4,5-Dimethoxycanthin-6-one Inhibits Glioblastoma Stem Cell and Tumor Growth by Inhibiting TSPAN1 Interaction with TM4SF1.

Glioblastoma stem cells (GSCs) have been implicated in the self-renewal and treatment resistance of glioblastoma (GBM). Our previous study found that 4,5-dimethoxycanthin-6-one has the potential to inhibit GBM cell proliferation. This current study aims to elucidate the molecular mechanism underlying the effects of 4,5-dimethoxycanthin-6-one in GBM development. The effect of 4,5-dimethoxycanthin-6-one on GSC formation and differentiation was explored in human GBM cell lines U251 and U87. Subsequently, 4,5-dimethoxycanthin-6-one binding to tetraspanin 1 (TSPAN1) / transmembrane 4 L six family member 1 (TM4SF1) was analyzed by molecular simulation docking. Co-immunoprecipitation (Co-IP) and immunofluorescence (IF) were used to assess the interactions between TSPAN1 and TM4SF1 in GSCs. Cell proliferation was detected by cell counting kit-8 (CCK-8) and colony formation assay. To evaluate cell migration, invasion and apoptosis, we employed wound healing assay, transwell and flow cytometry, respectively. Furthermore, subcutaneous xenograft tumor models in nude mice were constructed to evaluate the impact of 4,5-dimethoxycanthin-6-one on GSCs in vivo by examining tumor growth and histological characteristics. 4,5-Dimethoxycanthin-6-one inhibited GSC formation and promoted stem cell differentiation in a concentration-dependent manner. Molecular docking models of 4,5-dimethoxycanthin-6-one with TM4SF1 and TSPAN1 were constructed. Then, the interaction between TSPAN1 and TM4SF1 in GSC was clarified. Moreover, 4,5-dimethoxycanthin-6-one significantly inhibited the expressions of TM4SF1 and TSPAN1 in vitro and in vivo. Overexpression of TSPAN1 partially reversed the inhibitory effects of 4,5-dimethoxycanthin-6-one on GSC formation, proliferation, migration and invasion. 4,5-Dimethoxycanthin-6-one inhibited GBM progression by inhibiting TSPAN1/TM4SF1 axis. 4,5-Dimethoxycanthin-6-one might be a novel and effective drug for the treatment of GBM.

WNTA5-mediated miR-374a-5p regulates vascular smooth muscle cell phenotype transformation and M1 macrophage polarization impacting intracranial aneurysm progression.

miR-374a-5p expression and localization in intracranial aneurysm (IA) tissues were detected, and its correlation with vascular smooth muscle cells (VSMCs) and macrophage markers was analyzed. Using platelet-derived growth factor-BB (PDGF-BB) induced VSMC model, elastase-induced IA rat model. Subsequently, miR-374a-5p was knocked down or overexpressed. We investigated the effects of miR-374a-5p on phenotypic conversion, and in vivo experiments were also carried out to verify the findings. The targeted relationship between miR-374a-5p and WNTA5 was analyzed. The effect of WNT5A inhibition on VSMC phenotypic transformation and THP-1-derived macrophage polarization was explored. Clinical studies have shown that miR-374a-5p was upregulated in IA patients. miR-374a-5p was negatively correlated with SM22α, α-SMA, CD206, and positively correlated with CD86. In vitro experiments showed that knocking down miR-374a-5p reversed the promotion of SM22α and α-SMA expression by PDGF-BB, while overexpression of miR-374a-5p had the opposite effect. In addition, knocking down miR-374a-5p also reversed the decrease in Calponin, TIMP3, TIMP4, and IL-10 levels caused by PDGF-BB, and further reduced the levels of MMP1, MMP3, MMP9, IL-1ß, IL-6, and TNF-α. These findings were further validated in vivo. In IA rats, there were notable increases in both systolic and diastolic blood pressure, along with an elevated M1/M2 ratio and the occurrence of vascular lesions. However, these symptoms were improved after knocking down miR-374a-5p. Furthermore, miR-374a-5p could target the WNT signals (WNT2B, WNT3, and WNT5A). miR-374a-5p regulated the VSMC phenotypic conversion and M1 macrophage polarization by targeting WNT5A, thereby impacting the progression of IA.

The Signal Characteristics of Oil and Gas Pipeline Leakage Detection Based on Magneto-Mechanical Effects.

