help button home button Endocrine Society Endocrinology
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ben-Shlomo, I.
Right arrow Articles by Adashi, E. Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ben-Shlomo, I.
Right arrow Articles by Adashi, E. Y.
Endocrinology Vol. 138, No. 7 2680-2688
Copyright © 1997 by The Endocrine Society

ARTICLES

Interleukin (IL)-1ß Increases Glucose Uptake and Induces Glycolysis in Aerobically Cultured Rat Ovarian Cells: Evidence That IL-1ß May Mediate the Gonadotropin-Induced Midcycle Metabolic Shift1

Izhar Ben-Shlomo2, Shahar Kol3, Lois M. Roeder, Carol E. Resnick, Arye Hurwitz4, Donna W. Payne and Eli Y. Adashi5

Division of Reproductive Endocrinology (I.B.-S., S.K., C.E.R., A.H. D.W.P., E.Y.A.), Department of Obstetrics and Gynecology, and Division of Pediatric Research (L.M.R.), Department of Pediatrics, University of Maryland School of Medicine, Baltimore, Maryland 21201

Address all correspondence and requests for reprints to: Dr. Eli Y. Adashi or Dr. Donna W. Payne, The University of Utah, Department of Obstetrics and Gynecology, Health Sciences Center, Suite 2B200, 50 North Medical Drive, Salt Lake City, Utah 84132. E-mail: eadashi{at}hsc.utah.edu


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
This communication explores the possibility that interleukin (IL)-1ß, a putative intermediary in the ovulatory process, may take part in the gonadotropin-driven midcycle diversion of ovarian carbohydrate metabolism toward glycolysis. We examined the effect of treatment with IL-1ß on glucose metabolism in aerobically cultured whole ovarian dispersates from immature rats. Treatment with IL-1ß increased cellular glucose consumption/uptake, stimulated extracellular lactate accumulation and media acidification, and decreased extracellular pyruvate accumulation in a receptor-mediated, time-, dose- and cell density-dependent manner. Endogenous IL-1ß-like bioactivity was shown to mediate the ability of gonadotropins to exert these same metabolic effects. The IL-1ß effect was also (1) apparent over a broad range of glucose concentrations, inclusive of the putative physiological window; (2) relatively specific, because tumor necrosis factor-{alpha} and insulin were inactive; (3) contingent upon cell-cell cooperation (4) and reliant on de novo protein synthesis. Comparison of the molar ratios of lactate accumulation to glucose consumption in IL-1ß-replete vs. IL-1ß-deplete cultures suggests that IL-ß promotes the conversion of all available glucose to lactate but that other substrates for lactate production may also exist. However, no lactate was generated by cells grown under glucose-free conditions. Taken together, our data suggest that IL-1ß may act as a metabolic hormone in the ovary. Subject to the limitations of the in vitro paradigm, our data also suggest that IL-1ß may mediate the gonadotropin-associated midcycle shift in ovarian carbohydrate metabolism. By converting the somatic ovarian cells into a glucose-consuming glycolytic machinery, IL-1ß may establish glycolysis as the main energy source of the relatively hypoxic preovulatory follicle and the resultant cumulus-oocyte complex. The consequent oxygen sparing may conserve the limited supply of oxygen needed for vital biosynthetic processes such as steroidogenesis. This adaptational response may also provide the glycolytically incompetent oocyte with the obligatory tricarboxylic cycle precursors it depends on to meet the increased energy demands imposed upon it by the resumption of meiosis.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
A GROWING BODY of direct and indirect evidence supports the notion that intraovarian interleukin (IL)-1ß1 may be an intermediary in the ovulatory process (1). First, the provision of IL-1ß has been shown to bring about ovulation and to synergize with LH in this regard (2, 3). Second, the addition of an IL-1 receptor antagonist (IL-1RA) has been shown to attenuate LH-supported ovulation (4, 5). Third, some components of the intraovarian IL-1 system (e.g. IL-1ß and the type I IL-1 receptor) appear to be expressed only during a narrow periovulatory window (6, 7, 8, 9). Fourth, IL-1ß has been shown to induce a host of ovulation-associated phenomena (10, 11, 12, 13).

Yet another corollary of ovulation is the so called metabolic shift, a phenomenon first described more than 30 yr ago by Armstrong and Greep (14). This effect is characterized by the enhancement of glucose uptake as well as by the diversion of ovarian carbohydrate metabolism away from oxidative phosphorylation towards glycolysis (14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24). Although the precise teleologic rationale for the metabolic shift remains unknown, it is possible that it constitutes an adaptational response. Such an adaptational response may be designed to combat the anticipated relative hypoxia experienced by the follicle- or cumulus-enclosed oocyte (24, 25). Moreover, the resultant high throughput of monocarboxylate substrates may be designed to meet the increased energy needs of the meiotically active oocyte that cannot process glucose by itself and therefore relies on alternative energy substrates such as pyruvate and lactate (26, 27, 28, 29, 30).

Because intraovarian IL-1ß appears to be gonadotropin dependent (6) and IL-1ß has been shown to produce an aerobic glycolytic state in several extraovarian cell types (31, 32, 33), we hypothesized that IL-1ß may mediate the gonadotropin-induced midcycle conversion of ovarian metabolism towards glycolysis. To test this possibility, we examined the effect of treatment with IL-1ß on glucose consumption and uptake, and on lactate and pyruvate formation using aerobically cultured whole ovarian dispersates from immature rats. Our results indicate that IL-1ß, of either exogenous or endogenous origin, is capable of inducing glycolysis in aerobically cultured ovarian cells, an effect accompanied by up-regulation of glucose uptake. These findings support the proposition that ovarian IL-1ß-like activity mediates the periovulatory, gonadotropin-induced shift in ovarian glucose metabolism.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals
Immature (25–28 days old) Sprague-Dawley female rats, purchased from Zivic-Miller Laboratories (Zelienople, PA), were killed by CO2 asphyxiation. This project was approved by the Institutional Animal Care and Use Committee.

Reagents and hormones
Regular and glucose-free McCoy’s 5a media (modified, without serum), penicillin-streptomycin solution, L-glutamine, trypan blue stain (0.4%), BSA, and HEPES buffer were from Life Technologies Life Sciences (Grand Island, NY). Collagenase (Clostridium Histolyticum; CLS type I; 183 U/mg) was from Worthington Biochemical Corp. (Freehold, NJ). DNase (bovine pancreas; 2100 U/mg), aminoguanidine hemisulfate salt, and sodium nitroprusside were obtained from Sigma Chemical Co. (St. Louis, MO). [3H]L-glucose (20 Ci/mmol), [1,2-3H]2-deoxy-D-glucose (30.8 Ci/mmol) and [3H]3–0-methyl-D-glucose (60–90 Ci/mmol) were obtained from DuPont New England Nuclear Research Products (Boston, MA).

