The potential clinical utility of serial plasma albumin mRNA monitoring for the post-liver transplantation management.

OBJECTIVES: Elevated albumin (ALB) mRNA concentration has been reported in the plasma of patients with liver diseases. The plasma ALB mRNA measurement was shown to be an effective indicator of liver pathologies with superior diagnostic sensitivity and specificity when compared with alanine transaminase (ALT). We hypothesized that serial plasma ALB mRNA analysis would be helpful in the early detection and monitoring of post-liver transplantation complications. DESIGN AND METHODS: One hundred and five blood specimens were collected from 24 post-transplant recipients. Biochemical liver function test profiles and plasma ALB mRNA concentrations were assessed. RESULTS: Over the study period, the health status of 14 recipients (58%) remained stable (Stable group). Their plasma ALB mRNA concentrations remained within a low-concentration range. In contrast, 10 recipients (42%) developed 14 episodes of hepatic complications (Unstable group). The median plasma ALB mRNA concentration of the Unstable group was 6.5-times higher than that of the Stable group. Plasma ALB mRNA concentration was elevated on 13/14 (93%) episodes of the hepatic complications while ALT was elevated only on 8/14 (57%) episodes. CONCLUSIONS: The elevation of plasma ALB mRNA may allow sensitive detection of hepatic complications and monitoring of the clinical course in a dynamic fashion. Serial plasma ALB mRNA measurement is potentially useful for post-liver transplantation management.

Multiple atherosclerosis-related biomarkers associated with short- and long-term mortality after stroke.

OBJECTIVES: We investigated the relationships of biomarkers of various pathophysiologic pathways including high-sensitivity C-reactive protein (hs-CRP), lipocalin-2 (LCN2), myeloperoxidase (MPO) and matrix metalloproteinases 9 (MMP9) with mortality in stroke patients. DESIGN AND METHODS: hs-CRP, LCN2 and MPO concentrations in 92 patients were determined using enzyme-linked immunosorbent assays. MMP9 mRNA concentrations were determined using real-time quantitative reverse transcription-polymerase chain reaction. RESULTS: Twelve patients (13.0%) died at 6 months and 34 patients (37.0%) died at 5 years. The independent predictors for 6-month mortality were hs-CRP (adjusted OR=16.0) and LCN2 (adjusted OR=16.9), while for 5-year mortality was hs-CRP (adjusted OR=5.56). For patients with hs-CRP >3.4 mg/L, an increase in LCN2 was associated with 2.5-fold higher 6-month mortality, while an increase in normalized MMP9 mRNA was associated with 5.8-fold higher 6-month and 1.5-fold higher 5-year mortality. CONCLUSION: hs-CRP was the most significant independent predictor of both short- and long-term mortality after stroke, with LCN2 and MMP9 mRNA each adding further to the risk stratification.

Intrarenal cytokine gene expression in lupus nephritis.

BACKGROUND: Lupus nephritis is characterised by intrarenal inflammation and lymphocyte activation. AIM: To examine the profile of cytokine gene expression in glomerulus and tubulointerstitium in patients with lupus nephritis. METHODS: 36 consecutive patients with systemic lupus erythematosus having active renal disease were recruited, and they were required to undergo kidney biopsy. Glomerular and tubulointestitial cytokine expression of interleukin (IL)2, 4, 10, 12, 18, interferon gamma (IFN)gamma, T-bet (the Th1 transcription factor), GATA-3 (the Th2 transcription factor), transforming growth factor beta and monocyte chemoattractant protein (MCP)1 were studied by laser microdissection of the renal biopsy specimen, followed by real-time quantitative PCR. RESULTS: There were 13 patients with World Health Organization class III nephritis, 14 patients with class IV nephritis and 9 patients with class V nephritis. There was a significant correlation between serum C3, C4 and anti-double strand DNA antibody level with glomerular expression of T-bet, IFNgamma and IL2. There was a significant correlation between histological activity index and glomerular expression of IL12, IL18, IL10 and MCP1. In addition, the degree of glomerular leucocyte infiltration significantly correlated with glomerular expression of IFNgamma, IL10, IL12 and IL18. By contrast, histological chronicity index correlated with the tubulointerstitial expression of IL2, MCP1 and GATA-3. CONCLUSIONS: Intraglomerular expression of certain target genes correlate with the severity of systemic as well as histological activity, whereas the tubulointerstitial expression of other target genes correlate with the degree of chronic kidney scarring. This result may shed light on the immunopathogenesis of lupus nephritis.

Messenger RNA expression of RANTES in the urinary sediment of patients with lupus nephritis.

