Master of Science, The Ohio State University, 2022, Anatomy

Manzamine-A is a marine-derived alkaloid which has anti-viral and anti-proliferative properties and is currently being investigated for its efficacy in the treatment of certain viruses (malaria, herpes, HIV-1) and cancers (breast, cervical, colorectal). Manzamine-A has been found to exert effects via modulation of SIX1 gene expression, a gene critical to craniofacial development via the WNT, NOTCH, and PI3K/AKT pathways. To date, little work has focused on Manzamine-A and how its use may affect craniofacial development. We hypothesize that Manzamine-A, through SIX1, alters bone cell activity important to craniofacial skeletal development. As Manzamine-A is a drug that has great potential in a variety of therapeutics, it is critical that we asses potential side effects that this drug could have on patients and any developmental effects that may occur during use by expectant mothers. We aimed to assess the effects of Manzamine-A on four cell types, pre-osteoblasts, osteoblasts, pre-osteoclasts, and osteoclasts. PCR, RTQPCR, MTS cell viability assays and Caspase 3/7 apoptosis assay were used to test the effects of Manzamine-A on these cells. We also performed alkaline phosphatase (ALP) and Tartrate Resistant Acid Phosphatase (TRAP) assays to test the function of osteoblasts and osteoclasts respectively. We have found that Six-1 is highly expressed in osteoblasts and osteoclasts. Our studies have shown that osteoblast progenitors and osteoblasts exhibit great sensitivity to Manzamine-A treatment exhibited by a significant decrease in cell viability, increase in cellular apoptosis, and decrease in ALP activity. Similarly, osteoclast lineage cells also show great sensitivity to Manzamine-A. Overall, our data suggests Manzamine-A may have great effects on bone health overall and may disrupt skeletal development if exposed during pregnancy or postnatal development.

Committee: James Cray (Advisor); Melissa Quinn (Committee Member); Claudia Mosley (Advisor) Subjects: Anatomy and Physiology; Biology; Developmental Biology; Medicine; Pharmaceuticals; Pharmacology

Doctor of Philosophy, Case Western Reserve University, 2009, Biochemistry

Ski is the most studied member of a family of proteins all sharing a conserved Dachshund homology domain. It has been implicated in oncogenic transformation, myogenic conversion of avian embryo fibroblasts and also many aspects of vertebrate development, especially myogenesis. Ski-/- mice exhibit severe defects in skeletal muscle and die at birth, yet little is know about either the underlying mechanisms or the role of Ski in adult muscle regeneration. In these studies, I used Ski knockout mice and C2C12 myoblast cultures to address these issues, respectively. Detailed analysis of Ski-/- embryos revealed dramatically reduced hypaxial muscles but less affected epaxial muscles. The reduced number of myogenic regulatory factor positive cells in Ski-/- mice suggested an insufficient myogenic cell pool to support muscle formation. However, both the dermomyotomal hypaxial progenitors and myotomal epaxial progenitors formed and committed to myogenic fate appropriately. The hypaxial muscle defect in Ski-/- mice was not caused by abnormal proliferation, terminal differentiation or apoptosis of the myogenic cells either, but due to impaired migration of embryonic hypaxial progenitors. Surprisingly, the normal distribution of fetal/postnatal myogenic progenitors in Ski-/- mice suggested different effects of Ski on the behaviors of embryonic and fetal/postnatal myogenic progenitors. In addition, although not affecting the terminal differentiation of embryonic myogenic cells, Ski was necessary for that of adult satellite-cell derived C2C12 myoblasts as evidenced by impaired myotube formation and reduced induction of genes essential for myogenic differentiation in the absence of Ski. This function was mainly mediated by Ski's ability to form a complex with Six1 and Eya3 and activate Myog transcription through a MEF3 site. It is important in the future to further study mechanisms underlying the contrasting effects of Ski on embryonic, fetal and adult muscle development, to investi (open full item for complete abstract)

Committee: David Samols PhD (Committee Chair); Ed Stavnezer PhD (Advisor); Clemencia Colmenares PhD (Committee Member); Nikki Harter PhD (Committee Member); Lynn Landmesser PhD (Committee Member) Subjects: Biomedical Research