The additivity of substrate fragments in enzyme-ligand binding

BACKGROUND: Enzymes have evolved to recognise their target substrates with exquisite selectivity and specificity. Whether fragments of the substrate--perhaps never available to the evolving enzyme--are bound in the same manner as the parent substrate addresses the fundamental basis of specificity. An understanding of the relative contributions of individual portions of ligand molecules to the enzyme-binding interaction may offer considerable insight into the principles of substrate recognition. RESULTS: We report 12 crystal structures of Escherichia coli thymidylate synthase in complexes with available fragments of the substrate (dUMP), both with and without the presence of a cofactor analogue. The structures display considerable fidelity of binding mode and interactions. These complexes reveal several interesting features: the cofactor analogue enhances the localisation of substrate and substrate fragments near the reactive thiol; the ribose moiety reduces local disorder through additional specific enzyme-ligand interactions; the pyrimidine has multiple roles, ranging from stereospecificity to mechanistic competence; and the glycosidic linkage has an important role in the formation of a covalent attachment between substrate and enzyme. CONCLUSIONS: The requirements of ligand-protein binding can be understood in terms of the binding of separate fragments of the ligand. Fragments which are subsystems of the natural substrate for the enzyme confer specific contributions to the binding affinity, orientation or electrostatics of the enzymatic mechanism. This ligand-binding analysis provides a complementary method to the more prevalent approaches utilising site-directed mutagenesis. In addition, these observations suggest a modular approach for rational drug design utilising chemical fragments.     

"Tower vertebra": a new observation in sickle cell disease

PURPOSE: We studied the frequency of testicular microlithiasis (TM) in a select group of otherwise healthy infertile men with abnormal semen. METHODS: We reviewed scrotal sonography records of 180 consecutive patients with abnormal semen who were referred for evaluation of male infertility. The pampiniform plexus was evaluated for possible varicocele, and the testes were examined for the presence or absence of intratesticular calcifications and/or masses. Five patients with more than 5 echogenic foci per transducer field in either testis were identified. The medical records and sonograms of this subgroup of patients were analyzed. RESULTS: Of the 180 patients, 5 (2.8%) had TM. Three patients with TM had concomitant small varicoceles; 2 others had a history of testicular maldescent. Two patients eventually became fertile. CONCLUSIONS: Our results indicate a higher than expected frequency of TM in otherwise healthy infertile men. We speculate that the magnitude of hypospermatogenesis in patients with TM is variable and may relate to both the degree of testicular dysgenesis and the presence or absence of concomitant scrotal pathology (eg, scrotal varicocele and cryptorchidism).     

Digital color imaging colposcopy: a matter of choice

PURPOSE: The objective of this study was to correlate the findings of sarcoidosis on high resolution CT (HRCT) with indexes of disease activity as measured with 67Ga scan, bronchoalveolar lavage (BAL), and serum angiotensin-converting enzyme (SACE) assay. METHOD: Twenty-nine patients with proven sarcoidosis underwent HRCT scan, 67Ga scan, BAL, and SACE assay within a 1 month period. The extent of parenchymal involvement by nodules, consolidation, ground-glass attenuation, and linear opacities was quantified to the nearest 10% of surface area affected on the CT examination. Whole-lung gallium uptake was quantified and the percentage of BAL-recovered lymphocytes (BAL-%LC) and SACE levels obtained by chart review. CT scores of disease extent were correlated with measured indexes of activity using the Spearman rank correlation coefficient. RESULTS: The mean extent of nodules, consolidation, ground-glass attenuation, and linear opacities on HRCT images was 15.1 +/- 16.6, 1.6 +/- 4.0, 17.5 +/- 25.4, and 7.6 +/- 9.6%, respectively. The extent of nodules and consolidation correlated with the intensity of lung gallium uptake (r = 0.46, p < 0.02), BAL-%LC (r = 0.50, p < 0.01), and SACE levels (r = 0.38, p < 0.05). No significant correlation was found between extent of ground-glass attenuation or linear opacities with any indexes of disease activity. CONCLUSION: On HRCT scan, nodules and consolidation in sarcoidosis reflect disease activity as measured by 67Ga scan, BAL, and SACE assay.     

Association of MSX1 and TGFB3 with nonsyndromic clefting in humans

Nonsyndromic cleft lip with or without cleft palate (CL/P) and nonsyndromic cleft palate only (CPO) are common congenital anomalies with significant medical, psychological, social, and economic ramifications. Both CL/P and CPO are examples of complex genetic traits. There exists sufficient evidence to hypothesize that disease loci for CL/P and CPO can be identified by a candidate-gene linkage-disequilibrium (LD) strategy. Candidate genes for clefting, including TGFA, BCL3, DLX2, MSX1, and TGFB3, were screened for LD with either CL/P or CPO in a predominantly Caucasian population, with both case-control- and nuclear-family-based approaches. Previously reported LD for TGFA with both CL/P and CPO could not be confirmed, except in CL/P patients with a positive family history. Also, in contrast to previous studies, no LD was found between BCL3 and either CL/P or CPO. Significant LD was found between CL/P and both MSX1 and TGFB3 and between CPO and MSX1, suggesting that these genes are involved in the pathogenesis of clefting. In addition, a mutation search in the genes DLX2, MSX1, and TGFB3 was performed in 69 CPO patients and in a subset of the CL/P patients. No common mutations were found in the coding regions of these genes; however, several rare variants of MSX1 and TGFB3 were found that may alter the latters' normal function. These results form the basis for future research, including (a) mutation searches in the MSX1 and TGFB3 genes in Caucasian CL/P patients and (b) extension of the search for MSX1 mutations in CPO patients to the noncoding regions.     

