[Significance of Tim-3 and Its Ligand Galectin-9 in Th1/Th2 Imbalance in Patients with Multiple Myeloma].

OBJECTIVE: To investigate the significance of Tim-3 and Galectin-9 in Th1/Th2 imbalance in patients with multiple myeloma (MM). METHODS: 55 newly diagnosed MM patients and 20 healthy controls were included. Flow cytometry was used to detect the expression of Tim-3 on CD4+T cells, the proportion of Th1, Th2, Tim-3+Th1 and Tim-3+Th2 cells in peripheral blood. ELISA was used to detect the levels of cytokines IFN-γ and IL-4 in serum, and PCR was used to detect the level of Galectin-9 mRNA. Then the correlations between Galectin-9 mRNA expression and Th-cell subsets and related cytokine levels, as well as the relationship between Tim-3+Th1/Tim-3+Th2 ratio and corresponding clinical features were analyzed. RESULTS: Compared with the control group, the expression of Tim-3 on CD4+T cells in peripheral blood of MM patients was significantly increased (P<0.05), the proportions of Tim-3+Th1 cells, Tim-3+Th2 cells and Tim-3+Th1/Tim-3+Th2 ratio in MM patients were also increased (P<0.05), while the proportion of Th1 cells and Th1/Th2 ratio in MM patients were significantly decreased (P<0.05). The level of cytokine IFN-γ and IFN-γ/IL-4 ratio in MM patients were significantly decreased (P<0.05), while the level of cytokine IL-4 was increased (P<0.05). The mRNA levels of Galectin-9 in MM patients were significantly increased (P<0.05). The levels of Galectin-9 mRNA were positively correlated with Tim-3+CD4+T cells (r=0.663), Tim-3+Th2 cells (r=0.492) and IL-4 (r=0.470), while negatively correlated with IFN-γ (r=-0.593). The ratios of Tim-3+Th1/Tim-3+Th2 in MM patients were positively correlated with ISS stage (r=0.511), osteolytic damage (r=0.556) and chromosome abnormality (r=0.632). CONCLUSION: These results suggest that Tim-3 and Galectin-9 are involved in Th1/Th2 imbalance in MM patients, and the high ratio of Tim-3+Th1/Tim-3+Th2 is associated with poor clinical prognosis.

α-Mangostin Inhibited M1 Polarization of Macrophages/Monocytes in Antigen-Induced Arthritis Mice by Up-Regulating Silent Information Regulator 1 and Peroxisome Proliferators-Activated Receptor γ Simultaneously.

Background: α-Mangostin (MG) showed the potentials in alleviating experimental arthritis, inhibiting inflammatory polarization of macrophages/monocytes, and regulating peroxisome proliferators-activated receptor γ (PPAR-γ) and silent information regulator 1 (SIRT1) signals. The aim of this study was to analyze the correlations among the above-mentioned properties. Methods: Antigen-induced arthritis (AIA) was established in mouse, which was treated with MG in combination with SIRT1/PPAR-γ inhibitors to clarify the role of the two signals in the anti-arthritic actions. Pathological changes were systematically investigated. Phenotypes of cells were investigated by flow cytometry. Expression and co-localization of SIRT1 and PPAR-γ proteins in joint tissues were observed by the immunofluorescence method. Finally, clinical implications from the synchronous up-regulation of SIRT1 and PPAR-γ were validated by experiments in vitro. Results: SIRT1 and PPAR-γ inhibitors (nicotinamide and T0070097) reduced the therapeutic effects of MG on AIA mice, and abrogated MG-induced up-regulation of SIRT1/PPAR-γ and inhibition of M1 polarization in macrophages/monocytes. MG has a good binding affinity to PPAR-γ, and MG promoted the co-expression of SIRT1 and PPAR-γ in joints. Synchronously activating SIRT1 and PPAR-γ was revealed to be necessary by MG to repress inflammatory responses in THP-1 monocytes. Conclusion: MG binds PPAR-γ and excites this signaling to initiate ligand-dependent anti-inflammatory activity. Due to certain unspecified signal transduction crosstalk mechanism, it then promoted SIRT1 expression and further limited inflammatory polarization of macrophages/monocytes in AIA mice.

ent-Pimarane Diterpenes from the Aerial Parts of Siegesbeckia pubescens.

