X-ray-induced P-selectin localization to the lumen of tumor blood vessels

P-selectin is a cell adhesion molecule that is sequestered in Weibel-Palade storage reservoirs within the vascular endothelium and alpha granules in platelets. P-selectin is rapidly translocated to the vascular lumen after tissue injury to initiate the adhesion and activation of platelets and leukocytes. We studied the histological pattern of P-selectin expression in irradiated tumor blood vessels. We observed that P-selectin was localized within the endothelium of tumor vessels prior to treatment. At 1-6 h following irradiation, P-selectin was mobilized to the lumen of blood vessels. To determine whether radiation-induced vascular lumen localization of P-selectin was tumor type specific or species specific, we studied tumors in rats, C3H mice, C57BL6 mice, and nude mice. P-selectin localization to the vascular lumen was present in all tumors and all species studied. Irradiated intracranial gliomas showed P-selectin localization to the vascular lumen within 1 h, whereas blood vessels in normal brain showed no P-selectin staining in the endothelium and no localization to the irradiated vascular lumen. Radiation-induced P-selectin localization to the vascular lumen increased in time-dependent manner, until 24 h after irradiation. P-selectin in platelets may account for the time-dependent increase in staining within the vascular lumen after irradiation. We therefore used immunohistochemistry for platelet antigen GP-IIIa to differentiate between endothelial and platelet localization of P-selectin. We found that GP-IIIa staining was not present at 1 h after irradiation, but it increased at 6 and 24 h. P-selectin localization to the vascular lumen at 6-24 h was, in part, associated with platelet aggregation. These findings indicate that radiation-induced P-selectin staining in the vascular lumen of neoplasms is associated with aggregation of platelets. Radiation-induced localization of P-selectin to the vascular lumen is specific to the microvasculature of malignant gliomas and is not present in the blood vessels of the irradiated normal brain.     

The nature of the development of autoimmune hemolytic anemia in thymectomized animals exposed to the action of ionizing radiation

To study the character of the experimental autoimmune haemolytic anaemia formation in thymectomized mice, which were irradiated in 1.0 Gy dose. Shown, autoimmune process is result in most aggressive course in thymectomized irradiated mice with autoimmune anaemia than in irradiated mice with autoimmune anaemia. This pathology was connected with decrease of the T-suppressor function. Via 20th weeks the autoimmune process was practically depressed.     

Morphological study of endothelial cells in the human fetus during early period of gestation

During the formation of blood vessels, proliferation and migration of endothelial cells (ECs) are one of the most important mechanisms. The development of the vessels involves two different mechanisms: vasculogenesis i.e. differentiation of ECs in situ from mesenchymal precursors, and angiogenesis i.e. sprouting of ECs from pre-existing vessels. The frontal lobes from 20 brains of human fetuses ranging from 8 to 17 weeks of gestation (GW) were subjected to correlative light and electron microscopic studies. Our observations demonstrate the sprouting of ECs from pre-existing vessels in leptomeninges (angiogenesis) during the formation of a capillary network of the fetal human brain. In addition, the study did not reveal after the 8th GW the mitotic proliferation of ECs in examined specimens which allows to underline the importance of sprouting and elongation of ECs channels for this phase of vascularization of cerebral hemispheres.     

Microvascular system of the rat incisor enamel organ. A scanning electron microscopic study of vascular corrosion casts

The rat incisor is a commonly used model in studies of tooth eruption, amelogenesis and effects of mechanical loading on the dental and periodontal tissues. The purpose of this study was to assess the three-dimensional architecture of the microvascular bed of the rat incisor enamel organ, to describe the direction of blood flow, and to provide a histometric assessment of the vascular categories that can be statistically analyzed. Vascular corrosion casts were prepared and examined by scanning electron microscopy. The microvasculature of the labial periodontal space was arranged in three distinct layers. The inner layer in direct relation to the enamel organ consisted of a capillary network which was drained by short venules at the cemento-enamel junction. The intermediate layer consisted of arterioles oriented parallel to the long axis of the incisor mainly mid-labially, branching off smaller arterioles to the capillary network. The outer layer was formed by flattened sinusoid vessels of larger caliber. Blood supply was from the anterior superior alveolar artery branches through the arterioles into the capillary network. Drainage was postero-laterally along the cemento-enamel junction via short venules which emptied into the sinusoid vessels, finally to flow through Volkmann's canals into the alveolar bone via small venules. The findings demonstrate that the microvasculature of the rat incisor enamel organ has an exceptionally high level of physiologically-adapted structural organization.     

