Bilateral Strength Deficit Is Not Neural in Origin; Rather Due to Dynamometer Mechanical Configuration

During maximal contractions, the sum of forces exerted by homonymous muscles unilaterally is typically higher than the sum of forces exerted by the same muscles bilaterally. However, the underlying mechanism(s) of this phenomenon, which is known as the bilateral strength deficit, remain equivocal. One potential factor that has received minimal attention is the contribution of body adjustments to bilateral and unilateral force production. The purpose of this study was to evaluate the plantar-flexors in an innovative dynamometer that permitted the influence of torque from body adjustments to be adapted. Participants were identically positioned between two setup configurations where torques generated from body adjustments were included within the net ankle torque (locked-unit) or independent of the ankle (open-unit). Twenty healthy adult males performed unilateral and bilateral maximal voluntary isometric plantar-flexion contractions using the dynamometer in the open and locked-unit mechanical configurations. While there was a significant bilateral strength deficit in the locked-unit (p = 0.01), it was not evident in the open-unit (p = 0.07). In the locked-unit, unilateral torque was greater than in the open-unit (p<0.001) and this was due to an additional torque from the body since the electromyographic activity of the agonist muscles did not differ between the two setups (p>0.05). This study revealed that the mechanical configuration of the dynamometer and then the body adjustments caused the observation of a bilateral strength deficit.     

Desensitization of the Neurokinin 1 Receptor Is Mediated by the Receptor Carboxy-Terminal Region, but Is Not Caused by Receptor Internalization

Abstract: The carboxy-terminal cytoplasmic regions of the rat neurokinin 1 (substance P) and neurokinin 2 (neurokinin A) receptors have been exchanged to determine if this region of the neurokinin 1 receptor is involved in its desensitization. When expressed at similar levels in stably transfected Chinese hamster ovary (CHO) cell lines, receptors containing the carboxy-terminal region of the neurokinin 1 receptor desensitized significantly more (as measured by reduction of the inositol 1,4,5-trisphosphate response) when preexposed for 1 min to 1 µ M neurokinin, indicating a role for the carboxy-terminal region of the neurokinin 1 receptor in its desensitization. Measurement of receptor internalization using radiolabeled neurokinins (0.3 n M ) indicated that ∼75–80% of the receptors were internalized in each cell line after 10 min at 37°C, with no observable correlation between neurokinin receptor desensitization and internalization. Measurement of loss of receptor surface sites for cell lines CHO NK1 and CHO NK1NK2 following exposure to 1 µ M substance P also indicated no obvious relationship between the percent desensitization and percent of receptors internalized. Also, two inhibitors of neurokinin 1 receptor internalization, phenylarsine oxide and hyperosmolar sucrose, did not inhibit neurokinin 1 receptor desensitization. The protein kinase inhibitors Ro 31-8220, staurosporine, and Zn²⁺ had no effect on neurokinin 1 receptor desensitization, indicating that the kinases affected by these agents are not rate-limiting in neurokinin 1 receptor desensitization in this system.     

Force Production by a Bundle of Growing Actin Filaments Is Limited by Its Mechanical Properties

Bundles of actin filaments are central to a large variety of cellular structures such as filopodia, stress fibers, cytokinetic rings, and focal adhesions. The mechanical properties of these bundles are critical for proper force transmission and force bearing. Previous mathematical modeling efforts have focused on bundles’ rigidity and shape. However, it remains unknown how bundle length and buckling are controlled by external physical factors. In this work, we present a biophysical model for dynamic bundles of actin filaments submitted to an external load. In combination with in vitro motility assays of beads coated with formins, our model allowed us to characterize conditions for bead movement and bundle buckling. From the deformation profiles, we determined key biophysical properties of tethered actin bundles such as their rigidity and filament density.     

