ABSTRACT
Reprogramming of cancer cells into induced pluripotent stem cells (iPSCs) is a compelling idea for inhibiting oncogenesis, especially through modulation of homeobox proteins in this reprogramming process. We examined the role of various long noncoding RNAs (lncRNAs)-homeobox protein HOXA13 axis on the switching of the oncogenic function of bone morphogenetic protein 7 (BMP7), which is significantly lost in the gastric cancer cell derived iPS-like cells (iPSLCs). BMP7 promoter activation occurred through the corecruitment of HOXA13, mixed-lineage leukemia 1 lysine N-methyltransferase, WD repeat-containing protein 5, and lncRNA HoxA transcript at the distal tip (HOTTIP) to commit the epigenetic changes to the trimethylation of lysine 4 on histone H3 in cancer cells. By contrast, HOXA13 inhibited BMP7 expression in iPSLCs via the corecruitment of HOXA13, enhancer of zeste homolog 2, Jumonji and AT rich interactive domain 2, and lncRNA HoxA transcript antisense RNA (HOTAIR) to various cis-element of the BMP7 promoter. Knockdown experiments demonstrated that HOTTIP contributed positively, but HOTAIR regulated negatively to HOXA13-mediated BMP7 expression in cancer cells and iPSLCs, respectively. These findings indicate that the recruitment of HOXA13-HOTTIP and HOXA13-HOTAIR to different sites in the BMP7 promoter is crucial for the oncogenic fate of human gastric cells. Reprogramming with octamer-binding protein 4 and Jun dimerization protein 2 can inhibit tumorigenesis by switching off BMP7. Stem Cells 2017;35:2115-2128.
Subject(s)
Cellular Reprogramming Techniques/methods , Homeodomain Proteins/genetics , RNA, Long Noncoding/genetics , Stomach Neoplasms/genetics , Bone Morphogenetic Protein 7/genetics , Bone Morphogenetic Protein 7/metabolism , Cell Line, Tumor , Cell Proliferation , Homeodomain Proteins/metabolism , Humans , Promoter Regions, Genetic , RNA, Long Noncoding/metabolism , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathologyABSTRACT
The network of stemness genes and oncogenes in human patient-specific reprogrammed cancer stem cells (CSCs) remains elusive, especially in liver cancer. HepG2-derived induced pluripotent stem cell-like cells (HepG2-iPS-like cells) were generated by introducing Yamanaka factors and the knockdown vector shTP53. They exhibited features of stemness and a higher tumorigenesis after xenograft transplantation compared with HepG2 cells. The cancerous mass of severe combined immunodeficiency (SCID) mice derived from one colony was dissected and cultured to establish reprogrammed HepG2-derived CSC-like cells (designated rG2-DC-1C). A single colony exhibited 42% occurrence of tumors with higher proliferation capacities. rG2-DC-1C showed continuous expression of the OCT4 stemness gene and of representative tumor markers, potentiated chemoresistance characteristics, and invasion activities. The sphere-colony formation ability and the invasion activity of rG2-DC-1C were also higher than those of HepG2 cells. Moreover, the expression of the OCT4 gene and the c-JUN oncogene, but not of c-MYC, was significantly elevated in rG2-DC-1C, whereas no c-JUN expression was observed in HepG2 cells. The positive-feedback regulation via OCT4-mediated transactivation of the c-JUN promoter and the c-JUN-mediated transactivation of the OCT4 promoter were crucial for promoting cancer development and maintaining cancer stemness in rG2-DC-1C. Increased expression of OCT4 and c-JUN was detected in the early stage of human liver cancer. Therefore, the positive feedback regulation of OCT4 and c-JUN, resulting in the continuous expression of oncogenes such as c-JUN, seems to play a critical role in the determination of the cell fate decision from iPS cells to CSCs in liver cancer. Stem Cells 2016;34:2613-2624.
