MOF-Derived Oxygen-Deficient Titania-Mediated Photodynamic/Photothermal-Enhanced Immunotherapy for Tumor Treatment.

Immunotherapy has emerged as a revolutionizing therapeutic modality for cancer. However, its efficacy has been largely limited by a weak immune response and an immunosuppressive tumor microenvironment. Herein, we report a metal-organic framework (MOF)-derived titanium oxide nanoparticle (MCTx NP) as an immune booster that can greatly improve the immunotherapy efficacy by inducing "immunogenic cell death" (ICD) and remodeling the tumor microenvironment. The NPs, inheriting the characteristic structure of MIL-125 and enriched with oxygen vacancies (OVs), demonstrate both high photothermal conversion efficiency and a reactive oxygen species (ROS) generation yield upon near-infrared (NIR) activation. Moreover, the NPs can release O2 and reduce glutathione (GSH) in the tumor environment, showcasing their potential to reverse the immunosuppressive microenvironment. In vitro/vivo results demonstrate that MCTx NPs directly kill tumor cells and effectively eliminate primary tumors by exerting dual photodynamic/photothermal therapy under a single NIR irritation. At the same time, MCTx NPs augment the PD-L1 blockade efficacy by potently inducing ICDs and reversing the immunosuppressive tumor microenvironment, including promoting dendritic cell (DC) maturation, decreasing regulatory T cells (Tregs)' infiltration, and increasing cytotoxic T lymphocytes (CTLs) and helper T cells (Ths), resulting in effective distant tumor suppression. This work highlights MCTx NP-mediated photodynamic- and photothermal-enhanced immunotherapy as an effective strategy for tumor treatment.

Early changes of bone metabolites and lymphocyte subsets may participate in osteoporosis onset: a preliminary study of a postmenopausal osteoporosis mouse model.

Purpose: Metabolic and immune changes in the early stages of osteoporosis are not well understood. This study aimed to explore the changes in bone metabolites and bone marrow lymphocyte subsets and their relationship during the osteoporosis onset. Methods: We established OVX and Sham mouse models. After 5, 15, and 40 days, five mice in each group were sacrificed. Humeri were analyzed by microCT. The bone marrow cells of the left femur and tibia were collected for flow cytometry analysis. The right femur and tibia were analyzed by LC-MS/MS for metabolomics analysis. Results: Bone microarchitecture was significantly deteriorated 15 days after OVX surgery. Analysis of bone metabolomics showed that obvious metabolite changes had happened since 5 days after surgery. Lipid metabolism was significant at the early stage of the osteoporosis. The proportion of immature B cells was increased, whereas the proportion of mature B cells was decreased in the OVX group. Metabolites were significantly correlated with the proportion of lymphocyte subsets at the early stage of the osteoporosis. Conclusion: Lipid metabolism was significant at the early stage of the osteoporosis. Bone metabolites may influence bone formation by interfering with bone marrow lymphocyte subsets.

Omentin­1 induces osteoblast viability and differentiation via the TGF­ß/Smad signaling pathway in osteoporosis.

Osteoporosis is a bone­related disease that results from impaired bone formation and excessive bone resorption. The potential value of adipokines has been investigated previously, due to their influence on osteogenesis. However, the osteogenic effects induced by omentin­1 remain unclear. The aim of the present study was to determine the regulatory effects of omentin­1 on osteoblast viability and differentiation, as well as to explore the underlying molecular mechanism. The present study investigated the effects of omentin­1 on the viability and differentiation of mouse pre­osteoblast cells (MC3T3­E1) using quantitative and qualitative measures. A Cell Counting Kit­8 assay was used to assess the viability of MC3T3­E1 cells following treatment with different doses of omentin­1. Omentin­1 and bone morphogenetic protein (BMP) inhibitor were added to osteogenic induction mediums in different ways to assess their effect. The alkaline phosphatase (ALP) activity and Alizarin Red S (ARS) staining of MC3T3­E1 cells treated with omentin­1 and/or BMP inhibitor were used to examine the effects of omentin­1 on differentiation and mineralization. Western blotting was used to further explore its potential mechanism, and to study the role of omentin­1 on the viability and differentiation of osteoblasts. The results showed that omentin­1 altered the viability of MC3T3­E1 cells in a dose­dependent manner. Omentin­1 treatment significantly increased the expression of members of the TGF­ß/Smad signaling pathway. In the omentin­1 group, the ALP activity of the MC3T3­E1 cells was increased, and the ARS staining area was also increased. The mRNA and protein expression levels of BMP2, Runt­related transcription factor 2, collagen1, osteopontin, osteocalcin and osterix in the omentin­1 group were also significantly upregulated. All these effects were reversed following treatment with SIS3 HCl. These results demonstrated that omentin­1 can significantly promote osteoblast viability and differentiation via the TGF­ß/Smad signaling pathway, thereby promoting bone formation and preventing osteoporosis.

