The periosteum on the skeletal surface creates a unique micro-environment for cortical bone homeostasis, but how this micro-environment is formed remains a mystery. In our study, we observed the cells in the periosteum presented elongated spindle-like morphology within the aligned collagen fibers, which is in accordance with the differentiated osteoblasts lining on the cortical surface. We planted the bone marrow stromal cells(BMSCs), the regular shaped progenitor cells, on collagen-coated aligned fibers, presenting similar cell morphology as observed in the natural periosteum. The aligned collagen topology induced the elongation of BMSCs, whichfacilitated the osteogenic process. Transcriptome analysis suggested the aligned collagen induced the regular shaped cells to present part of the periosteum derived stromal cells(PDSCs) characteristics by showing close correlation of the two cell populations. In addition, the elevated expression of PDSCs markers in the cells grown on the aligned collagen-coated fibers further indicated the function of periosteal topology in manipulating cells' behavior. Enrichment analysis revealed cell-extracellular matrix interaction was the major pathway initiating this process, which created an osteo-friendly micro-environment as well. At last, we found the aligned topology of collagen induced mechano-growth factor expression as the result of Igf1 alternative splicing, guiding the progenitor cells behavior and osteogenic process in the periosteum. This study uncovers the key role of the aligned topology of collagen in the periosteum and explains the mechanism in creating the periosteal micro-environment, which gives the inspiration for artificial periosteum design.