The protein efficiency ratios of 30:70 mixtures of animal:vegetable protein are similar or higher than those of the animal foods alone

C57BL/6 mice receiving pretransplant immunization with C3H.SW spleen cells via the portal vein, but not the vena cava, show Ag-specific delayed rejection of allogeneic C3H.SW skin grafts. This delayed rejection is not seen if preimmunization is performed in gamma delta TCR knockout (C57BL/6-Tcrdtm1Mom) mice. gamma delta TCR+ and alpha beta TCR+ hybridoma cells were prepared from Peyer's patch cells harvested from C57BL/6 mice 4 days following portal venous immunization with 100 x 10(6) irradiated C3H.SW spleen cells and skin grafting with C3H.SW tail skin. After recloning, these hybridoma cells were tested for cytokine production in vitro following restimulation with irradiated C3H.SW spleen cells and for their ability to delay rejection of C3H.SW skin grafts after adoptive transfer to C57BL/6 mice. Delayed graft rejection was a function of cells that showed preferential production of IL-10, not IFN-gamma, in vitro, independent of the source (vena cava or portal vein immunized mice) or the TCR phenotype of the hybridoma. Simultaneous infusion of anti-IL-10 mAb abolished this graft prolongation effect of transferred gamma delta TCR+ hybridomas. Hybridoma cells producing IL-10 on restimulation could polarize cytokine production from freshly stimulated mesenteric lymph node away from production of IL-2 and IFN-gamma, and toward IL-4, IL-10, and TGF-beta production. This immunoregulation by hybridoma cells in vivo and in vitro was observed even for third party Ag-stimulated mice/cells as long as the hybridoma cells themselves received stimulation with their specific Ag.     

Properties and relationships of broad bean stain virus and Echtes Ackerbohnenmosaik-Virus

The effects of feeding irradiated wheat in mice on bone marrow and testis chromosomes, germ cell numbers and dominant lethal mutations were investigated. Feeding of freshly irradiated wheat resulted in significantly increased incidence of polyploid cells in bone marrow, aneuploid cells in testis, reduction in number of spermatogonia of types A, B and resting primary spermatocytes as well as a higher mutagenic index. Such a response was not observed when mice were fed stored irradiated wheat. Also there was no difference between the mice fed un-irradiated wheat and stored irradiated wheat.     

Effect of heme and vacuole deficiency on FRE1 gene expression and ferrireductase activity in Saccharomyces cerevisiae

Doppler sonography of the fetal descending aorta, renal artery, middle cerebral artery and umbilical artery in a population of 74 fetuses with a abdominal circumference below the fifth percentile of the reference limits were done. All fetuses were free from structural and chromosomal abnormalities. The pulsatility- and the resistance indices as well as the ratios between these indices from peripheral and cerebral vessels were calculated and correlated to the fetal distress. The measurement of the pulsatility index of the middle cerebral artery provided the best results in predicting the development of fetal distress. Better results were achieved by the use of ratios of pulsatility-indices of various vessels than by the examination of the vessels alone. Our results suggest the usefulness of the examination of the middle cerebral artery and their ratios compared to the renal artery.     

Functional and morphological damage of endothelium in rabbit ear artery following irradiation with cobalt60

1. The relaxant actions of acetylcholine and A23187 were examined in the rabbit central ear artery at different intervals following exposure to different doses of radiation with a cobalt60 unit. The artery was irradiated with a dose of 10 Gy, 20 Gy and 45 Gy. Radiation caused dose- and time-dependent impairment of the endothelium-dependent relaxations. The impaired endothelium-dependent relaxations occurred as early as 1 week postirradiation and persisted throughout the experimental period (10 weeks). 2. The endothelium-independent response to sodium nitroprusside was well preserved up to 6 weeks after irradiation. The contractile response to noradrenaline was unaltered by irradiation throughout the experimental period, but in contrast to control vessels, an increase in the sensitivity to noradrenaline in the presence of the nitric oxide synthase (NOS) inhibitor N(G)-nitro-L-arginine was not observed in the irradiated vessels. 3. The impaired endothelium-dependent relaxations in the irradiated vessels were not improved by pretreatment with the NOS substrate L-arginine, the cyclo-oxygenase inhibitor indomethacin or the free radical scavengers superoxide dismutase and catalase. 4. Scanning electron microscopy indicated morphologically intact endothelial cells within the first 4 weeks after irradiation. 5. Western blot analysis showed a significant decrease in the expression of endothelial NOS (eNOS) in the irradiated vessels. 6. These data indicate that endothelial cell function is specifically impaired in the irradiated vessels before morphological endothelial cell damage can be detected. This impairment may be related to diminished eNOS expression.     

