Trunk muscle activation characteristics in patients with severe haemophilia.

INTRODUCTION: Recurrent bleeding episodes in patients with haemophilia (PWH) lead to joint alterations and therewith disturbed muscle coordination patterns. Major weight-bearing joints are affected most. However, possible effects on trunk muscle activity have not been examined so far. The objective of this work was to study consequences of haemarthropathy on characteristics of trunk muscles in PWH while standing on surfaces with different mechanical properties. METHODS: Surface EMG of internal oblique (IO) and multifidus (MF) muscles were bilaterally recorded during a natural bilateral stance in 20 PWH with severe haemophilia A [age: 42 years (SD: 10)] and 25 non-haemophilic controls [NHC, 43 (12)]. Amplitude ratios, a symmetry index between sides and the co-activation ratio of IO over MF served as outcome measures and compared standing on three different surfaces (stable, soft, unsteady). RESULTS: PWH revealed markedly restricted lower extremity joints (P < 0.001), but without any hint of back pain. Neither result revealed significant main or interaction effects of 'group' (P > 0.24). Group-independent analyses showed amplitude ratios (MF: P < 0.05) as well as symmetry indices (MF: P < 0.02) significantly altered by 'surface' in NHC only. Effects of utilizing soft vs. unsteady surfaces were not detectable (P > 0.77). CONCLUSION: Utilizing unstable surfaces does not lead to altered trunk muscle activity in PWH. Differently than expected, a quite similar behaviour of lower trunk muscles in terms of applied indices can be found in PWH and NHC. Ascending alterations of muscle coordination in PWH could not be verified.

Fabrication and characterization of an electrically contacted vapor cell.

We demonstrate the use of electrically contacted vapor cells to switch the transmission of a probe laser. The excitation scheme makes use of electromagnetically induced transparency involving a Rydberg state. The cell fabrication technique involves thin-film-based electric feedthroughs, which are well suited for scaling this concept to many addressable pixels like in flat panel displays.

Ankle muscle activation in people with haemophilia.

Since normative surface EMG (SEMG) values for muscles acting at the knee joint are available for people with haemophilia, increasing interest is noticeable for other joints affected by haemophilic arthropathy. Adequate activity of shank muscles is an important key for appropriate postural control. The aim of this study was to determine differences in muscle activation patterns of lower leg muscles between people with and without haemophilia during upright standing. SEMG of tibialis anterior (TA), fibularis longus (FL), lateral (LG) and medial (MG) heads of gastrocnemius, and soleus (SO) muscles of both sides were recorded in 25 haemophilic patients (H) and 25 non-haemophilic control subjects (C) while standing on even ground. The Gilbert-Score was used to assign sides to major (H-MA) and minor (H-MI) affected ankle joints in H. To normalize the SEMG amplitudes, amplitude ratios (percentage of cumulated activity) were calculated. Compared to controls, TA ratios showed higher and MG reduced levels in both H groups (P < 0.01). In the H-MA subgroup of H, FL also joined the TA behaviour whereas SO had similar activation direction as MG. Although possible descending influences from the knee joints cannot be excluded, this can be interpreted as a compensational mechanism due to the severity of the orthopaedic status of the ankle, which with increasing heaviness is accompanied by reduced plantar flexion capability. However, ankle joint integrity appears to be reduced in H, with TA and MG seeming to play key roles for neuromuscular control of upright posture.

SEMG activation patterns of thigh muscles during upright standing in haemophilic patients.

Although electromyography (EMG) is a common method to evaluate muscle activity, studies utilizing EMG in haemophilic patients are rare. The haemophilic arthropathy, resulting in altered afferent information is expected to cause disturbed activation and inter-muscular coordination patterns in haemophilic subjects. The aim of this study was to determine differences of selected knee muscles between haemophilic patients and non-haemophilic subjects during upright standing. Surface EMG (SEMG) amplitudes of rectus femoris, vastus medialis (VM), vastus lateralis (VL) and biceps femoris (BF) muscles of both sides were measured in 27 haemophilic patients (H) and 26 control subjects (C) while standing on an even surface. Data from both sides were pooled in C, but data of H were subdivided further according to major (H-MA) and minor (H-MI) affected joints. To normalize the data, amplitude ratios (percentage of cumulated activity) were calculated as well. Regardless of whether H-MA or H-MI was compared with C, amplitudes of all extensor muscles reached significantly higher levels in H (P < 0.05). SEMG amplitude ratios also differed between H and C. Independent of subgroup, BF showed significantly reduced activation ratios (P < 0.01). Only the ratios of VM and VL of H-MA could replicate the observed amplitude differences to C (P < 0.05). These findings show that while standing, haemophiliacs maintain the necessary stability demands through increased extensor activities and modulated coordination patterns. Although all thigh muscles of haemophiliacs are characterized by distinct atrophy, increased amplitude levels could be proved for the knee extensor muscles only. Therefore, general atrophy-related effects cannot explain these results.

