Distinctive Hyaluronan Metabolism in Epidermal Keratinocytes: Roles of HYAL1 and HAS3

Hyaluronan (HA) is an essential extracellular matrix molecule found in the skin over half of total HA. HA plays an important role in several biological processes, including cell movement, differentiation and managing inflammation, and supports hydration, elasticity, and the barrier function in the skin. Epidermal keratinocytes make up most of the cells in the epidermis are responsible for producing and breaking down HA, however, the exact molecular pathways that regulate this process remain unclear. Unlike dermal fibroblasts, which use the HYBID/KIAA1199 system and hyaluronan synthase 2 (HAS2) for HA metabolism, early findings suggest that keratinocytes rely on a completely different set of enzymes and regulators. The acidic environment of the stratum corneum—the skin’s outermost layer—adds another layer of complexity and create both challenges and opportunities for these enzymes to function. Another challenge in HA research is figuring out how its two forms—high-molecular-weight HA (HMW-HA) and low-molecular-weight HA (LMW-HA)—affect the skin differently. HMW-HA is linked to hydration and anti-inflammatory properties, while LMW-HA often triggers inflammation and cancer cell invasion. The balance between these two forms is essential for keeping the skin healthy and addressing conditions that disrupt its natural state. However, the mechanisms that keratinocytes use to maintain this balance are still not well understood, leaving an important gap in research. Recognizing this, a team of researchers at Gifu Pharmaceutical University, led by Drs. Shintaro Inoue, and Yukiko Mizutani, investigated how keratinocytes regulate HA with detailed focus on the specific roles of HYAL1, which degrades HA, and HAS3, which synthesizes it.

The researchers compared how HA metabolism differs between dermal fibroblasts and epidermal keratinocytes. Using qPCR, they measured the expression of genes involved in HA degradation, such as HYBID and hyaluronidases, as well as those responsible for making it, like HASs. They were surprised that HYBID, a key enzyme for HA degradation in dermal fibroblasts, was barely expressed in keratinocytes. To compare the HA depolymerization activity of living cells, keratinocytes did not show HYBID-dependent extracellular HMW-HA depolymerization. Instead, the keratinocytes showed much higher levels of HYAL1, pointing to a unique process for HA degradation in the epidermis. To understand HYAL1’s role more clearly, the team studied its activity in the conditioned medium of cultured keratinocytes. Unexpectedly, HYAL1, known as an intracellular lysosomal enzyme, was secreted extracellularly. They added HMW-HA to this medium and observed its degradation , but only under acidic conditions in vitro. This matched the naturally acidic environment of the epidermis, particularly in the stratum corneum. When HYAL1 was silenced using small interfering RNA (siRNA), HA degradation almost completely stopped, confirming HYAL1 as the main enzyme responsible. HYAL2, another enzyme that could degrade HA, showed little to no activity under the same conditions, further highlighting HYAL1’s central role. They also noticed that HYAL1 secretion increased significantly during keratinocyte differentiation, linking its function to the skin’s maturation process.

On the other side of HA metabolism—its production—the authors explored the roles of the three known hyaluronan synthases: HAS1, HAS2, and HAS3. Their results showed that HAS3 was the dominant synthase in keratinocytes, with far higher levels compared to HAS2 or HAS1. To confirm its importance, they used siRNA to knock down HAS3, which caused an 81% reduction in HA production. In contrast, silencing HAS2 had no significant effect, demonstrating that HAS3 is the key enzyme driving HA synthesis in keratinocytes. According to the authors, this finding sets keratinocytes apart from dermal fibroblasts, where HAS2 takes the lead in producing HA. The team also looked at how external factors influence HA synthesis. When keratinocytes were exposed to interferon-gamma, a cytokine involved in immune responses, HAS3 expression increased significantly in a dose-dependent manner. This boost in HAS3 led to a noticeable rise in HMW-HA production which links immune signaling to HA metabolism. Surprisingly, knocking down TMEM2, a protein previously thought to help degrade HA, instead caused an increase in HAS3 expression and HA production. This revealed an unexpected regulatory connection between TMEM2 and HAS3. To ensure their findings reflected real physiological processes, the researchers examined how differentiation affects HYAL1 and HAS3 expression. They found that as keratinocytes matured, HYAL1 secretion into the extracellular space increased, consistent with its role in breaking down HA in the epidermis. In contrast, HAS3 expression slightly decreased, matching the observation that HMW-HA is more concentrated in the basal layers of the epidermis than in the upper layers. This balance between differentiation, enzyme activity, and HA metabolism highlighted the tightly controlled nature of HA turnover in the epidermis. One of the most intriguing parts of the study involved tracking how HYAL1 is transported outside the cell. Using specialized tools like deglycosylation assays with PNGase F, they discovered that newly made HYAL1 is glycosylated in the Golgi apparatus and secreted directly into the extracellular environment.

In conclusion, the study by Dr. Yukiko Mizutani and colleagues provided important new understanding on how HA is metabolized in the epidermis, setting it apart from the processes seen in the dermis.  The clinical applications of the findings of Gifu Pharmaceutical University scientists could be far-reaching, for instance, psoriasis, eczema, or age-related dryness could benefit from therapies targeting HYAL1 or HAS3 and controlling HYAL1 activity might prevent excessive HA breakdown and help the skin retain moisture and elasticity. On the other hand, a method to boost HAS3 could enhance HA production and support repair in damaged or aging skin. Additionally, HA is already widely used in dermal fillers and wound-healing products and with the better understanding how keratinocytes manage HA metabolism, it opens the door to develop more precise and natural therapies. Stimulating HAS3 through immune signaling, for instance, could encourage the skin to produce its own HA and by it reduces the need for synthetic replacements. Beyond dermatology, the findings also touch on cancer biology with HMW-HA has anti-inflammatory properties, while LMW-HA can fuel inflammation and tumor growth and how keratinocytes regulate the balance between these forms could lead to new strategies for preventing or treating skin cancers such as to target HYAL1 which will limit the harmful effects of LMW-HA in the skin.