RESUMO
BACKGROUND: Since the spread of Severe Acute Respiratory Syndrom Corona Virus 2 (SARS-CoV2) in Germany, intensive care beds have been kept free for patients suffering from Corona Virus Disease (COVID-19). Also, after the number of infections had declined, intensive care beds were kept free prophylactically; however, the percentage of beds reserved for COVID-19 differ in the individual federal states in Germany. The aim of this article is to define a necessary clearance quota of intensive beds for COVID-19 patients in Germany. An escalation and de-escalation scheme was created for rising and falling numbers of infected patients. METHODS: Data from the COVID-19 resource board of the state of Baden-Württemberg, the daily situation report of the Robert Koch Institute (RKI), the register of COVID-19 intensive care beds of the German Interdisciplinary Association for Intensive Care and Emergency Medicine (DIVI) as well as the daily report of COVID-19 Baden-Württemberg from April to November 2020 were used for the calculation. RESULTS: At the end of November 2020 approximately 13.5% of intensive care beds in Germany were used by COVID-19 patients. Of all persons tested positive for SARS-CoV2, 1.5% were admitted to an intensive care unit. The hospitalization rate was 6% and the mean age of infected persons was 43 years. Based on these numbers hospitals are recommended to keep 10% of intensive care beds available for COVID-19 patients in the case of less than 35 new infections/100,000 in the catchment area, 20% should be kept free in case of an advanced warning level of 35 new infections/100,000 inhabitants and 30% for a critical limit of 50 new infections/100,000 inhabitants. Further internal hospital triggers, such as the occupancy of the intensive care beds with COVID-19 patients, should be considered. CONCLUSION: If the number of infections is low a general nationwide retention rate of more than 10% of intensive care beds for COVID-19 patients is not justified. Locally increasing numbers of infections require a local dynamic approach. If the number of infections increases, the free holding capacity should be increased according to a step by step concept in close coordination with the local health authorities and other internal hospital triggers. In order not to overwhelm hospital capacities in the event of local outbreaks, a corresponding relocation concept should be considered at an early stage.
Assuntos
COVID-19 , Adulto , Cuidados Críticos , Hospitais , Humanos , Unidades de Terapia Intensiva , SARS-CoV-2RESUMO
Botrytis cinerea Pers. infects cut flower roses (Rosa × hybrida L.) during greenhouse production and gray mold symptoms are often expressed in the postharvest environment, resulting in significant economic losses. Disease management is based on cultural practices and preventative chemical treatments; however, gray mold outbreaks continue to occur. Rose tissues from six commercial shipments from two greenhouses in Colombia were evaluated to determine the Botrytis species composition as well as identify other pathogens present, gray mold incidence and severity, and fungicide resistance profiles. Botrytis isolates (49 total) were grouped into six morphological phenotypes, and all were identified to be B. cinerea sensu stricto. Disease incidence was higher in the petals than in the stem, stamen, ovary, sepal, or leaf tissues. Other fungi were isolated infrequently and included Alternaria alternata, Cladosporium cladosporioides, Epicoccum nigrum, Penicillium citrinum, Aspergillus brasiliensis, and Diplodia sp. Fungicide resistance profiles were determined using previously established discriminatory doses. Isolates resistant to thiophanate-methyl, iprodione, boscalid, and cyprodinil were found frequently in all shipments and in both greenhouses. The frequency of resistance to penthiopyrad, fenhexamid, fluopyram, isofetamid, and fludioxonil varied between shipments and greenhouses. No resistance to pydiflumetofen was observed at the discriminatory doses tested. Isolates with resistance to multiple chemical classes were commonly found. These results indicate that fungicide resistance management practices may improve preharvest and postharvest gray mold control of cut flower roses.
