Cytoskeletal changes during neurogenesis in cultures of avian neural crest cells

Summary Neural crest cells are motile and mitotic, whereas their neuronal derivatives are terminally post-mitotic and consist of stationary cell body from which processes grow. The present study documents changes in the cytoskeleton that occur during neurogenesis in cultures of avian neural crest cells. The undifferentiated neural crest cells contain dense bundles of actin filaments throughout their cytoplasm, and a splayed array of microtubules attached to the centrosome. In newly differentiating neurons, the actin bundles are disrupted and most of the remaining actin filaments are reorganized into a cortical layer underlying the plasma membrane of the cell body and processes. Microtubules are more abundant in newly-differentiating neurons than in the undifferentiated cells, and individual microtubules can be seen dissociated from the centrosome. Neuron-specific -III tubulin appears in some crest cells prior to cessation of motility and cell division, and expression increases with total microtubule levels during neurogenesis. To investigate how these early cytoskeletal changes might contribute to alterations in morphology during neurogenesis, we have disrupted the cytoskeleton with pharmacologic agents. Microfilament disruption by cytochalasin immediately arrests the movement of neural crest cells and causes them to round-up, but does not significantly change the morphology of the immature neurons. Microtubule depolymerization by nocodazole slows the movement of undifferentiated cells and causes retraction of processes extended by the immature neurons. These results suggest that changes in the actin and microtubule arrays within neural crest cells govern distinct aspects of their morphogenesis into neurons.     

Alpha 1-adrenergic effects and liver regeneration

The effects of several treatments involving alpha-adrenergic mechanisms upon the early stages of rat liver regeneration were examined. Catecholamine concentrations in rat plasma were measured at various times after hepatectomy and were found to be elevated relative to those in plasma from sham-operated rats. Surgical hepatic denervation or injection of an alpha 1-adrenergic receptor antagonist (prazosin) reduced incorporation of [3H]thymidine into liver DNA during the first 24 hr after partial hepatectomy. Chronic guanethidine injections (3 to 6 weeks) reduced liver catecholamine levels, but did not affect its ability to regenerate. The inhibition of regenerative DNA synthesis by prazosin was preceded by an alteration in the binding of epidermal growth factor to regenerating liver, which was apparently the result of an increased number of epidermal growth factor receptors. Thus, alpha 1-adrenergic blockade, which affects both epidermal growth factor receptor binding and subsequent DNA synthesis in hepatocyte primary cultures, can also modulate these processes during liver regeneration in vivo.     

Neurological Manifestations of COVID-19 Feature T Cell Exhaustion and Dedifferentiated Monocytes in Cerebrospinal Fluid

1. Download : Download high-res image (177KB)
2. Download : Download full-size image

     

Zur Kenntniss der Fohlenlähme

Ohne Zusammenfassung     

Immunization with apoptotic pseudovirus transduced cells induces both cellular and humoral responses: a proof of concept study in macaques

Dendritic cells are able to present viral antigens to T-cells after uptake of apoptotic bodies derived from virus-infected cells. Immunization with virus-infected apoptotic cells was previously shown to induce HIV-specific immune responses in mice. Here we evaluate the safety and immunogenicity of immunization with activated apoptotic cells in non-human primates using autologous T-cells infected with replication defective VSV pseudotyped SIV(mac239)Δenv. Animals were immunized with γ-irradiated activated T-cells carrying the VSVenvSIV(mac239)Δenv pseudovirus. SIV Gag-specific cellular immune responses were induced as early as two weeks after the first immunization eliciting a biased IFN-γ and IL-2 response. In addition, induction of SIV Gag-specific antibody responses and high titer neutralizing activity against the SIV pseudovirus harboring a VSV-env were detected after two immunizations. The vaccinated group and a control group of Chinese rhesus macaques were intravenously challenged with pathogenic SIV(mac251.) All animals became infected, but SIV-replication was effectively suppressed (below 100 copies/ml) in several animals in both groups. However the group immunized with apoptotic cells revealed better preservation of the gut CD4(+) T-cell compartment. Viral control was inversely correlated with an early (4 weeks) but transient increase in the percentage of Ki67(+)CD4(+) peripheral blood T-cells (Spearman -0.73). We here show that immunizations with activated apoptotic lymphocytes expressing transduced SIV genes result in induction of both cellular and humoral immune responses. This study provides evidence for an immunological principle demonstrating that certain apoptotic cells can be considered as carriers of antigens directing immune responses in macaques.     

