Multiple processes are involved in the uptake of chylomicron remnants by mouse peritoneal macrophages

The processes responsible for the uptake of chylomicron remnants by macrophages were investigated using freshly isolated cells from low density lipoprotein (LDL) receptor, very low density lipoprotein (VLDL) receptor and apolipoprotein E knockout mice. In peritoneal macrophages from normal mice, the metabolism of chylomicron remnants was inhibited 40% by anti-LDL receptor antibody and 60% by a high concentration of receptor-associated protein (RAP). Together they reduced the amount processed by 70%. Digestion of cell proteoglycans decreased remnant degradation by 20% while the addition of acetyl-LDL had no effect. When LDL receptors were absent, the absolute rates of metabolism were less than that of normal cells and were not inhibited by the anti-LDL receptor antibody; the rates, however, were reduced to less than half by RAP. These suggest that the LDL receptor-related protein (LRP) or another LDL receptor family member(s) contributes to chylomicron remnant uptake and becomes the major mechanism of uptake when LDL receptors are absent. In contrast, the VLDL receptor was not involved as its absence did not affect chylomicron remnant metabolism. Similarly, the absence of apoE production did not affect the amount of remnant uptake; however, the proportion that was sensitive to RAP was eliminated. The level of LRP expression was not altered in these cells whereas there was a decrease in LDL receptors. This suggests that the apoE content of chylomicron remnants is sufficient for its recognition by LDL receptors but additional apoE is required for its uptake by the LRP and that there is an up-regulation of a non-LDL receptor family mechanism in apoE deficiency. Together these studies suggest that even in the absence of LDL receptors or apoE secretion, chylomicron remnants could contribute to lipid accumulation in the artery wall during atherogenesis.     

Immortalization and characterization of bovine peritoneal macrophages transfected with SV40 plasmid DNA

A transformed bovine peritoneal macrophage cell line was developed and characterized. Primary peritoneal macrophages were transformed by calcium-phosphate transfection with SV40 plasmid DNA. The transformed cell line retained the morphology of resident peritoneal macrophages as determined by light microscopy and histochemical analysis showed non-specific esterase activity. In addition, immunohistochemical staining of transformed peritoneal macrophages for lysozyme activity was positive. Transformed cells phagocytized Staphylococcus aureus, lysed chicken red blood cell (RBC) targets with and without opsonization and produced hydrogen peroxide radicals and interleukin-6 upon stimulation with opsonized zymosan and lipopolysaccharide, respectively. Transformed cells were also able to ingest and kill Mycobacterium paratuberculosis, an acid-fast bacillus. These results suggest that this cell line should be useful to study interactions between the bovine and intracellular pathogens.     

Anandamide and other N-acylethanolamines in mouse peritoneal macrophages

Exposure to stressful events and elevated level of stress hormones are associated with impaired spatial memory and neuronal damage in the hippocampus. These neurons are considered to be maintained by neurotrophins such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) and trk family of neurotrophin receptors. Male Wistar rats (6 weeks old) were exposed to immobilization stress for 8 h and their brains were processed for in situ hybridization histochemistry. Exposure to long-lasting immobilization stress reduced mRNA levels for neurotrophins and their high affinity receptors in the brain, especially in the hippocampus. Our results provide, some new information that may be relevant to the pathogenesis of stress-induced disturbances of memory and learning.     

Nitric oxide production in mouse peritoneal macrophages enhanced with glycyrrhizin

The enhancement of nitric oxide (NO) production in glycyrrhizin (GL)-induced macrophages (M phi) in response to lipopolysaccharide (LPS) was investigated. No production in GL-induced macrophage culture supernatants was stimulated in response to LPS (10 micrograms/ml) for 24- or 48- h cultures, and these levels were compared three times with the levels in saline-induced peritoneal exudate cell cultures. Furthermore, M phi induced with proteose peptone (PP) containing GL could generate greater NO production than M phi induced with PP alone. However, no stimulation of NO production was observed by addition of GL in the cultures of M phi induced with thioglycollate or Bacillus Calmette Guerin. Moreover, GL-induced M phi showed cytostasis against such tumor target cells as L 1210 and P 388 lymphoma cell lines. These observations indicate that GL can activate the M phi in vivo system and stimulate NO production in response to LPS.     

