Claudie Hulin
email: claudie.hulin@cnrs-orleans.fr.

Lionel Mercury

Capillarity-driven supersolubility in dual-porosity systems

Keywords: Pore size-controlled solubility, Thermodynamics, Metastability, Tensile water properties, Phase transitions

Capillary phenomena are widespread in unsaturated media such as soils, building stone, deep depleted aquifers and still gas storage/sequestration reservoirs. Processes linked to capillarity depend on pore size and environmental conditions such as relative humidity. When capillary forces are high, water's internal pressure can reach negative values representative of a tensile state. In those

conditions, the capillary tensile water provokes compaction in moist granular materials. In geochemical terms, thermodynamics predict a significant effect on the solvent properties of tensile water. However, very little experimental work has validated thermodynamics models or shown the porosity arrangements that make capillarity a significant player to control the geochemical balance and poromechanics of porous media. In this study, we designed an experimental setup in glass microtubes (ø 200 µm) conducive to capillary tension. In the tubes, salts (halite and sodium sulfate) precipitated from an evaporating solution to build a strongly contrasted dual-porosity system. Large pore bodies (ø 200 µm) coexisted with nanometric pores that induced capillary tension in the whole volume. We observed mineral-liquid interactions (mass gain/loss, crystal shaping), especially how salt solubility changed as a function of capillary conditions. The water's tensile state increased salt solubility, changing the reactions' equilibrium constants at constant composition as predicted by thermodynamics. In addition, mineral grains showed clear evidence of poromechanical tensile stress, driving salts to move towards the interior of the tube or to crack. The results showed the potential significance of capillarity in heterogeneous porous media including nanopores. They should be considered in relation to how porous structures change, in the contexts of increasing droughts due to climate changes and increasing excavation work in deep sedimentary basins.

INTRODUCTION

Porous soils and rocks exposed to the atmosphere undergo weathering processes and erosion, which are heightened by frequent drying periods. Close to dry air, porous networks are characterized by the coexistence of air and water, and the fluid behavior is partly controlled by capillary forces. The so-called unsaturated zone (UZ) is widely present in near-surface environments, as soils and exposed rocks, as well as in engineering structures, monuments and artworks [START_REF] Desarnaud | Measurement of the Pressure induced by salt crystallization in confinement[END_REF]. Global climate change, especially regarding humidity cycles (more frequent rainfall, flooding and extreme aridity, sea level rise), may directly impact behavior in the UZ and the associated weathering-erosion processes. Secondly, industrial processes such as gas injection, nuclear waste disposal and fluid removal drive deep geological formations to UZ-like states (e.g. [START_REF] André | Well injectivity during CO 2 storage operations in deep saline aquifers-Part 2: Numerical simulations of drying, salt deposit mechanisms and role of capillary forces[END_REF]2015;Peysson et al., 2014;[START_REF] Ott | Salt precipitation due to supercritical gas injection: I. Capillary-driven flow in unimodal sandstone[END_REF], with possibly extremely dry (artificial) conditions.

Under drying conditions, water evaporates and the liquid-air interface retreats until it is retained in the thinnest pores by capillarity, where the internal liquid pressure decreases. The relative volume of capillary water results in poral geometry and dryness: for instance, multiscale porous networks retain larger volumes of capillary water, especially if large pores are connected through nanoscale pore throats (Fig. 1). In such a network, the nanometric capillary bridges in the pore throats generate high capillary forces that are transmitted to the whole volume of hydraulically connected water (outer pore throats and inner pore bodies) (Fig. 1). This situation can arise in sedimentary rocks (Fig. 1a), (e.g. [START_REF] Muller | Implication of fractal geometry for fluid flow properties of sedimentary rocks[END_REF][START_REF] Wang | Nanogeochemistry: Geochemical reactions and mass transfers in nano-pores[END_REF][START_REF] Anovitz | Diagenetic changes in macro-to nano-scale porosity in the St. Peter Sandstone: An (ultra) small angle neutron scattering and backscattered electron imaging analysis[END_REF] or in crustal rocks (Fig. 1b), [START_REF] Holness | Equilibrium dihedral angles in the system quartz-CO 2 -H 2 O-NaCl at 800°C and 1-15 kbar: the effect of pressure and fluid composition on the permeability of quartzites[END_REF][START_REF] Holness | The effect of feldspar on quartz-H 2 O-CO 2 dihedral angles at 4 kbar, with consequences for the behaviour of aqueous fluids in migmatites[END_REF]. Evaporation of in-pore water also leads to the precipitation of salt crystals initially dissolved in the poral solution. As they grow and accumulate in the poral network, those salt crystals can clog the pores and ultimately cause damages by exerting pressure against the pore wall (crystallization pressure, e.g. [START_REF] Correns | Growth and dissolution of crystals under linear pressure[END_REF][START_REF] Scherer | Crystallization in pores[END_REF][START_REF] Flatt | Salt damage in porous materials: how high supersaturations are generated[END_REF][START_REF] Benavente | Role of pore structure in salt crystallization in unsaturated stone[END_REF]Steiger, 2005a;[START_REF] Røyne | Pore-scale controls on reaction-driven fracturing[END_REF][START_REF] Coussy | Deformation and stress from in-pore drying-induced crystallization of salt[END_REF][START_REF] Shahidzadeh | Damage in porous media: role of the kinetics of salt (re)crystallization[END_REF]. Yet, in both arid environments and those with wetting and drying cycles, salts are recognized to be responsible for the alteration of porous media (e.g. [START_REF] Wellman | Salt weathering, a neglected geological erosive agent in coastal and arid environments[END_REF][START_REF] Cooke | Salt weathering in deserts[END_REF][START_REF] Goudie | Salt weathering hazards[END_REF]. While drying processes and evaporation have attracted a lot of attention, only a few studies have focused on the role of capillarity in weathering processes. However, capillarity is recognized to provoke the compaction of granular materials (e.g. [START_REF] Fraysse | Humidity effects on the stability of a sandpile[END_REF], possibly inducing cracks in clays (e.g. [START_REF] Weinberger | Initiation and growth of cracks during desiccation of stratified muddy sediments[END_REF] and cohesive sands [START_REF] Chavdarian | Origin and evolution of polygonal cracks in hydrous sulphate sands, White Sands National Monument, New Mexico[END_REF]. This effect can also be invoked to explain certain cracks in shotcrete (Ansell, 2010) and cementitious materials (e.g. [START_REF] Lagier | Numerical strategies for prediction of drying cracks in heterogeneous materials: Comparison upon experimental results[END_REF] and deformation observed in soft porous material (Bouzid et al., 2011b) and crystalline nano-channels [START_REF] Tas | Capillarity induced negative pressure of water plug in nanochannels[END_REF]. From the geochemical viewpoint, field observations suggest that a capillary state influences pedogenetic evolution over millions of years, accumulating preferential anhydrous/hydrous mineral phases depending on climate cycles and mean pore sizes (e.g. [START_REF] Tardy | Geochemistry of laterites, stability of Al-goethite, Al-hematite, and Fe 3+kaolinite in bauxites and ferricretes: an approach to the mechanism of concretion formation[END_REF]Trolard and Tardy;[START_REF] Trolard | A model of Fe 3+ -kaolinite, Al 3+ -goethite, Al 3+ -hematite equilibria in laterites[END_REF][START_REF] Roedder | Significance of monophase fluid inclusions in minerals[END_REF][START_REF] Tardy | Petrology of laterites and tropical soils[END_REF]. Capillary water's thermodynamic properties can be calculated either through a water activity framework (e.g., [START_REF] Trolard | The stabilities of gibbsite, boehmite, aluminous goethites and aluminous hematites in bauxites, ferricretes and laterites as a function of water activity, temperature and particle size[END_REF][START_REF] Trolard | A model of Fe 3+ -kaolinite, Al 3+ -goethite, Al 3+ -hematite equilibria in laterites[END_REF][START_REF] Wang | Nanogeochemistry: Geochemical reactions and mass transfers in nano-pores[END_REF] or by a liquid pressure approach (Mercury and Tardy, 1997a, b;[START_REF] Mercury | Negative pressure of stretched liquid water. Geochemistry of soil capillaries[END_REF][START_REF] Mercury | Thermodynamic properties in metastable systems under negative or positive pressures[END_REF]2004;[START_REF] Lassin | Geochemistry of unsaturated soil systems: aqueous speciation and solubility of minerals and gases in capillary solutions[END_REF]. Both models predict how the geochemical mass balance will change in any UZ holding capillary water.

The effects of capillarity are size-dependent, and so may be significant only for nanometric pores.

