Global transcriptome profiling reveals differential regulatory, metabolic and hormonal networks during somatic embryogenesis in Coffea arabica.

BACKGROUND: Somatic embryogenesis (SE) is one of the most promising processes for large-scale dissemination of elite varieties. However, for many plant species, optimizing SE protocols still relies on a trial and error approach. We report the first global scale transcriptome profiling performed at all developmental stages of SE in coffee to unravel the mechanisms that regulate cell fate and totipotency. RESULTS: RNA-seq of 48 samples (12 developmental stages × 4 biological replicates) generated 90 million high quality reads per sample, approximately 74% of which were uniquely mapped to the Arabica genome. First, the statistical analysis of transcript data clearly grouped SE developmental stages into seven important phases (Leaf, Dedifferentiation, Primary callus, Embryogenic callus, Embryogenic cell clusters, Redifferentiation and Embryo) enabling the identification of six key developmental phase switches, which are strategic for the overall biological efficiency of embryo regeneration. Differential gene expression and functional analysis showed that genes encoding transcription factors, stress-related genes, metabolism-related genes and hormone signaling-related genes were significantly enriched. Second, the standard environmental drivers used to control SE, i.e. light, growth regulators and cell density, were clearly perceived at the molecular level at different developmental stages. Third, expression profiles of auxin-related genes, transcription factor-related genes and secondary metabolism-related genes were analyzed during SE. Gene co-expression networks were also inferred. Auxin-related genes were upregulated during dedifferentiation and redifferentiation while transcription factor-related genes were switched on from the embryogenic callus and onward. Secondary metabolism-related genes were switched off during dedifferentiation and switched back on at the onset of redifferentiation. Secondary metabolites and endogenous IAA content were tightly linked with their respective gene expression. Lastly, comparing Arabica embryogenic and non-embryogenic cell transcriptomes enabled the identification of biological processes involved in the acquisition of embryogenic capacity. CONCLUSIONS: The present analysis showed that transcript fingerprints are discriminating signatures of cell fate and are under the direct influence of environmental drivers. A total of 23 molecular candidates were successfully identified overall the 12 developmental stages and can be tested in many plant species to optimize SE protocols in a rational way.

Persistent Spinal Pain Syndrome Type 2 (PSPS-T2), a Social Pain? Advocacy for a Social Gradient of Health Approach to Chronic Pain.

The Social Gradient of Health (SGH), or position in the social hierarchy, is one of the major determinants of health. It influences the development and evolution of many chronic diseases. Chronic pain dramatically affects individual and social condition. Its medico-economic impact is significant and worldwide. Failed Back Surgery Syndrome or Persistent Spinal Pain Syndrome type 2 (PSPS-T2) represents one of its most fascinating and disabling conditions. However, the influence of SGH on PSPS-T2 has been poorly explored. We designed a prospective multicentric study (PREDIBACK study) to assess the SGH prevalence, and to examine its association with medical and psychological variables, in PSPS-T2 patients. This study included 200 patients to determine the SGH association with pain (NPRS), Quality of life (EQ-5D-5L), kinesiophobia (FABQ-Work), catastrophism (CSQ), and functional capacity (ODI). Around 85.3% of PSPS-T2 patients in our study had low SGH. Low SGH patients had a higher FABQ-Work and CSQ-Catastrophizing score than high SGH patients (p < 0.05). High SGH patients have a higher ODI score than low SGH patients (p < 0.10). Our results suggest that SGH is a relevant factor to guide prevention, research, and ultimately intervention in PSPS-T2 patients and could be more widely transposed to chronic pain.

An innovative automated active compound screening system allows high-throughput optimization of somatic embryogenesis in Coffea arabica.

Somatic embryogenesis (SE) faces many challenges in fulfilling the growing demand for elite materials. A high-throughput approach is required to accelerate the optimization of SE protocols by multiplying experimental conditions within a limited time period. For the first time in plant micropropagation, we have developed a miniaturized and automated screening system to meet high-throughput standards. Coffea arabica embryo regeneration, classically achieved in 250-ml Erlenmeyer flasks, was successfully miniaturized in 24-well plates, allowing a volume downscaling factor of 100 and a space saving of 53 cm2/well. Cell clusters were ground and filtered to fit the automated pipetting platform, leading to fast, reproducible and uniform cluster distribution (23.0 ± 5.5 cell clusters/well) and successful regeneration (6.5 ± 2.2 embryos/well). Pilot screening of active compounds on SE was carried out. Compounds belonging to the histone deacetylase inhibitor family were tested for embryo regeneration efficiency. Cells treated with 1 µM Trichostatin A showed a marked 3-fold increase in the number of regenerated embryos. When re-tested in 250-ml flasks, the same enhancement was obtained, thereby validating the miniaturized and automated screening method. These results showed that our screening system is reliable and well suited to screening hundreds of compounds, offering unprecedented perspectives in plant micropropagation.

