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1.
Cell Reprogram ; 25(4): 171-179, 2023 08.
Artículo en Inglés | MEDLINE | ID: mdl-37590008

RESUMEN

Adipose-derived stem cells (ADSCs) are isolated from abundant adipose tissue and have the capacity to differentiate into multiple cell lineages. ADSCs have raised big interest in therapeutic applications in regenerative medicine and demonstrated to fulfill the criteria for a successful cell therapy. There are several methods for isolation of ADSCs from adipose tissue and cryopreservation of ADSCs. Here, novel methods for the isolation and cryopreservation of ADSCs are presented and focused. Microscopic pieces of adipose tissue were placed on transwell inserts, and the ADSCs were induced to migrate to the lower wells for 1 week. We compared the properties of our ADSCs with those isolated by enzymatic digestion and enzyme-free method of culture plate, and our ADSCs were found to be more stable and healthier. In addition, we proposed a novel cryoprotectant solution (FNCP) containing pectin and L-alanine, which was compared with standard cryoprotectant solution. Overall, our methods proved more useful for ADSCs isolation than other methods and did not require consideration of "minimal manipulation" by the U.S. Food and Drug Administration (FDA). Furthermore, our FNCP did not contain dimethyl sulfoxide and fetal bovine serum, therefore stable storage is possible in xeno-free and animal-free cryopreservation solutions.


Asunto(s)
Adipocitos , Tejido Adiposo , Estados Unidos , Humanos , Linaje de la Célula , Criopreservación , Separación Celular
2.
Cancers (Basel) ; 12(5)2020 May 11.
Artículo en Inglés | MEDLINE | ID: mdl-32403237

RESUMEN

BACKGROUNDS: Radioimmunotherapy (RIT) serves as a targeted therapy for non-Hodgkin lymphomas (NHL). Although HIF(Hypoxia-inducible factors)-1α is an important biomarker during radiation therapy, its role in NHL is unclear. Atorvastatin (ATV) is used as a combination drug for chemotherapy. METHODS: We investigated whether ATV downregulated tumor radio-resistance and enhanced the anticancer effect of 131I-RTX (rituximab) in Raji xenograft mouse models. First, the increased uptake and enhanced therapeutic effect of 131I-RTX by ATV was confirmed using molecular imaging in Raji xenograft subcutaneous model and orthotropic model with SPECT and IVIS images. Second, we examined the profile of differentially expressed miRNAs using miRNA array. RESULTS: We found that miR-346 inhibited HIF-1α/VEGF (Vascular endothelial growth factor) during ATV combination therapy with 131I-RTX. The underlying mechanism of ATV involved induction of anti-angiogenesis and radiosensitivity by downregulating HIF-1α in Raji cells. CONCLUSION: Our findings suggested that combination therapy with ATV and 131I-RTX is a promising strategy for enhancing the potency of 131I-RTX therapy in poorly responding patients and those with radio-resistance.

3.
J Clin Med ; 8(2)2019 Feb 11.
Artículo en Inglés | MEDLINE | ID: mdl-30754707

RESUMEN

The authors identified that chemo-brain was induced after trastuzumab (TZB) therapy. In addition, atorvastatin (ATV) could rescue chemo-brain during trastuzumab (TZB) therapy. Enhanced therapeutic effect of TZB was confirmed after ATV therapy. We also investigated that there was no hair loss side effect due to ATV therapy. In an animal model, 150 µg TZB and five serial doses of 20 mg/kg ATV were administered. 18F-fluorodeoxyglucose Positron Emission Tomography (PET) and Magnetic Resonance Imaging (MRI) data were acquired. Statistical parametric mapping analysis and voxel-based morphometry analysis were performed to identify differences in glucose metabolism and gray matter concentration. The enhanced therapeutic efficacy of TZB after ATV treatment was assessed using a human epidermal growth factor receptor 2-positive gastric cancer model. We found a decrease in cerebral glucose metabolism and gray matter concentration in the frontal lobe following TZB therapy (p < 0.005). After subsequent ATV administration, glucose metabolism and regional gray matter concentration were rescued (p < 0.005). Cognitive impairment due to TZB and the rescue effect of ATV were confirmed using a passive avoidance test and quantitative real-time reverse transcription PCR. Furthermore, the penetration and accumulation of TZB in tumors increased by 100% after ATV co-administration, which resulted in an enhanced anti-cancer effect. Our study collectively demonstrates that ATV co-administration with TZB rescued the TZB-induced chemo-brain and enhances the therapeutic efficacy of TZB in tumors. We also showed that there was no hair loss during ATV therapy.

