Troponin C modulates the activation of thin filaments by rigor cross-bridges

Extraction of troponin C (TnC) from skinned muscle fibers reduces maximum Ca2+ and rigor cross-bridge (RXB)-activated tensions and reduces cooperativity between neighboring regulatory units (one troponin-tropomyosin complex and the seven associated actins) of thin filaments. This suggests that TnC has a determining role in RXB, as well as in Ca(2+)-dependent activation processes. To investigate this possibility further, we replaced fast TnC (fTnC) of rabbit psoas fibers with either CaM[3,4TnC] or cardiac TnC (cTnC) and compared the effects of these substitutions on Ca2+ and RXB activation of tension. CaM[3,4TnC] substitution has the same effect on Ca(2+)- and RXB-activated tensions; they are reduced 50%, and cooperativity between regulatory units is reduced 40%. cTnC substitution also reduces the maximum Ca(2+)-activated tension and cooperativity. But with RXB activation the effects on tension and cooperativity are opposite; cTnC substitution potentiates tension but reduces cooperativity. We considered whether tension potentiation could be explained by increased activation by cycling cross-bridges (CXBs), but the concerted transition formalism predicts fibers will fail to relax in high substrate and high pCa when CXBs are activator ligands. It predicts resting tension, which is not observed in either control or cTnC-substituted fibers. Rather, it appears that cTnC facilitates RXB activation of fast fibers more effectively than fTnC. The order of RXB-activated tension facilitation is cTnC > fTnC > CaM[3,4TnC] > empty TnC-binding sites. Comparison of the structures of fTnC, CaM[3,4TnC], and cTnC indicates that the critical region for this property lies in the central helix or N-terminal domain, including EF hand motifs 1 and 2.     

Cholinergic strategies for Alzheimer's disease

Alzheimer's disease is a devastating degenerative disorder of the central nervous system that results in gradual deterioration of cognitive function and severe alteration of personality. Degeneration of neurons in the nucleus basalis Meynert, the origin of the major cholinergic projections to the neocortex, occurs early in the course of the disease, and is correlated with the cognitive decline. This link between cholinergic dysfunction in the basal-cortical system and cognitive deficits has focused scientific efforts on developing tools to elucidate the neurobiological role of the cholinergic system in cognition and to develop therapeutic interventions in the disorder. An important step in understanding the mechanisms underlying cognitive dysfunction has been the development of in vivo rodent models that mimic some of the features of Alzheimer's disease. Acute excitotoxic or immunotoxic lesions of the nucleus basalis in rodents have revealed a role of the basal-cortical system in attention, learning and memory. More recent advances in developing mouse gene technology offer newer models to systematically examine the underlying neuropathological cascade leading to dysfunctions in mnemonic processing. Using in vivo rodent models, several cholinergic enhancement strategies have been tested and proven to be effective in alleviating lesion-induced cognitive deficits, including neuropharmacological approaches (acetylcholinesterase inhibitors), neurotrophic factor administration (nerve growth factor), and transplantation of cholinergic-enriched fetal grafts. Successful results have also been obtained using ex vivo gene transfer to deliver nerve growth factor or acetylcholine to compromised regions of the basal-cortical system. Gene therapy may be of particular interest for clinical applications, because this approach provides a method for topographically restricted and selective delivery of therapeutic genes and their products to afflicted areas of the brain. Advanced techniques in molecular biology (e.g., exogenous regulatable gene transfer) and newly developed tools of modern neuroscience (e.g., neural precursor cells) will be important contributions for deciphering the biological bases of neuronal degeneration and for refining therapeutic strategies for Alzheimer's disease.     

