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In this study, the abilities of constitutive and conditional forms of the three Raf kinases to abrogate the cytokine dependency of FDC-P1 cells were examined. The constitutively active forms (delta) of all three Raf kinases were fused to the hormone-binding domain of the estrogen receptor (ER), rendering their activities conditionally dependent upon exogenous beta-estradiol. The vast majority of deltaRaf:ER-infected FDC-P1 cells remained cytokine-dependent; however, cells were obtained at low frequency in which expression of deltaRaf:ER abrogated cytokine dependency. Isoform specific differences between the Raf kinases were observed as cytokine-independent cells were obtained more frequently from deltaA-Raf:ER than either deltaRaf-1:ER or deltaB-Raf:ER infected cells. To determine whether the regulatory phosphorylation sites in the Raf proteins were necessary for abrogation of cytokine dependency, they were changed by site-directed mutagenesis. Substitution with phenylalanine eliminated the transforming ability of the deltaB-Raf:ER and deltaRaf-1:ER kinases. However, a similar substitution in A-Raf did not extinguish its transforming activity. The activated Raf proteins induced essential downstream MEK1 activity as treatment with the MEK1 inhibitor, PD98059, suppressed Raf-mediated growth. Activated MAP kinases (ERK1 and ERK2) were detected in deltaRaf:ER-transformed cells, and their presence was dependent upon a functional MEK1 protein. The cytokine-independent phenotype required the continued activity of the deltaRaf:ER proteins as removal of beta-estradiol caused the cells to stop growing and undergo apoptosis. The Raf-responsive cells were found to express autocrine growth factors, which promoted their growth. Constitutive activation of the Raf-1 oncogene resulted in malignant transformation as cytokine-independent FDC-P1 cells infected with a retrovirus encoding an activated Raf-1 protein formed tumors upon injection of immunocompromised mice. In summary, Raf kinases can abrogate cytokine dependency, prevent apoptosis and induce the tumorigenicity of a certain subpopulation of FDC-P1 cells by a MEK1-dependent mechanism.

A preferential solvation model that relates solute properties with solvent composition in binary mixtures has been applied to the dissociation pKa values of a set of 28 substituted phenols in methanol-water mixtures. The parameters of the model allow estimation of the pKa value of each phenol for any methanol-water composition. Moreover, it is demonstrated that the pKa values of the whole set of phenols at any methanol-water composition are linearly related to the pKa values of the phenols in water. Equations that relate the correlations' slope and intercept values with the solvent composition have been derived and tested with the set of phenols. The general parameters obtained for these equations allow an accurate calculation of the pKa value of any phenol, even of those not included in the original set, at any methanol-water composition solely from the pKa value of the phenol in water. These calculated pKa values can be used for quantitative structure-HPLC retention relationships. The method is tested by comparison of the calculated pKa values with the HPLC determined pKa values of 26 phenols in a polymeric column with a 50% methanol as mobile phase.

The 13 short tandem repeat (STR) loci D3S1358, vWA, FGA, D16S539, TH01, TPOX, CSF1PO, D8S1179, D21S11, D18S51, D5S818, D13S317 and D7S820 as well as the amelogenin locus, contained in AmpFlSTR Profiler Plus and/or AmpFlSTR Cofiler and/or AmpFlSTR Green I PCR amplification kits, were studied in four populations from the Iberian Peninsula, Basques, Catalans, Andalusians and Portuguese and two North African populations (Moroccan Arabs and Berbers). The aim of the study was to obtain accurate allele frequency data and other genetic parameters of forensic interest on the main representative human groups living in Iberia and Morocco using an automated method and commercial amplification kits.

The eight short tandem repeat (STR) polymorphic systems mapping on the male-specific region of the human Y chromosome, DYS19, DYS388, DYS389I, DYS389II, DYS390, DYS391, DYS392 and DYS393, were typed in four populations from northwest (NW) Africa (Moroccan Arabs, southern Moroccan Berbers, Saharawis and Mozabites). Allele frequency distributions showed statistically significant differences for all loci among all the populations except for DYS19. Complete typing was obtained for 185 chromosomes, which showed 74 different haplotypes. The two most frequent haplotypes were found in 16.2% and 15.1% of the individuals, although the latter was almost exclusively found in the Mozabites. Locus and haplotype informativeness were measured by means of the gene diversity (D). The haplotype diversity ranged from 0.856 (Mozabites) to 0.967 (southern Moroccan Berbers). For some loci, allele frequencies in NW Africans were clearly different from those in Europeans. The most common NW African haplotype was found only in one individual out of a total of 494 Europeans typed for the whole STR set. Thus, NW African and European Y chromosomes are clearly differentiated.

Eleven biallelic polymorphisms and seven short-tandem-repeat (STR) loci mapping on the nonrecombining portion of the human Y chromosome have been typed in men from northwestern Africa. Analysis of the biallelic markers, which represent probable unique events in human evolution, allowed us to characterize the stable backgrounds or haplogroups of Y chromosomes that prevail in this geographic region. Variation in the more rapidly mutating genetic markers (STRs) has been used both to estimate the time to the most recent common ancestor for STR variability within these stable backgrounds and to explore whether STR differentiation among haplogroups still retains information about their phylogeny. When analysis of molecular variance was used to study the apportionment of STR variation among both genetic backgrounds (i.e., those defined by haplogroups) and population backgrounds, we found STR variability to be clearly structured by haplogroups. More than 80% of the genetic variance was found among haplogroups, whereas only 3.72% of the genetic variation could be attributed to differences among populations-that is, genetic variability appears to be much more structured by lineage than by population. This was confirmed when two population samples from the Iberian Peninsula were added to the analysis. The deep structure of the genetic variation in old genealogical units (haplogroups) challenges a population-based perspective in the comprehension of human genome diversity. A population may be better understood as an association of lineages from a deep and population-independent gene genealogy, rather than as a complete evolutionary unit.

