Fucosidosis: four new mutations and a new polymorphism

Fucosidosis is a rare lysosomal storage disease due to a nearlycomplete deficiency of -L-fucosidase (EC 3.2.1.51 [EC] ). In thisstudy, all 8 exons of the -L-fucosidase structural gene (FUCA-1)were amplified by PCR methods, and the amplified products weresubcloned and sequenced. Five patient groups with fucosidosiswere selected according to their ethnic backgrounds and haplotypesfor RFLPs in FUCA-1. Four presumptive disease causing mutationswere detected: 1) A major deletion of DNA containing the lasttwo exons of FUCA-1 in two Algerian siblings. 2) A G to T mutationin exon 6 resulting in an in-frame termination codon (E375X)in eight Hispanic patients from Colorado and New Mexico. 3)A G to A mutation (G60D) in exon 1 in four Italian patientsand in three related French-American (Cajun) patients. ThisG60D mutation creates a unique site for Afl III. 4) A frameshiftmutation resulted from a two-base deletion in exon 2 (K151fs)in an Italian patient. This deletion obliterates a unique BstXIsite and creates a new BpmI site, and was found in only thispatient and in only one allele. The rationale for proposingthese defects as disease causing mutations includes pedigreeanalysis and the predicted consequences of each defect uponthe activity and the concentration of the enzyme. An A to Gtransition (Q281R) in exon 5 was found to be present in homozygousform in affected patients and also in normal subjects; it appearsto be a newly identified polymorphism. It causes a charge changeand may be responsible for the electrophoretic variant phenotypeof fucosidosis. This polymorphism is inherited concordant withthe RFLP PvuII—BglI haplotype 2 – 2, 2 – 2.The 4 new mutations identified here comprise 70% of allelesof the 25 fucosidosis patients in our study.     

Production of 5–15 µm Diameter Alginate-Polylysine Microcapsules by an Air-Atomization Technique

A novel method of preparing small-sized microcapsules using a Turbotak air-atomizer is reported. Alginate-polylysine microcapsules containing Bacillus Calmette Guérin vaccine have been prepared by an adaptation of the method of Lim (1) which allows the manufacture of small-sized microcapsules. A Turbotak is used to spray sodium alginate solution into calcium chloride solution to form temporary calcium alginate microgel capsules. These temporary microgel droplets are subsequently cross-linked with polylysine to form permanent membranes. Microcapules in the size range of 5–15 µm have been produced which can be compared to an average diameter of 300 µm obtained by the method reported by Lim. The microcapsule size is dependent on the conditions of operation of the Turbotak and the concentration of the sodium alginate solution. Particles within the size range 5–15 µm can be reproducibly manufactured using the conditions of operation reported here. Other size ranges below the minimum of 300 µm reported by Lim are also feasible using this technique.     

Thymic Function and Output of Recent Thymic Emigrant T Cells During Intracranial Glioma Progression

One of the hallmarks of patients with glioblastoma multiforme (GBM) is profound lymphopenia mostly confined to the T cell lineage. A deficiency in the production of naïve T cells from the thymus could contribute to the lymphopenia seen in GBM patients. In this study we asked whether thymic function and the production of recent thymic emigrant (RTE) T cells from the thymus was influenced by intracranial (i.c.) glioma progression. We found significant thymic involution in animals with progressive i.c. gliomas. Involuted thymi from animals with progressive i.c. T9.F gliomas showed dramatic losses of CD4⁺CD8⁺ (DP) thymocytes. Microscopic analysis complemented those findings by demonstrating a reversal of the typical cortico-medullary structure. Significant increases in apoptosis accompanied the rapid loss of viable thymocytes, which was prevented in part by adrenalectomy, suggesting a dominant role for endogenous glucocorticoids. This thymic involution was also associated with a significant decrease in peripheral RTE T cells, reflecting the diminished thymic function. Finally, we found that CD8⁺ RTE T cells were enriched in progressively growing T9 gliomas, which points to an immunological role for RTE's in anti-glioma immunity. Our findings may shed light on the significance of thymic function for anti-glioma immunity and the response to immunotherapeutic treatment paradigms.     

