Muscle phenotype is regulated by mechanical forces. However, it is not well understood how these forces are translated into intracellular signalling that influences gene expression. The purpose of this study was to test the hypothesis that muscles displaying a wide range of metabolic profiles and fibre-type composition exhibit differences in the detection and transmission of mechanical stimuli. A mechanical challenge in the form of passive stretch normalized to 3 N/g muscle weight was applied to the rat extensor digitorum longus (EDL), soleus (SOL), and plantaris (PLN) in situ for 5 min, following which activities of the mechanically-responsive p54 c-jun NH(2)-terminal kinase (JNK) and extracellular-regulated kinase (ERK) 1/2 were measured. EDL, SOL, and PLN were not different in their stretch-induced JNK (4.5, 5.2 and 6-fold baseline, respectively) or ERK (2.2, 2.2 and 1.9-fold baseline, respectively) responses, in spite of differing fibre-type compositions. The medial gastrocnemius (MG), a compartmentalized muscle with red (MGr) and white (MGw) regions, was subjected to the same normalized mechanical stretch protocol. The resulting JNK and ERK activities were significantly higher in MGr (13 and 4.5-fold baseline, respectively) than in MGw (5 and 1.2-fold baseline, respectively) and all other muscles. In contrast to stimulation by passive stretch, stimulation of the MG by isometric contractile activity did not result in a heterogeneous response between compartments. This study demonstrates an absence of difference among muscles of varying phenotype in their ability to transmit mechanical stimuli to the mitogen-activated protein kinase signalling pathways, and hence in their mechanosensitivity. Furthermore, the results highlight the importance of considering aspects of the functional organization of different muscles, such as compartmentalization and architecture, when studying mechanical signalling in vivo. 相似文献
A Hodgkin cell-specific antigen detected by the monoclonal antibody Ki- 1 was found on T helper lymphocytes after activation by autologous and allogeneic stimulator cells. About 50% of lymphoblasts generated by auto- and alloactivation reacted with the antibody. In contrast, only less than 6% of lymphoblasts stimulated with Con-A, phytohemagglutinin (PHA), or protein A, and none of lymphoblasts activated by oxidative mitogenesis, expressed this antigen. Among several permanent cell lines tested, the K562, MOLT-4, HL-60, and EBV transformed B lymphoblastoid cells reacted with the Ki-1 antibody. The results may indicate possible relationships between the autoreactive subset of T lymphocytes and the pathogenesis of Hodgkin's disease. 相似文献
Paroxysmal kinesigenic dyskinesia is an episodic movement disorder caused by dominant mutations in the proline-rich transmembrane protein PRRT2, with onset in childhood and typically with improvement or resolution by middle age. Mutations in the same gene may also cause benign infantile seizures, which begin in the first year of life and typically remit by the age of 2 years. Many details of PRRT2 function at the synapse, and the effects of mutations on neuronal excitability in the pathophysiology of epilepsy and dyskinesia, have emerged through the work of several groups over the last decade. However, the age dependence of the phenotypes has not been explored in detail in transgenic models. Here, we report our findings in heterozygous and homozygous Prrt2 knockout mice that recapitulate the age dependence of dyskinesia seen in the human disease. We show that Prrt2 deletion reduces the levels of synaptic proteins in a dose-dependent manner that is most pronounced at postnatal day 5 (P5), attenuates at P60, and disappears by P180. In a test for foot slippage while crossing a balance beam, transient loss of coordination was most pronounced at P60 and less prominent at age extremes. Slower traverse time was noted in homozygous knockout mice only, consistent with the ataxia seen in rare individuals with biallelic loss of function mutations in Prrt2. We thus identify three age-dependent phenotypic windows in the mouse model, which recapitulate the pattern seen in humans with PRRT2-related diseases.
