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R F Casper

Publications and source records attributed to R F Casper.

At least 19 recordsLinked to original sources
PubMed2006

Genetic variance modifies apoptosis susceptibility in mature oocytes via alterations in DNA repair capacity and mitochondrial ultrastructure.

Although the identification of specific genes that regulate apoptosis has been a topic of intense study, little is known of the role that background genetic variance plays in modulating cell death. Using germ cells from inbred mouse strains, we found that apoptosis in mature (metaphase II) oocytes is affected by genetic background through at least two different mechanisms. The first, manifested in AKR/J mice, results in genomic instability. This is reflected by numerous DNA double-strand breaks in freshly isolated oocytes, causing a high apoptosis susceptibility and impaired embryonic development following fertilization. Microinjection of Rad51 reduces DNA damage, suppresses apoptosis and improves embryonic development. The second, manifested in FVB mice, results in dramatic dimorphisms in mitochondrial ultrastructure. This is correlated with cytochrome c release and a high apoptosis susceptibility, the latter of which is suppressed by pyruvate treatment, Smac/DIABLO deficiency, or microinjection of 'normal' mitochondria. Therefore, background genetic variance can profoundly affect apoptosis in female germ cells by disrupting both genomic DNA and mitochondrial integrity.

Animals↗
PubMed2005

Inhibition of osteoclast differentiation by polycyclic aryl hydrocarbons is dependent on cell density and RANKL concentration.

We investigated the effect of representative polycyclic aryl hydrocarbons (PAHs), benzo[a]pyrene (BaP), and 7,12-dimethylbenz[a]anthracene (DMBA) on osteoclast differentiation and function by using dispersed cancellous bone derived rabbit osteoclasts and the RAW264.7 cells. These cells differentiate into osteoclasts when exposed to receptor activator of NF-kappaB ligand (RANKL). The rabbit osteoclasts were exposed to 10(-6) to 10(-9)M BaP or DMBA and the tartrate-resistant acid phosphatase (TRAP)-positive cells were counted. The effect of PAHs on osteoclast differentiation in dispersed rabbit osteoclast-containing stromal cell populations was cell density dependent, suggesting that the cell density of stromal cells, osteoclast precursors, and/or mature osteoclasts are factors regulating the effect of PAHs. To investigate the direct effect of BaP on osteoclast differentiation, RAW264.7 cells were exposed to 10(-5) to 10(-6) M BaP. Treatment of RAW264.7 cells cultured with 25 ng/ml soluble RANKL and 10(-5)M BaP for 5 days decreased osteoclast differentiation, TRAP activity levels, and resorption of bone-like substrata. The inhibition was prevented by 10(-6) to 10(-7) M resveratrol, an aryl hydrocarbon receptor (AhR) antagonist, and by higher concentrations of RANKL. To investigate the ability of RANKL to reverse BaP-mediated inhibition, gene expression was determined by RT-PCR. Cytochrome P450 1B1 (CYP1B1) mRNA, one of the genes activated by BaP, was present only in the groups exposed to BaP; the levels of CYP1B1 mRNA decreased in the presence of increasing concentrations of RANKL. These results suggest that the inhibitory effects of PAHs on osteoclastogenesis are direct and likely involve interaction of the RANKL and PAH signaling pathways.

Animals↗
PubMed2004

Alterations in mitochondrial membrane potential during preimplantation stages of mouse and human embryo development.

Mitochondria are cellular organelles regulating metabolism and cell death pathways. This study examined changes in mitochondrial membrane potential (deltapsim) throughout the stages of preimplantation development in mouse embryos conceived either in vivo or in vitro and human embryos donated to research from IVF. Embryos stained with the deltapsim-sensitive dye (JC-1) were quantified for the ratio of high- to low-polarized mitochondria using a deconvolution microscope. Overall, mouse zygotes and early embryos contain a subset of high-polarized mitochondria with a progressive increase in the ratio of deltapsim observed with increasing cleavage. A transient increase in the ratio of high to low deltapsim was observed in in vivo fertilized 2-cell stage embryos, coincident with embryonic genome activation in the mouse, but not in 2-cell embryos obtained through IVF. We further observed that arrested mouse 2-cell embryos possessed an increased ratio of deltapsim compared with non-arrested embryos. In human 8-cell embryos we observed an increased ratio of high- to low-polarized mitochondria with increasing degrees of embryo fragmentation. We concluded that the pattern of mitochondrial membrane potential progressively changes throughout preimplantation development, and that an aberrant shift in deltapsim could contribute to, or is associated with, decreased developmental potential.

