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S Sassa

Publications and source records attributed to S Sassa.

At least 19 recordsLinked to original sources
PubMed1995

The role of the erythroid-specific delta-aminolevulinate synthase gene expression in erythroid heme synthesis.

Using antisense technology, the effects of suppressed gene expression of the erythroid-specific delta-aminolevulinate (ALA) synthase (ALAS-E) on heme synthesis, expression of mRNAs encoding an erythroid-specific transcription factor NF-E2, other heme pathway enzymes, and beta-globin were examined in murine erythroleukemia (MEL) cells. In MEL cells in which an antisense ALAS-E RNA was expressed (AS clone), sense ALAS-E mRNA levels in both untreated and dimethylsulfoxide (DMSO)-treated cells were decreased compared with their respective controls. Heme synthesis in AS clones was decreased in proportion to the suppressed levels of ALAS-E mRNA. In addition, mRNAs for ALA dehydratase, porphobilinogen deaminase, ferrochelatase (FeC), and beta-globin were also decreased in AS clones. There was a strong correlation between the level of ALAS-E mRNA and most of the mRNAs of the heme pathway enzymes and beta-globin. There was a decrease in the mRNA level of p45, but not of mafK, which are the large and the small subunits of NF-E2, respectively, in AS clones. Treatment of AS cells with hemin and ALA in the presence of DMSO partially restored the suppressed mRNA levels for beta-globin and FeC and heme content, respectively. These findings thus indicate that heme formation, which is determined by the level of ALAS-E, plays an essential role on gene expression of many proteins necessary for erythroid development.

5-Aminolevulinate Synthetase↗
PubMed1995

Expression of cytochrome P450 mRNAs in the colon and the rectum in normal human subjects.

The expression of mRNA for three isozymes of cytochrome P450, CYP1A1, 1A2 and 3A3, was examined in biopsied samples of colon and rectum from five healthy subjects. Using RT-PCR, it was possible to detect the three CYP mRNA species at both sites in all samples examined. The expression of each CYP mRNA was site-specific in that rectum had higher levels of CYP1A1 mRNA than colon, while colon had higher levels of CYP3A3 mRNA than in rectum. In contrast, there was no consistent trend in CYP1A2 mRNA expression between the two sites. Variability of alimentary CYP expression may contribute to individual differences in susceptibility to carcinogens and/or drugs.

Aged↗
PubMed1995

The effect of porphyrin and radiation on ferrochelatase and 5-aminolevulinic acid synthase in epidermal cells.

The effects of ultraviolet A (UVA) and blue light on ferrochelatase protein, and its mRNA level, in 5-aminolevulinic acid (ALA)-loaded A431 cells was evaluated. Western blot analysis of ferrochelatase protein showed a protein band of 43 kDa. There was a decrease in the protein concentration 24 h and 48 h after irradiation of these cells. In contrast, as judged by Northern blot analysis, there was no change in ferrochelatase mRNA level. Measurement of ALA synthase activity showed an ALA dose-dependent but radiation-independent decrease of enzyme activity, suggesting an end-product feedback inhibition. Since reactive oxygen species generated by porphyrin-induced photochemical reaction may be involved in the decrease in ferrochelatase protein, the effect of scavengers of reactive oxygen species was evaluated by measuring porphyrin accumulation in irradiated, ALA-loaded A431 cells. Porphyrin accumulation was significantly decreased in the presence of singlet oxygen scavenger sodium azide (0.05 mM, 40.6% suppression) or hydroxyl radical scavenger mannitol (5.0 mM, 45.0% suppression). These data suggest that the photochemical reaction induced by porphyrin and irradiation resulted in a decrease in ferrochelatase protein content, but had no effect on ferrochelatase mRNA level nor on ALA synthase activity. The decrease in protein was partly mediated by the reactive oxygen species.

5-Aminolevulinate Synthetase↗
PubMed1995

The effect of ALA and radiation on porphyrin/heme biosynthesis in endothelial cells.

