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At least 361 records · Page 20Linked to original sources
PubMed1998

Is serum cystatin C a sensitive marker of glomerular filtration rate (GFR)? A preliminary study on renal transplant patients.

Human cystatin C is a basic low molecular mass protein (13,359 Dalton) freely filtered through the glomerulus and almost completely re-absorbed and catabolized by proximal tubular cells. We measured serum cystatin C in 38 kidney transplant patients (23 males, 15 females) aged between 6 and 32 years. To assess renal function, serum and urinary creatinine were also determined in all patients, and creatinine clearance was finally calculated. Cystatin C was determined by a particle-enhanced turbidimetric assay, and creatinine was measured by gas chromatography-mass spectrometry. To compare the diagnostic efficiency of cystatin C with that of creatinine, inulin clearance was performed on 12 renal transplant patients, and receiver operating characteristic (ROC) analysis was applied. The results of this study demonstrate that serum cystatin C significantly increases in renal transplant patients with reduced creatinine clearance (< 70 mL/min per 1.73 m2) and that the diagnostic accuracy of serum cystatin C is better than of serum creatinine. Cystatin C may be utilized as a very marker of reduced GFR.

Adolescent↗
PubMed1999

Cellular processing of the amyloidogenic cystatin C variant of hereditary cerebral hemorrhage with amyloidosis, Icelandic type.

An important gap in our understanding of the pathogenesis of the amyloidoses is the identification of the cellular events that lead from synthesis of an amyloid precursor protein to its conversion to the amyloid fiber subunit. We address this question by characterizing the effects of an amyloidogenic mutation on the intracellular processing of its protein product. The protein, a mutant of the cysteine protease inhibitor cystatin C, is the amyloid precursor protein in Hereditary Cerebral Hemorrhage with Amyloidosis--Icelandic type (HCHWA-I). The amyloid fibers are composed of mutant cystatin C (L68Q) that lacks the first 10 amino acids. We have previously shown that processing of wild-type cystatin C entails formation of a transient intracellular dimer that dissociates prior to secretion, such that extracellular cystatin C is monomeric. We report here that the cystatin C mutation engenders several alterations in its intracellular trafficking. It forms a stable intracellular dimer that is partially retained in the endoplasmic reticulum and degraded. The bulk of mutant cystatin C that is secreted does not dissociate and is secreted as an inactive dimer. Thus, formation of the stable mutant cystatin C dimer is an early event in the pathogenesis of this disease.

Amino Acid Sequence↗
PubMed2006

Evaluation of Dade Behring N Latex Cystatin C reagent on Abbott ci8200.

Plasma cystatin C has been shown in several studies to be superior to plasma creatinine for estimation of glomerular filtration rate (GFR). Reporting cystatin C results in mL/min using conversion formulas for transforming cystatin C expressed as mg/L to GFR expressed as mL/min has greatly facilitated the clinical use of the marker. At our hospital we have an increasing demand for cystatin C and at present we perform over 1400 cystatin C analyses a month. The test is available at all hours. This in combination with the volume emphasises the need to have the assay close to the routine chemistry instrument to reduce handling time per test and time to report test results. We have thus evaluated the Dade Behring N Latex Cystatin C assay (Dade Behring, Deerfield, IL, USA) on Architect ci8200 (Abbott Laboratories, Abbott Park, IL, USA). The nephelometric method on the ProSpec (Dade Behring) and the turbidimetric method on Architect ci8200 showed very good agreement (y = 1.0072x + 0.0042; R2 = 0.987). Accordingly, running the cystatin C analyses on a chemistry instrument (Architect ci8200) would be proper to increase the availability of the analysis and reduce turnaround times.

Cystatin C↗
PubMed2005

Cystatin C concentration as a risk factor for heart failure in older adults.

