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Biomedical subjects

C Richter

Publications and source records attributed to C Richter.

At least 55 records · Page 3Linked to original sources
PubMed2000

Direct localization of a beta-subunit domain on the three-dimensional structure of Escherichia coli RNA polymerase.

To identify the location of a domain of the beta-subunit of Escherichia coli RNA polymerase (RNAP) on the three-dimensional structure, we developed a method to tag a nonessential surface of the multisubunit enzyme with a protein density easily detectable by electron microscopy and image processing. Four repeats of the IgG-binding domain of Staphylococcus aureus protein A were inserted at position 998 of the E. coli RNAP beta-subunit. The mutant RNAP supported E. coli growth and showed no apparent functional defects in vitro. The structure of the mutant RNAP was determined by cryoelectron microscopy and image processing of frozen-hydrated helical crystals. Comparison of the mutant RNAP structure with the previously determined wild-type RNAP structure by Fourier difference analysis at 20-A resolution directly revealed the location of the inserted protein domain, thereby locating the region around position 998 of the beta-subunit within the RNAP three-dimensional structure and refining a model for the subunit locations within the enzyme.

Crystallization↗
PubMed2000

Hepatic steatosis and lactic acidosis caused by stavudine in an HIV-infected patient.

Lactic acidosis and hepatic steatosis caused by mitochondrial toxicity of nucleoside reverse transcriptase inhibitors (NRTI) is a rare cause of liver disease with a high mortality rate. This report describes a male, HIV-positive patient with a 4-week history of nausea, vomiting and abdominal pain. His medication consisted of prednisone 5 mg od (because of auto-immune thrombocytopenia), didanosine (for 2 years) and stavudine (for 3 months). Laboratory studies showed cholestasis and elevation of aminotransferases. Lactic level was not measured. Liver biopsy revealed steatosis and cholestatic hepatitis. In the absence of other causes of liver disease a probable diagnosis of stavudine-induced hepatic toxicity was made. After discontinuation of NRTI, he recovered completely. Because lactic acidosis had not been confirmed, stavudine was restarted and within 1 week the lactate level increased significantly. Therefore stavudine was discontinued again. One year later the patient is doing well on a double protease inhibitor regimen. In conclusion, clinicians treating patients with NRTI should be aware of the risk of lactic acidosis and hepatic steatosis. When this is suspected, all NRTI must be stopped. The diagnosis can be made when elevated lactate levels and hepatic steatosis are present in the absence of other causes of liver disease.

Abdominal Pain↗
PubMed2000

Cryptography with DNA binary strands.

Biotechnological methods can be used for cryptography. Here two different cryptographic approaches based on DNA binary strands are shown. The first approach shows how DNA binary strands can be used for steganography, a technique of encryption by information hiding, to provide rapid encryption and decryption. It is shown that DNA steganography based on DNA binary strands is secure under the assumption that an interceptor has the same technological capabilities as sender and receiver of encrypted messages. The second approach shown here is based on steganography and a method of graphical subtraction of binary gel-images. It can be used to constitute a molecular checksum and can be combined with the first approach to support encryption. DNA cryptography might become of practical relevance in the context of labelling organic and inorganic materials with DNA 'barcodes'.

Computer Communication Networks↗
PubMed2000

Peroxynitrite formed by mitochondrial NO synthase promotes mitochondrial Ca2+ release.

Mitochondria contribute to the maintenance of the intracellular Ca2+ homeostasis by taking up and releasing the cation via separate and specific pathways. The molecular details of the release pathway are elusive but its stimulation by the cross-linking of some vicinal thiols and consequently NAD+ hydrolysis are known. Thiol cross-linking and NAD+ hydrolysis can be achieved by addition of peroxynitrite (ONOO-), the product of the reaction between superoxide (O2-) and nitric oxide (nitrogen monoxide, NO*) to mitochondria. Mitochondria contain an NO synthase (mtNOS), which is stimulated by Ca2+, and are a copious source of O2-. We show here that intramitochondrially formed ONOO- stimulates the specific, NAD+-linked Ca2+ release from mitochondria. Our findings that upon Ca2+ uptake mtNOS is stimulated, that ONOO- is formed, and that Ca2+ is subsequently released from intact mitochondria suggest the existence of a feedback loop, which prevents overloading of mitochondria with Ca2+.

