RESUMEN

Ca2+-dependent gene regulation controls several functions to determine the fate of the cells. Proteins of the nuclear factor of activated T-cells (NFAT) family are Ca2+ sensitive transcription factors that control the cell growth, proliferation and insulin secretion in ß-cells. Translocation of NFAT proteins to the nucleus occurs in a sequence of events that starts with activating calmodulin-dependent phosphatase calcineurin in a Ca2+-dependent manner, which dephosphorylates the NFAT proteins and leads to their translocation to the nucleus. Here, we examined the role of IP3-generating agonists and near-UV light in the induction of NFATc3 migration to the nucleus in the pancreatic ß-cell line INS-1. Our results show that IP3 generation yields cytosolic Ca2+ rise and NFATc3 translocation. Moreover, near-UV light exposure generates reactive oxygen species (ROS), resulting in cytosolic Ca2+ spiking via the L-type Ca2+ channel and triggers NFATc3 translocation to the nucleus. Using the mitochondria as a Ca2+ buffering tool, we showed that ROS-induced cytosolic Ca2+ spiking, not the ROS themselves, was the triggering mechanism of nuclear import of NFATc3. Collectively, this study reveals the mechanism of near-UV light induced NFATc3 migration.

Asunto(s)

Señalización del Calcio , Factores de Transcripción NFATC/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Rayos Ultravioleta , Animales , Canales de Calcio Tipo L/metabolismo , Línea Celular Tumoral , Inositol 1,4,5-Trifosfato/metabolismo , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Ratas

RESUMEN

Insulin deficiency in type 2 diabetes mellitus (DM) involves a decline in both pancreatic ß-cell mass and function. Enhancing ß-cell preservation represents an important therapeutic strategy to treat type 2 DM. Far-infrared (FIR) radiation has been found to induce promyelocytic leukemia zinc finger protein (PLZF) activation to protect the vascular endothelium in diabetic mice. The influence of FIR on ß-cell preservation is unknown. Our previous study reveals that the biologically effective wavelength of FIR is 8-10 µm. In the present study, we investigated the biological effects of FIR (8-10 µm) on both survival and insulin secretion function of ß-cells. FIR reduced pancreatic islets loss and increased insulin secretion in nicotinamide-streptozotocin-induced DM mice, but only promoted insulin secretion in DM PLZF-/- mice. FIR-upregulated PLZF to induce an anti-apoptotic effect in a ß cell line RIN-m5f. FIR also upregulated mitochondrial function and the ratio of NAD+/NADH, and then induced Sirtuin1 (Sirt1) expression. The mitochondria Complex I inhibitor rotenone blocked FIR-induced PLZF and Sirt1. The Sirt1 inhibitor EX527 and Sirt1 siRNA inhibited FIR-induced PLZF and insulin respectively. Sirt1 upregulation also increased CaV1.2 expression and calcium influx that promotes insulin secretion in ß-cells. In summary, FIR-enhanced mitochondrial function prevents ß-cell apoptosis and enhances insulin secretion in DM mice through the Sirt1 pathway.

Asunto(s)

Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/radioterapia , Rayos Infrarrojos , Células Secretoras de Insulina/patología , Células Secretoras de Insulina/efectos de la radiación , Sirtuina 1/metabolismo , Sirtuina 1/efectos de la radiación , Animales , Apoptosis/genética , Apoptosis/efectos de la radiación , Canales de Calcio Tipo L/metabolismo , Canales de Calcio Tipo L/efectos de la radiación , Prueba de Tolerancia a la Glucosa , Secreción de Insulina/efectos de la radiación , Células Secretoras de Insulina/metabolismo , Islotes Pancreáticos/patología , Islotes Pancreáticos/efectos de la radiación , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Niacinamida , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/efectos de la radiación , Sirtuina 1/antagonistas & inhibidores , Análisis de Supervivencia , Regulación hacia Arriba

RESUMEN

In type 2 diabetes mellitus (T2DM), the overload of glucose and lipids can promote oxidative stress and inflammatory responses and contribute to the failure of beta cells. However, therapies that can modulate the function of beta cells and thus prevent their failure have not been well explored. In this study, beta cell injury model was established with palmitic acid (PA) to simulate the lipotoxicity (high-fat diet) found in T2DM. Sonodynamic therapy (SDT), a novel physicochemical treatment, was applied to treat injured beta cells. We found that SDT had specific effects on mitochondria and induced transient large amount of mitochondrial reactive oxygen species (ROS) production in beta cells. SDT also improved the morphology and function of abnormal mitochondria, inhibited inflammatory response and reduced beta cell dysfunction. The improvement of mitochondria was mediated by PINK1/Parkin-dependent mitophagy. Additionally, SDT rescued the transcription of PINK1 mRNA which was blocked by PA treatment, thus providing abundant PINK1 for mitophagy. Moreover, SDT also increased insulin secretion from beta cells. The protective effects of SDT were abrogated when mitophagy was inhibited by cyclosporin A (CsA). In summary, SDT potently inhibits lipotoxicity-induced beta cell failure via PINK1/Parkin-dependent mitophagy, providing theoretical guidance for T2DM treatment in aspects of islet protection.