At this scale, the capillary water is under absolute negative pressure (or under capillary tension) and able to exert a tensile force (e.g. be responsible for granular stack compaction). Nanometric pores can be quantitatively important in sedimentary rocks. Macropores are connected through nanometric pore throats (Muller and Mc Cauley, 1992;[START_REF] Anovitz | Diagenetic changes in macro-to nano-scale porosity in the St. Peter Sandstone: An (ultra) small angle neutron scattering and backscattered electron imaging analysis[END_REF] as in Figure 1. Water retained in the large pore bodies surrounded by nanothroats can reach and sustain the same negative pressure as the nanocapillary water in the pore throats before water cavitation.

Our laboratory experiments studied the physical and chemical behavior of water-solid interactions in such situations. We developed a set-up based on an existing protocol (Fig. 2; Bouzid et al., 2011b;[START_REF] Røyne | Pore-scale controls on reaction-driven fracturing[END_REF] where a bimodal pore system conducive to capillary tension was built.

Cylindrical microtubes were filled with a saline solution that evaporated in drying conditions. The solution became supersaturated with respect to its solid, which precipitated on both air-liquid interfaces (highest salt concentration). The salts grew until forming a cylinder that apparently clogged the tube, but a thin nanometric film persisted between the crystal and the glass wall (Fig. 2a). This film is assumed to have a zero contact angle with the silica walls, so the meniscus diameter is equal to the thickness of the open space. If the solution-air interfacial curvature became large enough to equilibrate with the imposed relative humidity (as expected from the Kelvin equation), capillary forces prevented evaporation and generated negative pressure in the entire trapped solution including the macrovolumeThe entire volume of solution was in a tensile state (Fig. 2b), which is supported by the final bubble nucleation appearing inside the trapped macrovolume (Fig. 2c). The water under tension was in a metastable state (superheated) until the nucleation of vapor returned the system to its most stable state (liquid-vapor assemblage at saturated vapor pressure). After bubble nucleation in the pore body, the liquid internal pressure is brought back at the atmospheric value. Consequently, there is no more pressure gradient across the liquid-air interface, which becomes flat (Fig. 2c).

. This method turns the initially monosized "saturated" microtube into an unsaturated bimodal pore system prone to high capillary conditions. Our goals were to experimentally validate the geochemical signatures of capillarity and to show that dual-porosity media can contain large capillary volumes.

MATERIAL AND METHODS

Materials

Micro-capillaries with 200 µm internal diameter (VitroTubes TM ) were filled with a saline solution (NaCl or Na 2 SO 4 ) that evaporated under controlled relative humidity.

NaCl and Na 2 SO 4 aqueous solutions were prepared from NormaPur powder and distilled water.

The NaCl solution was at 5.5 mol.L -1 (24.3 wt %) and had a relative supersaturation n/n 0 = 0.90 (molality at saturation n 0 = 6.15 mol.L -1 at 20°C). The Na 2 SO 4 solution was at 3.5 mol.L -1 (33.3 wt %), so undersaturated with respect to thenardite (relative supersaturation n/n 0 = 0.64 at 20°C) and supersaturated with respect to mirabilite (relative supersaturation n/n 0 = 1.19 at 20°C). The saline solutions were filtered with 0.450 µm filters (VWR® syringe filters) to avoid impurities and micro-crystallites. This means that there are no or very few potential seeds in the solution, so that during evaporation, the solution will reach a high supersaturation before crystals nucleate [START_REF] Desarnaud | Metastability limit for the nucleation of NaCl crystals in confinement[END_REF].

The evaporative demand, leading to evaporation or capillary conditions, was set by controlled relative humidity (RH) in a climate chamber (HPP108, Memmert technology). The preferred RH was 48%, to be compared to the equilibrium RH of a NaCl saturated solution (75.3%, [START_REF] Greenspan | Humidity Fixed Points of Binary Saturated Aqueous Solution[END_REF] or of a Na 2 SO 4 solution (93%, [START_REF] Winston | Saturated Solutions for the Control of Humidity in Biological Research[END_REF]. This RH gradient enables a moderate drying rate promoting the precipitation of massive salt [START_REF] Desarnaud | Metastability limit for the nucleation of NaCl crystals in confinement[END_REF] when the appropriate supersaturation was reached, at constant temperature. The microcapillaries were regularly observed using an optical microscope (Leica DM2500) in a room thermostated at 20°C. Observations were made in transmitted light using lens with 5x, 10x, 20x and 50x magnification. Pictures were captured with a Leica DFC295 digital camera allowing image resolution of 3 megapixels (2048 x 1536) and pixel size of 1.2 µm x 1.2 µm with a 5x magnification lens. To avoid any heat transfer, we reduced exposure to microscope lights to the minimum possible, so we can consider heating due to light as negligible.

Methods

Three types of experiments (Fig. 3; also supp. info.) were carried out, differentiated according to:

-two lengths of liquid column trapped in between the crystal plugs: long and short, corresponding to large and small volumes of tensile water; -two types of initial solutions, based on NaCl and Na 2 SO 4 electrolytes, because the solvent properties of the associated crystals phases are different when the water is in a tensile state.

The NaCl-short solution systems (Fig. 3a-d) were made from a highly supersaturated solution, using plasma-cleaned tubes (15 min) and filtered solution. This made the halite nucleate inside the solution and grow rapidly on both sides with a symmetric hopper (skeletal) shape [START_REF] Sunagawa | Growth and morphology of crystals[END_REF][START_REF] Desarnaud | Metastability limit for the nucleation of NaCl crystals in confinement[END_REF] (Fig. 3a). First, when the supersaturation was still high, several cubes of halite nucleated and crystallized on each other, with growth parallel to the tube. As the supersaturation decreased, the salt continued to grow at each side preferentially onto the inner wall of the tube. As the solution kept on evaporating, the salt continued to grow on each side and not in the middle, because at this point the solution was trapped and did not evaporate. This explaining the "bow-tie" shape (Fig. 3b-c). The solution on each side continued to evaporate until reaching first one side of the crystal (Fig. 3c) then the other (Fig. 3d). The liquid air interface stopped receding since no air entered in the trapped solution. We built 26 "bow-tie" systems.

For the long-NaCl solution systems (Fig. 3e), the NaCl solution was left to evaporate until reaching high supersaturation, without any crystal nucleation or growth. Then we broke the tube on each side close to the air-water menisci. This induced rapid evaporation at both liquid-air interfaces and led to the precipitation of two opposite crystals along the interfaces. The two crystals grew rapidly and apparently clogged the tube, trapping the large volume of solution (from 6.9 to 51 mm long, so from 0.157 to 1.88 mm 3 in volume). As in the bow-tie shape systems, the trapped solution did not evaporate. Twelve of these systems were built. Concerning Na 2 SO 4 , no useful distinction was made by tube length, so the experiments were performed irrespective of the trapped water volume. The protocol consisted of filling the tube with a 33.3%wt solution, undersaturated with respect to thenardite (anhydrous stable phase) and supersaturated with respect to mirabilite (decahydrate stable phase). After evaporation, the salt first crystallized into a dendritic metastable phase III (Fig. 3f) then transformed into stable phase V, thenardite (Fig. 3g-h), as commonly observed [START_REF] Steiger | Crystallization of sulfate phases in porous materials: The phase diagram Na 2 SO 4 -H 2 O and the generation of stress[END_REF][START_REF] Shahidzadeh | Damage in porous media: role of the kinetics of salt (re)crystallization[END_REF]. The crystallization sequence resulted in numerous thenardite crystals in solution (Fig. 3i), which coalesced to form plugs (Fig. 3j). The formation of massive plugs was encouraged by using highly saline solution (33.3% wt) introducing high salt mass. Ten such systems were built.

RESULTS

The dual-porosity systems were built in 48 tubes (table 1) either with one bow-tie shaped halite crystal (Fig. 3d), or with two distinct crystals (halite or thenardite). In each system, after the liquid-air interface receded and reached the external side of a crystal (Fig. 2b), we observed no further air entry in the trapped macrovolume. The liquid-air interface remained in the narrow space between the salt plugs and the glass wall and stopped evaporating. After one external side of one crystal was dry (and only after), we observed different reactions in the trapped volume: salt dissolution, gas nucleation and solid displacement. Over time (up to 2 years), we carefully monitored the events occurring within the trapped solution and/or affecting the crystal plugs.

