Histone deacetylase inhibitors in plasma cell leukemia treatment: effect of bone marrow microenvironment.

In the presented study we analysed the effect of histone deacetylase inhibitors (HDACi) suberoylanilide hydroxamic acid (SAHA) and valproic acid (VPA) on human plasma cell leukemia (PCL) cell line UHKT-944 in the presence of bone marrow microenvironment (BMM). For the analysis, the cells were cultured alone, with bone marrow stromal cells (BMSCs), with extracellular matrix (ECM) components or with interleukin-6, and treated with varied concentrations of SAHA and VPA for 24/48 hours. To study the effect of HDACi, we investigated cell proliferation, apoptosis, cell cycle and changes in selected signalling pathways. We found that both SAHA and VPA induced apoptosis, but had no effect on the cell cycle distribution of UHKT-944 cells. Investigation of the antiproliferative effect of SAHA and VPA revealed that BMSCs and high concentration of interleukin-6 had partial protective effect against SAHA or both inhibitors, respectively. No effect of ECM components on the efficiency of HDACi was observed. We further revealed that VPA down-regulated STAT3 phosphorylation while both inhibitors decreased Akt phosphorylation. In conclusion, VPA and SAHA might represent an additional therapeutic strategy in the PCL treatment. Protective effect of BMM should be taken into account when investigating prospective therapeutic agents against plasma cell disorders.

Mitochondrial uncoupling proteins--facts and fantasies.

Instead of a comprehensive review, we describe the basic undisputed facts and a modest contribution of our group to the fascinating area of the research on mitochondrial uncoupling proteins. After defining the terms uncoupling, leak, protein-mediated uncoupling, we discuss the assumption that due to their low abundance the novel mitochondrial uncoupling proteins (UCP2 to UCP5) can provide only a mild uncoupling, i.e. can decrease the proton motive force by several mV only. Contrary to this, the highly thermogenic role of UCP1 in brown adipose tissue is not given only by its high content (approximately 5 % of mitochondrial proteins) but also by the low ATP synthase content and high capacity respiratory chain. Fatty acid cycling mechanism as a plausible explanation for the protonophoretic function of all UCPs and some other mitochondrial carriers is described together with the experiments supporting it. The phylogenesis of all UCPs, estimated UCP2 content in several tissues, and details of UCP2 activation are described on the basis of our experiments. Functional activation of UCP2 is proposed to decrease reactive oxygen species (ROS) production. Moreover, reaction products of lipoperoxidation such as cleaved hydroperoxy-fatty acids and hydroxy-fatty acid can activate UCP2 and promote feedback down-regulation of mitochondrial ROS production.

Possible basic and specific functions of plant uncoupling proteins (pUCP).

Evidence has been provided that the plant uncoupling proteins (pUCP) play basic physiological roles similar to the other uncoupling protein subfamily members (mammalian UCP1,2,3,4 and BMCP) and are effective in the situations of slight uncoupling that leads to: (1) accelerated respiration and metabolic rates that are beneficial to plant growth and development; (2) decreased formation of reactive oxygen species in mitochondria; and, (3) mild thermogenesis, inevitably accompanying the previous two phenomena. Hypothetically, specific physiological roles of pUCP such as cut off of ATP synthesis could be manifested in connection with climacteric respiratory rise during fruit ripening, seed dormancy, and plant senescence. pUCP might also facilitate growth under low temperatures, e.g., during seed germination or in roots. The existence of these specific roles is suggested by the immunochemical and functional localization of pUCP in mitochondria of fruits, seeds and roots of various plant species.

Ketorolac vs tramadol in the treatment of postoperative pain during maxillofacial surgery.

BACKGROUND: This study aims to assess the best postoperative analgesia during maxillofacial surgery by using small doses of ketorolac or tramadol or their association and evaluates the presence of adverse effects due to NSAID or opioid use. METHODS: After their informed consent, 51 patients ASA I and II undergoing major maxillofacial surgery, were randomised in three groups and the following protocol was used: group K received ketorolac (30 mg i.v.) at the time of skin closure and repeated after 8 hrs and 16 hrs from the end of the operation. Group T received tramadol (100 mg i.v.) in the same condition; and group KT received first tramadol (100 mg i.v.) during surgery and then ketorolac (30 mg) was given in the administrations that followed. Meperidine 50 mg was used in case of unsatisfactory analgesia. Pain was evaluated using pain intensity scores 2, 4, 6, 12 and 24 hours from the end of the operation. Data was analysed using Anova and c2 test. RESULTS: The groups were comparable with regard to age, weight, duration of surgery. Very good postoperative analgesia was recorded in three groups. There is no difference statistically between K, T and KT groups in the pain scores measured. Only a low number of patients required opioids administration to achieve adequate analgesia. The patients were considered to have achieved excellent analgesia in 64.8% in T group, in 41.2% of the K group and in 58.8% of the KT group. There were no cases of insufficient analgesia. We did not find a significant difference considering BP, HR, respiratory depression in the post-operative period. Vomiting was registered in 41.2% of this T group vs 11.2% of the K group and in 35.5% of the KT group. CONCLUSIONS: Ketorolac and Tramadol produced comparable, effective and low cost postoperative analgesia during maxillofacial surgery. There are only statistically significant differences considering side effects.