Recombinant human IL-1ß (2 x 107 U/mg) was generously provided by Drs. Errol B. De Souza and C. E. Newton of DuPont-Merck Pharmaceutical Co. (Wilmington, DE). Recombinantly expressed, naturally occurring human IL-1 receptor antagonist (IL-1RA) was generously provided by Dr. Jerome F. Strauss III (Philadelphia, PA). Highly purified human CG (hCG; CR-127, 14, 900 U/mg) and ovine FSH (oFSH: NIH-FSH-S18, FSH potency equal to 65.6 NIH-FSH-S1 U/mg) were gifts of the National Hormone and Pituitary Program (Rockville, MD), through the NIADDK, NICHHD, and USDA. Bovine insulin was obtained from Sigma Chemical Co. (St. Louis, MO). Recombinant human tumor necrosis factor-{alpha} (TNF{alpha}; 5 x 107 U/ng) was generously provided by Dr. Jennie Mather (Genentech, Inc., San Francisco, CA).

Tissue culture
Whole ovarian dispersates were prepared as previously described (34). Purified granulosa cells were extracted by repeated follicular puncture as previously described (35). Highly purified theca-interstitial cell preparations from immature hypophysectomized rats were isolated as previously described (36). Cells were cultured in 35 x 10-mm tissue culture dishes (Falcon Plastics, Oxnard, CA) containing 1 ml of McCoy’s 5a medium (modified, without serum). Media were supplemented with L-glutamine (2 mM), penicillin (100 U/ml), and streptomycin (100 µg/ml). Cell cultures were maintained for up to 96 h at 37 C under a water-saturated atmosphere of 5% CO2 and 95% air. At the end of each experiment, collected conditioned media were stored at -20 C until assayed for their glucose or lactate content as described below. Pyruvate was assayed in freshly conditioned media.

Lactate assay
Lactate concentrations in conditioned media were determined by a modification of a previously described method (37) in which the NADH product is monitored at 340 nm after the NAD-linked conversion of lactate to pyruvate by lactate dehydrogenase with hydrazine trapping of pyruvate to ensure that the reaction goes to completion. The assay was adapted as follows to a microtiter plate system using premixed reagents from a commercially available kit (Sigma no. 826-A, Sigma Chemical Co., St. Louis, MO). Conditioned media (20 µl of 20-fold diluted) or varying volumes (20–100 µl) of L-lactic acid standard (50 µg/ml) were diluted in the microtiter plate wells to a volume of 100 µl after which were added NAD/glycine buffer solution, pH 9.2 (75 µl), and lactate dehydrogenase solution (25 µl). After a 30-min incubation period at ambient temperature, absorbance (340 nm) was read in a UVmax plate reader (Molecular Devices, Menlo Park, CA). The standard curve was linear in the range used (1–25 µg/ml), and media blanks were negligible. Lactate concentrations were calculated from the standard curve using a software package ({Delta}Soft, BioMetallics, Inc., Princeton, NJ) designed for the plate reader. The within-assay coefficients of variation were 14.5 and 6.9% and the between-assay coefficients of variation were 11.7 and 6.7% for the low and high standards, respectively.

Glucose assay
Glucose concentrations in conditioned media were determined by the modification of a previously described enzymatic method (38) wherein the glucose oxidase and peroxidase reactions are coupled with the chromogenic oxygen acceptor o-dianisidine. The assay was adapted as follows to a microtiter plate system using premixed reagents from a commercially available kit (Sigma no. 510-A, Sigma). Unconditioned or conditioned media (10 µl of 20-fold diluted) or varying volumes (1.5–30 µl) of standard glucose solution (50 µg/ml) were diluted in the microtiter plate wells to a volume of 30 µl to which were added the color reagent/enzyme solution (200 µl), prepared as described by the manufacturer. After a 45-min incubation period at ambient temperature, absorbance of oxidized o-dianisidine (450 nm) was read in the UVmax plate reader. The standard curve was linear in the range used (2.5–50 µg/ml). Glucose concentrations were calculated from the standard curve. The within-assay coefficients of variation were 2.7 and 1.2% and the between-assay coefficients of variation were 3.2 and 1.5% for the low and high standards, respectively. The values for glucose consumption reported herein were calculated by subtracting the value determined in conditioned media (a value containing assay variability, see below) from the glucose value in unconditioned media. This latter number is nominally 3 mg glucose/ml media (16.7 mM), but in reality varied slightly due to modest evaporation during culture. A value of 3.02 ± 0.13 mg glucose/ml (range 2.9–3.3 mg/ml) was actually measured in McCoy’s media after 96 h in cell-free cultures (n = 12).

In preliminary studies, glucose was also measured by a previously described coupled reaction (39) in which glucose is first phosphorylated and the glucose-6-phosphate formed is oxidized in the presence of NAD. When we compared glucose measurement by this technique to that of the glucose oxidase method described above, both methods gave comparable results. The correlation coefficient (r2) for the two methods, using data from five different experiments (n = 36 determinations for each method), was 0.66 (P = 0.0001). The slope (glucose oxidase method/NAD method) was 1.03 (P = 0.0001), indicating that the two methods gave essentially the same value for glucose concentration.

Pyruvate assay
Pyruvate concentrations in conditioned media were assayed by the same enzymatic reaction used to measure lactate as described above, but in the reverse direction (37). The loss of NADH is monitored at 340 nm after the NADH-linked conversion of pyruvate to lactate by lactate dehydrogenase. The assay was adapted as follows to a microtiter plate system using premixed reagents from a commercially available kit (Sigma no. 726-UV). Unconditioned or conditioned media (100 µl), or varying volumes (5–60 µl) of pyruvic acid standard (0.45 mM) were diluted in the microtiter plate wells to a volume of 100 µl after which an additional 100 µl of NADH (0.67 mM) in Tris buffer (1.5 M) was added. Initial absorbance (340 nm) was then read in a UVmax plate reader after which 25 µl lactate dehydrogenase solution (160 U/ml) was added to initiate the reaction. After a 15-min incubation period at ambient temperature, final absorbance (340 nm) was read and the change in absorbance was calculated for each sample. Pyruvate concentrations were calculated from the standard curve.

