Expression Levels Of TGF-β1 Research Articles

Effect of vaccination against Mycoplasma hyopneumoniae on divergent pig genetic groups

The bacterium Mycoplasma hyopneumoniae (Mhp) causes a chronic infectious respiratory disease in pigs, leading to important economic losses. This study aimed to compare the immune response of the local Piau breed and a commercial line to Mhp vaccination. For this, two phases were carried out. In the first, gene expression of toll-like receptors (TLR2, TLR4, TLR6, and TLR10) and cytokines (IL2, IL6, IL8, IL10, IL12, IL13, TNFα, and TGFβ) was assessed in porcine blood mononuclear cells (PBMC) from the two genetic groups before and after vaccination. In the second experiment, nitric oxide production, specific antibodies, and gene expression of toll-like receptors and cytokines were evaluated in bronchoalveolar lavage fluid (BALF) cells of vaccinated and unvaccinated pigs. After vaccination against Mhp, TLR2, TLR4, TLR6, TLR10, IL6, TNFα, and TGFβ expression levels were elevated in PBMC from commercial animals, and TLR6, TLR10, and TGFβ expression levels were elevated in PBMC from the Piau group. Vaccination also increased the production of Mhp-specific IgG antibodies in BALF cells in the Piau breed. Comparison of the two genetic groups revealed differences in TNFα and IL10 expression in BALF cells. These results show that Piau pigs have different immune responses to vaccination compared with commercial animals. It is worth noting that these genetic differences between both genetic groups may be related to phenotypic differences in Mhp resistance or susceptibility.

Construction a novel osteoporosis model in immune-deficient mice with natural ageing

Osteoporosis (OP) predominantly affects elderly individuals. Stem cells show potential for treating OP. However, animal models with normal immune function can eliminate implanted human cells. This study utilized naturally aging NOD/SCID mice, which exhibit immunodeficiency, to create a human osteoporosis model. This approach helps to minimize the premature immune clearance of transplanted allogeneic or xenogeneic cells in preclinical studies, allowing for a more accurate replication of the clinical pharmacological and pharmacokinetic processes involved in stem cell interventions for osteoporosis. NOD/SCID mice were fed until 12, 32, and 43 weeks of age, respectively, and then euthanized. We harvested lumbar vertebra for Micro‐Computed Tomography (Micro-CT) scanning and pathological examination. Additionally, we performed biomechanical testing of lumbar vertebra to assess the severity of osteoporosis. We utilized real-time RT-PCR to assess gene expression changes associated with bone metabolism, aging, inflammation, oxidative stress, and the Tgf-β1/Smad3 signaling pathway. In addition, the protein expression levels of P16, Tgf-β1 and Smad3 were detected using Western Blotting (WB). In comparison to 12-week-old mice, the 32-week-old and 43-week-old mice displayed significantly sparser and fractured trabeculae in their lumbar vertebra, lower bone mineral density (BMD), and changes in bone microstructural parameters (**P<0.01, ***P<0.001). Additionally, compared to 12-week-old mice, the 32-week-old and 43-week-old mice exhibited decreased expression of osteogenic genes (Alp, Opg, Sp7, Col1a1), increased expression of osteoclastic gene (Rankl), the number of TRAP-positive osteoclasts significantly increased in 32-week-old and 43-week-old mice compared to 12-week-old mice. The expression of genes related to aging and inflammatory (P16, Il-1β, Tnf-α) increases with advancing age (*P<0.05, **P<0.01, ***P<0.001). The expression of oxidative stress-related genes (Sod1, Sod2, Foxo3, Nrf2), as well as Tgf-β1 and Smad3 decreased with age (*P<0.05, **P<0.01, ***P<0.001). As age increases, the levels of P16 protein increase, Tgf-β1 and Smad3 proteins decrease. Our study successfully replicated osteoporosis models in NOD/SCID mice at both 32 and 43 weeks, with the latter exhibiting more severe osteoporosis. This condition seems to be driven by factors such as aging, inflammation, oxidative stress, and the Tgf-β1/Smad3 signaling pathway.

Early CSWT relieves myocardial fibrosis by inhibiting TGF-Smad2/3-MMP-9

Abstract Objective To investigate the correlation between the improvement of cardiac function and myocardial fibrosis and the TGF-β1-Smad2/3-MMP-9 signaling pathway in acute myocardial infarction (AMI) rats after early treatment with extracorporeal cardiac shock wave therapy (CSWT). Methods Thirty SD rats were randomly divided into five groups: sham group (6 rats), AMI group (6 rats), AMI+CSWT group (6 rats), AMI+SB431542 group (6 rats), and AMI+SB431542+CSWT group (6 rats). AMI was induced in all groups except the sham group. The AMI+CSWT group and AMI+SB431542+CSWT group received CSWT treatment. Echocardiography was performed at 4 days and 62 days postoperatively to assess left ventricular ejection fraction (LVEF) in all groups. Additionally, cardiac tissues were sampled and evaluated for pathological changes and degree of fibrosis by HE staining and Masson staining. The TGF-β1-Smad2/3-MMP-9 signaling pathway and protein expression of COL-I and COL-III were detected in the infarcted border area. Results (1) At 4 days postoperatively, EF and FS were significantly lower, and LVESV was enlarged in the AMI group, AMI+CSWT group, AMI+SB431542+CSWT group, and AMI+SB431542 group compared to the sham group (p &amp;lt; 0.05). (2) At 62 days postoperatively, EF and FS were significantly lower, and LVEDV and LVESV were significantly higher in the AMI group compared to the sham group (p &amp;lt; 0.05). EF and FS were significantly higher, and LVEDV and LVESV were significantly lower in the AMI+CSWT group compared to the AMI group (p &amp;lt; 0.05). There was no significant difference in EF, FS, LVEDV, and LVESV between the AMI+CSWT group and the AMI+SB431542+CSWT group (p &amp;gt; 0.05). (3) Histological analysis showed that the AMI group had more blue-stained areas compared to the sham group, while the AMI+CSWT group had fewer blue-stained areas and more neatly arranged red areas compared to the AMI group (p &amp;lt; 0.05). (4) Collagen volume fraction (CVF) was significantly higher in the AMI group compared to the sham group (p &amp;lt; 0.05). CVF was significantly lower in the AMI+CSWT group compared to the AMI group (p &amp;lt; 0.05). There was no significant difference in CVF between the AMI+CSWT group and the AMI+SB431542+CSWT group (p &amp;gt; 0.05). (5) The expression levels of TGF-β1, Smad3, MMP-9, COL-I, and COL-III were significantly higher in the AMI group compared to the sham group (p &amp;lt; 0.05). The expression levels of these proteins were significantly reduced in the AMI+CSWT group compared to the AMI group (p &amp;lt; 0.05). Conclusions Early CSWT improves cardiac function and inhibits myocardial fibrosis associated with the TGF-β1-Smad2/3-MMP-9 signaling pathway.pathological changesdegree of fibrosis

