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Biomedical subjects

Y Kimata

Publications and source records attributed to Y Kimata.

At least 19 recordsLinked to original sources
PubMed2001

Efficient radical trapping at the surface and inside the phospholipid membrane is responsible for highly potent antiperoxidative activity of the carotenoid astaxanthin.

The effects of the carotenoids beta-carotene and astaxanthin on the peroxidation of liposomes induced by ADP and Fe(2+) were examined. Both compounds inhibited production of lipid peroxides, astaxanthin being about 2-fold more effective than beta-carotene. The difference in the modes of destruction of the conjugated polyene chain between beta-carotene and astaxanthin suggested that the conjugated polyene moiety and terminal ring moieties of the more potent astaxanthin trapped radicals in the membrane and both at the membrane surface and in the membrane, respectively, whereas only the conjugated polyene chain of beta-carotene was responsible for radical trapping near the membrane surface and in the interior of the membrane. The efficient antioxidant activity of astaxanthin is suggested to be due to the unique structure of the terminal ring moiety.

Adenosine Diphosphate↗
PubMed2001

Translational control by the ER transmembrane kinase/ribonuclease IRE1 under ER stress.

Under conditions of endoplasmic reticulum (ER) stress, mammalian cells induce both translational repression and the unfolded protein response that transcriptionally activates genes encoding ER-resident molecular chaperones. To date, the only known pathway for translational repression in response to ER stress has been the phosphorylation of eIF-2alpha by the double-stranded RNA-activated protein kinase (PKR) or the transmembrane PKR-like ER kinase (PERK). Here we report another pathway in which the ER transmembrane kinase/ribonuclease IRE1beta induces translational repression through 28S ribosomal RNA cleavage in response to ER stress. The evidence suggests that both pathways are important for efficient translational repression during the ER stress response.

Amino Acid Sequence↗
PubMed2001

Diphtheria toxin receptor-mediated conditional and targeted cell ablation in transgenic mice.

Specific cell ablation is a useful method for analyzing the in vivo function of cells. We have developed a simple and sensitive method for conditional cell ablation in transgenic mice, called "toxin receptor-mediated cell knockout." We expressed the diphtheria toxin (DT) receptor in transgenic mice using a hepatocyte-specific promoter and found that injection of DT caused fulminant hepatitis. Three independently established transgenic lines demonstrated a good correlation between the sensitivity of hepatocytes to DT and the expression level of the DT receptors. Moreover, the degree of hepatocyte damage was easily controlled over a wide range of doses of injected DT without any obvious abnormalities in other cells or tissues. This system is useful for generating mouse models of disease and for studying the recovery or regeneration of tissues from cell damage or loss. As DT is a potent inhibitor of protein synthesis in both growing and non-growing cells, the method is applicable to a wide range of cells and tissues in mice or in other DT-insensitive animals.

Albumins↗
PubMed2001

Deep circumflex iliac perforator flap with iliac crest for mandibular reconstruction.

The deep circumflex iliac myocutaneous perforator (DCIP) flap with iliac crest was used to reconstruct oromandibular defects in 10 patients. In seven of the patients a dominant perforator was found preoperatively using a Doppler flowmeter; in five of these seven patients a DCIP flap was successfully transferred. In two of the seven patients the dominant perforators were too narrow: one patient underwent a standard osteocutaneous flap transfer and one patient underwent a second flap transfer. In three patients no dominant perforator was found before or during surgery. The freedom of the DCIP flap from the harvested iliac crest facilitates correct positioning. However, to ensure that the DCIP flap can be safely elevated, the presence of perforators must be confirmed preoperatively. Even when a perforator has been identified, complicated dissection may be necessary. We stress the importance of a thorough knowledge of the anatomy of second flaps and of obtaining informed consent to use them.

Adult↗
PubMed2000

Identification of a novel mammalian endoplasmic reticulum-resident KDEL protein using an EST database motif search.