To study porphyrin biosynthesis in human microvascular endothelial cells, HMEC-1 cells, a transformed human microvascular endothelial cell line, were incubated with 5-aminolevulinic acid (ALA), the precursor of endogenous porphyrins, and porphyrin accumulation was measured spectro-fluorometrically. The HMEC-1 cells accumulated porphyrin in a concentration-related and a time-dependent fashion. Protoporphyrin was the predominant porphyrin accumulated in the cells. The effect of light on protoporphyrin accumulation was evaluated by exposing the ALA-loaded HMEC-1 cells to ultraviolet-A (UVA) and blue light, followed by another incubation with ALA for 2-24 h. Enhancement of protoporphyrin accumulation in irradiated HMEC-1 cells was observed 2-24 h after irradiation, which was associated with a decrease in ferrochelatase protein and activity. Porphyrin accumulation from ALA after irradiation was significantly decreased when catalase (750-3000 U/mL, 29.3-44.3% suppression) or superoxide dismutase (270 U/mL, 36.4% suppression) was present during irradiation. These data demonstrate that HMEC-1 cells were capable of porphyrin biosynthesis, and that exposure of protoporphyrin-containing HMEC-1 cells to UVA and blue light, which includes the Soret band spectrum, decreased the ferrochelatase activity and its protein. These changes were mediated, at least in part, by reactive oxygen species.

Aminolevulinic Acid↗
PubMed1994

Suppression of cytochrome P450IA1 by interleukin-6 in human HepG2 hepatoma cells.

The effects of interleukin-6 (IL-6), the major inducer of the acute-phase reaction, on the expression of cytochrome P450IA1 (CYPIA1) were examined using human HepG2 hepatoma cells. Treatment of cells with IL-6 decreased the level of 3-methylcholanthrene-induced CYPIA1 protein and its mRNA. Nuclear runoff analysis revealed that the effect of IL-6 was largely transcriptional. IL-6 treatment of HepG2 cells increased mRNA for microsomal heme oxygenase, the rate-limiting enzyme in heme catabolism, suggesting that the suppressive effect of IL-6 on CYPIA1 mRNA may be due to a loss of heme. Consistent with this hypothesis, simultaneous treatment of cells with Sn-mesoporphyrin, an inhibitor of heme oxygenase, prevented the IL-6-mediated suppression of CYPIA1. These findings suggest that the suppression of P450IA1 mRNA by IL-6 appears to occur, at least in part, from the decline in free heme content as a result of the induction of heme oxygenase. Our results raise the possibility that other physiological as well as environmental stimuli which affect cellular heme concentrations may also modulate the expression of P450s.

Carcinoma, Hepatocellular↗
PubMed1994

[Long-term observation of serum hormone fluctuations in aging model of gonadectomized rat].

Serum LH increased rapidly in both sexes after gonadectomy, showing a peak at the 8th week. Afterwards, LH decreased gradually until the 75th week. In general, LH values in males were lower than those in females. FSH, as well as LH, increased rapidly after gonadectomy. In male serum FSH reached a peak at the 8th week and then decreased showing two peaks at the 34th and 92nd weeks. Serum FSH level decreased remarkably after the 96th week in males. In females FSH increased gradually with a subsequent rapid increase from the 28th week and reached a peak at the 42nd week. Afterwards, this FSH level was maintained to the 128th week. Serum progesterone (P4) decreased after gonadectomy in females, whereas it increased to 4 fold in males. In males this high level was maintained to the 66th week. Serum estradiol (E2) decreased in both sexes until the 3rd week after gonadectomy. However, in males the serum E2 level increased at the 4th and 5th weeks and reached the level of the 1st week after gonadectomy. Afterwards, the E2 level decreased to 10pg/ml at the 40th week which was maintained to the 128th week. In females, E2 increased from the 5th to 22nd week and decreased again from the 33rd week, and then the E2 level decreased rapidly to about 10pg/ml which was maintained to the 128th week. The present experiment indicated two results in male rats. 1. Serum FSH decreased remarkably after the 96th week of gonadectomy. 2. Serum P4 increased in spite of gonadectomy.

Aging↗
PubMed1994

Regulation of beta-globin mRNA accumulation by heme in dimethyl sulfoxide (DMSO)-sensitive and DMSO-resistant murine erythroleukemia cells.

