Rabies Virus Infection Induces Type I Interferon Production in an IPS-1 Dependent Manner While Dendritic Cell Activation Relies on IFNAR Signaling

As with many viruses, rabies virus (RABV) infection induces type I interferon (IFN) production within the infected host cells. However, RABV has evolved mechanisms by which to inhibit IFN production in order to sustain infection. Here we show that RABV infection of dendritic cells (DC) induces potent type I IFN production and DC activation. Although DCs are infected by RABV, the viral replication is highly suppressed in DCs, rendering the infection non-productive. We exploited this finding in bone marrow derived DCs (BMDC) in order to differentiate which pattern recognition receptor(s) (PRR) is responsible for inducing type I IFN following infection with RABV. Our results indicate that BMDC activation and type I IFN production following a RABV infection is independent of TLR signaling. However, IPS-1 is essential for both BMDC activation and IFN production. Interestingly, we see that the BMDC activation is primarily due to signaling through the IFNAR and only marginally induced by the initial infection. To further identify the receptor recognizing RABV infection, we next analyzed BMDC from Mda-5−/− and RIG-I−/− mice. In the absence of either receptor, there is a significant decrease in BMDC activation at 12h post infection. However, only RIG-I−/− cells exhibit a delay in type I IFN production. In order to determine the role that IPS-1 plays in vivo, we infected mice with pathogenic RABV. We see that IPS-1−/− mice are more susceptible to infection than IPS-1+/+ mice and have a significantly increased incident of limb paralysis

Enhancement of Sf9 cells and baculovirus production employing Grace's medium supplemented with milk whey ultrafiltrate

Animal cells can be cultured both in basal media supplemented with fetal bovine serum (FBS) and in serum-free media. In this work, the supplementation of Grace's medium with a set of nutrients to reduce FBS requirements in Spodoptera frugiperda (Sf9) cell culture was evaluated, aiming the production of Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) at a cost lower than those for the production using Sf900 II medium. In Grace's medium supplemented with glucose, Pluronic F68 (PF68) and yeast extract (YE), the effects of FBS and milk whey ultrafiltrate (MWU) on cell concentration and viability during midexponential and stationary growth phase were evaluated. In spite of the fact that FBS presented higher statistical effects than MWU on all dependent variables in the first cell passage studies, after cell adaptation, AgMNPV polyhedra production was comparable to that in Sf900 II. Batch cultivation in Grace's medium with 2.7 g l(-1) glucose, 8 g l(-1) YE and 0.1% (w/v) PF68 supplemented with 1% (w/v) MWU and 3% (v/v) FBS increased viable cell concentration to about 5-fold (4.7x10(6) cells ml(-1)) when compared to Grace's containing 10% (v/v) FBS (9.5x10(5) cells ml(-1)). AgMNPV polyhedra (PIBs) production was around 3-fold higher in the MWU supplemented medium (1.6x10(7) PIBs ml(-1)) than in Grace's medium with 10% FBS (0.6x10(7) PIBs ml(-1)). This study therefore shows a promising achievement to significantly reduce FBS concentration in Sf9 insect cell media, keeping high productivity in terms of cell concentration and final virus production at a cost almost 50% lower than that observed for Sf900 II medium.4911

Comparing BRIN-BD11 culture producing insulin using different type of microcarriers

This research was conducted to examine the growth profile, growth kinetics, and insulin-secretory responsiveness of BRIN-BD11 cells grown in optimized medium on different types of microcarriers (MCs). Comparisons were made on modified polystyrene (Hillex® II) and crosslinked polystyrene Plastic Plus (PP) from Solohill Engineering. The cell line producing insulin was cultured in a 25 cm2 T-flask as control while MCs based culture was implemented in a stirred tank bioreactor with 1 L working volume. For each culture type, the viable cell number, glucose, lactate, glutamate, and insulin concentrations were measured and compared. Maximum viable cell number was obtained at 1.47 × 105 cell/mL for PP microcarrier (PPMCs) culture, 1.35 × 105 cell/mL Hillex® II (HIIMCs) culture and 0.95 × 105 cell/mL for T-flask culture, respectively. The highest insulin concentration has been produced in PPMCs culture (5.31 mg/L) compared to HIIMCs culture (2.01 mg/L) and T-flask culture (1.99 mg/L). Therefore overall observation suggested that PPMCs was likely preferred to be used for BRIN-BD11 cell culture as compared with Hillex® II MCs