This was an exploratory-descriptive study of fifty children afflicted with myelomeningocele, ages one through six, who were known to the Myelomeningocele Clinic of the Crippled Children’s Division. The study identified the degree of multiple physical, emotional, and environmental stress factors that families must be prepared to cope with. The study identified eleven factors felt to play an important role in family dynamics and how they related to the families response to their child with myelomeningocele. The factors were tested and found to be valid by the use of a pre-test on ten case records. Medical records were then obtained from the Crippled Children's Division for chart review purposes and the appropriate material was recorded. Scores were developed that indicated the degree of stress ranging from minimal involvement to maximum involvement. The study found that a majority of the families in the sample live within commuting distance to needed medical services, have transportation available to them and generally utilize the necessary medical care appropriately. The remainder of the study showed, however, that families could be expected to face a variety of other problems that could only serve to increase family stress. Most of the families had limited financial resources. Over one-half of the families needed special education for their children. A majority of families had no medical insurance. Fifty-eight percent of the families were found to have additional stressful problems to cope with e. g., marital stress, sibling rivalry, additional ill members, etc. Added to this was the information that the child with myelomeningocele was found to be greatly involved in a multiplicity of medical problems at many different levels of functioning. e. g., orthopedic, bowel, neurosurgical, etc., that would be expected to add to the already stressful family dynamics. Among the recommendations developed was a plea for the expansion of the satellite clinic concept, development of parent groups on a geographical basis, development of educational programs for educators and community service personnel, brief orientation programs for parents with the goal of helping them understand and integrate the health care system that they find themselves in.

Prematurity is a worldwide problem. Every year, 15 million babies are born prematurely, and 1 million of those babies die because of related complications. The surviving premature babies are struggling to hold on to their lives, and even when they do live, most of them end up having various complications to survive and get stronger. There are physical complications faced on their journey such as having underdeveloped lungs, pneumonia, obesity, sepsis, retinopathy of prematurity, respiratory distress syndrome, bronchopulmonary dysplasia, asthma, wheezing, bronchiolitis, cerebral palsy, and motor impairment. They can also develop mental and behavioral health complications such as depression, seizures developmental delay, schizophrenia, autism spectrum disorder, psychological development disorders, behavioral problems, attention problems, and ADHD later in life. The purpose of this systemic review is to understand the impact of long-term complications of premature birth on individual life and society. We hypothesized that based on data from primary research, nearly one half of the infants will have either physical and/or cognitive/developmental health complications. We hypothesized that infants born premature have more physical complications than cognitive complications and infants born prematurely have more cognitive complications than physical complications. This research was carried out by finding cohort study design studies through Medline, Academic Search Premier, and APA PsychINFO, where the studies will be compiled from 2003 – 2020.

Cardiac arrhythmias are caused by a disruption of the normal initiation or propagation of electrical impulses in the heart. Hundreds of mutations in genes encoding ion channels or ion channel regulatory proteins are linked to congenital arrhythmia syndromes that increase the risk for sudden cardiac death. This dissertation focuses on how mutations in a gene (KCNQ1) that encodes a voltage-gated K+ ion channel (Kv7.1) can disrupt proper channel function and lead to abnormal repolarization of atrial and ventricular cardiomyocytes. In the heart, Kv7.1 coassembles with a regulatory protein to conduct the slowly activating delayed rectifier K+ current (IKs). Loss-of-function KCNQ1 mutations are linked to type 1 long QT syndrome (LQT1), and typically decrease IKs, which can lead to ventricular action potential (AP) prolongation. In patients, LQT1 is often characterized by an abnormally long corrected QT (QTc) interval on an electrocardiogram (ECG), and increases the risk for polymorphic ventricular tachycardias. KCNQ1 mutations are also linked to atrial fibrillation (AF), but cause a gain-of-function phenotype that increases IKs. Surprisingly, patients diagnosed with both LQT1 and AF are increasingly identified as genotype positive for a KCNQ1 mutation. The first aim of this dissertation was to determine a unique functional phenotype of KCNQ1 mutations linked to both arrhythmia syndromes by functional analyses via the whole-cell patch clamp technique in HEK293 cells. A proportion of patients with LQT1-linked KCNQ1 mutations do not have abnormal QTc prolongation known as latent LQT1. Interestingly, exercise can reveal abnormal QTc prolongation in these patients. During exercise, beta-adrenergic activation stimulates PKA to phosphorylate Kv7.1, causing an increase in IKs to prevent ventricular AP prolongation. Therefore, the second aim of this dissertation was to determine a molecular mechanism of latent LQT1 through functional analyses in HEK293 cells while incorporating pharmacological and phosphomimetic approaches to study PKA regulation of mutant Kv7.1 channels. The findings in this dissertation provide new insight into how KCNQ1 mutations disrupt the function of Kv7.1 in a basal condition or during beta-adrenergic activation. Also, this dissertation suggests these approaches will improve patient management by identifying mutation specific risk factors for patients with KCNQ1 mutations.

