RESUMO
Transgenic animals provide attractive systems for the production of valuable recombinant proteins. Previous studies indicate that milk is a suitable source of secreted recombinant proteins. In the current study, we examine whether excrement can be another source of recombinant proteins by using transgenic mice ubiquitously-expressing green fluorescent protein (GFP) as a model. We found that the surface of excrement from GFP-transgenic mice was fluorescent, and the supernatant after centrifugation of an excrement suspension was rich in undegraded, actively fluorescing GFP. GFP was successfully purified from stool as a fluorescent 27â¯kDa protein by using a common procedure. Finally, we observed that the fluorescence of digested materials began in the ileum and persisted throughout the remainder of the digestive tract. Our results demonstrate that excrement, which is produced daily regardless of the sex or age of the animal, may be another feasible source of recombinant proteins. The preparation method is simple, economical, and noninvasive.
Assuntos
Fezes/química , Proteínas de Fluorescência Verde/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Animais , Mucosa Gástrica/metabolismo , Proteínas de Fluorescência Verde/isolamento & purificação , Jejuno/metabolismo , Camundongos Endogâmicos ICR , Camundongos TransgênicosRESUMO
AIMS: We recently reported that a small subset (7%) of oesophageal squamous cell carcinomas completely lacking SOX2 expression had unique clinicopathological features and a dismal prognosis. The aim of the present study was to elucidate whether the findings obtained in oesophageal cancers are applicable to hypopharyngeal squamous cell carcinomas (HPSCCs) or oropharyngeal squamous cell carcinomas (OPSCCs). METHODS AND RESULTS: The study cohort consisted of consecutive patients with HPSCC (n = 130) and OPSCC (n = 65) who underwent surgery without preoperative therapy. On immunostaining, SOX2 was almost entirely negative in 10 of 130 HPSCCs (8%) and seven of 65 OPSCCs (11%). No significant differences were observed in clinicopathological features, including p16 status, between SOX2-positive and SOX2-negative cancers. However, patients with SOX2-negative HPSCC had significantly worse overall and recurrence-free survival than those with SOX2-positive HPSCC, whereas such a prognostic relationship was not confirmed in patients with OPSCC. In a multivariate analysis, the loss of SOX2 expression appeared to be an independent poor prognostic factor for patients with HPSCC. In a sequencing analysis, no mutation was found in SOX2. As SOX2 is known to contain an extensive CpG island before the transcription start site, methylation-specific polymerase chain reaction for the SOX2 promoter was performed. Methylated alleles were found in nine of 10 SOX2-negative HPSCCs but in none of SOX2-positive HPSCCs. CONCLUSIONS: Similarly to oesophageal cancers, a small subset (8%) of HPSCCs characteristically almost completely lacking SOX2 expression appeared to be aggressive neoplasms with high recurrence rates. Promoter hypermethylation was determined to be a major mechanism underlying epigenetic SOX2 silencing.
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Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias Hipofaríngeas/genética , Neoplasias Hipofaríngeas/patologia , Fatores de Transcrição SOXB1/genética , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/mortalidade , Metilação de DNA/genética , Intervalo Livre de Doença , Regulação para Baixo , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/mortalidade , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago , Feminino , Inativação Gênica , Neoplasias de Cabeça e Pescoço/mortalidade , Humanos , Neoplasias Hipofaríngeas/mortalidade , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Neoplasias Orofaríngeas/genética , Neoplasias Orofaríngeas/mortalidade , Neoplasias Orofaríngeas/patologia , Prognóstico , Regiões Promotoras Genéticas/genética , Modelos de Riscos Proporcionais , Carcinoma de Células Escamosas de Cabeça e PescoçoRESUMO
BACKGROUND: Concomitant chemoradiotherapy (CCRT) produces severe mucositis and swallowing dysfunction, often resulting in malnutrition. Intensive nutrition support (INS) in addition to percutaneous endoscopic gastrostomy (PEG) is reported to decrease adverse effects during CCRT. PATIENTS AND METHODS: Fifty-eight patients with oropharyngeal cancer treated by CDDP-based CCRT were retrospectively analyzed. Twenty-nine patients treated with INS in addition to PEG were classified as INS group, and other 29 patients treated with PEG but without INS were classified as control group. RESULTS: INS in addition to PEG significantly increased calorie intake in the second half of CCRT and reduced adverse events including mucositis (p = 0.0019), leukopenia (p = 0.04), and renal function (p = 0.006). Moreover, 21 out of 29 patients had successfully administration of 200 mg/m2 or more of CDDP, while only 10 out of 29 patients had enough amount of CDDP in control group. CONCLUSIONS: These results suggest that INS in addition to prophylactic PEG not only decreases adverse effects but also may potentially improve oncological outcome of the patients with oropharyngeal cancer treated by CCRT.