Evaluation effect of alginate hydrogel containing losartan on wound healing and gene expression.

Skin tissue engineering has become an increasingly popular alternative to conventional treatments for skin injuries. Hydrogels, owing to their advantages have become the ideal option for wound dressing, and they are extensively employed in a mixture of different drugs to accelerate wound healing. Sodium alginate is a readily available natural polymer with advantages such as bio-compatibility and a non-toxicological nature that is commonly used in hydrogel form for medical applications such as wound repair and drug delivery in skin regenerative medicine. Losartan is a medicine called angiotensin receptor blocker (ARB) that can prevent fibrosis by inhibiting AT1R (angiotensin II type 1 receptor). In this research, for the first time, three-dimensional scaffolds based on cross-linked alginate hydrogel with CaCl2 containing different concentrations of losartan for slow drug release and exudate absorption were prepared and characterized as wound dressing. Alginate hydrogel was mixed with 10, 1, 0.1, and 0.01mg/mL of losartan, and their properties such as morphology, chemical structure, water uptake properties, biodegradability, stability assay, rheology, blood compatibility, and cellular response were evaluated. In addition, the therapeutic efficiency of the developed hydrogels was then assessed in an in vitro wound healing model and with a gene expression. The results revealed that the hydrogel produced was very porous (porosity of 47.37 ± 3.76µm) with interconnected pores and biodegradable (weight loss percentage of 60.93 ± 4.51% over 14days). All hydrogel formulations have stability under various conditions. The use of CaCl2 as a cross-linker led to an increase in the viscosity of alginate hydrogels. An in vitro cell growth study revealed that no cytotoxicity was observed at the suggested dosage of the hydrogel. Increases in Losartan dosage, however, caused hemolysis. In vivo study in adult male rats with a full-thickness model showed greater than 80% improvement of the primary wound region after 2weeks of treatment with alginate hydrogel containing 0.1mg/mL Losartan. RT-PCR and immunohistochemistry analysis showed a decrease in expression level of TGF-β1 and VEGF in treatment groups. Histological analysis demonstrated that the alginate hydrogel containing Losartan can be effective in wound repair by decreasing the size of the scar and tissue remodeling, as evidenced by future in vivo studies.

Allicin Ameliorated High-glucose Peritoneal Dialysis Solution-induced Peritoneal Fibrosis in Rats via the JAK2/STAT3 Signaling Pathway.

Peritoneal fibrosis (PF) is one of the most serious complications of peritoneal dialysis (PD) and is the greatest obstacle to the clinical application of PD. Chinese herbal monomers have been effective in the prevention and treatment of PF. The aim of this study was to observe the effect of allicin on PF in rats induced by high glucose and to investigate its molecular mechanism of action. A rat model of PF was established by using a 4.25% glucose-based standard peritoneal dialysis solution. The degree of peritoneal pathological damage was assessed by Hematoxylin and eosin (H&E) staining. Peritoneal collagen deposition was detected by Masson's trichrome staining. The levels of Interleukin-6 (IL-6), Tumor necrosis factor-α (TNF-α), Interleukin-1β (IL-1β) and monocyte chemoattractant protein-1 (MCP-1) in the serum were measured by Enzyme Linked Immunosorbent Assay (ELISA). The expression levels of TGF-β, α-smooth muscle actin (α-SMA) and collagen I were examined by western blotting and immunohistochemistry. The protein expression levels and mRNA levels of E-cadherin, N-cadherin, vimentin, janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) in peritoneal tissue were determined by western blotting and qRT-PCR. TGF-β1 stimulated human peritoneal mesothelial cells (HPMCs), and the cells were treated with allicin and the JAK2/STAT3 pathway activator colivelin alone or in combination. A cell counting kit-8 (CCK-8) assay was used to measure cell viability. The role of JAK2/STAT3 in the effects of allicin was confirmed via in vitro mechanistic research by western blotting, wound healing assays and Transwell assays. Allicin relieves the inflammatory response by downregulating the levels of IL-1β, IL-6, MCP-1 and TNF-α. Furthermore, allicin decreased the expression of TGF-β, α-SMA and collagen I. Allicin also alleviated epithelial-to-mesenchymal transition (EMT), as specifically manifested by increased E-cadherin and reduced N-cadherin and vimentin. Further studies revealed that allicin reduced the protein levels of JAK2, STAT3, p-JAK2, and p-STAT3. The results of the cellular experiments verified the above results. The ability of allicin to inhibit fibrosis and the EMT process was significantly attenuated after HPMCs were treated with colivelin. Taken together, these findings suggest that allicin inhibits inflammation and EMT, thereby improving PF, and this protective effect may be achieved by inhibiting the JAK2/STAT3 signaling pathway.

