Tumor progression is closely related to tumor tissue metabolism and reshaping of the microenvironment. Oral squamous cell carcinoma (OSCC), a representative hypoxic tumor, has a heterogeneous internal metabolic environment. To clarify the relationship between different metabolic regions and the tumor immune microenvironment (TME) in OSCC, Single cell (SC) and spatial transcriptomics (ST) sequencing of OSCC tissues were performed. The proportion of TME in the ST data was obtained through SPOTlight deconvolution using SC and GSE103322 data. The metabolic activity of each spot was calculated using scMetabolism, and k-means clustering was used to classify all spots into hyper-, normal-, or hypometabolic regions. CD4T cell infiltration and TGF-β expression is higher in the hypermetabolic regions than in the others. Through CellPhoneDB and NicheNet cell-cell communication analysis, it was found that in the hypermetabolic region, fibroblasts can utilize the lactate produced by glycolysis of epithelial cells to transform into inflammatory cancer-associated fibroblasts (iCAFs), and the increased expression of HIF1A in iCAFs promotes the transcriptional expression of CXCL12. The secretion of CXCL12 recruits regulatory T cells (Tregs), leading to Treg infiltration and increased TGF-β secretion in the microenvironment and promotes the formation of a tumor immunosuppressive microenvironment. This study delineates the coordinate work axis of epithelial cells-iCAFs-Tregs in OSCC using SC, ST and TCGA bulk data, and highlights potential targets for therapy.
Humans ; Carcinoma, Squamous Cell/metabolism* ; Squamous Cell Carcinoma of Head and Neck ; Mouth Neoplasms/metabolism* ; Immunosuppression Therapy ; Transforming Growth Factor beta ; Head and Neck Neoplasms ; Gene Expression Profiling ; Tumor Microenvironment

Jiming Powder is a traditional ancient prescription with good therapeutic effect in the treatment of heart failure, but its mechanism lacks further exploration. In this study, a mouse model of coronary artery ligation was used to evaluate the effect and mechanism of Jiming Powder on myocardial fibrosis in mice with myocardial infarction. The study constructed a mouse model of heart failure after myocardial infarction using the method of left anterior descending coronary artery ligation. The efficacy of Jiming Powder was evaluated from multiple angles, including ultrasound imaging, hematoxylin-eosin(HE) staining, Masson staining, Sirius Red staining, and serum myocardial enzyme spectrum detection. Western blot analysis was performed to detect key proteins involved in ventricular remodeling, including transforming growth factor-β1(TGF-β1), α-smooth muscle actin(α-SMA), wingless-type MMTV integration site family member 3a(Wnt3a), β-catenin, matrix metallopeptidase 2(MMP2), matrix metallopeptidase 3(MMP3), TIMP metallopeptidase inhibitor 1(TIMP1), and TIMP metallopeptidase inhibitor 2(TIMP2). The results showed that compared with the model group, the high and low-dose Jiming Powder significantly reduced the left ventricular internal diameter in systole(LVID;s) and diastole(LVID;d), increased the left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), effectively improved cardiac function in mice after myocardial infarction, and effectively reduced the levels of myocardial injury markers such as creatine kinase(CK), creatine kinase isoenzyme(CK-MB), and lactic dehydrogenase(LDH), thus protecting ischemic myocardium. HE staining showed that Jiming Powder could attenuate myocardial inflammatory cell infiltration after myocardial infarction. Masson and Sirius Red staining demonstrated that Jiming Powder effectively inhibited myocardial fibrosis, reduced the collagen Ⅰ/Ⅲ ratio in myocardial tissues, and improved collagen remodeling after myocardial infarction. Western blot results showed that Jiming Powder reduced the expression of TGF-β1, α-SMA, Wnt3a, and β-catenin, decreased the levels of MMP2, MMP3, and TIMP2, and increased the level of TIMP1, suggesting its role in inhibiting cardiac fibroblast transformation, reducing extracellular matrix metabolism in myocardial cells, and lowering collagen Ⅰ and α-SMA content, thus exerting an anti-myocardial fibrosis effect after myocardial infarction. This study revealed the role of Jiming Powder in improving ventricular remodeling and treating myocardial infarction, laying the foundation for further research on the pharmacological effect of Jiming Powder.
Mice ; Animals ; Transforming Growth Factor beta1/metabolism* ; Matrix Metalloproteinase 2/metabolism* ; beta Catenin/metabolism* ; Matrix Metalloproteinase 3/therapeutic use* ; Powders ; Ventricular Remodeling ; Stroke Volume ; Ventricular Function, Left ; Myocardial Infarction/drug therapy* ; Myocardium/pathology* ; Heart Failure/metabolism* ; Collagen/metabolism* ; Creatine Kinase ; Fibrosis

