RESUMEN
Programmed cell death-1 (PD-1) is a co-inhibitory receptor that dampens immune responses upon interaction with PD-L1 and PD-L2. Although PD-1 expression on T cells is known to be activation-dependent, how cytokines modify its regulation is not fully resolved. Using polyclonal T-cell activation to study cytokine-dependent PD-1 regulation, we found that IL-2 inhibited transcriptional up-regulation of PD-1 despite the promotion of T-cell activation. The IL-2-mediated reduction in PD-1 expression augmented CD8+ T-cell activities against PD-L1-expressing target cells. To study the mechanism of PD-1 reduction, we focused on STAT5 activation in the IL-2 signaling pathway. Bioinformatic analysis suggested a novel conserved PD-1 promoter domain where NFAT and STAT5 can potentially compete with each other for binding. NFAT1 interaction with this domain revealed substantial potency in PD-1 transcription compared to STAT5A, and STAT5A overexpression could quench NFAT1-dependent PD-1 up-regulation in a sequence-specific manner. Chromatin immunoprecipitation analysis of activated T cells showed that IL-2 treatment significantly diminished the binding of NFAT1 and NFAT2 in the hypothesized competition site, while STAT5 binding to the same region was increased. These results raise the possibility that the competition of transcriptional factors might be involved in the fine-tuning of PD-1 expression by cytokines such as IL-2.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Receptor de Muerte Celular Programada 1/inmunología , Factor de Transcripción STAT5/inmunología , Animales , Células Cultivadas , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor de Muerte Celular Programada 1/deficiencia , Receptor de Muerte Celular Programada 1/genéticaRESUMEN
Azide-oligo(ethylene glycol)-terminated self-assembled monolayers (N3-OEG-SAMs) are promising interfacial structures for surface functionalization. Its many potential applications include chemical/bio-sensing and construction of surface models owing to its cross-coupling activity that originates from the azide group and oligo(ethylene glycol) (OEG) units for non-specific adsorption resistance. However, there are only a few studies and limited information, particularly on the molecular-scale structures and local cross-coupling activities of N3-OEG-SAMs, which are vital to understanding its surface properties and interfacial molecular design. In this study, molecular-scale surface structures and cross-coupling activity of azide-hexa(ethylene glycol)-terminated SAMs (N3-EG6-SAMs) were investigated using frequency modulation atomic force microscopy (FM-AFM) in liquid. The N3-EG6-SAMs were prepared on Au(111) substrates through the self-assembly of 11-azido-hexa(ethylene glycol)-undecane-1-thiol (N3-EG6-C11-HS) molecules obtained from a liquid phase. Subnanometer-resolution surface structures were visualized in an aqueous solution using a laboratory-built FM-AFM instrument. The results show a well-ordered molecular arrangement in the N3-EG6-SAM and its clean surfaces originating from the adsorption resistance property of the terminal EG6 units. Surface functionalization by the cross-coupling reaction of copper(I)-catalyzed azide-alkyne cycloaddition was observed, indicating a structural change in the form of fluctuating structures and island-shaped structures depending on the concentration of the alkyne molecules. The FM-AFM imaging enabled to provide information on the relationship between the surface structures and cross-coupling activity. These findings provide molecular-scale information on the functionalization of the N3-EG6-SAMs, which is helpful for the interfacial molecular design based on alkanethiol SAMs in many applications.
Asunto(s)
Azidas , Glicol de Etileno , Adsorción , Oro , Microscopía de Fuerza Atómica , Propiedades de SuperficieRESUMEN
Tumor hypoxia-driven accumulation of extracellular adenosine was shown to facilitate tumor evasion by engaging the immunosuppressive, intracellular cAMP-elevating A2 adenosine receptors (A2R) on tumor-reactive effector T cells, but there remains a need for careful evaluation of the limiting factors and properties of A2R blockade-enabled antitumor immunity. In studies of A2AR and/or A2BR gene-deficient mice, we found that A2AR deletion-but not A2BR deletion-liberates endogenous CD8+ T cell antitumor immunity against weakly immunogenic MCA205 sarcomas. Studies of adoptively transferred A2AR-/-, A2BR-/-, or A2AR-/-/A2BR-/- tumor-reactive T cells confirmed that immunosuppression in the tumor microenvironment was mediated by A2AR on CD8+ T cells. Treatment with A2AR antagonist mimicked A2AR gene deletion in adoptive T cell immunotherapy. This therapeutic benefit of targeting A2AR was independent of the anatomical location of tumor growth. The enhanced antitumor reactivity also led to the eradication of established intracranial tumors, which was associated with mouse survival and the maintenance of long-lasting, tumor-specific immunological memory. The blockade of the A2AR on adoptively transferred T cells by synthetic A2AR antagonist led to higher levels of IFN-γ secretion by tumor-infiltrating CD8+ T cells. These data clarify the mechanism of hypoxia-driven immunosuppression in the tumor microenvironment by A2AR on tumor-reactive CD8+ T cells and show that selective A2AR antagonists can be effective in improving the outcomes of T cell-based immunotherapies. Demonstration of the T cell dose dependency of tumor rejection points to a major limitation of current cancer immunotherapies, in which the presence of sufficient numbers of tumor-reactive T cells in a patient is not known.
