DOI: 10.5061/dryad.pg4f4qrpt
title: APOE4 is associated with elevated blood lipids and lower levels of innate immune biomarkers in a tropical Amerindian subsistence population

description: In post-industrial settings, APOE4 is associated with increased
 cardiovascular and neurological disease risk. However, the majority of
 human evolutionary history occurred in environments with higher pathogenic
 diversity and low cardiovascular risk. We hypothesize that in
 high-pathogen and energy-limited contexts, the APOE4 allele confers
 benefits by reducing innate inflammation when uninfected, while
 maintaining higher lipid levels that buffer costs of immune activation
 during infection. Among Tsimane forager-farmers of Bolivia (N=1266), APOE4
 is associated with 30% lower C-reactive protein, and higher total
 cholesterol and oxidized-LDL. Blood lipids were either not associated, or
 negatively associated with inflammatory biomarkers, except for
 associations of oxidized-LDL and inflammation which were limited to high
 BMI adults. Further, APOE4 carriers maintain higher levels of total and
 LDL cholesterol at low BMIs. These results suggest the relationship
 between APOE4 and lipids may be beneficial for pathogen-driven immune
 responses, and unlikely to increase cardiovascular risk in an active
 subsistence population.
description: Biomarker data used in this paper were collected by the THLHP
 between 2004 - 2015 (see Gurven et al., 2017; Kraft et al., 2020 for
 details). A Bolivian and Tsimane mobile medical team travel annually or
 biannually among study communities conducting clinical health assessments
 and collecting biochemical and anthropometric information from community
 members that want to participate. This sample includes all data from
 individuals for whom we have <i>APOE</i> genotyping and
 measurements of age, sex, and BMI - which is the base criteria for this
 study. Sample size varies by biomarker and over time for several reasons:
 sampling strategy varies by data type, absent or sick team personnel
 needed to collect data, the number of study villages and thus enrolled
 participants has increased over time, and the data types collected have
 changed over time (see Kraft et al., 2020). There are also fewer repeat
 measurements for a subset of biomarkers (i.e. C-reactive protein and
 oxidized LDL) that were assayed in the U.S., due to them being analyzed as
 part of a prior project. Specific sample sizes are reported in Table 1,
 and full tables report sample size for each model.
 <i>Ethics</i> This research has
 been approved by institutional review boards at the University of New
 Mexico (#07-157) and University
 of California Santa Barbara (#3-20-0740), as well as the Tsimane
 government (Tsimane Gran Consejo) and village leaders. Study participants
 give consent for each part of the research and data collection prior to
 participating, during every visit by the THLHP.
 <i>APOE genotyping </i> Whole
 blood samples were stored in cryovials (Nalgene, USA) and frozen in liquid
 nitrogen before transfer on dry ice to the University of California-Santa
 Barbara, where they were stored at -80°C until genotyping. Single
 nucleotide polymorphism (SNP) genotyping was used to identify
 <i>APOE</i> allelic variants in blood samples. Samples were
 shipped on dry ice to University of Southern California (2010 and 2013)
 and University of Texas-Houston (2016), where DNA was extracted,
 quantified, and haplotype coded for <i>APO- E2</i>,
 <i>E3</i>, and <i>E4</i> alleles using the TaqMan
 Allelic Discrimination system (Thermo-Fisher Scientific, Carlsbad, CA,
 USA). Determination of the <i>APOE2/E3/E4</i> alleles in the
 Tsimane derived from 2 SNPs of 20-30bp oligonucleotides surrounding the
 polymorphic site (Cys112Arg/rs429358 and Cys158Arg/rs7412) (Trumble et
 al., 2017; Vasunilashorn et al., 2011).
 <i>Measurement of blood lipids and immune function
 </i> Biomarkers were either assayed in the field
 at the time of collection, or in the Human Biodemography laboratory at UC
 Santa Barbara in 2016. Blood was collected by
 venipuncture in a heparin-coated vacutainer. Immediately following the
 blood draw, total leukocyte counts and hemoglobin were determined with a
 QBC Autoread Plus dry hematology system (QBC Diagnostics), with a QBC
 calibration check performed daily to verify QBC performance. Relative
 fractions of neutrophils, eosinophils, and lymphocytes were determined
 manually by microscopy with a hemocytometer by a certified Bolivian
 biochemist. ESR was calculated following the Westergren method
 (Westergren, 1957). Serum was separated and frozen in
 liquid nitrogen before transfer to the University of California-Santa
 Barbara where a commercial immunoassay was used to measure oxidized LDL
 (Mercodia, Winston Salem, NC). Serum high sensitivity C-Reactive Protein
 (hs-CRP) was assessed via immunoassay (Brindle et al., 2010), and was
 cross-validated by the University of Washington laboratory, using the
 protocols utilized for the National Health and Nutrition Evaluation Survey
 (NHANES). Oxidized LDL and hs-CRP assays use materials from the same lot
 across all measures. Total and LDL cholesterol levels from serum samples
 were measured (Stat Fax 1908, Awareness Technology, Palm City, FL) in the
 THLHP laboratory in San Borja, Beni, Bolivia.
description: Data is in long-format, as there are numerous repeated
 measurements per individual (pid). Given that there the number of
 observations vary by biomarker and individual, there are also many NAs in
 the data. Mixed effects linear regressions with restricted maximum
 likelihood estimation should used for analyses, with random intercept
 effects for individual ID, i.e. 'pid' (to deal with repeat
 observations). Because Tsimane villages vary in sanitation infrastructure,
 including access to soap and other hygienic products, and potentially
 prevalence by pathogen type (e.g. some living very close to the river
 versus farther out in the forest), individuals may also be clustered by
 community (i.e. community_id') to account for variation in such
 community-level factors. Immune and lipid measures are transformed to
 normalize their skewed distributions. Variables were transformed as
 follows: CRP, BMI, and triglycerides were natural log-transformed; total
 leukocytes and subsets (lymphocytes neutrophils, eosinophils), ESR, and
 remaining cholesterols (total cholesterol, LDL, HDL, ox-LDL) were
 square-root transformed.