Material-driven immunomodulation and ECM remodeling reverse pulmonary fibrosis by local delivery of stem cell-laden microcapsules.

Recent progress in stem cell therapy has demonstrated the therapeutic potential of intravenous stem cell infusions for treating the life-threatening lung disease of pulmonary fibrosis (PF). However, it is confronted with limitations, such as a lack of control over cellular function and rapid clearance by the host after implantation. In this study, we developed an innovative PF therapy through tracheal administration of microfluidic-templated stem cell-laden microcapsules, which effectively reversed the progression of inflammation and fibrotic injury. Our findings highlight that hydrogel microencapsulation can enhance the persistence of donor mesenchymal stem cells (MSCs) in the host while driving MSCs to substantially augment their therapeutic functions, including immunoregulation and matrix metalloproteinase (MMP)-mediated extracellular matrix (ECM) remodeling. We revealed that microencapsulation activates the MAPK signaling pathway in MSCs to increase MMP expression, thereby degrading overexpressed collagen accumulated in fibrotic lungs. Our research demonstrates the potential of hydrogel microcapsules to enhance the therapeutic efficacy of MSCs through cell-material interactions, presenting a promising yet straightforward strategy for designing advanced stem cell therapies for fibrotic diseases.

Cell volume regulation modulates macrophage-related inflammatory responses via JAK/STAT signaling pathways.

Cell volume as a characteristic of changes in response to external environmental cues has been shown to control the fate of stem cells. However, its influence on macrophage behavior and macrophage-mediated inflammatory responses have rarely been explored. Herein, through mediating the volume of macrophages by adding polyethylene glycol (PEG), we demonstrated the feasibility of fine-tuning cell volume to regulate macrophage polarization towards anti-inflammatory phenotypes, thereby enabling to reverse macrophage-mediated inflammation response. Specifically, lower the volume of primary macrophages can induce both resting macrophages (M0) and stimulated pro-inflammatory macrophages (M1) to up-regulate the expression of anti-inflammatory factors and down-regulate pro-inflammatory factors. Further mechanistic investigation revealed that macrophage polarization resulting from changing cell volume might be mediated by JAK/STAT signaling pathway evidenced by the transcription sequencing analysis. We further propose to apply this strategy for the treatment of arthritis via direct introduction of PEG into the joint cavity to modulate synovial macrophage-related inflammation. Our preliminary results verified the credibility and effectiveness of this treatment evidenced by the significant inhibition of cartilage destruction and synovitis at early stage. In general, our results suggest that cell volume can be a biophysical regulatory factor to control macrophage polarization and potentially medicate inflammatory response, thereby providing a potential facile and effective therapy for modulating macrophage mediated inflammatory responses. STATEMENT OF SIGNIFICANCE: Cell volume has recently been recognized as a significantly important biophysical signal in regulating cellular functionalities and even steering cell fate. Herein, through mediating the volume of macrophages by adding polyethylene glycol (PEG), we demonstrated the feasibility of fine-tuning cell volume to induce M1 pro-inflammatory macrophages to polarize towards anti-inflammatory M2 phenotype, and this immunomodulatory effect may be mediated by the JAK/STAT signaling pathway. We also proposed the feasible applications of this PEG-induced volume regulation approach towards the treatment of osteoarthritis (OA), wherein our preliminary results implied an effective alleviation of early synovitis. Our study on macrophage polarization mediated by cell volume may open up new pathways for immune regulation through microenvironmental biophysical clues.

Local delivery of stem cell spheroids with protein/polyphenol self-assembling armor to improve myocardial infarction treatment via immunoprotection and immunoregulation.

Stem cell therapies have shown great potential for treating myocardial infarction (MI) but are limited by low cell survival and compromised functionality due to the harsh microenvironment at the disease site. Here, we presented a Mesenchymal stem cell (MSC) spheroid-based strategy for MI treatment by introducing a protein/polyphenol self-assembling armor coating on the surface of cell spheroids, which showed significantly enhanced therapeutic efficacy by actively manipulating the hostile pathological MI microenvironment and enabling versatile functionality, including protecting the donor cells from host immune clearance, remodeling the ROS microenvironment and stimulating MSC's pro-healing paracrine secretion. The underlying mechanism was elucidated, wherein the armor protected to prolong MSCs residence at MI site, and triggered paracrine stimulation of MSCs towards immunoregulation and angiogenesis through inducing hypoxia to provoke glycolysis in stem cells. Furthermore, local delivery of coated MSC spheroids in MI rat significantly alleviated local inflammation and subsequent fibrosis via mediation macrophage polarization towards pro-healing M2 phenotype and improved cardiac function. In general, this study provided critical insight into the enhanced therapeutic efficacy of stem cell spheroids coated with a multifunctional armor. It potentially opens up a new avenue for designing immunomodulatory treatment for MI via stem cell therapy empowered by functional biomaterials.

