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Study of The Relationship Between 27-Hydroxycholesterol Levels and Hematopoietic Stem Cell Mobilization in Pregnant Women
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20-072-1
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Study of The Relationship Between 27-Hydroxycholesterol Levels and Hematopoietic Stem Cell Mobilization in Pregnant Women
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Dr. Fiona Campbell-Furlong
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Study Title: The Relationship Between 27-Hydroxycholesterol Levels and Hematopoietic Stem Cell Mobilization in Pregnant Women SPECIFIC AIMS The specific aims of this project are: To investigate the association between plasma 27-hydroxycholesterol (27HC) and total cholesterol levels during pregnancy progression. To investigate the association between plasma 27HC levels and mobilized hematopoietic stem cell (HSC) number during pregnancy progression. To investigate whether 27HC levels in the peripheral blood differ between pregnant and non-pregnant women. To investigate whether mobilized HSC number in the peripheral blood differ between pregnant and non-pregnant women. To investigate the association between CYP27A1 gene expression/mutation and complications during pregnancy. 4 STUDY DESIGN This prospective, single-site, observational study will enroll 50 healthy adult females in their first trimester of pregnancy and 13 adult healthy non-pregnant female controls. The purpose of this research study is to investigate the relationship between plasma 27HC levels and HSC mobilization during pregnancy. Participants will provide 6 mL of blood samples for the analyses of 27HC levels and HSC number at each research visit (single visit for non-pregnant female controls and 5 visits every 8-10 weeks for pregnant participants). Permission will also be obtained from the pregnant participants for the collection of placental and umbilical cord tissue discarded after delivery. 4.1 Characteristics of the Study Population Cohort A Number: 50 Age Range 18-40 years Sex: Females Health Status: Healthy, in their first trimester pregnancy Duration of Participation: 5 study visits every 8-10 weeks coinciding with routine prenatal visits, hospitalization for labor and delivery, and post-natal visit. First visit approximately 30 minutes in duration and the remaining 4 visits 10 minutes each. Placental tissue and umbilical cord specimens (surgical discard) will be collected following labor and delivery. Cohort B Number: 13 Age Range 18-40 years Sex: Females Health Status: Healthy, not-pregnant Duration of Participation: Single research visit, approximately 30 minutes in duration. 4.2 Sampling Plan The study sample will be drawn from healthy pregnant women and healthy non-pregnant women in the catchment area of UConn Health in Farmington, CT under the direction of Dr. Winston Campbell. 5 SELECTION AND ENROLLMENT OF PARTICIPANTS Fifty (50) healthy pregnant women and 13 healthy non-pregnant women, ages 18-40 years, from all ethnic and racial groups will be recruited in this study. Dr. Winston Campbell, the Principal Investigator of the clinical protocol, and his clinical team will recruit for this study from the patient population of UConn Health. All pregnant women presenting to the Department of Obstetrics and Gynecology at UConn Health in their first trimester for prenatal care will be considered candidates to participate in the study (Cohort A). Healthy controls (Cohort B) will be recruited from the patient population presenting in the Gynecology clinic. The UConn Health Women';s Center evaluates approximately 2,000 patients annually for routine gynecological care and the John Dempsey Hospital accounts for about 1,000 deliveries annually. Recruitment in this study will occur via direct clinician referral of the potential participant to the study team. Any recruitment material used will be presented to the IRB for approval before use. Vulnerable Populations Fifty (50) healthy adult females in their first trimester of pregnancy will be enrolled as part of this study. All five study visits will coincide with routine pre-natal and post-natal visits. Six milliliters (6 mL) of blood will be collected at each of the five visits by experienced and trained staff (a total of 30ml over 32-40 weeks). Permission will also be obtained to collect discarded placental tissue after delivery. The study imposes no greater than minimal risk to the pregnant women and fetuses. The researchers will not interfere/influence the standard of care, nor will they have any part in: 1) any decisions as to the timing, method, or procedures used to terminate a pregnancy, and 2) determining the viability of the fetus at the termination of the pregnancy. No other vulnerable populations will be enrolled in this study. Collaborating Sites/Investigators All prospective recruitment, enrollment, and clinical data abstraction will occur at UConn Health by Dr. Christine Nkemeh and the other Maternal-Fetal Medicine fellows under the direction of Dr. Winston Campbell, Professor, Dept. of Obstetrics and Gynecology, UConn Health. The blood sample collection, processing, and complete blood count, will be performed at the UConn Health - Department of Pathology & Laboratory Medicine. Plasma preparation and flow-cytometric analyses of coded blood samples will be performed at the UConn Health - Dept. of Cell Biology. Lipid panel and bile acid analyses will be performed at the UConn Metabolic Phenotyping Facility. The code key will be maintained at UConn Health with access restricted to Dr. Campbell and the UConn Health study team. Coded and deidentified plasma samples will be shared by Dr. Oguro with Dr. Jeffrey G. McDonald, Assoc. Prof., Center for Human Nutrition, University of Texas Southwestern Medical Center (UTSW) for analysis of plasma 27HC levels. 