Genes for chemokines MuMig and Crg-2 are induced in protozoan and viral infections in response to IFN-gamma with patterns of tissue expression that suggest nonredundant roles in vivo

MuMig and Crg-2 are IFN-inducible murine chemokines whose human homologues, HuMig and IP-10, respectively, share activity in vitro as T cell chemoattractants. We analyzed the expression of the genes Mumig, crg-2, and IFN-gamma during experimental infections with Plasmodium yoelii, Toxoplasma gondii, and vaccinia virus. Mumig, crg-2, and IFN-gamma were induced in multiple organs. During the acute phase of each infection as well as after i.p. injection of rIFN-gamma, levels of Mumig mRNA in the liver were as high or higher than levels in any of the other organs. In contrast, the organs showing the highest expression of crg-2 and IFN-gamma varied among the experimental models, with induction of these latter two genes colocalizing. Differences in relative levels of expression of Mumig and crg-2 in liver and spleen were not demonstrably due to expression of the genes in different cell types within these organs. We showed that both Mumig and crg-2 are induced in the liver in hepatocytes and in the spleen in CD11b+ cells. IFN-gamma was necessary for induction of Mumig during infections with T. gondii or vaccinia virus. In contrast, induction of crg-2 was not completely dependent on IFN-gamma. These data demonstrate that despite the overlap in activities within chemokine subsets, chemokine genes show differences in their patterns of expression and in their responses to inducers that suggest nonredundant roles in vivo. Moreover, the pattern of induction of crg-2 is consistent with Crg-2 acting primarily locally, while the pattern for Mumig induction suggests that MuMig may have a systemic role during infection.     

In vivo cardiotonic activity of pyridylmethylene-2-indolinones

Carboplatin hypersensitivity has rarely been reported in patients receiving repeated cycles of therapy, but has not been reported in transplant settings. We report a case of carboplatin hypersensitivity during conditioning for autologous PBSC transplantation. The patient suddenly developed chest tightness, hemoptysis, hypoxia and hypotension, resulting in a transient myocardial ischemia. The pathophysiologic mechanism for the event seemed to be non-immune-mediated direct histamine release given the lack of prior exposure to platinum. Contrary to advice not to continue further treatment with carboplatin by some authors, we successfully desensitized the patient and subsequently gave more carboplatin as a part of conditioning. Awareness of carboplatin as one of the causes of hypersensitivity may help avoid further problems either by substitution or desensitization, along with premedications.     

In vivo antispasmodic activity of Sulcotidil

An increase of oxygen concentration in a gaseous mixture passed through cell suspensions of Chlorella pyrenoidosa (from 21 to 100%) stimulates the production of extra-cellular organic substances and changes their composition. The cells accumulate in the medium glycolic acid rather than a variety of other substances. The accumulation of glycolic acid depends on the growth stage of the cells. The relation of production of extracellular organic substances by the algal cells to the ratio between the carboxylase and oxgenase activities of ribulose diphosphate carboxylase is discussed.     

In vivo and in vitro antiinflammatory activity of saikosaponins

Buddlejasaponin I and saikosaponin 1 and 2, biologically active compounds from Scrophularia scorodonia and Bupleurum rigidum respectively, exert potent in vivo antiinflammatory effects on mouse ear edema induced by phorbol myristate acetate (PMA). The effects of these compounds on swelling and other inflammatory parameters are described. In screening for in vitro effects of saikosaponins on cellular systems generating cyclooxygenase (COX) and lipoxygenase (LOX) metabolites, we observed that most saikosaponins showed a significant effect. The action is more marked on LOX metabolite LTC4. Our data support the inhibition of arachidonic acid metabolism as one of the biochemical mechanisms that might be the rationale for the putative antiphlogistic activity of these saikosaponins.     

Ribonucleoside and 2'-deoxyribonucleoside 5'-phosphonates: synthesis and antiviral activity

Groups of 5'-phosphonates of natural 2'-deoxynucleosides and ribonucleosides were synthesized by condensation of 3'-acetylated 2'-deoxynucleosides or 2',3'-substituted (2',3'-O-isopropylidene, 2',3'-O-methoxymethylene, or 2',3'-O-ethoxymethylene) ribonucleosides. As condensing agents, either N,N'-dicyclohexylcarbodiimide or 2,4,6-triisopropylbenzenesulphonyl chloride were used. Nucleoside 5'-ethoxycarbonyl-phosphonates were converted into corresponding nucleoside 5'-aminocarbonylphosphonates by the action of ammonia in methanol. All compounds were tested for inhibition of several viruses, including human herpes simplex virus type 2 and cytomegalovirus, but showed no activity. A few compounds insignificantly inhibited human immunodeficiency virus type 1 reproduction. Thymidine 5'-hydrogenphosphonate neutralized the anti HIV action of 3'-azido-3'-deoxythymidine, thus indirectly showing that it could be partly hydrolyzed in cell culture to corresponding thymidine.     