Assuntos
Botrytis , Rosa , Antifúngicos/farmacologia , Botrytis/efeitos dos fármacos , Botrytis/fisiologia , Colômbia , Farmacorresistência Fúngica , Rosa/microbiologiaRESUMO
Advanced materials, such as metal matrix composites (MMCs), are important for innovation, national security, and addressing climate change. MMCs are used in military, aerospace, and automotive applications because of their exceptional mechanical and thermal properties, however adoption has been slow due to costly and onerous manufacturing processes. A new process using fused filament fabrication 3D printing has been developed to make net shape MMCs without tooling or machining. The process involves printing an alumina preform and then using pressure-less infiltration with a molten aluminum alloy to form the composite. Arbitrary shapes can be formed in this process-a brake lever and a flange are demonstrated-and the properties can be tuned by varying the ceramic infill geometric pattern and ceramic loading. By using 35 vol% continuous fiber reinforcement over 800 MPa strength and 140 GPa modulus are achieved for the aluminum composite, 3.4 × and 2 × the matrix aluminum properties.
RESUMO
The rate of cholesterol synthesis from [14C]acetate was low in circulating blood lymphocytes freshly isolated from 17 normal subjects and 4 subjects with homozygous FH. On the other hand, the rate of cholesterol synthesis was two to fourfold above normal in freshly isolated lymphocytes from two subjects with abetalipoproteinemia. When the lymphocytes from subjects with all three genotypes were incubated for 48-72 h in the absence of lipoproteins, the rate of cholesterol synthesis increased by 5-15-fold. The subsequent addition of plasma LDL, but not HDL, rapidly suppressed cholesterol synthesis in the lymphocytes from normal subjects. In contrast, lymphocytes from the FH homozygotes, which have been shown previously to be deficient in cell surface LDL receptors, were resistant to LDL-mediated suppression of cholesterol synthesis. In addition to its ability to suppress cholesterol synthesis after it had been elevated by incubation of the cells in the absence of lipoproteins, LDL was able to suppress the induction of the enhanced rate of sterol synthesis when added to normal lymphocytes immediately after their isolation from the bloodstream. In contrast to the former action of LDL, the latter action of LDL-i.e., the suppression of induction of sterol synthesis-also occurred to a limited extent in lymphocytes from FH homozygotes. However, the FH lymphocytes, but not the normal cells, could be made resistant to this action of LDL by inclusion in the incubation medium of lipoprotein-deficient serum (30 percent, vol/vol) plus HDL (1 mg protein/ml). Considered together with previous data demonstrating a deficiency of LDL receptors in freshly isolated lymphocytes from FH homozygotes, the current studies provide evidence in support of the hypothesis that the interaction of plasma LDL with its cell surface receptor serves to regulate cholesterol synthesis in human lymphocytes.
Assuntos
Abetalipoproteinemia/genética , Colesterol/biossíntese , Hipercolesterolemia/genética , Lipoproteínas LDL/fisiologia , Linfócitos/metabolismo , Abetalipoproteinemia/sangue , Acetatos/metabolismo , Adolescente , Adulto , Criança , Depressão Química , Retroalimentação , Feminino , Fibroblastos/metabolismo , Homozigoto , Humanos , Hipercolesterolemia/sangue , Lanosterol/biossíntese , Lipoproteínas HDL/fisiologia , Lipoproteínas LDL/metabolismo , Lipoproteínas LDL/farmacologia , Masculino , Ácidos Oleicos/metabolismo , Receptores de DrogaRESUMO
Niemann-Pick disease type C (NPC) is characterized by substantial intracellular accumulation of unesterified cholesterol. The accumulation of unesterified cholesterol in NPC fibroblasts cultured with low density lipoprotein (LDL) appears to result from the inability of LDL to stimulate cholesterol esterification in addition to impaired LDL-mediated downregulation of LDL receptor activity and cellular cholesterol synthesis. Although a defect in cholesterol transport in NPC cells has been inferred from previous studies, no experiments have been reported that measure the intracellular movement of LDL-cholesterol specifically. We have used four approaches to assess intracellular cholesterol transport in normal and NPC cells and have determined the following: (a) mevinolin-inhibited NPC cells are defective in using LDL-cholesterol for growth. However, exogenously added mevalonate restores cell growth equally in normal and NPC cells; (b) the transport of LDL-derived [3H]cholesterol to the plasma membrane is slower in NPC cells, while the rate of appearance of [3H]acetate-derived, endogenously synthesized [3H]cholesterol at the plasma membrane is the same for normal and NPC cells; (c) in NPC cells, LDL-derived [3H]cholesterol accumulates in lysosomes to higher levels than normal, resulting in defective movement to other cell membranes; and (d) incubation of cells with LDL causes an increase in cholesterol content of NPC lysosomes that is threefold greater than that observed in normal lysosomes. Our results indicate that a cholesterol transport defect exists in NPC that is specific for LDL-derived cholesterol.
