Fc gamma RIII-mediated production of TNF-alpha induces immune complex alveolitis independently of CXC chemokine generation.

We recently demonstrated a codominant role of C5aR and FcgammaRIII in the initiation of IgG immune complex-mediated inflammation in mice. In this study, we investigated the relative contribution of FcgammaRIII in the generation of several cytokines during experimental hypersensitivity pneumonitis/alveolitis in vivo. Induction of immune complex-alveolitis in C57BL/6 mice resulted in strong accumulation of neutrophils into the lung and enhanced chemotactic activity within bronchoalveolar lavage fluid accompanied by an increased production of the proinflammatory cytokines TNF-alpha and IL-1beta as well as the ELR-CXC chemokines macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (KC). FcgammaRIII-deficient C57BL/6 mice (FcgammaRIII(-/-)) showed a marked reduction of the inflammatory response due to decreased production of TNF-alpha, IL-1beta, and MIP-2. Results obtained in C57BL/6 mice either lacking the TNF-alpha class I receptor (TNF-alphaRI(-/-)) or treated with neutralizing anti-TNF-alpha mAb demonstrated an essential contribution of TNF-alpha for mediating IL-1beta release, neutrophil influx, and hemorrhage. Surprisingly, MIP-2 and KC chemokine levels remained largely unaffected in TNF-alphaRI(-/-) mice or after functional inhibition of TNF-alpha. These data suggest that in immune complex alveolitis, the activation of FcgammaRIII may induce divergent downstream effector pathways with TNF-alpha acting independently of CXC chemokines to trigger the inflammatory response in C57BL/6 mice.

Endothelial-derived superoxide anions in pig coronary arteries: evidence from lucigenin chemiluminescence and histochemical techniques.

1. The generation of superoxide anions (O2-) by intact pig coronary artery rings was measured using a lucigenin-enhanced chemiluminescence technique and a histochemical technique with Nitroblue Tetrazolium (NBT) staining. 2. Isolated arteries with intact endothelium generated O2- at a rate of 9.0 +/- 0.8 pmol min-1 (mg dry weight)-1; this rate was diminished by about 24% when the endothelium was removed. The NBT staining of arterial ring preparations showed formazan precipitation mainly in the intima. Arterial rings were pretreated with diethylthiocarbamate in order to inhibit Cu-Zn superoxide dismutase (SOD) activity which increased the O2- generation by 184 +/- 55% (n = 10; P < 0.01). Stimulation of protein kinase C with phorbol 12-myristate 13-acetate (5 microM) enhanced endothelium-dependent O2- generation by 136 +/- 20% (n = 19; P < 0.01). Neither stimulation with bradykinin or substance P, nor inhibition with NG-nitro-L-arginine methyl ester of endothelial nitric oxide synthase had a significant effect on O2- generation. In contrast, the inhibition of flavoproteins with diphenyliodonium decreased concentration-dependent O2- generation (IC50, 1.85 +/- 5.33 microM). Inhibition of tetrahydrobiopterin synthesis with 2,4-diamino-6-hydroxy-pyrimidine resulted in a reduced generation of O2- by about 55%. 3. The addition of 100 microM NADH and 100 microM NADPH resulted in an excessive generation of O2- at a rate of 0.68 +/- 0.03 and 0.26 +/- 0.01 nmol O2- min-1 (mg protein)-1, respectively, in the membrane fraction, but not in the cytosolic fraction, of homogenates obtained from arteries. 4. The results suggest that intact coronary arteries do generate O2- under basal conditions and that the endothelial layer significantly contributes to this phenomenon. This generation of O2- is greatly influenced by intrinsic SOD activity. It is suggested that basal vascular O2- generation is mainly due to membrane-bound NAD(P)H oxidase activity and/or tetrahydrobiopterin-dependent processes.

Flavin enzymes, mitochondrial radicals and reduced glutathione in daily rhythmic dependency.

The diurnal rhythms of the microsomal flavoprotein NADPH-cytochrome c reductase activity, of diaphorase and of succinic dehydrogenase are presented. Minimum levels are ascertained at 09(00), maximum levels at 21(00). The concentration of mitochondrial radicals as a function of the time of day is also demonstrated. Here too the minimum is at 09(00) and the maximum between 15(00) and 21(00). On the other hand, GSH levels are found to be high between 09(00) and 12(00) and low in the evening. Thus a causative relationship between the concentration of cellular radicals, which originate in flavin enzymes, and the concentration of the tripeptide glutathione is assumed.

Effects of the scheduling of meal-feeding at different phases of the circadian system in rats.

This study was designed to determine whether or not a number of diverse rhythmic variables in the rat could be synchronized to meal timing. This was tested by restricting the availability of food; once during each 24-hour period an unrestricted quantity of food was made available for a 4-hour period to four different groups at different phases of the light-dark cycle, and the rhythms of the variables studied in the different groups were compared. Liver glycogen and serum glucose did synchronize to or were strongly influenced by feeding schedules; corticosterone and the several enzymes measured seemed to reflect an interaction of both the restricted feeding schedule and the light-dark cycle. The mitotic index in the corneal epithelium in all groups remained remarkably synchronized to the light-dark cycle and was altered only minimally by restricted meal timing. All groups on restricted feeding schedules gained less weight than the group fed ad libitum and maintained on a light-dark cycle. These studies caution against assuming that all body functions react in the same manner to different synchronizers; and they emphasize that one must not generalize about the synchronizing effect of meal-timing or even the light-dark cycle.

The manipulation of circadian rhythms.

In a study on male Wistar (TNO) rats kept under phase-shifted light-dark (12:12) conditions, it was demonstrated that the circadian rhythms of the following functions were effectively synchronized to the new lighting regimen: (a) body temperature, gross motor activity (after 1 week of acclimatization); (b) stomach weight, liver weight, liver glycogen, liver protein and acid phosphatase activity; serum corticosterone, glucose, total protein and inorganic phosphatase (after 4-6 weeks; as concluded from the data pooled of a sex of identical experiments); (c) the in vivo toxicity of approximately LD50 amounts (i.p.) for antimycin A and of E 600. In the individual experiments the various normal functions appeared to be different with respect to sensitivity to light. The overall pooled results indicated that, except for glycogen, the 24-h means of the normal values in the stomach, liver and serum were slightly decreased when compared to the respective means obtained from standard LD (= light: 6:00-18:00)-conditioned control groups (P less than 0.001 for liver weight, liver protein, serum protein and inorganic phosphate). The experimental animals revealed an intermediate or persisting decrease of the growth rate. From the serial biological and toxicological studies performed in the course of 1 year, it is concluded that the synchronizing effect of an altered lighting regiment may be influenced by seasonal factors...