Glucose uptake assay
The rate of glucose transport was measured using the glucose analogs [1,2-3H]2-deoxy-D-glucose or [3H]3–0-methyl glucose. Plated, pretreated cells were washed (x3) with HEPES buffer, pH 7.4, and the final wash immediately replaced with 1 ml of the reaction mix containing 1 mM of 3H-labeled and unlabeled substrate. The dishes were then placed on a rotary shaker and incubated at room temperature. At the end of the reaction time (up to 15 min), the reaction mixture was aspirated off and the cells quickly washed (x4) with stopping solution [NaCl (0.9%); HgCl2 (10-5 M); phloretin (10-4 M)]. The cells were allowed to dry overnight, then dissolved in NaOH (0.1 M). A portion of each sample was transferred to a plastic counting vial, scintillation fluor added, and the radioactivity determined. Nonspecific uptake (accounting for nontransporter-mediated transmembranal glucose passage) was measured for each time point by incubating the cells with a mixture of 3H-labeled and unlabeled L-glucose (final concentration = 1 mM). Net specific uptake was calculated by subtracting the nonspecific from the total uptake, the results being expressed as pmol/min·mg protein.

Data analysis
Data are presented as the mean ± SE of replicate experiments (n noted on figures), each performed in duplicate. Statistical significance is denoted in the figures and was determined by ANOVA analysis for comparison of multiple groups (Fisher’s protected least significant difference) or by t test for comparison of two groups, using Statview 512+ for MacIntosh (Brain Power, Inc., Calabasas, CA).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
IL-1ß-stimulated lactate accumulation and glucose consumption by cultured whole ovarian dispersates: dose dependence and specificity
To determine the effect of treatment with IL-1ß on glucose consumption and lactate generation, whole ovarian dispersates were cultured for 96 h under serum-free conditions in the absence or presence of increasing concentrations (0.1–50 ng/ml) of IL-1ß. As shown (Fig. 1Go), the basal consumption of glucose and the formation of lactate over the 96-h culture period were 2.6 ± 0.5 and 5.7 ± 1.1 mM, respectively. Treatment with IL-1ß produced dose-dependent increments in lactate accumulation (open circles) and glucose consumption (closed circles), the first statistically significant (P < 0.05) increments occurring at IL-1ß concentrations of 0.5 and 1.0ng/ml, respectively. A maximal IL-1ß effect (3- to 4-fold increase over untreated controls) was noted at 3–10 ng/ml, with an ED50 in the range of 0.3 to 0.4 ng/ml of IL-1ß. In this and subsequent experiments, it is apparent that at maximal IL-1ß stimulation, lactate accumulation (in molar terms) is 2- to 3-fold that of glucose consumption. This suggests that all available glucose is converted to lactate but that other substrates for lactate production may also exist.



View larger version (23K):
[in this window]
[in a new window]
  		Figure 1. IL-1ß-mediated glucose consumption and lactate accumulation by cultured whole ovarian dispersates: dose response. Whole ovarian dispersates (5 x 105 viable cells/dish) from immature rats were aerobically cultured for 96 h in the absence or presence of increasing concentrations (0.1–50ng/ml) of IL-1ß. At the conclusion of this treatment period, medium lactate accumulation (open circles) and glucose consumption (closed circles) were determined as described under Materials and Methods.

 
Cellular lactate concentrations were also determined after rapid (<10 sec) washing and acidification of cells (to prevent further metabolism). Under these conditions, no lactate could be detected, suggesting that lactate is efficiently secreted, most likely via the monocarboxylate transporter (40).

The pH of media (normally pH 7.4) conditioned by IL-1ß-treated cells was lowered by approximately 0.5 U as evidenced by the distinct yellowish color of the phenol red indicator dye. Media acidity apparently reflected the accumulation of lactic acid since IL-1ß-treated cells that did not accumulate lactate (e.g. those cultured in glucose-free media, see below) did not demonstrate a pH change. The change in pH did not account for the metabolic alterations under study in that qualitatively comparable findings were documented in appropriately buffered (pH stable) media (not shown).

The specificity of the IL-1ß metabolic effect was assessed using two other established ovarian agonists (34, 35). In contrast to IL-1ß, maximally effective doses of TNF{alpha} (a cytokine possessing an overlapping range of action) or insulin (an established metabolic hormone) were unable to stimulate glucose consumption or lactate accumulation (Fig. 2Go). The inability of insulin to promote ovarian glucose consumption is in keeping with preliminary data (not shown) suggesting that the ovary does not express insulin-regulatable glucose transporters.



View larger version (49K):
[in this window]
[in a new window]
  		Figure 2. IL-1ß-induced aerobic glycolysis: specificity. Whole ovarian dispersates (5 x 105 viable cells/dish) were aerobically cultured for 96 h in the absence or presence of IL-1ß (50 ng/ml), TNF{alpha} (10 ng/ml), or insulin (1 µg/ml) after which medium lactate concentration (shaded bars) and glucose consumption (hatched bars) were determined.

 
IL-1ß-stimulated aerobic glycolysis: Glucose dependence and receptor mediation
To assess the possibility of lactate-generating sources other than glucose, to ascertain the role of the ambient glucose concentration, and to examine the role of the IL-1 receptor, whole ovarian dispersates were cultured for 96 h in the absence or presence of IL-1ß, with and without the addition of a naturally occurring IL-1 receptor antagonist (IL-1RA). In addition, cultures were incubated in the absence or presence of various concentrations of glucose (1–3 g/liter). Under glucose-free conditions (Fig. 3Go), no lactate was produced, demonstrating that media glucose is required for IL-1ß-induced lactate accumulation. This phenomenon could not be attributed to hypoglycemia-induced cellular dysfunction since cells grown in the absence of glucose were judged normal by the following criteria: normal cellular morphology, IL-1ß-stimulated nitric oxide synthase activity, IL-1ß-stimulated glucose transporter transcripts, and stable transcripts corresponding to ribosomal large protein 19, a housekeeping gene (Kol S., and Ajashi E., unpublished observations).



View larger version (22K):
[in this window]
[in a new window]
  		Figure 3. IL-1ß-induced aerobic glycolysis: Glucose dependence and receptor mediation. Whole ovarian dispersates (5 x 105 viable cells/dish) were aerobically cultured for 96 h using the following treatment paradigms at each of the noted glucose concentrations (0, 1, 2, 3g/liter): Control (no treatment); IL-1ß (50ng/ml); IL-1RA (IL-1 receptor antagonist; 5 µg/ml); IL-1ß+IL-1RA (50ng/ml and 5 µg/ml, respectively). At the conclusion of the treatment period, medium lactate concentration (upper panel) and glucose consumption (lower panel) were determined.