Assuntos
Quimiorradioterapia , Nutrição Enteral , Gastrostomia , Desnutrição/prevenção & controle , Neoplasias Orofaríngeas/terapia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Apoio Nutricional , Prognóstico , Estudos RetrospectivosRESUMO
OBJECTIVE: To study the oncological outcome of the patients with unresectable locally advanced primary head and neck cancers invading the skull base, treated with particle radiotherapy. METHODS: Fifty-seven patients with unresectable primary head and neck cancers invading the skull base received proton or carbon ion radiotherapy as definitive treatment at Hyogo Ion Beam Medical Center between 2003 and 2009. Forty-seven patients were treated with proton radiotherapy and 10 patients were treated with carbon ion radiotherapy. A retrospective review was performed with clinical charts and recorded imagings. RESULTS: With a median follow-up of 32 months, the 3-year actual survival and local progression-free rates of all the patients were 61 and 56%, respectively. The 3-year actual survival rates of adenoid cystic carcinoma, squamous cell carcinoma, olfactory neuroblastoma, adenocarcinoma and malignant melanoma were 83, 44, 75, 0 and 38%, respectively. The 3-year actual local control rates of adenoid cystic carcinoma, squamous cell carcinoma, olfactory neuroblastoma, adenocarcinoma and malignant melanoma were 63, 31, 83, 50 and 0%, respectively. Distant metastasis was observed in 13 of 25 patients in adenoid cystic carcinoma, two of 14 patients in squamous cell carcinoma, one of six patients with olfactory neuroblastoma, two of four patients with adenocarcinoma, three of four patients with malignant melanoma and two of three patients with undifferentiated carcinoma. Mucositis and dermatitis were seen as acute toxicities. The most common late toxicity was visual disorder. Grades 2, 3 and 4 visual disorders were observed in seven, five and two patients, respectively. CONCLUSIONS: Proton and carbon ion radiotherapy resulted in satisfactory local control in patients with locally advanced unresectable primary head and neck cancers invading the skull base.
Assuntos
Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/radioterapia , Radioterapia com Íons Pesados , Terapia com Prótons , Base do Crânio/patologia , Adenocarcinoma/mortalidade , Adenocarcinoma/radioterapia , Adulto , Idoso , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/radioterapia , Cistadenocarcinoma/mortalidade , Cistadenocarcinoma/radioterapia , Intervalo Livre de Doença , Estesioneuroblastoma Olfatório/mortalidade , Estesioneuroblastoma Olfatório/radioterapia , Feminino , Seguimentos , Neoplasias de Cabeça e Pescoço/mortalidade , Radioterapia com Íons Pesados/efeitos adversos , Humanos , Masculino , Melanoma/mortalidade , Melanoma/radioterapia , Pessoa de Meia-Idade , Mucosite/etiologia , Invasividade Neoplásica , Terapia com Prótons/efeitos adversos , Radiodermite/etiologia , Estudos Retrospectivos , Resultado do Tratamento , Transtornos da Visão/etiologiaRESUMO
Coagulation is an important process in the context of water purification; and the seed protein of the moringa tree (Moringa oleifera) is a remarkably effective coagulant. The laboratory course described here is designed to provide high-school students with a stepwise, hands-on experience in investigating the protein-rich coagulant found in Moringa seeds. First, the seed powder was applied to model polluted water containing fine clay, food dyes, copper sulfate, and bacteria. This treatment changed the polluted water into clear water via coagulation; all students were convinced that the coagulation-inducing agent was a thermostable cationic protein. Finally, basic biochemical techniques (e.g., chromatographic separation and electrophoresis) were used to show that the target coagulant is a dimeric protein composed of 6.5 and 4.5 kDa subunits. Overall, this made it possible for the students to gain a deeper understanding (more comprehensive than the information taught in formal classes) of protein structure and its real-world implications. This stepwise exercise can be applied to research-based learning programs in high school, as it is an effective learning tool.
