Heat shock protein synthesis can be induced during recovery from cold treatment of Drosophila melanogaster larvae. Survival of larvae after a cold treatment is dramatically improved by a mild heat shock just before the cold shock. The conditions which induce tolerance to cold are similar to those which confer tolerance to heat.Heat shock proteins are a group of proteins which are universally expressed in response to heat, heavy metals, and a variety of chemical treatments (1, 5, 10, 11). The major heat shock proteins synthesized in bacteria, plants, and mammals show a high degree of sequence homology (1, 5). These proteins are also synthesized during some periods in normal development.Transient resistance to killing by heat stress has been observed to be induced in many organisms by an initial exposure to a sublethal heat treatment (2, 7, 11). We have shown that conditions which induce thermotolerance in Drosophila melanogaster also induce the synthesis of heat shock proteins (7), and this correlation has been repeated many times in a variety of systems (1,5). Because of this and the universality of the response, it has been suggested that heat shock proteins play a role in protecting cells from damage caused by these stresses. The precise function of the heat shock proteins as well their role in stress protection are still uncertain.We show here that heat shock proteins are synthesized during recovery from prolonged exposure to cold in the absence of heat shock and that a mild heat treatment of the same kind which protects against death from heat shock also prevents death from exposure to cold.Induction of heat shock proteins by cold treatment. Drosophila larvae recovering from cold treatments of more than 8 h at 0°C synthesized the same set of heat shock or stress proteins that are made in response to heat shock (7, 9, 12). Figure 1 shows the proteins synthesized in larval salivary glands during recovery from 14 h at 0°C. Salivary glands were dissected from cold-shocked larvae that had been allowed to recover for 0, 10, 20, 30, or 40 min at 25°C. The salivary glands were labeled for 30 min with ['5S] methionine, and the proteins were separated on sodium dodecyl sulfate-polyacrylamide gels as described previously (8). An untreated 25°C control is shown in Fig. 1, lane C, as is a typical heat shock pattern from animals heated to 37°C (lane H). Clearly, all the major heat shock proteins appeared in the 20-min sample. The strongest heat shock protein synthesis patterns were seen not during the 30 min immediately following the cold treatment, but rather when larvae were allowed to recover for 30 or 40 min before being labeled. This suggests that the induction of the heat shock proteins may be a response to the shift from 0 to 25°C rather than a response to the 0°C treatment itself. We are currently doing * Corresponding author.RNA blots to determine directly the levels of hsp70 mRNA present at 0°C and during the recovery at 25°C.Experiments designed to evaluate the effects of different durations of cold treatments ...

We have ablated peripheral lymph nodes in sheep and subsequently cannulated the pseudo-afferent lymphatic vessel that arises as a consequence of afferent lymphatic vessels reanastomosing with the former efferent duct. This technique allows the collection of lymph with a cellular composition that resembles true afferent fluid, and in particular, containing 1-10% dendritic cells. A 16-h collection of this lymph may contain between 10(6) and 10(7) dendritic cells. This dendritic cell population may be enriched to greater than 75% by a single-density gradient centrifugation step. We have generated a mAb that recognizes sheep CD1. This monoclonal not only reacts with afferent dendritic cells, but with dendritic cells in the skin and paracortical T cell areas of lymph nodes. The expression of CD1 suggests afferent dendritic cells are related to skin Langerhans' cells and other dendritic cells that act as accessory cells for T cell responses. Consistent with this is the high level of expression by dendritic cells of molecules involved in antigen recognition by T cells, including MHC class I and class II. Afferent dendritic cells express high levels of the cellular adhesion molecule LFA-3, and at the same time express a ligand for this molecule, namely CD2. The accessory functions of afferent dendritic cells resemble those displayed by mature Langerhans' cells and by lymph node interdigitating cells. These include clustering with resting T cells and stimulating their proliferation in a primary response to antigen. Afferent dendritic cells are capable of acquiring soluble protein antigen in vivo or in vitro and presenting the material directly to autologous T cells in an antigen-specific manner. We conclude that afferent dendritic cells represent a lymph-borne Langerhans' cell involved in antigen carriage to the lymph node.

