Biomedical subjects
Publications and source records attributed to M A Brown.
The goal of selective targeting of enediyne cytotoxins has been investigated using estrogenic delivery vehicles. A series of estrogen-enediyne conjugates were assembled, and affinity for human estrogen receptor [hERalpha] was determined. The most promising candidate induced receptor degradation following Bergman cycloaromatization and caused inhibition of estrogen-induced transcription in T47-D human breast cancer cells.
gamma-Carboxylated proteins were detected in normal human pancreas by immunohistochemistry with a monoclonal antibody (M3B) specific for gamma-carboxyglutamyl residues. Staining appeared to be localized to the glucagon-secreting alpha-cells in the islets of Langerhans. Consistent with this, sections from a glucagonoma were stained much more intensely with the M3B antibody than those from an insulinoma. A murine alpha-cell line (alphaTC1 Clone 9) was cultured and gamma-carboxylated polypeptides, identified immunologically as prothrombin, protein S and (tentatively) Gas6, were isolated from the intracellular compartment by chromatography on an M3B-coupled resin. As in liver, prothrombin is synthesized by alpha-cells as a gamma-carboxylated zymogen that can be cleaved by ecarin to form an active serine protease that is inhibited by hirudin. The pancreas thus appears to be a novel site of synthesis for certain vitamin K-dependent proteins.
Ankylosing spondylitis (AS) is a common inflammatory arthritis predominantly affecting the axial skeleton. Susceptibility to the disease is thought to be oligogenic. To identify the genes involved, we have performed a genomewide scan in 185 families containing 255 affected sibling pairs. Two-point and multipoint nonparametric linkage analysis was performed. Regions were identified showing "suggestive" or stronger linkage with the disease on chromosomes 1p, 2q, 6p, 9q, 10q, 16q, and 19q. The MHC locus was identified as encoding the greatest component of susceptibility, with an overall LOD score of 15.6. The strongest non-MHC linkage lies on chromosome 16q (overall LOD score 4.7). These results strongly support the presence of non-MHC genetic-susceptibility factors in AS and point to their likely locations.
gamma-carboxylated polypeptides were detected in the human kidney by immunohistochemistry with a monoclonal antibody (M3B) specific for gamma-carboxyglutamyl residues. An approximately 70-kDa gamma-carboxylated protein, subsequently identified as prothrombin, was isolated from the intracellular compartment of cultured human embryonic kidney (HEK293) cells by immunoaffinity chromatography on M3B-coupled resin. Immunohistochemical analyses demonstrated that prothrombin and another vitamin K-dependent protein, the growth arrest-specific protein 6, were detectable in human kidney. As in the liver, the kidney synthesizes prothrombin as a zymogen that can be cleaved by ecarin to an amidolytically active serine protease that is inhibited by hirudin. This demonstrates for the first time the de novo synthesis of a full-length, gamma-carboxylated, and functional prothrombin gene product by human kidney cells.
OBJECTIVE: To assess the role of genes and the environment in determining the severity of ankylosing spondylitis. METHODS: One hundred seventy-three families with >1 case of ankylosing spondylitis were recruited (120 affected sibling pairs, 26 affected parent-child pairs, 20 families with both first- and second-degree relatives affected, and 7 families with only second-degree relatives affected), comprising a total of 384 affected individuals. Disease severity was assessed by the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and functional impairment was determined using the Bath Ankylosing Spondylitis Functional Index (BASFI). Disease duration and age at onset were also studied. Variance-components modeling was used to determine the genetic and environmental components contributing to familiality of the traits examined, and complex segregation analysis was performed to assess different disease models. RESULTS: Both the disease activity and functional capacity as assessed by the BASDAI and the BASFI, respectively, were found to be highly familial (BASDAI familiality 0.51 [P = 10(-4)], BASFI familiality 0.68 [P = 3 x 10(-7)]). No significant shared environmental component was demonstrated to be associated with either the BASDAI or the BASFI. Including age at disease onset and duration of disease as covariates made no difference in the heritability assessments. A strong correlation was noted between the BASDAI and the BASFI (genetic correlation 0.9), suggesting the presence of shared determinants of these 2 measures. However, there was significant residual heritability for each measure independent of the other (BASFI residual heritability 0.48, BASDAI 0.36), perhaps indicating that not all genes influencing disease activity influence chronicity. No significant heritability of age at disease onset was found (heritability 0.18; P = 0.2). Segregation studies suggested the presence of a single major gene influencing the BASDAI and the BASFI. CONCLUSION: This study demonstrates a major genetic contribution to disease severity in ankylosing spondylitis. As with susceptibility to ankylosing spondylitis, shared environmental factors play little role in determining the disease severity.
