Background: Hypertension during pregnancy is a leading cause of birthing parent mortality and adverse pregnancy outcomes. Since non-metropolitan communities face higher rates of several risk factors for hypertension in pregnancy and shortages in obstetrical services, persons residing in non-metropolitan areas may be at increased risk for adverse outcomes compared to those living in metropolitan areas. Our study objectives were to examine by county of birthing parent residence (1) the prevalence of chronic hypertension (cHTN) and hypertensive disorders of pregnancy (HDP), and (2) the prevalence of adverse birthing parent and neonatal outcomes associated with hypertension. Methods: Using U.S. birth certificate data from 2016 to 2018, we described the prevalence of cHTN and HDP and the association of each with several birthing parent and neonatal outcomes, stratified by non-metropolitan versus metropolitan county of birthing parent residence. Multivariable Poisson regression models were used to calculate adjusted prevalence ratios for birthing parent and neonatal outcomes among individuals with cHTN or HDP who lived in non-metropolitan versus metropolitan U.S. counties. Results: The prevalence of cHTN and HDP for US live births was 2.2% and 7.4%, respectively, among non-metropolitan pregnant individuals and 1.8% and 6.6%, respectively, among metropolitan pregnant individuals. After adjusting for several sociodemographic characteristics among those with HDP, the prevalence ratio for an APGAR score < 7 at 5 minutes (aPR 1.34, 95% CI 1.29-1.38) and neonatal death (aPR 1.36, 95% CI 1.15-1.62) was increased among offspring born to women who resided in non-metropolitan counties. Similar results were seen among those with cHTN. Conclusion: The prevalence of cHTN and HDP is modestly more prevalent in non-metropolitan areas, but most pregnancy outcomes were similar among those residing in non-metropolitan areas compared to metropolitan areas. Further research should investigate the robustness of these findings using alternate definitions of rural and urban areas and the possible link between low APGAR score, low NICU admission, and neonatal death in non-metropolitan counties.

Preeclampsia (PE) is a disorder affecting 2-10% of pregnancies worldwide. Clinical signs include high blood pressure (HBP) and proteinuria in the mother after the 20th week of pregnancy. Currently, the only cure for PE is delivery of the fetus, which is often necessary preterm and thus dangerous for both mother and fetus. Maternal symptoms of PE are caused by excess anti-angiogenic proteins of placental origin called soluble Flt1s (sFlt1s). sFlt1 mRNA isoforms are produced by alternative polyadenylation (APA) of full-length Flt1 (fl-Flt1) pre- mRNA. While fl-Flt1 encodes a transmembrane protein, sFlt1s encode truncated proteins that are soluble. Multiple sFlt1 isoforms exist, and their respective contribution to the pathophysiology of PE is unclear. Furthermore, it is unknown whether there is a genome-wide role for APA in PE. In my thesis research, I developed a polyadenylation site sequencing method, and used this method to simultaneously quantify transcriptome-wide polyadenylation site usage and gene expression levels in normal, early-onset PE, and late-onset PE human placentae. I observed distinct expression profiles amongst the three groups, with differential expression of genes in several functional categories, including angiogenesis. I found that three sFlt1 isoforms account for >94% of all placental FLT1 transcripts, and that increased transcription of the entire FLT1 locus drives upregulation of both fl-Flt1 and sFlt1 in PE. I found that APA does not contribute substantially to PE pathophysiology. I also identified siRNAs that knock down sFlt1 mRNA efficiently in cell lines that pave the way for further development of novel RNAi based therapeutics to alleviate PE.

Healthy People 2020 was launched in December 2010 with a target human papilloma virus (HPV) vaccination rate of 80%. As of 2014, we were well short of this goal, especially among males, for whom the HPV vaccine became recommended in 2011. An estimated 14-20% of adolescent-aged males had completed the vaccine schedule as of 2015. This is particularly problematic in northeast Tennessee, as multiple risk factors for lower vaccination rates are characteristic of the population, including being white and living in the South. Reasons to decline vaccination vary, and usually involve concerns about safety, efficacy, and necessity. Worries about sexual disinhibition from being vaccinated are often cited by opponents. Parents also perceive a lack of benefit from getting the vaccination starting at age 11 before their sons are sexually active. The media and internet are also barriers to appropriate vaccination in males, as previous research has demonstrated that media coverage is more likely to focus on political controversies instead of benefits, and is more likely to emphasize the benefits to females. Research on improving vaccine uptake on males is currently limited. Doctors who appear knowledgeable and are willing to spend time talking about the HPV vaccine for male patients may increase vaccination rates. Other interventions that may also be effective include vaccinating as part of nurse visits or through school programs. Early studies have been mixed on the effect of patient and parent education on vaccine uptake, although a 2015 review demonstrated that most practice- and community-based educational interventions have some positive effect on uptake. The purpose of this project was to improve HPV vaccine uptake among male patients in a family practice residency program through patient and parent education. After informed consent was obtained, the patients and/or their parents were given a handout produced by the CDC highlighting the benefits of vaccination for males. A chart review was done to determine vaccine coverage among males before the intervention instituted in November of 2016. The intervention was completed in August 2017, and a repeat chart review is currently ongoing to determine vaccine coverage in the post-intervention period. Data collection and analysis is ongoing at the time of abstract submission. We expect a statistically significant increase in the number of male patients who have received any doses of vaccine, and in the number who have completed the vaccine series. Future research should involve broadening the intervention to include local family medicine and pediatrician’s offices to increase vaccine uptake in these populations as well.

