Idiopathic calcium nephrolithiasis. 1. Differences in urine crystalloids,urine saturation with brushite and urine inhibitors of calcification between persons with and persons without recurrent kidney stone formation.

The propensity of urine to promote calcium stone formation was compared in 64 patients with recurrent idiopathic calcium nephrolithiasis and 30 healthy individuals without such a history. The rates of excretion of urine crystalloids, the urine saturation with brushite (CaHPO4-2H2O), the ability of the urine to calcify collagen in vitro, and the concentration of urine inhibitors of collagen calcification were measured. The patients had a reduced urine citrate excretion rate in addition to an increased urine calcium excretion rate, while the rates for urine magnesium, phosphate, uric acid and oxalate were not significantly different in the two groups of subjects. The urine concentration of magnesium, phosphate and uric acid was decreased in the patients because of the higher urine volume. The urine creatinine excretion rate correlated with the rates of excretion of urine calcium, magnesium, phosphate, uric acid and oxalate in both groups, which suggested that increased lean body mass, possibly associated with greater food intake, may be an important determinant of crystalloid excretion. The urine of the patients was significantly more saturated with brushite than the urine of the control subjects and resulted in greater collagen calcification when incubated in vitro. The urine concentration of inhibitors of collagen calcification, however, was not significantly different in the two groups. Thus, the urine of patients with recurrent idiopathic calcium nephrolithiasis is more highly saturated with brushite, largely as a result of an increased urine calcium excretion rate, and contains a lower concentration of magnesium and citrate, substances that tend to prevent the precipitation and growth of crystals in urine.     

Sialylation of urinary prothrombin fragment 1 is implicated as a contributory factor in the risk of calcium oxalate kidney stone formation

Urinary glycoproteins are important inhibitors of calcium oxalate crystallization and adhesion of crystals to renal cells, both of which are key mechanisms in kidney stone formation. This has been attributed to glycosylation of the proteins. In South Africa, the black population rarely form stones (incidence < 1%) compared with the white population (incidence 12-15%). A previous study involving urinary prothrombin fragment 1 from both populations demonstrated superior inhibitory activity associated with the protein from the black group. In the present study, we compared N-linked and O-linked oligosaccharides released from urinary prothrombin fragment 1 isolated from the urine of healthy and stone-forming subjects in both populations to elucidate the relationship between glycosylation and calcium oxalate stone pathogenesis. The O-glycans of both control groups and the N-glycans of the black control samples were significantly more sialylated than those of the white stone-formers. This demonstrates a possible association between low-percentage sialylation and kidney stone disease and provides a potential diagnostic method for a predisposition to kidney stones that could lead to the implementation of a preventative regimen. These results indicate that sialylated glycoforms of urinary prothrombin fragment 1 afford protection against calcium oxalate stone formation, possibly by coating the surface of calcium oxalate crystals. This provides a rationale for the established roles of urinary prothrombin fragment 1, namely reducing the potential for crystal aggregation and inhibiting crystal-cell adhesion by masking the interaction of the calcium ions on the crystal surface with the renal cell surface along the nephron.     

Differences between scar and dermal cultured fibroblasts derived from a patient with recurrent abdominal incision wound herniation.

Fibroblasts were derived from dermis and scar of a 47-year-old white man with a recurrent incisional hernia as a result of fractured ribs. The scar was thin and stretched, suggesting a defect in the maturation of granulation tissue. After surgical repair, biopsy specimens of discarded scar and skin were used to generate fibroblast cell lines. Fibroblasts maintained in medium containing 10% fetal bovine serum and antibiotic were studied between their third and eighth passage. By phase contrast microscopy, no structural differences were obvious, but it was noted that to pass scar fibroblasts, a more aggressive trypsin regimen was required. Immunohistologic and Western blot analysis of patient scar fibroblasts showed (1) more a smooth muscle actin within stress fibers, (2) increased expression of the vitronectin integrin receptor alpha(v) (CD 51), and (3) reduced expression of the collagen integrin receptor alpha2 (CD 49b). The expression of vinculin from focal adhesions or a tubulin from microtubules was the same among cell lines. Contractions of scar and dermal fibroblast-populated collagen lattice were compared. At 24 hours, contractions were 69 percent with newborn fibroblasts (normal); 68 percent for patient dermal fibroblasts; and only 48 percent for patient scar fibroblasts. The retarded contraction of scar fibroblast-populated collagen lattice was significant (p > or = 0.002). Myosin ATPase activity, critical for lattice contraction, and cell migration were equivalent among all cell lines. A plausible mechanism for the retardation of scar lattice contraction is disruption of fibroblasts and collagen interactions, for which the attachment of cells to collagen is altered. It is proposed that either the decrease in the expression of collagen integrin receptor alpha2 (CD 49b), an increase in the expression of the vitronectin receptor alpha(v) (CD 51), or a combination of both is responsible for disruption of collagen fibroblast interactions.     

