The TCA cycle and tumorigenesis: the examples of fumarate hydratase and succinate dehydrogenase

It is well documented that disturbances in mitochondrial function are associated with rare childhood disorders and possibly with many common diseases of ageing, such as Parkinson's disease and dementia. There has also been increasing evidence linking mitochondrial dysfunction with tumorigenesis. Recently, heterozygous germline mutations in two enzymes of the Krebs tricarboxylic acid cycle (TCA cycle) have been shown to predispose individuals to tumours. The two enzymes, fumarate hydratase (FH) and succinate dehydrogenase (SDH), are ubiquitously expressed, playing a vital role in adenosine triphosphate (ATP) production through the mitochondrial respiratory chain. Germline mutations in FH are associated with leiomyomatosis and renal cell carcinoma, whilst SDH mutations are associated with predisposition to paraganglioma (PGL) and phaeochromocytoma (PCC). At present, there are few data to explain the pathway(s) involved in this predisposition to neoplasia through TCA cycle defects. We shall review the mechanisms by which mutations in FH and SDH might play a role in tumorigenesis. These include pseudo-hypoxia, mitochondrial dysfunction and impaired apoptosis, oxidative stress and anabolic drive. All of these mechanisms are currently poorly defined. To date, FH and SDH mutations have not been reported in non-familial leiomyomata, renal cancers, PCCs or PGLs. It remains entirely possible, however, that the underlying mechanisms of tumorigenesis in these sporadic tumours are the same as those in the Mendelian syndromes.     

Autoimmunity, Inflammatory Bowel Disease and Hyposplenism

The presence or absence of nine autoantibodies were assessedin 44 patients with ulcerative colitis (17 with hyposplenism)and 22 patients with Crohn's disease (eight with hyposplenism).The purpose of the study was to determine whether hyposplenismin inflammatory bowel disease is associated with an increasedtendency to autoimmunity, or whether autoimmunity is linkednot to hyposplenism itself but to the underlying bowel disease.The results strongly suggest that the latter hypothesis is correct.There was a much higher frequency of autoantibodies in patientswith ulcerative colitis than in those with Crohn's disease (P0.01),suggesting that autoimmune factors are more important in thepathogenesis of ulcerative colitis than in Crohn's disease.     

Six-week randomized controlled trial to compare the tolerabilities, pharmacokinetics, and antiviral activities of GW433908 and amprenavir in human immunodeficiency virus type 1-infected patients

This study compared the plasma amprenavir pharmacokinetics of the human immunodeficiency virus (HIV) protease inhibitors amprenavir (Agenerase) 1,200 mg twice daily (BID) and the amprenavir prodrug GW433908, a formulation that substantially reduces the number of tablets per dose compared with amprenavir, at doses of 1,395 mg and 1,860 mg BID, in combination with abacavir 300 mg BID and lamivudine 150 mg BID in patients with HIV infection. Overall, 78 patients received study treatment. Compared with amprenavir 1,200 mg BID, both GW433908 1,395 mg BID and GW433908 1,860 mg BID delivered equivalent steady-state (ss) values for area under the plasma amprenavir concentration-time curve (AUC) at the end of a dosing interval (tau), lower maximum plasma amprenavir concentrations (30% lower), and higher plasma amprenavir concentrations at the end of a dosing interval (28% higher for GW433908 1,395 mg BID and 46% higher for GW433908 1,860 mg BID). Time-variant plasma amprenavir pharmacokinetics were observed with reductions in plasma amprenavir exposure over the first 4 weeks of dosing; the decrease in plasma amprenavir AUC(tau,ss) versus the AUC from 0 h to infinity was 27% for GW43308 1,395 mg, 45% for GW433908 1,860 mg, and 23% for amprenavir 1,200 mg. All three regimens reduced plasma HIV-1 RNA ( approximately 2 log(10) copies/ml) and increased CD4(+) cell counts ( approximately 100 cells/mm(3)) over the initial 28 days. Adverse event profiles were consistent with those previously reported for amprenavir. Although not statistically tested, the GW433908 groups appeared to have fewer gastrointestinal symptoms. In conclusion, the protease inhibitor GW433908 delivered comparable plasma amprenavir concentrations to those delivered by amprenavir 1,200 mg BID. GW433908, in combination with abacavir and lamivudine, demonstrated potent antiviral activity and was generally well tolerated over a 4-week period.     

Mutational analysis of 105 mucopolysaccharidosis type VI patients

Mucopolysaccharidosis type VI (MPS VI; Maroteaux-Lamy syndrome) is a lysosomal storage disorder caused by mutations in the N-acetylgalactosamine-4-sulfatase (arylsulfatase B, ARSB) gene. ARSB is a lysosomal enzyme involved in the degradation of the glycosaminoglycans (GAG) dermatan and chondroitin sulfate. ARSB mutations reduce enzyme function and GAG degradation, causing lysosomal storage and urinary excretion of these partially degraded substrates. Disease onset and rate of progression is variable, producing a spectrum of clinical presentation. In this study, 105 MPS VI patients-representing about 10% of the world MPS VI population-were studied for molecular genetic and biochemical parameters. Direct sequencing of patient genomic DNA was used to identify ARSB mutations. In total, 83 different disease-causing mutations were found, 62 of which were previously unknown. The novel sequence changes included: 38 missense mutations, five nonsense mutations, 11 deletions, one insertion, seven splice-site mutations, and four polymorphisms. ARSB mutant protein and residual activity were determined on fibroblast extracts for each patient. The identification of many novel mutations unique to individuals/their families highlighted the genetic heterogeneity of the disorder and provided an appropriate cohort to study the MPS VI phenotypic spectrum. This mutation analysis has identified a clear correlation between genotype and urinary GAG that can be used to predict clinical outcome.     