5.1 Eligibility Criteria Eligibility Criteria for Cohort A- Pregnant Females (n=50) Inclusion Criteria: Female Age 18-40 years Pregnant <12 weeks of gestation Receiving prenatal care at UConn Health Able and willing to provide written informed consent Willing to provide placental tissue following delivery Exclusion Criteria: <18 years old Hypercholesterolemia Known history of any of the following conditions: Malignancy (eligible if no recurrence in the last 5 years) Congestive Heart Failure Cardiovascular Disease (unstable ≤ 6 months) Kidney disease Renal failure Impaired hepatic function Diabetes HIV, AIDS or other Immunodeficiency Autoimmune disease such as: Rheumatoid Arthritis, Inflammatory Bowel Disease, Systemic Lupus Erythematosus, etc. Currently taking any prescribed medication more than 3 times a week for longer than 2 weeks (other than pregnancy related vitamins or supplements) Recent (≤ 3 months) trauma or surgery Current substance and/or alcohol abuse Prisoners Eligibility Criteria for Cohort B- Healthy Controls (n=13) Inclusion Criteria: Female Age 18-40 years Not Pregnant (confirmed with Urine Pregnancy Test) Able and willing to provide written informed consent Exclusion Criteria: <18 years old Hypercholesterolemia Known history of any of the following conditions: Malignancy (eligible if no recurrence in the last 5 years) Congestive Heart Failure Cardiovascular Disease (unstable ≤ 6 months*) Kidney disease Renal failure Impaired hepatic function Diabetes HIV, AIDS or other Immunodeficiency Autoimmune disease such as: Rheumatoid Arthritis, Inflammatory Bowel Disease, Systemic Lupus Erythematosus, etc. Currently taking any prescribed medication more than 3 times a week for longer than 2 weeks (other than pregnancy related vitamins or supplements) Recent (≤ 3 months) trauma or surgery Current substance and/or alcohol abuse Prisoners 5.2 Study Enrollment Procedures Cohort A participants will be recruited from the population of pregnant women presenting at Department of Obstetrics and Gynecology for prenatal care. Cohort B participants will be recruited from the patient population presenting at the general gynecology clinic. A combination of IRB approved recruitment methods may be used; including, referral by clinicians, broadcast recruitment email messages, study flyers posted in public areas, clinics, and blood draw stations at UConn Health, video displays in clinics, etc. The majority of recruitment will occur via direct clinician referral of the potential participant to the study team. Dr. Campbell, or another UConn Health Dept. of Obstetrics and Gynecology clinician, will inform patient of the possibility of participating in a research study and ask if there is interest in learning more. If the patient is interested, the clinician or a Fellow will present the details of the study and administer the informed consent. If the patient prefers a separate visit for consent, this may be scheduled at their convenience. Interested patients will provide written consent following an informed consent discussion with an authorized member of the UConn Health study team prior to initiation of study activities. Since participants in Cohort A will receive ongoing prenatal care at the UConn Health Rheumatology clinic and study visits will be coordinated to coincide with routine prenatal/postnatal care visits and hospitalization for labor and delivery, a high rate of retention in this study is expected. Since participants in Cohort B will have a single study visit, a high rate of retention in this cohort is expected. 6 STUDY PROCEDURES 6.1 Schedule of Events Participants in Cohort A will have 5 study visits 8-10 weeks apart for a total study duration of up to 11 months. The study visits will coincide with their routine prenatal and postnatal care visits as follows: Visit 1: Coincides with the first prenatal visit (5-13weeks) Visit 2: Coincides with the prenatal visit for alpha-fetal protein analysis (17-24 weeks) Visit 3: Coincides with the prenatal visit for glucose tolerance test (28-32 weeks) Visit 4: At the time of admission into John Dempsey Hospital for labor and delivery Visit 5: Coincides with 4-6 weeks post-partum visit The researchers aim to collect blood specimens during each of the three trimesters, at the time of labor and delivery, and during their postpartum visit. Participants in Cohort B will have only one study visit. Event Cohort B Cohort A Visit 1 Visit 1 Visit 2 Visit 3 Visit 4 Visit 5 Informed Consent Prior to study activities x x Screening Form x x Medical History x x Demographics x x Current Medications x x Adverse Events x x x x x x Peripheral blood specimen (6 mL) x x x x x x Medical Notes on current pregnancy (from charts) x x x x x Medical Notes on outcome of pregnancy (from charts) x x Fresh placental tissue collection (following delivery) x Umbilical cord sample collection (following delivery) x 6.2 Description of Study Visits Visit 1: Approximate visit time = 30 minutes Consent Procedure Before any study activity, the IRB-approved Informed Consent Form (ICF) will be reviewed with the potential participant, section-by-section by a qualified member of