Hyaluronic acid-based multifunctional carriers for applications in regenerative medicine: A review.

Hyaluronic acid (HA) is an important type of naturally derived carbohydrate polymer with specific polysaccharide macromolecular structures and multifaceted biological functions, including biocompatibility, low immunogenicity, biodegradability, and bioactivity. Specifically, HA hydrogels in a microscopic scale have been widely used for biomedical applications, such as drug delivery, tissue engineering, and medical cosmetology, considering their superior properties outperforming the more conventional monolithic hydrogels in network homogeneity, degradation profile, permeability, and injectability. Herein, we reviewed the recent progress in the preparation and applications of HA microgels in biomedical fields. We first summarized the fabrication of HA microgels by focusing on the different crosslinking/polymerization schemes for HA gelation and the miniaturized fabrication techniques for producing HA-based microparticles. We then highlighted the use of HA-based microgels for different applications in regenerative medicine, including cartilage repair, bioactive delivery, diagnostic imaging, modular tissue engineering. Finally, we discussed the challenges and future perspectives in bridging the translational gap in the utilization of HA-based microgels in regenerative medicine.

Microfluidic-templated cell-laden microgels fabricated using phototriggered imine-crosslinking as injectable and adaptable granular gels for bone regeneration.

Injectable granular gels consisting of densely packed microgels serving as scaffolding biomaterial have recently shown great potential for applications in tissue regeneration, which allow administration via minimally invasive surgery, on-target cargo delivery, and high efficiency in nutrient/waste exchange. However, limitations such as insufficient mechanical strength, structural integrity, and uncontrollable differentiation of the encapsulated cells in the scaffolds hamper their further applications in the biomedical field. Herein, we developed a new class of granular gels via bottom-up assembly of cell-laden microgels via photo-triggered imine-crosslinking (PIC) chemistry based on the microfluidic technique. The particulate nature of the granular gels rendered them with shear-thinning and self-healing behavior, thereby functioning as an injectable and adaptable cellularized scaffold for bone tissue regeneration. Specifically, single cell-laden, monodisperse microgels composed of methacrylate- and o-nitrobenzene-functionalized hyaluronic acid and gelatin were prepared using a high-throughput microfluidic technique with a production rate up to 3.7 × 108 microgels/hr, wherein the PIC chemistry alleviated the oxygen inhibition on free-radical polymerization and facilitated enhanced fabrication accuracy, accelerated gelation rate, and improved network strength. Further in vitro and in vivo studies demonstrated that the microgels can serve as carriers to support the activity of the encapsulated mesenchymal stem cells; these cell-laden microgels can also be used as cellularized bone fillers to induce the regeneration of bone tissues as evidenced by the in vivo experiment using the rat femoral condyle defect model. In general, these results represent a significant step toward the precise fabrication of engineered tissue mimics with single-cell resolution and high cell-density and can potentially offer a powerful tool for the design and applications of a next generation of tissue engineering strategy. STATEMENT OF SIGNIFICANCE: Using microfluidic droplet-based technology, we hereby developed a new class of injectable and moldable granular gels via bottom-up assembly of cell-laden microgels as a versatile platform for tissue regeneration. Phototriggered imine-crosslinking chemistry was introduced for microgel cross-linkage, which allowed for the fabrication of microgels with improved matrix homogeneity, accelerated gelation process, and enhanced mechanical strength. We demonstrated that the microgel building blocks within the granular gels facilitated the proliferation and differentiation of the encapsulated mesenchymal stem cells, which can further serve as a cellularized scaffold for the treatment of bone defects.

Large-scale single-cell encapsulation in microgels through metastable droplet-templating combined with microfluidic-integration.

Current techniques for the generation of cell-laden microgels are limited by numerous challenges, including poorly uncontrolled batch-to-batch variations, processes that are both labor- and time-consuming, the high expense of devices and reagents, and low production rates; this hampers the translation of laboratory findings to clinical applications. To address these challenges, we develop a droplet-based microfluidic strategy based on metastable droplet-templating and microchannel integration for the substantial large-scale production of single cell-laden alginate microgels. Specifically, we present a continuous processing method for microgel generation by introducing amphiphilic perfluoronated alcohols to obtain metastable emulsion droplets as sacrificial templates. In addition, to adapt to the metastable emulsion system, integrated microfluidic chips containing 80 drop-maker units are designed and optimized based on the computational fluid dynamics simulation. This strategy allows single cell encapsulation in microgels at a maximum production rate of 10 ml h-1of cell suspension while retaining cell viability and functionality. These results represent a significant advance toward using cell-laden microgels for clinical-relevant applications, including cell therapy, tissue regeneration and 3D bioprinting.

