The ovariectomized, mature rat model of postmenopausal osteoporosis: an assessment of the bone sparing effects of curcumin.

Identification of natural health products that might benefit skeletal health could reduce the negative impact of osteoporotic bone fractures upon society. The objectives of this study were to evaluate an animal model of postmenopausal osteoporosis and to search for evidence that curcumin reduces bone mineral losses in a dose-dependent manner when endogenous estrogen levels are reduced. Bone mineral density was measured at the spine, femur and whole body before and at 2, 4 and 6 months after ovariectomy in each of 40 mature rats. Serum osteocalcin and C-telopeptide were measured as indicators of bone formation and resorption rates. Femoral compressive strength was measured at 6 months. Ovariectomy alone resulted in loss of mineral from the spine (p<0.005) and an increase in osteocalcin levels (p<0.05). At the same time, there was an increase in energy to fracture (p<0.01) due to an increased bone size. When ovariectomized animals were given etidronate there was no loss of mineral from the spine, the size of the femur increased (p<0.005), C-telopeptide levels were reduced (p<0.001) and femoral compressive strength increased (p<0.025). Administration of curcumin to ovariectomized animals resulted in changes that were intermediate between those produced by etidronate and by ovariectomy alone. The increase in femur size produced by the highest dose of curcumin was statistically significant (p< 0.01) and curcumin administration resulted in a significant, dose dependent, increase in energy to fracture. Curcumin produces beneficial changes in bone turnover and increases in bone strength using the ovariectomized mature rat model of postmenopausal osteoporosis.

CC chemokine I-309 is the principal monocyte chemoattractant induced by apolipoprotein(a) in human vascular endothelial cells.

BACKGROUND: Lipoprotein(a) [Lp(a)] is a risk factor for atherosclerosis; however, the mechanisms are unclear. We previously reported that Lp(a) stimulated human vascular endothelial cells to produce monocyte chemotactic activity. The apolipoprotein(a) [apo(a)] portion of Lp(a) was the active moiety. METHODS AND RESULTS: We now describe the identification of the chemotactic activity as being due to the CC chemokine I-309. The carboxy-terminal domain of apo(a) containing 6 type-4 kringles (types 5 to 10), kringle V, and the protease domain was demonstrated to contain the I-309-inducing portion. Polyclonal and monoclonal anti-I-309 antibodies as well as an antibody against a portion of the extracellular domain of CCR8, the I-309 receptor, inhibited the increase in monocyte chemotactic activity induced by apo(a). I-309 antisense oligonucleotides also inhibited the induction of endothelial monocyte chemotactic activity by apo(a). I-309 mRNA was identified in human umbilical vein endothelial cells. Apo(a) induced an increase in I-309 protein in the endothelial cytoplasm and in the conditioned medium. Immunohistochemical studies have identified I-309 in endothelium, macrophages, and extracellular areas of human atherosclerotic plaques and have found that I-309 colocalized with apo(a). CONCLUSIONS: These data establish that I-309 is responsible for the monocyte chemotactic activity induced in human umbilical vein endothelial cells by Lp(a). The identification of the endothelial cell as a source for I-309 suggests that this chemokine may participate in vessel wall biology. Our data also suggest that I-309 may play a role in mediating the effects of Lp(a) in atherosclerosis.

Megakaryocyte-targeted synthesis of the integrin beta(3)-subunit results in the phenotypic correction of Glanzmann thrombasthenia.

Glanzmann thrombasthenia is an inherited bleeding disorder characterized by qualitative or quantitative defects of the platelet-specific integrin, alphaIIbbeta(3). As a result, alphaIIbbeta(3) cannot be activated and cannot bind to fibrinogen, leading to a loss of platelet aggregation. Thrombasthenia is clinically characterized by mucocutaneous hemorrhage with episodes of intracranial and gastrointestinal bleeding. To develop methods for gene therapy of Glanzmann thrombasthenia, a murine leukemia virus (MuLV)-derived vector, -889Pl(A2)beta(3), was transduced into peripheral blood CD34(+) cells from 2 patients with thrombasthenia with defects in the beta(3) gene. The human alphaIIb promoter was used in this vector to drive megakaryocyte-targeted expression of the wild-type beta(3) subunit. Proviral DNA and alphaIIbbeta(3) biosynthesis were detected after in vitro differentiation of transduced thrombasthenic CD34(+) cells with megakaryocyte growth and development factor. Flow cytometric analysis of transduced patient samples indicated that 19% of megakaryocyte progeny expressed alphaIIbbeta(3) on the surface at 34% of normal receptor levels. Treatment of transduced megakaryocytes with a combination of agonists including epinephrine and the thrombin receptor-activating peptide induced the alphaIIbbeta(3) complex to form an activated conformation capable of binding fibrinogen as measured by PAC-1 antibody binding. Transduced cells retracted a fibrin clot in vitro similar to megakaryocytes derived from a normal nonthrombasthenic individual. These results demonstrate ex vivo phenotypic correction of Glanzmann thrombasthenia and support the potential use of hematopoietic CD34(+) cells as targets for alphaIIb promoter-driven MuLV vectors for gene therapy of platelet disorders. (Blood. 2000;95:3645-3651)

