The role of the bacterial microbiota on reproductive and pregnancy health.

Recent assessments have examined the composition of bacterial communities influencing reproductive, pregnancy and infant health. The Microbiome Project has made great strides in sequencing the microbiome and identifying the vast communities of microorganisms that inhabit our bodies and much work continues to examine the individual contribution of bacteria on health and disease to inform future therapies. This review explores the current literature outlining the contribution of important bacteria on reproductive health among sexually active men and women, outlines gaps in current research to determine causal and interventional relationships, and suggests future research initiatives. Novel treatments options to reduce adverse outcomes must recognize the heterogeneity of the bacteria within the microbiome and adequately assess long-term benefits in reducing disease burden and re-establishing a healthy Lactobacillus-dominant state. Recognizing other reservoirs outside of the lower genital track and within sexual partners as well as genetic and individual moderators may be most important for long-term cure and reduction of disease. It will be important to develop useful screening tools and comprehensively examine novel therapeutic options to promote the long-term reduction of high-risk bacteria and the re-establishment of healthy bacterial levels to considerably improve outcomes among pregnant women and sexually active men and women.

CYP17 genotype modifies the impact of anthropometric variation on salivary estradiol in healthy women.

Several studies demonstrate that human ovarian function is responsive to the energetic environment, which has led to the development of theoretical models that explain this phenomenon. Although many genes are involved in ovarian hormone production, the possibility that genetic polymorphism may affect ovarian response to energetic conditions has not been considered. Cytochrome P450c17α is an enzyme that produces androgen precursors used to make estrogens during ovarian steroidogenesis, and is encoded by the CYP17 gene. A functionally significant variant within the promoter region of CYP17 has been linked to variation in steroid production, and some evidence suggests that this polymorphism could alter transcription of CYP17 in an insulin-dependent manner. We tested the hypothesis that the CYP17 variant affected the relationship between anthropometric measurements and salivary estradiol in healthy women in the United States (n = 28). PCR-RLFP analysis was used to genotype women for the genetic variant, and estradiol was assayed from saliva by EIA. Moderated regression analysis of these preliminary data revealed a significant interaction between waist-to-hip ratio and CYP17 genotype (P = 0.004). Our study provides evidence that gene-environment interactions should be considered in future adaptive models for human ovarian function. Moreover, our results stand to illuminate possible associations between this genetic variant and reproductive disease.

Salivary proteomics in bisphosphonate-related osteonecrosis of the jaw.

OBJECTIVE: The objective of this study was to identify differentially expressed salivary proteins in bisphosphonate-related osteonecrosis of the jaw (BRONJ) patients that could serve as biomarkers for BRONJ diagnosis. SUBJECTS AND METHODS: Whole saliva obtained from 20 BRONJ patients and 20 controls were pooled within groups. The samples were analyzed using iTRAQ-labeled two-dimensional liquid chromatography-tandem mass spectrometry. RESULTS: Overall, 1340 proteins were identified. Of these, biomarker candidates were selected based on P-value (<0.001), changes in protein expression (≥1.5-fold increase or decrease), and unique peptides identified (≥2). Three comparisons made between BRONJ and control patients identified 200 proteins to be differentially expressed in BRONJ patients. A majority of these proteins were predicted to have a role in drug metabolism and immunological and dermatological diseases. Of all the differentially expressed proteins, we selected metalloproteinase-9 and desmoplakin for further validation. Immunoassays confirmed increased expression of metalloproteinase-9 in individual saliva (P = 0.048) and serum samples (P = 0.05) of BRONJ patients. Desmoplakin was undetectable in saliva. However, desmoplakin levels tended to be lower in BRONJ serum than controls (P = 0.157). CONCLUSIONS: Multiple pathological reactions are involved in BRONJ development. One or more proteins identified by this study may prove to be useful biomarkers for BRONJ diagnosis. The role of metalloproteinase-9 and desmoplakin in BRONJ requires further investigation.

Gene x environment interactions impact endometrial function and the menstrual cycle: PROGINS, life history, anthropometry, and physical activity.

