Search PubMed⌕ Search

Biomedical subjects

M J Reed

Publications and source records attributed to M J Reed.

At least 91 records · Page 5Linked to original sources
PubMed1996

In vivo activity of 4-methylcoumarin-7-O-sulfamate, a nonsteroidal, nonestrogenic steroid sulfatase inhibitor.

Steroid sulfatase regulates the formation of estrone from estrone sulfate (E1S) and dehydroepiandrosterone (DHA) from DHA sulfate. DHA can be converted to androstenediol, a steroid with potent estrogenic properties, and inhibition of steroid sulfatase activity is therefore an important therapeutic target. Because nonsteroidal steroid sulfatase inhibitors may offer some advantage for use in the treatment of breast cancer, 4-methylcoumarin-7-O-sulfamate (COUMATE) was synthesized and shown to be active in vitro. In this study, in vitro and in vivo techniques have been used to confirm that COUMATE, in contrast to the steroidal steroid sulfatase inhibitor estrone-3-O-sulfamate, is devoid of estrogenic activity. COUMATE did not stimulate the growth of MCF-7 breast cancer cells or uteri of ovariectomized rats, in contrast to estrone-3-O-sulfamate. COUMATE was orally active in vivo and after multiple dosing (10 mg/kg/day for 7 days) inhibited liver estrone sulfatase activity by 85%. Seven days after single or multiple dosing with COUMATE, liver estrone sulfatase activity was almost fully restored. Measurement of estrone sulfatase activity in WBCs revealed a degree of inhibition similar to that detected in liver samples. COUMATE was able to completely block the ability of E1S to stimulate uterine growth in ovariectomized rats. The development of a potent nonsteroidal, nonestrogenic steroid sulfatase inhibitor should allow the therapeutic potential of this type of therapy to be evaluated.

Animals↗
PubMed1996

Enhanced cell proliferation and biosynthesis mediate improved wound repair in refed, caloric-restricted mice.

Aged mice that have undergone long-term caloric-restriction (CR) have improved health and enhanced longevity in comparison to aged mice that are ad libitum-fed (AL). However, caloric-restriction does not benefit the impaired wound healing of aged mice. To test the hypothesis that CR mice have the capacity for enhanced wound repair, but require a short-term period of additional nutrient intake to show this advantage, we assessed wound healing in CR mice that had been refed (RF) an ad libitum diet for 4 weeks prior to wounding. Two strains of AL young (Y AL) (4-6 months), AL middle-aged (M AL) (15-17 months), and three different, matched cohorts of old mice (O) (30-33 months): O AL, O CR, and O RF were studied. Two full-thickness 4 mm diameter punch biopsy skin wounds were created on the dorsum of each mouse. Animals were sacrificed and wounds were harvested at 1,2,3,5, and 7 days post-wounding. Repair of wounds was slower in O AL and O CR mice compared to Y AL and M AL animals. In contrast, the O RF mice healed similarly to that of the Y AL and M AL mice, as assessed by measures of wound area and histologic criteria. O RF mice demonstrated enhanced synthesis of type I collagen mRNA in comparison to O AL and O CR mice. A greater number of endothelial cells and fibroblasts at the wound edge of the O RF mice exhibited replication in vivo as measured by uptake of BrdU. O RF mice had higher levels of insulin-like binding protein 3 (IGFBP-3). Furthermore, fibroblasts derived from the explant of the punch biopsy of O CR mouse skin revealed enhanced proliferation and contraction in vitro, in comparison to fibroblasts from the O AL mice. In conclusion, O RF mice demonstrate an enhanced capacity to undergo wound repair in comparison to O AL mice. This effect appears to be mediated, in part, by enhanced cell proliferation, contraction, and collagen biosynthesis. In addition, short-term refeeding induced an increase in the serum level of IGFBP-3, the major binding protein for IGF-1. These data confirm that cells from O CR animals have a preserved proliferative, biosynthetic, and contractile capacity, but that an adequate source of nutrients is necessary to demonstrate this advantage in wound healing.

Aging↗
PubMed1996

Contrast letter thresholds in the non-affected eye of strabismic and unilateral eye enucleated subjects.

