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1.
Micromachines (Basel) ; 10(12)2019 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-31835793

RESUMO

The clinical characteristics of excreted tumor cells can be found in the urine of bladder cancer patients, meaning the identification of tumor cells in urine can assist in bladder cancer diagnosis. The presence of white blood cells and epithelial cells in the urine interferes with the recognition of tumor cells. In this paper, a technique for detecting cancer cells in urine based on microfluidics provides a novel approach to bladder cancer diagnosis. The bladder cancer cell line (T24) and MeT-5A were used as positive bladder tumor cells and non-tumor cells, respectively. The practicality of the tumor cell detection system based on microfluidic cell chip detection technology is discussed. The tumor cell (T24) concentration was around 1 × 104 to 300 × 104 cells/mL. When phosphate buffer saline (PBS) was the diluted solution, the tumor cell detected rate was 63-71% and the detection of tumor cell number stability (coefficient of variation, CV%) was 6.7-4.1%, while when urine was the diluted solution, the tumor cell detected rate was 64-72% and the detection of tumor cell number stability (CV%) was 6.3-3.9%. In addition, both PBS and urine are tumor cell dilution fluid solutions. The sample was analyzed at a speed of 750 microns per hour. Based on the above experiments, a system for detecting bladder cancer cells in urine by microfluidic analysis chip technology was reported. The rate of recognizing bladder cancer cells reached 68.4%, and the speed reached 2 mL/h.

2.
Inflamm Res ; 68(7): 597-611, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31119302

RESUMO

OBJECTIVE: The present study was undertaken to validate whether TNF-α and calreticulin (CRT) serve as dual signaling to activate nucleotide-binding oligomerization domain-, leucine-rich repeat- and pyrin domain-containing 3 (NLRP3) inflammasome in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) and HUVECs. The effect of human antigen R (HuR) in NLRP3 inflammasome activation was also explored in RA FLS. METHODS: Immunofluorescence was used to determine the expression of NLRP3 and adaptor protein apoptosis associated speck-like protein containing a CARD (ASC) in RA synovial tissue and HuR location in RA FLS. Western blot and quantitative real-time PCR were employed to measure the priming effect of NLRP3 inflammasome in cells and HuR expression in synovial tissue. The concentrations of IL-1ß and IL-18 were detected by enzyme linked immunosorbent assay. Immunohistochemistry was used to visualize the expression of HuR in synovial tissue. HuR knockdown in RA FLS was achieved by siRNA-mediated gene silencing. RESULTS: Higher expression of NLRP3 and ASC in RA synovial tissue than those in osteoarthritis was detected. The staining of NLRP3, ASC and cleaved IL-1ß were observed in FLS and vascular endothelial cells in RA synovium. Expression of NLRP3 and pro-IL-1ß in RA FLS and HUVECs treated with TNF-α was increased. The pro-IL-18 expression was also enhanced in HUVECs, but not in RA FLS. TNF-α/CRT dual stimulation of cells gave rise to caspase-1 p20 expression and the secretion of IL-1ß. The secreted IL-18 was also elevated in HUVECs but not in RA FLS. HuR expression was significantly elevated in RA synovial tissue. TNF-α initiated the nucleocytoplasmic shuttling of HuR in both FLS and HUVECs. The knockdown of HuR in FLS incubated with TNF-α led to reduced caspase-1 p20 protein expression and further resulted in decreased secretion of IL-1ß in the presence of CRT. CONCLUSIONS: TNF-α/CRT dual signaling induced NLRP3 inflammasome activation, which could be suppressed by HuR knockdown presumably due to the block of HuR translocating from nucleus to cytoplasma.


Assuntos
Artrite Reumatoide/imunologia , Calreticulina/imunologia , Proteína Semelhante a ELAV 1/imunologia , Inflamassomos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Fator de Necrose Tumoral alfa/imunologia , Células Endoteliais da Veia Umbilical Humana/imunologia , Humanos , Transdução de Sinais , Membrana Sinovial/imunologia , Sinoviócitos/imunologia
3.
Oncol Lett ; 17(5): 4532-4544, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30944642

RESUMO

The detection of tumor cells and clusters in pleural effusion assists in the diagnosis of lung cancer. The proportion of tumor cells and clusters to the total number of cells in each patient varies substantially due to individual differences and the severity of the disease. The identification of one tumor cell or cluster from a large number of pleural effusions is the main challenge for hydrothorax tumor cell detection techniques. In the present study, by using A549 lung cancer and Met-5A mesothelial cell lines, a label-free microfluidic chip based on cell cluster size was designed. By setting the parameters of the chip, individual cells and clusters were able to enter different microfluidic channels. Subsequent to non-specific staining, the recovered components were stained using acridine orange (AO). A charge-coupled device camera was used to captured images of the cell, and the features of these cells were analyzed in their R and G channels using Matlab software to establish the characteristics and finally differentiate between the tumor and non-tumor cell or clusters. According to the results, when inlet A and B were under a velocity of 10 and 8.5 ml/h, respectively, the tumor cell clusters were successfully collected through microfluidic channels III-V, with a recovery rate of ~80%. Subsequent to staining with AO, the feature values in the R and G channels were identified, and initial differentiation was achieved. The present study combined the microfluidic chip, which is based on cluster size, with a computer identification method for pleural effusion. The successful differentiation of tumor cell clusters from non-tumor clusters provides the basis for the identification of tumor clusters in hydrothorax.