Asunto(s)
Neoplasias Encefálicas/inmunología , Linfocitos T CD8-positivos/inmunología , Hipoxia/inmunología , Inmunoterapia Adoptiva/métodos , Receptor de Adenosina A2A/metabolismo , Sarcoma/inmunología , Adenosina/metabolismo , Antagonistas del Receptor de Adenosina A2/farmacología , Animales , Linfocitos T CD8-positivos/trasplante , Línea Celular Tumoral , Modelos Animales de Enfermedad , Femenino , Humanos , Tolerancia Inmunológica , Inmunidad , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor de Adenosina A2A/genética , Escape del Tumor , Microambiente TumoralRESUMEN
Immunotherapy of cancer has finally materialized following the success of immune checkpoint blockade. Since down-regulation of immune checkpoint mechanisms is beneficial in cancer treatment, it is important to ask why tumors are infamously filled with the immunosuppressive mechanisms. Indeed, immune checkpoints are physiological negative feedback mechanisms of immune activities, and the induction of such mechanisms is important in preventing excessive destruction of inflamed normal tissues. A condition commonly found in tumors and inflamed tissues is tissue hypoxia. Oxygen deprivation under hypoxic conditions by itself is immunosuppressive because proper oxygen supply could support bioenergetic demands of immune cells for optimal immune responses. However, importantly, hypoxia has been found to up-regulate a variety of immune checkpoints and to be able to drive a shift toward a more immunosuppressive environment. Moreover, extracellular adenosine, which accumulates due to tissue hypoxia, also contributes to the up-regulation of other immune checkpoints. Taken together, tissue oxygen is a key regulator of the immune response by directly affecting the energy status of immune effectors and by regulating the intensity of immunoregulatory activity in the environment. The regulators of various immune checkpoint mechanisms may represent the next focus to modulate the intensity of immune responses and to improve cancer immunotherapy.
Asunto(s)
Neoplasias/inmunología , Neoplasias/patología , Oxígeno/metabolismo , Animales , Humanos , Inmunoterapia , Neoplasias/terapiaRESUMEN
T follicular helper (TFH) cells have been shown to be critically required for the germinal center (GC) reaction where B cells undergo class switch recombination and clonal selection to generate high affinity neutralizing antibodies. However, detailed knowledge of the physiological cues within the GC microenvironment that regulate T cell help is limited. The cAMP-elevating, Gs protein-coupled A2a adenosine receptor (A2aR) is an evolutionarily conserved receptor that limits and redirects cellular immunity. However, the role of A2aR in humoral immunity and B cell differentiation is unknown. We hypothesized that the hypoxic microenvironment within the GC facilitates an extracellular adenosine-rich milieu, which serves to limit TFH frequency and function, and also promotes immunosuppressive T follicular regulatory cells (TFR). In support of this hypothesis, we found that following immunization, mice lacking A2aR (A2aRKO) exhibited a significant expansion of T follicular cells, as well as increases in TFH to TFR ratio, GC T cell frequency, GC B cell frequency, and class switching of GC B cells to IgG1. Transfer of CD4 T cells from A2aRKO or wild type donors into T cell-deficient hosts revealed that these increases were largely T cell-intrinsic. Finally, injection of A2aR agonist, CGS21680, following immunization suppressed T follicular differentiation, GC B cell frequency, and class switching of GC B cells to IgG1. Taken together, these observations point to a previously unappreciated role of GS protein-coupled A2aR in regulating humoral immunity, which may be pharmacologically targeted during vaccination or pathological states in which GC-derived autoantibodies contribute to the pathology.
Asunto(s)
Autoanticuerpos/inmunología , Centro Germinal/inmunología , Inmunidad Humoral , Cambio de Clase de Inmunoglobulina/inmunología , Inmunoglobulina G/inmunología , Receptores Purinérgicos P1/inmunología , Linfocitos T Reguladores/inmunología , Adenosina/análogos & derivados , Adenosina/farmacología , Animales , Autoanticuerpos/genética , Linfocitos B/inmunología , Cambio de Clase de Inmunoglobulina/efectos de los fármacos , Inmunoglobulina G/genética , Ratones , Ratones Noqueados , Fenetilaminas/farmacología , Receptores Purinérgicos P1/genéticaRESUMEN
Germinal centers (GCs) are anatomic sites where B cells undergo secondary diversification to produce high-affinity, class-switched Abs. We hypothesized that proliferating B cells in GCs create a hypoxic microenvironment that governs their further differentiation. Using molecular markers, we found GCs to be predominantly hypoxic. Compared to normoxia (21% O2), hypoxic culture conditions (1% O2) in vitro accelerated class switching and plasma cell formation and enhanced expression of GL-7 on B and CD4+ T cells. Reversal of GC hypoxia in vivo by breathing 60% O2 during immunization resulted in reduced frequencies of GC B cells, T follicular helper cells, and plasmacytes, as well as lower expression of ICOS on T follicular helper cells. Importantly, this reversal of GC hypoxia decreased Ag-specific serum IgG1 and reduced the frequency of IgG1+ B cells within the Ag-specific GC. Taken together, these observations reveal a critical role for hypoxia in GC B cell differentiation.
