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Characterization of the major 68 kDa heat shock protein in a rat transformed astroglial cell line.

The heat shock response in a transformed astrocyte line was compared with nontransformed astrocytes. The synthesis of HSP 68, the major inducible heat shock protein (HSP 68) was induced by a non-lethal 45 degrees C, 10 min heat shock. Although the incorporation of [35S]methionine into HSP 68 suggested that similar amounts of protein were being synthesized after heat shock, Western immunoblotting demonstrated striking differences in the HSP immunostaining between the two cell types. By one- and 'two-dimensional gel electrophoresis the major 68 kDa heat shock protein (HSP 68) was similar in both

Dose-related effects of the hepatocarcinogen, Wy-14,643, on peroxisomes and cell replication.

The dose and time dependency of peroxisome proliferation and hepatocyte replication was evaluated in the liver of rats fed the peroxisome pr … to 1000 ppm throughout the treatment period. Hepatocellular replication, evaluated by nuclear histoautoradiography ([3H]thymidine labeling, 6-day infusion), was increased in all Wy-14,643 dose groups after 1 week of treatment (5 ppm, 4-fold; 10 ppm, 5-fold; 50 ppm, 13-fold; 100 ppm, 12-fold; and 1000 ppm, 13-fold over controls). However, in 5 and 10 ppm groups t …

Complementation of DNA repair in xeroderma pigmentosum group A cell extracts by a protein with affinity for damaged DNA.

Complementation group A of xeroderma pigmentosum (XP) represents one of the most prevalent and serious forms of this cancer-prone disorder. Because of a marked defect in DNA excision repair, cells from individuals with XP-A are hypersensitive to the toxic and mutagenic effects of ultraviolet light and many chemical agents. We report here the isolation of the XP-A DNA repair protein by complementation of cell extracts from a repair-defective human XP-A cell line. XP-A protein purified from calf thymus migrates on denaturing gel electrophoresis as a doublet of 40 and 42 kilodaltons. The XP-A pro

Amyotrophic lateral sclerosis: alterations in neurotransmitter receptors.

Loss of motor neurons is the primary pathological hallmark of amyotrophic lateral sclerosis. Drug and neurotransmitter receptors are neuronal markers and can be indicators of neuronal connectivity. Knowledge of alterations in receptors in amyotrophic lateral sclerosis should contribute to our understanding of normal spinal cord neurotransmitter systems as well as of the pathophysiology of amyotrophic lateral sclerosis. We therefore used a sensitive, light microscopic in vitro labeling receptor autoradiographic technique to map and quantitate muscarinic cholinergic, glycinergic, and benzodiazep

Quantitative cytochemistry of glycogen in blood cells. Methods and clinical application.

Quantitative glycogen determinations can be made in single blood and bone marrow cells, using microspectrophotometry or microfluorometry after staining with variants of the periodic acid--Schiff (PAS) reaction. These PAS variant reactions generally do not indicate the presence of non-glycogen PAS-positive substances, known to be prevalent in various hematopoietic cells, possibly due to masking of reactive groups. The specificity of the reaction in blood cells was ascertained by alpha-amylase digestion, which removed more than 95% of the PAS-positive material. Calibration of the PAS reaction wa

Selective migration of lymphocytes within the mouse small intestine.

The factors which determine the migration of lymphoid cells to lamina propria or Peyer's patches of mouse small intestine have been investigated by autoradiographic tracing of intravenously injected spleen, thymus and lymph node cells. The numbers of labelled cells found in antigen-free grafts of foetal small intestine were compared with the numbers in normally sited gut. Thymus, normal spleen and B spleen lymphocytes, labelled with [3H]adenosine or [5-3H]uridine, were confined to Peyer's patches in normal and grafted gut. [3H]Thymidine-labelled lymphoblasts from the mesenteric nodes of young

RNA метилтрансферазы METTL8 устанавливает m3C32 в митохондриальном tRNASThr/Ser (UCN) для оптимизации структуры trNA и митохондриального перевода.

