A hydrogel microcapsule with an intermediate thin oil layer is presented to achieve smart release of a broad range of cargoes triggered via diverse stimuli. A microfluidic technique is used to ...produce triple emulsion droplets with a thin oil layer that separates the innermost aqueous phase from the hydrogel prepolymer phase, which transforms into a hydrogel shell via photopolymerization. The intermediate oil layer within the hydrogel microcapsule acts as an effective diffusion barrier, allowing encapsulation of various small cargoes within a porous hydrogel shell until a stimulus is applied to destabilize the oil layer. It is demonstrated that diverse stimuli including chemical dissolution, mechanical stress, and osmotic pressure can be utilized to release the encapsulated cargo on‐demand. In addition, osmotic pressure and the hydrogel shell thickness can be independently tuned to control the onset time of release as well as the release behavior of multi‐cargo encapsulated hydrogel microcapsule. The release can be either simultaneous or selective.
A hydrogel microcapsule with an intermediate thin oil layer enables smart release of a broad range of cargoes via diverse stimuli. The oil layer in the microcapsules acts as an effective diffusion barrier until only when the oil layer is destabilized. The osmotic pressure and the overall stiffness of the microcapsule can be fine‐tuned to control the release behavior of the multi‐cargo encapsulated microcapsule.
Cas12a (also called Cpf1) is a representative type V-A CRISPR effector RNA-guided DNA endonuclease, which provides an alternative to type II CRISPR-Cas9 for genome editing. Previous studies have ...revealed that Cas12a has unique features distinct from Cas9, but the detailed mechanisms of target searching and DNA cleavage by Cas12a are still unclear. Here, we directly observe this entire process by using single-molecule fluorescence assays to study Cas12a from Acidaminococcus sp. (AsCas12a). We determine that AsCas12a ribonucleoproteins search for their on-target site by a one-dimensional diffusion along elongated DNA molecules and induce cleavage in the two DNA strands in a well-defined order, beginning with the non-target strand. Furthermore, the protospacer-adjacent motif (PAM) for AsCas12a makes only a limited contribution of DNA unwinding during R-loop formation and shows a negligible role in the process of DNA cleavage, in contrast to the Cas9 PAM.
Invariant natural killer T (iNKT), mucosal-associated invariant T (MAIT), and γδ T cells are innate T cells that acquire memory phenotype in the thymus and share similar biological characteristics. ...However, how their effector differentiation is developmentally regulated is still unclear. Here, we identify analogous effector subsets of these three innate T cell types in the thymus that share transcriptional profiles. Using single-cell RNA sequencing, we show that iNKT, MAIT and γδ T cells mature via shared, branched differentiation rather than linear maturation or TCR-mediated instruction. Simultaneous TCR clonotyping analysis reveals that thymic maturation of all three types is accompanied by clonal selection and expansion. Analyses of mice deficient of TBET, GATA3 or RORγt and additional in vivo experiments corroborate the predicted differentiation paths, while human innate T cells from liver samples display similar features. Collectively, our data indicate that innate T cells share effector differentiation processes in the thymus.
Blending two or more ingredients to create a better product that possesses the characteristics of all its ingredients has been actively studied in the food science and technology field. However, ...there has not been enough research on the blend of multiple vegetable ingredients in the area of high-moisture meat analogue (HMMA) manufacturing. Moreover, there has been no study on the development of HMMAs from a blend of soy protein isolate (SPI) and pea protein isolate (PPI). Thus, the aims of this study were i) to examine whether HMMA prepared by blending SPI and PPI has better characteristics than HMMA produced with only SPI or PPI; ii) to investigate the effect of interactions with SPI and PPI on the quality characteristics of HMMAs, when SPI and PPI were blended in various weight ratios 100:0, 80:20, 60:40, 50:50, 40:60, 20:80, and 0:100 (w/w). The quality characteristics of HMMAs were investigated via specific mechanical energy (SME), mass flow rate (MFR), visual appearance, cutting strength, degree of texturization, and moisture content. HMMAs made from blending SPI and PPI showed better visual and textural characteristics than HMMA made from PPI only, but these properties were still worse than HMMA made from SPI only. However, the phenomenon of weakened cutting strength by PPI addition when preparing SPI-based HMMAs suggested that PPI can be used as a tenderizer (texture modifier) in SPI-based HMMAs. Therefore, SPI and PPI-blended HMMAs suit the diets of the elderly because they require less force to masticate than SPI-only HMMA. In conclusion, our findings demonstrated that blending two or more protein sources can have a positive effect on the quality characteristics of HMMAs.
