Immunomodulatory and antimicrobial non-mulberry Antheraea mylitta silk fibroin accelerates in vitro fibroblast repair and regeneration by protecting oxidative stress.

The antimicrobial nature of Antharaea mylitta silk-fibroin (SF) is reported but antioxidant potential and the immunomodulatory role towards the fibroblast cell repair process is not explored. Polyurethane is reported to have inflammatory potential by mononuclear cells directed cytokine release, which can guide fibroblast repair. Present study demonstrates the conjunctive effect of inflammatory PU/SF to regulate the favorable shift from pro-inflammatory to anti-inflammatory cytokine stimulation for accelerated fibroblast repair. Minimal inhibitory concentration of SF was determined against pathogenic strains and the effect of SF was investigated for fibroblast NIH3T3 cell adhesion. SF doses (8, 8.5, 9 mg mL-1) were found to be greater than both the IC50 of DPPH scavenging and the ED50 for NIH3T3 proliferation. Anti-lipid peroxidase (ALP) activity of SF doses and citric acid-treated NIH3T3 cells were compared under hydrogen peroxide (H2O2) induced oxidative stress. 9 mg mL-1 SF showed greater ALP activity than the citric acid standard. SF-driven protection to oxidative damage was measured by viable cell fraction in trypan blue dye exclusion assay where 9 mg mL-1 SF showed the highest viability (p ≤ 0.05). 9 mg mL-1 SF was blended with PU for scaffold (w/v = 2 : 5, 2 : 7, 2 : 9) fabrication. The protective effect of PU/SF (2 : 5, 2 : 7, 2 : 9) against oxidative stress was verified by damaged cell survival in MTT assay and DNA quantification. The highest number of cells survived on PU/SF (2 : 9) at all intervals (p ≤ 0.01) upon oxidative damage; PU/SF (2 : 9) was also fabricated by employing the immobilization technique. Immobilized PU/SF (2 : 9) exhibited a greater zone of microbial inhibition, a higher extent of inhibition to microbial adherence, and caused more LDH release from bacterial cell membrane due to membrane rupture, resulting in bacterial cell death (E. coli, K. pneumoniae, P. aeruginosa, S. aureus) compared to the experimental results shown by blended PU/SF (2 : 9). The protective nature of PU/SF (2 : 9) against oxidative stress was ensured through the LDH activity of damaged NIH3T3 cells. Initial raised IL-6, TNF-alpha (pro-inflammatory cytokines) and lowered IL-8, IL-10 (anti-inflammatory cytokine) profiles coupled with fallen IL-6, TNF-alpha, and elevated IL-8, IL-10 at later hours synergistically progress the inflammatory phase of in vitro scratch wound repair in mononuclear culture treated by PU/SF (2 : 9).

Anti-inflammatory effect of epidermal growth factor conjugated silk fibroin immobilized polyurethane ameliorates diabetic burn wound healing.

Few studies are reported on immunomodulatory potential of non-mulberry (Antheraea mylitta) silk fibroin (SF) combined with polyurethane (PU) in diabetic wound healing. In this study, PU/SF (Antheraea mylitta) scaffolds were fabricated by blending and immobilization techniques. Effective SF dosage was determined and incorporated according to minimum inhibitory concentrations against wound associated bacterial strains while fabricating scaffold. Dermal fibroblast NIH3T3 cells were seeded on epidermal growth factor (EGF) treated and untreated PU/SF scaffolds, fabricated by blending and immobilization techniques. Fibroblast seeded PU/SF scaffolds were investigated for anti-inflammatory response in wound recovering potential comparing with Acticoat™ in third degree burn of streptozotocin induced diabetic rats. At 16th, 24th days, promising healing was achieved with faster granulation, enhanced collagenization, patterned re-epithelialization by EGF treated cellular immobilized PU/SF in normal, hyperglycemic burn. Biomarkers of different healing stages, CD31 (haemostasis), Ki67 (proliferative), alpha-sma, COL III (maturation) were examined. Since hyperglycemic burn is characterized by inflated pro-inflammatory cytokines, serum, tissue IL-6,8,10 were recorded, which revealed timely restoration of inflated IL-6,8 and protection against IL-10 elevation by cellular immobilized PU/SF compared to Acticoat™ (p ≤ 0.05), control (p ≤ 0.01). E-cadherin (gap junction protein), MMP 9 response suggested anti-inflammatory role of PU/SF on accelerated healing of thermal injury as potent dermal substitute.

User-friendly tool kits for protein gel electrophoresis techniques: A training program for high school students.

Recent advancements in biochemical sciences have helped the researchers to explore the molecular logic of life inclusive of its multifarious expressions and explain many facts about the structure and functions of cellular macromolecules. Due to its simple and cost-effective nature, polyacrylamide gel electrophoresis (PAGE) has become the most favored technique for qualitative and quantitative examination of macromolecules. Major drawbacks of such modifications are the cost and operational complexities faced by naïve students. Many interlinking laboratory equipment are needed in the school laboratories for the conduct of even simple scientific experiment. Some of these costly modern equipment are inaccessible for students of small laboratories, and their alternatives are not easily available. Many of these laboratory equipment required for routine gel electrophoresis technique can be fabricated in their simplest form using off-the-shelf components. A short term biochemistry training program was executed for high school students to provide them "hands-on" training using newly modified equipment, which was proved to be an exciting way of learning biochemical gel separation techniques. © 2018 International Union of Biochemistry and Molecular Biology, 46(5):566-577, 2018.