Several endoplasmic reticulum (ER)-resident proteins contain a unique C-terminal sequence (KDEL) which is required for the retention of these proteins in the ER. By searching a mouse EST database for records containing the nucleotide sequence encoding the KDEL motif, we extracted cDNAs encoding putative novel ER-resident proteins in addition to all of the known ER proteins bearing the KDEL motif. Using the sequence information obtained by this database search, we cloned the cDNA encoding a novel KDEL motif-bearing protein, ER protein 58 (EP58), sharing no significant homology to any of the known ER-resident proteins. Subcellular localization of EP58 in the ER was confirmed by cytoimmunofluorescence studies using epitope-tagged EP58. The EP58 gene was primarily expressed in embryo, placenta, and adult heart. Neither heat shock nor ER stress as tested here was sufficient to induce expression of the EP58 gene. A putative role of the N-terminal half of EP58 in protein-protein interaction is suggested by its similarity to the filamin rod domain. Similarity of the EP58 sequence with bacterial and fungus proteins suggests a possible role for EP58 in polysaccharide biosynthesis.

Amino Acid Sequence↗
PubMed2000

Dissociation of Kar2p/BiP from an ER sensory molecule, Ire1p, triggers the unfolded protein response in yeast.

The unfolded protein response (UPR) is a signal transduction pathway induced by a variety of endoplasmic reticulum (ER) stresses and functions to maintain homeostasis of the cellular membrane in eukaryotes. Various ER stresses result in the accumulation of unfolded proteins in the ER, which is sensed by the transmembrane protein kinase/ribonuclease Ire1p that transmits a signal from the ER to the nucleus in Saccharomyces cerevisiae. Here we report that the yeast ER chaperone Kar2p/BiP, a member of the HSP70 family found in the ER, directly regulates the UPR by the interaction with Ire1p. In the absence of ER stress, Kar2p binds the lumenal domain of Ire1p and keeps Ire1p in an inactive unphosphorylated state. Upon exposure of cells to ER stresses, Kar2p is released from Ire1p, resulting in activation of Ire1p and signal transduction to the nucleus. Subsequently, KAR2 mRNA is induced and Kar2p accumulates in the ER in a time-dependent manner, restoring the system to the basal state. This negative autoregulation is similar to the regulation of mammalian cytosolic chaperone Hsp70 via its interaction with heat shock factor 1.

Cloning, Molecular↗
PubMed2000

The Saccharomyces cerevisiae RuvB-like protein, Tih2p, is required for cell cycle progression and RNA polymerase II-directed transcription.

Two highly conserved RuvB-like putative DNA helicases, p47/TIP49b and p50/TIP49a, have been identified in the eukaryotes. Here, we study the function of Saccharomyces cerevisiae TIH2, which corresponds to mammalian p47/TIP49b. Tih2p is required for vegetative cell growth and localizes in the nucleus. Immunoprecipitation analysis revealed that Tih2p tightly interacts with Tih1p, the counterpart of mammalian p50/TIP49a, which has been shown to interact with the TATA-binding protein and the RNA polymerase II holoenzyme complex. Furthermore, the mutational study of the Walker A motif, which is required for nucleotide binding and hydrolysis, showed that this motif plays indispensable roles in the function of Tih2p. When a temperature-sensitive tih2 mutant, tih2-160, was incubated at the nonpermissive temperature, cells were rapidly arrested in the G(1) phase. Northern blot analysis revealed that Tih2p is required for transcription of G(1) cyclin and of several ribosomal protein genes. The similarities between the mutant phenotypes of tih2-160 and those of taf145 mutants suggest a role for TIH2 in the regulation of RNA polymerase II-directed transcription.

Bacterial Proteins↗
PubMed2000

Sfb2p, a yeast protein related to Sec24p, can function as a constituent of COPII coats required for vesicle budding from the endoplasmic reticulum.

The COPII coat is required for vesicle budding from the endoplasmic reticulum (ER), and consists of two heterodimeric subcomplexes, Sec23p/Sec24p, Sec13p/Sec31p, and a small GTPase, Sar1p. We characterized a yeast mutant, anu1 (abnormal nuclear morphology) exhibiting proliferated ER as well as abnormal nuclear morphology at the restrictive temperature. Based on the finding that ANU1 is identical to SEC24, we confirmed a temperature-sensitive protein transport from the ER to the Golgi in anu1-1/sec24-20 cells. Overexpression of SFB2, a SEC24 homologue with 56% identity, partially suppressed not only the mutant phenotype of sec24-20 cells but also rescued the SEC24-disrupted cells. Moreover, the yeast two-hybrid assay revealed that Sfb2p, similarly to Sec24p, interacted with Sec23p. In SEC24-disrupted cells rescued by overexpression of SFB2, some cargo proteins were still retained in the ER, while most of the protein transport was restored. Together, these findings strongly suggest that Sfb2p functions as the component of COPII coats in place of Sec24p, and raise the possibility that each member of the SEC24 family of proteins participates directly and/or indirectly in cargo-recognition events with its own cargo specificity at forming ER-derived vesicles.

