Safety and efficacy of intravenous sodium stibogluconate in the treatment of leishmaniasis: recent U.S. military experience

The efficacy and toxicity of sodium stibogluconate (SSG) at a dosage of 20 mg/(kg.d) for either 20 days (for cutaneous disease) or 28 days (for visceral, mucosal, or viscerotropic disease) in the treatment of leishmaniasis is reported. Ninety-six U.S. Department of Defense health care beneficiaries with parasitologically confirmed leishmaniasis were prospectively followed for 1 year. One patient was infected with human immunodeficiency virus; otherwise, comorbidity was absent. Clinical cure occurred in 91% of 83 cases of cutaneous disease and 93% of 13 cases of visceral/viscerotropic disease. Adverse effects were common and necessitated interruption of treatment in 28% of cases, but they were generally reversible. These included arthralgias and myalgias (58%), pancreatitis (97%), transaminitis (67%), headache (22%), hematologic suppression (44%), and rash (9%). No subsequent mucosal leishmaniasis was identified, and there were no deaths attributable to SSG or leishmaniasis.     

Effect of treatment with BCG on the course of visceral leishmaniasis in BALB/c mice

Intravenous inoculation of BCG was found to be both prophylactic and therapeutic in BALB/c mice against challenge with amastigotes of Leishmania donovani. Spleens and livers of mice inoculated with BCG maintained total parasite burdens at significantly lower levels when compared to controls. BCG administered intravenously 14 days prior to and on the same day of protozoan challenge was more protective than vaccine given 30 and 14 days prior to challenge. A level of 10(7) viable units of BCG provided more protection against challenge with parasites than did 10(6) viable units. BCG given the same route as the challenge dose of amastigotes provided more protection than if administered via some other route. BCG given to mice with an already established infection was shown to significantly reduce their parasite burdens.     

Distribution of cholecystokinin immunoreactivity in the BALB/c mouse forebrain: an immunocytochemical study

The present study describes cholecystokinin (CCK) immunoreactivity (CCK-IR) distribution in the brains of control and colchicine-treated mice. In the brains of control mice, the CCK-IR strongly revealed numerous axons and terminals. Perikarya exhibiting a faint to moderate immunoreactivity were also observed in areas such as cortices, hippocampus, amygdala, septum, and thalamus. The colchicine treatment did not seem to notably affect the brain CCK-IR innervation, but resulted in profound changes of the perikaryal staining. Indeed, the regions, which contained numerous moderately stained perikarya in the control animals, exhibited after colchicine treatment immunoreactive perikarya intensely stained but only in moderate number. This feature obviously appeared in the cortex in which, in addition to strongly stained perikarya, colchicine induced the appearance of numerous CCK-IR hillocks. In the lateral amygdala and thalamus of colchicine-treated animals, the somatic immunoreactivity was considerably decreased. The regions, such as paraventricular hypothalamic nucleus and bed nucleus of the stria terminalis, which in the control animals did not exhibit any stained perikaryon, showed a high number of strongly stained cell bodies after colchicine treatment. This study, mapping the mouse forebrain CCK-IR, demonstrated a wide distribution of this peptide. Moreover, CCK-IR is spontaneously visible in neurons of untreated mouse in some brain areas previously shown in the rat to exhibit CCK mRNA, but no clear perikaryal CCK-IR even after colchicine treatment.     

