Scar formation resulting from burns or severe trauma can significantly compromise the structural integrity of skin and lead to permanent loss of skin appendages,ultimately impairing its normal physiological function.A...Scar formation resulting from burns or severe trauma can significantly compromise the structural integrity of skin and lead to permanent loss of skin appendages,ultimately impairing its normal physiological function.Accumulating evidence underscores the potential of targeted modulation of mechanical cues to enhance skin regeneration,promoting scarless repair by influencing the extracellular microenvironment and driving the phenotypic transitions.The field of skin repair and skin appendage regeneration has witnessed remarkable advancements in the utilization of biomaterials with distinct physical properties.However,a comprehensive understanding of the underlying mechanisms remains somewhat elusive,limiting the broader application of these innovations.In this review,we present two promising biomaterial-based mechanical approaches aimed at bolstering the regenerative capacity of compromised skin.The first approach involves leveraging biomaterials with specific biophysical properties to create an optimal scarless environment that supports cellular activities essential for regeneration.The second approach centers on harnessing mechanical forces exerted by biomaterials to enhance cellular plasticity,facilitating efficient cellular reprogramming and,consequently,promoting the regeneration of skin appendages.In summary,the manipulation of mechanical cues using biomaterial-based strategies holds significant promise as a supplementary approach for achieving scarless wound healing,coupled with the restoration of multiple skin appendage functions.展开更多
Background: Motor neuron degeneration or loss in the spinal cord is the characteristic phenotype of motor neuron diseases or spinal cord injuries. Being proliferative and located near neurons, astrocytes are considere...Background: Motor neuron degeneration or loss in the spinal cord is the characteristic phenotype of motor neuron diseases or spinal cord injuries. Being proliferative and located near neurons, astrocytes are considered ideal cell sources for regenerating neurons.Methods: We selected and tested different combinations of the small molecules for inducing the conversion of human and mouse astrocytes into neurons. Microscopic imaging and immunocytochemistry analyses were used to characterize the morphology and phenotype of the induced neurons while RT-q PCR was utilized to analyze changes in gene expression. In addition, whole-cell patch-clamp recordings were measured to examine the electrophysiological properties of induced neurons.Results: The results showed that human astrocytes could be rapidly and efficiently converted into motor neuronlike cells by treatment with defined small molecules, with a yield of over 85% motor neuron-like cells attained. The induced motor neuron-like cells expressed the pan-neuronal markers TUJ1, MAP2, Neu N, and Synapsin 1 and motor neuron markers HB9, ISL1, CHAT, and VACh T. During the conversion process, the cells did not pass through a proliferative neural progenitor cell intermediate. The induced motor neurons were functional, showing the electrophysiological properties of neurons. The same chemical cocktail could induce spinal cord astrocytes from an amyotrophic lateral sclerosis mouse model carrying a SOD1 mutation to become motor neuron-like cells that exhibited a decrease in cell survival and an increase in oxidative stress compared to that observed in wild-type MNs derived from healthy mice. Moreover, the chemical induction reduced oxidative stress in the mutant astrocytes.Conclusions: The results of the present study demonstrated the feasibility of chemically converting human and mouse astrocytes into motor neuron-like cells that are useful for neurodegenerative disease modeling and regenerative medicine.展开更多
Background:Generation of neurons is essential in cell replacement therapy for neurodegenerative disorders like Parkinson’s disease.Several studies have reported the generation of dopaminergic(DA)neurons from mouse an...Background:Generation of neurons is essential in cell replacement therapy for neurodegenerative disorders like Parkinson’s disease.Several studies have reported the generation of dopaminergic(DA)neurons from mouse and human fibroblasts by ectopic expression of transcription factors,in which genetic manipulation is associated with potential risks.Methods:The small molecules and protein factors were selected based on their function to directly induce human fetal lung IMR-90 fibroblasts into DA neuron-like cells.Microscopical,immunocytochemical,and RT-qPCR analyses were used to characterize the morphology,phenotype,and gene expression features of the induced cells.The wholecell patch-clamp recordings were exploited to measure the electrophysiological properties.Results:Human IMR-90 fibroblasts were rapidly converted into DA neuron-like cells after the chemical induction using small molecules and protein factors,with a yield of approximately 95%positive TUJ1-positive cells.The induced DA neuron-like cells were immunopositive for pan-neuronal markers MAP2,NEUN,and Synapsin 1 and DA markers TH,DDC,DAT,and NURR1.The chemical induction process did not involve a neural progenitor/stem cell intermediate stage.The induced neurons could fire single action potentials,which reflected partially the electrophysiological properties of neurons.Conclusions:We developed a chemical cocktail of small molecules and protein factors to convert human fibroblasts into DA neuron-like cells without passing through a neural progenitor/stem cell intermediate stage.The induced DA neuron-like cells from human fibroblasts might provide a cellular source for cell-based therapy of Parkinson’s disease in the future.展开更多
To determine the prevalence and clinical features of olfactory and taste disorders among coronavirus disease 2019(COVID-19)patients in China.A cross-sectional study was performed in Wuhan from April 3,2020 to April 15...To determine the prevalence and clinical features of olfactory and taste disorders among coronavirus disease 2019(COVID-19)patients in China.A cross-sectional study was performed in Wuhan from April 3,2020 to April 15,2020.A total of 187 patients with confirmed severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)completed face-to-face interviews or telephone follow-ups.We found that the prevalence of olfactory and taste disorders was significantly lower in the Chinese cohort than in foreign COVID-19 cohorts.Females were more prone to olfactory and taste disorders.In some patients,olfactory and taste disorders precede other symptoms and can be used as early screening and warning signs.展开更多
Dear Editor,The large defects of the skin caused by burns or trauma can result in disruption of the structure and function of the skin,as well as irreversible loss of skin appendages,such as sweat glands(SG).As the im...Dear Editor,The large defects of the skin caused by burns or trauma can result in disruption of the structure and function of the skin,as well as irreversible loss of skin appendages,such as sweat glands(SG).As the important modulator of temperature homeostasis,SG damage triggers heat intolerance and thermoregulatory dysfunction,which poses a considerable threat to health and quality of life[1-3].The promotion of wound healing with SG restoration remains a challenging issue.It has been reported that there are urinary epithelial cells(UECs)in human urine samples,and they may most likely originate from renal proximal tubules[4].UECs are easy to obtain from urine samples at any age,sex,or ethnic origin except for renal failure,with a cost-effective,non-invasive and simple isolation method[4].More importantly,the extensive source of clinical urine sample makes it easy for urinary cell autogenous transplantation and individualized treatment.Therefore,it is of great significance to convert UECs into urinary epithelial stem cells(UESCs)with repair ability.In this context,we design a strategy for the pharmacological conversion of UECs into UESCs,and the repair ability of UESCs in functional skin wound healing is explored.展开更多
Background: Large skin defects severely disrupt the overall skin structure and can irreversibly damage sweat glands(SGs), thus impairing the skin’s physiological function. This study aims to develop a stepwise reprog...Background: Large skin defects severely disrupt the overall skin structure and can irreversibly damage sweat glands(SGs), thus impairing the skin’s physiological function. This study aims to develop a stepwise reprogramming strategy to convert fibroblasts into SG lineages, which may provide a promising method to obtain desirable cell types for the functional repair and regeneration of damaged skin.Methods: The expression of the SG markers cytokeratin 5(CK5), cytokeratin 10(CK10), cytokeratin 18(CK18), carcinoembryonic antigen(CEA), aquaporin 5(AQP5) and α-smooth muscle actin(α-SMA) was assessed with quantitative PCR(qPCR), immunofluorescence and flow cytometry. Calcium activity analysis was conducted to test the function of induced SG-like cells(iSGCs). Mouse xenograft models were also used to evaluate the in vivo regeneration of iSGCs.BALB/c nude mice were randomly divided into normal group, SGM treatment group and iSGC transplantation group.Immunocytochemical analyses and starch-iodine sweat tests were used to confirm the in vivo regeneration of iSGCs.Results: Ectodermal dysplasia antigen(EDA) overexpression drove human dermal fibroblast(HDF) conversion into i SGCs in SG culture medium(SGM). qPCR indicated significantly increased mRNA levels of the SG markers CK5, CK18and CEA in iSGCs, and flow cytometry data demonstrated(4.18±0.04)% of iSGCs were CK5 positive and(4.36±0.25)%of iSGCs were CK18 positive. The addition of chemical cocktails greatly accelerated the SG fate program. qPCR results revealed significantly increased mRNA expression of CK5, CK18 and CEA in iSGCs, as well as activation of the duct marker CK10 and luminal functional marker AQP5. Flow cytometry indicated, after the treatment of chemical cocktails,(23.05±2.49)% of iSGCs expressed CK5^(+) and(55.79±3.18)% of iSGCs expressed CK18^(+), respectively. Calcium activity analysis indicated that the reactivity of iSGCs to acetylcholine was close to that of primary SG cells [(60.79±7.71)% vs.(70.59±0.34)%, ns]. In vivo transplantation experiments showed approximately(5.2±1.1)% of the mice were sweat test positive, and the histological analysis results indicated that regenerated SG structures were present in iSGCs-treated mice.Conclusions: We developed a SG reprogramming strategy to generate functional iSGCs from HDFs by using the single factor EDA in combination with SGM and small molecules. The generation of iSGCs has important implications for future in situ skin regeneration with SG restoration.展开更多