Subject(s)
Feedback, Physiological , Gene Expression Regulation, Neoplastic , JNK Mitogen-Activated Protein Kinases/genetics , Liver Neoplasms/genetics , Neoplastic Stem Cells/metabolism , Octamer Transcription Factor-3/genetics , Aged , Animals , Antineoplastic Agents/pharmacology , Cell Differentiation , Cellular Reprogramming , Cisplatin/pharmacology , Doxorubicin/pharmacology , Drug Resistance, Neoplasm/genetics , Female , Fluorouracil/pharmacology , Hep G2 Cells , Humans , Induced Pluripotent Stem Cells/drug effects , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/pathology , JNK Mitogen-Activated Protein Kinases/metabolism , Liver Neoplasms/drug therapy , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Male , Mice , Mice, SCID , Middle Aged , Neoplastic Stem Cells/drug effects , Neoplastic Stem Cells/pathology , Octamer Transcription Factor-3/metabolism , Signal Transduction , Spheroids, Cellular/drug effects , Spheroids, Cellular/metabolism , Spheroids, Cellular/pathology , Transcriptional Activation , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
A fragment-based method was developed to investigate the binding conformations of peptide ligands. This method efficiently avoids the high degree of freedom (DOF) of peptide dockings by dividing a peptide into two half fragments. The fragments are separately docked on receptors and the results are used to rebuild a profile of massive possible docking conformations of the whole peptide. Through rapid scoring for filtering, the remaining peptide docking conformations are rigorously optimized by molecular dynamics (MD) and scored by molecular mechanics/generalized born surface area (MM/GBSA) method to predict the near-native binding conformations. This method has been tested on 17 cases of long peptide-protein interaction with known crystal structures, and also on 7 unbound protein receptors for which both the bound and unbound conformations are known. The resultant binding predictions fit very closely to the crystal structures.
Subject(s)
Molecular Docking Simulation , Peptides/metabolism , Proteins/metabolism , Algorithms , Amino Acid Sequence , Binding Sites , Peptides/chemistry , Protein Binding , Protein Structure, Secondary , Protein Structure, Tertiary , Proteins/chemistry , ThermodynamicsABSTRACT
During disease progression to AIDS, HIV-1 infected individuals become increasingly immunosuppressed and susceptible to opportunistic infections. It has also been demonstrated that multiple subsets of dendritic cells (DC), including DC-SIGN⺠cells, become significantly depleted in the blood and lymphoid tissues of AIDS patients, which may contribute to the failure in initiating effective host immune responses. The mechanism for DC depletion, however, is unclear. It is also known that vast quantities of viral envelope protein gp120 are shed from maturing HIV-1 virions and form circulating immune complexes in the serum of HIV-1-infected individuals, but the pathological role of gp120 in HIV-1 pathogenesis remains elusive. Here we describe a previously unrecognized mechanism of DC death in chronic HIV-1 infection, in which ligation of DC-SIGN by gp120 sensitizes DC to undergo accelerated apoptosis in response to a variety of activation stimuli. The cultured monocyte-derived DC and also freshly-isolated DC-SIGN⺠blood DC that were exposed to either cross-linked recombinant gp120 or immune-complex gp120 in HIV⺠serum underwent considerable apoptosis after CD40 ligation or exposure to bacterial lipopolysaccharide (LPS) or pro-inflammatory cytokines such as TNFα and IL-1ß. Furthermore, circulating DC-SIGN⺠DC that were isolated directly from HIV-1⺠individuals had actually been pre-sensitized by serum gp120 for activation-induced exorbitant apoptosis. In all cases the DC apoptosis was substantially inhibited by DC-SIGN blockade. Finally, we showed that accelerated DC apoptosis was a direct consequence of excessive activation of the pro-apoptotic molecule ASK-1 and transfection of siRNA against ASK-1 significantly prevented the activation-induced excessive DC death. Our study discloses a previously unknown mechanism of immune modulation by envelope protein gp120, provides new insights into HIV immunopathogenesis, and suggests potential therapeutic approaches to prevent DC depletion in chronic HIV infection.
Subject(s)
Apoptosis/physiology , Cell Adhesion Molecules/metabolism , Dendritic Cells/metabolism , HIV Envelope Protein gp120/metabolism , Lectins, C-Type/metabolism , MAP Kinase Kinase Kinase 5/metabolism , Receptors, Cell Surface/metabolism , Apoptosis/drug effects , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD40 Antigens/immunology , Cell Adhesion Molecules/immunology , Cells, Cultured , Dendritic Cells/immunology , Dendritic Cells/pathology , Gene Silencing , HIV Envelope Protein gp120/immunology , HIV Infections/blood , HIV Infections/immunology , Host-Pathogen Interactions , Humans , Lectins, C-Type/immunology , Lipopolysaccharides/pharmacology , MAP Kinase Kinase Kinase 5/immunology , Protein Binding , RNA, Small Interfering/genetics , Receptors, Cell Surface/immunology , TransfectionABSTRACT
Severe acute respiratory syndrome (SARS) is caused by infection of a previously undescribed coronavirus (CoV). L-SIGN, encoded by CLEC4M (also known as CD209L), is a SARS-CoV binding receptor that has polymorphism in its extracellular neck region encoded by the tandem repeat domain in exon 4. Our genetic risk association study shows that individuals homozygous for CLEC4M tandem repeats are less susceptible to SARS infection. L-SIGN is expressed in both non-SARS and SARS-CoV-infected lung. Compared with cells heterozygous for L-SIGN, cells homozygous for L-SIGN show higher binding capacity for SARS-CoV, higher proteasome-dependent viral degradation and a lower capacity for trans infection. Thus, homozygosity for L-SIGN plays a protective role during SARS infection.
