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Objective: To assess weight change and attempted weight loss during the 12-18 months before spontaneous conception in relation to the risk of pregnancy loss.Design: Prospective cohort study.Setting: United States, 2007-2011.Methods: Women (n = 629) who were attempting pregnancy reported at baseline any weight loss attempts over the past 12 months, and their minimum and maximum weights during that time. Follow up lasted one to six menstrual cycles and throughout pregnancy. Using bodyweight measured at 4 weeks' gestation, participants were categorised as having weight loss ≥5%, weight gain ≥5%, both, or neither, over the previous 12-18 months. Log-binomial models adjusted for potential confounders.Main Outcome Measures: Risk ratio (RR) and 95% confidence interval (CI) of pregnancy loss.Results: Attempted weight loss was reported by 44% of women and actual weight loss by 11%, but neither was consistently associated with pregnancy loss. The RR for recent weight gain ≥5% was 1.65 (CI 1.09, 2.49).Conclusions: Weight gain over the period spanning 12-18 months pre-conception to 4 weeks' gestation may increase the risk of pregnancy loss among fertile women with prior pregnancy losses. Attempted and actual weight loss were not associated with pregnancy loss; however, replication is needed from larger studies with data on particular weight-loss methods.Tweetable Abstract: Recent weight gain before and around the time of conception may increase the risk of pregnancy loss. [ABSTRACT FROM AUTHOR]

Interleukin-1 (IL-1) has been implicated as a participant in preterm labor that is induced by bacterial infection. Previously, we showed that serotonin-induced production of IL-1alpha by myometrial smooth muscle cells in vitro is also essential for the synthesis of interstitial collagenase. It is therefore likely that IL-1alpha production in uterine tissues has implications for both the normal physiology of involution and for the pathophysiological mechanisms of preterm labor. The objective of this study was to characterize the serotonin-induced production of IL-1alpha by myometrial cultures in vitro and to assess the production of IL-1alpha and its relationship to collagenase production in vivo during pregnancy and the postpartum period. Immunohistochemistry demonstrated IL-1alpha protein in the nuclei and cytoplasm of serotonin-treated myometrial cells. IL-1alpha levels were decreased by treatment with progesterone or IL-1-receptor antagonist but were unaffected by lipopolysaccharide. Western analysis of myometrium from pregnant rats showed low levels of IL-1alpha during midpregnancy with increased concentrations at days 21 and 22 and postpartum. IL-1alpha mRNA levels also increased from days 15 to 22. Levels of mRNA for IL-1beta also increased, although to a lesser degree than IL-1alpha. Both mRNAs decreased postpartum. Conversely, mRNA for interstitial collagenase was barely detectable at term but increased postpartum. Together, these data show that serotonin stimulates IL-1alpha production in vitro and indicate that normal myometrium from pregnant rats is an identifiable source of IL-1 during late pregnancy. The findings are consistent with the possibility that myometrial IL-1alpha participates in normal labor as well as the postpartum production of interstitial collagenase.

Primary cultures of uterine smooth muscle cells from post-partum rats express interstitial collagenase in response to serotonin and the serotonin-dependent production of interleukin-1 (IL-1) [Wilcox, B.D., Dumin, J.A. and Jeffrey, J.J. Serotonin regulation of interleukin-1 messenger RNA in rat uterine smooth muscle cells. Relationship to the production of interstitial collagenase J. Biol. Chem., 269, (1994a), 29658]. Transient transfections of these cells indicate that rat collagenase transcription is regulated via a proximal consensus AP-1 site within an extended palindrome. Mutation of either the AP-1 site or extended palindrome (EP) decreases promoter activity to approximately 30% of the wild-type. Electrophoretic mobility shift assays reveal the binding of smooth muscle cell nuclear proteins to the AP-1 EP. This binding is barely detectable after mutation of the EP and is completely eliminated by mutation of the AP-1 heptamer. Competition experiments demonstrate that binding to the AP-1 EP is specific and of higher affinity than binding to oligonucleotides containing a mutated EP. Binding to the AP-1 EP is higher when smooth muscle cells are cultured in the presence of serotonin than in its absence. Although IL-1 is required for collagenase transcription, binding to the AP-1 EP appears to be IL-1-independent. FosB, Fra-2, c-Jun, JunB and, most abundantly, JunD bind the AP-1 EP in the absence and presence of serotonin. In contrast, Fra-1 expression and binding are serotonin-dependent suggesting that the activation of Fra-1 may be a key component of collagenase transcriptional activation.

