Capacity For Self-repair Research Articles

Ursodesoxycholic acid alleviates liver fibrosis via proregeneration by activation of the ID1-WNT2/HGF signaling pathway.

BackgroundThe human liver possesses a remarkable capacity for self‐repair. However, liver fibrosis remains a serious medical concern, potentially progressing to end‐stage liver cirrhosis and even death. Liver fibrosis is characterized by excess accumulation of extracellular matrix in response to chronic injury. Liver regenerative ability, a strong indicator of liver health, is important in resisting fibrosis. In this study, we provide evidence that ursodesoxycholic acid (UDCA) can alleviate liver fibrosis by promoting liver regeneration via activation of the ID1‐WNT2/hepatocyte growth factor (HGF) pathway.MethodsBile duct ligation (BDL) and partial hepatectomy (PH) mouse models were used to verify the effects of UDCA on liver fibrosis, regeneration, and the ID1‐WNT2/HGF pathway. An Id1 knockdown mouse model was also used to assess the role of Id1 in UDCA alleviation of liver fibrosis.ResultsOur results demonstrate that UDCA can alleviate liver fibrosis in the BDL mice and promote liver regeneration via the ID1‐WNT2/HGF pathway in PH mice. In addition, Id1 knockdown abolished the protection afforded by UDCA in BDL mice.ConclusionsWe conclude that UDCA protects against liver fibrosis by proregeneration via activation of the ID1‐WNT2/HGF pathway.

Changes in proteins related to early nerve repair in a rat model of sciatic nerve injury

Peripheral nerves have a limited capacity for self-repair and those that are severely damaged or have significant defects are challenging to repair. Investigating the pathophysiology of peripheral nerve repair is important for the clinical treatment of peripheral nerve repair and regeneration. In this study, rat models of right sciatic nerve injury were established by a clamping method. Protein chip assay was performed to quantify the levels of neurotrophic, inflammation-related, chemotaxis-related and cell generation-related factors in the sciatic nerve within 7 days after injury. The results revealed that the expression levels of neurotrophic factors (ciliary neurotrophic factor) and inflammation-related factors (intercellular cell adhesion molecule-1, interferon γ, interleukin-1α, interleukin-2, interleukin-4, interleukin-6, monocyte chemoattractant protein-1, prolactin R, receptor of advanced glycation end products and tumor necrosis factor-α), chemotaxis-related factors (cytokine-induced neutrophil chemoattractant-1, L-selectin and platelet-derived growth factor-AA) and cell generation-related factors (granulocyte-macrophage colony-stimulating factor) followed different trajectories. These findings will help clarify the pathophysiology of sciatic nerve injury repair and develop clinical treatments of peripheral nerve injury. This study was approved by the Ethics Committee of Peking University People's Hospital of China (approval No. 2015-50) on December 9, 2015.

Endogenous Repair and Regeneration of Injured Articular Cartilage: A Challenging but Promising Therapeutic Strategy

Articular cartilage (AC) has a very limited intrinsic repair capacity after injury or disease. Although exogenous cell-based regenerative approaches have obtained acceptable outcomes, they are usually associated with complicated procedures, donor-site morbidities and cell differentiation during ex vivo expansion. In recent years, endogenous regenerative strategy by recruiting resident mesenchymal stem/progenitor cells (MSPCs) into the injured sites, as a promising alternative, has gained considerable attention. It takes full advantage of body’s own regenerative potential to repair and regenerate injured tissue while avoiding exogenous regenerative approach-associated limitations. Like most tissues, there are also multiple stem-cell niches in AC and its surrounding tissues. These MSPCs have the potential to migrate into injured sites to produce replacement cells under appropriate stimuli. Traditional microfracture procedure employs the concept of MSPCs recruitment usually fails to regenerate normal hyaline cartilage. The reasons for this failure might be attributed to an inadequate number of recruiting cells and adverse local tissue microenvironment after cartilage injury. A strategy that effectively improves local matrix microenvironment and recruits resident MSPCs may enhance the success of endogenous AC regeneration (EACR). In this review, we focused on the reasons why AC cannot regenerate itself in spite of potential self-repair capacity and summarized the latest developments of the three key components in the field of EACR. In addition, we discussed the challenges facing in the present EACR strategy. This review will provide an increasing understanding of EACR and attract more researchers to participate in this promising research arena.

Tissue Engineering of Cartilage Using a Random Positioning Machine.