A naturally occurring mutation near the amino terminus of alphaIIb defines a new region involved in ligand binding to alphaIIbbeta3.

Decreased expression of functional alphaIIbbeta3 complexes on the platelet surface produces Glanzmann thrombasthenia. We have identified mutations of alphaIIb(P145) in 3 ethnically distinct families affected by Glanzmann thrombasthenia. Affected Mennonite and Dutch patients were homozygous and doubly heterozygous, respectively, for a P(145)A substitution, whereas a Chinese patient was doubly heterozygous for a P(145)L substitution. The mutations affect expression levels of surface alphaIIbbeta3 receptors on their platelets, which was confirmed by co-transfection of alphaIIb(P145A) and beta3 cDNA constructs in COS-1 cells. Each mutation also impaired the ability of alphaIIbbeta3 on affected platelets to interact with ligands. Moreover, when alphaIIb(P145A) and beta3 were stably coexpressed in Chinese hamster ovary cells, alphaIIbbeta3 was readily detected on the cell surface, but the cells were unable to adhere to immobilized fibrinogen or to bind soluble fluorescein isothiocyanate-fibrinogen after alphaIIbbeta3 activation by the activating monoclonal antibody PT25-2. Nonetheless, incubating affected platelets with the peptide LSARLAF, which binds to alphaIIb, induced PF4 secretion, indicating that the mutant alphaIIbbeta3 retained the ability to mediate outside-in signaling. These studies indicate that mutations involving alphaIIb(P145 )impair surface expression of alphaIIbbeta3 and that the alphaIIb(P145A) mutation abrogates ligand binding to the activated integrin. A comparative analysis of other alphaIIb mutations with a similar phenotype suggests that these mutations may cluster into a single region on the surface of the alphaIIb and may define a domain influencing ligand binding. (Blood. 2000;95:180188)

Activation of proMMP-9 by a plasmin/MMP-3 cascade in a tumor cell model. Regulation by tissue inhibitors of metalloproteinases.

To examine MMP-9 activation in a cellular setting we employed cultures of human tumor cells that were induced to produce MMP-9 over a 200-fold concentration range (0.03 to 8.1 nM). The secreted levels of TIMPs in all the induced cultures remain relatively constant at 1-4 nM. Quantitation of the zymogen/active enzyme status of MMP-9 in the cultures indicates that even in the presence of potential activators, the molar ratio of endogenous MMP-9 to TIMP dictates whether proMMP-9 activation can progress. When the MMP-9/TIMP ratio exceeds 1.0, MMP-9 activation progresses, but only via an interacting protease cascade involving plasmin and stromelysin 1 (MMP-3). Plasmin, generated by the endogenous plasminogen activator (uPA), is not an efficient activator of proMMP-9. Plasmin, however, is very efficient at generating active MMP-3 from exogenously added proMMP-3. The activated MMP-3, when its concentration exceeds that to TIMP, becomes a potent activator of proMMP-9. Addition to the cultures of already-activated MMP-3 relinquishes the requirement for plasminogen and proMMP-3 additions and results in direct activation of the endogenous proMMP-9. The activated MMP-9 enhances the invasive phenotype of the cultured cells as their ability to transverse basement membrane is significantly increased following zymogen activation. That this enhanced tissue remodeling capability is due to the activation of MMP-9 is demonstrated through the use of a specific anti-MMP-9-blocking monoclonal antibody.

Activation of matrix metalloproteinase-9 (MMP-9) via a converging plasmin/stromelysin-1 cascade enhances tumor cell invasion.