In vivo micropuncture PCO2 measurements

An electrode is presented which permits in vivo PCO2 measurements using a micropuncture technique. The tip diameter of the electrode is only a few microns, the tip is specially designed for measuring PCO2 in small tissue compartements.     

In vitro and in vivo inhibition of murine gamma herpesvirus 68 replication by selected antiviral agents

We have evaluated the susceptibility of the murine gamma herpesvirus 68 (MHV-68) to a variety of antiviral agents. The acyclic nucleoside phosphonate analogs cidofovir [(S)-1-(3-hydroxy-2-phosphonylmethoxypropyl) cytosine], (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl)adenine (HPMPA), and adefovir [9-(2-phosphonylmethoxyethyl)adenine] efficiently inhibited the replication of the virus in Vero cells (50% effective concentrations [EC50s], 0.008, 0.06, and 2.2 microg/ml, respectively). Acyclovir, ganciclovir, and brivudin [(E)-5-(2-bromovinyl)-2'-deoxyuridine] had equipotent activities (EC50s, 1.5 to 8 microg/ml), whereas foscarnet and penciclovir were less effective (EC50s, 23 and > or =30 microg/ml, respectively). The novel N-7-substituted nucleoside analog S2242 [7-(1,3-dihydroxy-2-propoxymethyl)purine] inhibited MHV-68 replication by 50% at 0.2 microg/ml. The susceptibilities of MHV-68 and Epstein-Barr virus (EBV) to cidofovir, HPMPA, adefovir, and acyclovir were found to be comparable. However, for penciclovir, ganciclovir, brivudin, and S2242, major differences in the sensitivity of MHV-68 and EBV were observed, suggesting that MHV-68 is not always an optimal surrogate for the study of antiviral strategies for EBV. When evaluated with a model for lethal MHV-68 infections in mice with severe combined immunodeficiency, cidofovir proved to be very efficient in protecting against virus-induced mortality (100% survival at 50 days postinfection), whereas acyclovir, brivudin, and adefovir had little or no effect.     

Interferon-inducing and antiviral activity of levamisole

It was shown that levamisol administered orally to mice induced production of interferon with its maximum level in 4-6 hours and prolonged subsequent circulation in the host (the observation period of 5 days). Antiviral activity of levamisol in experimental forest-spring encephalitis was shown (protection of 35-40 per cent). When levamisol were used in combination with polyguacyl, an additive effect was recorded.     

In vivo and in vitro analysis of baculovirus ie-2 mutants

Upon transient expression in cell culture, the ie-2 gene of Autographa californica nuclear polyhedrosis virus (AcMNPV) displays three functions: trans activation of viral promoters, direct or indirect stimulation of virus origin-specific DNA replication, and arrest of the cell cycle. The ability of IE2 to trans stimulate DNA replication and coupled late gene expression is observed in a cell line derived from Spodoptera frugiperda but not in a cell line derived from Trichoplusia ni. This finding suggested that IE-2 may exert cell line-specific or host-specific effects. To examine the role of ie-2 in the context of infection and its possible influence on the host range, we constructed recombinants of AcMNPV containing deletions of different functional regions within ie-2 and characterized them in cell lines and larvae of S. frugiperda and T. ni. The ie-2 mutant viruses exhibited delays in viral DNA synthesis, late gene expression, budded virus production, and occlusion body formation in SF-21 cells but not in TN-5B1-4 cells. In TN-5B1-4 cells, the ie-2 mutants produced more budded virus and fewer occlusion bodies but the infection proceeded without delay. Examination of the effects of ie-2 and the respective mutants on immediate-early viral promoters in transient expression assays revealed striking differences in the relative levels of expression and differences in responses to ie-2 and its mutant forms in different cell lines. In T. ni and S. frugiperda larvae, the infectivities of the occluded form of ie-2 mutant viruses by the normal oral route of infection was 100- and 1,000-fold lower, respectively, than that of wild-type AcMNPV. The reduction in oral infectivity was traced to the absence of virions within the occlusion bodies. The infectivity of the budded form of ie-2 mutants by hemocoelic injection was similar to that of wild-type virus in both species. Thus, ie-2 mutants are viable but exhibit cell line-specific effects on temporal regulation of the infection process. Due to its effect on virion occlusion, mutants of IE-2 were essentially noninfectious by the normal route of infection in both species tested. However, since budded viruses exhibited normal infectivity upon hemocoelic injection, we conclude that ie-2 does not affect host range per se. The possibility that IE-2 exerts tissue-specific effects has not been ruled out.     