OBJECTIVE: To investigate the correlation of inducible co-stimulatory molecules (ICOS) with mesenteric vascular endothelial- mesenchymal transition (EndMT) and sclerosis in spontaneously hypertensive rats (SHR). METHODS: Twenty 4-week-old WKY rats and 20 SHRs of the same strain were both randomly divided into 4 groups for observation at 4, 6, 10 and 30 weeks of age. ICOS expression frequency in rat spleen CD4⁺T cells was analyzed using flow cytometry, and the expressions of ICOS, VE-cad, α-SMA and Col3 mRNA in rat mesentery were detected by RT-PCR. The distributions of ICOS, IL-17A and TGF-β in rat mesentery were detected by immunohistochemistry. The levels of IL-17A and TGF-β in rat plasma were measured using ELISA. The morphological changes of rat mesenteric vessels were observed with Masson staining. Spearman or Pearson correlation analyses were used to evaluate the correlation between ICOS expression and the expressions of the markers of vascular EndMT and sclerosis. RESULTS: Compared with the control WKY rats, the SHRs began to show significantly increased systolic blood pressure and ICOS expression frequency on CD4⁺T cells at 6 weeks of age (P < 0.05). In the SHRs, the mRNA and protein expressions of ICOS, α-SMA, Col3, IL-17A and TGF-β in the mesentery were significantly higher than those in control group (P < 0.05), while the mRNA and protein expressions of VE-cad started to reduce significantly at 10 weeks of age (P < 0.05). The plasma levels of IL-17A and TGF-β were significantly increased in SHRs since 6 weeks of age (P < 0.05) with progressive worsening of mesenteric vascular sclerosis (P < 0.05). ICOS mRNA and protein expression levels in the mesenteric tissues of SHRs began to show positive correlations with α-SMA and Col3 expression levels and the severity of vascular sclerosis at 6 weeks of age (P < 0.05) and a negative correlation with VE-cad expression level at 10 weeks (P < 0.05). CONCLUSION ICOS play an important pathogenic role in EndMT and sclerosis of mesenteric vessels in essential hypertension by mediating related immune responses.
Rats ; Animals ; Rats, Inbred SHR ; Rats, Inbred WKY ; Hypertension ; Interleukin-17 ; Sclerosis/pathology* ; Transforming Growth Factor beta ; Mesentery/pathology* ; RNA, Messenger/metabolism* ; Blood Pressure

Treating patients with esophageal squamous cell carcinoma (ESCC) is challenging due to the high chemoresistance. Growth differentiation factor 15 (GDF15) is crucial in the development of various types of tumors and negatively related to the prognosis of ESCC patients according to our previous research. In this study, the link between GDF15 and chemotherapy resistance in ESCC was further explored. The relationship between GDF15 and the chemotherapy response was investigated through in vitro and in vivo studies. ESCC patients with high levels of GDF15 expression showed an inferior chemotherapeutic response. GDF15 improved the tolerance of ESCC cell lines to low-dose cisplatin by regulating AKT phosphorylation via TGFBR2. Through an in vivo study, we further validated that the anti-GDF15 antibody improved the tumor inhibition effect of cisplatin. Metabolomics showed that GDF15 could alter cellular metabolism and enhance the expression of UGT1A. AKT and TGFBR2 inhibition resulted in the reversal of the GDF15-induced expression of UGT1A, indicating that TGFBR2-AKT pathway-dependent metabolic pathways were involved in the resistance of ESCC cells to cisplatin. The present investigation suggests that a high level of GDF15 expression leads to ESCC chemoresistance and that GDF15 can be targeted during chemotherapy, resulting in beneficial therapeutic outcomes.
Humans ; Esophageal Squamous Cell Carcinoma/drug therapy* ; Cisplatin/metabolism* ; Esophageal Neoplasms/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Carcinoma, Squamous Cell/genetics* ; Growth Differentiation Factor 15/therapeutic use* ; Receptor, Transforming Growth Factor-beta Type II/therapeutic use* ; Cell Line, Tumor ; Cell Proliferation ; Gene Expression Regulation, Neoplastic

