IU Indianapolis ScholarWorks :: Browsing by Author "Roper, Randall"

Browsing by Author "Roper, Randall"

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An Animal Model of Combined Pituitary Hormone Deficiency Disease
(2010-08) Colvin, Stephanie C.; Konieczny, Stephen F.; Rhodes, Simon J.; Walvoord, Emily C.; Belecky-Adams, Teri; Herring, B. Paul; Roper, Randall
LHX3 is a LIM-homeodomain transcription factor that has essential roles in pituitary and nervous system development in mammals. Children who are homozygous for recessive mutations in the LHX3 gene present with combined pituitary hormone deficiency disease (CPHD) characterized by deficits of multiple anterior pituitary hormones. Most LHX3 patients also present with additional defects associated with the nervous system including a characteristic limited head rotation and sometimes deafness. However, of the 10 types of LHX3 mutation described to date, one mutation type (W224ter) does not result in the limited head rotation, defining a new form of the disease. W224ter patients have CPHD but do not have nervous system symptoms. Whereas other mutations in LHX3 cause loss of the entire protein or its activity, the W224ter mutation causes specific loss of the carboxyl terminal of the LHX3 protein—a region that we have shown to contain critical regulatory domains for pituitary gene activation. To better understand the molecular and cellular etiology of CPHD associated with LHX3 gene mutations, I have generated knock-in mice that model the human LHX3 W224ter disease. The resulting mice display marked dwarfism, thyroid disease, female infertility, and reduced male fertility. Immunohistochemistry, real-time quantitative polymerase chain reaction (PCR), and enzyme-linked immunosorbant assays (ELISA) were used to measure hormones and regulatory factor protein and RNA levels, an approach which is not feasible with human patients. We have generated a novel mouse model of human pediatric CPHD. Our findings are consistent with the hypothesis that the actions of the LHX3 factor are molecularly separable in the nervous system and pituitary gland.
Correction to: Specific Susceptibility to COVID-19 in Adults with Down Syndrome
(Springer, 2021-05-04) Illouz, Tomer; Biragyn, Arya; Frenkel-Morgenstern, Milana; Weissberg, Orly; Gorohovski, Alessandro; Merzon, Eugene; Green, Ilan; Iulita, Florencia; Flores-Aguilar, Lisi; Dierssen, Mara; De Toma, Ilario; Lifshitz, Hefziba; Antonarakis, Stylianos E.; Yu, Eugene; Herault, Yann; Potier, Marie-Claude; Botté, Alexandra; Roper, Randall; Sredni, Benjamin; Sarid, Ronit; London, Jacqueline; Mobley, William; Strydom, Andre; Okun, Eitan; Biology, School of Science
The current SARS-CoV-2 outbreak, which causes COVID-19, is particularly devastating for individuals with chronic medical conditions, in particular those with Down Syndrome (DS) who often exhibit a higher prevalence of respiratory tract infections, immune dysregulation and potential complications. The incidence of Alzheimer’s disease (AD) is much higher in DS than in the general population, possibly increasing further the risk of COVID-19 infection and its complications. Here we provide a biological overview with regard to specific susceptibility of individuals with DS to SARS-CoV-2 infection as well as data from a recent survey on the prevalence of COVID-19 among them. We see an urgent need to protect people with DS, especially those with AD, from COVID-19 and future pandemics and focus on developing protective measures, which also include interventions by health systems worldwide for reducing the negative social effects of long-term isolation and increased periods of hospitalization.
