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Metastasis is the major cause of death and failure of cancer chemotherapy in patients with breast cancer (BC). Activation of TGF-β/lncRNA-MALAT1/miR-200c has been reported to play an essential role during the metastasis of BC cells. The present study aimed to validate the suppression of BC-cell migration and invasion by baicalin and explore its regulatory effects on the TGF-β/lncRNA-MALAT1/miR-200c signaling pathway. We found that baicalin treatment inhibited cell viability and migration and invasion. Mechanistically, baicalin treatment significantly downregulated the expression of TGF-β, ZEB1, and N-cadherin and upregulated E-cadherin on both mRNA and protein levels. Additionally, baicalin treatment significantly downregulated the expression of lncRNA-MALAT1 and upregulated that of miR-200c. Collectively, baicalin significantly suppresses cell viability, migration, and invasion of BC cells possibly by regulating the TGF-β/lncRNA-MALAT1/miR-200c pathway.

Purpose The mechanism of lymph node metastasis (LNM) of cervical squamous cell carcinoma (CSCC) remains unconfirmed. Here we investigate the effects and mechanisms of midkine (MDK) on CSCC LNM. Methods The expression of MDK in CSCC and normal cervical tissue was analyzed. In vivo and in vitro experiments explored the effects of MDK on LNM. A retrospective study of 134 CSCC and 53 healthy controls on the relationship of serum-MDK(s-MDK) and clinicopathological characteristics was performed to evaluate the clinical value of s-MDK in CSCC LNM. Results MDK was highly expressed in CSCC and overexpression of MDK was associated with CSCC LNM.MDK promoted LNM by enhancing proliferation, migration and invasion capacity of cervical cancer cells, facilitating lymphangiogenesis and down-regulating the expression of tight junction proteins of human lymphatic endothelial cells. MDK exerted these biological effects by interacting with syndecan-1 and activating PI3K/AKT and p38 MAPK pathway. s-MDK was related to LNM, stage, tumor size, vascular invasion and muscle invasion. s-MDK combined with serum-squamous cell carcinoma antigen improved the diagnostic accuracy of CSCC LNM. Conclusions These findings established a new mechanism of LNM and highlighted MDK as a candidate tumor biomarker and therapeutic target in CSCC.

Abstract Peritoneal metastasis is a common issue in the progression of high-grade serous ovarian cancers (HGSOCs), yet the underlying mechanism remains unconfirmed. We demonstrated that ZEB2, the transcription factor of epithelial–mesenchymal transition (EMT), was upregulated in ascites cells from HGSOC patients and in CD133+ cancer stem-like cells (CSLCs) from epithelial ovarian cancer (EOC) cell lines. SiRNA-mediated knockdown of ZEB2 in EOC cells decreased the percentage of CSLCs and reduced the colony forming potential, cell invasion capacity and expression of pluripotent genes Oct4 and Nanog. Inhibition of ZEB2 also induced cellular apoptosis and impacted the tumorigenicity of ovarian CSLCs. The mesenchymal markers N-cadherin and vimentin were downregulated, while the epithelial marker E-cadherin was upregulated after ZEB2 knockdown. MiR-200a, a molecule that downregulates ZEB2, had the opposite effect of ZEB2 expression in EOC-CSLCs. A retrospective study of 98 HGSOC patients on the relationship of ascites volume, pelvic and abdominal metastasis, International Federation of Gynecology and Obstetrics (FIGO) stage and the malignant involvement of abdominal organs and lymph nodes was performed. Patients with high expression of ZEB2 in tumour tissues had a higher metastasis rate and a poorer prognosis than those with low expression. The parameters of ZEB2 expression and ascites volume were strongly linked with the prognostic outcome of HGSOC patients and had higher hazard ratios. These findings illustrated that ZEB2 facilitates the invasive metastasis of EOC-CSLCs and can predict peritoneal metastasis and a poor prognosis in HGSOC patients.