RESUMO
We investigated the ability of type I collagen telopeptides to bind neighboring collagen molecules, which is thought to be the initial event in fibrillogenesis. Limited hydrolysis by actinidain protease produced monomeric collagen, which consisted almost entirely of alpha1 and alpha2 chains. As seen with ultrahigh resolution scanning electron microscopy, actinidain-hydrolyzed collagen exhibited unique self-assembly, as if at an intermediate stage, and formed a novel suprastructure characterized by poor fibrillogenesis. Then, the N- and C-terminal sequences of chicken type I collagen hydrolyzed by actinidain or pepsin were determined by Edman degradation and de novo sequence analysis with matrix-assisted laser desorption ionization-tandem time-of-flight mass spectrometry, respectively. In the C-telopeptide region of the alpha1 chain, pepsin cleaved between Asp(1035) and Phe(1036), and actinidain between Gly(1032) and Gly(1033). Thus, the actinidain-hydrolyzed alpha1 chain is shorter at the C terminus by three residues, Gly(1033), Phe(1034), and Asp(1035). In the alpha2 chain, both proteases cleaved between Glu(1030) and Val(1031). We demonstrated that a synthetic nonapeptide mimicking the alpha1 C-terminal sequence including GFD weakly inhibited the self-assembly of pepsin-hydrolyzed collagen, whereas it remarkably accelerated that of actinidain-hydrolyzed collagen. We conclude that the specific GFD sequence of the C-telopeptide of the alpha1 chain plays a crucial role in stipulating collagen suprastructure and in subsequent fibril formation.
Assuntos
Aminoácidos/química , Colágeno Tipo I/química , Colágeno/química , Cisteína Endopeptidases/química , Animais , Galinhas , Dicroísmo Circular , Frutas , Hidrólise , Metaloproteinases da Matriz/química , Microscopia Eletrônica de Varredura/métodos , Peptídeos/química , Estrutura Terciária de Proteína , Pele/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodosRESUMO
Cells recognize collagen fibrils as the first step in the process of adherence. Fibrils of chicken skin actinidain-hydrolyzed collagen (low adhesive scaffold collagen, LASCol), in which the telopeptide domains are almost completely removed, cause adhering cells to form spheroids instead of adopting a monolayer morphology. Our goal was to elucidate the ultrastructure of the LASCol fibrils compared with pepsin-hydrolyzed collagen (PepCol) fibrils. At low concentration of 0.2 mg/mL, the time to reach the maximum increasing rate of turbidity for LASCol was all slower than that for PepCol. Differential scanning calorimetry showed that the thermal stability of collagen self-assembly changed significantly between pH 5.5 and pH 6.6 with and without a small number of telopeptides. However, the calorimetric enthalpy change did not vary much in that pH range. The melting temperature of LASCol fibrils at pH 7.3 was 55.1 °C, whereas PepCol fibrils exhibited a peak around 56.9 °C. The D-periodicity of each fibril was the same at 67 nm. Nevertheless, the looseness of molecular packing in LASCol fibrils was demonstrated by circular dichroism measurements and immuno-scanning electron microscopy with a polyclonal antibody against type I collagen. As there is a close relationship between function and structure, loosely packed collagen fibrils would be one factor that promotes cell spheroid formation.
