RESUMEN
PURPOSE: The influence of the dipeptidyl peptidase-IV inhibitor gemigliptin alone or in combination with the histone deacetylase inhibitor PXD101 on survival of thyroid carcinoma cells was investigated. METHODS: SW1736, TPC-1, 8505C and BCPAP human thyroid carcinoma cells were used. To assess cell survival, cell viability, the percentage of viable cells and dead cells, cytotoxic activity, ATP levels and FACS analysis were measured. To validate the impact of gemigliptin combined with PXD101, the interactions were estimated by obtaining combination index in cells treated with two agents. RESULTS: In cells treated with gemigliptin or PXD101, cell viability, the percentage of viable cells and ATP levels were reduced, and the percentage of dead cells and cytotoxic activity were elevated. In cells treated with both gemigliptin and PXD101, compared with PXD101 alone, cell death was augmented, and all of the combination index values were lower than 1.0, suggesting the synergism between gemigliptin and PXD101. The percentage of apoptotic cells, and the protein levels of Bcl2 and cleaved poly (ADP-ribose) polymerase were elevated, and the protein levels of xIAP and survivin were reduced. The protein levels of phospho-Akt and phospho-AMPK were elevated, and cell migration was reduced. CONCLUSIONS: Our results demonstrate that gemigliptin induces cytotoxicity in thyroid carcinoma cells. Moreover, gemigliptin has a synergistic activity with PXD101 in the induction of cell death through involvement of Bcl2 family proteins, xIAP and survivin as well as mediation of Akt and AMPK in thyroid carcinoma cells.
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Biomarcadores de Tumor/metabolismo , Inhibidores de la Dipeptidil-Peptidasa IV/farmacología , Sinergismo Farmacológico , Inhibidores de Histona Desacetilasas/farmacología , Ácidos Hidroxámicos/farmacología , Piperidonas/farmacología , Pirimidinas/farmacología , Sulfonamidas/farmacología , Neoplasias de la Tiroides/patología , Apoptosis/efectos de los fármacos , Supervivencia Celular , Humanos , Transducción de Señal , Neoplasias de la Tiroides/tratamiento farmacológico , Neoplasias de la Tiroides/metabolismo , Células Tumorales CultivadasRESUMEN
AIMS: In this study, we compared the glucose-lowering effectiveness of insulin analogues and their combination according to baseline glycemic status in patients with type 2 diabetes (T2D) from the A1 chieve(®) study conducted in Korea. METHODS: This sub-analysis from the A1 chieve(®) study was a 24-week prospective, multicenter, non-interventional, open-labelled study. Of the 4058 patients, 3074 patients who had their HbA1c level measured at baseline were included in this sub-analysis. We classified patients into three groups according to baseline HbA1c levels: group I (HbA1c < 7.5%), group II (7.5% ≤ HbA1c < 9.0%) and group III (HbA1c ≥ 9.0%). RESULTS: Patients in group I showed no significant HbA1c reduction with any insulin regimens (detemir, aspart, detemir and aspart or biphasic aspart 30 (Novo Nordisk A/S, DK-2880 Bagsvaerd, Denmark) after 24 weeks of treatment. In group II, although HbA1c was decreased for all insulin regimens, there was no difference in mean HbA1c reduction among the four insulin regimens. In patients with a high baseline HbA1c level (group III), mean HbA1c reduction was the greatest in patients on a basal-bolus regimen (detemir and aspart, -3.50%) and lowest in patients on a bolus regimen (aspart, -1.81%; p < 0.001). CONCLUSION: For optimal glycaemic control, a basal-bolus regimen may be adequate for Korean patients with poorly controlled T2D (HbA1c ≥ 9.0%).
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Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Insulina/análogos & derivados , Adulto , Anciano , Femenino , Hemoglobina Glucada/análisis , Humanos , Insulina/uso terapéutico , Insulina Aspart/uso terapéutico , Insulina Detemir/uso terapéutico , Insulina Glargina/uso terapéutico , Insulina de Acción Prolongada/uso terapéutico , Masculino , Persona de Mediana Edad , Estudios ProspectivosRESUMEN
Aim of the present study was to evaluate the effect of apigenin in combination with BRAFV600E inhibitor PLX4032 on cell survival, and to investigate the influence of Akt inhibition on the combined effect of apigenin and PLX4032 in ATC cells harboring BRAFV600E. In 8505C and FRO cells harboring BRAFV600E, after treatment of apigenin and PLX4032, the cell viability decreased, and the percentage of dead cells increased in a time- and concentration-dependent manner, respectively. In apigenin- and PLX4032- treated cells, compared with apigenin alone-treated cells, the cell viability was lessened, and the percentage of dead cells was multiplied. In the addition of PLX4032 to apigenin, compared with the treatment of apigenin alone, the protein levels of cleaved PARP-1 and cleaved caspase-3 were elevated, and phospho-ERK protein levels were reduced, and the protein levels of total ERK, c-Myc, BRAF, phospho-Akt, phospho-p70S6K and phospho-4EBP1 were not varied. Compared with the treatment of PLX4032 alone, phosphop70S6K protein levels were reduced, and the other protein levels were not altered. Phospho-ERK protein levels were reduced only in 8505C cells. Under the co-treatment of apigenin and PLX4032, administration of the PI3K inhibitor wortmannin further decreased the cell viability, and increased the percentage of dead cells. In conclusion, our results suggest that PLX4032 augments apigenin-induced cytotoxicity in ATC cells harboring BRAFV600E. Moreover, Akt suppression potentiates the combined effect of apigenin and PLX4032 in ATC cells harboring BRAFV600E.