Articular cartilage is a skeletal tissue of avascular nature and limited self-repair capacity. Cartilage-degenerative diseases, such as osteoarthritis (OA), are difficult to treat and often necessitate joint replacement surgery. Cartilage is a tough but flexible material and relatively easy to damage. It is, therefore, of high interest to develop methods allowing chondrocytes to recolonize, to rebuild the cartilage and to restore joint functionality. Here we studied the in vitro production of cartilage-like tissue using human articular chondrocytes exposed to the Random Positioning Machine (RPM), a device to simulate certain aspects of microgravity on Earth. To screen early adoption reactions of chondrocytes exposed to the RPM, we performed quantitative real-time PCR analyses after 24 h on chondrocytes cultured in DMEM/F-12. A significant up-regulation in the gene expression of IL6, RUNX2, RUNX3, SPP1, SOX6, SOX9, and MMP13 was detected, while the levels of IL8, ACAN, PRG4, ITGB1, TGFB1, COL1A1, COL2A1, COL10A1, SOD3, SOX5, MMP1, and MMP2 mRNAs remained unchanged. The STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) analysis demonstrated among others the importance of these differentially regulated genes for cartilage formation. Chondrocytes grown in DMEM/F-12 medium produced three-dimensional (3D) spheroids after five days without the addition of scaffolds. On day 28, the produced tissue constructs reached up to 2 mm in diameter. Using specific chondrocyte growth medium, similar results were achieved within 14 days. Spheroids from both types of culture media showed the typical cartilage morphology with aggrecan positivity. Intermediate filaments form clusters under RPM conditions as detected by vimentin staining after 7 d and 14 d. Larger meshes appear in the network in 28-day samples. Furthermore, they were able to form a confluent chondrocyte monolayer after being transferred back into cell culture flasks in 1 g conditions showing their suitability for transplantation into joints. Our results demonstrate that the cultivation medium has a direct influence on the velocity of tissue formation and tissue composition. The spheroids show properties that make them interesting candidates for cellular cartilage regeneration approaches in trauma and OA therapy.

Primary Cilia Direct Murine Articular Cartilage Tidemark Patterning Through Hedgehog Signaling and Ambulatory Load.

Articular cartilage (AC) is essential for body movement but is highly susceptible to degenerative diseases and has poor self-repair capacity. To improve current subpar regenerative treatments, developmental mechanisms of AC should be clarified and, specifically, how its postnatal multizone organization is acquired. Primary cilia are cell surface organelles crucial for mammalian tissue morphogenesis. Although their importance for chondrocyte function is appreciated, their specific roles in postnatal AC morphogenesis remain unclear. To explore these mechanisms, we used a murine conditional loss-of-function approach (Ift88-flox) targeting joint-lineage progenitors (Gdf5Cre) and monitored postnatal knee AC development. Joint formation and growth up to juvenile stages were largely unaffected. However, mature AC (aged 2 months) exhibited disorganized extracellular matrix, decreased aggrecan and collagen II due to reduced gene expression (not increased catabolism), and marked reduction of AC modulus by 30%-50%. In addition, and unexpectedly, we discovered that tidemark patterning was severely disrupted, as was hedgehog signaling, and exhibited specificity based on regional load-bearing functions of AC. Interestingly, Prg4 expression was markedly increased in highly loaded sites in mutants. Together, our data provide evidence that primary cilia orchestrate postnatal AC morphogenesis including tidemark topography, zonal matrix composition, and ambulation load responses. © 2022 American Society for Bone and Mineral Research (ASBMR).

Applying a Three-dimensional Uniaxial Mechanical Stimulation Bioreactor System to Induce Tenogenic Differentiation of Tendon-Derived Stem Cells.

Tendinopathy is a common chronic tendon disease relating to inflammation and degeneration in an orthopaedic area. With high morbidity, limited self-repairing capacity and, most importantly, no definitive treatments, tendinopathy still influences patients' life quality negatively. Tendon-derived stem cells (TDSCs), as primary precursor cells of tendon cells, play an essential role in both the development of tendinopathy, and functional and structural restoration after tendinopathy. Thus, a method that can in vitro mimic the in vivo differentiation of TDSCs into tendon cells would be useful. Here, the present protocol describes a method based on a three-dimensional (3D) uniaxial stretching system to stimulate the TDSCs to differentiate into tendon-like tissues. There are seven stages of the present protocol: isolation of mice TDSCs, culture and expansion of mice TDSCs, preparation of stimulation culture medium for cell sheet formation, cell sheet formation by culturing in stimulation medium, preparation of 3D tendon stem cell construct, assembly of the uniaxial-stretching mechanical stimulation complex, and evaluation of the mechanical stimulated in vitro tendon-like tissue. The effectiveness was demonstrated by histology. The entire procedure takes less than 3 weeks. To promote extracellular matrix deposition, 4.4 mg/mL ascorbic acid was used in the stimulation culture medium. A separated chamber with a linear motor provides accurate mechanical loading and is portable and easily adjusted, which is applied for the bioreactor. The loading regime in the present protocol was 6% strain, 0.25 Hz, 8 h, followed by 16 h rest for 6 days. This protocol could mimic cell differentiation in the tendon, which is helpful for the investigation of the pathological process of tendinopathy. Moreover, the tendon-like tissue is potentially used to promote tendon healing in tendon injury as an engineered autologous graft. To sum up, the present protocol is simple, economic, reproducible and valid.

