Rapamycin facilitates healing of the tendon-bone interface in an aging rat model of chronic rotator cuff injury.

BACKGROUND: Tendon-bone interface (TBI) healing in chronic rotator cuff injury (CRCI) in older individuals is a common clinical challenge due to cellular senescence, as well as decreased tissue repair and regeneration. Many studies have demonstrated the antiaging, improved tissue repair, and bone regeneration properties of rapamycin (RPM) in multiple age-related diseases. This study aimed to explore the effects of RPM on TBI healing after CRCI in an aging rat model. METHODS: A CRCI model was established in 60 Sprague-Dawley rats (24 months old). Rats were then randomly allocated into the control, 0.1 µg RPM, and 1 µg RPM groups. At 4 and 8 weeks postreconstructive surgery, the supraspinatus tendon-humerus complexes were harvested for biomechanical, microimaging, histological, and immunohistochemical evaluations. RESULTS: Biomechanical testing results demonstrated that the failure load, ultimate strength, and stiffness of the 2 RPM groups were significantly higher than those of the control group at 4 and 8 weeks postoperatively. Microradiographically, both RPM groups had significantly higher values of bone mineral density and the ratio of trabecular bone volume to total volume than controls at each time point. Moreover, the RPM groups had higher histological scores and showed better regenerated TBI, characterized by better organizational tissue, more fibrocartilage cells, and more bone formation. Immunohistochemical evaluations showed that RUNX2-, SOX9-, and SCX-positive cells were significantly more in the 2 RPM groups than in the controls at each time point. CONCLUSIONS: RPM may effectively enhance CRCI healing after reconstruction by facilitating osteogenesis, tenogenesis, and fibrocartilage reformation at the TBI, as well as improving biomechanical properties.

Scalable and high-throughput production of an injectable platelet-rich plasma (PRP)/cell-laden microcarrier/hydrogel composite system for hair follicle tissue engineering.

BACKGROUND: Tissue engineering of hair follicles (HFs) has enormous potential for hair loss treatment. However, certain challenges remain, including weakening of the dermal papilla cell (DPC) viability, proliferation, and HF inducibility, as well as the associated inefficient and tedious preparation process required to generate extracellular matrix (ECM)-mimicking substrates for biomolecules or cells. Herein, we utilized gelatin methacryloyl (GelMA) and chitosan hydrogels to prepare scalable, monodispersed, and diameter-controllable interpenetrating network GelMA/chitosan-microcarriers (IGMs) loaded with platelet-rich plasma (PRP) and seeded with DPCs, on a high-throughput microfluidic chip. RESULTS: The ECM-mimicking hydrogels used for IGMs exhibited surface nano-topography and high porosity. Mass production of IGMs with distinct and precise diameters was achieved by adjusting the oil and aqueous phase flow rate ratio. Moreover, IGMs exhibited appropriate swelling and sustained growth factor release to facilitate a relatively long hair growth phase. DPCs seeded on PRP-loaded IGMs exhibited good viability (> 90%), adhesion, spreading, and proliferative properties (1.2-fold greater than control group). Importantly, PRP-loaded IGMs presented a higher hair inducibility of DPCs in vitro compared to the control and IGMs group (p < 0.05). Furthermore, DPC/PRP-laden IGMs were effectively mixed with epidermal cell (EPC)-laden GelMA to form a PRP-loaded DPC/EPC co-cultured hydrogel system (DECHS), which was subcutaneously injected into the hypodermis of nude mice. The PRP-loaded DECHS generated significantly more HFs (~ 35 per site) and novel vessels (~ 12 per site) than the other groups (p < 0.05 for each). CONCLUSION: Taken together, these results illustrate that, based on high-throughput microfluidics, we obtained scalable and controllable production of ECM-mimicking IGMs and DECHS, which simulate an effective micro- and macro-environment to promote DPC bioactivity and hair regeneration, thus representing a potential new strategy for HF tissue engineering.

Electrospun Aligned Nanofiber Yarns Constructed Biomimetic M-Type Interface Integrated into Precise Co-Culture System as Muscle-Tendon Junction-on-a-Chip for Drug Development.

The incorporation of engineered muscle-tendon junction (MTJ) with organ-on-a-chip technology provides promising in vitro models for the understanding of cell-cell interaction at the interface between muscle and tendon tissues. However, developing engineered MTJ tissue with biomimetic anatomical interface structure remains challenging, and the precise co-culture of engineered interface tissue is further regarded as a remarkable obstacle. Herein, an interwoven waving approach is presented to develop engineered MTJ tissue with a biomimetic "M-type" interface structure, and further integrated into a precise co-culture microfluidic device for functional MTJ-on-a-chip fabrication. These multiscale MTJ scaffolds based on electrospun nanofiber yarns enabled 3D cellular alignment and differentiation, and the "M-type" structure led to cellular organization and interaction at the interface zone. Crucially, a compartmentalized co-culture system is integrated into an MTJ-on-a-chip device for the precise co-culture of muscle and tendon zones using their medium at the same time. Such an MTJ-on-a-chip device is further served for drug-associated MTJ toxic or protective efficacy investigations. These results highlight that these interwoven nanofibrous scaffolds with biomimetic "M-type" interface are beneficial for engineered MTJ tissue development, and MTJ-on-a-chip with precise co-culture system indicated their promising potential as in vitro musculoskeletal models for drug development and biological mechanism studies.

A modular hydrogel bioink containing microsphere-embedded chondrocytes for 3D-printed multiscale composite scaffolds for cartilage repair.

