Skeletal system plays a crucial role in our life. Skeletal diseases and disorders unlike cancer, are not fatal, but affect the quality of our life. Cell-based therapeutic strategies to generate targeted desired cell types for repair or replacement of damaged skeletal tissues are ideal regenerative medicines. Because of the heterogeneous cell types generated from embryonic and mesenchymal stem cells, the ability of progenitor population to differentiate into a target cell type appear to be a better alternative for tissue regeneration. Osteo-chondroprogenitors uniquely co-expressing Sox9 and Runx2 with dual differentiation potential to become chondrocytes and osteoblasts is a progenitor cell which is suitable for cell based therapy of bone disease. Therefore, developing effective strategies to generate sufficient quantities of osteo-chondroprogenitors are essential. Toward this, we took advantage of two lineage conversion approaches. The first strategy was to interrogate the ability of osteoblasts to be reprogrammed into induced pluripotent stem (iPS) cells and another one was to use defined transcription factors to induce chondrocyte lineage from skin fibroblasts. The selection of osteoblasts is based on the fact that it is originally derived from osteo-chondroprogenitor lineage and the stochastic events of iPS induction might revert osteoblasts first to their progenitor state before becoming pluripotent. The second approach is based on a previous report using three transcription factors (Sox9, Klf4 and c-Myc) to reprogramme skin fibroblasts into chondrocyte lineage. Our aim is to examine whether osteo-chondroprogenitors would be formed during the two reprogramming processes using Sox9-EGFP knock-in mice as a reporter. We reasoned that osteoblasts can be reprogrammed into iPS cells by four Yamanaka’s factors with pluripotency as shown by their ability to form teratomas and contribute to chimeric embryos. However base on the limitation of selector marker of osteo-chondroprogenitor we still cannot capture this progenitor during iPS reprogramming. And because of the pluripotency potential, pluripotent reprogramming approach also brings high risk of teratoma formation. Therefore our second objective was performed to examine whether osteo-chondroprogenitors would be formed during lineage reprogramming. Transient appearance of Sox9-EGFP/Runx2+ve cells was observed in the intermediate stage of over 14 days of chondrocyte lineage induction from skin fibroblasts by Sox9, klf4 and c-Myc. Cells expressing Sox9-EGFP/Runx2+ve showed typical molecular markers of osteo-chondroprogenitors. In vitro and in vivo differentiation assays demonstrated that Sox9-EGFP/Runx2+ve cells can differentiate predominantly into osteoblasts and chondrocytes. Taken together our data indicate that cells with osteo-chondrogenic potential could be generated by defined transcription factors-mediated reprogramming processes. / published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

Esophageal squamous cell carcinoma (ESCC), the major histological subtype of esophageal cancer, is one of the most common malignancies with poor prognosis in the world. Despite continued development of diagnosis and treatment, ESCC remains the sixth leading cause of cancer death worldwide. Current treatment regimens in ESCC are often characterized by ineffectiveness and poor selectivity. Therapeutic methods directed at cancer-associated genes or cancer stem cells (CSCs) may be effective approaches to cure this deadly cancer. Therefore, this study aims to identify specific ESCC-related genes and cancer stemness genes which help us to develop new targeted agents to achieving objective, long-lasting therapeutic responses in ESCC. To obtain an accurate overview of genetic changes occurring in ESCC patients, our group performed microarray-based mRNA expression profiling and high-throughout transcriptome sequencing (RNA-Seq) to compare differentially expressed genes between ESCC tumors and their corresponding non-tumorous tissues. Prostate stem cell antigen (PSCA) was considered to be a candidate of primary interest due to significantly reduced expression in both microarray and RNA-Seq data. In this study, we examined the role of PSCA on the pathogenesis of esophageal cancer. Our results showed that PSCA was frequently down-regulated in ESCC. Its expression was negatively regulated by transcription factor SOX5. Also, we provided evidence that down-regulation of PSCA was associated with poor clinical outcomes of patients with ESCC. Both in vitro and in vivo assays revealed that PSCA could arrest cell cycle progression and promote differentiation. To further elucidate the mechanism involved in biological function of PSCA, we performed co-immunoprecipitation and mass spectroscopy to identify proteins that associate with PSCA. This study found that RB1CC1, a key signaling node to regulate cellular proliferation and differentiation, interacted specifically with PSCA both in vitro and in vivo. Binding of PSCA and RB1CC1 in cytoplasm resulted in stabilization and translocation of RB1CC1 into nucleus and then further regulates the crucial cell cycle and differentiation genes. Furthermore, in order to identify the cancer stemness genes specifically expressed in CSCs of ESCC, we utilized gene expression analysis to profile 34 stemness-associated genes in ESCC specimens. Developmental pluripotency associated 4 (DPPA4), a well known pluripotent marker of stem cell, was considered as the best candidate. Our following histopathological study demonstrated that DPPA4 rigorously marked the rare CSCs, in contrast to core stemness factors (OCT4 and SOX2) and previous reported CSC markers (CD90 and CD44), which expressed in a large population of cancer cells. Moreover, the expression of DPPA4 was also found to have prognostic value in ESCC, as the appearance of DPPA4+ cells was significantly associated with poor differentiation, advanced stage and higher incidences of lymph node metastasis. Finally, our functional studies showed that ESCC cells expressing exogenous DPPA4 conferred an enhanced ability to initiate tumor, self-renew, resist chemotherapy and metastasize through lymphatic system. In summary, this study provide evidence indicating that novel tumor suppressor gene PSCA and cancer stemness gene DPPA4 may contribute to the development and progression of ESCC. Additionally, they may serve as potential targets for development of effective therapeutic strategies. / published_or_final_version / Clinical Oncology / Doctoral / Doctor of Philosophy

No description available.

The cellular microenvironment consists of soluble and insoluble factors that provide signals that dictate cell behavior and cell fate. Limited characterization has hindered our ability to mimic the physiological or pathophysiological environment. While stem cells have vast promise in the areas of regenerative medicine and disease therapy, harnessing this potential remains elusive due to our limited understanding of differentiation mechanisms. Similarly, many in vitro cardiac disease models lack the critical structure- function relationships of healthy and diseased cardiac tissue. The goal of this work is to induce cardiomyogenesis and pathogenesis in vitro by recapitulating features of the native microenvironment during development and disease. / Engineering and Applied Sciences

published_or_final_version / Medicine / Master / Doctor of Medicine

published_or_final_version / Paediatrics / Master / Master of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

No description available.

No description available.

No description available.