Acquired immunity in rats against Angiostrongylus cantonensis infection

Acquired immunity against Angiostrongylus cantonensis was induced by immunizing rats with somatic antigens from fifth-stage larvae and adult worms and live third-stage larvae. Rats immunized twice had significantly fewer worms than rats immunized three times. Fewer worms were recovered from rats immunized with 200 live third-stage larvae than from any other groups. Rats immunized with somatic antigens had higher enzyme-linked immunosorbent assay (ELISA) antibody levels than rats immunized with live larvae. Rats immunized with live third-stage larvae of Angiostrongylus cantonensis were more strongly protected against challenge infections (62-92%) than rats immunized with antigens extracted from fifth-stage larvae (0-30%) and adult worms (11-24%).     

Strongyloides ratti: reversibility of immune damage to adult worms

Transplantation experiments were conducted to assess the reversibility or irreversibility of the damage sustained by Strongyloides ratti during infections in the rat host. Worms of different ages from primary and secondary infections were recovered from their original hosts and transplanted surgically into naive rats. The size and fecundity of normal (Days 6–11 postinfection) worms were maintained after transfer. Damaged worms from primary infection (Days 22–26) showed complete recovery of size and fecundity within 10 days of transfer; damaged worms from a secondary infection (Days 6–7) also showed functional recovery but to a lesser extent. The ultrastructural changes observed mainly in the intestine of damaged worms from primary infections, prior to their transfer, were, however, only partially ameliorated following transplantation into new naive hosts; there was no complete return to structural normality. On the other hand, second infection worms did show almost complete ultrastructural recovery. The course of a transplanted infection established with either damaged or normal worms was similar to infections established percutaneously. Increase in the size of transplanted infections from 100 to 250 worms per recipient did not alter the dynamics of the host/parasite relationship. There was no evidence of adaptation in S. ratti and damaged worms, when transplanted into naive rats, were as successful as normal worms in protecting the host against a subcutaneous larval infection. The implications of this work on the present understanding of the phenomenon of autoinfection in experimental rodent strongyloidiasis are discussed.     

Transplantation of young adult Angiostrongylus cantonensis into the rat pulmonary vessels and its application to the assessment of acquired resistance

Yoshimura K., Aiba H. and Oya H. 1979. Transplantation of young adult Angiostrongylus cantonensis into the rat pulmonary vessels and its application to the assessment of acquired resistance. International Journal for Parasitology9: 97–103. A simple procedure for the transplantation of young adult Angiostrongylus cantonensis into the rat pulmonary vessels was described and its potential usefulness was discussed. Acquired resistance to A. cantonensis was conferred on rats by infection with 30 third-stage larvae or by a similar infection followed by treatment with thiabendazole during 14–19 days postinfection. Nearly comparable levels of resistance to reinfection were also observed in rats that were transplanted with 30 young adult worms, collected from the brain surface of donor animals at 21 and 22 days postinfection. The time-course development of reaginic, hemagglutinating and precipitating antibodies differed in these groups of rats although there appeared to be no positive correlation between the appearance of these antibodies and acquired resistance.     

The influence of Mesocestoides corti on subsequent Angiostrongylus cantonensis infections in mice.

The influence of Mesocestoides corti on subsequent Angiostrongylus cantonensis infection in mice (C57BL/6 and BALB/c) was assessed. Both strains of mice infected with M. corti demonstrated a peak blood eosinophilia at around 3 weeks post-infection (p.i.). C57BL/6 and BALB/c mice primarily infected with M. corti were given A. cantonensis infection 18 days later, but pre-existing M. corti infection did not affect the recovery of intracranial worms of A. cantonensis at day 21 p.i. BALB/c mice with mixed parasite infections showed low morbidity and mortality as compared with mice singly infected with A. cantonensis and some mice demonstrated a pulmonary migration of intracranial worms. In C57BL/6 mice, intracranial worms were killed and thus all mice survived. C57BL/6 mice with mixed parasite infections failed to resist A. cantonensis reinfection. The blastogenic responses of spleen cells against A. cantonensis antigen were lower in BALB/c than in C57BL/6 mice and mixed parasite infections also resulted in less blastogenic responses against both concanavalin A and A. cantonensis antigen than monoinfection. The recovery of M. corti biomass was significantly higher in mice with mixed parasite infections than mice with monoinfection with M. corti. These data suggest a distinct difference in response to A. cantonensis infection between C57BL/6 and BALB/c mice, and the induction of immunosuppression in both mouse strains following M. corti infection. Blood eosinophilia provoked by M. corti infection is not directly associated with the killing of worms in subsequent A. cantonensis infection.     

