Mercurochrome Research Articles

Chlordane residues in rat and monkey tissues following subchronic inhalation exposure to technical chlordane.

Chlordane is a cyclodiene insecticide that was introduced in 1945. In recent years its use in the U.S.A. has been limited to termite control. The metabolism of chlordane and its residues in animals following oral intake has been documented. However, there are no published studies on chlordane residues in animals following inhalation exposure. This study sought to investigate the residues in the blood and tissues of rats and monkeys (both sexes) during 90-day subchronic inhalation exposure to technical chlordane. The effect of discontinued exposure on the depuration of residues in rats was also investigated.

The In Vivo Effects of Recombinant Human Interleukin-3: Demonstration of Basophil Differentiation Factor, Histamine-Producing Activity, and Priming of GM-CSF-Responsive Progenitors in Nonhuman Primates

Recently human interleukin-3 (IL-3) produced by molecular cloning was characterized as a growth factor for basophils and eosinophils in human bone marrow cultures. Since we found a similar activity of the human factor on simian bone marrow cells, we investigated the in vivo effects of recombinant human (rh) IL-3 in healthy rhesus monkeys (n = 10). rh IL-3 was administered subcutaneously (SC) to monkeys at different doses (11, 33, and 100 micrograms/kg/d) for 14 days followed by subsequent rh GM-CSF administration (5.5 micrograms/kg/d SC) for another two weeks. During the second week of rh IL-3 administration monkeys responded with a twofold to threefold increase of WBCs caused by a dose-dependent elevation of basophils (up to 40% of WBCs) and eosinophils. rh IL-3 also induced a dose-dependent increase of histamine (up to 700-fold above normal values) in monkey blood cells. Administration of rh GM-CSF to rh IL-3 pretreated monkeys resulted in a twofold enhanced increase in WBCs (due mainly to eosinophils and neutrophils) compared with animals treated with rh GM-CSF alone. Simultaneous administration of both cytokines (100 micrograms/kg rh IL-3 + 5.5 micrograms/kg rh GM-CSF SC) to two separate monkeys for 14 days induced a WBC elevation similar to that observed in monkeys treated with rh GM-CSF alone. In conclusion, our results indicate that rh IL-3 is a differentiation factor for blood basophils and primes the hematopoietic system for subsequent rh GM-CSF actions.

The in vivo effects of recombinant human interleukin-3: demonstration of basophil differentiation factor, histamine-producing activity, and priming of GM-CSF-responsive progenitors in nonhuman primates

Recently human interleukin-3 (IL-3) produced by molecular cloning was characterized as a growth factor for basophils and eosinophils in human bone marrow cultures. Since we found a similar activity of the human factor on simian bone marrow cells, we investigated the in vivo effects of recombinant human (rh) IL-3 in healthy rhesus monkeys (n = 10). rh IL-3 was administered subcutaneously (SC) to monkeys at different doses (11, 33, and 100 micrograms/kg/d) for 14 days followed by subsequent rh GM-CSF administration (5.5 micrograms/kg/d SC) for another two weeks. During the second week of rh IL-3 administration monkeys responded with a twofold to threefold increase of WBCs caused by a dose-dependent elevation of basophils (up to 40% of WBCs) and eosinophils. rh IL-3 also induced a dose-dependent increase of histamine (up to 700-fold above normal values) in monkey blood cells. Administration of rh GM-CSF to rh IL-3 pretreated monkeys resulted in a twofold enhanced increase in WBCs (due mainly to eosinophils and neutrophils) compared with animals treated with rh GM-CSF alone. Simultaneous administration of both cytokines (100 micrograms/kg rh IL-3 + 5.5 micrograms/kg rh GM-CSF SC) to two separate monkeys for 14 days induced a WBC elevation similar to that observed in monkeys treated with rh GM-CSF alone. In conclusion, our results indicate that rh IL-3 is a differentiation factor for blood basophils and primes the hematopoietic system for subsequent rh GM-CSF actions.

