Donadille

Keywords:

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TITLE OF CASE Do not include "a case report"

Gangliocytoma: outcome of a rare silent pituitary tumour

SUMMARY Up to 150 words summarising the case presentation and outcome (this will be freely available online)

The most common finding in front of a pituitary incidentaloma is a silent pituitary adenoma. We describe a 59 years old woman with a pituitary gangliocytoma and her follow up after one year. Hormonal exploration only evidenced partial corticotropic insufficiency. A transphenoidal surgery was performed due to the tumour's suprasellar expansion. Gangliocytoma is a benign tumour of unknown prevalence, belonging to CNS tumour with neuronal differentiation, and 129 cases have been reported in the literature. GH, ACTH or prolactin secretions have been reported, as these ganglion cell-like mature neurons are usually mixed with secreting pituitary endocrine cells. We report a case with a pure gangliocytoma devoid of symptomatic endocrine secretion, not surrounded by pituitary endocrine tumour cells. Immunochemistry of the tumour was positive for both hypothalamic GHRH and pituitary hormones, such as GH and ACTH. Hence, this immunoexpression was not associated with peripheral hormonal secretions, suggesting biologically inactive hypothalamo-pituitary hormones.

BACKGROUND Why you think this case is important -why did you write it up?

Gangliocytoma is a benign pituitary tumour, which contains mature neurons resembling hypothalamic ganglion-like cells intermixed with a fibrillary background without glial tumoral component. We report an unusual case of a patient presenting with a pituitary gangliocytoma revealed as an incidentaloma, by a magnetic resonance imaging (MRI). Histopathology could not identify any endocrine anterior pituitary tumour cells. Interestingly, immunohistochemistry revealed that ganglion-like neurons themselves were positive for ACTH as well as GH and GHRH. This inactive hormonal secretion may illustrate altered processing or bioactivity of those hormones within the gangliocytoma. This rare tumour represents a differential diagnosis of a non-secreting pituitary tumour in front of a pituitary incidentaloma.

CASE PRESENTATION Presenting features, medical/social/family history

A 59-year-old woman was referred to our endocrine unit for an incidentally discovered pituitary mass on encephalic MRI. This was performed as Sjögren syndrome was diagnosed in a context of inflammatory shoulder's arthralgia associated with headaches. She had delivered twice and experienced a normal menopause at the age of 50, with initial hot flushes shortly treated by paroxetine. Her only complaint was inflammatory shoulders' arthralgia at the time of the clinical examination in the endocrine unit. Her weight was 58 Kg and her height was 1.61m (BMI 22.4 Kg/m²). She had a normal blood pressure reaching 120/77 mmHg. Neither hypercortisolism nor acromegalic features were observed on clinical examination. She had no spontaneous nor provoked galactorrhea. She had no sign of diabetes insipidus.

INVESTIGATIONS If relevant

Pituitary focused MRI revealed a 14x11x15 mm pituitary tumour, close to the left cavernous sinus. A slight infrasellar as well as a large type IV supra-sellar invasion, lifting up the optic chiasm was observed. A holosellar hyper-T1 signal (Fig 1 ), and an iso/hyper-T2 heterogeneous signal in the absence of gadolinium uptake were noticed. This contrasted with the hypo-T1 and T2 signals without gadolinium uptake present in most cases of silent pituitary adenoma [5]. Her visual acuity was normal and the absence of a bitemporal hemianopsia was confirmed by kinetic perimetry, carried out with Goldman's perimeter.

She was admitted in our department and hormonal investigations were performed. Her serum prolactin was 15 ng/ml (N: <27 ng/ml); urinary cortisol was 73 nmol/24h (N: <260); morning cortisol plasma level was 218 nmol/l (N: 220-610 nmol/l). Her plasma ACTH level of 51 pg/ml was at the upper limit of the normal range (N: <52 pg/ml). Plasma sodium and potassium were in the normal range. After tetracosactide (synacthen*) test, her cortisol plasma level rose at 866 nmol/l (N after 60 mn: >550 nmol/l). TSH was 0.8 mUI/L (N: 0.4-4 mUI/L) and FT4 10.3 pmol/l (N: 7-18 pmol/l); alpha sub-unit plasma level was 2.84 UI/l (normal for menopausal women: N<1.6 UI/l), FSH and LH plasma levels were 62.5 and 52.1 UI/l respectively, with a low estradiol level of 22 pmol/l (normal for menopausal women: N<110 pmol/l). Plasma IgF1 was 160 ng/ml (N: 35-248 ng/ml). During an oral 75g glucose tolerance test, her plasma glucose rose to 8.3 mM and her lowest plasma GH was 0.4 mM after 120 min.

Hence, in front of the mildly low morning cortisolemia, hydrocortisone 10 mg per day was prescribed. Hypophysitis, in the context of Sjögren syndrome, could not be eliminated but no diabetes insipidus was present and no infundibulitis were noted on MRI.

DIFFERENTIAL DIAGNOSIS If relevant

. Silent gonadotropic pituitary adenoma, . Prolactinoma, . Acromegaly, . Intermittent Cushing's disease, . Hypophysitis.

TREATMENT If relevant

She was referred to our neurosurgeon in order to prevent optic chiasm compression, as a partially necrotic, non-secreting, suprasellar pituitary adenoma was suspected.

Transphenoidal surgery was performed and the presence of a tumour capsule and a granular cell lesion were noticed by the experienced neurosurgeon. Removal of the tumour was considered sub-total, since a zone of close contact with the cavernous sinus was noticed.

Histology (Fig 2a and2b) found a tumour exclusively composed of mature neurons with ganglion-cell aspect on a fibrillary background, with a fine vascularization and scattered zones of lymphocytic inflammation. Ki67 staining was 1%. No endocrine cell population was evidenced and some normally settled acini composed of corticotroph endocrine cells were seen, corresponding to the pars intermedia. Immunochemistry revealed a negative staining for prolactin, beta-TSH, beta-LH and beta-FSH, as well as Pit-1. Some ganglion cell-like neurons were positive for ACTH (Fig 2c) and cytokeratins (CAM5/2). A GH staining (Fig 2d)

was present in a small proportion of the ganglion cells-like neurons but GHRH was diffusely expressed. Neuronal differentiation was also confirmed by synaptophysin, NF and chromogranin A immunopositivity.

OUTCOME AND FOLLOW-UP

The immediate post-operative period was marked by a transient insipidus diabetes, controlled with desmopressin treatment. Postoperative IgF1 serum level remained normal (175 ng/ml. N: 35-248 ng/ml) and corticotropic insufficiency remained, as her postoperative morning cortisol level was 33 nmol/l with a plasma ACTH level of 13 pg/ml. Hydrocortisone treatment was therefore maintained. Three months later, persistent corticotropic insufficiency was noted and we noticed a positive desmopressin test, as her cortisol rose from 134 to 282 nmol/l (47%, N: <20%) and ACTH rose from 16 to 30 pg/ml (53%, N: <35%), respectively. Nine months after neurosurgery, the patient was still on hydrocortisone for corticotropic insufficiency: her last cortisol plasma level only rose from 72 nmol/l to 343 nmol/l (N after 60 mn: >550 nmol/l) after tetracosactide (synacthen*) test.

During another oral 75g glucose tolerance test, her plasma glucose rose to 8.1 mM and her lowest plasma GH was 0.3 mmol/l after 120 min. MRI revealed a stable millimetric paracavernous residue. One year after her initial consultation, she is now regularly monitored in order to detect a potential recurrence of the tumour.

DISCUSSION Include a very brief review of similar published cases

To date, 129 cases of gangliocytomas have been reported between 1990 and 2016 [2,3]. Since the exact pathogenesis was unknown, various authors used various ontologies in the past literature (ganglioneuroma, hamartoma, choristoma) [1]. Immune markers of both neuronal and glial cells have been identified in this particular tumour [4]. Few studies have addressed the origin of this tumour. One of the physiopathogenetic hypothesis relies on transdifferenciation of a common neuro-endocrine precursor or stem cell. Recently, an intriguing positive immunohistochemistry for anterior pituitary hormones on the ganglion cell-like component has been recently described [4]. In 2015, immunohistochemistry targeting a neuronal antigen protein have been also found on typical pituitary endocrine tumour: neuronal expression may also be an innate property of pituitary endocrine cells. The presence of ganglion cells inside pituitary endocrine tumour could represent a neuronal differentiation of an endocrine tumour, perhaps arising from an uncommitted progenitor cell [6]. In the literature, women are more frequently affected with gangliocytomas than men and acromegaly is the most frequent endocrine manifestation. The presence of a neural component does not seem to modify aggressiveness of the tumour or its risk of recurrence after resection [3]. No significant difference has been found between pure gangliocytoma and gangliocytoma mixed with pituitary endocrine tumor in terms of patients' characteristics [2], including age of onset, sex ratio, sellar mass syndrome, hormonology, (in)complete surgery. On MRI, authors have reported a sellar mass with hypoT1 signal (51% of cases), hyperT2 signal (69,6%), with less frequently cavernous sinus invasion (21%). Qiao described 23 patients, including 7 pure gangliocytoma [2]. Among them, three had hyperprolactinemia and all were controlled at follow-up (mean: 4.2 years), either spontaneously or with bromocriptine. The same outcome was noted for one more patient with GH/Prolactin co-secretion. Finally, three patients without any significant hormonal hypersecretion did not recur at follow-up, but no detailed hormonal description was provided. To our knowledge, this is the first report of a patient carrying a gangliocytoma without clinically significant hypersecretion, with a pure ganglion cell-like component, devoid of pituitary endocrine cell tumour, contrasting with positive immunochemistry for both hypothalamic and pituitary hormones (GHRH, GH and ACTH). Therefore, abnormal protein maturations and/or secretion of biologically inactive pituitary hormones in the neurons may explain the clinical setting.