Reactive Oxygen Species Production in Wheat Roots Is Not Linked with Changes in H+ Fluxes During Acidic and Aluminium Stresses

Aluminium stress induces peroxidation of lipids in the plasma membrane, the effect akin to that caused by reactive oxygen species (ROS). ROS have recently been proposed as regulators of redox-dependent ion transport across the plasma membrane during biotic and abiotic stresses, thus contributing to the plant defence system. The aim of this study was to discover whether ROS production is linked to redox-dependent H⁺ transport system located at the plasma membranes of two near-isogenic lines of wheat (Triticum aestivum L., ET8 = Al-resistant, ES8 = Al-sensitive).The activities of NADPH-dependent ROS synthase and SOD were increased in both wheat lines 15 and 30 min after Al treatments. However, the ROS production was also increased under acidic stress. There was no difference between the two wheat lines in the root-cell plasma membrane capacity to efflux H⁺ in response to potassium ferricyanide after Al and acidic treatments. In ET8, both stresses led to increases in ROS production and H⁺ influx.ROS production in wheat seedlings was activated primarily by low pH exposure rather than by the Al stress. ROS production and breakdown in wheat seedlings under Al and acidic stresses appear to be linked to the intracellular metabolic changes rather than to the increased activity of plasma membrane-based NADPH-dependent ROS synthase.Key Words: ion fluxes, reactive oxygen species (ROS), redox system, superoxide dismutase (SOD), Triticum aestivum L., wheat     

Triggering Dectin-1-Pathway Alone Is Not Sufficient to Induce Cytokine Production by Murine Macrophages

β-glucans (BG) are abundant polysaccharides of the Saccharomyces cerevisiae cell wall (Sc CW), an industry byproduct. They have immuno-stimulatory properties upon engagement of dectin-1 (Clec7a), their main receptor on particular immune cells, and they actually become of great interest because of their preventive or therapeutic potentials. Zymosan, a crude extract of Sc CW was studied as a prototypic BG, despite its miscellaneous PAMPs content. Here, we examined the response of murine wild type or Clec7a^-/- bone marrow-derived macrophages (BMDM) to products with increasing BG content (15, 65 or 75%) and compared their effects with those of other dectin-1 ligands. The enrichment process removed TLR ligands while preserving dectin-1 activity. The most enriched extracts have very low NFκB activity and triggered low amounts of cytokine production in contrast with crude products like zymosan and BG15. Furthermore, MyD88^-/- BMDM did not produce TNFα in response to crude Sc CW extracts, whereas their response to BG-enriched extracts was unaffected, suggesting that BG alone are not able to initiate cytokine secretion. Although Sc CW-derived BG stimulated the late and strong expression of Csf2 in a dectin-1-dependent manner, they remain poor inducers of chemokine and cytokine production in murine macrophages.     

Immune-Mediated Competition in Rodent Malaria Is Most Likely Caused by Induced Changes in Innate Immune Clearance of Merozoites

Malarial infections are often genetically diverse, leading to competitive interactions between parasites. A quantitative understanding of the competition between strains is essential to understand a wide range of issues, including the evolution of virulence and drug resistance. In this study, we use dynamical-model based Bayesian inference to investigate the cause of competitive suppression of an avirulent clone of Plasmodium chabaudi (AS) by a virulent clone (AJ) in immuno-deficient and competent mice. We test whether competitive suppression is caused by clone-specific differences in one or more of the following processes: adaptive immune clearance of merozoites and parasitised red blood cells (RBCs), background loss of merozoites and parasitised RBCs, RBC age preference, RBC infection rate, burst size, and within-RBC interference. These processes were parameterised in dynamical mathematical models and fitted to experimental data. We found that just one parameter , the ratio of background loss rate of merozoites to invasion rate of mature RBCs, needed to be clone-specific to predict the data. Interestingly, was found to be the same for both clones in single-clone infections, but different between the clones in mixed infections. The size of this difference was largest in immuno-competent mice and smallest in immuno-deficient mice. This explains why competitive suppression was alleviated in immuno-deficient mice. We found that competitive suppression acts early in infection, even before the day of peak parasitaemia. These results lead us to argue that the innate immune response clearing merozoites is the most likely, but not necessarily the only, mediator of competitive interactions between virulent and avirulent clones. Moreover, in mixed infections we predict there to be an interaction between the clones and the innate immune response which induces changes in the strength of its clearance of merozoites. What this interaction is unknown, but future refinement of the model, challenged with other datasets, may lead to its discovery.     

