Clinical Diagnosis Of Rejection Research Articles

Predicting Corneal Graft Rejection by Confocal Microscopy.

This study aims at estimating corneal healing by activated keratocyte (AK) counting with in vivo confocal microscopy after perforating keratoplasty (PK). It assesses the value of AK counting in predicting corneal graft rejection. This prospective single-center observational study included 45 patients who benefited from PK in 2013 and were followed up over 2 years. All were monitored by confocal microscopy at day 1, day 7, and monthly thereafter. The AKs were counted in 5 optical sections in each of the anterior, middle, and posterior stroma. The ability of AKs in predicting the occurrence of corneal rejection was assessed by comparison of AK counts between patients with and without clinical signs of rejection. In patients with graft rejection, the AK counts increased significantly 2 months before the clinical diagnosis of rejection, whereas it remained stable after 4 months in patients without rejection. In patient with graft rejection, the AK count reached a maximum at the rejection diagnosis and antirejection treatment initiation but decreased significantly 1 month after treatment initiation. This study confirmed the predictive value of AK counting in corneal graft rejection. The increase in the AK count allowed predicting graft rejection 2 months before the clinical diagnosis of rejection; it may then be the first sign of subclinical rejection.

Experimental study on the effects of rapamycin in prevention of rat corneal allograft rejection

To investigate the effects of rapamycin on prevention of corneal allograft rejection in murine corneal transplantation. The outbred strain SD and Wistar rats were used as donors and recipients, respectively. Sixty-eight Wistar rats were divided into four groups: Group A, autograft control; Group B, allograft control (the control groups were given placebo only); Group C and D, allograft groups, were treated with orally RAPA (3 mg.kg(-1).d(-1)) and CsA (10 mg.kg(-1).d(-1)), respectively. The drugs were delivered for 12 days beginning at the day of transplantation. Each animal was examined by operating microscopy. The grafts were evaluated clinically by means of Holland's scoring system and graft survival was assessed by Kaplan-Meier analysis. The neovascular indexes of rejection were compared among different groups. Histological examination on ocular tissues was performed on day 14 to confirm the clinical diagnosis of rejection. The average transplant survival time in the allogenic control (Group B) was (11.0 +/- 1.5) d. Treatment with RAPA (Group C) led to a statistically significant prolongation of transplant survival to (36.1 +/- 14.9) d (P < 0.05). Treatment with RAPA 3 mg.kg(-1).d(-1) prolonged transplant survival as compared with treatment with CsA (Group D), but the difference was not statistically significant (P > 0.05). Corneal neovascularization was induced after the surgery. In RAPA group, corneal neovascularization was markedly reduced as compared with allograft control (Group B) (P < 0.05) and CsA group (Group D) (P < 0.05). Fewer inflammatory cells were found in the corneal stroma of the RAPA group. These results show that oral immunosuppression with RAPA can prevent corneal graft rejection and corneal neovascularization.

Diagnostic value of urinary enzyme determination in renal transplantation

Abstract The measurement of enzyme activity in urine provides a sensitive assessment for renal tubular cell damage. The present study was undertaken to evaluate the clinical value of the determination of tubular brush-border-associated enzymes, alkaline phosphatase (AP), gamma-glutamyl transferase (GGT), leucine aminopeptidase (LAP), and dipeptidyl peptidase IV (DPP), of patients with normal graft function (NOR, n= 20), with acute tubular necrosis (ATN, n= ll), with an acute rejection episode (ARE, n= 17) after transplantation, and of healthy persons (n=20). The second urine of the morning was collected daily during the patients' stay in hospital. The enzyme activities were measured at 25 °C and were expressed as U/mmol creatinine. The enzymuria in NOR is higher than in healthy controls, but is still in the normal range. By 5 days after transplantation the initial increased excretion declines as the graft function improves. Elevated enzymuria (DPP 0.69 ± 0.56, AP 3.06 ± 3.24, GGT 4.16 ± 4.13, and LAP 1.39 ± 1.27) was observed during the rejection episodes. Two days before clinical diagnosis of rejection, the release of DPP-IV and GGT increases to double, and the AP and LAP increases to 3 times the value on the fourth day before rejection. Successful treatment of rejection coincided with a quick return by the third day of the rejection period to the previous enzyme distribution. In ATN no decrease of enzymuria occurs and the excretion is much higher than in ARE. Our method with the every day monitoring of kidney graft function offers the possibility for the early diagnosis of acute rejection.