Assuntos
Colágeno/química , Cisteína Endopeptidases/química , Peptídeo Hidrolases/química , Pele/química , Animais , Adesão Celular , Galinhas , Concentração de Íons de Hidrogênio , Hidrólise , Camundongos , Microscopia de Força Atômica , Células NIH 3T3 , Estabilidade Proteica , Análise Espectral , TermodinâmicaRESUMO
Smiling is believed to make people look younger. Ganel and Goodale (Psychon Bull Rev 25(6):612-616, 10.3758/s13423-017-1306-8, 2018) proposed that this belief is a misconception rooted in popular media, based on their findings that people actually perceive smiling faces as older. However, they did not clarify whether this misconception can be generalized across cultures. We tested the cross-cultural validity of Ganel and Goodale's findings by collecting data from Japanese and Swedish participants. Specifically, we aimed to replicate Ganel and Goodale's study using segregated sets of Japanese and Swedish facial stimuli, and including Japanese and Swedish participants in groups asked to estimate the age of either Japanese or Swedish faces (two groups of participants × two groups of stimuli; four groups total). Our multiverse analytical approach consistently showed that the participants evaluated smiling faces as older in direct evaluations, regardless of the facial stimuli culture or their nationality, although they believed that smiling makes people look younger. Further, we hypothesized that the effect of wrinkles around the eyes on the estimation of age would vary with the stimulus culture, based on previous studies. However, we found no differences in age estimates by stimulus culture in the present study. Our results showed that we successfully replicated Ganel and Goodale (2018) in a cross-cultural context. Our study thus clarified that the belief that smiling makes people look younger is a common cultural misconception.
RESUMO
Titanium is the only metal to which osteoblasts can adhere and on which they can grow and form bone tissue in vivo, resulting in a strong bond between the implant and living bone. This discovery provides the basis for the universal medical application of Ti. However, the biochemical mechanism of bond formation is still unknown. We aimed to elucidate the mechanism of bond formation between collagen, which constitutes the main organic component of bone, and TiO2, of which the entire surface of pure Ti is composed. We analysed the binding between the soluble collagen and TiO2 by chromatography with a column packed with Ti beads of 45 µm, and we explored the association between collagen fibrils and TiO2 (anatase) powders of 0.2 µm. We ran the column of chromatography under various elution conditions. We demonstrated that there is a unique binding affinity between Ti and collagen. This binding capacity was not changed even in the presence of the dissociative solvent 2M urea, but it decreased after heat denaturation of collagen, suggesting the contribution of the triple-helical structure. We propose a possible role of periodically occurring polar amino acids and the collagen molecules in the binding with TiO2.
Assuntos
Colágeno/química , Titânio/química , Ureia/química , Cromatografia Líquida , Colágeno/isolamento & purificação , Desnaturação ProteicaRESUMO
OBJECTIVE: This study aimed to investigate risk factors predictive of local recurrence and/or lymph node metastasis after transoral resection of early hypopharyngeal cancer. METHODS: Forty-nine consecutive patients who underwent transoral videolaryngoscopic surgery (TOVS) as an initial treatment for hypopharyngeal cancer were evaluated. On univariate and multivariate analysis, local recurrence rates were assessed respectively using log-rank test and cox regression analysis according to the following parameters: subsite, pT, mucosal margin, lymphatic invasion, vessel invasion, tumor thickness (> 4 mm vs ≤4 mm), history of esophageal cancer, and multiple Lugol-voiding lesions (LVLs) in the esophagus. Categorical variables were evaluated for their associations with lymph node metastasis using chi-squared test or Fisher's exact test. RESULT: The subsites of primary lesions were piriform sinus in 24 patients, posterior wall in 15 patients, and postcricoid in 10 patients. Thirty patients had esophageal cancer. Local recurrence occurred in 14 patients. Three patients had lymph node metastasis at the time of diagnosis and four patients developed lymph node metastasis after the initial treatment, resulting a total of seven patients having lymph node metastasis. While mucosal margin and LVLs showed significant associations with local recurrence on univariate analysis, only LVLs remained as a significant risk factor on multivariate analysis (P = .0395; hazard ratio = 8.897; 95% confidence interval, 1.113-71.15). Most cases of local recurrence were satisfactorily controlled by repeated TOVS. While multivariate analysis could not be performed due to the small number of the patients with lymph node metastases, venous invasion (P = .0166) and tumor thickness (P = .0092) were significantly associated with lymph node metastasis on univariate analysis. CONCLUSIONS: Local recurrence was more frequent in patients with LVLs, but most of them were salvaged by repeated TOVS. Patients with venous invasion and/or tumor thickness greater than 4 mm should be followed up with special attention to lymph node metastasis.Level of Evidence: 3.
