Down-regulation of human sialyltransferase gene expression during in vitro human keratinocyte cell line differentiation

We studied cytokines and anti-cytokine autoantibodies (Aabs) during T.b.brucei infections in IFN-gamma-/-, IFN-gammaR-/- and wild-type mice. Increased serum levels of IFN-gamma, TNF-gamma and IL-4 with decreased Aabs to these cytokines were recorded early during infections in all mice (except IFN-gamma in IFN-gamma-/- mice). Later, these responses were reversed, and surprisingly Aabs reacting to IFN-gamma in the IFN-gamma -/- mice were detected. To examine the possibility that an IFN-? immunoreactive molecule might be expressed due to infections and upon gene deletion, anti-IFN-gamma antibody was inoculated and resulted in abrogation of such Aabs. The scenario was different for IL-10 and TGF- since IFN-gammaR-/- and wild-type mice showed low cytokines and high Aabs early during infections, but later high cytokines and low Aabs were registered. Interestingly, IFN-gamma-/- mice exhibited reversed levels of both IL-10 and TGF-beta, and also of their Aabs. Fab fragments of purified serum immunoglobulins showed binding and neutralizing effects in biological assays. Pre-absorption of the Fab fragments with a cytokine inhibited the binding and neutralization effects of this cytokine, but not of other cytokines. These results highlight an important role for autoimmunity in cytokine regulation, and that genomic deletion of IFN-gamma modulates cytokines and their Aab responses in experimental African trypanosomiasis.     

Down-regulation of CXCR2 expression on human polymorphonuclear leukocytes by TNF-alpha

TNF-alpha is implicated in the initiation of cytokine cascades in various inflammatory settings. To assess the interactions of multiple cytokines at the level of inflammatory effector cells, we examined the effects of TNF-alpha on the expression of two IL-8Rs (CXCR1 and CXCR2) on polymorphonuclear leukocytes (PMNs). TNF-alpha decreased the surface expression of CXCR2 in a dose- and time-dependent manner. In contrast, CXCR1 expression was not affected by TNF-alpha. The release of CXCR2 into the supernatant of TNF-alpha-treated PMNs was detected by immunoblotting and immuno-slot-blot analyses, suggesting that the down-regulation of CXCR2 was caused mainly by shedding from the cell surface. The CXCR2 down-regulation was inhibited by PMSF and aprotinin, supporting the hypothesis that the shedding was mediated by serine protease(s). The intracellular Ca2+ mobilization and chemotaxis in response to IL-8 were suppressed by the pretreatment of PMNs with TNF-alpha, indicating that the decrease in CXCR2 was reflected in the decreased functional responses to IL-8. In contrast, the O2- release, which is mediated by CXCR1, was not suppressed by TNF-alpha. The treatment of whole blood with TNF-alpha also caused a significant reduction in CXCR2 and markedly suppressed intracellular Ca2+ mobilization and chemotaxis in response to IL-8, while enhancing the O2- release. These findings suggest that TNF-alpha down-regulates CXCR2 expression on PMNs and modulates IL-8-induced biologic responses, leading to the intravascular retention of PMNs with an enhanced production of reactive oxygen metabolites.     

Down-regulation of human FEN-1 gene expression during differentiation of promyelocytic leukemia cells

The presence of aflatoxin in corn and corn dust during relatively normal years and the increased risk of Aspergillus flavus infestation during drought conditions suggest that airborne agricultural exposures should be of considerable concern. Liquid extraction, thin layer chromatography, and high pressure liquid chromatography were used for the analysis of aflatoxin B1 in grain dust and bulk corn samples. A total of 24 samples of airborne dust were collected from 8 farms during harvest, 22 samples from 9 farms during animal feeding, and 14 sets of Andersen samples from 11 farms during bin cleaning. A total of 14 samples of settled dust and 18 samples of bulk corn were also collected and analyzed. The airborne concentration of aflatoxin B1 found in dust collected during harvest and grain unloading ranged from 0.04 to 92 ng/m3. Higher levels of aflatoxin B1 were found in the airborne dust samples collected from enclosed animal feeding buildings (5-421 ng/m3) and during bin cleaning (124-4849 ng/m3). Aflatoxin B1 up to 5100 ng/g were detected in settled dust collected from an enclosed animal feeding building; however, no apparent correlation was found between the airborne concentration of aflatoxin B1 and its concentration in settled dust or bulk corn. The data demonstrate that farmers and farm workers may be exposed to potentially hazardous concentrations of aflatoxin B1, particularly during bin cleaning and animal feeding in enclosed buildings.     

