An increase in glycoprotein concentration on extracellular virions dramatically alters vaccinia virus infectivity and pathogenesis without impacting immunogenicity

The extracellular virion (EV) form of Orthopoxviruses is required for cell-to-cell spread and pathogenesis, and is the target of neutralizing antibodies in the protective immune response. EV have a double envelope that contains several unique proteins that are involved in its intracellular envelopment and/or subsequent infectivity. One of these, F13, is involved in both EV formation and infectivity. Here, we report that replacement of vaccinia virus F13L with the molluscum contagiosum virus homolog, MC021L results in the production of EV particles with significantly increased levels of EV glycoproteins, which correlate with a small plaque phenotype. Using a novel fluorescence-activated virion sorting assay to isolate EV populations based on glycoprotein content we determine that EV containing either higher or lower levels of glycoproteins are less infectious, suggesting that there is an optimal concentration of glycoproteins in the outer envelope that is required for maximal infectivity of EV. This optimal glycoprotein concentration was required for lethality and induction of pathology in a cutaneous model of animal infection, but was not required for induction of a protective immune response. Therefore, our results demonstrate that there is a sensitive balance between glycoprotein incorporation, infectivity, and pathogenesis, and that manipulation of EV glycoprotein levels can produce vaccine vectors in which pathologic side effects are attenuated without a marked diminution in induction of protective immunity.

Assessment and Comparison of Subseasonal Bias and Forecast Skill in the Unified Forecast System (UFS) Benchmarks 3 and 5

<p>A series of reforecasts have been generated with prototype versions of the coupled Unified Forecast System (UFS) to evaluate progress in the model development. The forecast skill and biases of the UFS Prototypes 3 and 5 reforecast sets—called Benchmark 3 and Benchmark 5, respectively—are analyzed and compared with the NCEP Climate Forecast System version 2 (CFSv2) reforecasts from the Subseasonal Prediction Experiment (SubX). The evaluation focuses on surface variables typically provided in the subseasonal outlooks at weekly-averaged timescales, namely 2-meter air temperature, precipitation rate, and sea surface temperature. Additional assessment of the structure of the systematic error in total diabatic heating over three broad layers of the atmosphere (850-650 hPa, 650-450 hPa and 450-50 hPa) has been performed as a function of season and forecast lead. In terms of forecast skill, all models still experience a skill drop-off of varying degree by week 3. In general, however, the UFS prototypes considerably reduce the marked diminution of variability with lead time displayed in their predecessor, CFSv2. Moreover, the prototypes have reduced systematic error compared to CFSv2, particularly for 2-meter temperature and precipitation. A systematic overestimate of diabatic cooling is noted in the upper atmosphere (diabatic heating too negative compare to ERA-5 estimates) during boreal winter. </p>

Cystotoperitoneal shunt surgery during infancy in porencepahalic cyst located in frontal region led to regaining of developmental milestone

Porencepahalic cyst is considered as an extremely uncommon developmental disorder of the central nervous system, being characterized by the presence of a fluid-filled cysts or cavities located within the cerebral hemispheres. It can be associated with varied aetiology and can present with a spectrum of clinical presentation varying from asymptomatic to grossly spastic limbs, mental retardation, cognitive impairment and intractable seizure. Extensive Pubmed and Medline search did not yield any result when searched for term “infancy, porencepahalic cyst, cystoperitonreal shunt.” However, clear guideline for management is still lacking. Authors report an interesting case of giant porencepahalic cyst located in the right frontal region in infancy and underwent cystoperitonreal shunt surgery, which lead to good outcome with remarkable recovery of delayed milestones with adequate scholastic performances along with marked diminution in the size of porencepahalic cyst.

Splenic Infarction in a Patient with Spent-Phase Polycythemia Rubra Vera (PRV) Treated with Lenalidomide.

Introduction: Lenalidomide (LD) is an oral thalidomide analog with both immunomodulatory and anti-angiogenic properties. It is approved for use in myelodysplastic syndromes (MDS), specifically low-risk and intermediate 1 with 5Q-. LD has also been approved for relapsed multiple myeloma (MM). A major complication associated with LD includes venous thromboembolism (VTE), especially in combination with steroid and erythropoetin (EPO). VTE occurs in 2% of MDS, but range from 6.6% as monotherapy in MM to 8.5%–13.5% when combined with dexamethasone. Zonder reports a preliminary incidence of 75% (9/12 patients) in the SWOG 0302 trial of LD and dexamethasone in previously untreated myeloma. With the addition of ASA in these patients the incidence dropped to 19%. Patients with polycythemia rubra vera (PRV) have a higher incidence of VTE (5.1%> age 70). JAK-2 mutation (tyrosine kinase mutation) occurs as a sporadic event in 75% of patients with PRV and is associated with an older age group, inferior survival, myelofibrosis, acute leukemia and thromboembolic events. Recent studies with LD in this group of PRV and spent-phase myelofibrosis have shown encouraging results. LD has an activity of 46% in myelofibosis by decreasing transfusions and splenomegaly. It would seem to have activity in spent-phase PRV. We report a seminal event of splenic infarction in a patient with PRV and myelofibrosis treated with LD. Case Presentation: A 73 y/o male was diagnosed with PRV 16 yrs earlier. He underwent phlebotomy and received ASA. With a hyperproliferative phase he received hydrea as he was intolerant of IFN-A and anagrelide. Three months earlier, he developed fever, sweats, splenomegaly, and became transfusion dependent. His marrow confirmed myelofibrosis with dysplastic features. LD was begun at 25 mg daily. Three weeks later he developed pneumonia with bronchospasm and required hospitalization for antibiotics and steroids. He improved but returned 4 days later with left upper quadrant pain with CT findings consistent with multiple splenic infarcts. Prednisone was stopped and enoxaparin resulted in marked improvement. Currently, he has marked diminution in the size of his spleen from 13 cm to 4 cm. Currently, he is transfusion independent with marked clinical improvement. Discussion: This report points several dynamic issues within the field of myeloproliferative syndromes. The JAK-2 mutation offers fertile areas of research within MPD disorders as a causative or sporadic mutation and as an associated with several adverse events including VTE and myelofibrosis. We report here a unique occurrence of VTE in the form of multiple splenic infarctions within the context of treatment of the spent-phase of PVR with JAK-2 mutation receiving LD. VTE events associated with LD are associated with DVT or pulmonary embolus and have not been associated with splenic infarction. Splenic infarction has also been reported in myelofibrosis. As further studies confirm efficacy with myelofibrosis it should be suggested that prophylaxis with warfarin or enoxaparin, especially in JAK-2 mutation patients, be considered. Physicians should be encouraged to report their experience in patients with MPD and lenalidomide.

Temporal Analysis of the Antigenic Composition of Borrelia burgdorferi during Infection in Rabbit Skin

ABSTRACT The numbers of host-adapted Borrelia burgdorferi (HAB) organisms in rabbit skin were assessed by real-time PCR over the first 3 weeks of infection. Maximal numbers were found at day 11, while spirochete numbers decreased by more than 30-fold by day 21. The antigenic composition of HAB in skin biopsy samples was determined by use of a procedure termed hydrophobic antigen tissue Triton extraction. Immune sera from rabbits, sera from chronically infected mice, and monospecific antiserum to the antigenic variation protein, VlsE, were used to probe parallel two-dimensional immunoblots representing each time point. Individual proteins were identified using either specific antisera or by matching protein spots to mass spectrometry-identified protein spots from in vitro-cultivated Borrelia. There were significant changes in the relative expression of a variety of known and previously unrecognized HAB antigens during the 21-day period. OspC and the outer membrane proteins OspA and OspB were prominent at the earliest time point, day 5, when the antigenic variation protein VlsE was barely detected. OspA and OspB were not detected after day 5. OspC was not detected after day 9. VlsE was the most prominent antigen from day 7 through day 21. BmpA, ErpN, ErpP, LA7, OppA-2, DbpA, and an unidentified 15-kDa protein were also detected from day 7 through day 21. Immunoblot analysis using monospecific anti-VlsE revealed the presence of prominent distinct VlsE lower forms in HAB at days 9, 11, and 14; however, these lower forms were no longer detected at day 21. This marked diminution in VlsE lower forms paralleled the clearance of the spirochete from skin.

Changes in the Distribution of Anionic Constituents in Secretory Granules of Mouse Pancreatic Acinar Cells After Pilocarpine-induced Degranulation

We used cationized colloidal gold (CCG) to investigate the distribution of anionic sites in different secretory granules of mouse pancreatic acinar cell regranulation. Localization of anionic sites with CCG was carried out on ultrathin sections of a mouse pancreas, fixed in Karnovsky's fixative and OsO4 and embedded in Araldite. After pilocarpinestimulated degranulation, there was a marked diminution in the anionic charge density of immature and mature granules of the 4-hr group (≈43.0 gold particles/μm2) compared to the 8-hr mature granules group (≈64.6 gold particles/μm2). Scattergram analysis to investigate the correlation between section profile size and cationized gold labeling density revealed a reverse correlation, the small granule profiles demonstrated a higher density compared to the larger profiles of the same group. On the basis of these observations, it appears that a post-translational processing of secretory content influences the granule anionic charge and thus may affect the intragranular buffer capacity.

Long-Term Histologic Effects of Inferior Turbinate Laser Surgery

OBJECTIVE: In this study we sought to define the histologic changes produced by laser treatment of inferior turbinates. STUDY DESIGN: Eight inferior turbinates with prior laser treatment (mean, 26.8 months) were analyzed by light microscopy after turbinectomy for relief of refractory nasal obstruction. Histologic findings were compared with those of a group of 8 hypertrophic inferior turbinates that had no previous laser surgery. RESULTS: Laser-treated areas of the inferior turbinates demonstrated a histologically bland appearance, with marked diminution of seromucinous glands and relative preponderance of connective tissue matrix. Prominence of venous sinusoids was also significantly reduced in the laser-treated areas. Surface epithelium including goblet cells was reconstituted over the areas of laser application. CONCLUSION: Clinical laser surgery of the inferior turbinate produces striking long-term histologic changes. SIGNIFICANCE: The data suggest a differential response of turbinate histologic components to application of laser energy, with the glandular component being particularly sensitive. Further correlative study is needed to clarify the clinical significance of laser-induced histologic changes in inferior turbinates.

Peroxynitrite-induced apoptosis involves activation of multiple caspases in HL-60 cells

In this study, we show that caspases 2, 3, 6, and 7 were activated during peroxynitrite-induced apoptosis in human leukemia HL-60 cells and that processing of these caspases was accompanied by cleavage of poly(ADP-ribose) polymerase and lamin B. Treatment of cells with DEVD-fluoromethyl ketone (FMK), a selective inhibitor for caspase 3-like proteases, resulted in a marked diminution of apoptotic cells. VAVAD-FMK, an inhibitor of caspase 2, partially inhibited the apoptotic response to peroxynitrite. However, selective inactivation of caspase 6 by VEID-FMK did not affect apoptosis rates. These data suggest that caspase 3-like proteases and caspase 2, but not caspase 6, are required for peroxynitrite-induced apoptosis in this cell type. Moreover, we demonstrate that peroxynitrite treatment stimulated activation of caspases 8 and 9, two initial caspases in the apoptotic signaling pathway, and preincubation of cells with their inhibitor, IETD-FMK, inhibited activation of caspase 3-like proteases and caspase 2 at the concentration that prevents the apoptosis. These observations, together, suggest that caspase 8 and/or caspase 9 mediates activation of caspase 3-like proteases and caspase 2 during the apoptosis induced by peroxynitrite in HL-60 cells.

Fetal Music Perception: The Role of Sound Transmission

The fetal sound environment is now known to be rich and varied. Playback of tapes made from intrauterine recordings of sounds reveals some muffling, suggesting an attenuation of high-frequency sounds at the surface of the abdominal wall and during transmission through abdominal and uterine tissues and fluids. The present experiments show how the spectral features of synthesized musical sounds are altered once they reach the ear of the fetal sheep. Below 300 Hz, intrauterine sound pressure levels are nearly identical to those recorded outside the ewe. Between 315 and 2500 Hz, the attenuation increases at a rate of 5 dB per octave. Spectral analyses of trumpet and flugelhorn sounds recorded in utero show a marked diminution in sound pressure level in partials above 600 Hz; this diminution could be perceived by the fetus as an altered timbre.

The effect of extracellular calcium elevation on morphology and function of isolated rat osteoclasts

Osteoclasts are large multinucleate cells unique in their capacity to resorb bone. These cells are exposed locally to high levels of ionised calcium during the process of resorption. We have therefore examined the effect of elevated extracellular calcium on the morphology and function of freshly disaggregated rat osteoclasts. Cell size and motility were quantitated by time-lapse video recording together with digitisation and computer-centred image analysis. In order to assess the resorptive capacity of isolated osteoclasts, we measured the total area of resorption of devitalised cortical bone by means of scanning electron microscopy and computer-based morphometry. The results show that elevation of the extracellular calcium concentration causes a dramatic reduction of cell size, accompanied by a marked diminution of enzyme release and abolition of bone resorption. We propose that ionised calcium might play an important role in the local regulation of osteoclastic bone resorption.