Matrix metalloproteinases are a highly regulated family of enzymes, that together can degrade most components of the extracellular matrix. These proteins are active in normal and pathological processes involving tissue remodeling; however, their sites of synthesis and specific roles are poorly understood. Using in situ hybridization, we determined cellular distributions of matrix metalloproteinases and tissue inhibitor of metalloproteinase-1, an inhibitor of matrix metalloproteinases, in endometrium during the reproductive cycle. The mRNAs for all the metalloproteinases were detected in menstrual endometrium, but with different tissue distributions. The mRNA for matrilysin was localized to epithelium, while the others were detected in stromal cells. Only the transcripts for the 72-kD gelatinase and tissue inhibitor of metalloproteinases-1 were detected throughout the cycle. Transcripts for stromelysin-2 and the 92-kD gelatinase were only detected in late secretory and menstrual endometrium, while those for matrilysin, the 72-kD gelatinase, and stromelysin-3 were also consistently detected in proliferative endometrium. These data indicate that matrix metalloproteinases are expressed in cell-type, tissue, and reproductive cycle-specific patterns, consistent with regulation by steroid hormones, and with specific roles in the complex tissue growth and remodeling processes occurring in the endometrium during the reproductive cycle. (J. Clin. Invest. 1994.94:946-953.) Key words: in situ hybridization * reproductive cycle * matrilysin -stromelysin -RNA

Insulin-like growth factors (IGFs), IGF-binding proteins (IGFBPs), and insulin are believed to be important in the regulation of fetal and neonatal growth. We previously reported that the profiles of IGFBPs in fetal cord serum (FCS) were dependent on the growth/metabolic status of the fetus. The goals of the current study were to examine the IGF system in FCS from term fetuses with normal growth, those with intrauterine growth retardation (IUGR), and those who were large for gestational age (LGA) and in FCS from normal weight preterm (25-37 weeks) and term fetuses in the neonatal period from the day of birth (day 0) until 7 days of age (day 7). Western ligand blotting (WLB) of term FCS revealed IGFBPs with mol wt of 43 and 38 kilodaltons (kDa; IGFBP-3), 34 kDa (IGFBP-2), 28 kDa (IGFBP-1 and glycosylated IGFBP-4), and 24 kDa (IGFBP-4). In IUGR FCS, there was a 50% decrease in IGFBP-3 detected by WLB, which was shown not to be due to an IGFBP-3 protease in IUGR sera. In LGA FCS, IGFBP-3 levels were elevated 2-fold by densitometric analysis of ligand blots. In normal term FCS, the following levels (+/- SE) were present: IGF-I, 76 +/- 16; IGF-II, 401 +/- 38; IGFBP-3, 700 +/- 112; IGFBP-1, 77 +/- 10 ng/mL; and insulin, 3.8 +/- 1.6 microU/mL. In IUGR FCS, IGF-I, IGF-II, and IGFBP-3 were significantly reduced, and IGFBP-1 was 7-fold higher than in FCS from normal weight fetuses. In LGA FCS, IGF-I, insulin, and IGFBP-3 were significantly increased, whereas IGFBP-1 was significantly decreased. During the neonatal period, IGF-I levels on day 0 were 4-fold higher in FCS from term (38-40 weeks) compared to preterm (25-31 weeks) newborns. FCS IGF-II levels did not change significantly on day 0 between 25-40 weeks gestation. In the first 7 days of postnatal life, IGF-I levels were unchanged in preterm newborns, whereas in term neonates, IGF-I levels decreased precipitously on day 1, remained low during the first 3 days of life, and returned to birth levels by the end of the first week. In contrast, IGF-II and IGFBP-3 levels did not significantly change during the first week of life in preterm or term newborns.(ABSTRACT TRUNCATED AT 400 WORDS)

Oestradiol is important in the growth of uterine leiomyomata and may act primarily or secondarily through mediators such as growth factors, including the insulin-like growth factors (IGF-I and IGF-II), mitogenic peptides. IGF binding proteins (IGFBPs) modulate IGF actions at their target cells. The objective of this study was to examine the possible steroid dependence of IGF, IGFBP and IGF receptor gene expression and IGFBP synthesis in uterine leiomyomata, using tissues from women cycling normally and made hypo-oestrogenic by a gonadtrophin-releasing hormone agonist (GnRHa). Using a solution hybridization ribonuclease protection assay, anti-sense RNA probes for IGF-I, IGF-II and beta-actin (control) were hybridized with total RNA isolated from leiomyomata exposed in vivo to a range of serum oestradiol (< 40-240 pg/ml) and progesterone (0-10 ng/ml) concentrations. IGF-I gene expression was most abundant in leiomyomata obtained during the late proliferative phase of the cycle and was undetectable in leiomyomata from hypo-oestrogenic patients. IGF-II gene expression was not dependent on endogenous steroid concentrations or cycle stage. IGFBP gene expression was investigated by Northern blotting. The order of relative abundance of IGFBP mRNAs was IGFBP-4 >>> IGFBP-3 >> IGFBP-5 > IGFBP-2 and was not dependent on the in-vivo oestrogen status. Type I and type II IGF receptor gene expression was investigated by polymerase chain reaction using gene-specific primers. Type I and type II IGF receptor mRNAs were detected in leiomyomata and were not dependent on cycle stage or in-vivo oestrogen status. Explant cultures of leiomyomata and myometrium synthesized IGFBP-3 (mol. wt = 38-43 kDa), IGFBP-4, and binding proteins of mol. wt = 34 and 31 kDa. Identification of IGFBP-2 was inconclusive, and IGFBP-1 was not detected. These data support the hypothesis that IGF-I, but not IGF-II, may be a mediator of oestradiol action in the growth of uterine leiomyomata, and that IGFBPs may further modulate, by an autocrine or paracrine mechanism, IGF-I action in this tissue.

To study the cellular patterns of gene expression of the insulin-like growth factor (IGF) system in human endometrium during the menstrual cycle, we used in situ hybridization histochemistry to localize messenger ribonucleic acids (mRNAs) encoding IGF-I and -II, their receptors, and their binding proteins (IGFBPs) in fresh-frozen endometrial tissue obtained from cycling women. IGF-I and IGF-II mRNAs are both expressed diffusely throughout endometrial stroma and are not detected in endometrial epithelium. Endometrial IGF-I mRNA is significantly more abundant during the proliferative than the secretory phase of the menstrual cycle, whereas the reverse is true for IGF-II. Type I and type II IGF receptor mRNAs are both present in endometrial stroma, but are relatively more abundant in endometrial epithelium, and neither shows distinctive cyclic changes. IGFBP-2, -4, -5, and -6 mRNAs demonstrate a diffuse stromal pattern of expression, whereas IGFBP-1 and -3 are more focally concentrated in selected subpopulations of endometrial cells. IGFBP-1 mRNA is not detected in proliferative endometrium and demonstrates a very heterogeneous pattern of expression in secretory endometrium, where it is intensely abundant in a patchy distribution of stromal and epithelial cells. IGFBP-3 mRNA is primarily concentrated in endometrial capillaries and is increased in the secretory phase, largely due to the intense vascularization of endometrial glands during this phase. IGFBP-5 mRNA is more abundant in the proliferative phase, but all other IGFBP mRNAs are relatively increased in the secretory phase of the menstrual cycle. These findings support the view that the IGF system plays a fundamental role in endometrial biology, acting via autocrine and/or paracrine mechanisms, with IGF-I and IGFBP-5 being dominant in the proliferative phase, and IGF-II and the other IGFBPs predominant in the secretory phase of the menstrual cycle.