The hypothalamo-neurohypophysial model has been studied for many years. Of note, when the axons of the magnocellular, peptidergic neurons of the supraoptic nucleus (SON) and paraventricular nucleus (PVN) are transected or crushed, varying degrees of polydipsia and polyuria ensue as the result of measurable losses of vasopressin (AVP) within the organism's circulation. Following insult, these hypothalamic cells show a remarkable capacity to reorganize themselves within the proximal areas of the infundibular stalk and median eminence and form what has come to be known as a new 'mini neural lobe' . While the surviving neurons sprout new projections toward the level of the external zone, vascular hypertrophy is marked throughout the new neurohypophysis and new neurohemal contacts have been identified (at the ultrastructural level) associated with these vessels. In parallel with this vascular hypertrophy is a measurable re-release of vasopressin into the circulation. This new 'mini neural lobe' now has the morphological and physiological appearance of an intact neural lobe and is capable of releasing AVP in response to changes in water balance. While the ability of these axons to reorganize is more characteristic of the peripheral nervous system (PNS), this model system provides an unique opportunity to study axonal regeneration of the central nervous system (CNS). Not only the mechanisms underlying the restoration of AVP function following axotomy but the extent to which various magnocellular neuron populations are involved in the regenerative process may also be analyzed. Before attempting to identify putative markers associated with this regenerative process, it was necessary to carefully characterize the system following axonal injury. Using Sprague Dawley rats, we repeated previous physiological studies which had examined the intake of water and output of urine following hypophysectomy. In addition, we also correlated the restoration of water balance with the return of AVP release, as measured by radioimmunoassay. These data defined a temporal framework in which magnocellular AVP regeneration occurs. As a result of repeating these physiological studies, we noted several inconsistencies between other previously published work. First, the time course of AVP recovery did not agree with other published results, nor did the first appearance of AVP immunoreactivity . We did not observe a complete recovery of water balance as previously reported and the degree of magnocellular death was inconsistent with other reports. In light of these many conflicting observations between several historical reports and our own results, we did a basic physiological re-characterization of the hypothalamo-neurohypohysial system following hypophysectomy. By means of immunohistochemistry, we also demonstrated the re-appearance of AVP within the new the 'mini neural lobe ' concomitant with the increased appearance of synapsin I, a marker associated with the presence of mature and presumably functioning synapses to be no sooner than 28 days following surgical removal of the hypophysis. Immunocytochemistry was also used in conjunction with retrograde fluorescent labeling to extend the previous studies and include a 2-D analysis of cell survival throughout the PVN and SON following hypophysectomy or neurohypophysectomy. As reported previously, magnocellular neuronal loss is greater within the SON, particularly the hypophysectomized subject, and less so within the PVN; again with the greater loss in the PVN of the hypophysectomized animal. Based upon our observations and other recent reports, we suggest the possibility that some cells of the hypothalamo-neurohypophysial system or some other extrahypothalamic cell population may be capable of expressing vasopressin in response to neurohypophysectomy. We provide initial evidence that glial cells of the third ventricle may indeed be involved. Finally, one of the ultimate goals of using this as a model system of CNS regeneration is to understand the underlying mechanisms and components essential to central nervous tissue regeneration. Toward that end I have been involved with the initial studies to optimize an adenovirus delivery system which will be capable of incorporating various putative neurotransmitter and/or peptide anti-sense messages, being injected into the neurohypophysis and transported back into the cells of the hypothalamo-neurohypophysial system. Once these antisense sequences are expressed by the cells following axotomy, the sequence of expression of various proteins in response to injury may be elucidated.

