Altered distribution and levels of cathepsinD and cystatins in amyotrophic lateral sclerosis transgenic mice: possible roles in motor neuron survival

In amyotrophic lateral sclerosis (ALS) there is a selective degeneration of motor neurons leading to muscle paralysis and death. The mechanism underlying cell demise in ALS is not fully understood, but involves the activation of different proteolytic enzymes, including the caspase family of cysteine proteases. We have here studied whether other proteases, such as the cathepsins, residing in lysosomes, and the cathepsin inhibitors, cystatinB and -C are changed in ALS. The expression and protein levels of the cathepsinB, -L and -D all increased in the spinal cord in ALS mice, carrying the mutant copper/zinc superoxide dismutase (SOD1) gene. At the cellular level, cathepsinB and -L were present in ventral motor neurons in controls, but in the ALS mice cathepsinB was also expressed by glial fibrillary acidic protein (GFAP) positive astrocytes. The distribution of the aspartic protease, cathepsinD also changed in ALS with a loss of the lysosomal staining in motor neurons. Inhibition of caspases by means of X-chromosome-linked inhibitor of apoptosis protein (XIAP) overexpression did not inhibit cleavage of cathepsinD in ALS mice, suggesting a caspase-independent pathway. Expression of cystatinB and -C increased slightly in the ALS spinal cords. Immunostaining showed that in ALS, cystatinC was present in motor neurons and in GFAP positive astrocytes. CystatinB that is a neuroprotective factor decreased in motor neurons in ALS but was expressed by activated microglial cells. The observed changes in the levels and distributions of cathepsinD and cystatinB and-C indicate a role of these proteins in the degeneration of motor neurons in ALS.     

Fluid pressure driven fibril reinforcement in creep and relaxation tests of articular cartilage

Biological tissues exhibit diverse mechanical behaviors because of complex material properties. As has been shown for ligaments and intervertebral discs, mathematical models often appear to well predict load responses individually by adjusting model parameters, but likely fail to describe several different load responses simultaneously using the same model parameters. In the present study, we attempted to describe and explain both creep and relaxation responses of articular cartilage using a fibril-reinforced model, which has been successfully used to account for the load response of the relaxation tests of articular cartilage. Experiments were performed on bovine articular cartilage disks (n=8) using multi-step loading protocols, involving both creep and relaxation in each protocol. The experimental results indicated that mechanical changes, such as fiber recruitment in collagen network during stretch, recovered fully upon unloading. Creep loading did not affect relaxation properties, and vice versa. Relaxation proceeded much faster than creep, because of different fluid pressure profiles. The load sharing among the proteoglycan matrix, collagen network and fluid pressurization was predicted to differ for the creep and relaxation testing. The experimentally observed strong creep and relaxation responses in unconfined compression could not be predicted if either fibril reinforcement or fluid pressurization were neglected. It was essential to consider the interplay between nonlinear fibril reinforcement and fluid pressurization for the transient response (this interplay may be best termed as fluid pressure driven fibril reinforcement). Fibril reinforcement played a relatively insignificant role in the compressive load response at equilibrium, in agreement with previous findings for cartilage stress relaxation testing.     

Screening for cardiovascular risk factors and self-rated health in a community setting: a cross-sectional study in Finland

Background

Self-rated health is an independent predictor of mortality. However, general health checks in populations unselected for disease or risk factors have not been shown to reduce mortality or morbidity.

Aim

To describe new comorbidities and cardiovascular risk factors in apparently healthy people and to relate this to their self-rated health.

Design and setting

A targeted screening programme identified 462 middle-aged people with cardiovascular risk factors without previously diagnosed chronic disease in a Finnish community in 2005–2006.

Method

Home blood pressure monitoring, oral glucose tolerance test, estimated glomerular filtration rate, and ankle brachial index were used to detect previously undiagnosed conditions. The Short-Form Health Survey and Beck’s Depression Inventory were completed by participants before the diagnostic tests were performed.

Results

The prevalence of previously undiagnosed disease was: hypertension 113/462 (24% [95% confidence interval {CI} = 21% to 29%]), diabetes 19/462 (4% [95% CI = 2% to 6%]), renal insufficiency 23/462 (5% [95% CI = 3% to 7%]), and peripheral arterial disease 17/462 (4% [95% CI = 2% to 5%]). Of the 139 participants who regarded their health as ‘fair–poor’, 60 (43%) had a previously undetected condition affecting their vasculature.

Conclusion

Out of the screen-detected apparently healthy cardiovascular risk subjects, one in three had undiagnosed hypertension, diabetes, peripheral arterial disease, or renal insufficiency. Those individuals experiencing ill health tended to be at high risk of cardiovascular problems.     

