Research Article: Detecting early kidney damage in horses with colic by measuring matrix metalloproteinase -9 and -2, other enzymes, urinary glucose and total proteins

Date Published: January 23, 2007

Publisher: BioMed Central

Author(s): Bela M Arosalo, Marja Raekallio, Minna Rajamäki, Elina Holopainen, Tuulia Kastevaara, Hanna Salonen, Satu Sankari.


The aim of the study was to investigate urine matrix metalloproteinase (MMP-2 and -9) activity, alkaline phosphatase/creatinine (U-AP/Cr) and gamma-glutamyl-transpeptidase/creatinine (U-GGT/Cr) ratios, glucose concentration, and urine protein/creatinine (U-Prot/Cr) ratio and to compare data with plasma MMP-2 and -9 activity, cystatin-C and creatinine concentrations in colic horses and healthy controls. Horses with surgical colic (n = 5) were compared to healthy stallions (n = 7) that came for castration. Blood and urine samples were collected. MMP gelatinolytic activity was measured by zymography.

We found out that horses with colic had significantly higher urinary MMP-9 complex and proMMP-9 activities than horses in the control group. Colic horses also had higher plasma MMP-2 activity than the control horses. Serum creatinine, although within reference range, was significantly higher in the colic horses than in the control group. There was no significant increase in urinary alkaline phosphatase, gamma-glutamyltranspeptidase or total proteins in the colic horses compared to the control group. A human cystatin-C test (Dako Cytomation latex immunoassay® based on turbidimetry) did not cross react with equine cystatin-C.

The results indicate that plasma MMP-2 may play a role in the pathogenesis of equine colic and urinary MMP-9 in equine kidney damage.

Partial Text

Usually the detection of kidney damage can be made at a relatively late stage by measuring serum creatinine and urea concentrations because at the time of abnormal values 75% of nephrons are already damaged [1]. However, depending on the cause and extend of damage, kidney has ability to recover [2-5]. There is a need for tests that can measure the early stages of kidney damage when there is still a chance to prevent further damage. In the studies made with laboratory animals the release of endotoxins can affect kidney function [6,7]. In severe equine colic the release of endotoxins is presumed to happen, but to our knowledge there are no studies on effects of endotoxins on equine kidneys. The routine use of nephrotoxic drugs like gentamicin, and non-steroidal anti-inflammatory drugs preoperatively can further increase risk of damage to the kidneys. For instance, even though gentamicin nephrotoxicity appears to be related to high or low levels of the drug, it can occur at recommended doses. Additionally, dehydration, and other drugs can affect the toxicity of gentamicin. Moreover, a deterioration in renal function may become evident for the first time or may progress after the drug has been discontinued [8]. Gunson et al. [9] reported that horses with both water deprivation and concurrent administration of the non-steroidal drug developed acute necrosis of the renal papilla, whereas the groups of horses that received either non-steroidal drug or water deprivation alone did not develop kidney damage.

Plasma MMP-2 activity was higher in colic horses (p = 0.009) compared to the control group. (Fig. 1)

The horses with colic in this study had been administered potentially nephrotoxic drugs such as NSAIDs, and gentamicin and they were most probably also endotoxemic before the operation. Matrix metallo proteinases may play a role in the pathogenesis of endotoxemia. Raulo et al. [24] inoculated cows with endotoxin and detected an increase in MMP-2 activity in milk after two hours and an increase in capillary permeability, evidenced first by the penetration of small molecular weight proteins, and 6 to 12 hours post endotoxin inoculation, by neutrophilic leucocytes. Albert et al. [25] showed that in humans plasma activity of MMP-9 increased 30-fold in 2 hours after endotoxin stimulation and fell after 4 to 6 hours. In our study, plasma MMP-2 activity increased in all colic horses. MMP-2 is related to changes in cell permeability which is presumed to happen in colic. Molecules that are smaller than 68 kDa can pass through glomerular filtration. But in most healthy animals the proteins are reabsorbed in the proximal tubules and, thus, very little or no protein is detected in urine samples [1]. ProMMP-2 has a weight of 72 kDa and MMP-2 a weight of 59–62 kDa. Inactive proMMP-9 weighs 92 kDa, but after activation it weighs between 68 and 82 kDa [26]. In our study, urine MMP-9 complexes and both proMMP-9 and MMP-9 activities increased in colic horses which can be related to early tubular damage because these molecules cannot pass through glomerular basement membrane due to high molecular weight. Further studies need to be done to determine the time span over which gelatinolytic activity increases in acute tubular damage.

The results indicate that plasma MMP-2 may play a role in the pathogenesis of equine colic and urinary MMP-9 in equine kidney damage.

The author(s) declare that they have no competing interests.

BA participated in performing computer analysis on MMP, analyzing the data and drafting the manuscript. Marja R participated in the design of the study and performed the statistical analysis and helped to draft the manuscript. Minna R participated in supervising MMP analysis. EH and TK collected blood and urine samples and clinical data and helped to draft the manuscript. HS participated in the design of the study. SS carried out other laboratory assays and revised the manuscript.




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