Animals↗
PubMed2004

Inhibition of osteogenesis in vitro by a cigarette smoke-associated hydrocarbon combined with Porphyromonas gingivalis lipopolysaccharide: reversal by resveratrol.

BACKGROUND: Smoking and infection with Gram-negative bacterial pathogens are risk factors for alveolar bone loss. The aims of this study were: 1) to examine the combined effects of an aryl hydrocarbon, benzo[a]pyrene (BaP), that is concentrated in cigarette smoke, and lipopolysaccharide (LPS) extracted from Porphyromonas gingivalis on osteogenesis in a rat bone marrow cell (RBMC) model of osteogenesis; and 2) to determine whether resveratrol (Res), an aryl hydrocarbon receptor antagonist, could reverse the putative inhibitory effects of BaP + LPS on osteogenesis. METHODS: LPS of P. gingivalis strain 2561 was introduced in various concentrations to the RBMC in 96-well plates and kept in culture for 8 to 12 days. The same protocol was used for studying BaP and LPS + BaP combinations. Following the incubation periods, parameters of osteogenesis were measured, including formation of mineralized bone nodules, alkaline phosphatase activity, and total cell protein. Transcription of the pro-inflammatory cytokine interleukin (IL)-1beta in the cultures was determined by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Bone nodule formation generally decreased significantly with increasing LPS concentrations (P<0.05), whereas total cell protein decreased only slightly (P>0.05). BaP in previously high concentrations alone also caused a significant dose-dependent decrease in bone nodule formation (P<0.05) but when half maximal doses were used, significant decreases were most often seen when LPS was added. Hence, in combination, the inhibitory effects of LPS + BaP on osteogenesis were additive, inhibiting bone nodule formation up to 9-fold. Resveratrol partially reversed the inhibitory effects of low concentrations of LPS alone, and completely reversed the inhibition of nodule formation when low concentrations of LPS were combined with BaP. IL-1beta expression generally fluctuated inversely to the inhibitory activity of LPS, LPS + BaP, and LPS + BaP + Res combinations. CONCLUSIONS: Smoke-derived aryl hydrocarbons and bacterial LPS may act additively to inhibit bone formation. The findings may explain, in part, why net periodontal bone loss is greater and bone healing is less successful in smokers than non-smokers with periodontal infections. Reversal of the inhibitory effects in vitro by resveratrol suggests that this phytoalexin should be studied further for its potential therapeutic value, given its aryl hydrocarbon receptor antagonism and apparent anti-inflammatory activity.

Alveolar Bone Loss↗
PubMed2003

The aryl hydrocarbon receptor and its xenobiotic ligands: a fundamental trigger for cardiovascular diseases.

This review reconsiders a major cause of cardiovascular diseases, tobacco smoking, as the activation of the Aryl hydrocarbon Receptor (AhR), also known as the dioxin receptor, by aryl hydrocarbons from the tar fraction of tobacco in various organs of the cardiovascular domain. This concept sheds new light on well-known albeit controversial epidemiological concepts such as the Mediterranean diet and the French paradox. We also review the discovery that resveratrol, a natural AhR antagonist, may be of interest in the prevention and treatment of cardiovascular diseases.

Cardiovascular Diseases↗
PubMed2003

Aromatase inhibition reduces gonadotrophin dose required for controlled ovarian stimulation in women with unexplained infertility.

BACKGROUND: Adding clomiphene citrate (CC) to FSH for controlled ovarian stimulation (COS) decreases FSH dose required for optimum stimulation. However, because of its anti-estrogenic effects, CC may be associated with lower pregnancy rates offsetting the FSH-dose reduction benefit. Previously, we reported the success of aromatase inhibition in inducing ovulation without antiestrogenic effects. METHODS: A prospective pilot study that included women with unexplained infertility undergoing COS and intrauterine insemination. Thirty-six women received the aromatase inhibitor letrozole + FSH, 18 women received CC + FSH and 56 women received FSH only. Each woman received one treatment regimen in one treatment cycle. All patients were given recombinant or highly purified FSH (50-150 IU/day) starting on day 3 to 7 until day of hCG. RESULTS: The FSH dose needed was significantly lower in letrozole + FSH and CC + FSH groups compared with FSH-only without a difference in number of follicles >1.8 cm. Pregnancy rate was 19.1% in the letrozole + FSH group, 10.5% in the CC + FSH group and 18.7% in the FSH-only group. Both pregnancy rate and endometrial thickness were significantly lower in CC + FSH group compared with the other two groups. Estradiol (E2) levels were significantly lower in the letrozole + FSH group compared with the other two groups. CONCLUSIONS: Similar to CC, aromatase inhibition with letrozole reduces FSH dose required for COS without the undesirable antiestrogenic effects sometimes seen with CC.

