Massive dysregulation of genes involved in cell signaling and placental development in cloned cattle conceptus and maternal endometrium.

A major unresolved issue in the cloning of mammals by somatic cell nuclear transfer (SCNT) is the mechanism by which the process fails after embryos are transferred to the uterus of recipients before or during the implantation window. We investigated this problem by using RNA sequencing (RNA-seq) to compare the transcriptomes in cattle conceptuses produced by SCNT and artificial insemination (AI) at day (d) 18 (preimplantation) and d 34 (postimplantation) of gestation. In addition, endometrium was profiled to identify the communication pathways that might be affected by the presence of a cloned conceptus, ultimately leading to mortality before or during the implantation window. At d 18, the effects on the transcriptome associated with SCNT were massive, involving more than 5,000 differentially expressed genes (DEGs). Among them are 121 genes that have embryonic lethal phenotypes in mice, cause defects in trophoblast and placental development, and/or affect conceptus survival in mice. In endometria at d 18, <0.4% of expressed genes were affected by the presence of a cloned conceptus, whereas at d 34, ∼36% and <0.7% of genes were differentially expressed in intercaruncular and caruncular tissues, respectively. Functional analysis of DEGs in placental and endometrial tissues suggests a major disruption of signaling between the cloned conceptus and the endometrium, particularly the intercaruncular tissue. Our results support a "bottleneck" model for cloned conceptus survival during the periimplantation period determined by gene expression levels in extraembryonic tissues and the endometrial response to altered signaling from clones.

Hemolysis-induced lethality involves inflammasome activation by heme.

The increase of extracellular heme is a hallmark of hemolysis or extensive cell damage. Heme has prooxidant, cytotoxic, and inflammatory effects, playing a central role in the pathogenesis of malaria, sepsis, and sickle cell disease. However, the mechanisms by which heme is sensed by innate immune cells contributing to these diseases are not fully characterized. We found that heme, but not porphyrins without iron, activated LPS-primed macrophages promoting the processing of IL-1ß dependent on nucleotide-binding domain and leucine rich repeat containing family, pyrin domain containing 3 (NLRP3). The activation of NLRP3 by heme required spleen tyrosine kinase, NADPH oxidase-2, mitochondrial reactive oxygen species, and K(+) efflux, whereas it was independent of heme internalization, lysosomal damage, ATP release, the purinergic receptor P2X7, and cell death. Importantly, our results indicated the participation of macrophages, NLRP3 inflammasome components, and IL-1R in the lethality caused by sterile hemolysis. Thus, understanding the molecular pathways affected by heme in innate immune cells might prove useful to identify new therapeutic targets for diseases that have heme release.

Interferon-γ and granulocyte/monocyte colony-stimulating factor production by natural killer cells involves different signaling pathways and the adaptor stimulator of interferon genes (STING).

Natural killer (NK) cells are important for innate immunity in particular through the production of IFN-γ and GM-CSF. Both cytokines are important in restoration of immune function of tolerized leukocytes under inflammatory events. The expression of TLRs in NK cells has been widely studied by analyzing the mRNA of these receptors, rarely seeking their protein expression. We previously showed that murine spleen NK cells express TLR9 intracellularly and respond to CpG oligodeoxynucleotide (CpG-ODN) by producing IFN-γ and GM-CSF. However, to get such production the presence of accessory cytokines (such as IL-15 and IL-18) was required, whereas CpG-ODN or accessory cytokines alone did not induce IFN-γ or GM-CSF. We show here that TLR9 overlaps with the Golgi apparatus in NK cells. Furthermore, CpG-ODN stimulation in the presence of accessory cytokines induces the phosphorylation of c-Jun, STAT3, and IκBα. IFN-γ and GM-CSF production requires NF-κB and STAT3 activation as well as Erk-dependent mechanisms for IFN-γ and p38 signaling for GM-CSF. Using knock-out-mice, we show that UNC93b1 and IL-12 (produced by NK cells themselves) are also necessary for IFN-γ and GM-CSF production. IFN-γ production was found to be MyD88- and TLR9-dependent, whereas GM-CSF was TLR9-independent but dependent on STING (stimulator of interferon genes), a cytosolic adaptor recently described for DNA sensing. Our study thereby allows us to gain insight into the mechanisms of synergy between accessory cytokines and CpG-ODN in NK cells. It also identifies a new and alternative signaling pathway for CpG-ODN in murine NK cells.

