Antibodies against major histocompatibility complex class II antigens directly inhibit the growth of T cells infected with Theileria parva without affecting their state of activation

We have analyzed the effect of antibodies (Abs) directed against major histocompatibility complex (MHC) class II Abs on the proliferation of Theileria parva-infected (Tpi) T cells. Anti-MHC class II Abs exert a direct effect on Tpi T cells causing an acute block in their proliferation. The inhibition does not involve apoptosis and is also entirely reversible. The rapid arrest of DNA synthesis caused by anti-MHC class II Abs is not due to interference with the state of activation of the T cells since the transcriptional activator NF-kappa B remains activated in arrested cells. In addition, interleukin 2 (IL-2), IL-2R, and c-myc gene expression are also unaffected. By analyzing the cell-cycle phase distribution of inhibited cells, it could be shown that cells in all phases of the cell cycle are inhibited. The signal transduction pathway that results in inhibition was shown to be independent of protein kinase C and extracellular Ca2+. Tyrosine kinase inhibitors, however, partly reduced the level of inhibition and, conversely, phosphatase inhibitors enhanced it. The possible relevance of this phenomenon in other systems is discussed.

The characterization of PKC-isozyme specific EGFR-dependent Erk1/2 activation in androgen-independent prostate cancer cell lines

One growth factor receptor commonly altered during prostate tumor progression is the epidermal growth factor receptor (EGFR). EGFR signaling regulates Erk1/2 phosphorylation through multiple mechanisms. We hypothesized that PKC isozymes play a role in EGFR-dependent signaling, and that through PKC isozyme selective inhibition, EGFR-dependent Erk1/2 activation can be attenuated in AICaP cells. ^ To test the hypothesis, PKC activation was induced by 12-O-tetradecanoyi-phorbol-13-acetate (TPA) in PC-3 cells. As a result, Erk1/2 was activated similarly to what was observed upon EGF stimulation. EGF-induced Erk1/2 activation in PC-3 cells was PKC-dependent, as demonstrated through use of a selective PKC inhibitor, GF109203X. This provides evidence for PKC regulatory control over Erk1/2 signaling downstream of EGFR. Next, we demonstrated that when PKC was inhibited by GF109203X, EGF-stimulated Erk1/2 activation was inhibited in PC-3, but not DU145 cells. TPA-stimulated Erk1/2 activation was EGFR-dependent in both DU145 and PC-3 cells, demonstrated through abrogation of Erk1/2 activation by a selective EGFR inhibitor AG1478. These data support PKC control at or upstream of EGFR in AICaP cells. We observed that interfering with ligand/EGFR binding abrogated Erk1/2 signaling in TPA-stimulated cells, revealing a role for PKC upstream of EGFR. ^ Next, we determined which PKC isozymes might be responsible for Erk1/2 regulation. We first determined that human AICaP cell lines express the same PKC isozymes as those observed in clinical prostate cancer specimens (α, ϵ, &zgr;, ι and PKD). Isozyme-selective methods were employed to characterize discrete PKC isozyme function in EGFR-dependent Erk1/2 activation. Pharmacologic inhibitors implicated PKCα in TPA-induced EGFR-dependent Erk1/2 activation in both PC-3 and DU145 cells. Further, the cPKC-specific inhibitor, Gö6976 decreased viablilty of DU145 cells, providing evidence that PKCα is necessary for growth and survival. Finally, resveratrol, a phytochemical with strong cancer therapeutic potential inhibited Erk1/2 activation, and this correlated with selective inhibition of PKCα. These results demonstrate that PKC regulates pathways critical to progression of CaP cells, including those mediated by EGFR. Thus, PKC isozyme-selective targeting is an attractive therapeutic strategy, and understanding the role of specific PKC isozymes in CaP cell growth and survival may aid in development of effective, non-toxic PKC-targeted therapies. ^

A mechanism of insulin-like growth factor binding protein 2-induced cell mobility in glioma