RESUMO
BACKGROUND: Previously we found that a group of phosphorylated proteins (SIBLINGs) in bone binds with the Ti-device, and increases the early bone formation around the Ti-implants remarkably. From these results, we explained the biochemical mechanism of a strong bond between living bone and Ti, which was discovered by Brånemark and colleagues. For the clinical application of our findings, we need a large amount of these proteins or their substitutes. OBJECTIVE: We aimed to create a new molecule that equips with essential functions of SIBLINGs, Ti-binding, and bone enhancement around the Ti implant. METHODS: We chemically phosphorylated chitin and obtained a soluble form of phosphorylated chitin (P-chitin). In this solution, we immersed the Ti-devices of web-form (TW) which we previously developed and obtained the P-chitin coated TWs. Then we tested the P-chitin coated TWs for their calcification ability in vitro, and bone enhancing ability in vivo, by implanting them into rat calvaria. We compared the P-chitin coated TW and the non-coated TW in regard to their calcification and bone enhancing abilities. RESULTS: Ti-devices coated with phosphorylated-chitin induced a ten times higher calcification in vitro at 20 days, and four times more elevated amount of bone formation in vivo at two weeks than the uncoated Ti-device. CONCLUSIONS: Phosphorylated chitin could be a partial substitute of bone SIBLING proteins and are clinically applicable to accelerate bone formation around the Ti implants, thereby achieving the strong bond between living bone and Ti.
Assuntos
Quitina/farmacologia , Implantes Experimentais , Osteogênese/efeitos dos fármacos , Fosfoproteínas/farmacologia , Crânio , Titânio/química , Animais , Regeneração Óssea/efeitos dos fármacos , Calcificação Fisiológica/efeitos dos fármacos , Quitina/química , Quitina/metabolismo , Materiais Revestidos Biocompatíveis/química , Materiais Revestidos Biocompatíveis/farmacologia , Masculino , Teste de Materiais , Fosfoproteínas/química , Fosforilação , Ratos , Ratos Wistar , Crânio/efeitos dos fármacos , Crânio/metabolismo , Crânio/patologia , Crânio/fisiopatologia , Propriedades de Superfície/efeitos dos fármacos , Titânio/farmacologiaRESUMO
OBJECTIVE: To investigate the oncological and functional outcomes of the patients treated with transoral CO2 laser cordectomy for early glottic cancer. PATIENTS AND METHODS: Fifty-five consecutive patients who underwent CO2 laser cordectomy for early glottic cancer were retrospectively reviewed. RESULTS: Overall survival, larynx preservation, and relapse free local control rates were 96%, 100%, and 91%, respectively. Five patients with local recurrences were salvaged with re-cordectomy and/or radiotherapy. In type I cordectomy, VHI-10 consistently improved during postoperative course and VHI-10 at postoperative 12months was significantly better than preoperative value (2.3 vs. 9.4, p=0.02). Perceptual grading, MPT, MFR and AC/DC also improved and were better than preoperative values. In type III cordectomy, shimmer at 12months after cordectomy was significantly better than preoperative value (14.7 vs. 9.3, p=0.007). CONCLUSIONS: These results further support the rationale of CO2 cordectomy as initial and salvage surgery for early glottic cancer.