We have examined the worldwide distribution of a Y-chromosomal base-substitution polymorphism, the T/C transition at SRY-2627, where the T allele defines haplogroup 22; sequencing of primate homologues shows that the ancestral state cannot be determined unambiguously but is probably the C allele. Of 1,191 human Y chromosomes analyzed, 33 belong to haplogroup 22. Twenty-nine come from Iberia, and the highest frequencies are in Basques (11%; n=117) and Catalans (22%; n=32). Microsatellite and minisatellite (MSY1) diversity analysis shows that non-Iberian haplogroup-22 chromosomes are not significantly different from Iberian ones. The simplest interpretation of these data is that haplogroup 22 arose in Iberia and that non-Iberian cases reflect Iberian emigrants. Several different methods were used to date the origin of the polymorphism: microsatellite data gave ages of 1,650, 2,700, 3,100, or 3,450 years, and MSY1 gave ages of 1,000, 2,300, or 2,650 years, although 95% confidence intervals on all of these figures are wide. The age of the split between Basque and Catalan haplogroup-22 chromosomes was calculated as only 20% of the age of the lineage as a whole. This study thus provides evidence for direct or indirect gene flow over the substantial linguistic barrier between the Indo-European and non-Indo-European-speaking populations of the Catalans and the Basques, during the past few thousand years.

Eight Y-linked short-tandem-repeat polymorphisms (DYS19, DYS388, DYS389I, DYS389II, DYS390, DYS391, DYS392, and DYS393) were analyzed in four populations of Central Asia, comprising two lowland samples-Uighurs and lowland Kirghiz-and two highland samples-namely, the Kazakhs (altitude 2,500 m above sea level) and highland Kirghiz (altitude 3,200 m above sea level). The results were compared with mtDNA sequence data on the same individuals, to study possible differences in male versus female genetic-variation patterns in these Central Asian populations. Analysis of molecular variance (AMOVA) showed a very high degree of genetic differentiation among the populations tested, in discordance with the results obtained with mtDNA sequences, which showed high homogeneity. Moreover, a dramatic reduction of the haplotype genetic diversity was observed in the villages at high altitude, especially in the highland Kirghiz, when compared with the villages at low altitude, which suggests a male founder effect in the settlement of high-altitude lands. Nonetheless, mtDNA genetic diversity in these highland populations is equivalent to that in the lowland populations. The present results suggest a very different migration pattern in males versus females, in an extended historical frame, with a higher migration rate for females.

The solvation parameter model has been applied to the characterization of micellar electrokinetic chromatographic (MEKC) systems with mixtures of sodium dodecyl sulfate and Brij 35 as surfactant. The variation in MEKC surfactant composition results in changes in the coefficients of the correlation equation, which in turns leads to information on solute-solvent and solute-micelle interactions. Since the same solvation model can be used to describe many biological processes, particular MEKC surfactant compositions can be selected that model the solute-solvent interactions of some of these processes. Two different MEKC systems have been selected to model the solute-solvent interactions of two processes of biological interest (octanol-water partition and tadpole narcosis).

Central Asia is a vast region at the crossroads of different habitats, cultures, and trade routes. Little is known about the genetics and the history of the population of this region. We present the analysis of mtDNA control-region sequences in samples of the Kazakh, the Uighurs, the lowland Kirghiz, and the highland Kirghiz, which we have used to address both the population history of the region and the possible selective pressures that high altitude has on mtDNA genes. Central Asian mtDNA sequences present features intermediate between European and eastern Asian sequences, in several parameters-such as the frequencies of certain nucleotides, the levels of nucleotide diversity, mean pairwise differences, and genetic distances. Several hypotheses could explain the intermediate position of central Asia between Europe and eastern Asia, but the most plausible would involve extensive levels of admixture between Europeans and eastern Asians in central Asia, possibly enhanced during the Silk Road trade and clearly after the eastern and western Eurasian human groups had diverged. Lowland and highland Kirghiz mtDNA sequences are very similar, and the analysis of molecular variance has revealed that the fraction of mitochondrial genetic variance due to altitude is not significantly different from zero. Thus, it seems unlikely that altitude has exerted a major selective pressure on mitochondrial genes in central Asian populations.

