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IntroductionNanofibrous spheres, with their injectable format and biomimetic three-dimensional topologies that emulate the complexity of natural extracellular environments, have become increasingly attractive for applications in biomedical and regenerative medicine. Our research contributes to this growing field by detailing the design and fabrication of a novel series of polylactic acid/nano-hydroxyapatite (PLA/nHA) hybrid nanofibrous spheres.MethodsThese advanced structures were created by integrating electrospinning and electrospray techniques, which allowed for precise control over the nanofibrous spheres, especially in size. We have conducted a comprehensive investigation into the nanofibrous spheres’ capacity to deliver stem cells efficiently and maintain their viability post-implantation, as well as their potential to induce osteogenic differentiation.Results and DiscussionThe results show that these nanofibrous spheres are biocompatible and injectable, effectively supporting the attachment, growth, and differentiation of bone marrow-derived mesenchymal stem cells while aiding in their targeted transportation to bone defect areas to execute their regenerative functions. The findings of this study could significantly impact the future development of biocompatible materials for a range of therapeutic applications, including bone tissue engineering and regenerative therapy.

Recent advances in the field of cell therapy have proposed new solutions for tissue repair and regeneration using various cell delivery approaches. Here we studied ex vivo a novel topical delivery system of encapsulated cells in hybrid polyethylene glycol-fibrinogen (PEG-Fb) hydrogel microspheres to respiratory tract models. We investigated basic parameters of cell encapsulation, delivery and release in conditions of inflamed and damaged lungs of bacterial-infected mice. The establishment of each step in the study was essential for the proof of concept. We demonstrated co-encapsulation of alveolar macrophages and epithelial cells that were highly viable and equally distributed inside the microspheres. We found that encapsulated macrophages exposed to bacterial endotoxin lipopolysaccharide preserved high viability and secreted moderate levels of TNFα, whereas non-encapsulated cells exhibited a burst TNFα secretion and reduced viability. LPS-exposed encapsulated macrophages exhibited elongated morphology and out-migration capability from microspheres. Microsphere degradation and cell release in inflamed lung environment was studied ex vivo by the incubation of encapsulated macrophages with lung extracts derived from intranasally infected mice with Yersinia pestis, demonstrating the potential in cell targeting and release in inflamed lungs. Finally, we demonstrated microsphere delivery to a multi-component airways-on-chip platform that mimic human nasal, bronchial and alveolar airways in serially connected compartments. This study demonstrates the feasibility in using hydrogel microspheres as an effective method for topical cell delivery to the lungs in the context of pulmonary damage and the need for tissue repair.

Introduction: Nanofibrous spheres, with their injectable format and biomimetic three-dimensional topologies that emulate the complexity of natural extracellular environments, have become increasingly attractive for applications in biomedical and regenerative medicine. Our research contributes to this growing field by detailing the design and fabrication of a novel series of polylactic acid/nano-hydroxyapatite (PLA/nHA) hybrid nanofibrous spheres., Methods: These advanced structures were created by integrating electrospinning and electrospray techniques, which allowed for precise control over the nanofibrous spheres, especially in size. We have conducted a comprehensive investigation into the nanofibrous spheres' capacity to deliver stem cells efficiently and maintain their viability post-implantation, as well as their potential to induce osteogenic differentiation., Results and Discussion: The results show that these nanofibrous spheres are biocompatible and injectable, effectively supporting the attachment, growth, and differentiation of bone marrow-derived mesenchymal stem cells while aiding in their targeted transportation to bone defect areas to execute their regenerative functions. The findings of this study could significantly impact the future development of biocompatible materials for a range of therapeutic applications, including bone tissue engineering and regenerative therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Wang, Zhang, Liu, Chen, Yu, Yao, Chen, Yan, Wu and Wang.)