Smith-Magenis Syndrome (SMS; OMIM #182290) is a multiple congenital abnormality and intellectual disability (ID) disorder caused by either an interstitial deletion of the 17p11.2 region containing the retinoic acid induced-1 (RAI1) gene or a mutation of the RAI1 gene. Individuals diagnosed with SMS typically present characteristics such as ID, self-injurious behavior, sleep disturbance, ocular and otolaryngological abnormalities, craniofacial and skeletal abnormalities, neurological and behavioral abnormalities, as well as other systemic defects and manifestations. Previous work by Vyas in 2009 showed temporal expression of rai1 in zebrafish embryos as early as 9 hpf. We hypothesize that there is maternal rai1 expression as early as zero hours post fertilization in wild type embryos. Using end-point PCR, we found that in fact there is maternal rai1 expression is detectable as early as 2 hours post fertilization (hpf) in wild type zebrafish embryos. Furthermore, we quantified rai1 expression using qPCR and found that rai1 expression declines significantly after 6 hpf. We hypothesize that a down regulation of rai1 or loss of rai1 will lead to morphological phenotypes, especially if that loss of rai1 function occurs during the earliest stages of zebrafish embryogenesis. Using a rai1morpholino oligonucleotide (MO), we found a loss of rai1 expression did not induce a morphological phenotype in in wild type embryos; furthermore, we also found that a loss of maternal rai1 expression did not induce a morphological phenotype as well. Utilizing a mutant rai1 zebrafish line, we found that both rai1 +/fh370 progeny nor rai1 fh370/fh370 progeny exhibited a morphological phenotype and that downstream targets such as bdnf were not affected by a reduction or complete loss of rai1. Prior research has shown that retinoic acid (RA) can induce rai1 expression. We hypothesize that RA can induce expression of rai1 during zebrafish embryogenesis. Using wild type fish and a rai1 in situ hybridization probe, we found that RA treatment at 25 hpf induced expression of rai1. The construction of a rai1 overexpression vector used for overexpression studies was started. Further development of GFP expression vector and zebrafish rai1 antibody are needed to determine if the morpholino is reducing rai1 protein expression.

Congenital heart disease is the most common congenital anomaly, affecting approximately 1% of all live births each year. Although clinical interventions are improving, many affected infants do not survive to adulthood. Congenital cardiac defects originate from disturbances during development, making the study of mammalian cardiogenesis critical to improving outcomes for infants with congenital heart disease. Development of the mammalian heart involves epigenetically-driven specification and commitment of a diverse landscape of cardiac progenitors. Recent studies determined that chromatin modifying enzymes play a previously underappreciated role in the pathogenesis of congenital heart defects. This thesis investigates the functions of Hdac1 and Hdac2, highly homologous Class I histone deacetylases, during early murine cardiac development. We establish that Hdac1 and Hdac2 cooperatively regulate cardiogenesis in distinct cardiac progenitor populations during development. Together, our findings demonstrate that Hdac1 and Hdac2 are critical mediators of the earliest stages of mammalian cardiogenesis through a variety of spatiotemporally specific, redundant, and dose-sensitive roles and indicate they may play important roles in the pathogenesis of human congenital cardiac defects.

The model of basal ganglia function proposed by Albin, Young and Penney (1989) describes two anatomically independent motor pathways, the direct and indirect. However, under normal conditions striatal dopamine (DA) is required for the expression of motor behavior, and DAergic control of the two pathways (via D1 and D2 receptors, respectively) is dependent on co-activation. We tested for a possible breakdown of D1/D2 synergism using transgenic R6/1 mice bearing the human huntingtin allele (Htt). Motor stereotypy, observed prior to the onset of HD-related symptoms, was rated on a 5-point scale following activation of: A) D1 receptors alone, B) D2 receptors alone, and C) stimulation of both D1 and D2 receptors. Results revealed that mice with the HD allele, like their WT litter mates, depend on the co-activation of the indirect and direct motor pathways to facilitate deliberate behavior.

