Colorimetric detection of circulating tumor cells in breast cancer based on ladder-branch hybridization chain reaction and DFs/AuNCs nanozyme.

Breast cancer is the most common malignant tumor in women, which accounts for 6.9% of all cancer-related deaths. Early diagnosis is crucial for making the best clinical decision and improving the prognosis of patients. Circulating tumor cells (CTCs) have been regarded as significant tumor biomarkers. Herein, we designed a colorimetric biosensor for breast cancer CTCs quantification based on ladder-branch hybridization chain reaction (HCR) and DNA flowers/gold nanoclusters (DFs/AuNCs) nanozyme. With the assistance of complementary DNA labeled on magnetic beads (MBs), the cleavage products of RNA-cleaving DNAzymes (RCDs) could be rapidly captured, subsequently triggering ladder-branch HCR. In addition, the DFs/AuNCs nanozyme was applied for colorimetric analysis, which further improved the sensitivity for the detection of target CTCs. Benefiting from specific RCDs, ladder-branch HCR and DFs/AuNCs, we achieved a superior detection limit of 3 cells/mL as well as a broad linear range of 10 cells/mL to 104 cells/mL. Conclusively, this colorimetric biosensor achieved sensitively and selectively detection of breast cancer CTCs without the participation of enzymes at room temperature, which might provide new insight into the early detection of breast cancer.

Pretreatment with Daudi cells eliminates anti-CD47 monoclonal antibody interference in immunohematology testing.

BACKGROUND: Anti-CD47 monoclonal antibodies have increasing clinical applications in the treatment of cancer. However, anti-CD47 monoclonal antibodies interfere with immunohematology testing in patients who require blood transfusion. As the current approaches to removing any interferences have technical problems, new methods need to be developed to resolve anti-CD47 interference in immunohematology testing. MATERIALS AND METHODS: We evaluated the Daudi cell line for the adsorption of free anti-CD47 monoclonal antibodies from patients' plasma to facilitate immunohematology testing in patients treated with anti-CD47 monoclonal antibody. CD47 expression was identified on the Daudi cells using flow cytometry and confocal microscopy. Next, we tested the ability of intact Daudi cells mixed with simulating plasma and clinical samples to achieve efficient removal of interfering anti-CD47 monoclonal antibodies. The indirect antiglobulin test was used to verify whether interference from anti-CD47 monoclonal antibodies in plasma was eliminated and whether the detection of other irregular antibodies was affected. The effect of eliminating interference was also investigated in relation to the time that the Daudi cells were stored after having been fixed with paraformaldehyde. RESULTS: CD47 expression was higher on Daudi cells than on red blood cells. Analysis of the indirect antiglobulin test results revealed that anti-CD47 monoclonal antibody-treated patients' plasma absorbed by Daudi cells for 15 min at 37°C could completely prevent the interference of anti-CD47 monoclonal antibodies in immunohematology testing while the detection of the tested antibodies, including anti-D and anti-K, was unaffected. DISCUSSION: By decreasing the incubation time, we discovered that interferences in samples with agglutination strengths below 2+ could be eliminated after incubation for 5 min. Of importance, Daudi cells can be preserved with 4% paraformaldehyde for 14 days as short-term storage reagents. This is the first study in which Daudi cells were used to effectively resolve the interference of anti-CD47 monoclonal antibodies in pretransfusion tests.

Tumor necrosis factor-like cytokine 1A plays a role in inflammatory bowel disease pathogenesis.

The binding of tumor necrosis factor-like cytokine 1A (TL1A) to death receptor 3 (DR3) plays an important role in the interaction between dendritic cells (DCs) and T cells and contributes to intestinal inflammation development. However, the mechanism by which DCs expressing TL1A mediate helper T (Th) cell differentiation in the intestinal lamina propria (LP) during the pathogenesis of inflammatory bowel disease remains unclear. In this study, we found that TL1A/DR3 promoted Th1 and Th17 cell differentiation in T-T and DC-T cell interaction-dependent manners. TL1A-deficient CD4+ T cells failed to polarize into Th1/Th17 cells and did not cause colonic inflammation in a T cell transfer colitis model. Notably, TL1A was located in the cytoplasm and nuclei of DCs, positively regulated the DC-specific ICAM-grabbing nonintegrin/RAF1/nuclear factor κB signaling pathway, enhanced the antigen uptake ability of DCs, and promoted TLR4-mediated DC activation, inducing naive CD4+ T cell differentiation into Th1 and Th17 cells. Our work reveals that TL1A plays a regulatory role in inflammatory bowel disease pathogenesis.

CD30L is involved in the regulation of the inflammatory response through inducing homing and differentiation of monocytes via CCL2/CCR2 axis and NF-κB pathway in mice with colitis.