4.
BMC Cancer ; 19(1): 149, 2019 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-30760223

RESUMEN

BACKGROUND: Chemotherapy-induced alopecia has been well documented as a cause of distress to patients undergoing cancer treatment. Almost all traditional chemotherapeutic agents cause severe alopecia. Despite advances in the treatment of chemotherapy-induced alopecia, there is no effective treatment for preventing chemotherapy-induced alopecia. METHODS: In the present study, we investigated the potential role of a multi-target iron chelator, M30 in protecting against cyclophosphamide-induced alopecia in C57BL/6 mice implanted with an osmotic pump. M30 enhanced hair growth and prevented cyclophosphamide-induced abnormal hair in the mice. Furthermore, we examined the gene expression profiles derived from skin biopsy specimens of normal mice, cyclophosphamide-treated mice, and cyclophosphamide treated mice with M30 supplement. RESULTS: The top genes namely Tnfrsf19, Ercc2, Lama5, Ctsl, and Per1 were identified by microarray analysis. These genes were found to be involved in the biological processes of hair cycle, hair cycle phase, hair cycle process, hair follicle development, hair follicle maturation, hair follicle morphogenesis, regulation of hair cycle. CONCLUSION: Our study demonstrates that M30 treatment is a promising therapy for cyclophosphamide-induced alopecia and suggests that the top five genes have unique preventive effects in cyclophosphamide-induced transformation.


Asunto(s)
Alopecia/genética , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Antioxidantes/uso terapéutico , Ciclofosfamida/efectos adversos , Hidroxiquinolinas/uso terapéutico , Quimioterapia de Inducción/efectos adversos , Neoplasias/tratamiento farmacológico , Alopecia/inducido químicamente , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Ciclofosfamida/uso terapéutico , Modelos Animales de Enfermedad , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Humanos , Ratones , Ratones Endogámicos C57BL , Análisis por Micromatrices , Receptores del Factor de Necrosis Tumoral/genética
5.
Biomed Mater ; 10(3): 035014, 2015 Jun 24.
Artículo en Inglés | MEDLINE | ID: mdl-26107298

RESUMEN

The extracellular matrix (ECM) surrounding cells contains a variety of proteins that provide structural support and regulate cellular functions. Previous studies have shown that decellularized ECM isolated from tissues or cultured cells can be used to improve cell differentiation in tissue engineering applications. In this study we evaluated the effect of decellularized chondrocyte-derived ECM (CDECM) on the chondrogenesis of human placenta-derived mesenchymal stem cells (hPDMSCs) in a pellet culture system. After incubation with or without chondrocyte-derived ECM in chondrogenic medium for 1 or 3 weeks, the sizes and wet masses of the cell pellets were compared with untreated controls (hPDMSCs incubated in chondrogenic medium without chondrocyte-derived ECM). In addition, histologic analysis of the cell pellets (Safranin O and collagen type II staining) and quantitative reverse transcription-PCR analysis of chondrogenic markers (aggrecan, collagen type II, and SOX9) were carried out. Our results showed that the sizes and masses of hPDMSC pellets incubated with chondrocyte-derived ECM were significantly higher than those of untreated controls. Differentiation of hPDMSCs (both with and without chondrocyte-derived ECM) was confirmed by Safranin O and collagen type II staining. Chondrogenic marker expression and glycosaminoglycan (GAG) levels were significantly higher in hPDMSC pellets incubated with chondrocyte-derived ECM compared with untreated controls, especially in cells precultured with chondrocyte-derived ECM for 7 d. Taken together, these results demonstrate that chondrocyte-derived ECM enhances the chondrogenesis of hPDMSCs, and this effect is further increased by preculture with chondrocyte-derived ECM. This preculture method for hPDMSC chondrogenesis represents a promising approach for cartilage tissue engineering.