Efficacy and Safety of Anti-Nerve Growth Factor Antibody Therapy for Hip and Knee Osteoarthritis: A Meta-analysis.

Background: The efficacy and safety of anti-nerve growth factor (NGF) antibody therapy used for osteoarthritis (OA) pain are controversial. Purpose: To evaluate the efficacy and safety of anti-NGF antibody therapy via a meta-analysis of randomized controlled trials (RCTs). Study Design: Systematic review; Level of evidence, 1. Methods: PubMed, the Cochrane Central Register of Controlled Trials, Embase, and the Web of Science databases were searched for RCTs assessing anti-NGF antibody treatments for hip and knee OA. A total of 623 records were retrieved from the databases. A random-effects model was used to assess primary and secondary outcomes. Bias was assessed using the Cochrane Collaboration tool, funnel plots, and the Egger test. Subgroup analyses were used to assess the efficacy and safety of the independent variables. Sensitivity analysis was conducted to evaluate the effectiveness of tanezumab and the effectiveness of anti-NGF antibodies compared to active comparator drugs. We present the effects of dose, administration mode, and treatment duration on the efficacy and safety of anti-NGF antibody therapy. Results: There were 19 RCTs included in our meta-analysis. Anti-NGF antibody treatment showed significant improvements on the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) for pain, physical function, and stiffness as well as on a patient global assessment (PGA). The overall standardized mean differences were as follows: WOMAC pain (-0.31 [95% CI, -0.36 to -0.26]; Z = 11.75; P < .001; I 2 = 38%), WOMAC physical function (-0.36 [95% CI, -0.41 to -0.30]; Z = 12.67; P < .001; I 2 = 44%), WOMAC stiffness (-3.59 [95% CI, -4.87 to -2.30]; Z = 5.47; P < .001; I 2 = 98%), and PGA (-0.28 [95% CI, -0.34 to -0.22]; Z = 9.39; P < .001; I 2 = 50%). Anti-NGF antibody treatment resulted in a greater incidence of adverse events (risk ratio, 1.09 [95% CI, 1.06 to 1.12]; Z = 5.60; P < .001; I 2 = 0%). The incidence of serious adverse events was similar between the treatment and control groups (risk ratio, 1.15 [95% CI, 0.98 to 1.34]; Z = 1.71; P = .09; I 2 = 0%). Conclusion: Anti-NGF antibody treatment significantly relieved pain and improved function in patients with hip and knee OA. However, no conclusion could be drawn regarding the optimal treatment plan for anti-NGF antibodies when all 3 variables (dose, administration mode, and treatment duration) were combined in the analyses.

Microfluidic-templating alginate microgels crosslinked by different metal ions as engineered microenvironment to regulate stem cell behavior for osteogenesis.

Cell microenvironment is a collection of dynamic biochemical and biophysical cues which functions as the key factor in determining cell behavior. Encapsulating single cell into micrometer-scale hydrogels which mimics the cell microenvironment can be used for single cell analysis, cell therapies, and tissue engineering. Here, we developed a microfluidics-based platform to engineer the niche environment at single cell level using alginate microgels crosslinked by different metal ions to regulate stem cell behavior for bone regeneration. Specifically, we revealed that Ca2+ in the engineered microenvironment promoted osteogenic differentiation of encapsulated stem cells and substantially accelerated the matrix mineralization compared to Sr2+in vitro. However, the superior osteoinductive capacity of Ca2+ compared with Sr2+ led to comparable bone healing in a rat bone defect model. This attributed to Sr2+ in microgels to inhibit the osteoclast activity and bone resorption after implantation. In summary, the present study demonstrates metal ions as a critical factor in the environmental cues to affect cell behavior and influence the efficacy of stem cell-based therapy in tissue regeneration, and provides new insights to engineer an expecting microenvironment for regenerative medicine.

Continuous microfluidic encapsulation of single mesenchymal stem cells using alginate microgels as injectable fillers for bone regeneration.

