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Arthroplasty is used to relieve pain associated with degenerative or inflammatory joint disease, some post-traumatic joint problems, and avascular necrosis. Avascular necrosis, inflammatory and post-traumatic problems are seen on a regular basis in areas of high HIV seroprevalence. Degenerative arthritis is rare in younger HIV patients, however. Historically the only group of HIV patients in which arthroplasty has been common is that which received contaminated factor VIII transfusions in the 1980's. Haemophiliacs get a haemophilic arthropathy from repeated bleeds into joints and so is an additional complication. Much of the previous literature on this topic has focused on haemophiliac patients. This review examines the success of arthroplasty in HIV positive patients, with an emphasis on non-haemophiliac patients. We conclude that arthroplasty can be a safe procedure for HIV positive individuals if the surgery is carried out in good conditions, and early results are encouraging.

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The observation that LINE-1 transposable elements are enriched on the X in comparison to the autosomes led to the hypothesis that LINE-1s play a role in X chromosome inactivation. If this hypothesis is correct, loss of LINE-1 activity would be expected to result in species extinction or in an alternate pathway of dosage compensation. One such alternative pathway would be to evolve a karyotype that does not require dosage compensation between the sexes. Two of the three extant species of the Ryukyu spiny rat Tokudaia have such a karyotype; both males and females are XO. We asked whether this karyotype arose due to loss of LINE-1 activity and thus the loss of a putative component in the X inactivation pathway. Although XO Tokudaia has no need for dosage compensation, LINE-1s have been recently active in Tokudaia osimensis and show higher density on the lone X than on the autosomes.

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New World monkeys are valuable animal models to study human diseases. To determine the phenotype of cells involved in immune responses, we used flow cytometry to screen a large panel of anti-human monoclonal antibodies (mAb) for cross-reactivity with cells of the common marmoset and the cotton-top tamarin. Certain antigens (e.g., CD2, CD8, CD20) are well conserved. However, CD10, CD23, and CD33 showed a clear discrepancy in their reaction patterns in both species, indicating that significant differences on the epitope level occurred during evolution. Epstein-Barr virus-transformed B-cell lines were shown to be a valuable tool for screening B-cell-specific reagents. In some cases, fluorescein isothiocyanate (FITC) and phycoerythrin (PE) modification of mAbs had a negative effect on the binding capacity, which stressed the importance of choosing the right label. Despite the fact that some CD antigens were not detected, adequate numbers of cross-reactive mAbs were identified to perform extensive studies on immunological functions in both the common marmoset and the cotton-top tamarin.

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OBJECTIVE: To determine the predictive value(s) of beta-hCG serum levels for pregnancy outcome following blastocyst transfer. DESIGN: Retrospective review. SETTING: University-based assisted reproductive technology (ART) program. PATIENTS: All ART patients enrolled from January 1998 to December 1999. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Beta-hCG serum levels and pregnancy outcomes. RESULT(S): Of the 836 ART cycles initiated, 608 embryo transfers met study criteria and were assigned to one of two groups: 248 day 5 blastocyst transfers or 360 day 3 embryo transfers. In the day 5 blastocyst group, 147 pregnancies occurred (59.2%), and day 3 transfers resulted in 165 pregnancies (45.8%). For day 3 and day 5 transfers, mean values of beta-hCG on day 16 post-retrieval of spontaneous abortions were lower than ongoing pregnancies (P< .05). A beta-hCG value on day 16 of >300 mIU/mL predicted an ongoing pregnancy for day 5 transfer group in 97% of pregnancies compared with 92% for day 3 embryo transfers. A multiple gestation was observed in 70% of pregnancies with a beta-hCG level >400 mIU/mL in the day 5 group compared with 63% for the day 3 group. The incidence of higher-order multiple gestations was significantly lower in the day 5 blastocyst group (P< .05). CONCLUSION(S): Beta-hCG serum levels on day 16 post-retrieval were highly predictive of pregnancy outcome after a blastocyst transfer.

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A 3-day-old male infant with a 3-cm firm subcutaneous mass was found to have decreased platelets, decreased fibrin, and increased fibrin split products diagnostic of Kasabach-Merritt phenomenon. The vascular lesion was resected without complications. We suggest that early surgical intervention is an excellent therapeutic option for Kasabach-Merritt phenomenon.

