Creatinine and Cystatin C: A Measure of Renal Function in Men With Testosterone-Induced Muscle Hypertrophy

Abstract

Creatinine (Cr) is often used as a standalone gold standard marker of kidney function. Cystatin C (Cys C) is a less physiologically labile marker of renal function, particularly in certain subgroups. Herein, we analyze trends in cystatin C as compared to creatinine in men on testosterone replacement therapy with varying body mass indices and percent body fat (PBF). This retrospective analysis observes 227 men with testosterone-induced muscle hypertrophy who visited a men’s health tertiary care clinic. All participants were characterized as competitive or recreational athletes. In patients with a normal body mass index (BMI), there was no clinically significant correlation between Cr and Cys C. Slight correlation was seen with overweight (R² = .27) patients (p < .0001) and obese (R² = .29) patients (p < .0001). Patients with PBF of 0%–10% (n = 22) exhibited minimal (R² = .23) positive correlation between Cys C and Cr (p = .03). Positive correlation between Cys C and Cr in patients with PBF of 10%–20% was clinically negligible (R² = .17, n = 87), modest (R² = .49) in patients with PBF of 20%–30% (n = 42), and evident (R² = 1.00) in patients >30% (n = 3) (p < .0001, respectively). Cystatin C measurements display less variance compared with creatinine at differing BMI distinctions. At the upper limit of BMI or PBF in our patient population, cystatin C exhibits minimal to moderate variability compared with creatinine.

Introduction

Estimation of renal function continues to undergo evaluation throughout recent medical history. Glomerular filtration rate (GFR) is calculated with creatinine (Cr) and is the basis of diagnosing renal injury, renal failure, and renal function. We have identified men undergoing testosterone therapy (TTh) as a demographic of patients who may benefit from improved GFR calculation. Men who self-identify as elite athletes or engaging in competitive recreational activities while on testosterone are speculated to have a high level of muscle mass that may contribute to elevations in Cr. In this population, Cr may underestimate renal function. Herein, we focus on the utilization of cystatin C (Cys C) as a measure of renal function.

While complex procedures using exogenous markers of GFR such as inulin would be the physiologically ideal measurement, these studies are costly and ineffective for patient care. Instead, clinicians turn to calculated estimations to evaluate renal function, many of which utilize serum creatinine (Cr) as a filtration marker. However, novel studies have demonstrated the inaccuracies of Cr in estimated GFR (eGFR) due to systematic errors from muscle mass, tubular secretion, and intrinsic proportional bias, especially at extremes of patient muscularity (Nankivell et al., 2020). Clinicians have focused on implementing the Modification of Diet in Renal Disease (MDRD) and Chronic Kidney Disease Epidemiology (CKD-EPI) equations, used without the race component as of 2021 to more effectively utilize serum Cr. Refinements in the calculation of eGFR have identified Cys C as a candidate for an endogenous marker of renal function (Ferguson et al., 2015; Nankivell et al., 2020).

Previously published studies have demonstrated that calculating eGFR with Cys C can result in improved accuracy as compared to Cr alone without the consideration of sex or race (Pottel et al., 2023). Cys C is a protein produced by most human tissues and body fluids. It has a low molecular weight and is readily catabolized by tubular epithelial cells following glomerular filtration. For these reasons, it does not exhibit the interindividual variability seen with Cr measurements (Chew-Harris et al., 2013; Dharnidharka et al., 2002). The uniformity of Cys C and its efficacy in indicating declining renal function suggest it to be a valid option for calculating eGFR in patients with high body muscle mass compared with Cr. In addition, the use of Cys C circumvents the race component that has traditionally been incorporated into Cr-based eGFR calculations (Pottel et al., 2023).

Cr generation in the body is primarily determined by muscle mass and dietary intake which creates high variability depending on patient factors, making it a misleading marker and not universally generalizable (Stevens et al., 2006). Elite athletes are noted to have an elevated BMI, despite an overall favorable level of PBF and fitness. This is largely due to increased proportion of body muscle and lower PBF. We propose that Cys C may be used as a more accurate alternative to Cr in men with muscle hypertrophy.

Cr generation in the body is also impacted by steroid usage. Bodybuilders who use anabolic androgenic steroids and dietary supplements have significantly higher Cys C and Cr levels compared with those who do not use anabolic androgenic steroids (Ozkurt et al., 2023). Concomitant use of stimulants with anabolic steroids further increases Cr levels (Ostovar et al., 2017). Anabolic steroid use is also linked to glomerular abnormalities, proteinuria, and CKD such as focal segmental glomerulosclerosis (Ostovar et al., 2017; Parente Filho et al., 2020).