Dyrk1a Dynamics: The Influence of Gene Copy Number on Neurodevelopment in the Ts65dn Mouse Model of Down Syndrome
(2024-05) Hawley, Laura E.; Roper, Randall; Belecky-Adams, Teri; Cummins, Theodore; Goodlett, Charles; Hardy, Tabitha; Marrs, Kathleen
Down syndrome (DS) arises from the triplication of human chromosome 21 (Hsa21), leading to a spectrum of phenotypes characterized by neurodevelopmental and cognitive abnormalities. The Ts65Dn mouse model emulates DS by harboring three copies of genes found on Hsa21 resulting in trisomy 21 (Ts21)- like traits, including disruptions in neuronal pathways, delays in sensorimotor and behavior milestones, and deficits in learning and memory tasks. There is no cure for DS and available therapies primarily address symptoms stemming from Ts21-associated phenotypes. DYRK1A, a gene triplicated in Ts21, has a pivotal role in pathways of neurodevelopment and has been a focus of inhibition treatment research aimed at preempting abnormal brain phenotypes. This study aimed to find a point of substantial Dyrk1a expression dysregulation during a period of critical neonatal neurodevelopment and employ targeted pharmacological and genetic knockdown methods to alleviate the presence or severity of characteristically abnormal brain and behavior phenotypes. The hypothesis of this study was that administering a targeted intervention prior to a point of known overexpression in trisomic pups would ameliorate molecular, sensorimotor, and neurobehavioral deficits, redirecting growth trajectories of Ts65Dn neonatal pups towards more neurotypical outcomes. To test this hypothesis, the spatiotemporal pattern of DYRK1A expression was quantified during the first three weeks of neonatal development across the hippocampus, cerebral cortex, and cerebellum of the Ts65Dn mouse model and found to fluctuate according to the genotype, age, sex, and brain region of the subject. Dyrk1a protein and mRNA expression levels were delineated in trisomic animals by age, exploring the correlation between expression and age, sex, genotype, and brain region. Next a constitutive Dyrk1a knockdown model was integrated with the Ts65Dn model to investigate the impact of gene copy number reduction on protein and mRNA expression levels during phases of known DYRK1A dysregulation. On postnatal day 6, protein expression was rescued in all three brain regions of male animals but was rescued only in the cerebellum of females. There were no significant differences in mRNA transcript levels in either sex at this age. Finally, genetic elements were introduced into the Ts65Dn model to facilitate a spatiotemporally controlled functional reduction of Dyrk1a and discern how the timing of gene copy number reduction affects molecular and neurobehavioral development in a trisomic system. Results from these studies suggest that only functionally reducing Dyrk1a gene copy number on the day of birth is not sufficient to rescue the majority of deficits and delays present in the Ts65Dn mouse model of DS. These findings significantly enhance the understanding of trisomic Dyrk1a expression dynamics during neonatal development and shed light on tailored therapeutic approaches to modulate intrinsic DS characteristics based on age, sex, and phenotypic considerations.
DYRK1A-Related Trabecular Defects in Male Ts65Dn Mice Emerge During a Critical Developmental Window
(2021-08) LaCombe, Jonathan M.; Roper, Randall; Goodlett, Charles; Li, Jiliang; Wallace, Joseph; Meyer, Jason
Down syndrome (DS) is a complex genetic disorder caused by the triplication of human chromosome 21 (Hsa21). The presence of an extra copy of an entire chromosome greatly disrupts the copy number and expression of over 350 protein coding genes. This gene dosage imbalance has far-reaching effects on normal development and aging, leading to cognitive and skeletal defects that emerge earlier in life than the general population. The present study begins by characterizing skeletal development in young male Ts65Dn mice to test the hypothesis that skeletal defects in male Ts65Dn mice are developmental in nature.Femurs from young mice ranging from postnatal day 12- to 42-days of age (P12-42) were measured and analyzed by microcomputed tomography (μCT). Cortical defects were present generally throughout development, but trabecular defects emerged at P30 and persisted until P42. The gene Dual-specificity tyrosine-regulated kinase 1a (Dyrk1a) is triplicated in both DS and in Ts65Dn mice and has been implicated as a putative cause of both cognitive and skeletal defects. To test the hypothesis that trisomic Dyrk1a is related to the emergence of trabecular defects at P30, expression of Dyrk1a in the femurs of male Ts65Dn mice was quantified by qPCR. Expression was shown to fluctuate throughout development and overexpression generally aligned with the emergence of trabecular defects at P30. The growth rate in trabecular measures between male Ts65Dn and euploid littermates was similar between P30 and P42, suggesting a closer look into cellular mechanisms at P42. Assessment of proliferation of BMSCs, differentiation and activity of osteoblasts showed no significant differences between Ts65Dn and euploid cellular activity, suggesting that the cellular microenvironment has a greater influence on cellular activity than genetic background. These data led to the hypothesis that reduction of Dyrk1a gene expression and pharmacological inhibition of DYRK1A could be executed during a critical period to prevent the emergence of trabecular defects at P30. To tests this hypothesis, doxycycline-induced cre-lox recombination to reduce Dyrk1a gene copy number or the DYRK1A inhibitor CX-4945 began at P21. The results of both genetic and pharmacological interventions suggest that trisomic Dyrk1a does not influence the emergence of trabecular defects up to P30. Instead, data suggest that the critical window for the rescue of trabecular defects lies between P30 and P42.
