How do changes in the mtDNA and mitochondrial dysfunction influence cancer and cancer therapy? Challenges, opportunities and models.

Several mutations in nuclear genes encoding for mitochondrial components have been associated with an increased cancer risk or are even causative, e.g. succinate dehydrogenase (SDHB, SDHC and SDHD genes) and iso-citrate dehydrogenase (IDH1 and IDH2 genes). Recently, studies have suggested an eminent role for mitochondrial DNA (mtDNA) mutations in the development of a wide variety of cancers. Various studies associated mtDNA abnormalities, including mutations, deletions, inversions and copy number alterations, with mitochondrial dysfunction. This might, explain the hampered cellular bioenergetics in many cancer cell types. Germline (e.g. m.10398A>G; m.6253T>C) and somatic mtDNA mutations as well as differences in mtDNA copy number seem to be associated with cancer risk. It seems that mtDNA can contribute as driver or as complementary gene mutation according to the multiple-hit model. This can enhance the mutagenic/clonogenic potential of the cell as observed for m.8993T>G or influences the metastatic potential in later stages of cancer progression. Alternatively, other mtDNA variations will be innocent passenger mutations in a tumor and therefore do not contribute to the tumorigenic or metastatic potential. In this review, we discuss how reported mtDNA variations interfere with cancer treatment and what implications this has on current successful pharmaceutical interventions. Mutations in MT-ND4 and mtDNA depletion have been reported to be involved in cisplatin resistance. Pharmaceutical impairment of OXPHOS by metformin can increase the efficiency of radiotherapy. To study mitochondrial dysfunction in cancer, different cellular models (like ρ(0) cells or cybrids), in vivo murine models (xenografts and specific mtDNA mouse models in combination with a spontaneous cancer mouse model) and small animal models (e.g. Danio rerio) could be potentially interesting to use. For future research, we foresee that unraveling mtDNA variations can contribute to personalized therapy for specific cancer types and improve the outcome of the disease.

Patient-derived fibroblasts indicate oxidative stress status and may justify antioxidant therapy in OXPHOS disorders.

Oxidative phosphorylation disorders are often associated with increased oxidative stress and antioxidant therapy is frequently given as treatment. However, the role of oxidative stress in oxidative phosphorylation disorders or patients is far from clear and consequently the preventive or therapeutic effect of antioxidants is highly anecdotic. Therefore, we performed a systematic study of a panel of oxidative stress parameters (reactive oxygen species levels, damage and defense) in fibroblasts of twelve well-characterized oxidative phosphorylation patients with a defect in the POLG1 gene, in the mitochondrial DNA-encoded tRNA-Leu gene (m.3243A>G or m.3302A>G) and in one of the mitochondrial DNA-encoded NADH dehydrogenase complex I (CI) subunits. All except two cell lines (one POLG1 and one tRNA-Leu) showed increased reactive oxygen species levels compared with controls, but only four (two CI and two tRNA-Leu) cell lines provided evidence for increased oxidative protein damage. The absence of a correlation between reactive oxygen species levels and oxidative protein damage implies differences in damage prevention or correction. This was investigated by gene expression studies, which showed adaptive and compensating changes involving antioxidants and the unfolded protein response, especially in the POLG1 group. This study indicated that patients display individual responses and that detailed analysis of fibroblasts enables the identification of patients that potentially benefit from antioxidant therapy. Furthermore, the fibroblast model can also be used to search for and test novel, more specific antioxidants or explore ways to stimulate compensatory mechanisms.

Transcriptional changes in OXPHOS complex I deficiency are related to anti-oxidant pathways and could explain the disturbed calcium homeostasis.

Defective complex I (CI) is the most common type of oxidative phosphorylation disease, with an incidence of 1 in 5000 live births. Here, whole genome expression profiling of fibroblasts from CI deficient patients was performed to gain insight into the cell pathological mechanism. Our results suggest that patient fibroblasts responded to oxidative stress by Nrf2-mediated induction of the glutathione antioxidant system and Gadd45-mediated activation of the DNA damage response pathway. Furthermore, the observed reduced expression of selenoproteins, might explain the disturbed calcium homeostasis previously described for the patient fibroblasts and might be linked to endoplasmic reticulum stress. These results suggest that both glutathione and selenium metabolism are potentially therapeutic targets in CI deficiency.

Large scale mtDNA sequencing reveals sequence and functional conservation as major determinants of homoplasmic mtDNA variant distribution.

