[Banxia Xiexin Decoction inhibiting colitis-associated colorectal cancer infected with Fusobacterium nucleatum by regulating Wnt/ß-catenin pathway].

This paper investigates the intervention effect and mechanism of Banxia Xiexin Decoction(BXD) on colitis-associated colorectal cancer(CAC) infected with Fusobacterium nucleatum(Fn). C57BL/6 mice were randomly divided into a control group, Fn group, CAC group [azoxymethane(AOM)/dextran sulfate sodium salt(DSS)](AOM/DSS), model group, and BXD group. Except for the control and AOM/DSS groups, the mice in the other groups were orally administered with Fn suspension twice a week. The AOM/DSS group, model group, and BXD group were also injected with a single dose of 10 mg·kg~(-1) AOM combined with three cycles of 2.5% DSS taken intragastrically. The BXD group received oral administration of BXD starting from the second cycle until the end of the experiment. The general condition and weight changes of the mice were monitored during the experiment, and the disease activity index(DAI) was calculated. At the end of the experiment, the colon length and weight of the mice in each group were compared. Hematoxylin-eosin(HE) staining was used to observe the pathological changes in the colon tissue. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of interleukin(IL)-2, IL-4, and IL-6 inflammatory factors in the serum. Immunohistochemistry(IHC) was used to detect the expression of Ki67, E-cadherin, and ß-catenin in the colon tissue. Western blot was used to detect the protein content of Wnt3a, ß-catenin, E-cadherin, annexin A1, cyclin D1, and glycogen synthase kinase-3ß(GSK-3ß) in the colon tissue. The results showed that compared with the control group, the Fn group had no significant lesions. The mice in the AOM/DSS group and model group had decreased body weight, increased DAI scores, significantly increased colon weight, and significantly shortened colon length, with more significant lesions in the model group. At the same time, the colon histology of the model group showed more severe adenomas, inflammatory infiltration, and cellular dysplasia. The levels of IL-4 and IL-6 in the serum were significantly increased, while the IL-2 content was significantly decreased. The IHC results showed low expression of E-cadherin and high expression of Ki67 and ß-catenin in the model group, with a decreased protein content of E-cadherin and GSK-3ß and an increased protein content of Wnt3a, ß-catenin, annexin A1, and cyclin D1. After intervention with BXD, the body weight of the mice increased; the DAI score decreased; the colon length increased, and the tumor decreased. The histopathology showed reduced tumor proliferation and reduced inflammatory infiltration. The levels of IL-6 and IL-4 in the serum were significantly decreased, while the IL-2 content was increased. Meanwhile, the expression of E-cadherin was upregulated, and that of Ki67 and ß-catenin was downregulated. The protein content of E-cadherin and GSK-3ß increased, while that of Wnt3a, ß-catenin, annexin A1, and cyclin D1 decreased. In conclusion, BXD can inhibit CAC infected with Fn, and its potential mechanism may be related to the inhibition of Fn binding to E-cadherin, the decrease in annexin A1 protein level, and the regulation of the Wnt/ß-catenin pathway.

Roles of ß-catenin in innate immune process and regulating intestinal flora in Qi river crucian carp (Carassius auratus).

In mammals, ß-catenin participates in innate immune process through interaction with NF-κB signaling pathway. However, its role in teleost immune processes remains largely unknown. We aimed to clarify the function of ß-catenin in the natural defense mechanism of Qi river crucian carp (Carassius auratus). ß-catenin exhibited a ubiquitous expression pattern in adult fish, as indicated by real-time PCR analysis. Following lipopolysaccharide (LPS), Polyinosinic-polycytidylic acid (polyI: C) and Aeromonas hydrophila (A. hydrophila) challenges, ß-catenin increased in gill, intestine, liver and kidney, indicating that ß-catenin likely plays a pivotal role in the immune response against pathogen infiltration. Inhibition of the ß-catenin pathway using FH535, an inhibitor of Wnt/ß-catenin pathway, resulting in pathological damage of the gill, intestine, liver and kidney, significant decrease of innate immune factors (C3, defb3, LYZ-C, INF-γ), upregulation of inflammatory factors (NF-κB, TNF-α, IL-1, IL-8), and downregulation of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) activities, increase of Malondialdehyde (MDA) content. Following A. hydrophila invasion, the mortality rate in the FH535 treatment group exceeded that of the control group. In addition, the diversity of intestinal microflora decreased and the community structure was uneven after FH535 treatment. In summary, our findings strongly suggest that ß-catenin plays a vital role in combating pathogen invasion and regulating intestinal flora in Qi river crucian carp.

Polyphyllin I enhances tumor necrosis factor-related apoptosis-inducing ligand-induced inhibition of human osteosarcoma cell growth downregulating the Wnt/ß-catenin pathway.

