Vaginally Administered PEGylated LIF Antagonist Blocked Embryo Implantation and Eliminated Non-Target Effects on Bone in Mice

Female-controlled contraception/HIV prevention is critical to address health issues associated with gender inequality. Therefore, a contraceptive which can be administered in tandem with a microbicide to inhibit sexually transmitted infections, is desirable. Uterine leukemia inhibitory factor (LIF) is obligatory for blastocyst implantation in mice and associated with infertility in women. We aimed to determine whether a PEGylated LIF inhibitor (PEGLA) was an effective contraceptive following vaginal delivery and to identify non-uterine targets of PEGLA in mice

The therapeutic effect of LIF in EAE-associated axonal injury

© 2009 Estella AlexandrouAxonal degeneration is a major pathological feature of the central nervous system (CNS) inflammatory demyelinating disease multiple sclerosis (MS). This axonal degeneration has major consequences, as functional axonal regeneration in the CNS is largely absent. Cumulative axonal degeneration is the likely cause of the majority of progressive MS-related disability, and therefore, the need for novel neuroprotective therapies for MS exists. Experimental autoimmune encephalomyelitis (EAE), an animal model of MS pathology, also produces axonal injury. In particular, the optic nerve and spinal cord are key sites of neuroinflammation in mouse EAE. By utilizing this model, the short term and long term effects of the putative neuroprotective cytokine, leukaemia inhibitory factor (LIF), were investigated in the optic nerve and spinal cord utilising a number of outcome measures of axonal dysfunction. These included MRI measures of water diffusivity along (ADC ||) and across (ADC┴) the optic nerves, serum levels of phosphorylated neurofilament heavy chain subunit (pNF-H) and histological morphometric measures. LIF treatment reduced EAE grade and pNF-H plasma levels, decreased ADC┴, but had no effect on ADC ||, axon counts or inflammatory infiltration. In contrast, genetic deletion of LIF and its sister cytokine ciliary neurotrophic factor (CNTF), not only increased EAE grade and pNF-H levels, but also decreased optic nerve ADC|| and optic nerve and spinal cord axon densities. After reviewing current literature, we hypothesize that the target cell for endogenously upregulated LIF in EAE may be the neuron or axon, whereas the target cell for exogenously administered therapeutic LIF may be another cell type, possibly infiltrating macrophages and activated microglial cells. LIF antagonist treatment did not have any affect on EAE grade, pNF-H levels or MRI parameters. This lack of effect may be due to the inability of the LIF antagonist to enter the CNS, supporting the hypothesis that endogenous LIF has a centrally acting mechanism

Journal of economics & management strategy : JEMS

Leukemia inhibitory factor (LIF) and Ciliary Neurotrophic factor (CNTF) are members of the interleukin-6 family of cytokines, defined by use of the gp130 molecule as an obligate receptor. In the murine experimental autoimmune encephalomyelitis (EAE) model, antagonism of LIF and genetic deletion of CNTF worsen disease. The potential mechanism of action of these cytokines in EAE is complex, as gp130 is expressed by all neural cells, and could involve immuno-modulation, reduction of oligodendrocyte injury, neuronal protection, or a combination of these actions. In this study we aim to investigate whether the beneficial effects of CNTF/LIF signalling in EAE are associated with axonal protection; and whether this requires signalling through oligodendrocytes. We induced MOG(35–55) EAE in CNTF, LIF and double knockout mice. On a CNTF null background, LIF knockout was associated with increased EAE severity (EAE grade 2.1±0.14 vs 2.6±0.19; P<0.05). These mice also showed increased axonal damage relative to LIF heterozygous mice, as indicated by decreased optic nerve parallel diffusivity on MRI (1540±207 µm(2)−/s vs 1310±175 µm(2)−/s; P<0.05), and optic nerve (−12.5%) and spinal cord (−16%) axon densities; and increased serum neurofilament-H levels (2.5 fold increase). No differences in inflammatory cell numbers or peripheral auto-immune T-cell priming were evident. Oligodendrocyte-targeted gp130 knockout mice showed that disruption of CNTF/LIF signalling in these cells has no effect on acute EAE severity. These studies demonstrate that endogenous CNTF and LIF act centrally to protect axons from acute inflammatory destruction via an oligodendrocyte-independent mechanism