COP-Coated Vesicles↗
PubMed2000

Impaired proteasome function rescues thermosensitivity of yeast cells lacking the coatomer subunit epsilon-COP.

Formation of COPI-coated transport vesicles requires a cytosolic protein complex consisting of seven subunits: alpha-, beta-, beta'-, gamma-, delta-, epsilon- and zeta-COP, collectively designated coatomer. The yeast Saccharomyces cerevisiae gene encoding the epsilon-COP subunit is known as SEC28/ANU2. anu2 null mutant cells (anu2Delta) are temperature-sensitive, and alpha-COP is rapidly degraded in these cells when they are shifted to the restrictive temperature. We isolated extragenic suppressors that rescue the temperature-sensitive growth defect of anu2Delta cells. Genetic analysis revealed that one of the suppressors is allelic to PRE8 (PRS4), which encodes a 20 S proteasome subunit. In the presence of a proteasome inhibitor, MG132, anu2Delta cells did not cease growth even at the restrictive temperature. Furthermore, MG132 inhibited the rapid decrease of alpha-COP levels in anu2Delta cells shifted to the restrictive temperature. However, secretion of certain proteins by these cells was impaired even in the presence of MG132. In conclusion, impairment of proteasome-dependent proteolysis rescued some, but not all, temperature-sensitive defects of anu2Delta cells. These results are discussed in terms of evidence that epsilon-COP plays a critical role in maintaining the structural integrity of alpha-COP.

Adenosine Triphosphatases↗
PubMed2000

Anterolateral thigh flap donor-site complications and morbidity.

The authors examined donor-site complications and morbidity in 37 patients after reconstruction with free or pedicled anterolateral thigh flaps. Intraoperative assessment included damage to the vastus lateralis muscle and whether the main pedicle of the rectus femoris muscle had been killed. Postoperative assessment of the donor site included wound healing, range of motion, muscle strength, gait, and sensation. Patients were surveyed with a questionnaire about fatigue in their activities of daily life and the appearance of the donor site. All 32 patients who underwent primary skin closure could perform activities of daily life normally, and most (87.5 percent) reported that donor-site appearance was satisfactory. However, the severity of donor-site dysfunction was related to the degree of damage to the vastus lateralis muscle, and most patients (87.5 percent) had some loss of sensation at the anterolateral aspect of the thigh. Because of adhesions between the meshed skin graft and the underlying fascia, range of motion at the hip and knee was limited in significantly more patients who had received split-thickness skin grafts (60 percent) than patients who had undergone primary skin closure (3.1 percent). Therefore, wider flaps or flaps harvested nearer the knee may increase donor-site morbidity. The authors concluded that the incidence of long-term morbidity with the anterolateral thigh flap is low, although it is increased when the flap includes the vastus lateralis muscle or is wider and requires additional skin grafting at the donor site.

Adult↗
PubMed2000

Postoperative complications and functional results after total glossectomy with microvascular reconstruction.