OBJECTIVES: We assessed the impact of a high frequency, functionally significant allelic variant of the progesterone receptor gene (PROGINS) on endometrial function and menstrual cycle characteristics. Further we asked whether PROGINS moderates the impact of life history characteristics, anthropometric measures, and physical activity on endometrial function. METHODS: Fifty-two women were genotyped for the PROGINS variant, provided life history information, and had anthropometric measurements made. Women monitored their menstrual bleeding for three cycles, performed mid-cycle urinary ovulation tests, and recorded physical activity. A subset of women provided daily saliva samples and had mid-luteal endometrial thickness measurements taken during the third menstrual cycle. Salivary progesterone was assayed using ELISAs. The direct impact of PROGINS on endometrial and menstrual cycle characteristics was determined via independent t-tests with Bonferroni correction. Interactions between PROGINS and covariates were assessed by moderated regression. RESULTS: PROGINS did not directly impact any indicator of endometrial function. However, PROGINS caused an increase in menstrual cycle length with increasing mid-luteal progesterone levels; the opposite relationship was present in noncarriers (P < 0.05). Additionally, PROGINS interacted with four of six anthropometric measures (BMI, waist circumference, height, and waist-hip ratio) to impact endometrial function, however, interactions between PROGINS and life history variables, or physical activity was limited. CONCLUSIONS: The gene x environment interactions we report suggest that PROGINS alters endometrial sensitivity to maternal energetic condition. Thus, the possibility of genetically-based variation in sensitivity to energetic stress should be considered in future adaptive models of women's reproduction.

Worldwide distribution of allelic variation at the progesterone receptor locus and the incidence of female reproductive cancers.

OBJECTIVES: Global patterns of the incidence of cancer are often attributed to environmental and lifestyle differences between regions. Less attention has been given to global patterns of allelic variation of genes that may contribute to the risk of developing cancer. METHODS: We genotyped samples from 21 populations for four variants of the progesterone receptor (PR) gene. One is an Alu insertion in intron 7 which defines the PROGINS haplotype. The others include a promoter region SNP 331+ G/A (rs10895068), a haplotype defining T/C substitution in intron 6 (rs561650), and an A/T substitution (rs608995) in the 3' untranslated region of the gene. All variants have been investigated elsewhere in association with female reproductive cancers in western populations. RESULTS: We found population differences in the frequency of each of these alleles across study populations (P < 0.01, log-likelihood G statistic, computed in FSTAT) and therefore examined the correlation between the frequency of each genetic variant and the incidence of three female reproductive cancers (breast, uterine, and ovarian) obtained from the Globocan 2008 database. Breast and ovarian cancer incidence were significantly correlated with the frequency of the Alu insertion (r = 0.86 and 0.53) and the +331 A variant (r = 0.57 and 0.73). CONCLUSIONS: Our data expand the information on genetic variation at the PR locus in non-western populations and support an argument for more work on the genetic epidemiology of cancer among nonwestern populations.

Dexamethasone enhances fertility and preovulatory serum prolactin levels in eCG/hCG primed immature rats.

Glucocorticoids have heterogeneous effects on reproductive function. We used a gonadotropin-primed, immature rat model to study the influence of dexamethasone (1 mg/kg), given during the latter stages of follicular development, on litter size, the number of oocytes released, and pituitary hormone levels. Dexamethasone-treated females released a larger number of oocytes at ovulation and gave birth to larger litters indicating the oocytes were viable. Survival to weaning age was not affected but average weight at weaning was lower for pups born to DEX-treated females. Serum FSH and LH were assayed at 12, 24 and 48 h following eCG and did not differ between dexamethasone-treated and control animals, but prolactin showed a prolonged pattern of elevation in DEX-treated females. Prolactin, which normally exhibits an elevation on proestrous, may modulate follicular development. Dexamethasone enhances fertility and fecundity possible through an effect of prolactin on follicle development, or by other direct effects on the ovary. These results may improve our understanding of the usefulness of DEX in assisted reproductive therapies for women.

First record of Anguillicoloides crassus (Nematoda) in American eels (Anguilla rostrata) in Canadian estuaries, Cape Breton, Nova Scotia.

In the summer of 2007, American eels, Anguilla rostrata, from 2 localities on Cape Breton Island, were found to be infected with the swim bladder nematode Anguillicoloides crassus. This is the first documented report of this highly invasive parasite in Canadian waters. More than half of the yellow eels in Mira River (6 of 10), and 1 eel (of 5) from Sydney Harbour were infected. Parasite intensity ranged from 1 to 11 worms per eel. The occurrence of A. crassus at these 2 localities suggests the need for a more extensive survey on the distribution of this exotic parasite in eel populations throughout Cape Breton Island.

Chronic hypoxia diminishes the proliferative response of Guinea pig uterine artery vascular smooth muscle cells in vitro.