In order to solve the problem of the quantification of detection signals in the magnetic flux leakage (MFL) of defective in-service oil and gas pipelines, a non-uniform magnetic charge model was established based on magnetic effects. The distribution patterns of magnetic charges under different stresses were analyzed. The influences of the elastic load and plastic deformation on the characteristic values of MFL signals were quantitatively assessed. The experimental results showed that the magnetic charge density was large at the edges of the defect and small at the center, and approximately decreased linearly with increasing stress. The eigenvalues of the axial and radial components of the MFL signals were compared, and it was found that the eigenvalues of the radial component exhibited a larger decline rate and were more sensitive to stress. With the increase in the plastic deformation, the characteristic values of the MFL signals initially decreased and then increased, and there was an inflection point. The location of the inflection point was associated with the magnetostriction coefficient. Compared with the uniform magnetic charge model, the accuracy of the axial and radial components of the MFL signals in the elastic stage of the improved magnetic charge model rose by 17% and 16%, respectively. The accuracy of the axial and radial components of the MFL signals were elevated by 9.15% and 9%, respectively, in the plastic stage.

Propagation Characteristics of Magnetic Tomography Method Detection Signals of Oil and Gas Pipelines Based on Boundary Conditions.

The magnetic tomography method (MTM) is a non-contact external inspection method for detecting metal magnetic memory signals. It has great potential for application in long-distance oil pipeline and subsea pipeline inspection. However, the spatial distribution characteristics and propagation laws of magnetic signals are not yet clear, which makes the MTM passive detection. In this study, a three-dimensional mathematical model of the magnetic field distribution of the stress concentration zone outside the pipe was established based on the boundary conditions. For the two cases in which the stress concentration zone was located at the top and bottom of the inner wall of the pipe, the model was solved by finite element analysis. The variation law of the magnetic signal outside the pipe was analyzed, and experiments were designed to verify the model. The results show that the shape of the magnetic memory signal remained unchanged after passing through the pipe wall. As the magnetic permeability of the pipe medium is much larger than that of air, the magnetic memory signal is significantly attenuated after penetrating the pipe wall. As the detection height increases, the magnetic induction outside the tube decays exponentially. The results also prove that the magnetic tomography method can detect the stress concentration zone at any position of the pipeline, and the detection accuracy is higher when it is located at the top of the pipeline.

A pilot study: Gut microbiota, metabolism and inflammation in hypertensive intracerebral haemorrhage.

AIMS: In recent years, the incidence rate of hypertensive intracerebral haemorrhage (HICH) has been increasing, accompanied by high mortality and morbidity, which has brought a heavy burden to the social economy. However, the pathogenesis of HICH is still unclear. This study intends to explore the mechanism of gut microbiota metabolism and inflammation in the process of HICH to provide a theoretical basis for the diagnosis and treatment of HICH. METHODS AND RESULTS: HE staining showed that the brain tissues of model group had obvious oedema injury, which indicated that the HICH model was successfully constructed. ELISA analysis showed that IL-1ß and TNF-α levels in blood and brain tissues were significantly increased, and IL-10 level was significantly decreased in blood. IHC analysis showed that microglia and macrophages were activated in the model group. 16S rRNA sequence showed that the diversity of gut microbiota in HICH patients decreased. Also, the microbiota belonging to Firmicutes, Proteobacteria and Verrucomicrobia changed significantly. LC-MS/MS analysis showed that the metabolic phenotype of HICH patients changed. Also, the 3,7-dimethyluric acid- and 7-methylxanthine-related metabolic pathways of caffeine metabolism pathways were downregulated in patients with HICH. Bacteroides was negatively correlated with the IL-1ß and TNF-α levels. Blautia was negatively correlated with the IL-1ß and TNF-α levels, and positively correlated with the IL-10 level. Akkermansia was negatively correlated with the 3,7-dimethyluric acid and 7-methylxanthine. CONCLUSION: Our study suggested that HICH was accompanied by the increased inflammation marker levels in peripheral blood and brain, decreased gut microbiota diversity, altered gut metabolic phenotype and downregulation of caffeine metabolism pathway. SIGNIFICANCE AND IMPACT OF THE STUDY: Our study reported that HICH accompanied by the increased inflammation, decreased gut microbiota diversity and altered gut metabolic phenotype. Due to the number of patients, this work was a pilot study.

Nursing Practice Based on Evidence-Based Concepts to Prevent Enteral Nutrition Complications for Critically Ill Neurosurgical Patients.

