A novel scheme of dystrophin disruption for the progression of advanced heart failure

The precise mechanism of the progression of advanced heart failure is unknown. We assessed a new scheme in two heart failure models: (I) congenital dilated cardiomyopathy (DCM) in TO-2 strain hamsters lacking delta-sarcoglycan (SG) gene and (II) administration of a high-dose of isoproterenol, as an acute heart failure in normal rats. In TO-2 hamsters, we followed the time course of the histological, physiological and metabolic the progressions of heart failure to the end stage. Dystrophin localization detected by immunostaining age-dependently to the myoplasm and the in situ sarcolemma fragility evaluated by Evans blue entry was increased in the same cardiomyocytes. Western blotting revealed a limited cleavage of the dystrophin protein at the rod domain, strongly suggesting a contribution of endogenous protease(s). We found a remarkable up-regulation of the amount of calpain-1 and -2, and no change of their counterpart, calpastatin. After supplementing TO-2 hearts with the normal delta-SG gene in vivo, these pathological alterations and the animals' survival improved. Furthermore, dystrophin but not delta-SG was disrupted by a high dose of isoproterenol, translocated from the sarcolemma to the myoplasm and fragmented. These results of heart failure, irrespective of the hereditary or acquired origin, indicate a vicious cycle formed by the increased sarcolemma permeability, preferential activation of calpain over calpastatin, and translocation and cleavage of dystrophin would commonly lead to advanced heart failure.     

Four-dimensional model of the lower extremity after total hip arthroplasty

We have developed a four-dimensional (4D) model of the lower extremities after total hip arthroplasty in patients. The model can aid in preventing complications such as dislocation and wearing of the sliding surface. The skeletal structure and implant alignment were obtained from CT data. We applied registration method using CAD data to estimate accurate implant alignment from scattered CT data. The reconstructed three-dimensional (3D) skeletal model was combined with motion capture data that were acquired by an optical tracking system. We displayed the patient's skeletal movement and analyzed several parameters that relate to complications. The patient's skeletal model was superimposed onto video footage that was taken by a synchronized and calibrated digital video camera. For validation of the measurement error in this system, we used open MRI to evaluate the relative movement between skin markers and bones. This system visually represents not only the 3D anatomical structure, but also 4D dynamic functions that include the time sequential transitions of components and their positions. The open MRI results indicated that the average error in hip angle was within 5° for each static posture. This system enables clinicians to analyze patient's motions on the basis of individual differences. We found that our system was an effective tool in providing precise guidance of daily postoperative motions that was individualized for each patient. This system will be applicable for surgical planning, assessment of postoperative activities, and the development of new surgical techniques, materials, and prosthetic designs.     

Involvement of claudin-7 in HIV infection of CD4(-) cells

Background

The antibacterial activity of host defense peptides (HDP) is largely mediated by permeabilization of bacterial membranes. The lipid membrane of enveloped viruses might also be a target of antimicrobial peptides. Therefore, we screened a panel of naturally occurring HDPs representing different classes for inhibition of early, Env-independent steps in the HIV replication cycle. A lentiviral vector-based screening assay was used to determine the inhibitory effect of HDPs on early steps in the replication cycle and on cell metabolism.

Results

Human LL37 and porcine Protegrin-1 specifically reduced lentiviral vector infectivity, whereas the reduction of luciferase activities observed at high concentrations of the other HDPs is primarily due to modulation of cellular activity and/or cytotoxicity rather than antiviral activity. A retroviral vector was inhibited by LL37 and Protegrin-1 to similar extent, while no specific inhibition of adenoviral vector mediated gene transfer was observed. Specific inhibitory effects of Protegrin-1 were confirmed for wild type HIV-1.

Conclusion

Although Protegrin-1 apparently inhibits an early step in the HIV-replication cycle, cytotoxic effects might limit its use as an antiviral agent unless the specificity for the virus can be improved.     

Biochemical examination on the increased superoxide dismutase-like plasma substance observed in a state of acute gastric mucosal lesions

When experimental acute gastric mucosal lesions were produced in guinea pig by water-immersion and restraint stress, superoxide dismutase (SOD)-like substance in the plasma increased. On analysis by gel filtration, it was shown that the molecular weight of the increased SOD-like plasma substance was about 130,000, and even after treatment with trypsin, 84% of this substance remained. Since the molecular weight of intracellular SOD is about 40,000, it seems that this substance is similar to extracellular SOD, located on the endothelial cell-surface, as previously reported by Marklund et al. Our results suggest that in the presence of acute gastric mucosal lesions, SOD-like plasma substance is not identical to intracellular SOD, which derived from cell destruction by stress or free radical-induced microvascular damage or by hemolysis. Furthermore, this substance may itself work as a scavenger of free radicals generated under conditions, such as these described in the present experiment.     

Effect of self-association on the structural organization of partially folded proteins: inactivated actin

The propensity to associate or aggregate is one of the characteristic properties of many nonnative proteins. The aggregation of proteins is responsible for a number of human diseases and is a significant problem in biotechnology. Despite this, little is currently known about the effect of self-association on the structural properties and conformational stability of partially folded protein molecules. G-actin is shown to form equilibrium unfolding intermediate in the vicinity of 1.5 M guanidinium chloride (GdmCl). Refolding from the GdmCl unfolded state is terminated at the stage of formation of the same intermediate state. An analogous form, known as inactivated actin, can be obtained by heat treatment, or at moderate urea concentration, or by the release of Ca(2+). In all cases actin forms specific associates comprising partially folded protein molecules. The structural properties and conformational stability of inactivated actin were studied over a wide range of protein concentrations, and it was established that the process of self-association is rather specific. We have also shown that inactivated actin, being denatured, is characterized by a relatively rigid microenvironment of aromatic residues and exhibits a considerable limitation in the internal mobility of tryptophans. This means that specific self-association can play an important structure-forming role for the partially folded protein molecules.     

Synthesis and evaluation of N-acylsulfonamide and N-acylsulfonylurea prodrugs of a prostacyclin receptor agonist

N-Acylsulfonamide and N-acylsulfonylurea derivatives of the carboxylic acid prostacyclin receptor agonist 1 were synthesized and their potential as prodrug forms of the carboxylic acid was evaluated in vitro and in vivo. These compounds were converted to the active compound 1 by hepatic microsomes from rats, dogs, monkeys, and humans, and some of the compounds were shown to yield sustained plasma concentrations of 1 when they were orally administered to monkeys. These types of analogues, including NS-304 (2a), are potentially useful prodrugs of 1.