Insulin-like growth factor-1 inhibits apoptosis of rat gastric smooth muscle cells under high glucose condition via adenosine monophosphate-activated protein kinase (AMPK) pathway.

INTRODUCTION: Diabetic gastroparesis (DGP) is a common chronic complication of diabetes characterized by decreased gastric motility, and an effective number of gastric smooth muscle cells (GSMCs) ensures gastric motility. A previous study documented that apoptosis was present in gastric smooth muscles in rats with DGP and adenosine monophosphate-activated protein kinase (AMPK) was an important factor of apoptosis of rat GSMCs cultured under high glucose conditions. This study aimed to explore the effect of insulin-like growth factor-1 (IGF-1) on apoptosis of high glucose cultured rat GSMCs after silencing of AMPK and elucidate the underlying mechanism. MATERIAL AND METHODS: A total of 120 rats were divided into normal control (NC, n = 20), diabetic gastroparesis (DGP, n = 50) and DGP + IGF-1 (n = 50) groups. After establishing the rat model of DGP, rats in the DGP+IGF-1 group received an intraperitoneal injection of IGF-1 at a dose of 1.5 µg/kg/d for 10 weeks. The level of AMPK activity, liver kinase B1 (LKB1) activity, and calcium/calmodulin-dependent protein kinase b (CaMKKb) expression in rat gastric smooth muscle tissues was detected by Western blot analysis. Apoptosis in rat gastric smooth muscle tissues was detected by TUNEL assay. We also cultured rat GSMCs in vitro under high glucose (HG) condition (35 mM), incubated cells with IGF-1, and silenced AMPK with siRNA. The cells were divided into HG, HG + IGF-1, HG + siRNA, and HG + siRNA + IGF-1 groups. The apoptosis rates of rat GSMCs after silencing AMPK were detected by TUNEL assay and flow cytometry, and apoptosis-related protein expression in rat GSMCs was detected by Western blot. RESULTS: IGF-1 decreased LKB1 activity, CaMKKb expression, AMPK activity, and inhibited apoptosis in rat gastric smooth muscle tissues. Compared with rat GSMCs cultured in vitro under HG conditions, apoptosis rates were reduced after treatment with IGF-1 and AMPK silencing (both p < 0.01). Apoptosis rates were higher in the HG + siRNA group compared with the HG + IGF-1 group (p < 0.05). IGF-1 down-regulated the expression of calcium/calmodulin-dependent kinase II (CaMKII) and p53, up-regulated the expression of p21, PLC-b3, PI3K p110 Ser1070, and the activities of Akt, p70S6K, mTORC1, and mTORC2. IGF-1 also up-regulated Bcl-2 expression and down-regulated the expression of BAX and Caspase-3. CONCLUSIONS: IGF-1 can inhibit the apoptosis of rat GSMCs under high glucose conditions, its mechanism may be related to the regulation of expression and activity of p53, PI3K, TSC-2, Akt, mTOR, 4E-BP1, p70S6K, p21, CaMKII, and PLC-b3 in rat GSMCs acting through AMPK pathway.

Analysis of Factors Related to Distal Proximal Caries on the Distal Surface of the Mandibular Second Molar Induced by an Impacted Mandibular Third Molar.

OBJECTIVE: To analyze the factors related to distal proximal caries of the mandibular second molar (MSM) induced by an impacted mandibular third molar (MTM). METHODS: A total 500 panoramic radiographs of patients with impacted MTMs who were treated in the Department of Stomatology of the Affiliated Hospital of Yanbian University between October 2017 and October 2019 were selected. Descriptive and bivariate analyses were conducted, and the diagnosis of caries in the MSM and the position of the MTM were evaluated. RESULTS: The posterior margin space of the MSM was larger in males (13.5 mm) than females (11.1 mm, P < 0.001) and correlated with the MTM's impacted depth and eruption degree (P < 0.001). The prevalence of distal proximal caries of the MSM was 37.6%, and a χ 2 test showed that age, impacted depth, impacted direction, impacted angle, degree of occurrence, and CEJ distance were correlated with caries in the distal adjacent surface of the MSM (P < 0.001). Logistic multivariate analysis showed that the impacted depth was at position A or B, the impacted direction was mesioangular, and the impacted angle was <80°. Distal adjacent caries of MSMs were prone to occur when entirely or partially emergent. CEJ distance was not an independent factor for caries. The severity of caries in an MSM's distal adjacent surface had statistical significance on the impacted depth and impacted angle (P < 0.001). CONCLUSION: The posterior margin space of the MSM influences the impacted condition of the MTM. Preventive extraction can be considered if the impacted angle is <80°, especially in the case of a fully or partially emergent MTM at position A or B and mesioangular impacted areas.

Mechanism of AMPK-mediated apoptosis of rat gastric smooth muscle cells under high glucose condition.

To observe changes in AMP-activated protein kinase (AMPK) activity and phosphorylation changes in AMPK signaling pathway in gastric smooth muscle cells of rats with diabetic gastroparesis (DGP), investigate the effect of AMPK on apoptosis and explore the underlying mechanism. After establishing rat model of DGP, rats were divided into normal control (NC) and DGP groups. The phosphorylation changes in AMPK pathway were detected by AMPK Signaling Phospho-Antibody Array, and the apoptosis-related proteins were determined. Rat gastric smooth muscle cells were cultured in vitro under different glucose conditions, and divided into normal and high glucose groups. The AMPK activity and intracellular Ca2+ changes in cells were observed. After AMPK silencing, cells were divided into high glucose-24h, high glucose-48h and high glucose-48h+siRNA groups. Changes in expression of apoptosis-related proteins were observed. AMPK activity and apoptosis rates were both increased in gastric smooth muscle tissues in DGP rats (P<0.05, P<0.001, respectively). A total of 14 apoptosis-related differentially phosphorylated proteins were identified. Under high-glucose condition, AMPK activity and intracellular Ca2+ concentrations in rat gastric smooth muscle cells were increased (both P<0.05). After AMPK silencing, p53 expression was decreased, Akt and p70 S6 ribosomal protein kinase (p70S6K) activities were were increased, Bcl-2 expression was increased, CaMKII activity was decreased in the high glucose-48h group. Under high-glucose condition, activated AMPK can directly or indirectly promote cells apoptosis by regulating the expression and activity of p53, Akt, p70S6K, Protein kinase A (PKA), Phospholipidol C (PLC)-ß3, CaMKII, CaMKIV and eukaryotic translation initiation factor 4E binding protein1 (4E-BP1) in rat gastric smooth muscle cells.