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Neurosurgical procedures are the key therapeutic interventions for the cerebral hemorrhage and brain tumors. However, neurosurgical procedures inevitably cause surgical brain injury (SBI), which will induce hemorrhage and inflammation. Gelatin Sponges are still the primary hemostatic materials used in clinical, but their anti-inflammatory efficacy is poor. Herein, we developed a cross-linked gelatin hydrogel (GelMA) to load mesenchymal stem cells (MSC) and directly implant them to the SBI site. Upon contacting the SBI site, the GelMA showed better clotting performance than Gelatin Sponges. Moreover, the MSC can reduce oxidative stress and enhance mitochondrial fusion via mitochondria transfer, resulting in ameliorating mitochondrial damage and reducing inflammation. Thus, the GelMA containing MSC can effectively reduce brain edema and inflammation and improve neurological function in SBI mouse models. In addition, GelMA exhibits excellent hemocompatibility and low cytotoxicity. It also enhances the proliferation of MSCs and decelerates the rapid depletion of MSCs. Therefore, MSC-loaded GelMA exhibits excellent hemostatic and anti-inflammatory effects, making it a potential new-generation biomaterial for SBI., Competing Interests: Declaration of competing interest The authors declare no competing financial interest. Additionally, we would like to thank the imaging platform in core facility of the First Hospital of Jilin University, for their essential support and services, especially the confocal microscope, which significantly contributed to our research., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) is incurable and raises growing concerns. The main barrier to nerve repair is the complicated inhibitory microenvironment, where single-targeted strategies are largely frustrated. Despite the progress in combinatory therapeutic systems, the development and translation of effective therapies remain a challenge with extremely limited clinical materials. In this study, mesenchymal stem cells are transplanted in combination with sustained release of methylprednisolone through delivery in one composite matrix of a microsphere-enveloped adhesive hydrogel. All the materials used, including the stem cells, drug, and the matrix polymers gelatin and hyaluronan, are clinically approved. The therapeutic effects and safety issues are evaluated on rat and canine SCI models. The implantation significantly promotes functional restoration and nerve repair in a severe long-span rat spinal cord transection model. Distant spinal cord segments and the urinary system are effectively protected against pathologic damage. Moreover, the local sustained drug delivery mitigates the inflammatory microenvironment when overcoming the clinical issue of systemic side effects. The study presents an innovative strategy to achieve safe and efficient combinatory treatment of SCI., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

In this study, a novel macroporous composite biomaterial consisting of gelatin-hydroxyapatite-calcium sulphate for delivery of bone morphogenic protein-2 (rhBMP-2) and zoledronic acid (ZA) has been developed. The biomaterial scaffold has a porous structure and functionalization of the scaffold with rhBMP-2 induces osteogenic differentiation of MC3T3-e1 cells seen by a significant increase in biochemical and genetic markers of osteoblastic differentiation. In-vivo muscle pouch experiments showed higher mineralization using scaffold+rhBMP-2 when compared to an approved absorbable collagen sponge (ACS)+rhBMP-2 as verified by micro-CT. Co-delivery of rhBMP-2+ZA via the novel scaffold enabled a reduction in the effective rhBMP-2 doses. The presence of tartrate resistant acid phosphatase staining in the rhBMP-2 group indicates osteoclastic resorption, which could be stalled by adding ZA, which by speculation could explain the net increase in mineralization. The new scaffold allowed for slow release of rhBMP-2 in-vitro (3.3±0.1%) after 4weeks. Using single photon emission computed tomography (SPECT), the release kinetics of 125 I-rhBMP-2 in-vivo was followed for 4weeks and a total of 65.3±15.2% 125 I-rhBMP-2 was