To investigate the effects of visual disruption on contrast letter thresholds of the non-affected eye, subjects with one eye enucleated, strabismic subjects using the non-deviating eye and normal control subjects were asked to identify letters on eye charts and single letter cards which varied in contrast (between 4 and 96%) and size. At all contrast, contrast letter acuity of eye enucleated subjects was superior to both normal control subjects and strabismic subjects. Early onset strabismic subjects (onset < 24 months) showed inferior performance to normal control subjects at all contrasts of 25% and above. Late onset strabismic subjects showed normal performance at all contrasts, except for high contrast single letters, where performance was inferior to normal control subjects. Further, for all subjects groups, performance on letter charts was similar to performance on single letter cards. We conclude that disruption to the visual system caused by eye enucleation or strabismus is not equivalent. These differences may be due to intrinsic differences between the visual systems of eye enucleated subjects and strabismic subjects and/or to the profound differences in deprivation caused by the two conditions.

Adolescent↗
PubMed1996

Heteroatom-substituted analogues of the active-site directed inhibitor estra-1,3,5(10)-trien-17-one-3-sulphamate inhibit estrone sulphatase by a different mechanism.

Estrogens have a pivotal role in the growth and development of hormone-dependent breast cancers. In postmenopausal women, the hydrolysis of the conjugate estrone sulphate (E1S) to estrone (E1) by the enzyme estrone sulphatase is the major source of breast tumour estrogen. Inhibitors of estrone sulphatase should therefore have considerable therapeutic potential for the treatment of hormone-dependent tumours of the breast, either as the sole agent or in conjunction with aromatase inhibitors. Several inhibitors of estrone sulphatase have now been developed of which estra-1,3,5(10)-trien-17-one-3-sulphamate (EMATE) is the most potent and also inhibits the enzyme in a time- and concentration-dependent manner, showing that it acts as an irreversible inhibitor. Analogues of EMATE in which the 3-O-atom is replaced by other heteroatoms (S and N) were synthesized and tested for inhibition against estrone sulphatase. 4-Methoxyphenylsulphamide (1), 4-chlorothiophenyl-S-(N,N-dimethyl)sulphamate (2), estra-1,3,5(10)-trien-17-one-3-sulphamide (3), estra-1,3,5(10)-trien-17-one-3-S-sulphamate (4) and estra-1,3,5(10)-trien-17-one-3-S-(N,N-dimethyl)sulphamate (5) were found to inhibit estrone sulphatase weakly, but none of these compounds appears to behave as a time-dependent inhibitor. A model of the mechanism of enzyme inhibition by EMATE is proposed and we conclude that the sulphamate bridging oxygen atom of EMATE is essential for active site-directed inhibition of estrone sulphatase.

Chromatography, High Pressure Liquid↗
PubMed1996

Pharmacokinetics of oestrone-3-O-sulphamate.

The sulphatase pathway is thought to be the major route of oestrogen synthesis in breast tumours in postmenopausal women. There is currently considerable interest in developing a potent steroid sulphatase inhibitor to block oestrogen synthesis by this route. One of the most potent inhibitors discovered so far is oestrone-3-O-sulphamate (EMATE) which is active in vivo. In this study we report the preparation of a formulation for the administration of EMATE by the oral route. A method, using high-performance liquid chromatography (HPLC), was also established to measure concentrations of EMATE in rat plasma after its oral or i.v. administration. Using the oral formulation and HPLC assay, EMATE was readily detected in rat plasma after oral administration. Plasma EMATE concentrations were related to the dose of drug administered orally over the 10-40 mg/kg range. To examine the pharmacokinetics of EMATE, the compound (40 mg/kg, single dose) was administered either orally (in the formulation) or i.v. (in propylene glycol) with plasma samples being collected for up to 6 h. After oral administration, EMATE was rapidly absorbed, with the peak plasma concentration being detected at 30 min, after which plasma concentrations rapidly decreased. After i.v. administration a plasma EMATE concentration was detected at 1 h similar to that after oral administration. The clearance of EMATE from plasma followed a bi-phasic curve, showing an initial half-life of 30 min, followed by a slower half-life of 4 h 30 min. Little evidence was obtained for any metabolism of EMATE to oestrone. Rat liver sulphatase activity was almost completely inhibited (>99%) within 30 min of oral or i.v. administration of EMATE.

