Mesoderm induction and the control of gastrulation in Xenopus laevis: the roles of fibronectin and integrins

Exposure of isolated Xenopus animal pole ectoderm to the XTC mesoderm-inducing factor (XTC-MIF) causes the tissue to undergo gastrulation-like movements. In this paper, we take advantage of this observation to investigate the control of various aspects of gastrulation in Xenopus. Blastomeres derived from induced animal pole regions are able, like marginal zone cells, but unlike control animal pole blastomeres, to spread and migrate on a fibronectin-coated surface. Dispersed animal pole cells are also able to respond to XTC-MIF in this way; this is one of the few mesoderm-specific responses to induction that has been observed in single cells. The ability of induced animal pole cells to spread on fibronectin is abolished by the peptide GRGDSP. However, the elongation of intact explants is unaffected by this peptide. This may indicate that fibronectin-mediated cell migration is not required for convergent extension. We have investigated the molecular basis of XTC-MIF-induced gastrulation-like movements by measuring rates of synthesis of fibronectin and of the integrin beta 1 chain in induced and control explants. No significant differences were observed, and this suggests that gastrulation is not initiated simply by control of synthesis of these molecules. In future work, we intend to investigate synthesis of other integrin subunits and to examine possible post-translational modifications to fibronectin and the integrins.     

Influence of the pyruvate content of xanthan on macromolecular association in solution

The unusual increase in viscosity and pseudoplasticity often observed when salts are added to moderately concentrated aqueous solutions of xanthan gum is shown to arise from an increase in the extent of macromolecular association. The fractional change in viscosity on addition of KCl to salt-free 1% (w/v) solutions of purified polysaccharide in the K⁺ salt form is found to be positive only when the degree of pyruvate substitution (fraction of side chains which carry pyruvate ketal substituents) exceeds ≈0.31. Above this value, the fractional change in viscosity increases with further increase in the degree of pyruvate substitution. These differences cannot arise from different degrees of conformational ordering, since the magnitude of the thermally induced order disorder transition (monitored by optical rotation at low ionic strength) is independent of pyruvate content. The temperature at the transition midpoint, however, falls with increasing degree of pyruvate substitution. This is attributed to destabilization of the ordered structure by intramolecular electrostatic repulsion between pyruvate groups, and stabilization through apolar interactions of acetate methyl groups. Viscosity-concentration relationships show changes of slope which mark the onset of macromolecular association. Association commences at lower concentrations when ionic strength and degree of pyruvate substitution are high. It is suggested that once electrostatic repulsions have been diminished at high ionic strength, association is promoted by intermolecular apolar interactions of pyruvate methyl groups, which are suitably near the periphery of the helical conformation.     

Atypical B cells are part of an alternative lineage of B cells that participates in responses to vaccination and infection in humans

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PfSPZ-CVac efficacy against malaria increases from 0% to 75% when administered in the absence of erythrocyte stage parasitemia: A randomized,placebo-controlled trial with controlled human malaria infection

PfSPZ-CVac combines ‘PfSPZ Challenge’, which consists of infectious Plasmodium falciparum sporozoites (PfSPZ), with concurrent antimalarial chemoprophylaxis. In a previously-published PfSPZ-CVac study, three doses of 5.12x10⁴ PfSPZ-CVac given 28 days apart had 100% vaccine efficacy (VE) against controlled human malaria infection (CHMI) 10 weeks after the last immunization, while the same dose given as three injections five days apart had 63% VE. Here, we conducted a dose escalation trial of similarly condensed schedules. Of the groups proceeding to CHMI, the first study group received three direct venous inoculations (DVIs) of a dose of 5.12x10⁴ PfSPZ-CVac seven days apart and the next full dose group received three DVIs of a higher dose of 1.024x10⁵ PfSPZ-CVac five days apart. CHMI (3.2x10³ PfSPZ Challenge) was performed by DVI 10 weeks after the last vaccination. In both CHMI groups, transient parasitemia occurred starting seven days after each vaccination. For the seven-day interval group, the second and third vaccinations were therefore administered coincident with parasitemia from the prior vaccination. Parasitemia was associated with systemic symptoms which were severe in 25% of subjects. VE in the seven-day group was 0% (7/7 infected) and in the higher-dose, five-day group was 75% (2/8 infected). Thus, the same dose of PfSPZ-CVac previously associated with 63% VE when given on a five-day schedule in the prior study had zero VE here when given on a seven-day schedule, while a double dose given on a five-day schedule here achieved 75% VE. The relative contributions of the five-day schedule and/or the higher dose to improved VE warrant further investigation. It is notable that administration of PfSPZ-CVac on a schedule where vaccine administration coincided with blood-stage parasitemia was associated with an absence of sterile protective immunity.Clinical trials registration: {"type":"clinical-trial","attrs":{"text":"NCT02773979","term_id":"NCT02773979"}}NCT02773979.     

