Cell-based therapies in the clinic are limited by the number of cells that can be produced quickly and inexpensively. Whereas about one million cells are isolated from a single donor, existing cell-based therapies can require hundreds of millions to billions of cells. Rapid, exponential expansion of cell number would allow faster delivery of life-saving treatments, such as bone marrow transplants, to a greater number of patients.. Chaotic printing is a novel, patent-pending, high-resolution biofabrication technology that could dramatically improve cell expansion capabilities. It produces layered filaments with significantly higher Surface Area per unit Volume (SAV) than existing cell proliferation systems. Cell-laden hydrogel layers can be interspersed with “fugitive ink” layers. The fugitive ink dissolves and evacuates to leave open channels in between the cell-laden layers. The higher SAV should facilitate an exponentially larger interface between nutrient media and cells cultured in these filaments. We predicted this would dramatically improve the speed and yield of cell proliferation by improving nutrient availability and waste removal. In a first experiment, Bone Marrow-derived human Mesenchymal Stem Cells (BM-hMSCs) were cultured for 1 month in chaotically printed hydrogel filaments. To observe the open channels’ impact on cell expansion, hydrogel filaments with alternating cell-laden layers and open channels were compared to a control group of hydrogel filaments without any open channels. The group with open channels had significantly more cells than the control group on days 1, 7, 14, and 21 of culture. The largest difference between the groups occurred on day 7, the open channel group having 2.1 times as many cells as the control group. This experiment showed BM-hMSC viability for 1 month with our hydrogel formulation and chaotic printing method while also demonstrating increased cell expansion rate with open channels present. We also observed a peak expansion rate within the first week with the open channel design. We proceeded to develope and validate a novel bioreactor design for perfusing the open layers in our chaotically printed hydrogel filaments with flowing nutrient media. We hypothesized that adding flow to the system would further improve nutrient availability and waste removal. Co-axially extruding calcium chloride along with the hydrogel bioinks through our printhead allowed for chaotically printed hydrogel filaments to be solidified and extruded directly into small polystyrene tubes. This method results in consistent hydrogel filaments that contain open channels and are flush to the edges of the polystyrene “bioreactor” tubes into which they are extruded. This well-fitting bioreactor system ensures the flow of nutrient media through the open channels. These tubes can then be cultured within an incubator while connected to a flow circuit driven by a peristaltic pump located outside the incubator. A second experiment demonstrated that BM-hMSCs continued expanding in number for 1 week in chaotically printed hydrogel filaments housed in polystyrene bioreactor tubes when media was flowed through (i.e., replaced via flow) twice per day. Cell-laden filaments in tubes without flow regimens had virtually no viability by day 4. Work is ongoing on a flow regime that will optimize BM-hMSC expansion.