[Effect of Baicalin on Pyroptosis of Diffuse Large B-Cell Lymphoma Cell Lines DB and Its Mechanism].

OBJECTIVE: To investigate the effect of Baicalin on the proliferation and pyroptosis of diffuse large B-cell lymphoma cell line DB and its mechanism. METHODS: DB cells were treated with baicalin at different concentrations (0, 5, 10, 20, 40 µmol/L). Cell proliferation was detected by CCK-8 assay and half maximal inhibitory concentration (IC50) was calculated. The morphology of pyroptosis was observed under an inverted microscope, the integrity of the cell membrane was verified by LDH content release assay, and the expressions of pyroptosis-related mRNA and protein (NLRP3, GSDMD, GSDME, N-GSDMD, N-GSDME) were detected by real-time fluorescence quantitative PCR and Western blot. In order to further clarify the relationship between baicalin-induced pyroptosis and ROS production in DB cells, DB cells were divided into control group, baicalin group, NAC group and NAC combined with baicalin group. DB cells in the NAC group were pretreated with ROS inhibitor N-acetylcysteine (NAC) 2 mmol/L for 2 h. Baicalin was added to the combined treatment group after pretreatment, and the content of reactive oxygen species (ROS) in the cells was detected by DCFH-DA method after 48 hours of culture. RESULTS: Baicalin inhibited the proliferation of DB cells in a dose-dependent manner (r=-0.99), and the IC50 was 20.56 µmol/L at 48 h. The morphological changes of pyroptosis in DB cells were observed under inverted microscope. Compared with the control group, the release of LDH in the baicalin group was significantly increased (P<0.01), indicating the loss of cell membrane integrity. Baicalin dose-dependently increased the expression levels of NLRP3, N-GSDMD, and N-GSDME mRNA and protein in the pyroptosis pathway (P<0.05). Compared with the control group, the level of ROS in the baicalin group was significantly increased (P<0.05), and the content of ROS in the NAC group was significantly decreased (P<0.05). Compared with the NAC group, the content of ROS in the NAC + baicalin group was increased. Baicalin significantly attenuated the inhibitory effect of NAC on ROS production (P<0.05). Similarly, Western blot results showed that compared with the control group, the expression levels of pyroptosis-related proteins was increased in the baicalin group (P<0.05). NAC inhibited the expression of NLRP3 and reduced the cleavage of N-GSDMD and N-GSDME (P<0.05). Compared with the NAC group, the NAC + baicalin group had significantly increased expression of pyroptosis-related proteins. These results indicate that baicalin can effectively induce pyroptosis in DB cells and reverse the inhibitory effect of NAC on ROS production. CONCLUSION: Baicalin can inhibit the proliferation of DLBCL cell line DB, and its mechanism may be through regulating ROS production to affect the pyroptosis pathway.

A pairwise radiomics algorithm-lesion pair relation estimation model for distinguishing multiple primary lung cancer from intrapulmonary metastasis.

Background: Distinguishing multiple primary lung cancer (MPLC) from intrapulmonary metastasis (IPM) is critical for their disparate treatment strategy and prognosis. This study aimed to establish a non-invasive model to make the differentiation pre-operatively. Methods: We retrospectively studied 168 patients with multiple lung cancers (307 pairs of lesions) including 118 cases for modeling and internal validation, and 50 cases for independent external validation. Radiomic features on computed tomography (CT) were extracted to calculate the absolute deviation of paired lesions. Features were then selected by correlation coefficients and random forest classifier 5-fold cross-validation, based on which the lesion pair relation estimation (PRE) model was developed. A major voting strategy was used to decide diagnosis for cases with multiple pairs of lesions. Cases from another institute were included as the external validation set for the PRE model to compete with two experienced clinicians. Results: Seven radiomic features were selected for the PRE model construction. With major voting strategy, the mean area under receiver operating characteristic curve (AUC), accuracy, sensitivity, and specificity of the training versus internal validation versus external validation cohort to distinguish MPLC were 0.983 versus 0.844 versus 0.793, 0.942 versus 0.846 versus 0.760, 0.905 versus 0.728 versus 0.727, and 0.962 versus 0.910 versus 0.769, respectively. AUCs of the two clinicians were 0.619 and 0.580. Conclusions: The CT radiomic feature-based lesion PRE model is potentially an accurate diagnostic tool for the differentiation of MPLC and IPM, which could help with clinical decision making.

[Regulation of Baicalin on Growth of Extranodal NK/T Cell Lymphoma Cells through FOXO3/CCL22 Signaling Pathway].