Microsurgical reconstruction after total glossectomy can greatly improve quality of life; however, postoperative functional results are often unstable, and the effectiveness of total glossectomy remains questionable. To determine the problems of reconstruction after total glossectomy with laryngeal preservation and to examine the functional results of swallowing and speech, 30 patients who had undergone total glossectomy and reconstruction with free flaps were reviewed for this study. The patients ranged in age from 20 to 73 years, and 23 of the 30 had undergone reconstruction with a rectus abdominis musculocutaneous flap. Wider and thicker flaps were designed and transferred and were sutured to suspend the larynx. To maintain physiologic swallowing function after surgery, the extent of laryngeal suspension and cricopharyngeal myotomy was limited. Of the 30 patients, 21 (70 percent) could be decannulated with laryngeal preservation; 20 of these 21 could tolerate a normal/soft/pureed diet, and 1 was limited to a fluid diet. Speech was intelligible in 16 of the 19 patients evaluated. In 9 of the 30 patients, laryngeal function could not be preserved. In four of these nine patients, additional resection combined with total glossectomy caused severe aspiration and recurrent pneumonia. Two patients with preoperative cerebral dysfunction were also poor candidates for laryngeal preservation. Additionally, the transferred flap's lack of bulk in the oral cavity and the advanced age (73 years) of one patient and the poor motivation of another may have contributed to postoperative aspiration. Aspiration occurred in one patient because of local recurrence of a tumor. The presence of preoperative cerebral dysfunction (p = 0.025), resection of the epiglottis (p = 0.005), and postoperative orocutaneous fistulas (p = 0.04) were significantly associated with the failure of laryngeal preservation. However, because of the difficulty of enrolling a sufficient number of patients in the study and the inherent limitations of retrospective studies, multivariate analysis in this study showed that no factors, such as patient age, flap volume, and the type of neck dissection, were significant predictors of laryngeal preservation. Although prospective studies are necessary, the function of individual patients must be assessed so that the study experiences discussed here can be applied to subsequent patients.

Adult↗
PubMed2000

Elongation factor 2 in the liver and skeletal muscle of mice is decreased by starvation.

We examined whether starvation affected the amount of EF-2 protein as well as the level of its mRNA in the liver and skeletal muscle of mice, to understand the molecular mechanism for nutritional adaptation of protein-turnover. Although the amount of EF-2 was diminished by starvation in each of the tissues examined, the amount of EF-2 mRNA did not decrease in parallel with the protein.

Animals↗
PubMed1999

Free adipofascial flap for scalp reconstruction: case report.

The authors describe a case in which a large defect of the scalp was present after tumor excision. It was covered with a free adipofascial flap and a split-thickness skin graft, with satisfactory results. Numerous similar cases of scalp defects reconstructed with other free flaps have been described. But since donor-site morbidity is minimized with free adipofascial flaps, they should be used more often for reconstruction of scalp defects.

Adipose Tissue↗
PubMed1999

Anterolateral thigh flap for abdominal wall reconstruction.

The free or pedicled anterolateral thigh flap was introduced for the reconstruction of large abdominal wall defects. This flap is superior to the tensor fasciae latae musculocutaneous flap in several respects. These include the wide, reliable skin territory (which can reach the level of the knee) and the long pedicle. Therefore, a pedicled anterolateral thigh flap with reliable blood circulation can easily be positioned above the umbilicus. In addition, the free anterolateral thigh flap has greater freedom of orientation and can be used to repair larger abdominal wall defects than can the tensor fasciae latae flap. Seven patients in whom abdominal wall defects had been reconstructed with pedicled or free anterolateral thigh flaps were reviewed. Their average age was 47.1 years (range, 21 to 74 years), and the average follow-up period was 10.7 months (range, 2 to 21 months). The size of the abdominal wall defects ranged from 12 x 12 cm to 18 x 24 cm, and the size of the transferred flap ranged from 10 x 20 cm to 20 x 20 cm. Three flaps were pedicled and four were free, of which three incorporated the tensor fasciae latae flap. All flaps survived completely, and no postoperative abdominal hernias developed. Despite some variations in vascular anatomy and technical difficulties in elevating the anterolateral thigh flap, the authors conclude that the pedicled or free anterolateral thigh flap is superior to the tensor fasciae latae flap for reconstruction of large abdominal wall defects.

Abdominal Muscles↗
PubMed1999

Comparison of innervated and noninnervated free flaps in oral reconstruction.