Asunto(s)

Diabetes Mellitus Tipo 2/terapia , Células Secretoras de Insulina/metabolismo , Mitocondrias/efectos de la radiación , Mitofagia/efectos de la radiación , Proteínas Quinasas/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ondas Ultrasónicas , Animales , Proteínas Portadoras/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Supervivencia Celular/efectos de la radiación , Diabetes Mellitus Tipo 2/metabolismo , Inflamación/metabolismo , Inflamación/radioterapia , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/efectos de la radiación , Mitocondrias/genética , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , Mitofagia/efectos de los fármacos , Mitofagia/genética , Ácido Palmítico/farmacología , Ácido Palmítico/toxicidad , Proteínas Quinasas/genética , Protoporfirinas/metabolismo , Ratas , Especies Reactivas de Oxígeno/metabolismo , Receptores de GABA-A/metabolismo , Ubiquitina-Proteína Ligasas/genética

RESUMEN

FBXW7, a classic tumor suppressor, is a substrate recognition subunit of the Skp1-cullin-F-box (SCF) ubiquitin ligase that targets oncoproteins for ubiquitination and degradation. We recently found that FBXW7 is recruited to DNA damage sites to facilitate nonhomologous end-joining (NHEJ). The detailed underlying molecular mechanism, however, remains elusive. Here we report that the WD40 domain of FBXW7, which is responsible for substrate binding and frequently mutated in human cancers, binds to poly(ADP-ribose) (PAR) immediately following DNA damage and mediates rapid recruitment of FBXW7 to DNA damage sites, whereas ATM-mediated FBXW7 phosphorylation promotes its retention at DNA damage sites. Cancer-associated arginine mutations in the WD40 domain (R465H, R479Q and R505C) abolish both FBXW7 interaction with PAR and recruitment to DNA damage sites, causing inhibition of XRCC4 polyubiquitination and NHEJ. Furthermore, inhibition or silencing of poly(ADP-ribose) polymerase 1 (PARP1) inhibits PAR-mediated recruitment of FBXW7 to the DNA damage sites. Taken together, our study demonstrates that the WD40 domain of FBXW7 is a novel PAR-binding motif that facilitates early recruitment of FBXW7 to DNA damage sites for subsequent NHEJ repair. Abrogation of this ability seen in cancer-derived FBXW7 mutations provides a molecular mechanism for defective DNA repair, eventually leading to genome instability.

Asunto(s)

Reparación del ADN por Unión de Extremidades , Proteína 7 que Contiene Repeticiones F-Box-WD/genética , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli Adenosina Difosfato Ribosa/metabolismo , Factor de Células Madre/genética , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Sitios de Unión , Línea Celular , Línea Celular Tumoral , Supervivencia Celular/efectos de la radiación , Daño del ADN , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Proteína 7 que Contiene Repeticiones F-Box-WD/química , Proteína 7 que Contiene Repeticiones F-Box-WD/metabolismo , Fibroblastos/metabolismo , Fibroblastos/efectos de la radiación , Fibroblastos/ultraestructura , Rayos gamma , Células HCT116 , Humanos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Células Secretoras de Insulina/ultraestructura , Modelos Moleculares , Mutación , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Poli Adenosina Difosfato Ribosa/química , Unión Proteica , Dominios Proteicos , Dominios y Motivos de Interacción de Proteínas , Estructura Secundaria de Proteína , Factor de Células Madre/metabolismo , Ubiquitinación/efectos de la radiación

RESUMEN

CD19 chimeric antigen receptors (CARs) have demonstrated great efficacy against a range of B cell malignancies. However, antigen escape and, more generally, heterogeneous antigen expression pose a challenge to applying CAR therapy to a wide range of cancers. We find that low-dose radiation sensitizes tumor cells to immune rejection by locally activated CAR T cells. In a model of pancreatic adenocarcinoma heterogeneously expressing sialyl Lewis-A (sLeA), we show that not only sLeA+ but also sLeA- tumor cells exposed to low-dose radiation become susceptible to CAR therapy, reducing antigen-negative tumor relapse. RNA sequencing analysis of low-dose radiation-exposed tumors reveals the transcriptional signature of cells highly sensitive to TRAIL-mediated death. We find that sLeA-targeted CAR T cells produce TRAIL upon engaging sLeA+ tumor cells, and eliminate sLeA- tumor cells previously exposed to systemic or local low-dose radiation in a TRAIL-dependent manner. These findings enhance the prospects for successfully applying CAR therapy to heterogeneous solid tumors. Local radiation is integral to many tumors' standard of care and can be easily implemented as a CAR conditioning regimen.

