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Biomedical subjects

J Cohen

Publications and source records attributed to J Cohen.

At least 307 records · Page 17Linked to original sources
PubMed1999

Preimplantation genetic diagnosis of aneuploidy: were we looking at the wrong chromosomes?

PURPOSE: Our purpose was to study aneuploidy frequencies of chromosomes 1, 4, 6, 7, 14, 15, 17, 18, and 22 in cleavage-stage embryos. These frequencies were compared to spontaneous abortion data to determine differences in survival rate of their aneuploidies. METHODS: One hundred ninety-four embryos were analyzed with multicolor fluorescence in situ hybridization. Embryos were divided into three maternal age groups: 20 to 34.9 years, (2) 35 to 39.9 years, and (3) 40 years and older. Embryos were also divided into two developmental and morphological groups; arrested and nonarrested embryos. RESULTS: The rate of aneuploidy was 14.51%, 14.10%, and 31.48% for age groups 1, 2, and 3, respectively (P < 0.005). The chromosomes most frequently involved in aneuploidy events were 22, 15, 1, and 17. CONCLUSIONS: The chromosomes most involved in spontaneous abortions are not necessarily the ones causing a decrease in implantation rates with maternal age. Other aneuploidies, such as for chromosomes 1 and 17, may seldom implant or die shortly after implantation.

Adult↗
PubMed1999

Molecular analysis of the role of streptococcal pyrogenic Exotoxin A (SPEA) in invasive soft-tissue infection resulting from Streptococcus pyogenes.

Epidemiological studies strongly implicate the bacterial superantigen, streptococcal pyrogenic exotoxin A (SPEA), in the pathogenesis of necrotizing soft-tissue infection and toxic shock syndrome resulting from Streptococcus pyogenes. SPEA can act as a superantigen and cellular toxin ex vivo, but its role during invasive streptococcal infection is unclear. We have disrupted the wild-type spea gene in an M1 streptococcal isolate. Supernatants from toxin-negative mutant bacteria demonstrated a 50% reduction in pro-mitogenic activity in HLA DQ-positive murine splenocyte culture, and up to 20% reduction in activity in human PBMC culture. Mutant and wild-type bacteria were then compared in mouse models of bacteraemia and streptococcal muscle infection. Disruption of spea was not associated with attenuation of virulence in either model. Indeed, a paradoxical increase in mutant strain-induced mortality was seen after intravenous infection. Intramuscular infection with the SPEA-negative mutant led to increased bacteraemia at 24 h and a reduction in neutrophils at the site of primary muscle infection. Purified SPEA led to a dose-dependent increase in peritoneal neutrophils 6 h after administration. SPEA is not a critical virulence factor in invasive soft-tissue infection or bacteraemia caused by S. pyogenes, and it could have a protective role in murine immunity to pyogenic infection. The role of this toxin may be different in hosts with augmented superantigen responsiveness.

Animals↗
PubMed1999

Growth responses of different subpopulations of adult sensory neurons to neurotrophic factors in vitro.

Different subpopulations of adult primary sensory neurons in the dorsal root ganglia express receptors for different trophic factors, and are therefore potentially responsive to distinct trophic signals. We have compared the effect of the neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and NT-3, and of glial cell line-derived neurotrophic factor (GDNF) on neurite outgrowth in dissociated cultures of sensory neurons from the lumbar ganglia of young adult rats, and attempted to establish subset-specific effects of these trophic factors. We analysed three parameters of neurite growth (percentage of process-bearing neurons, length of longest neurite and total neurite length), which may correlate with particular types of axon growth in vivo, and may therefore respond differently to trophic factor presence. Our results showed that percentage of process-bearing neurons and total neurite length were influenced by trophic factors, whilst the length of the longest neurite was trophic factor independent. Only NGF and GDNF were found to enhance significantly the proportion of process-bearing neurons in vitro. GDNF was more effective than NGF on small, IB4- neurons, which are known to develop GDNF responsiveness early in postnatal development. NGF, and to a much lesser extent GDNF, enhanced the total length of the neurites produced by neurons in culture. BDNF exerted an inhibitory effect on growth, and both BDNF and NT-3 could partially block some of the growth-promoting effects of NGF on specific neuronal subpopulations.