Betulin gel alleviates esophageal stricture following endoscopic submucosal dissection: an animal study.

Esophageal stenosis is a troublesome complication after circumferential ESD. This study examined the efficacy of betulin gel in preventing esophageal stenosis after ESD in a porcine model. Twelve pigs were randomized to betulin group and control group evenly. At the distal esophagus, circumferential ESD was performed in all animals. In the betulin group, betulin gel was applied at days 1, 3, and 7. Endoscopy examination was performed at day 3, 1week, 2weeks, and 4weeks post-ESD. Then pigs were killed for macroscopic and histologic esophageal evaluation. The rate of esophageal stricture was lower in the betulin group (53.3 ± 12.5% vs 88.3% ± 2.9, p = 0.02). Betulin-treated pigs had lower dysphagia score (2.0 ± 0 vs 3.3 ± 0.5, p < 0.001), less weight loss (11.78% ± 2.16 vs 15.85% ± 3.63, p = 0.04), and better passability of the open and closed biopsies forceps (83.33% vs. 0%, p = 0.015, and 100% vs. 0%, p = 0.002) 4weeks post-ESD. Histologically, better re-epithelization (63.2 ± 10.7mm vs 22.8 ± 10.1mm, p < 0.001), slighter submucosal fibrosis (0.95 ± 0.17mm vs 2.32 ± 0.48mm, p = 0.002), lower muscularis propria damage score (1 vs 3, p < 0.001), and less inflammatory cells (307 vs 675 per high-power field, p = 0.002) were noted in the betulin group. The expression levels of TGF-β1, collagen i, collagen III, and α-SMA were significantly lower in the betulin group compared to the control group (p < 0.05). Betulin gel shows promise in reducing fibrosis, enhancing repair, and preventing esophageal stricture after ESD, suggesting a potential new strategy for prevention.

Transforming growth factor-beta and Epithelial-mesenchymal transition is considered an attractive pivotal Genetic-Biomarker for Hepatocellular carcinoma with hepatitis C virus-infection patients

Abstract Introduction/Objective Hepatocellular carcinoma (HCC) is the most common cause of cancer worldwide. HCC is induced by hepatitis C-virus (HCV) which represents a major health problem in Egypt. The transforming growth factor- beta (TGF-β) is a potent cytokine that has a multifunctional pro-fibrotic activity, for enhancing liver-inflammation, fibrosis, tumor-progression, and metastasis. TGF-β has a vital role in the regulation of tumorigenesis, by controlling epithelial-mesenchymal-transition (EMT), apoptosis, proliferation, and differentiation process. The dysregulation of TGF-β, Wnt/β-catenin signaling pathways, by HCV-core protein is implicated in the development of HCC. Methods/Case Report The study aims to determine the effect of the HCV-infection in HCC patients with a set of growth-factors, EMT, and anti-apoptotic protein expression levels by using western blotting and immunohistochemistry staining analysis. Results (if a Case Study enter NA) These results observed an insignificant increase in the expression of TGF-β protein for HCC/HCV-infected cells compared to the HCV-uninfected cells, and the expression of the β-catenin protein level is elevated in the nucleus. Also, it showed that Bcl-2 protein is a significant increase in HCV-infected when compared to HCV-free groups. Positive staining for p53 protein was detected in the HCC/HCV-infected, more than in HCC/HCV-uninfected cells. This data indicated that the core-protein of HCV can switch TGF-β from a tumor suppressor to a tumor promoter by suppressing apoptosis of hepatocytes and increasing EMT expression by activation of Wnt/β-catenin signaling pathway. Therefore, HCV-core protein enhances the nuclear accumulation of β-catenin expression protein to activate downstream target genes, which regulate cell-growth and cell-cycle progression. Conclusion It is speculated that HCV may exert an anti-apoptotic effect, and thus enhance tumorigenesis. Also, the accumulation of nuclear Wnt/β-catenin and p53 correlates with TGF-β expression, typically a late event in hepato- carcinogenesis. Taken together, the expression level of TGF-β, Wnt/β-catenin, Bcl-2, and p53 genotype in HCV is considered an indicator of shortened survival in patients, and may be an attractive clinically pivotal genetic biomarker to mediate liver pathogenesis.

Jianpi-Huatan-Huoxue-Anshen formula ameliorates gastrointestinal inflammation and microecological imbalance in chemotherapy-treated mice transplanted with H22 hepatocellular carcinoma.

