• Binding of 2, 3-DPG to high molecular weight substances in the red cell

      Chen, Clara Johsien; Department of Cell and Molecular Biology (1974-06)
    • Biocompatibility and mechanical/physical properties of 3D printed, milled, and conventionally processed denture base materials

      Ulmer, Mallory; Biomedical Sciences (Augusta University, 2019-12)
      According to the American College of Prosthodontists, over 36 million people in the USA are edentulous with a 2:1 predilection for geriatric patients1. Each year, an estimated 15% of edentulous Americans will seek denture treatment1. Conventional dentures require multiple visits and lab processing time. 3D printing technology offers the potential to reduce the number of appointments and speed up the time until patient rehabilitation. However, the newly FDA-certified 3D printer denture resins, featuring secretive and proprietary formulae, lack studies concerning their biocompatibility/safety and mechanical strength. This study aims to investigate the biocompatibility and physical properties of one such 3D printer resin, NextDent® Base (Vertex, Soesterberg, The Netherlands), and compare it to pre-existing conventional polymethyl methacrylate (PMMA) denture base (Lucitone 199, Dentsply Sirona, York, Pennsylvania) and milled PMMA denture base (IvoBase CAD®, Ivoclar Vivadent AG, Schaan, Liechtenstein). The cytotoxicity was examined using of 12 discs: conventional PMMA, milled PMMA, as-printed 3D printer resin, post-cured 3D printer resin, and Teflon controls. An MTT assay using human periodontal ligament (900L) cells was employed, and specimens were aged for 1, 3, 7, 10, and 14 days. After day 7, there were no statistically significant differences among the groups, excluding the Teflon control, which showed significantly less cell viability on day 14. Bars of conventional PMMA, milled PMMA, as-printed 3D printer resin, and post-cured 3D printer resin were subjected to a 3-point bend test to examine flexural strength and moduli differences. The mean flexural strength was 63.8 ± 3.06, 82.6 ± 1.9, 5.1 ± 0.4, and 22.1 ± 6.4 MPa, respectively, while the flexural moduli were 1757.3 ± 109.5, 2226.7 ± 76.3, 110.3 ± 20.3, and 537.0 ± 210.6 MPa, respectively. The flexural strength and modulus were significantly different among all groups. Weibull analyses for conventional PMMA, milled PMMA, as-printed 3D printer resin, and post-cured 3D printer resin revealed a Weibull modulus of 23.5, 42.8, 16.6, and 3.7, respectively, and a characteristic strength of 65.2, 83.5, 5.3, and 24.5 MPa, respectively. The characteristic strength was significantly different among all groups as well. The Weibull modulus was significantly different between all groups, except for conventional vs. as-printed, which were not significantly different. In summary, milled PMMA featured significantly greater mechanical properties. Both 3D printed groups proved to be very weak, with the as-printed group being the weakest of all. The differences between the as-printed and post-cured groups highlight the importance of properly post-curing the resin. While the biocompatibility results showed promise, the mechanical testing results were disappointing. Unfortunately, the findings suggest that 3D-printed denture base resin is not yet ready for clinical use.
    • Biomechanical behavior related to structure in normal and congenitally disordered elastic arteries

      Beall, Arthur C.; Department of Pharmacology and Toxicology (Augusta University, 1992-12)
    • Biosynthesis and Modification of Helicobacter pylori Lipid A

