Glucagon-Like Peptide-1 Receptor Agonist Ameliorates 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine (MPTP) Neurotoxicity Through Enhancing Mitophagy Flux and Reducing α-Synuclein and Oxidative Stress.

Parkinson disease (PD) is the second most common neurodegenerative disease without known disease modification therapy to slow down disease progression. This disease has pathological features of Lewy bodies with α-synuclein aggregation being the major component and selective dopaminergic neuronal loss over the substantia nigra. Although the exact etiology is still unknown, mitochondrial dysfunction has been shown to be central in PD pathophysiology. Type 2 diabetes mellitus has recently been connected to PD, and anti-diabetic drugs, such as glucagon-like peptide-1 receptor agonists (GLP-1RAs), have been shown to possess neuroprotective effects in PD animal models. The GLP-1RA liraglutide is currently under a phase 2 clinical trial to measure its effect on motor and non-motor symptoms in PD patients. In this study, we used an acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD to test the possible mechanism of the GLP-1RA liraglutide in the pathogenesis of PD. We show that the neurobehavioral and motor dysfunction caused by the mitochondrial complex I inhibitor, MPTP, can be partially reversed by liraglutide. The GLP-1RA can protect mice from apoptosis of substantia nigra neurons induced by MPTP. MPTP treatment led to imbalanced mitochondrial fusion and fission dynamics, altered mitochondrial morphology, impeded autophagy flux, increased α-synuclein accumulation, and elevated oxidative stress. Specifically, the normalizing of mitochondrial fusion-fission dynamic-related proteins and enhancement of autophagy flux after administration of liraglutide is associated with improving neuronal survival. This suggests that GLP-1RAs may provide potential beneficial effects for PD caused by mitochondrial dysfunction through improvement of mitochondrial morphology balance and enhancing damaged organelle degradation.

Mitochondrial Transfer of Wharton's Jelly Mesenchymal Stem Cells Eliminates Mutation Burden and Rescues Mitochondrial Bioenergetics in Rotenone-Stressed MELAS Fibroblasts.

Wharton's jelly mesenchymal stem cells (WJMSCs) transfer healthy mitochondria to cells harboring a mitochondrial DNA (mtDNA) defect. Mitochondrial myopathy, encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) is one of the major subgroups of mitochondrial diseases, caused by the mt.3243A>G point mutation in the mitochondrial tRNALeu(UUR) gene. The specific aim of the study is to investigate whether WJMSCs exert therapeutic effect for mitochondrial dysfunction in cells of MELAS patient through donating healthy mitochondria. We herein demonstrate that WJMSCs transfer healthy mitochondria into rotenone-stressed fibroblasts of a MELAS patient, thereby eliminating mutation burden and rescuing mitochondrial functions. In the coculture system in vitro study, WJMSCs transferred healthy mitochondria to rotenone-stressed MELAS fibroblasts. By inhibiting actin polymerization to block tunneling nanotubes (TNTs), the WJMSC-conducted mitochondrial transfer was abrogated. After mitochondrial transfer, the mt.3243A>G mutation burden of MELAS fibroblasts was reduced to an undetectable level, with long-term retention. Sequencing results confirmed that the transferred mitochondria were donated from WJMSCs. Furthermore, mitochondrial transfer of WJMSCs to MELAS fibroblasts improves mitochondrial functions and cellular performance, including protein translation of respiratory complexes, ROS overexpression, mitochondrial membrane potential, mitochondrial morphology and bioenergetics, cell proliferation, mitochondrion-dependent viability, and apoptotic resistance. This study demonstrates that WJMSCs exert bioenergetic therapeutic effects through mitochondrial transfer. This finding paves the way for the development of innovative treatments for MELAS and other mitochondrial diseases.

Expression of Toll-like receptor9 in diffuse large B-cell lymphoma: further exploring CpG oligodeoxynucleotide in NFκB pathway.

Human Toll-like receptors (TLRs) that recognize a variety of pathogen-associated molecular patterns are associated with activation and immunogenic response in lymphoid neoplasms, but rarely explored in diffuse large B-cell lymphoma (DLBCL). We conducted this study to evaluate the expression of TLR9 in and potential treatment of DLBCL with TLR9 agonist - CpG oligodeoxynucleotide (ODN). The real-time quantitative reverse transcription-polymerase chain reaction was carried out to detect TLR9 expression in 41 formalin-fixed paraffin-embedded samples. The transformation of immunophenotype and NFκB pathway of DLBCL upon CpG ODN stimulation were investigated by a DLBCL cell line. TLR9 was commonly detected in DLBCL with relative mRNA levels above 1.0 × 10(-2) in 35 of 41 cases (85.36%). It was suspected that a high proportion of DLBCL to be activated by CpG stimulation. In vitro study with a DLBCL cell line revealed an increased CD20, but decreased BCL-6 and MUM1/IRF4 expression after treatment with CpG ODN. The NFκB pathway was initially activated, but finally suppressed upon CpG ODN stimulation. The proliferation of tumor cells was also inhibited by long time incubation. These findings provide new insights into the role of TLR9 in DLBCL and potential implication of TLR9 agonist in the treatment of DLBCL.