Assuntos
LDL-Colesterol/metabolismo , Doenças de Niemann-Pick/metabolismo , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Fibroblastos/metabolismo , Humanos , Cinética , Lipoproteínas LDL/fisiologia , Lisossomos/metabolismo , Ácido Mevalônico/farmacologia , Valores de Referência , Frações Subcelulares/metabolismo , TrítioRESUMO
The transfer of normal human fibroblasts from medium containing whole serum to medium devoid of lipoproteins produced a 90 percent decrease in the cellular content of cholesteryl esters and a 30 percent decrease in the free cholesterol content. When these lipoprotein-deprived cells were subsequently incubated with human low density lipoprotein (LDL), there was a 7-fold increase in the cellular content of esterified cholesterol and a 1.6-fold increase in the cellular content of free cholesterol. The concentration at which LDL produced its half-maximal effect in elevating cellular sterol content (30 mug/ml of LDL-cholesterol) was similar to the half-maximal concentration previously reported for high affinity binding of LDL to its cell surface receptor. High density lipoprotein (HDL) and whole serum from a patient with abetalipoproteinemia (neither of which contains a component that binds to the LDL receptor) did not produce a significant increase in the content of either cholesterol or cholesteryl esters in normal cells. Furthermore, in fibroblasts from patients with the homozygous form of familial hypercholesterolemia, which lack functional LDL receptors, LDL had no effect in raising the cellular content of either free or esterified cholesterol even when present in the medium at concentrations as high as 450 mug sterol/ml. It is concluded that LDL-receptor interactions constitute an important biochemical mechanism for the regulation of the cholesterol content of normal human fibroblasts. Moreover, when considered in light of current concepts of LDL metabolism in intact mammals, the present data suggest that a major function of plasma LDL may be to transport cholesterol from its site of synthesis in liver and intestine to its site of uptake in peripheral tissues.
Assuntos
Colesterol/análise , Fibroblastos/análise , Lipoproteínas LDL/metabolismo , Receptores de Droga , Abetalipoproteinemia/sangue , Transporte Biológico , Criança , Colesterol/biossíntese , Colesterol/metabolismo , Ésteres , Feminino , Fibroblastos/efeitos dos fármacos , Homozigoto , Humanos , Hipercolesterolemia/genética , Recém-Nascido , Mucosa Intestinal/metabolismo , Lipoproteínas HDL/farmacologia , Lipoproteínas LDL/sangue , Fígado/metabolismo , Masculino , Ácidos Oleicos/farmacologia , Ligação ProteicaRESUMO
We used dual laser two-color flow cytometry to compare the expression of surface markers associated with activation and with differentiation in lung and peripheral blood T lymphocytes from normal subjects. T cell subsets, defined based on their reactivity with monoclonal antibodies (MAb) OKT3, OKT4, and OKT8, were analyzed for expression of activation antigens as detected by MAbs to the interleukin-2 receptor, the transferrin receptor, and HLA-DR determinants. Whereas circulating T lymphocytes expressed the three activation antigens at low levels, and the total of T4+ and T8+ cells always approximated the number of T3+ cells, lung T lymphocytes of the T3+, T4+, and T8+ populations expressed the activation antigens at variable levels in combinations not seen in circulating lymphocytes, and the sum of T4+ and T8+ cells always exceeded the T3+ total. A proportion of T4+T8+ cells was detected in lung lymphocytes.
