Brugia pahangi: differential induction and regulation of jird inflammatory responses by life-cycle stages

A novel, miniaturized high-throughput screening format is described for assay of combinatorial libraries generated on beads. This approach, which is ideally suited to encoded libraries synthesized on beads, utilizes the photolytic cleavage of individual compounds into a high-density well array (>6500 wells within a standard 96-well microtiter plate footprint) with well volumes as low as 0.37 microl. As a model study, an encoded dipeptide library (324 members) acylated with isobutyl succinate was assayed using this format to search for potential inhibitors of matrilysin, a member of the matrix metalloproteinase superfamily. In situ release of compounds from solid support was accomplished by photochemical cleavage after beads and enzyme were distributed to the wells. After the addition of a fluorogenic substrate to the array, the extent of enzyme inhibition and identification of active compounds was quantitated by imaging of the fluorescence emission upon uv irradiation. The structure-activity relationship data generated from the identified inhibitors in this study corroborate previous findings, thus validating the utility of this approach as a means of high-throughput screening of bead-based libraries.     

Neonatal and early life immune responses to various forms of vaccine antigens qualitatively differ from adult responses: predominance of a Th2-biased pattern which persists after adult boosting

Induction of neonatal immune responses to vaccine antigens is believed to be of limited efficacy because of immune immaturity and particular susceptibility to tolerogenic signals during this period of life. To characterize particular features of neonatal immune responses to vaccine antigens, we assessed the capacity of BALB/c mice at different stages of immunological maturation to respond to a selection of vaccine antigens and presentation systems. Significant B and T cell responses to vaccine antigens (tetanus and measles virus peptides, tetanus toxoid, live viral attenuated measles virus, canarypox recombinant measles vector or bacillus Calmette-Guérin) were obtained as early as the first week of life. However, these neonatal responses differed qualitatively from adult responses by a decreased IgG2a/IgG1 ratio of vaccine-specific antibodies, the secretion of significantly higher interleukin-5 and lower interferon-gamma levels by vaccine-specific T cells and an impaired induction of cytotoxic T cell precursors. This pattern of biased Th2 versus Th1 responses induced upon early exposure to vaccines was not reversed by decreasing the doses of vaccine antigens. It did not disappear with aging and was still reflected in adult responses to booster immunization with the corresponding antigen. Thus, neonatal immunization can induce significant vaccine specific responses with a predominance of a Th2 pattern which can persist in boosted adult mice.     

Malaria infection of the mosquito Anopheles gambiae activates immune-responsive genes during critical transition stages of the parasite life cycle

Six gene markers have been used to map the progress of the innate immune response of the mosquito vector, Anopheles gambiae, upon infection by the malaria parasite, Plasmodium berghei. In addition to four previously reported genes, the set of markers included NOS (a nitric oxide synthase gene fragment) and ICHIT (a gene encoding two putative chitin-binding domains separated by a polythreonine-rich mucin region). In the midgut, a robust response occurs at 24 h post-infection, at a time when malaria ookinetes traverse the midgut epithelium, but subsides at later phases of malaria development. In contrast, the salivary glands show no significant response at 24 h, but are activated in a prolonged late phase when sporozoites are released from the midgut into the haemolymph and invade the glands, between 10 and 25 days after blood feeding. Furthermore, the abdomen of the mosquito minus the midgut shows significant activation of immune markers, with complex kinetics that are distinct from those of both midgut and salivary glands. The parasite evidently elicits immune responses in multiple tissues of the mosquito, two of which are epithelia that the parasite must traverse to complete its development. The mechanisms of these responses and their significance for malaria transmission are discussed.     

