Postnatal development of cannabinoid receptor type 1 expression in rodent somatosensory cortex.

Endocannabinoids are powerful modulators of synaptic transmission that act on presynaptic cannabinoid receptors. Cannabinoid receptor type 1 (CB1) is the dominant receptor in the CNS, and is present in many brain regions, including sensory cortex. To investigate the potential role of CB1 receptors in cortical development, we examined the developmental expression of CB1 in rodent primary somatosensory (barrel) cortex, using immunohistochemistry with a CB1-specific antibody. We found that before postnatal day (P) 6, CB1 receptor staining was present exclusively in the cortical white matter, and that CB1 staining appeared in the gray matter between P6 and P20 in a specific laminar pattern. CB1 staining was confined to axons, and was most prominent in cortical layers 2/3, 5a, and 6. CB1 null (-/-) mice showed altered anatomical barrel maps in layer 4, with enlarged inter-barrel septa, but normal barrel size. These results indicate that CB1 receptors are present in early postnatal development and influence development of sensory maps.

Synthesis of carbapenam skeletons using a ruthenium-catalyzed cyclization.

Carbapenam is a very important skeleton of beta-lactam antibiotics, and it has a highly strained structure. When enynes 9 were treated with RuH(2)CO(PPh(3))(3) (10 mol %) in toluene upon heating, carbapenams 10 were obtained in good yields.

Participation of CD14 in the phagocytosis of smooth-type Salmonella typhimurium by the macrophage-like cell line, J774.1.

The role of CD14 in the phagocytosis and killing of microorganisms was investigated using macrophage-like cell lines, CD14-positive J774.1 cells and CD14-negative mutant J7.DEF.3 cells derived from J744.1 cells. The cells were infected with Salmonella typhimurium organisms of the smooth (S)-form LT2, mutant rough (R)-form TV148 or Staphylococcus aureus 248betaH. At 30 or 180 min incubation, the cells were washed and disrupted. Colony-forming units (CFUs) liberated from the disrupted cells were determined by quantitative cultivation, and the phagocytic index and killing rate were calculated. Both the phagocytic index and killing rate of J774.1 cells against LT2 organisms were greater than those of J7.DEF.3 cells. However, the index and rate of J774.1 cells against TV148 and 248betaH organisms were similar to those of the J7.DEF.3 cells. The phagocytosis of LT2 organisms by J774.1 cells was partially inhibited by S-form LPS (S-LPS) and anti-CD14 antibody, but not by R-chemotype LPS (R-LPS). These results suggest that CD14 participates in the phagocytosis of S-form Salmonella.

Non-specific resistance induced by a low-toxic lipid A analogue, DT-5461, in murine salmonellosis.

The ability of DT-5461, a chemically synthetic low-toxic lipid A analogue, to activate anti-Salmonella activity in C3H/HeN mice was examined. Previous intraperitoneal (i.p.) injection of DT-5461 (100 micrograms or more/mouse) significantly hindered the bacterial growth in the peritoneal cavities of the mice after the i.p. infection with Salmonella typhimurium LT2 strain. The effect was the maximum when DT-5461 was given 6 h before the challenge. The injection of DT-5461 6 h in advance could also confer protection against the infection. Bactericidal activity enhancement was also seen in mice that had been injected with a small amount of recombinant murine IFN-gamma (10(3) U per mouse) and non-effective dose (10 micrograms) of DT-5461 together 6 h before the challenge. Bactericidal effect enhancement was seen in mice that had been injected with IFN-gamma at 6 h and DT-5461 at 3 h before the challenge, while it could be hardly seen in mice injected with them in a reversed order. The i.p. injection of DT-5461 recruits the exudate cells into the peritoneal cavities, and the phagocytic and bactericidal abilities of the macrophages in the exudate cells are apparently elevated. The mechanisms of non-specific resistance enhancement induced by DT-5461 were discussed.

Sensitivity of bacteria to NaNO2 and to L-arginine-dependent system in murine macrophages.