Asunto(s)
Hipoxia de la Célula , Centro Germinal/inmunología , Centro Germinal/metabolismo , Cambio de Clase de Inmunoglobulina , Recombinación Genética , Animales , Linfocitos B/inmunología , Linfocitos B/fisiología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/fisiología , Diferenciación Celular , Centro Germinal/citología , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Inmunoglobulinas/metabolismo , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Oxígeno/metabolismo , Células Plasmáticas/inmunología , Células Plasmáticas/fisiologíaRESUMEN
Extracellular adenosine regulates inflammatory responses via the A2A adenosine receptor (A2AR). A2AR deficiency results in much exaggerated acute hepatitis, indicating nonredundancy of adenosine-A2AR pathway in inhibiting immune activation. To identify a critical target of immunoregulatory effect of extracellular adenosine, we focused on NKT cells, which play an indispensable role in hepatitis. An A2AR agonist abolished NKT-cell-dependent induction of acute hepatitis by concanavalin A (Con A) or α-galactosylceramide in mice, corresponding to downregulation of activation markers and cytokines in NKT cells and of NK-cell co-activation. These results show that A2AR signaling can downregulate NKT-cell activation and suppress NKT-cell-triggered inflammatory responses. Next, we hypothesized that NKT cells might be under physiological control of the adenosine-A2AR pathway. Indeed, both Con A and α-galactosylceramide induced more severe hepatitis in A2AR-deficient mice than in WT controls. Transfer of A2AR-deficient NKT cells into A2AR-expressing recipients resulted in exaggeration of Con A-induced liver damage, suggesting that NKT-cell activation is controlled by endogenous adenosine via A2AR, and this physiological regulatory mechanism of NKT cells is critical in the control of tissue-damaging inflammation. The current study suggests the possibility to manipulate NKT-cell activity in inflammatory disorders through intervention to the adenosine-A2AR pathway.
Asunto(s)
Adenosina/inmunología , Hepatitis Animal/inmunología , Células T Asesinas Naturales/inmunología , Receptor de Adenosina A2A/inmunología , Adenosina/análogos & derivados , Adenosina/farmacología , Agonistas del Receptor de Adenosina A2/inmunología , Agonistas del Receptor de Adenosina A2/farmacología , Antagonistas del Receptor de Adenosina A2/inmunología , Antagonistas del Receptor de Adenosina A2/farmacología , Animales , Células Cultivadas , Concanavalina A , Citometría de Flujo , Galactosilceramidas , Hepatitis Animal/inducido químicamente , Hepatitis Animal/genética , Interferón gamma/inmunología , Interferón gamma/metabolismo , Hígado/inmunología , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células T Asesinas Naturales/efectos de los fármacos , Células T Asesinas Naturales/metabolismo , Fenetilaminas/inmunología , Fenetilaminas/farmacología , Receptor de Adenosina A2A/metabolismo , Triazinas/inmunología , Triazinas/farmacología , Triazoles/inmunología , Triazoles/farmacología , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Tissue hypoxia plays a key role in establishing an immunosuppressive environment in vivo by, among other effects, increasing the level of extracellular adenosine, which then signals through A2A adenosine receptor (A2AR) to elicit its immunosuppressive effect. Although the important role of the adenosine--A2AR interaction in limiting inflammation has been established, the current study revisited this issue by asking whether hypoxia can also exert its T-cell inhibitory effects even without A2AR. A similar degree of hypoxia-triggered inhibition was observed in wild-type and A2AR-deficient T cells both in vitro and, after exposure of mice to a hypoxic atmosphere, in vivo. This A2AR-independent hypoxic T-cell suppression was qualitatively and mechanistically different from immunosuppression by A2AR stimulation. The A2AR-independent hypoxic immunosuppression strongly reduced T-cell proliferation, while IFN-γ-producing activity was more susceptible to the A2AR-dependent inhibition. In contrast to the sustained functional impairment after A2AR-mediated T-cell inhibition, the A2AR-independent inhibition under hypoxia was short lived, as evidenced by the quick recovery of IFN-γ-producing activity upon re-stimulation. These data support the view that T-cell inhibition by hypoxia can be mediated by multiple mechanisms and that both A2AR and key molecules in the A2AR-independent T-cell inhibition should be targeted to overcome the hypoxia-related immunosuppression in infected tissues and tumors.