The pregnancy-related size enlargement of the guinea pig uterine artery is partially accomplished by hyperplasia in all layers of the vessel wall. We sought to determine the separate and combined effects of chronic hypoxia and pregnancy on the proliferative capacity of uterine artery vascular smooth muscle cells (UA VSMCs). We established primary UA VSMC cultures from a total of 13 guinea pigs using an enzymatic digestion technique. Animals were bred and kept in normoxia or hypoxia (P(B) = 463 mmHg, simulated elevation = 3962 m) for 45 days, a duration equivalent to midpregnancy in the guinea pig 63-day gestation. Nonpregnant matched controls were included. The proliferative response of UA VSMCs to 1, 3, 5 or 7 days of serum stimulation in vitro was compared. Exposure to hypoxia reduced UA VSMC proliferative response to serum stimulation relative to that seen in cells harvested from normoxic females. The inhibitory effect was present both in cells harvested from nonpregnant and pregnant animals and resulted in a lower UA VSMC proliferative response in the cells harvested from hypoxic compared with normoxic pregnant animals. Our data were consistent with our hypothesis that chronic maternal hypoxia compromises the capacity for growth and remodeling of the uterine artery during pregnancy, perhaps by interfering with the ability of vascular smooth muscle cells to de-differentiate to a proliferative phenotype. Noteworthy was that such effects of chronic hypoxia were retained in cultured cells.

Human physiological adaptation to pregnancy: inter- and intraspecific perspectives.

Reproductive success requires successful maternal physiological adaptation to pregnancy. An interspecific perspective reveals that the human species has modified features of our haplorhine heritage affecting the uteroplacental circulation. We speculate that such modifications - including early implantation and deep, widespread invasion of fetal (trophoblast cells) into and resultant remodeling of maternal uterine vessels - are responses to or compensation for the biomechanical constraints imposed by bipedalism which, in turn, render our species susceptible to the pregnancy complication of preeclampsia. Preeclampsia is characterized by incomplete remodeling of maternal uterine vessels as the result of shallow trophoblast invasion, which in turn reduces uteroplacental blood flow and frequently leads to intrauterine growth restriction (IUGR). Using an intraspecific perspective, we consider the fitness-related consequences of variation in uteroplacental blood flow during high-altitude pregnancy. Although birth weights are reduced at high altitudes in Bolivia, multigenerational Andean residents are relatively protected from altitude-associated IUGR. Our preliminary data suggest that Andean women have greater uteroplacental oxygen delivery than European high-altitude residents due to more complete growth and remodeling of maternal uterine vessels. Identification of the physiological and genetic mechanisms involved in such inter- and intraspecific variations in pregnancy physiology will likely be useful for understanding human evolution and contemporary challenges to successful reproduction.

Inhibition of vascular endothelial growth factor/vascular permeability factor action blocks estrogen-induced uterine edema and implantation in rodents.

Estrogen induces a rapid increase in microvascular permeability in the rodent uterus, leading to stromal edema and a marked increase in uterine wet weight. This edema is believed to create an environment optimal for the growth and remodeling of the endometrium in preparation for implantation and pregnancy. Increased endometrial microvascular permeability also occurs in conjunction with implantation. Estrogen-induced uterine edema is immediately preceded by an increase in the expression of vascular endothelial growth factor (VEGF), a potent stimulator of microvascular permeability. The objective of this study was to determine to what degree immunoneutralization of VEGF would interfere with a) estradiol-induced uterine edema and b) pregnancy. In the first set of experiments, immature female rats were injected with either VEGF antiserum or normal rabbit serum (NRS) prior to 17beta-estradiol treatment. Rats treated with estradiol alone showed a 57% increase in uterine wet weight at 6 h compared with controls. Injection of 200 or 300 micro l of VEGF antiserum reduced the response to only 20% and 10% above controls, respectively. In the second set of experiments, young adult female mice were treated with 100 micro l of either VEGF antiserum or NRS at 1200 h on the fourth day after mating. NRS-treated mice had normal pregnancies. VEGF antiserum, however, completely blocked pregnancy. When VEGF antiserum-treated females were examined on Day 5 for the presence of implantation sites, none were found. These results show that a) VEGF is the major mediator of estrogen-induced increase in uterine vascular permeability and b) VEGF-induced edema is absolutely essential for implantation to take place.

Chronic hypoxia diminishes pregnancy-associated DNA synthesis in guinea pig uteroplacental arteries.