The plasma membrane calcium pump displays memory of past calcium spikes. Differences between isoforms 2b and 4b

To understand how the plasma membrane Ca(2+) pump (PMCA) behaves under changing Ca(2+) concentrations, it is necessary to obtain information about the Ca(2+) dependence of the rate constants for calmodulin activation (k(act)) and for inactivation by calmodulin removal (k(inact)). Here we studied these constants for isoforms 2b and 4b. We measured the ATPase activity of these isoforms expressed in Sf9 cells. For both PMCA4b and 2b, k(act) increased with Ca(2+) along a sigmoidal curve. At all Ca(2+) concentrations, 2b showed a faster reaction with calmodulin than 4b but a slower off rate. On the basis of the measured rate constants, we simulated mathematically the behavior of these pumps upon repetitive changes in Ca(2+) concentration and also tested these simulations experimentally; PMCA was activated by 500 nm Ca(2+) and then exposed to 50 nm Ca(2+) for 10 to 150 s, and then Ca(2+) was increased again to 500 nm. During the second exposure to 500 nm Ca(2+), the activity reached steady state faster than during the first exposure at 500 nm Ca(2+). This memory effect is longer for PMCA2b than for 4b. In a separate experiment, a calmodulin-binding peptide from myosin light chain kinase, which has no direct interaction with the pump, was added during the second exposure to 500 nm Ca(2+). The peptide inhibited the activity of PMCA2b when the exposure to 50 nm Ca(2+) was 150 s but had little or no effect when this exposure was only 15 s. This suggests that the memory effect is due to calmodulin remaining bound to the enzyme during the period at low Ca(2+). The memory effect observed in PMCA2b and 4b will allow cells expressing either of them to remove Ca(2+) more quickly in subsequent spikes after an initial activating spike.     

Concentration of a potent calcium oxalate monohydrate crystal growth inhibitor in the urine of normal persons and kidney stone patients by ELISA-based assay system employing monoclonal antibodies

Standardized calcium oxalate monohydrate (COM) crystal growth assay system was employed to study the ability of various test samples to influence growth rates of COM crystals. The inhibitory activity (IA) of various samples was expressed in terms of inhibitory units. Urine samples obtained from normal persons and kidney stone patients were found to have IA of 3.18 +/- 0.62 and 1.02 +/- 0.08, respectively. A potent inhibitor having molecular weight between 14.2 and 16.2 kDa was found to be primarily responsible for the differences observed in the urinary IAs between normal persons and kidney stone patients. The potent inhibitor was found to be tightly associated with a chromophore resembling Urobilirubin. An ELISA based assay system, using monoclonal antibodies against the above most potent inhibitor confirmed the difference observed in the urinary IA between the normal persons and kidney stone patients. This assay system has the potential to be routinely used to screen human beings for potential stone formers.     

Differences in Brain Morphological Findings between Narcolepsy with and without Cataplexy

Objective

Maps of fractional anisotropy (FA) and apparent diffusion coefficient (ADC) obtained by diffusion tensor imaging (DTI) can detect microscopic axonal changes by estimating the diffusivity of water molecules using magnetic resonance imaging (MRI). We applied an MRI voxel-based statistical approach to FA and ADC maps to evaluate microstructural abnormalities in the brain in narcolepsy and to investigate differences between patients having narcolepsy with and without cataplexy.

Methods

Twelve patients with drug-naive narcolepsy with cataplexy (NA/CA), 12 with drug-naive narcolepsy without cataplexy (NA w/o CA) and 12 age-matched healthy normal controls (NC) were enrolled. FA and ADC maps for these 3 groups were statistically compared by using voxel-based one-way ANOVA. In addition, we investigated the correlation between FA and ADC values and clinical variables in the patient groups.

Results

Compared to the NC group, the NA/CA group showed higher ADC values in the left inferior frontal gyrus and left amygdala, and a lower ADC value in the left postcentral gyrus. The ADC value in the right inferior frontal gyrus and FA value in the right precuneus were higher for NA/CA group than for the NA w/o CA group. However, no significant differences were observed in FA and ADC values between the NA w/o CA and NC groups in any of the areas investigated. In addition, no correlation was found between the clinical variables and ADC and FA values of any brain areas in these patient groups.