4.
Mol Immunol ; 107: 10-20, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30639474

RESUMO

The present study was undertaken to determine whether extracellular calreticulin (CRT) participates in the regulation of ICAM-1in rheumatoid arthritis (RA) and further explore the potential mechanism. Our results showed that ICAM-1 and VCAM-1 levels were positively correlated with CRT levels in RA serum and synovial fluid, respectively. In RA synovial tissue, increased co-expressions of CRT and ICAM-1 in vascular endothelium and perivascular areas and elevated co-location of CRT and VCAM-1 localized predominantly to lining layer were observed compared to those in OA. In in vitro HUVECs model, enhanced ICAM-1expression and increased phosphorylation levels of Akt and eNOS were detected in the presence of CRT. Increased phosphorylated eNOS was significantly inhibited by a PI3K inhibitor LY294002 and elevated ICAM-1expression was partially blocked by the inhibitors of both PI3K and eNOS (L-NAME). It has been certified that the RNA-binding protein TTP targets AU-rich elements in the ICAM-1 3'-UTR and suppresses ICAM-1 expression. Knocking down TTP in HUVECs led to an increased induction of ICAM-1 by CRT. We have currently known that activation of p38 downstream kinase MK-2 leads to phosphorylation and inactivation of human TTP. The block of p38 MAPK/MK-2 signaling led to decreased protein expression and mRNA stability of TTP and ICAM-1. Furthermore, L-NAME and/or LY294002 pre-treated HUVECs manifested decreased p38 and MK-2 phosphorylation, which was accompanied by reduced TTP and ICAM-1 protein expression as well as decreased mRNA stability. Our results suggested that CRT could promote ICAM-1 expression in endothelial cells through PI3K/Akt/eNOS/p38 MAPK signaling mediated TTP accumulation, probably in an inactive form, which may provide a possible proinflammatory mechanism of CRT in RA.


Assuntos
Artrite Reumatoide/imunologia , Calreticulina/imunologia , Células Endoteliais da Veia Umbilical Humana/imunologia , Molécula 1 de Adesão Intercelular/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Tristetraprolina/imunologia , Regulação para Cima/imunologia , Artrite Reumatoide/patologia , Cromonas/farmacologia , Feminino , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Morfolinas/farmacologia , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico Sintase Tipo III/antagonistas & inibidores , Óxido Nítrico Sintase Tipo III/imunologia , Fosfatidilinositol 3-Quinases/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Membrana Sinovial/imunologia , Membrana Sinovial/patologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia
5.
Biomed Pharmacother ; 110: 677-684, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30553194

RESUMO

Cervical spondylosis (CS), which is resulted from degeneration of cervical intervertebral disc, is a common disease seriously threatening human health and quality of life. However, there is still no effective clinic strategies for the treatment of this disease. The acupoint stimulation with needle-scalpel is a widely used approach to treat orthopedic diseases. In the present study, we evaluated the therapeutic effects of acupoint stimulation around neck with needle-scalpel on delaying the degeneration of cervical intervertebral discs and hopefully provided an approach for the precaution and early intervention of CS. We firstly established a rat model of CS by cervical static-dynamic imbalance to mimics disc degeneration and then stimulated the acupoints around neck with needle-scalpel. The cervical intervertebral disc samples were collected to measure type I and II collagen by quantitative PCR (qPCR), immunohistochemistry, and western blot. The changes in micro-structure and ultra-structure of nucleus pulposus were analyzed under the optical microscope and electron microscope respectively. Acupoint stimulation with needle-scapelon increased type I collagen production and decreased type II collagen production, and improved the micro-structure and ultra-structure of nucleus pulposus. Our results suggest that acupoint stimulation around neck with needle-scapelon could inhibit intervertebral disc degeneration through modulating the extracellular matrix collagen system and improving the changed structure of nucleus pulposus.