Assuntos
Antígenos de Superfície/análise , Pulmão/citologia , Linfócitos T/imunologia , Adulto , Anticorpos Monoclonais , Antígenos de Diferenciação de Linfócitos T , Ciclo Celular , Feminino , Citometria de Fluxo , Antígenos HLA-DR , Antígenos de Histocompatibilidade Classe II/análise , Humanos , Ativação Linfocitária , Masculino , Receptores Imunológicos/análise , Receptores de Interleucina-2RESUMO
A recombinant plasmid containing a full-length cDNA for hamster 3-hydroxy-3-methylglutaryl coenzyme A reductase was introduced by calcium phosphate-mediated transfection into UT-2 cells, a mutant line of Chinese hamster ovary cells that lack 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and thus require low density lipoprotein-cholesterol and mevalonate for growth. We selected a line of permanently transfected cells, designated TR-36 cells, that expressed high levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and thus grew in the absence of low density lipoprotein and mevalonate. Constitutive synthesis of reductase mRNA in TR-36 cells was driven by the simian virus 40 early promoter, and therefore the mRNA was not suppressed by sterols, such as 25-hydroxycholesterol or cholesterol derived from low density lipoprotein, which normally suppresses transcription of reductase mRNA when the reductase gene is driven by its own promoter. Although TR-36 cells continued to synthesize large amounts of reductase mRNA and protein in the presence of sterols, reductase activity declined by 50 to 60%. This decline was caused by a twofold increase in the rate of degradation of preformed enzyme molecules. The current data demonstrate that sterols accelerate the degradation of reductase protein independently of any inhibitory effect on the synthesis of the protein.
Assuntos
Hidroximetilglutaril-CoA Redutases/metabolismo , Lovastatina/análogos & derivados , Esteróis/farmacologia , Animais , Linhagem Celular , Células Cultivadas , Colesterol/farmacologia , Cricetinae , Cricetulus , DNA/genética , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hidroxicolesteróis/farmacologia , Hidroximetilglutaril-CoA Redutases/genética , Peso Molecular , Naftalenos/farmacologia , OvárioRESUMO
The t(11;14)(q13;q32) translocation has been associated with human B-lymphocytic malignancy. Several examples of this translocation have been cloned, documenting that this abnormality joins the immunoglobulin heavy-chain gene to the bcl-1 locus on chromosome 11. However, the identification of the bcl-1 gene, a putative dominant oncogene, has been elusive. In this work, we have isolated genomic clones covering 120 kb of the bcl-1 locus. Probes from the region of an HpaII-tiny-fragment island identified a candidate bcl-1 gene. cDNAs representing the bcl-1 mRNA were cloned from three cell lines, two with the translocation. The deduced amino acid sequence from these clones showed bcl-1 to be a member of the cyclin gene family. In addition, our analysis of expression of bcl-1 in an extensive panel of human cell lines showed it to be widely expressed except in lymphoid or myeloid lineages. This observation may provide a molecular basis for distinct modes of cell cycle control in different mammalian tissues. Activation of the bcl-1 gene may be oncogenic by directly altering progression through the cell cycle.