Heme induces programmed necrosis on macrophages through autocrine TNF and ROS production.

Diseases that cause hemolysis or myonecrosis lead to the leakage of large amounts of heme proteins. Free heme has proinflammatory and cytotoxic effects. Heme induces TLR4-dependent production of tumor necrosis factor (TNF), whereas heme cytotoxicity has been attributed to its ability to intercalate into cell membranes and cause oxidative stress. We show that heme caused early macrophage death characterized by the loss of plasma membrane integrity and morphologic features resembling necrosis. Heme-induced cell death required TNFR1 and TLR4/MyD88-dependent TNF production. Addition of TNF to Tlr4(-/-) or to Myd88(-/-) macrophages restored heme-induced cell death. The use of necrostatin-1, a selective inhibitor of receptor-interacting protein 1 (RIP1, also known as RIPK1), or cells deficient in Rip1 or Rip3 revealed a critical role for RIP proteins in heme-induced cell death. Serum, antioxidants, iron chelation, or inhibition of c-Jun N-terminal kinase (JNK) ameliorated heme-induced oxidative burst and blocked macrophage cell death. Macrophages from heme oxygenase-1 deficient mice (Hmox1(-/-)) had increased oxidative stress and were more sensitive to heme. Taken together, these results revealed that heme induces macrophage necrosis through 2 synergistic mechanisms: TLR4/Myd88-dependent expression of TNF and TLR4-independent generation of ROS.

Therapeutical targeting of nucleic acid-sensing Toll-like receptors prevents experimental cerebral malaria.

Excessive release of proinflammatory cytokines by innate immune cells is an important component of the pathogenic basis of malaria. Proinflammatory cytokines are a direct output of Toll-like receptor (TLR) activation during microbial infection. Thus, interference with TLR function is likely to render a better clinical outcome by preventing their aberrant activation and the excessive release of inflammatory mediators. Herein, we describe the protective effect and mechanism of action of E6446, a synthetic antagonist of nucleic acid-sensing TLRs, on experimental cerebral malaria (ECM) induced by Plasmodium berghei ANKA. We show that in vitro, low doses of E6446 specifically inhibited the activation of human and mouse TLR9. Tenfold higher concentrations of this compound also inhibited the human TLR8 response to single-stranded RNA. In vivo, therapy with E6446 diminished the activation of TLR9 and prevented the exacerbated cytokine response observed during acute Plasmodium infection. Furthermore, severe signs of ECM, such as limb paralysis, brain vascular leak, and death, were all prevented by oral treatment with E6446. Hence, we provide evidence that supports the involvement of nucleic acid-sensing TLRs in malaria pathogenesis and that interference with the activation of these receptors is a promising strategy to prevent deleterious inflammatory responses that mediate pathogenesis and severity of malaria.

Messenger RNAs in metaphase II oocytes correlate with successful embryo development to the blastocyst stage.