Chemotherapy accelerates the development of acquired immune responses to Schistosoma haematobium infection

Treatment of 41 Schistosoma haematobium-infected children, 5-16 years old, with the drug praziquantel induced a switch from a predominantly IgA-specific antibody response to a predominantly IgG1 response within 12 weeks. A cross-sectional survey suggests that the same switch occurs naturally, but over several years, as children age (n = 251). The switch may be driven by alterations in cytokine levels in response to the release of antigens by dead or damaged parasites. Adults are more resistant to schistosome infection than children, and the switch to an "adult" response suggests that praziquantel treatment may have an immunizing effect, with benefits extending beyond a transient reduction in levels of infection.     

Cellular immune responses to acute stress in female caregivers of dementia patients and matched controls.

This study investigated whether the stress of caregiving alters cellular immune responses to acute psychological stressors. Twenty-seven women caring for a spouse with a progressive dementia (high chronic stress) and 37 controls matched for age and family income performed a 12-min laboratory stressor. Cellular immune function was assessed by both functional and quantitative measures taken before (low acute stress), immediately after (high acute stress), and 30 min after (recovery from stress) exposure to the laboratory stressors. The laboratory challenges were associated with diminished proliferative responses but elevated natural killer (NK) cell cytotoxicity; however, subsequent analyses suggested that this elevated cytotoxicity was largely attributable to an increase in the number of NK cells in peripheral blood. The results suggest that although the stress of caregiving diminishes cellular immune function, caregiving appears to have little effect on cellular immune responses to or recovery from brief psychological challenges. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Differential expression of two prolactin and growth hormone genes during early development of tilapia (Oreochromis mossambicus) in fresh water and seawater: implications for possible involvement in osmoregulation during early life stages

The possible involvement of prolactin (PRL) and growth hormone (GH) in osmoregulation during early life stages of the tilapia (Oreochromis mossambicus) was examined by in situ hybridization and immunocytochemistry using synthetic oligonucleotide probes and homologous antisera to two tilapia PRLs (PRL188 and PRL177) and GH. Hybridization signals for PRL188 mRNA were detected for the first time in newly hatched larvae (5 days after fertilization), and were significantly greater in larvae in fresh water (FW) than those in seawater (SW) until 10 days after hatching. PRL177 mRNA was detected in the pituitary of embryos 1 day before hatching. Although PRL177 gene expression in the embryo and newly hatched larvae in FW was not significantly different from those in SW, the expression was significantly greater in FW than in SW from Day 2 until Day 10. Hybridization signals for GH mRNA were first detected in newly hatched larvae. No significant differences in GH mRNA expression were observed between larvae in FW and those in SW. A stronger immunoreaction, a significantly larger PRL cell size, and a pituitary area containing PRL cells were observed in larvae hatched and maintained in FW, in those transferred from SW to FW, compared to larvae hatched and maintained in SW, and in those transferred from FW to SW. No significant difference was observed in the activity of GH cells between larvae in FW and those in SW. These results suggest that both PRLs are involved importantly in FW adaptation, whereas GH does not seem to play a critical role in osmoregulation during early stages of tilapia.     

Heat transfer and microstructure during the early stages of metal solidification

Transient heat transfer in the early stages of solidification of an alloy on a water-cooled chill and the consequent evolution of microstructure, quantified in terms of secondary dendrite arm spacing (SDAS), have been studied. Based on dip tests of the chill, instrumented with thermocouples, into Al-Si alloys, the influence of process variables such as mold surface roughness, mold material, metal superheat, alloy composition, and lubricant on heat transfer and cast structure has been determined. The heat flux between the solidifying metal and substrate, computed from measurements of transient temperature in the chill by the inverse heat-transfer technique, ranged from low values of 0.3 to 0.4 MW/m² to peak values of 0.95 to 2.0 MW/m². A onedimensional, implicit, finite-difference model was applied to compute heat-transfer coefficients, which ranged from 0.45 to 4.0 kW/m² °C, and local cooling rates of 10 °C/s to 100 °C/s near the chill surface, as well as growth of the solidifying shell. Near the chill surface, the SDAS varied from 12 to 22 (μm while at 20 mm from the chill, values of up to 80/smm were measured. Although the SDAS depended on the cooling rate and local solidification time, it was also found to be a direct function of the heat-transfer coefficient at distances very near to the casting/chill interface. A three-stage empirical heat-flux model based on the thermophysical properties of the mold and casting has been proposed for the simulation of the mold/casting boundary condition during solidification. The applicability of the various models proposed in the literature relating the SDAS to heat-transfer parameters has been evaluated and the extension of these models to continuous casting processes pursued.     

