Biomedical subjects

I Choi

Publications and source records attributed to I Choi.

At least 73 records · Page 4Linked to original sources

PubMed1998

Effects of acanthoic acid on TNF-alpha gene expression and haptoglobin synthesis.

Tumour necrosis factor-alpha (TNF-alpha) is a major pro-inflammatory cytokine inducing the synthesis and release of many inflammatory mediators. It is involved in immune regulation, autoimmune diseases, and inflammation. Our previous study demonstrated that acanthoic acid, (-)-pimara-9(11), 15-dien-19-oic acid, a pimaradiene diterpene isolated from Acanthopanax koreanum, inhibited TNF-alpha production. To extend our understanding of inhibitory effects of acanthoic acid on TNF-alpha production, its effects on TNF-alpha gene expression was tested. Based on the results from RT-PCR and promoter analysis of TNF-alpha, it was found that acanthoic acid suppressed TNF-alpha gene expression. But the same concentration of acanthoic acid had no effect on IL-6 gene expression. Haptoglobin is an acute phase protein which is induced by TNF-alpha. When liver cells were treated with acanthoic acid, haptoglobin synthesis was blocked by acanthoic acid. These data confirmed that acanthoic acid inhibited gene expression and biological function of TNF-alpha.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed1998

Adult T-cell leukemia-lymphoma successfully treated with 2-chlorodeoxyadenosine.

Adult T-cell leukemia-lymphoma (ATL) is resistant to currently available chemotherapy and has a poor prognosis. We describe here a patient with ATL successfully treated with 2-chlorodeoxyadenosine (2-CdA). A 75-year-old Japanese male with an acute type of ATL, who had become resistant to the initial cytotoxic chemotherapy, was treated with 2-CdA administered by continuous drip infusion of 0.09 mg/kg/d for seven consecutive days in one month (one cycle). After three cycles of treatment, partial remission (PR) was achieved. Surprisingly, 249 days after the administration of 2-CdA, ATL cells completely disappeared from the peripheral blood. PR was maintained during 10 weeks until evidence of a new lymphadenopathy. No remarkable toxicity of 2-CdA occurred.

Aged↗

PubMed1998

Human interleukin 6 gene is activated by hepatitis B virus-X protein in human hepatoma cells.

Interleukin 6 (IL-6) is a pleiotropic cytokine that induces many biological activities, including some aspects of the immune reaction and inflammatory responses. In the liver, IL-6 regulates the synthesis of a broad spectrum of acute-phase proteins. IL-6 is also known to be a factor involved in the immunoregulatory perturbations in patients with chronic liver diseases (CLDs). Here, we report that IL-6 can be induced by hepatitis B virus (HBV)-X protein, as evidenced by high levels of serum IL-6 in patients with CLD with HBV infection, IL-6 productions observed in HBV-X-transfected cells, and transcriptional transactivations of the IL-6 gene by HBV-X. We determined serum levels of IL-6 in patients with chronic hepatitis B (CH-B), chronic hepatitis C (CH-C), liver cirrhosis (LC) caused by hepatitis B, and LC with hepatocellular carcinoma (HCC) caused by hepatitis B (LC+HCC). Mean serum levels of IL-6 in all CLD patients were higher than those in normal controls, and the difference was statistically significant (P < 0.05). Mean IL-6 levels of LC and LC+HCC patients were significantly higher than those of CH-B patients (P < 0.05). Because the etiological factor in all cases except CH-C (CH-B, LC, and LC+HCC) was HBV, we checked the possibility of HBV-transactivator-X activation of IL-6 promoter. Using deletion constructs of 5'-flanking regulatory regions of the IL-6 gene linked to the chloramphenicol acetyltransferase gene as a reporter, we found that the binding of nuclear factor-kappaB to a cis element is essential and sufficient for the induction of the IL-6 gene by HBV-X. We also found that HBV-X enhances the binding of two subunits of nuclear factor-kappaB (p65 and p52) to their target DNA binding sequences. These observations are relevant, in that HBV-X might play an important role in hepatic inflammation and diseases by up-regulating IL-6 production, which can eventually lead to LC and HCC.