Assuntos
Pontos de Acupuntura , Vértebras Cervicais , Degeneração do Disco Intervertebral/metabolismo , Degeneração do Disco Intervertebral/terapia , Agulhas , Núcleo Pulposo/metabolismo , Animais , Colágeno Tipo I/metabolismo , Colágeno Tipo II/metabolismo , Feminino , Degeneração do Disco Intervertebral/patologia , Núcleo Pulposo/patologia , Núcleo Pulposo/ultraestrutura , Ratos , Ratos Sprague-Dawley
6.
J Cell Physiol ; 234(1): 837-848, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-30078183

RESUMO

Diabetes mellitus (DM) comprises a group of metabolic diseases characterized by insulin deficiency or resistance and hyperglycemia. We previously reported the presence of abnormal differentiation of small intestinal epithelial cells (IECs) in diabetic mice, but the exact mechanism of this phenomenon has not been thoroughly elucidated to date. In this study, we found that H19 was markedly upregulated in IECs of DM mice. H19 knockdown significantly inhibited abnormal differentiation of IECs in DM mice. Bioinformatics analysis identified miR-141-3p as a candidate for H19. Based on luciferase reporter assays, we found that miR-141-3p directly targeted H19. Luciferase reporter assays also showed that miR-141-3p could directly target ß-catenin. Furthermore, H19 might act as an endogenous "sponge" by competing for miR-141-3p binding to regulate miRNA targets in vitro and in vivo. In summary, our findings provide the first evidence supporting the role of H19 in IECs of DM mice, and miR-141-3p targets not only protein-coding genes but also the lncRNA H19.


Assuntos
Diabetes Mellitus/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , beta Catenina/genética , Animais , Diferenciação Celular/genética , Diabetes Mellitus/metabolismo , Diabetes Mellitus/patologia , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Regulação da Expressão Gênica no Desenvolvimento/genética , Técnicas de Silenciamento de Genes , Humanos , Hiperglicemia/genética , Hiperglicemia/patologia , Resistência à Insulina/genética , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Camundongos , Camundongos Endogâmicos NOD , Ligação Proteica
7.
Clin Exp Rheumatol ; 36(5): 841-849, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29652658

RESUMO

OBJECTIVES: Fibroblast-like synoviocytes (FLS) play key roles in synovium hyperplasia and pannus formation in rheumatoid arthritis (RA). The present study was undertaken to explore the mechanisms that calreticulin (CRT) promoted anti-apoptosis of RA FLS. METHODS: The expression of CRT and anti-apoptotic proteins Bcl-XL and Mcl-1 in RA synovium were detected by immunohistochemistry. The expression of Bcl-XL and Mcl-1 in RA FLS by CRT were determined. The phosphorylation of Akt and STAT3 was detected by western blot. The effect of CRT on proliferation of RA FLS was examined by MTT assay. The ability of CRT to inhibit RA FLS apoptosis was assessed by flow cytometry. RESULTS: Increased expressions of CRT, Bcl-XL and Mcl-1 were detected in RA synovium compared with osteoarthritis (OA). Moreover, CRT expression correlated positively with Bcl-XL and Mcl-1 in RA, respectively. In vitro, CRT induced upregulation of Bcl-XL and Mcl-1 protein levels in RA FLS, in dose/time dependent manners. Upregulated expression of Bcl-XL and Mcl-1 induced by CRT were inhibited by PI3K/Akt or STAT3 pathways inhibitors in RA FLS, respectively. The increased phosphorylation levels of Akt and STAT3 were also detected with CRT incubation, in dose/time dependent manners. Additionally, CRT rescued apoptosis of RA FLS mediated by FasL. CONCLUSIONS: This study showed that upregulation of Bcl-XL and Mcl-1 expression in RA FLS by CRT were PI3K/Akt and STAT3 signal pathways dependent, and promoted the anti-apoptosis of RA FLS. Therefore, this may represent a therapeutic target for the treatment of RA.


Assuntos
Apoptose/efeitos dos fármacos , Artrite Reumatoide/tratamento farmacológico , Calreticulina/farmacologia , Fibroblastos/efeitos dos fármacos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Membrana Sinovial/efeitos dos fármacos , Sinoviócitos/efeitos dos fármacos , Proteína bcl-X/metabolismo , Artrite Reumatoide/enzimologia , Artrite Reumatoide/patologia , Calreticulina/metabolismo , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Fibroblastos/enzimologia , Fibroblastos/patologia , Humanos , Fosforilação , Membrana Sinovial/enzimologia , Membrana Sinovial/patologia , Sinoviócitos/enzimologia , Sinoviócitos/patologia , Regulação para Cima
8.
Clin Biochem ; 52: 167-170, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29129626

RESUMO

OBJECTIVE: To explore a panel of serum biomarkers for laboratory diagnosis of pediatric Henoch-Schönlein purpura (HSP). METHODS: The blood white blood cells (WBC) and serum levels of serum amyloid A (SAA), interleukin 6 (IL-6), immunoglobulin A (IgA), immunoglobulin G (IgG), immunoglobulin M (IgM), immunoglobulin E (IgE), C-reactive protein (CRP), complement component 3 (C3), complement component 4 (C4), and ASO (anti-streptolysin O) were detected in 127 patients with Henoch-Schonlein purpura (HSP), 110 cases of septicemia patients, and 121 healthy volunteers. The diagnostic ability of biomarkers selected from HSP and septicemia patients was analyzed by ROC curve. By designing the calculation model, the biomarker index was calculated for laboratory diagnosis of HSP and differential diagnosis between HSP and septicemia. RESULTS: The levels of serum WBC, CRP, IL-6 and SAA in the septicemia patients were significantly higher than those in the control group (p<0.05). Compared with the healthy individuals, serum levels of WBC, CRP, IL-6, SAA, IgA and IgM were significantly increased in patients with HSP (p<0.05). The area under the curve (AUC) of SAA, IgA, IgM, WBC, IL-6, and CRP in the patients with HSP was 0.964, 0.855, 0.849, 0.787, 0.765, and 0.622, respectively. The values of SAA, IgA, IgM, WBC, IL-6, and CRP in septicemia patients were 0.700, 0.428, 0.689, 0.682, 0.891, and 0.853, respectively. Biomarker index=SAA+IgA/4000+IgM/4000×0.4CRPmean valueCRPi. The biomarker index in HSP patients was significantly higher than that of the healthy controls. However, the biomarker index in septicemia patients was significantly lower than the control. CONCLUSION: The biomarker index of HSP patients is higher than that of the control group. While in the infectious disease represented by septicemia, it is decreased. The detection of biomarker index could exclude the interference of infection as the auxiliary examination to HSP patients.