Thermomorphogenic and Photomorphogenic Control of Stem Elongation in Fuchsia Is Not Mediated by Changes in Responsiveness to Gibberellins

Stem elongation in Fuchsia × hybrida was influenced by cultivation at different day and night temperatures or in different light qualities. Internode elongation of plants grown at a day (25°C) to night (15°C) temperature difference (DIF+10) in white light was almost twofold that of plants grown at the opposite temperature regime (DIF−10). Orange light resulted in a threefold stimulation of internode elongation compared with white light DIF−10. Surprisingly, internode elongation in orange light was similar for plants grown at DIF−10 and DIF+10. Flower development was accelerated at DIF−10 compared with DIF+10 in both white and orange light. To examine whether the effects of DIF and light quality on shoot elongation were related to changes in gibberellin metabolism or plant sensitivity to gibberellins (GAs), the stem elongation responses of paclobutrazol-treated plants to applied gibberellins were determined. In the absence of applied gibberellins paclobutrazol (>0.32 μmol plant⁻¹) strongly retarded shoot elongation. This inhibition was nullified by the application of about 10–32 nmol of GA₁, GA₄, GA₉, GA₁₅, GA₁₉, GA₂₀, GA₂₄, or GA₄₄. The results are discussed in relation to possible effects of DIF and light quality on endogenous gibberellin levels and gibberellin sensitivity of fuchsia and their effects on stem elongation. Received October 4, 1997; accepted December 17, 1997     

Tyrosinase Protein Is Expressed Also in Some Neural Crest Derived Cells Which Are Not Melanocytes

Some neural crest cells give rise to pigment cells in early ontogenesis. We tested here whether tyrosinase-a key enzyme in melanogenesis—was present in some nonpigment neural crest derivatives in adult hamsters. Interestingly enough, inactive tyrosinase protein was detected, using indirect immunofluorescence, in the satellite cells of spinal ganglia and Schwann cells of sciatic and facial nerves in normal adult animals. The results of cell blotting from spinal ganglia were similar to the fluorescence findings. Thus, our results seem to support the hypothesis that Schwann cells, satellite cells of spinal ganglia, and melanocytes may be more intimately related developmentally than other neural-crest-derived cells. Moreover, since we detected tyrosinase protein in cells which normally do not produce melanosomes, it could be deduced that, during the melanocyte's differentiation from its cell precursor, the expression of tyrosinase protein might precede the point when melanosomes begin to differentiate from known cytoplasmic structures.     

Time Course and Regional Basis of Pb-Induced Changes in MK-801 Binding: Reversal by Chronic Treatment with the Dopamine Agonist Apomorphine but Not the D1 Agonist SKF-82958