Asunto(s)
Adenosina/metabolismo , Hipoxia/inmunología , Receptor de Adenosina A2A/metabolismo , Subgrupos de Linfocitos T/inmunología , Linfocitos T Reguladores/inmunología , Animales , Proliferación Celular , Células Cultivadas , Femenino , Terapia de Inmunosupresión , Interferón gamma/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor de Adenosina A2A/genética , Microambiente TumoralRESUMEN
Metformin, a biguanide, is considered in EASD and ADA to be a first-line glucose-lowering agent for patients with type 2 diabetes. The effectiveness of metformin as an anti-diabetic drug is explained by its ability to lower blood glucose by decreasing hepatic glucose production, stimulating glucose uptake in the muscle, and increasing fatty acid oxidation in adipose tissue. The exact mechanism of this effect has not been fully understood, but metformin is thought to activate hepatic AMP-activated protein kinase (AMPK), and it suppresses liver glucagon signal by increasing AMP due to inhibition of mitochondrial respiratory complex I. Caution is advised to avoid use of metformin in patients at risk for lactic acidosis (e.g., in patients with advanced renal and liver insufficiency, infection, dehydration, alcoholism, or in those using diuretics or SGLT2 inhibitor).
Asunto(s)
Biguanidas/uso terapéutico , Biguanidas/farmacología , Diabetes Mellitus Tipo 2/tratamiento farmacológico , HumanosRESUMEN
Hypoxia-adenosinergic suppression and redirection of the immune response has been implicated in the regulation of antipathogen and antitumor immunity, with hypoxia-inducible factor 1α (HIF-1α) playing a major role. In this study, we investigated the role of isoform I.1, a quantitatively minor alternative isoform of HIF-1α, in antibacterial immunity and sepsis survival. By using the cecal ligation and puncture model of bacterial peritonitis, we studied the function of I.1 isoform in T cells using mice with total I.1 isoform deficiency and mice with T-cell-targeted I.1 knockdown. We found that genetic deletion of the I.1 isoform resulted in enhanced resistance to septic lethality, significantly reduced bacterial load in peripheral blood, increased M1 macrophage polarization, augmented levels of proinflammatory cytokines in serum, and significantly decreased levels of the anti-inflammatory cytokine IL-10. Our data suggest a previously unrecognized immunosuppressive role for the I.1 isoform in T cells during bacterial sepsis. We interpret these data as indicative that the activation-inducible isoform I.1 hinders the contribution of T cells to the antibacterial response by affecting M1/M2 macrophage polarization and microbicidal function.
Asunto(s)
Infecciones Bacterianas/genética , Infecciones Bacterianas/inmunología , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Peritonitis/genética , Peritonitis/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Animales , Infecciones Bacterianas/mortalidad , Modelos Animales de Enfermedad , Subunidad alfa del Factor 1 Inducible por Hipoxia/deficiencia , Activación de Linfocitos/inmunología , Macrófagos/inmunología , Masculino , Ratones , Ratones Noqueados , Especificidad de Órganos/genética , Peritonitis/mortalidad , Isoformas de Proteínas , Receptores de Antígenos de Linfocitos T/metabolismo , Sepsis/genética , Sepsis/inmunología , Sepsis/mortalidad , Activación TranscripcionalRESUMEN
BACKGROUND: Liver damage by ischemia and reperfusion injury is a risk factor for morbidity and mortality after liver surgery. Postoperative oxygen treatment is routinely applied in the postanesthesia and intensive care unit after liver surgery. The risks of aggravating the injury by increasing inspiratory oxygen from 21 to 60% in the postoperative period were investigated in mice. METHODS: Parameters of liver injury were compared after induction of hepatic ischemia-reperfusion injury, by clamping the left liver lobe for 45 min, and reperfusion for 24 h either under normoxic (21% oxygen) or hyperoxic (60% oxygen) conditions (n=22 per group). The extent of tissue injury and oxidative responses was analyzed in the presence or absence of polymorphonuclear leukocytes, functional Kupffer cells, and the p47phox unit of the nicotinamide adenine dinucleotide phosphate oxidase (n=6 to 11 per group). RESULTS: Compared with postoperative normoxic conditions, hyperoxia increased cell damage (glutamate-pyruvate transaminase: 1,870 [±968 SD] vs. 60% 2,981 [±1,038 SD], 21 vs. 60% oxygen, in U/l as mean±SD; P<0.01), liver weights (341±52 vs. 383±44, 21 vs. 60% oxygen, in mg as mean±SD; P=0.02), damage scores (1.9±0.8 vs. 3.1±1.0, 21 vs. 60% oxygen, score as mean±SD; P=0.02), and reactive oxygen species (15.0±12.0 vs. 30.4±19.2, 21 vs. 60% oxygen, in µmol/l as mean±SD; P<0.05). The aggravation of the tissue damaging effects as a result of hyperoxia was not seen in mice with depletions of polymorphonuclear leukocytes or Kupffer cells, or with nonfunctioning nicotinamide adenine dinucleotide phosphate oxidase. CONCLUSION: Liver injury after ischemia was significantly aggravated by hyperoxia as a consequence of immune cell-mediated oxidative burst. Further studies are needed to elucidate whether routine delivery of high inspirational oxygen concentrations postoperatively should be limited.