 
The IL-1ß effect on glucose consumption and lactate accumulation was apparent over a broad range of glucose concentrations (Fig. 3Go; solid circles) inclusive of the putative physiological window (100–200 mg/dl; 41). The IL-1ß-induced metabolic effect was glucose dose dependent up to 2 g/liter. No further increases were observed at the concentration of glucose in McCoy’s 5a media (3 g/liter) that was used for most of the experiments reported herein.

The IL-1ß action on glucose consumption and lactate accumulation was receptor-mediated because addition of IL-1RA (Fig. 3Go; open triangles), a reagent previously validated at the ovarian level (42), blocked the IL-1ß effect thereby approximating control levels (closed triangles) at all glucose concentrations. Treatment with IL-1RA alone (open squares) was without significant effect.

Comparison of lactate accumulation and glucose consumption under IL-1ß-replete and IL-1ß-deplete conditions
To compare the relative contribution of glucose consumption to lactate accumulation at the extremes of IL-1ß exposure, whole ovarian dispersates were cultured for varying durations with either IL-1ß (IL-1ß-replete) or a maximally inhibitory dose of IL-1RA (IL-1ß-deplete). Exposure of whole ovarian dispersates to maximally stimulatory levels of IL-1ß markedly increased the molar ratio of lactate produced to glucose consumed. The resultant ratio (2.5 ± 0.2; range 2.2–2.9; n = 3) was higher than 2 (the ratio expected if glucose is completely converted to lactate), suggesting, consistent with previous reports (16, 43), that substrates other than glucose may be used for lactate production. Nevertheless, if other sources (e.g. amino acids) exist, they are only processed in the presence of glucose because no lactate is produced in the total absence of glucose (cf. Fig. 3Go). All told, these data highlight the remarkable efficiency of the IL-1ß-stimulated conversion of glucose to lactate. In contrast, given IL-1ß-deplete conditions, i.e. an IL-1ß vacuum created by the addition of IL-1RA (42), the ratio of lactate accumulation to glucose consumption, (0.99 ± 0.4; range 0.5–1.8; n = 3) suggests a higher rate of oxidative phosphorylation or the possible diversion of glucose to nonglycolytic pathways. The large variability noted in this ratio for IL-1RA-treated cultures probably reflects the fact that the absolute consumption of glucose and the production of lactate in this state is very low, the values in question being at the limit of detection. These data also emphasize the stimulatory effect of both endogenous and exogenous IL-1ß on ovarian glycolysis.

IL-1ß-stimulated aerobic glycolysis: time and cell density dependence
To evaluate the time requirements of the IL-1ß effect, whole ovarian dispersates were cultured for the duration indicated in the absence or presence of a maximal stimulatory dose of IL-1ß. As shown (Fig. 4Go), treatment with IL-1ß resulted in marked enhancement (of up to 2.2-fold) of both parameters beginning with the 48-h time point. Whereas treatment with IL-1ß produced a statistically significant increment in lactate accumulation by 48 h, significant alteration in glucose consumption was only documented by the 72 h time point. No IL-1ß effect was noted for a series of time points during the first 48 h (not shown). This initial delay is suggestive of the induction of an intermediate factor(s) between IL-1ß stimulation and the final endpoint of glucose consumption and lactate production. In fact, when cells were cultured in the presence of cycloheximide (0.1 µg, an inhibitor of protein synthesis), IL-1ß-stimulated glucose consumption and lactate accumulation was completely inhibited (n = 3 experiments; not shown). These data suggest that protein synthesis is required for IL-1ß-driven glycolysis.



View larger version (23K):
[in this window]
[in a new window]
  		Figure 4. IL-1ß-induced aerobic glycolysis: time- and cell density dependence. Whole ovarian dispersates (5 x 105 viable cells/dish) were aerobically cultured for up to 96 h in the absence or presence of IL-1ß (50ng/ml), after which medium lactate concentration (upper panel) and glucose consumption (lower panel) were determined. Inset: Cells were cultured in the absence or presence of IL-1ß (50ng/ml) for 96 h at different cell densities (0.25–500 x 103 cells/dish).

 
Increases in glucose consumption and lactate accumulation were also cell density dependent (Fig. 4Go; inset for lactate only), over a cell density range of 0.25 to 5 x 105 cells/ml. However, a statistically significant (P < 0.01, n = 4) difference between control and IL-1ß-treated cells was observed only at a cell density of 5 x 105 cells/ml.

Effect of treatment with gonadotropins on glucose consumption and lactate accumulation: intermediary role of endogenous IL-1ß
Because gonadotropins have previously been implicated in the induction of the midcycle metabolic shift (14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24), we examined the impact of treatment with gonadotropins on ovarian glycolysis as well as on the intermediary paracrine role of endogenous IL-1ß in this regard. Whole ovarian dispersates were cultured for 96 h in the absence or presence of maximally stimulating doses of hCG or FSH, with or without IL-1RA. As shown (Fig. 5Go), treatment with either FSH or hCG produced significant (P < 0.05), albeit modest (30–40%), increments in glucose consumption and lactate accumulation. Furthermore, the concurrent provision of IL-1RA, significantly (P < 0.05) reduced the FSH and hCG effects to below control levels thereby implicating endogenous IL-1-like activity as an intermediary in this process.



View larger version (31K):
[in this window]
[in a new window]
  		Figure 5. Gonadotropin-stimulated glucose consumption and lactate accumulation. Whole ovarian dispersates (5 x 105 viable cells/dish) were aerobically cultured for 96 h in the absence or presence of hCG (10 ng/ml) or FSH (100 ng/ml), with or without IL-1RA (5 µg/ml). At the conclusion of the treatment period, medium lactate concentration (upper panel) and glucose consumption (lower panel) were determined (n is noted in parentheses for each treatment group).

 
The role of cell-cell interaction in IL-1ß-induced aerobic glycolysis
To assess the relative contribution of the different populations of ovarian somatic cells to the IL-1ß-induced metabolic effect, whole ovarian dispersates, isolated granulosa cells, purified theca-interstitial cells, or contact-dependent cocultures thereof were cultured for 96 h in the absence or presence of IL-1ß. As shown (Fig. 6Go), treatment of whole ovarian dispersates with IL-1ß, resulted in 1.5- and 2.2-fold increments (P < 0.05) in glucose consumption and lactate accumulation, respectively (cf Fig. 1Go). In contrast, treatment of isolated cellular populations of either granulosa cells or purified theca-interstitial cells produced negligible increments in glucose consumption and lactate accumulation. However, contact-dependent cocultures of these two cell types (granulosa cells/purified theca-interstitial cells), reconstituted at a putative physiological cellular ratio (4:1), allowed IL-1ß to produce significant (P < 0.05) marked increments in lactate accumulation (4.9-fold), an increase greater than expected from simple additivity. Alterations in glucose consumption followed a similar trend but did not reach statistical significance.