Metabolic pathways of WR-2721 (ethyol, amifostine) in the BALB/c mouse

This study investigated the metabolism of the radio- and chemoprotector compound, WR-2721 [amifostine; s-2-(3- aminopropylamino)ethylphosphorothioate], in the Balb/c mouse. The latter was selected for these studies because considerable radiation protection data have been published for this mouse strain using the WR-2721 dose, route of administration, and optimal time for protection following intraperitoneal injection used herein. It is known that protection requires conversion of the parent drug to its free thiol metabolite, WR-1065, in cultured cells. Because it is possible that metabolites of WR-1065 could be involved in protection and because thiols are metabolically very reactive molecules, we investigated the metabolism of WR-2721 using electrochemical detection-HPLC methods. The following are the major findings in this study: 1) WR-2721 drug was rapidly cleared from the bloodstream. Blood concentration of the parent drug decreased 10-fold 30 min after administration from the maximal observed value at 5 min 2) WR-1065 rapidly appeared in the perchloric acid (PCA)-soluble fraction of normal solid tissues. The highest WR-1065 concentrations in liver and kidney were 965 and 2195 mumol/kg, respectively, 10 min after parent drug administration, whereas for heart and small intestine the highest values were 739 and 410 mumol/kg at 30 min. 3) WR-1065 accumulated in the PCA-soluble fraction of two experimental tumors at a lower rate than for the other tissues.(ABSTRACT TRUNCATED AT 250 WORDS)     

Visceral leishmaniasis in the hoary zorro Dusicyon vetulus: a case of mistaken identity

The historical identification of the Brazilian 'north-eastern' zorro as Dusicyon vetulus is questioned in relation to its incrimination as a reservoir of Leishmania chagasi, the agent of American visceral leishmaniasis. Comparative cranial and dental morphology showed that specimens of this north-eastern species more closely resemble the crab-eating zorro Cerdocyon thous, conforming with the documented geographical ranges of the respective species. We conclude that the single 'wild' canid host of L. chagasi in the neotropics in C. thous.     

Immunization of BALB/c mice with Helicobacter urease B induces a T helper 2 response absent in Helicobacter infection

BACKGROUND & AIMS: Infection with Helicobacter induces a T helper type 1 response in mice and humans. Mice can be cured or protected from infection with Helicobacter by mucosal immunization with recombinant H. pylori urease B subunit (rUreB). This study characterizes the immune response of infected mice immunized with rUreB. METHODS: BALB/c mice were infected with H. felis. Two weeks later, they were orally immunized four times with rUreB and cholera toxin (CT) at weekly intervals. Controls were only infected or sham-immunized with CT. Animals were killed at various times after immunization. Splenic CD4(+) cells were obtained and cultured in vitro with rUreB to evaluate antigen-specific proliferation and induction of interferon gamma and interleukin 4 secretion. RESULTS: All rUreB-immunized mice (n = 8) were cured from infection 3 weeks after the fourth immunization. Immunization induced a proliferative response of splenic CD4(+) cells, a progressive decrease in interferon gamma secretion, and a concomitant increase in interleukin 4 secretion after each immunization. A simultaneous increase in rUreB specific serum immunoglobulin G1 levels was observed in infected/immunized mice. CONCLUSIONS: In BALB/c mice, therapeutic mucosal immunization with rUreB induces progressively a Th2 CD4(+) T cell response resulting in the elimination of the pathogen.     

Chest discomfort associated with liposomal amphotericin B: report of three cases and review of the literature

Liposomal formulations of amphotericin B are designed to maintain therapeutic efficacy of amphotericin B deoxycholate while reducing its associated toxicities. In three patients chest discomfort occurred during planned 1-hour infusions of liposomal amphotericin B (AmBisome) 3 mg/kg/day during an open-label trial. The first patient experienced chest tightness and difficulty breathing and the second had dyspnea and acute hypoxia, both within 10 minutes of the start of the infusion. The third patient complained of chest pain 5 minutes after the start of two infusions. All symptoms resolved on terminating therapy. Two patients were later rechallenged with slower infusions and tolerated the drug well. A review of the English-language literature revealed only two other case reports of infusion-related chest or pulmonary reactions with the drug, although similar reactions were noted in several reports of clinical trials. Further review of the literature revealed reports of chest and pulmonary adverse events with other liposomal formulations of amphotericin B, liposomal daunorubicin, liposomal doxorubicin, and liposomes. The pathophysiology of such reactions remains unclear, and premedication with diphenhydramine did not completely prevent this reaction in one of our patients. We recommend infusing liposomal amphotericin B over at least 2 hours with careful monitoring for adverse reactions.     