Asunto(s)
Lesión Pulmonar Aguda/etiología , Lesión Pulmonar Aguda/patología , Hiperoxia/complicaciones , Hiperoxia/patología , Complicaciones Posoperatorias/patología , Animales , Hepatocitos/patología , Macrófagos del Hígado/metabolismo , Macrófagos del Hígado/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Neutrófilos/metabolismo , Neutrófilos/patología , Oxígeno/análisis , Especies Reactivas de Oxígeno , Daño por Reperfusión/patologíaRESUMEN
We evaluated the effect of sitagliptin on glycemic control, endogenous insulin secretion, and beta cell function in Japanese patients with type 2 diabetes mellitus (T2DM) receiving a combination of oral antidiabetics and basal insulin analog glargine (basal-supported oral therapy [BOT]). Twenty-one patients showing inadequate glycemic control with BOT were given dipeptidylpeptidase-4 inhibitor (DPP-4I) sitagliptin at 50 mg/day for 12 weeks. Clinical markers of glycemic control, HbA1c, glycated albumin (GA), and 1,5-anhydroglucitol (1,5-AG), were measured before and 4 and 12 weeks after the start of sitagliptin. A 2-hour morning meal test was performed upon enrollment and at 12 weeks, and plasma glucose (PG), serum C-peptide, and plasma intact proinsulin (PI) were measured. HbA1c, GA, and 1,5-AG at 4 and 12 weeks were significantly improved over enrollment levels. The area under the PG concentration curve (AUC-PG) during the meal test at 12 weeks was significantly reduced (from 350 ± 17 mg ï½¥ hr/dL before sitagliptin treatment to 338 ± 21 mg ï½¥ hr/dL [mean ± SE], P < 0.05,); the AUC-C-peptide was unchanged (from 3.4 ± 0.4 ng ï½¥ hr/mL to 3.6 ± 0.5 ng ï½¥ hr/mL). However, both fasting and 2-hour PI/C-peptide ratios at 12 weeks were significantly decreased (from 13.3 ± 2.3 to 11.1 ± 2.0 [P < 0 .05] and from 9.5 ± 1.6 to 5.3 ± 0.9 [P < 0.01], respectively). Adding sitagliptin to BOT in Japanese T2DM patients appears to improve glycemic control without increasing endogenous insulin secretion and to reduce fasting and 2-hour postprandial PI/C-peptide ratios.
Asunto(s)
Diabetes Mellitus Tipo 2/tratamiento farmacológico , Inhibidores de la Dipeptidil-Peptidasa IV/uso terapéutico , Resistencia a Medicamentos , Hiperglucemia/prevención & control , Células Secretoras de Insulina/efectos de los fármacos , Proinsulina/metabolismo , Pirazinas/uso terapéutico , Triazoles/uso terapéutico , Administración Oral , Anciano , Algoritmos , Biomarcadores/sangre , Péptido C/sangre , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/etnología , Diabetes Mellitus Tipo 2/fisiopatología , Inhibidores de la Dipeptidil-Peptidasa IV/administración & dosificación , Monitoreo de Drogas , Quimioterapia Combinada , Ácidos Grasos no Esterificados/antagonistas & inhibidores , Ácidos Grasos no Esterificados/sangre , Femenino , Hemoglobina Glucada/análisis , Humanos , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/uso terapéutico , Insulina Glargina , Insulina de Acción Prolongada/administración & dosificación , Insulina de Acción Prolongada/uso terapéutico , Células Secretoras de Insulina/metabolismo , Japón , Masculino , Proinsulina/sangre , Pirazinas/administración & dosificación , Fosfato de Sitagliptina , Triazoles/administración & dosificaciónRESUMEN
The aim of the present study was to evaluate the effect of insulin glargine (Gla) (as part of basal-supported oral therapy) on endogenous insulin secretion and beta-cell function in type 2 diabetic patients. In 33 insulin-naive patients showing poor glycemic control on treatment with sulfonylurea (SU)-based OADs without DPP4 inhibitors, once-daily injection of Gla was added without changing OADs, and the dose of Gla was titrated to attain a fasting plasma glucose (FPG) <110 mg/dL over 24 weeks. Morning meal tests were done at baseline, 12 weeks and 24 weeks. FPG and 2-hour plasma glucose (2HPG) and serum C-peptide (FCPR and 2HCPR) were measured 3 times, while serum intact proinsulin (FPI and 2HPI) was measured at baseline and 24 weeks. Levels of FPG, FCPR, 2HPG, and HbA1c were significantly reduced from baseline at 24 weeks (176±52 to 117±27 mg/dL, p<0.01; 2.0±0.9 to 1.6±1.0 ng/mL, p<0.01; 257±53 to 202±27 mg/dL, p<0.01; and 8.4±0.9 to 7.3±0.6%, p<0.01, Mean±SD), but 2HCPR was unchanged. The patients were divided into two groups depending on whether FPG at 24 weeks was <110 mg/dL or not: attained group (n=15) and not attained group (n=18). The dose of Gla did not differ between the two groups, but the 2HPI/2HCPR ratio at 24 weeks showed a significant decrease from baseline in the attained group. Supplementation with Gla improved glycemic control and maintained intrinsic basal insulin secretion, without changing 2-hour postprandial secretion. Achieving good glycemic control with an FPG<110 mg/dL by adding Gla decreased the 2HPI/2HCPR ratio at 24 weeks.
