Somatic cell nuclear transfer is associated with altered expression of angiogenic factor systems in bovine placentomes at term

Low efficiency of somatic cell cloning by nuclear transfer has been associated with alterations of placental vascular architecture. Placental growth and function depend on the growth of blood vessels; VEGF-A and bFGF are the most important factors controlling neovascularization and vascular permeability in the placenta. We hypothesize that the VEGF-A and bFGF systems are disrupted in placentomes from cloned animals, contributing to the placental abnormalities that are common in these clones. We determined mRNA expression and protein tissue localization of VEGF-A, bFGF, and their receptors in placentomes from cloned and non-cloned bovine fetuses at term. Real-time RT-PCR revealed that VEGFR-2 mRNA was increased in cloned male-derived placentomes, while mRNA of bFGF and its receptors were decreased in placentomes of cloned females. VEGF-A system proteins were found to be located in placentomal endothelial, maternal and fetal epithelial and stromal cells; there was a variable pattern of cellular distribution of these proteins in both cloned and non-cloned animals. Alterations in the expression of VEGF-A and bFGF systems suggest that angiogenic factors are involved in abnormal placental development in cloned gestations, contributing to impaired fetal development and poor survival ratesFAPESP 02/07392-7CAPES (PROBRAL grant 272/7)\ud CAPES (PROBRAL grant D/06/33937

Crosstalk between Neospora caninum and the bovine host at the maternal-foetal interface determines the outcome of infection

19 páginas, 9 figuras, 1 tabla.Neospora caninum is an apicomplexan cyst-forming parasite that is considered one of the main causes of abortion. The pathogenic mechanisms associated with parasite virulence at the maternal-foetal interface that are responsible for the outcome of infection are largely unknown. Here, utilizing placentomes from cattle experimentally infected with high-virulence (Nc-Spain7) and low-virulence (Nc-Spain1H) isolates, we studied key elements of the innate and adaptive immune responses, as well as components of the extracellular matrix (ECM), at 10 and 20 days post-infection (dpi). The low-virulence isolate elicited a robust immune response characterized by upregulation of genes involved in pathogen recognition, chemokines and pro-inflammatory cytokines, crucial for its adequate control. In addition, Nc-Spain1H triggered the expression of anti-inflammatory cytokines and other mechanisms implicated in the maintenance of ECM integrity to ensure foetal survival. In contrast, local immune responses were initially (10 dpi) impaired by Nc-Spain7, allowing parasite multiplication. Subsequently (20 dpi), a predominantly pro-inflammatory Th1-based response and an increase in leucocyte infiltration were observed. Moreover, Nc-Spain7-infected placentomes from animals carrying non-viable foetuses exhibited higher expression of the IL-8, TNF-α, iNOS and SERP-1 genes and lower expression of the metalloproteases and their inhibitors than Nc-Spain7-infected placentomes from animals carrying viable foetuses. In addition, profound placental damage characterized by an alteration in the ECM organization in necrotic foci, which could contribute to foetal death, was found. Two different host-parasite interaction patterns were observed at the bovine placenta as representative examples of different evolutionary strategies used by this parasite for transmission to offspring.This work was supported by the Spanish Ministry of Economy and Competi‑ tiveness (AGL2013-44694-R and AGL2016-75935-C2-1-R) and the Community of Madrid (PLATESA2-CM P2018/BAA-4370). Laura Jiménez-Pelayo was fnan‑ cially supported by a fellowship from the University Complutense of Madrid (including two research stays in 2017 and 2018) and Marta García-Sánchez was fnancially supported through a grant from the Spanish Ministry of Economy and Competitiveness (BES-2014-070723). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewe

Two novel connexin32 mutations cause early onset X-linked Charcot-Marie-Tooth disease