The mitochondrial DNA (mtDNA) is highly variable, containing large numbers of pathogenic mutations and neutral polymorphisms. The spectrum of homoplasmic mtDNA variation was characterized in 730 subjects and compared with known pathogenic sites. The frequency and distribution of variants in protein coding genes were inversely correlated with conservation at the amino acid level. Analysis of tRNA secondary structures indicated a preference of variants for the loops and some acceptor stem positions. This comprehensive overview of mtDNA variants distinguishes between regions and positions which are likely not critical, mainly conserved regions with pathogenic mutations and essential regions containing no mutations at all.

Inherited mitochondrial variants are not a major cause of age-related hearing impairment in the European population.

Mitochondrial DNA (mtDNA) mutations have been implicated in various age-related diseases. To further clarify the role of mtDNA variants in age-related hearing impairment (ARHI), we determined the DNA sequence of the entire mitochondrial genome of 400 individuals using the Affymetrix Human Mitochondrial Resequencing Array. These were the 200 worst hearing and the 200 best hearing from a collection of 947 Belgian samples. We performed association tests with individual mitochondrial variants, comparison of the mutation load, and association with European haplogroups and their interaction with environmental risk factors. We also tested the influence of rare variants on ARHI. None of these tests showed any association with ARHI.

Enumeration of probiotic bacilli spores in animal feed: interlaboratory study.

Fourteen out of 17 laboratories completed an interlaboratory study comparing 2 pretreatment protocols of feed samples containing authorized probiotic bacilli spores. Both methods used tryptone soy agar for enumeration. Pretreatment A involved preparation of a suspension of the feed sample in 50% ethanol. For pretreatment B, the sample was suspended in peptone salt solution and heated at 80 degrees C for 10 min. Each laboratory analyzed 12 samples (6 per pretreatment), which represented duplicates of a high (10(9) colony-forming units [CFU]/g) and low (10(5) CFU/g) level of bacilli spores or a blank that contained vegetative probiotic bacteria only. For pretreatment A, the repeatability relative standard deviation (RSD(r)) was 2.9% for the low level and 2.5% for the high. The reproducibility relative standard deviation (RSDR) values were 7.8 and 5.9%, respectively. Pretreatment B revealed RSD(r) values of 1.1 and 1.0%, and RSDR values of 5.8 and 3.4%, respectively. The heat treatment (pretreatment B) of feed samples had better precision data, resulted in higher viable bacilli counts, and was more effective in deactivating vegetative background flora. It is therefore recommended for adoption for official control purposes and for CEN and ISO standards.

Percutaneous removal of a knotted pulmonary artery catheter using a tracheostomy dilator.

STATEMENT OF FINDINGS: This case report describes removal of a knotted, subclavian, pulmonary artery catheter using a tracheostomy dilator. With this simple method an invasive procedure might be averted.

Immunosenescence revisited. Does it have any clinical significance?

Immunosenescence refers to the influence of aging on the immune system. Numerous problems are encountered in studying this topic, the main one being the influence of concomitant disease. Despite the great efforts that have been devoted to research in this field, the results of studies performed to date have not been convincing and, until now, no sound scientific evidence has emerged to show that immunosenescence is clinically significant. The only possible exceptions to this are the discovery of a selective defect in cell-mediated immunity and the reactivation of varicella zoster virus. Therefore, many more, and better designed, studies will have to be conducted before the full clinical impact of immunosenescence can be delineated.

[Special attention intensive care in the Netherlands once again]. / Nogmaals het aandachtsgebied intensive care in Nederland.

The organizational structure of intensive care departments is still under debate. 'Open' (without) or 'closed format' (with intensivists), that is the question. However, various studies establish that the presence of full-time intensivists reduces mortality rates in intensive care units substantially. Currently, only a minority of Dutch hospitals have properly staffed intensive care units, despite clear guidelines. It is estimated that yearly 250 to 600 lives could be saved. While the capacity of training intensivists in the Netherlands has recently been increased enormously, only a faction of young intensivists will under the circumstances be able to find jobs. More jobs should be created, for the sake of patients and young doctors as well.

Synchroneously occurring lung cancer (stages I-II) and coronary artery disease: concomitant versus staged surgical approach.