View larger version (25K):
[in this window]
[in a new window]
  		Figure 6. Cell-cell interaction in IL-1ß-induced aerobic glycolysis. Whole ovarian dispersates (5 x 105 cells), purified granulosa (GC; 4 x 105 cells) or purified theca-interstitial (PT; 1 x 105 cells/ml) cells were prepared as described in Materials and Methods and aerobically cultured separately or in mixture (GC/PT) for 96 h in the absence or presence of IL-1ß (50 ng/ml) after which media were analyzed for lactate concentration (upper panel) or glucose consumption (lower panel).

 
Media pyruvate accumulation
The conversion of pyruvate to lactate by lactate dehydrogenase is the final metabolic step in glycolysis. Because pyruvate is the putative obligatory energy source for the developing denuded oocyte (26, 27, 28, 29), we examined the effect of treatment with IL-1ß on pyruvate accumulation in media conditioned by whole ovarian dispersates (Fig. 7Go). Media pyruvate content was considerably lower (two orders of magnitude) than lactate content in control cultures. Treatment with IL-1ß resulted in an 81% reduction in media pyruvate accumulation, an effect reversed by the concurrent application of IL-1RA. The ability of IL-1ß to inhibit pyruvate accumulation was time, dose, and cell density dependent (not shown). These data suggest that IL-1ß is capable of modulating the pyruvate content of conditioned media in a direction opposite to that observed for lactate and that this effect is also receptor-mediated. The decline in media pyruvate levels could not be ascribed to pyruvate instability because the content of pyruvate added to unconditioned (no cell) media was unchanged after a 96-h incubation period.



View larger version (38K):
[in this window]
[in a new window]
  		Figure 7. IL-1ß-mediated pyruvate depletion. Whole ovarian dispersates (5 x 105 viable cells/dish) from immature rats were aerobically cultured for 96 h in the absence or presence of a maximally stimulating dose of IL-1ß (50 ng/ml), with and without IL-1RA (5 µg/ml) after which media pyruvate concentrations were determined.

 
The effect of treatment with IL-1ß on glucose uptake by cultured whole ovarian dispersates
Glucose transport capacity constitutes a major regulatory step in the ability of tissues to increase their glucose utilization. To assess whether the IL-1ß-mediated increase in glucose consumption reflects an increase in glucose uptake, whole ovarian dispersates were incubated for 48 h in the absence or presence of a maximal stimulatory dose of IL-1ß. At the conclusion of this incubation period, cellular glucose uptake capacity was determined over a 15-min period. As shown (Fig. 8Go, left panel), the rate of specific [3H]2-deoxy-D-glucose uptake by control cells was linear throughout. Treatment with IL-1ß markedly increased the rate of glucose uptake, demonstrating an initial linear response for the first few min, and a progressive increase in [3H]2-deoxy-D-glucose uptake towards an apparent plateau. These findings indicate that the IL-1ß-induced increase in glucose consumption was attributable, in part, to an increase in glucose uptake.



View larger version (15K):
[in this window]
[in a new window]
  		Figure 8. IL-1ß-stimulated glucose uptake. Whole ovarian dispersates (5 x 105 viable cells/dish) were cultured for 48 h in the absence or presence of IL-1ß (50 ng/ml). At the conclusion of this incubation period, cellular glucose transport was determined as described under Materials and Methods. Left panel, Glucose uptake as a function of time (a representative experiment from two independent experiments is shown); right panel, the rate (V) of cellular glucose transport was determined using different concentrations (0.5–5.0 mM) of the [1,2-3H]2-deoxy-D-glucose substrate (S). The Eadie-Hofstee transformation of the data (right panel) represents one experiment from three similar, independent experiments.

 
The rate of cellular glucose uptake was determined with varying concentrations (0.5–5.0 mM) of the [3H]2-deoxy-D-glucose substrate. Eadie-Hofstee transformation of the data (Fig. 8Go, right panel) disclosed a modest elevation in Km (1.7-fold) and an increase in Vmax of 3.6-fold in IL-1ß-treated as compared with control cells. Although the variance for Vmax was high for both groups (mean ± SEM for controls = 6.5 ± 2.0 nmol/mg protein/min and for IL-1ß-treated cells = 22.6 ± 6.7 nmol/mg protein/min), the Vmax was elevated in the IL-1ß-treated group in each experiment. The mean ratio of IL-1ß/control was significantly higher than zero (P < 0.05) for both the Km and the Vmax. The increase in Vmax suggests an increase in the number of transporter proteins in IL-1ß-treated cells. The change in the apparent Km calculated in these experiments should be viewed with caution since this result may reflect, in part, the dissociation constant of 2-deoxy-D-glucose for hexokinase, rather than for the glucose transporters (44). Preliminary experiments using [3H]3–0-methyl glucose, a nonphosphorylatable substrate, also disclosed elevated Vmax values for IL-1ß-treated cells as compared with controls (not shown).


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Treatment with IL-1ß induces a significant increase in ovarian glucose consumption (and uptake) as well as in media lactate accumulation. Because the relative accumulation of lactate can be taken as a crude reflection of the intensity of the glycolytic process, the above constellation indicates a shift in cellular metabolism toward the glycolytic processing of glucose despite ample oxygen supply. The ovarian shift toward aerobic glycolysis mimics a similar shift that has long been noted in proliferating and tumor cells (45, 46).

Characterization of the IL-1ß metabolic effect revealed it to be receptor mediated (Fig. 3Go) and dose (Fig. 1Go), time (Fig. 4Go)- and cell density (Fig. 4Go; inset) dependent but glucose concentration-independent (Fig. 3Go). The IL-1ß action was relatively specific in that other ovarian effectors, TNF{alpha} (35), and insulin (34) were incapable of producing a similar effect (Fig. 2Go). Of particular note is the lack of effect by insulin on glucose consumption in the ovary. These data suggest that ovarian glucose transporters do not include the insulin-coupled variety (glucose transporter 4).

Importantly, the IL-1ß-induced increase in glucose consumption was associated with an increase in glucose uptake (Fig. 8Go). This phenomenon could be attributable to an increase in the number of glucose transporter proteins in IL-1ß-treated ovarian cells. Such a possibility is supported by the observation that IL-1ß up-regulates the steady-state levels of transcripts and of proteins corresponding to several glucose transporters in a number of extraovarian cell types (47, 48).