Do the diminishing efficacy and increasing toxicity of sodium stibogluconate in the treatment of visceral leishmaniasis in Bihar, India, justify its continued use as a first-line drug? An observational study of 80 cases

The results of recent investigations of the analgesic and the nonanalgesic effects of opioid glucuronides are relevant to the research on drug abuse in forensic toxicology. As has been shown for heroin, knowledge of the state of distribution and elimination of active and inactive metabolites and glucuronides offers new possibilities in forensic interpretation of analytic results. Because of similar metabolic degradation, calculation of the time-dependent ratio of the concentration of morphine and its glucuronide metabolites in blood or serum allows a rough estimation of increased dosage and of time elapsed since the last application. Drug effects can be examined with respect to individual case histories, including overdose and survival time if the patient died. However, different methods of administration and the strong influence of different volumes or compartments of distribution of parent compounds and metabolites on concentrations in human body tissues require careful use of glucuronide concentration data. In Germany, dihydrocodeine (DHC) is prescribed as a heroin substitute, and relative overdoses are needed to be effective. DHC metabolism was studied in three patients who died from overdoses. All metabolites (dihydrocodeine-6-glucuronide [DHC6], nor-DHC [NDHC], dihydromorphine [DHM], nor-DHM [NDHM], and DHM-3- and 6-glucuronide [DHM3G, DHM6G]) were determined using HPLC and fluorescence detection. Concentrations of DHM (0.16 mg/L to 0.22 mg/L serum) were found. The DHM glucuronide ratios were similar to those of morphine. Receptor binding studies showed that the binding affinity of DHM to porcine mu-receptor was higher than that of morphine, and DHM6G's binding affinity was as high as that of morphine-6-glucuronide (M6G). Metabolites may play an important role in the effectiveness of DHC in substitution and toxicity. Because of enzyme polymorphism, the formation of DHC poses a risk for proper dosage in patients who are either poor or extensive metabolizers. The distribution of opioid glucuronides in cerebral spinal fluid in relation to transcellular transport in central nervous tissue is discussed with respect to the receptor binding of opiates and drug effect.     

Successful treatment of visceral leishmaniasis with allopurinol plus ketoconazole in a renal transplant recipient after the occurrence of pancreatitis due to stibogluconate

A case of a renal transplant recipient who developed pancreatitis during stibogluconate treatment for visceral leishmaniasis and who was successfully treated with a combination of allopurinol and ketoconazole is reported. The features of this case are compared with those of the three previously reported cases of pancreatitis during stibogluconate treatment. Complete cure was achieved during the follow-up period of 15 months. If stibogluconate is used for treatment of renal transplant recipients, we advise extreme caution with close observation and combination therapy to be considered instead.     

Superior efficacy of liposomal amphotericin B with prolonged circulation in blood in the treatment of severe candidiasis in leukopenic mice

In leukopenic mice with severe systemic candidiasis, single-dose treatment (5 mg of amphotericin B [AMB]/kg of body weight) with long-circulating polyethylene glycol-coated AMB liposomes (PEG-AMB-LIP) resulted in zero mortality and a significant reduction in the number of viable Candida albicans in the kidney, whereas 70% mortality was seen in mice treated with five daily doses of AmBisome (5 mg of AMB/kg . day). When the first of five daily doses of AmBisome was combined with a single low dose of Fungizone (0.1 mg of AMB/kg), the efficacy was equal to that of PEG-AMB-LIP.     