Purification, partial characterization and biological effects of the XTC mesoderm-inducing factor

The mesoderm of Xenopus laevis is formed through an inductive interaction in which a signal from the vegetal hemisphere of the blastula acts on overlying animal pole cells. We have recently reported that the Xenopus XTC cell line secretes a mesoderm-inducing factor (MIF) which may resemble the natural signal. In this paper, we describe the purification and biological effects of XTC-MIF. XTC-MIF is a hydrophobic protein with an isoelectric point of 7.8 and an apparent relative molecular mass (Mr) of 23,500. On reduction, XTC-MIF loses its biological activity and the protein dissociates into two inactive subunits with apparent Mr of about 15,000. These properties closely resemble those of transforming growth factor type beta (TGF-beta), and it is interesting that TGF-beta 2 has recently been shown to have mesoderm-inducing activity. The biological response to XTC-MIF is graded. After exposure to 0.2-1.0 ng ml-1 XTC-MIF, stage-8 animal pole explants form mesenchyme and mesothelium. At higher concentrations, up to about 5 ng ml-1, muscle is formed, occasionally with neural tissue. In response to concentrations of XTC-MIF greater than 5-10 ng ml-1, notochord and neural tissue are usually formed. The formation of notochord and neural tissue in response to XTC-MIF represents a qualitative difference between this inducing factor and the other known group of MIFs, the heparin-binding growth factors.     

Mesoderm induction in Xenopus laevis: responding cells must be in contact for mesoderm formation but suppression of epidermal differentiation can occur in single cells

When Xenopus embryos are cultured in calcium- and magnesium-free medium (CMFM), the blastomeres lose adhesion but continue dividing to form a loose heap of cells. If divalent cations are restored at the early gastrula stage the cells re-adhere and eventually form muscle (a mesodermal cell type) as well as epidermis. If, however, the cells are dispersed during culture in CMFM, muscle does not form following reaggregation although epidermis does. This suggests that culturing blastomeres in a heap allows the transmission of mesoderm-induction signals from cell to cell while dispersion effectively dilutes the signal. In this paper, we have attempted to substitute for cell proximity by culturing dispersed blastomeres in XTC mesoderm-inducing factor (MIF). We find that dispersed cells do not respond to XTC-MIF by forming mesodermal cell types after reaggregation, but the factor does inhibit epidermal differentiation. One interpretation of this observation is that an early stage in mesoderm induction is the suppression of epidermal differentiation and that formation of mesoderm may require contact-mediated signals that are produced in response to XTC-MIF. We have gone on to study the suppression of epidermal differentiation in more detail. We find that this is a dose-dependent phenomenon that can occur in single cells in the absence of cell division. Animal pole blastomeres become more difficult to divert from epidermal differentiation at later stages of development and by stage 12 they are 'determined' to this fate. Fibroblast growth factor (FGF) also suppresses epidermal differentiation in isolated animal pole blastomeres and transforming growth factor-beta 1 acts synergistically with FGF in doing so.