Owing to its tumor tropism and prolonged transgene expression, mesenchymal stem cell (MSC) has been considered as an ideal delivery vehicle for cancer gene therapies or therapeutic vaccines. In this ...study, we demonstrated that intratumoral (i.t.) injection of MSCs expressing modified interleukin-12 (MSCs/IL-12M) exhibited stronger tumor-specific T-cell responses and antitumor effects as well as more sustained expressions of IL-12 and interferon (IFN)-γ in both sera and tumor sites than did IL-12M-expressing adenovirus (rAd/IL-12M) in mice bearing both solid and metastatic tumors. Subcutaneous (s.c.) injection of MSCs/IL-12M at contralateral site of tumor exhibited similar levels of serum IL-12 and IFN-γ as i.t. injection, but much weaker antitumor effects in both B16F10 melanoma and TC-1 cervical cancer models than i.t. injection. Although intravenous (i.v.) injection elicited earlier peak serum levels of cytokines, it induced weaker tumor-specific T-cell responses and antitumor effects than i.t. injection, indicating that serum cytokine levels are not surrogate indicators of antitumor effects. Taken together, these results indicated that MSC is more efficient than adenovirus as a cytokine gene delivery vehicle and that i.t. injection of MSCs/IL-12M is the best approach to induce strong tumor-specific T-cell responses that correlate with anti-metastatic effects as well as inhibition of solid tumor growth, although MSCs themselves have an ability to migrate into the tumor site. In addition, MSCs/IL-12M embedded in Matrigel (MSCs/IL-12M/Matrigel) exhibited significant antitumor effects even in immunodeficient mice such as SCID and BNX mice lacking T, B and natural killer (NK) cells, but not in IFN-γ knockout mice. Our findings provide an optimal approach for designing an efficient clinical protocol of MSC-based cytokine gene therapy to induce strong tumor-specific T-cell responses and therapeutic anticancer efficacy.
Summary
Background
Autophagy and genetic predisposition have been suggested to potentially play roles in the development of asthma. However, little is known about the role of autophagy in the ...pathogenesis of severe asthma.
Objective
We compared autophagy in the sputum granulocytes, peripheral blood cells (PBCs) and peripheral blood eosinophils (PBEs) between patients with severe asthma and those with non‐severe asthma and investigated the functional effects of autophagy.
Methods
We enrolled 36 patients with severe asthma, 14 with non‐severe asthma and 23 normal healthy controls in this study. Sputum granulocytes, PBCs and PBEs were isolated from each subject. Autophagy was evaluated based on the expression of microtubule‐associated protein light chain 3 (LC3) by Western blot, confocal microscopy, transmission electron microscopy and flow cytometry. IL‐8 levels were measured by ELISA. To induce autophagy, HL‐60 cells, human primary small airway epithelial cells (SAECs) and A549 cells were treated with IL‐5, IL‐1β and TNF‐α. To inhibit autophagy, PI3K inhibitors (LY29400 and 3‐methyladenine 3‐MA) and hydroxychloroquine (HCQ) were used. Knockdown of ATG5 and Beclin‐1 was performed in A549 cells, and the therapeutic effects of dexamethasone were evaluated.
Results
Higher autophagy levels were noted in sputum granulocytes, PBCs and PBEs from patients with severe asthma than from patients with non‐severe asthma and healthy controls (P < 0.05 for all). IL‐5 increased autophagy levels in both PBCs and PBEs (P < 0.05). 3‐MA attenuated the increased expression of LC3‐II and eosinophil cationic protein in HL‐60 cells induced by IL‐5 (P = 0.034 for both). Dexamethasone did not affect autophagy levels in PBEs. IL‐1β increased LC3‐II expression and IL‐8 production (P < 0.01) in SAECs, and this was attenuated by LY294002, 3‐MA, HCQ and knockdown of ATG5 and Beclin‐1 (in A549 cells) (P < 0.01).
Conclusions and Clinical Relevance
Autophagy could play a role in the pathogenesis of severe asthma. Autophagy modulation may be a novel therapeutic target for conventional therapy‐resistant severe asthma.
Despite ionizing radiation (IR) is being widely used as a standard treatment for lung cancer, many evidences suggest that IR paradoxically promotes cancer malignancy. However, its molecular ...mechanisms underlying radiation-induced cancer progression remain obscure. Here, we report that exposure to fractionated radiation (2 Gy per day for 3 days) induces the secretion of granulocyte-colony-stimulating factor (G-CSF) that has been commonly used in cancer therapies to ameliorate neutropenia. Intriguingly, radiation-induced G-CSF promoted the migratory and invasive properties by triggering the epithelial-mesenchymal cell transition (EMT) in non-small-cell lung cancer cells (NSCLCs). By irradiation, G-CSF was upregulated transcriptionally by β-catenin/TCF4 complex that binds to the promoter region of G-CSF as a transcription factor. Importantly, irradiation increased the stability of β-catenin through the activation of PI3K/AKT (phosphatidylinositol 3-kinase/AKT), thereby upregulating the expression of G-CSF. Radiation-induced G-CSF is recognized by G-CSFR and transduced its intracellular signaling JAK/STAT3 (Janus kinase/signal transducers and activators of transcription), thereby triggering EMT program in NSCLCs. Taken together, our findings suggest that the application of G-CSF in cancer therapies to ameliorate neutropenia should be reconsidered owing to its effect on cancer progression, and G-CSF could be a novel therapeutic target to mitigate the harmful effect of radiotherapy for the treatment of NSCLC.