Asunto(s)
Apigenina/farmacología , Indoles/farmacología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Sulfonamidas/farmacología , Neoplasias de la Tiroides/tratamiento farmacológico , Androstadienos/farmacología , Apigenina/uso terapéutico , Caspasa 3/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Humanos , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Proteínas Proto-Oncogénicas B-raf/metabolismo , Carcinoma Anaplásico de Tiroides , Vemurafenib , WortmaninaRESUMEN
OBJECTIVE: The efficient transfer of genes into intact islets is difficult since islets exist as clusters of differentiated cells with little replication potential. Cell proliferation in response to growth factors is known to be accompanied by loosening of cell-to-cell contacts and increasing paracellular permeability. In this study, we investigated whether gene delivery into intact islet cells was facilitated by modulating ß-cell proliferation. METHODS: Isolated rat islets were pretreated with glucagon-like peptide (GLP)-1 or human growth hormone for 24 hours, or with 300 mg/dL of glucose for 48 hours before transduction with a suboptimal dose of recombinant adenoviral vector expressing green fluorescent protein (GFP) and ß-galactosidase (multiplicity of infection of 25). Transduction efficiency was assessed by measuring ß-galactosidase activity and GFP expression using enzyme-linked immunosorbent assay, flow cytometry, and fluorescence microscopy. The numbers of 7-aminoactinomycin D-positive dead cells and 5-ethynyl-2-deoxyuridine (EdU)-positive proliferating cells were also monitored using flow cytometry and fluorescence microscopy. RESULTS: The transduction efficiency of rat islet cells by a suboptimal dose of viral vector was significantly improved by GLP-1 pretreatment, accompanied by enhanced cell viability and cell proliferation. An increased GFP expression in islet cells after GLP-1 pretreatment was observed among the increased numbers of EdU-positive proliferating cells. CONCLUSION: Pretreatment of rat islets with GLP-1 enhanced the transduction efficiency of an adenoviral vector, reducing viral dose burden while improving islet cell viability. From a therapeutic standpoint, genetic modification of pancreatic islets combined with GLP-1 pretreatment may be a promising option for ex vivo gene therapy prior to islet transplantation.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Péptido 1 Similar al Glucagón/farmacología , Islotes Pancreáticos/efectos de los fármacos , Transducción Genética , Transfección , Adenoviridae/genética , Animales , Supervivencia Celular/efectos de los fármacos , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Vectores Genéticos , Glucosa/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Hormona de Crecimiento Humana/farmacología , Islotes Pancreáticos/metabolismo , Masculino , Microscopía Fluorescente , Ratas , Ratas Sprague-Dawley , Técnicas de Cultivo de Tejidos , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
SU6656 is a small-molecule indolinone that selectively inhibits Src family kinase and induces death of cancer cells. The aim of the present study was to investigate the influence of SU6656 on cell survival and to assess the role of p21 and PI3K/Akt signaling in cell survival resulting from SU6656 treatment in anaplastic thyroid carcinoma (ATC) cells. When 8505C, CAL62, and FRO ATC cells were treated with SU6656, the viability of 8505C and CAL62 ATC cells decreased only after treatment with SU6656 at a dosage of 100 µM for 72 h, while the viability of FRO ATC cells decreased after treatment with SU6656 in a concentration- and time-dependent manner. Cell viability was not changed by pretreatment with the broad-spectrum caspase inhibitor z-VAD-fmk. Phospho-Src protein levels were reduced, and p21 protein levels were elevated. Phospho-ERK1/2 protein levels were multiplied without alteration of total ERK1/2, total Akt, and phospho-Akt protein levels. Regarding FRO ATC cells, the decrement of cell viability, the increment of cleaved PARP-1 protein levels, and the decrement of phospho-Src protein levels were shown in p21 siRNA- or LY294002-pretreated cells compared to SU6656-treated control cells. ERK1/2 siRNA transfection did not affect cell viability and protein levels of cleaved PARP-1, p21, and Akt. In conclusion, these results suggest that SU6656 induces caspase-independent death of FRO ATC cells by overcoming the resistance mechanism involving p21 and Akt. Suppression of p21 and Akt enhances the cytotoxic effect of SU6656 in FRO ATC cells.
