Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Adv Biol (Weinh) ; 5(5): e2000199, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34028212

RESUMO

The regulation of cell-cell adhesions in space and time plays a crucial role in cell biology, especially in the coordination of multicellular behavior. Therefore, tools that allow for the modulation of cell-cell interactions with high precision are of great interest to a better understanding of their roles and building tissue-like structures. Herein, the green light-responsive protein CarH is expressed at the plasma membrane of cells as an artificial cell adhesion receptor, so that upon addition of its cofactor vitamin B12 specific cell-cell interactions form and lead to cell clustering in a concentration-dependent manner. Upon green light illumination, the CarH based cell-cell interactions disassemble and allow for their reversion with high spatiotemporal control. Moreover, these artificial cell-cell interactions impact cell migration, as observed in a wound-healing assay. When the cells interact with each other in the presence of vitamin B12 in the dark, the cells form on a solid front and migrate collectively; however, under green light illumination, individual cells migrate randomly out of the monolayer. Overall, the possibility of precisely controlling cell-cell interactions and regulating multicellular behavior is a potential pathway to gaining more insight into cell-cell interactions in biological processes.


Assuntos
Comunicação Celular , Vitamina B 12 , Adesão Celular , Luz
2.
ACS Synth Biol ; 9(8): 2076-2086, 2020 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-32610009

RESUMO

The self-assembly of different cell types into multicellular structures and their organization into spatiotemporally controlled patterns are both challenging and extremely powerful to understand how cells function within tissues and for bottom-up tissue engineering. Here, we not only independently control the self-assembly of two cell types into multicellular architectures with blue and red light, but also achieve their self-sorting into distinct assemblies. This required developing two cell types that form selective and homophilic cell-cell interactions either under blue or red light using photoswitchable proteins as artificial adhesion molecules. The interactions were individually triggerable with different colors of light, reversible in the dark, and provide noninvasive and temporal control over the cell-cell adhesions. In mixtures of the two cells, each cell type self-assembled independently upon orthogonal photoactivation, and cells sorted out into separate assemblies based on specific self-recognition. These self-sorted multicellular architectures provide us with a powerful tool for producing tissue-like structures from multiple cell types and investigate principles that govern them.


Assuntos
Adesão Celular/efeitos da radiação , Luz , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Linhagem Celular Tumoral , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Humanos , Cinética , Optogenética , Fotorreceptores Microbianos/genética , Fotorreceptores Microbianos/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo
3.
Nano Lett ; 20(4): 2257-2263, 2020 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-31751141

RESUMO

Building tissue from cells as the basic building block based on principles of self-assembly is a challenging and promising approach. Understanding how far principles of self-assembly and self-sorting known for colloidal particles apply to cells remains unanswered. In this study, we demonstrate that not just controlling the cell-cell interactions but also their dynamics is a crucial factor that determines the formed multicellular structure, using photoswitchable interactions between cells that are activated with blue light and reverse in the dark. Tuning dynamics of the cell-cell interactions by pulsed light activation results in multicellular architectures with different sizes and shapes. When the interactions between cells are dynamic, compact and round multicellular clusters under thermodynamic control form, while otherwise branched and loose aggregates under kinetic control assemble. These structures parallel what is known for colloidal assemblies under reaction- and diffusion-limited cluster aggregation, respectively. Similarly, dynamic interactions between cells are essential for cells to self-sort into distinct groups. Using four different cell types, which expressed two orthogonal cell-cell interaction pairs, the cells sorted into two separate assemblies. Bringing concepts of colloidal self-assembly to bottom-up tissue engineering provides a new theoretical framework and will help in the design of more predictable tissue-like structures.