Jianpi-Huatan-Huoxue-Anshen formula [Tzu-Chi cancer-antagonizing & life-protecting II decoction (TCCL)] is a Chinese medical formula that has been clinically shown to reduce the gastrointestinal side effects of chemotherapy in cancer patients and improve their quality of life. However, its effect and mechanism on the intestinal microecology after chemotherapy are not yet clear. To discover the potential mechanisms of TCCL on gastrointestinal inflammation and microecological imbalance in chemotherapy-treated mice transplanted with hepatocellular carcinoma (HCC). Ninety-six mice were inoculated subcutaneously with HCC cells. One week later, the mice received a large dose of 5-fluorouracil by intraperitoneal injection to establish a HCC chemotherapy model. Thirty-six mice were randomly selected before administration, and feces, ileal tissue, and ileal contents were collected from each mouse. The remaining mice were randomized into normal saline, continuous chemotherapy, Yangzheng Xiaoji capsules-treated, and three TCCL-treated groups. After treatment, feces, tumors, liver, spleen, thymus, stomach, jejunum, ileum, and colon tissues, and ileal contents were collected. Morphological changes, serum levels of IL-1β, IL-6, IL-8, IL-10, IL-22, TNF-α, and TGF-β, intestinal SIgA, and protein and mRNA expression of ZO-1, NF-κB, Occludin, MUC-2, Claudin-1, and IκB-α in colon tissues were documented. The effect of TCCL on the abundance and diversity of intestinal flora was analyzed using 16S rDNA sequencing. TCCL treatment improved thymus and spleen weight, thymus and spleen indexes, and body weight, decreased tumor volumes and tumor tissue cell density, and alleviated injury to gastric, ileal, and colonic mucosal tissues. Among proteins and genes associated with inflammation, IL-10, TGF-β, SIgA, ZO-1, MUC-2, and Occludin were upregulated, whereas NF-κB, IL-1β, IL-6, TNF-α, IL-22, IL-8, and IκB-α were downregulated. Additionally, TCCL increased the proportions of fecal Actinobacteria, AF12, Adlercreutzia, Clostridium, Coriobacteriaceae, and Paraprevotella in the intermediate stage of treatment, decreased the proportions of Mucipirillum, Odoribacter, RF32, YS2, and Rikenellaceae but increased the proportions of p_Deferribacteres and Lactobacillus at the end of treatment. Studies on ileal mucosal microbiota showed similar findings. Moreover, TCCL improved community richness, evenness, and the diversity of fecal and ileal mucosal flora. TCCL relieves pathological changes in tumor tissue and chemotherapy-induced gastrointestinal injury, potentially by reducing the release of pro-inflammatory factors to repair the gastrointestinal mucosa, enhancing intestinal barrier function, and maintaining gastrointestinal microecological balance. Hence, TCCL is a very effective adjuvant to chemotherapy.

Macrophage polarization and macrophage-related factor expression in hypertrophy of the ligamentum flavum.

Owing to the unknow types of infiltrating macrophages and the corresponded factors, we aimed to investigate the specific types of infiltrating macrophages involved in HLF and the expression of macrophage-related factors. The ligamentum flavum was obtained from patients with lumbar spinal stenosis (HLF group; n = 15) and lumbar disc herniation (non-hypertrophic ligamentum flavum [NLF] group; n = 15). Ligamentum flavum specimens were paraffin embedded, followed by histological and immunohistochemical staining to identify the macrophage type and expression of macrophage-related factors. The HLF group demonstrated CD206 marker expression, while the NLF group did not (P < 0.0001; n = 11). CD68 marker was expressed in both groups (P > 0.05; n = 11). CCR7 was not expressed in either group. The expression levels of the extracellular matrix proteins aggrecan (Agg), type I collagen (Coll1), and type II collagen (Coll2) were higher in the HLF group than in the NLF group (P < 0.0001; n = 11). The aging markers p21, p16, and p53 were expressed in the HLF group, but not in the NLF group (P < 0.0001; n = 11). The expression levels of the inflammatory factors TNF-α and IL-1β were higher in the HLF group than in the NLF group (P < 0.0001; n = 11). Similarly, the expression level of the fibrosis factor TGF-β1 was higher in the HLF group than in the NLF group (P < 0.0001; n = 11). The infiltration of M2 macrophages may be involved in HLF, while involvement of M1 macrophages may only occur early in inflammation. The expression of extracellular matrix proteins and macrophage-related factors was increased. Aging may also be associated with HLF.

VSIG-3/IGSF11 silencing in A2058 melanoma cells simultaneously suppresses melanoma progression and induces anti-tumoral cytokine profile in human T cells: In silico and in vitro study.

VISTA is a newly discovered immune checkpoint whose functional mechanisms have become increasingly important to study due to its brilliant results in cancer immunotherapy. Despite VSIG-3/IGSF11 being identified as an inhibitory ligand for VISTA with potential as a target for cancer immunotherapy, very little is known of its functions. This study aimed to conduct a detailed analysis of VSIG-3/IGSF11 in melanoma, as well as to study the effects of its silencing on melanoma cell line progression and human T cell functions. Online databases were used to investigate VSIG-3/IGSF11 expression, its relationships, and prognostic value in melanoma. Then, the effects of VSIG-3/IGSF11 silencing on proliferation, migration, cell cycle arrest, and apoptosis in A2058 melanoma cells were assessed using MTT, colony formation, wound healing, cell cycle, and Annexin-VFITC/PI assays, respectively. Finally, A2058 cells transfected with VSIG-3/IGSF11 siRNA were co-cultured with human T cells, and the expression levels of T cell cytokines were evaluated using qRT-PCR. VSIG-3/IGSF11 expression was significantly increased in melanoma patients and cell lines; however, no correlation was found between VSIG-3/IGSF11 expression levels and clinicopathological characteristics, survival, or immune cell infiltration. Following VSIG-3/IGSF11 silencing in A2058 cells, viability, proliferation, and migration rates were decreased, while apoptosis was increased. T cells co-cultured with VSIG-3/IGSF11 siRNA-transfected A2058 cells exhibited increased expression levels of IFN-γ and IL-12 and decreased expression levels of IL-10, TGF-β, and TNF-α. The inhibitory effect of VSIG-3/IGSF11 silencing on A2058 melanoma cell progression, along with the alteration of T cell cytokines towards a pro-inflammatory phenotype, suggests that VSIG-3/IGSF11 is primarily involved in melanoma progression and modulating immune responses. Therefore, it may be a valuable target for immunotherapy in melanoma patients.