Type II [3H]estradiol binding sites play an important role in normal and malignant cell growth and proliferation and the delineation of the precise function of the type II site in cell growth has been hampered by the inability to purify, sequence and or clone this protein. The present manuscript describes methodology for the solubilization, purification and tentative identification of type II sites from the estrogen-treated rat uterus. This protein(s) chromatographs as a single major peak on DNA-cellulose, Affigel Blue dye affinity resin and during high performance liquid chromatography (HPLC) on hydroxyapatite. The purified fractions from these columns elicited classical [3H]estradiol binding characteristics (sigmoidal saturation curve, hyperbolic Scatchard plot, and Hill coefficient of approximately 4) for type II sites that are typically observed in crude or highly purified nuclear fractions or extracts. The type II binding activity also eluted as a single major component from a ligand affinity resin (GT-18-Sepharose) prepared by coupling 2,6-bis((3-methoxy-4-hydroxyphenyl)methylene)-cyclohexanone to epoxy-activated Sepharose (Pharmacia). The molecular weight of the type II site ([3H]estradiol binding activity) under non-denaturing or denaturing conditions was estimated to be approximately 10-15 kDa by gel filtration HPLC. Similarly, nuclear type II sites covalently labeled with the bioflavonoid, [3H]luteolin, migrated in the 10 kDa range on SDS PAGE. Thus, under these various experimental conditions, nuclear type II sites detected by [3H]estradiol or [3H]luteolin labeling techniques displayed little heterogeneity and appear much smaller than ERalpha or ERbeta or other steroid hormone receptors.
BACKGROUND: Managing resistant hypertension is difficult and mostly involves expensive testing seeking an underlying secondary cause. This study was undertaken to determine 1) the extent of the white-coat phenomenon in patients with resistant hypertension, and 2) whether 24-h ambulatory blood pressure (BP) monitoring (ABPM) or having BP recorded by a nurse instead of the referring doctor could clarify how many apparently resistant hypertensives actually have controlled BP. METHODS: This study involved 611 patients with BP > or = 140/90 mm Hg who were referred for 24-h ABPM by their specialist or general practitioner, including 277 patients who were taking no antihypertensives (group 1), 216 taking one or two antihypertensive drugs (group 2), and 118 taking at least three antihypertensives in combination (group 3). Each had BP recorded by one of two nurses before 24-h ABPM. Controlled BP was defined as awake ambulatory BP <135/85 mm Hg and the white-coat effect was the difference between the BP recorded by the referring doctor or nurse and the average awake ambulatory BP. RESULTS: Those with resistant hypertension (group 3) were older (61 years (12) v group 1: 46 years (14) and group 2: 56 (14) years; P < .001), but were of similar weight, height, and arm circumference to the other groups. Referral systolic, but not diastolic BP was higher in resistant hypertensives (mean 171/95 v 154/95 mm Hg and 164/94 mm Hg, respectively, P < .001 for systolic BP only). Twenty-eight percent of resistant hypertensives and 32% of those taking no antihypertensive drugs had normal awake ambulatory BP and the white-coat effect attributable to the referring doctor was always greater than that due to the nurse (range 16 to 26/12 to 14 mm Hg v 9 to 17/4 mm Hg, P < .001). Nurse recorded BP was highly sensitive (97%) in identifying awake hypertension but lacked specificity (57%) to replace ABPM. CONCLUSION: Our results show that approximately one in four patients with apparent resistant hypertension referred for ABPM have controlled BP and one-third of patients referred for initial evaluation of office or clinic hypertension have normal BP using ABPM, ie, white-coat hypertension. Twenty-four-hour ABPM appears an appropriate initial step before further investigating or treating patients with apparently resistant hypertension.