Conclusions

Several microstructural changes were noted in the inferior frontal gyrus and amygdala in the NA/CA but not in the NA w/o CA group. These findings suggest that these 2 narcolepsy conditions have different pathological mechanisms: narcolepsy without cataplexy form appears to be a potentially broader condition without any significant brain imaging differences from normal controls.     

Socio-economic factors and rheumatic fever occurrence. Differences between patients with and without frequent sore throat

One hundred and fourty-eight rheumatic fever patients and 444 controls matched by age, sex and place of residence, were interviewed about socio-economic and some other variables. Socio-economic factors recognized as risk factors for rheumatic fever (flat dampness, more than 2 persons per room, sleeping in bed with other person, low education of mother and undernourishment) were of lesser importance for persons with frequent sore throat in comparison to persons without frequent sore throat. According to the results obtained it seems that there is positive connection between host's propensity to clinical manifestation of throat infection and manifestation of rheumatic fever. The lesser susceptibility the more additional factors are needed for Rheumatic Fever to occur. The relative importance of socio-economic factors in rheumatic fever occurrence depends on host's susceptibility to infection.     

Circulating levels of matrix proteases and their inhibitors in pregnant women with and without a history of recurrent pregnancy loss

Background  

We have recently shown that serum relaxin-2 levels are attenuated in women with a history of recurrent pregnancy loss (RPL). We sought to determine whether a history of RPL is also associated with changes in serum matrix metalloproteases (MMPs) and tissue inhibitors of matrix metalloproteases (TIMP) -1 and -2.     

Exploring Differences in the Content of Job Interviews between Youth with and without a Physical Disability

Objective

Although people with disabilities have great potential to provide advantages to work environments, many encounter barriers in finding employment, especially youth who are looking for their first job. A job interview is an essential component of obtaining employment. The objective of this study is to explore the content of the answers given in job interviews among youth with disabilities compared to typically developing youth.

Methods

A purposive sample of 31 youth (16 with typical development and 15 with disability) completed a mock job interview as part of an employment readiness study. The interview questions focused on skills and experiences, areas for improvement, and actions taken during problem-based scenarios. Transcribed interviews were analyzed using a content analysis of themes that emerged from the interviews.

Results

We found several similarities and differences between youth with disabilities and typically developing youth. Similarities included giving examples from school, emphasizing their “soft skills” (i.e., people and communication skills) and giving examples of relevant experience for the position. Both groups of youth gave similar examples for something they were proud of but fewer youth with disabilities provided examples. Differences in the content of job interview answers between the two groups included youth with disabilities: (1) disclosing their condition; (2) giving fewer examples related to customer service and teamwork skills; (3) experiencing greater challenges in providing feedback to team members and responding to scenario-based problem solving questions; and (4) drawing on examples from past work, volunteer and extra curricular activities.

Conclusions

Clinicians and educators should help youth to understand what their marketable skills are and how to highlight them in an interview. Employers need to understand that the experiences of youth with disabilities may be different than typically developing youth. Our findings also help to inform employment readiness programs by highlighting the areas where youth with disabilities may need extra help as compared to typically developing youth.     

A comparison of throwing kinematics between youth baseball players with and without a history of medial elbow pain

Risk factors in throwing factors associated to little league elbow have not been adequately explored. Whether these factors also affect the players' performance is also important to elucidate while modifying throwing pattern to reduce injury. The purpose of this study was to compare the differences in throwing kinematics between youth baseball players with or without a history of medial elbow pain (MEP) and to determine the relationship between their throwing kinematics and ball speed. Fifteen players with previous MEP were matched with 15 healthy players by age, height and weight. Throwing kinematics was recorded by an electromagnetic motion analysis system. A foot switch was used for determining foot off and foot contact. Ball speed was recorded with a sports radar gun. The group with a history of MEP demonstrated less elbow flexion angle at maximum shoulder external rotation and had more lateral trunk tilt at ball release compared to the healthy group. The group with a history of MEP also had faster maximum upper torso rotation velocities, maximum pelvis rotation velocities and ball speeds. Maximum shoulder external rotation angle (r = 0.458, P = 0.011), elbow flexion angle at maximum shoulder external rotation (r = -0.637, P = 0.0003), and maximum upper torso rotation velocity (r = 0.562, P = 0.002) had significant correlation with ball speed. Findings of this study can be treated as elbow injury-related factors that clinicians and coaches can attend to when taking care of youth     

BRCA1 and BRCA2 germline mutations in Malaysian women with early-onset breast cancer without a family history

Background

In Asia, breast cancer is characterised by an early age of onset: In Malaysia, approximately 50% of cases occur in women under the age of 50 years. A proportion of these cases may be attributable, at least in part, to genetic components, but to date, the contribution of genetic components to breast cancer in many of Malaysia''s ethnic groups has not been well-characterised.

