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Biomedical subjects

Y Zhao

Publications and source records attributed to Y Zhao.

At least 703 records · Page 39Linked to original sources
PubMed1997

Chemiluminescence determination of tiopronin by flow injection analysis based on cerium(IV) oxidation sensitized by quinine.

A flow injection analysis method is proposed for the determination of tiopronin based upon the oxidation by cerium(IV) in dilute sulfuric acid medium and sensitized by quinine. With the peak height as a quantitative parameter applying optimum working conditions, tiopronin is determined over the 1-400 microM range (150 microliters per injection, n = 10, r = 0.9994) with a detection limit of 0.34 microM and an RSD (n = 10) less than 2% at 20 and 50 microM. The proposed method, combining the advantages of speed and sensitivity, was applied to the routine determination of tiopronin in a pharmaceutical preparation.

Cerium↗
PubMed1997

Genetic testing for male infertility: a postulated role for mutations in sperm nuclear matrix attachment regions.

Numerous reports have suggested that disturbances in nuclear condensation may result in male infertility. This notion has been supported by the observation of infertile individuals with a decrease or absence of the male sperm-specific chromatin packaging protamine proteins. To date, no correlation between the absence of protamine proteins and a mutation within the coding regions of the protamine genes has been documented. To address this issue, PCR-based mutation scanning analysis has been performed across the human male haploid expressed PRM1-->PRM2-->TNP2 domain in several oligozoospermic infertile individuals. This analysis identified a candidate mutation in a region of contact with the sperm nuclear matrix from 2 of 5 affected individuals. This is the first report of a mutation scan covering the entire PRM1-->PRM2-->TNP2 locus in affected individuals.

Chromosomal Proteins, Non-Histone↗
PubMed1997

The expression of transforming growth factor-beta s and TGF-beta receptor mRNA and protein and the effect of TGF-beta s on human myometrial smooth muscle cells in vitro.

In this study we investigated the expression of transforming growth factor-beta (TGF-beta) isoform and TGF-beta receptor mRNA and protein, and the effect of TGF-beta 1-3 on the rate of DNA synthesis and proliferation of human myometrial smooth muscle cells in vitro. To determine these, we utilized primary cultures of myometrial smooth muscle cells, standard and competitive quantitative reverse transcription-polymerase chain reaction (RT-PCR), immunocytochemistry, enzyme-linked immunoassay, radioreceptor assay, [3H] thymidine incorporation and cell proliferation assay. Standard RT-PCR and immunocytochemistry revealed that myometrial smooth muscle cells express TGF beta 1-3 and TGF-beta type I-III receptor (TGF-beta R) mRNA and protein. Quantitative RT-PCR, using an external synthetic RNA standard, indicated that the cells express 10 copies/cell of TGF-beta 1 and TGF-beta 2, less than one copy/cell of TGF-beta 3 and TGF-beta type IR, three copies/cell of type IIIR, and > 200 copies/cell, of TGF-beta type IIR mRNA. The cells also synthesized and released TGF-beta 1 at the rate of 7.8 +/- 0.7 ng/10(6) cells, of which 1.4 +/- 0.2 ng/10(6) cells was in an active form. The rate of [3H] thymidine incorporation or proliferation of subconfluent quiescent smooth muscle cells was not altered by TGF-beta s (0.1-10 ng/ml) under serum-free conditions, nor in the presence of 10% fetal bovine serum (FBS). TGF-beta 1-3 at 0.25-0.5 ng/ml in the presence of 2% FBS, which induces half maximal stimulation of these cells, stimulated the rate (P < 0.05), whereas at higher doses it reduced the rate of [3H]-thymidine incorporation compared to the controls. The effect of TGF-beta was partially reversible using neutralizing antibodies specific to TGF-beta 1, TGF-beta 2 (10 micrograms/ml) or TGF-beta 3 (3-6 micrograms/ml). TGF-beta s had no significant effect on cell proliferation determined by cell counting. The data indicate that human myometrial smooth muscle cells express the necessary components of the TGF-beta system, suggesting an autocrine/paracrine role for TGF-beta s in myometrium.

