The Mood, Mother and Child Study: Protocol for a Prospective Longitudinal Study and Randomized Controlled Trial

Background Perinatal depression affects >400,000 mother-child dyads in the United States every year and is associated with numerous adverse maternal and child developmental outcomes. Previous research implicates the dysregulation of oxytocin and the hypothalamic-pituitary-adrenal (HPA) axis functioning in mothers and children as potential mechanisms mediating or moderating the transmission of risk associated with maternal depression. Objective The Mood, Mother and Child study will examine the psychobiological sources of risk and resilience within mother-child dyads affected by maternal depression. This manuscript describes (1) the study rationale and aims, (2) the research design and procedures and how they were altered in response to the COVID-19 pandemic, and (3) the data analysis plan to test the study hypotheses. Methods This is a prospective longitudinal study with an embedded randomized controlled trial that examines (1) correlations among postpartum depression and anxiety symptoms, maternal and child oxytocin and HPA axis functioning, and child developmental outcomes and (2) the causal relationship between exogenous oxytocin and HPA reactivity. This study is funded by the National Institute of Child Health and Human Development with institutional review board approval. Results Recruitment and data collection have commenced, and the expected results will be available in 2024. Analyses are presented for testing the proposed hypotheses. Conclusions The unique combination of a prospective longitudinal research design with an embedded randomized controlled trial will allow the Mood, Mother and Child study to apply a developmental lens to the study of maternal depression and anxiety symptoms from birth to middle childhood and the psychobiological mechanisms promoting risk and resiliency for both mother and child outcomes. This will be the first study that simultaneously evaluates (1) the role of oxytocin using multiple methodologies, (2) the causal relationships between exogenous oxytocin and HPA axis functioning among mothers with differing levels of depression and anxiety symptoms, and (3) the multiple mediating and moderating roles of parenting behaviors and maternal and child psychobiological characteristics. The goals of these aims are to provide insights into the psychobiological effects of oxytocin in women and inform future clinical trials to treat perinatal mood disorders. Trial Registration ClinicalTrials.gov NCT03593473; https://classic.clinicaltrials.gov/ct2/show/NCT03593473 International Registered Report Identifier (IRRID) DERR1-10.2196/51132


PUBLIC HEALTH RELEVANCE:
The proposed research is relevant to public health because it will identify the psychobiological underpinnings of resilience among mother-child dyads exposed to maternal depression, a common and morbid condition that impacts the health of more than 400,000 mother-infant dyads every year.The project is relevant to NICHD's mission because it will identify developmental factors associated with psychosocial adjustment among children exposed to maternal depression, ensuring that all children have the chance to achieve their full potential for healthy and productive lives.

CRITIQUE 1
Significance: 3 Investigator(s): 1 Innovation: 4 Approach: 4 Environment: 2 Overall Impact: The proposed study will build on an existing cohort of mother-infant dyads to track the relationship between maternal depression and infant outcomes, and identify the moderating and mediating factors that contribute to infant risk and resilience.The premise of the project is strong and the methodology is rigorous in most cases.Strengths of the application include a multileveled psychobiological approach, excellent statistical methods, and an innovative intransal oxytocin (OT) trial to probe HPA and OT system function during stress in mothers.Major limitations are largely centered on insufficient detail regarding genomics and epigenomics methods and interpretation.Whole genome approaches are proposed, although only a few genes will be interrogated.Buccal cell DNA methylation is proposed as potential mediating mechanism of infant outcomes, but can only model, not imitate neuronal epigenomics.Minor limitations that nonetheless diminish impact include inadequate consideration of maternal partner support as a confound of dyad characteristics, and remaining concerns of the role of gene-environment correlations on results.

Strengths
 This is a highly significant area of research with great potential for identifying some of the neuroendocrine and behavioral mechanisms for the impact of maternal depression on child development.

Weaknesses
 The significance of the genomic aim is unclear.The mismatch between methodology and proposed analyses (i.e., whole genome approaches are proposed but analyses will only include a few genes) diminishes impact.The use of proxy tissue for neural development is necessary in human epigenetic studies, but more consideration must be given to the limits of interpretation of data collected from proxy tissue and impact on results.

Investigator(s):
Strengths  This is a superb team of investigators, well positioned to carry out the majority of procedures proposed. The Principal Investigators are established investigators, with many publications in the areas of oxytocin, maternal psychopathology, and child development. An advisory team of senior investigators has been assembled, which enhances confidence of project success. An experienced postdoctoral fellow with five publications in epigenetics and bioinformatics and statistical geneticist to be named will conduct genomics statistical analyses.