Adult↗
PubMed2002

Polycyclic aromatic hydrocarbons present in cigarette smoke cause bone loss in an ovariectomized rat model.

A number of epidemiological studies have suggested that cigarette smoking is a risk factor for osteoporosis. Benzo(a)pyrene (BaP) and 7,12-dimethylbenz(a)anthracene (DMBA) are polycyclic aromatic hydrocarbons (PAHs) found in the tar fraction of cigarette smoke, as well as in car exhaust and furnace gases. We hypothesized that BaP and DMBA are responsible, through interaction with the aryl hydrocarbon receptor (AhR), for the bone loss and fragility seen in smoking-related osteoporosis. In this study four groups of 9-month-old Sprague-Dawley rats were examined. An intact group served as controls. A second control was the ovariectomized (ovx) group. The third group (ovx + E(2)) were ovariectomized and also given a continuous basal dose of estrogen by implanted estrogen pellet (0.085 mg of 17beta-estradiol per rat). The fourth group (ovx + E(2) + BaP/DMBA) was ovariectomized with an estradiol pellet, and received subcutaneous injections of 250 microg/kg of BaP/DMBA weekly for 15 weeks. The loss of ovarian function allowed the study of a direct effect of BaP/DMBA on bone while the concomitant estrogen repletion prevented ovx-related bone loss. Dual-energy X-ray absorptiometry (DEXA), histomorphometry, image analysis, and mechanical testing were used to determine the effect of the treatments on bone. The DEXA results showed a significant (p < 0.05) decrease in bone mineral density compared with intact controls with both ovx alone and with ovx + E(2) + BaP/DMBA treatment. The ovx + E(2) rats were similar to the intact controls. The osteoid parameters showed a significant increase (p < 0.05) with BaP/DMBA addition vs. intact controls, mimicking the ovx rats. The ovx + E(2) rats had osteoid parameters comparable to those of intact rats. Bone connectivity was decreased in the ovx and ovx + E(2) + BaP/DMBA animals. Connectivity of the ovx + E(2) rats was comparable to that of intact animals. A decrease in failure force was seen in three-point bending for the ovx + E(2) + BaP/DMBA group and in vertebral compression in both the ovx and ovx + E(2) + BaP/DMBA groups vs. intact controls. The mechanical properties of the ovx + E(2) rats were similar to those of intact rats. These results demonstrate that BaP/DMBA causes a loss of bone mass and bone strength, possibly through an increase in bone turnover. This is the first in vivo study linking environmental toxicants, found in the tar fraction of cigarette smoke and in urban air pollution, to loss of bone mass and strength in estrogen-replete ovx rats.

Animals↗
PubMed2001

Resveratrol decreases hyperalgesia induced by carrageenan in the rat hind paw.

The effect of resveratrol, an aryl hydrocarbon receptor (AhR) antagonist, known to inhibit inducible cyclooxygenase-2 (COX2) and its transcription were examined in a model of hyperalgesia induced by carrageenan in the rat. Pretreatment with resveratrol did not reverse swelling and edema, but reversed the hyperalgesia induced by local tissue injury provoked by carrageenan. This reversal, occurring at resveratrol concentrations as low as 2 mg/kg, lasted for at least 48 hours. The link with COX2 activity inhibition and COX2 gene transcription, as well as a potential AhR inhibitory effect, remain to be established.

Animals↗
PubMed2001

Use of an aromatase inhibitor for induction of ovulation in patients with an inadequate response to clomiphene citrate.