Children with minimal sensorineural hearing loss: prevalence, educational performance, and functional status

OBJECTIVE: This study was designed to determine the prevalence of minimal sensorineural hearing loss (MSHL) in school-age children and to assess the relationship of MSHL to educational performance and functional status. DESIGN: To determine prevalence, a single-staged sampling frame of all schools in the district was created for 3rd, 6th, and 9th grades. Schools were selected with probability proportional to size in each grade group. The final study sample was 1218 children. To assess the association of MSHL with educational performance, children identified with MSHL were assigned as cases into a subsequent case-control study. Scores of the Comprehensive Test of Basic Skills (4th Edition) (CTBS/4) then were compared between children with MSHL and children with normal hearing. School teachers completed the Screening Instrument for Targeting Education Risk (SIFTER) and the Revised Behavior Problem Checklist for a subsample of children with MSHL and their normally hearing counterparts. Finally, data on grade retention for a sample of children with MSHL were obtained from school records and compared with school district norm data. To assess the relationship between MSHL and functional status, test scores of all children with MSHL and all children with normal hearing in grades 6 and 9 were compared on the COOP Adolescent Chart Method (COOP), a screening tool for functional status. RESULTS: MSHL was exhibited by 5.4% of the study sample. The prevalence of all types of hearing impairment was 11.3%. Third grade children with MSHL exhibited significantly lower scores than normally hearing controls on a series of subtests of the CTBS/4; however, no differences were noted at the 6th and 9th grade levels. The SIFTER results revealed that children with MSHL scored poorer on the communication subtest than normal-hearing controls. Thirty-seven percent of the children with MSHL failed at least one grade. Finally, children with MSHL exhibited significantly greater dysfunction than children with normal hearing on several subtests of the COOP including behavior, energy, stress, social support, and self-esteem. CONCLUSIONS: The prevalence of hearing loss in the schools almost doubles when children with MSHL are included. This large, education-based study shows clinically important associations between MSHL and school behavior and performance. Children with MSHL experienced more difficulty than normally hearing children on a series of educational and functional test measures. Although additional research is necessary, results suggest the need for audiologists, speech-language pathologists, and educators to evaluate carefully our identification and management approaches with this population. Better efforts to manage these children could result in meaningful improvement in their educational progress and psychosocial well-being.     

Development of seven new human prostate tumor xenograft models and their histopathological characterization

Seven human prostate tumor models were established by transplanting tumor fragments in NMRI athymic nude mice. Once established, the tumors were serially transplantable in both NMRI and BALB/c nude mice. The xenografts originated from primary prostatic carcinomas (prostatectomy specimens), transurethral resection material, and metastatic lesions (pelvic lymph nodes and scrotal skin). Histological examination revealed that, in the course of several mouse passages (8 to 23), tumors retained their resemblance to the original patient material. The PC-295, PC-310, PC-329, and PC-346 tumors are dependent on androgens for their growth. The PC-324, PC-339, and PC-374 tumors are androgen independent, although growth of PC-374 tumors still seemed androgen sensitive. All tumors are diploid, except for the PC-374, which is tetraploid. The diploid PC-295 tumor has an additional small population of tetraploid cells. All xenografts displayed a heterogeneous expression pattern of the androgen receptor except for the PC-324 and PC-339 tumors in which the androgen receptor could not be detected. Prostatic acid phosphatase and prostate-specific antigen were retained during serial transplantation in all tumors but the PC-324 and PC-339. This panel of permanent human prostate tumor models comprises tumors representing both the androgen-dependent and -independent stages of human prostate cancer with various degrees of differentiation and, therefore, is of great value for the study of many aspects of growth and progression of human prostate cancer.     

Nonlinear dynamics of epileptic seizures on basis of intracranial EEG recordings

PURPOSE: An understanding of the principles governing the behavior of complex neuronal networks, in particular their capability of generating epileptic seizures implies the characterization of the conditions under which a transition from the interictal to the ictal state takes place. Signal analysis methods derived from the theory of nonlinear dynamics provide new tools to characterize the behavior of such networks, and are particularly relevant for the analysis of epileptiform activity. METHODS: We calculated the correlation dimension, tested for irreversibility, and made recurrence plots of EEG signals recorded intracranially both during interictal and ictal states in temporal lobe epilepsy patients who were surgical candidates. RESULTS: Epileptic seizure activity often, but not always, emerges as a low-dimensional oscillation. In general, the seizure behaves as a nonstationary phenomenon during which both phases of low and high complexity may occur. Nevertheless a low dimension may be found mainly in the zone of ictal onset and nearby structures. Both the zone of ictal onset and the pattern of propagation of seizure activity in the brain could be identified using this type of analysis. Furthermore, the results obtained were in close agreement with visual inspection of the EEG records. CONCLUSIONS: Application of these mathematical tools provides novel insights into the spatio-temporal dynamics of "epileptic brain states". In this way it may be of practical use in the localization of an epileptogenic region in the brain, and thus be of assistance in the presurgical evaluation of patients with localization-related epilepsy.     

