Systemic hypoxia changes the organ-specific distribution of vascular endothelial growth factor and its receptors

Vascular endothelial growth factor (VEGF) plays a key role in physiological blood vessel formation and pathological angiogenesis such as tumor growth and ischemic diseases. Hypoxia is a potent inducer of VEGF in vitro. Here we demonstrate that VEGF is induced in vivo by exposing mice to systemic hypoxia. VEGF induction was highest in brain, but also occurred in kidney, testis, lung, heart, and liver. In situ hybridization analysis revealed that a distinct subset of cells within a given organ, such as glial cells and neurons in brain, tubular cells in kidney, and Sertoli cells in testis, responded to the hypoxic stimulus with an increase in VEGF expression. Surprisingly, however, other cells at sites of constitutive VEGF expression in normal adult tissues, such as epithelial cells in the choroid plexus and kidney glomeruli, decreased VEGF expression in response to the hypoxic stimulus. Furthermore, in addition to VEGF itself, expression of VEGF receptor-1 (VEGFR-1), but not VEGFR-2, was induced by hypoxia in endothelial cells of lung, heart, brain, kidney, and liver. VEGF itself was never found to be up-regulated in endothelial cells under hypoxic conditions, consistent with its paracrine action during normoxia. Our results show that the response to hypoxia in vivo is differentially regulated at the level of specific cell types or layers in certain organs. In these tissues, up- or down-regulation of VEGF and VEGFR-1 during hypoxia may influence their oxygenation after angiogenesis or modulate vascular permeability.

An Intact Dilysine-like Motif in the Carboxyl Terminus of MAL Is Required for Normal Apical Transport of the Influenza Virus Hemagglutinin Cargo Protein in Epithelial Madin-Darby Canine Kidney Cells

The MAL proteolipid, a component of the integral protein sorting machinery, has been demonstrated as being necessary for normal apical transport of the influenza virus hemagglutinin (HA) and the overall apical membrane proteins in Madin-Darby canine kidney (MDCK) cells. The MAL carboxy terminus ends with the sequence Arg-Trp-Lys-Ser-Ser (RWKSS), which resembles dilysine-based motifs involved in protein sorting. To investigate whether the RWKSS pentapeptide plays a role in modulating the distribution of MAL and/or its function in apical transport, we have expressed MAL proteins with distinct carboxy terminus in MDCK cells whose apical transport was impaired by depletion of endogenous MAL. Apical transport of HA was restored to normal levels by expression of MAL with an intact but not with modified carboxyl terminal sequences bearing mutations that impair the functioning of dilysine-based sorting signals, although all the MAL proteins analyzed incorporated efficiently into lipid rafts. Ultrastructural analysis indicated that compared with MAL bearing an intact RWKSS sequence, a mutant with lysine −3 substituted by serine showed a twofold increased presence in clathrin-coated cytoplasmic structures and a reduced expression on the plasma membrane. These results indicate that the carboxyl-terminal RWKSS sequence modulates the distribution of MAL in clathrin-coated elements and is necessary for HA transport to the apical surface.

In vivo tissue distribution of CD4 lymphocytes in mice determined by radioimmunoscintigraphy with an 111In-labeled anti-CD4 monoclonal antibody.

The tissue distribution of CD4 lymphocytes in normal C57/BL mice and CD4 knockout mice was determined by biodistribution measurements and gamma camera imaging with an 111In-labeled rat IgG2b monoclonal antibody directed against the murine CD-4 antigen. In normal mice high concentrations of antibody accumulated in the spleen and lymph nodes. At 45 hr after injection, the concentration of radiolabel in the spleen and lymph nodes of normal mice were 10- to 20-fold greater than in the corresponding tissue of the CD4 knockout mice and nonlymphoid tissues of both types of mice. At 24 and 45 hr, gamma camera images showed high concentrations of radiolabeled antibody in lymph node and spleen of normal but not knockout mice. These results indicate that radioimmunoscintigraphy with 111In-anti-CD4 is an excellent method for studying tissue distribution of CD lymphocytes in mice. Using an equivalent anti-human CD antibody, this method might be useful for studying the pathophysiology of conditions in which these cells play a critical role and for monitoring therapies for these disorders.