Five new ent-pimarane diterpenes (1-5) and five known analogs (6-10) were isolated from the aerial parts of Siegesbeckia pubescens. Their structures, including absolute configurations, were determined by comprehensive spectroscopic methods especially 1D and 2D NMR and quantum chemical electronic circular dichroism calculations. All the isolated compounds were evaluated for their cytotoxicity against human BT549, A549 and H157 cancer cell lines. Among them, compounds 1 and 2 showed mild cytotoxicity against lung cancer cell lines H157 with IC50 values of 16.35±2.59 and 18.86±4.83 µM, respectively.

Rapid and efficient isolation platform for plasma extracellular vesicles: EV-FISHER.

Extracellular vesicles (EVs) have found diverse applications in clinical theranostics. However, the current techniques to isolate plasma EVs suffer from burdensome procedures and limited yield. Herein, we report a rapid and efficient EV isolation platform, namely, EV-FISHER, constructed from the metal-organic framework featuring cleavable lipid probes (PO4 3- -spacer-DNA-cholesterol, PSDC). The EV-FISHER baits EVs from plasma by cholesterol and separates them with an ordinary centrifuge. The captured EVs could be released and collected upon subsequent cleavage of PSDC by deoxyribonuclease I. We conclude that EV-FISHER dramatically outperforms the ultracentrifugation (UC) in terms of time (∼40 min vs. 240 min), isolation efficiency (74.2% vs. 18.1%), and isolation requirement (12,800 g vs. 135,000 g). In addition to the stable performance in plasma, EV-FISHER also exhibited excellent compatibility with downstream single-EV flow cytometry, enabling the identification of glypican-1 (GPC-1) EVs for early diagnosis, clinical stages differentiation, and therapeutic efficacy evaluation in breast cancer cohorts. This work portrays an efficient strategy to isolate EVs from complicated biological fluids with promising potential to facilitate EVs-based theranostics.

Direct Targeting of CREB1 with Imperatorin Inhibits TGFß2-ERK Signaling to Suppress Esophageal Cancer Metastasis.

Metastasis accounts for 90% of cancer death worldwide, and effective therapeutic strategies are lacking. The aim of this work is to identify the key drivers in tumor metastasis and screen therapeutics for treatment of esophageal squamous cell carcinoma (ESCC). Gene Ontology analysis of The Cancer Genome Atlas (TCGA) gene expression datasets of ESCC patients with or without lympy metastasis identifies that TGFß2 is highly enriched in the pathways essential for tumor metastasis and upregulates in the metastatic ESCC tumors. High TGFß2 expression in ESCC correlates with metastasis and patient survival, and functionally contributes to tumor metastasis via activating extracellular signal-regulated kinases (ERK) signaling. By screening of a library consisting of 429 bioactive compounds, imperatorin is verified as a novel TGFß2 inhibitor, with robustly suppressive effect on tumor metastasis in multiple mice models. Mechanistically, direct binding of imperatorin and CREB1 inhibits phosphorylation, nuclear translocation of CREB1, and its interaction with TGFß2 promoter, represses TGFß2 expression and fibroblasts-secreted CCL2, and then inactivates ERK signaling to block cancer invasion and abrogates the paracrine effects of fibroblasts on tumor angiogenesis and metastasis. Overall, the findings suggest the use of TGFß2 as a diagnostic and prognostic biomarker and therapeutic target in ESCC, and supports the potential of imperatorin as a novel therapeutic strategy for cancer metastasis.

Drp1 is widely, yet heterogeneously, distributed in the mouse central nervous system.

OBJECTIVES: Drp1 is widely expressed in the mouse central nervous system and plays a role in inducing the mitochondrial fission process. Many diseases are associated with Drp1 and mitochondria. However, since the exact distribution of Drp1 has not been specifically observed, it is difficult to determine the impact of anti-Drp1 molecules on the human body. Clarifying the specific Drp1 distribution could be a good approach to targeted treatment or prognosis. METHODS: We visualized the distribution of Drp1 in different brain regions and explicated the relationship between Drp1 and mitochondria. GAD67-GFP knock-in mice were utilized to detect the expression patterns of Drp1 in GABAergic neurons. We also further analyzed Drp1 expression in human malignant glioma tissue. RESULTS: Drp1 was widely but heterogeneously distributed in the central nervous system. Further observation indicated that Drp1 was highly and heterogeneously expressed in inhibitory neurons. Under transmission electron microscopy, the distribution of Drp1 was higher in dendrites than other areas in neurons, and only a small amount of Drp1 was localized in mitochondria. In human malignant glioma, the fluorescence intensity of Drp1 increased from grade I-III, while grade IV showed a declining trend. CONCLUSION: In this study, we observed a wide heterogeneous distribution of Drp1 in the central nervous system, which might be related to the occurrence and development of neurologic disease. We hope that the relationship between Drp1 and mitochondria may will to therapeutic guidance in the clinic.

