Thuy Minh Giang

Sebastien Gaucel

Pierre Brestaz

Marc Anton

Anne Meynier

Ioan-Cristian C Trelea

Steven Le Feunteun
email: steven.le-feunteun@grignon.inra.fr

Dynamic modeling of in vitro lipid digestion: Individual fatty acid release and bioaccessibility kinetics

Keywords: Emulsion, Digestion, Pancreatic lipase, Substrate specificity, DHA, Simulation

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

Introduction

Human digestion of lipid emulsion is influenced by both physiological parameters and emulsion properties. Triacylglycerol (TAG) digestion occurs essentially in the intestine where about 80% of the lipolysis reaction take place [START_REF] Carriere | Secretion and contribution to lipolysis of gastric and pancreatic lipases during a test meal in humans[END_REF]. The reaction is mediated by the pancreatic lipase-colipase complex at the oil-water interface and releases the sn-2-monoacylglycerol (MAG) and two free fatty acids (FFA) [START_REF] Golding | The influence of emulsion structure and stability on lipid digestion[END_REF]. Lipolysis products are then incorporated into bile salt micelles before being transported to the gut wall and absorbed by the organism [START_REF] Smith | The digestive system[END_REF]. Hence, physiological parameters such as the concentrations of pancreatic lipase, colipase and bile salts can all modify lipid digestion. The structural characteristics of emulsions are also important to consider when evaluating lipolysis kinetics. For instance, the nature of emulsifiers influences the properties of the oil-water interface and can therefore affect lipase adsorption onto droplets. The size of the oil droplets is another key parameter since fine emulsions with high interfacial areas facilitate the enzyme activity compared to coarse emulsions with low interfacial areas [START_REF] Armand | Digestion and absorption of 2 fat emulsions with different droplet sizes in the human digestive tract[END_REF][START_REF] Golding | The influence of emulsion structure and stability on lipid digestion[END_REF][START_REF] Li | New mathematical model for interpreting pH-stat digestion profiles: impact of lipid droplet characteristics on in vitro digestibility[END_REF].

The TAG composition in fatty acid residues (FA), characterized by their carbon chain length and degree of unsaturation, also has a strong impact. Several studies have demonstrated that human and porcine pancreatic lipases exhibit a certain fatty acid specificity [START_REF] Berger | Lipases in organic solvents: The fatty acid chain length profile[END_REF][START_REF] Desnuelle | Specificities of Lipases[END_REF][START_REF] Mukherjee | Substrate specificities of lipases in view of kinetic resolution of unsaturated fatty acids[END_REF][START_REF] Yang | Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination[END_REF]. For instance, it is well known that pancreatic lipase is more active on medium chain TAG (MCT, up to C10) than on long chain TAG (LCT, from C12) [START_REF] Armand | Effects of droplet size, triacylglycerol composition, and calcium on the hydrolysis of complex emulsions by pancreatic lipase: an in vitro study[END_REF][START_REF] Desnuelle | Specificities of Lipases[END_REF][START_REF] Golding | Impact of gastric structuring on the lipolysis of emulsified lipids[END_REF][START_REF] Li | New mathematical model for interpreting pH-stat digestion profiles: impact of lipid droplet characteristics on in vitro digestibility[END_REF], and that its activity depends on the number and positions of unsaturations [START_REF] Mukherjee | Substrate specificities of lipases in view of kinetic resolution of unsaturated fatty acids[END_REF][START_REF] Yang | Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination[END_REF]. Long chain polyunsaturated fatty acids, such as docosahexaenoic acid (C22:6 n-3, DHA), have also been reported to be among the most resistant FA to pancreatic lipase [START_REF] Bottino | Resistance of certain long-chain polyunsaturated fatty acids of marine oils to pancreatic lipase hydrolysis[END_REF][START_REF] Yang | Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination[END_REF], possibly because of the short distance of the first double bond from the ester linkage [START_REF] Akanbi | Pancreatic lipase selectively hydrolyses DPA over EPA and DHA due to location of double bonds in the fatty acid rather than regioselectivity[END_REF][START_REF] Bottino | Resistance of certain long-chain polyunsaturated fatty acids of marine oils to pancreatic lipase hydrolysis[END_REF][START_REF] Lawson | Human absorption of fish oil fatty acids as triacylglycerols, free acids, or ethyl esters[END_REF].

Moreover, the extent of FFA and MAG that can be solubilized into the bile salt micelles also seems to be product dependent [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF][START_REF] Hofmann | The behavior and solubility of a number of pure monoglycerides in dilute, micellar bile-salt solution[END_REF]. For instance, the molar saturation ratio (mole of incorporated products/mole of bile salts) in a sodium glycodeoxycholate solution at 37°C has been reported to vary from 0.07 for stearic acid (C18:0) to 1.86 for lauric acid (C12:0) with intermediate values for long chain unsaturated FFA such as oleic (C18:1 n-9) and linoleic acids (C18:2 n-6) [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF]. Such values, which can be partly reasoned in terms of molecular polarities and amphiphilic properties, are however dependent on numerous parameters such as the type of bile acid used or the pH of the solution [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF][START_REF] Hofmann | The behavior and solubility of a number of pure monoglycerides in dilute, micellar bile-salt solution[END_REF]. Moreover, if the addition of sn-1-monoolein within the bile salt solution has been reported to improve the bile salt solubility of FFA, the opposite has been observed with the addition of oleic acid [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF]. Given the variety of lipolysis products and bile acids that coexist during the intestinal digestion of edible oil, it therefore seems difficult to predict the bioaccessibility of lipolysis products.

Modeling has proven to be a powerful tool for a better understanding of the intestinal digestion kinetics. Various models of lipid digestion have been published over the last decades. Several models are based on the Michaelis-Menten equation. Verger and co-workers were the first to adapt such approach to the biphasic nature of the lipolysis reaction by taking into account the interfacial concentrations of the substrates and enzymes [START_REF] Verger | Interfacial enzyme kinetics of lipolysis[END_REF][START_REF] Verger | Action of Phospholipase A at Interfaces[END_REF]. Several variants have been proposed since, as for instance by [START_REF] Jurado | Kinetics of the enzymatic hydrolysis of triglycerides in o/w emulsions[END_REF] for a bacterial lipase. These models are however quite complex and often require parameter values that are difficult to determine experimentally, in particular for the digestion of edible oils composed of different TAG species. In a much simpler approach, intestinal lipolysis can also be described by a first order kinetic model [START_REF] Ye | Effect of calcium on the kinetics of free fatty acid release during in vitro lipid digestion in model emulsions[END_REF]. If such models can be useful in comparing the initial enzyme activity in different conditions, they do not take into account the interfacial area of oil droplets. This is why [START_REF] Li | New mathematical model for interpreting pH-stat digestion profiles: impact of lipid droplet characteristics on in vitro digestibility[END_REF] proposed a model in which the rate constant is expressed per unit of interfacial area, and that accounts for a progressive decrease of the oil droplet size. This model provided good fits of pH-stat measurements performed on different emulsions with the assumptions that the number of droplet remains unchanged and all droplets have the same size at any given time. Recently, we resorted to a similar modeling approach to show, by integrating experimentally measured droplet sizes, that the reduction of the interfacial area induced by droplet coalescence was the main reason why our in vitro intestinal lipolysis seemed to stop before the substrate was fully exhausted [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF].

Nevertheless, all the above models assume a unique reaction rate constant without considering the FA composition of the studied oils. Hence, they cannot be used to reproduce the release kinetics of individual FA. In the present study, we develop a mathematical model of intestinal lipolysis that takes the composition of the oil substrate into account. It predicts the lipolysis kinetics of each FA residue and their in vitro bioaccessibility, defined as the concentration of FA (FFA + sn-2-MAG) measured within the bile salt micellar phase. The model is applied to a set of experimental data measured during the in vitro gastro-intestinal digestion of a whey protein stabilized emulsion. The estimated values for the model parameters (FA specific lipolysis rate constants and bile salt micellar fractions) are compared with the available literature and discussed according to their biochemical meanings.

Material and method

Materials

The oil containing medium-chain triacylglycerols (MCT), Miglyol 812S, was purchased from Sasol GmbH, Germany. The main fatty acids contained in this oil were C8:0 (54% w/w) and C10:0 (43%). The oil containing long-chain triacylglycerols (LCT), DHAsco, was obtained from Martek, via DSM Nutritional Products Ldt, Switzerland. It contained docosahexahenoic acid (DHA, C22:6 n-3) as major fatty acid (40% w/w) along with C12:0 (4%), C14:0 (12%), C16:0 (12%) and C18:1 n-9 (24%). Whey protein powder (Prolacta 95) was purchased from Lactalis Ingredients, France. Pepsin (P7012, 2,500 U mg -1 ), mucin (M2378), pancreatin (P7545, 8×USP specifications), pancreatic lipase (L3126, 120 U mg -1 ) and bile extract (B8631) were from porcine origin and obtained from Sigma-Aldrich, France. Water was Milli-Q water.

Solvents for liquid chromatography were chloroform for HPLC (Carlo Erba), methyl alcohol for HPLC (99.9%, Carlo Erba) and ammonia solution (30%, Carlo Erba). Acetone (Pure RE, Carlo Erba), sulfuric acid (RPE 96%, Carlo Erba), cyclohexane and heptadecanoic acid (Sigma) were used for the preparation of fatty acid methyl esters (FAME) of the micellar phase during intestinal digestion.

Emulsion preparation and digestion

A whey protein stabilized emulsion with droplet diameters below 1 µm was first prepared according to the procedure previously described in [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF]. Briefly, the oil phase (20% w/w) contained 37.5% (w/w) of MCT and 62.5% of LCT, and the aqueous phase (80% w/w) contained 4% (w/w) of whey protein powder, used as emulsifier, in a 0.1M sodium phosphate buffer (pH 7.0). After high pressure homogenization and addition of maltodextrin solution to improve the freeze-dried stability of oil droplets, the emulsion was freeze-dried.