      Stead, Christopher Michael; Department of Biochemistry and Molecular Biology (2010-05)
      The secondary acylation steps of Helicobacter pylori lipid A biosynthesis are poorly understood because H. pylori only has one homolog (Jhp0265) to the Escherichia coli secondary acyl transferases LpxL and LpxM. Jhp0265 was shown to be responsible for the transfer of a secondary C18 acyl chain to the 2′-linked acyl chain of lipid A, making Jhp0265 homologous to LpxL. An activity was also demonstrated for the addition of a secondary acyl chain to the 3′-linked acyl chain of H. pylori lipid A, although the enzyme responsible for the transfer remains unknown. After synthesis, H. pylori lipid A is modified by the action of five enzymes. Mutation of the candidate modification enzyme Jhp0634 demonstrated that the enzyme catalyzes the removal of the 3′-linked acyl chains of H. pylori lipid A, producing a tetra-acylated lipid A species. Continuing with the characterization of H. pylori lipid A modification enzymes, we were also able to demonstrate an activity for a Kdo trimming enzyme in vitro. Requirement for a Kdo hydrolase in vivo was confirmed after the Kdo transferase of H. pylori was shown to be bifunctional despite the presence of only one Kdo sugar in H. pylori lipopolysaccharide. Attempted identification of the Kdo hydrolase revealed that both Hp0579 and Hp0580 were required for the removal of the Kdo sugar, which occurred in the periplasm. A Kdo hydrolase mutant revealed two unexpected phenotypes related to interaction with the innate immune system. The first was an increased sensitivity to cationic antimicrobial peptides, which was explained by a downstream effect on modification to the 4′- phosphate group of lipid A. The second phenotype related to the expression of Oantigen on the bacterial cell surface. The Kdo hydrolase mutants produced a reduced amount of fully extended lipopolysaccharide and conversely, an increased amount of core-lipid A. The type of O-antigen epitope displayed was also affected by a Kdo hydrolase mutation, in a strain specific manner.
    • Biosynthesis and modification of helicobacter pylori lipid A

      Stead, Christopher Michael; Medical College of Georgia (Augusta University, 2010)
      The secondary acylation steps of Helicobacter pylori lipid A biosynthesis are poorly understood because H. pylori only has one homolog (Jhp0265) to the Escherichia coli secondary acyl transferases Lpxl and LpxM. Jhp0265 was shown to be responsible for the transfer of a secondary C18 acyl chain to the 2' -linked acyl chain of lipid A, making Jhp0265 homologous to Lpxl. An activity was also demonstrated for the addition of a secondary acyl chain to the 3'-linked acyl chain of H. pylori lipid A, although the enzyme responsible for the transfer remains unknown. After synthesis, H. pylori lipid A is modified by the action of five enzymes. Mutation of the candidate modification enzyme Jhp0634 demonstrated that the enzyme catalyzes the removal of the 3'-linked acyl chains of H. pylori lipid A, producing a tetra-acylated lipid A species. Continuing with the characterization of H. pylori lipid A modification enzymes, we were also able to demonstrate an activity for a Kdo trimming enzyme in vitro. Requirement for a Kdo hydrolase in vivo was confirmed after the Kdo transferase of H. pylori was shown to be bifunctional despite the presence of only one Kdo sugar in H. pylori lipopolysaccharide. Attempted identification of the Kdo hydrolase revealed that both Hp0579 and Hp0580 were required for the removal of the Kdo sugar, which occurred in the periplasm. A Kdo hydrolase mutant revealed two unexpected phenotypes related to interaction with the innate immune system. The first was an increased sensitivity to cationic antimicrobial peptides, which was explained by a downstream effect on modification to the 4'- phosphate group of lipid A. The second phenotype related to the expression of 0- antigen on the bacterial cell surface. The Kdo hydrolase mutants produced a reduced amount of fully extended lipopolysaccharide and conversely, an increased amount of core-lipid A. The type of 0-antigen epitope displayed was also affected by a Kdo hydrolase mutation, in a strain specific manner.
    • Biosynthesis of the Vibrio cholerae Kdo-lipid A Domain and its Role in Pathogenesis