Binding Sites↗

PubMed1997

Altered ligand binding properties and enhanced stability of a constitutively active estrogen receptor: evidence that an open pocket conformation is required for ligand interaction.

To elucidate the ligand binding properties of the estrogen receptor (ER) and how ligand access to and release from the ligand binding pocket is affected by the conformational state of the receptor, we have measured the rates of estradiol association and dissociation, the equilibrium binding, and the stability of estradiol binding to denaturants, comparing wild-type human ER and a point mutant (Y537S ER) that shows full constitutive activity, i.e., the same full transcriptional activity in the absence or presence of estrogen. Ligand binding kinetics and affinity were measured with the full-length (1-595) ERs and with truncated forms of both receptors containing domains C through F (including the DNA binding, hinge, and ligand binding domains, amino acids 175-595) or domains E and F (the ligand binding domain; amino acids 304-595). With all ERs, the rates of ligand association and dissociation were considerably slower with the Y537S mutant ER than with wild-type ER (6-fold and 3-4-fold, respectively). These marked differences in ligand on and off rates for the wild-type and Y537S receptors result in a predicted (k-1/k+1) and measured Kd that is 2-fold lower for Y537S ER compared to wild-type ER. The binding of estradiol by wild-type ER was disrupted by high concentrations of urea (above 2 M), whereas the Y537S ER was distinctly more resistant to this disruption. These results are consistent with a model in which wild-type ER in the absence of ligand adopts a transcriptionally inactive collapsed pocket conformation, stabilized by specific interactions of Y537 with nearby regions of ER. When estradiol is bound, the wild-type ER adopts a transcriptionally active, closed pocket (ligand occupied) conformation. By contrast, the Y537S mutant ER favors the transcriptionally active closed pocket conformation, whether occupied by ligand or not, the latter state (closed pocket but unoccupied) accounting for its constitutive activity. Our findings suggest that the entry or exit of ligand from the binding pocket requires that ER adopt an open pocket conformation. The reduced rates of ligand association and dissociation in the constitutively active form of the ER, as well as its greater resistance to disruption of ligand binding by urea, support the supposition that the rate at which this open pocket conformation can be accessed from the unoccupied or ligand-occupied Y537S ER is slower than from the unoccupied or occupied forms of wild-type ER. Thus, the binding and release of ligand by ER require that the receptor access an open pocket state, and the ease with which this state can be accessed is affected by mutations that alter receptor conformation.

Binding Sites↗

PubMed1997

Cloning and expression of a cDNA encoding a novel protein serine/threonine kinase predominantly expressed in hematopoietic cells.

We have isolated a cDNA clone of a new member of protein serine/threonine kinases, MPK38, from a cDNA library constructed from the murine teratocarcinoma PCC4 cell line by the polymerase chain reaction. MPK38 was transcribed as an approx. 2.5 kb mRNA encoding for a protein of 643 amino acids. N-terminus of MPK38 contains the kinase catalytic domain which exhibits approximately 60% protein sequence identity with the SNF1 serine/threonine kinase family. The MPK38 cDNA directs the in vitro translation of two protein species of approx. 70 and approx. 50 kDa, which appear to result from an internal initiation of translation. MPK38 was predominantly expressed in thymus and spleen, but was not detectable in kidney, liver, and muscle in the adult tissues. In addition, MPK38 was apparently expressed in T lineage cells and a macrophage/monocyte cell, but was not detectable in a B cell line and an embryonic cell line. However, a low level of MPK38 transcript was detectable in a mast cell line after a longer exposure. Furthermore, MPK38 gene product showed the kinase activity which was assessed by immune complex kinase assay. Thus, MPK38 gene product seems to play an important role in signal transduction of certain lineages of hematopoietic cells.

Amino Acid Sequence↗

PubMed1997

Abolition of substrate-dependent currents by tyrosine mutation in the transmembrane domain of glutamate transporter.