Tinglu Yixin granule inhibited fibroblast-myofibroblast transdifferentiation to ameliorate myocardial fibrosis in diabetic mice

Ethnopharmacological relevanceMyocardial fibrosis is one of the pathological characteristics of advanced diabetic cardiomyopathy (DCM) and serves as the strong evidence of poor prognosis. Among them, the transdifferentiation of cardiac fibroblasts (CFs) may play a crucial role in the development of myocardial fibrosis in DCM. Tinglu Yixin granule (TLYXG) has been clinically used for many years and can significantly improve cardiac function of patients with DCM. However, the effect of TLYXG on myocardial fibrosis in DCM remains unknown, and the underlying mechanisms of its efficacy have yet to be fully understood. Aim of the studyThis study aimed to investigate the impact and underlying mechanism of TLYXG on myocardial fibrosis in diabetes mice. Materials and methodsThe bioactive compounds in TLYXG were identified using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). The potential mechanism of TLYXG in treating DCM was predicted using network pharmacology combined with molecular docking and protein-protein docking. The mice model of type 2 diabetes were established by intraperitoneal injection of streptozotocin (STZ) and the high-fat diet (HFD). Indicators of pancreatic islet function, lipids, oxidative stress, and inflammatory factors were tested using kits. Cardiac function was assessed in diabetic mice using echocardiography. Histologic staining was performed to evaluate myocardial hypertrophy and fibrosis. Mechanistically, the hypothesis was tested through rescue experiments. The expression levels of transient receptor potential channel 6 (TRPC6), transforming growth factor-β1 (TGF-β1), collagen I (COL-I) and alpha-smooth muscle actin (α-SMA), along with the mRNA and phosphorylation levels of SMAD family member 3 (Smad3) and protein 38 mitogen-activated protein kinase (p38 MAPK), were assessed using quantitative RT-qPCR, western blot, immunohistochemistry, and immunofluorescence. Neonatal lactating mice were used to extract primary CFs for vitro experiments. Scratch and transwell assays were conducted to assess CFs migration and invasion abilities. Western blot and immunofluorescence were used to evaluate the expression levels of CFs transdifferentiation markers COL-I and α-SMA. ResultsA total of 168 active ingredients were detected in TLYXG based on UPLC-MS and databases. Network pharmacology indicated that TLYXG could improve DCM through inflammatory mediator regulation of TRP channels, TGF-beta signaling pathway, and MAPK signaling pathway. ELISA results showed that TLYXG could ameliorate metabolic levels, inflammation, and oxidative stress in diabetic mice. Echocardiography suggested that TLYXG improved cardiac systolic and diastolic dysfunction in diabetic mice. Histological analysis revealed that TLYXG alleviated myocardial fibrosis in diabetes mice. Additionally, molecular docking analysis indicated strong binding activity between the main active ingredients of TLYXG and TRPC6 of the TRP family. At the molecular level, TLYXG reduced the mRNA and protein expression levels of TRPC6 and TGF-β1 and inhibited the mRNA and phosphorylation levels of Smad3 and p38 MAPK. Furthermore, TLYXG inhibited CFs migration and invasion, and reduced the expression levels of the CFs transdifferentiation markers COL-I and α-SMA. ConclusionTLYXG inhibited the proliferation, migration, invasion and transdifferentiation of CFs by suppressing TGF-β1/Smad3/p38 MAPK signaling through down-regulation of TRPC6, thereby ameliorating myocardial fibrosis in diabetes mice.

Overexpression of ATF4 Inhibits Ferroptosis to Alleviate Anxiety Disorders by Activating the TGF-β Signaling Pathway.

Anxiety disorders seriously impair patients' mental health and quality of life, with limited effectiveness of current treatments. Dysregulation of activating transcription factor 4 (ATF4) is involved in various mental diseases, but the research on its potential roles in alleviating anxiety disorders remains limited. ATF4 was screened out by bioinformatic analysis and its expression was verified in vivo. Mice were treated with 21 d of chronic restraint stress to establish the anxiety mice model. The anxiolytic effect of ATF4 was assessed by a battery of behavior tests and evaluation of hippocampal tissue damage after overexpressing ATF4. Ferroptosis-related indicators were detected by enzyme-linked immunosorbent assay and Western blotting. Then the transforming growth factor beta (TGF-β) signaling pathway was predicted as the downstream regulatory pathway of ATF4 by bioinformatic methods. Western blotting was conducted to detect the protein expression level of TGF-β1, small mothers against decapentaplegic 3 (Smad3), and phospho-Smad3 (p-Smad3). ATF4 was screened out as a ferroptosis-related anxiolytic gene after bioinformatics analysis and was down-regulated in the anxiety mice model. Mice with ATF4 overexpression spent more time in the open arms in the elevated plus-maze test, appeared more frequently in the central area in the open-field test, and decreased the immobility time in the forced swimming and tail suspension tests. Hippocampal tissue damage was alleviated, ferroptosis was suppressed, and the levels of TGF-β1 and p-Smad3/Smad3 were increased by AFT4 overexpression. ATF4 overexpression can repress ferroptosis to improve anxiety disorders by activating the TGF-β signaling pathway.

Cloning and Expression Analysis of TGF-β Type I Receptor Gene in Hyriopsis cumingii.