On the day of the in vitro experiments, the dry emulsion was rehydrated in Milli-Q water to obtain a final oil concentration of 3.2% (w/w).

A volume of 3 mL of the rehydrated emulsion, corresponding to an oil mass of about 96 mg, was placed into hermetically sealed headspace vials (22.4 mL). The gastric and intestinal phases of the in vitro digestion experiments were performed sequentially at 37°C under magnetic stirring (400 rpm). The gastric phase duration was 60 min and was launched by adding 2.12 mL of simulated gastric fluid (SGF) and 40 µL of 1M HCl to reach a final pH of 2.5. The SGF solution contained 3.9 g L -1 of pepsin (corresponding to 10,000 U per mL of SGF using hemoglobin as substrate), 2.4 g L -1 of mucin, 120 mM of NaCl, 2 mM of KCl and 6 mM of CaCl 2 . The intestinal phase duration was then launched for 300 min maximum by adding 4.86 mL of simulated intestinal fluid (SIF) and 100 µL of 1M NaCO 3 to reach a final pH of 6.5. The SIF solution contained 30.8 g L -1 of bile extract powder, 0.82 g L -1 of pancreatin (activity: 8×USP specifications), 0.41 g L -1 of pancreatic lipase (corresponding to 100 U per mL of SIF using olive oil as the substrate), and the same electrolyte concentrations as the SGF.

Samples were taken at t = 0 min of the intestinal phase using a modified SIF that contained all constituents except pancreatin and lipase, and at 15, 30, 60, 120 and 300 min of intestinal digestion. One vial was used for one sampling time and one type of measurement (quantification of LCT and MCT by HPLC, quantification of the lipolysis products within the bile salt micellar phase by GC, or droplet size by laser granulometry) so that the contents of 18 vials in total were analyzed (3 methods times 6 sampling times) for one digestion. Three independent digestion experiments, further denoted as replicates, were performed.

Quantification of LCT and MCT by HPLC

HPLC paired with an evaporative light scattering detector (ELSD) was used to quantify the decrease in both LCT and MCT masses during the time course of the in vitro digestions. Total lipids were extracted according to Bligh and Dyer method with minor modifications in the ratio CHCl 3 /CH 3 OH/H 2 O 1/2/1, and dissolved in CHCl 3 to obtain a final lipid concentration of 0.7 mg mL -1 . HPLC operating conditions were similar to those described in Kenmogne-Domguia, Meynier, Viau, Llamas, & Genot (2012) using a Uptip-prep Strategy column (2.2 µm SI, 150×4.6 mm, Interchim, Montluçon, France) and 30 µL of injected lipid extract. The signal of LCT and MCT (retention times of 1.21 and 1.32 min, respectively) were deconvoluted using a specifically developed algorithm running with the Matlab™ software (The MathWorks Inc., Natick, MA), and converted into masses using a pre-established calibration curve. LCT and MCT masses were finally converted into lipolysis percentages using Equation 1:

𝑙𝑖𝑝𝑜𝑙𝑦𝑠𝑖𝑠 (𝑡) = 𝑚 𝑇𝐴𝐺 0 -𝑚 𝑇𝐴𝐺 (𝑡) 𝑚 𝑇𝐴𝐺 0 × 100 % (1)
where 𝑚 𝑇𝐴𝐺 0 and 𝑚 𝑇𝐴𝐺 (𝑡) are the masses (mg) of LCT or MCT initially present in the vials and measured by HPLC at time t, respectively.

Droplet size measurement

The volume-based distribution of oil droplet sizes was measured using a Mastersizer S (Malvern Instruments Ltd., Worcestershire, UK) equipped with a 2 mW He-Ne laser of λ = 633 nm and the 300RF lens with detection limits of 0.05 and 900 µm. The refractive index n 0 of the aqueous phase was 1.33 and the properties of the dispersed phase were 1.457 for the refractive index and 0.001 for the absorption. Samples were diluted with distilled water to reach an oil volume concentration near 0.01% (w/w) for the circulation in the measurement cell. The surface weighted mean diameter, 𝑑 32 , was calculated using Equation 2:

𝑑 32 = ∑ 𝑛 𝑖 3 𝑑 𝑖 3 ∑ 𝑛 𝑖 2 𝑑 𝑖 2 (2)
where 𝑛 𝑖 is the number of droplets of diameter 𝑑 𝑖 .

Quantification of lipolysis products within the bile salt micellar phase by GC

Intestinal aliquots were frozen immediately after sampling. The day after, they were centrifuged at 50,000 rpm for 45 min at 4°C with Aventi J26-XP (Beckman-Coulter). Three phases were observed after centrifugation: a lipidic phase at the top, a pellet at the bottom, and a major intermediate phase containing the bile salt mixed micelles and which will be further referred as the micellar phase. 500 µL of the micellar phase were sampled for the determination of its FA composition by direct trans-methylation as described previously by [START_REF] Berton | Quantification of unadsorbed protein and surfactant emulsifiers in oil-in-water emulsions[END_REF]. Briefly, 500 µL of the micellar phase was trans-methylated in presence of 100 µL of internal standard (heptadecanoic acid 1mg/mL in acetone/MeOH 2/1 v/v). 2 mL of methanol and 400 µL of H 2 SO 4 were then introduced. The mixture was mixed;

the tube screw capped and then heated à 100°C for 1 h. After cooling at room temperature, 500 µL of water and 1 mL of cyclohexane were added. After mixing, and decantation, the organic phase was removed for later analysis by GC. Fatty acid methyl ester (FAME)

analyses were performed on a gas chromatograph paired with a splitless injector and a FID (Clarus, Perkin Elmer). They were separated on a capillary column (DB 225, 30 m X 0.25 mm, film thickness 0.25 mm) after splitless injection. The carrier gas (H 2 ) was set at 2 m min - 1 . Temperature gradient was programmed as follows: 50°C for 3 min, increase to 180°C at 15°C min -1 , 7 min at 180°C, increase to 220°C at 10°C min -1 and finally 10 min at 220°C. The temperature of the injector and the detector were maintained at 250°C. Individual FA were identified by comparison of their retention time with those of standards FAME mix (cat 47885-U 37, Supelco); n-3 PUFA from menhaden oil (cat 4-7085 U-14, Supelco), and marine oil standard of the AOCS. Peak surfaces were integrated and corrected by response factors of individual fatty acids.

The FA composition of the native emulsion was also quantified by GC. As summarized in Table 1, 8 fatty acids representing 95.6% of the total FA mass, were considered for the modeling approach: C8:0, C10:0, C12:0, C14:0, C16:0, C16:1 n-7, C18:1 n-9, and C22:6 n-3.

These 8 FA showed coefficients of variation of less than 10.26% over the three replicate measurements. Since the emulsion was prepared from a mixture of two oils, the last 2 columns of Table 1 provide the fractional contributions of LCT (𝛼 𝐹𝐴 𝑖 ) and MCT (1-𝛼 𝐹𝐴 𝑖 ) for each FA residue. Results show that almost all C8:0 and C10:0 came from the MCT oil, whereas all the other fatty acids came essentially from LCT. These fractions will be used for TAG mass calculations in the model.

Mathematical modeling

Model assumptions and equations

The model was built to predict the lipolysis kinetics and the concentration profiles of the lipolysis products in the bile salt micellar phase, knowing that pancreatic lipase produces two FFA and one sn-2-MAG per TAG molecule. However, if the GC analyses of the samples make it possible to quantify the different acyl chains, they cannot be used to discriminate their molecular origin, i.e. free or esterified fatty acids. In the rest of this paper, the term fatty acid (FA) will therefore refer to FA carbon chains in any kinds of molecular products. The main modeling assumptions were as follows:

A1: the hydrolysis rate of a given fatty acid residue 𝐹𝐴 𝑖 was independent of the snposition within TAG molecules, with 𝐹𝐴 𝑖 referring to the fatty acid considered (C8:0, C10:0, C12:0, C14:0, C16:0, C16:1 n-7, C18:1 n-9, or C22:6 n-3).

A2: the hydrolysis rate of a given fatty acid residue 𝐹𝐴 𝑖 was proportional to the interfacial area occupied by this acyl chain.

𝑑𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (t) 𝑑𝑡 = -𝑘 𝐹𝐴 𝑖 . 𝐴 𝐹𝐴 𝑖 (𝑡) (3)
where 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 is the mass of 𝐹𝐴 𝑖 in the oil droplets (mg), 𝑘 𝐹𝐴 𝑖 is the lipolysis rate constant of 𝐹𝐴 𝑖 (mg m -2 min -1 ), and 𝐴 𝐹𝐴 𝑖 (𝑡) is the interfacial area occupied by 𝐹𝐴 𝑖 (m 2 ) at time t (min).

A3: the interfacial area occupied by 𝐹𝐴 𝑖 at time t was assumed to be proportional to its molar fraction within the oil droplets.

𝐴 𝐹𝐴 𝑖 (𝑡) = 𝑛 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) ∑ 𝑛 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) i . 𝐴 𝑇𝐴𝐺 (𝑡) (4)
where 𝑛 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) is the number of moles of 𝐹𝐴 𝑖 in the oil droplets (mol) at time t and 𝐴 𝑇𝐴𝐺 (t) is the interfacial area of oil droplets (m 2 ) at time t. This equation can be rewritten as

𝐴 𝐹𝐴 𝑖 (𝑡) = 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) 𝑀 𝐹𝐴 𝑖 ∑ 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) 𝑀 𝐹𝐴 𝑖 i . 𝐴 𝑇𝐴𝐺 (𝑡) (5)
where 𝑀 𝐹𝐴 𝑖 is the molar mass of 𝐹𝐴 𝑖 (g mol -1 ).

A4: oil droplets in the emulsion were considered as spheres. The droplet interfacial area is thus:

𝐴 𝑇𝐴𝐺 (𝑡) = 6. 𝑚 𝑇𝐴𝐺 (𝑡) 𝜌. 𝑑 32 (𝑡) (6)
where 𝑚 𝑇𝐴𝐺 (𝑡) is the total mass (mg) of TAG at time t, 𝜌 is the mean mass density of the TAG (taken as 0.92 mg mm -3 ) and 𝑑 32 (𝑡) is the surface weighted mean droplet diameter measured by laser diffraction (nm).

Combining assumptions A1-A4, Eq. ( 3) can be rewritten:

𝑑𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 𝑑𝑡 = -𝑘 𝐹𝐴 𝑖 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) 𝑀 𝐹𝐴 𝑖 ∑ 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) 𝑀 𝐹𝐴 𝑖 i . 6. 𝑚 𝑇𝐴𝐺 (𝑡) 𝜌. 𝑑 32 (𝑡) (7)
A5: our experimental results show that only a fraction of hydrolyzed products were transferred into the bile salt micellar phase. To calculate the masses of 𝐹𝐴 𝑖 within this phase, a "micellar fraction", 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 , was therefore introduced with the assumption that the transfer of lipolysis products into the micellar phase is instantaneous:

𝑑𝑚 𝐹𝐴 𝑖 𝑚𝑖𝑐 𝑑𝑡 = -𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 𝑑𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 𝑑𝑡 (8)
where 𝑚 𝐹𝐴 𝑖 𝑚𝑖𝑐 is the mass of 𝐹𝐴 𝑖 in the micellar phase (mg).

The micellar fraction 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 and the lipolysis rate 𝑘 𝐹𝐴 𝑖 are the unknown parameters of the model. They were estimated for each 𝐹𝐴 𝑖 by adjusting the model to the experimental data (see section 3.2). To this end, model predictions for TAG masses were calculated as follows.

The total TAG mass, 𝑚 𝑇𝐴𝐺 (𝑡), is the sum of the medium chain TAG (MCT) and long chain TAG (LCT) masses:

𝑚 𝑇𝐴𝐺 (𝑡) = 𝑚 MCT (𝑡) + 𝑚 LCT (𝑡) (9)
where 𝑚 𝑀𝐶𝑇 (𝑡) and 𝑚 𝐿𝐶𝑇 (𝑡) are the MCT and LCT masses (mg) at time t, respectively, calculated according to their fractional mass contributions (α FA i and 1-α FA i ) shown in Table 1 and by adding the adequate mass of glycerol moiety:

𝑚 MCT (𝑡) = � 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) i . (1 -𝛼 𝐹𝐴 𝑖 ) + 𝑚 𝐺𝑙𝑦𝑐𝑒𝑟𝑜𝑙 𝑀𝐶𝑇 (𝑡) (10) 𝑚 LCT (𝑡) = � 𝑚 𝐹𝐴 𝑖 𝑙𝑖𝑝 (𝑡) i . 𝛼 𝐹𝐴 𝑖 + 𝑚 𝐺𝑙𝑦𝑐𝑒𝑟𝑜𝑙 𝐿𝐶𝑇 (𝑡) ( 11
)
where 𝑚 𝐺𝑙𝑦𝑐𝑒𝑟𝑜𝑙 𝐿𝐶𝑇 (𝑡) and 𝑚 𝐺𝑙𝑦𝑐𝑒𝑟𝑜𝑙 𝑀𝐶𝑇 (𝑡) are the masses of glycerol moiety coming from MCT and LCT, respectively. They were calculated via a molar balance equation by considering that there is one mole of glycerol moiety per three moles of 𝐹𝐴 𝑖 .

Parameter estimation

The differential equations of the model (Eq. 7 and Eq. 8) were solved numerically using Matlab ® software (The MathWorks Inc., Natick, MA) equipped with the Statistics Toolbox.

The surface weighted mean diameter 𝑑 32 (𝑡) was integrated into the model using linear interpolations of the experimental values. Model outputs, namely the evolution of the MCT and LCT masses and masses of hydrolyzed products within the micellar phase, were confronted to the corresponding experimental results measured by HPLC and GC, respectively, to estimate the unknown model parameters: 𝑘 𝐹𝐴 𝑖 and 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 . Standard errors, coefficients of variation and 95% confidence intervals of the estimated parameter values were computed using the nonlinear regression software in the Matlab ® Statistics Toolbox and it was checked that estimated values were statistically significant at 0.05 level.

Results and Discussion

Experimental results

Results related to the evolution of droplet sizes during the gastric and intestinal phases were described in detail previously [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF]. It was found that droplet flocculation took place during the gastric phase but these aggregates where re-dispersed after the subsequent addition of bile. Fig. 1 summarizes the results related to the evolution of oil droplet sizes (𝑑 32 ) during the intestinal phase together with the corresponding TAG lipolysis kinetics. Fig. 1A shows that the lipolysis rate was much higher for MCT than for LCT as the hydrolysis of MCT was completely finished after 15 min, whereas about 20% of LCT could still be detected after 300 min of intestinal digestion. In good agreement with many studies [START_REF] Li | New mathematical model for interpreting pH-stat digestion profiles: impact of lipid droplet characteristics on in vitro digestibility[END_REF][START_REF] Ye | Effect of calcium on the kinetics of free fatty acid release during in vitro lipid digestion in model emulsions[END_REF]Zhu, Ye, Verrier, & Singh, 2013), the LCT lipolysis curve also showed two stages with a fast initial reaction rate that markedly slowed down after about 30 min. As previously shown in [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF], this can be attributed to the sudden increase of the surface weighted mean diameter induced by the coalescence of oil droplets, and which led to a sharp reduction of the interfacial area available for lipase adsorption (Fig. 1B).

Fig. 2 presents the bioaccessibility of the 8 fatty acids considered, i.e. the lipolysis product concentrations measured within the micellar phase, expressed as percentages of FA initially present in the oil phase. Three main observations can be made. First, fatty acids can be divided into two groups according to the evolution of their bioaccessibility. The first group contains the FA arising from lipolysis of MCT, C8:0 and C10:0. They were released more quickly than the second group which corresponds to the FA arising from the lipolysis of LCT (Table 1). One may also notice that the long chain FA showed two-stage shapes with a slower rate of appearance within the micellar phase after about 30 min. These results are therefore consistent with the MCT and LCT lipolysis kinetics deduced from the HPLC measurements (Fig. 1A). Second, both the rate of appearance and the concentration measured at the end of the experiment (300 min) depended on the FA considered. The rate and extent of FA appearance in the micellar phase tended to be higher for shorter FA. This confirms that all fatty acids do not behave in the same way in terms of lipolysis kinetics and/or incorporation into the bile salt micelles. Finally, since MCT were fully hydrolyzed in less than 15 min according to the HPLC measurements (Fig. 1A), the plateau value of about 65% for C8:0 and C10:0 (Fig. 2) demonstrates that only a fraction of hydrolyzed products was recovered within the micellar phase. This is in accordance with the study of [START_REF] Sek | Evaluation of the invitro digestion profiles of long and medium chain glycerides and the phase behaviour of their lipolytic products[END_REF] who showed that lipolysis products can also be found in the oil and pellet phases after the centrifugation step used to separate the micellar phase.

Modeling results

The model was designed to simulate the lipolysis kinetics as well as the bioaccessibility of individual FA, and considers 8 fatty acids representing 95.6% of the total FA mass. The parameters to be estimated were the reaction rate constant (𝑘 𝐹𝐴 𝑖 ) and the micellar mass fraction (𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 ) for each FA.

It appeared that the fatty acids coming from MCT, C8:0 and C10:0, were fully hydrolyzed in less than 15 min of intestinal digestion (Fig. 1A and Fig. 2), i.e. before the first sampling time.

The lipolysis rate constants, 𝑘 𝐶8 and 𝑘 𝐶10 , could therefore not be estimated due to lack of intermediate data. After preliminary model simulations, they were set at the minimum value enabling to reproduce a sufficiently fast MCT lipolysis, which was found to be 10 mg m -2 min -1 . According to the above considerations, 6 lipolysis rates 𝑘 𝐹𝐴 𝑖 and 8 micellar fractions 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 remained to be determined. In a first attempt, all parameters were estimated by fitting the model to the experimental data. This model version provided very good fits (data not shown) but it appeared that the 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 values were similar for all FA except C8:0. In other words, the bile salt solubilization ratio of the lipolysis products was about the same for FA longer than C8:0. To reduce the number of unknown parameters, it was therefore decided to keep only two 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 parameters, one for C8:0 and the other one common to all the other FA.

Hence, the final version of the model had 6 unknown lipolysis rates 𝑘 𝐹𝐴 𝑖 and 2 unknown micellar fractions 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 . Fig. 3 shows that this model version still provided very good fits of the experimental data, and Table 2 shows that the parameter values were properly estimated (coefficients of variation smaller than 10%). These results are further discussed according to their biological meanings.

Fractions of hydrolyzed products recovered within the bile salt micellar phase

Estimated values for the micellar fraction parameters (Table 2) can be interpreted as follows.

About 70% (w/w) of the C8:0 lipolysis products were released in the bile salt micellar phase, and about 56% for the other FA. The remaining amounts of products were retained in the oil and pellet phases during the centrifugation step. The order of magnitude of these values is consistent with studies in which centrifugation was also employed to measure the micellar content in lipolysis products [START_REF] Sek | Evaluation of the invitro digestion profiles of long and medium chain glycerides and the phase behaviour of their lipolytic products[END_REF]. Note also that the model assumes an instantaneous transfer of the products into the micellar phase. The very good fits of the bioaccessiblility kinetics (Fig. 3) obtained under this assumption therefore strongly suggest that the solubilization of the lipolysis products in the bile salt micelles was not rate limiting.

Similar micellar fraction values for all the lipolysis products considered may seem in contradiction with previously published data. Several studies have indeed shown that lipolysis products behave differently in terms of bile salt solubility [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF][START_REF] Hofmann | The behavior and solubility of a number of pure monoglycerides in dilute, micellar bile-salt solution[END_REF], with molar saturation ratios that can vary from 0.07 up to 1.86 [START_REF] Freeman | Properties of fatty acids in dispersions of emulsified lipid and bile salt and the significance of these properties in fat absorption in the pig and the sheep[END_REF]. In fact, the experimental protocol used in these studies was quite different from an in vitro digestion protocol since mixtures of pure molecules were used to obtain these results. During the digestion of complex natural oils, a great number of interactions take place between the different lipolysis products and bile acids, and this can greatly modify the solubility of the products in the bile salt mixed micelles. In our study, the products/bile salts molar ratio was of about 1.32 at the end of the experiment, thereby indicating that the saturation ratio for our mixture of lipolysis products was even greater.

One may also wonder why the micellar fraction obtained for C8:0 is different from those obtained for the other FA products. This can probably be attributed to the greater water solubility of this fatty acid. Indeed, using the formula proposed by [START_REF] Tzocheva | Solubility limits and phase diagrams for fatty acids in anionic (SLES) and zwitterionic (CAPB) micellar surfactant solutions[END_REF], it was found that C8:0 was the only free fatty acid with a significant water solubility (0.64 g L -1 ).

More precisely, within our experimental conditions, it can be estimated that about 6 mg of caprylic acid could dissolve if our reaction medium was made of pure water. This corresponds to 31% by weight of the total C8:0 mass introduced within the digestive tubes. According to our modeling results, a common micellar fraction of 56% provides good results for all FA except C8:0, for which a micellar fraction of 70% was estimated. The additional 14% for C8:0 (70 = 56 + 14) are therefore compatible with the contribution of a water soluble fraction, knowing that the micellar phase is not pure water and that the total C8:0 mass accounts for both FFA and sn-2-MAG molecular forms. According to this interpretation, the solubility of the lipolysis products in the bile salt mixed micelles would therefore be approximately the same for all the FA considered, C8:0 included.

FA specific lipolysis rate constants

The lipolysis rate constants estimated with the model are given in Table 2. The same data, but expressed as normalized values relatively to oleic acid, are also presented in Fig. 4 for comparison purposes with the available literature. We may first highlight the very good agreement of our results with those obtained by [START_REF] Berger | Lipases in organic solvents: The fatty acid chain length profile[END_REF] for C12:0, C14:0, C16:0 and C18:1n-9 in spite of very different experimental conditions. Indeed, if our results arise from the modeling of in vitro intestinal digestion of emulsified lipids made of a mixture of different TAG, [START_REF] Berger | Lipases in organic solvents: The fatty acid chain length profile[END_REF] used a protocol based on a lipase-catalyzed reaction in an organic solvent with a mixture of TAG that contained 3 identical acyl residues.

We may also stress that, according to our results, C8:0 and C10:0 presented rate constants that were at least 3 times the one estimated for oleic acid (Table 2), hence confirming the early study of [START_REF] Desnuelle | Specificities of Lipases[END_REF] who found the highest pancreatic lipase activity for C4:0 and reported a strong decrease in activity (by a factor of about 2) between C10:0 and C12:0.

Altogether, our findings are therefore in agreement with the reported tendency of pancreatic lipase to be less active on longer saturated fatty acid residues. These variations are usually ascribed to substrate dependent catalytic lipase activity, although other reaction steps, such as interfacial or diffusive mass transfers of substrates and products, may play a role [START_REF] Verger | Interfacial enzyme kinetics of lipolysis[END_REF][START_REF] Verger | Action of Phospholipase A at Interfaces[END_REF].

Besides, if DHA (C22:6 n-3) was not studied by [START_REF] Berger | Lipases in organic solvents: The fatty acid chain length profile[END_REF], slower rates of pancreatic lipolysis for long-chain polyunsaturated fatty acids have been reported by many authors [START_REF] Akanbi | Pancreatic lipase selectively hydrolyses DPA over EPA and DHA due to location of double bonds in the fatty acid rather than regioselectivity[END_REF][START_REF] Bottino | Resistance of certain long-chain polyunsaturated fatty acids of marine oils to pancreatic lipase hydrolysis[END_REF][START_REF] Ye | Effect of calcium on the kinetics of free fatty acid release during in vitro lipid digestion in model emulsions[END_REF]. For instance, the lipolysis rate constant of DHA should be about 0.17 times that obtained for C18:1 n-9 according to [START_REF] Yang | Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination[END_REF], and of about 0.6 times that obtained for C14:0 according to [START_REF] Mukherjee | Substrate specificities of lipases in view of kinetic resolution of unsaturated fatty acids[END_REF]. These values are represented by a triangle and a circle in Fig. 4, respectively, to show that our estimated rate constant for DHA is consistent with these findings. This higher resistance of long-chain polyunsaturated fatty acids has been attributed to an inhibitory effect induced by the presence of a double bond near the carboxyl group [START_REF] Akanbi | Pancreatic lipase selectively hydrolyses DPA over EPA and DHA due to location of double bonds in the fatty acid rather than regioselectivity[END_REF][START_REF] Bottino | Resistance of certain long-chain polyunsaturated fatty acids of marine oils to pancreatic lipase hydrolysis[END_REF][START_REF] Lawson | Human absorption of fish oil fatty acids as triacylglycerols, free acids, or ethyl esters[END_REF].

Evolution of the TAG composition during lipolysis

Fig. 5 presents the oil-water interfacial areas occupied by each FA residue of TAG molecules as given by the model. They were calculated by assuming proportionality with their molar concentrations within the lipid phase (Eq. 4). Therefore, the evolution of these FA specific interfacial areas is also representative of the predicted changes of the oil droplet composition during the course of digestion. Results show that about 50% of the oil surface area was occupied by MCT fatty acid residues (C8:0 and C10:0) at the beginning of the experiment.

After 15 min, MCT had totally disappeared (Fig. 1A) due to higher lipolysis rate constants for C8:0 and C10:0. This induced a considerable change in the composition of the remaining oil, with only long-chain fatty acid residues remaining. At the end of the experiment, DHA was by far the dominant FA residue (more than 70% by mole) because it had the lowest lipolysis rate constant (Table 2).

As a final remark, we may highlight that our previous model [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF]

Conclusion

In the present study, our previous model [START_REF] Giang | Dynamic modeling highlights the major impact of droplet coalescence on the in vitro digestion kinetics of a whey protein stabilized submicron emulsion[END_REF] was extended to take into account the fatty acid composition of the oil substrate, and to enable the modeling of the in vitro bioaccessibility of the lipolysis products. The model provided very good fits of the experimental data and shows that the differences observed in the bioaccessibility kinetics of the studied FA originated from different FA lipolysis rate constants. It was also used to simulate the compositional evolution of the remaining oil during the course of digestion.

Results related to the FA specific reaction rates were in good agreement with the available literature and confirm a general tendency towards a greater pancreatic lipase activity on shorter fatty acid residues. Our results also support the idea that lipolysis products are rapidly and equally solubilized (i.e. no fractionation) within the bile salt mixed micelles, with a probable additional contribution of water soluble products for fatty acids ≤ C8. The present model provides a significant improvement in comparison with models which assume an average lipolysis rate with a better representation of the intestinal lipid digestion mechanisms. [START_REF] Berger | Lipases in organic solvents: The fatty acid chain length profile[END_REF]. The circle and the triangle for C22:6 n-3 were estimated from the studies of [START_REF] Mukherjee | Substrate specificities of lipases in view of kinetic resolution of unsaturated fatty acids[END_REF] and [START_REF] Yang | Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination[END_REF], respectively.

  assumed an average lipolysis rate for the entire LCT oil fraction, which somewhat overestimated the LCT lipolysis rate at long times. By taking into account FA specific lipolysis rate constants, the newly developed model predicts an accumulation of most resistant FA residues in the remaining oil (Fig 5), which leads to a progressive decrease of the average reaction rate and a better agreement with experimental measurements of the remaining LCT at 300 min (Fig 3).

Fig 1 .

 1 Fig 1. Lipolysis of MCT (squares) and LCT (triangles) during intestinal digestion (A).

Fig 2 .

 2 Fig 2. Bioaccessibility of individual lipolysis products (lines are guides for eyes). Means and

Fig 3 .

 3 Fig 3. Comparison between calculated and experimental evolutions of the MCT, LCT, and

Fig 4 .

 4 Fig 4. Lipolysis rate constants for the different fatty acid residues according to the carbon

Fig 5 .

 5 Fig 5. Evolution of the interfacial areas (normalized values) occupied by each fatty acid

  Figure 1

Table 1 .

 1 Fatty acid composition of the native emulsion which was made of a mixture of MCT and LCT (37.5 and 62.5 % by weight, respectively).

		TAG composition	Origin of each FA
	Fatty acid	Mean of 3 replicates	STD of 3 replicates	Fraction coming from LCT	Fraction coming from MCT
		(% by weight)	(% by weight)	𝛼 𝐹𝐴 𝑖	1-𝛼 𝐹𝐴 𝑖
	C8:0	19.76	0.71	0.02	0.98
	C10:0	16.89	0.33	0.05	0.95
	C12:0	2.98	0.20	0.93	0.07
	C14:0	7.36	0.09	0.97	0.03
	C16:0	6.40	0.13	0.92	0.08
	C16:1 n-7	1.39	0.07	0.99	0.01
	C18:1 n-9	14.41	0.14	0.98	0.02
	C22:6 n-3	26.47	0.10	1.00	0.00
	Other FA	4.31	1.72	0.89	0.11

Table 2 .

 2 Results of the parameter estimation for the lipolysis rate 𝑘 𝐹𝐴 𝑖 and the micellar fraction 𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐 for each fatty acid.

	Fatty acid	𝑘 𝐹𝐴 𝑖 (mg min -1 m -2 )		𝑓 𝐹𝐴 𝑖 𝑚𝑖𝑐
		Value	STD	Value	STD
	C8:0	10*		0.70	0.01
	C10:0	10*
	C12:0	1.93	0.16
	C14:0	1.57	0.11
	C16:0	1.34	0.10	0.56	0.01
	C16:1 n-7	1.50	0.11
	C18:1 n-9	3.11	0.25
	C22:6 n-3	0.99	0.06

(*) minimum value providing a complete hydrolysis of MCT in 15 min.

Zhu, X.,Ye, A., Verrier, T., & Singh, H. (2013). Free fatty acid profiles of emulsified lipids during in vitro digestion with pancreatic lipase. FoodChemistry,[398][399][400][401][402][403][404]

Acknowledgements

This work was supported by the Institut National de la Recherche Agronomique and the Institut Carnot QUALIMENT (France). The authors are involved in the Food and Agriculture COST (European Cooperation in Science and Technology) Action FA1005 'Improving health properties of food by sharing our knowledge on the digestive process (INFOGEST)'.


1 Urban Ambiances as Common Ground?

Jean-Paul Thibaud In Lebenswelt. Aesthetics and philosophy of experience. n°4, 2014 I would like to start with an expression: "sharing ambiances"1 . There are at least two meanings underlying such an expression.

On the one hand, we can question whether it is possible to share ambiances and ask ourselves whether an ambiance can always be shared, whether certain conditions must be met for an ambiance to be shared, whether what is "shareable" takes on a particular expression when it is viewed through the lens of ambiances… These are what we can call "experienced ambiances", ambiances as they are experienced by the senses.

But we can also share conceptions about the idea of ambiances and test the extent to which it is possible to agree on the notion of ambiance, initiate a debate on the meaning and contribution of such a concept, experiment with types of dialogue between different disciplinary fields, etc. We can call these: "reflective ambiances", i.e., ambiances as they are addressed by thought.

To question the shared experience of an ambiance and open up the notion of ambiance for debate is quite simply two sides of the same coin: the two facets of sharing ambiances. This involves the potential crossover between the field of sensory experience and that of reflective thought, the continuity between empirical and theoretical realms. Here, we are already at the heart of the enigma since ambiances play on both sensing and knowing, and involve a certain "I-do-not-know-what" (je-ne-saisquoi as commented by Vladimir Jankélévitch, 1980) that can never be fully resolved. Like Saint Augustine with time, everyone has experiences with ambiances in everyday life and yet we have a lot of trouble explaining exactly what they are. In order to avoid an overly general and abstract argumentation, I will rely on short descriptions, lived experiences and concrete situations throughout the text. This bridge between experienced and reflective ambiances will enable me to approach this very intangible and quasi-indefinable domain.

The aim of this paper is to point out various arguments which question ambiance as a common ground of everyday urban experience. This foray into the problematic modes of existence of ambiances offers proof of both the great diversity in this field of research (Thibaud & Siret, 2012) and of the unique precautions necessary when addressing the topic. Indeed, ambiances demand special attention and require that we nurture them, otherwise they disappear or become subsumed into the broader fields of the environment or the landscape. Timothy [START_REF] Morton | Ecology without Nature: Rethinking Environmental Aesthetics[END_REF] even proposes to develop an ambient poetics, a special attention to the writing of aesthetic experience, "a way of conjuring up a sense of a surrounding atmosphere or world" (p. 22). Without doubt, ambiances are a fragile and unstable research topic and one that can be broached only after a lot of preliminary research. Ambiances bring to life the world around us; they make it truly inhabitable and conjure up extremely sensitive aspects of our human condition. What would a world devoid of ambiance be like? Simply asking the question makes it clear that now more than ever we need to nurture this vital aspect of living space and living together.

1-Bringing the sensory to the forefront

Using the notion of ambiance allows us to test ordinary cultures from a sensory perspective and to reframe social challenges through the lens of aesthetic issues. To do so, it is necessary to expand the usual frameworks of perception. Urban life is multifaceted and relies on a diversity of sensory universes and ways of experiencing them (Diaconu & al., 2001). Here, we are clearly close to an ethnographic approach which involves distancing ourselves from what seems obviousnon questionablein order to allow alterity to emerge. This is similar to using unfamiliaritybreaching experimentas a method and as a means to shift focus, like field anthropologists trying to give meaning to what they observe. Any investigation into urban culture as such, involves an attempt to dissolve stereotypes, an effort to shake up established categories, a certain "derangement of the senses" (Rimbaud). As Bernard Waldenfelds (2009) highlighted, phenomenology maintains close ties with ethnology because both test and deal with the pathic dimension of experience. I will return to the heuristic value of unfamiliarity and strangeness later. For now, let me just emphasize the importance of developing an approach that enables us to question the taken for granted and vary the way we deal with the world. In other words, dealing with urban cultures requires an ability to spur interaction between and amongst different ways of being in the world.

A second important element that brings us even closer to ambiance involves learning the lessons of an approach that is tuned into the forms and expressions of social life (Augoyard, 1979). We know that using the senses in everyday life invariably means perceiving things, identifying objects, interpreting scenes and engaging with the world. And yet alongside such perceivingsuch practical and cognitive perceptive activitythere is another, equally important facet to the sensory world that is: sensing. The sensing underlying an aesthetic experience is more about our ways of feeling the world; it is related to affective tonalities and ways of being together in a situation. In such "ambiental aesthetics" we are in the pre-reflective dimension of an experience, one which involves above all a bodily experience and an immediate sensation. Erwin [START_REF] Straus | Du Sens des Sens[END_REF] identified this nicely when he distinguished between the "pathic" (affective) and "gnostic" (cognitive) aspects of our relationship with the world. In our everyday lives, however, perceiving tends to, at least partially, obscure or mask sensing.

The challenge, therefore, is to bring the latter back to the forefront. In this respect, the notion of modal anthropology developed by François [START_REF] Laplantine | Le social et le sensible[END_REF] is a good way to incorporate ways of sensing into research on urban life by advancing the idea of a rhythmic epistemology. As he has noticed with regard to Japanese culture, the goal is not only to describe the sensory world based on the discontinuity of signs (which need to be interpreted as best they can), but also on the continuity of rhythms (which instead need to be modulated and varied). Such is the project of rhythmanalysis developed by Henri Lefebvre in his last book (2004). In other words, there is a shift here from semiotics to fundamental aesthetics, that is capable of returning to in-situ sensitivity. This is of central importance since it is one of the conditions for work on ambiances: the notion of ambiance loses all meaning if this sensing dimension is not acknowledged. As Gabor [START_REF] Csepregi | The clever body[END_REF] declared: « To detect a particular atmospheric quality means to reach beyond the factual, the objectively given: to hear, beyond the sound, the timbre of a voice, and to see, beyond the shape, the glimmer of a colour » (p. 43). Forays into the study of other cultural spheres, unsettling or disruptive experiences, focusing on unusual or extreme states of existence, and the careful study of breaches of everyday life, are all destabilising situations that bring us closer to the sensory, affective and atmospheric immanence of urban experience.

As we can see, we are currently moving towards a new research approachshared by sensory anthropology and the aesthetics of ambianceswhich allows us to understand culture as a lever for sensitivity, and urban cultures as diverse expressions of the sensory world in the making. Virginie Milliot (2013) gave a very good example of this argument with her thorough ethnographic study of the Goutte d'Or in Paris: « Anyone who goes to Goutte d'Or for the first time has the distinct impression that they are entering a very singular type of territory. It is not somewhere that you just stroll through, flitting nonchalantly over the surface. It is a neighbourhood that captivates the passer-by, plunging them into a sensorial world that resonates with emotions and reactions. Both the inhabitants and those who know it well refer to a highly-specific "ambiance"that it is difficult to put your finger on -to account for the conflicting feelings of attraction or repulsion and familiarity or strangeness that the neighbourhood evokes. » This line of thought involves the socialisation of sensory experience, the continuous learning and day-to-day creative processes at work within the human sensorium. Thus, it is not strictly limited to the artistic sphere since it shapes and renews itself more broadly alongside and according to everyday environments. If we wanted to push this further, we could say that contemporary urban space becomes the ideal place for shaping new modes of sensing. This argument has already been developed concerning the modern city. Georg [START_REF] Simmel | The Metropolis and Mental Life. The Sociology of Georg Simmel[END_REF] focused on the over-stimulating character of the metropolis, Walter Benjamin ( 2002) emphasized the experience of shock and loss of aura and Siegfried [START_REF] Kracauer | The Mass Ornament: Weimar Essays[END_REF] noted the various phenomena sustaining a culture of distraction.

But such thinking requires us to move beyond the practical and functional aspects of everyday life, and not focusing solely on the cognitive and interpretive angles of the perceptive experience; rather, they demand that we allow the human sensorium to flourish and be fully recognized as such. Under such conditions, sensory cultures emerge where feeling and acting come together, at the meeting point of praxis and aesthesis.

2-Putting common experience into perspective

Ambiances allow us to shed new light on the issue of being together by initiating a dual dynamic of socialising the sensory and sensitising social life. What happens to the distribution of the sensible when it is framed in terms of ambiance? How do different sensitivities deal with an ambiance? How do ambiances change the way we think about sensory forms in social life? It is worth pointing up two fundamental aspects of ambiances here which will help us address this issue.

First, as Gernot [START_REF] Böhme | Atmosphere as the Fundamental Concept of a New Aesthetics[END_REF] has clearly established, atmospheres are not only internal, they are also external, they involve a sensory medium, a third term between me and the world. Far from being a strictly personal experience or limited to a purely subjective mood, ambiances never cease to make affective tonalities public. They display affective tonalities as a common sensory domain to be seen, heard, smelled, touched and sensed. In other words, they make them perceptible to the eyes, ears and skin of everyone. Like the choir in the theatre of Ancient Greece, ambiances make the tonality of current situations perceptible and accompany our shared life.

Second, ambiances are the site par excellence in which sensing occurs and as such are inherently bound up with an empathetic relationship with the world. In other words, ambiances assume a means of synchronising and immediate integration into the surrounding environment. The public nature of an ambiance's tonalities and its empathetic power are two essential aspects at the root of the issue of common experience.

Numerous attempts have been made to clarify the unifying movement at work in an ambiance. Here, we are dealing with phenomena involving resonance, empathy, contagion, imitation, syntony, and sometimes even unison and fusion. Marc Breviglieri (2013) perfectly described the "spark of feeling" (étincelle du sentir) involved in assistance to the homeless persons in borderline states such as profound drunkenness, the sleeping mode or being chilled to the bone: « the borderline situation requests at first to find a complicity with the floating worlds of perception of the homeless person and to generate simultaneously threads of mutual echo. But then, the question is how the atmosphere spreads and reaches the context, undoing the insensible and bringing the sensitive to the foreground » (personal translation). From this perspective, ambiances are part of an atmospheric in-between, an envelopinga means of connectingthat highlights our ability to gel with others, to partake in a social situation. This is due to its all-encompassing nature and our ability to be submersed in a shared ambient milieu, regardless of how fleeting or established it may be. The famous Russian dramaturgist Anton [START_REF] Chekhov | On the technique of acting[END_REF] formulated it is very nicely: « Let us take another example: a catastroph on a crowded street. How many different personal moods are there? One person is afraid, another full of compassion, the third burns with a desire to help, a fourth is indifferent, but the objective Atmosphere of the horror of the catastroph prevails over all the people concerned, regardless of their personal moods. » (p. 31) While this vague background is part of any shared experience of an ambiance and in some respects constitutes its experiential basis, it must not mask the fact that we live in a diverse world and that our relationships with an ambiance are many and varied. We can therefore be all plunged together into a single ambiance (an atmospheric inbetween) whilst experiencing different ways of feeling submerged. To say that "we are not in the ambiance" (ne pas être dans l'ambiance) means that one's lived experienced is not necessarily in perfect harmony with the dominant tonality of a situation. Beyond the empathetic relationship underlying any experienced ambiance -I cannot not be attuned to the here and now of a sensory situation in which I find myselfis another level of experience that is related to my sense of presence in the situation, which can be more or less pronounced, more or less tinged with familiarity or strangeness. It is as such not simply a matter of being present together in a single situation, but of more or less feeling this shared presence. Rainer [START_REF] Kazig | Vers une culturalisation de la recherche sur les ambiances[END_REF] coined the term "cultural resonance ambiances" to designate a specific form of shared ambiance when people belonging to the same cultural background meet in particular public places such as bars, galleries, concerts.

A third aspect of common experience involves the form of the experience itself. Indeed, although we tend to assimilate ambiances with convivial collective experiences saying that "there is ambiance" (il y a de l'ambiance) usually means that a situation is lively, even friendlythey may just as easily be tense, heated, awful or conflictual. The full range of forms of sociability can be described in terms of ambiance. Here, too, we are still in the realm of common sensory experience, it is just that such experience may be problematic and unresolved, loaded with instability and precariousness over the make-up and cohesion of the group itself. The goal is not to conjure up a subjective feeling but rather the movement and flow of a social situation described by an affective tonality that is more or less positive and cohesive, euphoric or dysphoric, consonant or dissonant, and which is quite likely to evolve over time in one way or another (towards a resolution or deterioration).

In a nutshell, ambiances lead us to think about common experience within a complex web comprised of an "existential" dimension (empathy with the ambient world); a "contextual" dimension (degree of presence in the situation) ; and an "interactional" dimension (forms of sociability expressed in the tonality).

3-Initiating a political ecology of ambiances

Ambiances are part of current changes to lived-in spaces and are involved in the processes transforming urban territories. While ambiances provide an original way of thinking about inhabitant experiences, they also involve extremely important design and planning issues. What are the socio-political consequences of ambiance-based urban design? How can we test and implement the critical strength of the notion of ambiance? How do ambiances change the way we design everyday environments? When we ask such questions, it is not only the ambiance as a given or pre-existing milieu that is under the microscope, but also the creation of ambiances, the ambiantação according to the delightful term used in Brazil. Such interest in the processes, conditions and means of creating ambiances in urban space is a precursor for a possible political ecology of ambiances.

Let me first note that if sensory issues are gaining in importance in practices and representations of the contemporary worldranging from the private sphere to the public domainit is necessary to have a basic shared understanding of the term, otherwise we run the risk of depriving it of the elements underlying its heuristic force and critical strength. While the sensory world has an inherent affective side, it must not be confused with the realm of sentimentality, an avatar of contemporary compassion mongering, with political sentimentality or other spheres of collective resignation. Moreover, the sensory experience bound up with ambiances is not the same as that encountered in the gratuitous luxury, facile comfort or contemporary cosmetics far too often found in advertising discourses and other commercial contexts (Augoyard, 2011).

Similarly, the current rise of augmented reality and embedded technologies can by no means claim to cover the field of ambiances and replace fundamental reflections into contemporary living spaces. Obviously, the notion of ambiance that I wish to promote cannot and must not be written off as a simple product of postmodern sentimentality, something of purely commercial value bound up with a strictly hi-tech environment. That said, a political ecology of ambiances cannot ignore the increasing development of means of instrumentation and instrumentalisation of the sensory world. This involves updating how the everyday sensory environment is framed, programmed and governed (Amin & Thrift, 2002). We may therefore ask how an ambiance-based approach positions itself, amidst the tension between planning strategies and inhabitant tactics, between the spheres of power and resistance movements. In brief, to what extent can an ambiance-based approach avoid addressing the ethical and political issues inherent to all urban and planning work?

This could be the starting point for a whole new field of research focused on deconstructing the deep meaning behind the aestheticisation of public places currently underway [START_REF] Hajer | r eine neue sthetik unserer t dte[END_REF]. It would be necessary to point up the logic underpinning territorial and sensory branding, research the public and cultural policies underpinning sensory planning, and test hypotheses about the pacification [START_REF] Sennett | Espaces Pacifiants[END_REF], sanitization [START_REF] Thomas | écrire l'arrière-plan corporel de l'expérience urbaine[END_REF] and normalisation (Breviglieri, 2013) of shared sensory spaces. The goal would be to develop critical analysis of the current types of sensory implementation by combining a molar and molecular approach that aims to understand the socio-aesthetic issues embedded in the most ordinary urban contexts, and which would clarify the implicit distribution of the sensible that they mobilise or renew.

And yet, we must not forget or underestimate the regular, basic ongoing creation of urban ambiances by city dwellers themselves. Indeed, one of the lessons that ambiances teach us is that lived-in space is not a mere reflection of how designed spaces are received. Inhabitant activity itselfas witnessed in sensory perception, practical action, cultural expression and social interactionis part of ambiance creation in urban spaces. While going about their ordinary lives, city dwellers configure the ambient environment, demonstrate multiple competences and activate diverse know-how [START_REF] De Certeau | The Practice of Everyday Life, trans. Steven Rendall[END_REF][START_REF] Joseph | L'espace du public Les compétences du citadin[END_REF], and are at the centre of micro-resistance to space-time configurations seen as too restrictive or inhospitable [START_REF] Britto | Corpographies urbaines. Pour un urbanisme plus incorporé[END_REF].

There is already an abundance of scientific research that addresses this logic of opportunity, of kairos, of making-do. One less explored area, however, is the realm of ambiances itself. What exactly does it mean to acclimatise and become acclimatised to a situation? How do forms of social life set the tone of a territory? What are the vectors driving an intensification of experience? How much room should be left to improvisation in the creation of a sensory environment? Here too, lies a wealth of questions that could help us assess the breadth of everyday practices in the overall economy of urban ambiances.

Everything points to the fact that the notion of ambiance is in the process of becoming a key to the way we think about and understand transformations in the contemporary urban environment. Like Janus, ambiance appears to have two faces: it is the critical analysts and decoders of the most cutting-edge trends in sensory branding of all kinds; and it is a particularly efficient tool for conditioning and presenting diverse commercial and mercantile environments. This dichotomy is not of course as black and white as it appears, as illustrated by the place and role of artistic action, which can be both a sphere for the legitimisation of marketing logic and a space for destabilising the perceptively obvious. In other words, far from being neutral, the notion of ambiance appears to be bound up with the socio-aesthetic strategies underpinning changes to the sensory world of the future.

4-Questioning what in situ is about

As we saw previously, ambiances are intrinsically bound up with a relationship of immersion within the surrounding world. Describing an experience in terms of ambiance pushes us to define what it means to be submerged in a milieu and requires that we examine in situ issues under a new light. How can we think about situated experiences in terms of ambiance? What is the relationship between an ambiance and a situation? What role does the sensory play, given the contextual nature of shared experience?

A lot of research in the field of micro-sociology has emphasised the importance of situation as a register for analysis and as a blueprint for experience [START_REF] Goffman | Behavior in Public Places: Notes on the Social Organization of Gatherings[END_REF][START_REF] Joseph | L'espace du public Les compétences du citadin[END_REF]. An ambiance-centric approach is the most likely part of this general trend towards reassessing the micro-logic and contextual aspects of social life. Here too, we can schematically identify three research pointers which address in situ issues: the embodiment of situated experiences, the porous nature of sensory spaces, and the efficiency of equipment and the build environment.

-With regard to the embodiment of situated experience, we need to understand all of the consequences related to the fact that the body is the first site from which ambiances are experienced [START_REF] Schmitz | Emotions outside the boxthe new phenomenology of feeling and corporeality[END_REF]. The immediacy of sensing is experienced first and foremost in a physical experience and is part of what phenomenology describes as a pre-reflective experience. The immersion phenomenon is as such part of a dual movement of incorporation and expression, of physical feeling and sensory-motor phenomenon. We could perhaps talk about "ambient gestures" [START_REF] Thibaud | La fabrique de la rue en marche Essai sur l'altération des ambiances urbaines[END_REF], meaning the basic sensory-motor patterns that make up an ambiance: an ambiance can be heavy or relaxing, stimulating or reassuring, energising or hypnotic… But while such sensing by the body provides an initial key for understanding immersion phenomena (in terms of sensory-motor solicitation), we also need to examine its social and imagined repercussions. This is where the intercorporeal comes into play, the "place ballet" [START_REF] Seamon | Body-subject, time-space routines[END_REF], the tacit interactions that occur for example between passers-by in urban public spaces.

Is it relevant then to talk about a "common body"? How can we collectively be as one with an ambiance? In other words, not only the body itself, but also intra-body, inter-body, and body-to-body experiences. Further, the sensory-motor level of experience is not limited to the immediate here and now; it also calls on other phenomena tied to involuntary memory which link up past experiences and transport the subject to previously experienced space-time configurations. A smell will take me back in time; a sound will remind me of another situation… This involves an imaginary opening of the ambient world which uses processes of sensory association, metaphorisation and sedimentation. In other words, immersion here involves the choreography of close bodies and the interconnection of experienced situations.

-With regard to the porous nature of sensory spaces, it is more a matter of the ways situations are filtered and how the public pays attention. For one, all ambient situations are like envelopes, "atmospheric skins" (Hasse, 2000;[START_REF] Griffero | The atmospheric "skin" of the city[END_REF] or "interactional membranes" [START_REF] Bordreuil | Ambiances, partage d'ambiances et prisme interactionnel[END_REF] framing our relationships with the immediate environment. People involved together in a single situationfor example talking or walking together -share a special interaction which places them in an "interactional bath" or an atmospheric flow "that seizes them by encompassing them, directing the two of them together" (Tellenbach, 1992). Such experiences are not, however, impermeable to happenings and events in the surrounding environment (being questioned, an incident, etc.). The challenge is thus to describe the interaction and copenetration that occurs between a social situation and an urban context, how situational commitments are shaped and modelled by what goes on, and the adjustments that take place between the tonality of a social interaction and that of an ambient milieu. And yet the shape and scope of types of attention are also of vital importance since they lead to subtle changes based on the possibility of playing on peripheral attention and attentional thresholds (distracted listening, unfocused vision, divided attention, fringes, etc.). As Juhani [START_REF] Palasmaa | Selfhood and the World. Lived Space, Vision and Hapticity[END_REF] very accurately remarked: « Peripherical vision integrates us with space and its events, while focused vision pushes us out of the space and makes us mere observers » (p. 60).

-Finally, the efficiency of equipment and material apparatus highlights the active role of built environments in the production of immersive phenomena. This concerns how an ambiance is enveloped, and the sensory effects it produces. Depending on the morphological and material features of the built environment, the space is more or less open or closed, dilated or contracted, airtight or porous. A person's sense of being surrounded is more or less pronounced depending on the depth of their gaze, the extent of their movement or the range of their hearing.

Anne [START_REF] Jarrigeon | L'ambiance des foules anonymes[END_REF] well illustrates this overlap between the built forms, the social conducts in public and the atmosphere of the place: « Les Halles [in Paris] aspire, they absorb the flows of encounter, the inhabitants commuting, the determined paths they catalyze before expelling them to the surface. It is a transit point for many: in Les Halles, we run, we go, we find ourselves and then escape, even without reaching daylight. (...) The general deprivation of perspective and the lack of articulation between spaces contribute to the production of a particularly tiring social life, subject to a requirement of fluidity. (...) Les Halles is indeed living in melee, in a general atmosphere made of voltage, acceleration, rapid intersections governed by a true performance of avoidance. » (p. 264-265, personal translation) More generally, however, we may wonder whether we are witnessing the emergence of new fields of design or new means and conditions for the production of the city. The intentionality of designers is not limited to the production of ground-breaking built forms and new sustainable environments; it also touches upon the intentional creation of original sensory environments. Urban changes appear no longer to be the sole product of physical planning; they are also based on the "atmospherisation" [START_REF] Tellenbach | Goût et Atmosphère[END_REF] of living spaces and appear to benefit fully from the potentiality of the intangible world. In other worlds, more and more we tend to design also sensory atmospheres and affective tonalities.

Lastly, we should note that an ambiance-based approach affects the way we think about the role and place of inhabitants in the make-up of territories. This involves the question of living space present in any transformation of space. As a form of sensory life, ambiances place inhabitants at the very centre of urban debates. They also help point up how city dwellers contribute to shaping their living space on a daily basis through their most ordinary, everyday actions. In sum, ambiances contribute in two ways to thinking about embodiment and empowerment by making the sensory a showcase for different ways of living and helping inhabitants become ordinary experts of their living spaces.

So what can we say about the enigma of sharing ambiances? There is a constant search for new connections and other focuses (or better: defocalisation). There is a need to experiment, vary points of view, make art and science resonate, and to experiment with totally new types of writing. Do ambiances not sensitize of urban thought as well as social lifeforms? , Seuil, 1979 W. Benjamin, The Arcades Project, ed. Rolf Tiedemann, trans. Howard Eiland andKevin McLaughlin, New York, Belknap Press, 2002

BibliographyA.

  Amin & N. Thrift Cities. Reimagining the urban, Cambridge, Polity Press, 2002 J.-F. Augoyard Faire une ambiance ? in J.-F. Augoyard (ed.), « Faire une Ambiance », Bernin, À la Croisée, 2011, pp. 17-35 J.-F. Augoyard, Pas à Pas. Essai sur le cheminement en milieu urbain, Paris

This paper has been presented at the International Conference Atmosfere. Tra etica et estetica / Atmosphere. Between Ethics and Aesthetics. Università degli Studi di Torino, Galleria nazionale d'arte moderna Roma, Museo Hendrik Christian Andersen, Sensibilia. Roma, September

27th-28th 2013. It relies on a previous draft published in French(Thibaud, 2013).


MD Clémence Marois
email: clemence.marois@aphp.fr

MD Maria Del Mar Amador
email: mariadelmar.amador@aphp.fr

Christine Payan
email: christine.payan@aphp.fr

MD Lucette Lacomblez
email: lucette.lacomblez@aphp.fr

MD Anne-Marie Bonnet
email: anne-marie.bonnet@aphp.fr

MD Bertrand Degos
email: bertrand.degos@aphp.fr

Jean-Christophe Corvol
email: jean-christophe.corvol@aphp.fr

MD Marie Vidailhet
email: marie.vidailhet@aphp.fr

MD Nadine Le Forestier
email: nadine.leforestier@aphp.fr

MD Valérie Mesnage
email: valerie.mesnage@aphp.fr

MD Jean- Christophe Corvol

MD David Grabli
email: david.grabli@aphp.fr

: 1a

Jean-Christophe Corvol

Manuscript Marois

Outcome of gastrostomy in parkinsonism: A retrospective study

The main outcome measure was survival duration following gastrostomy insertion according to Kaplan-Meier estimate. Cox analysis was also performed to identify factors associated with survival. Finally, we described short term and long term adverse effects occurring during the follow-up period.

Results:

We identified 33 patients with Parkinsonism that received GTI during the study period. One patient was excluded from the analysis because of missing data. Among 32 patients, 7 (22%) had Parkinson's disease and 25 (78%) had atypical parkinsonism. The median survival following the procedure was 186 days (CI 95% [62-309]). In Cox model analysis, total dependency was the only factor negatively associated with survival (HR 0.1; 95 % CI [0.02-0.4], p = 0.001). Pneumonia was the most frequent adverse event.

Conclusion:

In this sample of patients with parkinsonian syndromes, survival after GTI was short particularly in totally dependent subjects. Aspiration pneumonia was not prevented by GTI. A larger prospective study is warranted to assess the potential benefits of gastrostomy, in order to identify the most appropriate indications and timing for the procedure.

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Introduction:

Dysphagia is a common feature in patients with parkinsonian syndromes (PS) [START_REF] Kalf | Prevalence of oropharyngeal dysphagia in Parkinson's disease: A meta-analysis[END_REF] which include idiopathic Parkinson's disease as well as atypical neurodegenerative Parkinson syndromes. Dysphagia results in significant morbidity by interfering with medication administration, nutrition and fluid intake, and is associated with aspiration pneumonia -the most frequent cause of death in patients with PS [START_REF] Akbar | Incidence and mortality trends of aspiration pneumonia in Parkinson's disease in the United States, 1979-2010[END_REF] . For PS patients presenting with dysphagia, the decision regarding gastrostomy tube insertion (GTI) for feeding is challenging since to date, few studies have addressed the procedural risks and outcomes in this patient population [START_REF] Yamazaki | Nutritional support by 'conventional' percutaneous endoscopic gastrostomy feeding may not result in weight gain in Parkinson's disease[END_REF][START_REF] Sarkar | Percutaneous Endoscopic Gastrostomy Tube Insertion in Neurodegenerative Disease: A Retrospective Study and Literature Review[END_REF] . In contrast, there are already established recommendations for GTI to maintain nutritional support in patients with amyotrophic lateral sclerosis (ALS) that present with severe dysphagia. [5-6-7] . Similarly, the guidelines are established for patients with advanced dementia which do not demonstrate a survival benefit, and consequently the procedure is not recommended. [START_REF] Volkert | ESPEN Guidelines on Enteral Nutrition: Geriatrics[END_REF] While the utility of GTI in PS patients remains unclear, our study aims to report the indications and outcomes for PS patients that received GTI at two French movement disorders centers, and to highlight any factors affecting these outcomes.

