[Signal transduction through SH2, SH3 and PH domains].
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Amino Acid Sequence↗
Biomedical subjects
H Miki
Publications and source records attributed to H Miki.
[A case of stage IVb gastric cancer effectively treated by neoadjuvant chemotherapy and total gastrectomy with no residual tumor (R0)].
PA 72-year-old man visited our hospital complaining of body-weight loss and appetite loss. The upper gastrointestinal series and gastrointestinal fibroscopy disclosed a Borrmann 3 type gastric cancer. Laparoscopic examination revealed peritoneal dissemination (P2). Continuous intravenous administration of 5-FU (500 mg/day) for three weeks and drip infusion of CDDP (10mg/day) five days per week every three weeks was given as neoadjuvant chemotherapy. This course was carried out twice. Then laparoscopic reexamination revealed complete disappearance of disseminated lesions of the peritoneum, and total gastrectomy with R0 was done. Histopathological finding of the resected specimen showed significant effects of the neoadjuvant chemotherapy. The patient has been doing well with keeping good performance status (PS) 0 for one year and seven months after first diagnosis.
Adenocarcinoma↗
Estimation of free calcium levels after thyroidectomy.
Total calcium is routinely measured after thyroidectomy in a clinical setting, while the measurement or calculation of the free calcium level is not generally performed. We reviewed total and free calcium levels in patients who underwent lobectomy (n = 15), subtotal thyroidectomy (n = 15) and total thyroidectomy (n = 15). Postoperative total calcium levels decreased significantly in comparison to preoperative levels in all thyroidectomies (p < 0.01), and this fall was significantly related to the extent of surgery (p < 0.01). In contrast, there was no significant difference between preoperative and postoperative free calcium levels in patients undergoing lobectomy, although we found a decrease in free calcium levels after both subtotal and total thyroidectomy. Total protein levels decreased regardless of the type of operation. Serum total calcium levels were thought to be altered by serum protein levels through the change of protein-bound calcium levels. When examined for free calcium levels, some patients were administered unnecessary calcium supplementation because hypocalcemia had been judged from the total calcium level. Since the wrong diagnosis may be given with regard to hypoparathyroidism by measurement of total calcium levels alone, we propose that free calcium levels should be routinely measured or calculated after thyroidectomy.
Biomarkers↗
Beta-tubulin binds Src homology 2 domains through a region different from the tyrosine-phosphorylated protein-recognizing site.
Src homology 2 (SH2) domains have been demonstrated to bind tyrosine-phosphorylated proteins that participate in signaling by growth factors and oncogenes by recognizing amino acid sequences containing phosphotyrosine residue. We found that SH2 domains such as Ash/Grb2, the 85-kDa subunit of phosphatidylinositol 3-kinase, and phospholipase Cgamma1 also bind beta-tubulin through a different region that recognizes phosphotyrosine in vitro and in vivo. Furthermore, binding occurs even when the SH2 domain is occupied by tyrosine-phosphorylated epidermal growth factor receptors. Using deleted constructs of Ash/Grb2 SH2, we found that carboxyl-terminal beta strands E and F, and alpha helix B (region "c") are required for binding. A synthetic peptide (FLWVVKFNSLNELVDYH) composed of region c inhibited the binding of beta-tubulin to the SH2 domains of Ash/Grb2, phosphatidylinositol 3-kinase, and phospholipase Cgamma1. The co-localization of SH2 proteins and microtubules is also confirmed by immunostaining. These data suggest that microtubules play important roles in the assembly of signaling molecules complexes containing SH2 proteins.
Adaptor Proteins, Signal Transducing↗
N-WASP, a novel actin-depolymerizing protein, regulates the cortical cytoskeletal rearrangement in a PIP2-dependent manner downstream of tyrosine kinases.
Here we identify a 65 kDa protein (N-WASP) from brain that binds the SH3 domains of Ash/Grb2. The sequence is homologous to Wiskott-Aldrich syndrome protein (WASP). N-WASP has several functional motifs, such as a pleckstrin homology (PH) domain and cofilin-homologous region, through which N-WASP depolymerizes actin filaments. When overexpressed in COS 7 cells, the wild-type N-WASP causes several surface protrusions where N-WASP co-localizes with actin filaments. Epidermal growth factor (EGF) treatment induces the complex formation of EGF receptors and N-WASP, and produces microspikes. On the other hand, two mutants, C38W (a point mutation in the PH domain) and deltaVCA (deletion of the actin binding domain), localize predominantly in the nucleus and do not cause a change in the cytoskeleton, irrespective of EGF treatment. Interestingly, the C38W PH domain binds less effectively to phosphatidylinositol 4,5-bisphosphate (PIP2) than the wild-type PH domain. These results suggest the importance of the PIP2 binding ability of the PH domain and the actin binding for retention in membranes. Collectively, we conclude that N-WASP transmits signals from tyrosine kinases to cause a polarized rearrangement of cortical actin filaments dependent on PIP2.