Dietary factors influence the recovery rates of Helicobacter pylori in a BALB/cA mouse model

We have identified a number of type I and type II keratins in the zebrafish Danio rerio by two-dimensional polyacrylamide gel electrophoresis, complementary keratin blot-binding assay and immunoblotting. These keratins range from 56 kDa to 46 kDa in molecular mass and from pH 6.6 to pH 5.2 in isoelectric point. Type II zebrafish keratins exhibit significantly higher molecular masses (56-52 kDa) compared with the type I keratins (50-48 kDa), but the isoelectric points show no significant difference between the two keratin subclasses (type II: pH 6.0-5.5; type I: pH 6.1-5.2). According to their occurrence in various zebrafish tissues, the identified keratins can be classified into "E" (epidermal) and "S" (simple epithelial) proteins. A panel of monoclonal anti-keratin antibodies has been used for immunoblotting of zebrafish cytoskeletal preparations and immunofluorescence microscopy of frozen tissue sections. These antibodies have revealed differential cytoplasmic expression of keratins; this not only includes epithelia, but also a variety of mesenchymally derived cells and tissues. Thus, previously detected fundamental differences in keratin expression patterns between higher vertebrates and a salmonid, the rainbow trout Oncorhynchus mykiss, also apply between vertebrates and the zebrafish, a cyprinid. However, in spite of notable similarities, trout and zebrafish keratins differ from each other in many details. The present data provide a firm basis from which the application of keratins as cell differentiation markers in the well-established genetic model organism, the zebrafish, can be developed.     

Induction of arthritis in BALB/c mice by cartilage link protein: involvement of distinct regions recognized by T and B lymphocytes

Both type II collagen and the proteoglycan aggrecan are capable of inducing an erosive inflammatory polyarthritis in mice. In this study we provide the first demonstration that link protein (LP), purified from bovine cartilage, can produce a persistent, erosive, inflammatory polyarthritis when injected repeatedly intraperitoneally into BALB/c mice. We discovered a single T-cell epitope, located within residues 266 to 290 of bovine LP (NDGAQIAKVGQIFAAWKLLGYDRCD), which is recognized by bovine LP-specific T lymphocytes. We also identified three immunogenic regions in bovine LP that contain epitopes recognized by antibodies in hyperimmunized sera. One of these B-cell regions is found in the most species-variable domain of LP (residues 1 to 36), whereas the other epitopes are located in the most conserved regions (residues 186 to 230 and 286 to 310). The latter two regions contain an AGWLSDGSVQYP motif shared by the G1 globulin domain of aggrecan core protein, versican, neurocan, glial hyaluronan-binding protein, and the hyaluronan receptor CD44. Our data reveal that the induction of arthritis is associated with antibody reactivities to B-cell epitopes located at residues 1 to 19. Together, these observations show that another cartilage protein, LP, like type II collagen and the proteoglycan aggrecan, is capable of inducing an erosive inflammatory arthritis in mice and that the immunity to LP involves recognition of both T- and B-cell epitopes. This immunity may be of importance in the pathogenesis of inflammatory joint diseases, such as juvenile rheumatoid arthritis, in which cellular immunity to LP has been demonstrated.     

In vitro activity of the echinocandin antifungal agent LY303,366 in comparison with itraconazole and amphotericin B against Aspergillus spp