Assuntos
Ciclinas/genética , Família Multigênica/genética , Oncogenes/genética , Proteínas Proto-Oncogênicas/genética , Sequência de Aminoácidos , Sequência de Bases , Northern Blotting , Southern Blotting , Clonagem Molecular , Ciclina D1 , Expressão Gênica/fisiologia , Genes Dominantes/genética , Humanos , Dados de Sequência Molecular , Alinhamento de Sequência , Translocação Genética/genética , Células Tumorais CultivadasRESUMO
PURPOSE: Preoperative chemoradiotherapy may increase the R0 (curative) resection rate, overall survival (OS) duration, and disease-free survival (DFS) duration. We evaluated paclitaxel-based induction chemotherapy and chemoradiotherapy in patients with localized gastric or gastroesophageal adenocarcinoma to determine its feasibility, impact on the R0 resection rate, type of pathologic response, OS, and DFS. PATIENTS AND METHODS: Patients with operable, localized gastric, or gastroesophageal adenocarcinoma were eligible. Staging included endoscopic ultrasonography (EUS) and laparoscopy. Patients received two 28-day cycles of induction chemotherapy of fluorouracil, paclitaxel, and cisplatin followed by 45 Gy of radiation and concurrent fluorouracil plus paclitaxel. The cancer was restaged and surgery was attempted. Postsurgery pathologic findings and R0 resection were correlated with OS and DFS. RESULTS: Forty-one patients were enrolled. Most carcinomas were proximal (83%) and pretreatment stage EUST3 (85%). Forty patients (98%) underwent surgery, and 78% had an R0 resection. We observed a pathologic complete response (pathCR) rate of 20% and a pathologic partial response (pathPR) rate of 15% (< 10% residual cancer cells in the resected specimen). No pretreatment parameter (sex, cancer location, baseline T stage, or baseline N stage) predicted the type of postsurgery pathologic response, OS, or DFS. However, pathCR (P = .02), pathCR + pathPR (P = .006), R0 resection (P < .001), and postsurgery T and N stages (P = .01 and P < .001, respectively) were associated with OS. Same parameters were significantly correlated with DFS. Toxicity was manageable. CONCLUSION: The type of pathologic response but not pretreatment parameters was associated with OS and DFS. Efforts to increase the rate of pathologic response and better systemic cancer control are warranted.
Assuntos
Adenocarcinoma/terapia , Quimioterapia Adjuvante , Paclitaxel/administração & dosagem , Radioterapia Adjuvante , Neoplasias Gástricas/terapia , Adenocarcinoma/mortalidade , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Terapia Combinada , Esquema de Medicação , Neoplasias Esofágicas/mortalidade , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/terapia , Feminino , Humanos , Masculino , Terapia Neoadjuvante , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Análise de SobrevidaRESUMO
The bcl-1 gene maps to chromosome 11q13 and has recently been shown to be a member of the cyclin gene family. Amplification of the chromosome region containing bcl-1 occurs frequently in breast cancer, squamous cell cancer, and other tumor types. We have hypothesized that amplification results in altered expression of the bcl-1 gene, contributing to carcinogenesis. In this work, we studied bcl-1 gene amplification and expression in a panel of human cell line. bcl-1 is expressed in all cell lines studied. The level of expression tends to be higher in amplified cell lines. We also screened these cell lines for int-2 and hst-1 expression, genes which are frequently coamplified with bcl-1. No int-2 expression was detected, and the two cell lines expressing hst-1 were unamplified. Our data provide support for the importance of bcl-1 in carcinogenesis.
Assuntos
Cromossomos Humanos Par 11 , Amplificação de Genes/genética , Regulação Neoplásica da Expressão Gênica/genética , Neoplasias/genética , RNA Mensageiro/análise , RNA Neoplásico/análise , Northern Blotting , Southern Blotting , Humanos , Células Tumorais CultivadasRESUMO
A low-nicotine cigarette smoke condensate, 12 fractions of the condensate, and a reconstituted sample were tested for their ability to induce transformation in the mouse cell line 3H/10T-1/2 CL-8. This cell line is noted for its remarkable low spontaneous rate of transformation. Both the crude condensate and the reconstituted sample as well as two specific fractions induced transformation in the mouse cells. These transformed cells produced fibrosarcomas when injected s.c. into antithymocyte serum-treated syngeneic mice.
