RESUMO
Cladosporium cladosporioides is one of the most ubiquitous dematiaceous fungi that seldomly occur human infection. Here, we demonstrate a rare case of pulmonary phaeohyphomycosis with a distinctive pulmonary lesion during the nadir period of outpatient chemotherapy against endometrial cancer. In addition to severe neutropenia, excessive exposure to C. cladosporioides at patient's residence was considered as dominant causative factor. More caution is considered necessary for pulmonary phaeohyphomycosis in patients who receive outpatient chemotherapy and are homebound during neutropenic status.
Assuntos
Abscesso Pulmonar , Feoifomicose , Humanos , Feoifomicose/tratamento farmacológico , Pacientes Ambulatoriais , CladosporiumRESUMO
AIM: To assess the pancreatic groove fat plane in the normal population and compare this with the fat plane in patients with groove pancreatitis or carcinoma using multidetector computed tomography (CT). MATERIAL AND METHODS: The pancreatic groove fat plane was evaluated retrospectively in 460 normal subjects (normal group), and in 25 patients with groove pancreatitis or carcinoma (pathology group) using 5 mm- and 1 mm-thick slices of unenhanced axial multidetector CT images. Two investigators independently assessed the degree of pancreatic groove fat plane visualisation using a four-point scale (grade 1: visualisation of 0-25%, grade 2: 26-50%, grade 3: 51-75%, grade 4: 76-100%). Pancreatic parenchymal condition, age, sex, body mass index, diabetes mellitus, and dyslipidaemia were also evaluated. RESULTS: The interobserver agreement for the visualisation grades was almost perfect (k-value = 0.95). In the normal group, grade 4 visualisation of the pancreatic groove fat plane was more common in those aged >80 years (78.6%) compared with younger age groups. Pancreatic atrophy and fatty infiltration significantly improved fat plane visualisation. In the pathology group, grade 4 visualisation of the pancreatic groove fat plane was not seen in either groove carcinoma or pancreatitis. A cut-off point of ≤50% visualisation of the pancreatic groove fat plane showed 95% sensitivity and 82% specificity for detecting possible abnormalities in older patients (>60 years). The clinical factors investigated were not significantly related to pancreatic groove fat plane visualisation. CONCLUSION: Pancreatic groove fat plane visualisation could be a good predictor for detecting groove abnormalities.
Assuntos
Neoplasias Pancreáticas/diagnóstico por imagem , Pancreatite Crônica/diagnóstico por imagem , Tecido Adiposo/diagnóstico por imagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Tomografia Computadorizada Multidetectores , Variações Dependentes do Observador , Estudos Retrospectivos , Adulto JovemRESUMO
BACKGROUND: Apheresis platelets (APs) have gained favour over whole blood-derived platelets on the presumption that they are less likely to provoke alloimmunization to red-blood-cell antigens. CASE REPORTS: Non-D Rh antibodies appeared in three patients after apheresis platelet transfusion. Anti-C and anti-E arose in two female patients with previous antigen exposure. Both anti-c and anti-E arose in a male recipient with no prior transfusion history. MATERIALS AND METHODS: Fifty APs were analysed for residual RBCs and RBC-derived microparticles, using samples obtained from a local blood centre. Cells and microparticles were quantified with a flow cytometry gating scheme, using PE-labelled anti-CD235a (glycophorin A) and FITC-labelled anti-CD41a (platelet gp IIb/IIIa) to distinguish lineage. RESULTS: Apheresis platelets were found to contain a mean of 7·5×10(6) (95% C.I. [6·3-8·5×10(6) ]) RBCs on one manufacturer's device and 5·2×10(6) (95% C.I. [4·0-6·3×10(6) ]) RBCs on another's. RBC-derived microparticles averaged 210·7×10(6) (95% C.I. [166·2-254·2×10(6) ]) on one manufacturer's device and 232·3×10(6) (95% C.I. [194·3-272·9×10(6) ]) on another's. These counts all correspond to volumes of <1 µl. CONCLUSION: Despite RBC contamination of APs below commonly accepted thresholds for Rh immunogenicity, AP transfusion can provoke non-D Rh antibody formation. RBC-derived microparticles, smaller but more numerous than RBCs, are volumetrically comparable and may be a hitherto underappreciated antibody stimulus. Further microparticle research will guide considerations of extended phenotypic matching of platelet components.
Assuntos
Incompatibilidade de Grupos Sanguíneos/sangue , Incompatibilidade de Grupos Sanguíneos/imunologia , Micropartículas Derivadas de Células/imunologia , Membrana Eritrocítica/imunologia , Isoanticorpos , Transfusão de Plaquetas , Sistema do Grupo Sanguíneo Rh-Hr , Idoso , Remoção de Componentes Sanguíneos , Incompatibilidade de Grupos Sanguíneos/etiologia , Feminino , Humanos , Isoanticorpos/sangue , Isoanticorpos/imunologia , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: This study aimed to analyse the perioperative results from a national dataset of rectal cancer resections in elderly patients. METHODS: The clinical records of patients undergoing rectal cancer surgery between 2012 and 2014 were retrieved from the Japanese National Clinical Database and analysed retrospectively. Patients were categorized according to age and those 80 years or older were defined as elderly. Subgroups were also defined according to the surgical approach (laparoscopy versus open surgery). The short-term outcomes, including mortality, anastomotic leak, surgical site infections and medical complications were compared between subgroups. RESULTS: Of 56 175 patients undergoing rectal cancer surgery, some 6717 patients were elderly and laparoscopy was performed in 46.8 per cent of the sample. When comparing laparoscopy and open surgery in elderly patients, the operative mortality rate (1.5 versus 2.8 per cent; P < 0.001), the incidence of anastomotic leakage (5.2 versus 6.5 per cent; P = 0.026), surgical site infections (6.0 versus 8.0 per cent; P = 0.001), pneumonia (1.4 versus 2.5 per cent; P = 0.001), renal failure (0.7 versus 1.3 per cent; P = 0.016) and cardiac events (0.3 versus 0.8 per cent; P = 0.008) were lower for laparoscopy than for open surgery. The overall complication rate in elderly patients (19.5 per cent) was comparable to that in the younger group (P = 0.07). However, incidence of systemic complications was significantly higher in elderly than in younger patients (all P < 0.001). CONCLUSION: Laparoscopy was safe and feasible in elderly patients compared with open surgery. However, the rates of systemic complications were significantly higher than in younger patients.
Assuntos
Laparoscopia/efeitos adversos , Complicações Pós-Operatórias/epidemiologia , Neoplasias Retais/cirurgia , Idoso , Idoso de 80 Anos ou mais , Fístula Anastomótica/epidemiologia , Bases de Dados Factuais , Estudos de Viabilidade , Feminino , Humanos , Japão/epidemiologia , Laparoscopia/mortalidade , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Pneumonia/epidemiologia , Neoplasias Retais/mortalidade , Estudos Retrospectivos , Infecção da Ferida Cirúrgica/epidemiologia , Resultado do TratamentoRESUMO
BACKGROUND: Robot-assisted laparoscopic surgery has several advantages over conventional laparoscopy. However, population-based comparative studies for low anterior resection are limited. This article aimed to compare peri-operative results of robot-assisted low anterior resection (RALAR) and laparoscopy. METHODS: This retrospective cohort study used data from patients treated with RALAR or conventional laparoscopic low anterior resection (CLLAR) between October 2018 and December 2019, as recorded in the Japanese National Clinical Database, a data set registering clinical information, perioperative outcomes, and mortality. Of note, the registry does not include information on the tumour location (centimetres from the anal verge) and diverting stoma creation. Perioperative outcomes, including rate of conversion to open surgery, were compared between RALAR and CLLAR groups. Confounding factors were adjusted for using propensity score matching. RESULTS: Of 21 415 patients treated during the study interval, 20 220 were reviewed. Two homogeneous groups of 2843 patients were created by propensity score matching. The conversion rate to open surgery was significantly lower in the RALAR group than in the CLLAR group (0.7 versus 2.0 per cent; P < 0.001). The RALAR group had a longer operating time (median: 352 versus 283 min; P < 0.001), less intraoperative blood loss (15 versus 20 ml; P < 0.001), a lower in-hospital mortality rate (0.1 versus 0.5 per cent; P = 0.007), and a shorter postoperative hospital stay (median: 13 versus 14 days; P < 0.001) compared with the CLLAR group. The CLLAR group had a lower rate of readmission within 30 days (2.4 versus 3.3 per cent; P = 0.045). CONCLUSION: These data highlight the reduced conversion rate, in-hospital mortality rate, intraoperative blood loss, and length of postoperative hospital stay for rectal cancer surgery in patients treated using robot-assisted laparoscopic surgery compared with laparoscopic low anterior resection.