the study team in a private area. Subjects will be given the opportunity to ask questions and to have them fully answered. Subjects who elect to enroll will sign and date the consent form. The member of the study team conducting the informed consent discussion will also sign and date the ICF. A copy of the ICF signed by the consenter and the subject will be provided to the subject. This process will be documented by the Documentation of Consent Form that will be stored in the research record under a unique Participant Identifier (PID). The original signed ICF will be stored separately from the research record and with other study documents that contain personal identifiers (HIPAA Authorization, or other). The HIPAA Authorization form will be provided to the participant to review and sign to authorize the use and disclosure of their Protected Health Information (PHI) collected for use in this study. We do not expect to encounter many non-English speaking patients in this study. However, in order to facilitate any non-English speaking patients who are unexpectedly encountered in the study, a short form consent will be made available as per the UConn Human Subjects Protection Program, Informed Consent - Short Form policy. This will only be presented to individuals who have a witness (who is fluent in English and the language of the subject) present and willing to translate for the patient. An IRB-approved Short Form consent document written in a language understandable to the subject, will be presented to the patient. The person obtaining consent will provide an oral presentation of the informed consent information to the subject and a copy of the IRB-approved consent form will be provided to the subject as a written summary. If the subjects consents to participating in the study, the subject will be asked to sign and date only the short form consent. The witness will sign both the short form consent as well as the IRB-approved informed consent form. The study consenter will only sign and date the IRB-approved informed consent form. A copy of the short form document as well as the IRB-approved informed consent form will be provided to the subject. If a larger non-English speaking population is encountered during the course of the study, the complete IRB-approved ICF will be translated into the language that this population understands. Enrollment The participant will be considered enrolled and will receive a unique participant identifier number (PID) number in the study once the ICF has been signed. The PID will not be derived from or include any direct personal identifiers (name, birthdate, medical record number, etc.). Screening After informed consent, participants will be screened for eligibility. For Cohort B participants, an on-site urine pregnancy test will be used to confirm non-pregnant status. Individuals that signed informed consent but do not meet all eligibility criteria will be withdrawn from participation and considered ";screen failures";. Data Collection Once all inclusion and exclusion criteria have been reviewed and the subject has been determined to have met all eligibility criteria, the following data collection forms (CRFs) will be completed: Demographic Information Form completed with subject by interview All the other data will be abstracted from medical charts by UConn Health IRB approved study team. Clinical data to be abstracted include: date of last menstrual period, expected date of delivery, gestational age at delivery, medical conditions, and medications will be obtained from the medical record. Infant information to be obtained from the medical record will include gestational age at delivery, date of delivery, mode of delivery, birth weight, 1 minute and 5 minute APGARS, and whether or not the baby is admitted to the NICU. Blood draw Blood (6 mL) will be collected peripherally following clinical care phlebotomy and blood draw by qualified clinical staff at UConn Health using best practices. Two EDTA (lavender top) Vacutainer tubes (6 mL) will be used for whole blood collection to obtain a total of 6 mL (3 mL in each tube). The research tubes will be labeled with sample ID (see Sample Processing, Labeling and Transfer) and date of collection. Once filled with blood, tubes will be immediately inverted 8-10 times to mix and ensure adequate anticoagulation of the specimen. The tubes will then be stored at 4°C until further processing at the UConn Health - Department of Pathology & Laboratory Medicine. The first EDTA tube (3 mL blood) will be used for complete blood count at UConn Health Dept. of Obstetrics and Gynecology and flow-cytometric analyses at Dept. of Cell Biology. The second EDTA tube (3 mL blood) will be used for plasma purification at Dept. of Cell Biology. The EDTA tubes will be stored on ice and securely transported to Dept. of Cell Biology for processing and analysis. Triple packaging consisting of a leak-proof primary receptacle (eg. blood tube), a leak-proof secondary package (eg. Sealable plastic bag), and an outer package clearly labeled with Biohazard sign will be used for secure transportation of the samples from Dept. of Obstetrics and Gynecology to Dept. of Cell Biology. At Dept. of Cell Biology, the plasma samples will be divided into 3 parts, labelled with sample ID (see Sample Processing, Labeling and Transfer) and date of collection. One part of the plasma sample will be shipped to UTSW (0.5 mL of plasma) for the analysis of plasma 27HC levels. Visits 2-5: Cohort A only - Approximate visit time = 10 minutes each Blood draw Blood (6 mL) will be collected peripherally by phlebotomy by qualified clinical staff at UConn Health using best practices at each study visit as detailed above. The blood draws at Visits 2, 3, and 4 will occur at the time of phlebotomy and blood draws performed for clinical care. Labelling, storage, processing, analysis, and transportation of samples to Dept. of Cell Biology will be as detailed above. Placental Tissue Collection Following delivery at the UConn John Dempsey Hospital, several samples of the placental tissue will be collected from the expelled placenta (to be discarded otherwise). Specifically, three 0.5cm3 blocks of placental tissue will be excised from the maternal side of the placenta. Collected specimens will be washed with sterile saline and placed into separate pre-labelled tubes (see Sample Processing, Labeling, and Transfer section) containing RNA stabilization reagent, DNA stabilization reagent, and fixative agent. All three pre-labelled tubes containing the reagents will be provided by Dept. of Cell Biology. The tubes will be stored on ice and securely transferred to Dept. of Cell Biology for processing and analysis. Umbilical Cord Sample Collection Following delivery of the placenta, a 1cm segment of the umbilical cord closest to the placental cord insertion site will be collected. This segment will be drained of all blood and then washed with saline. Collected specimen will be cut transversally into three pieces and placed into pre-labelled tubes (see Sample Processing, Labeling, and Transfersection) containing RNA stabilization reagent, DNA stabilization reagent, and fixative agent. All three pre-labelled tubes containing the reagents will be provided by Dept. of Cell Biology. The tubes will be stored on ice and securely transferred to Dept. of Cell Biology for processing and analysis. 6.3 Sample Processing, Labeling, and Transfer All specimens will be processed according to SOPs developed by the Dept. of Cell Biology team. Laminated copies of SOPs as well as labeling instructions will be provided to the UConn Health team prior to study initiation. All specimen tubes will be labeled with the sample ID and date of collection. The sample ID will be in the format 27HC-XXC-V#-S# where: 27HC is the study ID, XX is the two-digit sequential enrollment number and C is the cohort ";A"; or ";B";, V# is the visit number for blood samples (1, 2, 3, 4, or 5), S is the specimen type, and # is the tube number when multiple tubes of blood/placental tissue are collected (e.g., 27HC-01A-V1-B2, 27HC-01A-PM1, 27HC-04B-V1-P3). The Specimen type and tube number key is provided below: Specimen Type Description Tube Number B Blood- EDTA Tube 1, 2 PM Placenta-Maternal Side 1-3 U Umbilical cord 1-3 P Plasma 1-3 *Plasma separated from the blood in EDTA tube #2 The specimens will be prepared for transport using DOT/IATA compliant triple packaging consisting of a leak-proof primary receptacle (e.g. a closed tube), a leak-proof secondary package (e.g. sealable plastic bag), and an outer package clearly labeled with Biohazard sign (e.g. cardboard box or cooler). 6.4 Compensation There are no costs to study participants for taking part in the study. Participants in Cohort A will be provided a modest honorarium (total $20 for five visits: $10 gift cards at the end of visit 1 and $10 at visit 5). Participants in Cohort B will be provided a modest honorarium of $5 gift card. The gift cards will be presented directly to the participants by the study coordinator/fellow and the receipt will be documented. 6.5 Blood cell analysis Three (3) mL of fresh blood sample collected from participants (of the 6 mL) at each visit will be used for flow-cytometric analysis at Dept. of Cell Biology. Complete blood count will be performed at UConn Health either specifically for this study or as part of clinical care. The flow-cytometric analysis at Dept. of Cell Biology will evaluate the expression of CD34 and CD38 to obtain the frequency of HSCs that have CD34+CD38- cell-surface maker phenotype. 6.6 Plasma analysis The remaining three (3) mL of fresh blood samples collected from participants (of the 6 mL) will be processed at Dept. of Cell Biology for plasma purification. Purified plasma (~1.5 mL) will be divided into 3 tubes as follows: Tube #1: 500 uL for 27HC analysis (UTSW) Tube #2: 100 uL for lipid panel (UConn Metabolic Phenotyping Facility) Tube #3: Remaining volume (~900 uL) for storage for the backup (Dept. of Cell Biology) 6.7 Placental and umbilical cord tissue analysis The samples will be collected from 3 different loci from the maternal side of the placental tissue (a total of 3 pieces per tissue approximately 0.5 cm3 each) from pregnant women immediately after delivery at UConn John Dempsey Hospital. A 1cm section of umbilical cord will also be collected and cut transversely into three pieces. Specimens from the placenta and umbilical cord (one each) will be analyzed at Dept. of Cell Biology as follows: Piece #1: Place into a tube containing RNA stabilization reagent for mRNA expression analysis (mRNA expression analysis of CYP27A1 and other genes or RNA-sequencing at Dept. of Cell Biology) Piece #2: Place into a tube containing DNA stabilization reagent for genomic DNA mutation analysis (analysis of genomic mutations in CYP27A1 gene and other genes or whole genome sequencing at Dept. of Cell Biology) Piece #3: Place into a tube containing fixative reagent for immunohistochemistry (expression analysis of CYP27A1protein and