The TGF-β signaling pathway plays an important role in wound healing and immune response. In this study, a TGF-β type I receptor (TGF-βRI) homolog was cloned and characterized from freshwater mussel Hyriopsis cumingii. The full-length cDNA of the TGF-β RI gene was 2017 bp, with a 1554 bp open reading frame (ORF), and encoded 517 amino acids. The predictive analysis further identified distinct regions within the TGF-βRI protein: a signal peptide, a membrane outer region, a transmembrane region, and an intracellular region. Real-time quantitative PCR results showed that the TGF-β RI gene was expressed in all tissues of healthy mussels. The transcripts of TGF-β RI in hemocytes and hepatopancreas were significantly up-regulated at different periods after stimulation with Aeromonas hydrophila and peptidoglycan (PGN) (P < 0.05). The mRNA expression of TGF-β RI progressively increased from day 1 to day 10 after trauma (P < 0.05), and it returned to the initial level by day 15. The expression levels of TGF-β , Smad5, MMP1/19, and TIMP1/2, but not Smad3/4, were significantly up-regulated at different time points after trauma. However, the expression levels of TGF-β , MMP1/19, and TIMP2 were decreased after treatment with the inhibitor SB431542. Furthermore, the recombinant TGF-βRI proteins were expressed in vitro and existed in the form of inclusion bodies. Western blotting results showed that TGF-βRI proteins were expressed constitutively in various tissues of mussels, and their expression was up-regulated after trauma, which was consistent with the mRNA expression trend. These results indicate that TGF-β RI is involved in the process of wound repair and immune response.

Saffron improves the efficacy of immunotherapy for colorectal cancer through the IL-17 signaling pathway

Ethnopharmacological relevanceSaffron is one of the traditional medicinal herbs, which contains various active ingredients, such as safranal, crocin, saffron acid, etc. It has anti-inflammatory, antioxidant, and anti-cancer properties, and is widely used in clinical practice. The anti-cancer efficacy of saffron has been previously confirmed, but its anti-cancer mechanism in colorectal cancer remains unclear. ObjectiveWe investigated the effect of active compounds of saffron on the efficacy of immunotherapy for colorectal cancer. MethodsTCMSP and liquid chromatography-mass spectrometry analysis (LC-MS), GeneCards, and DisGeNET databases were used to identify the active compounds of saffron, drug targets and the disease targets of colorectal cancer. They were then subjected to Gene Ontology Enrichment (GO) and Signalling Pathway Enrichment (KEGG) analyses. The core targets and corresponding compounds were selected for molecular docking. The effect of active components of saffron on the proliferation of CT26 and HCT116 cells was investigated using the cell counting kit-8 (CCK-8). In vitro experiments were conducted by subcutaneous injection of CT26 cells to establish a colon cancer model. Enzyme-linked immunosorbent assay (ELISA), western blotting (WB), real-time polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), and flow cytometry (FCM) were employed to validate the effects of saffron on colorectal cancer immunotherapy. Results1. LC-MS analysis revealed that the main active component of saffron extract was crocin. The active chemicals of saffron intersected with 170 colorectal cancer targets, with 17 predicting targets for saffron treatment. GO and KEGG enrichment analyses revealed that the active components of saffron can prevent colorectal cancer development by enhancing Th17 cell differentiation and the IL-17 signaling pathway. 2. In vitro studies revealed that saffron alcohol extract, crocin, and safranal can suppress the proliferation of CT26 and HCT116 cells. 3. In vivo studies showed that crocin and safranal can increase the body mass and decrease the tumor mass of loaded mice, decrease the serum level of IL-17, and lower the mRNA expression level of IL-17, IL-6, TNF-α, TGF-β, and PD-L1 and IL-17, PD-L1 protein in tumors. This inhibitory effect was strengthened after combined immunotherapy. In addition, saffron modulated CD4+ and CD8+ T cells and the CD4+/CD8+T ratio in mouse spleens. ConclusionThe active components of saffron can reduce the expression of inflammatory factors and ameliorate the immunological microenvironment of tumors via the IL-17 signaling pathway, thereby improving the efficacy of immunotherapy for colorectal cancer. This study provides pharmacological support for the application of saffron in enhancing the efficacy of immunotherapy for colorectal cancer.

In vitro assessment of Dracocephalum lindbergii for growth inhibition, apoptosis induction, and cytokine modulation against Leishmaniamajor

Leishmaniasis is a vector-borne disease caused by several species of flagellate protozoan parasites belonging to the genus Leishmania, which are part of the Trypanosomatidae family. This study aims to evaluate the in vitro anti-Leishmania activity of Dracocephalum lindbergii Rech.f (D. lindbergii) on the growth, apoptosis induction, and proliferation of Leishmania major. To conduct this study, the aerial parts of D. lindbergii were collected during the flowering stage in May 2022 from North Khorasan province, Iran. The plant material was extracted using various solvents, starting with those of lower polarity and progressing to those of higher polarity. To assess the impact of the D. lindbergii fraction on L. major promastigotes, amastigotes, and macrophages (THP-1), cell viability was determined using the MTT assay. Additionally, flow cytometry with the Annexin V-PE apoptosis detection kit was employed to distinguish between viable, necrotic, and apoptotic promastigotes in response to treatment with the 100 % methanolic fraction of D. lindbergii (D). The expression levels of TNF-α, IFN-γ, IL-12, IL-10, TGF-β, IL4, iNOS, and GAPDH were quantified using real-time PCR (qPCR) on the macrophage cell line. Each treatment approach exhibited marked anti-leishmanial effects across different concentrations over 24, 48, and 72 h of incubation, showing statistically significant differences compared to the untreated control group (P < 0.001). Different concentrations of D, MAT, and AmpB, both individually and in combination, significantly reduced the total number of intramacrophage amastigotes compared to the untreated control groups at 24, 48, and 72 h. The results also showed time-dependent variations in the anti-Leishmania activity of the fraction. In terms of cellular morphology, treated cells exhibited changes such as shrinkage, cytoplasmic condensation, and reduced mobility, particularly noticeable after 24 h of treatment. Additionally, fraction D demonstrated significant antioxidant properties. This study highlights the potential of D. lindbergii as an anti-Leishmania agent, with the 100 % methanolic fraction emerging as a promising candidate for the development of novel treatments for leishmaniasis.