BACKGROUND: Previous studies that evaluated the association of kidney function with incident heart failure may be limited by the insensitivity of serum creatinine concentration for detecting abnormal kidney function. OBJECTIVE: To compare serum concentrations of cystatin C (a novel marker of kidney function) and creatinine as predictors of incident heart failure. DESIGN: Observational study based on measurement of serum cystatin C from frozen sera obtained at the 1992-1993 visit of the Cardiovascular Health Study. Follow-up occurred every 6 months. SETTING: Adults 65 years of age or older from 4 communities in the United States. PARTICIPANTS: 4384 persons without previous heart failure who had measurements of serum cystatin C and serum creatinine. MEASUREMENTS: Incident heart failure. RESULTS: The mean (+/-SD) serum concentrations of cystatin C and creatinine were 82 +/- 25 nmol/L (1.10 +/- 0.33 mg/L) and 89 +/- 34 micromol/L (1.01 +/- 0.39 mg/dL), respectively. During a median follow-up of 8.3 years (maximum, 9.1 years), 763 (17%) participants developed heart failure. After adjustment for demographic factors, traditional and novel cardiovascular risk factors, cardiovascular disease status, and medication use, sequential quintiles of cystatin C concentration were associated with a stepwise increased risk for heart failure in Cox proportional hazards models (hazard ratios, 1.0 [reference], 1.30 [95% CI, 0.96 to 1.75], 1.44 [CI, 1.07 to 1.94], 1.58 [CI, 1.18 to 2.12], and 2.16 [CI, 1.61 to 2.91]). In contrast, quintiles of serum creatinine concentration were not associated with risk for heart failure in adjusted analysis (hazard ratios, 1.0 [reference], 0.77 [CI, 0.59 to 1.01], 0.85 [CI, 0.64 to 1.13], 0.97 [CI, 0.72 to 1.29], and 1.14 [CI, 0.87 to 1.49]). LIMITATIONS: The mechanism by which cystatin C concentration predicts risk for heart failure remains unclear. CONCLUSIONS: The cystatin C concentration is an independent risk factor for heart failure in older adults and appears to provide a better measure of risk assessment than the serum creatinine concentration. *For a full list of participating Cardiovascular Health Study investigators and institutions, see http://www.chs-nhlbi.org.

Aged↗
PubMed1999

Changes in plasma cystatin C after renal transplantation and acute rejection in adults.

BACKGROUND: Cystatin C has recently been proposed as an alternative marker of glomerular filtration rate. The diagnostic value of plasma cystatin C for the longitudinal assessment of kidney function after renal transplantation, however, has not been addressed. METHODS: Renal function was evaluated in 30 adults receiving renal transplants (46 +/- 9 years, mean +/- SD) and in 56 healthy controls (38 +/- 10 years) using cystatin C. Plasma cystatin C was determined daily starting the day of surgery and for 3 weeks after surgery by an immunonephelometric assay. RESULTS: Plasma concentration significantly decreased during the first week (-44% vs -29% for creatinine). Plasma cystatin C correlated with plasma creatinine (r = 0.741; P <0.0001) and the reciprocal of the creatinine clearance estimated by the Cockcroft-Gault formula (r = 0.882; P <0.001). In all three cases of acute renal impairment, the increase in plasma cystatin C values was more prominent than that of creatinine. CONCLUSIONS: Plasma cystatin C is an alternative and accurate marker of allograft function in adult transplant patients. Increased sensitivity compared with creatinine for the detection of acute reduction in glomerular filtration rate allows in some cases a more rapid diagnosis of acute rejection or treatment nephrotoxicity.

Adult↗
PubMed1999

Cystatin C and creatinine after successful kidney transplantation in children.

BACKGROUND: Serum creatinine is commonly used for the monitoring of allograft function following renal transplantation (RTX). Due to lower muscle mass, creatinine production rate is reduced in children, thus decreasing its sensitivity for the detection of allograft dysfunction. In children, the serum concentration of cystatin C, a low molecular weight protein of 13.3 kDa, reflects glomerular filtration rate independent of age, height and body composition. We, therefore, sought to assess the potential of cystatin C as a marker of allograft function in children. METHODS: Cystatin C and creatinine were measured in parallel at least daily in 24 children (14 boys, 10 girls; mean age 10.5+/-5.1 years) during hospitalization after successful RTX. Cystatin was determined immunoturbidimetrically, creatinine enzymatically. RESULTS: Within one hour after RTX, cystatin C (mean+/-SE) almost halved from 6.69+/-0.45 mg/l to 3.69+/-0.38 mg/l while creatinine declined from 862 +/-65.4 to 633+/-62.9 micromol/l. Following a nadir of 1.82+/-0.18 mg/l on day 2, there was a secondary increase in cystatin C concentrations to 2.69+/-0.35 mg/l on day 10. Creatinine concentrations continued to decline until day 9 reaching 80.5+/-13.1 micromol/l. Day-to-day variation at steady-state was comparable. In the course of 9 acute rejection episodes, both parameters rose in parallel, the increase in creatinine concentration being much greater. CONCLUSION: Cystatin C was an early indicator of allograft function following successful RTX in children. It did not prove superior to creatinine for the recognition of acute allograft dysfunction, however.

