Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 11 de 11
Filtrar
1.
Nucleic Acids Res ; 52(8): 4295-4312, 2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38416579

RESUMO

5-Fluorouracil (5-FU) is the first-line chemotherapeutic agent in colorectal cancer, and resistance to 5-FU easily emerges. One of the mechanisms of drug action and resistance of 5-FU is through DNA incorporation. Our quantitative reverse-transcription PCR data showed that one of the translesion synthesis (TLS) DNA polymerases, DNA polymerase η (polη), was upregulated within 72 h upon 5-FU administration at 1 and 10 µM, indicating that polη is one of the first responding polymerases, and the only TLS polymerase, upon the 5-FU treatment to incorporate 5-FU into DNA. Our kinetic studies revealed that 5-fluoro-2'-deoxyuridine triphosphate (5FdUTP) was incorporated across dA 41 and 28 times more efficiently than across dG and across inosine, respectively, by polη indicating that the mutagenicity of 5-FU incorporation is higher in the presence of inosine and that DNA lesions could lead to more mutagenic incorporation of 5-FU. Our polη crystal structures complexed with DNA and 5FdUTP revealed that dA:5FdUTP base pair is like dA:dTTP in the active site of polη, while 5FdUTP adopted 4-enol tautomer in the base pairs with dG and HX increasing the insertion efficiency compared to dG:dTTP for the incorrect insertions. These studies confirm that polη engages in the DNA incorporation and bypass of 5-FU.


Assuntos
Neoplasias Colorretais , DNA Polimerase Dirigida por DNA , Fluoruracila , Fluoruracila/farmacologia , DNA Polimerase Dirigida por DNA/metabolismo , DNA Polimerase Dirigida por DNA/genética , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Humanos , Dano ao DNA , DNA/metabolismo , DNA/química , DNA/biossíntese , Reparo do DNA , Nucleotídeos de Desoxiuracil/metabolismo , Nucleotídeos de Desoxiuracil/química , Antimetabólitos Antineoplásicos/farmacologia , Antimetabólitos Antineoplásicos/uso terapêutico , Antimetabólitos Antineoplásicos/química , Cinética , Replicação do DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Síntese de DNA Translesão
2.
Am J Respir Crit Care Med ; 200(9): 1113-1125, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31265321

RESUMO

Rationale: The loss of pulmonary endothelial cells in emphysema is associated with increased lung ceramide. Ceramide perturbations may cause adaptive alterations in other bioactive sphingolipids, with pathogenic implications. We previously reported a negative correlation between emphysema and circulating glycosphingolipids (GSLs). Glucosylceramide (GlcCer), the initial GSL synthesized from ceramide by GCS (GlcCer synthase), is required for embryonic survival, but its role in the lung is unknown.Objectives: To determine if cigarette smoke (CS) alters lung GlcCer and to elucidate the role of GCS in lung endothelial cell fate.Methods: GlcCer was measured by tandem mass spectrometry in BAL fluid of CS- or elastase-exposed mice, and GCS was detected by Western blotting in chronic obstructive pulmonary disease lungs and CS extract-exposed primary human lung microvascular endothelial cells (HLMVECs). The role of GlcCer and GCS on mTOR (mammalian target of rapamycin) signaling, autophagy, lysosomal function, and cell death were studied in HLMVECs with or without CS exposure.Measurements and Main Results: Mice exposed to chronic CS or to elastase, and patients with chronic obstructive pulmonary disease, exhibited significantly decreased lung GlcCer and GCS. In mice, lung GlcCer levels were negatively correlated with airspace size. GCS inhibition in HLMVEC increased lysosomal pH, suppressed mTOR signaling, and triggered autophagy with impaired lysosomal degradation and apoptosis, recapitulating CS effects. In turn, increasing GlcCer by GCS overexpression in HLMVEC improved autophagic flux and attenuated CS-induced apoptosis.Conclusions: Decreased GSL production in response to CS may be involved in emphysema pathogenesis, associated with autophagy with impaired lysosomal degradation and lung endothelial cell apoptosis.


Assuntos
Células Endoteliais/patologia , Glucosilceramidas/metabolismo , Enfisema Pulmonar/etiologia , Enfisema Pulmonar/metabolismo , Fumar/efeitos adversos , Animais , Autofagia , Técnicas de Cultura de Células , Morte Celular , Modelos Animais de Doenças , Camundongos , Enfisema Pulmonar/patologia
3.
Microvasc Res ; 118: 144-154, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29601874

RESUMO

The objective of this study was to isolate the impact of hydrodynamics on selectin-mediated cell rolling in branched microvessels. Significant advancements have been made in furthering the understanding of complex interactions between biochemical and physical factors in the inflammatory cascade in simplified planar geometries. However, few studies have sought to quantify the effects of branched configurations and to isolate the effects of associated fluid forces. Experimental techniques were developed to perform in vitro adhesion experiments in Y-shaped micro-slides. The micro-slides were coated with P-selectin and microspheres coated with Sialyl-Lewisx were observed as they rolled in the chambers at different wall shear stresses. Study results revealed that microsphere rolling velocities and rolling flux were lowest in regions closest to the apex of a junctional region and were dependent on both branch angle and wall shear stress. The regions closest to the junctional region were shown to have low bulk flow velocities and shear stresses using computational fluid dynamics (CFD) modeling. Collectively, the study demonstrates that despite the presence of a uniform coating of P-selectin, hydrodynamic factors associated with the chamber geometry yield non-uniform effects on particle behavior. These findings could explain why cells have been observed to preferentially adhere or transmigrate near junctional regions. Future characterization of inflammatory processes in microvascular network configurations is therefore crucial for furthering our fundamental understanding of inflammation.