OBJECTIVE: To investigate the synergistic effects of polyphyllin I (PPI) combined with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on the growth of osteosarcoma cells through downregulating the Wnt/ß-catenin signaling pathway. METHODS: Cell viability, apoptosis and cell cycle distribution were examined using cell counting kit-8 and flow cytometry assays. The morphology of cancer cells was observed with inverted phase contrast microscope. The migration and invasion abilities were examined by xCELLigence real time cell analysis DP system and transwell assays. The expressions of poly (adenosine diphosphate-ribose) polymerase, C-Myc, Cyclin B1, cyclin-dependent kinases 1, N-cadherin, Vimentin, Active-ß-catenin, ß-catenin, p-glycogen synthase kinase 3ß (GSK-3ß) and GSK-3ß were determined by Western blotting assay. RESULTS: PPI sensitized TRAIL-induced decrease of viability, migration and invasion, as well as increase of apoptosis and cell cycle arrest of MG-63 and U-2 OS osteosarcoma cells. The synergistic effect of PPI with TRAIL in inhibiting the growth of osteosarcoma cells was at least partially realized through the inactivation of Wnt/ß-catenin signaling pathway. CONCLUSION: The combination of PPI and TRAIL is potentially a novel treatment strategy of osteosarcoma.

Conditional Deletion of ß-Catenin in the Mediobasal Hypothalamus Impairs Adaptive Energy Expenditure in Response to High-Fat Diet and Exacerbates Diet-Induced Obesity.

ß-Catenin is a bifunctional molecule that is an effector of the wingless-related integration site (Wnt) signaling to control gene expression and contributes to the regulation of cytoskeleton and neurotransmitter vesicle trafficking. In its former role, ß-catenin binds transcription factor 7-like 2 (TCF7L2), which shows strong genetic associations with the pathogenesis of obesity and type-2 diabetes. Here, we sought to determine whether ß-catenin plays a role in the neuroendocrine regulation of body weight and glucose homeostasis. Bilateral injections of adeno-associated virus type-2 (AAV2)-mCherry-Cre were placed into the arcuate nucleus of adult male and female ß-catenin flox mice, to specifically delete ß-catenin expression in the mediobasal hypothalamus (MBH-ß-cat KO). Metabolic parameters were then monitored under conditions of low-fat (LFD) and high-fat diet (HFD). On LFD, MBH-ß-cat KO mice showed minimal metabolic disturbances, but on HFD, despite having only a small difference in weekly caloric intake, the MBH-ß-cat KO mice were significantly heavier than the control mice in both sexes (p < 0.05). This deficit seemed to be due to a failure to show an adaptive increase in energy expenditure seen in controls, which served to offset the increased calories by HFD. Both male and female MBH-ß-cat KO mice were highly glucose intolerant when on HFD and displayed a significant reduction in both leptin and insulin sensitivity compared with controls. This study highlights a critical role for ß-catenin in the hypothalamic circuits regulating body weight and glucose homeostasis and reveals potential mechanisms by which genetic variation in this pathway could impact on development of metabolic disease.

Efficacy of Yisui granule on myelodysplastic syndromes in SKM-1 mouse xenograft model through suppressing Wnt/ß-catenin signaling pathway.

OBJECTIVE: To unmask the underlying mechanisms of Yisui granule (, YSG) for the treatment of Myelodysplastic syndromes (MDS). METHODS: Our study used an SKM-1 mouse xenograft model of MDS to explore the anti-tumor potential of YSG and its safety, assess its effect on overall survival (OS), and evaluate whether its mechanism is associated with the demethylation of the secreted frizzled related protein 5 (sFRP5) gene and suppressing Wnt/ß-catenin pathway. Bisulfite amplicon sequencing was applied to detect the level of methylation of the sFRP5 gene; western blotting, immunofluorescence staining, and real-time Polymerase Chain Reaction were performed to detect DNA methyltransferase 1 (DNMT1), sFRP5, and other Wnt/ß-catenin pathway-related mRNA and protein expression. RESULTS: The results showed that high-dosage YSG exerted an anti-tumor effect similar to that of decitabine, improved OS, and reduced long-term adverse effects in the long term. Mechanically, YSG reduced the expression of DNMT1 methyltransferase, decreased the methylation, and increased the expression of the Wnt/ß-catenin pathway antagonist-sFRP5. Furthermore, components of the Wnt/ß-catenin pathway, including Wnt3a, ß-catenin, c-Myc, and cyclinD1, were down-regulated in response to YSG, suggesting that YSG could treat MDS by demethylating the sFRP5 gene and suppressing the Wnt/ß-catenin pathway. CONCLUSIONS: Our findings demonstrated that YSG could be used alone or in combination with decitabine to improve outcomes in the MDS animal model, providing an alternative solution for treating MDS.

[Blaps rynchopetera combined with cyclophosphamide affects proliferation and apoptosis of lung cancer cells via Wnt/ß-catenin signaling pathway].

