The emergence of inflammatory microglia during gut inflammation is not affected by FFAR2 expression in intestinal epithelial cells or peripheral myeloid cells.

Gut inflammation can trigger neuroinflammation and is linked to mood disorders. Microbiota-derived short-chain fatty acids (SCFAs) can modulate microglia, yet the mechanism remains elusive. Since microglia do not express free-fatty acid receptor (FFAR)2, but intestinal epithelial cells (IEC) and peripheral myeloid cells do, we hypothesized that SCFA-mediated FFAR2 activation within the gut or peripheral myeloid cells may impact microglia inflammation. To test this hypothesis, we developed a tamoxifen-inducible conditional knockout mouse model targeting FFAR2 exclusively on IEC and induced intestinal inflammation with dextran sodium sulfate (DSS), a well-established colitis model. Given FFAR2's high expression in myeloid cells, we also investigated its role by selectively deleting it in these populations of cells. In an initial study, male and female wild-type mice received 0 or 2% DSS for 5d and microglia were isolated 3d later to assess inflammatory status. DSS induced intestinal inflammation and upregulated inflammatory gene expression in microglia, indicating inflammatory signaling via the gut-brain axis. Despite the lack of significant effects of sex in the intestinal phenotype, male mice showed higher microglial inflammatory response than females. Subsequent studies using FFAR2 knockout models revealed that FFAR2 expression in IECs or immune myeloid cells did not affect DSS-induced colonic pathology (i.e. clinical and histological scores and colon length), or colonic expression of inflammatory genes. However, FFAR2 knockout led to an upregulation of several microglial inflammatory genes in control mice and downregulation in DSS-treated mice, suggesting that FFAR2 may constrain neuroinflammatory gene expression under healthy homeostatic conditions but may permit it during intestinal inflammation. No interactions with sex were observed, suggesting sex does not play a role on FFAR2 potential function in gut-brain communication in the context of colitis. To evaluate the role of FFAR2 activated by microbiota-derived SCFAs, we employed the same knockout and DSS models adding fermentable dietary fiber (0 or 2.5% inulin for 8 wks). Despite no genotype or fiber main effects, contrary to our hypothesis, inulin feeding augmented DSS-induced inflammation and signs of colitis, suggesting context-dependent effects of fiber. These findings highlight microglial involvement in colitis-associated neuroinflammation and advance our understanding of FFAR2's role in the gut-brain axis. Although not integral, we observed that the role of FFAR2 differs between homeostatic and inflammatory conditions, underscoring the need to consider different inflammatory conditions and disease contexts when investigating the role of FFAR2 and SCFAs in the gut-brain axis.

PEGylated nanoparticles interact with macrophages independently of immune response factors and trigger a non-phagocytic, low-inflammatory response.

Poly-ethylene-glycol (PEG)-based nanoparticles (NPs) - including cylindrical micelles (CNPs), spherical micelles (SNPs), and PEGylated liposomes (PLs) - are hypothesized to be cleared in vivo by opsonization followed by liver macrophage phagocytosis. This hypothesis has been used to explain the rapid and significant localization of NPs to the liver after administration into the mammalian vasculature. Here, we show that the opsonization-phagocytosis nexus is not the major factor driving PEG-NP - macrophage interactions. First, mouse and human blood proteins had insignificant affinity for PEG-NPs. Second, PEG-NPs bound macrophages in the absence of serum proteins. Third, lipoproteins blocked PEG-NP binding to macrophages. Because of these findings, we tested the postulate that PEG-NPs bind (apo)lipoprotein receptors. Indeed, PEG-NPs triggered an in vitro macrophage transcription program that was similar to that triggered by lipoproteins and different from that triggered by lipopolysaccharide (LPS) and group A Streptococcus. Unlike LPS and pathogens, PLs did not increase transcripts involved in phagocytosis or inflammation. High-density lipoprotein (HDL) and SNPs triggered remarkably similar mouse bone-marrow-derived macrophage transcription programs. Unlike opsonized pathogens, CNPs, SNPs, and PLs lowered macrophage autophagosome levels and either reduced or did not increase the secretion of key macrophage pro-inflammatory cytokines and chemokines. Thus, the sequential opsonization and phagocytosis process is likely a minor aspect of PEG-NP - macrophage interactions. Instead, PEG-NP interactions with (apo)lipoprotein and scavenger receptors appear to be a strong driving force for PEG-NP - macrophage binding, entry, and downstream effects. We hypothesize that the high presence of these receptors on liver macrophages and on liver sinusoidal endothelial cells is the reason PEG-NPs localize rapidly and strongly to the liver.

Characterizing the amount and variability of intramuscular fat deposition throughout pork loins using barrows and gilts from two sire lines.

The objective was to determine the amount and variability of intramuscular fat (IMF) in a pork loin attributable to anatomical chop location, sex, and sire line. Pigs were sired by commercially available terminal Duroc boars selected for meat quality (MQ; n = 96) or lean growth (LG; n = 96) and equally split between barrows and gilts. After slaughter and fabrication, bone-in chops were removed from four locations of each left-side loin (A = 6th rib, B = 10th rib, C = last rib, and D = 4th lumbar vertebrae). An adjacent pair of chops from each location was collected and evaluated for visual color and marbling, subjective firmness, moisture and extractable lipid (IMF) (anterior chop), and Warner-Bratzler shear force (posterior chop). Data were analyzed using the MIXED procedure of SAS as a split-plot design. Homogeneity of variances was tested on raw data using Levene's test of the GLM procedure and found to be heterogeneous. Thus, a two-variance model was fit using the REPEATED statement of the MIXED procedure, grouped by pig. The mivque(0) option of the VARCOMP procedure was used to calculate the proportion of variability that each factor contributed to the total variance. Barrows (3.64%) produced chops with greater (P < 0.01) IMF content than gilts (3.20%), and barrows (2.14) had greater (P < 0.01) IMF variability than gilts (1.23). Chops from MQ pigs (4.02%) exhibited greater (P < 0.01) IMF content than LG (2.82%), and MQ (1.76) had greater IMF variability (P < 0.01) than LG pigs (0.97). Chops from locations A (3.80%) and D (3.77%) had greater IMF than B (3.34%; P < 0.01), and A, B, and D had greater IMF than C (2.77%; P < 0.01). Variances of IMF also differed (A = 1.44, B = 1.59, C = 1.05, and D = 2.18; P = 0.01) across chop locations. Of the variability in IMF, 33.0% was attributed to sire line, 10.16% to chop location, and 4.01% to sex, with 52.83% not accounted for by these three factors. Location A chops were the most (P < 0.01) tender (2.57 kg) and C chops the least (P < 0.01) tender (2.93 kg), while B and D chops were intermediate and not different from each other. No differences in variability (P = 0.40) of tenderness were observed among chop locations (A = 0.31, kg B = 0.24 kg, C = 0.24 kg, and D = 0.23 kg). These results demonstrated that variability in tenderness values did not reflect the variability of IMF. In conclusion, chop location, sex, and sire line all contribute to the amount and variability of pork loin marbling.