Natural and azido fatty acids inhibit phosphate transport and activate fatty acid anion uniport mediated by the mitochondrial phosphate carrier.

The electroneutral P(i) uptake via the phosphate carrier (PIC) in rat liver and heart mitochondria is inhibited by fatty acids (FAs), by 12-(4-azido-2-nitrophenylamino)dodecanoic acid (AzDA) and heptylbenzoic acid ( approximately 1 microm doses) and by lauric, palmitic, or 12-azidododecanoic acids ( approximately 0.1 mm doses). In turn, reconstituted E. coli-expressed yeast PIC mediated anionic FA uniport with a similar pattern leading to FA cycling and H(+) uniport. The kinetics of P(i)/P(i) exchange on recombinant PIC in the presence of AzDA better corresponded to a competitive inhibition mechanism. Methanephosphonate was identified as a new PIC substrate. Decanephosphonate, butanephosphonate, 4-nitrophenylphosphate, and other P(i) analogs were not translocated and did not inhibit P(i) transport. However, methylenediphosphonate and iminodi(methylenephosphonate) inhibited both electroneutral P(i) uptake and FA cycling via PIC. AzDA analog 16-(4-azido-2-nitrophenylamino)-[(3)H(4)]-hexadecanoic acid ((3)H-AzHA) bound upon photoactivation to several mitochondrial proteins, including the 30- and 34-kDa bands. The latter was ascribed to PIC due to its specific elution pattern on Blue Sepharose and Affi-Gel. (3)H-AzHA photolabeling of recombinant PIC was prevented by methanephosphonate and diphosphonates and after premodification with 4-azido-2-nitrophenylphosphate. Hence, the demonstrated PIC interaction with monovalent long-chain FA anions, but with divalent phosphonates of short chain only, indicates a pattern distinct from that valid for the mitochondrial uncoupling protein-1.

[Comparison of 0.125% bupivacaine and 0.2% ropivacaine in obstetric analgesia]. / Confronto tra bupivacaina 0.125% e ropivacaina 0.2% nell'analgesia del parto.

This comparative study of low doses of ropivacaine was conducted in order to identify the most effective form of analgesia during labour with the aid of supplementary low doses of fentanyl and clonidine. 60 ASA I and II parturient primipares who had asked for epidural analgesia were randomly assigned to two groups. Group R was given 5-7 ml 0.2% ropivacaine and Group B 0.125% bupivacaine with both groups receiving 75 ng clonidine and 50 ng fentanyl with their first bolus of local anaesthetic. The parameters measured included the speed and spread of the sensory blockade and the scale of any motor blockade. The material haemodynamics and VAS pain relief scores were also measured at 30-minute intervals during labour and all side-effects (nausea, vomiting, localised or generalised itching, headache etc) were also monitored. Apgar anaesthetics and other drugs was decided on the basis of the VAS score (a further dose was given to women with a VAS of > 3-4). The study was completed by a telephone interview 6 months after delivery and the data were analysed using the Student's t-test and the chi 2 test. The analgesic effect was satisfactory in both groups and no statistically significant differences were found between the two groups under most of the headings analysed, apart from the top-up doses needed to maintain adequate analgesia. The average time between the first VAS to parturition was 292 mns in Group B and 267 mns in Groups R. Top-up doses of local anaesthetic (2.35 vs 5.05) came on average to 15.8 ml in Group B compared to 24.1 ml in Group R. There were 20% Caesarian sections in Group R and 13.8% in Group B. Optimum analgesia was achieved in Group R, the level of analgesia was insufficient or barely sufficient in 3.3% of cases. There was no Apgar score < 7 in either group. It was therefore concluded that both bupivacaine and ropivacaine offer excellent analgesia during labour and have no significant side effects on mothers or babies.

Interaction of mitochondrial phosphate carrier with fatty acids and hydrophobic phosphate analogs.