Assuntos
Transformação Celular Neoplásica , Fibrossarcoma/induzido quimicamente , Fumar , Animais , Soro Antilinfocitário , Linhagem Celular , Terapia de Imunossupressão , Camundongos , Camundongos Endogâmicos C3H , Sarcoma Experimental/induzido quimicamente , Linfócitos T , AlcatrõesRESUMO
Two mouse anti-human monoclonal antibodies (S3.13 and S5.7) raised against cells of acute myelogenous leukemia were found to react with antigens expressed on the surface of subsets of monocytes and lymphocytes. S3.13 precipitates a peptide of Mr 29,000, and S5.7 precipitates a peptide of Mr 20,000 present on the surface of all the cell types tested. These two surface antigens were distributed on discrete subpopulations of normal hemopoietic cells. The antibodies reacted with all (S5.7) or a subpopulation (S3.13) of peripheral blood T-lymphocytes, and with a subset of monocytes. Both antibodies reacted with bone marrow blast cell progenitors of the myelomonocytes and erythroid lineage. S5.7 also reacted with non-T-lymphocytes and with cells of the eosinophilic lineage. Both antigens disappeared from the cell surface during normal myeloid and erythroid differentiation. Thus, these monoclonal antibodies define the molecular characteristics and the cellular distribution of two differentiation antigens present on cells of the hemopoietic lineage.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Superfície/imunologia , Células-Tronco Hematopoéticas/imunologia , Animais , Linhagem Celular , Humanos , Leucemia Mieloide Aguda/imunologia , Camundongos , Monócitos/imunologia , Linfócitos T/imunologiaRESUMO
The mechanism of biosynthesis of 4-methyl-5-beta-hydroxyethyl thiazole, the thiazole moiety of thiamine was studied in Salmonella typhimurium. Using the adenosine derepression technique the incorporation of various 14C-labeled precursors was determined. We found that;e1Me-14C]methionine, [2-14C]methionine, [U-14C]alanine, and [2-14C]glycine were not incorporated whereas [2-14C]tyrosine was incorporated. Degradation of the 4-methyl-5-beta-hydroxyethyl thiazole obtained after [2-14C]tyrosine incorporation revealed that all of the activity was located on carbon-2. These findings are discussed and compared with previous findings concerning 4-methyl-5-beta-hydroxyethyl thiazole biosynthesis.
Assuntos
Salmonella typhimurium/metabolismo , Tiamina/biossíntese , Tiazóis/metabolismo , Alanina/metabolismo , Glicina/metabolismo , Metionina/metabolismo , Tirosina/metabolismoRESUMO
Endoplasmic reticulum-like membranes (MAM) that are associated with mitochondria have been implicated as intermediates in the import of lipids, particularly phosphatidylserine, from the endoplasmic reticulum to mitochondria (Vance, J.E. (1990) J. Biol. Chem. 265, 7248-7256; Shiao, Y.-J. et al. (1995) J. Biol. Chem. 270, 11190-11198). We have now examined further the role of MAM in lipid metabolism using the mnd/mnd mouse, a model for the human degenerative disease neuronal ceroid lipofuscinosis. The biochemical phenotype of the mnd/mnd mutant mouse (in which lipids and proteins accumulate abnormally in storage bodies in cells of affected tissues) suggested that the mutation might lead to impaired mitochondrial import of lipids and proteins as a result of a defective linkage between MAM and mitochondria. We, therefore, investigated the status of MAM and phospholipid metabolism in mnd/mnd mice livers. Separation of MAM from livers of older, but not younger, mnd/mnd mice was aberrant. In addition, the amount of the MAM-specific protein, phosphatidylethanolamine N-methyltransferase-2 (PEMT2), was greatly reduced in homogenates and MAM from livers of mnd/mnd mice of all ages, although PEMT2 mRNA abundance was normal. Moreover, PEMT activity in MAM from mnd/mnd mice was 60% less than in control mice. Activities of two additional phospholipid biosynthetic enzymes-CTP:phosphocholine cytidylyltransferase and phosphatidylserine synthase-were also reduced by > 50% in mnd/mnd microsomes. Radiolabeling experiments in hepatocytes indicated that neither the mitochondrial import nor the subsequent metabolism of phosphatidylserine was grossly affected in mnd/mnd mice. However, 3 proteins (cytochrome b5, NADH:cytochrome b5 reductase and mitochondrial F1Fzero-ATP synthase c subunit) which are normally present in mitochondria were partially redistributed to microsomes in mnd/mnd mouse liver. These studies indicate that MAM are defective in the mnd/mnd mutant mouse in which the biochemical phenotype includes an abnormal accumulation of lipids and proteins in storage bodies.
