HIV-1 gp120 Induces Expression of IL-6 through a Nuclear Factor-Kappa B-Dependent Mechanism: Suppression by gp120 Specific Small Interfering RNA

In addition to its role in virus entry, HIV-1 gp120 has also been implicated in HIV-associated neurocognitive disorders. However, the mechanism(s) responsible for gp120-mediated neuroinflammation remain undefined. In view of increased levels of IL-6 in HIV-positive individuals with neurological manifestations, we sought to address whether gp120 is involved in IL-6 over-expression in astrocytes. Transfection of a human astrocyte cell line with a plasmid encoding gp120 resulted in increased expression of IL-6 at the levels of mRNA and protein by 51.3±2.1 and 11.6±2.2 fold respectively; this effect of gp120 on IL-6 expression was also demonstrated using primary human fetal astrocytes. A similar effect on IL-6 expression was observed when primary astrocytes were treated with gp120 protein derived from different strains of X4 and R5 tropic HIV-1. The induction of IL-6 could be abrogated by use of gp120-specific siRNA. Furthermore, this study showed that the NF-κB pathway is involved in gp120-mediated IL-6 over-expression, as IKK-2 and IKKβ inhibitors inhibited IL-6 expression by 56.5% and 60.8%, respectively. These results were also confirmed through the use of NF-κB specific siRNA. We also showed that gp120 could increase the phosphorylation of IκBα. Furthermore, gp120 transfection in the SVGA cells increased translocation of NF-κB from cytoplasm to nucleus. These results demonstrate that HIV-1 gp120-mediated over-expression of IL-6 in astrocytes is one mechanism responsible for neuroinflammation in HIV-infected individuals and this is mediated by the NF-κB pathway

Modified SOGI based shunt active power filter to tackle various grid voltage abnormalities

Shunt Active Power Filters (SAPF) have been effectively used to compensate the harmonics generated by the non-linear loads. The SAPF’s performance depends on the accurate generation of reference current, which is dependent greatly on the template of supply voltage. When the grid voltage (or its template) is characterized by different abnormalities like presence of harmonics, imbalance, dc-offset etc., some of the conventional techniques of frequency estimation may fail to correctly estimate the frequency. This ultimately affects the reference current generation and hence, the SAPF operation, ultimately leading to high distortion of the grid currents. The paper presents modified dual second-order generalized integrator (MDSOGI) based SAPF to ensure effective compensation of harmonics, even when the grid voltage is characterized by all the abnormalities mentioned above. It is highlighted with one case that when the sensed voltage is having dc-offset, DSOGI-SAPF results into the source current with THD, dc-offset and harmonic with values 5.82%, 0.8% and 4.5%, respectively. For the same case, the proposed technique yields grid current which is free of dc-offset and 2nd harmonic and has THD = 3.57%. The dynamic performance of the MDSOGI-SAPF is validated and its superior performance over DSOGI-SAPF is illustrated even with experimental results

Modified Dual Second-order Generalized Integrator FLL for Frequency Estimation Under Various Grid Abnormalities

Proper synchronization of Distributed Generator with grid and its performance in grid-connected mode relies on fast and precise estimation of phase and amplitude of the fundamental component of grid voltage. However, the accuracy with which the frequency is estimated is dependent on the type of grid voltage abnormalities and structure of the phase-locked loop or frequency locked loop control schemes. Among various control schemes, second-order generalized integrator based frequency- locked loop (SOGI-FLL) is reported to have the most promising performance. It tracks the frequency of grid voltage accurately even when grid voltage is characterized by sag, swell, harmonics, imbalance, frequency variations etc. However, estimated frequency contains low frequency oscillations in case when sensed grid-voltage has a dc offset. This paper presents a modified dual second-order generalized integrator frequency-locked loop (MDSOGI-FLL) for three-phase systems to cope with the non-ideal three-phase grid voltages having all type of abnormalities including the dc offset. The complexity in control scheme is almost the same as the standard dual SOGI-FLL, but the performance is enhanced. Simulation results show that the proposed MDSOGI-FLL is effective under all abnormal grid voltage conditions. The results are validated experimentally to justify the superior performance of MDSOGI-FLL under adverse conditions

