• Identification and characterization of CRIP1b: a novel CB₁ cannabinoid receptor interaction protein

      Niehaus, Jason Lance; School of Graduate Studies (2006-07)
      G protein-coupled receptors. (GPCRs) transduce extracellular stimuli to intracellular signals. through their interaction with heterotrimeric G proteins. Signaling diversity and specificity is imparted primarily through variations of G protein subunits. Protein-protein interactions between intracellular accessory proteins and GPCRs also modify signaling by altering receptor activity or signaling pathways. The ability of intracellular proteins to interact with the CB 1 cannabinoid receptor was investigated to determine whether particular signaling properties of CB 1 resulted from interaction with specific CB 1 interacting proteins. A novel protein named CRIP 1 b was discovered· to interact with the C-terminal tail of CB 1 ~- The interaction between CRIP 1 b and CB 1 was characterized using the yeast two-hybrid assay. Functional consequences of the CRIPlbCB 1 interaction were investigated by examining protein localization by confocal microscopy and measuring CB1 mediated N-type Ca2+ channel activity in the presence of CRIP 1 b by whole-cell patch clamp recordings. The ye_ast two-hybrid assay indicated that the last nine amino acids of the. CB 1 C-terminal tail were required for interaction with CRIP 1 b. Heterologous expression of CRIP 1 b and CB 1 in HEK 293 cells did not reveal evidence of co localization, nor was CB 1 able to significantly traffic CRIP 1 b to the plasma membrane. However, CRIP 1 b and CB1 were found to colocalize in superior cervical ganglion (SCG) neurons. Whole-cell voltage-clamp recordings ofN-type Ca2+ channels in SCG neurons indicated that CRIPlb had no.effect on agonist- or inverse agonist-induced modulation of Ca2 + current by CB1. Furthefrr1:ore, the level of CB 1 constitutive activity was· not significantly altered by CRIP 1 b. The high affinity of CB 1 for G proteins, as demonstrated by the ability of CB 1 to sequester G proteins from other Gu0 coupled receptors, was unaffected by expression ofCRIPlb. These results provide evidence that CRIP 1 b is a novel CB 1 accessory protein that interacts with the C-terminal tail of CB1. While CRIPlb and CB1 can apparently interact in a neuronal expression system, the ability of CRIP 1 b to modify CB 1 signaling was not detected in any of the pathways investigated. Thus, the distinctive signaling properties of CB1, such as constitutive activity and G protein seque.stration do not originate from nor are modified by CRIPlb.
    • Identification of insulin-responsive genes in rodent bone cells

      Blythe, Gregory A.; Department of Oral Biology (Augusta University, 1998-06)
    • Identification of Muscarinic Receptor Subtypes and Arachidonic Acid Metabolites Mediating the Effects of Actylcholine in The Pulmonary Circulation of Rabbits and Cats

