TBI risk stratification at presentation: a prospective study of the incidence and timing of radiographic worsening in the Parkland Protocol.

BACKGROUND: We have created a theoretical algorithm for venous thromboembolism prophylaxis after traumatic brain injury (TBI) known as the Parkland Protocol, which stratifies patients into low-, medium-, and high-risk categories for spontaneous progression of hemorrhage. This prospective study characterizes the incidence and timing of radiographic progression of the TBI patterns in these categories. METHODS: Inclusion criterion was presentation with intracranial blood between February 2010 and March 2011; exclusion was receipt of only one computed tomographic scan of the head during the inpatient stay or preinjury warfarin. At admission, all patients were preliminarily categorized per the Parkland Protocol as follows: low risk (LR), patients meeting the modified Berne-Norwood criteria; moderate risk (MR), injuries larger than the modified Berne-Norwood criteria without requiring a neurosurgical procedure; high risk (HR), any patient with a craniotomy/monitor. RESULTS: A total of 245 patients with intracranial hemorrhage were enrolled during the 13-month study period. Of patients preliminarily classified as LR at admission (n = 136), progression was seen in 25.0%. Spontaneous worsening was seen in 7.4% of LR patients at 24 hours after injury, and no LR patients progressed at 72 hours after injury. In patients initially classified as MR at admission (n = 42), progression was seen in 42.9%, with 91.5% of patients demonstrating stable computed tomographic head scans at 72 hours after injury. In patients initially classified as HR (n = 67), 64.2% demonstrated spontaneous progression of their TBI patterns, with 10.5% continuing to progress at 72 hours after injury. Most repeat scans were performed as routinely scheduled studies (81-91%). CONCLUSION: Increases in the incidence of spontaneous worsening were seen as severities of injury progressed from the Parkland Protocol's LR to MR to HR arms. The time frames for these spontaneous worsenings seem to be such that the protocol's theoretical recommendations for venous thromboembolism prophylaxis are worth pursuing as future points of investigation.

Antibody response of bluegill sunfish during development of acquired resistance against the larvae of the freshwater mussel Utterbackia imbecillis.

The larvae of freshwater mussels in the order Unionoida are obligate parasites on fishes, on which they metamorphose into juveniles. Bluegill sunfish (Lepomis macrochirus) acquire resistance against glochidia of the freshwater mussel Utterbackia imbecillis after 2 infections. In order to study the systemic and mucosal antibody response associated with acquired resistance, sera from experimentally infected fish were collected at 10-d intervals during 4 sequential infection periods and from naïve fish. Enzyme-linked immunosorbant assays (ELISA) revealed that fish exhibited a humoral and mucosal antibody response around day 20 after the 1st infection which was followed by second antibody response beginning at day 60 (day 20, 3rd infection) that persisted until the end of the collection period. Western blots of glochidial proteins probed with the sera revealed that the profile of proteins recognized by antibodies produced by fish changed over the course of multiple infections. Serum collected from fish at day 20 (peak of primary response) contained antibodies against approximately 39 and 91 kDa proteins. Immunohistochemical studies on whole-mount glochidia probed with serum from these fish demonstrated that the antibodies recognize granular structures located between the larval mantle and shell. Serum collected from fish during the secondary antibody response (days 60-80) bound additional protein bands in Western blots. Those antibodies recognized other cells of the larval mantle, most prominently in a ciliated region that contains the primordia of the gills and organs of the juvenile and adult mussel.

Persistence of host response against glochidia larvae in Micropterus salmoides.

Host fish acquire resistance to the parasitic larvae (glochidia) of freshwater mussels (Unionidae). Glochidia metamorphose into juvenile mussels while encysted on host fish. We investigated the duration of acquired resistance of largemouth bass, Micropterus salmoides (Lacepède, 1802) to glochidia of the broken rays mussel, Lampsilis reeveiana (Call, 1887). Fish received three successive priming infections with glochidia to induce an immune response. Primed fish were held at 22-23 degrees C and were challenged (re-infected) at intervals after priming. Metamorphosis success was quantified as the percent of attached glochidia that metamorphosed to the juvenile stage and were recovered alive. Metamorphosis success at 3, 7, and 12 months after priming was significantly lower on primed fish (26%, 40%, and 68% respectively) than on control fish (85%, 93%, and 92% respectively). A second group of largemouth bass was similarly primed and blood was extracted. Immunoblotting was used to detect host serum antibodies to L. reeveiana glochidia proteins. Serum antibodies were evident in primed fish, but not in naive control fish. Acquired resistance of host fish potentially affects natural reproduction and artificial propagation of unionids, many of which are of conservation concern.

