Cystatin C


Cystatin C or cystatin 3, a protein encoded by the CST3 gene, is mainly used as a biomarker of kidney function. Recently, it has been studied for its role in predicting new-onset or deteriorating cardiovascular disease. It also seems to play a role in brain disorders involving amyloid, such as Alzheimer's disease.
In humans, all cells with a nucleus produce cystatin C as a chain of 120 amino acids. It is found in virtually all tissues and body fluids. It is a potent inhibitor of lysosomal proteinases and probably one of the most important extracellular inhibitors of cysteine proteases. Cystatin C belongs to the type 2 cystatin gene family.

Role in medicine

Kidney function

, a marker of kidney health, is most accurately measured by injecting compounds such as inulin, radioisotopes such as 51chromium-EDTA, 125I-iothalamate, 99mTc-DTPA or radiocontrast agents such as iohexol, but these techniques are complicated, costly, time-consuming and have potential side-effects.
Creatinine is the most widely used biomarker of kidney function. It is inaccurate at detecting mild renal impairment, and levels can vary with muscle mass but not with protein intake. Urea levels might change with protein intake.
Formulas such as the Cockcroft and Gault formula and the MDRD formula try to adjust for these variables.
Cystatin C has a low molecular weight, and it is removed from the bloodstream by glomerular filtration in the kidneys. If kidney function and glomerular filtration rate decline, the blood levels of cystatin C rise. Cross-sectional studies suggest that serum levels of cystatin C are a more precise test of kidney function than serum creatinine levels.. Longitudinal studies are sparse, but some show promising results.
Cystatin C levels are less dependent on age, gender, ethnicity and muscle mass compared to creatinine. Cystatin C measurements alone have not been shown to be superior to formula-adjusted estimations of kidney function. As opposed to previous claims, cystatin C has been found to be influenced by body composition.
It has been suggested that cystatin C might predict the risk of developing chronic kidney disease, thereby signaling a state of 'preclinical' kidney dysfunction.
Studies have also investigated cystatin C as a marker of kidney function in the adjustment of medication dosages.
Cystatin C levels have been reported to be altered in patients with cancer, thyroid dysfunction and glucocorticoid therapy in some but not all situations. Other reports have found that levels are influenced by cigarette smoking and levels of C-reactive protein. However, inflammation does not cause an increase in the production of cystatin C, since elective surgical procedures, producing a strong inflammatory response in patients, do not change the plasma concentration of cystatin C. Levels seem to be increased in HIV infection, which might or might not reflect actual renal dysfunction. The role of cystatin C to monitor GFR during pregnancy remains controversial. Like creatinine, the elimination of cystatin C via routes other than the kidney increase with worsening GFR.

Death and cardiovascular disease

Kidney dysfunction increases the risk of death and cardiovascular disease.
Several studies have found that increased levels of cystatin C are associated with the risk of death, several types of cardiovascular disease and healthy aging. Some studies have found cystatin C to be better in this regard than serum creatinine or creatinine-based GFR equations.
Because the association of cystatin C with long term outcomes has appeared stronger than what could be expected for GFR, it has been hypothesized that cystatin C might also be linked to mortality in a way independent of kidney function. In keeping with its housekeeping gene properties, it has been suggested that cystatin C might be influenced by the basal metabolic rate.

Cystatin C, Shrunken Pore Syndrome and mortality

The glomerular sieving coefficients for 10–30 kDa plasma proteins in the human kidney are relatively high with coefficients between 0.9 and 0.07. These relatively high sieving coefficients, combined with the high production of ultrafiltrate in health, means that proteins ≤ 30 kDa in plasma normally are mainly cleared by the kidneys and at least 85% of the clearance of cystatin C occurs in the kidney. If the pores of the glomerular membrane shrink, the filtration of bigger molecules, e.g. cystatin C, will decrease, whereas the filtration of small molecules, like water and creatinine, will be less affected. In this case, cystatin C-based estimates of GFR, eGFRcystatin C, will be lower than creatinine-based estimates eGFRcreatinine, so that this condition, named Shrunken Pore Syndrome can be identified by a low eGFRcystatin C/eGFRcreatinine-ratio. This syndrome is associated with a very strong increase in mortality. The capacity of cystatin C to identify this syndrome might therefore explain the marked superiority of eGFRcystatin C, or cystatin C, over eGFRcreatinine, or creatinine, in predicting end-stage renal disease, cardiovascular manifestations, hospitalisation or death.