Animals↗
PubMed1997

Neonatal hypothyroidism alters the pattern of prostatic growth and differentiation, as well as plasminogen activator and metalloprotease expression, in the rat.

The purpose of this study was to examine whether changes in growth, ductal histology, and expression of plasminogen activator (PA) and matrix metalloproteases are associated with the increased prostatic weight and DNA content seen in adult rats that were treated neonatally with the goitrogen 6-propyl-2-thiouracil (PTU). Ventral prostatic weights were initially reduced in PTU-treated rats but were increased 40% over those of controls by Day 180; this increase in prostatic weight was also accompanied by increases in the number of prostatic ductal tips. In controls, prostatic PA and gelatinase A activities decreased after completion of morphogenesis at 21-28 days of age. In contrast to controls, PA and gelatinase A activities were maintained through puberty (42 days) in PTU-treated rats but declined by 90 days. The elevated PA activity in both prostatic lobes at 42 days of age in PTU-treated rats was inhibited by amiloride, indicating that it is the urokinase form of PA. These data show that the increased prostatic weight and DNA content in adult rats following neonatal PTU treatment results from a delayed but extended period of growth and the formation of new ductal elements. There is a temporal overexpression of urokinase and gelatinase A associated with the increased ductal branching, indicating as well an extended period of morphogenesis that results in their eventual increased adult size. The prostatic enlargement in PTU-treated rats may serve as a useful model to study regulation of both normal and abnormal prostatic growth and morphogenesis.

Amiloride↗
PubMed1997

Localization of lipid peroxidation-derived protein epitopes in the porcine corpus luteum.

Reactive oxygen species (ROS) generated within the corpus luteum (CL) are believed to play an integral role in luteolysis. Unsaturated lipids are susceptible to peroxidation by ROS, leading to the formation of toxic aldehydes such as malondialdehyde (MDA) and 4-hydroxy-nonenal (4-HNE) that can attack cellular and extracellular proteins. Aldehyde conjugation of proteins can have diverse effects on cell function, including cross-linking of cell surface receptors, alteration of enzyme activities, and modification of lipoproteins. The objectives of the present study were to 1) determine by immunological techniques whether lipid peroxidation-derived protein epitopes (i.e., MDA- and 4-HNE-lysine) could be detected in situ during the natural life span of the CL, and 2) determine the temporal and spatial localization of these epitopes. Fresh luteal tissue was collected from young gilts at distinct phases of the estrous cycle: early (Days 4-7), mid (Days 8-12), and late (Days 14-18). Immunocytochemistry was performed on tissue sections using monoclonal antibodies against MDA-lysine and 4-HNE-lysine. Positive immunostaining was evident in porcine CL at all stages of the estrous cycle. Immunoactivity was associated primarily with the steroidogenic cells, most consistently with the large luteal cells, and was most intense in regressing tissue (i.e., late). No staining was observed in sections incubated with nonimmune mouse serum or when anti-4-HNE-lysine was preadsorbed with 4-HNE-protein preparations. Immunoblotting of protein extracts from luteal tissue revealed three major reactive bands. At least two of these bands were closely associated with extracted oxidized human low-density lipoprotein (LDL) and with 4-HNE-conjugated human apoprotein B. These results demonstrate the presence of lipid peroxidation-derived protein epitopes within the porcine CL and show that one or more of these may be an oxidized-LDL moiety. This represents the first direct in situ evidence that protein modification by lipid peroxidation products occurs during the natural luteal life span. The physiological consequences remain to be determined.

Aldehydes↗
PubMed1997

Cytochromes P450 11: expression of the CYP19 (aromatase) gene: an unusual case of alternative promoter usage.

Family 19 of the P450 super family is responsible for the conversion of C19 androgenic steroids to the corresponding estrogens, a reaction known as aromatization because it involves conversion of the delta4-3-one A-ring of the androgens to the corresponding phenolic A-ring characteristic of estrogens. The gene encoding human aromatase has been cloned and characterized and shown to be unusual compared to genes encoding other P450 enzymes, because there are numerous untranslated first exons that occur in aromatase transcripts in a tissue-specific fashion due to differential splicing as a consequence of the use of tissue-specific promoters. Thus, expression in the ovary uses a proximal promoter that is regulated primarily by cAMP. On the other hand, expression in the placenta uses a distal promoter located at least 40 kb upstream of the start of transcription that is regulated by retinoids. Other promoters are used in brain and adipose tissue. In the latter case, class I cytokines such as IL-6 and IL-11, as well as TNF-alpha, are important regulatory factors. A common 3'-splice junction located upstream of the start of translation is used in all of the splicing events involved in the use of these various promoters. Thus, the coding region of the transcripts, and hence the protein, are identical regardless of the tissue site of expression; what differs in a tissue-specific fashion is the 5'-end of the transcripts. This pattern of expression has great significance both from a phylogenetic and ontogenetic standpoint, as well as for the physiology and pathophysiology of estrogen formation, as will be discussed in this review.

Adipose Tissue↗
PubMed1997

Tumor necrosis factor-alpha-angiotensin interactions and regulation of blood pressure.

OBJECTIVES: To compare the levels of tumor necrosis factor-alpha (TNF) produced by medullary thick ascending limb tubules (MTAL) obtained from normotensive and angiotensin II (Ang II)-dependent hypertensive rats and determine whether TNF participates in a mechanism that opposes elevation of blood pressure by Ang II. DESIGN: We have previously demonstrated that in-vitro administration of Ang II increases production of TNF and prostaglandin E2 (PGE2) by the MTAL. We hypothesize that production of TNF and PGE2 by the MTAL is elevated in in-vivo models of Ang II-dependent hypertension and acts to modulate the pressor effects of Ang II. Thus, inhibition of TNF should disclose whether this cytokine acts to modulate Ang II-induced hypertension. METHODS: MTAL tubules obtained from normotensive and Ang II-dependent hypertensive rats were isolated by enzymatic digestion and sieving. Tubules were cultured in the absence of exogenous Ang II. TNF and PGE2 levels were measured by enzyme-linked immunosorbent assay. Anti-TNF antiserum was administered intravenously to normotensive and Ang II-dependent hypertensive rats and their mean arterial pressures were measured. RESULTS: Production of TNF and PGE2 was significantly greater in MTAL tubules isolated from Ang II hypertensive rats than it was in those from normotensive controls. Administration of anti-TNF antiserum exacerbated the Ang II-mediated increase in mean arterial pressure. CONCLUSIONS: The higher levels of production of TNF and PGE2 by MTAL tubules isolated from Ang II hypertensive rats compared with those of normotensive controls are consistent with results of in-vitro experiments showing that administration of Ang II increases production of TNF and PGE2 by the MTAL. TNF and PGE2 participate in a counter-regulatory mechanism that opposes the pressor actions of Ang II.

Angiotensin II↗
PubMed1997

Inspiratory and expiratory steady flow analysis in a model symmetrically bifurcating airway.

Steady inspiratory and expiratory flow in a symmetrically bifurcating airway model was studied numerically using the finite element method (FIDAP). Flows of Reynolds number of 500 and 1000 during inspiration and a flow of Reynolds number of 500 during expiration were analyzed. Since the geometry of the bifurcation model used in this study is exactly the same as the model used in the experimental studies, the computed results were compared to the experimental findings. Results show that most of the important flow features that were observed in the experiment, such as the skewed velocity profiles in the daughter branches during inspiration and velocity peak in the parent tube during expiration, were captured in the numerical simulation. Quantitatively, the computed velocity profiles are in good agreement with the measured profiles. This comparison validates the computational simulations.

Bias↗
PubMed1997

Identification of envelope protein residues required for the expanded host range of 10A1 murine leukemia virus.

The 10A1 murine leukemia virus (MuLV) is a recombinant type C retrovirus isolated from a mouse infected with amphotropic MuLV (A-MuLV). 10A1 and A-MuLV have 91% amino acid identity in their envelope proteins yet display different host ranges. For example, CHO-K1 cells are resistant to A-MuLV but susceptible to infection by 10A1. We have now determined that retroviral vectors bearing altered A-MuLV envelope proteins containing 10A1-derived residues at positions 71 (A71G), 74 (Q74K), and 139 (V139M) transduce CHO-K1 cells at efficiencies similar to those achieved with 10A1 enveloped vectors. A-MuLV enveloped retroviral vectors with these three 10A1 residues were also able to transduce A-MuLV-infected NIH 3T3 cells. This observation is consistent with the ability of vectors bearing this altered A-MuLV envelope protein to recognize the 10A1-specific receptor present on NIH 3T3 cells and supports the possibility that residues at positions 71, 74, and 139 of the 10A1 envelope SU protein account for the expanded host range of 10A1.