Evidence for the involvement of a bone marrow-derived cell population in the immune expulsion of Trichinella spiralis

When mice were irradiated immediately before infection with Trichinella spiralis there was a profound and long-lasting interference with their ability to expel adult worms from the intestine. Irradiation given after the fifth day of infection was progressively less effective in this respect. The ability to expel worms was not restored when mesenteric lymph node cells (MLNC) were transferred (a) on the day of infection in mice irradiated one day previously, or (b) on day 7 of an infection in mice irradiated on day 6, even though the MLNC transferred immunity to intact recipients. Transfer of bone marrow (BM) alone was also without effect. However, worm expulsion was restored if, following irradiation and injection of BM, 10 days were allowed for BM differentiation before transfer of MLNC. This restoration was effective even after lethal levels of irradiation and was clearly dependent upon a donor-derived BM component cooperating with, or responding to, the activity of the transferred MLNC. The possibility that the BM component is non-lymphoid in nature is discussed.     

Mechanism of increasing dystrophin-positive myofibers by myoblast transplantation: study using mdx/beta-galactosidase transgenic mice

Female mdx/mdx mice were crossed with non-dystrophic transgenic males expressing the beta-galactosidase (beta-gal) gene under a muscle-specific promoter (TnILacZ1/29). All male offspring were mdx mice and about 50% of them also expressed the beta-gal gene. The beta-gal/mdx mice were selected as recipients for the transplantation of myoblasts from non-transgenic normal BALB/c mice. When host muscles were not irradiated before myoblast transplantation, 4.6% of the muscle fibers in host muscles were dystrophin positive 1 month after transplantation. Most of these dystrophin-positive muscle fibers were also beta-gal positive. About one quarter of these fibers are the result of reverse mutations; most of them have, however, been produced by fusion of donor myoblasts with host muscle fibers or with host myoblasts. The virtual absence of beta-gal-negative fibers indicates that there were no exclusively donor-donor fusions. When host muscles were irradiated before myoblast transplantation, roughly the same percentage (5.5%) of dystrophin-positive fibers were formed in the injected muscle, but 42% of them were beta-gal negative. These beta-gal-negative dystrophin-positive muscle fibers were formed by the exclusive fusion of donor myoblasts with one another rather than with host cells. This clearly indicates that myoblast transplantation can form completely new muscle fibers or muscle fiber segments when host satellite cell proliferation is reduced by irradiation. These newly formed muscle fibers had, however, a small diameter and additional myoblast transplantation may be required to increase their size. This situation has some similarities with findings in Duchenne muscular dystrophy patients of more than 6 years of age, who also have a limited proliferation capacity of their satellite cells.     

Mechanism of action of benzene on hematopoiesis (a study of hematopoietic stem cells)

A study was made of the effect of benzol on the colony-forming activity in the spleen and the bone marrow of mice in 4 different experimental variants. In benzol hypoplasia of hemopoiesis there was a decrease in the number of the CFU in the hemopoietic organs without any changes in the distribution of their cell types. Incubation of the cells of normal mouse bone marrow with benzol did not induce any decrease of the colony-forming efficiency. Administration of benzol to the lethally irradiated mice after the transplantation of normal bone marrow suspension led to a sharp decrease in the number of CFU with a preponderant depression of the granulocytic colony formation. These data afford to suppose the change in the properties of microenvironment of the hemopoietic stem cells in experimental benzol poisoning.     

