Investigating differential non-oxidative glucose-utilizing pathway gene expression as a novel diagnostic tool for type 2 diabetes

Thesis (MSc)--Stellenbosch University, 2013.ENGLISH ABSTRACT: Context: Despite the availability and accessibility of current diagnostic tools, diabetes remains largely under-diagnosed. Biological limitations, discordant assays and conflicting diagnostic thresholds together impede the accurate and successful diagnosis of diabetes, providing impetus into research for a novel diagnostic tool. Aim: Since flux through the five minor glycolytic pathways is increased during hyperglycemia, we hypothesized that the genes encoding the regulatory enzymes of such pathways may be differentially expressed between control, pre-diabetic and diabetic individuals setting the scene for an exploratory diagnostic avenue employing genetic biomarkers. Experimental procedures: Participants (n=60; n=20 Mixed Ancestry, n=40 Caucasian) were recruited from Stellenbosch and Paarl (Western Cape, South Africa) and classified as control, pre-diabetic or diabetic. RNA was purified from leukocytes isolated from blood samples and OGT, OGA, GFPT1, GFPT2, TKT, TKTL1 and AKR1B1 expression determined by quantitative real-time PCR. Results: Expression of OGA, OGT, GFPT2 and TKTL1 decreased in pre-diabetic and diabetic individuals; while GFPT1, TKT and AKR1B1 expression levels remained largely unaffected between the study groups. GFPT2 exhibited ethnic-dependent regulation. Conclusion: Differential expression of genes regulating non-oxidative glucose-utilizing pathways may offer diagnostic utility in the future and warrant further investigation.AFRIKAANSE OPSOMMING: Konteks: Teen spyte van die beskikbaarheid en toeganklikheid van huidige diagnostiese instrumente, word diabetes nogsteeds min gediagnoseer. Altesaam belemmer biologiese beperkings, teenstrydige toetse en botsende diagnostiese grense die akkurate en suksesvolle diagnose van diabetes, wat bydra tot die druifkrag in navorsing vir ‘n nuwe diagnostiese instrument. Doelstelling: Aangesien die vloei deur die 5 mineur glikolitiese pad weë toeneem gedurende hiperglukemie, veronderstel ons dat die gene wat regulatoriese ensieme kodeer in hierdie pad weë mag dalk differensieel uitgedruk word tussen kontrol, voor- en, diabetiese individue wat die toneel skep vir ‘n ondersoekende diagnotiese laan wat gebruik maak van genetiese merkers. Materiale en metodes: Deelnemers (n=60; n=20 Gemengde afkoms; n=40 Blankes) was uit Stellenbos en die Paarl gewerf (Wes-Kaap, Suid-Afrika) en geklassifiseer as ‘n kontrol, voorof diabeties. RNS was uit witbloodselle gesuiwer wat eers uit bloed monsters geisoleer was en OGT, OGA, GFPT1, GFPT2, TKT, TKTL1 en AKR1B1 uitdrukking was bepaal deur kwantitatiewe RT-PKR. Resultate: Uitdrukking van OGA, OGT, GFPT2 en TKTL1 het afgeneem in voor- en diabetiese individue; terwyl GFPT1, TKT en AKR1B1 utidrukkings vlakke meestal onaangeraak was tussen die studie groepe. GFPT2 het etiese-afhanglike regulasie vertoon. Gevolgtrekkings: Differensiele uitdrukking van gene wat glukose gebruik in nieoksidatiewe pad weë reguleer bied diagnotiese gebruikbaarheid in die toekoms en bevel verdere ondersoek

Differential hexosamine biosynthetic pathway gene expression with type 2 diabetes

The hexosamine biosynthetic pathway (HBP) culminates in the attachment of O-linked β-N-acetylglucosamine (O-GlcNAc) onto serine/threonine residues of target proteins. The HBP is regulated by several modulators, i.e. O-linked β-N-acetylglucosaminyl transferase (OGT) and β-N-acetylglucosaminidase (OGA) catalyze the addition and removal of O-GlcNAc moieties, respectively; while flux is controlled by the rate-limiting enzyme glutamine:fructose-6-phosphate amidotransferase (GFPT), transcribed by two genes, GFPT1 and GFPT2. Since increased HBP flux is glucose-responsive and linked to insulin resistance/type 2 diabetes onset, we hypothesized that diabetic individuals exhibit differential expression of HBP regulatory genes. Volunteers (n = 60; n = 20 Mixed Ancestry, n = 40 Caucasian) were recruited from Stellenbosch and Paarl (Western Cape, South Africa) and classified as control, pre- or diabetic according to fasting plasma glucose and HbA1c levels, respectively. RNA was purified from leukocytes isolated from collected blood samples and OGT, OGA, GFPT1 and GFPT2 expressions determined by quantitative real-time PCR. The data reveal lower OGA expression in diabetic individuals (P < 0.01), while pre- and diabetic subjects displayed attenuated OGT expression vs. controls (P < 0.01 and P < 0.001, respectively). Moreover, GFPT2 expression decreased in pre- and diabetic Caucasians vs. controls (P < 0.05 and P < 0.01, respectively). We also found ethnic differences, i.e. Mixed Ancestry individuals exhibited a 2.4-fold increase in GFPT2 expression vs. Caucasians, despite diagnosis (P < 0.01). Gene expression of HBP regulators differs between diabetic and non-diabetic individuals, together with distinct ethnic-specific gene profiles. Thus differential HBP gene regulation may offer diagnostic utility and provide candidate susceptibility genes for different ethnic groupings

UMW Theatre Remote Production of William Shakespeare\u27s Much Ado About Nothing

Beatrice and Benedick are meant for each other. Trouble is, they don’t see it that way. In one of Shakespeare’s wittiest and most romantic of comedies, mistaken identities, misdirected insults, devious fakery, and bumbling antics prove no match for the effervescent power of love. Will calculated swooning and conniving mischief succeed to find Beatrice and Benedick falling madly for each other, or will it all simply amount to Much Ado About Nothing? When campus closed on March 12 in order to transition to online learning in light of the rise of COVID-19, UMW Theatre was deep into preparation for the final offering of it’s 2019-20 season, Much Ado About Nothing by William Shakespeare. When there was a chance that we might return to in person classes on April 6, the faculty decided to put the production on hold and make arrangements for a delayed opening once classes resumed. When UMW announced that classes would remain online for the remainder of the academic year, the faculty and staff decided to present a performance of Much Ado About Nothing online. Gregg Stull, professor of theatre and chair of the department met with the Much Ado company on March 19 to share the plans. Rehearsals resumed on March 23. The cast presented a live stream performance on April 16 to a crowd of more than 1500 viewers watching from 37 states, Puerto Rico, the District of Columbia and five states. Within 24 hours of being posted as an on demand performance on YouTube, more than 2000 people have watched the UMW Theatre production of Much Ado. From the start of the production process until the performance, 107 UMW students contributed to bringing Much Ado About Nothing to life