Gas nucleation

The most common reaction, observed in 35 systems, was the formation of a gas bubble in the trapped solution after a certain amount of timefrom several hours to monthsafter at least one of the external sides of the salt(s) was dry. This gas nucleation has already been observed and was not the result of air entry (Bouzid et al., 2011b). Figure 4 illustrates two experiments in which gas nucleated in a solution trapped between two thenardite crystals (Na 2 SO 4 ). In the first experiment, gas nucleated 20 days after the external side of the right crystal dried (Fig. 4A) and in the second experiment, gas nucleated two months after the external sides of both crystals dried (Fig. 4B). We observed gas nucleation in the trapped solution in bimodal pore systems built with NaCl (halite) and Na 2 SO 4 (thenardite). In each of the 10 systems built with thenardite, gas nucleated from 20 days to 3 months after at least one external side of the salts dried. In systems built with halite crystals, we observed gas nucleation in 25 of the 38 experiments: in 20 of the 26 bow-tie shape systems (Fig. 3a-d), and in 5 of the 12 systems built with two distinct salts (Fig. 3e; see also supplementary information).

Inward movements of salt crystals

With two of the NaCl-long systems, crystal plugs were seen to move (Fig. 5). In both cases, we observed the inward motion of one or both salt plugs until a gas bubble appeared in the trapped solution. In the experiment shown in Fig. 5, the crystal on the left slid over 3.3 mm in four days (Fig. 5b-c). When a gas bubble appeared on the right (Fig. 5d), the crystal stopped and never moved again. In the experiment shown in Fig. 6, both salts moved inward for seven days (Fig. 6, step a to c) before a gas bubble nucleated (Fig. 6, step d). The salt on the left moved over 3.3 mm while the opposite one (on the right) moved over 0.13 mm (see the closer view 2, Fig. 6).

No plug motion was observed in any of the 10 Na 2 SO 4 based systems.

Dissolution of halite

Dissolution and traction

The moving crystal on the right in the NaCl-long experiment had been partially dissolved before the gas appeared (Fig. 6A, closer view 2). The simple cylindrical geometry of the system allowed us to measure the volume of the trapped solution and the total mass loss. The dissolved length of halite was 130 ± 13 µm, corresponding to a volume of 4.1.10 -3 ± 0.4.10 -3 mm 3 , and therefore a mass loss of 8.9 ± 0.9 µg (calculated with a halite density of 2.17 g/cm 3 ). The distance between the two salt plugs before (Fig. 6A, step a) and after crystals moved (Fig. 6A, step b) was respectively 10 ± 0.2 mm and 6.6 ± 0.2 mm, corresponding to a solution volume of 0.32 ± 0.01 mm 3 and 0.21 ± 0.01 mm 3 . Furthermore, a close examination of the same halite plug after the bubble nucleated showed clear lengthening over time, evidencing a reprecipitation process: the process appeared reversible.

Bow-tie shaped systems

Whereas salt dissolution was only observed in one experiment built with two distinct salts (large double plug systems, Fig. 3e,j), significant salt dissolution was observed in 16 of the 26 experiments built with bow-tie shape halite (Fig. 3A). Dissolution affected the salt in contact with the trapped solution at the thinnest part. In most cases, the bow-tie crystal was separated into two pieces and both pieces became rounded (Figs. 7 and8, steps c and d). Twenty of the 26 experiments built with bow-tie shaped halite showed gas nucleation after a certain period, between 5 days and several months (section 3.1). Gas nucleated in each of the 16 cases displaying salt dissolution, and there were no cases where salt dissolved after gas nucleation.

Dissolution in bow-tie shaped crystals lasted between 5 and 45 days. In most of the cases dissolution time was 10 to 15 days.

Due to the complex geometry of the bow-tie halite crystals, we could only calculate the volume of solution and the mass loss in one case (Fig. 8B). The volume of the trapped solution was estimated at 13.10 -3 ± 1.3.10 -3 mm 3 , and the mass loss of solid salt at 1.1 ± 0.1 µg (Fig. 8B).

The case of thenardite

In Na 2 SO 4 systems, we observed neither dissolution nor movement of the thenardite crystal.

However, as in NaCl systems, a gas bubble appeared in the trapped solution, from one day to months after one or both salt plugs dried (Fig. 9). A closer view of both surrounding salt plugs did not show any mass change with respect to the initial situation: no significant dissolution occurred during those two months.

Plug cracking

Another interesting observation was made with Na 2 SO 4 double plug systems, at the critical moment when one salt plug dries. In this experiment we cut the tube (Fig. 10, step a), enhancing the evaporation of the saline solution. After around ten minutes, the liquid-air interface reached the salt plug (Fig. 10, step b) and became confined in the small space between the salt plug and the tube wall.

As the interface continued to recede between the salt and the tube wall, cracks appeared and propagated on the crystal surface along preexisting discontinuities (probably the mark of crystal coalescence).

DISCUSSION

Tensile state and tensile stress

Gas nucleation and capillary film

The presence of a capillary film along a nano-scale space between the solid plugs and the tube walls is a first assumption of this work. We aspire to measure the properties and/or thickness of this film but have not achieved this yet. The Young-Laplace law (suppl. info.) gives the pressure drop across a curved interface between two phases. With concave air-liquid interfaces, the liquid pressure is lower than the air pressure, and increases as interface size decreases. This interface is itself related to the pore size by the value of its contact angle with both supporting solids (supp. info). As a second assumption in the calculations, the contact angle between the liquid and the solid host is assumed to be zero.

The thermodynamic driving force towards capillarity comes from air's RH when below the saturated vapor pressure (suppl. info.). If the water-air surface is flat, low RH drives evaporation that increases the local air vapor pressure. In small pores, low RH can drive the liquid to decrease its internal pressure (the water is in capillary state). If the local pore size is sufficiently small, the liquidair interfacial curvature becomes large enough to equilibrate water and dry air (Kelvin equilibrium is reached). The liquid pressure can decrease according to the Young-Laplace law (eq (2), suppl. info.).

Consequently, the decrease in liquid pressure, driven by dry air, is controlledand limited by the size of the nano-throats and is identical in the entire hydraulically connected volume. Yet the macropores are so large that they cannot prevent the boiling of tensile water. The formation of a sphere of vapor of radius r brings the system to its most stable state which gains energy equal to with P the pressure of the tensile liquid and P sat the saturation vapor pressure). However, the creation of the vapor-liquid interface of radius r requires an activation energy equal to (with γ the liquid-vapor surface tension). According to the Classical Nucleation Theory (CNT), the competition between these two opposite effects results in an energy barrier E b :

(1) which is the minimum energy required to create a sphere of vapor of critical radius r (e.g. Debenedetti, 1996, Caupin and[START_REF] Caupin | Cavitation in water: a review[END_REF]. In nanothroat, the size is too small for the vapor nucleus to reach the critical radius required for vapor nucleation. In contrast, the adjacent connected macropore is much larger than the critical radius, enabling vapor nucleation at any time. Before that, the occluded tensile water is metastable (superheated) during a limited lifetime that decreases with the superheating intensity, which is directly depending on .

During the course of our experiments, we observed vapor nucleation in 35 on 48 microtubes after a superheating period ranging for a few days to years.

At the nanoscale, repulsive forces are caused when a fluid sandwiched between two solids undergoes the overlapping of surface force fields emanating from the facing solids (e.g. [START_REF] Derjaguin | Surface forces[END_REF]. This disjoining effect accounts for the open film, even in the presence of a supersaturated solution (e.g. [START_REF] Desarnaud | Measurement of the Pressure induced by salt crystallization in confinement[END_REF]. However, impurities, wall roughness and excess supersaturation, for instance, could initiate precipitation of microcrystals that would seal the open film and totally clog the tube. This process could explain the absence of any change in the system, including gas nucleation, in six NaCl-short and 5 NaCl-long experiments (Table 1; supp. info). Water in the macrovolume would simply be trapped, without any capillary forces to put it under tensile stress, and therefore without gas nucleation, enhanced dissolution, or plug motion. It is worth saying that two NaCl-long experiments first showed slow inward halite plug motion for 3.5 and 7 months. Once the plugs stopped, no gas nucleation happened in the following year, meaning that the trapped macrovolume of solution can maintain a tensile state for a long time.

Mechanical effects

The inward motion of salt plugs (Figs 5 and6) demonstrates that the crystals did not obstruct the tube and were mechanically free to move. This is consistent with the macrovolume of solution having a tensile state before vapor nucleation. As vapor nucleated, the stretched water relaxed its tension, becoming unable to exert any tensile forces on the salts. In the two experiments where salt plugs moved inward, the two salt plugs moved differently (Fig. 5) or only one salt plug moved (Fig. 6). The liquid must exert an isotropic and homogenous tension in the macrovolume, but the friction forces between the glass wall and each solid plug should have been different, probably because each crystal had its own surface roughness.