Assuntos
Púrpura de Schoenlein-Henoch/diagnóstico , Adolescente , Proteínas de Bactérias/análise , Proteínas de Bactérias/sangue , Biomarcadores/sangue , Proteína C-Reativa/análise , Criança , Pré-Escolar , China , Complemento C3/análise , Complemento C4/análise , Feminino , Humanos , Imunoglobulina A/análise , Imunoglobulina A/sangue , Imunoglobulina E/análise , Imunoglobulina E/sangue , Imunoglobulina G/análise , Imunoglobulina G/sangue , Interleucina-6/análise , Interleucina-6/sangue , Masculino , Púrpura de Schoenlein-Henoch/sangue , Sepse/sangue , Sepse/diagnóstico , Proteína Amiloide A Sérica/análise , Estreptolisinas/análise , Estreptolisinas/sangue
9.
Biomicrofluidics ; 12(6): 064106, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30867867

RESUMO

Detecting the number of pathological lymphoma cells and lymphocyte subtypes in blood is helpful for clinical diagnosis and typing of lymphoma. In the current study, cell type is identified by cell morphological features and immunolabeled lymphocyte subtypes. Red blood cells and leukocytes were separated using a microfluidic cell chip based on physical blood cell parameters, and leukocytes were identified using five characteristic parameters: energy variance, entropy variance, moment of inertia variance, color mean, and cell area individually. The number of red blood cells that could come into contact with the leukocyte membrane was ≤2 based on the microfluidic injection flow rate of microfluidic chips. Anti-CD3 and anti-CD19 antibodies were used for immunofluorescence staining of T-lymphocyte and B-lymphocyte surface antigens, respectively. The results suggested that the microfluidic assay could detect lymphocyte surface antigen markers and intact leukocytes. Therefore, we report a one-step microfluidic chip for classifying hematological lymphoma cells based on the physical parameters of cells, which can simultaneously measure the overall morphology of blood cells and immunolabeling of lymphocyte surface antigens in one step, solving the current problem of detecting subtypes of hematological lymphoma cells based on multiple methods and multi-step detection.

10.
J Neurol Sci ; 383: 11-17, 2017 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-29246595

RESUMO

Cerebral amyloid angiopathy (CAA) is characterized by cerebrovascular amyloid deposition. It contributes to the rate of cognitive decline in older individuals and is present in >90% of patients with Alzheimer's disease (AD), with no cure so far. Molecular modifications during CAA should be elucidated to improve its diagnosis and treatment. In this study, amyloid-ß (Aß) aggregates in platelet membranes from 65 patients with CAA and 66 healthy volunteers (controls) were confirmed through thioflavin T (ThT) assay and Western blot analysis. Further, post-translational modifications (PTMs) of Aß in platelet membranes were analyzed using ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry (UPLC/ESI-Q-TOF/MS). ThT assay results indicated that there were amyloid components in platelets from both patients with CAA and controls. Western blot analysis showed that different molecular weight (MW) Aß aggregates were found in platelet membranes. LC-MS analysis showed that PTMs including methylation, phosphopantetheine, phosphorylation, deamidation, and acetylation, occurred in Aß peptide in platelet membranes from both patients with CAA and controls, while Met35 oxidation (MetOX) and Gln15 deamidation were identified only from patients with CAA. Thus, this study identified potential biomarkers of CAA and characterized the mechanism underlying amyloidogenesis in CAA.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Plaquetas/metabolismo , Membrana Celular/metabolismo , Angiopatia Amiloide Cerebral/metabolismo , Processamento de Proteína Pós-Traducional , Acetilação , Idoso , Peptídeos beta-Amiloides/química , Encéfalo/diagnóstico por imagem , Angiopatia Amiloide Cerebral/diagnóstico por imagem , Feminino , Humanos , Imagem por Ressonância Magnética , Masculino , Metilação , Pessoa de Meia-Idade , Peso Molecular , Oxirredução , Fosforilação , Agregação Patológica de Proteínas/metabolismo
11.
Biomed Eng Online ; 15: 14, 2016 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-26847685