OBJECTIVE: To investigate the effect of baicalin on the growth of extranodal NK/T cell lymphoma (ENKTCL) cells and its related mechanism. METHODS: Normal NK cells and human ENKTCL cells lines SNK-6 and YTS were cultured, then SNK-6 and YTS cells were treated with 5, 10, 20 µmol/L baicalin and set control. Cell proliferation and apoptosis was detected by Edu method and FCM method, respectively, and expressions of BCL-2, Bax, FOXO3 and CCL22 proteins were detected by Western blot. Interference plasmids were designed and synthesized. FOXO3 siRNA interference plasmids and CCL22 pcDNA overexpression plasmids were transfected with PEI transfection reagent. Furthermore, animal models were established for validation. RESULTS: In control group and 5, 10, 20 µmol/L baicalin group, the proliferation rate of SNK-6 cells was (56.17±2.96)%, (51.92±4.63)%, (36.42±1.58)%, and (14.60±2.81)%, respectively, while that of YTS cells was (58.85±2.98)%, (51.38±1.32)%, (34.75±1.09)%, and (15.45±1.10)%, respectively. In control group and 5, 10, 20 µmol/L baicalin group, the apoptosis rate of SNK-6 cells was (5.93±0.74)%, (11.78±0.34)%, (28.46±0.44)%, and (32.40±0.37)%, respectively, while that of YTS cells was (7.93±0.69)%, (16.29±1.35)%, (33.91±1.56)%, and (36.27±1.06)%, respectively. Compared with control group, the expression of BCL-2 protein both in SNK-6 and YTS cells decreased significantly (P<0.001), and the expression of Bax protein increased in SNK-6 cells only when the concentration of baicalin was 20 µmol/L (P<0.001), while that in YTS cells increased in all three concentrations(5, 10, 20 µmol/L) of baicalin (P<0.001). The expression of FOXO3 protein decreased while CCL22 protein increased in ENKTCL cell lines compared with human NK cells (P<0.001), but the expression of FOXO3 protein increased (P<0.01) and CCL22 protein decreased after baicalin treatment (P<0.001). Animal experiments showed that baicalin treatment could inhibit tumor growth. The expression of CCL22 protein in ENKTCL tissue of nude mice treated with baicalin decreased compared with control group (P<0.01), while the FOXO3 protein increased (P<0.05). In addition, FOXO3 silencing resulted in the decrease of FOXO3 protein expression and increase of CCL22 protein expression (P<0.01, P<0.001). CONCLUSION: Baicalin can inhibit proliferation and promote apoptosis of ENKTCL cell lines SNK-6 and YTS, up-regulate the expression of Bax protein, down-regulate the expression of BCL-2 protein, and down-regulate the expression of CCL22 protein mediated by FOXO3. Animal experiment shown that the baicalin can inhibit tumor growth. Baicalin can inhibit the growth and induce apoptosis of ENKTCL cells through FOXO3/CCL22 signaling pathway.

A novel inflammation-related prognostic model for predicting the overall survival of primary central nervous system lymphoma: A real-world data analysis.

Background: Primary central nervous system lymphoma (PCNSL) is a type of extranodal non-Hodgkin lymphoma. Although there are widely used prognostic scores, their accuracy and practicality are insufficient. Thus, a novel prognostic prediction model was developed for risk stratification of PCNSL patients in our research. Methods: We retrospectively collected 122 patients with PCNSL from two medical centers in China from January 2010 to June 2022. Among them, 72 patients were used as the development cohort to construct a new model, and 50 patients were used for the validation. Then, by using univariate and multivariate Cox regression analsis and Lasso analysis, the Xijing model was developed and composed of four variables, including lesion number, ß2-microglobulin (ß2-MG), systemic inflammation response index (SIRI) and Karnofsky performance status (KPS). Finally, we evaluated the Xijing model through internal and external validation. Results: Compared with the original prognostic scores, the Xijing model has an overall improvement in predicting the prognosis of PCNSL according to the time-dependent area under the curve (AUC), Harrell's concordance index (C-index), decision curve analysis (DCA), integrated discrimination improvement (IDI) and continuous net reclassification index (NRI). For overall survival (OS) and progression-free survival (PFS), the Xijing model can divide PCNSL patients into three groups, and shows more accurate stratification ability. In addition, the Xijing model can still stratify and predict prognosis similarly better in the elderly with PCNSL and subgroups received high-dose methotrexate (HD-MTX) or Bruton's tyrosine kinase inhibitors (BTKi). Finally, external validation confirmed the above results. Conclusions: Integrating four prognostic factors, including imaging findings, tumor burden, systemic inflammation response index, and comprehensive physical condition, we provided a novel prognostic model for PCNSL based on real-world data and evaluated its predictive capacity.

Metformin induces myeloma cells necrosis and apoptosis and it is considered for therapeutic use.

Accumulating evidence, especially in solid tumor, indicated that metformin possessed the potential ability in the proliferation of cancer cells. However, its effects on myeloma cells were relatively rarely clarified. To evaluate the anti-cancer effects of metformin against dexamethasone-resistant and -sensitive myeloma cells. The effects of metformin on myeloma cell lines, including dexamethasone-resistant U266, H929, RPMI 8226 and dexamethasone-sensitive MM.1s, were investigated using the cell counting kit-8 assay for cell proliferation. Apoptosis, necrosis, cell cycle arrest, and cell death mechanisms were explored via flow cytometry (FCM) and Western blot. In addition, the anti-myeloma activity was evaluated in vivo via non-obese diabetic/severe combined immunodeficiency xenograft mouse models. Metformin inhibited proliferation in a dose and time-dependent manner in all the cell lines, while dexamethasone only affected the viability of MM1.s cells. The FCM detection displayed that metformin induced apoptosis in H929, RPMI8226 and MM.1s cells, while for U266 cells, it induced necrosis with Annexin V-/Propidium iodide+. The cell cycle assays showed that metformin arrested G0/G1 phase of H929 and MM.1s cells, or G2/M phase of RPMI8226 cells, but showed no effect on U226 cells. Western blotting analyses demonstrated that the apoptosis-related protein of cleaved caspase 3 was activated; the expressions of Mcl-1, IGF-1R, PI3K, pAKT, and pmTOR proteins were inhibited by metformin in H929, RPMI8226, and MM.1s cells. The necrosis-related protein of iNOS increased in U266 cells while metformin treated. In vivo assay indicated metformin decreased U266 and H929 growth in bone marrow, and thus prolonged mice survival. These data suggested that metformin inhibited the proliferation of myeloma cells via inducing necrosis and apoptosis. This finding indicated that metformin may be served as a potent adjuvant in treating multiple myeloma.

Approach to multiplexing fiber communication with cylindrical vector beams.

Cylindrical vector beams (CVBs), including radial, azimuthal, and hybrid polarization vortex states, are based on the polarization singularity and regarded as the eigenmodes of fiber. In this Letter, we propose and demonstrate CVB (de)multiplexing communication in a few-mode fiber (FMF). We simulate the eight CVB modes, including the ±1 orders and the ±2 orders supported by the FMF. We measure the mode purities of the ±1-order and the ±2-order CVBs, which can reach 67.87%, 69.26%, 73.84%, and 71.95% after transmission in the 5 km FMF. The mode cross talk between the CVBs and their adjacent orders is less than -7.54 dB. In the experiment, we demonstrate four modes of coaxial CVB multiplexing communication with the orders of -2 and spatially orthogonal polarization states. The mode cross talk is less than -23.35 dB for all channels. CVB modes carrying 10 Gbit/s on-off keying signals are transmitted in the 5 km FMF and achieve bit-error rates below the forward error correction threshold of 3.8×10-3.

[Correlation between ALIP-like cluster and fibrous proliferation in bone marrow of patients with acute myeloid leukemia in CR phase].

This study was purposed to elucidate the prognostic values of ALIP (abnormal location of immature precursors)-like clusters and fibrous proliferation in bone marrow of AML patients in CR phase and the correlation between them. The bone marrow biopsy sections from 47 AML patients during admitting to relapse or till lost follow-up were examined retrospectively. The 47 patients were divided into pre-relapsed group and non-relapsed group according to relapse or not at end of follow-up. The concentration of ALIP-like cluster and reticulin fiber density (RFD) in sections were compared between the two groups respectively, the prognostic value of these two factors and the underlying relationship between them were estimated by statistical analysis. The results showed that ALIP-like cluster was (3.46 ± 2.71)/mm(2) during CR and RFD was (2.76% ± 1.50%) in pre-relapsed cases, which both were higher than those in non-relapsed cases (P < 0.05). Cases with ALIP-like cluster over 4/mm(2) or with RFD>1.68% showed high relapse rates of 89.5% or 95.2% respectively. RFD were (2.47% ± 2.48%) and (2.44% ± 2.23%) in cases with >4/mm(2) or ≤ 4/mm(2) respectively, there was no statistical significance (P > 0.05) . Meanwhile, the amount of ALIP-like cluster in CR not related with the paired RFD (r = 0.057, P > 0.05). It is concluded that both ALIP-like cluster in CR and RFD are poor prognostic factors for heralding early relapse. However, ALIP-like cluster and RFD show no correlation, and suggest that forming of ALIP not depends on fibrogenesis.