Enlargement of the uterine artery (UA) during pregnancy is diminished in women residing at a high altitude. We asked whether chronic hypoxia alters the rise in DNA synthesis in uteroplacental vessels and, if so, whether the reduction is related to the intrauterine growth retardation (IUGR) observed under conditions of chronic hypoxia. We used bromodeoxyuridine (BrdU) labelling to measure DNA synthesis in all vascular layers of the UA, mesometrial arteries (MA), thoracic aorta and mesenteric artery of guinea pigs, residing throughout pregnancy at a low (1600 m) or high (3962 m) altitude. Pregnancy increased DNA synthesis throughout the UA at both altitudes, yet the maximal value was less at high than low altitude (P<0.05). Likewise, pregnancy increased DNA synthesis throughout the MA, yet at high altitude pregnancy elevated levels returned to non-pregnant values after 42 days of gestation, whereas at low altitude DNA synthesis continued to be elevated until near term. Fetal weights were lower (P=0.01) and placental/fetal weight ratios tended to be greater in high than low altitude, near term pups (P = 0.09). We conclude that a diminished growt response by the uteroplacental vasculature to pregnancy may contribute to the previously reported reduced uterine artery blood flow and resulting IUGR at high altitude.

Relaxin stimulates uterine edema via activation of estrogen receptors: blockade of its effects using ICI 182,780, a specific estrogen receptor antagonist.

Relaxin's ability to stimulate uterine growth is well established. The mechanisms by which relaxin exerts this effect, however, remain unclear. In light of previous work demonstrating peptide growth factor activation of estrogen receptors (ERs), the present study was conducted to determine if relaxin similarly stimulates ERs. Twenty-five day-old female Sprague-Dawley rats were bilaterally ovariectomized and treated with estradiol or porcine relaxin alone or in combination with the ER antagonist ICI 182,780. Following treatment with 17beta-estradiol or relaxin alone, the uterine weight/body weight ratio (UtW/BW) increased significantly over control values (+98% and +77% respectively, p<0.0003). Pre-treatment of animals with ICI 182,780 (3 microg/g BW) prior to either estradiol or relaxin treatment completely inhibited the hormone-induced increases in uterine weight (p<0.0005). ICI 182,780 alone had no significant effect. Histological analysis of uterine cross-sections revealed that the edema present in the endometrium of animals treated with estradiol or relaxin alone was completely absent in the uteri of animals pre-treated with ICI 182,780. These data indicate that relaxin-induced uterine edema and growth is mediated by ERs.

In vitro cytotoxicity of amalgams made with binary Hg-In liquid alloys.

OBJECTIVE: Mercury vapor release from amalgams during setting significantly decreases when the amalgams are prepared with binary Hg-In liquid alloys. The objective of this study was to compare the cytotoxicity of amalgams made with experimental Hg-In alloys with that of amalgam without In and a commercial In-containing amalgam. METHODS: Amalgam specimens were prepared by triturating a high-Cu alloy powder (Tytin, Kerr) with pure Hg or Hg-In liquid alloy containing 5, 20 or 50% In and also by triturating an In-containing high-copper alloy powder (Indiloy, Shofu) with pure Hg. After the specimens were aged for 2 wk, a cylindrical specimen of each amalgam was immersed consecutively in cell culture medium for 0-8, 8-48 and 48-72 h. The cytotoxicity of the extracts was determined by placing them in contact with Balb/c 3T3 mouse fibroblasts for 24 h, after which the succinic dehydrogenase (SDH) activity was measured and expressed as a percentage of the Teflon negative controls. The results were statistically compared using ANOVA and Tukey's test (alpha = 0.05). The concentration of elements released into the extracts was determined by atomic absorption spectrophotometry and evaluated by Kruskal-Wallis and nonparametric multiple comparisons. RESULTS: For the 0-8 h and 8-48 h intervals, the 20% In amalgam was significantly (p < 0.05) less toxic than the other amalgams, and not different from the Teflon control. Results for the other amalgams were only slightly depressed compared to the Teflon control. For the 48-72 h interval, all amalgams were essentially no different from the control. Copper was the element dominantly released into the medium from all the amalgams tested. SIGNIFICANCE: For amalgam tested after aging, alloying indium to mercury did not deleteriously affect the cytotoxicity of the resultant amalgam compared to the amalgam without indium.

Determination of the flavor attributes of cooked beef from cross-bred Angus steers fed corn- or barley-based diets.

