Neutrophil gelatinase-associated lipocalin and associated damage markers in a chronic kidney disease model and an acute muscle injury model.

Date
2024-03
Journal Title
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Stellenbosch : Stellenbosch University
Abstract
ENGLISH ABSTRACT: Molecular markers of chronic pathology or acute tissue injury may be generalised or specific. Kidney and muscle are complex multicellular tissues predisposed to injury. The human immunodeficiency virus (HIV) causes kidney damage (KD) including neutrophil activation, as does acute exercise- induced muscle damage. This study aimed to assess the roles of matrix metalloproteinase 9 (MMP9), neutrophil gelatinase-associated lipocalin (NGAL) and fibroblast growth factor 21 (FGF21) in these conditions. Biobanked plasma and urine samples were obtained from 56 patients with confirmed KD and 10 healthy participants. Patient groups were HIV+ no KD, HIV⁺ with associated nephropathy (HIVAN), HIV⁺ with focal segmental glomerular sclerosis (FSGS) and KD-FSGS. Pathology data and laboratory data were collected from patients’ hospital records. Plasma and urine were analysed for MMP9, NGAL and FGF21. Immunohistochemistry staining was done on 6 kidney biopsies and 12 muscle biopsies to detect granulocytes (CD15), MMP9 and NGAL. The young healthy participants undertook plyometric exercise to induce acute muscle tissue damage. Blood was collected at baseline, post exercise, at 2 and 24 hours post exercise and analysed for the same biomarkers. Baseline and 24 hours post exercise muscle biopsies were stained using the same protocol as for the kidney biopsy samples. HIVAN patients had significantly higher urinary NGAL ng/mg compared to HIV-NKD (p<0.01), HIV-FSGS (p<0.05) and KD-FSGS (p<0.05). Similarly, plasma NGAL was significantly higher in HIVAN (p<0.05). Both plasma and urine MMP9 were variable in HIVAN and only significantly higher than HIV-NKD (p<0.05), not HIV-FSGS or KD-FSGS. HIVAN had a significantly higher urine MMP9/NGAL-complex ng.mg than HIV-NKD (p<0.05). Plasma MMP9/NGAL complex was significantly lower in the HIV-NKD group than in the HIVAN group and the HIV-FSGS group (p<0.01 and p<0.01 respectively). In the exercise study, NGAL and MMP9 were significantly higher post exercise than baseline (p<0.01 and p<0.05). Only MMP9 remained higher 2 hours post exercise (p<0.01). Elevations in NGAL after exercise-induced muscle damage did not reach the high levels seen with chronic KD, HIVAN. Similarly, p FGF21 was higher in HIVAN than at any time point after exercise. In contrast, MMP-9 for the HIVAN group was significantly lower than at 2 hours post exercise (p<0.05). MMP9 and NGAL colocalised in renal tissue of HIVAN participants. The overlap coefficient for MMP9 and NGAL was not significantly different to the other KD groups. In the HIVAN participants, the spatial ratio of MMP9 was significantly more than in the healthy control p<0.01. In HIVAN, the spatial ratio of NGAL was significantly more than healthy kidneys (p<0.01). In HIVAN, MMP9 and NGAL, colocalised more than healthy kidneys (p<0.001). Even in mild inflammation, the spatial ratio of MMP9 was more than in healthy kidneys (p<0.01) In muscle, the spatial ratio of MMP9 decreased significantly from baseline to 24 hours post exercise (p<0.05). In conclusion, NGAL, MMP9 and FGF21 are useful markers to differentiate between chronic diseases. In acute settings, NGAL, MMP9 and FGF21 increase. MMP9 and NGAL return to baseline levels after 24 hours. These markers are useful in following up on the recovery after an acute event.