Unravelling the Metabolic and Hormonal Machinery During Key Steps of Somatic Embryogenesis: A Case Study in Coffee.

Somatic embryogenesis (SE) is one of the most promising processes for large-scale dissemination of elite varieties. However, for many plant species, optimizing SE protocols still relies on a trial-and-error approach. Using coffee as a model plant, we report here the first global analysis of metabolome and hormone dynamics aiming to unravel mechanisms regulating cell fate and totipotency. Sampling from leaf explant dedifferentiation until embryo development covered 15 key stages. An in-depth statistical analysis performed on 104 metabolites revealed that massive re-configuration of metabolic pathways induced SE. During initial dedifferentiation, a sharp decrease in phenolic compounds and caffeine levels was also observed while auxins, cytokinins and ethylene levels were at their highest. Totipotency reached its highest expression during the callus stages when a shut-off in hormonal and metabolic pathways related to sugar and energetic substance hydrolysis was evidenced. Abscisic acid, leucine, maltotriose, myo-inositol, proline, tricarboxylic acid cycle metabolites and zeatin appeared as key metabolic markers of the embryogenic capacity. Combining metabolomics with multiphoton microscopy led to the identification of chlorogenic acids as markers of embryo redifferentiation. The present analysis shows that metabolite fingerprints are signatures of cell fate and represent a starting point for optimizing SE protocols in a rational way.

[Modesty in hospital and in care]. / La pudeur à l'hôpital et dans les soins.

Modesty is the expression of the fear of being seen in a position which the individual considers as strictly private. It is turned towards others and if it is breached, the individual experiences a feeling of shame. The hospital is a place where it is challenged.

Coffee Somatic Embryogenesis: How Did Research, Experience Gained and Innovations Promote the Commercial Propagation of Elite Clones From the Two Cultivated Species?

Since the 1990s, somatic embryogenesis (SE) has enabled the propagation of selected varieties, Arabica F1 hybrid and Robusta clones, originating from the two cultivated coffee species, Coffea arabica and Coffea canephora, respectively. This paper shows how mostly empirical research has led to successful industrial transfers launched in the 2000s in Latin America, Africa, and Asia. Coffee SE can be considered as a model for other woody perennial crops for the following reasons: (i) a high biological efficiency has been demonstrated for propagated varieties at all developmental stages, and (ii) somaclonal variation is understood and mastered thanks to intensive research combining molecular markers and field observations. Coffee SE is also a useful model given the strong economic constraints that are specific to this species. In brief, SE faced four difficulties: (i) the high cost of SE derived plants compared to the cost of seedlings of conventional varieties, (ii) the logistic problems involved in reaching small-scale coffee growers, (iii) the need for certification, and (iv) the lack of solvency among small-scale producers. Nursery activities were professionalized by introducing varietal certification, quality control with regard to horticultural problems and somaclonal variation, and sanitary control for Xylella fastidiosa. In addition, different technology transfers were made to ensure worldwide dissemination of improved F1 Arabica hybrids and Robusta clones. Innovations have been decisive for successful scaling-up and reduction of production costs, such as the development of temporary immersion bioreactors for the mass production of pre-germinated embryos, their direct sowing on horticultural soil, and the propagation of rejuvenated SE plants by rooted mini-cuttings. Today, SE is a powerful tool that is widely used in coffee for biotechnological applications including propagation and genetic transformation. Basic research has recently started taking advantage of optimized SE protocols. Based on -omics methodologies, research aims to decipher the molecular events involved in the key developmental switches of coffee SE. In parallel, a high-throughput screening of active molecules on SE appears to be a promising tool to speed-up the optimization of SE protocols.

Indirect somatic embryogenesis of Theobroma cacao L. in liquid medium and improvement of embryo-to-plantlet conversion rate.

