Year 2

Prematurity/preterm birth is the leading cause of perinatal morbidity and mortality both in the U.S. and in California. These babies are at increased risk for long-term disabilities, including cerebral palsy, gastrointestinal problems, and vision and hearing loss. Many premature babies also suffer from low birth weight, which not only increases complications in the perinatal period, but also leads to increased cardiovascular disease and diabetes in adulthood. The majority of these perinatal complications result from abnormal development and function of the placenta, a transient organ that forms the interface between mother and baby. Trophoblasts are the primary cells which carry out major placental functions such as establishing blood supply from the mother to the fetus. In this application, we proposed the placenta as a novel target for stem cell therapy and sought to generate human trophoblast stem (TS) cells, which give rise to all subtypes of trophoblasts in the placenta.
During the past year, we have made significant progress in two areas. First, we have learned that p63, a protein we had previously identified as a potential marker of trophoblast proliferation and regeneration, in fact regulates specific properties associated with stemness in trophoblast. We are currently attempting to prolong the lifespan of primary trophoblast in culture by introducing p63, along with other genes, into these cells. In addition, p63 is also expressed early after induction of human embryonic stem cell differentiation into trophoblast, and is maintained under low oxygen conditions, which are known to maintain trophoblast proliferation. We are currently testing the effect of down-regulation of this protein on trophoblast induction of human embryonic stem cells. We believe that this single protein plays a major role in developing and maintaining proliferating trophoblast in culture.
In order to discover additional markers for trophoblast proliferation and regeneration, we have collected samples from placentas of various gestational ages. It is known that trophoblast in early placenta (especially in first trimester) are more proliferative than those in later pregnancy (i.e. third trimester). By looking at total gene expression in placentas across gestation, we hope to identify more markers like p63, which play a role in trophoblast stemness. In addition to collection of placental samples, which include a mixture of trophoblast and other cell types, we have also worked on optimizing isolation of a pure population of trophoblast from placentas of different gestational ages. Aside from looking at gene expression, we are also optimizing culture conditions (including determining the best oxygen tension) for these cells.