Background: The ventral tegmental area (VTA) is well known for its role in cardiovascular control. It is demonstrated that about 20-30% of the VTA neurons are GABAergic though their role in cardiovascular control is not yet understood. This study is carried out to find the effects of GABA A and GABA B receptors on cardiovascular response of the VTA.
Methods: Experiments were performed on urethane anesthetized male Wistar rats. Drugs were microinjected unilaterally into the VTA. The average changes in mean arterial pressure (MAP) and heart rate (HR) were compared between the case and the control groups using t test and with the pre-injection values using paired t test.
Results: Microinjection of muscimol, a GABAA agonist (500, 1500 and 2500 pmol/100nl) into the VTA had no significant effect on MAP and HR compared with the saline group and pre-injection values. Injection of bicuculline methiodide (BMI, 100 and 200 pmol/100 nl), a GABAA antagonist, caused a significant increase in the MAP (11.1±1.95mmHg, P<0.5) and a decrease in HR (-32.07±10.2, P<0.01). Microinjection of baclofen a GABAB receptor agonist (500 or 1000 pmole/100 nl) and phaclofen a GABAB receptor antagonist (500 or 1000 pmole/100 nl) had no significant effects on MAP and HR.
Conclusion: For the first time it was demonstrated that GABA system of the VTA inhibits the cardiovascular system through the activation of GABAA but not the GABAB receptors.
Ventral tegmental area; GABA; Blood pressure; Heart rate
In Asia, Solanum nigrum fruit is traditionally used to manage, control, and treat diabetes.
This study was carried out to investigate the endothelium and nitric oxide roles in Solanum nigrum-induced vasorelaxation in non-diabetic and diabetic rat vessels.
Materials and Methods:
Diabetes was induced by a single i.p. injection of streptozotocin. Eight weeks later, superior mesenteric arteries of non-diabetic and diabetic groups were isolated and perfused according to the McGregor method. Solanum nigrum fruit extract (SNE) at concentrations of 0.00001 to 0.6 mg/ml was added to the medium and perfusion pressure was recorded.
Baseline perfusion pressure of diabetic group was significantly higher than non-diabetic rats in both intact and denuded endothelium. The low concentrations of SNE have vasodilatory effect in the diabetic and non-diabetic, but high concentrations of SNE produce initial significant contractions, followed by secondary relaxations in normal and diabetic rats. We observed vasorelaxation at low and high concentrations of SNE in both diabetic and non-diabetic groups after endothelium denudation. SNE-induced vasorelaxation in diabetic group is mediated by both endothelium and smooth muscle, but the relaxatory effect of SNE in non-diabetic group is not mediated by endothelium, and SNE has direct action on the smooth muscle.
Although the part of SNE-induced relaxation in diabetic vessel was mediated by endothelium, nitric oxide didn’t play any role in this action, and maybe we can use SNE in the management of diabetes vessel complications in future.
Diabetes mellitus; mesenteric bed; nitric oxide; Solanum nigrum fruit extract vasorelaxation
Chronic lung infection by Pseudomonas aeruginosa is the major severe complication in cystic fibrosis (CF) patients, where P. aeruginosa persists and grows in biofilms in the endobronchial mucus under hypoxic conditions. Numerous polymorphonuclear leukocytes (PMNs) surround the biofilms and create local anoxia by consuming the majority of O2 for production of reactive oxygen species (ROS). We hypothesized that P. aeruginosa acquires energy for growth in anaerobic endobronchial mucus by denitrification, which can be demonstrated by production of nitrous oxide (N2O), an intermediate in the denitrification pathway. We measured N2O and O2 with electrochemical microsensors in 8 freshly expectorated sputum samples from 7 CF patients with chronic P. aeruginosa infection. The concentrations of NO3− and NO2− in sputum were estimated by the Griess reagent. We found a maximum median concentration of 41.8 µM N2O (range 1.4–157.9 µM N2O). The concentration of N2O in the sputum was higher below the oxygenated layers. In 4 samples the N2O concentration increased during the initial 6 h of measurements before decreasing for approximately 6 h. Concomitantly, the concentration of NO3− decreased in sputum during 24 hours of incubation. We demonstrate for the first time production of N2O in clinical material from infected human airways indicating pathogenic metabolism based on denitrification. Therefore, P. aeruginosa may acquire energy for growth by denitrification in anoxic endobronchial mucus in CF patients. Such ability for anaerobic growth may be a hitherto ignored key aspect of chronic P. aeruginosa infections that can inform new strategies for treatment and prevention.
