Fufang-zhenzhu-tiaozhi formula protects islet against injury and promotes ß cell regeneration in diabetic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Fufang-zhenzhu-tiaozhi formula (FTZ) is a patented preparation of traditional Chinese medicine that has been used to treat hyperglycemia and hyperlipidemia in the clinic for almost 10 years. Our previous study had demonstrated that FTZ can protect islet ß cell injury in vitro. However, the efficacy of FTZ on ß cell regeneration in vivo and the involved anti-diabetic mechanism remains unknown. AIM OF THE STUDY: We aim to investigate the effects of FTZ as a good remedy for islet protection and ß cell regeneration, and to reveal the underlying mechanism. MATERIALS AND METHODS: C57BL/6 mice were fed with high-fat diet for 3 weeks and then intraperitoneally injected with streptozotocin (90 mg/kg/d × 1 d) to establish type 2 diabetes (T2D) models. Mice in each group were divided into three batches that sacrificed after 3, 7 and 28 days of FTZ administration. Body weight, blood glucose, and oral glucose tolerance test were measured at indicated time points. Fasting insulin was determined by enzyme-linked immunosorbent assay (ELISA) kit. Neonatal ß cell was assessed by insulin & PCNA double immunofluorescence staining, and the underlying mechanisms related to ß cell regeneration were further performed by hematoxylin-eosin staining, insulin & glucagon double immunofluorescence staining and Western blot. RESULTS: FTZ and metformin can significantly help with the symptoms of DM, such as alleviating weight loss, reducing blood glucose, improving the level of insulin in vivo, and relieving insulin resistance, suggesting FTZ and metformin treatment maintained the normal morphological function of islet. Notably, ß cell regeneration, which is indicated by insulin and PCNA double-positive cells, was promoted by FTZ, whereas few neonatal ß cells were observed in metformin group. Hematoxylin-eosin staining, and its quantification results showed that FTZ effectively prevented the invasion of inflammatory cells into the islets in diabetic mice. Most ß cells in the islets of diabetic model mice were devoid, and the islets were almost all α cells, while the diabetic mice administered FTZ could still maintain about half of the ß cells in the islet. Furthermore, FTZ upregulated the expression of critical transcription factors during ß cell development and maturation (such as PDX-1, MAFA and NGN3) in diabetic mice. CONCLUSIONS: FTZ can alleviate diabetes symptoms and promote ß cell regeneration in diabetic mice. Moreover, FTZ promotes ß cell regeneration by preserving islet (resisting inflammatory cells invading islets), maintaining the number of ß cells in islets, and increasing the expression of PDX-1, MAFA and NGN3.

Effects of Artemisia annua L. Essential Oil on Osteoclast Differentiation and Function Induced by RANKL.

Objective: This study aimed to assess the main components of Artemisia annua L. essential oil (AEO) and determine their effect on the proliferation and differentiation of RAW264.7 cells induced by receptor activator for nuclear factor-ligand (RANKL) in vitro. Then, we tried to explain part of the function of its possible mechanisms. Materials and Methods: Essential oil was extracted from Artemisia annua L. Osteoclasts were induced in vitro by RANKL in mouse RAW264.7 cells. The experimental group was treated with different concentrations of AEO, while the control group was not treated with AEO. CCK8 was used to detect osteoclast proliferation. The osteoclasts were stained with TRAP. Western blot was used to detect protein in the MAPK pathway and the NF-κB pathway after treatment with different concentrations of AEO. RT-PCR was used to determine the expression of osteoclast-related mRNA in cells. Results: The GC-MS analysis was used to obtain the main components of AEO, including camphor, borneol, camphor, borneol, terpinen-4-ol, p-cymene, eucalyptol, deoxyartemisinin, and artemisia ketone. The CCK8 results showed that the AEO volume ratio of 1 : 4000, 1 : 5000, and 1 : 6000 did not affect the proliferation of RAW264.7 cells. However, TRAP staining showed that AEO decreased osteoclast formation. Western blot results showed that the expression of protein TRAF6, p-p38, p-ERK, p-p65, and NFATc1 decreased in the MAPK pathway and the NF-κB pathway affected by AEO. Furthermore, RT-PCR results showed that the expression of osteoclast resorption-related mRNAs (MMP-9, DC-STAMP, TRAP, and CTSK) and osteoclast differentiation-related mRNAs (OSCAR, NFATc1, c-Src, and c-Fos) also decreased in the experimental group. Conclusions: AEO inhibits osteoclast differentiation in vitro, probably by reducing TRAF6 activation, acting on the MAPK pathway and NF-κB pathway, and inhibiting the expression of osteoclast-related genes.