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- Item1000 Genomes-based metaanalysis identifies 10 novel loci for kidney function(Nature Research, 2017) Gorski, Mathias; Van Der Most, Peter J.; Teumer, Alexander; Chu, Audrey Y.; Li, Man; Mijatovic, Vladan; Nolte, Ilja M.; Cocca, Massimiliano; Taliun, Daniel; Gomez, Felicia; Li, Yong; Tayo, Bamidele; Tin, Adrienne; Feitosa, Mary F.; Aspelund, Thor; Attia, John; Biffar, Reiner; Bochud, Murielle; Boerwinkle, Eric; Borecki, Ingrid; Bottinger, Erwin P.; Chen, Ming-Huei; Chouraki, Vincent; Ciullo, Marina; Coresh, Josef; Cornelis, Marilyn C.; Curhan, Gary C.; d’Adamo, Adamo Pio; Dehghan, Abbas; Dengler, Laura; Ding, Jingzhong; Eiriksdottir, Gudny; Endlich, Karlhans; Enroth, Stefan; Esko, Tonu; Franco, Oscar H.; Gasparini, Paolo; Gieger, Christian; Girotto, Giorgia; Gottesman, Omri; Gudnason, Vilmundur; Gyllensten, Ulf; Hancock, Stephen J.; Harris, Tamara B.; Helmer, Catherine; Hollerer, Simon; Hofer, Edith; Hofman, Albert; Holliday, Elizabeth G.; Homuth, Georg; Hu, Frank B.; Huth, Cornelia; Hutri-Kahonen, Nina; Hwang, Shih-Jen; Imboden, Medea; Johansson, Asa; Kahonen, Mika; Konig, Wolfgang; Kramer, Holly; Kramer, Bernhard K.; Kumar, Ashish; Kutalik, Zoltan; Lambert, Jean-Charles; Launer, Lenore J.; Lehtimaki, Terho; De Borst, Martin H.; Navis, Gerjan; Swertz, Morris; Liu, Yongmei; Lohman, Kurt; Loos, Ruth J. F.; Lu, Yingchang; Lyytikainen, Leo-Pekka; McEvoy, Mark A.; Meisinger, Christa; Meitinger, Thomas; Metspalu, Andres; Metzger, Marie; Mihailov, Evelin; Mitchell, Paul; Nauck, Matthias; Oldehinkel, Albertine J.; Olden, Matthias; Penninx, Brenda W. J. H.; Pistis, Giorgio; Pramstaller, Peter P.; Probst-Hensch, Nicole; Raitakari, Olli T.; Rettig, Rainer; Ridker, Paul M.; Rivadeneira, Fernando; Robino, Antonietta; Rosas, Sylvia E.; Ruderfer, Douglas; Ruggiero, Daniela; Saba, Yasaman; Sala, Cinzia; Schmidt, Helena; Schmidt, Reinhold; Scott, Rodney J.; Sedaghat, Sanaz; Smith, Albert V.; Sorice, Rossella; Stengel, Benedicte; Stracke, Sylvia; Strauch, Konstantin; Toniolo, Daniela; Uitterlinden, Andre G.; Ulivi, Sheila; Viikari, Jorma S.; Volker, Uwe; Vollenweider, Peter; Volzke, Henry; Vuckovic, Dragana; Waldenberger, Melanie; Wang, Jie Jin; Yang, Qiong; Chasman, Daniel I.; Tromp, Gerard; Snieder, Harold; Heid, Iris M.; Fox, Caroline S.; Kottgen, Anna; Pattaro, Cristian; Boger, Carsten A.; Fuchsberger, ChristianHapMap imputed genome-wide association studies (GWAS) have revealed >50 loci at which common variants with minor allele frequency >5% are associated with kidney function. GWAS using more complete reference sets for imputation, such as those from The 1000 Genomes project, promise to identify novel loci that have been missed by previous efforts. To investigate the value of such a more complete variant catalog, we conducted a GWAS meta-analysis of kidney function based on the estimated glomerular filtration rate (eGFR) in 110,517 European ancestry participants using 1000 Genomes imputed data. We identified 10 novel loci with p-value < 5 × 10−8 previously missed by HapMap-based GWAS. Six of these loci (HOXD8, ARL15, PIK3R1, EYA4, ASTN2, and EPB41L3) are tagged by common SNPs unique to the 1000 Genomes reference panel. Using pathway analysis, we identified 39 significant (FDR < 0.05) genes and 127 significantly (FDR < 0.05) enriched gene sets, which were missed by our previous analyses. Among those, the 10 identified novel genes are part of pathways of kidney development, carbohydrate metabolism, cardiac septum development and glucose metabolism. These results highlight the utility of re-imputing from denser reference panels, until whole-genome sequencing becomes feasible in large samples.
- Item2'-5'-Oligoadenylate synthetase-like protein inhibits intracellular M. tuberculosis replication and promotes proinflammatory cytokine secretion(Elsevier, 2019-12) Leisching, G.; Ali, A.; Cole, V.; Baker, B.Host cytoplasmic surveillance pathways are known to elicit type I interferon (IFN) responses which are crucial to antimicrobial defense mechanisms. Oligoadenylate synthetase-like (OASL) protein has been extensively characterized as a part of the anti-viral mechanism, however a number of transcriptomic studies reveal its upregulation in response to infection with a wide variety of intracellular bacterial pathogens. To date, there is no evidence documenting the role (if any) of OASL during mycobacterium tuberculosis infection. Using two pathogenic strains differing in virulence only, as well as the non-pathogenic M. bovis BCG strain, we observed that pathogenicity and virulence strongly induced OASL expression after 24 h of infection. Further, we observed that OASL knock down led to a significant increase in M. tb CFU counts 96 h post-infection in comparison to the respective controls. Luminex revealed that OASL silencing significantly decreased IL-1β, TNF-α and MCP-1 secretion in THP-1 cells and had no effect on IL-10 secretion. We therefore postulate that OASL regulates pro-inflammatory mediators such as cytokines and chemokines which suppress intracellular mycobacterial growth and survival.