Analyses of CNS cell responses: Representative images of CD3 labelled T-lymphocyte cells (A, B), IBA1 labelled microglia/macrophages (C, D), NIMP-R14 labelled neutrophils (E, F) and GFAP labelled astrocytes (G, H), in lumbar spinal cord sections from CNTF

<p>Cell counts were conducted in the dorsal column (Dorsal) and at the lateral edges (Lateral) for CD3 labelled T-lymphocytes (I, J), IBA1 labelled microglia/macrophages (K, L), NIMP-R14 labelled neutrophils (M, N), and GFAP labelled astrocytes (O, P). No differences in cell numbers were found between the groups, Data are presented as mean ± SEM. Scale bar represents 50 µm.</p

Representative images of methylene blue stained optic nerve sections used for analyses of axonal numbers.

<p>(A) Illustrates image selection of central (I) and peripheral (II) ROIs. Representative 100×magnification images taken in the centre of nerves from healthy CNTF KO/LIF HET (B), EAE induced CNTF KO/LIF HET (C), healthy CNTF KO/LIF KO (D), and EAE induced CNTF KO/LIF KO (E) mice. ROI’s were selected from within these larger images.</p

Spinal cord histological parameters for CNTF KO/LIF KO vs. CNTF KO/LIF HET healthy and EAE induced mice.

<p>Comparison of key morphometric measures in the dorsal column of the spinal cord, between CNTF KO/LIF KO (LIF KO) and CNTF KO/LIF HET (LIF HET) mice in the presence and absence of EAE. Data are presented as mean ± SEM values for each parameter for each group. Region of interest (ROI) represents the image area at 100×magnification excluding grey matter or artefacts.</p>*<p>P&lt;0.05 by 2-sample t-test, assuming equal variances.</p

The effects of deleting gp130 from oligodendrocytes on EAE disease grade: Average EAE disease severity grades for MBP-cre gp130^fl/fl (black) vs gp130^fl/fl littermate control (grey) mice.

<p>The deletion of gp130 from oligodendrocyte lineage cells resulted in a transient improvement in EAE grade from EAE day 16–18 (P&lt;0.05), but this effect was not sustained. Data are presented as mean ± SEM, and are analysed by Mann Whitney rank sum test.</p

Optic nerve histological parameters for CNTF KO/LIF KO vs. CNTF KO/LIF HET healthy and EAE induced mice.

<p>Comparison of key morphometric measures in the optic nerve between CNTF KO/LIF KO (LIF KO) and CNTF KO/LIF HET (LIF HET) mice in the presence and absence of EAE. Data are presented as mean ± SEM values for each parameter for each group. Region of interest (ROI) represents an area of 3480 µm².</p>*<p>P&lt;0.05 by 2-sample t-test, assuming equal variances.</p

Analyses of inflammatory cell responses: Using images of Hoechst stained lumbar spinal cord sections captured at 20×magnification, the percentage of spinal cord area covered by cellular nuclei was measured using ImageJ, to indicate changes in cellularity

<p>No differences were found between CNTF KO/LIF HET and CNTF KO/LIF KO mice during EAE day 15–16. Similarly, measures of the total area of inflammatory cell infiltrates in lumbar spinal cord sections captured at 5×magnification, showed no difference between the groups (B). Passive transfer EAE experiments showed no difference in EAE disease severity scores of CNTF WT/LIF WT recipient mice that received MOG primed T-lymphocytes harvested from CNTF KO/LIF HET or CNTF KO/LIF KO donor mice (C). Data are presented as mean ± SEM.</p

EAE disease severity grades.

<p>EAE disease severity scores for CNTF WT/LIF WT (n = 20) vs CNTF WT/LIF HET (n = 9) mice (A); CNTF WT/LIF HET (n = 30) vs CNTF WT/LIF KO (n = 21) mice (B); CNTF KO/LIF WT (n = 19) vs CNTF KO/LIF HET mice (n = 17) (C); and CNTF KO/LIF HET (n = 30) vs CNTF KOLIF KO (18) mice (D), over time. In CNTF gene deficient mice, the presence of the LIF gene was associated with reduced EAE disease severity scores relative to LIF gene deficient mice, from EAE day 15–18 (D)(*P&lt;0.05). All data are presented as mean ± SEM.</p