released from the scaffolds. In-vitro 14 C-ZA release curve shows an initial burst release on day 1 (8.8±0.7%) followed by a slow release during the following 4weeks (13±0.1%). In-vivo, an initial release of 43.2±7.6% of 14 C-ZA was detected after 1day, after which the scaffold retained the remaining ZA during 4-weeks. Taken together, our results show that the developed biomaterial is an efficient carrier for spatio-temporal delivery of rhBMP-2 and ZA leading to increased bone formation compared to commercially available carrier for rhBMP-2., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Administration of dendritic cells (DCs) combined with oncolytic adenovirus (Ad) expressing antitumor cytokines induces a potent antitumor effect and antitumor immunity by ameliorating the immunosuppressive tumor microenvironment. However, this combination therapy has significant limitations due to rapid dissemination and inactivation of the therapeutics at the tumor site, necessitating multiple injections of both therapeutics. To overcome these limitations, we have utilized gelatin-based hydrogel to co-deliver oncolytic Ad co-expressing interleukin (IL)-12 and granulocyte-macrophage colony-stimulating factor (GM-CSF) (oAd) and DCs for sustained release of both therapeutics. The injectable and biodegradable hydrogels were prepared by mixing the polymer solutions containing horseradish peroxidase and hydrogen peroxide. Gel matrix enabled sustained release of both oAd and DCs while preserving their biological activity over a considerable time period, leading to efficient retention of both therapeutics in tumor tissue. Further, tumors treated with oAd- and DC-loaded gel (oAd+DC/gel) showed a significantly greater expression level of IL-12, GM-CSF, and interferon-γ (IFN-γ) than either single treatment (oAd or DC) or oAd in combination with DC (oAd+DC), resulting in efficient activation of both endogenous and exogenous DCs, migration of DCs to draining lymph nodes, and tumor infiltration of CD4 + and CD8 + T cells. Moreover, oAd+DC/gel resulted in a significantly higher number of tumor-specific IFN-γ-secreting immune cells compared with oAd+DC. Lastly, oAd+DC/gel significantly attenuated tumor-mediated thymic atrophy, which is associated with immunosuppression in the tumor microenvironment, compared with oAd+DC. Taken together, these results demonstrate that gelatin gel-mediated co-delivery of oncolytic Ad and DCs might be a promising strategy to efficiently retain both therapeutics in tumor tissue and induce a potent antitumor immune response for an extended time period via a single administration., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Self-assembling cell sheets have shown great potential for use in cartilage tissue engineering applications, as they provide an advantageous environment for the chondrogenic induction of human mesenchymal stem cells (hMSCs). We have engineered a system of self-assembled, microsphere-incorporated hMSC sheets capable of forming cartilage in the presence of exogenous transforming growth factor β1 (TGF-β1) or with TGF-β1 released from incorporated microspheres. Gelatin microspheres with two different degrees of crosslinking were used to enable different cell-mediated microsphere degradation rates. Biochemical assays, histological and immunohistochemical analyses, and biomechanical testing were performed to determine biochemical composition, structure, and equilibrium modulus in unconfined compression after 3 weeks of culture. The inclusion of microspheres with or without loaded TGF-β1 significantly increased sheet thickness and compressive equilibrium modulus, and enabled more uniform matrix deposition by comparison to control sheets without microspheres. Sheets incorporated with fast-degrading microspheres containing TGF-β1 produced significantly more GAG and GAG per DNA than all other groups tested and stained more intensely for type II collagen. These findings demonstrate improved cartilage formation in microsphere-incorporated cell sheets, and describe a tailorable system for the chondrogenic induction of hMSCs without necessitating culture in growth factor-containing medium., (Copyright © 2011 Elsevier B.V. All rights reserved.)