      Hankins, Jessica V.; Department of Biochemistry and Molecular Biology (2011-05)
      Bacteria assemble remarkable surface structures that interface with their surrounding environment. One such structure is the glycolipid lipopolysaccharide (LPS) that covers the surface of Gram-negative bacteria. LPS is anchored to the bacterial cell by its lipid anchor known as lipid A. Since lipid A is the bioactive component of LPS, modulation of its structure can have a profound impact on disease by altering the host immune response. Additionally, LPS structure directly impacts the outer membrane permeability barrier and bacterial resistance to host antimicrobial peptides. Although the lipid A domain of Escherichia coli has been well characterized, the Vibrio cholerae lipid A biosynthetic pathway has received little attention. The late stages of lipid A biosynthesis include the transfer of the 3-deoxy-Dmanno- octulosonic acid (Kdo) sugars and the secondary acyl chains to the lipid A backbone. Here, the V. cholerae Kdo transferase (Vc0233) was shown to be monofunctional, transferring one Kdo residue to the lipid A precursor, lipid IVA. V. cholerae encode a Kdo kinase (Vc0227) responsible for the phosphorylation of the Kdo residue. The functionality of Vc0227 was shown to be required for the activity of the V. cholerae lipid A LpxL homologue, Vc0213. Interestingly, the addition of the phosphate group on the Kdo sugar was shown to be essential for lipid A secondary acylation in Haemophilus influenzae and Bordetella pertussis. Vc0213 was shown to catalyze the transfer of a myristate (C14:0) to the 2′-position of the V. cholerae phosphorylated Kdolipid A domain. A second protein, Vc0212, acts as an LpxM homologue and transfers 3- hydroxylaurate (3-OH C12:0) to the 3′-position creating hexa-acylated V. cholerae lipid A domain. Although lipid A secondary acyltransferases have been characterized among various Gram-negative bacteria, this is the first report of a lipid A secondary hydroxyacyltransferase. Further, the transfer of 3-hydroxylaurate (3-OH C12:0) was demonstrated to be essential for antimicrobial peptide resistance in V. cholerae and required for activation of the innate immune receptor TLR4.
    • Bisphosphonate-Related Osteonecrosis of the Jaw: From Mechanism to Treatment

      Howie, Rebecca; Department of Cellular Biology and Anatomy (2015-04-20)
      With 55 million prescriptions issued each year, bisphosphonates are the second most prescribed class of drug in the United States. They are widely used to treat diseases with excessive osteoclastic resorption, including post-menopausal osteoporosis, Paget’s disease, and tumor metastasis to bone. Unfortunately, with long term intravenous administration of nitrogen-containing bisphosphonates some patients develop bisphosphonate-related osteonecrosis of the jaw (BRONJ). This debilitating disease has limited treatment options once it has manifested and no mechanism for its development has been elucidated. This dissertation explores the novel concept that bisphosphonates cause osteonecrosis of the jaw by impairing osteocyte-induced osteoclastogenesis and, through the physical accumulation of bisphosphonates in bone, impairing the ability of recruited osteoclasts to attach thereby arresting bone healing. Furthermore, it explores the possibility that chelating agents can be used for the removal of bisphosphonate attachment from bone systemically and locally during extractions, potentially leading to a future preventive treatment. It was found that 13 weeks of 80µg/kg intravenous tail vein injections of Zoledronate followed by two mandibular molar extractions caused the clinical presentation of BRONJ as analyzed by the gross, radiographic, and histological methods. Bone dynamic parameters and TRAP staining suggested an impaired ability for the bone to remodel and defective osteoclast attachment in treated groups that persisted eight weeks after the cessation of treatment. Additionally, it was found through the use of a fluorescently tagged bisphosphonate, that the decalcifying agents cadmium, EDTA, and citric acid all had the ability to cause the significant release of bound bisphosphonate from bone. Finally, this dissertation showed that the migration of monocytes treated with low doses of Zoledronate had increased migration, while their migration to conditioned media of osteocytes treated with Zoledronate was impaired. Collectively, these data suggest that invasive trauma by itself consistently precipitated massive bone necrosis in Zoledronate treated animals, possibly through a bisphosphonate driven alteration of monocyte migration and that the use of decalcifying agents could acutely remove bisphosphonate from bone both systemically and locally. This study establishes and effective rodent model for BRONJ and a possible preventive strategy for the side-effects of bisphosphonates during high-risk situations.
    • Blood pressure impacts the renal T cell profile of male and female spontaneously hypertensive rats