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•HMMAs made from blending SPI and PPI showed better characteristics than HMMA made from PPI only.•PPI has potential as a tenderizer (texture modifier) in SPI-based HMMAs for the elderly.•Blending two or more protein sources can positively affect the quality characteristics of HMMAs.
Invariant natural killer T cells (iNKT cells) can produce copious amounts of interleukin 4 (IL-4) early during infection. However, indirect evidence suggests they may produce this immunomodulatory ...cytokine in the steady state. Through intracellular staining for transcription factors, we have defined three subsets of iNKT cells (NKT1, NKT2 and NKT17) that produced distinct cytokines; these represented diverse lineages and not developmental stages, as previously thought. These subsets exhibited substantial interstrain variation in numbers. In several mouse strains, including BALB/c, NKT2 cells were abundant and were stimulated by self ligands to produce IL-4. In those strains, steady-state IL-4 conditioned CD8(+) T cells to become 'memory-like', increased serum concentrations of immunoglobulin E (IgE) and caused dendritic cells to produce chemokines. Thus, iNKT cell-derived IL-4 altered immunological properties under normal steady-state conditions.
Tg2576 transgenic mice for Alzheimer's disease (AD) exhibited significant phenotypes for neuropathological constipation, but no research has been conducted on the association of the fecal microbiota ...with dysbiosis. The correlation between fecal microbiota composition and neuropathological constipation in Tg2576 mice was investigated by examining the profile of fecal microbiota and fecal microbiota transplantation (FMT) in 9-10-month-old Tg2576 mice with the AD phenotypes and constipation. Several constipation phenotypes, including stool parameters, colon length, and histopathological structures, were observed prominently in Tg2576 mice compared to the wild-type (WT) mice. The fecal microbiota of Tg2576 mice showed decreases in Bacteroidetes and increases in the Firmicutes and Proteobacteria populations at the phylum level. The FMT study showed that stool parameters, including weight, water content, and morphology, decreased remarkably in the FMT group transplanted with a fecal suspension of Tg2576 mice (TgFMT) compared to the FMT group transplanted with a fecal suspension of WT mice (WFMT). The distribution of myenteric neurons and the interstitial cells of Cajal (ICC), as well as the enteric nervous system (ENS) function, remained lower in the TgFMT group. These results suggest that the neuropathological constipation phenotypes of Tg2576 mice may be tightly linked to the dysbiosis of the fecal microbiota.
Three subsets of invariant natural killer T (iNKT) cells have been identified, NKT1, NKT2, and NKT17, which produce distinct cytokines when stimulated, but little is known about their localization. ...Here, we have defined the anatomic localization and systemic distribution of these subsets and measured their cytokine production. Thymic NKT2 cells that produced interleukin-4 (IL-4) at steady state were located in the medulla and conditioned medullary thymocytes. NKT2 cells were abundant in the mesenteric lymph node (LN) of BALB/c mice and produced IL-4 in the T cell zone that conditioned other lymphocytes. Intravenous injection of α-galactosylceramide activated NKT1 cells with vascular access, but not LN or thymic NKT cells, resulting in systemic interferon-γ and IL-4 production, while oral α-galactosylceramide activated NKT2 cells in the mesenteric LN, resulting in local IL-4 release. These findings indicate that the localization of iNKT cells governs their cytokine response both at steady state and upon activation.
•IL-4 secreting NKT2 cells are localized in the thymic medulla•Splenic NKT1 cells are in red pulp and NKT2 cells in T cell zone•Intravenous injection of αGalCer activates NKT1 cells in spleen and liver•Oral administration of αGalCer stimulates NKT2 cells in mesenteric LN
Little is known about where iNKT cell subsets are localized, and identifying these cells by current methods can be challenging. Hogquist and colleagues used histocytometry to precisely visualize iNKT cell subsets in tissues and find that the differential location of iNKT cell subsets impacts their cytokine response.
