The cardioprotective mechanism of phenylaminoethyl selenides (PAESe) against doxorubicin-Induced cardiotoxicity involves frataxin

Doxorubicin (DOX) is an anthracycline cancer chemotherapeutic that exhibits cumulative dose-limiting cardiotoxicity and limits its clinical utility. DOX treatment results in the development of morbid cardiac hypertrophy that progresses to congestive heart failure and death. Recent evidence suggests that during the development of DOX mediated cardiac hypertrophy, mitochondrial energetics are severely compromised, thus priming the cardiomyocyte for failure. To mitigate cumulative dose (5 mg/kg, QIW x 4 weeks with 2 weeks recovery) dependent DOX, mediated cardiac hypertrophy, we applied an orally active selenium based compound termed phenylaminoethyl selenides (PAESe) (QIW 10 mg/kg x 5) to our animal model and observed that PAESe attenuates DOX-mediated cardiac hypertrophy in athymic mice, as observed by MRI analysis. Mechanistically, we demonstrated that DOX impedes the stability of the iron-sulfur cluster biogenesis protein Frataxin (FXN) (0.5 fold), resulting in enhanced mitochondrial free iron accumulation (2.5 fold) and reduced aconitase activity (0.4 fold). Our findings further indicate that PAESe prevented the reduction of FXN levels and the ensuing elevation of mitochondrial free iron levels. PAESe has been shown to have anti-oxidative properties in part, by regeneration of glutathione levels. Therefore, we observed that PAESe can mitigate DOX mediated cardiac hypertrophy by enhancing glutathione activity (0.4 fold) and inhibiting ROS formation (1.8 fold). Lastly, we observed that DOX significantly reduced cellular respiration (basal (5%) and uncoupled (10%)) in H9C2 cardiomyoblasts and that PAESe protects against the DOX-mediated attenuation of cellular respiration. In conclusion, the current study determined the protective mechanism of PAESe against DOX mediated myocardial damage and that FXN is implicitly involved in DOX-mediated cardiotoxicity

Lactobacillus plantarum DK119 as a Probiotic Confers Protection against Influenza Virus by Modulating Innate Immunity

Lactobacillus plantarum DK119 (DK119) isolated from the fermented Korean cabbage food was used as a probiotic to determine its antiviral effects on influenza virus. DK119 intranasal or oral administration conferred 100% protection against subsequent lethal infection with influenza A viruses, prevented significant weight loss, and lowered lung viral loads in a mouse model. The antiviral protective efficacy was observed in a dose and route dependent manner of DK119 administration. Mice that were treated with DK119 showed high levels of cytokines IL-12 and IFN-c in bronchoalveolar lavage fluids, and a low degree of inflammation upon infection with influenza virus. Depletion of alveolar macrophage cells in lungs and bronchoalveolar lavages completely abrogated the DK119-mediated protection. Modulating host innate immunity of dendritic and macrophage cells, and cytokine production pattern appeared to be possible mechanisms by which DK119 exhibited antiviral effects on influenza virus infection. These results indicate that DK119 can be developed as a beneficial antiviral probiotic microorganism

Harnessing Liquid Crystal Sensors for High-Throughput Real-Time Detection of Structural Changes in Lysozyme during Refolding Processes

Despite the rapid advances in process analytical technology, the assessment of protein refolding efficiency has largely relied on off-line protein-specific assays and/or chromatographic procedures such as reversed-phase high-performance liquid chromatography and size exclusion chromatography. Due to the inherent time gap pertaining to traditional methods, exploring optimum refolding conditions for many recombinant proteins, often expressed as insoluble inclusion bodies, has proven challenging. The present study describes a novel protein refolding sensor that utilizes liquid crystals (LCs) to discriminate varying protein structures during unfolding and refolding. An LC layer containing 4-cyano-4′-pentylbiphenyl (5CB) intercalated with 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) is used as a sensing platform, and its proof-of-concept performance is demonstrated using lysozyme as a model protein. As proteins unfold or refold, a local charge fluctuation at their surfaces modulates their interaction with zwitterionic phospholipid DOPE. This alters the alignment of DOPE molecules at the aqueous/LC interface, affecting the orientational ordering of bulk LC (i.e., homeotropic to planar for refolding and planar to homeotropic for unfolding). Differential polarized optical microscope images of the LC layer are subsequently generated, whose brightness directly linked to conformational changes of lysozyme molecules is quantified by gray scale analysis. Importantly, our LC-based refolding sensor is compatible with diverse refolding milieus for real-time analysis of lysozyme refolding and thus likely to facilitate the refolding studies of many proteins, especially those lacking a method to determine structure-dependent biological activity

Pretreatment of a low dose of DK119(10⁷ CFU) confers improved protection by preventing severe weight loss.

<p>A group of mice (n = 6) was once treated intranasally with a low dose of DK119 (10⁷ CFU per mouse). After 4 days of DK119 treatment, mice were infected with A/PR8 H1N1 virus (2.5 LD₅₀) in the presence of DK119 (10⁹ CFU per mouse). The control group was infected with A/PR8 virus without DK119 and no pretreatment. Virus(V): mice infected with A/PR8 H1N1 virus, DK119(10⁷+10⁹)+V: mice pretreated with DK119 and infected with DK119 and A/PR8 H1N1 virus.DK119(10⁷+10⁹): mice treated with DK119 (10⁷ CFU and 10⁹ CFU per mouse). Bars indicate standard errors.</p

Effects of DK119 (10⁸ CFU) treatment on preventing weight loss against influenza H1N1 virus infection.