Weaknesses
 No limitations noted.

Strengths
 The use of intranasal oxytocin to probe HPA and OT function in mothers is highly innovative.

Weaknesses
 The genetic and epigenetic approaches proposed do not maximize potential innovation.

Approach:
Strengths  The sequalae of potential influences of maternal depression on child mental health are wellconsidered and operationalized.
 The use of multileveled measures of oxytocin function as potential moderators of mother-child relationships is excellent.
 The consideration of sex differences is a major strength.

Weaknesses
 Whole genome and epigenome methods do not seem warranted given the analyses focus on one system.The justification for buccal cell as a model of neurons is not strong and there is insufficient conceptualization of how data from proxy tissue will be interpreted.
 Gene-environment correlations still present a significant potential confound.The proposed method for addressing potential gene-environment correlations, using infant temperament as a proxy for genetic influences, is not sufficiently justified.
 A minor concern is that although the population is predominantly married women, there is no consideration in the methods or statistical analysis of a potential confound of maternal characteristics with availability or quality of partner support.
 Another minor concern is that resilience is defined as "better than expected" outcomes, but it is unclear how this will be statistically or conceptually defined within the population.

Strengths
 This is a superb environment for the proposed studies, and feasibility is enhanced by the fact that this location has housed prior projects which provide the basis for the proposed work as well as many other collaborative and related studies.

Weaknesses
 The location and equipment for proposed sequencing and array work is not adequately described.Overall Impact: This is a well-prepared and documented application that does a good job of clarifying and addressing concerns raised in the prior reviews.The project leverages a comprehensive earlier project with repeated visits of over 200 maternal-infant pairs, oversampled for mood disorder, that characterized the individuals and the dyad on multiple dimensions.This 5-year follow-up extends the focus on oxytocin as the principal linchpin in dyadic development and, through affective, behavioral, epigenetic and related neurohormonal systems, might translate the disruption imposed by maternal mood disorders to dysregulated child development.The research team is excellent and has already conducted high caliber work which forms the substrate for this project.Figure 1 provides a clear visual description of the conceptual model and the manner in which each of the three aims feeds into it.The revision also considers the role of sex as a biological variable and although the existing literature yields contradictory findings regarding differential vulnerability to maternal mood disorders by child sex, preliminary data provided from the parent study suggest that child sex does indeed modify associations between trajectories of maternal mood disorders and outcomes, as does variation in parenting behaviors.The revised application will explicitly test for sex effects, including in epigenetic markers.A prior concern regarding sociodemographic characteristics of the sample is discussed in Approach below, but the investigators also note that a follow-up study is planned for an existing, more diverse, cohort.The prior review noted that relatively few studies based on the parent project have been published to date.The investigators' response, that they are awaiting conclusion of data collection, reveals the double-edged sword that is inherent in the management of complex longitudinal studies in which each individual element (the still-face paradigm, for example) requires large amounts of processing time before data can even be input into a statistical database.Couple this with a neverending stream of incoming new data, each source with its own challenges, and the problem becomes exponential.Thus although the goal of avoiding piecemeal publication is laudable, inadequate identification of stand-alone questions that can be answered by already collected data can lessen the impact of this rigorously executed and important work.

Strengths
 In recent years, the role of the "4 th trimester" has taken a back seat to research on purported fetal programming via biological mediation during pregnancy.However, postnatal depression translated through effects on maternal-infant interaction and other social factors has clear sequelae for development that is often overlooked despite accrued evidence since the 1980s.The proposed study will reinvigorate this important avenue of inquiry through the examination of the role of oxytocin as the driver of dyadic success in relation to maternal mood disorders using modern technologies and current understanding of underlying processes.
 A child's developmental repertoire at age 5 is greatly expanded beyond that reportable at the last observation period at 12 months and is also more predictive of important subsequent outcomes.This allows testing on more meaningful outcome parameters while leveraging with the original study will allow reach-back to the peripartum period.

Weaknesses
 The influence of perinatal depression on children is repeatedly described as "devastating" which is a bit of hyperbole.Although it can cast a wide net on offspring outcomes, many children manage to not be significantly affected which is in fact a point of this work -to document the factors that underlie resilience and evaluate the continuum of outcomes.