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Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Indoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Sulfonamidas/farmacología , Neoplasias de la Tiroides/fisiopatología , Caspasas/genética , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/genética , Transducción de Señal/efectos de los fármacos , Neoplasias de la Tiroides/tratamiento farmacológico , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/metabolismoRESUMEN
SU5416, vascular endothelial cell growth factor receptor inhibitor, suppresses hypoxia-induced angiogenesis, growth, proliferation, and metastasis in cancer cells. CCAAT/enhancer-binding protein-homologous protein (CHOP) has pivotal roles in regulation of growth and survival. In the present study, we evaluated the effects of SU5416 on cell survival, p21, and PI3K/Akt signal pathway in FRO anaplastic thyroid carcinoma (ATC) cells. Moreover, we investigated the roles of CHOP in cell survival under condition of SU5416 treatment in FRO ATC cells. After SU5416 treatment, cell viability, PARP-1, and caspase-3 protein levels were not changed. p53 and p27 protein levels decreased while p21 protein levels increased. Phospho-Akt protein levels were not altered. In SU5416-treated situation, cell viability was not different before and after administration of either p21 siRNA or LY294002 whereas it was lessened after co-administration of p21 siRNA and LY294002. Compared to SU5416 treatment alone, cell viability was reduced with CHOP plasmid but it was unchanged with CHOP siRNA. PARP-1 and caspase-3 protein levels with CHOP plasmid were elevated whereas the protein levels with CHOP siRNA were similar. While CHOP plasmid transfection diminished p21 and phospho-Akt protein levels, CHOP siRNA transfection did not alter the protein levels. In conclusion, these results suggest that CHOP may sensitize FRO ATC cells to SU5416 thereby inhibiting cell survival by modulating p21 and PI3K/Akt signal pathway. Furthermore, these findings imply that CHOP may be a possible candidate as the chemosensitizing factor for induction of cytotoxicity in ATC cells exposed to SU5416.
Asunto(s)
Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Indoles/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Pirroles/farmacología , Transducción de Señal/efectos de los fármacos , Neoplasias de la Tiroides/enzimología , Neoplasias de la Tiroides/patología , Factor de Transcripción CHOP/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Técnicas de Silenciamiento del Gen , Humanos , Carcinoma Anaplásico de Tiroides , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Alpha-lipoic acid (ALA) has been shown to modulate cell death via PI3K/Akt signal pathway in various cells. In the present study, the effects of ALA on cell death and PI3K/Akt signal pathway linked to cell death-related proteins during endoplasmic reticulum (ER) stress in FRTL5 thyroid cells were evaluated. In FRTL5 thyroid cells, cell viability increased by ALA pretreatment in tunicamycin (TN)-treated cells. When TN was treated, CCAAT/enhancer-binding protein-homologous protein (CHOP) and Bax protein levels were elevated while Bcl-2 protein levels were reduced. ALA diminished CHOP and Bax protein levels, and augmented Bcl-2 protein levels in TN-treated cells. After exposure to TN, phospho-Akt protein levels were repressed whereas total Akt protein levels were not changed. ALA increased phospho-Akt protein levels but not total Akt protein levels in both non-TN-treated and TN-treated cells. After LY294002 administration in non-TN-treated cells, cell viability was reduced, and CHOP and Bax protein levels were elevated, and Bcl-2 protein levels were reduced. The CHOP, Bcl-2 and Bax protein levels were not different after LY294002 administration in TN-treated cells. LY294002 and wortmannin decreased cell viability, and increased CHOP and Bax protein levels, and decreased Bcl-2 protein levels in ALA-pretreated and TN-treated cells. In conclusion, these results suggest that ER stress may induce cell death by modulating PI3K/Akt signal pathway linked to cell death-related proteins in FRTL5 thyroid cells. Moreover, these findings imply that ALA may ameliorate ER stress-induced cell death by activating PI3K/Akt signal pathway and attenuating changes of cell death-related proteins in FRTL5 thyroid cells.
Asunto(s)
Retículo Endoplásmico/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , Estrés Fisiológico/efectos de los fármacos , Ácido Tióctico/farmacología , Glándula Tiroides/patología , Muerte Celular/efectos de los fármacos , Línea Celular , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Glándula Tiroides/efectos de los fármacos , Glándula Tiroides/metabolismo , Factor de Transcripción CHOP/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
In thyroid cells, the effects of all- TRANS retinoic acid (ATRA) on sodium/iodide symporter (NIS) and CCAAT/enhancer-binding protein-homologous protein (CHOP) under condition of endoplasmic reticulum (ER) stress have not been evaluated. In the present study, the relationships between NIS, CHOP, and p38 MAPK, and the effects of ATRA on NIS and CHOP expression as well as on p38 MAPK activation under condition of ER stress in thyroid cells were investigated. In FRTL5 thyroid cells, NIS mRNA and protein levels decreased following tunicamycin (TN) treatment, while CHOP mRNA and protein levels increased. In addition, while CHOP mRNA levels decreased after administration of tauro-UDCA and siCHOP, NIS mRNA levels were not altered. After pretreatment with SB203580, NIS mRNA levels decreased in non-TN-treated cells but increased in TN-treated cells. In contrast, CHOP mRNA levels decreased in both non-TN-treated and TN-treated cells. Exposure to ATRA decreased NIS mRNA levels in non-TN-treated cells but increased NIS mRNA levels in TN-treated cells. ATRA decreased CHOP mRNA levels in both non-TN-treated and TN-treated cells although the response was significant only in TN-treated cells. Phospho-p38 MAPK protein levels but not total p38 MAPK protein levels increased in TN-treated cells. ATRA attenuated this increase in phopho-p38 MAPK protein levels. In conclusion, our results demonstrate that ER stress may induce reciprocal changes in NIS and CHOP expression via p38 MAPK in FRTL5 thyroid cells, and that ATRA may attenuate ER stress-induced alterations in NIS and CHOP expression by modulating the phosphorylation of p38 MAPK in FRTL5 thyroid cells.
