RESUMO
OBJECTIVE: Hepatic artery (HA) reconstruction is an important part of resective surgery for advanced hepatobiliary and pancreatic malignancies, but few reports have been published. To identify indications for HA reconstruction, we retrospectively analyzed our surgical procedures and outcomes. METHODS: En-bloc resection of advanced hepatobiliary and pancreatic malignancies followed by HA reconstruction was performed in 35 patients. Patients ranged in age from 27 to 81 years and included 18 men and 17 women. The primary site of cancer included the bile duct in 22 patients, the pancreas in 7, and others in 6. Reconstruction of the HA was necessitated by HA resection due to direct cancer invasion in 29 patients and by accidental arterial injury during surgical procedure in 6 patients. RESULTS: The HA was reconstructed with end-to-end anastomosis between hepatic arteries in 17 patients. Transposition of an intra-abdominal artery, such as the gastroepiploic artery, was required in 14 patients, and arterial grafting was required in 4 patients. Although the HA patency was achieved in 30 patients, 4 cases of arterial thrombosis and 1 case of arterial rupture developed postoperatively. The overall RFS time was analyzed in all patients, and mean and median RFS times were 18 and 9 months, respectively. CONCLUSION: Although oncologic outcomes remain poor, HA resection and reconstruction can be performed in selected patients. We believe that the method of first choice for HA reconstruction is end-to-end anastomosis between HAs. A vascular autograft should be used only in selected cases.
Assuntos
Neoplasias do Sistema Biliar/cirurgia , Procedimentos Cirúrgicos do Sistema Biliar/efeitos adversos , Hepatectomia/efeitos adversos , Artéria Hepática/lesões , Artéria Hepática/cirurgia , Neoplasias Hepáticas/cirurgia , Pancreatectomia/efeitos adversos , Neoplasias Pancreáticas/cirurgia , Procedimentos Cirúrgicos Vasculares/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Anastomose Cirúrgica , Intervalo Livre de Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Procedimentos de Cirurgia Plástica/métodos , Resultado do TratamentoRESUMO
AIMS: Immediate maxillary reconstruction after malignant tumor extirpation differs from other types of maxillary reconstruction. Our reconstruction algorithm is described in this article. METHODS: One hundred ninety-four patients who had undergone maxillectomy for malignant tumors were reviewed, and maxillectomy defects were classified with the method of Cordeiro and Santamaria. RESULTS: Mean total blood loss was 848 ml, and 71 patients died within 2 years after surgery. For type IIIa defects of the orbital floor, titanium mesh or vascularized bone or cartilage was used for reconstruction, but the rate of postoperative complications did not differ between titanium and autografts. Therefore, to reconstruct orbital floor defects we have recently used only titanium mesh. For type I or II defects, we use autografts for only selected cases. CONCLUSIONS: We strive to perform less-invasive reconstructive surgery after resection for maxillary malignancy.
Assuntos
Neoplasias Maxilares/cirurgia , Procedimentos de Cirurgia Plástica/métodos , Idoso , Algoritmos , Feminino , Humanos , Masculino , Complicações Pós-Operatórias/cirurgia , Retalhos Cirúrgicos , Resultado do TratamentoRESUMO
A major response of eukaryotic cells to the presence of unfolded proteins in the lumen of the endoplasmic reticulum (ER) is to activate genes that encode ER-located molecular chaperones, such as the binding protein. This response, called the unfolded protein response, requires the transduction of a signal from the ER to the nucleus. In yeast (Saccharomyces cerevisiae) and mammalian cells, an ER-located transmembrane receptor protein kinase/ribonuclease called Ire1, with a sensor domain in the lumen of the ER, is the first component of this pathway. Here, we report the cloning and derived amino acid sequences of AtIre1-1 and AtIre1-2, two Arabidopsis homologs of Ire1. The two proteins are located in the perinuclear ER (based on heterologous expression of fusions with green fluorescent protein). The expression patterns of the two genes (using beta-glucuronidase fusions) are nearly nonoverlapping. We also demonstrate functional complementation of the sensor domains of the two proteins in yeast and show that the Ire1-2 protein is capable of autotransphosphorylation. These and other findings are discussed in relation to the involvement of these genes in unfolded protein response signaling in plants.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas Fúngicas/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases , Proteínas de Saccharomyces cerevisiae , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Transporte/metabolismo , Núcleo Celular/metabolismo , Núcleo Celular/ultraestrutura , Clonagem Molecular , Proteínas Fúngicas/genética , Regulação da Expressão Gênica de Plantas , Imuno-Histoquímica , Glicoproteínas de Membrana/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Chaperonas Moleculares/metabolismo , Dados de Sequência Molecular , Plantas Geneticamente Modificadas , Proteínas Quinases/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Transdução de Sinais , Tunicamicina/farmacologiaRESUMO