Strengths
 The Co-PIs reflect expertise in peripartum women and mood disorders (Steube) and developmental psychology within family systems (Mills-Koonce) and have previous experience working together on the existing R01 which forms the substrate for this project.Studies on the topic often are driven by expertise in one domain or the other; here the blended approach shows through the rigor with which both elements are represented in the existing and proposed work.
 The roles of a number of investigators listed are described as part of a "senior advisory team" without appreciable effort.Although this might be of concern in a new collaboration in a new field, the remarkable success of the prior study alleviates this as a concern and instead reveals the vibrant intellectual community within which this research is taking place.

Weaknesses
 No limitations were noted in the prior review and none are noted here.

Strengths
 The study will examine both the predictive and the protective factors that link PND to outcomes.
Inclusion of the latter is often overlooked but it is key to developing successful interventions that can mitigate deleterious effects.
 The oxytocin challenge is a unique feature of the project because it is believed to be the first to evaluate this response in women with older children within this research framework and few psychobiological studies that employ exogenous oxytocin delivery in relation to a stressor challenge designed to activate the HPA axis have included women at all.

Weaknesses
 None noted.

Strengths
 Although the study protocol imposes a high participant burden, participating women are described as identifying with the project's goals and enjoy the process.This resonates with those who implement longitudinal studies in this population and recognize that it suggests that the research team interacts with participants in a respectful fashion as partners.Retention/tracking is boosted by contacts in 6 month intervals for questionnaire completion.That the follow-up protocol for the proposed study is described as less intensive than the original further alleviates this concern.
 A prior concern related to the relatively homogenous nature of the sample population, particularly in terms of education level.Consistent with the investigators' response, unless there is reason to suspect that the underlying biobehavioral processes differ by race/SES as opposed to being moderated by contextual factors, this does not necessarily affect generalizability of the research question and eliminates the need to control for socioeconomic and related factors that can overwhelm the ability to detect associations.Practicalities of focusing on more educated women include higher rates of and more protracted duration of breastfeeding and higher participation in follow-up studies which contribute to study success.
 In response to the prior review, preliminary data showing differential findings by child sex have been presented and the consideration of child sex as a biological and social factor has been integrated into the current study. The inclusion of child temperament at the 5 year visit, and its integration into the analytic plan, is worthwhile.
 The rigor of the design allows evaluation of the role of oxytocin in terms of tonic, observational associations (both longitudinal and cross-sectional), phasic effects (response to stressor), a challenge model (placebo v oxytocin administration) as well as genetic/epigenetic variation.This is truly a remarkable undertaking.
 The protocols for women and children are comprehensive and reflect the state of the field but should not be overwhelming.The team is experienced in testing children.The physiological data collection systems afforded by MindWare provide strong measures of heart rate variability and PEP, yielding indications of parasympathetic and sympathetic activity.

Weaknesses
 Research on dyadic human systems is inherently complex.As a result, the investigators run the risk of generating too much complicated data without sufficient capacity to disseminate it in peer 1 R01 HD093901-01A1 8 MESH STUEBE, A reviewed journals, as opposed to abstracts.This can limit the impact of even the most wellsupported premise and rigorously constructed research plan.

Environment:
Strengths  UNC at both Chapel Hill and Greensboro have long-standing reputations for high quality research in all of the domains represented in this application.This includes a wealth of riches in development psychology at both campuses.
 The multidimensional nature of the project leverages significant infrastructure including a fully equipped laboratory for implementing experimental protocols and evaluating maternal and child behavioral and psychophysiological functioning.Based on the parent study that determines eligibility, the expected distribution by race/ethnicity is 79.5 percent white, 13 percent African American, 5 percent Asian; approximately 10 percent is of Hispanic ethnicity.

Vertebrate Animals:
Not Applicable (No Vertebrate Animals)

Biohazards:
Not Applicable (No Biohazards) Resubmission:  The application has done a thorough job of responding to the prior reviews.Features of the responses are integrated into the overall impact paragraph as well as other segments of this review.Overall Impact: This project is addressing an important public health issue.It is a continuation and extension of a project focused on maternal depression and anxiety trajectories (MDAT) across the first 5 years of the child's life.The research team is very strong.They are collecting a very extensive set of mediating and moderating variables for their complex model.Their basic plan is to continue the follow up from their existing R01 on perinatal effects on infant outcomes at age 1 to age 5 years.The proposed study would continue following the cohort online at a 6 month intervals until the child's age of 5 years.The study burden appears intense, covering psychosocial, psychiatric, biological and genetic assessments.To date their follow up response rate has been very good suggesting that their continuing response rate should be reasonable.Their response to previous reviews is appropriate.

