柠檬酸钠抗原修复液(50×)

Background: Taohong Siwu decoction (THSWT) has shown therapeutic effects on ischemia/reperfusion injury (IRI). This study tended to investigate the role of THSWT combined with the long non-coding RNA (LncRNA) H19 (H19)/miR-675-5p axis in improving limb IRI (LIRI). Methods: Hind LIRI rats and simulated IRI skeletal myoblasts models were constructed to evaluate the therapeutic effects of THSWT. The mechanism of THSWT treatment on LIRI was investigated by the regulation of the H19/miR-675-5p axis and the wingless/integrated (Wnt)/Ca 2+ signaling pathway. Various assessments, such as H&E staining, TUNEL staining, flow cytometry, cell counting kit-8 (CCK-8) assay, quantitative real-time polymerase chain reaction (qRT-PCR), western blot, immunohistochemistry (IHC) staining, enzyme-linked immunosorbent assay (ELISA), biochemical assay, and calcium fluorescence imaging, were conducted to observe skeletal muscle injury, cell apoptosis, skeletal myoblast proliferation, gene and protein expressions, cytokine levels, glucose (Glu) uptake, and Ca 2+ concentration. Results: THSWT intervention effectively improved skeletal muscle injury in LIRI rats, as evidenced by reduced muscle fiber damage and decreased cell apoptosis, accompanied by downregulation of H19, miR-675-5p, cleaved-Caspase3, Bax, PLC, and PKC expressions and upregulation of Bcl2 expression. Furthermore, silencing of H19 inhibited cell apoptosis of skeletal muscle and reduced IL-1 β , IL-6, and TNF- α levels in LIRI rats. Notably, THSWT intervention combined with the silencing of H19 synergistically promoted the repair of skeletal muscle injury in LIRI rats. Mechanistically, THSWT intervention combined with regulation of the H19/miR-675-5p axis promoted the proliferation of skeletal myoblasts damaged by IRI through the Wnt3a/Ca 2+ signaling pathway, increasing the levels of intracellular Bcl2, while decreasing the levels of Ca 2+ , CaMKⅡ, PLC, PKC, cleaved-Caspase3, Bax, TNF- α , IL-1 β , IL-6, Wnt3a, and β -catenin. Conclusions: THSWT combined with the regulation of the H19/miR-675-5p axis effectively improved LIRI by modulating the Wnt3a/Ca 2+ signaling pathway, providing insights for potential therapeutic strategies for LIRI.

Background Prostate cancer (PCa) is becoming the most common malignancy in men worldwide. We investigated the effect of astragaloside IV combined with PESV on the gut microbiota and metabolite of PCa mice and the process of treating PCa. Methods Nude mice were genetically modified to develop tumors characteristic of PCa. The treatment of PCa mice involved the administration of a combination of astragaloside IV and peptides derived from scorpion venom (PESV). Feces were collected for both 16 S rDNA and metabolic analysis. Fecal supernatant was extracted and used for fecal transplantation in PCa mice. Tumor development was observed in both PCa mice and nude mice. Tumor histopathology was examined, and the expression of inflammatory factors and the AGE-RAGE axis in PCa tissues were analyzed. Results PCa mice treated with Astragaloside IV in combination with PESV showed a significant reduction in tumor volume and weight, and stabilization of gut microbiota and metabolites. At the Genus level, significant differences were observed in Porphyromonas , Corynebacterium , Arthromitus and Blautia , and the differential metabolites were PA16_016_0 , Astragaloside+ , Vitamin A acid , Nardosinone , a-Nortestoster , D-Pantethine , Hypoxanthine , Pregnenolone , cinnamic acid , Pyridoxa , Cirtruline and Xanthurenate . There was a correlation between gut microbiota and metabolites. After the fecal transplantation, tumor growth was effectively suppressed in the PCa mice. Notably, both the mRNA and protein levels of the receptor for advanced glycation end products (RAGE) were significantly decreased. Furthermore, the expression of inflammatory factors, namely NF-κB, TNF-α, and IL-6, in the tumor tissues was significantly attenuated. Conversely, upregulation of RAGE led to increased inflammation and reversed tumor growth in the mice. Conclusion Astragaloside IV combined with PESV could treat PCa by intervening in gut microbiota composition and metabolite by targeting RAGE.

