## 4 Discussion  
Crerams and gels are among the most popular vehicles for the topical treatment of local pain and skin inflammation as well as for skin care. Ready-to-use base creams and gels are widely utilized, especially by small companies, to quickly obtain products with specific properties, adding active ingredients to the ready-made bases. Moreover, topical applications are highly appreciated by patients, especially for their quick application and ease of use, significantly increasing their compliance [28]. In our laboratories, we have studied the activity of compounds, especially natural ones, in cellular models to develop innovative formulations for human health and well-being. We have studied glucosamine derivatives for the treatment of osteoarthritis (OA) for a long time [23; 29], as well as substances such as _H. procumbens_ extract (HPE), which exhibit analgesic activities [24; 25; 30; 31]. In particular, we have previously shown that nNAG is much more effective than NAG in stimulating extracellular matrix production in human primary chondrocytes. Based on previous findings, we focused our studies on comparing commercial base creams and gels to which we added NAG, nNAG, and _H. procumbens_. However, through the evaluation of the sensory properties of creams and gels containing _H. procumbens_ extracts, we observed an intense, unpleasant brown color, leading us to exclude them from our subsequent studies. Moreover, analyzing the viscosity parameters, we observed many differences among all creams and gels containing 10% HPE. Considering that HPE root extract is a micrometric-sized powder containing several compounds, its behavior could be due to the association of these compounds with each other. For this reason, we decided to study NAG and nNAG, which are pure molecules, proposing to study HPE root extract in the future. In the present study, we analyzed the effects of nNAG in comparison with NAG, both in chondrocytes and fibroblasts in 3D cell cultures that resemble physiological conditions. The finding that mainly nNAG but also NAG are very effective at increasing collagens and decreasing MMPs in these two types of cells is very encouraging, also suggesting a higher  
Figure 4: Effects of NAG and nNAG on MMP production. Fibroblasts, cultured in spheroids, were left untreated (CTL) or treated with either 10 \(\mu\)M NAG or nNAG for 72 h. After the treatments, cell media were collected and analyzed by ELISA to measure the levels of MMP-3 and MMP-9 protein secretion. The results are reported as ng/mL and are expressed as the mean \(\pm\) SEM of data obtained from three independent experiments. * \(p\) < 0.05 vs. CTL. biological efficacy of these molecules when added in creams and gels. Chondrocytes are the only type of cell present in cartilage, where they produce and maintain the extracellular matrix (ECM) through the secretion of new ECM components, such as collagens, and the production of enzymes, such as MMPs, involved in the degradation of old ECM components, thereby maintaining homeostasis [32]. During the progression of OA, a loss of homeostasis leads to the degradation of cartilage [33]. Thus, the presence of nNAG in creams and gels for the topical treatment of OA is desirable. On the other hand, the ability of nNAG to stimulate the production of collagen type I--typical of skin--and inhibit MMP production suggests, once again, the suitability of these molecules as components of creams and gels used as topical treatments, as well as for wound repair, for various patients. The main components of commercial base creams are water and oil, and emulsifiers or surfactants are added to stabilize the product and improve its texture and properties. The type of emulsion (W/O or O/W) and texture are important for the end purpose, and they influence the flow and deformation properties [34; 35]. Topical gels are semi-solid systems that contain a liquid phase in a three-dimensional polymer matrix of natural or synthetic gums with high physical or chemical affinity.