Stem cell-based therapy appears as a promising strategy to induce regeneration of damaged and diseased tissues. However, low survival, poor engraftment and a lack of site-specificity are major drawbacks. Polysaccharide hydrogels can address these issues and offer several advantages as cell delivery vehicles. They have become very popular due to their unique properties such as high-water content, biocompatibility, biodegradability and flexibility. Polysaccharide polymers can be physically or chemically crosslinked to construct biomimetic hydrogels. Their resemblance to living tissues mimics the native three-dimensional extracellular matrix and supports stem cell survival, proliferation and differentiation. Given the intricate nature of communication between hydrogels and stem cells, understanding their interaction is crucial. Cells are incorporated with polysaccharide hydrogels using various microencapsulation techniques, allowing generation of more relevant models and further enhancement of stem cell therapies. This paper provides a comprehensive review of human stem cells and polysaccharide hydrogels most used in regenerative medicine. The recent and advanced stem cell microencapsulation techniques, which include extrusion, emulsion, lithography, microfluidics, superhydrophobic surfaces and bioprinting, are described. This review also discusses current progress in clinical translation of stem-cell encapsulated polysaccharide hydrogels for cell delivery and disease modeling (drug testing and discovery) with focuses on musculoskeletal, nervous, cardiac and cancerous tissues.

Severe skin wounds are often associated with large areas of damaged tissue, resulting in substantial loss of fluids containing electrolytes and proteins. The net result is a vulnerability clinically to skin infections. Therapies aiming to close these large openings are effective in reducing the complications of severe skin wounds. Recently, cell transplantation therapy showed the potential for rapid re-epithelialization of severe skin wounds. Here, we show the improved effects of cell transplantation therapy using a robust protocol of efficient expansion and delivery of epidermal cells for treatment of severe skin wounds. Human skin tissues were used to generate human epidermal organoids maintained under newly established culture conditions. The human epidermal organoids showed an improved capacity of passaging for at least 10 rounds, enabling organoids to expand to cell numbers required for clinical applications. A newly designed auto micro-atomization device (AMAD) was developed for delivery of human epidermal organoids onto the sites of severe skin wounds enhancing uniform and concentrated delivery of organoids, facilitating their engraftment and differentiation for skin reconstitution. With the optimal design and using pneumatic AMAD, both survival and functions of organoids were effectively protected during the spraying process. Cells in the sprayed human epidermal organoids participated in the regeneration of the epidermis at wound sites in a mouse model and accelerated wound healing significantly. The novel AMAD and out new protocol with enhanced effects with respect to both organoid expansion and efficient transplantation will be used for clincal treatments of complex, uneven, or large-area severe skin wounds.

BackgroundIntracoronary (IC) injection of mesenchymal stem cells (MSCs) results in a prompt decrease of absolute myocardial blood flow (AMF) with late and incomplete recovery of myocardial tissue perfusion. Here, we investigated the effect of decreased AMF on oxidative stress marker matrix metalloproteinase-2 (MMP-2) and its influence on the fate and homing and paracrine character of MSCs after IC or intramyocardial cell delivery in a closed-chest reperfused myocardial infarction (MI) model in pigs.MethodsPorcine MSCs were transiently transfected with Ad-Luc and Ad-green fluorescent protein (GFP). One week after MI, the GFP-Luc-MSCs were injected either IC (group IC, 11.00 ± 1.07 × 106) or intramyocardially (group IM, 9.88 ± 1.44 × 106). AMF was measured before, immediately after, and 24 h post GFP-Luc-MSC delivery. In vitro bioluminescence signal was used to identify tissue samples containing GFP-Luc-MSCs. Myocardial tissue MMP-2 and CXCR4 receptor expression (index of homing signal) were measured in bioluminescence positive and negative infarcted and border, and non-ischemic myocardial areas 1-day post cell transfer. At 7-day follow-up, myocardial homing (cadherin, CXCR4, and stromal derived factor-1alpha) and angiogenic [fibroblast growth factor 2 (FGF2) and VEGF] were quantified by ELISA of homogenized myocardial tissues from the bioluminescence positive and negative infarcted and border, and non-ischemic myocardium. Biodistribution of the implanted cells was quantified by using Luciferase assay and confirmed by fluorescence immunochemistry. Global left ventricular ejection fraction (LVEF) was measured at baseline and 1-month post cell therapy using magnet resonance image.ResultsAMF decreased immediately after IC cell delivery, while no change in tissue perfusion was found in the IM group (42.6 ± 11.7 vs. 56.9 ± 16.7 ml/min, p = 0.018). IC delivery led to a significant increase in myocardial MMP-2 64 kD expression (448 ± 88 vs. 315 ± 54 intensity × mm2, p = 0.021), and decreased expression of CXCR4 (592 ± 50 vs. 714 ± 54 pg/tissue/ml, p = 0.006), with significant exponential decay between MMP-2 and CXCR4 (r = 0.679, p