For this article, 178 consecutive cases of mandibular reconstruction using microvascular free flaps and performed from 1979 to 1997 were studied. The purpose of this report is to compare flap success rates, complications, and aesthetic and functional results. The ages of the 131 men and 47 women ranged from 13 to 85 years, with an average of 55 years. Donor sites included the rib (11 cases), radius (one case), ilium (36 cases), scapula (51 cases), fibula (34 cases), and soft-tissue flaps with implant (45 cases). Complications included total flap necrosis, partial flap necrosis, major fistula formation, and minor fistula formation. The rate of total flap necrosis involving the ilium and fibula was significantly higher than that of all other materials combined (p < 0.05). The overall rate of implant plate removal, which resulted from the exposure or fracture of the plate, was 35.6 percent (16 of 45 cases). Each mandibular defect was classified by the extent of the bony defect and by the extent of the soft-tissue defect. The extent of the mandibular bony defect was classified according to the HCL method of Jewer et al. The extent of the soft-tissue defect was classified into four groups: none, skin, mucosal, and through-and-through. According to these classifications, functional and aesthetic assessments of deglutition and contour were performed on 115 subjects, and speech was evaluated in 110. To evaluate the postoperative results, points were assigned to each assessment of deglutition, speech, and mandibular contour. Statistical analysis between pairs of bone-defect groups revealed that there was no significant difference in each category. Regarding deglutition, statistical analysis between pairs of soft-tissue-defect groups revealed there were significant differences (p < 0.05) between the none and the mucosal groups and also between the none and the through-and-through groups. Regarding speech, there was a significant difference (p < 0.05) between the none and the through-and-through groups. Regarding contour, there were significant differences (p < 0.01) between the none and the through-and-through groups and between the mucosal and the through-and-through groups. The points given for each function, depending on the reconstruction material, revealed that there was no significant difference between pairs of material groups. From this prospective study, the authors have developed an algorithm for oromandibular reconstruction. When the bony defect is lateral, the ilium, fibula, or scapula should be chosen as the donor site, depending on the extent of the soft-tissue defect. When the bony defect is anterior, the fibula is always the best choice. When the soft-tissue defect is extensive or through-and-through with an anterior bony defect, the fibula should be used with other soft-tissue flaps.
Assuntos
Mandíbula/cirurgia , Procedimentos de Cirurgia Plástica , Retalhos Cirúrgicos/irrigação sanguínea , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Algoritmos , Transplante Ósseo , Deglutição , Estética , Feminino , Humanos , Masculino , Neoplasias Mandibulares/cirurgia , Microcirurgia , Pessoa de Meia-Idade , Complicações Pós-Operatórias , Estudos Prospectivos , Procedimentos de Cirurgia Plástica/métodos , Inteligibilidade da FalaRESUMO
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.
Assuntos
Camundongos Transgênicos , Receptores de Superfície Celular/metabolismo , Albuminas/genética , Animais , Northern Blotting , Relação Dose-Resposta a Droga , Elementos Facilitadores Genéticos , Fator de Crescimento Semelhante a EGF de Ligação à Heparina , Hepatócitos/metabolismo , Humanos , Hibridização In Situ , Peptídeos e Proteínas de Sinalização Intercelular , Fígado/citologia , Fígado/metabolismo , Camundongos , Camundongos Knockout , Modelos Biológicos , Plasmídeos/metabolismo , Regiões Promotoras Genéticas , Regeneração , Fatores de Tempo , Distribuição Tecidual , Transaminases/sangue , TransfecçãoRESUMO
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.
Assuntos
Carcinoma de Células Escamosas/cirurgia , Mandíbula/cirurgia , Neoplasias Mandibulares/cirurgia , Retalhos Cirúrgicos/irrigação sanguínea , Adulto , Idoso , Feminino , Humanos , Fluxometria por Laser-Doppler , Masculino , Pessoa de Meia-Idade , Sobrevivência de Tecidos , Resultado do TratamentoRESUMO
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.
Assuntos
Difosfato de Adenosina/química , Antioxidantes/química , Cardiolipinas/química , Ferro/química , Peroxidação de Lipídeos , Lipossomos/química , Fosfatidilcolinas/química , beta Caroteno/análogos & derivados , beta Caroteno/química , Compostos Ferrosos/química , Radicais Livres/química , Ligação de Hidrogênio , Cinética , Modelos Moleculares , Conformação Molecular , Oxirredução , Propriedades de Superfície , Xantofilas , ZeaxantinasRESUMO
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.
Assuntos
Retículo Endoplasmático/metabolismo , Proteínas de Membrana , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Sequência de Aminoácidos , Animais , Antibacterianos/farmacologia , Apoptose/fisiologia , Northern Blotting , Western Blotting , Células Cultivadas , Doxiciclina/farmacologia , Endorribonucleases , Humanos , Imuno-Histoquímica , Dados de Sequência Molecular , Fosforilação , Biossíntese de Proteínas , Proteínas Serina-Treonina Quinases/química , Estrutura Terciária de Proteína , RNA Ribossômico 28S/metabolismo , Alinhamento de Sequência , Transfecção , Tunicamicina/farmacologia , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismoRESUMO
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.
