RESUMO
Gas explosion (GE)-induced traumatic brain injury (TBI) can affect thyroid hormone (TH) homeostasis in miners. This study evaluated the effects of hepatic transthyretin and hypothalamic-pituitary-thyroid (HPT) axis on thyroids and explored the protective effect and mechanism of curcumin on GE-induced TBI. Thirty rats were randomly divided into three groups (10 per group): first group (control group)-rats received GE treatment once; second group (GE group)-rats received GE treatment (200 m from the source of the explosion once); third group (GE + Cur group)-rats received curcumin (Cur) by lavage at a dose of 100 mg/kg/day once every other day for 7 days after receiving GE. After GE, the pathological changes were analyzed by hemotoxylin and eosin staining, and the levels of serum reactive oxygen species (ROS), urine iodine (UI), THs, nuclear factor-kappa B (NF-κB), superoxide dismutase (SOD), glutathione peroxidase (Gpx), and malondialdehyde (MDA) were analyzed using ELISA. Expression of proteins in the HPT axis of rats was examined by immunohistochemistry and Western blotting. We found that GE could induce pathologic changes in rat thyroid and liver. Serum levels of THs, NF-κB and serum redox state became unbalanced in rats after GE. GE could inhibit the biosynthesis and biotransformation of THs by affecting key HPT axis proteins. Additionally, GE reduced the level of hepatic transthyretin. Serum THs levels and thyroid sections were almost recovered to normal after curcumin treatment. The aforementioned key HPT axis proteins in the curcumin group showed opposite expression trends. In summary, GE affected THs balance while curcumin can protect against these injury effects by affecting TH biosynthesis, biotransformation, and transport, and inducing oxidative stress and inflammatory responses.
Assuntos
Lesões Encefálicas Traumáticas , Curcumina , Iodo , Animais , Curcumina/farmacologia , Amarelo de Eosina-(YS) , Explosões , Glutationa Peroxidase/metabolismo , Hematoxilina/farmacologia , Masculino , Malondialdeído/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo , Pré-Albumina/metabolismo , Pré-Albumina/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Glândula Tireoide/metabolismo , Hormônios Tireóideos/metabolismoRESUMO
Age-related wild-type transthyretin amyloidosis (wtATTR) is characterized by systemic deposition of amyloidogenic fibrils of misfolded transthyretin (TTR) in the connective tissue of many organs. In the heart, this leads to age-related heart failure with preserved ejection fraction (HFpEF). The hypothesis tested is that TTR deposited in vitro disrupts cardiac myocyte cell-to-cell and cell-to-matrix adhesion complexes, resulting in altered calcium handling, force generation, and sarcomeric disorganization. Human iPSC-derived cardiomyocytes and neonatal rat ventricular myocytes (NRVMs), when grown on TTR-coated polymeric substrata mimicking the stiffness of the healthy human myocardium (10 kPa), had decreased contraction and relaxation velocities as well as decreased force production measured using traction force microscopy. Both NRVMs and adult mouse atrial cardiomyocytes had altered calcium kinetics with prolonged transients when cultured on TTR fibril-coated substrates. Furthermore, NRVMs grown on stiff (~GPa), flat or microgrooved substrates coated with TTR fibrils exhibited significantly decreased intercellular electrical coupling as shown by FRAP dynamics of cells loaded with the gap junction-permeable dye calcein-AM, along with decreased gap junction content as determined by quantitative connexin 43 staining. Significant sarcomeric disorganization and loss of sarcomere content, with increased ubiquitin localization to the sarcomere, were seen in NRVMs on various TTR fibril-coated substrata. TTR presence decreased intercellular mechanical junctions as evidenced by quantitative immunofluorescence staining of N-cadherin and vinculin. Current therapies for wtATTR are cost-prohibitive and only slow the disease progression; therefore, better understanding of cardiomyocyte maladaptation induced by TTR amyloid may identify novel therapeutic targets.
