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Biomedical subjects

S Macpherson

Publications and source records attributed to S Macpherson.

16 recordsLinked to original sources
PubMed2003

Human 'testicular dysgenesis syndrome': a possible model using in-utero exposure of the rat to dibutyl phthalate.

BACKGROUND: The disorders comprising human 'testicular dysgenesis syndrome' (TDS) may be increasing in incidence. TDS originates in fetal life but the mechanisms are not known, and discerning them requires an animal model. METHODS AND RESULTS: The study investigated whether male rats exposed in utero to dibutyl phthalate [DBP; 500 mg/kg on gestational days (GD) 13-21] would provide a suitable model for human TDS. DBP induced a high rate (>60%) of cryptorchidism (mainly unilateral), hypospadias, infertility and testis abnormalities, similar to those in human TDS. Cell-specific immunohistochemistry and confocal microscopy were used to track development of Sertoli [anti-Müllerian hormone (AMH), Wilm's tumour (WT-1) protein, p27(kip)], Leydig [3beta-hydroxysteroid dehydrogenase (3beta-HSD)], germ (DAZL protein) and peritubular myoid (smooth muscle actin) cells from fetal life to adulthood. In scrotal and cryptorchid testes of DBP-exposed males, areas of focal dysgenesis were found that contained Sertoli and Leydig cells, and gonocytes and partially formed testicular cords; these dysgenetic areas were associated with Leydig cell hyperplasia at all ages. Suppression ( approximately 90%) of testicular testosterone levels on GD 19 in DBP-exposed males, coincident with delayed peritubular myoid cell differentiation, may have contributed to the dysgenesis. Double immunohistochemistry using WT-1 (expressed in all Sertoli cells) and p27(kip) (expressed only in mature Sertoli cells) revealed immature Sertoli cells in dysgenetic areas. DBP-exposed animals also exhibited Sertoli cell-only (SCO) tubules, sporadically in scrotal and predominantly in cryptorchid, testes, or foci of SCO within normal tubules in scrotal testes. In all SCO areas the Sertoli cells were immature. Intratubular Leydig cells were evident in DBP-exposed animals and, where these occurred, Sertoli cells were immature and spermatogenesis was absent. Abnormal Sertoli cell-gonocyte interaction was evident at GD 19 in DBP-exposed rats coincident with appearance of multinucleated gonocytes, although these disappeared by postnatal day 10 during widespread loss of germ cells. CONCLUSIONS: Abnormal development of Sertoli cells, leading to abnormalities in other cell types, is our hypothesized explanation for the abnormal changes in DBP-exposed animals. As the testicular and other changes in DBP-exposed rats have all been reported in human TDS, DBP exposure in utero may provide a useful model for defining the cellular pathways in TDS.

Age Factors↗
PubMed2002

Expression of oestrogen receptor beta (ERbeta1) protein in human breast cancer biopsies.

Oestrogen action is mediated via specific receptors that act as ligand-activated transcription factors. A monoclonal antibody specific to the C-terminus of human oestrogen receptor beta has been characterized and the prevalence of expression of oestrogen receptor beta protein investigated in a well defined set of breast cancers. Reverse transcription-polymerase chain reaction analysis of RNA from tissue biopsies detected oestrogen receptor beta in all samples examined. The anti-oestrogen receptor beta antibody cross reacted specifically with both long (approximately 59 Kd) and short (approximately 53 Kd) forms of recombinant oestrogen receptor beta. Western blot analysis of breast tumours contained both forms of oestrogen receptor beta protein although in some samples lower molecular weight species (32--45 Kd) were identified. Fifty-one breast cancer biopsies were examined using immunohistochemistry; 41 (80%) were immunopositive for oestrogen receptor alpha, 48 (94%) were immunopositive for oestrogen receptor beta and 38 (74.5%) co-expressed both receptors. Expression of oestrogen receptor beta was exclusively nuclear and occurred in multiple cell types. There was no quantitative relationship between staining for the two ERs although in tumours in which both receptors were present immunoexpression of oestrogen receptor alpha was invariably more intense. The significance of oestrogen receptor beta protein expression in breast cancers to therapy remains to be determined but the availability of a well characterized antibody capable of detecting oestrogen receptor beta in archive material will facilitate the process.

Antibodies, Monoclonal↗
PubMed2002

RNA binding protein Musashi1 is expressed in sertoli cells in the rat testis from fetal life to adulthood.