Products of the myc gene family integrate extracellular signals by modulating a wide range of their targets involved in cellular biogenesis and metabolism; the purpose of this integration is to regulate cell death, proliferation, and differentiation. However, understanding the regulation of myc at the transcription level remains a challenge. We performed rapid amplification of dmyc cDNA ends (5′ RACE) and mapped the transcription start site at P1 promoter, 18 base pairs upstream of the start of the known EST GM01143 and within the 5′ UTR. Our data show that the first TATA box, previously computationally predicted, is utilized to generate dmyc full length mRNA. The largest transcript contains all three exons, generated after the removal of the introns by constitutively regulated splicing events. Further investigation of Downstream Promoter Element (DPE) was achieved by studying lacZ reporter activity; investigation revealed that this element and its upstream cluster of binding sites are required for the dmyc intron 2 activity. These findings may provide valuable tools for further analysis of dmyc cis-elements.
dmyc; Drosophila; 5′ RACE; RNA splicing; TATA-box; Downstream Promoter Element (DPE)
Solanum nigrum fruit is traditionally used in Asia to manage, control, and treat diabetes but there is no scientific evidence of the efficacy of Solanum nigrum fruit in treatment of diabetes. We designed this study to investigate the effect of the administration of oral doses of aqueous extract from Solanum nigrum fruit on plasma glucose, lipid profiles, and the sensitivity of the vascular mesenteric bed to Phenylephrine in diabetic and non-diabetic rats.
Animals were divided into 5 groups (n=10): 2 groups served as non-diabetic controls (NDC), and the other groups had diabetes induced with a single injection of streptozotocin (STZ). Solanum nigrum-treated chronic diabetic (CD-SNE) and Solanum nigrum-treated controls (ND-SNE) received 1g/l of Solanum nigrum added to drinking water for 8 weeks. The mesenteric vascular beds were prepared using the McGregor method.
Administration of Solanum nigrum caused Ca/Mg ratio, plasma glucose, high-density lipoprotein (HDL), low-density lipoprotein (LDL), very low-density lipoprotein (VLDL), total cholesterol, and triglyceride concentrations to return to normal levels, and was shown to decrease alteration in vascular reactivity to vasoconstrictor agents.
Our results support the hypothesis that Solanum nigrum could play a role in the management of diabetes and the prevention of vascular complications in STZ-induced diabetic rats.
diabetes; lipid profile; Solanum nigrum; blood glucose; mesenteric bed; Ca/Mg ratio
The repetitive micro traumatic stresses placed on the athletes shoulder joint complex during the throwing motion challenge the surrounding tissues. The purpose of this study was to compare shoulder rotational strength, range of motion and proprioception between the throwing athletes and non-athletic persons.
Fifteen throwing athletes and 15 non-athletes participated in a nonrandom case – control study. Strength of shoulder rotational movements was tested with a hand held dynamometer. The ranges of internal and external rotation of shoulder were measured by a standard goniometer. The ability of subjects to replicate the target position and kinesthetic sense was examined on the subjects’ right shoulder by using a continuous passive motion device. Independent and paired t tests were used to statistically analyze between and within group differences.
No significant difference was detected on the range of internal rotation between throwing athletes and non-athletic candidates (P=0.3). The range of external rotation was significantly more in athletic subjects (P=0.03). The results also showed that throwing athletes demonstrated a significantly higher isometric strength of shoulder external and internal rotation than the non-athletic group (P<0.05). However, the comparison of the internal and external rotation strength of dominant side in each group showed that throwing athletes showed a significant lower isometric strength of shoulder external rotation than internal rotation (P<0.001). It was also demonstrated higher joint position acuity in the throwing athletes than non athlete subjects (P=0.01).