Respiratory responses to tracheobronchial stimulation during sleep and wakefulness in the adult cat

The response to tracheal stimulation (50 microliters of tap water) during wakefulness, non-rapid eye movement (NREM) sleep and rapid eye movement (REM) sleep was investigated in adult cats. In wakefulness, repetitive coughing occurred on 80% of the trials. In NREM and REM sleep, the most frequent response (approximately 69% and 58% of the trials, respectively) was arousal, followed by coughing. Apneas occurred following the stimulus and before arousal in 11% and 24% of the trials in NREM and REM sleep, respectively. In NREM sleep, the tracheal stimulus sometimes evoked expiratory efforts following a normal inspiratory effort (11% of the trials). These were much weaker than the expiratory efforts during coughing in wakefulness. In REM sleep, stimulation in 11% of the trials elicited increased inspiratory efforts. Although these may have been diminutive preparatory inspirations for coughing, they were much smaller than preparatory inspirations associated with coughing in wakefulness, and they were never followed by active expiratory efforts. Arousal from either NREM or REM sleep in response to tracheal stimulation was sometimes associated with an augmented breath. This response, which is common upon spontaneous arousal, may lead to deeper aspiration of the tracheal fluid. We conclude that in cats coughing requires wakefulness and that airway stimuli in sleep cause a variety of respiratory responses, some of which may be maladaptive.     

Effects of dietary NaCl deprivation during early development on behavioral and neurophysiological taste responses.

Investigated whether the gustatory system can be modified by restricting dietary NaCl during early development by recording neurophysiological taste responses in Sprague-Dawley rats at various times after deprivation (Exp I), and by measuring behavioral taste preferences in 3 groups of 7 NaCl deprived adult rats (Exp II). Overall findings indicate that Ss deprived of dietary NaCl from the 3rd day of gestation to 12 days postnatally and then placed on a NaCl-replete diet had chorda tympani nerve responses similar to those of nondeprived Ss when recordings were made at 28 days of age and older; however, preferences for NaCl solutions over water were significantly less than those of controls when tested at adulthood. NaCl deprivation in Ss from the 3rd day of gestation to approximately 35 days postnatally resulted in altered chorda tympani nerve responses to NaCl but not to other stimuli such as NH?Cl and KCl. Thus, it is concluded that restriction of dietary NaCl at a period in the rat's development when peripheral and central taste responses are changing results in short-term alterations in peripheral neural responses and in long-term changes in preference behaviors. (36 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Reproductive hormonal responses to ergotamine and ergonovine in cows during the luteal phase of the estrous cycle

We report the abnormal albumin in members of a Thai family that presented with high serum total T3 but not T4 when measured by radioimmunoassay. In contrast, total T3 values were very low when measured by ELISA and chemiluminescence. The subjects have no goiter, and clinically euthyroid. Their serum free T4, free T3, and TSH were normal. Spiking of T3 to affected serum showed good recovery by radioimmunoassay, but very poor recovery by ELISA and by chemiluminescence. The immunoprecipitation with labeled T3 bound to albumin showed high percent precipitation in affected serum. T3-binding studies showed that the association constant of serum albumin in affected subjects was 1.5 x 10(6) M-1 or 40-fold that of unaffected relatives of 3.9 x 10(4) M-1. In contrast, the Ka of HSA for T4 in an affected subject was only 1.5-fold that of a normal. Albumin complementary DNA from leukocytes of affected member was amplified and sequenced. We found the second nucleotide of normal codon 66 (CTT), a thymine, was substituted by a cytosine (CCT), resulting in the replacement of the normal leucine by proline. This is the first report of variant albumin causing familial dysalbuminemic hypertriiodothyroninemia.     

