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Vita-Salute San Raffaele University

5 ARTICLES PUBLISHED IN JoVE

JoVE Journal

Exploiting Live Imaging to Track Nuclei During Myoblast Differentiation and Fusion

Giorgia Careccia^*1,2, Federica Colombo^*1,3, Mario Tirone¹, Alessandra Agresti¹, Marco E. Bianchi^1,2, Samuel Zambrano^*1,2, Emilie Vénéreau^*1

¹Chromatin Dynamics Unit, Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) San Raffaele Scientific Institute, ²Vita-Salute San Raffaele University, ³Department of Electronics, Information and Bioengineering, Politecnico di Milano

Skeletal muscle differentiation is a highly dynamic process, which particularly relies on nuclear positioning. Here, we describe a method to track nuclei movements by live cell imaging during myoblast differentiation and myotube formation and to perform a quantitative characterization of nuclei dynamics by extracting information from automatic tracking.

Immunology and Infection

Purification and Expansion of Mouse Invariant Natural Killer T Cells for in vitro and in vivo Studies

Gloria Delfanti¹, Alessandra Perini^*1,2, Elisa Zappa^*1,2, Maya Fedeli^1,2

¹Experimental Immunology Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS San Raffaele Scientific Institute, ²Vita-Salute San Raffaele University

We describe a rapid and robust protocol to enrich invariant natural killer T (iNKT) cells from mouse spleen and expand them in vitro to suitable numbers for in vitro and in vivo studies.

Medicine

Fertility Sparing Procedure using Carbon Dioxide Fiber Laser Vaporization of Ovarian Endometrioma

Massimo Candiani¹, Jessica Ottolina¹, Iacopo Tandoi¹, Ludovica Bartiromo¹, Matteo Schimberni¹, Roberta Villanacci¹, Stefano Ferrari¹

¹Gynecological Surgery and Endometriosis Departmental Unit, San Raffaele Scientific Institute

In this protocol, CO₂fiber laser technique is demonstrated for the surgical treatment of ovarian endometriosis, which represents a viable alternative in terms of fertility preservation with the major advantage of not being dependent on the surgeon's skills and personal experience.

Medicine

Laparoscopic Oocyte Retrieval and Cryopreservation during Vaginoplasty for Treatment of Mayer-Rokitansky-Kuster-Hauser Syndrome

Valeria Stella Vanni^*1,2, Alessandra Alteri^*1,2, Lucia De Santis^1,2, Greta Chiara Cermisoni¹, Elisa Rabellotti¹, Diana Delprato¹, Marta Parma¹, Enrico Papaleo¹, Luigi Fedele³, Massimo Candiani^1,2

¹Obstetrics and Gynaecology Unit, IRCCS San Raffaele Scientific Institute, ²Vita-Salute San Raffaele University, ³Department of Obstetrics and Gynecology, Fondazione IRCCS Ca' Granda, Ospedale Maggiore Policlinico

A dedicated team can offer Mayer-Rokitansky-Kuster-Hauser patients the option to perform controlled ovarian stimulation and oocyte cryopreservation at the time of laparoscopic vaginoplasty.

Bioengineering

In Vitro Selection of Engineered Transcriptional Repressors for Targeted Epigenetic Silencing

Alessandro Migliara¹, Martino Alfredo Cappelluti¹, Francesca Giannese², Sara Valsoni¹, Alberto Coglot¹, Ivan Merelli^1,3, Davide Cittaro², Angelo Lombardo^1,4

¹San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS San Raffaele Scientific Institute, ²Center for Omics Sciences, IRCCS San Raffaele Institute, ³Institute for Biomedical Technologies, National Research Council, ⁴Vita-Salute San Raffaele University

Here, we present a protocol for the in vitro selection of engineered transcriptional repressors (ETRs) with high, long-term, stable, on-target silencing efficiency and low genome-wide, off-target activity. This workflow allows for reducing an initial, complex repertoire of candidate ETRs to a short list, suitable for further evaluation in therapeutically relevant settings.