OBJECTIVE: Huangqi Decoction (HQD), a classical traditional Chinese medicine formula, has been used as a valid treatment for alleviating liver fibrosis; however, the underlying molecular mechanism is still unknown. Although our previous studies showed that microRNA-663a (miR-663a) suppresses the proliferation and activation of hepatic stellate cells (HSCs) and the transforming growth factor-β/small mothers against decapentaplegic (TGF-β/Smad) pathway, whether long noncoding RNAs (lncRNAs) are involved in HSC activation via the miR-663a/TGF-β/Smad signaling pathway has not yet reported. The present study aimed to investigate the roles of lncRNA lnc-C18orf26-1 in the activation of HSCs and the mechanism by which HQD inhibits hepatic fibrosis. METHODS: The expression levels of lnc-C18orf26-1, miR-663a and related genes were measured by quantitative reverse transcription-polymerase chain reaction. HSCs were transfected with the miR-663a mimic or inhibitor and lnc-C18orf26-1 small interfering RNAs. The water-soluble tetrazolium salt-1 assay was used to assess the proliferation rate of HSCs. Changes in lncRNA expression were evaluated in miR-663a-overexpressing HSCs by using microarray to identify miR-663a-regulated lncRNAs. RNA hybrid was used to predict the potential miR-663a binding sites on lncRNAs. Luciferase reporter assays further confirmed the interaction between miR-663a and the lncRNA. The expression levels of collagen α-2(I) chain (COL1A2), α-smooth muscle actin (α-SMA) and TGF-β/Smad signaling pathway-related proteins were determined using Western blotting. RESULTS: Lnc-C18orf26-1 was upregulated in TGF-β1-activated HSCs and competitively bound to miR-663a. Knockdown of lnc-C18orf26-1 inhibited HSC proliferation and activation, downregulated TGF-β1-stimulated α-SMA and COL1A2 expression, and inhibited the TGF-β1/Smad signaling pathway. HQD suppressed the proliferation and activation of HSCs. HQD increased miR-663a expression and decreased lnc-C18orf26-1 expression in HSCs. Further studies showed that HQD inhibited the expression of COL1A2, α-SMA, TGF-β1, TGF-β type I receptor (TGF-βRI) and phosphorylated Smad2 (p-Smad2) in HSCs, and these effects were reversed by miR-663a inhibitor treatment. CONCLUSION Our study identified lnc-C18orf26-1 and miR-663a as promising therapeutic targets for hepatic fibrosis. HQD inhibits HSC proliferation and activation at least partially by regulating the lnc-C18orf26-1/miR-663a/TGF-β1/TGF-βRI/p-Smad2 axis.
Humans ; Transforming Growth Factor beta/pharmacology* ; Transforming Growth Factor beta1/metabolism* ; RNA, Long Noncoding/pharmacology* ; Drugs, Chinese Herbal/pharmacology* ; MicroRNAs/genetics* ; Hepatic Stellate Cells/pathology* ; Liver Cirrhosis/metabolism* ; Cell Proliferation ; Transforming Growth Factors/pharmacology*

OBJECTIVES: To investigate the effect of recombinant human fibroblast growth factor 21 (rhFGF21) on the proliferation and mineralization of cementoblasts and its mechanism. METHODS: Hematoxylin eosin, immunohistochemical staining, and immunofluorescence were used to detect the expression and distribution of fibroblast growth factor 21 (FGF21) in rat periodontal tissues and cementoblasts (OCCM-30), separately. Cell Counting Kit-8 was used to detect the proliferation of OCCM-30 under treatment with rhFGF21. Alkaline phosphatase staining and Alizarin Red staining were used to detect the mineralization state of OCCM-30 after 3 and 7 days of mineralization induction. The transcription and protein expression of the osteogenic-related genes Runx2 and Osterix were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot analysis. The expression levels of genes of transforming growth factor β (TGFβ)/bone morphogenetic protein (BMP) signaling pathway in OCCM-30 were detected through PCR array analysis. RESULTS: FGF21 was expressed in rat periodontal tissues and OCCM-30. Although rhFGF21 had no significant effect on the proliferation of OCCM-30, treatment with 50 ng/mL rhFGF21 could promote the mineralization of OCCM-30 cells after 7 days of mineralization induction. The transcriptional levels of Runx2 and Osterix increased significantly at 3 days of mineralization induction and decreased at 5 days of mineralization induction. Western blot analysis showed that the protein expression levels of Runx2 and Osterix increased during mineralization induction. rhFGF21 up-regulated Bmpr1b protein expression in cells. CONCLUSIONS rhFGF21 can promote the mineralization ability of OCCM-30. This effect is related to the activation of the TGFβ/BMP signaling pathway.
Humans ; Rats ; Animals ; Dental Cementum ; Core Binding Factor Alpha 1 Subunit/metabolism* ; Cell Differentiation ; Bone Morphogenetic Proteins/metabolism* ; Transforming Growth Factor beta/pharmacology*