- Item2020-12-11 Global transcriptomic investigation of the human macrophage response towards pathogenic/non-pathogenic mycobacteria(Stellenbosch : Stellenbosch University, 2019-12) Mishra, Abhilasha Madhvi; Baker, Bienyameen; Leisching, Gina; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Molecular Biology and Human GeneticsBackground:Tuberculosis (TB) is a major cause of infection-related mortalityworldwide. In 2017 an estimated 1.3 million people who were HIV-negative died of TB. An estimated 5-10% of infected individual develop active TB during their lifetime, while the remaining90% (of infected population) successfully control the bacteria. Also, some of the close household contacts of TB patients remain uninfected and healthy. Studying host immune response towards Mycobacterium tuberculosis(M. tb) can unfold the reason behind this enigma. Methods:We conducted a detailed investigation of in vitrohost response from human monocyte derived macrophages(hMDMs)towards different strains of mycobacteria(grown in detergent-freemedia), i.e. pathogenic (M. tbR179) andnon-pathogenic (M. smegmatisand M. bovisBCG). The host response was measured post-infection (at mRNA and protein levels) using AmpliSeq, quantitative real time polymerase chain reaction (qRT-PCR), multiplex ELISA (Luminex), intracellular mycobacterial survivaland cytotoxicity assay. Biological network analysis (ingenuity pathway analysis IPA) was performed to understand the gene regulatory networkinvolved in the pathophysiology associated with the host-immune system.Based on false discovery rate (FDR) and biological functions, we selected an inter-related gene family of interferon induced protein with tetratricopeptides (IFIT1, IFIT2 andIFIT3) from the list of 19 potential differentially expressed genes(DEGs)for knock-up (vector-based over-expression)/down experiments. This gene family is known to form a protein complex during viral infection to act against the antigen. Studyencompassing their role against bacteria is not well established.Therefore, we performed knocking-up of IFITsvia vector-based transfection and knocking-down via small interferingRNA (siRNA) approach to investigate their effect upon mycobacteria inside the host macrophages. Results:AmpliSeqanalysis found 19 DEGs at 12 hours post-infection across all three strains. We observed lower number of mycobacterial CFUs and higher host response (at both RNA and protein level) in hMDMs infected with M. smegmatisas compared to other two strains. Biological network analysis revealed interferon-interleukin associated signalling pathways as most prominent among the 19 differentially expressed genes.We found a differed host response towardsall three strains, which mayattributeto their pathogenicity. Messenger RNA and protein level comparisons at different time points, depicted strong role of interferon and interleukin associated gene network. This network was able to successfully counter M. smegmatisbut succumb to M. bovisBCG andM. tbR179. Most importantly, across all three strains, intra-cellular bacterial growth and survival measured through colony forming units (CFUs)decreased significantly upon knocking up of IFITs(IFIT1, IFIT2 andIFIT3),while we recordedan increase in CFUs upon knocking down ofIFITsin the host macrophages. Using multiplex ELISA, we found higher expression of key pro-inflammatory cytokines (i.e. IDO1, IFN-γ, IL-6, and IL-23) during knock-up (vector-based over-expression)of IFITsresulting in reduction of mycobacteria. Conclusion:Differentially expressed IFITs showed a strong effect against mycobacteria, which can be used as a promising therapeutic targetadjunct to anti-TB therapy. This knowledge will broaden the scope of host drug targets for resistance free bacteriostatic immuno-therapy.
- ItemA 3-year cytogenetic survey of 9661 patients in South Africa(Health and Medical Publishing Group (HMPG), 1983) Bester, Rina; Benjamin, Mercy; Retief, A. E.; Bernstein, Renee; Grace, H. J.; Nelson, Matilda M.; Jansen, S.; Benjamin, MercyDuring the period 1 January 1977 - 31 December 1979, 9661 patients underwent cytogenetic investigation at seven participating laboratories in South Africa. The chromosome data were coded using a standard protocol and the results tabulated, being listed according to the clinical signs which led to referral for investigation. Cytogenetic investigation was most commonly requested for prenatal studies, and 22% of the group's effort was directed towards this. One in 27 amniotic cell specimens was reported to have shown anomalous chromosomes, trisomy 21 being the most frequent abnormality. The majority of postnatal investigations were requested because congenital abnormalities suggested an underlying chromosomal defect. In 42.3% of 2420 patients a chromosome defect was confirmed. Results of chromosome studies are tabulated by indication for referral and the findings summarized. This collaborative study gives an indication of the nature and frequency of chromosome disorders in South Africa.