Objective: Cyclophosphamide (CTX), which is commonly used in clinical chemotherapy, has a damaging effect on ovarian follicles. This study aimed to establish a new method to count the number of follicles in mouse ovaries using benzyl alcohol–benzyl benzoate (BABB)-based tissue-clearing technology and evaluate the follicle-damaging effects of different doses of CTX. Methods: C57BL/6 mice were divided into four groups and administered intraperitoneal injections of 0, 40, 80, or 120 mg/kg CTX. The serum levels of estradiol (E2) and follicle-stimulating hormone (FSH) were detected using an ELISA kit. Mouse ovaries were subjected to BABB clearing and labeled with DAPI, β-actin, and DDX4 to observe the ovarian structure and follicles. A three-dimensional software, Imaris, was used to reconstruct the ovarian structure and automatically count the number of follicles. The effects of different CTX doses on the total follicle number and estrous cycle were determined. Results: As the CTX dose increased, E2 levels in CTX mice declined from 212.3 to 57.7 pg/mL; the FSH levels increased from 3.2 to 29 ng/mL. Mouse ovaries became transparent after BABB treatment. After fixation, microscopy, and Imaris processing, immunofluorescence signals of β-actin and DAPI from all levels in intact ovaries could be obtained and follicle number in half ovaries could be automatically counted using anti-DDX4 antibody labeling. In the NC, CTX40, CTX80, and CTX120 groups, the proportion of mice in the diestrus phase was 26.67%, 51.67%, 73.33%, and 95.00%, respectively, and the total follicle number was 2,603, 1,761, 1,043, and 262, respectively. E2 levels were positively correlated with follicle number and FSH levels were negatively correlated with follicle number, indicating that the damaging effect of CTX on follicle number may be dose dependent. Conclusion: BABB can be used to clear intact ovaries from adult mice, and follicle number in half ovaries can be automatically counted. The damaging effect of CTX on follicles and the endocrine system is dose dependent.

Background. Ovarian cancer (OC) is one of the leading lethal gynecologic cancers of women around the world. More than 70% of patients are diagnosed with stage III or IV with poor outcome. This is partly because of lacking early effective screening techniques and potential biomarkers of OC. CXC chemokines in tumor microenvironment (TME) and their interaction with relative receptors can excite the downstream signaling pathways to influence tumor progression. However, the role of CXC chemokines in OC has not been identified. Methods. ONCOMINE, GEPIA, Kaplan–Meier plotter, cBioPortal, TIMER, Metascape, and LinkedOmics were applied in our study. Results. The transcriptional levels of CXCL1/8/9/10/11/12/13/14/16/17 were significantly elevated while CXCL3 was obviously reduced in OC vs normal ovarian tissue. CXCL8/9/11/13 were correlated with clinic pathological stage. Patients with low expression of CXCL8/9/11/13 were associated with better prognosis. We also found that CXCL3 and CXC12 could be used as potential prognostic markers of OC through Kaplan–Meier plotter. Patients with high expression of CXCL3/12 had a significantly better prognosis. Their functions focus on locomotion, signaling, response to stimulus, undergoing the process of multiorganism, immune system, biological regulation, etc. The differentiated CXC chemokines mainly participate in cytokine-cytokine receptor interaction, chemokine signaling pathway, IL-17 signaling pathway, and toll-like receptor signaling pathway. Our results showed that CXC chemokines were highly correlated with infiltration of immune cells. The kinase targets of differentially expressed CXC chemokines are mainly in ATM, LYN, LCK, PLK1, FYN, CDK2, and ATR. Conclusions. Our results may provide a new insight for selecting precision biomarkers of targeted therapy of OC.

The diversity of the human microbiome heralds the difference of the impact that gut microbial metabolites exert on allogenic graft-versus-host (GVH) disease (GVHD), even though short-chain fatty acids and indole were demonstrated to reduce its severity. In this study, we dissected the role of choline-metabolized trimethylamine N-oxide (TMAO) in the GVHD process. Either TMAO or a high-choline diet enhanced the allogenic GVH reaction, whereas the analog of choline, 3,3-dimethyl-1-butanol reversed TMAO-induced GVHD severity. Interestingly, TMAO-induced alloreactive T-cell proliferation and differentiation into T-helper (Th) subtypes was seen in GVHD mice but not in in vitro cultures. We thus investigated the role of macrophage polarization, which was absent from the in vitro culture system. F4/80+CD11b+CD16/32+ M1 macrophage and signature genes, IL-1β, IL-6, TNF-α, CXCL9, and CXCL10, were increased in TMAO-induced GVHD tissues and in TMAO-cultured bone marrow–derived macrophages (BMDMs). Inhibition of the NLRP3 inflammasome reversed TMAO-stimulated M1 features, indicating that NLRP3 is the key proteolytic activator involved in the macrophage’s response to TMAO stimulation. Consistently, mitochondrial reactive oxygen species and enhanced NF-κB nuclear relocalization were investigated in TMAO-stimulated BMDMs. In vivo depletion of NLRP3 in GVHD recipients not only blocked M1 polarization but also reversed GVHD severity in the presence of TMAO treatment. In conclusion, our data revealed that TMAO-induced GVHD progression resulted from Th1 and Th17 differentiation, which is mediated by the polarized M1 macrophage requiring NLRP3 inflammasome activation. It provides the link among the host choline diet, microbial metabolites, and GVH reaction, shedding light on alleviating GVHD by controlling choline intake.