Assuntos
Neuropatias Amiloides Familiares , Insuficiência Cardíaca , Animais , Cálcio , Cálcio da Dieta , Camundongos , Miócitos Cardíacos , Pré-Albumina/química , Pré-Albumina/farmacologia , Ratos , Sarcômeros , Volume SistólicoRESUMO
BACKGROUND: Tafamidis is approved in many countries for the treatment of transthyretin amyloid cardiomyopathy. This study reports data on the long-term efficacy of tafamidis from an ongoing long-term extension (LTE) to the pivotal ATTR-ACT (Tafamidis in Transthyretin Cardiomyopathy Clinical Trial). METHODS: Patients with transthyretin amyloid cardiomyopathy who completed ATTR-ACT could enroll in an LTE, continuing with the same tafamidis dose or, if previously treated with placebo, randomized (2:1) to tafamidis meglumine 80 or 20 mg. All patients in the LTE transitioned to tafamidis free acid 61 mg (bioequivalent to tafamidis meglumine 80 mg) following a protocol amendment. In this interim analysis, all-cause mortality was assessed in patients treated with tafamidis meglumine 80 mg in ATTR-ACT continuing in the LTE, compared with those receiving placebo in ATTR-ACT transitioning to tafamidis in the LTE. RESULTS: Median follow-up was 58.5 months in the continuous tafamidis group (n=176) and 57.1 months in the placebo to tafamidis group (n=177). There were 79 (44.9%) deaths with continuous tafamidis and 111 (62.7%) with placebo to tafamidis (hazard ratio, 0.59 [95% CI, 0.44-0.79]; P<0.001). Mortality was also reduced in the continuous tafamidis (versus placebo to tafamidis) subgroups of: variant transthyretin amyloidosis (0.57 [0.33-0.99]; P=0.05) and wild-type transthyretin amyloidosis (0.61 [0.43-0.87]; P=0.006); and baseline New York Heart Association class I and II (0.56 [0.38-0.82]; P=0.003) and class III (0.65 [0.41-1.01]; P=0.06). CONCLUSIONS: In the LTE, patients initially treated with tafamidis in ATTR-ACT had substantially better survival than those first treated with placebo, highlighting the importance of early diagnosis and treatment in transthyretin amyloid cardiomyopathy. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT01994889 and NCT02791230.
Assuntos
Neuropatias Amiloides Familiares/tratamento farmacológico , Neuropatias Amiloides Familiares/mortalidade , Benzoxazóis/farmacologia , Cardiomiopatias/mortalidade , Tempo , Idoso , Idoso de 80 Anos ou mais , Neuropatias Amiloides Familiares/complicações , Cardiomiopatias/complicações , Cardiomiopatias/tratamento farmacológico , Feminino , Insuficiência Cardíaca/complicações , Insuficiência Cardíaca/tratamento farmacológico , Insuficiência Cardíaca/mortalidade , Hospitalização/estatística & dados numéricos , Humanos , Masculino , Pessoa de Meia-Idade , Pré-Albumina/farmacologia , Modelos de Riscos ProporcionaisRESUMO
Advances in cancer research have led to the development of new therapeutics with significant and durable responses such as immune checkpoint inhibitors. More recent therapies aim to stimulate anti-tumor immune responses by targeting the tumor necrosis factor (TNF) receptors, however this approach has been shown to require clustering of receptors in order to achieve a significant response. Here we present a perspective on using transthyretin, a naturally occurring serum protein, as a drug delivery platform to enable cross-linking independent clustering of targets. TTR forms a stable homo-tetramer with exposed termini that make TTR a highly versatile platform for generating multimeric antibody fusions to enable enhanced target clustering. Fusions with antibodies or Fabs targeting TRAILR2 were shown to have robust cytotoxic activity in vitro and in vivo in colorectal xenograft models demonstrating that TTR is a highly versatile, stable, therapeutic fusion platform that can be used with antibodies, Fabs and other bioactive fusion partners and has broad applications in oncology and infectious disease research.
Assuntos
Peptídeos , Pré-Albumina , Anticorpos , Análise por Conglomerados , Humanos , Pré-Albumina/metabolismo , Pré-Albumina/farmacologiaRESUMO
Dissociation of transthyretin (TTR) tetramers may lead to misfolding and aggregation of proamyloidogenic monomers, which underlies TTR amyloidosis (ATTR) pathophysiology. ATTR is a progressive disease resulting from the deposition of toxic fibrils in tissues that predominantly presents clinically as amyloid cardiomyopathy and peripheral polyneuropathy. Ligands that bind to and kinetically stabilize TTR tetramers prohibit their dissociation and may prevent ATTR onset. Drawing from clinically investigated AG10, we designed a constrained congener (14) that exhibits excellent TTR tetramer binding potency, prevents TTR aggregation in a gel-based assay, and possesses desirable pharmacokinetics in mice. Additionally, 14 significantly lowers murine serum retinol binding protein 4 (RBP4) levels despite a lack of binding at that protein's all-trans-retinol site. We hypothesize that kinetic stabilization of TTR tetramers via 14 is allosterically hindering all-trans-retinol-dependent RBP4-TTR tertiary complex formation and that the compound could present ancillary therapeutic utility for indications treated with RBP4 antagonists, such as macular degeneration.