Assuntos
Laparoscopia , Neoplasias Retais , Robótica , Humanos , Japão/epidemiologia , Neoplasias Retais/cirurgia , Estudos Retrospectivos , Resultado do TratamentoRESUMO
The antimicrobial peptide LL-37 is known to have a potent LPS-neutralizing activity in monocytes and macrophages. Recently, LL-37 in gingival crevicular fluids is suggested to be the major protective factor preventing infection of periodontogenic pathogens. In this study, we tried to address the effect of LL-37 on proinflammatory responses of human gingival fibroblasts (HGFs) stimulated with Toll-like receptor (TLR)-stimulant microbial compounds. LL-37 potently suppressed LPS-induced gene expression of IL6, IL8 and CXCL10 and intracellular signaling events, degradation of IRAK-1 and IkappaBalpha and phosphorylation of p38 MAPK and IRF3, indicating that the LPS-neutralizing activity is also exerted in HGFs. LL-37 also suppressed the expression of IL6, IL8 and CXCL10 induced by the TLR3 ligand poly(I:C). LL-37 modestly attenuated the expression of IL6 and IL8 induced by the TLR2/TLR1 ligand Pam(3)CSK(4), but did not affect the expression induced by the TLR2/TLR6 ligand MALP-2. Interestingly, LL-37 rather upregulated the expression of IL6, IL8 and CXCL10 induced by another TLR2/TLR6 ligand FSL-1. Thus, the regulatory effect of LL-37 is differently exerted towards proinflammatory responses of HGFs induced by different microbial stimuli, which may lead to unbalanced proinflammatory responses of the gingival tissue to infection of oral microbes.
Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Quimiocina CXCL10/biossíntese , Fibroblastos/efeitos dos fármacos , Interleucina-6/biossíntese , Interleucina-8/biossíntese , Infecções Bacterianas/imunologia , Células Cultivadas , Quimiocina CXCL10/genética , Quimiocina CXCL10/metabolismo , Fibroblastos/imunologia , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Gengiva/patologia , Humanos , Imunidade Inata , Quinases Associadas a Receptores de Interleucina-1/antagonistas & inibidores , Interleucina-6/genética , Interleucina-6/metabolismo , Interleucina-8/genética , Interleucina-8/metabolismo , Ligantes , Doenças Periodontais/imunologia , Transdução de Sinais/efeitos dos fármacos , Receptor 2 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor 3 Toll-Like/imunologia , Receptor 3 Toll-Like/metabolismo , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , CatelicidinasRESUMO
BACKGROUND: Laparoscopy-assisted distal gastrectomy (LAG) is gaining acceptance for treating early gastric cancer. However, the long-term quality of life after LAG for gastric cancer is unknown. This study compared the long-term quality of life after LAG versus open distal gastrectomy (ODG) for early gastric cancer. METHOD: This study included 53 patients who underwent LAG and 37 patients who underwent ODG for treatment of early gastric cancer. Quality of life was evaluated on the basis of a 22-item questionnaire that addressed food tolerance and mental and physical conditions, scored on a scale of 1-3. RESULTS: The mean follow-up periods after LAG and ODG were 99.3 and 97.0 months, respectively. Although the majority of patients who had undergone LAG were consuming a normal diet and had weight loss of less than 5 kg, all 22 items and the total score of the LAG group were comparable to those of the ODG group. However, the incidence of postoperative intestinal obstruction was significantly lower in the LAG group than in the ODG group (1% vs. 13%, p < 0.05). CONCLUSIONS: LAG is equivalent to ODG with respect to long-term quality of life and is associated with a reduced incidence of postoperative intestinal obstruction.