The anaphylatoxin C5a, a new parameter in the diagnosis of renal allograft rejection

Abstract In the underlying study the diagnostic value of the anaphylatoxin CSa was evaluated in kidney transplantation. In 49 transplant patients the following parameters were measured daily for a mean period of 25.1 days: plasma a [P-CSa], urine a [U-CSa], serum amyloid A (SAA], serum neopterin [S-NEOP] and urine neopterin [U-NEOP]. Sensitivity, specificity and day of first significant parameter increase (exceeding a cut-off level of > 50 YO) were evaluated retrospectively during 30 periods of rejection and 30 periods of stable graft function. U- a was the parameter with the highest sensitivity (84 %) and specificity (84 %), increasing in the mean 1.3 days before clinical diagnosis of rejection. Sensitivity and specificity of the other markers was lower: SAA 77 % and 77 %, U-NEOP 68 % and 65 %, S-NEOP 45 % and 77 %, and P- a 45 % and 48 %, respectively. During four instances of cy-tomegalovirus disease extremely high U-NEOP (≥= 1520 ± 518 μmol/mol creatinine) and slightly increased P- a levels (≥= 1.5 ± 1.4 ng/ml) occurred. Elevated urinary excretion of a seems to be a reliable and early marker of renal allograft rejection. In combination with SAA and U-NEOP, the daily assessment of U- a differentiates between viral infection and allograft rejection.

Diagnostic value of urinary enzyme determination in renal transplantation.

The measurement of enzyme activity in urine provides a sensitive assessment for renal tubular cell damage. The present study was undertaken to evaluate the clinical value of the determination of tubular brush-border-associated enzymes, alkaline phosphatase (AP), gamma-glutamyl transferase (GGT), leucine aminopeptidase (LAP), and dipeptidyl peptidase IV (DPP), of patients with normal graft function (NOR, n = 20), with acute tubular necrosis (ATN, n = 11), with an acute rejection episode (ARE, n = 17) after transplantation, and of healthy persons (n = 20). The second urine of the morning was collected daily during the patients' stay in hospital. The enzyme activities were measured at 25 degrees C and were expressed as U/mmol creatinine. The enzymuria in NOR is higher than in healthy controls, but is still in the normal range. By 5 days after transplantation the initial increased excretion declines as the graft function improves. Elevated enzymuria (DPP 0.69 +/- 0.56, AP 3.06 +/- 3.24, GGT 4.16 +/- 4.13, and LAP 1.39 +/- 1.27) was observed during the rejection episodes. Two days before clinical diagnosis of rejection, the release of DPP-IV and GGT increases to double, and the AP and LAP increases to 3 times the value on the fourth day before rejection. Successful treatment of rejection coincided with a quick return by the third day of the rejection period to the previous enzyme distribution. In ATN no decrease of enzymuria occurs and the excretion is much higher than in ARE. Our method with the every day monitoring of kidney graft function offers the possibility for the early diagnosis of acute rejection.

The anaphylatoxin C5a, a new parameter in the diagnosis of renal allograft rejection.

In the underlying study the diagnostic value of the anaphylatoxin C5a was evaluated in kidney transplantation. In 49 transplant patients the following parameters were measured daily for a mean period of 25.1 days: plasma C5a [P-C5a], urine C5a [U-C5a], serum amyloid A [SAA], serum neopterin [S-NEOP] and urine neopterin [U-NEOP]. Sensitivity, specificity and day of first significant parameter increase (exceeding a cut-off level of > 50%) were evaluated retrospectively during 30 periods of rejection and 30 periods of stable graft function. U-C5a was the parameter with the highest sensitivity (84%) and specificity (84%), increasing in the mean 1.3 days before clinical diagnosis of rejection. Sensitivity and specificity of the other markers was lower: SAA 77% and 77%, U-NEOP 68% and 65%, S-NEOP 45% and 77%, and P-C5a 45% and 48%, respectively. During four instances of cytomegalovirus disease extremely high U-NEOP (> or = 1520 +/- 518 mumol/mol creatinine) and slightly increased P-C5a levels (> or = 1.5 +/- 1.4 ng/ml) occurred. Elevated urinary excretion of C5a seems to be a reliable and early marker of renal allograft rejection. In combination with SAA and U-NEOP, the daily assessment of U-C5a differentiates between viral infection and allograft rejection.

