Potential of Biofermentative Unsulfated Chondroitin and Hyaluronic Acid in Dermal Repair.

Antonietta Stellavato,Maria D’Agostino,Antonella D’Agostino,Luisana Corsuto,Annalisa La Gatta,Chiara Schiraldi,Marcella Cammarota,Donatella Cimini

doi:10.3390/ijms23031686

Abstract

Chondroitin obtained through biotechnological processes (BC) shares similarities with both chondroitin sulfate (CS), due to the dimeric repetitive unit, and hyaluronic acid (HA), as it is unsulfated. In the framework of this experimental research, formulations containing BC with an average molecular size of about 35 KDa and high molecular weight HA (HHA) were characterized with respect to their rheological behavior, stability to enzymatic hydrolysis and they were evaluated in different skin damage models. The rheological characterization of the HHA/BC formulation revealed a G’ of 92 ± 3 Pa and a G″ of 116 ± 5 Pa and supported an easy injectability even at a concentration of 40 mg/mL. HA/BC preserved the HHA fraction better than HHA alone. BTH was active on BC alone only at high concentration. Assays on scratched keratinocytes (HaCaT) monolayers showed that all the glycosaminoglycan formulations accelerated cell migration, with HA/BC fastening healing 2-fold compared to the control. In addition, in 2D HaCaT cultures, as well as in a 3D skin tissue model HHA/BC efficiently modulated mRNA and protein levels of different types of collagens and elastin remarking a functional tissue physiology. Finally, immortalized human fibroblasts were challenged with TNF-α to obtain an in vitro model of inflammation. Upon HHA/BC addition, secreted IL-6 level was lower and efficient ECM biosynthesis was re-established. Finally, co-cultures of HaCaT and melanocytes were established, showing the ability of HHA/BC to modulate melanin release, suggesting a possible effect of this specific formulation on the reduction of stretch marks. Overall, besides demonstrating the safety of BC, the present study highlights the potential beneficial effect of HHA/BC formulation in different damage dermal models.

Highlights

Skin integrity and texture of the dermal matrix are correlated to the restoration of the biomacromolecules that contribute to preserve hydration and assembly of the extracellular matrix (ECM), and to the biochemical determinants able to counteract oxidative stress and more generally aging-related damages
Amplitude sweep tests revealed a fluid behavior for the high molecular weight HA (HHA)/biofermentative unsulfated chondroitin (BC) sample, with 92 ± 3 Pa and 116 ± 5 Pa G and G values, respectively
The mechanical spectrum (Figure 1) confirmed fluid behavior (G values higher than G ) at low frequencies. Both the moduli progressively increased with the oscillation frequency with G rising more markedly than G

Summary

Introduction

Skin integrity and texture of the dermal matrix are correlated to the restoration of the biomacromolecules that contribute to preserve hydration and assembly of the extracellular matrix (ECM), and to the biochemical determinants able to counteract oxidative stress and more generally aging-related damages. In our research unsulfated chondroitin were used and obtained by fermentation of the recombinant E. coli K4 strain EcK4r3, and purified by ultrafiltration/diafiltration, alcohol precipitation and active carbon treatments [9] This product is here addressed as biofermentative unsulfated chondroitin (BC), a CS variant, since it consists of the same repeating disaccharide unit of CS, formed by D-glucuronic acid and N-acetyl-D-galactosamine, but, like HA, without any sulfate groups. BC promoted cell proliferation and concurrently preserved the chondrocyte phenotype It showed improved anti-inflammatory properties compared to CS, reducing cytokines levels in a primary human chondrocyte inflammation model [10]. Previous studies, combining hyaluronic acid at higher and lower molecular weight (HCC), were investigated for dermal application and showed beneficial effects in tissue damage [11]. The filling ability and the hydration capacity of injective formulation of linear HA are known to be related to HA molecular weight and concentration

Methods

Results

Discussion

Conclusion