Assuntos
Comunicação Celular , Engenharia Tecidual/métodos , Linhagem Celular Tumoral , Movimento Celular , Humanos , Luz , Optogenética , Processos Fotoquímicos , Termodinâmica
4.
Adv Biosyst ; 3(4): e1800310, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-32627428

RESUMO

Controlling cell-cell interactions is central for understanding key cellular processes and bottom-up tissue assembly from single cells. The challenge is to control cell-cell interactions dynamically and reversibly with high spatiotemporal precision noninvasively and sustainably. In this study, cell-cell interactions are controlled with visible light using an optogenetic approach by expressing the blue light switchable proteins CRY2 or CIBN on the surfaces of cells. CRY2 and CIBN expressing cells form specific heterophilic interactions under blue light providing precise control in space and time. Further, these interactions are reversible in the dark and can be repeatedly and dynamically switched on and off. Unlike previous approaches, these genetically encoded proteins allow for long-term expression of the interaction domains and respond to nontoxic low intensity blue light. In addition, these interactions are suitable to assemble cells into 3D multicellular architectures. Overall, this approach captures the dynamic and reversible nature of cell-cell interactions and controls them noninvasively and sustainably both in space and time. This provides a new way of studying cell-cell interactions and assembling cellular building blocks into tissues with unmatched flexibility.


Assuntos
Comunicação Celular , Optogenética/métodos , Engenharia Tecidual/métodos , Comunicação Celular/genética , Comunicação Celular/fisiologia , Comunicação Celular/efeitos da radiação , Linhagem Celular Tumoral , Criptocromos/genética , Criptocromos/metabolismo , Humanos , Luz
5.
Iran Biomed J ; 22(3): 193-201, 2017 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-28941453

RESUMO

Background: Artificial oligonucleotides like DNA or RNA aptamers can be used as biodiagnostic alternatives for antibodies to detect pathogens. Comparing to antibodies, artificial oligonucleotides are produced easily at lower costs and are more stable. Neisseria meningitidis, the causative agent of meningitis, is responsible for about 1% of infections in an epidemic period. Specific DNA aptamers that bind to N. meningitidis serogroup B were identified by whole-cell Systemic Evolution of Ligands by EXponential Enrichment (SELEX). Methods: The SELEX begins with a library of labeled ssDNA molecules. After six rounds of selection and two rounds of counter-selection, 60 clones were obtained, of which the binding efficiency of 21 aptamers to the aforementioned bacterium was tested by flow cytometry. Results: The aptamers K3 and K4 showed the highest affinity to N. meningitidis serogroup B and no affinity to N. meningitidis serogroups Y, A, and C, or to other meningitis causing bacteria. The dissociation constant (Kd value) for K3 and K4 were calculated as 28.3±8.9 pM and 39.1±8.6 pM, respectively. K3 aptamer with the lowest Kd was chosen as the main aptamer. K3 could detect N. meningitidis in patients' cerebrospinal fluid (CSF) samples and in CSF from healthy volunteers inoculated with N. meningitidis serogroup B (ATCC 13090) at 200 and 100 CFU ml-1, respectively. Conclusion: The findings suggest the application of the developed aptamer in specific detection of N. meningitidis serogroup B amongst a group of meningitis causing bacteria.

6.
Can J Microbiol ; 63(2): 160-168, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28121169

RESUMO

Group A streptococcus (GAS) is an important Gram-positive pathogen that causes various human diseases ranging from peripheral lesions to invasive infections. The M protein is one of the main virulence factors present on the cell surface and is associated with invasive GAS infections. Compared with other M types, serotype M3 has a predominant role in lethal infections and demonstrates epidemic behaviors, including streptococcal toxic shock syndrome, bacteremia, and necrotizing fasciitis. Traditional methods for M typing are time-consuming, tedious, contradictory, and generally restricted to reference laboratories. Therefore, development of a new M-typing technique is needed. Aptamers with the ability to detect their target with a high degree of accuracy and specificity can be ideal candidates for specific M-typing of Streptococcus pyogenes. In this study DNA aptamers with a high binding affinity towards S. pyogenes serotype M3 were selected through 12 iterative rounds of the Systematic Evolution of Ligands by EXponential (SELEX) enrichment procedure using live cells as a target. We monitored the progress of the SELEX procedure by flow cytometry analysis. Of several aptamer sequences analyzed, 12L18A showed the highest binding efficiency towards S. pyogenes type M3, with an apparent dissociation constant (Kd) of 7.47 ± 1.72 pmol/L being the lowest. Therefore the isolated aptamer can be used in any tool, such as a biosensor, for the detection of S. pyogenes and can be used in the development of a novel M-typing system.