Increased serum neutral endopeptidase activity in acute renal allograft rejection.

Neutral endopeptidase (EC 3.4.24.11; NEP), originally isolated from renal tubular brush border, is a cell surface peptidase identical to the CD10 antigen (or CALLA; common acute lymphoblastic leukemia antigen) in lymphoid cells. We studied the serum NEP levels daily after transplantation (Tx) in 19 renal allograft recipients. The NEP activity was determined with a two-step enzymatic assay utilizing a fluorogenic substrate (Suc-Ala-Ala-Phe-AMC; see text) and related to clinical signs of graft rejection, to signs of immunoactivation in transplant fine-needle aspiration biopsy (FNAB) specimens, to renal function, and to serum levels of C-reactive protein. The serum NEP levels remained normal (peak level 10.3 +/- 1.8 micrograms/l on days 6-9 after Tx, initial level after Tx 7.3 +/- 1.4 micrograms/1 on day 2; mean values +/- SEM) in patients who neither showed clinical signs of rejection nor had findings of immunoactivation in FNAB samples. On the contrary, the serum NEP levels rose clearly in patients developing acute rejection verified clinically and in FNAB samples (peak value 90.4 +/- 18.7 micrograms/l on days 6-9 post-Tx; p < 0.001 compared with patients without sings of immunoactivation) and even in patients having immunoactivation in FNAB without clinical evidence of rejection (108.2 +/- 22.4 micrograms/l, p < 0.001). Serum NEP peak appeared 2-3 days before clinical diagnosis of rejection and a positive findings in FNAB samples. Serum NEP increments did not correlate with changes in serum creatinine, delayed onset of renal excretory function, blood leukocyte count, C-reactive protein level, or infections. Thus, the serum NEP activity was shown to increase after renal allotransplantation associated with early phases of immunoactivation and development of acute graft rejection. Because of the limited number of patients studied, the clinical implications of these preliminary observations for kidney transplant monitoring clearly need confirmation in larger studies.

Characterization of donor-directed antibody class in the post-transplant period using flow cytometry in renal transplantation

Over the past few years there has been increasing awareness of the importance of humoral mechanisms in the rejection of renal transplants. In this study we have monitored the development of antibodies directed against donor T and B lymphocytes using the sensitive flow cytometric technique. Forty-two cadaveric renal transplants were studied both before and for a maximum of 14 days after transplantation. Donor cells were separated from spleen on the day of transplantation and stored in liquid nitrogen until required. The dual colour flow cytometric assay was used to detect IgG or IgM directed against donor T or B lymphocytes. Using AB sera as controls, results were expressed as relative median fluorescence (RMF) and then correlated with the clinical performance of the grafts. Significant associations were found between the incidence of donor-directed antibodies and the development of clinical rejection. The magnitude of the rise in antibody levels was also related to graft performance. In patients showing severe graft rejection, high levels of antibodies of the IgG class developed before the clinical diagnosis of rejection was made. The routine use of this test allows the prediction of impending severe rejection to be made and may have important implications for immunosuppressive therapy.

Evidence that the systematic analysis of bile cytology permits monitoring of hepatic allograft rejection.