The plug cracks observed at the thenardite surface (Fig. 9) are assumed to be the result of capillary tension exert by the nanometric liquid-air interface. Cracks appeared as the nanometric interface receded between the salt and the tube wall, several minutes after we cut the tube. Therefore, the cracks were not related to the mechanical procedure of cutting the tubes. These cracks revealed the mechanical constraints exerted by the liquid against the solid surface. Yet the tensile stress in liquid is isotropic, so the mechanical effects depend primarily on the strength of the solids themselves. The glass wall was probably unaffected because of its high mechanical resistance. The thenardite crystals may have cracked because of the preexisting defects linked to the coalescence formation process (Fig. 9). Considering texture heterogeneity of pore walls in natural rocks, especially discontinuity in the structure or dislocations, even low tension can have strong mechanical effects.

We used two easy methods to ensure that the Na 2 SO 4 that crystals were thenardite (anhydrous crystals) and not mirabilite (decahydrate crystals). First, mirabilite rapidly dehydrates in microcrystals of thenardite in contact with dry air with RH of 93% or less (equilibrium vapor pressure of mirabilite).

This event is very characteristic and easily recognizable. As the experiments were conducted at 48% RH, mirabilite must dehydrate, so we can directly identify the crystals during the experiment.

Secondly, mirabilite dissolves at 32.4°C (e.g. [START_REF] Steiger | Crystallization of sulfate phases in porous materials: The phase diagram Na 2 SO 4 -H 2 O and the generation of stress[END_REF]. To be sure that the salt plugs were thenardite, at the end of experiments we heated the sample in a Linkam stage. As the crystal plugs did not dissolve above 32.4°C, we confirmed that the crystals observed were indeed thenardite. This has been confirmed using XRD analysis on two tubes containing Na 2 SO 4 salts that we identified as thenardite from morphology, behavior in dry environment, and heating.

Supersolubility effect

The dissolution of halite demonstrated that the trapped solution was becoming undersaturated with respect to halite. As temperature and composition remained constant, the change necessarily relates to the reaction's equilibrium constant. This change is attributed to the liquid pressure decreasing toward negative values, which is consistent with the other observations, like gas nucleation and salt plug traction. The tension was imposed by capillary forces settled by one or two nanometer scale menisci surrounding the macrovolume of solution. Thermodynamics involving capillarity (Mercury and Tardy, 1997 a, b;[START_REF] Mercury | Negative pressure of stretched liquid water. Geochemistry of soil capillaries[END_REF][START_REF] Mercury | Thermodynamic properties in metastable systems under negative or positive pressures[END_REF]2004;[START_REF] Lassin | Geochemistry of unsaturated soil systems: aqueous speciation and solubility of minerals and gases in capillary solutions[END_REF]; see also supp. info) predict how the equilibrium constant of any reaction changes when the liquid pressure changes.

(2) K P , K 0 are the equilibrium constant for reactions interacting with capillary and bulk liquid respectively, V° is the ionic volume of the subscribed phase at infinite dilution, n is the number of H 2 O molecules involved in the reaction (from 0 for halite to 10 for mirabilite dissolution), R is the gas constant and T is the temperature in kelvin. The equilibrium constant K P in capillary conditions was calculated considering that the chemical potentials of solids kept their values at atmospheric conditions (suppl.

info.).

The increase of solubility of halite, thenardite, and mirabilite can be calculated according to equations ( 2), ( 3) and (4). For halite, the solubility S is the square root of the solubility product K, so

with the halite solubility in capillary conditions (negative pressure P) and the halite solubility at atmospheric pressure. For thenardite and mirabilite, the solubility S is the cubic root of the solubility product K, so

The solubility ratio of halite, thenardite and mirabilite are reported in Fig. 11 as a function of capillary negative pressure. The geochemical behavior can be readily interpreted considering the volume properties of the solutes under tension [START_REF] Mercury | Thermodynamic properties in metastable systems under negative or positive pressures[END_REF]also suppl. Info.).

By calculating the mass of dissolved halite, it was possible to calculate the tension required for the new equilibrium constant. For that, we assumed that the halite + liquid system had reached the new equilibrium imposed by capillary tension. If so, the increased concentration we calculated with the mass loss corresponds to the increase in halite solubility. In some experiments (Figs 7 and8), the dissolution stopped largely before gas nucleation, supporting this assumption. In others, gas nucleated while the solid plugs were dissolving, meaning our calculations gave a minimal tension value, probably underestimated.

In two experiments (Figs 6 and 8), the tension could be estimated because of the simple geometry of crystal plugs and liquid macrovolume. In the NaCl-long experiment (Fig. 6), 8.9 ± 0.9 µg of halite dissolved in a volume of solution of 0.21 ± 0.01 mm 3 . Considering the assumptions stated above, this . The first assumption was that activity coefficients do not change with the increasing concentration linked to the higher solubility. The second assumption is that the system reached equilibrium during the observation time: both the macropore volume and the halite plugs retained their characteristic sizes. The thermodynamic model predicted a tension of 36 ± 6 MPa (Fig. 11, black circle shows the minimal tension 30 MPa). To reach that tension, the liquid-air interface must have a curvature radius of 2.4 ± 0.4 nm and a thickness of 4.8 ± 0.8 nm if the contact angle is equal to zero.

In one NaCl-short experiment (Fig. 8), 1.1 ± 0.1 µg dissolved in 13×10 -3 ± 1.3×10 -3 mm 3 of solution, corresponding to an increased mass concentration of 86 ± 16 µg.mm -3 . This corresponds to an increased solubility of 24 ± 5% ( due to a tension of 75 ± 18 MPa (Fig. 11, hollow square shows the maximum tension 93 MPa). To reach that tension, the liquid-air interface must have a curvature radius of 1.4 ± 0.3 nm and a thickness of 2.8 ± 0.6 nm if the contact angle is equal to zero. In this case, the solid dissolution stopped two weeks before gas nucleated, in close agreement with the assumption that the system had reached a new equilibrium state.

In most of the NaCl-short experiments, the amount of dissolved salt could not be calculated because of the complexity of the crystal shape and of the liquid volume. However, the increased solubility was clearly evidenced by the dissolution of the central part of the mono-solid, leading to two distinct halite plugs. This observed dissolution of the central part of the mono-solid is also consistent with minimizing surface energy of the solid-solution interface.

It is noteworthy that water and aqueous solutions can sustain even higher tensions (100 MPa and beyond) in closed microcavities than the presently recorded capillary tensions (30 to 93 MPa) (e.g. [START_REF] Roedder | Metastable superheated ice in liquid-water inclusions under high negative pressure[END_REF][START_REF] Green | Water and solutions at negative pressure: Raman spectroscopic study to -80 Megapascals[END_REF][START_REF] Zheng | Liquids at large negative pressures: water at the homogeneous nucleation limit[END_REF][START_REF] Alvarenga | Elastic properties of water under negative pressures[END_REF][START_REF] Shmulovich | Experimental superheating of water and aqueous solutions[END_REF][START_REF] Pallares | Equation of state for water and its line of density maxima down to -120 MPa[END_REF][START_REF] Qiu | Exploration of the phase diagram of liquid water in the low temperature metastable region using synthetic fluid inclusion[END_REF]. Interestingly, Chen et al. ( 2016) developed an ink-bottle pore geometry, by bonding a nanoporous silicon membrane onto a glass substrate patterned with large voids. They measured a maximum tension sustainable by the stretched superheated water in their macrovolume from 20 to 30 MPa, matching the present values.

Regarding thenardite (Na 2 SO 4 ), we never detected salt dissolution, while the liquid's tensile state was illustrated by the final water cavitation. The absence of dissolution can be interpreted by the limited increase of thenardite solubility with tension, which reaches a maximum of 5% at -60 MPa before decreasing (Fig. 11).

Re-equilibration after vapor nucleation

After vapor nucleation, the liquid returned to its most stable state (e.g. [START_REF] Caupin | Cavitation in water: a review[END_REF] and the solid salt solubility to its initial value. Consequently, after bubble nucleation, the aqueous solution is instantaneously supersaturated with respect to the solid, and recrystallization is expected to restore the new (though prior) equilibrium. However, especially in the bow-tie shape experiments, the crystal could not recover its initial shape because the secondary crystallization occurred through epitaxial overgrowth on the existing mineral. Considering the salt quantity in excess and the crystal surface area in contact with the solution, the thickness of the overgrown layer would be around 1 ± 0.5 µm along the whole surface area of the solid, or 7 µm with growth restricted to the two curved hemisphere surfaces (Fig. 12C). In both cases, we were not able to visualize the reprecipitated mass under the microscope (Fig. 12, see black arrow: a line 7 µm thick was drawn to appreciate the order of magnitude). In conclusion, salt dissolution was visible and calculable because it only occurred at the thinnest part of the salt where crystal morphology includes more kinks and steps whose surface energy is higher than a flat surface [START_REF] Chernov | Surface phenomena and parameters of crystal growth: simple basics[END_REF]. On the contrary, when the salt reprecipitated to reach the new equilibrium, it was deposited as a thin layer along the whole surface of the existing salt in contact with the saline solution. Consequently, the thickness of this secondary layer was so thin that we could not see it with an optical microscope. However, the reprecipitation materializesin the reshaping: the halite plugs changing from a round shape to halite's typical cubic habit (Fig. 12A,B).

CONCLUSION

This experimental approach illustrates that capillary tension up to tens of MPa can arise in a macrovolume of trapped aqueous solution. The capillarity tension is associated with a bimodal porosity synthesized in an initially monosized tube where sodium sulfate and halite precipitated in an evaporating solution. The dry air conditions additionally allow a nanometric liquid-air curved interface to exist in the thinnest part of the porous domain. These dual-porosity systems enabled us to monitor clear and significant mechanical and geochemical re-equilibration when put under capillary conditions. Increasing solubility, inward motion of grain crystals and crystal cracking were carefully observed in various situations and displayed convergent behaviors. Furthermore, a supersolubility effect can be readily interpreted using a thermodynamic framework that resulted in calculated tensions ranging from 30 to 93 MPa.

The thermodynamic framework was previously used to predict how capillarity might impact the geochemical budget along UZ either through indirect clues (e.g. [START_REF] Tardy | Geochemistry of laterites, stability of Al-goethite, Al-hematite, and Fe 3+kaolinite in bauxites and ferricretes: an approach to the mechanism of concretion formation[END_REF][START_REF] Trolard | The stabilities of gibbsite, boehmite, aluminous goethites and aluminous hematites in bauxites, ferricretes and laterites as a function of water activity, temperature and particle size[END_REF][START_REF] Trolard | A model of Fe 3+ -kaolinite, Al 3+ -goethite, Al 3+ -hematite equilibria in laterites[END_REF][START_REF] Tardy | Petrology of laterites and tropical soils[END_REF] or using thermodynamic predictions (e.g. Mercury and Tardy, 1997;[START_REF] Mercury | Thermodynamic properties in metastable systems under negative or positive pressures[END_REF]2004;[START_REF] Lassin | Geochemistry of unsaturated soil systems: aqueous speciation and solubility of minerals and gases in capillary solutions[END_REF][START_REF] Pettenati | Capillary geochemistry in non-saturated zone of soils. Water content and geochemical signatures[END_REF][START_REF] Pedretti | Analysis of the potential impact of capillarity on longterm geochemical processes in sulphidic waste-rock dumps[END_REF]. Recently Lassin et al. (2016) demonstrated experimentally on CO 2 -water equilibria that these predictions matched measurements. This work establishes that mineral-liquid interactions are significantly impacted and are associated with various mechanical effects that lead to a great variety of transport/reactivity couplings in geological systems.

Interestingly, the experiments we proposed showed both geochemical and poromechanical effects of the capillarity driving force, associated with the existence of a secondary thin porosity in a complex multiphasic system. The tensile stress can modify the pore architecture by displacing in-pore mineral grains, but also may play a role in terms of permeability with crack growth. However, the signatures depend on the chemical systems involved, and that substantially widens the potential variety of patterns in a multi-mineral porous media.

Conversely, our experiments demonstrated that salts precipitating within macro-scale pores or channels can build the required topology for the capillary tension to exist. This means that porous media cementation and the existence of nano-scale throats must be considered for transport property variation over time but should also be carefully examined to enable conclusions on the water-rock interaction processes. The required nano-scale pores are indeed largely encountered in geologic media, especially in sedimentary rocks where large pore bodies are connected through nanometric pore throats (e.g. [START_REF] Muller | Implication of fractal geometry for fluid flow properties of sedimentary rocks[END_REF][START_REF] Wang | Nanogeochemistry: Geochemical reactions and mass transfers in nano-pores[END_REF][START_REF] Anovitz | Diagenetic changes in macro-to nano-scale porosity in the St. Peter Sandstone: An (ultra) small angle neutron scattering and backscattered electron imaging analysis[END_REF]. The ratio of nanopores (let's say, 1 to 100 nm) to the total porosity can be quite large up to, for instance, 10-40% [START_REF] Görres | Soil pore manipulations to increase bioaccessible pore volume[END_REF], 20% [START_REF] Suetsugu | Effects of nanoscale pores in soils on carbon evolution under extremely dry conditions[END_REF], 28-56% [START_REF] Anovitz | Diagenetic changes in macro-to nano-scale porosity in the St. Peter Sandstone: An (ultra) small angle neutron scattering and backscattered electron imaging analysis[END_REF].

As a result, more broadly, capillarity may play a complex, readily hidden, role in how pore structures change when exposed to wetting-drying cycles or are subject to increasing aridity conditions. Global climate change with more frequent extreme events (e.g. [START_REF] Destouni | Screening long-term variability and change of soil moisture in a changing climate[END_REF][START_REF] Touhami | Assessment of climate change impacts on soil water balance and aquifer recharge in a semiarid region in south east Spain[END_REF][START_REF] Wanders | Human and climate impacts on the 21 st century hydrological drought[END_REF] increases the extent of arid cold and warm areas, possible hosts of the behaviors presented here. The anthropogenic interests for deep geological storage targeted complex sedimentary formations capped by thin porous rocks, which are typically prone to strong capillarity states.

Figure 1 .

 1 Figure 1. Dual-porosity systems. (a) in sedimentary oolithic rocks, (b) in crustal settings. (c) corresponding schematic view of a filled macropore surrounded by nanothroats (solid in brown, liquid in blue).

Figure 2 .

 2 Figure 2. Main experimental steps (see text). (Modified from Bouzid et al., 2011b)

Figure 3 .

 3 Figure 3. Crystallization sequences to build double plug systems (see text).

Figure 4 .

 4 Figure 4. Gas nucleation in a Na 2 SO 4 solution trapped by two thenardite crystals. (A) 20 days after one external side of the salt was dry (B) 2 months after both external sides of the salt became dry.

Figure. 5 .

 5 Figure. 5. Salt plug inward motion, evidenced with the large displacement of the left salt crystal that comes to an end with bubble nucleation.

Figure 6 .

 6 Figure 6. Salt traction during the 7 days after both salt plugs precipitated, trapping the saline solution. At the top: pictures of four stages of the experiment. Below: closer views of the salt plugs: the crystal on the right (closer view 2) first dissolved (step b) and then slightly reprecipitated (step d). At the bottom: illustration of the first three stages of the experiment for a better understanding. Step (a): salt plugs precipitated and entrapped the saline (NaCl) solution. Step (b): after 7 days, the crystal on the left has moved inward over 3.3 mm, while the one on the right has moved over 0.13 mm and been dissolved (see closer view 2). Step (c): gas nucleated in the trapped solution and the salts stopped moving and dissolving.

Figure 7 .

 7 Figure 7. (A) Salt dissolution in the trapped solution after one external side of the crystal (on the right) encounters air photos and interpretative sketch. (a) The NaCl solution evaporates. (b) Bow-tie shape crystal growing in the evaporating solution. (c) After 10 days, the right side of the crystal is dry. (d) Dissolution of the thinnest part of the salt in contact with the trapped solution.

Figure 8 .

 8 Figure 8. (A) System with two trapped compartments of saline solution. (B) Estimation of salt loss and volume of solution outlined by the red line drawn around the crystal (c and d).

Figure 9 .

 9 Figure 9. Absence of visible salt mass change in Na 2 SO 4 double plug systems. Double arrows (red and white) on the closer view show that salt mass had not changed (one color = one size).

Figure 10 .

 10 Figure 10. Crack development observed after a crystal plug dried (a) The tube is cut close to an immersed salt plug. (b) The liquid-air interface retreats and reaches the crystal which becomes dry (c) Cracks grow from the discontinuities on the crystal surface (see closer view in (b)).

Figure 11 .

 11 Figure 11. Thermodynamic change in equilibrium constant with increasing tension in the interacting aqueous solution. The black circle and hollow square show the positions of the extreme liquid tensions accounting for the enhanced dissolution respectively observed with NaCl-long (Fig. 6, 30 MPa) and NaCl-short (Fig. 8, 93 MPa) experiments.

  increased mass concentration of 43 ± 7 µg.mm -3 which corresponds to an increased solubility of 12 ± 2% (

Figure 12 .

 12 Figure 12. Changes in of two salts after the bubble nucleates, relaxing the solution's superheating state. The vertical black line at the bottom right is 7 µm thick and is drawn to show the maximum expectable thickness of the reprecipitation layer (see text).

Table 1 . Summary of dual-porosity systems and major observables (also suppl. info.).

 1

							No final gas nucleation
	Exp type	Number of exp	Solid motion	Salt dissolution	Gas nucleation	Time before gas nucleation	No changes	Only solid motion
	NaCl-short	26	0	16	20	2 days to 3 months	6
	NaCl-long	12	4	1	5	4 days to 3 months	5	2
	Na 2 SO 4	10	0	0	10	20 days to 3 months	0
	Total	48	4	17	35	-	11	2

ACKNOWLEDGMENTS

This work has received support from the French Agency for Research (Agence Nationale de la Recherche, ANR) through the grant CONGE BLAN-610-01, the Equipex Planex ANR-11-EQPX-36 and the labex Voltaire ANR-10-LABX-100-01. We are grateful to T. Dudock de Wit, C. Steefel, C.

Tournassat and two anonymous referees who greatly helped us to structure and rewrite an initial draft.

We would like to thank K. Tkaczyk, for polishing the English.


Rachid Tazi-Mezalek

Diane Frankel

Marc Fortin

Elise Kaspi

Julien Guinde

Alexandra Assolen

Sophie Laroumagne

Andrée Robaglia-Schlupp

Hervé Dutau

Patrice Roll

Andree Robaglia

Herve Dutau

Philippe Astoul
email: pastoul@ap-hm.fr

Chest ultrasonography to assess the kinetics and efficacy of talc pleurodesis in a model of pneumothorax: an experimental animal study

niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

The treatment of recurrent malignant pleural effusion or pneumothorax frequently aims to obliterate the pleural cavity to prevent recurrences. Pleurodesis is the symphysis of the pleural space by a fusion of the visceral and parietal pleura. It can be achieved by mechanical abrasion or injection of a sclerosing agent into the pleural space during a thoracoscopy (talc poudrage (TP)) or through a chest tube (talc slurry (TS)) [START_REF] Light | Pleural sclerosis for the treatment of pneumothorax and pleural effusion[END_REF][START_REF] Light | Pleural diseases. 3rd Edn[END_REF]. Sterile, medical-grade talc has been used as an effective and safe sclerosing agent for a long time [START_REF] Bethune | Pleural poudrage: new technique for the deliberate production of pleural adhesion as preliminary to lobectomy[END_REF][START_REF] Janssen | Safety of pleurodesis with talc poudrage in malignant pleural effusion: a prospective cohort study[END_REF]. It induces parietal and visceral inflammation leading to granuloma formation, pleural adhesions and fibrosis [START_REF] Rodriguez-Panadero | Mechanisms of pleurodesis[END_REF].

It is difficult to predict the efficacy of pleurodesis early after talc instillation. In malignant pleural effusion, pleural liquid reconstitution after talc application is a manifest sign of recurrence and failure of the pleurodesis. Chest radiography (CXR) demonstrates the recurrence of the effusion but cannot indicate if pleurodesis has occurred or not. Computed tomography (CT) also cannot predict the effectiveness of pleurodesis early after the procedure. However, talc is detected on CT as high-attenuation clusters along the pleura or into the fissure, characteristically described as split pleura sign [START_REF] Narayanaswamy | CT appearances of talc pleurodesis[END_REF]. Nevertheless, these features do not predict the success of pleurodesis. Conversely, the persistence of an air leak through the chest drain after the procedure is associated with a pneumothorax on imaging (CXR and/or CT) can predict procedure failure. Residual pneumothorax indicates an absence of the lung re-expansion. Finally, CT and CXR indirectly confirm the failure of pleurodesis by the presence of air or liquid in the pleural space, but cannot predict if the symphysis has occurred within the days following the procedure. Finally, pleural thickening and granulomas induced by talc pleurodesis are avid on fluorine-18 fluorodeoxyglucose positron emission tomography (PET), and can mimic neoplastic diseases even 20 years after the procedure [START_REF] Vandemoortele | Positive FDG-PET/CT of the pleura twenty years after talc pleurodesis: three cases of benign talcoma[END_REF]. Thus, PET is also not a reliable method for the assessment of talc pleurodesis.

Chest ultrasonography (CUS) has gained popularity in the last decade for the investigation of pleural diseases and the assessment of pleural procedures [8,[START_REF] Jones | Ultrasound-guided thoracentesis: is it a safer method[END_REF]. Small amounts of pleural fluid or air can easily be detected by sonography with more accuracy than by CXR [START_REF] Eibenberger | Quantification of pleural effusions: sonography versus radiography[END_REF][START_REF] Lichenstein | A bedside ultrasound sign ruling out pneumothorax in the critically ill: lung sliding[END_REF][START_REF] Soldati | Occult traumatic pneumothorax: diagnostic accuracy of lung ultrasonography in the emergency department[END_REF]. In fact, CUS has been proven to be a reliable imaging modality for the evaluation of a wide variety of chest diseases [START_REF] Havelock | Pleural procedures and thoracic ultrasound: British Thoracic Society Pleural Disease Guideline[END_REF]. Normal movement of the parietal pleura over the visceral pleura during the respiration cycle is recognised as the gliding sign [START_REF] Targhetta | Ultrasonic signs of pneumothorax: preliminary work[END_REF] or lung sliding sign [START_REF] Lichenstein | A bedside ultrasound sign ruling out pneumothorax in the critically ill: lung sliding[END_REF]. The absence of the lung sliding has been used to diagnose pneumothorax, in combination with the repetition of reverberation A lines, the lack of lung pulse and the presence of a lung point [START_REF] Volpicelli | International evidence-based recommendations for point-of-care lung ultrasound[END_REF]. Moreover, identification of pleural thickening as a hypoechoic structure is possible by CUS in the presence or lack of pleural effusion. Pleural thickening and pleural plaques appear as hypoechoic structures typically found along the posterolateral part of the chest. Finally, calcification can occur and appears with posterior acoustic shadows [START_REF] Wernecke | Sonographic features of pleural disease[END_REF][START_REF] Hallifax | State-of-the-art: Radiological investigation of pleural disease[END_REF].

Talc application aims to create a symphysis of the pleural space. Consequently, the sliding of both pleura will be reduced or vanished on CUS examination if the procedure has been successful. CUS has already been demonstrated as an efficient imaging modality for the evaluation of pleurodesis in rabbits [START_REF] Zhu | Efficacy of ultrasound in the diagnosis of pleurodesis in rabbits[END_REF].

No device and no method are currently available at the bedside to assess directly and to monitor the effects of pleurodesis. The objective of our study was to evaluate, in pigs, the capacity of CUS to predict the quality of a pleurodesis induced by TP and TS, and to compare the effectiveness of both techniques.

Materials and methods

Study design

The present study was conducted in compliance with the recommendations of the Federation of European Laboratory Animal Science Associations and was approved by the animal care committee of Aix-Marseille University (Marseille, France). All the animals ( pigs) were submitted to a strict veterinary protocol of anaesthesia: use of intramuscular premedication with ketamine and azaperone, intravenous prophylactic cefazolin before the procedure, intravenous propofol 2% during the procedure and intravenous tramadol before the talc application. During the procedure (thoracoscopy) animals were connected to mechanical ventilation through a veterinary oxygen mask. Systematic monitoring of blood pressure, cardiac pulse and saturation of oxygen was performed. Once the procedure ended, animals were left to recover in an individual enclosure with appropriate temperature, food, toys and veterinary surveillance.

The animals were randomised into two groups. In the first group, TP was performed first on the right side followed by TS on the left side 1 week later. In the second group, the sides were reversed.

Artificial pneumothorax was first induced on the operating table with a 2-mm Boutin's trocar (Novatech SA, La Ciotat, France) under direct CUS guidance and using local anaesthetic (lidocaine 2%). Successful introduction of air into the pleural space was confirmed audibly and on real-time ultrasonography. Blunt dissection of the chest wall, 7-mm trocar placement and direct-view rigid thoracoscope (Richard Wolf, Reims, France) insertion were performed safely, allowing examination of the pleural cavity.

For the TP technique, 2 g sterile French calibrated talc (Stéritalc; Novatech SA) were applied into pleural cavity with a double balloon insufflator (Richard Wolf ) as previously described [START_REF] Tschopp | ERS task force statement: diagnosis and treatment of primary spontaneous pneumothorax[END_REF][START_REF] Bridevaux | Short-term safety of thoracoscopic talc pleurodesis for recurrent primary spontaneous pneumothorax: a prospective European multicentre study[END_REF]. A 24 Fr chest tube (Vygon, Ecouen, France) was placed apically and connected to progressive wall suction (maximum of -50 cmH 2 O) for lung expansion. The chest tube was then removed.

For the TS technique, a dilution of 2 g sterile graded talc was mixed with 50mL saline and 5mL lidocaine 2% according to the literature [START_REF] Light | Pleural sclerosis for the treatment of pneumothorax and pleural effusion[END_REF][START_REF] Light | Pleural diseases. 3rd Edn[END_REF], and injected through the chest tube which was placed apically and clamped for 1 h. The chest tube was connected to progressive wall suction until the air leak stopped and was then removed.

Thoracic ultrasonographic evaluation

Nine points on each hemithorax were pre-defined for CUS examination: three points (superior, middle and inferior) along the anterior, mid-and posterior axillary lines (figure 1a). They were tattooed using subcutaneous permanent ink (figure 1b) to allow repetitive ultrasonographic examination during the study. Sonography (EnVisor C HD; Philips, Suresnes, France) was performed by experienced interventional pulmonologists blind to the talc technique used. All nine points were analysed by ultrasonography and classified as pneumothorax, pleural effusion, lung sliding or pleural symphysis (table 1). A total of 108 points of the chest were evaluated by ultrasonography during the study. Pleural thickening was measured in millimetres at the nine pre-defined points for each animal with high-frequency probe in B Mode. CUS was performed before, immediately after (day 0), and at days 1, 3, 14 and 21, on each side. Finally, autopsy was performed after sacrifice with a bolus of propofol and intravenous potassium chloride injection at day 21 according to the recommendations.

Pleurodesis evaluation

At the autopsy, a macroscopic pleurodesis score, initially proposed by LIGHT et al. [START_REF] Light | Talc slurry is an effective pleural sclerosant in rabbits[END_REF] and COLT et al. [START_REF] Colt | A comparison of thoracoscopic talc insufflation, slurry, and mechanical abrasion pleurodesis[END_REF], was used to grade the symphysis semiquantitatively (table 2). Investigators who performed the dissection of the pleura were blinded to the procedure previously made. The nine marked points were removed as a block (muscles, connective tissue, parietal pleura, adhesion and visceral pleura with underlying lung parenchyma) after a meticulous dissection in order not to rupture the symphyses that may have occurred.

A microscopic evaluation (histological examination) was performed for three points (superior, middle and inferior) of the midaxillary line as a predefined consensus. A total of 36 points (18 points for each procedure, TP and TS) among the 108 evaluated points of the study were analysed histologically. Tissue specimens were fixed in formalin 4% and embedded into paraffin. Finely cut sections of 7-μm thickness were placed on slides and stained with haematoxylin-eosin and Masson trichrome. The following pathological items were analysed: pleural symphysis, foreign body granuloma, inflammation, presence of talc particles and integrity of mesothelial cell sheet. The pathologists were blind to the procedure performed before and to the ultrasonographic findings.

Statistical analysis

Statistical analysis was performed with the Prism software (GraphPad Software Inc., San Diego, CA, USA). Significant differences were determined using nonparametric or parametric paired tests (Wilcoxon test or paired t-test, respectively) and Fisher's exact test. A p-value <0.05 was considered as significant.

Results

Six white Landrace pigs of 30±5 kg were submitted to both techniques. At baseline, all the pigs had a normal CUS exploration showing evident lung sliding and no area of pleural thickening.

A statistically significant difference in lung sliding between both techniques was observed starting from 72 h (day 3) after the procedure ( p=0.005) (figure 2a). TP achieved a higher sonographic score corresponding to reduced lung sliding (table 1). Symphysis was obtained at most of the points evaluated.

No statistically significant difference was observed between both techniques for pleural thickness except immediately after the procedure. Pleural thickness was higher after TP compared to TS at day 0 ( p=0.04) (figure 2b).

At autopsy (day 21), macroscopic symphysis at the defined points was evaluated during the dissection. Pleural adhesion was rarely observed after TS (figure 3a), whereas pleural adhesions were clearly visible after the TP technique (figure 3b). The macroscopic pleural score was higher for TP than TS ( p=0.0011) (figure 3c) and the representation of the pleural adhesions after the dissection showed that macroscopically, TP led to a higher number of pleurodesis areas than TS (figure 3d). Furthermore, we found a positive correlation between the 108 points evaluated by CUS prior to the sacrifice and macroscopic findings ( p<0.0001, r=0.55; Spearman test).

36 points were analysed at the histological level (figure 4). Granulation tissue formations were present in specimens from both techniques. Pleural symphysis was observed in 12 (66.7%) out of 18 points for TP and in five (27.8%) out of 18 points for TS with a significant difference ( p=0.044, Fisher test). The fusion between visceral and parietal pleura with a disruption or lack of the mesothelial cells sheet was statistically significant between both techniques ( p=0.018, Fisher test). We found the presence of a mesothelial cell sheet in five (27.8%) of 18 points for TP and in 13 (72.2%) of 18 points for TS. No significant differences were observed between the two techniques related to the presence of talc, inflammatory reaction and presence of granulomas (table 3).

For all these results, no difference was shown between the two groups of animals depending on the side of the TP and TS procedures.

Discussion

The management of malignant pleural effusion and recurrent pneumothorax are common clinical problems. These situations are frequently treated by an attempt to create a pleurodesis with TS or TP. After talc administration, the fusion of the visceral and parietal pleura should prevent the pleural surfaces from gliding over. CUS has recently emerged as an efficient technique to analyse pulmonary and pleural structures. The evaluation of lung sliding after talc application could be indicative of the effectiveness of a pleurodesis. In our study, we used an animal model ( pig) to evaluate the capacity of CUS to predict the quality of pleurodesis induced by talc after the creation of a pneumothorax and we compared the effectiveness of two technics, TP and TS. We confirm the efficacy of CUS to predict and to easily follow pleurodesis constitution after talc administration. We also identified TP as a superior technique to TS for the induction of pleurodesis in the context of pneumothorax. CXR and CT are usually used as standard techniques to identify pleural diseases. Typically, pleural and extrapleural lesions have obtuse angles with the chest wall, contrary to subpleural processes in the pulmonary parenchyma, in which the angle is more likely to be acute. Pleural plaque and thickening are easily identified as symmetric, smooth and soft-tissue density on CT scans. In recent years, ultrasonography has emerged as a new approach to evaluate chest diseases and it is now the method of choice in the guidance of pleural procedures [START_REF] Havelock | Pleural procedures and thoracic ultrasound: British Thoracic Society Pleural Disease Guideline[END_REF]. It has the advantage of providing real-time imaging of the parietal and visceral pleura, and demonstrating the presence or absence of lung sliding. Sonography is accurate in detecting pleural adhesions. Moreover, its use prior medical thoracoscopy facilitates trocar insertion and prevents visceral injuries [START_REF] Sasaki | Preoperative detection of pleural adhesions by chest ultrasonography[END_REF][START_REF] Wei | Use of transthoracic ultrasound to predict pleural adhesions: a prospective blinded study[END_REF][START_REF] Cassanelli | Accuracy of transthoracic ultrasound for the detection of pleural adhesions[END_REF]. Finally, CUS has been proposed to detect pleurodesis after talc application, as demonstrated in animal models by an absence of the lung sliding sign [START_REF] Zhu | Efficacy of ultrasound in the diagnosis of pleurodesis in rabbits[END_REF].

In our study, during dissection, we observed more macroscopic symphyses at the pre-defined points after TP than TS. Adhesions following TS were logically present in areas near the distal end of the chest tube and in gravity-dependent areas, while adhesions after talc insufflation were more uniform. We believe that obliteration of the pleural space would be more effective if the application of talc is more diffuse and if a larger pleural surface is treated, as induced by TP.