Assuntos
Neuropatias Amiloides Familiares/tratamento farmacológico , Pré-Albumina/farmacologia , Proteínas Plasmáticas de Ligação ao Retinol/antagonistas & inibidores , Neuropatias Amiloides Familiares/metabolismo , Animais , Relação Dose-Resposta a Droga , Cinética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Estrutura Molecular , Pré-Albumina/síntese química , Pré-Albumina/química , Proteínas Plasmáticas de Ligação ao Retinol/deficiência , Proteínas Plasmáticas de Ligação ao Retinol/metabolismo , Relação Estrutura-AtividadeRESUMO
Transthyretin (TTR) has been proved to repress neovascularization in diabetic retinopathy environment by regulating the molecules in and downstream of the STAT-4/miR-223-3p/FBXW7 signal pathway; however, the details of its direct targets are still not well understood. The interaction between TTR and a target in nucleus of human retinal microvascular endothelial cells (hRECs), heterogeneous nuclear ribonucleoprotein (hnRNP) A2B1, was screened by immunoprecipitation (IP) and mass spectrum (MS), and it was further confirmed by co-immunoprecipitation (co-IP). Regarding ZDOCK analysis using Discovery Studio, the interface and potential binding sites between TTR and hnRNPA2B1 were simulated; mutants were designed in these regions and five soluble ones were recombinantly expressed and prepared; the interaction between TTR and hnRNPA2B1 were disrupted by several mutated residues. In addition, for several mutated TTRs, the inhibition activities against the proliferation, migration and tube formation of hRECs were absent in vitro. Following the disruption of TTR-hnRNPA2B1, the molecules in and downstream of STAT-4/miR-223-3p/FBXW7 signal pathway, including STAT-4, miR-223-3p, FBXW7 p-Akt and Notch1 could not be regulated by TTR mutants; therefore, a TTR-hnRNPA2B1/STAT-4/miR-223-3p/FBXW7 was proposed. In conclusion, this work suggested that TTR should play a physiological role in diabetic environment by the direct binding with hnRNPA2B1, and it provided a theoretical basis for clinical diagnosis, therapy and further application.
Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/metabolismo , Hiperglicemia/metabolismo , Pré-Albumina/farmacologia , Retina/efeitos dos fármacos , Células Cultivadas , Retinopatia Diabética/metabolismo , Proteínas F-Box/metabolismo , Ontologia Genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/química , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/genética , Humanos , Espectrometria de Massas , MicroRNAs/genética , MicroRNAs/metabolismo , Mutação , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Pré-Albumina/química , Pré-Albumina/genética , Pré-Albumina/metabolismo , Proteínas Recombinantes , Retina/metabolismo , Fator de Transcrição STAT4/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Preeclampsia (PE) is a hypertensive disorder of pregnancy. Early diagnosis of PE is currently contingent on regular prenatal physical examinations and may be facilitated by identification of novel diagnostic markers. Transthyretin (TTR), also known as prealbumin, is primarily responsible for maintaining the normal levels of thyroxine and retinol binding protein. The expression of TTR is lower in patients with severe PE as compared with healthy controls. Here, we examined the suitability of TTR as a diagnostic marker in pregnant hypertensive rats. N'-nitro-l-arginine-methylesterhydrochloride (l-NAME) was used to generate a rat model of hypertension during pregnancy. Rat placental trophoblast cells were divided into control and TTR groups for in vitro experiments. Systolic blood pressure, diastolic blood pressure, mean blood pressure and urinary protein of hypertensive pregnant rats were higher than those of healthy pregnant rats, but these effects could be reversed by TTR treatment. There were no significant changes in blood pressure and urinary protein in healthy pregnant rats before or after TTR treatment. TTR levels in the serum and placental tissues of pregnant hypertensive rats were significantly reduced compared with those of healthy pregnant rats. Changes in placental and fetal weights in the hypertensive model could also be rescued by TTR treatment. TTR treatment significantly increased the level of matrix metalloproteinase-2/9 in hypertensive rats. Finally, in vivo and in vitro experiments demonstrated that TTR effectively increased the migration and invasion of rat placental trophoblast cells, as well as matrix metalloproteinase-2/9 levels in these cells. In conclusion, our data from a rat model suggest that TTR may have potential as a novel marker for PE diagnosis.