Cytologic analysis was performed on 128 bile specimens collected by schedule from 12 liver transplant recipients over a 4-month period. Clinical diagnoses at the time of specimen collection were determined retrospectively, as follows: clinically stable, 75; acute rejection, 15; CMV hepatitis, 1; systemic infection, 8; ischemic injury, 24 (all within the first 4 days postop); nonclassifiable, 5. Bile analysis was done by a blinded investigator. Specimens contained ductal epithelial cells (EC) and inflammatory cells (IC), which were counted using Cytospin slide preparations. Greater than 10 cells/slide were seen in 93.3% of rejections, 91.7% of ischemic injuries, 100% of systemic infections, and 14.6% of stable patients. In samples collected after POD 4, IC were seen in 86.7% of rejections, yielding a specificity of 94.4% (P less than 0.001). If lymphoblastic cells were also seen, the specificity increased to 96.6%. Five specimens were obtained the day before the clinical diagnosis of rejection; all demonstrated IC. Seven specimens were obtained 3 days after beginning therapy for rejection. In 5 the bile contained no IC, and clinical improvement occurred; in the 2 in whom IC were found, further therapy was subsequently required. IC were seen in 5 of 8 specimens taken when systemic infection was present; the clinical setting allowed differentiation from rejection. Only 1 case of CMV hepatitis was included, thus no conclusions can be drawn for this entity. Cytoplasmic vacuolization of EC was observed in 30% of cases, in these, cyclosporine levels were significantly higher (989.9 +/- 356.9 vs. 672.8 +/- 421.2, P = 0.02). In summary, bile cytology analysis aides in the monitoring of the onset and duration of rejection. It may be an indicator of persistent rejection, and it may help prevent overimmunosuppression in those cases with normal cytological findings.

EVIDENCE THAT THE SOLUBLE INTERLEUKIN 2 RECEPTOR LEVEL MAY DETERMINE THE OPTIMAL TIME FOR CYSTOSCOPICALLY-DIRECTED BIOPSY IN PANCREATICODUODENAL ALLOGRAFT RECIPIENTS

In 18 consecutive pancreaticoduodenal allograft recipients (15 combined kidney/pancreas and 3 pancreas only after a prior successful kidney transplantation) operated on between December 1987 and February 1989, we studied the soluble interleukin 2 receptor (SIL-2R) level over time. All pancreaticoduodenal allografts were transplanted with exocrine drainage via a duodenocystostomy that allowed for cystoscopically directed needle biopsies of the pancreas. Of these 18 recipients, at 6 weeks after transplantation, 6 had had no rejection episodes or cytomegalovirus disease (control group), an acute allograft rejection had developed in 7, CMV disease developed in 4, and both rejection and CMV disease developed in 1 by 12 days after transplantation. SIL-2R level increased in all patients during immunosuppressive induction therapy (preoperative mean +/- SE, 1637 +/- 284 U/mL; maximum, 4367 +/- 687 U/mL). After induction therapy, the mean was 2768 +/- 432 U/mL. In all 6 recipients in the control group, SIL-2R level continued to decrease. However, SIL-2R level was significantly higher compared with controls, in those who had CMV disease (levels were increased at a mean of 7 days before diagnosis of CMV disease) and in those who had acute rejection episodes (levels were increased a mean of 7 days before the clinical diagnosis of rejection). Factors that did not cause an increase in SIL-2R level included acute pancreatitis, wound infection, operative procedures, and CsA nephrotoxicity. SIL-2R level can be useful for monitoring pancreaticoduodenal allograft recipients. Increases predict impending rejection or CMV disease, prior to the onset of organ dysfunction. When SIL-2R level increases, we recommend cultures of blood and urine to exclude CMV and pancreaticoduodenal allograft biopsy to confirm early rejection prior to the initiation of potentially dangerous antirejection therapy.

Binding of antihuman globulin by exfoliated renal tubular cells following kidney transplantation.