Animals↗
PubMed2000

Effectiveness of recombinant erythropoietin and iron sucrose vs. iron therapy only, in patients with postpartum anaemia and blunted erythropoiesis.

BACKGROUND: To compare efficacy between recombinant human erythropoietin (rhEPO) plus parenteral iron vs. iron alone (parenteral vs. oral) in postpartum anaemia. METHODS: Sixty patients (haemoglobin 8.6 +/- 1.1 g dL-1) were randomized to rhEPO plus intravenous (i.v.) iron sucrose (group 1), rhEPO placebo plus i.v. iron sucrose (group 2), or oral iron alone (group 3), daily for 4 days beginning 48-72 h postpartum. Erythropoiesis and iron status were assessed before, and on 4, 7 and 14 days after, starting therapy. RESULTS: On day 7 the group 1 haematocrit increase was 7.7 +/- 3.1% vs. 5.3 +/- 1.9% (group 2, P < 0.01) and 4.4 +/- 3.2% (group 3, P < 0.01), and on day 14, 11.3 +/- 2.9% vs. 9.2 +/- 3.4% (group 2, P < 0.05) and 8 +/- 2.8% (group 3, P < 0.01). The odds of achieving a target haematocrit > 32% on day 7 and > 35% on day 14 were higher on rhEPO (1.5-2.7) than on either iron regimen alone. Group 1 reticulocyte counts were also higher on days 4 (P < 0.05 vs. oral iron) and 7 (P < 0.01 vs. oral and parenteral iron). CONCLUSION: All three regimens were effective in postpartum anaemia, but the haematocrit and reticulocyte responses to rhEPO plus parenteral iron were significantly greater than to iron alone. Benefit was greatest in the blunted erythropoiesis subgroup with elevated post-Caesarean section C-reactive protein levels.

Anemia↗
PubMed2000

Comparison of impact data in hockey, football, and soccer.

PURPOSE: To compare accelerational forces to the head in high school-level football, hockey, and soccer athletes. METHODS: Acceleration of impact was measured within the helmet of high school hockey and football players during actual game play. A triaxial accelerometer was placed at the vertex of the helmet immediately adjacent to the players head. Peak acceleration (in g's) was measured and the Gadd Severity Index and Head Injury Criterion score calculated during actual play periods in several games over four seasons. We also recorded acceleration of head impacts in high school-level soccer players who headed a soccer ball while equipped with a football helmet instrumented identically to the helmet used to record during football games. RESULTS: Peak accelerations inside the helmet for football averaged 29.2 g compared with 35 g for hockey (p = .004). There were no incidents of concussion or other traumatic brain injury during the recorded periods. In contrast, the peak accelerations associated with heading a soccer ball was 54.7 g (p = 2 x 10(-5) vs. hockey). CONCLUSION: Peak accelerations as measured at the surface of the head were 160 to 180% greater from heading a soccer ball than from routine (noninjurious) impacts during hockey or football, respectively. The effect of cumulative impacts at this level may lead to neurologic sequelae.

Acceleration↗
PubMed2000

Contrasting effects of NO and peroxynitrites on HSP70 expression and apoptosis in human monocytes.

The free radicals nitric oxide (.NO) and superoxide (O(2)(-).) react to form peroxynitrite (ONOO(-)), a highly toxic oxidant species. In this study we investigated the respective effects of NO and ONOO(-) in monocytes from healthy human donors. Purified monocytes were incubated for 6 or 16 h with a pure NO donor (S-nitroso-N-acetyl-DL-penicillamine, 0-2 mM), an.NO/ONOO(-) donor (3-morpholinosydnonimine chlorhydrate, 0-2 mM) with and without superoxide dismutase (200 IU/ml), or pure ONOO(-). We provide evidence that 3-morpholinosydnonimine chlorhydrate alone represents a strong stress to human monocytes leading to a dose-dependent increase in heat shock protein-70 (HSP70) expression, mitochondrial membrane depolarization, and cell death by apoptosis and necrosis. These phenomena were abolished by superoxide dismutase, suggesting that ONOO(-), but not.NO, was responsible for the observed effects. This observation was further strengthened by the absence of a stress response in cells exposed to S-nitroso-N-acetyl-DL-penicillamine. Conversely, exposure of cells to ONOO(-) alone also induced mitochondrial membrane depolarization and cell death by apoptosis and necrosis. Thus ONOO(-) formation may well explain the toxic effect generally attributed to.NO.