The repetitive nature of overhead throwing and the high forces that it causes result in adaptive changes of the dominant extremity. Throwing can lead to mobility, strength and neural adaptation.
Throwing Athletes; Muscle Weakness; Mobility Impairment; Proprioception
Myc is a crucial regulator of growth and proliferation during animal development. Many signals and transcription factors lead to changes in the expression levels of Drosophila myc, yet no clear model exists to explain the complexity of its regulation at the level of transcription. In this study we used Drosophila genetic tools to track the dmyc cis-regulatory elements. Bioinformatics analyses identified conserved sequence blocks in the noncoding regions of the dmyc gene. Investigation of lacZ reporter activity driven by upstream, downstream, and intronic sequences of the dmyc gene in embryonic, larval imaginal discs, larval brain, and adult ovaries, revealed that it is likely to be transcribed from multiple transcription initiation units including a far upstream regulatory region, a TATA box containing proximal complex and a TATA-less downstream promoter element in conjunction with an initiator within the intron 2 region. Our data provide evidence for a modular organization of dmyc regulatory sequences; these modules will most likely be required to generate the tissue-specific patterns of dmyc transcripts. The far upstream region is active in late embryogenesis, while activity of other cis elements is evident during embryogenesis, in specific larval imaginal tissues and during oogenesis. These data provide a framework for further investigation of the transcriptional regulatory mechanisms of dmyc.
dmyc; cis-regulatory module; enhancer; promoter; downstream promoter element; Drosophila
Diabetes is a worldwide high prevalence chronic progressive disease that poses a significant challenge to healthcare systems. The aim of this study is to provide a detailed economic burden of diagnosed type 2 diabetes mellitus (T2DM) and its complications in Iran in 2009 year.
This is a prevalence-based cost-of-illness study focusing on quantifying direct health care costs by bottom-up approach. Data on inpatient hospital services, outpatient clinic visits, physician services, drugs, laboratory test, education and non-medical cost were collected from two national registries. The human capital approach was used to calculate indirect costs separately in male and female and also among different age groups.
The total national cost of diagnosed T2DM in 2009 is estimated at 3.78 billion USA dollars (USD) including 2.04±0.28 billion direct (medical and non-medical) costs and indirect costs of 1.73 million. Average direct and indirect cost per capita was 842.6±102 and 864.8 USD respectively. Complications (48.9%) and drugs (23.8%) were main components of direct cost. The largest components of medical expenditures attributed to diabetes's complications are cardiovascular disease (42.3% of total Complications cost), nephropathy (23%) and ophthalmic complications (14%). Indirect costs include temporarily disability (335.7 million), permanent disability (452.4 million) and reduced productivity due to premature mortality (950.3 million).
T2DM is a costly disease in the Iran healthcare system and consume more than 8.69% of total health expenditure. In addition to these quantified costs, T2DM imposes high intangible costs on society in terms of reduced quality of life. Identification of effective new strategies for the control of diabetes and its complications is a public health priority.
Reliable evidence is the keystone for any noncommunicable disease (NCD) prevention plan to be initiated. In this study we carried out a risk factor investigation based on the WHO Stepwise approach to Surveillance (STEPS).
The study was conducted on 1000 adults between 15 and 64 years of age living in Ardabil province, north-west Iran during 2006, based on the WHO STEPS approach to surveillance of risk factors for NCD. At this stage only the first and second steps were carried out. Data were collected through standard questionnaires and methods analyzed using STATA version 8 statistical software package.
29.0% of men and 2.6% of women were current daily tobacco smokers. The mean number of manufactured cigarettes smoked per day was 18.9 among current daily smokers. Smoking was most prevalent among men of low-income families and those of lower education. The mean body mass index (BMI) was 26.6 kg/m2, and was significantly correlated with systolic blood pressure. 58.9% were overweight or obese; 18.0% had raised blood pressure and 3.7% had isolated systolic hypertension. The mean number of servings of fruit consumed per day was 1.1; 33.1% had low levels of activity. Combined risk factor analysis showed that 4.1% of participants were in the low-risk group (up to 5.1% among men and 3.2% among women). Those in the high-risk group comprised 25.6% in the 25- to 44-year age group and 49.7% in the 45- to 64-year age group. Mean BMI increased by age in both sexes at least at the first three decades of adult life.