OBJECTIVE: The assessment of the best surgical approach in patients with synchroneously occurring lung cancer (stages I and II) and coronary artery disease: concomitant or staged. METHODS: A retrospective, observational study was conducted in a tertiary centre for cardiothoracic surgery. From 1988-1995, 34 patients underwent pulmonary resection for stages I-II primary bronchogenic carcinoma and open-heart surgery (almost always coronary-artery bypass grafting), either concomitantly (n = 24) or in a staged procedure (n = 10). Mean interval between operations was 33.9 +/- 34.7 days (range: 12-120 days). Results were statistically computed. RESULTS: Preoperatively both groups were perfectly matched. Follow-up was 100%. Long term survival, median 4.2 years, was comparable in both groups (log-rank test: chi2 0.30; df = 1; P = 0.58), indicating no influence on survival from performing either a concomitant or staged procedure. No relation could be demonstrated between survival and age, histopathology or extent of tumour; nor in the concomitantly operated group between survival and timing of lung resection in relation to extra-corporeal circulation. Overall peri-operative mortality was 6/34, 17.6%, but a large difference was noted between the two groups (5/24, 20.8% vs. 1/10, 10%; P = 0.64), underscoring the greater risk involved in the concomitant procedure, although this difference was not statistically significant because of small numbers. CONCLUSIONS: No difference in survival between the two groups, one operated upon in a staged procedure, the other concomitantly, could be demonstrated. However, the greater perioperative risk makes the concomitant procedure less attractive, and the staged approach the preferred one. Interval between operations can be individualized according to the clinical status of the particular patient to a period as short as 2 weeks.

The influence of open-heart surgery on survival of patients with co-existent surgically amenable lung cancer (stages I and II).

OBJECTIVE: The evaluation of the influence of open-heart surgery on the survival of patients with co-existent surgically amenable lung cancer stages I and II. METHODS: A retrospective, observational study was conducted in a tertiary centre for cardiothoracic surgery. From 1988 to 1995, 121 consecutive patients underwent pulmonary resection for stages I-II primary non-small cell bronchogenic carcinoma. Eighty seven of them had merely a lung carcinoma necessitating resection, 34 had in addition defined coronary-artery disease and consequently were also subjected to open-heart surgery. Results were statistically computed. RESULTS: Follow-up was complete in 117/121 patients, 96.7% (83/87, 95.4% and 34/34, 100% in respective groups). Both groups were matched with regard to preoperative features possibly influencing survival. Median long term survival time was 4.3 years overall, 5.8 years for patients merely undergoing lung resection and 4.2 years for them undergoing open-heart surgery as well; this difference was not statistically significant (log-rank test: chi2 0.92, df= 1, P = 0.34), indicating no or limited influence of open-heart surgery on survival of patients with surgically amenable co-existent lung carcinoma. No relationship was found between survival and age, tumour stage, and histopathology. However, metastatic disease as cause of death was significantly increased in patients undergoing open-heart surgery (5/8 vs. 10/33, P = 0.0898), indicating a possible promotion of metastatic spread of co-existent lung carcinoma by this procedure. Overall perioperative mortality rate was 10/121, 8.3%, for the greater part the result of a relatively high mortality rate in the group of patients undergoing heart as well as lung surgery (6/34, 17.6%), underscoring the great risks involved in these patients, the mortality rate for lung resection alone being comparably low 4/87, 4.6% (P = 0.0191). CONCLUSION: Open-heart surgery for defined coronary-artery disease in patients with surgically amenable lung carcinoma carries a substantially higher perioperative risk, but has no influence on long term results. Metastatic spread is possibly promoted by open-heart surgery. Optimal treatment, consisting of complete revascularization and appropriate lung resection, is therefore sufficiently justified by these results.

Pharmacokinetics of cefradine, sulfamethoxazole and trimethoprim and their metabolites in a patient with peritonitis undergoing continuous ambulatory peritoneal dialysis.

Cefradine and co-trimoxazole pharmacokinetics were studied in a patient with peritonitis that complicated continuous ambulatory peritoneal dialysis (CAPD). Concentrations in the plasma reached after oral administration of 500 mg cefradine four times daily and 400/80 mg co-trimoxazole four times daily were for cefradine 100 micrograms/ml, for trimethoprim 15 micrograms/ml, and for sulfamethoxazole 100 micrograms/ml, respectively. In the dialysate concentrations were reached of 35-70 micrograms/ml cefradine, 2-5 micrograms/ml trimethoprim and 8-17 micrograms/ml sulfamethoxazole. The values for sulfamethoxazole are regarded too low to be clinically effective. Half-lives, protein binding values and CAPD clearances are presented. Low CAPD clearances were obtained during the night and high values during the day. The dosage yielded too high plasma trimethoprim concentrations, while sulfamethoxazole dialysate concentrations were too low. It seems questionable therefore whether co-trimoxazole can be used orally for the treatment of CAPD peritonitis.