Asunto(s)

Antígenos CD19/uso terapéutico , Inmunoterapia Adoptiva , Neoplasias Pancreáticas/inmunología , Neoplasias Pancreáticas/radioterapia , Ligando Inductor de Apoptosis Relacionado con TNF/genética , Animales , Antígenos CD19/inmunología , Antígenos de Neoplasias/química , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/efectos de la radiación , Antígeno CA-19-9 , Terapia Combinada , Modelos Animales de Enfermedad , Humanos , Células Secretoras de Insulina/inmunología , Células Secretoras de Insulina/efectos de la radiación , Ratones , Oligosacáridos/química , Oligosacáridos/inmunología , Oligosacáridos/uso terapéutico , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Radiación , Dosis de Radiación , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/uso terapéutico , Análisis de Secuencia de ARN , Ligando Inductor de Apoptosis Relacionado con TNF/inmunología

RESUMEN

Pancreatic ß-cell insulin production is orchestrated by a complex circuitry involving intracellular elements including cyclic AMP (cAMP). Tackling aberrations in glucose-stimulated insulin release such as in diabetes with pharmacological agents, which boost the secretory capacity of ß-cells, is linked to adverse side effects. We hypothesized that a photoactivatable adenylyl cyclase (PAC) can be employed to modulate cAMP in ß-cells with light thereby enhancing insulin secretion. To that end, the PAC gene from Beggiatoa (bPAC) was delivered to ß-cells. A cAMP increase was noted within 5 minutes of photostimulation and a significant drop at 12 minutes post-illumination. The concomitant augmented insulin secretion was comparable to that from ß-cells treated with secretagogues. Greater insulin release was also observed over repeated cycles of photoinduction without adverse effects on viability and proliferation. Furthermore, the expression and activation of bPAC increased cAMP and insulin secretion in murine islets and in ß-cell pseudoislets, which displayed a more pronounced light-triggered hormone secretion compared to that of ß-cell monolayers. Calcium channel blocking curtailed the enhanced insulin response due to bPAC activity. This optogenetic system with modulation of cAMP and insulin release can be employed for the study of ß-cell function and for enabling new therapeutic modalities for diabetes.

Asunto(s)

Secreción de Insulina/efectos de la radiación , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Luz , Optogenética , Adenilil Ciclasas/genética , Adenilil Ciclasas/metabolismo , Animales , Línea Celular , AMP Cíclico/metabolismo , Regulación de la Expresión Génica/efectos de la radiación , Humanos , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/efectos de la radiación , Ratones , Optogenética/métodos

RESUMEN

Type 2 diabetes mellitus is a complex metabolic disease that has reached epidemic proportions in the United States and around the world. This disease is characterized by loss of insulin secretion and, eventually, destruction of insulin-producing pancreatic beta cells. Controlling type 2 diabetes is often difficult as pharmacological management routinely requires complex therapy with multiple medications, and loses its effectiveness over time. The objective of this study was to explore the effectiveness of a novel, non-pharmacological approach that uses the application of ultrasound energy to augment insulin release from rat INS 832/13 beta cells. The cells were exposed to unfocused ultrasound for 5 min at a peak intensity of 1 W/cm2 and frequencies of 400 kHz, 600 kHz, 800 kHz and 1 MHz. Insulin release was measured with enzyme-linked immunosorbent assay and cell viability was assessed via the trypan blue dye exclusion test. A marked release (approximately 150 ng/106 cells, p < 0.05) of insulin was observed when beta cells were exposed to ultrasound at 400 and 600 kHz as compared with their initial control values; however, this release was accompanied by a substantial loss in cell viability. Ultrasound application at frequencies of 800 kHz resulted in 24 ng/106 cells released insulin (p < 0.05) as compared with its unstimulated base level, while retaining cell viability. Insulin release from beta cells caused by application of 800-kHz ultrasound was comparable to that reported by the secretagogue glucose, thus operating within physiological secretory capacity of these cells. Ultrasound has potential as a novel and alternative method to current approaches aimed at correcting secretory deficiencies in patients with type 2 diabetes.