Based on observed status of risk for cardiovascular health, burden of cardiovascular diseases is expected to increase if an effective prevention strategy is not undertaken.
cardiovascular health; noncommunicable diseases; WHO STEPS; smoking; obesity; physical activity
Mother's diet during pregnancy is important, since plant lignans and their metabolites, converted by the intestinal microflora to enterolignans, are proposed to possess multiple health benefits. Aim of our study was to investigate whether a dietary intervention affects lignan concentrations in the serum of pregnant women.
A controlled dietary intervention trial including 105 first-time pregnant women was conducted in three intervention and three control maternity health clinics. The intervention included individual counseling on diet and on physical activity, while the controls received conventional care. Blood samples were collected on gestation weeks 8-9 (baseline) and 36-37 (end of intervention). The serum levels of the plant lignans 7-hydroxymatairesinol, secoisolariciresinol, matairesinol, lariciresinol, cyclolariciresinol, and pinoresinol, and of the enterolignans 7-hydroxyenterolactone, enterodiol, and enterolactone, were measured using a validated method.
The baseline levels of enterolactone, enterodiol and the sum of lignans were higher in the control group, whereas at the end of the trial their levels were higher in the intervention group. The adjusted mean differences between the baseline and end of the intervention for enterolactone and the total lignan intake were 1.6 ng/ml (p = 0.018, 95% CI 1.1-2.3) and 1.4 ng/mg (p = 0.08, 95% CI 1.0-1.9) higher in the intervention group than in the controls. Further adjustment for dietary components did not change these associations.
The dietary intervention was successful in increasing the intake of lignan-rich food products, the fiber consumption and consequently the plasma levels of lignans in pregnant women.
A plasmid marker rescue system based on restoration of the nptII gene was established in Streptococcus gordonii to study the transfer of bacterial and transgenic plant DNA by transformation. In vitro studies revealed that the marker rescue efficiency depends on the type of donor DNA. Plasmid and chromosomal DNA of bacteria as well as DNA of transgenic potatoes were transferred with efficiencies ranging from 8.1 × 10−6 to 5.8 × 10−7 transformants per nptII gene. Using a 792-bp amplification product of nptII the efficiency was strongly decreased (9.8 × 10−9). In blood sausage, marker rescue using plasmid DNA was detectable (7.9 × 10−10), whereas in milk heat-inactivated horse serum (HHS) had to be added to obtain an efficiency of 2.7 × 10−11. No marker rescue was detected in extracts of transgenic potatoes despite addition of HHS. In vivo transformation of S. gordonii LTH 5597 was studied in monoassociated rats by using plasmid DNA. No marker rescue could be detected in vivo, although transformation was detected in the presence of saliva and fecal samples supplemented with HHS. It was also shown that plasmid DNA persists in rat saliva permitting transformation for up to 6 h of incubation. It is suggested that the lack of marker rescue is due to the absence of competence-stimulating factors such as serum proteins in rat saliva.