Actins↗
Glucagon-like peptide-1(7-36)amide enhances insulin-stimulated glucose uptake and decreases intracellular cAMP content in isolated rat adipocytes.
We investigated the effect of GLPs on glucose uptake in isolated rat adipocytes. GLP-1(7-36)amide significantly enhanced glucose uptake in the presence of 1 nM insulin. GLP-1(7-36)amide at 15 nM increased glucose uptake maximally by 56.4% as compared with 1 nM insulin alone (P < 0.01). In contrast, with less than 1 nM insulin or without insulin GLP-1(7-36)amide showed no effect on glucose uptake. Full-sequence GLP-1(1-37) at 15 nM in the presence of 1 nM insulin increased glucose uptake by 24.6% as compared with 1 nM insulin alone (P < 0.05). GLP-2 showed no effect on glucose uptake. Further, we examined the effect of GLP-1(7-36)amide on cAMP content in isolated rat adipocytes. Insulin at 1 nM caused a significant decrease of cAMP content. The combination of 15 nM GLP-1(7-36)amide and 1 nM insulin caused a further reduction of cAMP content. These data indicate that GLP-1(7-36)amide possesses augmentative effects on insulin action in isolated rat adipocytes. Furthermore, it is suggested that the stimulatory effect of GLP-1(7-36)amide occurs through the reduction of intracellular cAMP content.
1-Methyl-3-isobutylxanthine↗
Interaction of Ash/Grb-2 via its SH3 domains with neuron-specific p150 and p65.
We found that 180 kDa, 150 kDa (p150), 110 kDa, 100 kDa and 65 kDa (p65) proteins comprise the major Ash/Grb-2-binding proteins in bovine brain. Among these proteins, 180 kDa and 100 kDa proteins have already been identified as Sos and dynamin respectively. Here, p150 and p65 were affinity-purified with glutathione S-transferase-Ash fusion protein and their partial amino acid sequences were determined. Analysis showed p150 and p65 to be new proteins. These two proteins bind to both the N-terminal SH3 domain and the C-terminal SH3 domain of Ash. It was found that p150 and p65 are expressed predominantly in brain, although Ash is widely distributed in all tissues examined by Western blots. Immunohistochemical staining of rat brain showed p150 and p65 to be localized in a variety of neurons in the cerebellum and hippocampus, with p65 being especially concentrated in the nerve terminal. When the Ash-binding-motif peptide of the epidermal growth factor receptor was used to detect complexes formed with Ash in vivo, 180 kDa, 150 kDa, 110 kDa, 100 kDa and 65 kDa proteins were also bound; this shows that these proteins form complexes with Ash in brain. In addition, p150 and p65 co-immunoprecipitated with Ash. All these results suggest that Ash may function as a regulator of synaptic vesicle transport through dynamin, p150 and p65.
Adaptor Proteins, Signal Transducing↗
In situ hybridization analysis of substance P receptor in the rat retina.
Substance P receptor is known to provide a principal interface between tachykinin peptides and tachykinin-sensitive cells in retinal circuitry and to produce several physiological functions such as excitation of ganglion cells. We reported results of in situ hybridization analysis of substance P receptor in rat retina using digoxigenin-labeled RNA probes to yield discrete cell labeling. Distinct hybridization signal was present in a great majority of ganglion cells that provide retinal fibers to a central target. It was also present in a subpopulation of amacrine cells. Following optic nerve crush, ganglion cells lost their hybridization signal in a time-dependent manner, while hybridization-positive amacrine cells were persistently seen. From the results, we identified the hybridization message as distinctly localized to two systems, output cells and intrinsic cells in retinal circuitry.
Animals↗
High intraocular pressure-induced ischemia and reperfusion injury in the optic nerve and retina in rats.