Endogenous regeneration is becoming an increasingly important strategy for wound healing as it facilitates skin’s own regenerative potential for self-healing,thereby avoiding the risks of immune rejection and exogeno...Endogenous regeneration is becoming an increasingly important strategy for wound healing as it facilitates skin’s own regenerative potential for self-healing,thereby avoiding the risks of immune rejection and exogenous infection.However,currently applied biomaterials for inducing endogenous skin regeneration are simplistic in their structure and function,lacking the ability to accurately mimic the intricate tissue structure and regulate the disordered microenvironment.Novel biomimetic biomaterials with precise structure,chemical composition,and biophysical properties offer a promising avenue for achieving perfect endogenous skin regeneration.Here,we outline the recent advances in biomimetic materials induced endogenous skin regeneration from the aspects of structural and functional mimicry,physiological process regulation,and biophysical property design.Furthermore,novel techniques including in situ reprograming,flexible electronic skin,artificial intelligence,single-cell sequencing,and spatial transcriptomics,which have potential to contribute to the development of biomimetic biomaterials are highlighted.Finally,the prospects and challenges of further research and application of biomimetic biomaterials are discussed.This review provides reference to address the clinical problems of rapid and high-quality skin regeneration.展开更多
Electrospun nanofibrous membranes(eNFMs)have been extensively developed for bio-applications due to their structural and compositional similarity to the natural extracellular matrix.However,the emergence of antibiotic...Electrospun nanofibrous membranes(eNFMs)have been extensively developed for bio-applications due to their structural and compositional similarity to the natural extracellular matrix.However,the emergence of antibiotic resistance in bacterial infections significantly impedes the further development and applications of eNFMs.The development of antibacterial nanomaterials substantially nourishes the engineering design of antibacterial eNFMs for combating bacterial infections without relying on antibiotics.Herein,a comprehensive review of diverse fabrication techniques for incorporating antibacterial nanomaterials into eNFMs is presented,encompassing an exhaustive introduction to various nanomaterials and their bactericidal mechanisms.Furthermore,the latest achievements and breakthroughs in the application of these antibacterial eNFMs in tissue regenerative therapy,mainly focusing on skin,bone,periodontal and tendon tissues regeneration and repair,are systematically summarized and discussed.In particular,for the treatment of skin infection wounds,we highlight the antibiotic-free antibacterial therapy strategies of antibacterial eNFMs,including(i)single model therapies such as metal ion therapy,chemodynamic therapy,photothermal therapy,and photodynamic therapy;and(ii)multimodel therapies involving arbitrary combinations of these single models.Additionally,the limitations,challenges and future opportunities of antibacterial eNFMs in biomedical applications are also discussed.We anticipate that this comprehensive review will provide novel insights for the design and utilization of antibacterial eNFMs in future research.展开更多
Persistent inflammatory responses often occur when bacteria and other microorganisms frequently invade and colonize open wounds and eventually result in the formation of chronic wounds.Therefore,achieving real-time de...Persistent inflammatory responses often occur when bacteria and other microorganisms frequently invade and colonize open wounds and eventually result in the formation of chronic wounds.Therefore,achieving real-time detection of invasive bacteria accurately and promptly is essential for efficient wound management and accelerat-ing the healing process.Recently,flexible wearable sensors have garnered significant attention,especially those designed for monitoring real-time biophysical or biochemical signals in wound sites in a minimally invasive manner.They provide more precise and continuous monitoring data,making them as emerging tools for clinical diagnostics.In this review,we first discuss the species and community distribution of different types of bacteria in chronic wounds.Next,we introduce currently developed techniques for detecting bacteria at wound sites.Fol-lowing that,we discuss the recent progress and unresolved issues of various flexible wearable sensors in detecting bacteria at wound sites.We believe that this review can provide meaningful guidance for the development of flexible wearable sensors for bacteria detection.展开更多
Background:Human epidermal stem cells(hESCs)play an important role in re-epithelialization and thereby in facilitating wound healing,while an effective way to activate hESCs remains to be explored.Calcium silicate(CS)...Background:Human epidermal stem cells(hESCs)play an important role in re-epithelialization and thereby in facilitating wound healing,while an effective way to activate hESCs remains to be explored.Calcium silicate(CS)is a form of bioceramic that can alter cell behavior and promote tissue regeneration.Here,we have observed the effect of CS on hESCs and investigated its possible mechanism.Methods:Using a mouse full-thickness skin excision model,we explored the therapeutic effect of CS on wound healing and re-epithelialization.In vitro,hESCs were cultured with diluted CS ion extracts(CSIEs),and the proliferation,migration ability and stemness of hESCs were evaluated.The effects of CS on the epidermal growth factor(EGF),epidermal growth factor receptor(EGFR)and extracellular signal-related kinase(ERK)signaling pathway were also explored.Results:In vivo,CS accelerated wound healing and re-epithelialization.Immunohistochemistry demonstrated that CS upregulated cytokeratin 19 and integrinβ1 expression,indicating that CS improved hESCs stemness.In vitro studies confirmed that CS improved the biological function of hESCs.And the possible mechanism could be due to the activation of the EGF/EGFR/ERK signaling pathway.Conclusion:CS can promote re-epithelialization and improve the biological functions of hESCs via activating the EGF/EGFR/ERK signaling pathway.展开更多
Background:Sweat glands(SGs)have low regenerative potential after severe burns or trauma and their regeneration or functional recovery still faces many obstacles.In practice,restoring SG function requires not only the...Background:Sweat glands(SGs)have low regenerative potential after severe burns or trauma and their regeneration or functional recovery still faces many obstacles.In practice,restoring SG function requires not only the structural integrity of the gland itself,but also its neighboring tissues,especially blood vessels.Collagen triple helix repeat containing-1(CTHRC1)was first identified in vascular repair,and increasing reports showed a close correlation between cutaneous appendage specification,patterning and regeneration.The purpose of the present study was to clarify the role of CTHRC1 in SGs and their adjacent microvessels and find therapeutic strategies to restore SG function.Methods:The SGs and their adjacent microvascular network of Cthrc^(1−/−)mice were first inves-tigated using sweat test,laser Doppler imaging,tissue clearing technique and transcriptome analysis.The effects of CTHRC1 on dermal microvascular endothelial cells(DMECs)were further explored with cell proliferation,DiI-labeled acetylated low-density lipoprotein uptake,tube for-mation and intercellular junction establishment assays.The effects of CTHRC1 on SG function restoration were finally confirmed by replenishing the protein into the paws of Cthrc(1−/−)mice.Results:CTHRC1 is a key regulator of SG function in mice.At the tissue level,Cthrc1 deletion resulted in the disorder and reduction of the microvascular network around SGs.At the molecular level,the knockout of Cthrc1 reduced the expression of vascular development genes and functional proteins in the dermal tissues.Furthermore,CTHRC1 administration considerably enhanced SG function by inducing adjacent vascular network reconstruction.Conclusions:CTHRC1 promotes the development,morphogenesis and function execution of SGs and their neighboring vasculature.Our study provides a novel target for the restoration or regeneration of SG function in vivo.展开更多