RESUMO

BACKGROUND: Cervical cancer is the second leading cause of female-specific cancer-related deaths after breast cancer, especially in developing countries. However, the incidence of the disease may be significantly decreased if the patient is diagnosed in the pre-cancerous lesion stage or earlier. In recent years, computer-based algorithms are widely used in cervical cancer screening. Most of the proposed algorithms follow the procedure of segmentation, feature extraction, and then classification. Nevertheless, few of the existing segmentation methods are as flexible and robust as the human visual system, and the complexity of the algorithms makes it difficult for clinical application. METHODS: In this study, a computer-assisted analytical approach is proposed to identify the existence of suspicious cells in a whole slide cervical cell image (WSCCI). The main difference between our method and the conventional algorithm is that the image is divided into blocks with certain size instead of segmented cells, which can greatly reduce the computational complexity. Via data analysis, some texture and color histogram features show significant differences between blocks with and without suspicious cells. Therefore these features can be used as the input of the support vector machine classifier. 1100 non-background blocks (110 suspicious blocks) are trained to build a model, while 1040 blocks (491 non-background blocks) from 12 other WSCCIs are tested to verify the feasibility of the algorithm. RESULTS: The experimental results show that the accuracy of our method is about 98.98 %. More importantly, the sensitivity, which is more fatal in cancer screening, is 95.0 % according to the images tested in the study, while the specificity is 99.33 %. CONCLUSION: The analysis of the algorithm is based on block images, which is different from conventional methods. Although some analysis work should be done in advance, the later processing speed will be greatly enhanced with the establishment of the model. Furthermore, since the algorithm is based on the actual WSCCI, the method will be of directive significance for clinical screening.


Assuntos
Colo do Útero/patologia , Diagnóstico por Computador/métodos , Processamento de Imagem Assistida por Computador/métodos , Programas de Rastreamento , Imagem Molecular , Neoplasias do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/patologia , Automação , Feminino , Humanos , Máquina de Vetores de Suporte
12.
Zhen Ci Yan Jiu ; 40(5): 352-7, 2015 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-26669189

RESUMO

OBJECTIVE: To observe the effect of acupotomy lysis at acupoints around the neck on expression of matrix metalloproteinase 1 (MMP-1), MMP-2 and tissue inhibitor of metalloproteinase 1 (TIMP-1) genes and ultrastructure of pulpiform nucleus in cervical intervertebral disc degeneration (IVDD) rats, so as to explore its mechanism underlying easing IVDD. METHODS: SD rats were randomly allocated to control (n = 15), model (n = 14), Jiaji (EX-B 2, n = 13), cervico-acupoint (n = 14) and medication groups (n = 14). The cervical IVDD model was established by using static-dynamic imbalance method. For rats of the Jiaji (EX-B 2) and cervico-acupoint groups, EX-B 2-points of the cervical 2-7 segments, and peri-cervical acupoints: bilateral "Naokong" (GB 19) , "Naohu" (GV 17), "Dazhui" (GV 14), bilateral "Quyuan" (SI 13) and bilateral "Tianzong" (SI 11) were separately punctured with a needle-knife, once every 5 days for 3 times, and for rats of the medication group, Brufen Capsules (15 mg · kg(-1) · d(-1)) and Jingfukang Granule (0.5 mg · kg(-1) · d(-1)) were given by intragastric administration, once daily for 10 days. The expression levels of MMP-1, MMP-3 and TIMP-1 genes in the pulpiform nucleus of cervical intervertebral discs were detected by RT-PCR and changes of the ultrastructure of the pulpiform nucleus observed under transmission electron microscope. RESULTS: Compared to the control group, the expression levels of MMP-1 mRNA and MMP-3 mRNA of the cervical intervertebral disc tissues were significantly up-regulated in the model group (P < 0.05), and that of TIMP-1 mRNA was obviously down-regulated in the model group (P < 0.05). After the treatment, the increased expression of MMP-1 mRNA and MMP-3 mRNA and the decreased expression of TIMP-1 mRNA were reversed by acupotomy lysis and medication (P < 0.05) except TIMP-1 mRNA in the medication group (P > 0.05). No significant differences were found between the Jiaji (EX-B 2) and cervico-acupoint groups in down-regulating MMP-1 mRNA and MMP-3 mRNA expression and up-regulating TIMP-1 mRNA expression (P > 0.05). Results of electron microscope examinations showed that the ultrastructural injury changes of cells of the pulpiform nucleus were relatively milder in the Jiaji (EX-B 2) and cervico-acupoint groups, followed by the medication group in comparison with those of the model group. CONCLUSION: Acupotomy lysis at acupoints around the neck can improve the ultrastructural changes of cells of the pulpiform nucleus of cervical intervertebral discs in IVDD rats, which is possibly by regulating the expression of MMP-1, MMP-3 and TIMP-1 genes.


