Pollen carbohydrates and water content during development, presentation, and dispersal: a short review

Summary. Pollen accumulates starch reserves during development and the final stage of ripening. Before the anther opens, starch is totally or partially converted to pectins, glucose, fructose, sucrose, and to some unknown polysaccharides. Pollen is exposed to dispersing agents in an arrested developmental state which differs according to pollen water content. Pollen is classified as partially dehydrated or partially hydrated. The final water content may be reached before or after anther opening. Especially during exposure and dispersal, partially dehydrated pollen may interconvert soluble and insoluble reserves, modifying internal turgor pressure and hindering water loss or gain. Partially hydrated pollen is commonly devoid of mechanisms to conserve viability in time but has the advantage of quickly emitting pollen tubes on reaching the stigma. Correspondence and reprints: Department of Environmental Sciences, Siena University, 4 Via P.A. Mattioli, 53100 Siena, Italy.     

Multiplexed expression and screening for recombinant protein production in mammalian cells

Background  

A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit. To this end we have coupled an efficient recombinant protein production system based on transient transfection in HEK-293 EBNA1 (HEK-293E) suspension cells with a dot blot method allowing pre-screening of proteins expressed in cells in a high throughput manner.     

Tomato pollen tube development and carbohydrate fluctuations in the autotrophic phase of growth

Ripe pollen has different soluble and insoluble carbohydrates in variable amounts. Pollen germination and pollen tube growth were studied in a tomato cultivar (Solanum lycopersicum L. cv. Platense) with atypical pollen among tomatoes due to its very low amount or absence of sucrose. In vitro assays were performed using a culture medium without carbohydrates to explore whether there is an autotrophic phase of pollen tube growth, and if there is, describe it, and to analyze the fluctuations of endogenous carbohydrates (soluble carbohydrates, starch, pectins, and callose). Pollen germination was fast (ca. 10 min) and a definite autotrophic phase was observed. Soluble carbohydrates and pectins showed the most substantial changes during this period, even after 10 min. A small amount of callose was observed in the ripe pollen and pollen tubes. Pectins were the most abundant pollen tube wall component. Pollen can be considered starchless; starch was not involved in the autotrophic phase of growth. Other types of substances must be connected with the carbohydrate metabolism, because the fluctuations of the different substances did not follow balanced stoichiometric relationships. Pollen germination and pollen tube elongation was sustained autotrophically, even though sucrose was absent and starch was negligible in pollen grains. The type of pollen reserves and the fast pollen tube formation could be selective advantages in this cultivar.     

Post-pollination prefertilization drops affect germination rates of heterospecific pollen in larch and Douglas-fir

Pollen of larch (Larix?×?marschlinsii) and Douglas-fir (Pseudotsuga menziesii) was used in homospecific and heterospecific crosses. Germination of heterospecific pollen in ovulo was reduced in post-pollination prefertilization drops. This provides evidence of selection against foreign pollen by open-pollinated exposed ovules in these two sister taxa, which share the same type of pollination mechanism. Of the other prezygotic stages in pollen-ovule interactions, uptake of pollen by stigmatic hairs did not show any selection. Pollen tube penetration of the nucellus was similar for hetero- and homospecific pollen tubes, but heterospecific tubes only delivered gametes in one cross. To test for differences in the post-pollination prefertilization drops of each species, drops were gathered and analysed. Glucose and fructose were present in similar amounts in Douglas-fir and larch, while sucrose was found in larch only. Other carbohydrates such as xylose and melezitose were species-specific. In P. menziesii, sucrose is absent due to its conversion to glucose and fructose by apoplastic invertases. In contrast, Larix?×?marschlinsii drops have sucrose because they lack apoplastic invertases. The presence of invertase activity shows that the composition of gymnosperm post-pollination prefertilization drops is not static but dynamic. Drops of these two species also differed in their calcium concentrations.     

A new brace treatment similar for adolescent scoliosis and kyphosis based on restoration of thoracolumbar lordosis. Radiological and subjective clinical results after at least one year of treatment

Study design

A prospective treatment study with a new brace was conducted Objective. To evaluate radiological and subjective clinical results after one year conservative brace treatment with pressure onto lordosis at the thoracolumbar joint in children with scoliosis and kyphosis.

Summary of background data

Conservative brace treatment of adolescent scoliosis is not proven to be effective in terms of lasting correction. Conservative treatment in kyphotic deformities may lead to satisfactory correction. None of the brace or casting techniques is based on sagittal forces only applied at the thoracolumbar spine (TLI= thoracolumbar lordotic intervention). Previously we showed in patients with scoliosis after forced lordosis at the thoracolumbar spine a radiological instantaneous reduction in both coronal curves of double major scoliosis.

