Biochemical and immunomodulatory insights of extracellular matrix from decellularized human whole cervix: recellularization and in vivo ECM remodelling interplay.

Extracellular matrix (ECM) rich whole organ bio-scaffolds, preserving structural integrity and essential growth factors, has potential towards regeneration and reconstruction. Women with cervical anomalies or trauma can benefit from clinical cervicovaginal repair using constructs rich in site specific ECM. In this study, complete human cervix decellularization was achieved using a modified perfusion-based stir bench top decellularization method. This was followed by physico-chemical processes including perfusion of ionic agents, enzymatic treatment and washing using detergent solutions for a duration of 10-12 days. Histopathological analysis, as well as DNA quantification confirmed the efficacy of the decellularization process. Tissue ultrastructure integrity was preserved and the same was validated via scanning electron microscopy (SEM) and transmission electron microscopy (TEM) studies. Biochemical analysis and structural characterizations like FTIR, Raman spectroscopy of decellularised tissues demonstrated preservation of important proteins, crucial growth factors, collagen, and glycosaminoglycans (GAGs ). In vitro studies, using THP-1and HUVEC cells, demonstrated macrophage polarisation from M1 to M2 and vascular functional genes enhancement, respectively, when treated with decellularized cervical matrix. human decellularised cervical matrix (hdCM) () grafts were recellularized with site specific human cervical epithelial cells and HUVEC, showing non-cytotoxic cell viability and enhanced proliferation. Futhermore, hdCM scaffolds showed immunomodulatory effects in vivo on small rodent model via upregulation of M2 macrophage genes as compared to decellularised rat cervical matrix scaffolds (DRC). hdCM scaffolds underwent neo-vascularization followed by ECM remodelling with enhanced tissue integration.

Thermoresponsive keratin-methylcellulose self-healing injectable hydrogel accelerating full-thickness wound healing by promoting rapid epithelialization.

Chronic wounds suffer from impaired healing due to microbial attack and poor vascular growth. Thermoresponsive hydrogels gained attention in wound dressing owing to their gelation at physiological temperature enabling them to take the shape of asymmetric wounds. The present study delineates the development of thermoresponsive hydrogel (MCK), from hair-derived keratin (K) and methylcellulose (MC) in the presence of sodium sulfate. The gelation temperature (Tg) of this hydrogel is in the range of 30 °C to 33 °C. Protein-polymer interaction leading to thermoreversible sol-gel transition involved in MCK blends has been analyzed and confirmed by FTIR, XRD, and thermal studies. Keratin, has introduced antioxidant properties to the hydrogel imparted cytocompatibility towards human dermal fibroblasts (HDFs) as evidenced by both MTT and live dead assays. In vitro wound healing assessment has been shown by enhanced migration of HDFs in the presence of MCK hydrogel compared to the control. Also, CAM assay and CD31 expression by the Wistar rat model has shown increased blood vessel branching after the implantation of MCK hydrogel. Further, in vivo study, demonstrated MCK efficacy of hydrogel in accelerating full-thickness wounds with minimal scarring in Wistar rats, re-epithelialization, and reinstatement of the epidermal-dermal junction thereby exhibiting clinical relevance for chronic wounds.

Iodine functionalized 2,5-dimethoxy-2,5-dihydrofuran (DHFI) crosslinked whey protein-derived carbon nanodots (WCND) for antibacterial application.

Whey protein-derived carbon nanodots (WCND) were synthesized using the microwave irradiation method, and its amine-rich surface functionality was crosslinked with covalently bound Iodine functionalized 2,5-dimethoxy-2,5-dihydrofuran (DHFI) to produce WCND-DHFI. The physicochemical characterization of both WCND and WCND-DHFI was performed and compared to comprehend the consequence of iodination on the characteristics of WCND. The suitability of CND in biological environments was evaluated through in vitro cytocompatibility and Chorioallantoic Membrane (CAM) assay, as well as a hemocompatibility study. WCND-DHFI has shown enhanced cell viability against WCND. Further, the antibacterial properties of both CNDs were studied against both gram-positive and gram-negative bacterial strains, representing an enhancement in antibacterial activity after DHFI crosslinking. WCND-DHFI has depicted a stable and prominent bacteriostatic activity for up to 6 h for both strains of bacteria. WCND-DHFI has denoted a 99.996% and 99.999% loss of bacterial viability for gram-positive and negative strains, respectively. Novel surface functionalization portrays an improvement in antibacterial activity. Transmission and scanning electron microscopy represent the cell wall rupturing by the WCND-DHFI, resulting in bacterial death. The ROS-mediated bacteriostatic mechanism of WCND-DHFI has been explored through assessing lipid peroxidation and protein oxidation assay. Moreover, the oxidative damage of DNA also has been explored. WCND-DHFI is performing as a promising cytocompatible and hemocompatible material for antibacterial applications.