Animals↗
PubMed1996

Angiotensin-converting enzyme and enkephalinase in human breast cyst fluid.

Palpable breast cysts with an apocrine epithelial lining (type 1) are reported to be associated with a higher risk of developing breast cancer. The composition of breast cyst fluid (BCF) might include those factors involved in this increased risk. In this study peptidase activities that were active against the substrate [125I]metenkephalin-Arg-Phe were detected in BCF. The products were identified by reversed phase high-performance liquid chromatography (HPLC) as [125I]Tyr-Gly-Gly and [125I]Met-enkephalin. This proteolysis was not inhibited by PCMB, pepstatin A, leupeptin or aprotinin but was by EDTA, showing that the activity was due to metalloproteases. The production of [125I]Try-Gly-Gly was inhibited by phosphoramidon and thiorphan, whereas that of [125I]met-enkephalin was inhibited by captopril and Bothrops jararaca peptide, indicating that these activities are enkephalinase and angiotensin-converting enzyme (ACE) respectively. A fluorometric assay for ACE demonstrated that ACE levels are significantly higher in type 2 BCF than in type 1 BCF (30.8 vs 6.1 nmol hr-1 10 microliters-1, P < 0.001). As the increased risk of cancer is linked to type 1 cysts it is possible that higher levels of peptidase in type 2 BCF reflect a protective environment in the breast in which mitogenic peptide growth factors are neutralised by proteolysis.

Amino Acid Sequence↗
PubMed1996

SPARC is expressed in renal interstitial fibrosis and in renal vascular injury.

Tubulointerstitial inflammation and fibrosis are critical determinants for renal function and prognosis in a variety of human nephropathies. Yet, the pathophysiology of the injury remains obscure. We investigated the expression of SPARC (secreted protein acidic and rich in cysteine) by immunohistochemistry and in situ hybridization in experimental models characterized by tubulointerstitial fibrosis and matrix expansion in rats. SPARC is a secreted glycoprotein that has been demonstrated to affect cellular interaction with matrix proteins, modulate cell proliferation, bind to and/or inhibit growth factors such as PDGF and bFGF, and regulate angiogenesis. Interstitial expression of SPARC was most prominent in passive Heyman nephritis (PHN), chronic cyclosporine A (CsA) nephropathy, and the remnant kidney model and, to a lesser extent, in angiotensin II (Ang II)-infused animals. SPARC protein and mRNA were substantially increased at sites of tubulointerstitial fibrosis/matrix expansion. In the PHN model, SPARC protein was expressed by interstitial fibroblasts that also produced alpha-smooth muscle actin ("myofibroblasts") and correlated both temporally (r = 0.97) and spatially with sites of type I collagen deposition. Interstitial cell proliferation preceded the development of interstitial fibrosis, and maximal SPARC expression (d15) coincided with the initial decline in interstitial proliferation. In the Ang II-infusion model, which is characterized by arteriolopathy and tubulointerstitial injury, an increase in SPARC protein and mRNA was also seen in injured blood vessels. SPARC was shown to be expressed by vascular smooth muscle cells and also by cells in the adventitia of hypertrophied arteries. In summary, SPARC was transiently expressed by interstitial fibroblasts at sites of tubulointerstitial injury and fibrosis, and by smooth muscle cells and cells in the adventitia of injured arteries in the Ang II-model. In addition to its proposed role in extracellular matrix deposition. the antiproliferative properties of SPARC might contribute to the resolution of interstitial fibroblast proliferation in the PHN model.

Actins↗
PubMed1996

SPARC is expressed by mesangial cells in experimental mesangial proliferative nephritis and inhibits platelet-derived-growth-factor-medicated mesangial cell proliferation in vitro.