Subject(s)
Cell Adhesion Molecules/genetics , Lectins, C-Type/genetics , Receptors, Cell Surface/genetics , Severe Acute Respiratory Syndrome/genetics , Severe acute respiratory syndrome-related coronavirus/pathogenicity , Animals , CHO Cells/virology , Cell Adhesion Molecules/metabolism , Chlorocebus aethiops , Cohort Studies , Cricetinae , Cricetulus , Genetic Predisposition to Disease , Homozygote , Hong Kong/epidemiology , Humans , Intestine, Small/physiology , Lectins, C-Type/metabolism , Lung/physiology , Lung/virology , Molecular Sequence Data , Proteasome Endopeptidase Complex/metabolism , Receptors, Cell Surface/metabolism , Severe acute respiratory syndrome-related coronavirus/metabolism , Severe Acute Respiratory Syndrome/epidemiology , Tandem Repeat Sequences , Vero Cells/virologyABSTRACT
Severe dengue virus (DV) infections can cause the life-threatening condition dengue hemorrhagic fever, which is characterized by a severe plasma leak, thrombocytopenia, hemorrhage, and, in severe cases, circulatory collapse and death. There is now much evidence that pre-existing immunity to DV can enhance disease when an individual becomes infected on a second or sequential occasion. It has been shown that in contrast to infected dendritic cells (DC), noninfected bystander DC underwent maturation in dengue infection. In this study, we show that TNF-alpha and type I IFN contribute to the maturation of bystander DC, whereas the inhibition of DV-infected DC maturation can be overcome by activated T cells. Furthermore, IFN-gamma-inducible chemokines, CXCL9, 10, and 11 produced by infected DC are greatly amplified in the presence of DV-specific T cells. The chemokine secretion is also enhanced in coculture of HUVEC with either DV-infected DC or activated T cells. Finally, we found a close correlation between the serum level of these three chemokines and disease severity.
Subject(s)
Cytokines/physiology , Dendritic Cells/immunology , Dendritic Cells/virology , Dengue Virus/immunology , Dengue/immunology , Dengue/virology , T-Lymphocytes/immunology , T-Lymphocytes/virology , Bystander Effect/immunology , Cell Communication/immunology , Cell Differentiation/immunology , Cells, Cultured , Coculture Techniques , Cytokines/biosynthesis , Cytotoxicity, Immunologic , Dendritic Cells/cytology , Dendritic Cells/metabolism , Dengue/metabolism , Epitopes, T-Lymphocyte/immunology , Epitopes, T-Lymphocyte/metabolism , Humans , Lymphocyte Activation/immunology , T-Lymphocytes/cytology , T-Lymphocytes/metabolism , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/virologyABSTRACT
Effective vaccines should confer long-term protection against future outbreaks of severe acute respiratory syndrome (SARS) caused by a novel zoonotic coronavirus (SARS-CoV) with unknown animal reservoirs. We conducted a cohort study examining multiple parameters of immune responses to SARS-CoV infection, aiming to identify the immune correlates of protection. We used a matrix of overlapping peptides spanning whole SARS-CoV proteome to determine T cell responses from 128 SARS convalescent samples by ex vivo IFN-gamma ELISPOT assays. Approximately 50% of convalescent SARS patients were positive for T cell responses, and 90% possessed strongly neutralizing Abs. Fifty-five novel T cell epitopes were identified, with spike protein dominating total T cell responses. CD8(+) T cell responses were more frequent and of a greater magnitude than CD4(+) T cell responses (p < 0.001). Polychromatic cytometry analysis indicated that the virus-specific T cells from the severe group tended to be a central memory phenotype (CD27(+)/CD45RO(+)) with a significantly higher frequency of polyfunctional CD4(+) T cells producing IFN-gamma, TNF-alpha, and IL-2, and CD8(+) T cells producing IFN-gamma, TNF-alpha, and CD107a (degranulation), as compared with the mild-moderate group. Strong T cell responses correlated significantly (p < 0.05) with higher neutralizing Ab. The serum cytokine profile during acute infection indicated a significant elevation of innate immune responses. Increased Th2 cytokines were observed in patients with fatal infection. Our study provides a roadmap for the immunogenicity of SARS-CoV and types of immune responses that may be responsible for the virus clearance, and should serve as a benchmark for SARS-CoV vaccine design and evaluation.