Redesign of a weight-based heparin protocol for improved outcomes in a community hospital

Heparin has long been used to minimize complications of a variety of disorders and reaching a therapeutic level within 24 hours has been shown to improve patient outcomes. However, the dosing of heparin to reach a therapeutic level has been controversial. At our institution a weight based heparin nomogram was in place, however an initial data analysis revealed a large percent of the initial aPTT levels were above therapeutic range. This article will describe the performance improvement process and the overall goal to decrease the percentage of patients exceeding the initial therapeutic aPTT levels without compromising daily therapeutic aPTT levels.     

Detection of genes for enterotoxins, exfoliative toxins, and toxic shock syndrome toxin 1 in Staphylococcus aureus by the polymerase chain reaction.

Eight pairs of synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) protocol to detect genes for staphylococcal enterotoxins A to E, exfoliative toxins A and B, and toxic shock syndrome toxin 1 in Staphylococcus aureus strains isolated from clinical specimens and contaminated foods. Primers were targeted to internal regions of the toxin genes, and amplification fragments were detected after the PCR by agarose gel electrophoresis. Unequivocal discrimination of toxin genes was obtained by the PCR by using nucleic acids extracted from 88 strains of S. aureus whose toxigenicity was established biologically and immunologically. In immunological assays, two strains of S. aureus produced equivocal results for production of enterotoxin C or toxic shock syndrome toxin 1, giving an overall concordance between phenotypic and genotypic identification of 97.7%. Primer specificity was established in the PCR by using nucleic acids from known toxin-producing bacterial pathogens and from nontoxigenic S. aureus. Strains of Streptococcus spp., including some producers of pyrogenic exotoxin A carrying the speA gene, were negative by the PCR designed to detect staphylococcal toxins. The detection limits were established for all the staphylococcal toxin genes within their respective PCR protocols. The identification of staphylococcal toxin genes in strains of S. aureus by the PCR offers a very specific, sensitive, relatively rapid, and inexpensive alternative to traditional immunological assays which depend on adequate gene expression for reliability and sensitivity.     

Suppressor Lyt 2+ T-cells demonstrated in mice late after thermal injury

Biological properties of suppressor cells mediating second suppressive state (S-SupS) of IFN-gamma responsiveness in mice following thermal injury were investigated by their ability to inhibit [3H]thymidine incorporation by allogeneic cell-stimulated mononuclear cells (MNC) prepared from spleens of unburned normal mice (N-mice). Kinetic studies indicated that the suppressor cells appeared at approximately 15 days post-burn and persisted until 40 days after thermal injury. This cell population from the spleens of thermally injured mice (TI-mice) was inactivated by anti-Thy 1.2 monoclonal antibody (mAb) plus complement (C). Treatment with antimouse immunoglobulin and anti-Lyt 1.2 mAb followed by C failed to abrogate the activity. However, it could be eliminated by treatment with anti-Lyt 2.2 mAb and C. In addition, when Thy 1+ or Lyt 2+ T cells were depleted from spleen cells containing suppressor cell activities by treatment with mAbs and C, the suppressor activity for IFN-gamma production by N-mice splenic MNC stimulated with concanavalin A was also eliminated. However, no alteration in the suppression of IFN production was observed when this splenic cell population from TI-mice was treated with anti-Lyt 1.2 mAb and anti-mouse Ig followed by C. Since it has been reported [1] that the suppressor cell in spleens of TI-mice could neither be removed by plastic adherence nor by carbonyl-iron treatment, this secondarily-generated suppressor cell was concluded to be a T cell possessing the Lyt 1-2+ phenotype. The present report describes the demonstration, following a generation of suppressor macrophages [2], of suppressor Lyt 2+ T cells spleen of mice late after thermal injury.     

Ba2+-induced ATP release from adrenal medullary chromaffin cells is mediated by Ba2+ entry through both voltage- and receptor-gated Ca2+ channels

We have measured on-line the exocytotic secretion of ATP from adrenal medullary chromaffin cells induced by Ba2+ using a luciferin/luciferase assay. We have found that Ba2+-induced ATP release requires the entry of Ba2+ through either voltage- or receptor-gated Ca2+ channels. This conclusion is based on the observations that short preincubations with low concentrations of either nicotine or K+ greatly enhance Ba2+-induced ATP release and that this augmentation can be blocked with the nicotinic receptor antagonist, hexamethonium, and the Ca2+ antagonist, nifedipine, respectively. Moreover, both nicotine and K+ stimulate 133Ba2+ uptake, which in the case of K+ is inhibited by nifedipine. These results support the hypothesis that the cellular events leading to Ba2+-induced secretion coincide at least in part with the events leading to Ca2+-dependent exocytosis.     

Help-seeking patterns of community residents with depressive symptoms.

From a community sample, fifty-five individuals identified as having three or more symptoms of depression were asked if and where they had sought help and how they found or would find treatment. Only 20 subjects (33.9%) reported having sought professional help. No demographic differences were found between help-seekers and non-help-seekers. Eighty percent of the help-seekers had seen a mental health professional while 20% had seen a non-psychiatric physician. However, non-psychiatric physicians and friends or personal acquaintances were most frequently cited as the first point of contact for locating treatment. Results suggest that most individuals with a moderate number of depressive symptoms do not seek professional assistance but that, among those who do, a majority is eventually seen by a mental health professional. Virtually all utilize intermediaries other than mental health professionals, however, to locate assistance. The non-psychiatric physician plays a prominent role in locating help for depressed individuals, which suggests the need to educate physicians about appropriate treatment referral.