Colourimetric Determination of Pharmaceutical Thiocompounds and Allopurinol Using Mercurochrome

Abstract A new, simple and sensitive colourimetric method for determination of three pharmaceutical thiocompounds: cimetidine, thiabendazole and carbimazole, and also allopurinol, has been presented. the method is based on the reaction of each drug with mercurochrome (MER) in aqueous alkaline medium to give an intense red coloured chromogen. Optimization of the reaction conditions has been investigated. Obedience to Beer's law permitted the successful application of the proposed method for the assay of the investigated drugs in different dosage forms. Compared with the Pharmacopoeial methods, the proposed method gave equally accurate (t-test) and precise (F-test) results.

Inhibitory effect of a selective thromboxane A2receptor antagonist, EP 092, on platelet aggregation in whole bloodex vivoandin vivo

1. The inhibitory effect of a selective prostaglandin H2 (PGH2)/thromboxane A2 receptor antagonist, EP 092, on platelet aggregatory responses in whole blood ex vivo (guinea-pig: Rhesus monkey) and intravascular aggregation in vivo (rabbit) has been investigated. 2. Collagen (0.1-10.0 micrograms ml-1) caused a concentration-dependent decrease in single platelet count in samples of both guinea-pig and Rhesus monkey citrated whole blood incubated ex vivo. EP 092 administered to guinea-pigs by intravenous (0.1-3.0 mg kg-1) or oral (1.0-10.0 mg kg-1) routes significantly inhibited the platelet responses to collagen (ED50 values 1.3 +/- 0.2 and 1.4 +/- 0.2 mg kg-1 respectively). Similar potency against collagen-induced whole blood aggregation was observed in Rhesus monkey blood samples following EP 092 given orally (ED50 0.9 +/- 0.3 mg kg-1). 3. The duration of action of EP 092 against collagen aggregatory responses ex vivo in both guinea-pigs and Rhesus monkeys was between 3 and 6 h following oral administration at 3.0 mg kg-1. 4. The inhibitory activity demonstrated by EP 092 against collagen-induced aggregation of Rhesus monkey whole blood ex vivo was not accompanied by any significant reduction in thromboxane A2 formation except at the highest dose tested (10 mg kg-1). 5. The intravascular aggregatory response induced by collagen or thrombin in the anaesthetized rabbit was significantly inhibited by an intravenous infusion of EP 092 (10 mg kg-1). EP 092 appeared less potent and its effect was of shorter duration in this preparation compared with its inhibitory effect on ex vivo aggregation, being evident immediately after infusion of drug but not after a further 30 min. 6. It is concluded that collagen-induced platelet aggregatory response in guinea-pig and Rhesus monkey whole blood ex vivo and rabbit in vivo exhibit a thromboxane-dependent component which can be inhibited in a dose-related fashion by pretreatment with the thromboxane antagonist EP 092. In the rabbit, moreover, the data support the possibility of a role for thromboxane in the intravascular aggregatory response to thrombin.

Pharmacokinetics of Methylmercury in the Blood of Monkeys (Macaca fascicularis)

Pharmacokinetics of Methylmercury in the Blood of Monkeys (<i>Macaca fascicularis</i>)

Experimental american leishmaniasis and chagas' disease in the brazilian squirrel monkey: Cross immunity and electrocardiographic studies of monkeys infected with Leishmania braziliensis and Trypanosoma cruzi

Pung O. J., Hulsebos L. H. and Kuhn R. E. 1988. Experimental American leishmaniasis and Chagas' disease in the Brazilian squirrel monkey: cross immunity and electrocardiographic studies of monkeys infected with Leishmania braziliensis and Trypanosoma cruzi. International Journal for Parasitology 18: 1053–1059. Adult, laboratory-bred squirrel monkeys ( Saimiri sciureus) previously infected with either Leishmania braziliensis braziliensis or L. b. panamensis were challenge infected with blood-form trypomastigotes of Trypanosoma cruzi (Brazil strain). Monkeys previously infected with T. cruzi were challenged with stationary phase promastigote forms of L. b. braziliensis. Monkeys were examined during the course of challenge for evidence of infection, electrocardiographic alterations and parasite-specific antibody responses. T. cruzi epimastigotes were cultured from the blood of monkeys up to 3 months after challenge with this parasite. Unulcerated cutaneous lesions appeared and persisted in monkeys challenged with L. b. braziliensis. The formation of satellite lesions was observed in one monkey. Increased QRS intervals were not observed in T. cruzi challenged monkeys without prior cardiac irregularities and QRS left axis shifts were observed in only two of these monkeys. Elevated titers of parasite binding IgM and IgG specific for both T. cruzi and L. braziliensis were observed in all monkeys following challenge. These results indicate that prior infection with T. cruzi or L. braziliensis does not protect against heterologous challenge infection with these organisms. However, prior infection with Leishmania parasites may provide some protection against chagasic cardiopathies.