Matrix metalloproteinase-9 (MMP-9) may play a critical catalytic role in tissue remodeling in vivo, but it is secreted by cells as a stable, inactive zymogen, pro-MMP-9, and requires activation for catalytic function. A number of proteolytic enzymes activate pro-MMP-9 in vitro, but the natural activator(s) of MMP-9 is unknown. To examine MMP-9 activation in a cellular setting we employed cultures of human tumor cells (MDA-MB-231 breast carcinoma cells) that were induced to produce MMP-9 over a 200-fold concentration range (0.03-8.1 nM). The levels of tissue inhibitors of metalloproteinase (TIMPs) in the induced cultures remain relatively constant at 1-4 nM. Quantitation of the zymogen/active enzyme status of MMP-9 in the MDA-MB-231 cultures indicates that even in the presence of potential activators, the molar ratio of endogenous MMP-9 to TIMP dictates whether pro-MMP-9 activation can progress. When the MMP-9/TIMP ratio exceeds 1.0, MMP-9 activation progresses, but through an interacting protease cascade involving plasmin and stromelysin 1 (MMP-3). Plasmin, generated by the endogenous urokinase-type plasminogen activator, is not an efficient activator of pro-MMP-9, neither the secreted pro-MMP-9 nor the very low levels of pro-MMP-9 associated with intact cells. Although plasmin can proteolytically process pro-MMP-9, this limited action does not yield an enzymatically active MMP-9, nor does it cause the MMP-9 to be more susceptible to activation. Plasmin, however, is very efficient at generating active MMP-3 (stromelysin-1) from exogenously added pro-MMP-3. The activated MMP-3 becomes a potent activator of the 92-kDa pro-MMP-9, yielding an 82-kDa species that is enzymatically active in solution and represents up to 50-75% conversion of the zymogen. The activated MMP-9 enhances the invasive phenotype of the cultured cells as their ability to both degrade extracellular matrix and transverse basement membrane is significantly increased following zymogen activation. That this enhanced tissue remodelling capability is due to the activation of MMP-9 is demonstrated through the use of a specific anti-MMP-9 blocking monoclonal antibody.

Prenatal diagnosis of Glanzmann thrombasthenia using the polymorphic markers BRCA1 and THRA1 on chromosome 17.

Glanzmann thrombasthenia is an autosomal recessive bleeding disorder caused by mutations in the genes encoding platelet GPIIb or GPIIIa. Both genes map to chromosome 17q21 and polymorphisms within this chromosomal region have been identified. In the current study, prenatal diagnosis was performed for a family that already had one affected child, patient 1, who had a compound heterozygous mutation in GPIIb. At the time of prenatal diagnosis, the maternal GPIIb mutation had been identified but the paternal GPIIb mutation was unknown. By sequence analysis, the fetus was identified as a carrier of the mother's mutation. To determine the probability of the fetus inheriting the father's mutation, haplotype analysis of DNA samples from the fetus, mother, father and affected child were performed using polymorphic markers on chromosome 17q12-q21. These markers included polymorphisms within the thyroid hormone receptor alpha1 gene (THRA1), the breast cancer gene (BRCA1), GPIIb, GPIIIa, and an anonymous marker D17S579. Heterozygosity within the THRA1, BRCA1 and GPIIIa polymorphic markers predicted that the fetus carried the father's normal allele. Based on genetic linkage studies, no recombination was identified with any of the informative markers, and from the map distance between GPIIb and BRCA1 the accuracy of diagnosis was predicted to be >98%. The father's mutation was subsequently identified and direct sequence analysis of fetal DNA confirmed that the fetus did not inherit the fathers' mutant allele.

Glycoprotein IIb Leu214Pro mutation produces glanzmann thrombasthenia with both quantitative and qualitative abnormalities in GPIIb/IIIa.