Sensitivity of bacteria to NO2-/NO and to L-arginine-dependent system in murine macrophages was tested. The growth of all bacteria tested, Salmonella typhimurium, Pseudomonas aeruginosa and Staphylococcus epidermidis, was significantly inhibited by NaNO2 at the concentration of 5 to 10 mM. However, L-arginine-dependent effector mechanism of LPS-activated murine macrophages was ineffective in killing these bacteria. It seems that the concentration of NO2-/NO in phagolysosome in the activated macrophages is insufficient to kill the bacteria in situ. We concluded that L-arginine-dependent effector mechanism can hardly work on bacteria killing in activated macrophages.

Protective effect of recombinant tumor necrosis factor-alpha in murine salmonellosis.

The enhancement of resistance by i.p. administration of murine rTNF-alpha into mice against i.p. challenge with virulent Salmonella typhimurium was studied. Administration of TNF-alpha (5 x 10(4) U/mouse) into mice at 6 or 12 h before the challenge with S. typhimurium organisms enhanced the bactericidal capacity in the peritoneal cavities of the mice. The diminution of the infecting organism in the peritoneal cavities of the TNF-alpha-treated mice was not due to the systemic spread of the organism inasmuch as few organism were recovered from other areas such as the spleen and liver. The TNF-alpha treatment effected a slight increase of neutrophils in the peritoneal cavity, but did not effect an increase of macrophages, including Ia-bearing macrophages. The survival rate of mice infected with Salmonella was improved by the i.p. injection of TNF-alpha before infection. Co-administration of smaller doses of TNF-alpha (5 x 10(3) U) and murine rIFN-gamma (10(2) U) at 6 h before the challenge also effectively enhanced bactericidal activity and protectivity. The cooperative effect of TNF-alpha and IFN-gamma was only seen when these recombinant cytokines were injected together at the proper time before the challenge. Injection of rabbit anti-TNF-alpha serum abolished the effects of TNF-alpha and the cooperative effect of TNF-alpha and IFN-gamma. Furthermore, the serum could abolish the cooperative effect of IFN-gamma and LPS on bactericidal activity, suggesting participation of LPS-induced TNF-alpha in the cooperation.

Effect of murine recombinant interferon-gamma in the protection of mice against Salmonella.

The ability of recombinant murine (rMu) interferon (IFN)-gamma to activate anti-Salmonella-activity in normal mice and beige mutant (bg/bg) mice with Chediak-Higashi syndrome (CHS) was examined. Previous intraperitoneal (i.p.) injection of rMuIFN-gamma (10(4) U per mouse) significantly hindered the bacterial growth in the peritoneal cavities, spleens and livers of the mice after the i.p. infection with Salmonella enteritidis No. 11 strain. It was also effective on the beige mice that have phagocytic cells with a genetically impaired bactericidal function, suggesting that IFN-gamma activates the pathway irrelevant to the beige mutation. The effect was the maximum, when IFN-gamma was given 6 h before the challenge. The effect seemed to be due to the augmentation of bactericidal capacity rather than the prevention of systemic spread of bacteria. Recombinant human IFN-alphaA/D (10(2)-10(6) U per mouse), which produced effects identical to those of murine IFN-beta, did not show such a bactericidal effect. Bactericidal activity enhancement was also seen in mice that had been injected with a small amount of rMuIFN-gamma (10(2) U) and bacterial lipopolysaccharide (LPS) (10 ng) together at 6 h before the challenge, although the IFN-gamma or LPS alone at these doses produced very little if any effect. Bactericidal effect enhancement was seen in mice that had been injected with IFN-gamma at 6 h and LPS at 3 h before the challenge, while it could be hardly seen in mice injected with them in a reversed order.(ABSTRACT TRUNCATED AT 250 WORDS)

The adjuvant effect of a muramyl dipeptide (MDP) analog on temperature-sensitive Salmonella mutant vaccine.