The ability of IL-1ß to bring about a metabolic shift in ovarian cells was time dependent but not immediate (Fig. 4Go). Neither glucose consumption nor lactate generation could be detected up to 24 h in culture even in the presence of IL-1ß. The promotion of glucose uptake and the induction of the glycolytic state may require an as-yet-to-be defined set of transcriptional/translational events without that the cellular reprogramming may not be realized. For example, the modulation of distinct species of glucose transporter(s), monocarboxylate transporters, glutamate transporter(s), and/or of select glycolytic and oxidative enzymes may take place. Studies are currently underway to examine these possibilities.

Our observation that the IL-1ß-induced metabolic shift in ovarian glucose utilization is reversed by IL-1RA (Fig. 3Go) is an indication that this effect is receptor-mediated. In addition, treatment with IL-1RA attenuated the metabolic actions of FSH and hCG (Fig. 5Go), suggesting the existence of endogenously produced, IL-1-like bioactivity. Given the ability of LH/hCG to increase the steady state levels of intraovarian IL-1ß transcripts in the preovulatory rat ovary (6), it is conceivable that the in vivo ability of the midcycle gonadotropin surge to effect a metabolic shift may be due, in part, to the preovulatory induction of intraovarian IL-1ß.

Our findings (Fig. 6Go) suggest that the full magnitude of the metabolic shift requires both cellular components of the follicle (i.e. the granulosa and theca-interstitial cells). These observations are in keeping with previous reports that the ovarian action of IL-1ß is contingent on contact-independent cell-cell cooperation (11, 12, 13).

A metabolic shift of the nature we describe in this study may be the result of multiple regulatory effects on several metabolic pathways. Indeed, significant differences were noted in the molar ratio of lactate produced to glucose consumed when comparing IL-1ß-replete and IL-1ß-deplete circumstances. In the presence of maximally stimulating levels of IL-1ß, 2.5 ± 0.2 lactate molecules were produced per glucose molecule consumed. In an IL-1RA-induced IL-1ß vacuum, this ratio was only 0.9 ± 0.4. These observations suggest that in the presence of IL-1ß, every available glucose molecule is converted to lactate (expected ratio of 2). That the ratio is even higher (2.5 ± 0.2) suggests, consistent with previous reports (16, 43), that lactate-generating sources other than glucose may have been used in IL-1ß-treated cultures. The exact nature and relative contribution of such accessory precursors (e.g. glutamine, alanine, glycogen, or other substrates) is of interest and is currently under active investigation in our laboratory.

Our data further demonstrate an IL-1ß-mediated decrease in media pyruvate accumulation (Fig. 7Go), concordant with the IL-1ß-mediated increase in lactate accumulation. This decline in pyruvate most likely reflects a change in the redox state of the cell, resulting from IL-1ß-stimulated glycolysis. Indeed, the glycolytically induced depletion of NAD would be expected to drive the conversion of pyruvate to lactate to generate additional NAD to maintain the glycolytic process. However, pyruvate depletion could also be related to the activation and/or utilization of other metabolic pathways. For example, IL-1ß-treated cells may increase the uptake of pyruvate into the tricarboxylic acid cycle via pyruvate carboxylase, its conversion to alanine, or its utilization as a metabolic scavenger of oxygen-free radicals. In vivo, the requisite utilization of pyruvate by the glycolytically incompetent oocyte may drive the depletion of pyruvate from the glycolytically competent somatic cells.

The mechanism by which IL-1ß provokes the transition to glycolysis remains to be determined. It may be that the IL-1ß-stimulated increase in glucose uptake (Fig. 8Go) triggers the glucose-mediated enhancement of aerobic glycolysis as described by Crabtree (49). Whether IL-1ß also exerts direct stimulatory effects on glycolytic enzymes is a subject for future investigation.

In summary, these observations lend support to the notion that IL-1ß serves as a mediator of the gonadotropin-associated midcycle shift in ovarian carbohydrate metabolism. The rationale for such a shift remains a mystery. It could be argued that the conversion of the somatic ovarian cell into a glycolytic machinery is necessary to meet the energy needs of the maturing but glycolytically inactive (26, 27, 28, 29) oocyte, which depends on extracellular (cumulus cell-derived) sustenance (50, 51, 52, 53, 54) in general and on monocarboxylate glycolytic products in particular (26, 27, 28, 29). Conceivably, the relatively hypoxic oocyte environment (25) may require a sparing of the utilization of oxygen by the tricarboxylic acid cycle thereby conserving oxygen for other vital tasks such as steroidogenesis. Leese et al. (43) have suggested that the early shift toward aerobic glycolysis may prepare embryos for the relatively anoxic environment of implantation. Alternatively, metabolic antioxidants, such as pyruvate (55), may protect the oocyte from oxygen-free radicals, which may be generated during the dynamic periovulatory period. Undoubtedly, other possibilities exist. In any case, these studies, for the first time, implicate IL-1ß as a key player in modulating the energy needs of the cycling ovary, a topic first addressed in 1962 (14).


   Acknowledgments
 
The authors wish to thank Ms. Cornelia T. Szmajda for her assistance in the preparation of this manuscript and Ms. Susan Lautar for able technical assistance. We also thank Dr. Ian Simpson (NIH, MD), Dr. Steven Downs (Marquette University, Milwaukee, WI), Dr. Jerome F. Strauss (University of Pennsylvania, Philadelphia, PA), and Dr. Karl Brand (University of Erlangen-Nurenberg, Erlangen, Germany) for valuable advice and discussions.


   Footnotes
 
1 This work was supported in part by NIH Grants HD-19998 and HD-00697 (to E.Y.A.), HD-16596 (to L.R.), the Frank C. Bressler Research Award from the University of Maryland (to D.W.P.), as well as by the Lalor Foundation, Fulbright, and Harlea Charitable Trust Fellowship Awards (to I.B.-S.). Back

2 Current address: Department of Obstetrics and Gynecology, Haemek Medical Center, Afula, Israel. Back

3 Current address: Department of Obstetrics and Gynecology, Rambam Medical Center, Haifa, Israel. Back

4 Current address: Department of Obstetrics and Gynecology, Hadassa University Hospital, Mount Scopus, Jerusalem, Israel. Back

5 Current address: The University of Utah, Department of Obstetrics and Gynecology, Health Sciences Center, Suite 2B200, 50 North Medical Drive, Salt Lake City, Utah 84132. Back

