Using the whole-cell patch clamp technique, the role of actin microfilament in hyposmotic increase of voltage-operated calcium
channel current (IBa) was studied in guinea-pig gastric myocytes. Hyposmotic superfusate (212 mOsm) increased peak IBa amplitude by 32.7 ± 6.5%; when cytochalasin-D (Cyt-D, 20 μM), an actin cytoskeleton disruptor, was used, an increase of only
9.7 ± 3.1% was seen. IBaresponse to osmotic stress was potentiated (45.1 ± 4.1% increase) by 20 μM phalloidin, an actin microfilament stabilizer.
However, colchicine (100 μM), an microtubule cytoskeleton disruptor, had no effect on either IBa or its response to hyposmotic solution. Phalloidin also induced a rightward shift of the I/V relationship of IBa, while Cyt-D itself had no effect. These results suggest that actin cytoskeleton may mediate hyposmotic stretch-induced IBa increase in gastric smooth muscle.
Received: 26 March 1997 / Received after revision: 28 May 1997 / Accepted: 3 June 1997 相似文献
We have investigated the effects of intravitreal sciatic nerve (SN) and/or optic nerve (ON) grafts on the survival and the axonal regeneration of retinal ganglion cells (RGCs). Following transection of the ON, approximately 40% RGCs survived at 7 days post-axotomy (dpa). Results showed that the intravitreal ON graft significantly promoted the survival of RGCs at 7 dpa (39,063 vs 28,246). Intravitreal SN graft, however, did not rescue axotomized RGCs at 5, 7 or 14 dpa. Axotomized RGCs could be induced to regenerate axons along a segment of SN graft attached to the proximal stump of ON. On average, 608 axotomized RGCs were induced to regenerate axons along the attached SN graft. The presence of intravitreal SN graft promoted about 100% increase in the number of regenerating RGCs (1,227) relative to the control groups. The intravitreal ON graft, surprisingly, also induced about 100% more regenerating RGCs (1220) than in the control group. When SN and ON grafts were co-transplanted into the vitreous, about 200% more regenerating RGCs (1916) were observed than in the control group. These findings illustrated that the intravitreal ON graft rescued axotomized RGCs and enhanced the regeneration of retinal axons. This is the first report to show that ON promotes RGC axonal regeneration. The intravitreal SN graft did not rescue RGCs but promoted axonal regeneration. The differential effects of intravitreal ON and SN grafts on the survival and the RGC regeneration suggest that these might be two independently operating events. 相似文献
Trypanosoma brucei, the cause of African sleeping sickness, differentiates in the mammalian bloodstream from a long, slender trypanosome into a short, stumpy trypanosome. This event is necessary for infection of the tsetse fly and maintenance of the life cycle. We have previously shown that the stumpy form contains 10- to 15-fold-greater cysteine protease activity than either the slender form or the insect midgut procyclic, and we have isolated a cDNA encoding the protease. In order to determine whether the cDNA encodes the developmentally regulated cysteine protease, we have purified the protease from trypanosomes and have made a polyclonal antiserum against it. The trypanosomal protease gene was then expressed in Escherichia coli with three different methionines within the pre- and propeptides acting as initiation sites. In each case, a protein was synthesized that was recognized by an antiserum specific for the developmentally regulated trypanosomal cysteine protease. The protein synthesized from the more upstream initiation site within the propeptide was proteolytically active. The recombinant protease and the trypanosomal enzyme were identical with respect to peptide substrates and protease inhibitors. The protein remained active when synthesized in a truncated form lacking the nine consecutive prolines and carboxy-terminus extension, indicating that the terminal 108 amino acids are not necessary for proteolytic activity. 相似文献
X-linked severe combined immunodeficiency (X-SCID) is a rare, life-threatening immune disorder, caused by mutations in the gamma c chain gene, which encodes an essential component of the cytokine receptors for interleukin-2 (IL-2), IL-4, IL-7, IL-9, IL-15, and IL-21. A 13-month-old boy with recurrent infections who had reduced serum immunoglobulin levels and decreased numbers of CD3, CD16/56 cells was evaluated for gamma c chain gene mutation and protein expression. The patient had a C-to-T point mutation at nucleotide position 690, one of the hot spots, resulting in a single amino acid substitution of cysteine for arginine (R226C), as determined by direct sequencing and PCR-RFLP. The patient's mother was a heterozygous carrier. Percutaneous umbilical cord blood sampling was performed at the 6-month of gestation in a subsequent pregnancy. As the immunophenotype of the fetus showed an identical pattern, the pregnancy was terminated and genetic analysis of the abortus confirmed recurrence. This is the first report of the molecular diagnosis of X-SCID in Korea. Genetic analysis of the gamma c chain gene is useful for definite diagnosis and genetic counseling for X-SCID. 相似文献
In this review we focus on peptide- and peptidomimetic-based approaches that target autoimmune diseases and some pathologies of the central nervous system. Special attention is given to asthma, allergic rhinitis, osteoarthritis, and Alzheimer's disease, but other related pathologies are also reviewed, although to a lesser degree. Among others, drugs like Diacerhein and its active form Rhein, Pralnacasan, Anakinra (Kineret), Omalizumab, an antibody "BION-1", directed against the common beta-chain of cytokine receptors, are described below as well as attempts to target beta-amyloid peptide aggregation. Parts of the review are also dedicated to targeting of pathologic conditions in the brain and in other tissues with peptides as well as methods to deliver larger molecules through the "blood--brain barrier" by exploring receptor-mediated transport, or elsewhere in the body by using peptides as carriers through cellular membranes. In addition to highlighting current developments in the field, we also propose, for future drug targets, the components of the inflammasome protein complex, which is believed to initiate the activation of caspase- 1 dependent signaling events, as well as other pathways that signal inflammation. Thus we discuss the possibility of targeting inflammasome components for negative or positive modulation of an inflammatory response. 相似文献
