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1.
J Exp Clin Cancer Res ; 37(1): 180, 2018 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-30068373

RESUMEN

BACKGROUND: Glioma is the most common primary central nervous system tumor derived from glial cells. Kininogen-1 (KNG1) can exert antiangiogenic properties and inhibit proliferation of endothelial cells. The effect of KNG1 on the glioma is rarely reported, so our purpose in to explore its mechanism in glioma cells. METHODS: The differentially expressed genes (DEGs) were identified based on The Cancer Genome Atlas (TCGA) database. The KNG1-vector was transfected into the two glioma cells. The viability, apoptosis and cell cycle of glioma cells and microvessel density (MVD) were detected by cell counting kit-8 assay, flow cytometry and immunohistochemistry, respectively. The expression were measured by quantitative real-time PCR and Western blot, respectively. A tumor mouse model was established to determine apoptosis rate of brain tissue by terminal deoxynucleotidyl transfer-mediated dUTP nick end labeling (TUNEL) analysis. RESULTS: KNG1 was identified as the core gene and lowly expressed in the glioma cells. Overexpression of KNG1 inhibited cell viability and angiogenesis of glioma cells. Overexpression of KNG1 promoted the apoptosis and G1 phase cell cycle arrest of glioma cells. Moreover, the expressions of VEGF, cyclinD1, ki67, caspase-3/9 and XIAP were regulated by overexpression of KNG1. In addition, overexpression of KNG1 inhibited the activity of PI3K/Akt. Furthermore, overexpression of KNG1 decreased the tumor growth and promoted the apoptosis of decreased by overexpression of KNG1 in vivo. . CONCLUSIONS: Overexpression of KNG1 suppresses glioma progression by inhibiting the proliferation and promoting apoptosis of glioma cells, providing a therapeutic strategy for the malignant glioma.


Asunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Glioma/metabolismo , Glioma/patología , Quininógenos/biosíntesis , Animales , Apoptosis/fisiología , Neoplasias Encefálicas/sangre , Neoplasias Encefálicas/genética , Línea Celular Tumoral , Proliferación Celular/fisiología , Femenino , Glioma/sangre , Glioma/genética , Xenoinjertos , Humanos , Quininógenos/genética , Quininógenos/metabolismo , Masculino , Ratones , Ratones Desnudos , Persona de Mediana Edad , Transfección
2.
Anticancer Res ; 34(12): 6925-38, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-25503118

RESUMEN

The sera of patients with breast cancer have higher levels of des[Arg(9)]bradykinin, a kinin B1 receptor (B1R) agonist, than that from healthy individuals. Stimulation of breast cancer cells with the analog Lys-des[Arg(9)]bradykinin causes release of metalloproteinases-2 and -9 and increases cell proliferation. We examined the possibility that breast cancer cells, in addition to B1R, express the kinin-forming protease true tissue kallikrein (KLK1) and the endogenous proteins termed kininogens from which kinins are enzymatically released. Furthermore, we investigated whether stimulation of breast cancer cells with a B1R agonist would modify the cellular levels of KLK6, KLK10 and KLK11, three kallikrein-related peptidases with a still poorly-understood biological role in breast cancer. We found that breast cancer cells expressed KLK1 and kininogens, and that stimulation of estrogen-sensitive breast cancer cells with the B1R agonist produced down-regulation of KLK10 (a protease associated with growth suppression) but up-regulation of KLK11 and KLK6 (peptidases related to increased cell proliferation and invasiveness, respectively). Furthermore, we showed that the B1R agonist acts as a functional stimulus for the secretion of KLK1 and KLK6, an event relevant for kinin production and cell invasion, respectively.


Asunto(s)
Neoplasias de la Mama/metabolismo , Calicreínas/biosíntesis , Receptor de Bradiquinina B1/agonistas , Serina Endopeptidasas/biosíntesis , Bradiquinina/análogos & derivados , Bradiquinina/farmacología , Neoplasias de la Mama/sangre , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proliferación Celular , Regulación hacia Abajo , Femenino , Humanos , Calidina/análogos & derivados , Calidina/farmacología , Calicreínas/sangre , Calicreínas/genética , Quininógenos/biosíntesis , Células MCF-7 , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Invasividad Neoplásica , Interferencia de ARN , ARN Interferente Pequeño , Serina Endopeptidasas/sangre , Calicreínas de Tejido/biosíntesis , Calicreínas de Tejido/genética , Regulación hacia Arriba
3.
Int J Mol Med ; 31(3): 707-16, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23338533