AFRIKAANSE OPSOMMING: Molekulêre merkers van chroniese patologie of akute weefselbesering kan veralgemeen of spesifiek wees. Nier en spiere is komplekse meersellige weefsels wat geneig is tot besering. Die menslike immuniteitsgebreksvirus (MIV) veroorsaak nierbeskadiging (KD), insluitend neutrofiele aktivering, asook akute oefening-geïnduseerde spierskade. Hierdie studie het ten doel gehad om die rolle van matriksmetalloproteinase 9 (MMP9), neutrofiele gelatinase-geassosieerde lipokalien (NGAL) en fibroblastgroeifaktor 21 (FGF21) in hierdie toestande te assesseer. Plasma- en urinemonsters is verkry van biostoringsfasiliteite vir 56 pasiënte met bevestigde KD en 10 gesonde deelnemers. Pasiëntgroepe was MIV⁺ geen KD, MIV⁺ met gepaardgaande nefropatie (MIVAN), MIV⁺ met fokale segmentele glomerulêre sklerose (FSGS) en KD-FSGS. Patologiedata en laboratoriumdata is uit pasiënte se hospitaalrekords versamel. Plasma en urine is ontleed vir MMP9, NGAL en FGF21. Immunohistochemie-kleuring is gedoen op 6 nierbiopsies en 12 spierbiopsies om granulosiete (CD15), MMP9 en NGAL op te spoor. Die jong gesonde deelnemers het pliometriese oefening onderneem om akute spierweefselskade te veroorsaak. Bloed is by basislyn, na oefening, by 2 en 24 uur na oefening versamel en vir dieselfde biomerkers ontleed. Basislyn en 24 uur na oefening spierbiopsies is gevlek met dieselfde protokol as die nierbiopsiemonsters. MIV-pasiënte het aansienlik hoër urinêre NGAL ng/mg gehad in vergelyking met MIV-NKD (p<0.01), MIV-FSGS (p<0.05) en KD-FSGS (p<0.05). Net so was plasma NGAL aansienlik hoër in MIVAN (p<0.05). Beide plasma en urine MMP9 was veranderlik in MIVAN en slegs aansienlik hoër as MIV-NKD (p<0.05), nie MIV-FSGS of KD-FSGS nie. MIVAN het 'n aansienlik hoër urine MMP9/NGAL-kompleks ng.mg as MIV- NKD (p<0.05). Plasma MMP9/NGAL-kompleks was aansienlik laer in die MIV-NKD-groep as beide die MIVAN-groep en die MIV-FSGS-groepe (p<0.01 en p<0.01). In die oefenstudie was NGAL en MMP9 aansienlik hoër na-oefening as basislyn (p<0.01 en p<0.05). Slegs MMP9 het hoër gebly 2 uur na oefening (p<0.01). Verhogings in NGAL na oefening-geïnduseerde spierskade het nie die hoë vlakke bereik wat met kroniese KD, MIVAN, gesien is nie. Net so was plasma FGF21 hoër in MIVAN as enige tyd na oefening. Daarteenoor was MMP9 vir die MIVAN-groep laer as wat 2 uur na oefening gesien is. MMP9 en NGAL het in nierweefsel van MIVAN-deelnemers gekolokaliseer. Die oorvleuelingskoëffisiënt vir MMP9 en NGAL was nie beduidend anders as die ander KD-groepe nie. In die MIVAN-deelnemers was die ruimtelike verhouding van MMP9 aansienlik meer as in die gesonde nierbiopsies (p<0.01). So ook was die ruimtelike verhouding van NGAL aansienlik meer in MIVAN (p<0.01). Daarbenewens is MMP9 en NGAL, mede-gelokaliseerd tot ’n groter mate as in die MIVAN nierbiopsie as die gesonde nierbiopsie (p<0.001). Selfs in ligte inflamasie was die ruimtelike verhouding van MMP9 meer as in gesonde niere (p<0.01). In spiere het die ruimtelike verhouding van MMP9 aansienlik afgeneem van basislyn tot 24 uur na oefening (p<0,05). Ten slotte is NGAL, MMP9 en FGF21 nuttige biomerkers om tussen kroniese niersiektes te onderskei. In akute fisiologiese oefeningstres neem NGAL, MMP9 en FGF21 toe. MMP9 en NGAL keer na 24 uur terug na basislynvlakke. Hierdie merkers is nuttig om die herstel na 'n akute gebeurtenis op te volg.
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Thesis (PhD)--Stellenbosch University, 2024.
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