Mast cell numbers and passive cutaneous anaphylaxis in irradiated mice

IgE antibody levels in irradiated mice immunized with primed spleen cells are significantly higher than those measured in actively immunized intact mice. The possible role of mast cells in this elevated IgE response was assessed by the enumeration of mast cells and by determining their capacity to give rise to passive cutaneous anaphylactic (PCA) reactions in the skin of irradiated mice. The number of mast cells was found to be not significantly depleted over a period of 11 days by the irradiation doses used. Furthermore, the capacity of these mast cells to give PCA reactions was unimpaired. Radiation effects on mast cells in adoptively immunized mice therefore do not contribute to the high IgE levels observed in such animals.     

Helicobacter pylori, acid, and omeprazole revisited: bacterial eradication and rebound hypersecretion

Mouse blastocysts were exposed for 24 h to various concentrations of recombinant mouse tumor necrosis factor alpha (TNFalpha) and observed for their capacity to implant in vitro on a fibronectin-coated substrate or to develop in vivo after their transfer into surrogate females. Compared with findings in control blastocysts, exposure to TNFalpha resulted in a significant reduction in the average number of cells in the inner cell mass (ICM) lineage. This effect was associated with a significant increase in the frequency of cells identified as engaged in apoptosis by means of the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling technique. No difference was found in the incidence of nuclear fragmentation between control and TNFalpha-exposed blastocysts. When TNFalpha-pretreated blastocysts were allowed to implant in vitro, significantly fewer embryos were able to maintain a structured ICM cluster at the center of the trophectoderm outgrowth. Although no difference was found in the average surface area of the outgrowths, implants derived from TNFalpha-treated blastocysts contained significantly fewer nuclei than implants from control embryos. After transfer into recipient mice, TNFalpha-pretreated blastocysts implanted at about the same rate as control embryos, but a significantly higher rate of resorption was found among fetuses after exposure to the cytokine. In addition, the weight of the surviving fetuses was significantly lower than for control fetuses. These data indicate that the impact of TNFalpha on blastocysts is specifically aimed at the ICM lineage and that TNFalpha decreases the ability of embryos to differentiate into fetuses after implantation.     

The diagnostic value of serum lysozyme activity in inflammatory bowel disease

The inferior alveolar nerve was cut or the vessels entering the mandibular canal were damaged in 39 guinea pigs at 10 days of age. Possible changes in the incisors were recorded during a 100-day observation period whereafter the animals were killed and the skulls freed of soft tissues; at this time, 35 skulls were found suitable for further examination. The changes in the mandibular incisors on the operated side became observable 30-40 days post-operatively as discolourization, a 'trauma line' around the tooth, narrowing of the tooth or total loss of it's visible part. In the guinea pigs with the nerve cut (group A), such signs of maldevelopment were seen in 5 out of 19 animals, whereas the corresponding figure for the animals with the damaged vessels (group B) was 14/16. All incisors in group A and 13 out of 14 in group B recovered from the state of maldevelopment; one animal in group B showed complete loss of the incisor. The molar region was examined only from the cleaned skulls. In group A, the first molar was deformed in two animals; in group B, seven animals displayed either maldevelopment of the first molar or the first and second molar or complete loss of the first molar. No malformation of the third molar was observed. With two exceptions, the deformation or loss of molars occurred in combination with incisor maldevelopment. The malformations were considered to result from a disturbed blood supply. In group A, the dental changes may perhaps be ascribed to a short-term interference with the blood supply during the operation. The variable results in group B seem to imply that the mandibular teeth receive at least part of their blood supply from vessels besides those entering the mandibular canal. The innervation appears to have a subordinate role in the restoring of the morphology of continuously erupting teeth.     

Tissue preference and differentiation of malignant rat x mouse hybrid cells in chimaeric mouse fetuses

Mouse teratocarcinoma cells (OTT6050) deficient for thymidine kinase were fused with rat hepatoma cells ( Fu5AH ) deficient for hypoxanthine phosphoribosyltransferase using inactivated Sendai virus. The hybrid cells were selected and cultured in the presence of HAT medium. A clonally established hybrid cell line ( As3 ), which in addition to its mouse genome contains several rat chromosomes, expresses rat specific enzyme variants and produces large primarily undifferentiated tumors, with some hepatoma characteristics in athymic nude mice. To reveal the in vivo developmental potential of these cells and to determine whether, under different experimental conditions, they are capable of participating in tissue differentiation, the As3 cells were injected into mouse blastocysts from the C57BL/6 strain. The experimental blastocysts were then transferred into the uteri of pseudopregnant foster mothers to allow further development. From a total of 212 blastocysts transplanted, 61 fetuses developed and were analysed for As3 contributions between the 10th and 18th day of gestation. Four fetuses at day 18 showed hybrid cell participation in their livers and a few organs of only endo-mesodermal origin, as judged from the presence of rat-specific enzyme variants. The enzymes were organ-specifically expressed (e.g., lactate dehydrogenase) or appeared newly during in situ differentiation while being absent in the original hybrid cells (e.g., glycerol-3-phosphate dehydrogenase). During short in vitro culture of the chimaeric organs, it was possible to select for the hybrid cells which reverted to an enzyme pattern simiar to but not identical with the As3 cell line and different to that observed in situ.(ABSTRACT TRUNCATED AT 250 WORDS)     