Subject(s)
Antibodies, Viral/immunology , CD8-Positive T-Lymphocytes/immunology , Immunologic Memory , Severe Acute Respiratory Syndrome/immunology , Severe acute respiratory syndrome-related coronavirus/immunology , Th2 Cells/immunology , Adult , Cohort Studies , Cytokines/immunology , Female , Humans , Leukocyte Common Antigens/immunology , Lysosomal-Associated Membrane Protein 1/immunology , Male , Middle Aged , Proteome/immunology , Severe Acute Respiratory Syndrome/mortality , Tumor Necrosis Factor Receptor Superfamily, Member 7/immunology , Viral Vaccines/immunologyABSTRACT
Nasopharyngeal carcinoma (NPC) is a common neoplasm in Southeastern Asia, and cisplatin-containing regimens for combinational chemotherapy are widely used for treating locally recurrent or metastatic diseases. However, resistance to cisplatin is not infrequently seen and its associated side effects may be life-threatening. In this report, another metallo-pharmaceutical agent gold(III) porphyrin complex [Au(TPP)]Cl was investigated in comparison to cisplatin for its in vitro and in vivo anticancer effects. Through induction of the intrinsic apoptosis pathway, [Au(TPP)]Cl exhibited 100-fold higher potency than cisplatin in killing NPC cells, including cisplatin-sensitive and cisplatin-resistant variants, and also an variant harboring the Epstein-Barr virus. In addition, a safety concentration window was demonstrated, allowing [Au(TPP)]Cl to kill tumors with minimal cytotoxicity to noncancerous cells. More importantly, weekly intraperitoneal injection of 3 mg/kg [Au(TPP)]Cl was more effective than the same dose of cisplatin in inducing tumor apoptosis in vivo and remarkably inhibited tumor growth in animals without any noticeable side effect. [Au(TPP)]Cl therefore is a promising chemotherapeutic agent that deserves further development as a novel drug for the treatment of advanced NPC, in particular, for cases with cisplatin-resistance.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Carcinoma/drug therapy , Cisplatin/therapeutic use , Gold/administration & dosage , Nasopharyngeal Neoplasms/drug therapy , Porphyrins/administration & dosage , Animals , Cell Differentiation , Cell Line, Tumor , Female , Humans , In Vitro Techniques , Mice , Mice, Inbred BALB C , Models, Chemical , Treatment OutcomeABSTRACT
Nasopharyngeal carcinoma is a neoplasm with a high incidence in Southeast Asia, and it is strongly associated with Epstein-Barr virus (EBV) activation involving the expression of a weakly immunogenic protein, namely, latent membrane protein (LMP)-2. Previous immunological studies already identified the human leukocyte antigen (HLA)-A11 restricted peptide epitope (SSCSSCPLSK) in the LMP-2 antigen. In this work, we prepared gold nanoparticle (AuNP)-peptide conjugate 1 by treating the nanoparticles with the N-cysteinated LMP-2 epitope. The AuNP-peptide conjugates have been characterized by TEM (15-24 nm in diameter) and UV-vis spectroscopy (surface plasmon resonance absorption band at lambda(max) = 520 nm). In the presence of a CALNN capping peptide, the AuNP-peptide conjugates are stable in solution without aggregation at room temperature for at least 48 h. By ELIspot studies, AuNP-peptide conjugate 1 was found to elicit a significantly stronger INF-gamma response [number of spot forming cells (SPC) = 727 +/- 198] from peripheral blood mononuclear cells of healthy HLA-A11 donors when compared to that induced by the unconjugated LMP-2 peptides (SFC = 73 +/- 28). Further studies showed that dendritic cells treated with conjugate 1 can effect CD8+ T-cell activation leading to epitope-specific cytotoxic T lymphocyte killing responses in vitro.
