Screening of Multiple Myeloma by Polyclonal Rabbit Anti-Human Plasmacytoma Cell Immunoglobulin

Antibody-based immunotherapy has been effectively used for tumor treatment. However, to date, only a few tumor-associated antigens (TAAs) or therapeutic targets have been identified. Identification of more immunogenic antigens is essential for improvements in multiple myeloma (MM) diagnosis and therapy. In this study, we synthesized a polyclonal antibody (PAb) by immunizing rabbits with whole human plasmacytoma ARH-77 cells and identified MM-associated antigens, including enlonase, adipophilin, and HSP90s, among others, via proteomic technologies. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay showed that 200 µg/mL PAb inhibits the proliferation of ARH-77 cells by over 50% within 48 h. Flow cytometric assay indicated that PAb treatment significantly increases the number of apoptotic cells compared with other treatments (52.1% vs. NS, 7.3% or control rabbit IgG, 9.9%). In vivo, PAb delayed tumor growth and prolonged the lifespan of mice. Terminal deoxynucleotidyl transferase dUTP nick end labeling assay showed that PAb also induces statistically significant changes in apoptosis compared with other treatments (P<0.05). We therefore conclude that PAb could be used for the effective screening and identification of TAA. PAb may have certain anti-tumor functions in vitro and in vivo. As such, its combination with proteomic technologies could be a promising approach for sieving TAA for the diagnosis and therapy of MM.     

Cytokine Profiling in Immigrants with Clinical Malaria after Extended Periods of Interrupted Exposure to Plasmodium falciparum

Immunity to malaria is believed to wane with time in the absence of exposure to Plasmodium falciparum infection, but immunoepidemiological data on longevity of immunity remain controversial. We quantified serum cytokines and chemokines by suspension array technology as potential biomarkers for durability of immunity in immigrants with clinical malaria after years without parasite exposure. These were compared to serum/plasma profiles in naïve adults (travelers) and semi-immune adults under continuous exposure, with malaria, along with immigrant and traveler patients without malaria. Immigrants had higher levels of IL-2, IL-5 and IL-8 compared to semi-immune adults with malaria (P≤0.0200). Time since immigration correlated with increased IL-2 (rho=0.2738P=0.0495) and IFN-γ (rho=0.3044P=0.0282). However, immigrants did not show as high IFN-γ concentrations as travelers during a first malaria episode (P<0.0001). Immigrants and travelers with malaria had higher levels of IFN-γ, IL-6, and IL-10 (P<0.0100) than patients with other diseases, and IL-8 and IL-1β were elevated in immigrants with malaria (P<0.0500). Therefore, malaria patients had a characteristic strong pro-inflammatory/Th1 signature. Upon loss of exposure, control of pro-inflammatory responses and tolerance to P. falciparum appeared to be reduced. Understanding the mechanisms to maintain non-pathogenic effector responses is important to develop new malaria control strategies.     

Effect of Cadmium Ion on alpha-Glucosidase: An Inhibition Kinetics and Molecular Dynamics Simulation Integration Study

α-Glucosidase is a critical metabolic enzyme that produces glucose molecules by catalyzing carbohydrates. The aim of this study is to elucidate biological toxicity of Cd²⁺ based on α-glucosidase activity and conformational changes. We studied Cd²⁺-mediated inactivation as well as conformational modulation of α-glucosidase by using kinetics coupled with simulation of molecular dynamics. The enzyme was significantly inactivated by Cd²⁺ in a reversibly binding behavior, and Cd²⁺ binding induced a non-competitive type of inhibition reaction (the K _i was calculated as 0.3863 ± 0.033 mM). Cd²⁺ also modulated regional denaturation of the active site pocket as well as overall partial tertiary structural change. In computational simulations using molecular dynamics, simulated introduction of Cd²⁺ induced in a depletion of secondary structure by docking Cd²⁺ near the saccharides degradation at the active site, suggesting that Cd²⁺ modulating enzyme denaturation. The present study elucidated that the binding of Cd²⁺ triggers conformational changes of α-glucosidase as well as inactivates catalytic function, and thus suggests an explanation of the deleterious effects of Cd²⁺ on α-glucosidase.     