Furthermore, Cr generation in the body is both impacted by thyroid disease and impacts thyroid disease. Thyroid disorders influence kidney function parameters such as serum Cr and Cys C in opposite directions. Specifically, in hypothyroidism, serum Cr is comparatively higher and decreases after thyroid function is at the patient’s baseline (Echterdiek et al., 2022). Hypothyroidism impacts kidney function leading to reduced GFR, increased serum Cr, and altered kidney/glomerular structure. In contrast, cystatin C is lower in hypothyroidism and increases after thyroid function normalization (Echterdiek et al., 2022). In addition to PBF and BMI, patients who deviate from a euthyroid state may have altered Cr and Cys C laboratory values (Echterdiek et al., 2022). CKD has also been shown to impact thyroid function. Nephrotic syndrome is associated with changes in serum thyroid hormone concentrations (Iglesias et al., 2017). Subclinical hypothyroidism and euthyroid sick syndrome are characterized by low T3 and are common in patients with CKD (Iglesias et al., 2017). In kidney transplant patients, low T3 syndrome is the most common thyroid abnormality and has been linked to higher mortality risk and poorer graft outcomes. In summary, CKD and thyroid disease both have differing and negative effects on the other (Iglesias et al., 2017).

This study evaluates the relationship between Cr, Cys C, BMI, and PBF in elite athlete populations. By analyzing fluctuations in Cr and Cys C in patients of different BMI and PBF, we hope to identify the more consistent endogenous marker of renal function in patients with testosterone-induced muscle hypertrophy.

Currently, the particle-enhanced nephelometric immunoassay (PENIA) developed for the Dade Behring nephelometers is one of the most frequently used assay for Cys C (Ji et al., 2017). Studies show variation among assays, and as Cys C becomes more widely adopted, more assays are likely to become available (Ebert & Shlipak, 2020). As preference for Cys C as a tool for eGFR becomes more pronounced, demand for the test should reduce the high cost relative to Cr.

In this study, we retrospectively analyze the aggregate correlation between Cys C and Cr among self-identifying male elite athletes of varied BMI and PBF.

Methods

Our study included 227 male elite athletes presenting to a single men’s health tertiary clinic with testosterone-induced muscle hypertrophy. These men had no past medical history of renal disease excluding well-controlled diabetes mellitus, well-controlled hypertension, or treated nephrolithiasis. Data were analyzed via retrospective analysis per institutional review board (IRB) approval. Elite athletes were defined as those pursuing competition in professional or amateur sporting events. All included patients self-reported testosterone use. Evaluation consisted of chart review of pertinent past medical history, calculation of BMI, PBF via bioelectrical impedance analysis device, and laboratories including serum Cr and Cys C. Patients were categorized based on BMI into three subgroups: normal (BMI = 20–25), overweight (BMI = 25–30), and obese (BMI > 30). BMI was calculated using the standard Centers for Disease Control (CDC) defined calculation. In addition, patients were stratified into categories based on PBF (0%–10%, 10%–20%, 20%–30%, or >30%) where available (n = 154).

We then utilized linear regression analysis to correlate each patient’s Cys C and creatinine values across each BMI and PBF subset independently.

Results

In total, 227 elite athletes were identified and stratified by BMI and PBF. The majority of patients were classified as “overweight” (n = 88) or “obese” (n = 129) based on BMI. Demographic breakdown of the study population is outlined in Table 1. In total, 19.8% of the patients had medically stable hypertension (HTN) and 7% of patients had medically stable thyroid disease. In patients with a normal BMI, there was no statistically significant correlation between Cr and Cys C (R² = .0001, p = .98). Clinically minimal correlation was seen among overweight (R² = .27) patients (p < .0001) and obese (R² = .29) patients (p < .0001). Patients with PBF of 0%–10% (n = 22) exhibited clinically minimal (R² = .23) positive correlation between Cys C and Cr (p = .03). Positive correlation between Cys C and Cr in patients with PBF of 10%–20% was clinically negligible (R² = .17, n = 87), modest (R² = .49) in patients with PBF of 20%–30% (n = 42), and evident (R² = 1.00) in patients >30% (n = 3) (p < .0001), respectively. These findings are summarized in Figure 1A and 1B.

Table 1.

A Summary of Demographic Data. Interquartile Ranges are Represented in Parentheses

Demographics
Total number of patients	N = 227
Age	41 (34.4, 50)
BMI	30.64 (28.52, 33.12)
PBF	15.6 (13.1, 21.2)
Creatinine	1.46 mg/L (1.4, 1.55)

Figure 1.