BACKGROUND: Lupus nephritis is characterized by intra-renal inflammation. Patients with systemic lupus erythematosus (SLE) showed abnormal T-cell expression of RANTES (regulated upon activation, normal T cell expressed) and its level in their serum. The authors studied the mRNA expression of RANTES in the urinary sediment of lupus patients. METHODS: The authors studied 88 lupus patients, who were classified into active, remission and non-renal SLE groups according to the disease activity, 29 non-SLE and 10 healthy controls. Lupus activity was assessed by the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). Urinary mRNA expression of RANTES was studied by real-time quantitative polymerase chain reaction. RESULTS: The expression of RANTES in urinary sediment was significantly elevated in active group (P < 0.001). Expression level of RANTES correlated with the SLEDAI score (r = 0.57; P < 0.001) and renal score in SLEDAI (r = 0.60; P < 0.001). In addition, urinary expression of RANTES had significant correlation with degree of proteinuria, serum creatinine, albumin and estimated glomerular filtration rate. CONCLUSION: The authors conclude that the mRNA expression of RANTES was elevated in the urinary sediment of patients with active lupus nephritis. Measurement of urinary mRNA expression may be a novel non-invasive method for the assessment of lupus disease activity and the severity of renal involvement.

mRNA expression of target genes in the urinary sediment as a noninvasive prognostic indicator of CKD.

BACKGROUND: Study of messenger RNA (mRNA) expression of target genes in urinary sediment was suggested as a noninvasive marker of renal damage in patients with chronic kidney diseases (CKDs). We studied the relationship between urinary mRNA expression of target genes and risk for renal function deterioration in patients with CKD. METHODS: We studied 131 patients with CKD with kidney biopsy. mRNA expression of 11 target genes in urinary sediment was measured by means of quantitative polymerase chain reaction. Patients then were followed up for 27.4 +/- 10.1 months. The primary end point is doubling of serum creatinine concentration or end-stage renal disease. RESULTS: Thirty-six patients (27.5%) reached the primary end point during follow-up. Univariate analysis showed that sex, age, proteinuria, estimated glomerular filtration rate, histological diagnosis, degree of tubulointerstitial scarring, percentage of glomerulosclerosis, and urinary mRNA expression of hepatocyte growth factor (HGF) were predictors of the primary end point. At 24 months, event-free survival rates were 90.9% and 64.3% for patients with low and high urinary HGF expression, respectively (log rank test, P = 0.002). After adjusting for other confounding factors by using a Cox proportional hazard model, urinary HGF expression remained an independent predictor of the primary end point, and a 1-fold increase in expression was associated with a 4.0% (95% confidence interval, 0.5 to 7.5; P = 0.024) increase in risk. CONCLUSION: In the target genes examined, urinary HGF expression is an independent prognostic indicator of CKD after adjusting for confounding clinical and histological factors. Measurement of urinary HGF mRNA expression may be a useful noninvasive tool for risk stratification of patients with CKD.

The effect of immunosuppressive therapy on the messenger RNA expression of target genes in the urinary sediment of patients with active lupus nephritis.

BACKGROUND: Previous studies have shown that messenger RNA (mRNA) expression of target genes is increased in the urinary sediment of patients with active lupus. We study the effect of immunosuppressive therapy on the urinary gene expression profile in patients with active lupus nephritis. Method. We recruited nine patients with active systemic lupus erythematosus (SLE) and renal disease, and required corticosteroid, with or without cytotoxic treatment. They were followed for 6 months, urine samples were collected at 0, 4, 12 and 24 weeks and gene expression profile was determined by polymerase chain reactions. The pattern of gene expression was compared to clinical parameters of therapeutic response. RESULTS: Amongst the target genes studied, there was a progressive decline in the urinary expression of T-bet, interleukin (IL)-10, transforming growth factor-beta (TGF-beta), monocyte chemoattractant protein-1 (MCP-1), and interferon-gamma (IFN-gamma) after immunosuppressive treatment, although the change of IFN-gamma was not statistically significant. The time course of their urinary expression was parallel to the systemic activity as reflected by the systemic lupus erythematosus disease activity index (SLEDAI). Throughout the study period, the SLEDAI score correlated significantly with the expressions of IFN-gamma (r = 0.43, P = 0.009), T-bet (r = 0.40, P = 0.016), TGF-beta (r = 0.51, P = 0.002) and MCP-1 (r = 0.38, P = 0.022). The anti-double strand(anti-ds)DNA antibody titer correlated significantly with the expressions of IFN-gamma (r = 0.45, P = 0.009), T-bet (r = 0.37, P = 0.034), IL-10 (r = 0.59, P<0.001), TGF-beta (r = 0.44, P = 0.010) and MCP-1 (r = 0.49, P = 0.004). On the other hand, the expression level of IL-2, IL-4, IL-12, IL-18 and GATA-3 remained static throughout the study period. CONCLUSIONS: The mRNA expression of T-bet, IL-10, TGF-beta, MCP-1, and probably IFN-gamma in the urinary sediment of patients with active lupus nephritis improves with successful immunosuppressive therapy, and the change in gene expression profile is in phase with the clinical disease activity. Measurement of urinary mRNA expression of target genes may be a potential non-invasive tool for the monitoring of lupus disease activity.