Methods

A consecutive series of 91 children with adolescent scoliosis and kyphosis were treated with a modified symmetric 30 degrees Boston brace to ensure only forced lordosis at the thoracolumbar spine. Scoliosis was defined with a Cobb angle of at least one of the curves [greater than or equal to] 25 degrees and kyphosis with or without a curve <25 degrees in the coronal plane. Standing radiographs were made i) at start, ii) in brace at beginning and iii) after one year treatment without brace.

Results

Before treatment start ??in brace?? radiographs showed a strong reduction of the Cobb angles in different curves in kyphosis and scoliosis groups (sagittal n = 5 all p < 0.001, pelvic obliquity p < 0.001). After one year of brace treatment in scoliosis and kyphosis group the measurements on radiographs made without brace revealed an improvement in 3 Cobb angles each.

Conclusion

Conservative treatment using thoracolumbar lordotic intervention in scoliotic and kyphotic deformities in adolescence demonstrates a marked improvement after one year also in clinical and postural criteria. An effect not obtained with current brace techniques.     

P2X7 pre-synaptic receptors in adult rat cerebrocortical nerve terminals: a role in ATP-induced glutamate release

Although growing evidence suggests that extracellular ATP might play roles in the control of astrocyte/neuron crosstalk in the CNS by acting on P2X₇ receptors, it is still unclear whether neuronal functions can be attributed to P2X₇ receptors. In the present paper, we investigate the location, pharmacological profile, and function of P2X₇ receptors on cerebrocortical nerve terminals freshly prepared from adult rats, by measuring glutamate release and calcium accumulation. The preparation chosen (purified synaptosomes) ensures negligible contamination of non-neuronal cells and allows exposure of 'nude' release-regulating pre-synaptic receptors. To confirm the results obtained, we also carried out specific experiments on human embryonic kidney 293 cells which had been stably transfected with rat P2X₇ receptors. Together, our findings suggest that (i) P2X₇ receptors are present in a subpopulation of adult rat cerebrocortical nerve terminals; (ii) P2X₇ receptors are localized on glutamatergic nerve terminals; (iii) P2X₇ receptors play a significant role in ATP-evoked glutamate efflux, which involves Ca²⁺-dependent vesicular release; and (iv) the P2X₇ receptor itself constitutes a significant Ca²⁺-independent mode of exit for glutamate.     

A chromosomal translocation causing multiple abnormalities including open eyelids at birth and glomerulonephritis

We have characterized the phenotype of a mouse with a t(2;13) reciprocal translocation induced by chlorambucil. It results in abnormal eyelid formation as well as a series of neurological, physiological, and immunological abnormalities. This mutant has been termed T(2;13)1Fla/+. T(2;13)1Fla/+ mice exhibit open eyelids at birth, a dilute coat color, hyperactivity, and occasional circling and stargazing activity. At 1-6 months, T(2;13)1Fla/+ mice show signs of immune complex-mediated glomerulonephritis and die prematurely. Additionally, double-stranded DNA autoantibodies have been found in sera of T(2;13)1Fla/+ mice. Cytogenetic analysis situated the translocation breakpoint at the proximal end of Chromosome (chr) 2 at band A2, and on Chr 13 at band A4. The mutant phenotype completely correlated with the presence of the translocation. Additional genetic studies have mapped the mutation and translocation breakpoint to Chr 13 between D13Mit16 and D13Mit64, and to Chr 2 proximal to D2Mit5. By fluorescent in situ hybridization (FISH), the position of this mutation/translocation on Chr 13 has been mapped to a region less than 1cM from D13Mit61.     

Priming adult stem cells by hypoxic pretreatments for applications in regenerative medicine

The efficiency of regenerative medicine can be ameliorated by improving the biological performances of stem cells before their transplantation. Several ex-vivo protocols of non-damaging cell hypoxia have been demonstrated to significantly increase survival, proliferation and post-engraftment differentiation potential of stem cells. The best results for priming cultured stem cells against a following, otherwise lethal, ischemic stress have been obtained with brief intermittent episodes of hypoxia, or anoxia, and reoxygenation in accordance with the extraordinary protection afforded by the conventional maneuver of ischemic preconditioning in severely ischemic organs. These protocols of hypoxic preconditioning can be rather easily reproduced in a laboratory; however, more suitable pharmacological interventions inducing stem cell responses similar to those activated in hypoxia are considered among the most promising solutions for future applications in cell therapy. Here we want to offer an up-to-date review of the molecular mechanisms translating hypoxia into beneficial events for regenerative medicine. To this aim the involvement of epigenetic modifications, microRNAs, and oxidative stress, mainly activated by hypoxia inducible factors, will be discussed. Stem cell adaptation to their natural hypoxic microenvironments (niche) in healthy and neoplastic tissues will be also considered.     

Formation of adrenochrome by bovine cardiac sarcolemma

A sarcolemma preparation from bovine heart was able to promote adrenaline oxidation especially when NADH and NADPH were added. The superoxide anion O(2) was demonstrated to be involved in the activation of adrenochrome production.