Purpose: To explore the practical value of enteral nutrition care guided by evidence-based concepts in preventing enteral nutritional complications in critically ill neurosurgical patients. Methods: Three hundred critically ill patients from March 2020 to October 2021 from our neurosurgery department were included in the study. Patients were divided into a control group (March to December 2020, n = 150) and a study group (January to October 2021, n = 150) according to the order of their admission. The control group received conventional enteral nutrition care, and the study group received enteral nutrition care based on evidence-based concept guidance. The levels of serum nutritional indicators [hemoglobin (Hb), albumin (ALB), and total protein (TP)], feeding compliance rate, the incidence of complications (gastric retention, bloating, diarrhea, reflux, vomiting, aspiration, stress ulcers, etc.), and prognosis during the observation period were compared between the two groups. The scores of the questionnaire of knowledge, attitude, and practice on nutrition among neurosurgical nurses before and after the implementation of evidence-based care were compared among nursing staff in the study group. Results: At 1 and 2 weeks after enrollment, Hb, ALB, and TP levels were lower in both groups than before enrollment in the same group (P < 0.05). At 2 weeks after enrollment, Hb, ALB, and TP levels were higher in both groups than at 1 week after enrollment in the same group (P < 0.05). At 1 and 2 weeks after enrollment, Hb, ALB, and TP levels were higher in the study group than in the control group (P < 0.05). At 7 days after feeding, the feeding compliance rate was higher in the study group (94.67%) than in the control group (70.00%) (P < 0.05). The total complication rate was lower in the study group (8.00%) than in the control group (16.00%) (P < 0.05). The percentage of good prognosis was higher in the study group (34.00%) than in the control group (23.33%) (P < 0.05). After the implementation of evidence-based care, caregivers in the study group scored higher on nutrition knowledge, nutrition attitudes, and nutrition practices than those before the implementation (P < 0.05). Conclusion: The implementation of evidence-based nursing interventions in critically ill neurosurgical patients based on evidence-based concepts is of great clinical value in correcting their nutritional status, preventing enteral nutritional complications, improving prognosis, and enhancing the nutritional knowledge, attitudes, and practices of nursing staff.

4,5-Dimethoxycanthin-6-one is a novel LSD1 inhibitor that inhibits proliferation of glioblastoma cells and induces apoptosis and pyroptosis.

BACKGROUND: Glioblastoma is one of the most common fatal intracranial malignancies. Lysine-specific demethylase 1 (LSD1) reportedly has therapeutic effects on a variety of tumors. This study explored the therapeutic effect of LSD1 inhibition on glioblastoma cell lines and the possible underlying mechanisms. METHODS: The MTT assay was utilized to screen for the sensitivity of U87, U251 and T98G cells to 4, 5-dimethoxycarrageenin-6-one. qRT-PCR and western blot were used to measure the proliferation, apoptosis, and pyroptosis signaling pathway expression to observe the effect of LSD1 inhibition on U251 and T98G cells. Flow cytometry, immunofluorescence, immunohistochemistry, wound scratch, clone formation, and TUNEL assay were used to analyze the effects of 4, 5-dimethoxycanthin-6-one on glioblastoma cells. The effect of 4, 5-dimethoxycanthin-6-one was examined in vivo in BALB/c nude mice injected with U251 cells. HE staining was used to detect the histopathology of the tumor. RESULTS: LSD1 specifically catalyzes the demethylation of monomethylated and demethylated histone H3 lysine at position 4 (h3k4me1, h3k4me2, h3k4me3) and lysine at position 9 (h3k9me1). This regulated the transcriptional activity of proliferation, apoptosis, and pyroptosis signaling pathway genes. In vitro, the proliferation of glioblastoma cells was decreased in the 4, 5-dimethoxycanthin-6-one group. The expression of Caspase1 in glioblastoma cells treated with 4, 5-dimethoxycanthin-6-one increased, and the number of apoptotic cells increased. The tumor volume of mice injected with 4, 5-dimethoxycanthin-6-one decreased significantly. CONCLUSION: 4, 5-Dimethoxycanthin-6-one could act as a novel inhibitor of LSD1 to regulate glioblastoma, which could inhibit the proliferation of U251 and T98G cells and induce their apoptosis and pyroptosis. It is a potential drug for the treatment of glioblastoma.

A stomach like a utility room: Case report.