Methodology

Given that hereditary breast carcinoma is primarily due to germline mutations in one of two breast cancer susceptibility genes, BRCA1 and BRCA2, we have characterised the spectrum of BRCA mutations in a cohort of 37 individuals with early-onset disease (≤40 years) and no reported family history. Mutational analysis of BRCA1 and BRCA2 was conducted by full sequencing of all exons and intron-exon junctions.

Conclusions

Here, we report a total of 14 BRCA1 and 17 BRCA2 sequence alterations, of which eight are novel (3 BRCA1 and 5 BRCA2). One deleterious BRCA1 mutation and 2 deleterious BRCA2 mutations, all of which are novel mutations, were identified in 3 of 37 individuals. This represents a prevalence of 2.7% and 5.4% respectively, which is consistent with other studies in other Asian ethnic groups (4–9%).     

Differences between the prion protein and its homolog Doppel: a partially structured state with implications for scrapie formation

The key event in the pathogenesis of prion diseases is a conformational change in the prion protein (PrP). Models for conversion of PrP(C) into PrP(Sc) typically implicate an, as yet, unidentified intermediate. In an attempt to identify such an intermediate, we used native-state hydrogen exchange monitored with NMR. Although we were unable to detect an intermediate directly, we observed substantial protection above that expected based upon measurements of the global stability of PrP (>2 kcal mol(-1) super protection). This super protection implicates either structure in the denatured state or the presence of an intermediate. Similar experiments with Doppel, a homolog of PrP that does not form infectious prions, failed to demonstrate such super protection. This suggests that the partially structured state of PrP encompassing portions of the B and C helices, may be a significant factor in the ability of PrP to convert from PrP(C) to PrP(Sc).     

High-affinity formation of a 2:1 complex between gramicidin S and calmodulin.

Two molecules of gramicidin S, a very rigid cyclic decapeptide rich in beta-sheet structure, can bind in a Ca2+-dependent way to a calmodulin molecule in the presence as well as in the absence of 4 M-urea. The flow-microcalorimetric titration of 25 microM-calmodulin with gramicidin S at 25 degrees C is endothermic for 21.3 kJ.mol-1; the enthalpy change is strictly linear up to a ratio of 2, indicating that the affinity constant for binding of the second gramicidin S is at least 10(7) M-1. In 4 M-urea the peptide quantitatively displaces seminalplasmin from calmodulin, as monitored by tryptophan fluorescence. An iterative data treatment of these competition experiments revealed strong positive co-operativity with K1 less than 5 X 10(5) M-1 and K1.K2 = 2.8 X 10(12) M-2. A competition assay with the use of immobilized melittin enabled us to monitor separately the binding of the second gramicidin S molecule: the K2 value is 1.9 X 10(7) M-1. By complementarity, the K1 value is 1.5 X 10(5) M-1. In the absence of urea the seminalplasmin displacement is incomplete: the data analysis shows optimal fitting with K1 less than 2 X 10(4) M-1 and K1.K2 = 3.2 X 10(11) M-2 and reveals that the mixed complex (calmodulin-seminalplasmin-gramicidin S) is quite stable and is even not fully displaced from calmodulin at high concentrations of gramicidin S. The activation of bovine brain phosphodiesterase by calmodulin is not impaired up to 0.2 microM-gramicidin S. According to our model the ternary complex enzyme-calmodulin-gramicidin is relatively important and displays the same activity as the binary complex enzyme-calmodulin. Gramicidin S also displaces melittin from calmodulin synergistically, as monitored by c.d. Our studies with gramicidin S reveal the importance of multipoint attachments in interactions involving calmodulin and confirm the heterotropic co-operativity in the binding of calmodulin antagonists first demonstrated by Johnson [(1983) Biochem. Biophys. Res. Commun. 112, 787-793].     