Assuntos
Pontos de Acupuntura , Terapia por Acupuntura , Degeneração do Disco Intervertebral/metabolismo , Degeneração do Disco Intervertebral/terapia , Metaloproteinase 1 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Animais , Vértebras Cervicais/metabolismo , Vértebras Cervicais/ultraestrutura , Humanos , Degeneração do Disco Intervertebral/enzimologia , Degeneração do Disco Intervertebral/genética , Masculino , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 3 da Matriz/genética , Ratos , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/genética
13.
Zhongguo Dang Dai Er Ke Za Zhi ; 17(9): 918-21, 2015 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-26412170

RESUMO

OBJECTIVE: To screen biomarkers which can be used as an auxiliary method in the diagnosis of Henoch-Schönlein purpura (HSP) and to evaluate their diagnostic values by receiver operating characteristic (ROC) curve analysis. METHODS: A total of 127 children diagnosed with HSP between April 2012 and March 2014 were included in the HSP group and an equal number of healthy children were included in the control group. Twelve parameters, i.e., serum amyloid protein A (SAA), interleukin-6 (IL-6), immunoglobulins (IgA, IgG, IgM, and IgE), C-reactive protein (CRP), white blood cell (WBC) count, complements C3 and C4, anti-streptolysin O, and ferritin, were analyzed. The values of the screened biomarkers for diagnosis of HSP were assessed by ROC curve analysis. RESULTS: The HSP group had significantly higher levels of SAA, IL-6, CRP, WBC, IgA, and IgM than the control group (P<0.05). The areas under the ROC curve of SAA, IL-6, WBC, IgA, and IgM for the diagnosis of HSP were higher than 0.7 (P<0.05). The optimal cut-off values of SAA, IgA, IgM, WBC, and IL-6 for the diagnosis of HSP were 3.035 µg/mL, 1579.5 mg/L, 922.5 mg/L, 8.850 × 109/L, and 7.035 pg/mL, respectively; the corresponding sensitivities of the optimal cut-off values for the diagnosis of HSP were 95.1%, 75.6%, 72.3%, 78.0%, and 63.4%, respectively, and the corresponding specificities were 90.2%, 85.4%, 82.4%, 70.7%, and 80.5%, respectively. CONCLUSIONS: SAA, IgA, IgM, WBC, and IL-6 are valuable biomarkers for clinical diagnosis of HSP and among them SAA seems to be the best one.


Assuntos
Púrpura de Schoenlein-Henoch/diagnóstico , Adolescente , Biomarcadores/sangue , Proteína C-Reativa/análise , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Púrpura de Schoenlein-Henoch/sangue , Curva ROC , Proteína Amiloide A Sérica/análise
14.
Age Ageing ; 44(3): 458-64, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25673873

RESUMO

BACKGROUND: in vitro, it has been reported that amyloid ß (Aß) is bound to red blood cells (RBCs) and this process damages the red cell. Also, a possible relationship between RBCs and Alzheimer's disease (AD) is supported by the findings of RBC impairment in AD. Therefore, Aß fibrils bounding RBC are of great interest as potential biomarkers. METHODS: in this study, we focused on Aß amyloid fibrils and/or aggregation on the peripheral RBC from 50 subjects with AD and 50 healthy controls (HCs) through thioflavin T (ThT) staining followed by immunofluorescence assay to confirm the presence of Aß amyloid fibrils and/or aggregation on the RBC. Then we optimised fluorescence staining and imaging conditions and analysed the images obtained by image processing software. RESULTS: we have analysed RBC morphology in blood from 50 subjects with AD and 50 HCs found that 16.8% of the RBCs are elongated as compared with 6.7% in normal controls (P < 0.01), and there is a negative correlation between the two parameters (P < 0.05). Our study showed that 98% of AD peripheral RBCs were amyloid binding-positive (ranging from 2 to 30%), while only 38% that of RBCs (ranging from 2 to 3.4%) were in HCs. We also found four modified morphologies of RBCs triggered by Aß binding, which may serve as an ancillary investigation and indicate the progression of AD. CONCLUSION: we first directly prove the existence of Aß binding RBCs in peripheral blood. In addition, we observed new modified morphologies of RBC triggered by Aß binding, all of those can serve as a biomarker for the diagnosis and progression of AD.


Assuntos
Doença de Alzheimer/diagnóstico , Peptídeos beta-Amiloides/sangue , Eritrócitos/metabolismo , Idoso , Doença de Alzheimer/sangue , Peptídeos beta-Amiloides/metabolismo , Biomarcadores/sangue , Estudos de Casos e Controles , Imunofluorescência , Humanos , Testes Neuropsicológicos
15.
Mol Med Rep ; 11(2): 1528-34, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25352049