Assuntos
Membranas Intracelulares/enzimologia , Mitocôndrias Hepáticas/enzimologia , Lipofuscinoses Ceroides Neuronais/enzimologia , Lipofuscinoses Ceroides Neuronais/patologia , Transferases de Grupos Nitrogenados , Fosfolipídeos/biossíntese , Animais , Fracionamento Celular , Diacilglicerol Colinofosfotransferase/metabolismo , Modelos Animais de Doenças , Etanolaminofosfotransferase/metabolismo , Humanos , Membranas Intracelulares/ultraestrutura , Lipídeos/biossíntese , Fígado/enzimologia , Metiltransferases/metabolismo , Camundongos , Camundongos Mutantes , Mitocôndrias Hepáticas/ultraestrutura , Fosfatidiletanolamina N-Metiltransferase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Frações Subcelulares/enzimologia , Transferases/metabolismoRESUMO
We analyzed the phenotypic changes associated with monocyte activation and differentiation using a newly developed monoclonal antibody (B148.4). Among peripheral blood leukocytes, the antigen recognized by this antibody is expressed on monocytes and granulocytes at high and low density, respectively. Antigen expression is lost during in vitro differentiation of monocytes and is absent on tissue macrophages, indicating that expression of this antigen is related to monocyte differentiation. Only 1 alpha,25-dihydroxyvitamin D3 and phorbol diesters, of several inducers tested, up-regulate B148.4 antigen expression on cells (monocytes and certain myeloid cell lines) that constitutively bear the antigen, without, however, allowing its maintenance during monocytic differentiation or inducing it on negative cells. By immunoprecipitation from B148.4+ U937 cells, the antigen is a complex of a major 116-kd and two minor 38- and 46-kd molecules. Analysis of eight different tissues reveals that the antigen is shared with endothelial cells. Biochemical characteristics, cellular distribution, and expression pattern on monocytes, myeloid cell lines, and AML cells upon culture with different stimuli indicate that B148.4 is a novel monocyte differentiation antigen.
Assuntos
Antígenos de Diferenciação/análise , Monócitos/fisiologia , Anticorpos Monoclonais , Antígenos de Diferenciação/imunologia , Calcitriol/farmacologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Endotélio Vascular/citologia , Endotélio Vascular/fisiologia , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Linfócitos/citologia , Linfócitos/fisiologia , Monócitos/citologia , Monócitos/efeitos dos fármacos , Neutrófilos/citologia , Neutrófilos/fisiologia , Células Tumorais CultivadasRESUMO
Functioning bovine adrenocortical cells in monolayer culture were shown to obtain cholesterol for steroid synthesis from plasma low density lipoprotein (LDL). When grown in medium devoid of lipoproteins, the cells developed a minimal enhancement in steroid secretion in response to ACTH or cholera toxin. However, when LDL was available, steroid secretion was stimulated 4- to 9-fold. To determine the mechanism for this effect, we used LDL in which the protein component was labeled with 125I and the cholesteryl ester component was labeled with [3H]cholesteryl linoleate. These studies demonstrated that the cells derived cholesterol from LDL by binding the lipoprotein at a high affinity receptor site, internalizing it, and hydrolyzing its cholesteryl esters within lysosomes. The resultant free cholesterol was used for steroid synthesis and also acted to suppress the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase and cholesterol synthesis within the cell. LDL receptor activity was enhanced several-fold by treatment of the cells with ACTH or cholera toxin. High density lipoprotein, which did not bind to the LDL receptor, was not degraded with high affinity by the cells and did not support steroid synthesis. The current data suggest that the bovine adrenal cortex can obtain cholesterol for steroid hormone secretion from circulating LDL by means of a high affinity LDL receptor pathway. In a subsequent paper in this series, a similar high affinity LDL-binding site is demonstrated in membranes prepared from fresh bovine adrenocortical tissue.