Subject(s)
Antigens, Viral/immunology , Herpesvirus 4, Human/immunology , Metal Nanoparticles/chemistry , T-Lymphocytes, Cytotoxic/immunology , Viral Matrix Proteins/immunology , CD8-Positive T-Lymphocytes/immunology , Cytotoxicity, Immunologic , Gold , Humans , Lymphocyte Activation/immunology , Viral Matrix Proteins/chemistryABSTRACT
Nasopharyngeal carcinoma (NPC), a common neoplasm in Southeast Asia, is EBV-positive and expresses a limited number of antigens, including latent membrane protein 2. In this study, autologous monocyte-derived dendritic cells were cultured from patients with advanced NPC, matured with cytokine, pulsed with HLA-A1101-, A2402-, or B40011-restricted epitope peptides from EBV latent membrane protein 2 and injected into inguinal lymph nodes. Sixteen patients with local recurrence or distant metastasis after conventional therapies received four injections at weekly intervals. Epitope-specific CD8+ T-cell responses were elicited or boosted in 9 patients receiving HLA-A1101- or A2402-restricted peptides, with stronger responses seen to the A1101 peptide. Furthermore, epitope-specific cytotoxicity was detectable in peripheral blood T cells harvested at 3-months after vaccination from A1101-responsive patients, and in 2 patients, this coincided with partial tumor reduction. Approaches leading to stronger and more sustained EBV-specific T-cell responses, therefore, may have therapeutic potential in the context of NPC.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Dendritic Cells/immunology , Herpesvirus 4, Human , Immunotherapy, Adoptive/methods , Nasopharyngeal Neoplasms/immunology , Nasopharyngeal Neoplasms/therapy , Viral Matrix Proteins/immunology , Adult , CD8-Positive T-Lymphocytes/metabolism , Cytotoxicity, Immunologic , Female , HLA-A1 Antigen/immunology , Humans , Interferon-gamma/biosynthesis , Lymphocyte Activation/immunology , Male , Middle Aged , Nasopharyngeal Neoplasms/virologyABSTRACT
BACKGROUND: Chronic low back pain affects daily activities at home and workplaces and causes a huge economic burden. Current therapeutic options are very limited and the effects of available pharmacological agents are less than satisfactory. While NSAIDs might be effective for the short term and opioids might help with urgent pain relief and improving the life quality, their long-term use is associated with significant side effects and drug misuse or abuse. To seek alternative pharmacological agents for effective treatment, we examined the therapeutic potential of the extracts of Vaccinia variola-inoculated rabbit skin (Analgecine, abbreviated as AGC) in patients with chronic low back pain due to degenerative vertebral disorders. METHODS: In this randomized multi-center double-blind placebo-controlled phase 3 clinical trial (Chinese Clinical Trial Registry number 2009L01498), we enrolled patients (aged 26-70 years) with chronic low back pain for at least 3 months due to degenerative spinal (vertebral) disorders from 7 medical centers in China, and randomly allocated 459 participants to receive oral AGC or placebo for 28 days to study the efficacy and safety of AGC. Randomization was performed according to a centralized randomization schedule, which was blocked by study sites and generated by an unmasked statistician independent of study conduct and data analysis. Both participants and staff at each study site were masked to treatment assignment. The primary efficacy endpoint was the change of the mean pain intensity, based on an 11-point numerical rating scale, between the baseline and the last week of treatment, with the primary efficacy analysis of intention to treat. The ratio between exposed and unexposed groups was designed to be 3:1 in order to increase the likelihood of demonstrating the AGC effect upon repeated measures. RESULTS: 347 patients were assigned to receive AGC (4 units/tablet; 2 tablets twice a day) and 112 patients were to take placebo. Among them, 324 patients taking AGC and 112 receiving placebo completed the assessment. Patients receiving AGC reported significant pain relief at the end of week 2 and 3 compared to those taking placebo, with mean reduction of the pain scores as 1.7 vs. 0.9 at week 2 (p < 0.0001) and 2.8 vs. 1.2 at week 3 (p < 0.0001). A total of 47 AGC-treated patients reported 85 treatment emergent adverse events while 16 patients taking placebo reported 26 events, but no serious side effects were found to be related to AGC treatment. CONCLUSION: Analgecine (AGC, 8 units twice daily) effectively alleviates chronic low back pain due to degenerative vertebral disorders when compared to placebo and is well tolerated by tested individuals, and can be considered as a first-line treatment for chronic low pain due to degenerative vertebral diseases.