3T3 Cells↗
PubMed1997

The ICP0 protein of equine herpesvirus 1 is an early protein that independently transactivates expression of all classes of viral promoters.

To assess the role of the equine herpesvirus type 1 (EHV-1) ICP0 protein (EICP0) in gene regulation, a variety of molecular studies on the EICP0 gene and gene products of both the attenuated cell culture-adapted Kentucky A (KyA) strain and the Ab4p strain were conducted. These investigations revealed that (i) the ICP0 open reading frame (ORF) of the KyA virus strain is 1,257 bp in size and would encode a protein of 419 amino acids, and in comparison to the ICP0 gene (ORF63) of the Ab4p strain of 1,596 bp (E. A. Telford, M. S. Watson, K. McBride, and A. J. Davison, Virology 189:304-316, 1992), it has an internal in-frame deletion of 339 bp; (ii) one early transcript of 1.4 kb predicted to encode the EICP0 protein and a late transcript of 1.8 kb are detected in Northern blot analyses using probes containing the EICP0 ORF; (iii) the KyA EICP0 protein (50 kDa) and the Ab4p EICP0 protein (80 kDa) are expressed as several species of early proteins that are first detected at 3 to 4 h postinfection by Western blot analyses of infected-cell polypeptides, using an antiserum generated to a TrpE fusion protein that harbors amino acids 46 to 153 of the EICP0 protein; and (iv) the EICP0 protein of both EHV-1 strains is a potent transactivator of EHV-1 genes. Transient expression assays using a simian virus 40 expression construct of the EICP0 protein of the KyA strain showed that the EICP0 protein independently transactivated chloramphenicol acetyltransferase reporter constructs under the control of the immediate-early promoter (3.9-fold), the early thymidine kinase promoter (95-fold), the late (gamma1) IR5 promoter (85-fold), and the late (gamma2) glycoprotein K promoter (21-fold). The finding that the EICP0 protein of the KyA virus can function as an activator of gene expression indicates that amino acids corresponding to residues 319 to 431 of the Ab4p EICP0 protein are not essential for EICP0 transactivation of EHV-1 promoters.

Amino Acid Sequence↗
PubMed1997

Functional interactions between monomers of the retroviral envelope protein complex.

Retroviral vectors have been widely used in human gene therapy protocols. Entry into target cells is directed by the retroviral envelope protein, with receptor binding and postbinding fusion functions contributed mainly by the SU and TM subunits, respectively. We have generated mutants of the Moloney murine leukemia virus (MoMuLV) envelope protein with mutations in both the receptor binding domain of SU and throughout the TM subunit that are functionally inactive when expressed individually. However, the coexpression of these two classes of mutants partially restores envelope protein function and allows transduction. Several lines of evidence indicate that this complementation occurs in trans within envelope protein heterooligomers. The finding that the binding and postbinding functions of a retroviral envelope protein can be contributed by two different monomers should assist in the engineering of envelope proteins for tissue-specific gene delivery.

3T3 Cells↗
PubMed1997

Homozygosity by descent for a rare mutation in the myophosphorylase gene is associated with variable phenotypes in a Druze family with McArdle disease.

We examined a large consanguineous Druze family with McArdle disease for mutations in the glycogen myophosphorylase (PYGM) gene. All affected subjects were autozygous for a single G to A transition that abolishes the 5' consensus splice site in the first nucleotide of intron 14. The G to A transition is a rare mutation, with only one previous report in a single white subject heterozygous for this mutation and another, more common, mutation at codon 49. The kindred in our study is the first family reported in which disease is caused by homozygosity for this rare mutation. This kindred was originally reported as the first familial case of McArdle disease in the Druze.

Cell Line, Transformed↗
PubMed1997

Expression of transforming growth factor-beta receptors during hyperoxia-induced lung injury and repair.