Raf is a key serine-threonine protein kinase which participates in the transmission of growth, anti-apoptotic and differentiation messages. These signals can be initiated after receptor ligation and are transmitted to members of the MAP kinase cascade that subsequently activate transcription factors controlling gene expression. Raf is a member of a multigene family which includes: Raf-1, A-Raf and B-Raf. The roles that individual Raf kinases play in the regulation of normal and malignant hematopoietic cell growth are not clear. The following studies show that all three Raf kinases are functionally present in certain human hematopoietic cells, and their aberrant expression can result in abrogation of cytokine dependency. Cytokine-dependent TF-1 cells were infected with retroviruses encoding amino-terminal deleted (delta) A-Raf, B-Raf and Raf-1 proteins. These Raf proteins were conditionally inducible as they were fused to the hormone-binding domain of the estrogen receptor (ER). A hierarchy in the abilities of Raf-containing retroviruses to abrogate cytokine dependency was observed as deltaA-Raf:ER was 20- to 200-fold more efficient than either deltaRaf-1:ER or deltaB-Raf:ER, respectively. This result was unexpected as A-Raf is an intrinsically weaker kinase than either Raf-1 or B-Raf. The activated Raf proteins induced downstream MEK and MAP (ERK1 and ERK2) kinase activities in the cells which proliferated in response to Raf activation. Furthermore, a functional MEK signaling pathway was necessary as treatment of the cells with a MEK1-inhibitor suppressed Raf-mediated proliferation. To determine whether the regulatory phosphorylation residues contained in the modified Raf oncoproteins were necessary for transformation, they were altered by site-directed mutagenesis. Substitution of the regulatory phosphorylation tyrosine residues with phenylalanine in either A-Raf or Raf-1 reduced the capacity of these oncoproteins to abrogate cytokine dependency. In contrast, changing the critical aspartic acid residues of B-Raf to either tyrosine or phenylalanine increased the frequency of estradiol-responsive cells. Thus, the amino acids present in the regulatory residues modulated the capability of Raf proteins to abrogate the cytokine dependency of TF-1 cells. Differences in the levels of Raf and downstream kinase activities were observed between cytokine-dependent and estradiol-responsive deltaRaf:ER-infected cells as estradiol-responsive cells usually expressed more Raf and MEK activity than GM-CSF-dependent, deltaRaf:ER-infected cells. Abrogation of cytokine dependency by the activated deltaRaf:ER proteins was associated with autocrine growth factor synthesis which was sufficient to promote the growth of uninfected TF-1 cells. In summary, these observations indicate that the aberrant expression of certain activated deltaRaf:ER oncoproteins can alter the cytokine dependency of human hematopoietic TF-1 cells. These cells will be useful in evaluating the roles of the individual Raf oncoproteins in signal transduction, cell cycle progression, autocrine transformation, regulation of apoptosis and differentiation. Moreover, these Raf-infected cells may be important in evaluating the efficacy of novel anticancer drugs designed to inhibit Raf and downstream signal transduction molecules.

Osteoporosis, a common metabolic condition resulting in reduced bone mass, causes significant morbidity in affected individuals by predisposing them to osteoporotic fractures. To determine the relationship of scoliosis and osteoporosis, dual-energy X-ray absorptiometry (DXA) scans of 493 men and 762 women were analyzed according to their scoliotic status. No association was observed in the few scoliotic men without osteoarthritis of the spine compared to nonscoliotic men. Without osteoarthritis, scoliotic women had significantly decreased bone mass of the total hip and the femoral neck (p < 0.05) compared to nonscoliotic women, whereas in spine, bone mass was also decreased compared to nonscoliotic women, but not statistically significant. This suggests that scoliosis associates with bone loss, and affected women may benefit from early institution of therapeutic measures.

Crystalline electron donor-acceptor (EDA) complexes of various diarylacetylenes (DA) and dichlorobenzoquinone (DB) are isolated and structurally characterized by X-ray crystallography. Deliberate excitation of either the DB acceptor at lambda(DB) = 355 nm or the 1:2 [DA, 2DB] complex at lambda(CT) = 532 nm in the solid state leads to [2 + 2] cycloaddition and identical (isomeric) mixtures of the quinone methide products. Time-resolved (ps) diffuse reflectance spectroscopy identifies the ion-radical pair [DA(*+), DB(*-)] as the reactive intermediate derived by photoinduced electron transfer in both photochemical procedures. The effects of crystal-lattice control on the subsequent ion-radical pair dynamics are discussed in comparison with the same photocouplings of acetylenes and quinone previously carried out in solution.

Seven HLA class I and class II loci (HLA-A, B, C, DRB1, DQA1, DPA1 and DPB1) were typed at the DNA level in two populations of the Iberian Peninsula (100 Basque and 88 Catalan individuals) in order to unravel their genetic relationship and to compare these results with other European and Mediterranean populations. For the first time, the frequencies of alleles and haplotypes for the class I HLA loci at the DNA level in these populations are presented. The most frequent haplotype in both populations is A*29-Cw*1601-B*44-DRB1*0701-DQA1*0201-DPA1*0103-DPB 1*0401. Neither population differed markedly from the highly homogeneous European and Mediterranean genetic landscape. The Basques, a European outlier population according to classical genetic markers, appear to lie within the genetic European variation with a slight uniqueness and show no clear relationship to North African populations, as has been postulated in some previous HLA studies. Here, the range of possibilities provided by the highly polymorphic HLA system is stressed by using genetic distances, phylogenetic trees and principal component analyses in order to reconstruct population history.

The shoulder is a remarkable structure enjoying a greater range of motion than any other articulation in the human body. The shoulder is not a single joint but actually four separate articulations that act synergistically. The authors review the complex anatomy of the shoulder, presenting correlation between anatomic sections and MR arthrography.