Enhancing Our Genetic Knowledge of Human Iris Pigmentation and Facial Morphology
(2019-12) Eller, Ryan; Walsh, Susan; Berbari, Nicolas; Lapish, Christopher; Picard, Christine; Roper, Randall
The biological underpinnings that control iris pigmentation and facial morphology are two areas of research that over the last decade are becoming more thoroughly investigated due to the increased affordability of genotyping and advances in technology allowing for more advanced analysis techniques. Despite the ease of access to the data and the tools required to perform iris pigmentation and facial morphological studies, there are still numerous challenges researchers must overcome when exploring the genetics of these complex phenotypes. Some of these challenges include difficulty in working with the bioinformatic programs designed to analyze genetic associations, the inability to define a phenotype that captures the true nature of these traits, and analysis techniques that fail to model complex gene-gene interactions and their effect on a phenotype or phenotypes of interest. In this body of work, I attempted to address these challenges by designing a bioinformatic pipeline, Odyssey, that bridges the communication gaps between various data preparation programs and the programs that analyze genomic data. With this program, genome-wide association studies (GWAS) could be conducted in a quicker, more efficient, and easier manner. I also redefined iris color as a quantitative measurement of pre-defined color classes. In this way it is possible to define and quantify the unique and intricate mixtures of color, which allows for the identification of known and novel variants that affect individual iris color. I also improved upon current prediction models by developing a neural network model capable of predicting a quantitative output to four pre-defined classes; blue/grey, light brown (hazel), perceived green, and dark brown. I examined the effects of defining a simple facial morphology phenotype that more accurately captures the lower face and jaw shape. I then analyzed this phenotype via a GWAS and found several novel variants that may be associated with a square and diamond shaped face. Lastly, I demonstrated that structural equation modeling can be used in combination with traditional GWAS to examine interactions amongst associated variants, which unearths potential biological relationships that impact the multifaceted phenotype of facial morphology.
Examining Postnatal Retinal Thickness and Retinal Ganglion Cell Count in the Ts65Dn Mouse Model of Down Syndrome
(2023-05) Folz, Andrew; Roper, Randall; Goodlett, Charles; Belecky-Adams, Teri
Down syndrome (DS) is a genetic condition caused by the triplication of human chromosome 21 and presents with many phenotypes including decreased brain size, hypocellularity in the brain, and assorted ocular phenotypes. Some of the ocular phenotypes seen are increased risk of cataracts, accommodation difficulties, increased risk of refractive errors, and increased retinal thickness. The Ts65Dn mouse model of DS is a classically used mouse model as it presents a number of phenotypes also seen in those with DS. Some of these phenotypes include decreased brain volume, abnormal synaptic plasticity, and ocular phenotypes. These ocular phenotypes include decreased visual acuity, cataracts, and increased retinal thickness. The Ts65Dn mouse model is trisomic for Dyrk1a, a gene of interest in DS research. We hypothesize that there will be a genotypic and sex effect of retinal thickness and retinal ganglion cell (RGC) count at postnatal day 15 in the Ts65Dn mouse model of DS. Retinal slices were taken from male and female trisomic and euploid Ts65Dn mice at P15 and fluorescently labeled for RGCs and bipolar cells via immunohistochemistry. The retinas were measured for total retinal thickness and RNA-binding protein (RBPMS) positive cells in the RGC layer were counted. There was no genotypic or sex effect when comparing retinal thickness in trisomic mice as compared to euploid mice. There was a genotypic effect of RBPMS positive cell count in which the trisomic mice had a higher number of RBPMS positive cells than euploid mice. Increased retinal thickness along with increased RGC number have both been implicated with decreased apoptosis in the retina. In the Ts65Dn mouse model along with in individuals with DS, this could be due to an increase in DYRK1A protein levels reducing apoptosis. In future studies, determining DYRK1A’s influence in retinal thickness and RGC number could result in a treatment for overactive DYRK1A that could normalize retinal thickness and RGC number in those with DS.