other proteins at Dept. of Cell Biology) 7 RISKS AND PROTECTIONS 7.1 Potential Risks to Subjects Risk to Confidentiality: There is a potential risk to confidentiality due to the PHI that will be collected and stored in the subject';s research record. Risk from Blood draw: There may be a minor amount of discomfort due to the needle stick performed for the study blood draw. There is a minor risk of bruising (< 1%), infection at the phlebotomy site (< 1%) or dizziness following the blood draw (<1%). There is no additional risk to the participants from the additional low volume (6ml per visit) of blood drawn for this research. No more than 30ml of blood will be collected from the pregnant participants for purpose of this study over the approximately 40-week study period. No more than 6ml of blood will be collected from the participants in Cohort B. Risk from Placental and Umbilical cord specimens collection: The specimens collected for this study are placental and umbilical cord tissue that would otherwise be discarded. There is no additional risk to the participants in Cohort A or their new-born from the specimens collected for this research. 7.2 Adequacy of Protection against Risks The human subject components of this study will be conducted under the supervision of Dr. Campbell at UConn Health. UConn Health emergency care procedures will be followed if an adverse event or medical emergency occurs. The study site is located on the campus of a tertiary care hospital that is available for treatment of medical emergencies. Protection against Risk to Confidentiality All study visits will occur in a private area at UConn Health in Farmington, CT. Research records will be labeled with a participant ID (PID), an assigned unique identifier that is not derived from any patient identifiers. All contents of the research record will be labeled with the assigned PID. Research records will be stored in a secure area. A complete record of the subject's pertinent history and documentation of the visit will be kept on case report forms and will be stored in the research records. Research records will be accessible only to the IRB-approved UConn Health study team directly involved in conduct of the clinical protocol. Any study documents (Informed Consent Form, HIPAA Authorization) that contain the participant's name will be kept in a separate file apart from the research record and will be stored in a secure location. A master key that links participant names and PIDs will be maintained in a separate and secure location accessible only to the Site PI and approved UConn Health study team. At no time will this key or any identifiable information be provided to researchers at UConn Health Dept. of Cell Biology or UTSW. Samples will be labeled as per SOPs (see Sample Processing, Labeling, and Transfer) and will not contain any patient identifiers when they are transferred to Dept. of Cell Biology. Coded clinical data linked with study samples by the PID will be provided to the Dept. of Cell Biology study team for analysis. HIPAA personal identifiers will not be included in the clinical dataset/samples provided to Dept. of Cell Biology for use in analysis. The clinical data will be provided to Dept. of Cell Biology by secure file transfer or by the entering the data into a password-protected excel database. Dr. Oguro, as an investigator on the clinical protocol may be present at clinical team meetings when identifiable information is present. He will not record or disclose participant identifiers and will not receive the code key at any time. Publications and presentations about discoveries from the data generated from samples collected in this study will not identify subjects by name. Protection against Risk from Blood Draw Six (6) mL of blood will be drawn for participants at each research visit (five visits for Cohort A participants and one visit for Cohort B participants). Blood will be drawn by experienced, trained research staff in a clinic setting on a hospital campus. The site where the needle will be inserted will be wiped with alcohol before and after the draw. Only sterile needles will be used. An adhesive bandage will be placed over the site after the draw. Emergency treatment will be accessible on campus for any severe complications from blood draw. For participants in Cohort A, blood draws at visits 1, 2, 3, and 4 will follow phlebotomy and blood draws that are part of clinical care. Protection against Risk from Placental Specimen Collection Placental specimens will be collected following delivery by experienced, trained staff in appropriate settings. 7.3 Data and Safety Monitoring Data and Safety Monitoring Plan (DSMP) The Data & Safety Monitoring Plan for this study describes the components of the study that will be monitored by the study coordinators and consenters once annually. Recruitment, drop outs, unanticipated problems, data integrity and confidentiality, participant privacy and the general conduct of the study will be reviewed. Minutes of the annual DSMP review will be kept in the regulatory binder and provided to IRB at study continuation. Procedures in place to ensure confidentiality of research data are as follows: Only authorized individuals have access to any data, used or stored (via electronic format or as hard copy records). Only designated research staff and investigators will be granted access. All data collected on data forms are stored in a secure records room located in the Principle Investigator';s office at UConn Dept. of Ob/Gyn. CRF data and data abstracted from EMR will be entered into a excel database labeled only by the PID and the resulting dataset shared with Dr. Oguro at Dept. of Cell Biology. 