Background:Persistent hyperglycaemia in diabetes causes functional abnormalities of human dermal fibroblasts(HDFs),partially leading to delayed skin wound healing.Extracellular vesicles(EVs)containing multiple pro-hea...Background:Persistent hyperglycaemia in diabetes causes functional abnormalities of human dermal fibroblasts(HDFs),partially leading to delayed skin wound healing.Extracellular vesicles(EVs)containing multiple pro-healing microRNAs(miRNAs)have been shown to exert therapeutic effects on diabetic wound healing.The present study aimed to observe the effects of EVs derived from placental mesenchymal stem cells(P-MSC-EVs)on diabetic wound healing and high glucose(HG)-induced senescent fibroblasts and to explore the underlying mechanisms.Methods:P-MSC-EVs were isolated by differential ultracentrifugation and locally injected into the full-thickness skin wounds of diabetic mice,to observe the beneficial effects on wound healing in vivo by measuring wound closure rates and histological analysis.Next,a series of assays were conducted to evaluate the effects of low(2.28 x 1010 particles/ml)and high(4.56 x 1010 particles/ml)concentrations of P-MSC-EVs on the senescence,proliferation,migration,and apoptosis of HG-induced senescent HDFs in vitro.Then,miRNA microarrays and real-time quantitative PCR(RT-qPCR)were carried out to detect the differentially expressed miRNAs in HDFs after EVs treatment.Specific RNA inhibitors,miRNA mimics,and small interfering RNA(siRNA)were used to evaluate the role of a candidate miRNA and its target genes in P-MSC-EV-induced improvements in the function of HG-induced senescent HDFs.Results:Local injection of P-MSC-EVs into diabetic wounds accelerated wound closure and reduced scar widths,with better-organized collagen deposition and decreased p16INK4a expression.In vitro,P-MSC-EVs enhanced the antisenescence,proliferation,migration,and antiapoptotic abilities of HG-induced senescent fibroblasts in a dose-dependent manner.MiR-145-5p was found to be highly enriched in P-MSC-EVs.MiR-145-5p inhibitors effectively attenuated the P-MSC-EV-induced functional improvements of senescent fibroblasts.MiR-145-5p mimics simulated the effects of P-MSC-EVs on functional improvements of fibroblasts by suppressing the expression of cyclin-dependent kinase inhibitor 1A and activating the extracellular signal regulated kinase(Erk)/protein kinase B(Akt)signaling pathway.Furthermore,local application of miR-145-5p agomir mimicked the effects of P-MSC-EVs on wound healing.Conclusions:These results suggest that P-MSC-EVs accelerate diabetic wound healing by improv-ing the function of senescent fibroblasts through the transfer of miR-145-5p,which targets cyclin-dependent kinase inhibitor 1A to activate the Erk/Akt signaling pathway.P-MSC-EVs are promising therapeutic candidates for diabetic wound treatment.展开更多
Background:Promoting rapid wound healing with functional recovery of all skin appendages is the main goal of regenerative medicine.So far current methodologies,including the commonly used back excisional wound model(B...Background:Promoting rapid wound healing with functional recovery of all skin appendages is the main goal of regenerative medicine.So far current methodologies,including the commonly used back excisional wound model(BEWM)and paw skin scald wound model,are focused on assessing the regeneration of either hair follicles(HFs)or sweat glands(SwGs).How to achieve de novo appendage regeneration by synchronized evaluation of HFs,SwGs and sebaceous glands(SeGs)is still challenging.Here,we developed a volar skin excisional wound model(VEWM)that is suitable for examining cutaneous wound healing with multiple-appendage restoration,as well as innervation,providing a new research paradigm for the perfect regeneration of skin wounds.Methods:Macroscopic observation,iodine-starch test,morphological staining and qRT-PCR anal-ysis were used to detect the existence of HFs,SwGs,SeGs and distribution of nerve fibres in the volar skin.Wound healing process monitoring,HE/Masson staining,fractal analysis and behavioral response assessment were performed to verify that VEWM could mimic the pathological process and outcomes of human scar formation and sensory function impairment.Results:HFs are limited to the inter-footpads.SwGs are densely distributed in the footpads,scattered in the IFPs.The volar skin is richly innervated.The wound area of the VEWM at 1,3,7 and 10 days after the operation is respectively 89.17%±2.52%,71.72%±3.79%,55.09%±4.94%and 35.74%±4.05%,and the final scar area accounts for 47.80%±6.22%of the initial wound.While the wound area of BEWM at 1,3,7 and 10 days after the operation are respectively 61.94%±5.34%,51.26%±4.89%,12.63%±2.86%and 6.14%±2.84%,and the final scar area accounts for 4.33%±2.67%of the initial wound.Fractal analysis of the post-traumatic repair site for VEWM vs human was performed:lacunarity values,0.040±0.012 vs 0.038±0.014;fractal dimen-sion values,1.870±0.237 vs 1.903±0.163.Sensory nerve function of normal skin vs post-traumatic repair site was assessed:mechanical threshold,1.05±0.52 vs 4.90 g±0.80;response rate to pin-prick,100%vs 71.67%±19.92%,and temperature threshold,50.34◦C±3.11◦C vs 52.13◦C±3.54◦C.Conclusions:VEWM closely reflects the pathological features of human wound healing and can be applied for skin multiple-appendages regeneration and innervation evaluation.展开更多
The regeneration of sweat glands(SwGs)plays a pivotal role in the functional recovery of extensive skin wounds.Recent research has illuminated the possibility of reprogramming human epidermal ker-atinocytes(HEKs)into ...The regeneration of sweat glands(SwGs)plays a pivotal role in the functional recovery of extensive skin wounds.Recent research has illuminated the possibility of reprogramming human epidermal ker-atinocytes(HEKs)into induced SwG cells through the ectopic expression of ectodysplasin A.However,the clinical application of this genetic manipulation approach is inherently limited.In this study,we pre-sent findings demonstrating that a combination of six compounds can effectively and speedily reprogram HEKs in culture into fully functional SwG cells.These chemically induced SwG-like cells(ciSGCs)closely resemble the morphology,phenotypes,and functional properties of human primary SwG ductal cells.Furthermore,ciSGCs can be stimulated to differentiate into mature SwG cell types in vitro.In a 3D culture system,they can also generate SwG organoids that exhibit structural and biological features akin to native SwGs.Upon transplantation into scalded mouse paw skin,ciSGCs significantly expedited cuta-neous wound healing and completely restored the structural and functional aspects of the SwGs.In con-clusion,the small molecule cocktail-directed SwG reprogramming offers a non-transgenic and controllable strategy for producing high-quality,clinical-grade SwG cells,showing immense potential for the treatment of burn patients.展开更多
Loss of sweat glands(SwGs)commonly associated with extensive skin defects is a leading cause of hyperthermia and heat stroke.In vivo tissue engineering possesses the potential to take use of the body natural ability t...Loss of sweat glands(SwGs)commonly associated with extensive skin defects is a leading cause of hyperthermia and heat stroke.In vivo tissue engineering possesses the potential to take use of the body natural ability to regenerate SwGs,making it more conducive to clinical translation.Despite recent advances in regenerative medicine,reconstructing SwG tissue with the same structure and function as native tissue remains challenging.Elucidating the SwG generation mechanism and developing biomaterials for in vivo tissue engineering is essential for understanding and developing in vivo SwG regenerative strategies.Here,we outline the cell biology associated with functional wound healing and the characteristics of bioactive materials.We critically summarize the recent progress in bioactive material-based cell modulation approaches for in vivo SwG regeneration,including the recruitment of endogenous cells to the skin lesion for SwG regeneration and in vivo cellular reprogramming for SwG regeneration.We discussed the re-establishment of microenvironment via bioactive material-mediated regulators.Besides,we offer promising perspectives for directing in situ SwG regeneration via bioactive material-based cell-free strategy,which is a simple and effective approach to regenerate SwG tissue with both fidelity of structure and function.Finally,we discuss the opportunities and challenges of in vivo SwG regeneration in detail.The molecular mechanisms and cell fate modulation of in vivo SwG regeneration will provide further insights into the regeneration of patient-specific SwGs and the development of potential intervention strategies for gland-derived diseases.展开更多
The hair follicle(HF)is a highly conserved sensory organ associated with the immune response against pathogens,thermoregulation,sebum production,angiogenesis,neurogenesis and wound healing.Although recent advances in ...The hair follicle(HF)is a highly conserved sensory organ associated with the immune response against pathogens,thermoregulation,sebum production,angiogenesis,neurogenesis and wound healing.Although recent advances in lineage-tracing techniques and the ability to profile gene expression in small populations of cells have increased the understanding of how stem cells operate during hair growth and regeneration,the construction of functional follicles with cycling activity is still a great challenge for the hair research field and for translational and clinical applications.Given that hair formation and cycling rely on tightly coordinated epithelial-mesenchymal interactions,we thus review potential cell sources with HF-inducive capacities and summarize current bioengineering strategies for HF regeneration with functional restoration.展开更多
