Human embryonic stem (ES) cells are commonly derived from early embryos before they implant in the uterus, therefore, ES cells are not only able to grow unlimitedly but can also develop into different cell types, such as cells that form our heart, lungs, nerves, and blood vessels. The ability of human ES cells to grow with no restriction and to become functionally useful cells makes ES cells valuable to treat patients with heart attack, stroke, diabetes, and chronic lung diseases. The ES cells isolated from patients with cardiovascular and pulmonary diseases can also be used to define pathogenic mechanisms of the disease. Among cardiovascular and lung diseases, pulmonary hypertension (i.e., abnormally high blood pressure in the lungs) is a progressive and fatal disease that affects thousands of people annually in the United States. In addition, many patients with chronic obstructive pulmonary disease (COPD), congenital heart diseases, scleroderma, and AIDS also develop pulmonary hypertension, which may cause right heart failure and death. Although a variety of medical and surgical therapeutic strategies have been used clinically to treat patients with pulmonary hypertension, treatment is expensive and ineffective; the mortality of patients with pulmonary hypertension, albeit delayed, is still high. It is therefore urgent to explore the possibility to use stem cell therapy for patients with various forms of pulmonary hypertension, for example patients with primary pulmonary hypertension (PPH), patients with HIV/AIDS-mediated pulmonary hypertension, patients with blood clot-induced pulmonary hypertension, patients with pulmonary hypertension due to lung infection and smoking (e.g., COPD), and patients with pulmonary hypertension due to congenital heart disease. Although the therapeutic potential of stem cells is well recognized and promising, abnormal stem cells can be also be involved in the development of cardiopulmonary diseases. It is therefore important to understand the intrinsic mechanisms involved in stem cell growth and the regulatory mechanisms that guide stem cells to become different types of cells (e.g., blood vessel cells, heart cells, or lung cells). This study is designed to study: a) whether and how a special group of proteins, ion channel proteins, participate in regulating human ES cell growth, b) whether increased or decreased expression/function of these channel proteins affects the ability of human ES cells to become functional cells (e.g., blood vessel cells), and c) whether diseased cells isolated from patients with pulmonary hypertension affect normal human ES cell growth and alter the ability of ES cells to become functional cells (e.g., whether diseased cells can “misguide” normal human ES cells to become malfunctioned or dysfunctional cells). Completion of this project will provide useful information for developing new therapeutic approaches for patients with different forms of pulmonary hypertension.
Statement of Benefit to California:
California is the most populous state in the United States; there are millions of patients suffering from cardiopulmonary diseases in the state. Although it predominantly affects women, pulmonary vascular disease is a disease that affects people in all races; there is no significant difference in terms of incidence among different ethnic groups. Based on the demographic data from the UCSD Medical Center, the patients with pulmonary hypertension who were treated with lung transplantation and pulmonary endarterectomy include 76-78% Caucasians, 7-12% Hispanics, 8-13% African Americans, 3% Asians, and 3% Others. The current medical therapy for patients with pulmonary vascular disease is expensive and often ineffective. Development of more effective therapeutic approaches for pulmonary vascular disease would significantly improve healthcare quality and reduce healthcare expenses in California. The UCSD Medical Center, which is ranked fifth in the nation by US News & Report in the category of Respiratory Disorders, is a major referral center for patients with pulmonary hypertension. Developing stem cell-based therapeutic approaches for patients with pulmonary vascular disease would not only provide healthcare for California patients, but also definitely increase the visibility of the UCSD Medical Center and attract more patients from other states in the country and from other countries to obtain treatment at UCSD, which would financially benefit California. Finally, stem cell research and stem cell-based therapies are still in the budding stage. To become a leading authority in this field will eventually contribute to improvement and enhancement of economy, healthcare, and education in California.
This applicant proposes to investigate the role of Ca2+ and K+ channels in hESC. In the first 2 aims he proposes to define and charaterize ion channels functionally expressed in HESCs and to investigate their role in hESC proliferation, differentiation and/or fate decisions. RT-PCR and single cell RT-PCR and patch clamp will be used to investigate the expression and function of various channels using tchniques well established in this lab. In the third aim, he will explore whether feeder cells isolated from patients with pulmonary hypertension or chronic thrmboembolic disease alter the growth of hESC. In a final aim he proposes to turn patient fiblroblasts into hESC by overexpression of pluripotency genes such as Oct3/4, Sox 2 or Nanog. SIGNIFICANCE AND INNOVATION: The first two aims are descriptive and well within the investigator's expertise but are likely to yield data of interest with respect to the basic biology of hESC. There is no rationale provided for the 3rd aim since there is no evidence or plasible hypothesis for suggesting that hESC have any direct or indirect role in pulmonary hypertension or chronic thromboembolic disease. The evidence cited in the grant relate to more differented progenitor cells derived from the bone marrow/blood or the vessel wall. There is also no rationale for the the 4th aim. The challenge of turning fibroblasts into hESC is perhaps best explored in labs with experience at introducing and expressing transgenes in cultured cells. STRENGTHS: Strengths of this proposal include the combined experience of the research team and the broad array of techniques that will be employed to investigate ion channel expression and function. The survey experiments in aims one and two are likely to yield abundant data on expression of channel subunits and functional properties, but the manipulations described in aims three and four do not really address necessity or sufficiency of channel repertoire either in maintaining pluripotency or in sculpting differentiation. WEAKNESSES: The proposal fails to make a compelling connection between pulmonary vascular disease and hES cell differentiation. The applicants hypothesize that the niche environment of the hypertensive vascular wall may present signals to circulating stem cells in the blood. But these circulating stem cells are not hES cells and it is not clear that hES cells should be expected to respond to these hypothetical signals in the same way as circulating blood stem cells. Overall, the proposal does not do a good job of explaining whether the proposed experiments are mainly meant to provide new information about hES cells or about pulmonary vascular disease; or, whether hES cells are somehow envisioned to have some future therapeutic use in the treatment of vascular disease. The fourth aim, which tests whether expression of transcription factors present in undifferentiated hES cells confers pluripotency onto adult pulmonary vascular fibroblasts, seems particularly diffuse. The application asserts that "it is important to define whether the "misguided" or "dysproliferative" cells isolated from the pulmonary vasculature wall of patients with pulmonary hypertension are able to alter proliferation and differentiation of normal hES cells", but there is no clear statement of why this is important. Enthusiasm for this application would be much higher if it made clear whether the ultimate goal of these experiments is to develop a new therapy for pulmonary disease involving hES cells, to obtain new information about mechanisms that underlie hES cell differentiation, to identify the hypothetical signals emanating from the vascular wall niche, or some other objective. DISCUSSION: There was no further discussion following the reviewers' comments.