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1.
Pancreatology ; 22(7): 1028-1034, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-35948507

RESUMO

BACKGROUND AND AIMS: Pancreatic resection is associated with pancreatic exocrine insufficiency (PEI) leading to nutritional consequences. The Pancreatic Nutrition Clinic was established to diagnose and manage PEI through standardised nutritional assessment. In this prospective observational study, we aimed to define the rate of PEI, diabetes mellitus and nutritional abnormalities in patients who underwent pancreatic resection. METHODS: All Pancreatic Nutrition Clinic patients were included for analysis. Clinical data were prospectively obtained at initial assessment. Biochemical data included micronutrient levels, faecal elastase-1 and haemoglobin A1c. Bone mineral density and nutritional assessment were undertaken. RESULTS: Ninety-eight patients were included. Fifty-nine per cent (58/98) had undergone a pancreatoduodenectomy. Ninety-three patients had a faecal elastase-1 result, 65% (60/93) of which had a faecal elastase-1 less than 200 µg/g of faeces. Seventy-five patients (76%) of the total population required PERT, and thirty-nine (40%) were classified as malnourished using the patient-generated subjective global assessment tool. Seventy-two per cent (70/97) had a biochemical deficiency of one or more micronutrients. Thirty-eight people (39%) had diabetes mellitus. Of the seventy-eight patients with a bone mineral density scan available for analysis, 29% (23/78) had osteoporosis and 49% (38/78) osteopenia. CONCLUSIONS: Pancreatic exocrine insufficiency, micronutrient deficiency, bone disease, diabetes mellitus and malnutrition are highly prevalent in patients who have undergone pancreatic resection.


Assuntos
Diabetes Mellitus , Insuficiência Pancreática Exócrina , Desnutrição , Doenças Metabólicas , Humanos , Insuficiência Pancreática Exócrina/diagnóstico , Elastase Pancreática/análise , Micronutrientes
3.
Biochim Biophys Acta Mol Basis Dis ; 1866(12): 165970, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32950675

RESUMO

Di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) and its analogues are potent anti-cancer agents through their ability to target lysosomes. Considering this, it was important to understand the mechanisms involved in the Dp44mT-mediated induction of autophagy and the role of 5'-adenosine monophosphate-activated protein kinase (AMPK) as a critical autophagic regulator. As such, this investigation examined AMPK's role in the regulation of the transcription factor EB (TFEB), which transcribes genes involved in autophagy and lysosome biosynthesis. For the first time, this study demonstrated that Dp44mT induces translocation of TFEB to the nucleus. Furthermore, Dp44mT-mediated nuclear translocation of TFEB was AMPK-dependent. Considering that: (1) the mammalian target of rapamycin complex 1 (mTORC1) plays an important role in the regulation of TFEB; and (2) that AMPK is a known regulator of mTORC1, this study also elucidated the mechanisms through which Dp44mT regulates nuclear translocation of TFEB via AMPK. Silencing AMPK led to increased mTOR phosphorylation, that activates mTORC1. Since Dp44mT inhibits mTORC1 in an AMPK-dependent manner through raptor phosphorylation, Dp44mT is demonstrated to regulate TFEB translocation through dual mechanisms: AMPK activation, which inhibits mTOR, and inhibition of mTORC1 via phosphorylation of raptor. Collectively, Dp44mT-mediated activation of AMPK plays a crucial role in lysosomal biogenesis and TFEB function. As Dp44mT potently chelates copper and iron that are crucial for tumor growth, these studies provide insight into the regulatory mechanisms involved in intracellular clearance and energy metabolism that occur upon alterations in metal ion homeostasis.


Assuntos
Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Antineoplásicos/farmacologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Núcleo Celular/efeitos dos fármacos , Alvo Mecanístico do Complexo 1 de Rapamicina/antagonistas & inibidores , Tiossemicarbazonas/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Antineoplásicos/química , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Estrutura Molecular , Fosforilação/efeitos dos fármacos , Relação Estrutura-Atividade , Tiossemicarbazonas/química , Células Tumorais Cultivadas
4.
Biochim Biophys Acta Mol Basis Dis ; 1866(5): 165657, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31904416