The retina is a highly sophisticated piece of the neural machinery that begins the translation of incoming light signals into meaningful visual information. Several degenerative diseases of the retina are characterized by photoreceptor loss and eventually lead to irreversible blindness. Regenerative medicine, using tissue engineering-based constructs to deliver progenitor cells or photoreceptors along with supporting carrier matrix is a promising approach for restoration of structure and function. Fresh fibrin glue (FG) produced by the CryoSeal®FS system in combination with mouse retinal progenitor cells (RPCs) were evaluated in this study. In vitro expanded RPCs isolated from postnatal mouse retina were encapsulated into FG and cultured in the presence of the protease inhibitor, tranexamic acid. Encapsulation of RPCs into FG did not show adverse effects on cell proliferation or cell survival. RPCs exhibited fibroblast-like morphology concomitantly with attachment to the encapsulating FG surface. They expressed α7 and β3 integrin subunits that could mediate attachment to fibrin matrix via an RGD independent mechanism. The three dimensional environment and the attachment surface provided by FG was associated with a rapid downregulation of the progenitor marker SOX2 and enhanced the expression of the differentiation markers CRX and recoverin. However, the in vitro culture conditions did not promote full differentiation into mature photoreceptors. Nevertheless, we have shown that autologous fibrin, when fabricated into a scaffold for RPCs for delivery to the retina, provides the cells with external cues that could potentially improve the differentiation events. Hence, transient encapsulation of RPCs into FG could be a valid and potential treatment strategy to promote retinal regeneration following degenerative diseases. However, further optimization is necessary to maximize the outcomes in terms of mature photoreceptors.

Recent advances in the field of cell therapy have proposed new solutions for tissue repair and regeneration using various cell delivery approaches. Here we studied ex vivo a novel topical delivery system of encapsulated cells in hybrid polyethylene glycol-fibrinogen (PEG-Fb) hydrogel microspheres to respiratory tract models. We investigated basic parameters of cell encapsulation, delivery and release in conditions of inflamed and damaged lungs of bacterial-infected mice. The establishment of each step in the study was essential for the proof of concept. We demonstrated co-encapsulation of alveolar macrophages and epithelial cells that were highly viable and equally distributed inside the microspheres. We found that encapsulated macrophages exposed to bacterial endotoxin lipopolysaccharide preserved high viability and secreted moderate levels of TNFα, whereas non-encapsulated cells exhibited a burst TNFα secretion and reduced viability. LPS-exposed encapsulated macrophages exhibited elongated morphology and out-migration capability from microspheres. Microsphere degradation and cell release in inflamed lung environment was studied ex vivo by the incubation of encapsulated macrophages with lung extracts derived from intranasally infected mice with Yersinia pestis , demonstrating the potential in cell targeting and release in inflamed lungs. Finally, we demonstrated microsphere delivery to a multi-component airways-on-chip platform that mimic human nasal, bronchial and alveolar airways in serially connected compartments. This study demonstrates the feasibility in using hydrogel microspheres as an effective method for topical cell delivery to the lungs in the context of pulmonary damage and the need for tissue repair., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Cohen, Vagima, Mouhadeb, Toister, Gutman, Lazar, Jayson, Artzy-Schnirman, Sznitman, Ordentlich, Yitzhaki, Seliktar, Mamroud and Epstein.)