Mini-collagens in hydra nematocytes.

We have isolated and characterized four collagen-related c-DNA clones (N-COL 1, N-COL 2, N-COL 3, N-COL 4) that are highly expressed in developing nematocytes in hydra. All four c-DNAs as well as their corresponding transcripts are small in size (600-1,000 bp). The deduced amino acid sequences show that they contain a central region consisting of 14 to 16 Gly-X-Y triplets. This region is flanked amino-terminal by a stretch of 14-23 proline residues and carboxy-terminal by a stretch of 6-9 prolines. At the NH2- and COOH-termini are repeated patterns of cysteine residues that are highly conserved between the molecules. A model is proposed which consists of a central stable collagen triple helix of 12-14 nm length from which three 9-22 nm long polyproline II type helices emerge at both ends. Disulfide linkage between cysteine-rich segments in these helices could lead to the formation of oligomeric network structures. Electrophoretic characterization of nematocyst extracts allows resolution of small proline-rich polypeptides that correspond in size to the cloned sequences.

Androgens regulate the dendritic length of mammalian motoneurons in adulthood.

Sex steroid hormones have been thought to alter behaviors in adulthood by changing the activity of neural circuits rather than by inducing major structural changes in these pathways. In a group of androgen-sensitive motoneurons that mediate male copulatory functions, decreases in androgen levels after castration of adult rats produced dramatic structural changes, decreasing both the dendritic length and soma size of these motoneurons. These changes were reversed by androgen replacement. These results imply a surprising degree of synaptic plasticity in adult motoneurons and suggest that normal changes in androgen levels in adulthood are associated with significant alterations in the structure and function of these neurons.

Overexpression of a transporter gene in a multidrug-resistant human lung cancer cell line.

The doxorubicin-selected lung cancer cell line H69AR is resistant to many chemotherapeutic agents. However, like most tumor samples from individuals with this disease, it does not overexpress P-glycoprotein, a transmembrane transport protein that is dependent on adenosine triphosphate (ATP) and is associated with multidrug resistance. Complementary DNA (cDNA) clones corresponding to messenger RNAs (mRNAs) overexpressed in H69AR cells were isolated. One cDNA hybridized to an mRNA of 7.8 to 8.2 kilobases that was 100- to 200-fold more expressed in H69AR cells relative to drug-sensitive parental H69 cells. Overexpression was associated with amplification of the cognate gene located on chromosome 16 at band p13.1. Reversion to drug sensitivity was associated with loss of gene amplification and a marked decrease in mRNA expression. The mRNA encodes a member of the ATP-binding cassette transmembrane transporter superfamily.

Localization of a novel multidrug resistance-associated gene in the HT1080/DR4 and H69AR human tumor cell lines.

Two doxorubicin-selected human tumor cell lines, H69AR and HT1080/DR4, display a multidrug resistance phenotype but do not overexpress P-glycoprotein. Recently, a 6.5-kilobase mRNA encoding a novel member of the ATP-binding cassette superfamily of transport proteins, designated multidrug resistance-associated protein (MRP), has been identified in the H69AR cell line. In the present study, the levels of MRP mRNA were found to be 14-fold higher in HT1080/DR4 cells relative to sensitive HT1080 cells. Southern blotting indicates that gene amplification contributes to the overexpression of MRP in HT1080/DR4 cells. Using a 4-kilobase MRP complementary DNA probe, MRP genes were localized to 2-5 chromosomes bearing homogeneously staining regions and to multiple double minute chromosomes in H69AR cells. Resistant H69AR cells also contained a new der(16) with a structural aberration affecting 16p13.1, the normal cellular locus of the MRP gene. The MRP probe hybridized to two small homogeneously staining regions (hsr) in HT1080/DR4 cells including hsr(7)(p12p15). MRP localization was restricted to the normal cellular locus, 16p13.1, in the parental H69 and HT1080 cells and the drug-sensitive H69PR revertant cells. Our data provide combined evidence that amplification of the MRP gene is associated with the expression of drug resistance in selected solid tumor cell lines.