Mitochondrial transporters, in particular uncoupling proteins and the ADP/ATP carrier, are known to mediate uniport of anionic fatty acids (FAs), allowing FA cycling which is completed by the passive movement of FAs across the membrane in their protonated form. This study investigated the ability of the mitochondrial phosphate carrier to catalyze such a mechanism and, furthermore, how this putative activity is related to the previously observed HgCl(2)-induced uniport mode. The yeast mitochondrial phosphate carrier was expressed in Escherichia coli and then reconstituted into lipid vesicles. The FA-induced H(+) uniport or Cl(-) uniport were monitored fluorometrically after HgCl(2) addition. These transport activities were further characterized by testing various inhibitors of the two different transport modes. The phosphate carrier was found to mediate FA cycling, which led to H(+) efflux in proteoliposomes. This activity was insensitive to ATP, mersalyl or N-ethylmaleimide and was inhibited by methylenediphosphonate and iminodi(methylenephosphonate), which are new inhibitors of mitochondrial phosphate transport. Also, the HgCl(2) induced Cl(-) uniport mediated by the reconstituted yeast PIC, was found to be inhibited by these reagents. Both methylenediphosphonate and iminodi(methylenephosphonate) blocked unidirectional Cl(-) uptake, whereas Cl(-) efflux was inhibited by iminodi(methylenephosphonate) and phosphonoformic acid only. These results suggest that a hydrophobic domain, interacting with FAs, exists in the mitochondrial phosphate carrier, which is distinct from the phosphate transport pathway. This domain allows for FA anion uniport via the phosphate carrier and consequently, FA cycling that should lead to uncoupling in mitochondria. This might be considered as a side function of this carrier.

Occurrence of plant-uncoupling mitochondrial protein (PUMP) in diverse organs and tissues of several plants.

The presence of plant-uncoupling mitochondrial protein (PUMP), previously described by Vercesi et al. (1995), was screened in mitochondria of various organs or tissues of several plant species. This was done functionally, by monitoring purine nucleotide-sensitive linoleic acid-induced uncoupling, or by Western blots. The following findings were established: (1) PUMP was found in most of the higher plants tested; (2) since ATP inhibition of linoleic acid-induced membrane potential decrease varied, PUMP content might differ in different plant tissues, as observed with mitochondria from maize roots, maize seeds, spinach leaves, wheat shoots, carrot roots, cauliflower, broccoli, maize shoots, turnip root, and potato calli. Western blots also indicated PUMP presence in oat shoots, carnation petals, onion bulbs, red beet root, green cabbage, and Sedum leaves. (3) PUMP was not detected in mushrooms. We conclude that PUMP is likely present in the mitochondria of organs and tissues of all higher plants.

Existence of uncoupling protein-2 antigen in isolated mitochondria from various tissues.

Antibodies against Escherichia coli-expressed uncoupling protein-2 (UCP2) and uncoupling protein-3 (UCP3) were raised by operating the blotted proteins into the spleen of minipigs. The antisera reacted more intensively with the recombinant UCP2 and UCP3 than with uncoupling protein-1 (UCP1) isolated from brown adipose tissue. Moreover, anti-UCP2 and cross-reacting anti-UCP3 antibodies identified the presence of the UCP2/3 antigen in isolated mitochondria from rat heart, rat kidney, rat brain, rabbit epididymal white adipose tissue, hamster brown adipose tissue, and rabbit skeletal muscle. It has been concluded that UCP2 is expressed in these tissues (UCP3 in skeletal muscle); however their existence in mitochondria had not previously been demonstrated.

Fatty acid cycling mechanism and mitochondrial uncoupling proteins.

We hypothesize that fatty acid-induced uncoupling serves in bioenergetic systems to set the optimum efficiency and tune the degree of coupling of oxidative phosphorylation. Uncoupling results from fatty acid cycling, enabled by several phylogenetically specialized proteins and, to a lesser extent, by other mitochondrial carriers. It is suggested that the regulated uncoupling in mammalian mitochondria is provided by uncoupling proteins UCP-1, UCP-2 and UCP-3, whereas in plant mitochondria by PUMP and StUCP, all belonging to the gene family of mitochondrial carriers. UCP-1, and hypothetically UCP-3, serve mostly to provide nonshivering thermogenesis in brown adipose tissue and skeletal muscle, respectively. Fatty acid cycling was documented for UCP-1, PUMP and ADP/ATP carrier, and is predicted also for UCP-2 and UCP-3. UCP-1 mediates a purine nucleotide-sensitive uniport of monovalent unipolar anions, including anionic fatty acids. The return of protonated fatty acid leads to H+ uniport and uncoupling. UCP-2 is probably involved in the regulation of body weight and energy balance, in fever, and defense against generation of reactive oxygen species. PUMP has been discovered in potato tubers and immunologically detected in fruits and corn, whereas StUCP has been cloned and sequenced froma a potato gene library. PUMP is supposed to act in the termination of synthetic processes in mature fruits and during the climacteric respiratory rise.

[The ocular form of toxocariasis]. / Ocní forma larvální toxokarózy.

Ocular toxocariasis (ocular form of larval toxocariasis) arises mainly unilaterally and represents no rare disease. On 3rd Department of Infectology, 1st Faculty of Medicine, Charles University, Prague, 102 patients with proved larval toxocariasis were treated from 1981 to 1990. Ocular toxocariasis concerned only one third of this number. Most frequent form was the retinal toxocaral granuloma (in 55.2%), positioned by two thirds at the posterior pole of retina. In one case, endophthalmitis led to amaurosis of the eye. Clinical forms typical for ocular toxocariasis are presented. No statistically significant difference was observed in treatment effects using thiobendazole or diethylcarbamazine. All patients with ocular toxocariasis were treated with systematic steroids.