Local levels of interleukin-1beta, -4, -6 and tumor necrosis factor alpha in an experimental model of murine osteomyelitis due to staphylococcus aureus

The purpose of this study is to evaluate local levels of interleukin-1 beta (IL-1 beta), -4 (IL-4), -6 (IL-6), and tumour necrosis factor-alpha (TNF-alpha), in a model of murine osteomyelitis due to Staphylococcus aureus.Cytokine levels in supernatants derived from bone homogenates were determined by enzyme-linked immunosorbent assay, for 28 days following the direct implantation of murine tibiae with S.aureus. Levels of IL-1 beta and IL-6 in infected bone were elevated in the early post-infection period and then decreased. In contrast, TNF-alpha levels remained elevated 3 to 28 days post-infection, while IL-4 levels were elevated late in the course of infection. The histopathology of infected bone showed predominant infiltration of inflammatory cells and bone resorption 3 to 7 days after infection, and bone resorption and adjacent areas of formation 14 to 28 days after infection. These results suggest that the elevated IL-1 beta and IL-6 levels induced by infection may be related to bone damage mainly in the early phase of infection, and that TNF-alpha and IL-4 may at least in part be associated with histopathological changes, including both bone resorption and formation in the later phase of this osteomyelitis model.     

Effects of acidic pH on the formation of vinblastine-induced paracrystals in Chinese hamster ovary cells

Summary— The pH-related change in morphology of vinblastine (VLB)-induced paracrystals formed in Chinese hamster ovary (CHO) cells was examined immunohistochemically in order to determine both the mechanism of tubulin crystallization and the influence of acidic pHs on cytoskeletal microtubules. Lowering the extracellular pH (pHe) rapidly reduced the intracellular pH (pHi) in CHO cells. Lowering the pHi to near the neutral range significantly accelerated the growth of VLB-induced paracrystals, compared to that of paracrystals formed at a physiological pHe. However, further cytoplasmic acidification caused by the addition of sodium azide into the culture medium induced the disappearance of typical paracrystals and the appearance of a highly organized meshwork of tubulin appearing as short, thick filaments at the light microscopic level. Treatments using different concentrations of VLB at different pHe's showed that low pHi's (6.7 and 6.3) suppressed paracrystal-formation at lower concentrations of VLB (5×10^?6 M and 10^?5 M). At higher concentrations of VLB (5×10^?5 M and 10^?4 M), only short filaments were formed at pHi 6. 3. Electron microscopy revealed that the filaments had a ladder-like structure probably consisting of a stacked series of fused rings. This indicates that paracrystals may be modified by extremely low pH. These results show that paracrystals are unstable in living cells and that their formation is regulated by environmental pH.     

Requirement for the MAP kinase kinase/MAP kinase cascade in Xenopus oocyte maturation.

MAP kinase kinase (MAPKK) has been identified as a protein factor that can induce phosphorylation and activation of inactive MAP kinase in vitro. In this study, we produced an anti-Xenopus MAPKK antibody that can specifically inhibit Xenopus MAPKK activity in vitro. Microinjection of this antibody into immature oocytes prevented progesterone-induced MAP kinase activation. Moreover, progesterone-induced histone H1 kinase activation and germinal vesicle breakdown (GVBD) were inhibited in the oocytes injected previously with this antibody. Furthermore, when a bacterially expressed Mos was introduced into immature oocytes, Mos-induced MAP kinase activation and GVBD were blocked in the oocytes injected with the anti-MAPKK antibody. These results show that MAPKK is responsible for the activation of MAP kinase in vivo and that the MAPKK/MAP kinase cascade plays a pivotal role in the MPF activation during the oocyte maturation process.     

Temporal Profiles of Nerve Growth Factor β-Subunit Level in Rat Brain Regions After Transient Ischemia

To determine the role of nerve growth factor (NGF) in ischemic brain damage, we measured the temporal and regional changes in the level of NGF in the hippocampal subfields, the cerebral cortex, the striatum, and the septum at 1, 2, 7, and 30 days after transient forebrain ischemia using a highly sensitive sandwich-type enzyme immunoassay system for the beta-subunit of mouse 7S NGF (beta-NGF). We also analyzed glial fibrillary acidic protein immunoreactivity in the hippocampus to ascertain the contribution of reactive astrocytes to NGF production after an ischemic insult. In the CA1 subfield of the hippocampus, the level of beta-NGF decreased slightly 2 days after ischemia (not significant), at which time CA1 pyramidal cell loss began to occur, and increased by 40% 30 days after ischemia (p less than 0.05). A marked increase in glial fibrillary acidic protein-positive astrocytes in the CA1 subfield 2-30 days after ischemia suggests that the reactive astrocytes participated in a gradual increase in the level of beta-NGF after recirculation. The level of beta-NGF in the dentate gyrus decreased transiently 2 days (p less than 0.05) and 7 days (p less than 0.01) after ischemia, followed by recovery to the level of control animals 30 days after ischemia. The level of beta-NGF in the septum gradually decreased 7 days (-27%, p less than 0.05) and 30 days (-43%, p less than 0.01) after ischemia. The levels of beta-NGF in the cerebral cortex and striatum remained unaltered throughout the observation period.(ABSTRACT TRUNCATED AT 250 WORDS)