Microbial communities in sugarcane field soils with and without a sugarcane cropping history

Microbial communities in rhizosphere soil from sugarcane (Saccharum inter-specific hybrids) and bulk soil were compared at paired field sites with and without a sugarcane cropping history to determine whether monoculture affects soil microbial community composition. Differences were evaluated for culturable microorganisms and functional diversity indicated by community level physiological profiles (CLPP). Qualitative differences in rhizosphere bacterial communities were detected between sites with no sugarcane cropping history (N_site) and sites with a long-term sugarcane cropping history (L_site). More fluorescent pseudomonads were detected in N_site than L_site rhizosphere soil at two of three sites, and Actinobacteria were more numerous in N_site than L_site rhizosphere soil at one site. Fusarial fungi were more numerous in N_site than L_site rhizosphere soils. Bacteria were more numerous in rhizosphere soil compared to bulk soil. Total bacterial, pseudomonad, and Actinobacteria population densities were greater in bulk soil from an N_site compared to an L_site. CLPP distinguished bulk from rhizosphere soil at one of two sites and N_site and L_site rhizosphere soils at two of four sites. Site affected CLPP similarity more than cropping history. The results demonstrated that sugarcane monoculture can affect the composition of the microbial community in field soil. The findings have possible implications for reduced yields associated with sugarcane monoculture.     

Purification of a high-molecular-mass form of phospholipase A2 from rat kidney activated at physiological calcium concentrations.

Rat kidney contains a soluble phospholipase A2 (PLA2), which is chromatographically identical with a previously identified hormonally regulated form of the enzyme in rat renal mesangial cells. This kidney enzyme has been purified by sequential column fractionation. The purified enzyme is a 110 kDa polypeptide which can hydrolyse arachidonoyl phosphatidylcholine and arachidonoyl phosphatidylethanolamine, but has low activity towards arachidonoyl phosphatidylinositol. The enzyme is considerably larger than most previously isolated forms of secretory or intracellular PLA2, and is stimulated by physiological concentrations of Ca2+, with half-maximal activation occurring at 500 nM-Ca2+. The hormonal regulation and Ca2(+)-dependency of this enzyme strongly suggest that it plays a role in hormonally regulated arachidonic acid release and prostaglandin production in the kidney.     

The rat kidney acylase I, characterization and molecular cloning. Differences with other acylases I.

The soluble acylase I from rat kidney was purified to homogeneity using a five-step procedure. As the resulting protein was found to have a relative molecular mass of 125 kDa based on size-exclusion chromatography and 44 kDa based on SDS/PAGE, the native protein was taken to consist of three subunits. The amino-acid sequence of a peptide resulting from limited proteolysis of the polypeptide chain with proteinase K, which was determined by microsequencing (RHEFHALRAGFALDEGLA), was found to be very similar to the corresponding sequence of porcine kidney acylase I. However, as N-furyl-acryloyl-L-methionine, a synthetic substrate for porcine acylases, was not hydrolyzed by the rat enzyme, it was suggested that the polypeptide chain might differ in other respects from those of the other acylases I. A full length cDNA coding for the rat kidney acylase I was therefore isolated and found to contain a 1224-bp open reading frame encoding a protein consisting of 408 amino-acid residues, which corresponded to a calculated molecular mass of 45 847 Da per subunit. The deduced amino-acid sequence showed 93.6% and 87.2% identity with that of the human liver and porcine kidney, respectively.     

Differences in infantile growth patterns in Turner syndrome girls with and without spontaneous puberty.

OBJECTIVE: The role of prepubertal estrogen in child growth was modeled using Turner's syndrome, comparing growth patterns of girls who later did or did not enter puberty spontaneously. The hypothesis was that TS patients with normal prepubertal estrogen levels would have a different growth pattern from those with subnormal estrogen levels. STUDY DESIGN: Growth data from 78 full-term patients with Turner's syndrome were collected retrospectively. 24/78 later developed spontaneous puberty, (+Pub), and their growth data were compared to TS patients without spontaneous puberty (-Pub). A nonlinear mixed model was fitted using the bi-exponential model. RESULTS: The growth velocity difference between the -Pub and +Pub groups suggests an early infantile growth advantage in the -Pub group, which disappears before the end of the first year of life; growth velocity remains similar (+/- 1 cm/y) for the next 6 years and declines at age 7-8 years in the +Pub group faster than it does in the -Pub group. Bi-exponential analysis showed that both the 1st (restrictive) and 2nd exponent (forward) were different (p = 0.0003). CONCLUSIONS: Comparison of girls with or without spontaneous puberty suggests a role for estrogen in child growth. Estrogens restrict infantile growth, as well as growth during the mid-childhood spurt.