ABSTRACT
To study the mechanisms of gastric tumorigenesis, we have established CSN cell line from human normal gastric mucosa, and CS12, a tumorigenic and invasive gastric cancer cell line from CSN passages. Many stem cell markers were expressed in both CSN and CS12 cells, but LGR5 and NANOG were expressed only in CS12 cells. Increased expression of homeobox A13 (HoxA13) and its downstream cascades was significant for the tumorigenic activity of CS12 cells, and was associated with recruitment of E2F-1 to HoxA13 promoter accompanied with increased trimethylation of histone H3 lysine 4 (H3K4me3) at the hypomethylated E2F motifs. Knockdown of HoxA13 caused the downregulation of long non-coding RNA HOTTIP and insulin growth factor-binding protein 3 (IGFBP-3) genes, indicating that both were targets of HoxA13. Concurrent regulation of HoxA13-HOTTIP was mediated by the mixed lineage leukemia-WD repeat domain 5 complex, which caused the trimethylation of H3K4 and then stimulated cell proliferation. HoxA13 transactivated the IGFBP-3 promoter through the HOX-binding site. Activation of IGFBP-3 stimulated the oncogenic potential and invasion activity. Increased expression of HoxA13 (63.2%) and IGFBP-3 (28.6%) was detected in human gastric cancer tissues and was found in the gastric cancer data of The Cancer Genome Atlas. Taken together, the HoxA13-HOTTIP-IGFBP-3 cascade is critical for the carcinogenic characteristics of CS12 cells.
Subject(s)
Gene Expression Regulation, Neoplastic , Homeodomain Proteins/genetics , Insulin-Like Growth Factor Binding Protein 3/genetics , RNA, Long Noncoding/genetics , Stomach Neoplasms/genetics , Animals , Carcinogenesis/genetics , Cell Line , Cell Line, Tumor , DNA Methylation , Homeodomain Proteins/metabolism , Humans , Mice, SCID , Oncogenes/genetics , RNA Interference , RNA, Long Noncoding/metabolism , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Transplantation, HeterologousABSTRACT
Indoleamine 2, 3-dioxygenase (IDO) is a rate-limiting enzyme for the tryptophan catabolism. In human and murine cells, IDO inhibits antigen-specific T cell proliferation in vitro and suppresses T cell responses to fetal alloantigens during murine pregnancy. In mice, IDO expression is an inducible feature of specific subsets of dendritic cells (DCs), and is important for T cell regulatory properties. However, the effect of IDO and tryptophan deprivation on DC functions remains unknown. We report here that when tryptophan utilization was prevented by a pharmacological inhibitor of IDO, 1-methyl tryptophan (1MT), DC activation induced by pathogenic stimulus lipopolysaccharide (LPS) or inflammatory cytokine TNF-alpha was inhibited both phenotypically and functionally. Such an effect was less remarkable when DC was stimulated by a physiological stimulus, CD40 ligand. Tryptophan deprivation during DC activation also regulated the expression of CCR5 and CXCR4, as well as DC responsiveness to chemokines. These results suggest that tryptophan usage in the microenvironment is essential for DC maturation, and may also play a role in the regulation of DC migratory behaviors.
Subject(s)
Chemokines/metabolism , Chemotaxis , Dendritic Cells/physiology , Indoleamine-Pyrrole 2,3,-Dioxygenase/metabolism , CD40 Antigens/metabolism , Cell Proliferation , Cells, Cultured , Dendritic Cells/drug effects , Dendritic Cells/enzymology , Humans , Indoleamine-Pyrrole 2,3,-Dioxygenase/antagonists & inhibitors , Lipopolysaccharides/pharmacology , Receptors, CXCR4/biosynthesis , Receptors, CXCR5 , Receptors, Chemokine , Receptors, Cytokine/biosynthesis , T-Lymphocytes/physiology , Tryptophan/analogs & derivatives , Tryptophan/metabolism , Tryptophan/pharmacology , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
The water soluble oxovanadium(IV) tetraarylporphyrin has demonstrated excellent solution stability against glutathione reduction and high potency (5 microM, 97% inhibition) in inhibiting HIV-1 replication in Hut/CCR5 cells.
Subject(s)
Anti-HIV Agents/chemistry , Metalloporphyrins/pharmacology , Vanadium Compounds/pharmacology , Anti-HIV Agents/pharmacology , Cell Line , Dose-Response Relationship, Drug , Drug Stability , Glutathione/metabolism , Humans , Metalloporphyrins/chemistry , Oxidation-Reduction , Receptors, CCR5 , Solubility , Vanadium Compounds/chemistry , Virus Replication/drug effectsABSTRACT
Silver nanoparticles fabricated in Hepes buffer exhibit potent cytoprotective and post-infected anti-HIV-1 activities toward Hut/CCR5 cells.
