The dynamics of germinal centre selection as measured by graph-theoretical analysis of mutational lineage trees

We have developed a rigorous graph-theoretical algorithm for quantifying the shape properties of mutational lineage trees. We show that information about the dynamics of hypermutation and antigen-driven clonal selection during the humoral immune response is contained in the shape of mutational lineage trees deduced from the responding clones. Age and tissue related differences in the selection process can be studied using this method. Thus, tree shape analysis can be used as a means of elucidating humoral immune response dynamics in various situations.     

A collection of 33 novel human mtDNA homoplasmic variants

Mitochondria are involved in cellular energy production via oxidative phosphorylation and this function may be damaged by any mutation in mitochondrial DNA (mtDNA). To identify novel mtDNA mutations, we have developed a program to systematically screen the entire mitochondrial genome in a large number of individuals with clinical and/or morphological features of mitochondrial dysfunction, but still no genetic diagnosis. The sequence-data were obtained with an automated rapid system, which gave us a series of information: in the eleven mitochondrial genomes analyzed we observed the presence of 33 differences from the revised Cambridge Reference Sequence (Andrews et al., 1999), but they were all homoplasmic in the patients' tissues analyzed (skeletal muscle and blood), suggesting that they are unlikely to be primarily pathogenic though they may be co-responsible in the determination of the disease. This work can therefore help complete the already ample mtDNA polymorphism existent database.     

A compilation of chiropractic and complementary/alternative medicine (CAM) data from public-use national surveys: report on a health-services research resource for the chiropractic and CAM scientific community

BACKGROUND: Federally funded national surveys are routinely conducted to provide reliable, valid, and relevant data on health and health care, and these "public-use" survey data are typically made available for further study by the wider scientific community. The full potential for using such data to examine the delivery, utilization, organization, and costs of chiropractic or complementary/alternative (CAM) health care remains largely untapped. OBJECTIVE: To report on a project that identifies and indexes public-use survey databases that contain explicit reference to chiropractic and CAM health care, and compiles that information into a web-based resource for the scientific community. METHODS: Review of database source collections. RESULTS: The utility and efficiency of secondary analyses as a cost-effective research strategy are well appreciated within the larger health-services research community, creating many possible opportunities for productive cooperative research endeavors across scientific disciplines. CONCLUSION: The Chiropractic and Complementary/Alternative Compilation User's Manual is available for free download at http://w3.palmer.edu/carber/manualhome.asp, or by following the links at the Palmer Center for Chiropractic Research homepage.     

Evaluation of Toxoplasma gondii recombinant proteins for the diagnosis of recently acquired toxoplasmosis by an immunoglobulin G analysis

The value of T. gondii recombinant antigens rRop2, rGra4, rGra7 and rSAG1m (mature version) or rSAG1ct (C-terminal version) in differentiating recently acquired from chronic infections was determined by IgG-ELISA. The general highest sensitivity was observed with rRop2 whereas rSAG1m was not recognized by any of the serum samples, suggesting an incorrect folding. rGra4 and rGra7 showed significant higher sensitivity and absorbance values with serum samples from recently infected individuals compared to those with chronic infection. In contrast, rRop2 and rSAG1ct did not show differences in the reactivity pattern between both groups of serum samples.     

The differential contribution of granzyme A and granzyme B in cytotoxic T lymphocyte-mediated apoptosis is determined by the quality of target cells

Complementary approaches with purified molecules or transfected cytolytic effector cells have suggested that both, granzyme A (gzmA) and granzyme B (gzmB), similarly contribute to CTL-mediatedand perforin (perf)-dependent apoptotic nuclear damage (DNA fragmentation) in target cells. Studies employing gzmA or gzmB single-knockout mice on the other hand indicated that gzmB is the prominent CTL effector molecule for the rapid induction of DNA fragmentation, with gzmA playing only a minor part. We have now taken ex vivo-derived virus-specific or in vitro generated alloreactive CTL from mice deficient in either gzmA or gzmB and a panel of three target cells to reinvestigate this unresolved issue. We show that rapid CTL-mediated DNA fragmentation of L1210.3 target cells is solely dependent on gzmB, whereas the DNA fragmentation of EL4.F15 target cells by the same CTL population is mainly induced by gzmA and only marginally by gzmB. Moreover, CTL-induced apoptosis of a third target cell, MC57G, was partially dependent on both gzmA and gzmB activities. The differential contribution of the two gzms to apoptosis was further verified by their distinct sensitivity tocaspase inhibitors. The data suggest that both, gzmA and gzmB, have a similar potential to induce rapid perf-mediated apoptosis but that their individual contribution to the underlying intracellular processes is dictated by the quality of the target cell.     

