arup sen

Human monocytes are multifaceted cells with a wide range of immunoregulatory functions and distinct secretory products. This manuscript reports on initial attempts to identify specific early macromolecular synthetic events associated with various types of human monocyte activation by observing the patterns of RNA synthesis displayed by human monocytes that are exposed to well characterized activating stimuli. It was found that muramyl dipeptide (MDP), an activator of monocyte-derived fibroblast growth factor (MD-FGF) release from monocytes, also stimulates a reproducible increase in human monocyte total 3H-uridine incorporation and cytoplasmic messenger RNA (mRNA) synthesis at 4 h following activation. In contrast, polyriboinosinic acid:polyribocytidilic acid (poly I:C), an excellent stimulator of monocyte alpha interferon (IFN alpha) release, did not cause a change in either 3H-uridine incorporation or cytoplasmic mRNA production at any of the time points tested. Poly I:C was also found to be a poor stimulator of MD-FGF release. Conversely, MDP did not stimulate any detectable IFN release from human monocytes. The discrepancy between the patterns of macromolecular synthesis observed in human monocytes activated to secrete MD-FGF as compared with IFN indicates that divergent postactivation control mechanisms may be operative at the RNA level in the monocyte following activation of these two distinct functions.

The euryhaline marine yeast Debaromyces hansenii is a model system for the study of processes related to osmoadaptation. In this study, microarray-based gene expression analyses of the entire genome of D. hansenii was used to study its response to osmotic stress. Differential gene expression, compared to control, was examined at three time points (0.5, 3 and 6 h) after exposure of D. hansenii cultures to high salt concentration. Among the 1.72% of genes showing statistically significant differences in expression, only 65 genes displayed at least three-fold increases in mRNA levels after treatment with 2 M NaCl. On the other hand, 44 genes showed three-fold repression. Upregulated as well as the downregulated genes were grouped into functional categories to identify biochemical processes possibly affected by osmotic stress and involved in osmoadaptation. The observation that only a limited number of genes are upregulated in D. hansenii in response to osmotic stress supports the notion that D. hansenii is pre-adapted to survive in extreme saline environments. In addition, since more than one-half of the upregulated genes encode for ribosomal proteins, it is possible that a translational gene regulatory mechanism plays a key role in D. hansenii's osmoregulatory response. Validation studies for ENA1 and for hyphal wall/cell elongation protein genes, using real-time PCR, confirmed patterns of gene expression observed in our microarray experiments. To our knowledge, this study is the first of its kind in this organism and provides the foundation for future molecular studies assessing the significance of the genes identified here in D. hansenii's osmoadaptation.

Human monocytes are multifaceted cells with a wide range of immunoregulatory functions and distinct secretory products. This manuscript reports on initial attempts to identify specific early macromolecular synthetic events associated with various types of human monocyte activation by observing the patterns of RNA synthesis displayed by human monocytes that are exposed to well characterized activating stimuli. It was found that muramyl dipeptide (MDP), an activator of monocyte-derived fibroblast growth factor (MD-FGF) release from monocytes, also stimulates a reproducible increase in human monocyte total 3H-uridine incorporation and cytoplasmic messenger RNA (mRNA) synthesis at 4 h following activation. In contrast, polyriboinosinic acid:polyribocytidilic acid (poly I:C), an excellent stimulator of monocyte alpha interferon (IFN alpha) release, did not cause a change in either 3H-uridine incorporation or cytoplasmic mRNA production at any of the time points tested. Poly I:C was also ...