Assuntos
Gastrectomia/métodos , Laparoscopia , Qualidade de Vida , Neoplasias Gástricas/cirurgia , Idoso , Dieta , Feminino , Seguimentos , Gastrectomia/efeitos adversos , Humanos , Incidência , Obstrução Intestinal/epidemiologia , Obstrução Intestinal/etiologia , Masculino , Pessoa de Meia-Idade , Período Pós-Operatório , Redução de PesoRESUMO
It has been found that beta-catenin, a key regulator of the cadherin-mediated cell adhesion system, forms complexes with adenomatous polyposis coli (APC) tumor suppressor protein, and beta-catenin expression levels are affected by exogenously induced APC protein. The effects of intrinsic APC protein alteration on beta-catenin expression levels and its subcellular localization were examined in colonic epithelia of eight patients with familial adenomatous polyposis. In all eight patients, beta-catenin was immunostained at the membranes of the cell-to-cell borders in normal epithelial cells, whereas the nuclei and cytoplasms stained intensely in addition to the membranes in both adenoma and cancer cells. beta-Catenin expression levels in tumor tissues were over three times higher than those in corresponding normal mucosae of all of the three patients, whose resected specimens were available for quantitative immunoblot analysis. In these three patients, mutant truncated APC proteins were detected and shown to have lost the central region, including a known beta-catenin binding domain. beta-Catenin was not coimmunoprecipitated with these mutant APC proteins in tumor tissues but was able to be coprecipitated with glutathione S-transferase-fused APC protein containing a beta-catenin binding domain. These results suggest that the absence of wild type APC protein affects the subcellular localization and expression levels of beta-catenin in human tissues.
Assuntos
Polipose Adenomatosa do Colo/metabolismo , Colo/metabolismo , Proteínas do Citoesqueleto/biossíntese , Transativadores , Polipose Adenomatosa do Colo/genética , Polipose Adenomatosa do Colo/patologia , Colo/patologia , Proteínas do Citoesqueleto/genética , Regulação da Expressão Gênica , Humanos , beta CateninaRESUMO
We recently established a new human inflammatory breast cancer (IBC) xenograft (WIBC-9) originating from a patient with IBC. The graft was transplantable in BALB/c nude and severe combined immunodeficient (SCID) mice. WIBC-9 was frequently accompanied by lung metastasis and exhibited erythema of the overlying skin, reflecting its human counterpart. Histological study of the original tumor and WIBC-9 revealed invasive ductal carcinoma with a hypervascular structure of solid nests and marked lymphatic permeation in the overlying dermis. In the central part of the solid nests, absence of endothelial cells, central necrosis, and fibrosis were observed. In vitro, WIBC-9 formed tube-like structures and loops, reflecting its in vivo feature and its human counterpart. WIBC-9 exhibited aneuploidy, ErbB-2 gene amplification, and an absence of estrogen receptor and progesterone receptor, which is consistent with IBC. Comparative studies of WIBC-9, three established non-IBC xenografts, and a human breast cancer cell line (SK-BR3) by reverse transcription-PCR, ELISA, and immunohistochemistry indicated that certain human genes (interleukin 8, vascular epidermal growth factor, basic fibroblast growth factor, angiopoietin 13, Flt-1, Tie-2, and Tie-1) and certain murine genes (integrin alpha(v)beta3, flt-1, tie-2, vascular epidermal growth factor, and CD31) were overexpressed in exposure to tumor cells. The molecular basis and these unique histological features may be associated with aggressive IBC on angiogenic and nonangiogenic pathways.