- ItemAbnormalities of bone and mineral metabolism in patients with eating disorders(Stellenbosch : Stellenbosch University, 2001) Conradie, Maria Martha; Hough, F. S.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division Medical Physiology.ENGLISH ABSTRACT: Osteopenia is a well documented complication of anorexia nervosa (AN). The pathogenesis of this bone loss is presently poorly defined in the literature. Pathogenetic mechanisms that have been implicated include certain nutritional factors, exercise abuse, hypogonadism, hypercortisolism and/or vitamin 0 deficiency. We studied, 59 Caucasian eating disorder patients aged 15-45yr. The eating disorder was classified by a single, qualified psychiatrist according to OSM IV R criteria as either anorexia nervosa (AN: n =25), bulimia nervosa (BN: n = 17) or eating disorder not otherwise specified (EONOS: n = 17). All patients were subjected to a detailed dietary and general history. We assessed the prevalence and severity (OEXA), the nature (osteocalcin, deoxypyridinoline) and site (vertebral versus hip) of osteopenla in these patients. he role of nutritional factors (energy intake, weight, height, BMI, plasma albumin, lipids), physical activity, hypercortisolemia (plasma and urinary free cortisol), vitamin 0 deficiency (plasma 250HD) and hypogonadism (amenorrhoea, E2, LH, FSH) in the pathogenesis of bone loss were also evaluated. Mild osteopenia (BMO decreased by more than 1SO below age-matched controls) was documented in 46% of the total study population, with more marked osteopenia (Z-Score < -2 SO) present in 15%. Both vertebral and hip osteopenia were documented. In the study population those patients with AN (Lumbar BMO (q/cm") = 0.869 ± 0.121) were most likely to develop osteoporosis, although a significant percentage of patients with BN (Lumbar BMO (q/crn") = 0.975 ± 0.16) and EONOS (Lumbar BMO (g/cm2) = 0.936 ± 0.10) were also osteopenic (29% and 35% respectively). Twenty four percent (24%) of the total patient population had a history of fragility fractures. These fractures were reported more commonly amongst patients with AN and EONOS (28% and29.4%). Fracture prevalence was however similar in patients with normal and low bone mass. Conventional risk factors were similar in patients with normal and low bone mass, except for a significantly longer duration of amenorrhoea (p = 0.009), a lower BMI (p = 0.0001) and greater alcohol consumption (p = 0.05) in the osteopenic patients. Nutritional parameters (S-albumin, protein, Ca, and P04 intakes), physical activity, as well as 25(OH) vitamin D levels were similar in AN and BN subjects, as well as in patients with a low versus normal BMD. Plasma and urine cortisol levels were also similar in these subgroups. With the exception of two patients with borderline osteopenia, significant bone loss was only documented in those patients with a past or current history of amenorrhoea. In the total patient population the duration of amenorrhoea was significantly (p<0.009) longer in patients with osteopenia versus those with a normal bone mass. A significant negative correlation between BMD (Z-Score) and duration of amenorrhoea was also documented in the total patient population (r = -0.4, P = 0.001) as well as in all three eating disorder groups (AN r - -0.4, P = 0.03; BN r = - 0.6, P = 0.008; EDNOS r = -0.6, P = 0.005). In the total patient population, those patients with amenorrhoea, had lower BMD and BMI values and lower estrogen levels compared to those with a normal menstrual cycle. We conclude that osteopenia commonly attends AN, as well as BN and EDNOS. Nutritional (with the exception of alcohol consumption) and mechanical factors as well as hypercortisolemia did not appear to contribute significantly to bone loss in this study population. Hypogonadism appeared to be the main cause of the bone loss observed in these patients.
- ItemAccessory gene components for an HIV-1 subtype C vaccine : functional analysis of mutated Tat, Rev and Nef antigens(Stellenbosch : Stellenbosch University, 2002-12) Scriba, Thomas Jens; Van Rensburg, E. Janse; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Medicine.ENGLISH ABSTRACT: HIV has attained a global distribution and the number of infected people reached an estimated 28.1 million in sub-Saharan Africa at the end of 2001. HIV-1 subtype C is overwhelmingly prevalent in Botswana and South Africa and to date no interventions have been successful enough to curb the rapid spread of the virus. A number of HIV-1 vaccine strategies are being developed, however the breadth and efficacy of such candidate vaccines, many of which are based on the HIV-1 structural genes pol, gag and env, have mostly been found to be inadequate. The HIV-1 accessory genes are attractive components of HIV vaccines due to their role in viral pathogenesis, early expression and the high ratio of conserved CTl epitopes. Yet, because of undesirable properties questions regarding their safety as vaccine components are raised. In this study candidate tat, rev and nefmutants were assessed for efficient expression and inactivation of undesirable functionality. / Plasmid constructs that encode the South African HIV-1 subtype C consensus Tat, Rev and Nef proteins were constructed. The coding sequences of the genes were codon-optimised for optimum protein expression and these synthetic genes were constructed using overlapping 50-mer oligonucleotides. Furthermore, the proteins were mutated at previously described sites by PCR-based site-directed mutagenesis to render them inactive for their respective functions. Corresponding wild-type Tat, Rev and Nef constructs were also made from viral isolates that were least dissimilar to the respective consensus amino acid sequences. tn vitro expression of the different constructs were assessed in 293 cells by Western blotting with polyclonal mouse sera, which were generated by DNA immunisation with one of the Tat, Rev and Nef constructs. The transactivation activity of Tat variants and Rev-mediated nuclear export activity of RRE-containing transcripts were studied in cotransfection experiments using reporter-gene-based assays while Nef functionality was assessed in a cotransfection assay with subsequent flow cytometric analysis of surface CD4 and MHC-I expression on 293 cells. Sequence analysis of the South African HIV-1 subtype C consensus sequences of Tat, Rev and Nef revealed a high degree of similarity with a consensus sequence that was drawn up from a large number of viruses from southern Africa. These consensus sequences were also closer to individual viral isolate sequences than any individual sequences were, indicating that the use of a consensus sequence may serve to reduce genetic diversity between a vaccine and circulating viruses. Expression levels of the sequence-modified tat and nef gene constructs were not significantly higher than the wild-type constructs, however, the codon-optimised rev mutant exhibited markedly higher expression than the wild-type rev construct. Immunoreactivity of the protein with the mouse sera demonstrates expression and immunogenicity of the Tat, Rev and Nef immunogens in mice. In the background of the subtype C Tat, a single C22 mutation was insufficient to inactivate l TRdependent CAT expression in 293T and Hela cells. Yet, this activity was significantly impaired using the single mutation, C3?, or the double mutation, C22C3? Compared to the wild-type Rev, the function of the Rev with a double mutation, M5M10, was completely abrogated. Similarly, while the wild-type Nef and native, codon-optimised consensus Nef proteins mediated CD4 and MHC-I downregulation, CD4 downregulation was completely abrogated in one of the mutants, while both Nef mutants were entirely deficient for MHC-I downregulation. These data demonstrate the high expression levels and impaired functionality of sequence-modified HIV-1 subtype C consensus Tat, Rev and Nef DNA immunogens that may be used as single-standing vaccine components or form part of a multicomponent HIV-1 vaccine.