Asunto(s)
Condrogénesis/fisiología , Matriz Extracelular/fisiología , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Agrecanos/genética , Diferenciación Celular , Células Cultivadas , Condrocitos/fisiología , Condrogénesis/genética , Colágeno Tipo II/genética , Femenino , Glicosaminoglicanos/metabolismo , Humanos , Ensayo de Materiales , Placenta/citología , Embarazo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Transcripción SOX9/genética , Ingeniería de Tejidos , Andamios del Tejido
6.
J Invest Dermatol ; 135(1): 269-278, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25007043

RESUMEN

Diabetes mellitus disrupts wound repair and leads to the development of chronic wounds, likely due to impaired angiogenesis. We previously demonstrated that human proinsulin C-peptide can protect against vasculopathy in diabetes; however, its role in impaired wound healing in diabetes has not been studied. We investigated the potential roles of C-peptide in protecting against impaired wound healing by inducing angiogenesis using streptozotocin-induced diabetic mice and human umbilical vein endothelial cells. Diabetes delayed wound healing in mouse skin, and C-peptide supplement using osmotic pumps significantly increased the rate of skin wound closure in diabetic mice. Furthermore, C-peptide induced endothelial cell migration and tube formation in dose-dependent manners, with maximal effect at 0.5 nM. These effects were mediated through activation of extracellular signal-regulated kinase 1/2 and Akt, as well as nitric oxide formation. C-peptide-enhanced angiogenesis in vivo was demonstrated by immunohistochemistry and Matrigel plug assays. Our findings highlight an angiogenic role of C-peptide and its ability to protect against impaired wound healing, which may have significant implications in reparative and therapeutic angiogenesis in diabetes. Thus, C-peptide replacement is a promising therapy for impaired angiogenesis and delayed wound healing in diabetes.


Asunto(s)
Péptido C/metabolismo , Diabetes Mellitus Experimental , Neovascularización Fisiológica/fisiología , Cicatrización de Heridas/fisiología , Animales , Movimiento Celular/fisiología , Proliferación Celular/fisiología , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Experimental/fisiopatología , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patología , Diabetes Mellitus Tipo 1/fisiopatología , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patología , Diabetes Mellitus Tipo 2/fisiopatología , Células Endoteliales/metabolismo , Células Endoteliales/patología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Ratones Endogámicos C57BL , Úlcera Cutánea/metabolismo , Úlcera Cutánea/patología , Úlcera Cutánea/fisiopatología
7.
Cardiovasc Res ; 104(2): 234-44, 2014 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-25239825

RESUMEN

Lack of C-peptide, along with insulin, is the main feature of Type 1 diabetes mellitus (DM) and is also observed in progressive ß-cell loss in later stage of Type 2 DM. Therapeutic approaches to hyperglycaemic control have been ineffective in preventing diabetic vasculopathy, and alternative therapeutic strategies are necessary to target both hyperglycaemia and diabetic complications. End-stage organ failure in DM seems to develop primarily due to vascular dysfunction and damage, leading to two types of organ-specific diseases, such as micro- and macrovascular complications. Numerous studies in diabetic patients and animals demonstrate that C-peptide treatment alone or in combination with insulin has physiological functions and might be beneficial in preventing diabetic complications. Current evidence suggests that C-peptide replacement therapy might prevent and ameliorate diabetic vasculopathy and organ-specific complications through conservation of vascular function, as well as prevention of endothelial cell death, microvascular permeability, vascular inflammation, and neointima formation. In this review, we describe recent advances on the beneficial role of C-peptide replacement therapy for preventing diabetic complications, such as retinopathy, nephropathy, neuropathy, impaired wound healing, and inflammation, and further discuss potential beneficial effects of combined C-peptide and insulin supplement therapy to control hyperglycaemia and to prevent organ-specific complications.


Asunto(s)
Péptido C/uso terapéutico , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Angiopatías Diabéticas/prevención & control , Hipoglucemiantes/uso terapéutico , Animales , Péptido C/sangre , Péptido C/deficiencia , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 1/complicaciones , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/diagnóstico , Angiopatías Diabéticas/sangre , Angiopatías Diabéticas/etiología , Humanos , Transducción de Señal/efectos de los fármacos , Resultado del Tratamiento
8.
ScientificWorldJournal ; 2014: 539297, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25133241

RESUMEN

In the conventional DC-AC inverter consisting of two DC-DC converters with unipolar output capacitors, the output capacitor voltages of the DC-DC converters must be higher than the DC input voltage. To overcome this weakness, this paper proposes a single-phase DC-AC inverter consisting of two embedded Z-source converters with bipolar output capacitors. The proposed inverter is composed of two embedded Z-source converters with a common DC source and output AC load. Though the output capacitor voltages of the converters are relatively low compared to those of a conventional inverter, an equivalent level of AC output voltages can be obtained. Moreover, by controlling the output capacitor voltages asymmetrically, the AC output voltage of the proposed inverter can be higher than the DC input voltage. To verify the validity of the proposed inverter, experiments were performed with a DC source voltage of 38 V. By controlling the output capacitor voltages of the converters symmetrically or asymmetrically, the proposed inverter can produce sinusoidal AC output voltages. The experiments show that efficiencies of up to 95% and 97% can be achieved with the proposed inverter using symmetric and asymmetric control, respectively.