Neurologic disorders

s in the cystatin 3 gene are responsible for the Icelandic type of hereditary cerebral amyloid angiopathy, a condition predisposing to intracerebral haemorrhage, stroke and dementia. The condition is inherited in a dominant fashion.
Since cystatin 3 also binds amyloid β and reduces its aggregation and deposition, it is a potential target in Alzheimer's disease. Although not all studies have confirmed this, the overall evidence is in favor of a role for CST3 as a susceptibility gene for Alzheimer's disease. Cystatin C levels have been reported to be higher in subjects with Alzheimer's disease.
The role of cystatin C in multiple sclerosis and other demyelinating diseases remains controversial.

Other roles

Cystatin C levels are decreased in atherosclerotic and aneurysmal lesions of the aorta.
Genetic and prognostic studies also suggest a role for cystatin C.
Breakdown of parts of the vessel wall in these conditions is thought to result from an imbalance between proteinases and their inhibitors.
A few studies have looked at the role of cystatin C or the CST3 gene in age-related macular degeneration. Cystatin C has also been investigated as a prognostic marker in several forms of cancer. Its role in pre-eclampsia remains to be confirmed.

Laboratory measurement

Cystatin C can be measured in a random sample of serum using immunoassays such as nephelometry or particle-enhanced turbidimetry. It is a more expensive test than serum creatinine, which can be measured with a Jaffé reaction.
Reference values differ in many populations and with sex and age. Across different studies, the mean reference interval was between 0.52 and 0.98 mg/L. For women, the average reference interval is 0.52 to 0.90 mg/L with a mean of 0.71 mg/L. For men, the average reference interval is 0.56 to 0.98 mg/L with a mean of 0.77 mg/L.
The normal values decrease until the first year of life, remaining relatively stable before they increase again, especially beyond age 50. Creatinine levels increase until puberty and differ according to gender from then on, making their interpretation problematic for pediatric patients.
In a large study from the United States National Health and Nutrition Examination Survey, the reference interval was between 0.57 and 1.12 mg/L. This interval was 0.55 - 1.18 for women and 0.60 - 1.11 for men. Non-Hispanic blacks and Mexican Americans had lower normal cystatin C levels. Other studies have found that in patients with an impaired renal function, women have lower and blacks have higher cystatin C levels for the same GFR. For example, the cut-off values of cystatin C for chronic kidney disease for a 60-year-old white women would be 1.12 mg/L and 1.27 mg/L in a black man. For serum creatinine values adjusted with the MDRD equation, these values would be 0.95 mg/dL to 1.46 mg/dL.
Based on a threshold level of 1.09 mg/L, the prevalence of increased levels of cystatin C in the United States was 9.6% in subjects of normal weight, increasing in overweight and obese individuals. In Americans aged 60 and 80 and older, serum cystatin is increased in 41% and more than 50%.

Molecular biology

The cystatin superfamily encompasses proteins that contain multiple cystatin-like sequences. Some of the members are active cysteine protease inhibitors, while others have lost or perhaps never acquired this inhibitory activity. There are three inhibitory families in the superfamily, including the type 1 cystatins, type 2 cystatins and the kininogens. The type 2 cystatin proteins are a class of cysteine proteinase inhibitors found in a variety of human fluids and secretions, where they appear to provide protective functions. The cystatin locus on the short arm of chromosome 20 contains the majority of the type 2 cystatin genes and pseudogenes.
The CST3 gene is located in the cystatin locus and comprises 3 exons, spanning 4.3 kilo-base pairs. It encodes the most abundant extracellular inhibitor of cysteine proteases. It is found in high concentrations in biological fluids and is expressed in virtually all organs of the body. The highest levels are found in semen, followed by breastmilk, tears and saliva. The hydrophobic leader sequence indicates that the protein is normally secreted. There are three polymorphisms in the promoter region of the gene, resulting in two common variants. Several single nucleotide polymorphisms have been associated with altered cystatin C levels.
Cystatin C is a non-glycosylated, basic protein. The crystal structure of cystatin C is characterized by a short alpha helix and a long alpha helix which lies across a large antiparallel, five-stranded beta sheet. Like other type 2 cystatins, it has two disulfide bonds. Around 50% of the molecules carry a hydroxylated proline. Cystatin C forms dimers by exchanging subdomains; in the paired state, each half is made up of the long alpha helix and one beta strand of one partner, and four beta strands of the other partner.

History

Cystatin C was first described as 'gamma-trace' in 1961 as a trace protein together with other ones in the cerebrospinal fluid and in the urine of people with kidney failure. Grubb and Löfberg first reported its amino acid sequence. They noticed it was increased in patients with advanced kidney failure. It was first proposed as a measure of glomerular filtration rate by Grubb and coworkers in 1985.
Use of serum creatinine and cystatin C was found very effective in accurately reflecting the GFR in a study reported in the July 5, 2012 issue of the New England Journal of Medicine.