A sensitive and precise radioreceptor assay for determining plasma levels of human factor VIII/von Willebrand's factor (FVIII/vWF) has been developed by taking advantage of the FVIII/vWF receptor sites on human platelets. Paraformaldehyde-fixed platelets, which were processed and then stored, retained FVIII/vWF binding activity and therefore could be used as a convenient source of receptors. The human plasma samples to be tested were initially filtered on 4% agarose columns to concentrate the FVIII/vWF protein in the void volume and to remove the factor(s) that interferes with the assay. The percent recovery of FVIII/vWF in the pooled eluent was measured by the recovery of added trace 125I-FVIII/vWF. The coefficients of intra- and interassay variation were 6% and 10%, respectively. The plasma FVIII/vWF concentrations determined by the assay for pooled normal plasma, hemophilia A plasma, and plasmas from two patients with von Willebrand's disease were 16.3 +/- 0.5, 52.6 +/- 1.5, 6.8 +/- 0.8, and 3.2 +/- 0.2 microgram/ml, respectively. The range of plasma FVIII/vWF concentrations varied between 8.3 microgram/ml and 24.9 microgram/ml for 10 normal adults. The plasma FVIII/vWF concentrations determined by the radioreceptor assay correlated well with levels measured by the ristocetin-induced platelet aggregation method, thus demonstrating the functional relevancy of the radioreceptor assay for plasma FVIII/vWF. 相似文献
To understand the hematopoietic and nonhematopoietic responses to interleukin-3 (IL-3), expression of cell-surface IL-3 receptors (IL-3R) was examined on bone marrow (BM) cells and peripheral blood (PB) cells of rhesus monkeys during the course of in vivo IL-3 treatment. Whereas IL-3R expression is low in untreated monkeys, IL-3 administration led to a gradual increase in both low- and high-affinity binding sites for IL-3. This increase reflected the total number of cells expressing IL- 3Rs, as detected by flow cytometry using biotinylated IL-3. Most of these IL-3R+ cells in both BM and PB could be characterized as basophilic granulocytes that contained high levels of histamine. In contrast to the effect on these differentiated cells, IL-3 administration did not significantly alter the low level IL-3R expression on immature, CD34+ cells. Further flow cytometric analysis using biotinylated growth factors showed that the IL-3R+ basophils also expressed receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF), but not for IL-6 or Kit ligand. These findings indicated that the IL-3R+ cells included neither monocytes, which express GM-CSFRs and IL-6Rs abundantly, nor mast cells, which express c- kit. By combining flow cytometric and Scatchard data, it was calculated that the basophils contain as many as 1 to 2 x 10(3) high-affinity IL- 3Rs and 15 to 30 x 10(3) low-affinity sites. The finding that in vivo IL-3 treatment leads to the production of large numbers of cells that express high levels of IL-3R and are capable of producing histamine provides an explanation for the often severe allergic reactions that occur during prolonged IL-3 administration. It also indicates that IL- 3, in addition to its direct effects on hematopoietic cells, may also stimulate hematopoiesis through the release of secondary mediators such as histamine by IL-3-responsive mature cells. 相似文献
In hemophilia B, assays based on a monoclonal antifactor IX specific for the Thr-148 variant of an exonic polymorphism have diagnosed carriers in selected families by either establishing linkage or by indicating the presence or absence of a given normal factor IX. The sensitivity of the immunoassays for detecting heterozygous women was explored by comparing results from immunoassays with solid-phase polyclonal v the monoclonal antifactor IXs. Factor IX with the normal Ala-148 variant gave a flat dilution curve, qualitatively distinct from factor IX with the Thr-148 variant in the monoclonal assay. The two were indistinguishable in the polyclonal assay. Mixtures of equal amounts of the two types gave an intermediate result, about half as reactive in the monoclonal as compared with the polyclonal assay system. Whereas mixtures with 10% Ala-148 and 90% Thr-148 factor IXs could not readily be distinguished from Thr-148 factor IX plasma, as little as 1% of the Thr-148 protein was detected in Ala-148 factor IX plasma. The frequency of the Ala-148 variant varied in individuals with different ethnic backgrounds; it was found in 29% of white, 12% of black, and none of Asian blood donors' factor IX genes in Seattle. Only 4% of samples from South African black men were nonreactive (ie, Ala- 148). The Thr/Ala-148 dimorphism is in strong linkage disequilibrium with Taql restriction fragment length polymorphisms (RFLPs). Three recombinations were noted in normal white genes and one in a normal black factor IX gene (less than 2% of those examined). In 34 white families with at least one woman being a possible carrier, genetically, the immunoassay results were informative in 18. RFLP analyses were informative in eight of the 15 families tested. In five families each, assignment of carrier status was made to a woman by only DNA or only immunoassay results, whereas the other approach was noninformative. The immunoassays provide a rapid, inexpensive screening test and complement DNA analysis in white women who are potential carriers of hemophilia B. 相似文献