The encapsulation of cells in microscale hydrogels can provide a mimic of a three-dimensional (3D) microenvironment to support cell viability and functions and to protect cells from the environmental stress, which have been widely used in tissue regeneration and cell therapies. Here, a microfluidics-based approach is developed for continuous encapsulation of mesenchymal stem cells (MSCs) at the single-cell level using alginate microgels. This microfluidic technique integrated on-chip encapsulation, gelation, and de-emulsification into a one-step fabrication process, which enables scalable cell encapsulation while retaining the viability and functionality of loaded cells. Remarkably, we observed MSCs encapsulated in Ca-alginate microgels at the single-cell level showed significantly enhanced osteogenesis and accelerated mineralization of the microgels which occurred only after 7 days of induction. Furthermore, MSCs laden in alginate microgels displayed significantly enhanced bone formation compared to MSCs mixed with microgels and acellular microgels in a rat tibial ablation model. To conclude, the current microfluidic technique represents a significant step toward continuous single cell encapsulation, fabrication, and purification. These microgels can boost bone regeneration by providing a controlled osteogenic microenvironment for encapsulated MSCs and facilitate stem cell therapy in the treatment of bone defects in a minimally invasive delivery way. STATEMENT OF SIGNIFICANCE: The biological functions and therapeutic activities of single cells laden in microgels for tissue engineering remains less investigated. Here, we reported a microfluidic-based method for continuous encapsulation of single MSCs with high viability and functionality by integrating on-chip encapsulation, gelation, and de-emulsification into a one-step fabrication process. More importantly, MSCs encapsulated in alginate microgels at the single-cell level showed significantly enhanced osteogenesis, remarkably accelerated mineralization in vitro and bone formation capacity in vivo. Therefore, this single-cell encapsulation technique can facilitate stem cell therapy for bone regeneration and be potentially used in a variety of tissue engineering applications.

Negative regulators of STAT3 signaling pathway in cancers.

STAT3 is the most ubiquitous member of the STAT family and involved in many biological processes, such as cell proliferation, differentiation, and apoptosis. Mounting evidence has revealed that STAT3 is aberrantly activated in many malignant tumors and plays a critical role in cancer progression. STAT3 is usually regarded as an effective molecular target for cancer treatment, and abolishing the STAT3 activity may diminish tumor growth and metastasis. Recent studies have shown that negative regulators of STAT3 signaling such as PIAS, SOCS, and PTP, can effectively retard tumor progression. However, PIAS, SOCS, and PTP have also been reported to correlate with tumor malignancy, and their biological function in tumorigenesis and antitumor therapy are somewhat controversial. In this review, we summarize actual knowledge on the negative regulators of STAT3 in tumors, and focus on the potential role of PIAS, SOCS, and PTP in cancer treatment. Furthermore, we also outline the STAT3 inhibitors that have entered clinical trials. Targeting STAT3 seems to be a promising strategy in cancer therapy.

Association of HLA-DPB1 polymorphisms with rheumatoid arthritis: A systemic review and meta-analysis.

BACKGROUND: The current reports on the association between HLA-DPB1 alleles and rheumatoid arthritis (RA) results were controversial. Thus, we conducted a meta-analysis to assess whether DPB1 alleles are associated with increased risk of rheumatoid arthritis. METHODS: Systematic searches on PubMed, Embase, Elsevier, CNKI (China National Knowledge Infrastructure), Wanfang data and Cochrane Library prior to July 2017 were performed. The pooled odds ratios (ORs) and 95% confidence interval (95% CI) was used to assess the association between frequencies of DPB1 alleles and RA patients. RESULTS: Eight studies with 592 cases and 935 controls were included in this meta-analysis. Overall, the pooled ORs showed that frequencies of DPB1*0401 and *0601 were higher in the RA group compared with controls (*0401: OR: 1.586, 95%CI: 1.296-1.941, P<0.001; *0601: OR: 1.921, 95%CI: 1.142-3.229, P=0.014). Whereas, the frequencies of DPB1*0101, *0402 and *0501 were lower in the RA control than the controls (*0101: OR: 0.691, 95%CI: 0.481-0.993, P=0.046; *0402: OR: 0.707, 95%CI: 0.555-0.902, P=0.005; *0501: OR: 0.502, 95%CI: 0.329-0.767, P=0.001). No associations were observed for DPB1*0201, *0202, *0301 and *0901 (*0201: OR: 1.129, 95%CI: 0.882-1.446, P = 0.335; *0202: OR: 0.840, 95%CI: 0.940-1.441, P = 0.527; *0301: OR: 0.769, 95%CI: 0.577-1.026, P = 0.074; *0901: OR: 1.221, 95% CI: 0.541-2.755, P = 0.630). CONCLUSIONS: This meta-analysis demonstrates that high frequency expression of DPB1*0401 and *0601 are significantly associated with susceptibility to RA, it may be a risk factor for occurrence of RA. Low frequency expression of DPB1*0101, *0402 and *0501 may be negatively associated with RA, it may be a protective factor for occurrence of RA.