Assuntos
Neoplasias Laríngeas/cirurgia , Laringoscopia/métodos , Terapia a Laser/métodos , Complicações Pós-Operatórias/fisiopatologia , Prega Vocal/cirurgia , Distúrbios da Voz/fisiopatologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Glote/cirurgia , Humanos , Lasers de Gás , Masculino , Margens de Excisão , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/cirurgia , Estadiamento de Neoplasias , Satisfação do Paciente , Terapia de Salvação , Acústica da Fala , Resultado do TratamentoRESUMO
Back pain is a global health problem with a high morbidity and socioeconomic burden. Intervertebral disc herniation and degeneration are its primary cause, further associated with neurological radiculopathy, myelopathy, and paralysis. The current surgical treatment is principally discectomy, resulting in the loss of spinal movement and shock absorption. Therefore, the development of disc regenerative therapies is essential. Here we show reduced disc damage by a new collagen type I-based scaffold through actinidain hydrolysis-Low Adhesive Scaffold Collagen (LASCol)-with a high 3D spheroid-forming capability, water-solubility, and biodegradability and low antigenicity. In human disc nucleus pulposus and annulus fibrosus cells surgically obtained, time-dependent spheroid formation with increased expression of phenotypic markers and matrix components was observed on LASCol but not atelocollagen (AC). In a rat tail nucleotomy model, LASCol-injected and AC-injected discs presented relatively similar radiographic and MRI damage control; however, LASCol, distinct from AC, decelerated histological disc disruption, showing collagen type I-comprising LASCol degradation, aggrecan-positive and collagen type II-positive endogenous cell migration, and M1-polarized and also M2-polarized macrophage infiltration. Reduced nucleotomy-induced disc disruption through spontaneous spheroid formation by LASCol warrants further investigations of whether it may be an effective treatment without stem cells and/or growth factors for intervertebral disc disease.
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Degeneração do Disco Intervertebral , Deslocamento do Disco Intervertebral , Disco Intervertebral , Adesivos , Animais , Colágeno , RatosRESUMO
BACKGROUND: In 2008, Kano developed a new laryngeal closure technique (Kano's method) for the treatment of severe aspiration. The aim of this study was to evaluate the safety and efficacy of this technique in patients with head and neck cancer. METHODS: Since June 2014 until March 2018, six patients underwent Kano's method for management of severe aspiration after the treatment of head and neck cancers. The anterior parts of the thyroid and the cricoid cartilages were excised widely. The glottis was closed by suturing bilateral vocal folds and reinforced by the sternohyoid muscle. A tracheostoma was created with skin flaps, subglottic mucosal flaps, and stumps of cricoid and trachea cartilages. RESULTS: No severe complications were observed after the surgery. Oral intake improved without developing aspiration. CONCLUSIONS: Kano's method can provide satisfactory functional results with minimal invasion for treating severe aspiration after advanced surgery, chemotherapy, and/or chemoradiotherapy, in patients with head and neck cancer. LEVEL OF EVIDENCE: 4.
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OBJECTIVES: To evaluate the impact of retropharyngeal lymph node (RPLN) dissection in the surgical treatment of hypopharyngeal cancer. METHODS: Between 2007 and 2016, 98 previously untreated patients with pathological diagnosed hypopharyngeal squamous cell carcinoma underwent total pharyngolaryngectomy and bilateral neck dissection at Kobe University Hospital. Bilateral dissection of RPLN was simultaneously performed in all patients. Pharynx was reconstructed with free jejunal transfer in 94 patients and primarily closed in 4 patients. Postoperative chemoradiotherapy was performed in patients with high risk factors including metastasis to RPLN, multiple lymph node metastasis, extranodal invasion, and/or positive/close surgical margins. RESULTS: The median follow-up period was 25 months ranging from 1 to 105 months. RPLN adenopathy was preoperatively identified in 9 patients in FDG-positron emission tomography. All of them had pathologically diagnosed RPLN metastases, which had been controlled in all patients during the observation periods. Among the other 89 patients, 7 patients had RPLN metastasis. The 2-year overall survival rates of the patients with and without RPLN metastasis were 65.7% and 69.8% (P = .61), respectively. CONCLUSIONS: In the present study, patients with RPLN metastasis showed equally favorable oncological outcome compared with patients without RPLN metastasis. At least, ipsilateral RPLN dissection should be considered in the surgical treatment of advanced hypopharyngeal cancer and multiple neck lymph node metastasis regardless of primary subsite.