Transplanted Dipetalonema viteae in the jird: effect of worm burden on parturition rates and microfilaremia

Dipetalonema viteae was studied in the jird, Meriones unguiculatus, to determine the mechanism controlling the level of peripheral microfilaremia. Jirds killed 40 days after infection served as donors of female worms of known age and reproductive status. These worms were transplanted into uninfected jirds and the resultant microfilaremias were monitored. After approximately 100 days, the recipient jirds were killed and 58% of the transplanted worms were recovered alive but depleted of sperm and microfilariae, regardless of the total number implanted in a given host. A direct linear relationship between microfilaremia and the number of recovered adult worms was found. Based on the uniform absence of sperm and microfilariae in the recovered worms it was concluded that female worms, under the conditions of the present study, do not control the peripheral microfilaremia in multi-worm infections through a reduced parturition rate.     

Surface antigens of Angiostrongylus cantonensis developing in permissive and non-permissive hosts

The immunofluorescent antibody test and immunocytochemical method were employed to study the surface antigens of Angiostrongylus cantonensis obtained from infected rats, mice and guinea pigs. Positive results with intense fluorescence and brownish peroxidase staining were observed on the cuticular surface of A. cantonensis recovered from rats 22 days (late cerebral phase) and 34 days (lung phase) post-infection when tested with antisera against host (normal rat serum) antigens as well as crude extracts of A. cantonensis. However, host antigens were not observed on the surface of the nematode recovered from the brain of mice and guinea pigs 15 days post-infection.     

Nerve repair and behavioral recovery following brain transplantation in Notoplana acticola, a polyclad flatworm

Although Notoplana acticola, a marine polyclad, cannot regenerate brain tissue, neuronal repair is rapid. Brains were transplanted into decerebrate flatworms to determine the anatomical patterns and functionality of neural connections established between a new brain and the peripheral nerve network of the recipient animal. Sixty-nine transplants were performed. Four brain transplant orientations were used: normal, reversed, inverted, and reversed inverted. The functionality of the transplanted brains was tested and measured using both behavioral and electrophysiological criteria. Within 23 days, 56% of the transplants that survived and retained the transplants recovered the four behaviors tested: righting behavior, avoidance turning, ditaxic locomotion, and feeding. Nerves exiting the brain tended to join with the peripheral nerves closest to them. Anatomical connections were made within 24 hr of surgery. Some normal behavior was seen within the first 36 hrs after surgery. Control decerebrate worms did not recover behavior. Preliminary intracellular recordings from three types of identified brain sensory interneurons, in transplants, revealed normal electrophysiological properties and this implied that appropriate connections with peripheral sensory cells had been reestablished. Intracellular dye-marking of these neurons in reverse-oriented brains revealed that, although individual nerve processes apparently leave the brain and associate with inappropriate nerve cords, some of the processes turn 180 degrees to reinervate nerve cords, which they normally occupy in unoperated animals. Thus, although anatomical and functional neural connections apparently were made rapidly following brain transplantation, the specificity of the reconnections remains to be shown.     

Transplantation of adult Trichinella spiralis between hosts: worm survival and immunological characteristics of the host--parasite relationship.

A technique for the transplantation of Trichinella spiralis worms directly into the host intestine is described. Infections established by the direct transfer of adult worms were essentially normal both in terms of their survival and reproduction and in their stimulation of, and susceptibility to, host immune responses. Worms transplanted from NIH mouse donors at intervals after infection had an equal ability to survive in the recipient, even when taken from the donor shortly before or during the process of worm expulsion, showing that expulsion does not require worms to be irreversibly damaged. It was noted, however, that after 7 days in the donor the ability of the worm to reproduce in the recipient was temporarily impaired.     