Introduction: Current medical treatment for individuals diagnosed with schizophrenia (SCHZ) primarily relies on the inhibition of the dopamine D2 receptor that has been shown to be supersensitive in these patients. Treatment occurs through the use of antipsychotic medication which leads to a number of debilitating dose-dependent side effects, such as weight gain, agranulocytosis, and seizures. Patients diagnosed with SCHZ have also been shown to have increased inflammation in their central nervous system (CNS), particularly within specific brain regions such as the prefrontal cortex and hippocampus. This is in large part due to the interaction between a pro-inflammatory cytokine called tumor necrosis factor-alpha (TNFa) and microglia, which are resident CNS defense cells. TNFa is a cell-signaling protein, regulates a variety of immune cells, and is involved in the acute phase reaction of inflammation. Upon activation by TNFa secretion, microglial cells switch from being anti-inflammatory (M2) to pro-inflammatory (M1), thereby resulting in neuroinflammation as well as synaptic loss and neuronal death. In this project, we hypothesized oral administration through the diet of a novel TNFa modulator (PD2024) developed by P2D Biosciences, Inc. (Cincinnati, OH) would significantly reduce microglia activation in rats neonatally treated with Polyinosinic:polycytidylic acid (poly I:C). Methods and Results: To test our hypothesis, four groups (Neonatal Poly I:C/TNFa, Neonatal Poly I:C/Control, Neonatal Saline/TNFa, and Neonatal Saline/Control) were intraperitoneally injected with either poly I:C or saline during postnatal days (P)5-7. Poly I:C is an immunostimulant that mimics neonatal infection in humans, which also has been found to be a factor for the development of SCHZ later in life. Between days (P)30-(P)60, the Neonatal Poly I:C/TNFa and Neonatal Saline/TNFa groups were orally administered PD2024 through the diet. After (P)60, brain tissue was evaluated by immunohistochemistry (IHC) and confocal microscopy. Immunohistochemistry was used to label microglial cells in the prefrontal cortex and hippocampus with a green fluorescent dye attached to Iba1, a protein that specifically binds to these cells. Upon completion of IHC, tissue was evaluated using a confocal microscope and then analyzed with NIH ImageJ software. Analysis parameters included cell count, sampled cell body fluorescence, and overall image fluorescence. The results obtained showed a significant decrease in microglia activation for the Poly I:C/TNFa group when compared to the Poly I:C/Control group, as well as similarities in activation levels with the Saline/Control group. These results were demonstrated in both sampled cell body fluorescence and overall image fluorescence measurements. Conclusion: This data supports the hypothesis that PD2024 is successful in reducing microglia activation through the modulation of TNFa. Therefore, treatment with a TNFa modulator such as PD2024 alongside of current antipsychotic medication could mediate neuroinflammation and reduce the dose-dependent side effects. This approach could be a promising therapeutic treatment option for those diagnosed with schizophrenia, as well as potentially for other neurocognitive and behavioral disorders.

Dwarfism is considered one of the most recognized congenital defects of animals and humans and can be hereditary or sporadic in cause and expression. There are two general morphologic categories within this vastly diverse disease. These categories are disproportionate and proportionate dwarfism and within each of these there are numerous phenotypes which have been extensively described in humans, and to a lesser extent in dogs, cattle, mice, chickens, and other domestic species. Ponies and Miniature horses largely differ from full size horses only by their stature. Ponies are often defined as those whose height is not greater than 14.2 hands; however the maximum height for Miniature horses is constitutionally defined as 8.2 hands. Dwarfism is not considered a desirable genetic trait for Miniature horses. A majority of these conformationally inferior horses showed consistent physical abnormalities typical of disproportionate dwarfisms as seen in other mammal species. A whole genome scan with the Illumina Equine SNP50 chip clearly implicated a region on ECA1 as being associated with dwarfism of horses. The region implicated on the horse chromosome 1 (Equus Caballus; ECA1) contained a candidate gene for dwarfism, aggrecan (ACAN). Mutations were found in Exons 2, 6, 11 and 15 with each mutation associated with a distinct type of dwarfism. These mutations are independently transmitted throughout the population. Absence of normal homozygotes for these mutations and absence of normal horses which were heterozygous for these mutations indicated that these alleles caused dwarfism in those genotypes. These genotypes did not explain all observed dwarves in this population.