Assuntos
Adesão Celular , Inflamação/metabolismo , Migração e Rolagem de Leucócitos , Leucócitos/metabolismo , Modelos Cardiovasculares , Selectina-P/metabolismo , Vênulas/metabolismo , Animais , Humanos , Hidrodinâmica , Inflamação/patologia , Antígenos CD15/metabolismo , Microesferas , Antígeno Sialil Lewis X , Transdução de Sinais , Vênulas/patologia
4.
Cureus ; 16(3): e56371, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38633963

RESUMO

In this case report, we present an unusual complication linked to a hydrogel explant in a 72-year-old male presenting to the emergency department with persistent left eye pain, redness, and discharge for one month. The patient had a history of retinal detachment in 1989, which was managed with scleral buckle surgery and gas injection. Initial examination revealed an extruding scleral buckle in the superior temporal region, along with signs of an infection. CT scans revealed a 1.9 × 1.2 × 3.8 cm abscess accompanied by preseptal cellulitis. This case report highlights the importance of how hydrogel scleral buckle explants may mimic the presentation and symptoms of an abscess as a long-term complication. Nevertheless, there have been several reports of long-term issues associated with the expansion of the hydrophilic hydrogel material. This case report further illustrates how complications linked to hydrogel explants can resemble abscess symptoms, underscoring the significance of accurate diagnosis and appropriate management.

5.
Clin Immunol ; 142(3): 237-42, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22284392

RESUMO

Cell signaling initiated by the B cell receptor is critical to normal development of B lymphocytes, most notably at the transitional B cell stage. Inhibition of this signaling pathway with the syk inhibitor, fostamatinib, has produced significant efficacy in lymphoid malignancies and autoimmune conditions. Here, we demonstrate that short-term use of fostamatinib impairs B lymphocyte development at the transitional stage without affecting mature B cell populations. Additionally, IL-10 producing B cells remained relatively constant throughout the treatment period. These findings provide insight into the mechanism of action of B cell receptor inhibition in autoimmune disease. As the development of agents targeting B cell receptor signaling proceeds, monitoring for long-term consequences as well as functional evaluation of B cell subsets may further improve our understanding of this rapidly growing class of novel agents.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Oxazinas/farmacologia , Células Precursoras de Linfócitos B/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Piridinas/farmacologia , Aminopiridinas , Humanos , Interleucina-10/biossíntese , Interleucina-10/imunologia , Morfolinas , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/imunologia , Pirimidinas , Quinase Syk
6.
Sci Transl Med ; 11(480)2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30787169

RESUMO

Skin barrier dysfunction has been reported in both atopic dermatitis (AD) and food allergy (FA). However, only one-third of patients with AD have FA. The purpose of this study was to use a minimally invasive skin tape strip sampling method and a multiomics approach to determine whether children with AD and FA (AD FA+) have stratum corneum (SC) abnormalities that distinguish them from AD without FA (AD FA-) and nonatopic (NA) controls. Transepidermal water loss was found to be increased in AD FA+. Filaggrin and the proportion of ω-hydroxy fatty acid sphingosine ceramide content in nonlesional skin of children with AD FA+ were substantially lower than in AD FA- and NA skin. These abnormalities correlated with morphologic changes in epidermal lamellar bilayer architecture responsible for barrier homeostasis. Shotgun metagenomic studies revealed that the nonlesional skin of AD FA+ had increased abundance of Staphylococcus aureus compared to NA. Increased expression of keratins 5, 14, and 16 indicative of hyperproliferative keratinocytes was observed in the SC of AD FA+. The skin transcriptome of AD FA+ had increased gene expression for dendritic cells and type 2 immune pathways. A network analysis revealed keratins 5, 14, and 16 were positively correlated with AD FA+, whereas filaggrin breakdown products were negatively correlated with AD FA+. These data suggest that the most superficial compartment of nonlesional skin in AD FA+ has unique properties associated with an immature skin barrier and type 2 immune activation.