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Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/cirurgia , Neoplasias Hipofaríngeas/patologia , Neoplasias Hipofaríngeas/cirurgia , Excisão de Linfonodo , Faringectomia , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/mortalidade , Feminino , Humanos , Neoplasias Hipofaríngeas/mortalidade , Masculino , Pessoa de Meia-Idade , Esvaziamento Cervical , Estudos Retrospectivos , Taxa de Sobrevida , Resultado do TratamentoRESUMO
In molluscan shellfish, pigmentation is frequently observed in the calcified shell, but the molecular basis of this process is not understood. Here, we report two tyrosinase proteins (Pfty1 and Pfty2) found in the prismatic shell layer of the pearl oyster Pinctada fucata; this layer is recognized as the pigmented region in P. fucata. The protein sequences were deduced from the corresponding cDNAs and confirmed by MALDI-TOF/TOF analysis. The sequences suggest that both tyrosinases have two copper-binding sites in similar N-terminal domains that are homologous to tyrosinases of cephalopods and hemocyanins of gastropods. In turn, this suggests that bivalve tyrosinases are evolved from a common ancestral copper-binding protein in the mollusc. Pfty1 and Pfty2 were specifically expressed in the mantle, and their expression in the mantle is different from each other, suggesting that these tyrosinases have distinctive roles in melanogenesis in shells.
Assuntos
Perfilação da Expressão Gênica , Monofenol Mono-Oxigenase/química , Monofenol Mono-Oxigenase/genética , Pinctada/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Sítios de Ligação , Northern Blotting , Proteínas de Transporte/química , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Cobre/metabolismo , DNA Complementar/química , DNA Complementar/genética , Eletroforese em Gel de Poliacrilamida , Regulação Enzimológica da Expressão Gênica , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Dados de Sequência Molecular , Monofenol Mono-Oxigenase/metabolismo , Filogenia , Pinctada/enzimologia , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
BACKGROUND: Quality of voice after immediate recurrent laryngeal nerve (RLN) reconstruction in thyroid cancers has not been thoroughly studied. METHODS: Thirteen patients with fixed vocal cords (fixed vocal cord group) and 8 patients with intact or impaired mobile vocal cords (mobile vocal cord group) who had immediate RLN reconstruction simultaneously with total thyroidectomy, and patients who had arytenoid adduction and thyroplasty for vocal cord paralysis caused by previous surgery (arytenoid adduction thyroplasty group) were enrolled in this study. RESULTS: Preoperative phonation efficiency index was significantly lower (p = .008) in the fixed vocal cord group than in the mobile vocal cord group. One year after surgery, all voice parameters of the patients in the fixed vocal cord group had improved, compared with their preoperative data. The fixed vocal cord group had attained satisfactory voice qualities equivalent to those of the mobile vocal cord group in terms of various voice parameters. CONCLUSION: The present results support the idea that immediate RLN reconstruction at the time of surgery for thyroid cancers may spare the need for subsequent arytenoid adduction thyroplasty even in the patients with preoperatively fixed vocal cords. © 2016 Wiley Periodicals, Inc. Head Neck 39: 427-431, 2017.
Assuntos
Procedimentos de Cirurgia Plástica/métodos , Traumatismos do Nervo Laríngeo Recorrente/prevenção & controle , Neoplasias da Glândula Tireoide/cirurgia , Tireoidectomia/efeitos adversos , Paralisia das Pregas Vocais/prevenção & controle , Adulto , Idoso , Estudos de Coortes , Terapia Combinada/métodos , Feminino , Hospitais Universitários , Humanos , Japão , Laringoplastia/métodos , Masculino , Pessoa de Meia-Idade , Cuidados Pré-Operatórios/métodos , Traumatismos do Nervo Laríngeo Recorrente/etiologia , Estudos Retrospectivos , Neoplasias da Glândula Tireoide/patologia , Tireoidectomia/métodos , Fatores de Tempo , Resultado do Tratamento , Paralisia das Pregas Vocais/etiologia , Qualidade da VozRESUMO
Effects of salts on the activity and stability of actinidain were examined. With increasing salt concentration up to 0.5 M, the activity (kcat/Km) for N-alpha-Cbz-L-lysine p-nitrophenyl ester decreased to 40% of that in the absence of salt. The inhibitor constant Ki of LiCl, NaCl, and KCl was 0.16-0.43 M. With 3 M KCl and NaCl, the specificity constant kcat/Km recovered to 110 and 75%, respectively. No re-activation was observed with LiCl. The inhibition and re-activation were dependent on the changes in both Km and kcat, whereas no CD change was observed. The tryptophan fluorescence of actinidain was not affected by 0-0.5 M salt, but a considerable decrease in its intensity was observed with increasing salt concentration from 0.5 to 3.0 M. These results suggest that the inhibition observed with the lower salt concentration (<0.5 M) is due to attenuation of the electrostatic interaction between the enzyme and substrate, and the higher concentration (0.5-3.0 M) induces structural change in the states of tryptophan residues, which is associated with the re-activation. Actinidain keeps considerably high activity and stability even in the presence of 3 M salts.