Asunto(s)
Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/administración & dosificación , Insulina de Acción Prolongada/administración & dosificación , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/fisiología , Insulina/metabolismo , Anciano , Glucemia/análisis , Péptido C/sangre , Ayuno , Femenino , Hemoglobina Glucada/análisis , Humanos , Insulina Glargina , Secreción de Insulina , Japón , Masculino , Persona de Mediana Edad , Periodo Posprandial , Compuestos de Sulfonilurea/administración & dosificaciónRESUMEN
Background: Increased levels of plasminogen activator inhibitor-1 (PAI-1) in tumors have been found to correlate with poor clinical outcomes in patients with cancer. Although abundant data support the involvement of PAI-1 in cancer progression, whether PAI-1 contributes to tumor immune surveillance remains unclear. The purposes of this study are to determine whether PAI-1 regulates the expression of immune checkpoint molecules to suppresses the immune response to cancer and demonstrate the potential of PAI-1 inhibition for cancer therapy. Methods: The effects of PAI-1 on the expression of the immune checkpoint molecule programmed cell death ligand 1 (PD-L1) were investigated in several human and murine tumor cell lines. In addition, we generated tumor-bearing mice and evaluated the effects of a PAI-1 inhibitor on tumor progression or on the tumor infiltration of cells involved in tumor immunity either alone or in combination with immune checkpoint inhibitors. Results: PAI-1 induces PD-L1 expression through the JAK/STAT signaling pathway in several types of tumor cells and surrounding cells. Blockade of PAI-1 impedes PD-L1 induction in tumor cells, significantly reducing the abundance of immunosuppressive cells at the tumor site and increasing cytotoxic T-cell infiltration, ultimately leading to tumor regression. The anti-tumor effect elicited by the PAI-1 inhibitor is abolished in immunodeficient mice, suggesting that PAI-1 blockade induces tumor regression by stimulating the immune system. Moreover, combining a PAI-1 inhibitor with an immune checkpoint inhibitor significantly increases tumor regression. Conclusions: PAI-1 protects tumors from immune surveillance by increasing PD-L1 expression; hence, therapeutic PAI-1 blockade may prove valuable in treating malignant tumors.
Asunto(s)
Antígeno B7-H1 , Inhibidor 1 de Activador Plasminogénico , Escape del Tumor , Animales , Femenino , Humanos , Ratones , Antígeno B7-H1/metabolismo , Antígeno B7-H1/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Evasión Inmune , Ratones Endogámicos C57BL , Neoplasias/inmunología , Neoplasias/metabolismo , Inhibidor 1 de Activador Plasminogénico/metabolismo , Inhibidor 1 de Activador Plasminogénico/genética , Transducción de Señal , Microambiente Tumoral/inmunologíaRESUMEN
Cadmium (Cd) contamination of farmland soils is a growing concern because of its highly toxic impact on ecosystems and human health. Chelator-assisted washing and chemical immobilization are effective remediation strategies for Cd-contaminated soils. Ethylenediaminetetraacetic acid (EDTA) has traditionally been used for soil washing, but its persistence in the environment and subsequent toxicity have raised significant ecological concerns. Consequently, biodegradable chelators have gained increasing attention as eco-friendly alternatives to the persistent chelator, EDTA. Therefore, this study evaluated the performance and efficacy of three biodegradable chelators: L-glutamate-N,N'-diacetic acid (GLDA), methylglycine-diacetic acid (MGDA), and 3-hydroxy-2,2'-iminodisuccinic acid (HIDS) in comparison to EDTA for remediating a real Cd-contaminated agricultural soil. The influence of treatment parameters, including chelator variants, washing time, chelator concentration, solution pH, and liquid-to-soil ratio (L/S) on Cd extraction was studied and optimized to attain the maximum removal rate. Following chelator-assisted washing, the efficacy of a stabilization preference combining FeCl3 and CaO in reducing the leaching potential of residual Cd in chelator-washed soil residues was also investigated. GLDA demonstrated comparable Cd extraction efficiency to EDTA, and the Cd extraction efficiency was found to be positively correlated with the soil washing parameters. However, under the optimized conditions (chelator concentration: 10 mmol L-1; washing time: 3 h; solution pH: 3; L/S ratio: 10:1), GLDA exhibited a higher Cd extraction rate than EDTA or the other chelators. Furthermore, a post-treatment process incorporating FeCl3 and CaO substantially diminished the water-leachable Cd content in the resultant soil residues. The proposed remediation strategy, which combines chemically assisted washing and stabilization, could be a practical option for extracting bulk Cd from soil and reducing the leaching potential of residual Cd.