Received November 4, 1996.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Ben-Shlomo I, Adashi EY 1994 Interleukin-1 as a mediator in the ovulatory sequence: evidence for a meaningful role of cytokines in ovarian physiology. Curr Opin Endocrinol Diabetes. Current Science Series, pp 187–192
  2. Brännström M, Wang L, Norman RJ 1993 Ovulatory effect of interleukin-1ß on the perfused rat ovary. Endocrinology 132:399–404[Abstract/Free Full Text]
  3. Takehara Y, Dharmarajan AM, Kaufman G, Wallach EE 1994 Effect of interleukin-1ß on ovulation in the in vitro perfused rabbit ovary. Endocrinology 134:1788–1793[Abstract/Free Full Text]
  4. Peterson CM, Hales HA, Hatasaka HH, Mitchell MD, Rittenhouse L, Jones KP 1993 Interleukin-1ß (IL-1ß) modulates prostaglandin production and the natural IL-1 receptor antagonist inhibits ovulation in the optimally stimulated rat ovarian perfusion model. Endocrinology 133:2301–2306[Abstract/Free Full Text]
  5. Simón C, Tsafriri A, Chun S-Y, Piquette GN, Dang W, Polan ML 1994 Interleukin-1 receptor antagonist suppresses hCG-induced ovulation in the rat. Biol Reprod 51:662–667[Abstract]
  6. Hurwitz A, Ricciarelli E, Botero L, Rohan RM, Hernandez ER, Adashi EY 1991 Endocrine- and autocrine-mediated regulation of rat ovarian (theca-interstitial) interleukin-1ß gene expression: gonadotropin- dependent preovulatory acquisition. Endocrinology 129:3427–3429[Abstract/Free Full Text]
  7. Scherzer WJ, Ruutiainen-Altman KSL, Putowski LT, Kol S, Adashi EY, Rohan RM 1996 Detection and in vivo hormonal regulation of rat ovarian type I and type II interleukin-1 receptor mRNAs: increased expression during the periovulatory period. J Soc Gynecol Invest 3:131–140[CrossRef][Medline]
  8. Simón C, Frances A, Piquette G, Polan ML 1994 Immunohistochemical localization of the interleukin-1 system in the mouse ovary during follicular growth, ovulation, and luteinization. Biol Reprod 50:449–457[Abstract]
  9. Brännström M, Norman RJ, Seamark RF, Robertson SA 1994 Rat ovary produces cytokines during ovulation. Biol Reprod 50:88–94[Abstract]
  10. Brännström M, Wang L, Norman RG 1993 Effects of cytokines on prostaglandin production and steroidogenesis of incubated preovulatory follicles of the rat. Biol Reprod 48:165–171[Abstract]
  11. Kokia E, Hurwitz A, Ben-Shlomo I, Adashi EY, Yanagishita M 1993 Receptor mediated stimulatory effects of IL-1ß on hyaluronic acid and proteoglycan biosynthesis by cultured rat ovarian cells: role for heterologous cell-cell interactions. Endocrinology 133:2391–2394[Abstract/Free Full Text]
  12. Hurwitz A, Dushnik M, Solomon H, Ben-Shitrit A, Finci-Yeheskel Z, Milwidsky A, Meyer M, Adashi EY, Yagel S 1993 Cytokine-mediated regulation of rat ovarian function: interleukin-1 stimulates the accumulation of 92-kDa gelatinase. Endocrinology 132:2709–2714[Abstract/Free Full Text]
  13. Ben-Shlomo I, Kokia E, Adashi EY, Payne DW 1994 Interleukin-1ß stimulates nitrite production in the rat ovary: evidence for heterologous cell-cell interaction and for insulin-mediated regulation of the inducible isoform of nitric oxide synthase. Biol Reprod 51:310–318[Abstract]
  14. Armstrong DT, Greep RO 1962 Effect of gonadotrophic hormones on glucose metabolism by luteinized rat ovaries. Endocrinology 70:701–710
  15. Armstrong DT 1963 Stimulation of glycolytic activity of rat corpus luteum tissue by luteinizing hormone. Endocrinology 72:908–913[Abstract/Free Full Text]
  16. Armstrong DT, Kilpatrick R, Greep RO 1963 In vitro and in vivo stimulation of glycolysis in prepubertal rat ovary by luteinizing hormone. Endocrinology 73:165–169
  17. Channing CP, Villee CA 1966 Luteinizing hormone: Effects on uptake and metabolism of hexoses by luteinized rat ovaries. Biochim Biophys Acta 115:205–218[Medline]
  18. Hamberger LA, Ahren KEB 1967 Effects of gonadotropins in vitro on glucose uptake and lactic acid production of ovaries from prepubertal and hypophysectomized rats. Endocrinology 81:93–100[Abstract/Free Full Text]
  19. Flint APF, Denton RM 1969 Glucose metabolism in the superovulated rat ovary in vitro. Biochem J 112:243–254[Medline]
  20. Nilsson L 1974 Acute effects of gonadotropins and prostaglandins of the metabolism of isolated ovarian follicles from PMSG-treated immature rats. Acta Endocrinol 77:540–558
  21. Tsafriri A, Lieberman ME, Ahren KEB, Lindner HR 1976 Dissociation between LH-induced aerobic glycolysis and oocyte maturation in cultured graafian follicles of the rat. Acta Endocrinol 81:362–366
  22. Billig H, Hedin L, Magnusson C 1983 Gonadotropins stimulate lactate production by rat cumulus and granulosa cells. Acta Endocrinol 103:562–566
  23. Harlow CR, Winston RML, Margara RA, Hillier SG 1987 Gonadotropic control of human granulosa cell glycolysis. Human Reprod 2:649–653[Abstract/Free Full Text]
  24. Boland NI, Humpherson PG, Leese HJ, Gosden RG 1993 Pattern of lactate production and steroidogenesis during growth and maturation of mouse ovarian follicles in vitro. Biol Reprod 48:798–806[Abstract]
  25. Gosden RG, Byatt-Smith JT 1986 Oxygen concentration gradient across the ovarian follicular epithelium: model, predictions and implications. Human Reprod 1:65–68[Abstract/Free Full Text]
  26. Biggers JD, Whittingham DG, Donahue RP 1967 The pattern of energy metabolism in the mouse oocyte and zygote. Proc Natl Acad Sci USA 58:560[Free Full Text]
  27. Leese HJ, Barton AM 1984 Pyruvate and glucose uptake by mouse ova and preimplantation embryos. J Reprod Fertil 72:9–13[Abstract/Free Full Text]
  28. Tsutsumi O, Satoh K, Taketani Y, Kato T 1992 Determination of enzyme activities of energy metabolism in the maturing rat oocyte. Mol Reprod Develop 33:333–337[CrossRef][Medline]
  29. Reiger D, Loskutoff NM 1994 Changes in the metabolism of glucose, pyruvate, glutamine and glycine during maturation of cattle oocytes in vitro. J Reprod Fertil Steril 100:257–262[Abstract/Free Full Text]