Assuntos
Actinidia/enzimologia , Cisteína Endopeptidases/metabolismo , Esterases/metabolismo , Sais/farmacologia , Actinidia/efeitos dos fármacos , Dicroísmo Circular , Cisteína Endopeptidases/química , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Estabilidade Enzimática/efeitos dos fármacos , Esterases/química , Hidrólise/efeitos dos fármacos , Cinética , Cloreto de Lítio/farmacologia , Peptídeo Hidrolases/química , Peptídeo Hidrolases/metabolismo , Cloreto de Potássio/farmacologia , Cloreto de Sódio/farmacologia , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , Especificidade por SubstratoRESUMO
Random sequencing of molecules from a cDNA library constructed from mantle mRNA of the pearl oyster Pinctada fucata was used to obtain information on organic matrix proteins in the shell. In the determined sequences, we identified 7 distinct cDNAs encoding similar glycine-rich domains. Complete sequence analysis of these cDNAs showed that the predicted sequences of the proteins, which we named shematrins, possessed similar domains comprising repeat sequences of two or more glycines, followed by a hydrophobic amino acid. In addition, in shematrin-1, -2 and -3, a repeat domain designated as XGnX (where X is a hydrophobic amino acid) was conserved. It is of further note that all the shematrin proteins have RKKKY, RRKKY or RRRKY as their C-terminal sequence. According to northern blot analysis, all shematrins are exclusively expressed in the mantle, and particularly in the edge region of the mantle; furthermore, peptide fragments similar to shematrin-1 and -2 were detected in the prismatic layer of shells by MALDI-TOF/TOF MS analysis. These findings suggest that many of shematrins are synthesized in the mantle edge and secreted into the prismatic layer of the shell, where the protein family is thought to provide a framework for calcification.
Assuntos
Calcificação Fisiológica/genética , Proteínas da Matriz Extracelular/genética , Glicina/genética , Pinctada/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Proteínas da Matriz Extracelular/isolamento & purificação , Biblioteca Gênica , Dados de Sequência Molecular , Estrutura Terciária de Proteína/genética , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Distribuição TecidualRESUMO
BACKGROUND AND PURPOSE: The aim of this study was to retrospectively analyse the outcomes of cases of adenoid cystic carcinomas (ACCs) of the head and neck that were treated at a single institution with particle therapy consisting of either protons or carbon ions. METHODS AND MATERIALS: Between February 2002 and March 2012, 80 patients were treated with proton therapy (PT) or carbon ion therapy (CIT) alone. PT and CIT were employed in 40 (50%) patients each, and more than half of the patients received 65.0 GyE in 26 fractions (n=47, 59%). RESULTS: The median duration of follow-up was 38 months (range, 6-115 months). For all patients, the 5-year for overall survival (OS) rate, progression-free survival (PFS) rate, and local control (LC) rate were 63%, 39%, and 75%, respectively. No significant differences between PT and CIT were observed. The 5-year LC rates for T4 and inoperable cases were 66% and 68%, respectively. Twenty-one patients (26%) experienced grade 3 or greater late toxicities, including three patients who developed grade 5 bleeding from nasopharyngeal ulcers. CONCLUSIONS: Particle radiotherapy for ACC achieves favourable LC, and its efficacy in inoperable or T4 cases is promising. There were no significant differences between PT and CIT in terms of OS, PFS and LC.