Growth factor interactions between mouse mammary cell lines cocultured in collagen gels

Summary Three related mouse mammary cell lines were cultured in collagen gels and assayed for growth factor responsiveness and interaction via soluble factors. The CL-S1 cell line is nontumorigenic and grows poorly in collagen gel culture. The +SA and −SA cell lines exhibit different degrees of malignant behavior in vivo and have different growth properties in vitro. In collagen gel culture, +SA growth was stimulated by serum but not by epidermal growth factor (EGF), whereas both serum and EGF were required for optimal growth of −SA cells of early passage number as well as CL-S1 cells. −SA cells of later passage repeatedly exhibited a change so as to no longer require serum while retaining EGF responsiveness. [¹²⁵I]EGF binding analyses indicated that CL-S1 cells bound EGF with less affinity than did −SA cells whereas +SA cells bound almost to ligand. When cell lines were maintained in separate collagen gels but shared the same culture medium, growth of +SA or −SA cells was slightly enhanced in the presence of CL-S1 cells and −SA cell growth was enhanced by the presence of +SA cells. Using the normal rat kidney fibroblast line NRK (clone 49F) as an indicator, serum-containing conditioned media from each cell line and from each pair of cell lines cultured in collagen gels were tested for transforming growth factor (TGF) activity. Both the −SA and CL-S1 lines tested positive for TGF-α production and possibly released a TGF-β activity. These results suggest mechanisms by which cell populations in and around tumors can modify one another’s growth characteristics. The work was supported by a grant from the American Institute for Cancer Research, by American Cancer Society Institutional grant IN-119, by funds from the Poncin Trust (Seattle-First National Bank), and by grants CA-39611 and CA46885 from the National Institutes of Health, Bethesda, MD.     

Population genetics of grotto sculpin (Cottus specus), a new cave-adapted fish species

The grotto sculpin (Cottus specus) is a troglomorphic fish endemic to cave systems in Perry County, Missouri. These fish are state-threatened and were recently listed as federally endangered under the Endangered Species Act (ESA). Due to the unstable nature of the cave environment, grotto sculpins are highly susceptible to pollution via agricultural and waste runoff. Here we provide a summary of population genetic diversity within the range of the grotto sculpin. Samples from twenty-one cave, surface, and resurgence sites within and surrounding the Bois Brule drainage in Perry County, southeast Missouri, USA were collected and sequence data from the mitochondrial control were analyzed using analysis of molecular variance, maximum likelihood, parsimony, and Bayesian techniques. We found a substantial degree of divergence from surface populations of Midlands and Black River banded sculpin races (7.7–8.0 %). Within the Bois Brule drainage, two distinct lineages of grotto sculpin were identified which correspond with northern and southern cave system boundaries and may represent independent cave invasions.     

Categories and inheritance of resistance to Russian wheat aphid (Homoptera: Aphididae) biotype 2 in a selection from wheat cereal introduction 2401

The Russian wheat aphid, Diuraphis noxia (Kurdjumov) (Homoptera: Aphididae), is one of the most devastating insect pests of wheat (Triticum spp.) and barley (Hordeum spp.) in the world. Yield losses and control costs are valued at several hundred million dollars each year. The use of D. noxia-resistant cultivars is an ecologically, economically, and biologically sound method of managing this pest. Several D. noxia resistance (Dn) genes from wheat have been used to develop cultivars resistant to D. noxia. However, a new U.S. D. noxia biotype (biotype 2) in Colorado is virulent to all known Dn genes except the Dn7 gene from rye (Secale spp.). Hence, there is an immediate need to identify and characterize unique sources of D. noxia resistance to biotypes. In this article, we report resistance to D. noxia biotype 2, identified in a selection from wheat cereal introduction (CItr) 2401, that is controlled by two dominant genes. CItr2401 has a strong antibiosis effect that is exhibited as a reduced intrinsic rate of increase of D. noxia biotype 2. CItr2401 plants also exhibit tolerance to leaf rolling and chlorosis. No antixenosis was detected in CItr2401.     

Host tropism determination by convergent evolution of immunological evasion in the Lyme disease system

Pathogens possess the ability to adapt and survive in some host species but not in others–an ecological trait known as host tropism. Transmitted through ticks and carried mainly by mammals and birds, the Lyme disease (LD) bacterium is a well-suited model to study such tropism. Three main causative agents of LD, Borrelia burgdorferi, B. afzelii, and B. garinii, vary in host ranges through mechanisms eluding characterization. By feeding ticks infected with different Borrelia species, utilizing feeding chambers and live mice and quail, we found species-level differences in bacterial transmission. These differences localize on the tick blood meal, and specifically complement, a defense in vertebrate blood, and a polymorphic bacterial protein, CspA, which inactivates complement by binding to a host complement inhibitor, Factor H (FH). CspA selectively confers bacterial transmission to vertebrates that produce FH capable of allele-specific recognition. CspA is the only member of the Pfam54 gene family to exhibit host-specific FH-binding. Phylogenetic analyses revealed convergent evolution as the driver of such uniqueness, and that FH-binding likely emerged during the last glacial maximum. Our results identify a determinant of host tropism in Lyme disease infection, thus defining an evolutionary mechanism that shapes host-pathogen associations.     

