scholarly journals Analysis of naturally occurring delayed-type hypersensitivity reactions in leprosy by in situ hybridization.

1989 ◽  
Vol 169 (5) ◽  
pp. 1565-1581 ◽  
Author(s):  
C L Cooper ◽  
C Mueller ◽  
T A Sinchaisri ◽  
C Pirmez ◽  
J Chan ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fold Increase ◽  
Delayed Type Hypersensitivity ◽  
Cell Mediated Immunity ◽  
Serine Esterase ◽  
Ifn Gamma ◽  
Cytotoxic Cells ◽  
Selective Increase ◽  
Reversal Reaction

Analysis of tissue lesions of the major reactional states of leprosy was undertaken to study the immune mechanisms underlying regulation of cell-mediated immunity and delayed-type hypersensitivity (DTH) in man. In situ hybridization hybridization of reversal reaction biopsy specimens for INF-gamma mRNA expression revealed a 10-fold increase in specific mRNA-containing cells over that observed in unresponsive lepromatous patients. Expression of huHF serine esterase, a marker for T cytotoxic cells, were fourfold increased in reversal reaction and tuberculoid lesions above that detected in unresponsive lepromatous individuals. Immunohistology of reversal reactions confirmed a selective increase of Th and T cytotoxic cells in the cellular immune response. Of interest, the microanatomic location of these serine esterase mRNA-containing cells was identical to the distribution of CD4+ cells. Analysis of erythema nodosum leprosum (ENL) lesions revealed differences in the underlying immune processes in comparison with reversal reaction lesions. Although phenotypic Th cells predominated in ENL lesions, IFN-gamma and serine esterase gene expression were markedly reduced. We suggest that reversal reactions represent a hyperimmune DTH response characterized by a selective increase of CD4+ IFN-gamma producing cells and T cytotoxic cells, which result in the clearing of bacilli and concomitant tissue damage. In contrast, ENL reactions may be viewed as a transient diminution of Ts cells and activity leading to a partial and transient augmentation in cell-mediated immunity, perhaps sufficient to result in antibody and immune complex formation, but insufficient to clear bacilli from lesions.

scholarly journals Galanin-Like Peptide (GALP) Is a Target for Regulation by Leptin in the Hypothalamus of the Rat

Endocrinology ◽  
2000 ◽  
Vol 141 (7) ◽  
pp. 2703-2706 ◽  
Author(s):  
Anders Juréus ◽  
Matthew J. Cunningham ◽  
Molly E. McClain ◽  
Donald K. Clifton ◽  
Robert A. Steiner
Keyword(s):  
In Situ Hybridization ◽  
Leptin Receptor ◽  
Fold Increase ◽  
Female Rats ◽  
Dorsomedial Nucleus ◽  
Physiological Regulation ◽  
Tissue Sections ◽  
Galanin Receptors ◽  
Neuroendocrine Functions

Galanin-like peptide (GALP), which was recently isolated from the porcine hypothalamus, shares sequence homology with galanin and binds with high affinity to galanin receptors. To study the distribution and regulation of GALP-expressing cells in the brain, we cloned a 120 base-pair cDNA fragment of rat GALP and produced an antisense riboprobe. In situ hybridization for GALP mRNA was then performed on tissue sections throughout the forebrain of adult ovariectomized female rats. We found GALP mRNA-containing cells in the arcuate nucleus (Arc), caudal dorsomedial nucleus, median eminence and the pituitary. Because GALP mRNA in the Arc appeared to overlap with the known distribution of leptin receptor mRNA, we tested the hypothesis that GALP expression is regulated by leptin. Using in situ hybridization, we compared the number of GALP mRNA-containing cells among groups of rats that were fed ad lib or fasted for 48 h and treated with either leptin or vehicle. Fasting reduced the number of identifiable cells containing GALP mRNA in the Arc, whereas the treatment of fasted animals with leptin produced a 4-fold increase in the number of cells expressing GALP message. The presence of GALP mRNA in the hypothalamus and pituitary and its regulation by leptin suggests that GALP may have important neuroendocrine functions, including the physiological regulation of feeding, metabolism, and reproduction.

scholarly journals Cell-mediated immunity: delayed-type hypersensitivity and cytotoxic responses are mediated by different T-cell subclasses.