Matrix metalloproteinase 1 (MMP-1), a calcium-dependent zinccontaining collagenase, is involved in the initial degradation of native fibrillar collagen. Tissue necrosis factor-alpha (TNFα) is a ...pro-inflammatory cytokine that is rapidly produced by dermal fibroblasts, monocytes/macrophages, and keratinocytes and regulates inflammation and damaged-tissue remodeling. MMP-1 is induced by TNFα and plays a critical role in tissue remodeling and skin aging processes. However, the regulation of the MMP1 gene by TNFα is not fully understood. We aimed to find additional cis-acting elements involved in the regulation of TNFα-induced MMP1 gene transcription in addition to the nuclear factor-kappa B (NF-κB) and activator protein 1 (AP1) sites. Assessments of the 5'-regulatory region of the MMP1 gene, using a series of deletion constructs, revealed the requirement of the early growth response protein 1 (EGR-1)-binding sequence (EBS) in the proximal region for proper transcription by TNFα. Ectopic expression of EGR-1, a zinc-finger transcription factor that binds to G-C rich sequences, stimulated MMP1 promoter activity. The silencing of EGR-1 by RNA interference reduced TNFα-induced MMP-1 expression. EGR-1 directly binds to the proximal region and transactivates the MMP1 gene promoter. Mutation of the EBS within the MMP1 promoter abolished EGR-1-mediated MMP-1 promoter activation. These data suggest that EGR-1 is required for TNFα-induced MMP1 transcriptional activation. In addition, we found that all three MAPKs, ERK1/2, JNK, and p38 kinase, mediate TNFα-induced MMP-1 expression via EGR-1 upregulation. These results suggest that EGR-1 may represent a good target for the development of pharmaceutical agents to reduce inflammation-induced MMP-1 expression. BMB Reports 2020; 53(6): 323-328.
In the present report, three different shapes of chitosan-capped gold nanoparticles (nanospheres, nanostars, and nanorods) were synthesized to investigate the effects of shape on cytotoxicity and ...cellular uptake in cancer cells. Green tea extract was utilized as a reducing agent to reduce gold salts to gold nanospheres. Gold nanostars were prepared using an as-prepared nanosphere solution as a seed solution. Gold nanorods were synthesized using a conventional method. All three types of gold nanoparticles showed their characteristic surface plasmon resonance bands upon UV-visible spectrophotometry. In high-resolution transmission electron microscopy images, lattice structures were clearly observed in all three shapes, confirming the crystalline nature of the nanoparticles. All three colloidal solutions of gold nanoparticles retained colloidal stability in various solutions. To assess cytotoxicity, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed on four cancer cell lines. The cytotoxicity was the highest in nanorods, followed by nanostars and finally nanospheres. The cellular uptake of gold nanoparticles in human hepatocyte carcinoma cells (HepG2) was measured, and the results followed the order nanospheres > nanorods > nanostars. The outcomes of the current study may assist in the shape design of gold nanoparticles for therapeutic applications as drug delivery vehicles in the field of nanomedicine.
The anti-inflammatory and neuroprotective effects of trans-cinnamaldehyde (TCA) were investigated on the inflammatory cells and the dopaminergic degeneration in mice. TCA inhibited the up-regulation ...of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in the lipopolysaccharide (LPS)-induced inflammatory BV2 microglial cells. To investigate the TCA efficacy on the 6-hydroxydopamine (6-OHDA)-induced dopaminergic degeneration in mice, an intracerebroventricular injection of 6-OHDA was given to the mice, and TCA (30 mg/kg) was intraperitoneally administered. At 7 d after the 6-OHDA injection, 6-OHDA led to a severe loss of tyrosine hydroxylase (TH)-positive dopaminergic neurons in the striatum and substantia nigra (SN). On the other hand, TCA dramatically maintained the number of TH-positive dopaminergic neurons in the striatum and SN regions of the 6-OHDA-treated mice, which indicates that TCA is able to inhibit the 6-OHDA-induced reduction of TH expression in the dopaminergic neurons in the striatum and SN regions. TCA also inhibited the induction of iNOS and COX-2 in the 6-OHDA model, similarly as shown in the LPS-induced inflammatory BV2 microglial cells. These results indicate that TCA has a neuroprotective effect on dopaminergic neurons and that this effect may be associated with the inhibition of inflammatory responses. These findings suggest that TCA may be a therapeutic candidate for the prevention of inflammation-mediated neurodegenerative diseases.
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