Background:Hydrogels with tuneable mechanical properties are an attractive material platform for 3D bioprinting.Thus far,numerous studies have confirmed that the biophysical cues of hydrogels,such as stiffness,are kno...Background:Hydrogels with tuneable mechanical properties are an attractive material platform for 3D bioprinting.Thus far,numerous studies have confirmed that the biophysical cues of hydrogels,such as stiffness,are known to have a profound impact on mesenchymal stem cell(MSC)differentiation;however,their differentiation potential within 3D-bioprinted hydrogels is not completely understood.Here,we propose a protocol for the exploration of how the stiffness of alginate–gelatin(Alg-Gel)composite hydrogels(the widely used bioink)affects the differentiation of MSCs in the presence or absence of differentiation inducing factors.Methods:Two types of Alg-Gel composite hydrogels(Young’s modulus:50 kPa vs.225 kPa)were bioprinted independently of porosity.Then,stiffness-induced biases towards adipogenic and osteogenic differentiation of the embedded MSCs were analysed by co-staining with alkaline phosphatase(ALP)and oil red O.The expression of specific markers at the gene level was detected after a 3-day culture.Results:Confocal microscopy indicated that all tested hydrogels supported MSC growth and viability during the culture period.Higher expression of adipogenic and osteogenic markers(ALP and lipoprotein lipase(LPL))in stiffer 3D-bioprinted matrices demonstrated a more significant response of MSCs to stiffer hydrogels with respect to differentiation,which was more robust in differentiation-inducing medium.However,the LPL expression in stiffer 3D-bioprinted constructs was reduced at day 3 regardless of the presence of differentiation-inducing factors.Although MSCs embedded in softer hydrogels to some extent proceeded toward adipogenic and osteogenic lineages within a few days,their differentiation seemed to be slower and more limited.Interestingly,the hydrogel itself(without differentiation-inducing factors)exhibited a slight effect on whether MSCs differentiated towards an adipogenic or an osteogenic fate.Considering that the mechanoregulated protein Yes-associated protein(YAP)is involved in MSC fate decisions,we further found that inhibition of YAP significantly downregulated the expression of ALP and LPL in MSCs in stiffer constructs regardless of the induced growth factors present.Conclusions:These results demonstrate that the differentiation of MSCs in 3D-bioprinted matrices is dependent on hydrogel stiffness,which emphasizes the importance of biophysical cues as a determinant of cellular behaviour.展开更多
Background:Sweat glands(SGs)and hair follicles(HFs)are two important cutaneous appendages that play crucial roles in homeostatic maintenance and thermoregulation,and their interaction is involved in wound healing.SGs ...Background:Sweat glands(SGs)and hair follicles(HFs)are two important cutaneous appendages that play crucial roles in homeostatic maintenance and thermoregulation,and their interaction is involved in wound healing.SGs can be regenerated from mesenchymal stem cell-laden 3D bioprinted scaffolds,based on our previous studies,whereas regeneration of HFs could not be achieved in the same model.Due to the lack of an in vitro model,the underlying molecular mechanism of the interaction between SGs and HFs in regeneration could not be fully understood.The purpose of the present study was to establish an in vitro model of skin constructs with SGs and HFs and explore the interaction between these two appendages in regeneration.Methods:To investigate the interaction effects between SGs and HFs during their regeneration processes,a combined model was created by seeding HF spheroids on 3D printed SG scaffolds.The interaction between SG scaffolds and HF spheroids was detected using RNA expression and immunofluorescence staining.The effects of microenvironmental cues on SG and HF regeneration were analysed by altering seed cell types and plantar dermis homogenate in the scaffold.Results:According to this model,we overcame the difficulties in simultaneously inducing SG and HF regeneration and explored the interaction effects between SG scaffolds and HF spheroids.Surprisingly,HF spheroids promoted both SG and HF differentiation in SG scaffolds,while SG scaffolds promoted SG differentiation but had little effect on HF potency in HF spheroids.Specifically,microenvironmental factors(plantar dermis homogenate)in SG scaffolds effectively promoted SG and HF genesis in HF spheroids,no matter what the seed cell type in SG scaffolds was,and the promotion effects were persistent.Conclusions:Our approach elucidated a new model for SG and HF formation in vitro and provided an applicable platform to investigate the interaction between SGs and HFs in vitro.This platform might facilitate 3D skin constructs with multiple appendages and unveil the spatiotemporal molecular program of multiple appendage regeneration.展开更多
Unhealable diabetic wounds need to be addressed with the help of newer,more efficacious strategies.Exosomes combined with biomaterials for sustained delivery of therapeutic agents are expected to bring new hope for ch...Unhealable diabetic wounds need to be addressed with the help of newer,more efficacious strategies.Exosomes combined with biomaterials for sustained delivery of therapeutic agents are expected to bring new hope for chronic wound treatment.Here,the engineered exosomes modified for efficiently loading miR146a and attaching to silk fibroin patch(SFP)were demonstrated to promote diabetic wound healing.Silk fibroin binding peptide(SFBP)was screened through phage display,and SFBP-Gluc-MS2(SGM)and pac-miR146a-pac fusion protein were constructed.The designed exosomes(SGM-Exos,miR146a-Exos,and SGM-miR146a-Exos)were isolated from the engineered placental mesenchymal stem cells(PMSCs)transduced with SGM or/and pac-miR146a-pac protein.Gluc signals indicated SGM-Exo@SFP markedly increased the binding rate and the stability of SGM-Exo.Moreover,the loading efficiency of miR146a in SGM-miR146a-Exos was ten-fold higher than that in miR146a-Exos.Superior to untreated,SGM-miR146a-Exo-only treated,and SFP-only treated groups,SGM-miR146a-Exo@SFP drived wound healing associated with less inflammation,collagen deposition,and neovascularization.The transcriptomics analysis suggested anti-inflammatory and regenerative effects with SGM-miR146a-Exo@SFP treatment.Here,we show efficient exosome@biomaterial-based miRNA delivery systems for regenerative medicine and tissue engineering.展开更多
Neutrophil extracellular traps(NETs)have been considered a significant unfavorable factor for wound healing in diabetes,but the mechanisms remain unclear.The therapeutic application of small extracellular vesicles(sEV...Neutrophil extracellular traps(NETs)have been considered a significant unfavorable factor for wound healing in diabetes,but the mechanisms remain unclear.The therapeutic application of small extracellular vesicles(sEVs)derived from mesenchymal stem cells(MSCs)has received considerable attention for their properties.Hypoxic preconditioning is reported to enhance the therapeutic potential of MSC-derived sEVs in regenerative medicine.Therefore,the aim of this study is to illustrate the detailed mechanism of NETs in impairment of diabetic wound healing and develop a promising NET-targeting treatment based on hypoxic pretreated MSC-derived sEVs(Hypo-sEVs).Excessive NETs were found in diabetic wounds and in high glucose(HG)-induced neutrophils.Further research showed that high concentration of NETs impaired the function of fibroblasts through activating endoplasmic reticulum(ER)stress.Hypo-sEVs efficiently promoted diabetic wound healing and reduced the excessive NET formation by transferring miR-17-5p.Bioinformatic analysis and RNA interference experiment revealed that miR-17-5p in Hypo-sEVs obstructed the NET formation by targeting TLR4/ROS/MAPK pathway.Additionally,miR-17-5p overexpression decreased NET formation and overcame NET-induced impairment in fibroblasts,similar to the effects of Hypo-sEVs.Overall,we identify a previously unrecognized NET-related mechanism in diabetic wounds and provide a promising NET-targeting strategy for wound treatment.展开更多
基金supported in part by the National Nature Science Foundation of China(92268206,81830064)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)+4 种基金the Military Medical Research Projects(145AKJ260015000X,2022-JCJQ-ZB-09600)the Military Key Basic Research of Foundational Strengthening Program(2020-JCJQ-ZD-256-021)the Science Foundation of National Defense Science and Technology for Excellent Young(2022-JCJQ-ZQ-017)the Military Medical Research and Development Projects(AWS17J005,2019-126)the Specific Research Fund of The Innovation Platform for Academicians of Hainan Province(YSPTZX202317).
文摘Scar formation resulting from burns or severe trauma can significantly compromise the structural integrity of skin and lead to permanent loss of skin appendages,ultimately impairing its normal physiological function.Accumulating evidence underscores the potential of targeted modulation of mechanical cues to enhance skin regeneration,promoting scarless repair by influencing the extracellular microenvironment and driving the phenotypic transitions.The field of skin repair and skin appendage regeneration has witnessed remarkable advancements in the utilization of biomaterials with distinct physical properties.However,a comprehensive understanding of the underlying mechanisms remains somewhat elusive,limiting the broader application of these innovations.In this review,we present two promising biomaterial-based mechanical approaches aimed at bolstering the regenerative capacity of compromised skin.The first approach involves leveraging biomaterials with specific biophysical properties to create an optimal scarless environment that supports cellular activities essential for regeneration.The second approach centers on harnessing mechanical forces exerted by biomaterials to enhance cellular plasticity,facilitating efficient cellular reprogramming and,consequently,promoting the regeneration of skin appendages.In summary,the manipulation of mechanical cues using biomaterial-based strategies holds significant promise as a supplementary approach for achieving scarless wound healing,coupled with the restoration of multiple skin appendage functions.
