Miriplatin-loaded liposome, as a novel mitophagy inducer, suppresses pancreatic cancer proliferation through blocking POLG and TFAM-mediated mtDNA replication

Pancreatic cancer is a more aggressive and refractory malignancy. Resistance and toxicity limit drug efficacy. Herein, we report a lower toxic and higher effective miriplatin (MPt)-loaded liposome, LMPt, exhibiting totally different anti-cancer mechanism from previously reported platinum agents. Both in gemcitabine (GEM)-resistant/sensitive (GEM-R/S) pancreatic cancer cells, LMPt exhibits prominent anti-cancer activity, led by faster cellular entry-induced larger accumulation of MPt. The level of caveolin-1 (Cav-1) determines entry rate and switch of entry pathways of LMPt, indicating a novel role of Cav-1 in nanoparticle entry. After endosome-lysosome processing, in unchanged metabolite, MPt is released and targets mitochondria to enhance binding of mitochondria protease LONP1 with POLG and TFAM, to degrade POLG and TFAM. Then, via PINK1-Parkin axis, mitophagy is induced by POLG and TFAM degradation-initiated mitochondrial DNA (mtDNA) replication blocking. Additionally, POLG and TFAM are identified as novel prognostic markers of pancreatic cancer, and mtDNA replication-induced mitophagy blocking mediates their pro-cancer activity. Our findings reveal that the target of this liposomal platinum agent is mitochondria but not DNA (target of most platinum agents), and totally distinct mechanism of MPt and other formulations of MPt. Self-assembly offers LMPt special efficacy and mechanisms. Prominent action and characteristic mechanism make LMPt a promising cancer candidate.

Relevance between the occurrence of chronic obstructive pulmonary disease and mitochondria DNA haplogroups / 基础医学与临床

Objective To identify the relation between peripheral leukocytes mitochondria DNA(mtDNA) haplo-group and chromic obstructive pulmonary disease(COPD).Methods Sixty COPD patients and 45 healthy con-trols were recruited from Taihe Hospital .mtDNA hypervariable ( HVS ) region and 10171-10659 region were cloned and sequenced.Mutation sites were analyzed via aligning the target gene sequence with the rCRS ( revised Cambridge Reference Sequence) of mtDNA.Samples were divided into possible haplogroup according to mtDNA haplogroup classification tree of Han population.Results Both COPD group and control group were divided into 9 kinds of haplogroup, such as A, F and M7.A haplogroup has a higher proportion in COPD group than control group ( P<0.05) , while F haplogroup has a higher proportion in the control group than COPD group ( P<0.05) . Conclusions F haplogroup may reduce the COPD risk, while A haplogroup may be risk factor for COPD.The occurrence and development of COPD is potentially related to mtDNA haplogroup mutation .

Novel Mitochondrial DNA Variations Associated with Coronary Artery Disease in Type 2 Diabetes from an Asian Indian Population.

Background: Mitochondrial dysfunction leading to insulin resistance may contribute to metabolic and cardiovascular abnormalities and subsequent increase in coronary artery disease. Since mitochondria are involved in generation of ROS, we aimed to investigate the association of mtDNA mutations with T2DM and CAD in our population. Methods: We analyzed the complete mtDNA of South Indian subjects which included patients with angiographically documented CAD [n = 120], subjects with Type 2 Diabetes Mellitus and CAD [n = 150] and healthy control subjects without clinical manifestations of atherosclerotic disease and Type 2 Diabetes [n = 100]. We detected the association of common variants of the mitochondrial genes with both T2DM and CAD, which raises the possibility of a shared mitochondrial genetic background of these metabolic disorders in our population. Results: The complete mitochondrial analysis of the control group revealed several sequence variations but did not show any novel mutations. Mitochondrial analysis of individuals with CAD and T2DM revealed a total of 36 novel variations. Mutations were more prevalent in NADH Dehydrogenase [ND] genes that encode mitochondrial enzyme Complex I. Among the 20 novel mutations in the ND genes, 17 were missense, 2 synonymous and 1 frame shift variant were observed. In Cytochrome b [Cytb] gene, 7 variations observed were novel that included 5 missense mutations in cytochrome c oxidase [CO2] were novel mutations including 1 missense mutation and 1 synonymous mutation. In rRNA genes, we identified 1 novel variant in 12s RNA and 3 in 16s rRNA. Among the CAD patient group without T2DM, 3 novel variants in ND region were identified of which 2 were synonymous and one was missense. The variants observed are not reported to have any disease association so far by any studies. Conclusions: Presence of pathogenic known and novel mutations suggests mtDNA variations have a role in the pathophysiology of CAD associated with T2DM in our population.

