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C Chakraborty

Publications and source records attributed to C Chakraborty.

32 records · Page 2Linked to original sources
PubMed1990

Catecholoestrogen synthesis and metabolism in the rabbit uterus during the periimplantation period.

Microsomal oestradiol-2/4-hydroxylase (OE-2/4-H) and cytosolic catechol-O-methyltransferase (COMT) (EC 2.1.1.6) activity in the uteri of pregnant and pseudopregnant rabbits during the periimplantation period were studied. The apparent Km for the 4-hydroxylation of oestradiol (3.18 microM) was considerably less than for the 2-hydroxylation reaction (13.36 microM), whereas the Vmax were almost equal. This suggests that 4-hydroxyoestradiol (4-OH-OE2) is the predominant product of OE-2/4-H in the rabbit uterus. These reactions were inhibited by SKF-525A, indicating the involvement of cytochrome P450 dependent monooxygenases. Uterine cytosolic COMT utilized 2-hydroxyestradiol (2-OH-OE2) as the preferred substrate as compared to 4-hydroxyoestradiol (4-OH-OE2). Since the rabbit uterus has a considerable capacity to synthesize 4-OH-OE2 and a lower capacity to metabolize it, it could be suggested that more 4-OH-OE2 than 2-OH-OE2 could be available to the uterus for its physiological activities. Furthermore, an increase in OE-2/4-H in Day 6 pseudopregnant and pregnant uteri with a concomitant decrease in COMT suggests the involvement of catecholoestrogens in the implantation process in the rabbit.

Animals↗
PubMed1990

Estradiol-15 alpha-hydroxylation: a new avenue of estrogen metabolism in peri-implantation pig blastocysts.

Pig blastocysts have the capacity to convert estradiol into catechol estrogens. Our present study shows that they also have the capacity to hydroxylate estradiol in cycloaliphatic C-atom 15, and this aliphatic hydroxylation reaction is more predominate than the aromatic hydroxylations. The conversion of [4-14C]estradiol to [4-14C]15 alpha-hydroxyestradiol by mitochondrial-rich/microsomal fractions was examined by isolation of this product using reversed phase high-performance liquid chromatography (HPLC) attached to a radiometric flow detector, and its identification by gas chromatography-mass spectrometry. The enzyme kinetics for estrogen 15 alpha-hydroxylase were performed in the pig blastocyst obtained on Day 13 of pregnancy (Day 0 = first acceptance of the male). The enzyme follows classical Michaelis-Menten kinetics. The apparent Kms for estradiol were 2.47 and 1.85 microM, and the apparent Vmaxs were 0.25 and 0.197 nmol/mg/min in the mitochondrial-rich and microsomal fractions, respectively. The enzyme activity was inhibited by different steroidal compounds and non-steroidal estrogens, as well as by CO, SKF-525A, piperonyl butoxide and antibody to cytochrome P450 reductase. Ontogenesis of the blastocyst's estrogen 15 alpha-hydroxylase follows a similar pattern to that of estrogen-2/4-hydroxylase. Thus, highest activity was observed on Days 12 and 13 and lowest was on Day 15 of pregnancy. Furthermore, the enzyme is abundant primarily in the extraembryonic tissues rather than in the embryo proper. The abundance of the enzyme in the extraembryonic tissues, and its surge at a critical time of pregnancy recognition and just prior to implantation suggest that 15 alpha-hydroxylated estradiol could be involved in these processes.

Animals↗
PubMed1990

Estrogen regulates the synthesis of epidermal growth factor in mouse uterine epithelial cells.