RNA interference using small interfering RNA (siRNA) is a promising biological strategy for treatment of diverse diseases; however, application of siRNA is severely hindered by its poor stability and low cellular uptake efficiency. We have recently demonstrated that polyethyleneimine (PEI)-based amphiphilic core-shell particles have several distinguishing advantages over native PEI and its derivatives. This paper presents a novel type of PEI-based nanogels with a biodegradable gelatin core. The core-shell nanogels were synthesized via a two-stage reaction: (1) preparation of highly uniform gelatin nanoparticles through appropriate treatment of gelatin solution; and (2) conjugation of branched PEI to the preformed gelatin nanoparticles, followed by repeated cycles of desolvation and drying of the gelatin-PEI nanogels in ethanol/water mixture. The resulting nanogels have a well-defined nanostructure that contains a gelatin core and a PEI shell. They have an average diameter of 200 ± 40nm with high uniformity. The nanogel particles possess positive zeta-potential values of up to +40mV at neutral pH, indicating that they are highly positive and very stable in aqueous medium. The gelatin-PEI nanogels were able to completely condense siRNA at N/P ratios of as low as 5:1, and effectively protected siRNA against enzymatic degradation. Furthermore, the nanogels were four times less toxic than native PEI. Besides low toxicity, the nanogels were able to effectively deliver siRNA into HeLa cells. It was found that increasing the N/P ratio from 10 to 30 significantly increased the intracellular uptake efficiency of siRNA from 41 to 84%. Confocal laser scanning microscopic images confirmed that the nanogels were able to effectively deliver siRNA in the cytoplasm of HeLa cells. The delivered siRNA could inhibit 70% of human argininosuccinate synthetase 1 (ASS1) gene expression. This gene silencing percentage is much higher than that of the commercial Lipofectamine(TM) 2000. Our studies demonstrate that gelatin-PEI core-shell nanogels have promising potential to act as an effective siRNA carrier., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Wounds have the potential to become infected during any surgical procedure. Gelatin sponges that are commonly used to absorb blood during invasive surgeries would benefit tremendously if they released antibiotics. In this work, we have examined coating a commercial gelatin sponge with degradable polymer multilayer films containing vancomycin. The effect of the film on sponge absorption capabilities and the effect of the sponge on drug release kinetics were both examined. Application of vancomycin containing layer-by-layer assembled films to this highly porous substrate greatly increased drug loading up to approximately 880% compared to a flat substrate. Vancomycin drug release was extended out to 6 days compared to 2 days for film coated flat substrates. Additionally, the absorbent properties of the gelatin sponge were actually enhanced by up to 170% due to the presence of the vancomycin film coating. A comparison of film coated sponges with sponges soaked directly in vancomycin demonstrated the ability of the multilayer films to control drug release. Film released vancomycin was also found to remain highly therapeutic with unchanged antimicrobial properties compared to the neat drug, demonstrated by quantifying vancomycin activity against Staphylococcus aureus in vitro., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Catalase delivery can be effective in inhibiting reactive oxygen species (ROS)-mediated acceleration of tumor metastasis. Our previous studies have demonstrated that increasing the plasma half-life of catalase by pegylation (PEG-catalase) significantly increases its potency of inhibiting experimental pulmonary metastasis in mice. In the present study, a biodegradable gelatin hydrogel formulation was used to further increase the circulation time of PEG-catalase. Implantation of (111)In-PEG-catalase/hydrogel into subcutaneous tissues maintained the radioactivity in plasma for more than 14 days. Then, the effect of the PEG-catalase/hydrogel on spontaneous pulmonary metastasis of tumor cells was evaluated in mice with subcutaneous tumor of B16-BL6/Luc cells, a murine melanoma cell line stably expressing luciferase. Measuring luciferase activity in the lung revealed that the PEG-catalase/hydrogel significantly (P<0.05) inhibited the pulmonary metastasis compared with PEG-catalase solution. These findings indicate that sustaining catalase activity in the blood circulation achieved by the use of pegylation and gelatin hydrogel can reduce the incidence of tumor cell metastasis.

Injectable and thermo-reversible physical combination gels were formed in aqueous solution by preparing complex coacervate with two oppositely charged biomacromolecules that composed of negatively charged chondroitin 6-sulfate and positively charged high molecular weight gelatin type A and co-formulating with a negative, thermo-sensitive polysaccharide, methylcellulose containing a salting-out salt, ammonium sulfate. The combination of complex coacervation and a thermo-reversible gel demonstrated synergistic effects on the complex coacervate formation the release rates of model proteins and in situ gel depot formation. Gels indicated sustained release patterns of the protein over 25 days with minimal initial bursts. Optimized novel in situ gel depot systems containing dual advantages of complex coacervation and temperature responsiveness demonstrated a potential for efficient protein drug delivery in terms of high protein loading, sustained protein release, ease of administration, an aqueous environment without toxic organic solvents, and a simple fabrication method.