      Tipton, Ashlee J.; Department of Physiology (2014-03)
      Of the 68 million Americans with hypertension, fewer than 46% have their blood pressure (BP) adequately controlled and women are more likely than men to have uncontrolled hypertension. This underscores the critical need for new treatment options; however, this is a challenge due to our lack of knowledge regarding the mechanism(s) driving essential hypertension. T cells have been implicated in hypertension in males. Prior to our work, the role of T cells in hypertensive females had been unexplored. We demonstrate that female spontaneously hypertensive rats (SHR) have a decrease in BP in response to an immunosuppressant, supporting an immune component to their hypertension. We further defined a sex difference in the renal T cell and cytokine profile in SHR. Female SHR have a more anti-inflammatory immune profile in their kidneys than males. To gain insight into the mechanisms mediating sex differences in the immune profile, male and female SHR were gonadectomized. Gonadectomy increased pro-inflammatory markers in both sexes and attenuated anti-inflammatory markers particularly in females. Therefore, while both male and female sex hormones promote an anti-inflammatory immune profile, female ii sex hormones contribute greater to their more anti-inflammatory profile, but do not explain the sex difference. To determine the impact of hypertension on the renal immune profile, experiments measured renal T cells and cytokines in hypertensive male and female SHR, normotensive Wistar Kyoto rats (WKY), and SHR treated with antihypertensive therapy. All T cells and cytokines measured were higher in SHR compared to the same sex WKY. Moreover, antihypertensive therapy decreased renal Tregs only in female SHR. These data suggest that increased BP in both sexes is associated with an increase in renal inflammation; however female SHR have a compensatory increase in renal Tregs in response to increases in BP. TGF-β is a key cytokine regulating Treg and Th17 differentiation and we found that female SHR express more TGF-β than males. Experiments assessed if female SHR possessed a sex hormone or BP-mediated increase in renal TGF- β corresponding with increases in Tregs. We determined that loss of female sex hormones and increased BP in female SHR increase renal TGF-β expression. We conclude that BP status drive sex differences in the renal T cell and cytokine profile of SHR.
    • Bone and soft tissue regenerative response following alveolar ridge augmentation using polysulfone implants with and without demineralized bone powder in macaca fascicularis

      Fouad, Salama S; Department of Oral Biology (1987-06)
      Successful augmentation of bone surfaces has great clinical application, particularly to the face and oral cavity regionso More than 24 mi 11 ion Americans are edentulous and must depend upon dentures to eat and to restore their norma 1 speech and appearance o Porous po lysul fane (PPSF) is frequently used to fill osseous voidso The purpose of this study was to· test tooth soft, ti.ssue and bone response to porous po lysul fane (PPSF), with and without demineralized bone powder (DBP) in Macaca fascicularise Six adult female monkeys, 12-15 year~ of ~ge, were used in this study. One animal was sacrificed and used as a bone donor and the other five were recipientso All mandibular molar teeth were extracted and masstve alveolectomies were performedo The wounds were left to heal for 5 to 8 1/2 months postoperativelyo At the time of implantation~ PPSF with DBP was inserted subperiosteally into the left mandibular edentulous areas while PPSF alone was inserted into the right sideso The. animals were sacrificed at 42, .60, or 90 days following implantationo Each mandible was cut- into 3mm thick coronal sections which were then examined and photographed with a dissecting microscopeo Some specimens were then decalcified, embedded in paraffin and sect i a ned and stained With. H & Eo Other specimens were processed undecalcified in glycol and methylmethacrylate for histomorphometric measurements and tetracycline labe1ling·., Also, some specimens were processed for scanning electron microscopyo No inflammation or untoward reaction of the 'implantation sites were noted at the time of sacrifice. Histologically, the 42 day specimens of the' DBP-PPSF side· (experimental side) revealed penetration of fibrous tissue rich in fibroblasts and vessels into the pores of PPSF comparirig to the PPSF side (control side)., The fibrous tissue also surrounded the implant., Some multinucleated giant cells and macrophages were present., At 60 days, the PPSF side showed more organized fibrous tissue and bone grew only for a. short distance into the polysulfone. In contrast, the PPSF-DBP side showed large amounts of .bone formation within the pores of the polysulfone and almost covered the implant., The newly formed bone contained osteocytes and was -surrounded by osteob 1 asts. At 90 days, the PPSF side showed more bone tormation on the lower half of the. implant" These res~lts suggested that PPSF is a suitable non-resorbable material that accommodates bone and soft tissue formation. A 1 so :p the use of DBP enhanced both rate and amount of the new bone., In· conclusion; PPSF with and without DBP is a suitable material that can be used successfully for alveolar ridge augmentation.
    • Bone regenerative response following bone augmentation using hydroxyapatite with and without growth factors