RESUMEN

Polycystic ovary syndrome (PCOS) is a common endocrine-metabolic disorder, affecting 6-10% of women of reproductive age. The etiology remains poorly understood. To investigate the differentially expressed proteins from PCOS patients versus healthy women, the protein expression in follicular fluid was analyzed using two-dimensional electrophoresis (2-DE). Since follicular fluid contains a number of secretory proteins required for oocyte fertilization and follicle maturation, it is possible that follicular fluid can be used as a provisional source for identifying pivotal proteins associated with PCOS. In this study, six overexpressed proteins [kininogen 1, cytokeratin 9, antithrombin, fibrinogen γ-chain, apolipoprotein A-IV (apoA-IV) precursor and α-1-B-glycoprotein (A1BG)] in follicular fluids from PCOS patients were identified with matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF-MS) and nano LC-MS/MS. Western blot analysis confirmed that the protein expression levels of apoA-IV precursor and A1BG were increased in follicular fluid from PCOS patients compared with those from normal controls. The analysis of protein expression for other proteins revealed individual variation. These results facilitate the understanding of the molecular mechanisms of PCOS and provide candidate biomarkers for the development of diagnostic and therapeutic tools.


Asunto(s)
Apolipoproteínas A/biosíntesis , Apolipoproteínas A/genética , Líquido Folicular/metabolismo , Folículo Ovárico/metabolismo , Síndrome del Ovario Poliquístico/genética , Síndrome del Ovario Poliquístico/metabolismo , Adulto , Antitrombinas/análisis , Femenino , Fibrinógenos Anormales/biosíntesis , Glicoproteínas/biosíntesis , Glicoproteínas/genética , Humanos , Inmunoglobulinas/biosíntesis , Inmunoglobulinas/genética , Queratina-9/análisis , Queratina-9/biosíntesis , Quininógenos/biosíntesis , Proteómica
4.
Physiol Genomics ; 26(2): 152-7, 2006 Jul 12.
Artículo en Inglés | MEDLINE | ID: mdl-16837654

RESUMEN

We isolated PCR, RNA ligase-mediated rapid amplification of cDNA ends (RLM-RACE-PCR)-, and RT-PCR-generated clones from mouse kininogen family transcripts. DNA sequencing indicated that the clones were from two distinct genes. One set (K1) is from the previously reported mouse kininogen gene. The second set (K2) has an open reading frame, is 93% identical to K1 in the overlapping nucleotide sequence, and, unlike T-kininogens in the rat, encodes a bradykinin motif identical to K1. We discovered that K2 exists with two different 5' ends. We used RT-PCR to determine the distribution and relative abundance of K1 and K2 mRNA in mouse tissues. K2 is transcribed and K1 and K2 are generally both expressed in the same tissues; however, they differ in their regulation of the alternative splicing event that yields either low-molecular-weight kininogen (LMWK) or high-molecular-weight kininogen (HMWK). For example, in the liver K1 is expressed as both HMWK and LMWK, whereas K2 is only expressed as LMWK. Conversely, in the kidney K2 is strongly expressed as both HMWK and LMWK, whereas K1 is not expressed as HMWK and expressed only very weakly as LMWK.


Asunto(s)
Regulación de la Expresión Génica , Quininógenos/biosíntesis , Quininógenos/genética , Animales , Riñón/metabolismo , Quininógeno de Alto Peso Molecular/genética , Quininógeno de Bajo Peso Molecular/genética , Hígado/metabolismo , Ratones , Sistemas de Lectura Abierta , Reacción en Cadena de la Polimerasa , ARN Mensajero/metabolismo
5.
Biol Chem ; 387(2): 173-8, 2006 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-16497149

RESUMEN

Blood coagulation factor XII (FXII, Hageman factor) is a plasma serine protease which is autoactivated following contact with negatively charged surfaces in a reaction involving plasma kallikrein and high-molecular-weight kininogen (contact phase activation). Active FXII has the ability to initiate blood clotting via the intrinsic pathway of coagulation and inflammatory reactions via the kallikrein-kinin system. Here we have determined FXII-mediated bradykinin formation and clotting in plasma. Western blotting analysis with specific antibodies against various parts of the contact factors revealed that limited activation of FXII is sufficient to promote plasma kallikrein activation, resulting in the conversion of high-molecular-weight kininogen and bradykinin generation. The presence of platelets significantly promoted FXII-initiated bradykinin formation. Similarly, in vitro clotting assays revealed that platelets critically promoted FXII-driven thrombin and fibrin formation. In summary, our data suggest that FXII-initiated protease cascades may proceed on platelet surfaces, with implications for inflammation and clotting.