LY303,366 (LY) is a novel derivative of the echinocandin class of antifungal agents. The in vitro activities of LY, itraconazole (ITZ), and amphotericin B (AMB) were assessed against 60 Aspergillus isolates, including 35 isolates of A. fumigatus, eight isolates of A. terreus, eight isolates of A. flavus, eight isolates of A. niger and one isolate of A. nidulans. Four A. fumigatus isolates were resistant to ITZ. Susceptibility testing for all drugs was performed with a broth microdilution procedure. LY was tested in two media: antibiotic medium 3 (AM3) and Casitone with 2% glucose (CAS) with an inoculum of 2 x 10(3) spores/ml. ITZ and AMB were tested in RPMI 1640 with 2% glucose with an inoculum of 1 x 10(6) spores/ml. All tests were incubated at 37 degrees C for 48 h. A novel end point was used to determine a minimal effective concentration (MEC) for LY, i. e., almost complete inhibition of growth save a few tiny spherical colonies attached to the microplate. MICs were measured for ITZ and AMB with a no-growth end point. Ranges and geometric mean (GM) MECs were from 0.0018 to >0.5 and 0.0039 mg/liter and from 0.0018 to >0.5 and 0.008 mg/liter for LY in AM3 and LY in CAS, respectively. Differences between species were apparent, with A. flavus being significantly less susceptible to LY than any other species tested with both media (P 16 and 0.7 mg/liter for ITZ and from 0.25 to 16 and 1.78 mg/liter for AMB. Minimal fungicidal concentrations (MFCs) were also determined for all drugs. GM MFCs were 0.018, 0.09, 19.76, and 12.64 mg/liter for LY in AM3, LY in CAS, ITZ, and AMB, respectively. LY in AM3 and LY in CAS were fungicidal for 86.7 and 68% of isolates, respectively (98% killing). In comparison, ITZ and AMB were fungicidal for 35 and 70% of isolates, respectively (99.99% killing). A reproducibility study was performed on 20% of the isolates. For 12 isolates retested, the MEC or MIC was the same or was within 1 dilution of the original value for 11, 11, 10, and 9 isolates for LY in AM3, LY in CAS, ITZ, and AMB, respectively. In conclusion, LY seems to be a promising antifungal agent with excellent in vitro activity against Aspergillus spp.     

Growth of P511 mastocytoma cells in BALB/c mouse brain elicits CTL response without tumor elimination: a new tumor model for regional central nervous system immunity

We have developed a murine model to explore the tumor-specific CTL response in the immune-privileged central nervous system using P511 mastocytoma cells. Three strains with varying degrees of histocompatibility to P511 cells (CD-1, allogeneic; BALB/c, minor histoincompatible; DBA/2, syngeneic) received tumor cells (10(4)) into the putamen 7 days after cannula implantation, when the blood-brain barrier was functionally intact. Without exception, tumor formed reproducibly by day 7 in all strains. Tumor rejection occurred in CD-1 but not in BALB/c and DBA/2 mice. Using a flank injection site, both CD-1 and BALB/c, but not DBA/2 mice, ultimately rejected flank tumors. Analysis of tumor-specific CTL in BALB/c spleens revealed that P511 administration into brain or flank elicited similar responses: no fully activated CTL were detectable but a significantly expanded population of nonkilling precursors of CTL (pCTL) were present. A P511 cell-specific pCTL population was also identified at the brain tumor site 14 days post-tumor introduction, indicating that pCTL, generated in the periphery, traffic to the tumor site in brain. These data indicate that failure to reject tumor in brain is neither due to lack of afferent stimulation nor to inability of peripheral effectors (P511 cell-specific pCTL) to reach the tumor site. We hypothesize that these effector cells are prevented from developing into fully activated CTL by conditions within the central nervous system microenvironment that down-regulate CTL development.     

Suppression of the benzylpenicilloyl- (BPO) specific IgE formation with isologous anti-idiotypic antibodies in BALB/c mice

In vivo effects of actively produced or passively administered isologous anti-idiotypic antisera (aId) on the benzylpenicilloyl- (BPO) specific IgE and IgG formation in BALB/c mice have been studied. Isologous anti-BPO aId were raised in BALB/c mice by immunization with purified anti-BPO antibodies isolated from ascites induced with BPO-bovine gamma-globulin in the same mouse strain. Mice producing isologous anti-BPO aId exhibited long-term suppression of BPO-specific IgE and IgG antibody responses induced by BPO-ovalbumin (BPO-OVA) in aluminum hydroxide. Simultaneously, they produced increased amounts of anti-BPO aId after each challenge with the BPO-OVA antigens. Passive administration of isologous anti-BPO aId into syngeneic mice previously sensitized with BPO-OVA caused depression of BPO-specific IgE antibody levels for 2 to 3 weeks. When anti-BPO IgE had again reached its previous level, passively administered aId had decreased to the level of untreated mice. Passive administration of anti-BPO aId also depressed the primary anti-BPO IgE formation for 2 to 3 weeks. In all these experiments the IgE antibody formation against the carrier proteins used for BPO-antigens was not affected. These results show that IgE and IgG antibodies share major idiotypic determinants and that IgE production is accessible to regulation by aId.     