Highlights d The transcription factor Spic restrains inflammatory responses in macrophages d Spic promotes the expression of the iron exporter ferroportin in activated macrophages d NF-kB activity is required for the expression of Spic in activated macrophages d Interferon-gamma suppresses Spic expression in activated macrophages

The true potential of cytokine therapies in cancer treatment is limited by the inability to deliver optimal concentrations into tumor sites due to dose-limiting systemic toxicities. To maximize the efficacy of cytokine therapy, recombinant antibody-cytokine fusion proteins have been constructed by a number of groups to harness the tumor-targeting ability of monoclonal antibodies. The aim is to guide cytokines specifically to tumor sites where they might stimulate more optimal anti-tumor immune responses while avoiding the systemic toxicities of free cytokine therapy. Antibody-cytokine fusion proteins containing IL-2, IL-12, IL-21, TNFα, and interferons α, β and γ have been constructed and have shown anti-tumor activity in pre-clinical and early phase clinical studies. Future priorities for development of this technology include optimization of tumor targeting, bioactivity of the fused cytokine, and choice of appropriate agents for combination therapies. This review is intended to serve as a framework for engineering an ideal antibody-cytokine fusion protein, focusing on previously developed constructs and their clinical trial results.

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Macrophages in SHH subgroup medulloblastoma display dynamic heterogeneity that varies with treatment modality Graphical abstract Highlights d Sonic Hedgehog (SHH) subgroup of medulloblastoma (MB) recruits diverse macrophages d Radiation or molecular-targeted therapy alters macrophage distribution in SHH-MB d Radiation recruits immunosuppressive monocyte-derived macrophages (TAMoMacs) in SHH-MB d Radiation-induced TAMoMacs regulate CD8 T cell and neutrophil numbers in SHH-MB

Addition of fulvestrant to erlotinib was well tolerated, with increased activity noted among EGFR wild type patients compared to erlotinib alone, albeit in an unplanned subset analysis.

The LDL receptor-related protein 1 (LRP1) is a large endocytic receptor that binds and mediates the endocytosis of numerous structurally diverse ligands. Currently, the basis for ligand recognition by LRP1 is not well understood. LRP1 requires a molecular chaperone, termed the receptor-associated protein (RAP), to escort the newly synthesized receptor from the endoplasmic reticulum to the Golgi. RAP is a three-domain protein that contains the following two high affinity binding sites for LRP1: one is located within domains 1 and 2, and one is located in its third domain. Studies on the interaction of the RAP third domain with LRP1 reveal critical contributions by lysine 256 and lysine 270 for this interaction. From these studies, a model for ligand recognition by this class of receptors has been proposed. Here, we employed surface plasmon resonance to investigate the binding of RAP D1D2 to LRP1. Our results reveal that the high affinity of D1D2 for LRP1 results from avidity effects mediated by the simultaneous interactions of lysine 60 in D1 and lysine 191 in D2 with sites on LRP1 to form a bivalent D1D2-LRP1 complex. When lysine 60 and 191 are both mutated to alanine, the binding of D1D2 to LRP1 is ablated. Our data also reveal that D1D2 is able to bind to a second distinct site on LRP1 to form a monovalent complex. The studies confirm the canonical model for ligand recognition by this class of receptors, which is initiated by pairs of lysine residues that dock into acidic pockets on the receptor.The LDL receptor-related protein 1 (LRP1) is a member of the LDL receptor family and is a highly efficient endocytic and signal-transducing receptor that plays an important role in vascular development, lipoprotein metabolism, and inflammation (1-3). Originally identified as the hepatic receptor responsible for the removal of ␣ 2 -macroglobulin (␣ 2 M) 3 -protease complexes (4), we now know that LRP1 recognizes numerous ligands, including lipoproteins, matrix proteins, growth factors (1, 2, 5, 6), and extracellular proteases (7-11). Deletion of the Lrp1 gene in mice results in early embryonic lethality at E13.5 (12, 13) due to extensive hemorrhaging resulting from a failure to recruit and maintain vascular smooth muscle cells and pericytes in the vessels. Selective deletion of LRP1 in vascular smooth muscle cells (smLRP1 Ϫ/Ϫ mice) reveals that LRP1 protects against the development of atherosclerosis by attenuating PDGF receptor activation (14, 15) and prevents formation of aneurysms (11, 16) in part by regulating the levels of proteases that are known to degrade matrix components (11).The efficient delivery of LRP1 and certain other members of the LDL receptor family to the cell surface require their association with a 39-kDa protein termed the receptor-associated protein (RAP). RAP was initially identified when it co-purified with LRP1 (17,18). Subsequent work demonstrated that RAP binds tightly to LRP1 and prevents ligands from associating with this receptor (19,20). In the endoplasmic reticulum (ER), RAP functions a...