Primary Xenografts of Human Prostate Tissue as a Model to Study Angiogenesis Induced by Reactive Stroma

Characterization of the mechanism(s) of androgen-driven human angiogenesis could have significant implications for modeling new forms of anti-angiogenic therapies for CaP and for developing targeted adjuvant therapies to improve efficacy of androgen-deprivation therapy. However, models of angiogenesis by human endothelial cells localized within an intact human prostate tissue architecture are until now extremely limited. This report characterizes the burst of angiogenesis by endogenous human blood vessels in primary xenografts of fresh surgical specimens of benign prostate or prostate cancer (CaP) tissue that occurs between Days 6–14 after transplantation into SCID mice pre-implanted with testosterone pellets. The wave of human angiogenesis was preceded by androgen-mediated up-regulation of VEGF-A expression in the stromal compartment. The neo-vessel network anastomosed to the host mouse vascular system between Days 6–10 post-transplantation, the angiogenic response ceased by Day 15, and by Day 30 the vasculature had matured and stabilized, as indicated by a lack of leakage of serum components into the interstitial tissue space and by association of nascent endothelial cells with mural cells/pericytes. The angiogenic wave was concurrent with the appearance of a reactive stroma phenotype, as determined by staining for α-SMA, Vimentin, Tenascin, Calponin, Desmin and Masson's trichrome, but the reactive stroma phenotype appeared to be largely independent of androgen availability. Transplantation-induced angiogenesis by endogenous human endothelial cells present in primary xenografts of benign and malignant human prostate tissue was preceded by induction of androgen-driven expression of VEGF by the prostate stroma, and was concurrent with and the appearance of a reactive stroma phenotype. Androgen-modulated expression of VEGF-A appeared to be a causal regulator of angiogenesis, and possibly of stromal activation, in human prostate xenografts

Functional Stability of Unliganded Envelope Glycoprotein Spikes among Isolates of Human Immunodeficiency Virus Type 1 (HIV-1)

The HIV-1 envelope glycoprotein (Env) spike is challenging to study at the molecular level, due in part to its genetic variability, structural heterogeneity and lability. However, the extent of lability in Env function, particularly for primary isolates across clades, has not been explored. Here, we probe stability of function for variant Envs of a range of isolates from chronic and acute infection, and from clades A, B and C, all on a constant virus backbone. Stability is elucidated in terms of the sensitivity of isolate infectivity to destabilizing conditions. A heat-gradient assay was used to determine T90 values, the temperature at which HIV-1 infectivity is decreased by 90% in 1 h, which ranged between ∼40 to 49°C (n = 34). For select Envs (n = 10), the half-lives of infectivity decay at 37°C were also determined and these correlated significantly with the T90 (p = 0.029), though two ‘outliers’ were identified. Specificity in functional Env stability was also evident. For example, Env variant HIV-1ADA was found to be labile to heat, 37°C decay, and guanidinium hydrochloride but not to urea or extremes of pH, when compared to its thermostable counterpart, HIV-1JR-CSF. Blue native PAGE analyses revealed that Env-dependent viral inactivation preceded complete dissociation of Env trimers. The viral membrane and membrane-proximal external region (MPER) of gp41 were also shown to be important for maintaining trimer stability at physiological temperature. Overall, our results indicate that primary HIV-1 Envs can have diverse sensitivities to functional inactivation in vitro, including at physiological temperature, and suggest that parameters of functional Env stability may be helpful in the study and optimization of native Env mimetics and vaccines

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Late Recurrent Post-Transplant Primary Biliary Cirrhosis in British Columbia