Lung injury and repair processes involve many cellular activities, including cell growth, differentiation, and remodeling of extracellular matrix components. Transforming growth factor-beta (TGF-beta) is a major class of signaling peptide growth factors regulating these cellular activities. Type I (T beta RI) and type II (T beta RII) receptors for TGF-beta are transmembrane serine/threonine kinases that are essential for TGF-beta signaling. To gain insight into the possible molecular mechanisms of lung injury and repair, we investigated the expression of T beta RI and T beta RII in an acute hyperoxia-induced model of lung injury and repair. Localization of message expression of T beta RI and T beta RII in oxygen-exposed rat lung tissue was analyzed by using in situ hybridization. T beta RI mRNA expression was found in the interstitium, capillaries, and the alveolar septa of rat lungs exposed for 60 h to 100% oxygen. The distribution of T beta RII mRNA in oxygen-exposed rat lung tissue overlapped the localization of T beta RI mRNA. Temporal changes of T beta RI and T beta RII mRNA expressions in rat lung during hyperoxic exposure and repair were examined by Northern analysis. We found that expression of T beta RI was upregulated in adult rats undergoing prolonged exposure to 100% oxygen, and the increase of T beta RI expression persisted during 2 wk of repair of lung injury. The pattern of T beta RII expression during hyperoxic exposure and repair was distinct from that of T beta RI. The expression of T beta RII increased with a peak at 3 days postexposure and then declined after 7 days of repair. Changes of T beta RI and T beta RII protein expressions in rat lung during hyperoxic exposure and repair were examined further by Western blot analysis, which correlated with the mRNA expression. The results suggest that T beta RI and T beta RII may play important roles during the lung injury and repair by mediating signaling activity of TGF-beta and may regulate interactions between the mesenchyme and the epithelium.

Animals↗
PubMed1997

High altitude-induced changes in alpha1-adrenergic receptors and Ins(1,4,5)P3 responses in cerebral arteries.

In response to high-altitude long-term hypoxemia, the cerebral arteries of fetal and adult sheep show decreased contractile responses to norepinephrine (NE) and other agonists. To test the hypothesis that hypoxia-induced developmental and vessel specific cerebral artery contractility changes are mediated, in part, by changes in alpha1-adrenergic receptor (alpha1-AR) density and/or NE-induced inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] responses, we performed the following study. In common carotid (Com) and main branch cerebral (MBC) arteries from normoxic adult ewes and near-term fetuses and those acclimatized to high altitude (3,820 m), we quantified alpha1-AR density (maximal binding in fmol/mg protein) and affinity (dissociation constant in nM) with the alpha1-AR antagonist [3H]prazosin. In addition, we quantified NE-induced Ins(1,4,5)P3 responses in these arteries. With long-term hypoxemia, alpha1-AR density in fetal and adult Com decreased 75% (from 113 +/- 18 to 28 +/- 5 fmol/mg protein) and 66% (from 54 +/- 3 to 18 +/- 4 fmol/mg protein), respectively, from normoxic control values. alpha1-AR density of the fetal and adult MBC decreased 76% (from 47 +/- 4 to 11 +/- 1 fmol/mg protein) and 61% (from 23 +/- 3 to 9 +/- 3 fmol/mg protein), respectively, from controls. In hypoxemic adult Com, the NE-induced Ins(1,4,5)P3 response decreased 51% (from 309 +/- 38 to 151 +/- 24%) from the control value. In fetal and adult MBC, long-term hypoxemia was associated with decreases of 35% (from 345 +/- 40 to 225 +/- 30%) and 44% (from 355 +/- 55 to 199 +/- 16%), respectively, from control values. We conclude that in the adult Com and MBC vessels, acclimatization to high-altitude, long-term hypoxemia was associated with significant decreases in both alpha1-AR density values and Ins(1,4,5)P3 responses to NE. Similarly, in the fetal MBC arteries, high-altitude hypoxemia was associated with marked attenuation of both alpha1-AR density and NE-induced Ins(1,4,5)P3 responses. The magnitude of decreases in NE-induced Ins(1,4,5)P3 responses in these vessels correlated fairly well with the decreases in alpha1-AR density. These findings suggest that changes in noradrenergic receptor-second messenger coupling may play a role in altered cerebrovascular tone in association with high-altitude acclimatization and other forms of long-term hypoxia in both fetus and adult.