In vivo correlation between blood T2* and oxygen saturation

The in vivo relationship between blood T2* and oxygen saturation was investigated. A wide range of blood oxygen saturation levels was created in pigs by altering the ventilation rate. Blood T2* was measured in vivo in the abdominal aorta and the inferior vena cava, corresponding to each oxygen saturation level. Our results indicate that it is possible to measure blood T2* in vivo reliably with an MR technique we previously proposed. Blood T2* correlates closely with oxygen saturation levels over a wide range, and the relationship between blood T2* measured in vivo and oxygen saturation can be approximated by a linear function.     

Proliferative effects of CXC chemokines in rat hepatocytes in vitro and in vivo

The CXC chemokines have well-documented neutrophil chemotactic, angiogenic, and mitogenic properties. The current investigations evaluate the effects of interleukin-8 (IL-8), epithelial neutrophil activating protein (ENA-78), and macrophage inflammatory peptide-2 (MIP-2) on hepatocyte proliferation in vitro and liver regeneration in vivo. Primary rat hepatocytes were isolated by collagenase digestion and exposed to incremental doses of IL-8, ENA-78, or MIP-2, and cellular proliferation was measured via tritiated thymidine incorporation. These experiments demonstrated significant increases in hepatocyte proliferation in response to IL-8, ENA-78, and MIP-2. Next, rats were sacrificed in a time-dependent manner following 70% hepatectomy or sham laparotomy, and hepatic tissue levels of MIP-2 and ENA-78 were measured using an ELISA. ENA-78 and MIP-2 were significantly elevated following 70% hepatectomy as compared with sham-operated control animals. Rats undergoing 70% hepatectomy were then treated with neutralizing anti-ENA-78 serum, anti-MIP-2 serum, or preimmune control serum, and liver regeneration was evaluated. These experiments demonstrated that neutralization of ENA-78 or MIP-2 slowed the rate of liver regeneration. These data suggest that the CXC chemokines may be important agents for the induction of hepatocyte proliferation and may be important molecules in vivo in the setting of liver injury, repair, and regeneration.     

In vivo antinociceptive activity of anti-rat mGluR1 and mGluR5 antibodies in rats

Conflicting results of an association between the human platelet antigen 1b (HPA-1b or PlA2) allele and the risk of myocardial infarction and coronary artery disease have been reported. To assess the reason for this discrepancy, we determined the HPA-1 genotype in 298 men who had undergone coronary angiography, including 124 individuals with myocardial infarction, 83 individuals with coronary artery disease but no history of myocardial infarction, and 91 control patients. Among patients with acute or recent onset myocardial infarction (< 1 year), the prevalence of HPA-1b was higher than among patients with coronary artery disease but without myocardial infarction (33 percent vs. 14 percent, p = 0.016). In patients under 60 years of age this difference was even more pronounced (45 percent vs. 15 percent, p = 0.003). Unlike conventional risk factors HPA-1b does not represent a risk factor for coronary artery disease itself but appears to be associated with increased platelet thrombogenicity.     

New 11 beta-aryl-substituted steroids exhibit both progestational and antiprogestational activity

The syntheses of three 11 beta-aryl-19-norpregna-4,9-dien-3-one derivatives with 17-spirolactone and 17 beta-hydroxy-17 alpha-cyanoethyl substitutions are described. The progesterone agonist/antagonist activities of the new compounds are investigated using a recently developed tissue culture system that relies on the progesterone agonist up-regulation of the prostate-specific antigen (PSA) gene in female breast tumor cell lines. Two of the newly synthesized compounds exhibit mixed agonistic/antagonistic progestational activity.     

In vitro and in vivo activity of 3-(1-methyl 1-5-nitro-2-imidazolylmethylideneamino)-2-oxazolidinone against Pasteurella

Furazolidone was more active than 3-(1-methyl)-5-nitro-2-imidazolyl-methylideneamino)-2-oxazolidinone (MABN) against a series of 34 isolates of Pasteurella and 11 of Yersinia (formerly designated Pasteurella). However, the nitroimidazole was superior to furazolidone by both subcutaneous and oral routes against a series of mouse infections incited by strains of Pasteurella. It also was superior to furazolidone and sulfaquinoxaline when administered in the diet against two Pasteurella strains in a fowl cholera model infection in chickens. The good in vitro activity of MABN plus its low toxicity suggest its further study as an agent for fowl cholera and the shipping fever complex of cattle.     