Assuntos
Neoplasias da Mama/patologia , Endotélio Vascular/patologia , Indutores da Angiogênese/genética , Indutores da Angiogênese/metabolismo , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Caderinas/genética , Caderinas/metabolismo , Divisão Celular , Citocinas/genética , Citocinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Fibrose , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Integrinas/genética , Integrinas/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microscopia Eletrônica , Pessoa de Meia-Idade , Necrose , Transplante de Neoplasias , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Receptor TIE-2 , Receptores de Fatores de Crescimento/genética , Receptores de Fatores de Crescimento/metabolismo , Receptores de Fatores de Crescimento do Endotélio Vascular , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transplante Heterólogo/patologia , Células Tumorais Cultivadas/metabolismo , Células Tumorais Cultivadas/ultraestruturaRESUMO
PURPOSE: Total extraperitoneal preperitoneal (TEP) repair is widely used for inguinal, femoral, or obturator hernia treatment. However, mesh repair is not often used for strangulated hernia treatment if intestinal resection is required because of the risk of postoperative mesh infection. Complete mesh repair is required for hernia treatment to prevent postoperative recurrence, particularly in patients with femoral or obturator hernia. CASES: We treated four patients with inguinocrural and obturator hernias (a 72-year-old male with a right indirect inguinal hernia; an 83-year-old female with a right obturator hernia; and 86- and 82-year-old females with femoral hernias) via a two-stage laparoscopic surgery. All patients were diagnosed with intestinal obstruction due to strangulated hernia. First, the incarcerated small intestine was released and then laparoscopically resected. Further, 8-24 days after the first surgery, bilateral TEP repairs were performed in all patients; the postoperative course was uneventful in all patients, and they were discharged 5-10 days after TEP repair. At present, no hernia recurrence has been reported in any patient. CONCLUSION: The two-stage laparoscopic treatment is safe for treatment of strangulated inguinal, femoral, and obturator hernias, and complete mesh repair via the TEP method can be performed in elderly patients to minimize the occurrence of mesh infection.
Assuntos
Hérnia Femoral/cirurgia , Hérnia Inguinal/cirurgia , Hérnia do Obturador/cirurgia , Herniorrafia/métodos , Obstrução Intestinal/cirurgia , Laparoscopia/métodos , Idoso , Idoso de 80 Anos ou mais , Feminino , Hérnia Femoral/diagnóstico por imagem , Hérnia Inguinal/diagnóstico por imagem , Hérnia do Obturador/diagnóstico por imagem , Humanos , Obstrução Intestinal/diagnóstico por imagem , Intestinos/irrigação sanguínea , Intestinos/diagnóstico por imagem , Intestinos/cirurgia , Masculino , Telas Cirúrgicas , Tomografia Computadorizada por Raios XRESUMO
In order to confirm whether the binding sites for mu-calpain on the inner surface of erythrocyte membranes are substrate proteins themselves, we examined the binding properties of mu-calpain to mu-calpain-pretreated inside-out membranes. When native mu-calpain was incubated with mu-calpain-pretreated membranes, however, newly added calpain was degraded rapidly in a time- and Ca(2+)-dependent manner. Although the degradation of mu-calpain was not inhibited by various proteinase inhibitors, it was strongly inhibited by digestible substrates for calpain that possess the ability to inhibit the binding of mu-calpain to erythrocyte membranes. On the other hand, when mu-calpain inactivated by N-ethylmaleimide was incubated with mu-calpain-pretreated membranes, no degradation was observed. These results indicate that the degradation of mu-calpain occurs on the surface of mu-calpain-modified membranes and that it depends on the autoproteolytic activity of mu-calpain itself. It seems likely that the autoproteolytic activity of mu-calpain is accelerated markedly by some component(s) exposed on the surface of membranes during the pretreatment with mu-calpain. The possibility is thus proposed that cell membranes possess the ability to down-regulate calpain to protect cell membranes from overdegradation by excessively bound calpain. The active factor(s) in the membranes that can accelerate the autoproteolytic degradation of mu-calpain could be almost completely removed from mu-calpain-modified membranes by treatment with Triton X-100.
Assuntos
Calpaína/metabolismo , Membrana Eritrocítica/metabolismo , Animais , Regulação para Baixo , Etilmaleimida/metabolismo , Feminino , Humanos , Hidrólise , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Fosfolipídeos/metabolismo , Ligação Proteica , CoelhosRESUMO
The activities of phytase and alkaline phosphatase in the intestine gradually increased in parallel during development of rats, but the 70K and 90K subunits were expressed differentially; only the 70K subunit was detected at birth, whereas the 90K subunit appeared at the weaning period (3 weeks after birth). When rats were forced to wean at 18 days old and fed laboratory chow, the enzyme activity increased markedly and the 90K subunit appeared within 1 day. These findings suggest that weaning is involved in the change in the subunit composition. Increases in the enzyme activity and amount of the 90K subunit were significantly delayed by feeding weanling animals on casein diet, but induced significantly by feeding them on casein diet supplemented with phytate. Thus induction of the 90K subunit seems to be accelerated by intake of phytic acid in the diet. The Km value of the enzyme from suckling rats for phytate was 5.25 mM, while that of adult rats was 0.213 mM. In contrast, the Km value for p-nitrophenyl phosphate (PNPP) was constant during development. The phytase activity of suckling rats did not show a distinct pH-dependence. These findings suggest that the 90K subunit may play some important roles in expressing an efficient phytase activity.