Exploring the Effects of Ancestry on Inference and Identity Using Bioinformatics
(2023-08) Herrick, Noah; Walsh, Susan; Picard, Christine; Wilson, Jeremy; Balakrishnan, Lata; Roper, Randall
Ancestry is a complex and layered concept, but it must be operationalized for its objective use in genetic studies. Critical decisions in research analyses, clinical practice, and forensic investigations are based on genetic ancestry inference. For example, in genetic association studies for clinical and applied research, investigators may need to isolate one population of interest from a worldwide dataset to avoid false positive results, or in human identification, ancestry inferences can help reveal the identity of unknown DNA evidence by narrowing down a suspect list. Many studies seek to improve ancestry inference for these reasons. The research presented here offers valuable resources for exploring and improving genetic ancestry inference and intelligence toward identity. First, analyses with ‘big data’ in genomics is a resource-intensive task that requires optimization. Therefore, this research introduces a suite of automated Snakemake workflows, Iliad, that was developed to give the research community an easy-to-learn, hands-off computational tool for genomic data processing of multiple data formats. Iliad can be installed and run on a Google Cloud Platform remote server instance in less than 20 minutes when using the provided installation code in the ReadTheDocs documentation. The workflows support raw data processing from various genetic data types including microarray, sequence, and compressed alignment data, as well as performing micro-workflows on variant call format (VCF) files to merge data or lift over variant positions. When compared to a similar workflow, Iliad completed processing one sample’s raw paired-end sequence reads to a human-legible VCF file in 7.6 hours which was three-times faster than the other workflow. This suite of workflows is paramount towards building reference population panels from human whole-genome sequence (WGS) data which is useful in many research studies including imputation, ancestry estimation, and ancestry informative marker (AIM) discovery. Second, there are persistent challenges in ancestry inference for individuals of the Middle East, especially with the use of AIMs. This research demonstrates a population genomics study pertaining to the Middle East, novel population data from Lebanon (n=190), and an unsupervised genetic clustering approach with WGS data from the 1000 Genomes Project and Human Genome Diversity Project. These efforts for AIM discovery identified two single nucleotide polymorphisms (SNPs) based on their high allelic frequency differences between the Middle East and populations in Eurasia, namely Europe and South/Central Asia. These candidate AIMs were evaluated with the most current and comprehensive AIM panel to date, the VISAGE Enhanced Tool (ET), using an external validation set of Middle Eastern WGS data (n=137). Instead of relying on pre-defined biogeographic ancestry labels to confirm the accuracy of validation sample ancestry inference, this research produced a deep, unsupervised ADMIXTURE analysis on 3,469 worldwide WGS samples with nearly 2 million independent SNPs (r2 < 0.1) which provided a genetic “ground truth”. This resulted in 136/137 validation samples as Middle East and provided valuable insights toward reference samples with varying co-ancestries that ultimately affects the classification of admixed individuals. Novel deep learning methods, specifically variational autoencoders, were introduced for visualizing one hundred percent of the genetic variance found using these AIMS in an alternative method to PCA and presents distinct population clusters in a robust ancestry space that remains static for the projection of unknown samples to aid in ancestry inference and human identification. Third, this research delves into a craniofacial study that makes improvements toward key intelligence information about physical identity by exploring the relationship between dentition and facial morphology with an advanced phenotyping approach paired with robust dental parameters used in clinical practice. Cone-beam computed tomography (CBCT) imagery was used to analyze the hard and soft tissue of the face at the same time. Low-to-moderate partial correlations were observed in several comparisons of dentition and soft tissue segments. These results included partial correlations of: i) inter-molar width and soft tissue segments nearest the nasal aperture, the lower maxillary sinuses, and a portion of the upper cheek, and ii) of lower incisor inclination and soft tissue segments overlapping the mentolabial fold. These results indicate that helpful intelligence information, potentially leading towards identity in forensic investigations, may be present where hard tissue structures are manifested in an observable way as a soft tissue phenotype. This research was a valuable preliminary study that paves the way towards the addition of facial hard tissue structures in combination with external soft tissue phenotypes to advance fundamental facial genetic research. Thus, CBCT scans greatly add to the current facial imagery landscape available for craniofacial research and provide hard and soft tissue data, each with measurable morphological variation among individuals. When paired with genetic association studies and functional biological experiments, this will ultimately lead to a greater understanding of the intricate coordination that takes place in facial morphogenesis, and in turn, guide clinical orthodontists to better treatment modalities with an emphasis on personalized medicine. Lastly, it aids intelligence methodologies when applied within the field of forensic anthropology.