Biomarkers↗
PubMed2001

Identification and localization of retinal cystatin C.

PURPOSE: Cystatin C is a mammalian cysteine protease inhibitor, synthesized in various amounts by many kinds of cells and appearing in most body fluids. There are reports that it may be synthesized in the mammalian retina and that a cysteine protease inhibitor may influence the degradation of photoreceptor outer segment proteins. In the current study cystatin C was identified, quantitated, and localized in mouse, rat, and human retinas. METHODS: Enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (RT-PCR), DNA sequencing, Western blot analysis, and immunohistochemistry have been used on mouse, rat, and human retinas (pigment epithelium included). RESULTS: Cystatin C is present in high concentrations in the normal adult rat retina, as it is throughout its postnatal development. Its concentration increases to a peak at the time when rat pups open their eyes and then remains at a high level. It is mainly localized to the pigment epithelium, but also to some few neurons of varying types in the inner retina. Cystatin C is similarly expressed in normal mouse and human retinas. CONCLUSIONS: Cystatin C was identified and the localization described in the retinas of rat, mouse, and human using several techniques. Cystatin C is known to efficiently inactivate certain cysteine proteases. One of them, cathepsin S, is present in the retinal pigment epithelium and affects the proteolytic processing by cathepsin D of diurnally shed photoreceptor outer segments. Hypothetically, it appears possible that retinal cystatin C, given its localization to the pigment epithelium and its ability to inhibit cathepsin S, could be involved in the regulation of photoreceptor degradation.

Aged↗
PubMed2006

Does HAART improve renal function? An association between serum cystatin C concentration, HIV viral load and HAART duration.

AIM: The prevalence of renal disease in human HIV-infected individuals varies between 2% and 10%. Many reports have demonstrated the beneficial effect of antiretroviral (ARV) therapy on slowing the progression of renal diseases. The aim of our cross-sectional study was to determine serum cystatin C concentration in different stages of HIV infection and the relationship between cystatin C concentration and ARV treatment. METHODS: Cystatin C concentration was measured in the sera of 77 HIV-1-infected individuals and 18 HIV-seronegative volunteers. The glomerular filtration rate (GFR) was estimated using the Modification of Diet in Renal Disease Study formula. RESULTS: HIV infection resulted in a significant increase in serum cystatin C concentration compared with healthy individuals (933.4 +/- 32.1 vs 621.1 +/- 56.8 ng/ml, P < 0.001). There were no significant differences in urea, creatinine and GFR between those groups. On multivariate analyses serum cystatin C was independently associated with highly active antiretroviral therapy (HAART) duration (beta = -0.34, P = 0.04) and HIV viral load (beta = 0.33, P = 0.04), whereas there were no significant relationships with age, body mass index, HIV duration, CD4+ and CD8+ T-cell counts and serum high sensitivity C-reactive protein concentration. CONCLUSIONS: Our initial observations indicate that serum cystatin C, which may reflect mild renal dysfunction, is increased during HIV-infection and is associated with HIV viral load. Long-lasting HAART seems to decrease cystatin C concentration, thus potentially improves renal function.

Adult↗
PubMed2005

Comparative phylogenetic analysis of cystatin gene families from arabidopsis, rice and barley.

