19/04/10

Если я правильно понял, то весь прикол здесь в том, что плюрипотентные клетки гораздо дольше НЕ ДИФФЕРЕНЦИРУЮТСЯ во взвешенном состоянии (в суспензии). Логично, что для специализации необходимо достаточно твердое окружение клеток того органа, для которого идёт дифференцировка стволовых клеток. Неужели раньше это никто не замечал? Проверить! Видимо, сильно хотелось раздуть результат - надо обязательно прочесть статью и обсудить с коллегами.

ПИ 20/4/10 20:36К

BioTechniques Daily, 04/20/2010

Клетки остаются плюрипотентных дольше в новой клеточной среде

Новые культуральная среда для стволовых клетках продемонстрировала способность сохранять стволовые клетки живыми и недифференцированными дольше.

A new stem cell suspension developed at Hadassah University in Jerusalem, Israel has shown to keep stem cells alive and undifferentiated longer than current methods. The research—led by Benjamin Reubinoff, director of the Hadassah Human Embryonic Stem Cell Research Center and associate professor of obstetrics and gynecology at Hadassah University Medical Center—was published in Nature Biotechnology, and the authors believe that the technique may be useful in producing the large quantities of hESCs needed for clinical applications

"This research is an important step towards the development of an automated controlled process that produces the large quantities of cells needed for transplantaton in millions of patients and for other uses in industry and research," Reubinoff said in a press release.

To keep hESCs pluripotent and alive, they are normally cultured on feeder cells—such as a layer of mouse embryonic fibroblasts—or in clusters on small bead-like materials called microcarriers. But growing stem cells on these substrates is expensive, and can be frustrating since the cultures don’t remain undifferentiated for very long despite having all the necessary growth factors present.

Reubinoff’s team was studying the differentiation of hESC clusters into neural cells and had been using Invitrogen’s Neurobasal medium, which enables the long-term maintenance of the normal phenotype and growth of neuronal cells without a feeder layer. During the study, the researchers noticed that not all the cells differentiated. Suspecting that their choice of medium may have caused this unexpected behavior, they began to focus on how it might promote stem cell growth and inhibit differentiation.

To test their theory, the team customized the Neurobasal medium with commercial serum replacements that promote stem cell growth and inhibit differentiation. They also added proteins that stem cells need to live and grow including extracellular matrix components, neutrotrophins, fibroblast growth factor 2, and activin A. The researchers cultured the hESCs suspended in a regular serum-free medium and in their customized medium. After three weeks, more undifferentiated hESCs were present in their custom medium than in the control medium.

Using the new medium, the team tested three different hESC lines in three cultures. They analyzed the cultures after 7 and 20 weeks using fluorescence-activated cell sorting (FACS), and discovered that on both occasions, 90% of the cells in all three lines remained pluripotent. Furthermore, the medium also allowed the researchers to grow cell lines from abnormal human blastocysts and to direct the differentiation of the cells into neural spheres.

Reubinoff’s team admits that their method isn’t perfect. During their cell passaging step, they lost nearly 3 times as many cells than normal passaging steps in a feeder-layer based culturing method would lose. They will continue to refine the technique to ensure that the cultures yield as many cells as possible for research.

The paper “Derivation, propagation, and controlled differentiation of human embryonic stem cells” was published in the Mar. 28 edition of Nature Biotechnology