Asunto(s)
Coagulación Sanguínea/fisiología , Plaquetas/fisiología , Factor XII/fisiología , Sistema Calicreína-Quinina/fisiología , Bradiquinina/biosíntesis , Humanos , Quininógenos/biosíntesis , Peso Molecular , Valores de Referencia , Trombina/biosíntesis , Factores de Tiempo
6.
Arthritis Res Ther ; 7(4): R769-76, 2005.
Artículo en Inglés | MEDLINE | ID: mdl-15987478

RESUMEN

The human leukocyte antigen B27 (HLA-B27) transgenic rat is a model of human inflammatory bowel disease, rheumatoid arthritis and psoriasis. Studies of chronic inflammation in other rat models have demonstrated activation of the kallikrein-kinin system as well as modulation by a plasma kallikrein inhibitor initiated before the onset of clinicopathologic changes or a deficiency in high-molecular-mass kininogen. Here we study the effects of monoclonal antibody C11C1, an antibody against high-molecular-mass kininogen that inhibits the binding of high-molecular-mass kininogen to leukocytes and endothelial cells in the HLA-B27 rat, which was administered after the onset of the inflammatory changes. Thrice-weekly intraperitoneal injections of monoclonal antibody C11C1 or isotype IgG1 were given to male 23-week-old rats for 16 days. Stool character as a measure of intestinal inflammation, and the rear limbs for clinical signs of arthritis (tarsal joint swelling and erythema) were scored daily. The animals were killed and the histology sections were assigned a numerical score for colonic inflammation, synovitis, and cartilage damage. Administration of monoclonal C11C1 rapidly decreased the clinical scores of pre-existing inflammatory bowel disease (P < 0.005) and arthritis (P < 0.001). Histological analyses confirmed significant reductions in colonic lesions (P = 0.004) and synovitis (P = 0.009). Decreased concentrations of plasma prekallikrein and high-molecular-mass kininogen were found, providing evidence of activation of the kallikrein-kinin system. The levels of these biomarkers were reversed by monoclonal antibody C11C1, which may have therapeutic potential in human inflammatory bowel disease and arthritis.


Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antígeno HLA-B27/biosíntesis , Inflamación/metabolismo , Quininógenos/biosíntesis , Animales , Animales Modificados Genéticamente , Anticuerpos Monoclonales/farmacología , Artritis/tratamiento farmacológico , Artritis/genética , Artritis/metabolismo , Colitis/tratamiento farmacológico , Colitis/genética , Colitis/metabolismo , Antígeno HLA-B27/genética , Humanos , Inflamación/tratamiento farmacológico , Inflamación/genética , Quininógenos/genética , Masculino , Ratas , Ratas Endogámicas F344
8.
Biol Chem ; 385(3-4): 295-301, 2004.
Artículo en Inglés | MEDLINE | ID: mdl-15134344

RESUMEN

Kininogens serve dual functions by forming a scaffold for the assembly of the protein complex initiating the surface-activated blood coagulation cascade and as precursors for the kinin hormones. While rats have three kininogen genes, for mice, cattle, and humans only one gene has been described. Here, we present sequence and expression data of a second mouse kininogen gene. The two genes, kininogen-I and kininogen-II, are located in close proximity on chromosome 16 in a head-to-head arrangement. In liver and kidney, both genes are expressed and for each gene three alternative splice variants are synthesized. Two of them are the expected high and low molecular weight isoforms known from all mammalian kininogens. However, for both genes also a third, hitherto unknown splice variant was detected which lacks part of the high molecular weight mRNA due to splicing from the low molecular weight donor site to alternative splice acceptor sites in exon 10. The physiological functions of the six kininogen isoforms predicted by these findings need to be determined.


Asunto(s)
Regulación de la Expresión Génica/fisiología , Quininógenos/química , Quininógenos/genética , Empalme Alternativo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Orden Génico , Quininógeno de Alto Peso Molecular/química , Quininógeno de Alto Peso Molecular/genética , Quininógeno de Bajo Peso Molecular/química , Quininógeno de Bajo Peso Molecular/genética , Quininógenos/biosíntesis , Quininógenos/aislamiento & purificación , Masculino , Ratones , Datos de Secuencia Molecular , Isoformas de Proteínas/biosíntesis , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/aislamiento & purificación
9.
Int Immunopharmacol ; 3(3): 335-44, 2003 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-12639811