The Raf family of protein kinases display differences in their abilities to promote the entry of quiescent NIH 3T3 cells into the S phase of the cell cycle. Although conditional activation of deltaA-Raf:ER promoted cell cycle progression, activation of deltaRaf-1:ER and deltaB-Raf:ER elicited a G1 arrest that was not overcome by exogenously added growth factors. Activation of all three deltaRaf:ER kinases led to elevated expression of cyclin D1 and cyclin E and reduced expression of p27Kip1. However, activation of deltaB-Raf:ER and deltaRaf-1:ER induced the expression of p21Cip1, whereas activation of deltaA-Raf:ER did not. A catalytically potentiated form of deltaA-Raf:ER, generated by point mutation, strongly induced p21Cip1 expression and elicited cell cycle arrest similarly to deltaB-Raf:ER and deltaRaf-1:ER. These data suggested that the strength and duration of signaling by Raf kinases might influence the biological outcome of activation of this pathway. By titration of deltaB-Raf:ER activity we demonstrated that low levels of Raf activity led to activation of cyclin D1-cdk4 and cyclin E-cdk2 complexes and to cell cycle progression whereas higher Raf activity elicited cell cycle arrest correlating with p21Cip1 induction and inhibition of cyclin-cdk activity. Using green fluorescent protein-tagged forms of deltaRaf-1:ER in primary mouse embryo fibroblasts (MEFs) we demonstrated that p21Cip1 was induced by Raf in a p53-independent manner, leading to cell cycle arrest. By contrast, activation of Raf in p21Cip1(-/-) MEFs led to a robust mitogenic response that was similar to that observed in response to platelet-derived growth factor. These data indicate that, depending on the level of kinase activity, Raf can elicit either cell cycle progression or cell cycle arrest in mouse fibroblasts. The ability of Raf to elicit cell cycle arrest is strongly associated with its ability to induce the expression of the cyclin-dependent kinase inhibitor p21Cip1 in a manner that bears analogy to alpha-factor arrest in Saccharomyces cerevisiae. These data are consistent with a role for Raf kinases in both proliferation and differentiation of mammalian cells.

Eight human short tandem repeat polymorphisms (STRs) also known as microsatellites-DYS19, DYS388, DYS390, DYS391, DYS392, DYS393, DYS389I, and DYS389II, mapping in the Y chromosome-were analyzed in two Iberian samples (Basques and Catalans). Allele frequency distributions showed significant differences only for DYS392. Fst and gene diversity index (D) were estimated for the Y STRs. The values obtained are comparable to those of autosomal STR if corrections for the smaller effective population size on the Y chromosome are taken into account. This suggests that Y-chromosome microsatellites might be as useful as their autosomal counterparts to both human population genetics and forensics. Our results also reinforce the hypothesis that selective sweeps in the Y chromosome in recent times are unlikely. Haplotypes combining five of the loci were constructed for 71 individuals, showing 29 different haplotypes. A haplotype tree was constructed, from which an estimate of 7,000 to 60,000 years for the age of the Y-chromosome variation in Iberia was derived, in accordance with previous estimates obtained with mtDNA sequences and nuclear markers.

The catalytic domains of the Raf family of protein kinases (deltaRaf) differ in their ability to activate MEK in vitro and in vivo and in their ability to oncogenically transform mammalian cells. The kinase domain of B-Raf is more active than the equivalent portion of Raf-1 which in turn is more active than A-Raf. In Raf-1 the phosphorylation or mutation to aspartic acid of two key tyrosine residues upstream of the ATP binding site has been demonstrated to significantly potentiate catalytic activity. In A-Raf the analogous amino acids are also tyrosine whereas in B-Raf they are aspartic acid. To determine if these differences in amino acid sequence influence the relative catalytic activity of the Raf kinase domains we constructed forms of deltaA-Raf, deltaB-Raf and deltaRaf-1 that encode either aspartic acid [DD], phenylalanine [FF] or tyrosine [YY] at these positions. These proteins were expressed both in mammalian cells as fusions with the hormone binding domain of the estrogen receptor and as epitope-tagged proteins in Sf9 insect cells to test their oncogenic and catalytic potentials. When expressed in Rat1 or 3T3 cells in the presence of hormone all of the deltaRaf-1:ER and deltaA-Raf:ER proteins were transforming with the exception of the [FF] form of deltaA-Raf. In general the [DD] forms of the deltaRaf-1:ER and deltaA-Raf:ER proteins were the most potently oncogenic which correlated with their ability to elicit activation of the MAP kinase pathway. Consistent with the transformation data, the catalytic activity of the [DD] forms of deltaA-Raf:ER and deltaRaf-1:ER was about ten times greater than the cognate [FF] and [YY] forms of the proteins. By contrast all of the deltaB-Raf:ER proteins were highly transforming and deltaB-Raf catalytic activity was largely unaffected by mutation of the aforementioned aspartic acids to either tyrosine or phenylalanine. Similar results were obtained with epitope-tagged forms of deltaA-Raf, deltaB-Raf and deltaRaf-1 expressed in Sf9 cells. These data provide support for the model that key tyrosine residues in the protein kinase domains of A-Raf and Raf-1 are important in the regulation of catalytic activity. In addition they demonstrate that the higher intrinsic activity of B-Raf cannot be explained simply by the presence of aspartic acids at the analogous positions.