Prognostic Indicators for Squamous Cell Carcinoma of the Oral Cavity: A Clinicopathologic Correlation

Fifty-three patients with T1 squamous cell cancer of the floor of mouth and ventral surface of the tongue with a known clinical outcome were retrospectively analyzed and arbitrarily divided into “aggressive” and “nonaggressive” groups based on their clinical behavior. Various host and tumor factors were then evaluated in an attempt to determine whether the tumor behavior could have been predicted. The paraffin-embedded tumor specimens were evaluated for tumor differentiation, tumor thickness and tumor invasion, microvessel density, and p53 expression. In addition, a composite morphologic grading score was obtained by combining cell differentiation, nuclear polymorphism, mitosis activity, depth of infiltration, type of infiltration, and lymphatic infiltration. No single technique appeared capable of identifying “aggressive” behavior, although possibly an evaluation of composite factors might show promise in the future.     

Tracheal Transplantation: Superior and Inferior Thyroid Artery Perfusion Territory

Objective: To determine the perfusion territories of the superior and inferior thyroid arteries in humans. Tracheal transplantation is a potential option for management of long-segment tracheal stenosis. However, the maximum length of vascularized trachea that can be reliably transplanted has not been established. Study Design: The tracheal vascular territory of individual superior and inferior thyroid arteries was determined separately in 10 humans postmortem. Methods: India ink was infused unilaterally under controlled pressure into the superior (n = 5) and inferior (n = 5) thyroid arteries of cadaveric tracheas. Tracheas were sectioned longitudinally and the caudalmost extent of mucosal dye staining was determined via microscopic assessment. Results: The tracheal perfusion territory of the superior thyroid artery was two to five rings (1.7 ± 0.5 cm) and the inferior thyroid artery, nine to 13 rings (6.5 ± 1.1 cm). In both cases, the tracheal mucosa on the contralateral side was stained to the same caudal level. Conclusions: The inferior thyroid artery was shown to perfuse the trachea maximally to the 13th ring (8.1 cm). As such, the unilateral inferior thyroid artery would serve as a suitable vascular component for long-segment tracheal transplantation in humans.     

Intra-arterial administration of carboplatin and the blood brain barrier permeabilizing agent,RMP-7: A toxicologic evaluation in swine

RMP-7 is a bradykinin B2 receptor agonist shown to permeabilize the blood-brain barrier, especially that associated with brain tumors, when administered via both intracarotid and intravenous routes. Both routes of administration are currently being tested in human trials in combination with the chemotherapeutic agent carboplatin as therapy for gliomas. As an essential prerequisite to the initial intracarotid clinical trials, the potential neurotoxicity of intra-arterial administration of RMP-7 (at a high or low dose), alone and in combination with carboplatin, was assessed in anesthetized Red Duroc swine. Five treatment groups were evaluated with each pig receiving a series of alternating, intra-arterial infusions of RMP-7 (or saline) followed by carboplatin (or saline), as follows: (1) vehicle control: saline/saline; (2) carboplatin only control: saline/carboplatin (50 mg total); (3) RMP-7 only control: RMP-7 (750 ng/kg)/saline; (4) low dose combination: RMP-7 (75 ng/kg)/carboplatin (50 mg total); and (5) high dose combination: RMP-7 (750 ng/kg)/carboplatin (50 mg total). For each subject, one of the alternating dosing sequences (above) was repeated four times during a single dosing session which lasted approximately 40 minutes. Assessments during the in-life phase of the study in the pre- and post-treatment periods consisted of heart rate, arterial blood pressure (systolic, diastolic, and mean), blood gases, body weight, general clinical observations (including evaluation for neurological deficit) and clinical pathology (including a comprehensive battery of standard blood coagulation, hematological and serum chemistry tests). In addition, during the time of treatment, heart rate and arterial blood pressure were monitored. The animals were terminated two weeks after dosing and the brain and rete mirabile (distal to site of infusion) were evaluated for gross and histopathological abnormalities. The histopathology analysis included a reader-blinded analysis using low and high power light microscopic examination of both H&E and Kluver-Berrera stained sections through several key cortical and subcortical brain regions. Transient decreases in arterial blood pressure (mean of 10–25 mmHg) were observed in both groups receiving the high dose of RMP-7 (i.e., 750 ng/kg). No other side effects attributable to RMP-7 and/or carboplatin were observed, and clinical observations revealed no evidence of neurologic deficits. Post-mortem examination revealed no evidence of CNS or cerebral vascular pathology attributable to carboplatin and RMP-7. This study demonstrates that intracarotid administration of the maximum tolerated dose of RMP-7 (750 ng/kg) alone, or in combination with carboplatin (50 mg) is not accompanied by any serious adverse effect, apparent cerebrovascular abnormality or neuropathologic consequence and offers further evidence for the safety of this novel therapeutic approach for enhancing delivery of chemotherapeutics to brain tumors.     