Insulin-like growth factor binding protein 2 (IGFBP2) is a protein known to be overexpressed in a majority of glioblastoma multiforme (GBM) tumors. While it is known the IGFBP2 is involved in promoting GBM tumor cell invasion, no mechanism exists for how the protein is involved in signal transduction pathways leading to enhanced cell invasion. ^ We follow up on preliminary microarray data on IGFBP2-overexpressing GBM cells and protein sequence analysis of IGFBP2 in generating the hypothesis that IGFBP2 interacts with integnn α5 in regulating cell mobility. Microarray data showing upregulation of integrin α5 by IGFBP2 is validated and evidence of protein-protein interaction between IGFBP2 and integrin α5 is found. The exact binding domain on IGFBP2 responsible for its interaction with integrin α5 is also determined, confirming our initial findings and reaffirming that the IGFBP2/integrin α5 interaction is specific. Disruption of this interaction resulted in attenuation of IGFBP2-enhanced cell mobility. Further, we found that cell mobility is only enhanced when IGFBP2 and integrin α5 are both overexpressed and able to interact with each other. ^ We also determined fibronectin to be a critical player in the activation of the IGFBP2/integrin α5 pathway. The activation of this pathway appears to be progressive and initiates once GBM cells have sufficiently established anchorage. ^

Roles of insulin-like growth factor binding protein-3 in gastrointestinal stromal tumors

Imatinib mesylate, a selective inhibitor of KIT, PDGFR, and Abl kinases, has shown significant success as a therapy for patients with advanced gastrointestinal stromal tumors (GISTs). However, the underlying mechanisms of imatinib-induced cytotoxicity are not well understood. Using gene expression profiling and real-time PCR for target validation, we identified insulin-like growth factor binding protein-3 (IGFBP3) to be to be up-regulated after imatinib treatment in imatinib-sensitive GISTs. IGFBP3 is a multifunctional protein that regulates cell proliferation and survival and mediates the effects of a variety of anti-cancer agents through IGF-dependent and IGF-independent mechanisms. Therefore, we hypothesized that IGFBP3 mediates GIST cell response to imatinib. To test this hypothesis, we manipulated IGFBP3 protein levels in two KIT mutant, imatinib-sensitive GIST cell lines and assessed the resultant changes in cell viability, survival, and imatinib sensitivity. In GIST882 cells, endogenous IGFBP3 was required for cell viability. However, inhibiting imatinib-induced IGFBP3 up-regulation by RNA interference or neutralization resulted in reduced drug sensitivity, suggesting that IGFBP3 sensitizes GIST882 cells to imatinib. GIST-T1 cells, on the other hand, had no detectable levels of endogenous IGFBP3, nor did imatinib induce IGFBP3 up-regulation, in contrast to our previous findings. IGFBP3 overexpression in GIST-T1 cells reduced viability but did not induce cell death; rather, the cells became polyploid through a mechanism that may involve attenuated Cdc20 expression and securin degradation. Moreover, IGFBP3 overexpression resulted in a loss of KIT activation and decreased levels of mature KIT. Consistent with this, GIST-T1 cells overexpressing IGFBP3 were less sensitive to imatinib. Furthermore, as neither GIST882 cells nor GIST-T1 cells expressed detectable levels of IGF-1R, IGFBP3 is likely not exerting its effects by modulating IGF signaling through IGF-1R or IR/IGF-1R hybrid receptors in these cell lines. Collectively, these findings demonstrate that IGFBP3 has cell-dependent effects and would, therefore, not be an ideal marker for identifying imatinib response in GISTs. Nevertheless, our results provide preliminary evidence that IGFBP3 may have some therapeutic benefits in GISTs. ^

SEVERITY OF IRON DEFICIENCY ANEMIA AND ITS RELATIONSHIP TO GROWTH AND MORBIDITY IN A POPULATION OF PRE-SCHOOLERS IN RURAL GUATEMALA (IRON DEFICIENCY, MORBIDITY, GROWTH)