BACKGROUND: Foreign body (FB) ingestion is an emergency that is more common in children and adults with mental disorders. A wide array of FBs can be ingested, and most of them do not need to be treated. However, if the FB blocks the digestive tract or causes damage, it needs to be removed by endoscopy or even surgery.We describe here a stomach full of FBs, but these FBs did not cause serious damage. CASE PRESENTATION: A 9-year-old male child with mental retardation suffered abdominal pain after swallowing FBs. X-ray and computed tomography (CT) found a large number of FBs of different shapes. We tried to remove them under endoscopy but failed; we then changed to laparotomy and removed a large number of FBs. The patient started normal feeding on the 4th day and was discharged home. CONCLUSIONS: FB ingestion is very common. Symptoms are used to determine whether further treatment, which is usually feasible, is required. However, for patients who cannot accurately describe the ingestion of FBs, such as children, patients with mental disorders, and patients who are inebriated, FBs should still be treated with caution, especially when the clinical symptoms and related examinations are not typical, and adequate plans should be made, as shown in this case. There may be unexpected discoveries.

MiR-214 regulates CD3ζ expression in T cells.

INTRODUCTION: T-cell activation requires the T-cell receptor (TCR)-CD3 complex, which integrates and transduces signals. CD3ζ plays a vital role in TCR signalling by mediating T-cell activation. Abnormal CD3ζ expression is a common characteristic of haematological malignancies with T-cell immune dysfunction or autoimmune diseases. Targeted regulation of CD3ζ expression by either direct or indirect approaches is important for regulating T-cell activation. AIM OF THE STUDY: In this study, we focused on identifying miRNAs that may regulate CD3ζ expression. MATERIAL AND METHODS: Three microRNA target search algorithms (TargetScan, PicTar, and microrna.org) were used to identify hypothetical miRNAs that target CD3ζ in T cells. Of the predicted miRNAs, miR-214 was chosen and validated to determine whether miR-214 directly binds to the CD3ζ 3'-UTR and regulates CD3ζ expression by luciferase reporter assays, real-time PCR, and Western blotting. RESULTS: The results indicate that miR-214 specifically binds the CD3ζ 3'-UTR, and miR-214 mimics remarkably reduce the expression of CD3ζ in MOLT-4 cells. CONCLUSIONS: We identify for the first time that miR-214 targets expression in MOLT-4 cells, suggesting that miR-214 might negatively regulate T-cell activation by targeting CD3ζ.

Increased exhausted CD8+ T cells with programmed death-1, T-cell immunoglobulin and mucin-domain-containing-3 phenotype in patients with multiple myeloma.

AIM: The immunosuppressive microenvironment plays a crucial role in T-cell immunodeficiency in multiple myeloma (MM). Overexpression of T-cell immunosuppressive receptors, including programmed death-1 (PD-1) and T-cell immunoglobulin and mucin-domain-containing-3 (Tim-3), may be related to tumor immunosuppression and poor prognosis, and the malignant bone marrow (BM) microenvironment may contribute to such immunosuppression. The purpose of this study was to analyze the distribution of PD-1+ and/or Tim-3+ T cells in different T-cell subset in patients with MM. METHODS: The expression of PD-1 and Tim-3 with exhausted (CD244+ and CD57+ ) CD3+ , CD4+ and CD8+ T cells between BM and peripheral blood (PB) from 10 patients with untreated MM was detected by multicolor flow cytometry assay. RESULTS: A significant increase in both PD-1+ CD57+ and Tim-3+ CD57+ CD3+ T cells and PD-1+ Tim-3+ CD3+ T cells was detected in PB from patients with MM compared with 10 healthy individuals (HIs), and the alteration was mostly in the CD8+ T-cell subset. Significant higher percentage of PD-1+ CD3+ T cells was found in BM compared with PB from patients with MM. The level of PD-1+ Tim-3+ CD3+ , CD4+ , and CD8+ T cells was high in BM group compared with PB. Moreover, PD-1+ CD244+ or PD-1+ CD57+ CD3+ T cells, particularly PD-1+ CD244+ and PD-1+ CD57+ CD8+ T cells were significantly higher in BM than in PB. In addition, limited dynamic detection data from three MM cases who achieved complete remission after treatment showed that the numbers of either PD-1+ or PD-1+ Tim-3+ T cells in different T-cell subsets were decreased in both BM and PB. CONCLUSION: We characterized the distribution of PD-1 and TIM-3 concurrent with exhausted CD3+ , CD4+ and CD8+ T cells between BM and PB from patients with MM. Higher numbers of PD-1+ CD244+ or PD-1+ CD57+ CD3+ T cells in BM from patients with MM may contribute to mediate the BM immunosuppressive microenvironment. Although heterogeneous alterations in Tim-3+ T cells may represent a complex immunosuppressive pattern in MM. Overall, higher levels of PD-1+ CD244+ or PD-1/Tim-3+ CD57+ CD8+ T cells may be a major reason for lower T-cell activation and T-cell immunodeficiency in MM.