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An investigation of the possible interactions between combinations of vaccines and pyridostigmine bromide (PB) has been undertaken in the guinea pig. This study is part of a research programme funded by the UK Government to determine any effects of the pretreatment regimes given to UK Forces during the Persian Gulf conflict of 1990-1991. The study was designed to simulate PB administration and to model multiple vaccination protocols that were experienced by UK Forces, modelling a "worst case" situation in which all ten vaccines and PB were administered within a short period of time. Seven of the vaccines were health and hygiene (H+H) vaccines given to protect against endemic diseases and two vaccines to protect against the biological warfare agents anthrax and plague. In addition, pertussis vaccine was administered as an adjuvant to reduce the time to achieve immunity against anthrax. Four groups of eight animals were treated with 1/20th, 1/10th or 1/5th human doses of vaccines or vehicles, respectively. The PB or saline was delivered by implanted 28 day mini-osmotic pumps to achieve a mean red blood cell acetylcholinesterase (AChE) inhibition of around 30%. Body weight, temperature, immunological response, biochemical indices and spontaneous activity were monitored for 72 days. Although immunological responses to bacterial vaccines were observed, there were no remarkable findings in the parameters measured other than minor changes in body weight (4.9% decrease at the 1/5th human dose of vaccines) and temperature increases in response to vaccination. Animals in all groups remained generally healthy and active without visible adverse signs throughout the study. Reproduced with the permission of Her Majesty's Stationery Office. Published by John Wiley & Sons, Ltd.

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Genomes of the closely related bacteriophages phiX174 and S13 are 5386 bases long and differ at 114 nucleotides, affecting 28 amino acids. Both parental phages were adapted to laboratory culture conditions in replicate lineages and analysed for nucleotide changes that accumulated experimentally Of the 126 experimental substitutions, 90% encoded amino-acid changes, and 62% of the substitutions occurred in parallel in more than one experimental line. Furthermore, missense changes at 12 of the experimental sites were at residues differing between the parental phages; in ten cases the phiX174 experimental lineages were convergent with the S13 parent, or vice versa, at both the nucleotide and amino-acid levels. Convergence at a site was even obtained in both directions in three cases. These results point to a limited number of pathways taken during evolution in these viruses, and also raise the possibility that much of the amino-acid variation in the natural evolution of these viruses has been selected.

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The development of all low-order animals and noneutherian mammals follows an organized, polarized directional course from fertilization through fetal development. New evidence points to a fundamental polarization during all steps of mammalian development, from the early oocyte through fertilization and gastrulation. The generator of this polarization is primarily at the genetic level, with the results of gene expression and checkpoints being manifested in phenotype. Although cell-cell interactions reinforce the polarization of the embryo, they are not the underlying means of establishing axes in eutherian embryos. The ability of mammalian cells to remain totipotent is only partial, with little evidence that isolated blastomeres can result in full fetal development. The isolated blastomere can, however, contribute to development if reintroduced into a polarized environment. Polarization begins in the unovulated oocyte and is reinforced at fertilization. The axes and polarization established at fertilization endure through to the blastocyst stage and define the axes during gastrulation and fetal development.

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Health education graduate students were surveyed to assess perceptions of their professional responsibility to be role models of healthy behaviors, characteristics of a professional role model, and related socializing experiences during professional preparation. A total of 233 randomly selected health education graduate students participated in this study nationwide. Significant inverse associations were found between students' year in graduate school and sense of excellence as a role model, graduate program satisfaction, and professional commitment (all ps < 0.05). Students' sense of professional marketability and competence to role model were statistically significant in predicting their perception that role modeling healthy behaviors is a professional responsibility, F(2, 215) = 110.25, p = 0.00001. Positive associations also were found between students' desire to improve fitness behavior, nutrition, and weight and/or body fat ratio with self-ratings as role models (all ps < 0.05). Implications for the profession and preparation are provided.

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It is the responsibility of the profession to determine what is right, reasonable, and ethical health education practice. Opportunities within the profession abound to deliberate about the responsibility of health education specialists to role model positive health behaviors. Davis summarized the espoused perspectives nicely: We owe it to our profession and to our students to personally travel as far on the wellness continuum as behavioral choices will permit.... Health educators do have a special responsibility to be positive health role models by fulfilling their health potential and modeling the healthiest behaviors of which they are capable. All health educators need to accept for themselves the responsibilities that we assign to others.

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The molecular basis of adaptation is a major focus of evolutionary biology, yet the dynamic process of adaptation has been explored only piecemeal. Experimental evolution of two bacteriophage lines under strong selection led to over a dozen nucleotide changes genomewide in each replicate. At least 96 percent of the amino acid substitutions appeared to be adaptive, and half the changes in one line also occurred in the other. However, the order of these changes differed between replicates, and parallel substitutions did not reflect the changes with the largest beneficial effects or indicate a common trajectory of adaptation.