RESUMEN

The present study examined non-insulin-treated streptozotocin (STZ)-induced diabetic rats to determine the role of kinins in diabetic nephropathy. Their involvement in the renoprotective effect of the angiotensin-converting enzyme inhibitor (ACEI) ramipril was investigated using the bradykinin (BK) B(2)-receptor antagonist, icatibant (HOE 140), or a combination of the two drugs.Although, none of the treatments prevented the decline of the glomerular filtration rate (GFR) in diabetic rats, ramipril (3 mg/kg/day), but not icatibant (HOE 140; 500 microg/kg/day), prevented proteinuria in these animals. However, the antiproteinuric effect of ramipril was reduced by 45% when combined with icatibant. To explore whether the renal kallikrein-kinin system (KKS) belongs to the underlying mechanisms of these findings, we also determined urinary BK levels, renal kallikrein (KLK) and angiotensin-converting enzyme (ACE) activity as well as renal cortical mRNA levels of neutral endopeptidase 24.11 (NEP) and low-molecular weight (LMW) kininogen. STZ led to a reduction of renal KLK and ACE activity and NEP expression and to a three-fold increase of urinary BK excretion and renal kininogen expression. Icatibant given alone had no effect on these parameters. In contrast, ramipril treatment normalized urinary protein and BK excretion as well as kininogen mRNA expression without affecting NEP mRNA expression or KLK and ACE activity. Our data demonstrate that renal BK is increased in severe STZ-induced diabetes mellitus, but may affect glomerular regulation only to a minor degree under this condition. However, kinins are partly involved in the antiproteinuric action of ACEI at this stage of diabetic nephropathy.


Asunto(s)
Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Bradiquinina/análogos & derivados , Nefropatías Diabéticas/tratamiento farmacológico , Cininas/fisiología , Proteinuria/tratamiento farmacológico , Actinas/biosíntesis , Animales , Antiinflamatorios no Esteroideos/uso terapéutico , Presión Sanguínea/efectos de los fármacos , Bradiquinina/uso terapéutico , Antagonistas del Receptor de Bradiquinina B2 , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/patología , Nefropatías Diabéticas/complicaciones , Hipertensión Renal/complicaciones , Hipoglucemiantes/uso terapéutico , Insulina/uso terapéutico , Sistema Calicreína-Quinina/fisiología , Corteza Renal/efectos de los fármacos , Corteza Renal/enzimología , Pruebas de Función Renal , Quininógenos/biosíntesis , Masculino , Neprilisina/metabolismo , Peptidil-Dipeptidasa A/metabolismo , Proteinuria/etiología , Ramipril/uso terapéutico , Ratas , Ratas Wistar
10.
Toxicol Sci ; 73(1): 195-206, 2003 May.
Artículo en Inglés | MEDLINE | ID: mdl-12657746

RESUMEN

4-Aminophenol (4-AP), D-serine, and cisplatin are established rodent nephrotoxins that damage proximal tubules within the renal cortex. Using high throughput 2D gel proteomics to profile protein changes in the plasma of compound-treated animals, we identified several markers of kidney toxicity. Male F344 and Alpk rats were treated with increasing doses of 4-AP, D-serine, or cisplatin, and plasma samples were collected over time. Control groups received saline or nontoxic isomers, L-serine, and transplatin. Plasma proteins that displayed dose- and temporal-dependent regulation in each study were further characterized by mass spectrometry to elucidate the protein identity. Several isoforms of the rat-specific T-kininogen protein were identified in each study. T-kininogen was elevated in the plasma of 4-AP-, D-serine-, and cisplatin-treated animals at early time points, returning to baseline levels 3 weeks after treatment. The protein was not elevated in the plasma of control animals or those treated with nontoxic compounds. We propose that T-kininogen may be required to counteract apoptosis in proximal tubular cells in order to minimize tissue damage following a toxic insult. In addition, T-kininogen may be required to stimulate localized inflammation to aid tissue repair. We also identified several isoforms of the inter-alpha inhibitor H4P heavy chain in the 4-AP and D-serine studies. In each case, the protein expression levels in the blood samples paralleled the extent of kidney toxicity, highlighting the correlation between protein alterations and clinical chemistry endpoints. A further set of proteins correlating with kidney damage was found to be a component of the complement cascade and other blood clotting factors, indicating a contribution of the immune system to the observed toxicity. These observations underscore the value of proteomics in identifying new biomarkers and in the elucidation of mechanisms of toxicity.


Asunto(s)
Síndrome de Fanconi/inducido químicamente , Síndrome de Fanconi/patología , Proteoma/efectos de los fármacos , 4-Aminopiridina/toxicidad , Animales , Antineoplásicos/toxicidad , Proteínas Sanguíneas/metabolismo , Cisplatino/toxicidad , Creatinina/sangre , Relación Dosis-Respuesta a Droga , Electroforesis en Gel de Poliacrilamida , Procesamiento de Imagen Asistido por Computador , Pruebas de Función Renal , Quininógenos/biosíntesis , Masculino , Espectrometría de Masas , Peso Molecular , Bloqueadores de los Canales de Potasio/toxicidad , Proteoma/química , Ratas , Ratas Endogámicas F344 , Serina/toxicidad , alfa-Macroglobulinas/metabolismo
11.
Blood ; 101(11): 4430-6, 2003 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-12576314