By site-directed mutagenesis we examined the roles of tyrosine residues (Tyr127) in the putative transmembrane domain of rat glutamate transporter (GLAST). When expressed in Xenopus oocytes, Y127F mutant protein, which was localized in plasma membranes of oocytes, completely abolished glutamate uptake currents but did not affect the intrinsic substrate-independent currents. Coexpression of wild type and mutant transporters supports that the Y127F mutation did not elicit glutamate efflux. The efflux of glutamate by wild type or Y127F mutant transporters was measured under the condition of ion perturbation where transporters run in the reverse direction.

Amino Acid Transport System X-AG↗

PubMed1997

Roles of tyrosine kinases in the regulation of nitric oxide synthesis in murine liver cells: modulation of NF-kappa B activity by tyrosine kinases.

Nitric oxide (NO) synthesis is upregulated during chronic hepatic inflammation. The present study characterized the mechanisms involved in the induction of NO production and inducible NO synthase (iNOS) messenger RNA (mRNA) expression in murine embryonic liver cell line, BNL CL.2 cells. No production by BNL CL.2 cells was induced by interferon-r (IFN-r) plus lipopolysaccharide (LPS). However, other inflammatory cytokines such as interleukin (IL)-beta, tumor necrosis factor alpha (TNF-alpha), and IL-6 had no additional effects on it. The stimulatory effects of IFN-r and LPS were time- and dose-dependent. NO secretion was inhibited by treatment with inducible NOS inhibitors such as NG-monomethyl L-arginine, NG-amino-L-arginine, and diphenylene iodonium. iNOS mRNA was induced 3 hours after IFN-r plus LPS treatment, and iNOS expression was maximal in the presence of IFN-r and LPS. The protein tyrosine kinase inhibitors such as genistein and tyrphostin reduced IFN-r plus LPS-induced iNOS mRNA expression and NO production. In contrast, the inhibitors of protein kinase C, protein kinase A, and protein phosphatases did not affect iNOS expression induced by IFN-r plus LPS. In addition, iNOS mRNA expression was completely blocked by treatment with tyrphostin. However, mRNA expression of an early response gene, JunB, and constitutively expressed genes beta-actin and GAPDH were not inhibited by tyrphostin. Furthermore, tyrphostin inhibited the promoter activation of iNOS gene induced by IFN-gamma plus LPS, and it also suppressed IFN-gamma plus LPS-induced nuclear factor-kappa B-binding activity but not AP-1-binding activity. These results suggest that NO production and iNOS mRNA expression in this cell line is dependent on protein tyrosine kinases but does not require protein kinase C, protein kinase A, or protein phosphatases.

Animals↗

PubMed1997

Culture, category salience, and inductive reasoning.

The role of category salience in category-based induction was demonstrated in two ways: (i) temporarily increasing category salience facilitated category-based induction, and (ii) this effect was moderated by cultural differences that we predicted would be related to chronic category salience. Subjects for whom categories were presumed to be more accessible (Americans) were not as much influenced by manipulations to increase category salience as subjects who were presumed to have lower chronic accessibility of categories (Koreans). However, as anticipated, this pattern was reversed for inferences about behavioral properties of social categories. Due to the 'interdependent' nature of their culture, Koreans presumably have relatively higher chronic accessibility for social categories than do relatively 'independent' Americans, and hence were not influenced as much by increasing category salience.

Cross-Cultural Comparison↗

PubMed1997

Closely related genes encode developmental and tissue isoforms of porcine cytochrome P450 aromatase.