Discovery of Selenocysteine as aPotential Nanomedicine Promotes Cartilage Regeneration With Enhanced Immune Response by Text Mining and Biomedical Databases.

BackgroundUnlike bone tissue, little progress has been made regarding cartilage regeneration, and many challenges remain. Furthermore, the key roles of cartilage lesion caused by traumas, focal lesion, or articular overstress remain unclear. Traumatic injuries to the meniscus as well as its degeneration are important risk factors for long-term joint dysfunction, degenerative joint lesions, and knee osteoarthritis (OA) a chronic joint disease characterized by degeneration of articular cartilage and hyperosteogeny. Nearly 50% of the individuals with meniscus injuries develop OA over time. Due to the limited inherent self-repair capacity of cartilage lesion, the Biomaterial drug-nanomedicine is considered to be a promising alternative. Therefore, it is important to elucidate the gene potential regeneration mechanisms and discover novel precise medication, which are identified through this study to investigate their function and role in pathogenesis.MethodsWe downloaded the mRNA microarray statistics GSE117999, involving paired cartilage lesion tissue samples from 12 OA patients and 12 patients from a control group. First, we analyzed these statistics to recognize the differentially expressed genes (DEGs). We then exposed the gene ontology (GO) annotation and the Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway enrichment analyses for these DEGs. Protein-protein interaction (PPI) networks were then constructed, from which we attained eight significant genes after a functional interaction analysis. Finally, we identified a potential nanomedicine attained from this assay set, using a wide range of inhibitor information archived in the Search Tool for the Retrieval of Interacting Genes (STRING) database.ResultsSixty-six DEGs were identified with our standards for meaning (adjusted P-value < 0.01, |log2 - FC| ≥1.2). Furthermore, we identified eight hub genes and one potential nanomedicine - Selenocysteine based on these integrative data.ConclusionWe identified eight hub genes that could work as prospective biomarkers for the diagnostic and biomaterial drug treatment of cartilage lesion, involving the novel genes CAMP, DEFA3, TOLLIP, HLA-DQA2, SLC38A6, SLC3A1, FAM20A, and ANO8. Meanwhile, these genes were mainly associated with immune response, immune mediator induction, and cell chemotaxis. Significant support is provided for obtaining a series of novel gene targets, and we identify potential mechanisms for cartilage regeneration and final nanomedicine immunotherapy in regenerative medicine.

Advances in biomaterials for skeletal muscle engineering and obstacles still to overcome.

Repair of injured skeletal muscle is a sophisticated process that uses immune, muscle, perivascular, and neural cells. In acute injury, the robust endogenous repair process can facilitate complete regeneration with little to no functional deficit. However, in severe injury, the damage is beyond the capacity for self-repair, often resulting in structural and functional deficits. Aside from the insufficiencies in muscle function, the aesthetic deficits can impact quality of life. Current clinical treatments are significantly limited in their capacity to structurally and functionally repair the damaged skeletal muscle. Therefore, alternative approaches are needed. Biomaterial therapies for skeletal muscle engineering have leveraged natural materials with sophisticated scaffold fabrication techniques to guide cell infiltration, alignment, and differentiation. Advances in biomaterials paired with a standardized and rigorous assessment of resulting tissue formation have greatly advanced the field of skeletal muscle engineering in the last several years. Herein, we discuss the current trends in biomaterials-based therapies for skeletal muscle regeneration and present the obstacles still to be overcome before clinical translation is possible. With millions of people affected by muscle trauma each year, the development of a therapy that can repair the structural and functional deficits after severe muscle injury is pivotal.

Molecular Insights Into Lysyl Oxidases in Cartilage Regeneration and Rejuvenation.

