Introduction
Abnormalities in lipid and lipoprotein metabolism are associated with various diseases. It is well established that elevated levels of the low-density lipoprotein increase risk of cardiovascular diseases (CVD).[1,2] [3–7] [8–15]
However, the magnitude of this influence and whether it is positive or negative in the association of blood lipid on cancer has varied greatly among different cancer types, lipid components, and study populations in previous studies.[11] [13]
Esophageal cancer (EC) is one of the most frequent causes of cancer-related mortality worldwide and results in 508,000 deaths annually.[16] [17,18] [14]
Previously, we have reported the prevalence characteristics of dyslipidemia from a large-scale population-based randomized controlled trial in rural China.[19]
Methods
Ethical approval
Research protocols for the present study were approved by the Institutional Review Board of the Peking University School of Oncology, Beijing, China (No. 2011101110). Informed consent was obtained from all participants.
Parent study
In 2012, we initiated the Endoscopic Screening for Esophageal Cancer in China (ESECC) randomized controlled trial (Trial Registration: ClinicalTrials.gov, NCT01688908; https://clinicaltrials.gov/ct2/results?recrs=&cond=&term=01688908&cntry=&state=&city=&dist [20] [21]
Study design and participants
This study was a nested case-control study based on the ESECC trial. Eligibility criteria included: (1) ESECC cohort members; (2) questionnaire completed; (3) blood samples provided with valid test results for serum lipids. Cases were designated as malignant esophageal lesions (severe dysplasia, carcinoma in situ , or ESCC) identified at an endoscopic examination or at the annual follow-up to May 31, 2018. For each case, ten controls were randomly selected using incidence density sampling with matched gender, age at enrollment (5-year age group), family history of EC, and status of subject acceptance of screening from the ESECC population.
Measurement of serum lipids
A fasting blood sample of ∼5 mL was collected from each participant in a heparin sodium anticoagulant tube at enrollment. These tubes were centrifuged at 1000 r/min for 5 min and the supernatant was sent within 4 h for lipid measurement of TC, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglycerides (TG) to the clinical laboratory of the Hua County People's Hospital. Lipid measurements were conducted using a HITACHI7600 automatic biochemistry analyzer (Hitachi High Technologies Co., Tokyo, Japan) with commercially available reagents (Autobio Diagnostics Co., Ltd., Beijing, China) for analysis of TC, TG, LDL-C, and HDL-C.
Statistical analysis
The Chi-square test and Wilcoxon rank-sum test were used to compare baseline characteristics and risk factors of cases and matched controls. Univariable and Multivariable conditional logistic regression models were applied to evaluate the association of each blood lipid indicator and the risk of malignant esophageal lesions. In the multivariable model, adjustments were made for risk factors for ESCC previously identified in this population including age, gender, family history of EC, BMI, coal stove for heating, fume exposure in the kitchen, fast eating speed, and ingestion of leftovers.[20] vs. low serum lipid levels for this cutoff. A series of increasing stepwise cutoff points (every 10 mg/dL) from the 5% quantile to the 95% quantile in control subjects were then tested, and a curve of ORs together with a different set of cutoff points was generated to assess the effect size and consistency of the association which was detected. We also classified each lipid indicator into five categories by quintiles in control subjects and calculated the risk estimates for each quintile of subjects by setting the lowest quintile as the reference group.
Effect modification in this study was evaluated by adding an interaction term of the factor under assessment (age, gender, family history of EC, BMI, smoking, and alcohol consumption) and serum lipid indicators one at a time in the adjusted multivariable model. The coding form of variables assessed in effect modification analysis can be found in Supplementary Table 1, https://links.lww.com/CM9/A502 P values were detected. The optimal cutoff point, which maximized the heterogeneity of risk estimates between subgroups of individuals, was chosen as the cutoff point resulting in the minimum Akaike information criterion (AIC). Statistical analysis was performed using STATA (Version 13.1; Stata Corp LLC, College Station, USA). All tests were two-sided and P values < 0.05 were considered statistically significant.
Results
A total of 211 cases and 2101 matched controls were enrolled in this study [Supplementary Figure 1, https://links.lww.com/CM9/A502 Z = 2.370, P = 0.018) compared with control subjects. There were no significant differences identified for other ESCC risk factors in the ESECC population [Table 1 ]. The median levels of TC, TG, LDL-C, and HDL-C for cases were 189, 117, 97, and 53 mg/dL, and median levels for controls were 185, 113, 96, and 52 mg/dL, respectively. No statistically significant differences were found for these four lipid indices comparing cases and controls.