Methods:

All patients with a diagnosis of degenerative PS including idiopathic Parkinson's disease (IPD), multiple system atrophy (MSA), progressive supranuclear palsy (PSP), Lewy body dementia (LBD) or cortico-basal degeneration (CBD) that received GTI between January 2008 and December 2014 were identified from institutional databases at two movement disorders tertiary-care centers in Paris (see supplementary methods). Data was obtained retrospectively from anonymized routine medical charts and approval from our Institutional Review Board was waived. We also retrieved the number of PS patients seen at least once per year (caseload) in these centers. Informed consent for gastrostomy tube insertion was obtained from patients, or their relatives where the patient lacked capacity to provide informed consent at the time of gastrostomy insertion.

For each patient, data were extracted from medical records by two neurologists (CM and DG). Nutritional status was recorded according to the current French guidelines (see supplementary methods). Dependence was stratified in three levels based on the Schwab and England activities of daily Living Scale (level 1: completely independent (80-100%), level 2: partially dependent, able to walk (40-70%), level 3: totally dependent (0-30%)). Speech was assessed according to item 18 of the Unified Parkinson Disease Rating Scale (UPDRS).

Cognitive status was documented according to DSM IV criteria. Where possible, the indication for GTI from the clinician's perspective was determined from patient charts and was classified according to one or more of the following categories: undernutrition, impaired swallowing, aspiration pneumonia, inability to take drugs orally or unknown. Short-term adverse-events (AE) were defined as complications that occurred during the gastrostomy procedure or the related hospital stay. AE occurring after hospital discharge were classified as long-term AE. For the survival study, data were censored on the 31th December 2015. Documented follow-up was available for all included patients for at least one year post-GTI.

The main outcome measure was median survival time post-GTI, estimated by the Kaplan-Meier method. Cox proportional hazards regression analysis was performed to explore predictors of survival post-GTI. Covariates in the Cox regression were chosen among factors that might affect survival post-GTI in patients with a PS according to previous studies and our clinical judgment [START_REF] Oosterveld | Prognostic factors for early mortality in Parkinson's disease[END_REF] . These covariates were age at gastrostomy, disease duration, atypical parkinsonism, presence of dementia, under-nutrition and inability to walk. Hazard

Results:

From January 2008 to December 2014, between 4000 and 4500 PS patients were seen in clinic each year. Of these, approximately ten percent suffered from atypical parkinsonism.

Thirty-three patients were identified as having received gastrostomy tube insertion (mean 4.7 ± 1.3 procedures each year). One patient was excluded from the study because of missing data. The general features of these patients are presented in table 1. The indication for GTI was explicit for 32 patients and is reported in table 1. Although undernutrition was noted in 72 % of the patients, this feature was the specific indication from clinician's perspective in only 50 %. Other indications included swallowing impairment (n = 15, 47 %), aspiration pneumonia (n = 11, 34 %) or inability to take drugs orally (n = 7, 22 %). The median survival following the procedure was 186 days (CI 95% [62-309]) according to Kaplan-Meier estimate (figure 1A). Cox regression analysis (figure 1B and supplementary table S2) showed that being totally dependent was the only independent factor associated with decreased survival (HR 0.1; 95 % Confidence Interval [0.02-0.4], p = 0.001).

Two patients (6.25%) died during their hospital stay due to aspiration pneumonia. Survival rate was 40.6% at one year and 34.4% at two years. During the follow up period, 22 patients (69%) presented with at least one AE. Fourteen patients (46 %) developed aspiration

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pneumonia associated with acute pulmonary distress (9 patients), mechanical ventilation (3 patients) or tracheostomy (2 patients). Gastro-intestinal AEs, mainly gastrostomy related, accounted for 28% of the long term AEs (supplementary table S1). Median survival for MSA patients (n = 15) after disease onset was 10.9 [9-12.8] years as estimated by Kaplan-Meier method.

Discussion:

Despite the prevalence of swallowing disorders in patients with PS, very few studies have addressed the utility of GTI in this population. To our best knowledge, this is the first study that specifically describes the current use of gastrostomy in patients with PS, through the experience of two tertiary referral movement disorders centers.

Our study revealed that GTI was mainly utilized in patients with atypical parkinsonism. Of the seven patients with IPD, five presented with very advanced stages of disease and in the two other cases GTI was proposed specifically due to swallowing impairment albeit earlier in the disease course. Of course, the ability to facilitate therapeutic optimization of L-doparesponsive symptoms in IPD patients may have been an additional factor in the clinician's decision to perform GTI, even if not made explicit in the patient charts.

With respect to the primary outcome measure, our cohort demonstrated a strikingly short median survival of 6 months (186 days) following the procedure. This contrasts with ALS patients in two retrospective studies that demonstrated a median survival after GTI of 228 and 327 days respectively [START_REF] Russ | Percutaneous endoscopic gastrostomy in amyotrophic lateral sclerosis[END_REF][START_REF] Pena | What is the relevance of percutaneous endoscopic gastrostomy on the survival of patients with amyotrophic lateral sclerosis?[END_REF] . The short survival measured in our cohort may be partly explained by the fact that GTI was performed in patients at late stages of their disease course. At the time of GTI, 75% percent of all patients were completely dependent. Furthermore, in the subgroup of MSA patients (n=15,) gastrostomy was performed after median disease duration of 8.5 years which corresponds closely with overall median survival

M A N U S C R I P T A C C E P T E D ACCEPTED MANUSCRIPT Marois 9
in MSA [START_REF] Low | Natural history of multiple system atrophy in the USA: a prospective cohort study[END_REF][START_REF] Figueroa | Multiple system atrophy: prognostic indicators of survival[END_REF] . Naturally, no conclusions can be drawn as to an effect of GTI on survival owing to the absence of a control group. Identification of retrospective control patients in studies such as this (patients that declined gastrostomy) is difficult, as has been documented in other studies [START_REF] Group | Gastrostomy in patients with amyotrophic lateral sclerosis (ProGas): a prospective cohort study[END_REF] . Furthermore, any attempt at prospective randomization in this particular context raises ethical concerns.

Total dependency before gastrostomy was found to be the only factor independently associated with survival. With small numbers of participants, our study was unlikely to be sufficiently powered to demonstrate associations between survival and other factors measured such as age, disease duration and the presence of dementia.

The most frequent complication recorded was aspiration pneumonia which occurred in half of the patients and lends weight to the prevailing view that GTI does not prevent aspiration.

Interestingly, preventing aspiration pneumonia was the main indication from clinician's perspective for 34 % of our patients.

Given that dysphagia in PS is common, the small number of GTI procedures undertaken in our study centers was surprisingly low in comparison with our large case load [START_REF] Kalf | Prevalence of oropharyngeal dysphagia in Parkinson's disease: A meta-analysis[END_REF] . This may be either the result of clinicians' reluctance to propose GTI to this population given the uncertainties regarding the risks and benefits, patients or relatives' refusal of these invasive techniques or a combination of both. As emphasized previously, no guidelines endorse these procedures in PS patients. Furthermore, advance care planning is rarely discussed in relative slowly progressive diseases such as PS [START_REF] Kluger | Palliative care and Parkinson's disease: Meeting summary and recommendations for clinical research[END_REF] .

Our study presents several limits, mainly due to its retrospective nature. Incomplete information was available about patients' nutritional status after gastrostomy, so we could not draw any conclusions regarding the nutritional efficacy of GTI in PS. Another shortcoming is the lack of quality-of-life data for patients and caregivers following

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gastrostomy which, in a setting where survival may not be the primary goal, is crucial to guide future clinical decisions. As randomization is not conceivable in this context, these complex issues should be at best addressed in future research with mixed methods (both quantitative and qualitative). Finally, this study, performed at two French movement disorders centers, may reflect a regionally-specific practice and thus, not be generalized to the broader PS population.

In conclusion, this study is one of the first to give specific insight into outcomes of GTI in PS.

According to our results, ambulant patients are more likely to benefit from the procedure. A larger prospective study is warranted to assess the potential benefits of gastrostomy in terms of quality of life and nutritional efficacy, as well as to identify the most appropriate indications and timing for the procedure. This future research will enable clinicians and patients to discuss choices in an advance care planning setting with reference to an adequate evidence-base.

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  95 % confidence intervals were computed for each variable. Secondary outcome measures included the median survival time from disease onset. A Kaplan-Meir estimate was computed only for the MSA group (n = 15) and could not be calculated for other disease subgroups due to small sample sizes (IPD n = 7, PSP n = 5, CBD n = 3 and DLB n = 2). A p-value less than 0.05 was considered statically significant for all analyses. Statistical analyses were performed using SPSS 24 (IBM).

Table 1 :Figure 1 :

 11 Figure 1: Survival functions for patients that underwent gastrostomy

Acknowledgments: We would like to thank Dr Richard Blazé for correcting the English and Dr Céline Louapre for statistical advice and helpful criticism.

-The median survival following the gastrostomy tube insertion was 6 months.

-Being totally dependent was associated with a shorter survival.

-Aspiration pneumonia is no prevented by gastrostomy tube insertion.