The objectives of this study were to determine the nature of the relationship between severity of iron deficiency anemia, response to iron treatment, respiratory and gastrointestinal illness and weight change. Seventy-five pre-school children from rural Guatemala received daily oral iron therapy for an eleven week period, and were classified into one of three groups having different degrees of iron deficiency anemia. Anthropometric and biochemical data were collected prior and after iron treatment; morbidity data were collected throughout the period of treatment. The outcome variables were percentage weight change, percentage of total days ill with any type of symptom, percentage of total days ill with gastrointestinal symptoms, percentage of total days ill with respiratory symptoms, percentage of total days ill with combination syndrome symptoms. Age, sex and socio-economic status, were independent of any of the independent or outcome variables used. On the other hand, the level of hemoglobin covaried with the height of the children, the smallest children were the most severely anemic. The relationships between hemoglobin levels and weight change, frequency of morbidity (gastrointestinal, respiratory and combination syndrome) and total number of days ill with any symptomatology were investigated. No statistical significance was found in these analyses except when contrasting children with normal hemoglobin levels to iron deficient children, where the findings indicated the normal children experienced more gastrointestinal morbidity. The same relationship were again analyzed but including delta hemoglobin as covariate in the analysis, this latter one was found to be significant at 7% when the percentage of days ill from gastrointestinal morbidity was tested against the hemoglobin groups. The relationship found indicates that, all other covariates accounted for, the percentage of days ill from gastrointestinal morbidity will decrease approximately 1% for each 1% increase in delta of hemoglobin. ^

Growth rates and stable carbon and oxygen isotope record of corals from the Red Sea

Monthly delta18O records of 2 coral colonies (Porites cf. lutea and P. cf. nodifera) from different localities (Aqaba and Eilat) from the northern Gulf of Aqaba, Red Sea, were calibrated with recorded sea surface temperatures (SST) between 1988 and 2000. The results show high correlation coefficients between SST and delta18O. Seasonal variations of coral delta18O in both locations could explain 91% of the recorded SST. Different delta18O/SST relations from both colonies and from the same colonies were obtained, indicating that delta18O from coral skeletons were subject to an extension rate effect. Significant delta18O depletions are associated with high extension rates and higher values with low extension rates. The relation between coral skeletal delta18O and extension rate is not linear and can be described by a simple exponential model. An inverse relationship extends over extension rates from 1 to 5 mm/yr, while for more rapidly growing corals and portions of colonies the relation is constant and the extension rate does not appear to have a significant effect. We recommend that delta18O values be obtained from fast-growing corals or from portions in which the isotopic disequilibrium is fairly constant (extension rate >5 mm/yr). The results show that interspecific differences in corals may produce a significant delta18O profile offset between 2 colonies that is independent of environmental and extension-rate effects. We conclude that the rate of skeletal extension and the species of coral involved have an important influence on coral delta18O and must be considered when using delta18O records for paleoclimatic reconstructions.

Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)

Understanding the ecological implications of global climate change requires investigations of not only the direct effects of environmental change on species performance but also indirect effects that arise from altered species interactions. We performed CO2 perturbation experiments to investigate the effects of ocean acidification on the trophic interaction between the brown seaweed Fucus vesiculosus and the herbivorous isopod Idotea baltica. We predicted faster growth of F. vesiculosus at elevated CO2-concentrations and higher carbon content of the algal tissue. We expected that I. baltica has different consumption rates on algae that have been grown at different CO2 levels and that the isopods remove surplus carbon metabolically by enhanced respiration. Surprisingly, growth of F. vesiculosus as well as the C:N-ratio of the algal tissue were reduced at high CO2-levels. The changes in the elemental composition had no effect on the consumption rates and the respiration of the herbivores. An additional experiment showed that consumption of F. vesiculosus by the isopod Idotea emarginata was independent of ocean acidification and temperature. Our results could not reveal any effects of ocean acidification on the per capita strength of the trophic interaction between F. vesiculosus and its consumers. However, reduced growth of the algae at high CO2-concentrations might reduce the capability of the seaweed to compensate losses due to intense herbivory.