On signal sequence polymorphisms and diseases of distribution.

We report a previously unappreciated property of the signals that target organelle-specific proteins to their subcellular sites of action. Such targeting sequences are shown to be polymorphic. We discovered this polymorphism when we cloned the mitochondrial manganese-containing superoxide dismutase from cell lines of normal individuals and patients with genetic diseases of premature aging and compared their sequences to each other and to those previously reported. The polymorphism consists of a single nucleotide change in the region of the DNA that encodes the signal sequence such that either an alanine or valine is present. Subsequently, eight cell lines were analyzed and all three possible combinations of the two signal sequences were observed. Such signal sequence polymorphisms could result in diseases of distribution, where essential proteins are not properly targeted, thereby leading to absolute or relative deficiencies of critical enzymes within specific cellular compartments. Progeria and related syndromes may be diseases of distribution.

Distribution of endogenous type B and type D sheep retrovirus sequences in ungulates and other mammals.

The jaagsiekte sheep retrovirus (JSRV), which appears to be a type B/D retrovirus chimera, has been incriminated as the cause of ovine pulmonary carcinoma. Recent studies suggest that the sequences related to this virus are found in the genomes of normal sheep and goats. To learn whether there are breeds of sheep that lack the endogenous viral sequences and to study their distribution among other groups of mammals, we surveyed several domestic sheep and goat breeds, other ungulates, and various mammal groups for sequences related to JSRV. Probes prepared from the envelope (SU) region of JSRV and the capsid (CA) region of a Peruvian type D virus related to JSRV were used in Southern blot hybridization with genomic DNA followed by low- and high-stringency washes. Fifteen to 20 CA and SU bands were found in all members of the 13 breeds of domestic sheep and 6 breeds of goats tested. There were similar findings in 6 wild Ovis and Capra genera. Within 22 other genera of Bovidae including domestic cattle, and 7 other families of Artiodactyla including Cervidae, there were usually a few CA or SU bands at low stringency and rare bands at high stringency. Among 16 phylogenetically distant genera, there were generally fewer bands hybridizing with either probe. These results reveal wide-spread phylogenetic distribution of endogenous type B and type D retroviral sequences related to JSRV among mammals and argue for further investigation of their potential role in disease.

Immunolocalization of estrogen receptor protein in the mouse blastocyst during normal and delayed implantation.

We previously showed that estrogen receptor (ER) mRNA is present in preimplantation mouse embryos. The apparent synthesis of ER mRNA by the blastocyst at the time of implantation when estrogen is required was of special interest. A demonstration of the presence of ER protein would support the idea that estrogen can act directly on the embryo. The mouse embryo at the blastocyst stage is differentiated into two cell types, the trophectoderm and the inner cell mass. To determine whether ER mRNA is translated into ER protein and its cell-specific distribution, immunocytochemical analyses were performed in mouse blastocysts. ER protein was detected in all cell types of the normal, dormant, or activated blastocyst. To trace the fate of ER in these cell types, immunocytochemistry was performed in implanting blastocysts and early egg cylinder stage embryos developed in culture. Again, ER was detected in all cells of the implanting blastocyst. At the early egg cylinder stage, continued expression of ER was observed in cells derived from the inner cell mass or the trophoblast. In trophoblast giant cells, ER was concentrated in small regions of the nucleus, possibly the nucleoli, which was similar to that observed in dormant and activated blastocysts. The embryonic expression of ER at such early stages in a broad array of cells suggests that ER may have a general role during early development.

Repair of x-ray-induced DNA double-strand breaks in specific Not I restriction fragments in human fibroblasts: joining of correct and incorrect ends.