We examined the chromosomal basis for the synthesis of tissue (ovary, endometrium/placenta, and peri-implantation blastocyst) isoforms of cytochrome P450 aromatase in the pig. DNA fragments derived from three distinct porcine aromatase chromosomal genes were cloned and characterized. The porcine type III aromatase gene encoding the blastocyst aromatase isoform was found to consist of nine coding exons and two mutually exclusive, 5' untranslated exons (designated E1A and E1B), collectively spanning 30 kb or more. The porcine type II aromatase gene, encoding the endometrial/placental aromatase isoform, was identified by cloning of a genomic DNA fragment spanning the corresponding exons 7, 8, and 9. The DNA inserts of two other phage clones encompassed exons 2, 3, and 4 of a third chromosomal gene (type I) encoding the ovarian aromatase isoform. All intron-exon junctions in these genomic fragments were found to be identical in relative positions to those of the single-copy human aromatase gene. Comparisons of cDNA and genomic sequences indicated that nucleotide sequence variation was not uniform across the corresponding exons of these genes and that the corresponding intronic sequences were conserved. The type II and type III aromatase genes were localized to the same regional location (q16-17) on swine chromosome 1, which is homologous to the human chromosome 15 region (q21.1) in which the human aromatase gene resides. Results demonstrate that the three aromatase genes characterized in the present study appear to be similar in their overall structural organization and most likely are clustered, which could have resulted from at least two independent gene duplication events. The presence of multiple aromatase genes constitutes a newly described mechanism by which aromatase enzyme biosynthesis and functional activity can be regulated in a tissue and temporal fashion and serves to highlight further the complexity of aromatase gene expression in mammals. Moreover, the presence of a unique aromatase gene that is highly expressed in pig blastocysts may constitute a paradigm for other mammals (e.g., equids, rabbit, hamster) whose peri-implantation blastocysts are estrogenic.

Animals↗

PubMed1997

A developmental switch in expression from blastocyst to endometrial/placental-type cytochrome P450 aromatase genes in the pig and horse.

Pig blastocysts exhibit a transient period of estrogen production at periimplantation, with a second, more sustained period of estrogen synthesis occurring in endometrium and placenta at later pregnancy. Previously we reported the isolation of cDNA clones encoding a novel isoform of cytochrome P450 aromatase (the terminal enzyme in the estrogen biosynthetic pathway) from porcine periimplantation blastocysts. The present study investigated pregnancy-associated expression, in blastocysts and maternal reproductive tract tissues of this and an additional mRNA transcript encoding a distinct P450 aromatase isoform. Restriction endonuclease and nucleotide sequence analyses of 44 cDNA clones demonstrated that the major aromatase mRNA in periimplantation blastocysts and early-pregnancy endometrium and placenta (blastocyst-type) differed in sequence from the major aromatase mRNA expressed in endometrium and placenta at midpregnancy (endometrial-type). The deduced blastocyst and endometrial aromatase isoform protein sequences had 93% similarity. A third type of aromatase mRNA, deleted in exons 4-6 sequences, also was cloned from blastocysts. This cDNA was identical in nucleotide sequence to the blastocyst full-length aromatase cDNA and specified an open reading frame of 354 amino acids for a putative aromatase-related protein containing the heme-binding domain. Expression of this shorter mRNA in blastocysts was confirmed by reverse transcription polymerase chain reaction. The 5'-untranslated exon sequences in the transcripts encoding the blastocyst-type aromatase isoform were distinct from that of the endometrial type, consistent with differential expression of multiple chromosomal genes. In periimplantation equine embryos, however, embryonic and placental 5'-untranslated exon-containing transcripts were coexpressed. Results identify an aromatase isoform expressed in the endometrium and placenta at midpregnancy, demonstrate a transition in synthesis of aromatase isoform-specific mRNAs during placental development, and suggest the preferential involvement of the blastocyst aromatase isoform in synthesis of estrogenic molecules that may function in embryo-maternal signaling at periimplantation.

Amino Acid Sequence↗

PubMed1997

Expression of high-affinity neuronal and glial glutamate transporters in the rat optic nerve.

Recent studies have revealed that a dynamic axon-glial signaling occurs in the rat optic nerve, which is devoid of synapses. This interaction is postulated to be mediated by non-vesicular release of glutamate via a reversal of high-affinity glutamate transporters. Here we examined the expression of glial glutamate transporters (GLAST and GLT-1) and a neuronal transporter (EAAC1) in the rat optic nerve. RT-PCR analysis revealed the presence of mRNAs for GLT-1 and GLAST, but not EAAC1. RNase protection assays showed that of the two glial transporters, mRNA for GLAST was expressed at much higher level than was GLT-1. A similar expression pattern was found in primary astrocyte culture cells. GLAST mRNA level in the optic nerve was comparable to that in the cerebellum. Developmentally, GLAST mRNA level was highest at P2 and dropped slightly by adulthood. Nerve transection resulted in little or no change in mRNA levels for GLAST and GLT-1 assayed at 4 to 14 days post-transection, but GLAST mRNA level was decreased at 64 days. Western blot analysis revealed that the rat optic nerve showed immunoreactivity to antibodies against GLT-1, GLAST, and EAAC1. In conclusion, we suggest that glial and neuronal transporters are present in the rat optic nerve, where dynamic axon-glial interaction has been known to occur. In particular, the unusually high level of expression of GLAST in the optic nerve suggests a possible role for this glial transporter in protecting optic nerves from neurotoxicity during postnatal development.

