A double-blind, randomised, controlled trial to study the effects of an enteral feed supplemented with glutamine, arginine, and omega-3 fatty acid in predicted acute severe pancreatitis.

CONTEXT: Current best evidence is in favour of early institution of enteral feeding in acute severe pancreatitis with promising results from trials in immunonutrition on other patient groups. OBJECTIVE: To identify which groups of patients and products are associated with benefit, we investigated immunonutrition in patients with predicted acute severe pancreatitis. DESIGN: A randomised trial of a study feed containing glutamine, arginine, tributyrin and antioxidants versus an isocaloric isonitrogenous control feed was undertaken. PATIENTS: Thirty-one patients with a diagnosis of acute pancreatitis predicted to develop severe disease: 15 study feeds and 16 control feeds. INTERVENTIONS: Enteral feeding via nasojejunal tube for 3 days. If patients required further feeding the study was continued up to 15 days. MAIN OUTCOME MEASURES: Reduction in C-reactive protein (CRP) by 40 mg/L after 3 days of enteral feeding was the primary endpoint. Carboxypeptidase B activation peptide (CAPAP) levels were taken at regular intervals. RESULTS: After 3 days of feeding, in the study group 2/15 (13%) of patients had reduced their CRP by 40 mg/L or more. In the control group 6/16 (38%) of patients had reduced their CRP by this amount. This difference was found to be near the statistical significant limit (P=0.220). CONCLUSIONS: The cause of the unexpectedly higher CRP values in the study group is unclear. The rise in CRP was without a commensurate rise in CAPAP or outcome measures so there was no evidence that this represented pancreatic necrosis. The contrast between the CRP and CAPAP results is of interest and we believe that specific pancreatic indices such as CAPAP should be considered in larger future studies.

Physiologic impact of low-dose dopamine on renal function in the early post renal transplant period.

BACKGROUND: Low-dose dopamine (LDD) (< or =5.0 microg/kg/min) is often used in the early postrenal transplant period for its perceived improvements in renal function parameters. However, there is little published evidence to support its use. The aim of this study was to evaluate the effects of LDD on the physiologic parameters of the transplanted kidney. METHODS: With local ethics approval, 20 consecutive adult patients (age range, 27-74 years), who underwent cadaveric renal transplantation with cyclosporine immunosuppression, were randomized into two study groups, each with 10 patients. The study period was over 9 hrs on the first postoperative day. This 9-hr block was divided into three 3-hr periods. Patient group 1 received a dopamine infusion over the second 3-hr period only, and patient group 2 received a dopamine infusion over both the first and third 3-hr periods. During these periods, urine flow rate (UFR), effective renal plasma flow (ERPF), creatinine clearance (CC), and total urinary sodium excretion rate (tUNa) were measured. RESULTS: In both groups, there were significant (P<0.05, Wilcoxon rank sum test) increases in ERPF, UFR, CC, and tUNa during LDD infusion periods compared with periods of no LDD infusion. No changes in heart rate or mean arterial blood pressure were seen with LDD administration. CONCLUSIONS: LDD significantly increases ERPF, UFR, CC, and tUNa in the transplanted allograft kidney treated with cyclosporine immunosuppression in the early posttransplant period.

Quantitative detection of changes in cytokine gene expression in peripheral blood mononuclear cells correlates with and precedes acute rejection in renal transplant recipients.

Immunological monitoring of transplant recipients is an attractive concept. Cytokines provide an obvious focus for research, as they are central to the human immune response. This study aimed to identify cytokines whose sequential gene expression differentiated rejectors from non-rejectors immediately following renal transplantation. Forty-five renal transplant recipients (15 rejectors) and 13 living donors were recruited. Total RNA was extracted from the peripheral blood mononuclear cells and reverse transcribed. Cytokine gene expression levels of IL-4, IL-10, TNF-alpha and TGF-beta1 were measured using TaqMan. IL-10 expression increased significantly following donor surgery. IL-4 and TNF-alpha patterns clearly differentiated between rejectors and non-rejectors. In the rejectors significant increases occurred more than 48 h before clinical graft dysfunction. Negative predictive values were 76% and 80% for IL-4 and TNF-alpha, respectively. This study has identified two cytokines (IL-4 and TNF-alpha) whose gene expression patterns differentiate between rejecting and non-rejecting renal transplant recipients making immunological monitoring possible.