Table 1 -
Baseline characteristics and distribution of serum lipid levels among participants enrolled in the current study.
Variables
Controls (n = 2101)
Cases (n = 211)
Z or χ
2 value
P value∗
Age (years)
63 (60–66)
63 (60–66)
−0.052
0.958
Gender
0.0002
0.990
Male
1156 (55.02)
116 (54.98)
Female
945 (44.98)
95 (45.02)
Family history of EC
0.014
0.905
No
1690 (80.44)
169 (80.09)
Yes
411 (19.56)
42 (19.91)
BMI (kg/m2 )
24.80 (22.50–27.39)
24.09 (21.67–26.84)
2.370
0.018
Coal stove for heating
0.424
0.515
No
1256 (59.78)
131 (62.09)
Yes
845 (40.22)
80 (37.91)
Fume exposure in kitchen
0.432
0.511
No
534 (25.42)
58 (27.49)
Yes
1567 (74.58)
153 (72.51)
Fast eating speed
2.099
0.147
No
372 (17.71)
29 (13.74)
Yes
1729 (82.29)
182 (86.26)
Ingestion of leftovers
0.303
0.582
≤1/week
1236 (58.83)
120 (56.87)
>1/week
865 (41.17)
91 (43.13)
TC (mg/dL)†
189 (166–212)
185 (166–211)
0.724
0.469
TG (mg/dL)‡
117 (85–168)
113 (81–163)
1.006
0.314
LDL-C (mg/dL)†
97 (82–114)
96 (79–108)
1.805
0.071
HDL-C (mg/dL)†
53 (45–61)
52 (44–60)
0.948
0.343
Data are presented as mean (quartile) or n (%). ∗ The Chi-square test and rank-sum test were used to compare demographic characteristics and behavioral factors among cases and controls included in this study. † To convert cholesterol to mmol/L, multiply values by 0.0259. ‡ To convert TG to mmol/L, multiply values by 0.0113. BMI: Body mass index; EC: Esophageal cancer; HDL-C: High-density lipoprotein cholesterol; LDL-C: Low-density lipoprotein cholesterol; TC: Total cholesterol; TG: Triglycerides.
Almost no significant associations were found between either the dichotomized or the quintile-classified TC, TG, and HDL-C levels and malignant esophageal lesions in the study population as shown in Figure 1 and Supplementary Figure 2, https://links.lww.com/CM9/A502 Figure 1 ].
Figure 1: ORs and 95% CIs of each serum lipid indicator generated from the dichotomized classification with cutoff shifting algorithms ranging from 5% to 95% quartile∗ . (A) The ORs and 95% CIs of TC and malignant esophageal lesions. (B) The ORs and 95% CIs of TG and malignant esophageal lesions. (C) The ORs and 95% CIs of LDL-C and malignant esophageal lesions. (D) The ORs and 95% CIs of HDL-C and esophageal malignant lesions. ∗ In multivariable models, adjustments were made for risk factors for ESCC previously identified in this population including age, gender, family history of EC, BMI, coal stove for heating, fume exposure in kitchen, fast eating speed, and ingestion of leftovers. BMI: Body mass index; CI: Confidence interval; EC: Esophageal cancer; ESCC: Esophageal squamous cell carcinoma; HDL-C: High-density lipoprotein cholesterol; LDL-C: Low-density lipoprotein cholesterol; ORs: Odds ratios; TC: Total cholesterol; TG: Triglycerides.