Exfoliated renal tubular epithelial cells (RTCs) from kidney allograft recipients may bind antibody against human globular proteins. Urine from sixty consecutive transplant recipients was studied in the first month following transplantation to relate this binding to the clinical course and rejection. The spun, washed sediment was incubated with fluoresceinated goat antihuman globulin and examined under light and fluorescent microscopy for fluoresceinated RTCs. Of 28 patients who were never positive, 27 manifested no clinical rejection episodes. Of 22 total rejection episodes, 21 were preceded by the appearance of fluorescent RTCs. Five patients in this group did not revert to negative in this test, and all went on to loss of graft from acute rejection. Of 46 patients who were discharged from the hospital with negative RTCs, only four were readmitted within one month for treatment of rejection. In contrast, of the 11 patients who were positive at the time of discharge, 10 were readmitted in the first month. Graft survival was only 55% (6/11) in this latter group as compared with 91% (42/46) in the former. There were 11 patients with transiently positive tests who did not warrant a clinical diagnosis of rejection. In no case of acute tubular necrosis (ATN) alone or in obstructive uropathy was the assay positive. However, in some cases, in which the ATN merged imperceptibly into rejection, the RTCs started to fluoresce well in advance of the clinical suspicion of rejection. Information obtained from this examination may be used to assess the cause of renal failure in the early posttransplant period and to differentiate rejection from ATN and obstruction. This phenomenon of fluorescent RTCs may be an early manifestation of an immunological change occurring in a cell that is targeted by the host for rejection.

URINE i-TXB 2 IN RENAL ALLOGRAFT REJECTION

Immunoreactive thromboxane B 2 (i-TXB 2) was measured in daily urine samples from twelve patients after renal transplantation. In 21 of 30 rejection episodes, the increase in i-TXB 2 preceded both the increase in serum β 2-microglobulin (β 2-MG) and the clinical diagnosis of rejection. In 26 of 30 rejection episodes, the increase in urine i-TXB 2 preceded the increase in serum creatinine. The degree of change in i-TXB 2 is greater than that of either serum β 2-MG or creatinine. Urinary i-TXB 2 was very high in one patient with deep venous thrombosis, but it did not rise in patients with urinary tract infection, pneumonia, or acute tubular necrosis. Thus, urinary i-TXB 2 seems to be an early indicator of clinical renal allograft rejection.

Retrospective correlation of clinical and histologic findings of 189 exchanged kidneys

Tissues samples from 189 unsuccessful renal allografts, 47 recovered at autopsy and the others removed surgically, were examined histologically by light microscopy. Tissues samples were obtained from cadaver kidneys that had been exchanged regionally for transplantation. Each allograft tissue sample was rated as to extent of pathologic changes denoting rejection and was classified accordingly. Surgical and autopsy reports, as well as clinical data, were then obtained and these were compared with the retrospective pathologic findings of this study. Our pathologic findings agreed with the original pathologic diagnosis as to presence or absence of rejection changes in 180 cases, but disagreed with the clinical diagnosis of rejection in 28 of the 63 cases with minimal or no histologic evidence of rejection. There was less disagreement with the clinical diagnosis for the 87 cases with histologic evidence of rejection which had been judged as sufficient to cause allograft loss, 70 having been clinically diagnosed as rejected. Disagreement occurred most often where the allograft had never functioned or had been lost within 3 months. Retrospective analysis did not disclose any association between rejection histology and preformed antibodies or length of kidney perfusion time. Sufficient allografts appeared to have been lost for reasons other than rejection to cast doubt on the validity of interpreting renal allograft data only by graft survival statistics.

Urinary Cytodiagnosis of Acute Renal Allograft Rejection Using the Cytocentrifuge

The clinical diagnosis of acute renal allograft rejection in immunosuppressed recipients can often be predicted or confirmed on the basis of characteristic urinary cytologic findings. Use of cytocentrifugation permits a simple, rapid, reproducible and semiquantitative means of preparing cytologic urinary specimens of diagnostic quality from small quantities of urine. The cytodiagnosis of acute renal rejection was established before or on the same day a clinical diagnosis of rejection was made in the majority of renal transplant cases studied over a 12-month period. Renal tubular cells were found to be the exfoliated cells of greatest value in predicting an acute rejection episode, and their persistence has prognostic importance.

Ultrasonic measurement of blood flow changes in canine renal allografts

Renal allografts were carried out in dogs, the kidney being placed in the neck. After operation, ultrasonic and electromagnetic renal blood flow studies were carried out on alternate days. Postoperatively, an increase in blood flow was seen for the first 2–3 days, and thereafter a progressive fall in blood flow was observed until the animal died of rejection. A high degree of correlation was observed between the two methods of blood flow measurement. From these observations, it is concluded that ultrasonic methods of monitoring blood flow changes in rejection of human renal allografts is a useful adjunct in the clinical diagnosis of rejection.