Developmental study of neuropeptide Y-like immunoreactivity in the neurohypophysis and intermediate lobe of the rhesus monkey (Macaca mulatta).

The purpose of this study was to examine the development and distribution of neuropeptide Y-immunoreactive fibers in the neurohypophysis of the rhesus monkey (Macaca mulatta) throughout life and the relationship of these fibers to the hypothalamo-hypophyseal portal vasculature. In rhesus monkeys, which varied in age from fetal life to 34 years, neuropeptide Y-immunoreactive fibers were present at all ages examined. In adult monkeys, varicose neuropeptide Y-labeled fibers were concentrated in the upper infundibular stem in association with capillary loops of the portal vasculature and the long portal vessels. Other fibers travelled down the infundibular stem and were distributed at the junction of the lower infundibular stem and infundibular process in the vicinity of the short portal vessels. In the infundibular process, neuropeptide Y-immunoreactive fibers were concentrated along the border of the intermediate lobe. Other stained fibers were sparsely distributed in the infundibular process and were often associated with small vessels. Neuropeptide Y-immunoreactivity was also located in a few fibers and cells of the intermediate lobe. Very few labeled fibers were seen in the fetal neurohypophysis, but their number increased gradually during the first postnatal year. At two years of age, a high density of stained fibers was observed, especially in the infundibular process. The number of axons in the infundibular process was lower at 12 years and continued to decline until 34 years of age. Neuropeptide Y may modulate hormone release at these sites and may also be released directly into vessels in the infundibular process. The close association of neuropeptide Y-labeled fibers with capillaries of the portal vasculature strongly suggests that neuropeptide Y is released into the portal blood of monkeys throughout life and may influence hormone secretion from the anterior pituitary gland.

Natural killer cells in rhesus monkeys: Properties of effector cells which lyse Raji targets

Spontaneously occurring natural killer cell activity of rhesus monkey peripheral blood mononuclear cells was assayed against five human cell lines, three of which were Epstein-Barr virus (EBV) positive, including the human B cell line Raji. The lytic activity to Raji cells was high, significantly higher than to any other cell line tested. Raji cells are normally insensitive to spontaneous lysis by human NK cells, and the contrasting vigor of the rhesus monkey cytolytic activity to Raji prompted us to investigate the properties of this effector cell. We found the effector cell-mediating lysis of Raji to be nonadherent and phagocytic with lytic activity slightly enhanced in the E-rosette-forming cell (ERFC +) fraction and decreased in the ERFC − fraction. Further isolation of FcIgG receptor-positive and FcIgG receptor-negative subsets by rosetting resulted in significant enrichment of NK activity to Raji in the positive fraction and a loss of activity in the negative fraction. Depletion studies with various monoclonal antibodies (mAb's) confirmed that nearly all lytic activity was contained in the CD16 + (Leu 11b +) population, while subsets of effector cells expressed CD2 (9.6) and CD8 (OKT8). Depletion of CD4 (OKT4)-, HLADR (OKIa)-, or LFA1 (MAC-1)-positive populations failed to reduce NK activity. We compared the phenotypic properties of alloimmune effector cells exhibiting specificity for allogeneic donor targets with those exhibiting lysis of Raji targets. Results indicated that allospecific cytotoxic T lymphocytes expressed a CD16 −, CD2 + phenotype, a pattern distinct from that of the effector cell population recognizing Raji targets. The presence of CD2 mAb's in the culture had no effect on NK lytic activity. In contrast, mAbs CD8 and Leu 11b were inhibitory. This would suggest a functional role for CD8 and FcIgG molecules in the lysis of Raji cells by rhesus effectors. In summary, these studies describe a distinct population of effector cells in the blood of rhesus monkeys which exhibit spontaneous lytic activity to Raji cells and exhibit the properties of NK cells.