Research on ameliorating pulmonary fibrosis in silicosis mice of Cordyceps cicadae polysaccharides

Objective: A mouse silicosis model was constructed by injecting silicon dioxide (SiO(2)) particles into the trachea to explore the effect and mechanism of Cordyceps cicadae polysaccharides (CCP) on ameliorating pulmonary fibrosis in silicosis mice. Methods: In May 2023, CCP were extracted and isolated, the monosaccharide composition and functional group composition were analyzed by high performance liquid chromatography and Fourier transform infrared spectroscopy. C57BL/6J mice were injected with 50 μl 50 mg/ml SiO(2) suspension to construct silicosis mouse model, which were then randomly divided into model group, CCP intervention groups [low dose group (LCCP group), medium dose group (MCCP group) and high dose group (HCCP group) ], the control group was administered by physiological saline, 8 mice in each group. Mice in the CCP intervention groups received oral gavage administration once daily with CCP solution (100, 200 and 400 mg/kg), while control group and model group received physiological saline, lasted for 30 days. The body weight of mice was recorded and the lung coefficient was calculated. The pathomorphological changes of mouse lung tissue were determined by HE and Masson staining. The contents of fibrosis indexes [hydroxyproline acid (HYP), connective tissue growth factor (CTGF) and matrix metallopeptidase 2 (MMP-2) ] of lung tissue and the pro-inflammatory factors[tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) ] of lung tissue and alveolar lavage fluid were determined by ELISA. The expression level of Collagen Ⅰ was determined by immunohistochemistry. The relative protein expression levels of transforming growth factor-β1 (TGF-β1), P-Smad2, α-smooth muscle actin (α-SMA), Toll-like receptor 4 (TLR4), nuclear factor kappa-B p65 (NF-κBp65) and myeloid differentiation primary response gene 88 (MyD88) in lung tissue were determined by Western blot. Results: The total sugar content of the CCP was 86.78%, composed of D-mannose, D-rhamnose, D-glucose and D-galactose, with a molar ratio of 12.71∶1.53∶1.00∶12.64. The infrared spectrum indicated the characteristic groups of its polysaccharides. Compared with the control group, the body weight of mice in the model group was decreased, lung coefficient was increased, the contents of HYP, CTGF and MMP-2 in lung tissue were increased, and the contents of TNF-α, IL-1β and IL-6 in lung tissue and alveolar lavage fluid were increased (P<0.05). The mice lung showed massive inflammatory cell infiltration and collagen fiber deposition, and the silicosis fibrosis was severe. The expression of CollagenⅠin lung tissue of model group was increased, and the proteins expression levels of TGF-β1, P-Smad2/Smad2, α-SMA, TLR4, NF-κBp65 and MyD88 were increased in mouse lung tissue (P<0.05). Compared with the model group, the body weights of mice in the MCCP and HCCP groups were increased, the lung coefficients were decreased, the contents of HYP, CTGF and MMP-2 in lung tissue were decreased, and the contents of TNF-α, IL-1β and IL-6 in lung tissue and alveolar lavage fluid were decreased (P<0.05). The inflammatory cell infiltration in the lung was reduced, and the degree of fibrosis was improved to varying degrees. The expression level of CollagenⅠwas down-regulated in the lung tissue of MCCP and HCCP groups, and the protein expression levels of TGF-β1, P-Smad2/Smad2, α-SMA, TLR4, NF-κBp65 and MyD88 were decreased in lung tissue (P<0.05) . Conclusion: The CCP could reduce the levels of fibrosis-related indicators and pro-inflammatory factors in lung tissue, ameliorating mouse lung inflammation and silicosis fibrosis caused by SiO(2) particles by inhibiting the activation of TGF-β1/Smad pathway and TLR4/nuclear factor kappa-B (NF-κB) pathway.

The effect of different potency glucocorticosteroids treatments on tear inflammatory factors and corneal optical density after femtosecond-assisted laser in situ keratomileusis.

To assess the impact of glucocorticosteroids with varying potencies on inflammatory mediators in tears and corneal optical density after femtosecond-assisted laser in situ keratomileusis (FS-LASIK). In a prospective study, 110 patients (220 eyes) who underwent FS-LASIK were divided into 2 groups: 55 patients (110 eyes) received dexamethasone, and another 55 patients (110 eyes) received fluorometholone. Visual acuity, intraocular pressure, and corneal optical density were measured before, 1 week, and 1 month after surgery. Tear fluid samples were also collected to assess expression levels of TNF-α, IL-1α, IL-6, and TGF-β1. One week after the procedure, the dexamethasone group exhibited elevated intraocular pressure (IOP) levels (P > .05) and a decreased expression of TNF-α in tears (P < .001) compared to the fluorometholone group. Within the 0 to 2 mm range from the corneal apex, the anterior corneal layer's optical density in the fluorometholone group surpassed that of the dexamethasone group (P < .05). At 1 month post-surgery, the IOP in the fluorometholone group was higher than that in the dexamethasone group (P < .05). In both the 0 to 2 mm and 2 to 6 mm intervals from the corneal apex, the optical density of the anterior corneal layer was significantly higher in the fluorometholone group compared to the dexamethasone group (P < .05). There was no statistically significant difference in visual acuity between the 2 groups at any postoperative time point. Short-term use of potent corticosteroids after FS-LASIK can swiftly address ocular surface inflammation, enhance corneal wound healing, reduce corneal edema, and accelerate the restoration of corneal transparency, in contrast to prolonged use of milder corticosteroids post-surgery.