Rapid and Simple Screening of Mitochondrial DNA in Koreans by the Analysis of Highly Variable Control Region SNPs / 대한법의학회지

Human mitochondrial DNA (mtDNA) is generally used to identify highly degraded forensic samples, particularly when the extracted DNA is not sufficient for nuclear DNA analysis. However, direct sequencing, the most widely used mtDNA analysis method, is laborious and time-consuming, and precludes the simultaneous analysis of many samples. Here, we describe a rapid and simple screening method for mtDNA analysis in Koreans using single base extension (SBE) methods. Sixteen highly polymorphic mtDNA SNPs from the control region were selected, and a multiplex SBE system was constructed to analyze them. Because the developed system consists of two duplex PCRs, which produce small amplicons with fewer than 270 bp, it works well with highly degraded samples such as old skeletal remains. Using this multiplex SBE system, 145 different haplotypes were expected to be observed from 593 unrelated Koreans. Seventy-three haplotypes were expected to be observed only once, and the most frequent haplotype was expected to occur 80 times. Since the mean number of pairwise differences was estimated to be 4.55, the developed system could be useful to exclude samples that do not match evidence and reference samples. Therefore, the multiplex SBE system used in this study will be a useful tool to analyze many samples simultaneously and to efficiently screen out non-matching mtDNA sequences in forensic casework.

Establishment of a molecular tool for blood meal identification in Malaysia

Objective: To establish a polymerase chain reaction (PCR) technique based on cytochrome b (cytb) gene of mitochondria DNA (mtDNA) for blood meal identification. Methods: The PCR technique was established based on published information and validated using blood sample of laboratory animals of which their whole gene sequences are available in GenBank. PCR was next performed to compile gene sequences of different species of wild rodents. The primers used were complementary to the conserved region of the cytb gene of vertebrate’s mtDNA. A total of 100 blood samples, both from laboratory animals and wild rodents were collected and analyzed. The obtained unknown sequences were compared with those in the GenBank database using BLAST program to identify the vertebrate animal species. Results: Gene sequences of 11 species of wild animals caught in 9 localities of Peninsular Malaysia were compiled using the established PCR. The animals involved were Rattus (rattus) tanezumi, Rattus tiomanicus, Leopoldamys sabanus,Tupaia glis, Tupaia minor, Niviventor cremoriventor, Rhinosciurus laticaudatus, Callosciurus caniseps, Sundamys muelleri, Rattus rajah and Maxomys whiteheadi. The BLAST results confirmed the host with exact or nearly exact matches (>89% identity). Ten new gene sequences have been deposited in GenBank database since September 2010. Conclusions: This study indicates that the PCR direct sequencing system using universal primer sets for vertebrate cytb gene is a promising technique for blood meal identification.

Mutational Analysis of Mitochondria DNA in Children with IgA Nephropathy

PURPOSE: The association of mitochondrial DNA (mtDNA) mutations, deletions and copy number with progressive changes in patients with some glomerular disease and end-stage renal disease have been reported. In this study, we performed mtDNA mutation analysis in children with IgA nephropathy to investigate its role in progressive clinical course. METHODS: Seven children with IgA nephropathy were involved in this study. MtDNA isolated from platelet was amplified by PCR and sequenced entirely. RESULTS: The mean age at renal biopsy was 11.5+/-2.2 year and the mean age at latest evaluation was 17.9+/-3.2 year. The mean follow-up period were 7.8+/-3.1 years. Patients was divided into 2 groups according to the amount of proteinuria at presenting manifestation. Group 2 patients were nephrotic syndrome. Renal function reveals within normal range in all patients. In group 2 patients, the mean serum albumin level was significantly lower than those of group 1 (3.7+/-0.6 g/dL vs. 4.7+/-0.2 g/dL, P=0.0241) and the mean total cholesterol level was significantly higher than those of group 1 (222.7+/-35.7 mg/dL vs. 148.3+/-29.1 mg/dL, P=0.0283). In Group 2 patients, total amount of protein of 24 hour collected urine also significantly higher than those of group 1 (1,466.0+/-742.5 mg vs. 122.5+/-48.1 mg, P=0.0135). Pr/Cr ratio in random urine sample was also higher in group 2 than those of group 1 but the statistical significance was not noted (1.8+/-1.6 vs. 0.2+/-0.2, P=0.0961). Deletion of mtDNA nt 8272-8281 were observed in two patients, one patient in each groups, respectively. This is non-coding lesion. No patients demonstrated the mtDNA mutations. CONCLUSIONS: We have identified a deletion of mtDNA nt 8272-8281 in two children with IgA nephropathy. Further studies are needed to clarify the role of mitochondrial function in the progressive change of IgA nephropathy.