The identification of increased Dyrk1a protein levels in Ts65Dn mice guides the targeted administration of the novel Dyrk1a inhibitor CX-4945
(2018-04-27) Stringer, Megan; Goodlett, Charles; Roper, Randall
Down syndrome (DS) is caused by three copies of human chromosome 21 (Hsa21) and results in phenotypes including intellectual disability. Ts65Dn mice, the most extensively studied DS model, have three copies of ~50% of the genes on Hsa21 and display many phenotypes associated with DS, including cognitive deficits. DYRK1A is a dosage-sensitive gene found in three copies in humans with Trisomy 21 and in Ts65Dn mice and is involved in CNS development. Overexpression of DYRK1A is hypothesized to cause many of the cognitive and developmental deficits observed in DS and has been touted as a target for drug development in DS. Definitive evidence that excessive expression/activity of Dyrk1a directly contributes to specific phenotypes in DS mouse models is limited, and there is no direct evidence that verified pharmacological inhibition of Dyrk1a in vivo causes enduring improvement in DS cognitive phenotypes. In part, this reflects the remarkably limited knowledge of the temporal regulation of Dyrk1a expression and activity in different brain regions across development in DS mouse models. To establish the therapeutic potential of Dyrk1a inhibitors, the first aim of this study was to determine when and in what brain regions excessive Dyrk1a is evident and to identify developmental periods when elevated expression of Dyrk1a may be contributing to enduring aberrant functional development. This aim provided systematic quantification of Dyrk1a protein level at key postnatal (P) ages (P12, P15, P18, P24 P30, P42) in Ts65Dn mice, at ages of translational relevance to clinical applications in humans (birth, early adolescence, late adolescence, young adult). Western blot analysis showed that significant elevation of Dyrk1a with the largest effect sizes occurred in trisomic mice on P15. The second aim of this study was to test whether treating Ts65Dn with a novel Dyrk1a inhibitor (CX-4945) during the time of Dyrk1a elevation would rescue the behavioral and structural abnormalities observed. From P14-P18, Ts65Dn and euploid mice were treated with 75mg/kg CX-4945 or DMSO (vehicle) and tested on a homing task and locomotor activity in a novel arena on P17-P18, counterbalanced for order. At the cessation of treatment, hippocampal cell proliferation was assessed. While there was a lack of statistically significant improvements with CX-4945 treatment, there were modest effect sizes. In addition, several of the behavioral studies were significantly underpowered, making it difficult to conclusively ascertain the efficacy of CX-4945 on specific phenotypes. Nevertheless, this study demonstrates that Dyrk1a is dynamically expressed across development in mice, and suggests that consideration of the spatial and temporal expression of Dyrk1a may well be critical in the development of therapeutics for DS.
