Social Experiences and Demographic Factors in the Regulation of Immune Cells

Before the game, participants will be asked to complete the "Everyday Discrimination Scale" designed to measure their perceptions of discrimination encountered in their everyday lives. This instrument prompts respondents to rate the frequency of discriminatory events in their daily lives, such as "being treated with less courtesy than others." Everyday Discrimination Scale Participants provide their responses on a scale that ranges from "0" for "never" to "5" for "almost every day." The summed responses provide a measure of how frequently they experienced discrimination. The summed score can range from 0 to 50; the higher the score, the more frequently participants perceived that they experienced discrimination. Additionally, participants specify whether these experiences are due to various factors, including race and gender.

The "Experiences of Discrimination Scale" measures the type, timing, and frequency of discrimination. Participants answer "yes" or "no" to 9 types of unfair treatment, such as, "Have you ever been unfairly fired?" The sum of "yes" responses ranges from 0 to 9, with a score above 6 indicating many unfair treatments. Participants also specify if the unfair treatment was due to race, skin color, or other characteristics. The sum of responses related to ancestry, race, ethnicity, or skin color indicates the frequency of racial discrimination. Additionally, participants report the last time they experienced unfair treatment due to race or ethnicity, with responses from 1 ("past week") to 4 ("more than a year"). Lower scores indicate more recent occurrences.

The sum of response scores for the Recent Discrimination (a) and Ethnic Discrimination (b) subscale items of the "General Ethnic Discrimination Scale"provides the frequency of recent discrimination experience and includes a manipulation check for the validity of the recent Cyberball condition Recent Discrimination is calculated as the sum of responses to all (a) items ranging from a choice of 1 for "never" to 6 for "almost all the time. A summed minimum score of 18 corresponds to no lifetime discrimination, and a summed maximum score of 108 corresponds to the most frequent experience of perceived lifetime discrimination. Lifetime Discrimination is calculated as the sum of all (b) item responses ranging from a choice of 1 for "never" to 6 for "almost all the time. A summed minimum score of 18 corresponds to no lifetime discrimination, and a summed maximum score of 108 corresponds to the most frequent experience of perceived lifetime discrimination.

The Cumulative response score from the Stress from Discrimination (c) items on the General Ethnic Discrimination Scale measures a person's perception of the stress of recent and lifetime unfair treatment due to a participant's racial/ethnic identity. Participants are asked to respond to the statement "How stressful was this for you?" for each discrimination experience. Response choices range from 1 for "not at all stressful" to 6 for "extremely stressful." Discrimination Stress is calculated as the sum of all (c) items on the General Ethnic Discrimination (GED) scale, where a summed score of 17 indicates little to no stress, and 102 indicates the highest amount of stress experienced due to recent and lifetime discrimination experiences.

Multiplex enzyme-linked immunosorbent assays (ELISA) will allow assessment of the pg/mL concentration of inflammatory cytokines in saliva and serum. These data will be used to calculate correlation coefficients that depict the magnitude and direction of the relationship between saliva and serum inflammatory cytokine concentration. The cytokines to be measured are Interleukin-1 beta (IL-1β), Interleukin-6 (IL-6), Tumor Necrosis Factor-alpha (TNF-α, and c-Reactive Protein (CRP).

To determine the quantity of each type of peripheral blood mononuclear cells (PBMCs) available after the different Cyberball sequences, the PBMCs will be sorted and quantified from whole blood collected 30 minutes after the first and 30 minutes after the second Cyberball sequence. A Sony SH800 cell sorter is used to perform Fluorescence-Activated Cell Sorting (FACS) with appropriate antibodies to identify T cells (CD3+), B cells (CD19+), NK cells (CD56+), and monocytes (CD14+CD16-, CD14+CD16+) and counts provided of viable cells with the cell counter.

Cellular Cytokine release from (PBMCs) purified from the participant's whole blood is assessed in the presence or absence of stimulation with lipopolysaccharide (LPS) toxin. The LPS incubation time will vary in length, and ELISA will measure pg/ml concentration of IL-6, TNF-α, IL-1β, CRP

RNA expression counts and distribution will be measured in PBMCs purified from participant whole blood collected participants after completing the first and second Cyberball sequences (collection tines separated by up 2 hours)

The Dissimilarity Index or DI measures Race/Ethnic Residential Segregation. We aim to measure the DI between non-Hispanic Black and non-Hispanic White, Hispanic and non-Hispanic White, and Hispanic and non-Hispanic Black populations. Census tract data from the participant's childhood zip code will be used to calculate the three childhood dissimilarity indices. The 2020 census tract data for the current zip code will be used in the equation for calculating this index. The equation for the index is DI=100×0.5× SUM(PxiPx-PyiPy) where Pxi and Pyi are the populations of groups x and y in census tract i, and Px and Py are the total populations of groups x and y in the overall geographic area. The DI ranges from 0 to 100, with scores less than 30 indicating low segregation, 30-60 indicating moderate segregation, and greater than 60 indicating high segregation.

"The Communities That Care Youth Survey" assesses Community risk and protective factors. Calculated average risk scores are based on responses to risk items, and calculated average protection scores are based on the protective items, with minimum and maximum average scores equaling 1 and 4, respectively.

The average community risk is determined by calculating the average responses to items based on the five risk factors: Drug availability, norms for drug use, availability of handguns, neighborhood attachment, and community disorganization. A sample Community Risk item states:"If I wanted to get some marijuana, how easy would it be for me to get some?" response choices range from 1 for "Very hard to 4 for "Very easy." Average scores of less than 2 for the five factors indicate low community risk, and 3 or greater indicates high community risk.

"The Communities That Care Youth Survey" assesses Community risk and protective factors. Calculated average protective factors scores are based on responses to protective items, with minimum and maximum average scores equaling 1 and 4, respectively.

The Average Community Protection is determined by calculating the average responses to items on two protective factors: Opportunities and Rewards for prosocial involvement. A sample Community Protective item states: "My neighborhood offers lots of opportunities to participate in positive activities." With response choices ranging from 1 for "Strongly disagree" to 4 for "Strongly agree" Average scores lower than 2 indicate low community protection and a score of 3 or greater indicates high community protective factors.

The Brief Resilience Scale consists of six items that assess participants' resilience or buffer against stressful experiences. A sample item states, "I tend to bounce back quickly after hard times," with response choices ranging from 1 for "Strongly Disagree" to 5 for "Strongly Agree." The participant's Resilience Score is based on a calculated average of the response scores to the six items on the scale. The minimum possible average score is 1, and the maximum is 6. Scores below 3 indicate low resilience and scores above 4.31 indicate high resilience.

The frequency of recent stress is calculated by summing the response scores on the ten-item "Perceived Stress Scale." The response options for each feeling or thought indicate the frequency with which it occurred, ranging from 0 for "Never" to 4 for "Very Often." Scores can be made by summing across all 10 items. Scores can range from 0 to 40, with higher scores indicating greater stress.

The sum of response items on the "Comorbidity Questionnaire" provides a single measure of the magnitude of chronic medical conditions faced by participants. The questionnaire can have up to fifteen items with twelve listed illnesses and options for participants to list an additional three items. One point if given each is given for the presence of the condition, with an additional point for receiving treatment for the condition, and another if the condition affects daily function.

The Degree of Morbidity is calculated by summing the responses to the three components for each listed illness. The minimum score that can be earned is 0, and the maximum score is 45, with higher scores indicating greater morbidity.

The proportion of methylated to unmethylated cytosines at CpG sites around regulatory transcripts in the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway will be assessed in purified PBMCs