The Musashi1 (Msi1) gene identified in mouse is a member of a subfamily of RNA binding proteins that are highly conserved across species. Msi1 expression is highly enriched in proliferative cells within the developing central nervous system. Within the testis, proliferation and differentiation of germ cells takes place within the seminiferous epithelium, where these cells are supported physically and functionally by Sertoli cells that do not themselves proliferate following the onset of puberty. RNA binding proteins expressed in testicular germ cells are essential for normal fertility. Preliminary data suggested the mRNA for Msi1 was present in ovary; therefore, we used an Msi1-specific cRNA and monoclonal antibody to investigate whether Msi1 was expressed in the testis. Msi1 mRNA was expressed in rat testis from birth until adulthood; in situ hybridization revealed silver grains within the seminiferous epithelium. Immunohistochemical studies demonstrated that at all ages examined (from Fetal Day 14.5 until adulthood) Msi1 protein was expressed in Sertoli cells. In fetal and adult rat ovaries, Msi1 was detected in granulosa cells and their precursors. In Sertoli cells, protein was detected in both cytoplasmic and nuclear compartments; in adult testes, the immunointensity of the nuclear staining was stage dependent, with highest levels of expression in Sertoli cells at stages I-VI. In rat gonads, the RNA binding protein Msi1 is expressed in both proliferating and nonproliferating Sertoli and granulosa cells.

Aging↗
PubMed2002

Development and validation of a new monoclonal antibody to mammalian aromatase.

The biosynthesis of oestrogens from androgens is catalysed by the aromatase complex, an essential component of which is the aromatase cytochrome P450 (P450 arom) protein. Expression of a functional P450 arom is essential for normal fertility in males and females and the sequence of the protein is highly conserved. We have raised a new monoclonal antibody against a conserved peptide and validated it on fixed tissue sections of the rat, common marmoset (Callthrix jacchus) and human. The monoclonal antibody was used successfully for Western analysis and specifically reacted with a 55 kDa protein in microsomal extracts. On sections of ovaries in all three species, expression in follicles was specific to the mural granulosa cells of antral follicles and was present in corpora lutea. In the human and marmoset, staining of luteal cells was markedly heterogeneous and did not appear to vary consistently with the stage of the cycle. The intensity of immunostaining was elevated in corpora lutea from pregnant rats and following human chorionic gonadotropin rescue in the human. In the testis, the highest levels of expression were observed in the Leydig cells within the interstitium. In adult rat and marmoset, and possibly also in the human, some P450 arom was associated with the cytoplasm surrounding elongate spermatids but other germ cells were immunonegative. In conclusion, a new monoclonal antibody specific for P450 arom recognises the protein in rodent, primate and human. Its ability to work on fixed tissue sections will facilitate identification of individual cells expressing P450 arom within complex tissues.

Animals↗
PubMed2001

Differential expression of oestrogen receptor alpha and beta proteins in the testes and male reproductive system of human and non-human primates.

The role(s) oestrogens play in male adult reproductive function remains uncertain. We have used antibodies specific for oestrogen receptor- alpha (ERalpha) and - beta (ERbeta) to investigate their distribution within the male. In testes from adult human, macaque and marmoset, ERbeta protein was detected in Sertoli cells, Leydig cells and peritubular myoid cells. In germ cells, the intensity of immunostaining for ERbeta was variable between species. Immunoexpression in preleptotene, leptotene and zygotene spermatocytes was low/absent in all species. Elongated spermatids were consistently immunonegative. No ERalpha immunoexpression was detected in testes. ERbeta was detected in epithelial and stromal cell nuclei throughout the male reproductive system [efferent ductules (ED), epididymis, vas deferens, seminal vesicles] and in the bladder. ERalpha was detected in non-ciliated epithelial cells in the ED, but rarely in epithelial and basal cells within the epididymis. Epithelial cells from seminal vesicles and bladder were immunonegative for ERalpha. Expression of ERalpha in stromal cells was rare in the ED, epididymis and bladder but more frequent in seminal vesicles. Expression of ERalpha, and long and short forms of ERbeta, was confirmed by Western blotting. The widespread expression of ERbeta suggests that it is the primary target for modulation of tissue function via oestrogenic ligands in the male reproductive system.

Adult↗
PubMed2000

Differential expression of estrogen receptor-alpha and -beta and androgen receptor in the ovaries of marmosets and humans.