Assuntos
Pré-Albumina/farmacologia , Trofoblastos/efeitos dos fármacos , Animais , Movimento Celular/efeitos dos fármacos , Feminino , Humanos , Placenta , Gravidez , Ratos , Trofoblastos/metabolismo , Células Tumorais CultivadasRESUMO
Transthyretin (TTR) is an abundant homotetrameric serum protein and was selected here for engineering higher-valency molecules because of its compact size, simple structure, and natural propensity to tetramerize. To demonstrate this utility, we fused TTR to the C terminus of conatumumab, an antibody that targets tumor necrosis factor-related apoptosis-inducing ligand receptor 2, as heavy chains to form antibody dimers and Fab heavy chains to form Fab tetramers. Moreover, we used constant heavy domain 3 heterodimerization substitutions to create TTR-mediated conatumumab tetramers. The conatumumab-TTR fusions displayed substantially enhanced potency in cell-based assays, as well as in murine tumor xenograft models. We conclude that antibody-TTR fusions may provide a powerful platform for multimerizing antibody and Fab fragments to enhance the capabilities of human therapeutics that benefit from target clustering and higher-order antigen-binding valency.
Assuntos
Anticorpos Monoclonais , Antineoplásicos Imunológicos , Fragmentos Fab das Imunoglobulinas , Neoplasias Experimentais , Pré-Albumina , Multimerização Proteica , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/farmacocinética , Antineoplásicos Imunológicos/farmacologia , Linhagem Celular Tumoral , Humanos , Fragmentos Fab das Imunoglobulinas/genética , Fragmentos Fab das Imunoglobulinas/farmacologia , Camundongos , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Pré-Albumina/genética , Pré-Albumina/farmacocinética , Pré-Albumina/farmacologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/farmacocinética , Proteínas Recombinantes de Fusão/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Transthyretin (TTR) is a tetrameric protein, and its dissociation, aggregation, deposition, and misfolding are linked to several human amyloid diseases. As the main transporter for thyroxine (T4) in plasma and cerebrospinal fluid, TTR contains two T4-binding sites, which are docked with T4 and subsequently maintain the structural stability of TTR homotetramer. Affected by genetic disorders and detrimental environmental factors, TTR degrades to monomer and/or form amyloid fibrils. Reasonably, stabilization of TTR might be an efficient strategy for the treatment of TTR-related amyloidosis. However, only 10-25% of T4 in the plasma is bound to TTR under physiological conditions. Expectedly, T4 analogs with different structures aiming to bind to T4 pockets may displace the functions of T4. So far, a number of compounds including both natural and synthetic origin have been reported. In this paper, we summarized the potent inhibitors, including bisaryl structure-based compounds, flavonoids, crown ethers, and carboranes, for treating TTR-related amyloid diseases and the combination modes of some compounds binding to TTR protein.
Assuntos
Amiloide/antagonistas & inibidores , Amiloide/genética , Amiloidose , Pré-Albumina/farmacologia , Amiloide/metabolismo , Humanos , Modelos Moleculares , Estrutura Molecular , Pré-Albumina/químicaRESUMO
Peripheral neuropathy occurs in the setting of both hereditary and acquired amyloidosis. The most common form of hereditary amyloidosis is caused by 1 of 140 mutations in the transthyretin (TTR) gene, which can lead to neuropathic hereditary transthyretin amyloidosis (hATTR; previously referred to as transthyretin familial amyloid polyneuropathy), whereas acquired immunoglobulin light chain (AL) amyloidosis is the most common acquired form. Patients typically present with a sensorimotor polyneuropathy, focal neuropathy such as carpal tunnel syndrome, or autonomic neuropathy. When neuropathy is the sole or dominant presenting symptom, the diagnosis is commonly delayed. With the advent of new drug therapies for AL amyloidosis and hATTR amyloidosis, including proteasome inhibitors, TTR silencers, and TTR protein stabilizers, the neurologist is uniquely positioned to diagnose neurologic manifestations of systemic amyloidosis, leading to earlier disease identification and treatment. This article reviews the epidemiology, clinical presentations, pathophysiology, diagnostic workup, and treatment of neuropathy in the setting of amyloidosis.
Assuntos
Neuropatias Amiloides Familiares/terapia , Amiloidose de Cadeia Leve de Imunoglobulina/terapia , Mutação/genética , Neuropatias Amiloides Familiares/diagnóstico , Neuropatias Amiloides Familiares/genética , Humanos , Amiloidose de Cadeia Leve de Imunoglobulina/diagnóstico , Amiloidose de Cadeia Leve de Imunoglobulina/genética , Doenças do Sistema Nervoso/complicações , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/terapia , Pré-Albumina/farmacologiaRESUMO
Early detection of lung cancer offers an important opportunity to decrease mortality while it is still treatable and curable. Thirteen secretory proteins that are Stat3 downstream gene products were identified as a panel of biomarkers for lung cancer detection in human sera. This panel of biomarkers potentially differentiates different types of lung cancer for classification. Among them, the transthyretin (TTR) concentration was highly increased in human serum of lung cancer patients. TTR concentration was also induced in the serum, bronchoalveolar lavage fluid, alveolar type II epithelial cells, and alveolar myeloid cells of the CCSP-rtTA/(tetO)7-Stat3C lung tumor mouse model. Recombinant TTR stimulated lung tumor cell proliferation and growth, which were mediated by activation of mitogenic and oncogenic molecules. TTR possesses cytokine functions to stimulate myeloid cell differentiation, which are known to play roles in tumor environment. Further analyses showed that TTR treatment enhanced the reactive oxygen species production in myeloid cells and enabled them to become functional myeloid-derived suppressive cells. TTR demonstrated a great influence on a wide spectrum of endothelial cell functions to control tumor and immune cell migration and infiltration. TTR-treated endothelial cells suppressed T cell proliferation. Taken together, these 13 Stat3 downstream inducible secretory protein biomarkers potentially can be used for lung cancer diagnosis, classification, and as clinical targets for lung cancer personalized treatment if their expression levels are increased in a given lung cancer patient in the blood.
Assuntos
Células Endoteliais/imunologia , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/imunologia , Pré-Albumina/imunologia , Células Epiteliais Alveolares/imunologia , Animais , Biomarcadores Tumorais/sangue , Líquido da Lavagem Broncoalveolar/química , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Humanos , Neoplasias Pulmonares/classificação , Melanoma Experimental , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Mieloides/imunologia , Neoplasias Experimentais/imunologia , Pré-Albumina/farmacologia , Proteínas Recombinantes/farmacologia , Fator de Transcrição STAT3/genéticaRESUMO
Protein misfolding and aggregation are associated with a number of human degenerative diseases. In spite of the enormous research efforts to develop effective strategies aimed at interfering with the pathogenic cascades induced by misfolded/aggregated peptides/proteins, the necessary detailed understanding of the molecular bases of amyloid formation and toxicity is still lacking. To this aim, approaches able to provide a global insight in amyloid-mediated physiological alterations are of importance. In this study, we exploited Fourier transform infrared microspectroscopy, supported by multivariate analysis, to investigate in situ the spectral changes occurring in cultured intact HL-1 cardiomyocytes exposed to wild type (WT) or mutant (L55P) transthyretin (TTR) in native, or amyloid conformation. The presence of extracellular deposits of amyloid aggregates of WT or L55P TTR, respectively, is a key hallmark of two pathological conditions, known as senile systemic amyloidosis and familial amyloid polyneuropathy. We found that the major effects, associated with modifications in lipid properties and in the cell metabolic/phosphorylation status, were observed when natively folded WT or L55P TTR was administered to the cells. The effects induced by aggregates of TTR were milder and in some cases displayed a different timing compared to those elicited by the natively folded protein.
Assuntos
Mutação , Miócitos Cardíacos/citologia , Pré-Albumina/química , Pré-Albumina/farmacologia , Amiloide/efeitos dos fármacos , Amiloide/metabolismo , Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Lipídeos de Membrana/química , Análise Multivariada , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Fosforilação/efeitos dos fármacos , Pré-Albumina/genética , Agregados Proteicos , Dobramento de Proteína , Espécies Reativas de Oxigênio/metabolismo , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
BACKGROUND: TTR (transthyretin) cardiac amyloidosis is caused by dissociation of TTR into monomers, which misassemble into amyloid fibrils. TTR stabilizers act at the dimer-dimer interface to prevent dissociation. We investigated differences in survival among patients with TTR cardiac amyloidosis on stabilizer medications compared with those not on stabilizers. METHODS AND RESULTS: A retrospective study of patients with TTR cardiac amyloidosis presenting to a single center was conducted. Baseline characteristics were compared between those treated with stabilizers and those not treated with stabilizers. Cox proportional hazards modeling assessed for univariate predictors of the composite outcome of death or orthotopic heart transplant (OHT). Multivariable Cox proportional hazards assessed whether stabilizer treatment was independently associated with improved death or OHT after controlling for significant univariate predictors. One hundred twenty patients (mean age, 75±8, 88% male) were included: 29 patients who received stabilizers and 91 patients who did not. Stabilizer use was associated with a lower risk of the combined end point of death or OHT (hazard ratio, 0.32; 95% confidence interval, 0.18-0.58; P<0.0001). Subjects treated with stabilizers were more likely to be of White race (93% versus 55%; P<0.001), classified as New York Heart Association classes I and II (79% versus 38%; P=0.002), less likely to have a mutation (10% versus 36%; P=0.010), have lower troponin I (median 0.06 versus 0.12 ng/mL; P=0.002), and higher left ventricular ejection fraction (49% versus 40%; P=0.011), suggesting earlier stage of disease. In multivariable Cox analysis, the association between stabilizer and death or OHT persisted when adjusted for all noncollinear univariate predictors with P<0.05 (hazard ratio, 0.37; 95% confidence interval, 0.19-0.75; P=0.003). CONCLUSIONS: TTR stabilizers are associated with decreased death and OHT in TTR cardiac amyloidosis. These results need to be confirmed by ongoing randomized clinical trials.
Assuntos
Neuropatias Amiloides Familiares/tratamento farmacológico , Insuficiência Cardíaca/tratamento farmacológico , Pré-Albumina/farmacologia , Idoso , Feminino , Insuficiência Cardíaca/complicações , Insuficiência Cardíaca/genética , Transplante de Coração/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/efeitos dos fármacos , Troponina I/genética , Função Ventricular Esquerda/efeitos dos fármacosRESUMO
OBJECTIVE: Skin amyloid deposits can occur as part of systemic amyloidoses including those involving misfolded- aggregated transthyretins (agTTR). Pathological effects of agTTR on the skin are not well understood. The main objective of the current study was to examine the toxicity of agTTR upon a human keratinocyte cell line. METHODS: Cells were analyzed for indicators of oxidative stress after treatment with normal soluble TTR or the same pre-aggregation concentration of agTTR. Hydrogen peroxide production was analyzed as an indicator for the involvement of reactive oxygen species. RESULTS: Treatment of cells with agTTR significantly increased hydrogen peroxide production (p < 0.05 vs. controls). Glutathione (GSH) and catalase were analyzed as indicators of endogenous cellular antioxidant activity. Treatment of cells with agTTR resulted in significant decreases in both catalase activity and GSH levels (p < 0.05 vs. controls). CONCLUSION: agTTR disrupts redox balance and induces oxidative stress in these epidermoid cells.
Assuntos
Catalase/metabolismo , Glutationa/metabolismo , Queratinócitos/efeitos dos fármacos , Pré-Albumina/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/metabolismo , Catalase/efeitos dos fármacos , Linhagem Celular , Células Epiteliais/metabolismo , Glutationa/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio/metabolismo , Queratinócitos/metabolismo , Estresse Oxidativo , Pré-Albumina/químicaRESUMO
The cytotoxicity of amyloidogenic proteins such as transthyretin (TTR) has implications for neurodegeneration and other pathologies, but is not well understood. In the current study, potential effects of misfolded, aggregated TTRs (agTTR) upon a major cell membrane function-endocytosis-were assessed. Internalization of transferrin (Tf), a ligand whose receptor-mediated endocytosis is well characterized, was analyzed in different cell types after treatment with agTTR. The results indicate disruption of Tf endocytosis: 20-25% inhibition by agTTR relative to the same concentrations of normal soluble TTR, or relative to another control protein, albumin (p<0.05 for agTTR relative to controls). No statistically significant difference was observed for cell surface Tf binding between agTTR-treated and control cells. This is the first evidence for endocytic disruption by agTTR, and presents a novel cytotoxicity mechanism that may account for previously reported inhibitory effects of amyloidogenic TTR on neuronal growth.
Assuntos
Endocitose/efeitos dos fármacos , Pré-Albumina/farmacologia , Animais , Bioensaio , Transporte Biológico/efeitos dos fármacos , Linhagem Celular Tumoral , Colorimetria , Humanos , Camundongos , Pré-Albumina/química , Pré-Albumina/metabolismo , Ligação Proteica/efeitos dos fármacosRESUMO
Objective: To explore the effects of transthyretin (TTR) on biological behavior of retinal pigment epithelial cells (RPECs) and retinal microvascular epithelial cells (RMVECs). Methods: RPECs were cultured with exogenous TTR to explore the effect of TTR on the proliferation of RPECs. The expression of TTR of RPECs was silenced by TTR specific small interfering RNA and the expression of TTR was detected by using Western blotting to identify the efficacy of TTR silence. The level of vascular endothelial growth factor (VEGF) massage RNA was detected by using RT-PCR to identify the interaction between VEGF and TTR. The different proliferation and migration abilities of RMVECs with different expressions of TTR were measured by transwell system. Results: MTT assay showed that RPECs with 0 µmol/L TTR glowed faster than with 4 µmol/L TTR (t=18.08, P<0.0001). The expression level of TTR was decreased in the small interfering RNA group as compared with the negative control group (P<0.05). RT-PCR assay showed no differential expression of VEGF after the silencing of TTR (P>0.05). The transwell assay showed RMVECs with the silence of TTR proliferated more slowly than RMVECs without the treatment (t=4.901, P=0.0012), and also migrated more slowly (t=4.213, P=0.0029). Conclusions: TTR can inhibit the proliferation of RPECs and promote the proliferation and migration of RMVECs without the help of VEGF, the mechanism of which may be worth further study. (Chin J Ophthalmol, 2016, 52: 856-860).
Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Pré-Albumina/farmacologia , Epitélio Pigmentado da Retina/efeitos dos fármacos , Western Blotting , Células Cultivadas , Células Epiteliais/fisiologia , Inativação Gênica , Humanos , Pré-Albumina/metabolismo , RNA Mensageiro/metabolismo , RNA Interferente Pequeno , Epitélio Pigmentado da Retina/fisiologia , Pigmentos da Retina , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: The apoptosis of human umbilical vein endothelial cells has been reportedly induced by the protein transthyretin (TTR). In human ocular tissue, TTR is generally considered to be secreted mainly by retinal pigment epithelial cells (hRPECs); however, whether TTR affects the development of neovascularization in diabetic retinopathy (DR) remains unclear. METHODS: Natural and simulated DR media were used to culture human retinal microvascular endothelial cells (hRECs). Hyperglycemia was simulated by increasing the glucose concentration from 5.5 mM up to 25 mM, while hypoxia was induced with 200 µM CoCl2. To understand the effects of TTR on hRECs, cell proliferation was investigated under natural and DR conditions. Overexpression of TTR, an in vitro wound-healing assay, and a tube formation assay were employed to study the repression of TTR on hRECs. Real-time fluorescence quantitative PCR (qRT-PCR) was used to study the mRNA levels of DR-related genes, such as Tie2, VEGFR1, VEGFR2, Angpt1, and Angpt2. RESULTS: The proliferation of hRECs was significantly decreased in the simulated hyperglycemic and hypoxic DR environments. The cells were further repressed by added exogenous or endogenous TTR only under hyperglycemic conditions. The in vitro migration and tube formation processes of the hRECs were inhibited with TTR; furthermore, in the hyperglycemia and hyperglycemia/hypoxia environments, the levels of Tie2 and Angpt1 mRNA were enhanced with exogenous TTR, while those of VEGFR1, VEGFR2, and Angpt1 were repressed. CONCLUSIONS: In hyperglycemia, the proliferation, migration, and neovascularization of hRECs were significantly inhibited by TTR. The key genes for DR neovascularization, including Tie2, VEGFR1, VEGFR2, Angpt1, and Angpt2, were regulated by TTR. Under DR conditions, TTR significantly represses neovascularization by inhibiting the proliferation, migration and tube formation of hRECs.