Age Factors↗
PubMed1999

A statistically robust variance-components approach for quantitative trait linkage analysis.

Previously we showed (Wang, Guerra & Cohen 1998) that a statistically robust version of the Haseman & Elston (1972) sib-pair method greatly increased power to detect linkage in the presence of outliers. In this paper we report on M-estimation to accommodate outliers in the variance-components approach to linkage analysis developed by Amos (1994). Simulations show that in the presence of outliers the robust variance-components approach provides substantially greater power, more precise estimation of heritabilities, and better false-positive rates than the original Gaussian based approach. In the absence of outliers the performance of the robust variance-components approach is similar to that of the Gaussian based approach. For illustration we apply the method to two well characterized lipoprotein systems.

Alleles↗
PubMed1999

Mediterranean spotted fever in pregnancy.

Mediterranean spotted fever has rarely been reported in pregnancy. We report a case occurring in a young pregnant woman, which responded well to treatment with a combination of erythromycin and rifampicin. The treatment of spotted fevers in pregnancy is discussed in detail.

Adult↗
PubMed1999

Evidence for linkage of adolescent-onset idiopathic generalized epilepsies to chromosome 8-and genetic heterogeneity.

Several loci and candidate genes for epilepsies or epileptic syndromes map or have been suggested to map to chromosome 8. We investigated families with adolescent-onset idiopathic generalized epilepsy (IGE), for linkage to markers spanning chromosome 8. The IGEs that we studied included juvenile myoclonic epilepsy (JME), epilepsy with only generalized tonic-clonic seizures occurring either randomly during the day (random grand mal) or on awakening (awakening grand mal), and juvenile absence epilepsy (JAE). We looked for a gene common to all these IGEs, but we also investigated linkage to specific subforms of IGE. We found evidence for linkage to chromosome 8 in adolescent-onset IGE families in which JME was not present. The maximum multipoint LOD score was 3.24 when family members with IGE or generalized spike-and-waves (SW) were considered affected. The LOD score remained very similar (3.18) when clinically normal family members with SW were not considered to be affected. Families with either pure grand mal epilepsy or absence epilepsy contributed equally to the positive LOD score. The area where the LOD score reaches the maximum encompasses the location of the gene for the beta3-subunit of the nicotinic acetylcholine receptor (CHRNB3), thus making this gene a possible candidate for these specific forms of adolescent-onset IGE. The data excluded linkage of JME to this region. These results indicate genetic heterogeneity within IGE and provide no evidence, on chromosome 8, for a gene common to all IGEs.

Adolescent↗
PubMed1999

Microbiologic findings and correlations with serum tumor necrosis factor-alpha in patients with severe sepsis and septic shock.

To understand the microbiology of sepsis and its relationship with tumor necrosis factor (TNF)-alpha, 444 septic patients were studied in a phase II clinical trial. In total, 270 (61%) 444 of episodes were microbiologically documented. The most common isolates were Escherichia coli, Staphylococcus aureus, and Streptococcus pneumoniae. In some patients, obligate anaerobes were isolated in pure culture. Data on baseline TNF-alpha (n=409) showed that the geometric mean TNF-alpha concentrations in patients with severe sepsis and in those with late shock were 41 pg/mL (range, 5-3310) and 58 pg/mL (range, 5-1870), respectively. Patients with a positive culture had significantly higher TNF-alpha levels (65.9 vs. 29.2 pg/mL, P=.0001). Patients with a pure gram-negative infection had significantly higher TNF-alpha levels than those with a pure gram-positive or mixed infection, especially in the late shock group (142.6, 64.0, and 52.8 pg/mL, respectively, P=.004). These results provide further support for the concept that patients with sepsis are a heterogeneous group that require more precise definition.

Adolescent↗
PubMed1999

Potent induction of focused Th1-type cellular and humoral immune responses by RTS,S/SBAS2, a recombinant Plasmodium falciparum malaria vaccine.

