A Nationwide Survey Study of Recovery Community Centers Supporting People in Recovery From Substance Use Disorder.

OBJECTIVE: The medical community has become aware of its role in contributing to the opioid epidemic and must be part of its resolution. Recovery community centers (RCCs) represent a new underused component of recovery support. METHODS: This study performed an online national survey of all RCCs identified in the United States, and used US Census ZIP code tabulation area data to describe the communities they serve. RESULTS: Residents of areas with RCCs were more likely to be Black (16.5% vs 12.6% nationally, P = 0.005) and less likely to be Asian (4.7% vs 5.7%, P = 0.005), American Indian, or Alaskan Native (0.6% vs 0.8%, P = 0.03), or live rurally (8.5% vs 14.0%, P < 0.0001). More than half of RCCs began operations within the past 5 years. Recovery community centers were operated, on average, by 8.8 paid and 10.2 volunteer staff; each RCC served a median of 125 individuals per month (4-1,500). Recovery community centers successfully engaged racial/ethnic minority groups (20.8% Hispanic, 22.5% Black) and young adults (23.5% younger than 25 years). Recovery community centers provide addiction-specific support (eg, mutual help, recovery coaching) and assistance with basic needs, social services, technology access, and health behaviors. Regarding medications for opioid use disorder (MOUDs), RCC staff engaged members in conversations about MOUDs (85.2%) and provided direct support for taking MOUD (77.0%). One third (36.1%) of RCCs reported seeking closer collaboration with prescribers. CONCLUSIONS: Recovery community centers are welcoming environments for people who take MOUDs. Closer collaboration between the medical community and community-based peer-led RCCs may lead to significantly improved reach of efforts to end the opioid epidemic.

ARHGDIA Confers Selective Advantage to Dissociated Human Pluripotent Stem Cells.

Human pluripotent stem cells (hPSCs) have generated significant interest in the scientific community based on their potential applications in regenerative medicine. However, numerous research groups have reported a propensity for genomic alterations during hPSC culture that poses concerns for basic research and clinical applications. Work from our laboratory and others has demonstrated that amplification of chromosomal regions is correlated with increased gene expression. To date, the phenotypic association of common genomic alterations remains unclear and is a cause for concern during clinical use. In this study, we focus on trisomy 17 and a list of candidate genes with increased gene expression to hypothesize that overexpressing 17q25 located ARHGDIA will confer selective advantage to hPSCs. HPSC lines overexpressing ARHGDIA exhibited culture dominance in co-cultures of overexpression lines with nonoverexpression lines. Furthermore, during low-density seeding, we demonstrate increased clonality of our ARHGDIA lines against matched controls. A striking observation is that we could reduce this selective advantage by varying the hPSC culture conditions with the addition of ROCK inhibitor (ROCKi). This work is unique in (1) demonstrating a novel gene that confers selective advantage to hPSCs when overexpressed and may help explain a common trisomy dominance, (2) providing a selection model for studying culture conditions that reduce the appearance of genomically altered hPSCs, and (3) aiding in elucidation of a mechanism that may act as a molecular switch during culture adaptation.

Characterization of human fibroblast-derived extracellular matrix components for human pluripotent stem cell propagation.

Recent studies from our laboratory have shown that acellular substrates generated from human fibroblasts successfully maintained human pluripotent stem cells (hPSCs) in their undifferentiated state for extended periods. Aiming at better characterization, we conducted proteomic analyses to identify the extracellular matrix (ECM) proteins in mouse embryonic- and two human fibroblast-derived acellular substrates. Our studies identified heparan sulfate proteoglycan (HSPG) as a core component of these substrates and immunocytochemical analyses confirmed the presence of HSPG as well as other ECM proteins identified through proteomic analyses. In our attempt to develop surfaces that mimic fibroblast-deposited ECM and their self-renewal capabilities, substrates comprising HSPG and other core ECM proteins were formulated and assessed for the function of hPSC self-renewal. WA09 and BG01v hPSCs maintained on these substrates exhibit multiple characteristics of pluripotency, including (i) tight colony formation with typical stem cell morphology; (ii) positive expression of alkaline phosphatase, (iii) positive expression of SSEA3, SSEA4 and Oct4 based on immunocytochemical analyses; (iv) POU5F1, NANOG and SOX2 mRNA expression; and (v) in vitro differentiation and expression of germ-layer-specific markers. Our studies also reveal that although HSPG by itself-does not support hPSC self-renewal, a substrate that combines HSPG and fibronectin is sufficient for undifferentiated propagation of hPSCs. These studies form the basis for identification of appropriate ECM components in a substrate that synergistically promotes activation of adhesion and signaling pathways responsible for hPSC self-renewal.

Immunocytochemical analysis of human pluripotent stem cells using a self-made cytospin apparatus.

Human pluripotent stem cells (hPSCs) that include human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) are exciting cell sources due to their limitless self-renewal capabilities and their potential to differentiate into multiple cell types. The pluripotent state of hPSCs is typically assessed by techniques such as qPCR, immunocytochemistry, and by other in vitro and in vivo differentiation strategies into multiple cell types. Among these, immunocytochemical techniques have been developed for routine characterization of the undifferentiated state of hPSCs based on analysis of candidate intracellular and cell-surface biomarkers. Given the fact that hPSCs grow as colonies, problems arise in quantifying the expression of these markers at the individual cell level on a routine basis. Flow cytometry analyses serve to address this issue but require cell numbers and use of reagents that are not normally conducive for routine quality control assessment of hPSC cultures. Thus, the development of practical and reproducible means of creating monolayer cell samples with preserved integrity for marker evaluation has many advantages in stem cell research. This greatly benefits immunocytochemical analysis because individual cells from the monolayer can be easily observed and quantified for the expression of specific markers. Towards this goal, a self-made cytospin apparatus was constructed and optimized for use with immunocytochemical staining. Two cell-surface markers (SSEA3/SSEA4) expression were analyzed in a variant BG01 stem cell line for the purpose of this protocol.