      El-Kashef, Hassan; Department of Pharmacology (1987-05)
      Acetylch.oline (ACh) is known ·to reduce vascular pressure throughout the systemic. circulation; in. the·-- pulmonary circulatfon: ACh: can·- produce:either vasodi-1ationr or' ·vasoconstri-ction. For example,. ACh', produces: vasoconstrictfon · in· the, pulmonary ·circula~i on· of rabbits .. but dtlati on in· the· -feline. pu·lmof!ary-, vasculature·~-- The·· mechanism(s-) responsible·' for· the· actions of ACh in the pulmona.ry vasculature-- remain unc;lear. This study was designed to i·nve·st.igate poss·ible mechanisms. responsible for the actions. of ACh in the ra~bit ancf cat pulmonari_.circulation·~- Our hypothesi's is tbat . variations in· the· v.as.cular ·-response~ to: ACh may- be due to differe.nces in med-iators. (e.g. ·prostanoids)· released upon. activation .. of the muscarinic receptor~ These· di.ffe-rences cou 1 d be attributed,· either to. differences ... in muscarinic receptor subtypes.-. or to differences. in the·. metabolism of· arachidonic acid- tn rabbit. vs. cat. In anesthetized rabbits,_ ACh-induced pulmonary=- vasoconstriction-··. was,. totatly, inhibited::'·- by:: the: phosphotipasey A'2· inhibitor· qu-inacrine··,. the cycl o~oxygenase"' inhibitors:; indomethacin·,. and,.~ mec lofenamate·,, the- thtomboxane/ A:2:· synthetase=, i nh~i bi:tot· ]-;.;.{1'- Imidazolyl.)-· Heptano.fc. Acid .. (7;..IHA), and'- by,- the··' thromboxane·· A- 2 . receptor·· antagpntst. SQ:. 29-~.548:, .. but not·,. by·- the· 1 .ipoxygenase:-- inhibitor- nordihydrogua-iaret.ic: acid( NDGA}':. AGh·: s:igni-ftcantly; increased> plasma; thromboxane···B2.' (TXB'2J:· levels;. in·-. anesthetized:·,_ rabb_its:. A.Ch:~induced:. decrease'~ in, system.ic ar.teria:l pressure'" was, not· affected: by:·. any o.f'· the·· i nh'ibftors,. mentionedJ above--._ Small>. doses: of· the, select.i ve:· Mi- muscari nfc. receptor· subtype· antagonists:, pi renzepihe and_:. tr.ihexypheni:d~l,.. s:i gni-f.i cantly:- fnhi bjted:-: the· ACh.-i nduced:~- increase:'-·. i m pulmonary; vascuJa·r res:fstance- but. not: the~ ACh-induced=;· decrease Ht systemic. arterfal~: pressur.e··. The:.- s·ame" dose·· o.f secover.i ne:, .. a-~.- se·l ecti ve, M2-- muscar.i.nic: ii recept.or subtype. antagonist, dld ·not· change the · ACh ·effects on the pu 1 mona ry or· sys.temi c · v qSCu latu,re. Larger doses of pirenzepine, trihexyphen-idyl and\· secoverine,· totally. inhibited· the.- effects:: of ACh in· the .. pu:lmonary· and: systemic: circulatjons. Atrop.ine··. equally:- inh.i.bi ted:· the· effects. of ACn· on'" the pulmonary..- and.: systemic vascul atures- at.- any. do$es~ . . used': In .. fso_lated· rabbit·. lungs .. perfused·:·in:· situ~: w:ith · b]ood·-,_'free· medium;< . . . . . . . the ACh-fnduced· increases fn .. ·pulmonary v.ascular resistance, TXB2 level·s: a:nd ·6-Keto~PG·Fra were significa.nt.ly inhibited. by small concentrations.· of· pi.renzepine but .not· secove·rine. Larger concentrations. of· p.irenzepine~ and:'· · . . secover·ine totally ·inhibited· these· effects of ACh. In -anesthetized c~ts, the· ACh~inducec:l decre-ase in pulmonary vascular resfstance was partially~ · inhibited· by quinacrine and·' indomethacin and· significantly:· .potenti~ted by NOGA. The ACh-induced ·decrease in systemic arterial .pressure was not s·ignificantly affected by· any. of these· inhibitors. Small. doses. of secover-ine:'' but not . pfrenzepi ne;~" i nh.ibited·· the: pulmonary: but,. not .. the:· systemic vascular-. response·, tct.. ACh.,,_ i!!.=- v:ivo~ Howev-er,_. large··· doses; .. of·.· ptrenz:epi ne-, · tri·hexyphen:fdyl. and' s.ecoveri'ne' significantly:· inh.i bited~ the:, .. . . effects. of:. ACh~ in·.: the~: pulmonary;· and. systemic vascu:l ature\ In: isola ted:· cat: 1 ung_s:. perfused~- fm: sitw: w.ith; bJ ood.;.free- · medium~. 'ACh'" di-lated... the·· precons.tricted:: pulmonary: artery. bu.t> tt: did·: not ·s,tgnific.antly:· alter- TXB2 ori 6~ Keto~PGF 1 at 1 eveTs:~.. Smal:T: concentrat:i ons:- of:-· secoverfne-': but· not· pJ.renzep.i ne~-- par.t.ially;' i nhi bi'tedr the· ACh;.. induced~· decrease· in: pulinonar,Yvascular ·· resistance· •. The· pros.tacyc;ti n .. · synthetase fnhi bitor· tranylcypromine· si"gn.f$icantly~ i'nhi bited;'; the,· ACh- induced§ decrease:· in:· · pu]monary.:; va-scu..lar· resi.stance·~- . . i fi: ·These· results. ·indi-cate that l) ACh has . opposite actions in. the systemic {dilatory) versus pulmonary· (constrictor) circulation of rabbi-ts, 2r Arachidon-ic· acid.'metabo'lftes·. mediate. the-. pulmonary: . but. :not·· .the" systemic vascular respon.se- to. ACh, in rabbit, 3}. Thromboxane·· A2 mediates,. the:· pulmonary- vasoconstrictor- response·· to, ACh in rabbit·~ 4} The r.abbit pulmonary . v~scular· muscarinic recepto.rs. are· very·,· sensit:ive to·. p_irenzep.i-ne· . and thus be:have m9re · 1 ike Mi recept~rs, _5) · In· the cat·, the· ACh- induced·· decrease in- p~lmonary but not systemic vascular. pressure is partly mediated by prostacycli'n, 6)' In _the cat, the vascula-r· muscarinic receptors both in the· pulmon~ry. and the systemic beds are not selectively sensitive to . .pirenz.ep.ine ·and- thus behave like. non-Mt receptors and,· 7) In the .cat, the· ·muscarinic receptors. in the· pulmonary. and, systemic vasculat~re· represent different or heterogenous populations of M2. receptors since· they exhibit different affinities·. to secoverine.
    • Identification of professional competencies needed for practicing registered nurses in urban and rural hospitals