Downregulated miR-17, miR-29c, miR-92a and miR-214 may be related to BCL11B overexpression in T cell acute lymphoblastic leukemia.

AIM: BCL11B overexpression is a characteristic of most T cell acute lymphoblastic leukemia (T-ALL) cases, and downregulated BCL11B in leukemic T cells inhibits cell proliferation and induces apoptosis. The purpose of this study was to analyze the miRNA expression pattern that may be related to BCL11B regulation in T-ALL. METHODS: Quantitative real-time PCR was used to detect the miRNAs miR-17-3p, miR-17-5p, miR-29c-3p, miR-92a-3p, miR-214-3p and miR-214-5p, the BCL11B expression level in peripheral blood mononuclear cells which was obtained from 17 de novo and untreated T-ALL patients, and 15 healthy individuals (HIs) served as control. Correlations between the relative miRNA expression levels and BCL11B were analyzed. RESULTS: Based on the computational prediction that certain miRNAs bind the BCL11B 3'-UTR, miR-17-3p, miR-17-5p, miR-29c-3p, miR-92a-3p, miR-214-3p and miR-214-5p were found to be candidates for regulating BCL11B. The expression levels of the six miRNAs were decreased compared with HIs, and with the exception of miR-17-5p, statistically significant differences in expression levels were found in the T-ALL group. Moreover, while significantly higher BCL11B expression was found in the T-ALL group, a negative trend in the correlation level for all six miRNAs could be found in all groups; however, statistical significance was only found for miR-214-3p in the T-ALL group. CONCLUSION: miRNA downregulation together with BCL11B upregulation suggests that miR-17, miR-29c, miR-92a and miR-214 might be involved in BCL11B regulation. The therapeutic promise of regulating the expression of these miRNAs for T-ALL therapy may be considered in the future.

Effect of external periodic signals and electromagnetic radiation on autaptic regulation of neuronal firing.

An improved Hindmarsh-Rose (HR) neuron model, where the memristor is a bridge between membrane potential and magnetic flux, can be used to investigate the effect of periodic signals on autaptic regulation of neurons under electromagnetic radiation. Based on the improved HR model driven by periodic high-low-frequency current and electromagnetic radiation, the responses of electrical autaptic regulation with diverse high-low-frequency signals are investigated using bifurcation analysis. It is found that the electrical modes of neurons are determined by the selecting parameters of both periodic high and low-frequency current and electromagnetic radiation, and the Hamiltonian energy depends on the neuronal firing modes. The effects of Gaussian white noise on the membrane potential are discussed using numerical simulations. It is demonstrated that external high-low-frequency stimulus plays a significant role in the autaptic regulation of neural firing mode, and the electrical mode of neurons can be affected by the angular frequency of both high-low-frequency forcing current and electromagnetic radiation. The mechanism of neuronal firing regulated by high-low-frequency signal and electromagnetic radiation discussed here could be applied to research neuronal networks and synchronisation modes.

Different genetic alteration of A20 in a Sézary syndrome case with Vα2-Jα22 T cell clone.

BACKGROUND: The comprehensive genetic alterations underlying the pathogenesis of Sézary syndrome (SS) remains largely unknown. Previous studies showed that alterations of tumor necrosis factor-α-induced protein 3 gene (TNFAIP3; A20) are frequent in SS. In this study, we characterized the mutation and polymorphisms of A20 in a case with SS and compared with the genetic feature of A20 in T-cell acute lymphoblastic leukemia (T-ALL). METHODS: Using a novel approach based on the combination of fine-tiling array comparative genomic hybridization ( and ligation-mediated polymerase chain reaction (LM-PCR) to identify SS clone, the polymorphisms in the A20 gene (promoter, exons 2-9 [coding region] and 3'UTR) were detected by PCR and sequencing. RESULTS: The malignant SS clone was identified as TCR Vα2-Jα22 rearrangement without deletion at the A20 loci (6q23-27 region) in the SS case. Six polymorphisms were identified, all of them are belonging to single nucleotide polymorphisms (SNPs) that are recorded in genebank: rs5029924, rs5029937, rs2230926, rs582757 and rs77191406, while rs2307859 was not identified in the SS sample, which is found in all T-ALL. The alteration pattern of A20 in this case seemed different from the T-ALL samples, in contrast, it is similar to the alteration of A20 in samples from rheumatoid arthritis or systemic lupus erythematosus with poor clinical outcome and cancer developing. CONCLUSIONS: The genetic alteration of A20 in the SS case was different from the T-ALL samples and similar to the cases with refractory autoimmune disease and related to tumorigenesis. The findings lead to discuss whether such SNPs of A20 may link the refractory autoimmune inflammation and the tumorigenesis.