The RTS,S/SBAS2 vaccine confers sterile protection against Plasmodium falciparum sporozoite challenge. The mechanisms underlying this are of great interest, yet little is known about the immune effector mechanisms induced by this vaccine. The immune responses induced by RTS,S/SBAS2 were characterized in 10 malaria-naive volunteers. Several epitopes in the circumsporozoite protein (CSP) were identified as targets of cultured interferon (IFN)-gamma-secreting CD4+ T cells. RTS,S-specific IFN-gamma-secreting effector T cells were induced in 8 subjects; this ex vivo response mapped to a single peptide in Th2R. CSP-specific CD8+ cytotoxic T lymphocytes were not detected. RTS, S-specific IFN-gamma production was universal, whereas interleukin-4 and -5 production was rare. RTS,S-specific lymphoproliferative responses and antibodies to CSP were strongly induced in all volunteers. Responses waned with time but were boostable. Thus, RTS, S/SBAS2 is a potent inducer of Th1-type cellular and humoral immunity. These results highlight possible immune mechanisms of protection and have important implications for vaccine design in general.

Adult↗
PubMed1999

Hospital gangrene: the scourge of surgeons in the past.

Before the days of antisepsis, the infection rate on surgical wards was very high. Mortality from common operations such as limb amputations could run from 40% to 60%, mostly from infection. Hospital gangrene, a type of necrotizing infection, occurred frequently. It would spread rapidly, and the helpless surgeon could do nothing to stop it. The patient would be in agony for many days before death mercifully intervened.

Amputation, Surgical↗
PubMed1999

Rapid visualization of metaphase chromosomes in single human blastomeres after fusion with in-vitro matured bovine eggs.

The present study was aimed to facilitate karyotyping of human blastomeres using the metaphase-inducing factors present in unfertilized eggs. A rapid technique for karyotyping would have wide application in the field of preimplantation genetic diagnosis. When cryopreserved in-vitro matured bovine oocytes were fused with human blastomeres, the transferred human nuclei were forced into metaphase within a few hours. Eighty-seven human blastomeres from abnormal or arrested embryos were fused with bovine oocytes in a preclinical study. Fusion efficiency was 100%. In 21 of the hybrid cells, no trace of human chromatin was found. Of the remaining 66, 64 (97%) yielded chromosomes suitable for analysis. The method was used to karyotype embryos from two patients with maternal translocations. One embryo which was judged to be karyotypically normal was replaced in the first patient, resulting in one pregnancy with a normal fetus. None of the second patient's embryos was diagnosed as normal, and hence none was transferred. The results of the present study demonstrated that the ooplasmic factors which induce and maintain metaphase in bovine oocytes can force transferred human blastomere nuclei into premature metaphase, providing the basis for a rapid method of karyotyping blastomeres from preimplantation embryos and, by implication, cells from other sources.

Adult↗
PubMed1999

Positive outcome after preimplantation diagnosis of aneuploidy in human embryos.

usromosomal abnormalities are responsible for a great deal of embryo wastage, which is reflected, at least partially, in decreased implantation and increased miscarriage in older women. To address this problem the transfer of only chromosomally normal embryos previously selected by preimplantation genetic diagnosis (PGD) has been proposed. We designed a multi-centre in-vitro fertilization (IVF) study to compare controls and a test group that underwent embryo biopsy and PGD for aneuploidy. Patients were matched retrospectively, but blindly, for average maternal age, number of previous IVF cycles, duration of stimulation, oestradiol concentrations on day +1, and average mature follicles. All these parameters were similar in test and control groups. Only embryos classified as normal for those chromosomes were transferred after PGD. The results showed that the rates of fetal heart beat (FHB)/embryo transferred between the control and test groups were similar. However, spontaneous abortions, measured as FHB aborted/FHB detected, decreased after PGD (P < 0.05), and ongoing pregnancies and delivered babies increased (P < 0.05) in the PGD group of patients. Two conclusions were obtained: (i) PGD of aneuploidy reduced embryo loss after implantation; (ii) implantation rates were not significantly improved, but the proportion of ongoing and delivered babies was increased.

Aneuploidy↗
PubMed1999

Mitochondrial DNA rearrangements in human oocytes and embryos.