RESUMEN

A 6-year-old male with vertebral-basilar artery thrombosis was recognized to have high-molecular-weight kininogen (HK) deficiency. The propositus had no HK procoagulant activity and antigen (< 1%). Using monoclonal antibodies (Mabs) to kininogen domain 3, the propositus, family members, and Fitzgerald plasma were determined to have detectable low-molecular-weight kininogen. Mabs to HK domains 5 and 6 do not detect HK antigen in the propositus' plasma. The propositus has a single base pair (bp) deletion in cDNA position 1492 of exon 10 affecting amino acid 480 of the mature protein and resulting in a frameshift and a premature stop codon at position 1597 (amino acid 532). Unexpectedly, Mabs to the heavy chain and domain 5 of HK detect a 92-kDa form of HK in Fitzgerald plasma, the first HK-deficient plasma. The 92-kDa Fitzgerald HK has amino acid residues through 502, corresponding to domains 1 through 5, but lacks epitopes of domain 6 (positions 543 to 595). Fitzgerald DNA has a normal exon 10, but a 17-bp mutation in intron 9. These combined results indicate that mutations in the kininogen gene may differentially affect biosynthesis, processing, and/or secretion of HK.


Asunto(s)
Quininógeno de Alto Peso Molecular/deficiencia , Niño , Codón sin Sentido , Exones , Salud de la Familia , Mutación del Sistema de Lectura , Humanos , Immunoblotting , Intrones , Quininógeno de Alto Peso Molecular/biosíntesis , Quininógeno de Alto Peso Molecular/genética , Quininógenos/biosíntesis , Quininógenos/genética , Quininógenos/metabolismo , Masculino , Estructura Terciaria de Proteína , Trombosis/sangre , Trombosis/genética
12.
Biochemistry (Mosc) ; 67(1): 13-24, 2002 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-11841336

RESUMEN

This review considers the data of recent years concerning the contact system initiating the activation of blood plasma proteolytic systems, such as hemocoagulation, fibrinolysis, kininogenesis, and also complement and angiotensinogenesis. The main proteins of the contact system are the factors XII and XI, prekallikrein, and high-molecular-weight kininogen. The data on the structure, functions, and biosynthesis of these proteins and on their genes are presented. Studies in detail on the protein-protein interactions during formation of the ensemble of the contact system components on the anionic surface resulted in the postulation of the mechanism of activation of this system associated with generation of the XIIa factor and of kallikrein. This mechanism is traditionally considered a trigger of processes for the internal pathway of the hemocoagulating cascade. However, the absence of direct confirmation of such activation in vivo and the absence of hemorrhagia in the deficiency of these components stimulated the studies designed to find another mechanism of their activation and physiological role outside of the hemostasis system. As a result, a new concept on the contact system activation on the endothelial cell membrane was proposed. This concept is based on the isolation of a complex of proteins, which in addition to the above-mentioned proteins includes cytokeratin 1 and the receptors of the urokinase-like plasminogen activator and of the complement q-component. The ideas on the role of this system in the biology of vessels are developed. Some of our findings on the effect of leukocytic elastase on the key components of the contact system are also presented.


Asunto(s)
Coagulación Sanguínea , Sangre/metabolismo , Endotelio/citología , Quininógenos/biosíntesis , Leucocitos/metabolismo , Membrana Celular/metabolismo , Endotelio/metabolismo , Factor XI/biosíntesis , Factor XII/biosíntesis , Fibrinólisis , Humanos , Queratinas/biosíntesis , Modelos Biológicos , Precalicreína/biosíntesis , Conformación Proteica , Estructura Terciaria de Proteína , Relación Estructura-Actividad , Factores de Tiempo
13.
Biochem Biophys Res Commun ; 287(1): 301-4, 2001 Sep 14.
Artículo en Inglés | MEDLINE | ID: mdl-11549291

RESUMEN

Differential mRNA display showed that a cDNA band disappeared after treatment of mice with 3-methylcholanthrene (MC). The cDNA encoded low-molecular-weight (LMW) prekininogen, known to be the precursor of a potent vasodilator, bradykinin. MC is generally known to bind to aryl hydrocarbon receptor (AhR) as an initial event to cause effects in vivo. In accordance with the results, Northern blot analysis for LMW prekininogen mRNA using total RNAs from wild-type and AhR-null mice indicated that the suppression of the mRNA expression by MC was seen in wild-type mice but not in AhR-null mice. The expression of LMW prekininogen mRNA was almost completely lost within 1 h after treatment of mice with MC, while a clear increase of CYP1A2 mRNA, as a positive control, was noted 4 h after the treatment. The plasma concentration of bradykinin released from LMW prekininogen was decreased by MC in wild-type mice, but not in AhR-null mice. Based on these results, we conclude that AhR inhibits bradykinin synthesis in mice via suppression of the expression of LMW prekininogen. Possible mechanism(s) responsible for hypertension caused by treatment of mice with MC is also discussed.