Asunto(s)
Retículo Endoplásmico/metabolismo , Simportadores/metabolismo , Glándula Tiroides/metabolismo , Factor de Transcripción CHOP/metabolismo , Animales , Línea Celular , Retículo Endoplásmico/genética , Ratas , Simportadores/genética , Factor de Transcripción CHOP/genética , Tretinoina , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
BACKGROUND: Left ventricular filling pressure (LVFP) is related to the long-term prognosis in end-stage renal disease. The aims of this study were to evaluate the time course of the changes in LVFP, the predictors for the changes of LVFP, and the plasma N-terminal pro-brain natriuretic peptide (NT-proBNP) levels as indicators for the changes of LVFP in chronic hemodialysis (HD). METHODS: This study was designed prospectively. Doppler echocardiographic examinations and measurement of plasma NT-proBNP levels were performed in 37 consecutive patients on chronic HD and repeated at median of 43 months later. A ratio of peak early transmitral flow velocity to peak early diastolic mitral annular velocity (E/Em), an estimate of LVFP, was calculated. RESULTS: E/Em ratios were significantly increased during the follow-up period. In multivariate analysis, age and changes of LVMI were independently associated with the changes of E/Em ratios. The plasma NT-proBNP levels were independently associated with E/Em at baseline and at the end of follow-up. The changes of plasma NT-proBNP levels were independently associated with changes of E/Em ratios (b-coefficient 0.453, p = 0.003). CONCLUSIONS: Our data suggest that the deterioration of LVFP parallels with the progression of LV hypertrophy. Monitoring the plasma NT-proBNP levels might be useful for the detection of the LVFP changes in chronic HD.
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Hipertrofia Ventricular Izquierda/metabolismo , Hipertrofia Ventricular Izquierda/fisiopatología , Péptido Natriurético Encefálico/metabolismo , Fragmentos de Péptidos/metabolismo , Diálisis Renal , Presión Ventricular/fisiología , Monitoreo Ambulatorio de la Presión Arterial , Progresión de la Enfermedad , Ecocardiografía Doppler , Femenino , Humanos , Hipertrofia Ventricular Izquierda/diagnóstico por imagen , Fallo Renal Crónico/terapia , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Análisis de Regresión , Estadísticas no ParamétricasRESUMEN
We sought to determine the long-term outcomes in type 1 diabetic recipients of intraportal alloislet transplants on a modified immunosuppressive protocol. Six recipients with hypoglycemia unawareness received one to two islet infusions. Induction therapy was with antithymocyte globulin (ATG) plus etanercept for tumor necrosis factor-alpha blockade. Recipients received cyclosporine and everolimus for maintenance immunosuppression for the first year posttransplant, with mycophenolic acid or mycophenolate mofetil subsequently substituted for everolimus. Recipients have been followed for 1173 +/- 270 days since their last infusion for islet graft function (insulin independence, hemoglobin A(1c) levels and C-peptide production) and for adverse events associated with the study protocol. Of the six recipients, five were insulin-independent at 1 year, and four continue to be insulin-independent at a mean of 3.4 +/- 0.4 years posttransplant. None of the six recipients experienced recurrence of severe hypoglycemia. Measured glomerular filtration rate decreased from 110.5 +/- 21.2 mL/min/1.73 m(2) pretransplant to 82.6 +/-19.1 mL/min/1.73 m(2) at 1 year posttransplant. In conclusion, islet transplants restored insulin independence for a mean of >3 years in four of six recipients treated with ATG and etanercept induction therapy and with cyclosporine and, initially, everolimus for maintenance. Our results suggest this immunosuppressive protocol may allow long-term graft survival.