      Isler, Barbara A.; School of Nursing (1981-05)
      The purpose of this study was to determine if the hospital- location .s i gni fi cantly influenced the regis ter.ed nurses• competency needs .. A. sample of 112 urban and 80 rural .nurses was selected by Nursing Directors in 23 Georgia hospitals. Data were collected by use of a 113 item . competency questionnaire adapted from Clayton'·s (1978) work in thi.s area .. · Findings suggest the urban o~ rural hospital setting significantly . influences the competencies used by the registered nurse. Other findings include the acceptance percentage for ea-ch competency by the urban or. rural nurse indicating thei~ perception of nee~ and demographic characteristics of the two nurse populations.
    • Identification of Regulatory Elements in a Conserved Upstream Region of the Gene Encoding Interphotoreceptor Retinoid-Binding Protein (IRBP)

      Lu, Haiyan; Department of Ophthalmology (1999-06)
      (First Paragraph) IRBP is a large, single-subunit extracellular glycolipoprotein found in the interphotoreceptor matrix between the photoreceptor cells and retinal pigment epithelium cell layer (Fig. 1.). The protein is synthesized and secreted by the photoreceptor rods and cones, as well as pinealocytes, of all vertebrates. The molecular weight of human IRBP (1230 residues) is 133,400 daltons. This protein consists of four homologous segments of approximately 300 residues each. Each segment contains highly conserved hydrophobic domains among species. Ligands identified as bound to IRBP include retinoid isomers and fatty acids, and IRBP can also bind cholesterol, a-tocopherol and retinoic acid. The ability of IRBP to bind various retinoid isomers, fatty acids and many other hydrophobic ligands suggests multiple functions in the retina .
    • The Identification of the Requisite Knowledge and Relevant Role Activities by Nurse Administrators and the Relationship of These to Their Educational Preparation and Management Level

      Allen, Lori; Department of Nursing (1988-01)
      The purpose.of this descriptive correlational study was to identify the requisite knowledge and relevant role activities needed by nurse administrators to successfully enact their roles (first, mid, and top) and the relationship of these role knowledge areas and role.activities to their educational preparation and management level. To achieve the study purpose, two research questions and three hypotheses were tested. A total of 226 subjects participated in the study: 156 first-line nurse administrators; 44 mid level administrators.; and 26 top level nurse administrators. The study population consisted of a convenience sample of nurse administrators from five hospitals in a Southeastern metropolitan area. Data were collected through the use of the Nurse Administrative Role Aetivities ·and Knowledge (NARAA~) tool. Descriptive statistics were utilized to address the first research question. To test the hypptheses related to the second research question, ANOVA were utilized-. All three hypotheses were supported at the p=.05 level. More specifically, 13 role activities and four requisiteknowlege areas were found to be significantly different for first level .. and .top level administrators. The majority of these items were related to leadership, management, and administration of human resources. Seven role activities and 11 requisite knowledge areas were found to differ on the basis of educational preparation. Nurse administrators at each organizational level identified significantly different role activities·relevant for.their successful role enactment, but did not identify different requisite role knowledge areas. There was no significant difference in how nurses with different academic degrees rated the relevance of role activities to the success of their role enactment on the basis of academic degree. Academic degree did, however, discriminate among respondents in the degree to which they rated the relevance of requisite knowledge areas to the success of their role enactment. Other findings, limitations, implications, and suggestions for further study were discussed.
    • Identifying a vascoconstrictor role for interleukin-6, a pro-inflammatory cytokine.