The relationship between work rate (W˙) and time to exhaustion (t) during intense exercise is commonly described by either a hyperbolic function (NLin), t=W′/(W˙?W˙cp), or by its linear equivalent (LinW) Wlim=W′+W˙cp(t). The parameter W˙<INFcp (critical power) has been described as an inherent characteristic of the aerobic energy system, while W?′ has been shown to be a ralid estimate of anaerobic work capacity. Recent studies have demonstrated that oral supplementation of creatine monohydrate (CrH2O) increases total muscle creatine stores, and have linked these increases to improved performances in intense intermittent exercise. This study was conducted to determine the effect of CrH2O supplementation on estimates of W?′ and W˙<INFcp derived from the NLin and LinW equations, and to determine the effect of CrH2O on t in exhaustive constant power exercise of different intensities. Fifteen active but untrained university students completed three phases of testing on a cycle ergometer: (1) familiarization, three learning trials, (2) baseline determination of W?′ and W˙<INFcp, four bouts performed at a W˙ selected to elicit fatigue in 90–600?s, and (3) experimental determination of W?′ and W˙cp, four bouts performed at the same W˙ as baseline, but performed after 5 days of ingesting either a placebo (4?×?6?g of glucose/day) or CrH2O (4?×?5?g of CrH2O and 1?g glucose/day). Testing was administered in a double-blind manner. Analyses of covariance revealed a significant effect for CrH2O on both estimates of W?′ (NLin, P=0.04; LinW, P<0.01), but not on estimates of W˙cp (NLin, P=0.37; LinW; P=0.30). Within groups, t was significantly different for only CrH2O at the two highest W˙s (P=0.04). It is concluded that oral ingestion of CrH2O increases estimates of W?′ due to an improved t at the shorter, more intense exercise bouts.
The ion channel conductances that regulate the membrane potential was investigated by using a perforated patch-clamp technique in rabbit aorta endothelial cells (RAECs). The whole-cell current/voltage (I-V) relation showed a slight outward rectification under physiological ionic conditions. The resting membrane potential was -23.3 +/- 1.1 mV (mean +/- SEM, n = 19). The slope conductances at the potentials of -80 and 50 mV were 31.0 +/- 4.0 and 62.8 +/- 7.1 pS pF(-1), respectively (n = 15). Changes in the extracellular and intracellular Cl(-) concentrations did not affect the reversal potential on I-V curves. The background nonselective cationic (NSC) current was isolated after the K(+) current was suppressed. The relative permeabilities calculated from the changes in reversal potentials using the constant-field theory were P(K):P(Cs):P(Na):P(Li) = 1:0.87:0.40:0.27 and P(Cs):P(Ca) = 1:0.21. Increases in the external Ca(2+) decreased the background NSC current in a dose-dependent manner. The concentration for half block by Ca(2+) was 1.1 +/- 0.3 mM (n = 7). Through the continuous recording of the membrane potential in a current-clamp mode, it was found that the background NSC conductance is the major determinant of resting membrane potential. Taken together, it could be concluded that the background NSC channels function as the major determinant for the resting membrane potential and can be responsible for the background Ca(2+) entry pathway in freshly isolated RAECs. 相似文献
AIMS--To compare the haemoglobin (Hb) H inclusion test with immunocytochemical detection of embryonic zeta chains in screening for alpha thalassaemia. METHODS--Blood samples from 115 patients with relevant clinical history and hypochromic microcytic indexes were screened using the HbH inclusion test and the Variant Hemoglobin Testing System (BioRad, Hercules, CA, USA). RESULTS--The HbH inclusion test was positive in 61 of 115 cases, three of whom had HbH disease confirmed by electrophoresis. The remaining 58 had alpha thalassaemia 1. All three HbH cases and 56 of 58 cases of alpha thalassaemia 1 expressed embryonic zeta chains, giving a specificity of 96.7%. Fifty four of 115 cases had a negative HbH inclusion test, of whom 50 had beta thalassaemia trait and three had iron deficiency. No diagnosis was reached for the remaining patient. CONCLUSION--The immunocytochemical test is as sensitive as the HbH inclusion test in screening for alpha thalassaemia. The presence of zeta chains is highly specific for alpha thalassaemia 1 incorporating the (--/SEA) deletion. The specificity and simplicity of the immunocytochemical test make it the test of choice in screening for alpha thalassaemia. 相似文献
We have introduced concept of "chemical" embolization and have tried to develop a new agent which would enable us to embolize the lesion with one-shot injection. Such an agent must be able to occlude diffusely the lesion distal to the catheter. This has made it mandatory to develop a new catheter which can be introduced into the vessel as close to the lesion as possible with fewer risks of clot formation and/or vessel damage. A new superselective balloon catheter for angiography and infusion of liquid embolizing materials has been developed. This catheter consists of a proximal relatively stiff silicone catheter, a short distal thin-walled flexible silicone catheter and silicone balloon. These three silicone components are connected by silicone adhesives. The distal catheter allows us to catheterize fine arteries such as lenticulostriates, while the proximal catheter assures easy manipulation. This balloon catheter can be used for superselective angiography and infusion of liquid embolizing materials. It has been used on nine patients; one with a dural arterio-venous malformation (AVM), four with meningiomas, and four with brain and spinal cord AVMs. In the case of dural AVM and meningiomas, it was possible to easily introduce into the middle meningeal artery distal to the foramen spinosum. In addition, in one of the cases of meningioma, we were able to catheterize one of the main feeding pedicles beyond the pterion. Chemical embolization was carried out in 5 cases with good results. In the case of brain and spinal cord AVM, useful information was obtained from the superselective angiography.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献