PURPOSE: The purpose of this paper is to describe the damage caused to the retina and the axons of the optic nerve by acute ischemia-reperfusion injury and the extent to which optic nerve damage correlates with the duration if ischemia due to high intraocular pressure (IOP). METHODS: Acute ischemia in the retina and optic disc was induced in albino rats by increasing the IOP to 110 mmHg for a period of 45-120 min. Thereafter, the eyes were reperfused at normal IOP after 7 days. The retina and optic nerve were examined by light and electron microscopy, and morphometrical counts of the optic nerve axons were performed. RESULTS: After 45 min of ischemia, electron microscopic examination revealed swelling of mitochondria and degeneration of neurotubules on axons in cross sections of the optic nerve. The axonal counts in eyes subjected to 45 min of ischemia were 29% lower than in control eyes. After 60 min of ischemia, there were distinct disruptions of mitochondria and degeneration of the axons. After 90 min of ischemia, numerous axons showed degeneration with disordered myelin sheaths. Neuronal cell death was seen in the retina, mainly in the ganglion cell layer. CONCLUSION: Damage to the retinal ganglion cell layer and the optic nerve was evident after only 45 min of ischemia in normal eyes. This experiment suggests that seriously injured eyes must be protected from high IOP; if IOP elevation is required during vitrectomy, it is essential to reduce the duration of interruption of blood flow to a minimum.
Animals↗
Parathyroid carcinoma in patients with chronic renal failure on maintenance hemodialysis.
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Adult↗
Signal transductions of SH2/SH3: Ash/Grb-2 downstream signaling.
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Adaptor Proteins, Signal Transducing↗
Case report: lymphangioma of the oesophagus endoscopically resected.
Lymphangioma of the oesophagus is an extremely rare entity, with only nine cases having been reported worldwide. We report on a 52-year-old woman with oesophageal lymphangioma, diagnosed using endoscopic ultrasonography and endoscopically resected. No case of malignant transformation of the lymphangioma has been reported in the literature. Endoscopic resection seems to be a minimally invasive method that is appropriate both for the removal of the tumour and precise diagnosis.
Endoscopy↗
Pigmentary degeneration induced by N-methyl-N-nitrosourea and the fate of pigment epithelial cells in the rat retina.
Pigmentary degeneration of the retina was induced by a single intraperitoneal injection of 75 mg/kg of N-methyl-N-nitrosourea (MNU) in female Brown-Norway colored rats at 50 days of age, which were then observed at 24, 48 and 72 h and 7, 21, 35 and 150 days after the treatment. MNU-treated rats showed selective destruction of the photoreceptor cells by an apoptotic mechanism 24 h after the treatment, and the destruction was completed by day 7. During the photoreceptor cell degeneration, proliferation of Müller cells and infiltration of macrophages was prominent 72 h and 21 days after the treatment, respectively. Müller cell proliferation and macrophage infiltration corresponded to degenerative photoreceptor cell phagocytosis, and proliferating Müller cell processes responded to stabilize the damaged retina. Pigment epithelial cell detachment from the Bruch's membrane was seen 72 h after the treatment, and migration within all layers of the retina was seen at day 7 when photoreceptor cells were lost. At 21, 35 and 150 days after the treatment, lack of photoreceptor cells and deposition of pigment epithelial cells within the retina but not in contact to vascular endothelial cells were characteristic. MNU-induced photoreceptor apoptosis followed by Müller cell and macrophage reaction then pigment epithelial cells deposition within the retina partially resembles retinitis pigmentosa in humans.
Animals↗
Malignant fibrous histiocytoma of the pancreas.
An extremely rare case of malignant fibrous histiocytoma in the pancreas is reported. A 70-year-old man complained of upper abdominal discomfort. A tumor in the head of the pancreas was demonstrated by ultrasonography and computed tomography. The surgical specimen revealed a relatively well demarcated tumor, 9 x 7 x 6.5 cm in size. Microscopically, fibroblastic, histiocytic, and multinucleated giant tumor cells were observed in the myxoid area, but some tumor cells had proliferated in a storiform-pleomorphic pattern. Immunohistochemically, some tumor cells were positive for lysozyme, alpha-1-antitrypsin, alpha-1-antichymotrypsin, and vimentin. Electron microscopically, tumor cells showed a combination of fibroblastic and histiocytic features. The patient is currently well with no evidence of recurrence or metastasis 22 months after operation.
Aged↗
In vitro and in vivo antifungal activities of ER-30346, a novel oral triazole with a broad antifungal spectrum.