Модифицированные нуклеотиды в tRNA являются важными определяющими факторами сворачивания, структуры и функции. Здесь мы идентифицируем METTL8 как митохондриальный белок и активный РНК метилтрансферазы, ответственный за установку m3C32 в митохондриале человека (mt-)tRNAThr и mt-tRNAser (UCN). METTL8 имеет перекрестные связи с антикодоновой стволовой петлей (ASL) многих мт-тРНК в клетках, в связи с чем возникает вопрос о том, каким образом достигается целевая специфичность метилирования. Расчленение элементов распознавания mt-tRNA выявило наличие U34G35 и t6A37/(ms2)i6A37, которые одновременно п

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… RNA was also separated by denaturing PAGE, stained with ethidium bromide (EtBr) and labelled RNAs ( 3 H-Me) were detected by autoradiography. … RNA was also separated by denaturing PAGE, stained with ethidium bromide (EtBr) and labelled RNAs ( 3 H-Me) were detected by autoradiography. …

Transduction of light in the suprachiasmatic nucleus: evidence for two different neurochemical cascades regulating the levels of Per1 mRNA and pineal melatonin.

The suprachiasmatic nucleus (SCN) contains a circadian clock and regulates melatonin synthesis in the pineal gland. Light exposure during the subjective night acutely increases the mRNA levels of the Period (Per)1 gene in the SCN and acutely suppresses melatonin levels in the pineal gland. Activation of N-methyl-D-aspartate (NMDA) receptors in the SCN has been demonstrated to phase-shift the circadian clock in a manner similar to light. We tested the hypothesis that activation of excitatory amino acid (EAA) receptors in the SCN mediates the acute effects of light on Per1 mRNA levels and pineal

Early direct and transneuronal effects in mice with targeted expression of a toxin gene to D1 dopamine receptor neurons.

The neurochemical profile was examined at postnatal day 3-4 in mutant mice generated by in vivo Cre mediated activation of an attenuated diphtheria toxin gene inserted into the D1 dopamine receptor gene locus. An earlier study of this model had shown that D1 dopamine receptor, substance P and dynorphin were not expressed in the striatum. Quantitative in situ hybridization analysis showed an increase in D2 dopamine receptor and enkephalin messenger RNA expression. The nigrostriatal pathway in the mutant pups was intact with a normal number of dopaminergic neurons in the substantia nigra and the

Characterization of the human peroxisome proliferator activated receptor delta gene and its expression.

Peroxisome proliferator activated receptors (PPARs) are nuclear receptors regulating the expression of genes involved in lipid and glucose metabolism. Three different PPARs; alpha (PPARA), gamma (PPARG) and delta (PPARD) have been characterized and they are distinguished from each other by tissue distribution and cell activation. In this study, the structure and detailed chromosomal localization of the human PPARD gene was determined. Three genomic clones containing the PPARD gene was isolated from a human P1 library. The gene spans approximately 85 kb of DNA and consists of 9 exons and 8 intr

Disposition in rats of N-pyridinium-propyl-cyclam, N-triethylammonium-propyl-cyclam, and N-[Triethylammonium]-3-propyl-[15]ane-N5, potential cartilage imaging agents.

Quaternary ammonium compounds are known to highly concentrate in articular cartilages after i.v. administration. This property was used to synthesize new potential radiodiagnostic agents for joint imaging. Pharmacokinetic study was performed in rats for three new compounds: N-pyridinium-propyl-cyclam (NPPC), N-triethylammonium-propyl-cyclam (NTPC), and N-[triethylammonium]-3-propyl-[15]ane-N5 (NTP 15-5). After i.v. administration, [(3)H]NPPC and [(3)H]NTPC highly and rapidly concentrated in articular cartilage, this uptake being followed by a single exponential decrease with half-lives of, res

Chemical cross-linking of the substance P (NK-1) receptor to the alpha subunits of the G proteins Gq and G11.

We have previously shown that the high-affinity binding of substance P (SP) to its receptor is dependent on an interaction with a PTX-insensitive G protein. This G protein couples SP receptor activation to stimulation of its effector, phospholipase C. In this study, we combined photoaffinity labeling, chemical cross-linking techniques, and immunological characterization using sequence-specific antibody probes to identify G proteins that couple to the SP receptor. First we covalently labeled the SP receptor present on rat submaxillary gland membranes with a radioiodinated photoreactive derivati

Formation and persistence of benzo[a]pyrene-DNA adducts in mouse epidermis in vivo: importance of adduct conformation.

The formation and repair of benzo[a]pyrene diol epoxide-N2-deoxyguanosine adducts (BPDE-N2-dG) in DNA isolated from the skin of mice treated topically with benzo[a]pyrene (BP) was studied by 32P-postlabeling and by low-temperature fluorescence spectroscopy under low resolution and under high resolution fluorescence line narrowing (FLN) conditions. In agreement with earlier studies, total BP-DNA binding reached a maximum at 24 h after treatment (dose: 1 mumol/mouse), then declined rapidly until 4 days after treatment and much more slowly thereafter. An HPLC method was developed which resolved t