Encapsulation of attached ectoparasitic glochidia larvae of freshwater mussels by epithelial tissue on fins of naive and resistant host fish.

To metamorphose into juveniles and subsequently mature into adults, the glochidia larvae of freshwater mussels in the order Unionoida must temporarily parasitize the gills, fins, or other external structures of fish. Once attached to the fish, the glochidium is encapsulated by host fish epithelial tissue. The migration of epithelial cells of the bluegill sunfish Lepomis macrochirus over glochidia of Utterbackia imbecillis was examined by time-lapse video microscopy, and the morphology was examined by scanning electron microscopy. Initially, the leading edge epithelial cells migrating over the larvae became rounded and the cells moved as a sheet until the attached glochidium was completely covered. Cyst formation on host fish that had been repeatedly exposed to mussel larvae was significantly delayed and morphologically irregular compared to that on naïve fish. Cyst formation on other species of fish that are less successful as hosts was examined. In general, it took longer for glochidia to become encapsulated on these less suitable potential hosts. The delay and irregularities in cyst formation on resistant fish and nonhost fish species may result in increased mortality and reduced success of metamorphosis of glochidia.

A human T-cell lymphotropic virus type 1 enhancer of Myc transforming potential stabilizes Myc-TIP60 transcriptional interactions.

The human T-cell lymphotropic virus type 1 (HTLV-1) infects and transforms CD4+ lymphocytes and causes adult T-cell leukemia/lymphoma (ATLL), an aggressive lymphoproliferative disease that is often fatal. Here, we demonstrate that the HTLV-1 pX splice-variant p30II markedly enhances the transforming potential of Myc and transcriptionally activates the human cyclin D2 promoter, dependent upon its conserved Myc-responsive E-box enhancer elements, which are associated with increased S-phase entry and multinucleation. Enhancement of c-Myc transforming activity by HTLV-1 p30II is dependent upon the transcriptional coactivators, transforming transcriptional activator protein/p434 and TIP60, and it requires TIP60 histone acetyltransferase (HAT) activity and correlates with the stabilization of HTLV-1 p30II/Myc-TIP60 chromatin-remodeling complexes. The p30II oncoprotein colocalizes and coimmunoprecipitates with Myc-TIP60 complexes in cultured HTLV-1-infected ATLL patient lymphocytes. Amino acid residues 99 to 154 within HTLV-1 p30II interact with the TIP60 HAT, and p30II transcriptionally activates numerous cellular genes in a TIP60-dependent or TIP60-independent manner, as determined by microarray gene expression analyses. Importantly, these results suggest that p30II functions as a novel retroviral modulator of Myc-TIP60-transforming interactions that may contribute to adult T-cell leukemogenesis.

HIV-1 Tat interactions with p300 and PCAF transcriptional coactivators inhibit histone acetylation and neurotrophin signaling through CREB.

The human immunodeficiency virus type-1 (HIV-1) infects microglia, macrophages, and astrocytes in the central nervous system (CNS) and may cause severe neurological diseases, such as AIDS-related dementias or progressive encephalopathies, as a result of CNS inflammation and neurotrophin signaling defects associated with expression of viral antigens and HIV-1 replication in the brain. The HIV Tat protein can be endocytosed by surrounding uninfected cells; interacts with transcriptional coactivators/acetyltransferases, p300/CREB-binding protein, and p300/CREB-binding protein-associated factor (PCAF); and induces neuronal apoptosis. Since nerve growth factor (NGF) receptor and brain-derived neurotrophic factor receptor signaling through CREB requires p300 and PCAF histone acetyltransferases, we sought to determine whether HIV-1 Tat coactivator interactions interfere with neurotrophin receptor signaling in neuronal cells. Here, we demonstrate that Tat-coactivator interactions inhibit NGF- and brain-derived neurotrophic factor-responsive CRE trans-activation and neurotrophin protection against apoptosis in PC12 and IMR-32 neuroblastoma cells. Purified recombinant Tat or Tat-derived synthetic peptides, spanning p300- and PCAF-binding sequences, inhibit histone H3/H4 acetylation in vitro. A Tat mutant, TatK28A/K50A, defective for binding p300 and PCAF, neither repressed NGF-responsive CRE transactivation nor inhibited histone acetylation. HIV-1 Tat interacts in PCAF complexes in post-mortem CNS tissues from donor neuro-AIDS patients, as determined by fluorescence resonance energy transfer immunoconfocal microscopy. Importantly, these findings suggest that HIV-1 Tat-coactivator interactions may contribute to neurotrophin signaling impairments and neuronal apoptosis associated with HIV-1 infections of the CNS.