An assay that allows measurement of absolute induction frequencies for DNA double-strand breaks (dsbs) in defined regions of the genome and that quantitates rejoining of correct DNA ends has been used to study repair of dsbs in normal human fibroblasts after x-irradiation. The approach involves hybridization of single-copy DNA probes to Not I restriction fragments separated according to size by pulsed-field gel electrophoresis. Induction of dsbs is quantitated from the decrease in the intensity of the hybridizing restriction fragment and an accumulation of a smear below the band. Rejoining of dsbs results in reconstitution of the intact restriction fragment only if correct DNA ends are joined. By comparing results from this technique with results from a conventional electrophoresis assay that detects all rejoining events, it is possible to quantitate the misrejoining frequency. Three Not I fragments on the long arm of chromosome 21 were investigated with regard to dsb induction, yielding an identical induction rate of 5.8 X 10(-3) break per megabase pair per Gy. Correct dsb rejoining was measured for two of these Not I fragments after initial doses of 80 and 160 Gy. The misrejoining frequency was about 25% for both fragments and was independent of dose. This result appears to be representative for the whole genome as shown by analysis of the entire Not I fragment distribution. The correct rejoining events primarily occurred within the first 2 h, while the misrejoining kinetics included a much slower component, with about half of the events occurring between 2 and 24 h. These misrejoining kinetics are similar to those previously reported for production of exchange aberrations in interphase chromosomes.

Distribution of parthenogenetic cells in the mouse brain and their influence on brain development and behavior.

A systematic analysis of parthenogenetic (PG) cell fate within the central nervous system (CNS) was made throughout fetal development and neonatal and adult life. Chimeras were made between PG embryos carrying a ubiquitously expressed lacZ transgene and normal fertilized embryos. After detailed histological analysis, we find that the developmental potential of PG cells is spatially restricted to certain parts of the brain. PG cells are prevalent in telencephalic structures and are largely excluded from diencephalic structures, especially the hypothalamus. These spatial restrictions are established early in development. Behavioral studies with chimeras identified an increase in male aggression when the proportion of PG cells in the brain was high. These studies demonstrate that imprinted genes play key roles in development of the CNS and may be involved in behavior.

Normal development and function of natural killer cells in CD3 epsilon delta 5/delta 5 mutant mice.

The CD3 epsilon polypeptide contributes to the cell surface display as well as to the signal transduction properties of the T-cell antigen receptor complex. Intriguingly, the distribution of CD3 epsilon is not restricted to T cells, since activated mouse, human, and avian natural killer (NK) cells do express intracytoplasmic CD3 epsilon polypeptides. CD3 epsilon is also present in the cytoplasm of fetal thymic T/NK bipotential progenitor cells, suggesting that it constitutes a component of the NK differentiation program. We report here that the genetic disruption of CD3 epsilon exon 5 alters neither NK cell development nor in vitro and in vivo NK functions, although it profoundly blocked T-cell development. These results support the notion that CD3 epsilon is dispensable for mouse NK cell ontogeny and function and further suggest that the common NK/T-cell progenitor cell utilizes CD3 epsilon as a mandatory component only when differentiating toward the T-cell lineage.

Análise dos resultados do ensaio CBR realizado em condições variadas de umidade pós-compactação

O objetivo deste trabalho é avaliar a influência da variação do teor de umidade no valor do índice de suporte CBR de uma brita estabilizada granulometricamente e de seis solos coletados em rodovias do interior do Estado de São Paulo. Os seis solos estudados foram divididos em três pares, com curvas granulométricas próximas, mesma classificação HRB e USCS, sendo cada par formado por um solo de comportamento laterítico e um solo de comportamento não-laterítico. Os corpos-de-prova foram moldados na umidade ótima e massa específica seca máxima obtidos com os resultados do ensaio de Proctor normal. Os materiais foram analisados sob quatro condições de umidade: sem imersão, após alcançar metade da umidade de estabilização, após quatro e oito dias de imersão. Observou-se a distribuição da umidade interna dos corpos-de-prova para cada uma das situações. Foi analisada a influência do envelhecimento dos materiais solto e compactado por quatro dias e sem imersão, nos resultados dos ensaios de CBR. Com os resultados pôde-se correlacionar a influência da granulometria e da gênese dos materiais nos resultados dos ensaios de índice de suporte CBR nas diversas condições de umidade avaliadas.