      Daniels, Ericka R.; Medical College of Georgia (Augusta University, 2004-05)
      Increased plasma levels of interleukin-6 (IL-6) have been associated with elevated blood pressure and is predicative of risk for a firSt myocardial infarction. We therefore hypothesized that interleukin-6 has vasoconstrictive properties. Methods for this study included quantification of tissue levels of IL-6 , by capture ELISA from angiotensin 11- . I i hypertensivie male, C57BL/6J mice. Ti~sue coJcentratioti of IL-6 was compared to con~rol normotensiv¢ samples (thoracic aortic segments). ·Blood pressure was monitored by telemetry and the direct effect of IL-6 on v.ascu~ar contractility was studied in thoracic aortas isolated from normotensive mice. Our results show that IL-6 was significantly elevated (231.4 pg/mg ± 13.4 pg/mg . . ~ . . . SEM) in angiotensin II-hypertensiye mice compared to controls (129.9 pg/mg ± 11.2 · pg/mg SEM) and WT mice displayed a higher increase in MAP to angiotensin 11-infusion (60.35 mmHg ± 4.55 mmHg) at the end of infusion period compared to IL-6 KO mice (33.81 mmHg ± 6.67 mmHg). Pre-treatment of denuded aortic rings from normotensive mice with IL-6 for 24 hours caused a significant inhibition of their contractile response to phenylephrine and endothelin-1. In contrast, there was no significant effect on relaxation response to sodium niroprusside. In endothelium intact aortic rings, IL-6 caused contraction in the presence of L-NAME to inhibit nitric oxide synthase. We conclude that depending upon the experimental conditions, IL-6 has vasoconstrictive properties.
    • Identifying the Function of Nasal Embryonic LHRH factor (NELF) in Immortalized GnRH Neurons

      Ko, Eun Kyung; Department of Neuroscience and Regenerative Medicine (7/30/2018)
      Hypothalamic gonadotropin releasing hormone (GnRH) is crucial for the proper function of the hypothalamic-pituitary-gonadal (HPG) axis, puberty, and reproduction. When GnRH neuron migration or GnRH regulation is impaired, hypogonadotropic hypogonadism results. Mutations in the gene for nasal embryonic LHRH factor (NELF) have been identified in GnRH-deficient humans. NELF is a predominantly nuclear protein that may participate in gene transcription, but it is unlikely to be a transcription factor. Thus, our hypothesis is that NELF may indirectly regulate transcription via protein-protein interaction within a transcription complex. To address this question, RNA was extracted from NLT GnRH neuronal cells following either stable Nelf knockdown or scrambled control and subjected to cDNA arrays. Expression of transcription factors and cell migration gene expression was most commonly altered. Members of the Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway, including Stat1, Stat2, Stat5a, Jak2, Irf7 and Irf9, were significantly down-regulated as assessed by RT-qPCR. Protein levels of STAT1, phospho-STAT1, and JAK2 were reduced, but the levels of phospho-JAK2 were not. These findings suggest a role for NELF in the regulation of the JAK/STAT signaling pathway, which has important functions in GnRH neurons. Jacob, the rat orthologue of NELF, is phosphorylated at the serine 180 residue by phosphorylated Erk1/2 which is activated by synaptic NMDA receptor activation and then translocates to the nucleus. Phosphorylated Jacob in the nucleus interacts with CREB and regulates the expression of BDNF, which is associated with synaptic plasticity in the brain. Proteins, such as caldendrin, importin-α and α-interxin, have been identified as binding proteins with Jacob. However, binding proteins, phosphorylation sites and/or kinases for NELF have not yet been reported. To demonstrate novel and putative functions of NELF in the nucleus, identification of binding proteins and phosphorylation sites is required. To address this question, co-immunoprecipitation (Co-IP), mass spectrometry and in vitro kinase assays were performed. We found six putative binding proteins that could interact with NELF, including 14-3-3ε. We also identified phosphorylation sites on NELF, including serine 288, which could be phosphorylated by AKT1 kinase. These new findings will be helpful to understand the function of NELF in the nucleus
    • Immediate healing responses to ceramic endosseous dental implants in dog gingiva

      Long, William Gregory; Department of Oral Biology (1990-05)
    • Immune regulation of tumor cell plasticity: A promising molecular target in breast cancer metastasis