Enumeration of Lymphocyte Subsets in Peripheral Blood of Rhesus Monkeys Prior to and Following Monocyte Removal using a Modified Indirect Avidin-Biotin Immunoperoxidase Technique

A modified avidin-biotin immunoperoxidase technique has been used to enumerate B lymphocytes, T inducer/helper (T H) and T cytotoxic/suppressor (T S) cells in the peripheral blood of normal and immunosuppressed rhesus monkeys ( Macaca mulatta) prior to and following adherent-cell depletion. The levels of each of the B, T H and T S cells detected in the normal monkeys using monoclonal antibodies which recognized specific surface antigens on human lymphocytes were comparable to the levels reported in human peripheral blood using direct immunofluorescence, immunoperoxidase or flow cytometry techniques. Adherent cell depletion did not result in a significant loss of any of the lymphocyte subpopulations examined. The technique is reproducible and sensitive in detecting differences between normal and immunosuppressed animals, and would prove to be useful in studies pertaining to chemical and drug immunomodulation.

Formaldehyde concentrations in the blood of rhesus monkeys after inhalation exposure

The effect of subchronic exposure to formaldehyde (HCHO; 6 ppm; 6 hr/day, 5 days/wk for 4 wk) on the HCHO concentration in the blood of three rhesus monkeys was investigated. Immediately after the final exposure, the monkeys were sedated, and blood samples were withdrawn 7 min after the end of exposure. The HCHO concentration in the blood, determined by gas chromatography-mass spectrometry was 1.84 ± 0.15 μg/g blood and did not differ significantly after a further 45 hr without exposure to HCHO (2.04 ± 0.40 μg/g blood). The average concentration of HCHO in the blood of exposed monkeys was also not significantly different from that of three unexposed controls (2.42 ± 0.09 μg/g blood). However, individual monkeys differed significantly from one another with respect to their blood concentrations of HCHO. These results indicate that subchronic inhalation exposure of non-human primates to HCHO has no significant effect on the HCHO concentration in the blood, and that the average concentration of HCHO in the blood of monkeys is similar to that in the blood of humans.

Metabolism of native and acetylated low density lipoproteins in squirrel monkeys with emphasis on aortas with varying severities of atherosclerosis

We studied the clearance of 131I-labeled native low density lipoproteins (LDL) and 125I-acetyl LDL from the blood of hypercholesterolemic and atherosclerotic squirrel monkeys which had been fed a semipurified diet supplemented with cholesterol for 3 years and from control monkeys which had been fed the same diet without cholesterol. In agreement with previous observations in other species, acetyl LDL left the circulation much more rapidly than native LDL. The cholesterol supplemented monkeys removed native 131I-LDL to the liver, the major site of clearance of both LDL forms, more slowly than controls. The overall clearance of 125I-acetyl' LDL was similar for the two groups. The aortic intima plus inner media (AIM) cleared both LDL forms much more slowly than other organs, and the ratio of acetyl LDL to native LDL cleared was quite high. The outer media (OM) showed less selectivity for acetyl LDL than the AIM. While LDL clearance by the OM was not affected by diet, the LDL clearance per g of AIM tissue was increased by 2-fold for both native and acetyl LDL in the cholesterol supplemented monkeys. These monkeys also had a 3-fold increase in AIM weight (due to intimal and subintimal thickening), which resulted in a 6-fold increase in the total LDL cleared by the AIM. The clearance of both LDL forms by the AIM correlated with three indices of atherosclerosis: intimal thickness, AIM weight, and AIM cholesterol concentration. The correlations were higher in the case of acetyl LDL clearance which may be due to the high affinity of the acetylated form for macrophages.