This study aims to investigate the effects of Blaps rynchopetera Fairmaire and/or cyclophosphamide on the proliferation and apoptosis of lung cancer cells and decipher the underlying mechanism. B. rynchopetera and cyclophosphamide-containing serum and blank serum were prepared from SD rats. Cell counting kit-8(CCK-8) assay was employed to examine the proliferation of lung cancer cell lines A549 and Lewis treated with corresponding agents. The Jin's formula method was used to evaluate the combined effect of the two drugs. According to the evaluation results, appropriate drug concentrations and lung cancer cell line were selected for subsequent experiments, which included control, B. rynchopetera, cyclophosphamide, B. rynchopetera + cyclophosphamide, and B. rynchopetera + Wnt/ß-catenin pathway agonist lithium chloride(LiCl) groups. Immunocytochemistry was employed to measure the expression of proliferation-related proteins in Lewis cells after drug interventions. Flow cytometry was employed to determine the cell cycle and apoptosis. The expression levels of proliferating cell nuclear antigen(PCNA), cyclinD1, B-cell lymphoma 2(Bcl-2), Bcl-2-assiocated X protein(Bax), Wnt1, and ß-catenin were determined by Western blot. The results showed that B. rynchopetera and/or cyclophosphamide significantly inhibited the proliferation of A549 and Lewis cells. Compared with B. rynchopetera alone, the combination increased the inhibition rate on cell proliferation. The combination of B. rynchopetera and cyclophosphamide demonstrated a synergistic effect according to Jin's formula-based evaluation. Compared with the control group, the B. rynchopetera, cyclophosphamide, and B. rynchopetera + cyclophosphamide groups showed increased proportion of Lewis cells in G_0/G_1 phase, increased apoptosis rate, up-regulated expression of Bax, and down-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. Compared with the cyclophosphamide group, the combination group showed increased proportion of cells in G_0/G_1 phase, increased apoptosis rate, up-regulated expression of Bax, and down-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. Compared with the B. rynchopetera group, the B. rynchopetera + LiCl group had deceased proportion of cells in G_0/G_1 phase, decreased apoptosis rate, down-regulated expression of Bax, and up-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. The results indicated that B. rynchopetera could inhibit the proliferation, arrest the cell cycle, and induce the apoptosis of lung cancer cells by inhibiting the Wnt/ß-catenin signaling pathway. Moreover, B. rynchopetera had a synergistic effect with cyclophosphamide.

Mutant GGGGCC RNA prevents YY1 from binding to Fuzzy promoter which stimulates Wnt/ß-catenin pathway in C9ALS/FTD.

The GGGGCC hexanucleotide repeat expansion mutation in the chromosome 9 open reading frame 72 (C9orf72) gene is a major genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). In this study, we demonstrate that the zinc finger (ZF) transcriptional regulator Yin Yang 1 (YY1) binds to the promoter region of the planar cell polarity gene Fuzzy to regulate its transcription. We show that YY1 interacts with GGGGCC repeat RNA via its ZF and that this interaction compromises the binding of YY1 to the FuzzyYY1 promoter sites, resulting in the downregulation of Fuzzy transcription. The decrease in Fuzzy protein expression in turn activates the canonical Wnt/ß-catenin pathway and induces synaptic deficits in C9ALS/FTD neurons. Our findings demonstrate a C9orf72 GGGGCC RNA-initiated perturbation of YY1-Fuzzy transcriptional control that implicates aberrant Wnt/ß-catenin signalling in C9ALS/FTD-associated neurodegeneration. This pathogenic cascade provides a potential new target for disease-modifying therapy.

[Effect of acetylalkannin from Arnebia euchroma on proliferation, migration, and invasion of human melanoma A375 cells].

This study aimed to explore the effect and mechanism of acetylalkannin from Arnebia euchroma on the proliferation, migration, and invasion of human melanoma A375 cells. A375 cells were divided into a blank group, and low-, medium-, and high-dose acetylalkannin groups(0.5, 1.0, and 2.0 µmol·L~(-1)). The MTT assay was used to detect cell proliferation. Cell scratch and transwell migration assays were used to detect cell migration ability, and the transwell invasion assay was used to detect cell invasion ability. Western blot was used to detect the protein expression of migration and invasion-related N-cadherin, vimentin, matrix metalloproteina-se-9(MMP-9), and Wnt/ß-catenin pathway-related Wnt1, Axin2, glycogen synthase kinase-3ß(GSK-3ß), phosphorylated GSK-3ß(p-GSK-3ß), ß-catenin, cell cycle protein D_1(cyclin D_1), and p21. Real-time fluorescence-based quantitative polymerase chain reaction(real-time PCR) was used to detect the mRNA expression of E-cadherin, matrix metalloproteinase-2(MMP-2), N-cadherin, vimentin, ß-catenin, snail-1, and CD44. MTT results showed that the cell inhibition rates in the acetylalkannin groups significantly increased as compared with that in the blank group(P<0.01). The results of cell scratch and transwell assays showed that compared with the blank group, the acetylalkannin groups showed reduced cell migration and invasion, and migration and invasion rates(P<0.05, P<0.01) and weakened horizontal and vertical migration and invasion abilities. Western blot results showed that compared with the blank group, the high-dose acetylalkannin group showed increased expression of Axin2 protein(P<0.05), and decreased expression of N-cadherin, vimentin, MMP-9, Wnt1, p-GSK-3ß, ß-catenin, cyclin D_1, and p21 proteins(P<0.05, P<0.01). The expression of GSK-3ß protein did not change significantly. PCR results showed that the overall trend of MMP-2, N-cadherin, vimentin, ß-catenin, snail-1, and CD44 mRNA expression was down-regulated(P<0.01), and the expression of E-cadherin mRNA increased(P<0.01). Acetylalkannin can inhibit the proliferation, migration, and invasion of human melanoma A375 cells, and its mechanism of action may be related to the regulation of Wnt/ß-catenin signaling pathway.