The gene for polycomb group protein enhancer of zeste homolog 2 (EZH2) is amplified in late-stage prostate cancer

Overexpression of the polycomb group protein enhancer of zeste homologue 2 (EZH2) has been found in several malignancies, including prostate cancer, with an aggressive phenotype. Amplification of the gene has previously been demonstrated in several malignancies, but not in prostate cancer. Our goal was to evaluate the gene copy number and expression alterations of EZH2 in prostate cancer. The copy number of EZH2 in cell lines (LNCaP, DU145, PC‐3, 22Rv1), xenografts (n = 10), and clinical tumors (n = 191) was studied with fluorescence in situ hybridization. All cell lines had a gain of EZH2. Eight of the ten xenografts showed an increased copy number of the gene, including one case of high‐level amplification (≥5 copies of the gene and EZH2/centromere ratio ≥2). 34/125 (27%) of untreated prostate carcinomas showed increased copy number, but only one case of low‐level amplification (≥5 copies of the gene and EZH2/centromere ratio <2), whereas half (25/46) of the hormone‐refractory carcinomas showed increased copy number, including seven cases of low‐level amplification and three cases of high‐level amplification (P < 0.0001). Expression of EZH2 was significantly (P = 0.0009) higher in hormone‐refractory prostate cancer compared with that in benign prostatic hyperplasia or untreated cancer, according to quantitative real‐time RT‐PCR assay. Also, the expression of EZH2 protein was found to be higher in hormone‐refractory tumors than in hormone‐naïve tumors by immunohistochemistry. The EZH2 gene amplification was significantly (P < 0.05) associated with increased EZH2 protein expression. The data show that amplification of the EZH2 gene is rare in early prostate cancer, whereas a fraction of late‐stage tumors contains the gene amplification leading to the overexpression of the gene, thus indicating the importance of EZH2 in the progression of prostate cancer. © 2006 Wiley‐Liss, Inc.     

Cross‐Linked Poly(ε‐caprolactone/D,L‐lactide) Copolymers with Elastic Properties

Cross‐linked ε‐caprolactone (CL) and D ,L ‐lactide (DLLA) copolymers with elastic properties were synthesized in three steps. First, the monomers were copolymerized in ring‐opening polymerization to obtain telechelic star‐shaped oligomers with almost completely random monomer distribution. The oligomers were methacrylated with methacrylic anhydride in the second step and cured in a third. Molar CL/DLLA compositions of 30/70, 50/50, 70/30, 90/10, and 100/0 were used to obtain elastic structures with a wide range of properties. The effect of the average length of the copolymer block on the properties of the networks was evaluated with three different co‐initiator contents (0.5, 1.0, and 2.0/100) in the oligomer synthesis. The oligomers were characterized by ¹³C NMR spectroscopy, size‐exclusion chromatography (SEC), and differential‐scanning calorimetry (DSC). The formation of elastic networks was confirmed by the absence of a flow region in dynamic mechanical analysis (DMA), the increase in T_g in DSC, and the full recovery of the sample dimensions after tensile testing. In addition, gel contents were high and the samples swelled in CH₂Cl₂. The networks possessed break stresses from 0.7–9.7 MPa with elongations from 80–350%. Networks with 100 or 90% of ε‐caprolactone retained their form in vitro for 12 weeks, but an increase in lactide content made the networks more vulnerable to hydrolysis.

Water absorption of the polymers during hydrolysis.     

Importance of the superficial tissue layer for the indentation stiffness of articular cartilage

Indentation testing is a widely used technique for nondestructive mechanical analysis of articular cartilage. Although cartilage shows an inhomogeneous, layered structure with anisotropic mechanical properties, most theoretical indentation models assume material homogeneity and isotropy. In the present study, quantitative polarized light microscopy (PLM) measurements from canine cartilage were utilized to characterize thickness and structure of the superficial, collageneous tissue layer as well as to reveal its relation to experimental indentation measurements. In addition to experimental analyses, a layered, transversely isotropic finite element (FE) model was developed and the effect of superficial (tangential) tissue layer with high elastic modulus in the direction parallel to articular surface on the indentation response was studied. The experimental indentation stiffness was positively correlated with the relative thickness of the superficial cartilage layer. Also the optical retardation, which reflects the degree of parallel organization of collagen fibrils as well as collagen content, was related to indentation stiffness. FE results indicated effective stiffening of articular cartilage under indentation due to high transverse modulus of the superficial layer. The present results suggest that indentation testing is an efficient technique for the characterization of the superficial degeneration of articular cartilage.     