基金supported in part by the National Nature Science Foundation of China (81830064, 81721092)the National Key Research and Development Plan (2017YFC1103304)+1 种基金the CAMS Innovation Fund for Medical Sciences (CIFMS, 2019-I2M-5-059)the Military Medical Research and Development Projects (AWS17J005, 2019–126)。
文摘Background: Motor neuron degeneration or loss in the spinal cord is the characteristic phenotype of motor neuron diseases or spinal cord injuries. Being proliferative and located near neurons, astrocytes are considered ideal cell sources for regenerating neurons.Methods: We selected and tested different combinations of the small molecules for inducing the conversion of human and mouse astrocytes into neurons. Microscopic imaging and immunocytochemistry analyses were used to characterize the morphology and phenotype of the induced neurons while RT-q PCR was utilized to analyze changes in gene expression. In addition, whole-cell patch-clamp recordings were measured to examine the electrophysiological properties of induced neurons.Results: The results showed that human astrocytes could be rapidly and efficiently converted into motor neuronlike cells by treatment with defined small molecules, with a yield of over 85% motor neuron-like cells attained. The induced motor neuron-like cells expressed the pan-neuronal markers TUJ1, MAP2, Neu N, and Synapsin 1 and motor neuron markers HB9, ISL1, CHAT, and VACh T. During the conversion process, the cells did not pass through a proliferative neural progenitor cell intermediate. The induced motor neurons were functional, showing the electrophysiological properties of neurons. The same chemical cocktail could induce spinal cord astrocytes from an amyotrophic lateral sclerosis mouse model carrying a SOD1 mutation to become motor neuron-like cells that exhibited a decrease in cell survival and an increase in oxidative stress compared to that observed in wild-type MNs derived from healthy mice. Moreover, the chemical induction reduced oxidative stress in the mutant astrocytes.Conclusions: The results of the present study demonstrated the feasibility of chemically converting human and mouse astrocytes into motor neuron-like cells that are useful for neurodegenerative disease modeling and regenerative medicine.
基金supported in part by the National Nature Science Foundation of China(81830064,81721092)the National Key Research and Development Plan(2017YFC1103304)+1 种基金the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019–126)。
文摘Background:Generation of neurons is essential in cell replacement therapy for neurodegenerative disorders like Parkinson’s disease.Several studies have reported the generation of dopaminergic(DA)neurons from mouse and human fibroblasts by ectopic expression of transcription factors,in which genetic manipulation is associated with potential risks.Methods:The small molecules and protein factors were selected based on their function to directly induce human fetal lung IMR-90 fibroblasts into DA neuron-like cells.Microscopical,immunocytochemical,and RT-qPCR analyses were used to characterize the morphology,phenotype,and gene expression features of the induced cells.The wholecell patch-clamp recordings were exploited to measure the electrophysiological properties.Results:Human IMR-90 fibroblasts were rapidly converted into DA neuron-like cells after the chemical induction using small molecules and protein factors,with a yield of approximately 95%positive TUJ1-positive cells.The induced DA neuron-like cells were immunopositive for pan-neuronal markers MAP2,NEUN,and Synapsin 1 and DA markers TH,DDC,DAT,and NURR1.The chemical induction process did not involve a neural progenitor/stem cell intermediate stage.The induced neurons could fire single action potentials,which reflected partially the electrophysiological properties of neurons.Conclusions:We developed a chemical cocktail of small molecules and protein factors to convert human fibroblasts into DA neuron-like cells without passing through a neural progenitor/stem cell intermediate stage.The induced DA neuron-like cells from human fibroblasts might provide a cellular source for cell-based therapy of Parkinson’s disease in the future.
基金supported in part by the National Natural Science Foundation of China(81830064,81721092,81971841)the National Key Research and Development Plan(2017YFC1103304,2017YFC1104701)。
文摘To determine the prevalence and clinical features of olfactory and taste disorders among coronavirus disease 2019(COVID-19)patients in China.A cross-sectional study was performed in Wuhan from April 3,2020 to April 15,2020.A total of 187 patients with confirmed severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)completed face-to-face interviews or telephone follow-ups.We found that the prevalence of olfactory and taste disorders was significantly lower in the Chinese cohort than in foreign COVID-19 cohorts.Females were more prone to olfactory and taste disorders.In some patients,olfactory and taste disorders precede other symptoms and can be used as early screening and warning signs.
基金supported in part by the National Natural Science Foundation of China(92268206,81830064)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)+2 种基金the Military Medical Research Projects(145AKJ260015000X,2022-JCJQ-ZB-09600,2020-JCJQ-ZD-256-021)the Military Medical Research and Development Projects(AWS17J005,2019-126)the Specific Research Fund of the Innovation Platform for Academicians of Hainan Province(YSPTZX202317).
文摘Dear Editor,The large defects of the skin caused by burns or trauma can result in disruption of the structure and function of the skin,as well as irreversible loss of skin appendages,such as sweat glands(SG).As the important modulator of temperature homeostasis,SG damage triggers heat intolerance and thermoregulatory dysfunction,which poses a considerable threat to health and quality of life[1-3].The promotion of wound healing with SG restoration remains a challenging issue.It has been reported that there are urinary epithelial cells(UECs)in human urine samples,and they may most likely originate from renal proximal tubules[4].UECs are easy to obtain from urine samples at any age,sex,or ethnic origin except for renal failure,with a cost-effective,non-invasive and simple isolation method[4].More importantly,the extensive source of clinical urine sample makes it easy for urinary cell autogenous transplantation and individualized treatment.Therefore,it is of great significance to convert UECs into urinary epithelial stem cells(UESCs)with repair ability.In this context,we design a strategy for the pharmacological conversion of UECs into UESCs,and the repair ability of UESCs in functional skin wound healing is explored.
基金supported in part by the National Natural Science Foundation of China (81871569, 81830064, 81721092, 61803250)the National Key Research and Development Plan (2018YFC1105704, 2017YFC1103304, 2016YFA0101000, 2016YFA0101002)+2 种基金the CAMS Innovation Fund for Medical Sciences (CIFMS, 2019-I2M-5-059)the Military Key Basic Research of Foundational Strengthening Program (2020-JCJQ-ZD-256-021)the Military Medical Research and Development Projects (AWS17J005, 2019-126)。
文摘Background: Large skin defects severely disrupt the overall skin structure and can irreversibly damage sweat glands(SGs), thus impairing the skin’s physiological function. This study aims to develop a stepwise reprogramming strategy to convert fibroblasts into SG lineages, which may provide a promising method to obtain desirable cell types for the functional repair and regeneration of damaged skin.Methods: The expression of the SG markers cytokeratin 5(CK5), cytokeratin 10(CK10), cytokeratin 18(CK18), carcinoembryonic antigen(CEA), aquaporin 5(AQP5) and α-smooth muscle actin(α-SMA) was assessed with quantitative PCR(qPCR), immunofluorescence and flow cytometry. Calcium activity analysis was conducted to test the function of induced SG-like cells(iSGCs). Mouse xenograft models were also used to evaluate the in vivo regeneration of iSGCs.BALB/c nude mice were randomly divided into normal group, SGM treatment group and iSGC transplantation group.Immunocytochemical analyses and starch-iodine sweat tests were used to confirm the in vivo regeneration of iSGCs.Results: Ectodermal dysplasia antigen(EDA) overexpression drove human dermal fibroblast(HDF) conversion into i SGCs in SG culture medium(SGM). qPCR indicated significantly increased mRNA levels of the SG markers CK5, CK18and CEA in iSGCs, and flow cytometry data demonstrated(4.18±0.04)% of iSGCs were CK5 positive and(4.36±0.25)%of iSGCs were CK18 positive. The addition of chemical cocktails greatly accelerated the SG fate program. qPCR results revealed significantly increased mRNA expression of CK5, CK18 and CEA in iSGCs, as well as activation of the duct marker CK10 and luminal functional marker AQP5. Flow cytometry indicated, after the treatment of chemical cocktails,(23.05±2.49)% of iSGCs expressed CK5^(+) and(55.79±3.18)% of iSGCs expressed CK18^(+), respectively. Calcium activity analysis indicated that the reactivity of iSGCs to acetylcholine was close to that of primary SG cells [(60.79±7.71)% vs.(70.59±0.34)%, ns]. In vivo transplantation experiments showed approximately(5.2±1.1)% of the mice were sweat test positive, and the histological analysis results indicated that regenerated SG structures were present in iSGCs-treated mice.Conclusions: We developed a SG reprogramming strategy to generate functional iSGCs from HDFs by using the single factor EDA in combination with SGM and small molecules. The generation of iSGCs has important implications for future in situ skin regeneration with SG restoration.