Apoptosis↗
PubMed2000

Effect of ebselen on Ca2+ transport in mitochondria.

The seleno-organic compound ebselen mimics the glutathione-dependent, hydroperoxide reducing activity of glutathione peroxidase. The activity of glutathione peroxidase determines the rate of hydroperoxide-induced Ca2+ release from mitochondria. Ebselen stimulates Ca2+ release from mitochondria, accelerates mitochondrial respiration and uncoupling, and induces mitochondrial swelling, indicating a deterioration of mitochondrial function. These manifestations are abolished by cyclosporine A, a potent inhibitor of the mitochondrial permeability transition. However, when ebselen-induced Ca2+ cycling is prevented with ruthenium red, an inhibitor of the Ca2+ uniporter, or by chelation of extramitochondrial Ca2+ by EGTA, no detectable elevation of swelling or uncoupling is observed. The release of Ca2+ from mitochondria is delayed in the absence of rotenone, i.e. when pyridine nucleotides are maintained in the reduced state due to succinate-driven reversed electron flow. We suggest that ebselen induces Ca2+ release from intact mitochondria via an NAD+ hydrolysis-dependent mechanism.

Animals↗
PubMed2000

Anti- and pro-oxidative properties of PADMA 28, a Tibetan herbal formulation.

There is growing public interest in traditional medicine. PADMA 28, a multicompound herbal preparation derived from Tibetan medicine, has proven efficacy in some clinical trials and tests at the cellular level. We report here on studies of PADMA 28 at the molecular level. Extracts of PADMA 28 contain both reducing and metal ion-chelating substances. In this way, PADMA 28 acts as a powerful antioxidant or prooxidant, depending on its concentration and the reaction under study.

Antioxidants↗
PubMed2000

Shortening the duration of treatment for cervical tuberculous lymphadenitis.

The aim of the study was to determine the optimal duration of treatment for patients with tuberculous lymphadenitis. The Medline database was searched for relevant articles published between 1978-1997. Inclusion criteria were study populations of patients with predominantly cervical tuberculous lymphadenitis in whom the diagnosis had been confirmed bacteriologically and/or histologically, or was made probable by using clinical and laboratory markers. Treatment management had to include at least isoniazid, rifampicin and pyrazinamide and a follow-up of at least 12 months after the end of treatment. Patients with resistance to rifampicin and pyrazinamide and previous treatment for tuberculosis were excluded. The number of patients who relapsed after treatment was calculated. The study population in eight out of the 35 articles retrieved were suitable for analysis. Some concerned comparative studies. There were eight treatment schedules of 6 months' duration and three schedules of 9 months' duration. Treatment for 6 months resulted in a tuberculous lymphadenitis relapse rate of 13/422=3.3% (95% confidence interval: 1.7-5.5), with a mean follow-up of 31 months after completion of treatment. Treatment for 9 months resulted in a relapse rate of 3/112=2.7% (95% confidence interval: 0.6-7.8), with a mean follow-up of 20 months. Despite the limitations of the literature available, 6 months of therapy is probably sufficient for patients with tuberculous lymphadenitis.

Antitubercular Agents↗
PubMed1999

Mitochondrial nitric-oxide synthase stimulation causes cytochrome c release from isolated mitochondria. Evidence for intramitochondrial peroxynitrite formation.

