A histomorphometric study of the effect of Afriplex GRT(TM) on the pancreas, liver and kidney of rats fed a high-fat diet

Layman, Jodie (2018-12)

Thesis (MSc)--Stellenbosch University, 2018.

Thesis

ENGLISH ABSTRACT: Obesity causes a wide range of disorders including insulin resistance, hepatic steatosis as well as renal glomerular and tubular pathology. These disorders are typically associated with oxidative stress and inflammation. Rooibos, Aspalathus linearis, is a South African plant which has antioxidative, anti-inflammatory, antilipidemic, and antihyperglycemic properties as shown in vivo and in vitro. Afriplex GRT™ is an unfermented green rooibos extract which is postulated to have enhanced effects compared to fermented rooibos, mostly due to its high aspalathin content. The present study aimed to determine the ameliorative effects of Afriplex GRT™ on the histomorphometry of the pancreas liver and kidney in a diet-induced obese rat model. Male Wistar rats (N=28) were randomly sorted into four study groups (n=7) namely, control (C), GRT fed control (GRT), high-fat diet (HFD) and HFD supplemented with GRT (HFDGRT). Animals were either fed a control feed or a HFD which contained fat, cholesterol and fructose. Animals in the GRT and HFD-GRT groups were supplemented with Afriplex GRT™ at a concentration of 60 mg per kg body mass per day. Body mass, and pancreatic, liver and kidney mass were determined after which samples were processed to wax. Tissues were sectioned at 4 – 5 µm in thickness and stained using haematoxylin and eosin for analysis by a histopathologist. Immunohistochemistry using anti-insulin and anti-glucagon double antibody labelling was performed on the pancreas. Other staining techniques included Masson’s trichrome, Gordon and Sweet’s reticulin impregnation and periodic acid Schiff. Additionally, frozen sections of the liver and pancreas were stained using Oil red O for lipid infiltration. Morphometric techniques included area, diameter, radii, point counting and ratios. Body mass significantly decreased in the HFD-GRT group compared to the HFD group. Pancreas and kidney mass decreased significantly in the HFD-GRT group compared to the Cgroup, however liver mass significantly increased in both HFD groups. Islet area and β-cell area significantly decreased in the HFD-GRT group compared to all three control groups however, a greater number of islets was observed. Hepatic steatosis and liver pathology were reduced in the HFD-GRT group compared to the HFD group. Renal space and the proximal convoluted tubules were decreased in the HFD-GRT compared to C-group as indicated by trends. Additionally, a trend was observed in the arcuate artery media to lumen ratio which was increased in the GRT group compared to the HFD group. Similar to the present study, a decrease in body mass has been observed with fermented rooibos and its polyphenols in HFD-fed animals. In the HFD-GRT group, increased islets per section area and smaller islet areas are associated with increased insulin secretion, which suggests improved glucose tolerance. This study suggests that Afriplex GRT™ may effectively reduce hepatic steatosis, an effect previously seen in studies on fermented rooibos and rooibos polyphenols. More in-depth studies on the effects of Afriplex GRT™ in the kidney are needed, potentially for a longer study period. Therefore, Afriplex GRT™ has observable ameliorative effects on the histomorphology of the liver and the pancreas; the effects on the kidney requires further investigation.

AFRIKAANSE OPSOMMING: Vetsug veroorsaak 'n groot verskeidenheid van afwykings, wat insluit insulienweerstandigheid, hepatiese steatosis, asook renale glomerulêre en tubulêre patologie. Hierdie afwykings word tipies geassosieer met oksidatiewe stres en inflammasie. Rooibos, Aspalathus Linearis, is 'n Suid-Afrikaanse plant wat anti-oksidatiewe, anti-inflammatoriese, anti-lipidemiese en antihiperglukemiese eienskappe toon, volgens in vivo en in vitro studies. Daar word gepostuleer dat Afriplex GRT™, wat 'n ongefermenteerde groen rooibos uittreksel is, ‘n versterkte effekt toon in vergelyking met gefermenteerde rooibos, hoofsaaklik as gevolg van die hoë aspalatien inhoud. Die doel van die huidige studie was om die verbeterende effekte van Afriplex GRT™ op die histomorfometrie van die pankreas, lewer en nier in ‘n dieet-geïnduseerde obesitiet rotmodel vas te stel. Manlike Wistar rotte (N = 28) was verdeel in vier studiegroepe (n = 7) naamlik, kontrole groep (K), kontrole GRT groep (GRT), hoëvet diet groep (HVD) en HVD met GRT groep (HVDGRT). 'n Beheer voer of 'n HVD (wat vet, cholesterol en fruktose bevat) was vir rotter gevoer. Die voer van rotte in die GRT en HVD-GRT groepe was aangevul met Afriplex GRT™, met 'n konsentrasie van 60 milligram per kg liggaamsmassa per dag. Liggaamsmassa as ook pankreas, lewer en nier massa was bepaal, waarna weefsel in was verwerk was. Weefsels was gesny teen ‘n 4 - 5 μm dikte en gekleur met hematoksilien en eosien vir analise deur 'n histopatoloog. Immunohistochemiese kleuring vir anti-insulien en anti-glukagon was op die pankreas uitgevoer. Masson’s trichrome, Gordon en Sweet’s retikulien saturasie, en periodic acid Schiff was ook gedoen. Verder was bevrore snitte van die lewer en pankreas met Oil red O gekleur vir lipied infiltrasie. Morfometriese tegnieke het ingelsuit in area, deursnee, radius, punt telling en verhoudings. Liggaamsmassa het aansienlik afgeneem in die HVD-GRT-groep in vergelyking met die HVD-groep. Pankreas- en niermassa het aansienlik afgeneem in die HVD-GRT-groep in vergelyking met die K-groep, maar die lewermassa het aansienlik toegeneem in beide HVDgroepe. In die HVD-GRT-groep het die eiland- en beta-selgebied aansienlik afgeneem in vergelyking met al drie beheergroepe, maar 'n groter aantal eilandjies is waargeneem. Hepatiese steatosis en lewerpatologie was in die HVD-GRT-groep verlaag in vergelyking met die HVD-groep. Renale spasie en die proksimale kronkelbuisies is in die HVD-GRT verlaag in vergelyking met K-groep soos aangedui deur tendense. Daarbenewens was 'n tendens waargeneem in die arcuate arterie media tot lumen verhouding wat in die GRT groep vergroot is in vergelyking met die HVD groep. 'n Vermindering in liggaam gewig, vergelykbaar met die huidige studie, was geassosieer met gefermenteerde rooibos en sy polyfenole in die HVDgevoerde diere. In die HVD-GRT-groep het die toenemende eilandjies per area en aantal kleiner eilandjies per area gelei tot ‘n verhoogde insulienafskeiding, wat verbeterde glukosetoleransie aandui. Hierdie studie dui daarop dat Afriplex GRT™ hepatiese steatosis effektief kan verminder, 'n effek wat voorheen gesien is in studies oor gefermenteerde rooibos en rooibos polifenole. Meer in-diepte studies oor die gevolge van Afriplex GRT™ in die nier is nodig, ideal vir 'n langer tydperk. Dus, Afriplex GRT™ het waarneembare verbeterende effekte op die histomorfologie van die lewer en die pankreas. Hierdie effekte op die nier vereis verdere ondersoek.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/105195
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