Jhon Jairo Melchor-Moncada

This study evaluated the potential of Colombian silk fibroin hydrogels functionalized with recombinant LSECtin (rLSECtin) for intervertebral disc tissue engineering. In silico analysis indicated that rLSECtin may interact with glycans similar to those found in human adipose-derived stem cells (ADSCs) through residues Asn105, Asp106, Glu93, Asn87, and Glu85, which coordinate calcium to form hydrogen bonds with GlcNAc hydroxyl groups. rLSECtin was successfully produced, purified, and incorporated into silk fibroin hydrogels. Thermal analysis showed that the hydrogels were resistant to sterilization, and rheological studies demonstrated predominantly elastic, gel-like behavior. The secondary structure of the hydrogels was largely determined by α-helical content, which is related to silk I with β-sheets also contributing as a result of sonication-induced structuring. Metabolic activity assays indicated that rLSECtin was associated with increased ADSC metabolic activity under two-dimensional conditions, whereas hydrogels containing rLSECtin maintained cell viability, as confirmed by cell viability assays. Differentiated constructs exhibited glycosaminoglycan-rich extracellular matrix and chondrocyte-like morphology, whereas rLSECtin-functionalized hydrogels showed increased transcription of SOX9 , ACAN , and COL2A1 , along with elevated COL10A1 of uncertain significance for nucleus pulposus stability. Overall, Colombian silk fibroin hydrogels provide a mechanically stable and cytocompatible platform relevant for nucleus pulposus tissue engineering, in which rLSECtin appears to act primarily as a biochemical modulator rather than a structural modifier. These findings support a proof of concept for lectin-functionalized biomaterials in intervertebral disc regeneration. However, further studies, including mechanistic validation of glycan–lectin interactions, protein-level confirmation, quantitative extracellular matrix analysis, and evaluation of signaling pathways, are required to establish functional and translational relevance.

Antimicrobial resistance (AMR) poses a significant global health threat, necessitating the development of novel antibacterial strategies. Serratiopeptidase (SP), a metalloprotease produced by bacteria such as Serratia marcescens, has gained attention not only for its anti-inflammatory properties but also for its potential antibacterial activity. However, its protein nature makes it susceptible to pH changes and self-proteolysis, limiting its effectiveness. This study aimed to increase both the enzymatic stability and antibacterial activity of serratiopeptidase through immobilization on titanium oxide nanoparticles (TiO₂-NPs), leveraging the biocompatibility and stability of these nanomaterials. Commercial TiO₂-NPs were characterized using TGA/DTG, FT-IR, UV–Vis, and XRD analyses, and their biocompatibility was assessed through cytotoxicity studies. Serratiopeptidase was produced via fermentation using the C8 isolate of Serratia marcescens obtained from the intestine of Bombyx mori L., purified chromatographically, and immobilized on carboxylated nanoparticles via EDC/NHS coupling at various pH conditions. The optimal enzymatic activity was achieved by using pH 5.1 for nanoparticle activation and pH 5.5 for enzyme coupling. The resulting bioconjugate demonstrated stable proteolytic activity at 25 °C for 48 h. Immobilization was confirmed by FT-IR spectroscopy, and the Michaelis–Menten kinetics were determined. Notably, the bioconjugate exhibited two-fold greater antibacterial activity against E. coli than the free enzyme or TiO₂-NPs at 1000 µg/mL. This study successfully developed a serratiopeptidase–TiO₂ bioconjugate with enhanced enzymatic stability and antibacterial properties. The improved antibacterial activity of the immobilized enzyme presents a promising approach for developing new tools to combat antimicrobial resistance, with potential applications in healthcare, food safety, and environmental protection.

Serratiopeptidase, a bacterial metalloprotease known for its pain-relieving and anti-inflammatory properties, can be produced through fermentation with S. marcescens. This study aimed to identify key factors related to nutrient composition and physicochemical conditions for production in Erlenmeyer flasks and to scale up the mixture to a bioreactor to obtain the maximum proteolytic activity. A Plackett–Burman design was used to determine whether the presence of silkworm pupae (at 1.5%) was a significant parameter for serratiopeptidase production. Along with the variables pH, temperature, and time, they were optimized using a Taguchi experimental design, resulting in values of 7, 25 °C, and 36 h, respectively. Scaling up with a k_La of 25.45 ± 3.12 h⁻¹ showed the highest serratiopeptidase production at 24 h. A factorial design was used for ultrafiltration, resulting in an LMH (liters per square meter per hour) of 960 L/m²h, a TMP (transmembrane pressure) of 15 psi, and a concentration factor of five, with a specific activity of 24,325.81 ± 1515.69 U/mg. Afterward, the retentate was purified using strong anion exchange chromatography and ultrafiltration, yielding a 19.94 ± 3.07% recovery and a purification factor of 1.59 ± 0.31. In conclusion, waste from the sericulture industry can be used for serratiopeptidase production.

The development and evaluation of scaffolds play a crucial role in the engineering of hyaline cartilage tissue. This work aims to evaluate the performance of silk fibroin hydrogels fabricated from the cocoons of the Colombian hybrid in the in vitro regeneration of hyaline cartilage. The scaffolds were physicochemically characterized, and their performance was evaluated in a cellular model. The results showed that the scaffolds were rich in random coils and β-sheets in their structure and susceptible to various serine proteases with different degradation profiles. Furthermore, they showed a significant increase in ACAN, COL10A1, and COL2A1 expression compared to pellet culture alone and allowed GAG deposition. The soluble portion of the scaffold did not affect chondrogenesis. Furthermore, they promoted the increase in COL1A2, showing a slight tendency to differentiate towards fibrous cartilage. The results also showed that Colombian silk could be used as a source of biomedical devices, paving the way for sericulture to become a more diverse economic activity in emerging countries.