Our previous studies have shown that chalcones exhibit potent antileishmanial and antimalarial activities in vitro and in vivo. Preliminary studies showed that these compounds destroyed the ultrastructure of Leishmania parasite mitochondria and inhibited the respiration and the activity of mitochondrial dehydrogenases of Leishmania parasites. The present study was designed to further investigate the mechanism of action of chalcones, focusing on the parasite respiratory chain. The data show that licochalcone A inhibited the activity of fumarate reductase (FRD) in the permeabilized Leishmania major promastigote and in the parasite mitochondria, and it also inhibited solubilized FRD and a purified FRD from L. donovani. Two other chalcones, 2,4-dimethoxy-4′-allyloxychalcone (24m4ac) and 2,4-dimethoxy-4′-butoxychalcone (24mbc), also exhibited inhibitory effects on the activity of solubilized FRD in L. major promastigotes. Although licochalcone A inhibited the activities of succinate dehydrogenase (SDH), NADH dehydrogenase (NDH), and succinate- and NADH-cytochrome c reductases in the parasite mitochondria, the 50% inhibitory concentrations (IC50) of licochalcone A for these enzymes were at least 20 times higher than that for FRD. The IC50 of licochalcone A for SDH and NDH in human peripheral blood mononuclear cells were at least 70 times higher than that for FRD. These findings indicate that FRD, one of the enzymes of the parasite respiratory chain, might be the specific target for the chalcones tested. Since FRD exists in the Leishmania parasite and does not exist in mammalian cells, it could be an excellent target for antiprotozoal drugs.
Th1-type cellular immune responses play a critical role in protection against infection with Leishmania parasites, whereas activation of Th2-type cells results in progressive disease. Cutaneous leishmaniasis caused by Leishmania major is often a self-healing disease; however, persistent nonhealing forms are also known. In the present study, we have described cell-mediated immune responses in nonhealing patients by measuring T-cell proliferation, cytokine production, and phenotypic characterization of these cells. The responses were compared with those of patients with active lesions, patients who had recovered from infection, and healthy controls. Peripheral blood mononuclear cells from patients with active lesions and recovered donors proliferated vigorously and produced Th1-type cytokine when stimulated with L. major antigens, whereas in nonhealing patients the proliferative responses were significantly lower and showed a Th2-type response to Leishmania antigens. Interleukin-10 (IL-10) production was not a feature of L. major stimulation. Flow cytometric analysis revealed that L. major antigen induced proliferation of the CD4-positive population and that these cells were the major source of gamma interferon and IL-4. These results show a distinct dichotomy in the cytokine response to L. major infection.
A possible immunomodulatory role of granulocyte colony-stimulating factor (G-CSF) was investigated in an experimental pneumococcal meningitis model in rabbits. Animals were pretreated with G-CSF (10 μg/kg subcutaneously twice a day) starting 48 h before in vivo and ex vivo experiments, causing a five- to six-fold increase in the peripheral leukocyte level. Meningitis was induced by intracisternal inoculation of ∼4 × 105 CFU of Streptococcus pneumoniae type 3. Neutrophil pleocytosis and interleukin-8 (IL-8) levels were significantly attenuated in G-CSF-pretreated animals compared to untreated animals (P < 0.05). Furthermore, G-CSF pretreatment significantly delayed alterations in cerebrospinal fluid (CSF) tumor necrosis factor alpha and IL-1β levels, as well as protein and glucose levels (P < 0.05). No difference in CSF bacterial concentrations was found, whereas the blood bacterial concentration was significantly decreased in G-CSF-pretreated animals (P < 0.05). Ex vivo chemotaxis of neutrophils isolated from G-CSF-pretreated animals was significantly decreased compared to that of neutrophils from untreated animals (P < 0.05). In conclusion, G-CSF pretreatment attenuates meningeal inflammation and enhances systemic bacterial killing. Further preclinical studies are required to investigate whether this may affect the clinical course of meningitis and thus whether G-CSF treatment may have a beneficial role in pneumococcal meningitis.