Estrogens and androgens are essential for the maturation of the ovarian follicle and normal fertility in the female. We have used antibodies specific for both forms of estrogen receptor (alpha [ERalpha] and beta [ERbeta]) and androgen receptor (AR) to investigate the pattern of receptor expression in ovaries obtained from women and from a New World primate, the Common marmoset (Callthrix jacchus). On Western blots, three antibodies directed against different peptides within human ERbeta all recognized recombinant human (h) ERbeta but did not bind to recombinant hERalpha. The ERbeta protein was extracted from human ovary and prostate and marmoset ovary. In marmoset and human ovaries, ERbeta protein was detected in the nuclei of granulosa cells in all sizes of follicle (both before and after formation of the antrum), and it was also detected in thecal cells, corpora lutea, surface epithelium, and stroma. In contrast, ERalpha protein was not detected in the nuclei of granulosa cells in preantral follicles, was low/absent from stromal and thecal cells, but was expressed in granulosa cells of antral follicles and in the surface epithelium. The pattern of expression of AR protein more closely resembled that of ERbeta than ERalpha. In conclusion, three independent antibodies have demonstrated convincingly that ERbeta is expressed in a wide range of cells in the primate ovary. Granulosa cells in preantral follicles could contain ERbeta:beta dimers. In antral follicles, however, ERalpha is also expressed, and the formation of homo- or heterodimers containing ERalpha may influence the pattern of gene activation within these cells.

Amino Acid Sequence↗
PubMed2000

Study of the assembly of vesicular stomatitis virus N protein: role of the P protein.

To derive structural information about the vesicular stomatitis virus (VSV) nucleocapsid (N) protein, the N protein and the VSV phosphoprotein (P protein) were expressed together in Escherichia coli. The N and P proteins formed soluble protein complexes of various molar ratios when coexpressed. The major N/P protein complex was composed of 10 molecules of the N protein, 5 molecules of the P protein, and an RNA. A soluble N protein-RNA oligomer free of the P protein was isolated from the N/P protein-RNA complex using conditions of lowered pH. The molecular weight of the N protein-RNA oligomer, 513,879, as determined by analytical ultracentrifugation, showed that it was composed of 10 molecules of the N protein and an RNA of approximately 90 nucleotides. The N protein-RNA oligomer had the appearance of a disk with outer diameter, inner diameter, and thickness of 148 +/- 10 A, 78 +/- 9 A, and 83 +/- 8 A, respectively, as determined by electron microscopy. RNA in the complexes was protected from RNase digestion and was stable at pH 11. This verified that N/P protein complexes expressed in E. coli were competent for encapsidation. In addition to coexpression with the full-length P protein, the N protein was expressed with the C-terminal 72 amino acids of the P protein. This portion of the P protein was sufficient for binding to the N protein, maintaining it in a soluble state, and for assembly of N protein-RNA oligomers. With the results provided in this report, we propose a model for the assembly of an N/P protein-RNA oligomer.

Cloning, Molecular↗
PubMed1998

Abnormalities in functional development of the Sertoli cells in rats treated neonatally with diethylstilbestrol: a possible role for estrogens in Sertoli cell development.

Diethylstilbestrol (DES) was administered neonatally (Days 2-12; 10 microg on alternate days) to rats, and developmental changes in Sertoli cell function were evaluated at 18, 25, and 35 days of age and compared to those observed in rats administered a GnRH antagonist (GnRHa; Days 2 and 5; 10 mg/kg) or a vehicle (controls). DES and GnRHa treatments resulted in similar reductions in both Sertoli cell numbers (40% for DES, 48% for GnRHa) and suppression of testicular growth at 18 and 25 days, though by 35 days the suppression was more pronounced (p < 0.001) in DES-treated animals. Plasma FSH levels were suppressed markedly at 18 and 25 days, but not at 35 days, in GnRHa-treated rats, whereas in DES-treated rats the FSH levels were suppressed significantly only at 35 days. Both treatments suppressed plasma levels of inhibin B, though this was more pronounced (p < 0.05) in DES- than in GnRHa-treated rats. In controls, Sertoli cell immunoexpression of inhibin alpha, sulfated glycoprotein-1 (SGP-1), and androgen receptor (AR) increased in intensity and changed to an adult, stage-dependent pattern by 25 days. In GnRHa-treated rats these changes were reduced in intensity but were similar to those in controls at 35 days. In DES-treated rats, the increase in intensity and stage-dependent pattern of immunoexpression of inhibin alpha, SGP-1, and AR were virtually absent at 25 days but were present by 35 days. Germ cell volume per Sertoli cell was reduced in GnRHa- and DES-treated rats compared with controls at 18 and 25 days but was significantly greater (p < 0. 001) in DES- than in GnRHa-treated rats at 35 days. The proportion of apoptotic to viable germ cells was increased (p < 0.01) in GnRHa- and DES-treated rats compared with controls at 18 and 25 days; but at 35 days, values in GnRHa-treated rats had declined to control values whereas those for DES-treated rats remained 10-fold elevated (p < 0.001). In adulthood, testis weight and daily sperm production were reduced by 43% and 44%, respectively, in GnRHa-treated rats, but spermatogenesis was grossly normal. Comparable changes were observed in approximately 25% of DES-treated rats, but the majority exhibited > 60% reduction in testis weight with many Sertoli cell-only tubules and very low daily sperm production. Taken together, these data are interpreted as providing evidence for direct modulation of Sertoli cell (maturational) development by DES.