<p>Abstract</p> <p>Background</p> <p>X-linked Charcot-Marie Tooth (CMT) is caused by mutations in the connexin32 gene that encodes a polypeptide which is arranged in hexameric array and form gap junctions.</p> <p>Methods</p> <p>We describe two novel mutations in the connexin32 gene in two Norwegian families.</p> <p>Results</p> <p>Family 1 had a c.225delG (R75fsX83) which causes a frameshift and premature stop codon at position 247. This probably results in a shorter non-functional protein structure. Affected individuals had an early age at onset usually in the first decade. The symptoms were more severe in men than women. All had severe muscle weakness in the legs. Several abortions were observed in this family. Family 2 had a c.536 G>A (C179Y) transition which causes a change of the highly conserved cysteine residue, i.e. disruption of at least one of three disulfide bridges. The mean age at onset was in the first decade. Muscle wasting was severe and correlated with muscle weakness in legs. The men and one woman also had symptom from their hands.</p> <p>The neuropathy is demyelinating and the nerve conduction velocities were in the intermediate range (25–49 m/s). Affected individuals had symmetrical clinical findings, while the neurophysiology revealed minor asymmetrical findings in nerve conduction velocity in 6 of 10 affected individuals.</p> <p>Conclusion</p> <p>The two novel mutations in the connexin32 gene are more severe than the majority of previously described mutations possibly due to the severe structural change of the gap junction they encode.</p

Immune response of healthy horses to DNA constructs formulated with a cationic lipid transfection reagent

Background Deoxyribonucleic acid (DNA) vaccines are used for experimental immunotherapy of equine melanoma. The injection of complexed linear DNA encoding interleukin (IL)-12/IL-18 induced partial tumour remission in a clinical study including 27 grey horses. To date, the detailed mechanism of the anti-tumour effect of this treatment is unknown. Results In the present study, the clinical and cellular responses of 24 healthy horses were monitored over 72 h after simultaneous intradermal and intramuscular application of equine IL-12/IL-18 DNA (complexed with a transfection reagent) or comparative substances (transfection reagent only, nonsense DNA, nonsense DNA depleted of CG). Although the strongest effect was observed in horses treated with expressing DNA, horses in all groups treated with DNA showed systemic responses. In these horses treated with DNA, rectal temperatures were elevated after treatment and serum amyloid A increased. Total leukocyte and neutrophil counts increased, while lymphocyte numbers decreased. The secretion of tumour necrosis factor alpha (TNFα) and interferon gamma (IFNγ) from peripheral mononuclear blood cells ex vivo increased after treatments with DNA, while IL-10 secretion decreased. Horses treated with DNA had significantly higher myeloid cell numbers and chemokine (C-X-C motif) ligand (CXCL)-10 expression in skin samples at the intradermal injection sites compared to horses treated with transfection reagent only, suggesting an inflammatory response to DNA treatment. In horses treated with expressing DNA, however, local CXCL-10 expression was highest and immunohistochemistry revealed more intradermal IL-12-positive cells when compared to the other treatment groups. In contrast to non-grey horses, grey horses showed fewer effects of DNA treatments on blood lymphocyte counts, TNFα secretion and myeloid cell infiltration in the dermis. Conclusion Treatment with complexed linear DNA constructs induced an inflammatory response independent of the coding sequence and of CG motif content. Expressing IL-12/IL-18 DNA locally induces expression of the downstream mediator CXCL-10. The grey horses included appeared to display an attenuated immune response to DNA treatment, although grey horses bearing melanoma responded to this treatment with moderate tumour remission in a preceding study. Whether the different immunological reactivity compared to other horses may contributes to the melanoma susceptibility of grey horses remains to be elucidated

How firms respond to financial restatement: CEO successors and external reactions

Although past studies have paid considerable attention to firms' reputations, few have investigated the actions that firms take following a reputation-damaging event. We identify firms involved in financial earnings restatements and examine whether naming a successor CEO with specific qualities serves to signal the seriousness of a firm's efforts to restore its reputation. Using theories of market signaling, we argue that attributes of successor CEOs significantly influence the reactions of key external constituencies. In particular, firms with more severe restatement tend to name successors who have prior CEO or turnaround experience and a more elite education. The naming of such successors results in more positive reactions from the stock market, financial analysts, and mass media. We argue that these attributes send messages to stakeholders and the broader public about the CEO's credibility and the firm's efforts