Asunto(s)
Suero Antilinfocítico/uso terapéutico , Diabetes Mellitus Tipo 1/terapia , Inmunoglobulina G/uso terapéutico , Insulina/metabolismo , Trasplante de Islotes Pancreáticos/métodos , Islotes Pancreáticos/citología , Receptores del Factor de Necrosis Tumoral/uso terapéutico , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Adulto , Ciclosporina/uso terapéutico , Etanercept , Everolimus , Femenino , Humanos , Inmunosupresores/uso terapéutico , Masculino , Persona de Mediana Edad , Recurrencia , Sirolimus/análogos & derivados , Sirolimus/uso terapéutico , Resultado del TratamientoRESUMEN
OBJECTIVE: The aims of this study were to clarify changes in total ghrelin within the somatotropic axis in severe burn subjects with or without inhalation injury as well as the responsiveness of GH, IGF-1, and IGFBP-3 to the different severity of burn injuries. DESIGN: Twenty-three patients with severe burn injuries (>30% of 2nd degree burns or >10% of 3rd degree burns) were classified into 2 groups according to inhalation injury: group I with inhalation injury (n=9) and group II without inhalation injury (n=14). The evaluations of serum GH, IGF-1, IGFBP-3, and total ghrelin were done on post-burn injury days 3, 7, 14, 21, and 40. Cortisol levels were measured from 24-h urine collections on post-burn injury days 7 and 21. RESULTS: In all subjects, the levels of GH fluctuated throughout the observation period whereas IGF-1 showed an initial decline with nadir on day 7 and a subsequent increase through day 40. The levels of IGFBP-3 and total ghrelin showed a progressive increase with nadir on day 3. Compared with the group II, the GH levels were increased in the group I on post-burn days 3, 7, and 14, of which day 7 showed statistical significance (p<0.05). The levels of IGF-1 (days 7 and 21; p<0.05) and IGFBP-3 (days 7, 14, 21, and 40; p<0.05, p<0.01, p<0.05, p<0.05, respectively) were lower in the group I than in the group II throughout the study period. On post-burn injury days 3, 7, 14, and 21, total ghrelin levels were lower in the group I than in the group II with statistical significance on post-burn day 7 (p<0.001). CONCLUSIONS: Our present data show a concurrence of elevated GH levels and decreased IGF-I, IGFBP-3, and total ghrelin levels during the early burn injury period, in addition to more GH burst amplitude as well as greater falling of IGF-I, IGFBP-3 and total ghrelin levels proportional to the severity of burn injury. Further studies are needed to ascertain whether acyl- and desacyl-ghrelin instead of total ghrelin are completely independent of increased GH or other stress mediators, and whether GH-releasing hormone (GHRH) mainly stimulates the production and release of GH in acute critical conditions.
Asunto(s)
Quemaduras/sangre , Ghrelina/sangre , Hormona de Crecimiento Humana/sangre , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/sangre , Factor I del Crecimiento Similar a la Insulina/metabolismo , Lesión por Inhalación de Humo/sangre , Adulto , Anciano , Quemaduras/complicaciones , Quemaduras/metabolismo , Femenino , Hormona de Crecimiento Humana/metabolismo , Humanos , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Transducción de Señal/fisiología , Lesión por Inhalación de Humo/complicaciones , Lesión por Inhalación de Humo/metabolismo , Somatotrofos/metabolismo , Factores de TiempoRESUMEN
Diabetes mellitus is postulated to be associated with increased lipid peroxidation, which may contribute to vascular complications. One potential mechanism of the increased lipid peroxidation in diabetes is lipid-linked advanced glycosylation and oxidation. Aminoguanidine (AMGN), the prototype inhibitor of advanced glycosylation end product (AGE) formation, has been recently shown to prevent oxidative modification of low-density lipoprotein (LDL) in vitro at a moderate concentration. It is unknown whether AMGN may act as an antioxidant against lipid peroxidation under hyperglycemia in vivo. To investigate the in vivo effect of AMGN on lipid peroxidation in diabetes, we administered AMGN (1 g/L in drinking water) or vitamin E (400 mg/d for 5 d/wk) to streptozotocin (STZ)-induced diabetic rats for 9 weeks and measured plasma lipid hydroperoxides by ferrous oxidation with xylenol orange II (FOX method) and red blood cell (RBC) membrane malondialdehyde (MDA) and related aldehydes as thiobarbituric acid-reactive substances (TBARS). Plasma lipid hydroperoxide was higher in STZ-induced diabetic rats versus control rats (mean +/- SD, 7.53 +/- 2.03 v 5.62 +/- 0.44 micromol/L, P < .05; n = 8 to 14). RBC membrane TBARS were also higher in STZ-induced diabetic rats than in control rats (2.67 +/- 0.46 v 1.81 +/- 0.19 nmol/mL, P < .05). Plasma lipid hydroperoxide was lower in AMGN-treated (6.23 +/- 0.59 micromol/L, P < .05) and vitamin E-treated (5.29 +/- 0.27 micromol/L, P < .05) diabetic rats than in untreated diabetic rats. RBC membrane TBARS were also lower in AMGN-treated (1.93 +/- 0.12 nmol/mL, P < .05) diabetic rats than in untreated diabetic rats. There was no significant difference in plasma glucose, cholesterol, and triglyceride levels among diabetic groups. Although the mechanism(s) of action of AMGN on lipid peroxidation in vivo should be studied further, these results suggest that AMGN may have an additional beneficial effect as an antioxidant against lipid peroxidation in a prevention trial for diabetic vascular complications.
