Human and Murine Immune Responses to a Novel Leishmania major Recombinant Protein Encoded by Members of a Multicopy Gene Family

Vaccination of BALB/c mice with Leishmania major promastigote culture filtrate proteins plus Corynebacterium parvum confers resistance to infection with L. major. To define immunogenic components of this protein mixture, we used sera from vaccinated mice to screen an L. major amastigote cDNA expression library. One of the immunoreactive clones thus obtained encoded a novel protein of L. major with a molecular mass of 22.1 kDa. The predicted amino acid sequence of this clone exhibited significant homology to eukaryotic thiol-specific-antioxidant (TSA) proteins. Therefore, we have designated this protein L. major TSA protein. Southern blot hybridization analyses indicate that there are multiple copies of the TSA gene in all species of Leishmania analyzed. Northern blot analyses demonstrated that the TSA gene is constitutively expressed in L. major promastigotes and amastigotes. Recombinant TSA protein containing an amino-terminal six-histidine tag was expressed in Escherichia coli with the pET17b system and was purified to homogeneity by affinity chromatography. Immunization of BALB/c mice with recombinant TSA protein resulted in the development of strong cellular immune responses and conferred protective immune responses against infection with L. major when the protein was combined with interleukin 12. In addition, recombinant TSA protein elicited in vitro proliferative responses from peripheral blood mononuclear cells of human leishmaniasis patients and significant TSA protein-specific antibody titers were detected in sera of both cutaneous-leishmaniasis and visceral-leishmaniasis patients. Together, these data suggest that the TSA protein may be useful as a component of a subunit vaccine against leishmaniasis.     

Brain lysosomal glycosidase activity in immunosuppressed mice infected with avirulent Semliki forest virus.

Mice infected with an avirulent strain of Semliki forest virus show an increase in the activity of some of the brain lysosomal glycosidases. The increase in activity of these enzymes has been correlated with the histological, virological, and serological changes that result from the infection in the presence and absence of immunosuppression. Semliki forest virus alone caused the development of a mild encephalitis with perivascular infiltration, microgliosis, astrocyte hypertrophy, and a focal spongiform encephalopathy, together with an increased activity of brain N-acetyl-beta-D-glucosaminidase and beta-glucuronidase. Antilymphocyte serum given after infection marginally affected the course of the disease. Cyclophosphamide markedly delayed the development of the spongy changes in the increase in enzyme activities, but not the perivascular infiltration. It is suggested that the increased activity of the lysosomal glycosidases studied may be linked both to the development of a successful immune response and to the focal spongiform changes produced by the infection.     

Multiomics uncovers developing immunological lineages in human

The development of the human immune system during embryonic and fetal life has historically been difficult to research due to limited access to human tissue. Experimental animal models have been widely used to study development but cellular and molecular programmes may not be conserved across species. The advent of multiomic single-cell technologies and an increase in human developmental tissue biobank resources have facilitated single-cell multiomic studies focused on human immune development. A critical question in the near future is "How do we best reconcile scientific findings across multiple omic modalities, developmental time, and organismic space?" In this review, we discuss the application of single-cell multiomic technologies to unravel the major cellular lineages in the prenatal human immune system. We also identify key areas where the combined power of multiomics technologies can be leveraged to address specific immunological gaps in our current knowledge and explore new research horizons in human development.     

Concurrent de novo interstitial deletion of band 2p22 and reciprocal translocation (3;7)(p21;q22).

A child is described with a de novo interstitial deletion of band 2p22 and a reciprocal translocation (3;7)(p21;q22). The child has mild developmental delay, coloboma of the right eye, and Hirschsprung's disease. The clinical and cytogenetic findings are described.     

Putative role for interleukin-7 in the maintenance of the recirculating naive CD4+ T-cell pool

The capacity of the immune system to respond efficiently to new antigens depends upon a continuous source of naive CD4+ T cells. Such cells exit from the thymus and join the recirculated T-cell pool. Factors present at the sites of naive CD4+ T-cell circulation must be responsible for their survival, since upon removal from their host, naive CD4+ T cells die. However, such factors remain unknown. The presence of the cytokine interleukin-7 (IL-7) in secondary lymphoid organs and the continuous expression of its receptor on naive CD4+ T cells prompted us to examine the possibility that IL-7 might be a survival factor for naive CD4+ T cells. Using naive CD4+ T cells isolated from cord blood we show that IL-7, but not IL-2, can maintain naive CD4+ T-cell viability in vitro for at least 15 days. In addition, we find that IL-7 can induce modest proliferation of naive CD4+ T cells without affecting either their cell surface phenotype or their ability to respond to antigenic stimulation. We also find that after anti-CD3 stimulation, naive CD4+ T cells lose that ability to respond to IL-7. However, if cells are primed with IL-7 prior to antigenic stimulation, their proliferative responses are enhanced. Together, these data suggest a novel and important role for IL-7 in the maintenance and maturation of naive CD4+ T cells, ensuring that they can respond maximally when they first meet antigen in secondary lymphoid tissue.     