Clustered precursors in bone marrow sections predict early relapse in patients with acute myeloid leukemia within hematologic remission.

BACKGROUND: Bone marrow (BM) aspiration is largely used for relapse assessment in acute myeloid leukemia (AML). It remains unclear what roles that BM trephine biopsy plays on relapse assessment. METHODS: Bone marrow (BM) sections during complete remission (CR) from 60 acute myeloid leukemia (AML) patients were retrospectively analyzed. Computer image processing technology was performed for detection of the distance between precursors and endosteum, and density of precursors was also calculated under light microscopic image. Immunohistochemistry was used to identify the immunophenotype of clustered precursors. RESULTS: Except for single and double precursors, there existed clustered precursors of 3-5 cells during CR. Here, we demonstrated that clustered precursors, but not single and double precursors, were useful in risk factor of relapse. Area under the receiving operator curve (ROC) was of 0.007 (CI 95%, from 0.572 to 0.851). Using a standard cut-off value of >4.0/mm² for cluster density, early relapse was detected with a sensitivity of 51.5% and a specificity of 85.7%.Multivariate Cox regression analysis revealed that clustered precursor is an independent risk factor for early relapse (Adjusted HR: 0.325, 95% CI: 0.156-0.679, p = 0.003). CONCLUSIONS: Cumulatively, clustered precursors in BM sections during CR may serve as an independent risk factor of early relapse and poor outcome for AML patients in cluster density > 4.0/mm² in sections. Early aggressive interventions are needed to prevent hematologic relapse.

Clustered immature myeloid precursors in intertrabecular region during remission evolve from leukemia stem cell near endosteum and contribute to disease relapse in acute myeloid leukemia.

Most acute myeloid leukemia (AML) cannot be cured because leukemia stem cells (LSC) will contribute to eventual relapse. However, how LSC initiate relapse is not yet fully understood. We performed a retrospective study on bone marrow sections from AML patients during complete remission (CR), demonstrating that single and double immature myeloid precursors were located near endosteum and clustered precursors (≥ 3 cells/group) in intertrabecular region. Based on our observations, we hypothesize that after retrieval of myelotoxic regimen, LSC harboring near endosteum divide and differentiate into progeny cells which proliferate and then form colony clusters. Meanwhile, these clusters may migrate to intertrabecular region under the actions of cell migration factors. Without any interventions, clustered immature myeloid precursors may proliferate and hematologic relapse is then unavoidable.

[Real-time reverse-transcriptase polymerase chain reaction in monitoring BCR/ABL transcript levels in chronic myeloid leukemia patients after allogeneic hematopoietic stem cells transplantation].

OBJECTIVE: To monitor the gene expression of BCR/ABL in chronic myeloid leukemia patients after allogeneic hematopoietic stem cells transplantation with real-time RT-PCR for evaluating therapeutic efficacy and guiding further treatment. METHODS: Taqman real-time RT-PCR technique was performed to monitor peripheral blood BCR/ABL transcript levels in 10 patients at the time of diagnosis and serially at intervals in 25 patients after allogeneic hematopoietic stem cells transplantation. beta-actin mRNA was used as an endogenous reference. RESULTS: The sensitivity of real-time RT-PCR was 10 copies/microl. In 15 patients receiving transplantation, the median of N(BCR/ABL) (%) before transplantation, 1 month after transplantation and 2 months after transplantation was respectively: 6.57 (0.14 - 38.83), 0.10 (0 - 1.71) and 0 (0 - 0.52). 3 months later N(BCR/ABL) (%) all turned to 0. BCR/ABL transcripts after transplantation were detected to decrease gradually. N(BCR/ABL) (%) 1 month after transplantation and 2 months after transplantation was both lower than that before transplantation (chi(2) both = 13.07, P < 0.01). N(BCR/ABL) (%) at 2 months after transplantation was lower than that at 1 month after transplantation (chi(2) = 8.10, P < 0.01). In 10 other patients, N(BCR/ABL) (%) was 0 in consecutive tests from 3 to 43 months after transplantation. CONCLUSION: Real-time RT-PCR is sensitive, reliable and reproducible for monitoring CML relapse after transplantation. It is of help in detecting minimal residual disease, predicting the prognosis of the disease and providing practical indications for further treatment.