RESUMO

Determination of disease activity in patients with rheumatoid arthritis (RA) has become an important component for RA management. The aim of the present study was to investigate the association between circulating levels of serum amyloid A (SAA) and disease activity in RA patients. The types of disease and the respective number of patients enrolled in the present study were as follows: RA, 88; osteoarthritis (OA), 54; systemic lupus erythematosus (SLE), 43; and other autoimmune diseases, 30, as well as 50 healthy controls (HC). SAA levels were measured using an ELISA assay and western blot analysis was used to detect serum SAA levels. The correlations between SAA levels and disease activity score for 28 joints (DAS28), erythrocyte sedimentation rate (ESR) and C­reactive protein (CRP), respectively, were evaluated; in addition, the presence and absence of rheumatoid factor (RF) and anti­cyclic citrullinated peptide antibody (anti­CCP) were detected in respect to SAA levels. The results of the present study demonstrated that serum levels of SAA in RA patients were significantly increased compared to those of the OA, SLE, others and HC patients (P<0.05). SAA levels were found to be positively correlated with DAS28, ESR and CRP levels (R2=0.6174, 0.4422 and 0.3919, respectively). In addition, anti­CCP was not correlated with DAS28 (R2=0.0154). Furthermore, increased SAA levels were detected in patients with positive anti­CCP compared with those in anti­CCP negative subjects (P<0.01). In conclusion, the results of the present study provided further evidence for possible roles of SAA in RA, which indicated that it may be a useful biomarker for assessing disease severity and may provide additional information about disease activity.


Assuntos
Proteínas da Fase Aguda/metabolismo , Artrite Reumatoide/patologia , Autoanticorpos/sangue , Proteína Amiloide A Sérica/análise , Adulto , Idoso , Artrite Reumatoide/metabolismo , Doenças Autoimunes/metabolismo , Doenças Autoimunes/patologia , Sedimentação Sanguínea , Proteína C-Reativa/análise , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imuno-Histoquímica , Lúpus Eritematoso Sistêmico/metabolismo , Lúpus Eritematoso Sistêmico/patologia , Pessoa de Meia-Idade , Osteoartrite/metabolismo , Osteoartrite/patologia , Fator Reumatoide/metabolismo , Índice de Gravidade de Doença , Líquido Sinovial/metabolismo
16.
Biomed Rep ; 2(5): 765-769, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25054025

RESUMO

Satisfactory biomarkers for screening and early diagnosis of lung cancer remain scarce and require further investigation. The aim of the present study was to examine the changes of the biochemical and protein composition in the serum and pleural effusion from lung cancer and lung infection (bacterial pneumonia) patients. A total of 92 patients with lung cancer, 38 with bacterial pneumonia and 42 healthy controls were enrolled in the study. The serum levels of cholesterol, apolipoprotein A and transthyretin (TTR) in the lung cancer patients were higher than that of the lung infection patients (P<0.05). The levels of TTR were higher, whereas the activity of adenosine deaminase (ADA) was lower in the pleural effusion from the lung cancer patients compared to the lung infection patients (P<0.05). Furthermore, the pleural effusion/serum TTR ratios in the lung cancer patients were higher, whereas the ratios of ADA were lower (P<0.05). By matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis, four major peaks corresponding to native TTR, Sul-TTR, Cys-TTR and Cysgly-TTR were observed in the serum of the lung cancer and lung infection patients. A significant increase was found in the proportion of Cysgly-TTR in the pleural effusion from the patients with lung cancer. The data indicated that a combination of pleural effusion/serum TTR ratios and modified TTR may be beneficial for the differential diagnosis between lung cancer and lung infection.

17.
J Zhejiang Univ Sci B ; 15(1): 92-9, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24390749

RESUMO

OBJECTIVE: Our aim was to investigate clinical and laboratory characteristics of osteoarthritic patients who had amyloid deposition in their knee joints. METHODS: Synovial membranes were obtained from 36 patients with knee osteoarthritis (OA) who underwent joint replacement surgery. From this sample, the diagnosis of amyloid was determined by Congo red staining, which demonstrated apple-green birefringence under a polarized microscope. All synovial membranes were immunohistochemically characterized for the expressions of amyloid immunoglobulin light chain (AL-κ and AL-λ), serum amyloid-A (SAA), amyloidogenic transthyretin (ATTR), and amyloidogenic ß2-microglobulin (Aß2M). Matrix-assisted laser desorption-ionizaton/time of flight mass spectrometry (MALDI-TOF MS) was used to analyze transthyretin (TTR) isoforms in the serum of each patient. RESULTS: Nine cases (25%) were found to be amyloid-positive. Immunohistochemically, eight cases (88.9%) had ATTR deposition, and one sample (11.1%) was shown to be AL-κ-positive. MALDI-TOF MS identified that the TTR in the serum of the patients was unmodified wild-type TTR, TTR-Cys-S-S-Cys, and TTR-Cys-S-S-CysGly. The age at surgery and the disease duration were significantly higher in the ATTR-positive group than in the ATTR-negative group. Knee score and function score were significantly lower in the ATTR-positive group than in the ATTR-negative group. CONCLUSIONS: Amyloid deposition in synovial membranes of OA patients was found to be ATTR and AL-κ. TTR in the serum of the patients was unmodified wild-type TTR together with two isoforms. The high age at surgery, long disease duration, and a deteriorated knee function were associated with ATTR amyloid deposition in the osteoarthritic knee joints.


