While Mks were differentiating and maturing, a progressive increase in type I collagen Crotonoside release was observed as revealed by hydroxyproline quantification. Interestingly, when co-cultures were maintained in hypoxic conditions, the concentrations observed at each time point were significantly higher than those reported for hOST cultures alone or co-cultures at normoxic conditions. As expected, these results indicate that hemopoietic progenitors and Mks promote and sustain bone Orientin formation and matrix deposition. Likewise, oxygen tension also plays a crucial role in promoting type I collagen release. We further investigated the deposition of type I collagen with second harmonic generation imaging, a non-invasive optical method that exploits non-linear light scattering from fibrillar collagen. Using this technology, a time course analysis of the deposition of fibrillar type I collagen from hOSTs revealed that, hOSTs in co-culture with HSCs at 5% oxygen tension organized regularly oriented collagen fibers. The development of the present model to systematically study the functional interactions between HSCs and their niche, mimics and combines in vitro the biochemical and physical parameters of the osteoblastic niche. By including a new non-linear imaging approach based entirely on endogenous sources of optical contrast, this system allows for the non-invasive and dynamic characterization of key structural and architectural features of the model. To date, proplatelet formation and platelet release have been visualized by multiphoton intravital microscopy in intact bone marrow and HSC homing and proliferation within the bone marrow have been traced through the development of ex-vivo real time imaging technology. These studies have described the osteoblastic niche as a very dynamic environment where HSCs can be exposed to both endosteum and vascular signals that differently determine their fate. Despite this improvement in the knowledge of functional bone marrow niches, the mechanisms underlying the relationships between HSCs and their environment are not understood, especially in humans where invasive approaches are not possible.