Diabetic kidney disease is an important microvascular complication of diabetes and the leading cause of end-stage renal disease. Its pathological characteristics mainly include epithelial mesenchymal transition(EMT) in glomerulus, podocyte apoptosis and autophagy, and damage of glomerular filtration barrier. Transforming growth factor-β(TGF-β)/Smad signaling pathway is specifically regulated by a variety of mechanisms, and is a classic pathway involved in physiological activities such as apoptosis, proliferation and differentiation. At present, many studies have found that TGF-β/Smad signaling pathway plays a key role in the pathogenesis of diabetic kidney disease. Traditional Chinese medicine has significant advantages in the treatment of diabetic kidney disease for its multi-component, multi-target and multi-pathway characteristics, and some traditional Chinese medicine extracts, traditional Chinese medicines and traditional Chinese medicine compound prescription improve the renal injury of diabetic kidney disease by regulating TGF-β/Smad signaling pathway. This study clarified the mechanism of TGF-β/Smad signaling pathway in diabetic kidney disease by expounding the relationship between the key targets of the pathway and diabetic kidney disease, and summarized the research progress of traditional Chinese medicine in the treatment of diabetic kidney disease by interfering with TGF-β/Smad signaling pathway in recent years, to provide reference for drug research and clinical treatment of diabetic kidney disease in the future.
Humans ; Diabetic Nephropathies/genetics* ; Medicine, Chinese Traditional ; Kidney/pathology* ; Transforming Growth Factor beta/metabolism* ; Signal Transduction ; Epithelial-Mesenchymal Transition ; Smad Proteins/metabolism* ; Transforming Growth Factor beta1/metabolism* ; Diabetes Mellitus/genetics*

Objective To investigate the effect of insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) on the proliferation, migration and tumor immune microenvironment of colorectal cancer cells and its possible molecular mechanism. Methods The Cancer Genome Atlas (TCGA) database was used to analyze the expression levels of IGF2BP2 and MYC in colorectal cancer and adjacent tissues. The expression of IGF2BP2 in HCT-116 and SW480 human colorectal cancer cells was silenced by RNA interference (RNAi), and the silencing effect was detected by quantitative real-time PCR. After knocking down IGF2BP2, colony formation assay, CCK-8 assay and 5-ethynyl-2'-deoxyuridine (EdU) assay were employed to detect cell colony formation and proliferation ability. Transwell^TM assay was used to detect cell migration ability. Quantitative real-time PCR was used to detect the mRNA expression of IGF2BP2, MYC, tumor necrosis factor-α (TNF-α), transforming growth factor-β (TGF-β) and interleukin-10 (IL-10). The protein expression of IGF2BP2 and MYC was detected by western blot. The binding ability of IGF2BP2 and MYC in HCT-116 cells was detected by quantitative real-time PCR after RNA immunoprecipitation. Results The results of TCGA database showed that the expression of IGF2BP2 and MYC in colorectal cancer tissues was significantly higher than that in adjacent tissues, and the survival time of colorectal cancer patients with high expression of IGF2BP2 was shorter. After silencing IGF2BP2, the viability, proliferation and migration of HCT-116 and SW480 cells were decreased. The mRNA expression of MYC, TGF-β and IL-10 in IGF2BP2 knockdown group was significantly decreased, while the expression of TNF-α mRNA was increased. The expression of MYC protein and the stability of MYC mRNA were significantly decreased. RIP-qPCR results showed that IGF2BP2 could bind to MYC mRNA. Conclusion Knockdown of IGF2BP2 inhibits colorectal cancer cell proliferation, migration and promotes tumor immunity by down-regulating MYC expression.
Humans ; Cell Line, Tumor ; Cell Movement/genetics* ; Cell Proliferation/genetics* ; Colorectal Neoplasms/metabolism* ; Gene Expression Regulation, Neoplastic ; Interleukin-10/metabolism* ; RNA, Messenger ; RNA-Binding Proteins/metabolism* ; Transforming Growth Factor beta/genetics* ; Tumor Microenvironment/immunology* ; Tumor Necrosis Factor-alpha/metabolism* ; Proto-Oncogene Proteins c-myc/metabolism*