20 microsatellite polymorphisms: HUMHPRT, HUMD3S1358, HUMTH01, HUMACPP, HUMVWF, HUMD16S310, HUMD4S243, HUMTPO, HUMFES/FPS, HUMF13A1, HUMDHFRP2, HUMD11S2010, HUMD13S767, HUMD9S926, HUMD2S1328, HUMD14S306, HUMD18S848, HUMD5S818, HUMD7S820 and HUMFGA were analyzed in a worldwide survey covering five continents and allele frequencies are given. There is a high heterogeneity in allele frequencies among continents. A neighbor-joining tree based on Fst distance shows a pattern of differentiation that may reflect the role of drift in the development of genetic differences among humans. The variation found between continents confirms the usefulness of tetranucleotide microsatellites in human genetic variation studies.

After an intensive bibliographic search, we compiled all the available data on allele frequencies for classical genetic polymorphisms referring to North African populations and synthesized the data in an attempt to reconstruct the populations' demographic history using two complementary methods: (1) principal components analysis and (2) genetic distances represented by neighbor-joining trees. In both analyses the main feature of the genetic landscape in northern Africa is an east-west pattern of variation pointing to the differentiation between the Berber and Arab population groups of the northwest and the populations of Libya and Egypt. Moreover, Libya and Egypt show the smallest genetic distances with the European populations, including the Iberian Peninsula. The most plausible interpretation of these results is that, although demic diffusion during the Neolithic could explain the genetic similarity between northeast Africa and Europe by a parallel process of gene flow from the Near East, a Mesolithic (or older) differentiation of the populations in the northwestern regions with later limited gene flow is needed to understand the genetic picture. The most isolated groups (Mauritanians, Tuaregs, and south Algerian Berbers) were the most differentiated and, although no clear structure can be discerned among the different Arab- and Berber-speaking groups, Arab speakers as a whole are closer to Egyptians and Libyans. By contrast, the genetic contribution of sub-Saharan Africa appears to be small.

Sequences from the mitochondrial DNA (mtDNA) control region were analyzed in nine European and West Asian populations. They showed low genetic heterogeneity when compared to world populations. However, a Caucasoid population tree displayed a robust east-west gradient. Within-population diversity (ascertained through various parameters) and mean pairwise differences declined from east to west, in a pattern compatible with ancient population migration and expansion from the Middle East. Estimated expansion times indicate a Paleolithic event with important differences among populations according to their geographical position and thus a slower tempo than previously believed. The replacement of Neanderthals by anatomically modern humans, fully compatible with the present results, may have been a slower and more complex process than cultural change suggests.

The electrogram storage is a feature offered by the last generation of defibrillators, which enables the physician to establish whether the sensed episode and the electrical therapy delivered by the device was appropriate. The incidence of electromagnetic interferences may be higher than initially suspected with the latest generation of defibrillators. Such interference was documented in 4 out of 100 patients over a 24-month period. The electrical interference occurred in all four of them, while they were playing with slot machines. In the RR interval history, irregular QRS sensing of variable frequency was documented in some episodes. The intervals were irregular and nonphysiological satisfying the rate cut-off criteria in an intermittent and nonsustained way. There may be high risk of electromagnetic interference between slot machines and defibrillators. Our experience calls for a warning to all defibrillator patients and, perhaps some regulatory intervention.

A new model that relates the retention of a weak acid in HPLC columns with the pH and ionic strength of the mobile phase is derived and tested for different benzoic acids in methanol-water mobile phases. The proposed model uses the pH value in the mobile phase instead of the pH value in water, takes into account the effect of the activity coefficients, and considers different holdup times for the neutral and ionic species. The dependence of the holdup time of the ionic species on the mobile phase properties (pH, solvent composition, and ionic strength) is evaluated. It is demonstrated that the holdup time of the neutral species does not depend on the mobile phase properties, but the holdup time of the ionic species depends on the particular buffer used. The proposed equations can be combined with previously derived equations that relate the retention with the solvent composition of the mobile phase to establish a general model that relates the retention of the solute with the significant mobile phase properties:  composition, pH, and ionic strength.

Purpose: Our goal was to investigate the safety, effectiveness, and feasibility for the practicing physician of stepwise and low-dose administration of FSH in WHO group II anovulatory infertile women., Methods: Infertile female patients (n = 234) suffering from WHO group II anovulation, and who failed to became pregnant with clomiphene citrate, were included in a multicenter, prospective, clinical study of treatment with a protocol of chronic low-dose and small incremental rises with urinary purified or highly purified FSH. Follicular development was monitored with ultrasonographic scans., Results: The 234 patients received a total of 534 cycles of treatment, for a mean number of 2.3 treated cycles per patient. hCG was withheld in 65 (12.2%) cyles because of no response and in 28 (5.2%) cycles because of hyperresponse. Of the remaining 441 cycles, 419 (95%) were ovulatory, and in 198 (47.3%) of these cycles a single dominant follicle developed. There were 93 pregnancies (39.7% per patient), for a cycle fecundity rate of 17.4%. Cumulative conception rate after two treated cycles was 33.5%. There were 14 (15%) pairs of twins and 10 (10.8%) spontaneous miscarriages. The prevalence of complications was low with no cases of severe OHSS. Basal LH/FSH ratio was significantly higher in the pregnant group of patients than in nonpregnant women., Conclusions: Stepwise and chronic low-dose administration of FSH is a safe and effective method for treatment of WHO group II anovulatory infertility, mainly in those patients having high LH/FSH ratios.