Assuntos
Retinopatia Diabética/metabolismo , Células Endoteliais/efeitos dos fármacos , Pré-Albumina/farmacologia , Neovascularização Retiniana/metabolismo , Angiopoietina-1/genética , Angiopoietina-1/metabolismo , Angiopoietina-2/genética , Angiopoietina-2/metabolismo , Movimento Celular , Proliferação de Células , Células Cultivadas , Retinopatia Diabética/genética , Células Endoteliais/metabolismo , Glucose/farmacologia , Humanos , Hipóxia/metabolismo , Técnicas de Cultura de Órgãos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Neovascularização Retiniana/genética , Vasos Retinianos/fisiologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Transthyretin (TTR) is a blood and cerebrospinal fluid transporter of thyroxine and retinol. Gene expression profiling revealed an elevation of Ttr expression in the dorsomedial hypothalamus (DMH) of rats with exercise-induced anorexia, implying that central TTR may also play a functional role in modulating food intake and energy balance. To test this hypothesis, we have examined the effects of brain TTR on food intake and body weight and have further determined hypothalamic signaling that may underlie its feeding effect in rats. We found that intracerebroventricular (icv) administration of TTR in normal growing rats decreased food intake and body weight. This effect was not due to sickness as icv TTR did not cause a conditioned taste aversion. ICV TTR decreased neuropeptide Y (NPY) levels in the DMH and the paraventricular nucleus (P < 0.05). Chronic icv infusion of TTR in Otsuka Long-Evans Tokushima Fatty rats reversed hyperphagia and obesity and reduced DMH NPY levels. Overall, these results demonstrate a previously unknown anorectic action of central TTR in the control of energy balance, providing a potential novel target for treating obesity and its comorbidities.
Assuntos
Peso Corporal/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Hiperfagia/prevenção & controle , Obesidade/prevenção & controle , Pré-Albumina/farmacologia , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Núcleo Arqueado do Hipotálamo/metabolismo , Western Blotting , Células Cultivadas , Perfilação da Expressão Gênica/métodos , Hiperfagia/metabolismo , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Infusões Intraventriculares , Masculino , Neuropeptídeo Y/metabolismo , Obesidade/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Pré-Albumina/administração & dosagem , Pré-Albumina/fisiologia , Ratos Endogâmicos OLETF , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: Amyloid deposits are prevalent in osteoarthritic (OA) joints. We undertook this study to define the dominant precursor and to determine whether the deposits affect chondrocyte functions. METHODS: Amyloid deposition in human normal and OA knee cartilage was determined by Congo red staining. Transthyretin (TTR) in cartilage and synovial fluid was analyzed by immunohistochemistry and Western blotting. The effects of recombinant amyloidogenic and nonamyloidogenic TTR variants were tested in human chondrocyte cultures. RESULTS: Normal cartilage from young donors did not contain detectable amyloid deposits, but 7 of 12 aged normal cartilage samples (58%) and 12 of 12 OA cartilage samples (100%) had Congo red staining with green birefringence under polarized light. TTR, which is located predominantly at the cartilage surfaces, was detected in all OA cartilage samples and in a majority of aged normal cartilage samples, but not in normal cartilage samples from young donors. Chondrocytes and synoviocytes did not contain significant amounts of TTR messenger RNA. Synovial fluid TTR levels were similar in normal and OA knees. In cultured chondrocytes, only an amyloidogenic TTR variant induced cell death as well as the expression of proinflammatory cytokines and extracellular matrix-degrading enzymes. The effects of amyloidogenic TTR on gene expression were mediated in part by Toll-like receptor 4, receptor for advanced glycation end products, and p38 MAPK. TTR-induced cytotoxicity was inhibited by resveratrol, a plant polyphenol that stabilizes the native tetrameric structure of TTR. CONCLUSION: These findings are the first to suggest that TTR amyloid deposition contributes to cell and extracellular matrix damage in articular cartilage in human OA and that therapies designed to reduce TTR amyloid formation might be useful.