The plant cystatins or phytocystatins comprise a family of specific inhibitors of cysteine proteinases. Such inhibitors are thought to be involved in the regulation of several endogenous processes and in defence against pests and pathogens. Extensive searches in the complete rice and Arabidopsis genomes and in barley EST collections have allowed us to predict the presence of twelve different cystatin genes in rice, seven in Arabidopsis, and at least seven in barley. Structural comparisons based on alignments of all the protein sequences using the CLUSTALW program and searches for conserved motifs using the MEME program have revealed broad conservation of the main motifs characteristic of the plant cystatins. Phylogenetic analyses based on their deduced amino acid sequences have allowed us to identify groups of orthologous cystatins, and to establish homologies and define examples of gene duplications mainly among the rice and barley cystatin genes. Moreover, the absence of a counterpart between the two monocots, as well as strong variations in the motifs that interact with the cysteine proteinases, may be related to a species-specific evolutionary process. This cystatin classification should facilitate the assignment of proteinase specificities and functions to other cystatins as new information is obtained.

Amino Acid Motifs↗
PubMed1994

Monoclonal antibodies as probes to detect conformational changes in the rat cysteine proteinase inhibitor cystatin A.

Five monoclonal antibodies (MAbs), 77, 114, 138, 175 and 187, were established for rat cystatin A. MAbs 77, 114, 138 and 175 were shown to belong to the IgG1 subclass, whereas MAb 187 was an IgM. These MAbs partially suppressed inhibitory activity of rat cystatin A to papain. Their epitopes were mapped in detail on the molecule by examining the reactivities of the MAbs with NH2-terminally truncated forms and fragments of rat cystatin A by an enzyme-linked immunosorbent assay (ELISA), and by reactivity with the inhibitor on immunoblotting. In competitive binding assays the MAbs did not compete with each other, indicating that the epitopes recognized by these MAbs were substantially different. The conformational epitope recognized by the three MAbs 114, 138 and 175 belonged to one group that was highly sensitive to denaturation, but those epitopes were unchanged by NH2-terminal truncation. MAb 187 was able to recognize a linear epitope present in amino acid residues 15-50 in the NH2-terminal region. MAbs 77 and 114 reacted weakly with mouse cystatin A but not at all with human cystatin A, whereas MAb 187 reacted similarly with mouse cystatin A but at about half that level with human. The MAbs produced in this study should be useful tools for detecting conformational changes in the rat cystatin A molecule.

Amino Acid Sequence↗
PubMed1992

Immunolocalization of cystatin A in neoplastic, virus and inflammatory lesions of the uterine cervix.

Cystatin A was immunohistochemically demonstrated in the normal squamous epithelium of the uterine cervix, particularly in the parabasal and superficial cell layers whereas it was absent or scanty in the basal cells and in areas with parakeratosis. Cystatin A was also found in neoplastic lesions (dysplasia, carcinoma in situ and squamous cell carcinoma), but less abundant than in normal squamous epithelium. The immunoreaction in intraepithelial neoplasia was closely related to the degree of morphological maturation of the squamous cells with more abundant cystatin A in low grade dysplasia and less in high grade dysplasia and carcinoma in situ. In squamous cell carcinoma, cystatin A was often abundant in highly differentiated areas and almost absent in poorly differentiated ones. Cystatin A was found in the squamous epithelium in herpes and in condylomatous lesions. It was also found in the cytoplasm of neutrophils, but not in lymphocytes and plasma cells. In unspecific cervicitis, cystatin A was found extracytoplasmatically as small vesicles in the epithelial-stromal junction. The implications of cystatin A in neoplastic, virus, and inflammatory processes are discussed.

Carcinoma in Situ↗
PubMed2000

Purification and characterization of a cystatin from the leaves of methyl jasmonate treated tomato plants.