Chronic Pseudomonas aeruginosa lung infection in cystic fibrosis (CF) patients is almost impossible to eradicate with antibiotic treatment. In the present study, the effects of treatment with the Chinese herbal medicine ginseng on blood polymorphonuclear leukocyte (PMN) chemiluminescence and serum specific antibody responses were studied in a rat model of chronic P. aeruginosa pneumonia mimicking CF. An aqueous extract of ginseng was administered by subcutaneous injection at a dosage of 25 mg/kg of body weight/day for 2 weeks. Saline was used as a control. Two weeks after the start of ginseng treatment, significantly increased PMN chemiluminescence (P ≤ 0.001) and a decreased level in serum of immunoglobulin G (IgG) against P. aeruginosa (P < 0.05) were found. Furthermore, a higher IgG2a level (P < 0.04) but lower IgG1 level (P < 0.04) were found in the ginseng-treated infected group than in the control group. In the ginseng-treated group the macroscopic lung pathology was milder (P = 0.0003) and the percent PMNs in the cells collected by bronchoalveolar lavage was lower (P = 0.0006) than in the control group. However, the alveolar macrophage (AM) chemiluminescence values were not significantly different in the two groups infected with P. aeruginosa. The differences between the ginseng-treated noninfected rats and the control group (without P. aeruginosa lung infection) for the PMN chemiluminescence and AM chemiluminescence were not significant. These results suggest that ginseng treatment leads to an activation of PMNs and modulation of the IgG response to P. aeruginosa, enhancing the bacterial clearance and thereby reducing the formation of immune complexes, resulting in a milder lung pathology. The changes in IgG1 and IgG2a subclasses indicate a possible shift from a Th-2-like to a Th-1-like response. These findings indicate that the therapeutic effects of ginseng may be related to activation of a Th-1 type of cellular immunity and down-regulation of humoral immunity.
The predominant pathogen in patients with cystic fibrosis (CF) is Pseudomonas aeruginosa, which results in a chronic lung infection associated with progressive pulmonary insufficiency. In a rat model of chronic P. aeruginosa pneumonia mimicking that in patients with CF, we studied whether the inflammation and antibody responses could be changed by treatment with the Chinese herbal medicine ginseng. An aqueous extract of ginseng was injected subcutaneously, and cortisone and saline were used as controls. Two weeks after challenge with P. aeruginosa, the ginseng-treated group showed a significantly improved bacterial clearance from the lungs (P < 0.04), less severe lung pathology (P = 0.05), lower lung abscess incidence (P < 0.01), and fewer mast cell numbers in the lung foci (P < 0.005). Furthermore, lower total immunoglobulin G (IgG) levels (P < 0.01) and higher IgG2a levels (P < 0.025) in serum against P. aeruginosa sonicate and a shift from an acute type to a chronic type of lung inflammation compared to those in the control and cortisone-treated groups were observed. These findings indicate that ginseng treatment of an experimental P. aeruginosa pneumonia in rats promotes a cellular response resembling a TH1-like response. On the basis of these results it is suggested that ginseng may have the potential to be a promising natural medicine, in conjunction with other forms of treatment, for CF patients with chronic P. aeruginosa lung infection.
Our previous studies have shown that licochalcone A, an oxygenated chalcone, has antileishmanial (M. Chen, S.B. Christensen, J. Blom, E. Lemmich, L. Nadelmann, K. Fich, T.G. Theander, and A. Kharazmi, Antimicrob, Agents Chemother. 37:2550-2556, 1993; M. Chen, S.B. Christensen, T.G. Theander, and A. Khrazmi, Antimicrob. Agents Chemother. 38:1339-1344, 1994) and antimalarial (M. Chen, T.G. Theander, S.B. Christensen, L. Hviid, L. Zhai, and A. Kaharazmi, Antimicrob. Agents Chemother. 38:1470-1475, 1994) activities. We have observed that licochalcone A alters the ultrastructure of the mitochondria of Leishmania promastigotes (Chen et al., Antimicrob. Agents Chemother. 37:2550-2556, 1993). The present study was designed to examine this observation further and investigate the mechanism of action of antileishmanial activity of licochalcone A. Electron microscopic studies showed that licochalcone A altered the ultrastructure of Leishmania major promastigote and amastigote mitochondria in a concentration-dependent manner without damaging the organelles of macrophages or the phagocytic function of these cells. Studies on the function of the parasite mitochondria showed that licochalcone A inhibited the respiration of the parasite by the parasites. Moreover, licochalcone A inhibited the activity of the parasite mitochondrial dehydrogenase. The inhibition of the activity of the parasite mitochondrial enzyme correlated well with the changes in the ultrastructure of the mitochondria shown by electron microscopy. These findings demonstrate that licochalcone A alters the ultrastructure and function of the mitochondria of Leishmania parasites.