Assuntos
Córtex Suprarrenal/metabolismo , Colesterol/metabolismo , Lipoproteínas LDL , Lipoproteínas LDL/metabolismo , Receptores de Droga/metabolismo , Córtex Suprarrenal/efeitos dos fármacos , Hormônio Adrenocorticotrópico/farmacologia , Animais , Transporte Biológico , Bovinos , Células Cultivadas , Toxina da Cólera/farmacologia , Ésteres do Colesterol/metabolismo , Humanos , Cinética , Lipoproteínas HDL/metabolismo , Lipoproteínas LDL/sangue , EsteroidesRESUMO
A number of natural cytokines are characterized as having dipeptidyl peptidase (DP) IV susceptible N-terminal peptide sequences. Here we demonstrate that oligopeptides with sequences analogous to the N-terminal part of human IL-1 beta, IL-2, TNF-beta and murine IL-6 were hydrolyzed by purified DP IV and aminopeptidase N (AP-N). The rate of DP IV-catalyzed hydrolysis of these peptides was negatively correlated with their chain length. In contrast to these results, no degradation was found under our conditions for the intact recombinant cytokines, IL-1 alpha, IL-1 beta, IL-2, G-CSF and for natural IL-2, independent of whether DP IV and AP-N were used separately or in combination.
Assuntos
Aminopeptidases/metabolismo , Citocinas/metabolismo , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Peptídeos/metabolismo , Sequência de Aminoácidos , Animais , Antígenos CD13 , Catálise , Citocinas/química , Dipeptidil Peptidase 4 , Humanos , Hidrólise , Camundongos , Dados de Sequência Molecular , Peptídeos/químicaRESUMO
Recent data in the literature suggest that the HIV-1 Tat(1-86) protein exhibits immunosuppressive effects. Moreover, Tat was found to interact with dipeptidyl peptidase IV (DP IV), which is identical to the T cell activation marker CD26. Here we show that the N-terminal amino acid sequence of Tat is essential for the inhibition of DP IV-catalyzed IL-2(1-12) degradation. N-terminal modification of Tat with rhodamine prevented inhibition of enzymatic activity of DP IV as well as suppression of DNA synthesis of mitogen-stimulated human T cells. Moreover, natural peptides containing the X-X-Pro N-terminal motif of Tat also inhibited DP IV activity. These data suggest the existence of endogenous immunomodulatory oligopeptides which influence immune cell proliferation and differentiation via DP IV as does HIV-1 Tat.
Assuntos
Dipeptidil Peptidase 4/metabolismo , Produtos do Gene tat/farmacologia , HIV-1/metabolismo , Ativação Linfocitária , Mitógenos/farmacologia , Inibidores de Proteases/farmacologia , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Linhagem Celular , Produtos do Gene tat/química , Humanos , Interleucina-2/farmacologia , Rim , Mitógenos/antagonistas & inibidores , Dados de Sequência Molecular , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Relação Estrutura-Atividade , Suínos , Linfócitos T/efeitos dos fármacos , Produtos do Gene tat do Vírus da Imunodeficiência HumanaRESUMO
Various studies have shown that the membrane ectoenzyme dipeptidyl peptidase IV (DP IV, CD26), expressed on T, NK, and B cells in the human immune system, is involved in the regulation of DNA synthesis and cytokine production. Here, we clearly demonstrate that this enzyme is highly expressed also on human epidermal foreskin and split-skin keratinocytes and that the specific DP IV inhibitors Lys[Z(NO2)]-thiazolidide, Lys[Z(NO2)]-pyrrolidide inhibit the enzymatic activity as well as the DNA synthesis of these cells. These data demonstrate that CD26 plays a role also in regulation of DNA synthesis of epidermal keratinocytes and that the enzymatic activity is required for mediating these effects.