Role of kinases and G-proteins on arachidonate release induced by zymosan in mouse peritoneal macrophages

Each of the 15 Health Boards in Scotland maintains a computer file of its residents who are registered with a general practitioner; this is known as the Community Health Index or CHI. The CHI allows a variety of demographic data and indicators of health to be analysed on either a geographic or general practice base, or both simultaneously. The considerable potential of the CHI as a public health tool may be of interest to health authorities outside Scotland which are developing wider uses for their own family practitioner registers.     

Induction of DNA synthesis in rat peritoneal macrophages in culture by a pleural inflammatory exudate

Peritoneal macrophages in culture are blocked in the G0 phase of the cell cycle, but retain many of their functional characteristics such as phagocytic ability. Peritoneal macrophages have been thought to be a terminal cell type. It has been investigated whether such properties could be modified by a substance released in acute inflammatory exudates. For this purpose a pleural exudate obtained from rats injected with dextran (40,000) 4 hours before, was centrifuged to eliminate cells, sterilized by filtration on Millipore filter 0.22 mum and diluted 50% with 199 medium culture. This medium was used to treat normal and activated peritoneal macrophages in culture. The effects were observed 24, 48, 72, 96 hours after the beginning of treatment. An enhancement of spreading and capacity of phagocytosis was observed 24 hours after the beginning of treatment. After 48 hours, the number of cells incorporating tritiated thymidine increased and became highest 4 days later. These phenomena were also obtained with pleural exudate of inbred rats (Lewis, Wag) treating macrophages of the same strain and with rat pleural exudate treating mouse macrophages. No effects were observed with dextran alone. The chemical nature of the stimulatory factor remains to be elucidated.     

Interferon gamma upregulates its own gene expression in mouse peritoneal macrophages

Interferon gamma (IFN-gamma) exerts a variety of immunoregulatory effects on several cell targets. It is generally assumed that IFN-gamma is specifically produced by T and large granular lymphocytes. In this study, we show that IFN-gamma is constitutively expressed in resting mouse peritoneal macrophages (PM). Treatment of PM with cycloheximide results in a significant accumulation of IFN-gamma mRNA, suggesting that a short-lived IFN-gamma mRNA accumulates when protein synthesis is inhibited. Moreover, treatment of PM with IFN-gamma also results in a clear-cut accumulation of this mRNA. This effect is not observed in murine lymphocytes from mesenteric lymph nodes (which instead produce IFN-gamma after phytohemagglutinin treatment) and in mouse cell lines. The treatment of PM with IFN-gamma also results in secretion of IFN-gamma after 24-48 h. The upregulation of IFN-gamma expression is also found in PM from anti-asialo GM1-treated nude mice. We suggest that the ability of PM to produce this IFN-gamma is indicative of an autocrine mechanism. The macrophage IFN-gamma may play a role in the regulation of cell differentiation and immune response.     

Selective deimination of vimentin in calcium ionophore-induced apoptosis of mouse peritoneal macrophages

