Stress-induced neuroinflammation is widely regarded as one of the primary causes of depression. Gamma-aminobutyric acid (GABA)-enriched foods relieve stress and reduce inflammatory reactions. This study aimed to evaluate whether rice germ with 30% GABA (RG) reduced neuroinflammation in mice exposed to chronic unpredictable mild stress (CUMS). CUMS mice were administered 40, 90, and 140 mg/kg of RG. CUMS increased serum and hypothalamic pro-inflammatory cytokine (TNF-α and IL-6) levels, which were decreased by RG. In the hypothalamus, CUMS elevated M1-type microglia markers of CD86 and NF-κB, whereas RG lowered these levels. The expression levels of NLRP3 inflammasome complex (NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain, and caspase-1), IL-1β, and IL-18 were increased in the hypothalamus of CUMS mice and decreased by RG. RG attenuated depressive-like behaviors in CUMS mice, as measured by the forced swim test and tail suspension test. In conclusion, RG decreased hypothalamic inflammation-related signals, such as TNF-α, IL-6, M1 polarization, NF-κB, NLRP3 inflammasome complex, caspase-1, IL-1β, and IL-18, to diminish depressive-like behavior.

Poly-L-lactic acid (PLLA) fillers correct cutaneous volume loss by stimulating fibroblasts to synthesize collagen and by augmenting the volume. PLLA triggers the macrophage-induced activation of fibroblasts that secrete transforming growth factor-β (TGF-β). However, whether M2 macrophage polarization is involved in PLLA-induced collagen synthesis via fibroblast activation in aged skin is not known. Therefore, we evaluated the effect of PLLA on dermal collagen synthesis via M2 polarization in an H2O2-induced cellular senescence model and aged animal skin. H2O2-treated macrophages had increased expression levels of the M1 marker CD80 and decreased expression levels of the M2 marker CD163, which were reversed by PLLA. The expression levels of interleukin (IL)-4 and IL-13, which mediate M2 polarization, were decreased in H2O2-treated macrophages and increased upon the PLLA treatment. CD163, IL-4, and IL-13 expression levels were decreased in aged skin, but increased after the PLLA treatment. The expression levels of TGF-β, pSMAD2/SMAD2, connective tissue growth factor (CTGF), alpha-smooth muscle actin (α-SMA), collagen type 1A1 (COL1A1), and COL3A1 were also decreased in aged skin, but increased after the PLLA treatment. Moreover, PLLA upregulated phosphatidylinositol 3-kinase p85α (PI3-kinase p85α)/protein kinase B (AKT) signaling, leading to fibroblast proliferation. PLLA decreased the expression of matrix metalloproteinase (MMP) 2 and MMP3, which destroy collagen and elastin fibers in aged skin. The amount of collagen and elastin fibers in aged skin increased following the PLLA treatment. In conclusion, PLLA causes M2 polarization by increasing IL-4 and IL-13 levels and upregulating TGF-β expression and collagen synthesis in aged skin.

Angiogenesis promotes rejuvenation in multiple organs, including the skin. Heat shock protein 90 (HSP90), hypoxia-inducible factor-1 alpha (HIF-1α), and vascular endothelial growth factor (VEGF) are proangiogenic factors that stimulate the activities of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and extracellular signal-regulated kinase 1/2 (ERK1/2). Poly-D,L-lactic acid (PDLLA), polynucleotide (PN), and calcium hydroxyapatite (CaHA) are dermal fillers that stimulate the synthesis of dermal collagen. However, it is not yet known whether these compounds promote angiogenesis, which leads to skin rejuvenation. Here, we evaluated whether PDLLA, PN, and CaHA stimulate angiogenesis and skin rejuvenation using H2O2-treated senescent macrophages and endothelial cells as an in vitro model for skin aging, and we used young and aged C57BL/6 mice as an in vivo model. Angiogenesis was evaluated via endothelial cell migration length, proliferation, and tube formation after conditioned media (CM) from senescent macrophages was treated with PDLLA, PN, or CaHA. Western blot showed decreased expression levels of HSP90, HIF-1α, and VEGF in senescent macrophages, but higher expression levels of these factors were found after treatment with PDLLA, PN, or CaHA. In addition, after exposure to CM from senescent macrophages treated with PDLLA, PN, or CaHA, senescent endothelial cells expressed higher levels of VEGF receptor 2 (VEGFR2), PI3K, phosphorylated AKT (pAKT), and phosphorylated ERK1/2 (pERK1/2) and demonstrated greater capacities for cell migration, cell proliferation, and tube formation. Based on the levels of 4-hydroxy-2-nonenal, the oxidative stress level was lower in the skin of aged mice injected with PDLLA, PN, or CaHA, while the tumor growth factor (TGF)-β1, TGF-β2, and TGF-β3 expression levels; the density of collagen fibers; and the skin elasticity were higher in the skin of aged mice injected with PDLLA, PN, or CaHA. These effects were greater in PDLLA than in PN or CaHA. In conclusion, our results are consistent with the hypothesis that PDLLA stimulates angiogenesis, leading to the rejuvenation of aged skin. Our study is the first to show that PDLLA, PN, or CaHA can result in angiogenesis in the aged skin, possibly by increasing the levels of HSP90, HIF-1α, and VEGF and increasing collagen synthesis.