Asunto(s)
Quininógenos/genética , Precursores de Proteínas/genética , Receptores de Hidrocarburo de Aril/fisiología , Animales , Silenciador del Gen/efectos de los fármacos , Quininógenos/biosíntesis , Cininas , Masculino , Metilcolantreno/toxicidad , Ratones , Ratones Endogámicos C57BL , Dibenzodioxinas Policloradas/toxicidad , Precursores de Proteínas/biosíntesis , Teratógenos/toxicidad
14.
Life Sci ; 69(3): 359-68, 2001 Jun 08.
Artículo en Inglés | MEDLINE | ID: mdl-11441926

RESUMEN

The mRNAs for acute-phase proteins and kininogens were found to be increased in the submandibular gland (SMG) and extraorbital and intraorbital lacrimal gland (ELG and ILG) in response to experimentally induced inflammation in rats; i.e., 24 hours after subcutaneous injection of turpentine oil, mRNAs for C-reactive protein (CRP), serum amyloid P component (SAP), and H- and T-kininogens were induced in the SMG, ELG, and ILG of rats, whereas these mRNAs were not detected in the same tissues of normal control rats. The induction of mRNAs for these inflammatory proteins by turpentine oil was preceded by a transient increase in the level of mRNA for tumor necrosis factor-alpha (TNF-alpha) at 6 hours after subcutaneous injection of the oil. This was confirmed by injection of another inflammation inducer, lipopolysaccharide (LPS), which induced the TNF-alpha mRNA in the same way at 6 hours as turpentine oil did. The up-regulation of acute-phase proteins including kininogens in the SMG, ELG, and ILG suggest the existence of a strict defense system in the exocrine glands.


Asunto(s)
Proteína C-Reactiva/biosíntesis , Quininógenos/biosíntesis , Aparato Lagrimal/metabolismo , Glándula Submandibular/metabolismo , Animales , Proteína C-Reactiva/genética , Dacriocistitis/inducido químicamente , Dacriocistitis/metabolismo , Dacriocistitis/patología , Inyecciones Subcutáneas , Irritantes/administración & dosificación , Irritantes/toxicidad , Quininógenos/genética , Aparato Lagrimal/efectos de los fármacos , Aparato Lagrimal/patología , Masculino , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Sialadenitis/inducido químicamente , Sialadenitis/metabolismo , Sialadenitis/patología , Glándula Submandibular/efectos de los fármacos , Glándula Submandibular/patología , Enfermedades de la Glándula Submandibular/inducido químicamente , Enfermedades de la Glándula Submandibular/metabolismo , Enfermedades de la Glándula Submandibular/patología , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genética , Trementina/administración & dosificación , Trementina/toxicidad
15.
Biol Pharm Bull ; 23(10): 1239-42, 2000 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-11041259

RESUMEN

To characterize the local kallikrein-kinin system, tissue distribution of mRNAs for kininogens, precursor proteins of kinins, such as high-molecular-weight (H-), low-molecular-weight (L-) and T-kininogens, were studied in the rat by means of reverse-transcription polymerase chain reaction (RT-PCR) using a fluorophore Cy5-labeled 5'-primer. High levels of these three kininogen mRNAs were present in the liver. Relatively high levels of H-kininogen mRNA were also detected in the skin, lung, kidney, and testis in a descending order, whereas L-kininogen mRNA was detectable in the lung and brain, but not in the kidney, skin, testis, heart, adrenal gland, or skeletal muscle. T-Kininogen mRNA was present in these tissues, except for skeletal muscle. These findings suggest that the expression of each kininogen mRNA is regulated by the tissue-dependent mechanisms which is closely associated with functions of the kallikrein-kinin system in the tissue.