Effect of goose-derived adiponectin peptide gADP3 on LPS-induced inflammatory injury in goose liver

ABSTRACT 1. This study evaluated the effects and mechanisms of action of the peptide gADP3 on hepatic inflammatory injury induced by lipopolysaccharide (LPS). 2. Hepatic inflammatory injury was induced in geese by intraperitoneal injection of LPS and gADP3, and the adiponectin receptor agonist AdipoRon (positive control) was used for potential amelioration. Serum inflammatory factor levels, liver function-related biochemical indicators and oxidative stress-related biochemical parameters in the liver tissues were determined. The expression levels of adiponectin and its receptors, inflammation and oxidative stress-related genes and key signalling molecules involved in adiponectin, inflammation and oxidative stress signalling pathways in liver tissues were detected. 3. The peptide gADP3 alleviated inflammatory cell infiltration and hepatic inflammatory changes, reversed the decrease in serum albumin (ALB), total protein (TP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) content or activity induced by LPS and increased the activity of the antioxidant enzymes CAT (catalase), SOD (superoxide dismutase) and GSH-Px (glutathione peroxidase). 4. The peptide gADP3 upregulated the expression of antioxidant enzyme-related genes GCLC, HO-1 and NQO1 in liver tissues, decreased the levels of inflammatory factors like TNF-α, IL-1β, IL-6, IFN-γ and TGF-β and reduced mRNA expression levels of inflammatory-related genes TNF-α, IL-1β, iNOS and TGF-β. Additionally, it increased the mRNA and protein expression levels of adiponectin and its receptors, as well as key molecules in the adiponectin signalling pathway like AMPK and PPARα. In addition, gADP3 reversed the changes in mRNA or protein expression of inflammatory and oxidative stress signalling pathway-related genes P38MAPK, NF-κBP65, TLR4 and Nrf2 in liver tissues caused by LPS treatment. 5. In conclusion, goose-derived adiponectin peptide gADP3, similar to the adiponectin receptor agonist AdipoRon, attenuated LPS-induced hepatic inflammatory injury in geese.

Magnamosis improves the healing of gastrojejunal anastomosis and down-regulates TGF-β1 and HIF-1α in rats

This study elucidated the unique pathological features of tissue healing by magnamosis and revealed the changes in landmark molecule expression levels related to collagen synthesis and tissue hypoxia. Forty-eight male Sprague–Dawley rats were divided into the magnamosis and suture anastomosis groups, and gastrojejunal anastomosis surgery was performed. Rats were dissected at 6, 24, and 48 h and 5, 6, 8, 10, and 12 days postoperatively. Hematoxylin, eosin, and Masson’s trichrome staining were used to evaluate granulation tissue proliferation and collagen synthesis density at the anastomosis site. Immunohistochemistry was used to measure TGF-β1 and HIF-1α expression levels. Magnamosis significantly shortened the operation time, resulting in weaker postoperative abdominal adhesions (P < 0.0001). Histopathological results showed a significantly lower granulation area in the magnamosis group than in the suture anastomosis group (P = 0.0388), with no significant difference in the density of collagen synthesis (P = 0.3631). Immunohistochemistry results indicated that the magnamosis group had significantly lower proportions of TGF-β1-positive cells at 24 (P = 0.0052) and 48 h (P = 0.0385) postoperatively and HIF-1α-positive cells at 24 (P = 0.0402) and 48 h postoperatively (P = 0.0005). In a rat model of gastrojejunal anastomosis, magnamosis leads to improved tissue healing at the gastrojejunal anastomosis, associated with downregulated expression levels of TGF-β1 and HIF-1α.

Endothelial mesenchymal transition promotes pannus formation in ankylosing spondylitis byactivation of TGF-β/SMAD signalling pathway.

To explore the role of endothelial-mesenchymal transition (EndMT) mediated by the TGF-β/SMAD signalling pathway in the pathogenesis of ankylosing spondylitis (AS). Serum levels of TGF-β1 were measured by enzyme-linked immunosorbent assay (ELISA) in 48 patients with AS and 15 healthy subjects. The expression levels of TGF-β1, SMAD7, CTGF, CD34 and EndMT-related markers (α-SMA, vimentin, FSP-1, VE-cadherin) in the sacroiliac joint (SIJ) of three AS patients were detected by immunohistochemistry, and three non-spondyloarthritis (SpA) autopsy samples were used as controls. Serum TGF-β1 level of AS patients was significantly higher than that of healthy controls (22971 ± 7667 pg/ml vs. 14837±4653 pg/ml, p<0.01). Compared with the non-SpA control group, the microvascular density (MVD) at the pannus formation site of SIJ in AS patients was significantly increased, accompanied by respectively increased expressions of TGF-β1, CTGF, α-SMA, vimentin, and FSP-1 (all p<0.05), whereas respectively decreased expressions of VE-cadherin and SMAD7 (p<0.01). The expression level of FSP-1 was positively correlated with levels of TGF-β1 and MVD, and negatively correlated with SMAD7. Our findings show that EndMT is involved in the promotion of pannus formation by TGF-β/SMAD signalling pathway activation in AS.