A retrospective analysis was conducted on patients with cholangiocarcinoma (CCA) who underwent radical surgical excision at the Affiliated Hospital of Qingdao University between January 2017 and January 2022—the follow-up period extended until December 31, 2022. The selection criteria included: 1) Patients who underwent radical resection and were confirmed to have CCA through pathology after surgery; 2) Patients who underwent preoperative imaging with computed tomography (CT) that included imaging of the third lumbar vertebra level of the spine, with images suitable for body composition analysis; 3) Patients who had a complete set of clinicopathological and serological data collected within a month before surgery, including creatinine and cystatin C levels; 4) Patients with access to follow-up data and informed consent. Patients who received local and systemic adjuvant chemoradiotherapy before radical resection, those with concurrent tumors, and those unable to complete all follow-up assessments were excluded. The Ethics Committee of the Affiliated Hospital of Qingdao University approved this study, waiving the requirement for informed consent due to the retrospective study design and analysis of existing clinical data. (QYFYWZLL27646)

Extensive data was collected from the electronic medical record system of the Affiliated Hospital of Qingdao University. Patient demographic and serological data were obtained and recorded a month before surgery. The demographic data included age, sex, weight, and height. Serological data included assessments of renal function, cystatin C and gamma-glutamyl transferase levels, and the measurement of liver function markers such as alanine transaminase (ALT), aspartate transaminase (AST), albumin (ALB), globulin (GLB), total bilirubin (TBIL), and direct bilirubin (DBIL).

The details of the surgery and pathology were collected, including the location, diameter, and degree of differentiation of the tumor. The Creatinine/Cystatin C ratio (CCR) was calculated using the following formula: CCR = [creatinine (mg/dL)/cystatin C (mg/L)] × 10 (14).

The primary focus of the study was to measure the overall survival (OS) of the participants. The duration between the surgery date and the date of death or the end of the follow-up period was considered for calculating OS. The secondary endpoint was recurrence-free survival (RFS), defined as the period from the surgery date to the first recurrence or death. Recurrence was determined by either imaging-based or pathological validation of a tumor.

CT imaging data of patients collected within a month before surgery were analyzed using the sliceOmatic^® (v5.0; TomoVision, Canada) software. The software was used to quantify the skeletal muscle and adipose tissue content at the third lumbar vertebra level to evaluate sarcopenia and myosteatosis according to previously established parameters (15). Different specific Hounsfield unit (HU) thresholds were assigned for various tissues: -29 to +150 HU for skeletal muscle, -150 to -50 HU for visceral adipose tissue, and -190 to -30 HU for subcutaneous and intramuscular adipose tissue (16).

Each tissue type at the L3 level was evaluated in every CT image and marked with different colors ( Figure 1 ). Sarcopenia was assessed through the skeletal muscle index (SMI), calculated as [SMA/height squared (m²)], while myosteatosis was assessed by the average SMA degeneration (17). The myosteatosis thresholds were set at 41 HU for patients with a body mass index (BMI) < 25 kg/m² and 33 HU for patients with a BMI ≥ 25 kg/m² (18). Sarcopenia was defined as an SMI of < 41 cm²/m² for women and < 43 cm²/m² for men with a BMI < 25 and < 53 cm²/m² for men with a BMI ≥ 25 (19).

All measurements were carried out by a pair of researchers who analyzed the CT images of all patients blindly and with great attention to detail.

Using specialized software, measurement data, normally distributed data, and data with homogeneous variance underwent statistical testing. Non-parametric tests were employed for alternative data types. Depending on the type of data (parametric and non-parametric), the data are represented as either the mean ± standard deviation (SD) or the median (P25–P75). Fractional variables are represented as frequency and percentage.

Two-sample t-tests or Wilcoxon Mann-Whitney (U) tests were employed to compare numerical data. In contrast, categorical variables between the two groups were compared using Pearson Chi-square tests or Fisher exact tests. A receiver operating characteristic (ROC) curve of the CCR was drafted to calculate the critical value for predicting the survival status of patients with CCA.

Based on this cut-off value, all patients were grouped into either a low CCR group or a high CCR group. Kaplan-Meier curve analysis was conducted to evaluate OS and RFS, and the log-rank test was used to examine the differences between curves. Moreover, univariate and multivariate Cox regression analyses were conducted for OS and RFS, respectively, to analyze the prognostic value of the variables for OS and RFS based on the hazard ratio (HR) and 95% confidence interval (CI 95%).