Glanzmann thrombasthenia is an inherited bleeding disorder due to a functional reduction or absence of platelet GPIIb/IIIa (alphaIIbbeta3) integrin receptors. Based on a prolonged bleeding time and absence of platelet aggregation in response to physiologic agonists, a 55-year-old white man was diagnosed as having Glanzmann thrombasthenia. The patient's platelet fibrinogen level was approximately 5% of normal. As judged by complex-dependent monoclonal antibody (MoAb) binding, surface expression of platelet GPIIb/IIIa receptors was less than 5.5% of normal, whereas the binding of an anti-GPIIIa specific MoAb (7H2) was approximately 12% of normal. Immunoblot analysis of the patient's platelet lysates showed approximately 35% of normal levels of GPIIIa, approximately 30% of normal levels of GPIIb, and an abnormally migrating fragment of GPIIb. Biotinylation of the surface proteins on the patient's platelets followed by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed only GPIIb and GPIIIa subunits of normal size. Surface expression of platelet alphavbeta3 receptors was 192% of normal, suggesting that the patient's' defect was in GPIIb. Sequence analysis of the patient's GPIIb cDNA identified a T to C transition at nucleotide 643, predicting a Leu214Pro substitution. Direct sequencing of GPIIb exon 6 indicated that the patient is homozygous for the mutation. The nature of the Leu214Pro mutation was analyzed by expression in Chinese hamster ovary (CHO) cells. As judged by subunit-specific MoAb binding, surface expression of mutant receptors was approximately 60% of normal, but these receptors were not recognized by the complex-dependent monoclonal antibodies, 10E5 and 7E3. In addition, mutant receptors pretreated with the ligand-induced binding site MoAb AP5 were not recognized by the activation-dependent MoAb PAC-1 and mutant expressing CHO cells did not adhere to immobilized fibrinogen. These data suggest that the Leu214Pro mutation in GPIIb disrupts the structural conformation, and either directly or indirectly, the ligand binding properties of the heterodimeric complex. This is in accord with studies from other integrins that have implicated a beta-turn in a homologous region as important in ligand binding. Thus, the Leu214Pro mutation appears to produce the Glanzmann thrombasthenia phenotype by both qualitative and quantitative abnormalities. In addition, the mutation appears to confer susceptibility of the GPIIb subunit to proteolysis.

Distinct STAT structure promotes interaction of STAT2 with the p48 subunit of the interferon-alpha-stimulated transcription factor ISGF3.

Cells express a variety of STAT (signal transducer and activator of transcription) transcription factors that are structurally homologous and yet function specifically in response to particular cytokines. The functions of the individual STATs are dependent on distinct protein-protein interactions. STAT1 and STAT2 are activated by tyrosine phosphorylation in response to type I interferons-alpha/beta (IFN-alpha/beta) and subsequently form a multimeric transcription factor designated the IFN-alpha-stimulated gene factor 3 (ISGF3). ISGF3 is a unique STAT complex because it also contains a non-STAT molecule, p48, which is a critical DNA-binding component. We provide evidence that STAT2 specifically interacts with p48 in vivo before and after IFN-alpha stimulation. The specificity of ISGF3 formation is therefore a result of the distinct nature of the STAT2 molecule. Coimmunoprecipitation assays demonstrate p48 association with STAT2 but not STAT1. Hybrid STAT2. STAT1 molecules were used to identify a region of STAT2 which specifically associates with p48. The region of STAT2 interaction spans an amino-terminal region of two predicted coiled coils. The studies demonstrate the in vivo existence of a STAT2.p48 complex and a distinct STAT2.STAT1 complex after IFN-alpha stimulation. Data suggest that distinct bipartite complexes STAT2.p48 and STAT2.STAT1 translocate to the nucleus and associate on the DNA target site as ISGF3.

Elevated levels of 92-kd type IV collagenase (matrix metalloproteinase 9) in giant cell arteritis.

OBJECTIVE: To determine if circulating gelatinase activity and matrix metalloproteinase 9 (MMP-9) (gelatinase B, or 92-kd type IV collagenase) antigenic levels are elevated in sera of patients with giant cell arteritis (GCA), and to ascertain if MMP-9 messenger RNA (mRNA) is deposited in situ at sites of disease involvement. METHODS: Serum samples were collected from 12 patients with GCA and 12 healthy volunteers. Vascular tissue was obtained at the time of temporal artery biopsy. Type IV collagenase activity was determined by gelatin substrate zymography and the quantitative biotinylated gelatin substrate degradation assay. A double-sandwich immunoassay utilizing 2 different isotypes of monoclonal antibodies generated against MMP-9 was used for measuring serum MMP-9 antigenic levels. Finally, to localize sites of MMP-9 mRNA transcription in inflamed arteries, the method of reverse transcriptase in situ polymerase chain reaction (RTisPCR) was utilized. RESULTS: Serum gelatinase activity and MMP-9 titers were significantly increased in patients with GCA (mean +/- SEM 198.9 +/- 36.9 micrograms gelatin/hour/ml serum, versus 21.2 +/- 4.0 in controls; P = 0.0006). The differences in antigenic MMP-9 levels were even more prominent (3005.4 +/- 900.6 ng/ml and 31.6 +/- 9.8 ng/ml in GCA and control sera, respectively; P = 0.007). By RTisPCR, MMP-9 mRNA was mainly detected in cytoplasm of cells resembling smooth muscle cells and fibroblasts in regions of fragmented elastic tissue in the lamina media. CONCLUSION: Gelatinase activity, and specifically MMP-9 levels, are substantially elevated in sera of patients with GCA. Detection of MMP-9 mRNA in the lamina media of inflamed vasculature suggests that degradation of intercellular matrix, particularly elastic fibers, may play a key role in the pathogenesis of GCA. Further studies are needed to determine if the circulating MMP-9 level could be utilized as a clinical marker of disease activity.