Natural killer cells in the blood and bone marrow of the rhesus monkey

Natural killer (NK) cells in peripheral blood lymphocytes (PBL) and bone marrow (BM) cells of the rhesus monkey were detected by their functional activity against K562 cells. Animals could be grouped into “high” or “low” NK responders, a trait found to be consistent over a period of 2 years. NK active cells in PBL were in the nonadherent population, with the majority bearing Fc receptors and a further subdivision of these into CR + (complement receptor) and CR − NK cells. Of 10 monoclonal antibodies directed against different epitopes of human lymphocytes, OKT11, OKT10, and Leu 11 showed reactivity wih rhesus NK cells. Only OKT10 was reactive with the effector site of the cell, as shown by its capability to block NK function. Of the Leu 11 monoclonal antibodies (a, b, c), Leu 11c was nonreactive while Leu 11a and Leu 11b were shown by immunofluorescence to bind to 7 to 21% of PBL; Leu 11b was also cytotoxic to the NK cells. Leu 11b did not prevent binding of Leu 11a to PBL, suggesting reactivity of these antibodies with different epitopes. Percoll fractionation of PBL and BM revealed a greater enrichment of NK activity with BM; also, with PBL peak NK activity occurred in fractions 4 and 5 while this occurred in fraction 5 with BM. Although Percoll PBL fractions contained a higher percentage of Leu 11b cells, the NK activity of the BM fractions was proportionately greater. The majority of PBL cells with NK activity were FcR + while significant activity could be attributed to FcR − cells of BM, in both the unseparated and Percoll fractions of each tissue. The data suggest NK active cells of BM may be distinct from those found in PBL.

Demonstration of clonable alloreactive host T cells in a primate model for bone marrow transplantation.

The phenomenon of marrow rejection following supralethal radiochemotherapy was explained in the past mainly by non-T-cell mechanisms known to be resistant to high-dose irradiation. In the present study a low but significant number of radiochemoresistant-clonable T cells was found in the peripheral blood and spleen of Rhesus monkeys following the cytoreductive protocol used for treatment of leukemia patients prior to bone marrow transplantation. More than 95% of the clonable cells are concentrated in the spleen 5 days after transplant. The cells possess immune memory as demonstrated by the generation of alloreactive-specific cytotoxicity. The present findings suggest that host-versus-graft activity may be mediated by alloreactive T cells. It is hoped that elimination of such cells prior to bone marrow transplantation will increase the engraftment rate of HLA-nonidentical marrow in leukemia patients.

Influence of luliberin and chorionic gonadotropin on luteinizing hormone and testosterone in the blood of monkeys under conditions of acute stress.

1. A single administration of luliberin to intact animais in a dose of 100 μg leads to a pronounced transitory rise in the LH and testosterone levels in the peripheral blood plasma. 2. Administration of luliberin 30 min before stress stimulates the secretion of LH by the pituitary, but does not prevent the stress-induced decrease in the testosterone level in the blood. 3. A single administration of 4500 units of hCG leads, in intact animals, to a sharp rise in the testosterone concentration, which remains at a high level for three days after stimulation of the gonads. 4. Preliminary stimulation of the testes by hCG prevents the stress-induced suppression of the testosterone level in the blood plasma; however, the reaction of the testes to the stimulating effect of hCG is decreased in this case.

Concentration of Plasmodium ovale- and Plasmodium vivax-infected erythrocytes from nonhuman primate blood using Percoll gradients.

Plasmodium vivax and Plasmodium ovale schizont-infected erythrocytes were separated from peripheral blood by centrifugation using discontinuous Percoll (colloidal silica) gradients. Infected Aotus monkey or chimpanzee blood was diluted and placed on a discontinuous gradient containing 30%, 40%, 45%, and 50% Percoll (v/v in media) layers before centrifugation at 1,450 X g. Parasitized erythrocytes were concentrated to greater than 95% schizont-infected cells in two bands that contained an average of one leukocyte per 500 infected cells. Mononuclear cells and trophozoites were isolated in another band and noninfected red cells, ring-infected cells, and granulocytes were pelleted to the bottom. The yield of parasitized erythrocytes ranged from 50% to close to 100% of the estimated number of infected cells in the original whole blood. Use of this Percoll procedure results in a high yield of concentrated parasitized erythrocytes and separation of these cells from host white blood cells.