Assuntos
Neuropatias Amiloides Familiares/diagnóstico , Neuropatias Amiloides Familiares/fisiopatologia , Articulação do Joelho/fisiopatologia , Osteoartrite do Joelho/diagnóstico , Osteoartrite do Joelho/fisiopatologia , Pré-Albumina/metabolismo , Idoso , Neuropatias Amiloides Familiares/complicações , Biomarcadores/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Osteoartrite do Joelho/complicações , Amplitude de Movimento Articular
18.
Int J Clin Exp Pathol ; 7(11): 7795-800, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25550818

RESUMO

Transthyretin (TTR) is a major amyloid fibril protein found in patients with familial amyloidotic polynuropathy (FAP) and senile systemic amyloidosis (SSA). Mainly synthesized in the live, TTR is transferred in the form of tetramer bound with thyroxine, retinol-binding protein (RBP) and lipoprotein in the blood. The aim of this study was to demonstrate the presence of amyloid substances in the blood by investigated the hemocoelom amyloid in different tissue sections from autopsies such as brain, kidney, heart and aorta arch tissue. Congo red staining was employed following by application of polarized light examination, to verify the presence of amyloid deposition in the tissues. Immunohistochemical staining was then performed to identify the specific type of amyloid deposition. Matrix-assisted laser desorption-ionization/time of flight mass spectrometry (MALDI-TOF/MS) was also used to analyze TTR mutation in FAP patients. All subjects were FAP ATTR Val30Met patients. In FAP patients, TTR amyloid deposition was found mainly in the tunica intima of the aortic arch. Interestingly, amyloid substance was found in the blood of FAP patient. Our results suggest that amyloid substance was present in the blood of FAP ATTR Val30Met patients.


Assuntos
Neuropatias Amiloides Familiares/sangue , Neuropatias Amiloides Familiares/genética , Amiloide/sangue , Mutação , Pré-Albumina/genética , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
19.
Artigo em Inglês | MEDLINE | ID: mdl-23934172

RESUMO

This study explored a method that can rapidly detect bacteria in urine samples for the auxiliary determination of urinary tract infections (UTIs). Urine samples from patients with UTIs (230 cases) were obtained using aseptic technique. The urine biochemical assay was then carried out using an automated urine analyzer for all the urine samples. Bacterial species were identified by a combination of bacterial culture technique, morphological observation and the BACT-IST microbial identification/susceptibility analysis system. The most common seven species of bacteria in the study included Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter cloacae, Enterococcus faecalis, Staphylococcus aureus and Staphylococcus epidermidis. Bacterial samples were suspended in sample buffer solutions and separated by the "three-plug-injection" method using capillary electrophoresis (CE). Each species of bacteria appeared as a bacterial peak. The mixture of the seven species also provided only one peak. Further analysis showed that the concentration limit for the "three-plug-injection" method is 10(6) colony forming units (CFU)/mL, and there is a good linear relationship between the peak height and bacterial concentration (R(2)=0.99). The effect of urine composition on CE results was also investigated. The results showed that urine composition, i.e., proteins, white blood cells (WBCs) and red blood cells (RBCs), affected the peak retention time but could not affect the separation of bacteria. The results demonstrated that the bacteria in urine samples can be detected within 10min by the "three-plug-injection" method using CE. The "three-plug-injection" method is therefore suitable for the rapid detection of organisms in clinical urine samples from UTIs.


Assuntos
Bactérias/isolamento & purificação , Eletroforese Capilar/métodos , Infecções Urinárias/microbiologia , Infecções Urinárias/urina , Urina/microbiologia , Bactérias/química , Feminino , Humanos , Masculino
20.
Biochem Biophys Res Commun ; 400(4): 559-62, 2010 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-20807507

RESUMO

Recent studies clearly demonstrated that several types of pathogenic amyloid proteins acted as agents that could transmit amyloidosis by means of a prion-like mechanism. Systemic AA amyloidosis is one of the most severe complications of chronic inflammatory disorders, particularly rheumatoid arthritis. It is well known that, similar to an infectious prion protein, amyloid-enhancing factor (AEF) acts as a transmissible agent in AA amyloidosis. However, how AEF transmits AA amyloidosis in vivo remained to be fully elucidated. In the present study, we focused on finding cell-free forms of AEF and its carriers in circulation by using the murine transfer model of AA amyloidosis. We first determined that circulating cell-free AEF existed in blood and plasma in mice with systemic AA amyloidosis. Second, we established that plasma exosomes containing AA amyloid oligomers derived from serum amyloid A had AEF activity and could transmit systemic AA amyloidosis via a prion-like mechanism. These novel findings should provide insights into the transmission mechanism of systemic amyloidoses.


Assuntos
Amiloidose/metabolismo , Exossomos/metabolismo , Príons/metabolismo , Proteína Amiloide A Sérica/metabolismo , Amiloidose/sangue , Animais , Glicoproteínas/sangue , Glicoproteínas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C3H
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