Re-balance of memory T cell subsets in peripheral blood from patients with CML after TKI treatment.

T cell immune surveillance is considered an important host protection process for inhibiting carcinogenesis. The full capacity of T cell immune surveillance is dependent on T cell homeostasis, particularly for central memory T (TCM) cells and stem cell memory T (TSCM) cells. In this study, distribution of T cell subsets in peripheral blood from 12 patients with chronic myeloid leukemia (CML) and 12 cases with CML in complete remission (CR) was analyzed using a multicolor flow cytometer, and 16 samples from healthy individuals (HIs) served as control. The proportion of CD8+ TSCM and CD4+ and CD8+ TCM cells were lower, while CD4+ effector memory T (TEM) cells and CD4+ and CD8+ terminal effector T (TEF) cells were higher in CML patients compared with HIs. Moreover, the proportion of CD8+CD28- T cells, which were found to have the immune suppressive function, increased in the naive T (TN) cell and TCM subsets in CML patients compared with HIs. Our study reveals that elimination of leukemia cells by treating with tyrosine kinase inhibitors (TKIs) restores the memory T cell distribution from a skewed pattern in CML patients who are under leukemia burden, indicating that leukemia-specific immune responses mediated by T cells might be induced and maintained in CML patients, however, these responsive T cells might gradually become exhausted due to the continued existence of leukemia cells and their environment; therefore, T cell activation using a different approach remains a key point for enhancing global T cell immunity in CML patients, even for those with CR status.

Higher PD-1 expression concurrent with exhausted CD8+ T cells in patients with de novo acute myeloid leukemia.

OBJECTIVE: To investigate the association between the T cell inhibitory receptor programmed death 1 (PD-1) and T cell exhaustion status in T cells from patients with de novo acute myeloid leukemia (AML) and AML in complete remission (CR). METHODS: Surface expression of PD-1 and the exhaustion and immunosenescence markers CD244 and CD57 on CD3+, CD4+ and CD8+ T cells from peripheral blood samples from 20 newly diagnosed, untreated AML patients and 10 cases with AML in CR was analyzed by flow cytometry. Twenty-three healthy individuals served as control. RESULTS: A significantly higher percentage of PD-1+ cells were found for CD3+ T cells in the de novo AML group compared with healthy controls. In addition, an increased level of PD-1+CD8+ T cells, but not PD-1+CD4+, was found for CD3+ T cells in the de novo AML and AML-CR samples. A higher percentage of CD244+CD4+, CD244+CD8+, CD57+CD4+ and CD57+CD8+ T cells was found in CD3+ T cells in samples from those with de novo AML compared with those from healthy controls. Strong increased PD-1+CD244+ and PD-1+CD57+ co-expression was found for CD4+ and CD8+ T cells in the de novo AML group compared with healthy controls. CONCLUSIONS: We characterized the major T cell defects, including co-expression of PD-1 and CD244, CD57-exhausted T cells in patients with de novo AML, and found a particular influence on CD8+ T cells, suggesting a poor anti-leukemia immune response in these patients.

A kinetic model of multiple phenotypic states for breast cancer cells.