Irradiation induces severe damage in the hematopoietic system, which leads to bone marrow hyperplasia, pancytopenia, and aggravated tissue formation in bone marrow. Studies have shown that Toll-like receptor 4 (TLR4) has a protective effect against irradiation, but the underlying mechanism remains unclear. In this study, we used a TLR4 knockout (TLR4-/-) mouse irradiation model and found that the white blood cell and platelet counts in the peripheral blood of TLR4-/- mice recovered slowly after irradiation, with bone marrow hyperplasia and increased mortality. Additionally, we found that the proportion of CD11b+Gr1+ granulocytes in the peripheral blood and bone marrow of TLR4-/- mice was lower than that of wild-type mice after irradiation. Further, we found that the expression of NADPH Oxidases (NOXs) in the bone marrow was down-regulated after irradiation of TLR4-/- mice, and administration of the NOXs inhibitor VAS2870 reduced the proportion of CD11b+Gr1+ cells in the bone marrow and peripheral blood of wild-type mice after irradiation. Irradiation induced severe marrow adipocytes accumulation in TLR4-/- mice, TLR4 ligand lipopolysaccharide promoted proliferation and inhibited adipogenic differentiation of mesenchymal stromal cells. In summary, our data suggest that TLR4 promotes myeloid hyperplasia by up-regulating the expression of NOXs after irradiation, prohibits marrow adipogensis and increases the tolerance of mice to irradiation.

Aplastic anemia (AA) is generally considered as an immune-mediated bone marrow failure syndrome. Several studies show that bone marrow mesenchymal stem cells (BM-MSCs), as key cellular components of the bone marrow microenvironment, are also involved in the pathogenic mechanism of AA. Cyclosporin A (CsA) is a classic immunosuppressive drug for AA, and it specifically inhibits mammalian T cells by preventing activation of transcription factors involved in cytokine gene expression. However, little is known about the effect of CsA on the BM-MSCs. In this study, murine BM-MSCs were stimulated in the presence of CsA. Further, we found that CsA could inhibit murine BM-MSC proliferation and promote BM-MSC apoptosis, what's more CsA could inhibit adipogenic differentiation. Our study also showed that CsA could inhibit interleukin-6 expression in BM-MSCs, while promoting programmed death-ligand 2 expression. In conclusion, our results proposed that CsA may exert an effect on regulating the bone marrow environment by influencing BM-MSCs, which have a beneficial effect on treating AA.

Mesenchymal stem cells (MSCs) give origin to the marrow tromal environment that supports hematopoiesis. These cells present a wide range of differentiation potentials and a complex relationship with hematopoietic stem cells (HSCs) and endothelial cells. In addition to bone marrow (BM), MSCs can be obtained from other sites in the adult or the fetus. Recent studies have shown that cocultured endothelial cells and osteoblasts are mutually promotive in bone tissues repair. In this study, we observed the effects of coculture of endothelial cells and osteoblasts at different ratios on vasculogenesis and bone formation, and we found that angiogenic effect is more effective when endothelial cells are cocultured with osteoblasts at the ratio of 4:1, and osteogenic effect is more effective at the ratio of 1:4. It is concluded that the co-culture of human bone marrow mesenchymal stromal cells with human umbilical vein endothelial cells could be a promising culture system for bone tissue engineering applications.

Mesenchymal stem cells (MSCs) give origin to the marrow tromal environment that supports hematopoiesis. These cells present a wide range of differentiation potentials and a complex relationship with hematopoietic stem cells (HSCs) and endothelial cells. In addition to bone marrow (BM), MSCs can be obtained from other sites in the adult or the fetus. Recent studies have shown that cocultured endothelial cells and osteoblasts are mutually promotive in bone tissues repair. In this study, we observed the effects of coculture of endothelial cells and osteoblasts at different ratios on vasculogenesis and bone formation, and we found that angiogenic effect is more effective when endothelial cells are cocultured with osteoblasts at the ratio of 4:1, and osteogenic effect is more effective at the ratio of 1:4. It is concluded that the co-culture of human bone marrow mesenchymal stromal cells with human umbilical vein endothelial cells could be a promising culture system for bone tissue engineering applications.