Jiedu Recipe, a compound Chinese herbal medicine, inhibits cancer stemness in hepatocellular carcinoma via Wnt/ß-catenin pathway under hypoxia.

OBJECTIVE: Jiedu Recipe (JR), a Chinese herbal remedy, has been shown to prolong overall survival time and decrease recurrence and metastasis rates in patients with hepatocellular carcinoma (HCC). This work investigated the mechanism of JR in HCC treatment. METHODS: The chemical constituents of JR were detected using liquid chromatography-mass spectrometry. The potential anti-HCC mechanism of JR was screened using network pharmacology and messenger ribonucleic acid (mRNA) microarray chip assay, followed by experimental validation in human HCC cells (SMMC-7721 and Huh7) in vitro and a nude mouse subcutaneous transplantation model of HCC in vivo. HCC cell characteristics of proliferation, migration and invasion under hypoxic setting were investigated using thiazolyl blue tetrazolium bromide, wound healing and Transwell assays, respectively. Image-iT™ Hypoxia Reagent was added to reveal hypoxic conditions. Stem cell sphere formation assay was used to detect the stemness. Epithelial-mesenchymal transition (EMT) markers like E-cadherin, vimentin and α-smooth muscle actin, and pluripotent transcription factors including nanog homeobox, octamer-binding transcription factor 4, and sex-determining region Y box protein 2 were analyzed using Western blotting and real-time polymerase chain reaction. Western blot was performed to ascertain the anti-HCC effect of JR under hypoxia involving the Wnt/ß-catenin pathway. RESULTS: According to network pharmacology and mRNA microarray chip analysis, JR may potentially act on hypoxia and inhibit the Wnt/ß-catenin pathway. In vitro and in vivo experiments showed that JR significantly decreased hypoxia, and suppressed HCC cell features of proliferation, migration and invasion; furthermore, the hypoxia-induced increases in EMT and stemness marker expression in HCC cells were inhibited by JR. Results based on the co-administration of JR and an agonist (LiCl) or inhibitor (IWR-1-endo) verified that JR suppressed HCC cancer stem-like properties under hypoxia by blocking the Wnt/ß-catenin pathway. CONCLUSION: JR exerts potent anti-HCC effects by inhibiting cancer stemness via abating the Wnt/ß-catenin pathway under hypoxic conditions. Please cite this article as: Guo BJ, Ruan Y, Wang YJ, Xiao CL, Zhong ZP, Cheng BB, Du J, Li B, Gu W, Yin ZF. Jiedu Recipe, a compound Chinese herbal medicine, inhibits cancer stemness in hepatocellular carcinoma via Wnt/ß-catenin pathway under hypoxia. J Integr Med. 2023; 21(5): 474-486.

CTNNB1 p.D32A (c.95A > C) somatic mutation in stage I grade 1 endometrioid endometrial carcinoma with lung metastasis: a case report.

BACKGROUND: Most endometrial cancers are of low histological grade and uterine-confined, with a high 5-year survival rate. However, a small subset of women with low-grade and early-stage endometrioid endometrial cancer experience recurrence and death; thus, a more precise risk-stratification is needed. CASE PRESENTATION: A 29-year-old woman presented with abnormal vaginal bleeding and was diagnosed with FIGO grade 1 endometrioid endometrial carcinoma by curettage. Comprehensive cancer staging including pelvic and para-aortic lymphadenectomy was then performed. Postoperative pathological findings suggested an FIGO grade 1 endometrioid endometrial carcinoma infiltrating the superficial muscle layer. The patient did not receive adjuvant therapy. After 4 years of follow-up, the patient returned to our institution with lung metastasis. She underwent thoracoscopic resection of the affected lobes, followed by six cycles of combined chemotherapy of paclitaxel and carboplatin. Next-generation sequencing showed that the primary and lung metastatic tumors shared 4 mutations: PTEN (p.P248Lfs*8), CTNNB1 (p.D32A), BCOR (p.N1425S) and CBL (p.S439N). Immunohistochemistry revealed nuclear location of ß-catenin in the primary and lung metastatic tumor samples, indicating abnormal activation of ß-catenin. CONCLUSION: CTNNB1p.D32A (c.95A > C) mutation may be related to lung metastasis in this patient with low-grade early-stage endometrioid endometrial carcinoma.

eIF3a sustains non-small cell lung cancer stem cell-like properties by promoting YY1-mediated transcriptional activation of ß-catenin.

Cancer stem cells (CSCs) are the leading cause of recurrence and poor prognosis in non-small cell lung cancer (NSCLC). Eukaryotic translation initiation factor 3a (eIF3a) participates in many tumor development processes, such as metastasis, therapy resistance, and glycolysis, all of which are closely associated with the presence of CSCs. However, whether eIF3a maintains NSCLC-CSC-like properties remains to be elucidated. In this study, eIF3a was highly expressed in lung cancer tissues and was linked to poor prognosis. eIF3a was also highly expressed in CSC-enriched spheres compared with adherent monolayer cells. Moreover, eIF3a is required for NSCLC stem cell-like traits maintenance in vitro and in vivo. Mechanistically, eIF3a activates the Wnt/ß-catenin signaling pathway, promoting the transcription of cancer stem cell markers. Specifically, eIF3a promotes the transcriptional activation of ß-catenin and mediates its nuclear accumulation to form a complex with T cell factor 4 (TCF4). However, eIF3a has no significant effect on protein stability and translation. Proteomics analysis revealed that the candidate transcription factor, Yin Yang 1 (YY1), mediates the activated effect of eIF3a on ß-catenin. Overall, the findings of this study implied that eIF3a contributes to the maintenance of NSCLC stem cell-like characteristics through the Wnt/ß-catenin pathway. eIF3a is a potential target for the treatment and prognosis of NSCLC.