1976 ◽  
Vol 143 (6) ◽  
pp. 1534-1539 ◽  
Author(s):  
B Huber ◽  
O Devinsky ◽  
R K Gershon ◽  
H Cantor
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Division Of Labor ◽  
Delayed Type Hypersensitivity ◽  
Cell Mediated Immunity ◽  
Killer Activity ◽  
Cytotoxic Cells ◽  
Relative Contribution ◽  
Helper Function ◽  
Cytotoxic Responses

Cell-mediated immunity includes both the generation of cytotoxic cells and initiation of delayed-type hypersensitivity (DTH). The resting T-cell population, before stimulation by antigen, already contains cells of the Lyl subclass that are programmed to initiate DTH (and helper function) but not cytotoxic responses, as well as Ly23 cells which can generate killer activity (and suppressive function) but not DTH. The central implication of these findings is that the broad division between humoral and cell-mediated immune responses does not precisely correspond to the division of labor among T-cell subclasses. The relative contribution of DTH-competent Lyl cells and cytotoxic Ly23 cells to the classical homograft response remains to be determined.

Cholinergic agonist-induced pepsinogen secretion from murine gastric chief cells is mediated by M1 and M3 muscarinic receptors

2005 ◽  
Vol 289 (3) ◽  
pp. G521-G529 ◽  
Author(s):  
Guofeng Xie ◽  
Cinthia Drachenberg ◽  
Masahisa Yamada ◽  
Jürgen Wess ◽  
Jean-Pierre Raufman
Keyword(s):  
In Situ Hybridization ◽  
Fold Increase ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Double Knockout ◽  
Gastric Glands ◽  
Cholinergic Agonist ◽  
Chief Cells ◽  
Pepsinogen Secretion

Muscarinic cholinergic mechanisms play a key role in stimulating gastric pepsinogen secretion. Studies using antagonists suggested that the M3 receptor subtype (M3R) plays a prominent role in mediating pepsinogen secretion, but in situ hybridization indicated expression of M1 receptor (M1R) in rat chief cells. We used mice that were deficient in either the M1 (M1R−/−) or M3 (M3R−/−) receptor or that lacked both receptors (M1/3R−/−) to determine the role of M1R and M3R in mediating cholinergic agonist-induced pepsinogen secretion. Pepsinogen secretion from murine gastric glands was determined by adapting methods used for rabbit and rat stomach. In wild-type (WT) mice, maximal concentrations of carbachol and CCK caused a 3.0- and 2.5-fold increase in pepsinogen secretion, respectively. Maximal carbachol-induced secretion from M1R−/− mouse gastric glands was decreased by 25%. In contrast, there was only a slight decrease in carbachol potency and no change in efficacy when comparing M3R−/− with WT glands. To explore the possibility that both M1R and M3R are involved in carbachol-mediated pepsinogen secretion, we examined secretion from glands prepared from M1/3R−/− double-knockout mice. Strikingly, carbachol-induced pepsinogen secretion was nearly abolished in glands from M1/3R−/− mice, whereas CCK-induced secretion was not altered. In situ hybridization for murine M1R and M3R mRNA in gastric mucosa from WT mice revealed abundant signals for both receptor subtypes in the cytoplasm of chief cells. These data clearly indicate that, in gastric chief cells, a mixture of M1 and M3 receptors mediates cholinergic stimulation of pepsinogen secretion and that no other muscarinic receptor subtypes are involved in this activity. The development of a murine secretory model facilitates use of transgenic mice to investigate the regulation of pepsinogen secretion.

Translocation t(11;14)(q13;q32) is the hallmark of IgM, IgE, and nonsecretory multiple myeloma variants

Blood ◽  
2003 ◽  
Vol 101 (4) ◽  
pp. 1570-1571 ◽  
Author(s):  
Hervé Avet-Loiseau ◽  
Richard Garand ◽  
Laurence Lodé ◽  
Jean-Luc Harousseau ◽  
Régis Bataille
Keyword(s):  
Multiple Myeloma ◽  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Fold Increase ◽  
Major Interest ◽  
High Incidence ◽  
Very High

In an attempt to address the issue of cytogenetic features of multiple myeloma (MM) variants, we have analyzed a series of 8 IgM, 9 IgD, 2 IgE, and 14 nonsecretory (NS) MM cases using fluorescence in situ hybridization. A very high incidence (83%) of t(11;14)(q13;q32) was detected in the IgM (7 of 8), IgE (2 of 2), and NS (11 of 14) MM cases, but not in the IgD cases (2 of 9). Of note, no t(4;14) was observed in this cohort of patients. This increased incidence of t(11;14) was associated with 2 dominant features in these variants, namely, a “lymphoplasmacytic” presentation mainly in IgM MM and a lower secreting capacity in the others, 2 features previously associated with t(11;14). Of major interest, t(11;14) was never observed in Waldenström macroglobulinemia or in IgG/IgA “lymphoplasmacytic” lymphomas. Thus, for unknown reasons, t(11;14) is the hallmark of IgM, IgE, and NS MM, (but not IgD MM), with a 5-fold increase of its incidence compared to that of IgG and IgA MM.

scholarly journals Regression of an established tumor genetically modified to release granulocyte colony-stimulating factor requires granulocyte-T cell cooperation and T cell-produced interferon gamma.

