Carboxymethyl cellulose-agarose hydrogel in poly(3-hydroxybutyrate-co-3-hydroxyvalerate) nanofibers: A novel tissue engineered skin graft.

Healing chronic and critical-sized full-thickness wounds is a major challenge in the healthcare sector. Scaffolds prepared using electrospinning and hydrogels serve as effective treatment options for wound healing by mimicking the native skin microenvironment. Combining synthetic nanofibers with tunable hydrogel properties can effectively overcome limitations in skin scaffolds made only with nanofibers or hydrogels. In this study, a biocompatible hybrid scaffold was developed for wound healing applications using poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) nanofibers embedded with hydrogel made of 2 % carboxymethyl cellulose (CMC) blended with 1 % agarose. Hybrid scaffolds, characterized for surface morphology, swellability, porosity, and degradation, were found to be suitable for wound healing. Furthermore, the incorporation of CMC-agarose hydrogel into nanofibers significantly enhanced their mechanical strength compared to PHBV nanofibers alone (p < 0.05). Extract cytotoxicity and direct cytotoxicity tests showed that the hybrid scaffolds developed in this study are cytocompatible (>75 % viability). Furthermore, human adult dermal fibroblasts (HDFa) and human adult immortalized keratinocytes (HaCaT) adhesion, viability, and proliferation studies revealed that the hybrid scaffolds exhibited a significant increase in cell proliferation over time, similar to PHBV nanofibers. Finally, the developed hybrid scaffolds were evaluated in rat full-thickness wounds, demonstrating their ability to promote full-thickness wound healing with reepithelialization and epidermis closure.

Tuning thermoresponsive properties of carboxymethyl cellulose (CMC)-agarose composite bioinks to fabricate complex 3D constructs for regenerative medicine.

3D bioprinting has emerged as a viable tool to fabricate 3D tissue constructs with high precision using various bioinks which offer instantaneous gelation, shape fidelity, and cytocompatibility. Among various bioinks, cellulose is the most abundantly available natural polymer & widely used as bioink for 3D bioprinting applications. To mitigate the demanding crosslinking needs of cellulose, it is frequently chemically modified or blended with other polymers to develop stable hydrogels. In this study, we have developed a thermoresponsive, composite bioink using carboxymethyl cellulose (CMC) and agarose in different ratios (9:1, 8:2, 7:3, 6:4, and 5:5). Among the tested combinations, the 5:5 ratio showed better gel formation at 37 °C and were further characterized for physicochemical properties. Cytocompatibility was assessed by in vitro extract cytotoxicity assay (ISO 10993-5) using skin fibroblasts cells. CMC-agarose (5:5) bioink was successfully used to fabricate complex 3D structures through extrusion bioprinting and maintained over 80 % cell viability over seven days. Finally, in vivo studies using rat full-thickness wounds showed the potential of CMC-agarose bulk and bioprinted gels in promoting skin regeneration. These results indicate the cytocompatibility and suitability of CMC-agarose bioinks for tissue engineering and 3D bioprinting applications.

Oral administration of eugenol oleate cures experimental visceral leishmaniasis through cytokines abundance.

Visceral leishmaniasis (VL) is an endemic fatal infectious disease in tropical and subtropical nations. The limited treatment options, long treatment regimens, invasive mode of administration of drugs, and lack of effective vaccination are the main reasons for the search of new alternative therapeutics against VL. On this quest, from a series of eugenol derivatives, we had demonstrated eugenol oleate as a lead immunomodulatory anti-VL molecule earlier. In this report, the oral efficacy and mechanism of eugenol oleate in inducing immunomodulatory anti-VL activity has been studied in BALB/c mice model. The plasma pharmacokinetic and acute toxicity studies suggested that the eugenol oleate is safe with an appreciable pharmacokinetic profile. Eugenol oleate (30 mg/kg B.W.) showed 86.5% of hepatic and 84.1% of splenic parasite clearance. The increased Th1 cytokine profile and decreased Th2 cytokine profile observed from ELISA and qRTPCR suggested that the eugenol oleate induced the parasite clearance through the activation of the host immune system. Subsequently, the mechanistic insights behind the anti-leishmanial activity of eugenol oleate were studied in peritoneal macrophages in vitro by inhibitor response study and immunoblotting. The results inferred that eugenol oleate activated the PKC-ßII-p38 MAPK and produced IL-12 and IFN-γ which intern activated the iNOS2 to produce NO free radicals that cleared the intracellular parasite.

Eugenol derived immunomodulatory molecules against visceral leishmaniasis.

Visceral leishmaniasis (VL) is a life threatening infectious disease caused by Leishmania donovani. It leads to the severe immune suppression in the host defense system. Higher cytotoxicity, rigorous side effects and lower therapeutic indexes (TI) of current antileishmanial drugs have created a necessity to develop new molecules with better antileishmanial activity and high TI value. In this study, we have synthesized 36 derivatives of eugenol and screened them for their activity against promastigote and amastigote forms of L. donovani. Among the synthesized derivatives, comp.35 showed better antileishmanial activity against extra cellular promastigotes (IC50- 20.13 ± 0.91 µM) and intracellular amastigotes (EC50-4.25 ± 0.26 µM). The TI value (82.24 ± 3.77) was found to improve by 10-13 fold compared to Amphotericin B and Miltefosine respectively. Treatment with comp.35 (5 µg/ml) enhanced the nitric oxide (NO) generation, iNOS2 mRNA expression (∼8 folds increase) and decreased the arginase-1 activity (∼4 folds) in L. donovani infected peritoneal macrophages. Comp.35 had also increased the IL-12 (∼6 folds) and decreased the IL-10 (∼3 folds) mRNA expression and release in vitro. Results of in vivo studies revealed that comp.35 treatment at 25 mg/kg body weight efficiently cleared the hepatic and splenic parasite burden with enhanced Th1 response in L. donovani infected BALB/c mice. Hence, this study clearly represents comp.35, as an immunomodulatory molecule, can induce host protective immune response against visceral leishmaniasis through enhanced NO generation and Th1 response, which are essentials against this deadly disease.