Subject(s)
Cytoprotection/drug effects , HEPES , HIV-1/drug effects , HIV-1/physiology , Nanoparticles , Silver/pharmacology , Apoptosis/drug effects , Buffers , Cell Line , Gold/pharmacology , Humans , Microscopy, Electron, Transmission , Nanoparticles/ultrastructure , Virus Replication/drug effectsABSTRACT
Inflammation plays a key role in coronary artery disease (CAD) and other manifestations of atherosclerosis. Recently, urinary proteins were found to be useful markers for reflecting inflammation status of different organs. To identify potential biomarker for diagnosis of CAD, we performed one-dimensional SDS-gel electrophoresis followed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Among the proteins differentially expressed in urine samples, monocyte antigen CD14 was found to be consistently expressed in higher amounts in the CAD patients as compared to normal controls. Using enzyme-linked immunosorbent assays to analyze the concentrations of CD14 in urine and serum, we confirmed that urinary CD14 levels were significantly higher in patients (n = 73) with multi-vessel and single vessel CAD than in normal control (n = 35) (P < 0.001). Logistic regression analysis further showed that urinary CD14 concentration level is associated with severity or number of diseased vessels and SYNTAX score after adjustment for potential confounders. Concomitantly, the proportion of CD14+ monocytes was significantly increased in CAD patients (59.7 ± 3.6%) as compared with healthy controls (14.9 ± 2.1%) (P < 0.001), implicating that a high level of urinary CD14 may be potentially involved in mechanism(s) leading to CAD pathogenesis. By performing shotgun proteomics, we further revealed that CD14-associated inflammatory response networks may play an essential role in CAD. In conclusion, the current study has demonstrated that release of CD14 in urine coupled with more CD14+ monocytes in CAD patients is significantly correlated with severity of CAD, pointing to the potential application of urinary CD14 as a novel noninvasive biomarker for large-scale diagnostic screening of susceptible CAD patients.
Subject(s)
Biomarkers/urine , Coronary Artery Disease/diagnosis , Coronary Artery Disease/urine , Lipopolysaccharide Receptors/urine , Proteome/metabolism , Aged , Case-Control Studies , Coronary Artery Disease/metabolism , Humans , Inflammation/metabolism , Inflammation/urine , Monocytes/metabolism , Proteomics/methods , Tandem Mass Spectrometry/methodsABSTRACT
The development of peptide-MHC tetrameric complexes heralds a new era in the study of antigen-specific T cells and their role in viral infections. However, the frequencies of tetramer-staining CD8+ T cells in fresh peripheral blood mononuclear cells (PBMCs) are usually below 1% in patients with chronic hepatitis B and C viruses (HBV and HCV) as well as human immunodeficiency virus (HIV) infections, which makes difficult the comparison and sequential evaluation of different individuals. Thus, the development of a method to enumerate efficiently antigen-specific CD8+ T cells will be clinically beneficial in monitoring the antiviral cellular immunity during therapy. We report here a modified CRI-p culture method (cytotoxic T lymphocyte response index of the epitope-peptide method), using a panel of peptides to stimulate PBMCs in bulk culture. The modified CRI-p cultured cells were, in turn, subjected to fluorescence-activated cell sorter (FACS) analysis, tetramer staining or T-cell functional assays to quantify the antiviral immunity of HLA-A2 (+) HBV and HCV patients receiving antiviral therapies. The results obtained showed that patients with a sustained response had a significantly higher increase in the frequencies of tetramer staining of virus-specific CD8+ T cells than did nonresponders. This method permits semi-quantitative determination of the relative strength of CTL activity against a panel of peptides and provides a large number of cells for FACS analysis from a single blood sampling. Significantly, it achieves high frequencies of tetramer staining of CD8+ T cells allowing the data of different individuals to be easily compared and sequentially evaluated. The mechanisms involved in this method are discussed.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Cytotoxicity Tests, Immunologic/methods , Hepatitis B, Chronic/immunology , Hepatitis C, Chronic/immunology , T-Lymphocytes, Cytotoxic/immunology , Adult , Amino Acid Sequence , Antigens, Viral/genetics , Antigens, Viral/immunology , Antiviral Agents/therapeutic use , Female , Flow Cytometry/methods , HLA-A2 Antigen/metabolism , Hepatitis B, Chronic/drug therapy , Hepatitis C, Chronic/drug therapy , Humans , Male , Middle Aged , Oligopeptides/genetics , Oligopeptides/immunologyABSTRACT
BACKGROUND: The purpose of this study was to assess the usefulness, safety, and efficacy of intra-arterial (IA) infusion chemotherapy for patients with locally advanced oral commissure cancer. METHODS: Twenty-one patients with stages III and IV squamous cell carcinoma involving the mouth angle were recruited. Methotrexate (MTX; 50 mg/day) was continuously infused into the external carotid artery for a mean period of 8 days, followed by weekly IA bolus of 25 mg MTX for a mean period of 10 weeks. RESULTS: Thirteen patients (62%) achieved a complete response (CR) and 7 patients (33%) had a partial response (PR). At a median follow-up of 69 months, the estimated 1-year, 3-year, and 5-year survival rates of the patients with CR versus PR were 100% versus 57%, 92% versus 43%, and 80% versus 43%, respectively. CONCLUSION: Our data demonstrate that continuous IA chemotherapy could achieve a competitive acceptable survival rate and improved locoregional control of advanced oral commissure cancer.