Nitric oxide (NO) is synthesized by members of the NO synthase (NOS) family. Recently the existence of a mitochondrial NOS (mtNOS), its Ca(2+) dependence, and its relevance for mitochondrial bioenergetics was reported (Ghafourifar, P., and Richter, C. (1997) FEBS Lett. 418, 291-296; Giulivi, C., Poderoso, J. J., and Boveris, A. (1998) J. Biol. Chem. 273, 11038-11043). Here we report on the possible involvement of mtNOS in apoptosis. We show that uptake of Ca(2+) by mitochondria triggers mtNOS activity and causes the release of cytochrome c from isolated mitochondria in a Bcl-2-sensitive manner. mtNOS-induced cytochrome c release was paralleled by increased lipid peroxidation. The release of cytochrome c as well as increase in lipid peroxidation were prevented by NOS inhibitors, a superoxide dismutase mimic, and a peroxynitrite scavenger. We show that mtNOS-induced cytochrome c release is not mediated via the mitochondrial permeability transition pore because the release was aggravated by cyclosporin A and abolished by blockade of mitochondrial calcium uptake by ruthenium red. We conclude that, upon Ca(2+)-induced mtNOS activation, peroxynitrite is formed within mitochondria, which causes the release of cytochrome c from isolated mitochondria, and we propose a mechanism by which elevated Ca(2+) levels induce apoptosis.

Animals↗
PubMed1999

Crystal structure of Thermus aquaticus core RNA polymerase at 3.3 A resolution.

The X-ray crystal structure of Thermus aquaticus core RNA polymerase reveals a "crab claw"-shaped molecule with a 27 A wide internal channel. Located on the back wall of the channel is a Mg2+ ion required for catalytic activity, which is chelated by an absolutely conserved motif from all bacterial and eukaryotic cellular RNA polymerases. The structure places key functional sites, defined by mutational and cross-linking analysis, on the inner walls of the channel in close proximity to the active center Mg2+. Further out from the catalytic center, structural features are found that may be involved in maintaining the melted transcription bubble, clamping onto the RNA product and/or DNA template to assure processivity, and delivering nucleotide substrates to the active center.

Catalytic Domain↗
PubMed1999

Randomized trial comparing saquinavir soft gelatin capsules versus indinavir as part of triple therapy (CHEESE study).

OBJECTIVE: To compare efficacy and tolerability of saquinavir soft gelatin capsule (SQV-SGC) formulation and indinavir, both given as part of a triple drug regimen containing zidovudine and lamivudine, in HIV-1-infected individuals. DESIGN: Randomized, open label, multicentre study. PATIENTS: A total of 70 patients who were antiretroviral-naive and who had a CD4 cell count < 500 x 10(6)/I and/or > 10000 HIV RNA copies/ml plasma and/or HIV-related symptoms. Subjects were assigned randomly to zidovudine 200 mg three times per day plus lamivudine 150 mg twice per day plus either SQV-SGC 1200 mg three times per day (SQV-SGC group) or indinavir 800 mg three times per day (indinavir group). Data are presented for all patients up to week 24. RESULTS: Mean baseline CD4 cell counts (+/- SE) were 301+/-29 x 10(6) cells/l and 310 +/-43 x 10(6) cells/l in the SQV-SGC and indinavir groups, respectively. The log10 median baseline HIV RNA load was 5.00 copies/ml in the SQV-SGC group and 4.98 copies/ml in the indinavir group. No difference in antiretroviral effect between the treatment arms could be demonstrated. Intention-to-treat analysis (last observation carried forward [LOCF]) at week 24 revealed that RNA levels decreased to < 50 copies/ml in 74.3% of patients in the SQV-SGC group and in 71.4% of the patients in the indinavir group (P = 0.78). In the on-treatment analysis the proportion of patients < 50 copies/ml at week 24 was 88.0% in the SQV-SGC group and 84.6% in the indinavir group (P = 0.725). Intriguingly, the mean increase of CD4 cells in the first 24 weeks was 162+/-20 x 10(6) cells/l in the SQV-SGC group and 89+/-21 x 10(6) cells/l in the indinavir group (P = 0.01), but preliminary data indicate that this difference in CD4 cell count gain may disappear after 24 weeks of treatment. Both regimens were generally well tolerated. CONCLUSION: During the first 24 weeks of the study, we found no difference in antiviral potency between the indinavir group and the SQV-SGC group. A significantly higher CD4 response in the SQV-SGC group was observed.