Effects of larval treatment with the insect development inhibitor PH60:40 on the vectorial capacity of Aedes aegypti (L.) for Brugia pahangi (Buckley and Edeson)

The numbers of mature worms which developed in young rats after their mothers were injected with 4000 L3 late in lactation were 1% (Nippostrongylus brasiliensis) and 24% (Strongyloides ratti) of the dose administered. The low value for N. brasiliensis validates the conclusion that milk is a real and important vehicle for infection in S. ratti since possible errors from skin invasion of the young would have been common to both species. The level of mature S. ratti infection in lactating mothers in this experiment was negligible, 97-99% of the adult worms appearing in the offspring. These results may indicate that the milk route is possible with N. brasiliensis even though it is quantitatively insignificant.     

Cellular immune responses of human cadaver donor bone marrow cells and their susceptibility to commonly used immunosuppressive drugs in transplantation

BACKGROUND: The cascade of immunological effects brought about by donor bone marrow cell (DBMC) infusions in human organ transplantation, especially in the context of continuous pharmacologic immunosuppression, is not fully understood. Yet, in inbred rodents and even primates, administration of specific bone marrow cells has caused a state of acquired immunologic tolerance. METHODS: In vitro mixed lymphocyte culture (MLC) and cell-mediated lympholysis (CML) culture systems were used to compare the responding and regulatory properties of DBMC and individual bone marrow cell subsets versus spleen cells in the presence or absence of pharmacologic immunosuppression. RESULTS: In the absence of immunosuppressive drugs, the DBMC proliferated in MLC and in response to phytohemagglutinin, but to a lower magnitude than donor spleen cells. In CML assays, DBMC failed to function as cytotoxic cells. Removal of both CD3+ and CD34+ cells together (not just singly) had to occur for complete abrogation of the proliferative response of DBMC evoked in the presence of allogeneic stimulating cells. Testing several experimental variables using flow cytometric analysis led to the conclusion that when purified DBMC CD34+ cells were placed in coculture with irradiated allogeneic peripheral blood mononuclear cells, such CD34+ cells give rise both to CD3- TCRalphabeta+ as well as to dimly staining CD3+ TCRalphabeta+ cells. Low pharmacologic concentrations of tacrolimus/cyclosporine (CsA) and mycophenolic acid (MPA) singly or in combination had no effect on the spontaneous proliferation of DBMC and had significantly less inhibitory activity on MLC responses of DBMC and its purified CD3+ or CD34+ subpopulations, compared with the responses of spleen cells. Moreover, the previously described regulatory effects of DBMC on the MLC responses of peripheral blood or splenic responding cells were not inhibited by these immunosuppressive drugs. CONCLUSIONS: Taken together, these results support the notion that in vitro DBMC subpopulations, which proliferate as responding cells in co-culture with x-irradiated allogeneic cells and which cause regulatory effects when added as a third component to MLC reactions, seem to be culture-generated lymphoid cell lineage(s) progeny of CD34+ cells. This possibly includes unique CD3+ "primitive" (dimly staining) T cells, which are not as inhibited in their function by tacrolimus/CsA and MPA, as are postthymic (splenic) T cells.     