Previous studies have shown that IgG-coated erythrocytes (EIgG) augment the LPS-stimulated increase in serum TNF-alpha levels in animals and the LPS-stimulated secretion of TNF-alpha by isolated macrophages. The present study evaluated the mechanism for the effect of EIgG on LPS-stimulated TNF-alpha secretion in the murine macrophage cell line, RAW 264.7. Incubation of the macrophages with EIgG or IgG-coated glass beads caused a dose-dependent augmentation of LPS-stimulated TNF-alpha secretion. The addition of EIgG increased the rate of LPS-stimulated TNF-alpha protein secretion between 2 and 4 hr after LPS. Accordingly, EIgG increased the levels of TNF-alpha mRNA at 2 and 3 hr after LPS. The increase in the LPS-stimulated TNF-alpha mRNA levels caused by EIgG was associated with an increase in TNF-alpha mRNA stability. Thus, the augmentation of LPS-stimulated TNF-alpha secretion by EIgG was associated with an increase in TNF-alpha mRNA levels which at least partly resulted from an increase in the stability of TNF-alpha mRNA., (Copyright 2000 Academic Press.)

The regulation, by progesterone, of serotonin-induced interleukin-1alpha production was studied in primary cultures of rat uterine smooth muscle cells. Prior reports from this laboratory have demonstrated that these cells produce IL-1alpha and IL-1beta mRNAs in response to the hormonal action of serotonin. Results of the present study indicate that treatment of myometrial smooth muscle cells with medroxyprogesterone acetate (MPA) results in a marked decrease in IL-1alpha protein as measured by western blot analysis. These decreases occur even in the presence of maximally-inducing concentrations of serotonin. MPA-mediated changes in IL-1alpha protein are characterized by a rapid decline in IL-1alpha mRNA levels. This inhibition by medroxyprogesterone also occurs when cells are stimulated to produce IL-1alpha by PMA rather than serotonin. Thus, when cells are cultured in the presence of both inducer and inhibitor, the inhibitor, progesterone, clearly dominates in the control of IL-1alpha expression. This effect is concentration-dependent, can be mimicked by native progesterone or glucocorticoids, but is unaffected by estradiol. The ability of progestins to decrease IL-1alpha mRNA is blocked by both inhibitors of transcription and translation and by treatment with RU-486. Progesterone had no effect on chloramphenicol acetyl transferase (CAT) transcription from two different IL-1alpha promoter constructs, indicating that progesterone's action appears to be dependent on post-transcriptional rather than transcriptional regulation. Conversely, progesterone accelerated the normal rate of decay of IL-1alpha mRNA that occurs following the removal of serotonin from the cultures. These results suggest that progesterone decreases IL-1alpha levels by stimulating the production of an intracellular intermediate that decreases the stability of IL-1alpha mRNA.