Articular cartilage remains among the most difficult tissues to regenerate due to its poor self-repair capacity. The lysyl oxidase family (LOX; also termed as protein-lysine 6-oxidase), mainly consists of lysyl oxidase (LO) and lysyl oxidase-like 1-4 (LOXL1-LOXL4), has been traditionally defined as cuproenzymes that are essential for stabilization of extracellular matrix, particularly cross-linking of collagen and elastin. LOX is essential in the musculoskeletal system, particularly cartilage. LOXs-mediated collagen cross-links are essential for the functional integrity of articular cartilage. Appropriate modulation of the expression or activity of certain LOX members selectively may become potential promising strategy for cartilage repair. In the current review, we summarized the advances of LOX in cartilage homeostasis and functioning, as well as copper-mediated activation of LOX through hypoxia-responsive signaling axis during recent decades. Also, the molecular signaling network governing LOX expression has been summarized, indicating that appropriate modulation of hypoxia-responsive-signaling-directed LOX expression through manipulation of bioavailability of copper and oxygen is promising for further clinical implications of cartilage regeneration, which has emerged as a potential therapeutic approach for cartilage rejuvenation in tissue engineering and regenerative medicine. Therefore, targeted regulation of copper-mediated hypoxia-responsive signalling axis for selective modulation of LOX expression may become potential effective therapeutics for enhanced cartilage regeneration and rejuvenation in future clinical implications.

Tissue Engineered Materials in Cardiovascular Surgery: The Surgeon's Perspective

In cardiovascular surgery, reconstruction and replacement of cardiac and vascular structures are routinely performed. Prosthetic or biological materials traditionally used for this purpose cannot be considered ideal substitutes as they have limited durability and no growth or regeneration potential. Tissue engineering aims to create materials having normal tissue function including capacity for growth and self-repair. These advanced materials can potentially overcome the shortcomings of conventionally used materials, and, if successfully passing all phases of product development, they might provide a better option for both the pediatric and adult patient population requiring cardiovascular interventions. This short review article overviews the most important cardiovascular pathologies where tissue engineered materials could be used, briefly summarizes the main directions of development of these materials, and discusses the hurdles in their clinical translation. At its beginnings in the 1980s, tissue engineering (TE) was defined as “an interdisciplinary field that applies the principles of engineering and the life sciences toward the development of biological substitutes that restore, maintain, or improve tissue function” (1). Currently, the utility of TE products and materials are being investigated in several fields of human medicine, ranging from orthopedics to cardiovascular surgery (2–5). In cardiovascular surgery, reconstruction and replacement of cardiac and vascular structures are routinely performed. Considering the shortcomings of traditionally used materials, the need for advanced materials that can “restore, maintain or improve tissue function” are evident. Tissue engineered substitutes, having growth and regenerative capacity, could fundamentally change the specialty (6). This article overviews the most important cardiovascular pathologies where TE materials could be used, briefly summarizes the main directions of development of TE materials along with their advantages and shortcomings, and discusses the hurdles in their clinical translation.

A moldable thermosensitive hydroxypropyl chitin hydrogel for 3D cartilage regeneration in vitro and in vivo

Because of poor self-repair capacity, the repair of cartilage defect is always a great challenge in clinical treatment. In vitro cartilage regeneration provides a potential strategy for functional reconstruction of cartilage defect. Hydrogel has been known as an ideal cartilage regeneration scaffold. However, to date, in vitro cartilage regeneration based on hydrogel has not achieved satisfactory results. The current study explored the feasibility of in vitro 3D cartilage regeneration based on a moldable thermosensitive hydroxypropyl chitin (HPCH) hydrogel and its in vivo fate. The thermosensitive HPCH hydrogel was prepared and characterized. Goat auricular chondrocytes were encapsulated into the HPCH hydrogel to form a chondrocyte-hydrogel construct. The constructs were injected subcutaneously into nude mice or molded into different shapes for in vitro chondrogenic culture followed by in vivo implantation. The results demonstrated that the HPCH hydrogel possessed satisfactory gelation properties (gelation time < 18 s at 37 °C), biocompatibility (cell amount almost doubled within one week), and the ability to be applied as an injectable hydrogel for cartilage regeneration. All the constructs of in vitro culture basically maintained their original shapes (in vitro to initial: 110.8%) and displayed typical cartilaginous features with abundant lacunae and cartilage specific matrix deposition. These in vitro samples became more mature with prolonged in vivo implantation and largely maintained the original shape (in vivo to in vitro: 103.5%). These results suggested that the moldable thermosensitive HPCH hydrogel can serve as a promising scaffold for cartilage regeneration with defined shapes in vitro and in vivo. Statement of significanceBecause of avascular and non-nervous characteristic of cartilage, in vitro regeneration plays an important role in reconstructing cartilage function. Hydrogel has been known as an ideal cartilage regeneration scaffold. However, to date, in vitro cartilage regeneration based on hydrogel has not achieved satisfactory results. The current study demonstrated that the chondrocyte-hydrogel construct generated by high density of chondrocytes encapsulated into a thermosensitive HPCH hydrogel could successfully regenerate in vitro typical cartilage-like tissue with defined shapes and further mature to form homogeneous cartilage with their original shapes after in vivo implantation. The current study indicated that the moldable thermosensitive HPCH hydrogel could serve as a promising scaffold for in vitro and in vivo cartilage regeneration with different shapes.

