isografts in diabetic mice.

Fetal proislets (islet precursors) were transplanted to isogeneic diabetic CBA/H mice for analysis of their development and function post-transplant. Immunohistochemically, proislets contained scattered populations of insulin-, glucagon- and somatostatin-producing cells; following isotransplantation proislets developed into mature islet tissue with normal endocrine cell composition. Proislet isografts also developed normal endocrine function after transplantation to diabetic mice. Isografts of eight and four donor-equivalents of proislets reversed streptozotocin-induced diabetes by 30 +/- 7 and 74 +/- 26 days, respectively. Following stimulation with intragastric glucose (50 mg) mice carrying established isografts of four donor-equivalents of proislets exhibited normal blood glucose responses; those grafted with eight donor-equivalents of proislets showed decreased blood glucose levels. The beta cell mass of proislets continued to grow and differentiate post-transplant; established proislet grafts contained greater than 100-fold the total insulin content of the corresponding number of fetal pancreases from which the proislets were derived. The mean insulin content of eight and four donor-equivalent proislet grafts was 231% and 156%, respectively, that of the mean content of adult mouse pancreas. The data show that glucose homeostasis can be restored in diabetic mice following isotransplantation of fetal proislets and that the quantity of tissue transplanted is a critical parameter in achieving optimal control.

Animals↗
PubMed1990

Effect of GK1.5 monoclonal antibody dosage on survival of pig proislet xenografts in CD4+ T cell-depleted mice.

Treatment of CBA/H mice with 5 injections of anti-CD4 (GK1.5 mAb) terminating on day 10 posttransplant resulted in long-term survival (greater than or equal to 6 weeks) of fetal pig proislet (pancreatic islet precursor) xenografts. The GK1.5 mAb dose determined the duration of CD4+ T cell depletion and the extent to which the survival of pig proislet xenografts was prolonged. Sustained depletion of CD4+ T cells (0%, 1%, and 9% of total T cells in peripheral lymph nodes at 2, 4, and 6 weeks, respectively) and survival of proislet xenografts at 6 weeks posttransplant was observed when transplant recipients were treated with 5.4 mg GK1.5 mAb/injection. Treatment of transplanted mice with a suboptimal dose of GK1.5 mAb (0.2 mg/injection) resulted in the same level of depletion at 2 weeks posttransplant but a more rapid recovery of CD4+ T cells in the periphery (24% of total T cells at 4 weeks) and only temporary prolongation in xenograft survival (less than or equal to 4 weeks). Control xenografts showed evidence of graft destruction by as early as 6-7 days posttransplant and were completely rejected by 2 weeks. The rejection reaction consisted predominantly of CD4+ T cells, eosinophils and F4/80-positive macrophages. Only small numbers of CD8+ T cells were identified. CD4+ T cells therefore represented the major T cell component of the cellular infiltrate. In contrast, surviving xenografts in GK1.5 mAb-treated recipient mice showed essentially an absence of CD4+ T cells but presence of CD8+ T cells. This finding may be attributable to the increase (1.7-3.1-fold) in the absolute size of the population of CD8+ T cells in the periphery following GK1.5 mAb treatment in vivo. Compared with isolated fetal pig proislets, which contained only a small population of insulin-producing cells in addition to glucagon- and somatostatin-positive cells, surviving pig proislet xenografts contained mainly insulin-positive beta cells with smaller populations of glucagon- and somatostatin-positive cells. Fetal pig proislets therefore differentiate into insulin-producing islet tissue posttransplant and thus show evidence of normal development of endocrine function.

Animals↗
PubMed1990

Antibody-induced rejection of pig proislet xenografts in CD4+ T cell-depleted diabetic mice.

Reversal of diabetes in mice was achieved following in vivo depletion of host CD4+ T cells and transplantation of xenogeneic fetal pig proislets (pancreatic islet precursors). These procedures resulted in xenograft tolerance since established pig proislet xenografts were not rejected by antipig antibodies produced in the host, and rejection was not induced following the administration of donor major histocompatibility complex--specific pig lymphocytes. Proislet xenografts were rejected following the administration of donor MHC-specific hyper-immune antipig PBL serum raised in normal mice. Although established proislet xenografts in anti-CD4-treated mice are sensitive to antibody-mediated destruction, such hosts are unable to produce an antibody response that leads to graft rejection. The study indicates that the mechanism of preventing xenograft rejection by anti-CD4 treatment in vivo involves not only initial CD4+ T cell depletion but also quantitative and/or qualitative modulation of a CD4+ T cell-dependent antibody response. As a consequence, an apparent state of xenograft tolerance is produced.