[Interdisciplinary Assessment Criteria for Rehabilitation after Injuries of the Lower Extremity: A Function-Based Return to Activity Algorithm]. / Interdisziplinäre Beurteilungskriterien für die Rehabilitation nach Verletzungen an der unteren Extremität: Ein funktionsbasierter Return to Activity Algorithmus.

BACKGROUND: In the treatment of patients with lower extremity injuries, a paradigm shift is taking place: Time-dependent concepts are increasingly being replaced by function-based concepts. METHODS: A function-based Return to Activity Algorithm is presented which contains a level classification (I-IV). Qualitative and subsequent quantitative tests are assigned to every level. Within each level, first the respective qualitative test has to be passed before patients are allowed to perform the corresponding quantitative test. Criteria for success are qualitative and quantitative comparisons with the unaffected side. Before entering the next level, both tests have to be successfully passed. The levels are ordered according to increasing demands on the loco-motor system. These demands are adequate stability without impact interaction in sagittal plane for level I, followed by dynamic stability demands for level II. Impacts in frontal plane are added for level III and finally multidirectional impacts have to be compensated at level IV. The time expenditure per level is no more than five minutes. The case of a professional soccer player will serve to exemplify the Return to Activity Algorithm. Following a knee injury, he underwent arthroscopy with ACL reconstruction (patellar tendon) and a partial meniscectomy (lateral and medial). RESULTS: The athlete was able to successfully pass each level and finished his rehabilitation 203 days post injury. He returned to the team training 221 days post injury. 247 days post injury, the athlete completed his first game. CONCLUSION: The Return to Activity Algorithm is able to support the evaluation of the functional status of the loco-motor system after injury or surgery and is furthermore capable of uncovering deficits or asymmetries, which are a proven risk for re-injury. This function-oriented individual approach is able to adequately dose the therapeutic efforts on an individual basis. With this approach, the right timing for a safe return to sports activities can be detected with high certainty.

Response of Cultured Maize Cells to (+)-Abscisic Acid, (-)-Abscisic Acid, and Their Metabolites.

The metabolism and effects of (+)-S- and (-)-R-abscisic acid (ABA) and some metabolites were studied in maize (Zea mays L. cv Black Mexican Sweet) suspension-cultured cells. Time-course studies of metabolite formation were performed in both cells and medium via analytical high-performance liquid chromatography. Metabolites were isolated and identified using physical and chemical methods. At 10 [mu]M concentration and 28[deg] C, (+)-ABA was metabolized within 24 h, yielding natural (-)-phaseic acid [(-)-PA] as the major product. The unnatural enantiomer (-)-ABA was less than 50% metabolized within 24 h and gave primarily (-)-7[prime]-hydroxyABA [(-)-7[prime]-HOABA], together with (+)-PA and ABA glucose ester. The distribution of metabolites in cells and medium was different, reflecting different sites of metabolism and membrane permeabilities of conjugated and nonconjugated metabolites. The results imply that (+)-ABA was oxidized to (-)-PA inside the cell, whereas (-)-ABA was converted to (-)-7[prime]-HOABA at the cell surface. Growth of maize cells was inhibited by both (+)- and (-)-ABA, with only weak contributions from their metabolites. The concentration of (+)-ABA that caused a 50% inhibition of growth of maize cells was approximately 1 [mu]M, whereas that for its metabolite (-)-PA was approximately 50 [mu]M. (-)-ABA was less active than (+)-ABA, with 50% growth inhibition observed at about 10 [mu]M. (-)-7[prime]-HOABA was only weakly active, with 50% inhibition caused by approximately 500 [mu]M. Time-course studies of medium pH indicated that (+)-ABA caused a transient pH increase (+0.3 units) at 6 h after addition that was not observed in controls or in samples treated with (-)-PA. The effect of (-)-ABA on medium Ph was marginal. No racemization at C-1[prime] of (+)-ABA, (-)-ABA, or metabolites was observed during the studies.