Assuntos
Aptâmeros de Nucleotídeos/metabolismo , Técnica de Seleção de Aptâmeros , Streptococcus pyogenes/isolamento & purificação , Citometria de Fluxo , Humanos , Sorogrupo , Streptococcus pyogenes/classificação
7.
Avicenna J Med Biotechnol ; 8(3): 104-11, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27563422

RESUMO

BACKGROUND: Nowadays, highly specific aptamers generated by cell SELEX technology (systematic evolution of ligands by exponential enrichment) are being applied for early detection of cancer cells. Prostate Specific Membrane Antigen (PSMA), over expressed in prostate cancer, is a highly specific marker and therefore can be used for diagnosis of the prostate cancer cells. The aim of the present study was to select single-stranded DNA aptamers against LNCap cells highly expressing PSMA, using cell-SELEX method which can be used as a diagnostic tool for the detection of prostate cancer cells. METHODS: After 10 rounds of cell-SELEX, DNA aptamers were isolated against PSMA using LNCaP cells as a target and PC-3 cell lines for counter SELEX. Five DNA aptamers with more than 70% affinity were selected up on flow cytometry analysis of positive clones. RESULTS: Dissociation constants of two selected sequences (A12-B1) were estimated in the range of 33.78±3.77 and 57.49±2.214 pmol, respectively. Conserved secondary structures of A12 and B1 sequences suggest the necessity of these structures for binding with high affinity to native PSMA. Comparison of the secondary structures of our isolated aptamers and aptamer A10 obtained by protein SELEX showed similar stem-loop structures which could be responsible for the recognition of PSMA on LNCap cell surface. CONCLUSION: Our results indicated that selected aptamers may turn out to be ideal candidates for the development of a detection tool and also can be used in targeted drug delivery for future smart drugs.

8.
J Biotechnol ; 231: 46-54, 2016 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-27234880

RESUMO

Acinetobacter baumannii has turned into an important threat in nosocomial outbreak infections and multidrug resistance leading to high mortality rates in the 21st century. In recent years its mortality has increased by 15% which in part could be due to lack of a rapid and sensitive diagnostic test. In this work we introduced a new detection test for A. baumannii with two highly specific aptamer and nanobody molecules. High binding affinity DNA oligonucleotide aptamers toward A. baumannii were selected through 12 rounds of whole cell System Evolution of Ligands by EXponential enrichment process (SELEX). The SELEX procedures was monitored by flow cytometry. The dissociation constant and binding efficiency of the selected aptamer Aci49 was 7.547±1:353pM and 47.50%, respectively. A sandwich enzyme linked aptamer sorbent assay (ELASA) was designed with the biotinylated Aci49 aptamer and our previously developed nanobody against biofilm associated protein (Bap). The assay system was optimized with A. baumannii (ATCC 19606) and 47 clinical isolates of A. baumannii were tested. The threshold of detection in sandwich ELASA process was10(3) CFU/ml. The sensitivity of test toward the clinical isolates was 95.47%. Our results reveal that the sandwich ELASA is sensitive and specific enough for the rapid detection of A. baumannii from clinical isolates.


Assuntos
Acinetobacter baumannii/genética , Acinetobacter baumannii/isolamento & purificação , Aptâmeros de Nucleotídeos/genética , DNA Bacteriano/genética , Técnica de Seleção de Aptâmeros/métodos , Infecções por Acinetobacter/diagnóstico , Infecções por Acinetobacter/microbiologia , Aptâmeros de Nucleotídeos/química , DNA Bacteriano/análise , Citometria de Fluxo , Humanos , Limite de Detecção
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...