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Diabetes Mellitus Experimental/metabolismo , Guanidinas/farmacología , Peroxidación de Lípido/efectos de los fármacos , Animales , Antioxidantes/farmacología , Glucemia/análisis , Colesterol/sangre , Eritrocitos/metabolismo , Productos Finales de Glicación Avanzada/metabolismo , Peróxidos Lipídicos/sangre , Masculino , Ratas , Ratas Sprague-Dawley , Sustancias Reactivas al Ácido Tiobarbitúrico/análisis , Triglicéridos/sangre , Vitamina E/farmacologíaRESUMEN
In order to isolate the unidentified autoantigens in autoimmune diabetes, a human pancreatic islet cDNA library was constructed and screened with the sera from the diabetic patients. From the library screening, one clone (DRS-1) that strongly reacted with the sera was isolated. Subsequent sequence analysis revealed that the clone was a novel cDNA related to the diazepam binding inhibitor. DRS-1 was expressed in most tissues including liver, lung, tonsil, and thymus, in addition to pancreatic islets. DRS-1 was in vitro translated and the recombinant DRS-1 protein was expressed in Escherichia coli and purified. The size of the in vitro translated or bacterially expressed DRS-1 protein was in agreement with the conceptually translated polypeptide of DRS-1 cDNA. Further studies are required to test whether or not DRS-1 is a new autoantigen in autoimmune diabetes.
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ADN Complementario/biosíntesis , Isomerasas , Proteínas Recombinantes/genética , Secuencia de Aminoácidos , Autoantígenos/genética , Secuencia de Bases , Isomerasas de Doble Vínculo Carbono-Carbono , Clonación Molecular , ADN Complementario/química , ADN Complementario/aislamiento & purificación , Diabetes Mellitus Tipo 1/genética , Dodecenoil-CoA Isomerasa , Biblioteca de Genes , Humanos , Islotes Pancreáticos/metabolismo , Datos de Secuencia Molecular , ARN/aislamiento & purificación , Proteínas Recombinantes/química , Alineación de SecuenciaRESUMEN
We student basal, glucose- and glucagon-induced insulin secretion in non-insulin diabetes mellitus (NIDDM) patients in relation to body mass index (BMI) and fasting serum glucose (FBS) level. A total of 46 NIDDM patients and 22 control subjects with varying degrees of BMI and FBS were given 100 g of oral glucose and 1 mg of intravenous glucagon on separate days. C-peptide response to glucose, but not basal serum C-peptide and C-peptide response to glucagon, was significantly lower in NIDDM than in controls (P < 0.001). FBS was inversely correlated with C-peptide response to glucose in NIDDM patients (r = -0.67, P < 0.001), but not with basal C-peptide level and C-peptide response to glucagon. On the other hand, BMI was positively correlated with basal serum C-peptide level both in NIDDM (r = 0.60, P < 0.001) and in control subjects (r = 0.74, P < 0.001). In 15 poorly controlled NIDDM patients, the tests were repeated after insulin treatment for 10-14 days. C-peptide response to glucose significantly increase, but not to a level in control subjects, after glycemic control. Basal serum C-peptide level and the C-peptide response to glucagon decreased after glycemic control to significantly lower levels than those in the baseline and those in control subjects. These results suggest that beta cell secretory reserve is reduced in moderate to severe NIDDM patients.
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Glucemia/metabolismo , Péptido C/sangre , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus/sangre , Insulina/metabolismo , Obesidad , Administración Oral , Adulto , Anciano , Glucemia/análisis , Índice de Masa Corporal , Péptido C/efectos de los fármacos , Péptido C/metabolismo , Estudios de Cohortes , Diabetes Mellitus/metabolismo , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Ayuno , Femenino , Glucagón/administración & dosificación , Glucagón/farmacología , Glucosa/administración & dosificación , Glucosa/farmacología , Prueba de Tolerancia a la Glucosa , Humanos , Inyecciones Intravenosas , Inyecciones Subcutáneas , Insulina/administración & dosificación , Insulina/uso terapéutico , Secreción de Insulina , Masculino , Persona de Mediana Edad , Valores de ReferenciaRESUMEN
One hypothesis for the reduction in oxidative killing of neutrophils in diabetic patients is that increased polyol pathway activity during hyperglycemia reduces intracellular levels of nicotinamide adenine dinucleotide phosphate (NADPH), resulting in the reduction of neutrophil superoxide production during the respiratory burst. To test this hypothesis, we assessed the effect of tolrestat, an aldose reductase inhibitor, on neutrophil respiratory burst activity (NRBA) in diabetic patients. We measured fasting plasma glucose (FPG), hemoglobin A1 (HbA1), and NRBA levels in 79 diabetic patients and 48 normal controls. NRBA was reassessed in 34 patients after 4 weeks of tolrestat or placebo treatment, in seven controls after 4 weeks of tolrestat treatment, and in seven patients after 4 weeks of blood glucose control. NRBA was determined by flow cytometry, which detected fluorescent 2',7'-dichlorofluorescein (DCF) in neutrophils formed from 2',7'-dichlorofluorescein diacetate (DCF-DA) during phorbol myristate acetate (PMA)-induced respiratory bursts. Diabetic patients showed lower NRBA than the normal controls (mean cellular fluorescence, 438 +/- 103 v 668 +/- 101, mean +/- SD, P < .001). NRBA in diabetic patients showed a negative correlation with HbA1 (r = -.336, P < .005). Tolrestat treatment for 4 weeks in 17 patients restored the reduced NRBA to an almost normal level (relative NRBA, 0.55 +/- 0.20 v 0.99 +/- 0.36, P < .05) despite the fact that FPG level did not change (11.8 +/- 2.8 v 11.4 +/- 2.8 mmol/L). NRBA of these patients after tolrestat treatment was not significantly different from that of seven control subjects treated with tolrestat for 4 weeks. In 17 placebo-treated patients, there were no significant changes in NRBA and FPG level. The vigorous blood glucose control for 4 weeks in seven patients (16.6 +/- 2.1 v 8.6 +/- 2.3 mmol/L) also restored the reduced NRBA to almost normal (relative NRBA, 0.55 +/- 0.21 v 0.90 +/- 0.30, P < .05). The result that the reduced NRBA in diabetic patients was restored to almost normal either by tolrestat treatment or by blood glucose control strongly supports the hypothesis of this study.