RESUMO

Upon activation, the 5'-adenosine monophosphate-activated protein kinase (AMPK) increases catabolism, while inhibiting anabolism. The anti-cancer agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), activates AMPK in multiple tumor cell-types (Biochim. Biophys Acta 2016;1863:2916-2933). This acts as an initial cell "rescue response" after iron-depletion mediated by Dp44mT. Considering Dp44mT-mediated AMPK activation, the role of AMPK on Dp44mT cytotoxicity was examined. Dp44mT increased the p-AMPK/AMPK ratio in multiple tumor cell-types over short (24 h) and longer (72 h) incubations. Notably, Dp44mT was more effective in inhibiting tumor cell proliferation after AMPK silencing, potentially due to the loss of AMPK-mediated metabolic plasticity that protects cells against Dp44mT cytotoxicity. The silencing of AMPK-increased cellular cholesterol and stabilized lysosomes against Dp44mT-mediated lysosomal membrane permeabilization. This was substantiated by studies demonstrating that the cholesterol-depleting agent, methyl-ß-cyclodextrin (MßCD), restores Dp44mT-mediated lysosomal membrane permeabilization in AMPK silenced cells. The increased levels of cholesterol after AMPK silencing were independent of the ability of AMPK to inhibit the rate-limiting step of cholesterol synthesis via the inactivating phosphorylation of 3-hydroxy-3-methylglutaryl CoA reductase (HMGCR) at Ser872. In fact, Dp44mT did not increase phosphorylation of HMGCR at (Ser872), but decreased total HMGCR expression similarly in both the presence or absence of AMPK silencing. Dp44mT was demonstrated to increase autophagic initiation after AMPK silencing via an AMPK- and Beclin-1-independent mechanism. Further, there was increased cleaved caspase 3 and cleaved PARP after incubation of AMPK silenced cells with Dp44mT. Overall, AMPK silencing promotes Dp44mT anti-proliferative activity, suggesting a role for AMPK in rescuing its cytotoxicity by inhibiting autophagy and also apoptosis.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Antineoplásicos/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Tiossemicarbazonas/farmacologia , Proteínas Quinases Ativadas por AMP/genética , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/efeitos dos fármacos , Autofagia/genética , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/patologia , Fosforilação/efeitos dos fármacos , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Tiossemicarbazonas/uso terapêutico
5.
Biochim Biophys Acta Mol Basis Dis ; 1865(9): 2094-2110, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-30981813

RESUMO

The metastasis suppressor, N-myc downstream regulated gene-1 (NDRG1), is a stress response protein that is involved in the inhibition of multiple oncogenic signaling pathways. Initial studies have linked NDRG1 and the endoplasmic reticulum (ER) stress response. Considering this, we extensively examined the mechanism by which NDRG1 regulates the ER stress response in pancreatic and colon cancer cells. We also examined the anti-cancer agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), which induces NDRG1 expression and causes ER stress. The expression of NDRG1 was demonstrated to regulate the three main arms of the ER stress response by: (1) increasing the expression of three major ER chaperones, binding immunoglobulin protein (BiP), calreticulin, and calnexin; (2) suppressing the protein kinase, RNA-activated (PKR)-like ER kinase (PERK); (3) inhibiting the inositol-requiring kinase 1α (IRE1α) arm; and (4) increasing the cleavage of activating transcription factor 6 (ATF6). An important finding was that NDRG1 enhances the anti-proliferative and anti-migratory activity of Dp44mT. This increased efficacy could be related to the following effects in the presence of Dp44mT and NDRG1, namely: markedly increased activation of the PERK target, eukaryotic translation initiation factor 2α (eIF2α); the maintenance of activating transcription factor 4 (ATF4) expression; high cytosolic Ca+2 that increases the sensitivity of cells to apoptosis via activation of the calmodulin-dependent kinase II (CaMKII) signaling cascade; and increased pro-apoptotic C/EBP-homologous protein (CHOP) expression. Collectively, this investigation dissects the molecular mechanisms through which NDRG1 manipulates the ER stress response and its ability to potentiate the activity of the potent anti-cancer agent, Dp44mT.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Estresse do Retículo Endoplasmático , Retículo Endoplasmático/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fator 6 Ativador da Transcrição/antagonistas & inibidores , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Cálcio/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Calnexina/genética , Calnexina/metabolismo , Calreticulina/genética , Calreticulina/metabolismo , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Chaperona BiP do Retículo Endoplasmático , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Endorribonucleases/antagonistas & inibidores , Endorribonucleases/genética , Endorribonucleases/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Quelantes de Ferro/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , eIF-2 Quinase/antagonistas & inibidores , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismo
6.
Osteoporos Int ; 30(4): 741-761, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30903209

RESUMO

We undertook a systematic review and meta-analysis of published papers assessing dietary protein and bone health. We found little benefit of increasing protein intake for bone health in healthy adults but no indication of any detrimental effect, at least within the protein intakes of the populations studied. This systematic review and meta-analysis analysed the relationship between dietary protein and bone health across the life-course. The PubMed database was searched for all relevant human studies from the 1st January 1976 to 22nd January 2016, including all bone outcomes except calcium metabolism. The searches identified 127 papers for inclusion, including 74 correlational studies, 23 fracture or osteoporosis risk studies and 30 supplementation trials. Protein intake accounted for 0-4% of areal BMC and areal BMD variance in adults and 0-14% of areal BMC variance in children and adolescents. However, when confounder adjusted (5 studies) adult lumbar spine and femoral neck BMD associations were not statistically significant. There was no association between protein intake and relative risk (RR) of osteoporotic fractures for total (RR(random) = 0.94; 0.72 to 1.23, I2 = 32%), animal (RR (random) = 0.98; 0.76 to 1.27, I2 = 46%) or vegetable protein (RR (fixed) = 0.97 (0.89 to 1.09, I2 = 15%). In total protein supplementation studies, pooled effect sizes were not statistically significant for LSBMD (total n = 255, MD(fixed) = 0.04 g/cm2 (0.00 to 0.08, P = 0.07), I2 = 0%) or FNBMD (total n = 435, MD(random) = 0.01 g/cm2 (-0.03 to 0.05, P = 0.59), I2 = 68%). There appears to be little benefit of increasing protein intake for bone health in healthy adults but there is also clearly no indication of any detrimental effect, at least within the protein intakes of the populations studied (around 0.8-1.3 g/Kg/day). More studies are urgently required on the association between protein intake and bone health in children and adolescents.


Assuntos
Densidade Óssea/efeitos dos fármacos , Proteínas Alimentares/farmacologia , Envelhecimento/fisiologia , Densidade Óssea/fisiologia , Dieta/estatística & dados numéricos , Proteínas Alimentares/administração & dosagem , Humanos , Proteínas do Leite/administração & dosagem , Proteínas do Leite/farmacologia , Fraturas por Osteoporose/etiologia , Fraturas por Osteoporose/fisiopatologia , Fraturas por Osteoporose/prevenção & controle , Medição de Risco/métodos , Proteínas de Soja/administração & dosagem , Proteínas de Soja/farmacologia
7.
Osteoporos Int ; 29(9): 1933-1948, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29740667

RESUMO

A summary of systematic reviews and meta-analyses addressing the benefits and risks of dietary protein intakes for bone health in adults suggests that dietary protein levels even above the current RDA may be beneficial in reducing bone loss and hip fracture risk, provided calcium intakes are adequate. Several systematic reviews and meta-analyses have addressed the benefits and risks of dietary protein intakes for bone health in adults. This narrative review of the literature summarizes and synthesizes recent systematic reviews and meta-analyses and highlights key messages. Adequate supplies of dietary protein are required for optimal bone growth and maintenance of healthy bone. Variation in protein intakes within the "normal" range accounts for 2-4% of BMD variance in adults. In older people with osteoporosis, higher protein intake (≥ 0.8-g/kg body weight/day, i.e., above the current RDA) is associated with higher BMD, a slower rate of bone loss, and reduced risk of hip fracture, provided that dietary calcium intakes are adequate. Intervention with dietary protein supplements attenuate age-related BMD decrease and reduce bone turnover marker levels, together with an increase in IGF-I and a decrease in PTH. There is no evidence that diet-derived acid load is deleterious for bone health. Thus, insufficient dietary protein intakes may be a more severe problem than protein excess in the elderly. Long-term, well-controlled randomized trials are required to further assess the influence of dietary protein intakes on fracture risk.


Assuntos
Proteínas Alimentares/administração & dosagem , Osteoporose/prevenção & controle , Equilíbrio Ácido-Base/efeitos dos fármacos , Densidade Óssea/efeitos dos fármacos , Remodelação Óssea/efeitos dos fármacos , Cálcio da Dieta/administração & dosagem , Proteínas Alimentares/efeitos adversos , Proteínas Alimentares/farmacologia , Humanos , Fraturas por Osteoporose/prevenção & controle , Medição de Risco/métodos
10.
Biochim Biophys Acta Gen Subj ; 1861(5 Pt A): 995-999, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28219722

RESUMO

We recently demonstrated that a novel storage and transport mechanism for nitric oxide (NO) mediated by glutathione-S-transferase P1 (GSTP1) and multidrug resistance protein 1 (MRP1/ABCC1), protects M1-macrophage (M1-MØ) models from large quantities of endogenous NO. This system stores and transports NO as dinitrosyl-dithiol-iron complexes (DNICs) composed of iron, NO and glutathione (GSH). Hence, this gas with contrasting anti- and pro-tumor effects, which has been assumed to be freely diffusible, is a tightly-regulated species in M1-MØs. These control systems prevent NO cytotoxicity and may be responsible for delivering cytotoxic NO as DNICs via MRP1 from M1-MØs, to tumor cell targets.


Assuntos
Glutationa S-Transferase pi/metabolismo , Células Matadoras Naturais/metabolismo , Macrófagos/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Óxido Nítrico/metabolismo , Substâncias Protetoras/metabolismo , Glutationa/metabolismo , Humanos
11.
Biochim Biophys Acta ; 1853(5): 1130-44, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25661197

RESUMO

Iron is a crucial transition metal for virtually all life. Two major destinations of iron within mammalian cells are the cytosolic iron-storage protein, ferritin, and mitochondria. In mitochondria, iron is utilized in critical anabolic pathways, including: iron-storage in mitochondrial ferritin, heme synthesis, and iron-sulfur cluster (ISC) biogenesis. Although the pathways involved in ISC synthesis in the mitochondria and cytosol have begun to be characterized, many crucial details remain unknown. In this review, we discuss major aspects of the journey of iron from its initial cellular uptake, its modes of trafficking within cells, to an overview of its downstream utilization in the cytoplasm and within mitochondria. The understanding of mitochondrial iron processing and its communication with other organelles/subcellular locations, such as the cytosol, has been elucidated by the analysis of certain diseases e.g., Friedreich's ataxia. Increased knowledge of the molecules and their mechanisms of action in iron processing pathways (e.g., ISC biogenesis) will shape the investigation of iron metabolism in human health and disease.


Assuntos
Células/metabolismo , Doença , Ferro/metabolismo , Animais , Transporte Biológico , Humanos , Proteínas Ferro-Enxofre/metabolismo , Mitocôndrias/metabolismo , Modelos Biológicos
12.
Free Radic Biol Med ; 75: 14-29, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25035074

RESUMO

Nitrogen monoxide (NO) is vital for many essential biological processes as a messenger and effector molecule. The physiological importance of NO is the result of its high affinity for iron in the active sites of proteins such as guanylate cyclase. Indeed, NO possesses a rich coordination chemistry with iron and the formation of dinitrosyl-dithiolato iron complexes (DNICs) is well documented. In mammals, NO generated by cytotoxic activated macrophages has been reported to play a role as a cytotoxic effector against tumor cells by binding and releasing intracellular iron. Studies from our laboratory have shown that two proteins traditionally involved in drug resistance, namely multidrug-resistance protein 1 and glutathione S-transferase, play critical roles in intracellular NO transport and storage through their interaction with DNICs (R.N. Watts et al., Proc. Natl. Acad. Sci. USA 103:7670-7675, 2006; H. Lok et al., J. Biol. Chem. 287:607-618, 2012). Notably, DNICs are present at high concentrations in cells and are biologically available. These complexes have a markedly longer half-life than free NO, making them an ideal "common currency" for this messenger molecule. Considering the many critical roles NO plays in health and disease, a better understanding of its intracellular trafficking mechanisms will be vital for the development of new therapeutics.


Assuntos
Glutationa Transferase/metabolismo , Compostos de Ferro/metabolismo , Macrófagos/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Óxido Nítrico/metabolismo , Óxidos de Nitrogênio/metabolismo , Animais , Transporte Biológico , Resistencia a Medicamentos Antineoplásicos , Humanos , Ferro/metabolismo
13.
J Nutr Health Aging ; 15(2): 147-52, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21365169

RESUMO

BACKGROUND: Poor nutritional status is often present among older adults who experience a fall. However, dietary intake and weight loss are often overlooked as potential factors. The objective of this study was to test the association between dietary protein intake and risk of subsequent falls in a population-based cohort of elderly men and women. METHODS: Dietary intake and clinic data from 807 men and women (ages 67-93 years) from the Framingham Original Cohort Study were analyzed. Protein intake (total, animal and plant) was assessed as a continuous variable and by tertile of intake. Falls were reported by participants using a validated questionnaire at two time points. Weight was ascertained at each examination to examine the effect of weight loss over follow-up. RESULTS: Higher dietary protein intakes were associated with a reduced odds of falling, although of borderline statistical significance (OR=0.80, 95% CI: 0.60-1.07) and were not associated with the rate of falls over follow-up (RR=0.93, 95%CI: 0.73-1.19). Tertile analyses tended towards a protective association, but most did not achieve statistical significance; there was no dose-response. For those who lost ≥ 5% of their baseline weight, higher intakes of total, animal and plant protein showed a significantly lower rate of subsequent falls. CONCLUSION: This work highlights the importance of adequate protein intake as a potentially modifiable risk factor for fall prevention in older adults. Further exploration of the interaction of protein intake and weight loss as related to falls is needed.


Assuntos
Acidentes por Quedas/estatística & dados numéricos , Envelhecimento , Proteínas Alimentares/administração & dosagem , Estado Nutricional , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Humanos , Estudos Longitudinais , Masculino , Recidiva , Fatores de Risco , Redução de Peso
16.
Osteoporos Int ; 20(11): 1853-61, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19347239

RESUMO

UNLABELLED: Vitamin C may play a role in bone health. In the Framingham Study, subjects with higher total or supplemental vitamin C intake had fewer hip fractures and non-vertebral fractures as compared to subjects with lower intakes. Therefore, vitamin C may have a protective effect on bone health in older adults. INTRODUCTION: Dietary antioxidants such as vitamin C may play a role in bone health. We evaluated associations of vitamin C intake (total, dietary, and supplemental) with incident hip fracture and non-vertebral osteoporotic fracture, over a 15- to 17-year follow-up, in the Framingham Osteoporosis Study. METHODS: Three hundred and sixty-six men and 592 women (mean age 75 +/- 5 years) completed a food frequency questionnaire (FFQ) in 1988-1989 and were followed for non-vertebral fracture until 2003 and hip fracture until 2005. Tertiles of vitamin C intake were created from estimates obtained using the Willett FFQ, after adjusting for total energy (residual method). Hazard ratios were estimated using Cox-proportional hazards regression, adjusting for covariates. RESULTS: Over follow-up 100 hip fractures occurred. Subjects in the highest tertile of total vitamin C intake had significantly fewer hip fractures (P trend = 0.04) and non-vertebral fractures (P trend = 0.05) compared to subjects in the lowest tertile of intake. Subjects in the highest category of supplemental vitamin C intake had significantly fewer hip fractures (P trend = 0.02) and non-vertebral fractures (P trend = 0.07) compared to non-supplement users. Dietary vitamin C intake was not associated with fracture risk (all P > 0.22). CONCLUSION: These results suggest a possible protective effect of vitamin C on bone health in older adults.


Assuntos
Ácido Ascórbico/administração & dosagem , Suplementos Nutricionais , Fraturas do Quadril/prevenção & controle , Fraturas por Osteoporose/prevenção & controle , Idoso , Idoso de 80 Anos ou mais , Densidade Óssea/efeitos dos fármacos , Fatores de Confusão Epidemiológicos , Dieta/estatística & dados numéricos , Métodos Epidemiológicos , Feminino , Fraturas do Quadril/epidemiologia , Fraturas do Quadril/etiologia , Humanos , Masculino , Massachusetts/epidemiologia , Pessoa de Meia-Idade , Osteoporose/complicações , Osteoporose/epidemiologia , Osteoporose/prevenção & controle , Fraturas por Osteoporose/epidemiologia , Fraturas por Osteoporose/etiologia , Potássio na Dieta/administração & dosagem
17.
J Thromb Haemost ; 6(1): 176-83, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17949478

RESUMO

BACKGROUND: Fibroblast growth factor (FGF)-2 is a critical growth factor in normal and malignant cell proliferation and tumor-associated angiogenesis. Fibrinogen and fibrin bind to FGF-2 and modulate FGF-2 functions. Furthermore, we have shown that extrahepatic epithelial cells are capable of endogenous production of fibrinogen. OBJECTIVE: Herein we examined the role of fibrinogen and FGF-2 interactions on prostate and lung adenocarcinoma cell growth in vitro. METHODS: Cell proliferation was measured by (3)H-thymidine uptake and the specificity of FGF-2-fibrinogen interactions was measured using wild-type and mutant FGF-2s, fibrinogen gamma-chain (FGG) RNAi and co-immunoprecipitation. Metabolic labeling, immunopurification and fluorography demonstrated de novo fibrinogen production. RESULTS: FGF-2 stimulated DU-145 cell proliferation, whereas neither FGF-2 nor fibrinogen affected the growth of PC-3 or A549 cells. Fibrinogen augmented the proliferative effect of FGF-2 on DU-145 cells. The role of fibrinogen in FGF-2-enhanced DNA synthesis was confirmed using an FGF-2 mutant that exhibits no binding affinity for fibrinogen. FGG transcripts were present in PC-3, A549 and DU-145 cells, but only PC-3 and A549 cells produced detectable levels of intact protein. RNAi-mediated knockdown of FGG expression resulted in decreased production of fibrinogen protein and inhibited (3)H-thymidine uptake in A549 and PC-3 cells by 60%, which was restored by exogenously added fibrinogen. FGF-2 and fibrinogen secreted by the cells were present in the medium as a soluble complex, as determined by coimmunoprecipitation studies. CONCLUSIONS: These data indicate that endogenously synthesized fibrinogen promotes the growth of lung and prostate cancer cells through interaction with FGF-2.


Assuntos
Fibrinogênio/biossíntese , Fator 2 de Crescimento de Fibroblastos/farmacologia , Neoplasias/metabolismo , Proliferação de Células/efeitos dos fármacos , Sinergismo Farmacológico , Fibrinogênio/genética , Fibrinogênio/farmacologia , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Neoplasias/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , RNA Interferente Pequeno/farmacologia , Células Tumorais Cultivadas
18.
J Thromb Haemost ; 2(9): 1629-36, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15333041

RESUMO

Endothelial cell responses at sites of injury occur in a fibrin matrix and are regulated by growth factors including those of the FGF and VEGF families. The pericellular proteolytic balance is important in these responses, and FGF-2 and VEGF up-regulate endothelial cell u-PA, u-PAR and PAI-1. Because both VEGF and FGF-2 bind to fibrinogen, we have examined the capacity of fibrinogen to modulate the up-regulation of these proteins by FGF-2 and VEGF. Confluent cultures of endothelial cells were exposed to FGF-2, VEGF, and fibrinogen or to combinations of growth factors with fibrinogen. Changes in mRNA levels of u-PA, u-PAR and PAI-1 were measured by Northern blot. FGF-2 increased u-PA, u-PAR, and PAI-1 mRNA, but there was a significantly greater induction when fibrinogen was added to FGF-2 at all concentrations. The potentiation by fibrinogen was particularly evident at an FGF-2 concentration of 0.1 ng mL(-1), which resulted in non-significant change in transcript levels by itself, but significantly increased up to 2.6-fold with fibrinogen. VEGF also increased endothelial cell expression of u-PA, u-PAR and PAI-1, but this effect was not potentiated by fibrinogen. Addition of LM609, a monoclonal antibody to alphaVbeta3, significantly inhibited induction of u-PA mRNA and activity by fibrinogen-bound FGF-2 compared to FGF-2. A monoclonal antibody to FGFR1 also inhibited u-PA mRNA expression induced by fibrinogen-bound FGF-2. We conclude that fibrinogen increases the capacity of FGF-2, but not of VEGF, to up-regulate u-PA, u-PAR, and PAI-1 in endothelial cells and that fibrinogen-bound FGF-2 requires alphaVbeta3 binding to up-regulate endothelial cell u-PA.