P2U-purinergic receptor activation mediates inhibition of cAMP accumulation in cultured renal mesangial cells

Extracellular ATP has been reported to exert mitogenic and contractile effects on cultured renal mesangial cells (MCs). Since it is possible that these actions involve changes in the cAMP second messenger system, we examined the effect of extracellular nucleotides on the accumulation of cAMP in rat MCs. ATP, UTP and adenosine 5'-0-(3-thio)triphosphate (ATP gamma S) (100 microM) had no significant effects on baseline cAMP levels, but inhibited forskolin-stimulated accumulation of cAMP by 21-75% in the presence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX). Maximal inhibitory effects were observed at 100 microM of ATP gamma S with a threshold dose of 1 microM. ATP gamma S, ATP and UTP were the most potent inhibitors indicating stimulation of the P2u receptor. The P2x agonists adenosine 5'-(alpha, beta-methylene) triphosphate and adenosine 5'-(beta, gamma-methylene) triphosphate, and the P2y agonist 2-methylthio-ATP did not affect cAMP accumulation. Treatment with the P2 receptor antagonist suramin (200 microM) reduced the inhibition by 58%. The inhibitory effects of the nucleotides were significantly attenuated by preincubation with pertussis toxin (10-100 ng/ml). Inhibition of phospholipase C and protein kinase C did not prevent the inhibitory effect of the nucleotides. Inhibitors of forskolin-stimulated cAMP accumulation had different effects on DNA synthesis in cultured MCs as measured by 3H-thymidine uptake at 48 h: ATP, ATP gamma S and the inhibitor of adenylyl cyclase, SQ 22536, stimulated DNA synthesis in MCs, while UTP showed no significant mitogenic effect. Agents which increased baseline levels of intracellular cAMP (forskolin, IBMX, dibutyryl-cAMP) significantly diminished DNA synthesis in MCs. The results indicate that the P2u-purinergic receptor mediates inhibition of forskolin-induced cAMP accumulation which is likely due to inhibition of adenylyl cyclase. This effect appears to be partially mediated by PTX-sensitive G proteins. While the increase in cAMP accumulation is anti-mitogenic, inhibition of cAMP accumulation by P2u receptors is not correlated with MC growth control. Thus, additional mechanisms other than inhibition of cAMP accumulation by P2u receptors are likely to be involved in the mitogenesis of extracellular ATP.     

Regulation of cathepsin E expression during human B cell differentiation in vitro

Cathepsin E is an aspartic proteinase which has been implicated in antigen processing in the class II major histocompatibility complex pathway. In this study we show that cathepsin E, measured at both the protein and message level, is up-regulated late in human B cell activation. The implications of this observation in terms of cathepsin E function are discussed.     

Intracellular distribution of polyamines in human lymphoblastoid cell line during phorbol ester-induced differentiation

Polyamines such as putrescine, spermidine and spermine play an important role in nucleic acid metabolism. These aliphatic amines display a key role in cell-induced transformation by carcinogen substances. In particular, one of these, the phorbol myristate acetate, provokes cell differentiation and gives an increase of ornithindecarboxylase activity; enzyme regulating the pathways of polyamines. In this study we analyse the trend of the polyamines at cytoplasmic and nuclear level during phorbol treatment. Our results show a correlation between nuclear and cytoplasmic spermine, 3H-Thymidine, 3H-Leucine incorporation and cell cycle phases. These data remark that the polyamines are differently distributed into the cell during the phorbol myristate acetate-mediated differentiation process and that the spermine is down-regulated for to supply the increased protein biosynthesis.     