Asunto(s)
Electrónica/instrumentación , Electrónica/métodos
9.
Cardiovasc Res ; 101(1): 155-64, 2014 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-24142430

RESUMEN

AIMS: Human C-peptide has a beneficial effect on the prevention of diabetic neuropathy, nephropathy, and vascular complications; however, its role in protection against increased vascular permeability in diabetes remains unclear. Our purpose was to explore the potential protective role of C-peptide against microvascular permeability mediated by vascular endothelial growth factor (VEGF)-induced reactive oxygen species (ROS) generation in diabetes. METHODS AND RESULTS: Generation of intracellular ROS, real-time changes in intracellular Ca(2+), ROS-dependent stress fibre formation, and the disassembly of the adherens junctions were studied by a confocal microscopy in human umbilical vein endothelial cells (HUVECs). VEGF-induced vascular leakage was investigated in the skin of diabetic mice using a Miles vascular permeability assay. Microvascular leakage in the retina of streptozotocin diabetic mice was investigated using a confocal microscopy after left ventricle injection of fluorescein isothiocyanate (FITC)-dextran. C-peptide inhibited the VEGF-induced ROS generation, stress fibre formation, disassembly of vascular endothelial cadherin, and endothelial permeability in HUVECs. Intradermal injection of C-peptide prevented VEGF-induced vascular leakage. Consistent with this, intravitreal injection of C-peptide prevented the extravasation of FITC-dextran in the retinas of diabetic mice, which was also prevented by anti-VEGF antibody and ROS scavengers in diabetic mice. Conclusions/interpretation C-peptide prevents VEGF-induced microvascular permeability by inhibiting ROS-mediated intracellular events in diabetic mice, suggesting that C-peptide replacement is a promising therapeutic strategy to prevent diabetic retinopathy.


Asunto(s)
Péptido C/fisiología , Permeabilidad Capilar , Angiopatías Diabéticas/etiología , Uniones Adherentes/metabolismo , Animales , Cadherinas/metabolismo , Calcio/metabolismo , Células Cultivadas , Angiopatías Diabéticas/metabolismo , Células Endoteliales/metabolismo , Endotelio Vascular/metabolismo , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Especies Reactivas de Oxígeno/metabolismo , Retina/metabolismo , Fibras de Estrés/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo
10.
Diabetes ; 62(11): 3851-62, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-23884890

RESUMEN

Vasculopathy is a major complication of diabetes; however, molecular mechanisms mediating the development of vasculopathy and potential strategies for prevention have not been identified. We have previously reported that C-peptide prevents diabetic vasculopathy by inhibiting reactive oxygen species (ROS)-mediated endothelial apoptosis. To gain further insight into ROS-dependent mechanism of diabetic vasculopathy and its prevention, we studied high glucose-induced cytosolic and mitochondrial ROS production and its effect on altered mitochondrial dynamics and apoptosis. For the therapeutic strategy, we investigated the vasoprotective mechanism of C-peptide against hyperglycemia-induced endothelial damage through the AMP-activated protein kinase α (AMPKα) pathway using human umbilical vein endothelial cells and aorta of diabetic mice. High glucose (33 mmol/L) increased intracellular ROS through a mechanism involving interregulation between cytosolic and mitochondrial ROS generation. C-peptide (1 nmol/L) activation of AMPKα inhibited high glucose-induced ROS generation, mitochondrial fission, mitochondrial membrane potential collapse, and endothelial cell apoptosis. Additionally, the AMPK activator 5-aminoimidazole-4-carboxamide 1-ß-d-ribofuranoside and the antihyperglycemic drug metformin mimicked protective effects of C-peptide. C-peptide replacement therapy normalized hyperglycemia-induced AMPKα dephosphorylation, ROS generation, and mitochondrial disorganization in aorta of diabetic mice. These findings highlight a novel mechanism by which C-peptide activates AMPKα and protects against hyperglycemia-induced vasculopathy.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Apoptosis/efectos de los fármacos , Péptido C/uso terapéutico , Angiopatías Diabéticas/etiología , Dinámicas Mitocondriales/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Aminoimidazol Carboxamida/farmacología , Animales , Péptido C/farmacología , Células Cultivadas , Citosol/metabolismo , Diabetes Mellitus Experimental/tratamiento farmacológico , Activación Enzimática , Glucosa/administración & dosificación , Glucosa/farmacología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Metformina/farmacología , Ratones , Ratones Endogámicos C57BL , Especies Reactivas de Oxígeno/farmacología
11.
Diabetes ; 62(1): 243-53, 2013 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-22923476