The role of the CTLA-4 antigen in the development of autoimmune diseases is well documented, with several autoimmune disorders showing association or linkage with the CTLA-4 locus. Its role in the aetiology of rheumatoid arthritis (RA) however, remains unclear, as the functional studies of the B7-CTLA-4 pathway in mouse models of RA and genetic studies in humans have given contrasting results. We have studied the single nucleotide polymorphism at position +49 (A/G) of the CTLA-4 gene, in a cohort of 421 RA cases and 452 healthy controls from the UK. Despite the high statistical power to detect even a weak susceptibility effect, no significant association was found. We also analysed the distribution of the allele and genotype frequencies with respect to the presence of the shared epitope (a known RA susceptibility factor) and found no statistically significant differences. We conclude that, although the importance of the B7-CTLA-4 interaction in the development of RA can not be excluded, the CTLA-4 gene is unlikely to be a predisposing factor to this disease.
The role of interleukin (IL)-4 as an important immunomodulatory cytokine is well established. IL-4 exhibits a highly restricted pattern of expression by cells of distinct lineages. The cell types that produce IL-4 are located in anatomically distinct locations (e.g. circulating T cells vs. fixed tissue mast cells) and thus have access to different IL-4-responsive target cells. In addition, these cells appear to regulate IL-4 expression in cell-type-specific ways. These findings suggest that an understanding of IL-4 gene regulation in T and mast cells could provide the means to specifically control IL-4 release in a lineage- and site-specific manner. In this article we review the current knowledge regarding the cell-type specific regulation of IL-4 gene expression in mast cells and compare this to what has been defined in T cells. We show that there are distinct yet parallel events that control developmentally determined chromatin modifications, allowing accessibility of the locus, and provide the potential for transcription. In differentiated cells, a subset of unique cell activation signals initiates the cascade of events that lead to transcriptional activation of the IL-4 gene.
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OBJECTIVE: This study was undertaken to document vascular reactivity in both phases of normal menstrual cycles and anovulatory cycles of women receiving oral contraceptives and to compare vascular reactivity of women in the follicular phase of the normal menstrual cycle with that of men of a similar age group. STUDY DESIGN: Forearm blood flow was measured with plethysmography in 15 women with ovulatory cycles during the follicular and luteal phases of the normal cycle, at equivalent times in 14 women receiving oral contraceptives, and in 15 age-matched men. Changes in forearm blood flow, an index of vascular reactivity, were measured during a randomly ordered brachial artery infusion schedule of norepinephrine, sodium nitroprusside, angiotensin II, and Nomega-monomethyl-L -arginine and were analyzed with repeated measures analysis of variance. RESULTS: Forearm blood flow responses were similar in the two phases of the normal cycle. Responses were similar between women with ovulatory cycles and users of oral contraceptives, apart from a significantly greater response to angiotensin II in the oral contraceptive group during the medication phase than during the withdrawal phase (P =.027). Basal forearm blood flow was higher in men than in women (7 +/- 1.6 vs 5 +/- 1.4 mL/100 mL per minute; P =.015), and men demonstrated a significantly greater vasoconstrictor response to norepinephrine (P <.05) than did women. CONCLUSION: Vascular reactivity is similar in the two phases of the normal menstrual cycle, but women who use oral contraceptives have an exaggerated response to angiotensin II during the medication phase. Men demonstrate greater vascular reactivity to norepinephrine than do women, perhaps because of the lack of estrogen-mediated protective effects and increased adrenergic activity.