Tracking mesenchymal stem cells with Ir(III) complex-encapsulated nanospheres in cranium defect with postmenopausal osteoporosis.

Osteoporosis (OP) is a significant public health problem with associated fragility fractures, thereby causing large bone defects and difficulty in self-repair. The introduction of human mesenchymal stem cells (hMSCs) is the most promising platform in bone tissue engineering for OP therapy, which induces less side effects than conventional medication. However, the safety and efficiency of the cell-based OP therapy requires the ability to monitor the cell's outcome and biodistribution after cell transplantation. Therefore, we designed an in vivo system to track hMSCs in real time and simultaneously attempted to obtain a significant therapeutic effect during the bone repair process. In this study, we synthesized Ir(III) complex, followed by encapsulation with biodegradable methoxy-poly(ethylene glycol) poly(lactic-co-glycolic acid) nanospheres through double emulsions strategy. The Ir(III) complex nanospheres did not affect hMSC proliferation, stemness, and differentiation and realized highly efficient and long-term cellular labeling for at least 25 days in vivo. The optimal transplantation conditions were also determined first by injecting a gradient number of labeled hMSCs percutaneously into the cranial defect of the nude mouse model. Next, we applied this method to ovariectomy-induced OP mice. Results showed long-term optical imaging with high fluorescence intensity and computed tomography (CT) scanning with significantly increased bone formation between the osteoporotic and sham-operated bones. During the tracking process, two mice from each group were sacrificed at two representative time points to examine the bony defect bridging via micro-CT morphometric analyses. Our data showed remarkable promise for efficient hMSC tracking and encouraging treatment in bioimaging-guided OP stem cell therapy.

Ionizing radiation induces cutaneous lipid remolding and skin adipocytes confer protection against radiation-induced skin injury.

BACKGROUND: Radiation-induced skin injury is a serious concern during radiotherapy and radiation accidents. Skin fat represents the dominant architectural component of the human skin. However, the interplay between skin fat and the progression of radiation-induced skin injury remains largely unexplored. OBJECTIVE: This study aims to elucidate the interplay between skin fat and the progression of radiation-induced skin injury. METHODS: SD rats were irradiated with an electron beam. mRNA profiles were determined by RNA-Seq. The skin lipid mass was monitored by magnetic resonance imaging (MRI) and lipid profiles were measured by liquid chromatography-mass spectrometry (LC-MS). Human mature adipocytes isolated from dermal and subcutaneous white adipose tissues (WATs) were co-cultured with human keratinocytes (HaCaT) and skin fibroblasts (WS1) in the transwell culture system. Cell migration ability was measured by migration assay. RESULTS: Radiation modulated cutaneous lipid metabolism by downregulating multiple pathways. Moreover, radiation decreased skin fat mass with altered lipid metabolite profiles. The rats fed with a high-fat diet showed resistance to radiogenic skin injury compared with that with a control diet, indicating that skin lipid plays a radioprotective role. Mature adipocytes promoted the migration but not the proliferation of co-cultured skin keratinocytes and fibroblasts. Palmitic acid, the most abundant fatty acid in skin tissues, facilitated the migration of WS1 cells. Moreover, fatty acid-binding protein 4 (FABP4) could be incorporated into skin cells and promote DNA damage repair in irradiated skin fibroblasts. CONCLUSION: Radiation induces cutaneous lipid remolding, and skin adipocytes confer a protective role against radiation-induced skin injury.

Genome-Wide Analysis Reveals Zinc Transporter ZIP9 Regulated by DNA Methylation Promotes Radiation-Induced Skin Fibrosis via the TGF-ß Signaling Pathway.

Radiation-induced skin fibrosis is a detrimental and chronic disorder that occurs after radiation exposure. DNA methylation has been characterized as an important regulatory mechanism of multiple pathological processes. In this study, we compared the genome-wide DNA methylation status in radiation-induced fibrotic skin and adjacent normal tissues of rats by methylated DNA immunoprecipitation sequencing. Radiation-induced fibrotic skin showed differentially methylated regions associated with 3,650 protein-coding genes, 72 microRNAs, 5,836 long noncoding RNAs and 3 piwi-interacting RNAs. By integrating the mRNA and methylation profiles, the zinc transporter SLC39A9/ZIP9 was investigated in greater detail. The protein level of ZIP9 was increased in irradiated skin tissues of humans, monkeys, and rats, especially in radiogenic fibrotic skin tissues. Radiation induced the demethylation of a CpG dinucleotide in exon 1 of ZIP9 that resulted in recruitment of the transcriptional factor Sp1 and increased ZIP9 expression. Overexpression of ZIP9 resulted in activation of the profibrotic transforming growth factor-ß signaling pathway through protein kinase B in human fibroblasts. In addition, radiation-induced skin fibrosis was associated with increased zinc accumulation. The zinc chelator N,N,N',N'-tetrakis(2-pyridylmethyl)-1,2-ethylenediamine abrogated ZIP9-induced activation of the transforming growth factor-ß signaling pathway and attenuated radiation-induced skin fibrosis in a rat model. In summary, our findings illustrate epigenetic regulation of ZIP9 and its critical role in promoting radiation-induced skin fibrosis.