We aimed to evaluate the long-term clinical outcomes and prognostic factors of symptomatic intracranial unruptured vertebrobasilar artery dissection (siu-VBD).
A total of 191 patients (M:F = 127:64; ...median age, 46 years) with siu-VBD were treated between January 2001 and December 2008. Presentations, treatments, outcomes, and prognostic factors were retrospectively analyzed.
Clinical manifestations were ischemic symptoms with headache (n = 97) or without headache (n = 13) and headache without ischemic symptoms (n = 81). Forty-six patients (24.1%) underwent endovascular treatment. The remaining 145 patients (75.9%) were medically treated with anticoagulants (n = 49), antiplatelets (n = 48), or analgesics alone (n = 48). Clinical follow-up data were available in 178 patients (102 ischemic and 76 nonischemic) at 15 to 102 months (mean, 46 months). None of the siu-VBD hemorrhaged. All 76 patients without ischemic presentation had favorable outcomes (modified Rankin Scale, 0-1). Of the 102 patients with ischemic presentation, outcomes were favorable in 92 and unfavorable in 10 patients. Four patients died; 3 died of causes unrelated to VBD, and one died as a result of basilar artery (BA) dissection. Old age (odds ratio OR 1.099; 95% confidence interval CI 1.103-1.204; p = 0.042) and BA involvement (OR 11.886; 95% CI 1.416-99.794; p = 0.023) were independent predictors of unfavorable outcomes in siu-VBD with ischemic presentation.
Clinical outcomes for siu-VBD were favorable in all patients without ischemic symptoms and in most patients with ischemic presentation. None of the siu-VBD caused subarachnoid hemorrhage. Old age and BA involvement were independent predictors of unfavorable outcome in siu-VBD with ischemic presentation.
Abstract
Methylglyoxal is a reactive dicarbonyl compound produced by glycolytic processing and identified as a precursor of advanced glycation end products. The elevated methylglyoxal levels in ...patients with diabetes are believed to contribute to diabetic complications, including bone defects. The objective of this study was to evaluate the effect of methylglyoxal on the function of osteoblastic MC3T3-E1 cells. The data indicated that methylglyoxal decreased osteoblast differentiation and induced osteoblast cytotoxicity. Pretreatment of MC3T3-E1 cells with aminoguanidine (a carbonyl scavenger), Trolox (an antioxidant), and cyclosporin A (a blocker of the mitochondrial permeability transition pore) prevented methylglyoxal-induced cytotoxicity in MC3T3-E1 cells. However, BAPTA/AM (an intracellular Ca2+ chelator) and dantrolene (an inhibitor of endoplasmic reticulum Ca2+ release) did not reverse the cytotoxic effect of methylglyoxal. Methylglyoxal increased the formation of intracellular reactive oxygen species, mitochondrial superoxide, and cardiolipin peroxidation in osteoblastic MC3T3-E1 cells. Methylglyoxal also decreased the mitochondrial membrane potential and intracellular ATP and nitric oxide levels, suggesting that carbonyl stress-induced loss of mitochondrial integrity contributes to the cytotoxicity of methylglyoxal. Furthermore, the results demonstrated that methylglyoxal induced protein adduct formation, inactivation of glyoxalase I, and activation of glyoxalase II. Aminoguanidine reversed all aforementioned effects of methylglyoxal. Taken together, these data support the notion that high methylglyoxal concentrations have detrimental effects on osteoblasts through a mechanism involving oxidative stress and mitochondrial dysfunction.