The effect of 1,25-dihydroxyvitamin D3 on the Wnt signaling pathway in bovine intestinal epithelial cells is mediated by the DKK2 (dickkopf2) Wnt antagonist.

The Wnt/ß-catenin signaling pathway is aberrantly activated in most colorectal cancers. High-dose 1,25(OH)2D3 has anticancer effect by regulating Wnt signal pathway. However, it is not clear whether high-dose of 1,25(OH)2D3 have an effect on normal cells. The aim of the present study was to investigate the mechanism of high-dose 1,25(OH)2D3 on the Wnt signaling pathway in bovine intestinal epithelial cells. The potential mechanism of action was investigated after knockdown and overexpression of the Wnt pathway inhibitor, DKK2, in intestinal epithelial cells by observing the effects of 1,25(OH)2D3 on proliferation, apoptosis, pluripotency and the expression of genes related to the Wnt/ß-catenin signaling pathway. In the present study, we introduced the method of isolation and culture of primary bovine intestinal epithelial cells. After cells were treated with 50 ng/mL 1,25(OH)2D3 or DMSO for 48 h, total RNA was extracted, and six differentially expressed genes, including SERPINF1, SFRP2, SFRP4, FZD2, WISP1 and DKK2 were identified by transcriptome sequencing, which were related to Wnt signaling pathway. To further explore the mechanism of 1,25(OH)2D3 on the Wnt/ß-catenin signaling pathway, we constructed knockdown and overexpression plasmids of DKK2. After transfecting these plasmids into bovine intestinal epithelial cells, we measured the expression of DKK2 mRNA and protein through GFP expression, qRT-PCR and western blot analyses to verify the transfection efficiency. In addition, the CCK-8 assay was used to detect the cell proliferation rate after transfection. Subsequently, the transfected cells were treated with 1,25(OH)2D3 for 48 h, and the proliferation- (Ki67 and PCNA), apoptosis- (Bcl-2, p53, casp3 and casp8), pluripotency- (Bmi-1, Lrig1, KRT19 and TUFT1) and Wnt/ ß-catenin signaling pathway- related genes (LGR5, DKK2, VDR, ß- Catenin, SFRP2, WISP1 and FZD2) were detected by qRT-PCR and western blot analyses. Our results showed that the expression trend of some genes in bovine intestinal epithelial cells under high-dose 1,25(OH)2D3 was consistent with the sequencing results, including SFRP2 (P < 0.001), SFRP4 (P < 0.05), FZD2 (P < 0.01), WISP1 (P < 0.001) and DKK2 (P < 0.001). In addition, knockdown of DKK2 inhibited cell proliferation (P < 0.01), but DKK2 overexpression promoted cell proliferation (P < 0.01). Compared to the control group, 1,25(OH)2D3 promoted the expression of Wnt/ß-catenin signaling pathway-related proteins in bovine intestinal epithelium, thus maintaining intestinal homeostasis in normal intestinal epithelium. In addition, knockdown and overexpression of DKK2 indicated that 1,25(OH)2D3 weakened the inhibitory effect of DKK2 on the Wnt/ß-catenin signaling pathway. Together, these results suggest that high-dose 1,25(OH)2D3 has no killing effect on normal intestinal epithelial cells and regulates Wnt/ß-catenin signaling pathway through DKK2.

Papillary thyroid carcinoma with desmoid-type fibromatosis: the clinicopathological features with characteristic imaging and molecular correlation requiring comprehensive treatment.

The pathogenesis and treatment of papillary thyroid cancer with desmoid-type fibromatosis (PTC-DTF), a rare subtype of papillary thyroid carcinoma characterized by a mixed epithelial-mesenchymal structure, are still ill-defined. Previous reports on PTC-DTF have had limited follow-up and recurrence has been rarely reported. To better understand this condition, we conducted a thorough analysis of five cases of PTC-DTF from our institute, including clinical and pathological examinations, imaging, immunohistochemistry, and molecular analysis. We also reviewed relevant literature. The mean age of the patients was 51.8 years, with three women and two men included in the group. Ultrasound often showed a hypoechogenic and well-defined nodule in the thyroid gland, except for one individual who had distant lung metastases detected by PET-CT. The nodules ranged in width from 0.5 to 5.0 cm and were excised in each case. Following surgery, 131I therapy was used in two cases. The overall number of PTC-DTF cases has risen from the previously reported 55 to 60, with females being the most commonly affected and ranging in age from 19 to 82. Most masses underwent a thyroidectomy, and approximately half of the patients had lymph node metastases. Histologically, PTC-DTFs were composed of a predominant stromal component (65%-90%) and an intervening epithelial component. These spindle cells were arranged in parallel with abundant cytoplasm and vacuole-like nucleus but there wasn't evident atypia. The carcinoma cells were positively stained for CK and TTF-1 by immunohistochemistry, whereas mesenchymal cells were positive for SMA and displayed nuclear immunoreactivity for ß-catenin. BRAF, NRAS, and CTNNB1 mutations were identified in the epithelial and mesenchymal components through molecular testing, respectively. Perhaps because the mesenchyme harbors aberrant nuclear ß-catenin expression, PTC-DTF is more aggressive and prone to invasion and distant recurrence, as shown by our case 2, which is the first case to be reported thus far. PTC-DTF is typically treated with surgery, but clinicians may occasionally consider more holistic treatment plans that involve radioactive iodine and endocrine therapy.