Subject(s)
Antimetabolites, Antineoplastic/administration & dosage , Carcinoma, Squamous Cell/drug therapy , Lip Neoplasms/drug therapy , Methotrexate/administration & dosage , Adult , Aged , Antimetabolites, Antineoplastic/adverse effects , Carcinoma, Squamous Cell/mortality , Carcinoma, Squamous Cell/pathology , Carotid Artery, Internal , Disease-Free Survival , Drug Administration Schedule , Female , Follow-Up Studies , Humans , Infusions, Intra-Arterial/methods , Lip Neoplasms/mortality , Lip Neoplasms/pathology , Male , Methotrexate/adverse effects , Middle Aged , Mouth Mucosa/pathology , Retrospective StudiesABSTRACT
Senescent cells show a series of alterations, including a flat and enlarged morphology, increase in nonspecific acidic ß- galactosidase activity, chromatin condensation, and changes in gene expression patterns. The onset and maintenance of senescence are regulated by two tumor suppressor proteins, p53 and Rb, whose expression is controlled by two distinct proteins, p19(Arf) and p16(Ink4a), respectively, which are encoded by the cdkn2a locus. Transcription factor Jun dimerization protein 2 (JDP2) which binds directly to histones and DNA, inhibits the acetylation and methylation of core histones and of reconstituted nucleosomes that contain JDP2-recognition DNA sequences. JDP2-deficient mouse embryonic fibroblasts are known to be resistant to replicative senescence. Oxygen induces the expression of the JDP2 gene and JDP2 then inhibits the recruitment of polycomb repressive complexes (PRCs1 and 2) to the promoter of the gene encoding p16(Ink4a), resulting in the inhibition of methylation of lysine 27 of histone H3. These findings suggest that chromatin-remodeling factors, including the PRC complex controlled by JDP2, are important players in the senescence. The newly defined mechanisms that underlie the action of oxygen in the induction of JDP2 and cellular senescence are reviewed.
Subject(s)
Cellular Senescence/physiology , Oxidative Stress/physiology , Oxygen/pharmacology , Reactive Oxygen Species/metabolism , Repressor Proteins/metabolism , Animals , Cellular Senescence/drug effects , Cellular Senescence/genetics , DNA Damage , Fibroblasts/drug effects , Fibroblasts/physiology , Histones/metabolism , Histones/physiology , Humans , Mice , Oxidative Stress/genetics , Oxygen/metabolism , Protein Binding , Repressor Proteins/genetics , Signal Transduction/genetics , Signal Transduction/physiologyABSTRACT
CD209 (DC-SIGN) is an important C-type lectin which acts a receptor of many pathogens. The single nucleotide polymorphism (SNP) -336A>G in the CD209 promoter has been demonstrated to regulate promoter activity and to be associated with several important infectious diseases, such as human immunodeficiency virus-1 (HIV-1), Mycobacterium tuberculosis, and Dengue fever. CD209 facilitates severe acute respiratory syndrome (SARS)-coronavirus spike protein-bearing pseudotype driven infection of permissive cells in vitro. In keeping with previously published findings, our in vitro studies confirmed that this SNP modulates gene promoter activity. Genetic association analysis of this SNP with clinico-pathologic outcomes in 824 serologic confirmed SARS patients showed that the -336AG/GG genotype SARS patients was associated with lower standardized lactate-dehydrogenase (LDH) levels compared with the -336AA patients (p = 0.014, odds ratio = 0.40). High LDH levels are known to be an independent predictor for poor clinical outcome, probably related to tissue destruction from immune hyperactivity. Hence, SARS patients with the CD209 -336 AA genotype carry a 60% chance of having a poorer prognosis. This association is in keeping with the role of CD209 in modulating immune response to viral infection. The relevance of these findings for other infectious diseases and inflammatory conditions would be worth investigating.