Interleukin-1 has been shown to contribute to infection-induced inflammatory processes during pregnancy. Prior work from this laboratory has demonstrated that serotonin-induced IL-1alpha also is required for the in-vitro production of collagenase in uterine smooth muscle cells, a normal, non-inflammatory process that occurs in-vivo during post-partum uterine involution. To understand the molecular mechanisms that regulate transcription of the IL-1alpha gene in these cells, we isolated and characterized 1.6 kilobases of the 5'-flanking region of the rat IL-1alpha gene. Sequencing and primer extension identified a single transcription start site and multiple potential regulatory elements, including a TATA box at - 30 bp, a CAAT box at - 74 bp, and a conserved AP-1 site at - 9 bp. This 5'-flanking DNA exhibited low basal promoter activity that was inducible by serotonin. Serotonin-induced promoter activity was unaffected or induced by either medroxyprogesterone or IL-1 receptor antagonist. This occurred despite the ability of both of these hormones to markedly decrease IL-1alpha mRNA. Deletional analysis revealed a strong repressor in the region between - 147 and - 98 bp; removal of this sequence resulted in a fivefold higher basal promoter activity that was still serotonin responsive. Constitutive promoter activity appeared to reside between - 97 and - 22 bp. Deletion of this promoter region, which contained the TATA and CAAT boxes and an NF-IL-6/PEA-3 site, resulted in decreased basal transcriptional activity to the low level seen in larger constructs. Mutational analysis showed that serotonin-inducible transcriptional activity was mediated, at least in part, by the conserved AP-1 site at - 9 bp. This site is located within a larger extended palindromic region: 5'-AAGCCTGACTCAGACTT-3', that together effects both the basal and serotonin-inducible expression of the IL-1alpha gene.

We have investigated the mechanism by which thyroid hormone potentiates IFN-gamma-induced HLA-DR expression. IFN-gamma-induced HLA-DR expression requires activation of STAT1alpha and induction of the Class II trans-activator, CIITA. HeLa and CV-1 cells treated only with L-thyroxine (T4) demonstrated increased tyrosine phosphorylation and nuclear translocation (= activation) of STAT1alpha; this hormone effect on signal transduction, and T4 potentiation of IFN-gamma-induced HLA-DR expression, were blocked by the inhibitors CGP 41251 (PKC) and genistein (tyrosine kinase). Treatment of cells with T4-agarose also caused activation of STAT1alpha. In the presence of IFN-gamma, T4 enhanced cytokine-induced STAT1alpha activation. Potentiation by T4 of IFN-gamma action was associated with increased mRNA for both CIITA and HLA-DR, with peak enhancement at 16 h (CIITA), and 2 d (HLA-DR). T4 increased IFN-gamma-induced HLA-DR protein 2.2-fold and HLA-DR mRNA fourfold after 2 d. Treatment with actinomycin D after induction of HLA-DR mRNA with IFN-gamma, with or without T4, showed that thyroid hormone decreased the t(1/2) of mRNA from 2.4 to 1.1 h. HeLa and CV-1 cells lack functional nuclear thyroid hormone receptor. Tetraiodothyroacetic acid (tetrac) and 3,5,3'-triiodo-thyroacetic acid (triac) blocked T4 potentiation of IFN-gamma-induced HLA-DR expression and T4 activation of STAT1alpha. These studies define an early hormone recognition step at the cell surface that is novel, distinct from nuclear thyroid hormone receptor, and blocked by tetrac and triac. Thus, thyroid hormone potentiation of IFN-gamma-induced HLA-DR transcription is mediated by a cell membrane hormone binding site, enhanced activation of STAT1alpha, and increased CIITA induction.

The regulation of collagenase gene expression by serotonin and progesterone was investigated in primary cultures of rat uterine smooth muscle cells. Northern blot analysis demonstrates that serotonin (5-hydroxytryptamine (5-HT)), when administered to cells in serotonin-depleted medium, causes 6-8-fold increases in levels of collagenase mRNA. Selective serotonin 5-HT2 receptor agonists were able to mimic the effect of the natural hormone, while the induction by serotonin could be blocked by 5-HT2 receptor antagonists. Addition of phorbol ester (PMA) to 5-HT-depleted cultures fully mimicked the effect of 5-HT on collagenase mRNA induction. Treatment with progesterone analogs caused a decrease in collagenase mRNA, even in the presence of saturating levels of serotonin or PMA. In all experiments, levels of secreted collagenase were observed to correspond to levels of collagenase mRNA. Experiments with cycloheximide demonstrate that serotonin- and PMA-induced increases in collagenase mRNA are dependent on protein synthesis. Furthermore, nuclear run-on analysis shows that mRNA increases are accompanied by increases in initiation of transcripts. These data indicate that transcription of collagenase mRNA in myometrial smooth muscle cells is stimulated by serotonin, possibly via activation of protein kinase C, but is in some way prevented by the negative influence of progesterone.