The UK Committee on Radioactive Waste Management.

The UK Committee on Radioactive Waste Management is charged with recommending to Government, by July 2006, options for the long term management of the UK's radioactive waste legacy. These options should inspire public confidence. Now, more than halfway into the time allotted, we, as two former members of the Committee, express our concerns at the wayward approach that has been adopted. The Committee has placed emphasis on gaining public confidence but this has been done at the expense of recruiting the best scientific expertise in the management of radioactive waste, an act which we believe will actually undermine public confidence. Furthermore, given also the immense importance of this decision to public safety, national security and the national interest, we believe urgent steps should be taken to review the Committee's process, its management and its sponsorship.     

The case-case-control study design: addressing the limitations of risk factor studies for antimicrobial resistance.

OBJECTIVE: There are significant limitations of the standard case-control study design for identifying risk factors for resistant organisms. The objective of this study was to develop a study design to overcome these limitations. DESIGN: Theoretical analysis of different types of study designs that can be used in risk factor studies for resistant organisms. RESULTS: We developed the case-case-control study design, which uses two separate case-control analyses within a single study. The first analysis compares patients infected with resistant bacteria (resistant cases) with control-patients without infection caused by the target organism, who are therefore representative of the source population; and the second analysis compares patients infected with the susceptible phenotype of the target organism (susceptible cases) with the same control-patients without infection caused by the target organism. These two analyses provide risk models for (1) isolation of the resistant phenotype of the target organism as compared with the source population and (2) isolation of the susceptible phenotype of the organism as compared with the source population. When these two risk models are compared and contrasted, risk factors specifically associated with isolation of the resistant phenotype can be identified. CONCLUSIONS: The case-case-control study design is an effective method for identifying risk factors for antimicrobial-resistant pathogens. Although the case-case-control study design has limitations, it is, in our opinion, more informative and less flawed than the standard case-control study design.     

Daptomycin: a novel cyclic lipopeptide antimicrobial.

PURPOSE: The development, activity, pharmacokinetics, pharmacodynamics, clinical efficacy, adverse effects, and dosage and administration of daptomycin are reviewed. SUMMARY: Daptomycin, a novel cyclic lipopeptide antimicrobial, is bactericidal against a range of gram-positive bacteria, including many multiple-drug-resistant isolates. It has only minimal activity against anaerobic bacteria and no activity against gram-negative bacteria. Daptomycin exhibits linear pharmacokinetics, and the plasma concentration-versus-time relationship is best described by a two-compartment model with first-order elimination. The initial bactericidal activity is rapid, extensive, and concentration related. In clinical trials, daptomycin has shown efficacy in treating complicated skin and skin-structure infections (CSSSIs); the drug carries FDA-approved labeling for same. The adverse effects of daptomycin appear comparable to those of vancomycin and semisynthetic penicillins. The dosage for CSSSIs is 4 mg/kg by i.v. infusion every 24 hours. CONCLUSION: Daptomycin is bactericidal against gram-positive organisms and offers an option in the treatment of CSSSIs.