Asunto(s)
Aldehído Reductasa/antagonistas & inhibidores , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 2/sangre , Inhibidores Enzimáticos/farmacología , Naftalenos/farmacología , Neutrófilos/fisiología , Estallido Respiratorio/efectos de los fármacos , Glucemia/análisis , Neuropatías Diabéticas/sangre , Retinopatía Diabética/sangre , Femenino , Hemoglobina Glucada/análisis , Humanos , Masculino , Persona de Mediana Edad , Neutrófilos/efectos de los fármacos , Valores de Referencia , Análisis de Regresión , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Autoantibody to a rat islet cell-protein of 38 kilodalton was detectable at around 30 days of age in the sera of diabetes-prone Biobreeding (DP-BB) rats by both immunoprecipitation and differential Western blotting methods. Anti-38 kilodalton islet cell autoantibody was not, however, observed in the sera from 5- to 20-day-old DP-BB rats. Over 90% of DP-BB rats in which the antibody was detected, eventually developed Type 1 (insulin-dependent) diabetes mellitus. The antibody disappeared within 2 weeks after diabetes onset. However, it was preserved in the sera of DP-BB rats which had been treated with silica to prevent insulitis. The anti-38 kilodalton islet cell autoantibody was not detected in sera from control Wistar Furth (WF) rats. The autoantibody also cross-reacted with a rat insulinoma (RINm5F) cell protein of 38 kilodalton, but did not react with protein from mouse fibroblast (L-929 cells), rat pituitary cells (GH3 cells), or normal rat lymphocytes. The production of the autoantibody appears to be pancreatic Beta-cell dependent, since the autoantibody disappears after almost complete depletion of Beta cells, but is consistently present as long as Beta cells remain. Identification of the Beta-cell dependent anti-38 kilodalton islet cell autoantibody, which cross-reacts with a rat insulinoma cell protein of 38 kilodalton and precedes the onset of Type 1 diabetes in BB rats, will be invaluable for study of the molecular nature of a target islet cell autoantigen associated with the induction of autoimmunity in DP-BB rats.
Asunto(s)
Antígenos/análisis , Autoanticuerpos/análisis , Diabetes Mellitus Experimental/inmunología , Islotes Pancreáticos/inmunología , Estado Prediabético/inmunología , Animales , Antígenos/inmunología , Western Blotting , Línea Celular , Diabetes Mellitus Tipo 1/inmunología , Técnica del Anticuerpo Fluorescente , Insulinoma , Peso Molecular , Neoplasias Pancreáticas , Ratas , Ratas Endogámicas BB , Ratas Endogámicas Lew , Ratas Endogámicas , Ratas Endogámicas WFRESUMEN
The diabetic syndrome in BioBreeding (BB) rats is believed to result from the destruction of beta-cells by autoimmune responses. However, the initial events that cause the autoimmune destruction of beta-cells remain largely unknown. This investigation was initiated to see whether there are any antigenic changes on the beta-cells from neonatal to adult BB rats that may lead to the autoimmune destruction of beta-cells. Pancreatic grafts from neonatal BB rats remained largely intact without insulitis when transplanted into the renal subcapsular space of acutely diabetic BB rats. Similarly transplanted islet grafts from neonatal BB rats were also not subject to autoimmune destruction. In contrast, islet grafts obtained from adult BB rats, which had been treated with silica to prevent insulitis, were rapidly destroyed in diabetic recipients. These results indicate that beta-cells from neonatal BB rats are different from beta-cells from adult BB rats, at least regarding their recognition by immunologic effectors. Considering our observations and previous information on the initial role of macrophages/dendritic cells in the development of insulitis in BB rats, we suggest that beta-cell-specific antigenic changes that precede insulitis may result in the autoimmune destruction of beta-cells in BB rats.