Effects of oxygen concentration on differentiation of human IPS cells into cartilage tissue

Purpose: Cartilage is a type of tissue characterized by poor self-repair capacity due to the absence of blood vessels and nerve tissue. As such, cartilaginous tissue does not self-repair after sustaining extensive damage. Treatments to address cartilaginous tissue damage include bone perforation, osteochondral column transplantation, and autologous cultured cartilage transplantation. Each of these treatment methods has achieved a degree of success, but problems related to the number of procedures required and the quality of regenerated tissue remain to be overcome. In recent years, regenerative medical techniques have come to be regarded as promising new therapy options. Among these, induced pluripotent stem cells (iPSCs) have attracted attention as a potential new source of cells for therapeutic uses. iPSCs exhibit high self-renewal capacity and have excellent potential as a cell source owing to their maintenance of the ability to undergo cell division while remaining undifferentiated. Although the formation of cartilage tissues from iPSCs has been established, regenerative medicines utilizing iPSCs also face obstacles, such as long culture periods, the risk of oncogenesis, and compromised tissue purity. We hypothesized that if cartilaginous tissues could be prepared from iPSCs in a hypoxic environment, it would be possible to produce tissues more quickly than under a stable oxygen environment. In this study, the effects of oxygen concentration on the differentiation of human iPSC cells (hiPSCs) into cartilage were studied. Methods: The established hiPSC line Toe was maintained in feeder-free medium in 6-well dishes coated with laminin. The hiPSCs formed high-density cell colonies consisting of 1-2 × 105 cells at 10 days after the start of maintenance. Subsequently, chondrogenic differentiation of the iPSCs was induced. The hiPSCs were initially differentiated into mesendodermal cells in mesendodermal medium for 3 days (M-phase). On day 3, the medium was changed to chondrogenic medium, hiPSCs were cultured in chondrogenic medium for 10 days (C-phase) to induce differentiation into chondrocytes. Then, four groups were formed based on oxygen concentration in the M-phase and C-phase: NN (21%, 21%), HN (5%, 21%), NH (21%, 5%), and HH (5%, 5%). On day 14, total RNA was extracted from differentiated cells. The expression of sox9, aggrecan, and col2a1, which are cartilage differentiation markers, and of T and FOXF1, which are undifferentiated mesoderm markers, was assessed using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Moreover, cells were detached and digested to form a single-cell suspension using trypsin for flowcytometry, and the sox9-positive cell rate was measured. Results: In qRT-PCR analysis, sox9 expression was higher in NH than in NN and HN groups. col2a1 expression was higher in NH than in the other groups. Aggrecan expression was higher in NH group compared with HN group. T and FOXF1 expression was higher in HN group than NH group, and the groups cultured in hypoxic condition on C-phase (NH and HH groups) were lower than the groups cultured in normoxic condition on C-phase (NN and HN groups). In flowcytometry analysis, sox9-positive cell rate was higher in the NH and HH groups (5% C-phase oxygen concentration) than in the NN and HN groups (21% C-phase oxygen concentration). Conclusions: Chondrocyte differentiation was promoted by culture under hypoxic conditions during the production of cartilaginous tissues from human embryonic stem (ES) cells. Thus, it is possible that a hypoxic environment may also promote differentiation of cartilaginous tissue from iPSCs. In this study, a period of culturing in a hypoxic environment was provided during differentiation from hiPSCs into cartilage tissue. C-phase culturing with the oxygen concentration of 5% raised cartilage differentiation markers and lowered undifferentiated mesoderm markers. An oxygen concentration of 5% promotes differentiation from hiPSCs into cartilage tissue and raises purity levels. Therefore, adjusting the culture period or oxygen concentration (hypoxic environment) may allow engineering of high-purity cartilage tissues in a short time.