Eighteen commercial Angus cross-bred feedlot steers of similar hip height and live weight were randomly assigned to one of three dietary treatment groups: corn-, corn/barley, or barleybased diets (n = 6 per treatment). Steers were fed for 102-103 days on their respective diets prior to slaughter. Live animal performance traits, carcass characteristics, total lipid and descriptive flavor and descriptive palatability attributes of beef strip loin steaks were determined. End live weight (P = 0.88) did not differ between dietary treatments. Beef carcasses from steers fed corn-, barley-, and corn/barley-based diets did not differ in hot carcass weight (P = 0.18), ribeye area (P = 0.21), kidney, pelvic and heart fat (KPH) (P = 0.35), and yield grade (P = 0.14). However, adjusted preliminary yield grade was higher (P = 0.03) for carcasses from steers fed corn/barley-based diets than carcasses from steers fed barley as the dietary energy source. These data suggest that carcasses from steers fed barley-based diets were lower in external fat. Quality grade characteristics were not different in beef carcasses from steers fed either corn-, barley-, or a corn/barley-based diet. Cook time (P = 0.37), cooking loss (P = 0.83), descriptive meat palatability attributes (P > 0.27), Warner-Bratzler shear force (P = 0.25), and descriptive sensory flavor attributes (P ≥ 0.17) did not differ for steaks from steers fed the three diets prior to slaughter. The Japanese have claimed that feeding cattle barley-based high energy diets result in beef with different flavor than when cattle are fed high-energy corn-based diets. These results indicated that the eating quality, tenderness and flavor attributes of beef steaks were not influenced by the dietary grain source fed to young steers in this study prior to slaughter.

Epidemiology of adult sexually transmitted disease agents in children being evaluated for sexual abuse.

This prospective study describes the epidemiology of adult sexually transmitted disease agents in 1538 children ages 1 to 12 years being evaluated for possible sexual abuse. Infections with these agents were related to the presence or absence of a history of sexual contact. Neisseria gonorrhoeae (GC) was found in 2.8% (41 of 1469); human papillomavirus presenting as condyloma acuminata, 1.8%; Chlamydia trachomatis, 1.2% (17 of 1473); Treponema pallidum (syphilis), 0.1% (1 of 1263); and herpes simplex virus, 0.1%. Overall a history of sexual contact was present in 83% of children with N. gonorrhoeae; condyloma acuminata, 43%; Chlamydia trachomatis, 94%; syphilis, 0%; and herpes simplex virus, 50%. Selected vaginal discharges were examined for Trichomonas vaginalis and bacterial vaginosis. In children comprehending questions regarding sexual contact (i.e. were "verbal"), 89% with N. gonorrhoeae, 100% with Chlamydia trachomatis and 63% with condyloma acuminata had a history of sexual contact, indicating that in "verbal" children any infection with N. gonorrhoeae or C. trachomatis was highly associated with sexual contact.

Adaptive enhancement of amino acid uptake and exodus by thymic lymphocytes: influence of pH.

Entry of certain free amino acids (alpha aminoisobutyric acid (AIB), alanine and proline), but not of leucine into rat thymic lymphocytes increased progressively when the cells were incubated in amino acid deficient medium. Actinomycin D, cycloheximide, or a high concentration of AIB abolished the time-related increase in AIB accumulation, whereas exposure to a high concentration of leucine had no effect. This phenomenon could not be attributed to a progressive alteration in the nature of the incubation medium nor to reduced transinhibition of AIB uptake. The exodus of AIB also increased with time, but to a smaller degree than AIB entry. Initial rates of AIB entry and exodus increased with increases in the pH of the incubation medium over the range 6.5-8.0. The effects of pH on entry and exodus were time-related, increasing progressively oveb nullified the magnified time related increments in AIB transport caused by prolonged incubation at pH 8.0. The influence of a given pH on transport of AIB decreased rapidly when the cells were transferred to medium of another pH, but this tendency diminished the longer the cells were exposed to the initial pH. pH influenced the entry of alanine and proline in the same fashion as that of AIB, but did not affect leucine entry. These results indicate that thymic lymphocytes exhibit adaptive enhancement in the accumulation of free amino acids that are transported largley by the A or alanine-preferring system, and that the adaptive process involves both entry and exodus. Moreover, alterations in pH modify entry and exodus of these same amino acids, profoundly affect the magnitude of time-released increases, and may induce fundamental changes in the mechanism(s) serving amino acid transport.