The histone deacetylase inhibitor sodium butyrate induces DNA topoisomerase II alpha expression and confers hypersensitivity to etoposide in human leukemic cell lines.

The differentiating agent and histone deacetylase inhibitor, sodium butyrate (NaB), was shown previously to cause a transient, 3-17-fold induction of human DNA topoisomerase II alpha (topo II alpha) gene promoter activity and a 2-fold increase in topo II alpha protein early in monocytic differentiation of HL-60 cells. This observation has now been extended to other short chain fatty acids and aromatic butyrate analogues, and evidence is presented that human topo II alpha promoter induction correlates closely with histone H4 acetylation status. Because increased topo II alpha expression is associated with enhanced efficacy of topo II-poisoning antitumor drugs such as etoposide, the hypothesis tested in this report was whether NaB pretreatment could sensitize HL-60 myeloid leukemia and K562 erythroleukemia cells to etoposide-triggered DNA damage and cell death. A 24-72 h NaB treatment (0.4-0.5 mM) induced topo II alpha 2-2.5-fold in both HL-60 and K562 cells and caused a dose-dependent enhancement of etoposidestimulated, protein-linked DNA complexes in both cell lines. At concentrations with minimal effects on cell cycle kinetics (0.4 mM in HL-60; 0.5 mM in K562), NaB pretreatment also modestly enhanced etoposidetriggered apoptosis in HL-60 cells, as determined morphologically after acridine orange/ethidium bromide staining, and substantially increased K562 growth inhibition and poly(ADP-ribose)polymerase cleavage after etoposide exposure. Therefore, a temporal window may exist whereby a differentiating agent may sensitize experimental leukemias to a cytotoxic antitumor agent. These results indicate that histone deacetylase inhibitors should be investigated for etoposide sensitization of other butyrate-responsive hematopoietic and nonhematopoietic tumor lines in vitro and in vivo.

The primitive metazoan Hydra expresses antistasin, a serine protease inhibitor of vertebrate blood coagulation: cDNA cloning, cellular localisation and developmental regulation.

We have isolated and characterized cDNAs from Hydra which encode antistasin, a potent inhibitor of factor Xa in the vertebrate blood clotting cascade. Hydra antistasin is expressed in gland cells and represents a major class of transcripts from Hydra's head. Sequence analysis revealed that Hydra antistasin contains 6 internal repeats of a 25-26 amino acid sequence with a highly conserved pattern of 6 cysteine and 2 glycine residues identical to that in leech antistasin. Conservation of antistasin in a lower metazoan provides a potential link between the vertebrate and invertebrate coagulation systems.

Morphology of rat spinal motoneurons with normal and hormonally altered specificity.

Potential determinants of motoneuronal morphology were examined by using a sexually dimorphic, steroid-sensitive neuromuscular system in the rat spinal cord. In males, the spinal nucleus of the bulbocavernosus (SNB) innervates the perineal muscles bulbocavernosus (BC) and levator ani (LA), and the dorsolateral nucleus (DLN) innervates the ischiocavernosus muscle (IC). Adult females normally lack these motoneurons and the peripheral targets. Prenatal exposure of females to the androgen dihydrotestosterone propionate (DHTP) partially masculinizes this neuromuscular system and alters moto-neuron-to-muscle specificity, resulting in retained SNB target muscles anomalously innervated by motoneurons in the DLN. Because the morphology of SNB and DLN motoneurons normally differs significantly, the influence of spinal cord location and peripheral target on motoneuron morphology can be directly compared. Injection of cholera toxin conjugated to horseradish peroxidase (CTHRP) into the LA of DHTP-treated females labeled motoneurons predominantly in the SNB. These (SNB-LA) motoneurons in DHTP females were identical in all morphological measures to those of normal males. CTHRP injection into the BC of DHTP females labeled motoneurons in both the SNB and the DLN. SNB-BC motoneurons in DHTP females resembled those of normal males in process number and orientation, but were significantly smaller in dendritic length per motoneuron and in soma size. The DLN motoneurons anomalously projecting to the BC in DHTP females differed significantly from SNB-BC motoneurons in soma size and number and orientation of primary processes. However, these motoneurons were identical in all respects to DLN-IC motoneurons in DHTP females; DLN-IC motoneurons were similar to those of normal males in the orientation of their dendritic arbor, but were significantly smaller in dendritic length, soma size, and number of primary processes. These comparisons make it clear that DHTP selectively affects motoneuronal specificity and morphology in specific motoneuron classes. Further, motoneuronal morphology in the SNB/DLN system appears to be influenced more by spinal cord location than by peripheral target.