Can the urine dipstick test reduce the need for microscopy for assessment of systemic lupus erythematosus disease activity?

OBJECTIVE: Urine microscopic examination is an important component of the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). We investigated whether the urine dipstick test can reduce the need for microscopy for the assessment of SLEDAI. METHODS: We studied 269 urine samples from 259 SLE patients with Albustix and Hemastix reagent strips. The results were compared to concomitant microscopic examination of urinary sediment. RESULTS: When trace red blood cell was defined as the cutoff, the sensitivity, specificity, and negative predictive value (NPV) of the Hemastix urine test were 0.98, 0.53, and 0.99, respectively, for hematuria; 0.82, 0.47, and 0.90, respectively, for the presence of pyuria; and 0.91, 0.44, and 0.98, respectively, for the presence of casts by microscopic examination. When proteinuria of 1+ was defined as the cutoff, the sensitivity, specificity, and NPV of the Albustix test were 1.00, 0.46, and 0.99, respectively, for urinary casts; and 0.82, 0.49, and 0.90, respectively, for the presence of pyuria. When both Albustix and Hemastix were applied as screening test, urine microscopy could be reduced by 27%; however, 8% of cases with normal Albustix and Hemastix tests had at least one abnormality on urine microscopy examination. CONCLUSION: In patients with SLE, a combination of Albustix and Hemastix urine tests showed reasonable sensitivity to detect abnormalities in urine sediment. Based on these results, routine urine microscopy can be limited to SLE patients with abnormal Albustix or Hemastix tests. Rarer causes of abnormal renal function in lupus, such as tubulointerstitial nephritis or drug induced interstitial nephritis, would be manifested by pyuria and therefore would not necessarily be detected by changes in the blood and protein detectors on the urine dipstick.

Messenger RNA expression of target genes in the urinary sediment of patients with chronic kidney diseases.

BACKGROUND: The degree of renal scarring in kidney biopsy is an important prognostic factor in patients with chronic kidney diseases. We hypothesize that gene expression in the urinary sediment reflects the degree of renal damage. METHODS: We studied 29 patients with chronic kidney disease who underwent kidney biopsy (12 immunoglobulin-A nephropathy and 17 glomerulosclerosis) and 10 healthy controls. The mRNA expressions of a panel of target genes in urinary sediment were measured by real-time quantitative polymerase chain reaction. The results were compared with the degree of histological damage and renal function decline. RESULTS: There were significant differences in the urinary expression of transforming growth factor-beta (TGF-beta), monocyte chemotactic protein-1 (MCP-1) and collagen IV between disease groups and controls. Urinary TGF-beta mRNA expression correlated significantly with estimated glomerular filtration rate (r = -0.412, P = 0.029) and the degree of tubulointerstitial scarring (r = 0.418, P = 0.024). Urinary MCP-1 expression correlated with the degree of glomerulosclerosis (r = 0.450, P = 0.014), but not tubulointerstitial scarring. Urinary MCP-1 expression correlated with its corresponding level by enzyme-linked immunosorbent assay (ELISA) (r = 0.650, P<0.001), but TGF-beta expression did not correlate with its ELISA level. Urinary TGF-beta gene expression correlated with its intra-renal expression in glomeruli (r = 0.701, P<0.001) and tubulointerstitium (r = 0.573, P = 0.001) by immunohistochemistry, while urinary MCP-1 gene expression correlated with its staining in glomeruli (r = 0.576, P = 0.001) but not tubulointerstitium. After 12 months, there was a significant inverse correlation between the rate of renal function decline and urinary expression of connective tissue growth factor (r = -0.471, P = 0.010) and collagen I (r = -0.399, P = 0.032), but not TGF-beta or MCP-1. CONCLUSIONS: Amongst the target genes examined, the mRNA expression of TGF-beta in urinary sediment correlated with renal function, the degree of histological damage and intra-renal level in patients with chronic kidney diseases. Measurement of TGF-beta mRNA expression in urine may be a useful non-invasive tool for assessing the severity of renal damage in patients with chronic kidney diseases.