基金supported in part by the National Nature Science Foundation of China(92268206)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)+3 种基金the Military Medical Research Projects (2023-JSKY-SSQG-006)the National Key Research andDevelopment Program of China (2023YFC3011900)the Beijing NaturalScience Foundation (7232158)the Youth Autonomous InnovationScience Foundation of PLA general hospital (22QNFC019).
文摘Endogenous regeneration is becoming an increasingly important strategy for wound healing as it facilitates skin’s own regenerative potential for self-healing,thereby avoiding the risks of immune rejection and exogenous infection.However,currently applied biomaterials for inducing endogenous skin regeneration are simplistic in their structure and function,lacking the ability to accurately mimic the intricate tissue structure and regulate the disordered microenvironment.Novel biomimetic biomaterials with precise structure,chemical composition,and biophysical properties offer a promising avenue for achieving perfect endogenous skin regeneration.Here,we outline the recent advances in biomimetic materials induced endogenous skin regeneration from the aspects of structural and functional mimicry,physiological process regulation,and biophysical property design.Furthermore,novel techniques including in situ reprograming,flexible electronic skin,artificial intelligence,single-cell sequencing,and spatial transcriptomics,which have potential to contribute to the development of biomimetic biomaterials are highlighted.Finally,the prospects and challenges of further research and application of biomimetic biomaterials are discussed.This review provides reference to address the clinical problems of rapid and high-quality skin regeneration.
基金supported by the National Natural Science Foundation of China(82172211,92268206)the National Key Research and Development Programs of China(2022YFA1104300)+5 种基金the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research Projects(2022-JCJQ-ZB-09600,2023-JSKY-SSQG-006)the Natural Science Foundation of Sichuan Province(2023NSFSC0339)the 1·3·5 Project for Disciplines of Excellence of West China Hospital,Sichuan University(ZYGD22008)the China Postdoctoral Science Foundation(2022TQ0223,2022M722256)the Post-Doctor Research Project of West China Hospital,Sichuan University(2023HXBH031).
文摘Electrospun nanofibrous membranes(eNFMs)have been extensively developed for bio-applications due to their structural and compositional similarity to the natural extracellular matrix.However,the emergence of antibiotic resistance in bacterial infections significantly impedes the further development and applications of eNFMs.The development of antibacterial nanomaterials substantially nourishes the engineering design of antibacterial eNFMs for combating bacterial infections without relying on antibiotics.Herein,a comprehensive review of diverse fabrication techniques for incorporating antibacterial nanomaterials into eNFMs is presented,encompassing an exhaustive introduction to various nanomaterials and their bactericidal mechanisms.Furthermore,the latest achievements and breakthroughs in the application of these antibacterial eNFMs in tissue regenerative therapy,mainly focusing on skin,bone,periodontal and tendon tissues regeneration and repair,are systematically summarized and discussed.In particular,for the treatment of skin infection wounds,we highlight the antibiotic-free antibacterial therapy strategies of antibacterial eNFMs,including(i)single model therapies such as metal ion therapy,chemodynamic therapy,photothermal therapy,and photodynamic therapy;and(ii)multimodel therapies involving arbitrary combinations of these single models.Additionally,the limitations,challenges and future opportunities of antibacterial eNFMs in biomedical applications are also discussed.We anticipate that this comprehensive review will provide novel insights for the design and utilization of antibacterial eNFMs in future research.
基金supported by the National Natural Science Foundation of China(22205260,82172211,92268206)National Key Research and Development Programs of China(2022YFA1104303)+1 种基金the CAMS Innova-tion Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research Projects(145AKJ260015000X,2022-JCJQ-ZB-09600,2023-JSKY-SSQG-006).
文摘Persistent inflammatory responses often occur when bacteria and other microorganisms frequently invade and colonize open wounds and eventually result in the formation of chronic wounds.Therefore,achieving real-time detection of invasive bacteria accurately and promptly is essential for efficient wound management and accelerat-ing the healing process.Recently,flexible wearable sensors have garnered significant attention,especially those designed for monitoring real-time biophysical or biochemical signals in wound sites in a minimally invasive manner.They provide more precise and continuous monitoring data,making them as emerging tools for clinical diagnostics.In this review,we first discuss the species and community distribution of different types of bacteria in chronic wounds.Next,we introduce currently developed techniques for detecting bacteria at wound sites.Fol-lowing that,we discuss the recent progress and unresolved issues of various flexible wearable sensors in detecting bacteria at wound sites.We believe that this review can provide meaningful guidance for the development of flexible wearable sensors for bacteria detection.
基金supported in part by the National Nature Science Foundation of China(81830064,81721092)the National Key ResearchDevelopment Plan(2017YFC1103304)+2 种基金the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019-126)the Beijing Natural Science Foundation(7204309,7202197).
文摘Background:Human epidermal stem cells(hESCs)play an important role in re-epithelialization and thereby in facilitating wound healing,while an effective way to activate hESCs remains to be explored.Calcium silicate(CS)is a form of bioceramic that can alter cell behavior and promote tissue regeneration.Here,we have observed the effect of CS on hESCs and investigated its possible mechanism.Methods:Using a mouse full-thickness skin excision model,we explored the therapeutic effect of CS on wound healing and re-epithelialization.In vitro,hESCs were cultured with diluted CS ion extracts(CSIEs),and the proliferation,migration ability and stemness of hESCs were evaluated.The effects of CS on the epidermal growth factor(EGF),epidermal growth factor receptor(EGFR)and extracellular signal-related kinase(ERK)signaling pathway were also explored.Results:In vivo,CS accelerated wound healing and re-epithelialization.Immunohistochemistry demonstrated that CS upregulated cytokeratin 19 and integrinβ1 expression,indicating that CS improved hESCs stemness.In vitro studies confirmed that CS improved the biological function of hESCs.And the possible mechanism could be due to the activation of the EGF/EGFR/ERK signaling pathway.Conclusion:CS can promote re-epithelialization and improve the biological functions of hESCs via activating the EGF/EGFR/ERK signaling pathway.
基金supported by grants from the National Natural Science Foundation of China(81830064,81721092,32000969,82002056)Key Support Program for Growth Factor Research(SZYZ-TR-03)+3 种基金Chinese PLA General Hospital for Military Medical Innovation Research Project(CX-19026)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005)Beijing Natural Science Foundation(7204306).
文摘Background:Sweat glands(SGs)have low regenerative potential after severe burns or trauma and their regeneration or functional recovery still faces many obstacles.In practice,restoring SG function requires not only the structural integrity of the gland itself,but also its neighboring tissues,especially blood vessels.Collagen triple helix repeat containing-1(CTHRC1)was first identified in vascular repair,and increasing reports showed a close correlation between cutaneous appendage specification,patterning and regeneration.The purpose of the present study was to clarify the role of CTHRC1 in SGs and their adjacent microvessels and find therapeutic strategies to restore SG function.Methods:The SGs and their adjacent microvascular network of Cthrc^(1−/−)mice were first inves-tigated using sweat test,laser Doppler imaging,tissue clearing technique and transcriptome analysis.The effects of CTHRC1 on dermal microvascular endothelial cells(DMECs)were further explored with cell proliferation,DiI-labeled acetylated low-density lipoprotein uptake,tube for-mation and intercellular junction establishment assays.The effects of CTHRC1 on SG function restoration were finally confirmed by replenishing the protein into the paws of Cthrc(1−/−)mice.Results:CTHRC1 is a key regulator of SG function in mice.At the tissue level,Cthrc1 deletion resulted in the disorder and reduction of the microvascular network around SGs.At the molecular level,the knockout of Cthrc1 reduced the expression of vascular development genes and functional proteins in the dermal tissues.Furthermore,CTHRC1 administration considerably enhanced SG function by inducing adjacent vascular network reconstruction.Conclusions:CTHRC1 promotes the development,morphogenesis and function execution of SGs and their neighboring vasculature.Our study provides a novel target for the restoration or regeneration of SG function in vivo.
基金supported by the National Nature Science Foundation of China(82172211,81830064,82172231,81901971)National Key Research and Development Programs of China(2022YFA1104303)+2 种基金the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019-126)the Military Medical Science and Technology Youth Training Program(21QNPY128).