RESUMEN

C-peptide is a bioactive peptide with a potentially protective role in diabetes complications; however, its molecular mechanism of protection against cardiovascular damage caused by hyperglycemia-induced apoptosis remains unclear. We investigated the protective mechanism of C-peptide against hyperglycemia-induced apoptosis using human umbilical vein endothelial cells and streptozotocin diabetic mice. High glucose (33 mmol/L) induced apoptotic cell death in endothelial cells via sequential elevation of intracellular Ca(2+) and reactive oxygen species (ROS) as well as subsequent activation of transglutaminase 2 (TG2). C-peptide (1 nmol/L) prevented endothelial cell death by inhibiting protein kinase C- and NADPH oxidase-dependent intracellular ROS generation and by abolishing high glucose-induced TG2 activation, without affecting intracellular Ca(2+) levels. Consistently, in the aorta of streptozotocin diabetic mice, hyperglycemia stimulated transamidating activity and endothelial cell apoptosis that was inhibited by C-peptide replacement therapy (35 pmol/min/kg) using osmotic pumps (control and diabetes, n = 8; diabetes + C-peptide, n = 7). In addition, C-peptide prevented hyperglycemia-induced activation of transamidation activity and apoptosis in the heart and renal cortex of streptozotocin diabetic mice. Thus, C-peptide protects endothelial cells from hyperglycemia-induced apoptotic cell death by inhibiting intracellular ROS-mediated activation of TG2. Furthermore, TG2 may be a promising avenue of therapeutic investigation to treat diabetic vasculopathies.


Asunto(s)
Apoptosis , Péptido C/fisiología , Proteínas de Unión al GTP/fisiología , Células Endoteliales de la Vena Umbilical Humana/patología , Hiperglucemia/patología , Especies Reactivas de Oxígeno/metabolismo , Transglutaminasas/fisiología , Animales , Calcio/metabolismo , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Activación Enzimática , Proteínas de Unión al GTP/antagonistas & inhibidores , Humanos , Corteza Renal/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Miocardio/patología , Proteína Glutamina Gamma Glutamiltransferasa 2 , Estreptozocina , Transglutaminasas/antagonistas & inhibidores
12.
J Biol Chem ; 287(18): 14377-88, 2012 Apr 27.
Artículo en Inglés | MEDLINE | ID: mdl-22418443

RESUMEN

Transglutaminase 2 (TG2) is a versatile protein that is implicated in significant biological processes, including cell death and degenerative diseases. A possible role of TG2 in the apoptotic death of cancer cells induced by photodynamic therapy (PDT) was suggested recently; however, the mechanism by which TG2 regulates apoptotic responses to PDT remains to be elucidated. In this study, we investigated the key signaling pathways stimulated during apoptotic cell death following PDT and whether inhibition of TG2 activation using pharmacological approaches and siRNAs affects the signaling pathways. PDT caused the release of both cytochrome c and apoptosis-inducing factor (AIF) by damaging mitochondria, which resulted in caspase-dependent and caspase-independent apoptotic cell death, respectively. Released AIF translocated to the nucleus and, synergistically with the caspase-dependent pathway, led to apoptotic cell death. Both the caspase cascade and the activation of AIF following PDT were mediated by TG2 activation. In addition, PDT-activated calpain was responsible for the sequential events of Bax translocation, the collapse of ΔΨ(m), caspase-3 activation, and AIF translocation, all of which were provoked by TG2 activation. Together, these results demonstrate that PDT with a chlorin-based photosensitizer targets TG2 by activating calpain-induced Bax translocation, which induces apoptotic cell death through both caspase-dependent and AIF-mediated pathways. Moreover, these results indicate that TG2 may be a possible therapeutic target for PDT treatment of cancer.