  30. Zuelke KA, Brackett B 1992 Effects of luteinizing hormone on glucose metabolism in cumulus-enclosed bovine oocytes mature in vitro. Endocrinology 131:2690–2696[Abstract/Free Full Text]
  31. Taylor DJ 1990 Interleukin-1 stimulation of fibroblast glycolysis is accompanied by reduced glucose oxidation in the tricarboxylic acid cycle. Biochem Soc Trans 18:982–983[Medline]
  32. Mandrup-Poulsen T, Helqvist S, Molvig J, Wogensen LD, Nerup J 1989 Cytokines as immune effector molecules in autoimmune endocrine disease with special reference to insulin-dependent diabetes mellitus. Autoimmunity 4:191–218[Medline]
  33. Welsh N, Eizirik D, Bendtzen K, Sandler S 1991 Interleukin-1ß-induced nitric oxide production in isolated rat pancreatic islets requires gene transcription and may lead to inhibition of the krebs cycle enzyme aconitase. Endocrinology 129:2167–3173
  34. Hernandez ER, Resnick CE, Svoboda ME, Van Wyk JJ, Payne DW, Adashi EY 1988 Insulin as a regulator of and androgen biosynthesis by cultured rat ovarian cells: Cellular Mechanism(s) underlying physiological and pharmacological hormonal actions. Endocrinology 122:2034–2044[Abstract/Free Full Text]
  35. Adashi EY, Resnick CE, Croft CS, Payne DW 1989 Tumor necrosis factor {alpha} inhibits the differentiation of non-transformed ovarian granulosa cells: A cytodifferentiative non-cytotoxic property. J Biol Chem 264:11591–11597[Abstract/Free Full Text]
  36. Magoffin DA, Erickson GF 1982 Primary culture of differentiating ovarian androgen-producing cells in defined medium. J Biol Chem 257:4507–4513[Free Full Text]
  37. Gloster JA, Harris P 1962 Observations on an enzymatic method for the estimation of pyruvate in blood. Clin Chim Acta 7:206[CrossRef][Medline]
  38. Raabo E, Terkildsen TC 1960 On the enzymatic determination of blood glucose. Scand J Clin Invest 12:402[Medline]
  39. Bondar RJL, Mead DC 1974 Evaluation of glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides in the hexokinase method for determining glucose in serum. Clin Chem 20:586[Abstract]
  40. Garcia KG, Goldstein JL, Pathak RK, Anderson RGW, Brown MS 1994 Molecular characterization of a membrane transporter for lactate, pyruvate, and other monocarboxylates: Implications for the Cori Cycle. Cell 76:865–873[CrossRef][Medline]
  41. Kaku K, Fiedorek FT, Jr, Province M, Permutt MA 1988 Genetic analysis of glucose tolerance in inbred mouse strains. Diabetes 37:707–713[Abstract]
  42. Kokia E, Ben-Shlomo I, Adashi EY 1994 The ovarian action of IL-1 is receptor-mediated: reversal by a naturally occurring interleukin-1 receptor antagonist. Fertil Steril 63:176–181
  43. Leese HJ, Conaghan J, Martin KL, Hardy K 1993 Early human embryo metabolism. BioEssays 15:259–264[CrossRef][Medline]
  44. Maher F, Davies-Hill TM, Simpson I 1996 Substrate specificity and kinetic parameters of GLUT 3 in rat cerebellar granule neurons. Biochem J 315:827–831
  45. Greiner EF, Guppy M, Brand K 1994 Glucose is essential for proliferation and the glycolytic enzyme induction that provokes a transition to glycolytic energy production. J Biol Chem 269:31484–31490[Abstract/Free Full Text]
  46. Warburg O, Posener K, Negelein E 1924 Uber den stoffwechel der carcinomzelle. Biochem Z 152:309–344
  47. Bird TA, Davies A, Baldwin SA, Saklatvala J 1990 Interleukin-1 stimulates hexose transport in fibroblasts by increasing the expression of glucose transporters. J Biol Chem 265:13578–13583[Abstract/Free Full Text]
  48. Garcia-Welsh A, Schneiderman JS, Baly DL 1990 Interleukin-1 stimulates glucose transport in rat adipose cells: Evidence for receptor discrimination between IL-1ß and IL-1{alpha}. FEBS Lett 2:421–424
  49. Crabtree HD 1929 Observations on carbohydrate metabolism of tumours. Biochem J 23:536–545
  50. Donahue RP, Sterm S 1968 Follicular cell support of oocyte maturation: production of pyruvate in vitro. J Reprod Fertil 17:395–398[Abstract/Free Full Text]
  51. Brinster RI 1971 Oxidation of pyruvate and glucose by oocytes of the mouse and rhesus monkey. J Reprod Fertil 24:187–191[Abstract/Free Full Text]
  52. Eppig JJ 1976 Analysis of mouse oogenesis in vitro. Oocyte isolation and the utilization of exogenous energy sources by growing oocytes. J Exp Zool 198:375–382[CrossRef][Medline]
  53. Heller DT, Cahill DM, Schultz RM 1981 Biochemical studies of mammalian oogenesis: metabolic cooperativity between granulosa cells and growing mouse oocytes. Dev Biol 84:455–464[CrossRef]
  54. Downs SM, Mastropolo AM 1994 The participation of energy substrates in the control of meiotic maturation in murine oocytes. Dev Biol 162:154–168[CrossRef][Medline]
  55. DeBoer LW, Bekx PA, Han L, Steinke L 1993 Pyruvate enhances recovery of rat hearts after ischemia and reperfusion by preventing free radical generation. Am J Physiol 265:H1571–H1576



This article has been cited by other articles:


Home page
 IOVSHome page
S. F. Abcouwer, S. Shanmugam, P. F. Gomez, S. Shushanov, A. J. Barber, K. F. Lanoue, P. G. Quinn, M. Kester, and T. W. Gardner
Effect of IL-1{beta} on Survival and Energy Metabolism of R28 and RGC-5 Retinal Neurons
Invest. Ophthalmol. Vis. Sci., December 1, 2008; 49(12): 5581 - 5592.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Cell Physiol.Home page
N. Taneja, P. E. Coy, I. Lee, J. M. Bryson, and R. B. Robey
Proinflammatory interleukin-1 cytokines increase mesangial cell hexokinase activity and hexokinase II isoform abundance
Am J Physiol Cell Physiol, August 1, 2004; 287(2): C548 - C557.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ben-Shlomo, I.
Right arrow Articles by Adashi, E. Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ben-Shlomo, I.
Right arrow Articles by Adashi, E. Y.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals