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The process in which locally confined epithelial malignancies progressively evolve into invasive cancers is often promoted by unjamming, a phase transition from a solid-like to a liquid-like state, which occurs in various tissues. Whether this tissue-level mechanical transition impacts phenotypes during carcinoma progression remains unclear. Here we report that the large fluctuations in cell density that accompany unjamming result in repeated mechanical deformations of cells and nuclei. This triggers a cellular mechano-protective mechanism involving an increase in nuclear size and rigidity, heterochromatin redistribution and remodelling of the perinuclear actin architecture into actin rings. The chronic strains and stresses associated with unjamming together with the reduction of Lamin B1 levels eventually result in DNA damage and nuclear envelope ruptures, with the release of cytosolic DNA that activates a cGAS-STING (cyclic GMP-AMP synthase-signalling adaptor stimulator of interferon genes)-dependent cytosolic DNA response gene program. This mechanically driven transcriptional rewiring ultimately alters the cell state, with the emergence of malignant traits, including epithelial-to-mesenchymal plasticity phenotypes and chemoresistance in invasive breast carcinoma.

The hard processing conditions of natural and nature-derived hydrogels, whose potential in cellular applications is well established, strongly limit their employment in the realization of advanced systems for guiding cells behavior. In the first part of the thesis an overview on the main 3D micro-fabrication techniques to produce instructive platforms for cell engineering application is provided with special emphasis on the principles and main uses of the direct laser writing two-photon polymerization (DLW 2PP) process. The application of this technique to the fabrication of synthetic, natural and natural derived hydrogel 3D structures is presented, highlighting its current potentialities and limits. Special attention is devoted to the versatility of a biomaterial like gelatin and its broad spectrum of applications, as this is the core subject of the thesis which aims to further expand gelatin use by proposing new approaches to develop innovative platforms for specific cell engineering applications. In Chapter 2 we defined the composition of a gelatin-based photoresist to fabricate complex and high resolute microstructures with our 3D lithography system (Nanoscribe Professional GT). We first synthetized and characterized an acrylate gelatin which was mixed with an azobenzene based crosslinker and with a photoinitiator compatible with our system specifications. Then, the fabrication process was optimized in terms of process parameters and writing strategy. Moreover we proved the fundamental role of the azobenzene in the fabrication of stable structures. Finally, we characterized the polymerized gelatin showing physical properties compatible with many cellular applications. In Chapter 3 we developed a gelatin-based platform for the mechanical stimulation of cells; to this end we studied a strategy in which cells were first positioned and then exposed to mechanical stresses. To control cell positioning we used the Nanoscribe to introduce a novel topographic signal that strongly affected the cell adhesion process. Cells were mechanically stimulated by the light induced microstructures deformation due to the azobenzene crosslinker isomerization. As light source we employed the laser of a confocal microscope to promote a significant material photo-deformation. To get an in-depth comprehension of the photo-deformation mechanical properties of the material were analysed after light exposure. Adopting this strategy we were then able to selectively deform living cells down to the single cell level. In Chapter 4 we exploited same gelatin as instructive building blocks designed to guide, with specific topographic patterns, the production of anisotropic oriented micro-tissue for tissue engineering applications. To this endwe used the Nanoscribe as a rapid prototyping technique to test the efficiency of the aforementioned topographic signal. Once demonstrated the importance of the topography in determining the micro-tissue orientation, we tried to build up a droplet microfluidic device for the massive production of tubular gelatin patterned emulsions. More specifically, to pattern the gelatin surface, we performed an in-chip 3D lithography process to insert specific 3D extrusion micro-heads strategically positioned into the device. The large scale production of the gelatin blocks by exploiting such microfluidic device is under development.

Cinara (Cinara) palaestinensis palaestinensis Material examined. Italy, Tuscany, Florence, Bagno a Ripoli, loc. Terra Bianca, m 300 asl (43 �� 45 ��� N, 11 �� 21 ���E), Pinus halepensis, 5 oviparous females, 3 males, 06.xii.2015, 10.xii.2015, 12.xii. 2015, leg. F. Binazzi (A.B. coll., C 10 / 928, 929, 930); Italy, Umbria, Perugia, Vallo di Nera, loc. Geppa, m 500 asl (42 �� 45 ��� N, 12 �� 50 ��� E), P. halepensis, 2 oviparous females, 1 male, 13.xii. 2015, leg. F. Binazzi (A.B. coll., C 10 / 931, 932). Oviparous female (from seven specimens; Figs 1���2, Tab. 1). Small- to medium-sized body, 2.27���2.86 mm long, bronzy in colour when alive, with four rows of roundish intersegmental sclerites on dorsal abdomen. Siphuncular cones dark brown and small, with apical flange, their basal diameter 0.117���0.147 mm long. Head, pro- and mesothorax sclerotized. Mesosternal tubercle not prominent with 0���1 hairs. Metathorax and first abdominal tergite with two spinal sclerotized patches. Abdominal tergites 2 nd to 7 th without sclerotizations and the same absence in tergite 8 th which at most has basal sclerites of the setae. Cauda dark brown only on the external rim. Genital plate large and light colored. Antennae whitish with only segments I and II brown, while the distal end of III and IV, the distal third of V and the whole VI are pale brown. Rostrum reaches the fourth abdominal segment and has brown-blackish apical segments. Legs dark brown, slightly paler on proximal half of femora and basal third or half of tibiae except for a blackish part close to the tibial joints. The hind tibiae are not thickened and lack scent glands. Tarsi brown, with the same colour as the distal parts of tibiae. Antennal hairs rather hyaline, those on segment III up to 0.054 mm long, the longest 1.2���1.3 times the diameter of the segment at the insertion point. Abdominal tergites 1 st to 6 th with scarce, translucent short hairs, up to 0.050 mm long while those on tergite 7 th reach 0.072 mm of length; tergite 8 th with 8���10 setae; genital plate with 60���80 short hairs; fourth rostral segment with 6���8 accessory hairs; siphuncular cones with 0.048���0.060 mm long hairs; hind tibiae with pale brown hairs, 0.070���0.090 mm long, the longest on it 1.0��� 1.2 times as long as the diameter at the insertion point of the same hair on the tibia. The oviparous females show the same pigmentation of body and appendages as the aptera vivipara. However, the ovipara has shorter hind tibiae and smaller siphuncular cones than the apterous vivipara. Also the antennae are shorter, especially antennal segments, III and V. Another difference is the shorter first and second hind tarsal segments. The lengths of the body hairs are about the same, except that those on the antennae are a little shorter. As in many other aphids, the hairs of the genital plate are much more numerous in the ovipara than in the vivipara (data for comparison were obtained from 5 apterous viviparous females collected in the same localities and on the same dates as the oviparae) (Tab. 2). Apterous male (from four specimens; Fig. 3, Tab. 1). Small, narrow body, 1.68���2.27 mm long, bronzy in color when alive, with four rows of roundish intersegmental muscle-attached plates on dorsal abdomen. Siphuncular cones brown and small, with apical flange and one circular row of hairs, their basal diameter 0.072���0.120 mm long. Head, pro- and mesothorax sclerotized. Mesosternal tubercle prominent with 1���4 hairs. Metathorax and first abdominal tergite with two spinal sclerotized patches. Abdominal tergites 2 nd to 7 th without sclerotizations. Tergite 8 th with a narrow interrupted pale brown crossband with 8���10 hairs. Cauda dark brown with few hairs up to 0.090 mm long. Phallus blackish with dark brown sclerotized valvae covered with short dark setae and with dark brown lobed paramers bearing numerous longer setae up to 0.080 mm long. Genital plate bilobed and pale brown with about 30 hairs on each side, up to 0.080 mm long. Antennae rather well pigmented with segments I and II dark brown and with distal end of III, the distal half of IV and the whole V and VI brown. Rostrum reaches the fourth abdominal segment and has dark brown apical segments. Legs dark brown with slightly paler proximal half of femora and basal third or half of tibiae except for a blackish part close to the tibial joints. Tarsi brown, with the same pigmentation as the distal part of tibiae. Antennal hairs hyaline, those on segment III up to 0.0 72 mm long, the longest 1.5 times the diameter of the segment at the insertion point. Abdominal tergites 1 st to 7 th with scarce, translucent short hairs, 6���8 in one row on each segment, up to 0.075 mm long; tergite 8 th with 8���10 setae; fourth rostral segment with 6 accessory hairs; siphuncular cones with hairs 0.042���0.060 mm long; hind tibiae with rather hyaline hairs, 0.070���0.090 mm long, the longest on it, 1.5���1.7 times as long as the diameter at the insertion point of the same hair on the tibia. Ecological notes. The first and northernmost P. halepensis stand is located in hilly areas of Tuscany ranging from 300 to 400 m asl. This site is characterized by calcareous soil where pines were planted in the first half of the 20 th century. In these conditions, the Aleppo pine populations, associated with some nuclei of Italian stone pine (P. pinea) and maritime pine (P. pinaster), have become naturalized. The second stand is located on steep slopes in Nerina Valley (Umbria). This area, ranging from 500 to 700 m asl, is characterized by a similar calcareous soil but with frequent marly or siliceous interbeds (Catorci & Cesaretti 2016). In this site P. halepensis is assumed to be native or introduced in ancient times (Fenaroli & Gambi 1976)., Published as part of Binazzi, Francesco, Pennacchio, Fabrizio, Peverieri, Giuseppino Sabbatini & Roversi, Pio Federico, 2016, The sexuales of Cinara palaestinensis Hille Ris Lambers, the Aleppo pine aphid (Hemiptera: Aphididae Lachninae), pp. 417-424 in Zootaxa 4137 (3) on pages 418-421, DOI: 10.11646/zootaxa.4137.3.8, http://zenodo.org/record/271933, {"references":["Catorci, A. & Cesaretti, S. (2016) Parco fluviale del Nera. Analisi del sistema naturale, 44 pp. Available from http: // www. parcodelnera. it / wp-content / uploads / Relazione _ sistema _ naturale. pdf (accessed 23 Mar. 2016)","Fenaroli, L. & Gambi, G. (1976) Alberi, Dendroflora Italica. Museo Tridentino di Scienze Naturali, Trento, 717 pp."]}

Binazzi, Francesco, Pennacchio, Fabrizio, Peverieri, Giuseppino Sabbatini, Roversi, Pio Federico (2016): The sexuales of Cinara palaestinensis Hille Ris Lambers, the Aleppo pine aphid (Hemiptera: Aphididae Lachninae). Zootaxa 4137 (3): 417-424, DOI: http://doi.org/10.11646/zootaxa.4137.3.8

Mazza, Giuseppe, Pennacchio, Fabrizio, Gargani, Elisabetta, Franceschini, Italo, Roversi, Pio Federico, Cianferoni, Fabio (2014): First report of Ricania speculum (Walker, 1851) in Europe (Hemiptera: Fulgoromorpha: Ricaniidae). Zootaxa 3861 (3): 297-300, DOI: http://dx.doi.org/10.11646/zootaxa.3861.3.7