Dietary supplementation with probiotics increases growth performance, improves the intestinal mucosal barrier and activates the Wnt/ß-catenin pathway activity in chicks.

BACKGROUND: Probiotics comprise effective feed additives that can replace antibiotics in animal livestock production. However, mono-strain probiotics appear less effective because of their instability. Therefore, the present study aimed to investigate dietary supplementation with compound probiotics (CPP) on growth performance, diarrhea rate and intestinal mucosal barrier, as well as the possible molecular mechanism, in chicks. In total, 360 1-day-old chicks of the Hy-Line Brown Chicks were randomly divided into the control group (CON, basal diet), chlortetracycline group (500 mg kg-1 CTC) and compound probiotics group (1000 mg kg-1 CPP, consisting of Bacillus subtilis, Bacillus licheniformis, Enterococcus faecium and yeast). The experiment period was 56 days. RESULTS: The results showed that, in comparison with the CON group, CPP significantly increased the average daily feed intake and average daily gain of chicks and reduced diarrhea (P < 0.05). The probiotic group exhibited increased immune organ (i.e. spleen and thymus) mass and increased levels of serum immunoglobulin (Ig)A, IgM and IgG (P < 0.05) compared to the CTC group. In addition, the jejunal mass and morphology were improved in the probiotic group (P < 0.05). Moreover, CPP reinforced jejunal barrier function, as indicated by increased transepithelial electrical resistance, protein expression of occludin and claudin-1, and diamine oxidase levels in the jejunum (P < 0.05). Likewise, enhanced fluorescence signals of proliferating cell nuclear antigen-labeled mitotic cells and villin-labeled absorptive cells in the jejunum (P < 0.05) suggested that CPP promoted intestinal stem cells activity. Mechanistically, the Wnt/ß-catenin signaling pathway, including ß-catenin, TCF4, c-Myc, cyclin D1 and Lgr5, was amplified in the jejunum by CPP addition (P < 0.05). CONCLUSION: The present study demonstrated that dietary supplementation with CPP reinforced the jejunal epithelial integrity by activating Wnt/ß-catenin signaling and enhanced immune function in chicks. © 2023 Society of Chemical Industry.

[Effect of total flavonoids of buckwheat flower and leaf on myocardial cell apoptosis and Wnt/ß-catenin/PPARγ pathway in arrhythmic rats].

This paper aimed to investigate the effect of total flavonoids of buckwheat flower and leaf on myocardial cell apoptosis and Wnt/ß-catenin/peroxisome proliferator-activated receptor γ(PPARγ) pathway in arrhythmic rats. SD rats were randomly divided into a control group, a model group, a low-dose(20 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a medium-dose(40 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a high-dose(80 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a propranolol hydrochloride(2 mg·kg~(-1)) group, with 12 rats in each group. Except the control group, rats in other groups were prepared as models of arrhythmia by sublingual injection of 1 mL·kg~(-1) of 0.002% aconitine. After grouping and intervention with drugs, the arrhythmia, myocardial cells apoptosis, myocardial tissue glutathione peroxidase(GSH-Px), catalase(CAT), malondialdehyde(MDA), serum interleukin-6(IL-6), prostaglandin E2(PGE2) levels, myocardial tissue apoptosis, and Wnt/ß-catenin/PPARγ pathway-related protein expression of rats in each group were measured. As compared with the control group, the arrhythmia score, the number of ventricular premature beats, ventricular fibrillation duration, myocardial cell apoptosis rate, MDA levels in myocardial tissues, serum IL-6 and PGE2 levels, Bax in myocardial tissues, and Wnt1 and ß-catenin protein expression levels increased significantly in the model group, whereas the GSH-Px and CAT levels, and Bcl-2 and PPARγ protein expression levels in myocardial tissues reduced significantly. As compared with the model group, the arrhythmia score, the number of ventricular premature beats, ventricular fibrillation duration, myocardial cell apoptosis rate, MDA leve in myocardial tissues, serum IL-6 and PGE2 levels, Bax in myocardial tissues, and Wnt1 and ß-catenin protein expression levels reduced in the drug intervention groups, whereas the GSH-Px and CAT levels and Bcl-2 and PPARγ protein expression levels in myocardial tissues increased. The groups of total flavonoids of buckwheat flower and leaf were in a dose-dependent manner. There was no significant difference in the levels of each index in rats between the propranolol hydrochloride group and the high-dose group of total flavonoids of buckwheat flower and leaf. The total flavonoids of buckwheat flower and leaf inhibit the activation of Wnt/ß-catenin pathway, up-regulate the expression of PPARγ, reduce oxidative stress and inflammatory damage in myocardial tissues of arrhythmic rats, reduce myocardial cell apoptosis, and improve the symptoms of arrhythmia in rats.