ATP-Binding Cassette Transporters↗

PubMed1996

Transforming growth factor-beta1 inhibits human keratinocyte proliferation by upregulation of a receptor-type tyrosine phosphatase R-PTP-kappa gene expression.

We studied regulation of a receptor-type tyrosine phosphatase R-PTP-kappa gene expression in a human keratinocyte cell line, HaCaT. Addition of TGF-beta 1 to the HaCaT cells markedly induced the expression of R-PTP-kappa mRNA in a time- and dose-dependent manner. The induction of R-PTP-kappa mRNA expression was observed at a dose as low as 0.02 ng/ml TGF-beta1 and reached a peak at 2 ng/ml TGF-beta 1 after 6 h treatment. The TGF-beta 1-induced R-PTP-kappa mRNA expression was suppressed by sodium orthovanadate, a tyrosine phosphatase inhibitor, and H7, a serine/threonine kinase inhibitor, but not by genistein, a tyrosine kinase inhibitor. In addition, the inducing effect is not dependent on de novo protein synthesis. Taken together, these results suggest that TGF-beta 1 inhibits the human keratinocyte proliferation in vitro, possibly through induction of R-PTP-kappa gene expression.

Cell Division↗

PubMed1996

Suppression of interleukin-1 and tumor necrosis factor-alpha production by acanthoic acid, (-)-pimara-9(11),15-dien-19-oic acid, and it antifibrotic effects in vivo.

Interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha) are major proinflammatory cytokines inducing the synthesis and release of many inflammatory mediators. They are involved in immune regulation, autoimmune diseases, and inflammation. Acanthoic acid, (-)-pimara-9(11),15-dien-19-oic acid, is a pimaradiene diterpene isolated from the Korean medicinal plant, Acanthopanax koreanum. When human monocytes/macrophages stimulated with silica were treated with 0.1-10 microg/ml acanthoic acid, the production of IL-1 and TNF-alpha was inhibited up to 90%, but the production of interleukin-6 (IL-6) was not inhibited at all. At these concentrations, it had no cytotoxic effect on human monocytes/macrophages. It also suppressed the production of TNF-alpha by alveolar macrophages and lymphocytes stimulated with silica. In addition, acanthoic acid inhibited the release of superoxide anion and hydrogen peroxide from human monocytes/macrophages and neutrophils. To know the antifibrotic effects of acanthoic acid, its effects on fibroblast proliferation and collagen synthesis were tested. The proliferation of NIH3T3 cells was inhibited almost completely by the addition of the culture supernatants of human monocytes/macrophages treated with acanthoic acid, but not by the addition of acanthoic acid only. In vitro and in vivo treatment with acanthoic acid reduced collagen production by rat lung fibroblasts and lung tissue. Furthermore, acanthoic acid suppressed granuloma formation and fibrosis in the experimental silicosis. Acanthoic acid reduced serum GOT and GPT in the rats with cirrhosis induced by CCl4, and it was effective in reducing hepatic fibrosis and nodular formation. Taken together, these data indicate that acanthoic acid has a potent anti-inflammatory and antifibrosis effect by reducing IL-1 and TNF-alpha production.

Animals↗

PubMed1996

Roles of protein phosphatase 1 and 2A in an IL-6-mediated autocrine growth loop of human myeloma cells.