Human mitochondrial DNA (mtDNA) rearrangements, including more than 150 deletions and insertions, accumulate with age and are responsible for certain neuromuscular diseases. Human oocytes, arrested for up to 50 years, may express certain mtDNA rearrangements possibly affecting function. Investigations have previously shown a single mtDNA rearrangement (dmtDNA(4977)) in human oocytes. Sequencing of other rearrangements and their correlation with maternal age have not been performed in human oocytes or embryos. Here we use a nested PCR strategy of long followed by short polymerase chain reaction (PCR) that amplifies two-thirds of the mitochondrial genome. mtDNA rearrangements were detected in 50.5% of the oocytes (n = 295) and 32.5% of the embryos (n = 197). This represents a significant difference in the percentage of mtDNA rearrangements between oocytes and embryos (P < 0.0001). Twenty-three novel mtDNA rearrangements with deletions, insertions and duplications were found. There was no significant age-related increase in the percentage of human oocytes or embryos that contained mtDNA rearrangements. Significant reductions in the number of oocytes containing mtDNA rearrangements occurred as oocyte development progressed from germinal vesicle to the mature metaphase II oocyte (P < 0.05). These findings are discussed as they relate to mitochondria, mtDNA, and ATP production in human oocytes and embryos.

Adenosine Triphosphate↗
PubMed1999

Analysis of gene expression in single oocytes and embryos by real-time rapid cycle fluorescence monitored RT-PCR.

Rapid cycle DNA amplification is a refinement of the polymerase chain reaction (PCR) method that permits increased product specificity while reducing amplification time by an order of magnitude. Combined with the use of micro volume capillaries, minute samples can be examined by this technique. Thus, this approach is ideally suited to the analysis of gene expression in individual cells. As the current understanding of early developmental processes is still rudimentary, further characterization of transcription in single oocytes and embryos may provide additional insight into the molecular mechanisms directing these events. In this study, we examined the suitability of fluorescence monitored reverse transcription (RT)-PCR for the study of gene expression during oogenesis and embryogenesis using transcripts of the housekeeping gene, beta-actin, as an experimental model. Product accumulation was monitored by either the double-stranded DNA dye SYBR Green I or sequence-dependent hybridization of reporter molecules called molecular beacons. Dyes bind generically and are economical to use. However, both specific and non-specific products are labelled. Hybridization probes permit very specific and sensitive target recognition but they can be costly to manufacture. Once molecular markers indicative of optimal development are identified, this technology could be used in a clinical in-vitro fertilization laboratory as a diagnostic tool.

Animals↗
PubMed1999

Search for a human homologue of the mouse Ped gene.

The Ped gene influences the rate of cleavage division of preimplantation mouse embryos and subsequent embryonic survival. The mouse Ped gene product is a major histocompatibility complex (MHC) class Ib protein called Qa-2. Studies from many human in-vitro fertilization (IVF) clinics suggest that the mouse Ped gene has a human homologue because embryos fertilized at the same time have different cleavage rates, and those embryos that cleave at a faster rate are more likely to result in a viable pregnancy. Candidates for the human homologue of the mouse Ped gene include the MHC class Ib genes HLA-E, HLA-F, and HLA-G. The presence of mRNA for these three genes was tested in 108 spare day 3 human preimplantation embryos from 25 couples by using reverse transcription-polymerase chain reaction (RT-PCR). Of the 86 embryos tested for HLA-E mRNA, 72 were positive (84%), and of the 88 embryos tested for HLA-G mRNA, 39 were positive (44%). None of the 17 embryos tested for HLA-F mRNA were positive (0%). Studies of expression of HLA-G protein were undertaken to ascertain whether HLA-G was attached to the cell membrane via a glycosylphosphatidylinositol (GPI) linkage similar to that found in Qa-2 protein. Treatment of JEG-3 cells, an HLA-G expressing cell line, with phospholipase C did not result in removal of HLA-G showing that HLA-G, unlike Qa-2, is not GPI linked to the cell surface. The pros and cons of HLA-E, HLA-F, and HLA-G as candidates for the human Ped gene are discussed.

Animals↗
PubMed1999

Alternative splicing of the telomerase catalytic subunit in human oocytes and embryos.