Growth factor receptor expression during in vitro differentiation of partially purified populations containing murine stem cells

The Word Accentuation Test assesses the accentuation of 30 infrequent Spanish words written without the accentuation mark and is an easy-to-use tool for estimating premorbid intelligence of Spanish-speaking people. Its intraobserver (0.97) and interobserver (0.93) reliabilities and its correlation with the Wechsler Adult Intelligence Scale (.837) and Raven's Progressive Matrices (.655) are high, offering a good prediction of general intelligence. It is resistant to mental deterioration; 20 demented and 40 controls matched by sex, age, and education obtained similar scores. The discrepancies between current and predicted scores in Raven's scale can diagnose mild-moderate dementia with 0.79 accuracy (sensitivity, 0.78; specificity, 0.82).     

The desensitization of oxytocin receptors in human myometrial cells is accompanied by down-regulation of oxytocin receptor messenger RNA

Clinicians providing care to elderly patients must appreciate the subtle clinical manifestations that herald the onset of life-threatening infectious disease. Aged patients with an infection may have neither fever nor leucocytosis, making diagnosis challenging. Often, the early features of infectious disease are nonspecific and may resemble inflammatory or neoplastic processes, or there may be insufficient time to await definitive laboratory confirmation, and empirical antimicrobial treatment must be initiated. Aging involves inevitable deleterious alterations in biological processes and, in many elderly patients, this is most strongly characterised by diminished renal functional capacity. This has a major influence on antimicrobial prescribing in the elderly, because therapeutic efficacy must be achieved while minimising the risk of drug-related toxicity. Before prescribing an antibiotic to an aged patient with an infection, the clinician must be cognisant of the patient's drug allergy history and the other drugs that the patient is taking. Ignorance of potential drug-drug interactions can result in ineffective treatment or enhanced toxicity. The therapy of elderly patients with infections is being expanded. To reduce costs and enhance the efficiency of care, systems have been developed to provide antimicrobial care in the home and in long term care facilities. Home healthcare has burgeoned, and drugs that are well tolerated, have a broad spectrum of activity and are simple to administer (e.g. ceftriaxone and fluoroquinolones) appear to be eminently suitable for this therapeutic role. Physicians must also be informed of the factors responsible for the emergence of resistant bacteria that are contributing to infections in institutional and community settings. Clinicians should strive to curb inappropriate antibiotic use to stem the tide of infections that are caused by multidrug-resistant bacteria.     

Patterns of messenger RNA expression for alpha1-adrenoceptor subtypes in human corpus cavernosum

PURPOSE: The present study evaluated the expression of alpha1-adrenoceptor subtypes in human corpus cavernosum. MATERIALS AND METHODS: The mRNA encoding alpha1a, alpha1b and alpha1d subtypes were assessed by RNA-directed complementary cDNA synthesis followed by Taq DNA amplification. The level of alpha1 mRNA was calculated in arbitrary optical density units and normalized with respect to the length of the respective cDNA fragments. RESULTS: We found that alpha1a, alpha1b and alpha1d adrenoceptor subtypes are expressed in human corpus cavernosum, with a predominant expression of the alpha1a subtype. CONCLUSION: These results suggest that alpha1a-adrenoceptor subtype is important and that understanding the biochemical and functional characteristics of this subtype may lead to the development of specific antagonists in the treatment of impotence.     

Cellular localization of dopamine D2 receptor messenger RNA in the rat trigeminal ganglion

The actions of dopamine are mediated by specific, high-affinity, G protein-coupled receptors. Multiple subtypes of dopamine receptors have been characterized, including the D2 subtype (D2R). Cells within the dorsal root and petrosal ganglia of the rat express D2R messenger RNA (mRNA) consistent with D2R expression by primary sensory neurons. We hypothesized that neurons of the trigeminal ganglion express D2R mRNA. Total cellular RNA from rat trigeminal ganglia was analyzed on Northern blots under high stringency conditions. Hybridization of trigeminal ganglion RNA resulted in a signal which comigrated with striatal, pituitary, and hypothalamic D2R mRNA. To determine the distribution of D2R expressing cells in the trigeminal ganglion, cryostat sections were analyzed by in situ hybridization followed by emulsion autoradiography. We identified a population of clustered cells labeled with dense grain concentrations over their cytoplasms. These findings demonstrate the expression of D2 dopamine receptor mRNA in discrete subpopulations of neurons in the rat trigeminal ganglion. Our observations suggest that drugs active at dopamine receptors of the D2 subtype are potential modulators of sensory activity of neurons whose cell bodies reside in the trigeminal ganglion. D2 dopamine receptors may thus have a role in clinical pain syndromes involving the head and neck.     