Aminated gelatin microspheres (AGMS) was investigated as a nasal drug delivery system for peptide drugs. The in vitro drug release from microspheres was evaluated using a fluorescein-labeled insulin (RITC-insulin) and FITC-dextran with a molecular weight of 4.4 kDa (FD-4) as model drugs. RITC-insulin release from AGMS was significantly slower than from native gelatin microspheres (GMS), with a cumulate release of 18.4% and 32.4% within 30 min, and 56.9% and 75.1% within 8 h respectively. However, the release of FD-4 from both AGMS and GMS was quite rapid and no difference was observed for the two microspheres. The electrostatic interactions between the model drugs and the microspheres were supposed to be the main factor that controlled the release behavior. The absorption enhancing effect was estimated by measuring the changes of plasma glucose concentrations of healthy rats following intranasal administration of insulin-incorporated microspheres in both suspension and dry powder forms. AGMS could significantly increase the nasal absorption of insulin in rats when administered in a dry powder formulation, but no significant hypoglycemic effect was observed when given in suspensions. One of the mechanisms for the increased insulin absorption was attributed to the hydrogel nature of the microspheres that could absorb water from the nasal mucosa and thus resulted in a temporarily dehydration of the epithelium membrane and opening of the tight junctions. The positive charge of the AGMS has also evidently contributed to the absorption enhancing effect. In addition, the mucoadhesive properties of AGMS might also have played a role to the total effect. AGMS might be a new candidate carrier for the nasal delivery of peptide drugs.

This work describes the development of a non-invasive means of simultaneously delivering insulin-like growth factor-1 (IGF-1) and transforming growth factor-beta1 (TGF-beta1) to injured cartilage tissue in a controlled manner. This novel delivery technology employs the water-soluble polymer, oligo(poly(ethylene glycol) fumarate) (OPF), in the fabrication of biodegradable hydrogels which encapsulate gelatin microparticles. Release studies first examined the effect of gelatin isoelectric point (IEP) and crosslinking extent on IGF-1 release from these microparticles. In the presence of collagenase, highly crosslinked, acidic gelatin (IEP=5.0) provided sustained release of IGF-1, 95.2+/-2.9% cumulative release at day 28, while less crosslinked microparticles and microparticles of alternate IEP exhibited similar release values after only 6 days. Encapsulation of these highly crosslinked microparticles in a network of OPF provided a means to further control release, reducing final cumulative release to 70.2+/-4.7% in collagenase-containing PBS. Final release values from OPF-gelatin microparticle composites could be altered by incorporating less crosslinked, non-loaded microparticles within these constructs. Finally, this technology was extended to the dual delivery of IGF-1 and TGF-beta1 by loading these growth factors into either the OPF hydrogel phase or gelatin microparticle phase of composites. Release profiles were successfully manipulated by altering the phase of growth factor loading and microparticle crosslinking extent. For instance, by loading TGF-beta1 into the gelatin microparticle phase, a burst release of 10.8+/-0.7% was achieved, while loading this growth factor into the OPF hydrogel phase resulted in a burst release of 25.2+/-1.5%. With either system, simultaneous, slow release of IGF-1 over a 4-week period was accomplished by selectively loading this protein into highly crosslinked, encapsulated microparticles. These results demonstrate the utility of these systems in future studies to assess the interplay and time course of multiple growth factors in cartilage repair.