Hormonal control of neuron number in sexually dimorphic spinal nuclei of the rat: II. Development of the spinal nucleus of the bulbocavernosus in androgen-insensitive (Tfm) rats.

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus whose development is under the control of steroid hormones. The SNB contains many more motoneurons in adult male rats than in females, and this sex difference is produced by a sexually dimorphic motoneuron death which is regulated by androgens. To study further the role of androgens in the development of sex differences in SNB motoneuron number, we examined SNB development in males with the testicular feminization mutation (Tfm) which renders them insensitive to androgens. Counts of SNB motoneurons perinatally revealed that SNB development in normal male and female King-Holtzman rats was similar to that reported previously for Sprague-Dawley rats; SNB motoneuron number increased from initially low levels at embryonic day 18 through the day before birth, when motoneuron numbers in both sexes were substantially higher than adult levels. After this prenatal increase, motoneuron number declined in both sexes, until by postnatal day 10 motoneuron numbers were in their adult ranges and the sex difference was fully expressed. Females lost more motoneurons than did males during this period, and this loss was due to motoneuron death as revealed by counts of degenerating cells. SNB development in King-Holtzman Tfm males was similar to that of normal males through embryonic day 20, suggesting that androgens may not be necessary for the initial increase in motoneuron numbers in the SNB. Thereafter, SNB motoneuron numbers in Tfm males declined in a female-typical fashion; Tfm males and normal females did not differ at any postnatal age.(ABSTRACT TRUNCATED AT 250 WORDS)

Catalytic inhibition of human DNA topoisomerase IIalpha by hypericin, a naphthodianthrone from St. John's wort (Hypericum perforatum).

St. John's wort (Hypericum perforatum) is the most widely used herbal medicine for the treatment of depression. However, concerns have arisen about the potential of its interaction with other drugs due to the induction of cytochrome P450 isozymes 1A2 and 3A4 by the components hypericin and hyperforin, respectively. Structurally similar natural products are often employed as antitumor agents due to their action as inhibitors of DNA topoisomerases, nuclear enzymes that modify DNA during cellular proliferation. Preliminary findings that hypericin inhibited the DNA relaxation activity of topoisomerase IIalpha (topo II; EC 5.99.1.3) led us to investigate the mechanism of enzyme inhibition. Rather than stabilizing the enzyme in covalent complexes with DNA (cleavage complexes), hypericin inhibited the enzyme prior to DNA cleavage. In vitro assays indicate that hypericin is a potent antagonist of cleavage complex stabilization by the chemotherapeutics etoposide and amsacrine. This antagonism appears to be due to the ability of hypericin to intercalate or distort DNA structure, thereby precluding topo II binding and/or DNA cleavage. Supporting its non-DNA damaging, catalytic inhibition of topo II, hypericin was shown to be equitoxic to both wild-type and amsacrine-resistant HL-60 leukemia cell lines. Moreover, hypericin was incapable of stimulating DNA damage-responsive gene promoters that are activated by etoposide. As with the in vitro topo II assay, antagonism of DNA damage stimulated by 30 microM etoposide was evident in leukemia cells pretreated with 5 microM hypericin. Since many cancer patients experience clinical depression and concomitantly self-medicate with herbal remedies, extracts of St. John's wort should be investigated further for their potential to antagonize topo II-directed chemotherapy regimens.

An unexpected pathway for the metabolic degradation of 1,3-dialkyl-3-acyltriazenes.