SYCP1 and SYCP3 are essential testis-specific genes for centromere pairing during meiosis, as well as for spermatogenesis and fertility in male germ cells. However, it is still unclear regarding the expression patterns in the fertile reciprocal hybrid offspring of Megalobrama amblycephala (blunt snout bream, BSB, 2n = 48) × Culter alburnus (topmouth culter, TC, 2n = 48). This research elucidated the genetic and expression characteristics of SYCP1 and SYCP3 through molecular cloning, sequence alignment, Western blotting, and immunohistochemistry to assess their roles in both hybrids and parents. The findings revealed that SYCP1 and SYCP3 exhibited high homology between M. amblycephala and C. alburnus , with varying degrees of chimerism in the BT and TB hybrids. The expression level of SYCP1 in these hybrids was intermediate between parents, while SYCP3 was more similar to M. amblycephala and significantly different from C. alburnus ( p < 0.05). Western blotting confirmed the normal expression of both SYCP1 and SYCP3 proteins in the hybrid offspring. Immunohistochemistry verified the significant presence of these proteins in the testes of mature hybrids. These findings suggested that BT and TB hybrids retained the stability of the SYCP1 and SYCP3 genes inherited from their heterozygous parental origins, supporting independent protein expression despite slight variations in the CDS structure. Our results demonstrate that the normal expression of key meiotic genes plays an important role in overcoming reproductive barriers in distant hybridization, which is of great significance for genetic breeding in fish.

Background: Delayed wound healing, a common problem in patients with diabetes mellitus (DM), is associated with impaired keratinocyte autophagy. Epigallocatechin gallate (EGCG), a catechin, has been proven to promote diabetic wound healing. This study aims to explore the therapeutic mechanism of EGCG on diabetic wound healing. Methods: High glucose (HG)-induced keratinocytes and streptozotocin (STZ)-induced DM rats were prepared and intervened with EGCG to examine its therapeutic effects in in vivo and in vitro settings. The AMPK inhibitor, Compound C, was utilized to determine whether EGCG exerted its therapeutic effects through the AMPK/ULK1 pathway. Results: In vitro, EGCG improved HG-induced autophagy impairment in keratinocytes by increasing LC3II/LC3I, Becline1, and ATG5 levels and decreasing p62 level. Mechanically, EGCG activated the AMPK/ULK1 pathway, thereby promoting keratinocyte autophagy through the phosphorylation of AMPK and ULK1. Notably, EGCG promoted the proliferation, migration, synthesis and release of C-C motif chemokine ligand 2 (CCL2) in HG-treated keratinocytes. Furthermore, EGCG indirectly promoted the activation of fibroblasts, as evidenced by increased alpha-smooth muscle actin (α-SMA) and Collagen I levels. In vivo, EGCG promoted wound healing in DM rats, primarily by reducing inflammatory infiltration and increasing granulation tissue to promote wound epithelialization. Besides, EGCG promoted ATG5, KRT10, KRT14, TGF-β1, Collagen I, and α-SMA expressions in the neonatal epithelial tissues of DM rats. However, the use of Compound C reversed the effects of EGCG. Conclusions: These findings indicated that EGCG restored keratinocyte autophagy to promote diabetic wound healing through the AMPK/ULK1 pathway.

Objective: The use of immune checkpoint inhibitors (ICIs) provides promising strategies for hepatocellular carcinoma (HCC) treatment. This study aimed to explore impact and underlying mechanism of the combination therapy of quercetin and anti-programmed cell death 1 (anti-PD-1) antibody on HCC. Methods: Orthotopically transplanted HCC tumors in mice were treated with quercetin, anti-PD-1 antibody, or a combination of both therapies. Histopathological changes and programmed cell death ligand 1 (PD-L1) expression were characterized by hematoxylin and eosin (H&amp;E) and immunohistochemistry (IHC) staining. The diversity and differences of gut microbiota (GM) were evaluated through 16S rRNA sequencing. Levels of macrophage immunity-related cytokines were quantified by enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (RT-qPCR), and Western blot. Results: Combination therapy reduced necrosis, fibrosis, and PD-L1 expression in liver tissues. Additionally, combination therapy reduced GM imbalance and increased abundance of Firmicutes, Actinobacteria, and Verrucomicrobiota at the phylum level as well as Dubosiella and Akkermansia at the genus level. Combination therapy improved macrophage immunity, raised the expressions of CD8a, CD4, CD11b, interleukin (IL)-10, and interferon (IFN)-γ , and declined the expressions of IL-4, IL-6, toll-like receptor 4 (TLR4), an inhibitor of nuclear factor κBα (IκBα), and the NFκB subunit p65. Upon combination therapy, expressions of M2 macrophage-related genes arginase-1 (Arg-1), IL-10, transforming growth factor-β (TGF-β), and matrix metalloproteinase-9 (MMP-9) were upregulated. Instead, M1 macrophage-related genes IL-6, IL-12a, IL-1β, and tumor necrosis factor-α (TNF-α) were downregulated. Conclusions: Quercetin/anti-PD-1 antibody combination therapy reshaped HCC tumor microenvironment in mice in parallel with regulating the GM and macrophage immunity.