In accordance with several studies [START_REF] Rodriguez-Panadero | Mechanisms of pleurodesis[END_REF][START_REF] Zhu | Efficacy of ultrasound in the diagnosis of pleurodesis in rabbits[END_REF][START_REF] Sasaki | Preoperative detection of pleural adhesions by chest ultrasonography[END_REF][START_REF] Wei | Use of transthoracic ultrasound to predict pleural adhesions: a prospective blinded study[END_REF][START_REF] Cassanelli | Accuracy of transthoracic ultrasound for the detection of pleural adhesions[END_REF][START_REF] Leo | Can chest ultrasonography assess pleurodesis after VATS for spontaneous pneumothorax?[END_REF], the present work demonstrates that ultrasonography is an accurate imaging modality to predict pleural adhesions and to identify zones of ineffective pleurodesis. A significant concordance rate between ultrasonographic evaluation and macroscopic findings was found.

Consequently, the quality of the symphysis comparing both techniques seems to be nicely assessed by ultrasonography, although more studies are needed to confirm our results.

There is a general agreement that application of talc into the cavity creates diffuse inflammation, an imbalance of pleural coagulation and fibrinolysis, fibroblast proliferation, and collagen production. Pleural mesothelial cells, targeted by the talc, are the cornerstone of the pleurodesis process, releasing multiple molecules such as interleukin-8, transforming growth factor-β and fibroblast growth factor [START_REF] Hartman | Thoracoscopic talc insufflation increases pleural fluid IL-8 levels in patients with malignant pleural effusions[END_REF]. Following talc administration, there is a rapid neutrophil influx into the pleural space and activation of an inflammatory cascade reaction [START_REF] Rodriguez-Panadero | Mechanisms of pleurodesis[END_REF][START_REF] Hartman | Thoracoscopic talc insufflation increases pleural fluid IL-8 levels in patients with malignant pleural effusions[END_REF]. In our study, a statistically significant difference in the thickness measured by CUS was observed between both talc techniques in favour of TP, but only immediately after the procedure. This may be explained by the insufflation of the talc and its homogenous deposit over the pleural surface, which could produce an immediate diffuse and transient pleural inflammation, detected by ultrasonography as an increase in the pleural thickness.

Furthermore, a higher sonographic score was observed from the third day after talc instillation, corresponding to a persistent and substantial grade of symphysis. Histopathological changes caused by intrapleural talc administration are well described by others but to our knowledge, no study has assessed the chronology of the direct effect of the talc, in particular by using ultrasonography. In our daily practice, for the management of recurrent spontaneous pneumothorax, we rigorously wait until the third day for chest tube removal after talc poudrage, without any evidence-based data. Sonography represents a potential new tool to assess the kinetics of pleural symphysis and to predict the time of chest tube removal. However, more studies are needed to confirm these data.

The dissection shows that macroscopically, TP leads to a higher number of pleurodesis areas in comparison to TS. It is common sense to consider that the distribution of talc particles after insufflation under visual control during a thoracoscopy (TP) is more homogeneous and diffuse than the injection of a solution through a chest tube (TS), and consequently the distribution of the symphysis remains similar.

Based on these results, TP is probably more efficient than TS but this hypothesis should be demonstrated in large, randomised studies. Moreover, the fusion between the visceral and parietal pleura with a disruption of the mesothelial cell sheet observed in histological analysis was significantly different between both techniques, in favour of TP. However, no significant differences were observed between the two techniques in the presence of talc, inflammatory reaction or the presence of granulomas. The histological findings suggest that pleurodesis has no effect on the lung parenchyma itself. These data are consistent with previous reports of the safety and efficacy of talc pleurodesis [START_REF] Bethune | Pleural poudrage: new technique for the deliberate production of pleural adhesion as preliminary to lobectomy[END_REF][START_REF] Bridevaux | Short-term safety of thoracoscopic talc pleurodesis for recurrent primary spontaneous pneumothorax: a prospective European multicentre study[END_REF][START_REF] Colt | A comparison of thoracoscopic talc insufflation, slurry, and mechanical abrasion pleurodesis[END_REF][START_REF] Hartman | Thoracoscopic talc insufflation increases pleural fluid IL-8 levels in patients with malignant pleural effusions[END_REF][START_REF] Fraticelli | Distribution of calibrated talc after intrapleural administration: an experimental study in rats[END_REF].

Despite all these encouraging results, there are several limitations to our study. 1) Pleurodesis could not be achieved within a few centimetres of the diaphragm. This may be due to the choice of the lower point of the chest ( points 3, 6 and 9) and to the constant movement of the diaphragm with breathing. Not surprisingly, pleural adhesion distribution was less effective close to the diaphragm for both techniques.

2) We chose to reduce the microscopic analysis for 36 of the 108 defined initial points. We believe that microscopic evaluation will not change from one animal to another or between fibrotic and nonfibrotic areas.

3) This experimental study was limited to six animals (both side of the thorax). However, significant and encouraging results were obtained, which may be proved in a bigger cohort. 4) We used healthy pigs and created an artificial pneumothorax to study the pleurodesis, representing an experimental model of pneumothorax taking into account that in such large animals, the thicknesses of the visceral and parietal pleural are similar to the pleura in human, which is not true for small animals as rodents [29]. The same experimental design might be used using a model of malignant pleural effusions. 5) Pleural thickening is measured using a high-frequency probe in B mode. This measure is an operator-dependent datum and it will certainly difficult to evaluate in the malignant pleural effusion model.

Finally, we believe that the results of this experimental study have clinical implications. Indeed, ultrasonography could have multiple applications in humans, such as to demonstrate the quality of a talc pleurodesis, to compare the effectiveness of the sclerosing technique (TP versus TS), to study the kinetic of pleural symphysis and to assess with certainty the time for chest tube removal.

Conclusions

Sonography is a reliable tool used by pulmonologists to provide real-time imaging guidance for pleural procedures. Talc pleurodesis by poudrage or slurry can be easily assessed and repeated by ultrasonography days after the procedure. The quality of the pleurodesis and kinetics of the pleural symphysis can be evaluated indirectly by ultrasonography. It helps to detect possible recurrences and plans to remove properly the chest tube. Talc pleurodesis by poudrage seems to be a better technique to induce pleurodesis, as observed in our animal study. To confirm our results, comparison of pleurodesis efficacy and quality between slurry and poudrage need now to be compared using CUS in large, randomised clinical studies.

FIGURE 1

 1 FIGURE 1 Chest ultrasonography design. a) Schematic representation of the nine points on a hemithorax. b) Points tattooed with subcutaneous permanent ink for chest ultrasound follow-up.

FIGURE 2 FIGURE 3 FIGURE 4

 234 FIGURE 2 Induced pleurodesis depending on the two techniques. a) Comparison of the sonographic score over the study. A statistically significant difference for lung sliding between both techniques is observed starting from 72 h (day 3) after the procedure. Talc poudrage achieved a higher sonographic score than talc slurry. Day 3: p=0.005; day 14: p=0.0075; day 21: p<0.0001 (Wilcoxon test). b) Comparison of the pleural thickness evaluated by chest ultrasonography over the study. No statistically significant difference in the pleural thickness was observed between the techniques except immediately after the procedure: at day 0, pleural thickness was higher after poudrage than slurry ( p=0.04, paired t-test). *: p<0.05; **: p<0.01; ***: p<0.001.

TABLE 2

 2 Macroscopic grading score of the symphysis

	Grade	Pleurodesis
	0	Normal pleural space and lung
	1	No adhesion but pleural space inflamed as evidenced by redness and fibrin deposition
	2	Few scattered adhesions
	3	Generalised scattered adhesions
	4	Complete obliteration of the pleural space by adhesions

TABLE 1

 1 Grading of sonographic features

	Grade Value	Ultrasonic features
	0	Pneumothorax	B mode: lung sliding absent, repetitive A lines present, lung point present, no fluid detectable and B lines absent
			M mode: seashore sign absent and stratosphere sign present
	1	Pleural effusion	B mode: hypoechoic structure or echo-free space between both pleura, and gliding sign absent
			M mode: sinusoid sign if a small amount of liquid
	2	Lung sliding	B mode: gliding sign and B lines present
			M mode: seashore sign present
	3	Pleural symphysis B mode: gliding sign questionable or absent, B lines absent, and pleural thickening with posterior enhancement
			M mode: seashore sign absent

https://doi.org/10.1183/23120541.00158-2017

TABLE 3

 3 Results of histological examination

	Talc poudrage	Talc slurry	p-value

https://doi.org/10.1183/23120541.00158-2017

https://doi.org/10.1183/23120541.00158-2017

PNEUMOTHORAX | R. TAZI-MEZALEK ET AL.

Acknowledgements: We are indebted to S. Berdah and his team (Marie-Ange and Audrey) from the Surgical Teaching and Research Center (CERC, Faculty of Medicine of the North Hospital, Aix-Marseille University, Marseille, France) for their skilled technical assistance during this study.

Conflict of interest: None declared.