A multidomain cystatin was purified from the leaves of mature and seedling tomato plants (Lycopersicon esculentum, cv Bonnie Best) that had been sprayed with methyl jasmonate. For seedlings, cystatin purification was accomplished using EDTA washing, KCI extraction, 70 degrees C heat treatment, ammonium sulfate fractionation and gel filtration chromatography. For mature plants, DEAE chromatography was also needed to separate a protease, hydrolysis products of cystatin and serine proteinase inhibitors from the intact cystatin. Purified tomato cystatin has a molecular weight (Mr) of 88 kDa, eight papain binding domains, is a non-competitive inhibitor of papain with K1 of 1.4 nM and is not a glycoprotein. Tryptic peptides (Mr 26, 13 kDa) and most chymotryptic peptides (Mr 33, 13 kDa) of tomato cystatin retain inhibitor activity. Amino acid analysis revealed no Cys; Asx, Glx, Gly, Ser accounted for almost half the residues and there was some homology with potato multicystatin. Activity is stable at pH 4-11 at 4 degrees C, but unstable at neutral pH at > 60 degrees C (Ea = 92.5 kJ/mole). Extracts of mature plants treated with methyl jasmonate contain lower Mr cystatins that appear to result from the action of an endogenous 26 kDa protease on the 88 kDa inhibitor.

Acetates↗
PubMed2004

Extracellular production of a functional soy cystatin by Bacillus subtilis.

A recombinant Bacillus subtilis producing soy cystatin was developed by subcloning with a soy cystatin gene cloned in Escherichia coli. An active form of cystatin against the cysteine protease from Pacific whiting fillets contaminated with Myxosporidia parasite was constitutively expressed and secreted extracelluarly into the medium. Two gene fragments of signal peptides from kerA and sacB were introduced and compared for secretion efficiency of cystatin. The secretion level of active cystatin improved with the signal peptide of kerA when compared to that of sacB. Inhibitor activity was reduced rapidly after peak expression of the target protein at 36 h of fermentation. The addition of 1% glucose, a suppressor of protease, into the medium sustained the increase of the cystatin activity during fermentation. This study introduced a potential new method for fermentation production of cystatin.

Bacillus subtilis↗
PubMed1994

Differential changes in the association and dissociation rate constants for binding of cystatins to target proteinases occurring on N-terminal truncation of the inhibitors indicate that the interaction mechanism varies with different enzymes.

The importance of the N-terminal region of human cystatin C or chicken cystatin for the kinetics of interactions of the inhibitors with four cysteine proteinases was characterized. The association rate constants for the binding of recombinant human cystatin C to papain, ficin, actinidin and recombinant rat cathepsin B were 1.1 x 10(7), 7.0 x 10(6), 2.4 x 10(6) and 1.4 x 10(6) M-1.s-1, whereas the corresponding dissociation rate constants were 1.3 x 10(-7), 9.2 x 10(-6), 4.6 x 10(-2) and 3.5 x 10(-4) s-1. N-Terminal truncation of the first ten residues of the inhibitor negligibly affected the association rate constant with papain or ficin, but increased the dissociation rate constant approx. 3 x 10(4)- to 2 x 10(6)-fold. In contrast, such truncation decreased the association rate constant with cathepsin B approx. 60-fold, while minimally affecting the dissociation rate constant. With actinidin, the truncated cystatin C had both an approx. 15-fold lower association rate constant and an approx. 15-fold higher dissociation rate constant than the intact inhibitor. Similar results were obtained for intact and N-terminally truncated chicken cystatin. The decreased affinity of human cystatin C or chicken cystatin for cysteine proteinases after removal of the N-terminal region is thus due to either a decreased association rate constant or an increased dissociation rate constant, or both, depending on the enzyme. This behaviour indicates that the contribution of the N-terminal segment of the two inhibitors to the interaction mechanism varies with the target proteinase as a result of structural differences in the active-site region of the enzyme.

Animals↗
PubMed1998

Leukocystatin, a new Class II cystatin expressed selectively by hematopoietic cells.

We describe a new cystatin in both mice and humans, which we termed leukocystatin. This protein has all the features of a Class II secreted inhibitory cystatin but contains lysine residues in the normally hydrophobic binding regions. As determined by cDNA library Southern blots, this cystatin is expressed selectively in hematopoietic cells, although fine details of the distribution among these cell types differ between the human and mouse mRNAs. In addition, we have determined the genomic organization of mouse leukocystatin, and we found that in contrast to most cystatins, the leukocystatin gene contains three introns. The recombinant proteins corresponding to these cystatins were expressed in Escherichia coli as N-terminal glutathione S-transferase or FLAGTM fusions, and studies showed that they inhibited papain and cathepsin L but with affinities lower than other cystatins. The unique features of leukocystatin suggests that this cystatin plays a role in immune regulation through inhibition of a unique target in the hematopoietic system.