Common, complex disorders are polygenic and multifactorial traits representing interactions between environmental, genetic and epigenetic risk factors. More often than not, contributions of these risk factors have been studied individually and this is especially true for complex reproductive traits where application of genomic technologies has been challenging and slow to progress. This thesis explores the potential of genetic and epigenetic components contributing to a better understanding of the biological pathways underlying disease risk in two specific female complex reproductive traits - polycystic ovary syndrome (PCOS) and preterm premature rupture of membranes (PPROM). The PCOS projects focus on characterization of a gene, DENND1A, whose association to PCOS has been established by Genome Wide Association Studies (GWAS) and is known to contribute to PCOS steroidogenic phenotype. In addition, differential microRNAs expression contributing to DENND1A expression regulation in PCOS theca cells was identified. The studies on PPROM utilize a Whole Exome Sequencing approach to identify rare variants in fetal genes contributing to extracellular matrix composition and synthesis contributing to PPROM risk. The results suggest that fetal contribution to PPROM is polygenic and is driven by a significant genetic burden of potentially damaging rare variants in genes contributing to fetal membrane strength and integrity. Tissue and location specific expression patterns of the Chromosome 21 miRNA cluster (miR-99a, miR-125b, let-7c) in fetal membranes from term pregnancies with spontaneous rupture were investigated. The results suggest that these miRNAs play potential roles in fetal membrane rupture and fetal membrane defects associated with T21.

Stress urinary incontinence is prevalent among women. Pelvic floor muscle exercise has been found to be an effective treatment for management of women with stress urinary incontinence. Women's non-adherence to pelvic floor muscle exercise has been the major obstacle for achieving desirable treatment outcomes. The purpose of this study was to examine the relationships between the severity of stress urinary incontinence, impact of incontinence, self-esteem, and intrinsic motivation among Chinese women with stress urinary incontinence and to examine the implications of these relationships on Chinese women's adherence to pelvic floor muscle exercise. / The model developed from the study explained that Chinese women's adherence to pelvic floor exercise is influenced by the interaction among contextual, intrinsic and extrinsic components. Throughout learning and practicing pelvic floor muscle exercise, women's level of motivation to learn and adhere to exercise can be increased or decreased as these components interacted with each other. Finally, the study also highlights implications for nursing practice including the importance of assessment of women with urinary incontinence, awareness of factors influencing adherence to the exercise, and promoting women's adherence to the exercise by fostering women's exercise competency and commitment to exercise and raising women's awareness of the benefits of exercise. / The phase two findings showed that women's perception of severity of incontinence was influenced by sociocultural factors, self-esteem, and level of knowledge. The findings also indicated that about one-third of women were reluctant to disclose their incontinence problems to their partner. This was partly due to their beliefs of linkage between incontinence and deterioration of sexual function, and their conservative view regarding incontinence as a private matter which should not be discussed with their partner. Furthermore, the findings indicated that women's competency, commitment, perceived benefits of pelvic floor muscle exercise and perceived support from family and health professionals were the factors influencing women's adherence to pelvic floor muscle exercise. Importantly, the study revealed that motivation is dynamic in nature from women's initial learning to maintenance of the exercise. Furthermore, motivation was influenced by beliefs in effectiveness of the exercise, perceived exercise benefits and severity of incontinence. / The study employed a mixed method approach using a quantitative design in phase one and a qualitative design in phase two. The results of phase one of the study showed that the majority of women had low level of severity and impact of incontinence; however, 70% of women had high level of motivation to engage in pelvic floor muscle exercise. The results also demonstrated that women's self-esteem was influenced by the severity of incontinence in terms of consumption of pads, as well as the impact of incontinence on different aspects of life being affected by stress urinary incontinence. In addition, those women who had a higher level of severity in terms of increased wetting in previous year and previous week, as well as increased numbers of activities being affected by incontinence were more likely to have a higher level of motivation to adhere to pelvic floor muscle exercise. Furthermore, women with higher levels of impact of incontinence in terms of being affected on sexual life also demonstrated to have a higher level of motivation to adhere to pelvic floor muscle exercise. / Siu Lai Sheung Katherine. / "August 2006." / Adviser: Sheila Twinn. / Source: Dissertation Abstracts International, Volume: 68-03, Section: B, page: 1562. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 266-300). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Transforming growth factor -β (TGF-β) plays an important role in the progression of prostate cancer. It acts as a tumor suppressor in normal epithelial cells but as a tumor promoter in advanced prostate cancer cells. The PI3-kinase pathway has been shown to play integral roles in many cellular processes including cell proliferation, survival, and cell migration in many cell types. PI3-kinase pathway mediates TGF-β effects on prostate cancer cell migration and invasion. Phosphatase and tensin homolog (PTEN), a tumor suppressor gene, inhibits PI3-kinase pathway and is frequently mutated in prostate cancers. In this present study, we investigated possible roles of PTEN in TGF-β effects on proliferation, migration, and the activation of PI3-kinase/AKT pathway in prostate cancer cells. PTEN was expressed in DU145 cells; however PC3 cells lack PTEN expression. TGF-β1 and TGF-β3 had no effect on PTEN mRNA levels but both isoforms increased PTEN protein levels in DU145 and RWPE1 cells, suggesting that TGF-β may mediate regulation of PTEN protein stability. TGF-β1 and TGF-β3 increased PTEN protein levels even in the presence of cycloheximide, a protein synthesis inhibitor, in DU145 cells. In addition, TGF-β upregulated phosphorylation of PTEN, stabilizing PTEN protein. Increase of PTEN protein levels in these cells may also indicate that PTEN may mediate TGF-β effects on cell proliferation. Knockdown of PTEN in DU145 cells resulted in significant increase in cell proliferation which was no longer affected by TGF-β isoforms. PTEN overexpression in PC3 cells inhibited cell proliferation. Knockdown of endogenous PTEN enhanced cell migration in DU145 cells, whereas PTEN overexpression reduced migration in PC3 cells and reduced phosphorylation of AKT in response to TGF-β. Based on these results, we conclude that PTEN plays a role in inhibitory effects of TGF-β on cell proliferation whereas its absence may enhance TGF-β effects on activation of PI3-kinase pathway and cell migration.

Prostate cancer increases its incidence with age after men in their fifth decade as the ratio of estrogen to androgen rises. Epidemiological studies indicated that high levels of estrogens are associated with the high-risk ethnic groups for prostate cancer. Therefore, estrogens may be involved in prostatic carcinogenesis. It is widely believed that the actions of estrogens are mediated by estrogen receptors. However, expression of estrogen receptor in normal prostate and lesions of the gland was controversial. With the recent discovery of second estrogen receptor (ER-β), this issue became more complicated and it needs to be readdressed. In addition, the biological involvement of ER-β in human prostate remains to be investigated. In this study, we demonstrated that human normal prostate epithelial cells express ER-β but not ER-α, suggesting that estrogens act directly on these epithelial cells via ER-β. Using RT-PCR analysis, the transcripts of ER-β were detected in our primary human prostatic epithelial cell cultures that were derived from the ultrasound-guided peripheral zone biopsies and the cells express two estrogen-regulated genes such as progesterone receptor (PR) and pS2. Moreover, we had developed an ER-β antibody with fully characterizations and used it for immunohistochemistry. Results indicated that ER-p protein is expressed in the basal compartment of prostatic epithelium of the gland. Our findings lead to a new hypothesis that estrogens directly act on human prostatic epithelial cells to modulate its biological functions. To investigate expression of ERs in prostate cancer, RT-PCR analysis was used. We found that all three human prostate metastatic cancer cell lines, DU145, PC-3 and LNCaP, express ER-β transcripts while ER-α mRNA expression only in PC-3 cells. Expressions of PR and pS2 in these cell lines are various. LNCaP cells express both PR and pS2 mRNAs but DU145 cells with only PR and PC-3 cells with only pS2. Our immunohistochemical results on prostatic lesions revealed down-regulation of ER-β expression in high-grade of dysplasia and carcinoma of peripheral zone of the prostate compared to their low-grade lesions. This down-regulation in high-grade carcinoma was verified in transcriptional level by RT-PCR analysis on micro dissected normal epithelium and lesion samples of the gland. In the metastasis, ER-β was found to be reactivated as we observed ER-β mRNA expression in prostate cancer cell lines. Recent evidence suggests that ER-β may be antiproliferative factor for a protective effect against the mitogenic activity of estrogens in breast and androgens in prostate. Activation of the receptor may exhibit cell growth inhibition. We demonstrated that antiestrogens [ICI-182,780 (ICI) and 4-hydroxytamoxifen], raloxifene and phytoestrogen (resveratrol), but not estrogens (17β-estradiol and diethylstilbestrol), inhibit growth of DU145 cells which express only ER-β while PC-3 cells with both ERs showed growth inhibition in response to estrogen and antiestrogen treatments. In DU145 cells, the ICI-induced cell growth inhibition was prevented by blockade of ER-β expression using antisense oligonucleotide. It indicated that the inhibition is mediated via ER-p associated pathway. Using flow cytometry, we found that ICI-treatment could induce accumulation of cells at GO-G1 phase