Ethylene–Propylene Copolymerisations: Effect of Metallocene Structure on Termination Reactions and Polymer Microstructure

Summary: Ethylene–propylene (EP) copolymerisations were performed with two sterically different metallocenes activated by methylaluminoxane (MAO) in an attempt to better understand the effect of catalyst structure on termination reactions and polymer microstructure. The metallocene precursors under investigation were rac‐dimethylsilylbis(2‐methyl‐4‐phenyl‐1‐indenyl)zirconium dichloride ( 1 ) and a more sterically hindered counterpart rac‐dimethylsilylbis(2‐isopropyl‐4‐[3,5‐dimethylphenyl]indenyl) zirconium dichloride ( 2 ). For both catalyst systems, the most common termination mechanism was chain transfer to aluminium. In addition, for polymer samples polymerised with 1 /MAO, chain growth was terminated by chain transfer to Zr metal in propylene‐rich polymerisations and by chain transfer to ethylene monomer in ethylene‐rich polymerisations. The steric hindrance of 2 was able to suppress the chain transfer to the ethylene monomer, and chain transfer to Zr metal was also found in the ethylene‐rich polymerisations. The greater steric hindrance of 2 also affected the EP copolymer microstructure: regioregularity in the propylene‐rich copolymers was greater and isotacticity less with 2 /MAO than with 1 /MAO.

The catalyst precursors used: rac‐dimethylsilylbis(2‐methyl‐4‐phenyl‐1‐indenyl)zirconium dichloride ( 1 ) and rac‐dimethylsilylbis(2‐isopropyl‐4‐[3,5‐dimethylphenyl]indenyl) zirconium dichloride ( 2 ).     

Behavior of 3-year-old children in a prospective randomized trial of reduced saturated fat and cholesterol diet since infancy: The strip baby project

Interventions aimed at decreased exposure of children to known atherosclerosis risk factors may have untoward behavioral side effects. We examined how children’s behavior or parent’s perception of the behavior of the children at 3 years of age was influenced by the intervention in a prospective randomized trial that began in infancy and effectively decreased scrum cholesterol concentration. This Special Turku coronary Risk factor Intervention Project for babies (STRIP) began when the infant was 7 months old. Half of 1.062 children received individualized dietary counseling at 1-to 3-month intervals during the first 2 years of age and then half-yearly; the other half had an unrestricted diet. At 3 years of age a standardized questionnaire of the child’s behavior was sent to 791 families (76% returned the questionnaire). At the onset of the trial the sociodemographic data of the families and scrum lipid values of the intervention and control children were similar. Later, mean serum cholesterol values of the intervention children remained constantly at a level 6% to 10% below the values of the control children. At 3 years of age the parental perceptions of the child’s behavior suggested minimal differences between the intervention and control children. The intervention children were slightly less jealous and more active and creative, but showed slightly more negative signs of behavior (bed-wetting, problems in falling asleep, fears) than the controls. We conclude that long-term, individualized dietary and lifestyle intervention that begins in infancy slightly influences children’s behavior or parent’s recognition of the behavior of the children at the age of 3 years. This work was supported in part by grants from the Varsinais-Suomi Regional Fund of the Finnish Cultural Foundation, the Mannerheim League for Child Welfare, the Academy of Finland, and the Alli Paasikivi Foundation.     

Physical loading and performance as predictors of back pain in healthy adults A 5-year prospective study

We investigated muscle strength, aerobic power, and occupational and leisure-time physical loading as predictors of back pain in a 5-year follow-up study. A cohort of 456 adults aged 25, 35, 45 and 55 years, free of back pain, participated in measurements of anthropometric characteristics, aerobic power and muscle strength characteristics at baseline. The subjects' levels and types of physical activity and occupational physical loading were also determined. At 5 years after the baseline examinations 356 of these subjects (78.1 %) were reached by mail, and 262 of them (73.6%) properly completed and returned a questionnaire including a detailed back pain history for the 5 years following the baseline measurements. Of this number 56 subjects (21 %) who reported back pain ( > 30 on a scale from 0 to 100) and functional impairment during the 5-year follow-up composed the marked back pain group. Other subjects (n = 71, 27%) noting lesser symptoms were included in the mild back pain group; 135 subjects (52%) reported having had no back pain. The subjects with marked back pain were on average taller than the subjects without back pain, while no such difference was found in body mass. Heavy occupational musculoskeletal loading (P = 0.005) and high general occupational physical demands (P = 0.036) predicted future back pain. Leisuretime physical activity, aerobic power or muscle strength characteristics were not predictive of future back pain.