Figure 1A. Serum Cystatin C (Cys C) of Each Body Mass Index (BMI) Subgroup Plotted Against Serum Creatinine (Cr) for Each Patient Analyzed. Best Fit Lines Yielded From Linear Regression Analysis are Represented for Each BMI Subgroup: Normal, Overweight, and Obese

Discussion

The inaccuracy of GFR calculations using creatinine as a filtration maker in certain patient populations is well documented in the literature (Nankivell et al., 2020). Creatinine generation in the body is primarily determined by muscle mass and dietary intake. In populations of elite athletes, muscle mass is proportionally much higher than the general population, and our study population affirms that these elite athletes predominantly maintain an increased BMI with low PBF. Concordantly, our study confirms that elevated Cr is expected in men with high muscle mass, indicated by relatively high BMI and lean PBF. Cys C does not fluctuate as dramatically in this population indicated by clinically insignificant correlations between Cys C and Cr among various subgroups. However, a minimal to moderate correlative trend is seen in patients with markedly elevated BMI. Overall, we demonstrate Cys C as the marker less influenced by BMI and, therefore, more accurately reflective of renal function in this patient population.

The Kidney Disease: Improving Global Outcomes (KDIGO) 2012 CKD guidelines suggest using supplementary tests in addition to Cr-based estimates (such as Cys C or a clearance measurement) for confirmatory testing in specific circumstances when eGFR based on serum Cr is less accurate (Kellum et al., 2013). At least seven elite athletes of our cohort fit this category in which Cr would likely be less accurate. Currently, Cys C is mostly used as a confirmatory marker; however, in elite athletes, it may have a future role as the primary or sole renal filtration biomarker.

Accurate GFR estimation is critical in key clinical decisions including diagnosis of CKD, initiation of hemodialysis, or consideration for renal transplantation. Inaccurate estimations of kidney function can severely adversely affect certain patient populations in delaying appropriate renal interventions (Uppal et al., 2022). Cr, a product of creatinine phosphate, was implemented into basic medical practice due to its relatively constant secretion rate and insignificant reabsorption in the nephron. In addition, well-documented patient factors that influence Cr measurements include muscle mass, activity level, diet, chronic systemic conditions, and usage of specific medications that inhibit creatinine secretion (Shlipak et al., 2022). Prior studies have established that individuals with lower muscle mass have a conversely higher creatinine excretion rate and vice versa. These differences are assumed to be accounted for in equations for calculating eGFR, but data show this is ultimately inaccurate with Cr alone (Groothof et al., 2022). One confounding factor in many of these potentially GFR modifying factors we have identified is the subset of men on TTh. Testosterone contributes to increased muscle mass and has relatively little physiologic interplay with renal metabolic processes (Li et al., 2022). Therefore, we can assume that testosterone effects on Cr are exerted via its influences on muscle mass.

Contrastingly, Cys C has been identified as has having less historic evidence of fluctuations with PBF and BMI; however, ongoing research continues to elucidate this question (Andersen et al., 2009; Filler et al., 2005; Groesbeck et al., 2008; Knight et al., 2004). From a cell biology perspective, Cys C is produced by all nucleated cells in the body regardless of variations in BMI or PBF (Pasala & Carmody, 2017). While foundational cell biology would suggest muscle mass could not affect Cys C measurements, further research continues to elucidate the clinical nuance.

TTh to achieve physiologic levels has been a growing aspect of urological practice for decades now. Reported trends estimate that from 2000 to 2011 laboratory testing and prescription of testosterone in the United States and United Kingdom increased fourfold (Layton et al., 2014). Associated effects of TTh include increased muscle mass. When these effects are observed in elite-level athletes who deliberately attain an increased muscle mass, BMI, and PBF outside of the norm of renal study populations, unmeasurable physiologic variability prompts GFR inaccuracies when using Cr measurements. Consequently, additional, or newly innovative points of data such as Cys C can appropriately address clinical management of this growing patient base.

The limitations of our study are inherent to a single institution retrospective study, including limited data availability. A major limitation of this article is that the patients were not stratified by additional comorbidities that may affect renal function. This study represents a cross-sectional cohort of otherwise healthy individuals in the outpatient setting who engaged in routine physical activity, implying a relatively healthy population. This pooling may limit generalizability to the general population, which would include more comorbidities that could affect renal function. With the inclusion of patients with metabolic derangements such as hypertension, diabetes, and thyroid disease, it is important to keep in mind that these comorbidities can have a profound impact of patient Cr and Cys C levels. Because of this, the collection of thyroid function, hemoglobin A1c, and testosterone levels is desired in future studies. Future study can conduct research on patients who do not have pre-existing metabolic derangements, but the nature of the patient population in a urologic clinical setting is prone to bring concomitant medical complexity and this study aimed to adequately represent a typical patient base. Finally, future studies can also stratify the patient population based on their pre-existing comorbidities and can be analyzed within those groups separately. Ultimately, future research should focus on analyzing the accessibility of testing cys C across different institutions, expanding the study to larger, more geographically diverse patient populations and prospective study of renal outcomes.

We present data regarding our patient population of elite athletes undergoing TTh. Cys C measurements display less variance compared with Cr at differing BMI distinctions. At the upper limit of BMI or PBF in our patient population, cys C exhibits minimal to moderate variability compared with Cr. Findings in this specific population of men correlate with previously published data on broader patient populations.