      LEE, EUNMI; Department of Biochemistry and Molecular Biology / Cancer Center (2018-11-29)
      It is widely accepted that phenotypic plasticity of malignant cells is required during metastatic cascade. However, the specific mechanism of how the tumor microenvironment regulates tumor cell plasticity in metastasis is under intense investigation. We demonstrate here that monocytic and granulocytic subsets of myeloid-derived suppressor cells (MDSC), hereafter called mMDSCs and gMDSCs, infiltrate in the primary tumor and distant organs with different time kinetics and regulate spatiotemporal tumor plasticity. Using co-culture experiments and mouse transcriptome analyses in syngeneic mouse models, we provide evidence that tumor-infiltrating mMDSCs facilitate dissemination from the primary site by inducing the EMT/CSC phenotype. In contrast, pulmonary gMDSC infiltrates support metastatic growth by reverting the EMT/CSC phenotype and promoting tumor cell proliferation. We also observe that lung-derived gMDSCs isolated from tumor-bearing mice enhance metastatic growth of already disseminated tumor cells. Our ongoing studies reveal that calprotectin (S100A8 and S100A9 heterotetramer) is an important regulator of gMDSCs, which play a critical role in promoting breast cancer metastasis by inducing MET-like CSCs as well as suppressing anti-tumor immunity within the pre-metastatic niche. Furthermore, we develop a novel gMDSC-targeting compound that potentially binds to calprotectin and validate its therapeutic utility in a preclinical breast cancer model. Our goal for this study is to elucidate the molecular co-evolution of tumor and immune cells in cancer development and to identify molecular targets to provide alternative therapeutic options for women with metastatic disease.
    • Immune Response of Hamsters to Adenovirus

      Boudet, Robert; Department of Cell and Molecular Biology (1970-06)
    • Immunohistochemical evaluation of labial salivary glands in xerostomic patient

      Ma, Yat-Ho; Medical College of Georgia (Augusta University, 2011-12)
    • Immunometabolic Regulation of Myeloid-Lymphoid Interactions Following Traumatic Brain Injury

      Braun, Molly; Biomedical Sciences (Augusta University, 2019-09)
      Traumatic brain injury (TBI) is a major public health issue, producing significant patient mortality and poor long-term outcomes. Increasing evidence suggests an important, yet poorly defined, role for the immune system in the development of progressive secondary injury. Herein, we tested the hypothesis that peripheral macrophage infiltration initiates long-lasting adaptive immune responses after TBI. Using a murine controlled cortical impact model, we found increased infiltration and pro-inflammatory (M1) polarization of macrophages for up to three weeks post-TBI. Monocytes purified from the injured brain stimulated the proliferation of naïve T lymphocytes, enhanced the polarization of T effector cells (Teff: TH1/TH17), and decreased the production of regulatory T cells (TREG) in a mixed lymphocyte reaction. Similarly, elevated Teff polarization within both blood and brain tissue was attenuated by myeloid cell depletion after TBI. Functionally, C3H/HeJ (TLR4 mutant) mice reversed both M1 macrophage and TH1/TH17 polarization after TBI, as compared to C3H/OuJ (wild-type) mice. Moreover, brain monocytes isolated from C3H/HeJ mice were less potent stimulators of T lymphocyte proliferation and TH1/TH17 polarization, as compared to C3H/OuJ monocytes. To further elucidate the mechanism underlying this myeloid TLR4-mediated Teff activation, we examined the metabolic regulator, 5’-adenosine monophosphate-activated protein kinase (AMPK), within infiltrating macrophages following TBI. We determined that reduced activation of myeloid AMPK induced the generation of pro-inflammatory, myelin reactive T-cells. Similarly, we detected myelin-laden macrophages within the cerebrospinal fluid of severe TBI patients. Administration of the AMPK activator, metformin, attenuated pro-inflammatory Teff cell generation and enhanced counter-inflammatory TREGs in wild-type mice; however, these effects were lost in myeloid-specific AMPKα1 knockout mice. Activation of AMPK restored myeloid expression and activity of Ten-eleven translocase 2 (TET2), a demethylase that regulated the expression of myeloid PD-L1 after TBI. Moreover, TET2-/- mice exhibited exaggerated T-cell activation in response to TBI. In line with these data, activation of myeloid AMPK reduced the loss of white matter and improved neurobehavioral outcomes after TBI. Our studies identify that immunometabolic dysfunction drives epigenetic regulation of the myeloid-lymphoid transition after TBI; suggesting targeted interventions during the early stage of injury may prevent progressive neurodegeneration.