Ursolic Acid Suppresses Colorectal Cancer by Down-Regulation of Wnt/ß-Catenin Signaling Pathway Activity.

Overwhelming evidence points to an abnormally active Wnt/ß-catenin signaling as a key player in colorectal cancer (CRC) pathogenesis. Ursolic acid (UA) is a pentacyclic triterpenoid that has been found in a broad variety of fruits, spices, and medicinal plants. UA has been shown to have potent bioactivity against a variety of cancers, including CRC, with the action mechanism obscure. Our study tried to learn more about the efficacy of UA on CRC and its functional mechanism amid the Wnt/ß-catenin signaling cascade. We determined the efficacy of UA on CRC SW620 cells with respect to the proliferation, migration, clonality, apoptosis, cell cycle, and Wnt/ß-catenin signaling cascade, with assessment of the effect of UA on normal colonic NCM460 cells. Also, the effects of UA on the tumor development, apoptosis, cell cycle, and Wnt/ß-catenin signaling axis were evaluated after a subcutaneous SW620 xenograft tumor model was established in mice. In this work, we showed that UA drastically suppressed proliferation, migration, and clonality; induced apoptosis; and arrested the cell cycle at the G0/G1 phase of SW620 cells, without the influence on NCM460 cells, accompanied by weakened activity of the Wnt/ß-catenin signaling pathway. Besides, UA markedly deterred the growth of the xenograft tumor, ameliorated pathological features, triggered apoptosis, and arrested the cell cycle in xenograft CRC tissue, by lessening the Wnt/ß-catenin signaling cascade. Overall, UA may inhibit the malignant phenotype, induce apoptosis, and arrest the cell cycle of CRC, potentially by attenuating the Wnt/ß-catenin signaling axis, providing insights into the mechanism for the potency of UA on CRC.

Physical exercise suppresses hepatocellular carcinoma progression by alleviating hypoxia and attenuating cancer stemness through the Akt/GSK-3ß/ß-catenin pathway.

OBJECTIVE: Physical exercise, a common non-drug intervention, is an important strategy in cancer treatment, including hepatocellular carcinoma (HCC). However, the mechanism remains largely unknown. Due to the importance of hypoxia and cancer stemness in the development of HCC, the present study investigated whether the anti-HCC effect of physical exercise is related to its suppression on hypoxia and cancer stemness. METHODS: A physical exercise intervention of swimming (30 min/d, 5 d/week, for 4 weeks) was administered to BALB/c nude mice bearing subcutaneous human HCC tumor. The anti-HCC effect of swimming was assessed in vivo by tumor weight monitoring, hematoxylin and eosin (HE) staining, and immunohistochemistry (IHC) detection of proliferating cell nuclear antigen (PCNA) and Ki67. The expression of stemness transcription factors, including Nanog homeobox (NANOG), octamer-binding transcription factor 4 (OCT-4), v-Myc avian myelocytomatosis viral oncogene homolog (C-MYC) and hypoxia-inducible factor-1α (HIF-1α), was detected using real-time reverse transcription polymerase chain reaction. A hypoxia probe was used to explore the intratumoral hypoxia status. Western blot was used to detect the expression of HIF-1α and proteins related to protein kinase B (Akt)/glycogen synthase kinase-3ß (GSK-3ß)/ß-catenin signaling pathway. The IHC analysis of platelet endothelial cell adhesion molecule-1 (CD31), and the immunofluorescence co-location of CD31 and desmin were used to analyze tumor blood perfusion. SMMC-7721 cells were treated with nude mice serum. The inhibition effect on cancer stemness in vitro was detected using suspension sphere experiments and the expression of stemness transcription factors. The hypoxia status was inferred by measuring the protein and mRNA levels of HIF-1α. Further, the expression of proteins related to Akt/GSK-3ß/ß-catenin signaling pathway was detected. RESULTS: Swimming significantly reduced the body weight and tumor weight in nude mice bearing HCC tumor. HE staining and IHC results showed a lower necrotic area ratio as well as fewer PCNA or Ki67 positive cells in mice receiving the swimming intervention. Swimming potently alleviated the intratumoral hypoxia, attenuated the cancer stemness, and inhibited the Akt/GSK-3ß/ß-catenin signaling pathway. Additionally, the desmin+/CD31+ ratio, rather than the number of CD31+ vessels, was significantly increased in swimming-treated mice. In vitro experiments showed that treating cells with the serum from the swimming intervention mice significantly reduced the formation of SMMC-7721 cell suspension sphere, as well as the mRNA expression level of stemness transcription factors. Consistent with the in vivo results, HIF-1α and Akt/GSK-3ß/ß-catenin signaling pathway were also inhibited in cells treated with serum from swimming group. CONCLUSION: Swimming alleviated hypoxia and attenuated cancer stemness in HCC, through suppression of the Akt/GSK-3ß/ß-catenin signaling pathway. The alleviation of intratumoral hypoxia was related to the increase in blood perfusion in the tumor. Please cite this article as: Xiao CL, Zhong ZP, Lü C, Guo BJ, Chen JJ, Zhao T, Yin ZF, Li B. Physical exercise suppresses hepatocellular carcinoma progression by alleviating hypoxia and attenuating cancer stemness through the Akt/GSK-3ß/ß-catenin pathway. J Integr Med. 2023; 21(2): 184-193.