The pathogenesis of inflammatory bowel diseases (IBD) is complex, and dysregulated immune responses play a pivotal role in its occurrence and development. Our previous studies indicated that CD30L may participate in monocyte-mediated inflammation in patients with UC through the activation of circulating monocytes. However, it remains unclear how CD30L participates in monocyte-mediated inflammation in IBD by activation of circulating monocytes. In this study, we observed an increase in the expression of CD30L and chemokine receptor type 2 (CCR2) on circulating monocytes and pro-inflammatory monocytes in the colon lamina propria in mice with dextran sulfate sodium salt (DSS)-induced colitis. Moreover, there was a positive correlation between the expression levels of CCR2 and CD30L (r = 0.8817, p = 0.0480) in monocytes. In Cd30l-/- mice with DSS-induced colitis, the percentage and absolute number of circulating monocytes and pro-inflammatory monocytes decreased with the downregulation of CCR2. Stimulation via CD30L by immobilized anti-CD30L mAb suppressed the expression of pNF-κB p65, pIκBα, p65 and CCR2 and up-regulated the expression of IκBα in the sorted pro-inflammatory monocytes in Cd30l-/- mice with DSS-induced colitis. The mRNA levels of Ccr2 in the sorted pro-inflammatory monocytes were significantly down-regulated with the presence of immobilized RM153 and inhibitors of NF-κB (BAY 11-7082) in WT mice with DSS-induced colitis. Our results suggested that CD30L could promote the inflammatory response by inducing the homing and differentiation of monocytes via the chemokine ligand 2 (CCL2)/CCR2 axis and NF-κB signaling pathway in mice with colitis. These findings provide a novel target for monocyte-based immunotherapy against IBD.

Interleukin-36ß exacerbates DSS-induce acute colitis via inhibiting Foxp3+ regulatory T cell response and increasing Th2 cell response.

BACKGROUND: interleukin (IL)-36ß is a member of the IL-36 subfamily of the IL-1 family. Usually, serum levels of IL-36ß are higher in patients with inflammatory bowel disease (IBD), indicating that IL-36ß has a pathophysiological role in IBD. At the time of writing, no studies were published reporting the role of IL-36ß in modulating T cell-mediated immune responses in gastrointestinal inflammation. This research aimed to determine the function of IL-36ß in regulating T cells in mice with colitis caused by dextran sulfate sodium (DSS). METHODS: recombinant IL-36ß (rIL-36ß) was administered by intraperitoneal injection to mice with DSS-induced colitis. Clinical symptoms, colon length, and histological changes were determined. The production of cytokines was measured by ELISA. The help T (Th) cell subsets were measured by flow cytometry. The expression of mRNA of IL-36ß was measured by quantitative real-time PCR. RESULTS: there was an increased expression of IL-36 in the inflamed colonic mucosa of mice with DSS-induced acute colitis. Mice treated with recombinant IL-36ß (rIL-36ß) were more susceptible to DSS-induced colitis than PBS-treated mice. Moreover, spontaneous inflammatory cytokines produced by neutrophils greatly increased in the lamina propria lymphocytes (LPL) of rIL-36ß-treated animals with DSS-induced colitis. Besides, rIL-36ß-treatment dramatically elevated Th2 cell responses but significantly downregulated Foxp3+ regulatory T cell (Treg) responses. CONCLUSION: these findings indicate that IL-36ß enhances the pathology of DSS-induced colitis in mice by promoting Th2 responses in LPL while decreasing Foxp3+ Treg responses. Thus, we propose the regulation of the IL-36ß/IL-36R signaling pathway as a potential biological treatment for IBD.

Expression of Tspan-1 gene in patients with advanced gastric cancer.

The present study investigated the correlations of the Tspan-1 gene expression with the clinical characteristics and survival prognoses of patients with advanced gastric cancer. A total of 150 patients with advanced gastric cancer were enrolled in the present study, of whom 84 were at stage II and 66 were at stage III according to the tumor node metastasis (TNM) staging; the immunohistochemical staining method and the semi-quantitative PCR method were used to detect the positive expression rates and mRNA relative expression levels of Tspan-1, vascular endothelial growth factor (VEGF), E-cadherin and N-cadherin. The positive expression rates of Tspan-1, VEGF, E-cadherin and N-cadherin were 58.0% (87 patients), 50.0% (75 patients), 28.0% (42 patients) and 53.3% (80 patients), respectively. The positive expressions and mRNA levels of Tspan-1, VEGF, E-cadherin and N-cadherin were not correlated with sex or age (P>0.05), but associated with the cancer state (stage II or stage III) and maximum tumor diameter (P<0.05). With the increase of stage and tumor diameter, the positive rates and mRNA levels of Tspan-1, VEGF and N-cadherin were increased, while those of E-cadherin were decreased. Among patients with stage II/III advanced gastric cancer, those with positive expression of Tspan-1, VEGF and N-cadherin had lower median survival time and survival rates than patients with negative expressions, while patients with positive expression of E-cadherin had higher median survival time and survival rate than those with negative expression (P<0.05). The high expression of Tspan-1 gene is associated with the TNM staging of advanced gastric cancer and the tumor diameter, influences the survival prognosis, and may involve the processes of angiogenesis and epithelial-mesenchymal transition.