Deregulation of IL-6 production is one of the major causes for human multiple myeloma. Exogenous IL-6 stimulated the proliferation of fresh human myeloma cells and the myeloma cell line, U266, which produced IL-6 spontaneously. Anti-IL-6 antibody and IL-6 antisense oligonucleotide suppressed the IL-6 stimulated myeloma cell proliferation, indicating that IL-6 induced the myeloma cell proliferation via an autocrine loop. Okadaic acid, an inhibitor of protein phosphatase 1 and 2A, inhibited the U266 cell proliferation at a concentration of less than 1 ng/ml. At this concentration, okadaic acid suppressed the IL-6-induced IL-6 gene expression of myeloma cells. It seems that the okadaic acid blocked the myeloma cell proliferation by reducing IL-6 synthesis in myeloma cells. In addition, IL-6 itself also regulated IL-6 receptor expression. Analysis by FACScan and RT-PCR showed that anti-IL-6 antibody treatment up-regulated IL-6 receptor expression. Interestingly, the presence of okadaic acid induced the up-regulation of IL-6 receptor expression as well as the down-regulation of IL-6-induced gp130 phosphorylation in the myeloma cells. Taken together, these data suggest that protein phosphatase 1 and 2A are involved in IL-6-mediated autocrine growth of human myeloma cells by modulating IL-6 signaling and IL-6 receptor expression in myeloma cells.

Animals↗

PubMed1996

Molecular cloning and characterization of an estrogen-dependent porcine oviductal secretory glycoprotein.

A family of estrogen-dependent porcine oviductal secretory glycoproteins (POSPs) that exhibit structural similarities are synthesized and secreted into the oviductal lumen at proestrus, estrus, and metestrus. The objectives of this study were to clone the POSP cDNA, obtain the full-length cDNA and protein sequence, examine tissue specificity and species distribution, characterize its regulation, and establish its identity by comparison to other known protein, RNA, or DNA sequences. A full-length cDNA of 2022 base pairs was obtained with an open reading frame of 1581 nucleotides, coding for a deduced protein of 527 amino acids (57 970 M(r)). The deduced protein contained three potential N-glycosylation sites, a consensus heparin-binding site, and potential O-glycosylation sites. Amino acid analysis of POSP-E3 confirmed the presence of a 21-amino acid signal sequence. Northern blot analysis revealed an oviduct-specific mRNA species of 2.25 kb in the infundibulum (INF), ampulla (A), and isthmus (I). An mRNA of similar size was detected in the oviduct of the sheep, cow, and rabbit, and one of slightly greater size (2.8 kb) in the mouse and hamster oviduct but not in the horse or alligator oviduct. Dot blot analysis indicated that steady-state levels of POSP mRNA were significantly greater (p = 0.0001) in the A than in the INF or I regardless of day of the estrous cycle and were greater on Day 0 (estrus; p = 0.0001) regardless of location. Further, steady-state mRNA levels were significantly increased (p = 0.02) on Days 0 and 1, declining rapidly to Day 2 through Day 15 of the estrous cycle. Steady-state POSP mRNA levels were significantly greater (p < 0.003) in ovariectomized gilts treated with estradiol valerate than those treated with other steroid regimens, vehicle, or no treatment (Control), consistent with estrogen control of mRNA expression. The POSP protein exhibited significant identity to oviductal glycoproteins from the baboon, cow, hamster, human, mouse, and sheep, to several mammalian nonoviductal glycoproteins; and to several chitinases. POSP joins a growing subfamily of the chitinase gene family that lacks chitinase enzymatic activity.

Alligators and Crocodiles↗

PubMed1996

Interleukin 4-induced proliferation in normal human keratinocytes is associated with c-myc gene expression and inhibited by genistein.