OBJECTIVE: To use aromatase inhibition for induction of ovulation in women in whom clomiphene citrate (CC) treatment was unsuccessful. DESIGN: Prospective trial in infertility patients treated with CC. SETTING: Two tertiary-referral infertility clinics associated with the Division of Reproductive Sciences, University of Toronto. PATIENT(S): Twelve patients with anovulatory polycystic ovary syndrome (PCOS) and 10 patients with ovulatory infertility, all of whom had previously received CC with an inadequate outcome (no ovulation and/or endometrial thickness of < or =0.5 cm). INTERVENTION(S): The aromatase inhibitor letrozole was given orally in a dose of 2.5 mg on days 3-7 after menses. MAIN OUTCOME MEASURE(S): Occurrence of ovulation, endometrial thickness, and pregnancy rates. RESULT(S): With CC treatment in patients with PCOS, ovulation occurred in 8 of 18 cycles (44.4%), and all ovulatory cycles for the women included in this study had endometrial thickness of < or =0.5 cm. In 10 ovulatory patients, 15 CC cycles resulted in a mean number of 2.5 mature follicles, but all cycles had endometrial thickness of < or =0.5 cm on the day of hCG administration. With letrozole treatment in the same patients with PCOS, ovulation occurred in 9 of 12 cycles (75%) and pregnancy was achieved in 3 patients (25%). In the 10 patients with ovulatory infertility, letrozole treatment resulted in a mean number of 2.3 mature follicles and mean endometrial thickness of 0.8 cm. Pregnancy was achieved in 1 patient (10%). CONCLUSION(S): Oral administration of the aromatase inhibitor letrozole is effective for ovulation induction in anovulatory infertility and for increased follicle recruitment in ovulatory infertility. Letrozole appears to avoid the unfavorable effects on the endometrium frequently seen with antiestrogen use for ovulation induction.

Anovulation↗
PubMed2001

Effects of chilling to 0 degrees C on the morphology of meiotic spindles in human metaphase II oocytes.

OBJECTIVE: To determine the effects of chilling to 0 degrees C on the meiotic spindle of human metaphase II oocytes, as observed by optical sectioning microscopy. DESIGN: Laboratory study. SETTING: Academic research laboratory in a medical school. PATIENT(S): Seventy-two women undergoing infertility treatment donated a total of 108 oocytes. INTERVENTION(S): Metaphase II oocytes were stripped of their cumulus cells, cooled directly to 0 degrees C, and held for periods of 1 to 10 minutes. They were then fixed at 37 degrees C, stained for immunofluorescence, and examined microscopically. MAIN OUTCOME MEASURE(S): Morphology of the meiotic spindle in chilled and control oocytes. RESULT(S): Microscopic evaluations of 46 chilled oocytes revealed various time-dependent changes in microtubules compared to 9 control oocytes. After 1 minute at 0 degrees C, spindle damage was negligible, but in oocytes cooled for 2 or 3 minutes, there was obvious shortening of the spindle and loss of polarity. Cooling to 0 degrees C for 4 to 9 minutes resulted in increasingly more drastic changes; by 10 minutes the spindles had totally disappeared. Despite depolymerization of microtubular tubulin at 0 degrees C, the chromosomes did not become dispersed, but remained anchored even in the absence of spindles. CONCLUSION(S): Even brief exposure of human oocytes to temperatures near 0 degrees C causes profound alterations of the meiotic spindle.

Chromosomes, Human↗
PubMed2001

Aromatic hydrocarbon receptor-driven Bax gene expression is required for premature ovarian failure caused by biohazardous environmental chemicals.

Polycyclic aromatic hydrocarbons (PAHs) are toxic chemicals released into the environment by fossil fuel combustion. Moreover, a primary route of human exposure to PAHs is tobacco smoke. Oocyte destruction and ovarian failure occur in PAH-treated mice, and cigarette smoking causes early menopause in women. In many cells, PAHs activate the aromatic hydrocarbon receptor (Ahr), a member of the Per-Arnt-Sim family of transcription factors. The Ahr is also activated by dioxin, one of the most intensively studied environmental contaminants. Here we show that an exposure of mice to PAHs induces the expression of Bax in oocytes, followed by apoptosis. Ovarian damage caused by PAHs is prevented by Ahr or Bax inactivation. Oocytes microinjected with a Bax promoter-reporter construct show Ahr-dependent transcriptional activation after PAH, but not dioxin, treatment, consistent with findings that dioxin is not cytotoxic to oocytes. This difference in the action of PAHs versus dioxin is conveyed by a single base pair flanking each Ahr response element in the Bax promoter. Oocytes in human ovarian biopsies grafted into immunodeficient mice also accumulate Bax and undergo apoptosis after PAH exposure in vivo. Thus, Ahr-driven Bax transcription is a novel and evolutionarily conserved cell-death signaling pathway responsible for environmental toxicant-induced ovarian failure.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed2001

Effect of centrifugation on early embryonic development and parthenogenetic activation of bovine oocytes matured in vitro.