The crosslinking process of natural macromolecules with microwave energy should have the potentiality to overcome the problems due to the toxicity of the residuals of chemical crosslinking agents and moreover of the "in vivo" biodegradation products of the chemical crosslinked macromolecule. To evaluate the effective crosslinking of the gelatin forming the microspheres, the water-soluble fraction at 37 degrees C, the water absorption capability, the free amino and free carboxylic acid groups of the gelatin were determined. The structural change in the gelatin microspheres has been detected by the porosity studies. Moreover, both the "in vitro" biodegradability and the biocompatibility of the gelatin microspheres microwave-treated after a subcutaneous injection into female albino guinea pigs were tested. As the results suggest only the gelatin microspheres microwave-treated for 10 min at an inlet temperature of 250 degrees C could have been modified by the crosslink formation among the macromolecular chains. The gelatin microspheres treated with the microwave energy were very well biodegraded as indicated both by the "in vitro" enzymatic degradation studies and mainly by the histopathological examination. This latter study has also demonstrated the biocompatibility of the gelatin microspheres crosslinked with the microwave energy. In order to evaluate the feasibility of the microwave crosslinking process for pharmaceutical applications, both the drug loading and the drug release processes were evaluated using diclofenac as drug model, either as acidic form or as sodium salt. The microspheres were swollen in aqueous solution of diclofenac sodium salt, followed by a washing procedure with cool water to maintain the sodium salt into the microspheres or with pH 1.5 HCl to induce the diclofenac precipitation. To increase the amount of diclofenac acid form in the microspheres, the procedure was repeated three times washing with pH 1.5 HCl after each swelling process. Both the X-ray diffractometry and thermal analysis investigations showed a different physical state of the two drug forms in the microspheres, i.e. the amorphous state of the sodium salt and the crystalline state of the acidic form. According to the experimental results, the drug is released from gelatin microspheres according to the drug loading and the drug solubility.

The aim of present study was to prepare nanometer sized particles of gelatin via water-in-oil microemulsion system for drug and gene delivery applications. In this study, cross-linked gelatin nanoparticles encapsulating a fluorescent marker molecule fluorescein isothiocyanate-dextran (FITC-Dex, Mol. Wt. 19.3kDa) have been prepared, characterized and their influence on human fibroblasts has been assessed in terms of cell adhesion, cytotoxicity, light microscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM) and observation of cytoskeleton organisation. Gelatin nanoparticles were prepared inside the aqueous cores of sodium bis(2-ethylhexyl) sulfosuccinate (AOT)/n-hexane reverse micelles. Transmission electron microscopy image showed that the particles are spherical in shape with size of 37+/-0.84 nm diameter. The release of FITC-Dex from the nanoparticles in phosphate buffer saline (pH 7.4) is found to increase with time and about 80% of the encapsulated dye is released in 6 h. Cell adhesion studies with human fibroblasts have shown that gelatin nanoparticles do not affect the number of cells adhered to glass as compared to control cells with no particles. Standard cell viability assay demonstrated that cells incubated with gelatin nanoparticles remained more than 100% viable at concentration as high as 500 microg/ml. From SEM image, it was observed that the nanoparticles were internalised and the fibroblasts exhibited vacuoles in the cell body with cell membrane abnormalities. Endocytosis of nanoparticles was confirmed from TEM studies and it resulted in disruption of F-actin and beta-tubulin cytoskeleton. These studies show that the gelatin nanoparticles prepared by water-in-oil microemulsion systems are endocytosed by the fibroblasts without being toxic to cells even at high concentration of nanoparticles.

The gastric mucoadhesive properties of aminated gelatin microspheres were evaluated both in vitro and in vivo. The interactions of gelatin, aminated gelatin and microspheres with two kinds of commercial mucin were estimated in aqueous media. At a higher mucin concentration, aminated gelatin demonstrated a stronger interaction with mucin than either kind of the gelatin (isoelectric point (IEP): 5.0 and 9.0) under the same condition, although these interactions varied with varying media. At the same time, a larger amount of mucin was adsorbed to aminated gelatin microspheres than to either of the gelatin microspheres in the same condition. In the in vitro model of isolated and perfused rat stomach, the amount of aminated gelatin microspheres that remained in the stomach after perfusion was significantly larger than that of gelatin microspheres. However, no significant difference was observed whether the test was performed in simulated gastric fluid (SGF) or in phosphate-buffered saline (PBS, pH7.4). In the in vivo experiment, about 47% of the aminated gelatin microspheres remained in the stomach 2 h after oral administration in a capsule, whereas it was 29 and 34% for gelatin (IEP=5.0) and gelatin (IEP=9.0) microspheres, respectively. These results indicated that aminated gelatin microspheres demonstrated a higher gastric mucoadhesive ability than gelatin microspheres. The higher amino group content, improved chain flexibility and favorable polymer conformation were suggested to be the main factors that contributed to the stronger mucoadhesive properties of aminated gelatin microspheres than that of gelatin microspheres.