Assuntos
Dermatite Atópica/diagnóstico , Hipersensibilidade Alimentar/diagnóstico , Pele/patologia , Adolescente , Área Sob a Curva , Criança , Pré-Escolar , Células Dendríticas/metabolismo , Dermatite Atópica/patologia , Epiderme/metabolismo , Proteínas Filagrinas , Hipersensibilidade Alimentar/patologia , Humanos , Proteínas de Filamentos Intermediários/metabolismo , Queratinas/metabolismo , Lipídeos/análise , Microbiota , Pele/microbiologia , Fita Cirúrgica , Transcriptoma/genética , Perda Insensível de Água
7.
JCI Insight ; 3(4)2018 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-29467325

RESUMO

Lipids in the stratum corneum of atopic dermatitis (AD) patients differ substantially in composition from healthy subjects. We hypothesized that hyperactivated type 2 immune response alters AD skin lipid metabolism. We have analyzed stratum corneum lipids from nonlesional and lesional skin of AD subjects and IL-13 skin-specific Tg mice. We also directly examined the effects of IL-4/IL-13 on human keratinocytes in vitro. Mass spectrometric analysis of lesional stratum corneum from AD subjects and IL-13 Tg mice revealed an increased proportion of short-chain (N-14:0 to N-24:0) NS ceramides, sphingomyelins, and 14:0-22:0 lysophosphatidylcholines (14:0-22:0 LPC) with a simultaneous decline in the proportion of corresponding long-chain species (N-26:0 to N-32:0 sphingolipids and 24:0-30:0 LPC) when compared with healthy controls. An increase in short-chain LPC species was also observed in nonlesional AD skin. Similar changes were observed in IL-4/IL-13-driven responses in Ca2+-differentiated human keratinocytes in vitro, all being blocked by STAT6 silencing with siRNA. RNA sequencing analysis performed on stratum corneum of AD as compared with healthy subjects identified decreased expression of fatty acid elongases ELOVL3 and ELOVL6 that contributed to observed changes in atopic skin lipids. IL-4/IL-13 also inhibited ELOVL3 and ELOVL6 expression in keratinocyte cultures in a STAT6-dependent manner. Downregulation of ELOVL3/ELOVL6 expression in keratinocytes by siRNA decreased the proportion of long-chain fatty acids globally and in sphingolipids. Thus, our data strongly support the pathogenic role of type 2 immune activation in AD skin lipid metabolism.


Assuntos
Acetiltransferases/metabolismo , Dermatite Atópica/patologia , Epiderme/patologia , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Metabolismo dos Lipídeos/imunologia , Acetiltransferases/genética , Adulto , Animais , Biópsia , Diferenciação Celular/imunologia , Linhagem Celular , Dermatite Atópica/genética , Dermatite Atópica/imunologia , Modelos Animais de Doenças , Epiderme/imunologia , Epiderme/metabolismo , Elongases de Ácidos Graxos , Feminino , Perfilação da Expressão Gênica , Humanos , Interleucina-13/genética , Interleucina-13/imunologia , Interleucina-4/imunologia , Queratinócitos/imunologia , Queratinócitos/metabolismo , Queratinócitos/patologia , Lipídeos/análise , Masculino , Camundongos , Camundongos Transgênicos , Cultura Primária de Células , RNA Interferente Pequeno/metabolismo , Análise de Sequência de RNA , Células Th2/imunologia , Células Th2/metabolismo
9.
Biochimie ; 95(5): 1056-61, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23168250

RESUMO

Contradictory results for concentrations of vitamin B12, holotranscobalamin (holoTC), and methylmalonic acid (MMA) have been reported. We tested the hypothesis that the extracellular vitamin B12 markers are not reflecting the intracellular vitamin B12-dependent biochemical reactions in individuals with type 2 diabetes. The study included 92 patients with diabetes and 72 controls with similar age and sex distribution. We measured vitamin B12 markers [MMA, total serum vitamin B12, holoTC, total homocysteine (tHcy)], red blood cell (RBC)-B12, and the plasma concentrations of the methylation markers [S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH)]. In comparison to controls, diabetic patients showed significantly higher concentrations of plasma SAH (median 15.1 vs. 11.8 nmol/L; p < 0.001) and lower SAM/SAH ratio (9.1 vs. 8.2; p = 0.006). Concentrations of total vitamin B12 and holoTC did not differ significantly between the groups, but plasma MMA concentrations were significantly higher in diabetics (250 vs. 206 nmol/L). However, RBC-B12 was lower in diabetics compared to controls (median 230 vs. 260 pmol/L; p = 0.001). The inverse correlation between MMA and RBC-B12 was stronger in the controls compared to that in the patients (correlation coefficient in controls R = -0.446, p = 0.001; in patients R = -0.289, p = 0.022). Metformin treatment was associated with a lower total serum vitamin B12, but a comparable RBC-B12 and a slightly lower MMA and better methylation index. In conclusion, patients with type 2 diabetes showed normal extracellular vitamin B12, but disturbed intracellular B12-dependent biochemical reactions. Metformin treatment was associated with low serum vitamin B12 and improved intracellular vitamin B12 metabolism despite low serum vitamin B12.


Assuntos
Diabetes Mellitus Tipo 2/sangue , Deficiência de Vitamina B 12/sangue , Vitamina B 12/sangue , Idoso , Idoso de 80 Anos ou mais , Diabetes Mellitus Tipo 2/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Vitamina B 12/metabolismo , Deficiência de Vitamina B 12/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA