Investigation of HIV-1 Gag binding with RNAs and Lipids using Atomic Force Microscopy

Atomic Force Microscopy was utilized to study the morphology of Gag, {\Psi}RNA, and their binding complexes with lipids in a solution environment with 0.1{\AA} vertical and 1nm lateral resolution. TARpolyA RNA was used as a RNA control. The lipid used was phospha-tidylinositol-(4,5)-bisphosphate (PI(4,5)P2). The morphology of specific complexes Gag-{\Psi}RNA, Gag-TARpolyA RNA, Gag-PI(4,5)P2 and PI(4,5)P2-{\Psi}RNA-Gag were studied. They were imaged on either positively or negatively charged mica substrates depending on the net charges carried. Gag and its complexes consist of monomers, dimers and tetramers, which was confirmed by gel electrophoresis. The addition of specific {\Psi}RNA to Gag is found to increase Gag multimerization. Non-specific TARpolyA RNA was found not to lead to an increase in Gag multimerization. The addition PI(4,5)P2 to Gag increases Gag multimerization, but to a lesser extent than {\Psi}RNA. When both {\Psi}RNA and PI(4,5)P2 are present Gag undergoes comformational changes and an even higher degree of multimerization

A novel minimal in vitro system for analyzing HIV-1 Gag mediated budding

A biomimetic minimalist model membrane was used to study the mechanism and kinetics of cell-free in vitro HIV-1 Gag budding from a giant unilamellar vesicle (GUV). Real time interaction of Gag, RNA and lipid leading to the formation of mini-vesicles was measured using confocal microscopy. Gag forms resolution limited punctae on the GUV lipid membrane. Introduction of the Gag and urea to a GUV solution containing RNA led to the budding of mini-vesicles on the inside surface of the GUV. The GUV diameter showed a linear decrease in time due to bud formation. Both bud formation and decrease in GUV size were proportional to Gag concentration. In the absence of RNA, addition of urea to GUVs incubated with Gag also resulted in subvesicle formation but exterior to the surface. These observations suggest the possibility that clustering of GAG proteins leads to membrane invagination even in the absence of host cell proteins. The method presented here is promising, and allows for systematic study of the dynamics of assembly of immature HIV and help classify the hierarchy of factors that impact the Gag protein initiated assembly of retroviruses such as HIV.Comment: 27 pages, 9 Figures and 0 Table

A Discrete Two-Dimensional Model of a Loaded Cantilever Influenced by Time-Dependent Forces

We developed a discrete two-dimensional model of a cantilever which incorporates the effects of inhomogeneity, the geometry of an attached particle, and the influence of external time-dependent forces. We provide a comparison between the solutions for our discrete model and its continuous limit. The rotational-vibrational mode is studied in detail. The results of numerical simulations demonstrate usefulness of our model for many applications when a cantilever has a complicated geometry and is affected by time-depended and distributed external forces.Comment: Pages 15, Figures 1

Demonstration of the asymmetric lateral Casimir force between corrugated surfaces in the nonadditive regime

The measurement of the lateral Casimir force between two aligned sinusoidally corrugated Au-coated surfaces has been performed in the nonadditive regime. The use of deeper corrugations also allowed to demonstrate an asymmetry in the phase dependences of the lateral Casimir force, as predicted earlier. The measurement data are found to be in excellent agreement with the exact theoretical results computed at T=300 K including effect of real material properties. The deviations between the exact theory and the proximity force approximation are quantified. The obtained results are topical for applications in nanomachines.Comment: 9 pages, 3 figure

Epstein-Barr virus associated haemophagocytic lymphohistiocytosis treated with anakinra and rituximab: A case report