Optimization of vaccine responses in early life: the role of delivery systems and immunomodulators

Pooled donor fibrin with an ultimate fibrinogen concentration of 60 mg/ml was used to study its effect on wound healing of surgically created ulcers in a rabbit ear. Water soluble polymer (PEG Mw = 20 KD) beads of 100-150 microns were added (12% by volume) to the fibrinogen to obtain a porous and rough structure. Five 6 mm-diameter ulcers to the depth of bare cartilage were created on each rabbit ear. There were two periods of study (4 and 8 days), with 15 ulcers in each time period, 5 of which were treated with a modified fibrin scaffold, 5 with a non-modified fibrin scaffold, and 5 served as control ulcers. The ulcer sites were subjected to routine histological processing and histomorphometrical quantification. Data analysis revealed significant increases in volume fraction of fibroblast and number of blood vessels in the modified fibrin scaffold treated ulcers over control and non-modified fibrin scaffold treated groups.     

Effects of experimental suppression of active (REM) sleep during early development upon adult brain and behavior in the rat

In order to test the hypothesis that active sleep (AS) is important for the normal development of the central nervous system, 3 different deprivation methods were applied to male Wistar rat pups during the first month of life. Daily injection of clomipramine from 8 to 21 days of age reduced the high level of AS to less than the adult value throughout most of the experimental period. Administration of clonidine from 8 to 21 days of life induced an almost total suppression of AS. Instrumental deprivation, using the 'pendulum' method, led to a significant (but less severe) AS reduction during 2-4 weeks of postnatal age. Open-field behavior testing in adulthood revealed a higher than normal level of ambulation in all 3 experimental groups. Masculine sexual responses were deficient, due to a low level of both mounts and ejaculations, in both clomipramine- and clonidine-treated animals. Neither passive avoidance learning nor dark preference tests revealed any differences between the experimental and control rats. Sleep observations showed that there was an abnormally high incidence of large myoclonic jerks during AS in both clomipramine- and clonidine-treated rats. Subsequent measurement of regional brain weights showed a significant reduction in the cerebral cortex and medulla oblongata, as compared with the respective control groups, in both the clomipramine- and the clonidine-treated rats. In addition, DNA and protein determination in the affected brain areas showed a proportional reduction in the cortex and in the medulla. These results demonstrate that interference with normal functioning either of AS per se or of specific monoaminergic transmitter systems during early development can produce long-lasting behavioral as well as brain morphological and biochemical abnormalities in later life.     

Programmed cell death during the earliest stages of spinal cord development in the chick embryo: a possible means of early phenotypic selection

The spatiotemporal distribution of cell death in the chick embryo neural tube and spinal cord (brachial region) was examined between stage (St.) 12 and 22, in plastic semithin sections. Between St. 12 and 16, the total number of pycnotic cells per segment was low, whereas after St. 16 the number of pycnotic cells was substantially increased. Between St. 17 and 19 three cell death foci or regions could be recognized. One region, the dorsal pycnotic zone, was located in the most dorsal part of the spinal cord, including the neural crest, with the highest number of pycnotic cells observed at St. 18. The second region, or ventral pycnotic zone, was located between motoneurons and the floor plate and had the highest number of dying cells at St. 17. The third region, the floor plate pycnotic zone, was located in the midportion of the floor plate and had the greatest amount of cell death at St. 19. Although low numbers of pycnotic cells were also observed in other regions between St. 17 and 19, no pycnotic cells were found in the ventrolateral region that gives rise to motoneurons. Ultrastructural observations as well as data from in situ nick end labeling indicate that the pycnotic cells observed in the neural tube die by apoptosis and that the debris from the dead cells is phagocytized primarily by adjacent healthy neuroepithelial cells. Although the spatiotemporal distribution of pycnotic cells suggests that cell death at these early stages could play a role in establishing the pioneer axonal pathway for spinal commissural neurons, preliminary observations following perturbations of cell death do not support this notion. Alternatively, early cell death may be involved in the regulation of cellular patterning along the dorsoventral axis of the neural tube by a kind of negative selection of specific progenitor cells.     