1993 ◽  
Vol 178 (1) ◽  
pp. 151-161 ◽  
Author(s):  
A Stoppacciaro ◽  
C Melani ◽  
M Parenza ◽  
A Mastracchio ◽  
C Bassi ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
T Cell ◽  
Tumor Regression ◽  
Tnf Alpha ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Ifn Gamma ◽  
Established Tumor ◽  
Stimulating Factor

Using the murine colon adenocarcinoma C-26 cell line, engineered to release granulocyte colony-stimulating factor (G-CSF) (C-26/G-CSF), were studied the mechanisms responsible for inhibition of tumor take in syngeneic animals and of regression of an established tumor in sublethally irradiated mice injected with these cells. Immunocytochemistry and in situ hybridization, performed to characterize tumor-infiltrating leukocytes and their cytokine expression, respectively, indicated that polymorphonuclear leukocytes (PMN) were the major cells responsible for inhibition of tumor take and that they expressed mRNA for interleukin 1 alpha (IL-1 alpha), IL-1 beta, and tumor necrosis factor alpha (TNF-alpha). Expression of interferon gamma (IFN-gamma) and of IL-4 was undetectable, consistent with the absence of T lymphocytes at the site of tumor injection. In mice injected with C-26/G-CSF cells after 600-rad irradiation, the tumors grew to approximately 1.5 cm in 30 d, regressing completely thereafter in 70-80% of mice. During the growing phase, tumors were infiltrated first by PMN (between days 15 and 20), then by macrophages, and last by T lymphocytes. Both CD4+ and CD8+ T cells were present but only CD8 depletion significantly abrogated tumor regression. Depletion of PMN by the RB6-8C5 antigranulocytes monoclonal antibody reduced the number of T cells infiltrating the tumor and prevented tumor regression. In situ hybridization performed at the beginning of tumor regression revealed the presence of mRNA for IL-1 alpha, IL-1 beta, and TNF-alpha, but also the presence of cells, with lymphoid morphology, expressing IFN-gamma. Tumors from mice treated with recombinant IFN-gamma (between days 20 and 35) were rejected faster, whereas mice treated with antibodies to IFN-gamma (from day 20) died of progressive tumor. Cyclosporin A treatment (started at day 20) also abrogated tumor regression. These results indicate that inhibition of tumor take and regression in this model occurs through different mechanisms that involve PMN and PMN-T cell interactions, respectively, as well as a combination of cytokines that, for tumor regression, require IFN-gamma. Thus, gene transfer of a single cytokine gene such as G-CSF into tumor cells appears to be sufficient to trigger the cascade of cell interactions and cytokine production necessary to destroy a cancer nodule.

scholarly journals Medial Amygdala Kiss1 Neurons Mediate Female Pheromone Stimulation of Luteinizing Hormone in Male Mice

2018 ◽  
Vol 108 (3) ◽  
pp. 172-189 ◽  
Author(s):  
Sanya Aggarwal ◽  
Celion Tang ◽  
Kristen Sing ◽  
Hyun Wook Kim ◽  
Robert P. Millar ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Luteinizing Hormone ◽  
Fold Increase ◽  
Reproductive Physiology ◽  
Medial Amygdala ◽  
Male Mice ◽  
Reproductive Axis ◽  
Female Urine ◽  
Olfactory Stimuli

Background/Aims: The medial amygdala (MeA) responds to olfactory stimuli and alters reproductive physiology. However, the neuronal circuit that relays signals from the MeA to the reproductive axis remains poorly defined. This study aimed to test whether MeA kisspeptin (MeAKiss) neurons in male mice are sensitive to sexually relevant olfactory stimuli and transmit signals to alter reproductive physiology. We also investigated whether MeAKiss neurons have the capacity to elaborate glutamate and GABA neurotransmitters and potentially contribute to reproductive axis regulation. Methods: Using female urine as a pheromone stimulus, MeAKiss neuronal activity was analysed and serum luteinizing hormone (LH) was measured in male mice. Next, using a chemogenetic approach, MeAKiss neurons were bi-directionally modulated to measure the effect on serum LH and evaluate the activation of the preoptic area. Lastly, using in situ hybridization, we identified the proportion of MeAKiss neurons that express markers for GABAergic (Vgat) and glutamatergic (Vglut2) neurotransmission. Results: Male mice exposed to female urine showed a two-fold increase in the number of c-Fos-positive MeAKiss neurons concomitant with raised LH. Chemogenetic activation of MeAKiss neurons significantly increased LH in the absence of urine exposure, whereas inhibition of MeAKiss neurons did not alter LH. In situ hybridization revealed that MeAKiss neurons are a mixed neuronal population in which 71% express Vgat mRNA, 29% express Vglut2 mRNA, and 6% express both. Conclusions: Our results uncover, for the first time, that MeAKiss neurons process sexually relevant olfactory signals to influence reproductive hormone levels in male mice, likely through a complex interplay of neuropeptide and neurotransmitter signalling.