BACKGROUND: Haemophagocytic lymphohistiocytosis (HLH) is a severe, life-threatening syndrome characterised by hyperinflammation and macrophage activation. Viral infections such as Epstein-Barr virus (EBV) are a well-recognised trigger of HLH but the treatment of such cases is not well-defined. We present a case of primary EBV driven HLH that was successfully treated with the interleukin-1 inhibitor anakinra in addition to rituximab and high-dose steroids. CASE: A 22-year-old female with no past medical history developed a mononucleosis-like illness lasting five days characterised by fevers, sore throat and neck swelling. Two weeks following this she presented with fevers, night sweats, fatigue and right upper quadrant pain. She was diagnosed with HLH based on high fevers with hyperferritaemia, hypertriglyceridaemia, pancytopaenia, abnormal liver function tests and hepatosplenomegaly. Extensive investigation revealed an EBV viral load of 23,000,000 copies/ml with nil other obvious triggers. A diagnosis of primary-driven EBV HLH was made. She was treated with the interleukin-1 inhibitor anakinra, methylprednisolone and IVIG and a single dose of rituximab. Following the commencement of treatment, the patient made a dramatic improvement. Her EBV viral load reduced to 660 within nine days and her blood counts and liver function returned to normal. She was discharged from hospital on day sixteen. She continued the anakinra for 5 weeks at a weaning dose and completed a 12-week weaning dose of steroids. She has returned to her studies and has no lasting complications from her illness. DISCUSSION: This case highlights the potential of primary EBV infection to cause fulminant HLH. The prompt diagnosis and treatment of HLH using anakinra and rituximab in addition to conventional HLH treatment was safe, and associated with a dramatic clinical improvement. The use of anakinra has been documented in other cases of HLH but none, to our knowledge, of primary EBV-driven HLH with no underlying haematological or rheumatological condition

New constraints for non-Newtonian gravity in nanometer range from the improved precision measurement of the Casimir force

We obtain constraints on non-Newtonian gravity following from the improved precision measurement of the Casimir force by means of atomic force microscope. The hypothetical force is calculated in experimental configuration (a sphere above a disk both covered by two metallic layers). The strengthenings of constraints up to 4 times comparing the previous experiment and up to 560 times comparing the Casimir force measurements between dielectrics are obtained in the interaction range 5.9 nm$\leq\lambda\leq 115$nm. Recent speculations about the presence of some unexplained attractive force in the considered experiment are shown to be unjustified.Comment: 5 pages, 1 figur

5-fluorouracil and folinic acid-induced mucositis: no effect of oral glutamine supplementation.

In some clinical situations the endogenous production of glutamine may be insufficient to maintain optimal tissue structure and function such that glutamine becomes a conditionally essential amino acid. Studies in laboratory animals have demonstrated that glutamine supplementation can reduce the incidence and severity of cytotoxic-induced mucositis. This study examined the role of oral glutamine supplementation in the management of mucositis caused by 5-fluorouracil (5-FU) and folinic acid. Twenty-eight patients with gastrointestinal cancers were randomised to receive 16 g of glutamine per day for 8 days, or placebo, in a randomised double-blind trial before crossing over to the alternative supplement during the second treatment cycle. The supplement was well tolerated with no apparent adverse effects, but failed to have any significant effect on oral mucositis assessed by the patients or investigator. The possible reasons for this apparent lack of benefit are discussed

Experimental approaches to the difference in the Casimir force through the varying optical properties of boundary surface

We propose two novel experiments on the measurement of the Casimir force acting between a gold coated sphere and semiconductor plates with markedly different charge carrier densities. In the first of these experiments a patterned Si plate is used which consists of two sections of different dopant densities and oscillates in the horizontal direction below a sphere. The measurement scheme in this experiment is differential, i.e., allows the direct high-precision measurement of the difference of the Casimir forces between the sphere and sections of the patterned plate or the difference of the equivalent pressures between Au and patterned parallel plates with static and dynamic techniques, respectively. The second experiment proposes to measure the Casimir force between the same sphere and a VO${}_2$ film which undergoes the insulator-metal phase transition with the increase of temperature. We report the present status of the interferometer based variable temperature apparatus developed to perform both experiments and present the first results on the calibration and sensitivity. The magnitudes of the Casimir forces and pressures in the experimental configurations are calculated using different theoretical approaches to the description of optical and conductivity properties of semiconductors at low frequencies proposed in the literature. It is shown that the suggested experiments will aid in the resolution of theoretical problems arising in the application of the Lifshitz theory at nonzero temperature to real materials. They will also open new opportunities in nanotechnology.Comment: 23 pages of the text, 2 tables, and captions of 12 figures (to appear in Phys. Rev. A