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A retrospective analysis of results from 114 initiated in-vitro fertilization cycles utilizing pronuclear embryo transfer is presented. Patients were unselected for age or infertility criteria, constituted a continuous series and were grouped according to response to stimulation (Group 1, ideal; Group 2, suboptimal) or ovarian reserve (Group 3, poor). At 16-18 h post-insemination, embryos were scored for alignment of pronuclei and nucleoli and the appearance of the cytoplasm, generating an embryo score (ES). Transfers were performed 24-26 h post-insemination using two to six embryos with the highest ES. A corrected score was calculated (total score/number of embryos; CS). A total of 114 initiated cycles resulted in 97 oocyte retrievals with 38 clinical pregnancies (39%; 15% implantation). Pregnancy rates were significantly different between the three groups; 37 pregnancies in Group 1 (55% clinical pregnancy; 20% implantation), none in Group 2 and one in Group 3 (6%; 2% implantation: P < 0.001). The ES of transferred embryos correlated with groups. There was a strong correlation between CS and implantation and delivery rates. CS >15 resulted in a 28% implantation; 65% delivery rate. CS <14 resulted in four pregnancies, one delivered. The data show that oocyte quality and pronuclear embryo morphology are related to implantation and that pronuclear embryos can be successfully selected for embryo transfer.

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Plants form their gametes late in somatic development and, as a result, often pass somatic mutations on to their progeny. Classic examples of this process are the germinal revertants of unstable, Ac/Ds transposon-induced kernel mutations in maize: frequent and early reversion events during somatic development are generally correlated with a high frequency of revertant gametes. We have characterized a Ds allele of the maize waxy (wx) gene, wx-m5:CS7, for which the correlation between somatic and germinal reversion frequencies no longer holds. The ability of wx-m5:CS7 (CS7) to produce revertant gametes is suppressed approximately 100-fold in comparison with a second Ds allele, wx-m5:CS8 (CS8), which has an identical insertion at Wx and the same frequent and early somatic reversion pattern in endosperm. The excision of Ds from wx is not reduced 100-fold in the somatic tissues of CS7 plants as compared with CS8 plants. Suppressed formation of CS7 revertant gametes is independent of the Ac transposase source and is heritably passed to the embryos of progeny kernels; however, frequent and early somatic reversion is observed again in endosperms of these progeny kernels. This suppression appears to be caused by a dominant mutation in a trans-acting product that can suppress the germinal reversion of other Ds-induced alleles as well; the mutation is tightly linked to Wx but is not in the CS7 Ds itself. Taken together, the data suggest a novel mode of developmental control of Ac/Ds elements by the host plant, suppressing element excision in the shoot meristem.

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The purpose of this article is to provide clinicians information on some of the key issues in making decisions about the treatment process for children who stutter. Ten decisions that should be considered prior to, during, and at the time of dismissal are discussed. These decisions relate to a number of issues regarding treatment such as: increasing clinicians' confidence in treating stuttering in children, setting long- and short-term goals, selecting an approach to treatment, documenting progress, involving parents and teachers in the treatment process, and determining when the child is ready to be dismissed from treatment. Additionally, questions clinicians should ask themselves are presented with each of the ten decisions. The intent of this article is to show how an analysis of clinical information systematically collected over a period of time will help a clinician make accurate decisions about the treatment of children who stutter.

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Irreversibly sickled cells (ISCs) remain sickled even under conditions where they are well oxygenated and hemoglobin is depolymerized. In our studies we demonstrate that triton extracted ISC core skeletons containing only spectrin, protein 4.1, and actin also retain their sickled shape; while reversibly sickled cell (RSC) skeletons remodel to a round or biconcave shape. We also demonstrate that these triton extracted ISC core skeletons dissociate more slowly upon incubation at 37 degrees C than do RSC or control (AA) core skeletons. This observation may supply the basis for the inability of the ISC core skeleton to remodel its shape. Using an in vitro ternary complex dissociation assay, we demonstrate that a modification in beta-actin is the major determinant of the slow dissociation of the spectrin-protein 4.1-actin complex isolated from the ISC core skeleton. We demonstrate that the difference between ISC and control beta-actin is the inaccessibility of two cysteine residues in ISC beta-actin to labeling by thiol reactive reagents; due to the formation of a disulfide bridge between cysteine284 and cysteine373 in ISC beta-actin, or alternatively another modification of cysteine284 and cysteine373 which is reversible with DTT and adds less than 100 D to the molecular weight of beta-actin.