OBJECTIVE: The purpose of this study was to determine the predictive ability of parameters of 24-hour ambulatory blood pressure monitoring for the development of preeclampsia or gestational hypertension in women who are already considered at risk for these disorders. STUDY DESIGN: One hundred twenty-two pregnant women who were considered high risk for the development of preeclampsia underwent 24-hour ambulatory blood pressure monitoring between 18 and 30 weeks gestation, while their condition was normotensive according to routine mercury sphygmomanometry. One hundred sixty-four healthy primigravid women who were considered at usual risk for preeclampsia underwent the same tests as a parallel study. Routine blood pressure, awake and sleep average blood pressure, and 24-hour mean average blood pressure were entered into multiple logistic regression as predictors of either preeclampsia or gestational hypertension; significant variables were then tested by a series of receiver operator curves. RESULTS: Eight percent of usual risk and 45% of high risk women experienced the development of preeclampsia or gestational hypertension. In both groups, the average routine mercury blood pressure and awake, sleeping, and 24-hour ambulatory blood pressure monitoring-derived blood pressure were significantly higher in women who later experienced the development of preeclampsia or gestational hypertension. In usual risk women, 24-hour systolic blood pressure of >or=115 mm Hg and sleeping systolic blood pressure of >or=106 mm Hg were predictive of later preeclampsia or gestational hypertension, but sensitivities were low (77% and 54%, respectively). In high risk women, sleeping diastolic blood pressure of >or=62 mm Hg and sleeping mean arterial pressure of >or=79 mm Hg were predictive of preeclampsia or gestational hypertension, but again sensitivities were low (70% and 65%, respectively). CONCLUSION: Awake and sleeping blood pressure are higher in midpregnancy in women who later experience the development of preeclampsia or gestational hypertension. Twenty-four-hour ambulatory blood pressure monitoring provides a noninvasive method of selecting some of these women, but this test has a sensitivity no better than that of other predictive tests, even in women at high risk for preeclampsia.
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OBJECTIVE: To document the diversity of definitions and diagnoses of the hypertensive disorders of pregnancy reported in major medical journals as a preliminary step from which future documents may be developed concerning the classification and diagnosis of the hypertensive disorders of pregnancy. METHODS: A systematic review was undertaken of articles published between 1997 and 1998 researching "preeclampsia" in nine internationally recognized journals. Each article was assessed to establish the features used by the authors to define preeclampsia and its major defining variables (i.e., proteinuria and hypertension). Documentation of aspects of the blood pressure measurement technique was also noted. RESULTS: One hundred thirty-five articles were studied, demonstrating a wide variety in the authors' definitions of preeclampsia, "significant" proteinuria, and hypertension. In up to 13% of articles, one or more of these terms was not defined and documentation of the blood pressure measurement technique was generally poor. The diversity of reporting was of such magnitude that the groups of women in these studies could rarely be considered truly comparable. CONCLUSION: Scientific and clinical research groups are studying different preeclamptic groups as a result of using different definitions of this condition. As comparison of results among studies is fundamental to the correct elucidation of knowledge about preeclampsia, standardization of the classification and diagnostic criteria of the hypertensive disorders of pregnancy should be a major priority of societies devoted to the study of these disorders.