Control of motor-unit tension by rate modulation during sustained contractions in reinnervated cat muscle

1. The aim of this study was to describe the control of tension by rate modulation of single motor units in reinnervated muscle. 2. Single fast-twitch motor units were isolated from medial gastrocnemius (MG) muscles in two groups of anesthetized adult cats: one in which the MG nerve was left untreated and another in which that nerve was sectioned and immediately sutured together 10-33 mo before study. Together with conventional measures of isometric contractile properties, units were tested with the use of computer-controlled feedback regulation of stimulation rate to maintain tension during continuous isometric contraction at a constant submaximal level [25% of maximal tension (Pmax)]. 3. For motor units from both groups, stimulation rate began to decline after target tension was attained and then settled at lower values for variable durations before rapidly increasing, usually within the last 5% of the contraction's duration, until reaching the experimentally selected limit of 100 pulses/s (pps). 4. Measures of the declining phase in stimulation rate occurring at the beginning of sustained contraction were not significantly different in comparison of untreated versus reinnervated muscles. These measures included 1) the magnitude of the decrease in rate, 2) the minimum rate attained, and 3) the time taken to reach minimum stimulation rate expressed as a fraction of endurance time (Et, total duration of the sustained contraction). 5. Most fast-twitch units from reinnervated muscles fell within normal limits for both endurance time and the number of stimuli applied over that period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of Phagocytosis-Associated Chemiluminescence by Superoxide Dismutase

During the process of phagocytosis, human leukocytes emit a burst of luminescence which can be measured in a liquid scintillation spectrometer. The enzyme superoxide dismutase, which removes superoxide anions (O(2.)), inhibited this chemiluminescence by 70% at a concentration of 100 mug/ml. The enzyme did not inhibit phagocytosis. These results support other studies indicating that O(2.) is elaborated by phagocytizing leukocytes. They also indicate that O(2.) plays a major role in phagocytosis-associated chemiluminescence, though not necessarily as the luminescing agent. Catalase and benzoate inhibited the chemiluminescence of phagocytosis to a slight extent, suggesting that hydrogen peroxide and hydroxyl radical, respectively, might also be involved in this phenomenon. The relationship between the mediators of chemiluminescence and those responsible for phagocytic bactericidal activity remains to be defined.     

Composite articular cartilage engineered on a chondrocyte-seeded aliphatic polyurethane sponge

To circumvent the reconstructive disadvantages inherent in resorbable polyglycolic acid (PGA)/polylactic acid (PLA) used in cartilage engineering, a nonresorbable, and nonreactive polyurethane sponge (Tecoflex sponge, TS) was studied as both a cell delivery device and as an internal support scaffolding. The in vitro viability and proliferation of porcine articular chondrocytes (PACs) in TS, and the in vivo generation of new articular cartilage and long-term resorption, were examined. The initial cell attachment rate was 40%, and cell density increased more than 5-fold after 12 days of culture in vitro. PAC-loaded TS blocks were implanted into nude mice, became opalescent, and resembled native cartilage at weeks 12 and 24 postimplantation. The mass and volume of newly formed cartilage were not significantly different at week 24 from samples harvested at week 6 or week 12. Safranin O-fast green staining revealed that the specimens from cell-loaded TS groups at week 12 and week 24 consisted of mature cartilage. Collagen typing revealed that type II collagen was present in all groups of tissue-engineered cartilage. In conclusion, the implantation of PAC-TS resulted in composite tissue-engineered articular cartilage with TS as an internal support. Long-term observation (24 weeks) of mass and volume showed no evidence of resorption.     

Non-cholinergic effects of acetylcholinesterase in the substantia nigra: a possible role for an ATP-sensitive potassium channel

Summary Within the substantia nigra acetylcholinesterase has non-cholinergic actions that can be demonstrated at both behavioural and cellular levels: the aim of this study was, thus, to explore, in the in vitro guinea pig substantia nigra, the ionic mechanisms which mediate these non-classical phenomena. Acetylcholinesterase had a reversible hyperpolarizing action, via an opening of potassium channels, on a selective population of nigral neurons. These neurons could be identified by an ability to generate bursts of action potentials and by a sensitivity to either amphetamine or to a reduction of glucose in the perfusing medium. The acetylcholinesterase-induced hyperpolarization could not be attributed to a contaminant in the exogenous solution, since a highly purified preparation was even more potent. Furthermore, enzymatic action of any kind could be eliminated as boiled acetylcholinesterase was equally efficacious. The effect of acetylcholinesterase was not subject to tachyphylaxis and was resistant to blockade of potassium channels with tetraethylammonium: since both these phenomena are features of the D₂ autoreceptor for dopamine within the substantia nigra, it seems unlikely that acetylcholinesterase is operating on the same target as dendritically released local dopamine. On the other hand, the actions of acetylcholinesterase were enhanced by low glucose and blocked by the sulfonylurea, tolbutamide. These results strongly suggest that acetylcholinesterase can exert a nonenzymatic action and that this action, in the substantia nigra, is mediated by an ATP-sensitive potassium channel.