The euryhaline marine yeast Debaromyces hansenii is a model system for the study of processes related to osmoadaptation. In this study, microarray-based gene expression analyses of the entire genome of D. hansenii was used to study its response to osmotic stress. Differential gene expression, compared to control, was examined at three time points (0.5, 3 and 6 h) after exposure of D. hansenii cultures to high salt concentration. Among the 1.72% of genes showing statistically significant differences in expression, only 65 genes displayed at least three-fold increases in mRNA levels after treatment with 2 M NaCl. On the other hand, 44 genes showed three-fold repression. Upregulated as well as the downregulated genes were grouped into functional categories to identify biochemical processes possibly affected by osmotic stress and involved in osmoadaptation. The observation that only a limited number of genes are upregulated in D. hansenii in response to osmotic stress supports the notion that D. hansenii is pre-adapted to survive in extreme saline environments. In addition, since more than one-half of the upregulated genes encode for ribosomal proteins, it is possible that a translational gene regulatory mechanism plays a key role in D. hansenii's osmoregulatory response. Validation studies for ENA1 and for hyphal wall/cell elongation protein genes, using real-time PCR, confirmed patterns of gene expression observed in our microarray experiments. To our knowledge, this study is the first of its kind in this organism and provides the foundation for future molecular studies assessing the significance of the genes identified here in D. hansenii's osmoadaptation.

Affinity chromatography with actin-Sepharose conjugates of purified human fibronectin, normal human plasma, or serum-free culture fluid from human fibroblasts showed that fibronectin molecules can directly bind to actin. A quantitative recovery of soluble human fibronectin was accomplished by chromatography on actin immobilized on Sepharose beads. Human fibronectin molecules bound to actin-Sepharose were eluted with 0.25--0.35 M potassium bromide, and these molecules competed in a species-specific radioimmunoassay for human fibronectin. The subunits of fibronectin isolated by actin-Sepharose chromatography comigrated in SDS polyacrylamide gel electrophoresis with those of electrophoretically homogeneous fibronectin purified by conventional procedures. The efficient direct binding of fibronectin to actin suggests that interactions between these proteins might also take place in vivo but further studies are needed to elucidate the biological significance of this affinity.

The binding of type C viral p12 proteins to purified viral RNA has been examined in vitro with the use of a family of closely related infectious primate type C viruses--the woolly monkey (SSAV) and gibbon (GALV) group. This in vitro protein-RNA binding is type specific. The system should serve as a model for studies of the evolution of nucleic acid binding proteins.

A type C virus (OMC-1) detected in a culture of owl monkey kidney cells resembled typical type C viruses morphologically, but was slightly larger than previously characterized mammalian type C viruses. OMC-1 can be transmitted to bat lung cells and cat embryo fibroblasts. The virions band at a density of 1.16 g/ml in isopycnic sucrose density gradients and contain reverse transcriptase and a 60-65S RNA genome composed of approximately 32S subunits. The reverse transcriptase is immunologically and biochemically distinct from the polymerases of othe retroviruses. Radioimmunoassays directed to the interspecies antigenic determinants of the major structure proteins of other type C viruses do not detect a related antigen in OMC-1. Nucleic acid hybridization experiments using labeled viral genomic RNA or proviral cDNA transcripts to normal cellular DNA of different species show that OMC-1 is an endogenous virus with multiple virogene copies (20-50 per haploid genome) present in normal owl monkey cells and is distinct from previously isolated type C and D viruses. Sequences related to the OMC-1 genome can be detected in other New World monkeys. Thus, similar to the Old World primates (e.g., baboons as a prototype), the New World monkeys contain endogenous type C viral genes that appear to have been transmitted in the primate germ line.

The binding of type C viral p12 proteins to purified viral RNA has been examined in vitro with the use of a family of closely related infectious primate type C viruses-the woolly monkey (SSAV) and gibbon (GALV) group. This in vitro protein-RNA binding is type specific. The system should serve as a model for studies of the evolution of nucleic acid binding proteins.