Stem cell-based therapy appears as a promising strategy to induce regeneration of damaged and diseased tissues. However, low survival, poor engraftment and a lack of site-specificity are major drawbacks. Polysaccharide hydrogels can address these issues and offer several advantages as cell delivery vehicles. They have become very popular due to their unique properties such as high-water content, biocompatibility, biodegradability and flexibility. Polysaccharide polymers can be physically or chemically crosslinked to construct biomimetic hydrogels. Their resemblance to living tissues mimics the native three-dimensional extracellular matrix and supports stem cell survival, proliferation and differentiation. Given the intricate nature of communication between hydrogels and stem cells, understanding their interaction is crucial. Cells are incorporated with polysaccharide hydrogels using various microencapsulation techniques, allowing generation of more relevant models and further enhancement of stem cell therapies. This paper provides a comprehensive review of human stem cells and polysaccharide hydrogels most used in regenerative medicine. The recent and advanced stem cell microencapsulation techniques, which include extrusion, emulsion, lithography, microfluidics, superhydrophobic surfaces and bioprinting, are described. This review also discusses current progress in clinical translation of stem-cell encapsulated polysaccharide hydrogels for cell delivery and disease modeling (drug testing and discovery) with focuses on musculoskeletal, nervous, cardiac and cancerous tissues., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lee, Ma, Khoo, Abdullah, Nik Md Noordin Kahar, Abdul Hamid and Mustapha.)

Severe skin wounds are often associated with large areas of damaged tissue, resulting in substantial loss of fluids containing electrolytes and proteins. The net result is a vulnerability clinically to skin infections. Therapies aiming to close these large openings are effective in reducing the complications of severe skin wounds. Recently, cell transplantation therapy showed the potential for rapid re-epithelialization of severe skin wounds. Here, we show the improved effects of cell transplantation therapy using a robust protocol of efficient expansion and delivery of epidermal cells for treatment of severe skin wounds. Human skin tissues were used to generate human epidermal organoids maintained under newly established culture conditions. The human epidermal organoids showed an improved capacity of passaging for at least 10 rounds, enabling organoids to expand to cell numbers required for clinical applications. A newly designed auto micro-atomization device (AMAD) was developed for delivery of human epidermal organoids onto the sites of severe skin wounds enhancing uniform and concentrated delivery of organoids, facilitating their engraftment and differentiation for skin reconstitution. With the optimal design and using pneumatic AMAD, both survival and functions of organoids were effectively protected during the spraying process. Cells in the sprayed human epidermal organoids participated in the regeneration of the epidermis at wound sites in a mouse model and accelerated wound healing significantly. The novel AMAD and out new protocol with enhanced effects with respect to both organoid expansion and efficient transplantation will be used for clincal treatments of complex, uneven, or large-area severe skin wounds., (Copyright © 2020 Chang, Liu, Guo, Fang, Wang, Wang, Liu, Reid and Wang.)