文摘Background:Persistent hyperglycaemia in diabetes causes functional abnormalities of human dermal fibroblasts(HDFs),partially leading to delayed skin wound healing.Extracellular vesicles(EVs)containing multiple pro-healing microRNAs(miRNAs)have been shown to exert therapeutic effects on diabetic wound healing.The present study aimed to observe the effects of EVs derived from placental mesenchymal stem cells(P-MSC-EVs)on diabetic wound healing and high glucose(HG)-induced senescent fibroblasts and to explore the underlying mechanisms.Methods:P-MSC-EVs were isolated by differential ultracentrifugation and locally injected into the full-thickness skin wounds of diabetic mice,to observe the beneficial effects on wound healing in vivo by measuring wound closure rates and histological analysis.Next,a series of assays were conducted to evaluate the effects of low(2.28 x 1010 particles/ml)and high(4.56 x 1010 particles/ml)concentrations of P-MSC-EVs on the senescence,proliferation,migration,and apoptosis of HG-induced senescent HDFs in vitro.Then,miRNA microarrays and real-time quantitative PCR(RT-qPCR)were carried out to detect the differentially expressed miRNAs in HDFs after EVs treatment.Specific RNA inhibitors,miRNA mimics,and small interfering RNA(siRNA)were used to evaluate the role of a candidate miRNA and its target genes in P-MSC-EV-induced improvements in the function of HG-induced senescent HDFs.Results:Local injection of P-MSC-EVs into diabetic wounds accelerated wound closure and reduced scar widths,with better-organized collagen deposition and decreased p16INK4a expression.In vitro,P-MSC-EVs enhanced the antisenescence,proliferation,migration,and antiapoptotic abilities of HG-induced senescent fibroblasts in a dose-dependent manner.MiR-145-5p was found to be highly enriched in P-MSC-EVs.MiR-145-5p inhibitors effectively attenuated the P-MSC-EV-induced functional improvements of senescent fibroblasts.MiR-145-5p mimics simulated the effects of P-MSC-EVs on functional improvements of fibroblasts by suppressing the expression of cyclin-dependent kinase inhibitor 1A and activating the extracellular signal regulated kinase(Erk)/protein kinase B(Akt)signaling pathway.Furthermore,local application of miR-145-5p agomir mimicked the effects of P-MSC-EVs on wound healing.Conclusions:These results suggest that P-MSC-EVs accelerate diabetic wound healing by improv-ing the function of senescent fibroblasts through the transfer of miR-145-5p,which targets cyclin-dependent kinase inhibitor 1A to activate the Erk/Akt signaling pathway.P-MSC-EVs are promising therapeutic candidates for diabetic wound treatment.
基金supported in part by the National Nature Science Foundation of China[92268206,81830064]the CAMS Innovation Fund for Medical Sciences[CIFMS,2019-I2M-5-059]+2 种基金the Military Medical Research Projects[145AKJ260015000X,2022-JCJQ-ZB-09600,2020-JCJQZD-256-021]the Military Medical Research and Development Projects[AWS17J005,2019-126]the Specific Research Fund of The Innovation Platform for Academicians of Hainan Province[YSPTZX202317].
文摘Background:Promoting rapid wound healing with functional recovery of all skin appendages is the main goal of regenerative medicine.So far current methodologies,including the commonly used back excisional wound model(BEWM)and paw skin scald wound model,are focused on assessing the regeneration of either hair follicles(HFs)or sweat glands(SwGs).How to achieve de novo appendage regeneration by synchronized evaluation of HFs,SwGs and sebaceous glands(SeGs)is still challenging.Here,we developed a volar skin excisional wound model(VEWM)that is suitable for examining cutaneous wound healing with multiple-appendage restoration,as well as innervation,providing a new research paradigm for the perfect regeneration of skin wounds.Methods:Macroscopic observation,iodine-starch test,morphological staining and qRT-PCR anal-ysis were used to detect the existence of HFs,SwGs,SeGs and distribution of nerve fibres in the volar skin.Wound healing process monitoring,HE/Masson staining,fractal analysis and behavioral response assessment were performed to verify that VEWM could mimic the pathological process and outcomes of human scar formation and sensory function impairment.Results:HFs are limited to the inter-footpads.SwGs are densely distributed in the footpads,scattered in the IFPs.The volar skin is richly innervated.The wound area of the VEWM at 1,3,7 and 10 days after the operation is respectively 89.17%±2.52%,71.72%±3.79%,55.09%±4.94%and 35.74%±4.05%,and the final scar area accounts for 47.80%±6.22%of the initial wound.While the wound area of BEWM at 1,3,7 and 10 days after the operation are respectively 61.94%±5.34%,51.26%±4.89%,12.63%±2.86%and 6.14%±2.84%,and the final scar area accounts for 4.33%±2.67%of the initial wound.Fractal analysis of the post-traumatic repair site for VEWM vs human was performed:lacunarity values,0.040±0.012 vs 0.038±0.014;fractal dimen-sion values,1.870±0.237 vs 1.903±0.163.Sensory nerve function of normal skin vs post-traumatic repair site was assessed:mechanical threshold,1.05±0.52 vs 4.90 g±0.80;response rate to pin-prick,100%vs 71.67%±19.92%,and temperature threshold,50.34◦C±3.11◦C vs 52.13◦C±3.54◦C.Conclusions:VEWM closely reflects the pathological features of human wound healing and can be applied for skin multiple-appendages regeneration and innervation evaluation.
基金supported by the National Nature Science Foundation of China(92268206)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)+3 种基金the Military Medical Research Projects(2023-JSKY-SSQG-006)the Independent Research Project of State Key Laboratory of Trauma and Chemical Poisoning,the National Key Research and Development Program of China(2023YFC3011900)the Specific Research Fund of The Innovation Platform for Academicians of Hainan Province(YSPTZX202317)the Open Project of the State Key Laboratory of Trauma,Burn and Combined Injury,Third Military Medical University(SKLKF202301).
文摘The regeneration of sweat glands(SwGs)plays a pivotal role in the functional recovery of extensive skin wounds.Recent research has illuminated the possibility of reprogramming human epidermal ker-atinocytes(HEKs)into induced SwG cells through the ectopic expression of ectodysplasin A.However,the clinical application of this genetic manipulation approach is inherently limited.In this study,we pre-sent findings demonstrating that a combination of six compounds can effectively and speedily reprogram HEKs in culture into fully functional SwG cells.These chemically induced SwG-like cells(ciSGCs)closely resemble the morphology,phenotypes,and functional properties of human primary SwG ductal cells.Furthermore,ciSGCs can be stimulated to differentiate into mature SwG cell types in vitro.In a 3D culture system,they can also generate SwG organoids that exhibit structural and biological features akin to native SwGs.Upon transplantation into scalded mouse paw skin,ciSGCs significantly expedited cuta-neous wound healing and completely restored the structural and functional aspects of the SwGs.In con-clusion,the small molecule cocktail-directed SwG reprogramming offers a non-transgenic and controllable strategy for producing high-quality,clinical-grade SwG cells,showing immense potential for the treatment of burn patients.
基金supported in part by the National Nature Science Foundation of China[92268206,81830064]the CAMS Innovation Fund for Medical Sciences[CIFMS,2019-I2M-5-059]+2 种基金the Military Medical Research Projects[145AKJ260015000X,2022-JCJQ-ZB-09600,2020-JCJQ-ZD-256-021]the Military Medical Research and Development Projects[AWS17J005,2019-126]the Specific Research Fund of The Innovation Platform for Academicians of Hainan Province[YSPTZX202317].
文摘Loss of sweat glands(SwGs)commonly associated with extensive skin defects is a leading cause of hyperthermia and heat stroke.In vivo tissue engineering possesses the potential to take use of the body natural ability to regenerate SwGs,making it more conducive to clinical translation.Despite recent advances in regenerative medicine,reconstructing SwG tissue with the same structure and function as native tissue remains challenging.Elucidating the SwG generation mechanism and developing biomaterials for in vivo tissue engineering is essential for understanding and developing in vivo SwG regenerative strategies.Here,we outline the cell biology associated with functional wound healing and the characteristics of bioactive materials.We critically summarize the recent progress in bioactive material-based cell modulation approaches for in vivo SwG regeneration,including the recruitment of endogenous cells to the skin lesion for SwG regeneration and in vivo cellular reprogramming for SwG regeneration.We discussed the re-establishment of microenvironment via bioactive material-mediated regulators.Besides,we offer promising perspectives for directing in situ SwG regeneration via bioactive material-based cell-free strategy,which is a simple and effective approach to regenerate SwG tissue with both fidelity of structure and function.Finally,we discuss the opportunities and challenges of in vivo SwG regeneration in detail.The molecular mechanisms and cell fate modulation of in vivo SwG regeneration will provide further insights into the regeneration of patient-specific SwGs and the development of potential intervention strategies for gland-derived diseases.
基金supported,in part,by National Key Research and Development Plan(2018YFC1105704,2017YFC1103304,2016YFA0101000,2016YFA0101002)the National Nature Science Foundation of China(81871569,81830064,81721092)+1 种基金the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019-126).