The level of mRNA encoding beta-globin was examined in dimethyl sulfoxide (DMSO)-sensitive (DS), and DMSO-resistant (DR) murine erythroleukemia (MEL) cells. DR cells lack erythroid-specific delta-aminolevulinate (ALA) synthase (AL-AS-E), and fail to undergo erythroid differentiation following treatment with DMSO. Treatment of cells with DMSO markedly increased ALAS-E mRNA in DS cells, while the same treatment downregulated the nonspecific ALA synthase (ALAS-N) mRNA levels in both DS and DR cells. The levels of beta-globin mRNA, heme content, and hemoglobin in DS cells increased, while those in DR cells decreased following treatment with DMSO. Treatment of DR cells with hemin caused an increase in beta-globin mRNA and hemoglobin, and partially restored the DMSO-mediated suppression of beta-globin mRNA and hemoglobin synthesis. DMSO treatment decreased heme oxygenase (HO) mRNA in hemin-treated DS cells, but not in hemin-treated DR cells. These findings indicate that heme is necessary for accumulation of the beta-globin transcript during erythroid differentiation, and that hemin-mediated HO induction becomes markedly downregulated in differentiated erythroid cells, presumably because less free heme is available for HO induction by a greater demand for the synthesis of hemoglobin.

5-Aminolevulinate Synthetase↗
PubMed1994

Characterization and expression of cDNA encoding coproporphyrinogen oxidase from a patient with hereditary coproporphyria.

Hereditary coproporphyria (HCP) is an acute hepatic porphyria with autosomal dominant inheritance, but with a variable degree of clinical expression. Molecular cloning, sequencing and expression of the defective gene for coproporphyrinogen oxidase (CPO) in a patient with HCP were carried out. Enzyme assays revealed that CPO activity in EBV-transformed lymphoblastoid cells from the proband and one of her sisters was approximately 50% of normal. Nucleotide sequence analysis of CPO cDNAs isolated from the proband's cells demonstrated three base substitutions, and three accompanying amino acid substitutions. An A514-->C transition causing a Asn172-->His substitution occurred in one allele, while two other transitions, G265-->A and G580-->A, caused Gly89-->Ser and Val194-->Ile substitutions, respectively, in the other allele. The A514-->C and the G580-->A transitions are known genetic polymorphisms. Transfection of CPO cDNA into Escherichia coli demonstrated that cDNA with the G265-->A transition produced a protein with less than 5% of normal enzyme activity. These findings indicate that the G265-->A transition, involving the highly conserved glycine residue at the 89th position, is responsible for the CPO defect in the patient and accounts for the partial deficiency of CPO activity in this pedigree.

Amino Acid Sequence↗
PubMed1994

Changes with age of mammary glands in male and female soft-furred rat, Millardia meltada, in relation to prolactin and testosterone.

The soft-furred rat, millardia (Millardia meltada), is characterized by the development of androgen-dependent mammary tumours only in males. The age-related changes of the activities of thymidylate synthetase (TS) and thymidine kinase (TK), which contribute to DNA synthesis through de novo and salvage pathways, respectively, and structure in the mammary glands were studied in both males and females of this species between 5-28 months of age. While TK activity had no relation to age, TS activity decreased with age in males. In the females, TK activity increased with age, but not TS activity. These enzyme activities were generally higher in females than in males. The mammary glands of both sexes consisted of fine ducts with small end-buds and the glands of males contained mostly black pigments at any age examined. In either males or females, serum levels of prolactin and testosterone related little with age, DNA synthesizing enzyme activities or structure of the mammary glands. Furthermore, elevation by pituitary grafting of circulating prolactin affected neither DNA synthesizing enzyme activities nor structure of mammary glands in both sexes. The histological structures of adrenal, testis, ovary, ventral prostate and uterus of millardia were essentially similar to those of mice or rats.

Adrenal Glands↗
PubMed1993

Haem is necessary for a continued increase in ferrochelatase mRNA in murine erythroleukaemia cells during erythroid differentiation.