Evaluation of associations among subgroups defined by potential effect modification variables showed that a family history of EC was found to modify the association of serum lipids (TC and LDL-C) and esophageal malignancy (Z = 2.41, P interaction = 0.016 for TC; Z = 2.48, P interaction = 0.013 for LDL-C) [Supplementary Table 2, https://links.lww.com/CM9/A502 Figure 2 ]. Results were similar for quintile-based cutoffs, suggesting lipids play varying roles in subjects with or without a family history of EC [Supplementary Figure 3, https://links.lww.com/CM9/A502
Figure 2: ORs and 95% CIs of each serum lipid indicator generated from the dichotomized classification with cutoff shifting algorithms ranging from 5% to 95% quantile, stratified by the status of family history of EC† . (A) The ORs and 95% CIs of TC and malignant esophageal lesion, stratified by status of family history of EC. (B) The ORs and 95% CIs of TG and malignant esophageal lesion, stratified by status of family history of EC. (C) The ORs and 95% CIs of LDL-C and malignant esophageal lesion, stratified by status of family history of EC. (D) The ORs and 95% CIs of HDL-C and malignant esophageal lesions, stratified by status of family history of EC. ∗ Significant interaction P values (<0.05) were achieved under these cutoff points. † In multivariable models, adjustments were made for risk factors for ESCC previously identified in this population including age, gender, family history of EC, BMI, coal stove for heating, fume exposure in kitchen, fast eating speed, and ingestion of leftovers. BMI: Body mass index; CI: Confidence interval; EC: Esophageal cancer; ESCC: Esophageal squamous cell carcinoma; HDL-C: High-density lipoprotein cholesterol; LDL-C: Low-density lipoprotein cholesterol; ORs: Odds ratios; TC: Total cholesterol; TG: Triglycerides.
Using AIC as the selection criteria, the optimal cutoff points for maximizing heterogeneity between subgroups were set as 200, 230, 110, and 40 mg/dL for TC, TG, LDL-C, and HDL-C, respectively [Figure 2 ]. Table 2 demonstrates the association of serum lipid indicators and malignant esophageal lesions based on the selected cutoff values using AIC, stratified according to whether the study subject had a family history of EC or not. For individuals with a family history, high-level group of TC (≥200 mg/dL) and LDL-C (≥110 mg/dL) were accompanied by a significantly increased risk of malignant lesions (TC: ORHigh vs. Low = 2.22, 95% CI: 1.14–4.35; LDL-C: ORHigh vs. Low = 1.93, 95% CI: 1.01–3.65). However, a significantly negative association was observed for participants without a family history (TC: ORHigh vs. Low = 0.69, 95% CI: 0.48–0.98; LDL-C: ORHigh vs. Low = 0.50, 95% CI: 0.34–0.76), and distinct heterogeneity was found between individuals in these two subgroups (TC: Z = 3.11, P interaction = 0.002; LDL-C: Z = 3.62, P interaction < 0.001).
Table 2 -
Independent associations of serum lipids with EC risk in a stratification of family history of EC under the AIC-selected cutoff.
Variables
Participants without a family history of EC (n = 1859)
Participants with a family history of EC (n = 453)
Z value for interaction
P value for interaction
Cases (n )/controls (n )
Crude OR (95% CI)
Adjusted OR (95% CI)∗
Cases (n )/controls (n )
Crude OR (95% CI)
Adjusted OR (95% CI)∗
TC
3.11
0.002
Low (<200 mg/dL)
120/1058
Ref
Ref
17/243
Ref
Ref
High (≥200 mg/dL)
49/632
0.67 (0.47–0.96)
0.69 (0.48–0.98)
25/168
2.20 (1.14–4.26)
2.22 (1.14–4.35)
TG
−1.11
0.266
Low (<230 mg/dL)
146/1525
Ref
Ref
38/364
Ref
Ref
High (≥230 mg/dL)
23/165
1.47 (0.91–2.36)
1.62 (1.00–2.62)
4/47
0.81 (0.28–2.38)
0.83 (0.28–2.45)
LDL-C
3.62
<0.001
Low (<110 mg/dL)
138/1164
Ref
Ref
22/282
Ref
Ref
High (≥110 mg/dL)
31/526
0.50 (0.33–0.74)
0.50 (0.34–0.76)
20/129
1.94 (1.03–3.65)
1.93 (1.01–3.65)
HDL-C
0.04
0.968
Low (<40 mg/dL)
19/144
Ref
Ref
5/38
Ref
Ref
High (≥40 mg/dL)
150/1546
0.74 (0.44–1.22)
0.69 (0.41–1.15)
37/373
0.72 (0.26–2.03)
0.73 (0.26–2.07)
∗ In multivariable models, adjustments were made for risk factors for ESCC previously identified in this population including age, gender, family history of EC, BMI, coal stove for heating, fume exposure in kitchen, fast eating speed, and ingestion of leftovers. AIC: Akaike information criterion; BMI: Body mass index; EC: Esophageal cancer; ESCC: Esophageal squamous cell carcinoma; HDL-C: High-density lipoprotein cholesterol; LDL-C: Low-density lipoprotein cholesterol; OR: Odds ratio; TC: Total cholesterol; TG: Triglycerides.