CP-25 inhibits PGE2-induced angiogenesis by down-regulating EP4/AC/cAMP/PKA-mediated GRK2 translocation.

G protein-coupled receptor kinase 2 (GRK2), a type of cytosolic enzyme, transiently translocates to the plasma membrane upon G protein-coupled receptors (GPCRs) activation, and it also binds to extracellular signal-regulated kinase (ERK) to inhibit the activation of ERK. GRK2 deficiency in endothelial cells (ECs) leads to increased pro-inflammatory signaling and promotes recruitment of leukocytes to activated ECs. However, the role of GRK2 in regulating angiogenesis remains unclear. Here, we show that GRK2 is a novel regulatory molecule on migration and tube formation of ECs, vessel sprouting ex vivo and angiogenesis in vivo. We identify that EP4/AC/cAMP/protein kinase A (PKA)-mediated GRK2 translocation to cells membrane decreases the binding of GRK2 and ERK1/2 to inhibit ERK1/2 activation, which promotes prostaglandin E2 (PGE2)-induced angiogenesis. GRK2 small interfering RNA (siRNA) inhibits the increase in PGE2-induced HUVECs migration and tube formation. In vivo, PGE2 increases ECs sprouting from normal murine aortic segments and angiogenesis in mice, but not from GRK2-deficient ones, on Matrigel. Further research found that Lys220 and Ser685 of GRK2 play an important role in angiogenesis by regulating GRK2 translocation. Paeoniflorin-6'-O-benzene sulfonate (CP-25), as a novel ester derivative of paeoniflorin (pae), has therapeutic potential for the treatment of adjuvant arthritis (AA) and collagen-induced arthritis (CIA), but the underlying mechanism of CP-25 on angiogenesis has not been elucidated. In our study, CP-25 inhibits the migration and tube formation of HUVECs, and angiogenesis in mice by down-regulating GRK2 translocation activation without affecting GRK2 total expression. Taken together, the present results revealed that CP-25 down-regulates EP4/AC/cAMP/PKA-mediated GRK2 translocation, restoring the inhibition of GRK2 for ERK1/2, thereby inhibiting PGE2-stimulated angiogenesis.

VEGF Antagonism Attenuates Cerebral Ischemia/Reperfusion-Induced Injury via Inhibiting Endoplasmic Reticulum Stress-Mediated Apoptosis.

Endoplasmic reticulum (ER) stress-mediated apoptosis pathway is considered to play a vital role in mediating stroke and other cerebrovascular diseases. Previous studies have showed that vascular endothelial growth factor (VEGF) antagonism reduced cerebral ischemic-reperfusion (CI/R) damage, but whether attenuation of ER stress-induced apoptosis is contributing to its mechanisms remains elusive. Our study aimed to investigate the protective effect of VEGF antagonism on CI/R-induced injury. First, oxygen-glucose deprivation and re-oxygenation (OGD/R) BEND3 cell model was constructed to estimate small interfering RNA (siRNA)-VEGF on damage of endothelial cells. Next, in animal model, CI/R mice were induced by middle cerebral artery occlusion (MCAO) for 2 h followed by 24 h reperfusion to investigate cerebral tissue damage. For treatment group, mice received 100 µg/kg anti-VEGF antibodies at 30 min before MCAO, followed by 24 h reperfusion. Our findings demonstrated that pre-administration of siRNA-VEGF before OGD/R changed the biological characteristics of BEND3 cells, reversed the levels of X-box binding protein-1 (XBP-1) and glucose-regulated protein 78 (GRP78), showing siRNA-VEGF attenuated, at least in part, the oxidative damage in OGD/R cell by down-regulating ER stress. In mice experiment, pre-administration of anti-VEGF antibody reduced the brain infarct volume and edema extent and improved neurological scores outcome of CI/R injury mice. Pathological and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining results also confirmed this protective effect. The expressions of VEGF, CATT/EBP homologous protein (CHOP), inositol requiring enzyme 1α (IRE-1α), and cleaved-caspase12 and c-jun N-terminal kinase (JNK) phosphorylation were also prominently decreased. These results suggested that inhibition of endogenous VEGF attenuates CI/R-induced injury via inhibiting ER stress-mediated apoptosis.