Immunocytochemical analyses, using several mouse epidermal growth factor (EGF) polyclonal antibodies, detected immunoreactivity only in uterine luminal and glandular epithelia on late proestrus, estrus, and early on day 1 of pregnancy, but not late on day 1. This immunoreactivity was not detected in the ovariectomized uterus, but after estrogen stimulation it was detected first in the luminal epithelium between 12-24 h and then also in the glandular epithelium by 48 h. After 72 h of estrogen withdrawal, EGF immunoreactivity was no longer detected. This response was specific for estrogen and did not occur after progesterone injection (2 mg/day for 4 days). Using antipeptide antibodies specific for prepro-EGF, no immunoreactivity was detected in the ovariectomized uterus, weak reactivity was detected in the estrogenized uterus and submandibular gland, and strong reactivity was detected in the kidney. Northern blot analysis of uterine RNA failed to detect the expected 4.8-kilobase prepro-EGF mRNA, but, instead, a rare transcript of 2.4 kilobases was detected, which suggests that EGF mRNA is alternately processed in the uterus. The presence of an EGF-coding uterine transcript was further documented by hybridization of an EGF-coding region-specific oligodeoxyribonucleotide (oligo) to polymerase chain reaction-amplified uterine cDNA. In situ hybridization, using a prepro-EGF cRNA probe as well as an EGF-coding region-specific oligo, showed hybridization that colocalized with the EGF immunostaining (epithelia) and was absent from non-EGF-immunoreactive cells. Pulse labeling experiments coupled with immunoaffinity chromatography showed that estrogen induced an increase in the relative rate of synthesis of an acid-soluble immunoreactive protein which was the same size as authentic EGF. Furthermore, analysis of acid-soluble uterine proteins fractionated by DEAE-cellulose chromatography demonstrated a single coincident peak of antigenic activity and receptor-binding activity which coeluted from the column with authentic EGF. Electron microscopy localized EGF immunoreactivity to the Golgi of luminal epithelial cells. Taken together these results suggest that estrogen regulates expression of the EGF gene specifically in uterine epithelial cells. Increased expression of this gene results in an increase in the relative rate of synthesis of this protein and the accumulation of mature EGF.

Animals↗
PubMed1990

The O-methylation of catechol oestrogens by pig conceptuses and endometrium during the peri-implantation period.

Pig conceptuses display a surge in oestrogen and catecholoestrogen synthetic activity during the peri-implantation period. However, the pathways of catecholoestrogen metabolism in pig conceptuses and endometrium are unknown. O-Methylation is an important route of catecholoestrogen metabolism. Therefore, the O-methylations of 2- and 4-hydroxy-oestradiols (2- and 4-OH-oestradiol) by cytosol of pig conceptuses and endometrium during the peri-implantation period were studied. Kinetic studies performed in tissues obtained on day 13 of pregnancy (day 0 = first acceptance of the male) indicated that the O-methylation of 2-OH-oestradiol displayed simple Michaelis-Menten kinetics in both tissues. In blastocysts, the apparent Michaelis constant (Km) and maximum velocity (Vmax) for the O-methylation of 2-OH-estradiol were 1.4 mumol/l and 11.27 pmol/mg protein per min respectively, and when 4-OH-oestradiol was used as substrate, the values were 2.53 mumol/l and 9.86 pmol/mg protein per min respectively. The apparent Km and Vmax values for the O-methylation of 2-OH-oestradiol in endometrium were 0.77 mumol/l and 19.6 pmol/mg protein per min respectively, and for the O-methylation of 4-OH-oestradiol were 2.44 mumol/l and 10.38 pmol/mg protein per min respectively. Ontogenesis of catechol-O-methyltransferase (COMT) in conceptuses and endometrium was studied from day 10 to day 19 of pregnancy. Conceptus COMT activity was lowest on day 10 and increased gradually to day 19 of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Pattern and tissue distribution of catechol estrogen forming activity by pig conceptuses during the peri-implantation period.

The profile of catechol estrogen formation by pig conceptuses from d 14 to 20 of pregnancy (d 0 = 1st d of estrus) was studied using a direct product isolation assay for estrogen-2/4-hydroxylase (E-2/4-H). High performance liquid chromatography was coupled to detection of radioactive products in a flowing system to separate the 2- and 4-hydroxyestradiol (2- and 4-OH-E2) formed from estradiol. The major product was 2-OH-E2; its formation increased with increasing amounts of nicotinamide cofactors and exhibited a preference for NADPH. Maximum velocities were 909 and 154 pmol.mg protein-1.30 min-1, and apparent Michaelis constants were 7.14 and 5.12 microns, for 2- and 4-OH-E2 formation, respectively. Homogenates of d-13 conceptuses produced 1,151 +/- 142 pmol.mg protein-1.30 min-1 of 2-OH-E2. Estradiol-2-hydroxylase was affected (P less than .01) by day of pregnancy, and day effects from d 14 to 20 were described by a fourth-order equation (P less than .001). Conceptuses collected on d 14 produced 119 +/- 15.4 pmol of 2-OH-E2.mg protein-1.30 min-1. By d 15, E-2-activity had declined to 15.7 +/- 1.8 pmol.mg protein-1.30 min-1. Formation of 2-OH-E2 increased to 64.2 +/- 15.4 pmol.mg protein-1.30 min-1 on d 17, remained stable through d 19, and was 39.4 +/- 13.8 pmol.mg protein-1. 30 min-1 on d 20. Therefore, E-2/4-H activity appears to be regulated from d 14 to 20 of pregnancy in pigs; it may participate in the establishment of pregnancy.