Microsatellite in the beta-tubulin gene of Toxoplasma gondii as a new genetic marker for use in direct screening of amniotic fluids

To examine the correlation between Toxoplasma gondii genotype and congenital human toxoplasmosis, the polymorphism of the microsatellite consisting of a dinucleotide (TG) repeat in the intron of the beta-tubulin gene was investigated by PCR. Thirty-four reference strains were studied, including 7 strains virulent in mice and 27 strains avirulent in mice. The seven virulent strains had a (TG)8 microsatellite, and the avirulent strains had a (TG)7 microsatellite. This confirms the dichotomy already observed for virulent and avirulent strains. Additionally, 37 samples of amniotic fluid from infected fetuses were tested. All of them had the (TG)7 microsatellite marker. This result confirms that most of the human cases of congenital toxoplasmosis are due to strains avirulent in mice. Nevertheless, their virulence in human fetuses was obvious, as numerous abnormalities were observed on ultrasonic examination. The new genetic marker is the first one directly used for typing T. gondii isolates without any bias due to cultivation of the parasite. This microsatellite marker is not sufficient to type the strains which are avirulent in mice; however, seeking more polymorphic microsatellites should be worthwhile to obtain new genetic markers for direct screening of biological samples.     

Ultrastructural study on the hypothalamic-hypophysial-adrenal axis in fetal rats

The adrenal glands of decapitated and encephalectomized fetal rats were investigated electron microscopically and compared to those of normal intact fetal rats. Although the adrenal cortices did not show three zones (zona glomerulosa, fasciculata, and reticularis) on the 16.5th day of gestation when the decapitation or encephalectomy was carried out in utero, the zonation was recognized in fetuses operated on the 21.5th day of gestation. The same was true for normal control fetuses. However, cytoplasmic characteristics suggesting steroidogenesis in the cortical cells were reduced to various degrees in the encephalectomized or decapitated fetuses, especially in the latter ones. The change in cytoplasmic appearance was more conspicuous in the inner portion of the cortex. This result suggests that for the maintenance of normal adrenocortical function the hypothalamus may be indispensable even during the prenatal life of rats.     

Anatomical variations in the articular masses of the seventh cervical vertebra simulating fracture

Rats were exposed to gamma rays or helium ions on one of days 4-9 of gestation. Embryonic survival was recorded as a function of intrauterine position at autopsy on day 20 of gestation. Embryos located at the ovarian and cervical ends of the uterus experienced higher rates of mortality than did their littermates located at the middle of the uterine horn. This effect was observed in litters exposed to both radiation modalities on all days studied. The influence of intrauterine position on embryonic survival was directly proportional to radiation dose and to the number of fetuses occupying the uterus horn. Under the least advantageous conditions (i.e., a crowded uterine horn exposed to a moderately high radiation dose), the cervical embryo's probability of survival was less than half that of the litter as a whole. A disproportionately high rate of embryonic mortality at the cervical position was also observed in litters irradiated under hypoxic conditions, suggesting that the non-random distribution of radiation effect was not the result of variations in oxygen concentration within the uterus. In contrast, there was no indication that intrauterine position influenced the distribution of gross morphologic abnormalities in irradiated litters.     

Rabbit coronary vessels following a single irradiation of the heart region with a dose of 2400 rad

The effect of ionizing radiation on the coronary vessels was studied on rabbits whose cardiac areas were irradiated at a dose of 2400 rads. This exposure led to dilatation and stricture of small vessels. The most significant changes were found in the system of the left coronary artery at an early post-irradiation period.