Quantitative modeling of microscopic genes regulatory mechanisms in an individual cell is a crucial step towards understanding various macroscopic physiological phenomena of cell populations. Based on the regulatory mechanisms of genes zeb1 and cdh1 in the growth and development of breast cancer cells, we propose a kinetic model at the level of single cell. By constructing the effective landscape of underlying stationary probability for the genes expressions, it is found that (i) each breast cancer cell has three phenotypic states (i.e., the stem-like, basal, and luminal states) which correspond to three attractions of the probability landscape. (ii) The interconversions between phenotypic states can be induced by the noise intensity and the property of phenotypic switching is quantified by the mean first-passage time. (iii) Under certain conditions, the probabilities of each cancer cell appearing in the three states are consistent with the macroscopic phenotypic equilibrium proportions in the breast cancer SUM159 cell line. (iv) Our kinetic model involving the TGF-ß signal can also qualitatively explain several macroscopic physiological phenomena of breast cancer cells, such as the "TGF-ß paradox" in tumor therapy, the five clinical subtypes of breast cancer cells, and the effects of transient TGF-ß on breast cancer metastasis.

Salvia miltiorrhiza Bunge (Danshen) extract attenuates permanent cerebral ischemia through inhibiting platelet activation in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Danshen is a crude herbal drug isolated from dried roots of Salvia miltiorrhiza Bunge. This plant is widely used in oriental medicine for the treatment of cardiovascular and cerebrovascular diseases. The supercritical CO2 extract from Danshen (SCED) (57.85%, 5.67% and 4.55% for tanshinone IIA, tanshinone I and cryptotanshinone respectively) was studied in this article, whose potential molecular mechanism remains unclear, especially in anti-thrombosis. AIM OF THE STUDY: The present study was designed to observe the protective effect of SCED on ischemic stroke in rats and to explore the underlying anti-thrombosis mechanism. MATERIALS AND METHODS: Following induction of cerebral ischemia in rats by permanent middle cerebral artery occlusion (pMCAO). Neurological defect score, cerebral blood flow, infarct size, and brain edema were measured to evaluate the injury. Arteriovenous shunt thrombosis model and adenosine 5'-diphosphate (ADP) induced acute pulmonary embolism model were conducted to estimate the antithrombotic effect of SCED. In order to investigate the effects of SCED on platelet aggregation, rat platelet-rich-plasma (PRP) were incubated with SCED prior to the addition of the stimuli (ADP or 9, 11-dideoxy-11α, 9α-epoxymethanoprostaglandin F2α (U46619)). Aggregation was monitored in a light transmission aggregometer. Inhibitory effect of SCED on thromboxane A2 (TXA2) release was detected by ELISA kit. Phospholipase C (PLC)/ Protein kinase C (PKC) signaling pathway was analyzed by a Western blot technique. The effect of the SCED was also studied in vivo on bleeding time in mice. RESULTS: SCED improved the neurological defect score, increased cerebral blood flow, reduced infarct size and alleviated brain edema in rats exposed to pMCAO. After administration of SCED, thrombosis formation in arteriovenous shunt was inhibited and recovery time in pulmonary embolism was shortened. The inhibitory effect of SCED on platelet activation was further confirmed by TXB2 ELISA kit and Western blot analysis of PLC/PKC signaling pathway. CONCLUSIONS: SCED attenuates cerebral ischemic injury. The possible mechanism is that SCED inhibits thrombosis formation, platelet aggregation and activation of PLC/PKC pathway. On this basis, this new extract could be a promising agent to inhibit thrombosis formation and protect against cerebral ischemia injury.

[Changes of TRGV genes and prognosis-related chemokine/receptor gene expressions in T cell lymphoma].

Objective To analyze the expressions of T cell receptor γ subfamily variable region I, II, and III genes (TRGV I~III) and prognosis-related chemokines and their receptor genes (CCL20, CCR6, CCL17, CCL22 and CCR4) in the peripheral blood of T-cell lymphoma (non-γδ T cell type) patients, and to investigate the correlations between γδ T cells and chemokines/receptors and their relevance to disease prognosis. Methods Peripheral blood samples were collected from 27 patients with T-cell lymphoma (non-γδ T cell type) and 9 healthy individuals as controls. Expressions of TRGV I~III subfamily genes and prognosis-related chemokines/receptors (CCL20/CCR6 and CCL17/CCL22/CCR4) in the peripheral blood of patients with T-cell lymphoma and normal controls were detected by quantitative real-time PCR. Results Compared with the normal controls, the expression levels of TRGV I-III subfamily genes of patients at the stages of initial onset, relapse/refractoriness and complete remission (CR) were significantly higher, and the expression patterns of TRGV subfamilies were changed. The expression levels of CCL22 and CCR4 in initial onset group and relapse/refractory group were significantly higher than that in normal controls, while CCL17 expression was significantly lower than that in normal controls. There were unique correlation patterns in the expressions of TRGV subfamily genes and chemokine/receptor genes in the patients at the stages of initial onset, CR and relapse/refractoriness. There are low expressions of TRGV II subfamilies in newly diagnosed and relapse/refractory T-cell lymphoma patients, and their expressions are elevated after treatment. Conclusion TRGV II subfamilies might be the T cell functional subset against T-cell lymphoma, and might be associated with the outcome of the disease. And the high expression of CCL22/CCR4 might be related to the pathogenesis of T-cell lymphoma.