The mRNAs accumulated in oocytes provide support for embryo development until embryo genomic activation. We hypothesized that the maternal mRNA stock present in bovine oocytes is associated with embryo development until the blastocyst stage. To test our hypothesis, we analyzed the transcriptome of the oocyte and correlated the results with the embryo development. Our goal was to identify genes expressed in the oocyte that correlate with its ability to develop to the blastocyst stage. A fraction of oocyte cytoplasm was biopsied using micro-aspiration and stored for further expression analysis. Oocytes were activated chemically, cultured individually and classified according to their capacity to develop in vitro to the blastocyst stage. Microarray analysis was performed on mRNA extracted from the oocyte cytoplasm fractions and correlated with its ability to develop to the blastocyst stage (good quality oocyte) or arrest at the 8-16-cell stage (bad quality oocyte). The expression of 4320 annotated genes was detected in the fractions of cytoplasm that had been collected from oocytes matured in vitro. Gene ontology classification revealed that enriched gene expression of genes was associated with certain biological processes: 'RNA processing', 'translation' and 'mRNA metabolic process'. Genes that are important to the molecular functions of 'RNA binding' and 'translation factor activity, RNA binding' were also enriched in oocytes. We identified 29 genes with differential expression between the two groups of oocytes compared (good versus bad quality). The content of mRNAs expressed in metaphase II oocytes influences the activation of the embryonic genome and enables further develop to the blastocyst stage.

Associating Physical and Photocatalytic Properties of Recyclable and Reusable Blast Furnace Dust Waste.

Blast furnace dust waste (BFDW) proved efficient as a photocatalyst for the decolorization of methylene blue (MB) dye in water. Structural analysis unequivocally identified α-Fe2O3 as the predominant phase, constituting approximately 92%, with a porous surface showcasing unique 10-30 nm agglomerated nanoparticles. Chemical and thermal analyses indicated surface-bound water and carbonate molecules, with the main phase's thermal stability up to 900 °C. Electrical conductivity analysis revealed charge transfer resistance values of 616.4 Ω and electrode resistance of 47.8 Ω. The Mott-Schottky analysis identified α-Fe2O3 as an n-type semiconductor with a flat band potential of 0.181 V vs. Ag/AgCl and a donor density of 1.45 × 1015 cm-3. The 2.2 eV optical bandgap and luminescence stem from α-Fe2O3 and weak ferromagnetism arises from structural defects and surface effects. With a 74% photocatalytic efficiency, stable through three photodegradation cycles, BFDW outperforms comparable waste materials in MB degradation mediated by visible light. The elemental trapping experiment exposed hydroxyl radicals (OH•) and superoxide anions (O2-•) as the primary species in the photodegradation process. Consequently, iron oxide-based BFDW emerges as an environmentally friendly alternative for wastewater treatment, underscoring the pivotal role of its unique physical properties in the photocatalytic process.

Changes in WNT signaling-related gene expression associated with development and cloning in bovine extra-embryonic and endometrial tissues during the peri-implantation period.

We determined if somatic cell nuclear transfer (SCNT) cloning is associated with WNT-related gene expression in cattle development, and if the expression of genes in the WNT pathway changes during the peri-implantation period. Extra-embryonic and endometrial tissues were collected at gestation days 18 and 34 (d18, d34). WNT5A, FZD4, FZD5, LRP5, CTNNB1, GNAI2, KDM1A, BCL2L1, and SFRP1 transcripts were localized in extra-embryonic tissue, whereas SFRP1 and DKK1 were localized in the endometrium. There were no differences in the localization of these transcripts in extra-embryonic tissue or endometrium from SCNT or artificial insemination (AI) pregnancies. Expression levels of WNT5A were 11-fold greater in the allantois of SCNT than AI samples. In the trophoblast, expression of WNT5A, FZD5, CTNNB1, and DKK1 increased significantly from d18 to d34, whereas expression of KDM1A and SFRP1 decreased, indicating that implantation is associated with major changes in WNT signaling. SCNT was associated with altered WNT5A expression in trophoblasts, with levels increasing 2.3-fold more in AI than SCNT conceptuses from d18 to d34. In the allantois, expression of WNT5A increased 6.3-fold more in SCNT than AI conceptuses from d18 to d34. Endometrial tissue expression levels of the genes tested did not differ between AI or SCNT pregnancies, although expression of individual genes showed variation across developmental stages. Our results demonstrate that SCNT is associated with altered expression of specific WNT-related genes in extra-embryonic tissue in a time- and tissue-specific manner. The pattern of gene expression in the WNT pathway suggests that noncanonical WNT signal transduction is important for implantation of cattle conceptuses.

