Recessive amyotrophic lateral sclerosis families with the D90A SOD1 mutation share a common founder: evidence for a linked protective factor

Amyotrophic lateral sclerosis (ALS) is a progressive motor neurodegeneration resulting in paralysis and death from respiratory failure within 3-5 years. About 20% of familial cases are associated with mutations in the gene for copper/zinc superoxide dismutase ( SOD1 ), which catalyses the dismutation of the superoxide radical to hydrogen peroxide and oxygen. Experimental evidence suggests mutations act by a toxic gain of function but the mechanism is unknown. There are >60 known SOD1 mutations associated with ALS and all are dominant except for one in exon 4, a D90A substitution which is recessive. D90A pedigrees with dominant inheritance have now been reported and this apparent contradiction needs to be explained. We performed a worldwide haplotype study on 28 D90A pedigrees using six highly polymorphic microsatellite markers. We now show that all 20 recessive families share the same founder (alpha = 0.999), regardless of geographical location, whereas several founders exist for the eight dominant families (alpha = 0.385). This finding confirms that D90A can act in a dominant fashion in keeping with all other SOD1 mutations, but that on one occasion, a new instance of this mutation has been recessive. We propose a tightly linked protective factor which modifies the toxic effect of mutant SOD1 in recessive families.      

A clinical classification of the status of the pulp and the root canal system

Many different classification systems have been advocated for pulp diseases. However, most of them are based on histopathological findings rather than clinical findings which leads to confusion since there is little correlation between them. Most classifications mix clinical and histological terms resulting in misleading terminology and diagnoses. This in turn leads to further confusion and uncertainty in clinical practice when a rational treatment plan needs to be established in order to manage a specific pathological entity. A simple, yet practical classification of pulp diseases which uses terminology related to clinical findings is proposed. This classification will help clinicians understand the progressive nature of the pulp disease processes and direct them to the most appropriate and conservative treatment strategy for each condition. With a comprehensive knowledge of the pathophysiology of pain and inflammation in the pulp tissues, clinicians may accomplish this task with confidence.     

The use of MTT [3-(4, 5-dimethyl-2-thiazolyl) -2, 5-diphenyl -2H- tetrazolium bromide]-reduction as an indicator of the effects of strain-specific, polyclonal rabbit antisera on Candida albicans and C. krusei

There is only scanty data on the effects of specific antibody, with or without complement, on Candida albicans or Candida krusei in cell-free systems in vitro, although previously published work has shown that specific antibody mediates anti- Candida immunity in vivo by inhibition of adherence to host cells or surfaces and by the promotion of phagocytosis and intra-phagocytic killing. The MTT (3-[4, 5-dimethyl-2-thiazolyl] -2, 5-diphenyl -2H- tetrazolium bromide)-reduction method as a test of the viability of fungi was used to investigate the effect of complement, normal serum and immune serum on these two species of Candida that are of increasing importance as opportunistic pathogens. We report that normal rabbit serum or strain-specific, polyclonal anti- Candida rabbit antibody, with or without guinea pig complement, did not cause the reduction of total cell-mass or of the viability of either C. albicans or C. krusei, in vitro as determined by the MTT-reduction test. Complement alone without specific antibody, also, had no such effect on these two Candida species.     

联合抗击输血相关的GVHD

输血相关的GVHD(TA-GVHD)从发病率极低的意义上来说它是不常见的,但是从它是个值得我们协作的全球性问题的意义来说,它又是不容忽视的。本期本栏目刊登了两篇TA-GVHD文章:Aoun等报告贝鲁特美国大学医学中心(AUBMC,黎巴嫩)对10年记录回顾性研究10例GVHD病例,Leitman等报告美国4例自身免疫性疾病的患者用氟达拉     

Pain Management in Pediatric Age Group

The management of pain in palliative care of children is somewhat different from that in adults. The use of opioids in pediatric palliative care presents some unique challenges. Confident and rational use of opioids, illustrated by WHO Guidelines is essential for adequate management of pain in children with life limiting conditions.     

Surgical hypothermia in a patient with a cold agglutinin. Management by plasma exchange

Cold agglutinins are a potential danger to patients who must be subjected to hypothermia. A patient with a cold agglutinin of moderate titer but broad thermal amplitude was to undergo hypothermia during aortic valve replacement. He was managed preoperatively with an eight- liter plasma exchange by continuous-flow centrifugation to remove the cold agglutinin. There were no adverse effects during or after hypothermia.     

Structure-Activity Relationship for Thiirane-Based Gelatinase Inhibitors

An extensive structure-activity relationship study with the template of 2-(4-phenoxyphenylsulfonylmethyl)thiirane (1), a potent and highly selective inhibitor for human gelatinases, is reported herein. Syntheses of 65 new analogs, each in multistep processes, allowed for exploration of key structural components of the molecular template. This study reveals that the presence of the sulfonylmethylthiirane and the phenoxyphenyl group were important for gelatinase inhibition. However, para- and some meta-substitutions of the terminal phenyl ring enhanced inhibitory activity, and led to improve metabolic stability. This agrees with the result from metabolism studies with compound 1 that the primary route of biotransformation is oxidation, mainly at the para position of the phenyl ring and alpha position of the sulfonyl group in the aliphatic side chain.     