of cell cycle. Similarly, this GO-G1 cell accumulation was also induced by raloxifene in DU145 cells. For resveratrol, the treatment exhibited dual effects on cell cycle distribution in DU145 cells. In the early treatment, resveratrol induced cell cycle arrests at GO-G1phase. The prolonged treatment leads to S-phase cell cycle arrest. To study the molecular mechanism of this ER-p associated cell growth inhibition, real-time RT-PCR analysis was used to semi-quantitate the transcript levels of tentative ER-β regulated genes such as telomerase reverse transcriptase (TERT), survivin and thymidylate synthase (TS) in the treated cells compared to those in control. Results demonstrated that the treatment of ICI could down-regulate TERT and survivin mRNA expressions with dose-dependent fashion. As the ICI-treatment, resveratrol downregulated expression levels of TERT, survivin and TS in DU145 cells. Down-regulation of TS may be related to the S-phase cell cycle arrest observed in the prolonged treatment of resveratrol. Taken together, our findings support the concept that ER-β participates in cell cycle regulation in normal and malignant prostatic epithelial cells. Presence of ER-β in basal cells of the prostate acini indicates that the direct actions of estrogens may be involved in the normal physiology of the gland. Loss of this receptor in primary prostate cancer and its re-expression in metastasis suggests the roles of ER-β in the cancer progression. Activation of the receptor by antiestrogen and phytoestrogen induced cell growth inhibition in prostate cancer cells. The mechanism may be mediated by reduction of cell survival factors and eventually decrease in cell viability and induction of cell cycle arrests.

Prostate cancer is the most common noncutaneous cancer among men, yet current diagnostic methods are insufficient and more reliable diagnostic markers need to be developed. The answer that can bridge this gap and enable more efficient diagnoses may lie in microRNAs. These small, single stranded RNA molecules impact protein expression at the translational level and regulate important cellular pathways. Dysregulation of these small RNA molecules can have tumorigenic effects on cells and lead to many types of cancers. Currently the Prostate-Stimulating Antigen (PSA) is used as a diagnostic marker for prostate cancer. However, many factors can elevate PSA levels such as infections and certain medications, consequently leading to false positive diagnoses and unnecessary concern and over treatment with dire outcomes for the patient. Even worse, are the chances of false negative diagnoses, which result in prostate cancer not being diagnosed until its later stages. Therefore, although the use of the PSA level has had its uses in the clinic, it has failed to sufficiently bridge the gap or to distinguish indolent from aggressive disease. It has long been suggested in the literature that microRNAs are drastically altered throughout the course of cancer progression. Here, RNA sequencing was used to identify changes in miR expression profiles diagnostic for prostate cancer patients compared to non-patient controls. The RNA sequencing results were also used to identify normalization miRs to be used as endogenous controls. Confirmatory qRT-PCR was then used to corroborate these results for the top seven dysregulated miRs found from the RNA sequencing data. Data analysis of the Area Under the Curve (AUC) of the Receiver Operating Curves (ROC) of the selected miRs exhibited a better correlation with prostate cancer (AUC Range= 0.819- 0.950) than PSA (AUC of PSA=0.667). In summary, a panel of seven miRs are proposed, many of which have prostate specific targets, which would represent a significant improvement over current testing methods.

Candida albicans is an opportunistic pathogen that is present in the normal flora in a majority of individuals. One key factor in C. albicans virulence is the ability to change its morphology from yeast to an elongated or hyphal form. The regulation of this morphogenesis relies in part upon quorum sensing (QS) molecules. C. albicans often exists as part of a mixed culture alongside other microbes and is influenced by their presence as well as the presence of QS molecules that they produce. In this study, a library of diploid homozygous transcriptional regulator knockout (TRKO) mutants were screened to identify strains capable of forming hyphae in the presence of Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli. We identified three strains that showed increased hyphae development compared to wild type C. albicans. The strains identified had deletions of the transcriptional regulating genes Orf19.3928, Orf19.2842 (GZF3), and Orf19.3865 (RFX1). These strains were tested for alterations of filamentation in liquid media, and biofilm formation. All three strains showed increased rates of biofilm formation compared to the wild type. Orf19.3928 showed altered response to farnesol, a marked in biofilm formation and no inhibition of filamentation when farnesol was present in liquid media. The GZF3 deletion strain showed enhanced filamentation with all three bacterial species while the RFX1 deletion strain showed increased filamentation only with E. coli and S. aureus. In spent media, GZF3 showed slight increases in filamentation in E. coli and S. aureus while RFX1 had moderate increases in filamentation in E. coli and S. aureus and slight increases with P. aeruginosa.