LEARNING POINTS/TAKE HOME MESSAGES 3 to 5 bullet points -this is a required field

• Gangliocytoma is a rare cause of pituitary tumour and knowledge on this topic is scarce.

Its diagnosis may be difficult in the absence of any detectable hormonal secretion.

Hence, it may be revealed by a pituitary incidentaloma. Its MRI characteristics may be close to those of silent pituitary adenomas.

• If a detectable hormonal secretion is present, then gangliocytoma is most frequently revealed by acromegaly, hyperprolactinemia or Cushing's disease.

• Gangliocytoma is a very rare tumour and its diagnosis is often performed during histological examination. The most frequent histological finding is a mix of ganglion celllike neurons and pituitary endocrine cells.

• We describe a pure gangliocytoma: immunostaining was positive for ACTH, GH and GHRH. As it was not associated with peripheral hormonal secretions, this case suggests that hypothalamic and pituitary hormones from the tumour are biologically inactive. The pituitary MRI at presentation showed a type IV suprasellar mass of 14x11x15 mm, with a slight infrasellar invasion, very close to the optic chiasm. There was no obvious compression. On the T1-weighted sequence, a holosellar hyper-T1 signal was noted (A: upper image). No gadolinium uptake by this well limited mass (B: middle image), as well as an iso/hyper-T2 heterogeneous signal were noticed (C: lower image). The pituitary MRI at presentation showed a type IV suprasellar mass of 14x11x15 mm, with a slight infrasellar invasion, very close to the optic chiasm. There was no obvious compression. On the T1-weighted sequence, a holosellar hyper-T1 signal was noted (A: upper image). No gadolinium uptake by this well limited mass (B: middle image), as well as an iso/hyper-T2 heterogeneous signal were noticed (C: lower image).

Figure 1 :

 1 Figure 1: Initial pituitary MRI of the 59 yrs old patient

  Fig 1 209x297mm (300 x 300 DPI)

Figure 2 :

 2 Figure 2:Histopathology and immunochemistry a) x40 magnification -H&H stain: gangliocytoma. Histological examination showed a fibrillary background with abundant population of mature neurons resembling hypothalamic ganglion cells. A fine vascularization and scattered zones of lymphocytic inflammation were seen. b) x100 magnification -H&H stain: large mature neurons ganglion cell-like within an abundant neuropil. Mature neurons presented with a voluminous round nuclei and an evident eosinophilic nucleoli as well as a strong eosinophilic granular cytoplasm. Rare bi-nucleations were observed. c) x200 magnification -ACTH immunostaining: the ganglion cell-like neurons expressed ACTH in their cytoplasm. d) x200 magnification -GH immunostaining: some clusters of ganglion cell-like neurons also expressed GH. Fig 2a and 2b; Fig 2c; Fig 2d 254x190mm (300 x 300 DPI)

Vancouver style (Was the patient involved in a clinical trial? Please reference related

		articles)
	1	Louis DN, Perry A, Reifenberger G, et al. The 2016 World Health Organization
		Classification of Tumors of the Central Nervous System: a summary. Acta Neuropathol
		2016;131:803-20. doi:10.1007/s00401-016-1545-1
	2	Qiao N, Ye Z, Wang Y, et al. Gangliocytomas in the sellar region. Clin Neurol
		Neurosurg 2014;126:156-61. doi:10.1016/j.clineuro.2014.08.034
	3	Cossu G, Daniel RT, Messerer M. Gangliocytomas of the sellar region: A challenging
		diagnosis. Clin Neurol Neurosurg 2016;149:122-35.
		doi:10.1016/j.clineuro.2016.08.002
	4	Balci S, Saglam A, Oruckaptan H, et al. Pituitary adenoma with gangliocytic
		component: report of 5 cases with focus on immunoprofile of gangliocytic component.
		Pituitary 2015;18:23-30. doi:10.1007/s11102-013-0551-8
	5	Galland F, Vantyghem MC, Cazabat L, et al. Management of nonfunctioning pituitary
		incidentaloma. Ann Endocrinol (Paris) 2015;76:191-200.
		doi:10.1016/j.ando.2015.04.004
	6	Nguyen MT, Lavi E. Pituitary adenoma-neuronal choristoma is a pituitary adenoma
		with ganglionic differentiation. Exp Mol Pathol 2015;99:628-31.
		doi:10.1016/j.yexmp.2015.11.004
	FIGURE/VIDEO

CAPTIONS figures should NOT be embedded in this document Fig 1: Initial pituitary MRI of the 59 yrs old patient

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b) x100 magnification -H&H stain: large mature neurons ganglion cell-like within an abundant neuropil. Mature neurons presented with a voluminous round nuclei and an evident eosinophilic nucleoli as well as a strong eosinophilic granular cytoplasm. Rare bi-nucleations were observed. c) x200 magnification -ACTH immunostaining: the ganglion cell-like neurons expressed ACTH in their cytoplasm. d) x200 magnification -GH immunostaining: some clusters of ganglion cell-like neurons also expressed GH.

PATIENT'S PERSPECTIVE Optional -Copyright Statement

I, Dr BRUNO DONADILLE, M.D, The Corresponding Author, has the right to assign on behalf of all authors and does assign on behalf of all authors, a full assignment of all intellectual property rights for all content within the submitted case report (other than as agreed with the BMJ Publishing Group Ltd) ("BMJ Group")) in any media known now or created in the future, and permits this case report (if accepted) to be published on BMJ Case Reports and to be fully exploited within the remit of the assignment as set out in the assignment which has been read. http://casereports.bmj.com/site/misc/copyright.pdf.

Date


The

The Swiss Jura / Berne border dispute is linked to the discordance between the political border of the Swiss Bern and Jura Cantons, and the linguistic border between French and German speaking populations. From 1815, Bern Canton has had a large French speaking population. The French speakers gained the right to create their own new Canton called 'Canton of Jura' following a 1974 referendum. But the border between Bern and Jura Cantons is still contested. On November 24 th , 2013, a second referendum over selfdetermination has occurred making is clear that 71.8% of Berne Canton residents do not want to join in but for a primarily francophone city, Moutier, where 55.4% want to move over to the Canton of Jura.

Introduction:

The border dispute between the Jura and Bern Cantons is connected to the older problem of co-habitation between German speaking and French speaking population in Bern Canton. The German-speaking group forms the majority of the population, and has always had political power, while the French speaking population has always been, geographically and politically, at the periphery.

The Francophone population was integrated into Bern Canton during the Vienna congress (Actes d' Union, 1815). Political tension between the two groups became obvious after World War Two and took the form of a territorial dispute. During the following decades an important proportion of Bern francophone claimed the creation of their Canton was necessary. While the Bern central power tried to reach a compromise recognizing, for instance, the 'Jura people' as a political entity, and, making French the official language of the French speaking districts in 1950, the desire for a separate state grew among the French speaking population.

A first referendum was held in 1959 but the French speaking "separatists" did not gain the majority. But in 1974, the creation of the Jura Canton became a reality thanks to a second and successful referendum. Two additional referendums regarding boundaries were held in 1975. Since the creation of the Jura Canton as the 26th Canton of the Swiss Confederation, political tension have taken the form of a positional boundary dispute between two sovereign states. Swiss Cantons are independent states united into a confederation. Indeed, since then the Jura Canton and a part of the French speaking population remaining in Bern Canton have fought for 'reunification of the Jura people' but both the Bern government and a part of Francophones have refused this prospect.

History:

The Jura / Bern Canton border dispute did not exist before the creation of the Jura Canton (the 26th Canton of the Swiss confederation in existence since January 1st, 1979). Until then, the territorial 'Jura question' (question jurassienne) belonged to Bern exclusively. The presence of French speaking population across the Bern territory is a consequence of the Vienna congress in 1815. The 'Union acts' (articles 75 et 76 of the Vienna Congress Final Act) decided the transfer of the French speaking population formerly integrated to the Prince-Bishopric of Basel to the Bern Canton. The integration of Francophone population obviously became a problem after World War II, even if earlier tensions had riseen up both during the 19 th century and in particular in 1917.. The French political agitation was in protest against German language imposition and also fuelled by demands for political power.. Since the 1940's, the creation of a Jura Canton had become a key demand of French "separatists" of the Canton of Bern. In 1947, a separatist Comité de Moutier was created (in the eponym city which remained paradoxically in Bern after the 1974-75 referendums). Even if the Bern central power tried to reach a compromise by recognizing the existence of a French separate nation, i.e. a 'Jurassian people' and making French only official language in the French speaking districts, the political demands for partition remained strong. In September 1957, the Rassemblement Jurassien (RJ) launched a cantonal popular initiative to vote in favour of the creation of such a Canton. This first popular initiative occurred in July 1959, and was rejected by a majority of voters: 51,9% of 'no' in the six French-speaking districts at this time (Gasser 1978, 58). But 'separatism' was not dead. A separatist Jura Liberation Front (Front de libération du Jura (FLJ)) was created in 1962. Far from the peaceful commonly shared image of Switzerland, FLJ carried out various bomb and arson attacks on army buildings or on the houses of prominent anti-separatists leaders. Acting non-violently, the Ram Group" (Groupe béliers), a youth organization, was created in 1963 supporting the separatist demand.

On the other side, the French anti-separatist also organised. They founded a political party in 1953: the Union of Jura Patriots (Union des Patriotes Jurassiens (UPJ)), which in 1974 became Democratic Force (Force démocratique (FD)). They created their own youth organization: the Wild Boar Group (Groupe sanglier) in 1974 (on the model of the 'Ram Group').

As political agitation continued, in 1967 an official commission was set up to find a solution to the conflict. This commission (called 'Of the 24', because of its number of members), and a second commission set up in 1968, called 'The commission Petitpierre', both concluded in favour of a referendum for self-determination. This solution was accepted by Bern central power in 1970, and thereafter approved by popular vote.

As a result, a second referendum was organized in June 23, 1974, in the six French speaking districts of Berne Canton (but for the bilingual Biel-Bienne district that was excluded). This referendum of self-determination raised three questions and three different votes.

A first vote, held on the June 23 1974, decided with 51,9% of the 'yes' to launch the process to create the Jura Canton.

The districts voting 'no' i.e. across south Jura: Courtelary, Moutier and La Neuveville, however, voted a second time to confirm their will not to enter the Jura Canton; they did so on March 16, 1975. Finally, all contiguous municipalities to the new Cantonal border, along the border of the former districts, had the possibility to join the newly created Jura Canton. Ten municipalities changed side: eight municipalities from southern districts voted to enter the Canton of Jura (Courrendlin, Châtillon, Corban, Courchapoix, Les Genevez, Lajoux, Mervelier and Rossemaison), one of the northern municipality, Rebévilliers, however, chose to join Bern, and one entered the German district of Laufen.

In the end however, despite those referendums, the Jura question was not resolved. A part of the French speaking population (both found in the Cantons of Jura and of Bern) considered that instead of freedom, the process fostered the actual separation of the 'Jura people,' into two entities. These groups asserted their desire to continue the struggle until the 'Jura people' were reunited. The authorities of Jura Canton themselves were not indifferent to the situation of the francophone population in Bern Canton, and did not hesitate to defend the specific interests of that population when it was not under its jurisdiction (Pichard 2004, 93).

Physical Topographical/Physical features of the border, Culture: Despite the delineation of French and German speaker in cultural regions, the Jura question is made more complex because the Jura French speakers are divided into two different religious groups: Roman Catholics in the North (mostly in Jura Canton) and Protestants (dominant in Jura Bernois).

Because of this division between Catholics and Protestants, the French speaking population did not consider the geopolitical problem of the Jura question uniformly because of religious affiliation: the Catholics were much more in favour of a 'great Jura' than Protestants. The southern districts, which voted for Bern in 1974, were (and remain) dominantly Protestant. Interestingly, French Protestants share their religion with the Bern German speaking population. Indeed, there were no Catholics in Bern Canton before 1815, so the religious divide remained as important as the linguistic divide, and may explain why a part of French speaking population wished to remain in Bern; because they were protestants.

This religious factor was strengthened by an ideological factor: the symbiosis between a large part of French speaking Protestants and German speaking population symbolized by their common support to 'rightist' political parties, and especially the Democratic Union of the Center (Union Démocratique du Centre (UDC).. Despite its name claiming its affiliation to the 'center', UDC is morally strongly conservative and economically in favour of free market economy. UDC is also strongly in favour of Bern. This level of detail is confusing I think)

Regarding the 'Jura question' the UDC, one of the largest Swiss political party, and a prominent force in Bern (at the federal level) with 26,8% of the vote in the 2010 Cantonal elections, basically has been and remains in charge of the affairs at the Jura Bernois level. Its results were even slightly stronter in Jura Bernois, where it has been and remains the first political party (with 22,7% of votes in 2010 for Bern Canton).

The rightist Liberal Radical Party (Parti Libéral Radical) was also strongly anti-separatist. The PLR resulted from a fusion of the Radical Democratic Party (Parti Radicaldémocratique) and the Swiss Liberal Party (Parti libéral Suisse) which gained 12,7% of the votes in the 2010 Cantonal elections. In the 2010 legislative elections results, together UDC and PLR represented more than 35% of potential anti-separatist vote.

During most of the 20th century the pro-Jura vote was more often, but not always. a 'progressive' vote, but generally speaking it has been supported by lower economical class and left leaning parties. The bourgeoisie and its representatives did not (and today do not) accept pro-Jura vote.

However, here again the leftist political parties are not homogeneously in favour of a 'Great Jura'. For instance, the Socialist party is divided between anti and pro-Jura. The Socialist Party, (Parti socialiste du Jura bernois (PSJB)) is opposed to the 'great Jura' (It gained 13, 6% of the vote during the 2010 election in Jura Bernois). However, the Autonomous Socialist Party of South Jura (Parti Socialiste Autonome du Jura du Sud (PSA)), which is the second political party at the scale of the Jura Bernois with 20,4% of votes in 2010, is in favour of reunification (hence the use of 'South Jura' in its name, such an expression is typically pro-Jura and banned by pro-Bern politicians).

Finally, the Jura's Alliance (Alliance Jurassienne), a federation of smaller parties, is in favour of unification as well, but has represented and today still only represents a small percentage of the Jura Bernois electorate.

All in all, the three main political parties remain opposed to unification of Jura (UDC, PLR and PSJB) roughly representing … 49% of the votes in the Jura Bernois, which suggests that the probability for the 'yes' to win on Nov. 24 2013 to win remained rather low i .

Due to this religious and ideological situation, the scenario for a border readjustment in the period 2013-15 remains much more likely than a reunification. The city of Moutier, however, where Catholics are dominant, may join the Canton of Jura'. The eastern Jura bernois seems to be the only region were a significant border change is likely to take place. The Jura Bernois is about 541 kilometer square!

Conclusion (current state of affairs)

In Feb. 2012 the governments of Bern and Jura Cantons decided in common that a new referendum for self-determination was the best solution to solve the 'Jura question'. After a long time and 'slow down' position on this mater the government of Bern Canton agreed that a new referendum was an opportunity for the majority to say 'no' which may stop the nationalist agitation.

However, since 1974-1975, it is known that such votes are rather complicated. Indeed, the referendum of November 24, 2013 again witnessed the populations of both the Cantons of Jura and Bern say 'no' to reunification. The one exception is the city of Moutier, which confirmed 'yes' with more than 55% of the vote.

Because the 'No' won, this marks the end of the constitutional process. But, municipalities of Jura Bernois now have the option to choose individually to join in the Canton of Jura each one of them in small-scale referendums by the end of 2015. This Jura question is of great interest because Referendums give local population to choose its political destiny (sovereignty) underlying a 'rupture' with the traditional top down process of border drawing. Even when a territory is small, as is the case in the 'Jura question,' voting is a significant example of the use of Self-determination referendum to solve a territorial or a border dispute [START_REF] Cassese | Self-Determination of Peoples: A Legal Reappraisal[END_REF][START_REF] Cattaruzza | Les référendums d'autodétermination : démocratisation ou balkanisation du monde ?[END_REF][START_REF] Laponce | From Avignon To Schleswig And Beyond: Sovereignty And Referendums[END_REF]and 2004).

Text of law, international agreements,

No international agreement at the exception of Vienna Congress final act (1815), see the bibliography of key legal texts with web links below.

A glossary (about 50-75 words) Autonomist Key-word designating French speaking people today in favor of the unification of Jura bernois district with the Jura Canton. This word took place of the 'separatist' after 1974.

Canton of Bern

The Bern Canton is one of the 26 Swiss Cantons. Its 992,000 inhabitants (2012) are roughly 85% germanophones and 8% francophones. Before the creation of the Jura Canton the proportion of French speaker was more important (77,6% of Germanophones and 13,7% of Francophones in 1970) ii . Since the January 1st, 2010, Bern Canton is divided into 10 districts (Verwaltungskreise). One district has French as official language (Jura Bernois), one is bilingual (Biel/Bienne) and eight have German as official language (Emmental, Bern-Mittelland, Frutigen-Niedersimmental, Interlaken-Oberhasli, Oberargau, Obersimmental-Saanen, Seeland, Thun).

Biel-Bienne (district in Bern Canton)

Biel-Bienne is one of the 10 districts of the Bern Canton since January 1st, 2010. Instead of two municipalities as before (Bienne and Évilard, including 51,031 inhabitants among which 55,6% of germanophones and 28,4% of francophones) the up-to-date enlarged district incorporates 19 municipalities and a total population of 95,718 persons, among which only 15% are francophones. This makes impossible any extension of the Jura question on the neighboring and partially francophone municipalities of the Biel-Bienne district.

Jura (Republic and Canton)

The Republic and Canton of Jura is the most recent of the 26 Swiss Cantons. This new Canton was erected on January 1, 1979 as a conclusion of the June 23, 1974June 23, referendum (and border referendum of 1975) ) won by the partisans of its creation. Its capital is Delémont (11,590 inhabit. in 2010). The population of this Canton is 70,197 inhabit. (2010) Homogeneously francophone and Catholics in majority.

Jura bernois (district of the Bern Canton)

The Jura bernois is one of the 10 districts of the Bern Canton since January 1st, 2010. Its capital is Courtelary. It includes 49 municipalities and 51,539 inhabitants ( 2010) francophones (about 90%) and Protestants (about 65%) in majority.

Jura question ("question jurassienne")

This formula refers to the geopolitical problem linked with the political status of French speaking population in Bern Canton including the 'separatist' movement from 1947 to 1974, and since this period the 'autonomist' movement of Jura bernois district. All main political parties in Bern dream to put an end to the Jura question, that is in this perspective that the Nov. 24, 2013 referendum is organized.

Rassemblement jurassien (RJ) :

Is the first political movement openly claiming its separatism after World War Two. After the creation of the Republic and Canton of Jura (referendums of 1974-75), the RJ survives in the Jura Bernois as Mouvement Autonomiste Jurassien.

Separatist

Key-word designated the former supporters among the French speaking population of the creation of a Jura Canton before 1974. As this word sounded pejoratively to many Swiss people, the new movement in favor of the union between Jura Canton and Jura bernois refer to the concept of "autonomist".

Switzerland

The country has 7 783 000 inhabitants (2010) and a total area of 41,285 km2. 65.6% its total population speak German, 22.8% French, 8,4% Italian and 0,6% Romansh. Switzerland has a specific political organization; it is not a federation, but a confederation in which the 26 Cantons are sovereign states. Each Canton has its own constitution, parliament, government and courts. The sovereignty of Swiss Cantons emphasized by the rule of Cantonal linguistic sovereignty (Sprachhoheit) by which each Canton is free to determine its official(s) languages(s), for a determined area or domain, 'has the right to protect and defend its own linguistic character and to assure its survival' (Schmid, 1981, 21). Notably, most of Swiss Cantons are mono-language; only three of them are bilingual (Bern, Fribourg, and Valais; and, Grisons is unique with three official languages).

Legal bibliography:

List of key organizations:

Assemblée inter-jurassienne (AIJ) / Inter Jurassian Assembly The AIJ is consultative assembly created in March 1994 joining two delegations of 12 members from the Jura Canton and The Jura Bernois district. Its permanent seat is in Moutier (Jura Bernois). AIJ main role was to find a political solution to the 'Jura question' [START_REF] Gogniat | Assemblée interjurassienne, in dictionnaire historique du jura[END_REF]. In its May 2009 final report the AIJ proposed two solutions: to create a new Canton joining Jura and Jura Bernois district or the extension of autonomy for the Jura Bernois (the so-called "statu-quo+" solution). After hesitating, the Bern government finally approved the idea of a new vote in 2012. Indeed opinion surveys showed that the maintenance of Jura Bernois in Bern appeared to be dominant among francophones.

Conseil du Jura Bernois (CJB)

This consultative Council (Conseil) was created in 2006 by the Bern government to deal with cultural and scholar affairs within the Jura bernois. The CJB is supposed to play a more important (but imprecise) role if the Jura bernois remains in Bern Canton after Nov. 2013…


Valérie Bernard

Bruno Donadille

Tiphaine Le Poulennec

Mariana Nedelcu

Laetitia Martinerie

Sophie Christin-Maitre
email: sophie.christin-maitre@aphp.fr

Prof Christin-Maitre

MANAGEMENT OF ENDOCRINE DISEASE: Transition of care for young adult patients with Turner syndrome

Keywords: Turner syndrome, Karyotype, Transition, Rare disease

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

Introduction

Transition is defined by the Society for Adolescent Medicine as a « purposeful, planned movement of adolescents and young adults with chronic physical and medical conditions from child centred to adult oriented health care system » [START_REF] Blum | Transition from child-centered to adult health-care systems for adolescents with chronic conditions. A position paper of the Society for Adolescent Medicine[END_REF]. Most studies on transition have evaluated immediate transfer from paediatric units to adult units. However, mid to long term transfer in adult health care system of paediatric patients, should be assessed.

Turner syndrome (TS), affecting 1/2000 à 1/2500 live born girls, is a chromosomal aberration with total or partial loss of one of the X chromosomes [START_REF] Nielsen | Chromosome abnormalities found among 34,910 newborn children: results from a 13-year incidence study in Arhus, Denmark[END_REF][START_REF] Stochholm | Prevalence, incidence, diagnostic delay, and mortality in Turner syndrome[END_REF]. Short stature and primary ovarian insufficiency (POI) are classic features of TS. The prognosis is mainly related to cardiovascular diseases, involving congenital heart diseases, hypertension, aortic dilatation and potential aortic dissection [START_REF] Mortensen | Cardiovascular phenotype in Turner syndrome-integrating cardiology, genetics, and endocrinology[END_REF].

Recommendations to standardize TS medical care and follow-up have been initially published in 2001, followed by American guidelines and more recently international guidelines [START_REF] Saenger | Recommendations for the diagnosis and management of Turner syndrome[END_REF][START_REF] Bondy | Care of girls and women with Turner syndrome: a guideline of the Turner Syndrome Study Group[END_REF][START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF]. The most recent was established following a meeting in Cincinnati, sponsored primarily by the European Society of Endocrinology (ESE) and co-sponsored by the PaediatriEndocrine Society, the European Society of Paediatric Endocrinology (ESPE) and the Endocrine Society (ES) [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF].

Different modes of transition of patients with TS from paediatrics to adult units

In the nineties, the group of KR Rubin has proposed several modes of transition for patients with TS [START_REF] Rubin | Turner syndrome: transition from pediatrics to adulthood[END_REF]. The first one is a vertically integrated care model, with paediatric and adult units within the same hospital. It offers the advantage of providing a more simple transition and exchange of health care information from paediatric to adult care by sharing similar physicians, a familiar hospital to the transitioning patient, and the same electronic health records. In the second model, patients have access to paediatric and adult cares within the same health care system but not in a same site of care. The last model involves distinct paediatric and adult care systems. To date, around the world, many patients with TS do not have access to an integrated paediatric-adult multidisciplinary care program.

Studies evaluating TS transition

A lack of appropriate transition for patients with TS has been reported in several studies conducted in Belgium, Australia, France and Poland [START_REF] Verlinde | Health and psychosocial status of patients with turner syndrome after transition to adulthood: the Belgian experience[END_REF][START_REF] Pedreira | Health-care problems of Turner syndrome in the adult woman: a cross sectional study of a Victorian cohort and a case for transition[END_REF][START_REF] Devernay | Determinants of medical care for young women with Turner syndrome[END_REF][START_REF] Gawlik | Quality of medical follow-up of young women with Turner syndrome treated in one clinical center[END_REF]. The largest study included 568 women with TS, with an average age of 22.6 ± 2.6 years (range 18.3-31.2) [START_REF] Devernay | Determinants of medical care for young women with Turner syndrome[END_REF]. Only 3.5% of patients with TS underwent all recommended assessments within the 4-year period and 16% had undergone none of the recommended assessments. Multivariate analysis identified the type of physician, as the only factor associated with a better follow-up. This study illustrated the role of endocrinologists, in transition of patients with TS.

What are the key messages to be delivered to patients with TS during transition?

Several visits are usually necessary as many messages need to be delivered to the patient.

The karyotype

A special time should be dedicated to delivering the patient's karyotype. A study from the Turner Syndrome Life Course Project, including 656 women with TS, has recently reported a karyotypephenotype analysis [START_REF] Rosa | The Turner syndrome life course project: Karyotype-phenotype analyses across the lifespan[END_REF]. The authors mentioned that the most common karyotypes are the classical monosomy (45,X: 41.6% of participants) and the 45,X/46,XX mosaicism. Patients with a ring X chromosome (45,X/ 46,X,r(X): 7.3% of participants) were more likely to have a metabolic syndrome, elevated liver enzymes and higher HbA1C than patients with the classical 45,X karyotype. This study suggested that follow-up of patients with TS could be stratified according to their karyotype. However, according to the international recommendations for the care of patients with TS, information concerning the long term follow-up should be given to all patients with TS, irrespective of their karyotype [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF].

Hormonal replacement therapy

As the majority of patients with TS will experience primary ovarian insufficiency (POI) [START_REF] Bondy | Care of girls and women with Turner syndrome: a guideline of the Turner Syndrome Study Group[END_REF][START_REF] Hreinsson | Follicles are found in the ovaries of adolescent girls with Turner's syndrome[END_REF], a goal is having patients comply with taking hormonal replacement therapy (HRT), including estrogens and progestins, up to the age of 50 [START_REF] Klein | Estrogen Replacement in Turner Syndrome: Literature Review and Practical Considerations[END_REF]. The positive benefit-risk ratio of HRT on skin, bone and heart function should be discussed [START_REF] Eshre) Guideline Group On | ESHRE Guideline: management of women with premature ovarian insufficiency[END_REF]. An important issue is to define with the patient the route of HRT, with oral or transdermal estrogens [START_REF] Klein | Estrogen Replacement in Turner Syndrome: Literature Review and Practical Considerations[END_REF]. Assessing whether the patient wishes or not to have withdrawal bleedings should be taken into account.

Bone density

Measuring bone density can be helpful in convincing patients about the benefits of estrogens.

A recent Australian study has evaluated retrospectively bone density and bone fractures, in an adult population of 76 patients with TS. Despite a median age of 29 years, odds ratio of low bone mass in patients with TS was 9.8 times that of age-and gender-matched controls [START_REF] Nguyen | Delay in estrogen commencement is associated with lower bone mineral density in Turner syndrome[END_REF]. In this study, the prevalence of non-continuous HRT was high as it reached 40% of the population.

Reproduction

Initially, the physician should take into account the cardiac phenotype and the aortic diameters, as a pregnancy may be contra indicated in some patients with TS with cardiovascular diseases [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF]. If Y chromosome material is present in the karyotype, bilateral gonadectomy should be discussed in order to reduce the risk of gonadoblastoma [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF].

If the patient is still spontaneously menstruating, information should be given that the rate of spontaneous pregnancy is around 6% [START_REF] Bernard | Spontaneous fertility and pregnancy outcomes amongst 480 women with Turner syndrome[END_REF]. In such patients, oocyte cryopreservation should be proposed. Indeed, mature oocytes may be obtained after ovarian stimulation. The oocytes are retrieved by transvaginal retrieval and then vitrified for fertility preservation. No pregnancy has been reported, so far, after using cryopreserved oocytes, in patients with TS. When this technique is performed, because of age or non-oncological medical conditions, more than 10 to 12 oocytes are necessary to obtain a pregnancy [START_REF] Martinez | Update on fertility preservation from the Barcelona International Society for Fertility Preservation-ESHRE-ASRM 2015 expert meeting: indications, results and future perspectives[END_REF]. Therefore oocyte cryopreservation does not apply to patients with TS, when their ovarian reserve is too low.

In some cases, ovarian tissue has been preserved during childhood, before the occurrence of POI. No pregnancy has been reported, so far, in patients with TS after using frozen ovarian fragments. Furthermore, the potential impacts of ovarian tissue freezing performed during childhood, on oocyte competence remains quite unknown. Few studies have evaluated such oocyte competence in patients with TS [START_REF] Kavoussi | Oocyte cryopreservation in a woman with mosaic Turner syndrome: a case report[END_REF][START_REF] El-Shawarby | Oocyte cryopreservation after controlled ovarian hyperstimulation in mosaic Turner syndrome: another fertility preservation option in a dedicated UK clinic[END_REF][START_REF] Oktay | Fertility preservation by ovarian stimulation and oocyte cryopreservation in a 14-year-old adolescent with Turner syndrome mosaicism and impending premature ovarian failure[END_REF][START_REF] Balen | Conservation of fertility and oocyte genetics in a young woman with mosaic Turner syndrome[END_REF][START_REF] Doğer | Reproductive and obstetric outcomes in mosaic Turner's Syndrome: a cross-sectional study and review of the literature[END_REF]. Genetics of 11 oocytes obtained in ayoung woman with TS [START_REF] Balen | Conservation of fertility and oocyte genetics in a young woman with mosaic Turner syndrome[END_REF] have been evaluated by fluorescent in situ hybridization analysis. Among them, none had chromosomal abnormalities. Although this study is rather reassuring, few data are available concerning fertility preservation techniques in patients with TS. A single live birth has been reported, after autograft of ovarian tissue cryopreserved during childhood, in a patient with sickle cell anaemia [START_REF] Demeestere | Live birth after autograft of ovarian tissue cryopreserved during childhood[END_REF]. A single case of live birth after allografting of ovarian cortex between monozygotic twins with TS (45X, 46XX) and discordant ovarian function has been reported [START_REF] Donnez | Live birth after allografting of ovarian cortex between monozygotic twins with Turner syndrome (45,XO/46,XX mosaicism) and discordant ovarian function[END_REF].

In case of POI, patients should be oriented towards oocyte donation programs. However, repeated careful cardiovascular evaluation is necessary.

Cardiovascular follow-up

During transition, cardiovascular status needs to be re-evaluated [START_REF] Mortensen | Cardiovascular phenotype in Turner syndrome-integrating cardiology, genetics, and endocrinology[END_REF][START_REF] Donadille | Cardiovascular findings and management in Turner syndrome: insights from a French cohort[END_REF]. The need of a long term follow-up of aortic dilatation by cardiac magnetic resonance imaging (CMR) and/or transthoracic echocardiography (TTE) has to be explained. The frequency of cardiovascular evaluation is decided according to the initial aortic diameter at the age of 16, and the presence of associated cardiovascular risks such as aortic coarctation, hypertension and bicuspid aortic valve. According to the international guidelines, it ranges from every year to ten years [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF].

The main difficulty with TS transition lies in the multiplicity of the necessary exams. Care in adults includes evaluation of liver function, lipid and glucose levels, bone density, audiometric, ophthalmologic and dermatological evaluations as well as autoimmune screening, essentially for thyroid and celiac disease [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF]. Hearing problems are frequent and are associated with poor quality of life and social isolation. Motivating patients for their lifelong follow-up is a major issue of mid to long term transition.

What are the tools available in order to improve TS transition?

A tool for the transition readiness assessment is provided by The Endocrine Society. It includes 10 questions about health, 16 questions about using health care and 15 questions concerning social and emotional factors. A provider assessment of skill test, a clinical summary and transfer record, as well as a recommended approach for transitioning into adult practice are available on the website, (http://www.endocrinetransitions.org).

Partners of choice in the field of non-medical actors are advocacy groups (28) (CAIRN.Info).

Several of them are involved with TS, such as the Turner Syndrome Support Society in the UK (tss.org.uk), the Turner foundation (turnersyndromefoundation.org) or the Turner Syndrome Society of the United States in the USA (turnersyndrome.org) and AGAT in France (agat-turner.org), or in other countries (please see Orphanet links). Parental and family involvements are crucial for a successful TS transition.

Therapeutic education or personalised care planning may potentially increase the transition's success rate [START_REF] Who | Therapeutic patient education: continuing education programmes for health care providers in the field of prevention of chronic disesases[END_REF]. Among endocrine disorders, the most common chronic diseases where transition has been particularly studied are diabetes [START_REF]Children and Adolescents:Standards of Medical Care in Diabetes-2018[END_REF], 21 hydroxylase deficiency [START_REF] Speiser | Congenital adrenal hyperplasia due to steroid 21-hydroxylase deficiency: an Endocrine Society clinical practice guideline[END_REF][START_REF] Bachelot | Impact of transition on quality of life in patients with congenital adrenal hyperplasia diagnosed during childhood[END_REF] or hypopituitarism [START_REF] Ho | Consensus guidelines for the diagnosis and treatment of adults with GH deficiency II: a statement of the GH Research Society in association with the European Society for Pediatric Endocrinology[END_REF]. A Cochrane review on personalised care planning has shown that the effects are not large, but they appear greater when the intervention is more comprehensive, more intensive, and better integrated into routine care [START_REF] Coulter | Personalised care planning for adults with chronic or long-term health conditions. The Cochrane Database of Systematic Reviews[END_REF]. Our unit has developed a dedicated therapeutic education program for adult patients with TS. The goal of this program is a pragmatic approach for defining competencies and resources. Each session includes 3 to 5 patients and lasts one day, from 9 am to 4 pm. Patients come from all over France. A nurse trained for therapeutic education performs upon the patient's arrival, an educational diagnosis. Thereafter, a series of four thematic workshops, eaclasting 1 hour, is performed by physicians from the unit (an endocrinologist, a gynaecologist and a psychiatrist). The workshops deal respectively with feminine, dietary, somatic and psychological issues, related to TS. The discussions during each session are initiated by a homemade educational map (Figure 1). Each patient shares her own experience with her peers. At the end of the day, patients complete a knowledge assessment on TS. Then, the nurse establishes future targets in order to improve the acceptance of the disease and educative competencies (Supplemental Table 1). We are planning to include expert patients, during those sessions.

Specificity of the transition of patients with TS

A major issue in organising TS transition is to take into account the psychology of those patients.

Adolescence has been thought to be a high-risk period for girls with TS given their delay in pubertal and linear growth. Furthermore, TS is associated with a peculiar neurocognitive profile. In general, patients suffer from an excessive dissatisfaction with self-image and self-esteem [START_REF] Amundson | Impact of growth hormone therapy on quality of life in adults with turner syndrome[END_REF][START_REF] Gawlik | Transitions in endocrinology: treatment of Turner's syndrome during transition[END_REF].

Girls or young women with TS are less socially active than unaffected girls of similar age, in terms of time spent with friends or number of friends. This poor adolescent psychosocial integration is explained by a negative body image [START_REF] Schmidt | Shyness, social anxiety, and impaired self-esteem in Turner syndrome and premature ovarian failure[END_REF] and in some cases poor family acceptance of the diagnosis [START_REF] Mccauley | Psychosocial development in adolescents with Turner syndrome[END_REF]. Furthermore, attention deficit hyperactivity disorders (ADHD) are more frequent in TS. A recent study has reported an increased rate of 24% of ADHD in patients with TS, as compared to 1.3% in the general population [START_REF] Green | Attention deficit hyperactivity disorder (ADHD) in phenotypically similar neurogenetic conditions: Turner syndrome and the RASopathies[END_REF]. Therefore, some patients may have specific intellectual difficulties such as organizational, planning or task prioritization disorders [START_REF] Reiss | Neurodevelopmental effects of X monosomy: a volumetric imaging study[END_REF]. A psychological support is important to enhance the patient's self-esteem, and to reach a satisfactory quality of life level thus optimizing transition [START_REF] Kosteria | Turner Syndrome: transition from childhood to adolescence[END_REF].

On the other hand, inflexibility and preoccupation with keeping things in order are often reported in patients with TS [START_REF] Gawlik | Transitions in endocrinology: treatment of Turner's syndrome during transition[END_REF]. Those characteristics should be an advantage, as they should improve compliance for the mid and long term follow-up, when immediate transition has been successful.

Results of TS transition in our centre

In order to illustrate this review, we evaluated TS transition in adult patients, followed in our unit, in Saint-Antoine Hospital, Paris, France (Figure 2). Our unit belongs to a center of rare diseases, which includes children as well as adults units. It has been labelled by the French Ministry of Health in 2006. It is a member of the European Reference Network on Rare Endocrine conditions (Endo-ERN). Among the 258 patients in our adult unit, 220 (85.3%) have been diagnosed with TS before the age of 20 years. Among those, we distinguished patients who had an "organised transition", therefore coming directly from paediatric endocrine units (n=112) from patients referred by their general practitioner, their gynaecologists or coming spontaneously to our unit (n=108). In order to evaluate if the transition had been successful, each patient's electronic file was studied. The frequencies of visits and/or day hospitals were evaluated. Transition was defined as successful if the patient, after an initial visit, came in our unit, at least once, over the past 4 years. This figure was chosen, according to the mean frequency of cardiovascular evaluation, as recommended in the international guidelines [START_REF] Gravholt | Clinical practice guidelines for the care of girls and women with Turner syndrome: proceedings from the 2016 Cincinnati International Turner Syndrome Meeting[END_REF]. As shown in Figure 2, the mean age at TS diagnosis was similar in both groups. However, the percentage of patients lost to follow-up was statistically lower among the patients referred directly by paediatricians from our reference centre (p < 5x 10 -13 using a Fisher test). Age at first visit in the adult unit was older in the group without transition, illustrating a lack of follow-up. Although many biases may be present in this evaluation, a paediatric diagnosis followed by a synchronized adult follow-up using a common file was associated with a less frequent loss to follow-up and a higher frequency of visits performed in our adult reference centre. A remaining challenge is to understand why some patients and/or their family have not transferred directly from the paediatric care, where TS was initially diagnosed and followed. A recent review identified the barriers to transition from paediatric to adult care [START_REF] Gray | Barriers to Transition From Pediatric to Adult Care: A Systematic Review[END_REF]. The most common fall within the relationship domains, followed by access/insurance and belief/expectations. Carel et al. has shown previously that most socially vulnerable patients may be the one who will be lost at follow-up [START_REF] Devernay | Determinants of medical care for young women with Turner syndrome[END_REF]. However, the cost of care should not be involved in the success of transition in our evaluation, as in France full reimbursement of the patient's care is provided for every patients with TS.

Conclusion

Not so long ago, most young adults with TS were making their way through the healthcare system in an uncoordinated way [START_REF] Blum | Transition from child-centered to adult health-care systems for adolescents with chronic conditions. A position paper of the Society for Adolescent Medicine[END_REF]. Hence, inequity of care would affect patients already bearing the burden of a rare disease. A successful health care transition should help other aspects of life, such as work, school, or other social issues (). Young adults with a chronic rare condition like TS need a multidisciplinary approach to their care plan, including psychosocial aspects. Our preliminary data are in favour of an organised transition from paediatric to adult care units, within the same reference centre of rare diseases. Further studies are necessary in order to evaluate failure of transition, essentially the social and/or psychological characteristics of patients with TS.

Figure 1 .

 1 Figure 1. Educational map

Figure 2 .

 2 Figure 2. Evaluation of transition in adult patients with TS followed in Saint-Antoine Hospital

FIGURE 1 .

 1 FIGURE 1.


MD. Bruno Donadille
email: bruno.donadille@sat.aphp.fr

Alexandra Rousseau

Delphine Zenaty

Sylvie Cabrol

Carine Courtillot

Sylvie Salenave

Philippe Bouchard

Philippe Chanson

Michel Polak

Philippe Touraine

Yves Lebouc

Jean- Claude Carel

Juliane Léger

Sophie Christin-Maitre

Damien Bonnet

Nathalie Bourcigaux

Laure Cabanes

Lise Duranteau

Laurence Iserin

Guillaume Jondeau

Catherine Meuleman

Laurence Monnier-Cholley

CARDIOVASCULAR FINDINGS AND MANAGEMENT IN TURNER SYNDROME: INSIGHTS FROM A FRENCH COHORT

Keywords: Turner syndrome, POF, bicuspid aortic valve, aortic dilatation

Objective: Congenital cardiovascular malformations and aortic dilatation (AD) are frequent in patients with Turner syndrome (TS). The objective of this study was to investigate cardiovascular findings and management in a large cohort of patients, including children and adults.

Design/Methods: We recruited 336 patients with TS from a network of tertiary centers. We reviewed their files, checking for cardiovascular events, cardiac valve abnormalities, and aortic diameters indexed to body surface area (BSA) from magnetic resonance imaging (MRI) (n=110) or echocardiography (n=300).

Results: Informative cardiovascular data were available for only 233 patients.

Vascular surgery was reported in 7.4% of the cohort. The first cause of surgery was aortic coarctation, detected in 6.9% at a median age of 9.5 years (range: 0-60).

Aortic bicuspid valve (BAV) was detected in 21% at a median age of 20 years [25 th -75 th percentiles: 15-30]. At least one aortic diameter exceeded 32 mm in 12% of the cohort. This was detected at a median age of 19 years . When indexed to BSA, at least one aortic diameter exceeded 20 mm/m 2 in 39% of the cohort.

Conclusion:

Our study shows that cardiovascular monitoring for Turner syndrome patients is currently insufficient in France. BAV is present at birth, but often remains undiagnosed until later in life. Therefore improved management in cardiovascular monitoring is required and a more systematic approach should be taken.

INTRODUCTION

Turner syndrome, affecting 1/2500 liveborn girls, results from the total or partial absence of the X chromosome. Mortality rates are three times higher in women with Turner syndrome than in the general female population (SMR = 3.0; 95% CI 2.7-3.4) [START_REF] Schoemaker | Mortality in women with turner syndrome in Great Britain: a national cohort study[END_REF]. These higher mortality rates are largely due to cardiovascular complications, such as aortic dilatation and dissection. The estimated incidence of dissection is 36 per 100,000 Turner syndrome-years, versus only 6 per 100,000 in the general population [START_REF] Gravholt | Clinical and epidemiological description of aortic dissection in Turner's syndrome[END_REF].