3D-printable photocurable bioink for cartilage regeneration of tonsil-derived mesenchymal stem cells

Cartilage regeneration is challenging because of the poor intrinsic self-repair capacity of avascular tissue. Three-dimensional (3D) bioprinting has gained significant attention in the field of tissue engineering and is a promising technology to overcome current difficulties in cartilage regeneration. Although bioink is an essential component of bioprinting technology, several challenges remain in satisfying different requirements for ideal bioink, including biocompatibility and printability based on specific biological requirements. Gelatin and hyaluronic acid (HA) have been shown to be ideal biomimetic hydrogel sources for cartilage regeneration. However, controlling their structure, mechanical properties, biocompatibility, and degradation rate for cartilage repair remains a challenge. Here, we show a photocurable bioink created by hybridization of gelatin methacryloyl (GelMA) and glycidyl-methacrylated HA (GMHA) for material extrusion 3D bioprinting in cartilage regeneration. GelMA and GMHA were mixed in various ratios, and the mixture of 7% GelMA and 5% GMHA bioink (G7H5) demonstrated the most reliable mechanical properties, rheological properties, and printability. This G7H5 bioink allowed us to build a highly complex larynx structure, including the hyoid bone, thyroid cartilage, cricoid cartilage, arytenoid cartilage, and cervical trachea. This bioink also provided an excellent microenvironment for chondrogenesis of tonsil-derived mesenchymal stem cells (TMSCs) in vitro and in vivo. In summary, this study presents the ideal formulation of GelMA/GMHA hybrid bioink to generate a well-suited photocurable bioink for cartilage regeneration of TMSCs using a material extrusion bioprinter, and could be applied to cartilage tissue engineering.

Neuro-protection and neuro-regeneration of the optic nerve: recent advances and future directions.

Optic neuropathies refer to a collection of diseases in which retinal ganglion cells (RGCs), the specialized neuron of the retina whose axons make up the optic nerve, are selectively damaged. Blindness secondary to optic neuropathies is irreversible as RGCs do not have the capacity for self-renewal and have a limited capacity for self-repair. Numerous strategies are being developed to either prevent further RGC degeneration or replace the cells that have degenerated. In this review, we aim to discuss known limitations to regeneration in central nervous system (CNS), followed by a discussion of previous, current, and future strategies for optic nerve neuroprotection as well as approaches for neuro-regeneration, with an emphasis on developments in the past two years. Neuro-regeneration in the CNS is limited by both intrinsic and extrinsic factors. Environmental barriers to axon regeneration can be divided into two major categories: failure to clear myelin and formation of glial scar. Although inflammatory scars block axon growth past the site of injury, inflammation also provides important signals that activate reparative and regenerative pathways in RGCs. Neuroprotection with neurotrophins as monotherapy is not effective at preventing RGC degeneration likely secondary to rapid clearance of growth factors. Novel approaches involve exploiting different technologies to provide sustained delivery of neurotrophins. Other approaches include application of anti-apoptosis molecules and anti-axon retraction molecules. Although stem cells are becoming a viable option for generating RGCs for cell-replacement-based strategies, there are still many critical barriers to overcome before they can be used in clinical practice. Adjuvant treatments, such as application of electrical fields, scaffolds, and magnetic field stimulation, may be useful in helping transplanted RGCs extend axons in the proper orientation and assist with new synapse formation. Different optic neuropathies will benefit from neuro-protective versus neuro-regenerative approaches. Developing clinically effective treatments for optic nerve disease will require a collaborative approach that not only employs neurotrophic factors but also incorporates signals that promote axonogenesis, direct axon growth towards intended targets, and promote appropriate synaptogenesis.

Role of the Calcified Cartilage Layer of an Integrated Trilayered Silk Fibroin Scaffold Used to Regenerate Osteochondral Defects in Rabbit Knees.