Antibody-mediated targeting of liposomes to erythrocytes in the whole blood

F(ab′) 2 fragments derived from anti-rat erythrocyte antibody or normal rabbit serum IgG were covalently attached to the surface of liposomes consisting of equimolar amounts of egg phosphatidylcholine and cholesterol. These liposomes were interacted with rat, monkey or mouse blood, and their binding to both red and white blood cells was determined. Results of these studies show that coupling of liposomes to anti-rat erythrocyte F(ab′) 2 considerably enhances their binding to erythrocytes in rat blood. However, no such increase in the binding was observed with rat leukocytes or monkey and mouse erythrocytes. Besides, the interactions between the liposomes and target cells did not affect the permeability properties of the liposome bilayer. These observations indicate that liposomes coupled to cell-specific antibodies may serve as highly useful carriers for homing of drugs/enzymes to specific cells in biophase.

Inactivation and sedimentation of mercury from organomercurials byKlebsiella pneumoniae

Abstract A susceptibility of 63 clinical isolates of Klebsiella pneumoniae to inorganic and organic mercuric compounds was determined. 18 of them were found to be resistant to fluorescein mercuric acetate (FMA) and merbromin (MB). Moreover, all the resistant strains inactivate the antibacterial effect of FMA. The changes in the amount of organic mercury at the time of inactivation of the drug and the structures of the end products were examined in detail with the plasmid-bearing strain JK9 and its transconjugants of Escherichia coli . The results showed that FMA was inactivated by an intracellular enzyme produced inducively and was degraded to fluorescein (sodium salt, uranine), which led to the sedimentation of metallic mercury. The discovery of the genes conferring inducible organic mercury-inactivating enzymes determined by plasmids was the next step and their application in the recovery of metallic mercury from organomercurials is now imminent.

Dietary Protein Effects on Lipoproteins and on Sex and Thyroid Hormones in Blood of Rhesus Monkeys

We studied plasma lipoprotein and hormone concentrations in rhesus monkeys that had consumed either a low protein (3.8% of kilocalories) or a control protein (13.9%) purified diet since birth (6–10 yr before the beginning of this experiment) in order to test the hypothesis that chronic protein deficiency could influence plasma lipoproteins through an effect on the hepatic metabolism of gonadal or thyroid hormones. Protein-deficient monkeys had greater plasma concentrations of very low density lipoproteins (VLDL) plus high density lipoproteins (HDL) than controls. They also had lower serum albumin and greater alkaline phosphatase levels than the controls. Plasma thyroxine (T4) and free T4 concentrations were lower and the triiodothyronine (T3) levels tended to be greater in the protein-deficient group than in controls. This effect was apparent at two widely different levels of dietary iodide. Plasma T3 concentrations were elevated in other adult rhesus monkeys that were fed the low protein diet for only 6 wk. Monkeys injected with estradiol benzoate (100 µg/kg body weight) for 4 d had a marked reduction of VLDL concentrations. VLDL triglycerides were depressed more and plasma estrone levels were greater in deficient monkeys than in controls at 24 h after the last injection. In the control monkeys the T3 level rose and T4/T3 fell in response to estrogen injections, whereas the deficient monkeys did not respond.

Factors related to the appearance of alveolar macrophages in the developing lung.

The numbers of alveolar macrophages (AM) are known to be low in utero and to increase rapidly after birth. This study analyzed factors related to the appearance of AM in the lungs of term and preterm monkeys (Macaca nemestrina). We found that the numbers of AM were greater at birth in term than in preterm monkeys (p less than 0.01) and that they increased with postnatal age in term animals (r = 0.86) and in preterm animals without hyaline membrane disease (HMD) (r = 0.79). In contrast, the numbers of AM did not increase with postnatal age in preterm animals with HMD (r = 0.17). By multivariant analysis, AM numbers correlated most strongly with a combination of 2 variables: lavage fluid phospholipid and absence of HMD (r = 0.996). Preparations of alveolar surface-active material from newborn monkeys and to a lesser degree from adult monkeys were found to stimulate migration and polarization of monkey AM and blood monocytes in vitro. The migration-stimulating factor(s) in alveolar surface-active material had an apparent molecular weight greater than 10,000 and appeared not to be a lipoxygenase product of arachidonic acid metabolism. These data indicate a correlation between factors regulating lung pressure-volume stability and AM numbers and are consistent with a potential role for components of alveolar surface-active material in regulating the perinatal increase in AM.