Adjuvant effect of a synthetic muramyl dipeptide analog, MDP-Lys (L18), MurNAc-L-Ala-D-glu[Lys(CO-(CH2)16-CH3)-OH]NH2 on a live Salmonella enteritidis vaccine, a temperature-sensitive mutant Ts-O strain that was obtained from the virulent S. enteritidis No.11 strain and could not grow at 37 degrees C, but could multiply at 25 degrees C as fast as the parent strain, was examined. Although the Ts-O organisms were avirulent and could hardly multiply in vivo when the organisms were injected into C57BL/6 mice and its mutant beige strain, which has a malfunction of phagocytic cells, injection of these mice with Ts-O endowed them with some protective immunity against infection by the virulent No.11 strain. When MDP-Lys(L18) was injected with Ts-O vaccine, the protection and the bactericidal capacity in the peritoneal cavities and spleens of these mice were augmented. MDP-Lys(L18) was still effective when it was injected at 48 h before or after the inoculation of Ts-O vaccine. Its effect was also observed against infection by the virulent S. cholerae-suis Hokkaido strain, a strain that does not share common O-antigenic determinants with the S. enteritidis No.11 or Ts-O strain. In addition, the mice that were inoculated simultaneously with Ts-O organisms and MDP-Lys(L18) were examined 10 days later for their footpad delayed type hypersensitivity reactions against Ts-O antigen. Mice inoculated with MDP-Lys(L18) and Ts-O showed augmented footpad swelling in comparison with the controls. These findings indicate that MDP-Lys(L18) is capable of augmenting the cellular immunity by live vaccine.

Augmentation of protective and antibacterial activity induced by muramyl dipeptides in CBA/N defective mice with X-linked immunodeficiency for Salmonella enteritidis infection.

Stimulation of resistance induced by muramyl dipeptide (MDP) and its analog, N alpha-MDP-N epsilon-stearoyl-L-lysine [MDP-Lys(L18)], was examined in experimental salmonellosis in CBA/N defective mice with X-linked immunodeficiency to virulent Salmonella enteritidis no. 11. An injection of either MDP or MDP-Lys(L18) did not induce any effective protection, but repeated injections of MDP-Lys(L18) (100 micrograms per mouse per day for 3 days consecutively) to the mice before bacterial challenge gave some protection. Multiple injections with MDPs once a day for several days consecutively strongly increased bactericidal capacity in the peritoneal cavities and spleens of the mice. Moreover, previous injection of the MDPs could elevate the phagocytic function of the reticuloendothelial system in the defective mice. These results indicate that nonspecific resistance of CBA/N defective mice to salmonella infection can be improved by previous administrations of MDPs.

Effect of muramyl dipeptide analog on Salmonella enteritidis infection in beige mice with Chediak-Higashi syndrome.

Beige mutant (bg/bg) mice with Chediak-Higashi syndrome (CHS) were much more sensitive to virulent Salmonella enteritidis No. 11 strain than parental C57 BL/6 (+/+) or heterozygous (bg/+) mice, and they had weaker bactericidal activity against the organisms. Muramyl dipeptide (MDP) and N alpha-(N-acetyl-muramyl-L-alanyl-D-isoglutamyl)-N epsilon-stearoyl-L-lysine [MDP-Lys(L18)], a synthetic derivative of MDP, failed to confer any protection against the infection, but the MDPs showed some ability to stimulate the bactericidal activity in the peritoneal cavities and spleens of these mice. The bactericidal effect of MDP-Lys(L18) was dose-dependent, and the greatest effect was seen when it had been injected 24 hr before the infection. Multiple injections of MDP were much more beneficial than a single injection. Previous injection of N2,O2'-dibutyryl guanosine 3' : 5'-cyclic monophosphate (DB-cGMP) improved the impaired bactericidal capacity in beige mice, but the simultaneous injection of N6,O2-dibutyryl adenosine 3' : 5'-cyclic monophosphate (DB-cAMP) with DB-cGMP abolished the effect of DB-cGMP. The augmentation of bactericidal capacity by MDP-Lys(L18) was not affected by the injection of either DB-cGMP or DB-cAMP, suggesting that the effect of the MDPs was not related directly to cyclic nucleotide regulation in beige mice.