文摘The hair follicle(HF)is a highly conserved sensory organ associated with the immune response against pathogens,thermoregulation,sebum production,angiogenesis,neurogenesis and wound healing.Although recent advances in lineage-tracing techniques and the ability to profile gene expression in small populations of cells have increased the understanding of how stem cells operate during hair growth and regeneration,the construction of functional follicles with cycling activity is still a great challenge for the hair research field and for translational and clinical applications.Given that hair formation and cycling rely on tightly coordinated epithelial-mesenchymal interactions,we thus review potential cell sources with HF-inducive capacities and summarize current bioengineering strategies for HF regeneration with functional restoration.
基金This study was supported in part by the National Nature Science Foundation of China(81830064,81721092,81701906)the National Key Research and Development Plan(2017YFC1103300)+3 种基金Funds Chinese PLA General Hospital for Military Medical Innovation Research Project(CX19026)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019–126)Fostering Funds of Chinese PLA General Hospital for National Distinguished Young Scholar Science Fund(2017-JQPY-002).
文摘Background:Hydrogels with tuneable mechanical properties are an attractive material platform for 3D bioprinting.Thus far,numerous studies have confirmed that the biophysical cues of hydrogels,such as stiffness,are known to have a profound impact on mesenchymal stem cell(MSC)differentiation;however,their differentiation potential within 3D-bioprinted hydrogels is not completely understood.Here,we propose a protocol for the exploration of how the stiffness of alginate–gelatin(Alg-Gel)composite hydrogels(the widely used bioink)affects the differentiation of MSCs in the presence or absence of differentiation inducing factors.Methods:Two types of Alg-Gel composite hydrogels(Young’s modulus:50 kPa vs.225 kPa)were bioprinted independently of porosity.Then,stiffness-induced biases towards adipogenic and osteogenic differentiation of the embedded MSCs were analysed by co-staining with alkaline phosphatase(ALP)and oil red O.The expression of specific markers at the gene level was detected after a 3-day culture.Results:Confocal microscopy indicated that all tested hydrogels supported MSC growth and viability during the culture period.Higher expression of adipogenic and osteogenic markers(ALP and lipoprotein lipase(LPL))in stiffer 3D-bioprinted matrices demonstrated a more significant response of MSCs to stiffer hydrogels with respect to differentiation,which was more robust in differentiation-inducing medium.However,the LPL expression in stiffer 3D-bioprinted constructs was reduced at day 3 regardless of the presence of differentiation-inducing factors.Although MSCs embedded in softer hydrogels to some extent proceeded toward adipogenic and osteogenic lineages within a few days,their differentiation seemed to be slower and more limited.Interestingly,the hydrogel itself(without differentiation-inducing factors)exhibited a slight effect on whether MSCs differentiated towards an adipogenic or an osteogenic fate.Considering that the mechanoregulated protein Yes-associated protein(YAP)is involved in MSC fate decisions,we further found that inhibition of YAP significantly downregulated the expression of ALP and LPL in MSCs in stiffer constructs regardless of the induced growth factors present.Conclusions:These results demonstrate that the differentiation of MSCs in 3D-bioprinted matrices is dependent on hydrogel stiffness,which emphasizes the importance of biophysical cues as a determinant of cellular behaviour.
基金supported partially by the National Nature Science Foundation of China(81830064,81721092,81701906)the National Key Research and Development Plan(2017YFC1103300)+3 种基金Funds of Chinese PLA General Hospital for Military Medical Inno-vation Research Project(CX19026)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019-126)Fostering Funds of Chinese PLA General Hospital for National Distinguished Young Scholar Science Fund(2017-JQPY-002).
文摘Background:Sweat glands(SGs)and hair follicles(HFs)are two important cutaneous appendages that play crucial roles in homeostatic maintenance and thermoregulation,and their interaction is involved in wound healing.SGs can be regenerated from mesenchymal stem cell-laden 3D bioprinted scaffolds,based on our previous studies,whereas regeneration of HFs could not be achieved in the same model.Due to the lack of an in vitro model,the underlying molecular mechanism of the interaction between SGs and HFs in regeneration could not be fully understood.The purpose of the present study was to establish an in vitro model of skin constructs with SGs and HFs and explore the interaction between these two appendages in regeneration.Methods:To investigate the interaction effects between SGs and HFs during their regeneration processes,a combined model was created by seeding HF spheroids on 3D printed SG scaffolds.The interaction between SG scaffolds and HF spheroids was detected using RNA expression and immunofluorescence staining.The effects of microenvironmental cues on SG and HF regeneration were analysed by altering seed cell types and plantar dermis homogenate in the scaffold.Results:According to this model,we overcame the difficulties in simultaneously inducing SG and HF regeneration and explored the interaction effects between SG scaffolds and HF spheroids.Surprisingly,HF spheroids promoted both SG and HF differentiation in SG scaffolds,while SG scaffolds promoted SG differentiation but had little effect on HF potency in HF spheroids.Specifically,microenvironmental factors(plantar dermis homogenate)in SG scaffolds effectively promoted SG and HF genesis in HF spheroids,no matter what the seed cell type in SG scaffolds was,and the promotion effects were persistent.Conclusions:Our approach elucidated a new model for SG and HF formation in vitro and provided an applicable platform to investigate the interaction between SGs and HFs in vitro.This platform might facilitate 3D skin constructs with multiple appendages and unveil the spatiotemporal molecular program of multiple appendage regeneration.
基金This work was supported by the National Nature Science Foundation of China(81901971,82172211,81830064,82172231)Natural Science Foundation of Beijing Municipal(7194316,7202197)+3 种基金National Key Research and Development Programs of China(2022YFA1104303)the CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)the Military Medical Research and Development Projects(AWS17J005,2019-126)Military Medical Science and Technology Youth Training Program(21QNPY128).
文摘Unhealable diabetic wounds need to be addressed with the help of newer,more efficacious strategies.Exosomes combined with biomaterials for sustained delivery of therapeutic agents are expected to bring new hope for chronic wound treatment.Here,the engineered exosomes modified for efficiently loading miR146a and attaching to silk fibroin patch(SFP)were demonstrated to promote diabetic wound healing.Silk fibroin binding peptide(SFBP)was screened through phage display,and SFBP-Gluc-MS2(SGM)and pac-miR146a-pac fusion protein were constructed.The designed exosomes(SGM-Exos,miR146a-Exos,and SGM-miR146a-Exos)were isolated from the engineered placental mesenchymal stem cells(PMSCs)transduced with SGM or/and pac-miR146a-pac protein.Gluc signals indicated SGM-Exo@SFP markedly increased the binding rate and the stability of SGM-Exo.Moreover,the loading efficiency of miR146a in SGM-miR146a-Exos was ten-fold higher than that in miR146a-Exos.Superior to untreated,SGM-miR146a-Exo-only treated,and SFP-only treated groups,SGM-miR146a-Exo@SFP drived wound healing associated with less inflammation,collagen deposition,and neovascularization.The transcriptomics analysis suggested anti-inflammatory and regenerative effects with SGM-miR146a-Exo@SFP treatment.Here,we show efficient exosome@biomaterial-based miRNA delivery systems for regenerative medicine and tissue engineering.
基金supported by National Natural Science Foundation of China(82172211,92268206,22205260,81830064,82172231)National Key Research and Development Programs of China(2022YFA1104303)+2 种基金CAMS Innovation Fund for Medical Sciences(CIFMS,2019-I2M-5-059)Military Medical Research and Development Projects(AWS17J005,2019-126)Military Medical Science and Technology Youth Training Program(21QNPY128).
文摘Neutrophil extracellular traps(NETs)have been considered a significant unfavorable factor for wound healing in diabetes,but the mechanisms remain unclear.The therapeutic application of small extracellular vesicles(sEVs)derived from mesenchymal stem cells(MSCs)has received considerable attention for their properties.Hypoxic preconditioning is reported to enhance the therapeutic potential of MSC-derived sEVs in regenerative medicine.Therefore,the aim of this study is to illustrate the detailed mechanism of NETs in impairment of diabetic wound healing and develop a promising NET-targeting treatment based on hypoxic pretreated MSC-derived sEVs(Hypo-sEVs).Excessive NETs were found in diabetic wounds and in high glucose(HG)-induced neutrophils.Further research showed that high concentration of NETs impaired the function of fibroblasts through activating endoplasmic reticulum(ER)stress.Hypo-sEVs efficiently promoted diabetic wound healing and reduced the excessive NET formation by transferring miR-17-5p.Bioinformatic analysis and RNA interference experiment revealed that miR-17-5p in Hypo-sEVs obstructed the NET formation by targeting TLR4/ROS/MAPK pathway.Additionally,miR-17-5p overexpression decreased NET formation and overcame NET-induced impairment in fibroblasts,similar to the effects of Hypo-sEVs.Overall,we identify a previously unrecognized NET-related mechanism in diabetic wounds and provide a promising NET-targeting strategy for wound treatment.