Amino acid infusions during general anesthesia induce thermogenesis and prevent postoperative hypothermia. The effects of increased heat production during anesthesia on postoperative nitrogen balance have not been examined. Therefore, we studied the effect of perioperative amino acid infusions on postoperative nitrogen excretion in 24 patients scheduled for hysterectomy. Seven volunteers not subjected to anesthesia or surgery were used as awake controls. During isoflurane anesthesia, 8 patients received acetated Ringer's solution, and 16 patients received an IV amino acid mixture, 240 kJ/h, before and during anesthesia. Rectal temperature and energy expenditure were measured. The urinary nitrogen content was calculated from urea, creatinine, and urate the day before surgery and for 4 days postoperatively. Diets were recorded. In anesthetized control patients, postoperative nitrogen excretion was less than preoperative levels. Those patients also experienced the largest decrease in core body temperature during anesthesia (1.7+/-0.1 degrees C). All had postoperative shivering. In the amino acid-treated patients, the temperature decrease during anesthesia was less pronounced (1.0+/-0.1 degrees C; P < 0.001) and postoperative shivering disappeared. In addition, the nitrogen excretion was unchanged postoperatively, perhaps indicating an increase in protein turnover known to generate heat. In conclusion, the increase in heat production induced by amino acids reduced hypothermia, abolished shivering, and attenuated/normalized the postoperative nitrogen saving that occurred in patients who did not receive amino acids. IMPLICATIONS: We compared nitrogen excretion before and after surgery in patients who received a saline or amino acid infusion during isoflurane anesthesia. The increase in heat production induced by amino acids reduced hypothermia, abolished shivering, and attenuated/normalized the postoperative nitrogen saving that occurred in patients who did not receive amino acids.     

Characterization of VIP receptor-effector system antagonists in rat and mouse peritoneal macrophages

In the present study we show that the synthetic peptides [4-Cl-D-Phe6,Leu17]VIP and the growth hormone releasing factor (GRF) analog [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 inhibit in a competitive manner the specific [125I]VIP binding to both rat and mouse peritoneal macrophages. In rat peritoneal macrophages, the order of potency of the different peptides, as expressed by the IC50 values was: VIP (IC50 = 1.90 +/- 0.16 nM) > [4-Cl-D-Phe6,Leu17]VIP (IC50 = 125.8 +/- 13.2 nM) > [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 (IC50 = 354.8 +/- 21.2 nM). In mouse peritoneal macrophages a similar pattern of potency was observed: VIP (IC50 = 1.58 +/- 0.12 nM) > [4-Cl-D-Phe6,Leu17]VIP (IC50 = 110.8 +/- 10.7 nM) > [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 (IC50 = 251 +/- 19.2 nM). The behavior as VIP receptor antagonists of both [4-Cl-D-Phe6,Leu17]VIP and [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 in rat and mouse peritoneal macrophages was confirmed by: (a) the shift to the right of VIP dose-stimulated cyclic AMP production curves in the presence of the two antagonists; (b) the agreement between the order of efficacy of the two peptides in competition experiments with the corresponding inhibition of cyclic AMP production; (c) the inefficiency of the two antagonists on the stimulation of cyclic AMP production by the beta-adrenoceptor agonist isoproterenol, which indicates the specificity of the interaction; (d) the synergic effect of VIP on isoproterenol-stimulated cyclic AMP production was completely abolished by [4-Cl-D-Phe6,Leu17]VIP or [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2, suggesting that both antagonists acted via specific VIP receptors. Moreover, propranolol, a beta-adrenoceptor antagonist, did not affect the VIP-stimulated cyclic AMP production and the antagonist role of [4-Cl-D-Phe6,Leu17]VIP or [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2; (e) in cross-linking experiments, the intensity of the labeling of the [125I]VIP/receptor complexes was significantly lower with the antagonists than in the control experimental situation in both mouse and rat peritoneal macrophage membranes.     

Convenient plasmid for extrachromosomal DNA recombination in mouse cells

We constructed a convenient plasmid for DNA recombination assay. The plasmid, pMR1, contains a double prokaryotic terminator to decrease the background and two unique restriction enzyme sites on both sides of the double terminator to allow for easy construction. The assay is capable of selecting the bacterial cells containing recombined plasmid DNA on a selective plate containing ampicillin and chloramphenicol. We adapted pMR1 for V(D)J recombination and homologous recombination and detected both types of recombination in murine PreB cell line. As pMR1 has the double terminator, background on the selective plate decreases effectively and we select only the recombined clones. We consider the vector, pMR1, to be convenient for the analysis of homologous and non-homologous recombinations.     