Background: Intracoronary (IC) injection of mesenchymal stem cells (MSCs) results in a prompt decrease of absolute myocardial blood flow (AMF) with late and incomplete recovery of myocardial tissue perfusion. Here, we investigated the effect of decreased AMF on oxidative stress marker matrix metalloproteinase-2 (MMP-2) and its influence on the fate and homing and paracrine character of MSCs after IC or intramyocardial cell delivery in a closed-chest reperfused myocardial infarction (MI) model in pigs., Methods: Porcine MSCs were transiently transfected with Ad-Luc and Ad-green fluorescent protein (GFP). One week after MI, the GFP-Luc-MSCs were injected either IC (group IC, 11.00 ± 1.07 × 10 6 ) or intramyocardially (group IM, 9.88 ± 1.44 × 10 6 ). AMF was measured before, immediately after, and 24 h post GFP-Luc-MSC delivery. In vitro bioluminescence signal was used to identify tissue samples containing GFP-Luc-MSCs. Myocardial tissue MMP-2 and CXCR4 receptor expression (index of homing signal) were measured in bioluminescence positive and negative infarcted and border, and non-ischemic myocardial areas 1-day post cell transfer. At 7-day follow-up, myocardial homing (cadherin, CXCR4, and stromal derived factor-1alpha) and angiogenic [fibroblast growth factor 2 (FGF2) and VEGF] were quantified by ELISA of homogenized myocardial tissues from the bioluminescence positive and negative infarcted and border, and non-ischemic myocardium. Biodistribution of the implanted cells was quantified by using Luciferase assay and confirmed by fluorescence immunochemistry. Global left ventricular ejection fraction (LVEF) was measured at baseline and 1-month post cell therapy using magnet resonance image., Results: AMF decreased immediately after IC cell delivery, while no change in tissue perfusion was found in the IM group (42.6 ± 11.7 vs. 56.9 ± 16.7 ml/min, p  = 0.018). IC delivery led to a significant increase in myocardial MMP-2 64 kD expression (448 ± 88 vs. 315 ± 54 intensity × mm 2 , p  = 0.021), and decreased expression of CXCR4 (592 ± 50 vs. 714 ± 54 pg/tissue/ml, p  = 0.006), with significant exponential decay between MMP-2 and CXCR4 ( r  = 0.679, p  < 0.001). FGF2 and VEGF of the bioluminescence infarcted and border zone of homogenized tissues were significantly elevated in the IM goups as compared to IC group. LVEF increase was significantly higher in IM group (0.8 ± 8.4 vs 5.3 ± 5.2%, p  = 0.046) at the 1-month follow up., Conclusion: Intracoronary stem cell delivery decreased AMF, with consequent increase in myocardial expression of MMP-2 and reduced CXCR4 expression with lower level of myocardial homing and angiogenic factor release as compared to IM cell delivery.

The retina is a highly sophisticated piece of the neural machinery that begins the translation of incoming light signals into meaningful visual information. Several degenerative diseases of the retina are characterized by photoreceptor loss and eventually lead to irreversible blindness. Regenerative medicine, using tissue engineering-based constructs to deliver progenitor cells or photoreceptors along with supporting carrier matrix is a promising approach for restoration of structure and function. Fresh fibrin glue (FG) produced by the CryoSeal(®)FS system in combination with mouse retinal progenitor cells (RPCs) were evaluated in this study. In vitro expanded RPCs isolated from postnatal mouse retina were encapsulated into FG and cultured in the presence of the protease inhibitor, tranexamic acid. Encapsulation of RPCs into FG did not show adverse effects on cell proliferation or cell survival. RPCs exhibited fibroblast-like morphology concomitantly with attachment to the encapsulating FG surface. They expressed α7 and β3 integrin subunits that could mediate attachment to fibrin matrix via an RGD-independent mechanism. The three-dimensional environment and the attachment surface provided by FG was associated with a rapid down-regulation of the progenitor marker SOX2 and enhanced the expression of the differentiation markers cone-rod homeobox and recoverin. However, the in vitro culture conditions did not promote full differentiation into mature photoreceptors. Nevertheless, we have shown that autologous fibrin, when fabricated into a scaffold for RPCs for delivery to the retina, provides the cells with external cues that could potentially improve the differentiation events. Hence, transient encapsulation of RPCs into FG could be a valid and potential treatment strategy to promote retinal regeneration following degenerative diseases. However, further optimization is necessary to maximize the outcomes in terms of mature photoreceptors.