The extent to which the diagnostic performance of low-dosed storage phosphors can be improved by suitable image postprocessing was investigated. Storage phosphor radiographs and conventional 200- and 400-speed screen-film combinations of an anthropomorphic chest phantom with simulated nodules and linear structures were compared. The detection of test bodies was determined in a ROC analysis based on 16200 individual observations. The two analog screen-film systems did not differ significantly. As far as the storage phosphor radiographs are concerned, a higher diagnostic performance was achieved with a large filter kernel (S 40) whereas small filter kernels and edge-enhanced imaging modes reduced the detection of detail. The image quality of low-dosed storage phosphors is thus similar to high-amplification screen-film combinations if large filter kernels are used in postprocessing of the image.

The protein kinase domains of mouse A-Raf and B-Raf were expressed as fusion proteins with the hormone binding domain of the human estrogen receptor in mammalian cells. In the absence of estradiol, 3T3 and rat1a cells expressing delta A-Raf:ER and delta B-Raf:ER were nontransformed, but upon the addition of estradiol the cells became oncogenically transformed. Morphological oncogenic transformation was more rapid and distinctive in cells expressing delta B-Raf:ER compared with cells expressing delta A-Raf:ER. Biochemical analysis of cells transformed by delta A-Raf:ER and delta B-Raf:ER revealed several interesting differences. The activation of delta B-Raf:ER consistently led to the rapid and robust activation of both MEK and p42/p44 MAP kinases. By contrast, the activation of delta A-Raf:ER led to a weak activation of MEK and the p42/p44 MAP kinases. The extent of activation of MEK in cells correlated with the ability of the different Raf kinases to phosphorylate and activate MEK1 in vitro. delta B-Raf:ER phosphorylated MEK1 approximately 10 times more efficiently than delta Raf-1:ER and at least 500 times more efficiently than delta A-Raf:ER under the conditions of the immune-complex kinase assays. These results were confirmed with epitope-tagged versions of the Raf kinase domains expressed in insect cells. The activation of all three delta Raf:ER proteins in 3T3 cells led to the hyperphosphorylation of the resident p74raf-1 and mSOS1 proteins, suggesting the possibility of "cross-talk" between the different Raf kinases and feedback regulation of intracellular signaling pathways. The activation of either delta B-Raf:ER or delta Raf-1:ER in quiescent 3T3 cells was insufficient to promote the entry of the cells into DNA synthesis. By contrast, the activation of delta A-Raf:ER in quiescent 3T3 cells was sufficient to promote the entry of the cells into S phase after prolonged exposure to beta-estradiol. The delta Raf:ER system has allowed us to reveal significant differences between the biological and biochemical properties of oncogenic forms of the Raf family of protein kinases. We anticipate that cells expressing these proteins and other estradiol-regulated protein kinases will be useful tools in future attempts to unravel the complex web of interactions involved in intracellular signal transduction pathways.

Study Design: Controlled comparison of radiographic interpretive performance based on training and experience., Objectives: This study compared each of these groups in medicine and chiropractic by testing abilities to interpret abnormal plain film radiographs of the lumbosacral spine and pelvis., Summary of Background Data: Low back pain is a common and costly problem that is evaluated and treated primarily by medical physicians, orthopedists, and chiropractors. Although radiology is used extensively in patients with low back pain, the radiographic interpretations of students, clinicians, radiology residents, and radiologists have never been compared., Methods: Four hundred ninety-six eligible volunteers from nine target groups completed a test of radiographic interpretation consisting of nineteen cases with clinically important radiographic findings. The nine groups included 22 medical students, 183 chiropractic students, 27 medical radiology residents, 13 chiropractic radiology residents, 66 medical clinicians (including 12 general practice physicians, 25 orthopedic surgeons, 21 orthopedic residents, and 8 rheumatologists), 46 chiropractic clinicians, 48 general medical radiologists, 55 chiropractic radiologists, and 36 skeletal radiologists and fellows., Results: The test established a high level of internal consistency reliability (0.880) and revealed that, in the interpretation of abnormal plain film radiographs of the lumbosacral spine and pelvis, significant differences were found among professional groups (P < 0.0001). Post hoc tests (P < 0.05) revealed that skeletal radiologists achieved significantly higher test results than did all other medical groups; that the test results of general medical radiologists and medical radiology residents was significantly higher than those of medical clinicians; that test results of medical students was significantly poorer than that of all other medical groups; that the performance of chiropractic radiologists and chiropractic radiology residents was significantly higher than that of chiropractic clinicians and chiropractic students; that no significant differences was revealed in the mean values of performance of chiropractic clinicians and chiropractic students; that the test results of chiropractic radiologists, chiropractic radiology residents, and chiropractic students was significantly higher than that of the corresponding medical categories (general medical radiologists, medical radiology residents, and medical students, respectively); that no significant difference in test results was identified between chiropractic radiologists and skeletal radiologists or between chiropractic and medical clinicians; and that the length of time in practice for clinicians and radiologists was not a significant factor in the test results., Conclusions: These data demonstrate a substantial increase in test results of all radiologists and radiology residents when compared to students and clinicians in both medicine and chiropractic related to the interpretation of abnormal radiographs of the lumbosacral spine and pelvis. Furthermore, the study reinforces the need for radiologic specialists to reduce missed diagnoses, misdiagnoses, and medicolegal complications.