Biodegradable Multi-layered Silk Fibroin-PCL Stent for the Management of Cervical Atresia: In Vitro Cytocompatibility and Extracellular Matrix Remodeling In Vivo.

Cervical atresia is a rare congenital Müllerian duct anomaly that manifests as the absence or deformed nonfunctional presence of the cervix. Herein, a multi-layered biodegradable stent is fabricated using a homogeneous blend of silk fibroin with polycaprolactone using hexafluoroisopropanol as a common solution. Briefly, a concentric cylinder of 3D honeycomb layer is sandwiched within electrospun sheets for fixing at the cervico-uterine junction to pave the way of cervical reconstruction. An average length of 40 mm with 3 mm diameter is fabricated for the hybrid stent design. SEM evidences an evenly distributed pore architecture of the electrospun layer, and mechanical characterization of stent reveals a tensile strength of 1.7 ± 0.2 MPa, with a Young's modulus of 5.9 ± 0.1 MPa. Physico-chemical characterization confirms the presence of silk fibroin and poly caprolactone within the engineered stent. Following 14 days of pepsin enzymatic degradation, 18% degradation and a contact angle measurement of 97° are observed. In vitro cytocompatibility studies are performed using site-specific primary human cervical squamous, columnar epithelial cells, and human endometrial stromal cells. The study demonstrates non-cytotoxic cells' viability (no significant toxicity), improved cell anchoring, adherence among the stent layers, and proliferation in the 3D microenvironment. Furthermore, in vivo subcutaneous studies in the rodent model indicate that the implanted stent undergoes constructive remodeling, neo-tissue creation, neo-vasculature formation, and re-epithelialization while maintaining patency for 2 months.

High fibroin-loaded silk-PCL electrospun fiber with core-shell morphology promotes epithelialization with accelerated wound healing.

Silk fibroin (SF) is a widely explored biopolymer for wound-healing applications due to the presence of amino acids in the biodegradable polymer chain with superior mechanical properties. Herein, a high SF-loaded fibrous matrix along with poly(ε-caprolactone) (PCL) was fabricated using electrospinning of emulsion and blend compositions to modulate nanostructure morphology. A comparative study of the physicomechanical properties of electrospun fibers with emulsion (eS7P3) and homogenous blend (bS7P3) was performed as well. In both compositions, SF loading of up to 70% was successfully achieved in the spun fibers while emulsion yielded core-shell morphology, and the blend resulted in monolith fiber architecture as evidenced by TEM microscopy. Further characterization revealed superior mechanical properties in S7P3 fiber with core-shell morphology, as compared to those in the monolith in terms of a higher degree of crystallinity with Young's modulus of 60 MPa under tensile test and nanoindentation modulus of 1.59 ± 0.8 GPa. Further, eS7P3 nanostructure morphology containing silk in the core with a thin outer layer of PCL facilitated relatively faster biodegradation in the lysozyme medium, as compared to that in the monolith. Owing to the presence of a hydrophobic shell, protein adsorption on the fibrous mat presented slow but steady kinetics up to 24 h. When the scaffold was seeded with human placenta-derived mesenchymal stem cells (hPMSCs), in vitro study confirmed that the eS7P3 structure had marginally higher cell proliferation with superior cell infiltration than the monolith. Further, in vivo study involving a rodent model showed the potential of the eS7P3 fiber substrate with a core-shell structure for accelerating full-thickness wound healing by inducing hair follicle and wound closure with less scar formation after 15 days.