      Sohn, Jeong-Yeol; School of Graduate Studies (1997-09)
      The purpose of this study was to determine whether adding transforming growth factor-13 (TGF-13) which is known to promote osteogenesis or 15 amino acid polypeptide obtained from collagen type I (P-15) which is known to enhance DNA synthesis in cultured fibroblasts, would enhance the osseoint~gr.ation- of porous anorganic bovine bone block (HA-block) to mandibular bone surface. Total of 45 rabbits were divided into three equal groups. The buccal side of the mandible in ea.ch anesthetized animal was surgically exposed,. decorticated and 10 x 5 x 4 mm. HA (Group I), HA+ TGF-13 (Group II) or HA + P-15 (Group III) blocks. were affixed to .host bone using two, titanium screws. Animals were sacrificed, 1 wk, 4 wks and 8 wks after surgery. A core of the implant· and underlying host bone was used to determin.e osteocalcin using Western and Slot blot analyses before perfusing the animals with 4% forma}in. The contralateral sides were only decorticated and served as sham controls. The interface between the HAblock and the ho.st bone showed a signifi.cantly higher (P<0~05) number of mesenchymal cells in the HA + TGF--P. and HA + P-15 one week group when c.ompared. to HA-alone. This numb.er of mesenchymal ·cells declined significantly .by the 4th and 8th weeks. The proliferating cell nuclear antigen (PCNA) index using immunohistochemistry als.o showed a significant increase in the one week groups and· then also declined. Alkaline phosphatase, osteocalcin, bone volume, and new bone formation were all significantly increased by the 4th and 8th weeks and were significantly higher in the HA + TGF-13 and HA + P-15 at 4 wks after surgery. In addition, the macropores of the lower halves of all blocks were filled with new bo.ne. The den~ity of the newly formed bone was comparable to the control bone. These res.ults indicate that anorganic :bovine bQne blocks are osteoconductive and their pores permit growth of new bone. The adding of TGF-P or P-15 enhanced all parameters of osteogenesis at the mandibular bone surface.
    • Burnout and Job Satisfaction of Medical-Surgical Nurses

      Garbutt, Susan J.; School of Nursing (1981-12)
      Job satisfaction and burnout of medical-surgical nurses a n d t h e i r . r e 1 at i_ o n s h i p to . t h. e ~ ri u· r s .. e ' s o p t i. on t o w or k h e r I h i s s hi f t ·of choice was stud i e d . The Mas 1 a c h Burnout Inventory (Maslach and Jackson, 1981-), the Job Satisfaction-Instrument (Wall, ·1980; and Sorrow, 1980) and a Demographic Data Form were used to survey t~enty randomly selected registe~ed nurses f r o m e a c h . o f t h r e e s o u t h e a s t e r n h o s p l t a 1 s . ( t o t-a 1 N = 6 0 ) .. Result~ of this study indicated that as job satisfact}on increased, the intensity of burnout reported by medical-surgical nurses decreased. Frequency and intensity of burnout were positively correlated. Burnout occorred among n~rses who worked their choice of shift and among nurses who did not. However, nurses who worked their choice of shift reported a significantly lower intensity of burnout than nurses who did not work their choice of shift. This study prov.ides some interest-· ing data to be ~onside~ed by nurse administrators in working .with medical-sur9ical nurses. Replication and extension of this study is recommended.
    • The c-MYC oncogene deregulates global DNA methylation and hydroxymethylation to control genome-wide gene expression for tumor maintenance in leukemia/lymphoma