Selective activation of Toll-like receptor 7 in activated hepatic stellate cells may modulate their profibrogenic phenotype.

Cholestatic liver injury may activate HSCs (hepatic stellate cells) to a profibrogenic phenotype, contributing to liver fibrogenesis. We have previously demonstrated the involvement of TLR (Toll-like receptor) 7 in the pathogenesis of biliary atresia. In the present study we investigated the ability of TLR7 to modulate the profibrogenic phenotype in HSCs. Obstructive jaundice was associated with significant down-regulation of TLR7. Primary HSCs isolated from BDL (bile duct ligation) rats with obstructive jaundice exhibited reduced expression of TLR7 and increased expression of α-SMA (α-smooth muscle actin) and collagen-α1 compared with sham rats, reflecting HSC-mediated changes. Treatment of primary activated rat HSCs and rat T6 cells with CL075, a TLR7 and TLR8 ligand, significantly decreased expression of MCP-1 (monocyte chemotactic protein-1), TGF-ß1 (transforming growth factor-ß1), collagen-α1 and MMP-2 (matrix metalloproteinase-2), and inhibited cell proliferation and migration. In contrast, silencing TLR7 expression with shRNA (short hairpin RNA) in T6 cells effectively blocked the effects of CL075 stimulation, reversing the changes in MCP-1, TGF-ß1 and collagen-α1 expression and accelerating cell migration. Our results indicate that obstructive jaundice is associated with down-regulation of TLR7 and up-regulation of profibrogenic gene expression in HSCs. Selective activation of TLR7 may modulate the profibrogenic phenotype in activated HSCs associated with cholestatic liver injury.

Effects of hepatocyte CD14 upregulation during cholestasis on endotoxin sensitivity.

Cholestasis is frequently related to endotoxemia and inflammatory response. Our previous investigation revealed a significant increase in plasma endotoxin and CD14 levels during biliary atresia. We therefore propose that lipopolysacharides (LPS) may stimulate CD14 production in liver cells and promote the removal of endotoxins. The aims of this study are to test the hypothesis that CD14 is upregulated by LPS and investigate the pathophysiological role of CD14 production during cholestasis. Using Western blotting, qRT-PCR, and promoter activity assay, we demonstrated that LPS was associated with a significant increase in CD14 and MD2 protein and mRNA expression and CD14 promoter activity in C9 rat hepatocytes but not in the HSC-T6 hepatic stellate cell line in vitro. To correlate CD14 expression and endotoxin sensitivity, in vivo biliary LPS administration was performed on rats two weeks after they were subjected to bile duct ligation (BDL) or a sham operation. CD14 expression and endotoxin levels were found to significantly increase after LPS administration in BDL rats. These returned to basal levels after 24 h. In contrast, although endotoxin levels were increased in sham-operated rats given LPS, no increase in CD14 expression was observed. However, mortality within 24 h was more frequent in the BDL animals than in the sham-operated group. In conclusion, cholestasis and LPS stimulation were here found to upregulate hepatic CD14 expression, which may have led to increased endotoxin sensitivity and host proinflammatory reactions, causing organ failure and death in BDL rats.

No correlation between BRAFV600E mutation and clinicopathological features of papillary thyroid carcinomas in Taiwan.

OBJECTIVE: Genetic alterations in four oncogenes, namely RAS point mutations, RET rearrangements (RET/PTC), NTRK1 rearrangements (TRK) and BRAF point mutations have been identified in human papillary thyroid carcinomas (PTCs). These oncogenes act along the RET/PTC(TRK)-RAS-BRAF-MEK-MAPK kinase pathway, mediating a number of cellular fates including growth, proliferation and survival in thyroid cells. In this study, we analysed mutations of BRAF in a cohort of PTCs. METHODS: To screen for BRAF mutations, the genomic DNA of 105 PTCs were amplified by polymerase chain reaction (PCR) with primers flanking exon 15 and PCR products were directly sequenced with an automatic sequencer. These results, together with data from our previous studies on RAS, RET rearrangements and NTRK1 rearrangements in the same tumours, were compared to determine their individual significance in the pathogenesis of PTCs in Taiwan. RESULTS: BRAF mutations were detected in 49 of 105 (47%) tumour samples. All mutations involved a thymine-to-adenine transversion at nucleotide 1799 and were heterozygous. There was no overlap between papillary carcinomas harbouring RET rearrangements, NTRK1 rearrangements and BRAF mutations. In this cohort, correlation between BRAF mutations and various clinicopathological parameters in 101 papillary carcinomas did not reveal any association with age at diagnosis, sex, tumour size, histological variants of PTC, multicentricity, cervical lymph node metastases, extrathyroidal invasion, distant metastases and clinical stage. CONCLUSIONS: BRAFV600E mutation is the most prevalent oncogene in PTCs in Taiwan. Our data did not suggest that BRAFV600E mutation could be a potentially useful marker of prognosis in patients with papillary carcinomas in the population studied.