OBJECTIVE: To investigate the expression and significance of regulatory T cells (Tregs), FoxP3 and transforming growth factor-β (TGF-β) in different phase of chronic myeloid leukemia (CML). METHODS: Peripheral blood of 73 CML patients in Department of Hematology, Heze Municipal Hospital from March 2018 to March 2021 were collected. According to patient's period in CML, they were divided into ND CML group (newly diagnosed), CP CML group (chronic period), and BP CML group (blast phase). The percentage of Tregs, expression level of FoxP3 mRNA and TGF-β were detected by flow cytometry, RT-qPCR, and ELISA, respecitively. The roles of above indices in clinical pathogenesis of patients with CML were analyzed. RESULTS: The proportion of Treg in the ND CML group was slightly higher than the CP CML group, but the difference was not statistically significant (P =0.695), while the BP CML group was significantly higher than the other two groups (P =0.008, P <0.001). The expression levels of FoxP3 mRNA in ND CML group, CP CML group and BP CML group were 11.61±2.21, 6.46±1.35 and 8.54±2.13, respectively. Significant difference in FoxP3 mRNA levels was observed among patients in different phases of CML (F =55.199, P <0.001). The expression levels of FoxP3 mRNA both in ND CML group and BP CML group were significantly higher than that in CP CML group (P <0.001), and the ND CML group was the highest (P <0.001). However, the expression levels of TGF-β in different phases of CML showed no statistical differences (H =0.634, P =0.728). CONCLUSION The abnormal distribution of Treg subset in different phases of CML and the significant increase of the expression level of FoxP3 mRNA in the new onset and blast phase of CML suggest that Tregs may promote the occurrence and progression of CML through immune regulation.
Humans ; Blast Crisis/metabolism* ; Forkhead Transcription Factors/metabolism* ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics* ; RNA, Messenger/metabolism* ; T-Lymphocytes, Regulatory/metabolism* ; Transforming Growth Factor beta/metabolism*

OBJECTIVE: To observe the effect of amygdalin on liver fibrosis in a liver fibrosis mouse model, and the underlying mechanisms were partly dissected in vivo and in vitro. METHODS: Thirty-two male mice were randomly divided into 4 groups, including control, model, low- and high-dose amygdalin-treated groups, 8 mice in each group. Except the control group, mice in the other groups were injected intraperitoneally with 10% carbon tetrachloride (CCl₄)-olive oil solution 3 times a week for 6 weeks to induce liver fibrosis. At the first 3 weeks, amygdalin (1.35 and 2.7 mg/kg body weight) were administered by gavage once a day. Mice in the control group received equal quantities of subcutaneous olive oil and intragastric water from the fourth week. At the end of 6 weeks, liver tissue samples were harvested to detect the content of hydroxyproline (Hyp). Hematoxylin and eosin and Sirius red staining were used to observe the inflammation and fibrosis of liver tissue. The expressions of collagen I (Col-I), alpha-smooth muscle actin (α-SMA), CD31 and transforming growth factor β (TGF-β)/Smad signaling pathway were observed by immunohistochemistry, quantitative real-time polymerase chain reaction and Western blot, respectively. The activation models of hepatic stellate cells, JS-1 and LX-2 cells induced by TGF-β1 were used in vitro with or without different concentrations of amygdalin (0.1, 1, 10 µmol/L). LSECs. The effect of different concentrations of amygdalin on the expressions of liver sinusoidal endothelial cells (LSECs) dedifferentiation markers CD31 and CD44 were observed. RESULTS: High-dose of amygdalin significantly reduced the Hyp content and percentage of collagen positive area, and decreased the mRNA and protein expressions of Col-I, α-SMA, CD31 and p-Smad2/3 in liver tissues of mice compared to the model group (P<0.01). Amygdalin down-regulated the expressions of Col-I and α-SMA in JS-1 and LX-2 cells, and TGFβ R1, TGFβ R2 and p-Smad2/3 in LX-2 cells compared to the model group (P<0.05 or P<0.01). Moreover, 1 and 10 µmol/L amygdalin inhibited the mRNA and protein expressions of CD31 in LSECs and increased CD44 expression compared to the model group (P<0.05 or P<0.01). CONCLUSIONS Amygdalin can dramatically alleviate liver fibrosis induced by CCl₄ in mice and inhibit TGF-β/Smad signaling pathway, consequently suppressing HSCs activation and LSECs dedifferentiation to improve angiogenesis.
Rats ; Male ; Mice ; Animals ; Transforming Growth Factor beta/metabolism* ; Amygdalin/therapeutic use* ; Endothelial Cells/metabolism* ; Olive Oil/therapeutic use* ; Rats, Wistar ; Smad Proteins/metabolism* ; Liver Cirrhosis/metabolism* ; Liver ; Transforming Growth Factor beta1/metabolism* ; Signal Transduction ; Collagen Type I/metabolism* ; Carbon Tetrachloride ; Hepatic Stellate Cells