Transplantation of germ cells from rabbits and dogs into mouse testes.

Spermatogonial stem cells of a fertile mouse transplanted into the seminiferous tubules of an infertile mouse can develop spermatogenesis and transmit the donor haplotype to progeny of the recipient mouse. When testis cells from rats or hamsters were transplanted to the testes of immunodeficient mice, complete rat or hamster spermatogenesis occurred in the recipient mouse testes, albeit with lower efficiency for the hamster. The objective of the present study was to investigate the effect of increasing phylogenetic distance between donor and recipient animals on the outcome of spermatogonial transplantation. Testis cells were collected from donor rabbits and dogs and transplanted into testes of immunodeficient recipient mice in which endogenous spermatogenesis had been destroyed. In separate experiments, rabbit or dog testis cells were frozen and stored in liquid nitrogen or cultured for 1 mo before transplantation to mice. Recipient testes were analyzed, using donor-specific polyclonal antibodies, from 1 to >12 mo after transplantation for the presence of donor germ cells. In addition, the presence of canine cells in recipient testes was demonstrated by polymerase chain reaction using primers specific for canine alpha-satellite DNA. Donor germ cells were present in the testes of all but one recipient. Donor germ cells predominantly formed chains and networks of round cells connected by intercellular bridges, but later stages of donor-derived spermatogenesis were not observed. The pattern of colonization after transplantation of cultured cells did not resemble spermatogonial proliferation. These results indicate that fresh and cryopreserved germ cells can colonize the mouse testis but do not differentiate beyond the stage of spermatogonial expansion.     

Biological aspects of limb transplantation: I. Migration of transplanted bone marrow cells into recipient

The transplanted limb contains bone marrow tissue. The hematopoietic cells contained in the bone of the graft normally differentiate after transplantation and can be released to the recipient. The cells migrate to the recipient bone marrow cavities and lymphoid organs. This causes the immune reaction between the donor and the recipient, which develops not only in the graft itself but also in the recipient immune organs where donor bone marrow cells home. The purpose of this study was to investigate the process of migration of the hematopoietic cells from the donor limb to the recipient bone marrow cavities and lymphoid tissues. The questions the authors asked were: what is the rate of release of bone marrow cells from the transplanted bone, where do the released bone marrow cells home in the recipient, how fast are donor bone marrow cells rejected by the recipient, and can some bone marrow cells homing in the recipient tissues survive and create a state of microchimerism. Experiments were performed on Brown Norway and Lewis inbred rat strains (n = 30). Limb donors received intravenous chromium-51-labeled bone marrow cells. Twenty-four hours later, the limb with homing labeled bone marrow cells was transplanted to an allogeneic or syngeneic recipient. The rate of radioactivity of bone marrow cells released from the graft and homing in recipient tissues was measured after another 24 hours. To eliminate factors adversely affecting homing such as the "crowding effect" and allogeneic elimination of bone marrow cells by natural killer cells, total body irradiation and antiasialo-GM1 antiserum were applied to recipients before limb transplantation. In rats surviving with the limb grafts for 7 and 30 days, homing of donor bone marrow cells was studied by specific labeling of donor cells and flow cytometry as well as by detecting donor male Y chromosome. The authors found that transplantation of the limb with bone marrow in its natural spatial relationship with stromal cells and blood perfusion brings about immediate but low-rate release of bone marrow cells and their migration to recipient bone marrow and lymphoid tissues. Large portions of allogeneic bone marrow cells are rapidly destroyed in the mechanism of allogeneic elimination by radioresistant but antiasialo-GM1-sensitive natural killer cells. Some transplanted bone marrow cells remain in the recipient's tissues and create a state of cellular and DNA microchimerism. A low number of physiologically released donor bone marrow cells do not seem to adversely affect the clinical outcome of limb grafting. Quite the opposite, a slight prolongation of the graft survival time was observed.     

Stage-specific antigens of Nematospiroides dubius Baylis, 1926 (Nematoda: Heligmosomides)

Antigens were identified from Nematospiroides dubius recovered from outbred Quackenbush mice between 4 and 10 days postinoculation (PI). Parasite surface proteins were radioiodinated and extracts of the whole worms were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and reacted with normal and immune mouse sera followed by an avidin-biotin-peroxidase assay. Antigens ranged between 250,000 and 20,000 molecular weight (MW). A major surface antigen, 60,000 MW, which appeared to be a complex of different antigens, and a 250,000-MW internal antigen were found on fourth-stage (L4) and fifth-stage (L5) larvae 5-10 days PI but not earlier. A group of minor surface antigens (24,000-30,000 MW) were also expressed as larvae molted from L4 to L5, 6 and 7 days PI, but they differed from antigens of similar MW expressed by adult worms. An antigen, 45,000 MW, was detected in worms 5-10 days PI, but it was only expressed on the surface of L5 worms 9 and 10 days PI. We suggest that the antigen(s) common to adults and larvae may account for protective immunity.     

Mating behaviour of Echinostoma trivolvis and E. paraensei in concurrent infections in hamsters

Young adults of Echinostoma trivolvis and E. paraensei were recovered from hamsters previously infected with metacercarial cysts. Some worms of each species were exposed for 1 h to 3-H-tyrosine to label sperm and transplanted singly to uninfected hamsters with several unlabelled worms of the same or opposite species or both species. After 5 days, recovered worms were processed for paraffin sectioning and autoradiography. The resulting slides were observed for the location of radioactive sperm in the seminal receptacles of donor (labelled) and recipient (unlabelled) worms. When E. trivolvis was the donor with the recipient E. paraensei, self-insemination took place, but only one interspecies mating occurred out of 72 possible recipient worms. When E. paraensei served as the donor, self-insemination again occurred, but no cross-insemination was observed among the 59 E. trivolvis recipient worms. When single donor worms had a choice of either species of recipient worms, no interspecies mating took place, but both self- and cross-insemination occurred in the normal, unrestricted behaviour found in single species mating studies. Rates of both self- and cross-insemination were higher in concurrent infections of both recipient species than in single species mating studies. After transplant, both species localized in their natural habitat within the small intestine, with 1/3 overlapping in the duodenum, making interspecies mating a possibility. The correlation between mating and electrophoretic studies on the genetic relationship between 37-collar-spined echinostomes is discussed.     

Transplantation of Hymenolepis diminuta into naive, immune and irradiated mice.

Almost 100% of 7- to 10-day-old Hymenolepis diminuta became established when surgically transplanted from donor mice into the duodenum of naive recipient mice. Transplanted tapeworms survived 8-12 days, by which time they had survived much longer in total than they would have done in the donor. Mice previously immunized by a primary infection rejected transplants within 4 days. Sub-lethal irradiation (550 rad) delayed rejection by immune mice but such mice still rejected worms much more quickly than did naive mice. Surgery was shown to delay by 2-3 days the rejection of worms by naive mice, and the importance of circumventing surgery by administering the worms per os is emphasized. Prospects for reconstituting irradiated immune mice are considered vis-à-vis work with nematodes, and the differences which, on present knowledge, appear to exist between nematode and cestode rejection are briefly discussed.     