Asunto(s)
Cloruros , Restauración y Remediación Ambiental , Compuestos Férricos , Metales Pesados , Contaminantes del Suelo , Humanos , Cadmio , Ácido Edético/química , Metales Pesados/análisis , Ecosistema , Contaminantes del Suelo/análisis , Quelantes/química , Suelo/químicaRESUMEN
Antimicrobial treatment strategies must improve to reduce the high mortality rates in septic patients. In noninfectious models of acute inflammation, activation of A2B adenosine receptors (A2BR) in extracellular adenosine-rich microenvironments causes immunosuppression. We examined A2BR in antibacterial responses in the cecal ligation and puncture (CLP) model of sepsis. Antagonism of A2BR significantly increased survival, enhanced bacterial phagocytosis, and decreased IL-6 and MIP-2 (a CXC chemokine) levels after CLP in outbred (ICR/CD-1) mice. During the CLP-induced septic response in A2BR knockout mice, hemodynamic parameters were improved compared with wild-type mice in addition to better survival and decreased plasma IL-6 levels. A2BR deficiency resulted in a dramatic 4-log reduction in peritoneal bacteria. The mechanism of these improvements was due to enhanced macrophage phagocytic activity without augmenting neutrophil phagocytosis of bacteria. Following ex vivo LPS stimulation, septic macrophages from A2BR knockout mice had increased IL-6 and TNF-α secretion compared with wild-type mice. A therapeutic intervention with A2BR blockade was studied by using a plasma biomarker to direct therapy to those mice predicted to die. Pharmacological blockade of A2BR even 32 h after the onset of sepsis increased survival by 65% in those mice predicted to die. Thus, even the late treatment with an A2BR antagonist significantly improved survival of mice (ICR/CD-1) that were otherwise determined to die according to plasma IL-6 levels. Our findings of enhanced bacterial clearance and host survival suggest that antagonism of A2BRs offers a therapeutic target to improve macrophage function in a late treatment protocol that improves sepsis survival.
Asunto(s)
Inmunosupresores/antagonistas & inhibidores , Inmunosupresores/metabolismo , Macrófagos/inmunología , Fagocitosis/inmunología , Receptor de Adenosina A2B/metabolismo , Sepsis/inmunología , Regulación hacia Arriba/inmunología , Animales , Antígenos CD1/biosíntesis , Ciego , Femenino , Interleucina-6/antagonistas & inhibidores , Interleucina-6/metabolismo , Ligadura , Macrófagos/microbiología , Macrófagos/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Ratones Noqueados , Ratones Transgénicos , Fagocitosis/genética , Punciones , Receptor de Adenosina A2B/deficiencia , Receptor de Adenosina A2B/genética , Sepsis/genética , Sepsis/mortalidad , Tasa de Supervivencia , Regulación hacia Arriba/genéticaRESUMEN
Insulin glulisine (Glu) is a rapidly-acting insulin analog with a faster onset of action than the other insulin analogs of its class, which are insulin aspart (Asp) and insulin lispro (Lisp). While insulin Glu is usually injected just before meals, postprandial injection may help to avoid unexpected postprandial hypoglycemia or hyperglycemia by adjusting the insulin dosage according to food intake. However, the effect of postprandial insulin Glu on the glucose profile has not been evaluated. The aim of this study was to compare daily glucose excursion by continuous glucose monitoring (CGM) between multiple daily doses of preprandial insulin Asp or postprandial insulin Glu. In a randomized cross-over trial, we performed CGM to evaluate the 48-hour glucose profile during treatment with the same dosage of insulin Asp just before each meal in 12 hospitalized patients with type 2 diabetes. Patients also received the same dosage of long-acting insulin glargine at bedtime. The average glucose level, standard deviation of the glucose level, mean amplitude of glucose excursion, and daily glucose profile did not differ between preprandial Asp and postprandial Glu. The incidence of hypoglycemic episodes (glucose level<70 mg/dL with or without symptoms) and the area under the curve of glucose<70 mg/dL also did not differ between the two insulin regimens. Multiple daily injections of preprandial Asp and postprandial Glu achieved the same daily glucose excursion profile. Postprandial injection of Glu may provide greater flexibility for patients who require insulin therapy.