Reversing the polarization of tumor-associated macrophages inhibits tumor metastasis.

OBJECTIVE: The M2 phenotype is dominant in tumor associated macrophages (TAM), and plays a key role in promoting tumor growth, invasion and metastasis. Converting TAM polarization from M2 to M1 may contribute to eliciting anti-tumor-specific immune responses and inhibiting tumor metastasis. In this study, the effect of reversing the polarization of TAM on tumor metastasis was investigated. METHODS: Peritoneal macrophages were obtained from BABL/c mice, and M2 polarization was induced by IL-4. In an in vivo experiment, BABL/c mice were transplanted with 4T1 tumor cells. In vitro and in vivo experimental studies, M2 macrophage polarization was reversed with CpG-DNA or CpG-DNA combined with anti-IL-10R Ab. CD68, MHCII and FRß molecular expression in macrophages were examined with immunofluorescence staining. The mRNA expression of IL-2, IL-6, IL-13, VEGF and MMP-9 were detected with RT-PCR. VEGF and MMP-9 protein expression of tumors in situ was measured by western blot assay. Lung-metastasis of the tumor was observed and assessed by micro-CT. RESULTS: CpG-DNA and CpG-DNA combined with anti-IL-10R Ab could promote MHCII, IL-2, IL-6 and IL-13 molecular expression, and suppress the expression of FRß, MMP-9 and VEGF, in both freshly isolated peritoneal macrophages and M2 macrophages. In the CpG-DNA combined with anti-IL-10R Ab injecting group, the percentage of CD68+ MHCII+ cells were significantly higher than that of CD68+FRß+ cells (P<0.05). This was distinct from the result of the control group, which CD68+ FRß+ was higher than CD68+MHCII+cells (P<0.01). Furthermore, VEGF-A and MMP-9 level in primary tumor tissues in the experimental group was significantly lower (P<0.01), compared to the control group. Moreover, the number of detectable lung-metastasis foci was significantly lower in the experimental group than in the control group (P<0.05). CONCLUSION: Reversing the polarization of TAM from M2 to M1 phenotype can inhibit tumor metastasis.

Differential sensitivities of bladder cancer cell lines to resveratol are unrelated to its metabolic profile.

Resveratrol (RV) is a natural polyphenol compound with a wide range of activities, including inhibition of human bladder cancer (HBC) cell growth. Because RV is rapidly metabolized and has poor bioavailability, it is unclear whether the antitumor activity is due to RV or its metabolites. We therefore used liquid chromatography-mass spectroscopy, qRT-PCR, immunocytochemistry and western blotting to evaluate the metabolic profile and biotransformation of RV in the T24 and EJ HBC cell lines. Both T24 and EJ cells generated the same RV metabolite, RV monosulfate (RVS), and both exhibited upregulation of the RV-associated metabolic enzyme SULT1A1 (sulfotransferase). Despite these similarities, T24 cells were more sensitive to RV than EJ cells, yet T24 cells exhibited no sensitivity to an RVS mixture (84.13% RVS). Primary rat bladder epithelial cells showed no adverse effects when exposed to a therapeutic dose (100 µM) of RV. The differences in RV sensitivity between the two HBC cell lines did not reflect differences in the RV metabolic profile or SULT1A1 expression. Because RV exhibited stronger antitumor activity and better safety than RVS, we conclude that RV has significant therapeutic potential for HBC treatment, provided individual differences are considered during clinical research and application.

Knockdown of ST6Gal-I inhibits the growth and invasion of osteosarcoma MG-63 cells.

The upregulation of alpha-2,6-sialyltranferase 1 (ST6Gal-I) has been observed in several malignant tumors, including colon, ovarian and liver cancers, where its expression correlates with the invasion and metastasis of these tumors. However, the roles and molecular mechanisms by which ST6Gal-I mediates the growth and invasion of osteosarcoma cells still remain poorly unknown. In this study, we investigated the expression of ST6Gal-I in osteosarcoma MG-63 and Saos-2 cells which have different metastatic potential, and found that ST6Gal-I was highly expressed in MG-63 cells compared to Saos-2 cells. Downregulation of ST6Gal-I by shRNA in MG-63 cells significantly inhibited their malignant behaviors, including in cell proliferation and soft agar colony formation, as well as migration and invasion properties. In addition, we found that ST6Gal-I knockdown inhibited the expression levels of N-cadherin, vimentin, a-SMA, MMP-2, MMP-9 and VEGF. Together, our results suggest a role for ST6Gal-I to promote the growth and invasion of osteosarcoma cells through modulation of EMT-related molecules, and might be a promising marker for the prognosis and therapy of osteosarcoma.