Metabolism of low-dose inorganic arsenic in a central European population: influence of sex and genetic polymorphisms

BACKGROUND: There is a wide variation in susceptibility to health effects of arsenic, which, in part, may be due to differences in arsenic metabolism. Arsenic is metabolized by reduction and methylation reactions, catalyzed by reductases and methyltransferases. OBJECTIVES: Our goal in this study was to elucidate the influence of various demographic and genetic factors on the metabolism of arsenic. METHODS: We studied 415 individuals from Hungary, Romania, and Slovakia by measuring arsenic metabolites in urine using liquid chromatography with hydride generation and inductively coupled plasma mass spectrometry (HPLC-HG-ICPMS). We performed genotyping of arsenic (+III) methyltransferase (AS3MT), glutathione S-transferase omega 1 (GSTO1), and methylene-tetrahydrofolate reductase (MTHFR). RESULTS: The results show that the M287T (T-->C) polymorphism in the AS3MT gene, the A222V (C-->T) polymorphism in the MTHFR gene, body mass index, and sex are major factors that influence arsenic metabolism in this population, with a median of 8.0 microg/L arsenic in urine. Females < 60 years of age had, in general, higher methylation efficiency than males, indicating an influence of sex steroids. That might also explain the observed better methylation in overweight or obese women, compared with normal weight men. The influence of the M287T (T-->C) polymorphism in the AS3MT gene on the methylation capacity was much more pronounced in men than in women. CONCLUSIONS: The factors investigated explained almost 20% of the variation seen in the metabolism of arsenic among men and only around 4% of the variation among women. The rest of the variation is probably explained by other methyltransferases backing up the methylation of arsenic.     

Making the choice: the translation of global HIV and infant feeding policy to local practice among mothers in Pune, India

In 2003, India had over 5.1 million infected individuals living with HIV/AIDS. The percentage of all HIV cases attributed to perinatal transmission has been increasing steadily from 0.33% of total cases in 1999 to 2.80% in 2004. Recent statistics indicate that over 130,000 infants have been infected through this route. Despite recent advances in reducing in utero and interpartum transmission with the use of antiretrovirals, there is a critical need to make infant feeding safer. Current UNAIDS/WHO/UNICEF recommendations stress avoidance of all breast-feeding if replacement feeding fulfills the key requirements of being affordable, feasible, acceptable, sustainable, and safe. In this paper, we examine how the UNAIDS/WHO/UNICEF recommendations have been actualized within the context of an urban government hospital in India. The documented patterns of infant feeding by HIV-positive mothers in Pune, India, from 2000 to 2004, highlight the complexities of making an informed and healthy choice under suboptimal conditions. The data indicate that interpersonal variations in the key requirements greatly influence the optimal practice to minimize mortality risks. Moreover, local information on health outcomes is crucial to tailoring policy recommendations to save lives. We propose the development of a decision-making algorithm that includes factors affecting mother-to-infant transmission, including site-specific data on health risks to the mother and the child. Such an algorithm would allow identification of the healthiest feeding choice and would minimize the pitfalls of promoting homogeneous practices lacking site-specific evidence-based evaluation.     

Information and communication technology and community-based health sciences training in Uganda: perceptions and experiences of educators and students

Information and communication technology (ICT) has been advocated as a powerful tool for improving health education in low-resource settings. However, few evaluations have been performed of ICT perceptions and user experiences in low-resource settings. During late 2009, an internet-based survey on ICT was administered to students, tutors, and faculty members associated with a Community-Based Education and Service (COBES) program in Uganda. 255 surveys were completed. Response rates varied (students, 188/684, 27.5%; tutors, 14/27, 51.9%; faculty, 53/335, 15.8%). Most respondents owned mobile phones (98%). Students were less likely (p < 0.001) to own laptops (25%) compared to tutors (71%) and faculty (85%). Internet access at rural sites was uncommon; mobile phone coverage was almost universally present. Laptop ownership and internet and mobile phone access was not associated with high valuation of students' COBES experiences. Free text responses found that respondents valued ICT access for research, learning, and communication purposes. In summary, ICT penetration in this population is primarily manifest by extensive mobile phone ownership. Internet access in rural educational sites is still lacking, but students and educators appear eager to utilize this resource if availability improves. ICT may offer a unique opportunity to improve the quality of teaching and learning for COBES participants.     

Deletion of thioredoxin-interacting protein preserves retinal neuronal function by preventing inflammation and vascular injury

BACKGROUND AND PURPOSE

Retinal neurodegeneration is an early and critical event in several diseases associated with blindness. Clinically, therapies that target neurodegeneration fail. We aimed to elucidate the multiple roles by which thioredoxin-interacting protein (TXNIP) contributes to initial and sustained retinal neurodegeneration.

EXPERIMENTAL APPROACH

Neurotoxicity was induced by intravitreal injection of NMDA into wild-type (WT) and TXNIP-knockout (TKO) mice. The expression of apoptotic and inflammatory markers was assessed by immunohistochemistry, elisa and Western blot. Microvascular degeneration was assessed by periodic acid-Schiff and haematoxylin staining and retinal function by electroretinogram.

KEY RESULTS

NMDA induced early (1 day) and significant retinal PARP activation, a threefold increase in TUNEL-positive nuclei and 40% neuronal loss in ganglion cell layer (GCL); and vascular permeability in WT but not TKO mice. NMDA induced glial activation, expression of TNF-α and IL-1β that co-localized with Müller cells in WT but not TKO mice. In parallel, NMDA triggered the expression of NOD-like receptor protein (NLRP3), activation of caspase-1, and release of IL-1β and TNF-α in primary WT but not TKO Müller cultures. After 14 days, NMDA induced 1.9-fold microvascular degeneration, 60% neuronal loss in GCL and increased TUNEL-labelled cells in the GCL and inner nuclear layer in WT but not TKO mice. Electroretinogram analysis showed more significant reductions in b-wave amplitudes in WT than in TKO mice.

CONCLUSION AND IMPLICATIONS

Targeting TXNIP expression prevented early retinal ganglion cell death, glial activation, retinal inflammation and secondary neuro/microvascular degeneration and preserved retinal function. TXNIP is a promising new therapeutic target for retinal neurodegenerative diseases.     

New insight into active muscarinic receptors with the novel radioagonist [H]iperoxo

Activation of G protein-coupled receptors involves major conformational changes of the receptor protein ranging from the extracellular transmitter binding site to the intracellular G protein binding surface. GPCRs such as the muscarinic acetylcholine receptors are commonly probed with radioantagonists rather than radioagonists due to better physicochemical stability, higher affinity, and indifference towards receptor coupling states of the former. Here we introduce tritiated iperoxo, a superagonist at muscarinic M₂ receptors with very high affinity. In membrane suspensions of transfected CHO-cells, [³H]iperoxo – unlike the common radioagonists [³H]acetylcholine and [³H]oxotremorine M – allowed labelling of each of the five muscarinic receptor subtypes in radioagonist displacement and saturation binding studies. [³H]iperoxo revealed considerable differences in affinity between the even- and the odd-numbered muscarinic receptor subtypes with affinities for the M₂ and M₄ receptor in the picomolar range. Probing ternary complex formation on the M₂ receptor, [³H]iperoxo dissociation was not influenced by an archetypal allosteric inverse agonist, reflecting activation-related rearrangement of the extracellular loop region. At the inner side of M₂, the preferred G_i protein acted as a positive allosteric modulator of [³H]iperoxo binding, whereas G_s and G_q were neutral in spite of their robust coupling to the activated receptor. In intact CHO-hM₂ cells, endogenous guanylnucleotides promoted receptor/G protein-dissociation resulting in low-affinity agonist binding which, nevertheless, was still reported by [³H]iperoxo. Taken together, the muscarinic superagonist [³H]iperoxo is the best tool currently available for direct probing activation-related conformational transitions of muscarinic receptors.