Quantitative composition and distribution of the silt fraction in surface sediments of the North-West African continental margin

Surface sediments from the continental slope and rise of North-West Africa between the Canary lslands and the Cape Verde Islands are mainly composed of silt-sized material (2-63 µm). A number of sampling profiles were run normal to the coast and the composition of the silt fraction was determined quantitatively by scanning electron microscope analysis. The carbonate portion of the sediment was found to be nearly exclusively of biogenic origin. The most important contributors are planktonic foraminifers and coccoliths with minor contributions derived from pteropods. Plankton-produced biogenic opal such as diatoms and radiolarians play a very minor role. The high production rates of opal-silica plankton which exists in the surface waters of the NW-African upwelling system does not give rise to corresponding increases of opal accumulation in the bottom sediment. Benthic producers consist mainly of foraminifers and molluscs but the entire input from benthic producers is extremely small. An exception to this occurs in the prodelta sediments of the Senegal river. Downslope particle transport is indicated by the occurrence of shallow-water coralline algae, ascidian sclerites and cliona boring chips and can be traced as far down as the continental rise. The non-carbonate silt fraction mostly consists of quartz which is derived as eolian dust from the Sahara desert by the Harmattan and the NE-Trade-wind system. The percentage of carbonate in the surface sediments directly indicates the relative proportions of autochthonous biogenic components and terrigenous allochthonous quartz particles.

Some parameters of distribution of hydrocarbon paraffin fraction in bottom sediments of the Kara Sea and the Yenisey estuary

During Cruise 49 of R/V Dmitry Mendeleev in the Kara Sea (August-September, 1993) chemical-bitumenological studies of bottom sediments were carried out. Hydrocarbons were analyzed by gas-liquid chromatography. It was found on the basis of distribution of n-alkanes and isoprenoids (pristan and phytan) that organic matter is mainly terrigenous consisting of higher plant remains.

(Figure 2) Distribution of planktonic foraminifera in the upper Paleocene of DSDP Hole 93-605

Good faunal preservation in the upper part of the Planorotatites pseudomenardii Zone at Deep Sea Drilling Project Site 605, northwestern Atlantic, allows a biometric analysis of the upper Paleocene planktonic foraminiferal species Planorotatites pseudomenardii (Belli), a keeled species that probably developed from a middle Paleocene unkeeled Planorotalites form. Multivariate analysis shows a consistent separation of all Planorotatites specimens into two groups, which are differentiated by the presence or absence of a complete keel; other variables are only of minor importance. The keeled group coincides with P. pseudomenardii. We recognize only one unkeeled species, Planorotalites chapmani (Parr), with Planorotalites ehrenbergi (Bolli), Planorotalites imitata (Subbotina), Planorotalites planoconica (Subbotina), Planorotalites troelseni (Loeblich and Tappan), and Planorotalites hausbergensis (Gohrbrandt) as junior synonyms. P. chapmani ranges from the middle Paleocene to at least the top of the upper Paleocene. The morphology of P. pseudomenardii does not change significantly, and although the frequency of Planorotalites is variable, the proportion of P. pseudomenardii to all Planorotalites varies only slightly around 65% in the upper two-thirds of its range at Site 605. However, in the top 1.5 m of its range the proportion of P. pseudomenardii decreases; in the same section, all Planorotalites specimens show a reduction in the size of their tests, suggesting that a temporary change in environmental conditions led to the exit of P. pseudomenardii\ in Magnetozone C24R at Site 605-apparently higher than expected from current standard zonations. Unkeeled Planorotalites, in contrast to R. pseudomenardii, persisted and regained normal size. The entry of P. pseudomenardii at Site 605 cannot be described in the same detail because of low frequencies of Planorotalites specimens and an erratic distribution of P. pseudomenardii in the lower part of its range. Many of the washed residues of the samples from these sediments are dominated by radiolarians, and the poorly preserved foraminiferal faunas may have abundant benthics, indicating carbonate dissolution. The initially low frequencies of P pseudomenardii relative to the unkeeled Planorotalites show a strong negative correlation with the total amount of radiolarians per sample and could be the result of preferential preservation, as well as of the same environmental conditions that caused the abundance of radiolarians.

Locally asymptotically normal families of distributions; certain approximations to families of distributions and their use in the theory of estimation and testing hypotheses.

Bibliography: p. 98.

The natural history of plants, their forms, growth, reproduction, and distribution /

Plates numb. consecutively; Plates II, III, V, VI, IX, XII-XVI have outline diagrams on guard sheets.