Thirteen patients who had undergone ablative surgery for advanced squamous cell carcinoma in which more than half of the tongue had been resected underwent reconstruction in which the cutaneous nerve of a free flap was anastomosed to the stump of the transected lingual nerve. Eight of the patients underwent reconstruction with an innervated anterolateral thigh flap and five patients underwent reconstruction with an innervated rectus abdominis musculocutaneous flap. Sensory recovery of the flap at least 6 months postoperatively was compared in these 13 patients and in 16 additional patients who received noninnervated versions of the same flaps for the same defect. The degree of sensory recovery of innervated thigh flaps was significantly greater than that of noninnervated ones in all modalities and that of innervated rectus abdominis flaps was also greater than that of noninnervated flaps, except for hot and cold perception. These results indicate that sensory regrowth occurs in most areas through the surgically created pathways. However, results of Semmes-Weinstein testing showed that recovery did not reach the level of protective sensation in either type of innervated flap. Although these findings must be followed by additional objective and functional tests and the need for sensory reeducation should be considered, this simple operative procedure can improve postoperative intraoral function and should be attempted whenever possible after ablative surgery.

Abdominal Muscles↗
PubMed1999

Mutation of the yeast epsilon-COP gene ANU2 causes abnormal nuclear morphology and defects in intracellular vesicular transport.

Previously we reported an original method of visualizing the shape of yeast nuclei by the expression of green fluorescent protein (GFP)-tagged Xenopus nucleoplasmin in Saccharomyces cerevisiae. To identify components that determine nuclear structure, we searched for mutants exhibiting abnormal nuclear morphology from a collection of temperature-sensitive yeast strains expressing GFP-tagged nucleoplasmin. Four anu mutant strains (anu1-1, 2-1, 3-1 and 4-1; ANU=abnormal nuclear morphology) that exhibited strikingly different nuclear morphologies at the restrictive temperature as compared to the wild-type were isolated. The nuclei of these mutants were irregularly shaped and often consisted of multiple lobes. ANU1, 3 and 4 were found to encode known factors Sec24p, Sec13p and Sec18p, respectively, all of which are involved in the formation or fusion of intracellular membrane vesicles of protein transport between the endoplasmic reticulum (ER) and the Golgi apparatus. On the other hand, ANU2 was not well characterized. Disruption of ANU2 (delta anu2) was not lethal but conferred temperature-sensitivity for growth. Electron microscopic analysis of anu2-1 cells revealed not only the abnormal nuclear morphology but also excessive accumulation of ER membranes. In addition, both anu2-1 and delta anu2 cells were defective in protein transport between the ER and the Golgi, suggesting that Anu2p has an important role in vesicular transport in the early secretory pathway. Here we show that ANU2 encodes a 34 kDa polypeptide, which shares a 20% sequence identity with the mammalian epsilon-COP. Our results suggest that Anu2p is the yeast homologue of mammalian epsilon-COP and the abrupt accumulation of the ER membrane caused by a blockage of the early protein transport pathway leads to alteration of nuclear morphology of the budding yeast cells.

Amino Acid Sequence↗
PubMed1998

Loss of Hsp70-Hsp40 chaperone activity causes abnormal nuclear distribution and aberrant microtubule formation in M-phase of Saccharomyces cerevisiae.

The 70-kDa heat shock proteins, hsp70, are highly conserved among both prokaryotes and eukaryotes, and function as chaperones in diverse cellular processes. To elucidate the function of the yeast cytosolic hsp70 Ssa1p in vivo, we characterized a Saccharomyces cerevisiae ssa1 temperature-sensitive mutant (ssa1-134). After shifting to the restrictive temperature (37 degreesC), ssa1-134 mutant cells showed abnormal distribution of nuclei and accumulated as large-budded cells with a 2 N DNA content. We observed more prominent mutant phenotypes using nocodazole-synchronized cells: when cells were incubated at the restrictive temperature following nocodazole treatment, viability was rapidly lost and abnormal arrays of bent microtubules were formed. Chemical cross-linking and immunoprecipitation analyses revealed that the interaction of mutant Ssa1p with Ydj1p (cytosolic DnaJ homologue in yeast) was much weaker compared with wild-type Ssa1p. These results suggest that Ssa1p and Ydj1p chaperone activities play important roles in the regulation of microtubule formation in M phase. In support of this idea, a ydj1 null mutant at the restrictive temperature was found to exhibit more prominent phenotypes than ssa1-134. Furthermore, both ssa1-134 and ydj1 null mutant cells exhibited greater sensitivity to anti-microtubule drugs. Finally, the observation that SSA1 and YDJ1 interact genetically with a gamma-tubulin, TUB4, supports the idea that they play a role in the regulation of microtubule formation.

Adenosine Triphosphatases↗