Vascular targeting to the SST2 receptor improves the therapeutic response to near-IR two-photon activated PDT for deep-tissue cancer treatment

Background

Broader clinical acceptance of photodynamic therapy is currently hindered by (a) poor depth efficacy, and (b) predisposition towards establishment of an angiogenic environment during the treatment. Improved depth efficacy is being sought by exploiting the NIR tissue transparency window and by photo-activation using two-photon absorption (2PA). Here, we use two-photon activation of PDT sensitizers, untargeted and targeted to SST2 receptors or EGF receptors, to achieve deep tissue treatment.

Methods

Human tumor lines, positive or negative for SST2r expression were used, as well as murine 3LL cells and bovine aortic endothelial cells. Expression of SST2 receptors on cancer cells and tumor vasculature was evaluated in vitro and frozen xenograft sections. PDT effects on tumor blood flow were followed using in vivo scanning after intravenous injection of FITC conjugated dextran 150 K. Dependence of the PDT efficacy on the laser pulse duration was evaluated. Effectiveness of targeting to vascular SST2 receptors was compared to that of EGF receptors, or no targeting.

Results

Tumor vasculature stained for SST2 receptors even in tumors from SST2 receptor negative cell lines, and SST2r targeted PDT led to tumor vascular shutdown. Stretching the pulse from ~ 120 fs to ~ 3 ps led to loss of the PDT efficacy especially at greater depth. PDT targeted to SST2 receptors was much more effective than untargeted PDT or PDT targeted to EGF receptors.

General significance

The use of octreotate to target SST2 receptors expressed on tumor vessels is an excellent approach to PDT with few recurrences and some long term cures.     

An adaptive evolutionary shift in Fusarium head blight pathogen populations is driving the rapid spread of more toxigenic Fusarium graminearum in North America

Analysis of Fusarium head blight (FHB) pathogen diversity revealed that 3ADON producing Fusarium graminearum are prevalent in North America and identified significant population structure associated with trichothecene chemotype differences (F(ST)>0.285; P<0.001). In addition, we identified a trichothecene chemotype cline in Canada and documented a recent and significant shift in FHB pathogen composition by demonstrating that the 3ADON chemotype frequency in western Canada increased more than 14-fold between 1998 and 2004. On average, isolates from 3ADON populations produced significantly (P<0.05) more trichothecene and had significantly (P<0.005) higher fecundity and growth rates than isolates from the 15ADON population. These results indicate that selection is driving the rapid spread of an introduced pathogen population that is more toxigenic and potentially more vigorous. The discovery of this previously unrecognized pathogen diversity has significant implications for food safety and cereal production in North America.     

Systemic and brain macrophage infections in relation to the development of simian immunodeficiency virus encephalitis

The brains of individuals with lentiviral-associated encephalitis contain an abundance of infected and activated macrophages. It has been hypothesized that encephalitis develops when increased numbers of infected monocytes traffic into the central nervous system (CNS) during the end stages of immunosuppression. The relationships between the infection of brain and systemic macrophages and circulating monocytes and the development of lentiviral encephalitis are unknown. We longitudinally examined the extent of monocyte/macrophage infection in blood and lymph nodes of pigtailed macaques that did or did not develop simian immunodeficiency virus encephalitis (SIVE). Compared to levels in macaques that did not develop SIVE, more ex vivo virus production was detected from monocyte-derived macrophages and nonadherent peripheral blood mononuclear cells (PBMCs) from macaques that did develop SIVE. Prior to death, there was an increase in the number of circulating PBMCs following a rise in cerebrospinal fluid viral load in macaques that did develop SIVE but not in nonencephalitic macaques. At necropsy, macaques with SIVE had more infected macrophages in peripheral organs, with the exception of lymph nodes. T cells and NK cells with cytotoxic potential were more abundant in brains with encephalitis; however, T-cell and NK-cell infiltration in SIVE and human immunodeficiency virus encephalitis was more modest than that observed in classical acute herpes simplex virus encephalitis. These findings support the hypothesis that inherent differences in host systemic and CNS monocyte/macrophage viral production are associated with the development of encephalitis.