Irritable bowel syndrome (IBS) is one of the most common disorders seen by gastroenterologists. Visceral hypersensitivity is now well recognized as a clinical marker for the disease. Intrarectal lidocaine has been previously shown to decrease pain report from rectal distension in patients with IBS without any significant serum lidocaine levels. We conducted a prospective, double-blind, crossover trial on 10 patients with IBS to evaluate the effects of 300 mg intrarectal lidocaine jelly on abdominal pain. Ten Caucasian premenopausal women who met the Rome II criteria for diarrhea-predominant IBS were recruited into the study. All of the patients that participated had intermittent left lower quadrant pain and diarrhea. Each patient participated in 2 sessions in which saline jelly (placebo) and lidocaine jelly was administered on a double-blind, crossover basis. Patients participated in these sessions at a time when their ongoing pain was at least 3 on a 0 to 10 visual analogue scale. In comparison to placebo saline jelly, lidocaine jelly significantly decreased abdominal pain (P < .02) for at least 4 hours. None of the patients experienced any side effects. Intrarectal lidocaine may be a potentially useful treatment for chronic abdominal pain in IBS. The possible presence of abnormal sodium channels in the rectal and or colonic visceral afferents of patients with IBS might serve as a clue as to the effectiveness of rectal lidocaine. The dose of lidocaine used in this study may be of sufficient strength to normalize aberrant sodium channels that may be present in the colon of patients with IBS without affecting normal sodium channels of either IBS or control subjects.

Visual and quantitative monitoring of cell-to-cell variation in the expression of manganese superoxide dismutase (MnSOD) mRNA by using novel ratiometric imaging with molecular beacons (MB) reveals a distinct change in patterns following induction of human breast-carcinoma cells with lipopolysaccharide. Interestingly, the pattern of cell-to-cell variation in a cell line stably transfected with a plasmid bearing a cDNA clone of MnSOD and overproducing the enzyme is significantly different from the pattern associated with MnSOD induction. The levels and the patterns of cell-population heterogeneity for beta-actin mRNA expression do not show distinct changes either following induction or in stably transfected cells. These results are significant in light of the reported relationship between this enzyme and malignant phenotype of breast-carcinoma cells. Use of MBs in ratiometric image analyses for cytoplasmic mRNAs represents a novel means of directly examining the stochasticity of transcription of MnSOD and other genes implicated in cellular phenotype regulation.

A structural protein purified from the Rous sarcoma virus (RSV) can specificially bind in vitro to purified avian, but not mammalian, type C viral RNA. Following ultraviolet irradiation of viral particles under conditions which stabilize the polyploid 70S viral RNA, the same polypeptide can be directly purified from the RSV genome. Based on its electrophoretic mobility in polyacrylamide gels containing sodium dodecylsulfate, the RNA binding protein has been identified as the major phosphoprotein (p19) of avian type C viruses. Similar experiments show that the major phosphoproteins of mammalian type C viruses (p12 for murine viruses and p16 for endogenous primate viruses) are also the specific RNA binding proteins and, similarly, are found closely associated with the 70S RNA genomes in the intact viral particles.

A type C virus (OMC-1) detected in a culture of owl monkey kidney cells resembled typical type C viruses morphologically, but was slightly larger than previously characterized mammalian type C viruses. OMC-1 can be transmitted to bat lung cells and cat embryo fibroblasts. The virions band at a density of 1.16 g/ml in isopycnic sucrose density gradients and contain reverse transcriptase and a 60-65S RNA genome composed of approximately 32S subunits. The reverse transcriptase is immunologically and biochemically distinct from the polymerases of othe retroviruses. Radioimmunoassays directed to the interspecies antigenic determinants of the major structure proteins of other type C viruses do not detect a related antigen in OMC-1. Nucleic acid hybridization experiments using labeled viral genomic RNA or proviral cDNA transcripts to normal cellular DNA of different species show that OMC-1 is an endogenous virus with multiple virogene copies (20-50 per haploid genome) present in normal owl monkey cells and is distinct from previously isolated type C and D viruses. Sequences related to the OMC-1 genome can be detected in other New World monkeys. Thus, similar to the Old World primates (e.g., baboons as a prototype), the New World monkeys contain endogenous type C viral genes that appear to have been transmitted in the primate germ line.

... X30,000. One molecule is shown with a magnification of about X10D,009. 488 JAENISCH ANDLEVINE oligomers were observed. For the formu lation of the criteria to distinguish circular and catenated oligomers on micrographs, see Clayton et al. (1968) and Gordon et al. ...