In the presence of NADPH, rat liver microsomes catalyzed the degradation of a series of 1,3-dialkyl-3-acyltriazenes, and the extent of the reaction was correlated with compound lipophilicity. In the case of two methylcarbamoyltriazenes, 1-(2-chloroethyl)-3-benzyl-3- (methylcarbamoyl)triazene (CBzM) and 1-(2-chloroethyl)-3-methyl-3-(methylcarbamoyl)triazene (CMM), microsomal metabolites were isolated. Identification of the CBzM metabolites as 1-(2-chloroethyl)-3-benzyl-3-(hydroxymethylcarbamoyl)triazene and 1-(2-chloroethyl-3-benzyl-3-carbamoyltriazine, and the CMM metabolite as 1-(2-chloroethyl)-3-methyl-3-(hydroxymethylcarbamoyl)triazene indicated that the first metabolic step involves hydroxylation of the methylcarbamoyl substituent. Detailed studies of the metabolism of CBzM indicated that the Km for the reaction was 84 microM, and that metabolism was more efficient if microsomes were prepared from male than from female rats. During prolonged incubation, the metabolites of CBzM were also degraded. The degradation of CBzM and its metabolites was inhibited by SKF-525A and metyrapone, suggesting the involvement of a cytochrome P450 isozyme, and supporting the hypothesis that the process is oxidative rather than hydrolytic in both cases. Metabolic oxidation represents an alternative pathway to chemical or enzymatic hydrolysis for the in vivo decomposition of (methylcarbamoyl)triazenes. This mechanism may ultimately explain the antitumor efficacy and low acute toxicity of selected compounds.

Lithium chloride and avoidance of novel places.

Rats were exposed to a distinctive chamber (chamber A, part of a two-chamber apparatus), which was novel for half of the rats but familiar for the other half. Each rat was subsequently injected with lithium chloride or saline. In a test trial conducted 24 hr later, all rats were given a choice between chamber A and a second chamber (B), which was novel for all rats. The main result was that the group made familiar with chamber A and then given lithium showed a significant preference for that side or an avoidance of the novel side, a "spatial neophobia." A second experiment confirmed the spatial neophobia effect and demonstrated that it was not dependent on the particular conditioning procedure used in the first experiment. The spatial neophobia effect was related to similar effects in the taste aversion literature, and to the results of research on lithium-induced decreases in exploratory behavior.

A neuroanatomical correlate of paternal and maternal behavior in the biparental California mouse (Peromyscus californicus).

The medial preoptic area (MPOA) is important in the control of maternal behavior in rodents, and it is sexually dimorphic. This study demonstrates that the MPOA of nonparental virgin male Peromyscus californicus (n = 10) is larger than that of virgin females (n = 9) because of a significantly greater number of neurons in the MPOA of virgin males. However, this sex difference in MPOA volume disappeared when males and females became parents. Soma size increased significantly when females became mothers. These data suggest that maternal behavior may require a structural change in the MPOA, whereas paternal responsivity, which is normally inhibited in virgin males, may require a change in neuronal activity. Furthermore, these results underscore the importance of reproductive status in examination of sexually dimorphic nuclear structures.

Early handling increases hippocampal long-term potentiation in young rats.

The hippocampal formation undergoes major anatomical and physiological changes postnatally, and thus might be expected to be particularly sensitive to early handling effects. Long-term potentiation (LTP) in the hippocampal formation, a form of brain plasticity thought to be important in learning and memory, was examined in young rats following early handling or control treatments. The amplitude of LTP was reliably greater in rats receiving the early handling regime. Possible mechanisms and consequences of enhanced LTP were discussed.

FK506 prevents stroke-induced generation of ceramide and apoptosis signaling.

Ceramide is a key mediator of apoptosis during the cellular stress response which is also involved in stroke-induced death. Transient occlusion of the middle cerebral artery (MCA) in rats led to a strong generation of ceramide as measured in thalamus and entorhinal cortex of the ischemic brain tissue. Enhanced levels of ceramide may be involved in apoptosis signaling following stroke since exogenously added synthetic C2-ceramide increased expression of c-jun and the death-inducing ligands (DILs) CD95-L, TRAIL and TNF-alpha in neuroblastoma cells. DILs in turn mediated death via binding to their respective receptors as concluded from diminished apoptosis upon blocking of the common pathway by dominant negative FADD. C2-ceramide induced both necrosis and apoptosis in a concentration-dependent manner corresponding to the situation present in the ischemic brain. The immunosuppressant FK506 inhibited the release of ceramide, expression of CD95-L and apoptosis in an in vitro and in vivo model for ischemia/reperfusion. These data suggest that ceramide is a crucial initiator of death, e.g., by induction of DILs following stroke.