Asunto(s)

Diabetes Mellitus Tipo 2/terapia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Insulina/biosíntesis , Sonicación/métodos , Terapia por Ultrasonido/métodos , Animales , Línea Celular , Preparaciones de Acción Retardada/administración & dosificación , Diabetes Mellitus Tipo 2/diagnóstico por imagen , Ondas de Choque de Alta Energía , Hipoglucemiantes/administración & dosificación , Ratas , Resultado del Tratamiento

RESUMEN

Radiolabeled exendin is used for non-invasive quantification of beta cells in the islets of Langerhans in vivo. High accumulation of radiolabeled exendin in the islets raised concerns about possible radiation-induced damage to these islets in man. In this work, islet absorbed doses resulting from exendin-imaging were calculated by combining whole organ dosimetry with small scale dosimetry for the islets. Our model contains the tissues with high accumulation of radiolabeled exendin: kidneys, pancreas and islets. As input for the model, data from a clinical study (radiolabeled exendin distribution in the human body) and from a preclinical study with Biobreeding Diabetes Prone (BBDP) rats (islet-to-exocrine uptake ratio, beta cell mass) were used. We simulated 111In-exendin and 68Ga-exendin absorbed doses in patients with differences in gender, islet size, beta cell mass and radiopharmaceutical uptake in the kidneys. In all simulated cases the islet absorbed dose was small, maximum 1.38 mGy for 68Ga and 66.0 mGy for 111In. The two sources mainly contributing to the islet absorbed dose are the kidneys (33-61%) and the islet self-dose (7.5-57%). In conclusion, all islet absorbed doses are low (<70 mGy), so even repeated imaging will hardly increase the risk on diabetes.

Asunto(s)

Células Secretoras de Insulina/patología , Islotes Pancreáticos/patología , Riñón/metabolismo , Traumatismos por Radiación/diagnóstico , Radiometría/métodos , Adulto , Animales , Recuento de Células , Diabetes Mellitus/etiología , Diabetes Mellitus/genética , Modelos Animales de Enfermedad , Femenino , Radioisótopos de Galio/química , Radioisótopos de Galio/metabolismo , Humanos , Radioisótopos de Indio/química , Radioisótopos de Indio/metabolismo , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Péptidos y Proteínas de Señalización Intercelular , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/efectos de la radiación , Riñón/efectos de la radiación , Masculino , Persona de Mediana Edad , Péptidos/química , Péptidos/metabolismo , Radiación , Dosis de Radiación , Ratas , Ratas Mutantes , Adulto Joven

RESUMEN

The intestinal L cell is the principal source of glucagon-like peptide-1 (GLP-1), a major determinant of insulin release. Because GLP-1 secretion is regulated in a circadian manner in rodents, we investigated whether the activity of the human L cell is also time sensitive. Rhythmic fluctuations in the mRNA levels of canonical clock genes were found in the human NCI-H716 L cell model, which also showed a time-dependent pattern in their response to well-established secretagogues. A diurnal variation in GLP-1 responses to identical meals (850 kcal), served 12 h apart in the normal dark (2300) and light (1100) periods, was also observed in male volunteers maintained under standard sleep and light conditions. These findings suggest the existence of a daily pattern of activity in the human L cell. Moreover, we separately tested the short-term effects of sleep deprivation and nocturnal light exposure on basal and postprandial GLP-1, insulin, and glucose levels in the same volunteers. Sleep deprivation with nocturnal light exposure disrupted the melatonin and cortisol profiles and increased insulin resistance. Moreover, it also induced profound derangements in GLP-1 and insulin responses such that postprandial GLP-1 and insulin levels were markedly elevated and the normal variation in GLP-1 responses was abrogated. These alterations were not observed in sleep-deprived participants maintained under dark conditions, indicating a direct effect of light on the mechanisms that regulate glucose homeostasis. Accordingly, the metabolic abnormalities known to occur in shift workers may be related to the effects of irregular light-dark cycles on these glucoregulatory pathways.

Asunto(s)

Péptido 1 Similar al Glucagón/metabolismo , Células Secretoras de Glucagón/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Privación de Sueño/metabolismo , Adolescente , Adulto , Proteínas CLOCK/genética , Células Cultivadas , Ritmo Circadiano/fisiología , Células Secretoras de Glucagón/efectos de la radiación , Voluntarios Sanos , Humanos , Secreción de Insulina , Células Secretoras de Insulina/efectos de la radiación , Luz , Masculino , Redes y Vías Metabólicas/genética , Redes y Vías Metabólicas/efectos de la radiación , Factores de Tiempo , Adulto Joven

RESUMEN

Photoactivation of caged biomolecules has become a powerful approach to study cellular signalling events. Here we report a method for anchoring and uncaging biomolecules exclusively at the outer leaflet of the plasma membrane by employing a photocleavable, sulfonated coumarin derivative. The novel caging group allows quantifying the reaction progress and efficiency of uncaging reactions in a live-cell microscopy setup, thereby greatly improving the control of uncaging experiments. We synthesized arachidonic acid derivatives bearing the new negatively charged or a neutral, membrane-permeant coumarin caging group to locally induce signalling either at the plasma membrane or on internal membranes in ß-cells and brain slices derived from C57B1/6 mice. Uncaging at the plasma membrane triggers a strong enhancement of calcium oscillations in ß-cells and a pronounced potentiation of synaptic transmission while uncaging inside cells blocks calcium oscillations in ß-cells and causes a more transient effect on neuronal transmission, respectively. The precise subcellular site of arachidonic acid release is therefore crucial for signalling outcome in two independent systems.

Asunto(s)

Ácido Araquidónico/metabolismo , Membrana Celular/metabolismo , Animales , Ácido Araquidónico/química , Calcio/metabolismo , Señalización del Calcio/efectos de la radiación , Membrana Celular/efectos de la radiación , Cumarinas/química , Cumarinas/metabolismo , Células HeLa , Humanos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de la radiación , Luz , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Neuronas/efectos de la radiación

RESUMEN

The present study assessed the ability of optogenetics techniques to provide a better understanding of the control of insulin secretion, particularly regarding pancreatic ß-cell function in homeostasis and pathological conditions such as diabetes mellitus (DM). We used optogenetics to investigate whether insulin secretion and blood glucose homeostasis could be controlled by regulating intracellular calcium ion concentrations ([Ca(2+)]i) in a mouse pancreatic ß-cell line (MIN6) transfected with the optogenetic protein channelrhodopsin-2 (ChR2). The ChR2-transfected MIN6 (ChR2-MIN6) cells secreted insulin following irradiation with a laser (470 nm). The increase in [Ca(2+)]i was accompanied by elevated levels of messenger RNAs that encode calcium/calmodulin-dependent protein kinase II delta and adenylate cyclase 1. ChR2-MIN6 cells suspended in matrigel were inoculated into streptozotocin-induced diabetic mice that were then subjected to a glucose tolerance test. Laser irradiation of these mice caused a significant decrease in blood glucose, and the irradiated implanted cells expressed insulin. These findings demonstrate the power of optogenetics to precisely and efficiently controlled insulin secretion by pancreatic ß-cells 'on demand', in contrast to techniques using growth factors or chemical inducers. Optogenetic technology shows great promise for understanding the mechanisms of glucose homeostasis and for developing treatments for metabolic diseases such as DM.

Asunto(s)

Diabetes Mellitus Experimental/radioterapia , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Terapia por Luz de Baja Intensidad , Optogenética , Adenilil Ciclasas/metabolismo , Animales , Glucemia/metabolismo , Calcio/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Células Cultivadas , Channelrhodopsins , Diabetes Mellitus Experimental/metabolismo , Secreción de Insulina , Células Secretoras de Insulina/efectos de la radiación , Terapia por Luz de Baja Intensidad/métodos , Ratones , Estreptozocina

RESUMEN

Promotion of insulin-secreting ß-cell regeneration in patients with diabetes is a promising approach for diabetes therapy, which can contribute to rescue the uncontrolled hyperglycemia. Low-power laser irradiation (LPLI) has been demonstrated to regulate multiple physiological processes both in vitro and in vivo through activation of various signaling pathways. In the present study, we showed that LPLI promoted ß-cell replication and cell cycle progression through activation of Akt1/GSK3ß isoform-specific signaling axis. Inhibition of PI3-K/Akt or GSK3 with specific inhibitors dramatically reduced or increased LPLI-induced ß-cell replication, revealing Akt/GSK3 signaling axis was involved in ß-cell replication and survival upon LPLI treatment. Furthermore, the results of shRNA-mediated knock down of Akt/GSK3 isoforms revealed that Akt1/GSK3ß isoform-specific signaling axis regulated ß-cell replication and survival in response to LPLI, but not Akt2/GSK3α. The mechanism by which LPLI promoted ß-cell replication through Akt1/GSK3ß signaling axis involved activation of ß-catenin and down-regulation of p21. Taken together, these observations suggest that Akt1/GSK3ß isoform signaling axis play a key role in ß-cell replication and survival induced by LPLI. Moreover, our findings suggest that activation of Akt1/GSK3ß isoform signaling axis by LPLI may provide guidance in practical applications for ß-cell regenerative therapies.

Asunto(s)

Glucógeno Sintasa Quinasa 3/metabolismo , Células Secretoras de Insulina/fisiología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Regeneración/efectos de la radiación , Animales , Ciclo Celular/efectos de la radiación , Proliferación Celular/efectos de la radiación , Glucógeno Sintasa Quinasa 3/genética , Glucógeno Sintasa Quinasa 3 beta , Células Secretoras de Insulina/efectos de la radiación , Isoenzimas/genética , Isoenzimas/metabolismo , Terapia por Luz de Baja Intensidad , Masculino , Proteínas Proto-Oncogénicas c-akt/genética , Ratas , Transducción de Señal/efectos de la radiación

RESUMEN

Azobenzene photoresponsive elements can be installed on sulfonylureas, yielding optical control over pancreatic beta cell function and insulin release. An obstacle to such photopharmacological approaches remains the use of ultraviolet-blue illumination. Herein, we synthesize and test a novel yellow light-activated sulfonylurea based on a heterocyclic azobenzene bearing a push-pull system.

Asunto(s)

Compuestos Azo/química , Células Secretoras de Insulina/efectos de la radiación , Insulina/agonistas , Islotes Pancreáticos/efectos de la radiación , Compuestos de Sulfonilurea/química , Animales , Calcio/metabolismo , Células Cultivadas , Diazóxido/farmacología , Regulación de la Expresión Génica , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Células HEK293 , Humanos , Hipoglucemiantes/farmacología , Insulina/metabolismo , Secreción de Insulina , Células Secretoras de Insulina/citología , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Islotes Pancreáticos/citología , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Luz , Ratones , Procesos Fotoquímicos , Potasio/metabolismo , Transducción de Señal , Compuestos de Sulfonilurea/farmacología , Receptores de Sulfonilureas/genética , Receptores de Sulfonilureas/metabolismo

RESUMEN

Sulfonylureas are widely prescribed for the treatment of type 2 diabetes mellitus (T2DM). Through their actions on ATP-sensitive potassium (KATP) channels, sulfonylureas boost insulin release from the pancreatic beta cell mass to restore glucose homeostasis. A limitation of these compounds is the elevated risk of developing hypoglycemia and cardiovascular disease, both potentially fatal complications. Here, we describe the design and development of a photoswitchable sulfonylurea, JB253, which reversibly and repeatedly blocks KATP channel activity following exposure to violet-blue light. Using in situ imaging and hormone assays, we further show that JB253 bestows light sensitivity upon rodent and human pancreatic beta cell function. Thus, JB253 enables the optical control of insulin release and may offer a valuable research tool for the interrogation of KATP channel function in health and T2DM.

Asunto(s)

Hipoglucemiantes/farmacología , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Compuestos de Sulfonilurea/farmacología , Animales , Transporte Biológico/efectos de los fármacos , Transporte Biológico/efectos de la radiación , Línea Celular , Femenino , Humanos , Hipoglucemiantes/síntesis química , Secreción de Insulina , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/efectos de la radiación , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/efectos de la radiación , Canales KATP/metabolismo , Luz , Masculino , Ratones , Ratones Endogámicos C57BL , Compuestos de Sulfonilurea/síntesis química

RESUMEN

BACKGROUND: Ultraviolet irradiation by sun exposure has been associated with both harms and benefits to metabolic health. OBJECTIVE: The objective of this study was to determine whether unprotected daily sun exposure is associated with the prevalence of diabetes and explore the underlying mechanism. METHODS: We analyzed the Korean National Health and Nutrition Survey V from 2010 to 2011. Participants 19-60 years of age were asked about the average amount of time they had been exposed to direct sunlight per day since the age of 19. We categorized participants into three groups with different levels of lifetime daily sun exposure and explored the association of sun exposure with the prevalence of diabetes. RESULTS: The risk of diabetes was higher in subjects with more than 5h of unprotected sun exposure per day, with an odds ratio of 2.39 (95% CI 1.75-3.25), compared to those with less than 2h of sun exposure, and the association remained significant after adjusting for diabetes risk factors. Long-term sun exposure was associated with increased central obesity and the possibility of an increase in visceral adiposity, especially among women, and with decrease in beta cell function and peripheral adiposity or percent body fat in men. CONCLUSIONS: Our study provides a cutoff for upper limit of sun exposure and suggests unprotected daily sun exposure for more than 5h should be avoided to prevent diabetes. Increased central adiposity and decreased beta cell function were observed in women and men, respectively, who had long-term unprotected daily sun exposure.