Modified Dual Second-order Generalized Integrator FLL for Frequency Estimation Under Various Grid Abnormalities

Proper synchronization of Distributed Generator with grid and its performance in grid-connected mode relies on fast and precise estimation of phase and amplitude of the fundamental component of grid voltage. However, the accuracy with which the frequency is estimated is dependent on the type of grid voltage abnormalities and structure of the phase-locked loop or frequency locked loop control schemes. Among various control schemes, second-order generalized integrator based frequency- locked loop (SOGI-FLL) is reported to have the most promising performance. It tracks the frequency of grid voltage accurately even when grid voltage is characterized by sag, swell, harmonics, imbalance, frequency variations etc. However, estimated frequency contains low frequency oscillations in case when sensed grid-voltage has a dc offset. This paper presents a modified dual second-order generalized integrator frequency-locked loop (MDSOGI-FLL) for three-phase systems to cope with the non-ideal three-phase grid voltages having all type of abnormalities including the dc offset. The complexity in control scheme is almost the same as the standard dual SOGI-FLL, but the performance is enhanced. Simulation results show that the proposed MDSOGI-FLL is effective under all abnormal grid voltage conditions. The results are validated experimentally to justify the superior performance of MDSOGI-FLL under adverse conditions

Inhibition of gp120-induced IL-6 expression by gp120-specific siRNA.

<p>Four gp120 siRNA (A) were designed using Ambion software and commercially synthesized by Ambion Inc. Only positive strand sequences are shown in the figure. Astrocytes were transfected with 50 nmoles each of scrambled or one of the 4 different siRNA for 48 hours before gp120 transfection. IL-6 mRNA (B) and protein (C) was measured at 6 and 48 hours, post-gp120 transfection. The mRNA is presented as relative percent mRNA expression with gp120 transfected cells as positive control. The protein concentration is presented as relative percent expression. Each bar represents mean ± SE of 3 experiments with each experiment done in triplicates. The statistical significance was calculated using student's t test and ** denotes p value of ≤0.01.</p

gp120-mediated increase of IL-6 expression in SVGA astrocyte cells.

<p>1×10⁶ SVGA astrocytes were transfected with 1 µg gp120 JR-FL (R5 tropic) DNA using Lipofectamine™ 2000. The cells were harvested at the times described in the text. Mock transfection was performed with transfection of equal amount of empty human vector pcDNA3.1. All times noted on the figure and listed in the text are the times at which cells and supernatants were harvested after the end of the 5 hour transfection protocol. IL-6 mRNA (A) and IL-6 protein (B) expression levels were measured using real time RT-PCR and bioplex assays respectively. In (B), open bars show empty-vector transfected mock controls and closed bars show gp120 transfected samples. (C) shows the effect of exogenous gp120 on SVGA cells. SVGA were exposed to 20 nM recombinant gp120IIIB for various lengths of time and total mRNA was isolated. IL-6 expression levels were measured using real time RT-PCR. The mRNA levels are presented as fold difference between gp120 transfected cells and control cells transfected with empty plasmid. The protein concentration is presented as ng/ml protein in supernatant. Each bar represents mean ± SE of 3 experiments with each experiment done in triplicates. The statistical significance was calculated using student's t test and * and ** denotes p value of ≤0.05 and ≤0.01, respectively.</p

gp120 mediated increase in phosphorylation of IκB-α in primary human fetal astrocytes.

<p>Primary human astrocytes from two donors were treated with 20 nM gp120 IIIB, cells were collected and lysed using RIPA buffer at different times. These proteins were electrophoresed on 10% SDS gel and transferred to PVDF membrane. Antibodies against p-IκBα, total IκBα and β-actin were used for western blotting and membrane was read on densitometer. Values below the lanes show band intensities of the respective bands for both the donors (A & B). The increase in p-IκB-α was estimated by calculating the ratios of p-IκB-α to total IκB-α. Actin was used as loading control.</p