The establishment of cocoa embryogenic cell lines in liquid medium starting from high frequency somatic embryogenesis (HFSE) callus is described. The growth kinetics of the cultures during the multiplication and the expression steps conducted in 250 mL Erlenmeyer flasks were described for three genotypes selected for their agronomical traits (EET95, EET96, and EET103). The glucose and dissolved oxygen concentrations and the absorption of Murashige and Skoog medium macronutrients (nitrate, ammonium, potassium, sulfate, calcium, phosphorus, and magnesium) were monitored. The multiplication of the embryogenic calluses in a medium containing 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) at 1 mg L-1, initiated with an inoculation density of 20 g L-1 of callus, was achieved. The growth rate was characterized by two phases, with the second being concomitant with a depletion of phosphorus and magnesium, and a decrease in the embryogenic potential of the callus. The expression of the callus embryogenic capacity was conducted in an auxin-free medium. The embryo production starting from 1 and 5 g L-1 inoculation densities was compared. When placed in the optimal expression conditions in flasks, 1 g of callus produced 1000 to 1500 embryos within 5 to 7 wk. Finally, two paths for improving the plantlet regenerative capacities of cocoa SE produced in liquid medium were identified. Supplementing the expression medium with myo-inositol used as an osmotic agent at a concentration of 50 g L-1 increased the embryo-to-plantlet conversion rate from 13-16% to 40-48%. A 6-wk culture of the embryos on a maturation medium in Petri dishes optimized their subsequent development into plantlets.

[Different old ages]. / Des vieillesses multiples.

There is no single form of ageing. Some people age in a sort of peaceful and happy continuity of their existence, while others are confronted with illness or bereavements which can weaken them considerably. Ageing therefore takes many different forms. Autonomy, social ties and quality of life are key factors in determining how someone ages.

[When pain cannot be expressed]. / Quand la douleur ne peut se dire.

How pain is felt is a personal and private fact which is almost impossible to measure, which eludes any attempt to grasp and describe it and any desire to convey to another person its intensity and its nature. The challenge is to understand the pain, to assess it and manage it.

[Results and lessons of an innovating chemotherapy-induced nauseas and vomiting management strategy]. / Résultats et enseignements d'une stratégie innovante de prise en charge des nausées et vomissements chimio-induits.

Current chemotherapy-induced nausea and vomiting management guidelines recommend taking into account the emetogenic potential of the chemotherapy employed as well as individual risk factors to such effects. We performed an interventional prospective study to assess the impact of an innovating therapeutic optimization strategy. The latter combines current guidelines application to a specific consultation in order to individualize the treatment. This study included 170 patients and covered a total of 1,746 days of various chemotherapies. Among these patients, 86.5% never vomited and 53.8% never had any nausea or vomiting. These results seem generally better than the ones found in the literature with all kinds of chemotherapies. Regarding them, we have attempted to highlight the determining criteria for a successful antiemetic treatment.

[Reconstructing the face, an identity issue]. / Reconstruire le visage, enjeux d'identité.

The human being invests a physical body, a symbol of different significations and values. When this body is damaged, it is the individual's relationship with the world which is unsettled. The disfigured person is confronted with a loss of his or her identity and with a destabilising social relationship. A transplant is an option which leaves patients with an identity which they and others can find difficult to accept.

[The body and touch in nursing care]. / Le corps et le toucher en soins infirmiers.

The body is intimate, social and cultural. The care relationship is constructed through the body. The skin, a symbolic cover, exposes and protects. The touch between the patient and the caregiver expresses the story of a relationship between two bodies.

[Representation in life sciences (6). Cultural images of the body: Organism and flesh]. / Représentation en sciences du vivant (6). Images culturelles du corps : Entre organisme et chair.

In the social and historical plan, the « body ¼ is not still a datum of evidence. This formulation with a dualist connotation does not appear in all the human societies, certain do not distinguish the man from his flesh. The body is rather a question than an answer, a misleading evidence which reveals a multitude of different representations which assign to it a position determined within the general symbolism of the society, dependent on a social status, on a vision of the world, and inside this last one on a definition of the person. The body is a changeable reality from a society to an other one : the images which define it, the systems of knowledge which clarify its nature, the accomplished performances are strickingly diverse, contradictory. The body is a symbolic construction. It seems to be obvious, but nothing is more imperceptible.

[Pain and the care relationship]. / Douleur et relation de soins.

From one social and cultural condition to another, and depending on their personal history, people do not react in the same way to the same type of injury or disease. Each pain is unique. Making the patient a player in his/her treatment is an essential component of an effective care relationship.