In humans, Bloom’s Syndrome is caused by a mutation of the RecQ helicase BLM. Patients with Bloom’s Syndrome exhibit a high amount of genomic instability which results in a high incidence of cancer. Though Bloom’s Syndrome has been intensively studied, there are still many questions about the function of BLM which need to be answered. While it is clear that loss of BLM increases genomic instability, the other effects of genomic instability on the organism aside from cancer such as a potential effect on aging, have yet to be elucidated. In Chapter II, I identify new phenotypes in the C. elegans ortholog of BLM, him-6. him-6 mutants have an increased rate of cell death, a mortal germ line phenotype, and an increased rate of mutations. Upon further examination of the mutator phenotype, it was determined that the increased rate of mutations was caused by small insertions and deletions. The mutator phenotype identified in him-6 mutants closely mimics the cellular phenotype seen in Bloom’s Syndrome cells. This indicates that HIM-6 may behave in a similar fashion to BLM. In addition to the mutator phenotype, it was found that loss of him-6causes a shortened life span. This may provide evidence that there is a link between genomic stability and aging. In Chapter III, I identify a new role for the transcription factor DAF-16. DAF-16 in C. elegans has been intensively studied and regulates a wide variety of pathways. In this chapter, I demonstrate via the well established unc-93 assay that loss of daf-16 causes a subtle mutator phenotype in C. elegans. This indicates that DAF-16 may play a role in suppression of spontaneous mutation. When I examined other classic genomic instability phenotypes, I found at 25°C, the number of progeny in the DAF-16 mutants was significantly reduced compared to wild type worms. Additionally, I demonstrate daf-16(mu86)has a cell death defect. This study identifies several new phenotypes caused by a loss of him-6. These phenotypes provide further evidence that loss of him-6 causes genomic instability. In addition, this study also demonstrates that him-6 has a shortened life span which may be due to genomic instability. Secondly, this study identifies a new role for DAF-16 in preventing the occurrence of spontaneous mutations. This may indicate a novel function for DAF-16 in maintaining genomic stability.

The KCNE1 and KCNE3 type I transmembrane-spanning β-subunits assemble with the KCNQ1 voltage-gated K+ channel to afford membrane-embedded complexes with dramatically different properties. Assembly with KCNE1 produces the very slowly activating and deactivating IKs current that shapes the repolarization phase of cardiac action potentials. Genetic mutations in KCNQ1 or KCNE1 that reduce IKs current cause long QT syndrome and predispose affected individuals to potentially fatal cardiac arrhythmias. In contrast, complexes formed between KCNQ1 and KCNE3 produce rapidly activating and mostly voltage-independent currents, properties that are essential for function in K+ recycling and Cl−secretion in gastrointestinal epithelia. This thesis addresses how these two homologous accessory peptides impart their distinctive effects on KCNQ1 channel gating by examining two important protein regions: 1) a conserved C-terminal motif in the β-subunits themselves, and 2) the voltage sensing domain of KCNQ1 channels. Sequences in both the transmembrane domain and C-terminus of KCNE1 and KCNE3 have been identified as contributing to the divergent modulatory effects that these β-subunits exert. The homology of transmembrane-abutting C-terminal residues within the KCNE family and the presence of long QT-causing mutations in this region highlight its importance. A bipartite model of modulation was proposed that suggests the transmembrane domain of KCNE1 is passive, allowing the C-terminal domain to control modulation. Chapter II builds on this model by investigating the effect of mutating specific amino acids in the KCNE1 C-terminal domain. Point mutants that produce ‘high impact’ perturbations in gating were shown to cluster in a periodic fashion, suggesting an alpha-helical secondary structure that is kinked by a conserved proline residue and interacts with the Q1 channel complex. In Chapter III, the voltage sensing domain of Q1 channels is examined in the presence of either KCNE1 or KCNE3. To determine the influence of these two peptides on voltage sensing, the position of the S4 voltage sensor was monitored using cysteine accessibility experiments. In the slowly opening KCNQ1/KCNE1 complexes, voltage sensor activation appears to occur much faster than the onset of current, suggesting that slow channel activation is not due to slowly moving voltage sensors. KCNE3, on the other hand, shifts the voltage sensor equilibrium to favor the active state, producing open channels even at negative voltages. Taken together, these findings provide mechanistic detail to illustrate how two homologous peptides radically alter the gating properties of the same K+ channel and present a structural scaffold to map protein-protein interactions.