Acclimatization↗
PubMed1997

Ins(1,4,5)P3 receptors in cerebral arteries: changes with development and high-altitude hypoxia.

We and others have shown that adrenergic-mediated contractile responses in cerebral vessels in vitro differ with vessel segment, with developmental age, and with high-altitude, long-term hypoxia. This is associated with significant differences in alpha 1-adrenergic receptor density and norepinephrine (NE)-induced response of the second messenger inositol 1,4,5-trisphosphate [Ins(1,4,5)P3]. To test the hypothesis that vessel-specific, developmental, and hypoxic-associated contractility changes are mediated, in part, by changes in Ins(1,4,5)P3-receptor [Ins(1,4,5)P3-R] density or affinity, we performed the following study. In common carotid (Com), circle of Willis, and main branch anterior, middle, and posterior cerebral arteries (MBC) from normoxic fetal (approximately 140 days), newborn (3-5 days), and adult sheep and fetal and adult sheep acclimatized to high altitude, we quantified Ins(1,4,5)P3-R with [3H]Ins(1,4,5)P3. In normoxic Com, Ins(1,4,5)P3-R density values (fmol/mg protein) in fetus, newborn, and adult were 8 +/- 53, 150 +/- 18, and 357 +/- 21, respectively (P < 0.05). In normoxic MBC cerebral arteries, the receptor density values in the three age groups were 115 +/- 15, 105 +/- 9, 99 +/- 5 fmol/mg protein, respectively. For fetal and adult Com, high-altitude, long-term hypoxemia was associated with decreases in Ins(1,4,5)P3-R density of 32 (to 58 +/- 5) and 70% (to 109 +/- 12), respectively, from control values (P < 0.01). In MBC cerebral arteries of fetus and adult, hypoxic-associated decreases in Ins(1,4,5)P3-R density from control were 80 (to 23 +/- 3) and 47% (to 53 +/- 7), respectively (P < 0.01). Ins(1,4,5)P3 binding affinity to the receptor averaged 11.8 +/- 0.5 nM and did not vary significantly as a function of vessel type, developmental age, or hypoxia. In Com, but not in MBC, Ins(1,4,5)P3-R density increased dramatically with developmental age. This suggests that differences in Ins(1,4,5)P3-R density values may account, in part, for differences in contractile responses of the two artery types in the several age groups. In response to long-term, high-altitude hypoxia, Ins(1,4,5)P3-R density values in both fetal and adult Com and MBC decreased significantly, as did their NE-induced contraction. This suggests a cellular basis for changes in cerebrovascular contractility in response to long-term hypoxia and that Ins(1,4,5)P3-R may play a role in acclimatization responses to high altitude.

Aging↗
PubMed1997

Estrogen biosynthesis in THP1 cells is regulated by promoter switching of the aromatase (CYP19) gene.