This study examined the fertilization, early developmental competence and capacity for parthenogenetic activation of bovine oocytes matured in vitro after centrifugation. Immature oocytes were cultured in tissue culture medium 199 supplemented with 10% fetal bovine serum and 75 mIU mL(-1) FSH + LH at 5% CO2 to facilitate maturation. After culture for 24 or 30 h, the metaphase-II stage oocytes were centrifuged at 3000, 5000, 7000 or 10000g for 5 min before in vitro fertilization or parthenogenetic activation. Frozen-thawed bull semen was used for in vitro fertilization. For parthenogenetic activation, the oocytes were exposed to 20 microM calcium ionophore A23187 for 5 min at room temperature. Fertilization rates were not different between control and treatment groups (87.7% v. 74.6%, 73.4%, 75.9% and 76.4% respectively). Also, there were no differences in early embryonic development between control and treatment groups (rates of blastocyst formation were 21.1% v 20.2%, 28.8%, 31.2% and 24.1% respectively). When the oocytes were centrifuged at various speeds alone, the activation rate of oocytes was significantly higher (P < 0.05) in the 10000g treatment group compared with control (10.8% v 0.0%). There were no differences in the activation rates of oocytes between control and treatment groups at speeds up to 7000g (70.9% v. 71.9%, 78.3% and 77.2% respectively) after centrifugation and stimulation with Ca(2+)-ionophore. However, the activation rate of oocytes was significantly higher (P < 0.05) in the 10000g treatment group compared with control (70.9% v. 83.1%). In addition, the percentage of activated oocytes with diploid formation was significantly higher in the oocytes after centrifugation at 10000g and stimulation with calcium ionophore A23187 than in the control (18.4% v 7.1%). These results indicate that centrifugation of oocytes matured in vitro has no detrimental effect on fertilization and subsequent early embryonic development. They also indicate that the oocytes might be parthenogenetically activated after centrifugation and that high-speed centrifugation may induce activation of some oocytes. The results suggest that the optimal speed for centrifugation of bovine oocytes might be < or = 7000g to enhance the visibility of nuclear elements for further micromanipulation.

Animals↗
PubMed2001

Human UC-blood banking: impact of blood volume, cell separation and cryopreservation on leukocyte and CD34(+) cell recovery.

BACKGROUND: UC blood represents an increasingly useful source of hematopoietic stem cells for BMT, although, currently, low cell numbers generally limit its use to pediatric patients. We have determined parameters that influence the recovery of viable cells during processing and cryopreservation, in an effort to set guidelines for determining whether a sample will yield sufficient cells to be of use in the transplant setting. METHODS: UC blood was collected from donors from January 1996 to December 1999. Volume was reduced using Ficoll, followed by cryopreservation under liquid nitrogen. Total leukocyte and CD34(+)-cell counts were determined prior to processing and a subset of samples was also assessed post-processing and post-cryopreservation. RESULTS: Outcomes for 3816 samples were analyzed to determine the correlation between cell number, cell type, volume, and time between collection and processing. A positive relationship was observed between volume and cell number for both leukocytes and the CD34(+) cells. This correlation allowed us to determine the number of leukocytes and CD34(+) cells expected from a sample based on volume, and to set guidelines for determining the practicality of storing any given sample prior to processing and cryopreservation. DISCUSSION: Measuring blood volume gives a very useful indication of the total leukocyte and CD34(+) cell number. The majority (75%) of cord-blood samples contain sufficient leukocytes for a pediatric transplant, and the number of cells available can be determined prior to processing by measuring blood volume.

Antigens, CD34↗
PubMed2001

Controlling culture dynamics for the expansion of hematopoietic stem cells.

The ex vivo expansion of hematopoietic stem cells (HSCs) is the subject of intense commercial and academic interest due to the potential of HSCs to be a renewable source of material for cellular therapeutics. Unfortunately, because methodologies have not yet been developed to grow clinically relevant numbers of HSCs (or their derivatives) consistently, the potential of this technology is limited. Manipulation of the in vitro culture microenvironment, primarily through cytokine supplementation, has been the predominant approach in studies attempting to expand primary human HSC numbers in vitro. While promising results have been obtained, it is becoming clear that novel methods must be developed before cellular therapies using these stem cells can become routine. Ideally, bioprocesses must be designed to target specifically the growth of stem cell populations while incorporating positive and negative feedback from potentially dynamic mature and maturing cell populations. The product of these culture systems should consist of not only HSCs, but also of cells that allow the engraftment of HSCs and, ideally, cells responsible for the immediate or accelerated functional support of patients. Development of such "designer transplants" will require combining optimal culture conditions capable of amplifying HSC numbers with novel approaches for finely controlling the number, functional capabilities, and characteristics of potentially therapeutic cells in these very complex cell culture systems.