Effect of triethylamine on the recovery of selected South American alphaviruses,flaviviruses, and bunyaviruses from mosquito (Diptera: Culicidae) pools

We evaluated the effect of triethylamine (TEA) on the recovery of infectious virus from pools of mosquitoes for two South American alphaviruses (eastern equine encephalomyelitis and Venezuelan equine encephalomyelitis subtypes IIIC and ID), one flavivirus (Ilheus) and two bunyaviruses (Mirim [Guama group] and Itaqui [group C]). Mosquitoes were inoculated intrathoracically with virus, held for 7-10 d at 26 degrees C, and handled under one of four regimens before testing for the presence of virus by plaque assay. Mosquitoes were killed by freezing at - 70 degrees C for 3 min and tested immediately for the presence of virus; killed by freezing at -70 degrees C for 3 min and then held at room temperature for 1 h before testing for the presence of virus; anesthetized with TEA and assayed immediately for the presence of virus; or anesthetized with TEA and then held at room temperature for 1 h before being assayed for the presence of virus. For each of the viruses tested, viral titers in mosquitoes anesthetized with TEA were similar to those in mosquitoes killed by freezing at-70 degrees C. Likewise, there was no significant difference in viral titers in mosquitoes anesthetized with TEA and held at room temperature for 1 h or in mosquitoes frozen at -70 degrees C and held at room temperature for 1 h before being processed for virus by isolation. Triethylamine is advantageous for the handling of mosquitoes in a field environment. The elimination of the need for a cold chain, without compromising virus recovery, increases the feasibility of conducting research projects requiring the isolation of live virus from mosquitoes in remote tropical environments.     

Langerhans' cell histiocytosis: various manifestations with the same histopathologic base

Langerhans' cell histiocytosis (LCH), previously known as histiocytosis X, is a rare disease. It is characterized by the accumulation and proliferation of histiocytes, eosinophils and Langerhans' cells with Birbeck granules detected by electron microscopy. It involves single organs or systems or can present as a multisystem disease. The clinical presentation may vary widely, ranging from benign self-limiting types with spontaneous regression to slowly-progressive malignant disease. We report five cases of LCH with the same histopathologic basis but different outcome.     

Initiation of the breakage-fusion-bridge mechanism through common fragile site activation in human breast cancer cells: the model of PIP gene duplication from a break at FRA7I

Gene amplification plays a critical role in tumor progression. Hence, understanding the factors triggering this process in human cancers is an important concern. Unfortunately, the structures formed at early stages are usually unavailable for study, hampering the identification of the initiating events in tumors. Here, we show that the region containing the PIP gene, which is overexpressed in 80% of primary and metastatic breast cancers, is duplicated in the breast carcinoma cell line T47D. The two copies are organized as a large palindrome, lying 'in loco' on one chromosome 7. Such features constitute the landmark of the breakage-fusion-bridge (BFB) cycle mechanism. In hamster cells selected in vitro to resist cytotoxic drugs, common fragile site (CFS) activation has been shown to trigger this mechanism. Here, we characterize FRA7I at the molecular level and demonstrate that it lies 2 Mb telomeric to the PIP gene and sets the distal end of the repeated sequence. Moreover, our results suggest that the BFB process was frozen within the first cycle by healing of the broken chromosome. T47D cells thus offer a unique opportunity to observe the earliest products of the BFB cycle mechanism. Our findings constitute the first evidence that this amplification mechanism can be initiated in vivo by fragile site activation.