CD8+ T cells are activated during the early Th1 and Th2 immune responses in a murine Lyme disease model

T-helper responses following Borrelia burgdorferi infection in mice determine susceptibility to Lyme arthritis. The ratio of interleukin 4-positive, CD4+ to gamma interferon (IFN-gamma)-positive, CD4+ T cells was significantly greater in infected BALB/cJ mice than in infected C3H/HeJ mice. Increased numbers of IFN-gamma-producing cells predicted greater arthritis severity, and CD8+ T cells were the main source of IFN-gamma in both strains.     

Local immune responses to Chlamydia pneumoniae in the lungs of BALB/c mice during primary infection and reinfection

Cell-mediated immune (CMI) responses play a major role in protection as well as pathogenesis of many intracellular bacterial infections. In this study, we evaluated the infection kinetics and assessed histologically the lymphoid reactions and local, in vitro-restimulated CMI responses in lungs of BALB/c mice, during both primary infection and reinfection with Chlamydia pneumoniae. The primary challenge resulted in a self-restricted infection with elimination of culturable bacteria by day 27 after challenge. A mild lymphoid reaction characterized the pathology in the lungs. In vitro CMI responses consisted of a weak proliferative response and no secretion of gamma interferon (IFN-gamma). The number of lung-derived mononuclear cells increased substantially during the primary infection; the largest relative increase was observed in B cells (B220(+)). After reinfection, the number of lung-derived mononuclear cells increased further, and the response consisted mainly of T cells. The reinfection was characterized in vivo by significant protection from infection (fewer cultivable bacteria in the lungs for a shorter period of time) but increased local lymphoid reaction at the infection site. In vitro, as opposed to the response in naive mice, acquired immunity was characterized by a strongly Th1-biased (IFN-gamma) CMI response. These results suggest that repeated infections with C. pneumoniae may induce Th1-type responses with similar associated tissue reactions, as shown in C. trachomatis infection models.     

Phase I trial of recombinant adenovirus gene transfer in lung cancer. Longitudinal study of the immune responses to transgene and viral products

Animal studies indicate that the use of replication-deficient adenovirus for human gene therapy is limited by host antivector immune responses that result in transient recombinant protein expression and blocking of gene transfer when rechallenged. Therefore, we have examined immune responses to an adenoviral vector and to the beta-galactosidase protein in four patients with lung cancer given a single intratumor injection of 10(9) plaque-forming units of recombinant adenovirus. The beta-galactosidase protein was expressed in day-8 tumor biopsies from all patients at variable levels. Recombinant virus DNA was detected by PCR in day-30 and day-60 tumor biopsies from all patients except patient 1. A high level of neutralizing antiadenovirus antibodies was detected in patient 1 before Ad-beta-gal injection whereas it was low (patient 3) or undetectable in the other two patients. All patients developed potent CD4 type 1 helper T cell (Th1) responses to adenoviral particles which increased gradually over time after injection. Antiadenovirus cytotoxic T lymphocyte responses were consistently boosted in the two patients examined (patients 3 and 4). Sustained production of anti-beta-galactosidase IgG was observed in all patients except patient 1. Consistent with anti-beta-gal antibody production, all patients except patient 1 developed intense, dose-dependent Th1 responses to soluble beta-galactosidase which increased over time. Strong beta-galactosidase-specific cytotoxic T lymphocyte responses were detected in patients 2, 3, and 4. Our results clearly show that despite the intensity of antiadenovirus responses, transgene protein expression was sufficient to induce strong and prolonged immunity in three patients. Recombinant adenovirus injected directly into the tumor is a highly efficient vector for immunizing patients against the transgene protein.     

Cellular development of some embryonic organs and the chorion during the first trimester of human pregnancy

The DNA, RNA and protein content of chorionic tissue, heart, liver, kidney, lung and brain was estimated in 27 fetuses obtained at termination of pregnancy between 50 and 105 days of menstrual age. For brain, heart and liver, growth appeared to be most rapid at the earliest period studied (50 to 60 days) whereas kidney and lung development was most rapid later in the first trimester. It is suggested that these periods of particularly rapid growth may be the times at which those tissues are most vulnerable to injury.