All statistical analyses were conducted using the Statistical Package for the Social Sciences (SPSS) (version 26.0; IBM, Armonk, NY, USA). A two-sided P-value of less than 0.05 was considered statistically significant.

Among the 757 patients pathologically diagnosed with CCA, 608 were excluded based on specific criteria. The study focused on 149 patients diagnosed with cholangiocarcinoma (CCA). They were separated into two groups - one with myosteatosis and one without. The research flow chart is shown in Figure 2 .

The study classified the patients according to anatomic location, resulting in 80 intrahepatic cholangiocarcinomas and 69 extrahepatic cholangiocarcinomas. Patients with extrahepatic cholangiocarcinoma had significantly shorter OS and RFS than those with intrahepatic cholangiocarcinoma ( Supplementary Figure S1 ).

The study identified different factors that affect the survival rate of patients with cholangiocarcinoma. One of the factors is myosteatosis, which is when fat accumulates in muscle tissue. Patients with myosteatosis have almost twice the chance of not surviving compared to those without it. Another factor is the creatinine/cystatin C ratio (CCR). If the ratio is less than or equal to 10.834, it may lead to a lower survival rate. Lymph node involvement is also a significant factor for worse survival, while low albumin levels show poor nutritional status. Higher CA-125 levels may indicate cancer progression, while elevated white blood cell (WBC), neutrophil, and monocyte counts show systemic inflammation that may impact survival. Lower hemoglobin levels may indicate anemia from chronic disease, and higher levels of total bilirubin, direct bilirubin, indirect bilirubin, alkaline phosphatase, and uric acid may indicate impaired liver function and metabolic disturbances. Among all the variables, only lymph node metastasis showed a significant correlation with OS (Hazard Ratio [HR] = 3.126, P < 0.0001). Patients with lymph node metastasis had a 3.1 times higher chance of having a worse prognosis ( Table 2 ).

However, age, body mass index (BMI), nerve invasion, vascular invasion, skeletal muscle area (SMA), subcutaneous adipose tissue (SAT), visceral adipose tissue (VAT), intermuscular adipose tissue (IMAT), visceral to subcutaneous fat ratio (VSR), and sarcopenia did not affect the overall survival (OS) of individuals with cholangiocarcinoma ( Supplementary Table S1 ).

The research showed that certain factors were linked to cholangiocarcinoma recurrence in patients. These included the presence of myosteatosis, a CCR score of 10.834 or less, and various laboratory parameters such as alkaline phosphatase, hemoglobin, and cancer antigen 125. Only three factors independently influenced patients’ recurrence-free survival (RFS): lymph node metastasis, vascular invasion, and cancer antigen 125 levels. These three factors significantly predict recurrence or death, even when accounting for other variables ( Table 3 ).

Other factors, such as age, body mass index, nerve invasion, lymphocyte count, indirect bilirubin levels, and sarcopenia, were not found to have a significant impact ( Supplementary Table S2 ).

The study found that patients with myosteatosis had significantly shorter median survival than those without myosteatosis (33.4 months vs. 59.9 months, P = 0.022). It also found that myosteatosis was associated with a shorter median RFS than non-myosteatosis. This suggests that the presence of myosteatosis will increase the risk of disease recurrence or death in cholangiocarcinoma patients ( Figures 4A, B ).

The study found that if the ratio is less than or equal to 10.834, it may lead to a lower survival rate. Individuals with a creatinine/cystatin C ratio (CCR) of ≤ 10.834 had significantly shorter median survival than those with a CCR > 10.834 (37.2 months vs. 57.9 months, P = 0.038). Similarly, the study also found that a lower CCR score was associated with a shorter median RFS than a higher CCR score. This suggests that the presence of a lower CCR score increases the risk of disease recurrence or death in cholangiocarcinoma patients ( Figures 4C, D ).

This study found that patients with cholangiocarcinoma may have a worse prognosis if they have both myosteatosis and a CCR score of 10.834 or less. Patients with these two factors had the lowest overall and recurrence-free survival rates. Patients without myosteatosis and a CCR score of more than 10.834 had the highest survival rate. In contrast, those with both myosteatosis and a CCR score of 10.834 or less had the lowest survival rate. The difference in overall survival between these two groups was 31.8 months. The median recurrence-free survival for the group with both myosteatosis and a CCR score of 10.834 or less was 20.8 months, significantly less than the other groups. These findings suggest that combining myosteatosis and a CCR score of 10.834 or less may help predict the prognosis of patients with cholangiocarcinoma ( Figures 4E, F ).