ER-30346 is a novel oral triazole with a broad spectrum of potent activity against a wide range of fungi. ER-30346, with MICs at which 90% of the strains tested are inhibited (MIC90s) ranging from 0.025 to 0.78 microgram/ml, was 4 to 32 times more active than itraconazole, fluconazole, and amphotericin B against Candida albicans, Candida parapsilosis, and Candida glabrata. Against Candida tropicalis, ER-30346, with an MIC90 of 12.5 micrograms/ml, was 2 to > 8 times more active than itraconazole and fluconazole, but was 16 times less active than amphotericin B. ER-30346 (MIC90, 0.78 microgram/ml) was four to eight times more active than fluconazole and amphotericin B and had activity comparable to that of itraconazole against Trichosporon beigelli. The MIC90s of ER-30346 were 0.10 microgram/ml for Cryptococcus neoformans and 0.39 microgram/ml for Aspergillus fumigatus. ER-30346 was 2 to 8 times more active than itraconazole and amphotericin B and 32 to > 256 times more active than fluconazole. ER-30346 also showed good activity against dermatophytes, with MICs ranging from 0.05 to 0.39 microgram/ml, and its activity was comparable to or 2 to 16 times higher than those of itraconazole and amphotericin B and > 32 times higher than that of fluconazole. In vivo activity was evaluated with systemic infections in mice. Against systemic candidiasis and cryptococcosis, ER-30346 was comparable in efficacy to fluconazole and was more effective than itraconazole. Of the drugs tested, ER-30346 was the most effective drug against systemic aspergillosis. We studied the levels of ER-30346 in mouse plasma. The maximum concentration of drug in plasma and the area under the concentration-time curve for ER-30346 showed good linearity over a range of doses from 2 to 40 mg/kg of body weight.
Animals↗
Efficacy of ER-30346, a novel oral triazole antifungal agent, in experimental models of aspergillosis, candidiasis, and cryptococcosis.
ER-30346 is a novel oral triazole with a broad spectrum of potent activity against a wide range of fungi. In the present study, we investigated the therapeutic effects of oral ER-30346 on experimental local infections caused by Aspergillus fumigatus, Candida albicans, and Cryptococcus neoformans and compared them with those of itraconazole and fluconazole. In experimental murine models of pulmonary aspergillosis, candidiasis, and cryptococcosis, ER-30346 reduced the numbers of CFU in the lungs significantly compared with the numbers of CFU in the lungs of the controls (P < 0.05). ER-30346 was as effective as or more effective than itraconazole against pulmonary aspergillosis. Against pulmonary candidiasis and cryptococcosis, ER-30346 was more effective than itraconazole and was as effective as fluconazole. ER-30346 was also effective against pulmonary candidiasis caused by fluconazole-resistant C. albicans. In mice with intracranial cryptococcosis, ER-30346 reduced the numbers of CFU in the brains significantly compared with the numbers of CFU in the brains of the controls (P < 0.05) and was more effective than itraconazole and as effective as fluconazole. In an experimental model of oral candidiasis in rats, ER-30346 reduced the numbers of CFU in oral swabs significantly compared with the numbers of CFU in oral swabs from the controls (P < 0.05) and was more effective than itraconazole and as effective as fluconazole. Thus, ER-30346 shows efficacy in murine aspergillosis, candidiasis, and cryptococcosis models. Further studies are needed to determine the potential of ER-30346 for use in the treatment of these infections.
Animals↗
Intracranial aneurysms: diagnostic accuracy of MR angiography with evaluation of maximum intensity projection and source images.
PURPOSE: To determine whether evaluation of source images from magnetic resonance (MR) angiography in addition to maximum-intensity projection (MIP) images improves the detection of aneurysms. MATERIALS AND METHODS: Conventional and MR angiography were performed in 193 patients with various intracranial vascular lesions or normal findings. Images were evaluated in a blinded manner. Two readings were performed 6 weeks apart by evaluating MIP images with and without source images. Results were evaluated with receiver operating characteristic analysis. RESULTS: Sensitivity for the detection of aneurysms increased slightly when source images were included. The detection rate of internal carotid artery aneurysms was most improved with the addition of source images. No statistically significant differences in performance were found between the readings with MIP images alone and with source images. CONCLUSION: Sensitivity may improve with combined reading of nonselective MIP and source images.
Case-Control Studies↗
Effect of macroscopic deformation on lung microstructure.
Using an anisotropic theory of diffuse light scattering in lungs, we measured the fractional changes in geometric mean linear intercepts in orthogonal directions when freshly excised rabbit lungs were subjected to isovolume uniaxial strains. Results from the optical technique were compared with morphometric estimates of fractional changes in mean linear intercepts from the same strained and unstrained (control) lobes, with the conclusion that diffuse light scattering is adequate to estimate changes in mean free paths in different directions. We compared optical estimates of fractional changes in mean linear intercepts with the macroscopic strain field measured by displacements of pleural markers; this relationship did not significantly differ from the line of identity. We conclude that the microscopic strain field is closely matched to the macroscopic strain field during uniaxial distortion. This suggests that surface reorientation may not play a large role in the origin of the low shear modulus of the lung, but this cannot be definitively stated without comparison of these experimental results to specific model predictions of the changes in mean linear intercepts in shear deformation.
Animals↗