- ItemAccuracy and impact of the MTBDRplus v2 and MTBDRsl v2 line probe assays for the detection of first-line and second-line drug resistant tuberculosis(2023-02) Pillay, Samantha; Theron, Grant; de Vos, Margaretha; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics.ENGLISH ABSTRACT: Combating drug-resistant tuberculosis (DR-TB) remains a challenge globally. Treatment success rates are often derailed by under-diagnosis and under-reporting of disease. Patients remain contagious for prolonged periods prior to initiation of appropriate treatment which is further exacerbated by the amplification of drug resistance and poor treatment outcomes. Using current and new diagnostic tools effectively is key to rapid diagnosis of tuberculosis and early detection of drug resistance. Firstly, (chapter 2) line probe assays (LPAs) frequent inability to generate a resistance call in paucibacillary specimens is problematic. We showed that while MTBDRplus and MTBDRsl tests work well on smear-negative specimens for detecting drug resistance, failure rates remained high. We demonstrated with the use of routine key programmatic data how time-to-reporting of results improved with the use of molecular assays and provided evidence on how standard-of-care can be improved in a programmatic context. Secondly, (chapter 3) LPA testing on smear-negative specimens is not always performed causing diagnostic delays and hindering their role as a direct front-line diagnostic tests. Thus, by using Xpertgenerated data we determined the ratio of actionable-to-non-actionable results and the number of missed resistant cases at varying thresholds. We demonstrated that Xpert semiquantitation category is superior to informing reflex LPA testing than smear status. In short, this method provides a framework by which laboratories that currently do not test smear-negative specimens to expand testing. Thirdly (chapter 4) current pathways using Xpert MTB/RIF or Xpert Ultra as frontline tests for diagnosing TB and rifampicin resistance lack further treatment guidance. We did a systematic review and assessed the performance of Xpert MTB/XDR for the detection of pulmonary tuberculosis and resistance to isoniazid, fluoroquinolones, ethionamide, and amikacin. Participants consisted of 1228 for pulmonary tuberculosis detection and 1141 for drug resistance. We found Xpert MTB/XDR is unlikely to test positive as a follow-up test for the detection of Mycobacterium tuberculosis in samples that test Xpert Ultra
- ItemAccurate arterial path length estimation for pulse wave velocity calculation in growing children and adolescents(Gates Open Research, 2021-05) Witbooi, Lee-Roy C.; Page, Ben; Pitcher, Richard D.; Innes, SteveBackground: Most adult cardiovascular disease begins in childhood. Given the burgeoning obesity pandemic in children worldwide, there is a need for precise and scalable surveillance methods to detect subclinical cardiovascular disease in children and adolescents. Early detection allows early intervention and intensified primary prevention strategies in affected individuals. Carotid-femoral pulse wave velocity (PWV) directly measures arterial wall stiffness, an early feature of atherosclerosis. Calculation of PWV in growing children requires an accurate estimation of the true distance travelled by the aorto-femoral pressure wave, using surface anatomy landmarks. However, a variety of methods are used to estimate this distance, and these have not previously been investigated in growing children and adolescents. We sought to investigate this by comparing true arterial path length measured on computerized tomography (CT) scans, with a variety of estimations based on surface anatomy landmarks. Methods: Arterial path lengths were measured using multi-planar reformation (MPR) imaging software. These measurements were then compared with the surface anatomy measurements obtained using the same MPR imaging software. The fidelity of a variety of arterial path length estimation methods was tested. Results: The surface anatomy distance between the suprasternal notch and the angle of the mandible (PWV recording site in the neck), should be adjusted using the formula y=4.791+(1.0534*x). This value subtracted from the unadjusted distance from the suprasternal notch to the umbilicus, through the mid-inguinal crease to the femoral PWV recording site, provides the simplest reliable approximation of true intraluminal distance travelled. Conclusions: There is high correlation between the surface anatomy distances and the arterial path lengths they represent; however, these are not equal. Most surface anatomy measurements require adjustment using the formulae that we have provided, to accurately estimate the true distance travelled by the pulse wave.