Stimulation of oxidant generation by human sperm suspensions using phorbol esters and formyl peptides: relationships with motility and fertilization in vitro

OBJECTIVES: To investigate the influence of reactive oxygen species generated by human spermatozoa and contaminating leukocytes on sperm movement and fertilization in vitro. DESIGN: A chemiluminescence technique, using luminol and peroxidase, was used to monitor the generation of reactive oxygen species by human sperm suspensions and the results were correlated with sperm movement and the fertilization of human ova in vitro. SETTING: Diagnostic Andrology Laboratory and IVF Clinic. PATIENTS: Infertile couples undergoing IVF therapy. RESULTS: An N-formyl-methionyl-leucyl-phenylalanine (FMLP) provocation test was used to demonstrate that the presence of leukocytes in 28.5% of the sperm preparations was associated with elevated levels of spontaneous reactive oxygen species production, impaired movement, and a reduced capacity for fertilization in vitro. In the absence of leukocytes, exposure to phorbol ester stimulated a burst of reactive oxygen species generation by human spermatozoa, the magnitude of which was correlated highly with a loss of sperm motility but not with fertilization rates observed in the concurrent IVF cycle. CONCLUSION: Leukocyte contamination of human sperm preparations can be detected readily by FMLP-induced, luminol-dependent chemiluminescence and the results have an important bearing on the fertilizing capacity of the spermatozoa in vitro.     

Loss of KAI1 messenger RNA expression in both high-grade and invasive human bladder cancers

The molecular mechanisms responsible for metastasis are not fully understood. Recently, expression of the KAI1 gene on human chromosome 11p11.2 was found to be down-regulated in metastatic prostate cancer cell lines compared with normal human prostate, suggesting that KAI1 may be a metastasis suppressor gene. The aim of this study was to investigate whether there is reduced expression of KAI1 in late-stage bladder cancer. Sixty-six paraffin-embedded bladder tissue sections were analyzed for KAI1 mRNA by in situ hybridization. Nineteen of these were from patients with no histological evidence of bladder cancer, and 47 were from papillary transitional cell carcinomas (TCCs); of these, 16 were highly invasive. KAI1 mRNA was highly expressed in the specimens of normal bladder (11 of 11; 100%), inflammatory bladder (5 of 8; 63%), and noninvasive papillary TCCs of grades 1 and 2 (15 of 24; 63%), compared to grade 3 papillary TCCs (1 of 7; 14%) or invasive TCCs (1 of 16; 6%). The differences in expression between local and invasive disease were statistically significant (P 相似文献   

Lack of IL-2 cytokine expression despite Il-2 messenger RNA transcription in tumor-infiltrating lymphocytes in primary human breast carcinoma: selective expression of early activation markers

Tumor-infiltrating lymphocytes (TIL) are found in most human infiltrating ductal breast carcinomas. In studies of other tumors, TIL were capable of activation by IL-2, both in vitro and in vivo, to produce selective tumor cytolysis. Specific TIL-mediated tumor cytolysis in human breast tumors has recently been reported. The large numbers of TIL within human breast cancers imply that an immune response is occurring, since many of these cells express HLA class II as a late activation marker. However, the degree of early activation of the native TIL in breast tumors has not been fully investigated. Early activation markers CD69, CD43, and CD38 together with the IL-2R (CD25) and IL-2 cytokine were examined using mAbs and tissue section immunohistology. In situ hybridization was used to detect IL-2 mRNA (IL-2 mRNA) in parallel with immunohistochemical localization of IL-2. The results revealed the expression of CD69, CD43, and CD38, but markedly low CD25 (IL-2R) and IL-2 protein expression by the TIL. This strongly indicates that the TIL are an activated population of T cells that shows a deficiency in IL-2 protein and IL-2R expression despite adequate levels of IL-2 mRNA. The mechanism for apparent inhibition of IL-2 production and IL-2R expression in the presence of IL-2 mRNA is currently unclear; however, this may explain the relative anergic state of native TIL.