B-cell activation in the mesenteric lymph nodes of resistant BALB/c mice infected with the murine nematode parasite Trichuris muris

Immune responses in resistant BALB/c mice infected with the murine nematode parasite Trichuris muris were examined. Following the establishment of infection, worm burdens of T. muris were expelled by BALB/c mice by day 21 postinfection (p.i.). Specific immunoglobulin G1 (IgG1) antibodies to T. muris excretory/secretory (E/S) antigens were detected in sera from infected mice, though specific IgG2a antibodies were not observed during infection. Ig-producing cells increased in the mesenteric lymph nodes (MLN) of infected mice on days 7, 14, and 21 p.i., with the greatest increase in numbers of IgG- and IgA-producing cells occurring on day 14. Marked increases in the relative percentages of B220+ and surface Ig+ (sIg+) cells were observed in the MLN of infected mice on days 14 and 21 p.i. Furthermore, cellular expansion of the MLN in infected mice resulted in an increase in the absolute numbers of B220+ and sIg+ cells. The levels of interleukin 2 (IL-2), IL-4, and interferon-gamma (IFN-gamma) detected in the supernatants from concanavalin A-stimulated MLN cells of infected mice were higher than those found in normal mice. Consequently, the expulsion of T. muris in resistant BALB/c mice was concomitant with cytokine production and B-cell activation in the MLN of infected mice. These results suggest the involvement of B-cell responses in protective immunity to T. muris infection.     

Interests of the 'artificial stomach' techniques to study antacid formulations: comparison with in vivo evaluation

In this overview, the methods for assessing antacid activity in vitro are surveyed, and the problems of their comparison with in vivo methods of evaluation are discussed. In vitro assessment is based on two types of method: static and dynamic. The static method of titration, with end-of-titration pH values ranging between 3.0 and 1.0, has been used to quantify the number of sites capable of binding H+ ions at each end-of-titration pH, and to identify certain chemical mechanisms involved in this binding; in other words, this approach provides the pharmacological characteristics of the drugs tested. In contrast, it does not take into account physiological factors modulating antacid activity, such as gastroduodenal fluxes (including gastric emptying), drug adherence to the mucosa, and acid secretion. The dynamic method was initially based on an artificial stomach model, which has gradually been upgraded to a computer-controlled artificial stomach-duodenum model. This model overcomes certain weaknesses of the static method by simulating flux and pH conditions in the gastroduodenal tract, by taking into account interactions with the gastric mucosa and thereby reproducing the in vivo medium encountered by antacids. It is therefore capable of reflecting the characteristics of antacids, namely their effect on gastric pH and resistance to acidification, at the same time helping to identify the underlying chemicophysical mechanisms. In vivo, the antacid effect can be assessed qualitatively by means of pH-meter studies in healthy volunteers, both in baseline conditions and during secretory stimulation, and also quantitatively by methods based on intragastric titration in response to a liquid meal (IGT). pH-meter studies in baseline conditions come up against the variability of the basal pH and antacid homogenization with gastric contents, which results in a wide range of individual values. This variability is found in pH-meter studies during pentagastrin infusion and, to a lesser degree, in response to a meal. Close correlations have, however, been established between results obtained with the artificial stomach model and in healthy volunteers submitted to pH-metric or IGT studies, with several antacids. It seems that the artificial stomach method is sufficiently reproducible to make it the method of choice for investigating the antacid activity of all drugs aimed at treating acid hypersecretion disorders. In contrast, in vivo studies may be warranted for precise therapeutic indications, such as treatment of duodenal ulcer or gastro-esophageal reflux, in which the therapeutic effect is judged on the basis of an improvement in symptoms and endoscopic criteria, without the need to demonstrate the antacid effect itself.