scholarly journals Disruption of Brachypodium Lichenase Alters Metabolism of Mixed-linkage Glucan and Starch

2021 ◽  
Author(s):  
Mingzhu Fan ◽  
Jacob Krüger Jensen ◽  
Starla Zemelis-Durfee ◽  
Sang-Jin Kim ◽  
Jia-Yi Chan ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Brachypodium Distachyon ◽  
Fold Increase ◽  
Wild Type ◽  
Gene Encoding ◽  
Vegetative Tissues ◽  
Hybridization Data ◽  
Rna In Situ Hybridization ◽  
Germinating Seeds

Mixed-linkage glucan (MLG), which is widely distributed in grasses, is a polysaccharide highly abundant in cell walls of grass endosperm and young vegetative tissues. Lichenases are enzymes that hydrolyze MLG first identified in MLG-rich lichens. In this study, we identify a gene encoding a lichenase we name Brachypodium distachyon LICHENASE 1 (BdLCH1), which is highly expressed in the endosperm of germinating seeds and coleoptiles and at lower amounts in mature shoots. RNA in situ hybridization showed that BdLCH1 is primarily expressed in chlorenchyma cells of mature leaves and internodes. Disruption of BdLCH1 resulted in an eight-fold increase in MLG content in senesced leaves. Consistent with the in situ hybridization data, immunolocalization results showed that MLG was not removed in chlorenchyma cells of lch1 mutants as it was in wild type and implicate the BdLCH1 enzyme in removing MLG in chlorenchyma cells in mature vegetative tissues. We also show that MLG accumulation in lch1 mutants was resistant to dark induced degradation, and eight-week-old lch1 plants showed a faster rate of starch breakdown than wild type in darkness. Our results suggest a role for BdLCH1 in modifying the cell wall to support highly metabolically active cells.

DNA sequence mapping in interphase and metaphase chromosomes by fluorescence in situ hybridization

1992 ◽  
Vol 50 (1) ◽  
pp. 496-497
Author(s):  
Barbara Trask ◽  
Susan Allen ◽  
Anne Bergmann ◽  
Mari Christensen ◽  
Anne Fertitta ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequence ◽  
Dna Sequences ◽  
Dual Band ◽  
Nick Translation ◽  
Metaphase Chromosomes ◽  
Band Pass ◽  
Texas Red ◽  
Fluorescent Spot

Using fluorescence in situ hybridization (FISH), the positions of DNA sequences can be discretely marked with a fluorescent spot. The efficiency of marking DNA sequences of the size cloned in cosmids is 90-95%, and the fluorescent spots produced after FISH are ≈0.3 μm in diameter. Sites of two sequences can be distinguished using two-color FISH. Different reporter molecules, such as biotin or digoxigenin, are incorporated into DNA sequence probes by nick translation. These reporter molecules are labeled after hybridization with different fluorochromes, e.g., FITC and Texas Red. The development of dual band pass filters (Chromatechnology) allows these fluorochromes to be photographed simultaneously without registration shift.

Ultrastructural analysis of the spatial distribution of MRNA

1992 ◽  
Vol 50 (1) ◽  
pp. 552-553
Author(s):  
Gary Bassell ◽  
Robert H. Singer
Keyword(s):  
Electron Microscopy ◽  
Spatial Distribution ◽  
In Situ Hybridization ◽  
High Resolution ◽  
Nucleic Acids ◽  
Colloidal Gold ◽  
Dna Probe ◽  
Nucleotide Analogs ◽  
Gold Particles

We have been investigating the spatial distribution of nucleic acids intracellularly using in situ hybridization. The use of non-isotopic nucleotide analogs incorporated into the DNA probe allows the detection of the probe at its site of hybridization within the cell. This approach therefore is compatible with the high resolution available by electron microscopy. Biotinated or digoxigenated probe can be detected by antibodies conjugated to colloidal gold. Because mRNA serves as a template for the probe fragments, the colloidal gold particles are detected as arrays which allow it to be unequivocally distinguished from background.

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