Plasminogen binding proteins and plasmin generation on the surface of Leptospira spp.: the contribution to the bacteria-host interactions.

Leptospirosis is considered a neglected infectious disease of human and veterinary concern. Although extensive investigations on host-pathogen interactions have been pursued by several research groups, mechanisms of infection, invasion and persistence of pathogenic Leptospira spp. remain to be elucidated. We have reported the ability of leptospires to bind human plasminogen (PLG) and to generate enzimatically active plasmin (PLA) on the bacteria surface. PLA-coated Leptospira can degrade immobilized ECM molecules, an activity with implications in host tissue penetration. Moreover, we have identified and characterized several proteins that may act as PLG-binding receptors, each of them competent to generate active plasmin. The PLA activity associated to the outer surface of Leptospira could hamper the host immune attack by conferring the bacteria some benefit during infection. The PLA-coated leptospires obstruct complement C3b and IgG depositions on the bacterial surface, most probably through degradation. The decrease of leptospiral opsonization might be an important aspect of the immune evasion strategy. We believe that the presence of PLA on the leptospiral surface may (i) facilitate host tissue penetration, (ii) help the bacteria to evade the immune system and, as a consequence, (iii) permit Leptospira to reach secondary sites of infection.

Malaria primes the innate immune response due to interferon-gamma induced enhancement of toll-like receptor expression and function.

Malaria-induced sepsis is associated with an intense proinflammatory cytokinemia for which the underlying mechanisms are poorly understood. It has been demonstrated that experimental infection of humans with Plasmodium falciparum primes Toll-like receptor (TLR)-mediated proinflammatory responses. Nevertheless, the relevance of this phenomenon during natural infection and, more importantly, the mechanisms by which malaria mediates TLR hyperresponsiveness are unclear. Here we show that TLR responses are boosted in febrile patients during natural infection with P. falciparum. Microarray analyses demonstrated that an extraordinary percentage of the up-regulated genes, including genes involving TLR signaling, had sites for IFN-inducible transcription factors. To further define the mechanism involved in malaria-mediated "priming," we infected mice with Plasmodium chabaudi. The human data were remarkably predictive of what we observed in the rodent malaria model. Malaria-induced priming of TLR responses correlated with increased expression of TLR mRNA in a TLR9-, MyD88-, and IFNgamma-dependent manner. Acutely infected WT mice were highly susceptible to LPS-induced lethality while TLR9(-/-), IL12(-/-) and to a greater extent, IFNgamma(-/-) mice were protected. Our data provide unprecedented evidence that TLR9 and MyD88 are essential to initiate IL12 and IFNgamma responses and favor host hyperresponsiveness to TLR agonists resulting in overproduction of proinflammatory cytokines and the sepsis-like symptoms of acute malaria.

[Thoughts regarding researchers utilizing Grounded Theory]. / Reflexões sobre o pesquisador nas trilhas da Teoria Fundamentada nos Dados.

This descriptive-reflexive study was performed with the objective to present the characteristics of researchers who use the Grounded Theory method, and outline the development of aptitudes for the researcher to become a Grounded Theoretician. The theoretical discussion was based on the frameworks of this methodology and supported by the literature. The article presents the main demands of qualitative studies using Grounded Theory, and important behaviors, attitudes and characteristics developed by the researchers. It is concluded that learning about Grounded Theory involves more than operationalizing a group of procedures and techniques. It also involves facing challenges to change one's attitude as a researcher and develop new ways of thinking and researching, gathering knowledge based on data to form a theory.

Magnetic Zinc Oxide/Manganese Ferrite Composite for Photodegradation of the Antibiotic Rifampicin.

