Investigation of neuro-inflammatory parameters in a cuprizone induced mouse model of multiple sclerosis

Cuprizone, copper chelator, treatment of mouse is a toxic model of multiple sclerosis (MS) in which oligodendrocyte death, demyelination and remyelination can be observed. Understanding T and B cell subset as well as their cytokines involved in MS pathogenesis still requires further scrutiny to better understand immune component of MS. The study presented here, aimed to evaluate relevant cytokines, lymphocytes, and gene expressions profiles during demyelination and remyelination in the cuprizone mouse model of MS. Eighty male C57BL/6J mice fed with 0.2% cuprizone for eight weeks. Cuprizone has been removed from the diet in the following eight weeks. Cuprizone treated and control mice sacrificed biweekly, and corpus callosum of the brain was investigated by staining. Lymphocyte cells of mice analyzed by flow cytometry with CD3e, CD11b, CD19, CD80, CD86, CD4, CD25 and FOXP3 antibodies. IFN-gamma, IL-1alpha, IL-2, IL-5, IL-6, IL-10, IL-17, TNF-alpha cytokines were analyzed in plasma samples. Neuregulin 1 (Nrg1), ciliary neurotrophic factor (Cntf) and C-X-C chemokine receptor type 4 (Cxcr4) gene expressions in corpus callosum sections of the mice brain were quantified. Histochemistry analysis showed that demyelination began at the fourth week of cuprizone administration and total demyelination occurred at the twelfth week in chronic model. Remyelination occurred at the fourth week of following withdrawal of cuprizone from diet. The level of mature and activated T cells, regulatory T cells, T helper cells and mature B cells increased during demyelination and decreased when cuprizone removed from diet. Further, both type 1 and type 2 cytokines together with the proinflammatory cytokines increased. The level of oligodendrocyte maturation and survival genes showed differential gene expression in parallel to that of demyelination and remyelination. In conclusion, for the first-time, involvement of both cellular immune response and antibody response as well as oligodendrocyte maturation and survival factors having role in demyelination and remyelination of cuprizone mouse model of MS have been shown.     

Flavonol‐mediated stabilization of PIN efflux complexes regulates polar auxin transport

The transport of auxin controls the rate, direction and localization of plant growth and development. The course of auxin transport is defined by the polar subcellular localization of the PIN proteins, a family of auxin efflux transporters. However, little is known about the composition and regulation of the PIN protein complex. Here, using blue‐native PAGE and quantitative mass spectrometry, we identify native PIN core transport units as homo‐ and heteromers assembled from PIN1, PIN2, PIN3, PIN4 and PIN7 subunits only. Furthermore, we show that endogenous flavonols stabilize PIN dimers to regulate auxin efflux in the same way as does the auxin transport inhibitor 1‐naphthylphthalamic acid (NPA). This inhibitory mechanism is counteracted both by the natural auxin indole‐3‐acetic acid and by phosphomimetic amino acids introduced into the PIN1 cytoplasmic domain. Our results lend mechanistic insights into an endogenous control mechanism which regulates PIN function and opens the way for a deeper understanding of the protein environment and regulation of the polar auxin transport complex.     

Triton channels are sensitive to divalent cations and protons

Addition of Triton X-100 to planar bilayers composed of dioleoyl phosphatidyl choline, diphytanoyl phosphatidyl choline or mono-oleoyl glycerol induces single channel-like events when electrical conductivity across the bilayer is measured. Addition of divalent cations or protons causes channels to disappear; single channel conductance of remaining channels is not significantly altered; addition of EDTA or alkali (respectively) reverses the effect. It is concluded that sensitivity to divalent cations and protons need not be dependent on specific channel proteins or pore-forming toxins, but may be a feature of any aqueous pore across a lipid milieu.We are grateful to Dr. D.T. Edmonds and Prof. R.J.P. Williams for critical discussion, to Glenn Alder for technical assistance, to Ms. B. Bashford and Ms. S.G. Pelc for preparing the paper, and to the Cell Surface Research Fund, the Royal Society (A.A.L.), UNESCO (Molecular and Cell Biology Network) and The Wellcome Trust for financial support.     

Virus, toxin, complement: common actions and their prevention by Ca2+ or Zn2+

Membrane damage induced by haemolytic agents does not necessarily lead to lysis: the pores that are formed at low concentration of agent are formed at low concentration of agent are not large enough to allow leakage of cytoplasmic proteins, and in many instances the lesions become repaired with time. Quite different agents induce a similar type of lesion: in each case leakage is reduced at low ionic strenth, and is prevented by divalent cations such as Ca²⁺ or Zn²⁺, suggesting a possible therapeutic approach to the containment of several membrane-damaging diseases.     

Peripheral orphanin FQ/nociceptin analgesia in the mouse.

Orphanin FQ/Nociceptin (OFQ/N) administered peripherally was an effective analgesic in the tailflick test in mice (ED50 16.3 microg). It had a peak effect at 5 min and lasted up to 30 min. The kappa3 analgesic naloxone benzoylhydrazone was also active peripherally (ED50 3.8 microg). The analgesic actions of both agents were blocked by naloxone. Neither OFQ/N(1-11) nor OFQ/N(1-7) had appreciable peripheral activity. Antisense mapping both compounds against the murine orphan opioid receptor (KOR-3) confirmed the importance of this clone in their actions. Antisense probes targeting the second and third coding exons significantly lowered the analgesic effects of both compounds. However, the antisense targeting the first coding exon blocked only the actions of OFQ/N and not kappa3 analgesia.     

Lymphocyte culture supernatants prevent sickling of red blood cells

Metabisulphate-treated red blood cells from sickle-cell patients were protected from sickling by exposure to supernatants from cultures of phytohemagglutinin-stimulated normal human lymphocytes. Such supernatants, when not thus stimulated, afforded no protection.     

Dehydroascorbate and glutathione regulate the cellular development of <Emphasis Type="Italic">Nicotiana tabacum</Emphasis> L. SR-1 protoplasts

A Nicotiana tabacum L. SR-1 leaf protoplast system was used to study the effects of dehydroascorbate and glutathione on cellular development. We found that dehydroascorbate is readily taken up by protoplasts and internally reduced to ascorbate. Concomitantly, cell division was inhibited and cell expansion stimulated. In this respect, dehydroascorbate counteracted auxin-mediated leaf protoplast development. In contrast to the effects of dehydroascorbate, glutathione-induced cell dedifferentiation, and this effect is similar to that mediated by high auxin concentrations. We conclude that dehydroascorbate and glutathione affect the auxin-mediated regulation of cellular development. Therefore, the biological role of these compounds extends beyond stress tolerance and defense.