Summary Objective Cartilage regenerative procedures using the cell-based tissue engineering approach involving mesenchymal stem cells (MSCs) have been receiving increased interest because of their ...potential for altering the progression of osteoarthritis (OA) by repairing cartilage lesions. The aim of this study was to investigate the clinical and magnetic resonance imaging (MRI) outcomes of MSC implantation in OA knees and to determine the association between clinical and MRI outcomes. Design Twenty patients (24 knees) who underwent arthroscopic MSC implantation for cartilage lesions in their OA knees were evaluated at 2 years after surgery. Clinical outcomes were evaluated according to the International Knee Documentation Committee (IKDC) score and the Tegner activity scale, and cartilage repair was assessed according to the MRI Osteoarthritis Knee Score (MOAKS) and Magnetic Resonance Observation of Cartilage Repair Tissue (MOCART) score. Results The clinical outcomes significantly improved ( P < 0.001 for both). The cartilage lesion grades (as described in MOAKS grades for size of cartilage-loss area and percentage of full-thickness cartilage loss) at follow-up MRI were significantly better than the preoperative values ( P < 0.001 for both). The clinical outcomes at final follow-up were significantly correlated with the MOAKS and MOCART score at follow-up MRI ( P < 0.05 for all). Conclusions Considering the encouraging clinical and MRI outcomes obtained and the significant correlations noted between the clinical and MRI outcomes, MSC implantation seems to be useful for repairing cartilage lesions in OA knees. However, a larger sample size and long-term studies are needed to confirm our findings.
Claudins (CLDNs) are a family of integral membrane proteins central to the formation of tight junctions, structures that are involved in paracellular transport and cellular growth and ...differentiation, and are critical for the maintenance of cellular polarity. Recent studies have provided evidence that CLDNs are aberrantly expressed in diverse types of human cancers, including hepatocellular carcinomas (HCCs). However, little is known about how CLDN expression is involved in cancer progression. In this study, we show that CLDN1 has a causal role in the epithelial-mesenchymal transition (EMT) in human liver cells, and that the c-Abl-Ras-Raf-1-ERK1/2 signaling axis is critical for the induction of malignant progression by CLDN1. Overexpression of CLDN1 induced expression of the EMT-regulating transcription factors Slug and Zeb1, and thereby led to repression of E-cadherin, β-catenin expression, enhanced expression of N-cadherin and Vimentin, a loss of cell adhesion, and increased cell motility in normal liver cells and HCC cells. In line with these findings, inhibition of either c-Abl or ERK clearly attenuated CLDN1-induced EMT, as evidenced by a reversal of N-cadherin and E-cadherin expression patterns, and restored normal motility. Collectively, these results indicate that CLDN1 is necessary for the induction of EMT in human liver cells, and that activation of the c-Abl-Ras-Raf-1-ERK1/2 signaling pathway is required for CLDN1-induced acquisition of the malignant phenotype. The present observations suggest that CLDN1 could be exploited as a biomarker for liver cancer metastasis and might provide a pivotal point for therapeutic intervention in HCC.
Highlights ► We examined the mechanism of mitochondrial swelling and axonal degeneration. ► Energy failure occurred prior to mitochondrial swelling and axonal degeneration. ► Energy repletion ...prevented mitochondrial swelling and microtubule depolymerization. ► Perturbing microtubule dynamics enhanced axonal degeneration and mitochondrial swelling. ► Calcium was not related to mitochondrial swelling and microtubule depolymerization.
In stage I/II natural killer (NK)/T-cell lymphoma, concurrent chemoradiotherapy (CCRT) had previously been shown to result in superior outcome compared with anthracycline-containing regimens, which ...have since been considered ineffective. The role of CCRT in comparison with approaches employing nonanthracycline-containing chemotherapy (CT) and sequential radiotherapy (RT) in such patients remains to be defined.
Three hundred and three untreated patients (207 men, 96 women; median age: 51, 18–86 years) with stage I/II NK/T-cell lymphoma who had received nonanthracycline-containing regimens were collected from an international consortium and retrospectively analyzed. Treatment included single modality (CT and RT), sequential modalities (CT + RT; RT + CT) and concurrent modalities (CCRT; CCRT + CT). The impact of clinicopathologic parameters and types of treatment on complete response (CR) rate, progression-free-survival (PFS) and overall-survival (OS) was evaluated.
For CR, stage (P = 0.027), prognostic index for NK/T-cell lymphoma (PINK) (P = 0.026) and types of initial treatment (P = 0.011) were significant prognostic factors on multivariate analysis. On Cox regression analysis, ECOG performance score (P = 0.021) and PINK-EBV DNA (PINK-E) (P = 0.002) significantly impacted on PFS; whereas ECOG performance score (P = 0.008) and stage (P < 0.001) significantly impacted on OS. For comparing CCRT ± CT and sequential CT + RT, CCRT ± CT patients (n = 190) were similar to sequential CT + RT patients (n = 54) in all evaluated clinicopathologic parameters except two significantly superior features (higher proportion of undetectable circulating EBV DNA on diagnosis and lower PINK-E scores). Despite more favorable pre-treatment characteristics, CCRT ± CT patients had CR rate, PFS and OS comparable with sequential CT + RT patients on multivariate and Cox regression analyses.
In stage I/II NK/T-cell lymphomas, when effective chemotherapeutic regimens were used, CCRT and sequential CT + RT gave similar outcome.
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