Abstract: In the present study we attempted to further define the time course and regional specificity of lead (Pb)-induced changes in the NMDA receptor complex and the influence of dopaminergic system modulations on these changes. Autoradiographic measurements of alterations in MK-801 binding, as evaluated under four different activation conditions (none, spermidine, glycine, or maximal activation), were performed in medial frontal cortex, dorsal striatum, and nucleus accumbens of male rats after 2 weeks or 8 months of chronic postweaning (from 21 days of age on) exposure to 0, 50, or 150 ppm Pb acetate in drinking water. The 8-month groups also received chronic intermittent intraperitoneal injections of saline, or of the dopamine (DA) agonist apomorphine or the D₁ agonist SKF-82958 2–3 times per week beginning at 60 days of age. Two weeks of 50 ppm Pb exposure resulted in small but significant increases in MK-801 binding under conditions of glycine or spermidine activation, whereas decreases were observed in response to 150 ppm under conditions of no or maximal activation in all regions. After 8 months of Pb, concentration-dependent decreases in MK-801 binding were observed across regions under all activation conditions. These effects were noted at blood Pb concentrations averaging as low as 16 µg/dl. Pb-induced decreases in MK-801 binding were either partially or fully reversed by chronic intermittent treatment with the DA agonist apomorphine but not by the D₁ agonist SKF-82958, implicating D₂-based mechanisms in this reversal. Combined findings from this and previous studies based on this exposure protocol indicate a Pb-induced pattern of widespread hypoglutamatergic function accompanied by increased DA function in mesolimbic systems, a pattern of changes reminiscent of those proposed to underlie schizophrenia. Such findings suggest that Pb exposure, even at current environmental levels, could be a risk factor for behavioral and/or neurological disturbances arising from imbalances of glutamate/dopamine function in mesocorticolimbic systems.     

Autosomal-Dominant Multiple Pterygium Syndrome Is Caused by Mutations in MYH3

Multiple pterygium syndrome (MPS) is a phenotypically and genetically heterogeneous group of rare Mendelian conditions characterized by multiple pterygia, scoliosis, and congenital contractures of the limbs. MPS typically segregates as an autosomal-recessive disorder, but rare instances of autosomal-dominant transmission have been reported. Whereas several mutations causing recessive MPS have been identified, the genetic basis of dominant MPS remains unknown. We identified four families affected by dominantly transmitted MPS characterized by pterygia, camptodactyly of the hands, vertebral fusions, and scoliosis. Exome sequencing identified predicted protein-altering mutations in embryonic myosin heavy chain (MYH3) in three families. MYH3 mutations underlie distal arthrogryposis types 1, 2A, and 2B, but all mutations reported to date occur in the head and neck domains. In contrast, two of the mutations found to cause MPS in this study occurred in the tail domain. The phenotypic overlap among persons with MPS, coupled with physical findings distinct from other conditions caused by mutations in MYH3, suggests that the developmental mechanism underlying MPS differs from that of other conditions and/or that certain functions of embryonic myosin might be perturbed by disruption of specific residues and/or domains. Moreover, the vertebral fusions in persons with MPS, coupled with evidence of MYH3 expression in bone, suggest that embryonic myosin plays a role in skeletal development.     

p53 Regulation by TRP2 Is Not Pervasive in Melanoma

p53 is a central tumor suppressor protein and its inhibition is believed to be a prerequisite for cancer development. In approximately 50% of all malignancies this is achieved by inactivating mutations in the p53 gene. However, in several cancer entities, including melanoma, p53 mutations are rare. It has been recently proposed that tyrosinase related protein 2 (TRP2), a protein involved in melanin synthesis, may act as suppressor of the p53 pathway in melanoma. To scrutinize this notion we analyzed p53 and TRP2 expression by immunohistochemistry in 172 melanoma tissues and did not find any correlation. Furthermore, we applied three different TRP2 shRNAs to five melanoma cell lines and could not observe a target specific effect of the TRP2 knockdown on either p53 expression nor p53 reporter gene activity. Likewise, ectopic expression of TRP2 in a TRP2 negative melanoma cell line had no impact on p53 expression. In conclusion our data suggest that p53 repression critically controlled by TRP2 is not a general event in melanoma.     