The repair of osteochondral defects remains challenging, given the complexity of native osteochondral tissue and the limited self-repair capacity of cartilage. Osteochondral tissue engineering is a promising strategy. Here, we fabricated a biomimetic osteochondral scaffold using silk fibroin and hydroxyapatite, including a calcified cartilage layer (CCL). We studied the role played by the CCL in terms of cell viability in vivo. We established osteochondral defects in rabbit knees to investigate the effects of CCL-containing scaffolds with or without adipose tissue-derived stem cells (ADSCs). We evaluated osteochondral tissue regeneration by calculating gross observational scores, via histological and immunohistochemical assessments, by performing quantitative biochemical and biomechanical analyses of new osteochondral tissue, and via microcomputed tomography of new bone at 4, 8, and 12 weeks after surgery. In terms of surface roughness and integrity, the CCL + ADSCs group was better than the CCL and the non-CCL + ADSCs groups at all time points tested; the glycosaminoglycan and collagen type II levels of the CCL + ADSCs group were highest, reflecting the important role played by the CCL in cartilage tissue repair. Subchondral bone smoothness was better in the CCL + ADSCs group than in the non-CCL + ADSCs and CCL groups. The CCL promoted smooth subchondral bone regeneration but did not obviously affect bone strength or quality. In conclusion, a biomimetic osteochondral scaffold with a CCL, combined with autologous ADSCs, satisfactorily regenerated a rabbit osteochondral defect. The CCL enhances cartilage and subchondral bone regeneration.

THE EFFECT OF DECELLULARIZATION TECHNIQUE ON COLLAGEN TYPE II AND MATRICES POROSITY OF CARTILAGE BOVINE SCAFFOLD

Background: Cartilage has limited capacity for self-repair in case of defect. Stem cell based tissue engineering has become an alternative therapy for the treatment of cartilage defect. The use of decellularized cartilage bovine can serve as scaffold to support proliferation dan differentiation of the stem cell.Purpose: This research is to compare the effect of decellularization technique on collagen type II and matrices porosity of cartilage bovine scaffold.Methods: This experimental laboratory research used cartilage bovine scaffold which was decellularized using physical, chemical and enzymatic techniques with different time and concentration variations, then collagen type II measurement was performed by immunohistochemical method and evaluation of porosity size with electron microscopy.Results: In the preparations with the highest concentration and the longest immersion time of 5% SDS for 72 hours, the decellularization process occurs perfectly. The collagen type II content was found to have significantly higher concentrations than the controls and other preparations, and have interconnected porosities in large size.Conclusion: High concentrations of collagen type II and large diameter of pores after decellularization process in cartilage bovine scaffold confirm that this scaffold still has optimum biochemical cues that can be a good mechanical framework and microenvironment stem cell in effort to repair the cartilage defects. This result was expected to become a standard for the decellularization making of cartilage bovine scaffold.

Pollutant removal performance and microbial enhancement mechanism by water-lifting and aeration technology in a drinking water reservoir ecosystem

Here, the performance and mechanism of pollutant removal in the Zhoucun reservoir by water-lifting and aeration systems (WLAs) were explored. The hypolimnion anoxic layer disappeared and the reservoir was mixed after the WLAs were operated for approximately 35 days, providing a suitable environment for pollutant removal. Operation of the system enhanced the metabolic activity of the water microbes and their capacity for purification, which contributed to the removal of nitrogen, organic carbon, Fe, Mn, P, and S. Specifically, the total N concentration decreased from 2.55 to 0.48 mg/L, showing an 81.18% removal rate. Microbial metabolism and the diversity index increased following the operation of the WLAs in the Zhoucun Reservoir. Furthermore, the water reservoir clearly inhibited the performance of Fe, Mn, P, and S through the WLA operation, meeting the requirements for class III based on the Chinese Surface Water Environmental Quality Standard (GB3838–2002). High-throughput sequencing analysis revealed increased levels of indicator and keystone operational taxonomic units belonging to Flavobacterium, hgcI_clade, Rheinheimera, Dechloromonas, Pseudomonas, and Rhodocyclaceae, which are related to the degradation of organic carbon and removal of nitrogen and phosphorus. Moreover, total N, ammonia, total P, dissolved oxygen, temperature, and pH were the principal factors affecting the microbial community based on redundancy analysis and the Mantel test. Furthermore, network analysis showed that symbiotic relationships accounted for the major proportion of the microbial network. Our results provide a theoretical foundation for the efficiency of N removal and essential technical support for improving the self-repair capacity of water in drinking water reservoirs.

Advances in cartilage repair: The influence of inorganic clays to improve mechanical and healing properties of antibacterial Gellan gum-Manuka honey hydrogels.