INCLUSION OF MINORITIES PLAN: ACCEPTABLE
The expected distribution by race/ethnicity is 79.5 percent white, 13 percent African American, 5 percent Asian; approximately 10 percent is of Hispanic ethnicity; this is scientifically justified.

INCLUSION OF CHILDREN PLAN: ACCEPTABLE
The study will enroll children from infancy to five-years-old; this is scientifically justified.

COMMITTEE BUDGET RECOMMENDATIONS:
The budget was recommended as requested.
Footnotes for 1 R01 HD093901-01A1; PI Name: Stuebe, Alison M NIH has modified its policy regarding the receipt of resubmissions (amended applications).See Guide Notice NOT-OD-14-074 at http://grants.nih.gov/grants/guide/notice-files/NOT-OD-14-074.html.The impact/priority score is calculated after discussion of an application by averaging the overall scores (1-9) given by all voting reviewers on the committee and multiplying by 10.The criterion scores are submitted prior to the meeting by the individual reviewers assigned to an application, and are not discussed specifically at the review meeting or calculated into the overall impact score.Some applications also receive a percentile ranking.For details on the review process, see http://grants.nih.gov/grants/peer_review_process.htm#scoring.

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Qualified specialized services, including the Institutional Drug Service (IDS) for supporting the OT challenge, and the Biospecimen core for sample processing, are already in place and necessary for successful completion of the project Weaknesses  None noted Protections for Human Subjects: Acceptable Risks and/or Adequate Protections  The study team is experienced in working with populations of women with mood disorders and their children.The plan as stated is comprehensive and appropriate.Data and Safety Monitoring Plan (Applicable for Clinical Trials Only): appropriate Inclusion of Women, Minorities and Children:  Sex/Gender: Distribution justified scientifically  Race/Ethnicity: Distribution justified scientifically  For NIH-Defined Phase III trials, Plans for valid design and analysis:  Inclusion/Exclusion of Children under 18: Including ages <18; justified scientifically  The study includes 5 year old children, roughly equally distributed by sex, and their adult mothers.


Subjects: Acceptable Risks and/or Adequate Protections  The human subjects plan appears well thought out Data and Safety Monitoring Plan (Applicable for Clinical Trials Only): Not Applicable (No Clinical Trials) Inclusion of Women, Minorities and Children:  Sex/Gender: Distribution justified scientifically  Race/Ethnicity: Distribution not justified scientifically  For NIH-Defined Phase III trials, Plans for valid design and analysis: Not applicable  Inclusion/Exclusion of Children under 18: Including ages <18; justified scientifically  The sample inclusion criteria are appropriate and have been revised in response to previous critique They have responded well to previous critique Authentication of Key Biological and/or Chemical Resources: Not Applicable (No Relevant Resources) Budget and Period of Support: Recommend as Requested THE FOLLOWING SECTIONS WERE PREPARED BY THE SCIENTIFIC REVIEW OFFICER TO SUMMARIZE THE OUTCOME OF DISCUSSIONS OF THE REVIEW COMMITTEE, OR REVIEWERS' WRITTEN CRITIQUES, ON THE FOLLOWING ISSUES: PROTECTION OF HUMAN SUBJECTS: ACCEPTABLEINCLUSION OF WOMEN PLAN: ACCEPTABLEThe study will enroll children regardless of sex and their adult mothers; this is scientifically justified.

Protections for Human Subjects:
Acceptable Risks and/or Adequate Protections Data and Safety Monitoring Plan (Applicable for Clinical Trials Only):

Acceptable Inclusion of Women, Minorities and Children
:  Sex/Gender: Distribution justified scientifically  Race/Ethnicity: Distribution justified scientifically  For NIH-Defined Phase III trials, Plans for valid design and analysis:  Inclusion/Exclusion of Children under 18: Including ages <18; justified scientifically Vertebrate Animals: Not Applicable (No Vertebrate Animals) Biohazards: Acceptable Resource Sharing Plans: Acceptable Authentication of Key Biological and/or Chemical Resources: Not Applicable (No Relevant Resources) Budget and Period of Support: