Photo Augusto Domínguez Barros
ORAL DRYNESS IN SJÖGREN’S SYNDROME PATIENTS: NOT JUST A QUESTION OF WATER
María Julieta González Burgos
Simposio de Autoinmunidad en Reumatología Septiembre 9 , 2016, Bogotá, Colombia
The standard aetiopathogenic model employed to describe SS disease suggests that alterations in the immune system trigger a pathological response, which eventually destroys the exocrine glands. Apoptotic model
Some antecedents related with dryness mouth
Dawson et al proposed an alternative model of glandular dysfunction, referred to as the non-apoptotic model, in which mechanisms reducing or inhibiting water transport are thought to cause reduced salivary flow.
Perez et al. (2009) performed gene expression profiling of fractions enriched in epithelial cells from LSGs SS-patients and controls using cDNA microarrays. 528 genes were differentially expressed in SS-patients relative to controls. Among the up-regulated genes, 28 were pro-apoptotic and 15 were anti-apoptotic . These findings suggest that the balance between death and survival signals is altered on a large scale in LSG from SS-patients.
The severity of secretory dysfunction does not correlate with the degree of lymphocytic infiltration and glandular destruction in SS-patients (Fox RI).
In salivary glands of SS-patients, altered cholinergic signaling, calcium tunneling, among other factors, have been detected that may result in reduced water release. However, mouth dryness correlates only poorly with the USF.
Non-apoptotic model of glandular hypofunction
1. Cytokines induce inhibition of neurotransmitters
2. Degradation of acetylcholine by increase of cholinesterase
3. Anti-M3 antibodies block Muscarinic receptors
4. Impaired production of ON
5. Disturbance in the release of calcium induced calcium(CICR) by changes of cADPr
8. Altered expression and distribution of AQP5
Adapted of L. Dawson y cols., Rheumatology 2006.45:792–798
Autoimmunity Diseases: High Complexity and Difficult to Understand
Intercambiador Na+/H+
Intercambiador Cl-/HCO3-
Canal K+
Canal Cl-
Na+/K+ ATPasa
Intercambiador K+/H+
Na+/K+/2Cl-
AQP5
Canal de Na+
Receptor muscarínico
AcetilcolinaReceptores α- y β- adrenérgicosAdrenalina o noradrenalina
α β
NAACh
Dibujado por María José Barrera, PhD en Ciencias Biomédicas, 2015
Exocrine Epithelium Complexity Water and electrolytes
ACINUSIsotonic Saliva
DUCTHypotonic Saliva
IONIC COMPONENTSOF SALIVA
-140 mEq/L NaCl
25 mEq/L NaCl
Increased value(1.5 – 4) in SS patients
H2O
H2O
H2O
Components and functions of Saliva
Adapted of A. van Nieuw Amerongen y cols., Caries Res 2004.38:247–253
99% waterelectrolytes
(Na+, K+, Cl-, HCO3-)
proteins (immunoglobulins,
antibacterial products and antifungals,
enzymes)Glycoproteins/mucins
Non-stimulated salivary flow normal ≥
1,5ml/15minutes
SSTPGTAHTLTVLTTTATTPTATGSTATP
TTAAPPTPSATTPAPPSSSAPPE
GSTAPPAHGVTSAPDTRPAP
MUC1/TM
MUC5B
MUC7
GSTAPPAHGVTSAPDTRPAP
MUC1/SEC
Dominio citoplasmático
SEA
Secuencia de transmembrana
Secuencia VSIGLSFPMLP del C-terminal de MUC1/SEC
Sitio de clivaje
EGF
Dominio rico en Cisteína
Dominio D (D4)
Sitio de clivaje autocatalíticoSecuencia única
Secuencia señal
Dominio D (D1-D3)
Dominio rico en Cisteína
Repeticiones en tandem
Modificada de M. Hollingsworth y B. Swanson, Nat Rev Cancer, 2004. 4(1): p. 45-60
Características estructurales de las mucinas salivales
90-300 kDa
200-250 kDa
1 MDa
220 kDa
H2O
H2O
H2OH2O
H2OH2O
H2OH2O
H2OH2O
H2O
H2OH2O
H2O H2O
Mucinas poliméricas secretadasMucinas de membrana
C
N
Y
Y
YY Y Y YYY YN CC N
GalNAcGlcNAcGal
Ácido siálicoFucosaÉster sulfatoSO3
_
Y N-glicosilaciónDominio VNTRDominio citoplasmático Dominio Transmembrana
SO3
_SO3
_SO3
_
SO3
_SO3
_
SO3
_ SO3
_
SO3
_SO3
_ SO3
_
SO3
_
Mucinas salivales
Efecto higroscópico
Mucinas asociadas a membrana
Secuestro de factores solubles
Poder estequiométrico
Transducción de señales
Efecto de filtraciónEfecto de intercambio
iónico
Mucinas solubles
Célula
Modificada de M. Hollingsworth y B. Swanson, Nat Rev Cancer, 2004. 4(1): p. 45-60
Biosíntesis de mucinas
Modificada de D. Thornton y cols. Annu Rev Physiol, 2008. 70: p. 459-86.