OBJECTIVE: To determine (a) the prevalence of hypertension during sleep in pre-eclampsia and gestational hypertension, and (b) whether women with hypertension during sleep have worse pregnancy outcomes than hypertensive pregnant women with controlled (normal) blood pressure (BP) during sleep. DESIGN: Prospective double-blind cohort study. SETTING: Inpatients and outpatients managed in a day assessment unit (DAU) at St George Hospital, Sydney, Australia. PARTICIPANTS: A total of 186 hypertensive pregnant women, 158 of whom had successful 24 h BP monitoring; 40% had proteinuric pre-eclampsia (PE), 43% gestational hypertension (GH) and 17% essential hypertension (EH). INTERVENTIONS: Blood pressure, 24 h non-invasive, monitoring (Spacelabs 90207) was undertaken successfully in 158 women with PE, GH or EH, whether or not they were receiving antihypertensives. Women and clinicians were blinded to results of these BP monitors. Sleep hypertension was defined as BP > 117/68 mmHg at 26-30 weeks or > 123/72 mmHg after 30 weeks gestation. MAIN OUTCOME MEASURES: Maternal and fetal outcomes were compared between women with and without sleep hypertension and the prevalence of sleep hypertension was determined. RESULTS: Sleep hypertension was present in 59%, more commonly in PE (79%) than GH/EH (45%), P < 0.0001. Sleep hypertensives also had higher routine sphygmomanometer BPs [137(10)/91(7) mmHg; mean(SD)] than women with normal sleep BP [130(12)/ 87(8) mmHg] P = 0.007, and higher awake ambulatory blood pressure monitoring (ABPM) BPs [137(8)/88(7) versus 127(7)/79(6) mmHg], P < 0.0001. Awake, but not sleep, average heart rate was lower in sleep hypertensives [85(11) versus 91 (10) beats per minute, bpm], P = 0.002. Sleep hypertensives had a significantly greater frequency of renal insufficiency, liver dysfunction, thrombocytopenia and episodes of (awake) severe hypertension (P < 0.05), as well as lower birth weight babies [2715 (808) versus 3224(598) g, P < 0.0001]. CONCLUSIONS: Hypertension during sleep is a common finding in women with hypertensive disorders of pregnancy, particularly pre-eclampsia. These women also have higher awake BPs and a greater frequency of adverse maternal and fetal outcomes. These findings are largely explained by the greater likelihood of pre-eclamptics having sleep hypertension.
OBJECTIVE: To determine whether the insulin resistance syndrome and altered body composition are features of hypertensive pregnancy. DESIGN: Women were recruited in the third trimester of pregnancy from the antenatal clinic, day assessment unit, and maternity ward of St George Hospital, Sydney. POPULATION: Women with pre-eclampsia (n = 12), gestational hypertension (n = 12), essential hypertension in pregnancy (n = 11), and normotensive pregnancy (n = 10). METHODS: Energy metabolism was assessed by indirect calorimetry to measure basal metabolic rate and diet-induced thermogenesis. Body composition was measured as lean body mass, total body water and fat mass by bio-electrical impedance. Blood was collected for measurement of glucose, insulin and lipid profiles. Insulin resistance was indirectly assessed by the insulin and glucose concentrations and diet-induced thermogenesis. RESULTS: Women with essential hypertension and gestational hypertension were heavier than women with normotensive pregnancies both pre-pregnancy and in the third trimester, whereas women with pre-eclampsia were similar to those with normotensive pregnancy. Women with essential hypertension were otherwise similar to normotensive pregnancy but women with gestational hypertension had a reduced diet-induced thermogenesis and almost double insulin levels. Women with pre-eclampsia had a similar body composition and insulin levels but reduced basal metabolic rate, diet-induced thermogenesis and glucose levels compared with normotensive pregnancy. CONCLUSIONS: Women who develop gestational hypertension, but not pre-eclampsia, are more likely to be overweight. Women with essential hypertension are similar to women with normotensive pregnancy throughout pregnancy. Both gestational hypertension and pre-eclampsia appear to be associated with some degree of insulin resistance, greater than that occurring in normal pregnancy.
The objective of the study was to measure energy metabolism and body composition during pregnancy and postpartum, compared to non-pregnant women, using non-invasive techniques. A longitudinal study of eight normotensive pregnant women was carried out at 19 +/- 1 and 36+/-1 weeks gestation, and postpartum. A cross-sectional study was also performed comparing postpartum to 12 non-pregnant women. Indirect calorimetry was performed while fasting to measure basal metabolic rate (BMR) and postprandially to measure diet-induced thermogenesis (DIT). Body composition consists of fat mass, lean body mass (LBM), and total body water (TBW) and was measured by bio-electrical impedance. Insulin resistance was indirectly assessed by glucose and insulin concentration and DIT.