Animals↗
PubMed1990

Androgen receptor of rat penis is down-regulated by androgen.

To provide insight into the factors that control androgen receptor levels in rat penis, we assessed 5 alpha-[3H]-dihydrotestosterone binding in low-salt [10 mM tris(hydroxymethyl)aminomethane (Tris), 10 mM Na2M0O4] and high-salt (10 mM Tris, 10 mM Na2M0O4, 0.5 M KCl) extracts of rat penis using sucrose density gradients. Total receptor content decreased from approximately 729 +/- 114 fmol/g tissue at 3 wk of age to less than 50 fmol/g tissue at 10 wk of age. Castration of 3-wk-old rats prevented penile growth and the age-related decline in penile androgen receptor. Treatment of 3-wk-old castrated rats with 5 alpha-dihydrotestosterone caused an acceleration in the decline in receptor levels compared with intact animals. Castration of 10-wk-old rats (after androgen receptor levels had decreased) did not result in an increase in the amount of total androgen receptor by 16 wk of age. To determine the specificity of the androgen-mediated decline in receptor levels, the amounts of prostate androgen receptor were compared with those of the penis at different ages. When expressed as femtomoles per organ, the total androgen receptor level in the prostate increased fourfold from 3 to 10 wk of age, whereas the total androgen receptor in the penis declined approximately threefold. We conclude that the downregulation of the penile androgen receptor content that occurs in the rat between 3 and 10 wk of age is androgen mediated, does not occur in all androgen target tissues, and is prevented but not reversed by castration.

Aging↗
PubMed1990

Variation in inducibility of cytochrome P-450c and aryl hydrocarbon hydroxylase in rat liver, lung, kidney, pancreas and nasopharynx.

The effect of 3-methylcholanthrene (MC) treatment on the cytochrome P-450c content of various rat tissues was examined by measuring the level of immunodetectable P-450c in conjunction with its aryl hydrocarbon hydroxylase (AHH) activity. Immunoblots revealed that P-450d was induced in the nasopharynx and pancreas in addition to its previously reported induction in the liver, lung and kidney. In contrast to P-450c, induction of the immunorelated P-450d was observed only in the liver. The specific content of immunodetected P-450c in the tissue homogenates decreased in the order: liver, nasopharynx, pancreas, lung, kidney. The corresponding AHH activities decreased in the order: liver, kidney, lung, nasopharynx, pancreas. The ratio of AHH activity to P-450c content varied widely among tissues: ratios of 37.2:1.7:0.47:0.04:0.02 were obtained for the kidney, liver, lung, nasopharynx and pancreas, respectively. The absence of a direct relationship between the levels of AHH and P-450c indicates that the extrahepatic activity may partially derive from P-450 forms other than P-450c and/or the specific activity of P-450c varies among different tissues.

Animals↗
PubMed1990

A single nucleotide substitution introduces a premature termination codon into the androgen receptor gene of a patient with receptor-negative androgen resistance.

Mutations of the androgen receptor that impair the action of 5 alpha-dihydrotestosterone and testosterone result in abnormal male sexual development. The definition of the organization of the androgen receptor gene has permitted us to examine its structure in nine patients with androgen resistance that exhibit absent 5 alpha-dihydrotestosterone binding in cultured fibroblasts (receptor-negative androgen resistance). Using labeled probes specific for each individual coding exon, we find no gross rearrangements, insertions, or deletions of the androgen receptor gene in these patients. To analyze the genetic defect in these receptor-negative patients, we used the polymerase chain reaction to amplify each individual exon of the androgen receptor gene in nine affected patients. In all patients, the size of each amplified exon segment was identical to that in normal individuals. The nucleotide sequence of the entire coding region of the androgen receptor was determined in one of these patients. A single nucleotide substitution was identified that results in a premature termination codon in exon 6 at amino acid 794. S1 nuclease protection assays demonstrated that normal levels of androgen receptor mRNA are present in skin fibroblasts of this patient. Transfection of a mutated androgen receptor cDNA containing a termination codon at position 794 into eukaryotic cells resulted in formation of a normal amount of receptor protein, as indicated by immunoblotting, but the expressed protein does not bind 5 alpha-dihydrotestosterone. These findings suggest that the presence of a premature termination codon at amino acid 794 of the androgen receptor is the cause of androgen resistance in this patient.

Adolescent↗
PubMed1990

Testosterone at high concentrations interacts with the human androgen receptor similarly to dihydrotestosterone.