Ultrasonic Elastography Research Based on a Multicenter Study: Adding Strain Ratio after 5-Point Scoring Evaluation or Not

Background

This study aimed to confirm whether strain ratio should be added after evaluation of lesions with 5-point elasticity scoring for differentiating benign and malignant breast lesions on ultrasonographic elastography(UE).

Materials and Methods

From June 2010 to March 2012, 1080 consecutive female patients with breast lesions were recruited into a multicenter retrospective study, which involved 8 centers across China. Each institutional ethic review board approved the study, and all the patients gave written informed consent. All the patients underwent the UE procedure and the strain ratios were calculated and the final diagnosis was made by histological findings. The sensitivity, specificity, accuracy, PPV and NPV were calculated for each of the two evaluation systems and the areas under the ROC curve were compared.

Results

The strain ratios of benign lesions (mean, 2.6±2.0) and malignant lesions (mean,7.9±5.8) were significantly different (p <0.01). When the cutoff point was 3.01, strain ratio method had 79.8% sensitivity, 82.8% specificity, and 81.3% accuracy, while the 5-point scoring method had 93.1% sensitivity, 73.0% specificity, and 76.8% accuracy. The areas under the ROC curve with the strain ratio method and 5-point scoring method were 0.863 and 0.865, respectively(p>0.05). The strain ratio method shows better a diagnosis performance of the lesions with elasticity score 3 and 4.

Conclusions

Although the two UE methods have similar diagnostic performance, separate calculation of the strain ratios seems compulsory, especially for the large solid breast lesions and the lesions with elasticity score 3 and 4.     

QM/MM investigation of the catalytic mechanism of angiotensin-converting enzyme

Angiotensin-converting enzyme (ACE) converts angiotensin I to angiotensin II and degrades bradykinin and other vasoactive peptides. ACE inhibitors are used to treat diseases such as hypertension and heart failure. It is thus highly desirable to understand the catalytic mechanism of ACE, as this should facilitate the design of more powerful and selective ACE inhibitors. ACE exhibits two different active domains, the C-domain and the N-domain. In this work, we systematically investigated the inhibitor- and substrate-binding patterns in the N-domain of human ACE using a combined quantum mechanical and molecular mechanical approach. The hydrolysis of hippuryl–histidyl–leucine (HHL) as catalyzed by the N-domain of human somatic ACE was explored, and the effects of chloride ion on the overall reaction were also investigated. Two models, one with and one without a chloride ion at the first binding position, were then designed to examine the chloride dependence of inhibitor–substrate binding and the catalytic mechanism. Our calculations indicate that the hydrolysis reaction follows a stepwise general base/general acid catalysis path. The estimated mean free energy barrier height in the two models is about 15.6 kcal/mol, which agrees very well with the experimentally estimated value of 15.8 kcal/mol. Our simulations thus suggest that the N-domain is in a mixed form during ACE-catalyzed hydrolysis, with the single-chloride-ion and the double-chloride-ion forms existing simultaneously.

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Graphical Abstract Superposition of ACE C- and N- domains

     

Skin aging caused by intrinsic or extrinsic processes characterized with functional proteomics

The skin provides protection against environmental stress. However, intrinsic and extrinsic aging causes significant alteration to skin structure and components, which subsequently impairs molecular characteristics and biochemical processes. Here, we have conducted an immunohistological investigation and established the proteome profiles on nude mice skin to verify the specific responses during aging caused by different factors. Our results showed that UVB‐elicited aging results in upregulation of proliferating cell nuclear antigen and strong oxidative damage in DNA, whereas chronological aging abolished epidermal cell growth and increased the expression of caspase‐14, as well as protein carbonylation. Network analysis indicated that the programmed skin aging activated the ubiquitin system and triggered obvious downregulation of 14‐3‐3 sigma, which might accelerate the loss of cell growth capacity. On the other hand, UVB stimulation enhanced inflammation and the risk of skin carcinogenesis. Collectively, functional proteomics could provide large‐scale investigation of the potent proteins and molecules that play important roles in skin subjected to both intrinsic and extrinsic aging.