This report describes the natural history of the Segond fracture and documents the radiographic appearance of the healed Segond fracture. The clinical and radiographic records of 129 patients with acute anterior cruciate ligament (ACL) injuries were reviewed. Four (3.1%) of these patients had Segond fractures. On follow-up radiographic examination, seven patients demonstrated a characteristic bone excrescence arising 3-6 mm inferior to the lateral tibial plateau. In four of the five patients for whom acute injury films were available this excrescence arose at the site of the earlier Segond fracture. Healing of such fractures is associated with a characteristic bone excrescence distinct from an osteophyte. This excrescence implies significant internal derangement of the knee.

Rationale and Objectives: The frequency and morphology of central acetabular osteophyte formation were evaluated in patients with osteoarthritis., Methods: One hundred preserved acetabular specimens were evaluated for the occurrence of central osteophytes (excrescences present in the articulation of the hip joint), their relationship to marginal and femoral osteophytes of the hip, and their radiographic appearance., Results: In both specimen and patient analysis, the authors found that central excrescences occur commonly and appear to be related temporally to osteophytes of the femur and the margin of the acetabulum., Conclusions: The authors demonstrated a close relationship between the degree of marginal and femoral osteophytes with those found centrally, indicating a temporal relationship. The new radiographic signs discussed are: 1) a fine linear density associated with early osteoarthritis; 2) larger, more irregular densities associated with a more advanced stage of osteoarthritis; and 3) obliteration of the acetabular fossa by excrescences as found in severe osteoarthritis.

The shoulder joint is a remarkable structure enjoying a greater range of motion than any other articulation in the human body. The shoulder is not a single joint but actually four separate articulations that act synergistically. The authors review the complex anatomy of the shoulder.

The dissociation equilibria of di(2,4,4-trimethylpentyi) phosphinic acid, mono(2,4,4-trimethylpentyl) phosphinic acid, di(n-octyl)phosphinic acid and mono(n-octyl)phosphinic acid have been studied in ethanol-water mixtures by potentiometric titration at 25 degrees C. These data have been analysed both graphically numerically using the program LETAGROP-ZETA. The obtained pK(a) values have been correlated with the corresponding values in water, determined both indirectly by means of extraction measurements and by estimation using the suitable Hammett equation.

We studied phagocytosis of rod outer segments (ROS) by the retinal pigmented epithelium (RPE) using rapid freezing, freeze-drying, and electron microscopic immunocytochemistry. Phagocytosis of photoreceptor outer segment tips by the RPE occurs daily, and in rats the shedding of these tips is light-entrained to a circadian rhythm. We studied the phagocytic process 5, 30, 90, and 150 min after light onset or after subjective light onset in rats entrained to a 12-hr dark-12-hr light cycle. Lysosomes were labeled with antibodies to cathepsin D, a major lysosomal enzyme responsible for opsin degradation. Phagosomes and phagolysosomes were recognized because of the lamellar structure of their photoreceptor-derived contents. We found a population of lysosomes that fuse with one another before they interact with phagosomes. This fusion can be triggered either by light or by endogenous circadian mechanisms. We also found that lysosome-phagosome interaction occurs after the ingestion stage is completed and that this interaction occurs in two steps. First, smaller lysosomes fuse with phagosomes. Subsequently, larger lysosomes appear to interact with phagosomes via pore-like or bridge-like structures. It is proposed that interchange of contents takes place through these structures.

In brief MRI can prove diagnostically brief useful for a number of osseous injuries that active patients may incur. The authors weigh the benefits of MRI, such as early injury detection and excellent contrast and spatial resolution, versus its shortcomings, such as high cost and sensitivity to motion. In addition, they describe in detail the MRI appearances of stress fractures, bone bruises, occult fractures, and osteochondral fractures.

To compare the efficacy of intravenous diltiazem vs nitroglycerin in patients with unstable angina, we conducted a prospective, single blinded study enrolling 45 patients, 24 of them received diltiazem and 21 nitroglycerin. We considered drug failure if the patient experienced two or more anginal episodes or if one of them was longer than 30 minutes. There was a good response in 19 of 24 patients (79%) of the diltiazem group and 17 of 21 (81%) in the nitroglycerin group. Only one of the 5 non-response patients in the diltiazem group that was crossed to nitroglycerin was controlled, in opposition to the 4 patients crossed to diltiazem that were controlled all of them (p less than 0.05). Both drugs significantly (p less than 0.01) reduced systolic and diastolic blood pressure, but there was not difference between the two groups. Heart rate and double product decreased only in the diltiazem group (p less than 0.05). We did not find notorious side effects. We concluded that diltiazem is an alternative to nitroglycerin in patients with unstable angina.

170 patients who had suffered trauma to bones or soft tissues were examined by a digital and a conventional film/screen technique. In addition to "standard" and edge-enhanced images, post-processing was carried out and compared with conventional radiographs. Four radiologists and two surgeons evaluated the compacta, spongiosa, soft tissues, fractures and osteo-synthetic materials as shown on the two systems. The "standard" digital images are inferior to film images, but edge-enhanced images showed definite advantages for demonstrating soft tissues. Evaluation of osteo-synthetic material is limited by the creation of artifacts. Recognition of detail can be significantly improved by using post-processing with optimized parameters.

