Chronic wounds lead to morbidity, mortality, along with significant economic burden,
often due to the non-availability of suitable treatments. Diabetic wounds are most
common type of chronic wounds which exhibit complex pathophysiology and pose a
significant burden on the wound care industry. Wound dressings have emerged as an
effective treatment to maintain moisture, and protect from pathogens that impede
wound healing. Although different type of single layer dressings, such as gauze,
hydrogel, film, etc., are available yet none fulfill all the requirements for optimal
healing of chronic wounds. However, nanofibrous architectures have gained much
attention as they can mimic the collagen based fibrillar organization of the ECM.
Moreover, they possess high surface area and interconnected nanopores for efficient
exude accumulation and proper aeration. Several techniques have been used to fabricate
nanofibrous polymer matrices such as phase separation, wet spinning, self-assembly,
template-directed synthesis, and electrospinning. Among these, electrospinning is the

most simple, efficient, and promising method. Furthermore, in the case of deep/full-
thickness wounds single layer dressings cannot recover skin to its initial form and

functionality. Thus, full thickness diabetic wounds entail additional novel treatment and
management strategies due to complex and disturbed pathophysiology.

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In addition to the architectural requirements, the wound dressing should also have
active ingredients (biological, natural or chemical moieties) which can enhance wound
healing along with prevention of infection. Synthetic polymer-based dressings usually
fail to comprehensively address the optimal and quick regeneration requirements. The
synthetic polymer derived dressings are resistant to physical and chemical degradation
which causes deterioration of the environment through solid waste landfills. Despite
several advances in chronic wound management, natural product-based dressings with
high exude absorption and porous nature are still a hotspot in the medical field. The
animal derived commercial skin grafts have the risk of immune activation and disease
transmission along with costlier purification steps involved. The properties of a
dressing are primarily determined by its matrix composition so wound care domain is
focusing on the biomaterial used for fabrication of the dressings in addition to the
architecture of the dressings. Natural polymer-based dressings have gained
considerable attention. Natural polymers exhibit biocompatibility, natural abundance,
biological simulation, and steric configuration similar to ECM thereby rendering
effective cell-ECM interaction, signal transportation, and wound healing. The main
drawback associated with the use of natural polymers is their limited mechanical
strength and structural instability which can be overcome by blending it with a suitable
synthetic polymer. A variety of complex natural polymers such as carbohydrates are
extracted from bacteria (agrobacterium, etc.), seaweeds that have been used for
different biomedical applications. Recently, the carbohydrates (agarose, curdlan,
alginate, chitosan, etc.) derived dressings have upsurged within the healthcare industry
because of their biocompatible, biodegradable, antimicrobial, antioxidant, antiviral,
anticoagulant, and antidiabetic nature. Moreover, these possess peptide sequences on

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their surface as cell recognition sites and have increasingly emerged as substitute for
animal derived commercial skin grafts having the risk of immune activation and disease
transmission along with costlier purification steps involved
Agarose being a natural polysaccharide has been widely used for chromatography,
tissue engineering, electrophoresis, and regenerative medicine (angiogenesis,
spermatogenesis, neurogenesis, and wound healing, etc.). It possesses excellent
mechanical properties, swelling, biodegradability, biocompatibility, and natural
abundance. It also allows permeation of nutrients and oxygen, which can be controlled
as per the requirements. Moreover, it is an inert biopolymer, so it does not evoke the
immune system. In diabetic wounds, prolonged inflammation due to elevation of the

pro-inflammatory cytokines (IL-6, TNF-α and IL-1β) and downregulation of anti-
inflammatory cytokines (TGF-β and IL-10) results in delayed healing. Hence, the use

of biological response modifiers (BRM) like β-D glucans becomes important to combat

the prolonged inflammation and enhance the immune system. Curdlan (1,3-β-D-
glucopyranan) is a neutral exopolysaccharide that has been approved as food additive