In the last few years, several recommendations for the medical follow-up of patients with Turner syndrome have been published [START_REF] Bondy | Care of girls and women with Turner syndrome: A guideline of the Turner Syndrome Study Group[END_REF]. Monitoring is mostly based on expert consensus. Cardiovascular screening includes blood pressure measurement, echocardiography and cardiovascular MRI. It has been recommended that, in the absence of cardiac vascular disease, echocardiography or MRI should be carried out every five to 10 years. Annual imaging is recommended in cases of an aortic diameter greater than 32 mm or 20 mm/m 2 [START_REF] Bondy | Care of girls and women with Turner syndrome: A guideline of the Turner Syndrome Study Group[END_REF][START_REF] Cabanes | Turner syndrome and pregnancy: clinical practice. Recommendations for the management of patients with Turner syndrome before and during pregnancy[END_REF]. However, several studies have suspected suboptimal diagnosis in those patients with high cardiovascular mortality [START_REF] Schoemaker | Mortality in women with turner syndrome in Great Britain: a national cohort study[END_REF][START_REF] Bondy | Deficient medical care for adults with the Turner syndrome[END_REF][START_REF] Freriks | Standardized multidisciplinary evaluation yields significant previously undiagnosed morbidity in adult women with Turner syndrome[END_REF] The aim of this study was to investigate cardiovascular findings and management in a large cohort of French patients with Turner syndrome, including infants, children, adolescents and adults. For this purpose, we reviewed the medical charts of patients. We investigated the occurrence and timing of cardiovascular abnormalities, to facilitate the standardization of cardiovascular follow-up in patients with Turner syndrome. The patients with a standard peripheral leukocyte karyotype, with more than 10% of cells displaying a total or partial loss of the X-chromosome, were eligible. We considered the following data for each patient: age at diagnosis of Turner syndrome, timing of cardiac and aortic surgery, timing and results of the most recent echocardiogram and/or magnetic resonance imaging (MRI). Two-dimensional echocardiography of the aortic root was performed at enddiastole in the parasternal long-axis views, at four levels: the aortic annulus, the aortic sinuses, the supra-aortic ridge and the proximal ascending aorta, as described by Roman et al. [START_REF] Roman | Two-dimensional echocardiographic aortic root dimensions in normal children and adults[END_REF]. We also collected data concerning valve morphology, according to the European Association of echocardiography [START_REF] Lang | Recommendations for chamber quantification: a report from the American Society of Echocardiography's Guidelines and Standards Committee and the Chamber Quantification Writing Group, developed in conjunction with the European Association of Echocardiography, a branch of the European Society of Cardiology[END_REF].

SUBJECTS AND METHODS

The

Patients underwent imaging on a 1.5 Tesla MRI scanner. Aortic diameters were measured at the level of the aortic cusp, the aortic sinuses, the supra-aortic ridge, the proximal ascending aorta, the aortic arch and the descending aorta. Standardized imaging included morphologic ECG-gated sequences in the axial, coronal and left-anterior-oblique axes and cine-MR sequences in a coronal view of the ascending aorta and through the aortic valve, to check for aortic regurgitation or stenosis and to determine valve morphology [START_REF] Matura | Aortic dilatation and dissection in Turner syndrome[END_REF][START_REF] Kaiser | Normal values for aortic diameters in children and adolescents-assessment in vivo by contrast-enhanced CMR-angiography[END_REF][START_REF] Sakamoto | MR Imaging of the Aorta[END_REF].

For each patient, we recorded the largest aortic diameter (mm) at the most recent cardiovascular examination, together with the height, weight and body mass index (BMI) of the patient at the time of that examination. Body surface area (BSA) was calculated according to the formula of Dubois and Dubois [START_REF] Dubois | A formula to estimate the approximate surface area if eight and weight be known[END_REF]. According to previous recommendations [START_REF] Bondy | Care of girls and women with Turner syndrome: A guideline of the Turner Syndrome Study Group[END_REF][START_REF] Cabanes | Turner syndrome and pregnancy: clinical practice. Recommendations for the management of patients with Turner syndrome before and during pregnancy[END_REF], aortic dilatation (AD) was defined as an aortic diameter exceeding 32 mm or 20 mm/m². In cases of "normal" cardiac evaluations with no documentation of aortic diameter, the patient's file was excluded from the analysis.

We first focused on the most recent cardiological screening results and then searched the file to determine the time at which any abnormalities was first noted.

The original cardiovascular reports were looked for in each patient's file. We excluded patients (n=13) whose most recent cardiovascular examination had taken place before 1998 (Fig. 1).

Statistical analysis

The characteristics of the patients are expressed as frequencies and percentages for categorical variables, and as mean ± standard deviation (SD) or median [25 th -75 th percentiles] and range for continuous variables. Relationships between categorical variables were assessed with Chi-squared tests. We used the Kaplan-Meier method to generate survival curves from birth and to determine survival rates and their 95% confidence intervals (Greenwood variance).

The events studied were BAV, aortic coarctation and aortic dilatation. Analysis was carried out with the SAS V9.2 System (SAS Institute, Cary, NC, USA) and R software version 2.7.2 (R Foundation for Statistical Computing, Vienna, Austria; www.Rproject.org) for analyses of agreement and survival.

RESULTS

Our cohort included a total of 336 women with Turner syndrome seen at our center between 1999 and 2009. No detailed cardiovascular monitoring was carried out in 20 of the 336 cases (5.9%)(Fig. 1). Cardiovascular tests were presumably carried out but no cardiovascular data were available for 10 cases (3.0%) and data available before 1998 in 13 cases were excluded. In 60 cases (17.8%), cardiovascular imaging results were reported as "normal" with no written record of the aortic diameter measured. Informative cardiovascular data were therefore available for 233 patients .

The clinical characteristics of these patients are reported in Table 1. In summary, 73 children (31.3%) and 160 adults were analyzed. A monosomy was present in 108 of the 233 patients (46%). The other karyotypes observed were 46,Xi(Xq) (21%), 45,X mosaicism (16.3%), 45,X/46,XrX (9%), 45,X/Y+ mosaicism (4%), 46,XdelX and others (4%). The mean age at diagnosis of Turner syndrome was 8.0 ± 7.9 years (range: 0 to 50 years). Mean age at diagnosis was 4.9 ± 5.1 years for pediatric units and 9.4 ± 8.5 years for adult units (p=0.003). Mean age at most recent cardiac evaluation was 21.6 ± 11.8 years (range: 0 to 67 years).

Mean delays from the age at general diagnosis of the syndrome to the age at diagnosis of aortic coarctation and BAV were 3.94 years (range: 0 to 30 years) and 13 years (range: 0 to 30 years), respectively. Cardiovascular surgery had been carried out on 17 of our 233 patients (7.4%). No dissection was reported. Median age at surgery was 1 year (range: 0 to 48 years). Most (70%) of the surgical procedures were carried out to correct aortic coarctation (n=12), at a median age of 0.5 years. In the cohort, 59% of these operations occurred before the age of five years; ranging from 0 to 16 years. The second most frequent surgical procedure was valve surgery (n=5): three aortic commissurotomies, one aortic valve replacement and one transcutaneous pulmonary valvuloplasty. Combined surgery was carried out in seven patients. In three cases, atrial or interventricular septal defects were corrected together with aortic coarctation. In three other patients, bicuspid aortic valves were corrected together with aortic coarctation. The remaining patient underwent coronary artery bypass during aortic surgery for ischemic coronary disease at the age of 48 years. Three surgical procedures -pulmonary valve surgery or correction of aortic dilatation -were performed in patients, aged 19, 32 and 48 years.

One patient underwent surgery for aortic stenosis at the age of nine years. In one case, surgery for a dilated subclavian artery was performed at the age of 16 years.

Echocardiography and MRI data were available for 222 and 101 cases, respectively. In total, 90 patients (38.6% of the cohort) underwent both echocardiography and MRI. Cardiological evaluation revealed no abnormality in 131 cases (56.2%).

BAV were detected in 49 patients (21.0%), with an age at diagnosis ranging from 0 to 66 years. BAV was detected before the age of five years in only 10 cases.

For the remaining patients, the median age of detection of BAV evaluation was 20 years [25 th -75 th percentiles: 15-30]. Survival without the detection of BAV after 20 years of follow-up was 83.2% [IC95%: 77.6 -88.9] (Fig. 2a).

Aortic coarctation was detected in 16 cases (6.9%), at a mean age of 9.5  16 years (range: 0-60) and abnormalities were detected before the age of five years in 60% of cases. Other malformations were minor, and included pulmonary (n=17) and tricuspid (n=45) insufficiencies, trivial mitral insufficiency (n=32), aortic insufficiency (grade I-II)(n=29), persistent left superior vena cava (n=6), dysmorphic pulmonary veins (n=2) and transverse aortic arch elongation (n=1).

At least one aortic diameter exceeded 32 mm in 28 cases (12%) and 35 mm in 10 cases (4.3%). The median age at detection of aortic dilatation/BSA in our cohort was 19 [25 th -75 th percentiles: 7-30] (range: 0-67). Survival without the detection of AD after 20 years of follow-up was 77.8% [IC95%: 72.6 -83.5] (Fig. 2b). When aortic diameter was indexed to BSA, 91 (39%) patients from the cohort had at least one aortic diameter greater than 20 mm/m 2 (Fig. 3).

In total, 27 of the 49 cases of BAV presented an aortic diameter greater than 20 mm/m 2 . AD was therefore more frequent in patients with BAV than in patients without BAV (62.5% versus 31.3%, p<0.001). A 45,X karyotype was present in 63% of the patients with BAV and only 37% of those without BAV (p=0.007). Furthermore, a 45,X karyotype was present in 57.9% of patients with AD and 42% of patients without AD (p < 9.10 -3 ).

DISCUSSION

Our observational study included 336 patients with Turner syndrome. It constitutes a large population, including infants, children, adolescents and adult patients, followed over a long period of time. Most studies on cardiovascular status in patients with Turner syndrome have included less patients [START_REF] Sachdev | Aortic valve disease in Turner syndrome[END_REF](14)(15). The largest study by Mazzanti et al. included 594 patients with Turner syndrome, aged 1 month to 24 years. In our study, we included patients up to the age of 60 [START_REF] Mazzanti | Congenital heart disease in patients with Turner's syndrome. Italian Study Group for Turner Syndrome (ISGTS)[END_REF]. In our population, the mean age at diagnosis of Turner syndrome was 8.0 ± 7.9 years. The statistically significant difference between ages of diagnosis in pediatric and adults units (p=0.003) suggests that the age at diagnosis is improving over time, in France.

In other cohorts from US or Sweden, the age at diagnosis reached 14.5 (1) or 15.1 years (95% CI, 14.5-15.8) [START_REF] Stochholm | Prevalence, incidence, diagnostic delay, and mortality in Turner syndrome[END_REF].

A weakness of our study is the fact that it could be considered as an audit rather than research. However, we considered that it was very important to evaluate current care of patients with Turner syndrome. So far, few prospective studies on cardiac outcomes have been reported in patients with Turner syndrome. Among 17 frequently detected much later in life, at a mean age of 19.1  14.8 years (range: 0-66). Echocardiography was carried out by trained physicians at our reference center, but this examination can be difficult to carry out in infants and children and may fail to detect bicuspid valves due to pectus excavatum or limited acoustic windows.

In our cohort, aortic coarctation was detected much sooner than BAV, at a mean age of 9.1 years, with 10 cases identified before the age of five years.

Our present findings could be influenced by the well known general delay in diagnostic of Turner syndrome [START_REF] Stochholm | Prevalence, incidence, diagnostic delay, and mortality in Turner syndrome[END_REF]. Indeed, delayed cardio-vascular screening may be due to late diagnostic of Turner syndrome. This seems to be true for BAV, as the mean delay between Turner syndrome diagnosis and BAV diagnosis, in our cohort is 13 years, ranging from 0 to 30. However, it does not seem do be the case for aortic coarctation, as the mean delay is rather short (3.94 years). Data on BAV could justify that specifically cardiac diagnostic is poor, even in presence of a timely cardiac assessment for all patients in a reference center.

In our population, we found that 39% of the patients with BSA indexation had a dilated aorta (Fig. 3). This percentage is high, even though our observational study might have underestimated the amount of dilated aorta. The relationship between AD and BAV was significant (p<0.001), as suggested in previous studies [START_REF] Sachdev | Aortic valve disease in Turner syndrome[END_REF][START_REF] Hjerrild | Thoracic aortopathy in Turner syndrome and the influence of bicuspid aortic valves and blood pressure: a CMR study[END_REF]. All of our patients with an aortic diameter greater than 32 mm are alive and none underwent cardiac surgery between 2009 and 2011.

We confirmed statistically significant relationships between the 45,X karyotype and BAV and between 45,X and AD. We therefore conclude that patients with X monosomy should have more frequent cardiovascular follow-up. The natural history of AD remains unknown in patients with Turner syndrome. An intrinsic abnormality of elasticity in the ascending aorta has been suggested [START_REF] Sharma | Aortic distensibility and dilation in Turner's syndrome[END_REF], involving disruption of the transforming growth factor beta pathway [START_REF] Bondy | Aortic dissection in Turner syndrome[END_REF][START_REF] Habashi | Losartan, an AT1 antagonist, prevents aortic aneurysm in a mouse model of Marfan syndrome[END_REF].

In summary, our study sheds further light on the cardiovascular features in Turner syndrome. It emphasizes the fact that cardiovascular monitoring for Turner syndrome patients is currently insufficient in France. Furthermore, although BAV is present at birth, it often remains undiagnosed until later in life. Therefore, careful cardiovascular monitoring is thus required, and a more systematic approach should be taken.

TABLE & FIGURES LEGENDS Table 1: Population studied

Values are expressed as means  SD. All patients under the age of 18 years were considered to be children. BSA: Body surface area (10) -BMI: Body mass index. We used the Kaplan-Meier method to generate survival curves from birth and to determine survival rates without detection of an event and their 95% confidence intervals (Greenwood variance). The events studied were detection of BAV and aortic dilatation. The dotted line represents the limit of 32 mm (left) and 20 mm/m² (right). Figure 3a shows crude diameters, figure 3b shows indexed levels (mm/m 2 ).

  French Ministry of Health set up the Reference Center for Rare Growth Disorders (CRMERC) in Paris, in 2006. This network of specialized tertiary centers was established to improve and harmonize the care of patients with these rare diseases during the transition from adolescence to adulthood. It brings together the endocrine units of the Saint-Antoine, Robert-Debré, Pitié-Salpétrière, Armand Trousseau, Necker and Bicêtre Hospitals. Patients' files are recorded in a web database called CEMARA. This database has been declared to the French data protection agency (CNIL). We studied the cohort of patients followed up at this center between January 1999 and April 2009. This cohort includes patients with Turner syndrome diagnosed between 1954 and 2008. The study was approved by the institutional review board of our faculty.

Figure 1 :Figure 2 :

 12 Figure 1: Flow chart of the cohort

  Fig 2a: Diseasefree survival without bicuspid aortic valves (BAV). Fig 2b: Disease-free survival without aortic dilatation.

Figure 3 :

 3 Figure 3: Aortic diameters at the level of Valsalva sinus, according to age at most

FIGURE 3

 3 FIGURE 3

This research did not receive any specific grant from any funding agency in the public, commercial or not-forprofit sector.

patients, MRI detected aortic anomalies in seven patients, and the time to lesion detection was between two and six years in adults with TS [START_REF] Ilyas | Evaluation by magnetic resonance imaging of aortic dilatation and coarctation in adult Turner syndrome patients[END_REF]. In an Italian cohort of 80 adults, aortic diameter increased with growth rates of 0.1-0.4 mm/year [START_REF] Lanzarini | Prospective evaluation of aortic dimensions in Turner syndrome: a 2-dimensional echocardiographic study[END_REF].

Informative cardiovascular data were available for only 233 of our cases. Despite the follow-up of these patients at a reference center, cardiovascular evaluation was not carried out in 5.9% of cases. Furthermore, reports of evaluations summarized the findings as "normal" rather than giving precise aortic diameters in millimeters, for 18% of the patients. Our study highlights the need for improvement, with cardiovascular monitoring being made systematic in patients with Turner syndrome.

Interestingly, a recent study showed that only 3.5% of patients with Turner syndrome are being followed in accordance with NIH recommendations [START_REF] Devernay | Determinants of medical care for young women with Turner syndrome[END_REF]. The authors of this previous study highlighted the contribution of the loss of patients during the transition from pediatric to adult care. However, missing cardiovascular data rates in our cohort did not differ between children and adults. Freriks et al. have recently reported in 150 patients that standardized multidisciplinary evaluation yields significant previously undiagnosed morbidity in adult women with Turner syndrome [START_REF] Freriks | Standardized multidisciplinary evaluation yields significant previously undiagnosed morbidity in adult women with Turner syndrome[END_REF]. Since the results of our study, we have set up cardiovascular forms to be filled by physicians. We are planning to evaluate in 2 years from now the efficacy of this procedure on improving cardiovascular follow-up. As our study is only an assessment of referred patients, our findings cannot be extrapolated to the entire population of patients with Turner syndrome. Among our informative patients, no cardiovascular abnormalities were observed in 56.2% of the cohort. This percentage is not very different from previous studies [START_REF] Matura | Aortic dilatation and dissection in Turner syndrome[END_REF]14,[START_REF] Elsheikh | Hypertension is a major risk factor for aortic root dilatation in women with Turner's syndrome[END_REF].

Bicuspid aortic valves were detected in 21% of cases. The prevalence of this abnormality was identical in children and adults. Bondy et al. previously reported a prevalence of BAV, as assessed by highly focused echography and MRI, of 30% [START_REF] Sachdev | Aortic valve disease in Turner syndrome[END_REF]. Interestingly, although BAV are present at birth, we found that they were