RESUMEN
In the present study, we have attempted to elucidate the effects of the intra-arterial injection of microglia on the global ischemia-induced functional and morphological deficits of hippocampal CA1 neurons. When PKH26-labeled immortalized microglial cells, GMIR1, were injected into the subclavian artery, these exogenous microglia were found to accumulate in the hippocampus at 24 h after ischemia. In hippocampal slices prepared from medium-injected rats subjected to ischemia 48 h earlier, synaptic dysfunctions including a significant reduction of synaptic responses and a marked reduction of long-term potentiation (LTP) of the CA3-CA1 Schaffer collateral synapses were observed. At this stage, however, neither significant neuronal degeneration nor gliosis was observed in the hippocampus. At 96 h after ischemia, there was a total loss of the synaptic activity and a marked neuronal death in the CA1 subfield. In contrast, the basal synaptic transmission and LTP of the CA3-CA1 synapses were well preserved after ischemia in the slices prepared from the microglia-injected animals. We also found the microglial-conditioned medium (MCM) to significantly increase the frequency of the spontaneous postsynaptic currents of CA1 neurons without affecting the amplitude, thus indicating that MCM increased the provability of the neurotransmitter release. The protective effect of the intra-arterial injected microglia against the ischemia-induced neuronal degeneration in the hippocampus was substantiated by immunohistochemical and immunoblot analyses. Furthermore, the arterial-injected microglia prevented the ischemia-induced decline of the brain-derived neurotrophic factor (BDNF) levels in CA1 neurons. These observations strongly suggest that the arterial-injection of microglia protected CA1 neurons against the ischemia-induced neuronal degeneration. The restoration of the ischemia-induced synaptic deficits and the resultant reduction of the BDNF levels in CA1 neurons, possibly by the release of diffusible factor(s), might thus contribute to the protective effect of the arterial-injection of microglia against ischemia-induced neuronal degeneration.
Asunto(s)
Hipocampo/patología , Isquemia , Microglía/fisiología , Neuronas/fisiología , Animales , Western Blotting/métodos , Línea Celular , Medios de Cultivo Condicionados/farmacología , Modelos Animales de Enfermedad , Hipocampo/fisiopatología , Inmunohistoquímica/métodos , Técnicas In Vitro , Inyecciones Intraarteriales/métodos , Isquemia/patología , Isquemia/fisiopatología , Isquemia/terapia , Masculino , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Microglía/química , Proteínas del Tejido Nervioso/metabolismo , Neuronas/efectos de los fármacos , Técnicas de Placa-Clamp/métodos , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
The histologic pattern of tumor growth at tumor-nontumor boundaries was studied in 60 livers bearing hepatocellular carcinoma. Three growth patterns, arbitrarily described as "sinusoidal," "replacing," and "encapsulated," were distinguished. Cancer cells growing in the sinusoids between liver cell cords (sinusoidal pattern) were anaplastic; those growing in an expansile fashion and acquiring a fibrous capsule (encapsulated pattern) were most differentiated; and those growing into the cord of liver cells and replacing them (replacing pattern) were differentiated to an intermediate degree. There was certain relation between the histologic growth patterns and gross morphologic features of the tumors. Test results for 20 of the 60 cases were positive for serum hepatitis B surface antigen (HBsAg), and the livers also contained orcein-positive cells. Orcein-positive cells were frequently seen at the border between tumor and parenchyma. Cells containing HBsAg as an orcein-positive inclusion were present in cancer tissue in three cases. When serial sections were made from such areas and stained alternately with hematoxylin and eosin and orcein, it was found that these cells were hepatocytes blended with cancer cells. This phenomenon was related to the growth pattern of tumor cells. Orcein-positive cells were never found in metastatic lesions.
Asunto(s)
Carcinoma Hepatocelular/patología , Antígenos de Superficie de la Hepatitis B/análisis , Neoplasias Hepáticas/patología , Adulto , Anciano , Carcinoma Hepatocelular/inmunología , Femenino , Humanos , Neoplasias Hepáticas/inmunología , Masculino , Persona de Mediana Edad , Oxazinas , Coloración y EtiquetadoAsunto(s)
Carboxiliasas/análisis , Plantas/enzimología , Ribulosa-Bifosfato Carboxilasa/análisis , Rastreo Diferencial de Calorimetría , Fenómenos Químicos , Química , Dicroismo Circular , Estabilidad de Medicamentos , Concentración de Iones de Hidrógeno , Cinética , Luz , Sustancias Macromoleculares , Medicago sativa/enzimología , Peso Molecular , Conformación Proteica , Dispersión de Radiación , Espectrofotometría UltravioletaAsunto(s)
Transferrina , Animales , Apoproteínas , Sitios de Unión , Pollos , Dicroismo Circular , Cobre , Humanos , Rayos Infrarrojos , Unión Proteica , Conformación Proteica , Espectrofotometría , Transferrina/sangre , Rayos UltravioletaAsunto(s)
Aluminio , Conalbúmina , Proteínas del Huevo , Galio , Transferrina , Animales , Sitios de Unión , Pollos , Humanos , Unión Proteica , Conformación Proteica , Solventes , Espectrofotometría Ultravioleta , Triptófano/análisisAsunto(s)
Alquinos , Aminas , Acetileno , Ácidos , Aldehídos , Fenómenos Químicos , Química , Ésteres , Nitrógeno , Análisis EspectralAsunto(s)
Aldehídos , Aminas , Cetonas , Alquenos , Fenómenos Químicos , Química , Nitrógeno , Análisis EspectralAsunto(s)
Imidas , Maleatos , Fenómenos Químicos , Química , Cinamatos , Espectroscopía de Resonancia Magnética , Maleimidas , MetilaminasAsunto(s)
Fumaratos , Imidas , Maleatos , Pirazinas , Fenómenos Químicos , Química , Espectroscopía de Resonancia Magnética , Maleimidas , Modelos EstructuralesRESUMEN
Circular dichroism, differential scanning calorimetry and light-scattering measurements of ribulose 1,5-bisphosphate carboxylase (E.C. 4.1.1.39) from alfalfa, spinach and tobacco show: a) The conformation and thermal stability of the native carboxylases are sensitive to changes in pH and to activation of the enzyme with Mg(2+) and CO(2). The helical content, denaturation temperature (T(d)) and specific enthalpy of denaturation (Deltaq) decreased with increase in pH. Addition of Mg(2+) and CO(2) at pH 9 increased T(d) by 4 to 5 C; at pH 7.5 the changes in T(d) were smaller. b) Addition of mercurials produced changes in conformation and thermal stability. The decrease in helical content of the enzymes with increase in pH was enhanced by the addition of p-chloromercuribenzoate. At pH 9, addition of p-chloromercuribenzoate or of 1-(3-(chloromercuri)-2-methoxypropyl)urea decreased T(d) by 11.4 to 20.2 C and Deltaq by 2.1 to 2.8 calories per gram. c) The spinach carboxylase undergoes the largest and the tobacco the smallest changes in conformation and thermal stability upon change in pH or treatment with mercurials. d) The calorimetric data suggest that the large and small subunits are heat denatured independently but at the same temperature. e) Light scattering measurements at pH 9 of p-chloromercuribenzoate treated tobacco enzyme showed that there is no dissociation into subunits upon heating to temperatures greater than T(d). A ;ball and string' model for the carboxylase molecule is proposed to reconcile independence of subunit denaturation with apparent strong interactions between subunits.
RESUMEN
Fe(III), Cu(II), Co(III), and Mn(III) complexes of ovo- and human serum transferrins show resonance enhanced Raman bands near 1600, 1500, 1270, and 1170 cm-1 upon excitation with laser frequencies which fall within the visible absorption bands of those metalloproteins. Comparison of the visible absorption and resonance Raman spectra of the Cu(II)-transferrin complexes with those for the Cu(II) model compound, bis(2,4,6-trichlorophenolato)diimidazolecopper(II) monohydrate, indicates that the resonance Raman bands are due to enhancement of phenolic vibrational modes. For the model (Cu(II) compound, a normal coordinate analysis was used to aid our assignment of the observed resonance bands at 1562, 1463, 1311, and 1122 cm-1 to A1 vibrational modes of the 2,4,6-trichlorophenolato moiety. These assignments are consistent with those made for Cu(II)-transferrins. The latter assignments were based upon calculated A1 frequencies for p-methylphenol (Cummings, D.L., and Wood, J.L. (1974), J. Mol. Struct. 20, 1). The wavelength shifts in the resonance bands for the model compound from those for Cu(II)-transferrins are due to the influence of the chloro substituents on the planar vibrations of phenol. These results clearly identify tyrosine as a ligand in copper binding to transferrins.
Asunto(s)
Cobalto , Cobre , Hierro , Manganeso , Compuestos Organometálicos , Transferrina , Sitios de Unión , Rayos Láser , Modelos Químicos , Unión Proteica , Conformación Proteica , Espectrofotometría , Espectrofotometría Ultravioleta , Espectrometría RamanRESUMEN
Raman spectroscopy was used to study the anomalous decrease in the freezing temperature of water produced by an antifreeze glycoprotein obtained from the sera of an Antarctic fish. An active fraction of this glycoprotein has a molecular weight of approximately 18,000 by equilibrium sedimentation compared to an apparent weight of 20 by freezing temperature depression. The Raman spectra of water present in a 1% antifreeze glycoprotein solution and of ice frozen from this solution were indistinguishable from the spectra of pure water and ice, respectively. These results indicate that the bulk properties of water and ice are unaffected by the presence of the antifreeze glycoprotein. Raman measurements on ice grown slowly, using as seed an oriented single crystal of ice in contact with 1% glycoprotein solutions, showed that the active glycoprotein was not excluded from the ice phase. On the other hand, we found that a smaller, inactive glycoprotein was excluded. Comparison of the Raman spectra of active and inactive glycoprotein components as solids, in 5% solutions, and rapidly frozen 5% solutions, showed that the two components differ in conformation and possibly in the environment of their carbohydrate hydroxyls. These observations suggest that hydrogen bonding of the carbohydrate hydroxyls of the active glycoprotein at the ice-solution interface may physically prevent growth of the ice lattice.