Testosterone and dihydrotestosterone are believed to exert their androgenic effects by interacting with a single intracellular receptor protein in androgen target tissues. During fetal life, however, testosterone mediates the virilization of the Wolffian ducts into the epididymis, vas deferens, and seminal vesicles, whereas the urogenital sinus and external genitalia require the in situ conversion of testosterone to dihydrotestosterone to undergo male development. The reason why the signal provided by testosterone needs to be amplified in some androgen target tissues but not in others remains an enigma. To provide insight into the different actions of these androgens we studied their interaction with the human androgen receptor in fibroblasts cultured from the genital skin of a patient with 5 alpha-reductase deficiency. Dihydrotestosterone was formed in negligible amounts in these cells, and in some experiments the residual 5 alpha-reductase activity was further blocked with the 5 alpha-reductase inhibitor finasteride. Saturation analysis in fibroblast monolayers disclosed similar amounts of binding with testosterone and dihydrotestosterone, and the affinity of binding of dihydrotestosterone was, on the average, about 2-fold greater than that of testosterone. [3H]Testosterone also exhibited a 5-fold faster dissociation rate from the receptor than [3H]dihydrotestosterone. In thermolability experiments the [3H]testosterone-receptor complex displayed marked instability at 42 C with 2 nM [3H] testosterone, whereas with 20 nM [3H]testosterone, receptor stability was similar to that seen with [3H]dihydrotestosterone. In up-regulation experiments, 2 nM [3H]testosterone produced a 34% increase in specific androgen receptor binding after 24 h, whereas 20 nM [3H]testosterone produced an average increase of 64%. Our results suggest that the weaker androgenic potency of testosterone compared to that of dihydrotestosterone resides in its weaker interaction with the androgen receptor, most clearly demonstrable as an increase in the dissociation rate of testosterone from the receptor. When present in relatively high concentrations, however, testosterone overcomes this defect by mass action.(ABSTRACT TRUNCATED AT 400 WORDS)

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed1990

Antipeptide antibodies to two distinct regions of the androgen receptor localize the receptor protein to the nuclei of target cells in the rat and human prostate.

We have developed polyclonal antibodies to two synthetic peptides corresponding to the amino-(N-)terminal or carboxyl-(C-)terminal segments of the human androgen receptor (hAR) protein, as deduced from the nucleic acid sequence of the androgen receptor cDNA. Immunoreactive antisera were identified by solid phase enzyme-linked immunosorbent assay and purified by peptide affinity chromatography. Specific immunoreactivity with the hAR was confirmed by immunoblotting, using both a fusion protein produced in E. coli that contains the C-terminal 880-amino acid sequence of hAR and the full-length receptor protein produced in COS cells after transfection with a plasmid containing the entire hAR-coding region. Immunohistological evaluation of rat and human prostatic tissue using anti-C-terminal or anti-N-terminal antibodies demonstrated similar patterns of specific staining of the nuclei of epithelial and stromal cells. Castration resulted in a decrease in the amount of nuclear AR detected in the rat prostate after a short time of exposure to anti-C-terminal antibodies (less than 4 h), but did not alter the level of specific staining obtained with anti-N-terminal antibodies. This decrease in nuclear staining using anti-C-terminal antibodies could be reversed by treating castrated animals with dihydrotestosterone. When longer times of exposure to the primary antibodies were used, high levels of nuclear staining were obtained with both types of antibodies in prostate specimens from castrate as well as as intact rats. This immunohistochemical staining pattern contrasts with receptor measurements in rat prostate homogenates that indicate the partition of AR binding into the low salt (cytosolic) fraction in the castrate animal and into the high salt (nuclear) fraction in the intact animal. Our results suggest that the AR is predominantly a nuclear protein even in the absence of ligand and that dihydrotestosterone serves to tighten its association with the nucleus. These data also suggest that the immunoreactivity of anti-C-terminal antibodies is influenced by the presence of dihydrotestosterone, presumably via an alteration in the physical state of the receptor protein.

Amino Acid Sequence↗
PubMed1990

Estrogen and androgen production rates in two brothers with Reifenstein syndrome.

Defects of the androgen receptor in 46,XY individuals cause aberrant virilization that varies from a female phenotype to men with minor defects. More severely affected individuals also develop gynecomastia associated with enhanced estradiol secretion by the testis. However, the degree of breast development does not correlate with the rate of estrogen production, leading us to propose that feminization is a function of the degree of androgen resistance as well as the rate of estrogen formation. To test this hypothesis we measured estrogen and androgen formation in two brothers with perineoscrotal hypospadias and severe gynecomastia (the Reifenstein phenotype) due to a mutation that impairs androgen receptor function. Rates of estradiol production (60 and 70 micrograms/day) were elevated, but were not as high as in previously studied men with a similar phenotype. We conclude that the variable degree of feminization in this disorder cannot be explained by androgen resistance alone.

Adult↗