Amino Acid Sequence↗
PubMed1995

Immunolocalization of cystatin A in condylomatous and dysplastic lesions of the human uterine cervix: correlation with the presence and type of human papillomavirus infection.

Cystatin A is the major cysteine proteinase inhibitor in human squamous epithelia. We investigated the occurrence of cystatin A in normal, condylomatous, and dysplastic lesions of the cervix with or without human papillomavirus (HPV) infection. Cystatin A was detected by immunohistochemistry and HPV infection by DNA hybridization techniques. In the normal uterine cervix, cystatin A was seen throughout the epithelium, except in the basal and parabasal cell layers. In condylomatous lesions, the staining intensity was similar to that in normal epithelium. In low-grade cervical intraepithelial neoplasias (CINs), reduced staining was seen in the lower third of the epithelium; in high-grade CINs, a reduction in staining intensity was also seen in the middle and upper thirds. Cystatin A staining in epithelia and nuclei was negative in highly cellular and poorly differentiated CIN III. The cytoplasmic staining of cystatin A did not correlate with presence or type of HPV DNA. In the high-grade CINs infected with HPV types 16 and 18, however, cystatin A staining was more often confined to the nuclear compartment.

Cystatins↗
PubMed2006

Prevention of amyloid fibril formation of amyloidogenic chicken cystatin by site-specific glycosylation in yeast.

To address the role of glycosylation on fibrillogenicity of amyloidogenic chicken cystatin, the consensus sequence for N-linked glycosylation (Asn106-Ile108 --> Asn106-Thr108) was introduced by site-directed mutagenesis into the wild-type and amyloidogenic chicken cystatins to construct the glycosylated form of chicken cystatins. Both the glycosylated and unglycosylated forms of wild-type and amyloidogenic mutant I66Q cystatin were expressed and secreted in a culture medium of yeast Pichia pastoris transformants. Comparison of the amount of insoluble aggregate, the secondary structure, and fibrillogenicity has shown that the N-linked glycosylation could prevent amyloid fibril formation of amyloidogenic chicken cystatin secreted in yeast cells without affecting its inhibitory activities. Further study showed this glycosylation could inhibit the formation of cystatin dimers. Therefore, our data strongly suggested that the mechanism causing the prevention of amyloidogenic cystation fibril formation may be realized through suppression of the formation of three-dimensional domain-swapped dimers and oligomers of amyloidogenic cystatin by the glycosylated chains at position 106.

Amyloid↗
PubMed2006

Modulating the proteinase inhibitory profile of a plant cystatin by single mutations at positively selected amino acid sites.

Cysteine proteinase inhibitors of the cystatin superfamily have several important functions in plants, including the inhibition of exogenous cysteine proteinases during herbivory or infection. Here we used a maximum-likelihood approach to assess whether plant cystatins, like other proteins implicated in host-pest interactions, have been subject to positive selection during the course of their evolution. Several amino acid sites were identified as being positively selected in cystatins from either Poaceae (monocots) and Solanaceae (dicots). These hypervariable sites were located at strategic positions on the protein: on each side of the conserved glycine residues in the N-terminal trunk, within the first and second inhibitory loops entering the active site of target enzymes, and surrounding the larfav motif, a sequence of unknown function conserved among plant cystatins. Supporting the assumption that positively selected, hypervariable sites are indicative of amino acid sites implicated in functional diversity, mutants of the 8th cystatin unit of tomato multicystatin including alternative residues at positively selected sites in the N-terminal trunk exhibited highly variable affinities for the cysteine proteases papain, cathepsin B and cathepsin H. Overall, these observations support the hypothesis that plant cystatins have been under selective pressure to evolve in response to predatory challenges by herbivorous enemies. They also indicate the potential of site-directed mutagenesis at positively selected sites for the generation of cystatins with improved binding properties.

Amino Acids↗