Effect of cytochalasin B on the adhesion of mouse peritoneal macrophages

The adhesion of normal mouse macrophages to glass surfaces was reduced by nontoxic levels (1-50 mug/ml) of cytochalasin B in combination with a centrifugal force (1,000-8,000 g). Macrophages nonspecifically activated by Corynebacterium acnes were also detached by this treatment, but less effectively. The effects of cytochalasin B treatment on these cells were shown to be reversible. After detachment, the cells reattached to glass, appeared morphologically normal, and behaved like untreated cells as judged by adhesion, acid phosphatase levels, and phagocytosis. The effect of cytochalasin B on several parameters of phagocytosis by normal macrophages was also examined. The results demonstrate that cytochalasin B can be used to detach macrophages from surfaces and suggest a functional relationship between phagocytosis and macrophage adhesion to surfaces. Furthermore, the effect of cytochalasin B on adhesion of phagocytic cells provides a probe for further investigation of the adhesion of cells to surfaces.     

The in vitro effect of hydrocortisone on endocytosis in cultured mouse peritoneal macrophages

Three juvenile patients with cerebellar astrocytomas which have seeded the spinal subarachnoid space are presented. Histologic verification of the similarity between the posterior fossa tumor and its spinal implant was obtained in two of the three patients. The cerebellar tumors in all cases have been benign (grade I),and the behavior, other than their seeding has also been indolent. Review of pertinent literature discloses no similar experience with cerebellar astrocytomas. Aggressive therapy is advocated for the rare patient with subarachnoid seeding from this benign lesion.     

Chromosome-mediated resistance of Yersinia enterocolitica serotype O9 to intracellular killing by mouse peritoneal macrophages

The survival of Yersinia enterocolitica serotype O9 within mouse peritoneal macrophages was investigated. To evaluate the role of the virulence plasmid in the resistance to intracellular killing, an isogenic pair of virulent (plasmid-bearing) and avirulent (plasmid-less) O9 strains was used. The virulent strain was able to express plasmid-encoded outer membrane proteins and to colonize the Peyer's patches of orally infected mice. When mice were infected intraperitoneally, both strains were recovered at similar rates and over the same time from the peritoneal cavity. When in vitro assays were performed, both strains showed similar resistance to intracellular killing by monolayers of resident and inflammatory peritoneal macrophages. Previous opsonization of bacteria did not modify their survival within macrophage monolayers. We concluded that serotype O9 strains display a chromosome-mediated resistance to intracellular killing by mouse peritoneal macrophages. Moreover, macrophage resistance does not seem to be of importance for virulence of serotype O9 strains in mice.     

Regulation of prostaglandin synthesis and of the selective release of lysosomal hydrolases by mouse peritoneal macrophages

Macrophages isolated from the peritoneal cavity of untreated mice and maintained in tissue culture synthesize and release prostaglandins when challenged with zymosan. These cells also selectively release lysosomal acid hydrolases under the same conditions. The major prostaglandins released into the media are found to be prostaglandins E1, E2 and 6-oxoprostaglandin F1a, whereas prostaglandin F2a is not detected. Macrophages isolated from mice that have received an intraperitoneal injection of thioglycollate broth are far less responsive to zymosan challenge. These cells require 300 microgram of zymosan to synthesize and release one-third the amount of prostaglandins released from non-stimulated macrophages exposed to 50 microgram of zymosan. In addition, thioglycollate-stimulated macrophages release less than 10% of their lysosomal acid hydrolases when exposed to 300 microgram of zymosan whereas non-stimulated cells release approximately 50% of these enzymes after treatment with 50 microgram of zymosan. The zymosan-stimulated synthesis and release of prostaglandins are completely inhibited by indomethacin, whereas the increased selective release of lysosomal acid hydrolases is not affected. Macrophages, unlike fibroblasts, do not synthesize and release prostaglandins when exposed to serum or to bradykinin.     