How allosteric control of Staphylococcus aureus penicillin binding protein 2a enables methicillin resistance and physiological function

The expression of penicillin binding protein 2a (PBP2a) is the basis for the broad clinical resistance to the β-lactam antibiotics by methicillin-resistant Staphylococcus aureus (MRSA). The high-molecular mass penicillin binding proteins of bacteria catalyze in separate domains the transglycosylase and transpeptidase activities required for the biosynthesis of the peptidoglycan polymer that comprises the bacterial cell wall. In bacteria susceptible to β-lactam antibiotics, the transpeptidase activity of their penicillin binding proteins (PBPs) is lost as a result of irreversible acylation of an active site serine by the β-lactam antibiotics. In contrast, the PBP2a of MRSA is resistant to β-lactam acylation and successfully catalyzes the dd-transpeptidation reaction necessary to complete the cell wall. The inability to contain MRSA infection with β-lactam antibiotics is a continuing public health concern. We report herein the identification of an allosteric binding domain—a remarkable 60 Å distant from the dd-transpeptidase active site—discovered by crystallographic analysis of a soluble construct of PBP2a. When this allosteric site is occupied, a multiresidue conformational change culminates in the opening of the active site to permit substrate entry. This same crystallographic analysis also reveals the identity of three allosteric ligands: muramic acid (a saccharide component of the peptidoglycan), the cell wall peptidoglycan, and ceftaroline, a recently approved anti-MRSA β-lactam antibiotic. The ability of an anti-MRSA β-lactam antibiotic to stimulate allosteric opening of the active site, thus predisposing PBP2a to inactivation by a second β-lactam molecule, opens an unprecedented realm for β-lactam antibiotic structure-based design.The inexorable spread of bacterial resistance mechanisms against β-lactam antibiotics is a critical clinical concern. The resistance mechanism used by methicillin-resistant Staphylococcus aureus (MRSA) is acquisition of a set of genes that are induced on β-lactam exposure (1, 2). The key resistance enzyme is a unique, monofunctional dd-transpeptidase designated as penicillin binding protein 2a (PBP2a) that is refractory to inhibition by virtually all β-lactam antibiotics. In the MRSA bacterium, PBP2a catalyzes, in concert with the transglycosylase activities of other penicillin binding proteins (PBPs), the biosynthesis of the bacterial cell wall (3–5). The structure of this cell wall is a peptidoglycan polymer, comprised of glycan strands consisting of a repeating disaccharide motif [N-acetylglucosamine-N-acetylmuramylpentapeptide (NAG-NAM pentapeptide)], wherein adjacent glycan strands are cross-linked by PBP2a using peptide stems found on each NAM saccharide. The cell wall encases the entire bacterium as a single molecule, and its integrity is indispensible to the organism’s survival (6). The PBPs are the lethal targets of the β-lactam antibiotics as a result of irreversible acylation of the active site serine.The earlier structure determination for PBP2a (7) showed a closed active site conformation. Because the dd-transpeptidase site must accommodate two strands of peptidoglycan simultaneously—requiring an active site volume in excess of 1,000 ų (8, 9)—there must also exist an open conformation for PBP2a, which was preceded by structural studies with other PBPs (10–14). The mechanistic paradox in the case of PBP2a is not simply separate open and closed states but a mechanism that biases the open state to the peptidoglycan. Nature often resolves these paradoxes by allostery (15). Indeed, we have shown that synthetic samples of the peptidoglycan bind to PBP2a in a saturable manner and effect a conformational change that correlates to faster rates for PBP2a inactivation by β-lactams with enhanced PBP2a affinity (16). These observations suggested a structural model, wherein the binding of nascent peptidoglycan to a remote allosteric site opens the active site to both substrates and β-lactam inactivators. We disclose X-ray structures of PBP2a that confirm the presence of this allosteric site, reveal its location as 60 Å removed from the active site, and identify its ligands. Moreover, binding of these ligands to the allosteric site imparts conformational opening of the active site. Lastly, we document that ceftaroline, a new β-lactam antibiotic (Fig. 1A) that recently has received Food and Drug Administration approval for use in the treatment of MRSA infections, has the ability to trigger this conformational change and thus, enables access to the active site by a second ceftaroline molecule. These observations explain the mechanism for the manifestation of the physiological role of PBP2a, explain the advantageous anti-MRSA activity of ceftaroline, identify the basis for understanding emerging mutations in the gene for PBP2a that confer resistance to ceftaroline, and provide the context for future structure-based design of anti-MRSA β-lactams that will evade these mutations.Open in a separate windowFig. 1.Domains of PBP2a and key ligands. (A) The chemical structures of a synthetic NAG-NAM(pentapeptide) (1) and ceftaroline (2). The R1 and R2 groups of 2 are labeled. (B) Ribbon representation of PBP2a acylated by ceftaroline. The N-terminal extension is colored in green, the remaining allosteric domain is colored in gold, and the transpeptidase (TP) domain is colored in blue. These domain colors are retained in all other figures. Two molecules of ceftaroline (capped sticks in red) are found in complex with protein: one covalently bound as an acyl-enzyme in the TP domain (CFT1) and one intact at the allosteric domain (CFT2). A muramic acid saccharide (capped sticks in magenta) is found at the center of the allosteric domain. The arrow indicates the point of attachment of the membrane anchor. (C) The solvent-accessible surface representation for PBP2a is shown. The distance between the two ceftaroline molecules is 60 Å. (D) Ribbon representation of PBP2a in complex with 1 (black sticks). This view is rotated ∼45° on the y axis compared with the view of C.