In this study, a composite of zinc oxide and manganese ferrite was synthesized using co-precipitation and hydrothermal routes, to be used as photocatalysts in reactions with UV/Vis light source. The synthesized materials were characterized by FTIR, XRD, and SEM, where it was possible to verify the efficiency of the syntheses performed, through the identification of the resulting phases, the evaluation of the structural morphology of the particles, and the analysis of the detachments of the main vibration bonds present in these materials. The composite ZnO/MnFe2O4 was used in photodegradation reactions of the antibiotic rifampicin, with catalyst dosage of 0.20; 0.40, and 0.60 g and 10 ppm of rifampicin, reactions using pure ZnO as a catalyst were also performed as a comparative parameter of the influence of MnFe2O4 in this system. The composite ZnO/MnFe2O4 showed a maximum percentage of rifampicin decontamination of 94.72% and ZnO, 74.20%using 0.20 g of photocatalyst after 90 min, which indicates a positive influence on this process. The solution treated with ZnO/MnFe2O4 was subjected to magnetic field induction for attraction and consequently accelerated removal of the solids present, successfully, compacting for the application of ZnO/MnFe2O4 to be presented as a promising material for decontamination of emerging pollutants through photocatalytic reactions.

Evaluation of the Photocatalytic Activity of Distinctive-Shaped ZnO Nanocrystals Synthesized Using Latex of Different Plants Native to the Amazon Rainforest.

ZnO nanocrystals with three different morphologies have been synthesized via a simple sol-gel-based method using Brosimum parinarioides (bitter Amapá) and Parahancornia amapa (sweet Amapá) latex as chelating agents. X-ray diffraction (XRD) and electron diffraction patterns (SAED) patterns showed the ZnO nanocrystals were a pure hexagonal wurtzite phase of ZnO. XRD-based spherical harmonics predictions and HRTEM images depicted that the nanocrystallites constitute pitanga-like (~15.8 nm), teetotum-like (~16.8 nm), and cambuci-like (~22.2 nm) shapes for the samples synthesized using bitter Amapá, sweet Amapá, and bitter/sweet Amapá chelating agent, respectively. The band gap luminescence was observed at ~2.67-2.79 eV along with several structural defect-related, blue emissions at 468-474 nm (VO, VZn, Zni), green emissions positioned at 513.89-515.89 (h-VO+), and orange emission at 600.78 nm (VO+-VO++). The best MB dye removal efficiency (85%) was mainly ascribed to the unique shape and oxygen vacancy defects found in the teetotum-like ZnO nanocrystals. Thus, the bitter Amapá and sweet Amapá latex are effective chelating agents for synthesizing distinctive-shaped ZnO nanocrystals with highly defective and remarkable photocatalytic activity.

Multisite haplotype on cattle chromosome 3 is associated with quantitative trait locus effects on lactation traits.

The goal of this study was to identify candidate genes and DNA polymorphisms for quantitative trait loci (QTL) affecting milk yield (MY), fat yield (FY), and protein yield (PY) previously mapped to bovine chromosome 3 (BTA3). To accomplish this, 373 half-siblings sired by three bulls previously shown to be segregating for lactation trait QTL, and 263 additional sires in the U.S. Dairy Bull DNA Repository (DBDR) were genotyped for 2,500 SNPs within a 16.3 Mbp QTL critical region on BTA3. Targeted resequencing of ∼1.8 Mbp within the QTL critical region of one of the QTL heterozygous sires identified additional polymorphisms useful for association studies. Twenty-three single nucleotide polymorphisms (SNPs) within a fine-mapped region were associated with effects on breeding values for MY, FY, or PY in DBDR sires, of which five SNPs were in strong linkage disequilibrium in the population. This multisite haplotype included SNPs located within exons or promoters of four tightly linked genes: RAP1A, ADORA3, OVGP1, and C3H1orf88. An SNP within RAP1A showed strong evidence of a recent selective sweep based on integrated haplotype score and was also associated with breeding value for PY. Because of its known function in alveolar lumen formation in the mammary gland, RAP1A is thus a strong candidate gene for QTL effects on lactation traits. Our results provide a detailed assessment of a QTL region that will be a useful guide for complex traits analysis in humans and other noninbred species.

Challenges of nursing care in psychosocial rehabilitation: a study from the constructionist perspective.

OBJECTIVE: To identify the nursing staff's difficulties in providing care in psychosocial rehabilitation to CAPS users. METHODS: Qualitative study based on constructionism. Sixteen members of the CAPS nursing staff participated in the study in the city of Rio de Janeiro. The data collected in the interviews and observations were organized in Nvivo software and analyzed based on thematic content. RESULTS: The difficulties identified were: the territorial violence that imposes a silence to the actions of CAPS, low user education as a barrier to protagonism, impaired family adhesion, the technical-conceptual discomfort of nursing in acting in the crisis and material and human of CAPS dismantlement. FINAL CONSIDERATIONS: The challenges consist of overcoming staff exhaustion; however, they do not prevent nursing from betting on the possibility of protagonist care. Therefore, such care is dimensioned by the clinical-political-ethical relation in constant negotiation with the territory.

Interleukin-4 regulates the expression of CD209 and subsequent uptake of Mycobacterium leprae by Schwann cells in human leprosy.

The ability of microbial pathogens to target specific cell types is a key aspect of the pathogenesis of infectious disease. Mycobacterium leprae, by infecting Schwann cells, contributes to nerve injury in patients with leprosy. Here, we investigated mechanisms of host-pathogen interaction in the peripheral nerve lesions of leprosy. We found that the expression of the C-type lectin, CD209, known to be expressed on tissue macrophages and to mediate the uptake of M. leprae, was present on Schwann cells, colocalizing with the Schwann cell marker, CNPase (2',3'-cyclic nucleotide 3'-phosphodiesterase), along with the M. leprae antigen PGL-1 in the peripheral nerve biopsy specimens. In vitro, human CD209-positive Schwann cells, both from primary cultures and a long-term line, have a higher binding of M. leprae compared to CD209-negative Schwann cells. Interleukin-4, known to be expressed in skin lesions from multibacillary patients, increased CD209 expression on human Schwann cells and subsequent Schwann cell binding to M. leprae, whereas Th1 cytokines did not induce CD209 expression on these cells. Therefore, the regulated expression of CD209 represents a common mechanism by which Schwann cells and macrophages bind and take up M. leprae, contributing to the pathogenesis of leprosy.

Nursing and the leading role of the user in the CAPS: a study from the constructionist perspective.

OBJECTIVE: To analyze the actions of the Nursing team to promote the leading role of the patient in the Psychosocial Rehabilitation Center. METHOD: A qualitative study, with reference to Social Constructionism. The study was conducted with 16 members of the Nursing team in two CAPS in the city of Rio de Janeiro from September 2017 to January 2018. The data collected from interviews and observation were organized in the Nvivo Software and the thematic content was analyzed. RESULTS: The study analyzed leading care. This is constructed through creative communication, networking and the perception of the Nursing team as an "antenna". FINAL CONSIDERATIONS: The Nursing team does not reduce the users to the impossibilities of their psychiatric diagnoses, it uses creative communication and the construction of support networks in the territory. It constitutes itself as an antenna in care in psychosocial rehabilitation.

Detection of hepatitis C virus in platelets: evaluating its relationship to antiviral therapy outcome.

BACKGROUND/AIMS: Detection of HCV has been documented in extrahepatic sites such as platelets. However, its influence on antiviral therapy outcome is unknown. In this study, we investigated the relationship between the detection of HCV in platelets from a cohort of 48 chronically HCV-infected patients and response to antiviral therapy. METHODOLOGY: This study comprised of 19 males and 29 females, mean age 54.9 +/- 8.72 years, followed-up in Rio de Janeiro, Brazil, between August 2004 and October 2006. HCV-RNA was detected in serum and platelets (pre-treatment, end-of-treatment and 24 weeks after completion of therapy) by reverse transcription-nested polymerase chain reaction. Patients with genotype 1 or 4 were treated with peginterferon-alfa/ribavirin for 48 weeks, and patients with genotype 3 received interferon-alfa/ribavirin for 24 weeks. RESULTS: Baseline detection of HCV in platelets was found not to be related to therapy outcome. However, significant associations between detection rates of HCV in platelets and serum at the end-of-treatment (p = 0.0203), and 24 weeks after completion of therapy (p = 0.0016) were observed. Interestingly, HCV was detected in platelets from two patients with normal ALT who lost detectable serum HCV at the end-of-treatment and, after 24 weeks of followup, relapsed virologically in serum. CONCLUSIONS: Our data suggest that patients with HCV persistence in platelets by the end-of-treatment appear to be at an increased risk of recurrent HCV infection.