Oligoclonal expansion of TCR Vδ T cells may be a potential immune biomarker for clinical outcome of acute myeloid leukemia.

BACKGROUND: Recent data have shown that γδ T cells can act as mediators for immune defense against tumors. Our previous study has demonstrated that persisting clonally expanded TRDV4 T cells might be relatively beneficial for the outcome of patients with T cell acute lymphoblastic leukemia after hematopoietic stem cell transplantation (HSCT). However, little is known about the distribution and clonality of the TRDV repertoire in T cell receptor (TCR) of γδ T cells and their effects on the clinical outcome of patients with acute myeloid leukemia (AML). The aim of this study was to assess whether the oligoclonal expansion of TCR Vδ T cells could be used as an immune biomarker for AML outcome. FINDINGS: γδ T cells were sorted from the peripheral blood of 30 patients with untreated AML and 12 healthy donors. The complementarity-determining region 3 (CDR3) sizes of eight TCR Vδ subfamily genes (TRDV1 to TRDV8) were analyzed in sorted γδ T cells using RT-PCR and GeneScan. The most frequently expressed TRDV subfamilies in the AML patients were TRDV8 (86.67 %) and TRDV2 (83.33 %), and the frequencies for TRDV1, TRDV3, TRDV4, and TRDV6 were significantly lower than those in healthy individuals. The most frequent clonally expanded TRDV subfamilies in the AML patients included TRDV8 (56.67 %) and TRDV4 (40 %). The clonal expansion frequencies of the TRDV2 and TRDV4 T cells were significantly higher than those in healthy individuals, whereas a significantly lower TRDV1 clonal expansion frequency was observed in those with AML. Moreover, the oligoclones of TRDV4 and TRDV8 were independent protective factors for complete remission. Furthermore, the oligoclonal expansion frequencies of TRDV5 and TRDV6 in patients with relapse were significantly higher than those in non-recurrent cases. CONCLUSIONS: To the best of our knowledge, we characterized for the first time a significant alteration in the distribution and clonality of the TRDV subfamily members in γδ T cells sorted from AML patients. Clonally expanded TRDV4 and TRDV8 T cells might contribute to the immune response directed against AML, while oligoclonal TRDV5 and TRDV6 might occur in patients who undergo relapse. While the function of such γδ T cell clones requires further investigation, TRDV γδ T cell clones might be potential immune biomarkers for AML outcome.

Generation of V α13/ß21+T cell specific target CML cells by TCR gene transfer.

Adoptive immunotherapy with antigen-specific T cells can be effective for treating melanoma and chronic myeloid leukemia (CML). However, to obtain sufficient antigen-specific T cells for treatment, the T cells have to be cultured for several weeks in vitro, but in vitro T cell expansion is difficult to control. Alternatively, the transfer of T cell receptors (TCRs) with defined antigen specificity into recipient T cells may be a simple solution for generating antigen-specific T cells. The objective of this study was to identify CML-associated, antigen-specific TCR genes and generate CML-associated, antigen-specific T cells with T cell receptor (TCR) gene transfer. Our previous study has screened an oligoclonal Vß21 with a different oligoclonal Vα partner in peripheral blood mononuclear cells (PBMCs) derived from patients with CML. In this study, oligoclonally expanded TCR α genes, which pair with TCR Vß21, were cloned into the pIRES eukaryotic expression vector (TCR Vα-IRES-Vß21). Next, two recombinant plasmids, TCR Vα13-IRES-Vß21 and TCR Vα18-IRES-Vß21, were successfully transferred into T cells, and the TCR gene-modified T cells acquired CML-specific cytotoxicity with the best cytotoxic effects for HLA-A11+ K562 cells observed for the TCR Vα13/Vß21 gene redirected T cells. In summary, our data confirmed TCRVα13/Vß21 as a CML-associated, antigen-specific TCR. This study provided new evidence that genetically engineered antigen-specific TCR may become a druggable approach for gene therapy of CML.