      Poole, Candace Jean; Biomedical Sciences (Augusta University, 2019-05)
      Aberrant DNA methylation is a characteristic feature of tumor cells. However, our knowledge of how DNA methylation patterns are established and maintained to contribute to tumorigenesis is limited. Inactivation of the c-MYC oncogene triggers tumor regression in T-cell acute lymphoblastic leukemia (T-ALL) resulting in dramatic changes to the chromatin landscape including DNA methylation. In this study, I investigated how MYC regulates DNA methylation and hydroxymethylation patterns to contribute to gene expression programs important for tumor maintenance in T-ALL and Burkitt lymphoma. I report that MYC maintains 5-methylcytosine (5mC) and 5-hydroxy-methylcytosine (5hmC) patterns by regulating the DNA methylation machinery, which is important for gene expression in T-ALL. DNA methyltransferases (DNMTs) initiate 5mC marks, while Ten-eleven translocation methylcytosine dioxygenases (TETs) oxidize 5mC to produce 5hmC as an intermediate modification, ultimately leading to active DNA de-methylation. I demonstrated that DNMT1 and DNMT3B are MYC target genes and that their expression is dependent on high MYC levels. Knockdown of DNMT3B in T-ALL reduced cell proliferation through cell cycle arrest and caused the reactivation of gene transcription through reversing promoter/CpG island methylation. Furthermore, I demonstrated that TET1 and TET2 expression is MYC-dependent, as high TET1 and low TET2 levels depend on oncogenic MYC. Knockdown of TET1 in T-ALL reduced cell proliferation through cell cycle arrest and caused genome-wide changes in 5mC and 5hmC corresponding to changes in gene programs important for ribosomal biosynthesis and protein synthesis. In contrast, ectopic expression of TET2 reduced tumor cell proliferation through apoptosis/necrosis and caused genome-wide changes in 5mC and 5hmC corresponding to changes in transcriptional regulatory gene programs. My finding that a coordinated interplay between components of the DNA methylation machinery is necessary for MYC-driven tumor maintenance highlights the potential of targeting specific DNMT or TET proteins for therapeutic strategies.
    • Calcyon, a novel partner of clathrin light chain, stimulatesclathrin-mediated endocytosis

      Xiao, Jiping; Xiao, Jiping; School of Graduate Studies (2006-12)
      In the central nervous system, clathrin-meriate,d endocytosis (CME) is crucial for ' I • efficient synaptic transmission as CME regulates both presynaptic release of neurotransmitter and postsynaptic responses to .transmitter. Clathrin-coated vesicle assembly and disassembly are regulated by som~ 30 adaptor and accessory proteins, most I of which interact with clathrin heavy chain. Calcy_on, which is a single transmembrane protein predominantly expressed in brain, is localized to vesicular compartments within pre and postsynaptic structures. Calcyon has been implicated as a candidate gene for schizophrenia, but the function of calcyon is not yet described. Using the calcyon cytosolic domain as bait, we isolated clathrin light chain (LC) in a yeast two-hybrid screen. The interaction domains were mapped to the heavy chain binding domain and Cterminal regions in LC. In calcyon, residues123 to 155 of calcyon mediated the interaction with LC. Addition of a purified fusion protein containing the calcyon C terminus stimulated clathrin self-assembly in vitro in a dose-dependent fashion. There was a high degree of overlap in the distribution of LC and-calcyon in neuronal processes and cell bodies. Co-immunoprecipitation studies further suggested an association of calcyon with adaptor proteins that play a role in clathrin coated vesicle formation at the plasma membrane, trans Golgi Network and endosomes. Compared to control, HEK293 cells overexpressing calcyon exhibited significantly enhanced transferring (Tfn) uptake but equivalent levels.ofTfn recycli1:1g. Copversely, transferrin uptake was largely abolished in neocortical neurons obta;ined from mice homozygous for a calcyon null allele, whereas recycling proceeded at wild type levels. Deletion of the calcyon gene in mice also inhibited agonist-sti~ulated endocytosis of the GluRl and GluR2 subunits of the ligand gated AMP A type glutamate receptor in primary neurons in cultures. Collectively, these data indicate a role for calcyon in clathrin-mediated endocytosis in brain.