The level of mRNA encoding ferrochelatase (FeC) was examined in two murine erythroleukaemia (MEL) clones, DS and DR, a DMSO-sensitive, and a DMSO-resistant clone, respectively. DS cells undergo erythroid differentiation by DMSO treatment with a marked increase in haem synthesis, while DR cells fail to do so due to the lack of the erythroid-specific delta-aminolaevulinate synthase (ALAS-E). Both DS and DR cells showed an increase in the level of FeC mRNA within 18 h of DMSO treatment. The level of FeC mRNA in DR cells was then decreased, while that in DS cells continued to increase for 72 h. Treatment with haemin significantly increased FeC mRNA in DR cells. When cells were treated with both DMSO and haemin, the level of FeC mRNA in DR cells increased to a level comparable to that in DS cells. These findings suggest that the failure to maintain increased FeC mRNA DR cells after DMSO treatment may be due to a deficiency of haem in these cells.

Animals↗
PubMed1992

Interleukin-6 down regulates the expression of transcripts encoding cytochrome P450 IA1, IA2 and IIIA3 in human hepatoma cells.

Effects of human interleukin-6 (hIL-6), the major acute phase inducer, on the expression of transcripts encoding cytochrome P450s were examined in human hepatoma-derived cells. Using reverse-transcription polymerase chain reaction, it was demonstrated that three hepatoma cell lines, HepG2, HepG2f and Hep3B, express P450 mRNAs encoding IA1, IA2 and IIIA3, the major P450 isozymes involved in carcinogen metabolism, and that they also show induction responses to treatment with their specific inducers. When hepatoma cells were treated with hIL-6, the levels of IA1, IA2 and IIIA3 mRNAs were markedly suppressed. These findings suggest that significant down regulation of cytochrome P450s may occur during the acute phase reaction, which may result in alterations in drug biotransformation.

Base Sequence↗
PubMed1992

Heme oxygenase is a positive acute-phase reactant in human Hep3B hepatoma cells.

The effects of human interleukin-6 (hIL-6), the major acute-phase inducer, on the level of the transcript of microsomal heme oxygenase (HO) were examined in a human hepatoma cell line, Hep3B. Messenger RNAs (mRNAs) encoding HO and haptoglobin (Hpt) increased after hIL-6 treatment in a time- and dose-dependent manner. hIL-6 had no effect on the induction of heat-shock protein 70 (hsp70) mRNA, suggesting that the induction of HO by hIL-6 is regulated by a different mechanism from that which mediates the heat-shock induction of this enzyme. The hIL-6-mediated induction of HO mRNA was completely abrogated by simultaneous treatment of cells with actinomycin D, but not with cycloheximide, suggesting that the induction occurs at the level of transcription. A nuclear factor was shown both in untreated, and in the hIL-6-treated Hep3B cells that binds specifically to the IL-6-responsive element (IL6-RE) of the human HO gene. These findings suggest that HO is a positive acute-phase reactant in this human liver-derived cell line, and that the nuclear factor specific to the IL6-RE may be involved in the activation of the HO gene after hIL-6 treatment.

Acute-Phase Proteins↗
PubMed1992

Acylpeptide hydrolase: inhibitors and some active site residues of the human enzyme.

Acylpeptide hydrolase may be involved in N-terminal deacetylation of nascent polypeptide chains and of bioactive peptides. The activity of this enzyme from human erythrocytes is sensitive to anions such as chloride, nitrate, and fluoride. Furthermore, blocked amino acids act as competitive inhibitors of the enzyme. Acetyl leucine chloromethyl ketone has been employed to identify one active site residue as His-707. Diisopropylfluorophosphate has been used to identify a second active site residue as Ser-587. Chemical modification studies with a water-soluble carbodiimide implicate a carboxyl group in catalytic activity. These results and the sequence around these active site residues, especially near Ser-587, suggest that acylpeptide hydrolase contains a catalytic triad. The presence of a cysteine residue in the vicinity of the active site is suggested by the inactivation of the enzyme by sulfhydryl-modifying agents and also by a low amount of modification by the peptide chloromethyl ketone inhibitor. Ebelactone A, an inhibitor of the formyl aminopeptidase, the bacterial counterpart of eukaryotic acylpeptide hydrolase, was found to be an effective inhibitor of this enzyme. These findings suggest that acylpeptidase hydrolase is a member of a family of enzymes with extremely diverse functions.