Animals↗
PubMed1988

Epidermal growth factor binding in rat uterus during the peri-implantation period.

The profile of epidermal growth factor (EGF) binding to uterine membranes of rats on Day 1 through Day 7 of pregnancy was studied. The binding was lowest on Day 1 and increased gradually through the pre- and postimplantation periods. Binding affinity of the Day 7 uterine membranes was considerably higher than that of the Day 1. Apparent affinity constants (Ka) of Day 1 and Day 7 membranes were 0.29 X 10(-8) M and 1.03 X 10(-8) M respectively. To our knowledge, this is the first report of the modulation of EGF binding to uterine membranes by progesterone-estrogen interaction during early pregnancy.

Animals↗
PubMed1988

Characteristics of estrogen-2/4-hydroxylase in pig blastocysts: inhibition by steroidal and nonsteroidal agents.

The inhibitory potencies of steroidal and non-steroidal estrogens, catechol-estrogens, methoxyestrogen, haloestrogens, cholesterol and its side-chain-cleaved products, and inhibitors of steroid aromatase against the activity of estradiol-2/4-hydroxylase (E-2/4-H) in pig blastocysts were studied. All tested compounds, except cholesterol and 4-hydroxyandrostenedione, inhibited E-2/4-H in vitro. The fluctuation of E-2/4-H activity in pig blastocysts on different days of pregnancy may be due to the modulation of enzyme activity by steroids in the uterine lumen. Although alpha-naphthoflavone and aminoglutethimide did not affect E-2/4-H activity in vitro, inhibition by CO (95% CO + 5% O2), SKF-525A, piperonyl butoxide, and antibody to cytochrome P-450 reductase provides evidence for the involvement of cytochrome P-450 in E-2/4-H activity in pig blastocysts.

Animals↗
PubMed1988

The estrogen 2-hydroxylase activity of the gonadotropin-stimulated hypophysectomized immature rat ovary.

Using high-performance liquid chromatography and a combination of electrochemical and radiometric flow detection for 2-[14C]hydroxyestradiol, changes in estrogen 2-hydroxylase activity in the microsomal fraction of rat ovarian homogenates were followed. Injection of human chorionic gonadotropin (hCG) at 12-hr intervals to hypophysectomized immature rats stimulated hypertrophy of the theca-interstitial tissue and produced a profound increase in enzyme activity. With the last injection of hCG at 96 hr the peak serum concentration of hCG was reached 12 hr later and then decreased exponentially with a half-time of 13 hr. However, enzyme activity remained elevated for at least 60 hr before beginning to fall. Pregnant mare's serum gonadotropin (PMSG) also produced an increase in activity, which was apparently limited to the thecal-interstitial tissue because freshly removed granulosa cells from the mature follicles had undetectable activity levels. Administration of anti-PMSG antiserum after enzyme activity had been increased resulted in a prompt fall in activity, as did injection of hCG to mimic an ovulatory surge of LH. The results indicate that the thecal-interstitial tissue of the rat ovary has estrogen 2-hydroxylase activity that is dependent upon gonadotropic stimulation for expression.

Animals↗
PubMed1984

Inverse relationship between melanogenesis and endogenous hydroquinone.

Inhibition or stimulation of melanogenesis have been found to occur as a result of the alteration of hydroquinone levels in the body. Substances which stimulate melanogenesis are found to lower the level of hydroquinone in amphibia, and evidence for the relationship is also given by mammalian experiments.

Animals↗
PubMed1983

Abnormal tryptophan pyrrolase and amino acids related to melanogenesis in vitiligo.

A comparison of the serum tyrosine, DOPA, tryptophan, tyrosinase and tryptophan pyrrolase levels of vitiliginous patients with those of normal subjects show abnormalities in all these parameters. As the clinical diagnosis of vitiligo may be made without difficulty, these parameters appear to be of little diagnostic value in vitiligo. But they may be considered as additional biochemical parameters in vitiligo.

Adolescent↗