Allozyme Segregation Ratios in the Interspecific Cross CUCURBITA MAXIMAxC. ECUADORENSIS Suggest That Hybrid Breakdown Is Not Caused by Minor Alterations in Chromosome Structure

The parentals of the interspecific cross Cucurbita maxima x C. ecuadorensis had different isozyme phenotypes for 12 enzyme systems. Characterization of the systems demonstrated that the expression and intracellular distribution of the isozymes were similar to those in other plant taxa; however, a considerable number of duplicate loci were identified, indicative of a polyploid ancestry for Cucurbita. Genetic analysis provided evidence for 20 loci segregating in F₂ and backcross populations. Five linkage groups were identified, consisting of the loci Aat-mb – – Mdh-m2; Gal-1 – – Gal-2; Aat-p2 – – Gpi-c2; Acp-1 – – Pgm-c2 – – Pgm-p; and Est-1 – – Tpi-c2. Significant deviations from Mendelian segregation ratios were observed in 14% of the data sets for individual loci. However, these instances were scattered among the loci, no single locus consistently displaying skewed ratios. Recombination frequencies between linked loci were similar to those observed in intraspecific crosses, and the ratio of heterozygous to homozygous genotypes in backcross populations was very close to one. These results suggest that small differences in chromosome structure were not the major cause of the loss of fertility observed in F₂ and backcross populations.     

Changes in Antigenic Nature of Lymphocytes Caused by Common Viruses

Healthy human lymphocytes were incubated in the presence of influenza A2/Singapore/57, herpes simplex type 1, or adenovirus type 2. After two days the cultures were inactivated by irradiation. Fresh lymphocytes taken from the same donor were then found to react to the virus-treated cells in short-term cultures. We suggest that this reactivity is due to a change in the surface characteristics of the lymphocytes brought about by the presence of the virus. This may account for anomalous reactions in mixed lymphocyte cultures, and a similar effect in vivo might cause accelerated graft rejection.     

Sex Ratio Distortion Caused by Meiotic Drive in a Mosquito, Culex pipiens L

A genetic factor, distorter (d), has been discovered that upsets the normal sex ratio of 1:1 and results in a large excess of males in Culex pipiens. The effect can be explained by a sex-linked, recessive gene. Males homozygous for the gene (Md/md) produce few female offspring; the effect is not due to postzygotic mortality. During the first meiotic division in spermatogenesis, the shortest chromosome pair, which, according to Jost and Laven (1971), is associated with sex determination, can be seen to be abnormal. In a high proportion of spermatocysts, one of the dyads of the shortest bivalent fragments, and the pieces are distributed irregularly to the daughter cells. It is believed that the female-determining chromosomes fragment. This would give rise to an excess of male-determining sperm. The possible usefulness of this factor for control or for experimental purposes is discussed.     

Melanism in Peromyscus Is Caused by Independent Mutations in Agouti

Identifying the molecular basis of phenotypes that have evolved independently can provide insight into the ways genetic and developmental constraints influence the maintenance of phenotypic diversity. Melanic (darkly pigmented) phenotypes in mammals provide a potent system in which to study the genetic basis of naturally occurring mutant phenotypes because melanism occurs in many mammals, and the mammalian pigmentation pathway is well understood. Spontaneous alleles of a few key pigmentation loci are known to cause melanism in domestic or laboratory populations of mammals, but in natural populations, mutations at one gene, the melanocortin-1 receptor (Mc1r), have been implicated in the vast majority of cases, possibly due to its minimal pleiotropic effects. To investigate whether mutations in this or other genes cause melanism in the wild, we investigated the genetic basis of melanism in the rodent genus Peromyscus, in which melanic mice have been reported in several populations. We focused on two genes known to cause melanism in other taxa, Mc1r and its antagonist, the agouti signaling protein (Agouti). While variation in the Mc1r coding region does not correlate with melanism in any population, in a New Hampshire population, we find that a 125-kb deletion, which includes the upstream regulatory region and exons 1 and 2 of Agouti, results in a loss of Agouti expression and is perfectly associated with melanic color. In a second population from Alaska, we find that a premature stop codon in exon 3 of Agouti is associated with a similar melanic phenotype. These results show that melanism has evolved independently in these populations through mutations in the same gene, and suggest that melanism produced by mutations in genes other than Mc1r may be more common than previously thought.