Smart, Elastic, and Nanofiber-Based 3D Scaffolds with Self-Deploying Capability for Osteoporotic Bone Regeneration.

It has been a major challenge to treat osteoporotic bone defects with irregular shapes. Although bioactive glass offers an attractive material for bone regeneration, its inherent brittleness has greatly limited its scope of application. Herein, we report the fabrication of bioactive glass (SiO2-CaO) nanofibers with excellent flexibility to even allow for 180° bending. The bioactive glass nanofibers could be further assembled into 3D fibrous scaffolds with chitosan serving as the linkers. The scaffolds constructed from an assembly of 85SiO2-15CaO nanofibers and chitosan (85SiO2-15CaO NF/CS) possessed significantly better mechanical properties when benchmarked against both 75SiO2-25CaO nanofiber- and chitosan-based scaffolds. Moreover, the 85SiO2-15CaO NF/CS scaffolds exhibited an elastic behavior, with full recovery from 80% compression and good fatigue resistance over 1000 cycles of compression under water. Upon implantation, the elastic fibrous scaffolds were able to deform and fit irregularly shaped bone defects, followed by a self-deploying behavior to achieve a perfect match with the cavities. When applied to the repair of an osteoporotic calvarial defect in a rat model, the 85SiO2-15CaO NF/CS scaffolds showed substantial promotion of bone regrowth and vascularization. This new class of 3D fibrous scaffold provides a promising advancement in engineering smart materials for complex bone repair.

Bone microarchitectural parameters can detect oxytocin induced changes prior to bone density on mitigating bone deterioration in rabbit osteoporosis model using micro-CT.

BACKGROUND: This study is aimed to determine the efficacy of X-Ray Microtomography (micro-CT) in predicting oxytocin (OT) treatment response in rabbit osteoporosis(OP) model. METHODS: Sixty-five rabbits were randomly divided into three groups: control group, ovariectomy (OVX) -vehicle and OVX-oxytocin group. The controls underwent sham surgery. OVX-vehicle and OVX-oxytocin groups were subjected to bilateral OVX. The rabbits in OVX-oxytocin group were injected with oxytocin. In the 0th, 4th, 8th, 10th and 12th weeks post OVX operation, bone mineral density (BMD) and bone micro-architectural parameters were measured in three groups. RESULTS: Bone mineral density (BMD), bone volume fraction (BV/TV), Trabecular Number (Tb.N), and Trabecular Thickness (Tb.Th) decreased, while Trabecular Spacing (Tb.Sp) and Structure Model Index (SMI) increased overtime in all the three groups. In OVX-oxytocin group, the bone deterioration tendency is slowing down compared with that of the OVX-vehicle group. The BMD of the OVX-oxytocin group was significantly lower than those in the OVX-vehicle group at 12th week (P = 0.017). BV/TV and Tb.Sp in OVX-oxytocin group changed significantly from 8th week (P = 0.043) and 12th week (P = 0.014), which is earlier than that of BMD and other bone micro-architectural parameters. CONCLUSION: BV/TV and Tb.Sp changed prior to BMD and other bone micro-architectural parameters with oxytocin intervention, which indicate that they are more sensitive markers for predicting early osteoporosis and treatment monitoring when using micro-CT to evaluate osteoporosis rabbit model.

Comparison of semi-quantitative and quantitative dynamic contrast-enhanced MRI evaluations of vertebral marrow perfusion in a rat osteoporosis model.