Late recurrent primary biliary cirrhosis (PBC) following orthotopic liver transplant remains a controversial topic. The first documented case of recurrence occurring in 16 patients transplanted for PBC and followed at the authors' institution for longer than one year is presented. A 54-year-old man transplanted for PBC developed a cholestatic pattern of enzyme elevation on post-transplant day (PTD) 1305. Repeat antimitochondrial antibody was strongly positive (1:300 to 1:400). A liver biopsy revealed severe bile duct damage, lymphocytic cholangitis, focal periductal noncaseating granuloma and minimal endotheliitis. Recurrent PBC was diagnosed. At the time of orthotopic liver transplant this patient received induction immunosuppression with OKT3 crossed over to cyclosporine (CsA), azathioprine (AZA) and prednisone. AZA was discontinued early and maintenance CsA tapered to a target trough level of 150 to 200 ng/mL by PTD 365. Prednisone was withdrawn by PTD 664. CsA levels during PTDs 1225 to 1305 (before elevation of hepatobiliary enzymes) were below target at 114 to 166 ng/mL. Of the 16 patients, all but three were maintained on CsA, AZA and prednisone. One was on CsA (trough levels on target) and AZA; the other two, including the patient with recurrent PBC, were on CsA only. The trough CsA level of the patient without recurrent PBC has been within the target range. The authors speculate that the underlying defect in immunoregulation in PBC persists post-transplant and that in the patient without recurrent PBC this defect was unmasked by lowered maintenance immunosuppression - allowing recurrence of PBC in a previously stable liver allograft

T lymphocytes potentiate endogenous neuroprotective inflammation in a mouse model of ALS

Amyotrophic Lateral Sclerosis (ALS) is an adult-onset, progressive, motor neuron degenerative disease, in which the role of inflammation is not well established. Innate and adaptive immunity were investigated in the CNS of the Superoxide Dismutase 1 (SOD1)G93A transgenic mouse model of ALS. CD4+ and CD8+ T cells infiltrated SOD1G93A spinal cords during disease progression. Cell-specific flow cytometry and gene expression profiling showed significant phenotypic changes in microglia, including dendritic cell receptor acquisition, and expression of genes linked to neuroprotection, cholesterol metabolism and tissue remodeling. Microglia dramatically up-regulated IGF-1 and down-regulated IL-6 expression. When mutant SOD1 mice were bred onto a TCRβ deficient background, disease progression was significantly accelerated at the symptomatic stage. In addition, microglia reactivity and IGF-1 levels were reduced in spinal cords of SOD1G93A (TCRβ−/−) mice. These results indicate that T cells play an endogenous neuroprotective role in ALS by modulating a beneficial inflammatory response to neuronal injury

Substance P mediates inflammatory oedema in acute pancreatitis via activation of the neurokinin-1 receptor in rats and mice

1. Pancreatic oedema occurs early in the development of acute pancreatitis, and the overall extent of fluid loss correlates with disease severity. The tachykinin substance P (SP) is released from sensory nerves, binds to the neurokinin-1 receptor (NK1-R) on endothelial cells and induces plasma extravasation, oedema, and neutrophil infiltration, a process termed neurogenic inflammation. We sought to determine the importance of neurogenic mechanisms in acute pancreatitis. 2. Pancreatic plasma extravasation was measured using the intravascular tracers Evans blue and Monastral blue after administration of specific NK1-R agonists/antagonists in rats and NK1-R(+/+)/(−/−) mice. The effects of NK1-R genetic deletion/antagonism on pancreatic plasma extravasation, amylase, myeloperoxidase (MPO), and histology in cerulein-induced pancreatitis were characterized. 3. In rats, both SP and the NK1-R selective agonist [Sar(9) Met(O(2))(11)]SP stimulated pancreatic plasma extravasation, and this response was blocked by the NK1-R antagonist CP 96,345. Selective agonists of the NK-2 or NK-3 receptors had no effect. 4. In rats, cerulein stimulated pancreatic plasma extravasation and serum amylase. These responses were blocked by the NK1-R antagonist CP 96,345. 5. In wildtype mice, SP induced plasma extravasation while SP had no effect in NK1-R knockout mice. 6. In NK1-R knockout mice, the effects of cerulein on pancreatic plasma extravasation and hyperamylasemia were reduced by 60%, and pancreatic MPO by 75%, as compared to wildtype animals. 7. Neurogenic mechanisms of inflammation are important in the development of inflammatory oedema in acute interstitial pancreatitis