(stabilizer, thickener, and texturizer) by Food and Drug Administration (FDA), United
States. It has gained attention in biomedical industry due to its antiviral, anti-tumor,
antibacterial, anti-coagulatory, immunomodulatory, and wound healing properties. It
can interact with the dectin-1 receptor on macrophages and thus hasten the migration,
and proliferation of keratinocytes along with the re-epithelialization of wounds.
In the current study, agarose (10% w/v)/ PVA, Polyvinyl alcohol (12% w/v) based
multifunctional nanofibrous electrospun dressings were fabricated to address chronic
wounds. Zinc citrate (1, 3 and 5% w/w of the polymer) was used as a potential
antibacterial agent. The fabricated dressings exhibit a swelling of ~550% in Phosphate

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buffer saline (PBS) and mechanical strength of 10.11 ± 0.31MPa which is suitable for
most of the wound healing applications that require high strength. In vitro study
revealed an increased migration and proliferation of L929 fibroblasts with agarose
blends when compared to the control. The fabricated dressings exhibited antibacterial
properties against both Staphylococcus aureus (Gram-positive) and Escherichia coli
(Gram-negative) bacterial strains. Hence, a potential dressing with multifunctional
attributes such as ability to protect wounds from bacterial infections, effective swelling,
natural product based, and eco-friendly nature was fabricated.
In the second study, agarose/curdlan/polyvinyl alcohol based nanofibrous dressings
loaded with ciprofloxacin (0, 1, 3, and 5 wt.%) were fabricated using an electrospinning
technique with water and formic acid as solvents. In vitro evaluation revealed the
average diameter of the fabricated nanofibers were between 115-146 nm with high
swelling (~450-500%) properties. They exhibited enhanced mechanical strength (7.46
± 0.80 MPa -7.79 ± 0.007 MPa) and significant biocompatibility (~90-98%) with L929
and NIH 3T3 mouse fibroblasts. In vitro scratch assay showed higher proliferation and
migration of fibroblasts (~ 90-100% wound closure) when compared to electrospun
PVA and control. Significant antibacterial activity was observed against Escherichia
coli and Staphylococcus aureus. In vitro real-time gene expression studies with human
THP-1 cell line revealed a significant downregulation of pro-inflammatory cytokines
(8.64 fold decrease for TNF- α) and upregulation of anti-inflammatory cytokines (6.83
fold increase for IL-10) when compared to lipopolysaccharide. In brief, the results
advocate agarose-curdlan mat as a potential multifunctional, bioactive, and eco-friendly
dressing for healing diabetic wounds.

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Full-thickness diabetic wounds are chronic injuries characterized by bleeding,
excessive exude, and prolonged inflammation. Single-layer dressings fail to address
such disturbed pathophysiology. Therefore, bilayer dressings with structural and
compositional differences in each layer have gained attention. In full-thickness/deep
wounds dressing will be in contact with the blood, thus it should have haemostatic
agents also. Alginate is a carbohydrate which has great potential for haemostatic
applications and has been used extensively for various biomedical applications. It
hastens the wound healing process and reduces bacterial growth by maintaining moist
conditions. Sodium alginate (SA) have been used as a surgical sealant for effectively
sealing the bleeding wounds and hasten tissue regeneration. The crosslinked SA
hydrogels have been successfully used for the release of bioactive and other tissue
engineering applications. Also, SA derived gels have been used in large scale for wound
repair and healing due to their biocompatibility, hydrophilicity, haemostatic potential
and antimicrobial properties. However, hemocompatibility is the essential criteria
which limits the blood contacting biomaterials in clinical settings. The fabricated
materials during their application comes in contact with blood comprising of plasma
(55%), erythrocytes (44%), leukocytes (1%) and platelets. Hence, the adverse reactions
which might arise due to interaction between the blood and fabricated material needs
to be studied extensively.
In the third study we incorporated alginate along with earlier described polymers to
fabricate agarose/curdlan/alginate/polyvinyl alcohol-based bilayer dressings. The
bilayer dressings were fabricated by electrospinning agarose/curdlan/polyvinyl alcohol
blend (top layer) on an alginate/agarose/polyvinyl alcohol-based lyophilized porous
layer. Ciprofloxacin was incorporated in both layers as a potential antibacterial agent.

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The fabricated dressing exhibited high swelling (~1300%), biocompatibility (>90%
with NIH 3T3 and L929 mouse fibroblasts), and hemocompatibility (hemolysis