Resistance to beta-lactam antibiotics and aminoglycosides in gram negative bacteria. 1. Molecular and genetic characterization of R-factors (author's transl)

With frequent use of aminoglycoside antimicrobials and beta-lactam antibiotics in hospitals in the last few years, the number of bacterial strains resistant to these chemotherapeutics increased. Lately, strains of E. coli, Klebsiella, Enterobacter, Serratia, Proteus and Pseudomonas resistant to many antimicrobials (ampicillin, carbenicillin, cephalothin, chloramphenicol, gentamycin, tobramycin, sisomycin, neomycin, paromomycin, kanamycin, streptomycin, spectinomycin, tetracycline, sulphonamides) were isolated from patients of the university hospital in Zuerich. The resistant phenotype of two representative strains (Klebsiella pneumoniae 1 and Serratia marcescens 2) could be transferred by mixed cultivation to E. coli K-12. Multiple resistance of strain 1, and addition, could be transferred to Salmonella typhimurium, Serratia marcescens, Providencia, Proteus mirabilis and Klebsiella pneumoniae in varying frequencies. Transfer to Pseudomonas aeruginosa, however, could not be achieved. Spontaneous instability of resistance was observed in 0.15% of the cells of an overnight brothe culture and in 90% of the cells of a three months old culture. Conjugation, instability and the response to the sex phages MS-2 and If-1 suggested that resistance was mediated by a monomolecular R-factor, belonging to the fi+-type. This suggestion was confirmed by molecular characterization of the resistance plasmids. After transfer of the R-factors of K. pneumoniae 1 (R-FK 1) and Serratia marcescens 2 (R-FK2) into E. coli K-12, plasmid DNA was labelled with (methyl-3H) thymidine, and isolated by isopycnic centrifugation in cesiumchlorid-ethidium-bromide. Analysis of plasmid DNA then was carried out by sedimentation in a 5-20% neutral sucrose gradient together with reference plasmids of known molecular weights and sedimentation constants. The analysis revealed that R-FK1 had a molecular weight of 54 X 10(6) and R-FK2 of 50 X 10(6) daltons. The values were confirmed by contour length measurements of open circular forms with an electron microscope. A comparison of the sedimentation profile of labelled plasmid DNA from strain 1 and 14C-labelled DNA of E. coli K-12 (R-FK1) showed that the wild-type strain contained, besides the large resistance plasmid, at least two smaller "cryptic" plasmids. These smaller plasmid molecules were also found in antibiotic susceptible variants of strain 1, which did not contain the 54 X 10(6) dalton plasmid molecule, responsible for the resistant phenotype. The number of copies of R-FK1 in E. coli K-12 was determined to be 2, indicating stringent control of replication. It is discussed that the growing number of isolations of strains of Escherichia, Klebsiella, Serratia, Proteus, Providencia and Pseudomonas, exhibiting the same resistance phenotype, results from the spread of the R-factor described above among the hospital bacterial flora.     

Brain "ouabain", a neurosteroid, mediates sympathetic hyperactivity in salt-sensitive hypertension

This review addresses recent developments in the neurobiology of an endogenous inhibitor of brain Na+, K+ - ATPase, "ouabain". "Ouabain" is present in hypothalamic and medullary neurons and mediates sympathoexcitatory and pressor responses to acute and chronic increases in cerebrospinal fluid (CSF) sodium concentration as well as mediates the sympathoexcitatory and pressor responses to high dietary sodium intake in SHR and Dahl-S rats, and sympathetic hyperactivity in the congestive heart failure. Some of these actions of "ouabain" in the CNS take place in the median preoptic nucleus and ventral part of the AV3V region. Despite recent advances in unveiling a biological role for "ouabain" its structure, biosynthetic and metabolic pathways as well as actual control mechanisms remain unresolved.     

HLA (A*0201) mimicry by anti-idiotypic monoclonal antibodies

Soluble MHC Ags and anti-Id (anti-anti-MHC) Abs have both been shown to inhibit MHC alloantigen-specific B cell responses in vivo. We hypothesized that some anti-idiotypic Abs function as divalent molecular mimics of soluble HLA alloantigen. To test this idea, we studied two well-defined anti-idiotypic mAbs, T10-505 and T10-938, elicited in syngeneic BALB/c mice by immunization with CRll-351, an HLA-A2,24,28-specific mAb. Each anti-Id induced "Ab-3" Abs in rabbits that cross-reacted with HLA-A2 but not with HLA-B Ags. Furthermore, each anti-Id could bind to and block Ag recognition by Ha5C2.A2, a human homologue of mAb CRll-351. Both anti-Id mAb displayed weak reactivity with the human mAb SN66E3, which recognized an overlapping but distinct determinant of HLA-A2 Ags; neither reacted with human mAb MBW1, which recognized a nonoverlapping HLA-A2 determinant. Amino acid sequence comparison of mAb CRII-351 heavy and light chain variable region complementarity-determining regions (CDRs) with those of mAb Ha5C2.A2 and SN66E3 revealed short regions of homology with both human mAb; a large insert in the light chain CDR1 of mAb SN66E3 distinguished it from both CRll-351 and Ha5C2.A2. The amino acid sequences of mAb T10-505 and T10-938, which differed markedly from each other, revealed no homology to the alpha2 domain sequence of HLA-A*0201 that contains the CRll-351 mAb-defined epitope. We conclude that structurally different anti-Id Abs can mimic a polymorphic conformational epitope of an HLA Ag. In the case of T10-505 and T10-938 mimicry was not based on exact replication of the epitope by the hypervariable loops of the anti-Id mAb.     

Case report of unusual leukoencephalopathy preceding primary CNS lymphoma

A previously healthy 35 year old woman presented with bilateral uveitis associated with multiple, evolving, non-enhancing white matter lesions consistent with a progressive leukoencephalopathy such as multiple sclerosis. Thirty months after her initial presentation, she was diagnosed with primary CNS lymphoma and died 14 months later. The unusual clinical course preceding the diagnosis suggests that a demyelinating disease may have preceded, and possibly heralded, the development of primary CNS lymphoma. Cases of "sentinel lesions" heralding the diagnosis of primary CNS lymphoma have been reported, and this case further corroborates such instances and raises further issues regarding possible neoplastic transformation occurring in inflammatory diseases such as multiple sclerosis.     

A20 inhibits NF-kappaB activation in endothelial cells without sensitizing to tumor necrosis factor-mediated apoptosis

Expression of the NF-kappaB-dependent gene A20 in endothelial cells (EC) inhibits tumor necrosis factor (TNF)-mediated apoptosis in the presence of cycloheximide and acts upstream of IkappaBalpha degradation to block activation of NF-kappaB. Although inhibition of NF-kappaB by IkappaBalpha renders cells susceptible to TNF-induced apoptosis, we show that when A20 and IkappaBalpha are coexpressed, the effect of A20 predominates in that EC are rescued from TNF-mediated apoptosis. These findings place A20 in the category of "protective" genes that are induced in response to inflammatory stimuli to protect EC from unfettered activation and from undergoing apoptosis even when NF-kappaB is blocked. From a therapeutic perspective, genetic engineering of EC to express an NF-kappaB inhibitor such as A20 offers the mean of achieving an anti-inflammatory effect without sensitizing the cells to TNF-mediated apoptosis.