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We have previously cloned cDNAs that correspond to two different nonallelic preproinsulin genes in the amphibian, Xenopus laevis (Shuldiner et al., 1989, J. Biol. Chem. 264, 9428-9432). The coding regions of the two genes are very similar (i.e., 93% amino acid identity). While both preproinsulin genes are expressed coordinately in the adult pancreas, they are differentially expressed in prepancreatic embryos during neurulation (Shuldiner et al., 1991, Proc. Natl. Acad. Sci. USA 88, 7679-7683). We now report the use of the polymerase chain reaction (PCR) to investigate the genomic structures of both Xenopus preproinsulin genes. First, the nucleotide sequence of a portion of the 5' untranslated region that was lacking in our cDNA clones was obtained using rapid amplification of cDNA ends (5' RACE). Then, oligonucleotide primers were designed that flanked putative exon-intron splice boundaries, and intronic sequences in each of the two nonallelic genes were amplified. We determined that both Xenopus preproinsulin genes contain two introns, one interrupting the 5' untranslated region, and the second interrupting the region encoding the C-peptide. The introns are similar in position, but of greater length than those reported in most other species. Interestingly, dideoxy sequence analysis of the PCR-amplified introns revealed that the exon-intron splice junctions are well conserved between the two nonallelic genes, but internal to these junctions, the respective introns diverge significantly from each other. Thus, ample time has elapsed since the duplication of these two genes for marked divergence to occur within the introns suggesting that these regions are not important for expression in the adult pancreas. From these studies, we predict that elucidation of the sequences of the 5' flanking regions of the two nonallelic preproinsulin genes will reveal conserved regions that will be important for coordinate expression, as well as less conserved regions that will either be unimportant for coordinate expression (i.e., pancreatic expression) or important for differential expression (i.e., prepancreatic expression).

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To study the potential role of insulin-like growth factor-I (IGF-I) during early embryogenesis, we have used the amphibian Xenopus laevis, a versatile model of vertebrate development. Using polymerase chain reaction (PCR)-based cloning strategies, we have previously identified two different nonallelic Xenopus IGF-I genes (IGF-I' and IGF-I"). Both are expressed in similar quantities in adult liver. We now report the use of a modification of the reverse transcription-PCR method, designated RNA template-specific PCR (RS-PCR), to detect IGF-I messenger RNA (mRNA) from single oocytes and embryos. The same primer pair was used to amplify both IGF-I' and IGF-I" mRNAs. Slot blot analysis of the RS-PCR products with internal oligonucleotide probes that specifically recognize IGF-I' or IGF-I" sequences revealed that only IGF-I' mRNA was present in follicles surrounding mature (stage VI) oocytes; neither IGF-I mRNA was present in follicles surrounding less mature oocytes (stages I and IV) or within oocytes or unfertilized eggs. After fertilization, IGF-I' mRNA was first detected during early organogenesis (stages 21-23), and increased during subsequent stages of development. To localize early IGF-I' expression, a stage 27 embryo was sliced into 24-microns cross-sections; RS-PCR and slot blot analysis were performed on RNA extracts from consecutive sections. IGF-I' mRNA was expressed in all sections, but was most abundant in the body region from which the visceral organs were developing. In contrast, to the early expression of IGF-I', IGF-I" mRNA was not detected until stage 41, a period corresponding to premetamorphic growth. Reverse PCR for Xenopus GH mRNA demonstrated that the onset of GH gene expression was coincident with the onset of IGF-I" gene expression (stage 41). These data suggest that the early expression of IGF-I' may be GH independent, whereas later expression of IGF-I" is GH dependent. We conclude that the two nonallelic IGF-I genes are expressed differentially in a stage-specific manner and suggest that IGF-I' may play an important role during early organogenesis, whereas IGF-I" may be important for premetamorphic growth.

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The laboratory component of assisted reproductive technologies involves the manipulation and culture of male and female gametes in vitro, with the goal of producing viable embryos for transfer to a woman to enable the establishment of pregnancy. The biology of the gametes and resulting embryos dictates how they should be handled in vitro to maximize their potential at establishing a viable pregnancy. This article examines the biology of gametes and offers some suggestions as how best to proceed with the process of in vitro fertilization, embryo culture, and embryo transfer.

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Often, it is convenient to subclone polymerase chain reaction (PCR) products into a plasmid vector for subsequent replication in bacteria, but conventional subcloning methods often fail. We report a rapid and versatile method to subclone PCR products directionally into a specific site of virtually any plasmid vector. The procedure requires only four primers, does not require DNA ligase, and may be accomplished in a single day. Ligase-free subcloning is performed by incorporating into the PCR primers sequences at the 5' ends that result in PCR products whose 3' ends are complementary to the 3' ends of the recipient linearized plasmid. The PCR product and the linearized plasmid are spliced together in a second PCR reaction in which Taq polymerase extends the complementary overlapping 3' ends (ligation by overlap extension). Denaturation followed by heterologous reannealing and cyclization results in a cyclic recombinant plasmid with two nicks that may be used directly to transform competent Escherichia coli. In our hands, ligase-free subcloning is rapid, and offers many advantages over existing strategies.

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