- ItemAccurate estimation of large vessel length in growing children and adolescents for the purpose of pulse wave velocity calculation.(Stellenbosch : Stellenbosch University, 2018-12) Witbooi, Lee-Roy Cecil; Innes, Steve; Page, Ben; Pitcher, Richard; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Anatomy and Histology.Background Cardiovascular disease is a major cause of death in adults worldwide. Early detection allows for early intervention to prevent vascular events such as strokes, heart attacks, etc. Although these vascular events typically occur in late adulthood, the underlying atherosclerosis often begins during childhood. Early subclinical atherosclerosis can be detected by measuring the elasticity of the large arteries, particularly when performed serially over time. Normally, the elasticity of a healthy aorta helps to slow down the speed of the pressure wave created by contraction of the heart muscle. This is an important way of maintaining smooth laminar blood flow. Atherosclerosis causes the vessel wall to harden and lose elasticity. As the vessel wall hardens, the speed of the pressure wave increases. Pulse wave velocity (PWV) is a sophisticated method of detecting early elasticity changes, and is a preferred non-invasive technique to measure arterial wall stiffness. The velocity calculation requires accurate measurement of both distance travelled and time taken for the pulse wave to travel between two points. The distance used for pulse wave velocity calculation is an approximation of the intraluminal distance travelled by the pulse wave and is estimated by measuring the distance between various surface anatomy landmarks. The expert consensus document on arterial wall stiffness described carotid–femoral PWV as the “gold standard” measurement of arterial wall stiffness, yet there is no consensus on the arterial path length estimation method. A variety of arterial path length estimation methods exist, and this makes inter-study comparison of PWV very difficult. The purpose of the current study was to investigate the most accurate method of estimating the true distance travelled by the aorto-femoral pressure wave. We compared distances between a range of commonly used surface anatomy landmarks, and compared these to the true intraluminal distance measured on multi-planar reformations of archived computerized tomography imaging in children of varying ages. Our findings will allow standardization of PWV calculation in children and allow for inter-study comparisons. Methods Vessel lengths in children (aged 0-18 years) were measured with multi-planar reformation (MPR) imaging software. These measurements were then compared with the surface anatomy measurements also obtained using the MPR imaging software. The comparisons between vessel lengths and surface anatomy distances were performed in segments, since there were no whole body CT scans available on the Picture Archiving and Communication System (PACS) at the research site. Results The surface anatomy measurements from the suprasternal notch to the angle of the mandible (on the right) correlated well with the intraluminal vessel length from the origin of the brachiocephalic trunk to the external carotid at the angle of the mandible (r2=0.92; p<0.0001). The surface anatomy measurements from the suprasternal notch to the midpoint of the right inguinal crease, correlated well with the intraluminal vessel length from the origin of the brachiocephalic trunk to the right femoral artery at the right inguinal ligament (r2=0.98; p<0.0001). The surface anatomy measurements from the suprasternal notch to the xiphisternum, plus the surface distance between xiphisternum and the umbilicus, plus the surface distance between the umbilicus and the midpoint of the right inguinal crease, correlated well with the intraluminal vessel length from the origin of the brachiocephalic trunk to the right femoral artery at the right inguinal ligament (r2=0.97; p<0.0001). The surface anatomy measurement from the suprasternal notch to the xiphisternum, plus the surface distance between the xiphisternum and the midpoint of right inguinal crease, correlated well with the intraluminal vessel length from the origin of the brachiocephalic trunk to the right femoral artery at the right inguinal ligament (r2=0.97; p<0.0001). A regression equation is provided for each set of surface anatomy measurements, allowing further adjustment of measurements to more accurately represent the true intraluminal distance travelled by the pulse wave. Conclusions The surface anatomy distance between the suprasternal notch and the angle of the mandible, subtracted from the distance between the suprasternal notch and mid-inguinal crease, provides the closest approximation of true intraluminal distance travelled and would be the best method to standardize pulse wave velocity calculation in children and adolescents. However, surface anatomy estimations using the xiphisternum and umbilicus as landmarks produced very similar correlations.
- ItemAcrosome size and kinematics of human spermatozoa(Stellenbosch : University of Stellenbosch, 2007-03) Murray, George M.; Du Plessis, S. S.; Franken, D. R.; University of Stellenbosch. Faculty of Health Sciences. Dept. of Biomedical Sciences. Medical Physiology.For spermatozoa to gain access to the oocyte for fertilization, lytic enzymes need to be released during the acrosome reaction. These enzymes, which are stored and transported within an organelle termed the acrosome, make it possible for spermatozoa to collectively penetrate the layers of cells and glycoproteins that surround and protect an oocyte. Acrosomes may thus be viewed as essential for fertilization and their shape, size and volume were examined morphometrically by utilizing automated morphometric analysis equipment. In addition to the acrosome being necessary for normal unassisted fertilization, spermatozoa also need the ability to migrate to the oocyte. Following zona pellucida binding, sperm tail thrust movement initiates zona penetration into the space created by the digestive action of the acrosomal enzymes. Therefore the motion characteristics of spermatozoa were also quantified in terms of kinematic properties. In the treatment of male sub fertility, assisted reproductive techniques are applied. In the application of such techniques, a motile sub-population of spermatozoa was obtained by employing a procedure (swim-up selection) that selects cells on the basis of their kinematic ability. This study presents an analysis of the morphometric and kinematic qualities of spermatozoa populations that are subjected to swim-up selection and investigates the relationship of these morphometrical and kinematic qualities. Computer-assisted semen analysis, swim-up selection and automated sperm morphology analysis tests were all used to evaluate spermatozoa populations. Results indicated that, irrespective of acrosome size, higher kinematic parameter measurements were observed post-swim-up. A significant inverse relationship between the population’s average acrosome size and a number of kinematic parameters was observed. Our results indicated that for a post-swim-up population of spermatozoa an increase in the average acrosome size was significantly related to a decrease in the kinematic parameters VAP, VCL and the VSL within the same population.