Seawater carbonate chemistry, cell numbers and growth rate during experiments with dinoflagellates, 2007

The effects of dissolved inorganic carbon (DIC) on the growth of 3 red-tide dinoflagellates (Ceratium lineatum, Heterocapsa triquetra and Prorocentrum minimum) were studied at pH 8.0 and at higher pH levels, depending upon the pH tolerance of the individual species. The higher pH levels chosen for experiments were 8.55 for C. lineatum and 9.2 for the other 2 species. At pH 8.0, which approximates the pH found in the open sea, the maximum growth in all species was maintained until the total DIC concentration was reduced below ~0.4 and 0.2 mM for C. lineatum and the other 2 species, respectively. Growth compensation points (concentration of inorganic carbon needed for maintenance of cells) were reached at ~0.18 and 0.05 mM DIC for C. lineatum and the other 2 species, respectively. At higher pH levels, maximum growth rates were lower compared to growth at pH 8, even at very high DIC concentrations, indicating a direct pH effect on growth. Moreover, the concentration of bio-available inorganic carbon (CO2 + HCO3-) required for maintenance as well as the half-saturation constants were increased considerably at high pH compared to pH 8.0. Experiments with pH-drift were carried out at initial concentrations of 2.4 and 1.2 mM DIC to test whether pH or DIC was the main limiting factor at a natural range of DIC. Independent of the initial DIC concentrations, growth rates were similar in both incubations until pH had increased considerably. The results of this study demonstrated that growth of the 3 species was mainly limited by pH, while inorganic carbon limitation played a minor role only at very high pH levels and low initial DIC concentrations.

Data compilation of respiration, feeding, and growth rates of marine pelagic organisms

The metabolic rate of organisms may either be viewed as a basic property from which other vital rates and many ecological patterns emerge and that follows a universal allometric mass scaling law; or it may be considered a property of the organism that emerges as a result of the organism's adaptation to the environment, with consequently less universal mass scaling properties. Data on body mass, maximum ingestion and clearance rates, respiration rates and maximum growth rates of animals living in the ocean epipelagic were compiled from the literature, mainly from original papers but also from previous compilations by other authors. Data were read from tables or digitized from graphs. Only measurements made on individuals of know size, or groups of individuals of similar and known size were included. We show that clearance and respiration rates have life-form-dependent allometries that have similar scaling but different elevations, such that the mass-specific rates converge on a rather narrow size-independent range. In contrast, ingestion and growth rates follow a near-universal taxa-independent ~3/4 mass scaling power law. We argue that the declining mass-specific clearance rates with size within taxa is related to the inherent decrease in feeding efficiency of any particular feeding mode. The transitions between feeding mode and simultaneous transitions in clearance and respiration rates may then represent adaptations to the food environment and be the result of the optimization of tradeoffs that allow sufficient feeding and growth rates to balance mortality.

The Agrobacterium vitis T-6b oncoprotein induces auxin-independent cell expansion in tobacco

Among the Agrobacterium T-DNA genes, rolB, rolC, orf13, orf8, lso, 6b and several other genes encode weakly homologous proteins with remarkable effects on plant growth. The 6b oncogene induces tumors and enations. In order to study its properties we have used transgenic tobacco plants that carry a dexamethasone-inducible 6b gene, dex-T-6b. Upon induction, dex-T-6b plants develop a large array of morphological modifications, some of which involve abnormal cell expansion. In the present investigation, dex-T-6b-induced expansion was studied in intact leaves and an in vitro leaf disc system. Although T-6b and indole-3-acetic acid (IAA) both induced expansion and were non-additive, T-6b expression did not increase IAA levels, nor did it induce an IAA-responsive gene. Fusicoccin (FC) is known to stimulate expansion by increasing cell wall plasticity. T-6b- and FC-induced expansion were additive at saturating FC concentrations, indicating that T-6b does not act by a similar mechanism to FC. T-6b expression led to higher leaf osmolality values, in contrast to FC, suggesting that the T-6b gene induces expansion by increasing osmolyte concentrations. Metabolite profiling showed that glucose and fructose played a major role in this increase. We infer that T-6b disrupts the osmoregulatory controls that govern cell expansion during development and wound healing.

