It is generally believed that organized neural architecture is essential for nervous system development, function, and regeneration. In the absence of guidance cues, regenerating axons may lose their directions and become misaligned, resulting in the formation of neuromas and/or misappropriate connections. To help regenerate axons across damaged regions and guide them to appropriate targets, some bridging devices such as microgrooves are being intensively researched to achieve a better directional axonal growth. This paper reports a novel fabrication process to generate a highly aligned groove texture on the inner surface of semipermeable hollow fiber membranes (HFMs). HFMs have demonstrated promising results in guiding axonal regeneration. The fabrication process utilized a wet phase-inversion procedure with polyurethane (PU) as model polymer, dimethyl sulfoxide (DMSO) as solvent, and water as nonsolvent. Data indicated that highly aligned groove texture could be formed on the HFM inner surface by carefully controlling phase-inversion conditions such as the polymer solution flow rate, and/or nonsolvent flow rate, and/or polymer solution concentration ratio. The texture forming mechanism is qualitatively explained using a PU-DMSO-water ternary phase diagram and the dynamics of fluid instability. Axonal outgrowth on the HFM with aligned grooves showed the highly aligned orientation and improved axonal outgrowth length. This study may eventually lead to a new and effective way to fabricate nerve grafts for the spinal cord injury and nerve damage treatment based on this highly aligned three dimensional scaffold.