- ItemAdipose tissue as a possible therapeutic target for polyphenols : a case for Cyclopia extracts as anti-obesity nutraceuticals(Elsevier, 2019) Jack, Babalwa U.; Malherbe, Christiaan J.; Mamushi, Mokadi Peggy; Muller, Christo J. F.; Joubert, Elizabeth; Louwa, Johan; Pheiffer, CarmenENGLISH ABSTRACT: Obesity is a significant contributor to increased morbidity and premature mortality due to increasing the risk of many chronic metabolic diseases such as type 2 diabetes, cardiovascular disease and certain types of cancer. Lifestyle modifications such as energy restriction and increased physical activity are highly effective first-line treatment strategies used in the management of obesity. However, adherence to these behavioral changes is poor, with an increased reliance on synthetic drugs, which unfortunately are plagued by adverse effects. The identification of new and safer anti-obesity agents is thus of significant interest. In recent years, plants and their phenolic constituents have attracted increased attention due to their health-promoting properties. Amongst these, Cyclopia, an endemic South African plant commonly consumed as a herbal tea (honeybush), has been shown to possess modulating properties against oxidative stress, hyperglycemia, and obesity. Likewise, several studies have reported that some of the major phenolic compounds present in Cyclopia spp. exhibit anti-obesity effects, particularly by targeting adipose tissue. These phenolic compounds belong to the xanthone, flavonoid and benzophenone classes. The aim of this review is to assess the potential of Cyclopia extracts as an anti-obesity nutraceutical as underpinned by in vitro and in vivo studies and the underlying cellular mechanisms and biological pathways regulated by their phenolic compounds.
- ItemThe adrenal cortex in hypercholesterolaemic rabbits. Histochemical and electron microscopical changes(Health & Medical Publishing Group, 1978-2) Rossouw, D. J.; Chase, C. C.; Engelbrecht, F. M.The adrenals of rabbits on a cholesterol-rich diet for 35 days showed histopathological changes, a marked increase in weight and a lowering in the ascorbate content. A focal increase in the neutral lipid and cholesterol content was noted mostly in the inner cortical zones; and a characteristic acid phosphatase-positive pattern in areas of infiltrating cells, and an alkaline phosphatase-positive reaction in heterophils in the infiltrated areas. Electron microscopy confirmed that the zona glomerulosa cells were relatively normal in hypercholesterolaemic rabbits, while necrosis and fibrosis were very obvious in the inner two zones. The cellular infiltrate was shown to consist of large, granular mononuclear cells, heterophils, eosinophils, stromal phagocytes, lymphocytes and plasma cells. The possibility that the reaction was of an immunological nature is considered. The morphology of the adrenals of rabbits which were on a cholesterol-rich diet for 35 days and on a normal diet for 6 weeks afterwards, was indistinguishable from that of those rabbits killed after 35 days on a cholesterol-rich diet.
- ItemAfrica-wide evaluation of host biomarkers in QuantiFERON supernatants for the diagnosis of pulmonary tuberculosis(Nature Research, 2018-02-08) Chegou, Novel N.; Sutherland, Jayne S.; Namuganga, Anna-Ritah; Corstjens, Paul L. A. M.; Geluk, Annemieke; Gebremichael, Gebremedhin; Mendy, Joseph; Malherbe, Stephanus; Stanley, Kim; Van Der Spuy, Gian D.; Kriel, Magdalena; Loxton, Andre G.; Kriel, Belinda; Simukonda, Felanji; Bekele, Yonas; Sheehama, Jacob A.; Nelongo, Josefina; Van Der Vyver, Marieta; Gebrexabher, Atsbeha; Hailu, Habteyes; Esterhuyse, Maria M.; Rosenkrands, Ida; Aagard, Claus; Kidd, Martin; Kassa, Desta; Mihret, Adane; Howe, Rawleigh; Cliff, Jacqueline M.; Crampin, Amelia C.; Mayanja-Kizza, Harriet; Kaufmann, Stefan H. E.; Dockrell, Hazel M.; Ottenhoff, Tom H. M.; Walzl, Gerhard; AE-TBC consortiumWe investigated host-derived biomarkers that were previously identified in QuantiFERON supernatants, in a large pan-African study. We recruited individuals presenting with symptoms of pulmonary TB at seven peripheral healthcare facilities in six African countries, prior to assessment for TB disease. We then evaluated the concentrations of 12 biomarkers in stored QuantiFERON supernatants using the Luminex platform. Based on laboratory, clinical and radiological findings and a pre-established algorithm, participants were classified as TB disease or other respiratory diseases(ORD). Of the 514 individuals included in the study, 179(34.8%) had TB disease, 274(51.5%) had ORD and 61(11.5%) had an uncertain diagnosis. A biosignature comprising unstimulated IFN-γ, MIP-1β, TGF-α and antigen-specific levels of TGF-α and VEGF, identified on a training sample set (n = 311), validated by diagnosing TB disease in the test set (n = 134) with an AUC of 0.81(95% CI, 0.76–0.86), corresponding to a sensitivity of 64.2%(95% CI, 49.7–76.5%) and specificity of 82.7%(95% CI, 72.4–89.9%). Host biomarkers detected in QuantiFERON supernatants can contribute to the diagnosis of active TB disease amongst people presenting with symptoms requiring investigation for TB disease, regardless of HIV status or ethnicity in Africa.
- ItemAlcohol, hospital discharge, and socioeconomic risk factors for default from multidrug resistant tuberculosis treatment in rural South Africa : a retrospective cohort study(PLoS, 2013-12-13) Kendall, Emily A.; Theron, Danie; Franke, Molly F.; Van Helden, Paul; Victor, Thomas C.; Murray, Megan B.; Warren, Robin M.; Jacobson, Karen R.Background: Default from multidrug-resistant tuberculosis (MDR-TB) treatment remains a major barrier to cure and epidemic control. We sought to identify patient risk factors for default from MDR-TB treatment and high-risk time periods for default in relation to hospitalization and transition to outpatient care. Methods: We retrospectively analyzed a cohort of 225 patients who initiated MDR-TB treatment between 2007 through 2010 at a rural TB hospital in the Western Cape Province, South Africa. Results: Fifty percent of patients were cured or completed treatment, 27% defaulted, 14% died, 4% failed treatment, and 5% transferred out. Recent alcohol use was common (63% of patients). In multivariable proportional hazards regression, older age (hazard ratio [HR]= 0.97 [95% confidence interval 0.94-0.99] per year of greater age), formal housing (HR=0.38 [0.19-0.78]), and steady employment (HR=0.41 [0.19-0.90]) were associated with decreased risk of default, while recent alcohol use (HR=2.1 [1.1-4.0]), recent drug use (HR=2.0 [1.0-3.6]), and Coloured (mixed ancestry) ethnicity (HR=2.3 [1.1-5.0]) were associated with increased risk of default (P<0.05). Defaults occurred throughout the first 18 months of the two-year treatment course but were especially frequent among alcohol users after discharge from the initial four-to-five-month in-hospital phase of treatment, with the highest default rates occurring among alcohol users within two months of discharge. Default rates during the first two months after discharge were also elevated for patients who received care from mobile clinics. Conclusions: Among patients who were not cured or did not complete MDR-TB treatment, the majority defaulted from treatment. Younger, economically-unstable patients and alcohol and drug users were particularly at risk. For alcohol users as well as mobile-clinic patients, the early outpatient treatment phase is a high-risk period for default that could be targeted in efforts to increase treatment completion rates.
- ItemAltered mitochondrial respiration and other features of mitochondrial function in parkin-mutant fibroblasts from parkinson’s disease patients(Hindawi Publishing Corporation, 2016) Haylett, William; Swart, Chrisna; Van der Westhuizen, Francois; Van Dyk, Hayley; Van der Merwe, Lize; Van der Merwe, Celia; Loos, Ben; Carr, Jonathan; Kinnear, Craig; Bardien, SorayaMutations in the parkin gene are the most common cause of early-onset Parkinson’s disease (PD). Parkin, an E3 ubiquitin ligase, is involved in respiratory chain function, mitophagy, and mitochondrial dynamics. Human cellular models with parkin null mutations are particularly valuable for investigating the mitochondrial functions of parkin. However, published results reporting on patient-derived parkin-mutant fibroblasts have been inconsistent. This study aimed to functionally compare parkin-mutant fibroblasts from PD patients with wild-type control fibroblasts using a variety of assays to gain a better understanding of the role of mitochondrial dysfunction in PD. To this end, dermal fibroblasts were obtained from three PD patients with homozygous whole exon deletions in parkin and three unaffected controls. Assays of mitochondrial respiration, mitochondrial network integrity, mitochondrial membrane potential, and cell growth were performed as informative markers of mitochondrial function. Surprisingly, it was found that mitochondrial respiratory rates were markedly higher in the parkin-mutant fibroblasts compared to control fibroblasts (p = 0.0093), while exhibiting more fragmented mitochondrial networks (). Moreover, cell growth of the parkin-mutant fibroblasts was significantly higher than that of controls (). These unanticipated findings are suggestive of a compensatory mechanism to preserve mitochondrial function and quality control in the absence of parkin in fibroblasts, which warrants further investigation.
- ItemAlternative insulin mitogenic signaling pathways in immature osteoblast cell lines(Stellenbosch : Stellenbosch University, 2002-03) Langeveldt, Carmen Ronel; Hulley, P. A.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Medicine.ENGLISH ABSTRACT: Insulin is a mitogen for many cells and commonly signals through the classical, mitogenic Raf- MEK-ERK or metabolic PB-kinase pathways. Insulin deficiency or type I diabetes causes severe osteopenia. Obese patients with type II diabetes or insulin resistance, a disease associated with defective insulin signaling pathways and high levels of circulating insulin, have increased or normal bone mineral density. The question of whether hyperinsul inemia preserves bone mass is frequently raised. However, there is still a lot of controversy on the role of insulin as an osteoanabolic agent and this question still remains unanswered. A critical role for insulin signaling in bone building osteoblasts has recently been demonstrated with IRS-l knock-out mice. These mice developed low-turnover osteopenia due to impaired proliferation and differentiation, stressing the importance of osteoblastic IRS-l for maintaining normal bone formation. In the present study it was found that insulin does function in vitro as an osteoblast mitogen. This was illustrated in three relatively immature osteoblast (MBA-15.4, -15.6 mouse and MG- 63 human) cell lines, which responded to insulin with significant increases in proliferation. In the MBA -15.4 preosteoblasts insulin stimulation of proliferation was comparable to the welldescribed mitogen, TPA. The UMR-I06 cell line expresses markers of differentiated osteoblasts, and was much less responsive to insulin treatment. The difference in proliferative potential may be due to differences between spontaneously transformed cell lines, or the stage of cell differentiation. UOI26, a MEKI/2 inhibitor and wortmannin, a PB-kinase inhibitor, were used to investigate the pathway used by insulin to signal and activate ERK and osteoblast proliferation. In MBA-15.4 mouse preosteoblasts, GF-containing FCS was completely dependent on MEK for DNA synthesis. In contrast, in both MBA-15.4 and more mature MBA-15.6 osteoblasts, insulininduced proliferation was resistant to the inhibitors alone or in