Immune Dysregulation and the Increased Risk of Complications and Mortality Following Respiratory Tract Infections in Adults With Down Syndrome
(Sage, 2021-01) Illouz, Tomer; Biragyn, Arya; Iulita, Maria Florencia; Flores-Aguilar, Lisi; Dierssen, Mara; De Toma, Ilario; Antonarakis, Stylianos E.; Yu, Eugene; Herault, Yann; Potier, Marie-Claude; Botté, Alexandra; Roper, Randall; Sredni, Benjamin; London, Jacqueline; Mobley, William; Strydom, Andre; Okun, Eitan; Medicine, School of Medicine
The risk of severe outcomes following respiratory tract infections is significantly increased in individuals over 60 years, especially in those with chronic medical conditions, i.e., hypertension, diabetes, cardiovascular disease, dementia, chronic respiratory disease, and cancer. Down Syndrome (DS), the most prevalent intellectual disability, is caused by trisomy-21 in ~1:750 live births worldwide. Over the past few decades, a substantial body of evidence has accumulated, pointing at the occurrence of alterations, impairments, and subsequently dysfunction of the various components of the immune system in individuals with DS. This associates with increased vulnerability to respiratory tract infections in this population, such as the influenza virus, respiratory syncytial virus, SARS-CoV-2 (COVID-19), and bacterial pneumonias. To emphasize this link, here we comprehensively review the immunobiology of DS and its contribution to higher susceptibility to severe illness and mortality from respiratory tract infections.
An investigation of genetic variability in Lucilia cuprina and Musca domestica utilizing phylogenetic and population genetic approaches
(2020-08) Doll, Laura Catherine; Picard, Christine J.; Roper, Randall; Tarone, Aaron
Forensic entomology is a subdiscipline of entomology that involves the use of insect behavior and developmental data to aid in criminal investigations. Genetic data has become increasingly important to the field as there has been a push for DNA-based species identification methods of forensically relevant insects. Genetic data can also elucidate population structure and relatedness of these insects, and such knowledge can contribute to the development of more specific datasets for insects in different regions. The first study presented here investigated the phylogenetics of sister species Lucilia cuprina and Lucilia sericata to identify possible subspecies divisions and issues with DNA-based identifications in the United States. The initial aim of this study was to identify genetic differences between specimens of L. cuprina that preferred live versus carrion flesh. Flies collected from Indiana, USA and South Africa were sequenced and analyzed. Upon sequencing of the genes COI, Period, and 28s, our results indicated that L. cuprina from Indiana possess a unique combination of nuclear and mitochondrial haplotypes that suggest a unique lineage, possibly indicating modern hybridization with L. sericata. The inability of both nuclear and mitochondrial genes to distinguish between L. cuprina and L. sericata raises questions about the capabilities of DNA-based species identifications within this genus. Additionally, the inability of these genes to distinguish between specimens that preferred live versus carrion flesh highlights a need for continued research of these behavioral differences. The second study presented here investigated the population structure and relatedness of house flies in the American southwest in relation to a civil lawsuit where neighbors of a poultry farm alleged that flies were emanating from the farm to their homes. Musca domestica (house fly) specimens were collected from the chicken farm and from locations in varying directions and distances from the farm. Amplified fragment length polymorphism (AFLP) analysis was performed and the data were used in a number of analyses. Population reallocation simulations generally indicated that samples from different locations were not genetically different enough from other locations to allocate to their true origin population over others. Kinship analysis showed differences in samples collected in a later season that indicate a genetic bottleneck over time. Population structure analysis indicated the presence of two intermixing genetic populations in the dataset. AMOVA revealed that the majority of genetic variation laid within, rather than among, populations. A Mantel test revealed no significant correlation between genetic and geographic distances. These results indicate that the M. domestica population in this region of southwestern America is large and intermixing, with no clear genetic distinctions between specimens collected at the poultry farm versus the surrounding locations. In regard to the civil lawsuit, it was not possible to conclude that the flies did not emanate from the poultry farm. In a broader perspective, these data can be utilized to develop pest management strategies in this region. Overall, the data from both studies presented here will be useful to forensic investigations, development of more specific and detailed data and identification techniques, and pest control measures.

Browsing by Author "Roper, Randall"

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