Acetylation↗
PubMed1992

The molecular defect of ferrochelatase in a patient with erythropoietic protoporphyria.

The molecular basis of an inherited defect of ferrochelatase in a patient with erythropoietic protoporphyria (EPP) was investigated. Ferrochelatase is the terminal enzyme in the heme biosynthetic pathway and catalyzes the insertion of ferrous iron into protoporphyrin IX to form heme. In Epstein-Barr virus-transformed lymphoblastoid cells from a proband with EPP, enzyme activity, an immunochemically quantifiable protein, and mRNA content of ferrochelatase were about one-half the normal level. In contrast, the rate of transcription of ferrochelatase mRNA in the proband's cells was normal, suggesting that decreased ferrochelatase mRNA is due to an unstable transcript. cDNA clones encoding ferrochelatase in the proband, isolated by amplification using the polymerase chain reaction, were found to be classified either into those encoding the normal protein or into those encoding an abnormal protein that lacked exon 2 of the ferrochelatase gene, indicating that the proband is heterozygous for the ferrochelatase defect. Genomic DNA analysis revealed that the abnormal allele had a point mutation, C----T, near the acceptor site of intron 1. This point mutation appears to be responsible for the post-transcriptional splicing abnormality resulting in an aberrant transcript of ferrochelatase in this patient.

Amino Acid Sequence↗
PubMed1992

Differential induction responses of delta-aminolevulinate synthase mRNAs during erythroid differentiation: use of nonradioactive in situ hybridization.

Expression of mRNAs encoding the erythroid-specific delta-aminolevulinate synthase (ALAS-E) and the nonspecific delta-aminolevulinate synthase (ALAS-N) were examined in murine Friend virus-transformed erythroleukemia (MEL) cells using nonradioactive in situ hybridization. Following dimethyl sulfoxide (DMSO) treatment, ALAS-E mRNA increased markedly, while ALAS-N mRNA did not increase in wild-type MEL cells. In contrast, in a DMSO-resistant clone of MEL cells, ALAS-E was not detectable before and after DMSO treatment. These findings suggest that ALAS-E and ALAS-N mRNAs are under separate controls and that the expression of ALAS-E mRNA is a critical event in erythroid differentiation.

5-Aminolevulinate Synthetase↗
PubMed1992

Suppression of the development of uterine adenomyosis by danazol treatment in mice.

Inhibitory effects of danazol, an isoxazol derivative of synthetic steroid 17 alpha-ethinyl-testosterone, on the development of uterine adenomyosis, a pathological disorder of endometrial tissue defined as the presence of endometrial glands and stroma in the myometrium, were investigated in mice of SHN strain. Mice treated with 0.5 microgram danazol for 5 weeks during 4-9 weeks of age and killed at 21 weeks of age showed significantly lower incidence of the spontaneous development of adenomyosis than the age-matched intact control mice. The inhibitory effects of danazol were also evident in mice bearing pituitary isografts which were effective in inducing an early and a high incidence of adenomyosis. Furthermore, the treatment with danazol resulted in the decrease of serum levels of luteinizing hormone (LH) and prolactin (PRL) associated with hypofunction of ovaries and persistent diestrus. These results support the usefulness of danazol for the clinical treatment of gynecological disorders except for hypofunction of ovaries.

Animals↗
PubMed1992

Photosensitivity, abnormal porphyrin profile, and sideroblastic anemia.

Cutaneous photosensitivity in a 43-year-old man with idiopathic sideroblastic anemia associated with an abnormal porphyrin profile is reported. This condition was associated with elevated free erythrocyte porphyrin, plasma protoporphyrin, urine porphyrins (predominantly coproporphyrin), stool porphyrins (predominantly protoporphyrin), decreased ferrochelatase activity, and deletion of portions of the long arms of chromosomes 18 and 20. Five other patients with sideroblastic anemia and abnormal porphyrin profiles have been described; all but one of these patients had photosensitivity. The porphyrin profile of this patient is similar to that of three other previously described patients.

Adult↗