combination. Higher MEKinhibitor concentrations had no effect, and proliferation was also increased by the inhibitors in several experiments. This indicated that the classical, insulin mitogenic pathway was not involved in MBA-15.4 proliferation. Wortmannin had no effect on either insulin- or 20% FCSstimulated proliferation, but inhibited activation of Akt/PKB, the metabolic downstream target of PI3-kinase. Insul in signal ing to ERK was both MEK-and PI3-kinase- dependent, but this had no effect on proliferation. In contrast, FCS-stimulated ERK activation and proliferation was almost completely dependent on MEK-ERK activation. Proliferative signaling in the MG-63 human osteoblastic cell line in response to insulin was partially dependent on MEK and partially dependent on PB-kinase. In contrast, signaling in response to the phorbol ester, TPA, was partially dependent on PI3K but totally dependent on MEK-ERK. This indicates that the signal converges on ERK, suggesting the involvement of a PB-kinase upstream of a dominant MEK-ERK pathway. The differences found here between mouse and human insulin mitogenic signaling pathways indicate that there may be species differences between osteoblast signaling pathways, with mouse cells being independent and human cells being dependent on MEK for DNA synthesis in response to insulin. The effects of glucocorticoids on insulin mitogenic signaling in osteoblasts were also investigated, because chronic long-term steroid use results in excessive bone loss. The PTP inhibitor, sodium orthovanadate, reversed GC-impaired TPA- and FCS- induced proliferation in MBA-1SA and MG-63 preosteoblasts. PTPs, such as SHP-l and PTP-IB, dephosphorylate and inactivate phosphorylated kinases. Both SHP-l and PTPlB associated with kinases in the mitogenic signaling cascade of MBA-lS.4 preosteoblasts growing rapidly in 10% FCS. Further, SHP-I co-irnmunoprecipitated with active, tyrosine phosphorylated ERK, which may indicate that it can dephosphorylate and inactivate ERK. However, since the MEK-ERK or PB-kinase pathways are not important in insulin-induced proliferation in mouse osteoblasts, the PTPs are unlikely to be role players in the negative regulation of this signaling pathway. This was confirmed by the finding that vanadate was unable to reverse GC-induced decreases in insulinstimulated DNA synthesis. This suggests that vanadate-sensitive PTPs may not be important in the negative regulation of insulin-induced mouse osteoblast proliferation, and provides further evidence of a novel insulin mitogenic pathway in the MBA-lSA but not MG-63 osteoblastic cell line.
- ItemAmeliorative potentials of quercetin against cotinine-induced toxic effects on human spermatozoa(Hainan Medical University Journal Publisher, 2016) Goss, Dale; Oyeyipo, Ibukun P.; Skosana, Bongekile T.; Ayad, Bashir M.; Du Plessis, Stefan S.Objectives: Cotinine, the principal metabolite of nicotine found in smokers' seminal plasma, has been shown to adversely affect sperm functionality while quercetin, a flavonoid with diverse properties is associated with several in vivo and in vitro health benefits. The aim of this study was to investigate the potential benefits of quercetin supplementation against damage caused by the by-products of tobacco smoke in human sperm cells. Methods: Washed human spermatozoa from 10 normozoospermic donors were treated with nutrient medium (control), quercetin (30 mmol/L) and cotinine (190 mg/mL, 300 ng/mL) with or without quercetin for 60 and 180 min incubation periods. Computer-aided sperm analysis was used to assess sperm motility while acrosomereacted cells were identified under a fluorescent microscope using fluorescein isothiocyanate-labelled Pisum Sativum Agglutinin as a probe, viability was assessed by means of a dye exclusion staining technique (eosin/nigrosin) and oxidative stress by flow cytometry using dihydroethidium as a probe. Values were expressed as mean ± S.E.M. as compared by ANOVA. Results: Higher cotinine concentrations reduced the number of viable cells after 60 and 180 min of exposure while viability of cells was increased in the cotinine aliquots supplemented with quercetin after 180 min of exposure when compared with cotinine only treated group. Conclusion: This study indicates that the ameliorating ability of quercetin on cotinineinduced decline in sperm function is associated with increased number of viable cells.
- ItemAminoglycoside-induced hearing loss : South Africans at risk(Health and Medical Publishing Group (HMPG), 2009) Bardien, Soraya; De Jong, Greetje; Schaaf, H. Simon; Harris, Tashneem; Fagan, Johan; Petersen, Lucretia
- ItemAMP kinase activation and glut4 translocation in isolated cardiomyocytes(Clinics Cardive Publishing, 2010-04) Webste, Ingrid; Friedrich, Sven O.; Lochner, Amanda; Huisamen, BarbaraActivation of AMP-activated protein kinase (AMPK) results in glucose transporter 4 (GLUT4) translocation from the cytosol to the cell membrane, and glucose uptake in the skeletal muscles. This increased activation of AMPK can be stimulated by a pharmacological agent, AICAR (5’-aminoimidazole-4-carboxamide ribonucleoside), which is converted intracellularly into ZMP (5’-aminoimidazole-4-carboxamideribonucleosidephosphate), an AMP analogue. We utilised AICAR and ZMP to study GLUT4 translocation and glucose uptake in isolated cardiomyocytes. Adult ventricular cardiomyocytes were treated with AICAR or ZMP, and glucose uptake was measured via [3H]-2-deoxyglucose accumulation. PKB/Akt, AMPK and acetyl-CoA-carboxylase phosphorylation and GLUT4 translocation were detected by Western blotting or flow cytometry. AICAR and ZMP promoted AMPK phosphorylation. Neither drug increased glucose uptake but on the contrary, inhibited basal glucose uptake, although GLUT4 translocation from the cytosol to the membrane occurred. Using flow cytometry to detect the exofacial loop of the GLUT4 protein, we showed ineffective insertion in the membrane under these conditions. Supplementing with nitric oxide improved insertion in the membrane but not glucose uptake. We concluded that activation of AMPK via AICAR or ZMP was not sufficient to induce GLUT4-mediated glucose uptake in isolated cardiomyocytes. Nitric oxide plays a role in proper insertion of the protein in the membrane but not in glucose uptake.
- ItemAMP kinase activation and glut4 translocation in isolated cardiomyocytes(http://www.cvja.co.za/index.php, 2010-03) Webster, Ingrid; Huisamen, Barbara; Lochner, Amanda; Friedrich, Sven O; Biomedical Sciences: Medical Physiology