Cross-resistance of largemouth bass to glochidia of unionid mussels.

We tested whether host fish that acquired resistance to glochidia of one mussel species were cross-resistant to glochidia of other species. Largemouth bass (Micropterus salmoides) were primed with 4-5 successive infections of glochidia of Lampsilis reeveiana. The percentage of attached glochidia that survived and transformed to the juvenile stage (transformation success) was compared between primed fish and naïve controls. Transformation success of L. reeveiana, Lampsilis abrupta, Villosa iris, and Utterbackia imbecillis was significantly lower on primed fish (37.8%, 43.5%, 67.0%, and 13.2%, respectively) than on control fish (89.0%, 89.7%, 90.0%, and 22.2% respectively). Immunoblotting was used to analyze the binding of serum antibodies from primed fish with glochidia proteins. Antibodies bound to glochidia proteins of similar molecular weight from L. reeveiana and L. abrupta. Bound proteins of V. iris differed in molecular weight from those of the Lampsilis species. There was no binding to specific glochidia proteins of U. imbecillis or Strophitus undulatus. Our results indicate that host-acquired resistance can extend across mussel genera and subfamilies and might involve both specific and nonspecific mechanisms. Understanding the specificity of acquired resistance of hosts to glochidia could enhance understanding of the evolutionary and ecological relationships between mussels and their host fishes.

Nerve growth factor receptor signaling induces histone acetyltransferase domain-dependent nuclear translocation of p300/CREB-binding protein-associated factor and hGCN5 acetyltransferases.

The transcriptional coactivators, p300/CREB-binding protein-associated factor (PCAF) and hGCN5, are recruited to chromatin-remodeling complexes on enhancers of various gene promoters in response to growth factor stimulation. However, the molecular mechanisms by which surface receptor signals modulate the assembly of nuclear transcription complexes are not fully understood. Here we report that nerve growth factor receptor signaling induces nuclear translocation of PCAF and hGCN5 dependent upon the phosphorylation of Ser and Thr residues within their histone acetyltransferase domains, which requires activation of PI3K, Rsk2(pp90), and MSK-1. Neurotrophin stimulation induces p53(K320) acetylation by PCAF and transcriptionally activates p53-responsive enhancer elements within the p21(WAF/CIP1) promoter associated with G(1)/S arrest during neuronal differentiation. Most importantly, these findings represent the first evidence for signal-dependent nuclear translocation of PCAF and hGCN5 acetyltransferases and allude to a novel mechanism for ligand/receptor modulation of nuclear chromatin-remodeling complexes in neurons.

Ultrastructure of the calcium-sequestering gastrodermal cell in the hydroid Hydractinia symbiolongicarpus (Cnidaria, Hydrozoa).

Large, free-floating crystals of calcium carbonate occur in vacuoles of gastrodermal cells of the hydroid Hydractinia symbiolongicarpus. Here, morphological details about the process by which these cells accumulate and sequester calcium are provided by a cytochemical method designed to demonstrate calcium at the ultrastructural level. Electron-dense material presumably indicative of the presence of calcium was EGTA-sensitive and was shown by parallel electron energy loss spectroscopy (EELS) and energy spectroscopic imaging (ESI) to contain calcium. Calcium occurred in only one cell type, the endodermally derived gastrodermal cell. In these cells, the electron-dense material appeared first as a fine precipitate in the cytosol and nucleus and later as larger deposits and aggregates in the vacuole. During the life cycle, gastrodermal cells of the uninduced planula and the planula during metamorphic induction sequestered calcium. In primary polyps and polyps from established colonies, gastrodermal cells sequestered calcium, but the endodermal secretory cells did not. Our observations support the hypothesis that gastrodermal cells function as a physiological sink for calcium that enters the organism in conjunction with calcium-requiring processes such as motility, secretion, and metamorphosis.