Abnormal glycosylation of Tamm-Horsfall protein in patients with interstitial cystitis.

OBJECTIVE: To confirm abnormal glycosylation of Tamm-Horsfall protein (THP) in patients with interstitial cystitis (IC). PATIENTS, SUBJECTS AND METHODS: The sialic acid content of THP, a critical component of its biological activity, is reduced in patients with IC. N-glycan shows reduced levels of high molecular weight tri- and tetra-antennary sialylated oligosaccharides. These results are supported by quantitative monosaccharide analysis of neutral and amino sugars in patients vs control subjects. THP was isolated from urine samples of 23 patients with IC and 24 control subjects by salt precipitation. The sialic acid contents were measured using 1,2-diamino-4,5-methylene dioxybenzene-high performance liquid chromatography analysis. For N-glycan profiling, purified THP was treated with peptide:N-glycosidase F to release N-glycans. The purified N-glycans were labelled with 2-aminobenzamide and were profiled by high-pH anion exchange chromatography (HPAEC) with fluorescence detection. The neutral and amino sugars were determined by HPAEC with pulsed amperometric detection. RESULTS: The total sialic acid in patients was half of that in controls. There was a pattern of reduced level of high molecular weight sialylated oligosaccharide in 17 of 23 patients vs four of 24 controls. The total neutral and amino sugars showed a approximately 30% reduction in patients. The mean (sem) for the controls was 133.79 (6.51) vs 94.76 (6.67) nmol/200 microg of THP for patients (P < 0.001). CONCLUSIONS: THP in patients with IC has reduced sialylation and overall glycosylation, and by inference, THP has a role in the pathophysiology of IC.

Defective Tamm-Horsfall protein in patients with interstitial cystitis.

PURPOSE: Normal urinary Tamm-Horsfall protein shows a urothelial cytoprotective effect against potentially toxic compounds in urine that may injure the urothelium and cause bladder disease. One such disease is interstitial cystitis. In patients with interstitial cystitis this protective effect is decreased. We hypothesized that a difference in Tamm-Horsfall protein in patients with interstitial cystitis exists that may be involved in disease pathogenesis. MATERIALS AND METHODS: Using enzyme-linked immunosorbent assay the urinary Tamm-Horsfall protein concentration was determined in patients with interstitial cystitis and control subjects. Sialic acid content was measured by high performance liquid chromatography based assay. The structure of the protein glycosylation chains was analyzed using matrix assisted laser desorption/ionization-time of flight mass spectrometry. RESULTS: The mean Tamm-Horsfall protein concentration was not significantly different in patients with interstitial cystitis and controls (28.8 vs 28.2 mg/l urine and 36.8 vs 36.7 microg/mg creatinine, respectively, p = 0.6). The total mean sialic acid content of Tamm-Horsfall protein was almost 2-fold lower in 22 patients with interstitial cystitis compared with that in 20 controls (46.3 +/- 4.3 vs 75.3 +/- 4.1 nmol sialic acid per mg Tamm-Horsfall protein, respectively, p <0.0001). On matrix assisted laser desorption/ionization-time of flight mass spectrometry N-glycans released from Tamm-Horsfall protein revealed lower molecular weight di-antennary N-glycan structures and a resulting decrease in the number of terminal sialic acid residues in 10 patients with interstitial cystitis relative to those in 10 controls. CONCLUSIONS: Tamm-Horsfall protein is qualitatively different in patients with interstitial cystitis compared to controls. These data suggest that altered Tamm-Horsfall protein may be involved in interstitial cystitis pathogenesis and it may be useful for clinical diagnosis.

Prevalence of nephrolithiasis in human immunodeficiency virus infected patients on the highly active antiretroviral therapy.

BACKGROUND AND PURPOSE: Protease inhibitors, specifically indinavir, have historically been implicated as a cause of nephrolithiasis in the human immunodeficiency virus (HIV) infected patients. There is a paucity of data, however, on stone disease with nonindinavir etiologies since the introduction of highly active antiretroviral therapy (HAART). We sought to describe the prevalence of nephrolithiasis in the HIV population since the use of HAART. PATIENTS AND METHODS: We retrospectively reviewed HIV-positive patients currently receiving HAART treatment in whom image proven kidney and/or ureteral urolithiasis developed, between 1998 and 2010. A detailed analysis of patients' current treatment, surgical intervention, and metabolic studies was performed. RESULTS: A total of 436 HIV-positive patients were included and 46 (11%) patients had nephrolithiasis. Each patient included in this study was receiving nonindinavir-based antiretroviral therapy. There were 41 men of whom 36 were Caucasian. Eleven (24%) patients underwent 24-hour urine collections with 11 metabolic abnormalities identified. Stone analysis was available for seven patients (four calcium oxalate monohydrate, one cystine, one uric acid, and one atazanavir). CONCLUSIONS: We report the largest series of nephrolithiasis in an HIV population since the introduction of HAART and highlight not only the similar prevalence of nephrolithiasis to the non-HIV population but also the lack of consistent comprehensive metabolic evaluations in HIV patients with recurrent nephrolithiasis.

Role of sialic acid in urinary cytoprotective activity of Tamm-Horsfall protein.

OBJECTIVES: Tamm-Horsfall protein (THP) from normal urine has been shown to protect against the cytotoxic effects of toxic urinary cations (TFs) in vivo and in vitro. This study investigated the effect of desialylation on the cytoprotective activity of THP. METHODS: From pooled 24-hour urine specimens from healthy individuals, both TFs and THP were obtained. Sprague-Dawley rats received intravesical NaCl or KCl, and the baseline urodynamic percentage of nonvoiding contractions (NVCs) was recorded. Then, rehydrated TF, TF plus THP, or TF plus THP-desialylated (THP-d) were instilled, followed by KCl, and the urodynamic measurements were repeated. In vitro, human HTB4 bladder cells were incubated overnight with the rehydrated TF, TF plus THP, TF plus THP-d, or assay media alone, and the cytotoxicity levels were determined. RESULTS: Acid hydrolysis resulted in an 88% loss of sialic acid. TFs consistently demonstrated a greater than 50% toxicity for human HTB4 cells compared with cells incubated in media (P <0.01). TF cytotoxic activity was blocked by preincubation with THP but not by preincubation with THP-d. Similarly, rat bladder NVCs increased significantly over baseline when KCl was infused after TF infusion (1.68 NVCs/min; P <0.0001). NVCs were significantly reduced by preincubating TFs with THP (0.42 NVCs/min) but not by preincubating with THP-d (1.55 NVCs/min, P <0.0001). CONCLUSIONS: The cytoprotective function of urinary THP in the bladder can be compromised when a decrease is present in the sialic acid residues on THP. Such a decrease in THP cytoprotective activity may play a critical role in the pathophysiology of interstitial cystitis.