The effect of platelet-activating factor and its synthetic analogs on Ia-antigen expression by mouse peritoneal macrophages

Visual evoked potentials (VEPs) of the pattern shift reversal type were determined in a representative group of 57 prisoners of war (POWs) released in 1992 from detention camps in former Yugoslavia. The parameters were correlated with the conditions in four camps (1-4). All subjects were male, with a mean age of 34.75 years (SD +/- 8.92), average length of imprisonment 192.7 days (SD +/- 77.6), mean loss of body mass during imprisonment 19.32% (SD +/- 9.54), and the average number of reported blows to the head and neck was 25.7 (SD +/- 20.3). VEPs were determined on average 290.5 days after the last craniocerebral trauma caused by blows to the head and neck (SD +/- 152.0) i.e., on average 218.5 days after release from the camp (SD +/- 164.3). Although all the 57 POWs reported being maltreated to a certain extent, 14 reported being subjected to particularly brutal forms of torture, 5 had been held in solitary confinement and 25 had lost consciousness at least once. Solitary confinement and loss of consciousness had the most significant effect on VEPs, and the altered VEP parameters correlated significantly with the craniocerebral trauma experienced, loss of body mass and the length of time since the last craniocerebral trauma until examination, and from release until examination. However, the length of imprisonment and treatment in the camps did not have a significant effect on VEP parameters. The study confirmed that under such conditions the age of the subject is a risk factor. The results of this study also confirmed that prisoners in one camp had been subjected to the worst maltreatment.     

Destruction of allogeneic tumour cells by peritoneal macrophages

The population of peritoneal macrophages from mice immunised with allogeneic tumor cells contains two types of cytotoxic cell. One lyses the specific target cell, the other initiates activation of macrophages and hence leads to inhibition of growth of the specific target cell. The yield of lytic effector macrophages was found to depend on the route and the nature of the cell used for immunisation and the condition of the mice. The yield correlated with the yield of complement-dependent cytotoxic antibodies. In contrast, production of specific "activator" macrophages did not depend critically on these factors. The results underline the difference between the two types of cell and suggest that they are produced independently of one another.     

Release of superoxide anion from activated mouse peritoneal macrophages during Mycobacterium tuberculosis infection

Mouse peritoneal macrophage monolayers infected with M. tuberculosis were cultured in RPMI up to 7 days. Release of superoxide was assayed on different days in presence or absence of Phorbol myristate acetate (PMA), a known stimulator of NADPH oxidase which is involved superoxide production. Basal level of superoxide release was significantly higher in M. tuberculosis infected peritoneal mouse macrophages (P < 0.01) as compared to normal mouse macrophages. When normal and tuberculoid macrophage cultures were stimulated with PMA, increased superoxide anion release was observed in both the cultures but the increase of superoxide was significantly higher in normal macrophages as compared to tuberculoid stimulated macrophages. Superoxide release was maximum in 4 day old cultured macrophages and gradually it declined in older cultures by day 7, both in vitro and in vivo. A defective macrophage function in killing of M. tuberculosis bacilli was observed after 4 days of in vitro and in vivo cultures.     

Effect of esculentoside H on release of tumor necrosis factor from mouse peritoneal macrophages

Effect of esculentoside H (EH) on release of tumor necrosis factor (TNF) from murine peritoneal macrophage (Mphi) in vitro was studied. The results showed that EH (12.5-200 micrograms.ml-1) induced the thioglycolate-broth elicited peritoneal Mphi to release TNF into supernatants in a dose-dependent manner, and higher levels of TNF activity were detected in the supernatants from EH-stimulated calcimycin-primed M? culture. EH-induced TNF release had a different type of kinetics compared with that of lipopolysaccharides (LPS). LPS-induced release of TNF increased rapidly until 6 h after LPS stimulation, then declined gradually, while EH-induced TNF release increased gradually after EH stimulation and reached its peak at approximately 24 h later. These results suggested that the anti-tumor mechanisms of Phytolaccaceae may be related to the capacity of EH for TNF release.