In addition, we performed a sensitivity analysis by excluding prevalent ESCC cases detected within 12 months of blood lipid measurement, and the effect modification of family history status remained significant [Supplementary Table 3, https://links.lww.com/CM9/A502 https://links.lww.com/CM9/A502
Discussion
The association of serum lipids and cancer has been studied since the 1990s. To date, the associations identified in population-based prospective studies have varied markedly, primarily due to differences in populations, tissue specificity, and the complexity of lipid metabolism itself. Genetic and environmental factors and the potential interactions between them may also contribute to the inconsistencies in these associations, which may bias conclusions. This study is a prospective nested case-control study designed to investigate the association of serum lipids and the risk of malignant esophageal lesions in the squamous epithelium based on a large-scale population-based randomized controlled trial in rural China. We found almost no association between serum lipid levels and malignant esophageal lesions in the study population, which is consistent with a previous study.[14]
Serum lipid abnormalities are correlated with the development of many different diseases. In many cases, the determination of the specific role serum lipid(s) play in these diseases is not straightforward, as serum lipid is regulated by a complex network of other molecules. As such, a traditional pooled analysis that ignores interactions (heterogeneity) may be inappropriate, while stratified epidemiology has the power for the identification of real associations, which may serve to reduce controversial results in the study of serum lipids and cancer. Based on this concept, we suggest that genetic or environmental variables that have the potential for effect modification be subjected to more intensive evaluation in epidemiologic studies investigating the association of serum lipids and cancer.
Considering the design of ESCC screening projects, the interaction of associations detected in stratified analysis in this study may also be useful for the identification of individuals at high risk for ESCC. We previously established a prediction model for ESCC based on questionnaire investigation, and this model can significantly decrease endoscopic screenings by distinguishing a subgroup at high risk for ESCC in the general population.[22] [23] https://links.lww.com/CM9/A502
Two main strengths of this study when compared with previous hospital-based case-control studies should be noted. As this is a nested case-control study based on an ongoing large population-based randomized trial, the study sample is ideally representative and guarantees cases and controls can be compared. Moreover, serum samples were collected and tested before the onset of clinical-stage ESCCs in this study, which reduced the possibility of false positives due to the energy expenditure and metabolic disorder that may be found in advanced stages of cancer.
The biological mechanisms underlying the heterogeneous associations of serum lipids and the risk of malignant esophageal lesions in subjects with and without a family history of EC are thus far unclear. Here, we can propose a possible mechanistic explanation based on the observed heterogeneous effects and the biological significance of the family history of EC, which may imply this study for future genetic research. A family history of cancer is widely accepted as a collective representation of genetic susceptibility.[24] et al 's[25] [26] [26]
Our study has limitations. First, we are unable to conclude the causality of malignant esophageal lesions resulting from dyslipidemia, even though lipid measurement was performed before case identification. In addition, although the current study was based on a large-scale population-based trial with 34,000 participants, only 211 cases of malignant esophageal lesions from a single study area were included in the study, which limits the generalizability of conclusions from this study. Further evaluation in other study populations is needed to validate these results.
We conclude that high levels of TC or LDL-C were associated with an increased risk of malignant esophageal lesions in subjects with a family history of EC but were associated with decreased risk for subjects with no family history of EC. Moreover, these opposing effects yielded no association when subgroups of subjects were pooled together. This finding underlines the necessity of the application of stratified epidemiology in population-based studies evaluating the association of serum lipid and cancer. The mechanism involved in the modification of the association of serum lipids and malignant esophageal lesions with a family history of EC warrants investigation in additional family based genomic studies.
Acknowledgements
The authors would like to thank the Government of Hua County, the Health and Family Planning Commission of Hua County, Henan Province, and all the participants of the ESECC trial. The authors also would like to thank Dr. Michael A. McNutt for the editing and correction of this manuscript.
Funding
This work was supported by grants from the Charity Project of the National Ministry of Health (No. 201202014), the Natural Science Foundation of China (Nos. 82073626 and 81773501), the National Key R&D Program of China (No. 2016YFC0901404), the National Science & Technology Fundamental Resources Investigation Program of China (No. 2019FY101102), the Beijing-Tianjin-Hebei Basic Research Cooperation Project (No. J200016), the Digestive Medical Coordinated Development Center of Beijing Hospitals Authority (No. XXZ0204), the Beijing Natural Science Foundation (No. 7182033), and the Science Foundation of Peking University Cancer Hospital (No. 2020-7).
Conflicts of interest
None.
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