Lentiviral Vectors and Adeno-Associated Virus Vectors: Useful Tools for Gene Transfer in Pain Research.

Pain, especially chronic pain, has always been a heated point in both basic and clinical researches since it puts heavy burdens on both individuals and the whole society. A better understanding of the role of biological molecules and various ionic channels involved in pain can shed light on the mechanism under pain and advocate the development of pain management. Using viral vectors to transfer specific genes at targeted sites is a promising method for both research and clinical applications. Lentiviral vectors and adeno-associated virus (AAV) vectors which allow stable and long-term expression of transgene in non-dividing cells are widely applied in pain research. In this review, we thoroughly outline the structure, category, advantages and disadvantages and the delivery methods of lentiviral and AAV vectors. The methods through which lentiviral and AAV vectors are delivered to targeted sites are closely related with the sites, level and period of transgene expression. Focus is placed on the various delivery methods applied to deliver vectors to spinal cord and dorsal root ganglion both of which play important roles in primary nociception. Our goal is to provide insight into the features of these two viral vectors and which administration approach can be chosen for different pain researches. Anat Rec, 301:825-836, 2018. © 2017 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

p53 and mitochondrial dysfunction: novel insight of neurodegenerative diseases.

Mitochondria are organelles responsible for vital cell functions. p53 is a transcription factor that regulates the DNA stability and cell growth normality. Recent studies revealed that p53 can influence mitochondrial function changing from normal condition to abnormal condition under different stress levels. In normal state, p53 can maintain mitochondrial respiration through transactivation of SCO2. When stress stimuli presents, SCO2 overexpresses and leads to ROS generation. ROS promotes p53 inducing MALM (Mieap-induced accumulation of lysosome-like organelles within mitochondria) to repair dysfunctional mitochondria and MIV (Mieap-induced vacuole) to accomplish damaged mitochondria degradation. If stress or damage is irreversible, p53 will translocate to mitochondria, leading into apoptosis or necrosis. Neurodegenerative diseases including Parkinson's disease, Huntington's disease and Alzheimer's disease are still lack of clear explanations of mechanisms, but more studies have revealed the functional relationship between mitochondria and p53 towards the pathological development of these diseases. In this review, we discuss that p53 plays the vital role in the function of mitochondria in the aspect of pathological change metabolism. We also analyze these diseases with novel targeted treating molecules which are related to p53 and mitochondria, hoping to present novel therapies in future clinic.

Hepatic actinomycosis: report of one case and analysis of 32 previously reported cases.

Hepatic actinomycosis is rare, with few published cases. There are no characteristic clinical manifestations, and computed tomography (CT) shows mainly low-density images, making clinical diagnosis difficult, and leading to frequent misdiagnosis as primary liver cancer, metastatic liver cancer or liver abscess. Diagnosis normally requires examination of both the aetiology and pathology. This article reports one male patient aged 55 who was hospitalized because of repeated upper abdominal pain for more than 2 mo. He exhibited no chills, fever or yellow staining of the skin and sclera, and examination revealed no positive signs. The routine blood results were: haemoglobin 110 g/L, normal numbers of leukocytes and neutral leukocytes, serum albumin 32 g/L, negative serum hepatitis B markers and hepatitis C antibodies, normal tumour markers (alpha-fetoprotein and carcinoembryonic antigen). An abdominal CT scan revealed an 11.2 cm × 5.8 cm × 7.4 cm mass with an unclear edge in the left liver lobe. The patient was diagnosed as having primary liver cancer, and left lobe resection was performed. The postoperative pathological examination found multifocal actinomycetes in the hepatic parenchyma, which was accompanied by chronic suppurative inflammation. A focal abscess had formed, and large doses of sodium penicillin were administered postoperatively as anti-infective therapy. This article also reviews 32 cases reported in the English literature, with the aim of determining the clinical features and treatment characteristics of this disease, and providing a reference for its diagnosis and treatment.