Asunto(s)
Apoptosis/efectos de los fármacos , Calpaína/metabolismo , Caspasa 3/metabolismo , Neoplasias/tratamiento farmacológico , Fotoquimioterapia , Transducción de Señal/efectos de los fármacos , Transglutaminasas/metabolismo , Factor Inductor de la Apoptosis/metabolismo , Calpaína/genética , Caspasa 3/genética , Línea Celular Tumoral , Activación Enzimática/efectos de los fármacos , Proteínas de Unión al GTP , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Proteína Glutamina Gamma Glutamiltransferasa 2 , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/genética , Transducción de Señal/genética , Transglutaminasas/genética , Proteína X Asociada a bcl-2
13.
J Cell Biochem ; 112(10): 3061-71, 2011 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-21678478

RESUMEN

Here, we present differential cytotoxic responses to two different doses of photodynamic therapies (PDTs; low-dose PDT [LDP] and high-dose PDT [HDP]) using a chlorin-based photosensitizer, DH-II-24, in human gastric and bladder cancer cells. Fluorescence-activated cell sorting analysis using Annexin V and propidium iodide (PI) showed that LDP induced apoptotic cell death, whereas HDP predominantly caused necrotic cell death. The differential cytotoxic responses to the two PDTs were further confirmed by a DiOC(6) and PI double-staining assay via confocal microscopy. LDP, but not HDP, activated caspase-3, which was inhibited by Z-VAD, Trolox, and BAPTA-AM. LDP and HDP demonstrated opposite effects on intracellular reactive oxygen species (ROS)/Ca(2+) signals; LDP stimulated intracellular ROS production, contributing to a transient increase of intracellular Ca(2+) , whereas HDP induced a massive and prolonged elevation of intracellular Ca(2+) responsible for the transient production of intracellular ROS. In addition, the two PDTs also increased in situ transglutaminase 2 (TG2) activity, with a higher stimulation by HDP, and this increase in activity was prevented by Trolox, BAPTA-AM, and TG2-siRNA. LDP-induced apoptotic cell death was strongly inhibited by Trolox and TG2-siRNA and moderately suppressed by BAPTA-AM. However, HDP-mediated necrotic cell death was partially inhibited by BAPTA-AM but not by TG2-siRNA. Thus, these results demonstrate that LDP and HDP induced apoptotic and necrotic cell death by differential signaling mechanisms involving intracellular Ca(2+) , ROS, and TG2.


Asunto(s)
Fotoquimioterapia/métodos , Neoplasias Gástricas/metabolismo , Neoplasias de la Vejiga Urinaria/metabolismo , Apoptosis/efectos de los fármacos , Calcio/metabolismo , Caspasa 3/metabolismo , Línea Celular Tumoral , Proteínas de Unión al GTP/genética , Proteínas de Unión al GTP/metabolismo , Humanos , Necrosis/inducido químicamente , Porfirinas/farmacología , Proteína Glutamina Gamma Glutamiltransferasa 2 , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Transglutaminasas/genética , Transglutaminasas/metabolismo
14.
Cancer Sci ; 102(3): 549-56, 2011 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-21205075

RESUMEN

We investigated cellular responses to chlorin-based photosensitizer DH-II-24 under darkness in human gastric adenocarcinoma AGS cells. Cells were loaded with 0.5-10 µg/mL DH-II-24 for 12 h, and intracellular reactive oxygen species (ROS) and intracellular Ca(2+) levels, in situ tissue transglutaminase (tTGase) activity, cell viability, cell morphology and cell cycle were examined. DH-II-24 treatment had no effect on intracellular ROS production or cell morphology, and did not induce cell detachment at any concentrations tested. In addition, cell viability and cell cycle progression were not altered by the photosensitizer. However, DH-II-24 treatment elevated the basal level of intracellular Ca(2+) in a dose-dependent manner and inhibited tTGase activity without affecting tTGase expression levels. Furthermore, DH-II-24 inhibited lysophosphatidic acid-induced activation of tTGase in a dose-dependent manner. In contrast, photodynamic therapy (PDT) with 1 µg/mL DH-II-24 significantly elevated intracellular ROS and in situ tTGase activity in parallel with a rapid and large increase in intracellular Ca(2+) levels. DH-II-24-mediated PDT decreased cell viability and induced cell detachment. These results demonstrate that DH-II-24 treatment alone under darkness induced different cellular responses to DH-II-24-mediated PDT.


