Objective:

2

To investigate the mechanism of cAMP-PKA signaling pathway mediated by Chinese medicine formula Shaoyao Gancao Decoction (芍药甘草汤

SGD) on the regulation of aquaporin 5 (AQP5) and muscarinic receptor 3 (M3R) levels in Sjögren's syndrome (SS).

Methods:

2

Of the 30 mice

5 were randomly selected as control

and others were used for creating SS model. After successful modeling

mice were randomly divided into 5 groups (

n

=5 per group) and intragastrically administered with saline (8 mL/kg)

pilocarpine (1.4 mg/kg)

or low

medium and high doses SGD (0.14

0.21

0.35 g/kg

Radix paeoniae

with 0.01 g/kg

Radix glycyrrhizae

respectively) for 6 weeks. Human labial gland acinar cells were treated with pilocarpine or varying doses of SGD with saline as the placebo. Hematoxylin and eosin staining was used to observe the histopathological changes of the submandibular glands of mice. The serum levels of anti-SS antigen A (SS-A)

anti-SS antigen B (SS-B)

M3R

and α-fodrin in submandibular glands of mice were measured by enzyme-linked immunosorbent assay. Immunofluorescence staining was used to observe the spatial localization of AQP5 and M3R in acinar cells. Reverse transcriptase polymerase chain reaction and Western blot were used to detect the expressions of PKA

cAMP

Epac1

AQP5

M3R

nuclear factor kappa-B (NF-κB)

and tumor necrosis factor (TNF)-α in submandibular gland tissues and cells of each group.

Results:

2

Compared to normal mice

body weight

5-min salivary secretion

30-min secretion of tears and breakup time of tear film of model mice decreased at 1–6 weeks after immunization (all

P

<

0.05)

whereas water intake increased (all

P

<

0.05). In the model group

glands of the submandibular glands showed atrophy

accompanied by acini of different sizes

decreased numbers and loose arrangement

with catheter dilatation and different degrees of lymphocyte infiltration. Conditions of mice in SGD groups were improved. The positive expression of AQP5 and M3R were higher in the acinar cells treated with all doses SGD compared to the normal group; serum levels of SS-A

SS-B

and α-fodrin were lower

and that of M3R was higher in all doses SGD treated animals than the model or pilocarpine treated ones (all

P

<

0.05). Compared to the model and pilocarpine groups

the mRNA and protein levels of NF-κB and TNF-α were lower in mice or cells treated with medium or high-dose SGD (all

P

<

0.05)

while those of PKA

Epac1

AQP5 and M3R were higher (all

P

<

0.05).

Conclusion:

2

SGD can improve symptoms of SS by regulating the cAMP-PKA signaling pathway and increasing AQP5 and M3R levels.