Articular cartilage tissue engineering is being considered an alternative treatment strategy for promoting cartilage damage repair. Herein, we proposed a modular hydrogel-based bioink containing microsphere-embedded chondrocytes for 3D printing multiscale scaffolds integrating the micro and macro environment of the native articular cartilage. Gelatin methacryloyl (GelMA)/alginate microsphere was prepared by a microfluidic approach, and the chondrocytes embedded in the microspheres remained viable after being frozen and resuscitated. The modular hydrogel bioink could be printed via the gel-in-gel 3D bioprinting strategy for fabricating the multiscale hydrogel-based scaffolds. Meanwhile, the cells cultured in the scaffolds showed good proliferation and differentiation. Furthermore, we also found that the composite hydrogel was biocompatible in vivo. These results indicated that the modular hydrogel-based bioinks containing microsphere-embedded chondrocytes for 3D printing multiscale scaffolds could provide a 3D multiscale environment for enhancing cartilage repairing, which would be encouraging considering the numerous alternative applications in articular cartilage tissue engineering.

Global trends and hotspots in research on organoids between 2011 and 2020: a bibliometric analysis.

BACKGROUND: There is a lack of effective platforms that can rapidly screen drugs, for patients to achieve precision treatment. Since an organoid simulates the tissue or organ structure and function in vitro, it can be applied to predict the response to therapy, personalized medicine, and drug screening in clinical practice. However, the rapid development of this field meets several challenges. This study aimed to evaluate the current state of the organoid and prioritize future research areas using bibliometric analysis. METHODS: We selected articles and reviewers from the Web of Science database, using the search strategy syntax including "organoid" or "organoids", for the years 2011 to 2020. We conducted a general analysis and a thematic evolution analysis using the bibliometrix R package. Networks connecting productive countries/regions/institutions/authors were generated using VOSviewer. We performed a co-occurrence analysis using VOSviewer, burstness analysis using Citespace, and co-word biclustering analysis and landform map using BICOMB and gCLUTO to identify possible current and future directions and hotspots. RESULTS: We selected 3,168 publications for our analysis. We found that the number of publications in this field has increased sharply. The greatest contributions to organoid research have been made by the United States (among countries) and the University of Michigan (among institutions), and Hans Clevers is the most influential author. The journals with the highest number of selected articles and citations are Cancer Research and Nature. We observed the possibility of keyword classification into five clusters. Their trend changed from "methods to build organoids" (e.g., "lgr5+ stem cell" and "3D culture") to "practical applications of organoids" (e.g., "cystic fibrosis" and "Zika virus"). CONCLUSIONS: Our study used bibliometric analysis to provide an in-depth understanding of the trends and hotspots of organoid research. The primarily important subject matters are drug screening, disease modeling, personalized medicine, regenerative medicine, and developmental biology. However, this field still faces limitations in the form of lack of reproducibility, low levels of maturity and function, and the absence of appropriate readouts. Therefore, these five significant topics, and possible solutions to limitations (involving bioengineering strategies), might be noteworthy in the future.

Injectable composite hydrogels encapsulating gelatin methacryloyl/chitosan microspheres as ARPE-19 cell transplantation carriers.

Retinal pigment epithelial (RPE) cell transplantation is being explored as a feasible approach for treating age-related macular degeneration. The low aggregation ability of RPE cell suspensions or microtissues after transplantation has limited cell utilisation. Therefore, alternative transplantation strategies should be explored to induce cell aggregation and maintain cell viability. Herein, we propose a composite hydrogel that encapsulates gelatin methacryloyl (GelMA)/chitosan microspheres (GCMSs) as ARPE-19 cell transplantation carriers. The diameter of the GCMS was adjusted by tuning the parameters of the microfluidic devices, yielding a cell-adhering platform that induced uniform cell spreading. The live/dead assay and immunofluorescence results showed that ARPE-19 cells adhered and spread uniformly around the microspheres. Moreover, the hydrogel sheets were used to provide an aggregated protective shell, and the ARPE-19 cells on the microspheres encapsulated within these hydrogel sheets remained viable post-injection and produced fewer reactive oxygen species after cyclic stretching. Furthermore, we found that the composite hydrogel was biodegradable and biocompatible in vivo. Therefore, GCMSs provide an injectable microcarrier for ARPE-19 cells, and the hydrogel provides an aggregated protective shell in this novel platform, which has considerable potential for an alternative injectable and highly aggregated RPE cell transplantation strategy design.

Injectable microfluidic hydrogel microspheres based on chitosan and poly(ethylene glycol) diacrylate (PEGDA) as chondrocyte carriers.

Direct injection of chondrocytes in a minimally invasive way has been regarded as the significantly potential treatment for cartilage repair due to their ability to fill various irregular chondral defects. However, the low cell retention and survival after injection still limited their application in clinical transformation. Herein, we present chondrocyte-laden microspheres as cell carriers based on a double-network hydrogel by the combination of the chitosan and poly(ethylene glycol) diacrylate (PEGDA). The microfluidic technique was applied to prepare size-controllable chitosan/PEGDA hydrogel microspheres (CP-MSs) via the water-in-oil approach after photo-crosslinking and physical-crosslinking. The chondrocytes were laden on CP-MSs, which showed good cell viability and proliferation after long-term cell cultivation. The in vitro investigation further demonstrated that chondrocyte-laden CP-MSs were injectable and the cell viability was still high after injection. In particular, these cell-laden microspheres were self-assembled into a 3D cartilage-like scaffold by a bottom-up strategy based on cell-cell interconnectivity, which suggested that these injectable chondrocyte-laden microspheres showed potential applications as chondrocyte carriers for bottom-to-up cartilage tissue engineering.