A Cys374Tyr homozygous mutation of platelet glycoprotein IIIa (beta 3) in a Chinese patient with Glanzmann's thrombasthenia.

A 20-year-old woman from a consanguineous family in the Hunan Province of the People's Republic of China was diagnosed as having Glanzmann's thrombasthenia based on (1) nearly a lifelong history of epistaxis, gum bleeding, petechiae, and purpura; (2) severe menorrhagia resulting in anemia and need for whole-blood transfusion; (3) normal coagulation assays; (4) prolonged bleeding time; (5) absent clot retraction; (6) decreased glass bead retention; (7) absent platelet aggregation in response to adenine diphosphate, epinephrine, and collagen; and (8) normal initial slope of platelet aggregation in response to ristocetin, but with a diminished maximal extent. The patient's platelets had a decreased level of platelet fibrinogen, but the deficiency was not as severe as in other Glanzmann's thrombasthenia patients. As judged by monoclonal antibody binding studies, surface glycoprotein (GP) IIb/IIIa (alpha IIb beta 3) expression was less than 15% of normal and alpha v beta 3 vitronectin receptor expression was 15% to 19% of normal, suggesting that the defect was in GPIIIa (beta 3). Immunoblotting of platelet lysates demonstrated decreased levels of GPIIb (approximately 30% to 35% of normal) and GPIIIa (approximately 10% of normal), and the GPIIb had undergone normal maturational processing into GPIIb heavy and light chains. Sequence analysis of the patient's GPIIIa RNA identified a G to A mutation at nucleotide 1219, predicting a Cys to Tyr substitution at residue 374. The patient's parents, who are first cousins, are asymptomatic and have only minor reductions in platelet aggregation. Direct sequencing of polymerase chain reaction-amplified cDNA and GPIIIa exon VIII indicated that the patient is homozygous and her parents are heterozygous for the mutation. Transient transfection studies in Chinese hamster ovary cells indicated that the mutation results in an 85% to 90% reduction in GPIIb/IIIa surface expression, but these cells retain the ability to mediate adhesion to immobilized fibrinogen. The relative preservation of platelet fibrinogen despite the very low level of platelet surface GPIIb/IIIa expression in this patient raises some interesting questions regarding the mechanism of fibrinogen uptake and the pathophysiology of Glanzmann's thrombasthenia.

Markedly elevated serum MMP-9 (gelatinase B) levels in rheumatoid arthritis: a potentially useful laboratory marker.

In an attempt to find a potentially useful serum marker in rheumatoid arthritis (RA) which reflects underlying pathogenic mechanisms, we measured the circulating levels of matrix-degrading metalloproteinase-9 (MMP-9), also termed gelatinase B, in sera and synovial fluid (SF) from patients with RA and also quantitated the deposition and local synthesis of MMP-9 in RA synovium. Clinical samples, subjected to gelatin substrate zymography, antigenic immunoassay, and a quantitative substrate degradation assay, revealed elevated 92- and 72-kDa proenzyme forms of MMP-9 and MMP-2 in RA sera and SF compared with healthy controls. Immunostaining on fresh RA synovial specimens revealed MMP-9 within vascular walls in fibroblast-like cells and macrophages; mRNA synthesis was detected using reverse transcriptase in situ PCR. In summary, MMP-9 levels are substantially elevated in the sera and SF from patients with RA. The RA synovium is a source of this MMP-9 production, with abundant mRNA and protein observed within several different type of rheumatoid synovial cells.