RER
Dominio N-terminal (no plegado)Dominio N-terminal (plegado)Dominio C-terminal (no plegado)Dominio C-terminal (plegado)Puentes disúlfuro intermolecularesN-acetilgalactosaminaN-acetilglucosaminaGalactosaFucosaÁcido siálicoDominio de mucina glicosilado
Trans-Golgi
Cis-MedialGolgi
Altered secretion of salivary mucins in Sjögren's syndrome patients
Controls Patients
C1 C2 C3 C4 C5 P1 P2 P3 P4 P5 P6
193 kd
b-actin
stacking gelMUC5B
A B
p = 0.33
Rela
tive
leve
ls of
pro
tein
M
UC5
B/b-
actin
0
5
10
15
20
Controls Patients
Expression of MUC5B in LSG
A
E
B
m sm
s
C
d
ms
D
f
ms
f
Ann Rheum Dis 2008;67:1480–1487
Localization of MUC5B
MUC5B
Sulfo-Lewisa:SO3Galβ1-3[Fuc1-4]GlcNAc
SO3SO
3SO3 SO
3
Levels of Sulfo Lewisa -MUC5B
p < 0.001
Controls Patients
0
5
10
15
20
Rela
tive
leve
ls of
Sul
fo-M
UC
5B
BA
Controls Patients
193 kd
b-actin
Stacking gel
C1 C2 C3 C4 C5 P1 P2 P3 P4 P5 P6
SulfoMUC5B
Localization of Sulfo-Lewisa -MUC5B
m
A
m s
m
C
m
mf
B
a
B
a
C
Abso
rban
ce (A
U)
SS PatientsControl0.0
0.1
0.2
0.3 A p=0.04
Microdensitometric analysis of total sulfate groups and correlation with the basal lamina organization
Ann Rheum Dis 2008;67:1480–1487
Comparison between unstimulated salivary flow and the percentage of mucous acini Sulfo-Lewisa antigen from SS patients and controls
Ann Rheum Dis 2008;67:1480–1487
Disorganization of the basal lamina observed in patients with Sjögren syndrome may lead to dedifferentiation of acinar mucous cells and, as a consequence, alter sulfation of MUC5B. These changes are suggested to represent a novel mechanism that may explain xerostomia in these patients.