Licochalcone A, isolated from Chinese licorice roots, inhibited the in vitro growth of both chloroquine-susceptible (3D7) and chloroquine-resistant (Dd2) Plasmodium falciparum strains in a [3H]hypoxanthine uptake assay. The growth inhibition of the chloroquine-resistant strain by licochalcone A was similar to that of the chloroquine-susceptible strain. To examine the activity of licochalcone A on the different asexual blood stages of the parasite, licochalcone A was added to highly synchronized cultures containing rings, trophozoites, and schizonts. The growth of the parasites at all stages was inhibited by licochalcone A. The in vivo activity of licochalcone A was tested in a mouse model of infection with P. yoelii. Licochalcone A administered either intraperitoneally or orally for 3 to 6 days protected the mice from the otherwise lethal P. yoelii infection. These results demonstrate that licochalcone A exhibits potent antimalarial activity and might be developed into a new antimalarial drug.
This study was designed to examine the antileishmanial activity of the oxygenated chalcone licochalcone A in mice and hamsters infected with Leishmania parasites. Intraperitoneal administration of licochalcone A at doses of 2.5 and 5 mg/kg of body weight per day completely prevented lesion development in BALB/c mice infected with Leishmania major. Treatment of hamsters infected with L. donovani with intraperitoneal administration of licochalcone A at a dose of 20 mg/kg of body weight per day for 6 consecutive days resulted in a > 96% reduction of parasite load in the liver and the spleen compared with values for untreated control animals. The [3H]thymidine uptake by the parasites isolated from the treated hamsters was only about 1% of that observed in parasites isolated from the controls. Oral administration of licochalcone A at concentrations of 5 to 150 mg/kg of body weight per day for 6 consecutive days resulted in > 65 and 85% reductions of L. donovani parasite loads in the liver and the spleen, respectively, compared with those of untreated control hamsters. These data clearly demonstrate that licochalcone A is a promising lead for the development of a new drug against leishmaniases.
Licochalcone A, an oxygenated chalcone isolated from the roots of Chinese licorice plant, inhibited the growth of both Leishmania major and Leishmania donovani promastigotes and amastigotes. The structure of the licochalcone A was established by mass and nuclear magnetic resonance spectroscopies and by synthesis, and its purity was verified by high-pressure liquid chromatography. The 50% inhibition of growth of logarithmic- and stationary-phase promastigotes of L. major, as measured by [3H]thymidine uptake, were 4 and 2.5 micrograms/ml, respectively. The growth of L. major promastigotes was totally inhibited after a 20-h incubation period with licochalcone A at 5 micrograms/ml. At a concentration of 0.5 microgram/ml, licochalcone A markedly reduced the infection rate of human peripheral blood monocyte-derived macrophages and U937 cells with L. major promastigotes and exhibited a strong intracellular killing of the parasite. These data show that intracellular Leishmania amastigotes are more susceptible than promastigotes to licochalcone A. Results of studies on the site of action of licochalcone A indicate that the target organelle appears to be the parasite mitochondria. These findings demonstrate that licochalcone A in concentrations that are nontoxic to host cells exhibits a strong antileishmanial activity and that appropriate substituted chalcones might be a new class of antileishmanial drugs.
Infections in humans by Leishmania donovani parasites can result in a fatal disease, visceral leishmaniasis (VL), or in a self-limiting asymptomatic infection. In murine models of the infection employing Leishmania major, the course of the disease can be directed into a VL-like syndrome by interleukin-4 (IL-4)-producing Th2 cells, or cure may result by Th1 cells secreting gamma interferon (IFN-gamma). The present study examined the potential of human T cells to generate Th1 or Th2 responses to L. donovani. The profiles of IFN-gamma, IL-4, and lymphotoxin secretion after antigen stimulation were analyzed in a panel of L. donovani-reactive CD4+ human T-cell clones generated from individuals who had recovered from VL after antimonial treatment. Two of the T-cell clones produced large amounts of IL-4 without production of IFN-gamma, seven clones produced both IFN-gamma and IL-4, and eight produced only IFN-gamma. This is the first report of a Th1- and Th2-type response in human leishmaniasis. These results suggest that in analogy with murine models, there is a dichotomy in the human T-cell response to L. donovani infections. Preferential activation of IL-4-producing Th2-like cells may be involved in the exacerbation of human VL, whereas activation of IFN-gamma-producing Th1 cells may protect the host from severe disease. Identification of leishmanial antigens activating one or the other type of T cells will be important in the development of vaccines against leishmaniasis.