The human telomerase catalytic subunit (hTCS) is a ribonucleoprotein which synthesizes telomere repeats on the ends of chromosomes. Telomerase activity is thought to be essential in maintaining normal telomere length in immortal (cancer) and germ cells. The objective of this study was to determine the gene expression of telomerase mRNA in human oocytes at different meiotic stages and in embryos. Normal and abnormal human oocytes, preimplantation embryos, and blastocysts were analysed for the presence and expression of the hTCS transcripts. Multiple telomerase mRNA products were identified by reverse transcription-polymerase chain reaction (RT-PCR) using primers within the reverse transcriptase domain. DNA sequencing of these amplicons suggest that there are alternative splicing variants which align to other telomerase reverse transcriptase (RT) consensus domains. Surprisingly, in unfertilized and immature gametes, as well as preimplantation embryos, hTCS expression revealed three different PCR product sizes, 457, 421 and 275 bp. The frequency of the 275 bp DNA product was 6.6% in oocytes (two out of 30) compared with 56.6% (17 out of 30) in poorly developing human preimplantation embryos (P < 0.005). The presence of alternately spliced mRNA variants in human preimplantation embryos may suggest a lack of telomerase activity and thus chromosomes associated with shortened telomeres.

Alternative Splicing↗
PubMed1999

Community nephrology: audit of screening for renal insufficiency in a high risk population.

BACKGROUND: The rate of acceptance onto dialysis programmes has doubled in the past 10 years and is steadily increasing. Early detection and treatment of renal failure slows the rate of progression. Is it feasible to screen for patients who are at increased risk of developing renal failure? We have audited primary care records of patients aged 50-75 years who have either hypertension or diabetes, and are therefore considered to be at high risk of developing renal insufficiency. Our aim was to see whether patients had had their blood pressure measured and urine tested for protein within 12 months, and plasma creatinine measured within 24 months. METHODS: This was a retrospective study of case notes and computer records in 12 general practices from inner and greater London. A total of 16,855 patients were aged 50-75 years. From this age group, 2693 (15.5%) patients were identified as being either hypertensive or diabetic, or both. RESULTS: Of the 2561 records audited, 1359 (53.1%) contained a plasma creatinine measured within 24 months, and 11% of these (150) had a value > 125 micromol/l. This equates to a prevalence of renal insufficiency of > 110,000 patients per million in this group. Forty two patients (28%) had been referred to a nephrologist. Of records audited, 73% contained a blood pressure measurement and 29% contained a test for proteinuria within 12 months. CONCLUSIONS: There is a high prevalence of chronic renal insufficiency in hypertensive and diabetic patients. It is feasible to detect renal insufficiency at a primary care level, but an effective system will require computerized databases that code for age, ethnicity, measurement of blood pressure and renal function, as well as diagnoses.

Aged↗
PubMed1999

Clinical gram-positive sepsis: does it fundamentally differ from gram-negative bacterial sepsis?

OBJECTIVE: To review the basic differences between gram-positive and gram-negative sepsis and to assess the effect of these differences on current and future therapeutic strategies for sepsis. DESIGN: Literature review of the past 30 yrs of laboratory and clinical reports that analyze the microbial aspects of sepsis and the immunologic response to systemic infection. RESULTS: The increasing prevalence of sepsis from gram-positive bacterial pathogens necessitates reevaluation of many of the basic assumptions about the molecular pathogenesis of septic shock. It has been assumed that the initiation of the systemic inflammatory response with activation of the proinflammatory cytokine networks and other mediators results in a similar pathophysiologic process, regardless of the causative microbic pathogen. Yet, there is increasing experimental evidence that fundamental differences exist in the host response to gram-positive bacterial pathogens compared with the host response to gram-negative organisms. Systemic immune activation during sepsis may promote the clearance of the microbic pathogen; however, generalized inflammation also contributes to the pathogenesis of septic shock. The balance between these beneficial and deleterious effects may differ between gram-positive and gram-negative pathogens. CONCLUSIONS: Results of antimediator therapies in clinical trials in septic shock are inconclusive but suggest that the response may differ, depending on the type of microbic pathogen. The immune-mediated pathophysiologic mechanisms that underlie gram-positive sepsis and the potential interactions between the infecting microorganism and efficacy of anticytokine therapies require further investigation. Treatment strategies that explain the causative organism may be necessary for optimal use of immunoadjuvants in the future.

Anti-Inflammatory Agents↗