BACKGROUND: This study aims to investigate the technical feasibility of semi-quantitative and quantitative dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in the assessment of longitudinal changes of marrow perfusion in a rat osteoporosis model, using bone mineral density (BMD) measured by micro-computed tomography (micro-CT) and histopathology as the gold standards. METHODS: Fifty rats were randomly assigned to the control group (n=25) and ovariectomy (OVX) group whose bilateral ovaries were excised (n=25). Semi-quantitative and quantitative DCE-MRI, micro-CT, and histopathological examinations were performed on lumbar vertebrae at baseline and 3, 6, 9, and 12 weeks after operation. The differences between the two groups in terms of semi-quantitative DCE-MRI parameter (maximum enhancement, Emax), quantitative DCE-MRI parameters (volume transfer constant, Ktrans; interstitial volume, Ve; and efflux rate constant, Kep), micro-CT parameter (BMD), and histopathological parameter (microvessel density, MVD) were compared at each of the time points using an independent-sample t test. The differences in these parameters between baseline and other time points in each group were assessed via Bonferroni's multiple comparison test. A Pearson correlation analysis was applied to assess the relationships between DCE-MRI, micro-CT, and histopathological parameters. RESULTS: In the OVX group, the Emax values decreased significantly compared with those of the control group at weeks 6 and 9 (p=0.003 and 0.004, respectively). The Ktrans values decreased significantly compared with those of the control group from week 3 (p<0.05). However, the Ve values decreased significantly only at week 9 (p=0.032), and no difference in the Kep was found between two groups. The BMD values of the OVX group decreased significantly compared with those of the control group from week 3 (p<0.05). Transmission electron microscopy showed tighter gaps between vascular endothelial cells with swollen mitochondria in the OVX group from week 3. The MVD values of the OVX group decreased significantly compared with those of the control group only at week 12 (p=0.023). A weak positive correlation of Emax and a strong positive correlation of Ktrans with MVD were found. CONCLUSIONS: Compared with semi-quantitative DCE-MRI, the quantitative DCE-MRI parameter Ktrans is a more sensitive and accurate index for detecting early reduced perfusion in osteoporotic bone.

Fluorescent microspheres for one-photon and two-photon imaging of mesenchymal stem cells.

Stem cell-mediated therapy has emerged as a novel regenerative treatment for tissue repair in the last decade. However, noninvasive monitoring of stem cells remains a grand challenge for stem cell-based regenerative medicine for the complete understanding of the function and the behaviors of cells. Herein, acridine orange, a fluorescent dye was encapsulated into polymer nanospheres based on double emulsions. The fluorescent nanospheres with a diameter of around 200 nm were incubated with human mesenchymal stem cells (hMSCs) to produce nanosphere-endocytosed hMSCs. The fluorescent imaging of hMSCs can be directly and clearly captured using confocal microscopy in the one-photon and two-photon modes. The results indicated that the hMSCs adhered and spread well on the surface of the scaffolds with a high population and good distribution. Meanwhile, polystyrene microspheres were stained with dye complexes by using a gradual solvent evaporation method. The as-prepared fluorescent beads exhibited bright and multiple colors and a broad emission spectrum ranging from 500 to 800 nm, which was further used to quantitatively evaluate the one-photon and two-photon fluorescent signals of the hMSCs. Our study offers the possibility of direct monitoring of stem cells with high resolution, and encourages future quantitative clinical assessment in imaging-guided cell therapies.

Reduction of Longitudinal Vertebral Blood Perfusion and Its Likely Causes: A Quantitative Dynamic Contrast-enhanced MR Imaging Study of a Rat Osteoporosis Model.

Purpose To determine the longitudinal relationships among lumbar vertebral blood perfusion, bone mass, and marrow adipose tissue in a rat osteoporosis model after ovariectomy by using quantitative dynamic contrast agent-enhanced (DCE) magnetic resonance (MR) imaging, microcomputed tomography (micro-CT), and proton MR spectroscopy. Materials and Methods In this animal review committee-approved study, lumbar vertebrae were evaluated through MR spectroscopy, quantitative DCE MR imaging, micro-CT, and histopathologic analysis, and blood was examined at 0, 2, 6, 10, 14, 18, and 24 weeks after ovariectomy consisting of exposure of the ovaries but no excision (n = 35) or sham operation, defined as exposure of the ovaries but no excision (n = 35). Differences in the parameters of these examinations between two groups at the same time point were analyzed by an independent-sample t test. Results Significantly reduced volume transfer constant (Ktrans) and volume of extravascular extracellular space per unit volume of tissue (week 2 and 10, respectively; P = .036 and P = .014, respectively), decreased bone mineral density (week 2; P = .014), and increased fat fraction (week 6; P = .036) in the ovariectomy group were observed, compared with those in the sham group. Vascular endothelial growth factor and microvessel density values of the ovariectomy group decreased significantly compared with those of the sham group from weeks 18 (P = .005) and 14 (P = .018), respectively. Transmission electron microscopy revealed tighter gaps among vascular endothelial cells and more marrow fibrosis in the ovariectomy group. Conclusion Quantitative DCE MR imaging can directly reflect marrow perfusion. Ktrans is a promising parameter to demonstrate early reduced marrow perfusion. Enhanced vasoconstriction and tightened gaps among vascular endothelial cells may be the likely causes in the initial stage of osteoporosis. Increased marrow adipose tissue, decreased microvessel density, and increased marrow fibrosis may aggravate marrow ischemia in the late stage of osteoporosis. © RSNA, 2016 Online supplemental material is available for this article.