S
O
-O
O
ppGalNAcT
β1-3
Core 1
Esqueleto proteico
C1GalT
β1-3
2-3 β1-3
ST3GalTGal3ST
Serina o treonina del esqueleto proteicoGalNAc Gal Neu5Ac
Ser/Thr
Ser/Thr
Ser/Thr
Ser/Thr
Ser/Thr
Ser/Thr
C2GnT
β1-3
Ser/Thr
GlcNAcS
O
-O
OEster sulfato
β1-3
Ser/Thr
β3/4GalT
Core 2
C3GnT
β1-3
Ser/Thr Core 3
β1-3
Ser/Thr Core 4
C2GnT2
β1-3
Ser/Thr
β3/4GalT
β1-6 β1-6
β1-6 β1-6
β1-3/4 β1-3/4
Biosíntesis de oligosacáridos en GSL de pacientes SS
I. Castro y cols., Rheumatology. 2012. 51:482-490
p=0,92 p=0,085
Controles Pacientes SS0.0
0.5
1.0
1.5
2.0
Nive
les re
lativo
sGa
l3ST-
2/b-
actin
a
Controles Pacientes SS0
1
2
3
4
5
Nive
les re
lativo
sGa
l3ST-
4/b-
actin
a
I. Castro y cols., Rheumatology. 2012. 51:482-490Controls SS-Patients
0
5
10
15*
pmol
/h/m
gGa
l3ST
acti
vity
DrynessUSF TNF- Focus
Scoreeye mouth
Gal3ST activity -0.8660* -0.4880* 0.2205 -0.7204* -0.8271*
Correlation coefficients between Gal3ST activity and clinical parameters of SS-patients
Correlación actividad Gal3-O-sulfotransferasa y niveles de TNF-
The Gal3ST activity levels showed an inverse and significant correlation with the symptoms of oral and ocular dryness, with glandular levels of TNF-α and the degree of lymphocyte infiltration in the GS from SS patients.
These results suggest that in GS of SS patients there is an alteration in some of the machinery components involved in the sulfation of MUC5B and this alteration is correlated with the degree of inflammation presented glands.
Citoquinas pro-inflamatorias
CONTROL PACIENTES SS
?
Estos antecedentes sugieren que en GS de pacientes SS, el ambiente inflamatorio podría afectar la localización y sulfatación de mucinas.
H2O H2O
S/T S/T S/T S/T
SO3- SO3
- SO3-
SO3-
H2OH2O
S/T S/T S/T S/T
F. Wang y cols., International Immunopharmacology. 2015. 28:764–772
T. Kondo y cols., Modern Pathology. 2005. 18: 1199–1210
Células HeLa
Glándula endocervical normal teñida con Alcian Blue pH 1.0
Células HeLa estimuladas con TNF-
DMSO
TNF-
Detección de MUC5B y Sulfo-Lewisa en células HeLa estimuladas con TNF- o IFN- durante 48 horas.
0 1 10 1 10 ng/ml, 48hTNF-
SulfoLewisa
b -actina
kDa
IFN-
MUC5B
250 >
50 >
250 >
,
,
,
,
,
,
,
,
,
,
Estos resultados demuestran que TNF- e IFN- reducen la presencia de oligosacáridos sulfatados asociados a MUC5B y sugieren que en GSL de pacientes SS, los altos niveles de citoquinas pro-inflamatorias podrían explicar la hiposulfatación de esta mucina.
Citoquinas pro-inflamatoriasCONTROL PACIENTES SS
H2O H2O
S/T S/T S/T S/T
SO3- SO3
- SO3-
SO3-
H2OH2O
S/T S/T S/T S/T
Gracias
Colaborators
NationalSergio AguileraClaudio MolinaSergio GonzálezCecilia AlliendeMarcela HermosoUlises UrzúaAndrew QuestLisette Leyton Cecilia Leyton
InternacionalJuan Manuel Anaya Enno VeermanUlla MandelBruce BaumHenrik ClausenInka Brockhausen
FONDECYT Chile
Our StudentsPaola PérezEduardo GoicovichYoon-Jeoung KwonPatricia EwertMarianela SánchezMaría José BarreraRodrigo PintoMónica BritoJuan CortésIsabel CastroVerónica BahamondesHsiao Hsin SungCamilo TapiaDenisse SepúlvedaHery UrraJosé VelozoSebastián IndoKatherine BravoCamilo TapiaSebastián IndoSebastián PugaCarolina LagosPatricia CarvajalNicolás Albornoz
A heartfelt gratitude to our Patients
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