Background: Diabetic nephropathy (DN) is one of the most serious complications of diabetes. Rhein has been reported to be effective in treating DN. This study aimed to investigate the role and mechanism of rhein in the treatment of DN. Methods: High glucose-induced (HG) podocyte injury model and streptozocin-induced (STZ) DN mouse model were constructed and intervened with rhein. Cell viability was detected by Cell Counting Kit-8 (CCK-8) assay. The reactive oxygen species (ROS) level was measured by flow cytometry. The expression of Ras-related C3 botulinum toxin substrate 1 (Rac1), NADPH Oxidase 1 (NOX1), and β-catenin were measured by quantitative real-time PCR (RT-qPCR). The contents of glutathione peroxidase 4 (GPX4), α-smooth muscle actin (α-SMA), Nephrin, and Podocin were characterized by immunofluorescence (IF) staining. Hematoxylin-eosin (HE) staining and Masson staining were employed to observe the renal morphological changes and tubulointerstitial fibrosis. The contents of α-SMA and Nephrin were detected by immunohistochemistry (IHC) staining. The kits were utilized to analyze various biochemical indicators. Results: Rhein inhibited the HG-induced accumulation of ROS, malondialdehyde (MDA), and Fe2+, and the expression of α-SMA, Transferrin Receptor 1 (TFR1), acyl-CoA synthetase long-chain family member 4 (ACSL4), Vimentin, Snail, and Desmin. Rhein inhibited the expression of Rac1 and its downstream targets NOX1 and β-catenin. Rac1 silencing (si-Rac1) inhibited the accumulation of MDA and Fe2+ and the expression of Rac1, NOX1, β-catenin, α-SMA, TFR1, and ACSL4. Rac1 overexpression (oe-Rac1) resulted in the inhibition of superoxide dismutase (SOD), glutathione (GSH), GPX4 synthesis, and down-regulation of Recombinant Solute Carrier Family 7, Member 11 (SLC7A11) and Nephrin expression in HG-treated podocytes. Rac1 Lentivirus (LV-Rac1) injection significantly promoted the accumulation of MDA and Fe2+ and increased the expression of RAC1, NOX1, β-catenin, TFR1, ACSL4, and α-SMA in DN mice. Conclusions: Rhein inhibited ferroptosis and epithelial-mesenchymal transition (EMT) to attenuate DN by regulating the Rac1/NOX1/β-catenin axis.

Atherosclerosis may be caused or developed by an immune response and antioxidation imbalance. MicroRNA-375 (miR-375) or G-protein-coupled receptor 39 (GPR39) is involved in vascular endothelial cell injury, but their role in atherosclerosis is unknown. This experiment aimed to determine the action of the miR-375/GPR39 axis in atherosclerosis.Human aortic endothelial cells (HAECs) were treated with 10 ng/mL of oxidised low-density lipoprotein (ox-LDL) for 24 hours to induce HAEC injury, which was treated by the miR-375 inhibitor, GPR39 inhibitor, or agonist. High-fat diet (HFD) -induced ApoE−/− mice were made as an atherosclerosis model for miR-375 inhibitor treatment. Cell Counting Kit-8 was applied to detect HAEC viability. HAEC apoptosis and ROS levels were measured using flow cytometry. Vascular histopathology and the GPR39 expression were detected using hematoxylin-eosin and immunohistochemistry. The expressions of interleukin (IL) -6, IL-1β, and tumour necrosis factor-α (TNF-α) were assessed using an enzyme-linked immunosorbent assay. The miR-375, GPR39, NOX-4, and p-IκBα/IκBα levels were measured using quantitative reverse transcription polymerase chain reaction or western blot.MiR-375 and GPR39 levels increased and decreased in ox-LDL-treated HAECs, respectively. The miR-375 inhibitor or GPR39 agonist promoted cell viability and inhibited apoptosis in ox-LDL-induced HAEC injury. The miR-375 inhibitor also significantly downregulated the IL-6, IL-1β, TNF-α, p-IκBα/IκBα, ROS, and NOX-4 expressions to alleviate oxidative stress and inflammation, which were reversed by the GPR39 inhibitor. An in vivo experiment proved that the miR-375 inhibitor upregulated the GPR39 expression and improved inflammation, oxidative stress, and endothelial cell damage associated with atherosclerosis.The miR-375 inhibitor improved inflammation, oxidative stress, and cell damage in ox-LDL-induced HAECs and HFD-induced ApoE−/− mice by promoting the GPR39 expression, which provided a new theoretical basis for the clinical treatment of atherosclerosis.