Adult↗
PubMed1999

Ceramide induces cytochrome c release from isolated mitochondria. Importance of mitochondrial redox state.

In the present study we show that N-acetylsphingosine (C2-ceramide), N-hexanoylsphingosine (C6-ceramide), and, to a much lesser extent, C2-dihydroceramide induce cytochrome c (cyto c) release from isolated rat liver mitochondria. Ceramide-induced cyto c release is prevented by preincubation of mitochondria with a low concentration (40 nM) of Bcl-2. The release takes place when cyto c is oxidized but not when it is reduced. Upon cyto c loss, mitochondrial oxygen consumption, mitochondrial transmembrane potential (Delta Psi), and Ca2+ retention are diminished. Incubation with Bcl-2 prevents, and addition of cyto c reverses the alteration of these mitochondrial functions. In ATP-energized mitochondria, ceramides do not alter Delta Psi, neither when cyto c is oxidized nor when it is reduced, ruling out a nonspecific disturbance by ceramides of mitochondrial membrane integrity. Furthermore, ceramides decrease the reducibility of cyto c. We conclude that the apoptogenic properties of ceramides are in part mediated via their interaction with mitochondrial cyto c followed by its release and that the redox state of cyto c influences its detachment by ceramide from the inner mitochondrial membrane.

Animals↗
PubMed1999

Fragmented mitochondrial DNA is the predominant carrier of oxidized DNA bases.

Previous analyses indicated a high level of oxidative base modification in mitochondrial DNA, the extent of which raised questions about the methodological validity and biological implications. In the present study DNA was isolated from rat liver mitochondria under carefully controlled conditions, and the extent of base oxidation, DNA fragmentation, and nuclear DNA contamination were analyzed. DNA isolated from intact mitochondria treated with DNase consisted of 16.3 kilobase pairs, mostly circular, mitochondrial DNA molecules and a mixture of nuclear and mitochondrial DNA fragments, as identified by agarose gel electrophoresis and hybridization. High-performance liquid chromatography in combination with electrochemical detection confirmed that the overall level of 8-hydroxy-2'-deoxyguanosine, a marker commonly used in the analysis of base oxidation, is higher in mitochondrial than in nuclear DNA. Importantly, 8-hydroxy-2'-deoxyguanosine is relatively scarce in the 16.3 kilobase pair mitochondrial DNA molecules (0.051 pmol/microgram) but is present in high levels in mitochondrial DNA fragments (0.741 pmol/microgram). The fragments constitute about 18% of total mitochondrial DNA. The antitumor agent bleomycin, which binds to DNA, forms an iron complex capable of transferring electrons from Fe2+ to molecular oxygen. Exposure of mitochondria to bleomycin and iron resulted in nicking but not in a significant increase in base oxidation of 16.3 kilobase pair mitochondrial DNA, whereas the amount and the oxidation level of fragmented mitochondrial DNA significantly increased. These findings are relevant for a better understanding of the role of mitochondria in aging and various diseases and are consistent with the notion that despite the overall high DNA oxidation level, mitochondria can faithfully proliferate.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1999

Solar photocatalytic mineralization of commercial pesticides: methamidophos.

Aqueous solutions of methamidophos (O,S-dimethyl phosphoramidothioato) are mineralised in sunlight in the presence of dispersed particles of TiO2 in a pre-industrial pilot plant. A commercial pesticide (Tamaron 50, 50% Methamidophos) is used as a model to demonstrate the application of the treatment. Total Organic Carbon (TOC) and PO4(3-) are analysed to confirm the mineralization of the contaminants. The concentration of organics in water was set at TOC approximately 100 mg/L, but with only illuminated TiO2, mineralization was slow. Oxidation of the organics present in the formulation is enhanced 18 times by use of an appropriate electron scavenger (peroxydisulphate) compared to TiO2 alone.

Catalysis↗