Animals↗
PubMed2001

Tracking of oocyte dysmorphisms for ICSI patients may prove relevant to the outcome in subsequent patient cycles.

BACKGROUND: We determined whether oocyte dysmorphisms, especially repetition of specific dysmorphisms from cycle to cycle, had a prognostic impact on intracytoplasmic sperm injection (ICSI) outcome. METHODS: ICSI patients (n = 67) were grouped as follows: group 1 >50% phenotypically dysmorphic oocytes per cohort (cytoplasmic and extra-cytoplasmic dysmorphisms) with no repetition of a specific dysmorphism from cycle one to cycle two (36 cycles and 274 oocytes); group 2 >50% dysmorphic oocytes per cohort and repetition of the same dysmorphism from cycle one to cycle two (32 cycles and 313 oocytes); group 3 (control) <30% dysmorphic oocytes (33 cycles and 378 oocytes). RESULTS: In group 2 (repetitive), 47% of oocytes were observed to have organelle clustering versus 20.5% in group 1 and 17.3% in group 3 (P < 0.001). There was no difference between the groups in fertilization rates, cleavage rates or embryo quality. Embryos derived from normal oocytes were transferred in each group (57, 33 and 72% respectively). The clinical pregnancy and implantation rates in group 2 (3.1 and 1.7% respectively) were lower (P < 0.01, P = 0.005) than both group 1 (28 and 15% respectively) and group 3 (45.5 and 26.5% respectively). CONCLUSIONS: The low implantation rate in group 2, even though 33% of transferred embryos were derived from morphologically normal oocytes, suggests that repetitive organelle clustering may be associated with an underlying adverse factor affecting the entire follicular cohort.

Adult↗
PubMed2001

Sperm swim-up techniques and DNA fragmentation.

BACKGROUND: Swim-up techniques for sperm separation may have detrimental effects on sperm DNA. We wished to determine whether the normal swim-up method with centrifugation used in our laboratory, which involves a centrifugation step, was harmful to sperm compared with swim-up without centrifugation. METHODS: Semen samples were obtained from patients undergoing IVF or andrology assessment. An aliquot was removed for fixation and subsequent DNA fragmentation determination. The remaining sample was divided into two equal parts, which were subjected to swim-up either with (normal swim-up) or without (direct-swim-up) centrifugation. Semen analysis was performed both before and after swim-up. DNA fragmentation, in spermatozoa previously fixed in 4% paraformaldehyde, was assessed by the terminal transferase-mediated DNA end-labelling procedure (TUNEL). The percentage of spermatozoa with DNA damage after each swim-up technique was compared with that in the original semen sample. RESULTS: DNA damage was <5% in most samples. No significant change in DNA fragmentation was observed between the two swim-up procedures, although the 'normal' swim-up sample had significantly less DNA fragmentation than the pre-swim-up sample. CONCLUSIONS: We conclude that our normal swim-up technique caused no more DNA damage to spermatozoa from normal semen samples than a direct swim-up technique that involved no centrifugation step.

Adult↗
PubMed2000

7-ketocholesterol is an endogenous modulator for the arylhydrocarbon receptor.

We have identified 7-ketocholesterol (7-KC) as an endogenous modulator that inhibits transactivation by the arylhydrocarbon receptor (AhR) through competitive binding against xenobiotic ligands. 7-KC binds AhR and displaces labeled dioxin (2,3,7,8-tetrachlorodibenzo(p)dioxin (TCDD)). IC(50) is 5 x 10(-7) m in vivo and 7 x 10(-6) m in vitro. These figures are consistent with its concentration in human blood plasma and tissues. Association with 7-KC prevents AhR binding to DNA. 7-KC blocks the TCDD-mediated transactivation of stably expressed reporter gene constructs in T47-D cells as well as the expression of the endogenous CYP 1A1 gene in HepG2 cells and in primary porcine aortic endothelial cells. Injection of 7-KC to rats blocks the induction of CYP 1A1 messenger RNA and protein in endothelial cells from myocardial blood vessels. The differential sensitivity of mammalian species to toxic effects of AhR ligands, especially dioxin (TCDD), correlates with the expression of 7-hydroxycholesterol dehydrogenase, which synthesizes 7-KC from 7-hydroxycholesterol. The documented involvement of AhR ligands in cardiovascular diseases through lipid peroxidation and endothelium dysfunction can now be examined in the context of displacement of this protective modulator.

Animals↗