骨髓间质干细胞向大鼠损伤心肌组织的迁移

实验旨在动态观察骨髓间充质干细胞（mesenchymal stem cells,MSCs）向不同微环境下心肌组织的迁移特点,明确组织损伤在干细胞迁移中的作用,为提高干细胞治疗的靶向性和高效性奠定初步试验基础。分离纯化雄性Sprague-Dawley（SD）大鼠的骨髓MSCs,输注入雌性SD大鼠。实验分为4组：正常大鼠＋MSCs移植组,假手术＋MSCs移植组,心肌缺血＋MSCs移植组,心肌缺血对照组（心肌缺血＋培养基移植）。结扎冠状动脉前降支制造心肌缺血模型,将相等数量的雄性MSCs经尾静脉注射移植入前3组雌性大鼠体内,对照组注射等体积培养基,分别于移植后1周及8周取心脏组织标本,采用荧光原位杂交方法（fluorescence in situ hybridization,FISH）检测大鼠Y染色体雄性鉴别基因sty片段的表达,用透射电镜观察大鼠心肌组织超微结构改变。结果发现,移植后1周和8周,正常大鼠移植组和对照组大鼠的心肌组织中均未见sry基因的表达,但假手术移植组和心肌缺血移植组的心肌组织中均可见sty基因的表达,心肌缺血移植组的Y染色体sty基因阳性细胞数量在两个时间点均显著高于假手术移植组（P〈0．01）。分别比较心肌缺血移植组和假手术组在移植后1周和8周的Y染色体sry基因阳性细胞的数量,两个时间点无明显差异。心肌组织的超微结构观察发现心肌缺血移植组大鼠的心肌梗死周边区域可见一些细胞,其形态类似于体外培养的MSCs。研究结果提示MSCs具有向损伤心肌组织迁移的特性,迁移的高峰期可能在组织损伤1周左右,组织损伤及其程度在干细胞迁移中起重要作用。     

Lipids of Angiostrongylus cantonensis (Nematoda: Metastrongyloidea): a comparison between young adults and gravid worms

1. All major classes of lipids were found in the young adults in brain (22 days post-infection) and gravid Angiostrongylus cantonensis in lung of rats (34 days post-infection) comprising approximately 60% of phospholipids, 30% of neutral lipids and the rest, glycolipids. 2. The relative composition of phospholipids were quite similar between worms from the two different habitats, with phosphatidylcholine predominating. The glycolipid profiles were also similar. 3. More neutral lipids in the worms from brain existed as cholesterol and cholesterol esters than those from the lung. More than 20% of the fatty acids in these lipids of the brain were found as C10-C14 acids while oleic acid was the main component in the lung worm.     

Establishment, development and fecundity of Taenia crassiceps in the intestine of prednisolone-treated Mongolian gerbils and inbred mice

Worm establishment, development and fecundity of Taenia crassiceps in the intestine of prednisolone (PTBA)-treated, 15- and 4-week-old Mongolian gerbils and 9-week-old inbred mice of 4 strains (AKR/J, BALB/cAn, B10D2/oSn and C57BL/KsJ) were investigated following oral administration of metacestodes. Gerbils were divided into 5 groups of 4 animals each according to the host age and commencement day of PTBA-treatment (day -13, -7 or 0 relative to infection). Worm recovery from the intestine on day 35 postinfection was not affected by host age, but fewer worms were recovered the earlier the commencement of PTBA-treatment. Worm size, determined by wet weight, total length and proglottis number, correlated inversely with worm burden, suggesting they were affected by the crowding effect. Proglottides were released normally in the faeces but were markedly depressed in all groups except for that of young gerbils. Furthermore, egg production and its development in gravid proglottides were markedly depressed in all groups. In PTBA-treated mice of 4 strains, sexually mature but not gravid worms were recovered from all mice of the AKR/J strain on days 20-32 postinfection, but none or few worms from the intestine of the others. PTBA-treatment did not inhibit all protective host defence mechanism(s) in the unnatural or alternative rodent definitive host of T. crassiceps.     

Donor-recipient interconnections between giant nerve fibers in transplanted ventral nerve cords of earthworms

Twelve segments of ventral nerve cord (VNC) from donor earthworms, Eisenia foetida, were transplanted into recipient worms from which a comparable length of VNC had been removed. Within the first few days after transplantation, bud-like formations, containing outgrowths of the giant nerve fibers, were evident at the ends of transplanted and recipient VNC. Morphological and electrophysiological evidence indicated that by 4-10 days after transplantation, medial (MGF) and lateral (LGF) giant fibers within the transplanted VNC formed cell-specific connections with their counterparts in the recipient VNC. Although the diameters of the giant fiber connections in the transplant-recipient junctions were often larger than normal, spike conduction across the junction was initially slow (approximately 1.0 m/s) but gradually increased over the next 2-3 weeks. Within the transplant, giant fibers were initially normal in appearance, but spike conduction was slow (1-2 m/s). During the next few weeks velocities increased by as much as fourfold and then stabilized for the next several months. However, by 4-5 weeks after transplantation, giant fiber morphology within the transplant was altered significantly, as indicated by the formation of numerous branch-like extensions along the length of each giant fiber. By 9-10 months there were further morphological changes in the transplant, as indicated by decreased branching of the giant fibers and altered neuropile. Despite these morphological changes, through-conduction of giant fiber spikes remained reliable.     

The acquirement of growth ability for third-stage larvae of Strongyloides ratti during the head-passage in the rat

The role of larval passage through the head in the course of the migration of Strongyloides ratti in rats was investigated. Third-stage larvae (L3) recovered from various portions of donor rats were re-injected into the skin, cranial cavity and small intestine of recipient rats to check their ability for further growth. Cultured L3 (L3c) and the L3 recovered from the skin of donor rats (L3s) did not survive in the small intestine after intestinal inoculation. However, intestinal inoculation of L3 recovered from the head of donor rats (L3h) revealed growth to the adult stage. Cultured L3 injected into the cranial cavity of rats also became adult worms in the small intestine. L3 incubated in the cranial cavity for more than 24 h could grow in the small intestine of the recipient rats. These experiments suggest that S. ratti L3 acquire their ability to mature in the small intestine during their migration through the head of rats.     

Restoration of sensory and motor function in earthworm escape reflex pathways following ventral nerve cord transplantation

Twelve segments of earthworm ventral nerve cord (VNC) were excised from either segments 10-22 (i.e., within the MGF sensory field) or segments 75-87 (i.e., within the LGF sensory field) in donor worms and heterotopically, or homotopically, transplanted into recipient animals. Morphological evidence indicated that by four days after transplantation, peripheral connections were formed between the transplanted VNC and the body wall of the recipient, many of these connections involving novel pathways projecting ventrally from the transplant. Restoration of giant fiber touch sensitivity in the transplant occurred from 4-14 days after transplantation. Regardless of the site of transplantation, the restored sensitivity (i.e., MGF versus LGF sensory field) always reflected the origin of the donor VNC. Restoration of MGF-mediated motor activity in the transplant occurred approximately 17-22 days after transplantation. In the case of heterotopic transplants (i.e., anterior VNC into posterior segments), the restored MGF-mediated muscle potentials were facilitating, indicating at least some tendency for persistence of this feature after transplantation. Behavioral observations suggested that reconnections involving other reflex pathways (e.g., those controlling setal movements and peristaltic locomotion) were made within the transplant region and that properties of the restored reflexes reflected those of the donor VNC. The rapid restoration of sensory and motor connections, despite heterotopic placement, indicates a significant capacity for peripheral regeneration by the transplanted VNC. On the other hand, the maintenance of various properties of reflex function, despite heterotopic transplantation, suggests a limited capacity for rearrangement of established central connections in the transplanted VNC.     

Changes in worm burden, haematological and serological response in rats after single and multiple Angiostrongylus cantonensis infections.

Thirteen groups of rats were first sensitized with single or double doses of 5--30 third-stage larvae of Angiostrongylus cantonensis, followed by a challenge infection with 100 larvae at various periods after the primary infection. Seven other groups of rats receiving only the sensitizing infection served as the controls. In all the sensitized rats, a significantly (p less than 0.05) smaller mean number of adult worms was found established in the challenge infection as compared to the control. The frequency of the sensitizing dose and timing of the challenge infection appeared to influence the intensity of the host's response. There was no conclusive evidence to indicate that the immune response could retard the growth, development, or sex ratios of the worms established in subsequent infections. A positive haemagglutinating antibody response was first observed in some rats as early as four weeks post-infection with 100 larvae when the worms began migrating from the brain to the lungs. The antibody response and eosinophilia were most pronounced during the oviposition of the female worms and hatching of first-stage-larvae. Changes in white blood cell, lymphocyte, and neutrophil counts were also followed in some groups.