Asunto(s)
Quininógenos/biosíntesis , ARN Mensajero/biosíntesis , Animales , Southern Blotting , Electroforesis en Gel de Poliacrilamida , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Distribución Tisular
16.
Immunopharmacology ; 45(1-3): 121-6, 1999 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-10615000

RESUMEN

Expression of kininogen mRNAs has been studied in cultures of three different types of cells in rat brain, including neurons and astrocytes from cerebral cortex and meningeal cells from the leptomeninges/choroid plexus. T-kininogen mRNA was expressed by meningeal cells, but not by neurons and astrocytes, and the expression in meningeal cells was enhanced by culture with prostaglandin E2 (PGE2) or dibutyryl cAMP (Bt2cAMP). Low-molecular-weight kininogen mRNA was not detected in these cultures of cells, even after treatment with PGE2. Although expression of high-molecular-weight kininogen mRNA was very low in these cultures of cells, PGE2 or Bt2cAMP markedly stimulated its expression in cultures of meningeal cells and slightly in neurons, but not in astrocytes. We also found that expression of plasma kallikrein mRNA was strong in cultures of meningeal cells and slight in astrocytes, but absent in neurons. These results suggest that cells in the leptomeninges/choroid plexus are major sources of kininogens in rat brain which may function as precursor proteins for kinins and/or potent cysteine proteinase inhibitors during cerebral inflammation.


Asunto(s)
Encéfalo/citología , Encéfalo/metabolismo , Corteza Cerebral/metabolismo , Calicreínas/genética , Quininógenos/genética , ARN Mensajero/biosíntesis , Animales , Animales Recién Nacidos , Astrocitos/metabolismo , Encéfalo/enzimología , Células Cultivadas , Calicreínas/biosíntesis , Calicreínas/sangre , Quininógenos/biosíntesis , Quininógenos/sangre , Meninges/citología , Meninges/metabolismo , Neuronas/metabolismo , ARN Mensajero/sangre , Ratas
17.
Immunopharmacology ; 45(1-3): 135-9, 1999 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-10615002

RESUMEN

Components of the kallikrein-kinin system (KKS) have been shown to be synthesized in many tissues and cells, however, a systematic investigation on which of the KKS components are expressed in the various human tissues and cells and how their expression is regulated is not yet available. As a first step towards such a study we developed highly sensitive and specific reverse transcription polymerase chain reaction (RT-PCR) procedures for detecting mRNA expression of tissue kallikrein, high and low molecular weight kininogens, and kinin receptors B1 and B2. Analyses of a variety of human fibroblast and epithelial cell lines showed that they differ significantly in their individual expression profiles of KKS components indicating that the KKS participates in specific and diverse ways in the regulation of cellular functions. The RT-PCR procedures described here permit differentiation of cell lines and tissues according to their expression profiles of mRNAs of KKS components and thus provide a valuable means for selecting appropriate cells for studies on the functional significance of the KKS and its single components.


Asunto(s)
Sistema Calicreína-Quinina , Quininógenos/biosíntesis , Receptores de Bradiquinina/biosíntesis , Calicreínas de Tejido/biosíntesis , Línea Celular , Femenino , Humanos , Especificidad de Órganos , Receptor de Bradiquinina B1 , Receptor de Bradiquinina B2 , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Sensibilidad y Especificidad , Células Tumorales Cultivadas
18.
Immunopharmacology ; 44(1-2): 81-5, 1999 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-10604528

RESUMEN

To determine the existence of the kallikrein-kinin system in the heart, we have studied in vitro and in vivo whether rat heart expresses kininogens (KGNs). The reverse transcription-polymerase chain reaction (RT-PCR) for KGN mRNAs demonstrated that the cardiac tissue of adult male rats expresses T-KGN mRNA but not high-molecular-weight (H-) KGN mRNA. An intravenous injection of lipopolysaccharide (LPS) resulted in a significant increase in T-KGN mRNA levels of rat heart within 12 h. Polyacrylamide gel electrophoresis of cDNA products generated by RT-PCR from heart mRNA using primers specific for either T- or low-molecular-weigh (L-) KGN revealed that rat heart expressed not only T-KGN gene but also L-KGN gene, and that LPS injection exclusively stimulated the expression of T-KGN but not of L-KGN gene. T-KGN mRNA was also detected in cultured myocytes derived from fetal rat heart, and the expression was markedly enhanced by an addition of LPS to cultures. These results demonstrated that rat cardiomyocytes are the source of T- and L-KGNs but not of H-KGN, and that their expression of T-KGN mRNA is stimulated by LPS, probably via LPS-receptor CD14.


Asunto(s)
Regulación de la Expresión Génica , Quininógenos/genética , Miocardio/metabolismo , Animales , Células Cultivadas , Quininógenos/biosíntesis , Masculino , Peso Molecular , Miocardio/citología , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley
19.
Semin Thromb Hemost ; 25(6): 557-62, 1999.
Artículo en Inglés | MEDLINE | ID: mdl-10632478

RESUMEN

Administration of lipopolysaccharide (LPS) induces inflammation and tissue injuries that occasionally results in disseminated intravascular coagulation (DIC). This process is believed to be mediated by vasoactive molecules such as kinins and leads to endothelial damage and obstruction of the microcirculation. In this study, we evaluated the involvement of T-kininogen and macrophage migration inhibitory factor (MIF) in endotoxin-induced systemic inflammation. T-Kininogen is a protein unique to the rat and known as an acute-phase protein in response to endotoxins. Similarly, MIF functions as a proinflammatory cytokine and glucocorticoid-induced immunoregulator. First, we examined the effects of anti-MIF antibody on Wistar King male rats (ca 400 g) treated with intraperitoneal injection of LPS. At 6 hours after LPS injection (5 mg/kg), the platelet counts had decreased from 85 +/- 12.8 (x 10(4)/microL) to 8.8 +/- 2.6 (x 10(4)/microL). We treated these rats with the anti-rat MIF antibody (5 mg gamma G immunoglobulin [IgG] fraction/kg) 2 hours prior to LPS injection. This treatment prevented the decrease in platelet counts (45.6 +/- 5.6 [x 10(4)/microL]). Next, we examined the potential of MIF for production of T-kininogen. Intraperitoneal injection of rat MIF significantly upregulated the serum content of T-kininogen at the dose of 500 microg MIF/head. These results imply that MIF and T-kininogen might function in concert in the event of endotoxin-induced inflammation.


Asunto(s)
Quininógenos/biosíntesis , Factores Inhibidores de la Migración de Macrófagos/farmacología , Factor de Necrosis Tumoral alfa/biosíntesis , Animales , Anticuerpos/farmacología , Quininógenos/sangre , Quininógenos/efectos de los fármacos , Lipopolisacáridos/farmacología , Factores Inhibidores de la Migración de Macrófagos/inmunología , Masculino , Recuento de Plaquetas/efectos de los fármacos , Ratas , Ratas Wistar , Factores de Tiempo , Factor de Necrosis Tumoral alfa/efectos de los fármacos
20.
Biochim Biophys Acta ; 1404(3): 329-37, 1998 Sep 16.
Artículo en Inglés | MEDLINE | ID: mdl-9739161

RESUMEN

To identify the presence of a local kallikrein-kinin system in vascular wall, we have studied whether rat vascular smooth muscle cells (VSMC) express kininogen in vitro and in vivo. Western blots using anti-T-kininogen antibody revealed the presence of T-kininogen in conditioned medium of cultured VSMC. T-Kininogen secretion by VSMC was markedly enhanced by the addition of lipopolysaccharide (LPS), angiotensin II (AII) and phorbol 12-myristate 13-acetate (PMA) to the culture. Experiments using specific inhibitors for protein kinases and on the PMA-induced down-regulation of protein kinase C suggested that a protein kinase C-dependent or unidentified pathway is involved in AII or LPS action, respectively. The intravenous injection of LPS (0.5 mg/kg) resulted in an increase in T-kininogen mRNA levels in the vascular smooth muscle of rat aorta, peaking at 16 h. Polyacrylamide gel electrophoresis of cDNA products generated by reverse transcription-polymerase chain reaction (RT-PCR) from aortic mRNA using primers specific for either T- or low-molecular-weight kininogen revealed that rat vascular smooth muscle expressed T-kininogen gene but not low-molecular-weight kininogen gene, and that LPS exclusively stimulated T-kininogen expression. The mRNA for high-molecular-weight kininogen was undetectable in either aortic smooth muscle or cultured VSMC by means of RT-PCR analysis. RT-PCR using specific primers for rat tissue kallikrein genes showed that aortic smooth muscle expressed KLK1 (true kallikrein) mRNA, but not KLK10 (T-kininogenase) mRNA. These results demonstrated that rat VSMC are a source of T-kininogen but not of low-molecular-weight- or high-molecular-weight kininogen, in contrast to the expression of true kallikrein but not of T-kininogenase by these cells.


Asunto(s)
Angiotensina II/farmacología , Aorta/efectos de los fármacos , Quininógenos/biosíntesis , Lipopolisacáridos/farmacología , Músculo Liso Vascular/efectos de los fármacos , Angiotensina II/antagonistas & inhibidores , Animales , Aorta/metabolismo , Southern Blotting , Western Blotting , Células Cultivadas , Medios de Cultivo Condicionados/análisis , Inhibidores Enzimáticos/farmacología , Calicreínas/biosíntesis , Calicreínas/genética , Quininógeno de Alto Peso Molecular/genética , Quininógeno de Bajo Peso Molecular/genética , Quininógenos/genética , Lipopolisacáridos/antagonistas & inhibidores , Masculino , Músculo Liso Vascular/metabolismo , Reacción en Cadena de la Polimerasa , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , Acetato de Tetradecanoilforbol
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