Assuntos
Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Fibrinogênio/metabolismo , Fator 2 de Crescimento de Fibroblastos/farmacologia , Inibidor 1 de Ativador de Plasminogênio/genética , Receptores de Superfície Celular/genética , Ativador de Plasminogênio Tipo Uroquinase/genética , Fator A de Crescimento do Endotélio Vascular/farmacologia , Sequência de Bases , Células Cultivadas , DNA Complementar/genética , Expressão Gênica/efeitos dos fármacos , Humanos , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Ativador de Plasminogênio Tipo Uroquinase , Proteínas Recombinantes/farmacologia
19.
Eur J Clin Invest ; 32(12): 943-8, 2002 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-12534455

RESUMO

BACKGROUND: Infection with human papilloma virus (HPV) is an important etiological factor in the development of cervical cancer and it has been proposed that individuals homozygous for Arg/Arg at codon-72 of p53 are seven times more susceptible to HPV-mediated cancer. In this study, we have analyzed the genetic predisposition of the India population to HPV infection and cervical carcinogenesis. METHODS: We investigated 71 cases of squamous cell carcinoma of the cervix, 14 cases of low-grade squamous intraepithelial lesions, 25 cases of high-grade squamous intraepithelial lesions and 29 noncancer controls for presence of HPV16/18 infections by L-1-specific PCR assay and Southern hybridization, and its association with polymorphism at p53 codon 72. RESULTS: We observed that 69.1% (76/110) of the cervical cancer patients were HPV positive, among which the presence of HPV16, 18 and 16/18 coinfection was 40.9%, 8.2% and 13.6%, respectively. The allele frequencies of the three p53 genotypes Arg/Arg, Arg/Pro and Pro/Pro in the HPV-positive tumour samples were 0.34, 0.57 and 0.09 in comparison with frequencies of 0.18, 0.44 and 0.38 for HPV-negative tumours. Hence, there is a significant difference in the allelic frequency of p53 Arg/Arg in high-risk HPV-infected cervical carcinoma cases (0.34) and HPV-negative carcinomas (0.18). CONCLUSION: Our results indicate a striking over-representation of homozygous arginine at codon 72 of p53 in HPV-associated cervical carcinogenesis. We conclude that women with Arg/Arg homozygous allele are more prone to infection by HPV16/18, which leads to cervical carcinomas having poor prognosis.


Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/virologia , Genes p53 , Papillomaviridae , Infecções por Papillomavirus/complicações , Polimorfismo Genético , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/virologia , Estudos de Casos e Controles , Feminino , Frequência do Gene , Predisposição Genética para Doença , Homozigoto , Humanos , Índia , Reação em Cadeia da Polimerase/métodos , Prognóstico
20.
Med Phys ; 28(7): 1410-5, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11488572

RESUMO

Permanent implant of the prostate using I-125 and Pd-103 seeds is a popular choice of treatment for early-stage prostate cancer in the United States. Evaluation of the quality of the implant is best based on the calculated dose distribution from postimplant computed tomography (CT) images. This task, however, has been time-consuming and inaccurate. We have developed an algorithm for automatic source localization from postimplant CT images. The only requirement of this algorithm is knowledge of the number of seeds present in the prostate, thus minimizing the need for human intervention. The algorithm processes volumetric CT data from the patient, and pixels of higher CT numbers are categorized into classes of definite and potential source pixels. A multithresholding technique is used to further determine the number of seeds and their precise locations in the CT volume data. A graphic user interface was developed to facilitate operator review of and intervention in the calculation and the results of the algorithm. This algorithm was tested on two phantoms containing nonradioactive seeds, one with 20 seeds in discrete locations and another with 100 seeds with small distances between seeds. The tests showed that the algorithm was able to identify the seed locations to within 1 mm of their physical locations for discrete seed locations. It was further able to separate seeds at close proximity to each other while maintaining an average seed localization error of less than 2 mm, with no operator intervention required.


Assuntos
Braquiterapia/métodos , Radioisótopos do Iodo/uso terapêutico , Neoplasias da Próstata/radioterapia , Algoritmos , Humanos , Masculino , Modelos Estatísticos , Imagens de Fantasmas , Reprodutibilidade dos Testes , Software
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