Assuntos
6-Fitase/biossíntese , Dieta , Intestinos/enzimologia , Animais , Western Blotting , Eletroforese em Gel de Poliacrilamida , Indução Enzimática , Concentração de Íons de Hidrogênio , Intestinos/crescimento & desenvolvimento , Ácido Fítico/administração & dosagem , Ratos , Ratos EndogâmicosRESUMO
Calpain and its endogenous inhibitor, calpastatin, were isolated from erythrocytes of various mammals and their properties were compared. It has been widely believed that mammalian erythrocytes contain only mu-calpain. However, rat and human erythrocytes were found to contain two species of calpain, identified as mu-calpain and m-calpain from their elution positions on DEAE-cellulose column chromatography and their Ca(2+)-requirements. Thus, it is apparent that rat and human erythrocytes contain not only mu-calpain, but m-calpain as well. On the other hand, rabbit erythrocytes contain only mu-calpain. Western blot analysis showed that human and rabbit erythrocytes contain predominantly 70-kDa calpastatin (erythrocyte-type), but unnegligible amounts of 110-kDa calpastatin (tissue-type) are also present. Rat erythrocytes were shown to contain a calpastatin with a molecular mass of approx. 100 kDa almost exclusively; this molecular mass was in perfect coincidence with the mass of the calpastatin in rat lung. These results strongly suggest that rat erythrocytes contain a tissue-type calpastatin. No essential change in the calpain/calpastatin system during maturation of rabbit reticulocytes into mature erythrocytes was observed.
Assuntos
Proteínas de Ligação ao Cálcio/sangue , Calpaína/sangue , Eritrócitos/metabolismo , Animais , Western Blotting , Proteínas de Ligação ao Cálcio/isolamento & purificação , Calpaína/isolamento & purificação , Diferenciação Celular/fisiologia , Cromatografia DEAE-Celulose , Eritrócitos/química , Eritrócitos/enzimologia , Humanos , Coelhos , Ratos , Ratos Wistar , Reticulócitos/metabolismo , Especificidade da EspécieRESUMO
Intracellular calcium-activated neutral proteinase (CANP) in rabbit erythrocytes was activated by an influx of Ca2+ into the cells. The catalytic large subunit changed from the original 79 kDa from to the 77 kDa and 76 kDa forms on activation just in the same manner as occurs in the autolytic activation of purified CANP in vitro. The activation required both extracellular Ca2+ and A23187, and was accompanied by the degradation of some membrane proteins and morphological changes in erythrocyte shape from discocytes to echinodisks, echinocytes, and spherocytes. Exogenously added Cbz-Leu-Leu-Leu-aldehyde inhibited the activation of intracellular CANP as well as the degradation of membrane proteins and the morphological changes indicating that the latter two processes are due to the action of CANP. Leupeptin and E64d were without effect on intracellular CANP.
Assuntos
Calpaína/metabolismo , Eritrócitos/enzimologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais , Proteínas Sanguíneas/metabolismo , Calcimicina/farmacologia , Cálcio/farmacologia , Calpaína/antagonistas & inibidores , Ativação Enzimática , Eritrócitos/citologia , Feminino , Técnicas In Vitro , Leucina/análogos & derivados , Leucina/farmacologia , Leupeptinas/farmacologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , CoelhosRESUMO
The Shumiya cataract rat (SCR) is a hereditary cataract model in which lens opacity appears spontaneously in the nuclear and perinuclear portions at 11-12 weeks of age. It was found that the proteolysis of some crystallins and cytoskeletal proteins is significantly enhanced in cataractous SCR lenses. The calcium concentrations in cataractous lenses rise markedly with age as compared with control lenses and the autolytic product of calpain is also detected in cataractous lenses. In order to provide direct evidence for the involvement of calpain in the proteolytic modification of lens proteins, we developed antibodies exclusively specific to the proteolytic products of some lens proteins produced by the action of calpain and analyzed their degradation during cataractogenesis in SCR by Western blotting and immunohistochemical staining. The results demonstrate that calpain participates in the proteolytic modification of lens proteins, at least alpha-crystallin (A and B chain), betaB1-crystallin, and alpha-fodrin. The proteolytic products formed by the action of calpain on these proteins are detected in cataractous lenses of SCR as young as 8 weeks of age and accumulate with age. It was also found that betaB1-crystallin, originally a soluble protein, is converted to an insoluble form by limited calpain proteolysis. The chaperon-like activity of alpha-crystallin from control lens is markedly reduced by calpain proteolysis in vitro, and alpha-crystallin in opaque lens that has already undergone proteolysis by calpain shows significantly reduced chaperon-like activity. Immunohistochemical studies reveal that the area where the calpain-mediated alpha-crystallin proteolysis is in progress coincides well with the area developing and destined to develop the opacification. These results strongly suggest that calpain may contribute to lens opacification during cataract formation in SCR.
Assuntos
Calpaína/fisiologia , Catarata/metabolismo , Cristalino/metabolismo , Envelhecimento , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais , Cálcio/metabolismo , Proteínas de Transporte/metabolismo , Catarata/genética , Bovinos , Cristalinas/metabolismo , Proteínas dos Microfilamentos/metabolismo , Chaperonas Moleculares/metabolismo , Dados de Sequência Molecular , Fragmentos de Peptídeos/análise , Ratos , Ratos EndogâmicosRESUMO
The molecular mechanism that causes non-adhesive, discoid platelets to transform into sticky dendritic bodies that form blood clumps is a complex series of events. Recently it has become clear that lipid microdomains--also known as rafts--play a crucial role in this process. We have used a non-cytolytic derivative of perfringolysin-O, a cholesterol binding cytolysin, that binds selectively to cholesterol-rich membrane domains, combined with confocal- and immunoelectron microscopy to visualize cholesterol-raft dynamics during platelet adhesion. In resting platelets cholesterol was uniformly distributed on the cell surface and confined to distinct intracellular compartments (i.e. multivesicular bodies, dense granules, and the internal membranes of alpha-granules). Upon interaction with fibrinogen, cholesterol accumulated at the tips of filopodia and at the leading edge of spreading cells. Stimulation with thrombin receptor activating peptide (TRAP) resulted in a similar redistribution of cholesterol towards filopodia. The adhesion-dependent raft aggregation was accompanied by concentration of the tyrosine kinase c-Src and the tetraspanin CD63 in these domains, whereas glycoprotein Ib (GPIb) was not selectively targeted to the raft clusters. c-Src, the tetraspanin CD63, and GPIb were recovered in biochemically isolated low-density membrane fractions. Disruption of rafts by depleting membrane cholesterol had no effect on platelet shape change but inhibited platelet spreading on fibrinogen and TRAP-induced aggregation. Our results demonstrate that cholesterol rafts in platelets are dynamic entities in the membrane that co-cluster with the tyrosine kinase c-Src and the costimulatory molecule CD63 in specialized domains at the cell surface, thereby providing a possible mechanism in functioning as signaling centres.
Assuntos
Antígenos CD/metabolismo , Plaquetas/ultraestrutura , Microdomínios da Membrana/fisiologia , Fosfotransferases/metabolismo , Glicoproteínas da Membrana de Plaquetas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Pseudópodes/química , beta-Ciclodextrinas , Plaquetas/química , Plaquetas/fisiologia , Proteína Tirosina Quinase CSK , Tamanho Celular , Colesterol/metabolismo , Colesterol/fisiologia , Ciclodextrinas/farmacologia , Fibrinogênio , Humanos , Imuno-Histoquímica , Microdomínios da Membrana/química , Fosforilação , Ativação Plaquetária , Transporte Proteico , Proteínas Tirosina Quinases , Receptores de Trombina , Tetraspanina 30 , Quinases da Família srcRESUMO
Morules have been reported in pulmonary endodermal tumors (PET) resembling fetal lung, in thyroid carcinoma, and in endometrial and colonic neoplasms. A morule has biotin-containing optically clear nuclei (OCN) in PET and thyroid carcinoma. Biotin-containing OCN have been also reported in endometrial tissue during pregnancy and in endometrioid carcinoma of the ovary, and it has been postulated that morules or OCN develop under the influence of female sex hormones. The authors report here the first case, to their knowledge, of morules with OCN in a colonic adenoma from a 68-year-old man. The colonic polyp consisted of ordinary tubular adenomatous tissue and morules. Many cells in the morules contained OCN. The OCN were immunopositive for biotin and reacted with streptavidin. The neoplastic cells in the morules were immunopositive for oncofetal antigens. Serum levels of female sex hormones were within the normal range, and no cells in the adenoma were immunopositive for receptors for progesterone and estrogen. The results indicate that OCN are rich in biotin and that morules may be embryologically immature elements that develop independently of influence by female sex hormones.
Assuntos
Adenoma/patologia , Biotina/análise , Núcleo Celular/patologia , Pólipos do Colo/patologia , Idoso , Antígenos de Neoplasias/análise , Neoplasias do Ceco/patologia , Humanos , Técnicas Imunoenzimáticas , Pólipos Intestinais/patologia , Masculino , Estreptavidina/análiseRESUMO
The effect of various components of three semi-solid media on colony formation in two representative retinoblastoma cell lines, Y-79 and WERI, was determined. Diethylaminoethyl dextran was found to be toxic to the cells, and was deleted from the medium. Horse serum was also used without heat treatment. In the most improved culture medium, plating efficiency was 28% for Y-79 cells and 14% for WERI cells, an increase of more than 10 times that of the original formula. In the new medium, both Y-79 and WERI cells showed relatively constant plating efficiency within a certain range, showing that the medium is useful for quantitative clonogenic study of retinoblastoma cells.
Assuntos
Neoplasias Oculares/patologia , Células-Tronco Neoplásicas/citologia , Retinoblastoma/patologia , Ágar , Animais , Fenômenos Fisiológicos Sanguíneos , Linhagem Celular , Meios de Cultura/farmacologia , DEAE-Dextrano/farmacologia , Cavalos , Temperatura Alta , HumanosRESUMO
Establishment of new tumor cell lines is an important first step for biological studies of tumor cells. High success rates in establishing retinoblastoma cell lines have been reported when feeder-layer culture but not liquid-culture techniques were used. Liquid culture is, however, essential for studies in which feeder-layer cells are undesirable. In a previous study, we formulated a medium (RB-- medium), the components of which were almost identical to those of a soft-agar medium developed for colony formation of established retinoblastoma cell lines, in which one cell line from 12 primary retinoblastoma specimens was established. In this study, another medium (RB++ medium), RB-- medium to which 20 microM 2-mercaptoethanol and 375 microM asparagine were added, was tested for its ability to grow retinoblastoma cells from clinical specimens. In the RB++ medium, 6 cell lines from 16 primary sites, 2 from 2 intraocular-recurrent and 3 from 3 metastatic retinoblastomas grew for over a year. The major reason for the apparent improvement of the RB++ medium on the RB-- medium was demonstrated to be the addition of 2-mercaptoethanol.
Assuntos
Neoplasias Oculares/patologia , Mercaptoetanol/farmacologia , Retinoblastoma/patologia , Divisão Celular/efeitos dos fármacos , Pré-Escolar , Meios de Cultura/química , Meios de Cultura/farmacologia , Feminino , Humanos , Lactente , Masculino , Mercaptoetanol/antagonistas & inibidores , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/patologiaRESUMO
Cytogenic studies were performed in a 72-year-old male patient with triple primary cancers including breast, skin and lung. Left breast cancer was diagnosed at the age of 46 and he received mastectomy and thoracic irradiation. Squamous cell carcinoma and Bowen's disease were diagnosed from two separated parts of a skin lesion at the age of 70. Small-cell lung cancer was diagnosed 1 year later, and he received chemotherapy and radiotherapy. Chromosome analysis was carried out on both peripheral lymphocyte and skin fibroblast cultures at the age of 72. Out of 30 fibroblast cells karyotyped at the second passage, 7 cells (23%) consistently showed a reciprocal translocation t(Y;6)(q12;p21). The same translocation was found in one of 200 cells from lymphocyte cultures. The findings suggest that the translocation t(Y;6) might be inherent in nature, and that the patient was a mosaic of 46,XY/46,X,t(Y;6)(q12;p21). These results highlight the constitutional chromosomal abnormality as one of the possible high-risk factors for multiple primary cancers.