Asunto(s)
Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hiperglucemia/prevención & control , Hipoglucemia/prevención & control , Hipoglucemiantes/uso terapéutico , Insulina Aspart/uso terapéutico , Insulina/análogos & derivados , Adulto , Anciano , Glucemia/análisis , Estudios Cruzados , Complicaciones de la Diabetes/prevención & control , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/complicaciones , Esquema de Medicación , Quimioterapia Combinada/efectos adversos , Femenino , Hemoglobina Glucada/análisis , Humanos , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/efectos adversos , Insulina/administración & dosificación , Insulina/efectos adversos , Insulina/uso terapéutico , Insulina Aspart/administración & dosificación , Insulina Aspart/efectos adversos , Insulina Glargina , Insulina de Acción Prolongada/administración & dosificación , Insulina de Acción Prolongada/uso terapéutico , Masculino , Persona de Mediana Edad , Monitoreo AmbulatorioRESUMEN
The PD-1 receptor triggers a negative immunoregulatory mechanism that prevents overactivation of immune cells and subsequent inflammatory diseases. Because of its biological significance, PD-1 has been a drug target for modulating immune responses. Immunoenhancing anti-PD-1 blocking antibodies have become a widely used cancer treatment; however, little is known about the required characteristics for anti-PD-1 antibodies to be capable of stimulating immunosuppressive activity. Here, we show that PD-1 agonists exist in the group of anti-PD-1 antibodies recognizing the membrane-proximal extracellular region in sharp contrast to the binding of the membrane-distal region by blocking antibodies. This trend was consistent in an analysis of 81 anti-human PD-1 monoclonal antibodies. Because PD-1 agonist antibodies trigger immunosuppressive signaling by cross-linking PD-1 molecules, Fc engineering to enhance FcγRIIB binding of PD-1 agonist antibodies notably improved human T cell inhibition. A PD-1 agonist antibody suppressed inflammation in murine disease models, indicating its clinical potential for treatment of various inflammatory disorders, including autoimmune diseases.
Asunto(s)
Anticuerpos Monoclonales , Linfocitos T , Animales , Ratones , Anticuerpos Bloqueadores/metabolismo , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Monoclonales/metabolismo , HumanosRESUMEN
Excavated debris (soil and rock) contaminated with geogenic arsenic (As) is an increasing concern for regulatory organizations and construction stakeholders. Chelator-assisted soil flushing is a promising method for practical on-site remediation of As-contaminated soil, offering technical, economic, and environmental benefits. Ethylenediaminetetraacetic acid (EDTA) is the most prevalent chelator used for remediating As-contaminated soil. However, the extensive environmental persistence and potential toxicity of EDTA necessitate the exploration of eco-compliant alternatives. In this study, the feasibility of the conventional flushing method pump-and-treat and two newly designed immersion and sprinkling techniques were evaluated at the laboratory scale (small-scale laboratory experiments) for the on-site treatment of As-contaminated excavated debris. Two biodegradable chelators, L-glutamic acid-N,N'-diacetic acid (GLDA) and 3-hydroxy-2,2'-iminodisuccinic acid (HIDS), were examined as eco-friendly substitutes for EDTA. Additionally, this study highlights a useful post-treatment measure to ensure minimal mobility of residual As in the chelator-treated debris residues. The pump-and-treat method displayed rapid As-remediation (t, 3 h), but it required a substantial volume of washing solution (100 mL g-1). Conversely, the immersion technique demonstrated an excellent As-extraction rate using a relatively smaller washing solution (0.33 mL g-1) and shorter immersion time (t, 3 h). In contrast, the sprinkling technique showed an increased As-extraction rate over an extended period (t, 48 h). Among the chelators employed, the biodegradable chelator HIDS (10 mmol L-1; pH, 3) exhibited the highest As-extraction efficiency. Furthermore, the post-treatment of chelator-treated debris with FeCl3 and CaO successfully reduced the leachable As content below the permissible limit.
Asunto(s)
Arsénico , Restauración y Remediación Ambiental , Isópodos , Contaminantes del Suelo , Animales , Arsénico/análisis , Ácido Edético/química , Contaminantes del Suelo/análisis , Quelantes/química , Suelo/químicaRESUMEN
During the long-term storage of cells, it is necessary to inhibit ice crystal formation by adding cryoprotectants. Non-cell-permeable cryoprotectants have high osmotic pressure which dehydrates cells, indirectly suppressing intracellular ice crystal formation. However, the high osmotic pressure and dehydration often damage cells. Emerging polymer-type non-cell-permeable cryoprotectants form matrices surrounding cells. These matrices inhibit the influx of extracellular ice nuclei that trigger intracellular ice crystal formation. However, these polymer-type cryoprotectants also require high osmotic pressure to exert an effective cryoprotecting effect. In this study, we designed a poly(zwitterion) (polyZI) that forms firm matrices around cells based on their high affinity to cell membranes. The polyZI successfully cryopreserved freeze-vulnerable cells under isotonic conditions. These matrices also controlled osmotic pressure by adsorbing and desorbing NaCl depending on the temperature, which is a suitable feature for isotonic cryopreservation. Although cell proliferation was delayed by the cellular matrices, washing with a sucrose solution improved proliferation.