Journal: bioRxiv

Article Title: Epigenetic Regulation of Inflammation by Dopamine in Primary Human Macrophages

doi: 10.64898/2026.01.21.700899

Figure Lengend Snippet: Primary human monocyte-derived macrophages (hMDM) from sixteen donors were treated with dopamine (10 -6 M) for 3 hours with or without pre-treatment using hypomethylating agents 5-aza-2’-deoxycytidine (dAZA; 10 -6 M), 5-azacytidine (AZA; 10 -6 M), or vehicle control. RNA and DNA were isolated for analysis of IL-1β gene expression by qPCR and IL-1β DNA methylation, respectively. (A) Dopamine increased IL-1β mRNA expression, an effect inhibited by dAZA (n= 7-8 donors, *p<0.05). (B) AZA pretreatment produced a similar inhibitory effect on dopamine-induced IL-1β expression, trending toward statistical significance (n=5-6, p=0.0513). (C) Dopamine treatment significantly increased methylated IL-1β levels compared to vehicle control (n=16, p<0.05). (D) Lipopolysaccharide (LPS) increased IL-1β DNA methylation; however, this effect did not reach statistical significance in this donor cohort (n=16). (E) LPS-induced changes in IL-1β DNA methylation were significantly correlated with dopamine-induced changes within the same donors, indicating consistent donor-specific responsiveness across stimuli (n=16, **p<0.01).

Article Snippet: Both high-methylated and low-methylated human genomic DNA (80-8061-HGHM5 and 80-8062-HGUM5 from EpigenDx) were bisulfite treated and used as controls for the primer sets.

Techniques: Derivative Assay, Control, Isolation, Gene Expression, DNA Methylation Assay, Expressing, Produced, Methylation

Journal: Frontiers in Physiology

Article Title: Loss-of-function mitochondrial DNA polymerase gamma variants cause vascular smooth muscle cells to secrete a diffusible mitogenic factor

doi: 10.3389/fphys.2024.1488248

Figure Lengend Snippet: AlphaFold modeling of POLG variants and impact on the POLG/POLG2/dsDNA complex. (A) Crystal structure PDB8g5j of the human POLG holoenzyme complexed with DNA. (B) Zoomed view of the catalytic domain and residues of interest (magenta). Blues: POLG2s, gray: POLG, orange: POLG exonuclease domain, green: polymerase domain. (C) AlphaFold-predicted structure of human POLG (NP_001119603) complexed with two POLG2 subunits (NP_009146) and a short sequence of DNA. Residues shown as spheres were presumably too mobile to be resolved in the crystal structure. Insets of POLG and POLG2s illustrate model confidence (blue low; red high). (D–G) Catalytic domains modeled with POLG variants. Wild-type is shown as partially transparent. WT helices between Q976 and L1083 are colored green-olive–orange–red-brick. Residues associated with PEO are shown in hot pink. The variant in each panel is shown in red, with the WT residue in magenta.

Article Snippet: We introduced variants into the human POLG (NM_002693) coding sequence on a pCMV6 backbone (RC204456, OriGene) using the Q5 ® Site-Directed Mutagenesis Kit (E0552S, New England Biolabs (NEB)).

Techniques: Sequencing, Variant Assay, Residue