Assuntos
Retículo Endoplasmático/fisiologia , Proteínas Fúngicas/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Glicoproteínas de Membrana/metabolismo , Chaperonas Moleculares/metabolismo , Dobramento de Proteína , Proteínas Serina-Treonina Quinases , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/fisiologia , Clonagem Molecular , Proteínas Fúngicas/genética , Proteínas de Choque Térmico HSP70/genética , Glicoproteínas de Membrana/genética , Modelos Biológicos , Fosforilação , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Transdução de Sinais/fisiologiaRESUMO
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.
Assuntos
Glossectomia/reabilitação , Complicações Pós-Operatórias , Retalhos Cirúrgicos , Adulto , Idoso , Ingestão de Alimentos , Feminino , Humanos , Laringe/cirurgia , Masculino , Microcirurgia/efeitos adversos , Pessoa de Meia-Idade , Boca/cirurgia , Esvaziamento Cervical , Estudos Retrospectivos , Fala , Retalhos Cirúrgicos/efeitos adversos , Neoplasias da Língua/cirurgiaRESUMO
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.
Assuntos
Complicações Pós-Operatórias , Retalhos Cirúrgicos , Coxa da Perna/cirurgia , Adulto , Feminino , Marcha , Humanos , Masculino , Transtornos de Sensação/etiologia , Pele/patologia , Transplante de Pele , Inquéritos e Questionários , CicatrizaçãoRESUMO
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.
Assuntos
Proteínas de Transporte/genética , Ciclo Celular/genética , DNA Helicases , Proteínas Fúngicas/genética , RNA Polimerase II/genética , Saccharomyces cerevisiae/genética , Proteínas de Bactérias/genética , Proteínas de Transporte/biossíntese , Proteínas Fúngicas/biossíntese , Regulação Fúngica da Expressão Gênica , Saccharomyces cerevisiae/citologia , Transcrição GênicaRESUMO
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.
Assuntos
Retículo Endoplasmático/fisiologia , Proteínas Fúngicas/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/fisiologia , Vesículas Revestidas pelo Complexo de Proteína do Envoltório , Dimerização , Retículo Endoplasmático/ultraestrutura , Proteínas Ativadoras de GTPase , Regulação Fúngica da Expressão Gênica , Proteínas de Membrana/genética , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Transporte VesicularRESUMO
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.
Assuntos
Proteína Coatomer/genética , Saccharomyces cerevisiae/genética , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Clonagem Molecular , Proteína Coatomer/química , Proteína Coatomer/metabolismo , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/farmacologia , Genótipo , Leupeptinas/farmacologia , Substâncias Macromoleculares , Complexos Multienzimáticos/genética , Complexos Multienzimáticos/metabolismo , Complexo de Endopeptidases do Proteassoma , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/fisiologia , Supressão Genética , Temperatura , beta-Galactosidase/metabolismoRESUMO
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.
Assuntos
Retículo Endoplasmático/metabolismo , Etiquetas de Sequências Expressas , Receptores de Peptídeos/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Western Blotting , Células COS , Linhagem Celular , DNA Complementar/química , DNA Complementar/genética , DNA Recombinante , Bases de Dados Factuais , Embrião de Mamíferos/metabolismo , Feminino , Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Proteínas de Membrana/genética , Camundongos , Microscopia Confocal , Dados de Sequência Molecular , Oligopeptídeos , Peptídeos/genética , Peptídeos/metabolismo , Plasmídeos/genética , RNA/genética , RNA/metabolismo , Receptores de Peptídeos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Software , Distribuição TecidualRESUMO
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.
Assuntos
Fígado/metabolismo , Músculo Esquelético/metabolismo , Fator 2 de Elongação de Peptídeos/metabolismo , Inanição/metabolismo , Animais , Camundongos , Fator 2 de Elongação de Peptídeos/genética , RNA Mensageiro/metabolismoRESUMO
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.
Assuntos
Boca/cirurgia , Procedimentos de Cirurgia Plástica/métodos , Retalhos Cirúrgicos/inervação , Músculos Abdominais , Adulto , Idoso , Carcinoma de Células Escamosas/cirurgia , Glossectomia , Humanos , Pessoa de Meia-Idade , Neoplasias Bucais/cirurgia , Sensação , Coxa da PernaRESUMO
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.
Assuntos
Complexo I de Proteína do Envoltório/genética , Proteína Coatomer/genética , Proteínas Fúngicas/genética , Hexosiltransferases , Proteínas de Membrana , Sequência de Aminoácidos , Transporte Biológico , Carboxipeptidases/metabolismo , Catepsina A , Núcleo Celular/metabolismo , Clonagem Molecular , Galactose/metabolismo , Glucose/metabolismo , Proteínas de Fluorescência Verde , Immunoblotting , Proteínas Luminescentes/metabolismo , Microscopia Eletrônica , Modelos Genéticos , Dados de Sequência Molecular , Mutagênese , Plasmídeos/metabolismo , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Temperatura , Fatores de Tempo , Transferases/metabolismoRESUMO
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.