The interactive effects of temperature and osmotic potential on the growth of marines isolates of Fusarium solani

The mycelial growth of 18 Fusarium solani strains isolated from sea beds of the south-eastern coast of Spain was tested on potato-dextrose agar adjusted to different osmotic potentials with either KCl or NACl (-1.50 to -144.54 bars) in 10ºC intervals ranging from 15 to 35ºC. Fungal growth was determined by measuring colony diameter after 4 days incubation. Mycelial growth was maximal at 25ºC. The quantity and frequency pattern of mycelial growth of F. solani differ significantly at 15 and 25ºC, with maximal occurring at the highest water potential tested (-1.50 bars); and at 35ºC, with a maximal mycelial growth at -13.79 bars. The effect of water potential was independent of salt composition. The general growth pattern of F. solani showed declining growth at potentials below -41.79 bars. Fungal growth at 35ºC was always higher than that growth at 15ºC, of all the water potentials tested. Significant differences observed in the response of mycelia to water potential and temperature as main and interactive effects. The viability of cultures was increasingly inhibited as the water potential dropped, but some growth was still observed at -99.56 bars. These findings could indicate that marine strains of F. solani have a physiological mechanism that permits survival in environments with low water potential. The observed differences in viability and the magnitude growth could indicate that the biological factors governing potential and actual growth are affected by osmotic potential in different ways.

The Interactive Effects of Temperature and Osmotic Potential on the Growth of Aquatic Isolates of Fusarium culmorum.

The mycelial growth of 10 Fusarium culmorum strains isolated from water of the Andarax riverbed in the provinces of Granada and Almeria in southeastern Spain was tested on potato-dextroseagar adjusted to different osmotic potentials with either KCl or NaCl (−1.50 to−144.54 bars) at 10◦C intervals ranging from15◦ to 35◦C. Fungal growth was determined by measuring colony diameter after 4 d of incubation. Mycelial growth was maximal at 25◦C. The quantity and capacity of mycelial growth of F. culmorum were similar at 15 and 25◦C, with maximal growth occurring at −13.79 bars water potential and a lack of growth at 35◦C. The effect of water potential was independent of salt composition. The general growth pattern of Fusarium culmorum growth declined at potentials below −13.79 bars. Fungal growth at 25◦C was always greater than growth at 15◦C, at all of the water potentials tested. Significant differences were observed in the response ofmycelia to water potential and temperature as main and interactive effects. The number of isolates that showed growth was increasingly inhibited as the water potential dropped, but some growth was still observable at −99.56 bars. These findings could indicate that F. culmorum strains isolated from water have a physiological mechanism that permits survival in environments with low water potential. Propagules of Fusarium culmorum are transported long distances by river water, which could explain the severity of diseases caused by F.culmorum on cereal plants irrigated with river water and its interaction under hydric stress ormoderate soil salinity. The observed differences in growth magnitude and capacity could indicate that the biological factors governing potential and actual growth are affected by osmotic potential in different ways.

p21WAF1 is required for butyrate-mediated growth inhibition of human colon cancer cells