Asunto(s)

Adiposidad , Diabetes Mellitus/epidemiología , Encuestas Nutricionales , Obesidad/epidemiología , Luz Solar/efectos adversos , Adulto , Diabetes Mellitus/etiología , Diabetes Mellitus/patología , Femenino , Humanos , Resistencia a la Insulina/efectos de la radiación , Células Secretoras de Insulina/patología , Células Secretoras de Insulina/efectos de la radiación , Masculino , Persona de Mediana Edad , Obesidad/etiología , Obesidad/patología , Prevalencia , República de Corea/epidemiología , Caracteres Sexuales , Adulto Joven

RESUMEN

OBJECTIVE: Extensive use of mobile phones has been accompanied by a common public debate about possible adverse effects on human health. No study has been published so far to establish any association between the fastest growing innovation of mobile phone and fasting blood glucose. The aim was to determine the effects of exposure to electromagnetic field radiation generated by mobile phones on fasting blood glucose in Wistar Albino rats. MATERIALS AND METHODS: 40 Male Albino rats (Wistar Strain) were divided into 5 equally numerous groups. Group A served as the control one, group B received mobile phone radiation for less than 15 min/day, group C: 15-30 min/day, group D: 31-45 min/day, and group E: 46-60 min/day for a total period of 3 months. Fasting blood glucose was determined by using Spectrophotometer and serum insulin by Enzyme-linked Immunosorbent Assay (ELISA). The Homeostatic Model (HOMA-B) was applied for the assessment of ß-cell function and (HOMA-IR) for resistance to insulin. RESULTS: Wister Albino rats exposed to mobile phone radiation for longer than 15 min a day for a total period of 3 months had significantly higher fasting blood glucose (p < 0.015) and serum insulin (p < 0.01) compared to the control group. HOMA-IR for insulin resistance was significantly increased (p < 0.003) in the groups that were exposed for 15-30 and 46-60 min/day compared to the control rats. CONCLUSION: The results of the present study show an association between long-term exposure to activated mobile phones and increase in fasting blood glucose and serum insulin in Albino rats.

Asunto(s)

Glucemia/efectos de la radiación , Teléfono Celular , Campos Electromagnéticos , Exposición a Riesgos Ambientales , Resistencia a la Insulina/efectos de la radiación , Animales , Homeostasis/efectos de la radiación , Células Secretoras de Insulina/efectos de la radiación , Masculino , Ratas

RESUMEN

Hydrogels provide three-dimensional frameworks with tissue-like elasticity and high permeability for culturing therapeutically relevant cells or tissues. While recent research efforts have created diverse macromer chemistry to form hydrogels, the mechanisms of hydrogel polymerization for in situ cell encapsulation remain limited. Hydrogels prepared from chain-growth photopolymerization of poly(ethylene glycol) diacrylate (PEGDA) are commonly used to encapsulate cells. However, free radical associated cell damage poses significant limitation for this gel platform. More recently, PEG hydrogels formed by thiol-ene photo-click chemistry have been developed for cell encapsulation. While both chain-growth and step-growth photopolymerizations offer spatial-temporal control over polymerization kinetics, step-growth thiol-ene hydrogels offer more diverse and preferential properties. Here, we report the superior properties of step-growth thiol-ene click hydrogels, including cytocompatibility of the reactions, improved hydrogel physical properties, and the ability for 3D culture of pancreatic ß-cells. Cells encapsulated in thiol-ene hydrogels formed spherical clusters naturally and were retrieved via rapid chymotrypsin-mediated gel erosion. The recovered cell spheroids released insulin in response to glucose treatment, demonstrating the cytocompatibility of thiol-ene hydrogels and the enzymatic mechanism of cell spheroids recovery. Thiol-ene click reactions provide an attractive means to fabricate PEG hydrogels with superior gel properties for in situ cell encapsulation, as well as to generate and recover 3D cellular structures for regenerative medicine applications.

Asunto(s)

Química Clic/métodos , Hidrogeles/síntesis química , Hidrogeles/farmacología , Células Secretoras de Insulina/citología , Luz , Esferoides Celulares/citología , Compuestos de Sulfhidrilo/síntesis química , Animales , Fenómenos Biofísicos/efectos de los fármacos , Fenómenos Biofísicos/efectos de la radiación , Recuento de Células , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Células Inmovilizadas/citología , Células Inmovilizadas/efectos de los fármacos , Células Inmovilizadas/efectos de la radiación , Quimotripsina/metabolismo , Reactivos de Enlaces Cruzados/farmacología , Módulo de Elasticidad/efectos de los fármacos , Módulo de Elasticidad/efectos de la radiación , Hidrogeles/química , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/efectos de la radiación , Ratones , Polietilenglicoles/química , Polimerizacion/efectos de los fármacos , Polimerizacion/efectos de la radiación , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/efectos de la radiación , Compuestos de Sulfhidrilo/química