Asunto(s)
Adenocarcinoma/tratamiento farmacológico , Porfirinas/farmacología , Neoplasias Gástricas/tratamiento farmacológico , Adenocarcinoma/patología , Calcio/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Humanos , Fotoquimioterapia , Especies Reactivas de Oxígeno/metabolismo , Neoplasias Gástricas/patología , Transglutaminasas/metabolismo
15.
J Korean Med Sci ; 25(8): 1222-7, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20676337

RESUMEN

This study examined whether propofol and aminophylline affect the mobilization of intracellular calcium in human umbilical vein endothelial cells. Intracellular calcium was measured using laser scanning confocal microscopy. Cultured and serum-starved cells on round coverslips were incubated with propofol or aminophylline for 30 min, and then stimulated with lysophosphatidic acid, propofol and aminophylline. The results were expressed as relative fluorescence intensity and fold stimulation. Propofol decreased the concentration of intracellular calcium, whereas aminophylline caused increased mobilization of intracellular calcium in a concentration-dependent manner. Propofol suppressed the lysophosphatidic acid-induced mobilization of intracellular calcium in a concentration-dependent manner. Propofol further prevented the aminophylline-induced increase of intracellular calcium at clinically relevant concentrations. However, aminophylline reversed the inhibitory effect of propofol on the elevation of intracellular calcium by lysophosphatidic acid. Our results suggest that propofol and aminophylline antagonize each other on the mobilization of intracellular calcium in human umbilical vein endothelial cells at clinically relevant concentrations. Serious consideration should be given to how this interaction affects mobilization of intracellular calcium when these two drugs are used together.


Asunto(s)
Aminofilina/antagonistas & inhibidores , Anestésicos Intravenosos/antagonistas & inhibidores , Broncodilatadores/antagonistas & inhibidores , Calcio/metabolismo , Células Endoteliales/efectos de los fármacos , Propofol/antagonistas & inhibidores , Aminofilina/farmacología , Anestésicos Intravenosos/farmacología , Broncodilatadores/farmacología , Células Cultivadas , Células Endoteliales/metabolismo , Endotelio Vascular/citología , Humanos , Lisofosfolípidos/farmacología , Microscopía Confocal , Propofol/farmacología , Venas Umbilicales/citología
16.
Cancer Sci ; 100(12): 2431-6, 2009 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-19751236

RESUMEN

While photodynamic therapy (PDT) has been recognized as a promising therapeutic modality for the treatment of various cancers and diseases, developments of effective photosensitizers are highly desired to improve the prospect for the use of PDT. In this study, we evaluated DH-II-24, a new photosensitizer, for antitumor PDT in vitro and in vivo. Loaded into human colorectal carcinoma cells (HCT116), DH-II-24 was primarily accumulated in mitochondria, lysosomes, and endoplasmic reticula. Administration of DH-II-24 followed by light exposure induced necrotic cell death in a dose-dependent manner, whereas DH-II-24 in the absence of light induced minimal cell death. In order to investigate the distribution and phamacokinetics of the photosensitizer in vivo, DH-II-24 was intravenously injected to female BALB/c nude mice. Fluorescence imaging in vivo showed that DH-II-24 was rapidly distributed across the entire body and then mostly eliminated at 24 h. Next, effectiveness of DH-II-24-mediated PDT was examined on colorectal carcinoma xenografts established subcutaneously in BALB/c nude mice. DH-II-24 (1 mg/kg, i.v. administration) followed by light exposure significantly suppressed growth of xenograft tumors, compared to light exposure or DH-II-24 alone. Histological examination revealed necrotic damage in PDT-treated tumors, concomitantly with severe damage of tumor vasculature. These results suggest that DH-II-24 is a potential photosensitizer of photodynamic therapy for cancer.


Asunto(s)
Neoplasias Colorrectales/tratamiento farmacológico , Fotoquimioterapia , Fármacos Fotosensibilizantes/uso terapéutico , Porfirinas/uso terapéutico , Animales , Neoplasias Colorrectales/patología , Femenino , Células HCT116 , Humanos , Ratones , Ratones Endogámicos BALB C , Ensayos Antitumor por Modelo de Xenoinjerto
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