Effective treatment of cartilage defects represents a challenging problem, mainly due to the tissue's limited intrinsic self-repair capacity; the use of polymeric scaffolds as tissue substitute is rapidly increasing, but it is still limited by poor mechanical properties. Moreover, the onset of an infection can irreversibly affect the healing process. Accordingly, in this work we describe, for the first time, the preparation of composite scaffolds based on gellan gum, antibacterial Manuka honey and an inorganic clay (mesoporous silica, sodium‑calcium bentonite or halloysite nanotubes). The surface composition, morphology, mechanical and biological features of such composites are herein assessed, aiming to optimize the composition of a superior scaffold for cartilage repair. Results demonstrated that after 45days of in vitro incubation with human mesenchymal stem cells, the mesoporous silica-composite hydrogels exhibited significant changes in peak elastic and dynamic moduli over time thus demonstrating superior mechanical properties. Moreover, mesoporous silica provided the best performances in terms of in vitro cytocompatibility and antibacterial preventive activity in protection of cells in a co-culture model. Therefore, this selected composition was exploited for subcutaneous implantation in mice to investigate materials biocompatibility and infection prevention. Results demonstrated that composites did not cause severe immune response as well as they were able to restrain the infection. Accordingly, GG-MH-MS composites represent a very promising tool for cartilage tissue engineering.

Reciprocal Regulation of TRPS1 and miR-221 in Intervertebral Disc Cells.

Intervertebral disc (IVD), a moderately moving joint located between the vertebrae, has a limited capacity for self-repair, and treating injured intervertebral discs remains a major challenge. The development of innovative therapies to reverse IVD degeneration relies primarily on the discovery of key molecules that, occupying critical points of regulatory mechanisms, can be proposed as potential intradiscal injectable biological agents. This study aimed to elucidate the underlying mechanism of the reciprocal regulation of two genes differently involved in IVD homeostasis, the miR-221 microRNA and the TRPS1 transcription factor. Human lumbar IVD tissue samples and IVD primary cells were used to specifically evaluate gene expression and perform functional analysis including the luciferase gene reporter assay, chromatin immunoprecipitation, cell transfection with hTRPS1 overexpression vector and antagomiR-221. A high-level expression of TRPS1 was significantly associated with a lower pathological stage, and TRPS1 overexpression strongly decreased miR-221 expression, while increasing the chondrogenic phenotype and markers of antioxidant defense and stemness. Additionally, TRPS1 was able to repress miR-221 expression by associating with its promoter and miR-221 negatively control TRPS1 expression by targeting the TRPS1-3′UTR gene. As a whole, these results suggest that, in IVD cells, a double-negative feedback loop between a potent chondrogenic differentiation suppressor (miR-221) and a regulator of axial skeleton development (TRPS1) exists. Our hypothesis is that the hostile degenerated IVD microenvironment may be counteracted by regenerative/reparative strategies aimed at maintaining or stimulating high levels of TRPS1 expression through inhibition of one of its negative regulators such as miR-221.

Tendon tissue microdamage and the limits of intrinsic repair

The transmission of mechanical muscle force to bone for musculoskeletal stability and movement is one of the most important functions of tendon. The load-bearing tendon core is composed of highly aligned collagen-rich fascicles interspersed with stromal cells (tenocytes). Despite being built to bear very high mechanical stresses, supra-physiological/repetitive mechanical overloading leads to tendon microdamage in fascicles, and potentially to tendon disease and rupture. To date, it is unclear to what extent intrinsic healing mechanisms of the tendon core compartment can repair microdamage. In the present study, we investigated the healing capacity of the tendon core compartment in an ex vivo tissue explant model. To do so, we isolated rat tail tendon fascicles, damaged them by applying a single stretch to various degrees of sub-rupture damage and longitudinally assessed downstream functional and structural changes over a period of several days. Functional damage was assessed by changes in the elastic modulus of the material stress-strain curves, and biological viability of the resident tenocytes. Structural damage was quantified using a fluorescent collagen hybridizing peptide (CHP) to label mechanically disrupted collagen structures. While we observed functional mechanical damage for strains above 2% of the initial fascicle length, structural collagen damage was only detectable for 6% strain and beyond. Minimally loaded/damaged fascicles (2–4% strain) progressively lost elastic modulus over the course of tissue culture, despite their collagen structures remaining intact with high degree of maintained cell viability. In contrast, more severely overloaded fascicles (6–8% strain) with damage at the molecular/collagen level showed no further loss of the elastic modulus but markedly decreased cell viability. Surprisingly, in these heavily damaged fascicles the elastic modulus partially recovered, an effect also seen in further experiments on devitalized fascicles, implying the possibility of a non-cellular but matrix-driven mechanism of molecular repair. Overall, our findings indicate that the tendon core has very little capacity for self-repair of microdamage. We conclude that stromal tenocytes likely do not play a major role in anabolic repair of tendon matrix microdamage, but rather mediate catabolic matrix breakdown and communication with extrinsic cells that are able to effect tissue repair.