Imipenem induced high levels of beta-lactamase production in Pseudomonas aeruginosa biofilms. Piperacillin also induced beta-lactamase production in these biofilms but to a lesser degree. The combination of beta-lactamase production with other protective properties of the biofilm mode of growth could be a major reason for the persistence of this sessile bacterium in chronic infections.
In this paper we describe functional and phenotypic changes in T cells after in vitro coincubation of peripheral blood mononuclear cells (PBMC) and Leishmania donovani parasites at different parasite/peripheral blood mononuclear cell ratios. The phytohemagglutinin (PHA)-induced lymphoproliferative response was reduced by the coincubation, and at the maximal parasite/peripheral blood mononuclear cell ratio used (7.5:1), the average response was less than 40% of the response in the absence of parasites. The cause of the reduction in lymphoproliferation is not clear, but it requires live parasites. Interleukin-1 production was unaffected, the levels of soluble interleukin-2 receptor in supernatants were not changed by the coincubation, and the addition of exogenous interleukin-2 failed to revert the suppressive effect of the parasites. In addition to the reduction in lymphocyte proliferation, phenotypic lymphocyte changes were observed. Cell surface expression of the CD3 antigen, which is part of the CD3-T-cell receptor complex, was significantly reduced with increasing parasite/peripheral blood mononuclear cell ratios; the reduction was general in the sense that the parasites caused a shift in the fluorescent intensities of anti-CD3 labeled cells toward lower values, without affecting the distribution pattern. In contrast, the parasites altered the CD25 (interleukin-2 receptor) expression on PHA-stimulated cells from a homogenous CD25-positive population to two populations, one small and without CD25 expression and the other, larger population with only a slight reduction in size and CD25 expression. In addition to the changes in expression of surface antigens, a general reduction in the size of PHA-stimulated lymphocytes after coincubation with the parasites was observed. The data presented thus suggest that the inhibition of the proliferative response to PHA by live L. donovani in vitro is associated with early processes in lymphocyte activation. Further studies on the inhibitory phenomena described may be of potential significance in the investigation of the suppressive mechanisms in human visceral leishmaniasis.
Alginate, a viscous polysaccharide from mucoid Pseudomonas aeruginosa, may interfere with the host defenses in patients with cystic fibrosis and chronic P. aeruginosa lung infection. The alginate concentration in the sol phase of expectorated sputum was quantitated by a biochemical method and a newly developed enzyme-linked immunosorbent assay. There was a high degree of correlation between the methods, and the concentration of alginate ranged from 4 to 101 micrograms/ml with a median of 35.5 micrograms/ml when measured by enzyme-linked immunosorbent assay. Alginate could not be detected in the bronchial secretions from patients without P. aeruginosa infection. In vitro investigation of alginate did not show any activation of the alternative pathway of complement, as determined by a hemolytic kinetic assay and by testing for neutrophil chemotaxis. At a high concentration, P. aeruginosa alginate caused a slight activation of the classical pathway of complement. Alginate did not cause neutrophil chemotaxis by itself but was able to reduce the neutrophil chemotactic response to N-formylmethionylleucylphenylalanine and for zymosan-activated serum. P. aeruginosa and seaweed alginates were able to prime neutrophils for increased N-formylmethionylleucylphenylalanine-induced neutrophil oxidative burst, as determined by chemiluminescence. Because of its ability to prevent attraction of neutrophils to the site of infection, lack of complement activation, and ability to enhance neutrophil oxidative burst, alginate from P. aeruginosa may contribute to the persistence and pathogenesis of chronic P. aeruginosa infection in cystic fibrosis.