House keeping genes and gene expression analysis in transplant recipients: a note of caution.

BACKGROUND: House keeping genes are often used as a means of standardising results obtained in gene expression investigations. This study was performed to investigate whether beta-actin, beta2-microglobulin (two genes frequently quoted as house keeping genes) and/or transferrin receptor would be suitable house keeping genes for use in gene expression analysis of renal transplant recipients. METHODS: Sequential expression of all three genes was measured in the peripheral blood mononuclear cells of 13 living donors and 45 renal transplant recipients, pre-operatively and then daily for up to 2 weeks. Fifteen of the recipients experienced an episode of biopsy proven acute rejection. Gene expression measurement was performed using quantitative real time 'TaqMan' PCR technology. RESULTS: Gene expression of all three genes was unchanged in the living donor cohort. However, in the transplant recipients there were significant increases in expression following transplantation in the non-rejectors, and preceding the diagnosis of acute rejection. In the latter group, levels returned to pre-transplant values after the commencement of anti-rejection therapy. CONCLUSIONS: Beta-actin, beta2-microglobulin and transferrin receptor gene expression, although not influenced by surgery, is influenced by transplantation, acute rejection and anti-rejection therapy making these genes unsuitable as house keeping genes following renal transplantation. These findings may cast doubt on the results of some studies that used these genes for the purposes of standardisation when looking at cDNA measurement. We suggest that any group wishing to use a house keeping gene ensure that its expression is independent of study parameters prior to the start of the study.

Comparative evaluation of 'TaqMan' RT-PCR and RT-PCR ELISA for immunological monitoring of renal transplant recipients.

By sequentially monitoring cytokine gene expression (using RT-PCR ELISA technology) in peripheral blood cells of renal transplant recipients in the early post-operatively period we have shown that expression patterns correlate with clinical events, namely acute allograft rejection. This strategy may have the potential of predicting acute rejection prior to clinical detection. Unfortunately, the technique used was time consuming and only semi-quantitative and, therefore, not suitable for clinical application. In this study, we have sought to confirm the results of the early work using a real time quantitative RT-PCR technique ('TaqMan'), which may be applicable in the clinical laboratory. 'TaqMan' primers and probes were designed for Interleukin (IL)-4 and IL-10 using Primer Express software. Cytokine gene expression for both cytokines was re-measured in stored cDNA samples from 27 non-rejectors and 14 patients experiencing an episode of biopsy proven acute rejection. Compared to pre-transplant levels, IL-4 gene expression fell significantly on post-operative days 2 and 7 before returning to baseline values by day 14 in the non-rejectors. In the rejectors, the initial significant fall was again seen, but with an earlier return to pre-transplant levels at the time of rejection diagnosis. This was followed by a further significant fall in levels 48 h after the initiation of anti-rejection therapy. These different patterns for rejectors and non-rejectors were seen using both techniques. For IL-10, gene expression increased significantly following transplantation throughout the study period when compared to baseline values. This pattern was seen using both techniques. In the rejectors, there were different patterns seen depending on the technique used. When using RT-PCR ELISA, the initial rise was again seen followed by a return to baseline values at the time of rejection diagnosis followed by a further significant rise in gene expression after the start of anti-rejection treatment. The pattern resembled those of the non-rejectors when expression was measured using 'TaqMan'. This study has confirmed that sequential monitoring of cytokine gene expression, measured in peripheral blood mononuclear cells, detects significant changes that correlate with clinical events in renal transplant recipients, including acute rejection, although not all changes detected with RT-PCR ELISA were confirmed. Therefore, real time quantitative RT-PCR technology may be useful in monitoring the immunological status of these patients in the early post-operative period.