Controlled release of substituted benzoic and naphthoic acids using Carbopol gels: measurement of drug concentration profiles and correlation to release rate kinetics.

PURPOSE: The objective of this study is to correlate drug release mechanism with measured drug concentration profiles in gel layers of Carbopol matrices containing mesalamine or benzoic acid. METHODS: Release rate experiments with Carbopol matrices were performed using a rotating disk apparatus. Matrices were frozen and the gel layer in the matrices was sliced using a microtome in a cryostat. Drug concentration profiles were determined by direct measurement of the concentration of the drug in the gel slices. The pH of the slices was measured using microelectrodes, and water content was measured by Karl Fisher titration. RESULTS: The concentration gradient in mesalamine matrices decreased over time and correlated with square root of time release rate kinetics. The concentration profiles of benzoic acid were unchanged over time and correlated with zero order release rate kinetics. Carbopol gel layers were highly hydrated (93-95% water). Gel layers in matrices with mesalamine had a more alkaline microenvironmental pH. This higher pH resulted in increased growth of the thickness of the gel layer and a reduction drug diffusivity in comparison to benzoic acid matrices. CONCLUSIONS: The release rate kinetics of mesalamine and benzoic acid correlated to the measured concentration profiles. The shape of the concentration profiles is determined by the rate of growth of the Carbopol gel layer and drug diffusivity.

Correlation of the in situ detection of polymerase chain reaction-amplified metalloproteinase complementary DNAs and their inhibitors with prognosis in cervical carcinoma.

The purpose of this study was to correlate the presence of matrix metalloproteinase (MMP)-9 and MMP-2 and tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 mRNAs, detected in serial sections using the reverse transcriptase in situ PCR technique, with prognosis in 23 cases of cervical carcinoma. PCR-amplified MMP and TIMP cDNA were restricted to the invasive cancers cells and the surrounding stromal cells. The ratios of cancer and stromal cells expressing MMP-9 and MMP-2 to those expressing TIMP-1 and TIMP-2 were approximately 1 in those cancers with a good prognosis. This MMP:TIMP ratio in the cancer and stromal cells with a poor prognosis was significantly increased to 5.4 and 3.4 (P < 0.0001), respectively, reflecting a marked reduction in the TIMP detection rate in cancers with a poor prognosis. In cervical cancer cell lines SiHa and HeLa, the MMP:TIMP ratio was also close to 1 and, interestingly, these cell lines are invasive but rarely metastatic in nude mice. These data suggest that the balance of MMP-9 and MMP-2 to TIMP-1 and TIMP-2 expression is an essential factor in the aggressiveness of cervical cancer.

Interleukin-1 beta regulates human cytotrophoblast metalloproteinase activity and invasion in vitro.

During early human pregnancy, fetal cytotrophoblasts rapidly invade the uterus. This process has many similarities to tumor invasion, except that the extent and the timing of cytotrophoblast invasion are carefully regulated. Therefore, this system is particularly useful for studying mechanisms that regulate invasive processes. Previously, we showed that production and activation of the 92-kDa type IV collagenase (matrix metalloproteinase(MMP)-9) is necessary for cytotrophoblast invasion in vitro. In other systems, interleukin (IL)-1 beta is an important regulator of matrix-degrading metalloproteinases. Therefore, we investigated trophoblast production of IL-1 beta and its receptors, as well as the effects of this cytokine on cytotrophoblast metalloproteinase activity and invasion. The results showed that release of IL-1 beta parallels the invasive potential of the cytotrophoblasts; the highest levels are produced by first trimester cells and the lowest levels by term cells. Immunoprecipitation showed that cytotrophoblasts express the 80-kDa type I IL-1 receptor, suggesting that autocrine effects are possible. IL-1 beta stimulated trophoblast MMP-9 secretion (by a mechanism that required nascent mRNA and protein synthesis) as well as metalloproteinase activity and invasion of Matrigel. Increasing (by lipopolysaccharide treatment) or decreasing (by glucocorticoid treatment) IL-1 beta production had parallel effects on MMP-9 secretion, metalloproteinase activity, and invasion. Because IL-1 beta and corticosteroids are present in high concentrations at the maternal-fetal interface, normal trophoblast invasion may be regulated, in part, by their opposing actions. In contrast, stimulation of cytotrophoblast IL-1 beta secretion by lipopolysaccharide may play a role in the sequela of infected fetal membranes.