The effect of portal hypertension on transforming growth factor-alpha and epidermal growth factor receptor in the gastric mucosa of rats

BACKGROUND: The physical activity levels of US children are declining. Opportunities for physical activity within city schools are constrained by time and space limits. This study determined whether a supplemental program of physical activity would significantly alter the fitness levels of low-income, minority, urban elementary schoolchildren. METHODS: Ninety-nine students from two Cleveland Public Schools served as subjects. One school received a 15-week intervention program where teams of two medical students met with urban elementary schoolchildren three times a week for physical activity sessions. The other school served as a control and received no supplemental activity other than a regularly scheduled physical education class held once a week. We obtained field measurements of skinfold thickness, heart rate response to submaximal exercise, and sit and reach flexibility. RESULTS: The supplemental activity group showed significant improvements in flexibility, body composition, and heart rate response to submaximal exercise. CONCLUSIONS: This investigation indicates that a program of fitness activities conducted within the classroom can significantly improve levels of fitness in urban elementary schoolchildren.     

1,4-Cyclohexanecarboxylates: potent and selective inhibitors of phosophodiesterase 4 for the treatment of asthma

Evaluation of a variety of PDE4 inhibitors in a series of cellular and in vivo assays suggested a strategy to improve the therapeutic index of PDE4 inhibitors by increasing their selectivity for the ability to inhibit PDE4 catalytic activity versus the ability to compete for high affinity [3H]rolipram-binding sites in the central nervous system. Use of this strategy led ultimately to the identification of cis-4-cyano-4-[3-(cyclopentyloxy)-4-methoxyphenyl]cyclohexane-1-carboxyl ic acid (1, SB 207499, Ariflo), a potent second-generation inhibitor of PDE4 with a decreased potential for side effects versus the archetypic first generation inhibitor, (R)-rolipram.     

Biphasic spiral computed tomography versus digital subtraction angiography for evaluation of arterial thrombosis after orthotopic liver transplantation

Aseptic loosening of implant components is a common and important complication of both cemented and uncemented prosthetic joint replacements. Wear particles derived from organic polymer and metal implant biomaterials are commonly found within macrophages and macrophage polykaryons in the fibrous membrane between loose implant components and the host bone undergoing resorption. In order to determine whether biomaterial particle-containing, foreign-body macrophages may contribute to periprosthetic bone resorption, we cultured murine monocytes that had phagocytosed particles of biomaterials commonly employed in bone implant surgery [polymethylmethacrylate (PMMA), ultra-high molecular weight polyethylene (PE), titanium and chromium-cobalt] on bone slices and glass coverslips with UMR 106 osteoblast-like stromal cells in the presence of 1,25-dihydroxy-vitamin D3. Under these conditions, all biomaterial particle-containing, foreign-body macrophages differentiated into osteoclastic cells, i.e. tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells capable of extensive lacunar bone resorption. This study shows that particle phagocytosis by macrophages does not abrogate the ability of these cells to undergo osteoclast differentiation. These findings emphasise the importance of the foreign-body macrophage response to biomaterial wear particles in the pathogenesis of aseptic loosening.     

Acute coronary occlusion and in-stent thrombosis in a patient with essential thrombocythemia

BACKGROUND: Physical activity in old age probably protects against hip fracture. However, very little is known about the impact of occupation during working life, in terms of its physical activity level and the socio-economic status (SES) it indicates, on the risk of hip fracture in later life. METHOD: A population-based case-control study was conducted on 416 subjects to investigate relationships between risk of hip fracture and occupational physical activity and occupation-related SES. Occupational physical activity was coded according to the proportion of the working day at ages 20 and 50 years that the subjects were likely to have spent sitting. The Australian Classification of Standard Occupations (ASCO) was used as the basis for the coding of SES, indicated by job titles. RESULTS: Compared with those in the intermediate category, women with a sedentary job at age 50 years (odds ratio (OR) 7.2, 95 per cent confidence interval (CI) 1.2-41.9) or a mainly weight-bearing job (OR 5.1, 95 per cent CI 1.1-23.2) had an increased risk of hip fracture. A decreasing hip fracture risk with increasing SES of the longest held job was also observed. CONCLUSION: The finding of a J-shaped relationship in women between occupational physical activity and risk of hip fracture supports the need for research into the most effective types of exercise for maintenance of bone strength. Furthermore, the protective effect of SES demonstrated in this study still need to be corroborated in other studies before establishing evidence of a causal relationship.     

Lesion location influences perception of homonymous scotomata during flickering random dot pattern stimulation

An attempt was made to clarify whether the site of postchiasmal lesions affects subjective perception of homonymous visual field defects during stimulation with flickering random dot patterns (white noise-field). Out of 56 patients with homonymous hemianopia, 38 (68%) perceived scotomata in this situation, but 18 (32%) discerned none at all. Neuroradiologic superposition of cerebral lesions detected by computed tomography (CT) or magnetic resonance imaging (MRI) showed that nearly all patients who perceived their scotomata had lesions involving the primary visual cortex or the perigeniculate region, whereas those who received no scotoma had lesions centered within the optic radiation. Functional MRI of six normal subjects during stimulation with flickering random dot patterns indicated predominant activation of the primary visual cortex. Since noise-field defects were most frequently perceived by patients whose lesion involved the primary visual cortex, it appears that the sensitivity of noise-field campimetry depends on the site of damage in the visual pathway. The explanation for this may be that damage to long-range horizontal connections impairs filling-in processes.