<p>A group of mice (n = 6) was treated with DK119 (10⁸ CFU/mouse) at 4 days and 1 day prior to infection with A/PR8 virus (2.5 LD₅₀) in the presence of a low dose of DK119 (10⁸ CFU per mouse). Virus(V): mice infected with A/PR8 virus, DK119 (10⁸+10⁸+10⁸)+V: mice pretreated twice with DK119 and infected A/PR8 H1N1 virus in the presence of with DK119 (10⁸ CFU per mouse). Bars indicate standard errors.DK119 (10⁸+10⁸+10⁸): mice treated with DK119 alone (10⁸ CFU/mouse at 3 different time points without virus infection).</p

Oral administration of DK119 (10⁹ CFU) has protective effects against influenza virus infection.

<p>Mice (n = 5) were administered DK119 (10⁹CFU/mouse daily) orally for 10 days before infection and for 14 days after infection with A/PR8 H1N1 virus (2.5 LD₅₀), and their body weight changes were recorded. Virus(V): A/PR8 virus infection, DK119(10⁹)+V: Oral administration with DK119 and then virus infection. Bars indicate standard errors. DK119(10⁹): mice treated orally with DK119 alone (10⁹ CFU/mouse daily for 14 days).</p

Effects of DK119 pretreatment on cytokine production upon influenza virus infection.

<p>Levels of cytokines in bronchoalveolar lavage fluids (BALF) collected from mice (n = 3) at day 4 post infection with A/PR8 virus are presented in pg/ml as determined using cytokine ELISA. (A) IL-4, (B) IL-6, (C) TNF-α, (D) IL-12, (E) IFN-γ. DK119 (10?8): Intranasal DK119 10?8 control in the absence of A/PR8 virus infection. DK119 (10?9): Intranasal DK119 10?9 control in the absence of A/PR8 virus infection. (10?7)+V: DK119 (10?7) intranasal pretreatment and then A/PR8 virus infection in the presence of DK119 (10?9). (10?8)+V: DK119 (10?8) intranasal pretreatment and then A/PR8 virus infection in the presence of DK119 (10?9). Virus: Naïve mice infected with A/PR8 virus (2.5 LD₅₀). * indicates statistical significance compared to the control virus group (p<0.05). Bars indicate standard errors.</p

Intranasal administration of clodronate-liposomes selectively depletes CD11c⁺F4/80⁺alveolar macrophages in lungs and bronchoalveolar lavages.

<p>The cells from the bronchoalveolar lavage (BAL) and lungs from mice (n = 3) were harvested4 days after clodronate injection. CD3⁺ and CD11b⁺ cells were excluded to determine the effect of clodronate on depleting alveolar macrophages with CD3⁻CD11b⁻CD11c⁺F4/80⁺markers. (A) Representative profiles of flow cytometry gating of CD3⁻CD11b⁻CD11c⁺F4/80⁺phenotypic cells. (B) Percentages of CD3⁻CD11b⁻CD11c⁺F4/80⁺cells. (C) Cellularity of CD3⁻CD11b⁻CD11c⁺F4/80⁺cells. (−) Clodronate or (−)Clod: No clodronate control mice, (+) Clodronate or (+)Clod: Clodronate treated mice. BAL: Bronchoalveolar cells, Lung: Cells collected from lung tissues. *P<0.05, **P<0.001, ***P<0.0001 (Student T-test). Bars indicate standard errors.</p

Effects of clodronate treatment on DK119-mediated protection against H3N2 influenza virus infection.

<p>Groups of mice (n = 6) were two times treated with DK119 (10⁸ CFU/mouse) at 4 and 1 day earlier, then either received clodronate or PBS 4 hours prior to infection with A/Philippines/82H3N2 virus (2.5 LD₅₀) in the presence of a low dose of DK119 (10⁸ CFU per mouse). DK119+H3N2: mice pretreated with DK119 were infected with A/Philippines/82 (H3N2) virus. DK119+clodronate+H3N2: mice pretreated with DK119 were intranasally administered clodronate prior to infection with A/Philippines/82 (H3N2) virus. H3N2+Clodronate: mice treated with clodronate 4 hours prior to infection.DK119+Clodronate: mice treated with DK119 at the same doses and then clodronate administration. Bars indicate standard errors.</p

Intranasal co-administration of DK119 (10⁹CFU) and A/PR8 H1N1 virus confers protection.

<p>BALB/c mice (n = 5 per group) were infected with A/PR8 virus (2.5 LD₅₀) alone or co-administered with DK119 (10⁹ CFU per mouse) and A/PR8 H1N1 virus, and body weight changes were monitored daily for 14 days. Virus (V): mice infected with A/PR8 virus, DK119(10⁹)+V: mice infected with DK119 and A/PR8 H1N1 virus.DK119 (10⁹): mice treated with DK119 (10⁹ CFU per mouse) alone. Bars indicate standard errors.</p