Poly-D,L-lactic acid (PDLLA) filler corrects soft tissue volume loss by increasing collagen synthesis in the dermis; however, the mechanism is not fully understood. Adipose-derived stem cells (ASCs) are known to attenuate the decrease in fibroblast collagen synthesis that occurs during aging, and nuclear factor (erythroid-derived 2)-like-2 factor (NRF2) increases ASCs survival by inducing M2 macrophage polarization and IL-10 expression. We evaluated the ability of PDLLA to induce collagen synthesis in fibroblasts by modulating macrophages and ASCs in a H2O2-induced cellular senescence model and aged animal skin. PDLLA increased M2 polarization and NRF2 and IL-10 expression in senescence-induced macrophages. Conditioned media from senescent macrophages treated with PDLLA (PDLLA-CMMΦ) reduced senescence and increased proliferation and expression of transforming growth factor-β (TGF-β) and fibroblast growth factor (FGF) 2 in senescence-induced ASCs. Conditioned media from senescent ASCs treated with PDLLA-CMMΦ (PDLLA-CMASCs) increased the expression of collagen 1a1 and collagen 3a1 and reduced the expression of NF-κB and MMP2/3/9 in senescence-induced fibroblasts. Injection of PDLLA in aged animal skin resulted in increased expression of NRF2, IL-10, collagen 1a1, and collagen 3a1 and increased ASCs proliferation in aged animal skin. These results suggest that PDLLA increases collagen synthesis by modulating macrophages to increase NRF2 expression, which stimulates ASCs proliferation and secretion of TGF-β and FGF2. This leads to increased collagen synthesis, which can attenuate aging-induced soft tissue volume loss.

During skin aging, the volume of subcutaneous adipose tissue (sWAT) and the adipogenesis potential of adipose-derived stem cells (ASCs) decrease. It is known that the shortening of cilia length by pro-inflammatory cytokines is related to the decreased adipogenic differentiation of ASCs via increase in Wnt5a/β-catenin. High-intensity focused ultrasound (HIFU) is known to upregulate heat shock proteins (HSP), which decrease levels of pro-inflammatory cytokines. In this study, we evaluated whether HIFU modulates the cilia of ASCs by upregulating HSP70 and decreasing inflammatory cytokines. HIFU was applied at 0.2 J to rat skin, which was harvested at 1, 3, 7, and 28 days. All results for HIFU-applied animals were compared with control animals that were not treated. HIFU increased expression of HSP70 and decreased expression of NF-κB, IL-6, and TNF-α in sWAT. HIFU decreased the expression of cilia disassembly-related factors (AurA and HDAC9) in ASCs. Furthermore, HIFU increased the expression of cilia assembly-related factors (KIF3A and IFT88), decreased that of WNT5A/β-catenin, and increased that of the adipogenesis markers PPARγ and CEBPα in sWAT. HIFU increased the number of adipocytes in the sWAT and the thickness of sWAT. In conclusion, HIFU could selectively increase sWAT levels by modulating the cilia of ASCs and be used for skin rejuvenation.