Lateral-to-motor and medial-to-motor synapses in crayfish nerve cords are composed of an electrical and a chemical component. The presynaptic terminals showed localized clusters of synaptic vesicles, electron-dense areas, coated pits, and coated vesicles. In thin sections, active zones were defined by electron-dense regions where synaptic vesicles attached and, in freeze-fracture replicas, by clusters of intramembrane particles localized in bands with vesicle openings on the sides of these bands. The cytoplasmic surface of the postsynaptic membrane opposite the active zones was coated with electron-dense material that in freeze-fracture replicas was seen as an increase in intramembrane particles located in the external leaflet (EF-face). This specialization of the postsynaptic membrane may correspond to the neurochemical receptor. Also, pre- and postsynaptic membranes were separated by a wider extracellular gap than those of adjacent nonsynaptic regions and electrical synapses or gap junctions. Synaptic vesicles were located exclusively at the synaptic regions by means of a cytoskeleton that was different for the electrical and the chemical components. The vesicles associated with the electrical component were anchored to a cytoskeleton composed of a beaded layer of densities located parallel to the membrane. This cytoskeleton maintained the synaptic vesicles separated from the presynaptic membrane by a distance of 13 +/- 2 nm. The synaptic vesicles associated with the chemical component were anchored to electron-dense regions formed by filaments arranged in bundles, anchored to the presynaptic membrane. Vesicles lined both sides positioned to discharge their contents into the extracellular space and to replace the discharged vesicles.

The dissociation constants of several families of acids (substituted phenols and mercaptopyrimidines) in isopropyl alcohol medium have been determined by potentiometric titration with tetrabutylammonium hydroxide. Because of ion-pair formation the incomplete dissociation of the tetrabutylammonium salt has been taken into account in the calculation of pK(a). The dissociation constants of the salts were previously measured conductometrically. The resolution of acid strength in isopropyl alcohol relative to that in water has been determined for each series of acids by plotting the pK(a) values in isopropyl alcohol vs. those in water. The results show greater resolution in isopropyl alcohol than in water. The resolution of acid strength in tert-butyl alcohol relative to that in isopropyl alcohol has also been determined.

Titration curves have been simulated for the titration of acids, bases and salts in neutral amphiprotic solvents such as isopropyl and tert-butyl alcohols. Ranges of pK values and acid concentrations have been examined. The incomplete dissociation of salts, which increases the acid or basic strength, has been found to be the major factor modifying the shape of the curve and the pH break. The theoretical predictions have been checked by titrating several series of acids and good agreement has been obtained between computed and experimental results.

Alizarin, Alizarin S, quinizarin and quinalizarin have been compared as acid-base indicators with Bromocresol Green and Methyl Orange as reference indicators. The chromaticity co-ordinates, complementary chromaticity co-ordinates, pK(a) values, transition pH-range, pH of maximum colour change, optimum concentration for titrations and the quality of colour change were determined. The results show all four to be good indicators with a colour change quality similar to that of Bromocresol Green.

The ionic equilibria in neutral amphiprotic solvents (isopropyl and tert-butyl alcohols) have been established, and equations to calculate pH values in solutions of acids, bases, salts or their mixtures, developed. The effect, on the dissociation equilibria, of the presence of small quantities of water or other solvents in the bulk solvent used has been taken into account in the proposed equations. On the basis of these equations some buffer solutions have been studied and recommended for electrode standardization. The results, tested by experimental work, show the importance of the incompleteness of dissociation of salts in these solvents, which decreases the pH of acid buffers and increases the buffer capacity.

Dissociation constants (pK(a1) and pK(a2) in water-ethanol medium for 3-styryl-2-mercaptopropenoic and 3-(1-naphthyl)-2-mercaptopropenoic acid have been determined potentiometrically, and pK(a2) for both in aqueous medium, spectrophotometrically. Neutralization enthalpies in water-ethanol medium have been determined by thermometric titration. The reactions with metal ions have been studied, and the main reactions are described. The most sensitive reactions are with titanium(IV) (pD = 7.00) and nickel(II) (pD = 6.50).

The use of 1,2-naphthoquinone-2-semicarbazone, 1,2-naphthoquinone-2-semicarbzone-4-sulphonic acid and 1,2-naphthoquinone-2-thiosemicarbazone-4-sulphonic acid as acid-base indicators has been studied. The sharpness of the indicator transitions has been investigated by means of photometric titrations and the colour quality specified with the aid of the CIE chromaticity system. The results show that the three substances are satisfactory as neutralization indicators.

Half plasma disappearance time (HDT) of endogenous erythropoietin (Ep) was measured in single rats, using an in vitro assay system for EP. In rats treated with the hepatotoxic agent d-Galactosamine-HCl (GalN), a small but significant elevation of HDT was found as compared with control rats (164 and 105 min, respectively). In bilaterally nephrectomized rats mean HDT was significantly elevated: 266 min. Combination of nephrectomy and GalN treatment did not result in a significant further elongation of HDT (mean = 301 min). In experiments using isolated liver perfusion, rat livers (with and without GalN treatment) were shown to be unable to change perfusate Ep titre during 4 h of perfusion. It is concluded that hepatic degradation of Ep in rats is only minimal. The kidney however is important in the catabolism of Ep.