A diet high in fiber is associated with a decreased incidence and growth of colon cancers. Butyrate, a four-carbon short-chain fatty acid product of fiber fermentation within the colon, appears to mediate these salutary effects. We sought to determine the molecular mechanism by which butyrate mediates growth inhibition of colonic cancer cells and thereby to elucidate the molecular link between a high-fiber diet and the arrest of colon carcinogenesis. We show that concomitant with growth arrest, butyrate induces p21 mRNA expression in an immediate-early fashion, through transactivation of a promoter cis-element(s) located within 1.4 kb of the transcriptional start site, independent of p53 binding. Studies using the specific histone hyperacetylating agent, trichostatin A, and histone deacetylase 1 indicate that growth arrest and p21 induction occur through a mechanism involving histone hyperacetylation. We show the critical importance of p21 in butyrate-mediated growth arrest by first confirming that stable overexpression of the p21 gene is able to cause growth arrest in the human colon carcinoma cell line, HT-29. Furthermore, using p21-deleted HCT116 human colon carcinoma cells, we provide convincing evidence that p21 is required for growth arrest to occur in response to histone hyperacetylation, but not for serum starvation nor postconfluent growth. Thus, p21 appears to be a critical effector of butyrate-induced growth arrest in colonic cancer cells, and may be an important molecular link between a high-fiber diet and the prevention of colon carcinogenesis.

Mahogany (mg) stimulates feeding and increases basal metabolic rate independent of its suppression of agouti

The mahogany (mg) locus originally was identified as a recessive suppressor of agouti, a locus encoding a skin peptide that modifies coat color by antagonizing the melanocyte-stimulating hormone receptor or MC1-R. Certain dominant alleles of agouti cause an obesity syndrome when ectopic expression of the peptide aberrantly antagonizes the MC4-R, a related melanocyte-stimulating hormone receptor expressed in hypothalamic circuitry and involved in the regulation of feeding behavior and metabolism. Recent work has demonstrated that mg, when homozygous, blocks not only the ability of agouti to induce a yellow coat color when expressed in the skin of the lethal yellow mouse (AY), but also the obesity resulting from ectopic expression of agouti in the brain. Detailed analysis of mg/mg AY/a animals, presented here, demonstrates that mg/mg blocks the obesity, hyperinsulinemia, and increased linear growth induced by ectopic expression of the agouti peptide. Remarkably, however, mg/mg did not reduce hyperphagia in the AY/a mouse. Furthermore, mg/mg induced hyperphagia and an increase in basal metabolic rate in the C57BL/6J mouse in the absence of AY. Consequently, although mahogany is broadly required for agouti peptide action, it also appears to be involved in the control of metabolic rate and feeding behavior independent of its suppression of agouti.

The C-type lectin domains of lecticans, a family of aggregating chondroitin sulfate proteoglycans, bind tenascin-R by protein– protein interactions independent of carbohydrate moiety

The lecticans are a family of chondroitin sulfate proteoglycans including aggrecan, versican, neurocan, and brevican. The C-terminal globular domains of lecticans are structurally related to selectins, consisting of a C-type lectin domain flanked by epidermal growth factor and complement regulatory protein domains. The C-type lectin domain of versican has been shown to bind tenascin-R, an extracellular matrix protein specifically expressed in the nervous system, and the interaction was presumed to be mediated by a carbohydrate–protein interaction. In this paper, we show that the C-type lectin domain of brevican, another lectican that is specifically expressed in the nervous system, also binds tenascin-R. Surprisingly, this interaction is mediated by a protein–protein interaction through the fibronectin type III domains 3–5 of tenascin-R, independent of any carbohydrates or sulfated amino acids. The lectin domains of versican and other lecticans also bind the same domain of tenascin-R by protein–protein interactions. Surface plasmon resonance analysis revealed that brevican lectin has at least a 10-fold higher affinity than the other lectican lectins. Tenascin-R is coprecipitated with brevican from adult rat brain extracts, suggesting that tenascin-R and brevican form complexes in vivo. These results demonstrate that the C-type lectin domain can interact with fibronectin type III domains through protein–protein interactions, and suggest that brevican is a physiological tenascin-R ligand in the adult brain.