The expression of aromatase, the enzyme responsible for estrogen biosynthesis, has been studied in THP-1 cells of human mononuclear leukemic origin, which exhibit high rates of aromatase activity. These cells have the capacity to differentiate in the presence of vitamin D into cells with osteoclast-like properties. Differentiated cells displayed higher rates of aromatase than undifferentiated cells, and, in both cases, activity was stimulated 10- to 20-fold by dexamethasone. Phorbol esters also increased aromatase activity, but the effect was the same in differentiated as in undifferentiated cells. In a similar fashion to adipose stromal cells, serum potentiated the response to dexamethasone but had no effect on phorbol ester-stimulated activity. By contrast to its action in adipose stromal cells, (Bu)2cAMP markedly inhibited aromatase activity of THP-1 cells, as did factors whose actions are mediated by cAMP, such as PTH and PTH-related peptide. This was true of control cells, as well as of dexamethasone- and phorbol ester-stimulated cells. Previously we have shown that type 1 cytokines as well as tumor necrosis factor-alpha stimulate aromatase activity of adipose stromal cells in the presence of dexamethasone. By contrast, interleukin-6, interleukin-11, and leukemia-inhibitory factor had no effect on aromatase activity of THP-1 cells, whereas tumor necrosis factor-alpha, oncostatin M, and platelet-derived growth factor were slightly inhibitory of aromatase activity. Exon-specific Southern analysis of rapid amplification of cDNA ends-amplified transcripts was employed to examine the distribution of the various 5'-termini of aromatase transcripts. In the control group, most of the clones contained transcripts specific for the proximal promoter II, whereas in dexamethasone-treated cells, most transcripts contained exon I.4. In the phorbol ester-treated cells, a broader spectrum of transcripts was present, with equal proportions of I.4, II, and I.3-containing clones. Additionally, one clone containing a new sequence, exon I.6, was found. This was shown to be located about 1 kb upstream of exon II. By contrast, all clones from cells treated with (Bu)2cAMP contained promoter II-specific sequences. In addition to these transcripts, two clones in the library from the dexamethasone-treated cells contained the sequence previously defined as the brain-specific sequence, 1f. In one of these, the 1f sequence was fused downstream of exon I.4, indicative that its expression likely employed promoter I.4. These results point to similarities and important differences between aromatase expression in THP-1 cells and other cells such as adipose stromal cells, indicative of unique regulatory pathways governing aromatase expression in these cells.

Aromatase↗
PubMed1997

Prostaglandin E2 stimulates aromatase expression in endometriosis-derived stromal cells.

C19 steroids are converted to estrogens by aromatase P450 (P450arom). Aromatase expression in humans is regulated by use of tissue-specific promoters in the placenta (promoter I.1), adipose tissue (promoters I.4, I.3, and II), and gonads (promoter II). The use of each promoter gives rise to a population of P450arom messenger ribonucleic acid (mRNA) species with a unique untranslated 5'-terminus. Aromatase is not expressed in the endometrium of disease-free women. We demonstrated, however, the presence of P450arom mRNA in pelvic endometriotic implants and eutopic endometrial curettings of women with endometriosis. In the current report, aromatase activity and P450arom gene expression were investigated in cultured stromal cells derived from eutopic endometrium and ovarian endometriomas of women with pelvic endometriosis. We also investigated the hormonal regulation of aromatase expression and alternative promoter use in these cells. The effects of interleukin-1 beta (IL-1 beta), IL-2, IL-6, IL-11, oncostatin M, IL-15, tumor necrosis factor-alpha, PGE2, estradiol, R5020, dexamethasone, and dibutyryl cAMP (Bt2cAMP) on aromatase activity in endometriosis-derived stromal cells were assessed. We chose treatments with PGs and ILs because of the inflammatory nature of endometriosis. PGE2 stimulated aromatase activity in endometriosis-derived stromal cells by 19- to 44-fold (37-221 pmol/mg protein-4 h), whereas Bt2cAMP induction was 26- to 60-fold the baseline level. No stimulation was observed by estradiol or R5020 or by IL-1 beta, IL-2, IL-6, IL-11, IL-15, or TNF alpha in the presence or absence of glucocorticoids. A modest induction of aromatase activity (2-fold) was observed in dexamethasone- plus oncostatin M-treated cells. These changes in aromatase activity were accompanied by comparable changes in the levels of P450arom mRNA levels, determined by a quantitative reverse transcription-PCR method. Promoter-specific 5'-ends of P450arom transcripts in total RNA from endometriosis-derived stromal cells treated with PGE2 and Bt2cAMP were amplified employing a novel modified rapid amplification of cDNA5'-ends/Southern hybridization method using exon-specific oligonucleotide probes. The majority of P450arom transcripts in these cells contained the gonadal-type promoter II-specific sequences, whereas very few transcripts contained adipose-type promoter I.3- and I.4-specific sequences. PGE2 appears to be the most potent known stimulator of aromatase in endometriosis. Aromatase expression in PGE2-stimulated stromal cells of endometriosis is regulated primarily by the classically located promoter II, which, in turn, is regulated by cAMP. As PGE2 is known to increase intracellular cAMP levels, estrogen biosynthesis in endometriosis may be primarily regulated by PGE2 that is locally produced. Consequent local estrogen production may promote the growth of endometriotic implants.

Aromatase↗