RESUMEN

The magnetic flux density of MRI for clinical diagnosis has been steadily increasing. However, there remains very little biological data regarding the effect of strong static magnetic fields (SMFs) on human health. To evaluate the effects of strong SMFs on biological systems, we cultured insulin-secreting cells under exposure to sham and SMF conditions (3-10 T of magnetic flux density, and 0-41.7 T/m of magnetic field gradient) for 0.5 or 1 h, and analyzed insulin secretion, mRNA expression, glucose-stimulated insulin secretion, insulin content, cell proliferation and cell number. Exposure to SMF with a high magnetic field gradient for 1 h significantly increased insulin secretion and insulin 1 mRNA expression. Exposure to SMF with a high magnetic flux density for 0.5 h significantly enhanced responsiveness to glucose stimulation. Exposure to SMF did not affect the insulin content, cell proliferation or cell number. Our results suggested that MRI systems with a higher magnetic flux density might not cause cell proliferative or functional damages on insulin-secreting cells, and that SMF with a high magnetic field gradient might be used clinically after thorough in vivo investigations are conducted.

Asunto(s)

Células Secretoras de Insulina/fisiología , Células Secretoras de Insulina/efectos de la radiación , Insulina/metabolismo , Imagen por Resonancia Magnética , Animales , Línea Celular , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Relación Dosis-Respuesta en la Radiación , Campos Electromagnéticos , Secreción de Insulina , Células Secretoras de Insulina/citología , Dosis de Radiación , Ratas

RESUMEN

AIMS/HYPOTHESIS: Recent studies have shown that bone marrow transplantation reduces hyperglycaemia in a mouse model of diabetes induced by streptozotocin. However, the essential factors for the improvement of hyperglycaemia by bone marrow transplantation have not been fully elucidated. The aim of this study was to search for such factors. METHODS: We investigated the effect of irradiation to whole body, to abdomen alone or to whole body excluding abdomen, followed by infusion or no infusion of bone marrow cells. We also investigated the effect of bone marrow transplantation on beta cell-specific vascular endothelial growth factor-A gene (Vegfa) knockout mice. RESULTS: Bone marrow transplantation improved streptozotocin-induced hyperglycaemia and partially restored islet mass. This change was associated with increased islet vascularisation. Among the other methods investigated, low-dose irradiation of the whole body without infusion of bone marrow cells also improved blood glucose level. In streptozotocin-treated beta cell-specific Vegfa knockout mice, which exhibit impaired islet vascularisation, bone marrow transplantation neither improved hyperglycaemia, relative beta cell mass nor islet vascularisation. CONCLUSION/INTERPRETATION: Our results indicate that whole body irradiation is essential and sufficient for restoration of beta cell mass after streptozotocin treatment independent of infusion of bone marrow cells. Vascular endothelial growth factor-A produced in beta cells is also essential for this phenomenon.

Asunto(s)

Trasplante de Médula Ósea , Diabetes Mellitus Experimental/cirugía , Células Secretoras de Insulina/citología , Factor A de Crecimiento Endotelial Vascular/farmacología , Irradiación Corporal Total , Animales , Modelos Animales de Enfermedad , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/fisiología , Prueba de Tolerancia a la Glucosa , Hiperglucemia/prevención & control , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/efectos de la radiación , Masculino , Ratones , Ratones Endogámicos C57BL , Trasplante Isogénico

RESUMEN

To evaluate the effects of extremely low frequency magnetic field (ELFMF) on beta-cell survival and function, we cultured a hamster-derived insulin-secreting cell line (HIT-T15), which exhibits responsiveness to glucose in a semi-physiological range, under exposure to sham and ELFMF conditions, and assessed cell survival and function. We used our previously developed ELFMF exposure unit (a sinusoidal magnetic field at a frequency of 60 Hz, 5 mT) to culture cells under exposure to ELFMF conditions. We found that exposure to ELFMF for 5 days in the absence of glucose increased cell number, exposure for 2 days in the absence of glucose and for 5 days with 100 mg/dl glucose increased the insulin secretion to the culture medium, and exposure for 2 and 5 days with 40 and 100 mg/dl glucose increased intracellular insulin concentration in HIT-T15 cells. The increase in cell number under apoptotic culture conditions by exposure to ELFMF could lead to new therapeutic concepts in the treatment of diabetes. The ELFMF-induced increase in intracellular insulin concentration could be utilized to develop culture conditions to enhance intracellular insulin concentration in insulin-secreting cells that would be useful for cell transplantation to cure diabetes mellitus.

Asunto(s)

Glucosa/metabolismo , Células Secretoras de Insulina/fisiología , Células Secretoras de Insulina/efectos de la radiación , Insulina/metabolismo , Animales , Línea Celular , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Cricetinae , Relación Dosis-Respuesta en la Radiación , Campos Electromagnéticos , Células Secretoras de Insulina/citología , Dosis de Radiación