Expression of chemokine and fibrosing factor messenger RNA in the urinary sediment of patients with lupus nephritis.

OBJECTIVE: Lupus nephritis is characterized by intrarenal inflammation. To assess the extent and severity of disease activity and renal involvement, this study examined the expression of transforming growth factor beta (TGFbeta) and monocyte chemoattractant protein 1 (MCP-1) in the urinary sediment of patients with systemic lupus erythematosus (SLE). METHODS: We studied 106 patients with SLE who were classified according to their disease status as those with active disease, those with disease in remission, and those with nonrenal SLE. Ten healthy subjects were used as controls. Lupus activity was assessed by the SLE Disease Activity Index (SLEDAI). If renal biopsy was performed, the histologic activity index and chronicity index were determined, and a morphometry analysis of renal scarring was performed. The urinary expresssion of TGFbeta and MCP-1 messenger RNA (mRNA) was studied by real-time quantitative polymerase chain reaction, and the corresponding protein concentrations of TGFbeta and MCP-1 in the urine were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Expression of TGFbeta and MCP-1 mRNA in the urinary sediment was significantly elevated in the active disease group (P < 0.001 for both). These expression levels of TGFbeta and MCP-1 mRNA correlated with the SLEDAI score (TGFbeta r = 0.71, P < 0.001; MCP-1 r = 0.72, P < 0.001), and also significantly correlated with the histologic activity index (TGFbeta r = 0.487, P = 0.004; MCP-1 r = 0.357, P = 0.038). The urinary protein concentration of MCP-1, but not of TGFbeta, correlated with the SLEDAI score (r = 0.66, P < 0.001). However, neither the protein concentration of TGFbeta nor that of MCP-1 as measured by ELISA in the urine correlated with the histologic activity index. CONCLUSION: The measurement of urinary mRNA expression may be a noninvasive method for the assessment of lupus disease activity and the severity of renal involvement in patients with lupus nephritis.

Advances in the clinical laboratory assessment of urinary sediment.

Urinalysis has been used extensively in clinical practice to aid in the diagnosis of various renal and urologic diseases. The innovation of urinalysis is marching on right along with the rapid developments in biotechnology and astride from the solo urine cytology to sophisticated studies of individual component in the urinary sediment. In this review article, we focus on the use of flow cytometry and other technical advances in the examination of urinary sediment, the detection of urologic malignancies by the presence of microsatellite alteration in the urinary sediment, as well as the quantification of cytokine messenger RNA (mRNA) expression in urinary sediment by reverse transcription and real-time quantitative polymerase chain reaction (RT-QPCR). Notably, the study of cytokine mRNA expression in urinary sediment by RT-QPCR has recently been reported to provide important diagnostic information in kidney allograft recipients and patients with lupus nephritis. This simple and non-invasive method requires further study to determine its role in risk stratification and monitoring of therapeutic response in patients with other kidney diseases.

Inflammatory cytokine gene expression in the urinary sediment of patients with lupus nephritis.

OBJECTIVE: Lupus nephritis is characterized by intrarenal inflammation and lymphocyte activation. In the present study, the expression of cytokine genes in the urinary sediment of patients with systemic lupus erythematosus (SLE) was examined. METHODS: We studied 3 SLE patient groups (25 with active lupus nephritis [active group], 25 with inactive SLE and previous renal involvement [remission group], 20 with inactive SLE and no history of renal involvement [nonrenal SLE group]) and 2 control groups (10 patients with noninflammatory renal diseases [non-SLE group] and 10 healthy volunteers [healthy group]). Cytokine gene expression in the urinary sediment was studied by real-time quantitative polymerase chain reaction. RESULTS: Expression of interferon-gamma (IFNgamma) in urinary sediment was significantly higher in the active group than in all other groups (P < 0.001 by Kruskal-Wallis test). Among the SLE patient groups, there was a close correlation between IFNgamma expression and the overall SLE Disease Activity Index (SLEDAI) score (Spearman's r = 0.590, P < 0.001) and the SLEDAI renal score (r = 0.642, P < 0.001). Urinary expression of interleukin-2 (IL-2) in the active group was significantly higher than that in the healthy group (P = 0.046) but not in the remission or nonrenal SLE groups. There was no difference in the levels of IL-4 expression among the SLE groups. CONCLUSION: We found a predominance of Th1 cytokine in the urinary sediment of patients with active lupus nephritis. Measurement of cytokine gene expression in urinary sediment may be a useful noninvasive tool for assessing the severity of renal involvement in SLE.