Renal transplant fibrosis: Histomorphometric assessment of early renal transplant biopsies for markers of chronic rejection

ELSEVIER

Renal Transplant Fibrosis: Assessment of Early Renal Transplant Biopsies for Markers of Chronic Rejection M.L. Nicholson, S.J. Harper, T.J. Wheatley, T.A. McCulloch, J. Feehally, and P.N. Furness

L

ATE renal allograft graft failure is most commonly due to the process of chronic rejection.’ Chronic allograft damage occurs slowly and effective study of this problem would be enhanced by developing surrogate and interim end points for analysis. In this study, allograft fibrosis has been analysed using a histomorphometric method to assess the volume fraction of the interstitium and used as a surrogate marker of chronic graft damage.

lOll-

80 Graft survival(%)

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60-

>25% fibrosis (n=47) ~25% fibrosis (n=24)

PATIENTS AND METHODS A consecutive series of 107 renal transplants were studied. All patients received cyclosporin-(CsA) based immunosuppression either as dual therapy (CsA 17 mg/kg/d reducing to 5-7 mgkglday over 6 weeks plus reducing doses of prednisolone) or as triple therapy (CsA at an initial dose of 10 mg/kg/d reducing to 5 mg/kg/d over 6 weeks plus azathioprine 1 mg/kg/d plus reducing doses of prednisolone). Needle core transplant biopsies were performed preperfusion and at 1, 6 and 12 months posttransplant. Renal function was assessed by glomerular filtration rate measurements at the same time intervals. Graft fibrosis was assessed using a histomorphometric method: the microscope slide image was projected onto a wall mounted matrix consisting of 200 points. Twenty-five points from each of 8 randomly selected fields were counted for each biopsy and the interstitial volume fraction (IVFr) was calculated as the ratio of the number of points overlying the

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4oi 0

PzO.04

I 6

1 12

I I 18 24

I 30

1 I 36 42

I 48

I 54

I 60

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8 72

Time post-transplant (months)

Fig 2.

Graft survival from time of 6 month transplant

biopsy.

interstitium to the total number of points counted, expressed as a percentage. This method gave a standard error of the mean of less than 10%. Intra-observer error was also less than 10%. Groups were compared using a two-tailed student’s t-test. Differences were considered significant when P < .05. Kaplan-Meier graft survival curves were constructed and differences compared using the logrank statistic.

RESULTS

IVFr increased significantly at 1 and 6 months posttransplant, but then stabilized. There was no further rise by 12 months (Fig. 1). GFR was negatively correlated with IVFr at 6 months posttransplant (P < .OS). There were no statistical differences between IVFr measured in the high(dual) and low-(triple) dose CsA groups. For renal allografts surviving 6 months posttransplant in which the IVFr was 525%, the subsequent graft survival was 100%. In contrast, IVFr of >25% at 6 months predicted significantly reduced subsequent graft survival (P = .04; Fig. 2).

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IVFr % @em)

20 15

1

PR-OP Time

12

6

post-transplantmonths

Fig 1. Changes in renal transplant interstitial volume fraction (IVFr) with time. ‘P < .05 v. pre-op; tP < .05 v. 1 month.

0 1997 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010

Transplantation

Proceedings,

29, 2793-2794

(1997)

From the University Departments of Surgery (M.L.N., T.J.W.), Nephrology (S.J.H., J.F.), and Pathology, Leicester General Hospital, Leicester, United Kingdom. Reprint requests to Mr. M.L. Nicholson, Department of Surgery, Leicester General Hospital, Leicester LE5 4PW, United Kingdom.

0041-1345/97/$17.00 PII SO041 -1345(97)00680-5

2793

NICHOLSON,

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DISCUSSION

Histomorphometric measurement of IVFr can be used as a predictor of subsequent graft survival in allografts biopsied at the 6 months posttransplant. IVFr may therefore prove to be a useful surrogate endpoint in studies of therapeutic modalities that may prevent chronic allograft damage. The maximum increase in IVFr appears to be reached at 6 months posttransplant, a finding in agreement with the work of Ruiz et al.’ The method of assessing IVFr described here has a number of limitations. In particular, the point counting involved is time consuming and labor inten-

HARPER, WHEATLEY, ET AL

sive. If the method is to be used more widely, it will need to be modified to include a staining procedure to demonstrate the interstitium and the incorporation of an image analysis system to provide rapid automated results.

REFERENCES 1. Tilney NL, Whitley WD, Diamond JR, et al: Transplantation 52:389, 1991

2. Ruiz P, Kolbeck PC, Scroggs MW, et al: Transplantation 45:91, 1988