Aging↗
PubMed1998

Prospective care of elderly patients in family practice. Is screening effective?

OBJECTIVE: To evaluate cost and benefits of screening for and treating health and lifestyle risks among community-dwelling elderly. DESIGN: Randomized controlled trial. SETTING: Primary care. PARTICIPANTS: An opportunistic and prompted sample of 619 rostered elderly patients presenting for treatment who screened positive. INTERVENTIONS: One third (209) of experimental subjects had screening questionnaires placed in their charts with concerns highlighted for referrals. Two control groups received usual care. MAIN OUTCOME MEASURES: Yearly assessments of health service use and multidimensional functional capacity. RESULTS: Overall, screening and treatment of functionally active, elderly, middle-class people had no significant beneficial effect. Almost half of the experimental sample was ineligible because of treatment noncompliance. Generally ineligible subjects were older and more severely impaired. Subjects 75 years and older with risk factors showed improvement in daily living activities, and those living alone were found to have improved mental health and social functions (11% and 22%, respectively). CONCLUSIONS: Screening and treatment was ineffective in improving total functional capacity of all seniors 65 years and older. Elderly people 75 years and older, however, who were living alone or lonely did benefit from screening and treatment showing an improvement in daily activities, mental health scores, and social functions. This finding has implications for selective preventive health care spending for the elderly. A 2-year follow-up period could be too brief to detect long-term effects of early intervention with younger, middle-class seniors, especially those who are already functionally active.

Activities of Daily Living↗
PubMed1991

Management of tibial fractures with the orthofix dynamic external fixator.

We report our experience with the use of the Orthofix external fixator for the management of closed and open tibial fractures. One hundred fractures were treated in 96 patients between May 1985 and December 1989. There were 47 closed and 53 open fractures. Of the open fractures, 11 were grade I, 16 were grade II and 26 were grade III. Forty-five closed fractures and 49 open fractures went on to solid bony union in an average of 15.2 and 20.5 weeks, respectively. Non-union requiring operative intervention occurred in two closed and four open fractures (6%). The other main complications were pin track infection (30% of cases) and malunion which occurred in 14% of closed fractures and 32% of open fractures. We concluded that dynamic axial fixation is a useful method of treatment for open and difficult closed tibial fractures. Our experience indicates that malunion and pin track infection remain common problems and have still to be overcome.

Adolescent↗
PubMed1987

Neuropeptide Y in multiple endocrine neoplasia: release during surgery for phaeochromocytoma.

High plasma concentrations of neuropeptide Y (NPY) were found in a patient with bilateral adrenal phaeochromocytomas and medullary thyroid carcinoma associated with MEN IIa (32 pmol/l, normal less than 3.5 pmol/l). Both adrenal tumours contained and secreted NPY. Manipulation at operation produced a remarkable increase in plasma NPY concentrations (peak = 1631 pmol/l) coinciding with increases in plasma levels of catecholamines and arterial pressure. NPY was also shown to be present in thyroid tumour tissue: the concentration of NPY in tumour was 50-fold higher (0.9 nmol/g vs 0.004 nmol/g) than in adjacent normal thyroid tissue. It is possible that NPY from some phaeochromocytomas may contribute to hypertension during surgery.

Adrenal Gland Neoplasms↗
PubMed1981

Cancer of the head of the pancreas and choledochoduodenal junction: a clinical study of 88 Whipple resections.

The results of 88 pancreaticoduodenectomies performed at one institution over a period of 15 years (1960-1974) have been reviewed. Resection was undertaken in 62 patients with carcinoma of the head of the pancreas (HP) and in 26 patients with carcinoma of the choledochoduodenal junction (CDJ), with an operative mortality of 10% and 15% respectively. The 5-year survival rate for patients without positive lymph node histology was 13% for those with HP carcinoma and 21% for those with CDJ carcinoma. With positive nodes, the 5-year survival rates were 3% and 33% respectively, which suggests that a Whipple resection in patients with CDJ carcinoma has a relatively favourable prognosis even in the presence of lymph node metastases. In patients with HP carcinoma, on the other hand, resection was associated with a poor prognosis.

Adult↗