Asunto(s)
Antígenos/inmunología , Autoinmunidad/fisiología , Islotes Pancreáticos/citología , Envejecimiento/inmunología , Envejecimiento/fisiología , Animales , Supervivencia Celular/fisiología , Insulina/metabolismo , Islotes Pancreáticos/inmunología , Islotes Pancreáticos/fisiología , Trasplante de Islotes Pancreáticos/inmunología , Trasplante de Páncreas/inmunología , Enfermedades Pancreáticas/metabolismo , Enfermedades Pancreáticas/prevención & control , Ratas , Ratas Endogámicas BB , Dióxido de Silicio/farmacologíaRESUMEN
In NOD mice. 50-70% of females and 10-20% of males develop diabetes, although almost all the animals show insulitis. To see if environmental insults could induce diabetes in subjects with pre-clinical anti-Beta cell autoimmunity, non-diabetic NOD mice were selected and injected with a sub-diabetogenic dose of streptozotocin at 6 or 20 weeks of age. The streptozotocin failed to induce diabetes in 16 male and 16 female NOD mice within 4 weeks when they were injected at the age of 6 weeks. In contrast, 6 of 16 male and 10 of 16 female NOD mice developed diabetes within 4 weeks when they were injected at the age of 20 weeks. In untreated age-matched control NOD mice, none of the male and only 2 of 16 female mice became diabetic during the same 4 week period. On histologic examination, the degree of insulitis in streptozotocin-treated NOD mice (at the age of 24 weeks) was not significantly different from that of untreated control NOD mice. However, the streptozotocin-treated animals showed significantly lower pancreatic insulin content than the control mice. These results show that an anti-Beta cell autoimmune process in NOD mice has a predisposing effect on the induction of diabetes by a sub-diabetogenic dose of streptozotocin, and suggest that the precipitation of clinical diabetes by some environmental insults in subjects with pre-existing pre-clinical autoimmune Beta-cell destruction may be one mechanism of disease presentation in human Type 1 (insulin-dependent) diabetes.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Diabetes Mellitus Experimental/inmunología , Islotes Pancreáticos/inmunología , Animales , Femenino , Insulina/análisis , Islotes Pancreáticos/química , Masculino , Ratones , Ratones Endogámicos ICR , Ratones Mutantes , Valores de Referencia , EstreptozocinaRESUMEN
Multiple injections of low doses of streptozotocin to susceptible strains of mice produce an experimental autoimmune diabetes mellitus. To investigate the possible initial role of macrophages in the development of insulitis, we studied the effect of macrophage-toxic silica administration on the development of in vitro cellular cytotoxic immune response against pancreatic beta-cells. Multiple streptozotocin-treated mice developed hyperglycemia at day 12 and their splenocytes showed cytotoxicity against cultured rat insulinoma cells. Mice given silica and streptozotocin together remained normoglycemic and their splenocytes showed no cytotoxicity. In contrast, in vitro depletion of macrophages from the splenocytes of mice given multiple streptozotocin alone did not abolish the cytotoxicity. These results show that macrophages themselves contribute little to the cellular cytotoxicity, but are necessary for the development of cytotoxic cells. From these results we suggest that there are at least two different steps in the development of insulitis; the presentation of beta-cell autoantigen by macrophages to helper-T cells, followed by the development of beta-cell-specific cytotoxic cells.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Diabetes Mellitus Experimental/inmunología , Islotes Pancreáticos/inmunología , Macrófagos/inmunología , Animales , Enfermedades Autoinmunes/prevención & control , Línea Celular , Citotoxicidad Inmunológica , Diabetes Mellitus Experimental/prevención & control , Esquema de Medicación , Inmunidad Celular , Macrófagos/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Dióxido de Silicio/farmacología , Estreptozocina/administración & dosificaciónRESUMEN
NOD mice were treated with silica (which is selectively toxic to macrophages) from 4 or 20.5 wk of age. Syngeneic neonatal pancreases were transplanted into the renal subcapsular space of the NOD mice at 21 wk of age. Silica treatment was continued until 24 wk of age, and then the mice were killed for examination of islet morphology. Neither the islets in transplanted pancreases nor the host pancreatic islets from the early long-term silica-treated animals revealed insulitis. In contrast, most of the islets in transplanted pancreases from the late short-term silica-treated animals showed severe insulitis and beta-cell necrosis, as did the host islets. A further experiment was performed to compare the effect of late short-term silica treatment with that of anti-L3T4-antibody treatment of the same time and duration. In contrast to the late short-term silica-treated animals, the transplanted pancreases in the anti-L3T4-antibody-treated animals revealed intact islets, although most of the host islets showed insulitis. The control group, which received no treatment but did receive neonatal pancreases, revealed severe insulitis and beta-cell necrosis of both transplanted and host islets. These results suggest that early macrophage depletion can abolish the development of beta-cell-specific immunologic effectors but that late macrophage depletion, after the development of insulitis, does not affect the destruction of beta-cells by preexisting effectors other than macrophages. We conclude that macrophages are essential for the development of beta-cell-specific cytotoxic effectors in the initial phase of insulitis in NOD mice.