Aberrant gene expression patterns in placentomes are associated with phenotypically normal and abnormal cattle cloned by somatic cell nuclear transfer.

Transcription profiling of placentomes derived from somatic cell nuclear transfer (SCNT, n = 20), in vitro fertilization (IVF, n = 9), and artificial insemination (AI, n = 9) at or near term development was performed to better understand why SCNT and IVF often result in placental defects, hydrops, and large offspring syndrome (LOS). Multivariate analysis of variance was used to distinguish the effects of SCNT, IVF, and AI on gene expression, taking into account the effects of parturition (term or preterm), sex of fetus, breed of dam, breed of fetus, and pathological finding in the offspring (hydrops, normal, or other abnormalities). Differential expression of 20 physiologically important genes was confirmed with quantitative PCR. The largest effect on placentome gene expression was attributable to whether placentas were collected at term or preterm (i.e., whether the collection was because of disease or to obtain stage-matched controls) followed by placentome source (AI, IVF, or SCNT). Gene expression in SCNT placentomes was dramatically different from AI (n = 336 genes; 276 >2-fold) and from IVF (n = 733 genes; 162 >2-fold) placentomes. Functional analysis of differentially expressed genes (DEG) showed that IVF has significant effects on genes associated with cellular metabolism. In contrast, DEG associated with SCNT are involved in multiple pathways, including cell cycle, cell death, and gene expression. Many DEG were shared between the gene lists for IVF and SCNT comparisons, suggesting that common pathways are affected by the embryo culture methods used for IVF and SCNT. However, the many unique gene functions and pathways affected by SCNT suggest that cloned fetuses may be starved and accumulating toxic wastes due to placental insufficiency caused by reprogramming errors. Many of these genes are candidates for hydrops and LOS.

Molecular analysis and patterns of ALT and hepatitis C virus seroconversion in haemodialysis patients with acute hepatitis.

BACKGROUND: Haemodialysis (HD) continues to carry the risk of hepatitis C virus (HCV) transmission, with delayed seroconversion and often normal alanine aminotransferase (ALT) values increasing the likelihood of undetected infection and thus uninterrupted spread of HCV. The aim of this study was to identify the characteristic patterns of ALT changes and seroconversion during an outbreak of HCV in a HD unit. We also wanted to establish the relationship between infecting viruses using molecular analysis. METHODS: All patients (n = 72) and staff (n = 23) of the HD unit were prospectively followed for 14 months. Serial measurements for ALT, HCV antibody and HCV-RNA were performed besides HCV sequence analysis. RESULTS: The initial screening for anti-HCV and HCV-RNA confirmed chronic infection in 16/72 (22%) subjects and identified three subjects with recent seroconversion. In addition, five cases were reverse transcription-polymerase chain reaction positive alone for a total of eight recent cases. The interval between the initial observation of ALT changes and seroconversion varied from 1 to 8 months, and in several individuals ALT fluctuations only below the upper limit of normal were detected. However, relating each subject's ALT values to ALT at baseline, ALT levels increased between 1.6- and 4.7-fold. Molecular analysis provided evidence for transmission from two chronically infected source patients, probably because of inappropriate infection control measures. CONCLUSION: Our data highlight the importance of well-implemented safety precautions and regular HCV-RNA testing to prevent the further spread of HCV in this population, and suggest the use of ALT baseline values to identify infections that may remain unnoticed otherwise.