8 BENEFIT Participants will not directly benefit from the information gathered in the study but other people may benefit in the future. Investigators in this study will gain insight into the mechanism of HSC mobilization. It is possible that findings from this study could lead to research projects to understand the mechanism of HSC mobilization during pregnancy as well as the relationship between gene mutation and complications, to prevent anemia during pregnancy, and to improve HSC mobilization methods for transplantation. There is also the possibility that no benefit will come from this study. 9 INTEGRATIVE DATA ANALYSIS 9.1 Power Calculations This study is part of a powered study with two study sites: UConn Health and Chinese University of Hong Kong (CUHK). To determine approximate sample sizes for this study, a literature review was performed to identify estimates of effect size and variability that could be extrapolated to this research study. All sample size calculations are based on two-sided tests with alpha=0.05, with a goal of achieving 80% power to detect the association of interest. For example, linear regression results from two publications were used to inform sample size estimates for Specific Aims 1 and 2. Karuna et al. reported a significant association between 27HC and total cholesterol in healthy controls (beta coefficient=0.08, r= 0.54); based on these estimates, a sample size of 26 subjects would provide 80% power to detect an association of this magnitude [12]. For Aim 2, examining the association between 27HC and HSC number, we used data provided by Cimato et al. who found a significant relationship between LDL cholesterol and CD34+ HSPC in human subjects taking statins (beta coefficient=175, R-squared = 0.09573), and we hypothesize a similar relationship will be identified between total cholesterol and 27HC by the third trimester of pregnancy [11]. Using these estimates, a sample size of 82 individuals provides 80% power to detect such an association. Related to Aim 3, Ordovas et al. demonstrated a 49% increase in total cholesterol during pregnancy (mean 182 ± 35 ml/dL in the non-pregnant group and 265 ± 56 ml/dL by 37 weeks'; gestation) [19]. To detect such a large difference in means using a t-test, only 9 subjects are needed in each group (pregnant and non-pregnant). For Aim 4, Gao et al. measured hematopoietic stem/progenitor cells in the peripheral blood of human subjects, and found significantly increased levels in individuals with low compared to normal HDL Assuming similar variability in our sample of healthy women, we expect to see a difference in mean HSC levels of at least a similar magnitude (difference in means=11, standard deviation=15) between pregnant women in the third trimester and non-pregnant women [20]. With 82 pregnant women, 25 non-pregnant women are needed to have 80% power to detect this difference. Overall, sample sizes of 82 pregnant women and 26 non-pregnant women should provide sufficient power to investigate each of the aims. Accounting for expected dropouts, the study proposes to enroll 100 pregnant women and 26 non-pregnant women. Participants will be recruited at two study sites as follows: UConn Health: 50 pregnant women and 13 non-pregnant women CUHK: 50 pregnant women and 13 non-pregnant women The clinical study protocol at both sites will be reviewed by the respective institutional IRBs. 9.2 Statistical Analyses In addition to the hypothesis testing described above, descriptive analyses (mean, standard deviation, median and quartiles) will be performed to summarize 27HC, total cholesterol, and HSC number levels at each time point, and trajectories of their values over pregnancy progression will be plotted. The demographic characteristics of each cohort will be described. 10 DATA COLLECTION AND MANAGEMENT Data Collected from Human Subjects Specifically for Research Purposes Demographic information, medical history, date of last menstrual period, and current medication use will be provided by participants prior to the collection of blood at the research visit 1. Scanned copy of the CRFs redacted of PHI will be shared with the research team at Dept. of Cell Biology. The following medical data about current pregnancy (Cohort A only) will be abstracted from medical records into the study CRF: Gravida, Para, Abortion Date of Last Menstrual Period Expected Date of Delivery Gestational age Singleton pregnancy/Multiple pregnancy Ultrasound date and abnormal findings, if any Presence of any comorbidities (Preeclampsia, gestational diabetes, etc) Pregnancy Outcome- gestational age at delivery, date of delivery, mode of delivery Infant information- birth weight, 1 minute and 5 minute APGARS, and whether or not the baby is admitted to the NICU Participant Identification (PID) number A unique participant identifier (PID) not derived from any patient identifiers will be assigned to each participant once written informed consent for study participation has been obtained. The PID will consist of the study ID, ";27HC";, followed by two-digit sequential enrollment number and cohort (A-pregnant or B-non-pregnant); for e.g. 27HC-01A. Participant names or other personal identifiers will not be included in the study data set; all data will be identified only by PID. Access to individually identifiable private information about human