We studied the effect of IL-4 on the proliferation of cultured normal human keratinocytes. Keratinocyte proliferation was stimulated by IL-4 and inhibited by anti-IL-4 antibody in a concentration-dependent manner. Anti-IL-6 antibody did not inhibit normal human keratinocyte proliferation, suggesting that the IL-4 could directly induce proliferation of these cells. IL-4 significantly induced cell cycle G0/G1 to S phase progression. The keratinocyte proliferation by IL-4 was mediated through one of the growth control genes, c-myc protooncogene. The expression of c-myc mRNA was significantly increased after IL-4 treatment of the keratinocytes, suggesting that c-myc plays a key role in the control of proliferation. The signal transduction pathways induced by IL-4 in the keratinocytes were studied with inhibitors of signal transduction. Genistein, a tyrosine kinase inhibitor, suppressed the level of the induced c-myc mRNA expression, but H7, a serine/threonine kinase inhibitor, and okadaic acid, a protein phosphatase 1 and 2A inhibitor, did not block the induced c-myc gene expression. Taken together, these results suggest that IL-4 stimulates the proliferation of keratinocytes in vitro by promoting a transition from G0/G1 to S phase of the cell cycle. Induction of c-myc after IL-4 treatment could indicate an important role for c-myc in the proliferation of keratinocytes. Our observations also suggest that tyrosine kinases may be involved in IL-4-induced proliferation.

Base Sequence↗

PubMed1996

Molecular cloning of cytochrome P450 aromatase complementary deoxyribonucleic acid from periimplantation porcine and equine blastocysts identifies multiple novel 5'-untranslated exons expressed in embryos, endometrium, and placenta.

To facilitate studies on the molecular mechanisms involved in the unique temporal expression of the P450 aromatase gene in porcine periimplantation embryos, a complementary DNA (cDNA) library was prepared from day 12 porcine embryo messenger RNAs (mRNAs) and screened with a cDNA fragment encoding the amino-terminus of human aromatase. Two size variants of nearly full-length cDNA clones (designated clones 33F and 34B; 2470 and 2588 bp, respectively) were isolated and sequenced. Clones 33F and 34B encoded identical aromatase proteins of 503 amino acids, but differed in size due to alternative polyadenylation signal usage for the corresponding mRNAs. Using the 5'-rapid amplification of cDNA ends procedure, two classes of cDNA clones that contained distinct putative exon 1 sequences (E1A and E1B, respectively), but were otherwise identical in exon 2 and 3 sequences, were isolated. The E1A DNA sequence was identical to the 5'-end specified by clones 33F and 34B, except for an additional 23 nucleotides. The E1A and E1B exons showed no significant homology with each other or with the aromatase cDNA sequences from other species, including human. Expression of E1A-containing aromatase mRNA was higher than that of E1B in day 12 blastocysts, although both mRNAs were expressed at low levels in porcine endometrium and placenta of early pregnancy and in ovary at periestrus. To determine whether the E1A or E1B sequence was common to aromatase mRNAs in other steroidogenic preimplantation embryos, aromatase cDNA clones spanning exons 1-3 were isolated from equine embryos by a combination of RT-PCR and 5'-rapid amplification of cDNA ends. Equine embryonic exon 1 cDNA sequence had no homology with corresponding regions in aromatase transcripts of pigs or other species; in contrast, exon 2 and 3 cDNA sequences predicted an amino acid sequence with significant homology to aromatase. Results from the present study demonstrate alternative splicing of a novel first exon(s) in P450 aromatase gene transcripts in porcine preimplantation embryos and in endometrium and placenta of pregnancy. The lack of homology among equine, porcine, human, and bovine aromatase 5'-untranslated exons may indicate divergence in the corresponding regulatory motifs of these regions among mammalian species. These observations suggest the potential significance of activation of the E1A promoter and/or splicing of novel aromatase 5'-exons in the transient production of estrogens by periimplantation blastocysts.

Alternative Splicing↗

PubMed1996

An allograft kidney showing both features of IgA nephropathy and membranous glomerulonephritis--a case report.

We report a case of glomerular disease with both mesangial IgA and subepithelial IgG deposits in the allograft kidney. The patient was a 36 year-old man who had received a renal allograft 1 year previously. Fifteen days before admission, he discovered a microscopic hematuria without clinical evidences of allograft rejection. Light microscopy showed diffuse increase of mesangial matrix without mesangial cell proliferation. Capillary walls were diffusely and mildly thickened. Immunofluorescence microscopy demonstrated both granular deposits of IgA in the mesangium and IgG along the capillary walls. On electron microscopy, electron-dense deposits were identified not only in the mesangium but also on the epithelial side of the glomerular basement membrane.

Adult↗