In vivo antitumor activity of hexamethylmelamine against human breast, stomach and colon carcinoma xenografts

We have evaluated the antitumor activity of Altretamine (hexamethylmelamine, HMM) on human carcinoma xenografts serially transplanted in nude mice. Five human breast carcinoma xenografts, MX-1, T-61, MCF-7, R-27 and Br-10, were inoculated subcutaneously into female nude mice. Two human stomach carcinoma xenografts, SC-1-NU and St-4, and three human colon carcinoma xenografts, Co-3, Co-4 and Co-6, were inoculated subcutaneously into male nude mice. One pellet of 17 beta-estradiol (0.1 mg/mouse) was inoculated subcutaneously in the mice transplanted with MCF-7 when the tumors were inoculated. HMM was administered per os daily for 4 weeks. MX-1 and T-61 tumors regressed completely after treatment with HMM at a dose of 75 mg/kg (the maximum tolerated dose: MTD) for MX-1 and 25 mg/kg for T-61. Br-10 was sensitive, whereas MCF-7 and R-27 were resistant to HMM at its MTD. HMM exerted the most potent antitumor effect against T-61. Against MX-1, it exerted an antitumor effect equivalent to that of cisplatin or cyclophosphamide. In addition, this agent was effective against all stomach and colon carcinoma xenografts, in particular St-4 (T/C% = 10.7: the mean tumor weight of treated group/the mean tumor weight of control group) and Co-3 (T/C% = 31.5%) which are insensitive to presently available agents. HMM seems worthy of further clinical investigation as a candidate agent to treat breast, stomach, colon and other carcinomas.     

In vitro and in vivo inhibitory actions of morin on rat brain phosphatidylinositolphosphate kinase activity

Phosphatidylinositol-4,5-bisphosphate occupies a central role in signal transduction and in cellular transformation. Phosphatidylinositol-4,5-bisphosphate is produced by the enzymatic phosphorylation of phosphatidylinositol-4-phosphate by phosphatidylinositolphosphate kinase (EC 2.7.1.68). Inhibition of this enzyme might conceivably lowers the cellular pool of phosphatidylinositol-4,5-bisphosphate, thus constituting a feasible control point in regulating signal transduction and cellular transformation. Morin, a plant flavonoid, was demonstrated to exhibit in vitro inhibitory action on phosphatidylinositolphosphate kinase extracted from rat brain. This inhibition of enzymatic activity was found to be dose-dependent, with an IC50 value of approximately 10 microM morin. Lineweaver-Burk transformation of the inhibition data indicates that inhibition was competitive with respect to ATP. The Ki was calculated to be 5.15 x 10(-6) M. Inhibition was uncompetitive with respect to phosphatidylinositol-4-phosphate. The Ki was determined to be 0.94 x 10(-5) M. Administration of morin to rats led to a decrease in phosphatidylinositolphosphate kinase activity in brain extracts. This in vivo action of morin was found to be dose-dependent and time-dependent. These effects of morin on rat brain phosphatidylinositolphosphate kinase activity are discussed in relation to the other reported biological actions of this flavonoid.     

Synthesis and evaluation of novel rhodacyanine dyes that exhibit antitumor activity

Rhodacyanine dyes and several analogous delocalized lipophilic cations (DLCs) were synthesized and evaluated as novel antitumor agents. Rhodacyanine dye consists of two heteroaromatic rings such as thiazoles at both termini of the conjugate systems and 4-oxothiazolidine (rhodanine) in the middle of it. Compounds with such a unique double-conjugate structure were found to inhibit the growth of several tumor cell lines, such as colon carcinoma CX-1, and to exhibit relatively low toxicity against normal kidney cell line CV-1 (e.g., IC50(CX-1) = 50 nM, IC50(CV-1) = 17.3 microM; selectivity index = 346 for compound 5). These compounds were also found to be efficacious in the tumor-bearing nude mice model (e.g., against human melanoma LOX; T/C (%) = 168 for compound 5). Structural modifications on rhodacyanine, including deletion of a heteroaromatic ring involved in the merocyanine conjugate system and replacement of rhodanine with a structurally related moiety such as 4-oxoimidazolidine or 4-oxo-1,3-dithiolane, resulted in a loss of the selectivity and/or the activity. Our current structure-activity studies imply that the double-conjugate system with a rhodanine moiety is essential for the selective activity of rhodacyanine dyes, and we find this class of compounds as unique antitumor agents candidates.