subjects Access to identifiable study data will be restricted to the UConn Health study team involved in the conduct of the clinical protocol. Research records containing identifiable private information will be stored at the UConn Health and will only be accessed by IRB approved research coordinators/fellows. Coded clinical data will be provided to the Dept. of Cell Biology study team for analysis linked with study samples by the PID. The key linking the PID to participant identifiers will be stored separately and securely from the research records at the UConn Health Dept. of Obstetrics and Gynecology and will be accessible only to Site PI. The code key will not be provided to the Dept. of Cell Biology investigator or staff at any time. The Dept. of Cell Biology study team will have access to coded samples and data only. Dr. Oguro, as the Co-Investigator of this clinical protocol may be present at clinical team meetings when identifiable information is present. He will not record or disclose participant identifiers and he will not receive the code key at any time. Upon accessioning at Dept. of Cell Biology, the samples will be recoded and deidentified before being transferred to UTSW for 27HC analysis. Dr. Oguro will maintain the code key linking the UConn study PID and the new code assigned to samples shipped to UTSW for analysis. No genomic data generated in this study will be added to the participant';s medical record or returned to the UConn Health Dept. of Obstetrics and Gynecology study team. 10.1 Data Collection Forms (CRF) Forms to be labeled with the PID and completed for this study include: The Visit 1 Source Documentation Form The Visit 1 Source Documentation Form is labeled with the PID and includes the following data: Screening Form - This form lists all eligibility criteria. Participants must meet all inclusion criteria and not have any of the exclusion criteria to be enrolled into the study. Demographic Information Form-To be completed by participant Medical History and Medication Log Forms- Abstracted from EMR Clinical data abstracted from EMR Visits 2-5 (Cohort A only): Source Documentation Forms: The follow-up visit forms will document blood draws and the clinical data abstracted from EMR. Placental and umbilical specimens collection will also be documented. 10.2 Data Management Data Collection Forms (CRFs) will be labeled with the PID code and Visit Date and will be stored in the Research Record and/or electronically within the excel database. Paper CRF';s will be stored in a secure location in in the Principal Investigator';s office at the Obstetrics and Gynecology Clinic. Scans of completed CRFs, labelled only with PID, will be shared with Dept. of Cell Biology. Data from these scans will be entered into a password-protected excel database. Records will only be labeled with the PID and will not contain subject initials or any other identifier. 10.3 Quality Assurance Training A study initiation meeting will be attended by study staff in which the protocol, visit procedures, secure storage of research records, secure management of electronic databases and sharing of coded data with Dept. of Cell Biology, sample labeling protocol, and the process for sample transfer to Dept. of Cell Biology and UTSW will be reviewed. Delegation of Responsibilities A Delegation of Responsibilities log will be completed at the Initiation Visit for members of the CUHK study team that will be obtaining informed consent entering data on CRF, determining eligibility and labeling samples. This log will be kept in the study regulatory binder. 10.4 Protocol Deviations A cumulative Protocol Deviation Log will be kept electronically by the study coordinator with a copy added to the regulatory binder and provided to IRB at the next study continuation or closure submission, whichever occurs first. Deviations from the protocol will be entered on this log with a Note to File labeled with the PID and no personal identifiers added to the regulatory binder with a copy to the research record that describes the deviation, date when it was identified, the corrective action taken to prevent recurrence, whether or not the deviation met criteria of an Unanticipated Problem, and the date of IRB notification. Any unauthorized access to the database linking the research records to participant direct identifiers shall be reported to the IRB at the time that it is discovered as well as at annual review. Incidents of non-compliance, defined as any action that is taken or occurs that is not in accordance with an IRB approved study, IRB policies or regulations or failure to follow the requirements and determinations of the IRB, that is within the control of the study team must be reported to the IRB within 5 days of becoming aware of the occurrence. 10.5 Unanticipated Problems For the purpose of reporting Unanticipated Problems to IRB, internal adverse events that may also represent an unanticipated problem are defined as those events, experiences or outcomes that are: Unexpected (in terms of nature, severity or frequency) given (a) the research procedures that are described in the protocol-related documents, such as the IRB approved research protocol and informed consent document; and (b) the characteristics of the subject population being studied; and Related or possibly related to participation in the research (i.e., there is a reasonable possibility that the incident, experience, or outcome may have been caused by the procedures involved in the research). Any internal event meeting these criteria must be reported to the IRB, which will then make the final determination as to whether the research places subjects or others at a greater risk of harm (including physical, psychological, economic or social) than was previously recognized. 