Mesangial cell proliferation is a characteristic feature of many glomerular diseases and often precedes extracellular matrix expansion and glomerulosclerosis. This study provides the first evidence that SPARC (secreted protein acidic and rich in cysteine) could be an endogenous factor mediating resolution of experimental mesangial proliferative nephritis in the rat. SPARC is a platelet-derived-growth-factor-binding glycoprotein that inhibits proliferation of endothelial cells and fibroblasts. We now show that SPARC is synthesized by mesangial cells in culture and that SPARC mRNA levels are increased by platelet-derived growth factor and basic fibroblast growth factor. Recombinant SPARC or the synthetic SPARC peptide 2.1 inhibited platelet-derived-growth-factor-induced mesangial cell DNA synthesis in vitro. In a model of experimental mesangioproliferative glomerulonephritis, SPARC mRNA was increased 5-fold by day 7 and was identified in the mesangium by in situ hybridization. Similarly, SPARC was increased in glomerular mesangial cells and visceral epithelial cells by day 5 and reached maximal expression levels by day 7. Mesangial cell proliferation increased by 36-fold on day 5 and decreased abruptly on day 7. Maximal expression of SPARC was correlated with the resolution of mesangial cell proliferation. We propose that SPARC functions in part as an endogenous inhibitor of platelet-derived-growth-factor-mediated mesangial cell proliferation in glomerulonephritis and that it could account for the resolution of cellular proliferation in this disease.

Amino Acid Sequence↗
PubMed1996

Regulation of aromatase and sulphatase in breast tumour cells.

In situ oestrogen synthesis makes an important contribution to the high oestrogen concentration found in breast tumours. Cytokines, including interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF alpha), have been shown to regulate aromatase activity in fibroblasts derived from normal and malignant breast tissues. In the present study, the ability of other cytokines in the IL-6 superfamily (IL-11 and oncostatin M) to stimulate aromatase activity has been confirmed. Formation of oestrone via the oestrone sulphatase pathway may be the major route of tumour oestrogen synthesis and in the present study TNF alpha was found to stimulate sulphatase activity in a dose-dependent manner. Human serum albumin was also found to be a potent stimulator of oestrone sulphatase activity. Its stimulatory effect was blocked by basic fibroblast growth factor, but not by several other growth factors tested. Insight into the regulation of oestrogen synthesis in breast tumours should enable the development of novel compounds to inhibit oestrogen synthesis in women with breast cancer.

Antineoplastic Agents, Hormonal↗
PubMed1995

In vivo inhibition of oestrone sulphatase and dehydroepiandrosterone sulphatase by oestrone-3-O-sulphamate.

Many tumours in endocrine-sensitive tissues, such as the breast and endometrium, are hormone-dependent and the hydrolysis of oestrone sulphate (EIS) to oestrone by oestrone sulphatase (EI-STS) is a major source of oestrogen in such tumours. Oestrone-3-O-sulphamate (EMATE) has been shown to be a potent EI-STS inhibitor in vitro, and in this study its ability to inhibit enzyme activity in vivo was examined. EMATE was initially administered to female rats for 7 days, after which liver EI-STS activity was measured. As EMATE also inhibits a related sulphatase in vitro, dehydroepiandrosterone sulphatase (DHA-STS), its effect on the activity of this enzyme in vivo was also investigated. DHA-STS has a pivotal role in regulating the synthesis of another steroid with potent oestrogenic properties, androstenediol. Administration of EMATE almost completely inhibited liver EI-STS (99%) and DHA-STS (99%) activities and was active when given by the oral or subcutaneous routes. After a single dose of EMATE or following the cessation of multiple doses for 10 days, liver EI-STS activity remained inhibited ( > 95%) for up to 7 and 10 days, respectively. Other compounds, such as 4-hydroxytamoxifen and the "pure" antioestrogen ICI 182,780, which are reported to inhibit EI-STS activity in vitro, did not inhibit activity in vivo. In a preliminary study, EMATE, when injected over a 12-day period, effectively reduced the growth of EIS-stimulated nitrosomethyl-urea-induced mammary tumours in ovariectomised rats and inhibited tumour sulphatase activity in treated animals.

Animals↗