ß-Elemene regulates epithelial-mesenchymal transformation and inhibits invasion and metastasis of colorectal cancer cells.

OBJECTIVES: To study the inhibitory effect of ß-elemene on invasion and metastasis of colorectal cancer cells and its possible mechanism. METHODS: Human colon cancer HCT116 cells were treated with different concentrations of ß-elemene. The proliferation inhibition rate of the cells was detected by MTT assay, cell migration rate was detected by scratched assay, and cell invasion rate was evaluated by Transwell cell invasion assay. The expressions of Vimentin, E-cadherin, N-cadherin, and ß-catenin were detected by Western blotting. The mRNA expressions of Vimentin, E-cadherin, N-cadherin, and ß-catenin were detected by real-time PCR. RESULTS: Compared with the control group, the expressions of migration rate, invasion rate, scratch healing rate, N-cadherin, and Vimentin protein of HCT116 cells were decreased after ß-elemene treatment, while the expression of E-cadherin protein was increased, and the inhibition rate of cell proliferation was increased (p<0.05). CONCLUSIONS: ß-Elemene may inhibit cell proliferation and invasion and metastasis by inhibiting EMT signaling pathway in human colon cancer cell line HCT116.

Potential role of nutraceuticals via targeting a Wnt/ß-catenin and NF-κB pathway in treatment of osteoarthritis.

Osteoarthritis (OA) is a disease due to the aging of the articular cartilage, a post-mitotic tissue that stays functioning until primary homeostatic processes fail. Because of pain and disability, OA significantly influences national healthcare expenses and patient quality of life. It is a whole-joint illness characterized by inflammatory and oxidative signaling pathways and significant epigenetic alterations that cause cartilage extracellular matrix degradation. The canonical Wnt pathway (Wnt/ß-catenin pathway) and nuclear factor kappa B (NF-κB) signaling pathways may function in joint tissues by modulating the activity of synovial cells, osteoblasts, and chondrocytes. However, finding innovative ways to treat osteoarthritis and get the joint back to average balance is still a struggle. Nutraceuticals are dietary supplements that promote joint health by balancing anabolic and catabolic signals. New therapeutic methods for OA treatment have been developed based on many research findings that show nutraceuticals have strong anti-inflammation, antioxidant, anti-bone resorption, and anabolic properties. For the treatment of osteoarthritis, we explore the possible involvement of nutraceuticals that target the Wnt/ß-catenin and NF-κB pathways. PRACTICAL APPLICATIONS: In keeping with the aging population, osteoarthritis is becoming more widespread. In this extensive research, we studied the role of the Wnt/ß-catenin and NF-κB pathway in OA formation and progression. Nutraceuticals that target these OA-related signaling pathways are a viable therapy option. Wnt/ß-catenin and NF-κB signaling pathway are inhibited by polyphenols, flavonoids, alkaloids, and vitamins from the nutraceutical category, making them possible therapeutic drugs for OA therapy.

Identification of Hair Growth Promoting Components in the Kernels of Prunus mira Koehne and Their Mechanism of Action.

The application of the seed oil of Prunus mira Koehne (Tibetan name ཁམབུ།), a plant belonging to the Rosaceae family, for the treatment of alopecia has been recorded in Jingzhu Materia Medica (ཤེལ་གོང་ཤེལ་ཕྲེང་།) (the classic of Tibetan medicine) and Dictionary of Chinese Ethnic Medicine. This study aims to reveal the effective components and mechanism of hair growth promotion in the kernel of Prunus mira Koehne. Network pharmacology was used to predict the mechanism of action and effective components in the treatment of the kernel of Prunus mira Koehne. The contents of amygdalin in 12 batches of the kernel of Prunus mira Koehne were determined by HPLC. An animal model of the depilation of KM mice induced by sodium sulfide was created, and five effective components that promoted hair growth were initially screened. In the study of the effectiveness and mechanism of action, KM and C57BL/6 mice are selected as experimental objects, three screening tests for active components of the kernel of P. mira are performed, and three effective components are screened out from the eight components. HE staining was used to detect the number of hair follicles and the thickness of the dermis. RT-PCR and immunohistochemistry were used to evaluate the influence of the expression of indicators in the Wnt/ß-catenin signaling pathway in skin, including ß-catenin, GSK-3ß, and mRNA and protein expression levels of Cyclin D 1 and LEF 1. The network pharmacology study showed 12 signaling pathways involving 25 targets in the treatment of alopecia by the kernel of Prunus mira Koehne. vitamin E (3.125 mg/cm2/d), ß-sitosterol (0.061 mg/cm2/d), and linoleic acid (0.156 mg/cm2/d) in the kernel of Prunus mira Koehne can promote hair growth in mice, and the mechanism of action may be related to the Wnt/ß-catenin pathway.