11 PARTICIPANT RIGHTS AND CONFIDENTIALITY 11.1 Institutional Review Board (IRB) Review This protocol, the recruitment material, the informed consent form, study CRFs, and any subsequent modifications will be reviewed and approved by the UConn Health IRB before use. 11.2 Informed Consent The informed consent process will begin after an individual confirms interest in participating in the study. A copy of the IRB approved Informed Consent Form (ICF) will be provided to the subject in advance for review, whenever possible. At the research visit, a study clinician will review the ICF, section by section, with the subject in a private room prior to any study procedures. Questions from the participant will be encouraged and will be fully answered. Once the form has been fully reviewed and all questions answered, the volunteer will be asked if they would like to participate in the study. If they agree to participate, they will be asked to sign and date the ICF indicating their consent. The consenter member of the study team obtaining consent will sign the ICF as well and a copy of the form signed by both the subject and the consenter will be provided to the subject. The Informed Consent process will be documented by the consenter on the Documentation of Informed Consent Form or as a note in the subject';s Medical record. Original signed and dated ICFs will be stored separately and securely away from the research record with other identifiable documents that contain participant identifiers. 11.3 Genomic Data Sharing Participants will provide consent within the ICF for sharing of genomic data generated from their samples in this study in publicly available online scientific databases in the future. Before this genomic data is shared, PIDs will be replaced with new codes and a link will not be kept between the data and participant identity. Even if a participant wants to withdraw from the study, genomic data once shared will not be withdrawn since there will be no way to link data with the subject. Subjects who decline consent for sharing of re-coded genomic data in public access databases will be enrolled in this study, however, their genomic data will not be shared in public access databases. Any samples remaining after study analysis has been completed will be kept in storage at Dept. of Cell Biology and will be given a new unique code. This random code will link the study information to the remaining sample. These recoded samples may be shared with other researchers and/or used in other projects. Subjects will provide or decline consent within ICF for sharing of samples for future research use. Subjects who grant permission for sharing of samples for future research use will not be contacted when the samples are used. 11.4 Withdrawal Subjects may withdraw their permission to participate in this study at any time by contacting: Dr. Winston Campbell Dept. of Obstetrics and Gynecology, UConn Health, 263 Farmington Avenue, Farmington, CT 06030 Phone: 860-679-7605, Email: wcampbell@uchc.edu Dr. Campbell will inform Dr. Oguro of the PID of the sample to be removed, but will not provide the subject';s name. The Dept. of Cell Biology team will locate and destroy/delete all specimens/data, linked to the subject requesting to be withdrawn. Any samples that were shared with UTSW before the request for withdrawal will be retrieved and destroyed. Any samples that were recoded (without a link to the PID) and recoded sequencing data shared in public access databases will not be withdrawn as there will be no way to link the recoded data/samples with the participant. 11.5 HIPAA Authorization Study participants must provide written authorization at study entry for use and disclosure of their Protected Health Information that is recorded and used in this study in order to participate. Participants may withdraw their Authorization at any time, but will then be withdrawn from the study. HIPAA Authorization will include access of UConn Maternal-Fetal Medicine fellows to identified data in the EMR for abstraction into the study CRF and database. Study data will be stored in a password-protected database coded by Participant ID (PID). The key linking PID and participant name will not be shared with Dr. Oguro and Dept. of Cell Biology team performing laboratory analysis. As a co-investigator, Dr. Oguro will be actively engaged in the conduct of this research and although he will not be provided with the key linking PID and identity of participants, he may have identifiable information disclosed to him during meetings with the clinical study team. Dr. Oguro will be listed in the HIPAA Authorization form for this reason.
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Obstetrics and Gynecology
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Check with study contact
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Makayla Murphy. Telephone: 860-679-3331. Email: makmurphy@uchc.edu or Kristen Annis. Telephone: 860-679-3812. Email: annis@uchc.edu or Christine Nkemeh. Telephone: 860-679-2656. Email: nkemeh@uchc.edu
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Enrolling/recruiting. For current recruitment status, please check with study contact.
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